C Guidry1. 1. Department of Ophthalmology, Eye Foundation Hospital, University of Alabama at Birmingham, 35294, USA.
Abstract
PURPOSE: To characterize phenotypic and antigenic changes in isolated Müller cells during proliferation in extended culture. METHODS: Müller cells were isolated from porcine retina by sequential papain and DNase digestion, trituration, and density gradient centrifugation. The identity of the isolated cells was confirmed by immunodetection of carbonic anhydrase II (CA-II), cellular retinaldehyde-binding protein (CRALBP), glial fibrillary acidic protein (GFAP), vimentin, and delta smooth muscle actin (alpha SMA). Continuously proliferating cells established in culture were examined for changes in the expression of these antigens. RESULTS: Primary cultures of purified Müller cells, incubated under routine culture conditions, were proliferative and lost immunodetectable CRALBP within 2 weeks. The expression of CA-II also diminished with time, but at an apparently lower rate than that of CRALBP. Loss of GFAP expression was even more gradual and was complete by passage 5. De novo expression of alpha SMA was detectable in some cells within 12 days in culture and by all cells by passage 5. During this period, vimentin expression remained qualitatively unchanged. CONCLUSIONS: Isolated porcine Muller cells in culture undergo a phenotypic dedifferentiation to a fibroblast-like cell, which includes loss of detectable CRALBP, CA-II, and GFAP, and they acquire expression of the myoid marker alpha SMA.
PURPOSE: To characterize phenotypic and antigenic changes in isolated Müller cells during proliferation in extended culture. METHODS: Müller cells were isolated from porcine retina by sequential papain and DNase digestion, trituration, and density gradient centrifugation. The identity of the isolated cells was confirmed by immunodetection of carbonic anhydrase II (CA-II), cellular retinaldehyde-binding protein (CRALBP), glial fibrillary acidic protein (GFAP), vimentin, and delta smooth muscle actin (alpha SMA). Continuously proliferating cells established in culture were examined for changes in the expression of these antigens. RESULTS: Primary cultures of purified Müller cells, incubated under routine culture conditions, were proliferative and lost immunodetectable CRALBP within 2 weeks. The expression of CA-II also diminished with time, but at an apparently lower rate than that of CRALBP. Loss of GFAP expression was even more gradual and was complete by passage 5. De novo expression of alpha SMA was detectable in some cells within 12 days in culture and by all cells by passage 5. During this period, vimentin expression remained qualitatively unchanged. CONCLUSIONS: Isolated porcine Muller cells in culture undergo a phenotypic dedifferentiation to a fibroblast-like cell, which includes loss of detectable CRALBP, CA-II, and GFAP, and they acquire expression of the myoid marker alpha SMA.
Authors: Derek H Rosenzweig; Mourad Matmati; Ghazaleh Khayat; Sidharth Chaudhry; Boris Hinz; Thomas M Quinn Journal: Tissue Eng Part A Date: 2012-08-03 Impact factor: 3.845
Authors: Yun-Bi Lu; Kristian Franze; Gerald Seifert; Christian Steinhäuser; Frank Kirchhoff; Hartwig Wolburg; Jochen Guck; Paul Janmey; Er-Qing Wei; Josef Käs; Andreas Reichenbach Journal: Proc Natl Acad Sci U S A Date: 2006-11-08 Impact factor: 11.205