Journal of Cellular and Molecular Medicine

BackgroundThe etiology and molecular mechanisms of Moyamoya disease (MMD) remain unclear. Exosomes, as carriers of bioactive molecules, may reflect disease-specific alterations and serve as potential biomarkers. This study aimed to investigate disease mechanisms using proteomic profiling of serum-derived exosomes (SDEs) in MMD. Materials and MethodsPeripheral blood each from 15 MMD patients and 15 healthy-controls were used to isolate SDEs via ultracentrifugation. Proteins from pooled SDEs were extracted, digested, and analyzed by LC-MS/MS. Differentially expressed proteins were examined using MetaboAnalyst, DAVID, Enrichr, STRING, and Cytoscape. Key targets were validated at transcript and protein levels using RT-qPCR and ELISA in independent cohorts. ResultsA total of 2,554 proteins were identified, with 213 showing differential expression (118 upregulated, 95 downregulated; p [≤] 0.05). Functional and pathway analyses revealed enrichment in angiogenesis, cytoskeletal remodeling, and endothelial signaling. PRKG2 and MYC were upregulated, while RHOA was downregulated, highlighting their involvement in focal adhesion and PI3K-AKT pathways. Validation confirmed these findings. ConclusionDysregulated proteins were linked to RHOA-ROCK and PI3K-Akt signaling, suggesting their role in driving VSMC phenotypic switching, contributing to vascular occlusion. These findings indicate that altered exosomal-proteins may participate in maladaptive vascular remodeling, although the initial trigger for VSMC transition remains unknown.

Background: Atrial fibrillation (AF) burden is increasingly recognized as a determinant of clinical risk. Currently, AF burden can only be estimated using long-term rhythm monitoring. Bone morphogenetic protein 10 (BMP10) is a protein secreted from cardiac atria associated with AF and AF-related complications. This study evaluated whether BMP10 concentrations are associated with AF burden in a human atrial model: atrial engineered heart tissue (aEHT). Methods: Human induced pluripotent stem cell-derived atrial cardiomyocytes were cast into atrial engineered heart tissues (aEHTs). To mimic AF burden, mature aEHTs were optogenetically-paced at a high rate of 4 Hz, either intermittently for 4 hours every 2 days (~10% burden) or continuously for 24 hours per day (100% burden). After 18 days of high-rate pacing intervention, 7 days of recovery without pacing followed. BMP10 release was quantified by ELISA and contractile function was assessed by video analysis. EHT transcriptional remodeling in response to mimicked AF burden was assessed by RNA sequencing and the effect of recovery was analyzed by qPCR. Results: High-rate optogenetic pacing mimicking AF lead to a dynamic, burden-dependent BMP10 release: BMP10 concentrations in the medium were increased by intermittent optogenetic pacing (~10% burden) and highest under continuous optogenetic pacing (100% burden). BMP10 release declined toward control levels during recovery. Contractile dysfunction was most impaired after continuous pacing and showed only partial recovery within 7 days after pacing cessation. RNA sequencing revealed distinct burden-dependent transcriptional states. Pacing-regulated transcripts were related to BMP/TGF{beta} signaling, atrial identity, calcium handling, contractile phenotype, and electrophysiological remodeling. After recovery, BMP10 mRNA expression remained elevated despite normalization of BMP10 protein release. Conclusions: AF burden dynamically regulates BMP10 release and functional and molecular remodeling in human aEHTs. BMP10 release depicts a secreted protein-based readout of current or recent atrial high-rate stress, whereas persistent transcriptional changes indicate molecular memory of prior AF burden. These findings support BMP10 release as a burden-sensitive AF biomarker

Background: Platelets are critical drivers of thrombo-inflammatory responses in different cardiovascular diseases. Abdominal aortic aneurysm (AAA) is a progressive, life-threatening vascular disorder mainly characterised by chronic inflammation, extracellular matrix degradation, and the formation of a platelet-rich intraluminal thrombus (ILT). Experimental and clinical evidence identified platelets as main players in AAA pathology as evidenced by elevated platelet activation and procoagulant activity that critically contribute to AAA progression. Methods: The present study investigated the contribution of glycoprotein (GP)Ib alpha, the von Willebrand factor (VWF)-binding subunit of the platelet GPIb-IX-V complex, to AAA initiation and progression in experimental AAA using the ePPE mouse model and in patients. Results: Genetic ablation of platelet GPIb alpha significantly attenuated early aneurysm expansion in experimental AAA, indicating a critical role for GPIb alpha during the initial stages of aneurysm development. This initial effect was compensated at later time points showing no differences in aneurysm progression between groups. Notably, genetic deletion of GPIb alpha induced a constitutively hyperactive platelet phenotype already in naive mice that was further amplified during experimental AAA. This elevated platelet hyperactivity was mainly due to increased GPVI activation of platelets 28 days post-surgery. To assess the clinical relevance, spatial profiles of human ILT specimens from patients with AAA were analysed. In the ILT, we detected a highly compartmentalised distribution of GPIb alpha and VWF with pronounced enrichment within the luminal layer. In parallel, circulating VWF activity as well as platelet surface expression of GPIb alpha were significantly increased in patients with AAA. Conclusion: Collectively, these findings identify a dysregulated GPIb alpha-VWF axis in human AAA pathology, mainly characterised by enhanced platelet GPIb alpha surface expression and increased activity of circulating VWF.

