黑色素浓集激素受体2（MCHR2）基因表达及其功能研究
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摘要
肥胖症是由于体脂过度堆积而产生的慢性疾病,是心脑血管疾病、糖尿病、高血压、高脂血症以及癌症等多种疾病的高危因素,现已成为全球性的健康问题。多年来研究发现肥胖的发生与下丘脑对摄食的神经内分泌调控密切相关,由下丘脑分泌的促食肽黑色素浓集激素(MCH)及其受体(MCHR1和MCHR2)的发现为肥胖发生机制及防治的研究提供了新的靶点。通过构建转基因和基因敲除动物模型已对MCH及MCHR1与肥胖及能量代谢的关系进行了深入细致的研究,但有关MCHR2的功能及其与肥胖的关系至今仍不清楚。因此,本课题应用DNA重组技术构建人MCHR2真核表达载体,并稳定转染哺乳动物细胞,分析其表达、定位及生物学活性,进而以3T3-L1前脂肪细胞为研究对象,探讨MCHR2是否具备调控能量平衡的生理功能,为深入研究MCHR2与肥胖的关系奠定基础。研究内容包括以下三个部分:
1. MCHR2真核表达载体的构建与鉴定。方法:以人胎脑cDNA文库为模板PCR扩增获取MCHR2基因全长cDNA编码区序列,定向插入到真核表达载体pcDNA3.1(+)中,并转化大肠杆菌,经抗生素平板筛选获得阳性克隆,再分别进行酶切、PCR及DNA测序鉴定。结果:获得了长为1023bp的人MCHR2 cDNA全长的目的片段,并插入到pcDNA3.1(+)载体的CMV启动子下游构建了表达MCHR2的重组质粒,酶切和PCR鉴定证实重组质粒带有目的基因片段,DNA测序确认插入片段的序列准确无误。结论:表达人MCHR2蛋白的真核表达载体pcDNA3.1-MCHR2构建成功。
2. MCHR2在CHO细胞中的稳定表达及其生物学活性的分析。方法:脂质体转染法将pcDNA3.1-MCHR2真核表达载体导入CHO细胞,经G418筛选抗性细胞克隆,建立稳定转染细胞系,RT-PCR、Western blot检测MCHR2转录水平和蛋白水平的表达,免疫荧光检测MCHR2在细胞中的定位,放射性配体结合实验检测受体的密度和亲和力,通过测定细胞内cAMP、Ca~(2+)浓度的改变分析MCHR2的信号转导通路。结果:经脂质体转染和G418抗性筛选6~8周,建立了稳定转染pcDNA3.1-MCHR2质粒的CHO细胞系;在该细胞系中MCHR2基因能被有效地转录和翻译,并准确定位于细胞膜上,与125I-MCH的结合具有特异、高亲和及可饱和的配体-受体结合特性,其最大结合容量Bmax值为309.97±1.14fmol/mg protein,平衡解离常数Kd值为0.170±0.0006 nmol/L;MCH作用于MCHR2后不影响细胞内cAMP生成,可促使细胞内钙库释放Ca~(2+)进而使胞内Ca~(2+)浓度出现明显而短暂的升高,Ca~(2+)振荡过程在MCH刺激后30~50s内完成,其对Ca~(2+)释放的作用呈剂量依赖关系,半数有效浓度EC50为2.32±0.01nmol/L,提示MCHR2的信号转导通路是与Gq蛋白偶联。结论:建立了稳定转染MCHR2的CHO细胞系,从MCHR2的表达、定位、受体活性及信号转导通路等方面证实了真核表达载体pcDNA3.1-MCHR2稳定转染哺乳动物细胞后能有效表达具有生物学活性的MCHR2蛋白,为后续体外研究MCHR2基因的功能提供了可靠的实验依据。
3. MCHR2对脂肪细胞生物学特性的影响。方法:通过脂质体转染和G418筛选建立稳定转染pcDNA3.1-MCHR2质粒的3T3-L1前脂肪细胞系,命名为3T3-L1-MCHR2细胞,作为对照的稳定转染空质粒的细胞命名为3T3-L1-mock细胞;RT-PCR、Western blot、免疫荧光检测、放射性配体结合实验及细胞内Ca~(2+)检测鉴定MCHR2的表达及活性;MTT法检测细胞增殖,FCM检测细胞周期分布;激素鸡尾酒法(MIX+DEX+insulin)诱导前脂肪细胞分化,油红O染色观察脂肪细胞分化的形态学改变及量化分化程度,RT-PCR检测脂肪细胞分化成熟的标记基因;检测成熟脂肪细胞中TG的生成、FFA及Leptin的分泌。结果:建立了稳定转染细胞系——3T3-L1-MCHR2,并通过了MCHR2表达和活性的鉴定;对前脂肪细胞增殖和分化特性作比较分析,可见MCH对3T3-L1-MCHR2和3T3-L1-mock细胞的增殖无影响,对两株细胞的分化则有一定的促进作用,两者相比,3T3-L1-MCHR2细胞的分化速度更快、程度更高,具体表现为油红O比色在分化第9天开始出现脂质聚集增多(3T3-L1-mock细胞为第12天),分化成熟标记基因PPARγ2、C/EBPα和aP2从分化第3天起上调,Leptin自分化第6天起上调,而3T3-L1-mock细胞的PPARγ2和Leptin于分化第6天、C/EBPα和aP2于分化第9天才上调,且增幅较弱;对成熟脂肪细胞的脂质代谢特性的分析显示,3T3-L1-MCHR2和3T3-L1-mock细胞在MCH作用下FFA的含量无明显差异,而TG的含量明显升高,升高率分别为22.61%和12.17%;ELISA法检测成熟脂肪细胞Leptin的分泌,3T3-L1-MCHR2细胞在MCH作用6h及24h后Leptin分泌明显增多,增幅分别为28.17%和18.25%,而3T3-L1-mock细胞则在24h后才增多,增幅为14.57%。结论:MCHR2可介导MCH对前脂肪细胞分化、脂肪合成代谢及Leptin分泌的正调控作用,对前脂肪细胞增殖和脂肪分解代谢则无影响,初步证实MCHR2的生理功能参与了能量平衡的调控。
Obesity, a chronic disease characterized by excessive accumulation of body fat, has become a global health problem and is a significant risk factor for developing many serious diseases, including cardio- and cerebro- vascular diseases, diabetes, hypertension, dyslipidemias and some cancers. In recent years, it has been well established that obesity is associated with the neuroendocrine control of the hypothalamus. The identification of the hypothalamic orexigenic neuropeptide melanin-concentrating hormone (MCH) and its two receptors (MCHR1 and MCHR2) provide new targets for treatment of obesity. Major advances have been made in identifying the relevance of MCH and MCHR1 to obesity and energy homeostasis by transgenic animal and gene knockout animal. However, the function of MCHR2 and its relevance to obesity is still indistinct. In this study, we first construct the eukaryotic expression vector of human MCHR2 by DNA recombination technique. Then, we stably transfect mammalian cell line with the vector and analyse the expression, localization and biologic activity of MCHR2. Finally, we investigate the physiological function of MCHR2 in 3T3-L1 preadipocyte to evaluate whether it is involved in mediating the effects of MCH on energy balance and its relevance to obesity. This study includes three parts:
1. Construction and identification of eukaryotic expression vector of human MCHR2. Methods: The full-length MCHR2 cDNA fragment was amplified by PCR from the human fetal brain cDNA library and was oriently inserted into eukaryotic expression vector pcDNA3.1(+). The positive clone was obtained after transformation and antibiotic screening and was identified by restriction endonuclease digestion, PCR, and DNA sequencing. Results: The 1023bp MCHR2 cDNA fragment was obtained and cloned into the downstream of CMV promoter of pcDNA3.1(+). The recombinant plasmid was identified by restriction endonuclease digestion and PCR. DNA sequencing confirmed that the sequence of the inserted element was correct. Conclusion: The eukaryotic expression vector pcDNA3.1-MCHR2 was successfully constructed.
