bluuebunny/crossref_metadata_2025 huggingface dataset:
Social isolation during perinatal development alters the behavioral response to cocaine in juvenile rats
Authors: Lugo‐Escobar Natasha , Carreras Nicole , Segarra Annabell C.
Published: June 2021

Stress is a risk factor in drug addiction, enhancing development and increasing relapse. However its role during development has received little attention. This study will investigate if social isolation during early development alters the behavioral response to cocaine. Pregnant Sprague Dawley rats were housed 1 (prenatal isolation group) or 2 (control group) rats per cage. From postnatal day 2‐9, pups from the neonatal isolation group were isolated for 60 min in a separated cage. At day 21 (weaning), pups from the postnatal isolation group were housed individually. Another group was isolated during both prenatal and postnatal period. Control rats were maintained with dam and siblings and at weaning, housed 4‐5/cage of the same sex. At day 22, and for 5 consecutive days, pups were injected daily with saline or cocaine (15 mg/kg/i.p.) followed by 7 drug free days. A challenge injection of saline or cocaine (15 mg/kg,i.p.) was given on day 13. Locomotor activity was recorded on days 1, 5, and 13. Only rats that were isolated during both prenatal and postnatal period showed an increase in cocaine‐induced behavioral sensitization as compared to control rats. These data indicate that isolation stress during perinatal development enhances the behavioral response to drugs of adolescent rats. [Support contributed by NIH grants: U54NS39405 (SNRP); SO6‐GM08224 (MBRS/SCORE)]
Insulin‐like growth factor I downregulates proatherogenic 12/15‐lipoxygenase in vivo and in vitro
Authors: Vaughn Charlotte , Sukhanov Sergiy , Higashi Yusuke , Shai Shaw‐Yung , Delafontaine Patrick
Published: June 2021

We have shown that 12/15‐lipoxygenase (LOX) mediates important proatherogenic oxidized low density lipoprotein (OxLDL)‐induced effects in smooth muscle cells (SMC), namely, apoptosis and downregulation of the insulin‐like growth factor (IGF‐1) receptor, and that infusion of IGF‐1 into Apolipoprotein E‐null (ApoE‐/‐) mice suppresses macrophage (MF) plaque infiltration and atherosclerosis. To determine molecular mechanisms mediating IGF‐1's anti‐atherogenic effects ApoE‐\‐ mice were injected with IGF‐1 (0.7 mg/kg/day, 7d). IGF‐1 decreased LOX aortic gene expression by 64±6% (RT‐PCR). IGF‐1 dose‐dependently decreased LOX gene expression in THP‐1 derived MF, while no significant effect was seen in SMC. To examine IGF‐1's effects on LOX activity, SMC or MF were pre‐treated with IGF‐1 (10 or 50 ng/ml, 24h), and then exposed to OxLDL or control. 14,15‐leukotriene (a product of LOX‐specific enzymatic reaction) levels were measured in conditioned media by ELISA. OxLDL induced a five‐fold increase in LOX activity levels in MF, and a smaller increase in LOX activity in SMC. IGF‐1 partially blocked OxLDL‐induced LOX upregulation by 27±4%. In summary, IGF‐1 suppresses vascular LOX expression in ApoE‐\‐ mice and induces LOX downregulation in MΦ and SMC. Our data suggests that the ability of IGF‐1 to downregulate LOX plays an important role in its anti‐atherogenic effect.

Study supported by NIH HL 70241
Specific changes in TRPV4 expression in the vasopressinergic cells in supraoptic nucleus of the rat brain following bile duct ligation (BDL) induced cirrhosis
Authors: Carreño Flávia Regina , Ji Lisa , Cunningham J Thomas
Published: June 2021

Inappropriate vasopressin (AVP) results in hyponatremia, a risk factor in pathophysiological states such as cirrhosis. This study tested the hypothesis that changes in the expression of TRPV4, a transient receptor potential channel that has been shown to be activated by osmotic stimulation, in vasopressin neurons contributes to inappropriate vasopressin release in BDL rats. Four weeks after surgery, BDL rats demonstrated significantly increased plasma vasopressin and plasma renin activity (PRA), hypervolemia, and decreased plasma osmolality compared to sham ligated controls. Western Blot analysis revealed that TRPV4 expression was significantly increased in brain punches from BDL rats containing the supraoptic nucleus (SON) (100% ± 11 to 157% ± 4.8), but not the paraventricular nucleus (PVN) of the hypothalamus. Immunohistochemistry analysis demonstrated that TRPV4 is co‐localized in AVP neurons in both PVN and SON. The percentage of AVP neurons expressing TRPV4 was significantly increased (36 % ± 3 to 55.3% ± 4.3) only in the SON of BDL rats. It could be that increased expression of TRPV4 in AVP neurons in the SON resets their osmosensitivity allowing them to continue to release AVP in the face of hypoosmolality and hyponatremia contributing to the pathogenesis of inappropriate AVP release. NIH R01 HL55692
Influence of experimental early diabetic nephropathy on the functional subtypes of renal α1‐adrenoceptor in spontaneously hypertensive rats (SHR)
Authors: Khan Md Abdul H , Sattar Munavvar A , Abdullah Nor A , Majid Dewan S A , Johns Edward J
Published: June 2021

