Serine and cysteine proteinase inhibitors prevent nitric oxide production by activated macrophages by interfering with transcription of the inducible NO synthase gene.
Serine and cysteine proteinase inhibitors prevent nitric oxide production by activated macrophages by interfering with transcription of the inducible NO synthase gene.
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丝氨酸和半胱氨酸蛋白酶抑制剂通过干扰诱导型 NO 合酶基因的转录来防止活化的巨噬细胞产生一氧化氮。
DOI：
10.1006/bbrc.1995.2523
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Ignarro,LJ
中科院分区：
文献类型：
--
作者：
Griscavage,JM;Wilk,S;Ignarro,LJ
本研究的目的是确定丝氨酸和半胱氨酸蛋白酶抑制剂干扰LPS激活的大鼠肺泡巨噬细胞产生NO的机制。将巨噬细胞在存在LPS +测试试剂的情况下孵育24小时。分析培养基中的NO− X积累，测定收获的细胞的iNOS活性，并提取细胞RNA以通过北方印迹分析测定iNOS mRNA。TPCK、TLCK、calpain inhibitor 1（CPI-1）和calpain inhibitor 2（CPI-2）在1-100 μM浓度范围内均以浓度依赖性方式抑制NO− X生成和诱导型iNOS表达。TPCK和CPI-1的效力分别比TLCK和CPI-2高约10倍。这些数据表明，胰凝乳蛋白酶样丝氨酸或半胱氨酸蛋白酶是LPS诱导的iNOS基因表达所必需的，可能是通过涉及转录因子NF-κB激活的机制。因此，测试了NF-κB活化的有效抑制剂，其作用归因于抑制多催化蛋白酶复合物（MPC）的胰凝乳蛋白酶样活性。Z-IE（O-t-Bu）A-Leucinal在1 μM时可抑制NO− X的产生和诱导型iNOS的表达，在10 nM时抑制率超过50%。这些观察结果表明，MPC的抑制剂干扰iNOS的诱导，并提供了有力的证据，MPC的功能，重要的iNOS诱导巨噬细胞。
The objective of this study was to ascertain the mechanism by which serine and cysteine proteinase inhibitors interfere with production of NO by LPS-activated rat alveolar macrophages. Macrophages were incubated in the presence of LPS + test agent for 24 hr. Culture media were analyzed for NO−Xaccumulation, harvested cells were assayed for iNOS activity, and cellular RNA was extracted for determination of iNOS mRNA by Northern blot analysis. TPCK, TLCK, calpain inhibitor 1 (CPI-1) and calpain inhibitor 2 (CPI-2) each inhibited NO−Xproduction and inducible iNOS expression in a concentration-dependent manner at 1-100 μM. TPCK and CPI-1 were about 10-fold more potent than TLCK and CPI-2, respectively. These data suggested that a chymotrypsin-like serine or cysteine proteinase is required for the LPS-inducible expression of the iNOS gene, perhaps by mechanisms involving activation of transcription factor NF-κB. Accordingly, a potent inhibitor of NF-κB activation whose action is attributed to inhibition of the chymotrypsin-like activity of the multicatalytic proteinase complex (MPC) was tested. Z-IE(O-t-Bu)A-Leucinal abolished NO−Xproduction and inducible iNOS expression at 1 μM and showed over 50% inhibition at 10 nM. These observations indicate that inhibitors of MPC interfere with iNOS induction and provide strong evidence that MPC functions importantly in iNOS induction in macrophages.