2. Stably expression and biological activity of MCHR2 in CHO cells. Methods: CHO cells were transfected with the recombinant plasmids using lipofectamine. Stable cell line was established by G418 selection. Expression of MCHR2 in transcriptional level and protein level was detected by RT-PCR and western blot. Localization of MCHR2 in CHO cells was assayed by immunofluorence technique. Radioligand binding assays were performed to survey receptor density and affinity of MCHR2. Signal transduction pathways mediated by MCHR2 were analyzed by measurement of intracellular cAMP and calcium. Results: CHO cell line stably transfected with pcDNA3.1-MCHR2 plasmids was generated after liposome transfection and G418 selection. MCHR2 gene was efficiently transcribed and translated, and its protein exactly localized on the cytomembrane. MCHR2 could specific bind with ~(125)I-MCH with high-affinity and saturability (Bmax=309.97±1.14fmol/mg protein, Kd=0.170±0.0006nmol/L). MCH stimulation had no effect on the production of cAMP. MCH induced a clear and transient increase of intracellular calcium, and this procedure was completed within 30~50s. The effect of MCH on intracellular calcium showed dose-dependent response with an effective half-maximal concentration (EC50) of 2.32±0.01nmol/L. It is suggested that MCHR2 is only coupled to Gq protein. Conclusion: Stably transfected CHO cell line was established successfully. It is confirmed that MCHR2 expressed in mammalian cells is a functional receptor for the MCH by the analysis of its expression, localization, receptor characteristics, and signal pathways. This present study provides a reliable experimental foundation for further studies on the function of MCHR2 in vitro.
3. Influence of MCHR2 on the biological characteristics of adipocyte. Methods: 3T3-L1 cell line stably transfected with pcDNA3.1-MCHR2 plasminds was generated by transfection and G418 selection and named as 3T3-L1-MCHR2 while the control cell line transfected with pcDNA3.1(+) was named as 3T3-L1-mock. The expression and biological activity of MCHR2 were identified by RT-PCR, western blot, immunofluorence, radioligand binding and intracellular calcium assay. Cell proliferation was determined by MTT assay. Cell cycle was detected by flow cytometry. Cell differentiation was induced by hormone cocktail (MIX+DEX+insulin) and examined by Oil red O staining. The marker genes of adipocyte differentiation were examined by RT-PCR. The production of triglyceride and secretions of FFA and leptin were measured. Results: Stably transfected cell line named 3T3-L1-MCHR2 was established and passed the identification of expression and biological activity of MCHR2. MCH had no effect on the proliferation of 3T3-L1-MCHR2 and 3T3-L1-mock cells. Both the two cell lines were induced to differentiate to mature adipocytes by MCH. The speed and degree of differentiation in 3T3-L1-MCHR2 cells were higher than those in 3T3-L1-mock cells. Following was the detailed display of differentiation in 3T3-L1-MCHR2 cells: accumulation of lipid increased at day 9 (in 3T3-L1-mock it increased at day 12); marker genes of mature adipocyte including PPARγ2、C/EBPαand aP2 increased at day 3, leptin increased at day 6, while in 3T3-L1-mock cells, up-regulation of PPARγ2 and leptin appeared at day 6, up-regulation of C/EBPαand aP2 appeared at day 9, and the increasing levels were lower. There was no difference of the FFA secretion between MCH-treated group and control group. The triglyceride content in 3T3-L1-MCHR2 and 3T3-L1-mock mature adipocytes obviously increased by MCH, and the rising rate were 22.61% and 12.17%, compared with corresponding untreated cells. MCH stimulated leptin releases from 3T3-L1-MCHR2 mature adipocytes at 6h and 24h, the rising rate were 28.17% and 18.25% compared with the untreated cells. In contrast, secretion of leptin from 3T3-L1-mock mature adipocytes only increased by 14.57% at 24h. Conclusion: MCHR2 mediates the positive control of MCH on preadipocyte differentiation, triglyceride synthesis and leptin secretion without the influence on preadipocyte proliferation and lipolysis. The present study tentatively confirms that the physiological function of MCHR2 involvs in the regulation of energy balance.
