bluuebunny/crossref_metadata_2025 huggingface dataset:
Mentoring from Both Sides: How to Find, Be and Utilize a Great Mentor
Authors: Kamens Joanne
Published: April 2020

Mentoring should not be a scary or imposing concept ‐ it's really just about getting and giving advice, support and encouragement for ongoing learning. This workshop is intended for everyone: trainees and faculty ready to learn practical tactics in identifying mentors, making the "ask" for mentoring support and how to take the best advantage of mentoring relationships‐‐from both sides.

To learn more about how a strong mentoring culture benefits organizations of any size, Attend the "Re‐Imagining STEM: Who We Are and What We Do" workshop on Sunday, April 5 at 6 PM.
In Utero Exposure to Second‐Hand Cigarette Smoke Promotes Epigenetically‐Mediated Cardiac Dysfunction in High‐Fat Fed Male Mice
Authors: ronis martin , Xia Huijing , rodrigues samya lima , El Haji Eli , Noel Alexandra , Penn Arthur , Pulliam Casey , Gardner Jason , shankar kartik , Varner Kurt
Published: April 2020

We investigated whether in utero exposure to second‐hand cigarette smoke (SHS) would adversely affect cardiac structure/function in the exposed offspring fed a high‐fat diet. Pregnant mice (C57B1/6J) were exposed to SHS or air (4 hr/d) from gestational days 6–19. The male offspring (age: 65 days) from the SHS and air exposure groups were fed either normal chow, or high‐fat liquid diet (35% fat; HFD) for 16 weeks (n=12/group). Cardiac function was measured by echocardiography at baseline (30 days) and at the end of pair feeding. Mice were then sacrificed and heart tissue collected for histological and mRNAseq analysis. There was no significant difference in body weight or heart weigh/tibial length between the in utero SHS+HFD and in utero air+HFD treated offspring. However, compared to in utero air+HFD mice, in utero SHS+HFD mice had decreased left ventricular internal diameters and volumes during diastole (3.95±0.09 vs. 4.34±0.24 mm; 67.8±1.5 vs. 85.4±4.4 μl, P<0.05) and systole (2.91±0.07 vs. 3.34±0.15 mm; 32.5±1.9 vs. 46.3±5.1 μl, P<0.05), accompanied by increases in left ventricular wall thickness during diastole (0.72±0.03 vs. 0.66±0.03 mm, P<0.01) and septal thickness during systole (1.2±0.06 vs. 0.99±0.05 mm, P<0.05) and increased fractional shortening. There was also a significant increase in collagen in the hearts of the in utero SHS+HFD mice. These data are consistent with the development of concentric cardiac hypertrophy in in utero SHS+HFD mice. mRNAseq analysis of cardiac tissue revealed 100 up‐regulated and 120 down‐regulated genes in in utero air+HFD vs in utero SHS+HFD. Top Wiki Pathway terms included genes linked to adaptive cardiac hypertrophic responses: adrenergic receptor a1a, MAP2K1, MAP2K3, CDK7, CHUK, PDPK1 and IKBKE, microRNAs in cardiomyocytes, exercise induced changes in circadian rhythm (upregulated BMAL1, downregulated Per2) and signaling via estrogens, TNF alpha and MAP kinases. GO pathways identified included cardiac muscle contraction, protein acetylation and regulation of DNA methylation. We conclude that in utero exposure to SHS produces epigenetically‐regulated differential cardiac responses following challenge with obesogenic high fat diet. (NIH P20 GM103514).

Support or Funding Information

NIH P20 GM103514
Polyamine Regulation in Diabetic Breast Cancer Cells
Authors: Capellen Caleb , Ortega Jose , Dunworth Matthew , Casero Robert A. , Chandra Surabhi
Published: April 2020

Polyamines (spermine, spermidine, and putrescine) are involved in basic cellular processes such as cell growth, replication, and transcription. They are highly regulated cations that are elevated in cancer. Upregulation of polyamines can thus develop tumors due to hyper‐proliferation of cells as observed in cancers of the colon, skin, prostate and breast. Inhibitors of polyamine pathway could prevent tumor growth as shown in animal models of colon and skin cancer. Intracellular polyamine is regulated by biosynthesis, intestinal uptake and microbial production. Thus inhibitors of synthesis alone have not been very effective as cytotoxic agents but exhibit cytostatic properties. Diabetic conditions further accelerate the growth and metastasis of cancers such as pancreas, breast, liver, and colorectal, however the role of polyamines in such states has not been investigated. We hypothesized that polyamine regulation affects proliferation of breast cancer cells and normal breast epithelial cells in diabetic conditions. Breast cancer cells (MDA‐MB‐231) and normal breast epithelial cells (MCF‐10A) were treated with normal glucose (NG, 5mM) or high glucose (HG, 25 mM) in the presence/absence of polyamine pathway inhibitors. Cells were then treated with polyamine supplements, following depletion with polyamine pathway inhibitors. There was a time dependent increase in cell proliferation with HG from 48–72 hr compared to NG. This was correlated with an increase in polyamines (putrescine, spermidine) though spermine levels were not changed considerably. Simultaneous treatment with an ornithine decarboxylase (polyamine synthesis enzyme) inhibitor, difluoromethylornithine (DFMO) could prevent cell proliferation, restore polyamines, and normalize polyamine catabolic enzymes with high glucose exposure. In conclusion, polyamine regulation is responsible for proliferation of breast cancer cells and normal breast epithelial cells in diabetic states, which can be prevented using inhibitors affecting polyamine levels in cells.

