Search Results (1,967)

Smooth muscle cell (SMC) phenotypic transition contributes to several major vascular diseases such as intimal hyperplasia and restenosis, atherosclerosis, and aneurysm. However, the molecular mechanisms underlying this process are not fully understood. The objectives of this study are to determine the role of mRNA N⁶-methyladenosine (m6A) modification in SMC phenotypic modulation and injury-induced neointima formation. By using an m6A quantification kit, we found that m6A levels are altered during the early stage of SMC phenotypic modulation. RNA sequencing revealed that m6A modifications in the mRNAs of 708 genes are elevated while modifications in the mRNAs of 300 genes are decreased. These modifications occur in genes widely distributed in most chromosomes and involved in many cellular processes and signaling/gene regulations. Meanwhile, the regulators for m6A modifications are altered by PDGF-BB, a known factor inducing SMC phenotypic modulation. Although m6A writers and erasers are not altered during SMC phenotypic modulation, m6A reader YTHDF1 is dramatically reduced as early as 12 h following PDGF-BB treatment, a time much earlier than the downregulation of SMC contractile proteins. Importantly, the overexpression of YTHDF1 reverses the expression of SMC contractile proteins, suggesting a restoration of contractile SMC phenotype. By using a rat carotid artery balloon-injury model, we found that injury significantly decreases YTHDF1 levels in the medial SMCs while inducing neointima formation. Of significance, restoring YTHDF1 expression through lentiviral transduction blocks injury-induced neointima formation. Moreover, YTHDF1 delivery restores the expression of SMC contractile proteins that is diminished in arterial media layers due to the injury. These data demonstrate that YTHDF1 plays a protective role in maintaining the contractile SMC phenotype and vascular homeostasis during injury-induced pathological vascular remodeling. Full article

Disruptions in uterine tissue function contribute to disorders such as endometriosis, adenomyosis, endometrial cancer, and fibroids, which all significantly impact health and fertility. Advances in transcriptomics, particularly single-cell RNA sequencing, have revolutionized uterine biological research by revealing the cellular heterogeneity and molecular mechanisms underlying disease states. Single-cell RNA sequencing and spatial transcriptomics have mapped endometrial and myometrial cellular landscapes, which helped to identify critical cell types, signaling pathways, and phase-specific dynamics. Said transcriptomic technologies also identified stromal and immune cell dysfunctions, such as fibroblast-to-myofibroblast transitions and impaired macrophage activity, which drive fibrosis, chronic inflammation, and lesion persistence in endometriosis. For endometrial cancer, scRNA-seq uncovered tumor microenvironmental complexities, identifying cancer-associated fibroblast subtypes and immune cell profiles contributing to progression and therapeutic resistance. Similarly, studies on adenomyosis highlighted disrupted signaling pathways, including Wnt and VEGF, and novel progenitor cell populations linked to tissue invasion and neuroinflammation, while single-cell approaches characterized smooth muscle and fibroblast subpopulations in uterine fibroids, elucidating their roles in extracellular matrix remodeling and signaling pathways like ERK and mTOR. Despite challenges such as scalability and reproducibility, single-cell transcriptomic approaches may have potential applications in biomarker discovery, therapeutic target identification, and personalized medicine in gynecological disorders. Full article

Arterial stiffening is a significant risk factor for the development of cardiovascular diseases, including hypertension, atherosclerosis, and arteriopathy. The destruction of elastic fibers, accompanied by vascular inflammatory remodeling, is a key process in the progression of arterial stiffening and related pathologies. In young, healthy arteries, intact elastic fibers create a resilient microenvironment that maintains the quiescence of arterial cells. However, with advancing age, these elastic fibers undergo post-translational modifications, such as oxidation, glycosylation, and calcification, leading to their eventual degeneration. This degeneration results in the release of degraded peptides and the formation of an inflammatory, stiffened niche. Elastic fiber degeneration profoundly impacts the proinflammatory phenotypes and behaviors of various arterial cells, including endothelial cells, smooth muscle cells, macrophages, fibroblasts, and mast cells. Notably, the degraded elastic fibers release elastin-derived peptides (EDPs), which act as potent inflammatory molecules. EDPs activate various arterial cellular processes, including inflammatory secretion, cell migration, proliferation, and calcification, by interacting with the elastin receptor complex (ERC). These elastin-related cellular events are commonly observed with aging and in diseased arteries. These findings suggest that the degeneration of the elastic fiber meshwork is a primary event driving arterial inflammation, stiffening, and adverse remodeling with advancing age. Therefore, preserving elastic fibers and blocking the EDP/ERC signaling pathways may offer promising therapeutic strategies for mitigating age-related arterial remodeling and related arterial diseases. Full article

Background/Objectives: Alzheimer’s disease (AD) and related dementias (ADRD) disproportionately impact racial and ethnic minorities. Contributing biological factors that explain this disparity have been elusive. Moreover, non-invasive biomarkers for early detection of AD are needed. Endothelin-1 (ET-1), a vasoconstrictive factor linked to cerebral vascular disease pathology and neuronal injury, could provide insights to better understand racial disparities in AD. As a potent vasoconstrictive peptide that regulates contractions in smooth muscle, endothelial cells, and pericytes, ET-1 may result in cerebral vascular constriction, leading to cerebral hypoperfusion; over time, this may result in neuronal injury, contributing to the pathology of AD. The role of the ET-1 system as a driver of ethnic disparities in AD requires further investigation. In the United States (U.S.), ET-1 dysregulation in Hispanic/Latinx (H/L) ethnic populations has largely been unexplored. Genetics linking ET-1 dysregulation and racial disparities in AD also require further investigation. In this study, we examined the role of the ET-1 protein in human plasma as a potential biomarker with predictive value for correlating with the development of AD by age, race, and sex. Methods: We examined ET-1 protein levels using quantitative mass spectrometry in AA and NHW patients with AD, along with controls. Results: A partial correlation between age at draw and ET-1, stratified by race and sex, while controlling for AD status, was significant for female AAs (r = 0.385, p = 0.016). When the data were not stratified but controlled for AD status, the partial correlation between age at draw and ET-1 was not significant (r = 0.108, p = 0.259). Conclusions: Based on the small number of plasma specimens and no plasma specimens from H/L individuals with AD, we conclude that ET-1 was clearly not a significant factor in predicting AD in this study and will require a larger scale study for validation. Full article

Objective: In developed countries, stroke is the fifth cause of death, with a high mortality rate, and with recovery to normal neurological function in one-third of survivors. Atherosclerotic occlusive disease of the extracranial part of the internal carotid artery and related embolic complications are common preventable causes of ischemic stroke (IS), attributable to 7–18% of all first-time cases. Osteoprotegerin (OPG), a soluble member of the tumor necrosis factor receptor (TNFR) superfamily, is considered a modulator of vascular calcification linked to vascular smooth muscle cell proliferation and collagen production in atherosclerotic plaques. Therefore, OPG emerges as a potential biomarker (BM) of calcified carotid plaques and carotid artery stenosis (CAS). Methods: We performed a literature search of PubMed on OPG in CAS and atherosclerosis published until 2024. Results: Increased levels of serum OPG were reported in both patients with symptomatic and asymptomatic CAS, and higher values were observed in those with unstable atherosclerotic plaques. Notably, increased OPG levels were observed regardless of the location of atherosclerosis, including coronary and other peripheral arteries. In addition, chronic kidney disease, the most significant confounder disturbing the association between vascular damage and circulating OPG levels, decreases the usefulness of OPG as a BM in CAS. Conclusions: Osteoprotegerin may be considered an emerging BM of global rather than cerebrovascular atherosclerosis. Its diagnostic significance in identifying patients with asymptomatic CAS and their monitoring is limited. Full article

