bluuebunny/crossref_metadata_2025_split huggingface dataset:
Structure Activity Relationship of PEG‐Peptide Scavenger Receptor Inhibitors
Authors: Allen Rondine , Rice Kevin 
Published: November 2023

The rapid capture and degradation of viral and non‐viral gene delivery nanoparticles by scavenger receptors (SR) on Kupffer cells and fenestrated endothelial cells in the liver results in a decreased half life of all nanoparticles in an in vivo system, as well as a lower percentage of the dose available for delivery to the target organ. Current attempts to achieve SR inhibition involves co‐administering high molecular weight, polyanionic inhibitors. Polyinosinic acid (Poly‐I), the most widely used and potent inhibitor, competes for SR binding and successfully inhibits the uptake and metabolism of viral gene delivery nanoparticles in the liver. However, Poly‐I also activates the immune system, resulting in toxicity in mice, making Poly‐I clinically unacceptable.We have discovered peptide based SR inhibitors that block SR uptake of DNA nanoparticles and improves their metabolic half‐life by forming protein nanoparticles in the blood. These Poly (ethylene) glycol (PEG) polylysine peptides potently inhibit SR and allow DNA to transfect hepatocytes up to 12 hours after administration in mice.Radio‐iodinated PEG‐peptides were used to study the pharmacokinetics and biodistribution to understand the structural properties which influence transfection competency and potency. We hypothesize that the in vivo potency and activity is influenced by the stability of protein nanoparticles formed in the blood. Peptides which are able to form stable nanoparticles are able to effectively block liver uptake of DNA nanoparticles by saturating SRs on Kupffer cells in the liver. The results of this study provide a framework for the design and synthesis of future PEG‐peptide scavenger receptor inhibitors, as well as an assay to quickly determine the activity of each inhibitor.Support or Funding InformationThe Iowa Biosciences Academy is supported by the National Institute Of General Medical Sciences of the National Institutes of Health under Award Number R25GM058939, the University of Iowa (UI) Office of the Vice President for Research and the UI Chief Diversity Office. This work was also supported by the NIH Pharmacological Sciences Training Grant T32 GM067795 and NIH grant R01 GM087653.
Belinostat, A Clinically‐Used Histone Deacetylase Inhibitor, Inhibits Rifampicin Induced CYP3A4 and MDR1 Gene Expression
Authors: Pondugula Satyanarayana R , Abbott Kodye L , Chaudhury Chloe S , Schwartz Dean D 
Published: November 2023

Pregnane X receptor (PXR), a ligand‐dependent nuclear receptor, is primarily involved in regulating the gene expression of enzymes and proteins involved in endobiotic and xenobiotic metabolism. In particular, PXR plays a crucial role in regulating the gene expression of drug‐metabolizing enzymes and drug‐efflux pumps, including cytochrome p450 3A4 (CYP3A4) and multidrug resistance protein 1 (MDR1). In cancer patients, upregulation of the levels of the PXR target genes, including CYP3A4 and MDR1, can lead to chemoresistance as well as adverse drug interactions. It is possible to inhibit the activity of PXR in order to overcome PXR‐mediated chemoresistance and adverse drug interactions. However, a selective and less‐toxic PXR antagonist has yet to be developed. We sought to determine whether belinostat, a clinically‐used histone deacetylase inhibitor, inhibits the PXR target gene expression in human liver cells (primary hepatocytes & hepatocells) and human intestinal cells (LS174T colon cancer cells). Rifampicin, an agonist of human PXR, was used to activate PXR. Cell viability and CYQUANT cell proliferation assays were performed to determine cytotoxicity and cell proliferation, respectively. Quantitative RT‐PCR assays were conducted to study gene expression of CYP3A4 and MDR1. CYP3A4 P450‐Glo and Rhodamine 123 intracellular accumulation assays were performed to determine the function of CYP3A4 and MDR1, respectively. Belinostat, at its non‐cytotoxic and therapeutically‐relevant concentrations, inhibited rifampicin induced gene expression of CYP3A4 and MDR1 in the liver/intestinal cells. Belinostat also attenuated the PXR agonist‐induced activity of CYP3A4 and MDR1 in the liver/intestinal cells. Furthermore, belinostat decreased rifampicin‐induced proliferation of LS174T human colon cancer cells, suggesting that belinostat suppresses PXR‐mediated cell proliferation of the cancer cells. These results are consistent with the conclusion that belinostat, at its non‐cytotoxic and therapeutically relevant concentrations, downregulates PXR target gene expression by inhibiting the transcriptional activity of PXR. Future studies will be directed to study the selectivity of belinostat to PXR, mechanisms of belinostat inhibition of PXR, and belinostat inhibition of PXR target gene expression in PXR mouse models.Support or Funding InformationThe authors would like to thank Drs. Coleman and Tao for sharing their research facilities. This work was supported by the Auburn University Research Initiative in Cancer Grant, Animal Health and Disease Research Grant, and Auburn University Startup Funds to Pondugula SR.
Saikosaponins Induced Hepatotoxicity Through Disruption of Lipid Metabolism and Induction of Oxidative Stress in Mice: A Proteomic Analysis
Authors: Sun Rong , Li Xiaoyu , Yang Qian , Li Xiaojiaoyang , Liu Runping 
Published: November 2023

INTRODUCTIONRadix Bupleuri (RB) has been popularly used for treating many liver diseases such as chronic hepatic inflammation and viral Hepatitis in China. Increasing clinical and experimental evidences indicate the potential hepatotoxicity of RB or prescriptions containing RB. Recently, Saikosaponins (SS) has been identified as major bioactive compounds isolated from RB, which may be also responsible for RB‐induced liver injury.MATERIALS AND METHODSSerum AST, ALT and LDH were determined to evaluate SS‐induced liver injury in mice. Serum and liver total triglyceride and cholesterol were used to indicate lipid metabolism homeostasis. Liver ROS, GSH, MDA and iNOS were used to examine the oxidative stress level after SS administration. A 8‐Plex iTRAQ Labeling Coupled with 2D LC ‐ MS/MS technique was applied to analyze the protein expression profiles in livers of mice administered with different doses of SS for different time periods. Gene ontology analysis and comparative toxigenomics were employed to elucidate potential mechanism involved.RESULTSSS dose and time dependently induced liver injury in mice, indicated by increased serum AST, ALT and LDH levels. According to proteomic analysis, 487 differentially expressed proteins were identified in mice administrated with different dose of SS for different time periods. Altered proteins were enriched in pathways such as lipid metabolism, protein metabolism, macro molecular transportation, cytoskeleton structure and response to stress. Cluster analysis demonstrated that SS rapidly induced lipid transportation and lipid metabolism related genes in less than 1 hour, followed by stress‐induced proteins, occurred from 4–8 hours and further regulated proteins involved in protein metabolism and energy metabolism (8–24 hours). Lipid metabolism dysfunction and oxidative stress were significantly induced by SS in liver.CONCLUSIONThese results identified hepatotoxicity of SS administration and suggested potential mechanism, including impaired lipid and protein metabolism and oxidative stress. The proteomic approach gave us new insights to understand toxicity of traditional Chinese medicine and provided experimental evidence for the clinical safe usage of RB.Support or Funding InformationThis work was supported by grants from the National Natural Science Foundation of China (Grant No. 81073148, 30672649, 81374059); the National Major Fundamental Research Program of China (Grant No. 2009CB522802); the Project supported by the Shangdong Committee of Science and Technology, China(Grant No. 2008GG2NS02021); the Key Project of Shandong Natural Science Foundation of China (Grant No. ZR2011HZ005). R. Sun is supported by “Traditional Chinese medicine pharmacology and toxicology expert”, the post of Taishan scholar.
Cardiovascular regulation in mice with attenuation or overexpression of the vesicular acetylcholine transporter (VAChT) gene: parasympathetic function in heart failure
Authors: Tezini Geisa C. S. V. , Martins Dias Daniel P. , Oliveira Mauro , Guatimosim Silvia , Prado Marco A. M. , Salgado Helio C. 
Published: November 2023

Studies have shown that a parasympathetic hypoactivity has been identified as an independent risk factor in patients with myocardial infarction (Kleiger et al., 1987; La Rovere et al., 1998). Therefore, this study aims to evaluate the effects of the attenuation, or the overexpression, of the vesicular acetylcholine transporter (VAChT) gene on cardiovascular responses during the development of heart failure (HF). The study was conducted in mice knockdown homozygous for the VAChT gene (VAChT KDHOM, n=3) and transgenic subjects with overexpression of the VAChT gene (ChAT–ChR2–EYFP, n=4). Mice were implanted with probes for arterial pressure (AP) and heart rate (HR) recording by telemetry. Ten days after probe implantation the hemodynamics (AP and HR) and cardiac sympathovagal balance were assessed. Next, the mice were submitted to left coronary artery ligation for producing myocardial infarction (MI) and the development of HF. Data were collected 1 and 4 weeks after MI. Morphological analysis revealed similar infarct size among groups (32±4% in VAChT KDHOM vs. 28±2% in ChAT–ChR2–EYFP). VAChT KDHOM mice showed 1 week after MI increased mean AP (MAP) and HR; these parameters were back to normal after 4 weeks. ChAT–ChR2–EYFP mice presented 1 week after MI opposite results, with reduction of MAP and HR. MAP was back to normal after 4 weeks, while the HR remained at lower levels. Regarding the cardiac sympathovagal balance, there were no differences between groups. Overall, the results suggest that even though the extent of the MI was similar in both groups, the hemodynamic profile after this cardiac insult was opposite; VAChT KDHOM mice responded to MI with increased MAP and HR, whereas ChAT–ChR2–EYFP mice with reduction of both hemodynamic variables.Support or Funding InformationFAPESP
Methotrexate Scavenges Xanthine Oxidase‐Derived Superoxide without Inhibiting Xanthine Oxidase Activity
Authors: Sarmiento Cleafes , Chiou Anderw , Duryee Michael J. , Tian Jun , Thiele Geoffrrey M. , Anderson Daniel R. , Mikuls Ted R. , Zimmerman Matthew C. 
Published: November 2023

Background/HypothesisRheumatoid arthritis (RA), an autoimmune disorder affecting 1.5 million Americans, is associated with chronic inflammation and elevated levels of reactive oxygen species (ROS) in joints resulting in painful swelling, bone erosion, and joint deformity. Methotrexate (MTX), a disease‐modifying anti‐rheumatic drug, has anti‐inflammatory, anti‐proliferative, and anti‐fibrogenic properties; however, the precise mechanism(s) by which MTX provides relief to RA patients is unclear. Although previous studies have suggested that MTX inhibits ROS production, these studies failed to determine if MTX truly inhibits production or increases scavenging of ROS. Herein, we tested the hypothesis that MTX directly scavenges a specific ROS, that is, superoxide.MethodsIn a cell‐free in vitro system, superoxide generated by hypoxanthine (HX, 50mM) and xanthine oxidase (XO, 10mU/mL) was measured using electron paramagnetic resonance (EPR) spectroscopy and the EPR spin probe 1‐hydroxy‐3‐methoxycarbonyl‐2,2,5,5‐tetramethylpyrrolidine (CMH, 200mM) in the absence or presence of MTX (2mM). In addition, we examined the ability of MTX to inhibit XO activity, by measuring uric acid levels produced by the HX + XO reaction ± MTX.ResultsThe EPR spectra amplitude obtained from samples containing HX + XO were significantly elevated compared to control samples without HX and XO (3.69×106 ± 0.34×106 vs. 5.14×104 ± 0.69×104 EPR arbitrary units, p<0.05); thus, indicating increased levels of superoxide. MTX significantly attenuated levels of superoxide as determined by decreased EPR spectra amplitude (1.67×106 ± 0.24×106 EPR arbitrary units, p<0.05 vs. HX and XO alone). MTX did not inhibit XO activity as uric acid levels increased 8.77% ± 1.20% without MTX and 8.44% ± 1.00% with MTX.ConclusionsMethotrexate, a first line of defense for RA patients, directly scavenges XO‐derived superoxide, but does not inhibit XO activity. Future studies are needed to determine if MTX's anti‐inflammatory, anti‐proliferative, and/or anti‐fibrogenic actions are due to its ability to directly scavenge superoxide.Support or Funding InformationThis research was made possible by grants from the National Center for Research Resources (5P20RR016469) and the National Institute for General Medical Science (NIGMS) (8P20GM103427), a component of the National Institutes of Health (NIH) and its contents are the sole responsibility of the authors and do not necessarily represent the official views of NIGMS or NIH.
Lateral Hypothalamic Regulation of Energy Homeostasis and Sympathetic Outflow: The Role of Leptin and Melanocortin Signaling
Authors: Cruz Eva Nilda Rodriguez , Morgan Donald A , Davis Kevin Carroll , Saito Kenji , Toth Brandon A , Martinez Eduardo , Jiang Jingwei , Rahmouni Kamal , Cui Huxing 
Published: November 2023

Hypothalamus plays a fundamental role in both body weight homeostasis and sympathetic regulation, but the underlying neural basis of this association remains incompletely understood. Brain leptin and melanocortin systems have emerged as critical regulators of body weight homeostasis, sympathetic nerve activity (SNA) and blood pressure. However, the neural circuits by which leptin and melanocortin regulate SNA and blood pressure have not been fully defined. We have previously identified a unique subpopulation of GABAergic neurons in the lateral hypothalamic area (LHA), a historically recognized “feeding center” in the brain, that coexpress both leptin receptor (LepR) and melanocortin 4 receptor (MC4R), indicating that this unique subset of LHA inhibitory neurons are direct targets of endogenous leptin and melanocortin. Because of a well‐established role of LHA in both feeding and sympathetic regulation, we hypothesized that LHA MC4R signaling pathway may plays a key role in sympathetic overactivity and elevated blood pressure associated with obesity. Here we show that in vivo Cre/loxP system‐mediated re‐expression of endogenous MC4Rs in the LHA of severely obese MC4R‐null mice restores the blunted responses of MC4R agonist MTII‐mediated increase in sympathetic outflow to various beds, including brown adipose tissue, muscle, and kidney, and elevates blood pressure in obese normotensive MC4R‐TB mice without significantly affecting food intake and body weight. Finally, optogenetic and/or chemogenetic stimulation of LHA LepR‐positive neurons (some of which are also MC4R‐positive) suppresses food intake and deceases blood pressure and renal SNA in conscious mice. In conclusion, our findings identify a novel brain circuit by which leptin and melanocortin signaling regulate sympathetic traffics and blood pressure, which may contribute to the development of obesity‐associated hypertension.Support or Funding InformationNIH/NHLBIR01 HL127673‐01A1
Human Antigen R (HuR) Regulates Structure and Function of Brown Adipose Tissue
Authors: Yamamoto Kaila , Lanzillotta Lindsey , Anthony Sarah R , Jones Shannon , Yoshida George , Owens Albert Phillip , Tranter Michael 
Published: November 2023

Human antigen R (HuR) is an RNA binding protein widely expressed throughout the body, including in both white (WAT) and brown (BAT) adipose tissue. Recent work from our lab has shown that adipocyte‐specific HuR knockout (adipo‐HuR−/−) mice gain less weight following a high fat diet (HFD) than the wild type (WT) controls. In this study, we focused on the role of HuR deletion on the function of BAT, which mediates non‐shivering thermogenesis.Our results show that, when subjected to a cold stress (4°C), HFD‐fed adipo‐HuR−/− mice are less cold tolerant compared to their WT counterparts. However, when the cold challenge was repeated with chow fed mice, there was no difference between the HuR knockout and wildtype mice in their ability to thermoregulate, suggesting that a change induced by high fat feeding rendered the adipo‐HuR−/− mice less cold tolerant. Histological analysis revealed that, while the BAT of the chow fed adipo‐HuR−/− mice appeared normal, following HFD, the BAT from adipo‐HuR−/− mice appears less dense with a cell size that more closely resembles WAT. Since uncoupling protein‐1 (UCP1) plays a key role in BAT mitochondrial uncoupling and non‐shivering thermogenic regulation, we assessed the ability of HuR to directly mediate UCP1 mRNA expression. Interestingly, analysis of the UCP1 mRNA sequence predicts at least four distinct HuR binding sites in the UCP1 3′UTR, and our data shows that UCP1 mRNA expression is decreased in BAT from adipo‐HuR−/− mice.In conclusion, our results indicate that HuR regulates both the structure and function of BAT, at least in part through direct modulation of UCP1 mRNA expression.Support or Funding InformationThis work was supported by a University of Cincinnati Heart, Lung, and Vascular Institute Near Horizons Grant (MT, APO).
Effects of Lactobacillus rhamnosus on uptake and lipid accumulation in Caco‐2 cells
Authors: Ipinmoroti Ayodeji O , Onyango E M 
Published: November 2023