Support or Funding Information

Funding: Grants from the UNK Undergraduate Research Fellows Program and UNK Research Services Council Seed Grant
Equipment: Microplate Reader (Dr. Nuxoll’s lab), Grant from the National Institute for General Medical Science (NIGMS) (5P20GM103427), a component of the National Institutes of Health (NIH)
Clinical Significance of Activating Interleukin‐1 Ligands in HPV‐positive and HPV‐negative HNSCCs
Authors: Gibson-Corley Katherine N. , Coppock Joseph , Espinosa-Cotton Madelyn , Ofori-Amanfo Georgina , Simons Andreans
Published: April 2020

Interleukin‐1 alpha (IL‐1α) and interleukin‐1 beta (IL‐1β) are cytokines involved in the acute phase immune response and their expression is upregulated in a variety of solid tumors including head and neck squamous cell carcinomas (HNSCCs). Our previous data has shown that IL‐1α in particular is associated with poor prognosis but this has yet to be studied in the context of human papilloma virus (HPV) status. This study is aimed at determining differences in tumor expression and the clinical significance of IL‐1α and IL‐1β in HPV‐positive (HPV+) and HPV‐negative (HPV‐) tumors. Differences in tumor gene expression and the prognostic value of IL‐1α and IL‐1β were determined in HPV+ and HPV− tumors from publically available gene expression datasets. Tissue microarrays (TMAs) containing HPV+ (n=31) and HPV− (n=47) primary and metastatic HNSCCs were analyzed for IL‐1α and IL‐1β expression using immunohistochemical (IHC) staining. HPV status was confirmed using p16 IHC staining and RNAish. We found that both IL‐1α and IL‐1β gene expression were significantly increased in HPV− tumors compared to HPV+ tumors. However, tumor expression (as detected by IHC) of IL‐1α (but not IL‐1β) was significantly increased in HPV− tumors compared to HPV+ tumors. Comparison of matched primary and metastatic tumors revealed a decrease in IL‐1α expression and an increase in IL‐1β in metastatic tumors compared to primary tumors. Lastly, increased IL‐1α gene expression was significantly associated with worse survival in HPV− tumors but not in HPV+ tumors. IL‐1β held no prognostic value in HPV+ or HPV− tumors. Overall IL‐1α expression may possess more prognostic significance in HPV− tumors rather than HPV+ tumors and this work warrants further investigation in larger patient cohorts.
Characterization of Novel Intron Splice Mutations in Patients with Propionic Acidemia
Authors: Smith Lorenzo Jason , Stoeger Braden Frederick , King Kaitlyn , Caporale Diane
Published: April 2020

Propionic Acidemia (PA) is a rare genetic metabolic disorder characterized by a deficiency in propionyl CoA carboxylase (PCC), an enzyme responsible for the proper breakdown of certain proteins and lipids. As a result, patients exhibit an abnormal accumulation of toxic organic acids, which can cause detrimental health problems. PA derives from mutations in PCCA and PCCB genes, which are responsible for expressing the vital subunits of PCC. Mutations in intronic sequences may cause errors in RNA processing, specifically with RNA splicing. If intronic segments are improperly included in the final mRNA form, the resultant products become nonfunctional. The goal of this project was to determine whether novel intron splice site mutations induce improper mRNA processing and, thus, nonfunctional proteins. To achieve this, we collaborated with genetics testing company Prevention Genetics to construct minigene plasmids containing intron mutations from patients. The minigene system was transfected into mouse fibroblast cells to allow mRNA processing to occur. RNA was isolated, reverse‐transcribed into cDNA, and sequenced to determine whether the mutated introns are splicing differently from the wildtype. Initial results indicate several identified intronic mutations contribute to an altered splicing pattern in patients with PA. Knowledge of the effect of these mutations on gene expression will strengthen the genetic database to help doctors diagnose PA in their patients, as well as help develop preventive and personalized treatments.

Support or Funding Information

UWSP OSCAR Travel Grant (Up to $3,000 for three students)
CD4+ T Cells from RUPP rat model activate NK cells and cause mitochondrial oxidative stress and hypertension in normal pregnant rats
Authors: Deer Evangeline , Reeve Kristin E. , Amaral Lorena M. , Vaka Venkata Ramana , Franks Michael , Fitzgerald Sara , Herrock Owen , Ibrahim Tarek , LaMarca Babbette
Published: April 2020

Preeclampsia (PE) is new onset hypertension during pregnancy and is associated with elevated inflammatory response such as CD4+ T cells, NK cells, and cytokines. We have previously shown that women with PE exhibit increases in placental mitochondrial (mt) dysfunction and ROS compared to normal pregnant (NP) women. The Reduced Uterine Perfusion Pressure (RUPP) rat model produces many characteristics of PE such as hypertension, increases in CD4+ cells, renal and placental NK cells, and mt dysfunction/ROS. Moreover, we have shown an important role for mt ROS in hypertension in RUPPs. We have previously demonstrated that RUPP CD4+T cells cause hypertension in NP rats, however the role of RUPP CD4+ T cells in stimulating NK cells to cause mitochondrial dysfunction/ROS are unknown. Therefore, we examined the effect of adoptive transfer of RUPP CD4+ T cells to activate NK cells in NP rats and cause mt dysfunction and ROS as mechanisms of hypertension. Splenic CD4+ T cells were isolated from RUPP rats, cultured, and injected into NP rats on gestation day (GD) 13. On GD19, MAP blood and tissue samples were collected. Mt respiration and mtROS were measured in isolated mitochondria using the Oxygraph 2K and fluorescent microplate reader, respectively. A student’s t‐test was used for statistical analysis. On GD19, MAP increased to 110±2 mmHg (n=12) in RUPP CD4+ T cell recipients compared to control NP rats 101±2 mmHg (n=7, p<0.05). Circulating cytolytic NK cells increased to 3±0.6% in RUPP CD4+ T cell recipients (n=8) compared to NP controls 0.3±0.2% (n=7, p<0.05). Renal state 3 (194.3±62.33 vs 1004±251.9 pmol/sec/mg, p<0.05) and maximal (151.2±75.19 vs 531.5±99.97 pmol/sec/mg, p<0.05) respiration rates, indicative of ATP production and electron transport chain efficacy respectively, were reduced with RUPP CD4+ T cells (n=6) compared to NP (n=4). Renal mtROS increased from 251.5±81.75% in NP controls (n=4) to 416.9±47.68% in RUPP CD4+ T cell recipients (n=9, p<0.05). Collectively, the data indicate that the adoptive transfer of RUPP CD4+ T cells stimulates cytolytic NK cells and renal mitochondrial dysfunction/ROS during pregnancy as important mechanisms of hypertension in the pathophysiology of preeclampsia.

