基于TGF-β1/Smad信号通路探讨加味茵芍散含药血清对大鼠肝星状细胞增殖作用的影响及机制

Discussion about the Influence of Modified Yinshao Pulvis Drug with Serum on the Proliferation of Rat Hepatic Stellate Cell Based on TGF- β1/Smad Signaling Pathway

目的基于转化生长因子β1(TGF-β1)/Smad通路探讨加味茵芍散含药血清对大鼠肝星状细胞(HSC)增殖的影响。方法选成年雄性Wistar大鼠随机分为加味茵芍散组、己酮可可碱组和生理盐水组,分别给予加味茵芍散、己酮可可碱、生理盐水灌胃,喂养1周后,予以腹主动脉采血,制备含药血清。设生理盐水组,己酮可可碱组,空白组,加味茵芍散低、中、高剂量组,含药血清干预HSC-T6后,用MTT法检测加味茵芍散对HSC-T6增殖的影响,并予以PCR、Western blot检测TGF-β1、Smad3 mRNA和蛋白表达情况。结果加味茵芍散高、中、低剂量组与生理盐水同等稀释倍数组相比较,加味茵芍散含药血清对HSC的增殖存在较为明显的抑制作用(P<0.05),在5%~15%的血清稀释倍数范围内,随剂量增大或给药时间的延长而抑制作用增强。加味茵芍散能有效抑制TGF-β1/Smad通路中TGF-β1、Smad3的mRNA和蛋白表达,与空白组和生理盐水组比较差异有统计学意义(P<0.05),且具有量效与时效关系。结论加味茵芍散可抑制HSC增殖,其机制可能与下调TGFβ1、Smad3的表达、阻断TGF-β1/Smad信号传导通路有关。

Objective To discuss the influence of modified Yinshao Pulvis drug with serum on the proliferation of rat hepatic stellate cell based on TGF- β1 / Smad signaling pathway. Methods Adult male Wistar rats were randomly divided into modified Yinshao Pulvis drug group,pentoxifylline group and hysiological saline group and given hexanone theobromine lavage,traditional Chinese medicine and physiological saline,adaptive feeding after a week,to prepare drug serum from the abdominal aorta in pursuance. Then the rats were divided into Chinese medicine high dose group( Modified Yinshao pulvis bulk drug serum),traditional Chinese medicine( TCM) middle dose group( Modified Yinshao pulvis bulk drug serum),traditional Chinese medicine( TCM) low dose group( Modified Yinshao pulvis bulk drug serum) and the negative control group( normal saline serum intervention) and positive control group( hexanone theobromine serum intervention) and blank group. After intervention of HSC- T6 by serum drug,used MTT method to detect the influence on the increase of HSC- T6 and PCR,Western blot to detect Smad3,TGF- beta 1 mRNA and protein expression. Results Compared Modified Yinshao pulvis high,medium and low dose group with multiple dilution physiological saline groups,the Modified Yinshao pulvis bulk drug serum had obvious inhibiting effect on the hepatic stellate cell proliferation( P < 0. 05). Within the scope of 5% ~ 15% serum dilution ratio,the inhibiting effect was enhanced with the increase of the dose or with the extension of using time. Modified Yinshao pulvis can effectively restrain TGF- beta 1,Smad3 mRNA and protein expression. Compared with thecontrol group and blank group,there was significantly statistical difference( P < 0. 05),and related to the time and dose. Conclusion Modified Yinshao pulvis inhibits HSC activation,The mechanism of anti- hepatic fibrosis may be related to the expression of TGF- beta 1 and Smad3,and blocks TGF- beta 1 / Smad signaling pathway.