1. MCHR2真核表达载体的构建与鉴定。方法:以人胎脑cDNA文库为模板PCR扩增获取MCHR2基因全长cDNA编码区序列,定向插入到真核表达载体pcDNA3.1(+)中,并转化大肠杆菌,经抗生素平板筛选获得阳性克隆,再分别进行酶切、PCR及DNA测序鉴定。结果:获得了长为1023bp的人MCHR2 cDNA全长的目的片段,并插入到pcDNA3.1(+)载体的CMV启动子下游构建了表达MCHR2的重组质粒,酶切和PCR鉴定证实重组质粒带有目的基因片段,DNA测序确认插入片段的序列准确无误。结论:表达人MCHR2蛋白的真核表达载体pcDNA3.1-MCHR2构建成功。
2. MCHR2在CHO细胞中的稳定表达及其生物学活性的分析。方法:脂质体转染法将pcDNA3.1-MCHR2真核表达载体导入CHO细胞,经G418筛选抗性细胞克隆,建立稳定转染细胞系,RT-PCR、Western blot检测MCHR2转录水平和蛋白水平的表达,免疫荧光检测MCHR2在细胞中的定位,放射性配体结合实验检测受体的密度和亲和力,通过测定细胞内cAMP、Ca~(2+)浓度的改变分析MCHR2的信号转导通路。结果:经脂质体转染和G418抗性筛选6~8周,建立了稳定转染pcDNA3.1-MCHR2质粒的CHO细胞系;在该细胞系中MCHR2基因能被有效地转录和翻译,并准确定位于细胞膜上,与125I-MCH的结合具有特异、高亲和及可饱和的配体-受体结合特性,其最大结合容量Bmax值为309.97±1.14fmol/mg protein,平衡解离常数Kd值为0.170±0.0006 nmol/L;MCH作用于MCHR2后不影响细胞内cAMP生成,可促使细胞内钙库释放Ca~(2+)进而使胞内Ca~(2+)浓度出现明显而短暂的升高,Ca~(2+)振荡过程在MCH刺激后30~50s内完成,其对Ca~(2+)释放的作用呈剂量依赖关系,半数有效浓度EC50为2.32±0.01nmol/L,提示MCHR2的信号转导通路是与Gq蛋白偶联。结论:建立了稳定转染MCHR2的CHO细胞系,从MCHR2的表达、定位、受体活性及信号转导通路等方面证实了真核表达载体pcDNA3.1-MCHR2稳定转染哺乳动物细胞后能有效表达具有生物学活性的MCHR2蛋白,为后续体外研究MCHR2基因的功能提供了可靠的实验依据。
3. MCHR2对脂肪细胞生物学特性的影响。方法:通过脂质体转染和G418筛选建立稳定转染pcDNA3.1-MCHR2质粒的3T3-L1前脂肪细胞系,命名为3T3-L1-MCHR2细胞,作为对照的稳定转染空质粒的细胞命名为3T3-L1-mock细胞;RT-PCR、Western blot、免疫荧光检测、放射性配体结合实验及细胞内Ca~(2+)检测鉴定MCHR2的表达及活性;MTT法检测细胞增殖,FCM检测细胞周期分布;激素鸡尾酒法(MIX+DEX+insulin)诱导前脂肪细胞分化,油红O染色观察脂肪细胞分化的形态学改变及量化分化程度,RT-PCR检测脂肪细胞分化成熟的标记基因;检测成熟脂肪细胞中TG的生成、FFA及Leptin的分泌。结果:建立了稳定转染细胞系——3T3-L1-MCHR2,并通过了MCHR2表达和活性的鉴定;对前脂肪细胞增殖和分化特性作比较分析,可见MCH对3T3-L1-MCHR2和3T3-L1-mock细胞的增殖无影响,对两株细胞的分化则有一定的促进作用,两者相比,3T3-L1-MCHR2细胞的分化速度更快、程度更高,具体表现为油红O比色在分化第9天开始出现脂质聚集增多(3T3-L1-mock细胞为第12天),分化成熟标记基因PPARγ2、C/EBPα和aP2从分化第3天起上调,Leptin自分化第6天起上调,而3T3-L1-mock细胞的PPARγ2和Leptin于分化第6天、C/EBPα和aP2于分化第9天才上调,且增幅较弱;对成熟脂肪细胞的脂质代谢特性的分析显示,3T3-L1-MCHR2和3T3-L1-mock细胞在MCH作用下FFA的含量无明显差异,而TG的含量明显升高,升高率分别为22.61%和12.17%;ELISA法检测成熟脂肪细胞Leptin的分泌,3T3-L1-MCHR2细胞在MCH作用6h及24h后Leptin分泌明显增多,增幅分别为28.17%和18.25%,而3T3-L1-mock细胞则在24h后才增多,增幅为14.57%。结论:MCHR2可介导MCH对前脂肪细胞分化、脂肪合成代谢及Leptin分泌的正调控作用,对前脂肪细胞增殖和脂肪分解代谢则无影响,初步证实MCHR2的生理功能参与了能量平衡的调控。
Obesity, a chronic disease characterized by excessive accumulation of body fat, has become a global health problem and is a significant risk factor for developing many serious diseases, including cardio- and cerebro- vascular diseases, diabetes, hypertension, dyslipidemias and some cancers. In recent years, it has been well established that obesity is associated with the neuroendocrine control of the hypothalamus. The identification of the hypothalamic orexigenic neuropeptide melanin-concentrating hormone (MCH) and its two receptors (MCHR1 and MCHR2) provide new targets for treatment of obesity. Major advances have been made in identifying the relevance of MCH and MCHR1 to obesity and energy homeostasis by transgenic animal and gene knockout animal. However, the function of MCHR2 and its relevance to obesity is still indistinct. In this study, we first construct the eukaryotic expression vector of human MCHR2 by DNA recombination technique. Then, we stably transfect mammalian cell line with the vector and analyse the expression, localization and biologic activity of MCHR2. Finally, we investigate the physiological function of MCHR2 in 3T3-L1 preadipocyte to evaluate whether it is involved in mediating the effects of MCH on energy balance and its relevance to obesity. This study includes three parts:
1. Construction and identification of eukaryotic expression vector of human MCHR2. Methods: The full-length MCHR2 cDNA fragment was amplified by PCR from the human fetal brain cDNA library and was oriently inserted into eukaryotic expression vector pcDNA3.1(+). The positive clone was obtained after transformation and antibiotic screening and was identified by restriction endonuclease digestion, PCR, and DNA sequencing. Results: The 1023bp MCHR2 cDNA fragment was obtained and cloned into the downstream of CMV promoter of pcDNA3.1(+). The recombinant plasmid was identified by restriction endonuclease digestion and PCR. DNA sequencing confirmed that the sequence of the inserted element was correct. Conclusion: The eukaryotic expression vector pcDNA3.1-MCHR2 was successfully constructed.
