ABSTRACT
Aim To study the effect of puerarin( Pue) against myocardial ischemia/reperfusion injury and in-volved mitochondrial mechanism. Methods Anoxia/reoxygenation( A/R) injury model was established in H9c2 cell. Recombinant plasmid pFLAG-VDAC1 was constructed. Cells were randomly divided into 4 groups, normal control group ( Control) , A/R group, puerarin group ( Pue + A/R ) , and pFLAG-VDAC1-Pue group. Real-time PCR was used to investigate the expression of VDAC1 at mRNA level, and the expres-sion of protein level was detected by Western blot. LDH and CK activities were measured by automatic bi-ochemical analyzer. Mitochondrial membrane potential ( Δψm) and cell apoptosis were observed by flow cy-tometry method. Mitochondrial swelling test was used to detect the opening of mitochondria permeability tran- sition pore ( mPTP) . Results Compared with control group, the expression of VDAC1 and mRNA was up-regulated in A/R group, LDH and CK activity were el-evated, and then mPTP opened, Δψm collapsed, cell apoptosis was significantly increased. Puerarin pre-treatment can lower the expression of VDAC1, main-tain Δψm, prevent the opening of mPTP, and reduce apoptosis. However, the protective effect of Puerarin could be cancelled by transfection of pFLAG-VDAC1. Conclusions The cardioprotection of Puerarin against A/R injury is closely related to down-regulation of VDAC1 and prevention of mPTP opening.
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Aim To investigate the relationship be-tween the cardioprotection of apigenin ( Api ) from an-oxia/reoxygenation ( A/R) injury and Bcl-2 pathway. Methods H9 c2 cardiomyocytes were cultured and di-vided into normal control group, A/R group, Api pre-treatment group ( Api ) , Api + Bcl-2 inhibitor group ( Api + ABT-737 ) . Expression of Bcl-2 was deter-mined by Western blot,and cell viability was measured by MTT method. LDH, SOD, GSH-Px, MDA activity were determined by chromometry. ROS generation, mi-tochondrial membrane potential and apoptosis were de-termined by flow cytometry. Results 25h after apige-nin precondition,the expression of Bcl-2 was upregulat-ed in cardiomyocytes ( P <0. 01 ) . In the group pre-treated with 40 μmol · L-1 apigenin before A/R, the activity of LDH in culture medium decreased; the ac-tivity of intracellular SOD, GSH-Px increased; the content of MDA and ROS generation decreased; cell viability increased; mitochondrial membrane potential could be more stable and cell apoptosis decreased ( P<0. 01 ) . However, all these protective effects were attenuated significantly in the group pretreated with apigenin and Bcl-2 inhibitor ABT-737 . Conclusion The effect of apigenin against A/R injury in cardiomyo-cytes involves Bcl-2 pathway, and at least partly de-pends on its effect on upregulating the expression of Bcl-2 .
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OBJECTIVE: To study the protection effects of echshinone acid (EA) on the primary cultured rat cardiomyocytes subjected to anoxia-reoxygenation (A/R) injury. METHODS: The primary cultured neonatal rat cardiomyocytes were pretreated with EA (0.5, 5 and 50 μmol · L-1), EA (5 μmol · L-1) and L-NAME (0.1 mmol · L-1), or PD98059 (50 μmol · L-1) respectively for 1 h, and then subjected to A/R injury after 24 h. The cell viability, activities of SOD and GSH-Px, MDA contents, LDH activity in the medium and HSP70 protein expression were measured. RESULTS: Pretreatment with Ech decreased LDH activity and MDA contents, increased cell viability and SOD and GSH-Px activities in a concentration-dependent manner, and increased HSP70 protein expression. The heart protective effects of EA were partly abolished by L-NAME or PD98059. CONCLUSION: Pretreatment with EA for 1h before ischemia can induce delayed cardiomyocyte protective effects by activation of NO and MAPK signaling pathways and increasing expression of HSP70 in rat neonatal cardiomyocytes.
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Objective To observe the effects of 1,3-dipropyl-8-cyclopentylxanthine(DPCPX),an adenosine A1 receptor antagonist,on brain neurons damage induced by hypoxia and reoxygenation(H/R),and to elucidate the relevant mechanisms.Methods An in vitro cultured rat cerebral cortical neuronal H/R damage model was established;the effects of DPCPX were detected at final concentrations of 0(control),25,50,100nmol/L on the lactate dehydrogenase(LDH) release from normoxic neurons and H/R neurons which were treated with hypoxia for 8,12,24 hours followed by reoxygenation for 24 hours;the changes of malondialdehyde(MDA) content,activities of xanthine oxidase(XO) and Ca2+-ATPase in H/R neurons which were treated with hypoxia for 12 hours and reoxygenation for 24 hours brought by administration of DPCPX at the concentration of 100nmol/L were also determined by use of specific reagents.Results With addition of 100nmol/L DPCPX,the LDH release from H/R neurons which were treated with hypoxia for 12 hours and reoxygenation for 24 hours was significantly increased compared with that in control group(P
ABSTRACT
Aim To investigate the effects of chloride on the injury of the ventricular myocytes from anoxia-reoxygenation. Methods Under the conditions of anoxia-reoxygenation injury, primary cultured rat ventricular myocytes were treated with 4-acetanide- 4′-isothiocya- natostilbene -2,2′-disulfonic acid (SITS),4,4′,-dii sothiocya-nostilbene-2,2′-disulfonicacid (DIDS) or replaced Cl~- with equimolar gluconate, respectively. Then the cell viability and the contents of Lactate dehydrogenase (LDH), malondialdehyde (MDA), superoxide dismutase (SOD)and glutathione peroxidase (GSH-Px) in the media were measured. Results The release of LDH and MDA was significantly increased in the anoxia-reoxygenation group, while the cell viability and the activity of SOD, GSH-Px decreased significantly compared with those in the control group. In both Cl~--free+ A-R group and SITS+A-R group, LDH and MDA release was noticeably lower than those of the A-R group, while the cell viability and the activity of SOD, GSH-Px significantly increased compared with those in the anoxia-reoxygenation group. But the cell viability and the contents of LDH, MDA, SOD and GSH-Px in the DIDS+A-R group had no significant change compared with those in the anoxia-reoxygenation group.Conclusion Cl~- plays an important role in anoxia reoxygenation injury. SITS provides effective protection to the cardiac myocyte subjected to anoxia reoxygenation injury, while DIDS cannot provide such protection.