This study investigated the impact of early diabetic nephropathy (EDN) on the functional population of renal α1‐adrenoceptors in SHR rats. One week after streptozotocin, EDN was confirmed by hyperglycemia (14.8±2.3 vs 5.7 ±0.6 mg/dL), raised creatinine clearance (4.2±0.10 vs 0.5±0.07 mL/min/100g), albumin‐creatinine ratio (94.6±3.7 vs 39.4±2.6 µg albumin/mg creatinine) and fractional excretion of Na+ (1.2±0.01 vs 0.4±0.01 %) compared to non‐diabetic (NDN) rats. In acute studies, renal vasoconstrictions caused by renal nerve stimulation and intrarenal noradrenaline, phenylephrine and methoxamine were determined in the presence and absence of intrarenal amlodipine (AMP), 5‐methylurapidil (MEU), chloroethylclonidine (CEC) and BMY 7378. In both NDN and EDN rats, the renal vasoconstrictions were attenuated by all antagonists (AMP‐ NDN, 5‐15% and EDN, 8‐20%; MEU‐ NDN, 5‐14% and EDN, 9‐14%; BMY 7378‐ NDN, 5‐10% and EDN, 5‐8%). CEC enhanced these responses in EDN (5‐10%) but did not cause any effect in NDN SHR rats. The data showed that α1A and α1D are the renal functional α1‐adrenoceptors in SHR rat irrespective of the presence of EDN. A minor contribution of pre‐synaptic α1B‐adrenoceptors was observed in the EDN group.
Resuscitation strategy alters response to hemorrhage
Authors: Makley Amy Teres , Friend Lou Ann , Goodman Michael , Blanchard John , Lentsch Alex , Pritts Timothy
Published: June 2021

Objective

Hemorrhagic shock is associated with a systemic inflammatory response. Two common resuscitation strategies are crystalloid and blood component therapy. The effect of these strategies on inflammation is not fully understood. We hypothesized that crystalloid would exacerbate inflammation after hemorrhagic shock.

Methods

Mice underwent pressure controlled hemorrhage via femoral artery cannulation and resuscitation with whole blood (WB) or Ringers lactate (LR). Survival and fluid requirements were determined. Tissue myeloperoxidase (MPO) levels were analyzed.

Results

Resuscitation with LR resulted in less survival at 7 days vs WB or sham (Fig A). LR mice required more fluid (Fig B) and exhibited higher MPO activity in jejunum, but not liver or kidney (Fig C).

Conclusions

LR resuscitation led to increased fluid requirements, jejunal MPO and mortality compared to WB. These results are critical to understanding potential benefits of blood resuscitation following trauma.

Supported by a grant from the Henry Jackson Foundation
Differential Aquaporin Expression in Hepatic Stellate Cells During Transdifferentiation
Authors: Lakner Ashley , Gulledge Alyssa , McKillop Iain , Schrum Laura
Published: June 2021

Persistent activation and increased cell survival of the hepatic stellate cell (HSC) leads to the development and progression of liver fibrosis. Induction of HSC apoptosis and decreased proliferation can attenuate fibrosis. Aquaporins (AQP) play a critical role in both the initiation of apoptosis and proliferation. The expression profile and function of AQPs have not been determined in the daily progression of HSC activation. AQP 0, 1, 5, 8, 9, 11 and 12 mRNAs were detected in freshly isolated (dQ) cells, while activated (d14) cells exhibited a marked decrease in expression as demonstrated by semiquantitative and RealTime PCR. However, AQP mRNA expression profiles showed transient increases during culture activation (d1‐d10). Western blot analyses showed decreased AQP 8 and 9 protein expression in d14 versus dQ cells. Osmotic challenge significantly increased cell swelling in dQ cells which was abolished by pre‐treatment with HgCl2, an AQP channel blocker, while d14 cells did not swell. D1 and d14 HSCs exhibited decreased cellular viability after one hour exposure to the apoptotic agent gliotoxin, whereas dQ cells responded within 20 minutes. HgCl2 pretreatment abrogated these effects. These data suggest that HSC survival may be modulated by AQP expression which could be a potential therapeutic target. Supported by NIH AA14891.
PPAR‐alpha agonist increases skeletal muscle carnitine palmitoyltransferase‐1 activity and fatty acid oxidation in obese insulin‐resistant monkeys
Authors: Motz Kevin Michael , Hansen Barbara Caleen , Ortmeyer Heidi Karen
Published: June 2021

The selective PPAR‐alpha agonist K‐111 (Kowa Co. Ltd., Tokyo, Japan) decreases plasma triglycerides and improves insulin sensitivity in obese, insulin‐resistant rhesus monkeys. The purpose of this study was to determine whether K‐111 improves skeletal muscle fatty acid oxidation and enzymes involved in fatty acid metabolism. Five obese, insulin‐resistant adult monkeys were studied at baseline and again after treatment with K‐111 at 3 mg/kg μday for 7 weeks. Acyl‐CoA synthase (ACS), carnitine palmitoyltransferase‐1 (CPT‐1), β‐hydroxy‐acyl CoA dehydrogenase ( β‐HAD), citrate synthase (CS) and total fatty acid oxidation were measured in skeletal muscle homogenates at baseline and following K‐111. CPT‐1 activity significantly increased (0.012 ± 0.004 vs. 0.019 ± 0.004 pmol/min μ;mg protein, p<0.05) and total fatty acid oxidation tended to increase (2.1 ± 0.6 vs. 4.0 ± 1.1 pmol/min μmg protein, p<0.07) following treatment. ACS activity (128 ± 25 vs. 180 ± 11 pmol/min μmg protein), β‐HAD activity (0.182 ± 0.05 vs. 0.190 ± 0.03) and CS activity (0.108 ± 0.02 vs. 0.116 ± 0.01 µmol/min μmg protein) were not significantly affected by treatment. The change in total fatty acid oxidation was associated with the change in ß‐HAD and CS activity (p<0.01) but not to CPT‐1 activity. These results suggest that the mechanism by which PPAR‐alpha agonist treatment improves fatty acid oxidation is through changes in ß‐HAD and CS activity.
The anabolic effect of insulin is dependent on its ability to increase blood flow and muscle perfusion in human subjects
Authors: Timmerman Kyle L , Lee Jessica L , Dhanani Shaheen , Dreyer Hans C , Glynn Erin L , Fry Chris S , Drummond Micah J , Rasmussen Blake B , Volpi Elena
Published: June 2021

Insulin‐induced blood flow has been shown to correlate positively with insulin's ability to enhance muscle protein synthesis. Thus, we hypothesized that pharmacological inhibition of insulin‐induced vasodilation would reduce the anabolic response of muscle proteins to insulin.

