Hydroxysafflor Yellow A (HSYA) may reduce ischemia/reperfusion (I/R) injury. However, the underlying molecular mechanisms remain unclear. The present study explored the effect and the mechanisms of HSYA on myocardial injury
in vivo
and
in vitro
. Myocardial infarct size was assessed by Evans blue/2,3,5-triphenyltetrazoliumchloride staining. Levels of cardiac troponin I (cTnI), interleukin-6 (IL-6), lactate dehydrogenase (LDH), superoxide dismutase (SOD) and malondialdehyde (MDA) were measured using commercial kits. Alteration of mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) generation was determined by fluorescent signals. Apoptosis was detected by terminal deoxynucleotidyl-transferase-mediated dUTP nick-end labeling staining, flow cytometry assay and caspase-3 activity. Expression levels of the apoptosis-associated proteins were detected by reverse transcription quantitative polymerase chain reaction and western blot analysis.
In vivo
, animals treated with HSYA presented less severe myocardial injury and decreased janus kinase 2 (JAK2)/signal transducer and activator of transcription 1 (STAT1) activity, improved antioxidant capacity and decreased apoptosis.
In vitro
, compared with the hypoxia (H)/reoxygenation (R) + HSYA group, AG490 and S1491 treatment decreased the releases of cTnI, IL-6 and LDH and enhanced the resistance to oxidative stress by maintaining MMP and decreasing ROS generation. In addition, AG490 and S1491 were also identified to alleviate the H/R-induced apoptosis by inhibiting caspase 3 activity and modulating the expression levels of cleaved caspase-3, tumor necrosis factor receptor superfamily member 6 (Fas), Fas ligand, B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein. These data suggested that inactivation of the JAK2/STAT1 pathway strengthened the HSYA-induced protective effect in H/R-induced myocardial injury. In conclusion, the treatment of HSYA was effective in decreasing IR-induced myocardial injury, and this may be largely dependent on the JAK2/STAT1 pathway. Therefore, the present study provided a potential strategy to prevent myocardial I/R injury.
Aim:To explore the effect of neferine on angiotensin II (Ang II)-induced vascular smooth muscle cell (VSMC) proliferation. Methods: Human umbilical vein smooth muscle cells (HUVSMCs) were used. Cell proliferation was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry analysis. Heme oxygenase (HO)-1 protein expression was tested by Western blot analysis. Extracellular signal-regulated protein kinase 1/2 (ERK1/2) activation was determined by using immunoblotting. Results: Pre-incubation of HUVSMCs with neferine (0.1, 0.5, 1.0, and 5.0 µmol/L) significantly inhibited Ang II-induced cell proliferation in a concentration-dependent manner and neferine 5.0 µmol/L increased HO-1 expression by 259% compared with control. The antiproliferative effect of neferine was significantly attenuated by coapplication of zinc protoporphyrin IX (ZnPP IX, an HO-1 inhibitor) with neferine. Ang II-enhanced ERK1/2 phosphorylation was markedly reversed by neferine. By inhibiting HO-1 activity with ZnPP IX, the inhibitive effect of neferine on ERK1/2 phosphorylation was significantly attenuated. Cobalt-protoporphyrin (CoPP), an HO-1 inducer, significantly decreased Ang II-induced ERK1/2 phosphorylation and inhibited Ang II-induced cell proliferation. The ERK1/2 pathway inhibitor PD98059 significantly blocked Ang II-enhanced ERK1/2 phosphorylation and inhibited cell proliferation. Conclusion: These findings suggest that neferine can inhibit Ang II-induced HUVSMC proliferation by upregulating HO-1, leading to the at least partial downregulation of ERK1/2 phosphorylation.
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