LETTERSLeptin is an adipocyte-derived hormone that plays a key role inenergy homeostasis, yet resistance to leptin is a feature ofmost cases of obesity in humans and rodents. In vitro analysissuggested that the suppressor of cytokine signaling-3 (Socs3)is a negative-feedback regulator of leptin signaling involved inleptin resistance. To determine the functional significance ofSocs3 in vivo, we generated neural cell−specific SOCS3conditional knockout mice using the Cre–loxP system.Compared to their wild-type littermates, Socs3-deficient miceshowed enhanced leptin-induced hypothalamic Stat3 tyrosinephosphorylation as well as pro-opiomelanocortin (POMC)induction, and this resulted in a greater body weight loss andsuppression of food intake. Moreover, the Socs3-deficient micewere resistant to high fat diet−induced weight gain andhyperleptinemia, and insulin-sensitivity was retained. Thesedata indicate that Socs3 is a key regulator of diet-inducedleptin as well as insulin resistance. Our study demonstrates thenegative regulatory role of Socs3 in leptin signaling in vivo,and thus suppression of Socs3 in the brain is a potentialtherapy for leptin-resistance in obesity.It was initially proposed that leptin has a primary role as an anti-obesity hormone; however, high concentrations of serum leptin levelsare often associated with obesity, suggesting the presence of leptinresistance1−3.The leptin receptor is highly expressed in the hypothala-mus and belongs to the cytokine-receptor superfamily that activatesthe Janus kinase−signal transducers and activators of transcription(Jak-Stat) pathway4. Stat3 is apparently essential for leptin regulationof energy balance5−7.Several mechanisms for leptin resistance havebeen documented, including impaired leptin transport across theblood-brain barrier8and the presence of negative regulators of leptinsignaling, such as the tyrosine phosphatases SH2-containing proteintyrosine phospatase-2 (SHP-2) (ref. 9) and protein-tyrosine phos-phatase 1B (PTP1B)10−12as well as the suppressors of cytokine signal-ing family of molecules13.Peripheral leptin administration to micerapidly induces Socs3 mRNA in hypothalamic regions14.Socs3 bindsto the phosphorylated leptin receptor through its Src homology-2(SH2) domain and inhibits Jak tyrosine kinase activity through its N-terminal kinase inhibitory region, which functions as a pseudo-substrate13,15,16.Furthermore, the C-terminal Socs box recruits theubiquitin-transferase system and may mediate the degradation ofreceptor-Jak complexes17.Thus, Socs3 may be an important regulatorof leptin signaling and could be involved in leptin-resistance. However,because these conclusions were based on the analysis of cell lines, thephysiological significance of Socs3 in leptin signaling and its impacton obesity remained an open question.Because Socs3-deficient mice die during embryonic development asa result of placental defects18,a conditional knockout approach wasused to determine the tissue-specific roles of Socs3 (ref. 19; Fig. 1a). Togenerate a neural cell−specific deletion, cre transgenic mice, in whichCre recombinase is expressed specifically in neural precursor cellsunder the nestin promoter-enhancer (crenes)20or synapsin promoter(cresyn)21,were crossed with Socs3flox/floxmice. Genomic DNAextracted from the brain was analyzed with PCR to verify deletion ofSocs3.A 250−base pair band corresponding to the deleted Socs3 allele(Socs3del) appeared in the PCR analysis of brain DNA fromcrenesSocs3flox/flox(NesKO) mice and cresynSocs3flox/flox(SynKO) micebut not in the DNA from control Socs3flox/flox(wild type) mice (Fig. 1b). In these two types of conditional knockout mice, we con-firmed Socs3 deletion in the hypothalamus and cerebrum, but not inother tissues including heart, liver, kidney muscle and fat(Supplementary Fig. 1 online). RT-PCR analysis also indicated thatSocs3 expression was greatly reduced in the brain but not in the othertissues examined (Supplementary Fig. 1 online).After leptin administration, Socs3 is exclusively expressed in thearcuate nucleus (ARC) of the hypothalamus, which coexpress as leptinreceptors and neuropeptides that are considered to be essential in theregulation of food intake14.Consistent with previous reports, our realtime RT-PCR analysis revealed that leptin treatment led to a twofoldincrease in Socs3 mRNA in wild-type mice (Fig. 1c). However, expres-sion of Socs3 mRNA in NesKO and SynKO mice was much lower thanthat in wild-type mice, irrespective of leptin administration (Fig. 1c).Immunohistological analysis with an antibody to Socs3 confirmed amarked induction of Socs3 protein in the ARC of the brain in 1Division of Molecular and Cellular Immunology, Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.2Department of Molecular Genetics, Institute of Life Science, Kurume University, 2432-3 Aikawa-machi, Kurume 839-0861, Japan. 3Institute of Molecular Medicineand Department of Medicine, University of California San Diego, 9500 Gilman Drive, La Jolla, California 92093-0641, USA. 4Cardiovascular Research Institute andThe Third Department of Internal Medicine, Kurume University, 67 Asahi-machi, Kurume 830-0011, Japan. Correspondence should be addressed to A. Y.([email protected]).Published online 20 June 2004; doi:10.1038/nm1071Socs3 deficiency in the brain elevates leptin sensitivity andconfers resistance to diet-induced obesityHiroyuki Mori1,Reiko Hanada2,Toshikatsu Hanada1, Daisuke Aki1,Ryuichi Mashima1,Hitomi Nishinakamura1,Ta ke hiro Torisu1,Kenneth R Chien3,Hideo Ya s u k a w a4& Akihiko Yoshimura1NATURE MEDICINE VOLUME 10 |NUMBER 7 |JULY 2004 739© 2004 Nature Publishing Group http://www.nature.com/naturemedicineLETTERSwild-type mice after the leptin injection, but only a marginal level ofinduction in the two brain-specific Socs3-deficient mice (Fig. 1d).To clarify the effects of Socs3 deficiency on leptin-induced Stat3 acti-vation, recombinant leptin was injected into mice and the hypothala-mus was examined with immunoblotting and immunofluorescencestaining with phospho-specific (Tyr705) Stat3 antibodies. Stat3 phos-phorylation in the hypothalamus was evident within1 h, after leptininjection but decreased thereafter in wild-type mice (Fig. 2a,b).However, Stat3 activation 1 h after leptin stimulation was significantlygreater in both NesKO and SynKO mice com-pared to their control littermates, and