We recently found that myometrial distension stimulates maternal uterine vascular remodeling, and hypothesized that this may be a previously unrecognized mechanism for inducing arterial growth during pregnancy. The aim of this study was to further characterize a recently developed surgical model in which medical-grade silicone was infused into one uterine horn of a non-gravid rat to induce acute myometrial stretch, followed by an additional, gradual distension due to the secretion of an exudate into the uterine lumen. Our objectives were to better understand the effects of stretch on the myometrium and to look for the expression of proangiogenic and proinflammatory factors that may stimulate uterine vascular remodeling. Morphometric analysis showed hypertrophy of the uterine corpus that was primarily due to axial growth since the myometrial cross-sectional area was unchanged due to a thinning of the uterine wall secondary to stretch. This finding was supported by significantly increased myometrial smooth muscle cell mitosis. There was also an increase in the concentration of myometrial elastin but not collagen. The analysis showed modest increases in neutrophils, activated and unactivated macrophages, and the proinflammatory cytokines RANTES, MIP-3α, GRO-KC, and TNFα. The most dramatic change was the extremely high level of VEGF in the exudate, which was increased >900× above circulating levels. Full article

Ghrelin (GhRL) is an orexigenic hormone influenced by nutritional state. It plays a role in skin repair and diseases, though little information exists regarding its function in this organ. GhRL and its receptor were investigated in the skin of sheep under different feeding conditions to explore GhRL system presence and possible modifications due to diet. Three-year-old female sheep were free to graze from June to the pasture maximum flowering (MxF group) and from this period to maximum dryness addicted (Exp group) or not (MxD group) with 600 gr/die/head of barley and corn. Skin samples were processed for immunohistochemistry and real-time PCR. The immunostaining showed the presence of the GhRL system in skin appendages. Indeed, the ligand was localized in the hair follicles whereas the receptor was also observed in sweat glands and smooth muscle cells. The expression of both genes was significantly higher in the Exp group (3.6 and 2.9 folds respectively, p < 0.05) compared with the MxF group. These results suggest that the GhRL system is involved in the regulation of hair follicles and sweat glands. In addition, diet supplementation may positively modulate the expression of GhRL and its receptor in the skin. Full article

(1) Background: For the reconstruction of a human vagina, various surgical procedures are available that are often associated with complications due to their failure to mimic the physiology of the human vagina. We recently developed a vascularized, organ-specific matrix from healthy human vaginal wall tissue with suitable biomechanical properties. A superior graft would require further extensive colonization with autologous vaginal cells to reduce complications upon implantation. However, reports on isolation of vaginal cells from biopsies are scarce, and published protocols rarely contain sufficient details. In this study, we aimed to examine protocols for inconsistencies and identify (where possible) the optimal protocol in terms of reproducibility and efficiency for isolation of human vaginal fibroblasts (FBs), epithelial cells (VECs), and smooth muscle cells (SMCs). Overall, this study aims to guide other researchers and aid future tissue engineering solutions that rely on autologous cells. (2) Methods: A total of 41 isolation protocols were tested: four protocols specific to FBs, 13 protocols for VECs, and 24 protocols for SMCs. Protocols were derived from published reports on cell isolation by enzymes, with exclusion criteria including the need for specialized equipment, surgical separation of tissue layers, or missing protocol details. Enzymatic digestion with collagenase-I, collagenase-IV, and dispase-II was used for isolation of VECs, collagenase-IV for isolation of SMCs, and collagenase-IA for isolation of FBs. Fluorescent immunostaining was applied to identify VECs with cytokeratin, SMCs with desmin, endothelial cells with UEA-1, and FBs with vimentin. Protocols were assessed based on (>95%) homogeneity, duplicate consistency, cell viability, and time to first passage. (3) Results: A total of 9 out of the 41 protocols resulted in isolation and expansion of vaginal FBs. This involved 1 out of 13 VEC protocols, 6 out of 24 SMC protocols, and 2 out of 2 FB protocols. Isolation of vaginal SMCs or VECs was not achieved. The best results were obtained after digestion with 0.1% collagenase-IV, where pure FB colonies formed with high cell viability. (4) Conclusions: Today, vaginoplasty is considered the gold standard for surgically creating a neovagina, despite its considerable drawbacks and limitations. Tissue-engineered solutions carry great potential as an alternative, but cell seeding is desired to prevent complications upon implantation of grafts. In this study, we examined isolation of human vaginal FBs, SMCs, and VECs, and identified the most efficient and reliable protocol for FBs. We further identified inconsistencies and irreproducible methods for isolation of VECs and SMCs. These findings aid the clinical translation of cell-based tissue engineering for the reconstruction and support of vaginas, fulfilling unmet medic needs. Full article

