bluuebunny/crossref_metadata_2025 huggingface dataset:
Does Perfluorooctane Sulfonic Acid (PFOS) Affect the Mouse Immune System?
Authors: Torres Luisa , August Avery
Published: June 2021

Perfluorinated alkyl acids (PFAAs) have been used in numerous industrial applications as stain and water repellants, floor waxes, fire‐fighting foams, denture cleaners, shampoos, carpet spot cleaners, pharmaceutical products, and pesticides. PFAAs contain simple carbon chains bound to fluorine atoms as well as various substitution groups on the terminal end that dictate their active properties. The strength of the carbon‐fluorine bond makes perfluorinated compounds extremely resistant to environmental degradation, thereby leading to persistence in the environment. Perfluorooctane sulfonic acid (PFOS) is the PFAA most commonly found in both human and wildlife blood samples. PFOS exposure has been linked to hepatotoxicity, neurotoxicity, endocrine disruption, and carcinogenicity, but little is known regarding its impact on the immune system. To assess the effect of PFOS on populations of immune cells, we exposed adult male and female C57BL/6 mice to a low dose of PFOS (5μg/kg/day) via oral gavage every other day for 12 or 30 days. We observed a reduction in the number and percentage of lymphocytes as well as a reduction in the numbers of T cells and natural killer cells in the thymus and fat tissue of the mice after 12 days of exposure. However, these changes were not observed at 30 days. These results suggest either a transient reduction in immune cells, or short‐term redistribution of immune cells to other tissues upon oral exposure to PFOS. Future work will determine the impact of longer‐term PFOS exposure on populations of immune cells, as well as effects on response to viral infections, and antibody responses.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Glypican 3 (GPC3)‐CD81 axis regulates Ezrin mediated Hippo pathway via cross talking with HGF/c‐Met axis in hepatocytes and hepatocellular carcinoma (HCC)
Authors: Xue Yuhua , Mars Wendy , Bowen William , Orr Anne , Stoops John , Michalopoulos George
Published: June 2021

The experiments that integrate different pathways in normal and abnormal circumstances will help us to develop optimum targeted therapies because of multiple pathways involved in the solid tumor carcinogenesis. We have shown that activation of CD81 is associated with phosphorylation of Ezrin, which is overexpressed in many types of solid tumors and has been identified as metastasis related protein for many years. We have also shown that GPC3 binds to the membrane tetraspanin CD81, which is one of the portals of entry of hepatitis C virus (HCV). Previous studies have shown that GPC3 is associated with inhibition of hepatocyte growth and liver regeneration by inhibiting CD81‐mediated p‐Ezrin/Hippo/Yap pathway. It also has been shown that HGF/Met axis which promotes growth in HCC can phosphorylate Ezrin as well by different kinases. Since we have demonstrated the predominant GPC+++/CD81− pattern in HCC, the purpose of this study is to explore the regulation of expression and phosphorylation of Ezrin in HCC. We have found that CD81 expression could be detected in membrane and organelle fraction in human hepatoma cell lines HepG2, Huh7 and Hep3B, but immunoprecipitation experiment suggests that CD81 from latter two human hepatoma cell lines has lost association with GPC3. Human hepatoma cell lines have dramatically up regulated Ezrin expression and Yap activity. We found that treatment of primary rat hepatocytes with CD81 agonist antibody or HGF led to higher levels of phosphorylated Ezrin and lower Hippo activity. Mice overexpressing GPC3 had decreased p‐Ezrin but increased Hippo activity. In mice with post‐natal Met knock out there was less Yap expression at baseline level. Rat hepatoma cell lines JM1 and JM2 had dramatically down regulated CD81 expression and Hippo activity, and up regulated p‐Ezrin. Treatment with 20 umol of Ezrin inhibitor led to significant cell death selectively in JM2 cells in vitro.

Conclusions

Both HGF/Met and CD81 mediate p‐Ezrin activity and enhance nuclear Yap levels. These pathways in normal hepatocytes are inhibited by GPC3 and low HGF/Met activity. In human hepatocytes, GPC3 inhibits the CD81‐mediated p‐Ezrin effect on Hippo pathway and cause a decrease in nuclear Yap. The different distribution pattern of HGF/Met and GPC3/CD81 between normal hepatocytes and HCC cells provides the chance to develop precise anti‐HCC therapies. And Ezrin is a promising target of anti‐HCC therapies including primary lesion and metastasis.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Metabolic Profiling of Serum Reveals Significant Changes in Lipid and Amino Acid Metabolism in A Lipodystrophy Mouse Model
Authors: Zhou Yawen , Cheng Kenneth KY , Xu Aimin , Hoo Ruby LC
Published: June 2021

Objective

Adipose tissue serves as not only an energy reservoir to store excess energy in form of triglyceride but also as an important endocrine organ producing adipokines such as leptin and adiponectin. Lipodystrophy, characterized by abnormal or degeneration of adipose tissue, leads to metabolic diseases including hypertriglyceridemia, insulin resistance, and hepatic steatosis. Lipotoxicity is known as a major driver for the metabolic dysregulation in lipodystrophy, however, which specific lipid species and/or other factors contributes to the disease progression is not well characterized. In this study, we employed untargeted metabolomics to identify small metabolites that are altered in a lipodystrophy mouse model.

