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p21-Activated kinase-1 (Pak1) is frequently overexpressed and/or amplified in human breast cancer and is necessary for transformation of mammary epithelial cells. Here, we show that Pak1 interacts with and phosphorylates the Calcium/Calmodulin-dependent Protein Kinase II (CaMKII), and that pharmacological inhibition or depletion of Pak1 leads to diminished activity of CaMKII. We found a strong correlation between Pak1 and CaMKII expression in human breast cancer samples, and combined inhibition of Pak1 and CaMKII with small-molecule inhibitors was synergistic and induced apoptosis more potently in Her2 positive and triple negative breast cancer (TNBC) cells. Co-adminstration of Pak and CaMKII small-molecule inhibitors resulted in a dramatic reduction of proliferation and an increase in apoptosis in a 3D cell culture setting, as well as an impairment in migration and invasion of TNBC cells. Finally, mice bearing xenografts of TNBC cells showed a significant delay in tumor growth when treated with small-molecule inhibitors of Pak and CaMKII. These data delineate a signaling pathway from Pak1 to CaMKII that is required for efficient proliferation, migration and invasion of mammary epithelial cells, and suggest new therapeutic strategies in breast cancer.

Engagement of epidermal growth factor (EGF) with its receptor (EGFR) produces a broad range of cancer phenotypes. The overriding aim of this study was to understand EGFR signaling and its regulation by the Ca2+/calmodulin (CaM) dependent protein kinase kinase 2 (CaMKK2) in cancer cells.

Calcium/calmodulin-dependent protein kinase type II delta (CaMKIIδ), the predominant CaMKII isoform expressed in the heart, has been implicated in the progression of myocardial infarction- and pressure overload-induced pathological remodeling. However, the role of CaMKIIδ in volume overload (VO) has not been explored. We have previously reported an activation of CaMKII during transition to HF in long-term VO. Here, we address whether CaMKIIδ is critically involved in the mortality, myocardial remodeling, and heart failure (HF) progression in response to VO. CaMKIIδ knockout (δ-KO) and wild-type (WT) littermates were exposed to aortocaval shunt-induced VO, and the progression of adverse myocardial remodeling was assessed by serial echocardiography, histological and molecular analyses. The mortality rates during 10 weeks of VO were similar in δ-KO and WT mice. Both genotypes displayed comparable eccentric myocardial hypertrophy, altered left ventricle geometry, perturbed systolic and diastolic functions after shunt. Additionally, cardiomyocytes hypertrophy, augmented myocyte apoptosis, and up-regulation of hypertrophic genes were also not significantly different in δ-KO versus WT hearts after shunt. Therefore, CaMKIIδ signaling seems to be dispensable for the progression of VO-induced maladaptive cardiac remodeling. Accordingly, we hypothesize that CaMKIIδ-inhibition as a therapeutic approach might not be helpful in the context of VO-triggered HF.

Glucotoxicity is a critical component of the pathophysiology of type 2 diabetes mellitus; however, the molecular mechanisms of glucotoxicity are still not fully understood. We have attempted to determine the protein kinases involved in glucotoxicity in pancreatic β-cells by the use of a new technique. Using Multi-PK antibodies, which are capable of detecting a wide variety of protein kinases, we analyzed the protein kinase that correlated with insulin synthesis in INS-1 cells under glucotoxic conditions. When expression patterns of protein kinases in INS-1 cells were analyzed by Western blotting with Multi-PK antibodies, a kinase of 63 kd was significantly reduced concomitant with the decrease of insulin secretion under glucotoxic conditions. To identify the 63-kd kinase, we used a unique 2-dimensional gel electrophoretic technique and MicroRotofor (Bio-Rad Laboratories, Tokyo, Japan) electrophoresis. From the molecular size of a native kinase/cyanogen bromide fragment and pI value, the 63-kd protein kinase was deduced to be CaMKIV. This was confirmed by Western blotting analysis using anti-CaMKIV antibodies. The decreased CaMKIV levels under glucotoxic conditions recovered to original levels after changing the medium to a normal glucose concentration. Recombinant CaMKIV was degraded in a Ca²+-dependent manner by incubation with cell lysates from INS-1 cells under glucotoxic conditions, and degradation was protected by calpain inhibitor. Furthermore, CaMKIV was reduced in the pancreatic islets of diabetic Otsuka Long-Evans Tokushima fatty rats, whereas that of nondiabetic Long-Evans Tokushima Otsuka rats was not. This study suggests that the abnormal regulation of CaMKIV is a component of β-cell dysfunction caused by high glucose.

