Study Of Bifidobacterium Breve Acts As A Transfer Vector In The Genetherapy Of Breast Cancer

Objective To study the anti-tumor effect of IFNy and Bifidobacterium breve in mice bearing brenst cancinoma and its mechanism.Methods Restriction enzyme digestion and PCR using the recombinant plasmid. Detection of Bifidobacterium in the normal tissue and tumor tissues of mice in the distribution of 3H-TdR labeled with isotope Bifidobacterium,1,3,5 and 7d after injection mice were sacrificed on the neck dislocation, measured radioactivity of the liver, heart, lungs, kidney and tumor tissue.The 4T1 plasmids were injected locally into the tumors of the mice. When the tumor diameter is 5-8mm,the treatments began, the rats were randomly divided into four groups:control,Bifidobacterium breve(B-b),Bifidobacterium breve-pNZ44 and Bifidobacterium breve-pNZ44-IFNy。Measuring tumor size with vernier caliper, measuring every 3 days once tumor volume calculation.3 weeks, mice were killed, measuring tumor weight. Apoptosis using flow cytometry. Immunological parameters were detected in mice spleen CTL, NK cytotoxic activity, with the release of 3H-TDR methodResults Restriction analysis showed that the recombinant plasmid pNZ44-IFNγconstructed correctly. PCR identification showed that the recombinant prokaryotic expression vector pNZ44-IFNγhas been successfully transformed into Bifidobacterium. Experiment was divided into Normal+Bb (3H-TdR) and Tumor+ Bb (3H-TdR), pNZ44-IFNγin normal tissues of the changes over time is decreasing, the seventh days Tissue radioactivity measured activity on frist day has significantly decreased compared. Heart and liver tissue in the fifth days of radioactive activity has significantly reduced the lung and kidney in the first three days had significantly lower radioactive activity. pNZ44-IFNγactivity in tumor tissue radioactivity over time, changes are gradually enhanced, the threeth days of radioactive activity was significantly increased on seventh day tissue radioactivity measured on frist day compared with the activity has been a marked increase. In normal tissue radioactivity measured activity is reduced. liver, heart, lung, kidney radioactive activity in normal tissue and tumor tissue in mice with no significant difference. Tumor growth observed in each group size changes, the control group and blank plasmid group tumor growth fastest in relatively slow tumor plasmid alone, but pNZ44-IFNy plasmid group tumor growth was the slowest. B-b growth rate of tumor volume on fouteenth was significantly lower than in the control group, while the pNZ44-IFNy plasmid tumor volume growth rate in the fourth days had significantly lower than the control group. Detection of tumor weight in the control group and blank plasmid group the fastest tumor growth, tumor weight, the most important, while pNZ44-IFNy plasmid slowest tumor growth, compared to the control group significant difference. Detection of apoptosis by flow cytometry, Bb apoptosis rate compared with the control group were statistically significant, but Bb-pNZ44-IFNy group of the most significant tumor cell apoptosis, compared with the control group had significant differences. CTL were detected in mice spleen cells and NK cytotoxic activity, pNZ44-IFNy plasmid CTL and NK cell cytotoxic activity was significantly higher and the empty plasmid group, with statistical significance.Conclusions.This study, we constructed expression vector B-b-pNZ44-.IFNy, the plasmid has obvious anti-tumor activity. B-b-pNZ44-IFNy plasmid was superior in vivo antitumor effect of IFNy or B-b, inhibitory mechanisms may be combined to play the role of IFNy in immune regulation and Bifidobacterium in the area in tumor tissue hypoxia the role of selective enrichment. The innovation of this study is that the targeting vector Bifidobacterium and IFNy combination, give full play to their strengths and synergies, to enhance gene therapy has opened up new avenues and provide a new therapeutic agent for gene therapy The pre-clinical research laid the theoretical and experimental basis.The anti-tumor effect of double-gene-therapy is significantly better than that of single-gene-therapy.Its mechanism is perhaps associated with the expressions of IFNy and Bifidobacterium breve targeted inhibition of tumor, which enhance anti-tumor immunologic function and anti-angiogenesis function.