bluuebunny/crossref_metadata_2025 huggingface dataset:
The effects of strength training versus endurance exercise on blood volume expansion in rats with myocardial infarction (706.13)
Authors: Wason Emily , Garza Michael , Zhang John
Published: June 2021

After myocardial infarction (MI), cardiac remodeling causes sodium‐induced plasma expansion (PE), contributing to ventricular dilation. Evidence suggests Post‐MI endurance exercise (END) improves cardiac function (CF). However, it is known that END causes albumin (ALB)‐induced PE, a normal training adaptation. This may not be beneficial to an infarcted heart as it may worsen the MI‐induced PE. In contrast, strength training (ST) may increase myocardial strength without causing ALB‐induced PE. The aim of this study was to investigate the effect of ST versus END on PE in the post‐MI rat hearts. MI was induced by ligating the coronary artery. Survivors were assigned to 4 groups, Sham (n=9,), MI+SED (n=9, MI+no exercise), MI+END (n=7), and MI+ST (n=10). Exercise training was initiated 1 wk post‐MI on either a rodent treadmill (END) or 85° incline ladder (ST) for 10 wks. Our results showed total blood volume (TBV) and PE (ml·100g‐1) was significantly greater in MI‐END compared to MI‐ST (5.95±0.84 vs. 4.11±0.59, 3.74±0.49 vs. 2.49±0.24, p<0.05). Additionally, ALB concentration (g·dL‐1) was also significantly greater in MI‐END than MI‐ST (4.59±0.52 vs. 2.75±0.28, p<0.001). Our data suggests that compared to END, ST may favorably limit post‐MI TBV and PE.
Effects of progressive resistance and balance training 1‐3 years after stroke: a randomized controlled trial (706.14)
Authors: Vahlberg Birgit , Cederholm Tommy , Lindmark B , Zetterberg Lena , Hellström Karin
Published: June 2021

Objective: To compare the effect of a progressive functional resistance and balance training program with standard care on functional performance in individuals post stroke.

Methods: Thirty‐four community‐living individuals were randomized to progressive resistance and balance (PRB) training and 33 to standard care (C), for 3 months. About 75% were males and the age ranged from 65 to 85 years. Inclusion criteria: ability to walk 10 m, physical inactivity, and any of low fall‐related self‐efficacy, balance difficulties or repeated falls the last year. Primary outcomes: mobility (Short physical Performance Battery, SPPB) and balance (Bergs Balance Scale, BBS). Secondary outcomes: level of physical activity (Physical Activity Scale for the Elderly, PASE), walking capacity (6 Minutes Walking Test, 6MWT) and comfortable walking speed (m/s).

Results: Intention‐to‐treat analysis revealed significant differences in walking capacity (P=0.02, Effect Size; 0.68) and 10 meter self‐selected walking speed (P=0.014, ES; 0.68) in favor of the PRB group. Per protocol analyses also showed favorable effects for the PRB group in balance (P=0.02, ES; 0.68). No significant differences for the other secondary outcomes were observed. Within‐group analysis showed significant improvement in balance (BBS; P=0.002), mobility (SPPB; P=0.034), walking capacity (6MWT, P=0.005) and 10 meter self‐selected walking speed (10 mWT, P=0.036) in the PRB group only.

Conclusion Progressive resistance and balance training for three months performed in circuit class in individuals 1‐3 years after stroke are effective in improving walking capacity and walking speed.

Grant Funding Source: Uppsala University, Sweden
Casein hydrolysate supplementation enhances training effects on mitochondrial enzyme activities in slow muscle fibers but not in fast fibers in mice (706.18)
Authors: Matsunaga Yutaka , Tamura Yuki , Masuda Hiroyuki , Takahashi Yumiko , Hoshino Daisuke , Saito Noriko , Nakamura Hirohiko , Takeda Yasuhiro , Hatta Hideo
Published: June 2021

Aims: The aim of this study was to determine whether ingestion of casein hydrolysate enhances training effects on mitochondrial biogenesis.

Methods: Mice were randomly divided into three treatment groups: the control group (CON), the endurance training group (Tr), and the endurance training with casein hydrolysate group (CH+Tr). Mice were ingested either water or casein hydrolysate (0.2 mg/g BW, 7 times/week) and subjected to endurance training (20‐25 m/min×60 min, 5 times/week, for 6 weeks) 30 min after the supplementation. Muscular mitochondrial enzyme activities, citrate synthase (CS) and β‐hydroxyacyl‐CoA dehydrogenase (β‐HAD) were measured.

Results: In plantaris muscle, CS activities in Tr and CH+Tr groups were significantly higher than that in CON group (p<0.01) without significant differences between Tr group and CH+Tr group. On the other hand, CS activity in CH+Tr group was significantly higher than those in CON and Tr groups in soleus muscle (p<0.05). Similarly, in cardiac muscle, CS activity in CH+Tr group was significantly higher than that in Tr group (p<0.01) and tended to be higher comparing to CON group (p=0.051). β‐HAD activity was also increased in CH+Tr group in soleus and cardiac muscles.

Conclusion: Casein hydrolysate supplementation enhances training effects on mitochondrial enzyme activities in slow muscle fibers but not in fast fibers.