Background and AimsMutations in the desmin (DES) gene cause a variety of cardiomyopathies associated with arrhythmias, yet the electrophysiological consequences of these variants remain largely uncharacterized. The aim of this study was to investigate the pathogenic mechanisms of the de novo DES p.R406W variant, which was identified in a 9-year-old patient who suffered from severe ventricular arrhythmias and sudden cardiac death without overt structural heart disease. MethodsHuman induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) carrying the DES p.R406W variant (including the patients line) were compared to isogenic controls. Action potentials (AP) of hiPSC-CMs were recorded using patch-clamp. Furthermore, 3D engineered heart tissues (EHTs) were generated from hiPSC-CMs and their APs were recorded with sharp microelectrodes. Analytical techniques also included transmission electron microscopy (TEM) and integrated transcriptomic and proteomic profiling. Finally, a heterozygous knock-in (KI) mouse model carrying the Des p.R405W ortholog was evaluated through surface ECG, echocardiography and ex vivo cardiac optical mapping. ResultsThe DES p.R406W mutation prolonged AP duration in IM-R406W hiPSC-CMs and EHTs vs Control ones. Multi-omics analysis of EHTs revealed a dysregulation of genes and proteins involved in contractile function, cell adhesion, and electrical activity. TEM imaging revealed changes in Z-disc architecture in mutant tissues. Twenty-week-old Des p.R405W KI mice exhibited ventricular conduction slowing (prolonged QRS) and a high susceptibility to ventricular tachyarrhythmias, likely due to reentrant mechanisms. Mild hypertrophy was also observed, but only in females. ConclusionThe DES p.R406W variant is highly pathogenic, causing electrical and structural remodeling of the myocardium. This study highlights the effectiveness of hiPSC-CMs and EHTs in recapitulating the clinical phenotype of desminopathy, providing a platform for investigating the mechanisms of early-onset cardiac arrhythmias and SCD.

Background: Gastric carcinoma remains a major cause of cancer related mortality worldwide, with tumor progression increasingly recognized as a consequence of complex interactions within the tumor microenvironment. Hypoxia induced signaling, cancer associated fibroblast (CAF) heterogeneity, and immune checkpoint activation play critical roles in tumor progression and immune evasion. However, their integrated relationship in gastric carcinoma remains insufficiently characterized. Objectives: To evaluate the expression of Hypoxia inducible factor 1 alpha and its association with cancer-associated fibroblast subtypes and Programmed death-ligand 1 expression in gastric carcinoma. Methods: This cross sectional analytical study included 100 histologically confirmed gastric carcinoma cases from Satkhira Medical College. Immunohistochemistry was performed for HIF 1 alpha, smooth muscle actin (SMA), fibroblast activation protein (FAP), and PD L1. CAFs were subclassified into myofibroblastic CAFs (myCAFs) and inflammatory CAFs (iCAFs). Associations between biomarkers and clinicopathological variables were analyzed using chi square test, Spearman correlation, and multivariate logistic regression. Receiver operating characteristic (ROC) curve analysis was used to assess model performance. Result: High HIF 1 alpha expression was observed in 55% of cases and demonstrated significant association with poor differentiation (p = 0.001), advanced tumor stage (p = 0.002), and lymph node metastasis (p = 0.001). iCAF predominance was significantly associated with poor differentiation (p = 0.003), advanced stage (p = 0.004), and nodal metastasis (p = 0.004). High PD L1 expression was significantly associated with poor differentiation (p = 0.03), advanced stage (p = 0.001), and lymph node metastasis (p = 0.002). Multivariate logistic regression identified high HIF 1 alpha expression (OR = 3.8, p = 0.001), iCAF dominance (OR = 4.5, p < 0.001), and advanced tumor stage (OR = 2.9, p = 0.004) as independent predictors of high PD L1 expression. Combined high HIF 1 alpha expression and CAF activation demonstrated the highest rate of PD L1 positivity (76.7%, p < 0.001). ROC curve analysis demonstrated good predictive performance of the model with an area under the curve of 0.81. Conclusion: The present study demonstrates a significant interaction between hypoxia, stromal remodeling, and immune checkpoint activation in gastric carcinoma. High HIF 1 alpha expression and inflammatory CAF predominance are strongly associated with aggressive clinicopathological features and increased PD L1 expression, supporting the existence of a coordinated hypoxia stroma immune axis in gastric carcinoma progression. These findings may have potential implications for prognostic stratification and combined targeted therapeutic strategies.

