bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Role of Cystathionine Gamma Lyase and Endothelial Nitric Oxide Synthase for Cardiovascular Homeostasis
Authors: Al Shaarani Majd , Green Jonette , Traylor James , Bhuiyan Md Shenuarin , Orr A Wayne , Kevil Christopher G 
Published: November 2023

Nitric oxide (NO) and hydrogen sulfide (H2S) are two important vasoactive gasotransmitters. Both NO and H2S are essential in the genesis, development, repair and remodeling of blood vessels. They also have a cardio‐protective role that stems from their ability to regulate vasodilation, blood pressure and and the growth of new blood vessels. Genetic deficiency of either endothelial NO synthase (eNOS) or Cystathionine γ‐lyase (CSE, one of the genes implicated in H2S synthesis) has detrimental effects on tissue vascularization and vascular repair against shear forces and atherosclerotic changes. While the deletion of either gene alone has been previously studied; the effects of genetic deficiency of both genes together in the same animal has not been previously evaluated. In this study, we report that when both genes are deleted severe cardiovascular pathological changes emerge. Cardiac ejection fraction (EF) and fraction shortening (FS) measured on 2D/M mode echocardiogram performed on homozygous eNOS/CSE double knockout mice was severely diminished compared to that of homozygous CSE null/heterozygous eNOS knockout mice. H&E stained sections of the aorta showed significant changes with irregular, eosinophilic and less defined elastic laminae with loss of refractility and edematous tunica media with pauci‐cellularity and hypochromatic nuclei. These data are the first to reveal a critical relationship between eNOS and CSE expression for basal cardiac function and tissue histological organization.Support or Funding InformationThis work supported by HL113303
ENaC‐a mediates the protective effect of the TNF‐derived TIP peptide in pneumolysin‐induced capillary barrier dysfunction
Authors: Czikora Istvan , Sridhar Supriya , Alli Abdel , Verin Alexander , Chakraborty Trinad , Fulton David , Eaton Douglas C , Lucas Rudolf 
Published: November 2023

BackgroundStreptococcus pneumoniae is the major etiologic agent of bacterial pneumonia. Autolysis and antibiotic‐mediated lysis of pneumococci releases the pore‐forming toxin, pneumolysin (PLY), which is the primary cause of acute lung injury. PLY unleashes the perfect storm, by inhibiting alveolar liquid clearance (ALC) and by potently compromising alveolar‐capillary barrier function, which together cause potentially lethal permeability edema associated with pneumonia. As a consequence, alveolar flooding occurs, which can precipitate lethal hypoxemia by impairing gas exchange. Apart from being the entry step in Na+ reabsorption across Na+‐transporting epithelia, the epithelial sodium channel (ENaC) was recently suggested to affect stiffness in the large vasculature, but its role in barrier function in the microvasculature has not been investigated so far. In this study, we therefore investigated the consequences of specific ENaC activation or depletion on capillary barrier function, in the presence or absence of PLY.MethodsExpression of α, β and γ subunits of ENaC and protein phosphorylation status in HL‐MVEC were assessed in Western blotting. Barrier function in untreated and PLY‐treated human lung microvascular endothelial cell (HL‐MVEC) monolayers was monitored using electrical cell‐substrate impedance sensing (ECIS). In order to investigate the role of ENaC in barrier function in HL‐MVEC, we either activated the channel, using the TNF‐derived TIP peptide, which directly binds to the α subunit, or we depleted the crucial ENaC‐α subunit with specific siRNA.ResultsHL‐MVEC express all three subunits of ENaC. TIP peptide‐mediated ENaC activation significantly strengthened barrier function in PLY‐treated HL‐MVEC. By contrast, ENaC‐α depletion significantly increased sensitivity to PLY‐induced hyperpermeability and blunted the protective effect of the TIP peptide. PLY significantly increased phosphorylation of the actin‐binding protein filamin A, which was shown to directly bind to ENaC‐α and which promotes the interaction between ENaC with MARCKS, an important mechanism for activation of the channel's open probability. TIP peptide‐mediated activation of ENaC blunted phosphorylation of both calmodulin dependent Kinase II (CaMKII) and its substrate filamin A.ConclusionsTaken together, our results demonstrate a previously unrecognized role for ENaC, and more specifically for ENaC‐α, in strengthening barrier function of human lung capillaries. As such, these data suggest that strategies aiming to activate ENaC have the potential to not only improve ALC in the alveolar epithelium, but also barrier function in the capillary endothelium during pneumonia.
Epigenetic Regulation of Stab2 Expression in DBA Mice in Determining Atherosclerosis Susceptibility
Authors: Dong Sharlene , Kayashima Yukako , Maeda Nobuyo 
Published: November 2023

