Park Yoon Shin, Hwang Soojin, Jin Yoon Mi, Yu Yeonsil, Jung Sung-Ae, Jung Sung-Chul, Ryu Kyung-Ha, Kim Han Su, Jo Inho
Department of Molecular Medicine, School of Medicine, and Global Top 5 Research Program, Ewha Womans University, Mok-6-dong, Yangcheon-gu, Seoul 158-710, Republic of Korea.
J Cell Physiol. 2015 Jan;230(1):140-9. doi: 10.1002/jcp.24690.
CCN1 is highly expressed in cancer cells and has been identified in the secretome of bone marrow-derived mesenchymal stem cells (BM-MSC). Although secreted CCN1 is known to promote angiogenesis, its underlying mechanism remains unclear. Here, we examined whether our recently-established tonsil-derived MSC (T-MSC) secrete CCN1 and, if any, how CCN1 promotes the angiogenesis of human umbilical vein endothelial cells (HUVEC). Compared with untreated control T-MSC, a higher level of CCN1 was secreted by T-MSC treated with activin A and sonic hedgehog, drugs known to induce endodermal differentiation. Expectedly, conditioned medium collected from differentiated T-MSC (DCM) significantly increased HUVEC migration and tube formation compared with that from control T-MSC (CCM), and these stimulatory effects were reversed by neutralization with anti-CCN1 antibody. Treatment with recombinant human CCN1 (rh-CCN1) alone also mimicked the stimulatory effects of DCM. Furthermore, treatment with either DCM or rh-CCN1 increased the phosphorylation of AMP kinase (AMPK), and ectopic expression of siRNA of the AMPK gene inhibited all observed effects of both DCM and rh-CCN1. However, no alteration of intracellular ATP levels or phosphorylation of LKB1, a well-known upstream factor of AMPK activation, was observed under our conditions. Finally, the neutralization of integrin α(v) β(3) with anti-integrin α(v) β(3) antibody almost completely reversed the effects of CCN1 on AMPK phosphorylation, and EC migration and tube formation. Taken together, we demonstrated that T-MSC increase the secretion of CCN1 in response to endodermal differentiation and that integrin α(v) β(3) and AMPK mediate CCN1-induced EC migration and tube formation independent of intracellular ATP levels alteration.
CCN1在癌细胞中高表达，并且已在骨髓间充质干细胞（BM-MSC）的分泌蛋白组中被鉴定出来。尽管已知分泌的CCN1可促进血管生成，但其潜在机制仍不清楚。在此，我们研究了我们最近建立的扁桃体来源的间充质干细胞（T-MSC）是否分泌CCN1，以及如果分泌的话，CCN1如何促进人脐静脉内皮细胞（HUVEC）的血管生成。与未处理的对照T-MSC相比，用激活素A和音猬因子处理的T-MSC分泌的CCN1水平更高，激活素A和音猬因子是已知可诱导内胚层分化的药物。不出所料，与对照T-MSC（CCM）相比，从分化的T-MSC收集的条件培养基（DCM）显著增加了HUVEC的迁移和管形成，并且用抗CCN1抗体中和可逆转这些刺激作用。单独用重组人CCN1（rh-CCN1）处理也模拟了DCM的刺激作用。此外，用DCM或rh-CCN1处理均可增加AMP激酶（AMPK）的磷酸化，并且AMPK基因的siRNA异位表达抑制了DCM和rh-CCN1的所有观察到的作用。然而，在我们的条件下未观察到细胞内ATP水平的改变或AMPK激活的著名上游因子LKB1的磷酸化。最后，用抗整合素α(v)β(3)抗体中和整合素α(v)β(3)几乎完全逆转了CCN1对AMPK磷酸化、内皮细胞迁移和管形成的作用。综上所述，我们证明T-MSC响应内胚层分化而增加CCN1的分泌，并且整合素α(v)β(3)和AMPK介导CCN1诱导的内皮细胞迁移和管形成，而与细胞内ATP水平的改变无关。
