Anti-leukemia Effect And Mechanism Of Malaria Parasite Infection In A Murine Model

Leukemia,a group of hematologic malignancies disorders in leukocytes,is characterized by the uncontrolled proliferation and blocked in differentiation of hematopoietic cells.The classification of leukemia is complex,and its pathogenesis is poorly understood.The current clinical trials for leukemia include pharmaceutical medications,debilitating radiation,and bone marrow transplant therapies.Although lots of advances have been made in leukaemia treatment,survival rates remain low,with 5-year survival rates of 60%in young and 10%in elderly patients,respectively.Thus,new targets for treating leukemia are necessary.In the last few years,different research groups had independently reported an adverse relationship between some parasite infections and cancers.Anticancer activity of some parasites such as Trypanosoma cruzi,Toxoplasma gondii,Toxocara canis,Acantamoeba castellani and Plasmodium yoelii had been shown in experimental animals.Use of malaria infection as a treatment for other diseases was named as Malariotherapy.In 1981,Greentree LB put forward the idea of malariotherapy for cancer treatment.Malariotherapy had been successfully applied to the treatment of neurosyphilis in the history.In the 1950s,Professor Werner Zabel from Germany treated l9 cases of cancer patients with P.vivax infection.Chinese scholars such as Chen Xiaoping,as early as in 1980s,used malaria parasite infection for clinical treatment of some cancer patients,and also achieved some success.Statistical data from WHO in 1980 showed that the incidence of cancer in the malaria endemic areas of the world was the lowest.Giugni reported that the cancer mortality rate decreased almost half in the malaria endemic areas,compared with that in non-epidemic areas.In India,the incidence of malaria was high and cancer was very rare in the 1950s.However,cancer cases increased along with decreasing malaria cases in the 1980s.The literature was documented that hundreds of patients with malignant tumor were rehabilitated after infected with some pathogens,among them at least 150 patients suffered from malaria or typhoid fever.In the northern region of Henan Province,China,known as non-endemic areas,the incidence of esophageal cancer was high,while it was very low in the malaria endemic areas in its southern region.In animal models it had been shown that malaria parasite infection was able to inhibit some solid tumor growth.However,the use of malaria parasite infection for the treatment of leukemia has not been reported.In this study,murine leukemia model was established through inoculating BALB/c mice with acute myelomonocytic leukemia cell line WEHI-3,and the mice were infected with non-lethal type Plasmodium(Plasmodium yoelii 17XNL).The inhibition effect of malaria parasite infection on murine leukemia and its mechanism were explored.This paper includes two parts:I.Anti-leukemia Effect of Malaria Parasite Infection;Ⅱ.Mechanism of Anti-leukemia Effect of Malaria Parasite Infection.Part Ⅰ Anti-leukemia Effect of Malaria Parasite InfectionObjective:To study anti-leukemia activity of malaria parasite P.yoelii infection in leukemia WEHI-3 cells bearing mice.Methods:Female BALB/c mice were divided into the following 5 groups(n=10):control group(Group A);Plasmodium infection group(Group B);leukemia group(Group C);Plasmodium therapy group(Group D);Plasmodium prevention group(Group E).Group B i.p.inoculated with 1 x 10~5 P.yoelii parasitized erythrocytes(PRBC)per mouse;Group C i.p.inoculated with 1×10~5 WEHI-3 cells per mouse;Group D i.p.inoculated with 1×10~5 WEHI-3 cells and then i.p.inoculated with1×10~5 PRBC 1 week later;Group E i.p.inoculated with 1×10~5 PRBC and then i.p.inoculated with 1×10~5 WEHI-3 cells 2 weeks later.The general situation of each mouse was observed daily after WEHI-3 cells inoculated,including hair color,weight,mental state,activity and so on.Mice from each group were sacrified at days 22 post inoculation of WEHI-3 cells.And bone marrows were smeared,spleen and liver samples were used for histopathology.Results:Mice began to appear symptosis,back arch,piloerection,dispirited,abdominal swelling,moving slow and poor mental state from 2 weeks post inoculation of WEHI-3 cells.The percentage of myeloblastin in mice at day 22 post inoculation of WEHI-3 cells was more than 30%(leukemia can be diagnosed when the immature cells in the bone marrow≥20%according to diagnostic standard established by WHO).Abundant neoplastic cells were found in the liver and spleen tissues.All these results indicated that murine leukemia model was successfully established.After infected of P.yoelii 17XNL,the percentage of myeloblast in the mice incubated with WEHI-3 was significantly decreased(p<0.001).The neoplastic cells in the liver and spleen tissues from Plasmodium therapy group and Plasmodium prevention group were either obviously decreased or hard to find compared with leukemia group.Conclusion:These data showed that malaria parasite infection could not only treat but also prevent development of leukemia in a mouse model.Part Ⅱ Mechanism of Anti-leukemia Effect of Malaria Parasite InfectionObjective:To study the mechanism of anti-leukemia activity of Plasmodium yoelii infection on leukemia WEHI-3 cells bearing mice.Methods:Female BALB/c mice were divided into the following 5 groups(10animals per group):control group(Group A);Plasmodium infection group(Group B);leukemia group(Group C);Plasmodium therapy group(Group D);Plasmodium prevention group(Group E).Group B i.p.inoculated with 1 x 10~5 P.yoelii parasitized erythrocytes(PRBC)per mouse;Group C i.p.inoculated with 1×10~5 WEHI-3 cells per mouse;Group D i.p.inoculated with 1×10~5 WEHI-3 cells and then i.p.inoculated with 1×10~5 PRBC 1 week later;Group E i.p.inoculated with 1×10~5 PRBC and then i.p.inoculated with 1×10~5 WEHI-3 cells 2 weeks later.Mice from each group were sacrified at day 22 post inoculation of WEHI-3 cells.Leukocytes of blood in each mouse were separated,the levers of CD3、CD19、CD11b and Mac-3 were analyzed by flow cytometry.The levels of IFN-γand TNF-αin individual serum were measured by ELISA kits.Peritoneal macrophages were harvested,phagocytic activity was measured by phagocytic kit and analyzed by flow cytometry.The splenocytes were isolated from fresh spleen of each mouse,the effectorcells(splenocytes)and target cells(YAC-1 cells)were incubated,NK cell activity was determined by flow cytometry.Results:Malaria parasites infection significantly increased the levels of T cells(CD3~+)and B cells(CD19~+)in leukemia mice,and decreased the levels of monocytes(CD11b~+)and macrophages(Mac-3~+)in leukemia mice(p<0.001).The results indicated that malaria parasite infection promoted the differentiation of the precursor of T and B cells,and inhibited the differentiation of the precursor of monocytes and macrophages.Malaria parasites infection increased the productions of IFN-γand TNF-αin leukemia mice.Malaria parasites infection also increased phagocytic activity of peritoneal macrophagein and NK cell cytotoxicity in leukemia mice.Conclusion:Malaria parasites infection showed anti-leukemia effects in malaria parasites prevention group and malaria parasites therapy group.Anti-leukemia effects of malaria parasites infection maybe achieve through induction of innate and adaptive immunity.