Atherosclerosis is a progressive disease characterized by the accumulation of lipids and fibrous elements in the large arteries that often leads to acute occlusion and myocardial infarction. The aortic arch and root are two regions with distinct patterns of lesion development in a mouse model of atherosclerosis. A quantitative trait locus (QTL) analysis of the F2 generation from an intercross between ApoE‐null mice on 129S6 and DBA/2J backgrounds has revealed a significant locus on chromosome 10, Aath5, that affects plaque size in the aortic arch. The DBA allele of Aath5 confers resistance to aortic arch atherosclerosis. One candidate gene in the Aath5 locus is the Stab2 gene, which encodes a receptor for hyaluronan. The DBA allele of Stab2 was found to be associated with ten‐fold levels of plasma hyaluronan in DBA mice compared to 129 mice. The DBA allele of the Stab2 gene also includes a unique intracisternal A‐particle (IAP) retrotransposon inserted in a reverse orientation in the promoter region, driving the Stab2 expression from its long‐terminal repeat sequence (5′ LTR). Methylation status of the LTR of the IAP sequence was analyzed through allele‐specific bisulfite sequencing and found to be correlated with ectopic expression in the heart, aorta, macrophages and kidneys. Thus, epigenetic regulation of this IAP element may play a role in regulating Stab2 expression and possibly affect atherosclerotic susceptibility in the aortic arch in certain strains of mice.Support or Funding InformationThe work is supported by a National Institutes of Health (NIH) grant HL042630 to NM.
Development of compounds with mu‐opioid agonism and delta‐opioid antagonism as novel analgesics
Authors: Fernández Thomas Joaquín , Griggs Nicholas W , Traynor John R 
Published: November 2023

Opioid analgesics exert both their beneficial and unwanted effects by activation of the mu‐opioid receptor (MOPR). Furthermore, repeated administration of opioid analgesics and subsequent chronic MOPR activation leads to opioid tolerance and physical dependence. Many attempts to eliminate the adverse effects of opioids by the synthesis of more potent and selective MOPR agonists have proved to be unsuccessful. It has been proposed that activation of MOPR with concurrent antagonism of the delta‐opioid receptor (DOPR) can reduce the development of opioid tolerance and reward. Here we report on the development of “mixed efficacy compounds” that activate MOPR and inhibit DOPR. We evaluated potential mixed efficacy compounds using two in vitro assays; 1) [35S]‐GTPγS binding assays to measure G protein activation in membrane preparations of cloned cell lines expressing either MOPR, DOPR, or kappa‐opioid receptors (KOPR). Full agonists at each opioid receptor (DAMGO, DPDPE, and U69,593, respectively) were used as standards for [35S]‐GTPγS binding. Maximum binding and potency (EC50) values of the novel compounds were determined from concentration‐effect curves. 2) Competition binding assays using [3H]‐diprenorphine (opioid antagonist) to determine affinity (Ki) values. Comparison of affinity and degree of agonism of the novel compounds at MOPR and DOPR revealed structure‐activity relationships to guide further chemical modification and drug development to improve compounds devoid of tolerance and dependence liabilities.Support or Funding InformationSupported by DA‐03910
Inhibition of Opioid Induced Reward via Genetic and Pharmacological Manipulations
Authors: Davidson‐Knapp Rachel , Sandweiss Alesander J. , McIntosh Mary I. , Moutal Aubin , Hu Jackie , Giri Aswini , Yamamoto Takashi , Hruby Victor J. , Khanna Rajesh , Largent‐Milnes Tally M. , Vanderah Todd W. 
Published: November 2023

