bluuebunny/crossref_metadata_2025 huggingface dataset:
Developing Professionalism in Research through Journals Clubs
Authors: Mendez Gilberto , Prokopczuk Federico
Published: June 2021

Professional development in science begins with knowing how to communicate science effectively. Journal clubs are a great tool in supporting student's exposure to science and the way it is communicated to others. Breaking down articles helps improve a person's ability to critically appraise research while at the same time unveiling the vast world of scientific research. The California State University, Northridge (CSUN) ASBMB Student Chapter was established in 2017 to promote professionalism in biochemistry and molecular biology research. This chapter is based on specialized formal education activities, such as student‐led journal clubs to give participants the opportunity to develop essential skills and knowledge needed for a successful life as a future scientist. The structure for our journal clubs is very interactive: all participants must come to the meeting having read the paper, and a discussion leader is assigned beforehand to facilitate conversation. The leader primes the students with introductory knowledge on the subject and all students are assigned tasks while the leader asks the participants to answer specific questions on the concepts, figures, and discussion. Participants are free to ask questions and aid in the discussion throughout the meeting. Distinguished professors and researchers are also invited to give insight and an experienced perspective. The primary focus of the discussion is not to reiterate data, but instead understand experimental strategy and rationale. Journal clubs are hosted eight times a semester, and all members must be actively participating to maintain membership in the chapter. This method serves to ensure that all members improve their professional knowledge, competence, skill, and effectiveness in research. The club saw much success with this approach during our first semester, with nearly all 19 members saying that they saw much progress in their critical thinking and ability to communicate science. This experience is a gateway for many, if not all of the students who have never experienced this level of scientific engagement and discourse. Our mission is to help break the barriers of fear and self‐doubt that comes with delving into science, one journal article at a time.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Loss of Gα13 exercise‐mimetically reprograms skeletal muscle through Rock2
Authors: Kim Sang Geon , Koo Ja Hyun
Published: June 2021

Skeletal muscle is a key organ in energy homeostasis owing to its high requirement for nutrients. Heterotrimeric G proteins converge signals from cell‐surface receptors, potentiating or blunting responses against environmental changes. Here, we report that muscle‐specific deletion of Gα13 promotes reprogramming of myofibers to oxidative type, with resultant increases in mitochondrial biogenesis and cellular respiration. Mechanistically, Gα13 and its downstream effector RhoA suppressed nuclear factor of activated T‐cells 1 (NFATc1), a chief regulator of myofiber conversion, through Rho‐associated kinase 2 (Rock2)‐mediated phosphorylation at Ser243. Ser243 phosphorylation of NFATc1 was reduced after exercise, but was higher in obese animals or diabetic patients. Consequently, Gα13 deletion in muscles enhanced whole‐body energy metabolism and increased insulin sensitivity, thus affording protection from diet‐induced obesity and hepatic steatosis. Our results define Gα13 as a switch regulator of myofiber reprogramming, implying modulations of Gα13 and its downstream effectors in skeletal muscle as a potential therapeutic approach to metabolic diseases.

Support or Funding Information

This work was supported by grants from the National Research Foundation of Korea (NRF), funded by the Korean government (MSIP) (2017K1A1A2004511).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hepatocyte Specific Shp1 Deletion and Low Dose Rosiglitazone Treatment Act in Concert to Improve Liver Glucose Homeostasis in Diet‐induced Obese Mice
Authors: Leandro Joao Gabriel Bernardo , Forest Marie‐Pier , Lavallée‐Bourget Marie‐Hélène , Gomez Lilian , Moreira Rafael , Houde Vanessa , Schwab Michael , Bellmann Kerstin , Deshaies Yves , Sola‐Penna Mauro , Marette André
Published: June 2021

The protein‐tyrosine phosphatase Shp1 plays a key role in high‐fat diet (HFD)‐induced insulin resistance by negatively regulating insulin action on glucose homeostasis. Hepatocyte‐specific Shp1 knockout mice (Ptpn6H‐KO) on HFD show an improvement of hepatic insulin resistance and glucose homeostasis but also develop increased hepatic steatosis compared to their wild‐type littermates (Ptpn6fl/fl). Nevertheless, livers of Ptpn6H‐KO mice on HFD have reduced hepatocellular damage and a reduced inflammatory profile alongside with increased peroxisome proliferator‐activated receptor gamma (PPARγ) activity, suggesting a new role for Shp1 in the regulation of PPARγ and the control of hepatic lipid metabolism. Therefore, the aim of the present study was to determine whether activation of PPARγ activity using rosiglitazone could further ameliorate hepatic glucose and lipid homeostasis as well as the inflammatory profile in Ptpn6H‐KO mice. Male Ptpn6H‐KO and Ptpn6fl/fl mice were fed a HFD for 18 weeks. During the last 4 weeks the animals were treated with a very low dose of rosiglitazone (0.3 mg/kg/day). Low‐dose rosiglitazone treatment did not affect total body or adipose tissue weight, plasma levels of adiponectin or expression of PPARγ target genes in adipose tissue whereas PPARγ target genes expression was highly increased in liver. However, liver weight and liver triglycerides were increased in both rosiglitazone treated groups while plasma triglycerides were reduced by rosiglitazone. Furthermore, treatment with rosiglitazone led to an improvement in whole‐body glucose homeostasis and hepatic Akt activation both of which were even more ameliorated in HFD‐fed Ptpn6H‐KO compared to Ptpn6fl/fl mice. These results show that a low‐dose rosiglitazone treatment selectively target liver metabolism without detectable effects on adipose tissue, and our data further suggest that hepatic invalidation of Shp1 acts in concert with rosiglitazone to improve glucose metabolism in liver of HFD‐fed obese mice.

