bluuebunny/crossref_metadata_2025 huggingface dataset:
MUC17 expression is differently regulated by TLRs interaction with their ligands
Authors: Resta-Lenert Silvia , Ho Samuel
Published: June 2021

MUC17 is a membrane‐bound mucin highly expressed on the apical surface of intestinal epithelia. We have shown that MUC17 promotes epithelial restitution, is anti‐apoptotic, protects intestinal cells against gut pathogens, and that live probiotics protect against enteropathogens, in part, by promoting MUC17 levels upon activation of TLR 2, 4. We hypothesized that TLRs and their ligands may affect MUC17 and other mucins, MUC1, MUC2 and MUC3, in an interdependent manner. MUC17 scrambled (sc) and siRNA (si) Caco2 and HT29 cells were exposed to LPS, PGNs, Pam2CSK4, LTAs, MPLAs, FLAs and ODNs to stimulate TLR 2, 4, 5, 9. TLRs antagonists and inhibitors of kinases were used to test mechanisms regulating MUC17. Mucins levels were assayed in both sc and si MUC17 cells. Protein, and nucleic acid analysis, and confocal microscopy were performed by standard methods. TLRs stimulation resulted in diverse levels of induction of mucins RNA and protein. This effect was dependent on type and dose of ligand tested. However, in MUC17 si cells, expression of MUC1, MUC2, and MUC3 were significantly altered after TLRs ligands treatment compared to MUC17 sc cells (MUC1 increased x2; MUC2 and MUC3 decreased x1.5 and x2, respectively). Inhibition of MAPK, IKK, PI3K reversed these effects. Our data suggest that MUC17 expression is significantly regulated by TLRs interaction with their ligands, and that levels of gut mucins show some degree of cross‐regulation. We conclude that the intestinal mucin barrier is more complexly regulated than previously thought, and that modulation of TLRs may reverse, in part, altered mucin expression observed in diseases of the gut. Funding: UCSD‐DDRDC; Research Service‐Dept Veterans Affairs
Adiponectin enhances murine colitis via adiponectine receptor 1 pathway
Authors: Peng Yu-Ju , Liu Bing-Hsien , Mersmann Harry John , Ding Shin-Torng
Published: June 2021

Adiponectin (ADN) is an adipokine derived from adipocytes. It binds to adiponectin recetor1 and 2 (AdipoR1 and 2) to exert into function regulating in whole‐body energy homeostasis, anti‐cardiovascular disease and inflammatory response. The role of ADN in inflammation response is likely multifaceted. It may play an anti‐inflammatory (e.g. diabetes, atherosclerosis and obesity) or proinflammatory (reheumatic disease and emphysema) role in different diseases. As the effect of ADN on colitis is still controversial, the goal of this study was to determine the inflammatory role of AdipoR1 in murine colitis. We generated AdipoR1‐transgenic mice to increase the effect of ADN and treated dextran sulfate sodium (DSS) to induce colitis. After administration of 2% DSS in drinking water for 7 days, AdipoR1‐transgenic mice developed more severe colitis than wild‐type mice, the symptoms including weight loss, bloody‐diarrhea, short coloength, crypt disruption and immunity cell infiltration. Our data show that ADN plays a proinflammatory role in colitis. Therefore, treatment of reducing ADN function during colitis may help the animal to cope with the inflammation.
Macromolecular Complex of CFTR, NHERF2 and iNOS at the Plasma Membrane Contributes to Diarrhea in IBD
Authors: Arora Kavisha , Yarlagadda Sunitha , Ren Aixia , Zhang Weiqiang , Sinha Chandrima , Naren Anjaparavanda P.
Published: June 2021

Macromolecular complex formation is a common theme in regulation of protein function. The cystic fibrosis transmembrane conductance regulator (CFTR), a cGMP/cAMP‐regulated chloride (Cl−) channel, is a well‐known hub protein involved in multiple protein complexes. In this study, we find that a signaling macromolecular complex of CFTR, Na+/H+ exchanger regulatory factor 2 (NHERF2), and inducible Nitric Oxide Synthase (iNOS) is formed at the plasma membrane in Inflammatory Bowel disease (IBD). We demonstrated the protein interactions using biochemical methods (e.g., protein pairwise binding assay, macromolecular complex assembly assay, and co‐immunoprecipitation) and in vitro methods for studying protein interaction in live cell (e.g., FRET). We conclude based on our in vitro (polarized colonic epithelial monolayers) and in vivo (DSS‐induced colitis in mice) functional studies that the interaction is integrative to CFTR dependent diarrhea in IBD. To summarize our findings: (a) CFTR interacts with iNOS mediated by NHERF2 in a PDZ domain‐dependent manner. (b) Upregulation of iNOS increased CFTR channel function by at least two folds in in vivo and in vitro models of inflammation. (c). Diarrhea was attenuated in Nherf2−/− mice compared to Nherf2+/+mice in DSS‐induced colitis model which simulates IBD. Taken together, our results demonstrate that the macromolecular complex of CFTR, NHERF2 and iNOS play important roles in the pathogenic process of diarrhea in IBD. Our study will not only help understand the molecular mechanisms underlying the pathogenic process of diarrhea in IBD, but help to identify novel therapeutic targets and agents for IBD therapy.
Cardiac Inflammation and Dysfunction During Dengue Virus Infection in Mice
Authors: Kangussu Lucas , Costa Vivian , Olivon Vania , Melo Marcos , Soriani Frederico , Rachid Milene , Santos Robson , Teixeira Mauro , Souza Danielle , Bonaventura Daniella
Published: June 2021

The aim of this study is to investigate the mechanisms involved in cardiac function alterations in animal model of Dengue. Balb/c mice were infected with adapted DENV‐3. The study was performed on 3 and 6 days after infection. We evaluated survival, hematological and inflammatory parameters, vascular permeability, and tissue damage. Systolic blood pressure and heart rate were determined. Furthermore, the oxidative stress and the cardiac function were analyzed. Infected mice died 7 days after infection. At day 3 and 6 we observed thrombocytopenia, hemoconcentration, increased levels of transaminases, neutrophil and macrophage accumulation in heart and elevated levels of TNF‐α, IFN‐α, IL‐6, IL‐1β, CXCL1, CXCL2 and IL‐18 in serum and heart. These findings were accompanied by changes in hemodynamic parameters. There was a significant reduction in GSH levels, and increased lipid peroxidation, as well as, SOD and catalase activity in heart. Impaired cardiac function was verified with reduction in cardiac output, end‐diastolic diameter, endsystolic diameter, stroke volume, end‐diastolic volume, and endsystolic volume. Infection of mice with adapted DENV‐3 induced clinical manifestations of severe dengue infection, as observed in humans. Heart is target for Dengue virus infection, inducing intense inflammatory process and oxidative stress, associate with cardiac dysfunction.

