bluuebunny/crossref_metadata_2025 huggingface dataset:
Serine 363 is Required for NOPR Desensitization, Internalization, and Arrestin Signaling
Authors: Zhang Nancy , Planer William , Siuda Edward R , Zhao Hu‐Chen , Stickler Lucy , Chang Steven D , Baird Madison A , Cao Yu‐Qing , Bruchas Michael R
Published: June 2021

Nociceptin/Orphanin FQ peptide receptor (NOPR) plays a key role in pain modulation, opiate tolerance, and responsivity to stress and anxiety. C‐terminal phosphorylation at serine (S), threonine (T), and tyrosine (Y) residues are required for μ and κ opioid receptor internalization, desensitization, arrestin recruitment, and mitogen‐activated protein kinase phosphorylation (pMAPK), so we generated mutations in the NOPR c‐terminus. We showed that NOPR‐YFP internalized shortly after nociceptin treatment while NOPR‐S363A blocked internalization. We determined time course and concentration‐dependence of NOPR‐YFP‐mediated pMAPK. NOPR‐YFP‐induced pERK peaked at 10min following treatment and pJNK peaked at 30min. NOPR‐S363A showed reduced pJNK levels. We determined that NOPR‐YFP internalization is blocked by arrestin3 and GRK3 shRNAs and that NOPR‐S363A internalization and pJNK can be rescued via a dominant positive arrestin3. These data implicate GRK/arrestin in NOPR MAPK signaling and highlight the potential for the development of functionally selective NOPR‐ligands. This work was partially supported by NIH grant R00DA034929 (to MRB), T32DA007261 (to SDC) and HHMI SURF Program (to NZ).
The science of repurposing drugs in Alzheimer's disease therapeutics: The tale of rexinoid receptor ligand IRX4204
Authors: Pasinetti Giulio Maria
Published: June 2021

Alzheimer's disease (AD) is a multifaceted disease with different pathological features. Retinoid X receptor (RXR) is a nuclear receptor that can be activated by 9‐cis retionoic acid. RXR agonists are emerging as a new class of agents capable of promoting Aβ clearance by enhancing expression of ApoE that traffics Aβ. IRX4204 is a rexinoid agonist with high potency and selectivity to RXRs and an extensive history of safety in oncology clinical trials. In this study, we tested the effect of IRX4204 on amyloid‐mediated neuropathology and cognitive impairments in an animal model of AD. We found that IRX4204 can effectively pass the blood brain barrier and accumulate in the brain at a concentration sufficient to activate the CREB signaling pathway in vitro. IRX4204 can interfere with Aβ and tau oligomerization, promote ApoE protein expression, and improve synaptic plasticity. Moreover, in vivo IRX4204 treatment can significantly improve cognitive function and reduce neuropathology in TgCRND8 mouse model of AD. Our study demonstrated IRX4204 can be potentially developed into a disease‐modifying agent and provided impetus for immediate testing of IRX4204 in clinical settings for AD prevention and treatment.
INTERLEUKIN‐10 DEFICIENCY LIMITS THE DEVELOPMENT OF OBESITY AND INSULIN RESISTANCE PRODUCED BY A HIGH FAT DIET
Authors: Faulkner Jessica L , Gomolak Jessica R , Didion Sean P
Published: June 2021

The goal of this study was to test the hypothesis that deficiency of immune‐regulating cytokine interleukin‐10 (IL‐10) promotes the development of obesity and insulin resistance. Male C57BL/6 (wild‐type, WT) and IL‐10 deficient (IL‐10−/−) mice were fed either 10% (control) or 60% kcal high fat diet (HFD) for 36 wks. A HFD produced obesity and increased adiposity in both WT and IL‐10−/− mice, however the increases were markedly less (P<0.05) in IL‐10−/−. In addition, the increase in adipocyte size with HFD was less in IL‐10−/− mice as compared to WT. A HFD was associated with the development of insulin resistance in WT mice but not IL‐10−/− mice (eg, glucose levels in WT HFD and IL‐10−/− HFD were 187±47 and 70±44 mg/dl at 120 min following a glucose challenge, respectively). Fasting plasma insulin levels were also higher in WT HFD compared to IL‐10−/− HFD mice (2.6±0.9 vs. 1.0±0.3 ng/ml, respectively). Expression of GLUT4, IRS‐1 and PPARγ in adipose was reduced to a similar extent in response to a HFD in both WT and IL‐10−/− mice. GLUT4 and IRS‐1 expression in skeletal muscle was unaffected by genotype and HFD. Contrary to our hypothesis, these data suggest that IL‐10 contributes to insulin resistance and adiposity produced by a HFD. NIH HL‐089884 & HL‐107632.
Anthropometric assessment of young children in Retalhuleu, Guatemala
Authors: Alfonso Victor , de Ponce Silvia , Barrantes Lilliam , Villanueva Lisa , Casasola David , Reinhart Gregory A.
Published: June 2021

An anthropometric assessment was conducted on 1238 children (age 6 mo–7 yr) from 18 rural communities in Retalhuleu, Guatemala. The anthropometry scan included collection of weight, height or recumbent length, mid‐arm circumference, age and gender. The data was processed using WHO Anthro, WHO Anthro Plus and Epi Info. 907 children (73.21%) had a height‐for‐age Z score (HAZ) of −1.0 or lower. Of these, 429 children had a HAZ of −1 to −1.99, 335 had a HAZ score of −2.0 to −2.99 and 143 had a HAZ of <−3.0. For weight‐for‐height Z scores (WHZ), 800 children (64.6%) were within the normal range (WHZ −0.99 to 0.99), 27 (2.2%) were overweight (WHZ 2.0 to 3.0) and 14 (1.1%) were obese (WHZ >; 3.0). 231 children (18.7%) had WHZ of −1.0 or lower. Of these, 159 children had WHZ of −1.0 to −1.99, 51 had WHZ of −2.0 to −3.0, and 21 had WHZ <−3. The assessment demonstrates widespread stunting in this population and suggests a potential underlying malnutrition problem. This region in southwestern Guatemala is not generally recognized as an area afflicted with extensive nutritional inadequacies or food insecurity. Further evaluation of nutritional status in this population is warranted. This study was supported by the Mathile Institute for the Advancement of Human Nutrition®.
Relationship between egg consumption patterns and nutrient intake, diet quality, weight measures, and cardiovascular risk factors (CVRF)‐2001–2008 NHANES
Authors: Nicklas Theresa A. , O'Neil Carol E. , Fulgoni Victor L
Published: June 2021

