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Epigenetic modification contributes to the pathogenesis of cerebral ischemia. Piwil2 belongs to the PIWI proteins subfamily and has a key role in the regulation of gene transcription through epigenetics. However, the roles of Piwil2 in cerebral ischemia have not been investigated. In this study, we aim to elucidate the roles and the underlying molecular mechanisms of Piwil2 in ischemic tolerance induced by hypoxic postconditioning (HPC) against transient global cerebral ischemia (tGCI). We found that the expression of Piwil2 in CA1 was downregulated by HPC after tGCI. Silencing Piwil2 with antisense oligodeoxynucleotide (AS‐ODN) in CA1 after tGCI decreased the expression of apoptosis‐related proteins and exerted neuroprotective effects. Opposite results were observed after overexpression of Piwil2 induced by administration of Piwil2‐carried lentivirus. Furthermore, we revealed differentially expressed Piwil2‐interacting piRNAs in CA1 between HPC and tGCI groups by RNA binding protein immunoprecipitation (RIP) assay. Moreover, downregulating Piwil2 induced by HPC or AS‐ODN after tGCI caused a marked reduction of DNA methyltransferase 3A (DNMT3A), which in turn abolished the tGCI‐induced increase in the DNA methylation of cyclic AMP response element‐binding 2 (CREB2), thus increasing mRNA and protein of CREB2. Finally, downregulating Piwil2 restored dendritic complexity and length, prevented the loss of dentritic spines, thereby improving cognitive function after tGCI. These data firstly reveal that Piwil2 plays an important part in HPC‐mediated neuroprotection against cerebral ischemia through epigenetic regulation of CREB2. [ABSTRACT FROM AUTHOR]

Abstract: We demonstrated previously that 30min of hypoxic preconditioning (HPC) applied 1day before 10min of transient global cerebral ischemia (tGCI) reduced neuronal loss in the hippocampal CA1 subregion in adult rats. The aim of the present study was to investigate the role of Na+/K+-ATPase and protein kinase Mζ (PKMζ) in the protective effect of HPC against tGCI in adult rats. We found that the activity of Na+/K+-ATPase decreased in the hippocampal CA1 subregion after 10min of tGCI. This effect was not seen after 30min of HPC in adult rats. Corresponding to the changes in Na+/K+-ATPase activity, the surface expression of Na+/K+-ATPase α1 subunit increased after HPC. Furthermore, HPC dramatically reduced the number of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells in the hippocampal CA1 subregion after tGCI. However, neither PKMζ nor phosphorylation of PKMζ was changed after tGCI or HPC. The results of the present study are consistent with the hypothesis that both enhanced recovery of Na+/K+-ATPase activity due to preserved the protein levels of Na+/K+-ATPase α1 subunit and reduced DNA fragmentation after tGCI contribute to the protection afforded by HPC. However, PKMζ activation does not appear to play a role in this neuroprotection. [Copyright &y& Elsevier]

Hypoxic preconditioning (HPC) alleviates the selective and delayed neuronal death in the hippocampal CA1 region induced by transient global cerebral ischemia (tGCI). This type of cell death may include different programmed cell death mechanisms, namely, apoptosis and necroptosis. Although apoptotic signaling is well defined, the mechanisms that underlie neuronal necroptosis are yet to be fully elucidated. In this study, we investigated whether HPC protects neurons from cerebral ischemia-induced necroptosis. We observed that tGCI up-regulated the expression of receptor-interacting protein (RIP) 3 and increased the interaction of RIP1-RIP3 in CA1 at the early stage of reperfusion. The pretreatment with HPC or necrostatin-1 decreased the expression of RIP3 and the formation of RIP1-RIP3 after tGCI. We also found that HPC decreased the expression and the activity of caspase-8 in CA1 after tGCI, and notably, the pretreatment with Z-VAD-FMK, a pan-caspase inhibitor, did not trigger necroptosis but attenuated the tGCI-induced neuronal damage. Furthermore, we demonstrated that HPC decreased the activation of calcium-calmodulin kinase (CaMK) IIα and the interaction of RIP1 and CaMKIIα induced by tGCI. Intriguingly, the pretreatment with a CaMKs inhibitor KN-93 before tGCI resulted in significantly reduced RIP1-3 interaction and tGCI-induced neuronal damage. Finally, we ascertained that HPC prevented the dephosphorylation of dynamin-related protein 1 (Drp1)-Ser637 (serine 637) and inhibited the translocation of Drp1 to mitochondria induced by tGCI. Importantly, the treatment with a Drp1 inhibitor Mdivi-1 or necrostatin-1 before tGCI also abolished Drp1 dephosphorylation at Ser637 and mitochondrial translocation. Taken together, our results highlight that HPC attenuates necroptotic neuronal death induced by tGCI via Drp1-dependent mitochondrial signaling pathways mediated by CaMKIIα inactivation.-Zhan, L., Lu, Z., Zhu, X., Xu, W., Li, L., Li, X., Chen, S., Sun, W., Xu, E. Hypoxic preconditioning attenuates necroptotic neuronal death induced by global cerebral ischemia via Drp1-dependent signaling pathway mediated by CaMKIIα inactivation in adult rats.