Skeletal muscle fractional synthetic rate (FSR), net balance, blood flow (indocyanine green), and muscle perfusion (contrast enhanced ultrasound) were measured in 10 healthy, glucose tolerant subjects (31±2 yrs) in the basal state and during administration of insulin alone (N= 5) or insulin plus the NO synthase inhibitor NG‐monomethyl‐L‐arginine (L‐NMMA) (N= 5).

There were no differences in the basal period. FSR increased significantly (P<0.05) in subjects receiving insulin alone (basal: 0.059%/h vs. insulin: 0.079%/h) compared to subjects receiving insulin plus L‐NMMA (basal: 0.056%/h vs. insulin+ L‐NMMA: 0.045%/h). Those receiving insulin alone experienced a significantly (P<0.05) greater percent change from baseline (54.90%) in muscle perfusion than subjects receiving insulin + L‐NMMA (3.14%). There was also a trend (P=0.09) for increased blood flow in subjects receiving insulin alone.

These preliminary data suggest that the ability of insulin to stimulate blood flow and muscle perfusion is necessary for insulin to stimulate muscle protein synthesis in healthy, young adults.

Funded by NIH R01 AG018311 and UTMB Claude D. Pepper Older Americans Independence Center Grant #P30 AG024832.
Neuroprotective effects of antifreeze protein against hypothermia induced cell death in rat hippocampal slices
Authors: Rubinsky Liel , Zhang Jingli , Bickler Philip E
Published: June 2021

Antifreeze proteins (AFP) are a diverse family of molecules whose unusual properties include non‐colligative depression of the freezing temperature of water, and a yet unclear interaction with the cell membrane and function. To assess the neuroprotective effect of AFP type I / III we subjected rat hippocampal slices to deep hypothermic insult and measured cell survival with propidium iodide (PI) exclusion. Ionic currents (potassium channels, IH) were also measured with patch‐clamp electrodes. Our PI results demonstrate a significant protective effect of these proteins on neurons at deep hypothermic temperatures. While patch‐clamp recordings at 10°C, 18°C and 37°C in CA1 neurons in freshly made hippocampal slices revealed hyper‐polarization, consistent with enhanced potassium channel activity. These results suggest that AFP protected hippocampal neurons by influencing ionic flux across the cell membrane. The study provides new insight into the protective mechanism of action of these proteins and their function on neurons. This work was supported by the NIH GM52212.
Purinergic receptors in mouse and rat epididymis : Role of luminal ATP and adenosine in V‐ATPase activation
Authors: Belleannee Clemence , Da Silva Nicolas , Shum Winnie , Breton Sylvie
Published: June 2021

In the epididymis, V‐ATPase is expressed in clear cells and acidifies the lumen where sperm mature and are stored. We showed that luminal alkaline pH triggers the apical membrane accumulation of V‐ATPase and that this process is calcium‐dependent. However, the link between alkaline pH and intracellular calcium is not completely elucidated. Some purinergic receptors are sensitive to pH, and we examined their expression by RT‐PCR in rat epithelial cells isolated by laser capture microdissection. mRNA transcripts for P1 adenosine receptors (subtypes A1, A2B and A3) and P2 ATP receptors (subtypes P2X1, P2X2, P2X4, P2X11, P2Y1 and P2Y2) were detected. Ecto‐ATPase and alkaline phosphatase were also observed, indicating potential degradation of ATP into adenosine in the epididymal lumen. In vivo perfusion of rat and mouse epididymis showed that both ATP (600µM) and adenosine (300µM) induced apical membrane V‐ATPase recruitment even in the absence of an alkaline pH stimulus. BAPTA‐AM abolished the ATP‐mediated effect, indicating the participation of calcium in this response. These data show that ATP and adenosine are sufficient to cause the accumulation of V‐ATPase in the apical membrane of clear cells. The paracrine activation of purinergic receptors might, therefore, play a significant role in the regulation of proton secretion in the male reproductive tract. This work is supported by NIH grant HD40793.
In silico study of lactate metabolism in brain during visual stimulation
Authors: Occhipinti Rossana , Somersalo Erkki , Calvetti Daniela
Published: June 2021

Due to the lack of direct in vivo measurements of the partitioning of lactate and glucose between astrocytes and neurons during activation, interpretation of the available in vivo experimental data is all but straightforward. This has led to the formulation of different hypotheses on the metabolic role of astrocytes and neurons. The interpretation of lactate dynamics in the context of compartmentalized metabolism requires the formulation of mathematical models. In this work we employ a new dynamic computational model of the astrocyte‐glutamatergic neuron cellular complex, previously used and validated for steady state investigations (Occhipinti et al. 2008), which uses Michaelis‐Menten type kinetic expressions for the reactions and transport rates. The distributions of the kinetic parameters which identify the model are estimated by employing a new methodology, based on Bayesian statistics. The model predictions, trained on data collected by NMR spectroscopy in human visual cortex (Mangia et al. 2007), are in agreement with the suggestion already proposed by Gjedde et al. 2001, that neurons would be responsible for the observed increase in extracellular lactate concentrations during stimulation. The ensemble of model predictions is further investigated to identify important casual relations which may shade some light on the mystery of lactate metabolism in brain.
Hyperoxia attenuates the elevation in muscle sympathetic nerve activity following exposure to hypoxia
Authors: Querido Jordan Scott , Kennedy Paul M , Sheel A William
Published: June 2021

Hypoxia may sensitize the carotid chemoreceptors resulting in a sustained elevation of muscle sympathetic nerve activity (MSNA) which outlasts the hypoxic stimulus. To test this hypothesis we determined (n=4) the effect of carotid body deactivation on the sustained elevation in MSNA (common peroneal nerve) following isocapnic hypoxia (arterial O2 saturation 80% for 20 mins) in humans. Ventilation, heart rate and MSNA were elevated in hypoxia; however, only the elevation in MSNA outlasted the hypoxic stimulus, and remained elevated for 20 mins following the hypoxic stimulus. In contrast, carotid body deactivation with hyperoxia (100% O2 for 1 min) transiently reduced MSNA in eight of nine trials (mean decrease 12.6 ± 9.5%, p = 0.005). Our results suggest that input from the carotid chemoreceptors is obligatory for the sustained elevation in MSNA initiated by chemoreflex stimulation. We attribute the decrease in MSNA to a transient hyperoxia‐induced attenuation in carotid chemoreceptor sensitivity.