Support or Funding Information

CIHR

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Central Adiposity is a Strong Predictor of Non‐alcoholic Fatty Liver Disease (NAFLD) in South Asian Women
Authors: Albracht‐Schulte Kembra , Rosairo Shanthini , Ramalingam Latha , Wijetunge Sulochana , Ratnayake R.M.C.J. , Kotakadeniya H.M.S.R.B. , Dawson John A. , Kalupahana Nishan , Moustaid‐Moussa Naima
Published: June 2021

Incidence of non‐alcoholic fatty liver disease (NAFLD) is increasing with increasing obesity rates due to its relation with the metabolic syndrome. This is of concern in the South Asian population since they develop metabolic complications at lower levels of obesity compared to their Western counterparts. Furthermore, the relationship between NAFLD, insulin resistance and obesity is not well characterized in humans. To address this issue, we used a convenience sample of Sri Lankan adult females (n = 34) and collected anthropometric data, adipose tissue specimens (for histology), and fasted serum samples (for metabolic and inflammatory markers). Hepatic steatosis was assessed by ultrasound scanning and used to classify participants as NAFL 0, NAFL 1, and NAFL 2. Waist circumference significantly increased with increasing NAFLD grade. Participants with NAFLD had significantly higher body mass index, hip circumference, and fasting blood glucose, as well as a higher mean adipocyte area in both omental and abdominal areas, indicating a higher degree of adipocyte hypertrophy associated with fatty liver. There were, however, no differences in measures of dyslipidemia. Various adipokines were measured; however, resistin was the only pro‐inflammatory adipokine significantly elevated in NAFL 2. These findings indicate that measures of adiposity and fasting blood glucose may be important indicators of NAFLD in South Asian women.

Support or Funding Information

This research was funded by an International Research/Development Seed Grant from the Office of International Affairs at Texas Tech University, an International Research Seed Grant from the College of Human Sciences at Texas Tech University, and the Obesity Research Cluster at Texas Tech University. NMM is in part supported by the National Institutes of Health National Center for Complementary and Integrative Health (NIH NCCIH) award number 1 R15 AT008879‐01A1. KAS is a United States Department of Agriculture (USDA) National Institute of Food and Agriculture (NIFA) Fellow supported by an Agriculture and Food Research Initiative's (AFRI) Food, Agriculture, Natural Resources, and Human Sciences Education and Literacy Initiative (ELI) Predoctoral Fellowship under award number 2017‐67011‐26029. NSK was funded by the International Research Center, University of Peradeniya, Sri Lanka (InRC/2015/01).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Antioxidant and hypoglycemic effects of Justicia spicigera in kidney from diabetic rats
Authors: Martínez‐Mora José Alberto , Murillo‐Villicaña Marina , Mejía‐Barajas Jorge A. , Salgado‐Garciglia Rafael , Peña‐Montes Donovan J. , Montoya‐Pérez Rocío , Noriega‐Cisneros Ruth , Saavedra‐Molina Alfredo
Published: June 2021

Diabetes mellitus is a metabolic disease characterized by hyperglycemia, with alterations in the metabolism of carbohydrates, lipids and proteins, where there is an absolute or relative deficiency in the secretion and/or action of insulin, or both. Currently, the complications of DM with oxidative stress have been related, causing oxidative damage to biomolecules that can not be counteracted by the defense antioxidant systems, since they are diminished. Such oxidative damage can be counteracted with the administration of antioxidants such as those present in the leaves of Justicia spicigera. Solvents of different polarity were used: Hexane, ethyl acetate and ethanol, since each solvent carries different types of compounds and were tested in vitro antioxidants of: Antilipoperoxidation, reducing power, reduction of phosphomolybdenum and reduction of DPPH radical. In the reduction of the radical DPPH, the hexane and ethanolic extracts equalized the reduction of ascorbic acid with 100% and the ethyl acetate extract 70%. For the reduction of phosphomolybdenum, the hexane extract reduced it by 28%, the extract of ethyl acetate by 94%, and the extract ethanolic by 100%. The reduction of Fe3+ in the hexane extract was 9%; 17% ethyl acetate extract and the ethanolic extract reduced it by 25%. In the antilipoperoxidation technique, the hexanic and ethanolic extract inhibited lipoperoxidation by 82 and 79% respectively, while the extract of ethyl acetate by 53%. For the in vivo tests, male Wistar rats were used to obtain the kidneys. The GSH/GSSG ratio of the kidney mitochondria, the diabetic control group and the hexane and ethyl acetate extracts was of 0.8, unlike the ethanolic extract that significantly improved it to 2.0. Since in DM there is alteration in lipid and protein metabolism, it was of interest to analyze cholesterol, triglycerides, urea and serum creatinine. In the serological tests, the hexanic extract presented only hypolipidemic effect in the diabetic group. Ethyl acetate and ethanolic extracts had hypoglycemic and hypotriglyceridemic effects in diabetic rats. In conclusión, the hexane extracts, ethyl acetate and ethanol from the leaves of Justicia, are of beneficial effects of a serological and antioxidant level.

