Investigation of p21 in the molecular mechanisms of lung injury and pulmonary fibrosis

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1.Transforming Growth Factor-β1 as an Enhancer of Fas-mediated Apoptosis of Lung Epithelial CellsTransforming growth factor-β1(TGF-β1) has important roles in lung fibrosis and the potential to induce apoptosis in several types of cells. We previously demonstrated that apoptosis of lung epithelial cells induced by Fas ligation may be involved in the development of pulmonary fibrosis. Here we show that TGF-β1 induces apoptosis of primary cultured bronchiolar epithelial cells via caspase-3 activation and downregulation of cyclin-dependent kinase inhibitor p21. Concentrations of TGF-β1 that were not sufficient to induce apoptosis alone could enhance agonistic anti-Fas antibody or recombinant Fas ligand-mediated apoptosis of cultured bronchiolar epithelial cells. Soluble Fas ligand in the bronchoalveolar lavage fluid(BALF) from patients with idiopathic pulmonary fibrosis(IPF) also induced apoptosis of cultured bronchiolar epithelial cells that was significantly attenuated by anti-TGF-β anti … More body. Otherwise, BALF from patients with hypersensitivity pneumonitis(HP) could not induce apoptosis on bronchiolar epithelial cells despite its comparable amounts of soluble Fas ligand. The concentrations of TGF-β1 in BALF from patients with IPF were significantly higher compared with those in BALF from patients with HP or controls. Furthermore, co-incubation with the low concentration of TGF-β1 and HP BALF created pro-apoptotic effects comparable to the IPF BALF. In vivo, the administration of TGF-β1 could enhance Fas-mediated epithelial cell apoptosis and lung injury via caspase-3 activation in mice. Our results demonstrate a novel role of TGF-β1 in the pathophysiology of pulmonary fibrosis as an enhancer of Fas-mediated apoptosis of lung epithelial cells.2.In vivo gene transfer of mutant MCP-1 attenuates pulmonary fibrosisAlveolar epithelial cells are known to be present at the primary site of lung damage in pulmonary fibrosis. Apoptosis has been implicated as being involved in epithelial cell damage and pulmonary fibrosis. Since the cyclin-dependent kinase inhibitor p21 induces G1 arrest and DNA repair, and since it also prevents apoptosis in some cells, we hypothesized that p21 gene transfer may attenuate bleomycin-induced pulmonary fibrosis in mice, the pathogenesis of which likely involves epithelial cell apoptosis. Human p21 protein was expressed in mouse alveolar epithelial cells at 1 to 7 days in vitro and was detected predominantly in lung epithelial cells at 1 to 7 days in vivo after adenoviral transfer of the human p21 gene. Inflammatory cell infiltration and fibrosis had already begun at 7 days in this model. Adenoviral transfer of the human p21 gene at 7days after intratracheal instillation of bleomycin led to a decrease in the number of apoptotic cells, lung inflammation and fibrosis at 14 days. Therefore, the forced expression of p21 exerted both anti-apoptotic and anti-fibrotic effects, which would facilitate the ultimate goal of treatment for pulmonary fibrosis. Less