Ibuprofen regulates the expression and function of membrane-associated serine proteases prostasin and matriptase.
Ibuprofen regulates the expression and function of membrane-associated serine proteases prostasin and matriptase.
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DOI：
10.1186/s12885-015-2039-6
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发表时间：
2015-12-29
期刊：
影响因子：
3.8
通讯作者：
Chen LM
中科院分区：
文献类型：
--
作者：
Chai AC;Robinson AL;Chai KX;Chen LM
糖基磷脂酰肌醇锚定的细胞外膜丝氨酸蛋白酶前列腺素在正常膀胱尿路上皮细胞中表达。膀胱炎症减少前列腺素表达，前列腺素表达的丧失与人膀胱移行细胞癌中的上皮-间质转化（EMT）相关。非甾体类抗炎药（NSAID）可降低包括膀胱癌在内的各种癌症的发病率，但NSAID抗癌作用的分子机制尚未完全了解。使用正常人膀胱尿路上皮细胞系UROtsa、正常人滋养层细胞系B6 Tert-1、人膀胱移行细胞癌细胞系UM-UC-5和UM-UC-9以及人乳腺癌细胞系JIMT-1进行研究。通过逆转录/定量聚合酶链反应（RT-qPCR）和免疫印迹分析布洛芬处理后这些细胞中丝氨酸蛋白酶前列腺素和基质蛋白酶以及环氧合酶（考克斯-1和考克斯-2）的表达变化。通过测量B6 Tert-1细胞中的跨上皮电阻（TEER）和上皮通透性，评估了布洛芬调节的前列腺素在上皮紧密连接形成和维持中的功能作用。还在过表达重组人前列腺素、前列腺素表达沉默或用功能性阻断前列腺素抗体处理的B6 Tert-1细胞中评价了前列腺素对紧密连接的作用。在过表达前列腺素的细胞中检查间质蛋白酶酶原活化。前列腺素在UROtsa和B6Tert-1细胞中的表达增加。布洛芬以剂量依赖性方式上调UROtsa细胞中环氧合酶-2（考克斯-2）在mRNA和蛋白水平的表达，但不是上调前列腺素表达的必要条件。布洛芬诱导的前列腺素有助于B6 Tert-1细胞上皮紧密连接的形成和维持。在UROtsa细胞中，间质蛋白酶酶原被布洛芬下调，这可能是前列腺素表达增加的结果，因为过表达前列腺素导致间质蛋白酶活化和酶原在UROtsa、JIMT-1和B6 Tert-1细胞中下调。布洛芬对膀胱癌细胞中前列腺素和间质蛋白酶表达的差异调节。已经建议将Iceland用于治疗膀胱癌。我们的研究结果将上皮细胞外膜丝氨酸蛋白酶前列腺素和间质蛋白酶纳入NSAID抗癌作用的潜在分子机制。
The glycosylphosphatidylinositol-anchored extracellular membrane serine protease prostasin is expressed in normal bladder urothelial cells. Bladder inflammation reduces prostasin expression and a loss of prostasin expression is associated with epithelial-mesenchymal transition (EMT) in human bladder transitional cell carcinomas. Non-steroidal anti-inflammatory drugs (NSAIDs) decrease the incidence of various cancers including bladder cancer, but the molecular mechanisms underlying the anticancer effect of NSAIDs are not fully understood. The normal human bladder urothelial cell line UROtsa, the normal human trophoblast cell line B6Tert-1, human bladder transitional cell carcinoma cell lines UM-UC-5 and UM-UC-9, and the human breast cancer cell line JIMT-1 were used for the study. Expression changes of the serine proteases prostasin and matriptase, and cyclooxygenases (COX-1 and COX-2) in these cells following ibuprofen treatments were analyzed by means of reverse-transcription/quantitative polymerase chain reaction (RT-qPCR) and immunoblotting. The functional role of the ibuprofen-regulated prostasin in epithelial tight junction formation and maintenance was assessed by measuring the transepithelial electrical resistance (TEER) and epithelial permeability in the B6Tert-1 cells. Prostasin’s effects on tight junctions were also evaluated in B6Tert-1 cells over-expressing a recombinant human prostasin, silenced for prostasin expression, or treated with a functionally-blocking prostasin antibody. Matriptase zymogen activation was examined in cells over-expressing prostasin. Ibuprofen increased prostasin expression in the UROtsa and the B6Tert-1 cells. Cyclooxygenase-2 (COX-2) expression was up-regulated at both the mRNA and the protein levels in the UROtsa cells by ibuprofen in a dose-dependent manner, but was not a requisite for up-regulating prostasin expression. The ibuprofen-induced prostasin contributed to the formation and maintenance of the epithelial tight junctions in the B6Tert-1 cells. The matriptase zymogen was down-regulated in the UROtsa cells by ibuprofen possibly as a result of the increased prostasin expression because over-expressing prostasin leads to matriptase activation and zymogen down-regulation in the UROtsa, JIMT-1, and B6Tert-1 cells. The expression of prostasin and matriptase was differentially regulated by ibuprofen in the bladder cancer cells. Ibuprofen has been suggested for use in treating bladder cancer. Our results bring the epithelial extracellular membrane serine proteases prostasin and matriptase into the potential molecular mechanisms of the anticancer effect of NSAIDs.