This study examined the association of egg consumption patterns (ECP) with nutrient intake, diet quality, adiposity, and CVRF in adults participating in NHANES 2001–2008. 24‐hour dietary recalls and cluster analysis determined the ECP; BMI and waist circumference (WC) assessed adiposity; CVRF were serum lipids/blood pressure/triglycerides/glucose/insulin. Covariate adjusted LS means ± SE were generated. Eight ECP were created (34.5% of the variance in total energy intake). ECP were compared to the no egg pattern, which was the most common ECP (80.2% of the sample) followed by the egg/meat, poultry, fish [MPF]/grain/potato/fruit juice/vegetable pattern (7.8%). 1–3% of the sample consumed one of the six remaining ECP. Most ECP resulted in higher energy intake. Several ECP showed lower intakes of added sugars, but higher intakes of saturated and solid fats. Total nutrient intake varied in the ECP; three ECP had lower diet quality. Two ECP were associated with BMI and WC; when food groups consumed in those ECP were covariates, only one ECP remained significant (egg/MPF/grain; (p≤0.0063). Only one ECP was positively associated with diastolic blood pressure and LDL‐C (both p≤0.0063). Relationships between food patterns and health outcomes are complex. More studies are needed to determine what foods drive these associations and how foods consumed at other meals influence results. Support: USDA & Egg Nutrition Board.
Relationship of Cocaine Use with Markers of Body Mass Index and Body Composition and HIV Disease Progression
Authors: Pedamallu Santhipriya , Campa Adriana , Lai Shenghan , Li Yinghui , Page John Bryan , Martinez Sabrina Sales
Published: June 2021

Objective

To examine the relationship of cocaine use with HIV disease progression and measures of nutritional status such as energy and protein storages.

Methods

After consenting 231 HIV+ adults, they were given questionnaires on socio‐demographics, ART use and type and frequency of drugs of abuse. Body Mass Index and bioimpedance measures (fat mass [FM], lean body mass [LBM] and body cell mass [BCM]) were calculated. Blood was drawn for CD4 cell counts and HIV viral load.

Results

Seventy three percent of the participants used cocaine or crack cocaine and 62% were on ART. Linear regression analyses showed that frequency of cocaine use had deleterious effect on BCM (β=−6.805, p=0.01), LBM (β=−9.583, p=0.003) and FM (β=−6.418, p=0.01), after controlling for age, gender and ART. Moreover, frequency of cocaine use predicted lower CD4 cell count (β=−1.545, p=0.047) and higher HIV viral load (β=0.199, p=0.048).

Conclusion

Our findings show that cocaine use is associated with parameters of nutritional status and more rapid disease progression. The additive or synergistic effects of these relationships warrant further investigation to use results for treatment and programmatic decisions.

Funded by NIDA
Cultures with Unique Nutrition Concerns: Lesbian, Gay, Bisexual, Transgender
Authors: Bilyk Hope T , Wellington Christine , Kapica Cathy
Published: June 2021

Learning Outcome: There is a lack of data on nutrition issues of concern to the Lesbian, Gay, Bisexual and Transgender community.

Lesbian, Gay, Bisexual and Transgender (LGBT) individuals include all races, ethnicities, religions, and social classes. 7–10% of Canadians and 4% of Americans identify as LGBT. LGBT experience unique health disparities, as noted in Healthy People 2020. With increasing awareness and acceptance of LGBT, more individuals are acknowledging their identity. This study assessed the literature on nutrition issues of relevance to LGBT, and the availability of appropriate nutrition education materials. PUBMED was searched using the terms female and male homosexuality, gay, bisexual, transsexualism, transgender, and nutrition and health from 1998–2012. None of the articles addressed nutrition issues. Lesbians and bisexual females are more likely to be overweight or obese, contributing to an increased risk of heart disease, diabetes, and certain cancers. Gay and bisexual males have a higher rate of eating disorders and body dissatisfaction. Transgendered undergoing hormone therapy are at higher risk for cardiovascular disease, decreased bone density, and impaired glucose metabolism, thus requiring specialized nutrition counseling. LGBT have the highest rates of abuse of tobacco, alcohol, and other drugs, which may affect their nutrition status. Healthcare providers need to be aware of the special nutrition needs of LGBT. Despite increased risks, there is little research done or nutrition education materials available to address these concerns.

Funding Disclosure: None
Astaxanthin decreases inflammatory biomarkers associated with cardiovascular disease in human umbilical vein endothelial cells
Authors: Chew Weslee , Mathison Bridget , Kimble Lindsey , Mixter Philip F , Chew Boon P
Published: June 2021

As a potent antioxidant, astaxanthin may downregulate inflammation‐associated factors involved in endothelial dysfunction and cardiovascular disease progression. We studied the possible protective effects of astaxanthin against endothelial dysfunction in human umbilical vein endothelial cells (HUVEC) induced with H2O2. Cells were pre‐incubated with 0, 0.01, 0.1, or 1.0 μmol/L astaxanthin for 48 h, then oxidative stress induced with 100 μmol/L H2O2 overnight. Uptake kinetics showed a time‐dependent accumulation of astaxanthin (1.0 μmol/L) by HUVEC over the 48 h period. Oxidative stress induction with H2O2 decreased intracellular antioxidant activity (SOD and GPx) and increased the production of reactive oxygen species (ROS), inflammatory cytokines and mediators. In contrast, pre‐incubation of HUVEC with astaxanthin prior to H2O2 stress increased (P < 0.05) SOD, but not GPx activity, and decreased the production of ROS, PGE2, LTB4, NO, IL‐8, and IFN‐γ. Pre‐treatment with astaxanthin also downregulated transcriptional activation of NF‐κB and activator protein‐1, thereby inhibiting downstream production of inflammatory cytokines and mediators. Therefore, astaxanthin inhibits inflammatory biomarkers upregulated by H2O2, likely by scavenging ROS required for transcriptional activation. Supported by the University of Washington and Washington State University.
Biliverdin attenuates LPS‐induced pro‐inflammatory cytokine expression in whole human blood
Authors: Bisht kavita , Wegiel B , Tampe J , Shing Cecilia M. , Wagner K.H. , Otterbein L , Bulmer A.C.
Published: June 2021