Research support: NSERC, MSFHR
L2DTL, a cell cycle regulated gene is essential to tumor cell growth
Authors: Pan Hung‐Wei , Chen Mei‐Ling , Yuan Ray‐Hwang
Published: June 2021

The human homologue of Drosophila lethal (2) denticleless protein (L2DTL/DTL) or S. pombe Cdt2. CDT2, a regulator of DNA replication. Our previously results shown L2DTL is a nuclear protein. Its expression is regulated during cell cycle progression, increased during the G1 to S phase and decreased dramatically in mitosis. L2DTL mRNA level was increased in DNA synthesis period while liver regeneration, and was frequently over‐expressed in most common cancers specimens (from 25% to 70%). The expression level of L2DTL mRNA was also expressed in the all examined cell lines (hepatoma cell lines, MCF‐7, HeLa, 293 and 293T cells). Introduce the EGFP‐L2DTL into the NIH3T3 cells can increase almost 15% the S‐phase fraction. Up‐regulation of L2DTL at HeLa cells cause the bigger colonies, and down‐regulation of L2DTL by shRNA reduced HeLa cell colonies formation in vitro soft‐agar assay, moreover, interrupted L2DTL expression also depressed the tumor formation in NOD/SCID mice. While up‐ or down‐regulated the L2DTL protein in the cells, we find the PCNA level have same regulation. These results indicate that L2DTL are often overexpressed in tumor cells, and play important roles in tumor cell proliferation.
Oxidant Regulated Intersubunit Disulfide Bond Formation between ASIC1a Subunits
Authors: Zha Xiangming , Wang Runping , Collier Dan M , Snyder Peter M , Wemmie John A , Welsh Michael J
Published: June 2021

The acid‐sensing ion channel‐1a (ASIC1a) is composed of three subunits and is activated by a fall in extracellular pH. It plays an important role in diseases associated with a reduced pH and production of oxidants. Previous work showed that oxidants reduce ASIC1a currents. However, the effects on channel structure and composition are unknown. We found that ASIC1a formed intersubunit disulfide bonds and the oxidant H2O2 increased this link between subunits. Cys495 in the ASIC1a C‐terminus was particularly important for intersubunit disulfide bond formation, although other C‐terminal cysteines contributed. Intersubunit disulfide bonds produced some ASIC1a complexes larger than trimers. Intersubunit disulfide bond formation also reduced the proportion of ASIC1a located on the cell surface and contributed to the H2O2‐induced decrease in H+‐gated current. These results indicate that channel function is controlled by disulfide bond formation between intracellular residues on distinct ASIC1a subunits. They also suggest a mechanism by which the redox state can dynamically regulate membrane protein activity by forming intracellular bridges.
Kinin B2 Receptor‐Mediated Bradykinin Internalization and Metalloendopeptidase EP24.15‐Dependent Intracellular Bradykinin Degradation
Authors: Sandén Caroline , Enquist Johan , Bengtson Sara , Herwald Heiko , Leeb‐Lundberg Fredrik
Published: June 2021

Kinins are potent proinflammatory peptides that are produced extracellularly and rapidly degraded by extracellular and intracellular peptidases. In this study, we developed model cell systems expressing the kinin B2 receptor (B2R) and the metalloendopeptidase thimet oligopeptidase (EC 3.4.24.15; EP24.15) either individually or together to address 1) the cellular and functional relationship between these proteins and 2) the participation of EP24.15 in the metabolism of bradykinin (BK) after BK internalization via B2R. No direct colocalization of B2R and EP24.15 was observed using immunofluorescence microscopy. However, the two proteins coimmunoprecipitated specifically, and EP24.15 attenuated maximal B2R responsiveness without influencing the potency of BK to stimulate phosphoinositide hydrolysis and intracellular calcium mobilization. Cell surface‐bound BK remained intact in cells overexpressing EP24.15 but was degraded intracellularly in an EP24.15‐dependent manner upon B2R‐mediated endocytosis. These results show that EP24.15 acts to negatively regulate B2R responsiveness and as an intracellular peptidase in the degradation of BK specifically internalized via this receptor.
Identification Of Cysteine Residues In ALDH2 Responsible For Its Bioactivation Of Nitroglycerin
Authors: Tsou Pei‐Suen , Page Nathaniel A. , Qu Jun , Fung Ho‐Leung
Published: June 2021

The dehydrogenase, esterase, and nitrate‐bioactivating activities of mitochondrial aldehyde dehydrogenase ALDH2 were significantly inactivated by nitroglycerin (NTG). Tandem mass spectrometry sequencing of a trypsin or glu‐c‐digested NTG+ALDH2 mixture, after reduction by tris(2‐carboxyethyl)phosphine, indicated that oxidation occurred at C66 (to ‐SOH) and C386 (to ‐SOH, ‐SO2H, ‐SO3H). Modification at C319 was not detected, probably due to the low ionization efficiency of the C319‐contained tryptic or Glu‐c domains. Mutants of C66A, C386A, and C319A (the active site for dehydrogenase and esterase activities) were constructed. The specific ALDH activities (SA) of the WT, C66A, C386A and C319A were 2.51, 2.25, 3.56, 0 U/mg protein, respectively. Within 2 min of incubation, NTG was completely converted into 1,2‐glyceryl dinitrate (GDN) by WT, C66A, and C386A, but not by C319A.. None of these enzymes produced 1,3‐GDN. Consistent with these results, ALDH2 WT, C66A, and C386A catalyzed NO production from NTG with similar activities (cumulative NO production in μM*sec/SA: WT: 60.1 ± 15, C66A: 57.5 ± 24, C386A: 72.2 ± 32), while C319A did not liberate any detectable NO. These results suggest that the dehydrogenase, esterase, and NTG‐bioactivating activities of ALDH2 share the same active C319 site. (Supported in part by NIH HL81580).
Rosiglitazone Induces Cardiac Protein Kinase C beta2 Over‐expression And Increases Ventricular Mass in Type 2 Diabetic db/db Mice
Authors: Xia Zhengyuan , Carroll Rogayah , Wang Fang , Severson David L
Published: June 2021

