Developmental Angiogenesis Research Articles

Hypoxia regulate developmental coronary angiogenesis potentially through VEGF-R2- and SOX17-mediated signaling.

The development of coronary vessels in embryonic mouse heart involves various progenitor populations, including sinus venosus (SV), endocardium, and proepicardium. ELA/APJ signaling is known to regulate coronary growth from the SV, whereas VEGF-A/VEGF-R2 signaling controls growth from the endocardium. Previous studies suggest hypoxia might regulate coronary growth, but its specific downstream pathways are unclear. In this study, we further investigated the role of hypoxia and have identified SOX17- and VEGF-R2-mediated signaling as the potential downstream pathways in its regulation of developmental coronary angiogenesis. HIF-1α stabilization by knocking out von Hippel Lindau (VHL) protein in the myocardium (cKO) disrupted normal coronary angiogenesis in embryonic mouse hearts, resembling patterns of accelerated coronary growth. VEGF-R2 expression was increased in coronary endothelial cells under hypoxia in vitro and in VHL cKO hearts in vivo. Similarly, SOX17 expression was increased in the VHL cKO hearts, while its knockout in the endocardium disrupted normal coronary growth. These findings provide further evidence that hypoxia regulates developmental coronary growth potentially through VEGF-R2 and SOX17 pathways, shedding light on mechanisms of coronary vessel development.

Endothelial c-Src mediates neovascular tuft formation in Oxygen-Induced Retinopathy

Vascular retinopathy, characterised by abnormal blood vessel growth in the retina, frequently results in vision impairment or loss. Neovascular tufts, a distinctive pathological feature of this condition, are highly leaky blood vessel structures, exacerbating secondary complications. Despite their clinical significance, the mechanisms underlying tuft development are not fully elucidated, posing challenges for effective management and treatment of vascular retinopathy. This study investigates the role of c-Src in neovascular tuft formation. Although c-Src has been acknowledged as a pivotal regulator in developmental angiogenesis within the retinal vasculature, its specific role in governing pathological retinal angiogenesis remains to be fully understood.The Oxygen-Induced Retinopathy (OIR) model was used for neovascular tufts formation in both Cre-mediated vascular specific c-Src knockout mice and wildtype littermates. High-resolution imaging and analysis of isolated retinas was conducted. c-Src depletion demonstrated a significant reduction in neovascular tufts within the OIR model. This decrease in tuft formation was observed independently of any alterations in cell death, cell proliferation or cell adhesion and the absence of c-Src did not impact tuft pericyte coverage and junctional morphology. These findings underscore the critical role of c-Src in the pathogenesis of neovascular tufts in vascular retinopathy. Understanding the molecular mechanisms involving c-Src may offer valuable insights for the development of targeted therapies aimed at mitigating vision-threatening complications associated with retinopathy.

Platelet Angiopoietin-1 Protects Against Murine Models of Tumor Metastasis.

In addition to their fundamental roles in preserving vascular integrity, platelets also contribute to tumor angiogenesis and metastasis. However, despite being a reservoir for angiogenic and metastatic cytokines, platelets also harbor negative regulators of tumor progression. Angpt1 (angiopoietin-1) is a cytokine essential for developmental angiogenesis that also protects against tumor cell metastasis through an undefined mechanism. Although activated platelets release Angpt1 from α-granules into circulation, the contributions of platelet Angpt1 to tumor growth, angiogenesis, and metastasis have not been investigated. Using cytokine arrays and ELISAs, we first compared platelet Angpt1 levels in breast and melanoma mouse tumor models to tumor-free controls. We then assessed tumor growth and metastasis in mice lacking megakaryocyte and platelet Angpt1 (Angpt1Plt KO). The spontaneous metastasis of mammary-injected tumor cells to the lungs was quantified using RT-PCR (reverse transcription-polymerase chain reaction). The lung colonization of intravenously injected tumor cells and tumor cell extravasation were determined using fluorescent microscopy and flow cytometry. Platelet Angpt1 is selectively upregulated in the PyMT (polyoma middle tumor antigen) breast cancer mouse model, and platelets are the principal source of Angpt1 in blood circulation. While primary tumor growth and angiogenesis were unaffected, Angpt1Plt KO mice had both increased spontaneous lung metastasis and tumor cell lung colonization following mammary or intravenous injection, respectively. Although platelet Angpt1 did not affect initial tumor cell entrapment in the lungs, Angpt1Plt KO mice had increased tumor cell retention and extravasation. Serum from Angpt1Plt KO mice increased endothelial permeability and reduced VE (vascular endothelial)-cadherin expression at endothelial junctions compared with serum from control mice (Angpt1WT). Platelets provide an intravascular source of Angpt1 that restrains tumor metastasis by preserving the lung microvasculature to limit tumor cell extravasation.