Probiotic microorganisms have been shown to influence the digestion and absorption of various nutrients including lipids. Accumulation of triglyceride (TAG) in enterocytes of the small intestine follows two specific absorption pathways: the monoacylglycerol acyltransferase pathway (MGAT) and an alternative glycerol acyltransferase pathway (GPAT). Triglycerides are the main constituents of body fat in humans and other animals derived from the esterification of glycerol and free fatty acids. In this study, we investigated the effects of Lactobacillus rhamnosus MRS6AN, an isolate from “amabere amaruranu” a Kenyan traditional cultured milk, on triglyceride accumulation and expression of GPAT3 in Caco2 cell enterocyte model. In the present study, we examined the effects of L. rhamnosus MRS6AN on lipid absorption mediated by glycerol‐3‐phosphate acyltransferase. Caco‐2 cells were treated overnight with various bacterial cell extracts (cytoplasmic fraction, CF; filtered spent broth, FSB; heat inactivated bacteria, HIB) or with live bacteria (LB). Treated Caco‐2 cells were further divided into 3 treatment groups: Treatment 1 cells were directly overlaid with a lipid mixture without washing, Treatment 2 cells were washed in PBS then overlaid with the lipid mixture, while treatment 3 cells were washed with PBS, overlaid with lipid mixture and fresh treatments. The cells were subjected to incorporated lipid content assay through triglyceride accumulation analysis and Oil‐Red staining to determine the lipid content. Lipid content was measured using absorbance (OD 540 nm). Compared to the controls, TAG accumulation in cells treated with LB or its FSB was reduced by 0.143 and 0.097, respectively, in Treatment 1 cells; 0.151 and 0.136, respectively, in Treatment 2 cells; and 0.147 and 0.087 in Treatment 3 cells. HIB had no effect on lipid accumulation in Treatment 1 and 3 cells, but a slight reduction in Treatment 2 cells by 0.065. CF reduced TAG in Treatment 2 and 3 cells by 0.08 and 0.093 respectively. Data from this study suggest that L. rhamnosus MRS6AN strain may reduce lipid uptake and subsequent TAG accumulation in enterocytes.Support or Funding InformationEast Tennessee State University
Diaphragm neuromuscular transmission dysfunction in chronic heart failure
Authors: Kelley Rachel Catherine , Coblentz Philip David , Ferreira Leonardo Franklin 
Published: November 2023

Chronic heart failure (CHF) causes loss of inspiratory muscle pressure (~force) and endurance (inspiratory dysfunction), which contributes to dyspnea and exercise intolerance. Inspiratory dysfunction in patients may arise from phrenic nerve and diaphragm muscle abnormalities. Diaphragm contractile function is preserved in animals with mild CHF, yet patients with mild CHF have inspiratory dysfunction. Therefore, we hypothesized that CHF causes neuromuscular abnormalities that impair diaphragm function.MethodsMale Sprague‐Dawley (SD) rats underwent ligation of the coronary artery to induce CHF or sham surgery. Approximately twelve weeks post‐surgery, we dissected costal diaphragm bundles for tests of maximal force and fatigue characteristics using field (direct) muscle or phrenic nerve stimulation. After stimulations at maximal frequency (200 Hz), we conducted an isometric fatigue protocol (30 Hz, 500 ms trains, 0.5 train/second). In bundles stimulated through the phrenic nerve, we superimposed direct muscle stimulation every 15 s during the fatigue protocol.ResultsAnimals in the CHF groups demonstrated mild‐to‐moderate disease as evidenced by transmural infarct area <35% of left ventricle plus septum and normal right ventricle morphology. Phrenic nerve stimulation caused greater fatigue in diaphragm from animals with CHF (fatigue index at 90th contraction: 70 ± 1.4 % and 55 ± 3.1 %, n=3, p<0.05). CHF also exacerbated phrenic‐to‐diaphragm neuromuscular transmission failure compared to Sham (42 ± 1.1 % and 27 ± 1.4 %, p<0.05). Diaphragm fatigue characteristics were similar in Sham and CHF when contraction was stimulated via direct muscle stimulation. Maximal tetanic specific force (N/cm2) was also similar in Sham and CHF animals with either direct (Sham 29.4 ± 2.4, CHF 25.9 ± 2.6) or phrenic stimulation (Sham 23.5 ± 0.7, CHF 21.9 ± 1.4).ConclusionsOur data suggest that mild/moderate CHF causes neuromuscular transmission failure in the diaphragm. Neuromuscular transmission failure occurred in the absence of diaphragm weakness, which is evident in severe CHF, and likely contributes to inspiratory dysfunction seen in patients.Support or Funding InformationFunding support from NIH R01 HL130318