Grant Funding Source: Morinaga Milk Industry Co., Ltd
Cardiovagal activity following acute sprint interval exercise (706.6)
Authors: Olver T. , Abbott Kolten , Beaulieu Kristine , Lemon Peter , Shoemaker J.
Published: June 2021

The purpose of this study was to examine the impact of sprint interval exercise (SIE) on the time course of cardio‐autonomic recovery and vagal activity over a 24 h period. Eight healthy males (age=24.1±2.3 y) performed a single bout of SIE that included 4 x 30 s all out sprints interspersed by 4 min of recovery, performed on a manual treadmill. Following an overnight fast, measurements were taken at baseline (day 1; 8:00 a.m.), pre‐exercise (PRE‐X), immediately (PT‐X1), 90 (PT‐X2), 180 (PT‐X3) and 300 (PT‐X4) min post‐exercise as well as the following morning (day 2). Compared to PRE‐X, all spectral, time domain and Poincaré plot indices of heart rate variability (HRV) were reduced during PT‐X1‐3 (P蠄0.01). Baroreflex sensitivity was also depressed from PT‐X1‐4 (P<0.01). Relative to PRE‐X, plasma pancreatic polypeptide (PP; plasma indicator of vagal stimulation) was elevated at PT‐X1 (P=0.03). There were no differences in resting HRV, baroreflex sensitivity and plasma PP values between day 1 and 2 (P蠅0.12). These results indicate that despite an acute rise in plasma PP, there is a profound minimization of cardio‐vagal control following SIE. Cardio‐autonomic modulation recovers progressively and all indices of vagal activity are normalized by the following morning.

Grant Funding Source: Supported by NSERC
Changes to erythrocyte markers during lower body negative pressure and graded blood loss in humans (707.3)
Authors: Effertz Cary , Johnson Blair , Convertino Victor , Joyner Michael , Curry Timothy
Published: June 2021

We tested the hypothesis that erythrocyte count (RBC), hemoglobin (Hgb), and hematocrit (Hct) would be similar during lower body negative pressure (LBNP) and graded blood loss (BL) in humans.

Arterial blood pressure, central venous pressure and heart rate were measured in 12 healthy males (88 ± 3 kg) during LBNP and BL. The LBNP protocol consisted of 5 min stages at 0,‐15, ‐30, and ‐45 mmHg. The BL protocol consisted of removal of three 333 mL aliquots of blood (1 L total) which was stored for reinfusion. RBC, Hgb, and Hct were obtained via arterial catheter at baseline and at the end of the final stage for both LBNP and BL.

There were no differences (P > 0.05) in baseline values between treatment groups for RBC, Hgb, or Hct. RBC (4.81 ± 0.10 to 4.97 ± 0.10 1012/L), Hgb (14.18 ± 0.39 to 14.68 ± 0.36 g/dL), and Hct (40.88 ± 0.82 to 42.36 ± 0.83 %) increased from baseline to final stage (p<0.001 for all) during LBNP, while all values decreased from baseline to final stage (p<0.015 for all) during BL (4.82 ± 0.11 to 4.72 ± 0.11 1012/L; 14.33 ± 0.37 to 13.95 ± 0.39 g/dL; 41.09 ± 0.82 to 40.25 ± 0.86 %; respectively). The final stage values were greater in LBNP than BL for RBC, Hgb, and Hct (p< 0.001 for all).

LBNP results in relative hemoconcentration while BL results in relative hemodilution. The clinical significance of these findings remains to be elucidated.

Grant Funding Source: Supported by U.S. Army MRMC Combat Casualty Care Research Program Grant # W81XWH‐11‐1‐0823.
Influence of menstrual cycle phase on the cerebrovascular response to hypoxia: role of cyclooxygenase (708.1)
Authors: Peltonen Garrett , Rosseau Cameron , Harrell John , Schrage William
Published: June 2021

Menstrual phases influence resting cerebral blood flow (CBF), partially through cyclooxygenase (COX) mediated signaling. It is unknown if menstrual cycle hormone variations influence the CBF response to hypoxic stress. Compared to early follicular phase, we hypothesized females in early luteal phase would exhibit a larger, COX‐dependent, hypoxia‐mediated increase in CBF. We measured middle cerebral artery velocity (MCAv) with transcranial Doppler ultrasound in 4 healthy females (22 ± 3 yrs). Cerebral vascular conductance index (CVCi = MCAv*100/MABP) was calculated to normalize for potential differences in blood pressure. Females were studied twice each in early follicular (cycle day 1‐5) and late luteal (cycle day 12‐16) phases of the menstrual cycle, with and without COX inhibition (100 mg oral Indomethacin, Indo). Arterial O2 (SaO2) was reduced to 80% for 5 minutes while clamping end‐tidal CO2. On average, estrogen was higher (188 vs. 55 pg ml‐1) and progesterone was similar (790 vs. 603 pg ml‐1) during early luteal phase. Menstrual phase had no effect on basal CVCi (p=0.67) or the decrease in basal CVCi with Indo (p=0.45). Hypoxia increased CVCi (∆CVCi = 12 ± 1 cm s‐1 mmHg‐1; p<0.001), but the hypoxia‐induced ∆CVCi was not altered by menstrual phase (p=0.53) or Indo (p=0.873). These data suggest that menstrual phase does not alter basal or hypoxic CVCi and the role of COX is unchanged in either response.

Grant Funding Source: NIH R01 HL105820 & AHA PRE7390038
Tryptophan diet deficiency, intermittent hypoxia exposure and DNA methylation: implications in SIDS (710.10)
Authors: Lee Stella , Luikart Bryan , Nattie Eugene
Published: June 2021

The Sudden Infant Death Syndrome (SIDS) is the unexpected, sudden death of an infant in which an autopsy and a death scene investigation do not find a cause of death. Here, we focus on the role of brainstem serotonin (5‐HT) deficiency in SIDS pathogenesis as suggested by studies in SIDS cases. Further, a recent discovery of the effects of neonatal chronic intermittent hypoxia (CIH), a postulated occurrence in many SIDS cases, on DNA methylation as measured in adult rats suggests a potential role of epigenetics in the pathogenesis of SIDS. The present study investigated the effects of neonatal CIH on DNA methylation in rat pups treated with control vs. tryptophan deficient diets, the latter known to induce a brainstem 5‐HT deficiency. Our data suggest that global DNA methylation in the medulla increased significantly with tryptophan deficiency (Two way ANOVA, p<0.001) whereas exposure to CIH decreased it (p<0.001). Interestingly, there was a significant interaction effect between diet and CIH (p = 0.010). Similarly, DNA methylation in the raphe increased with tryptophan deficiency whereas the addition of CIH decreased it, demonstrating a significant interaction between diet and CIH (p = 0.029). Thus we find that two different risk factors associated with SIDS can result in epigenetic modification in the brainstem. It is unclear what the role of such epigenetic modification is in the context of 5‐HT deficiency, CIH and SIDS.

