Diabetic cardiomyopathy, especially myocardial ischemia reperfusion (I/R) injury, is a major cause of morbidity and mortality in type 2 diabetic patients. The increasing of basal p38 MAP Kinase (p38 MAPK) activation is a major factor that aggravates cardiac death on diabetic cardiomyopathy. In addition, metformin also shows cardio-protective effects on myocardial ischemia/reperfusion injury. In this study, we investigated the effect of the combination between metformin and p38 MAPK inhibitor (SB203580) in diabetic rats subjected to I/R injury. H9c2 cells were induced into a hyperglycemic condition and treated with metformin, SB203580 or the combination of metformin and SB203580. In addition, cells in both the presence and absence of drug treatment were subjected to simulated ischemia/reperfusion injury. Cell viability and cellular reactive oxygen species (ROS) were determined. Moreover, the Goto-Kakizaki (GK) rats were treated with metformin, SB203580, and the combination of metformin and SB203580 for 4 weeks. Diabetic parameters and cardiac functions were assessed. Finally, rat hearts were induced ischemia/reperfusion injury for the purpose of infarct size analysis and determination of signal transduction. A high-glucose condition did not reduce cell viability but significantly increased ROS production and significantly decreased cell viability after induced sI/R. Treatment using drugs was shown to reduce ROS generation and cardiac cell death. The GK rats displayed diabetic phenotype by increasing diabetic parameters and these parameters were significantly decreased when treated with drugs. Treatment with metformin or SB203580 could significantly reduce the infarct size. Interestingly, the combination of metformin and SB203580 could enhance cardio-protective ability. Myocardial I/R injury significantly increased p38 MAPK phosphorylation, Bax/Bcl-2 ratio and caspase-3 level. Treatment with drugs significantly decreased the p38 MAPK phosphorylation, Bax/Bcl-2 ratio, caspase-3 level and increased Akt phosphorylation. In conclusion, using the combination of metformin and SB203580 shows positive cardio-protective effects on diabetic ischemic cardiomyopathy.

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Myocardial ischemia/reperfusion (I/R) injury is a major cause of mortality and morbidity worldwide. Among factors contributing to I/R injury, proteolytic enzymes could also cause cellular injury, expand the injured area and induce inflammation, which then lead to cardiac dysfunction. Therefore, protease inhibition seems to provide therapeutic benefits. Previous studies showed the cardioprotective effect of secretory leukocyte protease inhibitor (SLPI) against myocardial I/R injury. However, the effect of a post-ischemic treatment with SLPI in an in vivo I/R model has never been investigated. In the present study, recombinant human (rh) SLPI (rhSLPI) was systemically injected during coronary artery occlusion or at the onset of reperfusion. The results show that post-ischemic treatment with rhSLPI could significantly reduce infarct size, Lactate Dehydrogenase (LDH) and Creatine kinase-MB (CK-MB) activity, inflammatory cytokines and protein carbonyl levels, as well as improving cardiac function. The cardioprotective effect of rhSLPI is associated with the attenuation of p38 MAPK phosphorylation, Bax, caspase-3 and -8 protein levels and enhancement of pro-survival kinase Akt and ERK1/2 phosphorylation. In summary, this is the first report showing the cardioprotective effects against myocardial I/R injury of post-ischemic treatments with rhSLPI in vivo. Thus, these results suggest that SLPI could be used as a novel therapeutic strategy to reduce myocardial I/R injury.

Protease enzymes contribute to the initiation of cardiac remodeling and heart failure after myocardial ischemic/reperfusion (I/R) injury. Protease inhibitors attenuate protease activity and limit left ventricular dysfunction and remodeling. Previous studies showed the cardioprotective effect of secretory leukocyte protease inhibitor (SLPI) against I/R injury. However, overexpression of SLPI gene in cardiovascular diseases has only been investigated in an in vitro experiment. Here, cardiac-selective expression of the human secretory leukocyte protease inhibitor (hSLPI) gene and its effect on I/R injury were investigated. Adeno-associated virus (AAV) serotype 9 carrying hSLPI under the control of cardiac-selective expression promoter (cardiac troponin, cTn) was intravenously administered to Sprague–Dawley rats for 4 weeks prior to coronary artery ligation. The results showed that myocardial-selective expression of hSLPI significantly reduced infarct size, cardiac troponin I (cTnI), creatine kinase-MB (CK-MB), and myoglobin levels that all served to improve cardiac function. Moreover, overexpression of hSLPI showed a reduction in inflammatory cytokines, oxidatively modified protein carbonyl (PC) content, ischemia-modified albumin (IMA), and necrosis and cardiac tissue degeneration. In conclusion, this is the first study to demonstrate cardiac-selective gene delivery of hSLPI providing cardioprotection against myocardial I/R injury in an in vivo model.

Background: Ischemic heart disease is a leading cause of death in patients with cardiovascular disease. Natural products containing high antioxidant activity have been used as an alternative therapy to improve the living conditions of patients. In this study, we examine the protective effect of tamarind seed (TS) on myocardial hypoxic injury. Methods: The hypoxia model was mimicked by mineral oil overlayed on H9c2 cardiomyoblasts for 4 h. TS extract was pretreated and administered during the hypoxic condition. Radical scavenging activity of TS extract was measured and exhibited very potent antioxidant activities on 2,2-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays. Results: TS extract at a concentration of 10 µg/ml significantly reversed the effect of hypoxia-induced cell death and intracellular reactive oxygen species (ROS) production. We also observed hypoxia-induced over-expression of both inflammatory cytokine mRNA and activation of cellular apoptosis. Pretreatment of TS extract significantly reduced hypoxia-induced HIF-1α and pro-inflammatory cytokine production, IL-1β and IL-6. The Western blot analysis for apoptotic regulatory molecules, caspase 3, caspase 8 and Bax proteins, also showed hypoxic injury reversal by TS extract treatment. Conclusions: The results suggest that the anti-ischemic effect of TS extract protects against hypoxia-induced injury and has potential to be an effective alternative therapy for ischemic heart disease and oxidative-damage related disease.

Inhibition of proteases shows therapeutic potential. Our previous studies demonstrated the cardioprotection by the Secretory Leukocyte Protease Inhibitor (SLPI) against myocardial ischaemia/reperfusion (I/R) injury. However, it is unclear whether the cardioprotective effect of SLPI seen in our previous works is due to the inhibition of protease enzymes. Several studies demonstrate that the anti-protease independent activity of SLPI could provide therapeutic benefits. Here, we show for the first time that recombinant protein of anti-protease deficient mutant SLPI (L72K, M73G, L74G) (mt-SLPI) could significantly reduce cell death and intracellular reactive oxygen species (ROS) production against an in vitro simulated I/R injury. Furthermore, post-ischaemic treatment of mt-SLPI is found to significantly reduce infarct size and cardiac biomarkers lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) activity, improve cardiac functions, attenuate I/R induced-p38 MAPK phosphorylation, and reduce apoptotic regulatory protein levels, including Bax, cleaved-Caspase-3 and total Capase-8, in rats subjected to an in vivo I/R injury. Additionally, the beneficial effect of mt-SLPI was not significantly different from the wildtype (wt-SLPI). In summary, SLPI could provide cardioprotection without anti-protease activity, which could be more clinically beneficial in terms of providing cardioprotection without interfering with basal serine protease activity.