川芎嗪抑制大鼠气道平滑肌细胞增殖和诱导凋亡的研究

Study on inhibiting proliferation and inducing apoptosis of tetramethylpyrazine on rat airway smooth muscle cells

目的研究川芎嗪对大鼠气道平滑肌细胞(ASMCs)增殖和凋亡的影响。方法采用改良组织块和胰酶消化联合培养原代ASMCs,以血小板源生长因子(PDGF)诱导ASMCs增殖建立模型。采用MTT法检测不同浓度川芎嗪作用不同时间对ASMCs增殖的抑制,Hoechst33342染色检测细胞凋亡,Western blot检测ERK1/2蛋白表达情况。结果 MTT检测川芎嗪处理(6,12,24,36和48 h)后,与对照组比较,12.5μmol/L组、25μmol/L组、50μmol/L组和100μmol/L组的细胞平均抑制率均显著增加(P均<0.05)。Hoechst 33342染色观察到50μmol/L组、100μmol/L组和200μmol/L组川芎嗪中强荧光细胞比例随川芎嗪剂量增大而增多,细胞核内多个不均一蓝染现象。川芎嗪(200μmol/L组)和PDGF(20 pg/L)共同处理后,随作用时间增加,ERK1/2蛋白表达水平显著降低。结论川芎嗪对ASMCs增殖有抑制作用,同时高浓度的川芎嗪可促进ASMCs凋亡,可能与下调ERK1/2的表达有关。

Objective It is to investigate the influence of tetramethylpyrazine （TMP） on the proliferation and apoptosis of airway smooth muscle ceils （ASMCs） in rat. Methods Primary ASMCs of rats were cultured by improved tissue-piece digestion inoculation. A model of proliferation ASMCs was established by PDGF. The anti-proliferation effect to ASMCs of different doses and time of TMP was detected by MTT colorimetric assay. Cell apoptosis after treated with TMP was assessed by stainning with Hoechst 33342. The expression of ERK1/2 was detected by Western Blot. Results After 6, 12, 24, 36 and 48 hours incubated with TMP, compared with control group, the average cell inhibition rates of 12.5 μmol/L, 25 μmol/L, 50 μmol/L and 100 μmol/L group were increased significantly （P 〈 0.05 ）. Hyperfluoreseenee could be observed by Hoechst 33342 after 24 hours being treated with 50 μmol/L, 100 μmol/L and 200 μmol/L TMP, and the hyperfluorescenee was increased with the increasing dose of TMP. The levels of ERK1/2 protein were decreased significantly after treated with 200μmol/L TMP and 20 pg/L PDGF. Conclusion The proliferation of rat ASMCs is inhibited by TMP through down-regulate the expression of ERK1/ 2 and the high dose of TMP can induce apoptosis of ASMCs.