Rosiglitazone (RSG) treatment in type 2 diabetes has been associated with increased incidence and worsening of heart failure, but the mechanism is unknown. It has been shown that protein kinase C (PKC) β2 is over‐expressed in failing human hearts (Circulation 1999;99:384‐91) and that RSG‐induced weight gain is attributable to increased activation of PKCβ in adipose tissue (FASEB J 2006;20:1203‐5). Further, left ventricular mass is reported to be a predictor of heart failure (Eur Heart J 2008;29:741‐747). We, therefore, hypothesized that RSG may induce PKCβ2 over‐expression and increase ventricular mass of the hearts from db/db (D) mice, a mongenic model of obesity and features of type 2 diabetes. Diabetic mice were either untreated (D), treated with RSG (D+RSG) at 26 mg/kg/day or with RSG plus N‐acetylcysteine (NAC, 1.4 g/kg/day) (D+RN), for 3 weeks; NAC can inhibit PKCβ via mechanisms independent of its antioxidant property. Both RSG or RSG+NAC reduced body weight (BW) relative to D group. However, RSG increased ventricular mass and heart weight/BW ratio, accompanied by 1.5‐fold increase of myocardial PKCβ2 protein expression and activation as well as increased protein kinase B/AKT phosphorylation at serine‐473 relative to D (all P<0.05). NAC prevented all these changes. RGS may have increased ventricular mass by inducing PKCβ2 over‐expression.

Supported by a CIHR grant (DLS) and an AHFMR postdoctoral fellowship (ZX)
Identification of two naturally occurring AMPKα2 variants exclusively expressed in the brain of amyotrophic lateral sclerosis
Authors: Wang Tingting , Cheng Huaiyan , Qiu Rongde , Wang Liqin , Feng Yinghong
Published: June 2021

5′‐AMP‐activated protein kinase (AMPK), a heterotrimeric complex consisting of catalyticα, scaffolding ?, and AMP‐binding ? subunits, is a highly conserved master fuel gauge that meets cell demands by monitoring and regulating cellular energy status. The objective of this study is to identify novel AMPKα2 variants and establish links to diseases. Methods of gene cloning, protein‐protein interaction assay, cell proliferation assay, gene expression, Western blot, and RT‐PCR detection of gene transcripts in human brain total RNA were employed. The result reveals two naturally occurring splicing variants of AMPKα2 that are exclusively expressed in the brains of all 5 cases of amyotrophic lateral sclerosis (ALS), but not in the brains and other tissues of 30 normal or non‐ALS human subjects. Gene analysis suggests that the variants result from differential skipping of two constitutive exons. Overexpression of the variants in COS‐1 cells and SH‐SY5Y neuronal cells induced cell death. Moreover, AICAR, an AMPK activator, and the full‐length AMPKα2 inhibited the variant‐ and glutamate‐induced neuro‐excitotoxicity, respectively. Mechanistic studies showed that the AMPKα2b variant binds AMPKβ subunits in a dominant negative fashion. The conclusion is that two ALS‐specific novel AMPKα2 variants have been uncovered.

Supported by NIH grant (HL065492) to YHF.
Effect of controlling for background diet in the interpretation of dietary fiber and plasma lipids relationship
Authors: Yu Winifred Wingyun , Lau Joseph , Trikalinos Thomas , Schmid Christopher H , Lichtenstein Alice H
Published: June 2021

Background

Participants in nutrition studies often consume background diets that include nutrients under investigation. Valid estimates of nutritional interventions therefore require proper control for backgrounds diet in study design.

Method

We assessed the effect of background diet in a systematic review of 34 trials studying the association of fiber and plasma lipids (total cholesterol [TC], high density lipoprotein cholesterol [HDL], low density lipoprotein cholesterol, and triglycerides [TG]). Background diet was defined to be adequately controlled when there were no reported statistically significant differences in energy and macronutrient intakes between experimental and control groups, or when results were adjusted for these variables. For each lipid outcome, the proportion of studies with significant findings was compared between studies with and without adequate diet control.

Result

Background diet control was adequate in 12 studies. The proportion of studies with significant findings appeared to be higher among studies with, than without, adequate diet control for TC (4/9 vs. 2/17), HDL (2/7 vs. 1/14) and TG (1/8 vs. 1/17) concentrations. These differences were not significant. More studies are needed to confirm these findings.

