bluuebunny/crossref_metadata_2025 huggingface dataset:
MLCP activation limits oscillatory pulse pressure‐induced rise in myogenic tone of isolated mouse cerebral arteries in the absence of NO
Authors: Bolduc Virginie , Colinas Olaia , Cole William C , Thorin Eric
Published: June 2021

In response to a static luminal pressure (SP), cerebral resistance arteries develop myogenic tone (MT) in vitro that is magnified by eNOS inhibition. The natural stimulus of MT, however, is the oscillatory pulse pressure (PP) induced by the cardiac cycle. We studied the impact of a PP (550 bpm) of 30 mm Hg on the MT of pressurized (60 mm Hg) mouse cerebral arteries (~120 μm diameter, n=7 per group) before and after eNOS inhibition. MT was magnified by PP compared to SP (14±2 vs. 6±3% reduction in diameter, p<0.05). As expected, inhibition of eNOS with L‐NNA (10 μM) increased MT in SP from 6±3 to 29±4% (p<0.05); PP, however, did not further increase MT (14±2 and 18±3%, respectively). MT is limited by the myosin light chain phosphatase (MLCP); phosphorylation of its MYPT1 subunit on Thr850 inhibits its activity and was unchanged between SP and PP. In contrast, eNOS inhibition led to a decrease in P‐MYPT1 by PP (−41%, p<0.05), while it was unchanged during SP (+12%, p>0.05). In conclusion, MT is maintained constant after eNOS inhibition under physiological conditions of oscillatory PP through a rise in MLCP activity. Supported by CIHR MOP 89733.
Acute alcohol effects on cardiac pro‐ and anti‐apoptotic pathways in the perfused adult rat heart
Authors: Walker Robin Kevina , Cousins Valerie , Haddad George E
Published: June 2021

Cardiovascular disease is among the major causes for increased morbidity and mortality rates. High alcohol consumption may lead to cardiomyopathy, cardiac arrhythmias, and a suite of other disorders (Movva R, Int J Cardiol, 2012). Previous studies in our lab have linked the AKT/PI3K pathway to cardiovascular disease. We also believe increased oxidative stress is a result of high alcohol consumption in cardiovascular disease. In this study we investigate the role of key genes (AKT, super oxide dismutase (SOD‐3), and Caspase‐9) that contribute to cardiac myopathy with acute (30min) alcohol retrograde coronary perfusion of the heart. Our results demonstrate that AKT expression is diminished with acute alcohol exposure which is consistent with the moderate alcohol; however, high alcohol treatment has increased expression over low and moderate alcohol exposure. The Capsase‐9 expression was decreased with high alcohol treatment while low and moderate alcohol had little effect on expression as compared to control. High alcohol treatment diminished SOD‐3 gene expression but low and moderate alcohol treatment had no effect on expression as compared to control. These results suggest that acute high alcohol consumption (e.g., binge drinking) can lead to increased oxidative stress, and malfunction of antioxidant proteins in cardiomyopathy. The data implies that the increased cell death in alcoholic cardiomyopathy may not be dependent on the Bcl‐xL pathway and downstream Caspase‐9. It is important to note that the data was collected from acute studies which may be different with chronic studies.
The effect of global hypo‐methylation in the cellular commitment and differentiation of cells derived from human neural stem cells
Authors: Alvarez Derry , Martinez Aglee , Monterrubio Jose , Rosenthal Joshua , Barreto Jennifer , Roig-Lopez Jose L.
Published: June 2021

Human Neural Stem Cells (hNSC) from fetal tissue was used to study how in vitro demethylation influences the cellular commitment and differentiation between neural or glial lineages. First we verify that neuronal like cells derived from the hNSC show typical electrophysiology profile of a neuron. Second after differentiation of the hNSC derived cells were label with glial (GFAP) and neuronal marker (Beta tubulin III) to score the lineage. Results shows that neurons derived from hNSC have ionic currents characteristic of physiological bona fide neurons. Second after treatment with hypo‐methylating agent 5‐Aza‐Deoxycytidine significantly increase % of neurons derives from hNSCs. No statistically difference is found for GFAP, however there is a tendency to show differences number of label cells between control and experimental conditions. Others markers such O2 and Glutamine synthetase was used to verify lineage commitment. Ongoing experiments will increase the number of replicates for the differentiation protocol. The results from these experiments will help to predict changes in commitment and differentiation of cells with a potential for replacement therapy after exposure to chemicals that can alter epigenetic state. This project was supported by grants from the NCRR (5P20RR016470‐12), NIGMS (8P20GM103475‐12) and MBRS‐RISE at UNE (2R25GM066250‐05A1) from NIH.
FOXO1 regulation of luteinizing hormone beta transcription
Authors: Skarra Danalea , Arriola David J. , Thackray Varykina
Published: June 2021

Luteinizing hormone (LH) is required for mammalian fertility and is produced exclusively by gonadotrope cells within the anterior pituitary. Insulin stimulation of gonadotropes induces LH production, although the mechanisms are unknown. One candidate that may transduce insulin signaling into functional changes in Lhb synthesis is the Forkhead box‐O (FOXO) family of transcription factors. In response to insulin, FOXOs are negatively regulated by Akt phosphorylation, which results in their export from the nucleus to the cytoplasm, thereby limiting their nuclear activity. We have identified FOXO1 expression in murine pituitary gonadotropes and in LbT2 cells, an immortalized gonadotrope cell line. Here, we investigate insulin regulation FOXO1 and the ability of FOXO1 to alter Lhb synthesis. Western analyses identified phosphorylation changes in FOXO1 in response to insulin, which correlated with FOXO1 cytoplasmic localization using immunofluorescence imaging. By luciferase assay, we determined that FOXO1 overexpression in LbT2 cells inhibits basal and gonadotropin‐releasing hormone induced Lhb transcription. Our results indicate that FOXO1 is regulated by insulin in gonadotropes and FOXO1 inhibits Lhb synthesis. In summary, our findings identify a novel mechanism for Lhb gene regulation.