Methods

We have recently generated the lipodystrophy mouse model by which adipocyte mouse double minute 2 homolog (Mdm2) is deleted by Cre‐loxP approach. Circulating metabolites were measured and compared in the serum of adipocyte‐specific Mdm2 knockout mice at 3‐week (Before lipodystrophy development) and 12‐week old (After lipodystrophy development) with age‐matched wild‐type mice, using an untargeted metabolomics profiling by LC‐MS. Data were explored using principal component analysis and statistical analysis.

Results

Our untargeted metabolomics profiling had successfully identified and quantified more than 300 circulating metabolites in the serum samples. The results promptly point towards a significant increase in metabolites groups including carnitine and amino acids in the lipodystrophy mouse mice. During the development of lipodystrophy, serum level of phosphatidylcholines species (PC) was significantly reduced whereas increased serum sphingolipid species (including sphingosine and sphinganine) were observed in Mdm2 knockout mice is positively associated with insulin resistance. The high rate of incomplete fatty acid oxidation and accumulation of acylcarnitine metabolites may indicate the damage of skeletal muscle due to an excessive fatty‐acid metabolites supply in the lipodystrophy mice.

Conclusion

Together, these data show that metabolic diseases in lipodystrophy mice is associated with the alteration of metabolites supply to other tissues. Moreover, the utilization of lipodystrophy model in investigating the mechanisms of novel therapeutic targets of metabolic disease via maintaining normal adipose tissue functions was demonstrated.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Intraperitoneal myriocin upregulates transforming growth factor b1 (tgf‐b1) expression in murine destabilization of the medial meniscus model of osteoarthritis—evidence for a role of ceramide species in cartilage homeostasis
Authors: Rose Brandon J , Goodman Michael A , Kooyman David L
Published: June 2021

Ceramides are biologically active sphingolipids known to play a role in numerous physiological and pathological processes. In particular, ceramides are known mediators in insulin resistance, and are found in increased abundance in osteoarthritic synovial fluid. Given the comorbidity between osteoarthritis and insulin resistance, we hypothesized that ceramides mediate cartilage catabolism in murine models of osteoarthritis. Further, that downregulation of ceramide synthesis would prevent structural damage and normalize biomarkers associated with osteoarthritis ‐ tgf‐b1, HtrA‐1, Ddr‐2, and Mmp‐13. To test this hypothesis, we performed surgical destabilization of the medial meniscus on the right knee of 8‐week‐old mice, treating one group with the serine palmitoyltransferase inhibitor myriocin and the other with a vehicle. An untreated sham group served as the control. We found myriocin administration failed to protect the joint from surgically‐induced cartilage degradation, and that no statistically significant difference occurred in the biomarkers measured with the notable exception of tgf‐b1, which was significantly increased in myriocin‐treated mice. Our findings lead us to the novel conclusion that ceramide species play an important role in cartilage homeostasis, acting specifically through regulating expression of tgf‐b1.

Support or Funding Information

Brigham Young University ORCA Grant

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MicroRNA‐17‐3p inhibits excessive post‐hypoxic autophagy and attenuates H9C2 cardiomyocytes reoxygenation injury via PTEN‐Akt‐mTOR signaling
Authors: He Yi , Cai Yin , Yan Dan , Zhang Dengwen , Chen Chen , Irwin Michael G. , Wang Sheng , Xia Zhengyuan
Published: June 2021

Background

The microRNA (miR)‐17~92 is one of the best‐characterized polycistronic miRNA clusters, which encodes six individual miRNAs and contributes to various cellular and biological processes. In particular, miR‐17‐3p (a passenger miRNA of miR‐17) has been shown to indirectly inhibit Phosphatase and tensin homolog (PTEN) and attenuate myocardial ischemia/reperfusion (I/R) injury, but the underneath mechanism is unclear. Inhibition of PTEN can upregulate Akt/mTOR signaling and therefore suppresses the excessive autophagy, which may be a mechanism that underlies myocardial I/R injury. We, thus, hypothesized that miR‐17‐3p may protect against hypoxia/reoxygenation (H/R) induced cell injury by inhibiting post‐hypoxic excessive autophagy in H9C2 cardiomyocytes via PTEN‐Akt‐mTOR signaling pathway.

Methods and results

In rat H9C2 cardiomyocytes, H/R (6 hours hypoxia followed by 6 hours reoxygenation) significantly enhanced the expression of miR‐17‐3p (P <0.05 vs. Control), which was concomitant by increased lactic acid dehydrogenase (LDH) leakage (cell injury marker, P<0.05 vs. Control) and the levels of p62 and the ratio of LC3II/I (autophagy markers, P<0.05 vs. Control), suggesting that miR‐17‐3p may be involved in the pathogenesis of H/R induced autophagic cell death. To explore the potential role of miR‐17‐3p in H/R‐induced cell injury, H9C2 cells were transfected with the miR‐17‐3p agomir or its negative control. The overexpression of miR‐17‐3p can significantly attenuate H/R‐induced cell injury (reduced LDH level, P <0.05 vs. H/R) and inhibit H/R‐induced excessive autophagy (decreased levels of p62 and LC3II/I, P <0.05 vs. H/R). Given the important role of PTEN‐Akt‐mTOR signaling in autophagy, the effects of miR‐17‐3p on the key events in the PTEN‐Akt‐mTOR signaling cascade during H/R were examined. As anticipated, miR‐17‐3p overexpression significantly down‐regulated PTEN expression and up‐regulated the levels of phosphorylated Akt1 (Thr308) and mTOR (P<0.05 vs. H/R), indicating that the miR‐17‐3p may protect against excessive post‐hypoxic autophagic cell death in H9C2 cardiomyocytes via PTEN‐Akt‐mTOR signaling pathway.