Therapeutic efficacy of ECs Foxp1 targeting Hif1α-Hk2 glycolysis signal to restrict angiogenesis

Endothelial cells (ECs) rely on glycolysis for energy production to maintain vascular homeostasis and the normalization of hyperglycolysis in tumor vessels has recently gained attention as a therapeutic target. We analyzed the TCGA database and found reduced Foxp1 expression in lung carcinoma. Immunostaining demonstrated reduced expression more restricted at tumor vascular ECs. Therefore, we investigated the function and mechanisms of Foxp1 in EC metabolism for tumor angiogenesis required for tumor growth. EC-Foxp1 deletion mice exhibited a significant increase of tumor and retinal developmental angiogenesis and Hif1α was identified as Foxp1 target gene, and Hk2 as Hif1α target gene. The Foxp1-Hif1α-Hk2 pathway in ECs is important in the regulation of glycolytic metabolism to govern tumor angiogenesis. Finally, we used genetic deletion of EC-Hif1α and RGD-peptide nanoparticles EC target delivery of Hif1α/Hk2-siRNAs to knockdown gene expression which reduced the tumor EC hyperglycolysis state and restricted angiogenesis for tumor growth. This study advances our understanding of EC metabolism for tumor angiogenesis, and meanwhile provides evidence for future therapeutic intervention of hyperglycolysis in tumor ECs for suppression of tumor growth.

The molecular mechanism responsible for HbSC retinopathy may depend on the action of the angiogenesis-related genes ROBO1 and SLC38A5.

HbSC disease, a less severe form of sickle cell disease, affects the retina more frequently and patients have higher rates of proliferative retinopathy that can progress to vision loss. This study aimed to identify differences in the expression of endothelial cell-derived molecules associated with the pathophysiology of proliferative sickle cell retinopathy (PSCR). RNAseq was used to compare the gene expression profile of circulating endothelial colony-forming cells from patients with SC hemoglobinopathy and proliferative retinopathy (n = 5), versus SC patients without retinopathy (n = 3). Real-time polymerase chain reaction (qRT-PCR) was used to validate the RNAseq results. A total of 134 differentially expressed genes (DEGs) were found. DEGs were mainly associated with vasodilatation, type I interferon signaling, innate immunity and angiogenesis. Among the DEGs identified, we highlight the most up-regulated genes ROBO1 (log2FoldChange = 4.32, FDR = 1.35E-11) and SLC38A5 (log2FoldChange = 3.36 FDR = 1.59E-07). ROBO1, an axon-guided receptor, promotes endothelial cell migration and contributes to the development of retinal angiogenesis and pathological ocular neovascularization. Endothelial SLC38A5, an amino acid (AA) transporter, regulates developmental and pathological retinal angiogenesis by controlling the uptake of AA nutrient, which may serve as metabolic fuel for the proliferation of endothelial cells (ECs) and consequent promotion of angiogenesis. Our data provide an important step towards elucidating the molecular pathophysiology of PSCR that may explain the differences in ocular manifestations between individuals with hemoglobinopathies and afford insights for new alternative strategies to inhibit pathological angiogenesis.

TMEM16F scramblase regulates angiogenesis via endothelial intracellular signaling.

