P7C3 ameliorates barium chloride-induced skeletal muscle injury activating transcriptomic and epigenetic modulation of myogenic regulatory factors.

Skeletal muscle injury affects the quality of life in many pathologies, including volumetric muscle loss, contusion injury, and aging. We hypothesized that the nicotinamide phosphoribosyltransferase (Nampt) activator P7C3 improves muscle repair following injury. In the present study, we tested the effect of P7C3 (1-anilino-3-(3,6-dibromocarbazol-9-yl) propan-2-ol) on chemically induced muscle injury. Muscle injury was induced by injecting 50 µL 1.2% barium chloride (BaCl2) into the tibialis anterior (TA) muscle in C57Bl/6J wild-type male mice. Mice were then treated with either 10 mg/kg body weight of P7C3 or Vehicle intraperitoneally for 7 days and assessed for histological, biochemical, and molecular changes. In the present study, we show that the acute BaCl2-induced TA muscle injury was robust and the P7C3-treated mice displayed a significant increase in the total number of myonuclei and blood vessels, and decreased serum CK activity compared with vehicle-treated mice. The specificity of P7C3 was evaluated using Nampt+/- mice, which did not display any significant difference in muscle repair capacity among treated groups. RNA-sequencing analysis of the injured TA muscles displayed 368 and 212 genes to be exclusively expressed in P7C3 and Veh-treated mice, respectively. There was an increase in the expression of genes involved in cellular processes, inflammatory response, angiogenesis, and muscle development in P7C3 versus Veh-treated mice. Conversely, there is a decrease in muscle structure and function, myeloid cell differentiation, glutathione, and oxidation-reduction, drug metabolism, and circadian rhythm signaling pathways. Chromatin immunoprecipitation-quantitative polymerase chain reaction (qPCR) and reverse transcription-qPCR analyses identified increased Pax7, Myf5, MyoD, and Myogenin expression in P7C3-treated mice. Increased histone lysine (H3K) methylation and acetylation were observed in P7C3-treated mice, with significant upregulation in inflammatory markers. Moreover, P7C3 treatment significantly increased the myotube fusion index in the BaCl2-injured human skeletal muscle in vitro. P7C3 also inhibited the lipopolysaccharide-induced inflammatory response and mitochondrial membrane potential of RAW 264.7 macrophage cells. Overall, we demonstrate that P7C3 activates muscle stem cells and enhances muscle injury repair with increased angiogenesis.

Combined Endurance and Resistance Exercise Mitigates Age-Associated Cardiac Dysfunction.

Aging is associated with a decline in cardiac function. Exercise has been shown to effectively reduce the risks of cardiovascular diseases. Here whether a combination of endurance and resistance exercises can improve cardiac function in aged mice during late life is investigated. Through transcriptome analysis, several signaling pathways activated in the hearts of 22-month-old mice after combined exercise, including cardiac muscle contraction, mitophagy, and longevity regulation are identified. Combined exercise training mitigated age-associated pathological cardiac hypertrophy, reduced oxidative stress, cardiac senescence, and enhanced cardiac function. Upstream stimulatory factor 2 (Usf2) is upregulated in the aged mouse hearts with combined exercise compared to sedentary mice. In the human cardiomyocytes senescent model, overexpression of Usf2 led to anti-senescence effects, while knockdown of Usf2 exacerbated cellular senescence. The results suggest that a combination of endurance and resistance exercises, such as swimming and resistance running, can mitigate age-related pathological cardiac remodeling and cardiac dysfunction in late life. These cardioprotective effects are likely due to the activation of Usf2 and its anti-senescence effect. Therefore, Usf2 can potentially be a novel therapeutic target for mitigating age-related cardiac dysfunction.

Association of Common Foods with Inflammation and Mortality: Analysis from a Large Prospective Cohort Study.

Some dietary patterns are associated with inflammation, while others lower inflammation and improve health. However, many people cannot follow a complete, healthy diet. Therefore, this study's aim was to identify specific foods associated chronic inflammation and mortality. The study used Multi-Ethnic Study of Atherosclerosis (MESA) research materials from the NHLBI Biologic Specimen and Data Repository Information Coordinating Center. Three plant-based and three animal-based MESA food categories were chosen based on perceived availability in the western diet. The assessed food categories were avocado, ham, sausage, eggs, greens, and broccoli. Inflammatory markers assessed were interleukin-6 (IL-6), fibrinogen antigen, C-reactive protein, D-Dimer, interleukin-2, matrix metalloproteinase 3, necrosis factor-a soluble receptors, oxidized LDL (oxLDL), and total homocysteine. The primary outcome was the multivariable association of foods and inflammatory markers with all-cause mortality. All inflammatory makers, except oxLDL, were associated with mortality in univariate analysis. The effect was largest with IL-6 and D-dimer. The category of broccoli had the most consistent association in univariate analyses with lower inflammation and lower mortality odds. Low and high broccoli consumption versus no consumption were associated with lower mortality odds in the multivariable models with IL-6 and D-dimer. Consumption of the MESA-defined food category "broccoli" (i.e., broccoli, cabbage, cauliflower, brussels sprouts, sauerkraut, and kimchee) was associated with lower inflammation and lower mortality odds. These findings should be validated in randomized controlled trials testing a "food is medicine" approach to identify which, if any, of these foods may have potential as an herbal therapeutic for chronic inflammation.

Development and testing of nanoparticles delivery for P7C3 small molecule using injury models.

The use of nanoparticles (NPs) has emerged as a potential tool for safe and effective drug delivery. In the present study, we developed small molecule P7C3-based NPs and tested its efficacy and toxicity along with the tissue specific aptamer-modified P7C3 NPs. The P7C3 NPs were prepared using poly (D, L-lactic-co-glycolic acid) carboxylic acid (PLGA-COOH) polymer, were conjugated with skeletal muscle-specific RNA aptamer (A01B P7C3 NPs) and characterized for its cytotoxicity, cellular uptake, and wound healing in vitro. The A01B P7C3 NPs demonstrated an encapsulation efficiency of 30.2 ± 2.6%, with the particle size 255.9 ± 4.3 nm, polydispersity index of 0.335 ± 0.05 and zeta potential of + 10.4 ± 1.8mV. The FTIR spectrum of P7C3 NPs displayed complete encapsulation of the drug in the NPs. The P7C3 NPs and A01B P7C3 NPs displayed sustained drug release in vitro for up to 6 days and qPCR analysis confirmed A01B aptamer binding to P7C3 NPs. The C2C12 cells viability assay displayed no cytotoxic effects of all 3 formulations at 48 and 72 h. In addition, the cellular uptake of A01B P7C3 NPs in C2C12 myoblasts demonstrated higher uptake. In vitro assay mimicking wound healing showed improved wound closure with P7C3 NPs. In addition, P7C3 NPs significantly decreased TNF-α induced NF-κB activity in the C2C12/NF-κB reporter cells after 24-hour treatment. The P7C3 NPs showed 3-4-fold higher efficacy compared to P7C3 solutions in both wound-closure and inflammation assays in C2C12 cells. Furthermore, the P7C3 NPs showed 3-4-fold higher efficacy in reducing the infarct size and protected mouse hearts from ex vivo ischemia-reperfusion injury. Overall, this study demonstrates the safe and effective delivery of P7C3 NPs.

Muscle-Brain crosstalk in cognitive impairment.

Sarcopenia is an age-related, involuntary loss of skeletal muscle mass and strength. Alzheimer's disease (AD) is the most common cause of dementia in elderly adults. To date, no effective cures for sarcopenia and AD are available. Physical and cognitive impairments are two major causes of disability in the elderly population, which severely decrease their quality of life and increase their economic burden. Clinically, sarcopenia is strongly associated with AD. However, the underlying factors for this association remain unknown. Mechanistic studies on muscle-brain crosstalk during cognitive impairment might shed light on new insights and novel therapeutic approaches for combating cognitive decline and AD. In this review, we summarize the latest studies emphasizing the association between sarcopenia and cognitive impairment. The underlying mechanisms involved in muscle-brain crosstalk and the potential implications of such crosstalk are discussed. Finally, future directions for drug development to improve age-related cognitive impairment and AD-related cognitive dysfunction are also explored.

Chemokine/ITGA4 Interaction Directs iPSC-Derived Myogenic Progenitor Migration to Injury Sites in Aging Muscle for Regeneration.

The failure of muscle to repair after injury during aging may be a major contributor to muscle mass loss. We recently generated muscle progenitor cells (MPCs) from human-induced pluripotent stem-cell (iPSC) cell lines using small molecules, CHIR99021 and Givinostat (Givi-MPCs) sequentially. Here, we test whether the chemokines overexpressed in injured endothelial cells direct MPC migration to the site by binding to their receptor, ITGA4. ITGA4 was heavily expressed in Givi-MPCs. To study the effects on the mobilization of Givi-MPCs, ITGA4 was knocked down by an ITGA4 shRNA lentiviral vector. With and without ITGA4 knocked down, cell migration in vitro and cell mobilization in vivo using aged NOD scid gamma (NSG) mice and mdx/scid mice were analyzed. The migration of shITGA4-Givi-MPCs was significantly impaired, as shown in a wound-healing assay. The knockdown of ITGA4 impaired the migration of Givi-MPCs towards human aortic endothelial cells (HAECs), in which CX3CL1 and VCAM-1 were up-regulated by the treatment of TNF-α compared with scramble ones using a transwell system. MPCs expressing ITGA4 sensed chemokines secreted by endothelial cells at the injury site as a chemoattracting signal to migrate to the injured muscle. The mobilization of Givi-MPCs was mediated by the ligand-receptor interaction, which facilitated their engraftment for repairing the sarcopenic muscle with injury.

Genetic deletion of Kvß2 (AKR6) causes loss of muscle function and increased inflammation in mice.

The voltage-gated potassium channels (Kv) are complex ion channels with distinct roles in neurotransmission, electrical conductivity of the heart, and smooth and striated muscle functions. Previously, we demonstrated that deletion of Kvß2 in mice results in decreased Pax7 protein levels, hindlimb muscles and body weights, and fiber type switching. In the present study, we tested the hypothesis that Kvß2 regulates skeletal muscle function in mice. The young and old Kvß2 knockout (KO) and wildtype (WT) mice were utilized to test the aging phenotype and skeletal muscle function. Consistent with our previous finding, we found a significant reduction in hindlimb skeletal muscles mass and body weight in young Kvß2 KO mice, which was also significantly reduced in old Kvß2 KO mice compared with age-matched WT mice. Forelimb grip strength, and the hindleg extensor digitorum longus (EDL) muscles force-frequency relations were significantly decreased in young and old Kvß2 KO mice compared to age-matched WT mice. Analysis of transmission electron microscopy images of EDL muscles in young mice revealed a significant reduction in the sarcomere length for Kvß2 KO vs. WT. Hematoxylin and eosin-stained tibialis anterior muscles cryosections displayed a significant decrease in the number of medium (2,000-4,000 µm2) and largest (>4,000 µm2) myofibers area in young Kvß2 KO vs. WT mice. We also found a significant increase in fibrotic tissue area in young Kvß2 KO mice compared with age-matched WT mice. Analysis of RNA Seq data of the gastrocnemius muscles (GAS) identified significant increase in genes involved in skeletal muscle development, proliferation and cell fate determination, atrophy, energy metabolism, muscle plasticity, inflammation, and a decrease in circadian core clock genes in young Kvß2 KO vs. WT mice. Several genes were significantly upregulated (384 genes) and downregulated (40 genes) in young Kvß2 KO mice compared to age-matched WT mice. Further, RT-qPCR analysis of the GAS muscles displayed a significant increase in pro-inflammatory marker Il6 expression in young Kvß2 KO mice compared to age-matched WT mice. Overall, the present study shows that deletion of Kvß2 leads to decreased muscles strength and increased inflammation.

Transformational Applications of Human Cardiac Organoids in Cardiovascular Diseases.

