Matrix stiffness controls lymphatic vessel formation through regulation of a GATA2-dependent transcriptional program

Tissue and vessel wall stiffening alters endothelial cell properties and contributes to vascular dysfunction. However, whether extracellular matrix (ECM) stiffness impacts vascular development is not known. Here we show that matrix stiffness controls lymphatic vascular morphogenesis. Atomic force microscopy measurements in mouse embryos reveal that venous lymphatic endothelial cell (LEC) progenitors experience a decrease in substrate stiffness upon migration out of the cardinal vein, which induces a GATA2-dependent transcriptional program required to form the first lymphatic vessels. Transcriptome analysis shows that LECs grown on a soft matrix exhibit increased GATA2 expression and a GATA2-dependent upregulation of genes involved in cell migration and lymphangiogenesis, including VEGFR3. Analyses of mouse models demonstrate a cell-autonomous function of GATA2 in regulating LEC responsiveness to VEGF-C and in controlling LEC migration and sprouting in vivo. Our study thus uncovers a mechanism by which ECM stiffness dictates the migratory behavior of LECs during early lymphatic development.Peer reviewe

Volumetric imaging reveals VEGF-C-dependent formation of hepatic lymph vessels in mice

International audienceThe liver is a major biosynthetic and detoxifying organ in vertebrates, but also generates 25%-50% of the lymph passing through the thoracic duct and is thereby the organ with the highest contribution to lymph flow. In contrast to its metabolic function, the role of the liver for lymph generation and composition is presently severely understudied. We took a rigorous, volume imaging-based approach to describe the microarchitecture and spatial composition of the hepatic lymphatic vasculature with cellular resolution in whole mount immune stained specimen ranging from thick sections up to entire mouse liver lobes. Here, we describe that in healthy adult livers, lymphatic vessels were exclusively located within the portal tracts, where they formed a unique, highly ramified tree. Ragged, spiky initials enmeshed the portal veins along their entire length and communicated with long lymphatic vessels that followed the path of the portal vein in close association with bile ducts. Together these lymphatic vessels formed a uniquely shaped vascular bed with a delicate architecture highly adapted to the histological structure of the liver. Unexpectedly, with the exception of short collector stretches at the porta hepatis, which we identified as exit point of the liver lymph vessels, the entire hepatic lymph vessel system was comprised of capillary lymphatic endothelial cells only. Functional experiments confirmed the space of Disse as the origin of the hepatic lymph and flow via the space of Mall to the portal lymph capillaries. After entry into the lymphatic initials, the lymph drained retrograde to the portal blood flow towards the exit at the liver hilum. Perinatally, the liver undergoes complex changes transforming from the main hematopoietic to the largest metabolic organ. We investigated the time course of lymphatic vessel development and identified the hepatic lymphatics to emerge postnatally in a process that relies on input from the VEGF-C/VERGFR-3 growth factor-receptor pair for formation of the fully articulate hepatic lymph vessel bed

Volumetric imaging reveals VEGF-C-dependent formation of hepatic lymph vessels in mice

International audienceThe liver is a major biosynthetic and detoxifying organ in vertebrates, but also generates 25%-50% of the lymph passing through the thoracic duct and is thereby the organ with the highest contribution to lymph flow. In contrast to its metabolic function, the role of the liver for lymph generation and composition is presently severely understudied. We took a rigorous, volume imaging-based approach to describe the microarchitecture and spatial composition of the hepatic lymphatic vasculature with cellular resolution in whole mount immune stained specimen ranging from thick sections up to entire mouse liver lobes. Here, we describe that in healthy adult livers, lymphatic vessels were exclusively located within the portal tracts, where they formed a unique, highly ramified tree. Ragged, spiky initials enmeshed the portal veins along their entire length and communicated with long lymphatic vessels that followed the path of the portal vein in close association with bile ducts. Together these lymphatic vessels formed a uniquely shaped vascular bed with a delicate architecture highly adapted to the histological structure of the liver. Unexpectedly, with the exception of short collector stretches at the porta hepatis, which we identified as exit point of the liver lymph vessels, the entire hepatic lymph vessel system was comprised of capillary lymphatic endothelial cells only. Functional experiments confirmed the space of Disse as the origin of the hepatic lymph and flow via the space of Mall to the portal lymph capillaries. After entry into the lymphatic initials, the lymph drained retrograde to the portal blood flow towards the exit at the liver hilum. Perinatally, the liver undergoes complex changes transforming from the main hematopoietic to the largest metabolic organ. We investigated the time course of lymphatic vessel development and identified the hepatic lymphatics to emerge postnatally in a process that relies on input from the VEGF-C/VERGFR-3 growth factor-receptor pair for formation of the fully articulate hepatic lymph vessel bed

