Lactate dehydrogenase isoenzymes during regeneration of the lens and retina in adult tritons

The range of lactate dehydrogenase (LDG) isozymes has been studied at the consecutive stages of retina regeneration from pigmented epithelium cells and lens regeneration from iris margin in adult crested newts. It was shown that the spectra of LDG isozymes peculiar to pigment epithelium cells and iris and characterized by the predominance of slowly migrating forms are replaced in the lens and retina regenerates by spectra characterized by the predominance of rapidly migrating isozymes which are peculiar to definitive lens and retina.     

Radioautographic study of the cellular proliferation of retinal pigment epithelium in axolotls

The proliferative activity of the pigment epithelium cells in the axolotl eyes was studied using 3H-thymidine in two types experiments: after the removal of lens, iris and retina and upon the cultivation of the pigment epithelium pieces in the cavity of lens-less eye. Irrespective of the operation type, the level of proliferation of the pigment epithelium cells changed regularly with respect to the time of observation. In the intact eye, the level of proliferation of the pigment epithelium cells was not high: the index of labelled nuclei equaled 0.5%, no mitoses were found. The highest values of the index of labelled nuclei (12.6-32.1%) and of the mitotic index (0.54-1.07%) were registered on the 10-20th days after the operation. After 40 days, the indices of proliferative activity of the pigment epithelium cells approached gradually those for the intact eye. The cultivation of the pigment epithelium cells in the cavity of a lens-less eye for 50 days did not result in their transdifferentiation into retina cells. The layered retina found in 7.7% of cases after the removal of lens, iris and retina could regenerate either from the cells of the retina growth zone localized in the region of embryonic split, or due to transdifferentiation of the pigment epithelium cells.     

Melanin granule metabolism in the iris and retinal pigment epithelium in adult tritions after completion of eye regeneration

It was concluded that the newly synthesized melanin granules were replaced in the pigmented tissues of the newt eye on the basis of redistribution of the cells of pigment epithelium of retina and iris labelled by 3H-DOPA 2.5 and 6.5 months after the isotope injection. The replacement of melanin granules and, correspondingly, melanin synthesis proceed more actively in the peripheral zones of the pigment epithelium of retina. The depigmentation of cells preceding the melanin synthesis appears to be realized with the participation of macrophages.     

Genetic mechanisms of cell transdifferentiation

Studies have been considered, which concern identification of regulatory genes in adult newts and their expression during retinal and lens regeneration. B.L. Astaurov repeatedly urged to join efforts of geneticists and embryologists in studies of the mechanisms underlying biological phenomena. This was also true for studies of regeneration. Such studies became possible only after introduction of molecular biology methods. Studies of the mechanisms underlying regeneration have been recently carried out jointly by geneticists and developmental biologists. This review presented at the conference dedicated to the 100th anniversary of B.L. Astaurov deals with these aspects in studies of regeneration.     

Study of β-III tubulin expression in human eye tissues during prenatal development

The expression of β-III tubulin, a marker protein of early neuronal cells, was studied by molecular genetic and immunochemical techniques. The study was performed with the eyes of human fetuses on weeks 8.5 to 27–28 of intrauterine development. Expression of β-III tubulin was detected immunochemically in the retina and lens fibers of fetuses at stages of 8.5 to 22–23 weeks. By means of PCR, a strong expression of the β-III tubulin gene was revealed in the retina of 9.5-week fetuses. Its level remained high until week 18, became slightly lower by week 24, and decreased to zero by week 27–28. The expression of this gene was also revealed in the lens of 9.5-week fetuses. Its level at stages of 15 to 24 weeks was very low, and no expression could be detected in 27-to 28-week fetuses. The results of PCR analysis were consistent with immunochemical data.     

Cultivation of the retinal pigment epithelium in the cavity of the lentectomized eye of newts

A study was made of proliferative activity and transdifferentiation of the cells of retinal pigment epithelium (RPE) cultivated in the cavity of the lensectomized eye of adult newt. Implantation of the newt RPE together with vascular membrane and scleral coat resulted in the regeneration of retina. In this process the character of changes in the proliferative activity of RPE and differentiation of retinal cells were the same as in the regeneration of retina in situ. RPE implanted with the vascular membrane alone, despite a high level of proliferation during the first ten days of cultivation, no differentiated retina was formed. Possible causes of these differences are discussed, and the comparison is made of the data obtained with those on RPE cultivation in vitro. After lens removal, with RPE implants present in the eye cavity, in addition to the regenerated lens, 2-3 extra lenses and retina were formed from the cells of the inner layer of the recipient's dorsal iris. Also some cases were revealed of lens formation from the cells of ventral iris. With a complete detachment of the recipient's retina (an after-effect of transplantation) a second differentiated retina regenerated in situ from the recipient's RPE cells.     

Radioautographic study of the cell proliferation of the pigment epithelium of the retina in albino clawed frogs

The proliferative activity of pigment epithelium was studied by means of 3H-thymidine autoradiography after the removal of retina, lens and iris with the ciliary-terminal zone in the adults. The cell population of pigment epithelium was shown to be heterogeneous on the level of proliferative activity. A low level of proliferation is characteristic of the cells of epithelial monolayer and the cells leaving it and forming aggregates. An intensive local proliferation leading to the formation of expansions was found in the pigment epithelium layer in 7% of cases. On the 20th day after the operation, the index of labelled nuclei in the expansions amounted to 43.4--59.3% and the mitotic index to 1.4--2.1%. On the 75th day elements of atypical retinal differentiation, besides the high proliferative activity, were observed in one expansion.