bluuebunny/crossref_metadata_2025 huggingface dataset:
Distilling Research Projects into Graphical Abstracts: Interactive Training During a Summer Research Fellowship
Authors: Aleksunes Lauren M , Laskin Debra L
Published: June 2021

Short‐term research experiences such as summer fellowship programs provide students with an opportunity to develop their research skills, explore career options, and improve written and oral communication skills. Most summer research programs for undergraduate students require a final poster or podium presentation. At Rutgers University, we developed an interactive session for undergraduates to conceptualize and draft written and graphical abstracts that would summarize the major findings of their research. These were distributed at our research symposium. During week 7 of a 10‐week summer undergraduate research fellowship, students learned about the major components of an abstract using a variety of examples. Students were also taught common pitfalls to avoid in communicating research findings. After the discussion of each of the opening sections (title, introduction, hypothesis, and methods), students were provided 5‐minutes for writing. After each portion of the abstract was composed, students volunteered to share their writings with the group, and the moderator provided constructive comments. Different examples of graphical abstracts were also presented to the students. Thirty students submitted abstracts during week 9 of the summer fellowship program which were assembled into booklets for the final research presentations. This interactive activity received the highest programmatic rating of the summer fellowship with a mean score of 4.58 (±0.59 SD) on a Likert scale of 1 to 5 with 5 representing the highest possible rating. Other weekly activities were also well‐received with mean scores ranging from 3.44 to 4.60. Taken together, an interactive session that includes didactic instruction and student responses allows undergraduate students to begin writing and designing their written and graphical abstracts as part of a summer research fellowship.

Support or Funding Information

Supported by NIEHS R25ES020721, P30ES005022, ASPET, and the SOT Intern Program.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Client‐based Classroom Undergraduate Research Experiences (CUREs) Benefit Faculty
Authors: Wisinski Jaclyn A , Cooper Scott T
Published: June 2021

Faculty are expected to establish and maintain research productivity, while demonstrating continuous innovation and improvement in teaching, especially at primarily undergraduate institutions (PUIs). These expectations can be quite daunting for early career faculty. However, a client‐based classroom undergraduate research experience (CUREs) can help faculty “clients” start research projects, provide professional development, recruit trained undergraduate research students and gain experience. At the University of Wisconsin‐La Crosse, each semester at least four different 300 and 400‐level biology and biochemistry courses, use the CURE model with fellow faculty members, often junior faculty, as clients. CURE instructors advise clients on the design and feasibility of specific projects in the multi‐week framework of the specific CURE course. With a reasonable research plan in mind, clients present the background, rationale and hypothesis as well as a basic outline of the experimental plan to students. In class sizes of 20–30, teams of 2–4 students work collaboratively on a research product (assay, mutant, clone, a set of data) and provide research documentation in the form of a complete lab notebook and final report. Clients and CURE instructors work together to guide students though each step of the experimental process, defining the molecular bases of techniques along the way. In the end, clients receive a formal lab report, research product and lab notebooks from students. In post CURE project surveys and interviews, clients reflected on the experience(s) in terms of usefulness to their research program, instructional development and career advancement. Instructor experiences were also evaluated in interviews. Based on responses by clients and CURE instructors, best practices could be established to make client‐based CURE a sustainable model for faculty development.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Caryophyllene Oxide is More Potent than Beta Caryophyllene in Attenuating the Abuse‐Related Effects of Ethanol
Authors: Oppong‐Damoah Aboagyewaah , Wood Bo Jarrett , Blough Bruce , Murnane Kevin Sean
Published: June 2021

17 million adults in the United States had an alcohol use disorder (AUD) in 2012 and current treatments yield disappointing remission rates. In recent years, the endocannabinoid system has been shown to modulate the abuse‐related effects of ethanol. In this study, we used the loss of righting reflex (LORR) assay to assess the direct pharmacodynamic interactions between beta‐caryophyllene (BCP), its derivative caryophyllene oxide (B Car 21), and ethanol. We investigated the effects of B Car 21 in two widely accepted animal models of ethanol abuse: two bottle choice ethanol consumption and ethanol‐induced conditioned place preference (CPP). Finally, we determined whether there is any pharmacokinetic interaction between B Car 21 and ethanol using blood and brain ethanol analysis. B Car 21 augmented the ethanol‐induced LORR at a dose tenfold lower than BCP. This effect was reversed by pretreatment with a selective CB2 antagonist, AM630. B Car 21 significantly decreased ethanol intake and preference without any effect on total fluid intake in mice that consumed high amounts of ethanol. In mice that consumed low amounts of ethanol, B Car 21 had no effect on ethanol intake, preference, or total fluid intake. B Car 21 significantly attenuated the expression of an ethanol‐induced CPP. Our findings show that B Car 21 is more potent than BCP in vivo and that its attenuation of the abuse‐related effects of ethanol is likely mediated through pharmacodynamic rather than pharmacokinetic interactions. This study expands the support for the CB2 receptor as a viable target for the treatment of AUD.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Protective Role of MCPIP1 in Brain Injury Following Hypoxic‐Ischemia in Neonatal Mice
Authors: Naik Radheya , Chang Yingzi , Han Byynghee Henry
Published: June 2021

