Angiogenesis

Somatic activating mutations in KRAS are found in the endothelium of the majority of sporadic brain arteriovenous malformations (bAVMs), yet the consequences of oncogenic KRAS signaling in endothelial cells during active vessel morphogenesis remain incompletely characterized. We expressed KRASG12V in primary human umbilical vein endothelial cells using a doxycycline-inducible lentiviral system and examined morphogenic behavior, proliferation, migration, and transcriptional output in a three-dimensional planar co-culture angiogenesis assay. KRASG12V-expressing cells failed to organize into vessel-like networks, instead forming compact sheet-like structures that persisted through day 12. A transient proliferative phase at days 3-5 resolved to control levels by day 12, consistent with preserved sensitivity to contact inhibition rather than unrestricted growth. Enhanced migration at day 5 was accompanied by upregulation of a focal adhesion and matrix remodeling program centered on ITGB3, PLAU, PLAUR, and PIK3CG. Translating ribosome-affinity purification sequencing (TRAP-seq) of the EC-specific translatome across four independent donor pools revealed progressive acquisition of an AVM-associated transcriptional identity by day 12, including upregulation of ACVRL1, ENG, JAG1, NOTCH1, ANGPT2, and TEK, with concordance to human bAVM nidus endothelium at both the gene and pathway level. Pharmacological inhibition with Alpelisib (PI3K), Trametinib (MEK), and Pazopanib (VEGFR2) demonstrated that PI3K is the principal organizer of the morphogenic phenotype. These findings characterize a KRASG12V-driven program in endothelial cells that recapitulates core transcriptional features of bAVM endothelium in a primary cell model.

Infantile hemangioma (IH), the most common benign vascular tumor in infants, is characterized by the aberrant proliferation of hemangioma endothelial cells (HemECs) and excessive angiogenesis. However, it is unclear how the tumor microenvironment sustains these active angiogenic signals. We performed single-cell RNA sequencing and spatial transcriptomic to profile 30 samples from proliferating IH, involuting IH, and normal skin. The results indicated that HemECs comprised distinct subpopulations. Among them, capillary endothelial cells (Cap.ECs) had an immature phenotype and functioned as the core drivers of IH. The endothelial-to-mesenchymal transition endothelial cells (endoMT-ECs) secreted collagen and engaged in CD44-mediated crosstalk with Cap.ECs to construct the angiogenesis-active niche in the tumor microenvironment (TME) of proliferating IH. Blockade of CD44-mediated intercellular communication effectively suppresses the angiogenesis of HemECs and promotes tumor regression. Beyond the endothelial layer, pericytes tightly enveloped Cap.ECs, with SERPINE2+ subtypes sustaining their angiogenic activity via ANGPT signaling. Pseudotime trajectory analysis revealed that the mesenchymal stem cells (MSCs)-enriched niche serves as a progenitor pool for Cap.ECs in proliferating IH. Our findings thus revealed that IH progression relies on CD44-mediated endoMT-EC-Cap.EC crosstalk and on both pericyte and MSC niches to sustain angiogenesis. HIGHLIGHTSO_LICapillary endothelial cells drive infantile hemangioma progression, with pericytes tightly ensheathing them. C_LIO_LICD44-mediated capillary and endoMT-derived endothelial cells crosstalk construct the angiogenic-active niche in the tumor microenvironment of proliferating infantile hemangioma. C_LIO_LIThe MSC-enriched niche provides a progenitor pool for the capillary endothelial cells in proliferating infantile hemangioma. C_LI

The Hippo signaling pathway is increasingly recognized as a key regulator of endothelial cell (EC) proliferation, migration and vascular development. However, the roles of its upstream scaffold proteins remain poorly understood. Although WWC family proteins are widely regarded as functionally redundant activators of LATS1/2 kinases, the human genome contains a third family member, WWC3, that is absent from mice, raising the possibility of species-specific regulation of endothelial Hippo signaling. Here, we assessed the roles of WWC2 and WWC3 in human ECs using siRNA-mediated knockdown. Surprisingly, we found that WWC3 is the predominant regulator of canonical Hippo signaling, with a substantially greater effect than WWC2 on LATS1/2 phosphorylation, YAP/TAZ localization and expression of Hippo target genes. Loss of WWC3 also altered endothelial morphology and induced a partial endothelial-to-mesenchymal transition-like (EndoMT-like) phenotype. By contrast, WWC2 had a lesser effect on canonical Hippo signaling, but it was required for normal VEGF signaling dynamics. Despite these distinct molecular functions, depletion of either WWC2 or WWC3 impaired EC proliferation, migration, and cord formation in vitro. Together, our findings demonstrate that WWC family proteins perform overlapping but distinct functions in human ECs, with WWC3 acting as the predominant canonical Hippo regulator, whereas WWC2 more efficiently modulates VEGF signaling. These results reveal unexpected functional specialization among WWC proteins and suggest that regulation of Hippo signaling in human ECs differs from that inferred from mouse studies.