Grant Funding Source: Supported by National Institute of Child Health and Human Development Grant PO1‐HD‐036379
Xanthine oxidase activation mediates intermittent hypoxia increased NADPH oxidase activity (710.3)
Authors: Nanduri Jayasri , Vaddi Damodara , Khan Shakil , Wang Ning , Makarenko Vladislav , Prabhakar Nanduri
Published: June 2021

We previously showed that NADPH oxidases (Nox) especially Nox2 mediate systemic and cellular responses to intermittent hypoxia (IH). However, the mechanisms by which IH activates Nox2 have not been examined. We recently reported that xanthine oxidase (XO) activation by IH generates ROS which leads to increased intracellular calcium. Given that calcium activated PKC is required for Nox2 activation, we tested the hypothesis that XO activation mediates IH increased Nox activity via ROS augmented calcium. Allopurinol, a XO inhibitor or silencing of XO in PC12 cells prevented IH‐induced Nox2 activation by inhibiting Nox2 complex formation at the membrane. Treatment of cells with Xanthine/XO under normoxia mimicked the effects of IH. Our results further demonstrate that ROS generated by XO, contribute to IH induced Nox activation by increasing intracellular calcium levels. IH augmented PKC activity by phosphorylation of alpha subunit which was inhibited by allopurinol and BAPTA (calcium chleator). More importantly, inhibition of PKC activity by allopurinol abolished Nox2 complex formation. These results demonstrate that ROS generated by XO activates PKC alpha via calcium resulting in IH augmented Nox activity. Supported HL‐ ‐90554.

Grant Funding Source: HL‐90554
Different sub‐populations of pre‐sympathetic neurons are revelead by acute hypoxia in brainstem slices of adult rats (710.7)
Authors: Karlen‐Amarante Marlusa , Almado Carlos , Accorsi‐Mendonça Daniela , Machado Benedito
Published: June 2021

In this study we evaluated, in brainstem slices from adult rats, the effects of acute hypoxia (AH) on the firing rate of pre‐sympathetic neurons in rostral ventrolateral medulla sending projections to the intermediolateral column of spinal cord (RVLM‐IML neurons). Anesthetized male Wistar rats (PO 30) were submitted to microinjection of fluorescent retrograde tracer into the spinal cord (T3‐T5) to label the RVLM‐IML neurons. Five days later the rats were anesthetized to obtain brainstem coronal slices. The RVLM‐IML neurons were recorded using whole‐cell patch‐clamp and the firing rate was analyzed before and after perfusion of slice to AH for 2 minutes with hypoxic solution (95% N2 and 5% CO2) in the presence of synaptic blockers. After the AH protocol the viability of RVLM‐IML neurons were verified by the perfusion with normal solution (95% O2 and 5% CO2). All recorded RVLM‐IML neuronsof adult rats (n=17) presented intrinsic spontaneous firing activity. Three sub‐populations of RVLM‐IML neurons were identified after AH exposure: a) 3 neurons decreased their firing rate after AH(‐ 34,10%, p= 0,0225); b) 4 neurons increased their firing rate after AH (+ 40,86%, p= 0,0249) and c) 10 neurons were not affected by AH(± 3,54%, p= 0,5380. Hypoxia produced no changes in input resistance and resting membrane potential in these neurons. The data show that pre‐sympathetic neurons in the brainstem slices of adult rats present pacemaker activity and different sub‐populations of pre‐sympathetic neurons are revealed in response to AH.

Grant Funding Source: Supported by FAPESP and CNPQ
Primary screening of siRNA to determine genes involved in vascular leakage (711.3)
Authors: Hoke Allison , Jibitu Meskerem , Gautam Aarti , Hammamieh Rasha , Jett Marti
Published: June 2021

Endothelial cells lining the vessels of the vasculature and the cell‐cell junctions provide the primary barrier for the passage of fluids, immune cells, and macromolecules between the bloodstream and tissues. Appropriate and dynamic regulation of this barrier is required for normal physiological processes. To identify genes implicated in vascular leakage, we combined siRNA knockdown screens with an in vitro assay that models the human endothelium by using primary human microvascular endothelial cells (HMVEC‐L) plated on a 96‐transwell collagen‐coated filter unit. To investigate genes related to vascular leakage, a primary screen of ~500 siRNAs against drugable target genes was performed and the cells were plated for 48 hours for monolayer formation. The monolayer permeability was evaluated using dextran‐FITC to detect leakage through small gaps present in the early stages of permeability induction. The viability of the siRNA knockdown HMVEC‐Ls was also determined. We have identified genes from the primary screen that lead to increased permeability when knocked down. These results need to be validated. With the application of this in vitro assay, genes relating to vascular leakage can be more fully investigated and the siRNA knockdown results will help identify genes with a potential for therapeutic roles.

