Possible involvement of alpha B-crystallin in the cardioprotective effect of n-butanol extract of Potentilla anserina L. on myocardial ischemia/reperfusion injury in rat

Background It has been reported that n-butanol extract of Potentilla anserina L (NP) had protective effect against acute myocardial ischemia/reperfusion (I/R) injury in mice. Because of limited phytochemical study on NP, its bioactive compounds and underlying protective mechanisms are largely unclear. Purpose The purpose of this study was to investigate the major bioactive compounds and possible mechanism for the cardioprotective effect of NP on rat with I/R injury. Methods We analyzed the phytochemical isolation of NP and identified the structure of compounds, which was elucidated by a combination of spectroscopic analyses. An I/R model was established by I-30 min/R-2 h in Sprage-Dawley rats. The rats were given intragastric administration of NP (49.3, 98.6, and 197.2 mg•kg−1) continuously for 10 days before I/R operation. The morphological changes and apoptosis of cardiomyocytes were observed by H&E staining, Transmission electron microscope and TUNEL staining respectively. The activities or contents of catalase (CAT), superoxide dismutase (SOD), malondialdehyde (MDA) and glutathione (GSH) in plasma were detected. Apoptosis related factors were also measured by RT-PCR and western blot. In order to discover the underlying mechanism of NP on I/R, we performed proteomic analysis using two-dimensional gel electrophoresis (2D-DIGE) and matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/MS) to describe differential proteins expression. Potential target protein resulted from 2D-DIGE coupled to MALDI-TOF/MS analysis were further confirmed by immunohistochemical staining, RT-PCR, and western blot. Results We isolated and identified 14 compounds, of which 7 compounds belong to triterpenes. Rats pretreated with NP showed a significant increase on the activities of GSH, SOD and CAT, and remarkable decrease on the content of MDA. NP significantly inhibited the apoptosis of cardiomyocyte and decreased the expression of Cyt C and cleaved-caspase-3. Proteomic analysis revealed that alpha B-crystallin (CryAB) might participate in the NP protective effect against I/R. NP enhanced the level of pCryAB Ser59, whereas the expression of CryAB was decreased. Conclusion NP was showed to alleviate I/R injury and inhibit myocardial apoptosis, which might be associated with reduction on oxidative stress and apoptosis. CryAB as a possible target involved in the NP protective effect. This study supplied valuable information to develop novel cardioprotective agents from NP extract.