Project description:How cells acquire their fate is a fundamental question in both developmental and regenerative biology. Multipotent progenitors undergo gradual cell fate restriction in response to temporal and positional cues from the microenvironment, the nature of which is far from being clear. In the case of the lymphatic system, venous endothelial cells are thought to give rise to lymphatic vessels, through a process of trans-differentiation. Upon expression of a set of transcription factors, venous cells acquire a lymphatic fate, and bud out to generate the lymphatic vasculature. In this work we challenge this view and show that while lymphatic endothelial cells (LECs) do arise in the Cardinal Vein (CV), they do so from a previously uncharacterized pool of multipotent angioblasts. Using lymphatic-specific transgenic zebrafish, in combination with endothelial photoconvertible reporters, and long-term live imaging, we demonstrate that these multipotent angioblasts can generate not only lymphatic, but also arterious, and venous fates. We further reveal that the underlying endoderm serves as a source of Wnt5b, which acts as a lymphatic inductive signal, promoting the angioblast-to-lymphatic transition. Moreover, Wnt5b induced lymphatic specification in human embryonic stem cells- derived vascular progenitors, suggesting that this process is evolutionary conserved. Our results uncover a novel mechanism of lymphatic vessel formation, whereby multipotent angioblasts and not venous endothelial cells give rise to the lymphatic endothelium, and provide the first characterization of their inductive niche. More broadly, our findings highlight the CV as a plastic and heterogeneous structure containing different cell populations, analogous to the hematopoietic niche in the aortic floor. Following Kaede photoconversion of dorsal or ventral halves of the PCV in Tg(fli1:gal4;uasKaede) embryos at 24 hpf, 6 embryos per group were used for FACS isolation of Kaede photconverted (red) ECs.
Project description:How cells acquire their fate is a fundamental question in both developmental and regenerative biology. Multipotent progenitors undergo gradual cell fate restriction in response to temporal and positional cues from the microenvironment, the nature of which is far from being clear. In the case of the lymphatic system, venous endothelial cells are thought to give rise to lymphatic vessels, through a process of trans-differentiation. Upon expression of a set of transcription factors, venous cells acquire a lymphatic fate, and bud out to generate the lymphatic vasculature. In this work we challenge this view and show that while lymphatic endothelial cells (LECs) do arise in the Cardinal Vein (CV), they do so from a previously uncharacterized pool of multipotent angioblasts. Using lymphatic-specific transgenic zebrafish, in combination with endothelial photoconvertible reporters, and long-term live imaging, we demonstrate that these multipotent angioblasts can generate not only lymphatic, but also arterious, and venous fates. We further reveal that the underlying endoderm serves as a source of Wnt5b, which acts as a lymphatic inductive signal, promoting the angioblast-to-lymphatic transition. Moreover, Wnt5b induced lymphatic specification in human embryonic stem cells- derived vascular progenitors, suggesting that this process is evolutionary conserved. Our results uncover a novel mechanism of lymphatic vessel formation, whereby multipotent angioblasts and not venous endothelial cells give rise to the lymphatic endothelium, and provide the first characterization of their inductive niche. More broadly, our findings highlight the CV as a plastic and heterogeneous structure containing different cell populations, analogous to the hematopoietic niche in the aortic floor.
Project description:The goal and objective of this study was to identify the transcriptional profiles differentiating the artery, vein, and lymphatic lineages in the adult rat vasculature with particular emphasis on the unique elements of the collecting lymphatic vessel transcriptome. A 2 x 3 experimental design was utilized in which parallel arteries, veins, and lymphatics from two different tissue beds were examined. The rat thoracic duct was selected as a large, post-nodal collecting lymphatic vessel that exhibits excellent conduit-type behavior while the rat mesenteric lymphatic was selected as a smaller, pre-nodal collecting lymphatic vessel that exhibits excellent pump behavior (see Gashev AA, et al. Microcirculation. 2004 Sep;11(6):477-92. [PMID: 15371129]). The axillary artery and vein were selected for comparison to the thoracic duct due to their similar anatomical position distal to the common junction of the lymphatic and venous vascular trees and represent a large artery and large vein, respectively. The mesentery artery and vein were selected for comparison to the mesenteric lymphatic vessels due to their parallel position within the mesenteric vasculature and represent a small atery and small vein, respectively. A 2 x 3, reference-based, experimental design was utilized consisting of both large (thoracic) and small (mesenteric) arteries, veins, and collecting lymphatic vessels for a total of 6 sample groups with n=6 biological replicates present in each group. All vessels acquired from the same donor animal have the same numerical label and were handled in parallel through all experimental steps. Each vessel sample RNA sample was amplified, labeled with Cy5, and compared to the same Rat Universal Reference RNA sample (Stratagene, La Jolla, CA) that was amplified and labeled with Cy3 dye. No dye swaps were utilized.
Project description:The goal and objective of this study was to identify the transcriptional profiles differentiating the artery, vein, and lymphatic lineages in the adult rat vasculature with particular emphasis on the unique elements of the collecting lymphatic vessel transcriptome. A 2 x 3 experimental design was utilized in which parallel arteries, veins, and lymphatics from two different tissue beds were examined. The rat thoracic duct was selected as a large, post-nodal collecting lymphatic vessel that exhibits excellent conduit-type behavior while the rat mesenteric lymphatic was selected as a smaller, pre-nodal collecting lymphatic vessel that exhibits excellent pump behavior (see Gashev AA, et al. Microcirculation. 2004 Sep;11(6):477-92. [PMID: 15371129]). The axillary artery and vein were selected for comparison to the thoracic duct due to their similar anatomical position distal to the common junction of the lymphatic and venous vascular trees and represent a large artery and large vein, respectively. The mesentery artery and vein were selected for comparison to the mesenteric lymphatic vessels due to their parallel position within the mesenteric vasculature and represent a small atery and small vein, respectively.
Project description:Whereas the cellular basis of the hematopoietic stem cell (HSC) niche in the bone marrow has been characterized, the nature of the fetal liver (FL) niche is not yet elucidated. We show that Nestin+NG2+ pericytes associate with portal vessels, forming a niche promoting HSC expansion. Nestin+NG2+ cells and HSCs scale during development with the fractal branching patterns of portal vessels, tributaries of the umbilical vein. After closure of the umbilical inlet at birth, portal vessels undergo a transition from Neuropilin-1+Ephrin-B2+ artery to EphB4+ vein phenotype, associated with a loss of peri-portal Nestin+NG2+ cells and emigration of HSCs away from portal vessels. These data support a model in which HSCs are titrated against a peri-portal vascular niche with a fractal-like organization enabled by placental circulation. Characterization of the transcriptome of fetal liver and adult bone marrow niche using RNA-seq
Project description:Whereas the cellular basis of the hematopoietic stem cell (HSC) niche in the bone marrow has been characterized, the nature of the fetal liver (FL) niche is not yet elucidated. We show that Nestin+NG2+ pericytes associate with portal vessels, forming a niche promoting HSC expansion. Nestin+NG2+ cells and HSCs scale during development with the fractal branching patterns of portal vessels, tributaries of the umbilical vein. After closure of the umbilical inlet at birth, portal vessels undergo a transition from Neuropilin-1+Ephrin-B2+ artery to EphB4+ vein phenotype, associated with a loss of peri-portal Nestin+NG2+ cells and emigration of HSCs away from portal vessels. These data support a model in which HSCs are titrated against a peri-portal vascular niche with a fractal-like organization enabled by placental circulation.