Objective

Inflammation is clearly implicated in the pathogenesis of cardiovascular disease, sepsis and respiratory disorders. The present study aimed to reveal the effect of biliverdin (BV) on the expression of pro/anti‐inflammatory cytokines in whole human blood.

Methods

Eight healthy males were recruited and 50 mLs of blood was collected into EDTA vacutainers. Whole blood was supplemented with BV (10 or 50 μM) in the presence or absence of 3 μg/mL lipopolysaccride for 4, 8 and 24 hours. Total RNA was purified at 4 h and plasma was collected at 4, 8 and 24 hours. Gene and protein expression of pro‐inflammatory cytokines (TNF‐α, IL‐1β, IL‐6, IL‐1Ra and IL‐10) were analyzed using qRT‐PCR and multiplex Luminex analysis.

Results

Biliverdin inhibited LPS‐mediated expression of IL‐1 β and IL‐1Ra in a dose dependent manner, showing a 50% decrease at 4 h (50 μM, P<0.05). However, BV showed only a slight non‐significant decrease (69 %) in IL‐6 expression under the same conditions while no significant effect was found for TNF‐α and IL‐10 expression.

Conclusion

Biliverdin reduces LPS‐induced expression of IL‐1 β and IL‐1Ra in whole human blood. This study is the first translational evidence to support BV's use as anti‐inflammatory agent in humans against LPS/sepsis.

Source of funding: FWF‐Austrian Science Fund (P21162‐K.‐H.W. and A.B.)
Angiotensin‐(1–7) stimulates angiogenesis in murine corpus cavernosum
Authors: Jarajapu Yagna , Wagh Anil , Pawar Rahul , Renownski Pawel , Sun Chengwen
Published: June 2021

Angiotensin (Ang)‐(1–7), a metabolite of Angiotensin‐converting enzyme‐2 (ACE2), is cardiovascular protective. Recent studies have implicated an important role of angiogenic progenitor cells (APCs) in the functions of Ang‐(1–7). This study evaluated the effect of Ang‐(1–7) in the tissue angiogenesis independent of APCs. Strips of corpus cavernosum (CC) or aortic rings were isolated from mice and cultured in matrigel. Formation of sprouts and tubes by endothelial cells (EC) were evaluated. In some experiments, assays were performed in CC‐strips from mice following treatment with Ang‐(1–7) (100μg/Kg, s.c., 4 weeks). ECs from sinusoidal CC have migrated into the matrix in 24 to 48 hours, and endothelial sprouts and tubes were observed in 3–5 days. Ang‐(1–7) treatment potentiated this response that was lower than the effect of VEGF (100nM). Effect of Ang‐(1–7) was inhibited by Mas receptor antagonist A‐779, and was not affected by nitric oxide synthase blocker L‐NAME. CC strips from Ang‐(1–7)‐treated mice have shown increased sprout and tube formations compared to the control. In mouse aortic rings, angiogenesis was observed in 3 to 5 days and the migration of smooth muscle cells was seen by day 4. Treatment with Ang‐(1–7) has no effect. These studies support that Ang‐(1–7) stimulates angiogenesis without the involvement of APCs by activating Mas receptor in the sinusoids of CC and not in a conduit blood vessel.
An exploration of sunitinb‐induced cardiotoxicity in male Sprague‐Dawley rats
Authors: Knapton Alan , Herman Eugene , Todd John , Estis Joel , Zhang Jun
Published: June 2021

Sunitinib (STB) inhibits tyrosine kinase and certain growth factors and has been associated with cardiotoxicity as an unexpected adverse effect. The characteristics of STB‐induced myocardial alterations have not been completely defined. The current study evaluated the effects of STB on serum troponin levels and myocardial histology in male Sprague‐Dawley rats (SD). Groups of 7–9 SD received daily oral doses of 12.5, 25.0 or 50.0 mg/kg STB for 14 days. Tissue and blood samples were taken 24 hours after the last dose. Serum levels of cardiac troponin I (cTnI) were monitored using the Erenna immunoassay system. STB‐induced changes in myocyte morphology (autophagy, necrosis and apoptosis) were evaluated (blinded) by light microscopy using a 0–3 scale. The most frequent alteration was an increased presence of autophagic vesicles. Mean autophagy lesion scores of 1.1, 1.6 and 2.5 occurred at doses of 12.5, 25 and 50 mg/kg, respectively. Myocyte necrosis and apoptosis were detected mainly after the high STB dose (mean severity scores of 1.5 and 1.4, respectively). Minor changes in cTnI levels occurred only at 50 mg/kg STB (3.3 vs 6.3 pg/ml in control and treated SD, p >; 0.05). These results indicate that STB induces myocyte alterations over a 2 week treatment period and the most prominent effect, increased autophagy, was not associated with significant changes in serum levels of the cardiac biomarker cTnI.
Pharmacological characterization of GPR120 ligands
Authors: NAYAK SURENDRA KUMAR , Adem Amina S , Rasoul Bejan , Polgar Willma , Zhou Wei , Gong Leyi , Thomsen William J
Published: June 2021

