In vitro comparison of human and murine trabecular meshwork cells: implications for glaucoma research.
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
23 Sep 2024
23 Sep 2024
Historique:
received:
22
03
2024
accepted:
12
09
2024
medline:
24
9
2024
pubmed:
24
9
2024
entrez:
23
9
2024
Statut:
epublish
Résumé
The trabecular meshwork (TM) is crucial for regulating intraocular pressure (IOP), and its dysfunction significantly contributes to glaucoma, a leading cause of vision loss and blindness worldwide. Although rodents are commonly used as animal models in glaucoma research, the applicability of these findings to humans is limited due to the insufficient understanding of murine TM. This study aimed to compare primary human TM (hTM) and murine TM (mTM) cells in vitro to enhance the robustness and translatability of murine glaucoma models. In this in vitro study, we compared primary hTM and mTM cells under simulated physiological and pathological conditions by exposing both cell types to the glucocorticoid dexamethasone (DEX) and Transforming Growth Factor β (TGFB2), both of which are critical in the pathogenesis of several ophthalmological diseases, including glaucoma. Phagocytic properties were assessed using microbeads. Cells were analyzed through immunocytochemistry (ICC) and Western blot (WB) to evaluate the expression of extracellular matrix (ECM) components, such as Fibronectin 1 (FN1) and Collagen IV (COL IV). Filamentous-Actin (F-Act) staining was used to analyze cross-linked actin network (CLAN) formation. Additionally, we evaluated cytoskeletal components, including Vimentin (VIM), Myocilin (MYOC), and Actin-alpha-2 (ACTA2). Our results demonstrated significant similarities between human and murine TM cells in basic morphology, phagocytic properties, and ECM and cytoskeletal component expression under both homeostatic and pathological conditions in vitro. Both human and murine TM cells exhibited epithelial-to-mesenchymal transition (EMT) after exposure to DEX or TGFB2, with comparable CLAN formation observed in both species. However, there were significant differences in FN1 and MYOC induction between human and murine TM cells. Additionally, MYOC expression in hTM cells depended on fibronectin coating. Our study suggests that murine glaucoma models are potentially translatable to human TM. The observed similarities in ECM and cytoskeletal component expression and the comparable EMT response and CLAN formation support the utility of murine models in glaucoma research. The differences in FN1 and MYOC expression between hTM and mTM warrant further investigation due to their potential impact on TM properties. Overall, this study provides valuable insights into the species-specific characteristics of TM and highlights opportunities to refine murine models for better relevance to human glaucoma.
Identifiants
pubmed: 39313534
doi: 10.1038/s41598-024-73057-9
pii: 10.1038/s41598-024-73057-9
doi:
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
22002Informations de copyright
© 2024. The Author(s).
Références
Quigley, H. A. & Broman, A. T. The number of people with glaucoma worldwide in 2010 and 2020. Br. J. Ophthalmol.90, 262–267 (2006).
pubmed: 16488940
pmcid: 1856963
doi: 10.1136/bjo.2005.081224
Hollows, F. C. & Graham, P. A. Intra-ocular pressure, glaucoma, and glaucoma suspects in a defined population. Br. J. Ophthalmol.50, 570–586 (1966).
pubmed: 5954089
pmcid: 506274
doi: 10.1136/bjo.50.10.570
Acott, T. S. & Kelley, M. J. Extracellular matrix in the trabecular meshwork. Exp. Eye Res.86, 543–561 (2008).
pubmed: 18313051
pmcid: 2376254
doi: 10.1016/j.exer.2008.01.013
Johnson, M. & Erickson, K. Mechanisms and routes of aqueous humor drainage. In Principles and Practice of Ophthalmology 2577–2595WB (Saunders Co., 2000).
