背景:半夏泻心汤(BXD)是一种应用于胃肠动力障碍的中药配方。先前的研究表明,miR-451-5p在胃电心律失常诱导的胃肠道运动障碍大鼠中下调。cajal的间质细胞(ICC)是胃肠道运动的起搏器,而ICCs的丧失是胃肠道运动性紊乱的原因。因此,BXD通过miR-451-5p调节ICCs凋亡的潜在相互作用机制仍有待探索.
目的:在这项工作中,主要目的是检查BXD通过miR-451-5p在胃肠道运动障碍大鼠模型和体外对ICC的功效,以及SCF/c-kit信令的潜在贡献。
方法:雄性SD大鼠采用单日饮食和双禁食法(期间饮用稀盐酸水),连续4周建立胃电心律失常大鼠。胃慢波(GSW)记录,RT-qPCR,采用免疫印迹法检测BXD对GED大鼠ICCs凋亡和miR-451-5p表达的影响。体外测定包括CCK-8,流式细胞术分析,RT-qPCR,并应用westernblot研究BXD通过miR-451-5p促进ICCs凋亡的潜在分子机制。
结果:BXD促进胃动力,减少ICCs凋亡,在GED大鼠中miR-451-5p升高。此外,与miR-451-5p抑制剂转染的ICC相比,miR-451-5p在BXD处理后的ICC中显著上调。同时,高miR-451-5p表达与BXD处理或miRNA模拟增强ICCs增殖和抑制凋亡。此外,miR-451-5p的过表达可以逆转BXD处理在ICCs中的G0/G1阻滞。Further,检测SCF和c-kit蛋白水平以证明BXD处理对miR-451-5p的调节涉及该信号传导。
结论:通过这项研究,我们证明BXD可以通过miR-451-5p促进ICCs增殖和抑制凋亡,并且可能涉及SCF/c-kit信号的调节,因此,从靶向miR-451-5p调节ICCs凋亡的角度,为胃肠道运动功能障碍提供了新的治疗基础。
BACKGROUND: Banxia Xiexin Decoction (BXD) is a traditional Chinese medical formula applied to gastrointestinal (GI) motility disorders. Previous studies showed that miR-451-5p was down-regulated in rats with GI motility disorders induced by gastric electrical dysrhythmia. Interstitial cells of Cajal (ICCs) are pacemakers for GI motility, while loss of ICCs is responsible for GI motility disturbance. Thus, the underlying interaction mechanisms for BXD regulating ICCs apoptosis via miR-451-5p remain to be explored.
OBJECTIVE: In this work, the main objectives were to examine the efficacy of BXD on ICCs via miR-451-5p both in GI motility disorders rats model and in vitro, as well as the potential contributions of SCF/c-kit signaling.
METHODS: Rats with gastric electrical dysrhythmia were established in male SD rats by using a single-day diet and a double fasting method (drinking diluted hydrochloric acid water during the period) for 4 weeks. The gastric slow wave (GSW) recording, RT-qPCR, and western blot were performed to examine the effects of BXD on ICCs apoptosis in rats with GED and miR-451-5p expression. In vitro assays included CCK-8, flow cytometry analysis, RT-qPCR, and western blot were applied to investigate the potential molecular mechanism of BXD on ICCs apoptosis via miR-451-5p.
RESULTS: BXD promoted gastric motility, reduced ICCs apoptosis, and elevated miR-451-5p in GED rats. In addition, miR-451-5p was significantly up-regulated in ICCs after BXD treatment compared with that in ICCs with miR-451-5p inhibitor transfection. Meanwhile, high miR-451-5p expression with either BXD treatment or miRNA mimics enhanced ICCs proliferation and inhibit apoptosis. Moreover, overexpression of miR-451-5p can reverse G0/G1 arrest in ICCs by BXD treatment. Further, SCF and c-kit protein levels were detected to demonstrate that modulation of miR-451-5p by BXD treatment was involved in this signaling.
CONCLUSIONS: Through this
study, we demonstrated that BXD could promote ICCs proliferation and inhibit apoptosis via miR-451-5p and may involve the modulations of SCF/c-kit signaling, thus suggesting a new therapy basis for GI motility dysfunction from the perspective of modulation of ICCs apoptosis by targeting miR-451-5p.