Neuroinflammation has been implicated with neurodegenerative diseases and acute brain injuries such as stroke. Monocyte chemotactic protein‐1‐induced protein‐1 (MCPIP1) is a multifunction inflammatory modulator known to have pro‐inflammatory or anti‐inflammatory effects depending on the nature of brain injury. However, its role in brain injury after asphyxia in the developing brain has not been studied. Therefore, we explored the role of MCPIP1 in acute neurodegeneration after hypoxic‐ischemia (HI) in the neonatal mouse model. Neonatal C57BL/6 wild type mice at postnatal day 7 (P7) underwent a carotid artery ligation and exposure to hypoxia (8% oxygen) for 42 minutes. After the HI insult, we determined the time‐course and the spatial distribution of MCPIP1 expression in the brain. To determine the impact of MCPIP1 in acute brain damage, we assessed brain tissue loss 1 week post‐HI brain injury in MCPIP1‐deficient versus wild type mice. Western blot analysis was used to determine MCPIP1 protein expression in the brain. Caspase‐3 activity assays were used to determine apoptotic cell death, and immunoflorescent double‐labeling was utilized to determine the scope of neuroinflammation in different types of cells including astrocytes, microglia, and neuronal cells. Our data revealed that MCPIP1 protein is expressed in the neonatal brain and its expression appears to increase 24 hours after HI insult. Immunostaining showed that MCPIP1 expressing cells are present in both neuronal and non‐neuronal cells of the brain, and that there are increased numbers of MCPIP1‐positive cells in the ipsi‐lateral hemisphere compared to the contra‐lateral hemisphere. Furthermore, there was significant change in brain tissue loss after HI in MCPIP‐1‐deficient mice compared with wild‐type mice. Our data suggest that HI brain injury appears to induce expression of MCPIP1, supporting a possible role of MCPIP1 in neuroinflammation after HI.

Support or Funding Information

This study is funded by KCOM‐ Biomedical Graduate Program, Warner/Ferumaturo Grant, and ATSU Start‐up Fund (BHH).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Assessing the Neuroprotective Effects of Docosahexaenoic Acid in 6‐Hydroxydopamine Induced Parkinsonism in Rats
Authors: Chitre Neha M , Wood Bo , Ray Azizi , Moniri Nader H , Murnane Kevin S
Published: June 2021

Parkinson's Disease (PD) is the second most common neurodegenerative disease throughout the United States. Dopamine (DA) Replacement Therapy is the standard treatment for the motor symptoms of PD. However, there is growing interest in studying newer therapeutic compounds. Polyunsaturated omega‐3 fatty acids have been associated with neuroprotective roles. In this study, we aimed to investigate the neuroprotective effects of Docosahexaenoic Acid (DHA) in 6‐Hydroxydopamine (6‐OH‐DA) unilaterally lesioned rats. Behavioral assays were conducted for the assessment of locomotor activity, stride length and limb asymmetry. Neurochemical analysis of DA levels and its major metabolite DOPAC in the striatum was performed using high performance liquid chromatography (HPLC‐ED). Biochemical estimation of oxidative stress markers like Glutathione and Reactive Oxygen Species (ROS) was conducted. Semiquantitative measurements of total Tyrosine Hydroxylase (TH) and Phospho‐TH (Ser 40) in striatal homogenates were assessed using Western Blot analysis. Our results demonstrate that DHA significantly improved stride length and limb asymmetry in the parkinsonian rats. No significant improvement was observed in locomotor activity. DHA significantly improved dopamine levels in the lesioned striatum along with a significant decrease in oxidative stress markers. There was a significant increase in expression of total Tyrosine Hydroxylase (TH) and Phospho‐TH (Ser 40) with DHA treatment. In conclusion, DHA protected the dopaminergic neurons and led to increased dopamine levels and improved motor function. We are currently studying the specific role and mechanism involved to establish a link between DHA mediated neuroprotection and increased levels of Tyrosine Hydroxylase. These results support targeting free fatty acid systems for the development of new therapeutics for PD.

Support or Funding Information

Studies were supported by the National Institute of Neurological Disorders and Stroke, a component of the National Institutes of Health, by grant number NS095239. Moniri N – Principal Investigator; Murnane KS – Co‐Investigator

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Dendrobii Officinalis Flower improved learning and memory impairment by mediating antioxidant effect in Senescent Rats
Authors: Lei shanshan , Yan Mei qiu , Lv Guiyuan , Chen Suhong
Published: June 2021

Aging is a necessary process, with the main feature of structural and functional decline, adaptability and resistance weakened, accompanied with obvious feature of memory dysfunction. At present, the aging of population has become the basic situation in China. Elderly people aged over 60 had reached 130 million in the year of 2007, accounting for 10% of the total population. It is expected that to 2020 year, the aging population will reach 2.24 to 3 million. To the year of 2050, 60 years of age will be more than 400 million. Therefore, it is important to study methods to delay senility to improve the quality of life of elderly and prevent and control the incidence of senile diseases.