Financial Support: INCT Dengue, CAPES, FAPEMIG and CNPQ.
Cardiac myocyte‐specific caveolin‐3 overexpression alters β‐adrenergic receptor activity
Authors: Busija Anna Rebecca , Roth David M. , Insel Paul A. , Patel Hemal H.
Published: June 2021

Caveolins are scaffolding proteins essential to the formation of caveolae, plasma membrane invaginations that compartmentalize signaling molecules. Mice with cardiac myocyte‐specific overexpression of caveolin‐3 (Cav‐3 OE) have increased caveolae number in cardiac myocytes and are protected from ischemia‐reperfusion injury, while mice lacking Cav‐3 (Cav‐3 KO) lack cardiac caveolae and are more sensitive to cardiac injury. β‐adrenergic receptors (β‐ARs) localize to caveolae and when stimulated by agonists alter cardiac inotropy and chronotropy but whether increased Cav‐3 expression alters β‐AR agonist response in the heart is unknown. We treated Langendorff‐perfused hearts from transgene‐negative (TGneg), Cav‐3 KO, and Cav‐3 OE mice with the β‐AR agonist isoproterenol (Iso). Cav‐3 OE mice had augmented Iso‐stimulated inotropy (+dP/dt) and lusitropy (−dP/dt) relative to TGneg mice but response was reduced in Cav‐3 KO mice. Cardiac myocytes (CM) isolated from Cav‐3 OE mice had enhanced Iso‐stimulated production of cAMP while Cav‐3 KO CM had reduced cAMP production. These data indicate that the expression of Cav‐3 is a key modifier of β‐AR‐promoted cAMP formation and contractile function. Physiological or pathological alteration in Cav‐3 expression thus has the potential to modulate cardiac response to sympathetic activation. Research supported by VMIRF and NIH research grants.
Antioxidants N‐acetylcysterine and Allopurinol synergistically enhance cardiac HIF‐1α and heme oxygenase‐1 and attenuate Postischemic Myocardial Injury in Diabetic Rats
Authors: Mao Xiaowen , Liu Yanan , Wang Tingting , Lei Shaoqing , Irwin Michael Garnett , Vanhoutte Paul M. , Xia Zhengyuan
Published: June 2021

Hypoxia inducible factor 1(HIF) α can reduce myocardial ischemia‐reperfusion injury(MIRI). However, hyperglycemia‐induced oxidative stress may reduce cardiac HIF‐1α and subsequently inhibit heme oxygenase 1 (HO‐1), a down‐stream protein of HIF‐1α that has anti‐ischemic properties. N‐acetylcysteine (NAC) and allopurinol (ALP) synergistically reduce MIRI in diabetes (PLoS One. 2011;6:e23967), but the role of HIF‐1α/HO‐1 in this process in unknown. Control or streptozotocin (STZ)‐induced diabetic rats were untreated (C, D) or treated with NAC (1.5g/kg/day) or ALP (100 mg/kg/day) or their combination (NAC+ALP) for four weeks starting one week after STZ injection. Cardiac and plasma 15‐F2t‐isoprostane (IsoP) were increased in D rats while cardiac HO‐1 and protein expression and activity were reduced, accompanied with reduced cardiac HIF‐1α, and increased post‐ischemic myocardial infarct size (IS) and cellular injury in D rats subjected to 30 minutes of coronary artery occlusion and 2 hours of reperfusion (all P<0.05 D vs. C). NAC+ALP normalized cardiac levels of HO‐1 and HIF‐1α protein expression, prevented the increase in IsoP, and reduced myocardial IS, but these effects of NAC+ALP were cancelled by either the HO‐1 blocker protoporphyrin or the HIF‐1α blocker 2‐Methoxyestradiol. It is concluded that HIF‐1α and HO‐1 activation play an important role in NAC and ALP mediated cardioprotection in diabetes.
Hyperbaric oxygen therapy effect in vascular reactivity
Authors: Estrada Norma , Guevara-Balcazar Gustavo , Villanueva-Lopez Guadalupe Cleva , Mera-Jimenez Elvia , Lara-Padilla Eleazar , Castillo-Hernandez Maria C
Published: June 2021

Introduction

The oxygen (O2)is a source of energy and for cellular respiratory support, deficient of oxygen delivery in tissue, can affect the cellular functions, We know that flow warranty the oxygen concentrations in tissue, is for that reason the new use of oxygen like therapy, because of we want to evaluate a possible changes with the hyperbaric oxygen therapy (HBO) in vascular reactivity, the information will be useful to suggest treatment with HBO in patients with cardiovascular problems.

Methods

Male rats were used between 250–300gr, tree months old, in two groups with and without hyperbaric oxygen therapy, rats were exposed to pure oxygen at 2 atmospheres absolute during 60 min for 3, 7, 10 and 20 sessions, the aortic rings were introduce immediately in the organ bath, after that we probe phenileprine (10−9 – 10−5 M), acetylcholine (10−9 – 10−5 M) and sodium nitroprusiate (10−11–10−6 M)

Results

The HBO increase relaxation dependent of the endothelium, after 20 sessions in the thoracic segment and early in the abdominal segment, we shown that the independent relaxation in the endothelium is mediated for sodium nitroprusiate is increase in the first days of exposition in both segments, regularization between 10 to 20 session of HBO, on the other hand phenyleprine contraction shown a decrease in the maximum effect after 20 sessions and this is related with the increment in the relaxant response in the endothelium, and this suggest a mayor ON release.
Bivalirudin Emulsions Demonstrate Efficacy of a Nanoparticle Strategy for Inhibition and Imaging of Thrombosis
Authors: Myerson Jacob Wheatley , He Li , Allen John Stacy , Williams Todd A , Tollefsen Douglas M , Lanza Gregory M , Caruthers Shelton D , Wickline Samuel A
Published: June 2021

A thrombin‐inhibiting perfluorocarbon nanoparticle (PFC NP), functionalized by PPACK (Phe(D)‐Pro‐Arg‐Chloromethylketone), was recently presented as a prototype for a novel class of targeted antithrombotic. Here, a NP functionalized with Bivalirudin (BVR), was compared to BVR and the PPACK NP.