Previously, we have reported that high mobility group box 1 (HMGB1), a proinflammatory mediator in sepsis, is released via the IFN-β-mediated JAK/STAT pathway. However, detailed mechanisms are still unclear. In this study, we dissected upstream signaling pathways of HMGB1 release using various molecular biology methods. Here, we found that calcium/calmodulin-dependent protein kinase (CaM kinase, CaMK) is involved in HMGB1 release by regulating IFN-β production. CaMK inhibitor, STO609, treatment inhibits LPS-induced IFN-β production, which is correlated with the phosphorylation of interferon regulatory factor 3 (IRF3). Additionally, we show that CaMK-I plays a major role in IFN-β production although other CaMK members also seem to contribute to this event. Furthermore, the CaMK inhibitor treatment reduced IFN-β production in a murine endotoxemia. Our results suggest CaMKs contribute to HMGB1 release by enhancing IFN-β production in sepsis.

CaMK4 has an important function in autoimmune diseases, and the contribution of CaMK4 in psoriasis remains obscure. Here, we show that CaMK4 expression is significantly increased in psoriatic lesional skin from psoriasis patients compared to healthy human skin as well as inflamed skin from an imiquimod (IMQ)-induced mouse model of psoriasis compared to healthy mouse skin. Camk4-deficient (Camk4-/-) mice treated with IMQ exhibit reduced severity of psoriasis compared to wild-type (WT) mice. There are more macrophages and fewer IL-17A+γδ TCR+ cells in the skin of IMQ-treated Camk4-/- mice compared to IMQ-treated WT mice. CaMK4 inhibits IL-10 production by macrophages, thus allowing excessive psoriatic inflammation. Deletion of Camk4 in macrophages alleviates IMQ-induced psoriatic inflammation in mice. In keratinocytes, CaMK4 inhibits apoptosis as well as promotes cell proliferation and the expression of pro-inflammatory genes such as S100A8 and CAMP. Taken together, these data indicate that CaMK4 regulates IMQ-induced psoriasis by sustaining inflammation and provides a potential target for psoriasis treatment.

Platelet-activating factor (PAF) is an important inflammatory lipid mediator affecting neural plasticity. In the present study, we demonstrated how PAF affects synaptic efficacy through activation of protein kinases in the rat hippocampal CA1 region. In cultured hippocampal neurons, 10 to 1000 nM PAF stimulated autophosphorylation of calcium/calmodulin-dependent protein kinase II (CaMKII) and phosphorylation of synapsin I and myristoylated alanine-rich protein kinase C substrate (MARCKS). In hippocampal CA1 slices, field excitatory postsynaptic potentials (fEPSPs) induced by stimulation of the Schaffer collateral/commissural pathways were significantly increased 10-50 min after exposure to 100 to 1000 nM PAF. Immunoblotting analysis showed that 100 nM PAF treatment for 10 or 50 min significantly and persistently increased CaMKII autophosphorylation in the hippocampal CA1 region. Increased protein kinase Calpha (PKCalpha) autophosphorylation was also seen at the same time point after PAF exposure. By contrast, extracellular signal-regulated kinase (ERK) phosphorylation was slightly but significantly increased at 10 min after PAF exposure. Consistent with increased CaMKII autophosphorylation, AMPA-type glutamate receptor subunit 1 (GluR1) (Ser-831) phosphorylation as a CaMKII postsynaptic substrate significantly increased after 10 or 50 min of treatment, whereas synapsin I (Ser-603) phosphorylation as a presynaptic substrate increased at 10 min in the hippocampal CA1 region. Phosphorylation of MARCKS (Ser-152/156) and NMDA receptor subunit 1 (NR1) (Ser-896) as PKCalpha substrates also significantly increased after 10 min but had not further increased by 50 min in the CA1 region. Increased of fEPSPs induced by PAF treatment completely and/or partly inhibited by KN93 and/or U0126 treatment. These results suggest that PAF induces synaptic facilitation through activation of CaMKII, PKC and ERK in the hippocampal CA1 region.

