GROWTH FACTOR EXPRESSION IN RENAL ENDOTHEIAL CELLS

血管内皮下部位的破坏性增生和硬化 肾微血管系统不可逆地损害肾功能， 管腔狭窄，肾小球结构破坏和 过滤面这些研究的目的是确定机制 调节肾内皮细胞产生生长因子， 与内皮细胞的增殖和硬化有关 层. 在破坏性肾小球疾病中尤其相关的是增殖 系膜细胞、成纤维细胞和基质沉积，所有这些都是 由血小板衍生生长因子（PDGF）刺激。我们的数据，使用 原代培养的人肾微血管内皮细胞，已显示 PDGF B/c-sis信使RNA（mRNA）表达和活性释放是 受血管部位相关因子的调节。PDGF的转录 B/c-sis mRNA由血小板（转化生长因子β）诱导， 凝血产物（凝血酶）和抑制激素，提高 cAMP水平（儿茶酚胺）。新数据显示肾内皮细胞 PDGF B mRNA的不同结构形式的表达。拟议 研究将确定不同PDGF B mRNA的相对贡献 利用体外和非洲爪蟾翻译活性PDGF蛋白产物， 卵母细胞翻译的分离和合成的结构形式的每一个 PDGF B/c-cic mRNA。另外的实验将确定是否表达 不同的PDGF B/c-sis mRNAs受转录调控的差异， 稳定机制。 基因组PDGF B/c-cic序列参与肾功能的调节 将通过DNA足迹技术鉴定内皮表达 和一系列侧翼序列缺失的瞬时表达 结构。参与cAMP介导的PDGF抑制的DNA序列 将通过凝胶位移鉴定和表征B/c-sis转录 分析，以及通过紫外线交联和作为 cAMP依赖性激酶的底物。PDGF B/c-sis的完整组织作用 使用PDGF B/c-sis的原位杂交来鉴定表达 肾活检样本中的mRNA。 这些研究将确定调节微血管的分子机制 PDGF的内皮生产，并可能提供潜在的目标， 干预破坏性的内皮下增殖过程。

Destructive proliferation and sclerosis at sites underlying endothelium in the kidney microvasculature irreversibly impair renal function through luminal narrowing, destruction of glomerular architecture and reduction of filtration surface. The objective of these studies is to define mechanisms regulating renal endothelial cell production of growth factors that are implicated in proliferation and sclerosis subjacent to the endothelial layer. Especially relevant in destructive glomerular diseases are proliferation of mesangial cells, fibroblasts and matrix deposition, all of which are stimulated by platelet-derived growth factor (PDGF). Our data, using primary cultured human renal microvascular endothelial cells, have shown PDGF B/c-sis messenger RNA (mRNA) expression and activity release are regulated by agents relevant at vascular sites. Transcription of PDGF B/c-sis mRNA is induced by platelet (transforming growth factor beta) and coagulation products (thrombin) and suppressed by hormones which elevate cAMP levels (catecholamines). New data has shown renal endothelial expression of distinct structural forms of PDGF B mRNA. The proposed studies will determine the relative contribution of different PDGF B mRNA's to translation of active PDGF protein product, using in vitro and xenopus oocyte translation of isolated and synthesized structural forms of each PDGF B/c-cic mRNA. Additional experiments will determine if expression of different PDGF B/c-sis mRNAs is differentially regulated by transcriptional and stabilization mechanisms. Genomic PDGF B/c-cic sequences participating in regulation of renal endothelial expression will be identified by DNA footprinting techniques and transient expression of a series of flanking sequence deletion constructs. DNA sequences participating in cAMP-mediated repression of PDGF B/c-sis transcription will be identified and characterized by gel shift assays, and DNA binding proteins identified by UV cross-linking and as substrates for cAMP dependent kinase. Intact tissue roles for PDGF B/c-sis expression will be identified using in situ hybridization of PDGF B/c-sis mRNA in renal biopsy samples. These studies will define molecular mechanisms regulating microvascular endothelial production of PDGF and may provide potential targets for intervention in destructive subendothlial proliferative processes.