bluuebunny/crossref_metadata_2025 huggingface dataset:
Mice with null mutation of Ceacam1 exhibit metabolic syndrome with cardiac hypertrophy
Authors: Morgan Eric E. , Ledford Kelly J , DeAngelis Anthony M , Buehler Jason , Patel Payal , Najjar Sonia M , Khouri Samer , Pierre Sandrine V
Published: June 2021

Metabolic syndrome is a risk factor for cardiovascular disease. Because the carcinoembryonic antigen‐related cell adhesion molecule 1 (Ceacam1) regulates insulin and lipid metabolism in liver, we hypothesized that null mutation of Ceacam1 (Cc1−/−) leads to metabolic syndrome and cardiovascular abnormalities. We characterized the metabolic and cardiovascular phenotypes of six month old male Cc1−/− mice.

Cc1−/− mice developed visceral obesity, elevated fasting plasma free fatty acids (FFA), and insulin resistance, as evidenced by fasting hyperinsulinemia and euglycemia (Table). Tail‐cuff measurements revealed higher systolic blood pressure in Cc1−/− mice, and echocardiography of left ventricular end‐diastolic diameter (LVDd), septal (SW) and posterior (PW) wall thicknesses revealed an increased relative wall thickness (RWT; calculated as SW+PW/LVDd). No differences in systolic function were observed (data not shown).

We conclude that null mutation of Ceacam1 mimics metabolic syndrome and results in elevated systolic blood pressure and left ventricular hypertrophy with preserved systolic function.

This work was supported by NIH grants DK054254 (SMJ) and HC36573 (SVP).
A lipid mediated mechanism increases protein O‐glycosylation in hearts of Type 2 diabetic mice
Authors: Fricovsky Eduardo Santiago , Suarez Jorge , Hu Yong , Belke Darrell , Scott Brian , Wang Hong , Dillmann Wolfgang H.
Published: June 2021

Increased nuclear protein O‐linked β‐N‐acetylglucosamine glycosylation (O‐GlcNAcylation) mediated by severe hyperglycemia of diabetes mellitus Type 1 contributes to cardiac myocyte dysfunction. However, it is unknown if increased O‐GlcNAcylation also occurs in the hearts of Type 2 diabetic (T2D) mice with mild hyperglycemia and insulin resistance. Type 2 diabetes was induced in mice by feeding a high fat diet (60% fat) and administering a single low dose of streptozotocin (50mg/Kg). Animals were studied after 10 weeks. Our results show that T2D mice have slightly increased fasting blood glucose levels compared with normal (N) mice (T2D; 5.25±1.43 vs. N; 4.01±0.4 mM). Insulin serum levels were 3 fold increased in T2D mice suggesting insulin resistance. Surprisingly, we found increased protein O‐GlcNAcylation in T2D mouse hearts. Cardiac UDP‐GlcNAc level was also increased. To investigate the basis for the increased protein O‐GlcNAcylation neonatal cardiomyocytes were cultured in 400 μM oleic acid (OA) and normal glucose (5mM) for 48 h. OA dramatically increased protein O‐GlcNAcylation which was reduced after expression of O‐GlcNAcase, which is the enzyme that removes O‐GlcNAc residues from protein. In conclusion, these data suggest that increased lipid utilization results in augmented hexosamine biosynthetic pathway as indicated by increase in protein UDP‐GlcNAcylation levels.
Aging is Associated with Increased Susceptibility to Western Diet‐Induced Glucose Intolerance and Endothelial Dysfunction in Mice
Authors: Folian Brian Joseph , Durrant Jessica R , Connell Melanie L , Russell Molly J , Seals Douglas R , Lesniewski Lisa A
Published: June 2021

Western diets (WD) are associated with impaired glucose tolerance (GT) and vascular endothelial dysfunction, but it is unknown if these deleterious effects are exacerbated in older age. To explore this, GT and endothelium dependent dilation (EDD) were measured in older (O: 29–34 mo) and young (Y: 4–7 mo) B6D2F1 mice (N= 4–5/group) fed WD (40% fat, 6–8 wk) or normal chow (NC). Body mass was greater in O (+3%) and Y (+6%) WD vs. NC (Y vs. O, ns). However, epididymal fat mass was ∼350% greater in O, but only 97% greater in Y (WD vs. NC, P<0.01). The area under the curve for glucose during a GT test (GTAUC) was greater in O WD vs. NC (1.63±0.13 vs. 1.27±0.05 (X 104) mg*min/ml, P=0.02), but not in Y (WD: 1.23±0.04 vs. NC:1.22±0.05). EDD (maximal carotid artery dilation to acetylcholine) was lower in O WD vs. NC (66±6 vs. 80±3%, P=0.03) but not in Y (WD: 91±8 vs. NC: 98±2%, P=0.43). Nitrotyrosine abundance, a marker of oxidative stress, was greater in the aortas of O vs. Y NC (1.8±0.4 vs. 1.0±0.2 AUs, P=0.02), but was increased in WD vs. NC only in O (+45%, 4.0±0.4 AUs, P<0.01). These preliminary observations suggest that aging is associated with impaired resistance to the potentially adverse effects of WD on glucose tolerance and vascular endothelial function in B6D2F1 mice, possibly via exacerbation of oxidative stress. This could contribute to the increased risk of cardiovascular diseases with aging. NIH AG015897, AG006537, AG013038, AG022241
Cholinergic location of delta opioid receptors in canine atria and sinoatrial node
Authors: Deo Shekhar H. , Barlow Matthew A. , Gonzalez Leticia , Yoshishige Darice , Caffrey James L. , Garcia Tiffany
Published: June 2021

Although delta opioid receptors (DORs) are associated with ischemic preconditioning and with vagal transmission in the sinoatrial (SA) node and atria, their precise location remains unconfirmed. DORs and the autonomic markers vesicular choline transporter (VAChT) and tyrosine hydroxylase (TH) were labeled in tissue slices and synaptosomes from canine atrium and SA node. Synapsin I verified the neural character of labeled structures. Acetylcholine and norepinephrine content indicated both cholinergic and adrenergic synaptosomes were present. VAChT and TH signals indicated more than 80% of synapsin positive synaptosomes were cholinergic and less than 8% were adrenergic. DORs co‐localized 75–85% with synapsin in tissue slices of atrium and SA node. Co‐localization was equally strong (85%) for nodal synaptosomes but less so for atrial synaptosomes (57%). Co‐localization between DOR and VAChT was 75–85% regardless of the source. DOR/TH overlap was uniformly low (8–17%). Western blots of synaptosomal extracts confirmed two DOR bands at molecular weights corresponding to reports for DOR monomers and dimers. DOR abundance was greater in nodal synaptosomes than in atrial synaptosomes due to more monomers in the SA node. The preferential association of DORs with cholinergic nerve terminals supports the hypothesis that post‐ganglionic prejunctional DORs regulate local vagal transmission within the heart.
EFFECTS OF CHRONIC HYPOXIA ON UPPER AIRWAY AND DIAPHRAGM MUSCLE ENDURANCE IN THE DEVELOPING RAT
Authors: O'Halloran Ken D , Mitchell Gordon S , Bradford Aidan , Carberry Jayne
Published: June 2021

Chronic hypoxia (CH) is common in respiratory disease and is known to alter skeletal muscle structure and oxidative capacity. We hypothesized that CH in early development would alter respiratory muscle function. Wistar rats were reared in normobaric normoxia or hypobaric hypoxia (PB=450mmHg) for 7 days beginning at P1, P11, P21 or P31. Isolated muscle function studies were conducted at P19, P29 or P39. Fatigue was assessed in response to repeated tetanic contractions (40Hz, 300msec) every 2 sec for 5 minutes. Chronic hypoxia during early development (beginning P1 and P11) increased sternohyoid specific force but significantly reduced muscle endurance (fatigue index ie ratio of force at 5 min of fatigue to initial force, measured at P19 was 83+/−7% vs. 59+/−7%* and 63+/−4%*; control vs. P1 and P11 hypoxic groups, *P<0.05 ANOVA). CH in older animals did not affect sternohyoid endurance. CH increased diaphragm specific force but had no effect on muscle endurance (eg fatigue index at P19 was 81+/−6% vs. 76+/−4% and 88+/−4%; control vs. P1 and P11 hypoxic groups, P>0.05 ANOVA). We conclude that CH elicits plasticity in respiratory muscles with differential effects in diaphragm and upper airway muscles. Increased fatigue of the sternohyoid muscle may have important consequences for the control of upper airway patency in vivo.
Renin angiotensin blockade attenuates proinflammatory cytokines activation in the microglia of paraventricular nucleus of heart failure rats
Authors: Kang Yu‐Ming , Gao Li‐Jing , Lei Jing‐Hui , Yang Zhi‐Ming , Lv Ji‐Yuan , Francis* Joseph
Published: June 2021

Microglia are activated in the paraventricular nucleus (PVN) of the hypothalamus, where they may contribute to the release of proinflammatory cytokines (PIC) that stimulate sympathoexcitatory systems. In this study we determined whether blockade of AT1‐R (valsartan), or inhibition of microglial activation (minocycline) or angiotensin‐converting enzyme (captopril) attenuates PIC in heart failure (HF) rats. Adult male Sprague‐Dawley rats were subjected to coronary artery ligation to induce HF or sham surgery (SHAM). Subsequently, animals were orally treated with valsartan (VAL, 30mg/kg/day), minocycline (MIN, 10mg/kg/day) or captopril (50mg/kg/day) for 4 weeks. Immunohistochemistry revealed activation PIC in the microglial cells and PVN neurons of HF rats compared with SHAM. Oral treatment with VAL, MIN or captopril inhibited microglial activation and attenuates PIC in the PVN of HF rats. Treatment with VAL, MIN or captopril also reduced plasma levels of NE and PIC. These findings suggest that activated microglia in the paraventricular nucleus of hypothalamus contribute to the pathogenesis of heart failure.
Impaired neurogenesis and activated caspase 3‐dependent apoptosis pathway in the rostral ventrolateral medulla cause hypertension in stroke‐prone spontaneously hypertensive rats
Authors: Kishi Takuya , Hirooka Yoshitaka , Sunagawa Kenji
Published: June 2021

Recent studies demonstrated that neural progenitor cells responsible for neurogenesis migrate to the sites of brain injury of epilepsy, stroke and neurodegenerative diseases. It is also well known that apoptosis is involved in regulating synaptic plasticity. It remains unknown the impact of balance between neurogenesis and apoptosis in the rostral ventrolateral medulla (RVLM) in the pathogenesis of hypertension. We examined neurogenesis and caspase 3‐dependent apoptosis initiated by mitochondrial translocation of Bax in the RVLM of stroke‐prone spontaneously hypertensive rats (SHRSP). We identified neuroblasts by staining for doublecortin (Dcx), cell proliferation by absorption of bromodeoxyuridine (BrdU), and apoptosis by staining for caspase‐3, Bax, PKC in membranous, cytosolic and mitochondorial fractions. In hypertensive states, SHRSP had significantly lower Dcx and BrdU expression than WKY (Dcx; −41±4%, BrdU; −33±5%, n=8 for each, P<0.01). In pre‐hypertensive states, the expressions of Dcx and BrdU in the RVLM were significantly decreased compared with WKY. The expression of Bax in mitochondorial fraction and caspase‐3 in cytosolic fraction were significantly increased in SHRSP than in WKY, in either pre‐hypertensive or hypertensive states. In conclusion, the impaired neurogenesis and activated caspase 3‐dependent apoptosis pathway in the RVLM might be involved in hypertension in SHRSP.
Circulatory Interleukin‐6 is Elevated in Overweight Compared to Healthy and Type 1 Diabetic Children
Authors: Rosa Jaime Sou , Oliver Stacy R. , Lee Jane S. , Flores Rebecca L. , Ibardolaza Mila , Zaldivar Frank P. , Galassetti Pietro R.
Published: June 2021

