The FASEB Journal

The uterus requires energy for sustained contractility during labor, to deliver the fetus and diminish the risk of postpartum hemorrhage. Our objective was to define energy requirements and assess metabolic flexibility in quiescent and contractile myometrial cells. Cells were treated with oxytocin to stimulate myometrial contractility. We found that myometrial cells rely on oxidative phosphorylation during quiescence and, when treated with oxytocin, can adapt to higher energy demands by shifting their energy production to glycolysis. Treatment with mitochondrial oxidation inhibitors revealed that in quiescent myometrial cells basal oxygen consumption rate decreased when treated with glucose oxidation inhibitor UK5099, but not the long chain fatty acid oxidation inhibitor etomoxir or the glutamine oxidation inhibitor BPTES. In oxytocin treated myometrial cells, this decrease was also observed upon BPTES treatment in addition to UK5099, suggesting that contractile myometrial cells can shift energy production from glucose to glutamine. Functionally, myometrial contractility was significantly reduced by UK5099 but not by etomoxir, further indicating dependence on glucose utilization.

Intervertebral disc degeneration is associated with loss of nucleus pulposus (NP) cell phenotype and extracellular matrix, both processes linked to changes in cytoskeletal contractility and cell shape. Here, we tested whether microenvironment-specific modulation of RhoA signaling can restore NP-like morphology and gene expression in NP cells cultured in 2D and in 3D alginate. In 2D monolayer culture, where cells are spread and mechanically activated, pharmacologic inhibition of RhoA with CT04 reduced RhoA activity, decreased actomyosin contractility gene expression, and shifted morphology toward a smaller, more circular phenotype. Bulk RNA sequencing showed that CT04 treatment increased expression of NP phenotypic and matrix-related genes including ACAN, GDF5, CHST3, and MUSTN1 while decreasing expression of catabolic and fibroblast-associated genes including ADAMTS1/9 and COL1, consistent with enrichment of extracellular matrix pathways. In contrast, RhoA activation with CN03 in 2D culture increased actin and phosphorylated myosin light chain intensity but produced limited phenotypic improvement. In 3D alginate, which minimizes integrin-mediated adhesion, baseline actomyosin markers were reduced relative to 2D culture. In alginate, RhoA activation with CN03 increased the amount of actin, phosphorylated myosin light chain, and actomyosin gene expression, yet also promoted a more compact, circular morphology and increased NP markers, including ACAN and KRT19 with repeated dosing. Across culture conditions, increased cell roundness was consistently associated with increased ACAN expression, indicating strong coupling between cytoskeletal state, morphology, and NP matrix programs. Together, these findings demonstrate that RhoA pathway perturbation can promote NP phenotypic gene expression in both 2D and 3D culture, but the direction of optimal modulation depends on the microenvironment, supporting RhoA signaling as a context-dependent therapeutic target for disc regeneration.

Equine endometrosis is a major cause of subfertility in mares characterized by fibrotic remodeling of the endometrium. Although transforming growth factor beta 1 (TGF-{beta}1) is implicated in fibrogenesis, the relationship between endometrosis severity and transcripts associated with tissue maintenance and proliferation remains incompletely defined. Present study evaluated endometrial mRNA expression of IGF1, MKI67, TGFB1, and ACTA2 in relation to endometrosis severity and defined histopathological features. Forty-seven endometrial samples were graded according to the modified Kenney and Doig (KD) categories. Relative mRNA expression was quantified by RT-qPCR and histopathology was extended using a standardized feature-based assessment. TGFB1 mRNA expression was higher in category I+ than in categories I and III (p = 0.041) and in samples with glandular basal lamina disruption (p = 0.020). MKI67 mRNA expression was lower in samples with luminal epithelial erosion (p = 0.049). IGF1 mRNA expression correlated negatively with KD category ({rho} = -0.401, p = 0.015), glandular degeneration ({rho} = -0.340, p = 0.043), overall inflammatory infiltration ({rho} = -0.387, p = 0.020), lymphocytic infiltration ({rho} = -0.426, p = 0.010), and neutrophilic infiltration ({rho} = -0.448, p = 0.006). MKI67 correlated positively with ESR1 ({rho} = 0.887, p < 0.001). These findings indicate that early endometrosis-compatible lesions are associated with increased TGFB1 transcription and that epithelial damage is accompanied by reduced MKI67 expression. The inverse associations between IGF1 expression and both lesion severity and inflammatory infiltration support a link between progressive histopathological changes and reduced expression of a growth factor involved in tissue maintenance.

