A novel telomerase expressing lung fibroblast phenotype

描述(申请人提供)：成纤维细胞数量的增加是肺纤维化的关键特征，它们在密集的细胞形态中聚集，称为成纤维细胞灶，是特发性肺纤维化(IPF)的诊断组织病理学特征。成纤维细胞的增殖增加可能是其表型不同于从正常肺分离的细胞的机制之一。有趣的是，在肺损伤和纤维化的动物模型中发现了一种新的成纤维细胞表型，其特征是端粒酶活性的表达。虽然端粒酶活性与某些发育早期的细胞和许多肿瘤细胞的增殖能力和寿命增加有关，但其在肺纤维化发病机制中的意义尚不清楚。在这个项目中，假设表达端粒酶的成纤维细胞表型的从头出现是由某些因素诱导的，从而导致细胞增殖和/或存活的增加。这将增加增殖细胞池的大小，作为可以终末分化为肌成纤维细胞表型的中等分化群体。为了检验这一假说，本文提出了四个具体目标。首先，我们将研究肺成纤维细胞端粒酶的诱导机制。端粒酶基因及其5‘侧翼序列将被克隆用于转录调控分析。其次，将通过分析端粒酶和肌动蛋白基因表达的关系，以及这两个基因的转录调控机制是否在细胞分化方面具有功能意义的方式，来研究端粒酶表达和肌成纤维细胞表型之间的关系。第三，通过比较端粒长度与端粒酶在体内和体外的表达，以及端粒酶的表达是否赋予细胞对生长因子和/或凋亡信号的反应的不同特征，来分析端粒酶表达的功能意义。最后，通过使用端粒酶(端粒酶逆转录酶或端粒酶RNA组分)基因敲除的小鼠以及特定的端粒酶抑制剂，在博莱霉素模型中活体检测端粒酶在肺纤维化中的作用。通过使用这些结合的方法，将揭示这种端粒酶表达表型的潜在作用和意义的新见解，这可能被证明是有助于设计未来的新的治疗方法。

DESCRIPTION (provided by applicant): Increased numbers of fibroblasts are a key feature of pulmonary fibrosis, and their coalescence in densely populated cell formations, termed fibroblastic foci, is a diagnostic histopathological feature of idiopathic pulmonary fibrosis (IPF). Increased proliferation is a likely mechanism for this increase in fibroblasts whose phenotype is distinct from cells isolated from normal lung. Interestingly a new fibroblast phenotype is found to emerge in animal models of lung injury and fibrosis characterized by the expression of telomerase activity. While telomerase activity is associated with increased proliferative capacity and life span in certain cells during early development and many tumor cells, its significance in the pathogenesis of pulmonary fibrosis is unknown. In this project it is hypothesized that emergence de novo of a telomerase-expressing fibroblast phenotype is induced by certain factors resulting in increased cell proliferation and/or survival. This would increase the pool size of proliferative cells serving as an intermediate differentiated population that can terminally differentiate to the myofibroblast phenotype. Four specific aims are proposed to test this hypothesis. First, the mechanism of telomerase induction will be studied in lung fibroblasts. The telomerase gene and its 5'-flanking sequences will be cloned for analysis of transcriptional regulation. Second, the relationship between the telomerase expressing and myofibroblast phenotypes will be examined by analyzing the relationship between telomerase and (-smooth muscle actin gene expression, and whether the transcriptional regulatory mechanisms for both genes are somehow related in a functionally meaningful way in terms of cellular differentiation. Third, the functional significance of telomerase expression will be analyzed by comparing telomere length with telomerase expression in vivo and in vitro, and whether telomerase expression endow cells with differential characteristics with respect to response to growth factors and/or apoptotic signals. Finally, the role of telomerase in pulmonary fibrosis will be examined in vivo in the bleomycin model by use of telomerase (either telomerase reverse transcriptase or telomerase RNA component) knockout mice as well as specific telomerase inhibitors. By using these combined approaches, new insights into the potential role and significance of this telomerase expressing phenotype will be uncovered, which may prove to be useful for devising future novel therapeutic approaches.