For overweight (OW) and type 1 diabetic (T1DM) adults, the development of micro/macro‐vascular complications is thought to be mediated by low‐grade, chronic inflammation, as reflected by elevated circulatory C‐reactive protein and interleukin‐6 (IL‐6). Whether IL‐6 is also elevated in OW and T1DM children is not well‐established. Baseline plasma IL‐6 was compared between 27 OW (12.2 ± 0.5 yrs; 12F; BMI% 97 ± 4), 37 non‐overweight T1DM (13.9 ± 0.4 yrs; 15F; BMI% 55 ± 3), and 33 healthy children (CTRL; 13.4 ± 0.4 yrs; 20F; BMI% 56 ± 4). Blood glucose of T1DM was maintained within 4.4‐6.1 mmol/L for 90 min prior to blood sampling. As results were not normally distributed, log transformation was required before one‐way ANOVA and the Tukey post‐hoc test were applied (α = 0.05). Progressively greater values of IL‐6 (pg/ml) were observed across groups: CTRL, 1.61 ± 0.36; T1DM, 2.20 ± 0.34; and OW 2.29 ± 0.40, p<0.05 vs CTRL. When subdivided by gender, this pattern was maintained, with statistical significance only detected, however, between male CTRL vs OW subgroups. Subclinical, low‐grade inflammation, as reflected by elevated IL‐6, may be present in OW and possibly T1DM children. This data suggest that for pediatric patients with dysmetabolic conditions, the pathogenetic mechanisms leading to vascular complications may be activated at an early age. Supported by NIH grants M01‐RR00827‐28; K‐23 RR018661‐01; & JDRF #11‐2003‐332.
Exercise Protects Against Endothelial Dysfunction During Oral Glucose and High Fat Load
Authors: Das Emon Kalyan , Gutterman David D. , Phillips Shane A.
Published: June 2021

The development of atherosclerosis and coronary disease is preceded by endothelial dysfunction which can be improved by chronic exercise. Ingestion of a single high glucose (HG) or high fat (HF) meal impairs endothelial function. We tested the hypothesis that chronic exercise protects against the reduction in flow‐mediated dilation (FMD, index of endothelial function) to a HG or HF meal. Sedentary (SED) subjects and athletes consumed either a HF or HG meal with a 7 day washout between. FMD was assessed before and 180 minutes after the meal (maximal ultrasound brachial diameter after reactive hyperemia). Endothelium‐independent dilation was determined via sublingual nitroglycerin (NTG; 0.4mg). Brachial artery FMD decreased in SED (n=9) following HF (10.3% to 6.7% at 120 minutes; and to 6.8% at 180 minutes; p<0.05 for both) and HG (10.3% to 7.0% at 60 minutes; p<0.05) meals. There was no change in FMD after HF and HG loading in athletes (n=5). NTG responses were similar among groups. We conclude that endothelial function is reduced in sedentary individuals after HG and HF diet. Chronic exercise protects against acute diet‐induced endothelial dysfunction.
IL‐10 KO female mice infused with TNF‐α show impaired ACh induced relaxation as compared to IL‐10KO male mice
Authors: Zemse Saiprasad M. , Hilgers Rob H. P. , Cleghorn Donald , Chiao Chin Wei , Brands Michael , Webb R. Clinton
Published: June 2021

Background: TNF‐α is a pro‐inflammatory cytokine, and is an important mediator of maternal endothelial dysfunction. In this study, we tested whether IL‐10 improves TNF‐α induced endothelial dysfunction.

Methods: C57BL6/IL‐10 KO female and male mice were treated with saline or TNF‐α (220ng/kg/min) for 14 days. Aortic rings were isolated and mounted in a wire myograph (Danish Myotech) and stretched to a tension of 5mN. Endothelium‐dependent relaxation was assessed by constructing cumulative concentration‐response curves to acetylcholine (ACh, 0.001–10 μmol/L) during phenylephrine PE, 10 μmol/L)‐induced contraction. Endothelium‐independent relaxations were assessed by the NO donor sodium nitroprusside (SNP).

Results: IL‐10 KO female mice treated with TNF‐ α showed significant impairment in ACh induced relaxation as compared to C57BL6 female mice treated with TNF‐ α (Emax = 52 ± 3% versus 76 ± 5% respectively; P<0.05). IL‐10 KO male mice treated with TNF‐α showed no significant impairment in ACh induced relaxation as compared to C57BL6 male mice treated with TNF‐α(Emax = 57 ± 3% versus 66 ± 3% respectively; P<0.05). TNF‐α infusion had no effect on endothelium‐independent relaxations by SNP in all the groups.

Conclusion: The anti‐inflammatory cytokine IL‐10 prevents impairment in endothelium‐dependent vasorelaxation in response to 14 days of TNF‐α infusion which is more in female mice but not male mice.
Hydrogen peroxide stimulates the Ca2+‐activated Big‐conductance K channels (BK) through cGMP signaling pathway in cultured human endothelial cells.
Authors: Dong Deli , Yue Peng , Wang Wenhui
Published: June 2021

We used the whole cell patch‐clamp technique to examine the effect of H2O2 on BK channels in human endothelial cells. We confirmed the previous finding that a 200 pS BK channel activity was detected when the cell membrane potential was clamped at 50 mV. Application of H2O2 (100 micromole) or adding 1 unit glucose oxidase stimulated BK channels. The stimulatory effect of H2O2 and GO was absent in cells treated with 10 micromole ebselen, a scavenger of superoxide anions. To determine whether the stimulatory effect of H2O2 and GO on BK channels was the result of increasing NO production in the endothelial cells, we examined the effect of H2O2 and GO on BK channels in the presence of 0.1 mM L‐NAME which inhibits NO synthase (NOS). Inhibition of NOS completely abolished the stimulatory effect of H2O2 on BK channels. In contrast, treatment of endothelial cells with D‐NAME did not block the effect of H2O2 on BK channels. Moreover, inhibiting soluble guanylate cyclase (sGC) with ODQ mimicked the effect of L‐NAME and abolished the effect of H2O2. Also, inhibiting protein kinase G with KT5823 abolished the stimulatory effect of H2O2 on BK channels. We conclude that H2O2 stimulates BK channels through NO/sGC/cGMP pathway in the cultured human endothelial cells.
Glu298Asp eNOS Gene Polymorphism Causes Attenuation in Exercise‐Induced Muscle Vasodilatation in Humans
Authors: Dias Rodrigo Goncalves , Alves Maria Janieire Nunes , Pereira Alexandre da Costa , Rondon Maria Urbana P. B. , Krieger José Eduardo , Krieger Marta Helena , Negrão Carlos Eduardo
Published: June 2021

We tested the hypothesis that the vasodilatation responses to exercise were attenuated in homozygous individuals for the allele Asp298.

Methods: from 287 volunteers genotyped, we studied 24 individuals: Glu/Glu (n=15, age 43±3 yr, BMI 22.9±0.3 kg/m2) and Asp/Asp (n=9, age 40±4 yr, BMI 23.5±0.9 kg/m2). Heart rate (HR), mean blood pressure (MBP) and forearm blood flow (FBF) were recorded during handgrip exercise (30% MVC).

Results: FVC responses to exercise were lower in Asp/Asp, compared with Glu/Glu (Δ=0.07±0.14 vs. 0.57±0.09 units, P=.002). To investigate the mechanisms underlying the lowered vasodilatation in Asp/Asp, we studied FBF during L‐NMMA and L‐NMMA + phentolamine in a subset of 8 Glu/Glu and 7 Asp/Asp. We measured muscle sympathetic nerve activity (MSNA). L‐NMMA did not change FVC responses to exercise in Asp/Asp, compared with saline (Δ=−0.05±0.20 vs. −0.09±0.18 units, P=.52). In contrast, L‐NMMA reduced FVC in Glu/Glu (Δ0.79±0.14 vs. 0.14±0.09 units) and the difference between genotypes were no longer observed (P=.62). L‐NMMA + phento increased similarly FVC responses to exercise in Glu/Glu and Asp/Asp (Δ=1.16±0.26 vs. 0.56±0.24 units, P=.43). MBP and MSNA were similar among genotypes.

Conclusion:Asp/Asp individuals have attenuated muscle vasodilatation responsiveness to exercise, which seems to be primarily mediated by a reduced nitric oxide bioavailability.
ACE2 over‐expression ameliorates glycemic homeostasis in diabetic mice
Authors: Bindom Sharell M , Feng Yumei , Raju Sammeta VM , Xia Huijing , Lazartigues Eric
Published: June 2021

ACE inhibitor therapy has been shown to prevent new onset diabetes and pancreatic â‐cell dysfunction. We hypothesized that ACE2 would ameliorate glycemia in diabetic mice. Diabetic obese mice (db/db) and control lean littermates (db/m) were infected with a hACE2 or control (GFP) adenovirus (2x10e6 pfu 100 ìL) to the pancreas. After 7d, fasting blood glucose (12 hrs) and glucose tolerance (GT, 2 g/kg IP) were measured as indices of glycemic control. ACE2 immunofluorescence (arbitrary units) revealed high expression of hACE2 in the liver of mice infected with Ad‐hACE2 vs. Ad‐GFP (120±6 vs. 14±1, n=9, P<0.01), while hACE2 in the pancreas was not different between groups. Body weight (g) in db/db mice was not affected by Ad‐ACE2 (35±1, n=5) or Ad‐GFP (36±1 n=5) delivery. Baseline glycemia (mg/dL) was significantly higher in db/db vs. db/m mice (246±18 vs. 102±6, n=10, P<0.01) and was dramatically reduced in Ad‐hACE2‐ vs. Ad‐GFP‐treated db/db mice (157±12 vs. 203±6, n=5, P<0.05). Ad‐hACE2 infection had no effect on glycemia in db/m (92±8, n=10) compared to Ad‐GFP‐infected mice (75±10). In addition, Ad‐hACE2 infection significantly improved GT in db/db compared to GFP‐treated‐mice (573±43 vs. 749±18 mg/dL respectively, P<0.01) 1 hr post glucose administration. These data confirm the role of ACE2 in diabetes and suggest that ACE2 gene therapy can ameliorate glucose homeostasis in diabetic mice. (NIH NS052479 and RR018766)
H2S‐induced pancreatic acinar cell apoptosis is mediated via JNK and p38 MAP Kinase
Authors: Bhatia Madhav , Adhikari Sharmila
Published: June 2021

Treatment of pancreatic acinar cells by hydrogen sulphide has been shown to induce apoptosis. However, a potential role of mitogen‐activated protein kinases (MAPKs) in this apoptotic pathway remains unknown. The present study examined the role of MAPKs in H2S‐induced apoptosis in mouse pancreatic acinar cells. Pancreatic acinar cells were treated with 10 μM NaHS (a donor of H2S) for 3 hours. For the evaluation of the role of MAPKs, PD98059, SP600125 and SB203580 were used as MAPKs inhibitors for ERK1/2, JNK1/2 and p38 MAPK respectively. We observed activation of ERK1/2, JNK1/2 and p38 when pancreatic acini were exposed to H2S. Moreover, H2S‐induced ERK1/2, JNK1/2 and p38 activation were blocked by pretreatment with their corresponding inhibitor in a dose‐dependent manner. H2S‐induced apoptosis led to an increase in caspase 3 activities and this activity was attenuated when caspase 3 inhibitor were used. Also, the cleavage of caspase 3 correlated with that of Poly‐(ADP‐Ribose)‐Polymerase (PARP) cleavage. H2S treatment induced the release of cytochrome c, smac from mitochondria into the cytoplasm, translocation of Bax into mitochondria and decreased the protein level of Bcl‐2. Inhibition of ERK1/2 using PD98059 caused further enhancement of apoptosis as evidenced by annexin V staining, while SP600125 and SB203580 abrogated H2S‐induced apoptosis. Taken together, the data suggest that activation of ERKs promotes cell survival, whereas activation of JNKs and p38 MAP kinase leads to H2S‐induced apoptosis.
Hindlimb Unloading Induces a Biphasic Temporal Response of Bcl‐2 Apoptotic Signaling in the Rat Soleus Muscle
Authors: Propst Gregory Keith , Kwak Hyo‐Bum , Kim Jong‐Hee , Dalton Ryan Lee , Lawler John M
Published: June 2021

It is now a widely accepted belief that oxidative stress and apoptosis contribute to muscle atrophy in a number of pathologies including severe disuse (e.g., hindlimb unloading (HU), mechanical ventilation), denervation, and congestive heart failure. Severe disuse induces skeletal muscle atrophy through activation of caspase‐3, apoptosis, calpains, and nuclear factor kappa‐B (NF‐κB) during 2–14 days of unloading. Recent data indicates apoptosis in muscle continues 28 days following denervation. We hypothesize that consistent with denervation data and the “constant proportion model,” pro‐apoptotic signaling would remain elevated after long‐term HU (28 days) compared with short‐term HU (2 days). Fischer 344 rats were divided into 3 groups: Control (CON), 2 days of HU (HU2), and 28 days of HU (HU28), after which soleus muscles were extracted. Bax (+117%) and caspase‐3 cleavage (+324%) were increased in the HU2 soleus, while Bcl‐2 levels trended lower (−18%). Contrary to our hypothesis, HU28 decreased Bax levels (−30%), while Bcl‐2 levels were elevated (+95%), and caspase‐3 cleavage trended lower (−15%) than CON. NF‐κB activation was higher than CON at 28 HU. The data indicates that apoptotic signaling during HU is biphasic, rapidly rising early and attenuating pro‐apoptotic signaling during prolonged HU. Our findings suggest a potential role for NF‐κB, and a more dynamic view of the constant domain model.
Inhibitory effects of Tumor Necrosis Factor (TNF)‐α on myoblast to myotube differentiation are attenuated by eicosapentaenoic acid (EPA)
Authors: Magee Peter , Pearson Stephen , Allen Jeremy
Published: June 2021

Elevated concentrations of TNF‐α are associated with muscle wasting, exerted through inhibition of myogenic differentiation and enhanced apoptosis. Previously, we found that EPA, an omega‐3 fatty acid, has a protective effect against apoptosis during myogenesis and therefore we hypothesized that EPA may ameliorate the apoptotic effects of TNF‐α.

To investigate this, murine C2C12 myoblasts were differentiated in culture medium containing 2% horse serum. EPA (50μM) was added at the start of differentiation and myotube formation allowed to progress for up to 5days in the presence or absence of TNF‐α (20ng/ml). Differentiation was evaluated by morphology and anti‐myosin antibody. Apoptosis was quantified by measurement of caspase activity.

It was found that EPA alone had no effect on myogenesis after 5 days compared with controls, whereas TNF‐α significantly reduced (P<0.05) myosin expression after 5 days and significantly increased (P<0.001) caspase‐8 activity. The effects of TNF‐α were ameliorated for both myosin (P<0.05) and apoptosis (P<0.008) by co‐treatment with EPA.

In conclusion, EPA has a protective action against the damaging effects of TNF‐α on myogenesis. Our findings support potential therapeutic applications for EPA in associated muscle wasting conditions such as ageing and during skeletal muscle regeneration following injury. Supported by the University of Salford.
IL‐6 Inhibits Hyperoxia Induced Bax Translocation Through Pi3kinase/AKT Mediated Bax Phosphorylation
Authors: Kolliputi Narasaiah V , Meloney Laura G , Kathryn Steiner M , Waxman Aaron B
Published: June 2021

We have shown that IL‐6 over expression protects mice from hyperoxic acute lung injury (HALI) in vivo and treatment with IL‐6 protects against oxidant mediated cell death in vitro. However, the mechanisms to account for these findings are not yet known. We hypothesized that Bax, a pro‐apoptotic member of Bcl‐2 family, may play an important role in oxidant mediated cell death and that IL‐6 may block this pathway. We characterized the expression, and localization of Bax and its regulation in WT and IL‐6 lung specific over expression transgenic (IL‐6 Tg(+)) mice exposed to 100% oxygen for 72 hours in vivo. We also analyzed the same proteins in HUVEC with and without H2O2 in the presence or absence of IL‐6. In control HUVEC treated with H2O2, or WT mice exposed to 100% oxygen, there was marked induction of Bax translocation and dimirization associated with increased pJNK and p38kinase activity. In contrast, JNK or p38 kinase inhibitors, or treatment with IL‐6 inhibited Bax mitochondrial translocation and apoptosis of HUVEC cells. IL‐6 Tg(+) mice exposed to 100% oxygen, have reduced pJNK, p38 levels, enhanced Pi3kinase/AKT kinase, and increased serine phosphorylation of Bax compared with WT mice. Moreover IL‐6 treated cells or S184 transfected cells have increased Bax phosphorylation and decreased translocation. IL‐6 potently blocks either hyperoxia or oxidant‐induced Bax insertion intomitochondrial membranes. We concluded that IL‐6 is cytoprotective, in part, by suppressing Bax translocation and dimirization through Pi3kinase/AKT mediated Bax phosphorylation.

This work is supported by NIH grant RO1HL074859 to Waxman A.B and NIH training grant 5 T32 HL 007874‐12 to Steiner M. K.
Skin permeability and water relations of sea snakes
Authors: Lillywhite Harvey B. , Babonis L. , Sheehy C. , Tu M.‐C.
Published: June 2021

We studied transepithelial evaporative water loss (TEWL) and skin resistance (Rs) in three species of sea kraits differing in both terrestrial tendency and water permeability of skin. Laticauda colubrina is semi‐terrestrial and its mean TEWL is 0.118 ± 0.11 mg cm−2 h−1; L. laticaudata is relatively more aquatic and TEWL = 0.37 ± 0.20 mg cm−2 h−1; L. semifasciata is the most fully aquatic and TEWL = 0.54 ± 0.31 mg cm−2 h−1 (P<0.0001, ANOVA for TEWL measured at 23–24 C). Corresponding Rs values are 401 ± 103, 176 ± 33, and 93 ± 12.9 s cm−1 respectively. Thus, Rs is significantly greater in the more terrestrial species and lowest in the more aquatic species (ANOVA, P<0.0001). Losses of water from snakes kept in sea water exhibit a reversed trend with lower rates of loss in the more aquatic species (mean % body mass lost per day = 0.68 ± 0.05 in L. c., 0.50 ± 0.04 in L. l., and 0.33 ± 0.02 in L. s.). External features of the stratum corneum are hypothesized to explain the lower rates of water loss observed in sea water, while lipids and ultrastructural features of the skin are also being investigated. The countergradient pattern of water losses in air vs. sea water appear to reflect adaptive traits favorable to the maintenance of water balance in amphibious species which differentially utilize both marine and nearshore terrestrial environments. Supported by the National Geographic Society (8058‐06) and Delfin Technologies Ltd.
Developmental changes in endothelium‐dependent relaxation of the chicken ductus arteriosus
Authors: Villamor Eduardo , Ågren Pia , van der Sterren Saskia , Frazziano Giovanna , Blanco Carlos E , Perez‐Vizcaino Francisco , Cogolludo Angel L
Published: June 2021