Support or Funding Information

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Hepatocyte Early Growth Response 1 (EGR1) Regulates Lipid Metabolism in Nonalcoholic Fatty Liver Disease
Authors: Magee Nancy S. , Zhang Yuxia
Published: June 2021

Objective

Nonalcoholic fatty liver disease (NAFLD) is a burgeoning health problem worldwide due to significant increases in the prevalence of obesity, insulin resistance diabetes, and hyperlipidemia. A risk factor for NAFLD is insulin resistance as it promotes the excessive accumulation of lipids into the hepatocytes, causing cellular stress and death. As a stress response gene, early growth response 1 (EGR1) plays critical roles in inflammation and tissue repair. The aim for this study is to explore a novel role of hepatocyte EGR1 in the regulation of lipid metabolism in NAFLD.

Methods

Human microarray data from NCBI GEO database (accession #GSE48452) was used to obtain relative EGR1 expression in human normal and NAFLD livers. To determine the expression of EGR1 during the development of NAFLD, C57Bl6/J mice were fed a high in fat, cholesterol, and fructose (HFCF) for up to 9 months and EGR1 levels were examined. An in vitro insulin stimulation experiment was performed in wild type and Egr1‐deficient primary hepatocytes to assess insulin response. In vivo hepatocyte‐specific Egr1 knockout (Egr1‐HKO) mice was accomplished in 2‐month‐old Egr1flox/flox male mice by tail vein administration of adeno‐associated viral vector 8 (AAV8) expressing cre recombinase driven by thyroxine binding globulin (TBG) promoter (AAV8‐Tbg‐Cre). An empty vector AAV8‐Tbg‐Null was included as a control. WT and Egr1‐HKO mice were fed either a normal chow or HFCF diet for 5 months and followed by the examination of body composition, adiposity, metabolic rates, insulin sensitivity, and liver histology.

Results

EGR1 was dramatically induced by insulin and abundantly expressed in the postprandial liver. The expression of EGR1 was significantly reduced in human and mouse NAFLD livers, suggesting a biological relevance of EGR1 downregulation during the pathogenesis of NAFLD. In vitro hepatocyte cultures showed that Egr1‐deficient hepatocytes have impaired activation of AKT but enhanced de novo lipogenesis in response to insulin. In vivo knockout of hepatocyte‐specific Egr1 led to liver steatosis and increased whole body adiposity under the chow diet. Additionally, in vivo EGR1 deficiency in hepatocytes exacerbated NAFLD progression by promoting liver inflammation and fibrosis in mice fed a HFCF diet.

Conclusions

Our studies reveal a previously unrecognized role of hepatic EGR1 in the regulation of lipid metabolism. Our findings suggest that hepatocyte EGR1 is crucial to maintain hepatic insulin response and loss of EGR1 in hepatocytes leads to liver steatosis that exacerbates NAFLD development and progression.

Support or Funding Information

NCI K22CA184146 and NIH P20 GM103549 to Yuxia Zhang.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Protective Effect of Curcumin against Nitrosamine‐induced Gastric Cancer in Rats
Authors: Waly Mostafa I. , Al‐Hinai Shaimaa , Al‐Bulushi Ismail , Rahman Mohammad Shafiur
Published: June 2021

Background

Nitrosamine acts as a carcinogenic agent and induces gastric cancer in rats. The mechanisms underlies nitrosamine‐mediated gastric cancer pathogenesis need to be investigated. Experimental studies continue to support the notion that curcumin combat cancer by acting as a dietary antioxidant.

Objective

Our study model was undertaken to investigate the preventive effect of curcumin supplementation against nitrosamine‐induced oxidative stress and carcinogenesis in rat stomach.

Methods

Forty Sprague‐Dawley rats (8 weeks old) were randomly divided into 4 groups (10 rats/group). Control group was fed a standard rat chow diet; nitrosamine‐treated group was fed a standard rat chow diet and also received intraperitonial injections of nitrosamine. The other two groups received oral supplementations of curcumin in the presence or absence of nitrosamine injection. All animals were fed ad‐libitum for 16 weeks. At the end of the experiment all rats were sacrificed and the gastric tissues were examined microscopically for gastric cancer lesions development, and for biochemical measurements of oxidative stress indices.

Results

Our results showed that nitrosamine causes gastric cancer development, as well as inducing oxidative stress in gastric tissue as evidenced by glutathione depletion and inhibition of antioxidant enzymes. The concomitant treatment of nitrosamine with curcumin significantly ameliorated the cytotoxic effects of nitrosamine.

Conclusion

The results of this study provide in vivo evidence that curcumin supplementation protects against nitrosamine‐induced gastric cancer in the used animal model.

