Journals
2018 EN
Sahu Soumyadip · Gu Chunfang · Shears Stephen B.
Inositol pyrophosphates (PP‐IPs) are unique, evolutionarily conserved signaling molecules, which act at the interface between signal transduction and cellular metabolism. 5‐IP 7 (5‐diphosphoinositol 1,2,3,4,6‐pentakisphosphate) is the most physiologically abundant PP‐IP (cytoplasmic concentration: 1–2 μM). 5‐IP 7 is a highly electronegative molecule with five monophosphate groups and one bisphosphate group, all packed around a six‐carbon inositol ring. Reported functions of 5‐IP 7 include non‐enzymatic pyrophosphorylation of proteins and non‐covalent binding to the highly electropositive domains in certain proteins (e.g., pleckstrin homology domains). Here, we provide evidence of a new mechanism of action for a PP‐IP, namely, as an enzyme inhibitor. The enzyme in question, encoded by NUDT3 , catalyzes mRNA 5′‐decapping activity. NUDT3 knockdown in MCF7 breast cancer cells cause an increase in the mRNA stability of several ‘migration marker proteins’, namely, β 6 ‐integrin, fibronectin, lipocalin and S100A8, which promote an elevated migratory phenotype compared to the wild type (WT) MCF7 cells. The NUDT3 gene product also hydrolyzes 5‐IP 7 , so we hypothesized that a persistent elevation of cellular levels of that PP‐IP would competitively inhibit mRNA decapping, and hence promote the expression of migration marker proteins. To pursue this idea, we used HCT116 colon cancer cells and HEK293 kidney epithelial cells in which we knocked out the IP 7 Ks that metabolize 5‐IP 7 , thereby increasing levels of this PP‐IP by 2 to 3‐fold. In the current study, qRT‐PCR analysis of IP 7 K −/− HCT116 cells showed an increase in the mRNA levels of β 6 ‐integrin by 2 ± 0.2‐fold, fibronectin by 2.5 ± 0.4‐fold, lipocalin by 5 ± 0.4‐fold, and S100A8 by 3 ± 0.4‐fold compared to WT cells (data are expressed as mean ± SE, n=5, p ≤ 0.01). Knockout of IP 7 K in HEK293 cells resulted in elevation of mRNA levels of β 6 ‐integrin by 6 ± 0.5‐fold, fibronectin by 3 ± 0.6‐fold, lipocalin by 7.5 ± 0.4‐fold, and S100A8 by 3.7 ± 0.35‐fold vs. WT HEK293 cells (data are expressed as mean ± SE, n=5, p ≤ 0.01). We are now developing techniques to load WT cells with 5‐IP 7 to more directly test our idea that this PP‐IP has a novel, enzymatic function as a regulator of expression of pro‐migratory genes that may have relevance to tumor metastasis. Support or Funding Information Intramural Research Program Funding for NIEHS/NIH. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Hearon Christopher M. · Sarma Satyam · Dias Katrin A.