Background: Tyrosine kinase inhibitors (TKIs) have dramatically improved cancer treatment but are known to cause cardiotoxicity. The pathophysiological consequences of TKI therapy are likely to manifest across different cell types of the heart, yet there is little understanding of the differential adverse cellular effects. Cardiac fibroblasts (CFs) play a pivotal role in the repair and remodeling of the heart following insult or injury, yet their involvement in anti-cancer drug induced cardiotoxicity has been largely overlooked. Here, we examine the direct effects of sunitinib malate and imatinib mesylate on adult rat CF viability, Ca2+ handling and mitochondrial function that may contribute to TKI-induced cardiotoxicity. In particular, we investigate whether Ca2+/calmodulin dependent protein kinase II (CaMKII), may be a mediator of TKI-induced effects. Methods: CF viability in response to chronic treatment with both drugs was assessed using MTT assays and flow cytometry analysis. Calcium mobilization was assessed in CFs loaded with Fluo4-AM and CaMKII activation via oxidation was measured via quantitative immunoblotting. Effects of both drugs on mitochondrial function was determined by live mitochondrial imaging using MitoSOX red. Results: Treatment of CFs with sunitinib (0.1-10 μM) resulted in concentration-dependent alterations in CF phenotype, with progressively significant cell loss at higher concentrations. Flow cytometry analysis and MTT assays revealed increased cell apoptosis and necrosis with increasing concentrations of sunitinib. In contrast, equivalent concentrations of imatinib resulted in no significant change in cell viability. Both sunitinib and imatinib pre-treatment increased Angiotensin II-induced intracellular Ca2+ mobilization, with only sunitinib resulting in a significant effect and also causing increased CaMKII activation via oxidation. Live cell mitochondrial imaging using MitoSOX red revealed that both sunitinib and imatinib increased mitochondrial superoxide production in a concentration-dependent manner. This effect in response to both drugs was suppressed in the presence of the CaMKII inhibitor KN-93. Conclusions: Sunitinib and imatinib showed differential effects on CFs, with sunitinib causing marked changes in cell viability at concentrations where imatinib had no effect. Sunitinib caused a significant increase in Angiotensin II-induced intracellular Ca2+ mobilization and both TKIs caused increased mitochondrial superoxide production. Targeted CaMKII inhibition reversed the TKI-induced mitochondrial damage. These findings highlight a new role for CaMKII in TKI-induced cardiotoxicity, particularly at the level of the mitochondria, and confirm differential off-target toxicity in CFs, consistent with the differential selectivity of sunitinib and imatinib.

