Effect of IGF-1 in the chondrogenesis of bone marrow mesenchymal stem cells in the presence or absence of TGF-β signaling
Effect of IGF-1 in the chondrogenesis of bone marrow mesenchymal stem cells in the presence or absence of TGF-β signaling
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DOI：
10.1359/jbmr.051213
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发表时间：
2006-04-01
影响因子：
6.2
通讯作者：
Spagnoli, A
中科院分区：
文献类型：
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作者：
Longobardi, L;O'Rear, L;Spagnoli, A
确定了胰岛素样生长因子 - I（IGF - I）在间充质干细胞（MSC）软骨形成中的一种新作用。通过条件性失活转化生长因子 - β II型受体，在有或无转化生长因子 - β信号的情况下评估了IGF - I的作用。我们发现IGF - I对MSCs具有强大的软骨诱导作用。IGF - I的作用独立于转化生长因子 - β，并且与转化生长因子 - β具有累加效应。
引言：间充质干细胞（MSCs）可从成年骨髓（BM）中分离出来，进行扩增，并分化为多种细胞类型，包括软骨细胞。IGF - I在MSCs软骨形成潜能中的作用知之甚少。转化生长因子 - β诱导MSC软骨分化，尽管其作用尚未明确界定。我们研究的目的是确定IGF - I在MSCs增殖、软骨凝聚、凋亡以及分化为软骨细胞过程中的生物学作用，包括单独作用以及与转化生长因子 - β联合作用，并且在有或无转化生长因子 - β信号的情况下。
材料和方法：从小鼠骨髓中分离出单核贴壁干细胞。通过在无血清和无胰岛素的特定培养基中培养高密度MSC团块长达7天，在有或无IGF - I和/或转化生长因子 - β的情况下诱导软骨分化。我们测量了胸苷掺入量，并对2天的团块用TUNEL、裂解的半胱天冬酶 - 3、花生凝集素和N - 钙黏蛋白进行染色。测量7天团块的大小，对蛋白聚糖合成进行染色，并通过实时定量PCR分析胶原蛋白II和Sox - 9的表达。我们从绿色荧光蛋白（GFP）位于胶原蛋白2启动子下的小鼠中获取MSCs，并通过共聚焦显微镜确定GFP的表达。我们使用cre - lox系统在MSCs中条件性失活转化生长因子 - β II型受体（TβRII），产生TβRII敲除的MSCs（RIIKO - MSCs）。
结果与结论：IGF - I通过刺激增殖、调节细胞凋亡以及诱导软骨细胞标志物的表达来调节MSC软骨形成。IGF - I的软骨诱导作用与转化生长因子 - β1同样强大，并且这两种生长因子具有累加效应。使用RIIKO - MSCs，我们表明IGF - I的软骨形成作用独立于转化生长因子 - β信号。我们发现细胞外信号相关激酶1/2丝裂原活化蛋白激酶（Erk1/2 MAPK）途径介导了转化生长因子 - β1的有丝分裂反应，并且在一定程度上介导了IGF - I的增殖作用。我们的数据通过展示IGF - I和转化生长因子 - β1在MSC软骨形成关键步骤中的作用，为优化MSCs在软骨疾病中的治疗应用提供了关键信息。
A novel role for IGF-I in MSC chondrogenesis was determined. IGF-I effects were evaluated in the presence or absence of TGF-beta signaling by conditionally inactivating the TGF-beta type II receptor. We found that IGF-I had potent chondroinductive actions on MSCs. IGF-I effects were independent from and additive to TGF-beta. Introduction: Mesenchymal stem cells (MSCs) can be isolated from adult bone marrow (BM), expanded, and differentiated into several cell types, including chondrocytes. The role of IGF-I in the chondrogenic potential of MSCs is poorly understood. TGF-beta induces MSC chondrogenic differentiation, although its actions are not well defined. The aim of our study was to define the biological role of IGF-I on proliferation, chondrogenic condensation, apoptosis, and differentiation of MSCs into chondrocytes, alone or in combination with TGF-beta and in the presence or absence of. TGF-beta signaling.Materials and Methods: Mononuclear adherent stem cells were isolated from mouse BM. Chondrogenic differentiation was induced by culturing high-density MSC pellets in serum- and insulin-free defined medium up to 7 days, with or without IGF-I and/or TGF-beta. We measured thymidine incorporation and stained 2-day-old pellets with TUNEL, cleaved caspase-3, peanut-agglutinin, and N-cadherin. Seven-day-old pellets were measured in size, stained for proteoglycan synthesis, and analyzed for the expression of collagen II and Sox-9 by quantitative real time PCR. We obtained MSCs from mice in which green fluorescent protein (GFP). was under the Collagen2 promoter and determined GFP expression by confocal microscopy. We conditionally inactivated the TGF-beta type II receptor (T beta RII) in MSCs using a cre-lox system, generating T beta RII knockout MSCs (RIIKO-MSCs).Results and Conclusions: IGF-I modulated MSC chondrogenesis by stimulating proliferation, regulating cell apoptosis, and inducing expression of chondrocyte markers. IGF-I chondroinductive actions were equally potent to TGF-beta 1, and the two growth factors had additive effects. Using RIIKO-MSCs, we showed that IGF-I chondrogenic actions are independent from the TGF-beta signaling. We found that the extracellular signal-related kinase 1/2 mitogen-activated protein kinase (Erk1/2 MAPK) pathway mediated the TGF-beta 1. mitogenic response and in part the IGF-I proliferative action. Our data, by showing the role of IGF-I and TGF-beta 1 in the critical steps of MSC chondrogenesis, provide critical information to optimize the therapeutic use of MSCs in cartilage disorders.