Support: T31 HD720329, R01 HD067448, K01 DK080467, P30 DK063491, and U54 HD012303.
The effect of chromosome substitution on endurance exercise capacity in inbred and F2 generation mice
Authors: Courtney Sean Michael , Massett Michael P
Published: June 2021

Previous research identified a region on chromosome 14 as an important regulator of endurance exercise capacity. The aim of this study was to identify the effect of chromosome substitution on endurance exercise capacity in inbred and hybrid mice. Mice from a chromosome substitution strain (CSS) derived from A/J (A) and C57Bl/6J (B6), denoted as B6.A14 were used to assess the contribution of chromosome 14 to endurance exercise capacity. To further investigate the heritability of the endurance exercise phenotype, 155 B6 x B6.A14 F2 mice were generated from a cross between B6 and B6.A14 CSS mice. All mice performed a graded exercise test to exhaustion to determine endurance exercise capacity. Endurance exercise capacity was expressed as time (min). B6 x B6.A14 F2 mice had an exercise capacity of 30.8 ± 0.1 minutes, which is significantly higher than parental B6.A14 CSS mice (28.5 ± 0.2 min) and similar to parental B6 mice (31.0 ± 0.1 min). Although the cohort mean is similar between F2 and B6 mice, the range of responses in the B6 x B6.A14 F2 mice (27.3 – 34.5 min) exceeded the range of the parental inbred strains. The range of responses in the B6 x B6.A14 F2 mice suggests that this population of mice is diverse in both phenotype and genotype and would be amenable to a chromosome‐wide scan for endurance exercise‐related genes.

Supported by NIH grant HL085918 to MPM
Low‐resolution structure of HIV‐1 genomic RNA regions obtained by small angle x‐ray scattering analysis
Authors: Olson Erik David , Jones Christopher P , Musier-Forsyth Karin
Published: June 2021

The 5′ untranslated region (5′UTR) of the human immunodeficiency virus type 1 (HIV‐1) RNA genome (vRNA) controls many aspects of virus replication, including vRNA dimerization, tRNA priming, and virus assembly. While the secondary structure of the 5′UTR is well annotated, a model for the tertiary structure of the entire 5′UTR remains elusive. In this work, using a divide‐and‐conquer approach, various portions of the 5′ UTR of the vRNA were examined using small angle x‐ray scattering (SAXS). SAXS requires homogeneous sample preparation, which was accomplished through a combination of optimizing construct design, buffer/refolding optimization, size‐exclusion chromatography, and point mutations. As expected, primer tRNALys3 adopts a compact L‐shaped fold, consistent with crystallographic data. The 2‐helix 104‐nt TAR/polyA RNA exists in an elongated conformation, with apparently co‐axially stacked helices. In contrast, the 95‐nt 3‐helix Psi RNA packaging signal adopts a less compact conformation, likely due to the flexible linker connecting stemloops SL2 and SL3. These preliminary models represent the first step toward our goal of building a low‐resolution model of the entire 5′UTR in the absence and presence of interacting partners. This research was supported by Ohio State University and by a fellowship to CPJ from the OSU Center for RNA Biology.
Meal and snack patterns associated with biomarkers of glucose metabolism, diet quality and weight status in US adults
Authors: Shin Dayeon , Song Su Jin , Paik Hee‐Young , Song Won O.
Published: June 2021

Epidemiological investigations established the associations between insulin resistance (IR), anthropometry and certain aspects of dietary intake. Our aim was to examine if meal and snack patterns are associated with biomarkers of glucose metabolism (fasting glucose and insulin, HbA1c, and HOMA‐IR), diet quality (HEI 2005), and weight (wt) status (BMI). Dietary recall data of 2175 men and 2191 women from the 2007–8 NHANES were subjected to factor analysis by gender by 19 food groups of MyPyramid Equivalents Database 2.0. Accounting for the complex survey design and sample wt, SAS v. 9.3 was used for logistic regressions. Of four meal patterns in men, two were associated (+) and one (−) with diet quality; the fourth(“grain and dairy”) was related with HbA1c and HOMA‐IR (p<0.05). Two meal patterns of women were associated (+) and one (−) with diet quality; the fourth (“fruit and alcohol”) indicated (+) for HbA1c, diet quality, and wt status. Snack patterns alone reflected total diet quality (p<0.05), but not biomarkers of glucose metabolism nor wt status in both gender. Meal and/or snack patterns of US adults are associated differentially with biomarkers of glucose metabolism, diet quality, and wt status.

Grant Funding Source: None
2010–2011 USDA fresh fruit and vegetable program (FFVP) improved Indiana elementary students’ consumption of fruit
Authors: Huang Li , Lin Yi‐Chun , Foland Elizabeth , Bai Yeon , Liu Yibin , Fly Alyce D.
Published: June 2021

U.S. children's fruit (F) and vegetable (V) consumption falls below recommendations. FFVP is a national program that addresses this problem. The FFVP 2010–11 in Indiana was evaluated to examine changes in students’ F and V intake using a quasi‐experimental, pre‐posttest design at 38 intervention and 8 control elementary schools with a 60 item questionnaire (4th–6th grades, n=11 138). Food intake frequencies (times/day) were grouped into 9 categories: F (in school, outside of school, total, 3 items each), V (in school, outside of school, total, 7 items each), and low nutrient density foods (LN) (in school, outside of school, total, 6 items each). The difference in daily F and V intake frequency between intervention and control overtime were determined with multilevel negative binomial regression using grade, gender, race, ethnicity and free/reduced lunch rate as covariates, post‐hoc tests were used to examine significant pairwise comparisons. Significant program by time interactions were found for total F (p=0.031) and total F in school (p=0.012) but not outside school (p=0.149). FFVP increased intake frequency of TF in intervention (p=0.038) but not in control (p=0.075) students. Total V and total LN food intake frequency did not change (FFVP × time, p=0.58 and 0.55, respectively). These data indicated FFVP may improve children's health. Further steps are needed to improve V consumption and reduce intake of LN food.
Standardized chungkookjang, short‐term fermented soybeans with Bacillus lichemiformis, improves cognitive function and insulin resistance in β‐amyloid‐infused diabetic rats
Authors: Park Sunmin , Kang Suna , Kim Hyun Jin , Kwon Dae Young , Daily James
Published: June 2021

