PHARMACOLOGY OF ANTIRHEUMATIC AGENTS

描述：（改编自申请人的摘要）- 研究者 首先提出了腺苷作用于特定受体的假设 对炎症细胞，介导抗炎作用 甲氨蝶呤，最常用和有效的二线抗风湿药 可用代理。 在该项目的前两年，他们取得了 四个最初目标中的三个。 他证明了腺苷的形成 细胞外的核苷酸去磷酸化，证实 腺苷介导甲氨蝶呤的抗炎作用 关节炎的体内模型，并描述了一种新的信号转导 中性粒细胞腺苷 A2A 受体。 他还展示了之前的 腺苷作用于其受体，在发病机制中发挥着意想不到的作用 甲氨蝶呤毒性（结节病），令人惊讶的是，炎症 细胞因子（IL-1 和 TNFα）下调抗炎 (A2A) 腺苷 受体 mRNA。 他现在建议研究： I. 生化机制 其中甲氨蝶呤促进腺苷释放。 他将决定是否 甲氨蝶呤促进细胞内腺嘌呤核苷酸的积累 (HPLC)，增加腺嘌呤核苷酸从细胞中的转运（HPLC， 腺嘌呤核苷酸库的放射性标记），或减少对 细胞的腺苷（放射性标记的腺苷摄取）。 二. 互动 腺苷受体和炎症之间：调节和 反监管。 他将研究甲氨蝶呤减少 A2A、A3 或两者均缺乏的小鼠的急性和慢性炎症 受体（基因敲除小鼠），以更好地确定这些受体中的哪一个是 负责腺苷的抗炎作用。 他还将 确定 IL-1 和 TNFα 是否调节 mRNA 稳定性（核径流 腺苷的测定）或转录（启动子构建体的转染） A2A 受体 mRNA。 三． 腺苷介导的甲氨蝶呤机制 毒性。 甲氨蝶呤治疗最可怕的并发症是肝脏并发症 纤维化。 他的数据表明，腺苷作用于其受体 成纤维细胞，促进基质生成并抑制基质分解。 他 将确定甲氨蝶呤是否促进腺苷释放 肝细胞以及这些细胞释放的腺苷是否影响 通过培养的星状细胞（肝细胞）生产胶原蛋白或胶原酶 成纤维细胞）。 拟议研究的结果将提出新颖的建议 开发治疗炎症药物的方法 关节炎并可能导致新治疗方法的开发 和预防甲氨蝶呤毒性。

DESCRIPTION: (Adapted from the applicant's abstract) - The investigator first proposed the hypothesis that adenosine, acting at specific receptors on inflammatory cells, mediates the antiinflammatory effects of methotrexate, the most commonly used and effective second line antirheumatic agent available. During the first 2 years of this project they achieved three of the four original aims. He demonstrated that adenosine is formed extracellularly from the dephosphorylation of nucleotides, confirmed that adenosine mediates the antiinflammatory effects of methotrexate in an in vivo model of arthritis, and described a novel signal transduction at neutrophil adenosine A2A receptors. He also demonstrated a previously unsuspected role for adenosine, acting at its receptors, in the pathogenesis of methotrexate toxicity (nodulosis), and that, surprisingly, inflammatory cytokines (IL-1 and TNFalpha) downregulate antiinflammatory (A2A) adenosine receptor mRNA. He now proposes to examine: I. The biochemical mechanism by which methotrexate promotes adenosine release. He will determine whether methotrexate promotes intracellular accumulation of adenine nucleotides (HPLC), increases transport of adenine nucleotides out of cells (HPLC, radiolabelling of adenine nucleotide pools), or diminishes uptake of adenosine by cells (radiolabelled adenosine uptake). II. Interaction between adenosine receptors and inflammation: Regulation and counterregulation. He will study the capacity of methotrexate to diminish acute and chronic inflammation in mice deficient in A2A, A3, or both receptors (knockout mice) to determine better which of these receptors is responsible for the antiinflammatory actions of adenosine. He will also determine whether IL-1 and TNFalpha regulate mRNA stability (nuclear run-off assays) or transcription (transfection of promoter constructs) of adenosine A2A receptor mRNA. III. Adenosine-mediated mechanisms of methotrexate toxicity. The most feared complication of methotrexate therapy is hepatic fibrosis. His data indicate that adenosine, acting at its receptors on fibroblasts, promotes matrix generation and inhibits matrix breakdown. He will determine whether methotrexate promotes adenosine release from hepatocytes and whether the adenosine released from these cells affects collagen or collagenase production by cultured stellate cells (hepatic fibroblasts). The results of the proposed studies will suggest novel approaches to the development of drugs for the treatment of inflammatory arthritis and may lead to the development of new approaches to the treatment and prevention of methotrexate toxicity.