Extracellular matrix (ECM) remodeling is essential for adaptation to changing mechanical demands, yet the mechanisms linking altered strain to functional outcomes remain poorly understood. This study aimed to define molecular and cellular programs driving the adaptation of tendon to increased (exercise) and decreased (disuse) strain. Male murine flexor tendon explants were cultured in incubator-housed tensile bioreactors and subjected to step changes in cyclic strain. After acclimation at 1% cyclic strain, exercise tendons experienced a step increase to 5% strain, while disuse tendons underwent stress deprivation. Increased strain produced significant mechanical adaptations, including increased elastic modulus and failure stress. Multiscale analyses of matrix organization, tissue composition, protein synthesis, signaling factors, and proteolytic activity revealed the mechanisms underlying these adaptations. Exercise-induced functional improvements were linked to an anabolic remodeling program characterized by TGF-{beta} and IL-6 signaling, small leucine-rich proteoglycan expression, MMP suppression, and enhanced collagen alignment. These findings indicate that regulators of matrix organization and turnover, beyond synthesis alone, are critical for functional adaptation. In contrast, mechanical unloading reduced collagen synthesis and alignment and promoted an MMP-dominant, catabolic phenotype favoring matrix breakdown. This study provides a comprehensive characterization of ECM remodeling, linking defined mechanical perturbations to molecular regulation and emergent structure-function relationships. These findings identify targetable mediators of adaptive remodeling and establish a framework for future studies of maladaptive ECM changes in aging, injury, and disease.

Human embryo implantation, occurring approximately one week after fertilization, remains poorly understood due to ethical and technical limitations of in vivo investigation. To overcome these barriers, and model this critical developmental event, encompassing peri- and early post-implantation stages, we used an in vitro embryo attachment model composed of donor-derived endometrial epithelial cells forming an open-faced endometrial layer (OFEL) and human stem cell-derived blastoids recapitulating human day 5 blastocysts in peri-implantation model. Following attachment, developmental progression was further investigated on laminin-coated substrates to capture early post-implantation dynamics. Despite its central role as the primary endocrine signal of early pregnancy, human chorionic gonadotropin (hCG) remains largely uncharacterized in this context. Here, we describe the transcriptomic profile of blastoid-endometrial co-cultures relative to OFEL alone, identifying CGA and CGB3/5/8 as among the most strongly upregulated genes following blastoid attachment to hormonally stimulated OFEL. Consistent with these findings, immunoassays and luteinizing hormone/choriogonadotropin receptor (LHCGR) activation assays of conditioned media confirmed the secretion of heterodimeric, biologically active hCG and its free subunits in co-cultures, but not in endometrial layers alone. Notably, the hyperglycosylated hCG heterodimer was the predominant isoform detected. Co-culture with the endometrial component significantly increased hCG secretion compared with blastoids cultured alone, an effect further enhanced by hormonal priming in the peri-implantation model. Collectively, these findings indicate that a hormonally primed endometrial environment not only promotes blastoid attachment but also amplifies embryonic hCG production and bioactivity, underscoring the importance of maternal endocrine cues in early embryo-endometrium communication. Furthermore, our peri- and early post-implantation models recapitulate key aspects of reciprocal endocrine signaling between embryonic and endometrial tissues, providing a tractable experimental framework to investigate embryo-endometrium crosstalk.