Both traditionally made chungkookjang (TFC, naturally fermented) and standardized chungkookjang (SFC, fermented by inoculating Bacillus lichemifomis for 48 h) have potent antidiabetic activities. It is interesting to investigate whether both chungkookjangs can protect against cognitive dysfunction in β‐amyloid‐infused diabetic rats since Alzheimer's disease is known as “brain diabetes”. Male rats with type 2 diabetes and an Alzheimer's‐like brain disorder induced by partial pancreatectomy and ICV β‐amyloid (25–35) infusion were fed either control diet, 10% cooked soybeans (CSB), 10% TFC, or SFC in a high fat diet for 8 weeks. Px rats infused β‐amyloid (35–25) as a normal‐control group. SFC, CSB and TFC improved cognitive function as measured by a water maze test in an ascending order. β‐amyloid deposition was much higher in the control rats than the normal‐control rats and SFC decreased its accumulation. Rats in the control group had lower whole body glucose infusion rates and increased hepatic glucose output at hyperinsulinemic state during euglycemic hyperinsulinemic clamp which was normalized by SFC. Interestingly, insulin secretion, especially at the second phase during hyperglycemic clamp, was higher in control rats in comparison to the normal control group while CSB, TFC, SFC lowered it. However, β‐cell mass was restored by SFC in β‐amyloid‐infused rats with reduced β‐cell mass by increased β‐cell apoptosis. In conclusion, cognitive dysfunction exacerbated insulin resistance and decreased β‐cell mass and SFC alleviated their exacerbation in β‐amyloid infused diabetic rats. The SFC‐improved memory loss may be due to increased genistein, daidzein, poly‐gamma‐glutamate, small peptides and other components in SFC.
Padiococcus Acidilactici (PA) Isolated from traditional Makgeolli inhibits lipid accumulation in 3T3‐L1 adipocyte and obesity in C57BL/6J mice fed a high‐fat diet
Authors: Moon Yeon‐Jeong , Baik Sang‐Ho , Cha Youn‐Soo
Published: June 2021

Korean traditional alcoholic beverage ‘Makgeolli’, is a type of rice beer typically prepared using steamed rice and nuruk. This study investigated the anti‐obesity effect of PA (Padiococcus Acidilactici) extract on in‐vitro and in‐vivo. PA extract treatment decreased the triglyceride (TG) level and the Oil red O staining in cultured 3T3‐ L1 adipocytes. For in‐vivo study C57BL/6J male mice, were randomly divided into ND (ND normal group), HD (HD high fat diet group), HDA (HD plus reference strain) and HDB (HD plus PA) groups. The lyophilized strains were orally administrated with a final concentration of 1.25 × 109 cfu/g for 12 weeks. HDB group showed lower final bodyweight, adipose tissue weight and serum lipid profile. Also, the activity of fatty acid synthase (FAS), a key enzyme of fatty acid synthesis, was markedly suppressed in the HDB, as compared to the HD group. We suggest PA extract may exert an anti‐obesity effect in C57BL/6J mice by enhancing the lipid metabolism (This research was supported by the Globalization of Korean Foods R&D program, funded by the Ministry of Food, Agriculture, Forestry and Fisheries, Republic of Korea).
Systematic review of Wild Rice: an ancient grain with modern benefits
Authors: Timm Derek Allen , Slavin Joanne
Published: June 2021

Wild rice (Zizania sp.) grows in shallow waters of North America and is distinct from white rice (Oryza sp.). The recent trend toward gluten free foods and ancient grains presents an opportunity for increased interest in the health benefits of wild rice. Wild rice is classified as a whole grain and typically contains 74% starch, 6.8% dietary fiber, 14% protein, and 0.7–0.9% of lipids. Wild rice is a good source of dietary fiber and has a good protein efficiency ratio (1.7), which is high for a grain. The lipid content is low; however most of the lipids are Ω‐6 (35–37%) and Ω‐3 (20–31%) fatty acids with a very favorable ratio of (1.1). In addition to macronutrient content, wild rice contains phytosterols levels several times higher than other grains. Furthermore, the antioxidant activity of wild rice is between 10–15 times higher than white rice. A new study of wild rice added to a high fat/cholesterol diet showed significant attenuation of serum free fatty acids, total cholesterol, and triglyceride levels while maintaining higher high density lipoprotein levels compared to control. The exact explanation of the protective mechanism of wild rice is uncertain, but the dietary fiber, phytosterol, or antioxidant capacity of wild rice may be the reason for this. Current interest in ancient grains and gluten‐free diets, as well as antioxidants and phytochemicals, makes wild rice an attractive grain addition to the diet.

