Muscle Cells Research Articles

Unveiling impaired vascular function and cellular heterogeneity in diabetic donor-derived vascular organoids.

Vascular organoids (VOs), derived from induced pluripotent stem cells (iPSCs), hold promise as in vitro disease models and drug screening platforms. However, their ability to faithfully recapitulate human vascular disease and cellular composition remains unclear. In this study, we demonstrate that VOs derived from iPSCs of donors with diabetes (DB-VOs) exhibit impaired vascular function compared to non-diabetic VOs (ND-VOs). DB-VOs display elevated levels of reactive oxygen species (ROS), heightened mitochondrial content and activity, increased proinflammatory cytokines, and reduced blood perfusion recovery in vivo. Through comprehensive single-cell RNA sequencing, we uncover molecular and functional differences, as well as signaling networks, between vascular cell types and clusters within DB-VOs. Our analysis identifies major vascular cell types (endothelial cells [ECs], pericytes, and vascular smooth muscle cells) within VOs, highlighting the dichotomy between ECs and mural cells. We also demonstrate the potential need for additional inductions using organ-specific differentiation factors to promote organ-specific identity in VOs. Furthermore, we observe basal heterogeneity within VOs and significant differences between DB-VOs and ND-VOs. Notably, we identify a subpopulation of ECs specific to DB-VOs, showing overrepresentation in the ROS pathway and underrepresentation in the angiogenesis hallmark, indicating signs of aberrant angiogenesis in diabetes. Our findings underscore the potential of VOs for modeling diabetic vasculopathy, emphasize the importance of investigating cellular heterogeneity within VOs for disease modeling and drug discovery, and provide evidence of GAP43 (neuromodulin) expression in ECs, particularly in DB-VOs, with implications for vascular development and disease.

GAS6 and AXL promote insulin resistance by rewiring insulin signaling and increasing insulin receptor trafficking to endosomes.

Growth-arrest specific 6 (GAS6) is a secreted protein that acts as a ligand for TAM receptors (TYRO3, AXL and MERTK). In humans, GAS6 circulating levels and genetic variations in GAS6 are associated with hyperglycemia and increased risk of type 2 diabetes. However, the mechanisms by which GAS6 influences glucose metabolism are not understood. Here, we show that Gas6 deficiency in mice increases insulin sensitivity and protects from diet-induced insulin resistance. Conversely, increasing GAS6 circulating levels is sufficient to reduce insulin sensitivity in vivo. GAS6 inhibits the activation of the insulin receptor (IR) and reduces insulin response in muscle cells in vitro and in vivo. Mechanistically, AXL and IR form a complex, while GAS6 reprograms signaling pathways downstream of IR. This results in increased IR endocytosis following insulin treatment. This study contributes to a better understanding of the cellular and molecular mechanisms by which GAS6 and AXL influence insulin sensitivity.

Isogenic patient-derived organoids reveal early neurodevelopmental defects in spinal muscular atrophy initiation.

Whether neurodevelopmental defects underlie postnatal neuronal death in neurodegeneration is an intriguing hypothesis only recently explored. Here, we focus on spinal muscular atrophy (SMA), a neuromuscular disorder caused by reduced survival of motor neuron (SMN) protein levels leading to spinal motor neuron (MN) loss and muscle wasting. Using the first isogenic patient-derived induced pluripotent stem cell (iPSC) model and a spinal cord organoid (SCO) system, we show that SMA SCOs exhibit abnormal morphological development, reduced expression of early neural progenitor markers, and accelerated expression of MN progenitor and MN markers. Longitudinal single-cell RNA sequencing reveals marked defects in neural stem cell specification and fewer MNs, favoring mesodermal progenitors and muscle cells, a bias also seen in early SMA mouse embryos. Surprisingly, SMN2-to-SMN1 conversion does not fully reverse these developmental abnormalities. These suggest that early neurodevelopmental defects may underlie later MN degeneration, indicating that postnatal SMN-increasing interventions might not completely amend SMA pathology in all patients.