The endometrium, the inner lining of the uterus, is a dynamic tissue that undergoes precise molecular and structural changes to achieve a receptive state capable of supporting embryo implantation. Although the uterine environment was long considered sterile, molecular studies have detected microbial signals and bioactive compounds that may influence endometrial function. Endometrial epithelial organoids (EEOs) provide a three-dimensional in vitro model that recapitulates the architecture, polarity, and hormonal responsiveness of native endometrial tissue. This study aimed to elucidate how bacterial-derived compounds, including D-lactate (D-lac), commonly associated with Lactobacillus communities, and lipopolysaccharides (LPS), a component of Gram-negative bacteria, affect the transcriptomic profile of the endometrial epithelium under a hormonally induced receptive state. EEOs were exposed to different concentrations of these compounds, and relative metabolic activity was monitored through resazurin-based assays, revealing no significant alterations across the conditions tested. Transcriptomics analysis of hormonally stimulated EEOs, mimicking the mid-secretory phase, revealed that D-lac modulated genes related to epithelial development, tissue remodelling and growth regulation, whereas LPS influenced genes associated with inflammatory signalling and immune response. While key markers of receptivity remained largely stable, small transcriptional changes suggest that microbial signals may modulate the functional balance of the receptive endometrium. These findings highlight a modulatory role of microbial signals on endometrial epithelial function and demonstrate that EEOs are a robust platform for exploring host-microbe interactions in the uterus, offering new insights into the mechanisms underlying uterine receptivity.

The human endometrium undergoes cyclic, hormone-driven remodeling that establishes a transient window of receptivity required for embryo implantation, placentation, and maintenance of pregnancy. Decidualization of endometrial stromal cells is a central component of this process and can be induced in vitro using cAMP alone or in combination with ovarian steroid hormones (EPC: estradiol, progesterone, and cAMP). Although cAMP activates the core decidual transcriptional program, whether hormone supplementation induces a more physiologically relevant response remains unclear, particularly in 3D endometrial organoid (Endo-organoid) models which have emerged as a new alternative methodology (NAM). Here, we compared morphological and transcriptomic responses of human endometrial stromal cell-derived Endo-organoids undergoing decidualization induced by cAMP or EPC stimulation. EPC-treated Endo-organoids exhibited enhanced structural remodeling and more advanced morphological transformation compared with cAMP-treated organoids. RNA-seq analysis revealed substantial overlap in canonical decidual gene expression between the two conditions, but EPC induced broader transcriptional and pathway-level changes, including enrichment of metabolic, stress-response, and differentiation-related processes. Together, these findings demonstrate that while cAMP activates the core decidual program, EPC elicits a broader and more physiologically relevant decidualization response in 3D human Endo-organoids, providing guidance for optimizing Endo-organoids to study endometrial receptivity, implantation, and early pregnancy success.

The human fallopian tube plays a critical role in reproductive biology, yet the structural organization and immune repertoire of this tissue remain incompletely characterized. Here, we performed an in-depth analysis of human fallopian tube tissue from women of reproductive age across three distinct anatomical regions (isthmus, ampulla, and fimbriae) across the menstrual cycle. Using antibody-based imaging for EPCAM, CD8A, and CD20 together with automated image analysis, the epithelial thickness and spatial distribution of T and B lymphocytes was assessed. No significant differences in epithelial thickness were observed between proliferative and secretory phases within any tubal region. In contrast, significant regional differences were identified, with the epithelium being thickest in the isthmus and thinnest in the ampulla. Both CD8A+ T lymphocytes and CD20+ B lymphocytes were detected throughout the fallopian tube, and a strong correlation between T and B lymphocyte abundance was observed across patients. Spatial analysis further revealed that both lymphocyte populations were preferentially localized within the mucosal compartment adjacent to the lumen. Notably, intraepithelial B lymphocytes were identified throughout the fallopian tube. Together, these findings provide new insight into epithelial organization and immune cell distribution in the human fallopian tube, highlighting the complexity of the tubal immune microenvironment and its potential relevance for reproductive biology.

