bluuebunny/crossref_metadata_2025 huggingface dataset:
Discerning the Mechanism of HLA Expression by Epigenetic Modulators in Breast Cancer Cell Lines
Authors: Terrigino Nicholas Taylor , Powers Rachel Elizabeth , O'Donnell Robert Watts
Published: June 2021

Our previous studies showed that the combination of a histone deacetylase inhibitor (Entinostat) and a DNA methyltransferase inhibitor (5‐Azacytidine) work in an additive fashion to upregulate surface Human Leukocyte Antigens (HLA) which are typically downregulated in cancer cells. The current study centers on Vorinostat, a histone deacetylase inhibitor, and 5‐Azacytidine, a DNA methyltransferase inhibitor, which allow us to study the regulation of HLA Class 1 expression in the breast cancer cell lines, MDA‐MB‐231 and MCF‐7. Using RT‐qPCR, we are testing for the expression of mRNAs for HLAs and the Antigen Processing Machinery (APM) responsible for placing peptides into these class I proteins and transporting them from the ER to the cell surface. The mRNA levels of APM genes TAP1, TAP2, and LMP2 showed a marked increase upon treatment with the drug combination when compared to the control. The LMP7 mRNA levels showed only a small increase upon treatment with the drug combination when compared to the control. Through flow cytometry, we have demonstrated that, in MCF‐7 cells, the combination of Vorinostat and 5‐Azacytidine caused the highest increase in HLA and Beta‐2 microglobulin expression when compared to each treatment alone. In the case of the MDA‐MB‐231 cells, the drug combination demonstrated the highest increase in Beta‐2 microglobulin expression, but not the highest increase in HLA expression when compared to each treatment alone. Future experiments are planned to determine the amount of HLA protein intracellularly, further assess the APM mRNA levels, and histone positioning at these genes. Discerning the mechanism of the HLA upregulation may help to design treatments which optimize HLA expression, and thus increase the efficacy of new immunotherapies that use checkpoint inhibitors like anti‐CTLA‐4 and anti‐PD‐1.

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 Plasmodium falciparum on Placental Expression of Inflammatory and Coagulation Factors
Authors: Dalapati Trisha
Published: June 2021

Plasmodium falciparum infection during pregnancy, which can develop into placental malaria (PM), is estimated to cause over 200,000 infant deaths annually. PM is characterized by sequestration of parasite infected red blood cells (iRBCs) and deposition of hemozoin (HZ) in the maternal blood (intervillous) space of the placenta. HZ is an insoluble, iron‐containing by‐product of the parasite's digestion of hemoglobin. Additionally, accumulation of maternal immune cells and excessive deposition of fibrin are seen in the intervillous space. The role of the syncytiotrophoblast (ST), the outermost cell layer of the placenta, during PM pathogenesis remains unclear. It is hypothesized that the ST responds to iRBCs, HZ, and inflammatory cytokines like tumor necrosis factor (TNF) produced by maternal immune cells to attenuate infection. However, it is ultimately harmed by its own response. Mechanistically, it is predicted that TNF leads to activation of tissue factor (TF), the initiator of the extrinsic coagulation pathway, which ultimately results in fibrin deposition. In addition to TNF, HZ released from iRBCs may also upregulate TF. Downstream coagulation factors activate protease activated receptors (PARs), which are G‐protein coupled receptors highly expressed on the ST. Activated PARs increase expression of TNF, propagating what is known as “the inflammation‐coagulation cycle”. The cycle's propagation adversely affects nutrient and gas exchange between the maternal blood and the ST, resulting in growth restriction, low birth weight, and occasionally death of the fetus. Using a human choriocarcinoma cell line, BeWo, and isolated human trophoblast cells, in vivo host (ST)‐parasite interactions are simulated. BeWos and human trophoblasts are left unstimulated or stimulated over a physiologically relevant time‐course with (1) lipopolysaccharide (LPS; positive control), (2) natural HZ, (3) TNF, or (4) natural HZ/TNF. RNA isolation is followed by cDNA generation and qPCR for TF, PAR‐1, and PAR‐2. Results indicate that BeWos respond to LPS and HZ with a modest upregulation of TF and PAR‐1 between 2 to 4 hours. Response to TNF occurs at 2 hours with a modest upregulation of TF. Gene expression of PAR‐2 was not found to be significant over the time‐course. qPCR results with cDNA generated from time‐course experiments using human trophoblast cells are forthcoming. Identifying the extent to which the ST is involved in PM pathogenesis will advanced the understanding of how this syndrome impacts fetal development and outcome.

