FIBROBLAST HETEROGENEITY IN NORMAL AND FIBROTIC LUNGS

间质性肺纤维化的特征是 肺实质结缔组织过多， 胶原蛋白的合成和改变I型：III型胶原蛋白的比例。 成纤维细胞被认为是主要的细胞类型， 产生过多的结缔组织成分。 我们和 其他人先前已经表明成纤维细胞是异质的， 并不是所有的成纤维细胞对环境介质的反应都一样， 并且成纤维细胞群体可以基于特定的 成纤维细胞亚群和物质之间的相互作用 导致细胞选择， 存在具有独特特性的成纤维细胞亚群 有助于纤维化的演变和发生。 我们建议 从成人、正常人肺和从肺分离成纤维细胞 早期和晚期纤维化，并描述其生长特征， 特点，生产胶原蛋白和弹性蛋白，收缩 属性和C1 q绑定。 我们将分离成纤维细胞 正常肺培养物的亚群， 基于C1 q差异的荧光激活细胞分选仪 约束，并确定其属性。 然后我们将纤维化 肺成纤维细胞培养并检查获得的群体。 我们 将决定是否获得成纤维细胞亚群， 这种方式在它们对三种炎性反应中不同， 已知参与成纤维细胞调节的介质：PDGF， TGF-β和γ-干扰素。 最后，我们建议确定 成纤维细胞亚群的特异性表型标志物， 开发探针来识别这些细胞。 我们将生成cDNA 使用从亲本培养物分离的mRNA的文库， 用来自成纤维细胞亚群的标记的cDNA将它们标记，和 鉴定含有特定DNA序列的克隆。 我们还将 试图利用一组抗体来区分 成纤维细胞亚群 使用这些探针，我们建议识别 正常和纤维化肺组织中的成纤维细胞亚群 标本 我们希望从这些结果中，我们能够 确定成纤维细胞亚群如何促进肺 纤维化

Interstitial pulmonary fibrosis is characterized by alterations in the lung parenchyma with excess connective tissue, increased rate of collagen synthesis and altered type I:III collagen ratio. Fibroblasts are believed to be the major cell type responsible for the production of excess connective tissue constituents. We and others have previously shown that fibroblasts are heterogeneous, that not all fibroblasts respond alike to environmental mediators, and that fibroblast populations can be separated based on specific interactions between fibroblast subpopulations and substances released by inflammatory cells lead to cell selection, and that presence of fibroblast subpopulations with distinctive properties contribute to the evolution and genesis of fibrosis. We propose to isolate fibroblasts from adult, normal human lungs and from lungs with early and late fibrosis, and to characterize their growth characteristics, production of collagen and elastin, contractile properties, and C1q binding. We will separate fibroblast subpopulations from cultures of normal lungs using a fluorescence-activated cell sorter based on differences in C1q binding, and determine their properties. We will then sort fibrotic lung fibroblast cultures and examine the populations obtained. We will determine whether the fibroblast subpopulations obtained in this manner differ in their response to three inflammatory mediators known to be involved in fibroblast regulation: PDGF, TGF-beta, and gamma-interferon. Finally, we propose to identify specific phenotypic markers for fibroblast subpopulations and develop probes to identify these cells. We will generate cDNA libraries using mRNA isolated from parent cultures, hybridize them with labeled cDNA from fibroblast subpopulations, and identify clones containing specific DNA sequences. We will also attempt to utilize a battery of antibodies to differentiate fibroblast subsets. Using these probes, we propose to identify fibroblast subpopulations in normal and fibrotic lung tissue specimens. We expect that from these results we will be able to determine how fibroblast subpopulations contribute to pulmonary fibrosis.