Maintenance of skeletal muscle function is essential for functional independence, quality of life and healthspan. Muscle RING-finger protein-1 (MuRF1) negatively regulates muscle function and mass through ubiquitination and degradation of muscle proteins. Accordingly, genetic and pharmacological inhibition of MuRF1 attenuates muscle wasting and weakness under catabolic stress. To explore the potential of MuRF1 inhibitors (e.g., MyoMed-205) to improve muscle health, we investigated here the long-term effects of MyoMed-205 on functional capacity and muscle physiology in rats under basal conditions. Wistar rats were randomized to control or MyoMed-205 groups and were followed for 4 or 8 weeks. Body weight, food and water intake, and exercise capacity were monitored weekly. At each endpoint, the soleus muscle was collected for histological analyses. MyoMed-205-treated rats showed normal basic survival-related behaviors and body growth. After 8 weeks, MyoMed-205-treated animals exhibited enhanced exercise capacity (speed (m/min): +45%, p = 0.01; endurance (min): +47%, p = 0.03; and distance covered (m): +87%, p = 0.04) compared with baseline performance. Conversely, no differences were found in soleus fiber type distribution, cross-sectional area, or lipid and collagen content. Our findings indicate that MyoMed-205 enhances functional exercise capacity independently of changes in soleus muscle structure in rats under basal conditions.

Diabetic foot ulcers (DFUs) remain a major clinical challenge as diabetes prevalence rises, emphasizing the need for improved therapeutics and relevant preclinical models. Common rodent wound-healing models poorly recapitulate human skin anatomy and repair. Although porcine skin is comparable to human skin, many studies employ young, healthy pigs that do not reflect typical chronic human wounds. Here, we evaluated wound healing in full-thickness skin wounds in non-diabetic and diabetic Yucatan minipigs. RNA sequencing identified key transcriptional differences in wounds of diabetic versus non-diabetic animals, including pathways linked to increased inflammation and oxidative stress, as well as decreased metabolism and extracellular matrix organization, known hallmarks of DFUs. These findings support this preclinical model as a powerful approach for discovery and therapeutic testing in diabetic wounds and provide a novel data set for further mining of potential gene targets for diabetic wound intervention.

The acceleration of hepatic lipid disposal during acute exercise has been proposed as a contributor to the anti-steatotic effects of exercise training. Ketogenesis, which produces acetoacetate (AcAc) and {beta}-hydroxybutyrate ({beta}OHB) from fatty acids, is among the lipid disposal pathways stimulated by exercise. This study tested the hypothesis that hepatic ketogenesis is necessary for exercise training to lower liver lipids. Liver-specific 3-hydroxymethylglutaryl-CoA synthase 2 knockout (HMGCS2 KO) mice and wild type (WT) littermates underwent sedentary, acute exercise, and exercise training protocols. Liver ketone bodies and lipids were determined via mass spectrometry platforms. Stable isotope infusions in conscious, unrestrained mice defined mitochondrial oxidative fluxes at rest and during exercise. Loss of hepatic HMGCS2 decreased liver AcAc and {beta}OHB concentrations and impaired their increase during exercise. Liver triacylglycerides (TAGs) were comparable between genotypes at rest (i.e., ad libitum fed and short fasted conditions). In contrast, liver TAGs were elevated in HMGCS2 KO mice following acute, non-exhaustive exercise. Liver TCA cycle flux was higher in KO mice at rest. During exercise, TCA cycle flux increased in both WT and KO mice but was not different between genotypes with greater exercise duration. This suggests that enhanced disposal of lipids via the TCA cycle may prevent liver lipid accumulation in HMGCS2 KO mice under sedentary conditions, but not during exercise. Unexpectedly, exercise training decreased liver TAGs similarly in both HMGCS2 KO and WT mice. In conclusion, hepatic ketogenesis supports liver lipid homeostasis during acute exercise, but is not required for exercise training to lower liver lipids. NEW & NOTEWORTHYExercise training has been proposed to mitigate liver steatosis partly through enhanced hepatic lipid disposal. During acute exercise, the disposal of fatty acids to ketone bodies is stimulated. This study tested the hypothesis that hepatic ketogenesis was required for exercise training to reduce liver fat in mice. The results show that hepatic ketogenesis is needed to prevent lipid accumulation during acute exercise, but is not necessary for exercise training to lower liver lipids.

