| Multi-drug resistance (MDR) is considered to be a major cause of failure of chemotherapy for tumor. MDR of tumor can be observed in most cancer cases. Therefore, it becomes one of the hot issues of cancer researches.At present, the recognized mechanisms of MDR include: P-glycoprotein (P-gp), multidrug-resistance protein (MRP), breast cancer resistance protein(BCRP), lung resistance-related protein(LRP). Moreover,DNA topoisomerases ,Protein kinase C (PKC), MGMT and Glutathione transferase(GST) also play an important role. However, we cannot fully explain the phenomena of MDR by these mechanisms. The latest studies suggest some other mechanisms probably contribute to MDR of tumor. With the development of molecular biology and correlated techniques, scientists keep discovering new mechanisms and multidrug resistance-related genes. To study the expression of the new gene of MDR can help us comprehend some new mechanisms of tumor's multi-drug resistance.The aim of our research is to study the expression of the two new drug resistance-related gene cDNA fragments, which our group cloned from SPC-A-1/CDDP with the suppression subtractive hybridization(SSH). The two cDNA fragments are named as fragment A1 and fragment A2.Methods: (1) We collected the samples from patients without chemotherapy, which included 12 cases of lung adenocarcinoma, 8 cases of lung squamous carcinoma, 2 cases of small cell lung cancer, 16 cases of gastric adenocarcinoma, 4 cases of gastric signet-ring cell carcinoma, 4 cases of Hepatocarcinoma, 3 cases of renal carcinoma, and the normal tissues around. These samples were stored in liquid nitrogen. (2) The plasmid with the cDNA fragment was amplified and extracted with the method of alkaline denaturation. The two cDNA fragments were cut from these plasmids by restricting enzyme (EcoRI) and harvested by V electrophoresis chamber. The two cDNAfragments were respectively labeled by using digoxigenin and 32P .(3) We used the In Situ Hybridization Histochemistry(ISHH) with the application of labeled digoxigenin to study the expressing location of the two fragments in these cancer tissues and the normal tissues of the samples.(4) We adopted the method of Northern blot with the application of label 32P to study the relative expressing quantity of the two fragments in these cancer tissues and the normal tissues of the samples.Results:(1) By ISHH, the expression of fragment A1 was observed in 10 cases of lung adenocarcinoma tissues among 12 cases detected, and the expression of fragment A1 was observed in 10 cases of gastric adenocarcinoma tissues among 16 cases detected. Meanwhile, the expression of fragment A2 was observed in 6 cases of lung adenocarcinoma tissues among 12 cases detected, and the expression of fragment A2 was observed in 7 cases of gastric adenocarcinoma tissues among 16 cases detected. The expression of fragment A1 and fragment A2 was not observed in the other cancer tissues and normal tissues.(2) By Northern blot, the expression of fragment A1 was found in 8 cases of lung adenocarcinoma tissues among 12 cases detected, and the expression of fragment A1 was found in 10 cases of gastric adenocarcinoma tissues among 16 cases detected, but not in other lung and gastric adenocarcinoma tissues, normal tissues and other carcinoma tissues. There was significant difference between lung adenocarcinoma tissues and normal lung tissues(P<0.01),and there was significant difference between gastric adenocarcinoma tissues and normal gastric tissues(P<0.01). At the same time, the expression of fragment A2 was found in 6 cases of lung adenocarcinoma tissues among 12 cases detected, and the expression of fragment A2 was found in 7 cases of gastric adenocarcinoma tissues among 16 cases detected, but not in other lung and gastric adenocarcinoma tissues, normal tissues and other carcinoma tissues. There was significant difference between lung adenocarcinoma tissues and normal lung tissues(P<0.01),and there was significant difference between gastric adenocarcinoma tissu...
|
PDF Full Text Request