Muscle Cells Research Articles

SGK1-Mediated Vascular Smooth Muscle Cell Phenotypic Transformation Promotes Thoracic Aortic Dissection Progression.

The occurrence of thoracic aortic dissection (TAD) is closely related to the transformation of vascular smooth muscle cells (VSMCs) from a contractile to a synthetic phenotype. The role of SGK1 (serum- and glucocorticoid-regulated kinase 1) in VSMC phenotypic transformation and TAD occurrence is unclear. Four-week-old male Sgk1F/F (Sgk1 floxed) and Sgk1F/F;TaglnCre (smooth muscle cell-specific Sgk1 knockout) mice were administered β-aminopropionitrile monofumarate for 4 weeks to model TAD. The SGK1 inhibitor GSK650394 was administered daily via intraperitoneal injection to treat the mouse model of TAD. Immunopurification and mass spectrometry were used to identify proteins that interact with SGK1. Immunoprecipitation, immunofluorescence colocalization, and GST (glutathione S-transferase) pull-down were used to detect molecular interactions between SGK1 and SIRT6 (sirtuin 6). RNA-sequencing analysis was performed to evaluate changes in the SIRT6 transcriptome. Quantitative chromatin immunoprecipitation was used to determine the target genes regulated by SIRT6. Functional experiments were also conducted to investigate the role of SGK1-SIRT6-MMP9 (matrix metalloproteinase 9) in VSMC phenotypic transformation. The effect of SGK1 regulation on target genes was evaluated in human and mouse TAD samples. Sgk1F/F;TaglnCre or pharmacological blockade of Sgk1 inhibited the formation and rupture of β-aminopropionitrile monofumarate-induced TADs in mice and reduced the degradation of the ECM (extracellular matrix) in vessels. Mechanistically, SGK1 promoted the ubiquitination and degradation of SIRT6 by phosphorylating SIRT6 at Ser338, thereby reducing the expression of the SIRT6 protein. Furthermore, SIRT6 transcriptionally inhibits the expression of MMP9 through epigenetic modification, forming the SGK1-SIRT6-MMP9 regulatory axis, which participates in the ECM signaling pathway. Additionally, our data showed that the lack of SGK1-mediated inhibition of ECM degradation and VSMC phenotypic transformation is partially dependent on the regulatory effect of SIRT6-MMP9. These findings highlight the key role of SGK1 in the pathogenesis of TAD. A lack of SGK1 inhibits VSMC phenotypic transformation by regulating the SIRT6-MMP9 axis, providing insights into potential epigenetic strategies for TAD treatment.

Long Noncoding RNA Function in Smooth Muscle Cell Plasticity and Atherosclerosis.

In the healthy mature artery, vascular cells, including endothelial cells, smooth muscle cells (SMCs), and fibroblasts are organized in different layers, performing specific functions. SMCs located in the media are in a differentiated state and exhibit a contractile phenotype. However, in response to vascular injury within the intima, stimuli from activated endothelial cells and recruited inflammatory cells reach SMCs and induce a series of remodeling events in them, known as phenotypic switching. Indeed, SMCs retain a certain degree of plasticity and are able to transdifferentiate into other cell types that are crucial for both the formation and development of atherosclerotic lesions. Because of their highly cell-specific expression profiles and their widely recognized contribution to physiological and disease-related biological processes, long noncoding RNAs have received increasing attention in atherosclerosis research. Dynamic fluctuations in their expression have been implicated in the regulation of SMC identity. Sophisticated technologies are now available to allow researchers to access single-cell transcriptomes and study long noncoding RNA function with unprecedented precision. Here, we discuss the state of the art of long noncoding RNAs regulation of SMC phenotypic switching, describing the methodologies used to approach this issue and evaluating the therapeutic perspectives of exploiting long noncoding RNAs as targets in atherosclerosis.

Islet-derived exosomal miR-204 accelerates insulin resistance in skeletal muscle by suppressing sirtuin 1: An in vivo study in a mouse model of high-fat diet-induced obesity.

The interaction between pancreatic islets and skeletal muscle plays a pivotal role in the development of insulin resistance. The present study aimed to elucidate the impact of non-hormonal molecules from islets on the insulin sensitivity of skeletal muscle cells. We developed a mouse model of obesity through a high-fat diet, assessing glucose tolerance and conducting miRNA sequencing on skeletal muscle samples. An in vitro model was established by treating cells with palmitic acid, and exosomes in the supernatant were characterized using scanning electron microscopy and CD63 expression analysis. Intracellular miR-204-5p levels were quantified by RT-PCR. Our in vivo model demonstrated a robust correlation between miR-204-5p level alterations and obesity-induced insulin resistance. Elevated fatty acid levels were observed to increase miR-204-5p in both skeletal muscle and islets. In cellular studies, palmitic acid increased miR-204-5p in MIN-6 islet β-cells but not in C2C12 skeletal muscle cells. Exosomes containing miR-204-5p, secreted by palmitic acid-treated MIN6 cells, were identified through morphological examination, immunoblotting for the exosomal marker CD63, and intraexosomal miR-204-5p level measurement. C2C12 cells were shown to uptake islet-derived miR-204-5p exosomes, as evidenced by the uptake of Exo-Red labeled exosomes. TargetScan analysis identified a highly conserved binding site for miR-204-5p in the 3' UTR of Sirt mRNA. Functional studies indicated that miR-204-5p overexpression reduced glucose consumption and uptake in C2C12 cells, decreased Sirt expression, and impaired insulin signaling, as evidenced by reduced Akt phosphorylation and membrane Glut4 levels. Our findings reveal that miR-204-5p contributes to the development of insulin resistance in obesity and acts as a signaling molecule in the crosstalk between pancreatic islets and skeletal muscle.

TRPC6 Channel Regulates Airway Remodeling in Chronic Obstructive Pulmonary Disease Causing Right Heart Failure.

The role of the canonical transient receptor potential 6 (TRPC6) channel in chronic obstructive pulmonary disease (COPD) remains poorly understood at the mechanistic level. Objects: This study aims to investigate the involvement of TRPC6 in COPD and its signaling mechanisms in human airway smooth muscle cells (HASMCs). Methods and Results: The study found that mRNA and protein expression of TRPC6 increased in cultured HASMCs that were incubated with nicotine, as measured by reverse transcription quantitative polymerase chain reaction and Western blot analysis. Nicotine treatment significantly enhanced TRPC6 transcriptional activity in HASMCs through nuclear factor (NF)-κB, as demonstrated by co-immunoprecipitation and electrophoretic mobility shift assays. Furthermore, miR-135a/b-5p was shown to downregulate TRPC6 expression in HASMCs at the mRNA and protein levels, as confirmed by luciferase reporter assays. Immunohistochemistry assays in a mouse model of cigarette-induced airway remodeling revealed a significant increase in smooth muscle (SM) cell proliferation and SM layer mass. Conclusion: These findings suggest that nicotine exposure increases HASMC proliferation and migration through NF-κB signaling, and that cigarette smoke inhalation causes airway SM layer remodeling via altered TRPC6-induced Ca2+ influx, which is abolished by miR-135a/b-5p both in vitro and invivo. Antioxid. Redox Signal. 00, 000-000.

Inflammation and Arterial Stiffness as Drivers of Cardiovascular Risk in Kidney Disease.

Patients with chronic kidney disease (CKD) have an increased cardiovascular (CV) risk. The lower the glomerular filtration rate, the higher the CV risk. Current data suggest that several uremic toxins lead to vascular inflammation and oxidative stress that, in turn, lead to endothelial dysfunction, changes in smooth muscle cells' phenotype, and increased degradation of elastin and collagen fibres. These processes lead to both functional and structural arterial stiffening and explain part of the increased risk of acute myocardial infarction and stroke reported in patients with CKD. Considering that, at least in patients with end-stage kidney disease, the reduction of arterial stiffness is associated with a parallel decrease of the CV risk, vascular function is a potential target for therapy to reduce the CV risk. in this review, we explore mechanisms of vascular dysfunction in CKD, paying particular attention to inflammation, reporting current data in other models of mild and severe inflammation, and discussing the vascular effect of several drugs currently used in nephrology.

Coordination among cytoskeletal organization, cell contraction, and extracellular matrix development is dependent on LOX for aneurysm prevention.

Distinct and seemingly independent cellular pathways affecting intracellular machinery or extracellular matrix (ECM) deposition and organization have been implicated in aneurysm formation. One of the key genes associated with this pathology in both humans and mice is lysyl oxidase (LOX), a secreted ECM-modifying enzyme, highly expressed in medial vascular smooth muscle cells. To dissect the mechanisms leading to aneurysm development, we conditionally deleted Lox in smooth muscle cells. We find that cytoskeletal organization is lost following Lox deletion. Cell culture assays and in vivo analyses demonstrate a cell-autonomous role for LOX affecting myosin light-chain phosphorylation and cytoskeletal assembly resulting in irregular smooth muscle contraction. These results not only highlight new intracellular roles for LOX, but notably, they provide a link between multiple processes leading to aneurysm formation, suggesting LOX coordinates ECM development, cytoskeletal organization, and cell contraction required for media development and function.

SMAD5 as a novel gene for familial pulmonary arterial hypertension.

Genetic diagnostic testing of 325 pulmonary arterial hypertension (PAH) patients using a PAH specific gene panel including 18 known PAH genes revealed mutations in 23 %. Further PAH candidate genes were sequenced in the remaining patients exposing two SMAD5 variants, which were clinically and functionally characterized. We first recorded familial cosegregation and clinical parameters. Functional tests were performed following transient over-expression of the two SMAD5 variants in pulmonary artery smooth muscle cells (PAMSCs). Expression of these variants was confirmed by qPCR, Sanger sequencing and Western blotting. Cell viability was evaluated using cell counting kit 8, cell proliferation by BrdU and apoptosis by annexin V assay. Both SMAD5 missense variants were absent in healthy controls and predicted to be pathogenic. The variant c.1175T>C p.(Leu392Pro) was identified in a heritable PAH patient and her healthy son. The mother had died of suspected PAH aged 42. Expression of this variant in PAMSCs led to significantly higher cell viability due to higher proliferation in comparison to SMAD5 wild type cells. The second variant c.277T>A p.(Trp93Arg) was identified in a patient with congenital heart disease associated PAH with a surgically repaired ventricle septal defect. Its expression led to significant lower cell viability due to increased apoptosis in comparison to wild type SMAD5 cells. Taking into account familial aggregation, clinical findings and functional evidence both variants could be classified as likely pathogenic. This is the first description of SMAD5 as a potential novel PAH gene for genetic diagnostic testing.

T follicular helper cell is essential for M2 macrophage polarization and pulmonary vascular remodeling in hypoxia-induced pulmonary hypertension

BackgroundHypoxia-induced pulmonary hypertension (HPH) is a subgroup of type 3 pulmonary hypertension that may cause early right ventricular failure and eventual cardiac failure, which lacks potential therapeutic targets. Our previous research demonstrated that T follicular helper (TFH) cells that produce IL-21 were involved in HPH. However, the molecular mechanisms of TFH/IL-21-mediated pathogenesis of HPH have been elusive. Here we investigate the role of TFH cells and IL-21 in HPH.MethodsStudies were performed in C57BL/6 mice or IL-21 knockout mice exposed to chronic hypoxia to induce PH, and examined by hemodynamics. Molecular and cellular studies were performed in mouse lung and pulmonary arterial smooth muscle cells (PASMCs). M2 signature gene (Fizz1), M1 signature genes (iNos, IL-12β and MMP9), GC B cell and its marker GL-7, caspase-1, M2 macrophages, TFH cells, Bcl-6 and IL-21 level were measured. Proliferation rate of PASMCs was measured by EdU. Pyroptosis was assessed using Hoechst 33,342/PI double fluorescent staining.ResultsIn response to chronic hypoxia exposure-induced pulmonary hypertension, IL-21−/− mice or downregulation of TFH cells in WT mice developed blunted pulmonary hypertension, attenuated pulmonary vascular remodelling. Furthermore, chronic hypoxia exposure significantly increased the germinal center (GC) B cell responses, which were not present in IL-21−/− mice or downregulation of TFH cells in WT mice. Importantly, IL-21 promoted the polarization of primary alveolar macrophages toward the M2 phenotype. Consistently, significantly enhanced expression of M2 macrophage marker Fizz1 were detected in the bronchoalveolar lavage fluid of HPH mice. Moreover, alveolar macrophages that had been cultivated with IL-21 promoted PASMCs proliferation and pyroptosis in vitro, while a selective CX3CR1 antagonist, AZD8797 (AZD), significantly attenuated the proliferation and pyroptosis of the PASMCs. Finally, ECM1 knockdown promoted IL-2–STAT5 signaling and inhibited Bcl-6 signaling to inhibit TFH differentiation in HPH.ConclusionsTFH/IL-21 axis amplified pulmonary vascular remodelling in HPH. This involved M2 macrophage polarization, PASMCs proliferation and pyroptosis. These data suggested that TFH/IL-21 axis may be a novel therapeutic target for the treatment of HPH.

SIX transcription factors are necessary for the activation of DUX4 expression in facioscapulohumeral muscular dystrophy

BackgroundFacioscapulohumeral muscular dystrophy (FSHD) is a common and progressive muscle wasting disease that is characterized by muscle weakness often first noticed in the face, the shoulder girdle and upper arms before progressing to the lower limb muscles. FSHD is caused by the misexpression of the Double Homeobox 4 (DUX4) transcription factor in skeletal muscle. While epigenetic derepression of D4Z4 macrosatellite repeats underlies DUX4 misexpression, our understanding of the complex transcriptional activation of DUX4 is incomplete.MethodsTo identify potential DUX4-regulatory factors, we used small interfering RNAs (siRNAs) to knockdown SIX family transcription factors (SIX1, 2, 4, 5) in patient-derived FSHD1 and FSHD2 myoblasts that were differentiated to form multinucleated myotubes. Quantitative real-time polymerase chain reaction was used to measure changes in DUX4 mRNA, DUX4 target gene expression and myogenic markers. Staining for SIX1 and SIX2 with specific antibodies was performed in FSHD myoblasts and myotubes. To assess reciprocal effects of DUX4 on SIX1, 2, and 4 expression, we utilized a doxycycline-inducible DUX4 myoblast cell line.ResultWe show that SIX1, 2 and 4 transcription factors, regulators of embryonic development, muscle differentiation, regeneration and homeostasis, are necessary for myogenic differentiation-dependent DUX4 expression in FSHD muscle cells. Using siRNA, we demonstrate SIX1, SIX2, and SIX4 to be critical factors involved in the induction of DUX4 transcription in differentiating FSHD myotubes in vitro. siRNA dual knockdown of SIX1 and SIX2 resulted in a ~ 98% decrease of DUX4 and DUX4 target genes, suggesting that SIX1 and SIX2 are the most critical in promoting DUX4 expression. Importantly, we show that DUX4 downregulates SIX RNA levels, suggesting negative feedback regulation.ConclusionsIn this study, we identified a family of developmental regulators that promote aberrant DUX4 expression in FSHD1 and FSHD2 differentiating muscle cells. Our findings highlight the critical involvement of SIX transcription factors (SIX1, 2, 4) in the pathogenesis of FSHD by serving as necessary factors that function in the promotion of DUX4 expression following epigenetic derepression of the D4Z4 repeats.

Single-cell view and a novel protective macrophage subset in perivascular adipose tissue in T2DM

BackgroundVasculopathy underlies diabetic complications, with perivascular adipose tissue (PVAT) playing crucial roles in its development. However, the changes in the cellular composition and function of PVAT, including the specific cell subsets and mechanisms implicated in type 2 diabetes mellitus (T2DM) vasculopathy, remain unclear.MethodsTo address the above issues, we performed single-cell RNA sequencing on the stromal vascular fraction (SVF) of PVAT from normal and T2DM rats. Then, various bioinformatics tools and functional experiments were used to investigate the characteristic changes in the cellular profile of diabetic PVAT SVF, their implications, and the underlying mechanisms.ResultsOur study reveals the single-cell landscape of the SVF of PVAT, demonstrating its considerable heterogeneity and significant alterations in T2DM, including an enhanced inflammatory response and elevated proportions of macrophages and natural killer (NK) cells. Moreover, macrophages are critical hubs for cross-talk among various cell populations. Notably, we identified a decreased Pdpn+ macrophage subpopulation in the PVAT of T2DM rats and confirmed this in mice and humans. In vitro and in vivo studies demonstrated that Pdpn+ macrophages alleviated insulin resistance and modulated adipokine/cytokine expression in adipocytes via the Pla2g2d-DHA/EPA-GPR120 pathway. This subset also enhances the function of vascular endothelial and smooth muscle cells, inhibits vascular inflammation and oxidative stress, and improves vasodilatory function, thereby protecting blood vessels.ConclusionPdpn+ macrophages exhibit significant vascular protective effects by alleviating insulin resistance and modulating adipokine/cytokine expression in PVAT adipocytes. This macrophage subtype may therefore play pivotal roles in mitigating vascular complications in T2DM. Our findings also underscore the critical role of immune-metabolic cross-talk in maintaining tissue homeostasis.Graphical

FSTL1 protects against acute aortic dissection by suppressing vascular smooth muscle cell phenotypic switching and degradation of the extracellular matrix

Acute aortic dissection (AAD) is a life-threatening cardiovascular emergency, which is closely related to the vascular smooth muscle cells (VSMCs) phenotypic switching and extracellular matrix (ECM) degradation. Previous studies have found that the secreted extracellular glycoprotein Follistatin-like 1 (FSTL1) is demonstrated as a protective factor for cardiovascular diseases. However, the role of FSTL1 in AAD remains elusive. We aimed to investigate whether FSTL1 could regulate VSMCs phenotypic switching and ECM degradation in AAD. Firstly, we found that FSTL1 expression in aorta was significantly decreased in human AAD examined by western blot and immunohistochemical staining. Then we established a mouse AAD model by administering β-aminopropionitrile (BAPN) dissolved in drinking water for 28 days. We found that FSTL1 expression in aorta was also decreased in mouse AAD. Exogenous supplement with recombinant human FSTL1 protein could rescue VSMCs phenotypic switching and ECM degradation to reduce the occurrence and progression of mouse AAD. In vitro, FSTL1 protein and adenovirus overexpressing FSTL1 (ad-FSTL1) reversed the primary VSMCs phenotypic switching and decreased the expression of MMP2 induced by PDGF-BB. Knocking down FSTL1 initiates VSMCs phenotypic switching and increases the expression of MMP2. In terms of mechanisms, AMPK phosphorylation was decreased and could be improved by FSTL1 protein in mouse AAD. FSTL1 protein and ad-FSTL1 reversed the decreased AMPK phosphorylation induced by PDGF-BB in primary VSMCs. These findings indicate that FSTL1 protects against VSMCs phenotypic switching and ECM degradation in AAD, and targeting FSTL1 may be a potential new strategy for prevention and treatment of AAD.

Dynamic three dimensional environment for efficient and large scale generation of smooth muscle cells from hiPSCs

BackgroundChronic ischemic limb disease often leads to amputation, which remains a significant clinical problem. Smooth-muscle cells (SMCs) are crucially involved in the development and progression of many cardiovascular diseases, but studies with primary human SMCs have been limited by a lack of availability. Here, we evaluated the efficiency of two novel protocols for differentiating human induced-pluripotent stem cells (hiPSCs) into SMCs and assessed their potency for the treatment of ischemic limb disease.MethodshiPSCs were differentiated into SMCs via a conventional two-dimensional (2D) protocol that was conducted entirely with cell monolayers, or via two protocols that consisted of an initial five-day three-dimensional (3D) spheroid phase followed by a six-day 2D monolayer phase (3D + 2D differentiation). The 3D phases were conducted in shaker flasks on an orbital shaker (the 3D + 2D shaker protocol) or in a PBS bioreactor (the 3D + 2D bioreactor protocol). Differentiation efficiency was evaluated via the expression of SMC markers (smooth-muscle actin [SMA], smooth muscle protein 22 [SM22], and Calponin-1), and the biological activity of the differentiated hiPSC-SMCs was evaluated via in-vitro assessments of migration (scratch assay), contraction in response to the treatment with a prostaglandin H2 analog (U46619), and tube formation on Geltrex, as well as in-vivo measurements of perfusion (fluorescence angiography) and vessel density in the limbs of mice that were treated with hiPSC-SMCs after experimentally induced hind-limb ischemia (HLI).ResultsBoth 3D + 2D protocols yielded > 5.6 × 107 hiPSC-SMCs/differentiation, which was ~ nine-fold more than that produced via 2D differentiation, and flow cytometry analyses confirmed that > 98% of the 3D + 2D-differentiated hiPSC-SMCs expressed SMA, > 81% expressed SM22, and > 89% expressed Calponin-1. hiPSC-SMCs obtained via the 3D + 2D shaker protocol also displayed typical SMC-like migratory, contraction, and tube-formation activity in-vitro and significantly improved measurements of perfusion, vessel density, and SMA-positive arterial density in the ischemic limb of mouse HLI model.ConclusionsOur dynamic 3D + 2D protocols produced an exceptionally high yield of hiPSC-SMCs. Transplantation of these hiPSC-SMCs results in significantly improved recovery of ischemic limb after ischemic injury in mice.

Functional Efficacy of Tissue-Engineered Small-Diameter Nanofibrous Polyurethane Vascular Grafts Surface-Modified by Methacrylated Sulfated Alginate in the Rat Abdominal Aorta.

Improved design to imitate natural vascular scaffolds is critical in vascular tissue engineering (VTE). Smooth muscle cells originating from surrounding tissues require larger pore sizes relative to those of endothelial progenitor cells found in the bloodstream. Furthermore, biofunctionalized scaffolds mimic the microenvironment, cellular function, and tissue morphogenesis. Here, we fabricated macroporous and nanofibrous polyurethane (PU) bilayer tissue-engineered vascular grafts (TEVGs) by a salt-leaching method to achieve high porosities up to 30 μm. These grafts have a low porosity on the luminal side and a high porosity on the abluminal side. To enhance their properties, we surface-modified the PU scaffolds using heparin-mimicking methacrylated sulfated alginate (PU-MSA). We then evaluated these tubular scaffolds for their anticoagulation effect, protein adsorption, and cell attachment in vitro. The results revealed that TEVGs modified with sulfated alginate (PU-MSA) exhibited better anticoagulation (25 ± 1 min) and higher VEGF protein adsorption (75 ± 5 ng/mL) compared to other scaffolds. Moving to in vivo testing, we examined the TEVGs in a rat model for either 1 or 5 months. Through ultrasonication and various histological analyses, we assessed the functionality and biocompatibility of the TEVGs. Notably, the PU-MSA scaffold created a microenvironment conducive to cell homing and regeneration in the field of VTE.

RO4929097 inhibits NICD3 to alleviate pulmonary hypertension via blocking Notch3/HIF-2α/FoxM1 signaling pathway.

Pulmonary hypertension (PH) is a condition in which the smooth muscle cells (SMCs) in the pulmonary arteries multiply excessively, causing the arteries to narrow. This can ultimately result in right heart failure and premature death. Notch3 is an important factor involved in pulmonary vascular remodeling in PH. RO4929097, as a γ-secretase inhibitor that inhibits Notch3 signaling pathway, may be a potential drug for the treatment of PH, but its feasibility and related mechanism of action need to be further investigated. In vitro modeling by hypoxic incubation of human pulmonary artery SMCs (HPASMCs). RO4929097 and plasmids including overexpression-NICD3 (oe-NICD3) and NICD3 small interfering RNA (siRNA) were used to alter the expression of NICD3, and HIF-2α inhibitor PT-2385 was used to alter the expression of HIF-2α. Western blot, EdU incorporation assay was used to investigate the alteration of NICD3, HIF-2α, FoxM1 protein expression, and cell proliferation. The severity of PH in rats was assessed by measuring the weight ratio of right ventricle (RV) to left ventricle (LV) and septum (S) (RV/[LV + S]) and hematoxylin-eosin (H&E) staining of lung tissues in a hypoxia-induced PH rat model. We first determined that hypoxia induction for 48h had the strongest induction of NICD3 and Notch3 in HPASMCs, and the strongest inhibition by 10μM RO4929097. Treatment of HPASMCs under hypoxic conditions with RO4929097 inhibited hypoxia-induced expression of NICD3, HIF-2α, FoxM1, and proliferation of HPASMCs. The inhibitory effect of RO4929097 was reversed after overexpression of NICD3 in HPASMCs. Further, we found that PT-2385 reversed the promotional effect of overexpression of NICD3 on the proliferation of HPASMCs. In vivo experiments, hypoxia-induced PH rats treated with RO4929097 showed a reduction in right ventricular hypertrophy index (RVHI) and a return to normal pulmonary artery morphology, indicating a reduction in the severity of PH. Our data suggest that RO4929097 regulates the Notch3/HIF-2α/FoxM1 signaling pathway by inhibiting the expression of NICD3, thereby inhibiting hypoxia-induced proliferation of HPASMCs. In vivo experiments also confirmed that RO4929097 could alleviate PH as a potential therapeutic strategy.

Exploring aortic stiffness in aging mice: a comprehensive methodological overview.

Stiffening of the vascular network is associated with the early stages of vascular aging, leading to cardiovascular disorders (hypertension), renal failures, or neurodegenerative diseases (Alzheimer's). Unfortunately, many people remain undiagnosed because diagnostic methods are either unsuitable for a large population or unfamiliar to clinicians which favor the hypertension evaluation. In preclinical research, stiffness studies are often partially conducted. We think that the evaluation of aortic stiffness is essential as it would improve our understanding of aging diseases progression. We propose here a systematic method using decision trees in a multi-scale and multimodal approaches. Our method was evaluated by analyzing the aortic situation in old and young mice. We demonstrate that both the endothelial and smooth muscle cells exhibit pronounced functional alterations in favor of constriction. Additionally, there is significant remodeling of the extracellular matrix, leading to a drastic degradation of elastic fibers and the accumulation of collagen in the aortic wall. This series of changes contributes to the development of vascular rigidity, a preliminary stage of arterial hypertension. Our results suggest that our method should improve preclinical understanding and encourage clinicians to equip themselves with tools for assessing vascular function, as it is an essential issue for preventing numerous pathologies.

Retraction: Nesfatin-1 regulates the phenotype transition of cavernous smooth muscle cells by activating PI3K/AKT/mTOR signaling pathway to improve diabetic erectile dysfunction.

Retraction: Nesfatin-1 regulates the phenotype transition of cavernous smooth muscle cells by activating PI3K/AKT/mTOR signaling pathway to improve diabetic erectile dysfunction.

Homozygous slc25a20 zebrafish mutant reveals insights into carnitine-acylcarnitine translocase deficiency pathogenesis

The SLC25A20 gene encodes carnitine-acylcarnitine translocase (CACT), facilitating the transport of long-chain acylcarnitine required for energy production via β-oxidation into the mitochondria. Loss-of-function mutations in this gene lead to CACT deficiency, a rare autosomal recessive disorder of fatty acid metabolism characterized by severe symptoms including cardiomyopathy, hepatic dysfunction, rhabdomyolysis, hypoketotic hypoglycemia, and hyperammonemia, often resulting in neonatal mortality. Here, we utilized CRISPR/Cas9 gene editing to isolate slc25a20 mutant zebrafish. Homozygous mutants displayed significant lethality, with the majority succumbing before reaching maturity. However, we identified a notably rare homozygous individual that survived into adulthood, prompting a histological examination. Firstly, we observed adipose tissue accumulation at various sites in the homozygous mutant. The mutant heart exhibited hypertrophy, along with degenerated myocardial and muscle cells containing numerous eosinophilic nuclei. Additionally, we found no large oil droplet vacuoles in the mutant liver; however, the hepatocytes displayed numerous small vacuoles resembling lipid droplets. Iron deposition was evident in the spleen and parts of the liver. Overall, our slc25a20 zebrafish mutant displayed tissue pathologies analogous to human CACT deficiency, suggesting its potential as a pathological model contributing to the elucidation of pathogenesis and the improvement/development of therapies for CACT deficiency.