ANGPT2 is widely recognized as a critical regulator of pathological neovascularization. By analyzing scRNA-seq data from neonatal retinas, we demonstrate that Angpt2 transcripts are highly enriched in tip cells relative to other endothelial subtypes, where Angpt1/4 expression is absent. However, mechanisms underlying ANGPT2 function at angiogenic fronts remain inadequately understood. Here, we show that endothelial Angpt2 deletion severely disrupted retinal vascularization, characterized by neovascular tufts and micro-hemorrhage. Similar angiogenic defects also occurred in the brain, but were less evident in other tissues examined. Mechanistically, ANGPT2 insufficiency attenuated retinal tip cell invasion with aberrant mural cell coverage, compromising sprouting into non-vascularized tissues. Retinal RNA-seq analysis revealed that transcripts associated with endothelial migration and junction assembly were reduced in Angpt2 mutants compared to littermate controls, while upregulated genes were enriched in hypoxia-responsive pathways and mural cell development. Notably, abnormal mural-tip cell associations were detected within 48 hours post-Angpt2 deletion, displaying also a hypoxia-driven transcriptomic signature. These closely resemble the vascular pathologies observed in human retinopathy of prematurity. In contrast, Angpt1 insufficiency or Angpt4 deficiency primarily affected venous morphogenesis. Collectively, our findings imply that ANGPT2 is essential for driving tip cell invasion during sprouting angiogenesis, and that its insufficiency triggers hypoxia-driven vascular anomalies.

Tumstatin and Endostatin are endogenous extracellular matrix-derived inhibitors of angiogenesis generated from the non-collagenous domains of type IV and type XVIII collagens, respectively. Although both molecules suppress angiogenesis and tumor growth in vivo, previous studies demonstrated that they engage distinct endothelial integrin receptors and activate different intracellular signaling pathways. In particular, Tumstatin inhibits endothelial proliferation through suppression of the focal adhesion kinase (FAK)/phosphatidylinositol 3-kinase (PI3K)/Akt/mTOR pathway, whereas Endostatin primarily inhibits endothelial migration through 5{beta}1 integrin-dependent signaling. Here we evaluated a key mechanistic distinction between these two angiogenesis inhibitors by examining Akt phosphorylation in human umbilical vein endothelial cells (HUVEC) cultured on fibronectin. Validating previous reports, recombinant human Tumstatin reduced Akt phosphorylation whereas recombinant human Endostatin did not alter Akt activation. These findings confirm a defining feature of Tumstatin signaling and reinforce the concept that collagen-derived angiogenesis inhibitors regulate endothelial cell behavior through distinct integrin-dependent mechanisms.

Background: Histidine-rich glycoprotein (HRG) is a regulator of coagulation that has been linked to experimental thrombosis, but its causal role in human thrombotic disease remains unclear. Objectives: To determine whether HRG has a role in thrombosis, we evaluated the association between genetically determined HRG variation and thrombosis risk using Mendelian randomisation (MR). Methods: We performed a two sample MR analysis in UK Biobank using non-overlapping samples. Separate genome wide association studies (GWAS) were conducted to identify single nucleotide polymorphisms (SNPs) associated with circulating HRG protein levels and to estimate SNP associations with thrombosis outcomes. Genetic instruments were derived from the HRG GWAS measurements and applied to assess associations with overall, venous, and arterial thrombosis. Sensitivity analyses using multiple MR methods were undertaken, alongside adjusted logistic regression models in participants with measured HRG levels. Results: Among 30,680 participants with HRG measurements, GWAS identified multiple loci associated with HRG levels, with the strongest signal at the rs9898 SNP ({beta} =0.52; P=1.1x10-306). Single instrument MR found no association between HRG protein levels predicted by the rs9898 SNP and risk of overall thrombosis ({beta} =-0.00660; P=0.622), venous thrombosis ({beta} =0.0101; P=0.650) and arterial thrombosis ({beta} =-0.0137; P=0.384). Similar null findings were observed using multi-instrument MR approaches. In complementary analyses, measured HRG levels were not associated with thrombosis after adjustment for age, sex, and C reactive protein, and results were unchanged after stratification by rs9898 genotype. Conclusion: Genetically determined variation in HRG is not associated with thrombotic risk, indicating that HRG related coagulation phenotypes do not translate into clinically meaningful thrombosis.

