吴茱萸碱对瘢痕疙瘩成纤维细胞缝隙连接通讯及胶原合成的影响

Effect of evodiamine on the gap junction intercellular communication and synthesis of collagen in keloid

目的观察吴茱萸碱对瘢痕疙瘩成纤维细胞增殖和细胞间缝隙连接蛋白43（Cx43）蛋白以及胶原合成的影响。方法取手术切除瘢痕疙瘩组织成纤维细胞原代培养，将瘢痕疙瘩成纤维细胞分为空白对照组和用含不同浓度（0．5、1．0、2．0、4．0μmol／L）吴茱萸碱培养液处理的实验组，给药24、48h后噻唑蓝（MTT）法检测细胞的活性，酶联免疫吸附试验（ELISA）法检测不同浓度吴茱萸碱对瘢痕疙瘩成纤维细胞Cx43蛋白、Ⅰ和Ⅲ型胶原合成的影响。结果在0．5、1．0、2．0、4．0μmol／L实验组中24h细胞抑制率分别为6．2％、9．1％、12．5％、17．6％，48h细胞抑制率分别为10．9％、21．7％、33．4％、43．3％，显示吴茱萸碱显著抑制瘢痕疙瘩成纤维细胞增殖，且存在一定的时间和浓度依赖性；ELISA结果显示：48h后在0．5、1．0、2．0、4．0μmol／L实验组中Cx43蛋白吸光度（A）值分别为0．252、0．315、0．383、0．547；I型胶原A值分别为0．507、0．412、0．336、0．226；m型胶原A值分别为0．246、0．228、0．179、0．115；吴茱萸碱作用后瘢痕疙瘩成纤维细胞的细胞缝隙连接Cx43蛋白表达增强，Ⅰ、Ⅲ型胶原表达减弱。结论吴茱萸碱有效促进细胞缝隙连接Cx43蛋白表达，提高细胞缝隙连接通讯（GJIC）水平，抑制瘢痕疙瘩成纤维细胞增殖和Ⅰ、Ⅲ型胶原合成。

Objective To investigate the effect of evodiamine on the proliferation of fibroblasts, synthesis of collagen, and gap junction protein connexin 43 （Cx43） in keloid. Methods The cultured in vitro fibroblasts from patients with keloid were harvested and divided into a control group and evodiamine - treated （0. 5, 1.0, 2. 0, and 4. 0/xmol/L） groups. After treatment for 24, and 48 h, cell activity was de- termined by methyl thiazol tetrazolium （MTT）. The expression of gap junction protein Cx43, and systhesis of collagen type I and m in vitro after treatment were measured by enzyme linked immunosorbent assay （ELISA）. Results At 24 h cell inhibition rate was 6. 2%, 9. 1%, 12. 5%, and 17.6% in 0. 5, 1.0, 2. 0, 4.0 μmol/L evodiamine - treated groups, respectively. Evodiamine could significantly inhibit the growth of fibroblasts in vitro in a concentration - and time - dependent manner. ELISA results revealed that Cx43 protein absorbance （A） values were 0. 252, 0. 315, 0. 383 and 0. 547, those of collagen type Ⅰ were 0. 507, 0. 412, 0. 336 and 0. 226, and those of Collagen type Ⅲ were 0. 246, 0. 228, 0. 179 and 0. 115 in the 0. 5, 1.0, 2. 0, 4. 0 μmol/L evodiamine - treated experimental groups, respectively. The ex- pression of gap junction protein Cx43 was up - regulated, and the systhesis of collagen type Ⅰ and Ⅲ in vitro was decreased after evodiamine treatment. Conclusion Evodiamine can effectively promote the pro- duction of gap junction protein Cx43, increase the level of gap junction intercellular communication （GJIC） , inhibit the growth of fibroblasts and the production of collagen type Ⅰ and Ⅲ of keloid in vitro.