Background: Cholesterol gallstone disease (CGS) is often accompanied by gallbladder contraction dysfunction and chronic inflammation, but effective therapeutic options remain limited. This study investigates whether a low-intensity pulsed ultrasound (LIPUS) treatment can improve gallbladder motility and alleviate chronic inflammation while exploring the underlying mechanisms. Methods: Gallbladder motility was assessed through in vitro and in vivo contraction tests, while bile condition was evaluated by observing bile crystal clearance. Tissue analysis and Western blotting were performed to examine the expression of the cholecystokinin A receptor (CCKAR) and α-smooth muscle actin (α-SMA) as markers of gallbladder smooth muscle health and the inflammatory microenvironment. Blood cholesterol levels were measured via biochemical assays. Results: LIPUS treatment obviously enhanced gallbladder contractility in response to CCK-8 stimulation and accelerated bile crystal clearance. It also reduced inflammatory cell infiltration and tissue edema, and promoted new capillary formation in the gallbladder, mitigating the progression of CGS. Furthermore, LIPUS restored CCKAR expression and improved the thickness of the gallbladder smooth muscle layer, providing a structural basis for increased smooth muscle contractility. Conclusion: LIPUS improves gallbladder motility and reduces chronic inflammation in CGS by enhancing CCKAR expression and smooth muscle integrity. These findings highlight the potential of LIPUS as a non-invasive therapeutic approach for managing CGS. Full article

Vascular smooth muscle cell (SMC) relaxation by guanylyl cyclases (GCs) and cGMP is mediated by NO and its receptor soluble GC (sGC) or natriuretic peptides (NPs) ANP/BNP and CNP with the receptors GC-A and GC-B, respectively. It is commonly accepted that cultured SMCs differ from those in intact vessels. Nevertheless, cell culture often remains the first step for signaling investigations and drug testing. Previously, we showed that even popular reference genes changed dramatically after SMC isolation from aorta. Regarding NP receptors, a substantial amount of data relies on cell culture. We hypothesize that the NP/cGMP system in intact aortic tunica media differs from isolated and cultured aortic SMCs. Therefore, we studied isolation and culturing effects on the expression of NP receptors GC-A, GC-B, and NP clearance receptor (NPRC) compared to sGC. We investigated intact tunica media and primary SMCs from the longitudinal halves of the same rat aorta. GC activity was monitored by cyclic guanosine monophosphate (cGMP). In addition, we hypothesize that there are sex-dependent differences in the NP/cGMP cascade in both intact tissue and cultured cells. We, therefore, analyzed a male and female cohort. Expression was quantified by RT-qPCR comparing aortic media and SMCs with our recently validated reference gene (RG) small nuclear ribonucleoprotein 2 (U2). Only GC-A was stably expressed. In intact media, GC-A exceeded GC-B and NPRC. However, GC-B, NPRC, and sGC were dramatically upregulated in cultured SMCs of the same aortae different from the stable GC-A. The expression was mirrored by NP-induced GC activity. In cultured cells, changes in GC activity were delayed compared to receptor expression. Minor differences between both sexes could also be revealed. Thus, isolation and culture fundamentally alter the cGMP system in vascular SMCs with potential impact on drug testing and scRNAseq. Especially, the dramatic increase in the clearance receptor NPRC in culture might distort all physiological ANP, BNP, and CNP effects. Full article