Mural cells are key regulators of vascular architecture, yet how their contractility and morphology jointly influence vessel diameter in vivo remains poorly understood. Here, using high-resolution live imaging and single-cell morphometric analysis in zebrafish, we show that vascular smooth muscle cells (vSMCs) and pericytes undergo a developmental reduction in cell size through the progressive retraction of actin-rich primary and secondary processes during vascular remodelling. By specifically manipulating RhoA activity to alter mural cell contractility and shape, we uncover vessel-specific roles for mural cells in diameter regulation. We find that vSMC contractility is dispensable for the initial constriction of the dorsal aorta but is required to stabilise its diameter following vasoconstriction and to maintain vascular tone. In contrast, pericyte contractility is dispensable for both the constriction and stabilisation of intersegmental vessels. In the brain vasculature, vessel diameter is governed by the balance between contractile force and the extent of vessel coverage by vSMCs. Together, our findings redefine the role of mural cell contractility in the control of vessel constraint, demonstrating that morphology and vessel coverage, rather than contractility alone, are key determinants of vascular diameter in vivo.

Tumors commonly exhibit a leaky vasculature and innate and/or adaptive immune cell infiltration. A recent study showed that a non-leaky and normalized mouse breast cancer vasculature after conditional deletion of the Shb gene in endothelial cells (EC) displayed a suppressive immune cell profile. To extend those findings further, regulation of immunomodulatory gene expression of normalized breast cancer EC was related to healthy human breast and human breast cancer EC gene expression. A considerable correlation was observed in the gene expression profiles of immunoregulatory genes between normal breast arterial EC and normalized (Shb-deficient) EC, and these changes did not primarily seem to involve gene products forming conduits for tumor leukocyte transendothelial migration but other aspects of immune suppression as for example cytokine expression. In addition to breast cancer, healthy lung and colonic EC exhibited an immune suppressive phenotype compared to their lung cancer and colorectal cancer counterparts. It is concluded that normalized EC may confer a non-leaky and immune suppressed phenotype, an insight that may be exploited to enhance the efficacy of immunotherapy.

Abstract Purpose: To expand the clinical and mechanistic understanding of the +57C>T seed-region mutation in miR-184 causing EDICT (endothelial dystrophy, iris hypoplasia, congenital cataract, and stromal thinning) syndrome. Design: Cross-sectional analysis and laboratory confirmation Participants: 18 members of a four-generation family with known +57C>T miR-184 status Methods: We used optical biometry, corneal topography, and medical history to characterize the clinical phenotype. Carrier effects on ocular biometric measurements were estimated using polygenic linear mixed models incorporating a pedigree-derived kinship matrix, adjusted for age and sex. Patient-derived and control induced pluripotent stem cells (iPSCs) were generated and differentiated into corneal endothelial cells (CECs). Main Outcome Measures: Axial length, keratometry (in diopters), white-to-white corneal diameter, topography mapping, central and peripheral corneal thickness, and history of retinal detachment or corneal transplant were compared between mutation carriers and noncarriers, adjusting for age and sex. Cellular analysis was conducted with immunostaining (ZO-1, ATP1A1), morphometric quantification, qRT-PCR of endothelial markers, and transendothelial electrical resistance (TEER). Results: 10 of 18 family members were heterozygous for +57C>T, with retinal detachment occurring in 5/10 affected individuals compared to 0/8 unaffected individuals (p=0.04). Affected eyes had 2.2 mm shorter axial length (p=0.02), 9.3 D steeper mean keratometry (p=0.004), 1.6 mm smaller horizontal corneal diameter (p=0.0001), and 139-micrometer thinner central corneas (p=0.003). Mutant iPSC-derived CECs were associated with irregular borders, increased cell and nucleus area, widened intercellular gaps, disrupted ATP1A1 membrane localization, and reduced barrier function on TEER (all p<0.05). Gene expression analysis showed downregulation of COL4A1, COL4A3, and AQP1 with upregulation of COL8A1. Conclusions: The miR-184 +57C>T mutation produces a broad ocular phenotype that includes smaller, thinner corneas and microphthalmia. Mechanistically, it disrupts CEC junctional integrity, extracellular matrix and pump-related genes, supporting a role for miR-184 in coordinated anterior-posterior eye morphogenesis.

