| INTRODUCTIONLung neoplasm has done harm to human health for many years. As the other solid tumors, chemotherapy is still the main method for controlling the recurrent and metastasis. The therapy of solid tumor was not very perfect because of the multidrug resistance. The tumor cell lines resisted one kind of anticancer agents and would resist many other drugs that are different from the first one on structure and function. Phenotype of multidrug resistance are classified as two types. One is herent resistance, and the other is acquired resistance. The tumor cells are sensitive to the anticancer drugs firstly, but after several courses they become to resistant the drugs. At present, the related factors included P-gp, MRP, LRP, Topo - H , GST, some changes of biochemical character in tumors. MRP was detected by Cole in 1992, and then the interrelated problems were attended.Recently, the studies on MRP expression and renal tumor, hema-tologic disease, liver tumor were more. But the study in lung neoplasm just began in broad, and the results existed many differences. In the present study we detected the MRP expression in NSCLC by im-munohistochemistry and explored the correlation between MRP expression and clinicopathological parameters, prognosis. At the same time we investigated the difference of MRP expression between the normal lung tissues and NSCLC by Western Blot and immunohistochemistry.Our study provided the theory basis for the clinical therapy of lung cancer, and made the therapy more pertinent.MATERIALS AND METHODS1. MaterialsAll the 62 samples used in immunohistochemical study were obtained from the patients admitted to the 1 st Affiliated Hospital of China Medical University during 1989 - 1992. All the patients were followed up, and follow - up period was from the operation to death of patient. All the samples were fixed in 10% formalin and embedded in paraffin. All the 30 samples used in Western Blot were obtained from the patients admitted to the Liaoning Province Tumor Hospital and the 1st Affiliated Hospital of China Medical University.2. ReagentsThe mouse-anti-human monoclonal antibody of MRP ( MRPm6) was provided by professor R. J. Scheper of Department of Pathology in Free University. The Ultro - sensitive S - P kit and DAB agent kit were bought from Fujian Maxin Biological Company. The goat - anti -mouse second antibody was bought from Huamei Biological Company.3. Methods3. 1 Western BlotRapidly homogenize tissues (1 -2g) in 5 volumes of lysis buffer (50mM Triscl PH 7.4, 150mM Nacl, 0.1% SDS, 1% Triton -100, ImM EDTA PH8.0, Aprotinin 12ul/10ml, PMSF 60ul/10ml) , Centrifuge the homogenate (13000rpm, 4℃) for 30 minutes to pellet insoluble material. (1) Electrophoresis (2) transfer proteins from gel to membrane (3)blocking (4) incubation with primary antibody (5)enzymeconjugate incubation (6)substrate incubation. (7)scan the results to computer3.2 ImmunohistochemistryDetect the MRP expression by immunohistochemistry S - P method. Scoring of the results: 100 cancer cells were counted in 10 fields which were selected at random. We used a scoring system to evaluate semiquantitatively the immunoexpression, judging the cases with brown plasma/membrane staining as positive. The sum score of positive cell was classified into the following 4 groups: 0, less than 10% positive; 1, more than 10% but less than 40% positive; 2, more than 40% but less than 70% positive; 3 , more than 70% positive. The intensity score was categorized into the following 3 groups; 0, no staining; 1, mild staining; 2, marked staining. The product of sum score and intensity score was classified as following 3 grades: negative, product score was 0; mild positive, product score was 1 - 3; marked positive, product score was 4-6.4. Statistical analysisSPSS for Windows 10. 0 statistical software was used to solve the data and Log ?rank test to analyze the differences. Cox model was used to solve the multivariate analysis. Paired - samples T test was used to solve the da...
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