Omega‐3 fatty acids (ω‐3 FAs) and other long chain free fatty acids activate GPR120, which has been shown to mediate some of the anti‐inflammatory effects of ω‐3 FAs. Loss‐of‐function mutation in human GPR120 confers increased risk for obesity and GPR120 deficient mice develop diet induced obesity, glucose intolerance and fatty liver disease. The goal of the current study was to understand GPR120 ligand selectivity by testing agonist activity of various fatty acids in cell based assays to measure G‐protein activated second messengers, β‐arrestin recruitment and inhibition of cytokine production. HEK 293 cells over‐expressing GPR120 and Gα15 were used to detect agonist mediated IP‐1 production and measure intracellular Ca2+. β‐arrestin recruitment was measured using DiscoveRx enzyme complementation technology. GPR120 agonist mediated inhibition of LPS‐induced cytokine production was measured in the RAW 264.7 cells. We confirmed the previously reported agonist activity of ω‐3 FAs including DHA, EPA, α‐linolenic acid and other polyunsaturated fatty acids such as palmitoleate and oleic acid. However, in contrast to earlier reports ω‐6 FAs linoleic acid, arachidonic acid and γ‐linolenic acid were found to robustly activate GPR120 signaling. Further characterization of GPR120 ligands will help understand its physiological role and determine its potential as a therapeutic target.
Statin mediated transactivation of the ERK5 mitogen activated protein kinase
Authors: Chu Uyen Bao , Yang Jay
Published: June 2021

Statins are potent inhibitors of cholesterol biosynthesis and are clinically beneficial in preventing coronary heart disease. Independent of their lipid‐lowering effects, these compounds have been shown to improve endothelial function and inhibit the thrombogenic response. Here, we show that the MAP kinase ERK5 that functions both as a kinase and as a transcriptional co‐activator is transactivated by statins. We used a Gal4 DNA binding domain‐ERK5 fusion co‐expressed with a Gal4‐luciferase reporter to examine the mechanism of the statin‐mediated transactivation of ERK5. At 10 μM, cerivastatin, fluvastatin, pitavastatin, and simvastatin exhibited ERK5 transactivation. Western blots revealed that the statin‐mediated ERK5 transactivation is through phosphorylation of the TEY motif. Supplementing the statin‐treated cells with geranylgeranyl pyrophosphate abrogated transactivation suggesting that transactivation involves inhibition of protein prenylation by geranylgeranyl transferase I which prenylates the Rho/Rac/CDC42 family of GTPases. Microarray analyses are underway to examine the downstream targets of statin‐mediated ERK5 transactivation via the Rho/Rac/CDC42 ERK5 axis. These data will aid in the future design of therapeutics mimicking the beneficial effects of ERK5 activation such as endothelial cell protection and enhanced recovery from myocardial ischemia.
Specificity of cocaine‐induced dopamine‐independent sigma agonist self‐administration
Authors: Hiranita Takato , Paul Soto L. , Tanda Gianluigi , Katz Jonathan L.
Published: June 2021

Sigma1 receptors (σ1Rs) are intracellularly‐mobile chaperone proteins implicated in several diseases, as well as psychiatric disorders and substance abuse. A previous study showed that cocaine self‐administration (SA) induced dopamine (DA)‐independent reinforcing effects of selective σ1R agonists. The present study assessed whether the induction was specific to SA of cocaine. Selective σ1R agonists (PRE‐084, (+)‐pentazocine) lacked reinforcing effects in drug‐naive rats. SA was not obtained with 28 experimental sessions with the opportunity to selfadminister either σ1R agonist. The same exposure to cocaine was sufficient for its SA, after which robust SA of σ1R agonists was obtained. As with cocaine, SA of the indirect‐acting DA receptor agonist d‐methamphetamine induced the reinforcing effects of PRE‐084. In contrast, experiences with food reinforcement, or SA of (−)‐heroin or (±)‐ketamine were ineffective as inducers of PRE‐084 reinforcement. The results indicate that experience with indirect‐acting DA receptor agonists induces reinforcing effects of previously inactive σ1R agonists whereas reinforcement through different modalities is ineffective. It is further suggested that induced σ1R mechanisms may play an essential role in treatment‐resistant stimulant abuse, suggesting new approaches for the development of effective medications for stimulant abuse. Supported by NIDA IRP.
Attenuation of methamphetamine‐induced neurotoxicity by selective sigma receptor ligands: in vivo and in vitro studies
Authors: Matsumoto Rae , Kaushal Nidhi , Robson Matthew , McCurdy Christopher , Coop Andrew
Published: June 2021

Methamphetamine (METH) is one of the most widely abused illicit substances worldwide. It is capable of causing hyperthermia and neurotoxicity after high and/or repeated dosing. Selective ligands for sigma receptors were developed by our research team and shown in preclinical studies in male, Swiss Webster mice to significantly attenuate hyperthermia as well as several classical markers of METH neurotoxicity in the striatum: depletion of dopamine and 5‐HT levels, reductions in dopamine and 5‐HT transporter expression. Use of an NG108–15 cell culture model system to further delineate mechanisms underlying the protective effects of the sigma ligands reveals significant attenuation of METH‐induced ROS/RNS and caspase dependent pathways. In contrast, METH‐induced endoplasmic reticulum stress pathways are less affected by intervention with sigma ligands in NG108–15 cells. The relative contribution of each of the sigma receptor subtypes in the neuroprotective effects are still being elucidated, with preliminary data suggesting the importance of the sigma‐2 subtype. Together, the data indicate that sigma receptor ligands can intervene at numerous mechanisms that contribute to METH neurotoxicity. [Funding from NIDA: DA013978, DA023205]
Effects of repeated treatment of phencyclidine on cognition and sensorimotor gating in female C57BL/6 mice of different ages
Authors: Zhu Shenghua , Wang Hongxing , He Jue , Wang Junhui , Kong Jiming , Li Xin‐Min
Published: June 2021