Japanese encephalitis virus infection causes reactive oxygen species-mediated skeletal muscle damage.

Skeletal muscle wasting is a clinically proven pathology associated with Japanese encephalitis virus (JEV) infection; however, underlying factors that govern skeletal muscle damage are yet to be explored. The current study aims to investigate the pathobiology of skeletal muscle damage using a mouse model of JEV infection. Our study reveals a significant increment in viral copy number in skeletal muscle post-JEV infection, which is associated with enhanced skeletal muscle cell death. Molecular and biochemical analysis confirms NOX2-dependent generation of reactive oxygen species, leading to autophagy flux inhibition and cell apoptosis. Along with this, an alteration in mitochondrial dynamics (change in fusion and fission process) and a decrease in the total number of mitochondria copies were found during JEV disease progression. The study represents the initial evidence of skeletal muscle damage caused by JEV and provides insights into potential avenues for therapeutic advancement.

Decoding the muscle transcriptome of patients with late onset Pompe disease reveals markers of disease progression.

Late-onset Pompe Disease (LOPD) is a rare genetic disorder caused by the deficiency of acid alpha-glucosidase leading to progressive cellular dysfunction due to the accumulation of glycogen in the lysosome. The mechanism of relentless muscle damage - a classic manifestation of the disease - has been extensively studied by analysing the whole muscle tissue; however, little, if any, is known about transcriptional heterogeneity among nuclei within the multinucleated skeletal muscle cells. This is the first report of application of single nuclei RNA sequencing to uncover changes in the gene expression profile in muscle biopsies from eight patients with LOPD and four muscle samples from age and gender matched healthy controls. We matched these changes with histology findings using GeoMx Spatial Transcriptomics to compare the transcriptome of control myofibers from healthy individuals with non-vacuolated (histologically unaffected) and vacuolated (histologically affected) myofibers of LODP patients. We observed an increase in the proportion of slow and regenerative muscle fibers and macrophages in LOPD muscles. The expression of the genes involved in glycolysis was reduced, whereas the expression of the genes involved in the metabolism of lipids and amino acids was increased in non-vacuolated fibers, indicating early metabolic abnormalities. Additionally, we detected upregulation of autophagy genes, and downregulation of the genes involved in ribosomal and mitochondrial function leading to defective oxidative phosphorylation. The upregulation of the genes associated with inflammation, apoptosis and muscle regeneration was observed only in vacuolated fibers. Notably, enzyme replacement therapy - the only available therapy for the disease - showed a tendency to restore metabolism dysregulation, particularly within slow fibers. A combination of single nuclei RNA sequencing and spatial transcriptomics revealed the landscape of normal and the diseased muscle, and highlighted the early abnormalities associated with the disease progression. Thus, the application of these two new cutting-edge technologies provided insight into the molecular pathophysiology of muscle damage in LOPD and identified potential avenues for therapeutic intervention.

Commentary: Effect of curcumin nanoparticles on proliferation and migration of mouse airway smooth muscle cells and airway inflammatory infiltration.

Commentary: Effect of curcumin nanoparticles on proliferation and migration of mouse airway smooth muscle cells and airway inflammatory infiltration.

Unraveling the Impact of miR-146a in Pulmonary Arterial Hypertension Pathophysiology and Right Ventricular Function

Pulmonary arterial hypertension (PAH) is a chronic disorder characterized by excessive pulmonary vascular remodeling, leading to elevated pulmonary vascular resistance and right ventricle (RV) overload and failure. MicroRNA-146a (miR-146a) promotes vascular smooth muscle cell proliferation and vascular neointimal hyperplasia, both hallmarks of PAH. This study aimed to investigate the effects of miR-146a through pharmacological or genetic inhibition on experimental PAH and RV pressure overload animal models. Additionally, we examined the overexpression of miR-146a on human pulmonary artery smooth muscle cells (hPASMCs). Here, we showed that miR-146a genic expression was increased in the lungs of patients with PAH and the plasma of monocrotaline (MCT) rats. Interestingly, genetic ablation of miR-146a improved RV hypertrophy and systolic pressures in Sugen 5415/hypoxia (SuHx) and pulmonary arterial banding (PAB) mice. Pharmacological inhibition of miR-146a improved RV remodeling in PAB-wild type mice and MCT rats, and enhanced exercise capacity in MCT rats. However, overexpression of miR-146a did not affect proliferation, migration, and apoptosis in control-hPASMCs. Our findings show that miR-146a may play a significant role in RV function and remodeling, representing a promising therapeutic target for RV hypertrophy and, consequently, PAH.

Dynamic phenotypic shifts and M2 receptor downregulation in bladder smooth muscle cells induced by mirabegron.

Mirabegron is available for treatment of overactive bladder (OAB). However, mechanisms underlying symptom improvements and long-term effects on bladder smooth muscle cells are uncertain. Contractility and growth of bladder smooth muscle contribute to OAB, and depend on smooth muscle phenotypes, and on muscarinic receptor expression. Here, we examined prolonged exposure to mirabegron (20-48h) on phenotype markers, muscarinic receptor expression, and phenotype-dependent functions in human bladder smooth muscle cells (hBSMC). Expression of markers for contractile (calponin, MYH11) and proliferative (MYH10, vimentin) phenotypes, proliferation (Ki-67), and of muscarinic receptors were assessed by RT-PCR. Proliferation, viability, actin organization and contractions in cultured hBSMC were examined by EdU, CCK-8, phalloidin staining and matrix contraction assays. Calponin-1 mRNA decreased with 100nM and 150nM mirabegron applied for 20h (0.56-0.6 fold of controls). Decreases were resistant to the β3-AR antagonist L-748,337 (0.34-0.55 fold, 100-150nM, 20h). After 40h, decreases occured in the presence of L-748,337, but not without L-748,337. MYH11 mRNA increased with 150nM mirabegron (40h, 1.9 fold). This was partly preserved with L-748,337, but not observed after 20h mirabegron exposure. Vimentin mRNA reduced with 150nM mirabegron after 20h, but not after 40h, with and without L-748,337 (0.71-0.63 fold). MYH10 mRNA expression remained unaffected by mirabegron. Exposure to 150nM mirabegron increased Ki-67 mRNA after 20h in the presence of, but not without L-748,337, and after 40h without, but not with L-748,337. Proliferation rates and actin organization were stable with 50-150nM mirabegron (24h, 48h). Viability increased significantly after mirabegron exposure for 20h, and by trend after 40h, which was fully sensitive to L-748,337. M2 mRNA was reduced by 20h mirabegron, which was resistant to L-748,337. Carbachol (3µM) enhanced time-dependent contractions of hBSMC, which was inhibited by mirabegron (150nM) in late phases (24h), but not in early phases of contractions. Conclusion: Mirabegron induces dynamic phenotype alterations and M2 downregulation in hBSMC, which is paralleled by time-shifted anticontractile effects. Phenotype transitions may be involved in improvements of storage symptoms in OAB by mirabegron.

Development of Serum-Reduced Medium for Mackerel Muscle Cell Line Cultivation

Development of Serum-Reduced Medium for Mackerel Muscle Cell Line Cultivation

Enhanced secretion of promyogenic exosomes by quiescent muscle cells.

Signaling interactions are important during skeletal muscle regeneration, where muscle cells in distinct states (quiescent, reactivated, proliferating and differentiated) must coordinate their response to injury. Here, we probed the role of secreted small extracellular vesicles (sEV/exosomes) using a culture model of physiologically relevant cell states seen in muscle regeneration. Unexpectedly, G0 myoblasts exhibited enhanced secretion of sEV (∼150nm) displaying exosome markers (Alix, TSG101, flotillin-1, and CD9), and increased expression of Kibra, a regulator of exosome biogenesis. Perturbation of Kibra levels confirmed a role in controlling sEV secretion rates. Purified sEVs displayed a common exosome marker-enriched proteome in all muscle cell states, as well as state-specific proteins. Exosomes derived from G0 cells showed an antioxidant signature, and were most strongly internalized by differentiated myotubes. Functionally, donor exosomes from all muscle cell states could activate an integrated Wnt reporter in target cells, but only G0-derived exosomes could induce myogenic differentiation in proliferating cells. Taken together, we provide evidence that quiescence in muscle cells is accompanied by enhanced secretion of exosomes with distinct uptake, cargo and signal activating features. Our study suggests the novel possibility that quiescent muscle stem cells in vivo may play a previously under-appreciated signaling role during muscle homeostasis.

