TIR-Domain-Containing Adaptor-Inducing Interferon- β (TRIF) Is Involved in Glucose Metabolism in Adipose Tissue through the Insulin/AKT Signaling Pathway

Abstract

Obesity significantly increases the risk of developing type 2 diabetes mellitus and other metabolic diseases. Obesity is associated with chronic low-grade inflammation in white adipose tissues, which is thought to play an essential role in developing insulin resistance. Many lines of evidence indicate that toll-like receptors (TLRs) and their downstream signaling pathways are involved in development of chronic low-grade inflammation and insulin resistance, which are associated with obesity. Mice lacking molecules positively involved in the TLR signaling pathways are generally protected from high-fat diet-induced inflammation and insulin resistance. In this study, we have determined the effects of genetic deficiency of toll/interleukin-1 receptor-domain-containing adaptor-inducing interferon-β (TRIF) on food intake, bodyweight, glucose metabolism, adipose tissue macrophage polarization, and insulin signaling in normal chow diet-fed mice to investigate the role of the TRIF-dependent TLR signaling in adipose tissue metabolism and inflammation. TRIF deficiency (TRIF^-/-) increased food intake and bodyweight. The significant increase in bodyweight in TRIF^-/- mice was discernible as early as 24 weeks of age and sustained thereafter. TRIF^-/- mice showed impaired glucose tolerance in glucose tolerance tests, but their insulin tolerance tests were similar to those in TRIF^+/+ mice. Although no difference was found in the epididymal adipose mass between the two groups, the percentage of CD206⁺ M2 macrophages in epididymal adipose tissue decreased in TRIF^-/- mice compared with those in TRIF^+/+ mice. Furthermore, activation of epididymal adipose AKT in response to insulin stimulation was remarkably diminished in TRIF^-/- mice compared with TRIF^+/+ mice. Our results indicate that the TRIF-dependent TLR signaling contributes to maintaining insulin/AKT signaling and M2 macrophages in epididymal adipose tissue under a normal chow diet and provide new evidence that TLR4-targeted therapies for type 2 diabetes require caution.

Figure 1

Effects of TRIF deficiency on bodyweight, food consumption, and plasma leptin levels. (a) Animals were weighed weekly from 16 weeks of age through 44 weeks. (b) Food consumption was recorded weekly from 36 weeks through 40 weeks. (c) Plasma samples were harvested at the age of 48 weeks, and plasma leptin was determined by leptin ELISA kit ( ^∗P < 0.05,  ^##P < 0.001) (n = 12/group).

Figure 2

Effects of TRIF deficiency on the IPGTT and ITT. (a) At the age of 46 weeks, mice were fasted for 16 h and injected i.p. with 1.5 g glucose/kg bodyweight, and (b) blood glucose and insulin were measured before and after glucose injection at the indicated time points. (c) At the age of 47 weeks, mice were fasted for 4 h and injected i.p. with 0.75 U insulin/kg bodyweight for the ITT. Blood glucose levels were determined and are shown as percent changes from (d) basal blood glucose levels ( ^∗P < 0.05,  ^##P < 0.001) (n = 12/group).

Figure 3

Effects of TRIF deficiency on macrophages in adipose tissue. (a) Flow cytometer analyses showing expression levels of CD206⁺ F4/80⁺ M2 and CD11c⁺ F4/80⁺ M1 macrophages in epididymal adipose tissue in wild-type mice (b) and TRIF KO mice. (c) Bar graphs present percentages of CD206⁺ F4/80⁺ M2 macrophages in F4/80⁺ macrophages. (d) The weights of right epididymal adipose tissue ( ^∗P < 0.05) (n = 4/group).

Figure 4

Effects of TRIF deficiency on activation of adipose tissue and liver AKT by insulin administration in mice. At 48 weeks of age, mice were fasted for 16 h and then intraperitoneally administered with PBS (a, c, e) or insulin (5 U/kg bodyweight) (b, d, f). Epididymal adipose tissue, muscle, and liver were collected 10 min after the administration. Levels of phospho-AKT (S473, P-Akt) and total-AKT (T-AKT) in adipose tissue (a, b), muscle (c, d), and liver (e, f) were determined by western blotting. (g–i) Bar graphs represent the results of densitometric analysis of western blots shown in (a) and (b), in (c) and (d), and in (e) and (f), respectively ( ^∗P < 0.05,  ^##P < 0.001) (n = 4/group).