PPACK or BVR were covalently attached to PFC NPs. Optical assay verified that PPACK and BVR selectivity and activity against thrombin was not diminished on the NPs. In vivo activity was assessed for PPACK NPs, PPACK, BVR, BVR NPs, heparin, non‐functionalized NPs, or saline in C57BL6 mice subjected to laser injury of the carotid artery. Time to thrombotic occlusion of the injured artery was assessed via Doppler flow measurement. Selected arteries were excised to assess NP retention via 19F magnetic resonance (MR). 3T MRI of thrombosis following administration of PPACK NPs was also demonstrated in cholesterol‐fed rabbits.

Previously, PPACK NPs outperformed both heparin (p=.001) and PPACK (p=.0006) in delaying occlusion of the carotid artery. PPACK or non‐functionalized NPs failed to delay occlusion of the carotid artery. BVR NPs significantly delayed occlusion (p=.02) whereas an equivalent dose of free BVR (120 mg/kg) did not (figure 1a). 19F MR captured thrombin‐specific PFC NP retention in occluded arteries of mice (figure 1b) and rabbits (figure 2).
Palmatine a Novel Anti‐adipogenic and Anti‐diabetic Alkaloid from an Indian Medicinal Plant Tinospora cordifolia
Authors: Vasanthi Hannah Rachel , Kannan Sangeetha Marimuthu
Published: June 2021

Palmatine a naturally existing alkaloid isolated from a medicinal plant Tinospora cordifolia commonly used in Indian System of Medicine for various ailments has been tested for its biological activities. Palmatine in 3T3‐L1 (murine pre‐adipocytes) cells effectively inhibited adipocyte differentiation and promoted glucose consumption. Quantification of genes (Glut‐4, PPARα, PPARγ, LPL, FAS and Adiponectin) involved in diabetes and obesity using Real Time PCR and comparing the activity of Palmatine with specific inhibitors of insulin pathway (IRTK, PI3K, Glut 4) reveals that Palmatine is a novel anti‐adipogenic and anti‐diabetic molecule. It is also evidenced by the up regulation of Glut‐4 and PPARα; and down regulation of PPARγ, LPL and FAS. Oxidative stress is the common pathway linking and intensifying in all metabolic disorders. However, the anti‐oxidant property of Palmatine studied by hydrogen peroxide induced cytotoxicity study in 3T3 L1 cells was found to be mild as compared to the proven antioxidant flavonoid Quercitin. In conclusion, the results strongly support the anti‐diabetic, anti‐adipogenic and mild antioxidant activity of the naturally occurring Palmatine in Tinospora cordifolia an antidiabetic‐medicinal plant used in India which could serve as a lead pharmaceutical agent to benefit the growing diabetic/obese population. The work was sponsored by the Department of Science & Technology, Govt. of India
Salivary gland transcriptome analysis of the Wild living and Captive Spotted Hyena
Authors: Osei-Boadi Kate , Linde Annika , Verreynne Erik , Melgarejo Tonatiuh
Published: June 2021

The African Spotted Hyena is a mammalian species with a strong immune system in spite of exposure to high bacterial loads in their diet. Hyenas are renowned for their unique adaptability and exceptional ability to fend off pathogens. This makes them a good model for studying the immune system in an attempt to discover immune related genes. Host defense peptides (HDPs) with antimicrobial activity against different micro‐organisms have been isolated from many species including human beings. Apart from being antimicrobial, HDPs also possess other biological activities such as chemotaxis and wound healing promoters. In an attempt to understand how the Spotted Hyena achieves its immunological dominance transcriptome analysis of the salivary gland of Wild living Hyena was carried out. Total RNA was isolated and purified from salivary gland samples from male and female wild living Spotted Hyena using the modified Ambion protocol. Total RNA was quantified using Agilent 2100 Bioanalyzer and the NanoDrop Spectrophotometer. Messanger RNA (mRNA) was selected from the total RNA of the female wild living Hyena and sequenced using 454 sequencing. The yield of total RNA as determined by the Bioanalyzer was 424ng/mL. A total of over 500 Mb with an average read length of 380 bps was obtained from the mRNA sequencing. Analyzing the salivary gland transcriptome of the Hyena has clinical relevance especially in infectious disorders and disease prevention. It will help us understand pathways and mechanisms that regulate disease progression and how altered gene expression contributes to diseases. (COBRE P20 – RR017686 from NCRR NIH)
Macrophage pro‐angiogenic miRNA, miR‐27b, is under NF‐kB transcription regulation
Authors: Pan Hua , Clarke Bradley , Wickline Samuel A
Published: June 2021

Atherosclerosis is the common trigger of cardiovascular diseases, which is a leading cause of mortality in US. It exhibits plaque development with chronic inflammation as a prominent feature. The NF‐κB signaling pathway is a pivotal mediator of the chronic inflammatory process that drives atherosclerotic plaque development, and has been proposed as a therapeutic target for clinical management. Additionally, microRNA species are rapidly being uncovered as equally prominent components of atherosclerosis. But little is known about the interplay between NF‐κB signaling and microRNA in this context. Here, we report that pro‐angiogenic miRNA, miR‐27b, is regulated by NF‐κB signaling in RAW267.4 cells under TNF‐α stimulation.

