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Monoclonal Anti-beta-Actin antibody produced in mouse

RRID:AB_476743

Antibody ID

AB_476743

Target Antigen

beta-Actin antibody produced in mouse bovine, rabbit, c elegans/worm, mouse, guinea pig, canine, feline, sheep, porcine, human, zebrafish/fish, rat, canine, guinea pig, rabbit, bovine, chicken, rat, sheep, wide range, carp, drosophila, hirudo medicinalis, cat, human, mouse, pig

Proper Citation

(Sigma-Aldrich Cat# A5316, RRID:AB_476743)

Clonality

monoclonal antibody

Comments

Vendor recommendations: IgG2a immunohistochemistry (formalin-fixed, paraffin-embedded sections): suitable, indirect ELISA: suitable, immunoblotting: 1:5,000

Vendor

Sigma-Aldrich

Cat Num

A5316

Publications that use this research resource

Histone Methylation by SETD1A Protects Nascent DNA through the Nucleosome Chaperone Activity of FANCD2.

  • Higgs MR
  • Mol. Cell
  • 2018 Jul 5

Literature context:


Abstract:

Components of the Fanconi anemia and homologous recombination pathways play a vital role in protecting newly replicated DNA from uncontrolled nucleolytic degradation, safeguarding genome stability. Here we report that histone methylation by the lysine methyltransferase SETD1A is crucial for protecting stalled replication forks from deleterious resection. Depletion of SETD1A sensitizes cells to replication stress and leads to uncontrolled DNA2-dependent resection of damaged replication forks. The ability of SETD1A to prevent degradation of these structures is mediated by its ability to catalyze methylation on Lys4 of histone H3 (H3K4) at replication forks, which enhances FANCD2-dependent histone chaperone activity. Suppressing H3K4 methylation or expression of a chaperone-defective FANCD2 mutant leads to loss of RAD51 nucleofilament stability and severe nucleolytic degradation of replication forks. Our work identifies epigenetic modification and histone mobility as critical regulatory mechanisms in maintaining genome stability by restraining nucleases from irreparably damaging stalled replication forks.

Funding information:
  • NCRR NIH HHS - RR024574(United States)

Human Cord Blood Serum-Derived APP α-Secretase Cleavage Activity is Mediated by C1 Complement.

  • Habib A
  • Cell Transplant
  • 2018 Jun 7

Literature context:


Abstract:

Alzheimer's Disease (AD) is the leading cause of dementia in the elderly. In healthy individuals, amyloid precursor protein (APP) is cleaved by α-secretase, generating soluble α-amyloid precursor protein (sAPPα), which contributes neuroprotective functions in the neuronal environment. In contrast, in the neurodegenerative environment of AD patients, amyloid-β-peptide (Aβ) of either 40 or 42 residues are generated by increased activity of β- and γ-secretase. These proteins amalgamate in specific regions of the brain, which disrupts neuronal functions and leads to cognitive impairment. Human umbilical cord blood cells (HUCBC) have proven useful as potential immunomodulatory therapies in various models of neurodegenerative diseases, including AD. Our most recent work studied the impact of umbilical cord blood serum (CBS) on modulation of sAPPα production. Heat-sensitive CBS significantly promoted sAPPα production, indicating that heat-sensitive factor(s) play(s) a role in this process. Liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis was used to determine the molecular source of α-secretase in purified CBS and aged blood serum (AgBS) fraction. Of the proteins identified, the subunits of C1 complex (C1q, C1r, and C1s) and alpha-2-macroglobulin showed significantly greater levels in purified α-CBS fraction (α-CBSF) compared with the AgBS fraction (AgBSF). Specifically, C1 markedly increased sAPPα and alpha-carboxyl-terminal fragment (α-CTF) production in a dose-dependent fashion, whereas C1q alone only minimally increased and C3 did not increase sAPPα production in the absence of sera. Furthermore, C1q markedly increased sAPPα and α-CTF, while decreasing Aβ, in CHO/APPwt cells cultured in the presence of whole sera. These results confirm our initial assumption that APP α-secretase activity in human blood serum is mediated by complement C1, opening a potential therapeutic modality for the future of AD.

Funding information:
  • Howard Hughes Medical Institute - GM084477(United States)

Dependence on the Pyrimidine Biosynthetic Enzyme DHODH Is a Synthetic Lethal Vulnerability in Mutant KRAS-Driven Cancers.

  • Koundinya M
  • Cell Chem Biol
  • 2018 Jun 21

Literature context:


Abstract:

Activating KRAS mutations are major oncogenic drivers in multiple tumor types. Synthetic lethal screens have previously been used to identify targets critical for the survival of KRAS mutant cells, but their application to drug discovery has proven challenging, possibly due in part to a failure of monolayer cultures to model tumor biology. Here, we report the results of a high-throughput synthetic lethal screen for small molecules that selectively inhibit the growth of KRAS mutant cell lines in soft agar. Chemoproteomic profiling identifies the target of the most KRAS-selective chemical series as dihydroorotate dehydrogenase (DHODH). DHODH inhibition is shown to perturb multiple metabolic pathways. In vivo preclinical studies demonstrate strong antitumor activity upon DHODH inhibition in a pancreatic tumor xenograft model.

Funding information:
  • Medical Research Council - (United Kingdom)

Regulation of Epithelial Plasticity Determines Metastatic Organotropism in Pancreatic Cancer.

  • Reichert M
  • Dev. Cell
  • 2018 Jun 18

Literature context:


Abstract:

The regulation of metastatic organotropism in pancreatic ductal a denocarcinoma (PDAC) remains poorly understood. We demonstrate, using multiple mouse models, that liver and lung metastatic organotropism is dependent upon p120catenin (p120ctn)-mediated epithelial identity. Mono-allelic p120ctn loss accelerates KrasG12D-driven pancreatic cancer formation and liver metastasis. Importantly, one p120ctn allele is sufficient for E-CADHERIN-mediated cell adhesion. By contrast, cells with bi-allelic p120ctn loss demonstrate marked lung organotropism; however, rescue with p120ctn isoform 1A restores liver metastasis. In a p120ctn-independent PDAC model, mosaic loss of E-CADHERIN expression reveals selective pressure for E-CADHERIN-positive liver metastasis and E-CADHERIN-negative lung metastasis. Furthermore, human PDAC and liver metastases support the premise that liver metastases exhibit predominantly epithelial characteristics. RNA-seq demonstrates differential induction of pathways associated with metastasis and epithelial-to-mesenchymal transition in p120ctn-deficient versus p120ctn-wild-type cells. Taken together, P120CTN and E-CADHERIN mediated epithelial plasticity is an addition to the conceptual framework underlying metastatic organotropism in pancreatic cancer.

Funding information:
  • NCI NIH HHS - F30 CA180601()
  • NCI NIH HHS - F32 CA221094()
  • NIDDK NIH HHS - P30 DK050306()
  • NIDDK NIH HHS - R01 DK060694()
  • NIDDK NIH HHS - R21DK090778(United States)

Anti-Depressant Fluoxetine Reveals its Therapeutic Effect Via Astrocytes.

  • Kinoshita M
  • EBioMedicine
  • 2018 Jun 12

Literature context:


Abstract:

Although psychotropic drugs act on neurons and glial cells, how glia respond, and whether glial responses are involved in therapeutic effects are poorly understood. Here, we show that fluoxetine (FLX), an anti-depressant, mediates its anti-depressive effect by increasing the gliotransmission of ATP. FLX increased ATP exocytosis via vesicular nucleotide transporter (VNUT). FLX-induced anti-depressive behavior was decreased in astrocyte-selective VNUT-knockout mice or when VNUT was deleted in mice, but it was increased when astrocyte-selective VNUT was overexpressed in mice. This suggests that VNUT-dependent astrocytic ATP exocytosis has a critical role in the therapeutic effect of FLX. Released ATP and its metabolite adenosine act on P2Y11 and adenosine A2b receptors expressed by astrocytes, causing an increase in brain-derived neurotrophic factor in astrocytes. These findings suggest that in addition to neurons, FLX acts on astrocytes and mediates its therapeutic effects by increasing ATP gliotransmission.

Funding information:
  • NHLBI NIH HHS - R01-HL112605(United States)

Brain phospholipid precursors administered post-injury reduce tissue damage and improve neurological outcome in experimental traumatic brain injury.

  • Thau-Zuchman O
  • J. Neurotrauma
  • 2018 May 17

Literature context:


Abstract:

Traumatic brain injury (TBI) leads to cellular loss, destabilisation of membranes, disruption of synapses and altered brain connectivity, and increased risk of neurodegenerative disease. A significant and long-lasting decrease in phospholipids (PL), essential membrane constituents, has recently been reported in plasma and brain tissue, in human and experimental TBI. We hypothesised that supporting PL synthesis post-injury could improve outcome after TBI. We tested this hypothesis using a multi-nutrient combination designed to support the biosynthesis of phospholipids and available for clinical use. The multi-nutrient Fortasyn® Connect (FC) contains polyunsaturated omega-3 fatty acids, choline, uridine, vitamins, co-factors required for PL biosynthesis, and has been shown to have significant beneficial effects in early Alzheimer's disease. Male C57BL/6 mice received a controlled cortical impact injury and then were fed a control diet or a diet enriched with FC for 70 days. FC led to a significantly improved sensorimotor outcome and cognition, reduced lesion size and oligodendrocyte loss, and it restored myelin. It reversed the loss of the synaptic protein synaptophysin and decreased levels of the axon growth inhibitor Nogo-A, thus creating a permissive environment. It decreased microglia activation and the rise in ß-amyloid precursor protein and restored the depressed neurogenesis. The effects of this medical multi-nutrient suggest that support of PL biosynthesis after TBI, a new treatment paradigm, has significant therapeutic potential in this neurological condition for which there is no satisfactory treatment. The multi-nutrient tested has been used in dementia patients, is safe and well-tolerated, which would enable rapid clinical exploration in TBI.

Funding information:
  • Medical Research Council - K-0912(United Kingdom)

Juvenile treatment with mGluR2/3 agonist prevents schizophrenia-like phenotypes in adult by acting through GSK3β.

  • Xing B
  • Neuropharmacology
  • 2018 May 14

Literature context:


Abstract:

Prodromal memory deficits represent an important marker for the development of schizophrenia (SZ), in which glutamatergic hypofunction occurs in the prefrontal cortex (PFC). The mGluR2/3 agonist LY379268 (LY37) attenuates excitatory N-methyl-D-aspartate receptor (NMDAR)-induced neurotoxicity, a central pathological characteristic of glutamatergic hypofunction. We therefore hypothesized that early treatment with LY37 would rescue cognitive deficits and confer benefits for SZ-like behaviors in adults. To test this, we assessed whether early intervention with LY37 would improve learning outcomes in the Morris Water Maze for rats prenatally exposed to methylazoxymethanol acetate (MAM), a neurodevelopmental SZ model. We found that a medium dose of LY37 prevents learning deficits in MAM rats. These effects were mediated through postsynaptic mGluR2/3 via improving GluN2B-NMDAR function by inhibiting glycogen synthase kinase-3β (GSK3β). Furthermore, dendritic spine loss and learning and memory deficits observed in adult MAM rats were restored by juvenile LY37 treatment, which did not change prefrontal neuronal excitability and glutamatergic synaptic transmission in adult normal rats. Our results provide a mechanism for mGluR2/3 agonists against NMDAR hypofunction, which may prove to be beneficial in the prophylactic treatment of SZ.

Funding information:
  • NIAID NIH HHS - R56AI1691785(United States)
  • NIMH NIH HHS - R01 MH085666()

Matrix Metalloproteinase 9 Displays a Particular Time Response to Acute Stress: Variation in Its Levels and Activity Distribution in Rat Hippocampus.

  • Aguayo FI
  • ACS Chem Neurosci
  • 2018 May 16

Literature context:


Abstract:

A single stress exposure facilitates memory formation through neuroplastic processes that reshape excitatory synapses in the hippocampus, probably requiring changes in extracellular matrix components. We tested the hypothesis that matrix metalloproteinase 9 (MMP-9), an enzyme that degrades components of extracellular matrix and synaptic proteins such as β-dystroglycan (β-DG43), changes their activity and distribution in rat hippocampus during the acute stress response. After 2.5 h of restraint stress, we found (i) increased MMP-9 levels and potential activity in whole hippocampal extracts, accompanied by β-DG43 cleavage, and (ii) a significant enhancement of MMP-9 immunoreactivity in dendritic fields such as stratum radiatum and the molecular layer of hippocampus. After 24 h of stress, we found that (i) MMP-9 net activity rises at somatic field, i.e., stratum pyramidale and granule cell layers, and also at synaptic field, mainly stratum radiatum and the molecular layer of hippocampus, and (ii) hippocampal synaptoneurosome fractions are enriched with MMP-9, without variation of its potential enzymatic activity, in accordance with the constant level of cleaved β-DG43. These findings indicate that stress triggers a peculiar timing response in the MMP-9 levels, net activity, and subcellular distribution in the hippocampus, suggesting its involvement in the processing of substrates during the stress response.

Funding information:
  • NIEHS NIH HHS - P42ES04911(United States)

mTORC1 Is Transiently Reactivated in Injured Nerves to Promote c-Jun Elevation and Schwann Cell Dedifferentiation.

  • Norrmén C
  • J. Neurosci.
  • 2018 May 16

Literature context:


Abstract:

Schwann cells (SCs) are endowed with a remarkable plasticity. When peripheral nerves are injured, SCs dedifferentiate and acquire new functions to coordinate nerve repair as so-called repair SCs. Subsequently, SCs redifferentiate to remyelinate regenerated axons. Given the similarities between SC dedifferentiation/redifferentiation in injured nerves and in demyelinating neuropathies, elucidating the signals involved in SC plasticity after nerve injury has potentially wider implications. c-Jun has emerged as a key transcription factor regulating SC dedifferentiation and the acquisition of repair SC features. However, the upstream pathways that control c-Jun activity after nerve injury are largely unknown. We report that the mTORC1 pathway is transiently but robustly reactivated in dedifferentiating SCs. By inducible genetic deletion of the functionally crucial mTORC1-subunit Raptor in mouse SCs (including male and female animals), we found that mTORC1 reactivation is necessary for proper myelin clearance, SC dedifferentiation, and consequently remyelination, without major alterations in the inflammatory response. In the absence of mTORC1 signaling, c-Jun failed to be upregulated correctly. Accordingly, a c-Jun binding motif was found to be enriched in promoters of genes with reduced expression in injured mutants. Furthermore, using cultured SCs, we found that mTORC1 is involved in c-Jun regulation by promoting its translation, possibly via the eIF4F-subunit eIF4A. These results provide evidence that proper c-Jun elevation after nerve injury involves also mTORC1-dependent post-transcriptional regulation to ensure timely dedifferentiation of SCs.SIGNIFICANCE STATEMENT A crucial evolutionary acquisition of vertebrates is the envelopment of axons in myelin sheaths produced by oligodendrocytes in the CNS and Schwann cells (SCs) in the PNS. When myelin is damaged, conduction of action potentials along axons slows down or is blocked, leading to debilitating diseases. Unlike oligodendrocytes, SCs have a high regenerative potential, granted by their remarkable plasticity. Thus, understanding the mechanisms underlying SC plasticity may uncover new therapeutic targets in nerve regeneration and demyelinating diseases. Our work reveals that reactivation of the mTORC1 pathway in SCs is essential for efficient SC dedifferentiation after nerve injury. Accordingly, modulating this signaling pathway might be of therapeutic relevance in peripheral nerve injury and other diseases.

