丹参含药血清调节大鼠肝星状细胞TGF-β1及细胞周期素CyclinD1的机制研究

Mechanism of Danshen drug-containing serum on TGF-β1 and CyclinD1 in hepatic stellate cells

目的:研究丹参含药血清调节大鼠肝星状细胞内转化生长因子β1(transforming growth factorβ1,TGF-β1)及细胞周期素CyclinD1的机制。方法:将20只Sprague-Dawley(SD)大鼠按照随机数字表法随机分为丹参低剂量组(1.04 g/kg)、中剂量组(2.08 g/kg)、高剂量组(4.17 g/kg)、空白组,并分别制备含药血清。体外传代培养肝星状细胞经10 ng/ml血小板衍生生长因子-BB(platelet-derived growth factor-BB,PDGF-BB)诱导后,再给予相应的含药血清干预,37℃培养24 h后收集细胞。经高效液相色谱法(HPLC)检测血清中丹参成分,q RT-PCR测定视黄酸受体-α(retinoic acid receptorα,RXR-α)及TGF-β1 mRNA,用Western blot法检测α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、Ⅰ型胶原蛋白(CollagenⅠ)及细胞周期素CyclinD1蛋白的表达情况。结果:丹参不同剂量处理组中,RXR-α mRNA表达量逐渐增加,而TGF-β1 mRNA、α-SMA蛋白、CollgenⅠ蛋白和CyclinD1蛋白表达逐渐降低,且与对照组比有统计学差异。结论:丹参含药血清可能通过促进RXR-α表达,抑制TGF-β1及CyclinD1表达,抑制肝星状细胞的增殖,进而抑制肝纤维化。

Objective：To investigate the mechanism of Danshen drug-containing serum on transforming growth factor β1（TGF-β1）and CyclinD1 in hepatic stellate cells. Methods： According to the random number table, 20 Sprague-Dawley（SD）rats were randomly divided into the Danshen（1.04 g/kg,2.08 g/kg,and 4.17 g/kg dosage）groups and normal control group. Blood was collected after gastric perfusion at 2 hours on 10 th day. The method of high performance liquid chromatography（HPLC）was used to detect the composition of Danshen in serum. HSCs were cultured in vitro and stimulated with platelet-derived growth factor-BB（PDGF-BB）,then were treated separately with the serum of Danshen（high,middle,and low dosage）,normal saline for 24 hours. After cells being collected,the expressions of retinoic acid receptor and TGF-β1 mRNA were measured by real-time PCR（q RT-PCR）,the content of α-smooth muscle actin（α-SMA）,typeⅠcollagen and CyclinD1 were detected by Western blot. Results：Compared with the normal control group,Danshen（low,middle,and high）groups could increase the expressions of retinoic acid receptor mRNA,and decrease the contents of TGF-β1 mRNA,α-SMA,type Ⅰ collagen and CyclinD1. Conclusion：Danshen may be achieved through increasing the expression of retinoic acid receptor and reducing TGF-β1 and CyclinD1 to inhibit the proliferation of HSCs,thus play a role of inhibition in liver fibrosis.