Support or Funding Information

NIH Grants: RO1HD067541‐06 and P20GM121334; T32HL105324
Oxygen Deficit Is a Sensitive Measure of Mild Gas Exchange Impairment at Inspired O2 between 12.5%–21%
Authors: Liu Matthew A. , Stark Phoebe C. , Prisk G. Kim , West John B.
Published: April 2020

The oxygen deficit (OD) is the difference between the end‐tidal alveolar PO2 and the estimated PO2 of arterial blood derived from the SpO2 that has the potential to be used clinically as a proxy for the alveolar‐arterial PO2 difference. Previous work has shown that a recently developed technology, the Alveolar Gas Meter, is accurate in measuring the efficiency of pulmonary gas exchange, via the OD, in patients with a history of lung disease as well as in normal subjects breathing a hypoxic mixture of 12.5% O2. This study sought to determine how the OD varied as the percent of inspired O2 changed from 21% to 12.5% in 36 healthy subjects split by age (young 22–31, n=23; old 42–90, n=13) to assess the possibility of use in a clinical setting. The PO2 and PCO2 were continually measured as the subject breathed through a mouthpiece using rapidly responding electrochemical cells, and the end‐tidal PO2 and PCO2 were calculated and averaged over 5 steady‐state breaths, while a fingertip pulse oximeter measured the SpO2. After correcting for any change in P50 resulting from the PCO2 differing from 40 mmHg, the arterial PO2 was calculated from the SpO2 by inverting the Hill‐equation. Subjects breathed 12.5, 15, 17.5 and 21% oxygen mixtures (balanced order) and their OD was measured in triplicate. Across all inspired O2 levels, the OD was higher in the older cohort than the young (p <0.0001, repeated measures ANOVA, Table
1). The older cohort had a larger OD at all four oxygen levels (all p<0.0001), with that difference and its variance being greater at higher inspired O2 values (age × O2 interaction p=0.002). The OD at an inspired O2 of 12.5% was significantly less than at 21% for both age cohorts. This is consistent with the known increase in ventilation‐perfusion mismatching that occurs in healthy aging. The decrease in OD magnitude as the inspired oxygen was reduced in both cohorts is consistent with the increased accuracy of the estimated arterial PO2 when based on saturation values on the steep part of the O2‐Hb dissociation curve. Further, as arterial saturation decreases there is an expected decrease in the alveolar‐arterial PO2 difference. The persisting higher OD seen in older subjects, irrespective of inspired O2, suggests that the measurement of oxygen deficit remains sensitive to mild gas exchange impairment, even when breathing air (21% O2).

Support or Funding Information

Financial support from UCSD

Measured Oxygen Deficit by Age

Young

Old

p‐value

All Inspired O2 (mean±SE)

−0.4±0.6

10.6±1.0

<0.0001

Inspired O2 by Age Interaction

0.002

12.5%

−1.4±0.7

5.7±0.6

‐

15%

−2.7±1.0

5.2±1.8

15% vs 12.5%: NS

17.5%

−2.7±1.0

9.4±1.8

17.5% vs 12.5%: NS

21%

5.1±1.3

21.9±2.1

21% vs 12.5%: <0.0001
Empagliflozin Reduces Arrhythmic Events and Improves Ca2+ Transient in Hypoxia‐induced Injury Rat Cardiomyocytes
Authors: dos Santos Danúbia Silva , Turaça Lauro Thiago , da Silva Coutinho Keyla Cristiny , Brunswick Tais Hanae Kasai , de Carvalho Antonio Carlos Campos , Girardi Adriana C.
Published: April 2020

Sodium glucose cotransporter 2 inhibitors (SGLT2i) emerged as promising antidiabetic drugs that decreasing glucose reabsorption as well as effectively reduce heart failure hospitalization and cardiovascular death in type 2 diabetes patients. Recently, it has been shown that the SGLT2i also exert beneficial cardiovascular effects in non‐diabetic heart failure patients. However, the cardioprotective mechanisms of these drugs remain unknow. The present study aimed to test the hypothesis that the cardioprotective effects of SGLT2i empagliflozin (EMPA) may be associated with changes of cardiac electrical activity and Ca2+ homeostasis in hypoxia‐induced injury ventricular myocytes. Cardiomyocytes isolated from neonatal rats submitted to chemical hypoxia (200 μM CoCl2) or normoxia were incubated with EMPA (1 μM) or vehicle during 24 hours. Electrophysiological recordings were obtained using multielectrode array (MEA) system and intracellular microelectrode. Cytoplasmic calcium transients were investigated via fura‐2 fluorescence. Compared with vehicle, MEA analysis showed a significant decrease in field potential duration in EMPA‐group in both conditions [hypoxia: 83 ± 4 (N=19) vs. 37 ± 3 (N=44); normoxia: 213 ± 5 (N=53) vs. 62 ± 4 (N=36) ms, p<0.001]. EMPA reduced action potential duration at 30%, 50% and 90% repolarization (APD30, APD50 and APD90, respectively) in hypoxia [APD30: 109 ± 2 (N=41) vs. 98 ± 1 (N=30); APD50: 150 ± 3 (N=41) vs. 134 ± 1 (N=30); APD90: 292 ± 4 (N=41) vs. 271 ± 5 (N=30) ms, p<0.05] and normoxia [APD30: 94 ± 2 (N=30) vs. 81 ± 1 (N=39); APD50: 129 ± 2 (N=30) vs. 111 ± 1 (N=39); APD90: 259 ± 4 (N=30) vs. 244 ± 4 (N=39) ms, p<0.05]. No differences were observed in amplitude and resting membrane potential. Action potential recordings also revealed that the percentage of arrhythmic cells was lower in EMPA‐group in both conditions [hypoxia: 19 (N=41) vs. 10 (N=30) %; normoxia: 7 (N=30) vs. 3 (N=39) %]. In addition, EMPA significantly increased in Ca2+ transient amplitude in hypoxia [F/F0: 0.16 ± 0.01 (N=21) vs. 0.21 ± 0.01 (N=19), p<0.01] and normoxia [F/F0: 0.17 ± 0.01 (N=19) vs. 0.23 ± 0.01 (N=20) μM, p<0.01]. These results suggest that EMPA modulates electrical activity, reduces arrhythmic events and improves Ca2+ transient, which may contribute to cardioprotective effects of SGLT2i.