The myometrium, the muscle layer of the uterus, undergoes profound phenotypic changes throughout pregnancy, labor, and postpartum, allowing for successful reproduction. Transcriptomic analysis of the murine myometrium revealed distinct gene expression signatures corresponding to these physiological stages, which reflect the dynamic remodeling processes occurring in the uterus during late gestation, term labor, and postpartum. These transcriptomic signatures at specific gestational points were accompanied by changes to the accessible chromatin landscape. Notably, increased chromatin accessibility was observed in the mouse myometrium at full term (gestational day 19) before the onset of active labor contractions. Accessible chromatin regions were bound by the transcription factor JUND, from the AP-1 family, and associated with progesterone receptor (PR) binding. Depletion of the progesterone receptor isoform B (PRB) from accessible chromatin at this prelabor stage implicates progesterone receptor isoform A (PRA) as a binding partner with JUND at prelabor and labor. During labor onset, accessible chromatin regions were associated with elevated production of enhancer RNAs and enriched in binding sites for transcription factors from the AP-1, SOX, and ETS families, implicating additional transcription factors in the labor process. Although PRB was strikingly absent from labor-associated accessible chromatin, it was found associated with histone H3 lysine 27 trimethylated (H3K27me3) repressed regions in late gestation and the postpartum period. These findings provide new insight into the dynamic transcriptional regulatory networks and chromatin-based mechanisms controlling gene expression in the myometrium across gestational stages providing new therapeutic targets for reproductive disorders.

Angiotensin-converting enzyme 2 (ACE2) is expressed in adipocytes, yet the mechanisms regulating its intracellular trafficking remain unclear. Here, we investigated whether ACE2 trafficking is coordinated with insulin-responsive vesicle dynamics mediated by the glucose transporter GLUT4 in 3T3L1 adipocytes. Subcellular localization analyses revealed that adipocyte differentiation promotes partial incorporation of ACE2 into insulin-responsive GLUT4-associated vesicular compartments, whereas ACE2 displayed a diffuse distribution in fibroblasts lacking a mature GLUT4 trafficking system. Reconstitution of insulin-responsive GLUT4 vesicle formation through exogenous expression of Sortilin and AS160 in fibroblasts was sufficient to partially recruit ACE2 into perinuclear GLUT4-positive compartments, indicating dependence on canonical GLUT4 vesicle machinery. NanoBiT assays demonstrated a regulated association between ACE2 and GLUT4 that was modestly enhanced by acute insulin stimulation but reduced following prolonged insulin exposure. Insulin stimulation also produced a slight increase in ACE2 surface exposure, while association with GLUT4 was accompanied by reduced ACE2 shedding, suggesting that recruitment into distinct trafficking routes may alter ACE2 accessibility to shedding machinery in adipocytes. Structural modeling further suggested that ACE2 and GLUT4 can form a membrane-compatible complex. Together, these findings indicate that ACE2 trafficking is coordinated with insulin-responsive GLUT4 vesicle dynamics, revealing a previously unrecognized association between metabolic signaling and ACE2 cellular dynamics in adipocytes, with potential implications for metabolic dysfunction and ACE2-associated disease processes.

