Defining the Transcriptional Targets of Leptin Reveals a Role for Atf3 in Leptin Action.

Leptin acts via its receptor (LepRb) to modulate gene expression in hypothalamic LepRb-expressing neurons, thereby controlling energy balance and glucose homeostasis. Despite the importance of the control of gene expression in hypothalamic LepRb neurons for leptin action, the transcriptional targets of LepRb signaling have remained undefined because LepRb cells contribute a small fraction to the aggregate transcriptome of the brain regions in which they reside. We thus employed translating ribosome affinity purification followed by RNA sequencing to isolate and analyze mRNA from the hypothalamic LepRb neurons of wild-type or leptin-deficient (Lepob/ob) mice treated with vehicle or exogenous leptin. Although the expression of most of the genes encoding the neuropeptides commonly considered to represent the main targets of leptin action were altered only following chronic leptin deprivation, our analysis revealed other transcripts that were coordinately regulated by leptin under multiple treatment conditions. Among these, acute leptin treatment increased expression of the transcription factor Atf3 in LepRb neurons. Furthermore, ablation of Atf3 from LepRb neurons (Atf3LepRbKO mice) decreased leptin efficacy and promoted positive energy balance in mice. Thus, this analysis revealed the gene targets of leptin action, including Atf3, which represents a cellular mediator of leptin action.

Tanshinone IIA attenuates cyclic strain-induced endothelin-1 expression in human umbilical vein endothelial cells.

1. Tanshinone IIA, one of the active components of the Radix of Salvia miltiorrhiza, is used in traditional Chinese medicine to treat cardiovascular diseases. However, the intracellular mechanism of action of tanshinone IIA remain to be determined. The aims of the present study were to test the hypothesis that tanshinone IIA alters strain-induced endothelin (ET)-1 expression and nitric oxide (NO) production, as well as to identify the putative signalling pathways involved, in human umbilical vein endothelial cells (HUVEC). 2. Cultured HUVEC were exposed to cyclic strain in the presence of 1-10 µmol/L tanshinone IIA. Expression of ET-1 was examined by reverse transcription-polymerase chain reaction and ELISA. Phosphorylation of endothelial NO synthase (eNOS) and activating transcription factor (ATF) 3 was assessed by western blot analysis. 3. Tanshinone IIA (3 and 10 µmol/L) inhibited strain-induced ET-1 expression. In contrast, NO production, eNOS phosphorylation and ATF3 expression were enhanced by tanshinone IIA. The eNOS inhibitor N(G) -nitro-L-arginine methyl ester (l-NAME; 100 µmol/L), the phosphatidylinositol 3-kinase inhibitor LY294002 (5 µmol/L) and the soluble guanylyl cyclase inhibitor 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one (ODQ; 10 µmol/L) inhibited tanshinone IIA-induced increases in ATF3 expression. Moreover, treatment of HUVEC with either an NO donor (3,3-bis [aminoethyl]-1-hydroxy-2-oxo-1-triazene; 500 µmol/L) or an ATF3 activator (carbobenzoxy-L-leucyl-L-leucyl-L-leucinal; 5 µmol/L) resulted in the repression of strain-induced ET-1 expression. The inhibitory effect of tanshinone IIA on strain-induced ET-1 expression was significantly attenuated by l-NAME, ODQ and the transfection of small interfering RNA for ATF3. 4. In conclusion, tanshinone IIA inhibits strain-induced ET-1 expression by increasing NO and upregulating ATF3 in HUVEC. The present study provides important new insights into the molecular pathways that may contribute to the beneficial effects of tanshinone IIA in the cardiovascular system.