Support or Funding Information

FAPESP and CNPq
Comparison of Neurotransmitter Release from Perivascular Nerves in C57BL/6J Mice and Wistar Rats
Authors: Mittal Astha , Mitchell Ray , Fang Hanwei , Bagher Pooneh
Published: April 2020

It is thought that sympathetic nerve activation stimulates arterial constriction predominately through the release of three neurotransmitters: norepinephrine (NE), adenosine triphosphate (ATP) and neuropeptide Y (NPY). The aim of the current study was to examine the composition of neurotransmitter released in response to sympathetic nerve stimulation in mesenteric arteries of two commonly used wild‐type animal models. Isolated mesenteric arteries from male C57BL/6J mice and Wistar rats were mounted in a Danish Myotechnology 2‐Channel Wire Myograph System and isometric force measurements were recorded. Electrical field stimulation (EFS) was used to stimulate sympathetic perivascular nerves (PVNs) to release neurotransmitters in the presence and absence of the adrenergic receptor blocker phentolamine and/or purinergic receptor blocker NF449. Phentolamine alone blocked a significant proportion of the contractile response evoked by EFS in both mice and rats, suggesting that NE is the main sympathetic neurotransmitter in both species. Treatment with NF449 alone blocked a small proportion of the contractile response evoked by EFS in mice but did not have a significant effect in rat, suggesting that ATP may play a more important role in mice. The application of both phentolamine and NF449 nearly completely blocked the contractile response to EFS in mouse mesenteric arteries; however, in rat, a proportion of the contractile response, roughly equivalent to the response obtained in the presence of phentolamine alone, was still observed. These data suggest that the neurotransmitter composition is different between mice and rats, in particular mouse mesenteric sympathetic PVNs release NE and ATP while rat mesenteric sympathetic PVNs release NE and another non‐purinergic neurotransmitter(s).
Anxiolytic Effects of Cannabinoid Receptor Agonists in the Zebrafish Species, Danio rerio
Authors: Prasad Aleya , Crowe Macey , Burton Grace , McGrew Lori
Published: April 2020

As of July 2019, 34 states in the U.S. have legalized medical marijuana and an additional 12 have legalized “low THC, high cannabidiol (CBD)” substances (Hanson & Garcia, 2019). While acute cannabis use has been associated with decreased anxiety levels, users have also experienced adverse side effects including increased anxiety, panic reactions, and psychotic symptoms (Hall & Degenhardt, 2009). Plant‐derived pharmacological agents, like cannabis, can vary in quality and composition (and thus efficacy) due to a variety of environmental factors (Atanasov et. al. 2015). In order to use cannabinoids to treat anxiety safely, a synthetic cannabinoid that can produce consistent therapeutic (anxiolytic) effects without producing psychoactive effects is necessary. Zebrafish are an increasingly popular model organism used due to their high sensitivity to chemicals as absorbed through their gills and quantifiable behavioral phenotypes (Stewart 2014). Humans and zebrafish have comparable endocannabinoid pathways that each contain CB1 and CB2 receptors (Krug and Clark 2015). When observed in a Novel Dive Tank, anxiety levels in zebrafish can be quantified. Like humans, male and female zebrafish differ in hormonal composition and therefore respond to treatments differently. In this study, male and female zebrafish were treated with CB receptor agonists anandamide and WIN 55,212‐2 and CB2 inverse agonist JTE‐907 in order to model the how the endocannabinoid system influences anxiety. The reduced amount of time spent in the upper zone by the JTE + Anandamide treated fish, combined with the increased time classified as highly mobile, suggests that JTE‐907 had an anxiogenic effect. Significant anxiolytic effects were observed the WIN treatment group, particularly in male treated fish. These results suggest that male treated fish are more receptive to the anxiolytic effects of a non‐selective CB agonist such as WIN 55, 212. Future research with a water‐soluble CB1 inverse agonist could provide insight into the differing roles of CB1 and CB2 receptors in cannabis‐based treatments.
17β‐Estradiol Induces Temozolomide Resistance through Redox Optimization in Glioblastoma
Authors: Lin Hong-Yi , Hsu Tsung-I , Ko Chiung-Yuan
Published: April 2020

