Dataset Information

TGF?2-induced outflow alterations in a bioengineered trabecular meshwork are offset by a rho-associated kinase inhibitor.

ABSTRACT: Members of the transforming growth factor beta (TGF?) cytokine family have long been associated with affecting several cellular functions, including cell proliferation, differentiation and extracellular matrix (ECM) turnover. Of particular interest to this work, TGF?2 has been linked to most types of glaucomas as a potential fibrotic agent that can cause elevation of intraocular pressure (IOP). Given that the trabecular meshwork (TM) provides most of aqueous humor outflow resistance in the eye, an in vitro bioengineered human TM (HTM) model has been created and validated by analyzing effects of TGF?2 on transcellular pressure changes and outflow facility. These changes were correlated with several biological alterations induced by this cytokine, including ECM production and overexpression of HTM-marker myocillin. Furthermore, this TM model has been used to extend current knowledge of gene expression of cytokines involved in TGF?-induced ECM turnover over time. In particular, the ability for a ROCK-inhibitor to diminish the effect of TGF? on TM was demonstrated. This work supports the notion that anti-fibrotic activities of ROCK-inhibitors could counteract the elevation of IOP and increased strain observed in glaucomatous TM.

SUBMITTER: Torrejon KY

PROVIDER: S-EPMC5141429 | biostudies-other | 2016 Dec

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

TGFβ2-induced outflow alterations in a bioengineered trabecular meshwork are offset by a rho-associated kinase inhibitor.

Torrejon Karen Y KY Papke Ellen L EL Halman Justin R JR Bergkvist Magnus M Danias John J Sharfstein Susan T ST Xie Yubing Y

Scientific reports 20161207

Members of the transforming growth factor beta (TGFβ) cytokine family have long been associated with affecting several cellular functions, including cell proliferation, differentiation and extracellular matrix (ECM) turnover. Of particular interest to this work, TGFβ2 has been linked to most types of glaucomas as a potential fibrotic agent that can cause elevation of intraocular pressure (IOP). Given that the trabecular meshwork (TM) provides most of aqueous humor outflow resistance in the eye, ...[more]

PMID: 27924833

Similar Datasets

Project description:Purpose:Continuous reductions in trabecular meshwork (TM) cellularity inhibit aqueous humor (AH) outflow, which is the main cause of primary open-angle glaucoma. Rho-associated protein kinase inhibitor (ROCKi) targets the TM to reduce intraocular pressure (IOP) and increase AH outflow facility. However, the underlying mechanisms are not entirely clear. Here, we aimed to investigate the effect of a ROCKi (Y-27632) on TM cell proliferation and phagocytosis. Methods:Immortalized human TM (iHTM) cells, glaucomatous TM (GTM3) cells, and primary human TM (pTM) cells were cultured and identified. The effects of various concentrations of Y-27632 on F-actin cytoskeleton were assessed using immunofluorescence. Cell proliferation effects were evaluated using a cell counting kit-8 (CCK8), cell counting, and Ki67 immunostaining. Cell phagocytosis was evaluated using immunofluorescence and flow cytometry in immortalized TM cells. C57BL/6J and Tg-MYOC Y437H mice were used to investigate the proliferative effects of Y-27632 on TM cells in vivo. The effect of Y-27632 on IOP was monitored for 2 weeks, and the outflow facility was detected 2 weeks after IOP measurement. TM cells in mice were counted using immunohistochemistry. Results:Y-27632 (100 ?M) significantly promoted the proliferation of both immortal TM cells and pTM cells. In GTM3 cells, phagocytosis was significantly greater in the Y-27632 group than in the control group, nearly reaching the level of phagocytosis in iHTM, as determined using immunofluorescence and flow cytometry. In Tg-MYOC Y437H mice, treatment with Y-27632 significantly decreased IOP and increased outflow facility, which greatly influenced the long-term IOP-lowering effect. The number of TM cells in Tg-MYOC Y437H mice was significantly improved after Y-27632 administration. Conclusion:Y-27632 promoted cell proliferation and phagocytosis of TM cells, and its proliferative effect was demonstrated in a transgenic mouse model. These results revealed a new IOP-lowering mechanism of Y-27632 through effects on TM cells, suggesting the potential for a correlation between TM cellularity and long-term recovery of IOP.

Project description:Key pointsTrabecular meshwork (TM) is a highly mechanosensitive tissue in the eye that regulates intraocular pressure through the control of aqueous humour drainage. Its dysfunction underlies the progression of glaucoma but neither the mechanisms through which TM cells sense pressure nor their role in aqueous humour outflow are understood at the molecular level. We identified the Piezo1 channel as a key TM transducer of tensile stretch, shear flow and pressure. Its activation resulted in intracellular signals that altered organization of the cytoskeleton and cell-extracellular matrix contacts and modulated the trabecular component of aqueous outflow whereas another channel, TRPV4, mediated a delayed mechanoresponse. This study helps elucidate basic mechanotransduction properties that may contribute to intraocular pressure regulation in the vertebrate eye.AbstractChronic elevations in intraocular pressure (IOP) can cause blindness by compromising the function of trabecular meshwork (TM) cells in the anterior eye, but how these cells sense and transduce pressure stimuli is poorly understood. Here, we demonstrate functional expression of two mechanically activated channels in human TM cells. Pressure-induced cell stretch evoked a rapid increase in transmembrane current that was inhibited by antagonists of the mechanogated channel Piezo1, Ruthenium Red and GsMTx4, and attenuated in Piezo1-deficient cells. The majority of TM cells exhibited a delayed stretch-activated current that was mediated independently of Piezo1 by TRPV4 (transient receptor potential cation channel, subfamily V, member 4) channels. Piezo1 functions as the principal TM transducer of physiological levels of shear stress, with both shear and the Piezo1 agonist Yoda1 increasing the number of focal cell-matrix contacts. Analysis of TM-dependent fluid drainage from the anterior eye showed significant inhibition by GsMTx4. Collectively, these results suggest that TM mechanosensitivity utilizes kinetically, regulatory and functionally distinct pressure transducers to inform the cells about force-sensing contexts. Piezo1-dependent control of shear flow sensing, calcium homeostasis, cytoskeletal dynamics and pressure-dependent outflow suggests potential for a novel therapeutic target in treating glaucoma.

Dataset Information

TGF?2-induced outflow alterations in a bioengineered trabecular meshwork are offset by a rho-associated kinase inhibitor.

Publications

TGFβ2-induced outflow alterations in a bioengineered trabecular meshwork are offset by a rho-associated kinase inhibitor.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets