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Leo A Kim, Lindsay L. Wong, Dhanesh Amarnani, Alex A. Bigger-Allen, Yang Hu, Christina Marko, Vinay Shah, Dean Eliott, Joseph F Arboleda-Velasquez, Patricia A D'Amore; Analysis and culture of cells from patient-derived membranes in proliferative diabetic retinopathy. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4683. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Formation of fibrovascular membranes (FVMs) is a hallmark of proliferative diabetic retinopathy (PDR). Complications of PDR such as retinal detachment and vitreous hemorrhage is addressed by surgical removal of FVMs. Here, we analyze and culture cells from surgically removed and patient-derived FVMs.
FVMs obtained from patients during vitrectomy surgery were analyzed by electron microscopy (EM), comparative genomic hybridization (CGH), immunohistology, and/or digested with collagenase II for cell isolation and culture. Antibody arrays and ELISA were used to profile angiogenesis-related proteins in cell culture supernatants.
Ultrastructural analysis of FVM showed abnormal vessels within fibrous stroma. FVMs can be stratified by proliferative activity according to the distribution of specific cell markers. The cellular constituents of FVMs lacked major chromosomal aberrations as shown by CGH analysis. Cells derived from fibrovascular membranes could be isolated and maintained in culture. FVM cells can be maintained in primary culture and retain the expression of cell markers detected in FVM, which define specific cell populations including CD31, alpha-smooth muscle actin, and glial fibrillary acidic protein-positive cells. Secretion of angiopoietin-1 and thrombospondin-1 by these cultured cells was found to significantly decrease in high glucose and under hyperosmolar conditions compared to human retinal pericytes derived from a non-diabetic donor.
Cultured FVM cells recapitulate some of the functional characteristics of FVMs from patients with PDR by expressing characteristic cell markers found within FVMs, exhibiting cellular proliferation, and secreting angiogenic factors following unique patterns in response to environmental alterations. We believe these characteristics make cultured FVMs a useful tool in the evaluation of the underlying molecular mechanisms of PDR.
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