Many studies have indicated that glutamatergic N‐methyl‐D‐aspartate (NMDA) neurotransmission is disturbed in schizophrenia. Phencyclidine (PCP), a NMDA receptor antagonist, can reproduce schizophrenia‐like symptoms in both humans and rodents. This present study investigated the behavioural performances of mice at different ages after repeated administration of high doses of PCP and possible changes of brain apoptotic regulation proteins. 2‐ and 6‐month‐old female C57Bl/6J mice received daily i.p. doses of PCP (20 mg/kg) or saline for 7 d. Testing was performed 24 h after the last day of treatment. Both age groups treated with PCP produced similar impairments in spatial working memory performed in the Y‐maze task and sensorimotor gating deficits in a prepulse inhibition (PPI) task. After behavioural tests, mice were decapitated for western blot analyses. PCP treatment altered the ratio of an anti‐apoptotic Bcl‐2 family member (Bcl‐2) to a proapoptotic analogue (Bax) in the frontal cortex. Our results demonstrate that age does not alter the behavioural sensitivity of mice to the high doses of PCP treatment and PCP‐induced cognitive and sensorimotor gating deficits in mice might be associated with the alterations in the ratio of anti‐apoptotic/proapoptotic proteins. These results suggest that the high‐dose PCP model may have significant potential in characterizing the behavioural and molecular mechanisms underlying cognitive deficits seen in schizophrenia patients.
Metabolomic Response to Sorafenib Treatment in Human Hepatocellular Carcinoma Cells
Authors: Zhou Shufeng , Luo Ruijuan
Published: June 2021

Untargeted metabolomics has enabled comprehensive overview on physiological system and provides chances for new discoveries linking cellular pathways to biological mechanism, which also is a good platform for better understanding of drug action. In this study, we employed a metabolomics method to examine how the multikinase inhibitor sorafenib affects metabolic pathways in human hepatocellular carcinoma cell line. The results showed that sorafenib treatment in HepG2 cells caused remarkable changes of several metabolic pathways in a concentration‐dependent manner. At a low concentration of sorafenib, the glycerophospholipid metabolism was the most affected pathway, while at a higher concentration, the purine metabolism was most significantly altered. The significant decrease of GTP level after sorafenib treatment could be associated with the toxicity of this drug. These findings have provided insights into how sorafenib acts on distinct metabolic and biochemical pathways that are associated with its therapeutic efficacy and side effects. Metabolic profiling studies for clinically approved kinase inhibitors can serve as a powerful tool to facilitate the understanding of their mechanism of actions, molecular targets and the interactome.
The Coxsackievirus and adenovirus receptor interacts with PDZ3 of the cellular scaffolding protein MAGI‐1
Authors: Martin Sterling , Yan Ran , Sharma Priyanka , Kolawoe Abimobola , Hostetler Heather , Excoffon Katherine
Published: June 2021

The Coxsackievirus and Adenovirus receptor (CAR) is an essential protein that is involved in cell‐cell adhesion, protein trafficking, and viral infection. The major isoform of CAR is selectively sorted to the basolateral membrane of polarized epithelial cells where it co‐localizes with the cellular scaffolding protein membrane‐associated guanylate kinase with inverted domain structure‐1 (MAGI‐1). We hypothesized that CAR directly interacts with MAGI‐1 via a PDZ‐based interaction and these two proteins interact in polarized epithelial cells. Each MAGI‐1 PDZ domain and the C‐terminus of CAR were subcloned into expression vectors and direct interactions were investigated by yeast‐2‐hybrid (Y2H) analysis and fluorescence resonance energy transfer (FRET). Interaction was investigated in polarized Madin‐Darby canine kidney (MDCK) epithelial cells. Y2H showed that CAR interacts with the third PDZ domain of MAGI‐1 (PDZ3) and FRET showed a Kd of 2.0 ± 0.9 nM. MAGI‐1 and CAR co‐immunoprecipate from polarized MDCK cells. These results show that CAR and MAGI‐1 interacts directly and this interaction occurs in polarized cells. Future work will investigate potential therapeutic interventions that may disrupt or enhance CAR‐MAGI‐1 interactions and alter epithelial susceptibility viral infection.

This work was supported by NIH grants R15AI090625 (KE), DK77573 (HH) and R25 GM086257.
Mucosal Expression of the Cytochromes P450 (CYP) and Nucleotide Kinases Involved in the Biotransformation of Drugs Used in Human Immunodeficiency Virus (HIV) Pre‐exposure Prophylaxis (PrEP)
Authors: To Elaine , Bumpus Namandjé N
Published: June 2021

Attempts to prevent HIV infection through PrEP have included topical application of anti‐HIV drugs to the mucosal sites of infection. However, clinical studies have revealed marked anatomic variability in drug exposure in vaginal tissue versus colorectal tissue. Thus, studies were performed to determine whether the CYPs and nucleotide kinases responsible for the metabolism and activation of drugs used in HIV PrEP were differentially expressed in human vaginal versus colonic tissue. Quantitative reverse transcriptase‐PCR revealed that colonic mRNA expression of CYPs 1A1, 2B6, and 3A4/5 were 14‐, 37‐ and 4‐fold greater, respectively, than vaginal mRNA. Immunoblotting confirmed the presence of CYPs 3A4 and 3A5 protein in vagina. The vaginal mRNA abundance of adenylate kinase 2, the primary enzyme responsible for the activation of the HIV PrEP drug tenofovir, was 1.5‐fold greater than in colonic tissue. Cytidine monophosphate kinase mRNAs were 8‐fold greater in colonic tissue and phosphoglycerate kinase mRNAs were 4‐fold greater in vaginal tissue. These kinases have been proposed to activate emtricitabine, which is also under development as a topical microbicide for HIV PrEP. These data indicate that enzymes that modulate exposure to anti‐HIV drugs may be differentially expressed in mucosal tissues that are sites of topical drug application for PrEP. Funding: NIH U01 AI068613 (HIV Prevention Trials Network).
A role for fibronectin matricryptic site signaling in short‐term responses of endothelium to flow
Authors: Okech William , Sarelius Ingrid H
Published: June 2021