First, we evaluated the dose and time dependence of TNF‐α stimulated NF‐κB signaling activation in RAW267.4 cells. We found that the nuclear translocation of NF‐kB protein, p65, peaked 30 minutes after treatment with 10 ng/ml TNF‐α. To investigate the interplay between NF‐κB signaling and microRNA, we specifically inhibited TNF‐α stimulated NF‐κB activation by treating TNF‐α activated RAW264.7 cells with Bay 11‐7082. The time dependent evaluation demonstrated that 30 minutes 5 μM Bay 11‐7082 inhibited TNF‐α induced NF‐κB activation in RAW264.7 cells. We then treated RAW264.7 with or without 10 ng/ml TNF‐α for 30 minutes and with or without Bay 11‐7082. Then total small RNA was extracted by using TRIzol reagent and evaluated for the expressions of 88 inflammation‐related microRNAs by polymerase chain reaction (PCR)‐based arrays. Among those 88 inflammation‐related microRNAs, miR19a, miR‐27b, and miR 195 are significantly upregulated (more than 7‐fold increase) after TNF‐α stimulated. After Bay 11‐7082 inhibition, miR‐27b expression level is not significantly different from the control cells. These results suggest that miR‐27b may represent a therapeutic target for clinical management of inflammation in atherosclerosis.
Anti‐inflammatory activities of Cassia alata extract in Complete Freund 's Adjuvant arthritis in rats
Authors: Lewis Anishka Triciana , Levy Arkene Stacy-Ann
Published: June 2021

Cassia alata (C. alata) is a plant belonging the family Caesalpiniaceae and native to tropical America. The anti‐inflammatory mechanism of a hexane extract of C. alata was investigated in Complete Freund's Adjuvant (CFA) arthritis, as a chronic model of inflammation. The extract was prepared and administered to CFA arthritic animals at 500mg/kg. The CFA model was created by the injection of 0.5ml CFA into the synovial cavity of the right knee joint of hind leg of rats. Changes in knee joint swelling, cartilage integrity and synovial fluid leukocyte counts were assessed in response to C. alata treatment. Assessment of the reduction in knee joint swelling was performed by obtaining the circumference of the knee joint. Histological analysis was done to assess the cartilage erosion. C. alata significantly (p = 0.009) reduced knee joint swelling and provided protection against cartilage degradation. The migration of leukocytes to the blood (p= 0.002) and synovial cavity (p=0.019, 0.012, 0.028 and 0.002) was also significantly reduced. These results suggest potent antiinflammatory activities for C. alata that could be potentially exploited for arthritis therapy. Funding was provided by the Graduated Studies Department at the University of the West Indies.
Manipulation of O‐linked‐β‐N‐acetylglucosamine (O‐GlcNAc) levels in HL‐1 cardiomyocytes
Authors: Medford Heidi M , Hall Kyla E , Marsh Susan A
Published: June 2021

The post‐translational attachment of O‐linked‐β‐N‐acetylglucosamine (O‐GlcNAc) to serine/threonine residues of proteins is a dynamic and reversible process analogous to phosphorylation. Previous in vivo and in vitro studies have shown that O‐GlcNAc can play both beneficial and detrimental roles in the heart; however, in vitro studies have been limited to primary cardiomyocytes and immortalized cardiomyoblasts. The immortalized mouse atrial myocyte line, HL‐1, has demonstrated promise in preliminary studies but it has not been used for O‐GlcNAc applications. Therefore, the goal of this study was to determine whether O‐GlcNAc is present in HL‐1s and, if so, whether O‐GlcNAc levels can be modified using standard approaches. HL‐1 cardiomyocytes were grown in culture to 100% confluence then treated with various concentrations of hexosamine biosynthesis pathway substrates (glucose, glucosamine), as well as inhibitors of O‐GlcNAcase (PUGNAc, Thiamet‐G), the enzyme that removes O‐GlcNAc, for 24 hours. As expected, all of these treatments increased O‐GlcNAc levels. These results are consistent with previous in vivo and in vitro findings. The ability to manipulate O‐GlcNAc levels in HL‐1s is an important finding as it demonstrates suitability of this cell line for preliminary studies without the challenges associated with isolating primary cardiomyocytes.
Neonatal growth restriction enhances central leptin sensitivity in adult mice
Authors: Peotta Veronica , Cushman Taylor , Dexter Benjamin , Aldape Gilbert , Hermann Gregory , Roghair Robert
Published: June 2021

Neonatal growth restriction (GR) leads to neonatal leptin deficiency and adult hypertension. We speculated leptin deficiency during a window of developmental plasticity permanently increases central leptin sensitivity. We thus hypothesized that neonatal GR enhances adult leptin signaling and that neonatal leptin supplementation would be protective. C57BL/6 mice were raised in litters of 6 or 12 to generate control and GR pups, respectively. Within each litter, pups were randomized to vehicle (0.9% NaCl) or leptin (1 mcg/g, ip) from P4 to P14. At 4 months, blood pressure and heart rate were measured by radiotelemetry. Additional mice underwent perfusion fixation after injection of vehicle or leptin (1 mcg/g ip). GR mice had decreased food intake (0.09±0.01 vs. 0.13±0.02g/g/d; p=0.02) and lower body weight (26±1.3 vs. 32±1.1g; p=0.01). Leptin supplementation normalized food intake (0.13±0.01g/g/d), but not adult body weight (28±1.4g; p=0.01). Adult GR mice had increased heart rates (576±5 vs. 546±bpm; p=0.004) and significantly increased pressor responses to leptin (2 mcg icv) that correlated with enhanced pSTAT3 signaling in the arcuate nucleus as well as increased c‐Fos expression in the subfornical organ. We conclude: 1) neonatal GR enhances central leptin sensitivity, and 2) leptin supplementation during a critical neurodevelopment window may be protective. (HL102659)
Resveratrol protects against diabetes‐induced endothelial dysfunction in high‐fat diet fed mice
Authors: Li Dai-Er , Hung Li-Man
Published: June 2021

Epidemiological studies have shown that red wine consumption is associated with reduced in cardiovascular mortality in the general population and diabetic patients. The present study was designed to investigate whether resveratrol (RSV, a red wine antioxidant) can attenuate diabetes progression, improves diabetes‐related vescular endothelial dysfunctions, and delineated further its underlying mechanisms.

Male C57Bl/6 mice were fed with high‐fat diet (HFD) for 17 weeks. Animals developed type 2 diabetes characterized by elevated body weight, hyperglycemia, hyperinsulinemia, and hyperlipidemia. Oral gavage fed with RSV (5 or 10 mg/kg/day for 17 weeks) significantly reduced body weight, plasma glucose, insulin, and triglyceride compared to non‐treated HFD mice. RSV treatment also attenuated plasma glucose elevation and improved insulin responses during glucose tolerance test. Furthermore, HFD diabetic mice exhibited an increasing the numbers of leukocyte rolling, adhering, and transmigrating in the post‐capillary venules of cremaster muscle. In contrast, treatment of RSV significantly attenuated leukocyte rolling, adhesion, and transmigration in HFD diabetic mice. The phenylephrine (PE)‐induced vasoconstriction was dramatically attenuated in HFD diabetic mice; whereas, RSV treatment significantly improved the vessel responsiveness to PE. Our result also shows that the phosphorylated AMPK (5′AMP‐activated protein kinase), Akt, and eNOS (endothelial nitric oxide synthase) protein levels were significantly reduced in aorta of HFD mice. Consistence with the observation on improvement of blood vessel responsiveness, RSV also elevated AMPK, Akt, and eNOS protein phosphorylation levels. Taking together, these results indicate that RSV attenuated diabetes‐related vascular endothelial dysfunctions, at least in part, by elevation of AMPK, Akt, and eNOS proteins phosphorylation.
Membrane localization of FK506‐binding proteins FKBP51 and FKBP52, immunophilins that are part of the endothelial store‐operated calcium entry heterocomplex
Authors: Kadeba Pierre I. , Scammell Jonathan G. , Cioffi Donna L.
Published: June 2021

Disruption of the endothelium leads to increased vascular permeability. Activation of store‐operated calcium (SOC) entry promotes inter‐endothelial cell gap formation. Canonical transient receptor potential proteins 1 and 4 (TRPC1/4) are subunits of the calcium‐selective SOC entry channel Isoc, and rat pulmonary artery endothelial cells (PAECs) and pulmonary microvascular endothelial cells (PMVECs) express the Isoc channel subunits. Data from our laboratory have implicated FK506 binding proteins (FKBPs) as regulators of SOC entry with FKBP52 facilitating and FKBP51 inhibiting activation. However, mechanisms by which immunophilins regulate SOC entry are unknown. We first sought to determine whether the immunophilins localize to the endothelial membrane where the TRPC1/4 heterocomplex is found. Membrane‐cytoskeleton preparations of PAECs and PMVECs resolved FKBP51 and FKBP52 in plasma membrane fractions, and immunocytochemistry revealed localization to cell‐cell borders. Next, co‐immunoprecipitation experiments were performed to determine whether these immunophilins contribute to the endothelial TRPC1/4 heterocomplex. We observed that FKBP52 co‐immunoprecipitated with TRPC4. Collectively, these data reveal membrane localization of FKBP51 and FKBP52, suggesting that they might contribute to the endothelial SOC entry heterocomplex. Supported by 5R00HL089361
Role of NO and Superoxide in Hyperhomocysteinemia Induced Cardiac Metabolic Dysfunction in Pregnant Cystathionine Beta‐synthase Heterozygote Mice
Authors: Song Su , Kertowidjojo Elizabeth , Ojaimi Caroline , KandhiI Sharath , Wolin Michael , Hintze Thomas H
Published: June 2021

Genetic defects in cystathionine beta‐synthase (CBS) can cause severe hyperhomocysteinemia (HHcy). HHcy causes cardiovascular dysfunction and is associated with many complications during pregnancy related to reduced NO bioactivity. This study assessed the mechanisms of HHcy on the NO‐dependent control of cardiac O2 consumption in pregnant CBS heterozygote (HET) mice. Pregnant CBS HET mice (n=8) were fed low‐folate diet for 20 days to increase plasma Hcy (8.2±0.5 of WT to 17.9±2.2 uM of HET). As compared to wild type (WT) (n=10), while heart weight/body weight increased by 18.2%, ejection fraction decreased from 85.7±0.8% to 80.3±1.8% in HET. Systolic blood pressure was significantly increased from 92.5±3.6 mmHg of WT to 114.5±4.0 mmHg in HET, and MBP from 66.8±9.2 mmHg to 82.9±5.5 mmHg, without hypertension. The total peripheral resistance significantly increased by 42.1%. The ability of bradykinin to reduce myocardial O2 consumption in vitro was impaired by 56% in heart. The response was restored by ascorbic acid, tempol and apocynin. O2− production was increased by 2.7 folds in Hcy heart using lucigenin chemiluminescence.

Conclusion

CBS deficiency caused HHcy during pregnancy, impairs cardiac function, increases O2− production and reduces the ability of NO to regulate myocardial O2 consumption probably through NADPH oxidase stimulation.
GLOBAL GENE EXPRESSION PROFILE IN RENAL ISCHEMIA/REPERFUSION INDUCED CARDIAC HYPERTROPHY
Authors: Nakama Karina Kaori , Carneiro-Ramos Marcela Sorelli
Published: June 2021

Objectives

The heart is a target organ in many regulatory systems, such as hormones, neuronal and immunological systems, modulating heart trophism. This study aimed to determine changes in gene expression of hypertrophied cardiac tissue in a renal ischemia‐induced systemic inflammation model.

Methods

C57BL/6 mice were submitted to unilateral occlusion of left renal pedicle for 60 min, followed by reperfusion of 5, 8, 15 and 20 days. The software dChip and GeneMAPP were used to separate and group genes in signaling pathways which had shown significant increase or decrease in expression. Three genes (H2‐Oa, Crtam and Ifnz) of very modulated pathways associated with immune response (MHC class II, Positive Regulation of Natural Killer Cell and Cytokine Activity Pathways, respectively) were selected. To validate array results, real time PCR was performed.

Results

The analysis results indicated 17,413 up‐regulated and 18,297 down‐regulated genes, and 405 up‐regulated and 300 down‐regulated pathways. According to the array analysis and real time PCR results the three selected genes were up regulated and showed a greater increase after 8 days of reperfusion.

