Abstract
BackgroundMetabolic syndrome is characterized by macrophage infiltration and inflammatory responses—metaflammation in adipose tissue. IL-6 and IL-1β could mediate the inflammatory responses in macrophage stimulated-preadipocytes by modulating MAPK and NF-κB pathways. To test this hypothesis we used antibodies to block IL-6 and IL-1β action in macrophage conditioned medium (MacCM)-stimulated human white preadipocytes. Moreover, as interventions that prevent this could potentially be used to treat or prevent metabolic syndrome, and 1α,25(OH)2D3 has previously been reported to exert an anti-inflammatory action on macrophage-stimulated adipocytes, in this study we also investigated whether 1α,25(OH)2D3 could attenuate inflammatory responses in MacCM-stimulated preadipocytes, and explored the potential anti-inflammatory mechanisms.MethodsHuman white preadipocytes were cultured with 25% MacCM for 24 h to elicit inflammatory responses. This was confirmed by measuring the concentrations and mRNA levels of major pro-inflammatory factors [IL-1β, IL-6, IL-8, monocyte chemoattractant protein (MCP)-1 and regulated on activation, normal T cell expressed and secreted (RANTES)] by ELISA and qPCR, respectively. IL-6 and IL-1β actions were blocked using IL-6 antibody (300 ng/ml) and IL-1β antibody (15 μg/ml), respectively. Potential anti-inflammatory effects of 1α,25(OH)2D3 were investigated by pre-treatment and treatment of 1α,25(OH)2D3 (0.01 to 10 nM) for 48 h in MacCM-stimulated preadipocytes. In parallel, western blotting was used to determine inflammatory signaling molecules including relA of the NF-κB pathway and p44/42 MAPK modified during these processes.ResultsMacCM enhanced the secretion and gene expression of IL-1β, IL-6, IL-8, MCP-1 and RANTES by increasing the phosphorylation levels of relA and p44/42 MAPK in preadipocytes, whereas blocking IL-6 and IL-1β action inhibited the inflammatory responses by decreasing p44/42 MAPK and relA phosphorylation, respectively. Furthermore, 10 nM of 1α,25(OH)2D3 generally inhibited the IL-6 and IL-1β-mediated inflammatory responses, and reduced both p44/42 MAPK and relA phosphorylation in MacCM-stimulated preadipocytes.Conclusions1α,25(OH)2D3 attenuates IL-6 and IL-1β-mediated inflammatory responses, probably by inhibiting p44/42 MAPK and relA phosphorylation in MacCM-stimulated human white preadipocytes.
Highlights
Metabolic syndrome is characterized by macrophage infiltration and inflammatory responses— metaflammation in adipose tissue
Cytokine release was screened using proteome array and the results revealed that (Fig. 1a, b) pro-inflammatory factors including IL-6, IL-8, monocyte chemoattractant protein-1 (MCP-1) and RANTES were secreted from macrophage conditioned medium (MacCM)-stimulated preadipocytes, compared to controls
The initial results show that in human white preadipocytes, MacCM massively enhanced the secretion and gene expression of major pro-inflammatory factors including IL-1β, IL-6, IL-8, monocyte chemoattractant protein (MCP)-1 and RANTES, which have been measured as indicators of adipose tissue metaflammation in keeping with published studies [14, 24, 28,29,30,31]
Summary
Metabolic syndrome is characterized by macrophage infiltration and inflammatory responses— metaflammation in adipose tissue. Metaflammation in adipose tissue, characterized by infiltration of macrophages, local gene expression and secretion of pro-inflammatory factors, especially IL-6 and IL-1β [2], is considered a potential factor contributing to development of the metabolic syndrome [1]. Accumulated evidence suggests that macrophages play a key role in influencing the proliferation, survival and differentiation of preadipocytes [6,7,8,9,10], which are essential in maintaining adipose tissue homeostasis [11] Most importantly they might even induce or aggravate metaflammation by stimulating preadipocytes to express and secrete a variety of cytokines, most of which are pro-inflammatory [12,13,14]. The anti-inflammatory effects of 1α,25(OH)2D3 on macrophage-stimulated preadipocytes, remains to be explored in detail
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