Weinreb, R. N., Aung, T. & Medeiros, F. A. The pathophysiology and treatment of glaucoma: a review. JAMA311, 1901–1911 (2014).
pubmed: 24825645
pmcid: 4523637
doi: 10.1001/jama.2014.3192
Costagliola, C. et al. How many aqueous humor outflow pathways are there? Surv. Ophthalmol.65, 144–170 (2020).
pubmed: 31622628
doi: 10.1016/j.survophthal.2019.10.002
Tamm, E. R. The trabecular meshwork outflow pathways: structural and functional aspects. Exp. Eye Res.88, 648–655 (2009).
pubmed: 19239914
doi: 10.1016/j.exer.2009.02.007
Brubaker, R. F. Measurement of uveoscleral outflow in humans. J. Glaucoma10, S45–S48 (2001).
pubmed: 11890274
doi: 10.1097/00061198-200110001-00017
Johnson, M. What controls aqueous humour outflow resistance? Exp. Eye Res.82, 545–557 (2006).
pubmed: 16386733
pmcid: 2892751
doi: 10.1016/j.exer.2005.10.011
Agarwal, P. & Agarwal, R. Trabecular meshwork ECM remodeling in glaucoma: could RAS be a target? Expert Opin. Ther. Targets22, 629–638 (2018).
pubmed: 29883239
doi: 10.1080/14728222.2018.1486822
Bermudez, J. Y., Montecchi-Palmer, M., Mao, W. & Clark, A. F. Cross-linked actin networks (CLANs) in glaucoma. Exp. Eye Res.159, 16–22 (2017).
pubmed: 28238754
pmcid: 5499152
doi: 10.1016/j.exer.2017.02.010
Lazarides, E. Actin, alpha-actinin, and tropomyosin interaction in the structural organization of actin filaments in nonmuscle cells. J. Cell Biol.68, 202–219 (1976).
pubmed: 1107334
doi: 10.1083/jcb.68.2.202
Osborn, M., Born, T., Koitsch, H. J. & Weber, K. Stereo immunofluorescence microscopy: I. Three-dimensional arrangement of microfilaments, microtubules and tonofilaments. Cell14, 477–488 (1978).
pubmed: 357010
doi: 10.1016/0092-8674(78)90234-9
Clark, A. F. et al. Glucocorticoid-induced formation of cross-linked actin networks in cultured human trabecular meshwork cells. Investig. Ophthalmol. Vis. Sci.35, 281–294 (1994).
Clark, A. F., Miggans, S. T., Wilson, K., Browder, S. & McCartney, M. D. Cytoskeletal changes in cultured human glaucoma trabecular meshwork cells. J. Glaucoma4, 183–188 (1995).
pubmed: 19920666
doi: 10.1097/00061198-199506000-00007
Clark, A. F. & Wordinger, R. J. The role of steroids in outflow resistance. Exp. Eye Res.88, 752–759 (2009).
pubmed: 18977348
doi: 10.1016/j.exer.2008.10.004
Clark, A. F. et al. Dexamethasone alters F-actin architecture and promotes cross-linked actin network formation in human trabecular meshwork tissue. Cell. Motil. Cytoskeleton60, 83–95 (2005).
pubmed: 15593281
doi: 10.1002/cm.20049
O’Reilly, S. et al. Inducers of cross-linked actin networks in trabecular meshwork cells. Investig. Ophthalmol. Vis. Sci.52, 7316–7324 (2011).
doi: 10.1167/iovs.10-6692
Gardel, M. L. et al. Elastic behavior of cross-linked and bundled actin networks. Science304, 1301–1305 (2004).
pubmed: 15166374
doi: 10.1126/science.1095087
Peng, M. et al. Cross-linked actin networks (CLANs) affect stiffness and/or actin dynamics in transgenic transformed and primary human trabecular meshwork cells. Exp. Eye Res.220, 109097 (2022).
pubmed: 35569518
pmcid: 11029344
doi: 10.1016/j.exer.2022.109097
Polansky, J. R., Weinreb, R. N., Baxter, J. D. & Alvarado, J. Human trabecular cells. I. Establishment in tissue culture and growth characteristics. Investig. Ophthalmol. Vis. Sci.18, 1043–1049 (1979).
