InVitro Effects of Thymoglobulin in Human Embryonic Kidney Cell Line (HEK293) in Culture.

Abstract

Anti-thymocyte globulins are polyclonal T-cell-depleting immunoglobulins used in induction of immunosuppression in kidney transplant recipients. Thymoglobulin is purified rabbit immunoglobulin (Ig)G, obtained by immunization of rabbits with fetal human thymus, which depletes T lymphocytes by complement-dependent lysis and apoptosis, reduces production of cytokines, and decreases expression of adhesion molecules in endothelial cells. To determine possible direct effects of Thymoglobulin on kidney cells during transplantation, we used the Human Embryonic Kidney cell line (HEK293) in culture. We measured membrane potential of the cells by use of the slow whole patch-clamp technique. We determined effects of Thymoglobulin on cell death and proliferation during hypoxia/re-oxygenation injury, using a hypoxic chamber. Depolarizations of HEK293 cells caused by Thymoglobulin were concentration-dependent and membrane potential-dependent, showing direct effects of Thymoglobulin on the HEK293 cells, whereas rabbit anti-thymocyte globulin produced against Jurkat cells (ATG-F) and normal rabbit IgG had no effects. To determine the effects of Thymoglobulin in hypoxia/re-oxygenation conditions, cells were incubated for 24 hours with Thymoglobulin in an atmosphere with 5% CO2-95% N2 at 37°C followed by 1 hour in atmosphere with 5% CO2-95% air at 37°C. The effects of hypoxia/re-oxygenation were detected by calculating cell death and determining the cell growth, using scratch test. Thymoglobulin prevented the cell death induced by hypoxia and re-oxygenation conditions. In addition, it accelerated the cell growth (improved scratch wound-healing). This is the first study to show the direct effects of Thymoglobulin on kidney-derived epithelial cells, which may lead to better understanding of its effects in kidney transplantation.