Adiponectin and Liver harvested from AKO mice liver (Figure 3, A and B). To further corroborate the cell type-specific effect of adiponectin, human umbilical vein endothelial cells (HUVEC), hepatoma H4IIE, macrophage RAW 264.7 and stellate LX-2 cell lines were treated with or without adiponectin. QPCR analysis revealed that adiponectin significantly increased UCP2 mRNA level by ,3.5- and ,1.8-fold in HUVEC and LX-2, respectively (Figure 3C). Consistent with the in vivo data, the UCP2 expression in hepatoma cell H4IIE and macrophage cell RAW 264.7 was not affected by adiponectin (Figure 3C). Adiponectin enhanced the mRNA stability of UCP2 and promoted hnRNP K translocation to mitochondria To investigate the mechanism underlying adiponectin-induced UCP2 expression, PBS or adiponectin was administered into the portal vein of AKO mice in the presence or absence of transcriptional inhibitor actinomycin D (ActD) or protein synthesis inhibitor cycloheximide (CHX) (Figure 4). ActD treatment did not block the effects of adiponectin on UCP2 expression. In the presence of ActD, adiponectin increased the UCP2 protein and mRNA levels by ,2.3- and ,3.2-fold, respectively (Figure 4, A and B). The elevation of UCP2 protein expression was observed in both PC and NPC fractions. On the other hand, CHX completely suppressed the stimulatory effect of adiponectin on UCP2 protein expression, which was measured in the lysates derived from both the AKO liver tissues and the NPC fractions (Figure 4, A and D). However, CHX did not prevent adiponectin-induced elevation of UCP2 mRNA expression (Figure 4B). Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is involved in regulating UCP2 mRNA stability and protein synthesis [32]. While the total hnRNP K protein expression was not Figure 4. Adiponectin enhances the mRNA stability of UCP2 and promotes its protein synthesis. Two inhibitors, actinomycin D (ActD, A, B and C) or cycloheximide (CHX, A, B and D), were administered together with or without adiponectin protein into liver tissues of AKO mice livers. The protein (A) and mRNA (B) abundance of UCP2 was monitored by Western blotting and QPCR, respectively. The relative protein expression was also monitored in PCs/NPCs lysates (C and D). *, P,0.05 vs corresponding controls, n = 3. doi:10.1371/journal.pone.0032349.g004 PLoS ONE | www.plosone.org 4 February 2012 | Volume 7 | Issue 2 | e32349