| Bladder carcinoma is the most common tumor in urinary system.Worldwide, bladder carcinoma incidence rate is ninth place in malignant tumors, ranked sixth in men ranked tenth place after in women; In China, male bladder carcinoma incidence ranked eighth in systemic tumors,ranked twelfth place after in women,which is severely threatening people's health. The mechanism of bladder carcinoma is still not fully understood, but with other organs tumors, many risk factors for cancer work for a long time in normal cells, causing proto-oncogene activation, then after a series of complex signaling affect the process of cell cycle. Because of tumor suppressor gene inactivation or missing in the duplication and transcription process, neither making the damaged DNA repaired,nor making the cells into normal apoptosis, the cells that carrying abnormal DNA unrestricted copy. proliferate and cause cancer.As the member of the inhibition of apoptosis gene family,livin gene was found in 2000 ,located on human chromosome 20 q13.3, full-length 4.6 kb, contains seven exons and six introns, the gene transcription product mRNA has two subtypes, separately encoding 298 amino acid protein (α) and 280 amino acid protein (β). Like other IAPs family members,livin gene contains BIR and RING finger domain,mainly through interaction with the caspase family, inhibits caspase mediated cascade activation reaction of apoptosis, prompting tumors. Livin is not expressed in the majority of normal adult tissues (except placenta).In epidermis, lymph nodes, spleen and melanoma cells it has a very low expression. But in the process of fetal development,livin has a high expression in variety tissues, such as fetal brain, thymus. kidney and liver.It is found that some cancer cell line at high livin expression, such as lymphoma, HaCat cells, MCF7 breast cancer cells , especially melanoma cells expression in the highest level. In bladder carcinoma cells, both livinαandβsubtypes expressed,however, livin, caspase-3, MVD expression and tumor recurrence study in bladder carcinoma has yet to see reports. In the present study,we detected the expression of livin protein,caspase-3 protein as well as the expression of MVD in normal bladder tissue and bladder carcinoma tissue by immunohistochemical SP method, analyzing its correlation.ObjectiveTo explore the role of livin, caspase-3 and MVD in bladder cancer occurrence and development, hoping to provide the new methods to the clinical diagnosis ,treatment and prognosis of bladder carcinoma.Materials and MethodsThe 65 cases of surgically resected bladder carcinoma tissues were collected in the first affiliated hospital of zhengzhou university from January 2006 to December 2006 .There were 38 cases of males and 27 females, the age ranged from 29 to 78 years old, average 62.5 years old. None had received preoperative radiotherapy or chemotherapy. Pathological types were bladder transitional cell carcinoma. According to WHO 1973 years histological grade: I grade 25 cases, II grade 28 cases,III grade 12 cases;clinical stage: Tis-T1 period of 39 cases, T2-T4 period of 26 cases.Postoperative follow-up of 1.5～2 years, one of 28 cases of postoperative recurrence, 37 patients without recurrence. At the same time take 10 cases of normal tissue adjacent to cancer (All adjacent normal tissues were obtained more 5 centimeters away from the edge of tumor) for the contrast group.All the specimens were fixed with formalin, and embedded routinely with paraffin, every section was 4um.Polyclonal rabbit anti-livin antibody, polyclonal rabbit anti-caspase-3 antibody and monoclonal mouse anti-CD34 antibody were used to detect the expression of livin, caspase-3 and CD34 protein in 65 cases of bladder carcinoma, 10 normal bladder tissues by immunohistochemistry SP method. SPSS10.0 software were used to analyze the date.All bladder carcinoma specimens according to the pathological grade, clinical stage, with or without recurrence classify statistics,each indicator using X~2 test, Fisher's exact test or t test analyzed.The correlation between livin and caspase-3 using of Pearson product-moment correlation (Pearson) correlation analysis, a = 0.05 was considered the level of significance.Results1.The positive staining of livin protein was mostly located in the cytoplasm and/or nucieus,mainly in cytoplasm of cancer cell. 53 cases in 65 cases of bladder carcinoma showed positive expression, the positive rate was 81.54% (53/65),10 cases of normal bladder tissue were not positive expression. Livin protein expressions in bladder carcinoma were significantly higher than those in normal bladder tissues, there was significant difference(P = 0.000). In I , II, III grade, the positive rates of livin protein were respectively 76.00% (19/25), 85.71% (24/28) and 83.33% (10/12), livin protein expressions had no significant difference in the tumor grading(P = 0.651); In