F E Kruse1, S C Tseng. 1. Department of Ophthalmology, Bascom Palmer Eye Institute, University of Miami School of Medicine, FL 33101.
Abstract
PURPOSE: To determine if vitamin A could be one of the factors in serum responsible for the previously observed effect of 20% fetal bovine serum on stimulating clonal growth of an additional subpopulation in limbal cultures, possibly stem cells, but inhibiting that of transient amplifying cells in peripheral corneal cultures. METHODS: A reported serum-free clonal growth assay was used. The mitogenic response was measured by colony-forming efficiency (CFE), colony size, and BrdU labeling index; the differentiation was assessed by colony morphology, AE-5 monoclonal antibody staining, and cornified envelope formation. RESULTS: HPLC analyses revealed that added retinoic acid (RA) was rapidly taken up by cultured cells. As compared to the control without RA, low concentrations of RA (10(-9) M to 10(-7) M) stimulated the CFE of limbal cultures but did not change that of peripheral corneal cultures. Furthermore, 10(-8) M RA induced the emergence of two new types of colonies, one of which was almost exclusively present in limbal cultures, and allowed continuous clonal growth of some colonies in late limbal cultures. RA also dose dependently reduced colony size and BrdU labeling index in both limbal and peripheral corneal cultures. RA in concentrations above 10(-8) M stimulated normal differentiation of both limbal and peripheral corneal epithelial cells, as evidenced by increased AE-5 staining, but inhibited the formation of cornified envelopes, an index for abnormal, squamous metaplasia, in late cultures. CONCLUSION: These results suggest that RA has a differential dose-dependent effect on subpopulations of corneal and limbal epithelial cells. Although RA stimulates the conversion of limbal stem cells to transient amplifying cells, it inhibits the amplification of corneal and limbal transient amplifying cells and prevents abnormal terminal differentiation. These data further support the role of vitamin A as a physiological modulator of proliferation and differentiation of the ocular surface epithelium.
PURPOSE: To determine if vitamin A could be one of the factors in serum responsible for the previously observed effect of 20% fetal bovine serum on stimulating clonal growth of an additional subpopulation in limbal cultures, possibly stem cells, but inhibiting that of transient amplifying cells in peripheral corneal cultures. METHODS: A reported serum-free clonal growth assay was used. The mitogenic response was measured by colony-forming efficiency (CFE), colony size, and BrdU labeling index; the differentiation was assessed by colony morphology, AE-5 monoclonal antibody staining, and cornified envelope formation. RESULTS: HPLC analyses revealed that added retinoic acid (RA) was rapidly taken up by cultured cells. As compared to the control without RA, low concentrations of RA (10(-9) M to 10(-7) M) stimulated the CFE of limbal cultures but did not change that of peripheral corneal cultures. Furthermore, 10(-8) M RA induced the emergence of two new types of colonies, one of which was almost exclusively present in limbal cultures, and allowed continuous clonal growth of some colonies in late limbal cultures. RA also dose dependently reduced colony size and BrdU labeling index in both limbal and peripheral corneal cultures. RA in concentrations above 10(-8) M stimulated normal differentiation of both limbal and peripheral corneal epithelial cells, as evidenced by increased AE-5 staining, but inhibited the formation of cornified envelopes, an index for abnormal, squamous metaplasia, in late cultures. CONCLUSION: These results suggest that RA has a differential dose-dependent effect on subpopulations of corneal and limbal epithelial cells. Although RA stimulates the conversion of limbal stem cells to transient amplifying cells, it inhibits the amplification of corneal and limbal transient amplifying cells and prevents abnormal terminal differentiation. These data further support the role of vitamin A as a physiological modulator of proliferation and differentiation of the ocular surface epithelium.
Authors: Nilanjana Sengupta; Sergio Caballero; Sean M Sullivan; Lung-Ji Chang; Aqeela Afzal; Sergio Li Calzi; Jennifer L Kielczewski; Sabrina Prabarakan; E Ann Ellis; Leni Moldovan; Nicanor I Moldovan; Michael E Boulton; Maria B Grant; Edward W Scott; Jeffrey R Harris Journal: Mol Ther Date: 2009-07-07 Impact factor: 11.454
Authors: Gabriel Gonzalez; Yuzuru Sasamoto; Bruce R Ksander; Markus H Frank; Natasha Y Frank Journal: Wiley Interdiscip Rev Dev Biol Date: 2017-11-03 Impact factor: 5.814