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Study On The Regulation Mechanism Of Hematopoiesis By Ly6C~+ Monocyte Progenitor Cells In Bone Marrow

Posted on:2019-02-27Degree:MasterType:Thesis
Country:ChinaCandidate:J M GaoFull Text:PDF
GTID:2354330548457622Subject:Cell biology
Abstract/Summary:PDF Full Text Request
Hematopoietic stem cells are the progenitor cells of various blood cells and have the important physiological function of continuing and rebuilding hematopoiesis.Niche as a suitable microenvironment for the survival of HSCs,through the adhesion molecules and membrane surface signaling molecules in direct contact with HSCs or through the secretion of cytokines and other stimuli in HSCs signal activation,thereby regulating HSCs self-renewal,proliferation,differentiation and survival of cells Biological behavior.The phenotype of granulocytes in mouse bone marrow is CD11b~+Gr-1~+and is divided into two subtypes:granulocyte-like(CD11b~+Ly6C~-Ly6G~+)and monocyte-like(CD11b~+Ly6C~+Ly6G~+).The regulation of HSCs by granulocytes in bone marrow has not been reported at present.This paper first explored the specific mechanism of Ly6C~+granulocytes in the bone marrow to regulate the proliferation and differentiation of HSCs.The article is divided into three parts.(1)Study the effects of Ly6C~+granulocytes on hematopoietic reconstitution in bone marrow;(2)Study the mechanism of action of Ly6C~+granulocytes on hematopoietic regulation in bone marrow;(3)Study the expansion of HSCs by specific expression factors in Ly6C~+granulocytes of bone marrow influences.During this period,we used in vivo antibody removal,flow sorting,immunofluorescence staining,and PCRArray to complete research tasks.It was found that after Ly6C~+granulocytes in bone marrow were cleared by antibodies,the hematopoietic reconstitution capacity of mice was significantly enhanced,indicating that Ly6C~+granulocytes inhibited mouse HSCs;next we used immunofluorescence staining to find Ly6C~+granulocytes and HSCs.The positional relationship of the seed cells was marked with different color antibodies.The experiments showed that Ly6C~+granulocytes and HSCs cells in the bone marrow were closely adjacent to each other,suggesting that Ly6C~+granulocytes themselves or their secretory factors may have a certain regulatory effect on hematopoietic stem cells;To further explore which factors in Ly6C~+granulocytes play a key role in the regulation of HSCs,we used PCR Arrays to screen out specific cytokines expressed in Ly6C~+granulocytes.Among the many factors,we studied the in vitro culture of B cells based on previous studies.We found that Tnfsf12 and Tnfsf13,two kinds of cytokines that we would like to study further,added Tnfsf12 and Tnfsf13 to the hematopoietic stem cell system in vitro.After six days of culture,we analyzed the total and absolute number of hematopoietic stem cells by flow cytometry,The results showed that the hematopoietic stem cells were differentiated to a certain extent in the cell group to which these factors were added compared to the control group,indicating that these factors may have the effect of promoting the differentiation of hematopoietic stem cells.This study provides a new idea for revealing the regulatory mechanism of Niche cells on HSCs.It was found for the first time that Ly6C~+granulocytes in bone marrow inhibit the self-renewal ability of HSCs by secreting Tnfsf12 and Tnfsf13.In the later period,we will continue to study the regulatory role of Ly6C~+granulocytes in HSCs and provide information for further revealing the cellular components in the bone marrow microenvironment.
Keywords/Search Tags:hematopoietic stem cells, Ly6C~+, flow cytometry, PCRArray, immunofluorescence staining, in vitro culture
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