2. Stably expression and biological activity of MCHR2 in CHO cells. Methods: CHO cells were transfected with the recombinant plasmids using lipofectamine. Stable cell line was established by G418 selection. Expression of MCHR2 in transcriptional level and protein level was detected by RT-PCR and western blot. Localization of MCHR2 in CHO cells was assayed by immunofluorence technique. Radioligand binding assays were performed to survey receptor density and affinity of MCHR2. Signal transduction pathways mediated by MCHR2 were analyzed by measurement of intracellular cAMP and calcium. Results: CHO cell line stably transfected with pcDNA3.1-MCHR2 plasmids was generated after liposome transfection and G418 selection. MCHR2 gene was efficiently transcribed and translated, and its protein exactly localized on the cytomembrane. MCHR2 could specific bind with ~(125)I-MCH with high-affinity and saturability (Bmax=309.97±1.14fmol/mg protein, Kd=0.170±0.0006nmol/L). MCH stimulation had no effect on the production of cAMP. MCH induced a clear and transient increase of intracellular calcium, and this procedure was completed within 30~50s. The effect of MCH on intracellular calcium showed dose-dependent response with an effective half-maximal concentration (EC50) of 2.32±0.01nmol/L. It is suggested that MCHR2 is only coupled to Gq protein. Conclusion: Stably transfected CHO cell line was established successfully. It is confirmed that MCHR2 expressed in mammalian cells is a functional receptor for the MCH by the analysis of its expression, localization, receptor characteristics, and signal pathways. This present study provides a reliable experimental foundation for further studies on the function of MCHR2 in vitro.
3. Influence of MCHR2 on the biological characteristics of adipocyte. Methods: 3T3-L1 cell line stably transfected with pcDNA3.1-MCHR2 plasminds was generated by transfection and G418 selection and named as 3T3-L1-MCHR2 while the control cell line transfected with pcDNA3.1(+) was named as 3T3-L1-mock. The expression and biological activity of MCHR2 were identified by RT-PCR, western blot, immunofluorence, radioligand binding and intracellular calcium assay. Cell proliferation was determined by MTT assay. Cell cycle was detected by flow cytometry. Cell differentiation was induced by hormone cocktail (MIX+DEX+insulin) and examined by Oil red O staining. The marker genes of adipocyte differentiation were examined by RT-PCR. The production of triglyceride and secretions of FFA and leptin were measured. Results: Stably transfected cell line named 3T3-L1-MCHR2 was established and passed the identification of expression and biological activity of MCHR2. MCH had no effect on the proliferation of 3T3-L1-MCHR2 and 3T3-L1-mock cells. Both the two cell lines were induced to differentiate to mature adipocytes by MCH. The speed and degree of differentiation in 3T3-L1-MCHR2 cells were higher than those in 3T3-L1-mock cells. Following was the detailed display of differentiation in 3T3-L1-MCHR2 cells: accumulation of lipid increased at day 9 (in 3T3-L1-mock it increased at day 12); marker genes of mature adipocyte including PPARγ2、C/EBPαand aP2 increased at day 3, leptin increased at day 6, while in 3T3-L1-mock cells, up-regulation of PPARγ2 and leptin appeared at day 6, up-regulation of C/EBPαand aP2 appeared at day 9, and the increasing levels were lower. There was no difference of the FFA secretion between MCH-treated group and control group. The triglyceride content in 3T3-L1-MCHR2 and 3T3-L1-mock mature adipocytes obviously increased by MCH, and the rising rate were 22.61% and 12.17%, compared with corresponding untreated cells. MCH stimulated leptin releases from 3T3-L1-MCHR2 mature adipocytes at 6h and 24h, the rising rate were 28.17% and 18.25% compared with the untreated cells. In contrast, secretion of leptin from 3T3-L1-mock mature adipocytes only increased by 14.57% at 24h. Conclusion: MCHR2 mediates the positive control of MCH on preadipocyte differentiation, triglyceride synthesis and leptin secretion without the influence on preadipocyte proliferation and lipolysis. The present study tentatively confirms that the physiological function of MCHR2 involvs in the regulation of energy balance.
引文
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[6] Ito M, Gomori A, Ishihara A, et al. Characterization of MCH-mediated obesity in mice[J]. Am J Physiol Endocrinol Metab, 2003, 284(5): E940-E945.
[7] Segal-Lieberman G, Bradley RL, Kokkotou E, et al. Melanin-concentrating hormone is a critical mediator of the leptin deficient phenotype[J]. Proc Natl Acad Sci USA, 2003, 100(17): 10085-10090.
[8] Elliott JC, Harrold JA, Brodin P, et al. Increases in melanin-concentrating hormone and MCH receptor levels in the hypothalamus of dietary-obese rats[J]. Brain Res Mol Brain Res, 2004, 128(2): 150-159.
[9] Ludwig DS, Tritos NA, Mastaitis JW, et al. Melanin-concentrating hormone overexpression in transgenic mice leads to obesity and insulin resistance[J]. J Clin Invest, 2001, 107(3): 379-386.
[10] Kokkotou E, Jeon JY, Wang X, et al. Mice with MCH ablation resist diet-induced obesity through strain-specific mechanisms[J]. Am J Physiol Regul Integr Comp Physiol, 2005, 289(1): R117-R124.
[11] Hill J, Duckworth M, Murdock P, et al. Molecular cloning and functional characterization of hMCH2, a novel human MCH receptor[J]. J Biol Chem, 2001, 276(23): 20125-20129.
[12] Tan CP, Sano H, Iwaasa H, et al. Melanin-concentrating hormone receptor subtypes 1 and 2: species-specific gene expression[J]. Genomics, 2002, 79(6): 785-792.
[13] 杨俊霞, 易发平, 宋方洲等. 豚鼠 MCHR2 基因外显子 1 的克隆及序列分析[J].第三军医大学学报, 2006, 28(21):2167-2170.
[14] Chen Y, Hu CK, Hsu CK, et al. Targeted disruption of the melanin-concentrating hormone receptor-1 results in hyperphagia and resistance to diet-induced obesity[J]. Endocrinology, 2003, 143(7): 2469-2477.
[15] Astrand A, Bohlooly YM, Larsdotter S, et al. Mice lacking melanin-concentratinghormone receptor 1 demonstrate increased heart rate associated with altered autonomic activity[J]. Am J Physiol Regul Integr Comp Physiol, 2004, 287(4): R749- R758.
[16] Meyre D, Lecoeur C, Delplanque J, et al. A genome-wide scan for childhood obesity-associated traits in French families shows significant linkage on chromosome 6q22.31-q23.2[J]. Diabetes, 2004, 53(3): 803-811.
[1] Eberle AN, Mild G, Schlumberger S, et al. Expression and characterization of melanin-concentrating hormone receptors on mammalian cell lines[J]. Peptides, 2004, 25(10): 1585-1595.
[2] Schlumberger SE, Talke-Messerer C, Zumsteg U, et al. Expression of receptors for melanin-concentrating hormone (MCH) in different tissues and cell lines[J]. J Recept Signal Transduct Res, 2002, 22(1-4): 509-531.
[3] J.萨姆布鲁克, D.W.那塞尔著, 黄培堂等译校. 分子克隆实验指南(第三版)[M].北京: 科学出版社, 1996:627-628.