Neuropathic and inflammatory pain are major clinical challenges due to their ambiguous mechanisms and limited treatment approaches. N-methyl-D-aspartate receptor (NMDAR) and calcium-calmodulin-dependent protein kinase II (CaMKII) are responsible for nerve system sensation and are required for the induction and maintenance of pain. However, the roles of NMDAR and CaMKII in regulating orofacial pain are still less well known. Here, we established a neuropathic pain model by transecting a mouse inferior alveolar nerve (IAN) and an inflammatory pain model by injecting complete Freund's adjuvant (CFA) into its whisker pad. The Cre/loxp site-specific recombination system was used to conditionally knock out (KO) NR2B in the trigeminal ganglion (TG). Von Frey filament behavioral tests showed that IANX and CFA-induced mechanical allodynia were altered in NR2B-deficient mice. CFA upregulated CaMKIIα and CaMKIIβ in the mouse TG and spinal trigeminal caudate nucleus (SpVc). CaMKIIα first decreased and then increased in the TG after IANX, and CaMKIIβ decreased in the TG and SpVc. CFA and IANX both greatly enhanced the expression of phospho (p)-NR2B, p-CaMKII, cyclic adenosine monophosphate (cAMP), p-ERK, and p-cAMP response element binding protein (CREB) in the TG and SpVc. These neurochemical signal pathway alterations were reversed by the conditional KO of NR2B and inhibition of CaMKII. Similarly, IANX- and CFA-related behavioral alterations were reversed by intra-ganglionic (i.g.) -application of inhibitors of CaMKII, cAMP, and ERK. These findings revealed novel molecular signaling pathways (NR2B-CaMKII-cAMP-ERK-CREB) in the TG- and SpVc-derived latent subsequent peripheral and spinal central sensitization under nerve injury and inflammation, which might be beneficial for the treatment of orofacial allodynia.

Lamins are nuclear envelope proteins involved in various cellular functions, such as DNA modulation, cellular differentiation, and development. In this study, we investigate the role of histamine in lung cancer biology. Since it is known that lamin-A/C is negatively regulated in lung cancer, we hypothesize that histamine signaling is related to nuclear lamin-A/C regulation and cancer progression. Our findings reveal that histamine stimulation enhances lamin-A/C expression in lung cancer cells. Lamin-A/C expression is dependent on histamine-mediated intracellular calcium signaling and subsequent calcium/calmodulin-dependent kinase II (Ca/CaMKII) activation. The nuclear protein nestin, which stabilizes lamin-A/C expression, is also modulated by Ca/CaMKII. However, histamine-mediated lamin-A/C expression is independent of Akt/focal adhesion kinase or autophagy signaling. Histamine stimulation attenuates lung cancer motility in the presence of enhanced lamin-A/C expression. In conclusion, we propose a regulatory mechanism that accounts for the modulation of lamin-A/C levels through the involvement of Ca/CaMKII in cancer cells and provides molecular evidence of histamine signaling in lamin-A/C biology.

The function, trafficking and synaptic signaling of AMPA receptors are tightly regulated by phosphorylation. Ca(2+)/calmodulin-dependent kinase II (CaMKII) phosphorylates the GluA1 AMPA receptor subunit at Ser831 to increase single-channel conductance. We show that CaMKII increases the conductance of native heteromeric AMPA receptors in mouse hippocampal neurons through phosphorylation of Ser831. In addition, co-expression of transmembrane AMPA receptor regulatory proteins (TARPs) with recombinant receptors is required for phospho-Ser831 to increase conductance of heteromeric GluA1-GluA2 receptors. Finally, phosphorylation of Ser831 increases the efficiency with which each subunit can activate, independent of agonist efficacy, thereby increasing the likelihood that more receptor subunits will be simultaneously activated during gating. This underlies the observation that phospho-Ser831 increases the frequency of openings to larger conductances rather than altering unitary conductance. Together, these findings suggest that CaMKII phosphorylation of GluA1-Ser831 decreases the activation energy for an intrasubunit conformational change that regulates the conductance of the receptor when the channel pore opens.