Glioblastoma is the most fatal cancer of all brain tumors, and the standard treatment of glioblastoma patients is surgical tumor resection followed by radiotherapy and temozolomide (TMZ)‐mediated chemotherapy. According to the TCGA databases, female patients exhibit poorer prognosis compared with male patients, suggesting that the gender‐specific hormone plays an important role in glioblastoma malignancy. In addition to acting as a female sex hormone, 17β‐estradiol is an important neuro‐steroid for brain health, including learning and memory. Numerous studies indicate that 17β‐estradiol aggressively participates in the pathogenesis of endocrine‐related cancers, such as breast and prostate cancers. However, the role of 17β‐estradiol is studied poorly in glioblastoma which exhibits strong neurosteroidogenesis. In this study, we attempt to elucidate whether 17β‐estradiol is involved in acquired resistance of glioblastoma to TMZ. Herein, we found that the level of 17β‐estradiol is significantly increased in TMZ‐resistant glioblastoma. Moreover, 17β‐estradiol attenuated TMZ‐induced cell death in A172 and U87MG cells. To dissect mechanisms underlying 17β‐estradiol‐induced resistance, we investigated whether 17β‐estradiol affected TMZ‐induced DNA damage and ROS production. Particularly, 17β‐estradiol significantly attenuated TMZ‐induced ROS production and hydrogen peroxide accumulation without affecting DNA damage. Further, 17β‐estradiol obviously prevented TMZ‐induced mitochondria dysfunctions characterized by the elevation of mitochondrial ROS and the reduction of ATP production. In addition, 17β‐estradiol treatment increased expression of multiple anti‐oxidant proteins, including superoxide dismutase (SOD) 1, SOD2, catalase (CAT), glutathione reductase, and nuclear factor erythroid 2 like 2. In particular, 17β‐estradiol promoted degradation of hydrogen peroxide through activating CAT, not through glutathione‐ and redox‐in‐mediated catabolic pathways. Based on these findings, we conclude that 17β‐estradiol counteracts TMZ‐induced oxidative stress through enhancing mitochondrial activity and CAT, leading to TMZ resistance.

Support or Funding Information

Tsung‐I Hsu
Pharmacological activation of AMPK prevents aircraft noise induced oxidative stress, endothelial dysfunction and vascular inflammation
Authors: Kvandová Miroslava , Kalinovic Sanela , Schmal Isabella , Stamm Paul , Frenis Katie , Daiber Andreas , Oelze Mathias Matthias , Schulz Eberhard , Steven Sebastian , Jansen Thomas , Münzel Thomas , Kröller-Schön Swenja
Published: April 2020

Background

Aircraft noise exposure is responsible for the emergence of oxidative stress, endothelial dysfunction and induction of vascular inflammatory response leading to cardiovascular complications like heart attacks and strokes. The AMP‐dependent‐Kinase (AMPK) has important protective roles in the vasculature especially in situations of oxidative stress and vascular inflammation. In the present study, we investigated the activation of AMPK using daily AICAR injections as a preventive strategy against noise induced destructive effects.

Methods and Results

AICAR (5‐Aminoimidazole‐4‐carboxamide‐1‐β‐D‐ribofuranoside; =AMP analog) was used for stimulation of AMPK activity and administered daily by intraperitoneal injections (200 mg/kg/day) for 7 days. After 3 days AICAR pre‐treatment, the C57BL/6j mice were exposed to aircraft noise (maximum sound level of 85 dB(A), average sound pressure level of 72 dB(A)), which does not impair hearing significantly.

We could show that aircraft noise exposure lead to a significant endothelial dysfunction, which was significantly improved by AMPK activation by AICAR. In addition, we observed that AICAR administration was able to normalize the aircraft noise induced blood pressure increment. Similarly, noise exposure induced oxidative stress as well as in whole blood samples was reduced by AICAR administration. Endothelial dysfunction and oxidative stress is often correlated with vascular inflammation processes. Induction of an inflammation (MCP‐1, PAI‐1, IL‐6) by aircraft noise exposure was abrogated using AICAR injection in the vasculature and brain. Furthermore, we could show that aircraft noise exposure induces circadian rhythm dysregulation, which can be reduced by AICAR administration.

Conclusion

We here present novel data of AMPK activation with AICAR as a protective strategy against negative effects of aircraft noise exposure. Our data suggest that activation of AMPK might be a future therapeutic strategy to prevent traffic noise‐induced cardiovascular complications.

Support or Funding Information

German Research Foundation ‐
Deutsche Forschungsgemeinschaft (DFG).
Characterization of HIV‐1 Unmyrostolated Gag Polyprotein Interactions with the MAL HIV‐1 Genome
Authors: Arefeayne Nahum Fekadu , Swanson Canessa , Getachew Noel , Lafaver Brittany , Gilchrist Darin , Chioma Kennedy , Summers Michael
Published: April 2020

Human Immunodeficiency Virus (HIV‐1) is a retrovirus that targets CD4 T‐cells within the immune system which leads to compromising the immune system and effectively resulting in the development of Acquired Immune Deficiency Syndrome, AIDS. Current antiretroviral gene therapies target areas of the HIV‐1 replication cycle that are susceptible to mutations, resulting in reduced efficiency of these treatments. Also, the cocktail of drugs that are typically prescribed to patients can result in adverse effects such as drug toxicity after long‐term use. There is a need for a new drug target that targets a highly conserved region of the viral replication cycle to allow for more effectivity in treatment. One region of the replication cycle known to be highly conserved is genomic recognition where the viral genome is recognized and bound by the viral Gag polyprotein. This highly conserved step stems from the minimal packaging region of the genome termed the Core Encapsidation Signal (CES) which is a portion of the viral genome located in the 5′‐Leader (5′‐L). It is proposed that the CES selectively recognizes and promotes the formation of Gag hexamers which become form the viral particle core. This research aims to understand the interactions of the CES and Gag polyprotein through the use of electrophoretic mobility shift assays (EMSAs) as well as isothermal titration calorimetry (ITC). These experiments allow for the identification of binding sites that may be occurring between the viral genome and the Gag proteins. Initial ITC data has shown CES to have seven binding sites however, this was only done with the Nucleocapsid (NC) domain of the Gag polyprotein, which is only a small domain of the Gag that only plays a role in RNA binding. Whereas the other structural domains play a role in packaging, assembly, and Using the full viral protein allows for a more biologically relevant evaluation for understanding the viral packaging cycle. In addition to EMSAs and ITC experiments, the future goal is to obtain a structure of the nucleation complex through the use of x‐ray crystallography and cryogenic electron microscopy. By understanding the protein‐RNA interaction that occurs during the formation of the nucleation complex of the virus and eventually elucidating the structure of this complex, antiretroviral therapies can begin to target this highly conserved region of the replication cycle.

Support or Funding Information

This research was funded by NIH/NIGMS grants RO1OM042561, 8RO1AI50498 and NIH/NIGMS MARC U*STAR T34 HHS 00026 National Research Service Award to UMBC. This research was supported in part by a grant to UMBC from the Howard Hughes Medical Institute through the HHMI Adaptation Project. We would like to thank the MARC U*STAR program and UMBC Summer Biomedical Training Program for the strong support.
Cervix Pathology For Medical Students: Morphing‐Based Animation
Authors: King Thomas Scott , Nelson Erin , Sharma Ramaswamy
Published: April 2020

Cervical cancer is the third most common cancer in US women, resulting in approximately 4,000 deaths per year. Preventative screening of women will be part of the future clinical practices of many of our medical students. Therefore, microscopic examination of cervical pathology is an important component of histopathology teaching laboratories. Traditionally, students examine the normal cervical structure, including the transformation zone within which most cervical squamous cell carcinomas arise. Presentation of these topics may be typically incongruous in that normal and abnormal are taught sequentially but as separate topics and in two‐dimensional formats (i.e., histopathology micrographs). It is all too often incumbent on the student to “connect the dots”, and conceptualize the progress from the normal exocervix to metaplasia, dysplasia and ultimately neoplasia, within the “transformation zone”. Visualization of this process is not always well‐understood by all students. We hypothesized that students will assimilate pathological processes more quickly and effectively when these are viewed in transition, that is, by “morphing” from one stage of development to the next.

We utilized morphing software to animate a series of drawings of preadolescent to post‐adolescent cervix with eversion and development of the transformation zone (squamous metaplasia). The three‐dimensional animations continued through dysplastic and neoplastic development. The video included colposcopic images of the transformation zone, and demonstrated findings of ectropion, dysplasia and neoplasia in the cervical surface. Morphing sequences were created using software that calculated multiple transitions by the processes of warping and cross‐dissolving between corresponding frames and generated an output movie by using a single intermediate morph from each pair along the sequence. Images were processed as three‐dimensional signals with third‐dimension being time. The resultant animated and morphing video was disseminated as AVI movie format for Windows applications, as animated GIF and Flash animation for WEB pages and PowerPoint presentations or to QuickTime format for Mac computers. The video is currently an integral component of the curriculum for second year students at our university and has been very well‐received by these students in each of the last two years.

Analysis of the effectiveness of the video in facilitating learning is ongoing over a two‐year period. We are currently comparing two study groups of students using a self‐paced, formative assessment quiz to evaluate the student’s understanding of the cervical transformation zone and its clinical significance. Participating students are also asked to complete a brief survey addressing the overall usefulness of the video and perceived effectiveness of the video in gaining understanding of cervical transformation zone and its clinical importance.
Investigating the Role of Myo/Nog Cells in Tissue Injury in the Liver and Stellate Cell Cultures
Authors: Delancy Matthew M. , Milton LaBraya , Gerhart Jacquelyn , Abdalla Fathma , George-Weinstein Mindy , Bhave Vishakha , Behling Kathryn C.
Published: April 2020

Myo/Nog cells, named for their expression of the skeletal muscle specific transcription factor Myo D, and the bone morphogenic protein inhibitor noggin, are critical regulators of normal development, progenitors of myofibroblasts in the ocular lens and neuroprotective in the retina. They also rapidly respond to wounding of the skin. The role of Myo/Nog cells in the modulation of tissue injury and fibrosis in the liver has not been studied. We examined the response of Myo/Nog cells to acetaminophen induced acute liver toxicity and their relationship to human hepatic stellate cells already known to play a role in acute and chronic liver injury responses. Liver tissue was obtained from mice following acetaminophen overdose over a time course of 0 to 96 hours. Additionally, LX‐2 human hepatic stellate cells were grown to confluence and subjected to a mechanical injury (scratch assay) and examined at several time points 0–24 hours after wounding. Immunofluorescent staining with the G8 monoclonal antibody was used to identify Myo/Nog cells in tissue sections of liver and stellate cell cultures. Preliminary analyses showed that in the mouse model of acetaminophen toxicity, there is a resident population of Myo/Nog cells in the liver that decline in number during the first 24 hours after the acetaminophen overdose and then increase, particularly in the region near the central vein (zone 3) where hepatocyte necrosis was greatest. The highest number of Myo/Nog cells was seen at 48 hours post acetaminophen administration, the time of maximal liver injury as previously documented by serum biomarkers. With the in vitro human stellate cell scratch assay, we found that a subset of these cells express the Myo/Nog marker G8, and the proportion of cells that express this marker increases after wounding, peaking at 24 hours after the scratch. G8 positive cells consistently clustered in proximity to the margins of the wound, with the greatest densities counted within the first high‐power field of the scratch. These studies suggest that Myo/Nog cells respond to acute liver injury by increasing in number and/or migrating toward the site of injury. Whether these cells play a role in mediating progression of liver injury or promote recovery via liver regeneration needs to be further evaluated in the model. The in vitro studies show that Myo/Nog cells may represent a subpopulation of human stellate cells in the liver, and this relationship should be further explored as a potential target for future therapeutics for acute and chronic liver injury.

