静脉血管平滑肌细胞中的多倍体细胞分析

目的探讨流式细胞术分析静脉血管平滑肌细胞多倍体细胞的方法和意义。方法选取普通级健康成年日本大耳白兔2只，雄性，8月龄，大约3 kg;普通级SD大鼠2只，雄性，8周龄，大约200 g。人大隐静脉为2例心脏冠状动脉旁路移植术中修补血管所得。在麻醉日本大耳白兔和大鼠后开腹腔获取下腔静脉，采用多种酶组合的消化液消化兔、大鼠以及人的静脉血管得到适合流式细胞检测的静脉血管平滑肌单细胞悬液，用碘化丙啶标记细胞，利用细胞流式仪分析兔、大鼠以及人的静脉血管平滑肌单细胞悬液各5 000个细胞，检测细胞DNA含量，DNA含量翻倍的细胞为多倍体细胞。并利用荧光原位杂交技术（FISH）验证阳性对照HEK293细胞系中的多倍体的存在。结果流式结果显示分析的5 000个细胞中，阳性对照HEK293细胞的多倍体细胞为1 355个（27.1%），从2例患者中取得的大隐静脉多倍体细胞为310个（6.2%）和250个（5.0%）;2只大鼠的下腔静脉平滑肌细胞中多倍体细胞为360个（7.2%）和450个（9%）;2只兔的下腔静脉平滑肌细胞中多倍体细胞分别为270个（5.4%）和305个（6.1%）。结论成年的静脉血管包括人大隐静脉的平滑肌细胞中普遍存在一定比例的多倍体细胞。

Detection of polyploid cells in venous vascular smooth muscle cells

ObjectiveTo confirm the polyploid cells in animal and human venous vein with the use of flow cytometry. MethodsThe animals were 2 healthy Japanese white rabbit （male, 8 months old, about 3 kg） and 2 SD rats （male, 8 weeks old, about 200 g）. 2 human saphenous veins were from the patients who received coronary artery bypass grafting operation. The rabbits and rats were anesthetized and the abdominal cavity was opened, then the inferior vena cava was obtained. We used multiple enzymes to digest animal and human vein and to get dispersed smooth muscle cells. The DNA was stained by propidium iodide （PI）, then 5 000 cells of rabbits were analyzed by flow cytometry. The confirmation of polyploid cells in tumor cell HEK293 by using Fluorescence in situ hybridization was as a positive control. ResultsThe percentage of polyploid cells of tumor cell HEK293 was 27.1%（1 355 cells）. The percentage of polyploid cells of vascular smooth muscle cells in human great saphenous vein was 5.0% and 6.2% ,respectively. The percentage of polyploid cell in vascular smooth muscle cells of inferior vena cava in two rats were 7.2% （360 cells）and 9% （450 cells） ,respectively. The percentage of polyploid cells of vascular smooth muscle cells in two rabbits were 5.4% （270 cells）and 6.1% （305 cells）, respectively. ConclusionVein blood vessel contains a certain proportion of polyploid cells in vascular smooth muscle cells, including human saphenous vein.