Conclusion

Upregulating PTEN‐Akt‐mTOR axis and the subsequent inhibition of excessive autophagy may represent the major mechanism whereby miR‐17‐3p attenuates H/R injury in H9C2 cardiomyocytes.

Support or Funding Information

Studied supported by RGC/GRF grants (17158616M, 17117217 M)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inhibition of autophagy in cardiomyocytes by calmodulin antagonists
Authors: Lopez Vanessa , Keomanivong Faithe , Wauson Eric , Tran Quang‐Kim
Published: June 2021

Macroautophagy (hereafter referred to as autophagy) is a catabolic process that regulates important aspects of myocardial metabolism in which cytoplasmic contents including proteins, other macromolecules, and organelles are sequestered in double membrane vesicles and delivered to lysosomes for destruction. While numerous studies have indicated involvement of signaling via Ca2+ in autophagic flux, the precise mechanisms remain unknown. Calmodulin (CaM) is the ubiquitous transducer of intracellular Ca2+ signals. CaM binds to an estimated network of over 300 cellular proteins, yet is expressed insufficiently for its targets. We have begun to examine the role of CaM in autophagic flux in cardiomyocytes. In H9C2 cardiomyocytes, amino acid starvation for 2 hrs triggers autophagic flux, as assessed by the formation of LC3‐II and the decrease in p62 protein levels. Chloroquine dose‐dependently reduces starvation‐induced autophagic flux. Competitive binding assays using purified CaM and FRET‐based CaM biosensors demonstrate direct CaM binding by chloroquine. Consistently, W‐7 and calmidazolium, two other structurally distinct CaM antagonists, dose‐dependently inhibit starvation‐induced formation of autophagic flux. These data demonstrate a role for CaM in autophagic flux in cardiomyocytes.

Support or Funding Information

AHA 15SDG25090279 Grant (to E Wauson) and NIH HL112184 grant (to Q‐K Tran)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Signaling Pathways Contributing to Central Kappa Opioid Diuresis
Authors: Ebert Kevin M , Kapusta Daniel R
Published: June 2021

Central activation of alpha‐2 receptors and nociceptin opioid peptide receptors (NOP) produce marked diuresis in rats, which is mediated in part by a downstream G‐alpha z protein signaling pathway that inhibits antidiuretic hormone (ADH) secretion. These studies investigated whether central G‐alpha protein signaling pathways also contribute in producing the diuretic and/or antinatriuretic response to central administration of the kappa opioid agonist, U‐50488H. Sprague Dawley rats were implanted with an intracerebroventricular (i.c.v.) cannula and pretreated with vehicle or pertussis toxin (PTX), which inhibits the Gi/o family of proteins except for Gz. After 48‐hrs pretreatment, rats were instrumented with cannula in the femoral artery, vein, and bladder and infused i.v. with isotonic saline (55 μl/min). After recovery, blood pressure (BP), heart rate (HR), and urine output (V) were collected in conscious rats before (control) and for 90‐min after (10‐min collections) i.c.v. U‐50488H (n=6). In vehicle pretreated rats, i.c.v. U‐50488H produced a marked increase in V (max Δ, 101±8 μl/min) and decrease in urinary sodium excretion (UNaV; max Δ, 5±0.5 μeq/min). The diuretic response produced by U‐50488H was significantly (p<0.05) blunted by pretreatment with PTX (max Δ, 57±3 ul/min). In contrast, PTX did not alter the decrease in UNaV. U‐50488H did not alter BP or HR in either group. To explore which G‐alpha proteins may participate in the kappa opioid mediated diuresis, separate rats were pretreated i.c.v. (24‐hrs) with an oligodeoxynucleotide (ODN), which selectively targeted a specific G‐alpha protein. In these studies, pretreatment of groups of rats (n=6/group) with a single ODN for either Gi1, Gi2, Gi3, Go, or Gz, failed to blunt the maximal diuresis as compared to the increase in V produced by i.c.v. U‐50488H in scrambled ODN treated rats. These results demonstrate that central kappa opioids produce a pertussis toxin sensitive diuresis, but not antinatriuresis. This suggests that a combination of Gi/o proteins may be required to produce the aquaretic effects of kappa opioid agonists.

Support or Funding Information

Supported by NIH P30GM106392 to DRK

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Sphingosine‐1‐phosphate induced vasoconstrictor responses decreased in cerebral arteries of endotoxemic rats
Authors: SOYDAN GURAY
Published: June 2021

Introduction

Sepsis is the leading cause of mortality in intensive care units. Vascular dysfunction is the most important cause of death in sepsis. There are some data in the literature for vascular hyporesponsiveness to miscellaneous vasopressors. However most of the data are about resistance mesenteric artery and aorta which is the biggest artery in the body. The responses of cerebral arteries to vasopressors were not investigated in vitro while the brain blood flows were investigated in sepsis.