Bradycardia occurs when the heart rate is lower than normal resulting in reduced cerebral blood flow and contributing to neurodegeneration in adults but how it affects embryonic cerebrovascular development is not well studied. We induce bradycardia by targeting the heart pacemaker channel Hcn4 via chemical (ivabradine) and genetic (hcn4 mutant) methods. Bradycardia results in reduced brain vessel diameter and mural cell (pericyte and vascular smooth muscle cell) number. Endothelial cells are the first responders in sensing changes in blood flow, and we show that signalling through the canonical endothelial-autonomous mechanosensitive pathway (Piezo1, Mek5, Erk5, Klf2) is reduced in bradycardia. To identify the ligand-receptor combination that transmits signals to developing mural cells, we show that expression of the Notch ligand jagged2b is decreased in the brain of both hcn4 and klf2 mutants. jag2b knockdown reduces mural cell numbers in brain vessels. Restoring jag2b levels increases mural cell numbers in both wildtype and hcn4 mutants. Our work connects bradycardia, mechanosensitive signaling and mural cell recruitment demonstrating that mural cell numbers can be increased in bradycardia by restoring Notch signalling via upregulating endothelial Jag2b. SummaryBradycardia models show reduced blood flow, Piezo1-klf2-jag2b-notch3 mechanosensing and mural cell recruitment to developing brain vasculature. Restoration of jag2, an endogenous endothelial cell ligand, rescues mural cell numbers in bradycardia mutants.

Pathogenic variants in COL3A1 cause Vascular Ehlers-Danlos syndrome (vEDS), a rare connective tissue disorder characterised by vascular fragility, increasing the risk of arterial ruptures/dissection. Advances in genomic sequencing have led to an increasing number of COL3A1 variants where the clinical significance is unclear, with these being termed variants of uncertain significance (VUS). VUS creates challenges for diagnosis and clinical management. Thus major efforts have been made to reclassify these to either pathogenic or benign variants in disease causality. Functional data from model systems can provide significant evidence to clinicians on the pathogenicity of a variant. To address the increasing numbers of VUS in COL3A1, we developed a fast pipeline using F0 crispant zebrafish to provide functional evidence for variant classification despite there being no direct orthologue of COL3A1 in zebrafish. Loss of col5a1 resulted in cardiac defects, dysmorphic blood vessel structures and delayed angiogenic sprouting. Trunk haemorrhage prevalence under physical stress increased in col5a1 knockout zebrafish, recapitulating vEDS patients. Remarkably, co-injection of F0 col5a1 knockout crispants with human wildtype COL3A1 mRNA partially rescued cardiac and vascular phenotypes, indicating a level of functional conservation between zebrafish type V and human type III collagen. These findings establish a tractable in vivo platform for functional assessment of COL3A1 VUS. Phenotypic rescue with wildtype COL3A1 provides a benchmark against which the pathogenicity of variants can be evaluated, generating functional evidence for VUS reclassification. Our model provides both a valuable tool for investigating vEDS disease mechanisms and a clinically relevant platform to improve diagnoses for patients with suspected vEDS.

