ABSTRACT
<p><b>OBJECTIVE</b>To investigate the reversal effect of O-(4-ethoxyl-butyl)-berbamine (EBB) on multidrug resistance (MDR) in MCF-7/ADR cell.</p><p><b>METHODS</b>3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay was used to assess the antitumor effect of EBB and determine the reversal effects of different concentrations ( < or = IC20) of EBB on MCF-7/ADR cell. Flow cytometry was applied to observe the intracellular accumulation of Rh123 and cell cycle in the presence of EBB. The expressions of MDR-related genes mdr 1 and topoisomerase II b (top II b) were evaluated by reverse transcription-polymerase chain reaction.</p><p><b>RESULTS</b>The sensitivity of MCF-7/ADR to adriamycin (ADR) was enhanced up to 50. 40, 89.80, and 14.88 folds after exposure of the cells to 3 micromol/L EBB, 7.5 micromol/L EBB, and 10 micromol/L verapamil (VPL), respectively. After 2 hours of incubation with 6 micromol/L EBB, intracellular Rh123 accumulation in MCF-7/ADR cells was increased to the level comparable to that in MCF-7 cells. When 6 micromol/L EBB was added together with 2 micromol/L ADR, MCF-7/ ADR cells showed to be arrested in the G2/M phase. The declination of mdr 1 gene expression was observed when 6 micromol/L EBB, 12 micromol/L EBB, and 10 micromol/L VPL were added for 48 hours; meanwhile, the expression of top II b mRNA showed no significant change.</p><p><b>CONCLUSION</b>EBB has a strong reversal effect on MDR in MCF-7/ ADR cell, which may be achieved by enhancing the arrestment of MCF-7/ADR cells at G2/M phase and increasing intracellular drug concentration.</p>
Subject(s)
Female , Humans , ATP Binding Cassette Transporter, Subfamily B, Member 1 , Metabolism , Antineoplastic Agents , Pharmacokinetics , Pharmacology , Benzylisoquinolines , Pharmacology , Breast Neoplasms , Pathology , Calmodulin , Cell Cycle , Cell Line, Tumor , Doxorubicin , Pharmacokinetics , Pharmacology , Drug Interactions , Drug Resistance, Multiple , Drug Resistance, NeoplasmABSTRACT
<p><b>OBJECTIVE</b>To investigate the potential effect of EBB, a calmodulin antagonist, on invasion of human fibrosarcoma cells HT1080.</p><p><b>METHODS</b>The antitumor effect of EBB was assessed by 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Activities of matrix metalloproteinase (MMP)-2 and MMP-9 were measured by Zymogrophy analysis. The mRNA levels, of MMP-2, MMP-9, and tissue inhibitor of metalloproteinases (TIMP)-1 were evaluated by reverse transcriptionpolymerase chain reaction (RT-PCR). Transwell chamber assay was applied to measure the effect of EBB on the invasion of HT1080 cells.</p><p><b>RESULTS</b>Calmodulin antagonist EBB inhibited the proliferation of HT1080 cells with an IC50 of (8.2 +/- 1.2) microg/ml. EBB down-regulated the activities of MMP-2 and MMP-9, and down-regulated the mRNA levels of MMP-2 and MMP-9, while up-regulated the mRNA levels of TIMP-1. The invasive ability of HT1080 cells was decreased to (31.13 +/- 2.265)%, (59.91 +/- 2.566)%, and (71.58 +/- 0.5960)% after exposure of the cells with 2, 5, and 10 microg/ml EBB, respectively.</p><p><b>CONCLUSION</b>Treatment with calmodulin antagonist EBB is effective in suppressing tumor invasion. The possible mechanism is the down-regulation of MMPs.</p>
Subject(s)
Humans , Antineoplastic Agents , Pharmacology , Benzylisoquinolines , Pharmacology , Calmodulin , Cell Line, Tumor , Down-Regulation , Fibrosarcoma , Pathology , Matrix Metalloproteinase 2 , Genetics , Matrix Metalloproteinase 9 , Genetics , Neoplasm Invasiveness , RNA, Messenger , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To investigate whether fetal bone marrow stromal cells have hemangioblastic characteristics.</p><p><b>METHODS</b>Human fetal bone marrow stromal cells (hfMSCs) were isolated and cultured. Immunophenotypes of hfMSCs were tested by FACS. hfMSCs seeded in the matrigel were induced with vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in vitro. Vascularization and hematopoiesis were detected with immunohistochemistry and electron microscope.</p><p><b>RESULTS</b>The typical properties of this CD34- stromal cell population were that 99% cells expressed Flk1 (vascular endothelial cell growth factor receptor 2) and tube structure was formed. In the process of induction, hfMSCs could give rise to CD34+ round cells.</p><p><b>CONCLUSIONS</b>We have demonstrated that fetal bone marrow stroma-derived Flk1+ CD34- cells could differentiate into vascular endothelial cells and hematopoietic cells, indicating that fetal bone marrow stroma-derived Flk1+ CD34- cells have hemangioblastic characteristics.</p>
Subject(s)
Humans , Antigens, CD34 , Allergy and Immunology , Bone Marrow Cells , Cell Biology , Allergy and Immunology , Metabolism , Cell Differentiation , Cells, Cultured , Fetus , Fibroblast Growth Factor 2 , Metabolism , Hematopoiesis , Hematopoietic Stem Cells , Cell Biology , Allergy and Immunology , Metabolism , Immunophenotyping , Stromal Cells , Cell Biology , Allergy and Immunology , Metabolism , Vascular Endothelial Growth Factor Receptor-2 , PhysiologyABSTRACT
Inhibitory effects of the O - ( 1 - ethoxvl - butyl ) berbarmine (EBB) on cultured fibroblast was studied by observating calmodulin (CaM ) content of cultured fibroblast with ELISA and DNA content at each phase of cell cycle with flow - cvtometerv. The CaM con-tent in the test group . compared to control . decreased markedly and DNA content increased significantly in the G0+Gi phase but reduced in the S phase. These results suggested that inhibitory mechanism of EBB on fibroblast proliferation may be closely related to CaM decrease in cells.