[1] Hawes BE, Kil E, Green B, et al. The melanin-concentration hormone receptor couples to multiple G proteins to activate diverse intracellular signaling pathways[J]. Endocrinology, 2000, 141(12): 4524-4532.
[2] Macdonald D, Murgolo N, Zhang R, et al. Molecular characterization of the melanin-concentration hormone/ receptor complex: identification of critical residues involved in binding and activation[J]. Mol Pharmacol, 2000, 58(1): 217-225.
[3] Chambers J, Ames RS, Bergsma D, et al. Melanin-concentrating hormone is the cognate ligand for the orphan G-protein-coupled receptor SLC-1[J]. Nature, 1999, 400(6741): 261-265.
[4] Tetsuka M, Satio Y, Imai K, et al. The basic residues in the membrane-proximal C-terminal tail of the rat melanin-concentrating hormone receptor 1 are required for receptor function[J]. Endocrinology, 2004, 145(8): 3712-3723.
[5] Murdoch H, Feng GJ, Bachner D, et al. Periplakin interferes with G protein activation by the melanin-concentrating hormone receptor-1 by binding to the proximal segment of the receptor C-terminal tail[J]. J Biol Chem, 2005, 280(9):8208-8220.
[6] Francke F,Ward RJ,Jenkins L, et al. Interaction of neurochondrin with the melanin concentrating hormone-receptor 1 interferes with G protein-coupled signal trans- duction but not agonist-mediated internalization[J]. J Biol Chem, 2006, 281(43): 32496-32507.
[7] Liu S, Carrillo JJ, Pediani J, et al. Effective information transfer from the α1b-adrenoceptor to Gα11 requires both β/γ interactions and an aromatic group four amino acids from the C terminus of the G protein[J]. J Biol Chem, 2002, 277(28): 25707-25714.
[8] Satio Y, Tetsuka M, Li Y, et al. Properties of rat melanin-concentrating hormone receptor 1 internalization[J]. Peptides, 2004, 25(10): 1597-1604.
[9] Abbott CR, Kennedy AR, Wren AM, et al. Identification of hypothalamic nuclei involved in the orexigenic effect of melanin concentrating hormone[J]. Endocrinology, 2003, 144(9): 3943-3949.
[10] Saito Y, Nothacker HP, Wang Z, et al. Molecular characterization of the melanin- concentrating-hormone receptor[J]. Nature, 1999, 400(6741): 265-269.
[11] Bachner D, Kreienkamp HJ, Weise C, et al. Identification of melanin concentrating hormone (MCH) as the natural ligand for the orphan somatostatin-like receptor 1 (SLC-1)[J]. FEBS Lett, 1999, 457(3): 522-524.
[12] Lembo PM, Grazzini E, Cao J, et al. The receptor for the orexigenic peptide melanin-concentrating hormone is a G-protein-coupled receptor[J]. Nat Cell Biol, 1999, 1(5): 267-271.
[13] Shimomura Y, Mori M, Sugo T, et al. Isolation and identification of melanin- concentrating hormone as the endogenous ligand of the SLC-1 receptor[J]. Biochem Biophys Res Comm, 1999, 261(3): 622-626.
[14] Mori M, Harada M, Terao Y, et al. Cloning of a novel G protein-coupled receptor, slt, a subtype of the melanin-concentrating hormone receptor[J]. Biochem Biophys Res Comm, 2001, 283(5): 1013-1018.
[15] Hill J, Duckworth M, Murdock P, et al. Molecular cloning and functionalcharacterization of hMCH2, a novel human MCH receptor[J]. J Biol Chem, 2001, 276(23): 20125-20129.
[16] Sailer AW, Sano H, Zeng Z, et al. Identification and characterization of a second melanin-concentrating hormone receptor, MCHR-2[J]. Proc Natl Acad Sci USA, 2001, 98(13): 7564-7569.
[17] An S, Cutler G, Zhao JJ, et al. Identification and characterization of a melanin- concentrating hormone receptor[J]. Proc Natl Acad Sci USA, 2001, 98(13): 7576 -7581.
[18] Wang S, Behan J, O’Neill K, et al. Identification and pharmacological characteri- zation of a novel human melanin-concentrating hormone receptor, mch-r2[J]. J Biol Chem, 2001, 276(37): 34664-34670.
[19] Rodriguez M, Beauverger P, Naime I, et al. Cloning and molecular characterization of the novel human melanin-concentrating hormone receptor (MCH2)[J]. Mol Pharmacol, 2001, 60(4): 632-639.
[20] Pissios P, Trombly DJ, Tzameli I, et al. Melanin-concentrating hormone receptor 1 activates extracellular signal-regulated kinase and synergizes with G(s)-coupled pathways[J]. Endocrinology, 2003, 144(8): 3514-3523.
[21] Eberle AN, Mild G, Schlumberger S, et al. Expression and characterization of melanin-concentrating hormone receptors on mammalian cell lines[J]. Peptides, 2004, 25(10): 1585-1595.
[1] Kim S, Moustaid-Moussa N. Secretory, endocrine and autocrine/paracrine function of the adipocyte[J]. J Nutr, 2000, 130(12): 3110S-3115S.
[2] Ahima, RS, Flier, JS. Adipose tissue as an endocrine organ[J]. Trends Endocrinol Metab, 2000, 11(8): 327-332.
[3] Serradeil-Le, Gal C, Lafontan M, et al. Characterization of NPY receptors controlling lipolysis and leptin secretion in human adipocytes[J]. FEBS Lett, 2000, 475(2): 150-156.
[4] Martinez JA, Aguado M, Fruhbeck G. Interactions between leptin and NPY affecting lipid mobilization in adipose tissue[J]. J Physiol Biochem, 2000, 56(1): 1-8.
[5] Norman D, Isidori AM, FrajeseV, et al. ACTH and α-MSH inhibit leptin expression and secretion in 3T3-L1 adipocytes: model for a central-/peripheral melanocortin- leptin pathway[J]. Molecular and Cellular Endocrinology, 2003, 200(1-2): 99-109.
[6] Bonston BA. The role of melanocortins in adipocyte function[J]. Ann NY Acad Sci, 1999, 885: 75-84.
[7] Bradley RL, Kokkotou EG, Maratos-Flier E, et al. Melanin-concentrating hormone regulates leptin synthesis and secretion in rat adipocytes[J]. Diabetes, 2000, 49(7): 1073-1077.
