Kaposi's sarcoma is one of the myriad of diseases that affect AIDS patients. Features of KS, which include a frequent multifocal presentation, and resolution upon restoration of immune competence, suggest a systemic, environmental component to this disease. Other aspects of KS, such as the male gender predilection, and involvement of the microvasculature, imply an association between KS and other angiogenic dise es. Because prior studies conducted in this laboratory have shown an association between the cellular thiol redox status and cell cycle progression, this study will investigate the contribution of this putative intracellular regulatory parameter, the cellular thiol redox bioenergetic status (TRBS) in cell cycle progression in KS. This proposal hypothesizes that the growth deregulation that occurs in KS is attributable, at least in part, to an environmentally mediated perturbation of the cellular TRBS/Ca2+ storage state. The experimental design includes studies of microvascular endothelial (MVE) cells cultured in endothelial cell growth medium (ECGM), KS cells cultured in ECGM supplemented to simulate the in vivo KS milieu, and then a reciprocal change in the culture conditions.
Specific Aims I and II studies will biochemically characterize the KS and MVE cells and investigate the contribution of environmental influences on the cellular phenotype. the bioenergetic status will be determined by an HPLC nucleotide profile, and the glutathione quantitated by a kinetic, dual beam spectrophotometric assay.
Specific Aim III will address whether there is an association between the cellular TRBS and mitogenic responsiveness. Experiments will be conducted to evaluate the cellular Ca2+ storage state by monitoring endoplasmic reticulum (e.r.) Ca2+ loading, and inositol triphosphate initiated Ca2+ release (45Ca2+ used for e.r. assays), and Ca2+ mobilization in response to the mitogen BFGF (intact cells, fura-2 AM fluorimetric and microscopic assays). Flow cytometric DNA analyses and 3H- thymidine incorporation will be used to follow cell cycle progression and DNA synthesis, respectively.
Specific Aim I V studies will determine the effects of specific sex steroids on the proliferative/cell cycle progression responses, and autocrine interleukin-6 production, in KS and MVE cells. Although this study is designed to provide insight into the pathogenesis of KS, it will also generate information that is applicable to other diseases that result from capillary growth deregulation. A future goal of these studies is the development of treatment modalities to address/alleviate these growth disturbances.
