Dopamine receptor D2 antagonism normalizes profibrotic macrophage-endothelial crosstalk in non-alcoholic steatohepatitis.

BACKGROUND & AIMS: Currently there is no effective treatment for liver fibrosis, which is one of the main histological determinants of non-alcoholic steatohepatitis (NASH). While Hippo/YAP (Yes-associated protein) signaling is essential for liver regeneration, its aberrant activation frequently leads to fibrosis and tumorigenesis. Unravelling "context-specific" contributions of YAP in liver repair might help selectively bypass fibrosis and preserve the pro-regenerative YAP function in hepatic diseases. METHODS: We used murine liver fibrosis and minipig NASH models, and liver biopsies from patients with cirrhosis. Single-cell RNA-sequencing (scRNA-Seq) was performed, and a G-protein-coupled receptor (GPCR) ligand screening system was used to identify cell-selective YAP inhibitors. RESULTS: YAP levels in macrophages are increased in the livers of humans and mice with liver fibrosis. The increase in type I interferon and attenuation of hepatic fibrosis observed in mice specifically lacking Yap1 in myeloid cells provided further evidence for the fibrogenic role of macrophage YAP. ScRNA-Seq further showed that defective YAP pathway signaling in macrophages diminished a fibrogenic vascular endothelial cell subset that exhibited profibrotic molecular signatures such as angiocrine CTGF and VCAM1 expression. To specifically target fibrogenic YAP in macrophages, we utilized a GPCR ligand screening system and identified a dopamine receptor D2 (DRD2) antagonist that selectively blocked YAP in macrophages but not hepatocytes. Genetic and pharmacological targeting of macrophage DRD2 attenuated liver fibrosis. In a large animal (minipig) NASH model recapitulating human pathology, the DRD2 antagonist blocked fibrosis and restored hepatic architecture. CONCLUSIONS: DRD2 antagonism selectively targets YAP-dependent fibrogenic crosstalk between macrophages and CTGF+VCAM1+ vascular niche, promoting liver regeneration over fibrosis in both rodent and large animal models. LAY SUMMARY: Fibrosis in the liver is one of the main histological determinants of non-alcoholic steatohepatitis (NASH), a disease paralleling a worldwide surge in metabolic syndromes. Our study demonstrates that a macrophage-specific deficiency in Yes-associated protein (YAP) attenuates liver fibrosis. Dopamine receptor D2 (DRD2) antagonism selectively blocks YAP in macrophages and thwarts liver fibrosis in both rodent and large animal models, and thus holds potential for the treatment of NASH.

Selective Targeting of Vascular Endothelial YAP Activity Blocks EndMT and Ameliorates Unilateral Ureteral Obstruction-Induced Kidney Fibrosis.

Kidney fibrosis is accompanied by vascular dysfunction. Discovering new ways to ameliorate dysfunctional angiogenesis may bypass kidney fibrosis. YAP (Yes-associated protein) plays a multifaceted role during angiogenesis. Here, we found that selectively targeting YAP signaling in the endothelium ameliorates unilateral ureteral obstruction (UUO)-induced kidney fibrosis. Genetic deletion of Yap1, encoding YAP protein, in VE-cadherin+ endothelial cells inhibited endothelial-to-mesenchymal transition (EndMT) and dysfunctional angiogenesis and improved obstructive nephropathy and kidney fibrosis. Treatment with the systemic YAP inhibitor verteporfin worsened kidney fibrosis symptoms because of its lack of cell specificity. In an attempt to identify endothelial-specific YAP modulators, we found that G-protein-coupled receptor coagulation factor II receptor-like 1 (F2RL1) was highly expressed in vessels after UUO-induced kidney fibrosis. The F2RL1 peptide antagonist FSLLRY-NH2 selectively blocked YAP activity in endothelial cells and ameliorated kidney fibrosis. Thus, selective antagonization of endothelial YAP activity might bypass kidney fibrosis and provide new avenues for the design of antifibrotic therapies.

Identification of Interferon Receptor IFNAR2 As a Novel HCV Entry Factor by Using Chemical Probes.

Upon sensing pathogen-associated patterns and secreting interferons (IFNs) into the environment, host cells perceive extracellular type I IFNs by the IFNα/ß receptors IFNAR1 and IFNAR2 to stimulate downstream innate immune signaling cascades. Through the use of chemical probes, we demonstrated that IFNAR2 facilitates hepatitis C virus (HCV) entry. Silencing of IFNAR2 significantly attenuated HCV proliferation. IFNAR2 binds infectious HCV virions through a direct interaction of its D2 domain with the C-terminal end of apolipoprotein E (apoE) on the viral envelope and facilitates virus entry into host cells. The antibody against the IFNAR2 D2 domain attenuates IFNAR2-apoE interaction and impairs HCV infection. The recombinant IFNAR2 protein and the chemical probe potently inhibit major HCV genotypes in various human liver cells in vitro. Moreover, the impact of a chemical probe on HCV genotype 2a is also documented in immune-compromised humanized transgenic mice. Our results not only expand the understanding of the biology of HCV entry and the virus-host relationship but also reveal a new target for the development of anti-HCV entry inhibitors.

Single-cell transcriptomic landscape of nucleated cells in umbilical cord blood.

BACKGROUND: For both pediatric and adult patients, umbilical cord blood (UCB) transplant is a therapeutic option for a variety of hematologic diseases, such as blood cancers, myeloproliferative disorders, genetic diseases, and metabolic disorders. However, the level of cellular heterogeneity and diversity of nucleated cells in UCB has not yet been assessed in an unbiased and systemic fashion. In the present study, nucleated cells from UCB were subjected to single-cell RNA sequencing to simultaneously profile the gene expression signatures of thousands of cells, generating a rich resource for further functional studies. Here, we report the transcriptomes of 17,637 UCB cells, covering 12 major cell types, many of which can be further divided into distinct subpopulations. RESULTS: Pseudotemporal ordering of nucleated red blood cells identifies wave-like activation and suppression of transcription regulators, leading to a polarized cellular state, which may reflect nucleated red blood cell maturation. Progenitor cells in UCB also comprise 2 subpopulations with activation of divergent transcription programs, leading to specific cell fate commitment. Detailed profiling of cytotoxic cell populations unveiled granzymes B and K signatures in natural killer and natural killer T-cell types in UCB. CONCLUSIONS: Taken together, our data form a comprehensive single-cell transcriptomic landscape that reveals previously unrecognized cell types, pathways, and mechanisms of gene expression regulation. These data may contribute to the efficacy and outcome of UCB transplant, broadening the scope of research and clinical innovations.