Chronic pain affects approximately 100 million Americans for which opioids are the mainstay therapy. While physicians and patients alike are apprehensive about using opioids due to their side effects including respiratory depression and addiction, 259 million opioid prescriptions were written in 2012. Although opioids are the best available analgesics, they increase both positive and negative reinforcement, ultimately leading to addiction. The pro‐nociceptive or pain promoting, neurotransmitter Substance P (SP) and its corresponding receptor (NK1R) are found associated with dopamine neurons in the ventral tegmental area (VTA). Evidence exists suggesting that SP potentiates positive reinforcement of opiates leading to reward. Using in vivo microdialysis, we show that systemic morphine significantly increases SP release in the VTA, an effect mediated by inhibition of VTA GABAergic neurons. SP administered in the VTA significantly increases levels of dopamine in the nucleus accumbens (NAc), a reward processing center in the ventral striatum. Using CRISPR‐Cas9 knockdown of NK1R in the VTA, we show this prevents the activation of opiate reward, corroborating the results from a conditioned place preference paradigm (CPP). Finally, we developed a novel opioid agonist/NK1R antagonist, TY032, which was shown to inhibit acute and chronic pain in male rats. Importantly, our study found TY032 microinjection into the VTA did not result in an increase in extracellular dopamine release in the NAc. These data support targeting the dopamine reward circuitry and pain pathways with multifunctional opioid‐NK1R compounds could be a path to developing analgesics that lack the potential for abuse.Support or Funding InformationFunding NIH/NIDA DA018717, NIH/NIDA DA013449.
CNS‐Permeant Tamoxifen Analog Modulates The Dopamine Transporter And Reduces Amphetamine Reinforcing Effects
Authors: Carpenter Colleen , Zestos Alexander , Altshuler Rachel , Sorenson Roderick , Guptaroy Bipasha , Jutkiewicz Emily , Kennedy Robert , Showalter Hollis , Gnegy Margaret 
Published: November 2023

Amphetamine (AMPH) is the second most widely abused drug worldwide, yet an effective therapeutic for AMPH abuse and addiction remains elusive. The reinforcing properties of amphetamine are due to its ability to increase extracellular dopamine in the nucleus accumbens. This process is facilitated by protein kinase C (PKC) activation and blocking PKC reduces AMPH activities.The selective estrogen receptor modulator tamoxifen remains the only available brain‐permeant PKC inhibitor. We have created a small library of novel tamoxifen analogs with increased selectivity for PKC inhibition and reduced affinity at the estrogen receptor. With an IC50 of 80 nM, our lead compound (6c) inhibits PKC 250 times more potently than tamoxifen and displays no binding at the estrogen receptor α. In determining the efficacy of 6c to reduce the neurochemical and behavioral effects of AMPH, we firstly investigated the ability of the compound to modulate the dopamine transporter in both cellular and rat striatal synaptosomal preparations.6c showed negligible effects in blocking dopamine uptake or AMPH‐induced dopamine release in HEK cells stably expressing the dopamine transporter. However, 0.3–3 μM 6c dose dependently inhibits dopamine release stimulated by 10 μM AMPH in synaptosomes. 6c less potently blocks dopamine uptake and does not displace [3H]WIN 35428 binding at the dopamine transporter in synaptosomes, showing that it does not bind near the active site. 6 mg/kg 6c s.c. does not affect baseline dopamine levels or locomotion but significantly reduces both dopamine release and locomotion induced by 2 mg/kg i.p. AMPH as measured with microdialysis in freely‐moving rats. 6 mg/kg s.c. 6c reduces i.v. AMPH self‐administration (0.032 mg/kg AMPH per infusion) by ~40% but does not affect food self‐administration in rats.6c effectively reduces the actions of AMPH both in vitro and in vivo. The fact that 6c modulates the dopamine transporter in synaptosomes but not in cells overexpressing DAT, suggests its action is via a mediator versus a direct action at the transporter. This study also highlights the importance of choosing appropriate models of the dopaminergic terminal in screening compounds for AMPH abuse.Support or Funding InformationNIH grant 1R01 DA11697, HHMI International Student Research Fellowship
Regulation of Goal‐directed Action Selection by Cocaine, MDMA, and Orbitofrontal BDNF‐trkB
Authors: Pitts Elizabeth , Gourley Shannon 
Published: November 2023