We tested the hypothesis that endothelium‐dependent relaxation in the chicken ductus arteriosus (DA) is developmentally regulated. Isolated DA rings from 15‐, 19‐ and 21‐day‐old chicken embryos relaxed to acetylcholine (ACh). This relaxation was unaffected by indomethacin but impaired by endothelium removal, by the NO synthase inhibitor L‐NAME, and by the soluble guanylate cyclase inhibitor ODQ, suggesting the involvement of NO. This NO production was confirmed with the fluorescent probe DAF‐2DA. The combination of apamin and charybdotoxin with L‐NAME produced a further inhibition of ACh‐induced relaxation, suggesting the participation of a putative EDHF. In the 21‐day DA, the relaxations induced by ACh and sodium nitroprusside SNP were markedly reduced and scanning electron microscopy demonstrated an irregular endothelial lining with protrusion and detachment of endothelial cells. The relaxations induced by BAY 41‐2272 and 8‐Br cGMP were not affected by age. In summary, we found that ACh induces endothelium‐dependent relaxation of the chicken DA and that NO and EDHF are involved in this response. During chicken DA closure, endothelial cells undergo morphologic and functional alterations that result in the lack of endothelium‐dependent relaxation.

Supported by “Fundación de Investigación Médica Mutua Madrileña”
The effect of temperature on cellular Ca+ flux in ventricular myocytes from bluefin tuna
Authors: Shiels Holly A , Thompson S. H. , Block B. A.
Published: June 2021

Bluefin Tuna (Thunnus orientalis) have the capacity to move from warm surface waters to cooler waters beneath the thermocline where they will be faced with the challenge of a cold heart delivering blood to metabolically warmed tissues. We used confocal microscopy to investigate the temporal and spatial characteristics of cellular Ca2+ flux in bluefin tuna ventricular myocytes to further our understanding of the thermal tolerance of heart function in these animals. Tuna were acclimated to either 15°C or 23°C for a minimum of 3 weeks. Cardiac myocytes were isolated from the spongy myocardium and loaded with the Ca2+indicator dye Fluo‐4 and field‐stimulated to contract. Line scan images across the width of the myocyte revealed spatial inhomogeneities in the temporal properties of the Ca2+ transients. The Ca2+ wavefront initiated faster, rose faster and reached larger peak amplitude at the periphery of the cell compared with the centre. This pattern was exacerbated during acute cooling and when the sarcoplasmic reticulum was inhibited with ryanodine and thapsigargin.
Comparative analysis of DNA repair pathways in mammals.
Authors: Podlutsky Andrej , Podlutskaya Natalia , Bakri Imad , Csiszar Anna , Ungvari Zoltan , Austad Steven
Published: June 2021

In our study we used three groups of animals: rodents, wild mice and white footed mice; bats, little brown bat and Brazilian free‐tailed bat; primate, marmoset and humans. These groups have different longevity quotient: rodents (0.8 and 1.6); bats (8.1 and 2.7?); primates (4.2 and 2.3) respectively. We hypothesize that DNA repair ability and DNA stability might correlate with species maximum longevity. In order to study different pathways of DNA repair we treated cells with gamma‐radiation (targeting base excision repair, BER) or UV‐C light (targeting nucleotide excision repair, NER). We monitored cell survival as well as DNA‐damage level with comet‐assay. Our results indicate that bat's cells are similar to primate cells following gamma radiation – they survive better and have less damage then rodent's cells. This suggests that bats and primates have superior BER compared with rodents. However, following UV‐C treatment, bat's cells are similar to rodent's cells – they are very sensitive to UV‐type of damage; at same time both primate species show comparable resistance to UV, superior to rodent or bat. These results are indicating that NER pathway is not efficient in repairing DNA in rodents and bats. Study with caspase activation and ROS production are supporting our conclusion ‐ exceptional bat's longevity is not dependant from NER pathway.
Null of Se‐glutathione peroxidase‐1 (GPX1) or Cu, Zn‐superoxide dismutase (SOD1) suppresses murine pancreatic insulin synthesis and secretion
Authors: Wang Xiaodan , Vatamaniuk Marko Z. , Roneker Carol A. , Lei Xin Gen
Published: June 2021

Physiological roles of antioxidant enzymes in body insulin metabolism still remain elusive. We compared impacts of null of GPX1 and SOD1 alone or in combination on pancreatic insulin synthesis, secretion, and function. Male mice (8‐wk old) of GPX1 null (GPX1−/−), SOD1 null (SOD1−/−), double knockout of both enzymes (DKO), and their wild‐type (WT) were fed a Se‐adequate diet and used for various tests (n = 6 per genotype by assay). Compared with the WT, the 3 knockout groups exhibited lower (P < 0.05) pancreatic β cell mass (44 to 59%), plasma insulin concentration (46 to 68%), glucose stimulated insulin secretion in cultured islets and plasma (27 to 79%), and ATP production in islets (52 to 67%), along with elevated (P < 0.05) hydroperoxide production in islets. However, these 3 null groups had glucose tolerance similar to the WT. Despite showing a higher (P < 0.05) fasted blood glucose concentration, the SOD1−/− actually had improved insulin tolerance over the WT. Although both GPX1−/− and SOD1−/− islets had elevated phosphor‐p53 and phosphor‐p38 MAPK protein levels than the WT, only the SOD1−/− islets showed lower (P < 0.05) mRNA and protein levels of pancreatic duodenal homeobox‐1 and its upstream regulator forkhead protein a2. In conclusion, null of SOD1 exerted more pronounced impact on pancreatic insulin metabolism and function than that of GPX1, and the DKO did not worsen the scenario. [NIH DK 53018 to XGL]
Impact of selenium and selenoproteins on regulation of glucose homeostasis and insulin sensitivity in mice
Authors: Labunskyy Vyacheslav M. , Hatfield Dolph L. , Gladyshev Vadim N.
Published: June 2021