Background: Chronic obstructive pulmonary disease (COPD) is characterized by progressive and incurable airflow obstruction and chronic inflammation. Both TGF-β1 and CXCL8 have been well described as fundamental to COPD progression. DNA methylation and histone acetylation, which are well-understood epigenetic mechanisms regulating gene expression, are associated with COPD progression. However, a deeper understanding of the complex mechanisms associated with DNA methylation, histone post-translational changes and RNA methylation in the context of regulatory pathways remains to be elucidated. We here report on how DNA methylation and histone acetylation inhibition differentially affect CXCL8 signaling in primary human non-COPD and COPD airway cells. Methods: Airway smooth muscle (ASM) cells, a pivotal cell type in COPD, were isolated from the small airways of heavy smokers with and without COPD. Histone acetylation and DNA methylation were inhibited before the TGF-β1 stimulation of cells. Subsequently, CXCL8 production and the abundance and activation of pertinent transcription regulatory proteins (NF-κB, p38 MAPK and JNK) were analyzed. Results: TGF-β1-stimulated CXCL8 release from ASM cells from ‘healthy’ smoker subjects was significantly modulated by DNA methylation (56.32 pg/mL and 56.60 pg/mL) and acetylation inhibitors (27.50 pg/mL and 48.85 pg/mL) at 24 and 48 h, respectively. However, modulation via the inhibition of DNA methylation (34.06 pg/mL and 43.18 pg/mL) and acetylation (23.14 pg/mL and 27.18 pg/mL) was observed to a lesser extent in COPD ASM cells. These changes were associated with differences in the TGF-β1 activation of NF-κB and MAPK pathways at 10 and 20 min. Conclusions: Our findings offer insight into differential epigenetics in controlling COPD ASM cells and provide a foundation warranting future studies on epigenetic differences associated with COPD diagnosis. This would provide a scope for developing therapeutic interventions targeting signaling and epigenetic pathways to improve patient outcomes. Full article

Oxidative stress is the key cause of the etiopathogenesis of several diseases associated with constipation. This study examined whether the green pine cone can improve the symptoms of constipation based on the antioxidant activities. The changes in the key parameters for the antioxidant activity and laxative effects were examined in the loperamide (Lop)-induced constipation of Sprague-Dawley (SD) rats after being treated with the methanol extracts of green pine cone (MPC, unripe fruits of Pinus densiflora). MPC contained several bioactive compounds, including diterpenoid compounds such as dehydroabietic acid, taxodone, and ferruginol. In addition, it exhibited high scavenging activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radicals. These effects of MPC successfully reflected the improvement in nicotinamide adenine dinucleotide phosphate oxidase (NADP) H oxidase transcription, superoxide dismutase (SOD) levels, and nuclear factor erythroid 2-related factor 2 (Nrf2) phosphorylation levels in the mid colon of Lop+MPC-treated SD rats. Furthermore, significant improvements in the stool parameters, gastrointestinal (GI) transit, intestine length, and histopathological structure of the mid colon were detected in the Lop-induced constipation rats after MPC treatment. The other parameters, including the regulators for the adherens junction (AJ) and tight junction (TJ), and GI hormone secretion for laxative effects, were improved significantly in Lop+MPC-treated SD rats. These effects were also verified in Lop+MPC-treated primary rat intestine smooth muscle cells (pRISMCs) through analyses for antioxidant defense mechanisms. Overall, the finding of this study offers novel scientific evidence that MPC could be considered as a significant laxative for chronic constipation based on its antioxidant activity. Full article