This study reports the identification of splice variants for the calcium/calmodulin-dependent protein kinase II (CaMKII) gene from Nilaparvata lugens, Laodelphax striatellus, and Sogatella furcifera. CaMKII is a multifunctional serine/threonine protein kinase that transduces Ca2+ signals in cells to control a range of cellular processes in the nervous system and muscular tissue. Sequence analysis showed that CaMKII was 99.0% identical at the amino acid level among three rice planthoppers, with the exception of a variable region located in the association domain. Four kinds of 20-81 amino acid "inserts" were found in the variable region. The phylogenetic tree of the deduced amino acid sequences showed that the NlCaMKII isoforms were more closely related to the LsCaMKII isoforms and were slightly distinct from SfCaMKII. CaMKII-E was the dominant type among the five main isoforms. CaMKII genes were constitutively expressed in various nymphal and adult stages and in tested tissues with the predominant transcription occurring in the head. There was no major tissue specificity of isoform expression, but the expression pattern and relative abundance of isoforms varied when compared with the RT-PCR between tissues. In addition, RNAi in N. lugens with dsRNA at a concentration of 200 ng nymph-1 induced a mortality of 77.7% on the 10th day and a reduction in the mRNA expression level of 67.2%. Unlike the holometabolous insect Helicoverpa armigera, the knockdown of NlCaMKII did not suppress the expression of 20E response genes, such as ECR, USP1, and HR3, in N. lugens. These results indicate that the role of CaMKII in hemimetabolous insects may be different from that in holometabolous insects.

Phosphatidylinositol-4-phosphate 5-kinase type-1 gamma (Pip5k1c) is a lipid kinase that plays a pivotal role in the regulation of receptor-mediated calcium signaling in multiple tissues; however, its role in the skeleton is not clear. Here, we show that while deleting Pip5k1c expression in the mesenchymal stem cells using Prx1-Cre transgenic mice does not impair the intramembranous and endochondral ossification during skeletal development, it does cause osteopenia in adult mice, but not rapidly growing young mice. We found Pip5k1c loss dramatically decreases osteoblast formation and osteoid and mineral deposition, leading to reduced bone formation. Furthermore, Pip5k1c loss inhibits osteoblastic, but promotes adipogenic, differentiation of bone marrow stromal cells. Pip5k1c deficiency also impairs cytoplasmic calcium influx and inactivates the calcium/calmodulin-dependent protein kinase, which regulates levels of transcription factor Runx2 by modulating its stability and subsequent osteoblast and bone formation. In addition, Pip5k1c loss reduces levels of the receptor activator of nuclear factor-κB ligand, but not that of osteoprotegerin, its decoy receptor, in osteoblasts in bone and in sera. Finally, we found Pip5k1c loss impairs the ability of bone marrow stromal cells to support osteoclast formation of bone marrow monocytes and reduces the osteoclast precursor population in bone marrow, resulting in reduced osteoclast formation and bone resorption. We conclude Pip5k1c deficiency causes a low-turnover osteopenia in mice, with impairment of bone formation being greater than that of bone resorption. Collectively, we uncover a novel function and mechanism of Pip5k1c in the control of bone mass and identify a potential therapeutic target for osteoporosis.

Metabotropic GABA(B) receptors are known to modulate the activity of voltage-dependent calcium channels. Previously, we have shown that GABA(B) receptors couple to a non-Gi/o G-protein to enhance calcium influx through L-type calcium channels by activating protein kinase C in neonatal rat hippocampal neurons. In this study, the components of this signaling pathway were investigated further. Gαq was knocked down using morpholino oligonucleotides prior to examining GABA(B) -mediated enhancement of calcium influx. When Gαq G-proteins were eliminated using morpholino-mediated knockdown, the enhancing effects of the GABA(B) receptor agonist baclofen (10 μM) on calcium current or entry were eliminated. These data suggest that GABA(B) receptors couple to Gαq to regulate calcium influx. Confocal imaging analysis illustrating colocalization of GABA(B) receptors with Gαq supports this hypothesis. Furthermore, baclofen treatment caused translocation of PKCα (protein kinase C α) but not PKCβ or PKCε, suggesting that it is the α isoform of PKC that mediates calcium current enhancement. Inhibition of calcium/calmodulin-dependent kinase II did not affect the baclofen-mediated enhancement of calcium levels. In summary, activation of GABA(B) receptors during development leads to increased calcium in a subset of neurons through Gαq signaling and PKCα activation without the involvement of calcium/calmodulin-dependent kinase II. Activation of GABA(B) receptors in the neonatal rat hippocampus enhances voltage-dependent calcium currents independently of Gi/o . In this study, knockdown of Gαq with morpholino oligonucleotides abolished enhancement of calcium influx and protein kinase Cα was activated by GABA(B) receptors. Therefore, we hypothesize that GABA(B) receptors couple to Gq to activate PKCα leading to enhancement of L-type calcium current.

