Abstract
OBJECTIVE: Inflammatory activity in fat tissue has recently been implicated in mechanisms of insulin resistance and obesity-related metabolic dysfunction. Toll-like receptors (TLRs) play a key role in innate immune responses and recent studies implicate the TLR pathway in mechanisms of inflammation and atherosclerosis. The aim of this study was to examine differential TLR expression and function in human adipose tissue. METHODS AND PROCEDURES: We biopsied subcutaneous abdominal fat from 16 obese subjects (age 39+/-11 years, BMI 49+/-14 kg/m2) and characterized TLR expression using quantitative real-time PCR and confocal immunofluorescence imaging. In tissue culture, we stimulated isolated human adipocytes with Pam3CSK4 and lipopolysaccharide (LPS) (TLR2 and TLR4 agonists, respectively) and quantified TLR activity, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) production, and nuclear factor-kappaB (NF-kappaB) p65 nuclear activation using real-time PCR, enzyme-linked immunosorbent assay (ELISA), and immunofluorescence. RESULTS: TLR1, 2, and 4 protein colocalized with adiponectin in human adipocytes with TLR4 exhibiting the highest immunohistochemical expression. Using real-time PCR, we confirmed higher level of gene expression for TLR4 as compared to other members of the TLR family (TLR1, 2, 7, 8) in human adipose depots (P<0.001). In tissue culture, adipocyte TLR2/TLR4 mRNA expression and protein increased significantly following Pam3CSK4 and LPS (P<0.001). TLR2/TLR4 stimulation was associated with NF-kappaB p65 nuclear translocation and proinflammatory cytokine production. DISCUSSION: The findings demonstrate that TLRs are inducible in adipose tissue and linked with downstream NF-kappaB activation and cytokine release. Adipose stores may play a dynamic role in the regulation of inflammation and innate immunity in human subjects via modulation of the TLR/NF-kappaB regulatory pathway.