Sphingosine‐1‐phosphate (S1P) is a bioactive lipid mediator which has several physiological functions in cell growth, inflammation, endothelial barrier integrity and vascular tone. Serum S1P levels decrease in sepsis and this decrement is correlated with the severity of sepsis. S1P is a vasoconstrictor lipid mediator which has more vasoconstrictor responses in small sized vessels and brain arteries. Our aim is to study the vasoconstrictor responses of the brain and mesenteric arteries to S1P in sepsis.

Material and Methods

10 mg/kg lipopolysaccharide (serotype O55:B5) were given intraperitoneally to rats to make an animal model os sepsis. Rats were anesthetized with gaseous carbon dioxide and decapitated 20 hours after injection, Basilar, anterior cerebral, middle cerebral and 3rd branch of mesenteric arteries were mounted in a wire myograph. Arteries were stimulated with 124 nM K+‐PSS after 30 min stabilization. Concentration‐response curves for S1P were evaluated.

Results

High K+ responses did not change in brain and peripheral arteries. S1P‐induced vasoconstrictor responses decreased in anterior and prominently in middle cerebral arteries. S1P‐induced vasoconstrictor responses in basilar and mesenteric arteries did not change.

Conclusions

This is the first study that show the alterations of S1P‐induced vasoconstrictor responses of brain arteries in sepsis. The difference in the alteration of S1P responses of cerebral arteries may explain the alteration and redistribution of cerebral perfussion in sepsis.

Support or Funding Information

This study was funded by Hacettepe University Scientific Research Projects Coordination Unit (TSA‐2017‐12848).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Increased Infarct Volume and Altered Repair Following Ischemic Stroke in Mice Lacking the Brain‐Expressed Orphan G Protein‐Coupled Receptor GPR37
Authors: Owino Sharon , Giddens Michelle , Jiang Micheal , Gu Xiaohuan , Wei Ling , Yu Shan Ping , Hall Randy
Published: June 2021

GPR37 is an orphan G protein‐coupled receptor expressed almost exclusively in the nervous system. Since its initial discovery as a substrate for parkin, the majority of studies on GPR37 have focused on its role in Parkinson's disease. Remarkably little is known about GPR37 in other models of neuronal damage/disease. We've previously shown that GPR37 mediates protection from oxidative stress in cultured astrocytes, which led us to assess the role of GPR37 in vivo in a rodent model of ischemic stroke. Cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) in WT and GPR37−deficient mice. Infarct size, histochemical observations, and protein expression levels were analyzed within the brain 72 hrs. following MCAO. Following occlusion, infarct volume was markedly increased in GPR37 deficient mice compared to WT mice. Interestingly, this coincided with a decrease in the abundance of reactive astrocytes (as assessed by the upregulation of GFAP) within the stroke penumbra region of the GPR37‐deficient mice relative to WT mice. Examination of receptor expression levels within the brain also revealed that GPR37 was massively upregulated within multiple brain regions following ischemia, suggesting a critical role for this receptor in the ischemic response. Furthermore, GPR37‐deficient mice exhibited substantially decreased levels of HIF‐1α and nest in positive cells, indicative of diminished reparative processes following hypoxia. Taken together, these studies demonstrate that GPR37 plays an important role in regulating the generation of reactive astrocytes in vivo and modulating repair following ischemic insults.

Support or Funding Information

This study is supported by grant R01‐NS088413 from the National Institutes of Health.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
L‐arginine Supplementation and Thromboxane Synthase Inhibition Reverse Cerebral Ischemia in Murine Cerebral Malaria by Plasmodium berghei ANKA
Authors: da Silva Moreira Aline , de Freitas Almeida Vanessa Estato , Bianco‐Júnior Cesare , Daniel‐Ribeiro Claudio Tadeu , Tibiriçá Eduardo Vera , de Moura Carvalho Leonardo José
Published: June 2021

Cerebral malaria (CM) is one of the most severe and lethal complications of Plasmodium falciparum infection. The mainstay treatment for CM is intravenous artesunate, nevertheless 15–20% of the patients receiving this drug still die. Vascular dysfunction, with vasoconstriction, leads to decreased cerebral blood flow, ischemia, tissue hypoxia and death in CM. Nitric oxide (NO) and arachidonic acid metabolites are important regulators of physiological cerebral blood flow through their vasodilator or vasoconstrictive properties. The objective of this study was to determine the effects of L‐arginine supplementation and pharmacological inhibition of thromboxane A2 (TXA2) synthesis on the reversal of ischemia in experimental cerebral malaria (ECM). C57BL/6 mice infected with Plasmodium berghei ANKA (PbA) and showing clinical signs of ECM were treated subcutaneously with the NO precursor L‐arginine (50mg/kg) and/or the thromboxane synthase inhibitor ozagrel (100mg/kg). Cerebral blood flow was measured by Laser Speckle Contrast Imaging (LSCI). The adjuvant effect of L‐arginine in combination with artesunate on survival of mice with late‐stage ECM was evaluated. Animals that developed ECM showed a significant decrease (25%) in cerebral blood flow compared to non‐infected control animals. L‐arginine induced a marked increase (22.5%) in cerebral blood flow in mice with ECM one hour after administration. The administration of ozagrel, which prevents the generation of the vasoconstrictor metabolite TXA2, also led to increased (16.5%) cerebral blood flow in animals with ECM, and this was consistent with decreased levels of brain TXA2 in ozagrel‐treated compared to saline‐treated animals. The combination of ozagrel and L‐arginine showed no synergistic effect on cerebral blood flow. Treatment of animals with late‐stage ECM with artesunate plus L‐arginine increased survival compared to artesunate plus saline. Conclusion: L‐arginine and ozagrel increase cerebral blood flow, and L‐arginine increase survival in mice with cerebral malaria and show potential as adjuvant therapy for this malaria complication.