IntroductionThere remains considerable debate as to the cause of the epidemic of Mesoamerican Nephropathy (MeN). We have previously reported early loss of estimated glomerular filtration rate (eGFR) as a surrogate for disease onset in a population-representative cohort study of young-adults at risk of disease from Northwest Nicaragua. Using a nested case-control approach we analysed urine and serum proteins surrounding this timepoint with the aim of gaining insight into the primary disease aetiology. MethodsWe conducted label-free ultra high-performance liquid chromatography mass-spectrometry based proteomics using urine samples collected at the study visit before, and at, first observed eGFR loss amongst cases and compared results to matched controls. We then performed direct protein measurements in a discovery cohort followed by quantification of serum total immunoglobulin E (stIgE) at multiple timepoints in a replication cohort. ResultsProteomic analysis demonstrated no differences in the levels of any single protein between cases and controls (n=25 each), at either timepoint, after correction for multiple comparisons. However, functional enrichment analysis demonstrated upregulation of adaptive immune pathways amongst cases. Direct measurements in the discovery cohort using high-sensitivity PCR-based immunoassay (n=21 controls, 19 cases) demonstrated higher stIgE in cases at the study visit immediately prior to first observed eGFR loss (mean difference 810kU/L, 95% confidence interval (CI): 162-1457kU/L). In the replication cohort (n=22 cases, 21 controls) an stIgE level >500kU/L measured by electrochemiluminescence in study samples from any timepoint in the 3 years prior to the first observed loss of eGFR was independently associated with case status when compared to samples from controls at matched visits (adjusted Odds Ratio: 8.1, 95% CI: 1.4-47.8). DiscussionA high level of stIgE precedes loss of eGFR in those at risk of MeN. Understanding what leads to this rise is likely to be key to understanding the cause of the MeN epidemic. Lay SummaryMesoamerican nephropathy describes an epidemic-level chronic kidney disease impacting rural working age adults in Central America. Although a number of exposures, including occupational heat exposure, have been proposed the cause of the epidemic, there remains much debate as to the primary aetiology of the disease. In this study we interrogated urine and blood samples from individuals from affected communities at risk of disease both before and after they develop kidney dysfunction. Using two different approaches, analysis of both urine and blood samples provide evidence of upregulation of immunoglobulin-E (IgE) related pathways in the 2-3 years before individuals develop evidence of kidney disease. Infections (particularly those involving parasites) and allergic reactions, but not heat exposure, have been reported to increase IgE levels. Going forwards, understanding the cause of this increase in IgE in individuals at risk of disease is likely to provide insight into the cause of Mesoamerican Nephropathy epidemics.

Ferroptosis is linked to various diseases, but the role of transferrin (TF) in endometriosis (EM) remains unclear. Expression levels of ferroptosis-related proteins, including transferrin (TF), transferrin receptor (TFRC), and glutathione peroxidase 4 (GPX4), were analyzed by western blotting. Compared to normal endometrial stromal cells, eutopic and ectopic endometrial stromal cells from EM patients exhibited significantly enhanced proliferative and migratory abilities, accompanied by a marked reduction in glutathione (GSH) levels in both eutopic and ectopic tissues. TF and TFRC expression was upregulated in ectopic endometrium relative to normal controls, while GPX4 expression was downregulated. To evaluate the functional role of TF, siRNA-mediated knockdown was performed in endometrial stromal cells, with knockdown efficiency confirmed by western blotting. Functional assays demonstrated that TF knockdown not only suppressed cell proliferation (CCK-8 and clonogenic assays) and migration (wound healing assay) but also significantly increased apoptosis rate (flow cytometry with Annexin V-FITC/PI staining).These findings implicate TF in the pathogenesis and progression of endometriosis, likely through modulating endometrial stromal cell proliferation, migration, and apoptosis.

Disturbances of 24-hour or circadian rhythms imposed by everyday irregular work and/or social schedules have been linked to vascular disease, including ischemic stroke. Using an established shift work-like paradigm and preclinical model for ischemic stroke, we have shown that environment-induced circadian dysregulation exacerbates stroke outcomes differentially to a greater extent in male than female rats. Because more severe stroke outcomes and circadian rhythm disturbances have been linked to gut pathophysiology, present study examined the effects of chronic LD cycle shifting on gut cytoarchitecture, microbiota composition, metabolites, and gut-derived inflammatory mediators. Adult (5-7mo) rats were divided into 2 groups and exposed for 50d to a fixed or shifted (lights-on advanced by 12hr/5d) LD 12:12 cycle. Circadian entrainment of activity rhythms was stable in all rats on the fixed LD 12:12 cycle but was severely disrupted during exposure to shifted LD cycles. Significant changes in the composition of the gut microbiome including reduced alpha diversity, shifts in beta diversity and correlations between the abundance of beneficial gut bacteria and stroke survival were observed in male but not female rats exposed to shifted LD cycles relative to fixed LD controls. This effect of circadian dysregulation on gut microbiota was accompanied by evidence of pathologic gut morphology (i.e., shorter and blunted villi, crypt hyperplasia disruption of tight junction proteins and gut barrier integrity), elevated serum endotoxin concentrations, decreased levels of the short-chain fatty acid (SCFA) butyrate, and increased circulating levels of the inflammatory cytokine IL-17A in shifted LD male rats. These results suggest that alterations in gut morphology, microbiota and metabolites may contribute to sex differences in the effects of shift work-related circadian dysregulation on ischemic stroke outcomes.

