Biology subjects

Kiyan, Y.

Publications and source records attributed to Kiyan, Y..

4 recordsLinked to original sources

bioRxiv2025

Macrophages induce stromal differentiation and endothelialization in iPSC-derived kidney organoids.

Human induced pluripotent stem cell (iPSC) - derived kidney organoids are powerful models of kidney development and disease but lack mature vasculature. Here, we show that co-culture with human monocyte-derived macrophages (MDM) induces robust endothelial and stromal differentiation in iPSC-derived kidney organoids. MDM promoted the formation of lumenized capillary networks, an effect reproduced by iPSC-derived macrophages and the THP-1 monocyte line. In contrast, organoids without MDM displayed only transient endothelial differentiation that coincided with transient VEGFA upregulation and that regressed during maturation. MDM counteracted this transient phase by releasing soluble Neuropilin-1 (sNRP1), which sequestered VEGFA and stabilized late endothelial development, evidenced by sustained CD31 promoter activity. Transcriptomic profiling revealed that MDM redirected mesodermal fate suppressing lateral plate and cardiac mesoderm while promoting paraxial and intermediate mesoderm and the generation of FoxD1 stromal progenitors via CXCL5 (ENA-78) secretion. These stromal cells supported late endothelial maturation and enhanced overall organoid differentiation. Our findings uncover a macrophage-driven mechanism coupling stromal and vascular development, providing a strategy to achieve functional vascularization in kidney organoids and improving their physiological relevance for research applications.

cell biology↗

bioRxiv2024

Heparanase 2 regulates endothelial permeability and prevents from proteinuria via VEGFA and FGF signaling

BackgroundHeparan sulfates (HS) attached to the apical surface of vascular endothelial cells (ECs) play an important role in regulating endothelial permeability and ligand recognition by cell-surface receptors. Shedding of heparan sulfate (HS) from the EC surface increases vascular leakage and is associated with vascular diseases. Recently, heparanase 2 (Hpa2) was described as a novel regulatory molecule that controls HS shedding. However, its role in regulating HS physiology in the vascular endothelium is largely unknown. Here, we characterize the role of endogenous Hpa2 in the vertebrate vascular system. MethodsWe use zebrafish larvae as our primary animal model. Hpa2 expression and localization was examined by in situ hybridization and immunofluorescence. Hpa2 loss-of-function (LOF) was induced by CRISPR-Cas9 or morpholino antisense strategies. We assessed vascular permeability, blood vessel architecture, and EC morphology using transgenic zebrafish and transmission electron microscopy. EC expression profiles and HS quantity were analyzed in Hpa2-LOF larvae. The capacity of recombinant Hpa2 to modulate signaling in ECs by the HS-binding growth factors fibroblast growth factor 2 (FGF2) and vascular endothelial growth factor A165 (VEGFA165) was tested by western blotting and immunofluorescence. Attenuation of the Hpa2-LOF phenotype was tested in vivo in zebrafish larvae via use of recombinant Hpa2 and pharmacological inhibition of FGF and VEGFA signaling. ResultsWe detected hpse2 expression in hepatic tissue and localized the protein in blood vessels. Hpa2-LOF larvae exhibited increased vascular permeability, occasional hypersprouting, and altered EC and extracellular matrix (ECM) morphology. Hpa2-LOF also reduced HS levels and caused changes in the endothelial transcriptome characterized by dysregulated genes involved in ECM-receptor interaction and signal transduction regulation. Recombinant hHpa2 rescued the Hpa2-LOF phenotype in zebrafish. We showed in vitro that Hpa2 competes with FGF2 and VEGFA165 for binding on the EC surface and consequently reduces the cellular response these factors elicit. Pharmacological inhibition of these pathways alleviated the Hpa2-LOF phenotype in zebrafish. ConclusionWe conclude that Hpa2 is a circulating molecule that maintains vascular integrity by regulating HS-dependent processes on the EC surface. These results may translate into novel strategies applying recombinant Hpa2 to treat microvascular diseases.