Grant Funding Source: MN Wild Rice Council
Discriminative stimulus effects of the synthetic cannabinoid JWH‐018 in rhesus monkeys
Authors: Rodriguez Jesse S , McMahon Lance R
Published: June 2021

JWH‐018 produces greater maximal stimulation of G‐proteins at cannabinoid CB1 receptors than Δ9‐tetrahydrocannabinol (Δ9‐ THC); whether this results in a difference in maximum behavioral effects is not clear. The aim of this study was to establish a behavioral assay requiring high agonist efficacy by training a relatively large dose (0.1 mg/kg i.v.) of JWH‐018 as a discriminative stimulus in rhesus monkeys (n=5). The potency (ED50 value) of JWH‐018 to produce discriminative stimulus effects and to decrease response rate was 0.01 mg/kg and 0.06 mg/kg, respectively. Other cannabinoids substituted for the JWH‐ 018 discriminative stimulus with the following rank order potency (ED50 in mg/kg): CP‐55940 (0.005) = JWH‐018 >; Δ9‐THC (0.04) = WIN‐55212–2 (0.04) = JWH‐073 (0.07). The CB1 receptor‐selective antagonist rimonabant surmountably antagonized the discriminative stimulus effects of JWH‐018 and Δ9‐THC; Schild and single‐dose apparent affinity analyses yielded pA2 and pKB values that were not significantly different from each other. These data strongly suggest that the same receptor type mediates the discriminative effects of JWH‐018 and Δ9‐THC. However, the reported difference in efficacy at this receptor type does not result in different levels of discriminative stimulus effect in a situation where such a difference is expected.
Epac2 and PLCε contribute to the inflammatory response to cigarette smoke in vivo
Authors: Oldenburger Anouk , Bos Sophie , Smrcka Alan , Meurs Herman , Maarsingh Harm , Schmidt Martina
Published: June 2021

β2‐Agonists increase cellular cAMP by binding to Gs‐coupled receptors, leading to activation of PKA as well as the cAMP‐regulated exchange factors Epac1 and ‐2, the latter known to signal to PLCε. We showed recently that Epac activation reduces cigarette smoke (CS) extract‐induced IL‐8 release from airway smooth muscle cells by inhibiting Nf‐κB. We now studied the role of the Epac signalosome in CS‐induced airway inflammation in vivo. Mice were pretreated with vehicle, the β2‐agonist fenoterol or the Epac1 and ‐2 activator 8‐pCPT and exposed to CS or filtered air for 5 days. In bronchoalveolar lavage fluid of CS exposed wild‐type mice, total inflammatory cells, macrophages and neutrophils were all increased. Treatment with fenoterol or 8‐pCPT did not alter the inflammatory response induced by CS, although fenoterol slightly increased total inflammatory cells in air exposed mice. Compared to wild‐types, the CS‐induced increase in total inflammatory cells, macrophages and neutrophils were reduced in Epac2−/− mice, whereas specifically the neutrophils were reduced in PLCε−/− mice. In conclusion, Epac2 enhances CS‐induced airway inflammation in vivo presumably by activating PLCε. (NAF grant 3.2.09.034)
NAD(P)H autofluorescence induction by Compound C in rat carotid chemoreceptor cells
Authors: Kim Insook , Fite Jaclyn Lasha , Kim Donghee , Donnelly David F. , Carroll John L.
Published: June 2021

Compound C (CC) has been known as an inhibitor of AMP‐activated kinase (AMPK). Recently other roles of CC have been reported. CC inhibits hypoxia dependent HIF‐1α activation independent of AMPK but dependent on mitochondrial electron transport chain (ETC) (Emerling 07) and CC functions as an inhibitor of complex I of ETC (Chua 11). In addition, we have found that CC (4μM) produces ~10 fold increase in NAD(P)H autofluorescence (AutoFlu) as compared to hypoxia or cyanide in rat carotid body (CB) glomus cells. Autoflu was detected with 360/50 excitation(x) and 460/50 emission(m) filters. The effect of CC on the hypoxic [Ca2+]i response was observed by using 340&380x with 540/50m filter for fura‐2. Rat CB glomus cells were identified with rhodamine‐tagged peanut agglutinin.

Results

1) AutoFlu produced by CC is dose dependent, reaching a plateau at 0.04μM in both CB type cells. 2) The magnitude of AutoFlu induced by CC was greater in glomus cells compared to non‐glomus cells. 3) AutoFlu induced by 0.4μM CC was reduced ~50% by hypoxia but only < 10% by 20mM KCl. 4) CC did not increase calcium despite the increase in AutoFlu. 5) 0.4μM CC reduced the hypoxia‐induced [Ca2+]i increase by ~50%, but did not block 20mM KCl‐induced [Ca2+]i increase. These preliminary results suggest that CC induced significant NAD(P)H AutoFlu, which may significantly affect the measured [Ca2+]i response to hypoxia in glomus cells. (Funded by NIH HL54621)
Caprylic triglyceride as a novel therapeutic approach to effectively improve the performance and attenuate the symptoms due to the motor neuron loss in ALS disease
Authors: Pasinetti Giulio Maria
Published: June 2021

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder causing progressive muscle weakness and ultimately death. ALS patients have impaired energy balance linked to lower activity of mitochondrial electron transport chain enzymes in ALS spinal cord, suggesting that improving mitochondrial function may present a therapeutic approach for ALS. When fed a ketogenic diet, the G93A ALS mouse shows significant increase in serum ketones as well as significantly slower progression of weakness and lower mortality rate. In this study, we treated SOD1‐G93A mice with caprylic triglyceride, a medium chain triglyceride that is metabolized into ketone bodies and can serve as an alternate energy substrate for neuronal metabolism. Treatment with caprylic triglyceride attenuated progression of weakness and protected spinal cord motor neuron loss in the transgenic mice, significantly improving their performance even though there was no significant benefit regarding survival. Our results demonstrate that caprylic triglyceride alleviates ALS‐type motor impairment through restoration of energy metabolism, indicating feasibility of using it as a treatment with a high impact on quality of life of ALS patients.
Link between the prorenin/renin receptor and glucose homeostasis in mice fed a high‐fat/high‐carbohydrate diet
Authors: Shamansurova Zulaykho , Tan Paul , Bisotto Sandra , Michel Catherine , Nguyen Thi M‐D , Shiller Peter W , Gutkowska Jolanta , Lavoie Julie L
Published: June 2021

Objective

Elucidate the role of the (P)RR in glucose homeostasis in mice fed high‐fat/high‐carbohydrate diet (HF/HC).

Material and methods

12 week‐old C57BL/6 male mice were placed on a HF/HC or normal diet (ND) and were treated with either (P)RR blocker [(P)RR‐B] or saline for 10 weeks. Body weight was assessed weekly. At the end of treatment, mice were euthanized and fat tissues from subcutaneous (SCF), peri‐gonadal (PGF), peri‐renal (PRF) localisation were separated, weighed and flash frozen. Blood was collected and plasma was flash frozen for future measurement of glucose, insulin and triglycerides. Gene expression for glucose transporters GLUT1 and GLUT4 were assessed by real‐time PCR.

Results

(P)RR‐B administration lowered weight gain in the HF/HC fed mice, whereas food intake didn't differ between the groups. Increases produced by the HF/HC in circulating glucose, insulin and triglycerides were significantly decreased. Insulin sensitivity measured by glucose‐insulin ratio was also improved by the (P)RR‐B. Altered GLUT1 and GLUT4 mRNA expression in mice on a HF/HC was improved by the (P)RR‐B, in line with the circulating metabolites data.

Conclusion

(P)RR‐B administration seems to improve glucose homeostasis, insulin sensitivity and GLUTs gene expression profile in mice fed a HF/HC.

Research supported by CIHR, HFSC, CDA and Merck Frosst Canada
Long term hypoxia impairs ryanodine receptor function and regulation by cyclic nucleotides in immature and mature pulmonary arterial myocytes
Authors: Hough Steven , Rubalcava Monica , Manjunath Chetas , Blood Quintin , Longo Lawrence D , Wilson Sean M
Published: June 2021

Calcium sparks are due to ryanodine receptor (RyR) activation and are intimate to local and global Ca2+ signals in pulmonary arterial (PA) myocytes. One possibility is that impaired RyR activity contributes to altered PA reactivity following long‐term hypoxia (LTH) in developing arteries. Cyclic nucleotides (CNs), including cAMP and cGMP, can augment localized Ca2+ sparks and global Ca2+ waves due to RyR activation. Yet, the influence of LTH on CN regulation of RyRs of PA myocytes is unknown. These studies tested the hypothesis that cAMP and cGMP increase Ca2+ sparks and waves in PA myocytes and that LTH moderates this effect. Spark and wave activity was evaluated by visual analysis of line‐scan confocal recordings of PA myocytes from term‐fetal, ~ 10 day old, or adult sheep that lived at low (720 m) or at high altitude (3,200 m) for >;100 days. Similar percentages of term‐fetal, newborn, and adult myocytes had sparks and waves before CN treatment. LTH blunted spark activity and impaired CN reduction of sparks in fetus and adult. However, CNs inhibited sparks of LTH but not normoxic newborns. Ca2+ wave activity and regulatory influences by CNs were diminished by LTH in all age groups. The data suggest the effects of LTH on CN dependent regulation of RyRs may contribute to PA dysfunction and are highly dependent on the developmental age examined. (NSF MRI923559, NIH R03HD69746, P01HD31226, R01HD3807, LLU basic sciences fellowship)
Perivascular adipose tissue impairs H2O2‐mediated vasodilation in the coronary circulation
Authors: Owen Meredith Kohr , Krenzke Rebecca , Dick Gregory M , Tune Johnathan David
Published: June 2021

This investigation examined the effects of perivascular adipose tissue (PVAT) on vasodilatory mechanisms in the coronary circulation. Isometric tension studies were performed on isolated coronary arteries from lean and obese Ossabaw swine. After a 30 min incubation period with 0.3 g PVAT, arteries were constricted with U46619 (1 μM) or PGF2α (10 μM) and responses to H2O2 (1 μM–1 mM) were examined. Compared to untreated‐control arteries, addition of coronary PVAT markedly attenuated relaxation to 1 mM H2O2 in lean swine (78 ± 9% vs. 17 ± 11%). This effect of PVAT was similar in intact and denuded arteries or in arteries pre‐contracted with PGF2α or U46619. However, pre‐constriction with KCl (60 mM) abolished relaxation to H2O2, implicating K+ channels in the vasodilatory response. Inhibition of K+ channels with tetraethylammonium (3 mM), penitrem A (1 μM), barium chloride (30 μM) or glibenclamide (1 μM) did not prevent relaxation to H2O2. In contrast, 4‐aminopyridine (10 mM) prevented relaxation to H2O2 to a similar degree as PVAT. Coronary arteries from obese swine demonstrated similar relaxation to H2O2, however, PVAT did not attenuate this response (98 ± 2% vs. 82 ± 8%, respectively). These findings indicate that coronary PVAT releases factors that diminish coronary vascular smooth muscle vasodilation via inhibition of K+ channels, which is absent in obesity. Support 5T32DK064466–09 and HL092245.
KEY ROLE OF TRPC3 CHANNELS IN CALCIUM HOMEOSTASIS ALTERATION RELATED TO NO DEFICIENCY IN AGED PORCINE ENDOTHELIAL CELLS
Authors: Banquet S , Bourguignon MP , Garry A , Royere E , Gosgnach W , Thollon C , Villeneuve N , Vilaine JP
Published: June 2021

NO controls the activity of canonical transient receptor potential channel (TRPC) in endothelial cells. Our group already provided evidences that reduced NO bioavailability was directly linked to increased basal cytosolic Ca2+ in a cellular model of endothelial dysfunction. The aim of this study was to evaluate the role of TRPC3 channels in Ca2+ homeostasis of aortic porcine endothelial cells (PAEC). In PAEC at P1, producing NO, specific inhibition of TRPC3 by Pyr3 induced a concentration‐dependent decrease in Ca2+ entry as shown by reduced basal [Ca2+]i and decreased Ca2+ peak amplitude in response to bradykinin. Similar modifications were observed in the presence of Orai1 inhibitor or in a Ca2+‐free buffer while OAG, a specific TRPC3 activator, didn't modify Ca2+ homeostasis. In dysfunctional PAEC at P4, model of endothelial dysfunction associated with basal increased of [Ca2+]i, Pyr3 normalized the basal [Ca2+]i in a concentration‐dependent manner while the activation of TRPC3 by OAG over increased it. These results indicate that calcium entry by TRPC3 and Orai1 channels is involved in regulation of cytosolic calcium in functional endothelial cells. In dysfunctional endothelial cells with limited NO production, an increased expression or activity of TRPC3 may participate to the abnormal Ca2+ homeostasis
Comparison of neurohumoral responses to lower body negative pressure and hemorrhage in baboons
Authors: Shade Robert E , Hinojosa‐Laborde Carmen , Muniz Gary , Bauer Cassondra , Convertino Victor
Published: June 2021