[8] Tadayyon M, Welters HJ, Haynes AC, et al. Expression of melanin-concentrating hormone receptors in insulin-producing cells: MCH stimulates insulin release in RINm5F and CRI-G1 cell-lines[J]. Biochem Biophys Res Commun, 2000, 275(2): 709-712.
[9] Bradley RL, Mansfield JP, Maratos-Flier E, et al. Melanin-concentrating hormone activates signaling pathways in 3T3–L1 adipocytes[J]. Am J Physiol Endocrinol Metab, 2002, 283(3): E584-592.
[10] Hill J, Duckworth M, Murdock P, et al. Molecular cloning and functional characterization of hMCH2, a novel human MCH receptor[J]. J Biol Chem, 2001, 276(23): 20125-20129.
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[14] Cowherd RM, Lyle RE and Mcgeheejr RE. Molecular regulation of adipocyte differentiation[J]. Cell Dev Bio, 1999,10(1): 3-10.
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[16] Gregoire FM, Smas CM and Sul HS. Understanding adipocyte differentiation[J]. Physiol Rev, 1998, 78(3): 783-809.
[17] Bradley RL, Mansfield JP, Maratos-Flier E. Neuropeptides, including neuropeptide Y and melanocortins, mediate lipolysis in murine adipocytes[J]. Obes Res, 2005, 13(4): 653-661.
[18] Ahima RS, Prabakaran D, Mantzoros C, et al. Role of leptin in the neuroendocrine response to fasting[J]. Nature, 1996, 18(6588): 250-252.
[19] Segal-Lieberman G, Bradley RL, Kokkotou E, et al. Melanin-concentrating hormone is a critical mediator of the leptin-deficient phenotype[J]. Proc Natl Acad Sci USA, 2003, 100(17): 10085-10090.
[20] Kokkotou EG, Tritos NA, Mastaitis JW, et al. Melanin-concentrating hormone receptor is a target of leptin action in the mouse brain[J]. Endocrinology, 2001, 142(2): 680-686.
[21] Gavrila A, Chan JL, Miller LC, et al. Circulating melanin-concentrating hormone, agout-related protein, and α-melanocytes-stimulating hormone levels in relation to body composition: alterations in response to food deprivation and recombinant human leptin administration[J]. J Clin Endocrinol Metab, 2005, 90(2): 1047-1054.
[1] WHO. World Health Organization 1998. Obesity: Preventing and managing the global epidemic. Report of a WHO consultation on obesity. Geneva, Switzerland.
[2] J.萨姆布鲁克,D.W.那塞尔著,黄培堂等译校.分子克隆实验指南(第三版)[M].北京:科学出版社,1996:627-628.
[3] Qu D, Ludwig DS, Gammeltoft S, et al. A role for melanin-concentrating hormone in the central regulation of feeding behaviour [J]. Nature, 1996, 380(6571): 243-247.
[4] Elliott JC, Harrold JA, Brodin P, et al. Increases in melanin-concentrating hormone and MCH receptor levels in the hypothalamus of dietary-obese rats [J]. Brain Res Mol Brain Res, 2004, 128(2): 150-159.
[5] Shimada M, Tritos NA, Lowell BB, et al. Mice lacking melanin-concentrating hormone are hypophagic and lean [J]. Nature, 1998, 396(6712): 670-674.
[6] Ludwig DS, Tritos NA, Mastaitis JW, et al. Melanin-concentrating hormone overexpression in transgenic mice leads to obesity and insulin resistance [J]. J ClinInvest, 2001, 107(3): 379-386.
[7] Katsuura G and Inui A. Melanin-Concentrating Hormone as a Metabolic and Cognitive Regulatory Factor [J]. Curr Med Chem–Central Nervous System Agents, 2003, 3(3): 217-227.
[8] Marsh DJ, Weingarth DT, Novi DE, et al. Melanin-concentrating hormone 1 receptor-deficient mice are lean, hyperactive, and hyperphagic and have altered metabolism [J]. Proc Natl Acad Sci USA, 2002, 99(5): 3240-3245.
[9] Chen Y, Hu CK, Hsu CK, et al. Targeted disruption of the melanin-concentrating hormone receptor-1 results in hyperphagia and resistance to diet-induced obesity [J]. Endocrinology, 2002, 143(7): 2469-2477.
[10] Sailer AW, Sano H, Zeng Z, et al. Identification and characterization of a second melanin-concentrating hormone receptor, MCH-2R [J]. Proc Natl Acad Sci USA, 2001, 98(13): 7564-7569.
[1] Shi Y. Beyond skin color: emerging roles of melanin-concentrating hormone in energy homeostasis and other physiological functions. Peptides, 2004, 25:1605-1611.
[2] Katsuura G,Inui A. Melanin-concentrating hormone as a metabolic and cognitive regulatory factor. Curr Med Chem, 2003, 3:217-227.
[3] Tan CP, Sano H, Iwaasa H, et al. Melanin-concentrating hormone receptor subtypes 1 and 2: species-specific gene expression. Genomics, 2002, 79:785-792.
[4] Hill J, Duckworth M, Murdock P, et al. Molecular cloning and functional characterization of hMCH2, a novel human MCH receptor. J Biol Chem, 2001,276:20125-20129.
[5] Eberle AN, Mild G, Schlumberger S, et al. Expression and characterization of melanin-concentrating hormone receptors on mammalian cell lines. Peptides, 2004, 25:1585-1595.
[6] Pissios P, Trombly DJ, Tzameli I, et al. Melanin-concentrating hormone receptor 1 activates extracellular signal-regulated kinase and synergizes with G(s)-coupled pathways. Endocrinology, 2003, 144:3514-3523.
[7] Della-Zuana O, Presse F, Ortola C, et al. Acute and chronic administration of melanin-concentrating hormone enhances food intake and body weight in Wistar and Sprague-Dawley rats. Int J Obes Relat Metab Disord, 2002, 26:1289-1295.
[8] Gomori A, Ishihara A, Ito M, et al. Chronic intracerebroventricular infusion of MCH causes obesity in mice. Am J Physiol Endocrinol Metab, 2003, 284: E583-E588.
[9] Ito M, Gomori A, Ishihara A, et al. Characterization of MCH-mediated obesity in mice. Am J Physiol Endocrinol Metab, 2003, 284: E940-E945.
[10] Segal-Lieberman G, Bradley RL, Kokkotou E, et al. Melanin-concentrating hormone is a critical mediator of the leptin deficient phenotype. Proc Natl Acad Sci USA, 2003, 100:10085-10090.
[11] Elliott JC, Harrold JA, Brodin P, et al. Increases in melanin-concentrating hormone and MCH receptor levels in the hypothalamus of dietary-obese rats. Brain Res Mol Brain Res, 2004, 128:150-159.
