Microglial dysfunction is increasingly recognized as a key contributor to the pathogenesis of Alzheimer's disease (AD). Environmental enrichment (EE) is well documented to enhance neuronal form and function, but almost nothing is known about whether and how it alters the brain's innate immune system. Here we found that prolonged exposure of naive wild-type mice to EE significantly altered microglial density and branching complexity in the dentate gyrus of hippocampus. In wild-type mice injected intraventricularly with soluble Aβ oligomers (oAβ) from hAPP-expressing cultured cells, EE prevented several morphological features of microglial inflammation and consistently prevented oAβ-mediated mRNA changes in multiple inflammatory genes both in vivo and in primary microglia cultured from the mice. Microdialysis in behaving mice confirmed that EE normalized increases in the extracellular levels of the key cytokines (CCL3, CCL4, TNFα) identified by the mRNA analysis. Moreover, EE prevented the changes in microglial gene expression caused by ventricular injection of oAβ extracted directly from AD cerebral cortex. We conclude that EE potently alters the form and function of microglia in a way that prevents their inflammatory response to human oAβ, suggesting that prolonged environmental enrichment could protect against AD by modulating the brain's innate immune system.

Environmental enrichment (EE) is a rodent behavioral paradigm that can model the cognitive benefits to humans associated with intellectual activity and exercise. We recently discovered EE's anti-inflammatory protection of brain microglia against soluble oligomers of human amyloid β-protein (oAβ). Mechanistically, we report that the key factor in microglial protection by EE is chronically enhanced β-adrenergic signaling. Quantifying microglial morphology and inflammatory RNA profiles revealed that mice in standard housing (SH) fed the β-adrenergic agonist isoproterenol experienced similar protection of microglia against oAβ-induced inflammation as did mice in EE Conversely, mice in EE fed the β-adrenergic antagonist propranolol lost microglial protection against oAβ. Mice lacking β1/β2-adrenergic receptors showed no protection of microglia by EE In SH mice, quantification of norepinephrine in hippocampus and interstitial fluid showed that oAβ disrupted norepinephrine homeostasis, and microglial-specific analysis of β2-adrenergic receptors indicated a decreased receptor level. Both features were rescued by EE Thus, enhanced β-adrenergic signaling at the ligand and receptor levels mediates potent benefits of EE on microglial inflammation induced by human Aβ oligomers in vivo.

Post-hemorrhagic hydrocephalus (PHH) refers to a life-threatening accumulation of cerebrospinal fluid (CSF) that occurs following intraventricular hemorrhage (IVH). An incomplete understanding of this variably progressive condition has hampered the development of new therapies beyond serial neurosurgical interventions. Here, we show a key role for the bidirectional Na-K-Cl cotransporter, NKCC1, in the choroid plexus (ChP) to mitigate PHH. Mimicking IVH with intraventricular blood led to increased CSF [K+] and triggered cytosolic calcium activity in ChP epithelial cells, which was followed by NKCC1 activation. ChP-targeted adeno-associated viral (AAV)-NKCC1 prevented blood-induced ventriculomegaly and led to persistently increased CSF clearance capacity. These data demonstrate that intraventricular blood triggered a trans-choroidal, NKCC1-dependent CSF clearance mechanism. Inactive, phosphodeficient AAV-NKCC1-NT51 failed to mitigate ventriculomegaly. Excessive CSF [K+] fluctuations correlated with permanent shunting outcome in humans following hemorrhagic stroke, suggesting targeted gene therapy as a potential treatment to mitigate intracranial fluid accumulation following hemorrhage.

Regulation of the volume and electrolyte composition of the cerebrospinal fluid (CSF) is vital for brain development and function. The Na-K-Cl co-transporter NKCC1 in the choroid plexus (ChP) plays key roles in regulating CSF volume by co-transporting ions and mediating same-direction water movements. Our previous study showed ChP NKCC1 is highly phosphorylated in neonatal mice as the CSF K+ level drastically decreases and that overexpression of NKCC1 in the ChP accelerates CSF K+ clearance and reduces ventricle size [1]. These data suggest that NKCC1 mediates CSF K+ clearance following birth in mice. In this current study, we used CRISPR technology to create a conditional NKCC1 knockout mouse line and evaluated CSF K+ by Inductively Coupled Plasma Optical Emission spectroscopy (ICP-OES). We demonstrated ChP-specific reduction of total and phosphorylated NKCC1 in neonatal mice following embryonic intraventricular delivery of Cre recombinase using AAV2/5. ChP-NKCC1 knockdown was accompanied by a delayed perinatal clearance of CSF K+. No gross morphological disruptions were observed in the cerebral cortex. We extended our previous results by showing embryonic and perinatal rats shared key characteristics with mice, including decreased ChP NKCC1 expression level, increased ChP NKCC1 phosphorylation state, and increased CSF K+ levels compared to adult. Collectively, these follow up data support ChP NKCC1's role in age-appropriate CSF K+ clearance during neonatal development.

