Genetic Engineering Mouse Dp(16)1Yey/mir-155-Was Builded And Used To Study The Effect Of Mir-155 Gene Dose On The Proliferation And Differentiation Of Hematopoietic Cells In DS Mice And Related TPO Signaling Pathway

Aim: Down syndrome,also known as Down syndrome,is a disease caused by an unusually more than one chromosomes in chromosome 21.This study was designed to build a mouse model,Dp(16)1Yey/ mir-155-,which is used to study the effect of mir-155 gene dose on the proliferation and differentiation of hematopoietic cells in DS mice and related TPO signaling pathway.Methods: mi R155 knockout mice(mi R155 + /-)mice and DS(Dp16 / +)was mated to acquire contains two copies mi R155 DS mice(Dp16 / mi R155-)and contains three copies mi R155 DS mice(Dp16 / +),which brood offspring mice also contains wild-type mice(+ / +).First,the abnormal peripheral blood of three genotype mice was analyzed.The spleen weight of different genotype mice was compared,and the blood distribution was observed in the spleen,liver and bone marrow tissue.In order to further validate hematopoietic cell distribution in cytology,the spleen and bone marrow hematopoietic cell of different genotype mice were analyzed by flow cytometry.Using flow cytometry analysis of the different genotypes of 15 months mice bone marrow hematopoietic stem cells(Sca1 + lineage-,c-Kit +)(LSK)and myeloid progenitor cells(Sca1,lineage--,c-Kit +)quantity,used to determine whether the change of peripheral blood and spleen and bone marrow cytology associated with changes of hematopoietic stem cells or progenitor cells.The expression of STAT protein in the hematopoietic system of different genotype mice was analyzed using Westernblot method.Result: These three genotypes mice were successfully constructed.These three genotype 3 months to 15 months of age mice peripheral blood complete blood count results show that compared with wild type mice,Dp16 / + mice since 3 months,the number of peripheral blood red blood cell(RBC)began to reduce;Since the age of 9 months,the hemoglobin concentration(HGB)has decreased.Dp16/mi R155-mice was almost identical to the wildtype mice.In addition,the peripheral blood count results also show that Dp16 / + mice since 3 months average red blood cell volume(MCV)and average red blood cell hemoglobin(MCH)content significantly higher than the wild type controls,and Dp16 / mi R155-no significant difference with wild type mice.Since the age of 6 months,the number of platelets(PLT)and platelet volume(MPV)in Dp16/+ mice increased significantly,while Dp16/mi R155-was not significantly different from that in wild-type mice.The above results indicated that peripheral blood abnormality of Dp16/+ mice may be related to mi R155 level.In 16-24 months age mice,Compared with the wild-type control mice(104.07 ± 25.34 mg,n = 6),the spleen weight of Dp16/+ mice(225.67 ± 38.41 mg,n = 6)increased significantly.Compared with the wild-type mice,Dp16/mi R155-(142.55 + + 39.28 mg,n = 6)was not very different from the wild-type mice.The results showed that the separation between the white and red pulp of wild-type mice was obvious,and the difference between Dp16/+ mice and Dp16/mi R155-was not obvious.The results showed that the hepatic cord around the central vein of the liver tissue of the wild-type mice was obvious,and the structure was not obvious in Dp16/mi R155-and Dp16/+ mice.In Dp16/mi R155-and Dp16/+ mice,CD41+ megakinocytes were significantly higher than those in the wild-type control mice.It was also found that the number of Ter119+ red cells decreased significantly in Dp16/mi R155-and Dp16/+ mice.No differences were found between Dp16/mi R155-and Dp16/+ mice.Compared with the wild-type control mice,the number of Dp16/mi R155-and Dp16/+ mouse hematopoietic stem cells(LSK)increased and the number of myeloid progenitor cells(Sca1-,c-kit +)decreased,and there was no significant difference between Dp16/mi R155-and Dp16/+ mice.In myeloid progenitor cells,although common myeloid progenitor cells(CMP)(Sca1,lineage--,c-Kit + andCD34 +,Fc gamma R-)number have no significantly difference among genotypes in mice,the Megakaryocyte Erythroid precursors(MEP)(Sca1,lineage--,c-Kit + and CD34-,Fc gamma R-)quantity in Dp16 / mi R155-and Dp16 / + mice was increased,and the Granulopoietic Mononuclea progenitors(GMP)(Sca1,lineage--,c-Kit + and CD34 +,Fc gamma R +)quantity in Dp16 / mi R155-and Dp16 / + mice was reduced,Dp16 / mi R155 – and Dp16 / + mice had no significant difference.The expression of STAT3 protein was highest in DS mice Dp16/+,and there was no significant difference in wild type and Dp16/mi R155-mice.Conclusion: Genetic engineering mouse Dp(16)1Yey/ mir-155-was successfully constructed.In Dp16/mi R155-and Dp16/+ mice,mir-155 gene abnormality resulted in the increase of hematopoietic stem cells in bone marrow,and in the medullary system,the Megakaryocyte Erythroid precursors is increased,and the Granulopoietic Mononuclea progenitors is decreased.The number of CD41+ megakaryocytes in the bone marrow was significantly higher,and the number of Ter119+ red cells decreased significantly,Spleen and hepatic cord structure,bone marrow tissue structure disorder,These evidences indicate that the increase or abnormality of mir-155 gene leads to the pathogenesis of acute macronuclear leukemia(ds-amkl)in mice with down syndrome.The spleen and liver were infiltrated with leukemia.At the same time,the number of red blood cells in the peripheral blood of the three mir-155 genes was decreased and the volume increased,Giant cell changes,The number of platelets increased and the volume increased,There are also giant cell changes.Erythrocytes and platelets have a teratogenic reaction.There is an enlargement of the spleen(associated with leukemia infiltration).The expression of STAT3 protein in DS mouse Dp16/+ was the highest,and the abnormal expression of mir-155 resulted in the excessive expression of protein STAT3 in TPO signaling pathway,which was the important cause of abnormal peripheral blood.