Human Bone Marrow Microenvironment in Health and Disease

Abstract: Hematopoietic stem cells (HSCs) are safeguarded from various threats such as stress, injury, or radiationwithin specialized microenvironments or niches within the bone marrow. In this thesis, we investigatedvarious facets of the bone marrow (BM) microenvironment and its critical role in hematopoiesis andrelated disorders.Initially, we elucidated the role of the Early Growth Response 1 (EGR1) gene in bone marrowmesenchymal stem cells (MSCs). EGR1 expression was found to be significantly elevated in specific BMMSC populations, particularly lin-CD45-CD271+ CD140a- BM MSCs, exerting a pivotal role inhematopoietic stroma support. This support function was mediated through both cell-cell interactions andsoluble factors, where EGR1-overexpressing BM MSCs exhibited enhanced secretion of chemokineligand 28 and increased expression of vascular cell adhesion molecule 1, crucial for hematopoiesissupport. EGR1 also played a dual role in BM MSC proliferation regulation. Understanding thesemechanisms can improve hematopoietic stem cell transplantation and regenerative medicine.Further, employing single-cell RNA sequencing, we provided a comprehensive analysis of the cellularcomposition of human BM stroma, revealing diverse cell populations and stromal progenitors with varyingdifferentiation capacities. We identified and characterized multipotent stromal stem cells (MSSCs), highlyadipocytic gene-expressing progenitors (HAGEPs), pre-osteoblasts, and other stromal clusters, offeringvaluable insights into BM stromal heterogeneity and its structural organization. We employed in silicocluster interaction analysis and found that the different stromal populations are predicted to interact andsupport HSCs differentially in different niches.Next, we introduced a meticulous methodology utilizing multicolor immunofluorescence staining and 3Danalysis to investigate human BM architecture. We illustrated the potential of sequential staining,emphasizing specific structural changes associated with myeloproliferative neoplasms (MPNs) and theircorrelation with CD271 expression.Finally, we investigated human BM architecture and cytokine expression patterns in patients with AcuteLymphoblastic Leukemia (ALL) and Primary Myelofib(PMF) with elaborate immunofluorescence andmRNA-based staining methods and compared them to healthy controls. We revealed significantalterations, including differential numbers of megakaryocytes, differences in cellularity, alteredmesenchymal stem cell density, and distinct cytokine expressions offering critical insights into diseasepathogenesis and progression.Collectively, these findings illuminate various facets of the BM microenvironment, offering valuableinsights into its critical role in hematopoiesis, stromal heterogeneity, and disease pathology, paving theway for potential therapeutic advancements in regenerative medicine and hematological disorders.