Support or Funding Information

This work is supported by National Institutes of Health grant 1R21AI111242 to Dr. Julie M. Moore.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Assisted Insertion of Pseudomonas aeruginosa Type Three Secretion Translocator PopD in Membranes by Translocator PopB
Authors: Tang Yuzhou , Heuck Alejandro P.
Published: June 2021

Type III secretion (T3S) system is deployed by a wide range of pathogens to manipulate host cell response and establish infection. The T3S system is a syringe‐like apparatus that spans across the double membrane of bacteria, protruding 40nm–80nm into the extracellular space and connecting with target cell membrane. In Pseudomonas aeruginosa, the proteins PopB and PopD are secreted and found associated with the target eukaryotic cell membrane. These two proteins are believed to form a transmembrane complex or translocon to allow effector protein translocation. We have recently determined that PopD alone can spontaneously oligomerize and form mostly homo‐complexes of six subunits in the membrane. Interestingly, PopB and PopD together form membrane‐inserted hetero‐complexes that contain eight subunits of PopB and eight of PopD. However, the assembly mechanism to form these complexes remains unknown.

Here, we investigated how PopD interacts with membranes when assembled into the PopB/D hetero‐complex using a combination of biochemical and biophysical techniques that report on the environment and location of individual amino acid. Our data suggested that the presence of PopB triggers conformational change in membrane associated PopD in lipid bilayers. Using a cell based phosphorylation assay, we also found a similar conformational change of PopD in infected HeLa cell membranes. Together, our results showed that PopB assists the membrane insertion of PopD.

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 Chronic, Intermittent Sucrose Consumption on the Discriminative Stimulus Effects of Dopamine or Opioid Antagonists
Authors: Marek Morgan A. , Rothbauer Dylan R. , Schulz Emily N. , Barton Allison C. , Tryhus Aaron M. , Hardorn‐Papke Damin J. , Brandt Lucas J. , Jewett David C.
Published: June 2021

Chronic, intermittent sucrose access has been shown to induce neurochemical changes in the brain, including increasing the levels of dopamine and D3‐receptor mRNA, decreasing the levels of D2‐receptor mRNA, and increasing μ‐opioid receptor binding. We wondered if chronic, intermittent sucrose consumption would alter the ability of rats to discriminate drugs that affect endorphin and dopamine functioning. We successfully trained rats with chronic, intermittent sucrose access to discriminate various naltrexone training doses (ranging from 0.32 – 3.2 mg/kg) from saline. In the present series of studies, we sought to determine if chronic, intermittent sucrose consumption alters the ability of haloperidol to serve as a discriminative stimulus. Sprague‐Dawley rats were trained to discriminate between haloperidol [0.018–0.56 mg/kg, 30 min pretreatment (PT)] and vehicle in a two‐lever, operant choice procedure. Most subjects reached the discrimination criteria (80% or greater condition‐appropriate responding for 8 of 10 consecutive sessions). Among subjects that acquired the discrimination, there were no significant differences in acquisition between subjects with chronic, intermittent sucrose access and subjects with 24‐hr water access. Haloperidol (0.032 – 0.056 mg/kg) produced significant rate suppression. Increasing the PT to 60 minutes increased rates of lever pressing and reinforcers earned, but did not alter haloperidol generalization functions. Taken together, chronic sucrose consumption enhanced naltrexone's ability to function as a discriminative stimulus, but did not affect the discriminative stimulus effects of haloperidol.