Endothelial cell (EC) responses to mechanical forces exerted by flowing blood are well established; a principal response is realignment of the EC cytoskeleton in the direction of flow. The existing paradigm holds that this realignment is signaled directly from changes in perceived wall shear stress by mechanosensors (e.g. integrins and the glycocalyx) on the EC surface. We sought to determine whether short‐term responses of the EC cytoskeleton to flow have an ECM fibronectin (FN) ‐dependent component. Human umbilical vein endothelial cells (HUVEC) were cultured in perfusable microslides and subjected to flow (10 dyn/cm2 for 4 hrs) in the presence or absence of 9D2 (an anti‐FNIII‐1 mAb that blocks FN matrix assembly). HUVECs were then fixed and labeled for FN and F‐Actin and immunofluorescence images from 20 sites in 4 microslides per condition were obtained. In ECs incubated with 9D2 for 2 hrs prior to flow, there was a decrease in F‐Actin alignment compared to non‐treated controls (42.04 ± 0.37° vs 35.02 ± 0.38°, p< 0.01). This finding demonstrates that when FN signaling is blocked by 9D2, realignment of F‐ Actin stress fibers is significantly attenuated. Therefore, these data suggest that ECM FN plays a role in regulating F‐Actin realignment of endothelial cells due to flow. Supported by NIH HL105909
Ca2+‐activated K+ channels of small and intermediate conductance control the activation of endothelial cell connexin‐ and pannexin‐formed hemichannels
Authors: Lillo Mauricio , Gaete Pablo , Puebla Mariela , Figueroa Xavier
Published: June 2021

Nitric Oxide (NO), Ca2+‐activated K+ channels (KCa) of small (SKCa) and intermediate (IKCa) conductance and hemichannels formed by connexins (Cx‐HC) or pannexins (Panx‐HC) are involved in the control of vasomotor tone. However, the mechanisms of Cx‐ and Panx‐based hemichannel activation in endothelial cells are unknown. In mesenteric vessels, we evaluated the expression and activity of Cx‐HC and Panx‐HC by immunofluorescence, biotinylation of membrane proteins of endothelial cell luminal surface and ethidium uptake. Cx37, Cx40, Cx43 and Panx‐1 were detected in endothelial cells. Stimulation with acetylcholine (ACh) or the NO‐donor S‐nitroso‐N‐acetyl‐D‐penicillamine (SNAP) increased ethidium uptake in endothelial cells, which was abolished by the NO synthase inhibitor NG‐nitro‐L‐arginine (L‐NA) or by the Panx‐HC blockers probenecid and Panx10 and attenuated by the Cx‐HC blocker La3+. In addition, ACh‐activated ethidium uptake was also abolished by blockade of KCa with tetraethylammonium (TEA) or IKCa with charybdotoxin (ChTX) or triarylmethane‐34(TRAM‐34), but only attenuated by the SKCa blocker apamin. TEA did not affect the SNAP‐activated ethidium uptake. Treatment with TEA, ChTX or apamin also inhibits the ACh‐induced NO production. These results indicate that SKCa and IKCa regulate the activity of Cx‐HC and Panx‐HC through the modulation of NO production.

FONDECYT 1100850, ANILLOS ACT‐71
Reponses to IL6 and IL8 are blocked by BQ788 in Cremaster and Mesentery of control and diabetic mice
Authors: Vigilance Jacqueline , Dewar Anthony M. , Frame Mary D.
Published: June 2021

We examined responses to interleukin‐6 (IL6) and IL8 in the mesentery vs cremaster muscle of leptin deficient diabetic mice (db/db), compared to genetic back ground controls (C57BL/6). To determine whether the responses occurred through the endothelin‐ B (ET‐B) receptor, an ET‐B inhibitor, BQ788 was used. The cremaster or mesentery of anesthetized (5% isoflurane) mice (C57, N=25; db/db, N =16) were prepared for intravital microscopy. Arteriole (A) / venule (V) pairs were studied. Endothelial dysfunction was apparent in db/db, as diminished dilation to acetylcholine (33±8% of baseline, A) vs C57 (86±5%*). Maximal dilation (MAX) to IL6 was significantly less in V compared to A; in cremaster, MAX was decreased by half in db/db mesenteric A. EC50 in C57 cremaster A (−7±0.7 Log mg/ml) was 100‐fold greater than in C57 mesentery (−9±0.6*), db/db cremaster (−10±0.2*) or mesentery (−9±2*). MAX to IL8 was less in V compared to A; in cremaster, db/db did not respond to IL8, in mesenteric A, dilation was 50% less in the db/db v. C57. EC50 in C57 cremaster A (−7±0.2 Log mg/ml) was 100‐fold greater than EC50 of C57 mesentery (−9±1.2*) or db/db mesentery (−9±0.3*). BQ788 blocked dilations to IL6 and IL8. Thus vasodilatory responses to IL6 and IL8 are mediated by ET‐B receptors in the mesentery and cremaster. There is tissue specific distribution of responses, compromised response by the venules and in db/db vs C57BL/6. Thanks to NIH DK68401.
Sulforaphane alters cerebral leukocyte‐endothelial cell interactions post global ischaemia reperfusion
Authors: Gillespie Scarlett , Milton Nathaniel , Ashioti Maria , Gavins Felicity NE
Published: June 2021

Introduction

Sulforaphane (SFN) is a dietary available isothiocyanate, previously shown to have anti‐inflammatory properties. Since discovery of its ability to induce a battery of phase II detoxification enzymes, its potential as an anti‐inflammatory agent has been investigated in a variety of pathologies.

Aim

Investigate effect of SFN pre‐treatment on cerebral leukocyte‐endothelial cell (L‐E) interactions following bilateral common carotid artery occlusion (BCCAo).

Methods and Results

Male C57Bl6 mice, underwent 5 min BCCAo, followed by 2 h reperfusion, with/without SFN pretreatment. Little L‐E interactions were present in sham mice. BCCAo significantly increased in L‐E interactions, as measured using intravital microscopy. 24h pre‐treatment with SFN (5mg/kg) decreased these interactions, bringing the values back to levels observed in sham mice. These data are complemented with work from an in vitro oxygen glucose deprivation (OGD) model using SHSY5Y neuronal cell line. Preliminary results indicate SFN (6μM) reduces cell death associated with OGD challenge.

