title: Human beta-defensin-2 controls cell cycle in malignant epithelial cells: in vitro study.
abstract: OBJECTIVE
  In the present research we analyze the mechanism of human beta-defensin-2 (hBD-2) influence on cultured malignant epithelial cell growth.
  
  
  METHODS
  The analysis of a concentration-dependent effect of recombinant hBD-2 (rec-hBD-2) on cell growth patterns and cell cycle distribution has been performed in vitro with 2 cell lines (human lung adenocarcinoma A549 cells and human epidermoid carcinoma A431 cells) using MTT test, flow cytometry and direct cell counting. To study intracellular localization of hBD-2 immunocytofluorescent and immunocytochemical analyses were applied, and effect of hBD-2 on signal cascades involved in cell cycle regulation has been studied by Western blotting.
  
  
  RESULTS
  According to our data, rec-hBD-2 exerts a concentration-dependent effect on the viability of cultured A549 and A431 cells. It causes proproliferative effect at concentrations below 1 nM, significant suppression of cell proliferation at concentration range from 10 nM to 1 μM (p<0.05), and cell death at higher concentrations. Using flow cytometry we have demonstrated that hBD-2 dependent growth suppression is realized via cell cycle arrest at G1/S phase (p<0.05). Also, we have registered significant activation of pRB and decreased expression of Cyclin D1 in cells treated with the defensin compared to untreated control cells, while the expression of p53 remains unaffected. The study of intracellular localization of hBD-2 in these cells has revealed that exogeneously added defensin molecules enter the cells, are distributed throughout the cytoplasm and could be detected in cell nuclei. The model study using A549 cells treated with 1,25-(OH)(2)D(3) has shown similar cell growth suppression effect of native endogenously produced hBD-2.
  
  
  CONCLUSIONS
  The results of our study suggest that in malignant epithelial cells hBD-2 may control cell growth via arrest of G1/S transition and activation of pRB.
authors.affiliation: R.E. Kavetsky Institute of Experimental Pathology, Oncology and Radiobiology, NAS of Ukraine, Kyiv,Ukraine.
authors.forename: E
authors.initials: E
authors.lastname: Zhuravel
authors.forename: T
authors.initials: T
authors.lastname: Shestakova
authors.forename: O
authors.initials: O
authors.lastname: Efanova
authors.forename: Yu
authors.initials: Y
authors.lastname: Yusefovich
authors.forename: D
authors.initials: D
authors.lastname: Lytvin
authors.forename: M
authors.initials: M
authors.lastname: Soldatkina
authors.forename: P
authors.initials: P
authors.lastname: Pogrebnoy
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D002453:Cell Cycle; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019938:Cyclin D1 / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology; D016193:G1 Phase / Q000187:drug effects; D006801:Humans; D009369:Neoplasms; D016160:Retinoblastoma Protein / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D023083:beta-Defensins / Q000378:metabolism* / Q000494:pharmacology
chemical_list: C106641:DEFB4A protein, human; D016160:Retinoblastoma Protein; D016159:Tumor Suppressor Protein p53; D023083:beta-Defensins; D019938:Cyclin D1