TMEM16F (also known as ANO6), a Ca2+-activated lipid scramblase (CaPLSase) that dynamically disrupts lipid asymmetry, plays a crucial role in various physiological and pathological processes, such as blood coagulation, neurodegeneration, cell-cell fusion and viral infection. However, the mechanisms through which it regulates these processes remain largely elusive. Using endothelial cell-mediated angiogenesis as a model, here we report a previously unknown intracellular signaling function of TMEM16F. We demonstrate that TMEM16F deficiency impairs developmental retinal angiogenesis in mice and disrupts angiogenic processes in vitro. Biochemical analyses indicate that the absence of TMEM16F enhances the plasma membrane association of activated Src kinase. This in turn increases VE-cadherin phosphorylation and downregulation, accompanied by suppressed angiogenesis. Our findings not only highlight the role of intracellular signaling by TMEM16F in endothelial cells but also open new avenues for exploring the regulatory mechanisms for membrane lipid asymmetry and their implications in disease pathogenesis.

Nogo-B modulates physiological and pathological angiogenesis during development and disease

Angiogenesis, i.e., the formation of new blood vessels, is essential for normal development and functional vessel formation in ischemic diseases. During angiogenesis, endothelial cells (ECs) coordinate migration, proliferation, and metabolic fluxes to adapt to branching vessels' increasing energy demands. Nogo-B is a ubiquitously expressed reticulon family protein mainly localized to the endoplasmic reticulum (ER) membrane. Our previous studies have shown that the genetic loss of Nogo-B results in aberrant vascular remodeling and dysregulated endothelial barrier function. The current study aims to examine the role of endothelial Nogo-B in regulating developmental angiogenesis and pathological angiogenesis in retinopathy of prematurity (ROP). We hypothesized that endothelial Nogo-B is required for physiological and pathological angiogenesis, and Nogo-B depletion attenuates pathological angiogenesis. Using tamoxifen-inducible, endothelial cell-specific Nogo-B−/− (Nogo-BiECKO) mice, Nogo-B global knockout mice, and C57Bl/6 (WT) mice, our results show that loss of Nogo-B in ECs significantly delayed Isolectin B4 (IB4)-labeled retinal blood vessel sprouting. Impaired EC proliferation was observed by the staining of and incorporation of 5-ethynyl-2′-deoxyuridine (EdU). Furthermore, using the Oxygen Induced Retinopathy (OIR) model to mimic ROP, we could demonstrate that loss of Nogo-B in EC resulted in reduced neovascularization compared to WT. Mechanistically, using Percoll gradients, ultra-resolution microscopy, proximity ligation assay, and electron-microscopy, we show that Nogo-B is localized in the mitochondria-associated ER membrane (MAM) and regulated MAM remodeling. Using seahorse assay, quantitative RT-PCR, and metabolomics analysis, we determined that loss of Nogo-B altered VEGF-mediated EC glycolysis, which is needed for the energy demand during angiogenesis. These data suggest that Nogo-B regulates glucose metabolism in EC and plays a pivotal role in physiological and pathological angiogenesis. Targeting endothelial Nogo-B may provide a plausible approach to controlling the formation of pathological neovascularization in retinal diseases, such as ROP, diabetic retinopathy, and wet forms of age-related macular degeneration. This work was supported by the National Institutes of Health (R01HL153599) and awards from the American Heart Association award to JY. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Phosphoproteomic profiling identifies DNMT1 as a key substrate of beta IV spectrin-dependent ERK/MAPK signaling in suppressing angiogenesis