Conclusion

This data suggests that BCCAo leads to an inflammatory response within the cerebral microcirculation. Furthermore, pre‐treatment with SFN reduced L‐E interactions following pre‐treatment, suggesting its ability to protect the cerebral microcirculation from subsequent inflammatory challenges. This is further supported by the early results showing SFN's protection in the OGD model. These preliminary results suggest a potential therapeutic role for SFN in stroke.

Funded by the Medical Research Council, UK.
Three‐dimensional reconstructions of human embryos: vertebrobasilar system evolutive morphogenesis.
Authors: LABROUSSE Marc , LHUAIRE Martin , MICARD Émilien , SCHARBARG Emeric , CENDRE Romain , DELMAS Vincent , FELBLINGER Jacques , AVISSE Claude , BRAUN Marc
Published: June 2021

OBJECTIVES

The purpose of this study was to describe the morphogenesis of the vertebrobasilar system from three‐dimensional reconstructions of 4 human embryos, and to compare our results with literature.

MATERIALS AND METHODS

We investigated four embryos belonging to the French collection of Paris‐Descartes University: 14mm (Carnegie stage 17), 15mm (CS 18), 22.5mm (CS 21) and 27mm (CS 23) respectively, serially sectioned. Staining was Masson's trichrome. Scanning was performed using an Epson V750 PRO ®, (resolution 2400 to 3200 dpi). Registration of image stacks and export in DICOM files were performed using original software developed by IADI Laboratory (INSERM U947, NANCY). The images obtained were exploited with ORS ® Visual.

RESULTS

From the embryo of 14mm to 27mm, the vertebrobasilar vascularization was well visualized. Its collaterals were seen. The main relationships with the mesencephalon and rhombencephalon were analysed.

CONCLUSION

This original software allows a new method of three‐dimensional reconstruction, and a better understanding of vertebrobasilar system evolutive morphogenesis. It brings a new perspective to the study of human embryology.
Dietary Cocoa Reduces Adipose Tissue Inflammation in High‐ Fat Fed Obese Mice
Authors: Gu Yeyi , Harvatine Kevin , Lambert Joshua D.
Published: June 2021

In diet‐induced obesity, adipose tissue (AT) is in a chronic state of inflammation predisposing the development of insulin‐resistance, type 2 diabetes and cardiovascular complications. Cocoa (Theobroma cacao) is a polyphenol‐rich food with putative anti‐inflammatory activities; however, little research has been done to support the role of cocoa in modulating inflammation in obese subjects, and the mechanism of action remains to be fully elucidated. The objective of the present study is to determine the effects of dietary cocoa supplementation on AT inflammation in high‐fat (HF)‐fed mice. Male C57BL/6J mice were fed a HF diet (HF group), a HF diet with 8% (w/w) unsweetened cocoa powder (HFC group), or a low‐fat diet for 18 weeks. Cocoa supplementation significantly decreased pro‐inflammatory gene expression (Il6, Il12b, Nos2 and Emr1) in the stromal vascular fraction of AT by 40–62% compared to HF group (P < 0.05). Protein expression of adipose‐specific phospholipase A2 was reduced by 53% in HFC mice (P < 0.05). Moreover, AT arachidonic acid (AA) levels were positively correlated with adiposity (r = 0.56, P < 0.01) and cocoa‐supplemented mice had 33% lower levels of AA compared to HF control (P < 0.05). These results demonstrate the anti‐inflammatory effects of cocoa in obese mice and provide mechanistic biomarkers for further investigation.

Grant Funding Source: Dept. of Food Science of The Pennsylvania State University
Student Perceptions and Effectiveness of Team‐Based Learning in Anatomy
Authors: Palmer Phillip B
Published: June 2021

Team‐based learning (TBL), a method of instruction involving both independent study and group activities, is purported to improve student participation in learning, promote development of skills required for teamwork, and foster independent learning. Following examples in the literature, team‐based learning was implemented in a gross anatomy and embryology course for doctor of physical therapy students. The intent of the study was to assess students’ perceptions of TBL and to determine the effectiveness of this teaching approach when compared to a more traditional lecture‐ and lab‐based approach. Student perceptions were assessed utilizing a 17‐item questionnaire modified from an instrument designed by Vasan, DeFouw, and Compton. In general, the students perceived that TBL helped them increase understanding of anatomical content, develop information synthesizing skills, and to study more consistently. Perceptions concerning the importance of working as a part of a team were also positive, and students indicated that the team‐based activities enhanced their learning. A comparison between two cohorts of students (1 exposed to the traditional lecture/lab approach; the other participants in TBL) was conducted to assess the difference in student learning. Student scores on a set of 143 multiple‐choice test questions utilized in both classes were used for the comparison. No significant difference was found between the two cohorts (t(56)=1.085; p=.283). It appears that TBL is at least as effective as the traditional lecture/laboratory method in learning anatomical concepts. Also, students responded positively to the use of TBL as a teaching strategy and its use for teaching anatomy.
Anti‐cancer Property of Aegle marmelos Leaves: Finding the Facts Against Hepatocellular Carcinoma HepG2 cell.
Authors: Alshatwi Ali A. , Hasan Tarique N , Syed Naveed A , Alagal Reham I , Shafi Gowhar
Published: June 2021