Rohen, J. W., Schachtschabel, O. O. & Matthiessen, P. F. In vitro studies on the trabecular meshwork of the primate eye. Albrecht Von Graefes Arch. Klin. Exp. Ophthalmol. Albrecht Von Graefes Arch. Clin. Exp. Ophthalmol.193, 95–107 (1975).
doi: 10.1007/BF00419354
Mao, W., Liu, Y., Wordinger, R. J. & Clark, A. F. A magnetic bead-based method for mouse trabecular meshwork cell isolation. Investig. Ophthalmol. Vis. Sci.54, 3600–3606 (2013).
doi: 10.1167/iovs.13-12033
Overby, D. R. et al. Ultrastructural changes associated with dexamethasone-induced ocular hypertension in mice. Investig. Ophthalmol. Vis. Sci.55, 4922–4933 (2014).
doi: 10.1167/iovs.14-14429
van Zyl, T. et al. Cell atlas of aqueous humor outflow pathways in eyes of humans and four model species provides insight into glaucoma pathogenesis. Proc. Natl. Acad. Sci. USA117, 10339–10349 (2020).
pubmed: 32341164
pmcid: 7229661
doi: 10.1073/pnas.2001250117
Whitlock, N. A., McKnight, B., Corcoran, K. N., Rodriguez, L. A. & Rice, D. S. Increased intraocular pressure in mice treated with dexamethasone. Investig. Ophthalmol. Vis. Sci.51, 6496–6503 (2010).
doi: 10.1167/iovs.10-5430
Zode, G. S. et al. Ocular-specific ER stress reduction rescues glaucoma in murine glucocorticoid-induced glaucoma. J. Clin. Investig.124, 1956–1965 (2014).
pubmed: 24691439
pmcid: 4001532
doi: 10.1172/JCI69774
Patel, G. C. et al. Dexamethasone-induced ocular hypertension in mice: effects of myocilin and route of administration. Am. J. Pathol.187, 713–723 (2017).
pubmed: 28167045
pmcid: 5397678
doi: 10.1016/j.ajpath.2016.12.003
Tripathi, R. C., Li, J., Chan, W. F. & Tripathi, B. J. Aqueous humor in glaucomatous eyes contains an increased level of TGF-beta 2. Exp. Eye Res.59, 723–727 (1994).
pubmed: 7698265
doi: 10.1006/exer.1994.1158
Gajda-Deryło, B. et al. Comparison of cytokine/chemokine levels in aqueous humor of primary open-angle glaucoma patients with positive or negative outcome following trabeculectomy. Biosci. Rep.39, BSR20181894 (2019).
pubmed: 30967499
pmcid: 6499456
doi: 10.1042/BSR20181894
Shepard, A. R. et al. Adenoviral gene transfer of active human transforming growth factor-{beta}2 elevates intraocular pressure and reduces outflow facility in rodent eyes. Investig. Ophthalmol. Vis. Sci.51, 2067–2076 (2010).
doi: 10.1167/iovs.09-4567
Tamm, E. R., Russell, P. & Piatigorsky, J. Development of characterization of a immortal and differentiated murine trabecular meshwork cell line. Investig. Ophthalmol. Vis. Sci.40, 1392–1403 (1999).
Binter, M. et al. A simple dissection method for the isolation of mouse trabecular meshwork cells. PLoS One18, e0296124 (2023).
pubmed: 38128042
pmcid: 10734917
doi: 10.1371/journal.pone.0296124
Keller, K. E. et al. Consensus recommendations for trabecular meshwork cell isolation, characterization and culture. Exp. Eye Res.171, 164–173 (2018).
pubmed: 29526795
pmcid: 6042513
doi: 10.1016/j.exer.2018.03.001
Gasiorowski, J. Z. & Russell, P. Biological properties of trabecular meshwork cells. Exp. Eye Res.88, 671–675 (2009).
pubmed: 18789927
doi: 10.1016/j.exer.2008.08.006
Begley, C. G., Yue, B. Y. & Hendricks, R. L. Murine trabecular meshwork cells in tissue culture. Curr. Eye Res.10, 1015–1030 (1991).
pubmed: 1782800
doi: 10.3109/02713689109020340
Matsumoto, Y. & Johnson, D. H. Dexamethasone decreases phagocytosis by human trabecular meshwork cells in situ. Investig. Ophthalmol. Vis. Sci.38, 1902–1907 (1997).