βIV-spectrin is a membrane-associated cytoskeletal protein that maintains the structural stability of cell membranes and integral proteins such as ion channels and transporters. Its biological functions are best characterized in the brain and heart, although recently we discovered a fundamental new role in the vascular system. Using cellular and genetic mouse models, we reported that βIV-spectrin acts as a critical regulator of developmental and tumor-associated angiogenesis. βIV-spectrin was shown to selectively express in proliferating endothelial cells (EC) and suppress VEGF/VEGFR2 signaling by enhancing receptor internalization and degradation. Here we examined how these events impact the downstream kinase signaling cascades and target substrates. Based on quantitative phosphoproteomics, we found that βIV-spectrin significantly affects the phosphorylation of epigenetic regulatory enzymes in the nucleus, among which DNA methyltransferase 1 (DNMT1) was determined as a top substrate. Biochemical and immunofluorescence results showed that βIV-spectrin inhibits DNMT1 function by activating ERK/MAPK, which in turn phosphorylates DNMT1 at S717 to impede its nuclear localization. Given that DNMT1 controls the DNA methylation patterns genome-wide, and is crucial for vascular development, our findings suggest that epigenetic regulation is a key mechanism by which βIV-spectrin suppresses angiogenesis.

Canthin-6-One Inhibits Developmental and Tumour-Associated Angiogenesis in Zebrafish.

Tumour-associated angiogenesis play key roles in tumour growth and cancer metastasis. Consequently, several anti-angiogenic drugs such as sunitinib and axitinib have been approved for use as anti-cancer therapies. However, the majority of these drugs target the vascular endothelial growth factor A (VEGFA)/VEGF receptor 2 (VEGFR2) pathway and have shown mixed outcome, largely due to development of resistances and increased tumour aggressiveness. In this study, we used the zebrafish model to screen for novel anti-angiogenic molecules from a library of compounds derived from natural products. From this, we identified canthin-6-one, an indole alkaloid, which inhibited zebrafish intersegmental vessel (ISV) and sub-intestinal vessel development. Further characterisation revealed that treatment of canthin-6-one reduced ISV endothelial cell number and inhibited proliferation of human umbilical vein endothelial cells (HUVECs), suggesting that canthin-6-one inhibits endothelial cell proliferation. Of note, canthin-6-one did not inhibit VEGFA-induced phosphorylation of VEGFR2 in HUVECs and downstream phosphorylation of extracellular signal-regulated kinase (Erk) in leading ISV endothelial cells in zebrafish, suggesting that canthin-6-one inhibits angiogenesis independent of the VEGFA/VEGFR2 pathway. Importantly, we found that canthin-6-one impairs tumour-associated angiogenesis in a zebrafish B16F10 melanoma cell xenograft model and synergises with VEGFR inhibitor sunitinib malate to inhibit developmental angiogenesis. In summary, we showed that canthin-6-one exhibits anti-angiogenic properties in both developmental and pathological contexts in zebrafish, independent of the VEGFA/VEGFR2 pathway and demonstrate that canthin-6-one may hold value for further development as a novel anti-angiogenic drug.

Inhibiting anti-angiogenic VEGF165b activates a miR-17-20a-Calcipressin-3 pathway that revascularizes ischemic muscle in peripheral artery disease

BackgroundVEGF165a increases the expression of the microRNA-17-92 cluster, promoting developmental, retinal, and tumor angiogenesis. We have previously shown that VEGF165b, an alternatively spliced anti-angiogenic VEGF-A isoform, inhibits the VEGFR-STAT3 pathway in ischemic endothelial cells (ECs) to decrease their angiogenic capacity. In ischemic macrophages (Møs), VEGF165b inhibits VEGFR1 to induce S100A8/A9 expression, which drives M1-like polarization. Our current study aims to determine whether VEGF165b inhibition promotes perfusion recovery by regulating the microRNA(miR)-17-92 cluster in preclinical PAD.MethodsFemoral artery ligation and resection was used as a preclinical PAD model. Hypoxia serum starvation (HSS) was used as an in vitro PAD model. VEGF165b was inhibited/neutralized by an isoform-specific VEGF165b antibody.ResultsHere, we show that VEGF165b-inhibition induces the expression of miR-17-20a (within miR-17-92 (miR-17-18a-19a-19b-20a-92) cluster) in HSS-ECs and HSS-Møs vs. respective normal and/or isotype-matched IgG controls to enhance perfusion recovery. Consistent with the bioinformatics analysis that revealed RCAN3 as a common target of miR-17 and miR-20a, Argonaute-2 pull-down assays showed decreased miR-17-20a expression and higher RCAN3 expression in the RNA-induced silencing complex of HSS-ECs and HSS-Møs vs. respective controls. Inhibiting miR-17-20a induced RCAN3 levels to decrease ischemic angiogenesis and promoted M1-like polarization to impair perfusion recovery. Finally, using STAT3 inhibitors, S100A8/A9 silencers, and VEGFR1-deficient ECs and Møs, we show that VEGF165b-inhibition activates the miR-17-20a-RCAN3 pathway independent of VEGFR1-STAT3 or VEGFR1-S100A8/A9 in ischemic-ECs and ischemic-Møs respectively.ConclusionsOur data revealed a hereunto unrecognized therapeutic ‘miR-17-20a-RCAN3’ pathway in the ischemic vasculature that is VEGFR1-STAT3/S100A8/A9 independent and is activated only upon VEGF165b-inhibition in PAD.

