Anti-tumor Effect Of Trichinella Spiralis IRNA On SP2/0 Tumor In BalB/C Mice

Trichinella spirilas was first discovered in human body by English Peacock in 1825 in Londen,given the name by Owen in 1835,called Trichinella for short.It can caused trichinelliasis that was a worldwide parasite desease between people and animals,infected people and more than 150 animals,did harm to them .Though it did bad to the people and animals very much ,there were reports that Trichinella could inhance the resistance of host to the tumor ,there was anti-tumor effect to tumor after the mice were infected the Trichinella spirilas and attenuated Trichinella spirilas or immunized by excretory-secretory (ES) antigen and different crude antigen of Trichinella,but different effect to different tumors.According to the foreigen reports , the mechanism of Trichinella spirilas anti-tumor may be that after Trichinella spirilas infecting the animals,it also can provocate immunity system of body ,so educe the anti-tumor effect.However nowadays it is not safe and mature to inject the Trichinella spirilas and attenuated Trichinella spirilas or excretory-secretory (ES) antigen and different crude antigen of Trichinella directly, for finding a safe and effective methed that can adopt immunity and be accepted by the people,this research combined with the features of iRNA together,used muscle larvae antigen as the immunizing antigen ,in voiding the worry that iRNA of the normal tissue caused autoimmunity damage, preparedTrichinella spiralis iRNA to observe the anti-tumor effect of Trichinella spirilas iRNA on SP2/0 tumor in BalB/C mice. Through immunizing animals,checking the valence of antibody to collect tissues for preparing the Trichinella spiralis iRNA,at last we checked the tumor morphology index,cellular immunity difference of animals and humoral immunity difference of animals.The experiment found that Trichinella spiralis iRNA inhibited the SP2/0 tumor growth in BalB/C mice successfully.The result of experiment established foundations for further study anti-tumor drug of Trichinella spiralis .The preparation of Trichinella spiralis iRNA Cultivated the Trichinella spiralis in the mice,used washing precipitation method to collect the muscle larva,then freezed and thawed it three times , broke it into pieces by ultrasound,then to prepare the muscle larva crude antigen,used different muscle larva crude antigen of protein dose to immunze the mice,rabbits and goats,used indirect ELISA to check the valence of antibody,as the standard of 1024 of valence of antibody to make sure the best immunizing dose ,meanwhile sacrificed the animals to collect the lymphoid node,spleen and liver, compared with the cold phonel and hot phonel to alternate for preparing the Trichinella spiralis iRNA,moreover checked the PH,appearance,quality,integrity,nucleic acid concentration,protein concentration,toxity and activity.The experiment found that for male 18-20g mice 300μg was the best immunizing dose,for male2.5-3kg rabbit 2mg is the best immunizing dose,for the male 15kg one-year goat 20mg is the best immunizing dose.Compared with two methods ,choosed the cold phenol for preparing the Trichinella spiralis iRNA at last because degradation was not easy to happen in this process of methed.The results ofcheking:the PH was from 6 to 8, OD260/OD280>2.0, OD260 / OD230>2.0, agarose gel electrophoresis show the 28s and 18s straps were obvious but 5s was not obvious,the nucleic acid concentration was high,protein concentration was smaller than 1%,non -undue toxicity, sterile, leucocyte cell adhesion inhibition ratio was mare than 20%.Anti-tumor effect of Trichinella spirilas iRNA on SP2/0 tumor in BalB/C mice Separated the mice for two groups,one was injected Trichinella spirilas iRNA ,then inoculated with SP2/0 myeloma cell,the other one was inoculated with SP2/0 myeloma cell, then injected Trichinella spirilas iRNA, there were 4 groups in the the former,there were 5 mice in each group,three of it were injected for 7days by 0.5mg,0.75mg and 0.1mg ,then inoculated with SP2/0 myeloma cell,the forth group was the control group in common, corresponding with the physiologic saline;there are 4groups in the latter too ,the same to the former ,there were 5 mice in each one,three of the groups were inoculated with SP2/0 myeloma cell,then injected the iRNA as 2mg,3mg and 4mg in the next day .The mice in the two groups were all sacrificed in 20 days later after inoculating with SP2/0 myeloma cell,then checked the shape of tumors,T lymphocyte of spleen of mice , humoral immunity difference.Accoding to the clinical observation,there was no one dead in 20 days,the bearing cancer ratio was 100%,one of the text groups did not grow tumor,the volume and weight of tumor in text groups were smaller than the control groups that were injected saline,moreover differences were significant(p<0.05). In the groups that were first injected iRNA,the anti-tumor effect of the group of 1mg/ml was the best.In the groups that were first inoculated with SP2/0 myeloma cell , the anti-tumoreffect of the group of 4mg/ml was thebest.The study used flow cytometry to check the T lymphocyte of spleen of mice difference, the quantity of CD3+,CD4+ and the ratio of CD4+/CD8 were higher than the control groups, particular that the ratio of CD4+/CD8 inhanced or degraded was direct ratio to anti-tumor effect.