Source of research support: NHLBI T32 HL69772
Lack of effect of omega‐3 fatty acid (PUFA) dietary supplement on clinical measures of periodontitis in humans
Authors: Parulkar Madhura , Dawson Dolphus R. , Kryscio Richard , Novak Michael J. , Ebersole Jeffrey L. , Boissonneault Gilbert A.
Published: June 2021

Previous research in experimental animals suggests that PUFA may improve resolution of periodontitis by inhibiting inflammation and modulating alveolar bone loss, but human data are limited. In the present study 65 patients with chronic periodontitis (mean age 43.9 ± 10.9, 37.8% female and 71.9% Caucasian) were randomized to receive 3 g of PUFA or placebo (PL). In addition, all patients received oral health instruction at weeks 0 and 16. Clinical periodontal measures (pocket depth, bleeding on probing, attachment loss and plaque index) and blood samples for serum and red blood cell fatty acids (FA), were collected at weeks 0, 8, 16 and 28 with data presented for weeks 0,8 and 16. PUFA subjects had significantly greater amounts of DHA and total PUFA in serum at week 8 and 16 when compared to week 0 or to PL at the corresponding time. Similar changes were seen with EPA, DHA and total PUFA in RBC membranes. Despite significant effects of PUFA supplementation on serum and RBC FA content, clinical measures were not significantly associated with lipid changes. These results suggest that PUFA administration may not enhance oral health outcomes in humans with periodontitis, or their effect is dwarfed by the impact of oral hygiene. This research was supported by NIH‐NCRR grant P20RR020145 and GCRC grant #M01RR02602.
Bloomed dark chocolate still contains the biological function in lowering serum lipids and C‐reactive protein inflammation marker
Authors: Hong Mee Young , Nemoseck Tricia , Marsh Tanya , Chapin Andrea , Castaneda Debra
Published: June 2021

Chocolate blooming occurs during melting due to exposure to high temperatures (i.e. hot weather or cooking) and subsequent cooling. Although bloomed chocolate is known to be safe for human consumption, it is not known if the biological function of chocolate, particularly related to its lipid‐lowering property, remains unchanged. In the present study, both regular and bloomed dark chocolate groups decreased the levels of serum triglycerides, total cholesterol and LDL‐cholesterol compared to control (no chocolate) (P<0.05). Moreover, chocolate‐fed animals, regardless if bloomed or not, C‐reactive protein (CRP) was lower than control (P<0.01). Total antioxidant capacity was determined by Oxygen Radical Absorbance Capacity (ORACTotal) and was slightly lower in bloomed chocolate (422 μmoleTE/g) compared to regular chocolate (546 μmoleTE/g) but bloomed chocolate still contained most of the original ORAC value. These results suggest that heat treated thus bloomed chocolate still contains the beneficial properties of chocolate in regard to lipid profiles and CRP values. Chocolates, bloomed or not, may be beneficial for improvement of the risk factors of cardiovascular disease. Additional research is warranted to examine the long‐term effect of chocolates and the mechanisms underlying their lipid‐lowering effects. Supported by CHNR08‐810 and NUTR 302L class.
Reduced levels of oxidative stress in methionine restricted F344 rats
Authors: Nichenametla Sailendra Nath , Gonsar Kristin , Calcagnotto Ana , Richie John P
Published: June 2021

Dietary methionine restriction (MR) extends life span by 40‐50% in both rats and mice. Previous studies suggest that the anti‐aging properties of MR may be mediated, in part, by reduced oxidative stress. Our current objective was to determine the effect of MR on biomarkers of oxidative stress and endogenous antioxidants and related enzymes in the rat liver. Male F344 rats (6‐week old) were fed diets containing either 0.87% (control) or 0.17% (MR) methionine for up to 6 weeks. Five animals per group were sacrificed weekly and blood and liver were processed for analysis of glutathione (GSH), antioxidant enzymes and biomarkers of oxidative stress. In blood, levels of 8‐OHdG were 31% lower, 8‐isoporstane were 39% lower, and the levels of protein bound GSH were 24% lower in MR rats compared to controls indicating a reduction in levels of oxidative stress (p < 0.05). MR was also associated with 3‐ to 5‐fold reductions in glutathione peroxidase activity and a 2‐fold reduction in GSH levels in liver, where as, blood GSH levels were increased > 2‐fold (p < 0.01). Activity levels of glutathione reductase and superoxide dismutase were unchanged by MR. These results indicate that reductions in the levels of oxidative stress by MR are not mediated by increases in antioxidants and antioxidant related enzymes in liver but rather by decreased ROS production.
Subclassification of Leukotriene‐Based Asthmatics
Authors: Broughton Kenneth Shane , Ross Erica , Johnson Kurt
Published: June 2021

Previously we identified the idea of four subtypes of asthmatics, three of which were cytokine‐based asthmatics and another larger leukotriene‐based family. Previous studies identified that the leukotriene specific subtype is n‐3 PUFA responsive. It was our intent to determine if a leukotriene‐specific family of asthmatics could be further subclassified into families of asthmatics or not.

Twenty‐two asthmatics went through a methacholine bronchoprovocation challenge on two occasions separated by one month. Saliva for one half hour, a blood sample four hours post‐bronchoprovocation, and 24 urine samples were collected. Saliva and urine were analyzed by reverse‐phase high‐pressure liquid chromatography (RP‐HPLC) coupled with a diode array‐UV detector at 280 nm and a flow rate of 1.0 mL/min for analysis of cysteinyl leukotrienes on a Schimadzu HPLC.

Ten individuals were identified as being non‐LT based asthmatics while twelve were identified as being LT‐based asthmatics. Based on saliva LT production, three individuals were identified as being overlapping LT asthmatics: a C4/D4, a B4/C4/D4, and a B4/C4 based asthmatic. However, the remaining asthmatics separated almost evenly into each subfamily of LTB4, LTC4, LTD4 and LTE4 based asthmatics. While this is a small study, these data indicate differing types of LT based asthmatics at the mechanistic level.
Lung protection against the chemotherapeutic agent bleomycin by dietary flaxseed oil.
Authors: Kamal Afrin , Herndon Betty , Molteni Agostino , Lawrenz Josh , Smith Brendon , Addington Stephen , Smith Brad , Alajajian Sarah , Gasper David , Baybutt Richard C
Published: June 2021