Selenium is an essential trace element, which has been shown to be effective in cancer prevention. Whereas increased levels of selenium in the diet have important health benefits, recent clinical trials have demonstrated that supplemental intake of selenium above the adequate level raises the risk of type 2 diabetes mellitus. However, the molecular mechanisms for the effect of dietary selenium on the development of this disease are not well understood. In the present study, we examined the contribution of selenoproteins to increased risk of developing diabetes. C57BL/6J mice (n=6–8 per group) were fed either selenium deficient Torula yeast based diet or diets supplemented with 0.1 and 0.4 ppm selenium. Our data show decreased insulin sensitivity in mice maintained on selenium supplemented compared to selenium deficient diets. As expected, we observed elevated levels of stress‐related selenoproteins, such as GPx1 and MsrB1, while expression of housekeeping selenoproteins, such as TR3, was not significantly changed upon increased concentration of selenium in the diets. In addition, our preliminary data show a dysregulation of glucose homeostasis in transgenic mice encoding an i6A− mutant Sec tRNA, which are characterized by reduced synthesis of selenoproteins. Together with previous reports showing impaired insulin sensitivity in mice that overexpress GPx1, these data suggest a possible role for the stress‐related selenoproteins in regulation of glucose homeostasis and insulin sensitivity in mice.
Lipid effects of prescription omega‐3‐acid ethyl esters plus simvastatin in subjects with hypertriglyceridemia
Authors: Maki Kevin C. , McKenney James , Lubin Barry C. , Reeves Matthew S.
Published: June 2021

Prescription omega‐3‐acid ethyl esters (P‐OM3) are approved as an adjunct to diet for treatment of very high triglyceride (TG) levels, and are frequently used in combination with a statin to lower persistent hypertriglyceridemia. The present randomized, crossover trial evaluated 6 weeks of combination therapy with simvastatin 20 mg/d (S) plus P‐OM3 4 g/d (Lovaza™) or placebo in 39 men and women with TG 200 to 600 mg/dL and non‐HDL cholesterol (C) above the subject's treatment goal after a 5‐week diet lead‐in. Median baseline concentrations of non‐HDL‐C, TG, LDL‐C, and HDL‐C were 209, 304, 163 and 38 mg/dL, respectively. Non‐HDL‐C decreased by 40.0% with S+P‐OM3 compared to 34.3% for S + placebo (p<0.001). The following also showed favorable changes (p<0.05) with S + P‐OM3 compared to S + placebo: VLDL‐C (−41.5% vs. −22.4%), TG (−43.6% vs. −28.7%), total‐C (−31.0% vs. −26.4%), HDL‐C (+16.4% vs. +10.6%), apolipoprotein B (−31.6% vs. −28.4%), total‐C:HDL‐C ratio (−39.1% vs. −32.6%) and TG:HDL‐C ratio (−50.6% vs. −36.9%). LDL‐C (−37.2% vs. −38.4%) and apolipoprotein A‐I (+0.9% vs. +4.3%) responses did not differ significantly between treatments. Thus, S + P‐OM3 and dietary counseling improved non‐HDL‐C and other lipid and lipoprotein parameters to a greater extent than S alone.

This was funded as an Investigator‐Initiated Trial (NCT00487591) by Reliant Pharmaceuticals, Inc.
Effects of zinc (Zn) supplementation and phytate‐reduced maize in 6‐12 mo infants in Guatemala
Authors: Krebs Nancy F , Mazariegos Manolo , Westcott Jamie E , Wright Linda , Das Abhik , Goco Norman , Hartwell Tyler , Solomons Noel W , Raboy Victor , Angel Luis , Hambidge K Michael
Published: June 2021

Attaining adequate zinc intake from complementary foods for older infants is a challenge in populations primarily dependent on cereal based diets. Objective: To test the effects of a zinc supplement (ZS) and/or phytate reduced maize for use as a complementary food on growth and other functional outcomes. Methods: In a 2x2 design, a total of approximately 400 primarily breastfed infants’ families from the Western Highlands of Guatemala were randomized to receive phyate‐reduced maize or wild‐type hybrid maize; infants were further randomized to receive a zinc supplement (5 mg/d) or placebo from 6‐12 mo of age. Anthropometry was obtained monthly; morbidity weekly; Bayley Scales of Infant Development (BSID) at 6 and 12 mo; and plasma [Zn] at 12 mo. Results: Stunting was highly prevalent in all groups by 6 mo of age. Linear growth (and other anthropometry) did not differ among treatment groups. No differences were observed by treatment group for morbidity due to diarrhea, or BSID motor or mental scales. The BSID Behavior Scale was higher in the ZS group (p = 0.02), and mean plasma [Zn] was higher in the ZS group (p = 0.05). Conclusion: The lack of impact of the interventions in this population likely reflects a multifactorial basis for the high rates of stunting. Support: Global Network for Women's and Children's Health Research, Bill and Melinda Gates Foundation, and NIH Office of Dietary Supplements.
Dietary iron deficiency increases Zip14 expression in rats
Authors: Nam Hyeyoung , Gulec Sukru , Knutson Mitchell D
Published: June 2021

Zip14 (Slc39a14) is a transmembrane protein that can transport iron and zinc. We have found that Zip14 expression is markedly up‐regulated in the liver, spleen, and duodenum of rats made anemic by repeated phlebotomy, suggesting that Zip14 responds to body iron status. The present study aimed to determine how Zip14 expression is affected by iron deficiency and overload. Weanling male Sprague‐Dawley rats (n=18) were randomized to 3 dietary groups: iron deficient (9 ppm Fe), control (215 ppm Fe), or iron‐loaded (2.9% carbonyl Fe). After 3 weeks, mean hemoglobin levels were 7.2, 13.2, and 14.5 g/dL, respectively, and mean liver non‐heme iron levels were 0.27, 1.56, and 93.5 μmol/g. Immunoblot analysis reveals that Zip14 protein levels increased dramatically in iron‐deficient liver, pancreas, and heart, tissues that express abundant Zip14. In iron‐deficient liver, zinc concentrations did not differ from controls, indicating that the marked increase in Zip14 levels did not result from altered zinc status. Zip14 expression also increased in duodenum in response to iron deficiency. The iron transport capacity of Zip14 along with its robust induction by iron deficiency strongly implicates Zip14 in physiologic iron metabolism. Supported by NIH grant DK065064 (MDK).
The adaptor for stress‐related selenoprotein synthesis does not require aminoacylation for its maturation
Authors: Carlson Bradley A , Xu Xue‐Ming , Lee Byeong Jae , Gladyshev Vadim N , Hatfield Dolph L
Published: June 2021

