| 曹晓倩,董文珠,王璐,等.鳖甲煎丸对AngⅡ-ROS诱导下HSC-LX2细胞中PKC-Pyk2/SRC通路的影响[J].浙江中医药大学学报,2019,43(4):297-304. |
| 鳖甲煎丸对AngⅡ-ROS诱导下HSC-LX2细胞中PKC-Pyk2/SRC通路的影响 |
| Effect of Biejiajian Pill on PKC-Pyk2/SRC Pathway in HSC-LX2 Cell Induced by AngⅡ-ROS |
| DOI：10.16466/j.issn1005-5509.2019.04.001 |
| 中文关键词: 肝纤维化 HSC-LX2 AngⅡ ROS PKC Pyk2 SRC 鳖甲煎丸 |
| 英文关键词: liver fibrosis HSC-LX2 AngⅡ ROS PKC Pyk2 SRC Biejiajian pill |
| 基金项目:浙江省自然科学基金项目(LY16H280009)；国家自然科学基金项目(81173640)；浙江中医药大学预研项目(2016ZG11) |
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| 中文摘要: |
| [目的]探讨鳖甲煎丸含药血清对血管紧张素Ⅱ(angiotensinⅡ，AngⅡ)诱生的活性氧簇(reactive oxygen species，ROS)作用下HSC-LX2细胞中蛋白激酶C(protein kinase C，PKC)-富含脯氨酸的酪氨酸激酶2 (proline-rich tyrosine kinase 2，Pyk2)/SRC通路的影响。 [方法]以人源肝星状细胞HSC-LX2为研究对象，以AngⅡ诱导产生ROS，再以鳖甲煎丸含药血清进行干预，以MTT法检测HSC-LX2细胞的增殖情况，荧光探针DCFH-DA检测细胞内ROS含量，Western blot 检测PKC、Pyk2、SRC蛋白表达情况。[结果] MTT结果显示，与AngⅡ组比较，5%鳖甲煎丸含药血清培养24h后，鳖甲煎丸高、中剂量组显著抑制HSC-LX2细胞增殖(P＜0.05，P＜0.01)；培养48h后，鳖甲煎丸高、中、低剂量组均能显著抑制HSC-LX2细胞增殖(P＜0.05，P＜0.05，P＜0.01)；培养72h后，中剂量组有显著抑制效果(P＜0.01)。此外，除了15%中剂量鳖甲煎丸含药血清培养48h后有显著抑制效果(P＜0.05)，其余各组、各时间段均无明显抑制作用，差异无统计学意义(P＞0.05)。荧光探针DCFH-DA结果显示，与空白对照组比较，AngⅡ组ROS产生明显增多，经鳖甲煎丸含药血清高、中剂量组干预后，ROS均不同程度减少。Western blot显示，鳖甲煎丸含药血清干预后，PKC(P＜0.001，P＜0.001，P＜0.001)、Pyk2(P＜0.05， P＜0.001，P＜0.001)、SRC(P＜0.001，P＜0.001，P＜0.01)蛋白表达均受到显著抑制。[结论] HSC-LX2细胞经AngⅡ刺激后可以产生ROS，且细胞增殖明显，此过程与PKC、Pyk2、SRC蛋白的激活有关。鳖甲煎丸可通过减少ROS的产生来阻止HSC-LX2细胞的增殖，其机制与抑制PKC、Pyk2、SRC蛋白的表达有关。 |
| 英文摘要: |
| [Objective] To investigate the effect of serum containing Biejiajian pill on PKC-Pyk2/SRC pathway in HSC-LX2 cell under effect of reactive oxygen species(ROS) induced by angiotensin Ⅱ(AngⅡ). [Methods] Human hepatic stellate cell HSC-LX2 cell was used as research object, ROS was induced by AngⅡ, and serum containing Biejiajian pill was used for intervention. The proliferation of HSC-LX2 cell was detected by MTT assay. The intracellular ROS content was detected by fluorescent probe DCFH-DA, and the expression of PKC, Pyk2 and SRC protein was detected by Western blot. [Results] The MTT results showed that compared with the AngⅡgroup, the high and medium dose groups of Biejiajian pill significantly inhibited the proliferation of HSC-LX2 cell after cultured by 5% serum containing Biejiajian pill for 24 hours(P＜0.05, P＜0.01). After 48h, the high, medium and low dose groups could significantly inhibit the proliferation of HSC-LX2 cell(P＜0.05, P＜0.05, P＜0.01). After 72h, the medium dose group had significant inhibition(P＜0.01). In addition, there was a significant inhibitory effect of the 15% serum containing medium dose Biejiajian pill for 48h(P＜0.05), and no significant inhibition was observed in the other groups and time periods. The results of fluorescent probe DCFH-DA showed that compared with blank control group, the ROS production in AngⅡ group increased significantly. After the intervention of the high and medium dose serum containing Biejiajian pill, ROS decreased to varying degrees. Western blot results showed that the expression of PKC(P＜0.001, P＜0.001, P＜0.001), Pyk2(P＜0.05, P＜0.001, P＜0.001) and SRC(P＜0.001, P＜0.001, P＜0.01) was significantly inhibited after intervention with serum containing Biejiajian pill. [Conclusion] HSC-LX2 cell can produce ROS after stimulation with AngⅡ, and the cell proliferation is obvious. This process is related to the activation of PKC, Pyk2 and SRC proteins. Biejiajian pill can prevent the proliferation of HSC-LX2 cell by reducing the production of ROS, and its mechanism is related to the inhibition of PKC, Pyk2, SRC protein expression. |
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