Dendrobii Officinalis Flower (DOF), which is belong to Dendrobium officinale, is very popular to people for the effect of anti‐aging in China, but lack of corresponding pharmacological experiments. The aim of was to evaluate whether DOF has effect on antioxidant enzymes and improving the brain function in aging rats. DOF was administrated at a dosage of 1.38, or 0.46 g · kg−1 once a day, as well as with a normal control group and senescent control group for 8 weeks. The results showed that DOF could improving the learning function of the escape latency, total distance and target quadrant time at 1.38 g · kg−1 and 0.46 g · kg−1 in Morris water maze experiment, as well as increasing level of γ‐GABA. In addition, DOF (1.38 g · kg−1) could decrease the levels of MDA, Glu and MAO‐B, improving locomotors, SOD and GSH‐P x. Histopathological study showed DOF also could significantly reduce structural lesions and neurodegeneration in the hippocampus. In conclusion, DOF could alleviate brain aging and ameliorate the bodily functions, which may be due to attenuating body's oxidative stress, improving hippocampus damage, and balance the release of neurotransmitters.

Support or Funding Information

This study was supported by the National Key Research and Development Program (No. 2017YFC1702200), the National Science Foundation of China (No. 81673638, No. 81874352, No. 81703772 and No.81803760), the China Postdoctoral Science Foundation (No. 2018M632506)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Elucidating Anti‐Inflammatory Signaling Paradigm at the 5‐HT2A Receptor
Authors: Billac Gerald , Nichols Charles
Published: June 2021

Given the recent resurgence of clinical interest in the application of classical serotonergic psychedelics as treatment for various disease states, a comprehensive analysis of the structure‐activity relationship (SAR) is necessary to fully understand the differential expression of behavioral and physiological effects elicited by these compounds. While the field is mostly focused on action within the central nervous system, previously published data from the Nichols laboratory supports the use of serotonin (5‐hydroxytryptamine; 5‐HT) 2A receptor (5‐HT2AR) agonists as potent modulators of tumor necrosis factor alpha (TNF‐α)‐mediated inflammatory responses in the periphery. Whereas whole organism models will provide insight into the physiological response profile, the mechanism of action and structural basis mediating the anti‐inflammatory activity have yet to be elucidated on a molecular level. In this study, we are characterizing the signaling pathway activation profile of a comprehensive panel of 5HT2AR ligands comprising three chemical classes: tryptamines, ergolines, and phenethylamines. We hypothesize that a distinct signaling pathway elicited by 5‐HT2AR activation is mediating the differential anti‐inflammatory response to these ligands. With this data, the varying degree of effector pathway bias can also be determined for each ligand, which, from a pharmacological standpoint, will support better understanding of adverse and favorable therapeutic activities, as well as, to inform structure‐activity studies and drug candidate selection matrices. Initial results from examination of the canonical Gαq pathway – using a calcium mobilization measure – and recruitment of the multi‐adaptor protein β‐arrestin2 – using a luciferase reporter gene assay – exhibit no significant correlation between potency and efficacy for either pathway with in vivo measures of anti‐inflammatory activity. Ongoing studies are examining additional GPCR‐mediated signaling (Gi, Gs) via comparative analysis of cAMP accumulation. The collective analysis of a comprehensive panel of 5HT2AR ligands has never been examined in this fashion; thus, we aim to elucidate the SAR to inform drug design for novel molecules that are biased to pathways targeting a biologically therapeutic response.

Support or Funding Information

Eleusis PBC

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inter‐Subunit Conformational Dynamics and Allosteric Modulation of Metabotropic Glutamate Receptors
Authors: Levitz Joshua , Gutzeit Vanessa , Thibado Jordana
Published: June 2021

Metabotropic glutamate receptors (mGluRs) are class C G protein‐coupled receptors (GPCRs) which function in synapses throughout the central nervous system where they respond to the excitatory neurotransmitter glutamate to initiate intracellular signaling cascades. A comprehensive understanding of mGluR activation requires a mechanistic description of the effects of both extracellular ligand binding domain(LBD)‐targeting “orthosteric” and transmembrane domain (TMD)‐targeting “allosteric” compounds, both of which have therapeutic potential. Here we use a combination of high resolution, time‐resolved functional and conformational assays to dissect the mechanisms by which allosteric compounds activate and modulate mGluRs. First, using an electrophysiological assay we find that mGluR2 can be activated by both orthosteric or allosteric ligands and that allosteric activation does not require LBD closure. We then use a single molecule fluorescence‐based subunit counting assay to find that mGluR transmembrane domains efficiently dimerize with variable efficiency to a degree that is consistently higher than that observed for class A GPCRs and is not sensitive to allosteric drugs. Next, we develop an inter‐TMD FRET assay that reveals that activation‐associated inter‐TMD rearrangement occurs independently of allosteric input from the LBDs. Using this assay, we test a panel of allosteric compounds to uncover diversity in apparent affinity, efficacy and kinetics between these compounds. Strikingly, we find that negative allosteric modulators are either neutral or induce TMD rearrangement which correlates with degree of inverse agonism. Together this data is consistent with a model where multiple (>2) intersubunit reorientations are part of the activation process and motivates further work on the role of multimeric assembly and inter‐domain coupling in the activation of class C GPCRs.