Theabrownin/whey protein isolate complex coacervate strengthens C2C12 cell proliferation via modulation of energy metabolism and mitochondrial apoptosis.

Theabrownin (TB)-whey protein isolate (WPI) complex coacervates (TW) were firstly prepared to investigate the regulatory effects on skeletal muscle. The binding of TB to WPI reached saturation with the strongest electrostatic interaction at the ratio of 10:1. The formation of TW was driven by electrostatic interactions with the aid of hydrogen bonding and hydrophobic interactions, and the digestion behavior of TW was investigated based on in vitro gastrointestinal and CaCO2 cell models. The regulatory effect of TW on muscle cells was investigated by C2C12 cell assay. Cell cycle analysis showed that TW promoted the transition of skeletal muscle cells from proliferative state to differentiated state. Immunofluorescence and gene expression revealed that TW positively regulated myogenic regulatory factors, contributing to myofiber formation. Moreover, TW activated the intracellular TCA cycling and oxidative phosphorylation, providing energy for skeletal muscle regeneration and repair. Mechanistically, TW inhibited the release of cytochrome C from mitochondria to cytoplasm through the Bcl-2/Cytochrome C/Cleaved-Caspase-3 pathway, exhibiting a protective effect on skeletal muscle cells. In the future, the molecular mechanism of TW enhancing skeletal muscle function should be validated through aging animal models and clinical trials and expand its therapeutic application for muscle health in functional food and dietary supplements.

A Deep Dive into PDE5 Inhibition: Innovative Discoveries via Virtual Screening

Background: PDE5 inhibitors have had a surge in popularity over the last decade owing to their efficacy in the treatment of erectile dysfunction, coronary vasculopathy, and pulmonary arterial hypertension. These inhibitors exhibit competitive binding with phosphodiesterase type 5 and inhibit the hydrolysis of cyclic guanosine monophosphate, hence elevating the levels of cGMP in smooth muscle cells and prolonging the duration of an erection. However, due to production costs and side effects, further research is needed to discover new PDE5 inhibitors. Objective: The study aimed to identify potent PDE5 inhibitors by employing the extensive application of computer-aided drug design. Methods: Three different databases, named Million Molecules Database, Natural Product Database, and NCI Database, have been screened, which has been followed by filtering based on various druglikeness rules, docking, ADME, toxicity, consensus molecular docking, and 100 ns molecular dynamics simulation. Results: Three compounds (ZINC05351336, ZINC12030898, and ZINC17949426) have exhibited stable-binding characteristics at the active site of PDE5, demonstrating a robust binding affinity. These molecules have been found to possess drug-like capabilities, effective ADME features, low toxicity, and high stability. Conclusion: The study has delved into the realm of PDE5 inhibitors, which have been found to be effective in treating erectile dysfunction, but high production costs and side effects necessitate new ones. Through computer-aided drug design and screening, three compounds have been identified with promising binding characteristics, drug-appropriate properties, effective ADME profiles, minimal toxicity, and stability, making them potential candidates for future PDE5 inhibitors

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ON THE INNER FRONT COVER: A genetic mosaic C. elegans adult stained for myosin A and UNC‐89 (vertebrate homolog is obscurin) in body‐wall muscle shows disorganized UNC‐89/obscurin in a muscle cell that does not express myosin A.Credit: Sarah Almuhanna (Imam Abdulrahman Bin Faisal University College of Medicine). image