Porcine circovirus type 2 (PCV2) induces autophagy via the 5' adenosine monophosphate-activated protein kinase (AMPK)/extracellular signal-regulated kinase (ERK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) pathway in pig kidney PK-15 cells. However, the underlying mechanisms of AMPK activation in autophagy induction remain unknown. With specific inhibitors and RNA interference (RNAi), we show that PCV2 infection upregulated calcium/calmodulin-dependent protein kinase kinase-beta (CaMKKβ) by increasing cytosolic Ca(2+) via inositol 1,4,5-trisphosphate receptor (IP3R). Elevation of cytosolic calcium ion (Ca(2+)) did not seem to involve inositol 1,4,5-trisphosphate (IP3) release from phosphatidylinositol 4,5-bisphosphate (PIP2) by phosphoinositide phospholipase C-gamma (PLC-γ). CaMKKβ then activated both AMPK and calcium/calmodulin-dependent protein kinase I (CaMKI). PCV2 employed CaMKI and Trp-Asp (WD) repeat domain phosphoinositide-interacting protein 1 (WIPI1) as another pathway additional to AMPK signaling in autophagy initiation. Our findings could help better understanding of the signaling pathways of autophagy induction as part of PCV2 pathogenesis. Further research is warranted to study if PCV2 interacts directly with IP3R or indirectly with the molecules that antagonize IP3R activity responsible for increased cytosolic Ca(2+) both in PK-15 cells and PCV2-targeted primary cells from pigs.

Calcium/calmodulin-dependent kinase II (CaMKII) facilitates L-type calcium channel (LTCC) activity physiologically, but may exacerbate LTCC-dependent pathophysiology. We previously showed that CaMKII forms stable complexes with voltage-gated calcium channel (VGCC) beta(1b) or beta(2a) subunits, but not with the beta(3) or beta(4) subunits (Grueter et al. 2008). CaMKII-dependent facilitation of Ca(V)1.2 LTCCs requires Thr498 phosphorylation in the beta(2a) subunit (Grueter et al. 2006), but the relationship of this modulation to CaMKII interactions with LTCC subunits is unknown. Here we show that CaMKII co-immunoprecipitates with forebrain LTCCs that contain Ca(V)1.2alpha(1) and beta(1) or beta(2) subunits, but is not detected in LTCC complexes containing beta(4) subunits. CaMKIIalpha can be specifically tethered to the I/II linker of Ca(V)1.2 alpha(1) subunits in vitro by the beta(1b) or beta(2a) subunits. Efficient targeting of CaMKIIalpha to the full-length Ca(V)1.2alpha(1) subunit in transfected HEK293 cells requires CaMKII binding to the beta(2a) subunit. Moreover, disruption of CaMKII binding substantially reduced phosphorylation of beta(2a) at Thr498 within the LTCC complex, without altering overall phosphorylation of Ca(V)1.2alpha(1) and beta subunits. These findings demonstrate a biochemical mechanism underlying LTCC facilitation by CaMKII.