BackgroundPremature ovarian insufficiency (POI) is a condition affecting female fertility e.g., in the context of autoimmune disease. A recent proof-of-concept study showed that B-cell depletion with rituximab (RTX) temporarily restored ovarian function in patients with autoimmune POI. However, the underlying immunological mechanisms behind this response is unknown. ObjectivesTo characterize B- and T-cell phenotypes before and after RTX therapy in the above-mentioned patients and associations with autoantibody levels and treatment response. MethodsCryopreserved cell samples were available from six autoimmune patients with POI, four of whom were positive for autoantibodies against steroidogenic enzymes (21OH, 17OH and/or SCC). Patients received RTX and blood samples were collected at baseline, second infusion (2 weeks), 3, 8 and 12 months. Peripheral B- and T-cell phenotypes were analysed using spectral flow cytometry and correlated with autoantibody levels. ResultsAt baseline, patients displayed signs of B-cell activation by alteration in subset composition compared to matched controls. Upon closer analysis, patients positive for steroidogenic enzyme autoantibodies had significantly increased frequency of CXCR3+switched memory (SWM) B-cells than autoantibody-negative patients, which correlated positively with anti-17OH and-SCC autoantibody levels. In parallel, increased Th1 cell frequencies were also observed in the same patient group. Longitudinal analyses showed expected B-cell depletion followed by repopulation after RTX. Changes in T-cell subsets were observed at 8 months with reduced frequencies of Th1 and Tfh cells. ConclusionAutoimmune POI displayed altered B- and T-cell subsets implicating cell crosstalk, immune cell infiltration to sites of inflammation such as ovaries, and presence of Th1-driven immunopathogenesis. Key messagesO_LIIncreased CXCR3+SWM B cells and Th1 cells were found in patients with autoimmune POI who were positive for autoantibodies against steroidogenic enzymes. C_LIO_LICXCR3 expression on SWM B cells positively associated with anti-17OH and anti-SCC autoantibody levels. C_LIO_LIRTX induced rapid B-cell depletion followed by repopulation and a delayed T-cell effect at 8 months with reduced Th1 and Tfh cell frequencies. C_LI Capsule summaryRTX has recently been shown to restore fertility in autoimmune POI. Lymphocyte profiling identified Th1 bias accompanied by CXCR3+memory B cells that correlate with autoantibody levels. B-cell depletion appears to temporarily alleviate ovarian autoimmunity and ameliorate physiological function.

Objective: Aortic dissection (AD) is a serious, life-threatening cardiovascular crisis. While vascular proteoglycans normally buffer hemodynamic stress, their aberrant accumulation in the AD aorta predisposes to rupture. Aggrecan (ACAN) is among the most frequently elevated proteoglycans in this setting, yet its pathogenic relationship to AD remains undefined. This study aimed to investigate the role of ACAN in AD and identify potential therapeutic targets. Approach and Results: We employed human tissues, animal models, and cell cultures. Western blotting and immunohistochemistry (IHC) were performed on aortic samples from AD patients and controls to assess contractile markers, MMPs, ACAN, and SOX9. In vivo, AD was induced in mice using {beta}-aminoisobutyric acid (BAPN) and Ang II, with AAV-mediated ACAN knockdown in VSMCs. Histological staining confirmed model validity and enabled assessment of AD incidence, mortality, and protein expression changes. In vitro, human aortic vascular smooth muscle cells (HAVSMCs) were stimulated with Ang II, and SOX9 and ACAN were silenced via siRNA to evaluate the functional impact of ACAN downregulation. In human AD aortas, elevated ACAN and its upstream transcription factor SOX9 drove VSMC phenotypic switching and ECM degradation. In mice, ACAN knockdown corrected AD-induced medial structural disruption, elastin fragmentation, and collagen deposition, markedly reducing incidence and mortality while curbing excessive mitochondrial fission and rescuing functional integrity. ACAN silencing attenuated Ang II-induced mitochondrial damage in HAVSMCs via the RNA-seq-identified YAP1/TAZ pathway, and Verteporfin restored homeostasis. Conclusion: ACAN is aberrantly expressed in AD and murine aortas. ACAN downregulation delays disease progression by preserving mitochondrial homeostasis and function via the YAP1/TAZ pathway.