Support or Funding Information

This research was supported by Strategic Grant Funding (CL/SQU‐UAEU/15/03) and Internal Grant Funding (IG/AGR/FOOD/17/1), College of Agricultural and Marine Sciences, Sultan Qaboos University

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 IMPACT OF THE AFFORDABLE CARE ACT ON RURAL HEALTHCARE
Authors: Boyd‐Seng Tina L. , Einstein George P. , Tulp Orien L.
Published: June 2021

In 2010 The Affordable Care Act (ACA) was passed by the U.S. Congress, in a governmental attempt to address the healthcare issues faced by the country. Designed to provide greater coverage to more American citizens, the law has caused an irreversible change in the healthcare systems of the U.S., and in many cases, costs have increased while healthcare quality has decreased. Rural regions suffer additional issues as low competition drives high costs in generally poor areas. In order to gauge the impact of these changes on some rural communities, a local rural community was surveyed using a researcher‐developed quantitative descriptive survey focused on assessing the effects on the quality of the healthcare patients receive, as well as their expenses and level of satisfaction (affordability, availability, accessibility, accommodation, acceptability) following the passage of the ACA. Also, the study sought to compare the results obtained from each different insurance option (employer‐based, self‐paying, personal based, or ACA) in terms of the view of access to healthcare since passing of the ACA. Through comparisons of average scores in each category before and after the passage of the ACA, assessments of the effects of the law on the patient‐perceived quality of healthcare can be made. Analysis of the results showed a statistically committed relationship between age and healthcare provider as well as between income and healthcare provider (both with p< 0.001), and all the comparisons between the perceptions of affordability, availability, accessibility, accommodations and acceptability before the implementation of ACA and after the implementation of ACA were significant (all p's < 0.01). An inspection of the mean differences reveals that on all questions the perceptions and/or beliefs became more negative. These results indicate that among the sampled population, the passing of the ACA has, in most cases, decreased the quality of healthcare provided to patients, at a higher cost than before the law was enacted

Support or Funding Information

Supported by Institutional Resources of USAT Montserrat and the Einstein Medical Institute, N. Palm Beach, FL, USA.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Generating 3D artificial melanoma tumor tissues using CellMate™ hydrogels.
Authors: Lyden Timothy , Dahlberg Declan , Martin Mike
Published: June 2021

We have previously reported on the successful use of natural 3D ECM materials as artificial tissue scaffolds to generate tissue‐like constructs of melanoma using the ATCC cell lines B16F1and B16F10. In that earlier work we observed that 3D constructs like these undergo a progressive scaffold colonization, tissue development and maturation in‐vitro which closely parallels tumor progression in‐vivo. Now, we have extended that work by applying a new and unique 3D hydrogel matrix called CellMate™. This material is an embedding hydrogel comprised of hyaluronate and chitosan which crosslink to generate a matrix scaffolding for cell attachment and tissue development. In these new experiments, B16F1 and B16F10 cell lines were each seeded into CellMate™ at densities of 10 and 20 million cells per scaffold. The resulting structures where then maintained for 6 months of continuous culture with periodic new plates and regular media exchanges at 3 day intervals. This approach resulted in even more physiological artificial tumors, than previously developed on ECM scaffolds. As previously seen on natural scaffolds, these new artificial tumors develop two distinct cell populations which closely reflect the normal in‐vivo tumor structures seen in the clinical setting. These two cell types include pigmented cells with a relatively dendritic appearance and non‐pigmented cells with relatively less dendritic and more rounded cell bodies. These cellular phenotypes also gave rise to distinct tissue‐level structural phenotypes as well. Pigmented tissue regions tended to present with less regular surfaces and more tendency to produce branching structures that extend away from the construct. In contrast, non‐pigmented cell regions generated large rounded and bulging features which often resembled ridges of tissue and which had very smooth surface features. Additionally these structures also appeared to be highly friable and prone to peeling off the main pigmented tumor structure. Colonies and later cell‐strains were developed from these shed structures and these retained their non‐pigmented phenotypes through multiple passages in 2D monolayers. Collectively these studies illustrate the efficacy of using 3D artificial tissues to create better in‐vitro tumor models of melanoma.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inhaled Corticosteroids Stimulate cAMP Production and Enhance βAR Signaling In a Non‐Genomic Fashion In Human Airway Smooth Muscle Cells
Authors: Corpuz Maia , Johnstone Timothy , Kazarian Austin , Ostrom Rennolds
Published: June 2021