Funding information:
  • Medical Research Council - MC_U120081321(United Kingdom)

A Glial Signature and Wnt7 Signaling Regulate Glioma-Vascular Interactions and Tumor Microenvironment.

  • Griveau A
  • Cancer Cell
  • 2018 May 14

Literature context:


Abstract:

Gliomas comprise heterogeneous malignant glial and stromal cells. While blood vessel co-option is a potential mechanism to escape anti-angiogenic therapy, the relevance of glial phenotype in this process is unclear. We show that Olig2+ oligodendrocyte precursor-like glioma cells invade by single-cell vessel co-option and preserve the blood-brain barrier (BBB). Conversely, Olig2-negative glioma cells form dense perivascular collections and promote angiogenesis and BBB breakdown, leading to innate immune cell activation. Experimentally, Olig2 promotes Wnt7b expression, a finding that correlates in human glioma profiling. Targeted Wnt7a/7b deletion or pharmacologic Wnt inhibition blocks Olig2+ glioma single-cell vessel co-option and enhances responses to temozolomide. Finally, Olig2 and Wnt7 become upregulated after anti-VEGF treatment in preclinical models and patients. Thus, glial-encoded pathways regulate distinct glioma-vascular microenvironmental interactions.

Funding information:
  • Intramural NIH HHS - ES016005(United States)

Kir4.1-Dependent Astrocyte-Fast Motor Neuron Interactions Are Required for Peak Strength.

  • Kelley KW
  • Neuron
  • 2018 Apr 18

Literature context:


Abstract:

Diversified neurons are essential for sensorimotor function, but whether astrocytes become specialized to optimize circuit performance remains unclear. Large fast α-motor neurons (FαMNs) of spinal cord innervate fast-twitch muscles that generate peak strength. We report that ventral horn astrocytes express the inward-rectifying K+ channel Kir4.1 (a.k.a. Kcnj10) around MNs in a VGLUT1-dependent manner. Loss of astrocyte-encoded Kir4.1 selectively altered FαMN size and function and led to reduced peak strength. Overexpression of Kir4.1 in astrocytes was sufficient to increase MN size through activation of the PI3K/mTOR/pS6 pathway. Kir4.1 was downregulated cell autonomously in astrocytes derived from amyotrophic lateral sclerosis (ALS) patients with SOD1 mutation. However, astrocyte Kir4.1 was dispensable for FαMN survival even in the mutant SOD1 background. These findings show that astrocyte Kir4.1 is essential for maintenance of peak strength and suggest that Kir4.1 downregulation might uncouple symptoms of muscle weakness from MN cell death in diseases like ALS.

Funding information:
  • FIC NIH HHS - K01 TW000001(United States)

The Factor Inhibiting HIF Asparaginyl Hydroxylase Regulates Oxidative Metabolism and Accelerates Metabolic Adaptation to Hypoxia.

  • Sim J
  • Cell Metab.
  • 2018 Apr 3

Literature context:


Abstract:

Animals require an immediate response to oxygen availability to allow rapid shifts between oxidative and glycolytic metabolism. These metabolic shifts are highly regulated by the HIF transcription factor. The factor inhibiting HIF (FIH) is an asparaginyl hydroxylase that controls HIF transcriptional activity in an oxygen-dependent manner. We show here that FIH loss increases oxidative metabolism, while also increasing glycolytic capacity, and that this gives rise to an increase in oxygen consumption. We further show that the loss of FIH acts to accelerate the cellular metabolic response to hypoxia. Skeletal muscle expresses 50-fold higher levels of FIH than other tissues: we analyzed skeletal muscle FIH mutants and found a decreased metabolic efficiency, correlated with an increased oxidative rate and an increased rate of hypoxic response. We find that FIH, through its regulation of oxidation, acts in concert with the PHD/vHL pathway to accelerate HIF-mediated metabolic responses to hypoxia.

Funding information:
  • NIGMS NIH HHS - GM062791(United States)

A Circular RNA Protects Dormant Hematopoietic Stem Cells from DNA Sensor cGAS-Mediated Exhaustion.

  • Xia P
  • Immunity
  • 2018 Apr 17

Literature context:


Abstract:

Disrupting the balance between self-renewal and differentiation of hematopoietic stem cells (HSCs) leads to bone marrow failure or hematologic malignancy. However, how HSCs sustain their quiescent state and avoid type I interferon (IFN)-mediated exhaustion remains elusive. Here we defined a circular RNA that we named cia-cGAS that was highly expressed in the nucleus of long-term (LT)-HSCs. Cia-cGAS deficiency in mice caused elevated expression of type I IFNs in bone marrow and led to decreased numbers of dormant LT-HSCs. Under homeostatic conditions, cia-cGAS bound DNA sensor cGAS in the nucleus to block its synthase activity, thereby protecting dormant LT-HSCs from cGAS-mediated exhaustion. Moreover, cia-cGAS harbored a stronger binding affinity to cGAS than self-DNA did and consequently suppressed cGAS-mediated production of type I IFNs in LT-HSCs. Our findings reveal a mechanism by which cia-cGAS inhibits nuclear cGAS by blocking its enzymatic activity and preventing cGAS from recognizing self-DNA to maintain host homeostasis.

Funding information:
  • NIAID NIH HHS - T32 AI060546(United States)

Erythropoietin Intensifies the Proapoptotic Activity of LFM-A13 in Cells and in a Mouse Model of Colorectal Cancer.

  • Tankiewicz-Kwedlo A
  • Int J Mol Sci
  • 2018 Apr 23

Literature context:


Abstract:

The Bruton’s tyrosine kinase (BTK) inhibitor LFM-A13 has been widely employed as an antileukemic agent, but applications in solid cancer have been found recently. The compound promotes apoptosis, has an antiproliferative effect, and increases cancer cell sensitivity to chemotherapy drugs. We decided to assess the impact of the simultaneous use of erythropoietin (Epo) and LFM-A13 on signal transduction in colon DLD-1 and HT-29 cells, as well as in tumor xenografts. The induction of apoptosis by Epo and LFM-A-13 in the cells was confirmed by phosphatidylserine externalization, loss of mitochondrial membrane potential, and modulation of the expression of apoptotic protein BAX and antiapoptotic protein BCL-2 in colon adenocarcinoma cells. Nude mice were inoculated with adenocarcinoma cells and treated with Epo and LFM-A13 in order to evaluate the degree of tumor regression. The simultaneous use of Epo and LFM-A13 severely inhibited cell growth, activated apoptosis, and also inhibited tumor growth in xenografts. The addition of Epo to LFM-A13 intensified the antiproliferative effect of LFM-A13, confirmed by the loss of mitochondrial membrane potential and the accumulation of apoptotic colon cancer cells with externalized phosphatidylserine (PS). These preclinical results suggest that the combination of Epo and LFM-A13 has a high proapoptotic activity and should be tested in the clinic for the treatment of solid tumors such as colon cancer.

Funding information:
  • Intramural NIH HHS - Z99 CA999999(United States)

CC-401 Promotes β-Cell Replication via Pleiotropic Consequences of DYRK1A/B Inhibition.

  • Abdolazimi Y
  • Endocrinology
  • 2018 Mar 5

Literature context:


Abstract:

Pharmacologic expansion of endogenous β-cells is a promising therapeutic strategy for diabetes. To elucidate the molecular pathways that control β-cell growth we screened ∼2,400 bioactive compounds for rat β-cell replication-modulating activity. Numerous hit compounds impaired or promoted rat β-cell replication, including CC-401, an advanced clinical candidate previously characterized as a c-Jun N-terminal kinase (JNK) inhibitor. Surprisingly, CC-401 induced rodent (in vitro and in vivo) and human (in vitro) β-cell replication via dual specificity tyrosine-phosphorylation-regulated kinases (DYRK1A/B) inhibition. In contrast to rat β-cells, which were broadly growth responsive to compound treatment, human β-cell replication was only consistently induced by DYRK1A/B inhibitors. This effect was enhanced by simultaneous glycogen synthase kinase-3β (GSK-3β) or transforming growth factor-β (ALK5/TGF-β) inhibition. Prior work emphasized DYRK1A/B inhibition-dependent activation of nuclear factor of activated T-cells (NFAT) as the primary mechanism of human β-cell replication induction. However, inhibition of NFAT activity had limited impact on CC-401-induced β-cell replication. Consequently, we investigated additional effects of CC-401-dependent DYRK1A/B inhibition. Indeed, CC-401 inhibited DYRK1A-dependent phosphorylation/stabilization of the β-cell replication-inhibitor p27Kip1. Additionally, CC-401 increased expression of numerous replication-promoting genes normally suppressed by the dimerization partner, RB-like, E2F and multi-vulval class B (DREAM) complex, which depends upon DYRK1A/B activity for integrity, including MYBL2 and FOXM1. In summary, we present a compendium of compounds as a valuable resource for manipulating the signaling pathways that control β-cell replication and leverage a novel DYRK1A/B inhibitor (CC-401) to expand our understanding of the molecular pathways that control β-cell growth.

Funding information:
  • NCATS NIH HHS - UL1 TR001085()
  • NIDDK NIH HHS - R01 DK101530()
  • NIDDK NIH HHS - T32 DK007217()
  • NIGMS NIH HHS - R01 GM099801(United States)
  • NIGMS NIH HHS - T32 GM113854()

Gut Epithelial Vitamin D Receptor Regulates Microbiota-Dependent Mucosal Inflammation by Suppressing Intestinal Epithelial Cell Apoptosis.

  • He L
  • Endocrinology
  • 2018 Feb 1

Literature context:


Abstract:

Recent studies show that colonic vitamin D receptor (VDR) signaling protects the mucosal epithelial barrier and suppresses colonic inflammation, but the underlying molecular mechanism remains to be fully understood. To investigate the implication of colonic VDR downregulation seen in patients with inflammatory bowel disease, we assessed the effect of gut epithelial VDR deletion on colonic inflammatory responses in an experimental colitis model. In a 2,4,6-trinitrobenzenesulfonic acid-induced colitis model, mice carrying VDR deletion in gut epithelial cells [VDRflox/flox (VDRf/f);Villin-Cre or VDRΔIEC] or in colonic epithelial cells (VDRf/f;CDX2-Cre or VDRΔCEC) developed more severe clinical colitis than VDRf/f control mice, characterized by more robust T-helper (TH)1 and TH17 responses, with greater increases in mucosal interferon (IFN)-γ+, interleukin (IL)-17+, and IFN-γ+IL-17+ T cells. Accompanying the severe mucosal inflammation was more profound colonic epithelial cell apoptosis in the mutant mice. Treatment with caspase inhibitor Q-VD-OPh dramatically reduced colitis severity and attenuated TH1 and TH17 responses in VDRΔCEC mice. The blockade of cell apoptosis also prevented the increase in mucosal CD11b+CD103+ dendritic cells (DCs), known to be critical for TH17-cell activation. Moreover, depletion of gut commensal bacteria with antibiotics eliminated the robust TH1 and TH17 responses and CD11b+CD103+ DC induction. Taken together, these observations demonstrate that gut epithelial VDR deletion aggravates epithelial cell apoptosis, resulting in increases in mucosal barrier permeability. Consequently, invading luminal bacteria activate CD11b+CD103+ DCs, which promote mucosal TH1 and TH17 responses. Therefore, gut epithelial VDR signaling controls mucosal inflammation by suppressing epithelial cell apoptosis.

Funding information:
  • Intramural NIH HHS - ZID BC010990-06(United States)
  • NCI NIH HHS - R01 CA180087()
  • NIDDK NIH HHS - P30 DK042086()

Chromatin Modification and Global Transcriptional Silencing in the Oocyte Mediated by the mRNA Decay Activator ZFP36L2.

  • Dumdie JN
  • Dev. Cell
  • 2018 Feb 5

Literature context:


Abstract:

Global transcriptional silencing is a highly conserved mechanism central to the oocyte-to-embryo transition. We report the unexpected discovery that global transcriptional silencing in oocytes depends on an mRNA decay activator. Oocyte-specific loss of ZFP36L2 an RNA-binding protein that promotes AU-rich element-dependent mRNA decay prevents global transcriptional silencing and causes oocyte maturation and fertilization defects, as well as complete female infertility in the mouse. Single-cell RNA sequencing revealed that ZFP36L2 downregulates mRNAs encoding transcription and chromatin modification regulators, including a large group of mRNAs for histone demethylases targeting H3K4 and H3K9, which we show are bound and degraded by ZFP36L2. Oocytes lacking Zfp36l2 fail to accumulate histone methylation at H3K4 and H3K9, marks associated with the transcriptionally silent, developmentally competent oocyte state. Our results uncover a ZFP36L2-dependent mRNA decay mechanism that acts as a developmental switch during oocyte growth, triggering wide-spread shifts in chromatin modification and global transcription.

Funding information:
  • NICHD NIH HHS - K12 HD001259()
  • NIGMS NIH HHS - R35 GM118069()
  • NIMH NIH HHS - R33MH083521(United States)

Basal Cells Show Increased Expression of Aromatase and Estrogen Receptor α in Prostate Epithelial Lesions of Male Aging Rats.

  • Morais-Santos M
  • Endocrinology
  • 2018 Feb 1

Literature context:


Abstract:

Besides androgens, estrogen signaling plays a key role in normal development and pathologies of the prostate. Irreversible synthesis of estrogens from androgens is catalyzed by aromatase. Interestingly, animals lacking aromatase do not develop cancer or prostatitis, whereas those with overexpression of aromatase and, consequently, high estrogen levels develop prostatitis and squamous metaplasia via estrogen receptor 1 (ERα). Even with this evidence, the aromatase expression in the prostate is controversial. Moreover, little is known about the occurrence of age-dependent variation of aromatase and its association with histopathological changes commonly found in advanced age, a knowledge gap that is addressed herein. For this purpose, the immunoexpression of aromatase was evaluated in the prostatic complex of young adult to senile Wistar rats. ERα was also investigated, to extend our understanding of estrogen responsiveness in the prostate. Moderate cytoplasmic immunoreactivity for aromatase was detected in the glandular epithelium. Eventually, some basal cells showed intense staining for aromatase. The expression pattern for aromatase appeared similar in the normal epithelium when young and senile rats were compared; this result was corroborated by Western blotting. Conversely, in senile rats, there was an increase in the frequency of basal cells intensely stained for aromatase, which appeared concentrated in areas of intraepithelial proliferation and prostatitis. These punctual areas also presented increased ERα positivity. Together, these findings suggest a plausible source for hormonal imbalance favoring estrogen production, which, by acting through ERα, may favor the development of prostatic lesions commonly found in advanced age.

The soluble form of LOTUS inhibits Nogo receptor-mediated signaling by interfering with the interaction between Nogo receptor type 1 and p75 neurotrophin receptor.