Background: Platelets play a critical role in thrombo-inflammation following acute myocardial infarction (AMI). Objectives: Their impact on hepatic thrombopoietin (TPO) regulation after AMI remains poorly understood to date. Methods: Wildtype, thrombocytopenic or GPVI deficient mice underwent ischemia/reperfusion (I/R) injury. Platelet activation and turnover, and hepatic expression of different receptors involved in TPO regulation were analyzed. Results: Here, we identify a heart-liver axis that dynamically regulates platelet production post AMI. An elevated platelet turnover with increased reticulated as well as desialylated platelets early after AMI was detected. This was accompanied by the upregulation of specific receptors such as Asgr1/2 and IL-6R and increased phosphorylation of STAT3 in the liver and elevated numbers of megakaryocytes in spleen tissue. Consequently, increased TPO plasma levels at 24h post AMI were detected and platelet counts were rapidly restored after AMI. Platelet depletion induces a compensatory increase in hepatic TPO expression and plasma TPO levels, accompanied by dysregulated STAT3 signaling. This response contrasts with GPVI-deficient (Gp6-/-) mice, which exhibit no major alterations in TPO regulation, suggesting distinct mechanisms between acute thrombocytopenia and chronic platelet receptor deficiency in platelet homeostasis after AMI. These findings provide the first evidence of a direct heart-liver axis regulating platelet homeostasis after AMI, driven by inflammatory and hepatic signaling pathways. Conclusions: Our study highlights a previously unrecognized role of the heart-liver axis in controlling platelet turnover, with hepatic STAT3 signaling as a key mediator. These insights enhance our understanding of post AMI platelet homeostasis and may be critical for future therapeutic strategies.

IntroductionPreeclampsia is characterized by a pro-inflammatory, anti-migratory and anti-angiogenic placental phenotype. Impaired spiral artery remodeling stemming from trophoblast dysfunction is a key pathogenic mechanism. Little is known about the processes that govern trophoblast function normally and in preeclampsia. In preeclampsia, placental Let-7b-5p is reduced. The objectives of this study were to determine the normal function of Let-7b-5p in human trophoblast cells, to examine whether the ssRNA sensors, Toll-like receptor (TLR) 7 and/or TLR8 are mediators of trophoblast Let-7b-5p function, and whether disruption of this pathway promotes a preeclampsia-like phenotype in the trophoblast. MethodsThe human first trimester trophoblast cell line, Sw.71, was transfected with a Let-7b- 5p mimic, a Let-7b-5p inhibitor, or scramble control. Cells were treated with or without the TLR7 inhibitor IRS661 or the TLR8 inhibitor CUCPT9a. Trophoblast migration was measured using a two-chamber assay and interactions with human endometrial endothelial cells (HEECs) was measured using a 3D matrigel model. Trophoblast anti-angiogenic sFlt-1 release was measured by ELISA and sFLT1 mRNA measured by RT-qPCR. ResultsTransfection of trophoblast cells with a Let-7b-5p mimic elevated migration through activation of TLR7 and TLR8, while in a TLR7-dependent manner, the Let-7b-5p mimic negatively regulated sFlt-1 production. Furthermore, inhibition of trophoblast Let-7b-5p reduced migration, elevated FLT1 mRNA expression and sFlt-1 release, and reduced trophoblast-endometrial endothelial cell interactions. ConclusionsThis study highlights a role for TLR7/TLR8-activating Let-7b-5p in promoting normal trophoblast function and endothelial interactions and that disruption in this miR-driven signaling pathway may be relevant to processes driving a pre-eclamptic placental phenotype. HighlightsTrophoblast migration is positively driven by Let-7b-5p activating TLR7 and TLR8 Let-7b-5p, via TLR7, negatively regulates trophoblast anti-angiogenic sFlt-1 production. Inhibition of trophoblast Let-7b-5p reduces trophoblast migration and normal interactions with endometrial endothelial cells, while sFlt-1 production is elevated. TLR7/TLR8-activating Let-7b-5p promotes normal trophoblast function and endothelial interactions and disruption in this miR-driven signaling pathway may promote a preeclamptic placental phenotype.