Support or Funding Information

University of Wisconsin‐Eau Claire Office of Research and Sponsored Programs

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Characterization of a Novel Series of D4 Dopamine Receptor Ligands Reveals Structure‐Activity Relationships for Selective Partial Agonists
Authors: Day Marilyn M. , Free R. Benjamin , Stang Christopher R.T. , Boateng Comfort A. , Keck Thomas M. , Sibley David R.
Published: June 2021

D4 dopamine receptors (D4R) are G protein‐coupled receptors that play important roles in cognition, attention, and decision making. D4R ligands have been shown to alter cognition and behavior in animal models of drug addiction and variations in the DRD4 gene have been associated with novelty‐seeking and risk behavior, as well as ADHD. A better understanding of D4R‐mediated signaling is essential to understanding and treating D4R‐associated disorders, including substance abuse disorders. Despite its clinical implications, there are currently few compounds that selectively modulate the D4R receptor. Such compounds are important to study D4R function and identify favorable pharmacology for disease treatment. We created a small next‐generation compound library using computational modelling to design D4R ligands based on the parental scaffold of the prototypical D4R partial agonist A‐412997. This library was analyzed for D2R, D3R, and D4R activity using both agonist and antagonist radioligand competition binding assays and functional signaling readouts (β‐arrestin recruitment and cAMP inhibition). We found that substitutions on the piperidine and pyridine moieties, as well as modifications in linker length, led to several novel lead compounds with improved selectivity and potency, and varying degrees of agonist efficacy. Compounds were identified that displayed high binding affinity (<100 nM) for the D4R coupled with a >1000‐fold selectivity vs. the D3R and D2R. Importantly, these ligands displayed a range of agonist efficacies from highly efficacious partial agonists to full antagonists of the D4R. Interestingly, some compounds with similar binding affinities showed divergent functional efficacies, allowing for the generation of a preliminary structure‐activity relationship underlying functional efficacy. Further optimization may allow for directed tuning of functional efficacy that, when coupled to in vivo behavioral studies, may provide insight into the importance of relative efficacy levels as a means to treat D4R‐related disorders.

Support or Funding Information

NINDS IRP

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Th2‐driven Innate Immune Responses in Carbon Nanotube‐induced Lung Fibrosis
Authors: Ma Qiang , Dong Jie
Published: June 2021

Aberrant tissue responses to persistent deposition of foreign bodies frequently result in organ fibrosis through orchestrated, but poorly understood processes. Carbon nanotubes are new nano materials that have found a broad range of applications in industrial and commercial products. Animal studies indicate that certaincarbon nanotubes behave as insoluble fibers that deposit in the lungs upon inhalation and stimulate a rapid onset pulmonary fibrotic response, characterized by inflammation, interstitial fibrosis, and granuloma formation. We have used multi‐walled carbon nanotubes (MWCNTs)‐induced mouse lung fibrosisas a model to analyze the molecular underpinnings of lung fibrosis induced by foreign bodies. Genome‐wide microarray analyses of the lungs exposed to MWCNTs revealed a range of differentially expressed genes involved in pathways of immune and inflammatory regulation, response to stress and extracellular stimuli, and cell migration and adhesion. In particular, the T helper 2(Th2)‐driven innate immune responses were significantly enriched. MWCNTs significantly induced the expression of Th2 cytokines, typified by interleukin(IL)‐4 and IL‐13, and a panel of Th2 signature downstream genes, such as Il4i1, Chia, and Ccl11/Eotaxin. Induction involved the activation of STAT6 via the phosphorylation of STAT6 and the up‐regulation of GATA‐3 that directly controls the transcription of Th2 target genes. These findings demonstrate the activation of Th2‐driven innate immune responses during the development of lung fibrosis induced by carbon nanotubes, providing a molecular link between Th2functions and the tissue response to foreign body deposition in mammalian lungs.