  • Kawakami Y
  • J. Neurosci.
  • 2018 Feb 9

Literature context:


Abstract:

Nogo receptor type 1 (NgR1) is known to inhibit neuronal regeneration in the CNS. We have previously identified lateral olfactory tract usher substance (LOTUS) interacts with NgR1 and inhibits its function by blocking its ligand binding. Therefore, LOTUS is expected to have therapeutic potential for the promotion of neuronal regeneration. However, it remains unknown whether the soluble form of LOTUS (s-LOTUS) also has an inhibitory action on NgR1 function as a candidate for therapeutic agents. Here, we show that s-LOTUS inhibits NgR1-mediated signaling by inhibiting the molecular interaction between NgR1 and its co-receptor p75 neurotrophin receptor (p75NTR). In contrast to the membrane-bound form of LOTUS, s-LOTUS did not block ligand binding to NgR1. However, we identified p75NTR as a novel LOTUS binding partner, and found that s-LOTUS suppressed the interaction between p75NTR and NgR1. s-LOTUS inhibited myelin-associated inhibitor (MAI)-induced RhoA activation in murine cortical neurons. Functional analyses revealed that s-LOTUS inhibited MAI-induced growth cone collapse and neurite outgrowth inhibition in chick DRG neurons. In addition, while olfactory bulb (OB) neurons of lotus-KO mice are sensitive to MAI due to a lack of LOTUS expression, treatment with s-LOTUS inhibited MAI-induced growth cone collapse in these neurons. Finally, we observed that s-LOTUS promoted axonal regeneration in optic nerve crush injury of mice (either sex). These findings suggest that s-LOTUS inhibits NgR1-mediated signaling possibly by interfering with the interaction between NgR1 and p75NTR Thus, s-LOTUS may have potential as a therapeutic agent for neuronal regeneration in the damaged CNS.SIGNIFICANCE STATEMENTNogo receptor type 1 (NgR1) is a well-known receptor to inhibit neuronal regeneration in the CNS. Because the membrane-bound form of LOTUS antagonizes NgR1 through a cis-type molecular interaction between LOTUS and NgR1, the soluble form of LOTUS (s-LOTUS) is expected to be a therapeutic agent for neuronal regeneration. In our present study, we show that s-LOTUS inhibits the interaction between NgR1 and p75NTR, NgR1 ligand-induced RhoA activation, growth cone collapse and neurite outgrowth inhibition, and promotes axonal regeneration. Our results indicate that s-LOTUS inhibits NgR1-mediated signaling through a trans-type molecular interaction between LOTUS and NgR1, and therefore, s-LOTUS may have therapeutic potential for neuronal regeneration.

Funding information:
  • NIGMS NIH HHS - R01-GM083204(United States)

Altered Baseline and Nicotine-Mediated Behavioral and Cholinergic Profiles in ChAT-Cre Mouse Lines.

  • Chen E
  • J. Neurosci.
  • 2018 Feb 28

Literature context:


Abstract:

The recent development of transgenic rodent lines expressing cre recombinase in a cell-specific manner, along with advances in engineered viral vectors, has permitted in-depth investigations into circuit function. However, emerging evidence has begun to suggest that genetic modifications may introduce unexpected caveats. In the current studies, we sought to extensively characterize male and female mice from both the ChAT(BAC)-Cre mouse line, created with the bacterial artificial chromosome (BAC) method, and ChAT(IRES)-Cre mouse line, generated with the internal ribosome entry site (IRES) method. ChAT(BAC)-Cre transgenic and wild-type mice did not differ in general locomotor behavior, anxiety measures, drug-induced cataplexy, nicotine-mediated hypolocomotion, or operant food training. However, ChAT(BAC)-Cre transgenic mice did exhibit significant deficits in intravenous nicotine self-administration, which paralleled an increase in vesicular acetylcholine transporter and choline acetyltransferase (ChAT) hippocampal expression. For the ChAT(IRES)-Cre line, transgenic mice exhibited deficits in baseline locomotor, nicotine-mediated hypolocomotion, and operant food training compared with wild-type and hemizygous littermates. No differences among ChAT(IRES)-Cre wild-type, hemizygous, and transgenic littermates were found in anxiety measures, drug-induced cataplexy, and nicotine self-administration. Given that increased cre expression was present in the ChAT(IRES)-Cre transgenic mice, as well as a decrease in ChAT expression in the hippocampus, altered neuronal function may underlie behavioral phenotypes. In contrast, ChAT(IRES)-Cre hemizygous mice were more similar to wild-type mice in both protein expression and the majority of behavioral assessments. As such, interpretation of data derived from ChAT-Cre rodents must consider potential limitations dependent on the line and/or genotype used in research investigations.SIGNIFICANCE STATEMENT Altered baseline and/or nicotine-mediated behavioral profiles were discovered in transgenic mice from the ChAT(BAC)-Cre and ChAT(IRES)-Cre lines. Given that these cre-expressing mice have become increasingly used by the scientific community, either independently with chemicogenetic and optogenetic viral vectors or crossed with other transgenic lines, the current studies highlight important considerations for the interpretation of data from previous and future experimental investigations. Moreover, the current findings detail the behavioral effects of either increased or decreased baseline cholinergic signaling mechanisms on locomotor, anxiety, learning/memory, and intravenous nicotine self-administration behaviors.

Funding information:
  • NIAID NIH HHS - AI07090-31(United States)

Lithium Inhibits GSK3β and Augments GluN2A Receptor Expression in the Prefrontal Cortex.

  • Monaco SA
  • Front Cell Neurosci
  • 2018 Feb 17

Literature context:


Abstract:

Glycogen synthase kinase 3β (GSK3β) is a highly conserved serine/threonine kinase that has been implicated in both psychiatric and neurodegenerative diseases including schizophrenia, bipolar disorder, and Alzheimer's disease; therefore regulating its activity has become an important strategy for treatment of cognitive impairments in these disorders. This study examines the effects of lithium on GSK3β and its interaction with β-catenin and NMDA receptors within the prefrontal cortex. Lithium, a clinically relevant drug commonly prescribed as a mood stabilizer for psychiatric disorders, significantly increased levels of phosphorylated GSK3β serine 9, an inhibitory phosphorylation site, and decreased β-catenin ser33/37/thr41 phosphorylation in vitro, indicating GSK3β inhibition and reduced β-catenin degradation. GluN2A subunit levels were concurrently increased following lithium treatment. Similar alterations were also demonstrated in vivo; lithium administration increased GSK3β serine 9 phosphorylation and GluN2A levels, suggesting a reduced GSK3β activity and augmented GluN2A expression. Correspondingly, we observed that the amplitudes of evoked GluN2A-mediated excitatory postsynaptic currents in mPFC pyramidal neurons were significantly increased following lithium administration. Our data suggest that GSK3β activity negatively regulates GluN2A expression, likely by mediating upstream β-catenin phosphorylation, in prefrontal cortical neurons. Furthermore, our biochemical and electrophysiological experiments demonstrate that lithium mediates a specific increase in GluN2A subunit expression, ultimately augmenting GluN2A-mediated currents in the prefrontal cortex.

Funding information:
  • NCI NIH HHS - CA100193(United States)

Impaired Processing of Prohormones in Secretogranin III-Null Mice Causes Maladaptation to an Inadequate Diet and Stress.

  • Maeda Y
  • Endocrinology
  • 2018 Feb 1

Literature context:


Abstract:

Secretogranin III (SgIII), a member of the granin family, binds both to another granin, chromogranin A (CgA), and to a cholesterol-rich membrane that is destined for secretory granules (SGs). The knockdown of SgIII in adrenocorticotropic hormone (ACTH)-producing AtT-20 cells largely impairs the regulated secretion of CgA and ACTH. To clarify the physiological roles of SgIII in vivo, we analyzed hormone secretion and SG biogenesis in newly established SgIII-knockout (KO) mice. Although the SgIII-KO mice were viable and fertile and exhibited no overt abnormalities under ordinary rearing conditions, a high-fat/high-sucrose diet caused pronounced obesity in the mice. Furthermore, in the SgIII-KO mice compared with wild-type (WT) mice, the stimulated secretion of active insulin decreased substantially, whereas the storage of proinsulin increased in the islets. The plasma ACTH was also less elevated in the SgIII-KO mice than in the WT mice after chronic restraint stress, whereas the storage level of the precursor proopiomelanocortin in the pituitary gland was somewhat increased. These findings suggest that the lack of SgIII causes maladaptation of endocrine cells to an inadequate diet and stress by impairing the proteolytic conversion of prohormones in SGs, whereas SG biogenesis and the basal secretion of peptide hormones under ordinary conditions are ensured by the compensatory upregulation of other residual granins or factors.

Funding information:
  • Intramural NIH HHS - (United States)

Glutamate-induced and NMDA receptor-mediated neurodegeneration entails P2Y1 receptor activation.

  • Simões AP
  • Cell Death Dis
  • 2018 Feb 20

Literature context:


Abstract:

Despite the characteristic etiologies and phenotypes, different brain disorders rely on common pathogenic events. Glutamate-induced neurotoxicity is a pathogenic event shared by different brain disorders. Another event occurring in different brain pathological conditions is the increase of the extracellular ATP levels, which is now recognized as a danger and harmful signal in the brain, as heralded by the ability of P2 receptors (P2Rs) to affect a wide range of brain disorders. Yet, how ATP and P2R contribute to neurodegeneration remains poorly defined. For that purpose, we now examined the contribution of extracellular ATP and P2Rs to glutamate-induced neurodegeneration. We found both in vitro and in vivo that ATP/ADP through the activation of P2Y1R contributes to glutamate-induced neuronal death in the rat hippocampus. We found in cultured rat hippocampal neurons that the exposure to glutamate (100 µM) for 30 min triggers a sustained increase of extracellular ATP levels, which contributes to NMDA receptor (NMDAR)-mediated hippocampal neuronal death through the activation of P2Y1R. We also determined that P2Y1R is involved in excitotoxicity in vivo as the blockade of P2Y1R significantly attenuated rat hippocampal neuronal death upon the systemic administration of kainic acid or upon the intrahippocampal injection of quinolinic acid. This contribution of P2Y1R fades with increasing intensity of excitotoxic conditions, which indicates that P2Y1R is not contributing directly to neurodegeneration, rather behaving as a catalyst decreasing the threshold from which glutamate becomes neurotoxic. Moreover, we unraveled that such excitotoxicity process began with an early synaptotoxicity that was also prevented/attenuated by the antagonism of P2Y1R, both in vitro and in vivo. This should rely on the observed glutamate-induced calpain-mediated axonal cytoskeleton damage, most likely favored by a P2Y1R-driven increase of NMDAR-mediated Ca2+ entry selectively in axons. This may constitute a degenerative mechanism shared by different brain diseases, particularly relevant at initial pathogenic stages.

Funding information:
  • NCRR NIH HHS - 5P20-RR021940(United States)

Bcl-xL dependency coincides with the onset of neurogenesis in the developing mammalian spinal cord.

  • Fogarty LC
  • Mol. Cell. Neurosci.
  • 2018 Jan 12

Literature context:


Abstract:

The bcl-2 family of survival and death promoting proteins play a key role in regulating cell numbers during nervous system development. Bcl-xL, an anti-apoptotic bcl-2 family member is highly expressed in the developing nervous system. However; the early embryonic lethality of the bcl-x germline null mouse precluded an investigation into its role in nervous system development. To identify the role of bcl-x in spinal cord neurogenesis, we generated a central nervous system-specific bcl-x conditional knockout (BKO) mouse. Apoptotic cell death in the BKO embryo was initially detected at embryonic day 11 (E11) in the ventrolateral aspect of the spinal cord corresponding to the location of motor neurons. Apoptosis reached its peak at E13 having spread across the ventral and into the dorsal spinal cord. By E18, the wave of apoptosis had passed and only a few apoptotic cells were observed. The duration and direction of spread of apoptosis across the spinal cord is consistent with the spatial and temporal sequence of neuronal differentiation. Motor neurons, the first neurons to become post mitotic in the spinal cord, were also the first apoptotic cells. As neurogenesis spread across the spinal cord, later born neuronal populations such as Lim2+ interneurons were also affected. The onset of apoptosis occurred in cells that had exited the cell cycle within the previous 24h and initiated neural differentiation as demonstrated by BrdU birthdating and βIII tubulin immunohistochemistry. This data demonstrates that spinal cord neurons become Bcl-xL dependent at an early post mitotic stage in developmental neurogenesis.

Funding information:
  • NIAID NIH HHS - R01 AI074847(United States)

An Image-Based miRNA Screen Identifies miRNA-135s As Regulators of CNS Axon Growth and Regeneration by Targeting Krüppel-like Factor 4.

  • van Battum EY
  • J. Neurosci.
  • 2018 Jan 17

Literature context:


Abstract:

During embryonic development, axons extend over long distances to establish functional connections. In contrast, axon regeneration in the adult mammalian CNS is limited in part by a reduced intrinsic capacity for axon growth. Therefore, insight into the intrinsic control of axon growth may provide new avenues for enhancing CNS regeneration. Here, we performed one of the first miRNome-wide functional miRNA screens to identify miRNAs with robust effects on axon growth. High-content screening identified miR-135a and miR-135b as potent stimulators of axon growth and cortical neuron migration in vitro and in vivo in male and female mice. Intriguingly, both of these developmental effects of miR-135s relied in part on silencing of Krüppel-like factor 4 (KLF4), a well known intrinsic inhibitor of axon growth and regeneration. These results prompted us to test the effect of miR-135s on axon regeneration after injury. Our results show that intravitreal application of miR-135s facilitates retinal ganglion cell (RGC) axon regeneration after optic nerve injury in adult mice in part by repressing KLF4. In contrast, depletion of miR-135s further reduced RGC axon regeneration. Together, these data identify a novel neuronal role for miR-135s and the miR-135-KLF4 pathway and highlight the potential of miRNAs as tools for enhancing CNS axon regeneration.SIGNIFICANCE STATEMENT Axon regeneration in the adult mammalian CNS is limited in part by a reduced intrinsic capacity for axon growth. Therefore, insight into the intrinsic control of axon growth may provide new avenues for enhancing regeneration. By performing an miRNome-wide functional screen, our studies identify miR-135s as stimulators of axon growth and neuron migration and show that intravitreal application of these miRNAs facilitates CNS axon regeneration after nerve injury in adult mice. Intriguingly, these developmental and regeneration-promoting effects rely in part on silencing of Krüppel-like factor 4 (KLF4), a well known intrinsic inhibitor of axon regeneration. Our data identify a novel neuronal role for the miR-135-KLF4 pathway and support the idea that miRNAs can be used for enhancing CNS axon regeneration.

Funding information:
  • NCI NIH HHS - R01 CA112054(United States)

Inhibition of Inwardly Rectifying Potassium (Kir) 4.1 Channels Facilitates Brain-Derived Neurotrophic Factor (BDNF) Expression in Astrocytes.

