目的 探索龙鳖胶囊含药血清对大鼠骨髓间充质干细胞（bone marrow stromal cells，BMSCs）迁移和Hippo-Yes相关蛋白（Yes-associated protein，YAP）信号通路的影响。方法 通过全骨髓贴壁法提取大鼠原代BMSCs，取第3代细胞，采用流式细胞术检测CD29、CD44、CD45和CD90的表达情况；利用成脂和成骨分化诱导剂对BMSCs进行诱导，成骨分化后采用茜素红染色观察形成矿化结节的情况，成脂分化后采用油红O染色观察脂滴的形成情况。SD大鼠ig龙鳖胶囊（0.625 g/kg），2次/d，连续7 d，腹主动脉取血，制备含药血清；使用YAP抑制剂Verteporfin（2、5 μmol/L）或5.0%含药血清处理BMSCs，采用划痕实验和Transwell迁移实验检测24 h后细胞的迁移情况。给予2.5%、5.0%含药血清处理BMSCs 24 h，采用Western blotting法检测Hippo-YAP信号通路相关蛋白的表达情况。结果 BMSCs分离培养后正常贴壁生长，呈长梭形；经成骨诱导茜素红染色后，可见大量被染成红色的矿化结节；经成脂诱导油红O染色后，可见大量被染成橘红色的脂滴；流式结果显示，表达CD29的阳性率为99.68%，表达CD44的阳性率为99.03%，表达CD45的阳性率为0.14%，CD90的阳性率为95.38%，符合BMSCs的特征。与对照组比较，Verteporfin显著抑制BMSCs迁移（P<0.05、0.01、0.001），5.0%含药血清显著促进BMSCs迁移（P<0.05）；与等剂量的Verteporfin组比较，5.0%含药血清＋Verteporfin组BMSCs迁移率明显上升（P<0.01、0.001）。2.5%、5.0%含药血清均显著下调大型肿瘤抑制因子2（large tumor suppressor homolog 2，LATS2）蛋白表达水平（P<0.05），上调YAP及其下游蛋白结缔组织生长因子（connective tissue growth factor，CTGF）蛋白表达水平（P<0.01）。结论 龙鳖胶囊含药血清具有促进BMSCs迁移的作用，其作用机制可能与调控Hippo-YAP信号通路相关。

Objective To explore the effect of Longbie Capsule (龙鳖胶囊) containing serum on migration of rat bone marrow mesenchymal stem cells (BMSCs) and Hippo-Yes-related protein (YAP) signaling pathway. Methods Primary rat BMSCs were extracted by whole bone marrow adherence method, and third generation cells were taken, expressions of CD29, CD44, CD45 and CD90 were detected by flow cytometry; BMSCs were induced with adipogenic and osteogenic differentiation inducers, alizarin red staining was used to observe the formation of mineralized nodules after osteogenic differentiation, oil red O staining was used to observe the formation of lipid droplets after adipogenic differentiation. SD rats were ig Longbie Capsule (0.625 g/kg), twice a day for 7 d, blood was taken from abdominal aorta to prepare drug-containing serum; YAP inhibitor Verteporfin (2, 5 μmol/L) or 5.0% drug-containing serum were used to treat BMSCs, scratch test and Transwell migration test were used to detect cell migration after 24 h. BMSCs were treated with 2.5% and 5.0% drug-containing serum for 24 h, Western blotting was used to detect the expressions of proteins related to Hippo-YAP signaling pathway. Results After isolation and culture, BMSCs grew normally adherently and showed a long spindle shape. After osteogenic induction of Alizarin Red staining, a large number of mineralized nodules were stained in red; After lipogenic induction oil red O staining, a large number of orange-red lipid droplets could be seen; Flow cytometry results showed that positive rate of CD29, CD44, CD45 and CD90 were 99.68%, 99.03%, 0.14% and 95.38%, which were in line with the characteristics of BMSCs. Compared with control group, Verteporfin significantly inhibited the migration of BMSCs (P < 0.05, 0.01, 0.001), 5.0% drug-containing serum significantly promoted the migration of BMSCs (P < 0.05); Compared with the corresponding dose of Verteporfin group, migration rate of BMSCs in 5.0% drug-containing serum + Verteporfin group was significantly increased (P < 0.01, 0.001). Both 2.5% and 5.0% drug-containing serum significantly down-regulated large tumor suppressor homolog 2 (LATS2) expression (P < 0.05), up-regulated expressions of YAP and its downstream protein connective tissue growth factor (connective tissue growth factor, CTGF) (P < 0.01). Conclusion Longbie capsule containing serum can promote the migration of BMSCs, and its mechanism may be related to the regulation of Hippo-YAP signaling pathway.

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国家自然科学基金资助项目（82004386）；国家自然科学基金资助项目（81873314）；国家自然科学基金资助项目（82004383）；国家自然科学基金资助项目（81974574）；国家自然科学基金资助项目（81473698）；广东省中医药局科研项目（20201129）；广东省普通高校重点科研平台和科研项目（2018KQNCX041）；广东省财政厅项目（[2014]157号，[2018]8号）；广东省中医院中医药科学技术研究专项（YN2019ML08，YN2015MS15，YK2013B2N19）；广东省医学科学技术研究基金资助项目（A2020105，B2019091）；广州市科技计划项目（202102010273）