The choroid plexus (ChP) epithelium is a source of secreted signaling factors in cerebrospinal fluid (CSF) and a key barrier between blood and brain. Here, we develop imaging tools to interrogate these functions in adult lateral ventricle ChP in whole-mount explants and in awake mice. By imaging epithelial cells in intact ChP explants, we observed calcium activity and secretory events that increased in frequency following delivery of serotonergic agonists. Using chronic two-photon imaging in awake mice, we observed spontaneous subcellular calcium events as well as strong agonist-evoked calcium activation and cytoplasmic secretion into CSF. Three-dimensional imaging of motility and mobility of multiple types of ChP immune cells at baseline and following immune challenge or focal injury revealed a range of surveillance and defensive behaviors. Together, these tools should help illuminate the diverse functions of this understudied body-brain interface.

Protein-truncating variants (PTVs) have important impacts on phenotype diversity and disease. However, their population genetics characteristics in more globally diverse populations are not well defined. Here, we describe patterns of PTVs in 1320 genes sequenced in 10,539 healthy controls and 9434 patients with psoriasis, all of Han Chinese ancestry. We identify 8720 PTVs, of which 77% are novel, and estimate 88% of all PTVs are deleterious and subject to purifying selection. Furthermore, we show that individuals with psoriasis have a significantly higher burden of PTVs compared to controls (P = 0.02). Finally, we identified 18 PTVs in 14 genes with unusually high levels of population differentiation, consistent with the action of local adaptation. Our study provides insights into patterns and consequences of PTVs.

Transmission and secretion of signals via the choroid plexus (ChP) brain barrier can modulate brain states via regulation of cerebrospinal fluid (CSF) composition. Here, we developed a platform to analyze diurnal variations in male mouse ChP and CSF. Ribosome profiling of ChP epithelial cells revealed diurnal translatome differences in metabolic machinery, secreted proteins, and barrier components. Using ChP and CSF metabolomics and blood-CSF barrier analyses, we observed diurnal changes in metabolites and cellular junctions. We then focused on transthyretin (TTR), a diurnally regulated thyroid hormone chaperone secreted by the ChP. Diurnal variation in ChP TTR depended on Bmal1 clock gene expression. We achieved real-time tracking of CSF-TTR in awake TtrmNeonGreen mice via multi-day intracerebroventricular fiber photometry. Diurnal changes in ChP and CSF TTR levels correlated with CSF thyroid hormone levels. These datasets highlight an integrated platform for investigating diurnal control of brain states by the ChP and CSF.

Norcantharidin (NCTD) is a Chinese FDA approved, chemically synthesized drug for cancer treatment. The effect of NCTD on signaling proteins of EGFR and c-Met was systematically elucidated in current study.

With the speedy development of sequencing technologies, noninvasive prenatal testing (NIPT) has been widely applied in clinical practice for testing for fetal aneuploidy. The cell-free fetal DNA (cffDNA) concentration in maternal plasma is the most critical parameter for this technology because it affects the accuracy of NIPT-based sequencing for fetal trisomies 21, 18 and 13. Several approaches have been developed to calculate the cffDNA fraction of the total cell-free DNA in the maternal plasma. However, most approaches depend on specific single nucleotide polymorphism (SNP) allele information or are restricted to male fetuses.

Melanoma is one of the most malignant and aggressive types of cancer worldwide. Fibroblast growth factor 2 (FGF2) is one of the critical regulators of melanoma angiogenesis and metastasis; thus, it might be an effective anti-cancer strategy to explore FGF2-targeting drug candidates from existing drugs. In this study, we evaluate the effect of the marine drug propylene glycol alginate sodium sulfate (PSS) on FGF2-mediated angiogenesis and invasion. The data shows that FGF2 selectively bound to PSS with high affinity. PSS inhibited FGF2-mediated angiogenesis in a rat aortic ring model and suppressed FGF2-mediated invasion, but not the migration of murine melanoma B16-F10 cells. The further mechanism study indicates that PSS decreased the expression of activated matrix metalloproteinase 2 (MMP-2) and matrix metalloproteinase 9 (MMP-9), and also suppressed their activity. In addition, PSS was found to decrease the level of Vimentin in B16-F10 cells, which is known to participate in the epithelial-mesenchymal transition. Notably, PSS did not elicit any changes in cancer cell viability. Based on the results above, we conclude that PSS might be a potential drug to regulate the tumor microenvironment in order to facilitate the recovery of melanoma patients.

