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Tina T. Wong, Padmapriya Sathiyanathan, Stephanie Chu, Li Fong Seet, Lawrence W. Stanton; In Vitro Expansion Of Mesenchymal Stem Cell-like Cells From The Human Trabecular Meshwork. Invest. Ophthalmol. Vis. Sci. 2011;52(14):5997.
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Development of primary open-angle glaucoma (POAG) is associated with increased resistance to aqueous humour outflow through the trabecular meshwork. The decrease in trabecular meshwork (TM) cell population and cellular dysfunction resulting in excessive deposition of extracellular matrix proteins, both characteristics of glaucomatous TM tissue, are believed to contribute to increased outflow resistance and disease development. Putative stem cells are thought to exist in the TM. The purpose of the study is to culture cells derived from the TM and characterize them for stem cell characteristics.
The TM was stripped from cadaver eyes and treated with collagenase overnight. The samples were trypsinized and seeded in DMEM (low glucose) with 10% FBS, 4 mM L-glutamine, 1 mM sodium pyruvate and 1 mM non-essential amino acids. The expanded cells were characterized for markers of TM tissue as well as stem cells by real-time quantitative PCR (qPCR), immunofluorescent analyses and flow cytometry.
qPCR showed that the expanded cells were positive for the TM markers, CHI3L1, ANK3, HMFG-1, MMP1, MGP and LDL-R. Both qPCR and immunofluorescent analyses confirmed the expression of the stem cell markers, Nanog, Oct4 and Nestin. qPCR also revealed the expression of the mesenchymal stem cell (MSC) markers CD73, CD90, CD105, and CD146, and the absence of CD31, CD34, and CD45 expression which are not found in MSCs. Flow cytometry analyses confirmed the expression of the MSC markers in a significant population of the cultured cells. Morphologically, the cells resembled MSCs in appearance.
Cells derived from the TM tissue and cultured in vitro express stem cell as well as MSC markers. These cells may have the potential to help replace lost or dysfunctional cells in the TM tissue associated with glaucoma.
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