Support or Funding Information

CAPES, FAPERJ, IOC‐Fiocruz

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
In Silico Molecular Docking of Fisetin with the Amyloidogenic Peptides Human Amylin and Aβ42
Authors: Villanueva James Amador , Bamigboje Tobiloba , Hari Saran Krishnan , Boluwaduro Adeyemi , Gautham Sakthi , Ibrahim Adam
Published: June 2021

Amylin is a 37‐residue polypeptide which is co‐secreted with insulin from pancreatic beta cells. In diabetics, this protein has been shown to misfold and form amyloids which cause pancreatic cell death. The exact mechanism by which hIAPP causes cell death remains unknown. However, it has been associated with an aggregation of the protein and an increase in the oxidative stress in the cell. Amylin has similarities with Aβ42, the main protein implicated in the pathogenesis of Alzheimer's disease. Fisetin, a naturally occurring substance which is found in strawberries, apples and cucumbers has been shown to slow down the rate of aggregation of Aβ42. Fisetin has also been proven to have antioxidant properties. Based on the structural and molecular similarities between Aβ42 and human IAPP, we have used molecular alignment and docking experiments to study and show the possibility that fisetin may also slow down the rate of aggregation of human IAPP by binding to single IAPP chains before they form aggregates. Using Autodock Vina, Autodock Tools and Discovery Studio Visualizer, the binding sites of fisetin to both proteins were determined. Binding energies and protein conformations were also determined and analyzed.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
DESIGN, SYNTHESIS AND IN‐VITRO EVALUATION OF ATP‐COMPETITIVE INHIBITORS OF AURORA KINASE‐A USING STRUCTURE‐BASED DRUG DESIGN (SBDD)
Authors: Abdullah Omeima Alhaj , Mehanna Ahmed
Published: June 2021

Aurora‐A kinase inhibitors are small molecule ATP – competitive inhibitors that have been emerged as promising chemotherapeutic agents for cancer. In this study, Structure‐based drug design (SBDD), is mainly used to investigate and to study the molecular interactions between selective ATP‐competitive inhibitor (alisertib) along with the proposed compounds and the existing X‐ray structure of Aurora‐A kinase protein (PDB: 2W1C). Based on SBDD, different proposed compounds (p‐aminobenzoic acid derivatives) were designed from alisertib (reference compound) by replacing the lipophilic moiety (benzazepine) with simpler lipophilic cyclic moieties, such as naphthalene, fluorene, and benzothiazole through amide bond as a linker. This structure‐based design was based on the primary interactions of alisertib with key residues of the hinge and hydrophobic regions of ATP‐binding site (Glu‐211, Ala‐213, Lys‐162, Val‐147, Leu‐139, and Leu‐263). p‐aminobenzoic acid derivatives were synthesized and their inhibitory activity was in‐vitro tested by PhosphoSens ® assay. All the designed p‐aminobenzoic acid compounds bound to the ATP‐active site and exhibited similar interactions as other scaffolds of Aurora‐A inhibitors. However, they presented less inhibitory activity against Aurora‐A compared to alisertib (IC50= 6 nM). Compound‐5 (IC50= 46μM) was the most active compound and will be further modified to enhance the inhibitory activity of Aurora‐A kinase.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A Methodology for Prioritizing Pediatric Pharmacokinetic Studies and Pharmacogenomic Implementation using Evidence from Patient Drug Safety Data
Authors: Eugene Andy Roger , Eugene Beata , DiCenzo Robert , Harralson Arthur
Published: June 2021

Introduction

Physicians attempting to integrate pharmacogenomic testing into routine patient care have been limited due to insurance company reimbursements. However, hospitals committed to improving patient safety outcomes are integrating pharmacogenomic consultations from board‐certified pharmacologists and clinical pharmacists to provide personalized dosing via pharmacogenomic and pharmacometrics tools. Therefore we sought to identify and prioritize drug‐gene pairs associated with the most frequently reported adverse drug reactions (ADRs).

Methods

Drug safety data from the first and second quarter of 2017 were downloaded as ASCII text files from the United States Food and Drug Administration's Adverse Events Reporting System (FAERS). The quarterly FAERS files were merged with the ‘primaryid’ column defining the unique case sequence identifier and manufacturer version numbers. Data homogenization and summary statistics were computed in R for statistical programming (version 3.3.2, Vienna, Austria). The most frequently reported drugs and adverse drug events (ADRs) are reported and stratified using the following pediatric age groups: neonate, infant, child, and adolescent. Drug‐Gene associations were referenced from GeneCards data base (http://www.genecards.org/).