  • Kinboshi M
  • Front Mol Neurosci
  • 2018 Jan 24

Literature context:


Abstract:

Inwardly rectifying potassium (Kir) 4.1 channels in astrocytes regulate neuronal excitability by mediating spatial potassium buffering. Although dysfunction of astrocytic Kir4.1 channels is implicated in the development of epileptic seizures, the functional mechanisms of Kir4.1 channels in modulating epileptogenesis remain unknown. We herein evaluated the effects of Kir4.1 inhibition (blockade and knockdown) on expression of brain-derived neurotrophic factor (BDNF), a key modulator of epileptogenesis, in the primary cultures of mouse astrocytes. For blockade of Kir4.1 channels, we tested several antidepressant agents which reportedly bound to and blocked Kir4.1 channels in a subunit-specific manner. Treatment of astrocytes with fluoxetine enhanced BDNF mRNA expression in a concentration-dependent manner and increased the BDNF protein level. Other antidepressants (e.g., sertraline and imipramine) also increased the expression of BDNF mRNA with relative potencies similar to those for inhibition of Kir4.1 channels. In addition, suppression of Kir4.1 expression by the transfection of small interfering RNA (siRNA) targeting Kir4.1 significantly increased the mRNA and protein levels of BDNF. The BDNF induction by Kir4.1 siRNA transfection was suppressed by the MEK1/2 inhibitor U0126, but not by the p38 MAPK inhibitor SB202190 or the JNK inhibitor SP600125. The present results demonstrated that inhibition of Kir4.1 channels facilitates BDNF expression in astrocytes primarily by activating the Ras/Raf/MEK/ERK pathway, which may be linked to the development of epilepsy and other neuropsychiatric disorders.

Funding information:
  • NINDS NIH HHS - R56 NS021072(United States)

Dual effect of serotonin on the dendritic growth of cultured hippocampal neurons: Involvement of 5-HT1A and 5-HT7 receptors.

  • Rojas PS
  • Mol. Cell. Neurosci.
  • 2017 Dec 4

Literature context:


Abstract:

Serotonin acts through its receptors (5-HTRs) to shape brain networks during development and modulates essential functions in mature brain. The 5-HT1AR is mainly located at soma of hippocampal neurons early during brain development and its expression gradually shifts to dendrites during postnatal development. The 5-HT7R expressed early during hippocampus development, shows a progressive reduction in its expression postnatally. Considering these changes during development, we evaluated in cultured hippocampal neurons whether the 5-HT1AR and 5-HT7R change their expression, modulate dendritic growth, and activate signaling pathways such as ERK1/2, AKT/GSK3β and LIMK/cofilin, which may sustain dendrite outgrowth by controlling cytoskeleton dynamics. We show that mRNA levels of both receptors increase between 2 and 7 DIV; however only protein levels of 5-HT7R increase significantly at 7 DIV. The 5-HT1AR is preferentially distributed in the soma, while 5-HT7R displays a somato-dendritic localization at 7 DIV. Through stimulation with 5-HT at 7 DIV during 24h and using specific antagonists, we determined that 5-HT1AR decreases the number of primary and secondary dendrites and restricts the growth of primary dendrites. The activation of 5-HT1AR and 5-HT7R promotes the growth of short secondary dendrites and triggers ERK1/2 and AKT phosphorylation through MEK and PI3K activation respectively; without changes in the phosphorylation of LIMK and cofilin. We conclude that 5-HT1AR restricts dendritogenesis and outgrowth of primary dendrites, but that both 5-HT1AR and 5-HT7R promote secondary dendrite outgrowth. These data support the role of 5-HT in neuronal outgrowth during development and provide insight into cellular basis of neurodevelopmental disorders.

Down-regulation of NTPDase2 and ADP-sensitive P2 Purinoceptors Correlate with Severity of Symptoms during Experimental Autoimmune Encephalomyelitis.

  • Jakovljevic M
  • Front Cell Neurosci
  • 2017 Nov 23

Literature context:


Abstract:

The present study explores tissue and cellular distribution of ectonucleoside triphosphate diphosphohydrolase 2 (NTPDase2) and the gene and protein expression in rat spinal cord during the course of experimental autoimmune encephalomyelitis (EAE). Given that NTPDase2 hydrolyzes ATP with a transient accumulation of ADP, the expression of ADP-sensitive P2 purinoceptors was analyzed as well. The autoimmune disease was actively induced in Dark Agouti female rats and the changes were analyzed 10, 15 and 29 days after the induction. These selected time points correspond to the onset ( Eo ), peak ( Ep ) and recovery ( Er ) from EAE. In control animals, NTPDase2 was confined in the white matter, in most of the glial fibrillary acidic protein (GFAP)-immunoreactive (ir) astrocytes and in a considerable number of nestin-ir cells, while the other cell types were immunonegative. Immunoreactivity corresponding to NTPDase2 decreased significantly at Eo and Ep and then returned to the baseline levels at Er . The preservation of the proportion of GFAP single-labeled and GFAP/NTPDase2 double-labeled elements along the course of EAE indicated that changes in NTPDase2-ir occurred at fibrous astrocytes that typically express NTPDase2 in normal conditions. Significant downregulation of P2Y1 and P2Y12 receptor proteins at Eo and several-fold induction of P2Y12 and P2Y13 receptor proteins at Ep and/or Er were observed implying that the pathophysiological process in EAE may be linked to ADP signaling. Cell-surface expression of NTPDase2, NTPDase1/CD39 and ecto-5'-nucleotidase (eN/CD73) was analyzed in CD4+ T cells of a draining lymph node by fluorescence-activated cell sorting. The induction of EAE was associated with a transient decrease in a number of CD4+ NTPDase2+ T cells in a draining lymph node, whereas the recovery was characterized by an increase in NTPDase2+ cells in both CD4+ and CD4- cell populations. The opposite was found for NTPDase1/CD39+ and eN/CD73+ cells, which slightly increased in number with progression of the disease, particularly in CD4- cells, and then decreased in the recovery. Finally, CD4+ NTPDase2+ cells were never observed in the spinal cord parenchyma. Taken together, our results suggest that the process of neuroinflammation in EAE may be associated with altered ADP signaling.

Epigenetic mechanisms underlying NMDA receptor hypofunction in the prefrontal cortex of juvenile animals in the MAM model for schizophrenia.

  • Gulchina Y
  • J. Neurochem.
  • 2017 Nov 19

Literature context:


Abstract:

Schizophrenia (SCZ) is characterized not only by psychosis, but also by working memory and executive functioning deficiencies, processes that rely on the prefrontal cortex (PFC). Because these cognitive impairments emerge prior to psychosis onset, we investigated synaptic function during development in the neurodevelopmental methylazoxymethanol (MAM) model for SCZ. Specifically, we hypothesize that N-methyl-D-aspartate receptor (NMDAR) hypofunction is attributable to reductions in the NR2B subunit through aberrant epigenetic regulation of gene expression, resulting in deficient synaptic physiology and PFC-dependent cognitive dysfunction, a hallmark of SCZ. Using western blot and whole-cell patch-clamp electrophysiology, we found that the levels of synaptic NR2B protein are significantly decreased in juvenile MAM animals, and the function of NMDARs is substantially compromised. Both NMDA-mEPSCs and synaptic NMDA-eEPSCs are significantly reduced in prelimbic PFC (plPFC). This protein loss during the juvenile period is correlated with an aberrant increase in enrichment of the epigenetic transcriptional repressor RE1-silencing transcription factor (REST) and the repressive histone marker H3K27me3 at the Grin2b promoter, as assayed by ChIP-quantitative polymerase chain reaction. Glutamate hypofunction has been a prominent hypothesis in the understanding of SCZ pathology; however, little attention has been given to the NMDAR system in the developing PFC in models for SCZ. Our work is the first to confirm that NMDAR hypofunction is a feature of early postnatal development, with epigenetic hyper-repression of the Grin2b promoter being a contributing factor. The selective loss of NR2B protein and subsequent synaptic dysfunction weakens plPFC function during development and may underlie early cognitive impairments in SCZ models and patients. Read the Editorial Highlight for this article on page 264.

Funding information:
  • NIMH NIH HHS - R01 MH085666()

Extracellular ATP induces graded reactive response of astrocytes and strengthens their antioxidative defense in vitro.

  • Adzic M
  • J. Neurosci. Res.
  • 2017 Nov 16

Literature context:


Abstract:

It is widely accepted that adenosine triphosphate (ATP) acts as a universal danger-associated molecular pattern with several known mechanisms for immune cell activation. In the central nervous system, ATP activates microglia and astrocytes and induces a neuroinflammatory response. The aim of the present study was to describe responses of isolated astrocytes to increasing concentrations of ATP (5 µM to 1 mM), which were intended to mimic graded intensity of the extracellular stimulus. The results show that ATP induces graded activation response of astrocytes in terms of the cell proliferation, stellation, shape remodeling, and underlying actin and GFAP filament rearrangement, although the changes occurred without an apparent increase in GFAP and actin protein expression. On the other hand, ATP in the range of applied concentrations did not evoke IL-1β release from cultured astrocytes, nor did it modify the release from LPS and LPS+IFN-γ-primed astrocytes. ATP did not promote astrocyte migration in the wound-healing assay, nor did it increase production of reactive oxygen and nitrogen species and lipid peroxidation. Instead, ATP strengthened the antioxidative defense of astrocytes by inducing Cu/ZnSOD and MnSOD activities and by increasing their glutathione content. Our current results suggest that although ATP triggers several attributes of activated astrocytic phenotype with a magnitude that increases with the concentration, it is not sufficient to induce full-blown reactive phenotype of astrocytes in vitro. © 2016 Wiley Periodicals, Inc.

Targeting Interleukin-1β Protects from Aortic Aneurysms Induced by Disrupted Transforming Growth Factor β Signaling.

  • Da Ros F
  • Immunity
  • 2017 Nov 21

Literature context:


Abstract:

Aortic aneurysms are life-threatening conditions with effective treatments mainly limited to emergency surgery or trans-arterial endovascular stent grafts, thus calling for the identification of specific molecular targets. Genetic studies have highlighted controversial roles of transforming growth factor β (TGF-β) signaling in aneurysm development. Here, we report on aneurysms developing in adult mice after smooth muscle cell (SMC)-specific inactivation of Smad4, an intracellular transducer of TGF-β. The results revealed that Smad4 inhibition activated interleukin-1β (IL-1β) in SMCs. This danger signal later recruited innate immunity in the adventitia through chemokine (C-C motif) ligand 2 (CCL2) and modified the mechanical properties of the aortic wall, thus favoring vessel dilation. SMC-specific Smad4 deletion in Il1r1- or Ccr2-null mice resulted in milder aortic pathology. A chronic treatment with anti-IL-1β antibody effectively hampered aneurysm development. These findings identify a mechanistic target for controlling the progression of aneurysms with compromised TGF-β signaling, such as those driven by SMAD4 mutations.

Funding information:
  • NINDS NIH HHS - R01 NS039600(United States)

Epigenetic control of early neurodegenerative events in diabetic retinopathy by the histone deacetylase SIRT6.

  • Zorrilla-Zubilete MA
  • J. Neurochem.
  • 2017 Oct 20

Literature context:


Abstract:

Diabetic retinopathy (DR) is one of the common complications associated with diabetes mellitus and the leading cause of blindness worldwide. Recent research has demonstrated that DR is not only a microvascular disease but may be a result of neurodegenerative processes. Moreover, glucose-induced neuron and glial cell damage may occur shortly after the onset of diabetes which makes the disease hard to diagnose at early stages. SIRT6, a NAD-dependent sirtuin deacylase, modulates aging, energy metabolism, and neurodegeneration. In previous studies we showed that SIRT6 deficiency causes major retinal transmission defects, changes in the expression of glycolytic genes, and elevated levels of apoptosis. Given the importance of glucose availability for retinal function and the critical role of SIRT6 in modulating glycolysis, we aimed to analyze SIRT6 participation in the molecular machinery that regulates the development of experimental DR. Using non-obese diabetic mice, we determined by western blot that 2 weeks after the onset of the disease, high glucose concentrations induced retinal increase in a neovascularization promoting factor (vascular endothelial growth factor, VEGF), and the loss of a neuroprotective factor (brain-derived neurotrophic factor, BDNF) associated with reduced levels of SIRT6 and increased acetylation levels of its substrates (H3K9 and H3K56) suggesting a deregulation of key neural factors. Noteworthy, retinas from CNS conditionally deleted SIRT6 mice showed a resemblance to diabetic retinas exhibiting lower protein levels of BDNF factor and increased protein levels of VEGF. Moreover, cultured Müller glial cells subjected to high glucose concentrations exhibited decreased levels of SIRT6 and increased levels of H3K56 acetylation. In addition, the increment of VEGF levels induced by high glucose was reverted by the over-expression of SIRT6 in this cell type. Accordingly, siRNA experiments showed that, when SIRT6 was silenced, VEGF levels increased. Our findings suggest that epigenetically regulated neurodegenerative events may occur at an early diabetic stage prior to the characteristic proliferative and vascular changes observed at a later diabetic stage.

Activation of Ras-ERK Signaling and GSK-3 by Amyloid Precursor Protein and Amyloid Beta Facilitates Neurodegeneration in Alzheimer's Disease.

  • Kirouac L
  • eNeuro
  • 2017 Oct 27

Literature context:


Abstract:

It is widely accepted that amyloid β (Aβ) generated from amyloid precursor protein (APP) oligomerizes and fibrillizes to form neuritic plaques in Alzheimer's disease (AD), yet little is known about the contribution of APP to intracellular signaling events preceding AD pathogenesis. The data presented here demonstrate that APP expression and neuronal exposure to oligomeric Aβ42 enhance Ras/ERK signaling cascade and glycogen synthase kinase 3 (GSK-3) activation. We find that RNA interference (RNAi)-directed knockdown of APP in B103 rat neuroblastoma cells expressing APP inhibits Ras-ERK signaling and GSK-3 activation, indicating that APP acts upstream of these signal transduction events. Both ERK and GSK-3 are known to induce hyperphosphorylation of tau and APP at Thr668, and our findings suggest that aberrant signaling by APP facilitates these events. Supporting this notion, analysis of human AD brain samples showed increased expression of Ras, activation of GSK-3, and phosphorylation of APP and tau, which correlated with Aβ levels in the AD brains. Furthermore, treatment of primary rat neurons with Aβ recapitulated these events and showed enhanced Ras-ERK signaling, GSK-3 activation, upregulation of cyclin D1, and phosphorylation of APP and tau. The finding that Aβ induces Thr668 phosphorylation on APP, which enhances APP proteolysis and Aβ generation, denotes a vicious feedforward mechanism by which APP and Aβ promote tau hyperphosphorylation and neurodegeneration in AD. Based on these results, we hypothesize that aberrant proliferative signaling by APP plays a fundamental role in AD neurodegeneration and that inhibition of this would impede cell cycle deregulation and neurodegeneration observed in AD.

Global Inhibition with Specific Activation: How p53 and MYC Redistribute the Transcriptome in the DNA Double-Strand Break Response.

  • Porter JR
  • Mol. Cell
  • 2017 Sep 21

Literature context:


Abstract:

In response to stresses, cells often halt normal cellular processes, yet stress-specific pathways must bypass such inhibition to generate effective responses. We investigated how cells redistribute global transcriptional activity in response to DNA damage. We show that an oscillatory increase of p53 levels in response to double-strand breaks drives a counter-oscillatory decrease of MYC levels. Using RNA sequencing (RNA-seq) of newly synthesized transcripts, we found that p53-mediated reduction of MYC suppressed general transcription, with the most highly expressed transcripts reduced to a greater extent. In contrast, upregulation of p53 targets was relatively unaffected by MYC suppression. Reducing MYC during the DNA damage response was important for cell-fate regulation, as counteracting MYC repression reduced cell-cycle arrest and elevated apoptosis. Our study shows that global inhibition with specific activation of transcriptional pathways is important for the proper response to DNA damage; this mechanism may be a general principle used in many stress responses.

Dual function of the PI3K-Akt-mTORC1 axis in myelination of the peripheral nervous system.

