RESUMO
BACKGROUND: Ginsenoside Rg1 (Rg1) has been well documented to be effective against various cardiovascular disease. The aim of this study is to evaluate the effect of Rg1 on mechanical stress-induced cardiac injury and its possible mechanism with a focus on the calcium sensing receptor (CaSR) signaling pathway. METHODS: Mechanical stress was implemented on rats through abdominal aortic constriction (AAC) procedure and on cardiomyocytes and cardiac fibroblasts by mechanical stretching with Bioflex Collagen I plates. The effects of Rg1 on cell hypertrophy, fibrosis, cardiac function, [Ca2+]i, and the expression of CaSR and calcineurin (CaN) were assayed both on rat and cellular level. RESULTS: Rg1 alleviated cardiac hypertrophy and fibrosis, and improved cardiac decompensation induced by AAC in rat myocardial tissue and cultured cardiomyocytes and cardiac fibroblasts. Importantly, Rg1 treatment inhibited CaSR expression and increase of [Ca2+]i, which similar to the CaSR inhibitor NPS2143. In addition, Rg1 treatment inhibited CaN and TGF-ß1 pathways activation. Mechanistic analysis showed that the CaSR agonist GdCl3 could not further increase the [Ca2+]i and CaN pathway related protein expression induced by mechanical stretching in cultured cardiomyocytes. CsA, an inhibitor of CaN, inhibited cardiac hypertrophy, cardiac fibrosis, [Ca2+]i and CaN signaling but had no effect on CaSR expression. CONCLUSION: The activation of CaN pathway and the increase of [Ca2+]i mediated by CaSR are involved in cardiac hypertrophy and fibrosis, that may be the target of cardioprotection of Rg1 against myocardial injury.
RESUMO
Porcine epidemic diarrhea virus (PEDV) is an important pathogen causing severe watery diarrhea, vomiting, dehydration, and death in sucking piglets. Attenuated vaccines have been used widely in sows in order to protect piglets through passive lactogenic immunity. Rapid and sensitive detection methods for differentiating attenuated vaccine strains from virulent ones are essential and practical in PEDV prevention and control. Based on the deletion mutation in ORF3 gene sequence, a TaqMan probe-based real-time quantitative PCR (TaqMan qPCR) was developed to distinguish PEDV virulent strains from attenuated vaccine ones in this study. The TaqMan qPCR could specifically detect PEDV virulent strain but not attenuated vaccine strain and other viruses. At least 37 DNA copies and PEDV of 0.995 TCID50 could be detected by TaqMan qPCR. The reproducibility was evaluated using various dilution of plasmids carrying PEDV ORF3 gene and virulent PEDV, and the inter-assay coefficient of variation (CV) was less than 0.44%. The TaqMan qPCR was further applied to detect 38 samples including intestines and their contents, fecal swabs, and mesenteric lymph nodes. Meanwhile, indirect immunofluorescence assay (IFA) was employed to detect PEDV-specific antigen. PEDV positive rates were 31.58% (12/38) and 26.32% (10/38) by TaqMan PCR and IFA, respectively, which suggested that the former was more sensitive than the latter. The TaqMan qPCR based on PEDV ORF3 gene could be a valuable tool in diagnose of porcine epidemic diarrhea and in molecular epidemiological study of the virulent PEDV.