Human lung myofibroblast-derived inducers of alveolar epithelial apoptosisidentified as angiotensin peptides

Earlier work from this laboratory found that fibroblasts isolated from fibr otic human lung [human interstitial pulmonary fibrosis (HIPF)] secrete a so luble inducer(s) of apoptosis in alveolar epithelial cells (AECs) in vitro [B. D. Uhal, I. Joshi, A. True, S. Mundle, A. Raza, A. Pardo, and M. Selman . Am. J. Physiol. 269 (Lung Cell. Mol. Physiol. 13): L819-L828, 1995]. The cultured human fibroblast strains most active in producing the apoptotic ac tivity contained high numbers of stellate cells expressing alpha-smooth mus cle actin, a myofibroblast marker. The apoptotic activity eluted from gel-f iltration columns only in fractions corresponding to proteins. Western blot ting of the protein fraction identified immunoreactive angiotensinogen (ANG EN), and two-step RT-PCR revealed expression of ANGEN by HIPF fibroblasts b ut not by normal human lung fibroblasts. Specific ELISA detected angiotensi n II (ANG II) at concentrations sixfold higher in HIPF-conditioned medium t han in normal fibroblast-conditioned medium. Pretreatment of the concentrat ed medium with purified renin plus purified angiotensin-converting enzyme ( ACE) further increased the ELISA-detectable ANG II eightfold. Apoptosis of AECs in response to HIPF-conditioned medium was completely abrogated by the ANG II receptor antagonist saralasin (50 mu g/ml) or anti-ANG II antibodie s. These results identify the protein inducers of AEC apoptosis produced by HIPF fibroblasts as ANGEN and its derivative ANG II. They also suggest a m echanism for AEC death adjacent to HIPF myofibroblasts [B. D. Uhal,, I. Jos hi, C. Ramos, A. Pardo, and M. Selman. Am. J. Physiol. 215 (Lung Cell. Mol. Physiol. 19): L1192-L1199, 1998].