Lower body negative pressure (LBNP), an experimental model of hemorrhage (Hem), causes shifts in blood volume to the legs and elicits central hypovolemia. This study defined the levels of LBNP which equate to actual Hem. Male baboons (n=10) were sedated and instrumented for measurement of arterial pressure (AP), pulse pressure (PP), and central venous pressure (CVP). Blood was removed in four steps (7 min/step): 6.25%, 12.5%, 18.75%, and 25% of total blood volume. Shed blood was returned and the animals were allowed to recover. Four weeks after the Hem protocol, the same animals were subjected to four levels of LBNP which elicited equivalent changes in PP and/or CVP observed during Hem. Blood samples were collected before Hem or LBNP( baseline, B), after the second Hem step or LBNP (midpoint, MP), and after the fourth Hem step or LBNP (pre‐syncope, PS). Blood samples were analyzed for plasma renin activity (PRA), catecholamines ( Nepi, Epi) and vasopressin (AVP). The results for PRA (mean±sem) are shown below:

PRA (ng AngI/ml h)

Study
B
MP
SP

Hem
2.8±0.7
3.9±0.9
7.9±1.8

LBNP
4.1±1.0
5.4±1.7
9.9±2.7

Repeated measures ANOVA of the log transformed PRA data indicated that PRA was significantly increased relative to B for the MP and PS samples and that PS was higher than MP for both the Hem and LBNP studies. However, there was no significant difference between Hem and LBNP for PRA at the B, MP and SP time points. Similar results were obtained for plasma Nepi, Epi and AVP. We conclude that LBNP levels can be used as surrogates for actual blood loss. This study supports the validity of LBNP as a human model of central hypovolemia associated with Hem. Funding Source: U.S. Army MRMC Combat Casualty Care Research Program.
Calcium regulates Na+:K+:2Cl− cotransporter function in Malpighian tubules of Rhodnius prolixus
Authors: Gioino Paula , Ianowski Juan P
Published: June 2021

Rhodnius prolixus is a haematophagous insect, vector of Chagas’ disease. During each blood meal the insect ingests large volumes of blood that may be up to 10 times its unfed weight. The excess of water and ion ingested is expelled thanks to a rapid post‐prandial diuresis triggered by the diuretic hormone serotonin, that stimulate the Malpighian (renal) tubules (MT). Until now, serotonin was believed to stimulate MTs via the second messenger cAMP and Ca2+ was considered to play no significant role. Here we tested the role of Ca2+ on serotonin‐stimulated MTs. Our results show that serotonin stimulation triggered Ca2+ waves in MTs. In addition, incubating tubules in Ca2+‐free saline containing BAPTA‐AM (3 μM) reduced fluid transport by 50%, decreased Na+ flux by 70% and decreased pH of the secreted fluid from 7.10±0.05 to 6.20±0.21. Measurements of transepithelial potential suggest a reduced activity of the basolateral Na+:K+:2Cl− cotransporter (NKCC) when chelating Ca2+. The results indicate that Ca2+ is important for normal functioning of the tubules. We hypothesize that Ca2+ may be involved in NKCC regulation. Disruption of Ca2+ signalling may alter NKCC activity by changing its phosphorylation state or its insertion to the membrane. Funded by a Natural Sciences and Engineering Research Council of Canada grant to JPI.
Building better animal models of disease through integration of multiple disease phenotypes
Authors: Dwinell Melinda Ryan , Nigam Rajni , Liu Weisong , Smith Jennifer R , De Pons Jeff , Shimoyama Mary
Published: June 2021

Animal models of disease are used to test hypotheses of disease mechanisms as well as for therapeutic development. Effective animal models must sufficiently reflect human disease and should be classified carefully as disease models. Integration of genomic and phenotypic data provides the foundation to study the genetic and environmental influences on the risk, onset and progression of disease. PhenoMiner, at the Rat Genome Database (http://rgd.mcw.edu), is an interactive data‐mining tool to compare clinical measurements used as indices of disease across multiple rat strains, measured under a variety of conditions. Data are integrated through the use of four ontologies (rat strain, clinical measurement, measurement method, experimental conditions). PhenoMiner provides a solid foundation to distinguish phenotypes by strains and strains by phenotypes. Currently more than 42,000 records can be viewed graphically or in tabular format, allowing easy comparison across strains. Graphical summaries (bar charts and heat maps) provide a rapid method to identify strains with abnormal phenotypes or groups of phenotypes that correspond to disease traits. In addition to providing phenotype driven model identification, PhenoMiner generates standard ranges for clinical measurements under control conditions to be used to identify strains with potential disease traits.