Inhaled corticosteroids have been shown to be highly beneficial when combined with β‐adrenergic receptor (βAR) agonists in treatment of asthma. The mechanism for this synergy has never been fully clear, but corticosteroids have been shown to increase βAR expression and reverse βAR desensitization via canonical genomic actions. The current study investigated whether inhaled corticosteroids have an effect on cAMP production in both primary and immortalized human airway smooth muscle (HASM) cells. We measured real‐time cAMP dynamics in live HASM cells using the downward cAMP difference detector in situ (cADDis) cAMP sensor (Montana Molecular, Bozeman MT). Addition of various concentrations of either prednisone, fluticasone or allopregnanolone increased cAMP production over basal within 3 min without phosphodiesterase inhibitors present. We examined various passages of HASM cells and cells from different patients and found fluticasone consistently increased cAMP production at concentrations between 10 nM and 10 μM. We also examined cAMP responses to fluticasone in human embryonic kidney (HEK‐293) cells and observed no increases in cAMP production in response to fluticasone, prednisone, or progesterone. We repeated studies in the presence of glucocorticoid receptor GR‐α antagonists, RU‐486 and GSK‐9027, to understand how corticosteroids may be increasing cAMP levels. Pretreatment with either antagonist attenuated cAMP responses to all activators, including forskolin and βAR agonists, implying these drugs have considerable non‐specific effects. We also evaluated how corticosteroids would alter responses to other cAMP‐elevating agents. Fluticasone (0.1 to 1 μM) increased the potency and maximal effect of isoproterenol, PGE2 and forskolin. These findings are consistent with the idea that corticosteroids directly stimulate cAMP signaling in a non‐genomic manner and can enhance signaling by βAR and prostaglandin E receptors. Future studies will identify the molecular mechanism for this effect and determine if GR‐α is required. These findings reveal an unappreciated action of inhaled corticosteroids that may help explain how these drugs are beneficial in combination with β‐adrenergic receptor agonists for the treatment of asthma.

Support or Funding Information

This work was supported by NIH grant GM107094 (RO)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Poly(ADP‐ribosyl)ation of Signal Transducer and Activator of Transcription (STAT)6 by poly(ADP‐ribose) polymerase (PARP)‐1 is critical for its integrity, nuclear retention, and subsequent regulation of Th2 cytokines during asthma
Authors: Wang Jeffrey , Ghonim Mohamed A. , Ibba Salome Valentina , Pyakurel Kusma , Lammi Matthew R. , Boulares Hamid
Published: June 2021

Background

Although asthma is often controlled by corticosteroids and β2‐adrenergic agonists, many individuals cannot control the disease with these therapies. Identifying the key molecular mechanisms behind the disease is crucial for establishing strategies to improve the quality of life for asthmatics. Our laboratory performed a series of pioneering studies revealing the critical role of PARP‐1, classically regarded as a DNA repair enzyme, in asthma pathogenesis by controlling NF‐κB nuclear retention and STAT6 integrity. PARP‐1 inhibition destabilizes STAT6, a transcription factor central to Th2 cytokine signaling, upon IL‐4 or allergen exposure, potentially through a calpain‐dependent mechanism.

Hypothesis

PARP‐1 interacts with IL‐4‐activated STAT6 and modifies it for maintaining STAT6 integrity and trafficking, leading, ultimately, to the expression of Th2 cytokines.

Methods

I utilized cell‐free recombinant protein‐based enzymatic reactions, in vitro stimulation of primary splenocytes and PBMCs with IL‐4, histidine‐tag pulldown, immunofluorescence, and co‐immunoprecipitation with subsequent immunoblot to conduct the studies.

Results

PARP‐1 was activated in lungs and PBMCs of human asthmatics. STAT6 interacted with and was poly(ADP‐ribosyl)ated (PARylated) by PARP‐1 in both the cell‐free system and in IL‐4‐treated mouse splenocytes. PARylation of STAT6 was enhanced upon by JAK3. μ‐calpain‐mediated degradation is highly selective for STAT6, and phosphorylation of STAT6 rendered it more susceptible to degradation, but PARylation protected it from such degradation. Intriguingly, these events occurred in the complete absence of damaged DNA, as IL‐4 stimulation‐induced PARylation of STAT6 by PARP‐1 was not accompanied with a DNA damage response as assessed by p53 phosphorylation. PARP inhibition also prevented STAT6 translocation after IL‐4 stimulation in PBMCs.

Conclusion

Modification of STAT6 by PARP‐1 through a DNA damage‐independent mechanism is critical in Th2 responses and overall asthma pathogenesis. Based on the prediction that the IL4/STAT6 pathway is upregulated in asthma, concomitant with elevated phosphorylation of STAT6, activated STAT6 in cells of patients with the disease should be highly susceptible to degradation upon PARP inhibition, leading to an extensive reduction in the expression of Th2 genes.

Support or Funding Information

THIS WORK WAS SUPPORTED IN PART BY AMERICAN HEART ASSOCIATION GRANT# 149740335A/JEFFREY WANG/2017 and the Louisiana Cancer Research Center Stanley S. Scott Cancer Center

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mutations in Lyn Kinase Causes Changes in Neutrophil Function and Migration
Authors: Chini Julia A. , Bennin David A. , de Jesus Adriana Almeida , Goldbach‐Mansky Raphaela , Huttenlocher Anna
Published: June 2021

The Src Family Kinase (SFK) Lyn plays important regulatory roles in immune cells including neutrophils. As a signaling protein, Lyn has been shown to regulate neutrophil migration and endothelial adhesion, contributing to inflammation. Lyn contains a tyrosine kinase domain as well as SH2 and SH3 domains allowing for adaptor functions. A set of patients with systemic neutrophil‐induced inflammation have been shown to have de novo germline mutations in Lyn, Y508* and Y508F, affecting the negative regulatory tyrosine residue of Lyn. The purpose of this study is to evaluate the physiological, cellular, and biochemical effects of this mutation, and further understand the role of Lyn Kinase in neutrophil function. Patients were found to have a gain‐of‐function mutation and presented with skin vasculitis that improved with administration of the SFK inhibitor, dasatinib. We generated stable expression lines of WT, Y508*, and Y508F pMX‐Lyn‐GFP in neutrophil‐like PLB‐985 cells. Kinase activity assays show increased phosphorylation of SFK substrate, SAM68, and overall tyrosine phosphorylation with both Y508*and Y508F expression compared to WT, indicating higher kinase activity in the mutants. Further experiments using kinase‐dead mutants indicate that the higher kinase activity is caused by increases in Lyn's kinase activity and not its adaptor function. In addition, we evaluated the chemotaxis of PLB‐985 cells towards the chemoattractant fmlp using transwell migration assays and microfluidic devices that allow for the generation of chemoattractant gradients. Y508* and Y508F PLB‐985 cells were found to have significantly increased chemotaxis towards 10 mM fmlp (p<0.05) than WT Lyn. Our data supports the role of Lyn as a positive regulator of migration, demonstrating that disruption in the regulatory domain of Lyn can cause cellular and physiological changes. Furthermore, it highlights the role of the de novo germline mutations in Lyn as a cause of human inflammatory disease.

Support or Funding Information

Supported by the Hilldale Undergraduate Research Fellowship

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Reversal of myofibroblast differentiation by phorbol 12‐myristate 13‐acetate is PKC‐independent
Authors: Talbott George , Le Helen , Lor Paia , Jin Zhuqiu
Published: June 2021

Fibrosis often results in organ dysfunction and failure in diseases, such as: chronic heart failure, hepatic cirrhosis, pulmonary fibrosis, and end‐stage renal disease. The differentiation of fibroblasts into myofibroblasts results in the secretion of collagens and other extracellular matrix proteins that limit organ function and underlie the fundamental basis of fibrosis. TGF‐β1 induces the differentiation of α‐SMA‐expressing myofibroblasts from fibroblasts. However, the mechanism to induce reversal of myofibroblast differentiation remains elusive. Phorbol 12‐myristate 13‐acetate (PMA) has been used for the stimulation of lymphocytes and splenocytes. It is also involved in multiple cellular functions by potently activating protein kinase C (PKC). Nevertheless, the effect of PMA on the formation of myofibroblasts is unknown. To investigate whether PMA plays a role in myofibroblast differentiation, NIH 3T3 fibroblasts were cultured in DMEM medium for 48 hours in the presence of DMSO (vehicle control), PMA, or PKC inhibitors (calphostin C, chelerythrine, and staurosporine) alone or plus PMA. The expression of α‐smooth muscle actin (SMA, a hallmark of the myofibroblastic phenotype), fibroblast‐specific protein 1 (FSP1, a biomarker of fibroblasts), and TGF‐β1 in cultured fibroblasts was detected by western blotting and immunofluorescence. Expression of α‐MARCK phosphorylation was also detected to examine the activity of PKC activation. Positive staining of α‐SMA was observed in some cells in control culture conditions, and these α‐SMA positive cells exhibited significant morphology changes with large nuclei and cytoplasm, compared with α‐SMA negative fibroblasts. Treatment with PMA 50 ng/mL for 48 hours reduced the expression of α‐SMA and TGF‐β1, and pretreatment with the three PKC inhibitors mentioned above could not abrogate PMA‐induced reduction of α‐SMA and TGF‐β1. These results indicate that some myofibroblasts are derived from fibrolbasts under basic culture conditions. PMA induces reversal of myofibroblast differentiation via a PKC‐independent mechanism.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effects Of Nitrite In Goldfish (Carassius Auratus) Olfactory Function
Authors: Martinez Melody , Huertas Mar
Published: June 2021

In the last decade, freshwater ecosystems have experienced a dramatic increase of nitrogenous compounds as a result of farming and industrial activity. Aquatic ammonium (NH4+) is naturally transformed to nitrite (NO−2) by nitrifying bacteria. Nitrite is toxic to vertebrates and is known to cause disruption of physiological processes such as ion homeostasis, respiration, and cardiovascular activity. However, little is known about the effects of NO−2 on the olfactory function in teleost, which is critical for the survival of the species. This study aims to determine the physiological effects of sub‐lethal concentrations of nitrite in the olfactory function of goldfish (Carassius auratus). Thus, goldfish were exposed to 6 different NO−2 concentrations (1000 mM, 500 mM, 300 mM, 100 mM, 10 mM and 0 mM.) for 72 h. At the end of the experiment, blood, brain and nose samples were obtained for histological analysis. Additionally, in order to determine the reduction of olfactory function, olfactory responses to several food odorants were recorded by electro‐olfactogram (EOG) after exposure of the nose to several NO−2 concentrations. No mortality was observed in any treatment. Goldfish exposed to the two highest concentrations of NO−2 exhibited metahemoglobemia (darkening in the blood due to irreversible oxidation of hemoglobin). Histological analysis of the olfactory epithelium showed an increase in mucus production and mucus cells. Moreover, the olfactory epithelia of these fish showed hyperplasia of the olfactory sensory neurons. Hyperplasia of epithelium cells occurred in goldfish exposed to high NO−2 concentrations. Preliminary electrophysiology recordings showed a decrease in the olfactory function due to NO−2 treatments. These results suggest that sub‐lethal concentrations of nitrite are neurotoxic and alter the olfactory function in fish.

Support or Funding Information

Research support: Texas State University start‐up funding to Dr. Mar Huertas.

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 Clinical Implication of Compounds which could induce the function of human organic cation transporter 2 (hOCT2), while inhibit the function of human Multidrug and Toxin Extrusion Proteins 1 (hMATE1)
Authors: YANG HONG , Guo Dong , Shu Yan
Published: June 2021

The human organic cation transporter 2 and human multidrug and toxin extrusion protein 1 (hOCT2/hMATE1) transport system is a pair of transporter proteins highly expressed at the basolateral and apical membrane of renal proximal tubules respectively. They function collaboratively to facilitate elimination of a variety of compounds into urine, thus playing a crucial role in drug disposition and response. Preliminary data from our lab showed that acute exposure (20 minutes) of Cd2+ could significantly upregulate the activities of human hOCT2, while inhibit the function of hMATE1, both in a dose‐dependent manner (Fig. 1A&1B). Beside Cd2+, we identified imatinib, an anticancer drug, could induce the function of hOCT2 while inhibit the function of hMATE1 (Fig. 2). In vivo study also showed that pre‐treatment of imatinib could significantly increase the accumulation of metformin in mice kidney while had no impact on the plasma exposure of metformin (Fig. 3A&3B). It implicated that the index of plasma drug exposure will not be able to correctly reflect drug‐drug interaction when a drug could induce hOCT2 while inhibit hMATE1 at the same time, which potentially could cause significant drug accumulation in kidney tubular cells.

Support or Funding Information

NIH: R01GM099742FDA: U01FD004320

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Interaction between Adenosine and β‐adrenergic receptors in regulating vascular tone in mouse mesenteric arteries.
Authors: Yadav Vishal R. , Zhang HanTing , Mustafa S. Jamal S.
Published: June 2021

Adenosine (AR) and β‐adrenergic receptors (βR) are GPCRs involved in the regulation of cardiovascular function. Adenosine is an endogenous nucleoside which controls local blood flow through activation of four subtypes of ARs. Stimulation of A1 and A3 ARs produce contraction while A2A and A2B ARs cause relaxation in mouse aorta and mesenteric arteries (MAs). Stimulation of β1R causes increase in heart rate and contractile force whereas β2R produces relaxation of blood vessels.

Reportedly, A1AR, β1R, and β2R form heterodimers in human heart. Hetero‐dimerization between A1AR and βR leads to altered receptor pharmacology, functional coupling, and intracellular signaling pathways. Importantly, stimulation of the myocytes with isoproterenol (βR agonist) inhibits the effect of A1AR activation on the co‐localization of protein kinase C‐ɛ involving protein kinase A. Thus, a complex interaction between signaling cascades of A1AR and β1R exists, which may regulate cardiovascular function. In the present study, we aimed to determine whether interaction between β1R and ARs exists in regulating vascular tone in mouse aorta and MAs.

We measured mean arterial blood pressure (MABP), protein expression, organ bath (aorta) and DMT (MA) muscle tension in β1R KO and WT mice. Our non‐invasive CODA tail cuff BP (mmHg) data showed significant reduction in MABP in β1R KO mice. The MABP in β1R WT mice was 83.4 ± 3; whereas 68.9 ± 3 (p<0.05) in β1R KO mice. Our DMT experiments revealed that β1R was partly involved in the vascular relaxation response to isoproterenol. Significantly reduced relaxation response to isoproterenol was observed in A1AR KO mice MAs. Importantly, significantly enhanced contraction to CCPA (A1AR agonist) was observed in β1R KO mice MAs. These contractile responses were blocked by HET0016 (Cyp4A inhibitor). NECA (non selective ARs agonist) induced responses were comparable in MAs. Furthermore, our vascular studies revealed significantly reduced PE and ACh induced contraction in MAs of β1R KO mice whereas KCl induced responses were comparable. Our western blot analysis revealed higher A1AR and comparable Cyp4A expression levels in MAs. Our studies in aorta revealed comparable CCPA induced contraction in β1R WT and KO mice. In addition, comparable A1AR and Cyp4A expression levels and vascular responses to KCl, ACh and PE were observed in aorta.

In conclusion, our data revealed a possible interaction between β1R and A1AR at a vascular level in MAs. This unique receptor cross‐talk may enlighten the important signaling responses controlling vascular function.