Support or Funding Information

This work is supported by an NIGMS R35 (R35 GM124731) grant to JL. JKT is supported by an NSF GRFP.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
YM‐254890 is a General Inhibitor of G Proteins
Authors: Peng Qianman , Shen Jianzhong
Published: June 2021

Small molecular chemicals that target individual subtype of G proteins including Gs, Gi and Gq has been lacking, except for pertussis toxin that has been validated as a selective peptide inhibitor of the Gi protein. Recently, a cyclic depsipeptide compound termed YM‐254890 was isolated from the culture broth of Chromobacterium sp., which was reported as a selective inhibitor for the Gq protein by blocking GDP exchange of GTP on the α subunit of Gq complex; however, functional selectivity of YM‐254890 towards various G proteins remains unclear, primarily due to its restricted availability before 2017. Here, using primary human coronary artery endothelial cells (HCAEC) as a testing model, we performed a systemic pharmacological study on the functional selectivity of YM‐254890 on multiple G protein‐mediated receptor signaling. First, we confirmed that YM‐254890, at 30 nM, abolished 100 μM UTP activated P2Y2 receptor‐mediated Ca2+ signaling, indicating its potent inhibition on the Gq protein. However, unexpectedly, pretreatment of HCAEC with 30 nM YM‐254890 also significantly suppressed (>50%) 100 μM adenosine‐induced cAMP elevation by Gs‐coupled adenosine A2 receptors; furthermore, we found that 30 nM YM‐254890 significantly reversed CXCL12 activated CXCR4‐Gi protein mediated suppression of cellular cAMP level, suggesting a blockage of Gi proteins. Western blotting assay indicated that YM‐254890, at 30 nM, abolished MAPK signaling triggered by P2Y2, CXCR4 and adenosine A2 receptors. Nonetheless, no cellular toxicity was observed for YM‐254890 up to 1 μM, and it neither affected A23187‐induced Ca2+ signaling, nor forskolin‐induced cAMP elevation and growth factor‐induced MAPK signaling. We conclude that YM‐254890 is not a selective inhibitor for the Gq protein, instead it acts as a broader spectrum inhibitor for Gq, Gs and Gi with a high potency, without affecting non‐GPCR‐mediated cellular signaling.

Support or Funding Information

Supported in part by NIH grant 1R01HL125279‐01A1 (to J.S.)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Palmatine Identified from Malaysian Local Plant Possess In vitro and In vivo Antidiabetic and Antioxidant Effect on High‐Fat Diet‐Induced Diabetic Rat Model
Authors: Okechukwu Patrick Nwabueze , Ekeuku Sophia Ogechi , Sharma Mridula , Fromming Gabriele Ruth Anisah
Published: June 2021

Background

Coscinium fenestratum (CF) (Gaertn.) Colebr is from the family Menispermaceae, is a woody plant found in Southeast Asia including Malaysia. Traditionally CF has been used for the treatment of many diseases including diabetes Mellitus. Our previous research identified Palmatine a protoberberine alkaloid to be the active bioactive compound present in CF.

Objective

The aim of this study was to investigate in vitro and in vivo anti‐diabetic, antioxidant Effect on High Fat Diet Induced Diabetic Rat Module.

Method

The ability of Palmatine to inhibit reactive oxygen specie (ROS) such as Lipid peroxidation, DPPH, Nitric oxide, Hydroxyl, free radical scavenging activity was evaluated. The in vitro antidiabetic activity of Palmatine was evaluated using alpha glycosidase, alpha amylase and Dipeptidyl peptidase IV enzyme. Sprague dawley rats weighing 180–200g was fed with high fat diet (HFD) for 60 days to induce diabetes. Rats with plasma glucose level of 11mmol/L or 200mg/dL and above after 60 days HFD feeding, were orally administered with Palmatine (2mg/kg) for 90 days. The plasma glucose level and body weight of the rats were measured weekly. After treatment, the rat blood was withdrawn via cardiac puncture for hematological studies. Liver and pancreas was harvested for antioxidant, histology, biochemical and lipid profile analysis.

Result

Palmatine was able to inhibit free radicals generated by Nitric oxide, DPPH, Hydroxyl, and alpha glucosidase, and alpha amylase activities. Palmatine was able to reduce plasma glucose level, activate antioxidant enzymes such as superoxide dismutase, catalase, glutathione reductase and inhibit lipid peroxidation in HFD induced diabetic rats. Biochemical and hematological analysis showed a reduction of diabetic markers in rats such HbA1c, urea, alkaline phosphatase, aspartate transaminase, alanine aminotransferase, LDL‐cholesterol, Triglycerides, but increase HDL level.

Conclusion

Palmatine possess antidiabetic and antioxidant in HFD induced diabetic rat model and able to offer protection to kidney, liver and cardiovascular organs by reducing the disease markers (Diabetes Complication Prevention).

Support or Funding Information

Centre of Excellence for Research, Value, Innovation and Entrepreneurship Research Grant Scheme UCSI University (UCSI‐CERVIE‐RGS Proj‐in‐FAS 039)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Impact of Prenatal Dexamethasone on Circulating Cytokines and Chemokines in Neonatal and Adult Rats
Authors: Madhavpeddi Lakshmi , Hammond Bradley , Hale Taben , Handa Robert
Published: June 2021

Dexamethasone (DEX) is a synthetic glucocorticoid administered to mothers at risk for pre‐term labor. We and others have shown that prenatal DEX produces long‐term changes in the hypothalamic pituitary adrenal axis and cardiovascular function in adult offspring. However, there is limited information regarding the longitudinal impact of in utero DEX exposure on the immune system. In the present study, we determined the plasma levels of 20 cytokines and chemokines in male and female neonatal and adult rats that had been exposed to DEX in utero. Pregnant dams were administered DEX (0.4mg/kg or 0.1mg/kg per day, s.c.) or vehicle on gestation days 18–21. A subset of rats was sacrificed at post‐natal day 0 (PND0) and remaining rats were weaned at post‐natal day 21 and sacrificed at 2.5 – 3 months of age for measurement of plasma cytokine and chemokine levels. At PND0, plasma Il‐1β was reduced only in DEX‐exposed males, while the lymphocyte chemoattractant IP‐10 was only increased in DEX‐exposed females. In contrast, the levels of monocyte chemoattractant protein‐1 (MCP‐1) at PND0 were elevated in both males and females that were exposed to DEX in utero. Although increased MCP‐1 levels continued to be elevated in adult females that were prenatally exposed to DEX, this chemokine was significantly reduced in DEX‐exposed adult males. Moreover, DEX‐exposed male, but not female adult offspring displayed reduced circulating levels of Il‐2, Il‐17α, Il‐18, and TNFα. Taken together, prenatal DEX results in persistent and sex‐specific changes in circulating levels of inflammatory mediators. Further studies will determine the long‐term consequence of this age‐ and sex‐related changes of DEX on immune function.

Support or Funding Information

ABRC ADHS14‐082990

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
ANTIMICROBIAL ASSAY OF CARESOO ELECTROLYZED WATER: Part II. Comparative Study on the Antimicrobial Activity of Sulfamylon solution, Dakin's solution, commercially available hypochlorous acid and the electrolyzed water from Caresoo hypochlorous acid water generator
Authors: Atanacio Ronald , Malbas Fedelino , Mondoy Melisa , Cavo Betty , Galit Salvacion Rosario , Alvarado‐Delacruz Kristine , Sarte Roan May , Atanacio Enriquito
Published: June 2021

In the initial study conducted, the electrolyzed water from Caresoo hypochlorous acid water generator affirmed to have bioactivity against pathogenic microorganisms. Part II of this study is to compare the bioactivity of sulfamylon solution, Dakin's solution, and commercially available hypochlorous acid against electrolyzed water from Caresoo hypochlorous acid water generator.

Electrolyzed water was prepared and tested against Staphylococcus aureus, Methicillin Resistant Staphylococcus aureus (MRSA), Methicillin Resistant Staphylococcus epidermidis, (MRSE) Pseudomonas aeruginosa, Streptococcus pyogenes, Proteus mirabilis and Candida albicans. The antimicrobial activity of the electrolyzed water was compared with the bioactivity of the full strength Dakin's solution, 100% sulfamylon solution and commercially available hypochlorous acid in 3 replicates using Mac Farland.

The electrolyzed water exhibited bioactivity in all test organisms less than 15 seconds after inoculation. The same results were obtained in commercially available HOCl (V). Full strength Dakin's solution and 100% sulfamylon solution showed complete antimicrobial activity after 30 minutes of inoculation.

The antimicrobial activity of the electrolyzed water from Caresoo® Hypochlorous Acid Water Generator affirms its potential as a pharmacological agent in treating and controlling infection as compared to full strength Dakin's solution and sulfamylon solution.

Support or Funding Information

Researchers funded this project.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Histone Deacetylase Inhibitors Attenuate Hepatic Fibrosis through Suppression of Group 2 Innate Lymphoid Cells and Type 2 Inflammation
Authors: Iyer Abishek , Loh Zhixuan , Fitzsimmons Rebecca L , Reid Robert C , Ramnath Divya , Clouston Andrew , Irvine Katharine M , Powell Elizabeth E , Schroder Kate , Stow Jennifer L , Sweet Matthew J , Fairlie David P
Published: June 2021

Deposition of collagen and matrix proteins is essential for tissue repair and remodelling. Chronic liver diseases typically show excessive matrix protein deposition or crosslinking that leads to scar tissue, obstructs blood flow and disrupts liver functions, all characteristics of advanced liver damage in cirrhosis. There are no effective anti‐fibrotic therapies. Immunomodulatory mechanisms that contribute to fibrogenesis in chronic liver disease are not fully elucidated. Here we examine thioacetamide‐induced liver injury in mice, which develop progressive hepatic inflammation and fibrosis similar to clinical features of chronic human liver disease. We find that inhibitors of class I, but not class II, histone deacetylase (HDAC) enzymes suppress hepatic inflammation and fibrosis in this model. Inhibitors of class I HDACs attenuate accumulation and activation of group 2 innate lymphoid cells (ILC2), specifically interleukin (IL)‐33 (but not IL‐25) dependent CD45+/Lin−/Thy‐1+/ST2+/KLRG1int ILC2 cells, associated with type 2 inflammation that drives hepatic stellate cells to secrete excessive matrix proteins. Some HDAC inhibitors that are registered anti‐cancer drugs might be repurposed for treating chronic liver fibrosis.

Support or Funding Information

ZL acknowledges salary support from the Centre for Inflammation and Disease Research, University of Queensland. The National Health and Medical Research Council of Australia is acknowledged for a Career Development Fellowship to KS (1141131), a Senior Research Fellowship to MJS (1107914) and a Senior Principal Research Fellowship to DPF (1117017).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Silencing Fatty Acid Binding Protein‐4 Through Percutaneous Retro‐orbital Injections Attenuates Inflammation in ApoE−/− Mice Liver
Authors: Prathipati Priyanka , Rodriguez‐Aguayo Cristian , Walton Brian , Sood Anil , Lopez Gabriel Berestein
Published: June 2021

Background

Fatty acid binding protein‐4 (FABP4) is a lipid chaperon is an important regulator of atherosclerosis and inflammation. We hypothesize that silencing FABP4 could be a novel therapeutic treatment for atherosclerosis, in turn liver inflammation. Hence a liposomal small interfering RNA (siRNA) delivery system to silence FABP4 was developed.

