title: Regulation of integrin-mediated adhesion by muscarinic acetylcholine receptors and protein kinase C in small cell lung carcinoma.
abstract: OBJECTIVE
  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.
  
  
  METHODS
  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.
  
  
  METHODS
  Experiments were conducted in the Molecular Pharmacology Laboratory, Guthrie Research Institute.
  
  
  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.
authors.affiliation: Department of Cardiothoracic Surgery, Guthrie Clinic, Sayre, PA, USA.
authors.forename: R L
authors.initials: RL
authors.lastname: Quigley
authors.forename: S H
authors.initials: SH
authors.lastname: Shafer
authors.forename: C L
authors.initials: CL
authors.lastname: Williams
mesh_terms: D001285:Atropine / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D018288:Carcinoma, Small Cell / Q000378:metabolism / Q000503:physiopathology*; D002448:Cell Adhesion / Q000502:physiology; D016326:Extracellular Matrix Proteins; D005455:Fluorescent Antibody Technique; D066298:In Vitro Techniques; D016023:Integrins / Q000378:metabolism / Q000502:physiology*; D008175:Lung Neoplasms / Q000378:metabolism / Q000503:physiopathology*; D018721:Muscarinic Agonists / Q000494:pharmacology; D018727:Muscarinic Antagonists / Q000494:pharmacology; D011493:Protein Kinase C / Q000502:physiology*; D011976:Receptors, Muscarinic / Q000502:physiology*; D012710:Serum Albumin, Bovine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D016326:Extracellular Matrix Proteins; D016023:Integrins; D018721:Muscarinic Agonists; D018727:Muscarinic Antagonists; D011976:Receptors, Muscarinic; D012710:Serum Albumin, Bovine; D001285:Atropine; D002217:Carbachol; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate