Muscle Cell Growth Research Articles

Production of cultured meat by culturing porcine smooth muscle cells in vitro with food grade peanut wire-drawing protein scaffold

Cultivating meat is a promising solution to the negative problems brought by traditional animal husbandry. To make cultured meat have the sensory and nutritional characteristics of conventional meat as much as possible, many studies have been conducted on various cell types and scaffold characteristics. Therefore, this study aims to produce a low-cost cultured meat with a quality closer to that of conventional meat. Tissue generation requires three-dimensional (3D) scaffolds to support cells and simulate extracellular matrix (ECM). Here, we used peanut wire-drawing protein (a biomaterial based on edible porous protein) as a new culture meat scaffold to culture cells. The scaffold can support cell attachment and proliferation to create 3D engineered porcine muscle tissue. The differentiation of smooth muscle cells (SMCs) was induced by a low serum medium to produce more extracellular matrix proteins. After differentiation, it was found that peanut wire-drawing protein scaffolds could be used for porcine smooth muscle cell adhesion and growth. The ECM protein and muscle protein produced by SMCs can endow cultured meat with better quality. This technology provides an innovative pathway for the industrialized production of cultured meat.

Gut Microbiota-Derived Indole-3-Carboxylate Influences Mucosal Integrity and Immunity Through the Activation of the Aryl Hydrocarbon Receptors and Nutrient Transporters in Broiler Chickens Challenged With Eimeria maxima.

Two studies were conducted to evaluate the effects of indole-3-carboxylate (ICOOH) as a postbiotic on maintaining intestinal homeostasis against avian coccidiosis. In the first study, an in vitro culture system was used to investigate the effects of ICOOH on the proinflammatory cytokine response of chicken macrophage cells (CMCs), gut integrity of chicken intestinal epithelial cells (IECs), differentiation of quail muscle cells (QMCs), and primary chicken embryonic muscle cells (PMCs) and anti-parasitic effect against Eimeria maxima. Cells to be tested were seeded in the 24-well plates and treated with ICOOH at concentrations of 0.1, 1.0, and 10.0 µg. CMCs were first stimulated by lipopolysaccharide (LPS) to induce an innate immune response, and QMCs and PMCs were treated with 0.5% and 2% fetal bovine serum, respectively, before they were treated with ICOOH. After 18 h of incubation, cells were harvested, and RT-PCR was performed to measure gene expression of proinflammatory cytokines of CMCs, tight junction (TJ) proteins of IECs, and muscle cell growth markers of QMCs and PMCs. In the second study, in vivo trials were carried out to study the effect of dietary ICOOH on disease parameters in broiler chickens infected with E. maxima. One hundred twenty male broiler chickens (0-day-old) were allocated into the following four treatment groups: 1) basal diet without infection (CON), 2) basal diet with E. maxima (NC), 3) ICOOH at 10.0 mg/kg feed with E. maxima (HI), and 4) ICOOH at 1.0 mg/kg feed with E. maxima (LO). Body weights (BWs) were measured on 0, 7, 14, 20, and 22 days. All groups except the CON chickens were orally infected with E. maxima on day 14. Jejunal samples were collected for lesion score and the transcriptomic analysis of cytokines and TJ proteins. In vitro, ICOOH increased the expression of TJ proteins in IECs and decreased IL-1β and IL-8 transcripts in the LPS-stimulated CMCs. In vivo, chickens on the HI diet showed reduced jejunal IL-1β, IFN-γ, and IL-10 expression and increased expression of genes activated by aryl hydrocarbon receptors and nutrient transporters in E. maxima-infected chickens. In conclusion, these results demonstrate the beneficial effects of dietary ICOOH on intestinal immune responses and barrier integrity in broiler chickens challenged with E. maxima. Furthermore, the present finding supports the notion to use microbial metabolites as novel feed additives to enhance resilience in animal agriculture.

HOXA cluster antisense RNA 2 elevates KIAA1522 expression through microRNA-520d-3p and insulin like growth factor 2 mRNA binding protein 3 to promote the growth of vascular smooth muscle cells in thoracic aortic aneurysm.

