人大隐静脉与胸廓内动脉平滑肌细胞生长特性的比较

目的：探讨人大隐静脉（saphenous vein，SV）与胸廓内动脉（internal thoracic artery，ITA）血管平滑肌细胞（VSMCs）的原代培养及传代培养的方法，并比较二者的生长特性。方法：取冠状动脉搭桥术后剩余的SV与ITA血管标本，分为SV组与ITA组，用植块法进行VSMCs的原代培养，用免疫荧光染色法鉴定细胞，并比较两组细胞培养时间的差异。去血清培养48h后，在DMEM／F12、100ml／LFBS及10ng／ml血小板源性生长因子（PDGF）-BB不同培养条件下，观察SV的VSMCs与ITA的VSMCs生长情况。用MTY比色法检测细胞增殖并进行比较。结果：原代及传代培养均获成功。免疫荧光染色法显示平滑肌肌动蛋白（SMα-actin）位于细胞质，总阳性率〉95％。SV组原代培养植块周围细胞爬出时间为（9．1±1．1）d，大于ITA组的（5．8±1．0）d（P〈0．05）。sV的VSMCs再次传代培养时间为（34．9±3．4）d大于ITA的VSMCs首次传代培养时间（29．1±4．4）d（P〈0．05）。SV的VSMCs再次传代培养时间为（9．0±4．2）d，与ITA的VSMCs再次传代培养时间（9．6±3．9）d相似。两组细胞在相同培养条件时，未见明显的形态学差异，细胞生长曲线的差异无统计学意义。结论：采用植块法进行SV与ITA的VSMCs原代培养简便可行，细胞纯度高，具有良好的细胞表型转变特性，是研究冠状动脉搭桥术后血管桥再狭窄分子机制良好的细胞模型。SV的VSMCs可能较ITA的VSMCs具有更强的增殖潜能，二者内在属性的差异仍有待于进一步研究。

Comparison of growth characteristics of smooth muscle cells from saphenous vein and internal thoracic artery in in vitro culture

AIM： To explore primary culture and subculture methods of vascular smooth muscle cells （VSMCs） from human saphenous vein （SV） and internal thoracic artery （ITA） and to compare their growth characteristics in vitro. METHODS： SV and ITA tissues were obtained from patients undergoing coronary artery bypass grafting. VSMCs were primarily cultured by explant attached method and then were identified by immunofluorescence staining. Cell culture time of SV VSMCs and ITA VSMCs were compared and analyzed. After 48 h serum deprived culture, SV VSMC and ITA VSMC growth was observed under different conditions of DMEM/F12, 10% FBS and 10 ng/ml PDGF-BB and cell proliferations were compared by MTT assay. RESULTS： The primary and subculture were successfully completed. Positive staining of SMα-actin in the cytoplasm was shown by immunofluorescence （ positive rate of 〉 95 % ）. Primary cultured VSMC growth from the edge of SV tissue clumps after incubation was slower than that of ITA [（9.1±1.1） vs. （5.8±1.0） days, P〈0.05]. The first passage time ofSV VSMCs （34.9±3. d） days was longer than that of ITA VSMCs （29. 1 ±4. 4） days, P 〈 0. 05 and the repassage time of SV VSMCs （9.0 ±4. 2） days was similar to that of ITA VSMCs （9.6 ±3.9） days. No significant morphological difference was observed between SV VSMCs and ITA VSMCs under the same culture conditions and no significant difference was found in the relative cell growth curves. CONCLUSION： VSMC primary culture of SV and ITA by explant attached method is feasible. VSMCs in vivo present good cell phenotype transformation characteristics and can be used as a good cell model for molecular mechanism study of restenosis after coronary artery bypass surgery. SV VSMCs may have stronger proliferation potentials than ITA VSMCs. The intrinsic difference of VSMCs from SV and ITA still needs further study.