The Heterogeneity Of Hemocytes In Oncomelania Hupensis And The Role Against Schistosoma Japonicum Infection

Objective The hemocytes of Oncomelania hupensis is the main effector of immune defense against Schistosoma japonicum.The morphological and functional characterization of hemocytes remains unclear.In this study,the basic morphologic and classification were deeply investigated.The dynamic change of hemocytes after S.japonicum challenge was evaluated,including composition of subpopulations,the phagocytosis and apoptosis,and the expression profiles of NOX(NADPH oxidase)and IL 17(Interleukin 17)m RNA in hemocytes,to understand the role of hemocytes against S.japonicum infection.Methods The hemocytes were collected from 0 h to 8 d after S.japonicum infection.The morphology of normal haemocytes was observed under Giemsa staining and electronic microscope.The dynamic change in amount of haemocytes and subpopulations were evaluated after S.japonicum challenge by Giemsa staining and flow cytometry.In addition,the ability of hemocytes in phagocytosis by devouring the latex beads and apoptosis was checked by flow cytometry.The dynamic expression of NOX and IL 17 in hemocytes were estimated through real-time PCR.Results The hemocytes in Oncomelania hupensis were divided into granulocytes and hyalinocytes according to existence of granules,accounted for 6.93±0.40% and91.74±0.41% of total hemocytes,respectively.The granulocytes could be further divided into basophilic and eosinophilic granulocytes,accounted for 2.86±0.12% and4.10±0.29% of total hemocytes,respectively.While the hyalinocytes were divided into small hyalinocytes(SH),medium hyalinocytes(MH),and large hyalinocytes(LH)based on the size,accounted for 87.83±0.88%,4.65±1.13%,and 0.26±0.04% of total hemocytes,respectively.The total hemocytes count(THC)increased dramatically at 6h after challenge,and the granulocytes and large hyalinocytes increased most,accounted for 0.29% and 11.59% of total hemocytes at 6 h post-infection.The percentage of hemocytes in phagocytosis and apoptosis increased to highest at 6 h after challenge,accounted for 60.7% and 68.1% of total hemocytes,respectively.The expression of NOX-1 m RNA in hemocytes increased after challenge,and up to highest at 48 h post-infection and decreased to lowest at 8 d post-infection.The expression of NOX-2 increased to highest at 24 h post-infection and then decreased to lowest at 4 d post-infection followed by higher expression at 8 d post-infection.The expression of IL 17-1 increased to highest at 12 h post-infection and decreased to lowest at 4 d postinfection,but increased to higher at 8 d post-infection.The expression of IL 17-2continuously increased after challenge,and the expression was 60.88 times higher at 8d post-infection than 0 h.Conclusion The hemocytes in O.hupensis show the heterogeneity in morphology and function.The hemocytes can be divided into basophilic granulocytes,eosinophilic granulocytes,small hyalinocytes(SH),medium hyalinocytes(MH)and large hyalinocytes(LH)based on the granules and the size.Among them,the granulocytes and LH play key roles against S.japonicum infection.Phagocytosis and apoptosis may help hemocytes defense against S.japonicum,especially at the early 12 h after infection.The expression of NOX and IL 17 m RNA increased after infection and reached to the highest level at different times.The NOX and IL 17 related signal pathways were thought to play roles to adjust the ability of hemocytes in phagocytosis and apoptosis,to help snails defense against S.japonicum.