Recently, long non-coding RNAs (lncRNAs) have been revealed to mediate smooth muscle dysfunction in thoracic aortic aneurysm (TAA). LncRNA HOXA-AS2 has been proposed to engage in the regulation of diverse diseases. However, its function in TAA remains unknown. This study aimed to reveal the role and mechanism of HOXA-AS2 in VSMCs which were implicated in TAA formation. RT-qPCR or western blot was performed to detect RNA or protein expression levels. The role of HOXA-AS2 in VSMCs was explored by functional assays. The relationship among HOXA-AS2/miR-520d-3p/KIAA1522/IGF2BP3 was analysed via mechanism assays. HOXA-AS2 was detected to have significantly high expression in TAA tissues and function as an oncogene to promote proliferation of VSMCs, while inhibiting cell apoptosis (Figure 1, **P<0.01). HOXA-AS2 was unveiled to bind with miR-520d-3p (Figure 2, *P<0.05, **P<0.01) and further up-regulate KIAA1522 to facilitate the growth of VSMCs (Figure 3-4, *P<0.05, **P<0.01). HOXA-AS2 was also found to recruit IGF2BP3 to stabilize KIAA1522 mRNA (Figure 5, **P<0.01). All data were displayed as mean±standard deviation. HOXA-AS2 up-regulates KIAA1522 through targeting miR-520d-3p/IGF2BP3 to drive VSMC growth in TAA.

Circular RNA circ_0021001 regulates miR-148b-3p/GREM1 axis to modulate proliferation and apoptosis of vascular smooth muscle cells.

Intracranial aneurysm (IA) is an abnormal expression in the intracranial arteries, which is related to the growth and apoptosis of vascular smooth muscle cells (VSMCs). Circular RNA (circRNA) circ_0021001 (also named circARFIP2) has been identified to mediate the regulation of VSMCs proliferation. However, the molecular mechanism of circ_0021001 involved in VSMC dysfunction in IA is poorly defined.The expression levels of circ_0021001, microRNA-148b-3p (miR-148b-3p), and Gremlin 1 (GREM1) were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Cell viability, proliferation, cell cycle progression, and apoptosis were detected by Cell Counting Kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), and flow cytometry assays. Protein levels of proliferating cell nuclear antigen (PCNA), p21, B-cell lymphoma-2 (Bcl-2), Bcl-2 related X protein (Bax), and GREM1 were examined by western blot assay. The binding relationship between miR-148b-3p and circ_0021001 or GREM1 was predicted by StarBase and then verified using a dual-luciferase reporter assay.The expression levels of circ_0021001 and GREM1 were increased, and that of miR-148b-3p was decreased in IA tissues and HUASMCs. Moreover, the downregulation of circ_0021001 could repress proliferation ability and induce apoptosis of HUASMCs. The mechanical analysis uncovered that circ_0021001 served as a sponge of miR-148b-3p to regulate GREM1 expression.Circ_0021001 silencing could suppress cell growth and induce apoptosis of HUASMCs partially through modulating the miR-148b-3p/GREM1, presented circ_0021001 as a promising therapeutic target for IA.

Micro-Vibration Environment Promotes Bone Marrow Mesenchymal Stem Cells (BMSCs) Healing of Fracture Ends and Matrix Metalloproteinase-9 (MMP-9) Expression

When the muscle is stimulated by micro-vibration environment, it will produce a mechanical effect on skeletal muscle, thereby promoting growth of skeletal muscle cells. Bone marrow mesenchymal stem cells (BMSCs), as mechanically sensitive cells, have ability to multipolarize and multiple tropisms. This experiment explores the effect of BMSC cells on fracture end healing in fracture rats in a micro-vibration environment, and further explores whether it promotes the healing of fracture end to provide biological treatment ideas for the clinical repair of fracture patients. After modeling, SD rats were assigned into blank group, control group, and experimental group (treated with BMSCs) followed by analysis of bone volume fraction and bone trabecular thickness and number by Micro-CT, callus growth by H&amp;E staining, and expression of p38 and MMP-9 by immunohistochemical staining. The BV/TV value of experimental group was (0.41±0.06), Tb.Th value (0.08±0.01), Tb.N value (3.96±0.48) and was higher than other two groups (P &lt; 0.05). The growth of capillaries, trabecular bone, fibrous callus and cartilaginous callus in experimental group showed increased growth and the calcification was observed at the edge of cartilage callus. In addition, experimental group showed increased distribution area of MMP-9 and elevated expression of MMP-9 and p38MAPK. In conclusion, the micro-vibration environment can effectively promote the chemotaxis of BMSC cells to the fracture site to activate ossification, thereby promoting the proliferation and ossification and differentiation of BMSC, and further promoting the repair of fracture ends.

Epigenetics: Through the pediatric urology looking glass

Epigenetics: Through the pediatric urology looking glass

Impaired expression of BCAT1 relates to muscle atrophy of mouse model of sarcopenia

BackgroundThe underlying mechanism of muscle atrophy in sarcopenia is still not fully understood; branched chain aminotransferase 1(BCAT1) isocitrate dehydrogenase-1 encodes an evolutionarily conserved cytoplasmic aminotransferase for glutamate and branched-chain amino acids (BCAAs), thus constituting a regulatory component of cytoplasmic amino and keto acid metabolism. In human gliomas carrying wild-type isocitrate dehydrogenase-1, BCAT1 promotes cell proliferation through amino acid catabolism. Hence, the goals of this study were to unravel the potential role of BCAT1 expression in muscle atrophy and to explore the mechanisms underlying this process.MethodsWe first measured Bcat1 expression by RT-qPCR and western blotting in murine and cellular models of muscle atrophy. To understand how the Bcat1-driven changes sustained muscle cell growth, we analyzed reactive oxygen species (ROS) levels and activation of the mTORC1/S6K1 pathway in muscle cells. Furthermore, we performed Cell Counting Kit-8(CCK8) assays and fluorescence staining to evaluate growth rate of cells and ROS levels. Finally, we verified that depletion of Bcat1 impairs the growth rate of muscle cells and increases ROS levels, indicating that muscle atrophy resulted from the downregulation of the mTORC1/S6K1 pathway. Data were analyzed by two-tailed unpaired Student’s t-test or Mann-Whitney U test for two groups to determine statistical significance. Statistical analyses were performed using GraphPad Prism version 6.0 and SPSS 16.0 software.ResultsBcat1 expression level in skeletal muscles was lower in murine and cellular models of sarcopenia than in the control groups. Bcat1 knockdown not only suppressed the growth of muscle cells but also increased the production of ROS. Impaired cell growth and increased ROS production was rescued by co-introduction of an shRNA-resistant Bcat1 cDNA or addition of the mTORC1 stimulator MYH1485. Muscle cells with Bcat1 knockdown featured lower mTORC1 and S6K1 phosphorylation (pS6K1) than NT muscle cells. Addition of either shRNA-resistant Bcat1 cDNA or MYH1485 rescued the suppression of cell growth, increase in ROS production, and decrease in pS6K1.ConclusionsThe branched chain amino acids catabolic enzyme BCAT1 is essential for the growth of muscle cells. BCAT1 expression contributes to sustained growth of muscle cells by activating mTOR signaling and reducing ROS production.

Hemocompatibility of all-trans retinoic acid-loaded citrate polymer coatings for vascular stents.

Current strategies implementing drug-eluting polymer stent coatings fail to fully address the lasting effects of endothelial suppression which ultimately result in delayed reendothelialization and thrombogenic complications. The present study investigates the in vitro hemocompatibility of all-trans retinoic acid loaded poly (1,8-octanediol-co-citrate) coatings (AtRA-POC coatings) for advanced intravascular stent technology. The ability of these materials in supporting endothelial restoration via migration and proliferation while inhibiting smooth muscle cell growth is also explored. Using in vitro models, the hemocompatibility of AtRA-loaded POC-coated cobalt chromium (CoCr) vascular stents was evaluated in terms of platelet and inflammatory activity. Platelet activity was quantified by platelet adhesion and platelet activation, further supported by SEM visualization. Inflammatory activity was quantified by the production of proinflammatory cytokines by THP1 monocytes. Lastly, in vitro wound healing and an 5-Ethynyl-2'deoxyuridine (EdU) and pico green DNA assays were used in quantitating endothelial and smooth muscle cell migration and proliferation. Experimental examinations of platelet adhesion and activation demonstrate significant reductions in the platelet response to POC coated AtRA loaded stents when compared to bare CoCr stents. Such findings reveal AtRA-POC coatings to have significantly improved hemocompatibility compared to that of bare metal stents and at least as good as POC alone. Similarly, in reference to LPS-stimulated controls, Human monocyte-like THP1 cells in culture with AtRA-POC-CoCr stents for 24 hours showed reduced detection of proinflammatory cytokines, comparable to that of bare CoCr and untreated controls. This result supports AtRA-POC coatings as possessing limited immunological potential. Observations from in vitro endothelial and smooth muscle cell investigations demonstrate the ability of the drug AtRA to allow cell processes involved in restoration of the endothelium while inhibiting smooth muscle cell processes. This study demonstrates AtRA loaded POC coatings are hemocompatible, noninflammatory, and provide a promising strategy in enhancing vascular stent techniques and clinical integration. Possessing hemocompatibility and immunological compatibility that is at least as good as bare metal stents as clinical standards support the use of AtRA-POC coatings for vascular applications. Additionally, selectively reducing smooth muscle cell proliferation while supporting endothelial cell proliferation and migration further demonstrates the potential of these materials in significantly improving the state of vascular stent technology in the area of stent thrombosis and neointimal hyperplasia.

A trial design to maximize knowledge of the effects of rodatristat ethyl in the treatment of pulmonary arterial hypertension (ELEVATE 2).

Serotonin plays a key role in the development and maintenance of the pathobiology associated with pulmonary arterial hypertension (PAH). Platelet‐driven and locally produced serotonin from lung tissue and arterial endothelial cells induce excessive growth of pulmonary artery smooth muscle cells. The unchecked growth of these cells is a major driver of PAH including the remodeling of pulmonary arteries that dramatically reduces the diameter and flexibility of the arterial lumen. Tryptophan hydroxylase 1 (TPH1) is the rate‐limiting enzyme for biosynthesis of serotonin and is upregulated in PAH arterial endothelial cells, supporting TPH1 inhibition to treat PAH. Targeting the serotonin pathway via inhibition of peripheral serotonin and local production in diseased tissues, rather than individual receptor‐mediated or receptor‐independent mechanisms, may result in the ability to halt or reverse pulmonary vascular remodeling. Rodatristat ethyl, a prodrug for rodatristat, a potent, peripheral inhibitor of TPH1, has demonstrated efficacy in monocrotaline and SUGEN hypoxia nonclinical models of PAH and robust dose‐dependent reductions of 5‐hydroxyindoleacetic acid, the major metabolite of serotonin in plasma and urine of healthy human subjects. ELEVATE 2 (NCT04712669) is a Phase 2b, double‐blind, multicenter trial where patients with PAH are randomized to placebo, 300 or 600 mg twice daily of rodatristat ethyl. The trial incorporates endpoints to generate essential clinical efficacy, safety, pharmacokinetic, and pharmacodynamic data needed to evaluate the ability of rodatristat ethyl to ameliorate PAH by halting or reversing pulmonary vascular remodeling through its unique mechanism of TPH1 inhibition. Herein we describe the experimental design highlighting the trial's unique features.

MEOX2-mediated regulation of Cathepsin S promotes cell proliferation and motility in glioma

Nuclear transcription factor Mesenchyme Homeobox 2 (MEOX2) is a homeobox gene that is originally discovered to suppress the growth of vascular smooth muscle and endothelial cells. However, whether or not it is connected to cancer is yet unknown. Here, we report that MEOX2 functions as a tumor-initiating element in glioma. Bioinformatic analyses of public databases and investigation of MEOX2 expression in patients with glioma demonstrated that MEOX2 was abundant at both mRNA and protein levels in glioma. MEOX2 expression was shown to be inversely linked with the prognosis of glioma patients. MEOX2 inhibition changed the morphology of glioma cells, inhibited cell proliferation and motility, whereas had no effect on cell apoptosis. Besides, silencing MEOX2 also hampered the epithelial-mesenchymal transition (EMT), focal adhesion formation, and F-actin assembly. Overexpression of MEOX2 exhibited opposite effects. Importantly, RNA-sequencing, ChIP-qPCR assay, and luciferase reporter assay revealed Cathepsin S (CTSS) as a novel transcriptional target of MEOX2 in glioma cells. Consistently, MEOX2 causes glioma tumor development in mice and greatly lowers the survival period of tumor-bearing mice. Our findings indicate that MEOX2 promotes tumorigenesis and progression of glioma partially through the regulation of CTSS. Targeting MEOX2-CTSS axis might be a promising alternative for the treatment of glioma.

Calcium Channels in the Heart: Disease States and Drugs.

Calcium ions are the major signaling ions in the cells. They regulate muscle contraction, neurotransmitter secretion, cell growth and migration, and the activity of several proteins including enzymes and ion channels and transporters. They participate in various signal transduction pathways, thereby regulating major physiological functions. Calcium ion entry into the cells is regulated by specific calcium channels and transporters. There are mainly six types of calcium channels, of which only two are prominent in the heart. In cardiac tissues, the two types of calcium channels are the L type and the T type. L-type channels are found in all cardiac cells and T-type are expressed in Purkinje cells, pacemaker and atrial cells. Both these types of channels contribute to atrioventricular conduction as well as pacemaker activity. Given the crucial role of calcium channels in the cardiac conduction system, mutations and dysfunctions of these channels are known to cause several diseases and disorders. Drugs targeting calcium channels hence are used in a wide variety of cardiac disorders including but not limited to hypertension, angina, and arrhythmias. This review summarizes the type of cardiac calcium channels, their function, and disorders caused by their mutations and dysfunctions. Finally, this review also focuses on the types of calcium channel blockers and their use in a variety of cardiac disorders.

MiR-22-3p Inhibits Proliferation and Promotes Differentiation of Skeletal Muscle Cells by Targeting IGFBP3 in Hu Sheep.

Simple SummaryThe meat production of Hu sheep affects the mutton supply, and how to increase the meat production is the primary focus of genetic breeding. Therefore, in order to explore the key factors affecting the proliferation and differentiation of Hu sheep skeletal muscle cells, we performed functional verification of miRNA at the cellular level. Our findings are helpful to clarify the molecular regulation mechanism of proliferation and differentiation of skeletal muscle cells, which will benefit the molecular breeding of Hu sheep.The growth and development of skeletal muscle require a series of regulatory factors. MiRNA is a non-coding RNA with a length of about 22 nt, which can inhibit the expression of mRNA and plays an important role in the growth and development of muscle cells. The role of miR-22-3p in C2C12 cells and porcine skeletal muscle has been reported, but it has not been verified in Hu sheep skeletal muscle. Through qPCR, CCK-8, EdU and cell cycle studies, we found that overexpression of miR-22-3p inhibited proliferation of skeletal muscle cells (p < 0.01). The results of qPCR and immunofluorescence showed that overexpression of miR-22-3p promoted differentiation of skeletal muscle cells (p < 0.01), while the results of inhibiting the expression of miR-22-3p were the opposite. These results suggested that miR-22-3p functions in growth and development of sheep skeletal muscle cells. Bioinformatic analysis with mirDIP, miRTargets, and RNAhybrid software suggested IGFBP3 was the target of miR-22-3p, which was confirmed by dual-luciferase reporter system assay. IGFBP3 is highly expressed in sheep skeletal muscle cells. Overexpression of IGFBP3 was found to promote proliferation of skeletal muscle cells indicated by qPCR, CCK-8, EdU, and cell cycle studies (p < 0.01). The results of qPCR and immunofluorescence experiments proved that overexpression of IGFBP3 inhibited differentiation of skeletal muscle cells (p < 0.01), while the results of interfering IGFBP3 with siRNA were the opposite. These results indicate that miR-22-3p is involved in proliferation and differentiation of skeletal muscle cells by targeting IGFBP3.

Biomimetic Design and Biocompatibility of Biomimetic Calcium Carbonate Nanocomposites for Skeletal Muscle Injury Repair

As an important part of the human body, skeletal muscle is easily damaged in the process of exercise. The purpose of this study is to investigate the controllable construction and biocompatibility of biomimetic calcium carbonate nanocomposites in exercise skeletal muscle repair. In this study, adult healthy mice were selected as the research objects. First, the mice were anesthetized with 2.5% pentobarbital sodium. Then, the mice were fixed and hit with steel balls on the middle part of the gastrocnemius muscle abdomen to establish a skeletal muscle contusion model. The mice were killed by cervical dislocation under anesthesia. The injured gastrocnemius muscles were quickly removed to prepare the cell suspension. 0.8600 g chitosan was weighed and dissolved in 200 ml deionized water. Then, 0.0660 g of calcium chloride dihydrate was added and stirred to dissolve to prepare chitosan calcium carbonate composite. The calcium carbonate composite material with a length of 8.0 cm is cut into a square sheet of stainless steel. The stainless steel plate was sterilized and then placed in a 12 culture plate to dilute the full cells in the culture bottle. The cells were seeded in 12 well culture plates, and 100 μL cells and 600 μL medium were added to each well. The samples were cultured in the incubator (there were three control holes in each group for SEM test, three control holes for fluorescence measurement, and six control holes for MTT measurement). The plates were taken out and placed in a new 24 plate. 1000 μL medium was added. Then. 100 μL MTT solution was added into each well. The biocompatibility test was conducted for 4 h. The GSH PX level in the chitosan calcium carbonate composite group was lower than that in the natural recovery group (P &lt; 0.01). The results showed that chitosan calcium carbonate composite could promote the growth of skeletal muscle cells.

LDHA mediated degradation of extracellular matrix is a potential target for the treatment of aortic dissection

Aortic dissection (AD) is a disease with high mortality and lacks effective drug treatment. Recent studies have shown that the development of AD is closely related to glucose metabolism. Lactate dehydrogenase A (LDHA) is a key glycolytic enzyme and plays an important role in cardiovascular disease. However, the role of LDHA in the progression of AD remains to be elucidated. Here, we found that the level of LDHA was significantly elevated in AD patients and the mouse model established by BAPN combined with Ang II. In vitro, the knockdown of LDHA reduced the growth of human aortic vascular smooth muscle cells (HAVSMCs), glucose consumption, and lactate production induced by PDGF-BB. The overexpression of LDHA in HAVSMCs promoted the transformation of HAVSMCs from contractile phenotype to synthetic phenotype, and increased the expression of MMP2/9. Mechanistically, LDHA promoted MMP2/9 expression through the LDHA-NDRG3-ERK1/2-MMP2/9 pathway. In vivo, Oxamate, LDH and lactate inhibitor, reduced the degradation of elastic fibers and collagen deposition, inhibited the phenotypic transformation of HAVSMCs from contractile phenotype to synthetic phenotype, reduced the expression of NDRG3, p-ERK1/2, and MMP2/9, and delayed the progression of AD. To sum up, the increase of LDHA promotes the production of MMP2/9, stimulates the degradation of extracellular matrix (ECM), and promoted the transformation of HAVSMCs from contractile phenotype to synthetic phenotype. Oxamate reduced the progression of AD in mice. LDHA may be a therapeutic target for AD.

EphrinB2 promotes the human aortic smooth muscle cell growth and migration via mediating F-actin remodeling.

To evaluate the potential effect of EphrinB2 in human thoracic aortic dissection (TAD) and to illustrate the mechanisms governing the role of EphrinB2 in the growth of human aortic smooth muscle cells (HASMC). In the study, EphrinB2 expression was investigated by qRT-PCR and immunohistochemistry in 12 pairs of TAD and adjacent human tissues. HASMCs were used for in vitro experiments. Next, EphrinB2 overexpression and depletion in HASMCs were established by EphrinB2-overexpressing vectors and small interfering RNA, respectively. The transfection efficiency was evaluated by qRT-PCR and Western blot. The effects of overexpression and depletion of EphrinB2 on cell proliferation, migration, and invasion were tested in vitro. Cell Counting Kit-8, flow cytometry and transwell migration/invasion, and wound healing assay were used to explore the function of EphrinB2 on HASMC cell lines. The relationship between EphrinB2 and F-actin was assessed by Western blot, immunofluorescence, and Co-IP. We found that EphrinB2 was a prognostic biomarker of TAD patients. Moreover, EphrinB2 expression negatively correlated to aortic dissection tissues, and disease incidence of males, suggesting that EphrinB2 might act as a TAD suppressor by promoting proliferation or decreasing apoptosis in HASMC. Next, over-expression of EphrinB2 in HASMC lines drove cell proliferation, migration, and invasion, and inhibited apoptosis while knockdown EphrinB2 showed the opposite phenomenon, respectively. Furthermore, the level of F-actin in mRNA, protein, and distribution in HASMC cell lines highly matched with the expression of EphrinB2, which indicated that EphrinB2 could mediate the HASMC cytoskeleton via inducing F-actin. In conclusion, our results first provided the pivotal role of EphrinB2 in HASMC proliferation initiated by mediating F-actin and demonstrated a prognostic biomarker and the potential targets for therapy to prevent thoracic aortic dissection.

The effect of surface morphology on endothelial and smooth muscle cells growth on blow-spun fibrous scaffolds

This study aimed to analyze the growth of two types of blood vessel building cells: endothelial cells (ECs) and smooth muscle cells (SMCs) on surfaces with different morphology. Two types of materials, differing in morphology, were produced by the solution blow spinning technique. One-layer materials consisted of one fibrous layer with two fibrous surfaces. Bi-layer materials consisted of one fibrous-solid layer and one fibrous layer, resulting in two different surfaces. Additionally, materials with different average fiber diameters (about 200, 500, and 900 nm) were produced for each group. It has been shown that it is possible to obtain structures with a given morphology by changing the selected process parameters (working distance and polymer solution concentration). Both morphology (solid versus fibrous) and average fiber diameter (submicron fibers versus microfibers) of scaffolds influenced the growth of ECs. However, this effect was only visible after an extended period of culture (6 days). In the case of SMCs, it was proved that the best growth of SMCs is obtained for micron fibers (with an average diameter close to 900 nm) compared to the submicron fibers (with an average diameter below 900 nm).

Serum from Patients with Severe Alcoholic Liver Cirrhosis Inhibits Proliferation and Migration of Human Coronary Artery Smooth Muscle Cells.

Liver cirrhosis has been associated with an increased risk of coronary artery disease and clinical complications following percutaneous coronary revascularization. The present study is based on the hypothesis that cirrhosis may influence intimal hyperplasia following PCI. Sera from 10 patients with alcoholic liver cirrhosis and 10 age-matched healthy controls were used to stimulate cultured human coronary artery smooth muscle cells (HCASMC) for 48 h. HCASMC proliferation, migration, gene expression and apoptosis were investigated. Serum concentrations of growth factors and markers of liver function were also determined in patients and healthy controls. Treatment of HCASMC with patient sera reduced cell proliferation and migration (p < 0.05 vs. healthy controls), whereas apoptosis was unaffected (p = 0.160). Expression of genes associated with a synthetic vascular smooth muscle cell phenotype was decreased in cells stimulated with serum from cirrhotic patients (RBP1, p = 0.001; SPP1, p = 0.003; KLF4, p = 0.004). Platelet-derived growth factor-BB serum concentrations were lower in patients (p = 0.001 vs. controls). The results suggest the presence of circulating factors in patients with alcoholic liver cirrhosis affecting coronary smooth muscle cell growth. These findings may have implications for clinical outcomes following percutaneous coronary revascularization in these patients.

Abstract 12771: HDAC Inhibition With Quisinostat Reduces Pulmonary Vascular Remodeling in Experimentally Induced Pulmonary Arterial Hypertension

