SERUM AMYLOID A GENE EXPRESSION: A KINETIC ANALYSIS

急性期反应是一个复杂的生理过程，包括 系统性、新陈代谢、体液和营养改变，这被设置为 组织损伤和细胞坏死后的运动。其中一个这样的事件是 大量血浆浓度的变化，通常是增加 蛋白质，包括两种诱导蛋白，血清淀粉样蛋白A(SAA)和 C反应蛋白(CRP)。对SAA合成的诱导部分是 其特征是一系列细胞和分子事件，包括 巨噬细胞产生循环介质白介素1的研究 在肝脏中SAA mRNA转录和SAA合成之前。这个 这项研究的总体目标是定义细胞和分子 调节SAA基因表达的过程。 这项研究将从两个体内诱导SAA的研究开始 动物模型，小鼠和兔子，使用两种炎症刺激， 皮下注射脂多糖和松节油。这个 体内研究的目标将是确定药物的顺序动力学。 IL-1的产生、SAA mRNA的转录、SAA mRNA的翻译以及 翻译后将前SAA修饰为成熟apoSAA。除了……之外 确定了涉及肝脏SAA诱导的事件的分子链， 这项建议将研究可能的肝外SAA合成。此外， IL-1介导的SAA诱导与急性期的关系 对C反应蛋白的诱导合成将进行研究。使用分子细节 测定对体内SAA诱导、IL-1等因素的影响 调节SAA合成将在细胞水平上进行评估 肝脏和其他合成SAA组织的培养，目的是 定义了模拟1000的活体情况所必需的条件 SAA合成成倍增加。在分子水平上，调节和 比较SAA基因无细胞翻译和细胞翻译的动力学 用兔网织红细胞裂解液系统、非洲爪哇卵母细胞和原代 肝细胞培养。最后，在基因组水平上，数量和 人类和小鼠SAA基因的分布将被确定，DNA 正常人和继发性淀粉样变性者SAA基因的多态性 将会被比较。将采用的方法包括放射免疫分析， Northern和Southern DNA杂交分析、免疫细胞化学、电子 显微镜、原代细胞培养和游离蛋白质合成。这 这项研究将为未来的治疗改进提供必要的信息 在组织损伤和细胞坏死后的宿主防御，并将支持 未来对慢性炎症的研究，其中一个序列是 严重的淀粉样变性。

The acute phase response is a complex physiologic process consisting of systemic, metabolic, humoral and nutritional alterations, that is set into motion following tissue injury and cell necrosis. One such event is the change, usually an increase, in concentration of a large number of plasma proteins, including two inducible proteins, serum amyloid A (SAA) and C-reactive protein (CRP). Induction of SAA synthesis has been partially characterized as a sequence of cellular and molecular events involving macrophage production of a circulating mediator Interleukin 1 (IL-1) which precedes transcription of SAA mRNA and synthesis of SAA in liver. The overall goal of this study is to define the cellular and molecular processes regulating SAA gene expression. The investigation will begin with studies of in vivo SAA induction in two animal models, mice and rabbits, using two inflammatory stimuli, lipopolysaccharide (LPS) and subcutaneous turpentine oil injection. The goal of the in vivo studies will be to determine the sequential kinetics of IL-1 production, of transcription of SAA mRNA, translation of SAA mRNA, and post-translational modification of preSAA to mature apoSAA. In addition to defining the molecular chain of events involved in hepatic SAA induction, this proposal will study possible extrahepatic SAA synthesis. Furthermore, the relationship between IL-1 mediated SAA induction and acute phase induction of CRP synthesis will be studied. Using the molecular details determined for in vivo SAA induction, the effect of IL-1 and other factors regulating SAA synthesis will be evaluated at the cellular level in primary cultures of liver and other SAA synthesizing tissues, with the aim of defining those conditions necessary to mimic the in vivo situation of 1000 fold increase in SAA synthesis. At the molecular level, the regulation and kinetics of cell-free and cellular translation of SAA mRNA will be compared using the rabbit reticulocyte lysate system, Xenopus oocytes and primary liver cell cultures. Finally, at the genomic level, the number and distribution of human and mouse SAA genes will be determined and the DNA polymorphism of SAA in normal individuals and those with secondary amyloid will be compared. Methodology to be employed includes radioimmunoassay, Northern and Southern DNA blot analysis, immunocytochemistry, electron microscopy, primary cell culture and cell free protein synthesis. This study will provide information essential to future therapeutic enhancement of host defense following tissue injury and cell necrosis and will support future studies of chronic inflammation, one sequence of which is the devastating condition amyloidosis.