Background: Activation of the renin-angiotensin system (RAS) contributes to aortic diseases caused by fibrillin-1 haploinsufficiency (Fbn1C1041G/+). However, whether RAS activation influences mitral valve disease associated with FBN1 variants remains undefined. This study determined whether inhibition or augmentation of the RAS affects mitral valve pathology in Fbn1C1041G/+ mice. Methods: Angiotensinogen (AGT) antisense oligonucleotides (ASOs) were administered for 12 weeks to inhibit the RAS in male and female Fbn1C1041G/+ mice. To augment the RAS, mice were infused with angiotensin II (AngII; 1,000 ng/kg/min) for 28 days. Mitral valve and heart function were monitored by ultrasound. At termination, hearts were dissected free and tissue sectioned to assess mitral valve leaflet thickness and pathology. Results: Mitral valve leaflet thickness was significantly increased in Fbn1C1041G/+ mice compared to Fbn1+/+ littermates of both sexes. ASO-mediated AGT inhibition did not alter this phenotype despite markedly reduced plasma AGT concentrations. AngII infusion increased mitral valve leaflet thickness in Fbn1C1041G/+ mice with increased proteoglycan and collagen content, while exerting minimal effects in Fbn1+/+ mice. Twenty-five percent of AngII-infused Fbn1C1041G/+ mice died of non-vascular causes. Ultrasound showed mitral valve dysfunction in AngII-infused Fbn1C1041G/+ mice. Grossly expanded left atria were common in AngII-infused Fbn1C1041G/+ mice. These markedly enlarged left atria contained increased fibrosis, macrophage accumulation, and platelet-rich masses. Conclusions: Mitral valves of Fbn1C1041G/+ mice did not respond to AGT inhibition but were markedly more susceptible to AngII infusion, with greater increases in thickness and compositional changes than mitral valves of Fbn1+/+ mice. In addition, AngII promoted mitral valve dysfunction that were associated with pronounced left atrial pathology.

Background Intervertebral disc degeneration (IDD) is a leading cause of chronic low back pain and disability worldwide, affecting most individuals over 50 years of age. Despite its prevalence, no disease-modifying therapies exist, and current interventions are limited to reducing pain. Cellular senescence and the associated secretory phenotype (SASP) have been increasingly recognized as major drivers of disc matrix degradation and inflammation. However, the upstream molecular mechanisms that lead to IDD degeneration are still unknown. Connexin 43 (Cx43), a gap junction protein implicated in progression of age-related diseases, has emerged as a key regulator of cellular senescence and inflammatory signalling in musculoskeletal tissues. Methods Human primary cells were isolated from intervertebral disc samples obtained from patients classified into clinically meaningful groups: healthy controls, chronic/mechanical degeneration (DDD, ADJ, ASD), and acute/inflammatory event (herniated nucleus pulposus, HNP). Cx43 expression was assessed by qPCR and Western blotting. Cellular senescence was evaluated through SA-{beta}-gal staining and analysis of p53/p21 expression. SASP factors and EMT-related markers were measured by qPCR. Protein expression was quantified by immunoblotting across different age groups and degeneration grades. Results In this current study Cx43, was identified as the most abundant connexin isoform in human intervertebral discs, showing a progressive increase in expression with age and disc degeneration. Also, high Cx43 expression correlated with increased expression of the senescent markers p53 and p21 and increased SA-{beta}-gal activity. Besides, increased expression of EMT-related and differentiation markers has been correlated with high Cx43 levels in human IDD samples, consistent with fibrotic remodeling processes. Conclusions These findings identify aberrant upregulation of Cx43 signaling as a potential mechanistic link between intervertebral disc cellular senescence and extracellular matrix degradation, with the ensuing inflammatory response, representing a novel potential therapeutic target to modulate senescence-driven pathogenesis and modulate IDD progression.

Lymphedema is a chronic disease characterized by impaired lymph drainage and accumulation of protein-rich interstitial fluid, which progresses to develop irreversible fibrosis. Importantly, effective therapies and treatments are lacking to alleviate and mitigate the disease. The pathological inflammation and fibrogenesis underlying lymphedema prompted us to evaluate a preclinical medicine IVA337, a pan-peroxisome proliferator-activated receptor (PPAR) agonist that improves liver fibrosis in patients with metabolic dysfunction-associated steatohepatitis (MASH) by activating three PPAR isoforms (, {beta}/{delta}, {lambda}), which play critical roles in lipid metabolism, anti-inflammation responses, and anti-fibrogenesis. Here, we investigate the therapeutic effects of IVA337 during the early stage of surgery-induced secondary lymphedema in mice and explored the underlying mechanisms. IVA337 administration alleviated lymphedema progression, improved lymphatic drainage, reduced dermal thickness, and resolved lymphatic vessel dilation. Mechanistically, IVA337 suppressed the TGF{beta}/SMAD2/3 signaling pathway, reduced immune cells infiltration, and improves lymphatic vessels integrity. In human dermal lymphatic endothelial cells (HDLECs), IVA337 attenuated TGF{beta} induced SMAD2/3 phosphorylation and preserved the expression of cell junction Claudin5, reduced VE-Cadherin-stained cell-cell gaps. Collectively, our findings demonstrate that IVA337 protects against early stage lymphedema by inhibiting TGF{beta}/SMAD2/3-mediated inflammatory and fibrotic responses. This study provides a potential therapeutic strategy to improve lymphatic function during the early phase of lymphedema and prevent progressive fibrosis in patients with lymphedema and related disorders.