Results

In the first and second quarters of 2017, the most frequently reported drugs (% of total reported cases) [top 5 ranked genes] within the pediatric age groups were: Neonate – Ondansetron (71%) [HTR3A, CYP2D6, ABCB1, TACR1, CCK], Infant – Valproate Sodium (8%) [SCN1A, SCN4A, SLC2A1, SCN5A], Child – Risperidone (43%) [PRL, DRD2, HTR2A, CYP2D6, ABCB1], and Adolescent – Risperidone (28%) [PRL, DRD2, HTR2A, CYP2D6, ABCB1]. The top reported ADRs were: Neonate – Foetal exposure during pregnancy (8%), Infant – off label use (4%), Child – gynaecomastia (12%), and Adolescents – gynaecomastia (8%).

Conclusion

In the pediatric aged patients, drugs modulating serotonergic receptors, transported via ABCB1, and metabolized by CYP2D6 (Ondansetron and Risperidone) are most frequently associated with reported adverse drug reactions in children. Genotyping pregnant mothers prior to prescribing Ondansetron for anti‐nausea and vomiting may improve drug safety outcomes by mitigating unnecessarily high fetal drug exposure in utero in potential poor metabolizers. Similarly, genotyping children for CYP2D6 prior to prescribing, the antipsychotic, Risperidone may improve drug safety outcomes and should be communicated as an option to parents. Pediatric drug safety opportunities exist for Obstetrics & Gynecology and Psychiatry clinics for integrating clinical pharmacogenomics into routine patient care.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Acetaminophen and Aspirin Overdose in an Extremely Preterm Neonate
Authors: Pavlek Leeann , Rogers Lynette
Published: June 2021

Introduction

Acetaminophen (N‐acetyl‐p‐aminophenol; APAP) is a widely available over‐the‐counter analgesic which is considered safe at standard dosing, but can produce significant drug‐induced liver injury and acute liver failure when taken in excess. Aspirin (acetylsalicylic acid) is a commonly used non‐steroidal anti‐inflammatory and anti‐platelet medication, which can produce significant respiratory and metabolic derangements when taken in large amounts. Accidental and intentional overdoses of these two medications are often encountered in children and adults and have widely accepted treatment protocols. There are currently no reported cases describing the presentation and treatment of a combined acetaminophen and aspirin overdose in a premature neonate.

Case Report

This case describes a premature infant born at 26 weeks gestation one day after her mother is hospitalized for management of an overdose of acetaminophen and aspirin. The mother had significantly elevated blood levels of both drugs, and underwent treatment with N‐acetylcysteine (NAC) and sodium bicarbonate. Acetaminophen and aspirin are both known to cross the placenta and can cause neonatal hepatotoxicity and coagulopathy respectively. NAC acts as an antidote to acetaminophen toxicity and also crosses the placenta, so administration is recommended to treat acetaminophen overdose in pregnancy. In this case, the baby was delivered before the treatment course was completed and subsequently showed signs of liver injury. The infant had laboratory results showing elevated acetaminophen and aspirin levels, metabolic acidosis, and prolonged PT, PTT, and INR. The neonate had resolution of her metabolic acidosis with administration of fluids containing sodium acetate. It was recommended by our neonatal pharmacist and toxicology experts to administer NAC, but the standard dosing regimen and concentration of NAC would provide an unsafe excess of fluid. This case report describes our novel treatment protocol that was formulated to provide this extremely low birth weight neonate with appropriate weight‐based doses of NAC. She clinically improved and did well while in the neonatal intensive care unit, with no further consequences of the overdose.

Conclusion

This is the first case report describing the presentation and unique management strategies of an acetaminophen and aspirin overdose in an extremely premature baby.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Aqueous extract of Alhagi Maurorum protects from norfloxacin‐induced hepato‐nephrotoxicity in rats
Authors: Khalifa Hesham Ahmed Mohammed Ismail , Abdelrahman Ahmed Shaban Abdelaziz , Sahu Ravi P
Published: June 2021

Objectives

To determine the effects of aqueous extract of Alhagi maurorum against the adverse effects including hepato‐nephrotoxicity induced by a broad‐spectrum antibiotic norfloxacin in rats.

Methods

Adult male albino rats were treated with or without norfloxacin (10 mg/kg) and Alhagi maurorum (300 mg/kg). Serum samples were collected to evaluate liver and kidney function tests, and hepatic and renal tissue samples were obtained to assess antioxidant activity and histopathological examination.

Results

Alhagi maurorum ameliorated norfloxacin induced elevated levels of serum alkaline phosphatase (ALP), aminotransferases (AST), urea, creatinine, and uric acid. In addition, Alhagi maurorum significantly reduced the tissue levels of malondialdehyde (MDA), and increased glutathione peroxidase (GPx) and superoxide dismutase (SOD) compared to norfloxacin alone treated group. Histological analysis demonstrate that Alhagi maurorum conserved the hepatic and renal tissue damages induced by norfloxacin.

Conclusion

These findings indicate that Alhagi maurorum aqueous extract possesses a potent antioxidant activity and could be used to attenuate norfloxacin induced oxidative stress and hepatorenal damage in experimental models.