Conclusion

Validation of H2‐Oa, Crtam and Ifnz indicates a possible relationship of these genes in the development of cardiac hypertrophy due to a systemic condition of inflammation caused by renal failure.
Association of mitogen activated protein kinase genetic polymorphism with carotid intima medial thickness (CIMT) in women enrolled in the Kronos Early Estrogen Prevention Study (KEEPS)
Authors: Miller Virginia May , Petterson Tanya M , Jeavons Elysia N , Heit John A , Harman S Mitchell , de Andrade Mariza
Published: June 2021

Menopausal hormone treatments (MHT) may limit progression of CIMT, a measure of atherosclerosis, but pose a risk for ischemic stroke. To test candidate gene variation for an association with CIMT, DNA from 714 women participating in KEEPS, a clinical trial of MHT to prevent progression of atherosclerosis, was genotyped using custom Illumina Infinium iSelect chemistry. Single nucleotide polymorphisms (SNPs; 13,229) were genotyped within 769 genes within the anticoagulant, procoagulant, fibrinolytic or innate immunity pathways. Of these SNPs, 75 were removed due to call rate<0.95, relatedness, duplication, and misidentification; 1275 were removed for quality control. There were 504 ancestry informative markers (AIM). Percent of European ancestry using AIM, age at enrollment and pulse pressure correlated positively with CIMT. Using logistic regression to adjust for these, one SNP (rs2236935 of MAP4K4 on chromosome 2) associated positively with CIMT (p‐value = 5.771e‐06). This gene encodes a protein of the serine/threonine protein kinase family involved in cytokine signaling pathways related to innate immunity, development of type II diabetes, and response to environmental stress. Whether this SNP remains associated with changes in CIMT in women randomized to hormonal treatment in KEEPS remains to be determined. (Supported by grants from the Aurora Foundation, NIH HL90639 and the Mayo Foundation)
Mutations of HOMEZ gene in Congenital Heart Diseases
Authors: XUAN Chao , JIA Ke-Gang , WANG Bin-Bin , BAI Xiao-Yan , GAO Ge , YANG Qin , LIU Xiao-Cheng , MA Xu , HE Guo-Wei
Published: June 2021

Genome‐wide linkage analysis revealed that a potential congenital heart disease (CHD) susceptibility locus on Homeodomain leucine zipper‐encoding (HOMEZ) gene in a South Indian population. We aimed to identify potential pathogenic mutations for HOMEZ in isolated VSD in the Chinese population. Case‐control mutational analysis was performed in 400 patients with isolated VSD and 400 healthy controls by using PCR and sequencing. Two novel heterozygous missense mutations (c.116 C>T; c. 630T>A) were identified in HOMEZ gene exon‐2. The two mutations lead to alanine to valine substitution at the position 39 and serine to arginine at position 210 which are highly conservative among many species. The hydropathicity of the valine and arginine residue at the position 39 and 210 were significantly different from the wild type. We have identified two novel heterozygous missense mutations in HOMEZ gene exon‐2 in isolated VSD patients and have found that these two mutations resulted in alteration of the hydropathicity of the HOMEZ protein. Therefore, the two missense mutations of HOMEZ gene are directly linked with the etiology of isolated VSD in the Chinese population.

Supported by National Basic Research Program of China 2010CB529502; International S&T Cooperation Program of China 2009DFB30560; Tianjin Municipal S&T Com 09ZCZDSF04200 & 10JCYBJC26400; NSFC 81170148; GRF CUHK4789/09M; CUHK2041688
CD4+ T cells Regulate Cardiac Ischemia/Reperfusion Injury
Authors: Zuidema Mozow Yusof , Li Jun , Lee Sewon , Meininger Gerald A , Hill Michael A , Dellsperger Kevin C , Zhang Cuihua
Published: June 2021

The mechanisms of healing following myocardial infarction are multifaceted. We tested the hypothesis that chronic, post‐ischemic, cardiac inflammation is regulated by CD4+ T cells through cytokine release and metabolism, and this leads to impaired cardiac function. For these studies, a mouse model of myocardial I/R (30 minutes/24 hours) was employed using wild type (WT) C57Bl/6J mice and CD4+ −/− mice. Based on several hemodynamic measures (EF, SW, dP/dtmax), the cardiac function of CD4+ −/− mice was impaired. Significant induction of monocyte chemotactic protein‐1 (1.9x; MCP‐1) and interferon gamma (3.4x; IFNγ) protein expression occurred following I/R in WT but was absent in CD4+ −/− (n=4; p<0.05). These observations suggested that CD4+ T cells play a pivotal role in the recruitment of innate immune responses following I/R. CD4+ −/− mice had increased cardiac TNFα (3x), ADAM10 (3x), ADAM17 (1.5x), and TLR‐2 (2.3x) mRNA expression relative to WT (n=5; p<0.05). Collectively, these results suggest that CD4+ T cells mediate cardiac dysfunction via TNFα metabolism and inflammatory cascades linked to MCP‐1, IFNγ, and TLR‐2. We propose that CD4+ T cells may bridge the innate immune response to the adaptive immune response in cardiac I/R. This may provide novel mechanistic strategies to target CD4+ T cells and treat chronic reperfusion injury syndromes. Supported by HL077566, HL085119, AHA SDG and AHA 10POST3870022.
Uterine artery vasoconstriction during exercise in rat pregnancy
Authors: OˈHagan Kathleen P , Lashley Christopher , Supik David A , Atkinson James T , Murphy Robert J
Published: June 2021

Dynamic exercise initiates a decrease in uterine artery conductance (UtC) that is reduced at term pregnancy in rabbits [−40% (SD20) vs. −7% (SD15)]. We hypothesized that a gestational attenuation of the uterine vasoconstrictor response occurs in rats, a common model for pregnancy research. Nonpregnant (8NP) and pregnant (12P) rats were familiarized w/treadmill running and instrumented with a left carotid arterial catheter to measure blood pressure (BP) and a Transonic flowprobe for measurement of right UtBF. At day20 gestation (7–10d postsurgery), rats underwent exercise at 7 m/min (6%grade) for 5 min. Values are mean (SD). Pre‐exercise BP was lower [126 (13) vs. 111 (13) P=0.02], HR similar [456 (34) bpm] and UtBF [0.46 (0.25) vs 2.1 (1.6) ml/min, P<0.01] and UtC [0.35 (0.17) vs. 2.1 (1.7) ml‐min−1/mmHg × 10−2P<0.01] elevated in the P rats. BP and HR increased with exercise in P and NP. Averaged over min 2–5 of exercise, the reduction in UtBF [−47% (12) vs. −25% (20), P=0.02)] and UtC [−49% (12) vs. −30% (20), P=0.02] was mildly attenuated in the P rats. Similar to rabbits, P rats demonstrate a reduced uterine vasoconstrictor response to dynamic exercise, although it appears less pronounced in the rat.