Introduction: Podocyte injury is central to the pathogenesis of most glomerulonephritides (GN) and causes segmental glomerulosclerotic lesions that predict progression in IgA Nephropathy (IgAN). Recent advances in high-resolution microscopy and AI-assisted image analysis have enabled detailed quantification of podocyte foot process (FP) morphology. However, whether nanoscale podocyte morphometrics can predict disease progression or treatment response in GN has not been investigated. Aim: To evaluate whether nanoscale podocyte morphometric parameters predict clinical characteristics, disease progression, and treatment response in GN, with a focus on IgAN. Method: Podocyte morphometrics were analyzed in kidney biopsies from patients with GN using high-resolution microscopy and the deep learning-based tool Automatic Morphometric Analysis of Podocytes (AMAP). Four morphometric parameters were quantified: slit diaphragm length (SDL), FP area, FP circularity and FP perimeter. These parameters were correlated with clinical characteristics, conventional electron microscopy (EM) findings and longitudinal follow-up data. Results: The study included 37 patients with GN from Danderyd University Hospital (Stockholm, Sweden), with IgAN representing the largest diagnostic subgroup (n = 19). The median follow-up for the cohort was 3.0 years. SDL correlated significantly with urine albumin-to-creatinine ratio (uACR; p = 0.021), whereas conventional EM measurements did not (p = 0.22). Within the IgAN subgroup, lower SDL was associated with a steeper decline in eGFR, higher FP area with increased long-term proteinuria, and higher FP circularity with improvement in uACR during the first year. The association between lower SDL and eGFR decline remained as a trend in IgAN patients not treated with corticosteroids (p = 0.068) but was absent in the treatment group (p = 0.59). Conclusion: In this proof-of-concept study, nanoscale podocyte morphometrics demonstrated greater sensitivity than conventional EM in quantifying podocyte injury and predicting progression in IgAN. These findings suggest that high-resolution morphometrics may improve risk stratification in IgAN but require validation in larger, independent cohorts before clinical implementation.

BackgroundSystemic sclerosis (SSc) is a severe autoimmune disease characterised by progressive fibrosis driven by fibroblast activation. Primary cilia, key hubs for profibrotic signalling, are markedly shortened in SSc fibroblasts, but the mechanisms underlying this phenotype remain unclear. This study aimed to define the signalling pathways responsible for primary cilia shortening and fibroblast activation in SSc. MethodsPrimary dermal fibroblasts from SSc patients and healthy controls were analysed for cilia incidence and length by immunofluorescence, profibrotic marker expression by qPCR, and contractility using gel contraction assays. Cells were treated with TGF{beta}1 and pharmacological inhibitors targeting AURKA, HDAC6, ROCK2, and Smad3 signalling. CAV1-silenced fibroblasts were used as an in vitro model of SSc. ResultsMaintenance of the constitutively short primary cilia phenotype in SSc fibroblasts did not require active TGF{beta} signalling. However, TGF{beta}1 induced reversible cilia shortening in healthy fibroblasts and further shortened cilia in SSc fibroblasts to a similar final length, mediated by Rho/ROCK2 rather than canonical Smad3-dependent signalling. Constitutive cilia shortening in SSc was driven by aberrant AURKA activity upstream of HDAC6, promoting ciliary disassembly. Pharmacological inhibition of AURKA or HDAC6 selectively elongated cilia in SSc fibroblasts, reduced profibrotic marker expression, and abrogated fibroblast contractility. CAV1-silenced fibroblasts similarly exhibited constitutive cilia shortening that was reversed by AURKA inhibition without affecting healthy cells. ConclusionsAberrant activation of the AURKA/HDAC6 axis maintains short primary cilia and promotes fibroblast activation in SSc. These findings reveal a mechanistic link between cilia morphology and fibrosis and identify AURKA as a potential therapeutic target for SSc-associated tissue remodelling.