Grant Funding Source: Supported by the Transformational Medical Technologies program contract TMTI.DRUG.02.10.WR.023.
Induction of ectopic inflammasome function in fetal lung mesenchyme (715.4)
Authors: McCoy Alyssa , Greer Rachel , Prince Lawrence
Published: June 2021

Bronchopulmonary Dysplasia (BPD) commonly affects preterm infants. Inflammation and the soluble inflammatory mediator interleukin‐1 beta (IL‐1beta) play major roles in BPD pathogenesis and inhibit normal lung airway formation. IL‐1beta release requires both NF‐kappaB dependent mRNA expression and post‐translational cleavage by caspase‐1 containing inflammasomes. Best characterized in immune cells, the inflammasome has not been investigated in the developing lung. Here we test the hypothesis that the endotoxin lipopolysaccharide (LPS) can induce ectopic inflammasome expression in fetal lung mesenchyme, leading to IL‐1beta release. In primary fetal mouse lung mesenchymal cells, LPS rapidly induced IL‐1beta mRNA expression, but neither LPS nor the inflammasome activator ATP stimulated the release of IL‐1beta peptide. However, treating mesenchymal cells with LPS for at least 6h and then stimulating with ATP did release IL‐1b peptide, suggesting LPS‐dependent priming. These results were consistent with immunoblots showing caspase‐1 cleavage when mesenchymal cells were primed with LPS and then activated with ATP. LPS priming also increased caspase‐1 mRNA expression. Mesenchymal cells expressing a constitutively active IKKbeta contained higher levels of IL‐1beta and inflammasome component mRNA, but did not release IL‐1beta peptide unless first primed with LPS and then activated with ATP. These data suggested that IKKbeta activity might not be sufficient to assemble a fully functional inflammasome. These data show that LPS can generate functional inflammasome expression and function in lung mesenchymal cells, potentially propagating the release of inflammatory cytokines within the lung.

Grant Funding Source: Supported by 2R25GM059994‐14 and 5R01HL097195‐02
Role of hepatic microsomal oxidation in regulation of monoamine oxidase mediated processes in rat brain (727.1)
Authors: Tseylikman Vadim , Manukhina Eugenia , Downey H , Tseylikman Olga , Kozochkin Denis , Deev Roman , Misharina Mariya , Misharina Mariya , Komelkova Mariya
Published: June 2021

Monoamine oxidase (MAO) plays a key role in metabolism of biogenic amines and serves as a marker of alcoholism and depressive disorders. Various isoforms of cytochrome This study was focused on the relationship between MAO activity and hepatic content of cytochrome P450 which reflects the state of microsomal oxidation. Methods. For vital integrative evaluation of hepatic microsomal oxidation in rats, the hexobarbital sleep test was used. Content of cytochrome P450 was measured in hepatic microsomes. Activities of MAO‐A and MAO‐B were measured in the whole brain and brain structures. Content of serotonin, a MAO substrate was measured in the brain and plasma. Intensity of oxidative stress was evaluated by measuring lipid peroxidation products and carbonylated proteins in brain. Results. Rats with short hexibarbital sleep time (SHST) had higher content of microsomal cytochrome P450 than rats with long hexobarbital sleep time (LHST). Whole brain MAO‐A and MAO‐B activities, serotonin and carbonylated protein levels were higher in SHST than LHST rats. MAO‐A and MAO‐B activities were higher in brain cortex of SHST rats; only MAO‐A activity was higher in hypothalamus and medulla of LHST. The same brain regions of LHST rats had higher concentrations of carbonylated proteins and lipid peroxidation products. Conclusion. Data obtained in this study showed a correlation between MAO activity and microsomal oxidation phenotype. Rats with higher hepatic content of cytochrome P450 had higher activities of MAO‐A and MAO‐B in the brain and higher plasma serotonin levels than rats with lower microsomal oxidation. These correlations may reflect contribution of biogenic amines to regulation of cytochrome P450‐dependent monooxygenases and a role of cytochrome P450 in metabolism of biogenic amines and hormones regulating MAO activity including glucocorticoids

Grant Funding Source: Russian Foundation for Basic Research grant 11‐04‐01378‐а&14‐04‐01381‐a
Automatic sketching during manic writing episodes elicited by moment‐to‐moment shifts in the level of conscious awareness (728.20)
Authors: Canolty Nancy , Jennings Marcus
Published: June 2021

In this case study, subtle shifts in conscious awareness experienced by one of the authors (nlc) on the 5th day of a 29‐day manic episode were evaluated. While making handwritten stream‐of‐consciousness journal entries with a ballpoint pen, time (hr:min:sec) was recorded when conscious awareness shifted; subsequent arm movements not under conscious control were recorded by keeping the pen in hand and in contact with the journal page. Time (hr:min:sec) was again recorded when arm movements ceased. Tracings generated by arm movements were labeled autosketches and the duration of an autosketching session was the elapsed time between transitioning into and out of a given level of conscious awareness. Autosketches were assigned to one of 3 groups. Group 1 had light free‐form lines (low intensity); Group 2 had dark pleated lines (medium intensity); Group 3 had heavy superimposed lines constrained to a given area (high intensity). In the low, medium and high intensity groups, respectively, there were 142, 32 and 14 autosketching sessions with mean durations of 18, 30 and 38 seconds. Compared to groups of low or medium intensity, the high intensity group had the smallest number of autosketching sessions but those sessions had the highest mean duration. This case study documented that mania evokes moment‐to‐moment shifts in conscious awareness that vary in both duration and intensity.
Anti‐ inflammatory effects of thymoquinone in LPS‐ stimulated BV‐2 murine microglia cells (730.2)
Authors: Taka Equar , Mazzio Elizabeth , Goodman Carl , Reeams Renee , Soliman Karam
Published: June 2021