physiology↗

bioRxiv2020

Urokinase receptor associates with TLR4 interactome to promote LPS response

GPI-anchored uPAR is the receptor for the extracellular serine protease urokinase-type plasminogen activator (uPA). Binding of uPA to uPAR localizes proteolytic cascade activation at the cell surface and can induce intracellular signaling. As uPAR possesses no transmembrane domain, it relies on uPAR cross-talk with various membrane receptors. Though uPAR role in inflammatory processes is well documented, underlying mechanisms are not fully understood. In this study we demonstrate that uPAR is a part of Toll-like receptor 4 (TLR4) interactome. GPI-uPAR and soluble uPAR colocalized with TLR4 on the cell membrane and interacted with scavenger receptor CD36. We show that downregulation of uPAR expression resulted in diminished LPS-induced TLR4 signaling, less activation of NF{kappa}B, and decreased secretion of inflammatory mediators in myeloid and non-myeloid cells in vitro. In vivo uPAR-/- mice demonstrated strongly diminished inflammatory response and better organ functions in cecal ligation and puncture mouse polymicrobial sepsis model. Our data show that uPAR can interfere with innate immunity response via TLR4 and this mechanism represents a potentially important target in inflammation and sepsis therapy.

cell biology↗

bioRxiv2019

Heparanase-2 protects from endothelial injury by inhibiting TLR4 signaling

ObjectiveThe endothelial glycocalyx and the regulation of its shedding are important to vascular health. Endo-{beta}-D-glucuronidase heparanase-1 (HPSE1) is the only enzyme that can shed heparan sulfate. However, the mechanisms are not well understood.\n\nApproach and resultsTo investigate HPSE1 and its endogenous inhibitor, heparanase-2 (HPSE2), we used cell culture, lentiviral protein overexpression, a microfluidic chip model of cell culture under shear stress conditions, and lipopolysaccharide (LPS) injections in mice. We show that HPSE1 activity aggravated Toll-like receptor 4 (TLR4)-mediated response of endothelial cells to LPS. On the contrary, HPSE2 overexpression was protective. The microfluidic chip flow model confirmed that HPSE2 prevented heparan sulfate shedding by HPSE1. Furthermore, heparan sulfate did not interfere with cluster of differentiation-14 (CD14)-dependent LPS binding, but instead reduced the presentation of the LPS-CD14 complex to TLR4. HPSE2 reduced LPS-mediated TLR4 activation by LPS, subsequent cell signaling, and cytokine expression. Moreover, HPSE2-overexpressing endothelial cells remained protected against LPS-mediated loss of cell-cell contacts. In vivo, expression of HPSE2 in plasma and kidney medullary capillaries was decreased in mouse sepsis model. We next applied purified HPSE2 in mice and observed decreases in TNF and IL-6 plasma concentrations after intravenous LPS injections.\n\nConclusionsOur data demonstrate the important role of heparan sulfate and the glycocalyx in endothelial cell activation and suggest a protective role of HPSE2 in microvascular inflammation. HPSE2 offers new options for protection against HPSE1-mediated endothelial damage and preventing microvascular disease.\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC=\"FIGDIR/small/705939v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (50K):\norg.highwire.dtl.DTLVardef@1bae526org.highwire.dtl.DTLVardef@11a6039org.highwire.dtl.DTLVardef@a36deforg.highwire.dtl.DTLVardef@1869ea8_HPS_FORMAT_FIGEXP M_FIG Graphical abstract\n\nC_FIG O_TBL View this table:\norg.highwire.dtl.DTLVardef@11a3deborg.highwire.dtl.DTLVardef@1d71919org.highwire.dtl.DTLVardef@b0c8a3org.highwire.dtl.DTLVardef@4477f3org.highwire.dtl.DTLVardef@11f1588_HPS_FORMAT_FIGEXP M_TBL O_TABLECAPTIONNon standard abbreviations\n\nC_TABLECAPTION C_TBL

cell biology↗