Ning Su

Up-regulation of heme oxygenase-1 attenuates brain damage after cerebral ischemia via simultaneous inhibition of superoxide production and preservation of NO bioavailability.

Cerebral ischemia exacerbates neuronal death and neurological dysfunction. Evidence supports the involvement of oxidative/nitrative stress in the pathophysiology of cerebral ischemia. Heme oxygenase-1 (HO-1) is a rate-limiting enzyme in heme catabolism, possessing potent anti-oxidant and anti-apoptosis effects. In transgenic mice, HO-1 overproduction is neuroprotective against cerebral ischemia injury, but by unclear mechanisms. The present study determined whether treatment with adenoviral vector overexpressing HO-1 (Ad-HO-1) attenuates post-ischemic brain damage via reduction of oxidative/nitrative stress. After focal cerebral ischemia, Ad-HO-1 reduced lipid peroxidation and protein nitration, decreased infarct volume, and attenuated neurologic deficits. Zinc protoporphyrin IX (ZnPP IX, a specific HO-1 inhibitor) blocked Ad-HO-1 mediated effects against ischemic brain damage. Although Ad-HO-1 slightly reduced ischemic brain NO concentrations, Ad-HO-1 treatment significantly inhibited cerebral expression of iNOS protein expression, without significant effect upon nNOS or eNOS expression compared to vehicle after focal cerebral ischemia. Ad-HO-1 preserved NO bioavailability by increasing eNOS phosphorylation during ischemia compared to vehicle. Together, our results suggest that Ad-HO-1 attenuates post-ischemic brain damage via simultaneous reduction of oxidative/nitrative stress and preservation of NO bioavailability.

The brain-derived neurotrophic factor is associated with alcohol dependence-related depression and antidepressant response

Brain-derived neurotrophic factor (BDNF) plays an essential role in neuronal survival, proliferation, and synaptic remodeling and modulates the function of many other neurotransmitters. Additionally, it likely underlies neurodegenerative and psychiatric disorders, including alcohol dependence-related depression (AD-D). Here, we investigated the possible association between three single nucleotide polymorphisms (SNPs) of the BDNF gene (rs13306221, rs6265, rs16917204) and AD-D. Of 548 patients with alcohol dependence (AD), 166 had AD-D and 312 healthy controls. Response to 8-week sertraline treatment was also assessed. The frequency of the A allele of rs6265 (Val66Met) was significantly higher in AD-D patients than in the healthy controls (p=0.009 after Bonferroni correction). The analysis revealed a strong association between the rs6265 genotype distribution and AD-D (p=0.005 after Bonferroni correction), and the A allele of rs6265 was significantly overrepresented in AD-D patients compared to AD without depression (AD-nD) patients (p=0.001 after Bonferroni correction). Additionally, carriers of the A allele of rs6265 responded better to sertraline treatment (p=0.001). Our results suggested a novel association between BDNF rs6265 and AD-D. These findings might lead to earlier detection of AD-D, perhaps providing better tools for clinical care of these patients in the future.

Thioredoxin-1 attenuates post-ischemic neuronal apoptosis via reducing oxidative/nitrative stress

Recent studies show that Thioredoxin (Trx) possesses a neuronal protective effect and that Trx inactivation is closely related to cerebral ischemia injury. Peroxynitrite (ONOO⁻) formation may trigger oxidative/nitrative stress and represent a major cytotoxic effect in cerebral ischemia. The present study was conducted to validate whether treatment with recombinant human Trx-1 (rhTrx-1) would attenuate ONOO⁻ generation and oxidative/nitrative stress in focal transient cerebral ischemia. The results showed that intravenously administered rhTrx-1 (10 mg/kg) significantly improved neurological functions and reduced cerebral infarction and apoptotic cell death following cerebral ischemia. Neuronal ONOO⁻ formation was significantly attenuated after rhTrx-1 treatment. Moreover, rhTrx-1 resulted in a significant decrease in antioxidant capacity and p38 mitogen activated protein kinase (MAPK) activity in ischemic brain tissue. Furthermore, the suppression on ONOO⁻ formation by either rhTrx-1 or an ONOO⁻ scavenger uric acid reduced cerebral infarct size in mice subjected to cerebral ischemia. Peroxynitrite donor SIN-1 not only blocked the neuronal protection of rhTrx-1 but also markedly attenuated rhTrx-1-induced antioxidative/antinitrative effect. We concluded that rhTrx-1 exerts an antioxidative/antinitrative effect against cerebral ischemia injury by blocking ONOO⁻ and superoxide anion formation. These results provide the information that thioredoxin is much more likely to succeed as a therapeutic approach to diminish oxidative/nitrative stress-induced neuronal apoptotic cell death in the ischemic brain.

Blockade of SOCE protects HT22 cells from hydrogen peroxide-induced apoptosis.