BACKGROUD: Sepsis is a life-threatening condition characterized by a dysregulated host response to infection. It frequently precipitates sepsis-associated acute kidney injury (SA-AKI). 20-Hydroxyeicosatetraenoic acid (20-HETE), is an important regulator of vascular tone. We aim to investigate whether disruption of 20-HETE signaling contributes to septic renal dysfunction. METHODS: In the animal study, Sprague-Dawley rats underwent the cecal ligation and puncture to induce sepsis and SA-AKI. Comprehensive cardiac and renal ultrasonographic assessments were performed in all animals. Blood samples were collected for biochemical assessment of renal function and quantification of circulating 20-HETE levels. Kidney tissues were harvested for analysis of the 20-HETE-GPR75-PLC/PKC signaling pathway and histological evaluation by hematoxylin and eosin staining. In the clinical study, circulating 20-HETE levels and renal hemodynamic parameters derived from point-of-care ultrasound were evaluated in healthy individuals and patients with sepsis. Venous blood samples were collected for quantification of serum 20-HETE concentrations. RESULTS: Firstly, CLP-induced septic rats produced a biphasic hemodynamic response, with early compensation followed by late cardiovascular decompensation, accompanied by progressive renal dysfunction, tubular injury, depletion of circulating and renal 20-HETE, and reduced renal GPR75 expression. Secondly, Restoration of 20-HETE signaling with a pharmacological analog improved systemic and renal hemodynamics, increased MAP, reduced serum creatinine, blood urea nitrogen, and ameliorated renal pathological injury. These protective effects were partially attenuated by PKC inhibition, implicating PKC-dependent signaling in 20-HETE-mediated renal protection. Moreover, in septic patients, circulating 20-HETE concentrations were reduced compared with healthy individuals and were lowest among those with SA-AKI, paralleling alterations in renal hemodynamic parameters measured by point-of-care ultrasound. CONCLUSIONS: Sepsis-associated depletion of 20-HETE accompanies impaired systemic and renal hemodynamic regulation and is associated with the development of SA-AKI. Restoration of 20-HETE signaling improves hemodynamic function and attenuates renal injury, potentially involving GPR75-associated PLC/PKC signaling. KEY WORDS: sepsis-associated acute kidney injury, 20-hydroxyeicosatetraenoic acid, GPR75, renal hemodynamics, point-of-care ultrasonography

Marfan syndrome is a multisystem connective tissue disorder with age-dependent and sex-dependent phenotypic variability. This study characterized early cardiovascular, valvular, transmitral flow, and skeletal changes in Fbn1C1041G/+ mice. Male and female wild-type and Fbn1C1041G/+ mice were examined from 4 to 20 weeks of age. Transthoracic echocardiography was used to measure aortic root and ascending aortic diameters, anterior mitral leaflet length, and mitral inflow Doppler parameters. Lateral X-ray imaging was used to quantify kyphosis angle. Fbn1C1041G/+ mice of both sexes developed progressive aortic dilation, with earlier and broader aortic root involvement in males and delayed annular dilation in females. Anterior mitral leaflet elongation was detected from 8 weeks in males and at 20 weeks in females. In contrast, reduced E-wave velocity and E/A ratio were present from 4 weeks in both sexes, without changes in A-wave velocity. Kyphosis was detectable earlier in females than in males. These findings show that early Fbn1-related phenotypes follow organ-specific and sex-dependent temporal patterns, supporting age-resolved and sex-stratified assessment in preclinical Marfan syndrome studies.

Background: Cardiac fibrosis is a hallmark of many cardiovascular diseases, driven by sustained fibroblast activation and excessive extracellular matrix deposition, leading to myocardial stiffening and impaired contractility. Current therapies inadequately address this process. This study evaluated the antifibrotic potential of lanifibranor, a balanced pan-peroxisome proliferator-activated receptors (PPARs) agonist, in TGF-beta1-induced cardiac fibrosis. Methods: Human cardiac microtissues, along with 2D and 3D cardiac fibroblast and cardiomyocyte cultures, were used to assess cell viability, structure, metabolism, contractility, and gene expression. Results: Lanifibranor reduced TGF-beta1-induced fibrosis by limiting fibroblast activation and matrix deposition without affecting viability. In fibroblasts, these effects were associated with partial restoration of mitochondrial respiration and reduced focal adhesion maturation. In cardiac microtissues, lanifibranor improved contraction kinetics, decreased profibrotic transcriptional activity, and preserved bioenergetic homeostasis despite altered nucleotide balance. In cardiomyocytes, treatment normalized contractility and calcium handling while maintaining metabolic stability. Conclusions: Lanifibranor attenuates TGF-beta1-driven cardiac fibrosis by combining antifibrotic effects with metabolic and functional improvements in human models.