In earlier work we showed a reduction in bleomycin‐induced pathology by a diet high in flax seed oil. To investigate the protective mechanisms, 36 rats were fed a diet with 15% flaxseed oil 4 wk, and 36 controls received 15% corn oil. These received either 8 units/kg intratracheal bleomycin or vehicle, and lungs were harvested at 2, 7 and 21 days. Tissue was fixed and stained (H&E) for histopathology. Two investigators evaluated inflammatory cell infiltrate, septal thickness, emphysema, medial vasculitis, adventitial vasculitis and edema. Animals receiving flaxseed oil showed a delay in edema formation (p=0.025), and a significant decrease in inflammatory cell infiltrate and vasculitis (p=0.04 and 0.007 respectively) compared to corn oil controls. Rats receiving bleomycin and flaxseed oil had significantly reduced pulmonary septal thickness (p=0.013) over positive controls. No differences were observed among groups at 21 days. The time course for injury (highest at 7 and even 2 days) suggests protection for Clara cells. The significant reduction in septal thickness in the flaxseed oil‐treated groups further suggests protection against fibrotic effects of bleomycin. This model presents a dietary regimen that may prove beneficial against the side effects of this highly effective chemotherapeutic agent and its known toxic effects on the lung. Supported by Wheaton College General Endowment Fund
Serum antioxidant capacity in head and neck cancer patients receiving chemoradiation therapy.
Authors: Hine R Jean , Prior Ronald L , Gardner Kenneth E , Maddox Anne Marie , Siegel Eric R. , Stack Brendan C , Vural Emre A , Suen James Y.
Published: June 2021

Measurements of in vivo antioxidant capacity might help to resolve a controversy about the safety of antioxidant supplements during concurrent chemoradiation therapy for cancers of the head and neck. Some data suggest that antioxidant supplements chemoradiation therapy may reduce treatment toxicities; other evidence indicates they may protect tumor cells from oxidative damage. The study defined serum antioxidant capacity before, during and after outpatient therapy. Total, hydrophilic and lipophilic oxygen radical absorbance capacity was measured in serum using the ORACFL microplate method. The procedure is based on the inhibition of peroxyl radical‐ induced oxidation initiated by thermal decomposition of azo‐compounds using fluorescein as the fluorescent probe. The median age of the subjects (n=27) was 58. Forty four percent took supplements. Most were smokers and reported smoking during therapy. Quantifying antioxidant intakes was complicated by treatment toxicities. Total (P=0.0081) and hydrophilic (P=0.0222) antioxidant capacity declined significantly between the pretreatment and end of radiation visits; no changes were detected in the lipophilic fraction. There was, however, marked interindividual variability in the temporal patterns of the lipophilic fraction. To our knowledge, this is the first report of antioxidant capacity in cancer patients receiving similar treatments.
The short term effects of exercise and omega‐3 fatty acids on the plasma lipid profile in Wheaton College students
Authors: Thomas Rachael M , Johnson Hannah , Smith Maria , Baybutt Richard
Published: June 2021

Dyslipidemia plays a major role in the development of atherosclerosis, an underlying factor in cardiovascular disease. Both exercise and dietary modifications are key to positively influence lipid profile. The purpose of this study was to examine the immediate effects of a single bout of moderate‐intensity exercise and short‐term consumption of omega‐3 fatty acids on the plasma lipid profile in men and women. Fourteen Wheaton College students (7 males and 7 females) participated in 2 single bouts of 60 minutes, moderate‐intensity exercise (55‐60% of estimated HRmax), separated by a 14 day consumption of 3 g of fish oil per day. Plasma lipid profile was measured before and after each bout of exercise; measurements were grouped as baseline (BA), post‐exercise (PE), post‐fish oil (PF), or post‐exercise and fish oil (PEF). Significant differences were found in female TRIG, male HDL, female VLDL, and male ratio TC/HDL (P < 0.05). From BA to PEF, males had significantly higher HDL values by 16%, and significantly lower TC/HDL values by 13% (p < 0.017). TRIG and VLDL levels in females were significantly increased by 47% and 55% (p < 0.017) from BA to PEF, respectively. Separately, acute exercise and short‐term fish‐oil intake were not shown to significantly change the lipid profile; however, when combined, HDL increased in males, and TC/HDL decreased in males.
Consensus Measures of Diet for Genome‐wide Association Studies of Gene‐Nutrient Interactions in Health and Disease
Authors: Stover Patick J. , Ordovas Jose M. , McCalla Anne‐Lyne , Hammond Jane , Maiese Debbie , Wagner Diane , Hamilton Carol M , Ramos Erin
Published: June 2021

Gene‐nutrient interactions influence risk for initiation and progression of some common chronic diseases. Genome‐wide association studies (GWAS) permit comprehensive identification of genetic variation that interacts with dietary components to confer risk or protective phenotypes. Data pooling is essential to achieve statistical power to identify and confirm gene‐diet associations. Therefore GWAS require replication in multiple populations, but such approaches are often hindered by lack of consistency in quantifying dietary exposures and nutritional status among studies, including choice of measures and methodologies. The PhenX (consensus measures for Phenotypes and eXposures) was initiated to enhance cross‐study analyses of GWAS and large‐scale genomic research through definition standardization of disease phenotypes, risk factors, and environmental exposures. The PhenX "Nutrition and Dietary Supplements" Working Group finalized measures and protocols to be recommended to the research community will be available in the PhenX Toolkit (http://www.phenx.org). This will enable comparability and replication of GWAS studies to investigate gene‐nutrient interactions.
The effects of whole mushrooms during inflammation
Authors: Martin Keith R , Yu Sanhong , Weaver Veronika , Cantorna Margherita
Published: June 2021