Support or Funding Information

Anonymous donation from a private funder.
NON‐PSYCHOACTIVE CANNABINOIDS (CBD/CBG) ACT VIA CANNABINOID CB2/CB1 RECEPTORS TO REGULATE INTESTINAL MYOFIBROBLAST METABOLIC ACTIVITY AND TO INHIBIT FORSKOLIN‐MEDIATED ELEVATION OF CYCLIC‐AMP
Authors: Venu Vivek Krishna Pulakazhi , Saifeddine Mahmoud , Mihara Koichiro , Kamal Brishna , Lantela Daniel , Hollenberg Morley D.
Published: April 2020

Intestinal fibrosis, a common complication of inflammatory bowel disease (IBD), comprises an excessive accumulation of scar tissue in the intestinal wall. Fibrosis is thought to result from an aberrant response to injury resulting in excessive extracellular matrix (ECM) deposition, which can lead to tissue stiffening and intestinal stricture formation. Activation of the cannabinoid‐receptor‐CB2 has been reported to regulate mesenchymal cell function and to dampen fibrogenic signaling. More recently, non‐psychoactive cannabinoids, Cannabdiol (CBD) and Cannabigerol (CBG), have been found to be associated with anti‐inflammatory signalling that dampens inflammation. Here, we tested the hypothesis that CBD and CBG, via cannabinoid CB1/CB2 receptors, could function as potentially negative regulators of intestinal inflammation‐associated fibrosis through modulating intestinal myofibroblast function and triggering CB2/CB2 signalling. The effects of CBD and CBG on TGF‐β induced cell metabolic activity (MTT assay) were examined using cultured myofibroblast cells isolated from mouse and human intestine. Cannabinoid‐regulated signal transduction pathways were assessed using HEK‐293 cells transfected to express human CB2 and CB2 – red‐fluorescent‐protein‐tagged receptors. We found that at 48h and 72h TGF‐β induced cell metabolic activity was attenuated by 5% and 12% compared to TGF‐β‐treatment alone by increasing concentrations of CBG (10μM). Further, CBD (500nM) was able to inhibit TGF‐β induced cell metabolic activity by 12% and 25%, compared to TGF‐β alone at 48h and 72h. In CB1/CB2‐expressing HEK cells, preliminary data showed that both CBD and CBG, acting via CB2, were able to diminish forskolin (0.5 μM)‐increased cyclic‐AMP production, without affecting changes in intracellular calcium, indicating activation of Gi. In cells co‐expressing CB1/CB2 receptors, both CBG (100 nM) and CBD (20 nM) inhibited high forskolin concentration(5–10 μM)‐stimulated cAMP production. Nontransfected HEK cells showed no responses to either CBD or CBG. We conclude that the non‐psychoactive cannabinoids, CBD/CBG can regulate non‐neuronal cells, including intestinal myofibroblasts and kidney HEK cells via both CB2 and CB1 receptors.

Support or Funding Information

Supported by grants from the Canadian Institutes of Health Research for MDH and Mitacs postdoctoral fellowships for VKPV
Sex Differences in Renal T cells in DOCA‐salt Hypertension are not Blood Pressure Dependent.
Authors: Belanger Kasey , Sullivan Jennifer
Published: April 2020

Background

There is increasing evidence implicating T cells in the development and maintenance of hypertension. Our lab has previously shown that there is a sex difference in renal T cells in deoxycorticosterone acetate salt (DOCA) hypertension, with females having greater T regulatory cells than males. Moreover, Tregs attenuate DOCA‐induced increases in blood pressure (BP) only in female rats. Therefore, the goal of the current study was to test the hypothesis that the sex difference in renal Tregs in DOCA‐salt rats is BP dependent.

Methods

At 10 wks of age, male and female Sprague‐Dawley rats were anesthetized, a right uni‐nephrectomy was performed, and all rats were implanted with telemetry devices for the continuous measurement of BP. After one week of recovery, all rats received a subcutaneous 21‐day slow‐release DOCA pellet (200 mg) with 0.9% saline water to drink. To prevent DOCA‐salt induced increases in BP, a subset of rats (N=9–12/group) were randomized to receive hydrochlorothiazide (HCTZ; 55 mg/kg/day) and reserpine (4.5 mg/kg/day) combined with 0.9% saline in the drinking water for 3 weeks. Rats were individually housed throughout the study. Water intake and body weights were measured every two days and the dose of drugs were altered, as needed, to maintain baseline BP levels. After 3 wks of treatment, the remaining kidney was isolated for flow cytometric analysis of T cells.

Results

After 3 wks of DOCA‐treatment, males had a higher BP than females (179 ± 3 mmHg vs. 157 ± 4 mmHg; n=6–7; Psex=0.0001). Treatment with HCTZ and reserpine prevented DOCA‐induced increases in BP in both males (110 ± 5 mmHg) and females (101 ± 5 mmHg; Ptreatment<0.0001; Pinteraction=0.307). With DOCA‐salt induced hypertension, males had more renal CD4+ T cells (expressed as %CD3+ T cells: 63 ± 2 vs 60 ± 1) and Th17 (expressed as %CD3+CD4+ cells: 14 ± 1 vs 11 ±1) than females (Psex=0.04). HCTZ and reserpine decreased both CD4+ cells (expressed as %CD3+ T cells: 61 ± 3.9 vs 60 ±2.9; Ptreatment<0.0001) and Th17 cells (expressed as %CD3+CD4+ cells: 5.15 ±0.93 vs 5.1 ± 1.3; Ptreatment<0.0001) and the decrease was comparable between sexes (CD4+: Pinteraction=0.86; Th17: Pinteraction=0.55). With DOCA alone, females had a higher number of Tregs than males (expressed as %CD3+CD4+ cells: 3.9 ±0.21 vs 2.6 ±0.2; Psex<0.0001). Preventing DOCA‐salt induced increases in BP with HCTZ and reserpine did not significantly change renal Tregs in either sex (Ptreatment=0.11; Pinteraction=0.38).

