| Part one The biological significance of CD40 in lung cancerObjective To investigate the expression of CD40 with associate molecules p53 and C-raf-1 on lung cancer and its clinical significance .To measure the serum level of soluble CD40 (sCD40) in healthy volunteers as well as in patients with lung cancer and evaluate the sCD40 level in both benign pleural effusion and malignant pleural effusion. The significance of sCD40 in differentiation of benign pleural effusion between malignant pleural effusion was analyzed. The effects of cross-linking of agonist CD40 monoclonal antibody (5C11) and rhCD40L on the growth, the proliferation, cell cycle and cell apoptosis in lung cancer cell lines (including CD40 positive and negative cell lines) as well as the phenotype changes were explored. The possible mechanism of CD40 involved in the pathogenesis of lung cancer and CD40 as a potential therapeutic target was assessed.Methods Surgical biopsy specimens were obtained from 51 patients who underwent pulmonary resection for primary lung cancer. The expression of CD40 with associate molecules p53 and C-raf-1 on different types of malignant lung cancer tissuewere examined by immunohistochemistry. The correlation of CD40, p53 and C-raf- 1 with clinical features and pathological pattern were analyzed. The expression of CD40 on the surface of cancer cells from 10 fresh lung cancer tissues and 4 lung tumor cell lines were also measured by immunofluorescence technique and flow cytometry. The level of sCD40 expression was tested by enzyme linked immunosorbent assay (ELISA). In order to investigate the possible function of CD40 in lung cancer, agonist CD40 monoclonal antibody (5C11, 1 Dug/ml) and recombinant CD40 ligand (rhCD40L, 2u.g/ml) were used to co-incubate with lung cancer cell lines in vitro, then phenotype changes on the cell surface and alterations of tumor cell cycle or apoptosis were monitored. [3H]-TdR incorporation method was used to determine the proliferative activity of tumor cells after using 5C11 and rhCD40L.Results 1. CD40 positive cells showed a staining on cell membrane, p53 positive cells in cell nucleus while C-raf-1 positive cells on endoplasm. The rate of CD40 positive cells in the tumor biopsy tissues was 76.5% (39/51) with an overexpression of 54.9.1% (28/51). The expression of CD40 had a close relation to clinical stage and lymph nodule metastasis. There was no relationship with pathological grade and pathological type (p>0.05). CD40 positive cells varied from 2.0% to 88.2% among four lung tumor cell lines while among 10 fresh tissues of lung cancers including 4 adenocarcinoma samples, 5 squamous carcinoma samples and 1 small cell lung carcinoma sample, the rate of CD40 positive cells varied from 1.6% to 71.0%.2.The rate of p53 positive cells in the tumor biopsy tissues was 74.5%(38/51) with an over-expression of 45.1% (23/51). There was a close relation of p53 overexpression to local lymph nodule metastasis (p <0.05). But p53 expression did not had a close relation to pathological type, pathological grade and clinical stage of lung cancer (p>0.05).3.The rate of C-raf-1 positive cells in the tumor resection tissues was68.6%(35/51) with an over-expression 53.0%(27/51). The C-raf-1 expression had a close relation neither to pathological type, pathological grade nor to clinical stage of lung tumor (p>0.05)).4. CD40 expression had a positive correlation with p53 expression (p <0.05) while C-raf-1 had not (p>0.05).5.The serum concentration of sCD40 in patients with lung cancer was 623.23 180.69pg/ml, significantly higher than those in healthy volunteers and in healthy donors which was 117.52+17.8 pg/ml and 116.25 18.72 pg/ml respectively(p O.01). No close relations among the serum concentration of sCD40, stage and pathological type of lung cancers were found. The serum concentration of sCD40 in patients with benign lung diseases was 656.37 304.92pg/ml and had not a significant difference compared with those in lung cancers (p>0.05). The concentration of sCD40 in benign pleural e... |
PDF Full Text Request