Consumption of edible mushrooms has been suggested to improve health. A number of isolated mushroom constituents can also modulate immunity. Five commonly consumed edible mushrooms were tested to determine whether they stimulate the immune system in vitro and in vivo. White button (WB) extracts readily stimulated macrophage production of TNF‐α. Crimini, maitake, oyster and shiitake extracts also stimulated TNF‐α production but the levels were lower than WB. Primary cultures of murine macrophage and ovalbumin (OVA) specific T cells showed that whole mushroom extracts alone did not affect cytokine production but co‐stimulation with either lipopolysacharide or OVA (respectively) induced TNF‐α, IFN‐γ, and IL‐1β while decreasing IL‐10. WB‐fed mice (2%; 4 weeks) showed no effect on ex vivo immune responsiveness or associated toxicity (pathology of liver, kidney and gastrointestinal tract). Mice fed 1% WB and stimulated with dextran sodium sulfate (DSS) were protected from DSS‐induced weight loss. In addition, 2% WB feeding protected mice from transient DSS‐ induced colonic injury. The TNF‐α response in the colon and serum of the DSS challenged and 2% WB fed mice was higher than controls. Data support a model whereby edible mushrooms regulate immunity in vitro. In vivo effects of edible mushrooms required a DSS challenge to detect TNF‐α changes and transient protection from colonic injury. There were modest effects of in vivo consumption of edible mushrooms on induced inflammatory responses. The result is not surprising since it certainly would be harmful to strongly induce or suppress immune function following ingestion of a commonly consumed food.
Improved small volume resuscitation from hemorrhagic shock with acellular hemoglobin
Authors: Cabrales Pedro , Tsai Amy G. , Intaglietta Marcos
Published: June 2021

Systemic and microvascular hemodynamic responses to hemorrhagic shock volume resuscitation with hypertonic saline (HTS) followed by infusion of polymerized bovine Hb (PBH) at different concentrations were studied in the hamster window chamber model, to determine role of plasma oxygen carrying capacity and vasoactivity during resuscitation. Moderate hemorrhagic shock was induced by arterial controlled bleeding of 50% of blood volume (BV), and a hypovolemic state was maintained for one hour. Volume was restituted by infusion of HTS (7.5% NaCl) 3.5% of BV of followed by 10% of BV of PBH at two different concentrations. Resuscitation was followed for 90 minutes and was carried out using 13 gPBH/dl [PBH13], PBH diluted to 4 gPBH/dl in albumin solution at matching colloidal osmotic pressure (COP) [PBH4], and albumin only solution at matching COP [PBH0]. Systemic parameters, microvascular hemodynamics and functional capillary density (FCD) were determined during hemorrhage, hypovolemic shock and resuscitation. PBH13 caused higher arterial pressure without reverting vasoconstriction and hypoperfusion. PBH4 and PBH0 had lower MAP and partially reverted vasoconstriction. Only treatment with PBH4 restored perfusion and FCD when compared to PBH13 and PBH0. Blood gas parameters and acid‐base balance recovered proportionally to microvascular perfusion. Tissue pO2 was significantly improved in the PBH4 group showing that limited restoration of oxygen carrying capacity is beneficial and compensates for the effects of vasoactivity characteristic of molecular hemoglobin solutions proposed as blood substitutes.
Enhanced cyclooxygenase 2‐mediated vasodilatation in coronary arteries from insulin resistant obese Zucker rats
Authors: PRIETO DOLORES , SÁNCHEZ PINA ANA , CONTRERAS CRISTINA , VILLALBA NURIA , PILAR MARTINEZ
Published: June 2021

Despite selective inhibitors of COX‐2 are highly effective anti‐inflammatory and analgesic drugs, they have been reported to increase the risk of myocardial infarction and atherothrombotic events. In the current study we investigated the role of arachidonic acid (AA) metabolism via cyclooxygenase 1 (COX)‐1) and 2 (COX‐2) in coronary arteries from prediabetic obese Zucker rats (OZR). Branches from the coronary artery from OZR and from lean Zucker (LZR) rats were mounted in microvascular myographs to assess vascular function and COX expression was determined by both immunohistochemistry and Western blot. AA elicited relaxations of similar magnitude in arteries from LZR and OZR which were abolished by mechanical endothelial cell removal. The AA relaxant responses were enhanced by selective inhibition of COX‐1 in both LZR and OZR. Inhibition of COX‐2 markedly reduced the vasodilatation induced by AA in OZR, but not in LZR. COX‐1 was distributed in the coronary endothelium from both LZR and OZR, while COX‐2 was predominantly expressed in the endothelium from OZR. Whereas COX‐1 protein levels were similar in both strains, COX‐2 protein content was increased by 2.5‐ fold in arteries from OZR. AA is mainly metabolised to vasoconstrictor prostanoids via COX‐1 in coronary arteries from healthy animals. In arteries from insulin resistant OZR, endothelial COX‐2 is up‐regulated to produce vasodilator prostaglandins.
Differential Translational Regulation of SLUG mRNA By a uORF In SLUG‐High and SLUG‐Low Cancer Cells
Authors: Yarlagadda Vidhush K , Misra Smita , Mittal Mukul K , Chaudhuri Gautam
Published: June 2021

The metastasis regulator protein SLUG is stringently regulated during cellular growth and development. This gene is expressed from a single promoter and produces an mRNA with a 164 nucleotide 5′‐untranslated region (5′‐UTR). There is a highly conserved tricodon upstream open reading frame (uORF) at the 5′‐UTR in human SLUG mRNA. uORFs are known to regulate the translation of mRNAs. Here we report differential inhibition of SLUG mRNA translation by this uORF in SLUG‐high and SLUG‐low breast cancer cells. We cloned the 164 base‐pair 5′‐UTR sequence in front of the firefly luciferase gene in pGL3‐Control plasmid in the appropriate orientation. Dual luciferase assay with transiently transfected cells showed 50–60% inhibition of the expression of luciferase activity in the SLUG‐low but not in the SLUG‐high breast cancer cells. Mutagenesis of the ATG codon in the uORF to TAG abrogated this inhibition. This data indicates that human SLUG gene expression is differentially regulated by the uORF. We are currently investigating the mechanism behind this differential regulation of SLUG expression. Understanding the molecular mechanism of this inhibition will contribute towards the development of chemical intervention measures against the expression of this important metastasis regulatory protein and thus against breast cancer metastasis.