MtDNA release promotes cGAS-STING activation and accelerated aging of postmitotic muscle cells

The mechanism regulating cellular senescence of postmitotic muscle cells is still unknown. cGAS-STING innate immune signaling was found to mediate cellular senescence in various types of cells, including postmitotic neuron cells, which however has not been explored in postmitotic muscle cells. Here by studying the myofibers from Zmpste24−/− progeria aged mice [an established mice model for Hutchinson-Gilford progeria syndrome (HGPS)], we observed senescence-associated phenotypes in Zmpste24−/− myofibers, which is coupled with increased oxidative damage to mitochondrial DNA (mtDNA) and secretion of senescence-associated secretory phenotype (SASP) factors. Also, Zmpste24−/− myofibers feature increased release of mtDNA from damaged mitochondria, mitophagy dysfunction, and activation of cGAS-STING. Meanwhile, increased mtDNA release in Zmpste24−/− myofibers appeared to be related with increased VDAC1 oligomerization. Further, the inhibition of VDAC1 oligomerization in Zmpste24−/− myofibers with VBIT4 reduced mtDNA release, cGAS-STING activation, and the expression of SASP factors. Our results reveal a novel mechanism of innate immune activation-associated cellular senescence in postmitotic muscle cells in aged muscle, which may help identify novel sets of diagnostic markers and therapeutic targets for progeria aging and aging-associated muscle diseases.

Ginger protects against vein graft remodeling by precisely modulating ferroptotic stress in vascular smooth muscle cell dedifferentiation

Vein graft failure (VGF) is associated with vein graft (VG) intimal hyperplasia, which is characterized by abnormal accumulation of vascular smooth muscle cells (VSMCs). Most neointimal VSMCs are derived from pre-existing VSMCs via a process of VSMC phenotypic transition, also known as dedifferentiation. There is increasing evidence to suggest that ginger or its bioactive ingredients may block VSMC dedifferentiation, exerting vasoprotective functions; however, the precise mechanisms have not been fully characterized. Therefore, we investigated the effect of ginger on VSMC phenotypic transition in VG remodeling after transplantation. Ginger significantly inhibited neointimal hyperplasia and promoted lumen opening in a 3-month VG, which was primarily achieved by reducing ferroptotic stress. Ferroptotic stress is a pro-ferroptotic state. Contractile VSMCs did not die but instead gained a proliferative capacity and switched to the secretory type, forming neointima after vein transplantation. Ginger and its two main vasoprotective ingredients (6-gingerol and 6-shogaol) inhibit VSMC dedifferentiation by reducing ferroptotic stress. Network pharmacology analysis revealed that 6-gingerol inhibits ferroptotic stress by targeting P53 while 6-shogaol inhibits ferroptotic stress by targeting 5-lipoxygenase (Alox5), both promoting ferroptosis. Furthermore, both ingredients co-target peroxisome proliferator-activated receptor gamma (PPARγ), decreasing PPARγ mediated nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (Nox1) expression. Nox1 promotes intracellular reactive oxygen species (ROS) production and directly induces VSMC dedifferentiation. In addition, Nox1 is a ferroptosis-promoting gene that encourages ferroptotic stress production, indirectly leading to VSMC dedifferentiation. Ginger, a natural multi-targeted ferroptotic stress inhibitor, finely and effectively prevents VSMC phenotypic transition and protects against venous injury remodeling.