+2 more
Background The amount of time needed to increase oxygen utilization to sufficiently meet metabolic demand (V̇O 2 kinetics) is impaired in heart failure (HF) with reduced ejection fraction, and is an independent predictor of HF mortality. However, it is not known if V̇O 2 kinetics are slowed in heart HF patients with preserved ejection fraction (HFpEF). We tested the hypothesis that V̇O 2 kinetics are slowed during submaximal exercise (equivalent to activities of daily living) in HFpEF patients, and that slower V̇O 2 kinetics are related to impaired peripheral oxygen extraction. Methods Eighteen healthy controls (69±6 years, 10 females) and eighteen HFpEF patients (68±7 years, 10 females) were studied during submaximal (~30% predicted max) and peak treadmill exercise. Cardiac output (Q c , acetylene rebreathing), pulmonary oxygen uptake (V̇O 2 , Douglas bags), and arterial‐venous oxygen difference (a‐v O 2 diff ) were measured during exercise. Breath‐by‐breath V̇O 2 uptake was measured continuously at the onset and throughout submaximal exercise, and V̇O 2 kinetics quantified as the time needed to for V̇O 2 to rise to ~63% of steady state V̇O 2 (mean response time, MRT). Results During submaximal exercise, absolute oxygen demand was similar between groups (V̇O 2 : Control: 0.81±0.1, HFpEF: 0.85±0.2 L/min; P=0.54) however, HFpEF patients had substantially slowed V̇O 2 kinetics (MRT: Control: 40.1±14.2, HFpEF: 65.4±27.7 s; P=0.002), and >50% greater accumulated oxygen deficit compared to controls (Control: 400±173, HFpEF: 693±392 mL; P=0.007). At peak exercise, HFpEF patients had lower relative V̇O 2 (Control: 22.2±4.0, HFpEF: 14.6±3.1 mL/kg/min; P60s demonstrated impaired peripheral oxygen extraction that was apparent during submaximal exercise compared to HFpEF patients with a MRT 60s: 7.9±1.1 mL/dL; P=0.03). Conclusions HFpEF patients have slowed V̇O 2 kinetics during exercise that are due in part to impaired peripheral oxygen utilization. This finding supports growing evidence that peripheral oxidative capacity is a primary contributor to exercise intolerance in HFpEF leading to substantial oxygen deficit even during activities of daily living. Further, MRT can lend important insight into the pathophysiology of exercise intolerance in individual HFpEF patients, and identify patients with severe peripheral limitations to exercise capacity. Support or Funding Information NIH: AG017479, F32‐HL137285 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Sahu Bhavani Shankar · Cero Cheryl · Han Ruijun
+3 more
Obesity results in fat mass accumulation and is associated with serious comorbidities such as type 2 diabetes and cardiovascular disease. Targeting excess fat by lipolysis has been a major challenge due to side effects. TLQP‐21, a neuropeptide encoded by the pro‐peptide VGF (nonacronymic), that binds the complement 3a receptor 1 (C3aR1) on the adipocyte membrane, is emerging as a novel modulator of adipocyte functions and a potential target for obesity. However, the molecular mechanism is still largely uncharacterized. In current study, we aim to understand the role of C3aR1 in murine, human adipocytes and in vivo in diet‐induced obese mice. In vitro experiments using loss and gain of function approaches in 3T3‐L1 adipocytes demonstrate that C3aR1 knockdown prevents TLQP‐21‐induced prolipolytic response. Likewise, TLQP‐21 treatment decreases body weight and fat mass in diet induced obese mice by a mechanism requiring C3aR1 activation. Finally, we demonstrate that this lipolytic pathway is conserved in human adipocytes and can be activated by the rodent but not the human TLQP‐21 peptide, a finding in line with our previous demonstration of a low potency of the human compared to the rodent TLQP‐21 at the human C3aR1 (Cero et al., Structure 2014). In summary, our results identified a novel lipolytic pathway involving TLQP‐21 and C3aR1, a receptor classically studied for its role in innate immunity thus offering a novel target to develop new effective pharmacotherapies for obesity and associated diseases. Support or Funding Information NIH grant (NIH/NIDDK R01DK102496) This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
LIU BING · Barrero Oscar Salgado · Sahu Chinmayi
+3 more
Regulatory T (Treg) cells are a distinct lineage of T lymphocytes that control immunological self‐tolerance and homeostasis. The use of Treg cells for immunotherapy has been challenging, partially because of the functional heterogeneity and instability of Treg cells. The transcription factor FOXP3 is crucial for Treg lineage specification during development and is continuously required for the suppressive function of mature Treg cells. FOXP3 can be regulated by various post‐translational modifications, which enable transient regulation of FOXP3 functionality in response to environmental cues. Here, we investigate the novel O‐linked N‐Acetylglucosamine (O‐GlcNAc) modification on FOXP3 in Treg cell function and stability. Protein O‐GlcNAcylation is enriched in induced Treg cells, compared to naïve T cells. Chemical augmentation of O‐GlcNAcylation promotes the induction of Treg cell signature genes. O‐GlcNAcylation, controlled by O‐GlcNAc cycling enzymes O‐GlcNAc transferase (OGT) and O‐GlcNAcase (OGA), modifies and stabilizes FOXP3 by counteracting with ubiquitination. Treg cell‐specific OGT deletion in mice does not affect Treg cell lineage specification, but results in fewer suppressive effector Treg cells and the development of an aggressive autoimmune syndrome. O‐GlcNAc‐deficient Treg cells show reduced FOXP3 protein expression and increased effector T‐cell signatures. In summary, we demonstrate that protein O‐GlcNAcylation, by modulating FOXP3, is indispensable for Treg cell suppressive function and lineage stability. Manipulating O‐GlcNAc levels may represent a novel approach to translate Treg cell immunotherapy into clinical practice. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Dias Katrin A. · Lawley Justin S. · Hearon Christopher M.
+4 more
Objective Xenon is an inhalational anesthetic with the potential to improve athletic performance and cardiorespiratory fitness via proposed increases in total blood volume. This study aimed to determine the effect of a four‐week xenon supplementation protocol on total blood volume, maximal oxygen uptake (V̇ 2max ) and 3‐kilometre time trial performance in trained individuals. Methods Fourteen subjects (32 ± 12 years; 71% male) were assessed for total blood volume through the carbon monoxide rebreathing method, cardiorespiratory fitness via V̇O 2max on a treadmill test, and athletic performance using a 3 km time trial. Following baseline assessments, subjects were matched for sex, age and V̇O 2max , and were randomised to 12 sessions of gas inhalation over four weeks (2 minutes of inhalation on 3 days per week); 1) xenon, n = 7 (70% xenon, 21% oxygen, 9% nitrogen) or 2) sham gas, n=7 (7% carbon dioxide, 21% oxygen, 62% nitrogen) dosing. Subjects were blinded to group allocation and all outcomes were re‐assessed following the four‐week dosing period. Within‐group mean differences and 95% confidence intervals for outcomes of interest are presented in Table 1. Differences between the xenon and sham gas group over time were tested using two‐way repeated measures ANOVA and are represented by P values. Results Xenon and sham groups were equal at baseline for all outcomes of interest (Table 1). Twelve subjects completed the four‐week dosing intervention and are included in the analyses; one subject withdrew after a single xenon dose due to nausea and one subject was withdrawn after one week of xenon inhalation due to adverse symptoms during and after dosing which resembled sleep paralysis. Blinding was equally effective between groups; subjects in the xenon group were 65% certain of group allocation, while subjects in the sham group were 63% certain of group allocation. There were no significant differences between xenon or sham gas groups for changes in total blood volume, V̇O 2max , or 3 km time trial performance (Table 1). Conclusions Four weeks of xenon inhalation did not result in significant changes in total blood volume, cardiorespiratory fitness or athletic performance in comparison to a sham gas. Given the presence of adverse symptoms in approximately 30% (2 out of 7) subjects and a clear absence of physiological and performance benefits, our findings do not support the use of xenon as a performance enhancing substance. Support or Funding Information This study was supported in part by funding from the Partnership for Clean Competition Research Collaborative. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Sahu Divya · Klemke Richard · Boss Gerry R.
