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Targeting endothelial coagulation signaling ameliorates liver obstructive cholestasis and dysfunctional angiogenesis.

ABSTRACT: Cholestatic fibrogenesis is a pathobiological process in which cumulative injury to the bile ducts coincides with progressive liver fibrosis. The pathobiologic mechanisms underlying fibrogenesis and disease progression remain poorly understood. Currently, there is no effective treatment for liver fibrosis. In this study, we discovered that components of the coagulation cascade were associated with the advanced progression of obstructive cholestasis, and anticoagulant therapy could improve liver cholestasis-induced fibrosis. In a mouse model of common bile duct ligation (BDL), which mimics cholestatic liver injury, RNA sequencing analysis revealed an increased expression of coagulation factors in endothelial cells. Pharmacological targeting of the coagulation signaling by hirudin, an anticoagulatory antagonist of thrombin, ameliorated obstructive cholestasis, and attenuated liver fibrosis symptoms. Hirudin attenuated fibrosis-associated angiogenesis, endothelial-to-mesenchymal transition (EndMT), and tissue hypoxia and reduced liver inflammation after BDL. Furthermore, hirudin suppressed YAP (Yes-associated protein) signaling and its downstream effectors in vascular endothelial cells, which are considered with profibrotic characteristics. In conclusion, we demonstrated that pharmacological targeting of coagulation signaling by hirudin has the potential to alleviate liver obstructive cholestasis and fibrosis.

SUBMITTER: Yang X

PROVIDER: S-EPMC10621472 | biostudies-literature | 2023 Jul

REPOSITORIES: biostudies-literature

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Targeting endothelial coagulation signaling ameliorates liver obstructive cholestasis and dysfunctional angiogenesis.

Yang Xue X Ou Yangying Y Yang Ying Y Wang Lu L Zhang Yuwei Y Zhao Fulan F Shui Pixian P Qing Jie J

Experimental biology and medicine (Maywood, N.J.) 20230701 14

Cholestatic fibrogenesis is a pathobiological process in which cumulative injury to the bile ducts coincides with progressive liver fibrosis. The pathobiologic mechanisms underlying fibrogenesis and disease progression remain poorly understood. Currently, there is no effective treatment for liver fibrosis. In this study, we discovered that components of the coagulation cascade were associated with the advanced progression of obstructive cholestasis, and anticoagulant therapy could improve liver ...[more]

PMID: 37644866

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Project description:RationaleThe highly conserved NOTCH (neurogenic locus notch homolog protein) signaling pathway functions as a key cell-cell interaction mechanism controlling cell fate and tissue patterning, whereas its dysregulation is implicated in a variety of developmental disorders and cancers. The pivotal role of endothelial NOTCH in regulation of angiogenesis is widely appreciated; however, little is known about what controls its signal transduction. Our previous study indicated the potential role of post-translational SUMO (small ubiquitin-like modifier) modification (SUMOylation) in vascular disorders.ObjectiveThe aim of this study was to investigate the role of SUMOylation in endothelial NOTCH signaling and angiogenesis.Methods and resultsEndothelial SENP1 (sentrin-specific protease 1) deletion, in newly generated endothelial SENP1 (the major protease of the SUMO system)-deficient mice, significantly delayed retinal vascularization by maintaining prolonged NOTCH1 signaling, as confirmed in cultured endothelial cells. An in vitro SUMOylation assay and immunoprecipitation revealed that when SENP1 associated with N1ICD (NOTCH1 intracellular domain), it functions as a deSUMOylase of N1ICD SUMOylation on conserved lysines. Immunoblot and immunoprecipitation analyses and dual-luciferase assays of natural and SUMO-conjugated/nonconjugated NOTCH1 forms demonstrated that SUMO conjugation facilitated NOTCH1 cleavage. This released N1ICD from the membrane and stabilized it for translocation to the nucleus where it functions as a cotranscriptional factor. Functionally, SENP1-mediated NOTCH1 deSUMOylation was required for NOTCH signal activation in response to DLL4 (Delta-like 4) stimulation. This in turn suppressed VEGF (vascular endothelial growth factor) receptor signaling and angiogenesis, as evidenced by immunoblotted signaling molecules and in vitro angiogenesis assays.ConclusionsThese results establish reversible NOTCH1 SUMOylation as a regulatory mechanism in coordinating endothelial angiogenic signaling; SENP1 acts as a critical intrinsic mediator of this process. These findings may apply to NOTCH-regulated biological events in nonvascular tissues and provide a novel therapeutic strategy for vascular diseases and tumors.

Project description:BackgroundAging leads to vascular endothelial cell senescence. Decreased expression of VEGFA and VEGFR2 plays a crucial role in impairing angiogenesis in senescent endothelial cells. Noncoding RNAs, including circular RNAs (circRNAs) and microRNAs (miRNAs), regulate endothelial cell proliferation, differentiation, apoptosis, and migration and participate in the occurrence and development of vascular diseases. However, the mechanism of noncoding RNAs in age-related vascular endothelial dysfunction remains unclear. Here, we aimed to identify the circRNA that is associated with VEGF/VEGFR2 signaling pathway activation in angiogenesis.MethodsImmunoblotting, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), in vitro and in vivo experiments, luciferase assays, and chromatin immunoprecipitation followed by qRT-PCR (ChIP-qPCR) assays were performed to clarify the roles played by circCRIM1 in mouse aortic endothelial cell (MAEC) angiogenesis.ResultsCircCRIM1 expression was downregulated in both an aging mouse model of lower limb ischemia in vivo and aging MAECs in vitro. Overexpressing circCRIM1 mediated through a plasmid or adeno-associated virus (AAV) reversed the downregulation of angiogenesis-related phenotype acquisition during aging. MiR-455-3p was confirmed to be a potential target of circCRIM1 through luciferase assays followed by RNA fluorescence in situ hybridization (FISH), which revealed the colocalization of circCRIM1 and miR-455-3p. CircCRIM1 was found to be a competitive endogenous RNA that sponged miR-455-3p and regulated angiogenesis-related phenotypes in MAECs. Furthermore, Twist1 was found to be downstream of miR-455-3p. A ChIP-qPCR assay showed that Twist1 promoted VEGFR2 expression by binding to the promoter region, playing a vital role in angiogenesis.ConclusionsDecreased expression of circCRIM1 impaired angiogenesis in aging via the miR-455-3p/Twist1/VEGFR2 axis. Our findings suggest that overexpression of circCRIM1 may be an effective therapeutic strategy for promoting ischemic lower limb blood flow recovery.

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Targeting endothelial coagulation signaling ameliorates liver obstructive cholestasis and dysfunctional angiogenesis.

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Targeting endothelial coagulation signaling ameliorates liver obstructive cholestasis and dysfunctional angiogenesis.

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