Sirt7 protects against vascular calcification via modulation of reactive oxygen species and senescence of vascular smooth muscle cells

Vascular calcification is frequently seen in patients with chronic kidney disease (CKD), and significantly increases cardiovascular mortality and morbidity. Sirt7, a NAD+-dependent histone deacetylases, plays a crucial role in cardiovascular disease. However, the role of Sirt7 in vascular calcification remains largely unknown. Using in vitro and in vivo models of vascular calcification, this study showed that Sirt7 expression was significantly reduced in calcified arteries from mice administered with high dose of vitamin D3 (vD3). We found that knockdown or inhibition of Sirt7 promoted vascular smooth muscle cell (VSMC), aortic ring and vascular calcification in mice, whereas overexpression of Sirt7 had opposite effects. Intriguingly, this protective effect of Sirt7 on vascular calcification is dependent on its deacetylase activity. Unexpectedly, Sirt7 did not alter the osteogenic transition of VSMCs. However, our RNA-seq and subsequent studies demonstrated that knockdown of Sirt7 in VSMCs resulted in increased intracellular reactive oxygen species (ROS) accumulation, and induced an Nrf-2 mediated oxidative stress response. Treatment with the ROS inhibitor N-acetylcysteine (NAC) significantly attenuated the inhibitory effect of Sirt7 on VSMC calcification. Furthermore, we found that knockdown of Sirt7 delayed cell cycle progression and accelerated cellular senescence of VSMCs. Taken together, our results indicate that Sirt7 regulates vascular calcification at least in part through modulation of ROS and cellular senescence of VSMCs. Sirt7 may be a potential therapeutic target for vascular calcification.

Specificities of primary and secondary prevention of lower extremity artery disease in patients with diabetes mellitus.

Diabetes mellitus (DM) is a major risk factor for lower extremity arterial disease (LEAD) and about 20% of symptomatic patients with LEAD have DM. In subjects with DM, LEAD is a cause of morbidity and mortality. DM typically causes complications in the form of macro- and microangiopathy. In these patients, macroangiopathy manifests as atherosclerosis like in non-diabetic patients. However, its course is accelerated due to accompanying risk factors like hyperlipidemia and hypertension, with cumulative effects. Other factors are also relevant such as inflammation, endothelial dysfunction, platelet activation, blood rheological properties, hypercoagulability, and factors stimulating vascular smooth muscle cell proliferation. Additionally, DM is a risk factor for restenosis and amputation. DM is strongly associated with femoral-popliteal and tibial LEAD, which manifests earlier in patients with DM and may progress more rapidly to critical limb ischemia. Diabetic microangiopathy is characterized by arteriolosclerosis and interstitial fibrosis which additionally affects progression and outcomes of angiopathy of lower limbs. Glycemic control particularly decreases microangiopathic complications, while prevention of macrovascular complications requires treatment of accompanying risk factors like hypertension and dyslipidemia.

MiR-196a-5p hinders vascular smooth muscle cell proliferation and vascular remodeling via repressing BACH1 expression

Hyperproliferation of vascular smooth muscle cells (VSMCs) is a driver of hypertensive vascular remodeling. This study aimed to uncover the mechanism of BTB and CNC homology 1 (BACH1) and microRNAs (miRNAs) in VSMC growth and hypertensive vascular remodeling. With the help of TargetScan, miRWalk, miRDB, and miRTarBase online database, we identified that BACH1 might be targeted by miR-196a-5p, and overexpressed in VSMCs and aortic tissues from spontaneously hypertensive rats (SHRs). Gain- and loss-of-function experiments demonstrated that miR-196a-5p suppressed VSMC proliferation, oxidative stress and hypertensive vascular remodeling. Double luciferase reporter gene assay and functional verification showed that miR-196a-5p cracked down the transcription and translation of BACH1 in both Wistar Kyoto rats (WKYs) and SHRs. Silencing BACH1 mimicked the actions of miR-196a-5p overexpression on attenuating the proliferation and oxidative damage of VSMCs derived from SHRs. Importantly, miR-196a-5p overexpression and BACH1 knockdown cooperatively inhibited VSMC proliferation and oxidative stress in SHRs. Furthermore, miR-196a-5p, if knocked down in SHRs, aggravated hypertension, upregulated BACH1 and promoted VSMC proliferation, all contributing to vascular remodeling. Taken together, targeting miR-196a-5p to downregulate BACH1 may be a promising strategy for retarding VSMC proliferation and hypertensive vascular remodeling.