Cerebrospinal fluid (CSF) provides vital support for the brain. Abnormal CSF accumulation, such as hydrocephalus, can negatively affect perinatal neurodevelopment. The mechanisms regulating CSF clearance during the postnatal critical period are unclear. Here, we show that CSF K+, accompanied by water, is cleared through the choroid plexus (ChP) during mouse early postnatal development. We report that, at this developmental stage, the ChP showed increased ATP production and increased expression of ATP-dependent K+ transporters, particularly the Na+, K+, Cl-, and water cotransporter NKCC1. Overexpression of NKCC1 in the ChP resulted in increased CSF K+ clearance, increased cerebral compliance, and reduced circulating CSF in the brain without changes in intracranial pressure in mice. Moreover, ChP-specific NKCC1 overexpression in an obstructive hydrocephalus mouse model resulted in reduced ventriculomegaly. Collectively, our results implicate NKCC1 in regulating CSF K+ clearance through the ChP in the critical period during postnatal neurodevelopment in mice.

Reissner's fiber (RF) is an extracellular polymer comprising the large monomeric protein SCO-spondin (SSPO) secreted by the subcommissural organ (SCO) that extends through cerebrospinal fluid (CSF)-filled ventricles into the central canal of the spinal cord. In zebrafish, RF and CSF-contacting neurons (CSF-cNs) form an axial sensory system that detects spinal curvature, instructs morphogenesis of the body axis, and enables proper alignment of the spine. In mammalian models, RF has been implicated in CSF circulation. However, challenges in manipulating Sspo, an exceptionally large gene of 15,719 nucleotides, with traditional approaches has limited progress. Here, we generated a Sspo knockout mouse model using CRISPR/Cas9-mediated genome-editing. Sspo knockout mice lacked RF-positive material in the SCO and fibrillar condensates in the brain ventricles. Remarkably, Sspo knockout brain ventricle sizes were reduced compared to littermate controls. Minor defects in thoracic spine curvature were detected in Sspo knockouts, which did not alter basic motor behaviors tested. Altogether, our work in mouse demonstrates that SSPO and RF regulate ventricle size during development but only moderately impact spine geometry.

Seizures are due to excessive, synchronous neuronal firing in the brain and are characteristic of epilepsy, the fourth most prevalent neurological disease. We report handling-induced and spontaneous seizures in mice deficient for CD39, a cell-surface ATPase highly expressed on microglial cells. CD39-/- mice with handling-induced seizures had normal input-output curves and paired-pulse ratio measured from hippocampal slices and lacked microgliosis, astrogliosis or overt cell loss in the hippocampus and cortex. As expected, however, the cerebrospinal fluid of CD39-/- mice contained increased levels of ATP and decreased levels of adenosine. To determine if immune activation was involved in seizure progression, we challenged mice with lipopolysaccharide (LPS) and measured the effect on microglia activation and seizure severity. Systemic LPS challenge resulted in increased cortical staining of Iba1/CD68 and gene array data from purified microglia predicted increased expression of interleukin-8, triggering receptor expressed on myeloid cells 1, p38, pattern recognition receptors, death receptor, nuclear factor-κB , complement, acute phase, and interleukin-6 signalling pathways in CD39-/- versus CD39+/+ mice. However, LPS treatment did not affect handling-induced seizures. In addition, microglia-specific CD39 deletion in adult mice was not sufficient to cause seizures, suggesting instead that altered expression of CD39 during development or on non-microglial cells such as vascular endothelial cells may promote the seizure phenotype. In summary, we show a correlation between altered extracellular ATP/adenosine ratio and a previously unreported seizure phenotype in CD39-/- mice. This work provides groundwork for further elucidation of the underlying mechanisms of epilepsy.

G protein-coupled receptors (GPCRs) are key regulators of human physiology and are the targets of many small molecule research compounds and therapeutic drugs. While most of these ligands bind to their target GPCR with high affinity, selectivity is often limited at the receptor, tissue, and cellular level. Antibodies have the potential to address these limitations but their properties as GPCR ligands remain poorly characterized. Here, using protein engineering, pharmacological assays, and structural studies, we develop maternally selective heavy chain-only antibody ("nanobody") antagonists against the angiotensin II type I receptor (AT1R) and uncover the unusual molecular basis of their receptor antagonism. We further show that our nanobodies can simultaneously bind to AT1R with specific small-molecule antagonists and demonstrate that ligand selectivity can be readily tuned. Our work illustrates that antibody fragments can exhibit rich and evolvable pharmacology, attesting to their potential as next-generation GPCR modulators.