Figure 4

APOA4-stimulated glucose uptake was mediated by LRP1 in 3T3-L1 mature adipocytes. (A) APOA4 stimulated glucose uptake by 22–44% in a dose-dependent manner. 3T3-L1 mature adipocytes were incubated with 0, 1, and 2 μM r-m-APOA4 for 1 h (n = 5 per treatment). (B) SiRNA targeting Lrp1 gene was transfected into 3T3-L1 mature adipocytes. Left, a representative western blot image showing reduced LRP1 levels in Lrp1 siRNA-treated cells, compared to control siRNA-treated cells. GAPDH was used as internal control. Right, quantification of the band intensity revealed ~ 75% decrease in average band intensity for LRP1 protein in Lrp1 siRNA compared to control siRNA. *p < 0.05, Student’s t-test. (C) Lrp1 knockdown abrogated 1 µM and 2 µM APOA4-stimulated glucose uptake by ~ 93% and 67%, respectively, compared with 1 and 2 µM controls. Data are presented as percentage increase of basal glucose uptake levels under control siRNA treatment. (D) Insulin-stimulated glucose uptake increased in adipocytes at 5 nM and 100 nM versus untreated. APOA4 (1 µM) treatment further augmented insulin-stimulated glucose uptake at 5 nM, suggesting they act synergistically to promote glucose uptake. Two-way analysis of variance (ANOVA) revealed significant main effects of Apo-A-IV concentration (p < 0.0001) and siRNA treatment (p < 0.0001) as well as interaction between treatments (p = 0.0007). Sidak’s multiple comparisons test revealed statistically significant changes. ****p < 0.0001, 0 µM vs. 1 and 2 µM (Control), and ***p < 0.001, 1 µM and 2 µM control vs. 1 µM and 2 µM Lrp1 siRNA, respectively (n = 4–5/treatment).