Three-dimensional pore structure of the decellularized parsley scaffold regulates myogenic differentiation for cell cultured meat.

Decellularized plant scaffolds have been used to develop edible scaffolds for cell cultured meat because of their natural structures similar to that of mammalian tissues. However, their diverse three-dimensional (3D) porous structures may lead to differences in myogenic differentiation of skeletal muscle cells. In this study, parsley plant tissues were decellularized and modified by type A gelatin and transglutaminase while retaining, respectively, longitudinal fibrous and transverse honeycomb pore structures. The effects of the structure of the decellularized parsley scaffold on the proliferation and myogenic differentiation of C2C12 cells were investigated and the quality of cell cultured meat was evaluated. The results showed that fibrous pore structure guided cells to be arranged in parallel, whereas honeycomb pore structure connected cells in a circular pattern. After induced differentiation, the fibrous scaffolds were more inclined to form multinucleated myotubes with higher expression of myogenic genes and proteins, and the final cell-based meat contained higher total protein content. Decellularized plant scaffolds with fibrous pore structure were more suitable for myogenic differentiation of C2C12 cells, providing support to the development of edible scaffolds for cultured meat. PRACTICAL APPLICATION: This study investigated the different three-dimensional (3D) pore structure of parsley parenchyma to gain insight into how the 3D pore structure of decellularized plant scaffolds regulates myogenic differentiation, which is expected to address the unstable myogenic differentiation of skeletal muscle cells on decellularized plant scaffolds in cell culture meat production.

IL-33 Facilitates Fibro-Adipogenic Progenitors to Establish the Pro-Regenerative Niche after Muscle Injury.

During the process of muscle regeneration post-injury in adults, muscle stem cells (MuSCs) function is facilitated by neighboring cells within the pro-regenerative niche. However, the precise mechanism triggering the initiation of signaling in the pro-regenerative niche remains unknown. Using single-cell RNA sequencing, 14 different muscle cells are comprehensively mapped during the initial stage following injury. Among these, macrophages and fibro-adipogenic progenitor cells (FAPs) exhibit the most pronounced intercellular communication with other cells. In the FAP subclusters, the study identifies an activated FAP phenotype that secretes chemokines, such as CXCL1, CXCL5, CCL2, and CCL7, to recruit macrophages after injury. Il1rl1, encoding the protein of the interleukin-33 (IL-33) receptor, is identified as a highly expressed signature surface marker of the FAP phenotype. Following muscle injury, autocrine IL-33, an alarmin, has been observed to activate quiescent FAPs toward this inflammatory phenotype through the IL1RL1-MAPK/NF-κB signaling pathway. Il1rl1 deficiency results in decreased chemokine expression and recruitment of macrophages, accompanied by impaired muscle regeneration. These findings elucidate a novel mechanism involving the IL-33/IL1RL1 signaling pathway in promoting the activation of FAPs and facilitating muscle regeneration, which can aid the development of therapeutic strategies for muscle-related disorders and injuries.

Transcriptional, developmental, and functional parallels of lymphatic and venous smooth muscle.

