CORE--ANIMAL PHYSIOLOGY

蛋白质化学/动物生理学核心将提供极具成本效益的服务 该项目涉及蛋白质化学、分析生物化学和动物生理学等领域。这个核心的蛋白质化学部分将提供组织中通过己糖胺途径的通量的测量，并确定其表达可能受己糖胺途径调控的关键蛋白质的血浆水平。特别是，核心将对主要的己糖胺途径产物的细胞内水平进行高效液相色谱测量，这将表明该途径在不同代谢条件下在脂肪、肌肉和内皮组织中的激活程度(Rossetti，Barzilai，Hawkins，Brownlee项目)。作为对这一途径激活程度的另一种直接测量，核心将使用一种高度特异的免疫沉淀和Western blotting技术来量化关键细胞内蛋白质SP-1的糖基化数量。众所周知，这种转录因子的激活受其糖基化程度的影响，调节着许多与代谢综合征相关的蛋白质的转录， 包括多种细胞因子和急性期反应物。由于这种相互作用对所有项目都至关重要，核心将测量血浆中纤溶酶原激活物抑制物-1的水平 (PAI-1)、血管紧张素原、肿瘤坏死因子-α、白细胞介素1-β和6、Acrp30、抵抗素、血清淀粉样蛋白A和瘦素(Rossetti，Barzilai，Hawkins，ShaMoon)，以及主动脉提取液中PAI-1水平(Brownlee)，采用包括放射免疫分析、ELISA分析、酶免疫分析和Western blotting在内的各种技术。该核心的动物生理学部分将为动物准备进行留置动静脉插管以及内脏脂肪和主动脉的外科切除提供最好的服务(由Rossetti，Barzilai，Brownlee项目)。

The Protein Chemistry/Animal Physiology Core will provide highly cost-effective services to the program project in the areas of protein chemistry, analytical biochemistry, and animal physiology. The Protein Chemistry component of this core will provide measurements of the amount of flux through the hexosamine pathway in tissues, and determinations of plasma levels of key proteins whose expression is likely to be regulated by the hexosamine pathway. In particular, the core will perform HPLC measurements of intracellular levels of the major hexosamine pathway products, which will indicate the degree of activation of this pathway in fat, muscle and endothelial tissue under various metabolic conditions (projects by Rossetti, Barzilai, Hawkins, Brownlee). As an additional, direct measurement of the degree of activation of this pathway, the core will use a highly specific technique ofimmtmoprecipitation and Western blotting to quantify the amount of glycosylation of a key intracellular protein, sp-1. This transcription factor, whose activation is known to be affected by its degree of glycosylation, regulates the transcription of many proteins associated with the metabolic syndrome, including many cytokines and acute phase reactants. Since this interaction is of central importance to all projects, the Core will measure plasma levels ofplasminogen activator inhibitor-1 (PAI- 1), angiotensinogen, TNF-alpha, Interleukins 1-beta and 6, Acrp30, resistin, serum amyloid A and leptin (projects by Rossetti, Barzilai, Hawkins, Shamoon), and PAI-1 levels in aortic extracts (project by Brownlee), by a variety of techniques including radioimmunoassays, ELISA assays, enzyme immunoassays and Western blotting. The Animal Physiology component of this core will provide a superlative service for animal preparation in performing indwelling arterial and venous catheterizations and surgical resections of visceral fat and aortas (projects by Rossetti, Barzilai, Brownlee).