脐带静脉内皮细胞诱导去分化重编排脂肪干细胞成骨分化的研究

Experimental study on the osteogenic differentiation of dedifferentiation reprogrammed adipose-derived stem cells induced by human umbilical vein endothelial cells

目的观察脐带静脉内皮细胞（HUVEC）诱导去分化重编排脂肪干细胞成骨分化的效果并探讨其分子机制。方法分离人脂肪干细胞（hASCs），取第3代hASCs进行去分化重编排并标记为去分化重编排脂肪干细胞（De．hASCs）。分离人HUVEC，将hASCs与HUVEC以1：1比例进行共培养（hASCs组），将De—hASCs与HUVEC以1：1比例进行共培养（De—hASCs组）。并于共培养4d和7d后检测碱性磷酸酶（ALP）活性，Runt相关基因2（Runx2）和骨形态发生蛋白-2（BMP-2）的mRNA表达水平，骨钙素的mRNA表达水平。结果共培养4d后，De—hASCs组的ALP活性高于hASCs组，但差异无统计学意义（P〉0．05）。共培养7d后，De—hASCs组ALP活性高于hASCs组，且差异有统计学意义（P〈0．05）。共培养4d和7d后，De—hASCs组Runx2和BMP一2的mRNA表达水平均明显高于hASCs组，且差异有统计学意义（P〈0．05）。共培养4d和7d后，De-hASCs组骨钙素的mRNA表达水平高于hASCs组，但差异无统计学意义（P〈0．05）。结论去分化重编排hASCs与HUVEC共培养条件下具有比普通脂肪干细胞更强的成骨分化能力，且与其上调BMP一2基因表达相关。

Objective To study the osteogenic differentiation and related molecular mechanisms of dedifferentiation reprogrammed hASCs （De-hASCs） induced by human umbilical vein endothelial cells （HUVECs）. Methods HASCs were isolated from 6 volunteers. HASCs passaged three times were then selected to undergo the dedifferentiation reprogrammed process. These cells were denoted as the De-hASCs. Both De-hASCs and hASCs were then co-cultured with HUVECs for a total period of one week. Alkaline phosphatase （ALP） activity of cells cultured for 4 and 7 days were measured, runt related tran- scription factor-2 （ Runx2 ） and bone morphogenetic protein-2 （BMP-2） mRNA levels of cells cultured for 4 and 7 days were detected. Osteocalcin mRNA expression levels of cells cultured for 4 and 7 days were ex- amined. Results ALP activity of the De-hASCs measured 4 days after cell culture was stronger than that of hASCs, but there was no significant difference. ALP acitivty of the De-hASCs measured 7 days after cell culture was stronger than that of hASCs with significant difference. Runx2 and BMP-2 mRNA expression levels of the De-hASCs were higher than those of hASCs both 4 and 7 days after cell culture with siginificant difference. Osteocalcin mRNA expression levels of De-hASCs were higher than those of hASCs both 4 and 7 days after cell culture but without significant difference. Conclusion When co-cultured with HUVECs, De-hASCs possessed a stronger osteogenic differentiation capacity than hASCs by up-regulating the expres- sion of BMP-2.