Dataset Information

TGF?2-Hepcidin Feed-Forward Loop in the Trabecular Meshwork Implicates Iron in Glaucomatous Pathology.

ABSTRACT: Purpose:Elevated levels of transforming-growth-factor (TGF)-?2 in the trabecular meshwork (TM) and aqueous humor are associated with primary open-angle glaucoma (POAG). The underlying mechanism includes alteration of extracellular matrix homeostasis through Smad-dependent and independent signaling. Smad4, an essential co-Smad, upregulates hepcidin, the master regulator of iron homeostasis. Here, we explored whether TGF-?2 upregulates hepcidin, implicating iron in the pathogenesis of POAG. Methods:Primary human TM cells and human and bovine ex vivo anterior segment organ cultures were exposed to bioactive TGF-?2, hepcidin, heparin (a hepcidin antagonist), or N-acetyl carnosine (an antioxidant), and the change in the expression of hepcidin, ferroportin, ferritin, and TGF-?2 was evaluated by semiquantitative RT-PCR, Western blotting, and immunohistochemistry. Increase in reactive oxygen species (ROS) was quantified with dihydroethidium, an ROS-sensitive dye. Results:Primary human TM cells and bovine TM tissue synthesize hepcidin locally, which is upregulated by bioactive TGF-?2. Hepcidin downregulates ferroportin, its downstream target, increasing ferritin and iron-catalyzed ROS. This causes reciprocal upregulation of TGF-?2 at the transcriptional and translational levels. Heparin downregulates hepcidin, and reduces TGF-?2-mediated increase in ferritin and ROS. Notably, both heparin and N-acetyl carnosine reduce TGF-?2-mediated reciprocal upregulation of TGF-?2. Conclusions:The above observations suggest that TGF-?2 and hepcidin form a self-sustained feed-forward loop through iron-catalyzed ROS. This loop is partially disrupted by a hepcidin antagonist and an anti-oxidant, implicating iron and ROS in TGF-?2-mediated POAG. We propose that modification of currently available hepcidin antagonists for ocular use may prove beneficial for the therapeutic management of TGF-?2-associated POAG.

SUBMITTER: Ashok A

PROVIDER: S-EPMC7401420 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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TGFβ2-Hepcidin Feed-Forward Loop in the Trabecular Meshwork Implicates Iron in Glaucomatous Pathology.

Ashok Ajay A Chaudhary Suman S Kritikos Alexander E AE Kang Min H MH McDonald Dallas D Rhee Douglas J DJ Singh Neena N

Investigative ophthalmology & visual science 20200301 3

<h4>Purpose</h4>Elevated levels of transforming-growth-factor (TGF)-β2 in the trabecular meshwork (TM) and aqueous humor are associated with primary open-angle glaucoma (POAG). The underlying mechanism includes alteration of extracellular matrix homeostasis through Smad-dependent and independent signaling. Smad4, an essential co-Smad, upregulates hepcidin, the master regulator of iron homeostasis. Here, we explored whether TGF-β2 upregulates hepcidin, implicating iron in the pathogenesis of POAG ...[more]

PMID: 32182331

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Project description:Ocular hypertension is a causal risk-factor to developing glaucoma. This is associated with stiffening of the trabecular meshwork (TM), the primary site of resistance to aqueous-humor-outflow. The mechanisms underlying this stiffening or how pathologic extracellular matrix (ECM) affects cell function are poorly understood. It is recognized that mechanotransduction systems allow cells to sense and translate the intrinsic biophysical properties of ECM into intracellular signals to control gene transcription, protein expression, and cell behavior. Using an anterior segment perfusion model, we document that there are significantly more low flow regions that are much stiffer, and fewer high flow regions that are less stiff in glaucomatous TM (GTM) when compared to non-glaucomatous TMs (NTM). GTM tissue also has fewer cells overall when compared with NTM tissue. In order to study the role of pathologic ECM in glaucoma disease progression, we conducted studies using cell derived matrices (CDM). First, we characterized the mechanics, composition and organization of fibronectin in ECM deposited by GTM and NTM cells treated with glucocorticosteroids. Then, we determined that these GTM-derived ECM are able to induce stiffening of normal NTM cells, and alter their gene/protein expression to resemble that of a glaucomatous phenotype. Further, we demonstrate that GTM-derived ECM causes endoplasmic reticular stress in NTM. They also became resistant to being reorganized by these NTM cells. These phenomena were exacerbated by ECMs obtained from steroid treated glaucoma model groups. Collectively, our data demonstrates that CDMs represent a novel tool for the study of bidirectional interactions between TM cells and their immediate microenvironment.Statement of significanceExtracellular matrix (ECM) changes are prevalent in a number of diseases. The precise mechanisms by which changes in the ECM contribute to disease progression is unclear, primarily due to absence of appropriate models. Here, using glaucoma as a disease model, we document changes in cell derived matrix (CDM) and tissue mechanics that contribute to the pathology. Subsequently, we determine the effect that ECMs from diseased and healthy individuals have on healthy cell behaviors. Data emanating from this study demonstrate that CDMs are a potent tool for the study of cell-ECM interactions.

Dataset Information

TGF?2-Hepcidin Feed-Forward Loop in the Trabecular Meshwork Implicates Iron in Glaucomatous Pathology.

Publications

TGFβ2-Hepcidin Feed-Forward Loop in the Trabecular Meshwork Implicates Iron in Glaucomatous Pathology.

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