Discussion

Our data suggests that while preventing DOCA‐salt induced increases in BP attenuates the increase in renal pro‐inflammatory T cells, sex differences in renal T cells is not dependent on hypertension.
Examination of Cortactin‐ and Membrane‐Associated Proteins at Listeria Membrane Protrusions
Authors: Bruzzini Kaitlin M. , Guttman Julian A.
Published: April 2020

Listeria monocytogenes is an intracellular pathogen that manipulates its host’s cytoskeleton to polymerize actin together with an array of actin‐associated proteins, including VASP and cortactin to one pole of the bacteria forming comet/rocket tails for intracellular propulsion. These structures also enable the microbes to move from one cell to another through a cell‐to‐cell spreading process involving the bacteria pressing into the host’s plasma membrane, forming L. monocytogenes protrusions which are internalized into corresponding invaginations in neighbouring cells. Recently, a siRNA screen listed genes involved in comet/rocket tail formation and cell‐to‐cell spreading. Here we examined a subset of those proteins (Endophilin, Ephrin A2, Anti‐Ephrin B1) as well as cortactin‐associated proteins (FGD1, SHK, VAV2) by immunolocalization at actin‐rich structures generated during L. monocytogenes infections.
Muscle Function During Feeding Through Infancy
Authors: Mayerl Christopher James , Tobin Hieronymus L. , Chava Almasi M. , Edmonds Chloe E. , Gould Francois D. H. , Stricklen Bethany M. , Bond Laura E. , German Rebecca Z.
Published: April 2020

Infant mammalian feeding is a complex process that requires the integration of different behaviors and over twenty‐five muscles controlled by multiple cranial nerves. Despite extensive characterization of muscle activity during a feeding sequence, from suckling to swallowing, specific biomechanics of muscle function are unknown. The function of a muscle is determined by both when the muscle is active and the line of action of the muscle, as well as the change in length during activity. In mammals, there is potential for the line of action of the muscles involved in swallowing to change through ontogeny, as infants have a compressed oropharyngeal anatomy relative to adults. Additionally, while we understand when muscles are active during infant feeding, little has been done to understand whether those activity patterns correspond to concentric, eccentric, or isometric contractions. Here, we used contrast‐based microCT scanning coupled with electromyography (EMG) and fluoromicrometry to test the possibility that muscle function changes through infancy using our validated infant pig model. We collected anatomical data from pigs at infancy and just prior to weaning (21 days old), and isolated muscles of interest in Avizo. We collected EMG and fluoromicrometry data from five muscles involved in feeding (Geniohyoid, Digastric, Stylohyoid, Thyrohyoid, and Omohyoid) at the same ages. We found substantive changes in muscle anatomy through infancy and found extensive variation in muscle firing patterns and function within muscles. Some muscles (eg thyrohyoid) exhibited concentric contractions, whereas other muscles were eccentric (eg omohyoid). Still others exhibited multiple functions, and were eccentric during sucking EMG bursts, but isometric during swallowing EMG bursts (eg geniohyoid). Additionally, we found regional heterogeneity in muscle firing patterns in some muscles (eg stylohyoid), whereby electrodes placed in the middle of the muscle showed activity during swallows, but electrodes placed closer to the hyoid (more ventrally) recorded activity during sucking and swallowing. These results highlight the complexity of infant mammal feeding and demonstrate that muscle activity does not necessarily indicate muscle function. Instead, many muscles that are traditionally thought to function to pull the hyoid along their line of action may actually be functioning to stabilize the hyoid during swallowing. This suggests that control and coordination of the oropharyngeal structures is likely to be more important for successful feeding function than strength or the velocity of moving those structures.

Support or Funding Information

AAA Postdoctoral Fellowship
Role of Mechanical Force on the Growth Plate
Authors: Serra Rosa
Published: April 2020

The shape and size of the limb is determined by genetic, biochemical, and mechanical factors resulting from weight bearing activity or muscle contraction. Changes in bone shape in response to mechanical forces was first reported by Wolf, 1892. Observations by Hueter‐Volkman indicated that supraphysiological forces lead to reduced skeletal growth. More recently, Frost (1990) proposed the concept of chondral modeling to describe physiological loading. In this case, both tension and compression lead to increased skeletal growth up to a peak then compression beyond that peak leads to inhibition of growth. Recent experiments using zebra fish, chick, and muscle‐less Spd mice indicated a specific connection between mechanical load and endochondral bone formation in embryonic skeleton. A role for mechanical forces on postnatal skeletal development is apparent in clinical manifestations of children with hemiplegia resulting in unequal loading of developing limbs. A large proportion of children with hemiplegia demonstrated significant limb length discrepancy with the paralyzed limb being shorter than the other limb. An age dependent effect of force on limb development in mice during spaceflight and tail suspended rats indicated a critical developmental window where loading regulates growth plate function. Longitudinal growth of limbs occurs through cartilaginous structures called the growth plate in a process called endochondral bone formation. One important feature of the growth plate is that the cells align into columns that represent a continuum of differentiation. It was suggested that the magnitude of bone growth is dependent on chondrocyte proliferation, matrix deposition, and hypertrophy; however, without the columnar structure, growth would not be longitudinal and the proper length and shape of skeletal elements would be disrupted. Recent experimental data indicate that mechanical load is required to regulate growth plate function including proliferation of chondrocytes and column formation. The molecular mechanism governing proliferation and column formation in response to load are not known but may involve chondrocytes sensing mechanical changes in the environment thought their actin cytoskeleton. Unloading mouse hind limb results in disorganization of the actin structure in chondrocytes, loss of columnar structure in the growth plate, and reduced limb length. Mice with conditional deletion of genes associated with regulation of the actin cytoskeleton demonstrate similar alterations in their growth plates. These observations highlight the importance of mechanical loading on growth plate function but there is still limited understanding of the molecular mechanisms of how limb length is regulated in response to mechanical loads.

Support or Funding Information

NIH/NIAMS R21 AR070097 and the US‐Israel Bi‐national Science Foundation