Abstract

In this study, it was aimed to analyze behavioral changes of adrenergic receptors (ARs) in first three passages and osteogenic/adipogenic differentiation of mesenchymal stem cells (MSCs) derived from placenta fetal membrane (FM) and bone marrow (BM). It was also aimed to evaluate effects of receptor blockade on differentiation. We obtained first three passages of MSCs from placenta and BM samples. For cell identification, the cells were analyzed by flow cytometry using CD34, CD45 and CD3, CD105 antibodies in each passage. The effects of propranolol and phenoxybenzamine at incremental doses were analyzed by MTT. In addition, cell cultures were separately maintained with the blockers or without after second passage. After each passage and differentiation, α1A, α1B, α2A, α2B, β1, β2, β3 AR-mRNA expressions analyzed by RT-qPCR technique. BMP6 and PPARG mRNA expressions only after differentiation and passage 3 were analyzed. A microscopic examination was also performed. Our results showed that AR expression behaviors were different in MSCs obtained from different tissue sources. In particular, α_1A-AR and α_2A-AR were expressed with considerably high coefficients in differentiation under blocker effect in BM-derived MSCs. No such coefficients were observed in any group of placental MSCs. In addition, it was found that the blockers stimulated adipogenesis in BM-derived MSCs during osteogenic differentiation. MSCs exhibit protein expressions that vary according to source of tissue and differentiation. Given that MSCs from different sources are used for repair and modulation, our study makes implications of this variable expression intriguing in the clinical practice.

摘要

在本研究中，旨在分析前三代肾上腺素受体 (ARs) 的行为变化以及源自胎盘胎膜 (FM) 和骨髓 (BM) 的间充质干细胞 (MSCs) 的成骨/成脂分化。它还旨在评估受体阻断对分化的影响。我们从胎盘和 BM 样本中获得了前三代 MSC。对于细胞鉴定，通过流式细胞术分析细胞，在每个传代中使用 CD34、CD45 和 CD3、CD105 抗体。通过 MTT 分析增加剂量的普萘洛尔和苯氧苯甲胺的作用。此外，细胞培养物在第二次传代后分别用或不用阻断剂维持。每次传代分化后，通过RT-qPCR技术分析α1A、α1B、α2A、α2B、β1、β2、β3 AR-mRNA的表达。仅在分化和第 3 代后分析 BMP6 和 PPARG mRNA 表达。还进行了显微镜检查。我们的结果表明，从不同组织来源获得的 MSCs 中的 AR 表达行为是不同的。特别是，α_1A -AR 和 α _2A -AR 在 BM 衍生的 MSC 中在阻断剂作用下以相当高的分化系数表达。在任何一组胎盘 MSCs 中都没有观察到这样的系数。此外，发现阻断剂在成骨分化过程中刺激了 BM 衍生的 MSCs 中的脂肪生成。MSCs 表现出根据组织来源和分化而变化的蛋白质表达。鉴于来自不同来源的 MSCs 用于修复和调节，我们的研究使这种可变表达的含义在临床实践中很有趣。