Effect of mifepristone under acute stress on thymocyte apoptosis in mice

Abstract

In 1936, the Canadian pathologist Hans Selye, in experiments with adrenalectomy, has shown adrenal cortex hormones to be involved in development of thymic acute involution under stress. By 1970s, the idea of lysis of lymphoid cells by glucocorticoids dominated among researchers as the main mechanism of these changes. Later on, increased migration of T lymphocytes from thymus as well as decreased migration of bone marrow precursors to the thymus were considered. Since 1990s, apoptosis induced by glucocorticoids was also studied in this respect. To elucidate the in vivo contribution of glucocorticoids these events, a convenient tool was developed in experimental endocrinology and pharmacology, i.e., treatment of male rats or mice with antagonist of glucocorticoid and progesterone receptors mifepristone, also known as RU-38486 or RU-486. The purpose of this work is to investigate the effect of mifepristone on thymocyte apoptosis in mice under acute stress. Experimental studies were performed in male white noninbred mice. Mifepristone was injected once subcutaneously 30 min before immobilization at a dose of 50 mg/kg body weight prepared in olive oil solution. Control mice and the animals from comparison group were injected once subcutaneously with an equivalent amount of the drug solvent. A classical model of 24-hour immobilization stress in a plastic restrainer (supine position) was used for experimental simulation of acute stress. Thymocyte apoptosis was assessed by flow laser cytometry with BD PE Annexin V Apoptosis Detection Kit I (BD Pharmingen) reagent kit adapted for Guava EasyCyte flow laser cytometer by Millipore Corporation. It was found that acute stress induced by 24-hour immobilization of mice leads to an increase in the total relative number of 7-AAD-positive cells as well as proportion of cells stained with 7-AAD, but not annexin V-PE (nuclear debris, annexin V(-), 7-AAD (+)). Administration of mifepristone alleviated these changes, thus confirming involvement of glucocorticoids in increasing number of necrotic thymocytes. The number of thymocytes in early apoptosis (i.e., annexin V-PE-positive, 7-AAD-negative), as well as total relative number of cells with phosphatidylserine exposed at the surface (annexin V-PE-positive thymocytes) under acute stress and at stress on the background of the introduction of mifepristone did not differ from the control.