Organoid technology has significantly advanced in recent years and revolutionized the field for generation of organs using in vitro systems (a.k.a "organs in a dish"). The use of pluripotent stem cells or tissue derived cells for generating a 3-dimensional culture system to recapitulate the architecture and function of the organ is central in achieving and improving organoid systems. Unlike most organs in the body, very little progress has been made in cardiac organoid due to its structural complexity and vascularization. In this review, we will discuss the current applications of human cardiac organoids for cardiac disease modeling, drug discovery, drug cardiotoxicity testing, and clinical applications.

Cardioprotective Effects of 1-(3,6-Dibromo-carbazol-9-yl)-3-Phenylamino-Propan-2-Ol in Diabetic Hearts via Nicotinamide Phosphoribosyltransferase Activation.

Diabetes is associated with increased cardiac injury and sudden death. Nicotinamide phosphoribosyltransferase (Nampt) is an essential enzyme for the NAD+ salvage pathway and is dysregulated in diabetes. Nampt activation results in rescued NADH/NAD+ ratios and provides pharmacological changes necessary for diabetic cardioprotection. Computer docking shows that 1-(3,6-Dibromo-carbazol-9-yl)-3-phenylamino-propan-2-ol (P7C3) allows for enhanced Nampt dimerization and association. To test the pharmacological application, we used male leptin receptor-deficient (db/db) mice and treated them with Nampt activator P7C3. The effects of 4-week P7C3 treatment on cardiac function were evaluated along with molecular signaling changes for phosphorylated protein kinase B (p-AKT), phosphorylated endothelial nitric oxide synthase (p-eNOS), and sirtuin 1 (SIRT1). The cardiac function evaluated by ECG and echocardiography were significantly improved after 4 weeks of P7C3 treatment. Biochemically, higher NADH/NAD+ ratios in diabetic hearts were rescued by P7C3 treatment. Moreover, activities of Nampt and SIRT1 were significantly increased in P7C3-treated diabetic hearts. P7C3 treatment significantly decreased the blood glucose in diabetic mice with 4-week treatment as noted by glucose tolerance test and fasting blood glucose measurements compared with vehicle-treated mice. P7C3 activated Nampt enzymatic activity both in vitro and in the 4-week diabetic mouse hearts, demonstrating the specificity of the small molecule. P7C3 treatment significantly enhanced the expression of cardioprotective signaling of p-AKT, p-eNOS, and Beclin 1 in diabetic hearts. Nampt activator P7C3 allows for decreased infarct size with decreased Troponin I and lactose dehydrogenase (LDH) release, which is beneficial to the heart. Overall, the present study shows that P7C3 activates Nampt and SIRT1 activity and decreases NADH/NAD+ ratio, resulting in improved biochemical signaling providing cardioprotection. SIGNIFICANCE STATEMENT: This study shows that 1-(3,6-Dibromo-carbazol-9-yl)-3-phenylamino-propan-2-ol (P7C3) is effective in treating diabetes and cardiovascular diseases. The novel small molecule is antiarrhythmic and improves the ejection fraction in diabetic hearts. The study successfully demonstrated that P7C3 decreases the infarct size in hearts during myocardial infarction and ischemia-reperfusion injury. Biochemical and cellular signaling show increased NAD+ levels, along with Nampt activity involved in upregulating protective signaling in the diabetic heart. P7C3 has high therapeutic potential for rescuing heart disease.

Novel biomarkers of inflammation in heart failure with preserved ejection fraction: analysis from a large prospective cohort study.

BACKGROUND: Heart failure with preserved ejection fraction (HFpEF) is a syndrome with a heterogeneous cluster of causes, including non-resolving inflammation, endothelial dysfunction, and multi-organ defects. The present study's objective was to identify novel predictors of HFpEF. METHODS: The study analyzed the Multi-Ethnic Study of Atherosclerosis (MESA) to assess the association of specific markers of inflammation with new onset of HFpEF (interleukin-2 [IL-2], matrix metalloproteinase 3 [MMP3], large low-density lipoprotein cholesterol [LDL-C], and medium high-density lipoprotein cholesterol [HDL-C]). The study included men and women 45 to 84 years of age without cardiovascular disease at baseline. The primary outcome was the multivariate association of the hypothesized markers of inflammation with new-onset of HFpEF versus participants without new-onset heart failure. Participants with missing data were excluded. RESULTS: The present analysis included 6814 participants, 53% female, with a mean age of 62 years. Among the entire cohort, HFpEF was diagnosed in 151 (2.2%) participants and heart failure with reduced ejection fraction (HFrEF) was diagnosed in 146 (2.1%) participants. Participants were followed for the outcome of heart failure for a median 13.9 years. Baseline IL-2 was available for 2861 participants. The multivariate analysis included 2792 participants. Of these, 2668 did not develop heart failure, 62 developed HFpEF, 47 developed HFrEF, and 15 developed unclassified heart failure. In the multivariate regression model, IL-2 was associated with new-onset HFpEF (OR, 1.00058; 95% confidence interval, 1.00014 to 1.00102, p = 0.009) but not new-onset HFrEF. In multivariate analysis, MMP3, large LDL-C, and medium HDL-C were not associated with HFpEF or HFrEF. CONCLUSION: These findings portend IL-2 as an important component of suboptimal inflammation in the pathogenesis of HFpEF.

NAD+ centric mechanisms and molecular determinants of skeletal muscle disease and aging.

The nicotinamide adenine dinucleotide (NAD+) is an essential redox cofactor, involved in various physiological and molecular processes, including energy metabolism, epigenetics, aging, and metabolic diseases. NAD+ repletion ameliorates muscular dystrophy and improves the mitochondrial and muscle stem cell function and thereby increase lifespan in mice. Accordingly, NAD+ is considered as an anti-oxidant and anti-aging molecule. NAD+ plays a central role in energy metabolism and the energy produced is used for movements, thermoregulation, and defense against foreign bodies. The dietary precursors of NAD+ synthesis is targeted to improve NAD+ biosynthesis; however, studies have revealed conflicting results regarding skeletal muscle-specific effects. Recent advances in the activation of nicotinamide phosphoribosyltransferase in the NAD+ salvage pathway and supplementation of NAD+ precursors have led to beneficial effects in skeletal muscle pathophysiology and function during aging and associated metabolic diseases. NAD+ is also involved in the epigenetic regulation and post-translational modifications of proteins that are involved in various cellular processes to maintain tissue homeostasis. This review provides detailed insights into the roles of NAD+ along with molecular mechanisms during aging and disease conditions, such as the impacts of age-related NAD+ deficiencies on NAD+-dependent enzymes, including poly (ADP-ribose) polymerase (PARPs), CD38, and sirtuins within skeletal muscle, and the most recent studies on the potential of nutritional supplementation and distinct modes of exercise to replenish the NAD+ pool.

Nampt activator P7C3 ameliorates diabetes and improves skeletal muscle function modulating cell metabolism and lipid mediators.

BACKGROUND: Nicotinamide phosphoribosyltransferase (Nampt), a key enzyme in NAD salvage pathway is decreased in metabolic diseases, and its precise role in skeletal muscle function is not known. We tested the hypothesis, Nampt activation by P7C3 (3,6-dibromo-α-[(phenylamino)methyl]-9H-carbazol-9-ethanol) ameliorates diabetes and muscle function. METHODS: We assessed the functional, morphometric, biochemical, and molecular effects of P7C3 treatment in skeletal muscle of type 2 diabetic (db/db) mice. Nampt+/- mice were utilized to test the specificity of P7C3. RESULTS: Insulin resistance increased 1.6-fold in diabetic mice compared with wild-type mice and after 4 weeks treatment with P7C3 rescued diabetes (P < 0.05). In the db-P7C3 mice fasting blood glucose levels decreased to 0.96-fold compared with C57Bl/6J wild-type naïve control mice. The insulin and glucose tolerance tests blood glucose levels were decreased to 0.6-fold and 0.54-folds, respectively, at 120 min along with an increase in insulin secretion (1.76-fold) and pancreatic ß-cells (3.92-fold) in db-P7C3 mice. The fore-limb and hind-limb grip strengths were increased to 1.13-fold and 1.17-fold, respectively, together with a 14.2-fold increase in voluntary running wheel distance in db-P7C3 mice. P7C3 treatment resulted in a 1.4-fold and 7.1-fold increase in medium-sized and larger-sized myofibres cross-sectional area, with a concomitant 0.5-fold decrease in smaller-sized myofibres of tibialis anterior (TA) muscle. The transmission electron microscopy images also displayed a 1.67-fold increase in myofibre diameter of extensor digitorum longus muscle along with 2.9-fold decrease in mitochondrial area in db-P7C3 mice compared with db-Veh mice. The number of SDH positive myofibres were increased to 1.74-fold in db-P7C3 TA muscles. The gastrocnemius and TA muscles displayed a decrease in slow oxidative myosin heavy chain type1 (MyHC1) myofibres expression (0.46-fold) and immunostaining (6.4-fold), respectively. qPCR analysis displayed a 2.9-fold and 1.3-fold increase in Pdk4 and Cpt1, and 0.55-fold and 0.59-fold decrease in Fgf21 and 16S in db-P7C3 mice. There was also a 3.3-fold and 1.9-fold increase in Fabp1 and CD36 in db-Veh mice. RNA-seq differential gene expression volcano plot displayed 1415 genes to be up-regulated and 1726 genes down-regulated (P < 0.05) in db-P7C3 mice. There was 1.02-fold increase in serum HDL, and 0.9-fold decrease in low-density lipoprotein/very low-density lipoprotein ratio in db-P7C3 mice. Lipid profiling of gastrocnemius muscle displayed a decrease in inflammatory lipid mediators n-6; AA (0.83-fold), and n-3; DHA (0.69-fold) and EPA (0.81-fold), and a 0.66-fold decrease in endocannabinoid 2-AG and 2.0-fold increase in AEA in db-P7C3 mice. CONCLUSIONS: Overall, we demonstrate that P7C3 activates Nampt, improves type 2 diabetes and skeletal muscle function in db/db mice.

Post hoc assessment of the relationship among coronary stenosis, electrocardiography, and ventricular function in patients with heart disease.

Cardiovascular diseases including cardiac arrhythmias lead to fatal events in patients with coronary artery disease; however, clinical associations from echocardiography, electrocardiography (ECG), and biomarkers remain unknown. We sought to identify the factors that may be related to elevated QRS intervals in patients with risk for coronary artery disease. In this study, we performed analysis of clinical data from 503 patients divided into two groups, i.e., patients with either <50% coronary artery stenosis or >50% coronary artery stenosis. We further examined patients with elevated ECG parameters such as QRS > 100 ms and QTc > 440 ms. Patients with >50% coronary artery stenosis exhibited significant increases in age, triglycerides, and troponin levels. Further, ECG parameters demonstrated increased QRS and QTc durations, while echocardiographic parameters highlighted a decrease in ejection fraction (EF) and fractional shortening (FS). Patients with QTc > 440 ms exhibited increased brain natriuretic peptide and creatinine levels with a decrease in estimated glomerular filtration rate clearance rates. Patients with QRS > 100 ms had greater left ventricular (LV) mass and LV internal diameter in systole and diastole. Multimodal logistic regression showed significant relation between QTc, age, and creatinine. These findings suggest that patients with significant coronary stenosis may have lower EF and FS with prolonged QRS intervals, demonstrating greater risk for arrhythmic events.

Deletion of Kvß2 (AKR6) Attenuates Isoproterenol Induced Cardiac Injury with Links to Solute Carrier Transporter SLC41a3 and Circadian Clock Genes.