Methods

Male ApoE mice (N=16) were fed high fat diet from 6 weeks of age until 12–16 weeks. At 11 weeks of age, the mice were injected with 0.1ml (500µg/ml) of FABP4/control siRNA liposomes tagged with fluorescent Qdot (QD) 625. The injections were administered through percutaneous retro‐orbital route twice a week for four consecutive weeks. Starting in the 12th week, two mice from each group were sacrificed every week. Aortas, serum and livers were collected at the time of sacrifice, parts of these organs were stored in formalin and processed for paraffin sections and part of them were flash frozen for protein analysis. FABP4 in aortas and livers along with the pro‐inflammatory markers (IL‐6) in livers were analyzed by western blot and immunohistochemistry. This investigates the effect of FABP4 siRNA liposomes on atherosclerosis and inflammation in liver.

Results

Silencing FABP4 in ApoE mice attenuates the inflammation in liver compared to the control siRNA treatment (Figure).

Conclusion

Percutaneous retro‐orbital injections of FABP4 siRNA liposomes for four consecutive weeks might help to attenuate liver inflammation in ApoE mice.

Effect of FABP4 silencing on liver inflammation

Silencing FABP4 attenuated the expression of pro‐inflammatory marker in the livers of ApoE−/− mice.

Support or Funding Information

This work was supported by Gillson Longenbaugh Foundation (9091‐001‐N‐1 & G2015)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Vitamin D Promotes Anti‐inflammatory Phenotype as Opposed to Proinflammatory State of Macrophages
Authors: Othman Tesneem , Jarjes Lelein , Hermes Erin L , Tauseef Mohammed , Siddiqui Rizwan , Faridi Hafeez M , Khan Mohammed , Newaz Mohammad , Wu Mei X , Shahid Mohd
Published: June 2021

Vitamin D (Vit‐D) deficiency is linked to several metabolic diseases such as atherosclerosis and obesity. However, the underlying mechanism remains elusive. Immediate early response gene x‐1 (IEX‐1)—which is negatively regulated by Vit‐D—is recently shown to play a crucial role in the pathogenesis of obesity and atherosclerosis. Its paucity prevents anti‐to‐pro‐inflammatory shift in macrophage phenotype and thereby protecting against these diseases. We hypothesize that Vit‐D protects against the metabolic disease partly by inhibiting pro‐inflammatory macrophages via repressing IEX‐1. We cultured RAW 264.7 cells, a murine macrophage cell line in DMEM, and pretreated them with different doses of Vit‐D (0.1, 1, and 10 nM) for 8 hours. The cells were then differentiated into pro‐ or anti‐inflammatory macrophages by treatment with LPS (10 ng/ml) and IL‐4 (10 nM), respectively, for 24 hours in the absence/presence of Vit‐D. Total cellular RNA was extracted using Trizole and cDNA was synthesized. The mRNA levels of IEX‐1, pro‐, and anti‐inflammatory genes were determined using quantitative RT‐PCR. The protein levels of TNF‐α and IL‐10 were measured in culture supernatant using ELISA. LPS treatment increased mRNA levels of proinflammatory genes IL‐6 (60±6‐fold, p<0.001), TNF‐α (8±0.7‐fold, p<0.01), and iNOS (10±0.7‐fold, p<0.01) in RAW cells. Pretreatment with 10 nM Vit‐D but not 0.1 or 1 nM significantly inhibited LPS‐induced increases in expression levels of IL‐6 (24±2‐fold, p<0.01), TNF‐α (4±0.3‐fold, p<0.05), and iNOS (4.7±0.3‐fold, p<0.05 versus LPS alone). Vit‐D also suppressed LPS‐induced TNF‐α protein production by macrophages (18±0.8 versus 27±1.4 ng/ml, p<0.05). On the other hand, IL‐4 treatment induced expression levels of anti‐inflammatory genes Arg‐1 (93±18‐fold, p<0.01), MRC‐1 (2±0.1‐fold, p<0.05), and Clec10a (2.2±0.2‐fold, p<0.001), suggesting the formation of alternatively activated macrophages. Interestingly, pretreatment with Vit‐D dosedependently potentiated IL‐4‐induced increases in Arg‐1, MRC‐1, and Clec10a mRNA levels. Specifically, Vit‐D at 10 nM markedly increased expression levels of Arg‐1 (287±51‐fold, p<0.01), MRC‐1 (6±0.8‐fold, p<0.05), and Clec10a (6±0.8‐fold, p<0.001 versus IL‐4 alone) induced by IL‐4. Vit‐D also increased IL‐4‐induced IL‐10 protein production (510±41 versus 748±36 pg/ml, p<0.05). Concurrently, Vit‐D treatment dose‐dependently repressed IEX‐1 expression in RAW cells, confirming negative regulation of IEX‐1 by Vit‐D. These results suggest that Vit‐D promotes the anti‐inflammatory state of macrophages likely in part, by repressing IEX‐1 expression.

