Lung granulomas are associated with numerous conditions, including inflammatory disorders, exposure to environmental pollutants, and infection. Osteopontin is a chemotactic cytokine produced by macrophages, and is implicated in extracellular matrix remodeling. Furthermore, osteopontin is up-regulated in granulomatous disease, and osteopontin null mice exhibit reduced granuloma formation. Animal models currently used to investigate chronic lung granulomatous inflammation bear a pathological resemblance, but lack the chronic nature of human granulomatous disease. Carbon nanoparticles are generated as byproducts of combustion. Interestingly, experimental exposures to carbon nanoparticles induce pulmonary granuloma-like lesions. However, the recruited cellular populations and extracellular matrix gene expression profiles within these lesions have not been explored. Because of the rapid resolution of granulomas in current animal models, the mechanisms responsible for persistence have been elusive. To overcome the limitations of previous models, we investigated whether a model using multiwall carbon nanoparticles would resemble chronic human lung granulomatous inflammation. We hypothesized that pulmonary exposure to multiwall carbon nanoparticles would induce granulomas, elicit a macrophage and T-cell response, and mimic other granulomatous disorders with an up-regulation of osteopontin. This model demonstrates: (1) granulomatous inflammation, with macrophage and T-cell infiltration; (2) resemblance to the chronicity of human granulomas, with persistence up to 90 days; and (3) a marked elevation of osteopontin, metalloproteinases, and cell adhesion molecules in granulomatous foci isolated by laser-capture microdissection and in alveolar macrophages from bronchoalveolar lavage. The establishment of such a model provides an important platform for mechanistic studies on the persistence of granuloma.

ContextThe NRLP3 inflammasome is a multiprotein danger-sensing complex that serves as a critical link between obesity-related adipose inflammation and insulin resistance and has been shown in animal models to be inhibited by fish oil-derived long chain omega-3 polyunsaturated fatty acids (n-3 PUFA).ObjectiveWe conducted a clinical trial and in vitro experiments to test our hypothesis that n-3 PUFA suppress NLRP3 inflammasome in human obesity through downregulation of inflammasome gene expression in adipocytes and macrophages.DesignPlacebo-controlled clinical trial and in vitro coculture experiments with primary human adipocytes (from biopsy specimens) and human THP-1 monocyte-derived macrophages treated with eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) vs vehicle control.SettingGeneral community, research laboratory.Patients and Other ParticipantsObese (body mass index ≥ 30 kg/m2), nondiabetic males and females age 18 to 50. N = 25.InterventionsClinical trial: Eight-week treatment with 4 g Lovaza (EPA and DHA) or placebo. Cells culture: EPA and/or DHA at 100 µg/mL or vehicle control in culture medium.Main Outcome MeasuresAdipose tissue or adipocyte/macrophage mRNA expression of IL-1β and IL-18 and circulating IL-18 levels.ResultsTreatment of obese human subjects with fish oil supplements reduced expression of adipose inflammatory genes including inflammasome-associated IL-18 and IL-1β and circulating IL-18 levels. Both EPA and DHA reduced inflammasome gene expression in obese human adipose and human adipocyte and macrophages.Conclusions N-3 PUFA reduce NLRP3 inflammasome in human adipose through downregulation of gene expression in adipocytes and monocytes/macrophages and has potential as nutritional therapeutic agent in prevention of obesity-related inflammation.

Background: Omega-3 polyunsaturated fatty acids, specifically the fish oil derived eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have been proposed as inflammation resolving agents via their effects on adipose tissue. Objective: We proposed to determine the effects of EPA and DHA on human adipocyte differentiation and inflammatory activation in vitro. Methods: Primary human subcutaneous adipocytes from lean and obese subjects were treated with 100μM EPA and/or DHA throughout differentiation (differentiation studies) or for 72 hours post-differentiation (inflammatory studies). THP-1 monocytes were added to adipocyte wells for co-culture experiments. Subcutaneous and visceral adipose explants from obese subjects were treated for 72 hours with EPA and DHA. Oil-RedO staining was performed on live cells. Cells were collected for mRNA analysis by qPCR and media collected for protein quantification by ELISA. Results: Incubation with EPA and/or DHA attenuated inflammatory response to LPS and monocyte co-culture with reduction in post-LPS mRNA expression and protein levels of IL6, CCL2, and CX3CL1. Expression of inflammatory genes was also reduced in the endogenous inflammatory response in obese adipose. Both DHA and EPA reduced lipid droplet formation and lipogenic gene expression without alteration in expression of adipogenic genes or adiponectin secretion. Conclusions: EPA and DHA attenuate inflammatory activation of in vitro human adipocytes and reduce lipogenesis.

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