SUMOylation Fine-Tunes Endothelial HEY1 in the Regulation of Angiogenesis.

Angiogenesis, which plays a critical role in embryonic development and tissue repair, is controlled by a set of angiogenic signaling pathways. As a TF (transcription factor) belonging to the basic helix-loop-helix family, HEY (hairy/enhancer of split related with YRPW motif)-1 (YRPW motif, abbreviation of 4 highly conserved amino acids in the motif) has been identified as a key player in developmental angiogenesis. However, the precise mechanisms underlying HEY1's actions in angiogenesis remain largely unknown. Our previous studies have suggested a potential role for posttranslational SUMOylation in the dynamic regulation of vascular development and organization. Immunoprecipitation, mass spectrometry, and bioinformatics analysis were used to determine the biochemical characteristics of HEY1 SUMOylation. The promoter-binding capability of HEY1 was determined by chromatin immunoprecipitation, dual luciferase, and electrophoretic mobility shift assays. The dimerization pattern of HEY1 was determined by coimmunoprecipitation. The angiogenic capabilities of endothelial cells were assessed by CCK-8 (cell counting kit-8), 5-ethynyl-2-deoxyuridine staining, wound healing, transwell, and sprouting assays. Embryonic and postnatal vascular growth in mouse tissues, matrigel plug assay, cutaneous wound healing model, oxygen-induced retinopathy model, and tumor angiogenesis model were used to investigate the angiogenesis in vivo. We identified intrinsic endothelial HEY1 SUMOylation at conserved lysines by TRIM28 (tripartite motif containing 28) as the unique E3 ligase. Functionally, SUMOylation facilitated HEY1-mediated suppression of angiogenic RTK (receptor tyrosine kinase) signaling and angiogenesis in primary human endothelial cells and mice with endothelial cell-specific expression of wild-type HEY1 or a SUMOylation-deficient HEY1 mutant. Mechanistically, SUMOylation facilitates HEY1 homodimer formation, which in turn preserves HEY1's DNA-binding capability via recognition of E-box promoter elements. Therefore, SUMOylation maintains HEY1's function as a repressive TF controlling numerous angiogenic genes, including RTKs and Notch pathway components. Proangiogenic stimuli induce HEY1 deSUMOylation, leading to heterodimerization of HEY1 with HES (hairy and enhancer of split)-1, which results in ineffective DNA binding and loss of HEY1's angiogenesis-suppressive activity. Our findings demonstrate that reversible HEY1 SUMOylation is a molecular mechanism that coordinates endothelial angiogenic signaling and angiogenesis, both in physiological and pathological milieus, by fine-tuning the transcriptional activity of HEY1. Specifically, SUMOylation facilitates the formation of the HEY1 transcriptional complex and enhances its DNA-binding capability in endothelial cells.