Background/Objective: Diabetic macular oedema (DMO) is a leading cause for visual impairment primarily managed with intravitreal anti-VEGF agents such as ranibizumab (RBZ). Levosulpiride (LSP), a prokinetic medication, was recently repositioned as a safe oral treatment for naive DMO. Here, we investigated the adjuvant effect of oral LSP in combination with intravitreal RBZ injections for treating persistent DMO. Subjects/Methods: Double-blinded, dual-centre, phase 2 trial in patients with centre-involving DMO randomly assigned to be orally treated with placebo (15 patients, 18 eyes) or LSP (18 patients, 19 eyes) along with 3 successive (4 weeks apart) RBZ intravitreal injections and a 24-week follow-up. Results: Baseline best-corrected visual acuity (BCVA) improved (p[&le;]0.04) at week 12 in both RBZ+placebo and RBZ+LSP, but improvement was maintained (p=0.009) at week 24 only in RBZ+LSP. In agreement, longitudinal changes from baseline in BCVA from weeks 12 to 24 defined superior (p=0.02) visual gains measured by the Area Under the Curve (AUC) in RBZ+LSP vs. RBZ+placebo. The baseline value of mean central foveal thickness (CFT) decreased (p[&le;]0.002) in both groups at week 12 and CFT reduction was significant (p=0.006) at week 24 only in RBZ+LSP. Also, longitudinal changes from baseline in CFT resulted in a higher AUC reduction (p[&le;]0.04) at weeks 4 to12 in RBZ+LSP vs. RBZ+placebo. No significant adverse side effects were detected. Conclusions: Adjunctive LSP showed functional and anatomical benefits over the first-line therapy with RBZ. Adjuvant properties may involve the LSP-induced intraocular upregulation and downregulation of vasoinhibin and VEGF, respectively. Larger clinical trials are warranted.

Anti-angiogenic cancer therapies attempt to withhold necessary nutrients and oxygen from growing tumors by targeting the major promoters of endothelial cell (EC) angiogenesis: vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR-2). Unfortunately, these treatments are often insufficient, even when coupled with chemotherapies, and fail to significantly increase survival rates. The tumor microenvironment (TME) is mechanically distinct compared to normal tissue, including increased matrix deformations or strains caused by cancer-associated fibroblasts (CAFs). In this report, we detail the specific and independent roles of two tyrosine residues, Y1054 and Y1214, on mechanical activation of VEGFR-2. Furthermore, we characterize CAF biochemical and mechanical signaling and demonstrate how ECs exhibit decreased vessel growth when co-cultured with CAFs and treated with a contractility inhibitor. Using non-phosphorylatable VEGFR-2 mutants, we reveal Y1054 and Y1214 are each necessary for EC angiogenesis, particularly in response to strain. Overall, this research highlights the need to study how mechanics in the TME promote vessel growth and thus tumor progression, which is important to consider when developing future anti-angiogenic therapies.

Preeclampsia is a major pregnancy complication driven by placental dysfunction, yet research is limited by reliance on patient-derived tissues and models that do not fully capture human disease. Here, we develop a genetically engineered human trophoblast organoid model of preeclampsia that can be generated without access to placental tissue. Using a CRISPR-based Prime Integrase strategy, we engineered induced trophoblast stem cells to express the preeclampsia-associated soluble fms-like tyrosine kinase-1 (sFlt-1) exon 15a isoform. Engineered organoids showed a transcriptional shift towards primary preeclamptic placentae and developed several features of disease. These included reduced PlGF, increased IL-6 and soluble endoglin, oxidative stress, impaired growth and an elevated sFlt-1/PlGF ratio comparable to primary preeclamptic trophoblast organoids. These broader changes were not reproduced by adding recombinant human sFlt-1 to control organoids. Conditioned media from engineered organoids impaired endothelial network formation, demonstrating a functional effect of the altered trophoblast secretome. Treatment with sulfasalazine and metformin also restored angiogenic balance and organoid growth. Together, these findings establish a tractable human model that reproduces molecular and functional features of preeclampsia and provides a platform to study disease mechanisms and test potential therapies.

Purpose: To investigate corneal subbasal nerve plexus alterations using in vivo corneal confocal microscopy (IVCM) in patients with Autosomal Recessive Spastic Ataxia of Charlevoix-Saguenay (ARSACS) and Spastic Paraplegia Type 7 (SPG7). Methods: This cross-sectional pilot study included eight ARSACS patients, five SPG7 patients, and twenty age- and sex-matched healthy controls. All participants underwent neurological and ophthalmological examination followed by central corneal imaging using IVCM. Quantitative corneal nerve parameters were analyzed with automated software, and correlations with clinical severity scales were assessed. Results: The mean age was 34.2 +/- 3.4 years in controls, 34.5 +/- 0.7 years in the ARSACS group, and 38.2 +/- 3.5 years in the SPG7 group. Corneal nerve branch density (CNBD) and corneal nerve total branch density (CTBD) were significantly lower in ARSACS and SPG7 patients compared with healthy controls. CNFD, CNFL, CNFA, CNFW, and CNFrD were lower in ARSACS and SPG7 patients compared with healthy controls; however, these differences did not reach statistical significance. No statistically significant differences in IVCM parameters were detected between ARSACS and SPG7 patients. Spearman correlation analysis did not show significant correlations between corneal nerve parameters and FARS, SARA, ADL scores, or disease duration. Conclusion: IVCM revealed reduced corneal nerve branching parameters in patients with ARSACS and SPG7. These findings indicate involvement of the corneal subbasal nerve plexus and support the potential role of corneal confocal microscopy as a non-invasive ocular imaging modality for evaluating peripheral neural alterations in hereditary spastic ataxias.