Background: Lumbar disc herniation, with or without concomitant disc degeneration, is a major cause of lumbar radiculopathy and low back pain, which also a key public musculoskeletal disorder without an exact pathophysiology. Studies have suggested that inflammatory cells and biochemical markers of inflammation also play an important role in lumbar radiculopathy in addition to nerve compression. The aim of the present study was to assess the association of selected circulatory inflammatory markers (CRP, hs-CRP and E-selectin) in patients with lumbar disc herniation without radiological degeneration (LDH) and lumbar disc herniation with radiological degeneration (LDHD). Materials & methods: This case-control study included 208 participants, comprising 104 patients with lumbar disc pathology and 104 controls. Patients were further stratified into LDH (n=67) and LDHD (n=37). Serum CRP, hs-CRP and E-selectin concentrations were measured. Results: Among the patients, 35.6 % presented with LDHD while 64.4 % had only LDH. Significantly increased median hs-CRP (p<0.001) and CRP (p<0.001) were observed in patients groups compared to controls, while CRP showing a consistent independent association across the combined disease (OR=1.68, 95% CI=1.33-2.14, p<0.001), LDHD (OR=1.62, 95% CI=1.16-2.20, p=0.005) and LDH (OR=1.69, 95% CI=1.30-2.20, p<0.001) multivariable models. No significant difference was observed in serum E-selectin between the study groups. Multivariable models incorporating inflammatory and clinical variables demonstrated substantially greater discriminatory performance than individual biomarkers alone. Conclusion: Elevated circulating CRP and hs-CRP concentrations were associated with lumbar disc pathology, with CRP showing a consistent independent association across the combined disease, LDH and LDHD multivariable models, whereas E-selectin showed no significant association. Multivariable models incorporating inflammatory and clinical variables demonstrated greater discriminatory performance than individual biomarkers. These findings support a potential systemic inflammatory component in lumbar disc pathology, although the cross-sectional nature of the measurements does not establish causality or a local inflammatory response within the disc.

Background Hypertrophic and dilated cardiomyopathies (HCM and DCM) are the most common inherited cardiomyopathies. However, genotype-specific molecular and functional cardiomyocyte phenotypes and responses to neurohormonal stimulation remain incompletely understood. Here, we investigated whether patient-derived HCM and DCM cardiomyocytes exhibit distinct baseline phenotypes or differential responses to hypertrophic stimulation and pharmacological treatment. Methods Three human-induced pluripotent stem cell (hiPSC) lines were used: a control line, an HCM patient-derived line carrying a MYBPC3 mutation, and a DCM patient-derived line carrying an LMNA mutation. The cells were differentiated into hiPSC-cardiomyocytes, which were exposed to endothelin-1 and the GATA4-targeted compound 3i-1262, followed by transcriptional and protein expression analyses. In addition, engineered heart tissues (EHTs) were generated and cultured for 40 days, with endothelin-1 and 3i-1262 treatment applied during the final 20 days. Lastly, {beta}-adrenergic stimulation with isoprenaline was performed. EHT contractile function was quantified using MUSCLEMOTION. Results Patient-derived hiPSC-cardiomyocytes exhibited genotype-dependent responses to endothelin-1 at the transcriptional and protein levels. DCM-cardiomyocytes failed to maintain structural integrity in the EHTs, resulting in tissue fracture or cessation of beating. Functional analyses demonstrated distinct baseline contractile properties between control and cardiomyopathy EHTs, as well as differential responses to endothelin-1 and isoprenaline. Conclusions Patient-derived hiPSC-cardiomyocytes exhibit genotype-specific molecular and functional phenotypes. EHTs generated from HCM hiPSC-derived cardiomyocytes showed a progressive decline in apparent force, whereas DCM EHTs fractured over time, suggesting mutation-associated phenotypes in 3D cardiac tissue models. These findings highlight the utility of hiPSC-based cardiac models for investigating molecular and functional disease mechanisms and pharmacological responses in inherited cardiomyopathies. Keywords: hypertrophic cardiomyopathy, dilated cardiomyopathy, engineered heart tissues, hiPSC-derived cardiomyocytes, GATA4

