Effect Of H22 Tumor-bearing Mice Tumor Growth Inhibition And Expression Of Related Genes Sea Cucumber Acid Mucopolysaccharide

Objective:To observe the effect of Stichopus japonicas acidic mucopolysaccharide (SJAMP) in mice bearing neoplasia of H22hepatoma cells, and then, to study the effect and mechanism of SJAMP in H22hepatoma cells on cell proliferation and apoptosis by detecting the expression of related genes and proteins from the angle of molecular biology, to provide the basis for the further application of SJAMP.Methods:Model of mice bearing neoplasia of H22hepatoma cells was established, and then the50Kunming mice were randomly divided into5groups:negative control group(group A, normal saline), positive control group (group B,5-FU), low-SJAMP-dose group(group C, SJAMP6.25mg/kg), medium-SJAMP-dose group(group D, SJAMP12.5mg/kg), high-SJAMP--dose group(group E, SJAMP25mg/kg),10in each group and sex in half. Every mouse in each group was received an intraperitoneal injection administration of0.2ml/d with its equivalent medicine and dose, once daily lasting for12days. And then took off the cervical vertebra of mice to death. The histogenesis and growth pattern of liver cancer were observed. The changes in the tumor tissue and cells of mice in group A and SJAMP groups were observed by light microscope and transmission electron microscope. The immunohistochemical method was used to detect the expression of PCNA, P53, P21, C-myc, CyclinD1, CDK4, pRb, E2F-1, Bcl-2and Bax protein. The expression of P53, P21, C-myc, CyclinD1, CDK4, pRb, E2F-1, Bcl-2, and Bax mRNA were detected by Real-time PCR.Results:After the administration of intraperitoneal injection for12days, took off the mice to death. All tumors were separated and weighted under aseptic conditions. Tumors were appeared in each mouse. The average weight of other group was less than group A, and the difference between B,E groups and group A had statistical meaning (P<0.05). HE staining showed that except group A, SJAMP groups tumor cells were less, arranged loosely, karyopyknosis and necrotic area were increased. The result of transmission electron microscope showed each dose group of tumor cells were in different degrees of apoptosis morphology, including cell shrinkage, chromosomes condensation, and apoptotic body. Immunohistochemical results showed that compared with group A, SJAMP groups the expression of PCNA, P53, C-myc, Cyclin D1, CDK4, E2F-1and Bcl-2protein decreased, and the differences were significant (P<0.05),the expression of P21, pRb and Bax protein decreased, and the differences were significant (P<0.05). Real-time PCR results showed that compared with group A, SJAMP groups the expression of P53, C-myc, Cyclin D1, CDK4, E2F-1and Bcl-2mRNA decreased, and the differences were significant (P<0.05), the expression of P21, pRb and Bax mRNA decreased, and the differences were significant (P<0.05).Conclusion:A dose of SJAMP could inhibit the growth of H22hepatoma cells. The mechanism might be SJAMP could adjust the expression of PCNA protein, P53, p21and C-myc mRNA and protein and affect the Rb-E2F pass way by regulating the expression of CyclinD1, CDK4, pRb, E2F-1made H22hepatoma cells arrested in cell cycle, to inhibit tumor cell proliferation. And SJAMP can also regulate the expression of Bax, Bcl-2mRNA and protein to promote tumor cell apoptosis.