Support or Funding Information

Supported by HL 027339

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Rapamycin PFC Nanoparticles Pre‐Treatment Mitigates Cisplatin Induced Acute Kidney Injury
Authors: Doherty Justin , Fazal Jawad , Wickline Samuel , Pan Hua
Published: June 2021

Cisplatin, belonging to the class of platinum‐containing anti‐cancer drugs, is widely used in a variety of cancers, including sarcomas, lymphomas, small cell lung cancer, squamous cell carcinoma of the head and neck, bladder cancer, testicular cancer, cervical cancer and ovarian cancer. Despite its chemotherapeutic efficacy, cisplatin induces acute kidney injury (AKI) in approximately one‐third of treated patients, which further impairs their quality of life, and thwarts successful outcomes of cisplatin treatment. Although it is known that early enhancement of autophagy and inhibition of cisplatin induced inflammation is beneficial in mitigating cisplatin induced AKI, treatment targeting both mechanisms remains to be developed. Accordingly, our hypothesis is that rapamycin PFC nanoparticles could serve this purpose via rapamycin targeting of mTOR signaling to induce autophagy and inhibit its downstream NF‐κB signaling pathway, a convergent gate keeper in inflammation. Rapamycin delivery by PFC nanoparticles enables sufficient drug delivery to the site of injury with minimum systemic exposure as well as sustained local drug release.

Fluorescence microscopic images demonstrated the delivery of rapamycin PFC nanoparticles to the kidneys, where they remained for 24 hours following i.v. injection (n=3). Therefore, we performed rapamycin PFC nanoparticles (1 ml/kg) treatment 24 hours before cisplatin injection (10 mg/kg) (n=5). Mice receiving cisplatin injection only (n=5) and mice receiving no injection (n=5) served as controls. On the day of rapamycin PFC nanoparticles and cisplatin injections as well as 24, 48, and 72 hours after the cisplatin injection, blood was collected from each mouse for a BUN test to assess kidney function. The results (Fig. A) illustrated that cisplatin treatment induced mild kidney function impairment, which was mitigated by rapamycin PFC nanoparticles pre‐treatment. Furthermore, using proximal tubule cells (NRK‐52E), we demonstrated that the benefit of rapamycin PFC nanoparticles pre‐treatment could come from 1) enhancing autophagy (Fig. B), where p62 level was reduced in the groups with the rapamycin nanoparticles treatment regardless of cisplatin treatment; 2) preventing cisplatin induced inflammation (Fig. C) through NF‐κB signaling pathway activation, where p65 upregulation was only observed in the group receiving cisplatin treatment alone; and 3) inhibiting apoptosis (Fig. C), where rapamycin pre‐treatment inhibited cisplatin induced Bax expression.

In conclusion, rapamycin PFC nanoparticles pretreatment could potentially mitigate cisplatin induced AKI in cancer patients.

Support or Funding Information

This work was supported by the NIH grant R01 DK102691.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Brain Microvascular Endothelial Derived Microparticles as a Non‐Invasive Diagnostic Tool for Multiple Sclerosis
Authors: Yun J. Winny , Boyer Chris , Minagar Alireza , Alexander J. Steven
Published: June 2021

Background

Multiple sclerosis (MS), an autoimmune‐neurodegenerative disease, exhibits features of neurovascular inflammation including endothelial activation and shedding of microparticles (MPs). While the brain lacks ‘classical’ lymphatics, we previously reported expression of venous‐lymphatic endothelial markers (LYVE‐1, podoplanin, prox‐1, VEGFR‐3, VEGF‐C, VEGF‐D) in both brain tissue and sera, with altered expression of these proteins in MS. Based on differential expression of these markers in brain tissue, we propose that these markers may originate within brain microvascular endothelium and undergo changes in disease which can be analyzed diagnostically.

Objectives

To validate the neurolymphatic biomarker matrix as a novel, non‐invasive and cost‐effective tool for diagnosing MS and discriminating MS subtypes.

Methods

Neurolymphatic biomarker expression was determined in human cerebral microvascular endothelial cells (hCMECs) following 24–48 hours of TNF‐α (20 ng/ml) exposure +/− IFN‐γ (1000 U/ml) using western blotting. MPs from these cells were visualized using scanning electron microscopy. Human sera were analyzed by flow cytometry, western and dot blotting to characterize relative partitioning of neurolymphatic biomarkers in MP.

Results

hCMECs released MPs that contain lymphatic and junctional markers as well as caveolin‐1. Plasma MPs were seen to liberate soluble (non‐particulate) neurolymphatic biomarkers upon storage, indicating that serum represents a more stable MP‐derived pool of these markers. MP flow analysis of intracellular actin revealed that permeabilization destabilizes phosphatidylserine‐Annexin V interaction which is used to designate MP identity, suggesting that flow cytometry of MPs may require more judicious interpretation. All neurolymphatic markers so far tested were able to distinguish MS from healthy control serum samples; FOXC2 expression can further distinguish subtypes of MS: RRMS from SPMS.

Conclusion

Lymphatic biomarkers in plasma and sera are expressed by brain microvascular endothelial cells and exhibit changes in expression in response to inflammatory stimuli (cytokines). Such alterations parallel transfer of these markers into caveolin‐1 ‐enriched microparticles, which represent a circulating ‘snapshot’ of the vascular surface that can be analyzed by several immune approaches, particularly following storage‐dependent solubilization of these biomarkers.

Support or Funding Information

Supported by the Annette Funicello Research Fund for Neurological Diseases and the Malcolm Feist Predoctoral Fellowship from Center for Cardiovascular Diseases and Sciences, LSU Health Sciences Center‐Shreveport

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