BACKGROUND: Subarachnoid hemorrhage (SAH) induces inflammatory responses and subsequent immune cell activation, which may contribute in cerebral vasospasm, microcirculatory impairment and poor neurological outcomes. Although cerebral vasospasm has traditionally been considered a major cause of delayed cerebral ischemia after SAH, therapies targeting angiographic vasospasm have not consistently improved functional outcomes. Early inflammatory responses may contribute to microcirculatory impairment, cerebral vasospasm, and subsequent neurological injury. Herein, we investigated whether interleukin-10 (IL-10), an anti-inflammatory cytokine, improves these outcomes in an experimental SAH model. METHODS: Mice received intramuscular injections of either an adeno-associated virus encoding IL-10 (AAV/IL-10) vector or an AAV expressing green fluorescent protein (AAV/GFP) vector (control). India ink angiography was performed to assess the diameter of the sphenoidal segment of the middle cerebral artery (MCA), the total length of the visible cortical arteries, and cortical staining intensity, as indices of cerebral vasospasm, microcirculatory impairment, and cerebral perfusion, respectively. Perivascular inflammatory cell infiltration and cytokine levels were assessed using immunohistochemistry and ELISA. We also evaluated the therapeutic efficacy of the AAV/IL-10 vector when administered immediately after SAH induction. RESULTS: IL-10 overexpression significantly improved neurological outcomes after SAH and was associated with attenuated cerebral vasospasm and microcirculatory impairment, as well as preservation of cerebral perfusion. It also significantly reduced neutrophil and macrophage infiltration around the internal carotid artery and attenuated SAH-induced elevations in IL-6 and matrix metalloproteinase-3 levels. Mice treated with the AAV/IL-10 vector immediately after SAH induction showed significant improvements in neurological scores and cerebral perfusion. CONCLUSIONS: AAV-mediated IL-10 overexpression improves neurological outcomes after SAH, likely by attenuating inflammatory responses, cerebral vasospasm, and microcirculatory impairment. These findings suggest that IL-10-based anti-inflammatory therapy is a promising therapeutic strategy for SAH.

Background: Autophagy is a homeostatic pathway supporting stress adaptation and is dysregulated in atherosclerosis. Its potential as a biomarker or therapeutic target in atherosclerotic vascular disease (ASVD) remains incompletely defined. We measured autophagic flux in peripheral blood mononuclear cells (PBMCs) from patients with peripheral arterial disease (PAD) or carotid stenosis (CS), compared with healthy controls, and explored clinical outcome associations. Methods: Ninety-four patients with PAD or CS and 19 healthy controls were studied. Autophagic flux was quantified from fresh blood using a validated ex vivo chloroquine inhibition ELISA measuring LC3BII accumulation. Major adverse cardiovascular events (MACE) and major adverse limb events (MALE) were ascertained over a median follow up of 828 days. Results: The ASVD cohort comprised claudication (n = 16), chronic limb threatening ischemia (CLTI; n = 49), and CS (n = 29). Autophagic flux was higher in ASVD than controls (mean 281.4 vs. 182.3 ng LC3BII/mg protein/h; p < 0.0001) and remained independently associated after multivariable adjustment. Within CLTI, concurrent infection was associated with lower flux (p = 0.001), approaching control levels (p = 0.327). In CLTI, higher flux quartiles were associated with lower MACE risk, most strongly for quartile 3 (hazard ratio 0.07 vs. quartile 1, 95% CI 0.01 to 0.50; p = 0.009). Conclusion: Autophagic flux is elevated in PBMCs from ASVD patients, independent of age and sex. Attenuated flux in CLTI with concurrent infection may indicate autophagic exhaustion in advanced disease. The association between higher flux and lower MACE in CLTI suggests prognostic utility, warranting evaluation in larger prospective studies.

Background: Pulmonary arterial hypertension (PAH) is a serious disease with poor prognosis, especially in infants or preterm babies and there is still no optimal treatment for this disease. Noradrenalin (NE) is a vasoactive mediator which is released by sympathetic ganglion. According to previous studies, NE/1-adrenoreceptors is not only in regulating normal physiologic responses, but also in the pathogenesis of PAH. However, the mechanisms of NE in PAH are not fully understood. Methods: Human PASMC (PASMC) was used in this study. Cell viability assay and Wound healing assay were used to evaluate the proliferation and migration of PASMC. Immunoprecipitation and western blots analysis were used to investigate the mechanisms which involved in NE-induced PASMC proliferation. Results: We investigated that NE could induce human PASMC proliferation and migration. Furthermore, we first find that endothelin 1 (ET-1) signaling pathway plays an important role in NE-induced PASMC proliferation. ET1 is a critical molecular which is known for regulating cell growth and migration. We investigated that NE could increase NE-1 secretion, further enhancing ET-1 bind to its receptors. For further clarifying the downstream signals in NE/ET-1 induced PASMC proliferation, we detected the phosphorylation and expression levels of ERK and JNK. Conclusions: By combining the results from ours and previous studies, we believed that JNK/c-jun pathway may play an important role in NE-induced PASMC proliferation. Key Words: Noradrenaline; Pulmonary Arterial Hypertension; Pulmonary Artery Smooth Muscle Cells; Endothelin-1; JNK/c-Jun Signaling.