Hypoxic postconditioning (HPC) is an innovative neuroprotective strategy with cytoprotective effects on the hippocampal neurons against transient global cerebral ischemia (tGCI) in adult rats. However, its molecular mechanisms have not yet been adequately elucidated. Neuroglobin (Ngb) is an endogenous neuroprotectant with hypoxia-inducible property, and its role in experimental stroke has been increasingly attractive. Hence, the purpose of this study is to explore the involvement of Ngb in HPC-mediated neuroprotection and to further investigate its underlying molecular mechanism. We found that HPC increased Ngb expression in CA1 subregion after tGCI. Also, the inhibition of Ngb expression with Ngb antisense oligodeoxynucleotide (AS-ODNs) eliminated the neuroprotective effect mediated by HPC, whereas overexpression of Ngb ameliorated neuronal damage in CA1 after tGCI, indicating that HPC conferred neuroprotective effects via upregulation of Ngb. We further showed that HPC increased the membranous level of Na + /K + ATPases β1 subunit (Atp1b1) in CA1 after tGCI. Furthermore, we demonstrated that Ngb upregulation in CA1 after HPC maintained the membranous level of Atp1b1 through Ngb-Atp1b1 interaction and reduced the glutathionylation of membranous Atp1b1 via suppression of reactive oxygen species (ROS), ultimately preserving the activity of NKA. Taken together, these data indicate that Ngb is involved in the neuroprotection of HPC against tGCI via maintenance of NKA activity in the hippocampal CA1.

Objective: To investigate whether lithium modifies open-field and elevated plus maze behavior, and brain phospho-glycogen synthase kinase 3 (P-GSK3beta) expression in Fmr1 knockout mice., Methods: One hundred and eighty FVB mice, including knockout and wild type, with an age of 30 days were used. An open-field and elevated plus maze was utilized to test behavior, while western blot was used to measure the P-GSK3beta expression. Six groups were formed: control (saline), lithium chloride 30, 60, 90, 120, and 200 mg/kg. The experiments were carried out in the Institute of Neuroscience, Second Affiliated Hospital of Guangzhou Medical University, Guangzhou, China between January and June 2012., Results: Lithium significantly decreased total distance, crossing, central area time, and center entry in the open-field test (p<0.05), and significantly reduced open-arm tracking, open-arm entry, and open-arm time in the elevated plus maze (p<0.05) in knockout mice. In wild type mice, significant changes were observed in both behavior tests in some treatment groups. Lithium ameliorated P-GSK3beta expression in the hippocampus of all the treatment groups in knockout mice (p<0.05). However, lithium did not modify either GSK3beta expression in tissues of knockout mice, or P-GSK3beta or GSK3beta expression in tissues of wild type mice., Conclusion: Lithium ameliorated open-field and elevated plus maze behaviors of Fmr1 knockout mice. This effect may be related to its enhancement of P-GSK3beta expression. Our findings suggest that lithium might have a therapeutic effect in fragile X syndrome.

To investigate the protective effects of blocking rennin-angiotensin system (RAS) on the progression of renal injury in glomerulosclerosis, a glomerulosclerosis model was made for SD rats by unilateral nephrectomy and being injected with Adriamycin into caudal vein. The rats with glomerulosclerosis were randomly divided as ten per group into those without further treatment (group D) and those treated with Benazepril (group DB), Losartan (group DL), or sham-operation (group C), respectively. After 6 weeks of administration of Benazepril or Losartan, the mRNA expressions of TGF-beta1, Col IV, Fn, ET-1 and iNOS in renal cortex were measured by RT-PCR. Besides, the expressions of TGF-beta1, ET-1 and iNOS at protein level were detected by Western blotting and the concentrations of Col IV and Fn were analyzed with immunohistochemistry respectively. Results showed that the rats in group D appeared as obvious proteinuria, hypoalbuminemia and hypercholesterolemia, which had a significant difference compared with group C (p < 0.05), and most of their mesangiums were detected with cellular proliferation and significant increasing for extracellular matrix. Renal cortex TGF-beta1, Col IV, Fn, ET-1 and iNOS in rats of group D were increased by 3.59, 2.57, 2.21, 2.58 and 3.28 times at mRNA level, and by 2.60, 1.40, 0.75, 1.83 and 2.15 times at protein level, respectively, compared with group C. When the animals were treated with Benazepril (group DB) or Losartan (group DL), however, the biochemical and pathological damages were significantly recovered, and protein expressions of TGF-beta1, Col IV, Fn, ET-1 and iNOS were also significantly diminished (p < 0.05). This study suggested that blocking RAS using Benazepril or Losartan can have protective effects on the renal injury in glomerulosclerosis by down-regulating the expressions of TGF-beta1, Col IV, Fn, ET-1 and iNOS.