Introduction: Histone deacetylases (HDACs) expression and activity are increased in pulmonary arterial hypertension (PAH). Although inhibiting HDACs has been promoted as a treatment for PAH, results from preclinical studies were equivocal and raised concerns regarding safety. Quisinostat is a “second generation” HDAC inhibitor, which is highly potent against HDAC1 and elicits a prolonged pharmacodynamic response. Hypothesis: Quisinostat may exert therapeutic effects in experimentally-induced PAH. Methods: HDAC1 expression was measured in lungs and microvascular endothelial cells (MVECs) from control donors and PAH patients. HDAC activity, proliferation and inflammation were measured after quisinostat treatment of PAH MVECs. Chronic quisinostat treatment was tested in three PAH rat models: the Sugen + hypoxia (SuHx), the Monocrotaline shunt flow (MF) and pulmonary artery banding (PAB) model. Results: HDAC1 expression is increased in lungs of PAH patients. Quisinostat decreased proliferation and mitigated inflammatory cytokine expression in MVECs. Conditioned medium from quisinostat treated PAH MVECs inhibited the growth of healthy pulmonary artery smooth muscle cells. In vivo , quisinostat treatment reduced right ventricular (RV) systolic pressure, RV afterload and total pulmonary resistance in SuHx rats. Further analysis by histology revealed that quisinostat reduced pulmonary vascular remodeling in both SuHx and MF rat models by reducing proliferation and perivascular inflammation. The PAB rat model demonstrated that quisinostat had no independent effect on RV function or remodeling. Conclusions: Quisinostat can partly reverse RV afterload and pulmonary vascular remodeling in established experimental PAH and may be a promising intervention for PAH.

Knockdown of long noncoding RNA GAS5 reduces vascular smooth muscle cell apoptosis by inactivating EZH2-mediated RIG-I signaling pathway in abdominal aortic aneurysm

BackgroundAbdominal aortic aneurysm (AAA), an irreversible cardiovascular disease prevalent in the artery, causes the increase of the aneurysm diameter over time, and is a fatal phenomenon inducing sidewall rupture. Long noncoding RNAs (lncRNAs) serve as promising biomarkers for AAA. In the present study, we sought to define the role of lncRNA growth-arrest-specific transcript 5 (GAS5) in growth of smooth muscle cells (SMC) and progression of AAA.MethodsInitially, we established angiotensin II (Ang II)-induced AAA mouse models and Ang II-treated vascular SMC model. RT-qPCR and Western blot analysis were adopted to determine expression of GAS5 and zeste homolog 2 (EZH2). After ectopic expression and depletion experiments in Ang II-treated mice and vascular SMCs, cell apoptosis was detected in SMCs using flow cytometry and in mice using TUNEL staining. The binding of GAS5 and EZH2 was evaluated using RNA binding protein immunoprecipitation (RIP) and Co-IP assays.ResultsIncreased GAS5 and RIG-I but decreased EZH2 were found in aortic tissues of AAA mice. EZH2 overexpression inhibited AAA formation and suppressed SMC apoptosis. Functionally, EZH2 blocked the RIG-I signaling pathway and consequently inhibited SMC apoptosis. GAS5 regulated EZH2 transcription in a negative manner in SMCs. Knockdown of GAS5 attenuated SMC apoptosis, which was reversed by EZH2 inhibition or RIG-I overexpression.ConclusionsThe current study demonstrated that GAS5 induced SMC apoptosis and subsequent AAA onset by activating EZH2-mediated RIG-I signaling pathway, highlighting GAS5 as a novel biomarker for AAA.

CircRNA Chordc1 protects mice from abdominal aortic aneurysm by contributing to the phenotype and growth of vascular smooth muscle cells

Circular RNAs (circRNAs) have important potential in modulating vascular smooth muscle cell (VSMC) activity, but their roles in abdominal aortic aneurysm (AAA) are unknown. We performed in situ hybridization and immunohistochemistry and determined that circChordc1 (cysteine and histidine-rich domain containing 1) was markedly downregulated in aneurysm tissue compared with normal arteries. A gene gain and loss strategy was used to confirm that circChordc1 transformed VSMCs into a contracted phenotype and improved their growth, which significantly suppressed aneurysm formation and reduced the risk of rupture in mouse models of angiotensin (Ang) II- and CaCl2-induced AAA. RNA pull-down, immunoprecipitation, and immunoblotting indicated that circChordc1 facilitated the VSMC phenotype and growth determination by binding to vimentin and ANXA2 (annexin A2), which not only increased vimentin phosphorylation to promote its degradation but also promoted the interaction between ANXA2 and glycogen synthase kinase 3 beta (GSK3β) to induce the nuclear entry of β-catenin. Thus, our present study revealed that circChordc1 optimized the VSMC phenotype and improved their growth by inducing vimentin degradation and increasing the activity of the GSK3β/β-catenin pathway, thereby extenuating vascular wall remodeling and reversing pathological aneurysm progression.