Oxidative stress is an established event in the pathology of neurobiological diseases. Previous studies indicated that store-operated Ca(2+) entry (SOCE) has been involved in oxidative stress. The present study was carried out to investigate the effects of SOCE inhibition on neuronal oxidative stress injury induced by hydrogen peroxide (H2O2) in HT22 cells, a murine hippocampal neuronal model. H2O2 insult induced significant intracellular Ca(2+) overload, mitochondrial dysfunction and cell viability decrease. Inhibition of SOCE by pharmacological inhibitor and STIM1 RNAi significantly alleviated intracellular Ca(2+) overload, restored the mitochondrial membrane potential (MMP), decreased cytochrome C release and eventually inhibited H2O2-induced cell apoptosis. These findings suggest that SOCE inhibition exhibited neuroprotection against oxidative stress induced by H2O2 and SOCE might be a useful therapeutic target in neurobiological disorders.

Downregulation of STIM2 improves neuronal survival after traumatic brain injury by alleviating calcium overload and mitochondrial dysfunction.

Although store-operated calcium entry (SOCE) has been implicated in several neurological disorders, the exact mechanism for its role in traumatic brain injury (TBI) has not been elucidated. In this study, we found that TBI upregulated the expression of a calcium sensor protein called stromal interactive molecule 2 (STIM2); however, the levels of its homologue, STIM1, were unaffected. Both STIM1 and STIM2 are crucial components of SOCE, both in vivo and in vitro. Using shRNA, we discovered that downregulation of STIM2, but not STIM1, significantly improved neuronal survival in both an in vitro and in vivo model of TBI, decreasing neuronal apoptosis, and preserving neurological function. This neuroprotection was associated with alleviating TBI-induced calcium overload and preserving mitochondrial function. Additionally, downregulation of STIM2 not only inhibited Ca(2+) release from the endoplasmic reticulum (ER), but also reduced SOCE-mediated Ca(2+) influx, decreased mitochondrial Ca(2+), restored mitochondrial morphology and improved mitochondrial function, including MMP maintenance, ROS production and ATP synthesis. These results indicate that inhibition of STIM2 can protect neurons from TBI by inhibiting calcium overload and preserving mitochondrial function. This suggests that STIM2 might be an effective interventional target for TBI.

Mitofusin 2 ameliorates hypoxia-induced apoptosis via mitochondrial function and signaling pathways.

Mitochondrial dynamics play a critical role in mitochondrial function and signaling. Although mitochondria play a critical role in hypoxia/ischemia, the further mechanisms between mitochondrial dynamics and ischemia are still unclear. The current study aimed to determine the role of mitofusin 2, a key regulator of mitochondrial fusion, in a hypoxic model and to explore a novel strategy for cerebral ischemia via modulation of mitochondrial dynamics. To the best of our knowledge, this is the first study to investigate both mitochondrial function and molecular pathways to determine the role of mitofusin 2 in hypoxia-induced neuronal apoptosis. In vivo, C57BL/6 mice (male, 19-25g) underwent a permanent middle cerebral artery occlusion for 12 or 24h (n=6 per group). In vitro, cobalt chloride was used to mimic hypoxia in immortalized hippocampal neurons. Down- or up-regulation of Mfn2 was induced to investigate the role of Mfn2 in hypoxia, especially in mitochondrial function and signaling pathways. The findings demonstrated that decreased mitofusin 2 occurred both in vivo and in vitro hypoxic models; second, the anti-apoptotic effect of Mfn2 may work via restoration of mitochondrial function; third, the modulation of the B Cell Leukemia 2/Bcl-2 Associated X protein and extracellular signal-regulated kinase 1/2 signaling pathways highlight the role of Mfn2 in signaling pathways beyond fusion. In summary, depletion of mitofusin 2 would lead to apoptosis both in normal or hypoxic conditions; however, mitofusin 2 overexpression could attenuate hypoxia-induced apoptosis, which represents a potential novel strategy for neuroprotection against ischemic brain damage.

Neuroprotective effects of crocin against traumatic brain injury in mice: Involvement of notch signaling pathway

This study investigated the protective effects and mechanisms of crocin, an extract of saffron, on brain damage after traumatic brain injury (TBI) in mice. C57BL/6 mice were subjected to controlled cortical impact (CCI)-induced TBI. Pretreatment with crocin (20mg/kg) had protective effects against TBI, demonstrated by improved neurological severity score (NSS) and brain edema, decreased microglial activation and release of several pro-inflammatory cytokines, and decreased cell apoptosis. TBI activated Notch signaling, as shown by upregulated levels of Notch intracellular domain (NICD) and Hes1 mRNA, and pretreatment with crocin further increased Notch activation. However, pretreatment with DAPT (100mg/kg), a gamma-secretase inhibitor, significantly suppressed crocin-induced activation of Notch signaling and attenuated the ability of crocin to protect mice against TBI-induced inflammation and apoptosis. Therefore, these results suggest that crocin has neuroprotective effects against TBI in mice, and these effects are at least partially dependent on activation of Notch signaling.