  • Figlia G
  • Elife
  • 2017 Sep 7

Literature context:


Abstract:

Myelination is a biosynthetically demanding process in which mTORC1, the gatekeeper of anabolism, occupies a privileged regulatory position. We have shown previously that loss of mTORC1 function in Schwann cells (SCs) hampers myelination. Here, we genetically disrupted key inhibitory components upstream of mTORC1, TSC1 or PTEN, in mouse SC development, adult homeostasis, and nerve injury. Surprisingly, the resulting mTORC1 hyperactivity led to markedly delayed onset of both developmental myelination and remyelination after injury. However, if mTORC1 was hyperactivated after myelination onset, radial hypermyelination was observed. At early developmental stages, physiologically high PI3K-Akt-mTORC1 signaling suppresses expression of Krox20 (Egr2), the master regulator of PNS myelination. This effect is mediated by S6K and contributes to control mechanisms that keep SCs in a not-fully differentiated state to ensure proper timing of myelination initiation. An ensuing decline in mTORC1 activity is crucial to allow myelination to start, while remaining mTORC1 activity drives myelin growth.

Chronic Stress Triggers Expression of Immediate Early Genes and Differentially Affects the Expression of AMPA and NMDA Subunits in Dorsal and Ventral Hippocampus of Rats.

  • Pacheco A
  • Front Mol Neurosci
  • 2017 Aug 29

Literature context:


Abstract:

Previous studies in rats have demonstrated that chronic restraint stress triggers anhedonia, depressive-like behaviors, anxiety and a reduction in dendritic spine density in hippocampal neurons. In this study, we compared the effect of repeated stress on the expression of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptor subunits in dorsal and ventral hippocampus (VH). Adult male Sprague-Dawley rats were randomly divided into control and stressed groups, and were daily restrained in their motion (2.5 h/day) during 14 days. We found that chronic stress promotes an increase in c-Fos mRNA levels in both hippocampal areas, although it was observed a reduction in the immunoreactivity at pyramidal cell layer. Furthermore, Arc mRNAs levels were increased in both dorsal and VH, accompanied by an increase in Arc immunoreactivity in dendritic hippocampal layers. Furthermore, stress triggered a reduction in PSD-95 and NR1 protein levels in whole extract of dorsal and VH. Moreover, a reduction in NR2A/NR2B ratio was observed only in dorsal pole. In synaptosomal fractions, we detected a rise in NR1 in dorsal hippocampus (DH). By indirect immunofluorescence we found that NR1 subunits rise, especially in neuropil areas of dorsal, but not VH. In relation to AMPA receptor (AMPAR) subunits, chronic stress did not trigger any change, either in dorsal or ventral hippocampal areas. These data suggest that DH is more sensitive than VH to chronic stress exposure, mainly altering the expression of NMDA receptor (NMDAR) subunits, and probably favors changes in the configuration of this receptor that may influence the function of this area.

SIRT1 Functions as a Negative Regulator of Eukaryotic Poly(A)RNA Transport.

  • Shan P
  • Curr. Biol.
  • 2017 Aug 7

Literature context:


Abstract:

Most eukaryotic mRNAs are polyadenylated in the nucleus, and the poly(A)-tail is required for efficient mRNA export and translation. However, mechanisms governing mRNA transport remain unclear. Here, we report that the nicotinamide adenine dinucleotide (NAD)-dependent deacetylase SIRT1 acts as an energy sensor and negatively regulates poly(A)RNA transport via deacetylating a poly(A)-binding protein, PABP1. Upon energy starvation, SIRT1 interacts with and deacetylates PABP1 and deactivates its poly(A)RNA binding, leading to nuclear accumulation of PABP1 and poly(A)RNA and thus facilitating eukaryotic cells to attenuate protein synthesis and energy consumption to adapt to energy stress. Moreover, AMPK-directed SIRT1 phosphorylation is required for energy starvation-induced PABP1-SIRT1 association, PABP1 deacetylation, and poly(A)RNA nuclear retention. In addition, the SIRT1-PABP1 association is not specific to energy starvation but represents a common stress response. These observations provide insights into dynamic modulation of eukaryotic mRNA transport and translation, suggesting that the poly(A)-tail also provides a basis for eukaryotes to effectively shut down mature mRNA transport and thereby tailor protein synthesis to maintain energy homeostasis under stress conditions.

Molecular Memory of Morphologies by Septins during Neuron Generation Allows Early Polarity Inheritance.

  • Boubakar L
  • Neuron
  • 2017 Aug 16

Literature context:


Abstract:

Transmission of polarity established early during cell lineage history is emerging as a key process guiding cell differentiation. Highly polarized neurons provide a fascinating model to study inheritance of polarity over cell generations and across morphological transitions. Neural crest cells (NCCs) migrate to the dorsal root ganglia to generate neurons directly or after cell divisions in situ. Using live imaging of vertebrate embryo slices, we found that bipolar NCC progenitors lose their polarity, retracting their processes to round for division, but generate neurons with bipolar morphology by emitting processes from the same locations as the progenitor. Monitoring the dynamics of Septins, which play key roles in yeast polarity, indicates that Septin 7 tags process sites for re-initiation of process growth following mitosis. Interfering with Septins blocks this mechanism. Thus, Septins store polarity features during mitotic rounding so that daughters can reconstitute the initial progenitor polarity.

Cdc25A Is a Critical Mediator of Ischemic Neuronal Death In Vitro and In Vivo.

  • Iyirhiaro GO
  • J. Neurosci.
  • 2017 Jul 12

Literature context:


Abstract:

Dysregulation of cell cycle machinery is implicated in a number of neuronal death contexts, including stroke. Increasing evidence suggests that cyclin-dependent kinases (Cdks) are inappropriately activated in mature neurons under ischemic stress conditions. We previously demonstrated a functional role for the cyclin D1/Cdk4/pRb (retinoblastoma tumor suppressor protein) pathway in delayed neuronal death induced by ischemia. However, the molecular signals leading to cyclin D/Cdk4/pRb activation following ischemic insult are presently not clear. Here, we investigate the cell division cycle 25 (Cdc25) dual-specificity phosphatases as potential upstream regulators of ischemic neuronal death and Cdk4 activation. We show that a pharmacologic inhibitor of Cdc25 family members (A, B, and C) protects mouse primary neurons from hypoxia-induced delayed death. The major contributor to the death process appears to be Cdc25A. shRNA-mediated knockdown of Cdc25A protects neurons in a delayed model of hypoxia-induced death in vitro Similar results were observed in vivo following global ischemia in the rat. In contrast, neurons singly or doubly deficient for Cdc25B/C were not significantly protective. We show that Cdc25A activity, but not level, is upregulated in vitro following hypoxia and global ischemic insult in vivo Finally, we show that shRNA targeting Cdc25A blocks Ser795 pRb phosphorylation. Overall, our results indicate a role for Cdc25A in delayed neuronal death mediated by ischemia.SIGNIFICANCE STATEMENT A major challenge in stroke is finding an effective neuroprotective strategy to treat cerebral ischemic injury. Cdc25 family member A (Cdc25A) is a phosphatase normally activated during cell division in proliferating cells. We found that Cdc25A is activated in neurons undergoing ischemic stress mediated by hypoxia in vitro and global cerebral ischemia in rats in vivo We show that pharmacologic or genetic inhibition of Cdc25A activity protects neurons from delayed death in vitro and in vivo Downregulation of Cdc25A led to reduction in retinoblastoma tumor suppressor protein (pRb) phosphorylation. An increase in pRb phosphorylation has been previously linked to ischemic neuronal death. Our results identify Cdc25A as a potential target for neuroprotectant strategy for the treatment of delayed ischemic neuronal death.

Oncogenic Activation of the RNA Binding Protein NELFE and MYC Signaling in Hepatocellular Carcinoma.

  • Dang H
  • Cancer Cell
  • 2017 Jul 10

Literature context:


Abstract:

Global transcriptomic imbalance is a ubiquitous feature associated with cancer, including hepatocellular carcinoma (HCC). Analyses of 1,225 clinical HCC samples revealed that a large numbers of RNA binding proteins (RBPs) are dysregulated and that RBP dysregulation is associated with poor prognosis. We further identified that oncogenic activation of a top candidate RBP, negative elongation factor E (NELFE), via somatic copy-number alterations enhanced MYC signaling and promoted HCC progression. Interestingly, NELFE induces a unique tumor transcriptome by selectively regulating MYC-associated genes. Thus, our results revealed NELFE as an oncogenic protein that may contribute to transcriptome imbalance in HCC through the regulation of MYC signaling.

Funding information:
  • Intramural NIH HHS - Z01 BC010876-01()
  • Intramural NIH HHS - ZIA BC010877-09()
  • NCI NIH HHS - Z01 BC010876()
  • NCI NIH HHS - Z01 BC010877()
  • NHLBI NIH HHS - P01 HL131471()

Exophilin-8 assembles secretory granules for exocytosis in the actin cortex via interaction with RIM-BP2 and myosin-VIIa.

  • Fan F
  • Elife
  • 2017 Jul 4

Literature context:


Abstract:

Exophilin-8 has been reported to play a role in anchoring secretory granules within the actin cortex, due to its direct binding activities to Rab27 on the granule membrane and to F-actin and its motor protein, myosin-Va. Here, we show that exophilin-8 accumulates granules in the cortical F-actin network not by direct interaction with myosin-Va, but by indirect interaction with a specific form of myosin-VIIa through its previously unknown binding partner, RIM-BP2. RIM-BP2 also associates with exocytic machinery, Cav1.3, RIM, and Munc13-1. Disruption of the exophilin-8-RIM-BP2-myosin-VIIa complex by ablation or knockdown of each component markedly decreases both the peripheral accumulation and exocytosis of granules. Furthermore, exophilin-8-null mouse pancreatic islets lose polarized granule localization at the β-cell periphery and exhibit impaired insulin secretion. This newly identified complex acts as a physical and functional scaffold and provides a mechanism supporting a releasable pool of granules within the F-actin network beneath the plasma membrane.

Purinergic receptor P2Y6 contributes to 1-methyl-4-phenylpyridinium-induced oxidative stress and cell death in neuronal SH-SY5Y cells.

  • Qian Y
  • J. Neurosci. Res.
  • 2017 Jul 29

Literature context:


Abstract:

Oxidative stress and neural degeneration have been shown to be involved in the pathogenesis of Parkinson's disease (PD). The P2Y6 purinergic receptor (P2Y6R) has been shown to participate in the activation of microglia and the production of pro-inflammatory factors induced by lipopolysaccharide to cause neuronal loss. However, the function of P2Y6R during oxidative stress in neurons is unclear. In the present study, 1-methyl-4-phenylpyridinium (MPP+ ) treatment increased the level of UDP/P2Y6R on neuronal SH-SY5Y cells. Importantly, pharmacological inhibition of P2Y6R or knockdown of P2Y6R using a siRNA exerted an increased protective effect by preventing MPP+ -induced increases in the levels of reactive oxygen species (ROS), superoxide anion, inducible nitric oxide synthase (iNOS), and malondialdehyde (MDA) and down-regulation of superoxide dismutase 1 (SOD1) expression. UDP, an agonist of P2Y6R, enhanced the effects of MPP+ , which was also inhibited by apyrase or MRS2578. Additionally, P2Y6R knockdown also significantly reversed both the loss of cell viability and the increase in the levels of phosphorylated extracellular signal-regulated protein kinase (p-ERK1/2) and p38 (p-p38) caused by MPP+ stimulation. However, the inhibition of the ERK1/2 and p38 kinase signaling pathways had no effect on P2Y6R expression. Taken together, these results support the hypothesis that P2Y6R expressed on neuronal SH-SY5Y cell is associated with the progression of oxidative stress and cell death induced by MPP+ , suggesting that P2Y6R may play an important role in the pathogenesis of PD.

Hypothalamic AMPK-ER Stress-JNK1 Axis Mediates the Central Actions of Thyroid Hormones on Energy Balance.

  • Martínez-Sánchez N
  • Cell Metab.
  • 2017 Jul 5

Literature context:


Abstract:

Thyroid hormones (THs) act in the brain to modulate energy balance. We show that central triiodothyronine (T3) regulates de novo lipogenesis in liver and lipid oxidation in brown adipose tissue (BAT) through the parasympathetic (PSNS) and sympathetic nervous system (SNS), respectively. Central T3 promotes hepatic lipogenesis with parallel stimulation of the thermogenic program in BAT. The action of T3 depends on AMP-activated protein kinase (AMPK)-induced regulation of two signaling pathways in the ventromedial nucleus of the hypothalamus (VMH): decreased ceramide-induced endoplasmic reticulum (ER) stress, which promotes BAT thermogenesis, and increased c-Jun N-terminal kinase (JNK) activation, which controls hepatic lipid metabolism. Of note, ablation of AMPKα1 in steroidogenic factor 1 (SF1) neurons of the VMH fully recapitulated the effect of central T3, pointing to this population in mediating the effect of central THs on metabolism. Overall, these findings uncover the underlying pathways through which central T3 modulates peripheral metabolism.

Funding information:
  • NHLBI NIH HHS - R01 DK107220()
  • NIDDK NIH HHS - R01 DK112698()
  • Wellcome Trust - P01 HL084207()

Physiological and pathophysiological control of synaptic GluN2B-NMDA receptors by the C-terminal domain of amyloid precursor protein.

  • Pousinha PA
  • Elife
  • 2017 Jul 6

Literature context:


Abstract:

The amyloid precursor protein (APP) harbors physiological roles at synapses and is central to Alzheimer's disease (AD) pathogenesis. Evidence suggests that APP intracellular domain (AICD) could regulate synapse function, but the underlying molecular mechanisms remain unknown. We addressed AICD actions at synapses, per se, combining in vivo AICD expression, ex vivo AICD delivery or APP knock-down by in utero electroporation of shRNAs with whole-cell electrophysiology. We report a critical physiological role of AICD in controlling GluN2B-containing NMDA receptors (NMDARs) at immature excitatory synapses, via a transcription-dependent mechanism. We further show that AICD increase in mature neurons, as reported in AD, alters synaptic NMDAR composition to an immature-like GluN2B-rich profile. This disrupts synaptic signal integration, via over-activation of SK channels, and synapse plasticity, phenotypes rescued by GluN2B antagonism. We provide a new physiological role for AICD, which becomes pathological upon AICD increase in mature neurons. Thus, AICD could contribute to AD synaptic failure.

PTEN controls glandular morphogenesis through a juxtamembrane β-Arrestin1/ARHGAP21 scaffolding complex.

  • Javadi A
  • Elife
  • 2017 Jul 27

Literature context:


Abstract:

PTEN controls three-dimensional (3D) glandular morphogenesis by coupling juxtamembrane signaling to mitotic spindle machinery. While molecular mechanisms remain unclear, PTEN interacts through its C2 membrane-binding domain with the scaffold protein β-Arrestin1. Because β-Arrestin1 binds and suppresses the Cdc42 GTPase-activating protein ARHGAP21, we hypothesize that PTEN controls Cdc42 -dependent morphogenic processes through a β-Arrestin1-ARHGAP21 complex. Here, we show that PTEN knockdown (KD) impairs β-Arrestin1 membrane localization, β-Arrestin1-ARHGAP21 interactions, Cdc42 activation, mitotic spindle orientation and 3D glandular morphogenesis. Effects of PTEN deficiency were phenocopied by β-Arrestin1 KD or inhibition of β-Arrestin1-ARHGAP21 interactions. Conversely, silencing of ARHGAP21 enhanced Cdc42 activation and rescued aberrant morphogenic processes of PTEN-deficient cultures. Expression of the PTEN C2 domain mimicked effects of full-length PTEN but a membrane-binding defective mutant of the C2 domain abrogated these properties. Our results show that PTEN controls multicellular assembly through a membrane-associated regulatory protein complex composed of β-Arrestin1, ARHGAP21 and Cdc42.

