A study of the proliferating activity in lens epithelium and the identification of tissue-type stem cells

Using a newly established fixation method and immunohistochemical methods, we precisely described the regions of cells stained with various antibodies relating to cell proliferation; this method enabled us to make cellular-level diagrams of the epithelium in which the position of every lens epithelial cell (LEC) was determined in reference to the cell located at the top of the bow area. The proliferating activity of LECs of 4-week-old (4W) mice was examined either by labeling with 5-bromodeoxyuridine (BrdU) in vivo or by measuring the amount of mRNA prepared from LECs, which had been separated into the posterior part, containing the germinative zone, and the anterior part and then cultured. The epithelial region stained with antibody for proliferating cell nuclear antigen (PCNA) and cyclin D1 remained relatively constant during the study period, although the positive region was reduced a little from embryonic day 18 (E18) to 12W. This region at 4W overlapped well with the DNA synthesizing region. Therefore, we reasoned that this region would correspond to the germinative zone of the adult mouse. Considering together with results of the reactivation pattern of genes, we considered that the location of tissue-type stem cells in lens epithelium (LE) as immediately anterior to the germinative zone.