Wordinger, R. J. et al. Effects of TGF-beta2, BMP-4, and gremlin in the trabecular meshwork: implications for glaucoma. Investig. Ophthalmol. Vis. Sci.48, 1191–1200 (2007).
doi: 10.1167/iovs.06-0296
Takahashi, E., Inoue, T., Fujimoto, T., Kojima, S. & Tanihara, H. Epithelial mesenchymal transition-like phenomenon in trabecular meshwork cells. Exp. Eye Res.118, 72–79 (2014).
pubmed: 24291802
doi: 10.1016/j.exer.2013.11.014
Yemanyi, F., Baidouri, H., Burns, A. R. & Raghunathan, V. Dexamethasone and glucocorticoid-induced matrix temporally modulate key integrins, caveolins, contractility, and stiffness in human trabecular meshwork cells. Investig. Ophthalmol. Vis. Sci.61, 16 (2020).
doi: 10.1167/iovs.61.13.16
Bronte, G., Puccetti, M., Crinò, L. & Bravaccini, S. Epithelial-to-mesenchymal transition and EGFR status in NSCLC: the role of vimentin expression. Ann. Oncol.30, 339–340 (2019).
pubmed: 30576405
doi: 10.1093/annonc/mdy548
Hann, C. R., Springett, M. J., Wang, X. & Johnson, D. H. Ultrastructural localization of collagen IV, fibronectin, and laminin in the trabecular meshwork of normal and glaucomatous eyes. Ophthalmic Res.33, 314–324 (2001).
pubmed: 11721183
doi: 10.1159/000055687
Tawara, A., Tou, N., Kubota, T., Harada, Y. & Yokota, K. Immunohistochemical evaluation of the extracellular matrix in trabecular meshwork in steroid-induced glaucoma. Graefes Arch. Clin. Exp. Ophthalmol. Albrecht Von Graefes Arch. Klin. Exp. Ophthalmol.246, 1021–1028 (2008).
doi: 10.1007/s00417-008-0800-0
Medina-Ortiz, W. E., Belmares, R., Neubauer, S., Wordinger, R. J. & Clark, A. F. Cellular fibronectin expression in human trabecular meshwork and induction by transforming growth factor-β2. Investig. Ophthalmol. Vis. Sci.54, 6779–6788 (2013).
doi: 10.1167/iovs.13-12298
Shepard, A. R. et al. Delayed secondary glucocorticoid responsiveness of MYOC in human trabecular meshwork cells. IInvestig. Ophthalmol. Vis. Sci.42, 3173–3181 (2001).
Polansky, J. R., Fauss, D. J. & Zimmerman, C. C. Regulation of TIGR/MYOC gene expression in human trabecular meshwork cells. Eye Lond. Engl.14(Pt 3B), 503–514 (2000).
Faralli, J. A., Schwinn, M. K., Gonzalez, J. M., Filla, M. S. & Peters, D. M. Functional properties of fibronectin in the trabecular meshwork. Exp. Eye Res.88, 689–693 (2009).
pubmed: 18835267
doi: 10.1016/j.exer.2008.08.019
Gospodarowicz, D. & Ill, C. Extracellular matrix and control of proliferation of vascular endothelial cells. J. Clin. Investig.65, 1351–1364 (1980).
pubmed: 7410547
pmcid: 371473
doi: 10.1172/JCI109799
Mao, Y. & Schwarzbauer, J. E. Fibronectin fibrillogenesis, a cell-mediated matrix assembly process. Matrix Biol. J. Int. Soc. Matrix Biol.24, 389–399 (2005).
doi: 10.1016/j.matbio.2005.06.008
Tripathi, B. J. & Tripathi, R. C. Neural crest origin of human trabecular meshwork and its implications for the pathogenesis of glaucoma. Am. J. Ophthalmol.107, 583–590 (1989).
pubmed: 2729407
doi: 10.1016/0002-9394(89)90253-5
Buffault, J., Labbé, A., Hamard, P., Brignole-Baudouin, F. & Baudouin, C. The trabecular meshwork: structure, function and clinical implications. A review of the literature. J. Fr. Ophtalmol43, e217–e230 (2020).
pubmed: 32561029
doi: 10.1016/j.jfo.2020.05.002