Extending Peripheral Retinal Vascularization in Retinopathy of Prematurity Through Regulation of VEGF Signaling

Experimental studies provide evidence that regulation of VEGF receptor-2 signaling in endothelial cells orders cell divisions and extends developmental angiogenesis while inhibiting pathologic intravitreal angiogenesis and has relevance to retinopathy of prematurity (ROP). We tested the hypothesis that intravitreal anti-VEGF would extend vascularization into peripheral avascular retina in human type 1 ROP compared with controls. Retrospective, nonrandomized treatment comparison. The study was conducted at an academic institution, with the study population comprising all premature infants screened for ROP from January 2019 through December 2022. The experimental group included type 1 ROP treated with bilateral bevacizumab (0.25 mg) and had adequate fundus imaging by a certified ophthalmic photographer at 2 examinations: within 2 weeks of treatment and 1-3 weeks later. A control group included gestational age- and birthweight-matched infants with ROP less severe than type 1 ROP. The main outcome measure was extent of temporal retinal vasculature measured by a masked analyst between treated and control eyes. Paired and nonpaired t tests were used. Of 382 screened infants, 34 developed type 1 ROP; 11 comprised the experimental group and 11 the control group. At baseline, there was a trend toward shorter temporal vascular extent in treatment compared with control groups (3667±547 vs 4262±937 pixels, 95% CI -1277, 88; P = .084) but no difference between groups at follow-up (P = .945). Vascular extension was significantly greater in the treatment than control (872±521 vs 253±151 pixels, 95% CI 262, 977; P = .003), showing catch-up growth. This clinical evidence supports laboratory-based studies that regulation of VEGF using an intravitreal anti-VEGF agent increases developmental angiogenesis into the peripheral avascular retina while inhibiting pathologic intravitreal angiogenesis in ROP.

Differentially expressed genes in orbital adipose/connective tissue of thyroid-associated orbitopathy.

Thyroid-associated orbitopathy (TAO) is a disease associated with autoimmune thyroid disorders and it can lead to proptosis, diplopia, and vision-threatening compressive optic neuropathy. To comprehensively understand the molecular mechanisms underlying orbital adipogenesis in TAO, we characterize the intrinsic molecular properties of orbital adipose/connective tissue from patients with TAO and control individuals. RNA sequencing analysis (RNA-seq) was performed to measure the gene expression of orbital adipose/connective tissues of TAO patients. Differentially expressed genes (DEGs) were detected and analyzed through Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, and Gene Set Enrichment Analysis (GSEA). The protein-protein interaction (PPI) network was constructed using the STRING database, and hub genes were identified by the Cytoscape plug-in, cytoHubba. We validated several top DEGs through quantitative real-time polymerase chain reaction (qRT-PCR). We identified 183 DEGs in adipose tissue between TAO patients (n=3) and control patients (n=3) through RNA sequencing, including 114 upregulated genes and 69 downregulated genes. The PPI network of these DEGs had 202 nodes and 743 edges. PCR-based validation results of orbital adipose tissue showed multiple top-ranked genes in TAO patients (n=4) are immune and inflammatory response genes compared with the control individual (n=4). They include ceruloplasmin isoform x3 (CP), alkaline tissue-nonspecific isozyme isoform x1 (ALPL), and angiotensinogen (AGT), which were overrepresented by 2.27- to 6.40-fold. Meanwhile, protein mab-21-like 1 (MAB21L1), phosphoinositide 3-kinase gamma-subunit (PIK3C2G), and clavesin-2 (CLVS2) decreased by 2.6% to 32.8%. R-spondin 1 (RSPO1), which is related to oogonia differentiation and developmental angiogenesis, was significantly downregulated in the orbital muscle tissues of patients with TAO compared with the control groups (P=0.024). Our results suggest that there are genetic differences in orbital adipose-connective tissues derived from TAO patients. The upregulation of the inflammatory response in orbital fat of TAO may be consistent with the clinical phenotype like eyelid edema, exophthalmos, and excess tearing. Downregulation of MAB21L1, PIK3C2G, and CLVS2 in TAO tissue demonstrates dysregulation of differentiation, oxidative stress, and developmental pathways.

