CALCIUM THIOL REDOX INTERACTION IN KAPOSI'S CELL CYCLE

卡波西细胞周期中硫醇钙氧化还原相互作用

• 批准号: 2224629
• 负责人: Susan R Mallery
• 金额: $ 21.18万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-04-01 至 1996-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2224629
• 关键词: DNA replication; Kaposi's sarcoma; autocrine; bioenergetics; calcium; calcium flux; calcium indicator; cell cycle; cell growth regulation; endoplasmic reticulum; environment; flow cytometry; glutathione; high performance liquid chromatography; human subject; inositol phosphates; interleukin 6; nucleotides; oxidation reduction reaction; radiotracer; sex hormones; spectrometry; thiols; thymidine; tritium; vascular endothelium

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 IV 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.

卡波西肉瘤是影响艾滋病的无数疾病之一 患者 KS的特征，包括频繁的多灶性 呈现和恢复免疫能力后的消退，表明 一种系统性的环境因素 KS的其他方面， 比如男性的性别偏好， 微血管，暗示KS和其他血管生成性疾病之间的联系 es. 因为在这个实验室进行的先前研究表明， 细胞巯基氧化还原状态与细胞周期的关系 进展，这项研究将调查这一假定的贡献 细胞内调节参数，细胞巯基氧化还原生物能 状态（TRBS）在KS细胞周期进程中的作用。 该提案假设 在KS中发生的生长失调是可归因的，至少 部分，环境介导的细胞TRBS/Ca2+扰动 存储状态。 实验设计包括微血管的研究 在内皮细胞生长培养基（ECGM）中培养的内皮（MVE）细胞， 在ECGM中培养的KS细胞补充以模拟体内KS环境， 然后是培养条件的相互变化。 具体目标一 和II研究将对KS和MVE细胞进行生物化学表征， 研究环境对细胞的影响， 表型 生物能量状态将由HPLC确定 核苷酸谱，并通过动力学双光束定量谷胱甘肽 分光光度测定 具体目标三将涉及是否存在 细胞TRBS和促有丝分裂反应之间的关联。 将进行实验以评估细胞Ca2+储存状态 通过监测内质网（e.r.）Ca2+负载和肌醇 三磷酸盐引发的Ca 2+释放（45Ca 2+用于e.r.测定）和Ca2 + 响应于促分裂原BFGF（完整细胞，fura-2 AM）的动员 荧光测定和显微镜测定）。 流式细胞术DNA分析和3H- 胸苷掺入将用于跟踪细胞周期进程， DNA合成。 具体目标IV研究将确定 特异性类固醇对增殖/细胞周期的影响 进展反应和自分泌白细胞介素-6的产生， MVE细胞。 尽管这项研究旨在深入了解 KS，它还将产生适用于其他疾病的信息 这是由毛细血管生长失调引起的。 这些未来的目标 研究是发展治疗方式，以解决/缓解 这些生长障碍。