Aplexone Targets the HMG-CoA Reductase Pathway and Differentially Regulates Arteriovenous Angiogenesis
Ontology highlight
ABSTRACT: Arterial and venous endothelial cells exhibit distinct molecular characteristics at early developmental stages. These lineage-specific molecular programs are instructive to the development of distinct vascular architectures and physiological conditions of arteries and veins, but their roles in angiogenesis remain unexplored. Here, we show that the caudal vein plexus in zebrafish forms by endothelial cell sprouting, migration and anastomosis, providing a venous-specific angiogenesis model. Using this model, we identified a novel compound, aplexone, which effectively suppresses venous, but not arterial, angiogenesis. Multiple lines of evidence indicate that aplexone differentially regulates arteriovenous angiogenesis by targeting the HMG-CoA reductase (HMGCR) pathway. Treatment with aplexone affects the transcription of enzymes in the HMGCR pathway and reduces cellular cholesterol levels. Injecting mevalonate, a metabolic product of HMGCR, reverses the inhibitory effect of aplexone on venous angiogenesis. In addition, aplexone treatment inhibits protein prenylation and blocking the activity of geranylgeranyl transferase induces a venous angiogenesis phenotype resembling that observed in aplexone-treated embryos. Furthermore, endothelial cells of venous origin have higher levels of proteins requiring geranylgeranylation than arterial endothelial cells and inhibiting the activity of Rac or Rho Kinase effectively reduces the migration of venous, but not arterial, endothelial cells. Taken together, our findings indicate that angiogenesis is differentially regulated by the HMGCR pathway via an arteriovenousdependent requirement for protein prenylation in zebrafish and human endothelial cells.
Project description:Formation and maturation of a functional blood vascular system is required for the development and maintenance of all tissues in the body. During the process of blood vessel development, primordial endothelial cells are formed and become specified toward arterial or venous fates to generate a circulatory network that provides nutrients and oxygen to, and removes metabolic waste from, all tissues. Specification of arterial and venous endothelial cells occurs in conjunction with suppression of endothelial cell cycle progression, and endothelial cell hyperproliferation is associated with potentially lethal arterial-venous malformations. However, the mechanistic role that cell cycle state plays in arterial-venous specification is unknown. Herein, studying retinal vascular development in Cdh5-CreERT2;R26FUCCI2aR reporter mice, we found that venous and arterial endothelial cells are in distinct cell cycle states during development and in adulthood. That is, venous endothelial cells reside in early G1 state, while arterial endothelial cells reside in late G1 state. Single cell RNA sequencing of developing retinal endothelial cells revealed that BMP signaling and early G1 state are enriched in venous endothelial cells, while TGF-b signaling and late G1 state are enriched in arterial endothelial cells. Cultured endothelial cells in early vs. late G1 exhibited significant differences in gene expression and activity, especially among BMP/TGF-b signaling components. The early G1 state was found to be essential for BMP4-induced venous gene expression, whereas late G1 state is essential for TGF-b1-induced arterial gene expression. In a mouse model of endothelial cell hyperproliferation and disrupted arterial-venous specification, pharmacological inhibition of endothelial cell cycle prevented the vascular defects. Collectively, our results show that endothelial cell cycle control plays a key role in arterial-venous network formation, and distinct cell cycle states provide distinct windows of opportunity for the molecular induction of arterial vs. venous specification.
Project description:Formation and maturation of a functional blood vascular system is required for the development and maintenance of all tissues in the body. During the process of blood vessel development, primordial endothelial cells are formed and become specified toward arterial or venous fates to generate a circulatory network that provides nutrients and oxygen to, and removes metabolic waste from, all tissues. Specification of arterial and venous endothelial cells occurs in conjunction with suppression of endothelial cell cycle progression, and endothelial cell hyperproliferation is associated with potentially lethal arterial-venous malformations. However, the mechanistic role that cell cycle state plays in arterial-venous specification is unknown. Herein, studying vascular development in Cdh5-CreERT2;R26FUCCI2aR reporter mice, we find that venous and arterial endothelial cells exhibit a propensity for different cell cycle states during development and in adulthood. That is, venous endothelial cells are predominantly FUCCI-Negative, while arterial endothelial cells are enriched for the FUCCI-Red reporter. Single cell RNA sequencing analysis of developing retinal endothelial cells reveals that venous endothelial cells are enriched for the FUCCI-Negative state and BMP signaling, while arterial endothelial cells are enriched for the FUCCI-Red state and TGF-b signaling. Further transcriptional analyses and live imaging of cultured endothelial cells expressing the FUCCI reporter show that reporter-negative corresponds to an early G1 state and reporter-red corresponds to late G1 state. We find the early G1 state is essential for BMP4-induced venous gene expression, whereas late G1 state is essential for TGF-b1-induced arterial gene expression. In a mouse model of endothelial cell hyperproliferation and disrupted arterial-venous specification, pharmacological inhibition of endothelial cell cycle prevents the vascular defects. Collectively, our results show that endothelial cell cycle control plays a key role in arterial-venous network formation, and distinct cell cycle states provide distinct windows of opportunity for the molecular induction of arterial vs. venous specification.