Supported by MWU.
NOS1‐dependent S‐nitrosylation of cardiac calcium‐handling proteins (RyR2, SERCA, and L‐type Ca2+ channel) modulates basal contractility and adrenergic inotropism
Authors: Vielma Alejandra Z , Eller Gisela , González Daniel R , Boric Mauricio P
Published: June 2021

We tested the hypothesis that NOS‐1 activity regulates basal and β‐adrenergic‐stimulated cardiac contractility by S‐nitrosylation of Ca2+‐handling proteins.

We used the isolated rat heart preparation and biotin‐switch assay, to study the effect of selective NOS‐1 inhibitor (S‐methyl‐L‐thiocitrulline, SMTC) on the contractile response to isoproterenol (ISO) and S‐nitrosylation of sarcoplasmic Ca2+ channel (RyR2) and pump (SERCA2), L‐type Ca2+ channel (LTCC), and unspecific total cardiac proteins (TCP).

SMTC (300nM, 20‐min) significantly decreased basal contractility (−15%) and the S‐nitrosylation of RYR2 (−70%), SERCA2 (−50%), LTCC (−80%), and TCP (−55%). ISO (10nM, 3‐min) increased S‐nitrosylation of RyR2 (+140%) and TCP (+30%), but did not change S‐nitrosylation of SERCA2 and LTCC. In the presence of SMTC, ISO caused proportionally similar changes in S‐nitrosylation of all these proteins, but the maximal inotropic response was reduced (−30%). Furthermore, in isolated mice cardiomyocytes, SMTC reduced ISO‐elicited maximal sarcomere shortening (−50%) and [Ca2+]i transient amplitude (−50%), and increased the time‐constant for [Ca2+]i decay (+30%).

Thus, rapid and reversible NOS‐1 dependent S‐nitrosylation of specific Ca2+‐handling proteins is essential to keep basal cardiac contractility, and to accomplish a full inotropic response.

Supported by Grants Fondecyt 1090757 & Anillos‐ACT71
Iron supplementation recommendations during pregnancy: Case study of WHO, CDC and India Government policies
Authors: Young Melissa Fox , Wendt Amanda , Ramakrishnan Usha , Martorell Reynaldo
Published: June 2021

Iron deficiency and anemia remain significant public health problems affecting pregnant women worldwide, despite the existence of guidelines for iron supplementation since the 1960s. The purpose of this paper was to examine current policy recommendations for prenatal iron supplementation. We reviewed current recommendations and evaluated evidence used to develop policy using three cases studies, World Health Organization (WHO), Centers for Disease Control and Prevention (CDC), and the Government of India. Information was obtained by searching websites, PubMed databases and contacting key informants. All cases set separate guidelines for routine iron supplementation (dose: 60, 30, 100 mg respectively; duration/initiation: immediately to delivery (at least 6 mo), 1st prenatal visit to delivery, after 1st trimester for 100 days) and for those considered anemic and/or severely anemic (dose: 120, 60–120, 200mg; duration: 3mo, until normalized, minimum 100 days). The evidence used to inform policy decisions varied but focused on estimates of iron requirements, anemia burden and reduction/improvement of anemia/hemoglobin. The quality of evidence from trials was constrained by methodological concerns and limited attention to clinical outcomes, compliance and adverse events (hemocentration). A reevaluation of high dose iron recommendations may be warranted. Funding support provided by a BMGF grant through a subcontract with CARE‐India.
Iron supplementation during lactation increases hemoglobin without an increase in iron status or oxidative stress
Authors: Jorgensen Josh M , Yang Zhenyu , Lonnerdal Bo , Chantry Caroline J , Allen Lindsay H , Dewey Kathryn G
Published: June 2021

The RDA for iron (Fe) decreases to 9 mg/d during lactation, yet breastfeeding women are counseled to continue taking prenatal vitamin‐mineral supplements (PNV) postpartum. Our objective was to examine the effect of Fe‐containing PNV taken postpartum on biomarkers of Fe status and oxidative stress. Lactating women (n=114) were randomly assigned to consume daily one Fe‐free PNV plus either 27 mg of Fe (as Fe sulfate) or placebo for 3 mo. The placebo group took the tablets between meals, while those in the Fe group took the tablets either with (Fe‐with) or between meals (Fe‐between). Blood and urine samples were collected before and after the supplementation period to analyze hemoglobin (Hb), Fe status (ferritin, hepcidin, transferrin saturation (TfSat), and total plasma Fe) and oxidative stress (isoprostane and 8‐ hydroxy‐2‐deoxyguanosine (8‐OHdG)). The change from baseline in Hb was greater in the Fe‐with and Fe‐between groups compared to placebo (0.04, 0.25, and −0.37g/dL, respectively; P=0.027), yet only 1 woman in the placebo group had Hb < 12 g/dL at the end of the intervention period. There were no significant differences in ferritin (P=0.09), hepcidin (P=0.84), TfSat (P=0.08), or total plasma Fe (P=0.07). There were no group‐wise differences in isoprostane (P=0.64) or 8‐OHdG (P=0.67). Fe consumption during lactation increased Hb without a significant increase in Fe status and did not affect oxidative stress.

Grant Funding Source: Center for Health and Nutrition Research at UC Davis
Titin is not the weak link in the Z‐disk streaming induced by plyometric exercise
Authors: Macaluso Filippo , Isaacs Ashwin W , Myburgh Kathy H
Published: June 2021

The purpose was to analyze the skeletal muscle changes induced by an acute bout of plyometric exercise (PlyEx) before and after a PlyEx training.

Healthy untrained individuals (N=13) completed an acute bout of PlyEx (10×10 squat‐jumps, 1 min rest). Thereafter, 8 subjects completed 8 weeks of PlyEx, while 5 subjects were instructed to refrain from doing any jumping activity for 8 weeks. Seven days after the last training session all subjects repeated a second acute bout of PlyEx.