Support or Funding Information

None

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Expression Regulation of Hox Genes by Perfluorochemicals in Mouse Liver
Authors: ZHANG YUE , CHENG XINGGUO
Published: June 2021

Homeobox (Hox) genes encode homeodomain proteins, which play important roles in the development and morphological diversification of plants and animals. Perfluorochemicals (PFCs), which are the persistent environmental pollutants and frequently detected in human blood, can interfere with animal development. PFCs produce numerous hepatic adverse effects, such as increased peroxisomes, hepatomegaly and altered lipid metabolism, primarily via activation of peroxisome proliferator‐activated receptors (PPARs) and/or constitutive androstane receptor (CAR). No study has reported that PFCs can regulate the expression of Hox genes. This study was designed to determine the expression regulation of Hox and paraHox genes by perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), and perfluorodecanoic acid (PFDA) in mouse liver. A single dose of 0.1 mmole/kg PFOA, PFNA and PFDA was intraperitoneally administered to adult male wild‐type, and age‐matched male PPARα‐null and male CAR‐null mice (n=6/treatment). Four days later, mouse livers were collected and processed. Messenger RNA expression of Hox genes (including Hox‐a to ‐d subfamily members) and paraHox genes (including Gsx, Pdx, and Cdx gene family members) were determined. Our results showed that PFOA, PFNA, and PFDA all induced mRNA expression of Hoxb7, c5, d10 and Zeb2 in wild‐type mouse liver, but not in PPARα‐null mouse liver. We further showed that PFNA induced mRNA expression of Hoxa5, b7, c5, c8, and d10 in livers of wild‐type and CAR‐null mice, but not in PPARα‐null mouse liver, indicating a PPARα‐dependent mechanism. Furthermore, knocking‐out of PPARα or CAR in mouse liver decreased constitutive mRNA expression of Hoxc5. Wy‐14643, a classical PPARα agonist, induced mRNA expression of Hoxc5, but not of Hoxa5, b7 or d10 in wild‐type mouse liver. 1,4‐bis‐[2‐(3,5,‐dichloropyridyloxy)] benzene (TCPOBOP), a classical CAR agonist, also induced mRNA expression of Hoxc5, but not of Hoxa5, b7 or d10 in wild‐type mouse liver. Messenger RNA expression of studied paraHox genes were not apparently altered by PFCs in mouse liver. In conclusion, perfluorochemicals induced mRNA expression of certain Hox genes such as Hoxc5, maybe via activation of PPARα and/or CAR.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterization of the Expression and Function of Rheostat Locations within the Na+/Taurocholate Cotransporting Polypeptide
Authors: Ruggiero Melissa , Swint‐Kruse Liskin , Hagenbuch Bruno
Published: June 2021

Background

The Na+/taurocholate cotransporting polypeptide (NTCP) is a sodium dependent bile acid transporter expressed at the basolateral membrane of human hepatocytes. NTCP is important for the enterohepatic circulation of bile acids, such as taurocholate. It also transports the steroid hormone estrone‐3‐sulfate and the cholesterol lowering drug rosuvastatin. We have previously identified rheostat positions in NTCP. These are positions at which replacement of an amino acid to all other 19 amino acids results in a gradual change in function, whereas all‐or‐nothing changes are known as toggle locations. One of these rheostat positions is the naturally occurring polymorphic position S267.

Aim

The purpose of the current study is to characterize the effects of amino acid replacements at S267 with respect to expression and function.

Methods

NTCP mutants were generated using site‐directed mutagenesis, and the mutated proteins were transiently expressed in HEK293 cells. Surface biotinylation assays were performed to determine plasma membrane expression levels. Functional characterization was achieved using radiolabeled model substrates: [3H]‐taurocholate, [3H]‐estrone‐3‐sulfate, and [3H]‐rosuvastatin. After determining initial linear rate conditions, concentration dependent uptake was measured and kinetic parameters were calculated using the Michaelis‐Menten equation.

Results

Surface biotinylation experiments suggest that many mutants are expressed at a similar level as wild‐type NTCP. However, S267Q which showed slightly decreased taurocholate uptake, normal estrone‐3‐sulfate transport, and significantly decreased transport of rosuvastatin in our initial experiments, was expressed in extremely low amounts at the protein and membrane level. Thus, after normalization, S267Q exhibited a two‐ to ten‐fold increase in function. Another mutant, S267W, showed decreased taurocholate uptake but stimulated transport of estrone‐3‐sulfate and rosuvastatin. Comparison of this mutant's kinetics with that of wild‐type NTCP suggests that the affinity for taurocholate is decreased while the affinity for rosuvastatin is increased, explaining the observed initial altered transport.

Conclusions

Our results suggest that the functional outcome of mutations at S267 is mainly due to changes in substrate affinity rather than protein expression. Future experiments will assess whether this is also true for other rheostat positions.