The DYT6 Dystonia Protein THAP1 Regulates Myelination within the Oligodendrocyte Lineage.

  • Yellajoshyula D
  • Dev. Cell
  • 2017 Jul 10

Literature context:


Abstract:

The childhood-onset motor disorder DYT6 dystonia is caused by loss-of-function mutations in the transcription factor THAP1, but the neurodevelopmental processes in which THAP1 participates are unknown. We find that THAP1 is essential for the timing of myelination initiation during CNS maturation. Conditional deletion of THAP1 in the CNS retards maturation of the oligodendrocyte (OL) lineage, delaying myelination and causing persistent motor deficits. The CNS myelination defect results from a cell-autonomous requirement for THAP1 in the OL lineage and is recapitulated in developmental assays performed on OL progenitor cells purified from Thap1 null mice. Loss of THAP1 function disrupts a core set of OL maturation genes and reduces the DNA occupancy of YY1, a transcription factor required for OL maturation. These studies establish a role for THAP1 transcriptional regulation at the inception of myelination and implicate abnormal timing of myelination in the pathogenesis of childhood-onset dystonia.

Ywhaz/14-3-3ζ Deletion Improves Glucose Tolerance Through a GLP-1-Dependent Mechanism.

  • Lim GE
  • Endocrinology
  • 2017 Jun 5

Literature context:


Abstract:

Multiple signaling pathways mediate the actions of metabolic hormones to control glucose homeostasis, but the proteins that coordinate such networks are poorly understood. We previously identified the molecular scaffold protein, 14-3-3ζ, as a critical regulator of in vitro β-cell survival and adipogenesis, but its metabolic roles in glucose homeostasis have not been studied in depth. Herein, we report that Ywhaz gene knockout mice (14-3-3ζKO) exhibited elevated fasting insulin levels while maintaining normal β-cell responsiveness to glucose when compared with wild-type littermate controls. In contrast with our observations after an ip glucose bolus, glucose tolerance was significantly improved in 14-3-3ζKO mice after an oral glucose gavage. This improvement in glucose tolerance was associated with significantly elevated fasting glucagon-like peptide-1 (GLP-1) levels. 14-3-3ζ knockdown in GLUTag L cells elevated GLP-1 synthesis and increased GLP-1 release. Systemic inhibition of the GLP-1 receptor attenuated the improvement in oral glucose tolerance that was seen in 14-3-3ζKO mice. When taken together these findings demonstrate novel roles of 14-3-3ζ in the regulation of glucose homeostasis and suggest that modulating 14-3-3ζ levels in intestinal L cells may have beneficial metabolic effects through GLP-1-dependent mechanisms.

Funding information:
  • NIAID NIH HHS - R01 AI118985(United States)
  • NIDDK NIH HHS - DK43140(United States)

Pharmacological enhancement of mGlu5 receptors rescues behavioral deficits in SHANK3 knock-out mice.

  • Vicidomini C
  • Mol. Psychiatry
  • 2017 Jun 29

Literature context:


Abstract:

SHANK3 (also called PROSAP2) genetic haploinsufficiency is thought to be the major cause of neuropsychiatric symptoms in Phelan-McDermid syndrome (PMS). PMS is a rare genetic disorder that causes a severe form of intellectual disability (ID), expressive language delays and other autistic features. Furthermore, a significant number of SHANK3 mutations have been identified in patients with autism spectrum disorders (ASD), and SHANK3 truncating mutations are associated with moderate to profound ID. The Shank3 protein is a scaffold protein that is located in the postsynaptic density (PSD) of excitatory synapses and is crucial for synapse development and plasticity. In this study, we investigated the molecular mechanisms associated with the ASD-like behaviors observed in Shank3Δ11-/- mice, in which exon 11 has been deleted. Our results indicate that Shank3 is essential to mediating metabotropic glutamate receptor 5 (mGlu5)-receptor signaling by recruiting Homer1b/c to the PSD, specifically in the striatum and cortex. Moreover, augmenting mGlu5-receptor activity by administering 3-Cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide ameliorated the functional and behavioral defects that were observed in Shank3Δ11-/- mice, suggesting that pharmaceutical treatments that increase mGlu5 activity may represent a new approach for treating patients that are affected by PMS and SHANK3 mutations.

Funding information:
  • Wellcome Trust - WT098418MA(United Kingdom)

3-Iodothyronamine Induces Tail Vasodilation Through Central Action in Male Mice.

  • Gachkar S
  • Endocrinology
  • 2017 Jun 1

Literature context:


Abstract:

3-Iodothyronamine (3-T1AM) is an endogenous thyroid hormone (TH)-derived metabolite that induces severe hypothermia in mice after systemic administration; however, the underlying mechanisms have remained enigmatic. We show here that the rapid 3-T1AM-induced loss in body temperature is a consequence of peripheral vasodilation and subsequent heat loss (e.g., over the tail surface). The condition is subsequently intensified by hypomotility and a lack of brown adipose tissue activation. Although the possible 3-T1AM targets trace amine-associated receptor 1 or α2a-adrenergic receptor were detected in tail artery and aorta respectively, myograph studies did not show any direct effect of 3-T1AM on vasodilation, suggesting that its actions are likely indirect. Intracerebroventricular application of 3-T1AM, however, replicated the phenotype of tail vasodilation and body temperature decline and led to neuronal activation in the hypothalamus, suggesting that the metabolite causes tail vasodilation through a hypothalamic signaling pathway. Consequently, the 3-T1AM response constitutes anapyrexia rather than hypothermia and closely resembles the heat-stress response mediated by hypothalamic temperature-sensitive neurons. Our results thus underline the well-known role of the hypothalamus as the body's thermostat and suggest an additional molecular link between TH signaling and the central control of body temperature.

De Novo-Synthesized Retinoic Acid in Ovarian Antral Follicles Enhances FSH-Mediated Ovarian Follicular Cell Differentiation and Female Fertility.

  • Kawai T
  • Endocrinology
  • 2017 Jun 5

Literature context:


Abstract:

Retinoic acid (RA) is the active form of vitamin A and is synthesized from retinol by two key enzymes, alcohol dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH). As the physiological precursor of RA, retinol impacts female reproductive functions and fertility. The expression of Adh1 and Adh5 as well as Aldh1a1 and Aldh1a7 are significantly increased in the ovaries of mice treated with equine chorionic gonadotropin/FSH. The RA receptor is expressed and localized in granulosa cells and is activated by endogenous RA as indicated by LacZ expression in granulosa cells of RA-responsive transgene-LacZ transgenic mice (RA reporter mice). Coinjection of the ADH inhibitor, 4-methylpyrazole, with equine chorionic gonadotropin significantly decreases the number and developmental competence of oocytes ovulated in response to human chorionic gonadotropin/LH as compared with controls. Injections of RA completely reverse the effects of the inhibitor of ovulation and oocyte development. When mice were fed a retinol-free, vitamin A-deficient diet that significantly reduced the serum levels of retinol, the expression of the LH receptor (Lhcgr) was significantly lower in the ovaries of the vitamin A-deficient mice, and injections of human chorionic gonadotropin failed to induce genes controlling ovulation. These results indicate that ovarian de novo biosynthesis of RA is required for the follicular expression of Lhcgr in granulosa cells and their ability to respond to the ovulatory LH surge.

Funding information:
  • NIAMS NIH HHS - R01 AR061569(United States)

Mid-life environmental enrichment increases synaptic density in CA1 in a mouse model of Aβ-associated pathology and positively influences synaptic and cognitive health in healthy ageing.

  • Stuart KE
  • J. Comp. Neurol.
  • 2017 Jun 1

Literature context:


Abstract:

Early-life cognitive enrichment may reduce the risk of experiencing cognitive deterioration and dementia in later-life. However, an intervention to prevent or delay dementia is likely to be taken up in mid to later-life. Hence, we investigated the effects of environmental enrichment in wildtype mice and in a mouse model of Aβ neuropathology (APPSWE /PS1dE9 ) from 6 months of age. After 6 months of housing in standard laboratory cages, APPSWE /PS1dE9 (n = 27) and healthy wildtype (n = 21) mice were randomly assigned to either enriched or standard housing. At 12 months of age, wildtype mice showed altered synaptic protein levels and relatively superior cognitive performance afforded by environmental enrichment. Environmental enrichment was not associated with alterations to Aβ plaque pathology in the neocortex or hippocampus of APPSWE /PS1dE9 mice. However, a significant increase in synaptophysin immunolabeled puncta in the hippocampal subregion, CA1, in APPSWE /PS1dE9 mice was detected, with no significant synaptic density changes observed in CA3, or the Fr2 region of the prefrontal cortex. Moreover, a significant increase in hippocampal BDNF was detected in APPSWE /PS1dE9 mice exposed to EE, however, no changes were detected in neocortex or between Wt animals. These results demonstrate that mid to later-life cognitive enrichment has the potential to promote synaptic and cognitive health in ageing, and to enhance compensatory capacity for synaptic connectivity in pathological ageing associated with Aβ deposition.

Differential DARC/ACKR1 expression distinguishes venular from non-venular endothelial cells in murine tissues.

  • Thiriot A
  • BMC Biol.
  • 2017 May 19

Literature context:


Abstract:

BACKGROUND: Intravascular leukocyte recruitment in most vertebrate tissues is restricted to postcapillary and collecting venules, whereas capillaries and arterioles usually support little or no leukocyte adhesion. This segmental restriction is thought to be mediated by endothelial, rather than hemodynamic, differences. The underlying mechanisms are largely unknown, in part because effective tools to distinguish, isolate, and analyze venular endothelial cells (V-ECs) and non-venular endothelial cells (NV-ECs) have been unavailable. We hypothesized that the atypical chemokine receptor DARC (Duffy Antigen Receptor for Chemokines, a.k.a. ACKR1 or CD234) may distinguish V-ECs versus NV-ECs in mice. METHODS: We generated a rat-anti-mouse monoclonal antibody (MAb) that specifically recognizes the erythroid and endothelial forms of native, surface-expressed DARC. Using this reagent, we characterized DARC expression and distribution in the microvasculature of murine tissues. RESULTS: DARC was exquisitely restricted to post-capillary and small collecting venules and completely absent from arteries, arterioles, capillaries, veins, and most lymphatics in every tissue analyzed. Accordingly, intravital microscopy showed that adhesive leukocyte-endothelial interactions were restricted to DARC+ venules. DARC was detectable over the entire circumference of V-ECs, but was more concentrated at cell-cell junctions. Analysis of single-cell suspensions suggested that the frequency of V-ECs among the total microvascular EC pool varies considerably between different tissues. CONCLUSIONS: Immunostaining of endothelial DARC allows the identification and isolation of intact V-ECs from multiple murine tissues. This strategy may be useful to dissect the mechanisms underlying segmental microvascular specialization in healthy and diseased tissues and to characterize the role of EC subsets in tissue-homeostasis, immune surveillance, infection, inflammation, and malignancies.

Funding information:
  • NIAID NIH HHS - P01 AI112521()
  • NIAID NIH HHS - R01 AI111595()

CaMKII Autophosphorylation Is Necessary for Optimal Integration of Ca2+ Signals during LTP Induction, but Not Maintenance.

  • Chang JY
  • Neuron
  • 2017 May 17

Literature context:


Abstract:

CaMKII plays a critical role in decoding calcium (Ca2+) signals to initiate long-lasting synaptic plasticity. However, the properties of CaMKII that mediate Ca2+ signals in spines remain elusive. Here, we measured CaMKII activity in spines using fast-framing two-photon fluorescence lifetime imaging. Following each pulse during repetitive Ca2+ elevations, CaMKII activity increased in a stepwise manner. Thr286 phosphorylation slows the decay of CaMKII and thus lowers the frequency required to induce spine plasticity by several fold. In the absence of Thr286 phosphorylation, increasing the stimulation frequency results in high peak mutant CaMKIIT286A activity that is sufficient for inducing plasticity. Our findings demonstrate that Thr286 phosphorylation plays an important role in induction of LTP by integrating Ca2+ signals, and it greatly promotes, but is dispensable for, the activation of CaMKII and LTP.

Funding information:
  • NIMH NIH HHS - R01 MH080047()
  • NINDS NIH HHS - DP1 NS096787()

FGF21 Regulates Metabolism Through Adipose-Dependent and -Independent Mechanisms.

  • BonDurant LD
  • Cell Metab.
  • 2017 Apr 4

Literature context:


Abstract:

FGF21 is an endocrine hormone that regulates energy homeostasis and insulin sensitivity. The mechanism of FGF21 action and the tissues responsible for these effects have been controversial, with both adipose tissues and the central nervous system having been identified as the target site mediating FGF21-dependent increases in insulin sensitivity, energy expenditure, and weight loss. Here we show that, while FGF21 signaling to adipose tissue is required for the acute insulin-sensitizing effects of FGF21, FGF21 signaling to adipose tissue is not required for its chronic effects to increase energy expenditure and lower body weight. Also, in contrast to previous studies, we found that adiponectin is dispensable for the metabolic effects of FGF21 in increasing insulin sensitivity and energy expenditure. Instead, FGF21 acutely enhances insulin sensitivity through actions on brown adipose tissue. Our data reveal that the acute and chronic effects of FGF21 can be dissociated through adipose-dependent and -independent mechanisms.

Funding information:
  • NHLBI NIH HHS - R01 HL111190()
  • NIDDK NIH HHS - K01 DK111758()
  • NIDDK NIH HHS - R01 DK106104()
  • NIGMS NIH HHS - T32 GM067795()

The Pseudomonas aeruginosa lectin LecA triggers host cell signalling by glycosphingolipid-dependent phosphorylation of the adaptor protein CrkII.

  • Zheng S
  • Biochim. Biophys. Acta
  • 2017 Apr 21

Literature context:


Abstract:

The human pathogen Pseudomonas aeruginosa induces phosphorylation of the adaptor protein CrkII by activating the non-receptor tyrosine kinase Abl to promote its uptake into host cells. So far, specific factors of P. aeruginosa, which induce Abl/CrkII signalling, are entirely unknown. In this research, we employed human lung epithelial cells H1299, Chinese hamster ovary cells and P. aeruginosa wild type strain PAO1 to study the invasion process of P. aeruginosa into host cells by using microbiological, biochemical and cell biological approaches such as Western Blot, immunofluorescence microscopy and flow cytometry. Here, we demonstrate that the host glycosphingolipid globotriaosylceramide, also termed Gb3, represents a signalling receptor for the P. aeruginosa lectin LecA to induce CrkII phosphorylation at tyrosine 221. Alterations in Gb3 expression and LecA function correlate with CrkII phosphorylation. Interestingly, phosphorylation of CrkIIY221 occurs independently of Abl kinase. We further show that Src family kinases transduce the signal induced by LecA binding to Gb3, leading to CrkY221 phosphorylation. In summary, we identified LecA as a bacterial factor, which utilizes a so far unrecognized mechanism for phospho-CrkIIY221 induction by binding to the host glycosphingolipid receptor Gb3. The LecA/Gb3 interaction highlights the potential of glycolipids to mediate signalling processes across the plasma membrane and should be further elucidated to gain deeper insights into this non-canonical mechanism of activating host cell processes.

