Experimental Study On The Effects Of Mad1 To Rat Bladder Tumor Model In Vivo

OBJECTIVE : The aim of this study was to construct the orthotopicmodel of bladder tumor in rats and transfect expression plasmidpcDNA3.1(+)/Mad1 into bearing tumors rats and to observe if overexpression of MAD1 can inhibit the proliferation of bladder tumor and toexplore the TERT-targeted bladder tumor therapeutic approaches. METHODS AND RESULTS : Bladder tumors were induced in SDrats in situ and expression plasmid pcDNA3.1(+)/Mad1 was instilled intothe bladders. We observed if over expression of MAD1 can inhibit theproliferation of bladder tumor in situ . The first part of this paper was experimental study of MNU-inducedbladder tumor in rats. Bladder tumors were induced in SD rats byintravesical instillation of MNU through self designed metal catheters.2.5mg carcinogen was instilled into experimental animal by three weeks'vacation for 12 weeks. By a series of observations on the macroscopes andthe histopathological characteristics of experimental animals,we cancompare the differences between model group and conrtrol. We sucessfullyestablished model of MNU-induced bladder tumor. The histologicalchanges and pathological features of tumors are similar to those of human.A 100% induced rate of bladder tumor was achieved in model group after12 weeks. The rates of transitional carcinoma,squamous cell carcinomaand their mixed type were 81.0%,14.3% and 4.7%,respectively. The second part of this paper was experimental study on the effects ofMAD1 to bladder tumor in vivo. Bladder tumors were induced in SD ratsby intravascular instillation of MNU. The rats bearing tumor wererandomly divided into 3 groups: transfected with pcDNA3.1(+)/Mad1(group A),transfected with empty vector(group B) and transfected withsaline (group C). Tumor-bearing rats were transfected using Lipofectamine.Rat body weight (RBW), Bladder absolute weight (BAW) ,bladderrelative weight (BRW) and expression levels of mRNA and protein ofMad1 and TERT were assayed and electron microscope examination andflow cytometer analysis were used to observe the inhibition effect of Mad1to bladder tumor. Compared with group B and group C,in group A,though there was no diference in RBW (P>0.05), BAW and BRW weresignificantly decreased (P<0.01 and P<0.05, respectively). Mad1 mRNAand protein expression levels were markedly improved. TERT mRNAexpression levels were markedly decreased. Morphologic changes ofapoptosis were found in tumor cells by electron microscope in Mad1transfected group. Flow cytometry showed an increase of G0/G1-phase cellsand a decrease of S-phase cells after transfected with Mad1. CONCLUSIONS : The presented method was simple,rapid andeffctive for constructing the orthotopic model of bladder tumor in rats. Thehistopathological features of SD rat bladder tumors resembled those ofhuman,most of which were bladder transitional cell carcinomas. Theconstruction of bladder tumor model induced by MNU in SD rat lays afavorably foundation for further study on the therapy effect by bladderinstillation of drugs and prevention of bladder tumor. Animal exprementhas proved over expression of MAD1 can inhibit the proliferation ofbladder tumor. The correlative mechanisms include: 1. down-regulateTERT expression of bladder tumor cells; 2. stagnate bladder tumor cells inthe Gl-phase and induce S-phase arrest and accelerate differentiation; 3.enhance apoptosis of bladder tumor cells.