The mammalian selenocysteine (Sec) tRNA population contains two isoforms that differ by a 2′‐O‐methyl group on the ribose at position 34, designated Um34. Um34 synthesis at position 34 is dependent on the presence of selenium (Se) and the resulting Um34‐modified isoform in turn is required for expression of stress‐related selenoproteins (SRSPs). Since Se is involved in 1) Sec synthesis that occurs on its tRNA, 2) the Se donor, SPS2, a selenium‐containing protein, and 3) Um34 synthesis, a question is raised regarding the number of steps Se is required before the SRSP adaptor is generated, i.e., must this isoform be aminoacylated before the final methylation event? If so, then Se is required at multiple steps for Um34 synthesis. To address this question, we mutated the discriminator base at position 73 (G73‐>A73) that prevents aminoacylation of Sec tRNA, or the 3′terminal base (A76‐>G76) that is aminoacylated poorly, and compared the rate of synthesis of Um34 following microinjection into Xenopus oocytes of wild type and mutant Sec tRNAs. Um34 was synthesized on each tRNA, but interestingly, the mutant isoforms appeared to add Um34 faster. Thus, the adaptor for SRSPs expression can mature independent of its aminoacylation status.
Serotomide and safflomide modulate forskolin‐stimulated cAMP formation via 5‐HT1 receptor
Authors: Park Jae B
Published: June 2021

Serotomide (N‐caffeoylserotonin) and safflomide (N‐caffeoyltryptamine) are serotonin‐ derived phenylpropenoid amides found in plants (e.g., Coffea canephora and Carthamus tinctorius). Serotomide and safflomide were investigated to determine their effects on serotonin receptor 5‐HT1 in renal epithelial (OK) cells. At the concentration of 10 μM, serotomide was able to inhibit forskolin‐stimulated cAMP formation in the OK cells by 31% (P < 0.011). The inhibition was repressed by Nan‐190 and spiperone (5‐HT1 antagonists), suggesting that serotomide suppresses cAMP formation via binding to 5‐HT1 receptors on the OK cells. Safflomide did not inhibit forskolin‐stimulated cAMP formation at the same concentration (10 μM), but repressed the inhibition of forskolin‐stimulated cAMP by serotonin agonists (e.g., serotonin and 8‐OH‐DPAT) by 33 % (P < 0.012), suggesting that safflomide may block 5‐HT1 receptors in a similar way to Nan‐190 and spiperone. Taken together, the data indicate that serotomide and safflomide may be potent compounds that respectively act to activate and to block 5‐HT1 receptors on OK cells. Potential actions of serotomide and safflomide in neurological and other dysfunctions are discussed. This study was funded by ARS/USDA.
Resveratrol Phenocopies the Suppressive Effects of Insulin‐like Growth Factor Receptor‐1 siRNA on IGF‐1 Promoted Colon Cancer Cell Proliferation
Authors: Vanamala J , Reddivari L , Tarver C C , Murano P S , Lupton J R , Singh P
Published: June 2021

The insulin‐like growth factor (IGF) system is believed to mediate obesity‐related increase in the risk of colon cancer. IGFs activate the β‐catenin pathway, which plays a central role in the growth of colon cancer cells. Dietary resveratrol exerts inhibitory effects on cancers; however, its molecular mechanisms are not fully elucidated, particularly in relation to obesity‐promoted cancers. We hypothesized that resveratrol suppresses colon cancer cell proliferation by inhibiting activation of IGF receptor‐1 (IGF‐1R) in response to IGFs (elevated during obesity). IGF‐1 (10 nM) promoted proliferation of colon cancer cells, in vitro, as expected. Resveratrol (50–150 μM) or IGF‐1R siRNA (100 nM) inhibited (p < 0.05) proliferation of colon cancer cells in response to both exogenous and autocrine IGFs. The relative levels of IGF‐1R were reduced (p < 0.05) in cells treated with either resveratrol or IGF‐1R siRNA, in association with a loss (p < 0.05) in Akt activation, GSK3β phosphorylation, and cyclin D1 levels. These results suggest that resveratrol effectively phenocopies the inhibitory properties of IGF‐1R siRNA and may thus be a potent chemopreventive/therapeutic agent against obesity‐associated increase in colon cancer risk. We will examine this possibility in future studies using in vivo models. Funded by TAES and AICR 058094.
Primary roles for human neutrophil Fc receptors in the initiation of nephrotoxic glomerulonephritis
Authors: Tsuboi Naotake , Mayadas Tanya
Published: June 2021

Antibody‐antigen complexes promote inflammatory and autoimmune diseases. Mice deficient in γ‐chain of Fc‐receptors are protected in antibody mediated glomerulonephritis and lupus nephritis and FcR‐bearing macrophages have been implicated as the primary initiators of disease. However, how well studies of diseases mediated by murine receptors accurately reflect human inflammation remains unclear. Here we show that neutrophil selective transgenic expression of the two uniquely human FcγRIIA and FcγRIIIB in γ‐chain deficient mice (IIA+IIIBtg/γ−/−) was sufficient to promote progressive glomerulonephritis associated with glomerular neutrophil accumulation, glomerulosclerosis, macrophage and T cell accumulation, renal failure and mortality. These responses far exceeded those in wild‐type mice despite similar expression levels of activating receptors suggesting that the human and mouse FcγRs are not functionally equivalent. Both FcγRIIIB and FcγRIIA promoted robust neutrophil recruitment while FcγRIIA alone mediated tissue injury. The latter was consistent with the robust adhesion dependent H2O2 production in IIAtg/γ−/− and the lack there of in IIIBtg/γ−/ −. Thus, human neutrophil FcγRs serve as molecular links between immune complexes and renal injury suggesting a new paradigm for FcγRs on neutrophils in the initiation of antibody mediated‐glomerulonephritis.

Funded by HL065095, AR050800 and Arthritis Foundation Postdoctoral Fellowship.