The Regulatory Mechanism Of Trichinella Spiralis Co-infection On The Immunopathology Of Liver And Small Intestine In Mice Infected With Plasmodium Berghei

Malaria is still one of the parasitic diseases that seriously threaten human health.Malaria infection can manifest as a febrile illness that is not severe,but can also develop into severe malaria such as cerebral malaria,severe anemia,hypoglycemia,acidosis,acute renal failure,jaundice,pulmonary edema,or acute respiratory distress syndrome.The complications of severe malaria are often associated with the host immune imbalances such as excessive immune responses in the body and/or overgrowth of Plasmodium parasites.Although current antimalarial drugs are effective in controling the parasite’s reproduction,they are not effective in curing severe malaria complications.Therefore,maintaining the immune balance in the host is very important for controlling the progress of malaria.A large number of epidemiological investigation have shown that the geographical distribution of malaria-endemic areas and helminth endemic areas are overlapped,and the co-infection of Plasmodium and helminth is widespread worldwide,especially in the poor areas of Africa,South America,and Southeast Asian countries.Co-infection can affect host susceptibility to parasite infection and host immune regulation.Currently,studies on the co-infection of Plasmodium and helminth are mainly focused on the epidemiological investigations,while studies on the interactions between the co-infected parasites and the effects of these interactions on the host and the mechanisms that may be involved are relatively rare.Objective:In this paper,a Kunming mouse model of co-infection of Trichinella spiralis(Ts)and Plasmodium berghei ANKA(PbANKA)was established to explore the immunomodulatory effects of Ts co-infection on the immunopathology of liver and the immune response of small intestine in PbANKA infected mice.Methods:1.One hundred specific pathogen free Kunming mice(6-8 weeks,18-22 g)were randomly divided into four groups(n=25),e.g.,the naive group;Ts-mono-infected group(Ts group),orally infected with 20 T.spiralis larvae;PbANKA-mono-infected group(Pb group),intraperitoneally injected with 1×106 PbANKA-infected red blood cells;Ts+Pb-co-infected group(Ts+Pb group),orally infected with 20 T.spiralis larvae 9 days previously and then challenged with 1 ×106 PbANKA-infected red blood cells.2.The survival of mice was monitored daily.The body weight and parasitemia of mice were monitored from day 3 after PbANKA infection.3.On the 22 days after Ts-infection and/or 13 days after PbANKA-infection,the liver and spleen of each group were collected for the detection of the liver and spleen indexes,alanine aminotransferase(ALT)and aspartate aminotransferase(AST),liver tissue parasite burden,liver pathology,immunohistochemical staining of galectin(Gal)-1、Gal-3、neutrophils elastase and CD68+macrophages in liver and spleen tissues.In addition,qRT-PCR was used to detect the mRNA expression levels of eosinophil-associated cytokines and inflammatory-associated cytokines in liver and spleen tissues of mice in each group.4.The spleen tissues of mice in each group were collected on the 22 days after Ts-infection and/or 13 days after PbANKA-infection,and the mRNA expression levels of CD200/CD200R were detected by using qRT-PCR.5.The small intestine tissues of mice in each group were collected on 22 days after Ts-infection and/or 13 days after PbANKA-infection,qRT-PCR was used to detect the mRNA expression levels of M1-type/M2-type macrophage markers in each group.6.Peritoneal macrophages of mice in each group were isolated on the 22 days after Ts-infection and/or 13 days after PbANKA-infection,which were observed by transmission electron microscopy and the mRNA expression levels of Gal-1,Gal-3,and macrophage polarization marker were detected by using qRT-PCR.Results:1.In vivo experiments showed that Ts co-infection had no significant effect on the survival rate of PbANKA-infected mice,but significantly reduced the parasitemia.Ts co-infection significantly reduced the parasite burden in liver tissue of PbANKA-infected mice.Ts co-infection significantly increased the liver and spleen indexes,and the liver pathology of PbANKA-infected mice.Ts co-infection significantly increased the mRNA and protein expression levels of Gal-1 and Gal-3 in both liver and spleen tissues of PbANKA-infected mice.Ts co-infection significantly increased the number of neutrophils and CD68+macrophages in both liver and spleen tissues of PbANKA-infected mice.2.Ts co-infection significantly increased the mRNA expression level of inflammatory cytokines in the liver and spleen tissues of PbANKA-infected mice.3.Ts co-infection resulted in significantly increased mRNA expression levels of CD200 and CD200R in the spleen of PbANKA-infected mice.4.Ts co-infection resulted in significantly increased mRNA expression levels of M2-type macrophage markers[mannose receptor,C type 2(Mrc-2)and chitinase-like 3(Yml)]in small intestinal tissue of PbANKA infected mice.5.In vitro experiments showed that Ts co-infection resulted in significantly increased mRNA expression levels of Gal-1 and Gal-3 as well as M1-type polarization marker in peritoneal macrophages of PbANKA-infected mice.Conclusions:1.Ts co-infection did not affect the survival rate of PbANKA-infected mice,but significantly reduced the parasitemia in PbANKA-infected mice and parasite burden in liver tissue,and significantly aggravated the liver immunopathological injury of PbANKA-infected mice.2.Further studies found that Ts co-infection significantly increased the expression of neutrophil elastase and CD68+macrophages in the liver and spleen of PbANKA infected mice,and significantly lincreased the expression of Gal-1 and Gal-3 and pro-inflammatory factors in the liver and spleen.This suggests that Ts co-infection can enhance the immune response of PbANKA infected mice against plasmodium,kill and eliminate plasmodium by activating immune cells such as macrophages and neutrophils,and produce a large number of inflammatory factors to aggravate the immune damage to host organs.3.Ts co-infection can regulate the immune response of PbANKA infected mice,and promote M2-type polarization of macrophages in the small intestine.4.This study explored the effect of Ts co-infection on malaria outcomes,and the results are of great significance in understanding the pathogenesis of malaria in areas where helminth and malaria are common.