Kvß subunits belong to the aldo-keto reductase superfamily, which plays a significant role in ion channel regulation and modulates the physiological responses. However, the role of Kvß2 in cardiac pathophysiology was not studied, and therefore, in the present study, we hypothesized that Kvß2 plays a significant role in cardiovascular pathophysiology by modulating the cardiac excitability and gene responses. We utilized an isoproterenol-infused mouse model to investigate the role of Kvß2 and the cardiac function, biochemical changes, and molecular responses. The deletion of Kvß2 attenuated the QTc (corrected QT interval) prolongation at the electrocardiographic (ECG) level after a 14-day isoproterenol infusion, whereas the QTc was significantly prolonged in the littermate wildtype group. Monophasic action potentials verified the ECG changes, suggesting that cardiac changes and responses due to isoproterenol infusion are mediated similarly at both the in vivo and ex vivo levels. Moreover, the echocardiographic function showed no further decrease in the ejection fraction in the isoproterenol-stimulated Kvß2 knockout (KO) group, whereas the wildtype mice showed significantly decreased function. These experiments revealed that Kvß2 plays a significant role in cardiovascular pathophysiology. Furthermore, the present study revealed SLC41a3, a major solute carrier transporter affected with a significantly decreased expression in KO vs. wildtype hearts. The electrical function showed that the decreased expression of SLC41a3 in Kvß2 KO hearts led to decreased Mg2+ responses, whereas, in the wildtype hearts, Mg2+ caused action potential duration (APD) shortening. Based on the in vivo, ex vivo, and molecular evaluations, we identified that the deletion of Kvß2 altered the cardiac pathophysiology mediated by SLC41a3 and altered the NAD (nicotinamide adenine dinucleotide)-dependent gene responses.

Metabolic regulation of Kv channels and cardiac repolarization by Kvß2 subunits.

Voltage-gated potassium (Kv) channels control myocardial repolarization. Pore-forming Kvα proteins associate with intracellular Kvß subunits, which bind pyridine nucleotides with high affinity and differentially regulate channel trafficking, plasmalemmal localization and gating properties. Nevertheless, it is unclear how Kvß subunits regulate myocardial K+ currents and repolarization. Here, we tested the hypothesis that Kvß2 subunits regulate the expression of myocardial Kv channels and confer redox sensitivity to Kv current and cardiac repolarization. Co-immunoprecipitation and in situ proximity ligation showed that in cardiac myocytes, Kvß2 interacts with Kv1.4, Kv1.5, Kv4.2, and Kv4.3. Cardiac myocytes from mice lacking Kcnab2 (Kvß2-/-) had smaller cross sectional areas, reduced sarcolemmal abundance of Kvα binding partners, reduced Ito, IK,slow1, and IK,slow2 densities, and prolonged action potential duration compared with myocytes from wild type mice. These differences in Kvß2-/- mice were associated with greater P wave duration and QT interval in electrocardiograms, and lower ejection fraction, fractional shortening, and left ventricular mass in echocardiographic and morphological assessments. Direct intracellular dialysis with a high NAD(P)H:NAD(P)+ accelerated Kv inactivation in wild type, but not Kvß2-/- myocytes. Furthermore, elevated extracellular levels of lactate increased [NADH]i and prolonged action potential duration in wild type cardiac myocytes and perfused wild type, but not Kvß2-/-, hearts. Taken together, these results suggest that Kvß2 regulates myocardial electrical activity by supporting the functional expression of proteins that generate Ito and IK,slow, and imparting redox and metabolic sensitivity to Kv channels, thereby coupling cardiac repolarization to myocyte metabolism.

Nanoparticle drug delivery characterization for fluticasone propionate and in vitro testing 1.

Glucocorticoids, such as fluticasone propionate (FP), are used for the treatment of inflammation and alleviation of nasal symptoms and allergies, and as an antipruritic. However, both short- and long-term therapeutic use of glucocorticoids can lead to muscle weakness and atrophy. In the present study, we evaluated the feasibility of the nanodelivery of FP with poly(dl-lactide-co-glycolide) (PLGA) and tested in vitro function. FP-loaded PLGA nanoparticles were prepared via nanoprecipitation and morphological characteristics were studied via scanning electron microscopy. FP-loaded nanoparticles demonstrated an encapsulation efficiency of 68.6% ± 0.5% with a drug loading capacity of 4.6% ± 0.04%, were 128.8 ± 0.6 nm in diameter with a polydispersity index of 0.07 ± 0.008, and displayed a zeta potential of -19.4 ± 0.7. A sustained in vitro drug release pattern was observed for up to 7 days. The use of fluticasone nanoparticle decreased lipopolysaccharide (LPS)-induced lactate dehydrogenase release compared with LPS alone in C2C12 treated cells. FP also decreased expression of LPS-induced inflammatory genes in C2C12 treated cells as compared with LPS alone. Taken together, the present study demonstrates in vitro feasibility of PLGA-FP nanoparticle delivery to the skeletal muscle cells, which may be beneficial for treating inflammation.

Nanodrug delivery platform for glucocorticoid use in skeletal muscle injury.

Glucocorticoids are utilized for their anti-inflammatory properties in the skeletal muscle and arthritis. However, the major drawback of use of glucocorticoids is that it leads to senescence and toxicity. Therefore, based on the idea that decreasing particle size allows for increased surface area and bioavailability of the drug, in the present study, we hypothesized that nanodelivery of dexamethasone will offer increased efficacy and decreased toxicity. The dexamethasone-loaded poly(lactic-co-glycolic acid) (PLGA) nanoparticles were prepared using nanoprecipitation method. The morphological characteristics of the nanoparticles were studied under scanning electron microscope. The particle size of nanoparticles was 217.5 ± 19.99 nm with polydispersity index of 0.14 ± 0.07. The nanoparticles encapsulation efficiency was 34.57% ± 1.99% with in vitro drug release profile exhibiting a sustained release pattern over 10 days. We identified improved skeletal muscle myoblast performance with improved closure of the wound along with increased cell viability at 10 nmol/L nano-dexamethasone-PLGA. However, dexamethasone solution (1 µmol/L) was injurious to cells because the migration efficiency was decreased. In addition, the use of dexamethasone nanoparticles decreased lipopolysaccharide-induced lactate dehydrogenase release compared with dexamethasone solution. Taken together, the present study clearly demonstrates that delivery of PLGA-dexamethasone nanoparticles to the skeletal muscle cells is beneficial for treating inflammation and skeletal muscle function.

Metformin's impact on statin-associated muscle symptoms: An analysis of ACCORD study data and research materials from the NHLBI Biologic Specimen and Data Repository Information Coordinating Center.

Statins are widely prescribed, yet statin muscle pain limits their use, leading to increased cardiovascular risk. No validated therapy for statin muscle pain exists. The goal of the study was to assess whether metformin was associated with reduced muscle pain. A secondary analysis of data from the ACCORD trial was performed. An ACCORD sub-study assessed patients for muscle cramps and leg/calve pain while walking, typical non-severe statin muscle pain symptoms. We compared muscle pain between patients using a statin (n = 445) or both a statin and metformin (n = 869) at baseline. Overall patient characteristics were balanced between groups. Unadjusted analysis showed fewer reports of muscle cramps (35%) and leg/calve pain while walking (40%) with statins and metformin compared to statin only (muscle cramps, 42%; leg/calve pain while walking, 47%). Multivariable regression demonstrated a 22% odds reduction for muscle cramps (P = 0.049) and a 29% odds reduction for leg/calve pain while walking (P = 0.01). Metformin appears to reduce the risk of non-severe statin muscle pain and additional research is needed to confirm the findings and assess metformin's impact on statin adherence and related cardiovascular outcomes.

Assessment of Cardiovascular Function and Microvascular Permeability in a Conscious Rat Model of Alcohol Intoxication Combined with Hemorrhagic Shock and Resuscitation.

Hypotension, cardiac depression, and elevated microvascular permeability are known problems that complicate resuscitation of patients following traumatic injury, particularly those who are also intoxicated from alcohol consumption. A conscious rat model of combined alcohol intoxication and hemorrhagic shock has been used to study the hemodynamic mechanisms involved. Here, we describe using this model to study microvascular leakage and cardiac electrical activity.