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Objective To investigate the expression of Twist,E-cadherin and N-cadherin in bladder urothelial carcinoma and evaluate their relationships with tumor.Methods From January 1998 to June 2007,the expression of Twist,E-cadherin and N-cadherin protein were detected in 180 bladder urothelial carcinoma tissue specimens in paraffin blocks and 60 normal bladder tissue specimens by tissue microarray and immunohistochemistry.Results Compared with normal hladder tissues,the expression of Twist and N-cadherin was up-regulated in bladder carcinoma tissues and the positive percentage was 63.9％ and 47.8％ respectively.The percentage of high E-cadherin expression in bladder carcinoma tissues was only 27.8％,which was significantly lower than that in normal bladder tissues.The expression of Twist was significantly related with age,tumor number,vessel invasion,distant metastasis and lymph node metastasis (P＜0.05).The expression of E-cadherin was significantly related with distant metastasis and lymph node metastasis (P＜ 0.05).The expression of N-cadherin was significantly related with distant metastasis and lymphnode metastasis (P＜0.05).There was significant positive correlation between Twist and E-cadherin expression (P＜ 0.05).There was significant positive correlation between Twist anl N-cadherin expression (P＜0.05).There was negative correlation between E-cadherin and N-cadherin expression.All the 180 bladder carcinoma patients were followed up after the surgery,and the mean follow-up time was (35.2±9.1) months.The 5-year survival rates of patients with low and high Twist expressions were 67.7％ and 46.1％ respectively,and there was significant difference between the two groups (P＜0.05).The multivariate analysis indicated that the Twist expression was an independent prognostic factor of bladder carcinoma.Conclusions The high expression of Twist,E-cadherin and N-cadherin are involved in the processes of invasion and metastasis of bladder urothelial carcinoma.The Twist expression was one of the independent prognostic factors of bladder urothelial carcinoma.
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Objective To predict the secondary structure and the B cell epitopes of human heparanase protein, and to identify its immunogenicity. Methods The flexible regions of secondary structure and the B cell epitopes of human heparanase amino acid sequence were predicted by DNAStar and Bcepred software. The multiple antigenic peptides (MAP) of the epitopes were synthesized in 8-branch form. Rabbits were immunized with the 8-branch MAPs mixed with a universal T-helper epitope human IL-1β peptide (VQGEESNDK, amino acid 163-171 ). The immunogenicity of the synthesized peptides was evaluated by ELISA, Western blot and immunohistochemistry. Results Amino acid 1 -15 ( MAP1), 279-293 (MAP2) and 175-189(MAP3) of large-subunit of human heparanase protein was predicted as the most potential epitopes of human heparanase protein. All the three synthesized MAPs induced high titer of antibodies. ELISA, Western blot and immunohistochemistry analysis showed all the three MAPs could produce high titer serum antibodies, antibodies induced by MAP1 and MAP2 had high specific binding activity , and MAP2 antibody showed the strongest binding activity with liver cancer tissues. Conclusion The large-subunit No. 1-15, 279-293 amino acid of human heparanase protein may be the B cell preponderant epitopes and the strongest immunogenicity may be No. 279-293 peptide, which provided a theoretic basis for the antibody and vaccine development of heparanase subunit peptide.
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potential use in clinical therapy and prognosis estimation of the tumor.
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Objective To select and identify human leukocyte antigen(HLA)-A2-restricted T-cell epitope within heparanase(Hpa),and to provide aeademic bases for peptide-based immunotherapy with malignant tumor.Methods The Hpa sequence was scanned for prediction of the immunogenic peptides-based CTL epitopes using the HLA-binding prediction software SYFPEITHI and BIMAS.Ten the affinity of candidate epitopes to HLA-A2 was evaluated by T2 binding test.Acfivation of T cell was assessed by ELIS-POT and cytotoxictiy by lactate dehydrogenase(LDH)release assay.Results of the six predicted nona-peptides,Hpa(310-318,FLNPDVLDI)showed high affinity of binding to HLA-A2 and led to IFN-γ secre-tion in vitro.Furthermore,Hpa(310-318) FLNPDVILDI could induce PBMC from a HLA-A2 positive healthy donor to lyse specifically HCC-LM6 and SW-480 cells which expressing both Hpa and HLA-A2.Conclusion The nona-peptide Hpa(310-318)FLNPDVLDI may be all HLA-A2-restricted CTL epitope.which is capable of inducing Hpa-specific CTL in vitro.
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Objective To explore available immune strategy to improve immunological effects of the B cell epitopes of human heparanase protein.Methods Based on predicted potential B cell epitopes of human heparanase protein,three candidates of multiple antigenic peptides(MAP)with 8-branches were synthesized and identified by conjugation to antibody against full human heparanase protein.C57BL/6 mice were immunized with the 8-branch MAPs mixed with or without the universal human T-helper IL-1β peptide.The antibody titers of immune serum was assayed by ELISA.Results Most potential epitopes of human heparanase protein were predicted as No.1-15(MAP1),279-293(MAP2)and 175-189(MAP3)in large subunit of human heparanase protein.All three synthesized MAPs mixed with the T-helper epitope induced higher titers of antibodies than them without the T-helper epitope.Conclusion The three B cell epitopes of human heparanase protein could he its B cell preponderant epitopes.The 8-branch MAPs mixed TH cell epitope may be an available and optimizing immune strategy.