Support or Funding Information

Supported by a fund to QM from National Institute for Occupational Safety and Health, Health Effects Laboratory Division and Nanotechnology Research 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.
Utilizing SULT4A1 Mutant Mouse Models to Characterize SULT4A1
Authors: Garcia Patrick L. , Hossain Mohammed I. , Andrabi Shaida A. , Falany Charles N.
Published: June 2021

Identified by our lab in 2000, sulfotransferase 4A1 (SULT4A1) belongs to the cytosolic sulfotransferase (SULT) gene superfamily of phase II conjugation enzymes based on key sequence and structural similarities. Broadly, SULTs catalyze the transfer of a sulfonate moiety (SO3−) to a diverse set of endogenous and exogenous substrates in tandem with the obligate cofactor 3′‐phosphoadenosine‐5′‐phosphosulfate (PAPS). Expression studies have shown SULT4A1 to be localized to neuronal tissues, and SULT4A1 orthologs have been found in all vertebrate species examined. Remarkably, human SULT4A1 shares a high degree of sequence homology with other vertebrate SULT4A1 orthologs, more than any other human SULT. Despite the conserved nature of SULT4A1 among vertebrates, all attempts to demonstrate sulfotransferase activity have failed and the neuronal function of this orphan enzyme remains unclear. To elucidate an understanding of SULT4A1, CRISPR‐Cas9 gene editing technology was performed to disrupt SULT4A1 function in a C57BL/6J mouse background and two mutations of interest were pursued. A 28 bp deletion (Δ28) in exon 1 resulted in a frameshift mutation and a premature stop codon at amino acid (AA) 62. A second mutation of interest was a 12 bp (Δ12) in‐frame deletion in exon 3 of four AA immediately adjacent to the active site histidine. Dynamic modeling studies of the Δ12 mutation suggested a structural disruption to the substrate binding pocket. Interestingly, homozygous mutant pups of both lines displayed a severe and progressive neurological pathology beginning 8–10 days post‐natal, until death at day 21–25. Neurological symptoms included persistent tremor, ataxia, absence seizures, and decreased weight gain. Homozygous mutant mice of both strains were confirmed to lack detectable SULT4A1, while heterozygous mice of both strains showed a modest decrease compared to wild‐type (WT) mice. Since little is known about SULT4A1 function, subcellular fractionation was performed to determine if SULT4A1 localizes to other cellular compartments. Indeed, SULT4A1 was detected in mitochondrial, cytosolic, and microsomal fractions, a previously unreported finding. Due to the early onset of neurological symptoms, SULT4A1 expression was assessed in primary cortical neurons (PCNs) harvested from gestational day 15 mice. PCNs were cultured in vitro to day 12 and robust expression of SULT4A1 was observed in these cells by immunostaining. In summary, SULT4A1 is a critical protein for neurofunction and normal development, as mice lacking functional SULT4A1 displayed a progressively severe and lethal neurological pathology. These mutant mouse models will be instrumental into characterizing the function of SULT4A1.

Support or Funding Information

This work was supported by GM113980 to CNF and NS08695301 to SAA.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Ocular Retention and Bioavailability Studies of Bromfenac Mucoadhesive Nanoparticles by Sustained Ophthalmic Delivery
Authors: Bhavani B. , Swathi P. Kumar , Kilari Eswar Kumar
Published: June 2021