stage Tis-T1 and T2-T4, the positive expression rates were respectively 79.49% (31/39), 84.62 ( 22/26), there wasn't significant difference between stage Tis-T1 and T2-T4 (P = 0.845); In the recurrence group and the non-recurrent group, livin protein positive expression rates were respectively 92.86% (26/28), 70.27 (26 / 37), and there was significant difference between the group with and without recurrence(P = 0.024).2.The positive staining of caspase-3 protein was mostly located in the cytoplasm and/or nucleus,mainly in cytoplasm of cancer cell. 30 cases in 65 cases of bladder carcinoma showed positive expression, the positive rate was 46.15% (30/65). The positive rate in normal bladder tissues was 90.00%(9/10). Caspase-3 protein expressions in normal bladder tissues were significantly higher than those in bladder carcinoma, there was significant difference(P = 0.000). In I , II, III grade, the positive rates of caspase-3 were respectively 68.00% (17/25),35.71% (10/28) and 25.00% (3/12), caspase-3 protein expressions had significant difference(P = 0.017); by X2 partition, the standard of testingα= 0.0167, there was significant difference between grade I and III(P = 0.014). There was no significant difference between grade I and II (P = 0.019), neither no significant difference between grade II and III (P = 0.768). In stage Tis-T1 and T2-T4, the positive expression rates were respectively 48.72% (19/39), 42.31 (11/26), there wasn't significant difference between stage Tis-T1 and T2-T4 (P = 0.612); In the recurrence group and the non-recurrent group, caspase-3 protein positive expression rate were 32.14% (9 / 28), 64.86 (24/37), and there was significant difference between the group with and without recurrence (P = 0.009).3. In bladder carcinoma tissues, the negative expressions of caspase-3 protein(34 cases) in the positive expressions of livin protein(53 cases), there were the positive expressions of caspase-3(ll cases) in the negative expressions of livin protein(12 cases), both positive expressions were 19 cases, there was 1 cases having a common negative expression.Using Pearson product-moment correlation (Pearson) correlation analysis showed that: the expression of livin protein was a negatively correlation with caspase-3 protein in bladder carcinoma tissues (P = 0.000, r = -0.434).4.Vascular endothelial cells stained brown for CD34 positive. There is selecting five visions of the largest number of vascular in a microscope at 400 times perspective, counting the number of blood vessels, and calculating the average for MVD. In 65 cases of bladder carcinoma tissues, the MVD ranged from 6.20 to 38.25 at each high power field, average was 18.42±1.96. In 10 cases of normal bladder tissues, the MVD ranged from 1.25 to 3.21 at each high power field, average was 2.14±0.42. there was significant difference (P = 0.000). The averages of MVD expression in I , II, III grade were respectively 13.25±1.17,16.95±1.87 and 32.64±1.66, comparing to three groups had statistically significant differences (P <0.05); There was significant difference between each other group(P <0.05). In stage Tis-T1 and T2-T4, the averages of MVD expression were respectively 14.25±1.30,24.69±3.26, there were statistical significance between stage Tis-T1 and T2-T4 (P = 0.000); In the recurrence group and the non-recurrent group, The averages of MVD expression were respectively 22.63±2.61,12.86±1.15, comparing to two groups had statistical significance (P = 0.000).Conclusions1. Livin protein showed high expressions in bladder carcinoma tissues, but no expression in normal bladder tissues. The expression of livin protein may had no correlation with pathological grade and clinical stage; but livin expression had correlation with the availability of tumor recurrence, and it could be a marker to evaluate the prognosis of bladder carcinoma.2. Caspase-3 protein indicated high expression in normal, and low expressions in bladder carcinoma tissues. The expression of caspase-3 protein had no correlation with clinical stage, but it was correlated with pathological grade and tumor recurrence. It was suggested that down-regulation of caspase-3 expressions played an important role in carcinogenesis and prognosis of bladder carcinoma.3. In bladder carcinoma tissues, the expression of livin protein was negatively correlated with the expression of caspase-3 protein. It was revealed that the expression of livin inhibited caspase-3,which could be one of the mechanisms in the carcinogenesis of bladder carcinoma.4. In bladder carcinoma tissues.the MVD value was significantly higher than in normal bladder tissues. There were differences with pathological grade,clinical stage and tumor recurrence, it is suggested that MVD can be used as an indicator of biological behavior, to predict the risk of bladder carcinoma recurrence. |
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