Background: Thoracic aortopathy is a disorder with genetic influence usually presenting in adulthood for which family screening is potentially desirable. Family screening is recommended, but the predictors of a positive screen and real-world pickup rates are unknown. Methods: This was a retrospective cohort of 1022 probands (first affected family member identified) with thoracic aortopathy and one or more features suggestive of a genetic etiology, and their presenting family members, assessed in a cardiac clinic (2009?2024). Imaging and genetic testing were employed in family screening. The prespecified outcomes were uptake and pickup rate of family screening, and proband and family member specific characteristics that predicted a positive family screen. Results: Among probands, 43.5% had one or more family member screened, with an average of 3 relatives per successful proband. 27.6% of family members screened positive. A pre-existing family history of aortopathy was the only variable predicting a higher incidence rate for positive family screen (p = 0.0003). Age of presentation < 60 was not predictive. For family members, extravascular features (p < 0.0001), closer relation to the proband (p < 0.02), male sex (p < 0.0001) and older age (p< 0.0001) all predicted a positive screen. Family members were eight times more likely to screen positive through imaging as compared to genetic testing. Probands with a genetic diagnosis of Marfan and Loeys Dietz syndromes accounted for only 4% of the total. Conclusions: Proband-initiated family screening for thoracic aortopathy has a high yield of affected individuals, even among older probands.

Uterine fibroids (leiomyomas) are highly prevalent benign tumors defined by excessive extracellular matrix (ECM) deposition, altered vascular structure, and progressive tissue stiffening, yet the cellular programs that coordinate these features remain poorly understood. Cellular senescence has been implicated in fibroid biology, but whether senescence represents a uniform state or distinct, functionally specialized cell identities within fibroids is unknown. Here, we identify the distinct heterogeneous populations of senescent cells ("senotypes") present in human fibroids and characterize their role in shaping the fibroid microenvironment. Using single-cell RNA sequencing (scRNA-seq) integrated with a senescence gene signature and protein-level validation, we identify senescent cells (SnC) distributed across fibroblast, mural, and endothelial compartments, each exhibiting distinct transcriptional programs. SnC endothelial cells (ECs) are enriched in fibroids relative to matched myometrium and activate TEAD4-associated mechanosensing, angiogenic, and immune signaling pathways, despite being associated with impaired vessel maturation in situ. In parallel, SnC fibroblast and mural populations in fibroids upregulated SRF-associated cytoskeletal and ECM programs, accompanied by increased COL6A3 expression and collagen VI deposition, consistent with tissue stiffening. Ligand-receptor and spatial analyses reveal that these SnC populations function as interconnected signaling hubs, coordinating immune cell recruitment and stromal remodeling. Importantly, analysis of human fibroids treated with collagenase demonstrated a reduction in both ECM density and SnC burden, supporting a reinforcing relationship between matrix mechanics and senescence. Together, these findings establish senescence in fibroids as a heterogeneous, mechanically reinforced, and network-driven process that links vascular dysfunction, immune signaling, and fibrosis, highlighting distinct SnC states as potential translational targets for non-surgical therapies.

