ROLE OF MACROPHAGE RECEPTORS FOR OXLDL IN ATHEROGENESIS

拟议研究的长期目标是改善我们的 了解脂蛋白-巨噬细胞相互作用， 有助于泡沫细胞形成和动脉粥样硬化形成。 重点将 将其置于乙酰LDL受体（SRA）、hCD 36及其小鼠上， 同源物（mDC 36）和94-97 kDa氧化LDL（OxLDL）- 小鼠腹腔巨噬细胞的结合蛋白， 以前被表征，现在被鉴定为macrosialin。 Macrosialin及其人类同源物DC 68，之前都是 克隆，主要存在于晚期内体部分， 其功能尚不清楚。 我们建议研究它们的运输 在亚细胞区室之间，以制备稳定转染的细胞 为了表征它们的配体结合特性，鉴定配体 通过制备嵌合蛋白和通过位点- 定向诱变，并制备麦胡萝卜素敲除小鼠， 确定巨噬细胞的表型结果 功能、免疫功能和动脉粥样硬化形成。 CD 36是一种 OxLDL结合受体，在结合和吞噬中起作用 与玻连蛋白受体合作， 血小板反应蛋白 我们将检验一个假设， 氧化低密度脂蛋白可以通过这种相互作用来调节。 使用细胞 来自SRA敲除小鼠（T.儿玉），我们将 评估受体在体外和体内OxLDL代谢中的作用， vivo. 最后，我们将继续使用表达克隆， 非洲爪蟾卵母细胞寻找额外的巨噬细胞蛋白， 可能参与摄取OxLDL或/和受损或 凋亡细胞已经分离出五个克隆体。 没有一 它们代表SRA、mCD 36或巨唾液酸蛋白。 这些将 进行测序和鉴定。

The long term goal of the proposed studies is to improve our understanding of lipoprotein-macrophage interactions that may contribute to foam cell formation and atherogenesis. Emphasis will be placed on the acetyl LDL receptor (SRA), hCD36 and its mouse homologue (mDC36), and a 94-97 kDa oxidized LDL (OxLDL)- binding protein from mouse peritoneal macrophages which we have previously characterized and have now identified as macrosialin. Macrosialin and its human homologue, DC68, both previously clones, are found predominantly in the late endosomal fraction and their functions are not known. We propose to study their transport between subcellular compartment, to prepare stably transfected cells to characterize their ligand-binding properties, identify ligand binding sites by preparation of chimeric proteins and by site- directed mutagenesis, and prepare a mcarosialin knockout mouse to ascertain phenotypic consequences with regard to macrophage function, immune function and atherogenesis. CD36, an established OxLDL-binding receptor, functions in the binding and phagocytosis of apoptotic cells in cooperation with the vitronectin receptor and thrombospondin. We will test the hypothesis that binding of OxLDL may be modulated by interactions of this kind. Using cells from the SRA knockout mouse (gift of Dr. T. Kodama), we will assess the role of the receptor in OxLDL metabolism in vitro and in vivo. Finally, we will continue to use expression cloning in Xenopus oocytes to search for additional macrophage proteins that may participate in uptake of OxLDL or/and of damaged or apoptotic cells. Five clones have already been isolated. None of them represents either SRA, mCD36, or macrosialin. These will be sequenced and characterized.