Human biallelic MFN2 mutations induce mitochondrial dysfunction, upper body adipose hyperplasia, and suppression of leptin expression.

  • Rocha N
  • Elife
  • 2017 Apr 19

Literature context:


Abstract:

MFN2 encodes mitofusin 2, a membrane-bound mediator of mitochondrial membrane fusion and inter-organelle communication. MFN2 mutations cause axonal neuropathy, with associated lipodystrophy only occasionally noted, however homozygosity for the p.Arg707Trp mutation was recently associated with upper body adipose overgrowth. We describe similar massive adipose overgrowth with suppressed leptin expression in four further patients with biallelic MFN2 mutations and at least one p.Arg707Trp allele. Overgrown tissue was composed of normal-sized, UCP1-negative unilocular adipocytes, with mitochondrial network fragmentation, disorganised cristae, and increased autophagosomes. There was strong transcriptional evidence of mitochondrial stress signalling, increased protein synthesis, and suppression of signatures of cell death in affected tissue, whereas mitochondrial morphology and gene expression were normal in skin fibroblasts. These findings suggest that specific MFN2 mutations cause tissue-selective mitochondrial dysfunction with increased adipocyte proliferation and survival, confirm a novel form of excess adiposity with paradoxical suppression of leptin expression, and suggest potential targeted therapies.

Funding information:
  • Wellcome Trust - P30 DK072488()

Cellular distribution of the fragile X mental retardation protein in the mouse brain.

  • Zorio DA
  • J. Comp. Neurol.
  • 2017 Mar 1

Literature context:


Abstract:

The fragile X mental retardation protein (FMRP) plays an important role in normal brain development. Absence of FMRP results in abnormal neuronal morphologies in a selected manner throughout the brain, leading to intellectual deficits and sensory dysfunction in the fragile X syndrome (FXS). Despite FMRP importance for proper brain function, its overall expression pattern in the mammalian brain at the resolution of individual neuronal cell groups is not known. In this study we used FMR1 knockout and isogenic wildtype mice to systematically map the distribution of FMRP expression in the entire mouse brain. Using immunocytochemistry and cellular quantification analyses, we identified a large number of prominent cell groups expressing high levels of FMRP at the subcortical levels, in particular sensory and motor neurons in the brainstem and thalamus. In contrast, many cell groups in the midbrain and hypothalamus exhibit low FMRP levels. More important, we describe differential patterns of FMRP distribution in both cortical and subcortical brain regions. Almost all major brain areas contain high and low levels of FMRP cell groups adjacent to each other or between layers of the same cortical areas. These differential patterns indicate that FMRP expression appears to be specific to individual neuronal cell groups instead of being associated with all neurons in distinct brain regions, as previously considered. Taken together, these findings support the notion of FMRP differential neuronal regulation and strongly implicate the contribution of fundamental sensory and motor processing at subcortical levels to FXS pathology. J. Comp. Neurol. 525:818-849, 2017. © 2016 Wiley Periodicals, Inc.

Funding information:
  • Howard Hughes Medical Institute - (United States)
  • NCRR NIH HHS - P40 RR012546(United States)

The Putative Drp1 Inhibitor mdivi-1 Is a Reversible Mitochondrial Complex I Inhibitor that Modulates Reactive Oxygen Species.

  • Bordt EA
  • Dev. Cell
  • 2017 Mar 27

Literature context:


Abstract:

Mitochondrial fission mediated by the GTPase dynamin-related protein 1 (Drp1) is an attractive drug target in numerous maladies that range from heart disease to neurodegenerative disorders. The compound mdivi-1 is widely reported to inhibit Drp1-dependent fission, elongate mitochondria, and mitigate brain injury. Here, we show that mdivi-1 reversibly inhibits mitochondrial complex I-dependent O2 consumption and reverse electron transfer-mediated reactive oxygen species (ROS) production at concentrations (e.g., 50 μM) used to target mitochondrial fission. Respiratory inhibition is rescued by bypassing complex I using yeast NADH dehydrogenase Ndi1. Unexpectedly, respiratory impairment by mdivi-1 occurs without mitochondrial elongation, is not mimicked by Drp1 deletion, and is observed in Drp1-deficient fibroblasts. In addition, mdivi-1 poorly inhibits recombinant Drp1 GTPase activity (Ki > 1.2 mM). Overall, these results suggest that mdivi-1 is not a specific Drp1 inhibitor. The ability of mdivi-1 to reversibly inhibit complex I and modify mitochondrial ROS production may contribute to effects observed in disease models.

Funding information:
  • NIGMS NIH HHS - R01 GM067180()
  • NIGMS NIH HHS - R01 GM089853()
  • NINDS NIH HHS - R01 NS064978()
  • NINDS NIH HHS - R01 NS085165()
  • NINDS NIH HHS - R21 NS096538()

Endosulfine-alpha inhibits membrane-induced α-synuclein aggregation and protects against α-synuclein neurotoxicity.

  • Ysselstein D
  • Acta Neuropathol Commun
  • 2017 Jan 10

Literature context:


Abstract:

Neuropathological and genetic findings suggest that the presynaptic protein α-synuclein (aSyn) is involved in the pathogenesis of synucleinopathy disorders, including Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy. Evidence suggests that the self-assembly of aSyn conformers bound to phospholipid membranes in an aggregation-prone state plays a key role in aSyn neurotoxicity. Accordingly, we hypothesized that protein binding partners of lipid-associated aSyn could inhibit the formation of toxic aSyn oligomers at membrane surfaces. To address this hypothesis, we characterized the protein endosulfine-alpha (ENSA), previously shown to interact selectively with membrane-bound aSyn, in terms of its effects on the membrane-induced aggregation and neurotoxicity of two familial aSyn mutants, A30P and G51D. We found that wild-type ENSA, but not the non-aSyn-binding S109E variant, interfered with membrane-induced aSyn self-assembly, aSyn-mediated vesicle disruption and aSyn neurotoxicity. Immunoblotting analyses revealed that ENSA was down-regulated in the brains of synucleinopathy patients versus non-diseased individuals. Collectively, these results suggest that ENSA can alleviate neurotoxic effects of membrane-bound aSyn via an apparent chaperone-like activity at the membrane surface, and a decrease in ENSA expression may contribute to aSyn neuropathology in synucleinopathy disorders. More generally, our findings suggest that promoting interactions between lipid-bound, amyloidogenic proteins and their binding partners is a viable strategy to alleviate cytotoxicity in a range of protein misfolding disorders.

Funding information:
  • NIA NIH HHS - P50 AG005134()

Neuroprotective effect of bee venom is mediated by reduced astrocyte activation in a subchronic MPTP-induced model of Parkinson's disease.

  • Kim ME
  • Arch. Pharm. Res.
  • 2016 Aug 30

Literature context:


Abstract:

Bee venom (BV), also known as apitoxin, is widely used in traditional oriental medicine to treat immune-related diseases. Recent studies suggest that BV could be beneficial for the treatment of neurodegenerative diseases. Parkinson's disease (PD) is the second most common neurodegenerative disease next to Alzheimer's disease, and PD pathologies are closely associated with neuroinflammation. Previous studies have suggested the neuroprotective effects of BV in animal models of PD are due to the modulation of inflammation. However, the molecular mechanisms responsible for the anti-neuroinflammatory effect of BV have not been elucidated in astrocytes. Here, the authors investigated the neuroprotective effects of BV and pramipexole (PPX; a positive control) in a subchronic MPTP-induced murine PD model. Both BV and PPX prevented MPTP-induced impairments in motor performance and reduced dopaminergic neuron loss, and furthermore, these neuroprotective effects of BV and PPX were found to be associated with reduced astroglial activation in vivo PD model. However, in MPP(+) treated primary cultured astrocytes, BV modulated astrocyte activation, whereas PPX did not, indicating that the neuroprotective effects of PPX were not mediated by neuroinflammation. These findings suggest that BV should be considered a potential therapeutic or preventive agent for PD and other neuroinflammatory associated disorders.

Executive function deficits and glutamatergic protein alterations in a progressive 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mouse model of Parkinson's disease.

  • Pflibsen L
  • J. Neurosci. Res.
  • 2016 Jul 27

Literature context:


Abstract:

Changes in executive function are at the root of most cognitive problems associated with Parkinson's disease. Because dopaminergic treatment does not necessarily alleviate deficits in executive function, it has been hypothesized that dysfunction of neurotransmitters/systems other than dopamine (DA) may be associated with this decrease in cognitive function. We have reported decreases in motor function and dopaminergic/glutamatergic biomarkers in a progressive 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) Parkinson's mouse model. Assessment of executive function and dopaminergic/glutamatergic biomarkers within the limbic circuit has not previously been explored in our model. Our results show progressive behavioral decline in a cued response task (a rodent model for frontal cortex cognitive function) with increasing weekly doses of MPTP. Although within the dorsolateral (DL) striatum mice that had been given MPTP showed a 63% and 83% loss of tyrosine hydroxylase and dopamine transporter expression, respectively, there were no changes in the nucleus accumbens or medial prefrontal cortex (mPFC). Furthermore, dopamine-1 receptor and vesicular glutamate transporter (VGLUT)-1 expression increased in the mPFC following DA loss. There were significant MPTP-induced decreases and increases in VGLUT-1 and VGLUT-2 expression, respectively, within the DL striatum. We propose that the behavioral decline following MPTP treatment may be associated with a change not only in cortical-cortical (VGLUT-1) glutamate function but also in striatal DA and glutamate (VGLUT-1/VGLUT-2) input.

A Dietary Medium-Chain Fatty Acid, Decanoic Acid, Inhibits Recruitment of Nur77 to the HSD3B2 Promoter In Vitro and Reverses Endocrine and Metabolic Abnormalities in a Rat Model of Polycystic Ovary Syndrome.

  • Lee BH
  • Endocrinology
  • 2016 Jan 31

Literature context:


Abstract:

Hyperandrogenism is the central feature of polycystic ovary syndrome (PCOS). Due to the intricate relationship between hyperandrogenism and insulin resistance in PCOS, 50%-70% of these patients also present with hyperinsulinemia. Metformin, an insulin sensitizer, has been used to reduce insulin resistance and improve fertility in women with PCOS. In previous work, we have noted that a dietary medium-chain fatty acid, decanoic acid (DA), improves glucose tolerance and lipid profile in a mouse model of diabetes. Here, we report for the first time that DA, like metformin, inhibits androgen biosynthesis in NCI-H295R steroidogenic cells by regulating the enzyme 3β-hydroxysteroid dehydrogenase/Δ5-Δ4-isomerase type 2 (HSD3B2). The inhibitory effect on HSD3B2 and androgen production required cAMP stimulation, suggesting a mechanistic action via the cAMP-stimulated pathway. Specifically, both DA and metformin reduced cAMP-enhanced recruitment of the orphan nuclear receptor Nur77 to the HSD3B2 promoter, coupled with decreased transcription and protein expression of HSD3B2. In a letrozole-induced PCOS rat model, treatment with DA or metformin reduced serum-free testosterone, lowered fasting insulin, and restored estrous cyclicity. In addition, DA treatment lowered serum total testosterone and decreased HSD3B2 protein expression in the adrenals and ovaries. We conclude that DA inhibits androgen biosynthesis via mechanisms resulting in the suppression of HSD3B2 expression, an effect consistently observed both in vitro and in vivo. The efficacy of DA in reversing the endocrine and metabolic abnormalities of the letrozole-induced PCOS rat model are promising, raising the possibility that diets including DA could be beneficial for the management of both hyperandrogenism and insulin resistance in PCOS.

Funding information:
  • Intramural NIH HHS - (United States)
  • NIDCD NIH HHS - T32 DC009401(United States)

Association of N-cadherin levels and downstream effectors of Rho GTPases with dendritic spine loss induced by chronic stress in rat hippocampal neurons.

  • Castañeda P
  • J. Neurosci. Res.
  • 2015 Oct 20

Literature context:


Abstract:

Chronic stress promotes cognitive impairment and dendritic spine loss in hippocampal neurons. In this animal model of depression, spine loss probably involves a weakening of the interaction between pre- and postsynaptic cell adhesion molecules, such as N-cadherin, followed by disruption of the cytoskeleton. N-cadherin, in concert with catenin, stabilizes the cytoskeleton through Rho-family GTPases. Via their effector LIM kinase (LIMK), RhoA and ras-related C3 botulinum toxin substrate 1 (RAC) GTPases phosphorylate and inhibit cofilin, an actin-depolymerizing molecule, favoring spine growth. Additionally, RhoA, through Rho kinase (ROCK), inactivates myosin phosphatase through phosphorylation of the myosin-binding subunit (MYPT1), producing actomyosin contraction and probable spine loss. Some micro-RNAs negatively control the translation of specific mRNAs involved in Rho GTPase signaling. For example, miR-138 indirectly activates RhoA, and miR-134 reduces LIMK1 levels, resulting in spine shrinkage; in contrast, miR-132 activates RAC1, promoting spine formation. We evaluated whether N-cadherin/β-catenin and Rho signaling is sensitive to chronic restraint stress. Stressed rats exhibit anhedonia, impaired associative learning, and immobility in the forced swim test and reduction in N-cadherin levels but not β-catenin in the hippocampus. We observed a reduction in spine number in the apical dendrites of CA1 pyramidal neurons, with no effect on the levels of miR-132 or miR-134. Although the stress did not modify the RAC-LIMK-cofilin signaling pathway, we observed increased phospho-MYPT1 levels, probably mediated by RhoA-ROCK activation. Furthermore, chronic stress raises the levels of miR-138 in accordance with the observed activation of the RhoA-ROCK pathway. Our findings suggest that a dysregulation of RhoA-ROCK activity by chronic stress could potentially underlie spine loss in hippocampal neurons.

Pregnancy induces resistance to the anorectic effect of hypothalamic malonyl-CoA and the thermogenic effect of hypothalamic AMPK inhibition in female rats.

  • Martínez de Morentin PB
  • Endocrinology
  • 2015 Mar 21

Literature context:


Abstract:

During gestation, hyperphagia is necessary to cope with the metabolic demands of embryonic development. There were three main aims of this study: Firstly, to investigate the effect of pregnancy on hypothalamic fatty acid metabolism, a key pathway for the regulation of energy balance; secondly, to study whether pregnancy induces resistance to the anorectic effect of fatty acid synthase (FAS) inhibition and accumulation of malonyl-coenzyme A (CoA) in the hypothalamus; and, thirdly, to study whether changes in hypothalamic AMPK signaling are associated with brown adipose tissue (BAT) thermogenesis during pregnancy. Our data suggest that in pregnant rats, the hypothalamic fatty acid pathway shows an overall state that should lead to anorexia and elevated BAT thermogenesis: decreased activities of AMP-activated protein kinase (AMPK), FAS, and carnitine palmitoyltransferase 1, coupled with increased acetyl-CoA carboxylase function with subsequent elevation of malonyl-CoA levels. This profile seems dependent of estradiol levels but not prolactin or progesterone. Despite the apparent anorexic and thermogenic signaling in the hypothalamus, pregnant rats remain hyperphagic and display reduced temperature and BAT function. Actually, pregnant rats develop resistance to the anorectic effects of central FAS inhibition, which is associated with a reduction of proopiomelanocortin (POMC) expression and its transcription factors phospho-signal transducer and activator of transcription 3, and phospho-forkhead box O1. This evidence demonstrates that pregnancy induces a state of resistance to the anorectic and thermogenic actions of hypothalamic cellular signals of energy surplus, which, in parallel to the already known refractoriness to leptin effects, likely contributes to gestational hyperphagia and adiposity.