Endothelial senescence is increasingly recognized as a driver of vascular pathology, while immunoglobulin G (IgG) has recently been reported to accumulate in aging tissues and induce senescence in macrophages and microglia. In cerebral cavernous malformations (CCMs), IgG accumulation has been obviously observed in CCM lesions, but the contribution of IgG to endothelial injury remains unclear. Using multi-omic profiling, endothelial models, and CCM mice, we identified IgG-secreting plasma cells enriched in lesions associated with endothelial senescence, hemorrhage, and disease severity. CCM loss-associated mTOR activation impaired lysosomal acidification and IgG processing, promoting intracellular IgG accumulation. IgG, in turn, induced NF-kB-dependent endothelial senescence. In vivo, BCMA-mediated plasma cell depletion attenuated lesion progression, whereas IgG supplementation partially restored disease severity. Anti-CD38 treatment likewise reduced IgG accumulation, endothelial senescence, hemorrhage, and lesion progression. These findings identify lysosomal dysfunction-mediated IgG as a pathogenic trigger of endothelial senescence and support targeting the plasma cell-IgG axis in CCM.

Fibroblast growth factor 2 (FGF2) is frequently induced during ischemic retinal injury and has traditionally been considered a pro-angiogenic factor based largely on studies using exogenous FGF2 administration. However, its endogenous cellular origin and physiological role remain incompletely understood. Here, we used single-cell transcriptomic analysis combined with spatial validation and rod photoreceptor-specific genetic approaches to define the endogenous role of FGF2 during oxygen-induced retinopathy (OIR). We identify rod photoreceptors as a major cellular source of ischemia-induced FGF2. Notably, Fgf2 expression remained elevated during the regression of pathological neovascularization, revealing a temporal dissociation between neuronal stress responses and vascular remodeling. Single-cell analysis further showed that Fgf2 induction occurred within a coordinated photoreceptor stress-response program involving endothelin 2 (Edn2) and B-cell lymphoma 3 (Bcl3). This transcriptional signature was independently reproduced in the N-methyl-N-nitrosourea (MNU)-induced photoreceptor degeneration model. Rod-specific deletion of Fgf2 markedly increased photoreceptor apoptosis, indicating that endogenous FGF2 contributes to photoreceptor survival under ischemic stress. In contrast, neither genetic depletion nor overexpression of FGF2 altered pathological neovascularization or vaso-obliteration. Bidirectional manipulation of FGF2 further modulated the expression of representative stress-associated genes Edn2 and Bcl3, supporting FGF2 involvement in this injury-response program. Finally, receptor expression analysis revealed relatively limited endothelial expression of Fgfr1-Fgfr4 compared with VEGF receptors, suggesting a cellular basis for the distinct effects of endogenous FGF2 and VEGF signaling. Together, these findings identify endogenous retinal FGF2 as a photoreceptor-derived survival factor that is induced during stress but is insufficient to drive pathological angiogenesis. These results support a model in which neuronal adaptation and vascular remodeling represent partially distinct responses during ischemic retinal injury.

Background: Spontaneous cervical artery dissection (sCeAD) is a rare vasculopathy whose pathophysiology remains incompletely understood. Impaired vascular extracellular matrix integrity, including elastic fibers, may contribute to its development. We investigated whether serum fibrillin-1 and soluble elastin fragments (sELF) differ between patients with sCeAD and controls during the acute and chronic stages. Methods: Patients with acute sCeAD were prospectively enrolled at four German stroke centers. Blood samples were collected at baseline and after 6{+/-}1 months. Patients with a first acute ischemic stroke unrelated to sCeAD and healthy individuals served as controls. Serum fibrillin-1 and sELF concentrations were measured using enzyme-linked immunosorbent assays. Results: 61 patients with sCeAD, 53 patients with first non-CeAD ischemic stroke, and 79 healthy controls were included. After sex-matching, serum fibrillin-1 concentrations were significantly lower in patients with acute sCeAD than in healthy controls (97 [60; 192] vs. 176 [113; 269] ng/mL; p=0.009). Fibrillin-1 concentrations were also lower in both male and female patients with sCeAD than in respective healthy controls. In patients with sCeAD, fibrillin-1 concentrations increased significantly after 6 months compared with baseline (171 [130; 270] vs. 104 [67; 205] ng/mL; p=0.021). Serum fibrillin-1 concentrations were higher in men than in women across all study groups. No significant differences in sELF concentrations were observed between groups or time points. Discussion: Serum fibrillin-1 concentrations were lower during acute sCeAD and increased significantly during follow-up, whereas sELF concentrations remained unchanged. These findings support an association between circulating fibrillin-1 and acute sCeAD and warrant further investigation of its role in sCeAD pathophysiology. Pronounced sex-related differences in fibrillin-1 concentrations highlight the importance of sex-specific analyses in future.