Aegle marmelos (Beal in Hindi) is commonly found in South Asia. The leaves and other part of plant have been used in traditional medicinal system like Ayurveda and Unani for treatment of diabetes and cardiovascular diseases. Present study was sought to find the facts for anticancer activity of methanol extract (ME) of Aegle marmelos leaves. GC‐MS analysis of ME revealed 1,2,3‐ Propanetriol was major component followed by Cinnamic acid methyl ester, 3,4‐dihydro‐4,5‐dihydroxy‐ 1(2H)‐Naphthalenone, Phytol, Nicotinamide, etc. After 24 hours treatment with ME, MTT assay was performed and found that 55.67 % of the cells were dead at the highest concentration tested (640μg/mL) whereas, the IC50 of ME was achieved at 512.38μg/mL. Up to the concentration 5 μg/mL the extract showed a growth supporting activity. As the cell viability was 7.55% more than that of untreated control. The induction of apoptosis was observed when HepG2 cells were treated with ME. Total 1000 counts were made and 231±14 (21.3%) cells were found to be apoptotic at 320μg/mL (IC40) ME. DNA bands and a streak in the ME terated lane suggest that there was DNA fragmentation in HepG2 carcinoma hepatocytes. By this preliminary study we conclude that leaf of Aegle marmelos may have an strong candidature to be the source of anticancer drugs. However this extract must be tested against other cancer cell lines before any firm conclusion.
Vasorelaxant activity of p‐cymene in superior mesenteric artery of rats
Authors: Barbosa Renata Lisboa , Santos Julyane Hora , Cunha Patricia Santos , Quintans‐Júnior Lucindo José , Bonjardim Leonardo Rigoldi , de Souza Araújo Adriano Antunes
Published: June 2021

p‐Cymene is an alkylbenzene related to a monoterpene very promising for their biological properties, including actions on the cardiovascular system. Given this, the aim of the present study was to evaluate the vasorelaxant effect induced by p‐cymene in rat mesenteric artery. Artery rings with functional endothelium pre‐contracted with phenylephrine (PHE) (control), p‐cymene (10–8 – 3 × 10–2M) induced relaxation (Emax = 104 ± 6 %; n= 6) was not different from that obtained after removal of endothelium (Emax = 95 ± 7 %; n= 6) or pre‐contracted with KCI 80 mM (Emax = 72 ± 4 %; n= 6), suggesting a possible involvement of Ca2 + channels. In preparations without functional endothelium and pre‐contracted with PHE, incubation with tetraethylammonium, a non‐selective blocker of potassium channels, was able to reduce relaxations induced by p‐cymene in concentrations 10–3, 3.10–3, and 10–2M (p < 0.05 and p < 0.01). Furthermore, the incubation with p‐cymene (3 × 10–3, 10–2, and 3 × 10–2 M) was able to antagonize the contractions induced by CaCl2 (from 10–7 to 3 × 10–2 M) and sodium orthovanadate (from 10–5 to 3 × 10–2 M), an general inhibitor of protein‐tyrosine phosphatase. Taken together, these results suggest that p‐cymene produces a vasorelaxant effect by an endothelium‐independent mechanism in the rat mesenteric artery, possibly involving voltage sensitive Ca2+, K+ channels and Ca2+ desensitization of contractile elements.

Support: CNPq, Capes, Fapitec
Pericytes and Astrocytes Crosstalk: Understanding Perivascular Synergism at the BBB
Authors: Prasad Shikha , Cucullo Luca
Published: June 2021

The Blood brain barrier (BBB) plays a critical role in maintaining brain homeostasis. Astrocytes (AC) lying adjacent to BBB endothelial cells (EC) have been reported to play a major role in ECs polarization and differentiation into a BBB phenotype. Pericytes (PC) have been reported to synthesize most elements of the basement membrane and provide structural support and vasodynamic capabilities to the brain microvasculature thus suggesting a more primary role in BBB differentiation than previously anticipated. Whether PC and AC modulate each other functions and activities (thus affecting the BBB) are still poorly understood. For this purpose we analyzed the conditioned media from human AC, PC and AC‐PC cultures and assessed the morphological changes in correspondingly treated human BBB EC. Analysis of culture media have shown that Ang‐1 (important for BBB TJ formation) was primarily released by PC rather than AC (release of Ang‐2) thus suggesting that PC play an important role in BBB maturation. Furthermore, PC induced AC redistribution (cytosol to membrane) of glut‐1, important for glucose transport across the BBB was seen. This suggests that PC can modulate AC functions thus affecting the BBB. In addition, EC culture in presence of PC and/or AC‐PC conditioned media (but not AC alone) reassembled in a capillary like structure formation. This suggests that PC also play a role during angiogenesis of brain microcapillary.
Drug susceptibility influences macronutrient intake and body composition in tuberculosis patients (1014.4)
Authors: Frediani Jennifer , Tukvadze Nestan , Sanikidze Eka , Kipiani Maia , Hebbar Gautam , Tangpricha Vin , Blumberg Henry , Ziegler Thomas
Published: June 2021

Background: Malnutrition and lean tissue wasting are common in patients with tuberculosis (TB) disease, yet little information is available on habitual dietary intake or links with body composition.

Methods: Dietary intake was obtained in a randomized clinical trial of high‐dose vitamin D (Vit D) in patients with pulmonary TB in Tbilisi, Georgia. Food intake was obtained (0, 8 and 16 weeks) using a validated, culture‐specific tool. Serial body composition was by BMI and bioelectrical impedance analysis (BIA; % fat mass and % fat‐free mass). Descriptive statistics and repeated measures ANOVA were used.

Results: 192 subjects (mean age 35 y; 100 Vit D, 99 placebo) were studied. Mean daily intake of kcal, fat and protein were adequate at diagnosis and increased (with BMI) over time (time effect p<0.0001, treatment effect NS). There were no differences in % of fat mass or % fat‐free mass between groups over time. Multi‐drug resistant TB patients (n=23) had lower body weight, BMI (p=0.05) and fat‐free mass vs. drug‐susceptible patients, despite increased intake of kcal, protein, fat and CHO at 8 and 16 weeks.

Conclusions: Macronutrient intake was adequate in this TB patient cohort. Increased macronutrient intake occurred concomitant with increased BMI and maintained proportions of body fat and fat‐free mass. MDR‐TB patients demonstrated less efficient anabolism than drug‐susceptible TB patients. 1

Grant Funding Source: Supported by grants from the National Institutes of Health Fogarty grant D43 TW007124 and K24 RR0233