Background: Atherosclerotic calcification (AC) is a common feature of atherosclerotic cardiovascular disease. β-Hydroxybutyrate (BHB) has been identified as a molecule that influences cardiovascular disease. However, whether BHB can influence AC is still unknown. Methods and Results: In this study, ApoE^−/− mice, fed a Western diet, were used to examine the effects of BHB on AC. Rat vascular smooth muscle cells (VSMCs) were used to verify the impacts of BHB on AC and to explore the underlying mechanisms. The results show that Western diet-challenged ApoE^−/− mice, supplemented with BHB for 24 weeks, exhibited reduced calcified areas, calcium content, and alkaline phosphatase (ALP) activity in the aortas, as well as ameliorated severity of AC. Furthermore, BHB downregulated the expression of glucose-regulated protein 78 (GRP78) and C/EBP homologous protein (CHOP), thereby reducing endoplasmic reticulum stress (ERS) and ERS-mediated apoptosis in the aortas of the mice. Consistently, in vitro studies showed that BHB reduced ALP activity and calcium content in VSMCs, and inhibited VSMC calcification. Additionally, BHB suppressed ERS-mediated apoptosis in VSMCs. Conclusions: In summary, the present results demonstrate that BHB can alleviate atherosclerotic calcification by inhibiting ERS-mediated apoptosis. Therefore, BHB may serve as a viable therapeutic agent for AC. Full article

The prevalence of cardiovascular disease varies with sex, and the impact of intrinsic sex-based differences on vasculature is not well understood. Animal models can provide important insights into some aspects of human biology; however, not all discoveries in animal systems translate well to humans. To explore the impact of chromosomal sex on proteomic phenotypes, we used iPSC-derived vascular smooth muscle cells from healthy donors of both sexes to identify sex-based proteomic differences and their possible effects on cardiovascular pathophysiology. Our analysis confirmed that differentiated cells have a proteomic profile more similar to healthy primary aortic smooth muscle cells than iPSCs. We also identified sex-based differences in iPSC-derived vascular smooth muscle cells in pathways related to ATP binding, glycogen metabolic process, and cadherin binding as well as multiple proteins relevant to cardiovascular pathophysiology and disease. Additionally, we explored the role of autosomal and sex chromosomes in protein regulation, identifying that proteins on autosomal chromosomes also show sex-based regulation that may affect the protein expression of proteins from autosomal chromosomes. This work supports the biological relevance of iPSC-derived vascular smooth muscle cells as a model for disease, and further exploration of the pathways identified here can lead to the discovery of sex-specific pharmacological targets for cardiovascular disease. Full article

The octapeptide angiotensin II (Ang II) is a circulating hormone as well as a locally formed agonist synthesized by the angiotensin-converting enzyme (ACE) of endothelial cells. It forms a powerful mechanism to control the amount and pressure of body fluids. All main effects are directed to save body salt and water and ensure blood pressure under basic conditions and in emergencies. All blood vessels respond to stimulation by Ang II; the immediate response is smooth muscle contraction, increasing vascular resistance, and elevating blood pressure. Such effects are conveyed by type 1 angiotensin receptors (AT₁Rs) located in the plasma membrane of both endothelial and vascular smooth muscle cells. AT₁Rs are heterotrimeric G protein-coupled receptors (GPCRs), but their signal pathways are much more complicated than other GPCRs. In addition to G_q/11, the G_12/13, JAK/STAT, Jnk, MAPK, and ERK 1/2, and arrestin-dependent and -independent pathways are activated because of the promiscuous attachment of different signal proteins to the intracellular G protein binding site and to the intracellular C terminal loop. Substantial changes in protein expression follow, including the intracellular inflammation signal protein NF-κB, endothelial contact proteins, cytokines, matrix metalloproteinases (MMPs), and type I protocollagen, eliciting the inflammatory transformation of endothelial and vascular smooth muscle cells and fibrosis. Ang II is an important contributor to vascular pathologies in hypertensive, atherosclerotic, and aneurysmal vascular wall remodeling. Such direct vascular effects are reviewed. In addition to reducing blood pressure, AT₁R antagonists and ACE inhibitors have a beneficial effect on the vascular wall by inhibiting pathological wall remodeling. Full article