Homer1a attenuates glutamate-induced oxidative injury in HT-22 cells through regulation of store-operated calcium entry

Calcium disequilibrium is extensively involved in oxidative stress-induced neuronal injury. Although Homer1a is known to regulate several neuronal calcium pathways, its effects on, or its exact relationship with, oxidative stress-induced neuronal injury has not yet been fully elucidated. We found that Homer1a protected HT-22 cells from glutamate-induced oxidative stress injury by inhibiting final-phase intracellular calcium overload and mitochondrial oxidative stress. In these cells, stromal interactive molecule 1 (STIM1) puncta, but not the protein level, was significantly increased after glutamate treatment. Store-operated calcium entry (SOCE) inhibitors and cells in which a key component of SOCE (STIM1) was knocked out were used as glutamate-induced oxidative stress injury models. Both models demonstrated significant improvement of HT-22 cell survival after glutamate treatment. Additionally, increased Homer1a protein levels significantly inhibited SOCE and decreased the association of STIM1-Orai1 triggered by glutamate. These results suggest that up-regulation of Homer1a can protect HT-22 cells from glutamate-induced oxidative injury by disrupting the STIM1-Oria1 association, and then by inhibiting the SOCE-mediated final-phrase calcium overload. Thus, regulation of Homer1a, either alone or in conjunction with SOCE inhibition, may serve as key therapeutic interventional targets for neurological diseases in which oxidative stress is involved in the etiology or progression of the disease.

Inhibition of Na+-K+-2Cl- Cotransporter-1 attenuates traumatic brain injury-induced neuronal apoptosis via regulation of Erk signaling

Traumatic brain injury (TBI) is the leading cause of mortality and morbidity worldwide and is characterized by immediate brain damage and secondary injuries, such as brain edema and ischemia. However, the exact pathological mechanisms that comprise these associated secondary injuries have not been fully elucidated. This study aimed to investigate the role of the Na(+)-K(+)-2Cl(-) cotransporter-1 (NKCC1) in the disruption of ion homeostasis and neuronal apoptosis in TBI. Using a traumatic neuron injury (TNI) model in vitro and a controlled cortex injury (CCI) model in vivo, the present study investigated changes in the expression and effects of NKCC1 in TBI using western blot, RNA interference, a lactate dehydrogenase (LDH) release assay, TdT-mediated dUTP Nick end-labeling (TUNEL) analysis, sodium imaging, brain water content, and neurological severity scoring. TBI induced the expression of NKCC1 to be significantly upregulated in the cortex, both in vitro and in vivo. Pharmacological inhibitor bumetanide (Bume) or NKCC1 RNA interference significantly attenuated TBI-induced intracellular Na(+) increase, inhibited neuronal apoptosis, and improved brain edema and neurological function. Furthermore, NKCC1 inhibition also significantly inhibited TBI-induced extracellular signal-regulated kinase (Erk) activation. Erk inhibition significantly protected neurons from TBI injury; however, Erk inhibition had no effect on NKCC1 expression or the neuroprotective effect of NKCC1 inhibition against TBI. This study demonstrates the role of NKCC1 in TBI-induced brain cortex injury, establishing that NKCC1 may play a neurotoxic role in TBI and that the inhibition of NKCC1 may protect neurons from TBI via the regulation of Erk signaling.

Upregulation of Homer1a Promoted Retinal Ganglion Cell Survival After Retinal Ischemia and Reperfusion via Interacting with Erk Pathway

Retinal ischemia and reperfusion (I/R) is extensively involved in ocular diseases, causing retinal ganglion cell (RGCs) death resulting in visual impairment and blindness. Homer1a is considered as an endogenous neuroprotective protein in traumatic brain injury. However, the roles of Homer1a in RGCs I/R injury have not been elucidated. The present study investigated the changes in expression and effect of Homer1a in RGCs both in vitro and in vivo after I/R injury using Western blot, TUNEL assay, gene interference and overexpression, and gene knockout procedures. The levels of Homer1a and phosphorylated Erk (p-Erk) increased in RGCs and retinas after I/R injury. Upregulation of Homer1a in RGCs after I/R injury decreased the level of p-Erk, and mitigated RGCs apoptosis. Conversely, downregulation of Homer1a increased the level of p-Erk, and augmented RGCs apoptosis. Furthermore, inhibition of the p-ERK reduced RGCs apoptosis, and increased the expression of Homer 1a after I/R injury. Finally, the retinas of Homer1a KO mice treated with I/R injury had significantly less dendrites and RGCs, compared with Homer1a WT mice. These findings demonstrated that Homer1a may contribute to RGCs survival after I/R injury by interacting with Erk pathway.