The Loss Of Chemokine Receptor ? Affects The Hematopoietic System And Hematopoietic Reconstitution

Hematopoietic stem cells(HSCs)are the origin cells of all cell lines in the blood system of the body.These cells have the ability of self-renewal,proliferation and continuous differentiation,and are responsible for maintaining the dynamic balance of the blood system in the body and rebuilding under the stress response of the body.Important physiological functions such as the hematopoietic system.The bone marrow microenvironment(Niche)of hematopoietic stem cells is located around the vascular sinus near the trabecular bone area.It is a series of special cell groups composed of bone marrow mesenchymal stromal cells(MSC)and endothelial cells and is responsible for maintaining and regulating the physiological functions of hematopoietic stem cells.A special microenvironment.In the bone marrow microenvironment,its constituent cells directly or indirectly contact with hematopoietic stem cells through cell adhesion molecules and cell membrane surface signal molecules to regulate a series of biological functions such as self-renewal,proliferation,and differentiation of hematopoietic stem cells.Under normal circumstances,CXCR3 can induce chemotaxis and proliferation of different types of cells,and can also inhibit migration and proliferation and induce apoptosis.In some cases,it may be beneficial for tumor growth and progression.G-CSF can specifically regulate the differentiation,proliferation and survival of granulocyte progenitor cells and downstream cells after binding to receptors in vivo.It is also related to the secretion of vascular endothelial cells and enhances the formation of blood vessels,and it can mobilize hematopoietic stem cells and mature granulocytes in the bone marrow to peripheral blood.This thesis includes four parts:(1)CXCR3,GCSFR single gene knockout and G-CSFR & CXCR3 double gene knockout mice hematopoietic cell phenotype detection;(2)CXCR3 gene knockout mice cultured in vitro to isolate C-kit positive cells to observe the effect of gene defects on the differentiation of stem cells invitro;(3)CXCR3 gene knockout mice and G-CSFR & CXCR3 double knockout mice in vivo full bone marrow competitive transplantation and hematopoietic reconstruction ability under body stress response(4)The deletion of CXCR3 gene affects the molecular mechanism of hematopoietic stem / progenitor cell division and differentiation.The research results are as follows:1: We first tested the proportion and number of hematopoietic cells in the peripheral blood and bone marrow of CXCR3 gene knockout mice,G-CSFR gene knockout mice and CXCR3,G-CSFR double knockout mice,and found that: CXCR3 gene The loss of hematopoietic stem cells resulted in fewer cells dividing than ordinary wild-type mice,and it was found that the number of B cells was significantly reduced during the differentiation of the blood system downstream lineage,which means that the deletion of the CXCR3 gene is.To a certain extent,it inhibited the ability of hematopoietic stem cells to self-proliferate and differentiate into mature B cells.The defect of GCSFR gene led to a significant decrease in the number of granulocytes andT cells differentiated by their downstream lineages.However,when both were knocked out,it was found that the granulocytes,T cells or B cells of the downstream lineage,as well as the upstream hematopoietic stem and progenitor cells,had returned to levels similar to those of wild-type mice.2: C-ki + cells isolated from CXCR3 gene knockout mice were cultured in vitro,and it was found on the tenth day of culture that the number of C-kit + cells of CXCR3 gene knockout mice was much more than normal wild type,and B cells were cultured after ten days.The number is still significantly less than normal wild-type cells.It further illustrates the ability of the CXCR3 gene to inhibit stem cell proliferation and differentiate into mature B cells.3: We competitively transplanted whole bone marrow cells of CXCR3 gene knockout mice into recipient CD45.1 mice,and found that the chimerism rate,B lymphocyte,The ratio of T lymphocytes and Gr-1 + CD11 b + granulocytes islower than that of normal wild group.What about hematopoietic stem / progenitor cells?After treatment of CXCR3 knockout mice and normal wild-type mice with5-fluorouracil,bone marrow cells were collected on days 3,5,7,8,9,and 10 and tested for their hematopoietic recovery function.Among them,B cell grouping was obvious.Compared with wild-type mice,the differentiation ability of CXCR3 knockout mice B cells was significantly reduced.The number of hematopoietic stem and progenitor cells in CXCR3 knockout mice was lower than that of normal wild-type mice.Changes in other cells during hematopoietic reconstruction.4: Finally,we sorted out the hematopoietic stem cells of the CXCR3 geneknockout mice,and extracted and traced the transcriptome level using a small amount of cellular RNA.It was found that the developmental Gpr55 and Ubqln3 genes in CXCR3 gene knockout mice were up-regulated,and genes such as Gbx2 and Emilin2 have been down-regulated.This study is the first to explore the role of CXCR3 and G-CSFR genes in the process of normal hematopoiesis,revealing the impact of CXCR3 gene on the growth and development of hematopoietic stem cells and their differentiation.Later,the gene defects will be explored at the transcriptome level.The impact will provide information for further revealing how CXCR3 regulates the function of hematopoietic stem cells.