Thymoquinone (TQ) is one the main pharmacologically active constituents of the black cumin seeds (Nigella sativa) believed to be responsible for therapeutic effects on chronic inflammatory conditions such as arthritis and asthma. In this study, we investigate the possible neuroprotective action of TQ in activated microglia cells. We hypothesized that the anti‐inflammatory properties of TQ are mediated in part by inhibiting the formation of pro‐inflammatory cytokines, chemokine’s and other inflammatory mediators. In this study, we evaluated the anti‐inflammatory activity of TQ in lipopolysaccharide (LPS) (1µg/ml) stimulated microglia cells in the presence or absence of TQ (10µM). Cytokines/chemokine's were profiled using a RayBio antibody protein array and NO release was evaluated using the Griess reagent assay. In addition, cytokine antibody array results were validated using PCR array and ELISA. Our results showed that TQ significantly inhibited Pro‐inflammatory cytokines including IL‐6, IL‐1β, IL‐12p40/70, Growth Colony Stimulating Factor (G‐CSF), and pro‐inflammatory chemokine’s such as MCP‐5, IP‐10, MCP‐1 and inflammatory mediators NO production in LPS stimulated BV‐2 cells. In conclusion, the results obtained suggest that TQ could have a neuroprotction potential and may provide a mean for the treatment of neuro‐inflammation that occurs in neurodegenerative diseases such as Alzheimer and Parkinson’s diseases.

Grant Funding Source: Supported by NIH grants NIMHD G12 MD007582 and P20 MD006738
Features of finger and palm papillary pattern as morphological markers of individual predisposition to type 2 diabetes (734.6)
Authors: Sarapultsev Alexey , Sarapultsev Petr , Yakusheva Marina
Published: June 2021

The goal of the research is detecting morphological markers of individual predisposition to development of type 2 diabetes metabolic disorders. At the first stage of the research, features of papillary pattern of 126 people were observed ‐ 63 type 2 diabetes patients were examined and 63 people of the control group formed by the method of “directional selection” according to such parameters as age, gender, nationality, no diabetes. It included studying the dermatoglyphic pattern taken by the specially designed fingerprint scanner. The second stage studied interconnections of the features of papillary pattern and anthropometric measurements (body weight, body mass index, waist circumference, hip circumference) with laboratory parameters normal ranges (С‐peptide, immunoreactive insulin, glycosylated hemoglobin, fasting glycemia, insulin resistance index, cholesterol, triglycerides, lipoproteins) for 70 type 2 diabetes patients.

The results received testify to the existence of papillary pattern features with type 2 diabetes patients, distinguishing them from practically healthy people having no obesity or carbohydrate metabolism disorder. The presence of complicated palm patterns indicates possible disorders in the hypothalamo‐pituitary axis and predisposition to development of dysmetabolism.
An asymmetric gene evolution mechanism via replication‐transcription conflicts (735.11)
Authors: Merrikh Houra , Million‐Weaver Samuel , Nakta Samadpour Ariana
Published: June 2021

Bacterial genomes are generally biased such that most genes are encoded on the leading strand, in a co‐directional manner to DNA replication. This presumably avoids the potentially detrimental head‐on collisions that occur between the replication and transcription machineries when genes are encoded on the lagging strand. However, despite this co‐orientation bias, a significant number of highly conserved and essential genes remain head‐on to replication and are encoded for on the lagging strand. Our work, in the model organism Bacillus subtilis, led to the hypothesis that bacterial cells keep some genes in the head‐on orientation, in part, to accelerate their adaptive evolution in a targeted manner. Using both bioinformatics analyses and experimental data, we proposed that this targeted evolution mechanism is driven, at least in part, by the collisions between replication and transcription machineries. Our recent unpublished results are consistent with this hypothesis, and indicate that the asymmetric nature of how these conflicts impact the genome is indeed the major driving force behind this targeted and orientation‐dependent mutagenesis mechanism. Altogether, these findings have deepened our understanding of the impact of replication‐transcription conflicts on asymmetric mutagenesis, genome organization and evolution in bacteria. Because the underlying mechanisms are highly conserved in many species, this work potentially has far‐reaching implications beyond the bacterial world.

Grant Funding Source: Supported by NIH DP2GM110773
MLH3, a minor player in mismatch repair, is a major force in GAA•TTC repeat expansion (736.9)
Authors: Fuselier Kayla , Grabczyk Ed
Published: June 2021

Friedreich ataxia (FRDA) is a progressive neurodegenerative disorder caused by GAA·TTC repeat expansion in the first intron of the frataxin (FXN) gene. Disease severity correlates with the length of the expanded repeats and the reduction of FXN mRNA. Our objective is to refine our understanding of the role mismatch repair proteins play in the repeat expansion that causes FRDA. We have previously shown that the expansion rate is associated with transcription within the repeat and requires MutSβ and the subsequent action of a MutL complex. We now demonstrate that the necessary complex is MutLγ, a heterodimer of MLH1 and MLH3. We report here that our studies on MLH3 indicate that it is pivotal in GAA•TTC expansion in our human cell model system. MLH1 has a central role in all MutL complexes; whereas MLH3 is only a component of the MutLγ complex. Therefore, MLH3 may be the better potential therapeutic target for FRDA.

Grant Funding Source: Supported by the Friedreich's Ataxia Research Alliance and the LSUHSC Research Enhancement Fund
Imaging of molecular oxygen at subcellular resolution in mammalian cell culture: from 2D to 3D models (764.1)
Authors: Dmitriev Ruslan , Kondrashina Alina , Foley Tara , Papkovsky Dmitri
Published: June 2021

Oxygen monitoring is necessary for studying cell metabolism, stem cell development and ROS production, however most of the current methods suffer from low sensitivity and signal drift (electrodes), lack of quantitative context and end‐point nature (hypoxia stains) or inability to probe O2 inside the cell (extracellular probes). Phosphorescence lifetime imaging microscopy (PLIM) allows for quantitative, sensitive and specific detection of O2, by means of phosphorescent indicator dyes. We developed phosphorescent O2 probe (Pt‐Glc) which demonstrates efficient cytosolic staining of different cell lines (2D cultures), and 3D tissue models such as spheroids (primary neurospheres) and live brain slices from embryonic tissue. The PLIM analysis of neurospheres from rat brain cortex with Pt‐Glc confirmed presence of hypoxic niches. We correlated oxygenation map of spheroids with their size, distribution of live cell (cholera toxin, TMRM), proliferating (BrdU) and stem cell markers (nestin), thus performing multiplexed metabolic imaging of live neurosphere. A comparison with pimonidazole, HIF‐a immunostaining and nanoparticle probes was also made. Thus, Pt‐Glc represents valuable tool for studying stem cell niche within ex vivo and in vitro 3D models.

Grant Funding Source: Supported by the Science Foundation Ireland.
Differential effect of high fat diet on tissue morphology and hormonal levels in male and female mice (765.1)
Authors: Wong Nicole , Donthireddy Laxminarasimha , Babu Sathyapriya , Adhikary Debasis , Chakraborty Sanjoy , Chakraborty Tandra
Published: June 2021

The pathology of obesity produces many diseases. It has a far ranging negative effect on health including some type of cancers, non‐alcoholic fatty liver disease(NAFLD), diabetes and many other diseases that are related with obesity. This project was undertaken to observe the differences in the tissue morphology (liver, pancreas, adipose, heart, brain, kidney, spleen, ovaries and testes) in male and female mice when exposed to high fat diet for a period of 6 months. Within 3 months animals in VHFD showed signs of NAFLD and with longer treatment the liver showed deposition of lipid droplets in hepatocytes, which leads to non‐ alcoholic steatohepatitis (NASH) subsequently cirrhosis and hepatocellular carcinoma. Morphometric analyses of pancreas showed robust signs of inflammation, increase in pancreatic islets size and number in both obese male and female. Adipose tissue showed signs of hypertrophy than hyperplasia. The effect of high fat was more pronounced in males than females. obesity‐associated hyperleptinemia and hyperinsulinemia was observed in both males and female. Thus, these results suggests that high fat diet not only affects adiposity and hormonal imbalance but changes the cellularity and morphology of the tissue to such an extent that can lead to diseases that are fatal affecting men more than women.
Exposure to gasoline power generator exhaust fume negatively affected hematological, liver and kidney function parameters in female albino rats (766.7)
Authors: Igwo‐Ezikpe Miriam , Awodele Olufunsho , Anyakora Chimezie , Ifegwu Clinton , Owolabi Bukola , Oyewale Ayodele
Published: June 2021

The lack of constant electricity power supply to meet utility usage in developing countries like Nigeria has warrant the use of petroleum powered generators to supply electricity. There has also been dearth research on the effects of exposure to fumes from the generators thus necessitating the need for this research. This study therefore investigated hematological, liver, kidney, hormonal and histopathological changes in female albino rats exposed to gasoline power generator exhaust fume at varied distance of 100, 200 and 300 meter respectively after 42 days. Present study showed that there was significant (p < 0.05) increase in packed cell volume, white blood cells, lymphocytes, neutrophiles, monocytes and platelets in the exposed animals compared to control unexposed rats. Liver and kidney function parameters were also significantly (p < 0.05) increased whereas there were no significant (p <0.05) difference in follicle stimulating hormone, prolactin, progesterone and estradiol amongst the animals, however, luteinizing hormone was increased in 300 meter exposed rats. Histopathological investigation showed degenerated organs in the exposed animals compared to control. Generally, the observed variations in the parameters were proportional to the distance from source of exposure. This study demonstrated that constant exposure to gasoline power generator as is the case in Nigeria could induce some biochemical changes which may be toxic to exposed individuals and may lead to adverse health conditions such as cancer.
Overexpression and FPLC‐purification of the DNA polymerase from T. aquaticus (779.7)
Authors: Vidmar Kristen , Parente Amy
Published: June 2021

Systematic evolution of ligands by exponential enrichment (SELEX) is a methodology in biochemistry for identifying nucleic acid sequences capable of adopting a shape that binds a target molecule. The process involves several rounds of the polymerase chain reaction (PCR) to amplify DNA sequences followed by steps that ultimately separate and enrich the population of DNAs with sequences having a high affinity for the target molecule. Four main reagents required for any PCR reaction include: 1 ‐ DNA template, 2 ‐ DNA primers, 3 ‐ dNTPs, and 4 ‐ DNA polymerase. PCR optimization is frequently performed to identify experimental conditions that are optimal for the chosen template and primers. This can be a costly step of the experimental procedures, as commercially available DNA polymerases can range from a few hundred to thousands of dollars. We have undertaken the process of overexpressing and purifying the heat‐stable DNA polymerase from Thermus aquaticus. A plasmid containing the gene for Taq DNA polymerase was transformed to BL21(DE3) E. coli cells. Cell cultures of these transformed cells can then be grown to saturation at 37°C and overexpression initiated with the addition of IPTG. The desired Taq polymerase will be purified from a portion of the cleared lysate using ion‐exchange chromatography. PCR activity using a standard template and primers can then be evaluated using samples of purified and unpurified Taq polymerase with comparison to several commercial samples.
Role of SET oncoprotein in epithelial‐mesenchymal transition and pancreatic cancer progression (784.3)
Authors: Hung Sau Wai , Lee Haesung , Gilbert Caitlin , Hoang Toan , Naidu Kineta , Govindarajan Rajgopal
Published: June 2021

One of the key reasons for why pancreatic cancer has such a poor prognosis is that >80% of diagnoses occur when metastasis has already presented. At the cellular level, activation of the epithelial‐mesenchymal transition (EMT) is significant because it allows for the dissemination of primary tumor cells to distant sites. In this study, we sought to determine the role of a novel oncoprotein, SET, in EMT and pancreatic cancer cell progression. We found that silencing SET reverted EMT by inducing a cuboidal, epithelial morphology and reducing colony formation, cellular proliferation, migration, and invasion. Consistently, SET overexpression was detected in a subset of pancreatic tumors, especially poorly‐differentiated pancreatic ductal adenocarcinomas. Higher levels of SET protein expression were also identified in all pancreatic cancer cell lines compared with normal human pancreatic ductal epithelial cells. SET‐mediated EMT was determined to be a result of cadherin switching during which the SPARC/Slug and Rac1/JNK/c‐Jun/AP‐1 signaling pathways inhibit epithelial E‐cadherin and promote mesenchymal N‐cadherin, respectively. In brief, we elucidated a novel role for the SET protein in inducing EMT through cadherin switching and revealed the molecular pathways involved. These findings have implications for the design and targeting of pathways necessary for intervening pancreatic tumor progression.

Grant Funding Source: Supported by NIH 1021RR571367 and 1021RR571381.
Necessity for EFGR kinase enzymatic activity in the coordinate upregulation of a chemoattractant gene suite by vasoactive intestinal peptide in mouse lymphocytes (784.5)
Authors: Saunders Jenna , Wanjara Steve , Hoselton Scott , Schuh Jane , Dorsam Glenn
Published: June 2021

Vasoactive intestinal peptide (VIP) is a potent chemoattractant for immune cells that is delivered by the peripheral nervous system to immune organs. However, the molecular mechanism regulating VIP’s chemoattractant properties is not known. Previously, our group published the VIP transcriptome in resting CD4 T cells, which identified a chemoattractant gene set (e.g. EGFR, Snail and syt XIII) that was coordinately upregulated. We hypothesize that these gene products control the chemoattractant activity of VIP in immune cells. Moreover, we propose that this chemoattractant gene set is transcriptionally upregulated by the transactivation of EGFR by VIP as is observed in non‐immune cells, including breast and colon cancer cells. This research will utilize primary mouse splenocytes treated +/‐ VIP over various time intervals in an attempt to identify which FACS‐sorted splenocyte subpopulations are sensitive to VIP and the extent to which the chemoattractant gene set is upregulated. Optimal conditions will be repeated +/‐ EGFR neutralizing antibody or a specific EGFR kinase inhibitor to assess the contribution of EGFR enzymatic activity for the chemoattractant gene expression changes. Lastly, Boyden‐chamber chemotaxis assays will be employed to test whether pharmacological inhibition against EGFR kinase activity and/or other chemoattractant gene products suppresses VIP’s chemoattractant activity using identified splenocyte subpopulations. The significance of the proposed research is to better understand the molecular control of lymphocyte trafficking mediated by the nervous system.

Grant Funding Source: This research was supported by NIH/NIAID (R15 to GD).
O‐GlcNAcylation modifies the metastatic properties of prostate cancer cells (789.5)
Authors: Liu Xin , Yang Pengyuan , Hart Gerald
Published: June 2021

Prostate cancer is one of the most common forms of cancer in men and the second leading cause of cancer deaths in men in the United States. O‐linked β‐N‐acetylglucosamine (O‐GlcNAc) is a ubiquitous and dynamic post‐translational modification that exists on serine and threonine residues of nuclear and cytoplasmic proteins. This modification is regulated by O‐GlcNAc transferase (OGT), which attaches O‐GlcNAc to proteins, and O‐GlcNAcase (OGA), which removes O‐GlcNAc. O‐GlcNAc serves as a nutrient sensor to regulate virtually all cellular processes as well as playing roles in the various diseases, including Alzheimer's disease, diabetes, and cancer. The roles of O‐GlcNAc in the regulation of prostate cancer remains unclear. Herein, we investigate how the regulation of O‐GlcNAc may affect prostate cancer, progression using normal prostate epithelial cell (PrEc) and prostate cancer cell lines (LNCaP and PC‐3) as models. Using western blotting techniques, we show that O‐GlcNAc, OGT, and OGA are highly elevated in LNCaP and PC‐3 when compared to PrEc. LNCaP exhibited the highest levels of O‐GlcNAc, OGT, and OGA. OGT and OGA are differentially localized in lines with different metastatic properties. Additionally, 2D‐gel electrophoretic analyses show significant differences in O‐GlcNAcylated protein profiles among the three cell lines. In order to answer how O‐GlcNAc affects the biological properties of these cancer cells, including shapes, proliferation, invasion and metastasis, several assays were used. The result of soft agar assays indicate that alterations in O‐GlcNAc influence the metastasis ability of prostate cancer.
Gas chromatographic‐mass spectroscopic identification of the bioactive compounds in Hydrocotyle bonariensis (794.8)
Authors: Adebawo Olugbenga , Oladipo Akinwunmi , Obaseki Oluwatofunmi , Adesegun Oluwanbo , Gisarin Olusola , Anyasor Godswill , Osilesi Odutola
Published: June 2021

Hydrocotyle bonariensis Comm. Ex Lam (Apiacae) commonly called Large Leaf Pennyworth is a herbaceous, hairless, perennial plant which is prostrate with creeping lateral stem. It is found in the tropical and subtropical regions of the world. The plant’s importance in agriculture is in its usefulness in land conservation. However, the leaf of the plant has been reported to be useful as emetic, diuretic and laxative. Also, the leaf has been used in the treatment of tuberculosis, relieving the pain of rheumatism and arthritis, to increase brain capacity and for longevity. We have previously reported that the leaves of H. bonariensis contain alkaloids, flavonoids, tannins, phenolics and saponins as bioactive components. The present work is a preliminary investigation into the elucidation of the bioactive compounds responsible for the curative effects ascribed to the plant. Gas chromatographic‐mass spectrometric analytical method was adopted for the chemical identification of the compounds. Result revealed that a number of anti‐oxidant, anti‐inflammatory and anti‐cancer agents including caryophellene, falcarinol and 2‐methoxybenzyl‐amine were present in the methanolic and hexane extracts of the leaves of the herb.
The role of myocyte‐enriched calcineurin‐inhibiting protein (MCIP1) in estrogen mediated repression of cardiac hypertrophy (795.5)
Authors: Turpin Courtney , Waikel Rebekah
Published: June 2021

