Regulation of integrin-mediated adhesion by muscarinic acetylcholine receptors and protein kinase C in small cell lung carcinoma.
Regulation of integrin-mediated adhesion by muscarinic acetylcholine receptors and protein kinase C in small cell lung carcinoma.
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DOI：
10.1378/chest.114.3.839
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发表时间：
1998-09
期刊：
影响因子：
9.6
通讯作者：
R. Quigley;S. Shafer;Carol L. Williams
中科院分区：
文献类型：
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作者：
R. Quigley;S. Shafer;Carol L. Williams
研究目的了解小细胞肺癌(SCLC)细胞的表型特征可能有助于这种早期转移的支气管源性恶性肿瘤的新疗法的发展。在此，我们研究了在SCLC细胞上表达的M3亚型M受体(MAChR)的激活是否影响这些细胞的β1整合素介导的黏附。研究了小细胞肺癌细胞系SCC-9和NCI-H345与细胞外基质(ECM)蛋白的粘附性。用甲苯胺蓝标记细胞并测量光密度，或用~3H-胸腺嘧啶核苷标记细胞并测量β-活性来量化细胞粘附性。采用荧光激活细胞分选技术定量检测小细胞肺癌细胞表面β1整合素的表达。背景实验在格思里研究所分子药理实验室进行。测量和结果用激动剂卡巴胆碱(10微米，1.5h)激活mAChR可显著增加SCC-9细胞对ECM蛋白、层粘连蛋白和I型和IV型胶原的黏附。相反，mAChR激活不改变SCC-9细胞与玻璃体连接蛋白、纤维连接蛋白、多L赖氨酸或牛血清白蛋白的黏附。卡巴胆碱也不会改变缺乏功能性mAChR的NCI-H345小细胞肺癌细胞的粘附性。用针对β1整合素的AIIB2封闭抗体预先与SCC-9细胞孵育可抑制mAChR诱导的与ECM蛋白的黏附。免疫荧光分析表明，mAChR激活并不改变SCC-9细胞表面β1整合素的表达。佛波酯(PMA)(10 nM，1.5h)直接刺激蛋白激酶C(PKC)可增加SCC-9和NCI-H345细胞对ECM蛋白的黏附。这些结果表明，直接激活PKC或刺激M3mAChR(导致PKC活性增加)会增加β1整合素的结合活性，从而增加SCLC细胞对ECM蛋白的黏附。结论mAChR能够调节小细胞肺癌的增殖和黏附，提示这些受体的激活可能参与改变小细胞肺癌的发生和转移。
STUDY OBJECTIVES Improved understanding of the phenotypic characteristics of small cell lung cancer (SCLC) cells may facilitate the development of new therapies for this bronchogenic malignancy with early metastases. Herein we investigate whether activation of the M3 subtype of muscarinic acetylcholine receptor (mAChR) expressed on SCLC cells affects beta1-integrin-mediated adhesion of these cells. DESIGN Adhesion of the SCLC cell lines SCC-9 and NCI-H345 to extracellular matrix (ECM) proteins was investigated. Cell adhesion was quantified by labeling the cells with either toluidine blue dye and measuring optical density or 3H-thymidine and measuring beta-activity. Fluorescence-activated cell sorting was used to quantify the SCLC cell surface expression of beta1-integrins. SETTING Experiments were conducted in the Molecular Pharmacology Laboratory, Guthrie Research Institute. MEASUREMENTS AND RESULTS Activation of mAChR with the agonist carbachol (10 microM, 1.5 h) significantly increases adhesion of the SCC-9 SCLC cell line to the ECM proteins laminin and collagen types I and IV. In contrast, mAChR activation does not alter the adhesion of SCC-9 cells to vitronectin, fibronectin, poly-L-lysine, or bovine serum albumin. Carbachol also does not alter the adhesion of NCI-H345 SCLC cells that lack functional mAChR. Preincubation of SCC-9 cells with the AIIB2 blocking antibody to beta1-integrin inhibits mAChR-induced adhesion to ECM proteins. Immunofluorescence analysis indicates that mAChR activation does not alter the surface expression of beta1-integrins by SCC-9 cells. Direct stimulation of protein kinase C (PKC) by treatment with phorbol 12-myristate 13-acetate (PMA) (10 nM, 1.5 h) increases the adhesion of both the SCC-9 and NCI-H345 cell lines to ECM proteins. These results indicate that direct activation of PKC or stimulation of M3 mAChR (which results in increased PKC activity) increases the binding activity of beta1-integrins, resulting in increased adhesion of SCLC cells to ECM proteins. CONCLUSIONS The ability of mAChR to regulate SCLC proliferation and adhesion suggests that activation of these receptors may be used to alter SCLC tumorigenesis and metastasis.