Funding information:
  • NIH HHS - P40 OD018537(United States)

GluA1 phosphorylation contributes to postsynaptic amplification of neuropathic pain in the insular cortex.

  • Qiu S
  • J. Neurosci.
  • 2014 Oct 1

Literature context:


Abstract:

Long-term potentiation of glutamatergic transmission has been observed after physiological learning or pathological injuries in different brain regions, including the spinal cord, hippocampus, amygdala, and cortices. The insular cortex is a key cortical region that plays important roles in aversive learning and neuropathic pain. However, little is known about whether excitatory transmission in the insular cortex undergoes plastic changes after peripheral nerve injury. Here, we found that peripheral nerve ligation triggered the enhancement of AMPA receptor (AMPAR)-mediated excitatory synaptic transmission in the insular cortex. The synaptic GluA1 subunit of AMPAR, but not the GluA2/3 subunit, was increased after nerve ligation. Genetic knock-in mice lacking phosphorylation of the Ser845 site, but not that of the Ser831 site, blocked the enhancement of the synaptic GluA1 subunit, indicating that GluA1 phosphorylation at the Ser845 site by protein kinase A (PKA) was critical for this upregulation after nerve injury. Furthermore, A-kinase anchoring protein 79/150 (AKAP79/150) and PKA were translocated to the synapses after nerve injury. Genetic deletion of adenylyl cyclase subtype 1 (AC1) prevented the translocation of AKAP79/150 and PKA, as well as the upregulation of synaptic GluA1-containing AMPARs. Pharmacological inhibition of calcium-permeable AMPAR function in the insular cortex reduced behavioral sensitization caused by nerve injury. Our results suggest that the expression of AMPARs is enhanced in the insular cortex after nerve injury by a pathway involving AC1, AKAP79/150, and PKA, and such enhancement may at least in part contribute to behavioral sensitization together with other cortical regions, such as the anterior cingulate and the prefrontal cortices.

Funding information:
  • NINDS NIH HHS - 1K99NS087086(United States)

Opposing effects of membrane-anchored CX3CL1 on amyloid and tau pathologies via the p38 MAPK pathway.

  • Lee S
  • J. Neurosci.
  • 2014 Sep 10

Literature context:


Abstract:

Several Alzheimer's disease (AD) risk genes are specifically expressed by microglia within the CNS. However, the mechanisms by which microglia regulate the pathological hallmarks of AD--extracellular deposition of β-amyloid (Aβ) and intraneuronal hyperphosphorylation of microtubule-associated protein tau (MAPT)--remain to be established. Notably, deficiency for the microglial CX3CR1 receptor has opposing effects on Aβ and MAPT pathologies. CX3CL1, the neuronally derived cognate ligand for CX3CR1, signals both in membrane-anchored and soluble forms. In this study, we sought to determine the relative contribution on membrane-anchored versus soluble CX3CL1 in regulating the microglia-mediated amelioration of Aβ pathology, as well as provide insight into the potential downstream microglial-based mechanisms. As expected, CX3CL1 deficiency reduced Aβ deposition in APPPS1 animals in a similar manner to CX3CR1 deficiency. Surprisingly, however, CX3CL1-deficient APPPS1 animals exhibited enhanced neuronal MAPT phosphorylation despite reduced amyloid burden. Importantly, neither of these phenotypes was altered by transgenic expression of the soluble CX3CL1 isoform, suggesting that it is the membrane-anchored version of CX3CL1 that regulates microglial phagocytosis of Aβ and neuronal MAPT phosphorylation. Analysis of transcript levels in purified microglia isolated from APPPS1 mice with the various CX3CL1/CX3CR1 genotypes revealed increased expression of inflammatory cytokines and phagocytic markers, which was associated with activation of p38 mitogen-activated protein kinase and Aβ internalization within microglia. Together, these studies challenge the "frustrated phagocytosis" concept and suggest that neuronal-microglial communication link the two central AD pathologies.

MicroRNA-7 protects against 1-methyl-4-phenylpyridinium-induced cell death by targeting RelA.

  • Choi DC
  • J. Neurosci.
  • 2014 Sep 17

Literature context:


Abstract:

Parkinson's disease (PD) is characterized by the progressive loss of dopaminergic neurons in the substantia nigra. Mitochondrial complex I impairment in PD is modeled in vitro by the susceptibility of dopaminergic neurons to the complex I inhibitor 1-methyl-4-phenylpyridinium (MPP+). In the present study, we demonstrate that microRNA-7 (miR-7), which is expressed in tyrosine hydroxylase-positive nigral neurons in mice and humans, protects cells from MPP+-induced toxicity in dopaminergic SH-SY5Y cells, differentiated human neural progenitor ReNcell VM cells, and primary mouse neurons. RelA, a component of nuclear factor-κB (NF-κB), was identified to be downregulated by miR-7 using quantitative proteomic analysis. Through a series of validation experiments, it was confirmed that RelA mRNA is a target of miR-7 and is required for cell death following MPP+ exposure. Further, RelA mediates MPP+-induced suppression of NF-κB activity, which is essential for MPP+-induced cell death. Accordingly, the protective effect of miR-7 is exerted through relieving NF-κB suppression by reducing RelA expression. These findings provide a novel mechanism by which NF-κB suppression, rather than activation, underlies the cell death mechanism following MPP+ toxicity, have implications for the pathogenesis of PD, and suggest miR-7 as a therapeutic target for this disease.

Funding information:
  • NCRR NIH HHS - RR-17072(United States)

Nicotine improves obesity and hepatic steatosis and ER stress in diet-induced obese male rats.

  • Seoane-Collazo P
  • Endocrinology
  • 2014 May 21

Literature context:


Abstract:

Nicotine, the main addictive component of tobacco, promotes body weight reduction in humans and rodents. Recent evidence has suggested that nicotine acts in the central nervous system to modulate energy balance. Specifically, nicotine modulates hypothalamic AMP-activated protein kinase to decrease feeding and to increase brown adipose tissue thermogenesis through the sympathetic nervous system, leading to weight loss. Of note, most of this evidence has been obtained in animal models fed with normal diet or low-fat diet (LFD). However, its effectiveness in obese models remains elusive. Because obesity causes resistance towards many factors involved in energy homeostasis, the aim of this study has been to compare the effect of nicotine in a diet-induced obese (DIO) model, namely rats fed a high-fat diet, with rats fed a LFD. Our data show that chronic peripheral nicotine treatment reduced body weight by decreasing food intake and increasing brown adipose tissue thermogenesis in both LFD and DIO rats. This overall negative energy balance was associated to decreased activation of hypothalamic AMP-activated protein kinase in both models. Furthermore, nicotine improved serum lipid profile, decreased insulin serum levels, as well as reduced steatosis, inflammation, and endoplasmic reticulum stress in the liver of DIO rats but not in LFD rats. Overall, this evidence suggests that nicotine diminishes body weight and improves metabolic disorders linked to DIO and might offer a clear-cut strategy to develop new therapeutic approaches against obesity and its metabolic complications.

Funding information:
  • Medical Research Council - G1100356(United Kingdom)
  • NINDS NIH HHS - R01 NS085093(United States)

Nesfatin-1 in human and murine cardiomyocytes: synthesis, secretion, and mobilization of GLUT-4.

  • Feijóo-Bandín S
  • Endocrinology
  • 2013 Dec 25

Literature context:


Abstract:

Nesfatin-1, a satiety-inducing peptide identified in hypothalamic regions that regulate energy balance, is an integral regulator of energy homeostasis and a putative glucose-dependent insulin coadjuvant. We investigated its production by human cardiomyocytes and its effects on glucose uptake, in the main cardiac glucose transporter GLUT-4 and in intracellular signaling. Quantitative RT-PCR, Western blots, confocal immunofluorescence microscopy, and ELISA of human and murine cardiomyocytes and/or cardiac tissue showed that cardiomyocytes can synthesize and secrete nesfatin-1. Confocal microscopy of cultured cardiomyocytes after GLUT-4 labeling showed that nesfatin-1 mobilizes this glucose transporter to cell peripherals. The rate of 2-deoxy-D-[(3)H]glucose incorporation demonstrated that nesfatin-1 induces glucose uptake by HL-1 cells and cultured cardiomyocytes. Nesfatin-1 induced dose- and time-dependent increases in the phosphorylation of ERK1/2, AKT, and AS160. In murine and human cardiac tissue, nesfatin-1 levels varied with diet and coronary health. In conclusion, human and murine cardiomyocytes can synthesize and secrete nesfatin-1, which is able to induce glucose uptake and the mobilization of the glucose transporter GLUT-4 in these cells. Nesfatin-1 cardiac levels are regulated by diet and coronary health.

Funding information:
  • NICHD NIH HHS - R03HD077483(United States)
  • NIDDK NIH HHS - DK081545(United States)

Ghrelin requires p53 to stimulate lipid storage in fat and liver.

  • Porteiro B
  • Endocrinology
  • 2013 Oct 23

Literature context:


Abstract:

Ghrelin, a stomach-derived peptide, stimulates feeding behavior and adiposity. For its orexigenic action, ghrelin triggers a central SIRT1/p53/AMPK pathway. The tumor suppressor p53 also plays an important role in white adipose tissue (WAT), where it is up-regulated in the adipocytes of obese mice. It is not known, however, whether p53 has any role in mediating the peripheral action of ghrelin. In the present study, chronic peripheral ghrelin treatment resulted in increased body weight and fat-mass gain in wild-type mice. Correspondingly, mRNA levels of several adipogenic and fat-storage-promoting enzymes were up-regulated in WAT, whereas hepatic triglyceride content and lipogenic enzymes were also increased in wild-type mice following ghrelin treatment. In contrast, mice lacking p53 failed to respond to ghrelin treatment, with their body weight, fat mass, and adipocyte and hepatic metabolism remaining unchanged. Thus, our results show that p53 is necessary for the actions of ghrelin on WAT and liver, leading to changes in expression levels of lipogenic and adipogenic genes, and modifying body weight.

Funding information:
  • NIA NIH HHS - T32 AG020494(United States)
  • NINDS NIH HHS - R56 NS050525(United States)

Synaptic alterations in the rTg4510 mouse model of tauopathy.

  • Kopeikina KJ
  • J. Comp. Neurol.
  • 2013 Apr 15

Literature context:


Abstract:

Synapse loss, rather than the hallmark amyloid-β (Aβ) plaques or tau-filled neurofibrillary tangles (NFT), is considered the most predictive pathological feature associated with cognitive status in the Alzheimer's disease (AD) brain. The role of Aβ in synapse loss is well established, but despite data linking tau to synaptic function, the role of tau in synapse loss remains largely undetermined. Here we test the hypothesis that human mutant P301L tau overexpression in a mouse model (rTg4510) will lead to age-dependent synaptic loss and dysfunction. Using array tomography and two methods of quantification (automated, threshold-based counting and a manual stereology-based technique) we demonstrate that overall synapse density is maintained in the neuropil, implicating synapse loss commensurate with the cortical atrophy known to occur in this model. Multiphoton in vivo imaging reveals close to 30% loss of apical dendritic spines of individual pyramidal neurons, suggesting these cells may be particularly vulnerable to tau-induced degeneration. Postmortem, we confirm the presence of tau in dendritic spines of rTg4510-YFP mouse brain by array tomography. These data implicate tau-induced loss of a subset of synapses that may be accompanied by compensatory increases in other synaptic subtypes, thereby preserving overall synapse density. Biochemical fractionation of synaptosomes from rTg4510 brain demonstrates a significant decrease in expression of several synaptic proteins, suggesting a functional deficit of remaining synapses in the rTg4510 brain. Together, these data show morphological and biochemical synaptic consequences in response to tau overexpression in the rTg4510 mouse model.

Funding information:
  • Wellcome Trust - (United Kingdom)

Development and distribution of neuronal cilia in mouse neocortex.

  • Arellano JI
  • J. Comp. Neurol.
  • 2012 Mar 1

Literature context:


Abstract:

Neuronal primary cilia are not generally recognized, but they are considered to extend from most, if not all, neurons in the neocortex. However, when and how cilia develop in neurons are not known. This study used immunohistochemistry for adenylyl cyclase III (ACIII), a marker of primary cilia, and electron microscopic analysis to describe the development and maturation of cilia in mouse neocortical neurons. Our results indicate that ciliogenesis is initiated in late fetal stages after neuroblast migration, when the mother centriole docks with the plasma membrane, becomes a basal body, and grows a cilia bud that we call a procilium. This procilium consists of a membranous protrusion extending from the basal body but lacking axonemal structure and remains undifferentiated until development of the axoneme and cilia elongation starts at about postnatal day 4. Neuronal cilia elongation and final cilia length depend on layer position, and the process extends for a long time, lasting 8-12 weeks. We show that, in addition to pyramidal neurons, inhibitory interneurons also grow cilia of comparable length, suggesting that cilia are indeed present in all neocortical neuron subtypes. Furthermore, the study of mice with defective ciliogenesis suggested that failed elongation of cilia is not essential for proper neuronal migration and laminar organization or establishment of neuronal polarity. Thus, the function of this organelle in neocortical neurons remains elusive.

Funding information:
  • NIDA NIH HHS - R01 DA031833(United States)

Expression and localization of the calmodulin-binding protein neurogranin in the adult mouse olfactory bulb.

  • Gribaudo S
  • J. Comp. Neurol.
  • 2009 Dec 10

Literature context:


Abstract:

Neurogranin (Ng) is a brain-specific postsynaptic protein involved in activity-dependent synaptic plasticity through modulation of Ca(2+)/calmodulin (CaM)-dependent signal transduction in neurons. In this study, using biochemical and immunohistochemical approaches, we demonstrate Ng expression in the adult mouse olfactory bulb (OB), the first relay station in odor information processing. We show that Ng is principally associated with the granule cell layer (GCL), which is composed of granule cell inhibitory interneurons. This cell type is continuously renewed during adult life and plays a key role in OB circuits, integrating and modulating the activity of mitral/tufted cells. Our results indicate that Ng localizes in the soma and dendrites of a defined subpopulation of mature GABAergic granule cells, enriched in the deep portion of the GCL. Ng-immunopositive cells largely coexpress the Ca(+)/CaM-dependent kinase IV (CaMKIV), a downstream protein of CaM signaling cascade, whereas no colocalization was observed between Ng and the calcium-binding protein calretinin. Finally, we demonstrate that adult neurogenesis contributes to the Ng-expressing population, with more newly generated Ng-positive cells integrated in the deep GCL. Together, these results provide a new specific neurochemical marker to identify a subpopulation of olfactory granule cells and suggest possible functional implications for Ng in OB plasticity mechanisms.

Funding information:
  • Howard Hughes Medical Institute - R01 NS062798-03(United States)