+1 more
Background Despite accounting for around 17,000 deaths and 79,000 new diagnoses annually in the United States, treatment options for bladder cancer remain limited. The key driver of bladder cancer stage progression is increasing invasion into the bladder wall, which has limited response to current chemotherapy options and hence newer therapies directed against invasion are key to improving the treatment of bladder cancer. We have recently shown that arginine, the nitric oxide (NO) precursor, is implicated in bladder cancer progression. Our laboratory has also identified mammalian target of rapamycin complex 2 (mTORC2) as a driver of bladder cancer cell invasion. We evaluated the interaction between these two pathways to promote invasion. Methods Expression of inducible and endothelial nitric oxide synthases (iNOS and eNOS) in FFPE tissue sections of progressive disease was assessed by IHC and correlated with histopathology, progression and stage based on moderate to strong expression (2+, 3+) compared to absent to weak expression (0, 1+). We utilized gene silencing methods and NOS inhibitors and NO scavenger for effects on bladder cancer invasion and migration. We assessed mTORC2 pathway activity and NOS levels in invasive cell tip protrusions called “invadopodia” and evaluated if mTORC2 regulates NOS localization. We used a novel zebrafish model to characterize the effects of mTORC2 and NO on bladder cancer metastases. Result We found that eNOS and iNOS are elevated in invasive human bladder tumors and cell lines and their ablation reduces bladder cancer cell migration and invasion by reorganizing the actin cytoskeleton and reducing invadopodia. mTORC2 silencing can affect levels of iNOS and eNOS in bladder cancer cell lines. Silencing of mTORC2, eNOS or iNOS reduced metastases of bladder cancer cells within zebrafish. Conclusion mTORC2 pathway is a key driver of bladder cancer invasion and metastases by regulating the NO pathway and both mTORC2 and NO can be targets for bladder cancer therapy, which would benefit patient outcomes. Support or Funding Information Prevent Cancer Foundation Fellowship to DS Case Western Reserve University/Cleveland Clinic CTSA Grant Number UL1 RR024989 from the National Center for Research Resources (NCRR), a KL2 career development award (RR024990) and UCSD start‐up funding to DEH This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Samuel T. Jake · Beaudry Rhys · Haykowsky Mark
+2 more
Cycle echocardiography is currently recommended for assessing cardiac involvement (i.e. left ventricular diastolic dysfunction) in patients with unexplained dyspnea on exertion. However, cycle echocardiography is often limited by movement and respiratory artifact, particularly in obese patients with dyspnea. Our group has recently shown that isometric handgrip echocardiography is a powerful diastolic discriminator, capable of unmasking subclinical stress induced diastolic dysfunction. How isometric handgrip echocardiography compares to conventional cycle echocardiography however, remains unclear and was the focus of the present investigation. We hypothesized that isometric handgrip echocardiography would unmask more diastolic dysfunction, given its markedly different hemodynamic load compared to cycle echocardiography. To test this hypothesis, we recruited 24 individuals from the community (9 male, 15 female, age range: 18 – 80), who all performed 3 minutes of isometric handgrip echocardiography followed by 3 minutes of dynamic cycle exercise (20 W). At rest and during the final minute of each exercise protocol heart rate (HR), mean arterial pressure (MAP) and Doppler derived E/e′ were recorded. Consistent with our previous work, and that of others, responders were defined as ΔE/e′ > 1.5 while non‐responders were defined as ΔE/e′ 0.05), while isometric handgrip resulted in a larger increase in MAP (ΔMAP: 28 ± 14 vs . 16 ± 12, handgrip vs . cycle exercise, P = 0.0003). Remarkably, the isolated increase in afterload experienced by the myocardium during isometric handgrip exercise was more sensitive at unmasking sub‐clinical diastolic dysfunction in community dwelling individuals compared to cycle exercise (handgrip: n = 14 vs . n = 10; and cycle: n = 10 vs . n = 14, responders vs . non‐responders, respectively). Taken together, these data highlight the usefulness of isometric handgrip echocardiography at isolating myocardial diastolic relaxation abnormalities in community dwelling individuals, beyond that of dynamic cycle exercise. Future work should focus on confirming the sensitivity of this method in individuals at risk for or with diagnosed heart failure. Support or Funding Information AHA 16SDG27260115 and the Harry S. Moss Heart Trust. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Moralez Gilbert · Hieda Michinari · Sarma Satyam
