Bearing Human Cervical Cancer In Nude Mice Model, Il-8 And Il-8 Neutralizing Antibody, Il-8 Of Shrna On Tumor Growth

Cervical cancer is the global disease incidence rate occupies the second woman malignant tumor, comes first in of feminine malignant tumor in our country. our previous study have been selected the differential expression gene by cDNA gene chip and found many genes correlated strongly with lymph node metastasis of early squamous cervical cancer. In these genes, the expression of IL-8 in early squamous cervical cancer with lymph node metastased was higer 3.4 times than with no metastatic lymph node. our previous study also have been shown that IL-8 might enhance early cervical cancer to invase and metastases by stimulate directly cervical cancer cells proliferation, migration,enhance tumor-endothelial adhesion and promoting angiogenesis in vitro experiment. Also using IL-8 shRNA silences the IL-8 gene in the Caski cell line, confirmed that IL-8 shRNA can suppress the cervical cancer cell proliferation, migration and the adherency effectively. To further expound that IL-8 gene expression in the cervical cancer carcinogenesis development function, this experiment plans through the establishment IL-8 related bearing human cervical cancer nude mouse model, observes IL-8 in nude mouse in vivo the influence which grows to cervical cancer nude mouse transplant lump and whether to have the lymph metastases. Using the IL-8 monoclonal antibody and IL-8shRNA in nude mouse in vivo, observes it the inhibitory action which grows to cervical cancer nude mouse transplant lump.Purpose: (1) Establishes the IL-8 related bearing human cervical cancer nude mouse model, studies IL-8 in nude mouse the influence which grows to cervical cancer nude mouse transplant lump and lymph migration. So it might be one of the the theory basis for the clinical discussion cervical cancer metastases mechanism.(2) Using the IL-8 monoclonal antibody and IL-8shRNA in nude mouse in vivo, observes it the inhibitory action which grows to cervical cancer nude mouse transplant lump,to treats cervical cancer for the clinical gene target to provide the theory basis.Method: (1) Using high express IL-8 the IL-8pcDNA3.1-IL-8-SiHa cell line, the CaSki cell line and silences IL-8 the PGenesil-1-IL-8shRNA-Caski cell line to vaccinate the nude mouse hypodermic, establishes the IL-8 related bearing human cervical cancer nude mouse model. (2)Observe the different cell vaccination in nude mouse hypodermic the situation of tumor growth, such as regular survey tumor volume size to plan tumor growth curve and weigh tumor after nude mouse death, computation tumor promotion rate or suppression rate and life time. (3) Observe tumor and the lymph node cytomorphology change by the HE staining; With RT-PCR, ELISA, TUNEL, immunohistochemistry methods further examines in the nude mouse tumor and serum the IL-8 expression(.4)To expresses IL-8 with the IL-8 monoclonal neutralizing antibody in the high expression of IL-8 in two models in nude mice bearing human cervical cancer to carry on the treatment. The 4th day after the vaccination, we starts intraperitoneal injection the IL-8 monoclonal antibody at the high, middle and low three dose group. Anti-tumor effects were observed using the forementioned similar method plan tumor growth curve, the rate of inhibition , the HE staining observation cytomorphology change and examination in the nude mouse tumor and serum the IL-8 expression with RT-PCR, ELISA, TUNEL, immunohistochemistry methods.Results: (1) Successfully established IL-8 related bearing human cervical cancer xenograft model in nude mice. (2) Tumors in nude mice found that high expression of IL-8 in pcDNA3.1-IL-8-SiHa cells significantly enhance the ability of tumor volume and weight were higher, and promote tumor rate was 84.5%; axillary lymph nodes increases in the experimental group, HE staining found that cell volume is big, the nuclear staining is deep but the control group had no significant changes;RT-PCR, immunohistochemistry and ELISA results showed that nude mice tumor and serum IL-8 expression compared with the control group increased significantly. (3) silences IL-8 the PGenesil-1-IL-8shRNA-Caski cell was significantly reduced tumor volume and weight was significantly lower than the control group, the inhibition rate was 71.0%; HE staining of cell division found rare, organizations have more necrotic area; TUNEL revealed that apoptosis significantly increased; RT-PCR, immunohistochemistry and ELISA results showed that nude mice tumor and serum IL-8 expression compared with the control group decreased significantly. (4) CaSki cells and pcDNA3.1-IL-8-tumor SiHa cells after IL-8 monoclonal antibody treatment was significantly reduced tumor volume and weight, the inhibition rates respectively were 76.6% and 72.0%; HE staining tumor growth that is not the experimental group better, smaller cell size, cell division of the rare and organizations have more necrotic area; TUNEL Fluorescence display apoptosis significantly increased; RT-PCR, immunohistochemistry and ELISA result discovered that in the nude mouse tumor and the serum IL-8 expression compares with the control group reduces obviously.Conclusion: (1) IL-8 gene that can promote cervical cancer xenografts in nude mice induced tumor effect and lymph node metastasis in nude mice in vivo. (2) IL-8 gene silence in CaSki cells significantly inhibited the growth of cervical cancer xenografts in nude mice. (3) IL-8 monoclonal antibody significantly inhibited the growth of cervical cancer xenografts in nude mice,and along with IL-8 monoclonal antibody dosage's increase, the tumor volume also reduces, proved that the anti-tumor effect and between the injection antibody dosage has the quantity - effect relations. So they might be one of the ideal strategies for gene therapy of cervical cancer.