Mutation of key signaling regulators of cerebrovascular development in vein of Galen malformations

To elucidate the pathogenesis of vein of Galen malformations (VOGMs), the most common and most severe of congenital brain arteriovenous malformations, we performed an integrated analysis of 310 VOGM proband-family exomes and 336,326 human cerebrovasculature single-cell transcriptomes. We found the Ras suppressor p120 RasGAP (RASA1) harbored a genome-wide significant burden of loss-of-function de novo variants (2042.5-fold, p = 4.79 x 10−7). Rare, damaging transmitted variants were enriched in Ephrin receptor-B4 (EPHB4) (17.5-fold, p = 1.22 x 10−5), which cooperates with p120 RasGAP to regulate vascular development. Additional probands had damaging variants in ACVRL1, NOTCH1, ITGB1, and PTPN11. ACVRL1 variants were also identified in a multi-generational VOGM pedigree. Integrative genomic analysis defined developing endothelial cells as a likely spatio-temporal locus of VOGM pathophysiology. Mice expressing a VOGM-specific EPHB4 kinase-domain missense variant (Phe867Leu) exhibited disrupted developmental angiogenesis and impaired hierarchical development of arterial-capillary-venous networks, but only in the presence of a “second-hit” allele. These results illuminate human arterio-venous development and VOGM pathobiology and have implications for patients and their families.

ScDiffCom: a tool for differential analysis of cell–cell interactions provides a mouse atlas of aging changes in intercellular communication

Dysregulation of intercellular communication is a hallmark of aging. To better quantify and explore changes in intercellular communication, we present scDiffCom and scAgeCom. scDiffCom is an R package, relying on approximately 5,000 curated ligand–receptor interactions, that performs differential intercellular communication analysis between two conditions from single-cell transcriptomics data. Built upon scDiffCom, scAgeCom is an atlas of age-related cell–cell communication changes covering 23 mouse tissues from 58 single-cell RNA sequencing datasets from Tabula Muris Senis and the Calico murine aging cell atlas. It offers a comprehensive resource of tissue-specific and sex-specific aging dysregulations and highlights age-related intercellular communication changes widespread across the whole body, such as the upregulation of immune system processes and inflammation, the downregulation of developmental processes, angiogenesis and extracellular matrix organization and the deregulation of lipid metabolism. Our analysis emphasizes the relevance of the specific ligands, receptors and cell types regulating these processes. The atlas is available online (https://scagecom.org).

Retinoic acid alters metalloproteinase action in red deer antler stem cells.

Metalloproteinases (MMP)s regulate developmental processes, control angiogenesis and wound healing, participate in the formation of immune receptors, and are expressed in stem cells. Retinoic acid (RA) is a potential modulator of these proteinases. The aim was to determine (1) MMPs' action in antler stem cells (ASCs) before and after differentiation into adipo-, osteo-, and chondrocytes and (2) the effect of RA on modifying MMP action in ASCs. Antler tissue from pedicle was collected approximately 40 days after antler casting, post mortem from healthy breeding five year old males (N = 7). The cells were isolated from the pedicle layer of periosteum after skin separation and cultured. The pluripotency of the ASCs was evaluated by mRNA expression for NANOG, SOX2, and OCT4. ASCs were stimulated with RA (100nM) and differentiated for 14 days. The MMP (1-3) and TIMP(1-3) (tissue inhibitor of MMPs) mRNA expression was determined in the ASCs, their concentrations in the ASCs and the medium after RA stimulation as well as profiles of mRNA expression for MMPs: 1-3 and TIMPs: 1-3 during differentiation of ASC to osteocytes, adipocytes and chondrocytes. RA increased MMP-3 and TIMP-3 mRNA expression and output (P < 0.05) and not influenced on MMP-1 and TIMP-1 mRNA expression and output in ASC (P > 0.05). Depending on differentiation of ASC to osteocytes, adipocytes or chondrocytes, MMPs`and TIMPs`expression profile fluctuates for all studied proteases and its inhibitors. The studies demand continuation considering the role of proteases in stem cells physiology and differentiation. The results may be relevant for the study of cellular processes during the cancerogenesis of tumor stem cells.

The role of blood flow in vessel remodeling and its regulatory mechanism during developmental angiogenesis.