Volumetric imaging reveals VEGF-C-dependent formation of hepatic lymph vessels in mice

The liver is a major biosynthetic and detoxifying organ in vertebrates, but also generates 25%–50% of the lymph passing through the thoracic duct and is thereby the organ with the highest contribution to lymph flow. In contrast to its metabolic function, the role of the liver for lymph generation and composition is presently severely understudied. We took a rigorous, volume imaging-based approach to describe the microarchitecture and spatial composition of the hepatic lymphatic vasculature with cellular resolution in whole mount immune stained specimen ranging from thick sections up to entire mouse liver lobes. Here, we describe that in healthy adult livers, lymphatic vessels were exclusively located within the portal tracts, where they formed a unique, highly ramified tree. Ragged, spiky initials enmeshed the portal veins along their entire length and communicated with long lymphatic vessels that followed the path of the portal vein in close association with bile ducts. Together these lymphatic vessels formed a uniquely shaped vascular bed with a delicate architecture highly adapted to the histological structure of the liver. Unexpectedly, with the exception of short collector stretches at the porta hepatis, which we identified as exit point of the liver lymph vessels, the entire hepatic lymph vessel system was comprised of capillary lymphatic endothelial cells only. Functional experiments confirmed the space of Disse as the origin of the hepatic lymph and flow via the space of Mall to the portal lymph capillaries. After entry into the lymphatic initials, the lymph drained retrograde to the portal blood flow towards the exit at the liver hilum. Perinatally, the liver undergoes complex changes transforming from the main hematopoietic to the largest metabolic organ. We investigated the time course of lymphatic vessel development and identified the hepatic lymphatics to emerge postnatally in a process that relies on input from the VEGF-C/VERGFR-3 growth factor—receptor pair for formation of the fully articulate hepatic lymph vessel bed

An Evolutionarily Conserved Role for Polydom/Svep1 during Lymphatic Vessel Formation

Rationale: Lymphatic vessel formation and function constitutes a physiologically and pathophysiologically important process, but its genetic control is not well understood. Objective: Here, we identify the secreted Polydom/Svep1 protein as essential for the formation of the lymphatic vasculature. We analyzed mutants in mice and zebrafish to gain insight into the role of Polydom/Svep1 in the lymphangiogenic process. Methods and Results: Phenotypic analysis of zebrafish polydom/svep1 mutants showed a decrease in venous and lymphovenous sprouting, which leads to an increased number of intersegmental arteries. A reduced number of primordial lymphatic cells populated the horizontal myoseptum region but failed to migrate dorsally or ventrally, resulting in severe reduction of the lymphatic trunk vasculature. Corresponding mutants in the mouse Polydom/Svep1 gene showed normal egression of Prox-1+ cells from the cardinal vein at E10.5, but at E12.5, the tight association between the cardinal vein and lymphatic endothelial cells at the first lymphovenous contact site was abnormal. Furthermore, mesenteric lymphatic structures at E18.5 failed to undergo remodeling events in mutants and lacked lymphatic valves. In both fish and mouse embryos, the expression of the gene suggests a nonendothelial and noncell autonomous mechanism. Conclusions: Our data identify zebrafish and mouse Polydom/Svep1 as essential extracellular factors for lymphangiogenesis. Expression of the respective genes by mesenchymal cells in intimate proximity with venous and lymphatic endothelial cells is required for sprouting and migratory events in zebrafish and for remodeling events of the lymphatic intraluminal valves in mouse embryos

An Evolutionarily Conserved Role for Polydom/Svep1 During Lymphatic Vessel Formation

Rationale: Lymphatic vessel formation and function constitutes a physiologically and pathophysiologically important process, but its genetic control is not well understood. Objective: Here, we identify the secreted Polydom/Svep1 protein as essential for the formation of the lymphatic vasculature. We analyzed mutants in mice and zebrafish to gain insight into the role of Polydom/Svep1 in the lymphangiogenic process. Methods and Results: Phenotypic analysis of zebrafish polydom/svep1 mutants showed a decrease in venous and lymphovenous sprouting, which leads to an increased number of intersegmental arteries. A reduced number of primordial lymphatic cells populated the horizontal myoseptum region but failed to migrate dorsally or ventrally, resulting in severe reduction of the lymphatic trunk vasculature. Corresponding mutants in the mouse Polydom/Svep1 gene showed normal egression of Prox-1+ cells from the cardinal vein at E10.5, but at E12.5, the tight association between the cardinal vein and lymphatic endothelial cells at the first lymphovenous contact site was abnormal. Furthermore, mesenteric lymphatic structures at E18.5 failed to undergo remodeling events in mutants and lacked lymphatic valves. In both fish and mouse embryos, the expression of the gene suggests a nonendothelial and noncell autonomous mechanism. Conclusions: Our data identify zebrafish and mouse Polydom/Svep1 as essential extracellular factors for lymphangiogenesis. Expression of the respective genes by mesenchymal cells in intimate proximity with venous and lymphatic endothelial cells is required for sprouting and migratory events in zebrafish and for remodeling events of the lymphatic intraluminal valves in mouse embryos

