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: 3367657
• 负责人: Susan R Mallery
• 金额: $ 16.66万
• 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/3367657
• 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众多疾病之一 病人。KS的特征，包括频繁的多焦点 提出，以及恢复免疫能力后的解决方案，建议 这种疾病的系统的、环境的组成部分。KS的其他方面， 例如男性的性别偏好，以及 微血管系统，暗示KS与其他血管生成疾病之间存在关联 艾斯。 因为之前在这个实验室进行的研究表明 细胞硫醇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，细胞内硫醇氧化还原生物能 KS细胞周期进程中的状态(TRBS)。这项提议假定 在KS发生的增长放松管制是可归因于，至少在 部分，与环境介导的细胞内TRBS/钙离子的扰动有关 存储状态。实验设计包括对微血管的研究 内皮细胞(MVE)在内皮细胞生长介质(ECGM)中培养， 在ECGM中培养KS细胞以模拟体内KS环境， 然后是培养条件的互惠变化。具体目标： 和II研究将对KS和MVE细胞和 调查环境影响对细胞的贡献 表型。生物能量状态将通过高效液相色谱来确定。 核苷酸图谱和双束动力学定量谷胱甘肽 分光光度分析。具体目标III将解决是否存在 细胞内TRBS和有丝分裂反应之间的联系。 将进行实验以评估细胞内钙的储存状态 通过监测内质网(E.R.)钙离子负载和肌醇 三磷酸引发钙离子释放(45Ca2+用于E.R.分析)，和钙离子 有丝分裂原碱性成纤维细胞生长因子(完整细胞，Fura-2 AM)的动员 荧光分析和显微分析)。流式细胞仪DNA分析和~3H- 胸腺嘧啶核苷掺入将用于跟踪细胞周期进程和 DNA合成。特定目标IV研究将确定 特异性类固醇对细胞增殖/细胞周期的影响 KS和KS的进行性反应和自分泌白介素6的产生 MVE细胞。 尽管这项研究的目的是为了深入了解糖尿病的发病机制 KS，它还将生成适用于其他疾病的信息 这是毛细血管增长放松管制的结果。这些项目的未来目标 研究是开发治疗方式来解决/缓解 这些生长障碍。