Support or Funding Information

P20 GM103549, P30 GM118247, GM077336, and W. M. Keck Foundation

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inhibition of Cyclic Nucleotide Efflux by Placental MRP Transporters Enhances Trophoblast Syncytialization
Authors: Gorczyca Ludwik , Aleksunes Lauren
Published: June 2021

In the placenta, multinucleated syncytiotrophoblasts arise from the fusion of cytotrophoblasts, a process regulated by cyclic nucleotide (cAMP and cGMP) signaling. Syncytiotrophoblasts secrete hormones and express efflux transporters, including the multidrug resistance‐associated proteins (MRP) 1 and 5. Understanding the interplay of cyclic nucleotides, efflux transporters, and syncytialization is important because aberrant cytotrophoblast functioning contributes to the pathogenesis of placental diseases and toxicities. In the present study, we sought to determine whether MRP transporters regulate the syncytialization of human placental cells. BeWo b30 trophoblasts were treated with vehicle, MK‐571 (25 μM), 8‐Bromo‐cAMP (100 μM), forskolin (25 μM), IBMX (200 μM), or their combination for up to 24 hr. Markers of trophoblast cell fusion (glial cells missing homolog 1, GCM1; syncytin 2) and hormone secretion (human chorionic gonadotropin, hCGa and hCGb) were quantified by qPCR. Treatment of BeWo cells with MK‐571, a pharmacological inhibitor of MRPs, increased intracellular concentrations of cAMP and cGMP by 15‐ and 19‐fold, respectively. Moreover, MK‐571 treatment up‐regulated cell fusion and hormone secretion markers (GCM1, hCGa, hCGb, syncytin 2) by 20–200% under basal conditions. Similarly, MRP inhibition with MK‐571 further enhanced syncytialization and hormone markers following stimulation with the permeable cAMP analog 8‐Bromo‐cAMP (2‐ to 6‐fold), activation of cAMP synthesis with forskolin (20–160%), or inhibition of cAMP/cGMP breakdown by IBMX (50–130%). In addition, shRNAs were used to generate a stable BeWo B30 trophoblast cell line with reduced MRP5 expression. Control and MRP5‐knockdown cells were treated with vehicle or 8‐Bromo‐cAMP for 24 hr. In response to treatment with 8‐Bromo‐cAMP, the induction of GCM1, hCGa, and hCGb mRNA expression was higher in MRP5 knockdown cells as compared to control cells. Furthermore, MRP5‐knockdown cells were larger in size (including greater numbers of multinucleated cells) and exhibited decreased staining of the tight junction protein E‐cadherin, pointing to enhanced syncytialization. In conclusion, MRP transporters participate in trophoblast cell fusion, which likely represents a novel mechanism regulating placentation and placental toxicities.

Support or Funding Information

Supported by F31ES029794, R01ES029275, T32ES007148, P30ES005022.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Involvement of organic cation transporter 2 (hOCT2) and multidrug and toxin extrusion proteins (hMATEs) in renal elimination of meta‐iodobenzylguanidine (mIBG)
Authors: López Quiñones Antonio J , Wagner Dave J , Wang Joanne
Published: June 2021

Radiolabeled meta‐iodobenzylguanidine (mIBG) is utilized as both a diagnostic imaging agent (123I‐mIBG) and targeted radiotherapy (131I‐mIBG) for neuroblastoma. mIBG is cleared predominantly by the kidney through glomerular filtration and active secretion. Whole‐body scintigraphy showed rapid uptake and extensive accumulation in non‐tumor tissues, which can interfere with mIBG tumor imaging quality and lead to peripheral toxicities. Although mIBG is known to be transported into neuroblastoma tumor cells by the norepinephrine transporter (NET), the molecular mechanisms underlying mIBG transport in the kidney and non‐tumor tissues are unclear. We hypothesize that human organic cation transporters 1–3 (hOCT1–3) as well as multidrug and toxin extrusion proteins (hMATE1/2‐K) are involved in mIBG renal secretion and uptake in non‐tumor tissues. To test this hypothesis, the uptake of mIBG was measured in HEK293 cell lines stably transfected with hNET, hOCT1, hOCT2, hOCT3, hMATE1 and hMATE2‐K followed by LC‐MS/MS quantification. The transepithelial transport of mIBG was further evaluated utilizing a MDCK cell line co‐transfected with hOCT2 and hMATE1 to model secretion in renal proximal tubule cells. Uptake studies in the HEK293 system demonstrated that mIBG is a universal substrate for hOCT1, 2, 3, hMATE1 and 2‐K. Kinetic analysis further revealed that hOCT1, hOCT2, and hOCT3 transport mIBG with similar apparent affinities (Km of 17.5 ± 5.4, 17.2 ± 2.8, 15.5 ± 6.6 μM, respectively) which are comparable to hNET (Km of 35.7 ± 8.9 μM). In the hOCT2/hMATE1 MDCK cells, the basal‐to‐apical (B‐to‐A) flux of mIBG is much greater than its A‐to‐B flux and increased by 20‐fold as compared to B‐to‐A flux in vector transfected control cells. The apparent B‐to‐A permeability of mIBG in the hOCT2/hMATE1 cells was 12.6 ± 2.7 cm/sec, which is 18‐fold greater than that in control cells. Interestingly, the intracellular accumulation in the hOCT2/hMATE1 cells was increased only by 1.9‐fold as compared to the control cells, suggesting that apical hMATE1 efficiently effluxes mIBG from renal cells after mIBG uptake via OCT2. Together, our data demonstrated that mIBG is efficiently transported by hOCTs and hMATEs and that the hOCT2/hMATEs pathway is involved in active secretion of mIBG in the kidney. Inhibition of hOCT2 and/or hMATEs by other drugs may alter mIBG renal clearance and/or intrarenal accumulation, leading to clinically significant drug‐drug interactions during 131I‐mIBG therapy.

Support or Funding Information

This work is supported by NIH Grants GM066233 and T32‐GM007750.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.