SERUM AMYLOID A GENE EXPRESSION: A KINETIC ANALYSIS

急性期反应是一个复杂的生理过程

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.