The Correlation Between MAPK,PKC Signal Transduction Pathways And MDR In MGC803 Cells

Multidrug resistance (MDR) is a major factor in the failure of many forms of chemptherapy. Several different molecular mechanisms will switch on in MDR cells, the most investigated mechanisms with known clinical significance are: (1) activation of transmembrane proteins effluxing different chemical substance from the cells, in which P - glycoprotein( P - gp) encoding by MDR1 and multi-drug resistance associated protein (MRP) ; (2) activation of the enzymes of the glutathione detoxification system ( especially GST ; ( 3 ) alteration of the genes and the proteins involved into the control of apotosis ( especially p53 Bcl - 2) MDR associated genes are expressed in a large proportion of human tumors, and its expression in several different forms cancer was shown to be associated with a lack of response to combination chemotherapy. MDR1 expression is usually low or undetectable prior to treatment, but it is frequently increased during the progression of the disease and , most noticeably, after chemotherapy . The increased expression of MDR1 mRNA can be found in some drug - sensitive cancer cells by transient exposure to different chemotherapeutic drugs.The signal transduction pathway of the mitogen - activated protein kinanses (MAPKs) play a critical role in cell proliferation differatiation and apoptosis. The ERK1/2 ( Ras/Raf - 1/MEK1/2/ERK1/2) signal transduction pathway is a subfamily of MAPK. The expression of MDR1 and the activation of MAPK increase in cancer cells after treatment with various chemotherapeutic drugs. The selective inhibitor of MEK1/2, PD098058, has been shown to significant reverse the drug resistance of drug resistance cell line L1210/VCR, but its accuratemechanism is needed to elucidate that whether MAPK plays a role in MDR, and whether the regulation of MEK can regulate the expression of MDR.Recently accumulated evidence indicates that there is relationship between the function of MDR and protein kinase C(PKC). PKC comprises a family of at least 13 distinct serine/threonine kinase isoenzymes involved in signal transduc-tion pathways that govern a wide range of physiological processes including differentiation, proliferation, gene expression, pathogenesis and development of tumor, activation of oncogene , phosphorylation of protein and so on. Some studies show that the activity of PKC in MDR tumor cells with overexpression P - gp is significantly higher than that in drug - sensitive cells, but Schwartz, et al. have found that the activity of PKC in human acute lymphocyte leukemia drug - resistance cells MOLT - 3/TQ2500 is lower than that in drug - sensitive cells. The phosphorylation of P - gp can be enhanced and drug - resistance can be partially reversed by the activator of PKC in human leukemia and some solid tumors , but the drug - resistance can not be changed by the activator and inhibitor in human acute lymphocyte leukemia cells MOLT - 3. Besides, some reports show that the phosphorylation of P - gp by PKC can not modulate the fuction of P - gp pump, the drug - resistance revers by inhibiting PKC is correlation with inducing apoptosis in cells.Our study was to observe the expressions of associated genes of MDR of human gastric cancer cell line MGC803 by transient exposure to Vincristine( VCR) and the effect on MDR by the specific inhibitors of MEKl/2( PD098059) and PKC(myr-PKC).Materials and MethodsCell line and cell cultureHuman gastric cancer cell line MGC803 were grown as monolayer in RP-MI1640 medium supplemented with 10% fetal calf serum, gentamicin (12U/ ml) at 37 ina5% CO2 atmosphere.Morphological analysis of cellsAfter treatment with VCR (20ng/ml) or VCR (20ng/ml) + PD09805910nM/mlK VCR(20ng/ml) + myrPKC (50nM/ml) 24h,48h,MGC803 cells were analyzed by wright - Giemsa staining, and the morphology of cells were detected by optic microscope.MTT assay to drug sensibilityCells (1 X 105ml) pretreated with VCR(20ng/ml) for72h were plated into 96 - well plate and cultured in 100l RPMI1640 medium. After cultured 4h, cells were divided i...