BackgroundEffective skin wound healing is essential for restoring tissue integrity following injury. Repair proceeds through phases of hemostasis, inflammation, proliferation, and remodeling, but molecular mechanisms governing these stages remain poorly defined. Vascular niche cells (VNCs)-including endothelial cells, vascular smooth muscle cells (SMCs), and fibroblasts-are central regulators of healing, but the lack of longitudinal in vivo human data has limited identification of VNC-derived signals that distinguish effective repair from pathological healing such as ulcers. Thus, defining the regulation of VNCs in wound healing addresses a critical knowledge gap. MethodsWe developed a protocol for wound healing using dermal forearm punch biopsies to track longitudinal repair in healthy volunteers. Single-cell and spatial transcriptomics were performed to identify and validate signaling activities within VNCs. ResultsWe spatiotemporally defined the inflammation, proliferation, and remodeling phases of human skin wound healing with a focus on VNCs. Spatial analysis localized this activity for VNCs and immune cells within a heterogenous granulation zone that later led to re-epithelializion. Angiogenesis was dominated by Vegf, Egf and Hif1 signaling. Extracellular matrix (ECM) remodeling occurred through Collagen, Laminin, Thrombospondin, and Fibronectin. SMCs emerged as dominant drivers of injury-induced remodeling including basement membrane and interstitial ECM components compared to fibroblasts. This SMC-led program was further defined by robust induction of TIMP1, an inhibitor of matrix degradation, which localized to granulation tissue and correlated with re-epithelialization and wound resolution. Lastly, we compared remodeling factors between healing and non-healing human diabetic foot ulcers (DFUs). SMCs in non-healing DFUs had deficient expression for core remodeling factors, including TIMP1, indicating SMC activity is needed for effective healing. ConclusionWe identified an SMC-driven model of wound repair in which TIMP1-dependent activity underpins granulation zone formation. Failure of this program defined a mechanistic basis for impaired healing in ulcers, identifying SMCs and TIMP1 as therapeutic targets.

ObjectivesThe 2025 Global Consensus recommends continuing biologics throughout pregnancy in women with inflammatory bowel disease (IBD). Real-world evidence on biologic treatment patterns and outcomes remains limited. This study compared maternal and neonatal outcomes across different biologic use trajectories during pregnancy. MethodsA retrospective study was performed in pregnant women with IBD, treated and/or delivering at the University Hospitals Leuven, Belgium, between 2017 and 2025. Patients were categorized as continuers, discontinuers, non-users or initiators of biologics during pregnancy ResultsAmong 255 pregnancies, 103 (40.4%) were continuers, 68 (26.7%) discontinuers, 77 (30.2%) non-users, and 7 (2.7%) initiators. Before conception, 67.1% used biologics. Third-trimester disease activity was most frequent in initiators (42.9%, 3/7) and discontinuers (19.1%, 13/68), followed by non-users (14.3%, 11/77) and continuers (13.6%, 14/103). C-sections occurred more often in non-users (41.3%, 26/63) and discontinuers (39.4%, 26/66) than continuers (31.1%, 23/74). Preterm birth was more common among initiators (14.3%, 1/7) and discontinuers (12.1%, 8/66) than continuers (8.0%, 6/75) and non-users (3.2%, 2/62). Low birthweight occurred most in initiators (14.3%, 1/7), continuers (8.1%, 6/74) and discontinuers (6.1%, 4/66). Small-for-gestational-age infants were most frequent among continuers (14.9%, 11/74) and initiators (14.3%, 1/7) than discontinuers (7.6%, 5/66). ConclusionsWomen who discontinued biologics during pregnancy had higher rates of C-sections, preterm birth, and third-trimester disease activity than continuers, supporting continuation of biologics in pregnancy. The higher SGA rates among continuers, however, require further investigation. Initiators showed the poorest outcomes, highlighting the need for adequate disease control before and during pregnancy.