Thursday, April 25, 2024
4/25/2024
DESCRIPTION (provided by applicant): Pulmonary granuloma formation is a key feature of sarcoidosis, a chronic disease of unknown etiology. Sarcoidosis has been linked to environmental risk factors (wood-burning stoves, fireplaces and firefighting) that favor carbon nanotube formation. We have shown that lungs instilled with multiwall carbon nanotubes (MWCNT) produce chronic inflammation and granuloma formation resembling sarcoidosis. Sarcoidosis alveolar macrophages exhibit elevated pro-inflammatory cytokines such as CCL5 and IFN-¿, but are deficient in PPAR¿, a lipid regulator that regulates the ABC transporters A1/G1 and dampens inflammation. This phenotype appears in macrophages from MWCNT-instilled mice. In macrophage-specific PPAR¿-KO mice, inflammatory mediators and granulomatous disease are exacerbated, suggesting that PPAR¿ is protective. Preliminary data indicate elevated microRNA (miR)-33 in macrophages and granulomatous tissues from both sarcoid patients and MWCNT-instilled mice. The role of miR-33 in the lung is unknown, but elsewhere miR-33 is shown to increase inflammation and repress ABCA1 and ABCG1 lipid transporters (which have anti- inflammatory properties). Strikingly, both transporters are depressed in macrophages from sarcoid patients and MWCNT-instilled mice. Further, in vitro miR-33 overexpression in wild type alveolar macrophages decreases ABCA1 and ABCG1 but not PPAR¿. These results suggest that two pathways may dysregulate transporters in granulomatous disease - one associated with intrinsic PPAR¿ status and the other via environmentally triggered miR-33. We hypothesize that granuloma-protective pathways of PPAR¿ are mediated by anti-inflammatory actions of ABCA1 and/or ABCG1. Aim 1 will determine functional significance of lipid transporters in granulomatous disease. MWCNT will be instilled in macrophage-specific KO models of: PPAR¿, ABCA1, ABCG1, and ABCA1/ABCG1 (double KO) plus wild-type mice. Sixty days later, untreated, sham-(vehicle) treated, or MWCNT-instilled mice will be examined for granuloma size/frequency, and alveolar macrophage expression of ABCA1, ABCG1, miR-33, CCL5, IFN¿, PPAR¿, and intracellular cholesterol. Aim 2 will explore the role of miR-33 as a mediator in granulomatous disease and alveolar macrophage expression of ABCA1 and ABCG1. MWCNT-instilled wild-type and macrophage-specific PPAR¿ KO mice will receive lenti-control, lenti-miR-33, or lenti-miR-33 antagomir. Granuloma size/frequency and alveolar macrophage markers will be evaluated. Subaims will examine miR-33 and lipid transporters in murine and healthy human alveolar macrophages cultured with MWCNT. Aim 3 will determine whether ABCA1, ABCG1, and miR-33 are intrinsically dysregulated in sarcoid alveolar macrophages compared to healthy controls. Subaims will examine effects of lenti-ABCA1, lenti-ABCG1, or lenti-miR-33 antagomir on alveolar macrophages. Our murine MWCNT granuloma model provides a unique opportunity for defining mechanisms and components perpetuating chronicity in environmental lung disease.
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