PPARγ Deficiency Exacerbates Fibrotic Response to Mycobacteria Peptide in Murine Sarcoidosis Model.

We established a murine model of multiwall carbon nanotube (MWCNT)-elicited chronic granulomatous disease which bears similarities to human sarcoidosis pathology including alveolar macrophage deficiency of peroxisome-proliferator-activated receptor gamma (PPARγ). Because lymphocyte reactivity to mycobacterial antigens has been reported in sarcoidosis, we hypothesized that addition of mycobacterial Early Secreted Antigenic Target Protein 6 (ESAT-6) to MWCNT might exacerbate pulmonary granulomatous pathology. MWCNT with or without ESAT-6 peptide-14 were instilled by oropharyngeal route into macrophage-specific PPARγ KO or wild-type mice. Controls received PBS or ESAT-6. Lung tissues, bronchoalveolar lavage (BAL) cells and fluid, were evaluated 60 days post instillation. PPARγ-KO mice receiving MWCNT+ESAT-6 had increased granulomas and significantly elevated fibrosis (trichrome staining) vs wild-type mice or PPARγ-KO mice receiving only MWCNT. Immunostaining of lung tissues noted elevated fibronectin and Siglec F expression on CD11c(+) infiltrating alveolar macrophages in the presence of MWCNT+ESAT-6 compared to MWCNT. Analyses of BALF proteins indicated increased levels of Transforming Growth Factor-β (TGF-β) and the TGF-β pathway mediator IL-13, in PPARγ-KO mice receiving MWCNT+ESAT-6 compared to wild-type or PPARγ KO receiving MWCNT. Similarly, mRNA levels of matrix metalloproteinase (MMP)-9, another requisite factor for TGF-β production, was elevated in PPARγKO by MWCNT+ESAT-6. Analysis of ESAT-6 in lung tissues by mass spectrometry revealed ESAT-6 retention in lung tissues of PPARγ-KO but not wild-type mice. Data indicate that PPARγ deficiency promotes pulmonary ESAT-6 retention, exacerbates macrophage responses to MWCNT+ESAT-6, and intensifies pulmonary fibrosis. Findings suggest that the model may facilitate understanding of the effects of environmental factors on sarcoidosis-associated pulmonary fibrosis.