Publicação
The role of pericytes in the generation hematopoietic stem cells in the mouse embryo
| Resumo: | Hematopoiesis is a complex process that gives rise to hematopoietic stem cells (HSCs) and progenitors responsible for the production of all blood cells in the organism. This process is defined by two major waves: primitive and definitive haematopoiesis. In the primitive haematopoiesis, the yolk sac gives rise to erythrocytes, macrophages and megakaryocytes during early embryonic development. This wave is transitory and contribute to the survival of the early embryo. In the definitive haematopoiesis that occurs in the mid-gestation of mouse embryos hematopoietic stem cells are generated from hemogenic endothelial cells found in the aortic wall. These are adult-type HSCs, that are multilineage, are able to reconstitute hematopoietic system of both primary and secondary host irradiated mice upon transplantation in vivo. It is unclear how these HSCs born in the aorta. The microenvironment surrounding the dorsal aorta was shown to influence their birth. However, what are the exact cell types involved in this process remains unclear. We here found that the dorsal aorta is surrounded by three layers of perivascular cells, including pericytes, that are phenotypically and genetically distinct. Recent work showed that pericytes support adult HSCs but whether they also support their birth is not known. The proximal layers express PDGFRβ, known to be involved in pericyte recruitment and to control smooth muscle cell proliferation. Our general hypothesis is that PDGFRβ is required to generate HSCs in the mouse embryo. To address this, we used the PDGFRβ knock-out mouse model. The deletion of this receptor in mice leads to a reduced number of pericytes and vascular smooth muscle cells in the developing embryo following by death before birth. In this study, we performed in vitro and in vivo hematopoietic assays, as well as flow cytometry and immunohistochemistry to characterise wild-type and PDGFRβ mutant embryos at the time of HSC generation. Our preliminary data indicate that both hematopoietic progenitor and stem cell activities are impaired in mutant embryos. Blood vessel integrity is not affected at this stage. However, we found that the percentage of hemogenic endothelial cells, which are HSC precursors significantly decrease in the mutant embryos. Altogether, my data demonstrate that PDGFRβ signalling is required to generate the first HSCs in the mouse embryo. This study is of high importance for the understanding of how hematopoietic cells are generated in vivo and for the identification of the key cells in its surrounding environment responsible for its production, so that this process can be transposed into in vitro assays for its production. |
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| Autores principais: | Ventura, Telma Filipa da Cruz |
| Assunto: | PDGFRβ Célula estaminal hematopoiética Célula endothelial hemogénica Nicho Embrião de ratinho Teses de mestrado - 2018 |
| Ano: | 2018 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório da Universidade de Lisboa |
| Resumo: | Hematopoiesis is a complex process that gives rise to hematopoietic stem cells (HSCs) and progenitors responsible for the production of all blood cells in the organism. This process is defined by two major waves: primitive and definitive haematopoiesis. In the primitive haematopoiesis, the yolk sac gives rise to erythrocytes, macrophages and megakaryocytes during early embryonic development. This wave is transitory and contribute to the survival of the early embryo. In the definitive haematopoiesis that occurs in the mid-gestation of mouse embryos hematopoietic stem cells are generated from hemogenic endothelial cells found in the aortic wall. These are adult-type HSCs, that are multilineage, are able to reconstitute hematopoietic system of both primary and secondary host irradiated mice upon transplantation in vivo. It is unclear how these HSCs born in the aorta. The microenvironment surrounding the dorsal aorta was shown to influence their birth. However, what are the exact cell types involved in this process remains unclear. We here found that the dorsal aorta is surrounded by three layers of perivascular cells, including pericytes, that are phenotypically and genetically distinct. Recent work showed that pericytes support adult HSCs but whether they also support their birth is not known. The proximal layers express PDGFRβ, known to be involved in pericyte recruitment and to control smooth muscle cell proliferation. Our general hypothesis is that PDGFRβ is required to generate HSCs in the mouse embryo. To address this, we used the PDGFRβ knock-out mouse model. The deletion of this receptor in mice leads to a reduced number of pericytes and vascular smooth muscle cells in the developing embryo following by death before birth. In this study, we performed in vitro and in vivo hematopoietic assays, as well as flow cytometry and immunohistochemistry to characterise wild-type and PDGFRβ mutant embryos at the time of HSC generation. Our preliminary data indicate that both hematopoietic progenitor and stem cell activities are impaired in mutant embryos. Blood vessel integrity is not affected at this stage. However, we found that the percentage of hemogenic endothelial cells, which are HSC precursors significantly decrease in the mutant embryos. Altogether, my data demonstrate that PDGFRβ signalling is required to generate the first HSCs in the mouse embryo. This study is of high importance for the understanding of how hematopoietic cells are generated in vivo and for the identification of the key cells in its surrounding environment responsible for its production, so that this process can be transposed into in vitro assays for its production. |
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