Differential expression of extracellular matrix remodeling genes in a murine model of bleomycin-induced pulmonary fibrosis.

Exposure to the chemotherapeutic drug bleomycin leads to pulmonary fibrosis in humans and has been widely used in animal models of the disease. Using C57BL/6 bleomycin-sensitive mice, pulmonary fibrosis was induced by multiple intraperitoneal injections of the drug. An increase in the relative amounts of steady-state alpha1(I) procollagen, alpha1(III) procollagen, and fibronectin mRNA as well as histopathological evidence of fibrosis was observed. The effect of bleomycin on the expression of the enzymes responsible for extracellular matrix degradation, the matrix metalloproteinases (MMPs), and their inhibitors (TIMPs), was selective and showed temporal differences during the development of fibrosis. Of the MMPs tested, bleomycin treatment resulted in the up-regulation of gelatinase A and macrophage metalloelastase gene expression in whole-lung homogenates, whereas gelatinase B, stromelysin-1, and interstitial collagenase gene expression was not significantly changed. Timp2 and Timp3, the murine homologues of the respective TIMP genes, were constitutively expressed, whereas Timp1 was markedly up-regulated during fibrosis. The strong correlation between enhanced extracellular matrix gene expression, differential MMP and TIMP gene expression, and histopathological evidence of fibrosis suggest that dysregulated matrix remodeling is likely to contribute to the pathology of bleomycin-induced pulmonary fibrosis.

E1A-induced immortalization of rat type II alveolar epithelial cells.

Using a retroviral vector expressing the adenoviral 12S E1A gene product the authors have immortalized rat type II alveolar epithelial cells. For a period of time, the immortalized cells retain many of the ultrastructural characteristics of type II cells in situ, including the presence of lamellar bodies. By 250 days in culture, however, neither lamellar bodies, SP-A, nor a phospholipid profile characteristic of surfactant were present. The cell bind the lectin Maclura pomifera and stably express cytokeratins and the E1A gene product. The cell line also has a diploid karyotype, exhibits contact inhibition of growth, and does not grow in soft agar. E1A-immortalized cell lines should prove useful as models for study of certain aspects of type II alveolar epithelial cell function.

Relative release of interleukin-1 beta and interleukin-1 receptor antagonist by alveolar macrophages. A study in asbestos-induced lung disease, sarcoidosis, and idiopathic pulmonary fibrosis.

We examined the influence of untreated interstitial lung disease (ILD) on the in vitro release of interleukin-1 beta (IL-1 beta) and interleukin-1 receptor antagonist (IL-Ira) from alveolar macrophages (AM); AM were harvested from normal volunteers, ILD patients, and patients with asbestos-related pleural disease but no ILD. AM were cultured for 24 h and assays for IL-1 beta and IL-1ra were done using sensitive and specific enzyme-linked immunosorbent assay. A greater amount of IL-1 beta was detected in AM supernatants from asbestosis, sarcoidosis, and IPF patients than in those from normal subjects. The IL-1 beta:IL-1ra ratio (IL-1 beta activity index [IL-1AI]) was significantly lower in supernatants of normal macrophages compared with macrophage supernatants from individuals with ILD. The IL-1AI correlated with bronchoalveolar lavage cellularity, a marker of disease activity. Current smoking was associated with lower IL-1 beta and IL-1ra release in ILD. The IL-1AI is a convenient method for comparison of IL-1 beta activity between patient populations.

Isolation and immortalization of rat pre-type II cell lines.

The fetal respiratory distress syndrome is due, in part, to the presence of abundant pre-type II alveolar epithelial cells that have not yet differentiated into mature type II cells. Studies of this syndrome have been limited somewhat by the lack of an adequate in vitro model. In the present study we immortalized pre-type II cells by infecting primary isolates obtained from fetal rat lung with a retroviral construct expressing the adenoviral 12S E1A gene product. The immortalized pre-type II cells retained many of the ultrastructural features typical of pre-type II cells in primary culture, most notably lamellar bodies were not detected and the cells contained abundant stores of glycogen, expressed cytokeratin filaments, and bound the lectin Maclura pomifera. Karyotyping revealed that the cells are diploid. Growth studies demonstrate log phase growth in the presence of serum with a markedly decreased growth rate shortly after the cells reach confluence. Exposure of the immortalized pre-type II cells to hydrocortisone and dibutyryl cAMP resulted in the induction of lamellar bodylike organelles; however, these cells did not secrete surfactant or express surfactant protein A. These cells may serve as useful models for some in vitro studies of fetal type II cell maturation or the fetal respiratory distress syndrome, or both.