Cross-talk between miR-29 and transforming growth factor-betas in trabecular meshwork cells.

Glaucoma is second only to cataracts as the greatest blinding disorder worldwide, and it is estimated that it will affect 60.5 million people by 2010.1 The main risk factor for primary open-angle glaucoma (POAG) is elevated intraocular pressure (IOP), presumably due to changes in the conventional outflow pathway (trabecular meshwork/Schlemm's canal). Alterations in the composition of the extracellular matrix (ECM) of the trabecular meshwork (TM) are known to be associated with POAG and are believed to play an important role in the abnormal increase in outflow resistance that leads to elevated IOP in this disease.2,3 The transforming growth factor beta (TGFβ) subfamily of cytokines includes three isoforms, TGFβ1, β2, and β3, which regulate a wide range of essential cellular activities, including proliferation, differentiation, and ECM dynamics in many cell types.4–6 Because of their pivotal role in cell regulation, alteration of TGFβs is a characteristic of many diseases and pathologies,7–10 including glaucoma. TGFβ2 has been found to be elevated in the aqueous humor of glaucoma patients,11,12 and elevation of TGFβ1 has been associated with pseudoexfoliative glaucoma.13 Experiments in vitro and in vivo have shown that TGFβs might play an important role in the pathogenesis of the TM in glaucoma. For instance, TM cells treated with TGFβ2 showed senescence-associated changes14 and increased synthesis of several ECM components.15,16 In addition, perfusion of human anterior segments with TGFβ2 resulted in increased ECM and IOP,17 and overexpression of the active form of TGFβ2 increased IOP and reduced outflow facility in mice and rats.18 Similarly, TGFβ1 overexpression changed the morphology of the anterior segment of rat eyes and affected IOP.19 The miR-29 family of microRNAs (miRNAs) is composed of three highly similar ortologs (miR-29a, miR-29b, and miR-29c) that share identical seed sequences. The members of the miR-29 family are known to repress posttranscriptional expression of several mRNAs that encode proteins involved in fibrosis, including multiple ECM components such as collagens, fibrillins, and elastin.20–23 We have previously demonstrated that miR-29b negatively regulates the expression of genes involved in ECM synthesis and deposition in TM cells and that downregulation of miR-29b, under chronic oxidative stress conditions, contributes to an increase in expression of multiple ECM components.23 Some recent reports have suggested that downregulation of members of the miR-29 family by TGFβs might contribute to the fibrogenic effects of these cytokines. TGFβ1 has been implicated in the reduction of the levels of miR-29a observed in fibroblasts from systemic sclerosis patients, which is believed to contribute to increased expression of multiple collagen genes targeted by miR-29a.24 Similarly, TGFβ1 has been shown to induce a significant downregulation of miR-29a in proximal tubule cells leading to an increase in collagen IV.25 However, our knowledge about the interactions between miR-29 and TGFβs is still very limited. Therefore, we investigated the potential effects of TGFβ1 and TGFβ2 on the expression of the miR-29 family and evaluated whether alterations in miR-29 expression might contribute to the effects mediated by these cytokines on the expression of ECM genes in human TM cells. In addition, we analyzed whether miR-29 can, in turn, affect the expression of TGFβ1 and TGFβ2.