浙江中医药大学
纸质出版:2018
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[1]夏雪皎,黄棪,鲁军,杨东升,周凝,张俊杰.疏肝健脾活血方含药血清对肝纤维化模型大鼠肝窦内皮细胞失窗孔化的影响[J].中医杂志,2018,59(23):2037-2042.
XIA Xuejiao, HUANG Yan, LU Jun, et al. Effects of Serum Containing Shugan Jianpi Huoxue Decoction(疏肝健脾活血方) on Defenestration of Sinusoidal Endothelial in Hepatic Fibrosis Model Rats[J]. Journal of Traditional Chinese Medicine2018, 59(23): 2037-2042.
[1]夏雪皎,黄棪,鲁军,杨东升,周凝,张俊杰.疏肝健脾活血方含药血清对肝纤维化模型大鼠肝窦内皮细胞失窗孔化的影响[J].中医杂志,2018,59(23):2037-2042. DOI: 10.13288/j.11-2166/r.2018.23.015.
XIA Xuejiao, HUANG Yan, LU Jun, et al. Effects of Serum Containing Shugan Jianpi Huoxue Decoction(疏肝健脾活血方) on Defenestration of Sinusoidal Endothelial in Hepatic Fibrosis Model Rats[J]. Journal of Traditional Chinese Medicine2018, 59(23): 2037-2042. DOI: 10.13288/j.11-2166/r.2018.23.015.
目的探讨疏肝健脾活血方治疗肝纤维化的可能作用机制。方法 10只SD大鼠以10 ml/kg疏肝健脾方浓缩液(浓度2. 7 g/ml)灌胃
每日1次
连续3天后制备含药血清。中药血清设5%、10%、15%、20%、25%共5个浓度
采用MTT法筛选实验药物浓度。采用四氯化碳(CCl4)腹腔注射建立肝纤维化大鼠模型后分离肝纤维化肝窦内皮细胞(SEC)
同时集正常大鼠肝星状细胞(HSC)和SEC。实验分为正常组、模型组、DAPT组、NF组、中药组、中药加DAPT组、中药加NF组
共7组。各组均加入HSC
正常组加入正常大鼠SEC
其余组加入肝纤维化大鼠SEC
常规培养24 h。吸出上清液
DAPT组、中药加DAPT组加入0. 2μmol/L DAPT (Notch通路阻断剂)
NF组、中药加NF组加入1 gsu/ml rh NF-κB (Notch通路激动剂)。前4组加入含10%FBS DMEM/12培养基
后3组加入含预筛选浓度中药血清、10%FBS DMEM/12培养基
培养72 h后MTT法检测各组HSC活力
Western Blot法检测SEC中CD31、vWF蛋白表达情况。结果筛选出10%浓度的中药血清对细胞增殖有抑制作用。与正常组比较
造模2、4、8周模型组HSC活力上升
造模4、8周CD31蛋白表达升高(P <0. 05或P <0. 01);与模型组比较
NF组HSC活力上升
DAPT组和中药组HSC活力、CD31、vWF蛋白表达下降(P <0. 05或P <0. 01);与NF组比较
中药加NF组HSC活力、CD31、vWF蛋白表达下降(P <0. 05或P <0. 01);与DAPT组比较
中药加DAPT组HSC活力、CD31、vWF蛋白表达均下降(P <0. 05或P <0. 01)。结论疏肝健脾活血方可通过干预Notch通路
抑制HSC活力及SEC失窗孔化
可能是其治疗肝纤维化的作用机制之一。
Objective To explore the possible mechanism of Shugan Jianpi Huoxue Decoction(疏肝健脾活血方)(STHD) in treating hepatic fibrosis.Methods Ten SD rats were orally administered with 10 ml/kg Shugan Jianpi Huoxue Decoction(concentration 2.7 g/ml) once a day for 3 consecutive days to prepare drug-containing serum.The serum of traditional Chinese medicine is set at 5 concentrations of 5%
10%
15%
20% and 25%.The concentration of the experimental drug was screened by MTT method
and the liver fibrosis sinusoidal endothelial cells(SEC)were isolated by intraperitoneal injection of carbon tetrachloride(CCl4).The normal rat hepatic stellate cells(HSC)and SEC were collected.In this experiment
the rats were divided into 7 groups including normal group
model group
DAPT group
NF group
Chinese medicine group
Chinese medicine plus DAPT group and Chinese medicine plus NF group.HSC was added to each group
and SEC of normal rats was added to the normal group
and SEC of liver fibrosis rats were added to the other groups for 24 hours.The supernatant was aspirated
and the DAPT group
the Chinese medicine plus DAPT group were added with Notch pathway blocker DAPT 0.2 μmol/L
and the NF group and the Chinese medicine plus NF group were added with Notch pathway agonist rhNF-κB 1 gsu/ml.The first4 groups were added with 10% FBS DMEM/12 medium
and the last 3 groups were added with pre-screened concentration Chinese medicine serum
10% FBS DMEM/12 medium.After 72 hours of culture
the MTC activity of each group was detected by MTT method
and the expression of CD31 and vWF proteins in SEC was detected by Western Blot method.Results The screening out 10% concentration of traditional Chinese medicine in serum had an inhibitory effect on cell proliferation.Compared with the normal group
the HSC activity of the model group was increased after 2
4
and 8 weeks
and the CD31 protein expression was increased after 4 and 8 weeks since model establishment(P<0.05 or P<0.01).Compared with the model group
the HSC activity of the NF group was increased.The HSC activity
CD31 and vWF protein expression were decreased in DAPT group and Chinese medicine group(P <0.05 or P<0.01).Compared with the NF group
the HSC activity
CD31 and vWF protein expression decreased in traditional Chinese medicine plus NF group(P<0.05 or P<0.01).Compared with the DAPT group
the expression of HSC
CD31 and v WF protein in Chinese medicine plus DAPT group were decreased(P<0.05 or P<0.01).Conclusion Shugan Jianpi Huoxue Decoction could inhibit HSC activity and the defenestration of SEC by interfering plu Notch pathway
which may be one of the mechanisms of its treatment of liver fibrosis.
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