Support or Funding Information

No funding support

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Trichloroacetate‐Induced Developmental Toxicity and the Production of Reactive Oxygen Species in Zebrafish Embryos (Danio rerio)
Authors: Issa Omar N , Hassoun Ezdihar A , Williams Frederick E
Published: June 2021

Trichloroacetic acid (TCA) is one of the toxic haloacetate by‐products formed as a result of the chlorination of drinking water as a part of the disinfection process. In this report, the developmental toxic effects of TCA were investigated in a nonmammalian vertebrate species: zebrafish (Danio rerio) embryos. The exposure of zebrafish embryos to (24, 32, 40, and 48 mM) TCA occurred at 4 hours post‐fertilization (hpf), and the embryos were monitored at 8, 24, 32, 55, 80, and 144 hpf for developmental landmarks from proper gastrulation to swimming and feeding success. At 32 hpf, TCA elicited a significant YSE at all of TCA concentrations when compared with the untreated controls. However, when compared with the untreated controls at the 55 and 80 hpf, a significant amount of CSE and YSE was observed in animals exposed to all of the tested TCA concentrations. Hatching delay was also produced in zebrafish embryos due to exposure to TCA concentrations of 24 and 40 mM at the 55hpf developmental time point. The hatching delay was more pronounced at the 55 hpf than 80 hpf time point. In addition, embryos developed lordosis resulting in behavioral abnormalities like abnormal feeding and swimming. The lethal dose‐50 (LD50) of TCA in zebrafish embryos was calculated as 29.72 mM. Along with the induced developmental toxicity, TCA elicited the release of superoxide anion (SA) in zebrafish embryos at 32, 55, and 80 hpf of development.

The reactive oxygen species (ROS) produced by TCA exposure coincides with the developmental abnormalities which were observed in exposed embryos. This indicates that the ROS generated may cause the developmental toxic effects of TCA exposure in zebrafish embryos.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Dynamic Culturing of Rat Hepatocytes in Sandwich‐Configuration Enhances and Maintains Formation of Biliary Networks
Authors: Deferm Neel , Stam Janine , de Witte Peter , Bouillon Thomas , Annaert Pieter
Published: June 2021

Primary hepatocytes are currently the ‘gold standard’ for evaluating hepatotoxicity of drugs and other xenobiotics in vitro, as they ensure a close resemblance to the in vivo situation in terms of activity levels of drug metabolizing enzymes and transport proteins. Hepatocyte phenotype and cell polarity is commonly maintained for several days by culturing human or animal hepatocytes between two layers of collagen in a sandwich‐configuration. Despite the pronounced positive effects of culturing hepatocytes in sandwich‐configuration, decreases in metabolic enzyme expression and activity occur rapidly. Moreover, this static in vitro system does not adequately resemble the in vivo microenvironment where hepatocytes are in close proximity with the microvascular network, which continuously exposes the cells to hemodynamic stresses such as wall shear stress. We therefore hypothesized that restoring hemodynamics by dynamically culturing hepatocytes using a shaking incubator would improve metabolic function and hepatocyte phenotype. Rat hepatocytes were cultured for 11 days either in sandwich‐configuration under static (i.e. non‐shaking) or dynamic conditions (i.e. continuous shaking). Hepatocyte phenotype, functionality, viability as well as bile canalicular morphology were assessed at multiple days of culture. Hepatocytes undergoing continuous shaking exhibited a uniform and polygonal morphology. Moreover, both static and dynamic cultures developed extensive biliary networks at day 3 of culture time. Mean (± SEM) biliary excretion index (BEI) values of dynamically cultured cells, as determined by the quantitative evaluation of multidrug‐resistance associated protein 2 (Mrp 2; Abcc2) mediated excretion of 5‐(6)‐carboy‐2′,7′‐dichlorofluorescein (CDF), did not decrease through 11 days in culture (day 3: 46 ± 3%, day 11: 45 ± 1%), whereas mean BEI values (± SEM) of static cultures declined from day 3 onwards (day 3: 45 ± 3%, day 11: 34 ± 2%). No significant difference in overall biochemical integrity between both cultures was observed, as measured by the capability of the hepatocytes to convert toxic ammonia to non‐toxic urea. Consistently, viability of the cultured cells, which was monitored longitudinally by determining the cellular release of lactate dehydrogenase (LDH), did not differ significantly between static and dynamic cultures. In conclusion, although both cultures did not differ in terms of biochemical integrity and viability, our results show that dynamically culturing rat hepatocytes in sandwich‐configuration improves maintenance of functional biliary networks for up to 11 culture days.

Support or Funding Information

Financial support for this research was provided by the Research Foundation ‐ Flanders (FWO)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The marine natural product coibamide targets expression of HER family receptors
Authors: Kazemi Soheila , Serrill Jeffrey D. , McPhail Kerry L. , Ishmael Jane E.
Published: June 2021

Natural products have historically been a significant source of inspiration for drug design and development, particularly in the areas of cancer and infectious disease. The utility of natural products as drug leads is attributed to the fact that these molecules have evolved to bind specific biological targets and have the potential to reveal new aspects of cell signaling. We previously reported the discovery of coibamide, a cytotoxic N‐methyl‐stabilized depsipeptide isolated from a marine cyanobacterium growing within Coiba National Park, Panama. Initial testing of coibamide in the National Cancer Institute in vitro 60 (NCI60) cancer cell line panel revealed pM to nM potency as a growth inhibitor, an unmatched selectivity profile and histological selectivity for breast, CNS‐derived tumors, ovarian and colon cancer cells.