Volumetric imaging reveals VEGF-C-dependent formation of hepatic lymph vessels in mice

The liver is a major biosynthetic and detoxifying organ in vertebrates, but also generates 25%–50% of the lymph passing through the thoracic duct and is thereby the organ with the highest contribution to lymph flow. In contrast to its metabolic function, the role of the liver for lymph generation and composition is presently severely understudied. We took a rigorous, volume imaging-based approach to describe the microarchitecture and spatial composition of the hepatic lymphatic vasculature with cellular resolution in whole mount immune stained specimen ranging from thick sections up to entire mouse liver lobes. Here, we describe that in healthy adult livers, lymphatic vessels were exclusively located within the portal tracts, where they formed a unique, highly ramified tree. Ragged, spiky initials enmeshed the portal veins along their entire length and communicated with long lymphatic vessels that followed the path of the portal vein in close association with bile ducts. Together these lymphatic vessels formed a uniquely shaped vascular bed with a delicate architecture highly adapted to the histological structure of the liver. Unexpectedly, with the exception of short collector stretches at the porta hepatis, which we identified as exit point of the liver lymph vessels, the entire hepatic lymph vessel system was comprised of capillary lymphatic endothelial cells only. Functional experiments confirmed the space of Disse as the origin of the hepatic lymph and flow via the space of Mall to the portal lymph capillaries. After entry into the lymphatic initials, the lymph drained retrograde to the portal blood flow towards the exit at the liver hilum. Perinatally, the liver undergoes complex changes transforming from the main hematopoietic to the largest metabolic organ. We investigated the time course of lymphatic vessel development and identified the hepatic lymphatics to emerge postnatally in a process that relies on input from the VEGF-C/VERGFR-3 growth factor—receptor pair for formation of the fully articulate hepatic lymph vessel bed

The role of fatty acid beta-oxidation in lymphangiogenesis

Lymphatic vessels are lined by lymphatic endothelial cells (LECs), and are critical for health. However, the role of metabolism in lymphatic development has not yet been elucidated. Here we report that in transgenic mouse models, LEC-specific loss of CPT1A, a rate-controlling enzyme in fatty acid β-oxidation, impairs lymphatic development. LECs use fatty acid β-oxidation to proliferate and for epigenetic regulation of lymphatic marker expression during LEC differentiation. Mechanistically, the transcription factor PROX1 upregulates CPT1A expression, which increases acetyl coenzyme A production dependent on fatty acid β-oxidation. Acetyl coenzyme A is used by the histone acetyltransferase p300 to acetylate histones at lymphangiogenic genes. PROX1-p300 interaction facilitates preferential histone acetylation at PROX1-target genes. Through this metabolism-dependent mechanism, PROX1 mediates epigenetic changes that promote lymphangiogenesis. Notably, blockade of CPT1 enzymes inhibits injury-induced lymphangiogenesis, and replenishing acetyl coenzyme A by supplementing acetate rescues this process in vivo.status: publishe

The role of fatty acid beta-oxidation in lymphangiogenesis.

Lymphatic vessels are lined by lymphatic endothelial cells (LECs), and are critical for health. However, the role of metabolism in lymphatic development has not yet been elucidated. Here we report that in transgenic mouse models, LEC-specific loss of CPT1A, a rate-controlling enzyme in fatty acid beta-oxidation, impairs lymphatic development. LECs use fatty acid beta-oxidation to proliferate and for epigenetic regulation of lymphatic marker expression during LEC differentiation. Mechanistically, the transcription factor PROX1 upregulates CPT1A expression, which increases acetyl coenzyme A production dependent on fatty acid beta-oxidation. Acetyl coenzyme A is used by the histone acetyltransferase p300 to acetylate histones at lymphangiogenic genes. PROX1-p300 interaction facilitates preferential histone acetylation at PROX1-target genes. Through this metabolism-dependent mechanism, PROX1 mediates epigenetic changes that promote lymphangiogenesis. Notably, blockade of CPT1 enzymes inhibits injury-induced lymphangiogenesis, and replenishing acetyl coenzyme A by supplementing acetate rescues this process in vivo