[12] Kokkotou E, Jeon JY, Wang X, et al. Mice with MCH ablation resist diet-induced obesity through strain-specific mechanisms. Am J Physiol Regul Integr Comp Physiol, 2005, 289:R117-R124.
[13] Chen Y, Hu CK, Hsu CK, et al. Targeted disruption of the melanin-concentrating hormone receptor-1 results in hyperphagia and resistance to diet-induced obesity. Endocrinology, 2003, 143:2469-2477.
[14] Astrand A, Bohlooly YM, Larsdotter S, et al. Mice lacking melanin-concentrating hormone receptor 1 demonstrate increased heart rate associated with altered autonomic activity. Am J Physiol Regul Integr Comp Physiol, 2004, 287:R749-R758.
[15] Meyre D, Lecoeur C, Delplanque J, et al. A genome-wide scan for childhood obesity-associated traits in French families shows significant linkage on chromosome 6q22.31-q23.2. Diabetes, 2004, 53:803-811.
[16] Gavrila A, Chan JL, Miller LC, et al. Circulating melanin-concentrating hormone, agout-related protein, and α-melanocytes-stimulating hormone levels in relation to body composition: alterations in response to food deprivation and recombinant human leptin administration. J Clin Endocrinol Metab, 2005, 90:1047-1054.
[17] Burdyga G, Varro A, Dimaline R, et al. Feeding-dependent depression of melanin-concentrating hormone and melanin-concentrating hormone receptor-1 expression in vagal afferent neurones. Neuroscience, 2006, 137:1405-1415.
[18] Abbott CR, Kennedy AR, Wren AM, et al. Identification of hypothalamic nuclei involved in the orexigenic effect of melanin-concentrating hormone. Endocrinology, 2003, 144:3943-3949.
[19] Challis BG, Coll AP, Yeo GS, et al. Mice lacking pro-opiomelanocortin are sensitive to high-fat feeding but respond normally to the acute anorectic effects of peptide-YY3-36. Proc Natl Acad Sci USA, 2004, 101:4695-4700.
[20] Su J, McKittrick BA, Tang H, et al. Discovery of melanin-concentrating hormone receptor R1 antagonists using high-throughput synthesis. Bioorg Med Chem, 2005, 13:1829-1836.
[2] Shi Y. Beyond skin color: emerging roles of melanin-concentrating hormone in energy homeostasis and other physiological functions[J]. Peptides, 2004, 25(10): 1605-1611.
[3] Katsuura G, Inui A. Melanin-concentrating hormone as a metabolic and cognitive regulatory factor[J]. Curr Med Chem, 2003, 3(3):217-227.
[4] Della-Zuana O, Presse F, Ortola C, et al. Acute and chronic administration of melanin-concentrating hormone enhances food intake and body weight in Wistar and Sprague-Dawley rats[J]. Int J Obes Relat Metab Disord, 2002, 26(10): 1289-1295.
[5] Gomori A, Ishihara A, Ito M, et al. Chronic intracerebroventricular infusion of MCH causes obesity in mice[J]. Am J Physiol Endocrinol Metab, 2003, 284(3): E583-E588.
[6] Ito M, Gomori A, Ishihara A, et al. Characterization of MCH-mediated obesity in mice[J]. Am J Physiol Endocrinol Metab, 2003, 284(5): E940-E945.
[7] Segal-Lieberman G, Bradley RL, Kokkotou E, et al. Melanin-concentrating hormone is a critical mediator of the leptin deficient phenotype[J]. Proc Natl Acad Sci USA, 2003, 100(17): 10085-10090.
[8] Elliott JC, Harrold JA, Brodin P, et al. Increases in melanin-concentrating hormone and MCH receptor levels in the hypothalamus of dietary-obese rats[J]. Brain Res Mol Brain Res, 2004, 128(2): 150-159.
[9] Ludwig DS, Tritos NA, Mastaitis JW, et al. Melanin-concentrating hormone overexpression in transgenic mice leads to obesity and insulin resistance[J]. J Clin Invest, 2001, 107(3): 379-386.
[10] Kokkotou E, Jeon JY, Wang X, et al. Mice with MCH ablation resist diet-induced obesity through strain-specific mechanisms[J]. Am J Physiol Regul Integr Comp Physiol, 2005, 289(1): R117-R124.
[11] Hill J, Duckworth M, Murdock P, et al. Molecular cloning and functional characterization of hMCH2, a novel human MCH receptor[J]. J Biol Chem, 2001, 276(23): 20125-20129.
[12] Tan CP, Sano H, Iwaasa H, et al. Melanin-concentrating hormone receptor subtypes 1 and 2: species-specific gene expression[J]. Genomics, 2002, 79(6): 785-792.
[13] 杨俊霞, 易发平, 宋方洲等. 豚鼠 MCHR2 基因外显子 1 的克隆及序列分析[J].第三军医大学学报, 2006, 28(21):2167-2170.
[14] Chen Y, Hu CK, Hsu CK, et al. Targeted disruption of the melanin-concentrating hormone receptor-1 results in hyperphagia and resistance to diet-induced obesity[J]. Endocrinology, 2003, 143(7): 2469-2477.
[15] Astrand A, Bohlooly YM, Larsdotter S, et al. Mice lacking melanin-concentratinghormone receptor 1 demonstrate increased heart rate associated with altered autonomic activity[J]. Am J Physiol Regul Integr Comp Physiol, 2004, 287(4): R749- R758.
[16] Meyre D, Lecoeur C, Delplanque J, et al. A genome-wide scan for childhood obesity-associated traits in French families shows significant linkage on chromosome 6q22.31-q23.2[J]. Diabetes, 2004, 53(3): 803-811.
[1] Eberle AN, Mild G, Schlumberger S, et al. Expression and characterization of melanin-concentrating hormone receptors on mammalian cell lines[J]. Peptides, 2004, 25(10): 1585-1595.
[2] Schlumberger SE, Talke-Messerer C, Zumsteg U, et al. Expression of receptors for melanin-concentrating hormone (MCH) in different tissues and cell lines[J]. J Recept Signal Transduct Res, 2002, 22(1-4): 509-531.
[3] J.萨姆布鲁克, D.W.那塞尔著, 黄培堂等译校. 分子克隆实验指南(第三版)[M].北京: 科学出版社, 1996:627-628.
[1] Hawes BE, Kil E, Green B, et al. The melanin-concentration hormone receptor couples to multiple G proteins to activate diverse intracellular signaling pathways[J]. Endocrinology, 2000, 141(12): 4524-4532.