We have undertaken studies to determine the mechanism of action of coibamide and have determined that coibamide selectively targets the ER secretory pathway. To date, these relatively rare natural products are known for their ability to selectively inhibit biosynthesis receptor tyrosine kinases, growth factors and cell adhesion proteins. We studied the relative expression of human epidermal growth factor receptor (HER) proteins in human HER2‐amplified and triple negative breast cancer cells following exposure to coibamide. Coibamide inhibited expression of HER family members with differential sensitivity (EGFR=HER3<<HER2). Time‐ and concentration‐dependent decreases in HER protein expression were accompanied by a dampening in AKT/MAPK signaling. Coibamide promoted proteasomal degradation of HER family proteins consistent with the action of an ER secretory pathway inhibitor. These results suggest that coibamide preferentially inhibits the biosynthesis of a subset of ER secretory pathway proteins to induce cancer cell death rather than a total inhibition of cellular secretory function. The HER family of proteins are associated with drug resistance and treatment failure in response to many modern, targeted anti‐cancer medicines and thus this work has the potential to reveal a new way to target HER signaling.

Support or Funding Information

This work was supported by the Oregon State University (OSU) College of Pharmacy, the American Brain Tumor Association (JEI) and an American Foundation for Pharmaceutical Education (AFPE) Pre‐Doctoral Fellowship in the Pharmaceutical Sciences (JDS)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Vitamin E analogs as chemotherapeutic agents and potentiators of chemotherapy in PC‐3 human prostate cancer cells
Authors: Graham Andrew , Oblad Richard , Barnes Bradley , Harmon Molli , Kenealey Jason
Published: June 2021

Vitamin E ethyl acetate (VEEA),ethyl butyrate (VEEB), and succinate (VES) are three analogs of vitamin E that have anticancer activity. VES and VEA have been shown to selectively induce apoptosis in cancer cells across a number of cell lines via mitochondrial destabilization. We have shown that VEEA is more effective at reducing cell viability than VEEB and VES in PC‐3 human prostate cancer cells at 24, 48, 72, and 96 hour time points. Using mixture design response surface methodology (MDRSM), we have identified that 90% VEEA and 10% VEEB is the ideal combination for reducing cell viability at a 48 hour time point. We have also found that a mixture of VEEA and VEEB increases the effectiveness of chemotherapeutic drugs. We conclude that Vitamin E analogs have the potential to be chemotherapeutic agents, and increase the effectiveness of chemotherapeutics in PC‐3 human prostate cancer cells.

Support or Funding Information

Bryant Adams Foundation

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inverse agonist of ERRγ inhibits the cell growth through the regulation of cell cycle and apoptosis in anaplastic thyroid cancer cells
Authors: Hwang Ji Sun , Cho Sung Jin , Hwang Hayoung
Published: June 2021

In the previous study, we investigated that inverse agonist of ERRγ (estrogen‐related receptor gamma), GSK5182, enhances the responsiveness of radioiodine therapy by the modulation of sodium iodide symporter, NIS function in anaplastic thyroid cancer cells via the regulation of ERRγ and the MAPK signaling pathway. However, there is no reports showing the direct molecular mechanisms supporting the anti‐ cancer effect of inverse agonist of ERRγ. By the screening of inverse agonist of ERRγ, NDDC‐1228 which has more potent activity compared to GSK5182 was identified. Next, the functional activity of NDDC‐1228 was evaluated in the anaplastic thyroid cancer cells. At first, we evaluated the role of NDDC‐1228 in the regulation of thyroid cancer cell proliferation. NDDC‐1228 specifically inhibited the proliferation of CAL62, anaplastic thyroid carcinoma cell line under the cancer environment mimic condition but did not affect the proliferation of other organ derived cancer cell lines. When we examine the effect on the cell cycle, NDDC‐1228 treatment effectively induced the GI‐arrest by enhancing the expression of p21 and reducing the expression of Cyclin D and CDK proteins. In addition, NDDC‐1228 suppressed the expression of survivin, XIAP1 which are the inhibitors of apoptosis as well as activated the pro‐apoptotic protein, Caspase‐3. These results suggest that inverse agonist of ERRγ has anti‐cancer effects on anaplastic thyroid cancer cells via the regulation of cell cycle and apoptosis pathways and NDDC‐1228 could be used as a lead compound for the thyroid cancer therapy.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Dynamics and Bifunctional Substrate Activation by an Arginine Drive Catalysis in Plant Chalcone Isomerases
Authors: Burke Jason , Noel Joseph
Published: June 2021

In organic synthesis, bifunctional chiral catalysts are an increasingly popular tool to position and stabilize a nucleophile and activate an electrophile for asymmetric addition reactions. It turns out that organic chemists were not first to employ this synthetic strategy, plants have been using an similar method for quite some time (~500 million years). By using techniques in structural biology, enzymology and NMR, we characterize a novel mechanism by which the enzyme chalcone isomerase (CHI) catalyzes oxa‐Michael cyclizations of 2′‐hydroxychalcones into flavanones; a key early step in the synthesis of flavonoid compounds in plants. The enzyme enhances these reaction rates by utilizing the guanidine of a conserved arginine in a bifunctional capacity to position the nucleophilic phenoxide and activate the electrophilic enone of the chalcone substrate. In addition, we reveal the structural and biochemical basis of an evolutionary neofunctionalization event, in which a divergent clade of CHIs unique to Legumes expanded it's substrate repertoire to include 6′‐deoxychalcone substrates. Together these results reveal the details of key evolutionary mechanistic innovations unique to this class of plant enzymes.

Support or Funding Information

HHMI

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.