Premenopausal women have a lower incidence of left ventricular hypertrophy (LVH) and heart failure, which is thought to be the result of estrogen signaling. However, the mechanism for estrogen’s cardioprotective effects is not well understood. We seek to define the role of MCIP1, in estrogen‐mediated inhibition of LVH. It is known that estrogen induces the expression of MCIP1, which inhibits prohypertrophy protein calcineurin. In our study, LVH was surgically induced in 10‐12 week old mice using transverse aortic constriction (TAC). Two weeks post‐surgery, hearts were excised and total RNA was extracted from the apex of the heart for gene analysis. Given that estrogen is known to induce MCIP1, we expected MCIP1 levels to be higher in females. Surprisingly, mRNA levels of MCIP1 in SHAM females were 50% that of males and TAC levels of MCIP1 decreased in females to 20% of their SHAM value. To better understand regulation and the role of MCIP1 in estrogen mediated inhibition of hypertrophy, we are currently performing a series of in vitro experiments using neonatal cardiomyocytes exposed to phenylephrine to induce hypertrophy in the presence or absence of estrogen. Gene expression of MCIP1 and known modulators, including targeting miRNAs are being assessed at the mRNA and protein levels. Together these studies will define MCIP1’s role in estrogen repression of cardiac hypertrophy.

Grant Funding Source: NIH 5P20RR016481‐12 8P20GM103436‐12, R15HL108192‐02, R25GM102776
Retinoic acid and sodium butyrate attenuate renal fibrosis and inflammation in guanylyl cyclase‐A/natriuretic peptide receptor‐A gene‐targeted mice (796.13)
Authors: Kumar Prerna , Periyasamy Ramu , Das Subhankar , Pandey Kailash
Published: June 2021

Mice lacking functional guanylyl cyclase/natriuretic peptide receptor‐A (GC‐A/NPRA) gene (Npr1) exhibit hypertension, kidney disease, and heart failure. The objective of the present study was to elucidate the effect of all‐trans retinoic acid (ATRA) and histone deacetylase inhibitor, sodium butyrate (NaBu) on attenuation of renal fibrosis and remodeling in Npr1 gene‐targeted mutant mice. Mice (24‐week old) were injected intraperitonealy with ATRA (0.5 mg/Kg/day), NaBu (0.5 mg/Kg/day), and ATRA‐NaBu (1.0 mg/Kg/day) for 2‐week. The results showed that Npr1 gene‐disrupted heterozygous (1‐copy; +/‐) mice exhibited renal fibrosis and augmented levels of proinflammatory cytokines compared with wild‐type (2‐copy; +/+) and gene‐duplicated heterozygous (3‐copy; ++/+) mice. Retinoic acid and NaBu treatment significantly lowered systolic blood pressure in 1‐copy mice and increased plasma cGMP levels in Npr1 gene dose‐dependent manner with pronounced effects in 1‐copy mice. Treatment with ATRA‐NaBu combination synergistically attenuated renal fibrosis by 70%, immunoexpression of renal α‐SMA by 75%, and plasma and renal levels of proinflammatory cytokines TNF‐α and IL‐6 in 1‐copy mice compared with vehicle‐treated control mice. The present results provide direct evidence that ATRA and NaBu act as potent immunosuppressive and antifibrotic agents and repair the abnormal renal pathology in mice with reduced Npr1 gene copy number, which will have important implications in prevention of hypertension‐related renal pathophysiological conditions.
Preclinical efficacy of the dual sigma receptor antagonist dopamine uptake inhibitor, CM699, as a medication for stimulant abuse (803.3)
Authors: Katz Jonathan , Hiranita Takato , Lopez Jessica , Brellenthin Angelique , Mesangeau Christopher , Alsharif Walid , Kopajtic Theresa , Coggiano Mark , Jamalapuram Seshulatha , Tsai Shang‐Yi , Avery Bonnie , Su Tsun‐Ping , Tanda Gianluigi , McCurdy Christopher
Published: June 2021

Sigma receptor (σR) antagonists block many effects of stimulants, but not reinforcing effects assessed with self‐administration (SA). Yet, a recent study suggests that σR antagonism with dopamine transporter (DAT) inhibition selectively blocks stimulant SA. A compound with potential for dual σR‐DAT inhibition was synthesized by treating 1‐methyl‐2‐benzimidazolinone with 1,4‐dibromobutane in the presence of K2CO3 in dimethylformamide (DMF), yielding a compound that was then coupled with 3H‐spiro[isobenzofuran‐1,4'‐piperidine] in the presence of K2CO3 in DMF. The resulting compound, CM699, had affinities of 14, 2.30 and 318 nM at σ1Rs, σ2Rs and DAT, respectively. Antagonism at σ1Rs was confirmed by Western blots of σ1R‐EYFP bound to the ER chaperone, Binding immunoglobulin Protein (BiP). CM699 had no effect whereas the agonist, (+)‐pentazocine decreased BiP association. CM699 (5.0 mg/kg, iv) in rats produced a Cmax of 1.84 µg/mL in plasma, 5 min after injection which declined exponentially (overall T½ = 4.4 hr). CM699 produced dose‐dependent insurmountable and selective antagonism of cocaine SA in rats (ED50 = 3.99 mg/kg, ip) and blunted effects of cocaine on dopamine (DA) levels in the rat nucleus accumbens, indicating minimal abuse liability. The results provide preclinical proof of concept for dual σR/DAT inhibition as a novel approach for the development of medications to treat stimulant abuse.

Grant Funding Source: NIH Intramural Research Program