The first acute bout of PlyEx induced a rise of creatine kinase activity (CK = 2421 ± 4978 U/l), Z‐line streaming mainly in fast twitch fibers, a stretch of the c‐terminus of titin (observed by immunogold) not limited to fast twitch fibers, and an increase in calpain‐3 autolysis. The second acute bout of PlyEx in the trained group did not induce: a rise of CK (110 ± 58 U/l), Z‐line streaming, or calpain‐3 activation; while in the control group only two subjects exhibited Z‐line streaming, although all subjects presented a stretch of titin.

Eccentric explosive exercise induced a stretch of titin, which is not responsible for the weakness of the sarcomere and of the calpain‐3 activation involved in sarcomeric remodelling.

Grant Funding Source: Claude Leon Foundation, National Research Foundation (NRF)
Human skeletal muscle microvascular blood volume: effects of ageing, feeding and exercise training
Authors: Phillips Bethan , Varadhan Krishna , Atherton Philip , Rennie Michael , Smith Kenneth , Williams John
Published: June 2021

Perfusion of the skeletal muscle microvasculature in response to hyperaemic stimuli such as feeding and exercise is crucial for whole‐body (e.g. glycaemia) and local (e.g. protein turnover) metabolic control, and in the case of exercise; performance, recovery and adaptation. Using the novel technique of contrast‐enhanced ultrasound (CEUS) we explored the effects of feeding on muscle microvascular blood volume (MBV) in 10 young (23 y, BMI; 24 kg·m2) and 12 older (71 y, BMI; 26 kg·m2) men, and in a sub‐group of 4 older men (73 y, BMI; 26 kg·m2) after 20 wk resistance exercise training (RET). Under both postabsorptive and postprandial (IV Glamin 102 mg·kg·h−1 and 20% dextrose; blood glucose at ~7 mmol·l−1) conditions, femoral artery leg blood flow (LBF) was measured over 40 min, before infusion of Definity™perflutren microbubbles to assess regional quadriceps MBV by CEUS. Increases in LBF were evident only in the young men after feeding (0.51±0.02 vs. 0.57±0.04 l.min−1, P<0.05). Similarly, increases in MBV after feeding were observed in young (+25±10%, P<0.05) but not older men (+5.3±6%; P>0.05). Intriguingly, in older men, RET faithfully restored youth‐like increases in MBV after feeding (+31±24%, P<0.05). We conclude that: (i) RET rejuvenates age‐related declines in postprandial microvascular hyperaemia and (ii) RET‐induced improvements in MBV may underlie some of the established metabolic benefits of exercise.

This work was supported the Dunhill Medical Trust.
Mild exercise elicits de novo synthesis of hippocampal dihydrotestosterone and enhanced neurogenesis in adult male rats
Authors: Okamoto Masahiro , Hojo Yasushi , Inoue Koshiro , Matsui Takashi , Lee Min chul , Kawato Suguru , Soya Hideaki
Published: June 2021

We accurately determined the concentrations of hippocampus‐derived androgens and estrogens by employing novel derivatization methods to improve the sensitivity of liquid chromatography with tandem mass spectrometry (LC‐MS/MS), despite that the hormones physiological significance is not well understood. Gonadal hormones modulate adult hippocampal neurogenesis (AHN). Androgens, potent neuroprotective steroids, exert permissive action via their receptors (AR) together with several trophic factors such as BDNF and VEGF. These allow us to postulate that exercise increases AHN through hippocampal androgen. We examined the modulatory roles of hippocampal androgens in exercise‐induced AHN in male rats. We demonstrate that treadmill running significantly increased AHN in both orchidectomized (ORX) and sham‐operated (Sham) rats. However, exercised‐induced AHN was fully inhibited by flutamide (AR antagonist) in ORX rats (Fig. 1A, B). Analysis of mRNA expression for 5a‐redctases, which converts testosterone into dihydrotestosterone (DHT), and for AR revealed that both increased in the hippocampus with exercise (Fig. 1C, D). Further, a mass‐spectrometric analysis showed that hippocampal DHT was significantly increased, even if circulating androgens were depleted (Fig. 1E, F). This suggests that exercise‐induced AHN mediates through the paracrine role of newly synthesized hippocampal DHT through AR.
TNF‐α‐induced loss of IKK‐α in skeletal muscle: implications for skeletal muscle oxidative phenotype
Authors: Remels Alexander , Langen Ramon , Schols Annemie , Gosker Harry
Published: June 2021

Molecular mechanisms underlying lower limb muscle dysfunction in chronic inflammatory disorders, e.g. chronic obstructive pulmonary disease (COPD), are unclear. Here we describe that IKK‐α, a key kinase in the alternative NF‐κB pathway, is a novel regulator of skeletal muscle oxidative phenotype (OXPHEN), which is targeted by inflammation. In C2C12 myotubes, IKK‐α knock‐down decreased myosin heavy chain (MyHC) slow protein and MyHC I and IIA (oxidative) mRNA levels while MyHC IIB (glycolytic) mRNA levels increased. Conversely, IKK‐α overexpression increased MyHC slow protein. Importantly, IKK‐α knock‐down decreased PGC‐1α and PGC‐1β mRNA levels, two key regulators of muscle OXPHEN. We previously demonstrated that TNF‐α potently impairs skeletal muscle OXPHEN (Remels et. al. FASEB J 2010). Interestingly, TNF‐α reduced IKK‐α protein, mRNA and promoter activity and diminished phosphorylation of the down‐stream target of IKK‐α in the alternative NF‐κB pathway (P100) in a classical NF‐κB dependent manner. IKK‐α protein levels in skeletal muscle tissue of COPD patients (n=34) were lower (−50%) compared to healthy controls (n=10). This data shows that IKK‐α controls skeletal muscle OXPHEN and suggests that TNF‐α‐induced loss of IKK‐α protein may contribute to loss of muscle OXPHEN in COPD.

This study was supported by the Netherlands Asthma Foundation (3.2.09.068) and ENIGMA grant QLK6‐CT‐2002‐02285.