溃结灵含药血清对大鼠结肠上皮细胞损伤的影响

Effect of Kuijieling medicated serum on the rat colonic epithelial cell damage model

目的:探讨溃结灵含药血清对TNF-α引起的大鼠结肠上皮细胞损伤模型的影响。方法:采用本实验室建立的方法进行大鼠结肠上皮细胞(CEC)的分离及原代培养,MTT法观察含药血清(10%,5%,2.5%)对CEC增殖的影响,分别以25,50,100,150,200ng/ml的TNF-α干预CEC,测定不同干预时间(3h,6h,24h)细胞的增殖情况,WesternBlot法检测50ng/ml TNF-α干预6h后结肠上皮细胞pMEK1/2,pERK1/2蛋白表达水平。结果:空白大鼠血清对CEC生长无影响,柳氮磺(10%含药血清)及溃结灵中剂量(5%含药血清)作用24h、48h均明显促进CEC增殖(P<0.05)。50ng/ml TNF-α干预CEC 6h后可明显抑制细胞增殖,造成细胞损伤模型,含药血清各组对细胞损伤有一定的促增殖作用。50ng/ml TNF-α作用于CEC6h后细胞内pMEK1/2、pERK1/2蛋白表达均有增加,溃结灵高剂量(10%含药血清)、中剂量(5%含药血清)明显促进pMEK1/2蛋白表达(P<0.05),溃结灵中剂量(5%含药血清)及柳氮磺(10%含药血清)明显促进pERK1/2蛋白表达(P<0.05)。结论:溃结灵含药血清可促进CEC增殖,上调TNF-ɑ引起的损伤细胞模型pMEK1/2,pERK1/2表达,可能进一步激发细胞内MEK/ERK信号通路,进而促进细胞增殖效应。

To explore the effects of Kuijieling Decoction medicated serum on the cell damage model of primary rat colonic epithelial ceils （CECs）. Methods： Cell separation and culture were made in our lab. MTT assay was used to determine ceil gyowth of CECs under the effects of Knijieling Decoction medicated serum（ 10% ,5% ,2.5% ）. CECs were treated with different concentration of TNF-a（25,50,100, 150,200ng/mL） for 3h, 6h and 24h. The proliferous activity and protein expression of pMEK1/2 and pERK1/2 in CECs were measured after intervened with 50ng/mL TNF-afor 6h. Results： 1. There was no effect on the CECs proliferation with rat serum without medicine. And the CECs proliferation was promoted significantly after intervened with Liudanhuang（ 10% Decoction medicated serum）and Kuijieling（5% Decoc- tion medicated serum） for 24h and 48h, （ P 〈 0.05 ） ;2. The CECs proliferation was inhibited significantly after intervened with 50ng/ml TNF-u for 6h, and ceil damage model was made by this method. Different concentration of medicated sermn could stimulate colonic epithelial ceils to reproduce in a certain degree after damaged by TNF-a; 3. The protein expression of pMEK1/2,pEtLK1/2 in the CECs induced by TNF-a for 6h were obviously increased. And the protein expression of pMEK1/2 in the medium （5% Decoction medicated serum） and high （ 10% Decoction medicated serum） groups of Kuijieling were obviously higher than that in m（xtel group（ P 〈 0.05 ）, liudanhuang（ 10% De- coction medicated serum） and Kuijieling （5% Decoction medicated serum） could enhance the protein expression of pERK1/2 significantly（ P 〈0.05）. Conclusion： Kuijieling Decoction medicated senun could promote CECs proliferation and up-regulated the protein expression of pMEK1/2,pERK1/2 in the CEC damage model induced by TNF-a. This maybe further activate the intraceilular MEK/ERK Signal Pathway to promote the CECs proliferation.