Supported by R01HL094271 and R01HL064541.
Altered Glutamate Receptor Expression Following Carotid Body Denervation (CBD) in Goats
Authors: Miller Justin Robert , Neumueller S , Muere C , Olesiak S , Pan L G , Hodges M R , Forster H V
Published: June 2021

After CBD, goats hypoventilate and their CO2 sensitivity is reduced for several days followed by a partial recovery (Pan et al, 1998). Even after partial recovery 30 days post‐CBD, histochemical markers demonstrate a down regulation of the serotonergic system (Miller et al, EB 2012); thus, the mechanisms which contribute to the recovery of ventilation following CBD remain unclear. Recently, AMPA‐type glutamate receptors have been implicated with a form of activity‐dependent plasticity suggested to stabilize the output of neuronal networks (homeostatic plasticity) (Turrigiano and Nelson, 2004). Accordingly, to test the hypothesis that goats sacrificed 30 days post‐CBD exhibit altered AMPA receptor expression, we quantified immunolabeled neuronal GluR2 optical density (OD) at sites that contribute to ventilatory control. Relative to a control goat (n=1), in CBD goats (n=3) we observed increased GluR2 OD within the nucleus tractis solitarii (NTS), decreased GluR2 OD within the midline raphé (MR), and no change in the dorsal motor nucleus of the vagus or the hypoglossal motor nucleus. These preliminary findings suggest that increased glutamatergic drive within the NTS is associated with the partial recovery of ventilation post‐CBD and decreased glutamatergic drive within the MR may contribute to the down regulation of the serotonergic system. Supported by NIH 25739 and VA
Oral administration of anti‐sPLA2‐IB antibodies reduces sepsis mortality in murine endotoxemia
Authors: Sand Jordan , Butz Daniel E , Tsao Francis , Cook Mark E
Published: June 2021

Annually 750,000 people develop severe sepsis and 215,000 die at a cost of more than $28 billion. Evidence suggests sepsis progresses when bacteria translocate across a compromised intestinal epithelial barrier. Secretory intestinal phospholipase (sPLA2‐IB) may play an important role in regulation of gutmediated inflammation. The objective of this study was to demonstrate reduced sepsis mortality by oral administration of antibodies against sPLA2‐IB. We made peptide antibodies to ten 8 amino acid sequences of the sPLA2‐IB molecule and tested their efficacy in inhibiting sPLA2 enzyme activity and in reducing endotoxin (LPS) induced TNF production in RAW264.7 macrophages in vitro. Three antibodies inhibited sPLA2 activity and four antibodies decreased LPS induced TNFα by more than 50%. We orally administered these antibodies to female Balb/c mice given 10mg/kg LPS. Mice fed antibody that inhibited TNF (anti‐ TYSYSCSG) had 100% endotoxin‐induced mortality while antibody that inhibited PLA2 activity (anti‐VPYNKEYK) had 0% mortality and mice receiving no oral antibody had 45% mortality. Antibody #6 is so specific that changing the primary sequence target by 4 amino acids in either the amino or carboxyl direction makes it ineffective. In summary, oral administration of an antibody that inhibits sPLA2‐IB enzyme activity prevents sepsis related mortality caused by endotoxemia.

Research supported by a Draper Technology Innovation Fund Grant.
GALLIC ACID MODIFIES THE EXPRESSION OF INTESTINAL MALTASE‐GLUCOAMYLASE mRNA BUT NOT ENZYME ACTIVITY
Authors: Simsek Meric , Hamaker Bruce R , Nichols Buford L , Quezada‐Calvillo Roberto
Published: June 2021

Natural polyphenols modify the expression of mRNA coding for proteins involved in nutritional, metabolic or immune processes. Gallic Acid (GL) inhibits competitively the activities of intestinal Maltase‐Glucoamylase (Mgam) and Sucrase‐Isomaltase (Si). However, it is not clear whether GL also regulates the expression of mRNA coding for Mgam and Si.

Objective

To explore the possibility of GL regulates the expression of mRNA coding for Mgam.

Methods

Small intestinal explants from 8 weeks old BALB/c mice were cultured 24 h in DMEM − 10% FBS media, with or without 0.1 or 0.5 mM GL. Maltase activity, molecular size distribution and quantity of Mgam mRNA were determined in the tissues.

Results

No changes in maltase activity were induced by GL. However, qRT‐PCR showed a increment of Mgam mRNA induced by 0.1 mM GL. Western Blot analysis showed that 0.5 mM GL modified the proportions of the different molecular forms of Mgam, suggesting an effect on their posttranslational processing.

Conclusions

GL modifies the expression of mRNA and the postranslational processing of Mgam. However, these modifications have no effects on the expression of maltase activity derived from Mgam in the intestinal epithelium.

Grant Funding Source: CONACYT, Mex
Biochemical and Physical Correlates of Serum [25(OH)D] and the IGF system in Men
Authors: Darr Rachel L , Savage Kathleen J , Wilding Gregory , Wu Dawei , Millen Amy , Burton Harold , Browne Richard W , Horvath Peter J
Published: June 2021

Serum [25(OH)D] and IGF system components are shown to have relationships with one another and physical performance capabilities. Fasting blood specimens, anthropometric (body mass index [BMI], waist circumference [WC], body fat 7 sites [%BF]) and physical fitness measures (% body weight lifted for one‐repetition maximum bench press [%BP], predicted VO2), were taken from 36 healthy middle‐aged men (mean age 45.8±9.7 years). Serum [25(OH)D] and IGF system measures (Free IGF‐1, IGFBP‐3) were determined by CMIA and ELISA, respectively. Pearson correlation coefficients (r) were estimated. Free IGF‐1 correlated negatively with BMI (p=0.04, r=0.35), but there were no associations with %BF (p=0.08, r=0.3) or WC (p=0.11, r=0.26). Serum [25(OH)D] correlated positively with IGFBP‐3 (p=0.04, r=0.35) and predicted VO2 (p=0.02, r=0.41), but not with %BP (p=0.6, r=0.08). Consequently, individuals with a higher BMI present with lower levels of circulating Free IGF‐1 available to tissues. As Serum 25(OH)‐Vitamin D levels increase, greater levels of IGFBP‐3and a higher aerobic oxygen capacity are present; speculating more IGFBP‐3 present in aerobically inclined individuals.