[2] Macdonald D, Murgolo N, Zhang R, et al. Molecular characterization of the melanin-concentration hormone/ receptor complex: identification of critical residues involved in binding and activation[J]. Mol Pharmacol, 2000, 58(1): 217-225.
[3] Chambers J, Ames RS, Bergsma D, et al. Melanin-concentrating hormone is the cognate ligand for the orphan G-protein-coupled receptor SLC-1[J]. Nature, 1999, 400(6741): 261-265.
[4] Tetsuka M, Satio Y, Imai K, et al. The basic residues in the membrane-proximal C-terminal tail of the rat melanin-concentrating hormone receptor 1 are required for receptor function[J]. Endocrinology, 2004, 145(8): 3712-3723.
[5] Murdoch H, Feng GJ, Bachner D, et al. Periplakin interferes with G protein activation by the melanin-concentrating hormone receptor-1 by binding to the proximal segment of the receptor C-terminal tail[J]. J Biol Chem, 2005, 280(9):8208-8220.
[6] Francke F,Ward RJ,Jenkins L, et al. Interaction of neurochondrin with the melanin concentrating hormone-receptor 1 interferes with G protein-coupled signal trans- duction but not agonist-mediated internalization[J]. J Biol Chem, 2006, 281(43): 32496-32507.
[7] Liu S, Carrillo JJ, Pediani J, et al. Effective information transfer from the α1b-adrenoceptor to Gα11 requires both β/γ interactions and an aromatic group four amino acids from the C terminus of the G protein[J]. J Biol Chem, 2002, 277(28): 25707-25714.
[8] Satio Y, Tetsuka M, Li Y, et al. Properties of rat melanin-concentrating hormone receptor 1 internalization[J]. Peptides, 2004, 25(10): 1597-1604.
[9] Abbott CR, Kennedy AR, Wren AM, et al. Identification of hypothalamic nuclei involved in the orexigenic effect of melanin concentrating hormone[J]. Endocrinology, 2003, 144(9): 3943-3949.
[10] Saito Y, Nothacker HP, Wang Z, et al. Molecular characterization of the melanin- concentrating-hormone receptor[J]. Nature, 1999, 400(6741): 265-269.
[11] Bachner D, Kreienkamp HJ, Weise C, et al. Identification of melanin concentrating hormone (MCH) as the natural ligand for the orphan somatostatin-like receptor 1 (SLC-1)[J]. FEBS Lett, 1999, 457(3): 522-524.
[12] Lembo PM, Grazzini E, Cao J, et al. The receptor for the orexigenic peptide melanin-concentrating hormone is a G-protein-coupled receptor[J]. Nat Cell Biol, 1999, 1(5): 267-271.
[13] Shimomura Y, Mori M, Sugo T, et al. Isolation and identification of melanin- concentrating hormone as the endogenous ligand of the SLC-1 receptor[J]. Biochem Biophys Res Comm, 1999, 261(3): 622-626.
[14] Mori M, Harada M, Terao Y, et al. Cloning of a novel G protein-coupled receptor, slt, a subtype of the melanin-concentrating hormone receptor[J]. Biochem Biophys Res Comm, 2001, 283(5): 1013-1018.
[15] Hill J, Duckworth M, Murdock P, et al. Molecular cloning and functionalcharacterization of hMCH2, a novel human MCH receptor[J]. J Biol Chem, 2001, 276(23): 20125-20129.
[16] Sailer AW, Sano H, Zeng Z, et al. Identification and characterization of a second melanin-concentrating hormone receptor, MCHR-2[J]. Proc Natl Acad Sci USA, 2001, 98(13): 7564-7569.
[17] An S, Cutler G, Zhao JJ, et al. Identification and characterization of a melanin- concentrating hormone receptor[J]. Proc Natl Acad Sci USA, 2001, 98(13): 7576 -7581.
[18] Wang S, Behan J, O’Neill K, et al. Identification and pharmacological characteri- zation of a novel human melanin-concentrating hormone receptor, mch-r2[J]. J Biol Chem, 2001, 276(37): 34664-34670.
[19] Rodriguez M, Beauverger P, Naime I, et al. Cloning and molecular characterization of the novel human melanin-concentrating hormone receptor (MCH2)[J]. Mol Pharmacol, 2001, 60(4): 632-639.
[20] Pissios P, Trombly DJ, Tzameli I, et al. Melanin-concentrating hormone receptor 1 activates extracellular signal-regulated kinase and synergizes with G(s)-coupled pathways[J]. Endocrinology, 2003, 144(8): 3514-3523.
[21] Eberle AN, Mild G, Schlumberger S, et al. Expression and characterization of melanin-concentrating hormone receptors on mammalian cell lines[J]. Peptides, 2004, 25(10): 1585-1595.
[1] Kim S, Moustaid-Moussa N. Secretory, endocrine and autocrine/paracrine function of the adipocyte[J]. J Nutr, 2000, 130(12): 3110S-3115S.
[2] Ahima, RS, Flier, JS. Adipose tissue as an endocrine organ[J]. Trends Endocrinol Metab, 2000, 11(8): 327-332.
[3] Serradeil-Le, Gal C, Lafontan M, et al. Characterization of NPY receptors controlling lipolysis and leptin secretion in human adipocytes[J]. FEBS Lett, 2000, 475(2): 150-156.
[4] Martinez JA, Aguado M, Fruhbeck G. Interactions between leptin and NPY affecting lipid mobilization in adipose tissue[J]. J Physiol Biochem, 2000, 56(1): 1-8.
[5] Norman D, Isidori AM, FrajeseV, et al. ACTH and α-MSH inhibit leptin expression and secretion in 3T3-L1 adipocytes: model for a central-/peripheral melanocortin- leptin pathway[J]. Molecular and Cellular Endocrinology, 2003, 200(1-2): 99-109.
[6] Bonston BA. The role of melanocortins in adipocyte function[J]. Ann NY Acad Sci, 1999, 885: 75-84.
[7] Bradley RL, Kokkotou EG, Maratos-Flier E, et al. Melanin-concentrating hormone regulates leptin synthesis and secretion in rat adipocytes[J]. Diabetes, 2000, 49(7): 1073-1077.
[8] Tadayyon M, Welters HJ, Haynes AC, et al. Expression of melanin-concentrating hormone receptors in insulin-producing cells: MCH stimulates insulin release in RINm5F and CRI-G1 cell-lines[J]. Biochem Biophys Res Commun, 2000, 275(2): 709-712.
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