+4 more
Recent studies have identified that well‐healed burn survivors have altered resting cardiac responses that persist years after the initial burn injury. In healthy non‐burned individuals, prolonged exercise leads to beneficial cardiac remodeling and improved cardiac function. However, it is unknown whether burn survivors likewise demonstrate beneficial cardiac adaptations to long‐term aerobic exercise training. The purpose of this study was to test the hypothesis that six months of aerobic exercise training improves left ventricular systolic function in well‐healed burn survivors. Eleven participants (8 males; aged 43 ± 13 years) with well‐healed burn injuries covering an average total body surface area of 53 ± 25% (range: 22–88%), participated in a progressive 6‐month aerobic‐focused exercise training regimen. An index of cardiac systolic function was assessed via speckle tracking analysis to measure local longitudinal peak systolic strain (Tomtec software) at rest, prior to and after the 6‐month aerobic‐focused exercise training regimen. Exercise training improved resting left ventricular longitudinal systolic strain (Pre, −13.6 ± 2.9% vs. Post, −16.3 ± 2.9%, P=0.013). These data show that despite impaired resting cardiac responses that persist well past complete recovery, cardiovascular adaptations to long‐term aerobic exercise training can be obtained in well‐healed burned subjects. Support or Funding Information Work funded by NIH GM068865 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Salem Gamal A · Shaban Ahmed · Diab Hussain
+4 more
Objectives To determine the effects of leaf extract of Phoenix dactylifera against the oxidative stress response induced by a wide‐spectrum analgesic paracetamol in rats. Methods Adult Sprague dawley rats were treated with or without paracetamol (1500mg/kg) and leaf extract of Phoenix dactylifera (300mg/kg). Serum samples were collected to evaluate liver function tests, and hepatic tissue samples were obtained to assess oxidative stress‐related markers and histopathological examination. Results Phoenix dactylifera ameliorated paracetamol‐induced elevated serum liver markers such as alkaline phosphatase (ALP), aminotransferases (AST), alanine aminotransferase (ALT) and bilirubin. In addition, Phoenix dactylifera restored paracetamol‐induced reduced levels of liver tissue antioxidant enzymes such as glutathione peroxidase (GPx), catalase (CAT), and superoxide dismutase (SOD). Histological analysis demonstrate that Phoenix dactylifera conserved the hepatic tissue against lesions such as inflammation, centrilobular necrosis, and hemorrhages induced by PCM. Conclusion These findings indicate that Phoenix dactylifera possesses protective effects against paracetamol‐induced liver injury and oxidative stress, and could be used to explore against drug induced deleterious effects in experimental models. Support or Funding Information No funding support This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology
Journals
2018 EN
Khalifa Hesham Ahmed Mohammed Ismail · Abdelrahman Ahmed Shaban Abdelaziz · Sahu Ravi P
Objectives To determine the effects of aqueous extract of Alhagi maurorum against the adverse effects including hepato‐nephrotoxicity induced by a broad‐spectrum antibiotic norfloxacin in rats. Methods Adult male albino rats were treated with or without norfloxacin (10 mg/kg) and Alhagi maurorum (300 mg/kg). Serum samples were collected to evaluate liver and kidney function tests, and hepatic and renal tissue samples were obtained to assess antioxidant activity and histopathological examination. Results Alhagi maurorum ameliorated norfloxacin induced elevated levels of serum alkaline phosphatase (ALP), aminotransferases (AST), urea, creatinine, and uric acid. In addition, Alhagi maurorum significantly reduced the tissue levels of malondialdehyde (MDA), and increased glutathione peroxidase (GPx) and superoxide dismutase (SOD) compared to norfloxacin alone treated group. Histological analysis demonstrate that Alhagi maurorum conserved the hepatic and renal tissue damages induced by norfloxacin. Conclusion These findings indicate that Alhagi maurorum aqueous extract possesses a potent antioxidant activity and could be used to attenuate norfloxacin induced oxidative stress and hepatorenal damage in experimental models. Support or Funding Information No funding support This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
The Federation of American Societies for Experimental Biology