Cocaine dependence is characterized by compulsive drug use and maladaptive decision‐making. Adolescents are particularly vulnerable to the effects of cocaine; for example, cocaine exposure during adolescence increases the risk of developing lifelong addictions. Here we show that subchronic cocaine exposure during adolescence results in increased responding during acquisition of cocaine self‐administration and a bias towards stimulus‐driven habits in adulthood in mice. Changes in Brain‐derived Neurotrophic Factor (BDNF) in the orbitofrontal prefrontal cortex (oPFC) could underlie, in part, this habit bias, while strategies that stimulate cortical BDNF systems could be protective. We utilized viral‐mediated gene transfer to interfere with the activity of BDNF's high‐affinity receptor tyrosine kinase receptor B (trkB) selectively in the oPFC. This manipulation induced habit‐like behavior in an instrumental contingency degradation task. Next, we hypothesized that stimulating BDNF expression in the oPFC could block habits by enhancing response‐outcome learning and memory. We report that 3,4‐methylenedioxymethamphetamine (MDMA) increases BDNF levels in the oPFC, but not the amygdala or dorsal striatum, and also “breaks” habits resulting from adolescent cocaine exposure. Interfering with trkB activity in the oPFC blocks MDMA‐mediated rescue of goal‐directed decision making. MDMA was delivered immediately following instrumental contingency degradation, suggesting that it enhances the consolidation of new response‐outcome learning and memory. Finally, 7,8‐dihydroxyflavone (7,8‐DHF), a trkB agonist, also reverses cocaine‐induced habits, further suggesting that BDNF‐trkB systems are a point of intervention in combatting maladaptive decision making following repeated cocaine exposure.Support or Funding InformationThis work was supported by DA036737 and DA042358. The Yerkes National Primate Research Center is supported by the Office of Research Infrastructure Programs: OD P51OD011132
Dimethyl Fumarate Improves Depression and Anxiety by Reducing Brain Catalase Levels in Mice
Authors: Iniaghe Loretta Oghenekome , Ilondu Chinenye Amara , Eseka Ewere Ogechuwu , Gabriel Benjamin 
Published: November 2023

IntroductionDepression and anxiety are psychiatric disorders which are leading causes of disability and are estimated to be major contributors to overall global burden of disease; both often accompany chronic diseases. This study investigated the effects of dimethyl fumarate (DMF) in animal models of depression and anxiety.MethodsMice (n=40) were treated with either the vehicle ‐dimethyl sulfoxide/distilled water (DMSO solution), 50 and 100 mg/kg DMF and Imipramine and subjected to either the forced swim test (FST), n=20 or the tail suspension test (TST) after they were euthanized, whole brains isolated, homogenized and assayed for brain catalase levels. Another set of mice (n=40) were treated daily with either the vehicle, DMF 50 and 100 mg/kg and Imipramine for two weeks and subjected to FST and TST on the fourteenth day. Thereafter, they were sacrificed, whole brains isolated, homogenized and assayed for brain catalase levels. Changes in body weight were also recorded daily.The same procedure was followed for determination of anxiety using the stair case and hole board tests instead of the FST and TST and Diazepam as the reference drug.ResultsIn the test for depression, 100 mg/kg DMF significantly (p<0.05) reduced periods of immobility in both the FST and TST after acute and chronic drug administration. DMF also significantly (p<0.05) decreased brain catalase levels and improved body weights when compared to the control.In the test for anxiolysis, 50 mg/kg DMF significantly (p<0.05) improved upward climbs and number of head dips in both the stair case and hole board tests respectively.ConclusionDMF exhibited anti‐depressant activity and anxiolytic properties at doses used in this studySupport or Funding InformationPersonal Funds of the Authors
DSS‐induced intestinal damage exacerbates liver injury by acute alcohol exposure in mice: role of intestinal hypoxia‐inducible factor 1α
Authors: Shao Tuo , McClain Craig , Feng Wenke 
Published: November 2023