Grant Funding Source: University Funds
Purified recombinant human β‐carotene 15–15′‐oxygenase 1 (BCO1) cleaves β‐apocarotenals and lycopene
Authors: Sena Carlo dela , Narayanasamy Sureshbabu , Curley Robert W , Harrison Earl H
Published: June 2021

β‐Carotene 15–15′‐oxygenase 1 (BCO1) cleaves provitamin A carotenoids at the 15–15′ bond to produce retinal (vitamin A). This study aimed to elucidate the substrate specificity using pure BCO1, rather than partially purified preparations as used in previous studies. The enzyme was cloned into an expression vector with a C‐terminal His‐tag, expressed in E.coli, and purified using Co2+ ion affinity chromatography. The pure enzyme is soluble in detergent‐free buffer, however greater enzyme activity was observed if Triton X‐100 was present during purification and storage. BCO1 cleaved ß‐carotene, ß‐cryptoxanthin, α‐carotene, ß‐apo‐8′‐ carotenal, ß‐apo‐10′‐carotenal and ß‐apo‐12′‐carotenal to yield retinal. BCO1 cleavage of lycopene yielded acycloretinal. Lycopene and ß‐apo‐12′‐carotenal were previously reported to be unreactive with BCO1. BCO1 did not catalyze cleavage of lutein or zeaxanthin. We have calculated apparent kcat and KM values for each substrate; however, some substrates yielded kinetics that did not fit well with Michaelis‐Menten kinetics. However, among 40 carbon substrates activity was greatest with ß‐carotene, followed by ß‐cryptoxanthin, α‐carotene, and lycopene. For the ß‐apocarotenals activity decreased with decreasing length of the molecule.

(NIH Grant R01‐HL49879 and a grant from the Ohio Agricultural Research Development Center (OARDC)).
Bioinformatics analysis of Mycobacterium tuberculosis‐specific genomic regions to identify immunodominant proteins and peptides
Authors: Mustafa AbuSalim
Published: June 2021

This study was aimed to identify immunodominant proteins and peptides encoded by Mycobacterium tuberculosis‐specific genomic regions of differences (RDs) by using bioinformatics to predict for HLA‐DR binding regions. The analysis of RD proteins and peptides by in silico methods (using computational programs to predict major and HLA‐promiscuous antigenic proteins and peptides) and wet‐lab experiments (using peripheral blood mononuclear cells and sera from TB patients and BCG‐vaccinated healthy subjects to asses antigen‐specific cellular and humoral immune responses in vitro) identified several major antigens and peptides. To evaluate them for in vivo reactivity, the genes of immunodominant antigens were cloned and expressed in DNA vaccine vectors (pUMVC6 and pUMVC7) and mycobacterial hosts, i.e. BCG, M. vaccae and M. smegmatis. Immunizations of mice and guinea‐pigs with the recombinant constructs induced antigen‐specific cellular and humoral immune responses in these animal models of TB. Each of these proteins had several T and B cell epitopes scattered throughout the sequences, which confirmed their strong immunogenicity and appropriateness for diagnostic and vaccine applications.

Supported by Kuwait University Research Administration grants MI01/10 and GM01/01.
ENDOCARDIAL FIBROLASTOSIS IN CHICK MODEL OF HYPOPLASTIC LEFT HEART SYNDROME
Authors: Pesevski Zivorad , Stopkova Tereza , Sedmera David
Published: June 2021

Endocardial fibroelastosis (EFE) is a diffuse thickening of the ventricular endocardium, causing myocardial dysfunction and presenting as unexplained heart failure in infants and children. It is believed to be caused by persistent and increased wall tension in the ventricles. . Its frequent association with hypoplastic left heart syndrome (HLHS) lead us to hypothesize abnormal hemodynamic loading is an important factor in its pathogenesis. We have tested this theory in a chick model of HLHS induced by left atrial ligation (LAL). At ED8 and 12 modifications of myocardial architecture and fibrosis were studied by histology and immunoconfocal microscopy, and the amount of collagen was quantified by image analysis.

Histology with H&E / Alcian Blue staining did not reveal any significant fibrosis in the LAL hearts with the exception of cardiac skeleton and valves. Immunohistochemistry with collagen I antibody clearly showed a thickening of the layer of subendocardial fibrous tissue. Quantification of staining area normalized by endocardial circumference showed a significant increase at ED12, but not ED8 in the LAL group. We conclude that abnormal hemodynamic loading stimulates fibrous production in the subendocardium of the hypoplastic left ventricle. Therefore, EFE in HLHS clearly appears to be a secondary effect of abnormal hemodynamics.

Grant Funding Source: Supported by Ministry of Education, Youth and Sports PRVOUK‐P35/LF1/5, Academy of Sciences RVO: 67985823, and Grant Agency of the Czech Republic P302/11/1308
Comparative proteomic analysis of an isogenic pair of lung normal and lung cancer cell line
Authors: Alam Md Maksudul , Hooda Jagmohan , Cadinu Daniela , Henke R. Michael , Ma Xiaotu , Zhang Li
Published: June 2021

Lung cancer is the leading cause of cancer‐related deaths in both men and women. Non‐small cell lung cancer (NSCLC) represents ~85% of lung cancer cases. Understanding how NSCLC cells differ from normal lung cells can help divulge the underlying causes of this disease. In this study, we analyze the protein expression pattern of an isogenic pair of normal (HBEC30KT) and NSCLC (HCC 4017) lung cell line. iTRAQ‐based quantitative differential MALDI TOF MS/MS was performed on eight cancer (NSCLC) samples and eight normal controls. We have identified 1465 proteins, 282 of which were differentially expressed with 1.3‐ fold change, with 119 up‐regulated and 163 down‐regulated. The protein expression pattern was verified by western blot for some randomly selected proteins. The up‐regulated proteins are involved in various functions including ribosome function, protein translation, mitochondrial functions and so on. However, among the down‐regulated proteins, most of them are involved in cytoskeleton and cell motility. The predicted cytoskeletal structures were also examined; as predicted from proteomic data, the cancer cell line showed thin and short actin stress fibers with reduced focal adhesion.