The Expression Of Co-stimulatory Molecule CD40 On Human Cervical Carcinoma And The Biological Roles Of CD40 Signaling On SiHa

Invasive cervical carcinoma is one of the common malignant tumors in woman, ranking the 2nd place among all of female malignant carcinomas, only second to breast cancer. It was reported that incidence and mortality of cervical carcinoma trended to be younger in recent years. The incidence among women of less than 35 has been increasing. So it was important and necessary to investigate the disease etiologically and pathogenesis, in order to explore more sensitive was for diagnosis and more effective therapy of cervical carcinoma.CD40, a 48KD cell membrane molecule, belongs to the tumor necrosis factor receptor(TNFR) superfamily. CD40 is expressed in many cell types including B cell, dendritic cells, monocytes, endothelial cell, epithelia cell and also many carcinoma cells. CD40 ligand is expressed mainly in activated CD4+T helper cell. Interaction by CD40L-CD40 is a main co-stimulatory signal during antigen-specific immune response and during immune regulation. To date, an increasing number of studies showed that signals mediated by CD40 could contribute to B cell proliferation, Ig secretion and class switching; dendritic cell activation and maturation; adhesion molecules expression on endothelial cell, inflammatory factor secretion, and the accumulation of white cells to the inflammatory sites. These indicated that CD40 is a very important molecule during immune response.In this study, the expression of CD40 on human cervical carcinoma and their clinical significance were focused. The expression of CD40 on cervical carcinoma cell line SiHa and its biological effects were studied. Moreover we studied the effects of CD40 mAb on the chemosensitivity of human cervical tumor cell line SiHa to Gemcitabin. We hope to find a new method of tumor immunotherapy. 1. the expression and clinical significances of CD40 and related molecules in human cervical carcinomaObjective: To study the expression of CD40 and associated molecules HPV16/18 E6, P16INK4a, VEGF and CD34 on different types of malignant cervical cancer, and to study their clinical significance. Methods: Surgical biopsy specimens were obtained from 239 patients (includes: 56 cases cervical squamous epithelium cancer, 36 cases of cervical intraepithelial neoplasia (CIN I), CIN II 39 cases of, CIN III 27 of, 43 cases of chronic cervical inflammation and 38 normal controls) . The expression of CD40 and associated molecules HPV, P16INK4a, VEGF and CD34 on different types of malignant cervical cancer were examined by immunohistochemistry. The relation of CD40 and associated molecules and clinical significance were analyzed. Results: The expression of CD40, HPV16/18 E6, P16INK4a, VEGF and CD34 on cervical cancer and CIN III have significant differences from the normal controls, and CIN I~II. The rates of CD40 positive cell in the 56 cases of cancer and in the 27 cases of CIN III were 55.55% and 67.86%, respectively (p<0.01). The positive expression rates of CD40, HPV16/18 E6, P16INK4a and VEGF were 78.6%, 80.36 and 87.50% respectively, and the CD34 MVD was 27.02±10.68. The positive expression rates of P16INK4a were 90.40% and 97.72% in CIN III and cervical cancer respectively. The expression of P16INK4a was significantly associated with HPV16/18 E6 in CIN III and cervical cancer (r=0.362 and r=0. 837, p<0.01 ). CD40 expression had a positive correlation with HPV16/18 E6 (r=0.57, p<0.01), P16INK4a(r=0.52, p<0.01), VEGF(r=0.32, p<0.05) and CD34 MVD(r=0.37, p<0.05). Further more, the expression of CD40 has a close correlation to lymph nodule metastasis(r=0.44, p<0.01) and invasive extent of muscular layer(r=0.30, p<0.05) in cervical cancer. Conclusion: These results suggested that CD40 might be a valuable factor for diagnosis and judgment of prognosis and lymphonudule metastasis in cervical cancer. These data also indicated that CD40 was an important molecule to tumor development.2. the biological effects of CD40 ligation on human cervical squamous epithelium cancer cell line Objective: To study the effect of CD40 mAb on the chemosensitivity of human cervical tumor cell line SiHa to Gemcitabin. Methods: Flow cytometric analysis was used to detected expression of CD40 in SiHa. Agonistic anti-human CD40 monoclonal antibody (5C11) was added to the cell culture system. PI staining and Annexin V-PI assay were used to study the biological effects of 5C11 on cervical tumor cell. Cell proliferation was analyzed by MTT assay after treatment with chemotherapeutic drugs. Expression of apoptotic genes mRNA was evaluated by quantitative RT-PCR. Results: The cervical tumor cell line SiHa highly expressed CD40. 5C11 or Gemcitabin could not effectively induced SiHa proliferation arrest, but combination of 5C11 and Gemcitabin could do. CD40 activation can induce arresting the cells at G2/M interphase and down-regulating anti-apoptotic genes BCL-XL expression and up-regulating apoptotic genes BAX expression weakly. Conclusion: CD40 activation could enhance chemosensitivity of human cervical tumor cell to Gemcitabin by inducing arresting the cells at G2/M phase and affecting apoptotic genes expression.