Project description:Formation and maturation of a functional blood vascular system is required for the development and maintenance of all tissues in the body. During the process of blood vessel development, primordial endothelial cells are formed and become specified toward arterial or venous fates to generate a circulatory network that provides nutrients and oxygen to, and removes metabolic waste from, all tissues. Specification of arterial and venous endothelial cells occurs in conjunction with suppression of endothelial cell cycle progression, and endothelial cell hyperproliferation is associated with potentially lethal arterial-venous malformations. However, the mechanistic role that cell cycle state plays in arterial-venous specification is unknown. Herein, studying vascular development in Cdh5-CreERT2;R26FUCCI2aR reporter mice, we find that venous and arterial endothelial cells exhibit a propensity for different cell cycle states during development and in adulthood. That is, venous endothelial cells are predominantly FUCCI-Negative, while arterial endothelial cells are enriched for the FUCCI-Red reporter. Single cell RNA sequencing analysis of developing retinal endothelial cells reveals that venous endothelial cells are enriched for the FUCCI-Negative state and BMP signaling, while arterial endothelial cells are enriched for the FUCCI-Red state and TGF-b signaling. Further transcriptional analyses and live imaging of cultured endothelial cells expressing the FUCCI reporter show that reporter-negative corresponds to an early G1 state and reporter-red corresponds to late G1 state. We find the early G1 state is essential for BMP4-induced venous gene expression, whereas late G1 state is essential for TGF-b1-induced arterial gene expression. In a mouse model of endothelial cell hyperproliferation and disrupted arterial-venous specification, pharmacological inhibition of endothelial cell cycle prevents the vascular defects. Collectively, our results show that endothelial cell cycle control plays a key role in arterial-venous network formation, and distinct cell cycle states provide distinct windows of opportunity for the molecular induction of arterial vs. venous specification.
Project description:We applied single-cell RNA-sequencing (scRNA-seq) of vascular endothelial cells isolated from zebrafish embryos at the 24 hours post fertilization (hpf) stage. Six distinct clusters or subclusters related to vascular endothelial cells were identified which include arterial, two venous, cranial, endocardial and endothelial progenitor cell subtypes
Project description:Molecular pathways regulating the development of arterial and venous endothelial cells (ECs) are now well-established, but control of parallel arterial-venous (A-V) alignment is unclear. We report that arterial-venous alignment in the skin is determined by apelin receptor (APJ) expression in venous ECs. We used microarrays to detail the global programme of gene expression in endothelial cells that has relationship with the deficient of APJ. Endothelial cells were marked and isolated by Fluorescence-activated cell sorting in Trizol.
Project description:Molecular pathways regulating the development of arterial and venous endothelial cells (ECs) are now well-established, but control of parallel arterial-venous (A-V) alignment is unclear. We report that arterial-venous alignment in the skin is determined by apelin receptor (APJ) expression in venous ECs.
Project description:The vascular tree has considerable diversity, with discrete regions having different physiologic characteristics and permeability. Of note are venules that are significantly more sensitive to pro-inflammatory cytokines than arterioles. We used microarrays to identify molecular signatures that distinguish primary human venous endothelial cells from arterial endothelial cells. We used microarrays to identify genes differentially expressed by venous vs arterial human endothelial cells.
Project description:Distinct endothelial cell cycle states (early G1 vs. late G1) provide different “windows of opportunity” to enable the differential expression of genes that regulate venous and arterial specification, respectively. Endothelial cell cycle control and arterial-venous identities are disrupted in vascular malformations including arteriovenous (AV) shunts which is a hallmark of hereditary hemorrhagic telangiectasia (HHT). We show how endothelial cell late G1 arrest induced by Palbociclib modulates the expression of genes regulating arterio-venous identity and prevents AVM development induced by BMP9/10 inhibition.
Project description:Background: Hemorrhoids are common diseases of the anorectal system and are usually accompanied by vascular proliferation and edema. Curcumin is a natural molecule with potential anti-inflammatory, antitumor and antioxidant effects. This study aimed to explore the effect and molecular mechanism of curcumin in alleviating angiogenesis in rat hemorrhoids. Methods: A rat model of hemorrhoids was established via glacial acetic acid induction. Human venous endothelial cells were collected to perform cell experiments. The rats received curcumin at 25, 50, or 100 mg/kg/d. The rats were scored for perianal symptoms and inflammation levels, and the expression of angiogenesis-related factors was evaluated by immunofluorescence. Bioinformatics analysis, dual-luciferase, RT‒qPCR and Western blotting were used to verify the molecular mechanism of curcumin in the treatment of hemorrhoids. Results: Curcumin inhibited angiogenesis in endothelial cells and relieved the symptoms of hemorrhoids in rats. The study revealed that the expression of miR-190a-5p was significantly downregulated in hemorrhoids, whereas curcumin promoted the expression of miR-190a-5p. The results revealed that miR-190a-5p could inhibit angiogenesis and inflammation in endothelial cells by targeting TGFBR2, thereby alleviating the development of hemorrhoids. Mechanistically, curcumin inhibits the expression of TGFBR2 by upregulating miR-190a-5p, effectively blocking the angiogenesis ability of endothelial cells and thereby attenuating the progression and development of hemorrhoids in rats. Conclusion: Curcumin effectively inhibits angiogenesis in endothelial cells and relieves inflammation and the progression of hemorrhoids by regulating the miR-190a-5p/TGFBR2 molecular pathway.
Project description:scRNA-seq was used to investigate the different endothelial cell population and differential gene expression between arterial and venous cells.