Role of beta-citryl-L-glutamate in the differentiation of retina pigmented epithelial cells to lens fiber cells

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1. When adult bovine lens epithelial cells were cultured in the presence of fetal calf serum, they differentiated into fiber cells and formed lentoid bodies at about 6 weeks in the culture. beta-citryl-L-glutamate (beta-CG) concentration increased from 2 to 4 weeks of the culture, reached to the highest level and then decreased gradually. alpha-crystallin, a marker of differentiation into lense fiber cells, increased extremely from 4 to 5 weeks of the culture and then continued to increase gradually. The cells of lentoid bobies were stained strongly by the anti-alpha-crystallin antibody, while cells around the structures were stained by the anti-beta-CG antibody.2. Retina pigmented epithelial cells were dissociated from eyes of 8-to 9-day-old chick embryos and were cultured in the medium containing phenylthiourea and testicular hyaluronidase. They rapidly lost metanosomes as they proliferated and dedifferentiated in culture, but they could not transdifferentiated into lens fiber cells in the medium containing phenylthiourea, hyaluronidase and ascorbic acid at extremely high cell density. The pigmented epithelial cells were transdifferentiated directly into lens fiber cells in the medium containing hyaluronidase at high cell density according to the method of Itoh et al. The epithelial cells differentiated into fiber cells and formed lentoid bodies efficiently, but neither cells of lentoid bodies nor cells around the lentoidbodies were stained by the anti-beta-CG antibody.Sixteen tryptic-digested peptides from the purified beta-CG-hydrolysing enzyme were prepared and their amino acid sequence were determined. None of their amino acid sequences was corresponded to the known amino acid sequence of proteins, but the sequence of some peptide from 100 and 115 kDa subunits showed 100% homology for cytoskeletal keratin.