Vessel remodeling is essential for a functional and mature vascular network. According to the difference in endothelial cell (EC) behavior, we classified vessel remodeling into vessel pruning, vessel regression and vessel fusion. Vessel remodeling has been proven in various organs and species, such as the brain vasculature, subintestinal veins (SIVs), and caudal vein (CV) in zebrafish and yolk sac vessels, retina, and hyaloid vessels in mice. ECs and periendothelial cells (such as pericytes and astrocytes) contribute to vessel remodeling. EC junction remodeling and actin cytoskeleton dynamic rearrangement are indispensable for vessel pruning. More importantly, blood flow has a vital role in vessel remodeling. In recent studies, several mechanosensors, such as integrins, platelet endothelial cell adhesion molecule-1 (PECAM-1)/vascular endothelial cell (VE-cadherin)/vascular endothelial growth factor receptor 2 (VEGFR2) complex, and notch1, have been shown to contribute to mechanotransduction and vessel remodeling. In this review, we highlight the current knowledge of vessel remodeling in mouse and zebrafish models. We further underline the contribution of cellular behavior and periendothelial cells to vessel remodeling. Finally, we discuss the mechanosensory complex in ECs and the molecular mechanisms responsible for vessel remodeling.

Mesenchymal Stromal Cells Facilitate Tip Cell Fusion Downstream of BMP-Mediated Venous Angiogenesis-Brief Report.

The goal of this study was to identify and characterize cell-cell interactions that facilitate endothelial tip cell fusion downstream of BMP (bone morphogenic protein)-mediated venous plexus formation. High resolution and time-lapse imaging of transgenic reporter lines and loss-of-function studies were carried out to study the involvement of mesenchymal stromal cells during venous angiogenesis. BMP-responsive stromal cells facilitate timely and precise fusion of venous tip cells during developmental angiogenesis. Stromal cells are required for anastomosis of venous tip cells in the embryonic caudal hematopoietic tissue.

ZFP36-mediated mRNA decay regulates metabolism

Cellular metabolism is tightly regulated by growth factor signaling, which promotes metabolic rewiring to support growth and proliferation. While growth factor-induced transcriptional and post-translational modes of metabolic regulation have been well defined, whether post-transcriptional mechanisms impacting mRNA stability regulate this process is less clear. Here, we present the ZFP36/L1/L2 family of RNA-binding proteins and mRNA decay factors as key drivers of metabolic regulation downstream of acute growth factor signaling. We quantitatively catalog metabolic enzyme and nutrient transporter mRNAs directly bound by ZFP36 following growth factor stimulation-many of which encode rate-limiting steps in metabolic pathways. Further, we show that ZFP36 directly promotes the mRNA decay of Enolase 2 (Eno2), altering Eno2 protein expression and enzymatic activity, and provide evidence of a ZFP36/Eno2 axis during VEGF-stimulated developmental retinal angiogenesis. Thus, ZFP36-mediated mRNA decay serves as an important mode of metabolic regulation downstream of growth factor signaling within dynamic cell and tissue states.

MicroRNA-216a is essential for cardiac angiogenesis

While it is experimentally supported that impaired myocardial vascularization contributes to a mismatch between myocardial oxygen demand and supply, a mechanistic basis for disruption of coordinated tissue growth and angiogenesis in heart failure remains poorly understood. Silencing strategies that impair microRNA biogenesis have firmly implicated microRNAs in the regulation of angiogenesis, and individual microRNAs prove to be crucial in developmental or tumor angiogenesis. A high-throughput functional screening for the analysis of a whole-genome microRNA silencing library with regard to their phenotypic effect on endothelial cell proliferation as a key parameter, revealed several anti- and pro-proliferative microRNAs. Among those was miR-216a, a pro-angiogenic microRNA which is enriched in cardiac microvascular endothelial cells and reduced in expression under cardiac stress conditions. miR-216a null mice display dramatic cardiac phenotypes related to impaired myocardial vascularization and unbalanced autophagy and inflammation, supporting a model where microRNA regulation of microvascularization impacts the cardiac response to stress.