Selective expression of mitogen-inducible cyclooxygenase in macrophages stimulated with lipopolysaccharide.
Selective expression of mitogen-inducible cyclooxygenase in macrophages stimulated with lipopolysaccharide.
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DOI：
10.1016/s0021-9258(18)35698-9
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发表时间：
1992-12
期刊：
影响因子：
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通讯作者：
Soo-Hee Lee;E. Soyoola;P. Chanmugam;Suzanne B. Hart;Wenqin Sun;Hua Zhong;Shuenn S. Liou;D. Simmons;D. Hwang
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文献类型：
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作者：
Soo-Hee Lee;E. Soyoola;P. Chanmugam;Suzanne B. Hart;Wenqin Sun;Hua Zhong;Shuenn S. Liou;D. Simmons;D. Hwang
已知存在于真核生物中的两种形式的环加氧酶：首先从公羊精囊中纯化的由2.8-脱氢酶mRNA编码的环加氧酶（考克斯-1）和新发现的由4-脱氢酶mRNA编码的促分裂原诱导的环加氧酶（考克斯-2）。通过1）测定用阿司匹林灭活内源性酶后新合成的酶的活性; 2）比较用或不用脂多糖处理的细胞中新合成的酶蛋白的水平;和3）评估编码考克斯-1和考克斯-2的mRNA的表达，研究用脂多糖刺激的大鼠肺泡巨噬细胞中这两种形式的酶的表达。通过Western印迹分析和使用两种不同抗体的35 S标记的酶的免疫沉淀来评估酶蛋白的水平，一种抗体对考克斯-2具有特异性，另一种抗体识别两种形式的酶，但优先识别考克斯-1。我们在这里报告的增强环氧合酶活性诱导的细菌脂多糖在大鼠肺泡巨噬细胞是由选择性表达的考克斯-2。地塞米松完全抑制脂多糖刺激的巨噬细胞中考克斯-2的表达，而考克斯-1则不受影响。在静息的未受刺激的巨噬细胞中，仅检测到考克斯-1而未检测到考克斯-2。逆转录聚合酶链反应检测发现，巨噬细胞中考克斯-2的mRNA水平升高，而脂多糖对考克斯-1的mRNA水平无影响。这些结果表明，脂多糖刺激的巨噬细胞中，由于考克斯-2的选择性表达，导致了胰高血糖素和血栓素合成的增加。
Two forms of cyclooxygenase are known to be present in eukaryotic organisms: a cyclooxygenase (COX-1) first purified from ram seminal vesicles encoded by a 2.8-kilobase mRNA, and a newly discovered mitogen-inducible cyclooxygenase (COX-2) encoded by a 4-kilobase mRNA. Expression of these two forms of the enzyme in rat alveolar macrophages stimulated with lipopolysaccharide was investigated by 1) determining the activity of newly synthesized enzyme after inactivating the endogenous enzyme with aspirin; 2) comparing levels of newly synthesized enzyme proteins in cells treated with or without lipopolysaccharide; and 3) assessing the expression of the mRNAs encoding COX-1 and COX-2. Levels of enzyme proteins were assessed by Western blot analysis and immunoprecipitation of 35S-labeled enzyme using two different antibodies, one specific for COX-2 and the other recognizing both forms of the enzyme but preferentially recognizing COX-1. We report here that the enhanced cyclooxygenase activity induced by the bacterial lipopolysaccharide in rat alveolar macrophages is caused by selective expression of the COX-2. Expression of COX-2 in macrophages stimulated by lipopolysaccharide was completely inhibited by dexamethasone, whereas COX-1 was unaffected. In resting unstimulated macrophages, only COX-1 but not COX-2 was detected. Levels of mRNA for the COX-2 in macrophages were increased, but those of the COX-1 were not affected by lipopolysaccharide as assessed by reverse transcription coupled with polymerase chain reaction. These results indicate that increased synthesis of prostaglandins and thromboxanes in lipopolysaccharide-stimulated macrophages results from selective expression of COX-2.