The p32 protein plays a crucial role in the regulation of cytosolic Ca2+ concentrations ([Ca2+]c) that contributes to the Ca2+‑dependent signaling cascade. Using an adenovirus and plasmid p32‑overexpression system, the aim of the study was to evaluate the role of p32 in the regulation of [Ca2+] and its potential associated with Ca2+‑dependent endothelial nitric oxide synthase (eNOS) activation in endothelial cells. Using electron and confocal microscopic analysis, p32 overexpression was observed to be localized to mitochondria and the endoplasmic reticulum and played an important role in Ca2+ translocation, resulting in increased [Ca2+] in these organelles and reducing cytosolic [Ca2+] ([Ca2+]c). This decreased [Ca2+]c following p32 overexpression attenuated the Ca2+‑dependent signaling cascade of calcium/calmodulin dependent protein kinase II (CaMKII)/AKT/eNOS phosphorylation. Moreover, in aortic endothelia of wild‑type mice intravenously administered adenovirus encoding the p32 gene, increased p32 levels reduced NO production and accelerated reactive oxygen species (ROS) generation. In a vascular tension assay, p32 overexpression decreased acetylcholine (Ach)‑induced vasorelaxation and augmented phenylephrine (PE)‑dependent vasoconstriction. Notably, decreased levels of arginase II (ArgII) protein using siArgII were associated with downregulation of overexpressed p32 protein, which contributed to CaMKII‑dependent eNOS phosphorylation at Ser1177. These results indicated that increased protein levels of p32 caused endothelial dysfunction through attenuation of the Ca2+‑dependent signaling cascade and that ArgII protein participated in the stability of p32. Therefore, p32 may be a novel target for the treatment of vascular diseases associated with endothelial disorders.

EPA, an omega-3 polyunsaturated fatty acid, exerts beneficial effects on human health. However, the molecular mechanisms underlying EPA function are poorly understood. The object was to illuminate molecular mechanism underlying EPA's role. Here, 1H-NMR-based metabolic analysis showed enhanced branched-chain amino acids (BCAAs) and lactate following EPA treatment in skeletal muscle cells. EPA regulated mitochondrial oxygen consumption rate. Furthermore, EPA induced calcium/calmodulin-dependent protein kinase kinase (CaMKK) through the generation of intracellular calcium. This induced the phosphorylation of AMP-activated protein kinase (AMPK) and p38 mitogen-activated protein kinase (p38 MAPK) that led to glucose uptake, and the translocation of glucose transporter type 4 (GLUT4) in muscles. In conclusion, EPA exerts benign effects on glucose through the activation of AMPK-p38 MAPK signaling pathways in skeletal muscles.

Alzheimer's disease (AD), characterized by cognitive impairments, is considered to be one of the most widespread chronic neurodegenerative diseases worldwide. We recently introduced a novel therapeutic agent for AD treatment, the T-type calcium channel enhancer ethyl-8-methyl-2,4-dioxo-2-(piperidin-1-yl)-2H-spiro[cyclopentane-1,3-imidazo[1,2-a]pyridin]-2-ene-3-carboxylate (SAK3). SAK3 enhances calcium/calmodulin-dependent protein kinase II and proteasome activity, thereby promoting amyloid beta degradation in mice with AD. However, the antioxidative effects of SAK3 remain unclear. We investigated the antioxidative effects of SAK3 in olfactory bulbectomized mice (OBX mice), compared with the effects of donepezil as a positive control. As previously reported, single oral administration of both SAK3 (0.5 mg/kg, p.o.) and donepezil (1.0 mg/kg, p.o.) significantly improved cognitive and depressive behaviors in OBX mice. Single oral SAK3 administration markedly reduced 4-hydroxy-2-nonenal and nitrotyrosine protein levels in the hippocampus of OBX mice, which persisted until 1 week after administration. These effects are similar to those observed with donepezil therapy. Increased protein levels of oxidative stress markers were observed in the microglial cells, which were significantly rescued by SAK3 and donepezil. SAK3 could ameliorate oxidative stress in OBX mice, like donepezil, suggesting that the antioxidative effects of SAK3 and donepezil are among the neuroprotective mechanisms in AD pathogenesis.