Inflammatory bowel diseases (IBD) are characterized by repeated injury of the mucosa and loss of the intestinal epithelial barrier function, which has also been considered one of the major contributors to the development of alcoholic liver disease (ALD). Hypoxia induced factor 1α (HIF1α) plays a critical role in the intestinal barrier integrity by regulating several mucus and tight junction protecting genes. In this study, we hypothesize that intestinal HIF1α deletion increases intestinal permeability resulting in an elevation of endotoxemia in the dextran sulfate sodium (DSS)‐induced colitis murine model, and this exacerbates acute alcohol‐induced liver injury. To test this hypothesis, Wide type (WT) and intestinal epithelial‐specific HIF1α knockout (IEhif‐1α−/−) mice were fed 3% dextran sodium sulfate (DSS) in drinking water for 7 days and a bolus of alcohol at 6g/kg was gavaged 6 hours before sacrificing. DSS decreased body weight of the WT mice, and this decrease was more significant in the IEhif‐1α−/− mice. Intestinal inflammation and mucosal abnormality were increased in the IEhif‐1α−/− mice than that in the WT mice, along with an elevated serum LPS concentration, indicating an increase in intestinal barrier permeability. Binge alcohol, in addition to DSS, further damaged barrier function and increased serum LPS concentrations. Further analyses showed that DSS+alcohol induced a decrease in the expression of caludin‐1 and intestinal trefoil factor, which play an important role in intestinal barrier integrity. Intestine epithelial HIF1α deletion aggravated this effect. Unlike alcohol exposure, DSS treatment alone did not affect liver function in either WT or IEhif‐1α−/− mice, as determined by serum ALT and AST analysis. However, ALT and AST levels were significantly elevated in DSS + alcohol treatment groups, and these elevations were more pronounced in IEhif‐1α−/− mice. HIF‐1α deletion also enhanced alcohol‐induced hepatic triglyceride concentration in DSS‐treated mice. Hepatic histology analysis showed that IEhif‐1α−/− mice had an increased fat accumulation and inflammatory cell infiltration, confirming the hepatic steatosis and liver injury. In conclusion, our results demonstrated that DSS‐induced colitis increases intestinal permeability and endotoxemia, resulting in an worsened liver steatosis and injury due to binge alcohol exposure. Deleting intestinal HIF1α further exacerbates these deleterious effects. Targeting intestinal HIF1α might be an effective strategy for the treatment of colitis‐associated ALD.Support or Funding InformationSupported by NIH and Veterans Administration.
Repairing Interstrand DNA Crosslinks (ICL): Characterization of an ICL Incision/Lesion Bypass Polymerase Complex regulated by the Fanconi anemia pathway
Authors: Jasti Vijay Prasad , Sharma Shilpy , Althaus Irene , Weinstein Abby , Ramani Shreyas , Canman Christine 
Published: November 2023

The ‘REVersionless’ genes, REV1, REV3, and REV7, were discovered as being necessary for both spontaneous and DNA damage‐induced mutagenesis in yeast. REV3 is the catalytic subunit of DNA polymerase Zeta (ζ) and together with its accessory subunit REV7 extends nucleotides inserted opposite multiple types of DNA lesions. REV1 is a deoxycytidyl transferase and performs a scaffolding function in orchestrating the switch between insertion TLS polymerases directly opposite a DNA adduct and Polζ for the extension step. REV1 and Polζ are also involved in ICL repair and homologous recombination (HR) repair of DNA double stranded breaks (DSB). The phenotypes of cells deficient in expression of REV1, REV3, or REV7 are similar to cells derived from patients diagnosed with Fanconi Anemia (FA), a genetic disorder characterized by bone marrow failure, ICL hypersensitivity, and increased risk for cancer. Previous studies have demonstrated an epistatic relationship between REV1 or REV3 and genes encoding proteins comprising the FA core signaling complex in resistance to cisplatin. Additionally, REV1, Polζ, and the FANCD2/FANCI heterodimer have been shown individually to be involved in homologous recombination (HR) repair, which is crucial for resolution of DNA DSBs and ICL repair during S phase. We demonstrate that human REV1 and REV3 exhibit an epistatic relationship with FANCD2 and FANCI in cellular resistance and ICL repair following mitomycin C (MMC) treatment, as well as HR repair. MMC‐induced chromosomal anomalies are increased in REV1 or REV3‐depleted cells and FANCD2 depletion does not further exacerbate this phenotype. We further show that the FANCA core complex protein regulates MMC‐induced chromatin association of REV7 and by extension REV3 to sites of ICL repair. Co‐immunoprecipitation studies show that REV1, REV3 and REV7 interact with FANCD2 and FANCI independent of DNA damage or monoubiquitination of FANCD2 by the FA core complex. In addition, REV1, REV3, and REV7 co‐immunoprecipitate with the SLX4/FANCP scaffold protein, which regulates the unhooking step of an ICL DNA adduct by the ERCC1/XPF (FANCQ) endonuclease and HR repair of a DSB. We propose a model where REV1/Polζ‐dependent activities during ICL and HR repair are regulated by multiple FA proteins including FANCA, FANCD2 and SLX4 (FANCP). Together, these results suggest that REV1 and Polζ are part of a macromolecular complex involved in resistance to many anticancer agents and interactions within this complex may be viable targets for small molecule inhibitors to sensitize resistant tumors to chemotherapy.Support or Funding InformationR01 CA133046