Different Effects of Regulatory Genes (tat, nef) of Human Immunodeficiency Virus Type 1 (HIV-1) on the Proliferation and Differentiation of Mouse Embryonic Stem Cells in vitro.

To examine the effects of the tat and nef regulatory genes of human immunodeficiency virus (HIV-1) on cell differentiation we used the mouse embryonic stem cells (ESC) as a model. Proliferation, embryoid bodies (EB) formation and subsequent differentiation into cardiomyocytes, glial and neuronal cells were investigated in ESC lines transfected with these genes. It has been shown that the transfection of ESC by the tat gene increased their proliferating activity, whereas the nef gene transfected ESC showed its decrease. The number of embryoid bodies formed was higher in the cultures of ESC transfected by the nef and lower in the cells transfected by the tat in comparison with controls. The percentage of embryoid bodies with contracting cardiomyocytes was higher against control in the nef transfected cells and lower in ESC transfected with the tat. There were no reliable differences in the appearance of glial cells between control and the nef and tat transfected cell lines. Spontaneous differentiation of ESC into neuronal cells was almost not observed in the nef transfected cells, in contrast to control and the tat transfected cells. However, addition of retinoic acid (RA) to the nef transfected cells caused even a slight increase in neuron formation as compared to control ESC treated with RA. Thus, for the first time we have shown that the tat and nef regulatory genes of HIV-1 had a visible effect on proliferation of ESC and some first steps of their differentiation. In general, the reverse correlation between the effects of these two viral genes on ESC proliferation and differentiation were observed.

[Influence of regulatory genes of type 1 human immunodeficiency virus on proliferation and differentiation of murine embryonic stem cells]. / Vliianie reguliatornykh genov virusa immunodefitsita cheloveka tipa 1 na proliferatsiiu i differentsirovku émbrional'nykh stvolovykh kletok myshi.

Spontaneous formation of embryoid bodies and subsequent differentiation of some cells into cardiomyocytes were demonstrated on murine embryonic stem cells of R1 line. The lines of embryonic stem cells were obtained that had been transfected with genetic constructs carrying expressing regulatory genes of the human immunodeficiency virus tat and nef and "green protein" gene (GFP). The transfection of embryonic stem cells with the gene tat stimulated their proliferative activity, while this activity decreased in the cells transfected with the gene nef. The time necessary for the formation of embryoid bodies by all lines of transfected cells was similar to that in the control cells. In the cultures of cells transfected with nef and tat, the number of embryoid bodies and the percentage of embryoid bodies with contracting cardiomyocytes were higher and lower than in the control, respectively. Thus, an inverse correlation was observed between the effects of regulatory genes of the human immunodeficiency virus on proliferation and differentiation embryonic stem cells.

[Effects of human immunodeficiency virus type 1 nef and tat genes on rat PC12 pheochromocytoma cells]. / Effekty genov nef i tat virusa immunodefitsita cheloveka tipa 1 na kletki feokhromotsitomy krysy PC12.

The regulatory genes nef and tat of the human immunodeficiency virus type 1 (HIV-1) were transferred into the rat pheochromocytoma cells (line PC12) under the control of the eukaryotic promoters. Proliferative activity of the PC12 cells transfected with the tat HIV-1 gene was substantially increased as compared to the control. Conversely, the nef gene introduced into the cultivated PC12 cell caused inhibition of their proliferative activity and formation of cell agglomerates resembling in morphology the multinuclear syncytial cells. Thus, our results suggest that the tat gene activates proliferation of the cultivated PC12 cells, whereas the nef gene inhibits proliferation of the same cells. We have obtained for the first time a direct indication for the possible role of the nef gene in formation of multinuclear T-lymphocyte and macrophage syncytium in HIV-1-infected patients. The HIV-1 nef and tat genes had no significant effect on the neuronal differentiation of the PC12 cells induced by the nerve growth factor (NGF).

[Effects of the nef and tat genes of the human immunodeficiency virus type I on rodent cells in vivo and in vitro]. / Vliianie genov nef i tat virusa immunodefitsita cheloveka tipa 1 na kletki gryzunov in vivo i in vitro.

The tat and nef regulatory genes of the human immunodeficiency virus type I (HIV-1) under the control of eukaryotic promoters were transferred in vivo into mice and in vitro into rat cell cultures. The development was disturbed and adenocarcinomas of the lacrimal glands and pancreas appeared in transgenic mice carrying the HIV-1 tat gene. Transfection with the tat gene altered morphology and increased proliferative activity of Rat-2 pseudonormal cells. The tat gene also induced the formation of neoplastic foci in a primary rat embryo fibroblast culture. The results obtained showed that the HIV-1 tat gene can act as an oncogene and activate the proliferation of cultured cells. Cell proportions in peripheral blood and bone marrow were altered and mitogen-induced lymphocyte proliferation was decreased in transgenic mice carrying the HIV-1 nef gene. This gene also significantly suppressed proliferation but had no effect on morphology of Rat-2 cells. Thus, the HIV-1 nef gene appeared to suppress proliferation of various animal cells.