Publications (6)14.23 Total impact
- [Show abstract] [Hide abstract] ABSTRACT: DJ-1 protein, as a multifunctional intracellular protein, has an important role in transcriptional regulation and anti-oxidant stress. A recent study by our group showed that DJ-1 can regulate the expression of certain anti‑oxidant enzymes and attenuate hypoxia/re‑oxygenation (H/R)‑induced oxidative stress in the cardiomyocyte cell line H9c2; however, the detailed molecular mechanisms have remained to be elucidated. Nuclear factor erythroid 2‑like 2 (Nrf2) is an essential transcription factor that regulates the expression of several anti‑oxidant genes via binding to the anti‑oxidant response element (ARE). The present study investigated whether activation of the Nrf2 pathway is responsible for the induction of anti‑oxidative enzymes by DJ‑1 and contributes to the protective functions of DJ‑1 against H/R‑induced oxidative stress in H9c2 cells. The results demonstrated that DJ‑1‑overexpressing H9c2 cells exhibited anti‑oxidant enzymes, including manganese superoxide dismutase, catalase and glutathione peroxidase, to a greater extent and were more resistant to H/R‑induced oxidative stress compared with native cells, whereas DJ‑1 knockdown suppressed the induction of these enzymes and further augmented the oxidative stress injury. Determination of the importance of Nrf2 in DJ‑1‑mediated anti‑oxidant enzymes induction and cytoprotection against oxidative stress induced by H/R showed that overexpression of DJ‑1 promoted the dissociation of Nrf2 from its cytoplasmic inhibitor Keap1, resulting in enhanced levels of nuclear translocation, ARE‑binding and transcriptional activity of Nrf2. Of note, Nrf2 knockdown abolished the DJ‑1‑mediated induction of anti‑oxidant enzymes and cytoprotection against oxidative stress induced by H/R. In conclusion, these findings indicated that activation of the Nrf2 pathway is a critical mechanism by which DJ-1 upregulates anti-oxidative enzymes and attenuates H/R-induced oxidative stress in H9c2 cells.
- [Show abstract] [Hide abstract] ABSTRACT: We have recently shown that DJ-1 is implicated in the delayed cardioprotective effect of hypoxic preconditioning (HPC) against hypoxia/reoxygenation (H/R) injury as an endogenous protective protein. This study aims to further investigate the underlying mechanism by which DJ-1 mediates the delayed cardioprotection of HPC against H/R-induced oxidative stress. Using a well-characterized cellular model of HPC from rat heart-derived H9c2 cells, we found that HPC promoted nuclear factor erythroid 2-related factor 2 (Nrf2) and its cytoplasmic inhibitor Kelch-like ECH-associated protein-1 (Keap1) dissociation and resulted in increased nuclear translocation, antioxidant response elements (ARE)-binding and transcriptional activity of Nrf2 at 24 h after HPC, with subsequent upregulation of manganese superoxide dismutase (MnSOD) and heme oxygenase-1 (HO-1), which provided the delayed protection against H/R-induced oxidative stress in normal H9c2 cells. However, the aforementioned effects of HPC were abolished in DJ-1-knockdown H9c2 cells, which were restored by restoration of DJ-1 expression. Importantly, we showed that inhibition of the Nrf2 pathway in H9c2 cells mimicked the effects of DJ-1 knockdown and abolished HPC-derived the induction of antioxidative enzymes (MnSOD and HO-1) and the delayed cardioprotection. In addition, inhibition of Nrf2 also reversed the effects of restored DJ-1 expression on induction of antioxidative enzymes and delayed cardioprotection by HPC in DJ-1-knockdown H9c2 cells. Taken together, the present work revealed that activation of Nrf2 pathway and subsequent upregulation of antioxidative enzymes could be a critical mechanism by which DJ-1 mediates the delayed cardioprotection of HPC against H/R-induced oxidative stress in H9c2 cells.
- [Show abstract] [Hide abstract] ABSTRACT: It has been well demonstrated that hypoxic preconditioning (HPC) can attenuate hypoxia/reoxygenation (H/R)-induced oxidant stress and elicit delayed cardioprotection by upregulating the expression of multiple antioxidative enzymes such as heme oxygenase-1 (HO-1), manganese superoxide dismutase (MnSOD) and so on. However, the underlying mechanisms of HPC-induced upregulation of antioxidative enzymes are not fully understood. Nuclear factor erythroid 2-related factor 2 (Nrf2) is an essential transcription factor that regulates expression of several antioxidant genes via binding to the antioxidant response element (ARE) and plays a crucial role in cellular defence against oxidative stress. Here, we wondered whether activation of the Nrf2-ARE pathway is responsible for the induction of antioxidative enzymes by HPC and contributes to the delayed cardioprotection of HPC. Cellular model of HPC from rat heart-derived H9c2 cells was induced 24 h prior to H/R. The results showed that HPC efficiently attenuated H/R-induced viability loss and lactate dehydrogenase leakage. In addition, HPC increased nuclear translocation and ARE binding of Nrf2 during the late phase, upregulated the expression of antioxidative enzymes (HO-1 and MnSOD), inhibited H/R-induced oxidant stress. However, when Nrf2 was specifically knocked down by siRNA, the induction of antioxidative enzymes by HPC was completely abolished and, as a result, the inhibitory effect of HPC on H/R-induced oxidant stress was reversed, and the delayed cardioprotection induced by HPC was also abolished. These results suggest that HPC upregulates antioxidative enzymes through activating the Nrf2-ARE pathway and confers delayed cardioprotection against H/R-induced oxidative stress.
