Effect Of Overexpression Of CD40L Gene On Extramedullary Heart And Kidney Infiltration In Nude Mice Leukemia Model

Objective:In this study,we detected the expression of CD13,MMP-2 and VEGF in the heart and kidney of extramedullary organs of NOD-SCID mice with CD40L gene overexpression constructed by THP-1 cells,and to explore the effect of CD40L overexpression on extramedullary heart and kidney infiltration of acute myeloid leukemia.Methods:1)NOD-SCID mice were injected intravenously with THP-1 cells,1×10~7THP-1 cells transfected with empty vector and 1×10~7 THP-1 cells transfected with CD40L gene to establish the NOD-SCID mouse model of acute myeloid leukemia.The stable THP-1 cell lines expressing CD40L have been successfully screened in this research group.The mice were divided into three groups:THP-1 group,empty vector transfection THP-1 group and CD40L transfection THP-1 group,with 8 mice in each group.2)Blood routine,peripheral blood smear and bone marrow morphology of mice in each group were detected.3)Tumor-bearing mice were selected to peel off their hearts and kidneys,fix them with 4%formaldehyde,paraffin-embed and slice them,and perform HE staining to observe the extramedullary heart and kidney infiltration in mice with acute myeloid leukemia;4)Immunohistochemistry was used to detect the expression of CD13,MMP2and VEGF in heart and kidney of mice.Results:(1)Stable THP-1 cell line overexpressing CD40L:stable fluorescent expression of green fluorescent protein was observed under microscope,CD40L overexpression in THP-1 cell line was verified by PCR method in this experimental group.(2)THP-1 group,idling group and CD40L group showed leukocytosis,hemoglobin and thrombocytopenia in peripheral blood before and after 4 weeks of modeling,and there was no obvious trend change among the three groups.(3)THP-1 cells were found in peripheral blood smears and bone marrow smears of 7 mice in THP-1 group,6 mice in idle group and 6 mice in CD40L group.(4)HE staining results of heart and kidney tissue sections of mice in three groups:there were 4 mice in THP-1 group with heart infiltration and kidney infiltration,2 mice with heart infiltration and 6 mice with kidney infiltration in idling group,and 3 mice with heart and kidney infiltration in CD40L group.(5)the expression of CD13,MMP-2 and VEGF in heart and kidney of three kinds of mice was detected by immunohistochemical method:CD13 was positive in heart and kidney of all mice,and the expression of MMP-2in heart and kidney of CD40L group was lower than that of idle group and THP-1group,the difference was statistically significant(p<0.05).The expression of VEGF in CD40L group was lower than that in idling group and THP-1 group,the difference was statistically significant(p<0.05),and the expression in kidney was also lower than that of the other two groups,but the difference was not statistically significant(p>0.05).Conclusion:(1)After modeling the mouse acute myeloid leukemia model,peripheral blood WBC increased,HB and PLT decreased;(2)HE staining and CD13 expression of extramedullary heart and kidney tissue in acute myeloid leukemia mice were found.Acute myeloid leukemia was prone to extramedullary heart and kidney infiltration;(3)The expressiont of MMP-2 and VEGF in the extramedullary heart and kidney tissue of leukemia mice overexpressing CD40L gene was lower than that of leukemia mice not overexpressing CD40L gene,suggesting that overexpression of CD40L gene may affect the extramedullary of leukemia mice the degree of heart and kidney tissue infiltration,the specific mechanism needs to be further studied.