Coordinated regulation of extracellular matrix synthesis by the microRNA-29 family in the trabecular meshwork.
Coordinated regulation of extracellular matrix synthesis by the microRNA-29 family in the trabecular meshwork.
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DOI：
10.1167/iovs.10-6165
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发表时间：
2011-05
影响因子：
4.4
通讯作者：
G. Villarreal;Dong-Jin Oh;M. Kang;D. Rhee
中科院分区：
文献类型：
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作者：
G. Villarreal;Dong-Jin Oh;M. Kang;D. Rhee
目的 microRNA-29 (miR-29) 家族已在多种组织中成为细胞外基质 (ECM) 稳态的关键调节剂。在这项研究中，作者研究了 miR-29 家族在基础和 TGF-β2 刺激条件下调节小梁网 (TM) 中 ECM 合成的作用。方法 将人 TM 细胞与 2.5 ng/mL 活化的重组人 TGF-β2 一起孵育 24、48 和 72 小时。在 TGF-β2 刺激的情况下，使用特定的药物抑制剂来阻断 SMAD3 功能。通过实时 PCR 评估 miR-29 家族表达的变化。通过免疫印迹分析确定 miR-29 分子和抑制剂对 ECM 水平的影响。结果 miR-29 家族的所有三个成员均在培养的 TM 细胞中表达。尽管TM细胞与TGF-β2一起孵育可诱导miR-29a并抑制miR-29b水平，但未观察到对miR-29c表达的显着影响。其他研究表明，SMAD3 在基础条件和 TGF-β2 条件下调节 miR-29b 表达。随后的功能获得和丧失实验表明，miR-29 家族在基础和 TGF-β2 刺激条件下充当各种 ECM 蛋白的关键抑制因子。结论 本研究的结果表明 miR-29 家族是 TM 中 ECM 表达的关键调节因子，并表明 TGF-β2 对其的调节可能在控制 ECM 合成中发挥重要作用。总之，这些数据进一步深入了解了 TM 中介导 TGF-β2 信号传导和 ECM 产生的复杂调节机制。
PURPOSE The microRNA-29 (miR-29) family has emerged, in various tissues, as a key modulator of extracellular matrix (ECM) homeostasis. In this study, the authors investigate the role of the miR-29 family in the regulation of ECM synthesis in the trabecular meshwork (TM) under basal and TGF-β2 stimulatory conditions. METHODS Human TM cells were incubated with 2.5 ng/mL activated, recombinant human TGF-β2 for 24, 48, and 72 hours. A specific pharmacologic inhibitor was used to block SMAD3 function in the context of TGF-β2 stimulation. Changes in the expression of the miR-29 family were assessed by real-time PCR. The effect of miR-29 molecules and inhibitors on ECM levels was determined by immunoblot analysis. RESULTS All three members of the miR-29 family were expressed in cultured TM cells. Although the incubation of TM cells with TGF-β2 induced miR-29a and suppressed miR-29b levels, no significant effect was observed on miR-29c expression. Additional studies revealed that SMAD3 modulates miR-29b expression under basal and TGF-β2 conditions. Subsequent gain- and loss-of-function experiments demonstrated that the miR-29 family functions as a critical suppressor of various ECM proteins under basal and TGF-β2 stimulatory conditions. CONCLUSIONS The findings derived from this study identify the miR-29 family as a critical regulator of ECM expression in the TM and suggest that its modulation by TGF-β2 may be important in controlling ECM synthesis. Together, these data provide further insight into the complex regulatory mechanisms mediating TGF-β2 signaling and ECM production in the TM.