Lymphatic muscle cells (LMCs) are indispensable for lymphatic vessel contraction and their aberrant recruitment or absence is associated with both primary and secondary lymphedema. Despite their critical role in lymphatic vessel function, the transcriptomic and developmental basis that confer the unique contractile properties to LMCs are largely undefined. In this study, we employed single-cell RNA sequencing (scRNAseq), lineage tracing and in vivo imaging to investigate the basis for the hybrid cardiomyocyte and blood vascular smooth muscle cell (SMC) characteristics that have been described for LMCs. Using scRNAseq, the transcriptomes of LMC and venous SMCs from the murine hindlimb exhibited more similarities than differences, although both were markedly distinct from that of arteriole SMCs in the same tissue. Functionally, both lymphatic vessels and blood vessels in the murine hindlimb displayed pulsatile contractility. However, despite expressing genes that overlap with the venous SMC transcriptome, through lineage tracing we show that LMCs do not originate from Myh11+ SMC progenitors. Previous studies have shown that LMCs express cardiac-related genes, whereas in our study we found that arteriole SMCs, but not LMCs, expressed cardiac-related genes. Through lineage tracing, we demonstrate that a subpopulation of LMCs and SMCs originate from WT1+ mesodermal progenitors, which are known to give rise to SMCs. LMCs, however, do not derive from Nkx2.5+ cardiomyocyte progenitors. Overall, our findings suggest that venous SMCs and LMCs and may derive from a related mesodermal progenitor and adopt a similar gene expression program that enable their contractile properties.

Proteomic biomarkers related to obesity in heart failure with reduced ejection fraction and their associations with outcome.

Heart failure (HF) pathophysiology in patients with obesity may be distinct. To study these features, we identified obesity-related biomarkers from 4210 circulating proteins in patients with HF with reduced ejection fraction (HFrEF) and examined associations of these proteins with HF prognosis and biological mechanisms. In 373 patients with trimonthly blood sampling during a median follow-up of 2.1 (25th-75th percentile: 1.1-2.6) years, we applied an aptamer-based multiplex approach measuring 4210 proteins in baseline samples and the last two samples before study end. Associations between obesity (BMI > 30 kg/m2) and baseline protein levels were analyzed. Subsequently, associations of serially measured obesity-related proteins with biological mechanisms and the primary endpoint (PEP; composite of cardiovascular mortality, HF hospitalization, left ventricular assist device implantation, and heart transplantation) were examined. Obesity was identified in 26% (96/373) of patients. A total of 30% (112/373) experienced a PEP (with obesity: 26% [25/96] vs. without obesity: 31% [87/277]). A total of 141/4210 proteins were linked to obesity, reflecting mechanisms of neuron projection development, cell adhesion, and muscle cell migration. A total of 50/141 proteins were associated with the PEP, of which 12 proteins related to atherosclerosis or hypertrophy provided prognostic information beyond clinical characteristics, N-terminal pro-B-type natriuretic peptide, and high-sensitivity troponin T. Patients with HFrEF and obesityshow distinct proteomic profiles compared to patients with HFrEF without obesity. Obesity-related proteins are independently associated with HF outcome. These proteins carry potential to improve management of obesity-related HF and could be leads for future research.

Smooth Muscle Cells-Expressed Bmal1 Regulates Vascular Calcification Independent of the Canonical Circadian Pathway.

Vascular calcification is a major cardiovascular issue that increases morbidity and mortality in diabetes patients. While dysregulation of the circadian master regulator Basic Helix-Loop-Helix ARNT-Like Protein 1 (Bmal1) in vascular smooth muscle cells (VSMC) under diabetic conditions has been suggested, its role in vascular calcification is unclear. In VSMC, Bmal1 was upregulated under high glucose treatment and in aortic tissues from a diabetic mouse model. RNA sequencing from isolated VSMC between Bmal1 deletion and wildtype mice indicated Bmal1's pro-calcification role. Indeed, reduced levels of the osteogenic master regulator, Runt-Related Transcription Factor 2 (Runx2), were found in Bmal1 deletion VSMC under diabetic conditions. Alizarin red staining showed reduced calcification in Bmal1 deletion VSMC in vitro and vascular rings ex vivo . Furthermore, in a diabetic mouse model, SMC-Bmal1 deletion showed reduced calcium deposition in aortas. Collectively, diabetes-upregulated circadian regulator Bmal1 in VSMC contributes to vascular calcification. Maintaining normal circadian regulation may improve vascular health in diabetes.