- [Show abstract] [Hide abstract] ABSTRACT: To investigate the protective effects of preconditioning human umbilical vein endothelial cells (HUVECs) with Polygonum multiflorum stilbeneglycoside (PMS) under anoxia/reoxygenation (A/R), and the mechanism of protection. Prior to A/R, HUVECs were incubated with PMS (0.6 x 10(-11), 1.2 x 10(-11), or 2.4 x 10(-11) mol/L) for 3 h. Cell injury was subsequently evaluated by measuring cell viability with an MTT assay and lactate dehydrogenase (LDH) release, whereas lipid peroxidation was assayed by measuring malondialdehyde (MDA) content. Antioxidant capacity was quantified by superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity. Nitric oxide (NO) production was determined by nitrite accumulation. Endothelial NO synthase (eNOS) and inducible NOS (iNOS) protein expression was detected by Western blotting. Guanylate cyclase activity and cyclic GMP (cGMP) activity were assessed by an enzyme immunoassay kit. PMS incubation attenuated A/R-induced injury in a concentration-dependent manner, as evidenced by a decrease in LDH activity and an increase in cell viability. PMS exerted its protective effect by inhibiting the A/R-mediated elevation of MDA content, as well as by promoting the recovery of SOD and GSH-Px activities. Additionally, PMS incubation enhanced NO and cGMP formation by increasing iNOS expression and guanylate cyclase activity. The protective effects of PMS were markedly attenuated by NOS inhibitor L-NAME, soluble guanylate cyclase inhibitor ODQ or PKG inhibitor KT5823. PMS preincubation resulted in the enhancement of antioxidant activity and anti-lipid peroxidation. The NO/cGMP/cGMP-dependent protein kinase (PKG) signaling pathway was involved in the effect of PMS on HUVECs.
- [Show abstract] [Hide abstract] ABSTRACT: Development of intracellular calcium overload is an important pathophysiological factor in myocardial ischemia/reperfusion or anoxia/reoxygenation injury. Recent studies have shown that Sodium Ferulate (SF) stimulates nitric oxide (NO) production and exerts a cardioprotective effect in the ischemia-reperfused heart. However, it has not been determined whether the cardioprotection of SF is associated with suppression of Ca(2+) overload via NO/cyclic GMP (cGMP)/cGMP-dependent protein kinase (PKG) pathway. In this work, after cardiomyocytes were incubated with 100, 200, 400, or 800 microM SF for 3 h, anoxia/reoxygenation injury was induced and intracellular Ca(2+) concentration, NO synthase (NOS) activity, guanylate cyclase activity, NO, and cGMP formation were measured appropriately. The results showed that treatment with SF concentration-dependently inhibited calcium overload induced by anoxia/reoxygenation. We also demonstrated that SF (100-800 microM) concentration dependently enhanced NO and cGMP formation through increasing NOS activity and guanylate cyclase activity in the cardiomyocytes. On the contrary, inhibition of calcium overload by SF was markedly attenuated by addition of an NOS inhibitor, an NO scavenger, an soluble guanylate cyclase inhibitor, and a PKG inhibitor: N(G)-nitro-l-arginine methyl ester (L-NAME, 100 microM), 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazole-1-oxyl-3-oxide (c-PTIO, 1.0 microM), 1H-[1, 2, 4] oxadiazolo [4, 3-alpha] quinoxalin-1-one (ODQ, 20 microM) and KT5823 (0.2 microM), respectively. Our findings indicate that SF significantly attenuates anoxia/reoxygenation-induced Ca(2+) overload and improves cell survival in cultured cardiomyocytes through NO/cGMP/PKG signal pathway.
- [Show abstract] [Hide abstract] ABSTRACT: Anoxic preconditioning (APC) attenuates myocardial injury caused by ischemia/reperfusion. The protective mechanisms of APC involve up-regulation of the protective proteins and inhibition of apoptosis. 14-3-3 protein, as a molecular chaperone, plays an important role in regulating cell survival and apoptosis. However, the role of 14-3-3 protein in cardioprotection of APC and the pathways determining 14-3-3 protein expression during APC are not clear. In this work, Western blotting analysis was used to detect the 14-3-3 protein expression and activity of extracellular signal-regulated protein kinase 1/2 (ERK1/2) in cardiomyocytes subjected to anoxia-reoxygenation injury with and without APC and control. The cardiomyocytes from APC group were more resistant to injury induced by anoxia-reoxygenation and had much stronger phosphorylation of ERK1/2 than the control. The 14-3-3 protein expression was positively correlated with the phosphorylation of ERK1/2. Furthermore, inhibition of the ERK1/2 with PD98059 abolished the 14-3-3 protein up-regulation in cardiomyocytes induced by APC. The results indicate that APC up-regulates 14-3-3 protein expression through the ERK1/2 signaling pathways.
Nanchang UniversityNan-ch’ang-shih, Jiangxi Sheng, China