Antibodies against the melanocortin-4 receptor act as inverse agonists in vitro and in vivo
Antibodies against the melanocortin-4 receptor act as inverse agonists in vitro and in vivo
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DOI：
10.1152/ajpregu.00878.2006
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发表时间：
2007-06-01
影响因子：
2.8
通讯作者：
Hofbauer, Karl G.
中科院分区：
文献类型：
--
作者：
Peter, Jean-Christophe;Nicholson, Janet R.;Hofbauer, Karl G.
针对中枢G蛋白偶联受体的功能活性抗体（Abs）尚未报道。我们选择下丘脑黑皮质素-4受体（MC 4-R）作为靶点，因为它在能量稳态调节中起着至关重要的作用。将N-末端（NT）结构域的15个氨基酸序列用作抗原。该肽在表面等离子体共振实验和对过表达人MC 4-R（hMC 4-R）的HEK-293细胞的研究中显示出功能活性。对NT肽免疫的大鼠产生特异性抗体，其在体外纯化和表征。在HEK-293细胞中，大鼠抗NT Ab显示hMC 4-R的特异性免疫荧光标记。它们在基础条件下和用合成MC 4-R激动剂刺激后减少cAMP的产生。针对NT肽免疫的大鼠产生了与MC 4-R阻断一致的表型，即增加的食物摄入和体重、增加的肝脏和脂肪垫重量以及升高的血浆甘油三酯。在对大鼠的单独实验中，在将纯化的Ab注射到第三脑室中之后，可以产生食物摄入的增加。在注射兔抗NT抗体的大鼠中获得了类似的结果。我们的数据表明，第一次，主动免疫大鼠对NT序列的MC 4-R的结果在特定的抗体，这似乎刺激食物的摄入，作为下丘脑中的反向激动剂。
Functionally active antibodies (Abs) against central G-protein-coupled receptors have not yet been reported. We selected the hypothalamic melanocortin-4 receptor (MC4-R) as a target because of its crucial role in the regulation of energy homeostasis. A 15 amino acid sequence of the N-terminal (NT) domain was used as an antigen. This peptide showed functional activity in surface plasmon resonance experiments and in studies on HEK-293 cells overexpressing the human MC4-R (hMC4-R). Rats immunized against the NT peptide produced specific antibodies, which were purified and characterized in vitro. In HEK-293 cells, rat anti-NT Abs showed specific immunofluorescence labeling of hMC4-R. They reduced the production of cAMP under basal conditions and after stimulation with a synthetic MC4-R agonist. Rats immunized against the NT peptide developed a phenotype consistent with MC4-R blockade, that is, increased food intake and body weight, increased liver and fat pad weight, and elevated plasma triglycerides. In a separate experiment in rats, an increase in food intake could be produced after injection of purified Abs into the third ventricle. Similar results were obtained in rats injected with anti-NT Abs raised in rabbits. Our data show for the first time that active immunization of rats against the NT sequence of the MC4-R results in specific Abs, which appear to stimulate food intake by acting as inverse agonists in the hypothalamus.