The present study was designed to formulate the sustained ocular delivery of bromfenac loaded mucoadhesive chitosan nanoparticles (NPs). The NPS were prepared by different ratios of chitosan and sodium alginate by ionic gelation method. The prepared nanoparticles were characterized by its particle size, zeta potential, entrapment efficiency, in vitro release and ocular irritation test. The NPs are employed for transcorneal permeation and in vivo evaluation was carried out in rat eye for ocular toxicity studies. The pharmacokinetic data of selected NPs were compared with marketed bromfenac formulation (Megabrom). The selected NPs were characterized by their mean particle size 202±2.02 nm, encapsulation efficiency 73.4±1.25 % and zetapotential +32.3±0.12 mV. In vitro release exhibited biphasic drug release profile with initial burst followed by slow drug release. The NPs exhibited significant mucin adhesion. The Draize's test showed that, there was no irritation upon topical instillation of prepared NPs. The AUC (0‐α) of NPs was increased up to 4.02 fold and clearance was decreased up to 5.5 fold as compared to marketed ophthalmic formulation and were found to be safe in histopathological studies. It is concluded that he formulated bromfenac NPs showed prolong ocular residence and reducing dosing frequency.

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 Inhibitive Effect of Salidroside on Apoptosis of EA.hy926 Cells Induced by H2O2 under Simulated Microgravity
Authors: Yuan Min , Jia Mingwen , Wang Jingyu , Yuan Ming
Published: June 2021

Microgravity or simulated microgravity(SM) can induce cardiovascular dysfunction in which vascular endothelial cells play an important role. Studies showed that oxidative stress was common in vascular tissues under simulated microgravity and was also increased in vascular endothelial cells during spaceflight. The purpose of this experiment is to study whether vascular endothelial cells have different biological response to oxidative stress under simulated microgravity with new generation transcriptome sequencing technology and discuss the cell protection effect of salidroside. EA.hy926 cells, a fusion of HUVECs and the lung carcinoma cell line A549, were randomly divided into control group(Con72) and clinostat group(Z72), and after clinostating for 72 hours, the two groups were randomly given the following treatment at the same time: stimulated by H2O2, stimulation by H2O2 and salidroside intervention. Apoptosis was measured by flow cytometry and transcriptome sequencing was performed by new generation sequencing technology, RT‐qPCR was used to verify the differentially expressed genes and detect the effect of salidroside on the expression of these genes. The results showed that there were obvious apoptosis in EA.hy926 cells treated by H2O2 under simulated microgravity, which was significantly inhibited by salidroside. There were six significant differentially expressed genes screened by transcriptome sequencing, including BCL‐2A1, FAM196B, TMEM158, PPP1R16B, CXCL8, PPP1R3B. RT‐qPCR results showed that BCL‐2A1 and FAM196B were consistent with the transciptome sequencing results, meanwhile, BCL2A1 and FAM196B genes were significantly increased after salidroside intervention in H2O2‐treated EA.hy926 cells under simulated microgravity. These results showed that apoptosis was induced significantly in H2O2‐treated EA.hy926 cells under simulated microgravity, which can be inhibited by salidroside, the active ingredient of Rhodiola rosea, through the regulation of BCL‐2A1 and FAM196B gene expression.

Support or Funding Information

Funded by State Key Laboratory of Space Medicine Fundamentals and Application, China Astronaut Research and Training Center, grant No.SMFA14B01

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Actions of Nitric Oxide Synthase Inhibitors on Mitochondrial Respiration in Isolated Rat Heart Mitochondria
Authors: Dholakia Monica H. , Sakamuri Siva SVP. , Sure Venkata N. , Sperling Jared A. , Satou Ryousuke , Katakam Prasad VG.
Published: June 2021

Objective

The existence of a nitric oxide (NO) producing enzyme, nitric oxide synthase (NOS), within cardiac mitochondria has been speculated in the most recent decades. There is conflicting data regarding the identity, physiological function, and the pathophysiological significance of mitochondrial NOS. Nevertheless, mitochondrial preparations have been shown to display an ability to produce NO that is sensitive to NOS inhibitors. Our objective was to evaluate the impact of pharmacological NOS inhibition on mitochondrial respiration. We hypothesized that pharmacological inhibition of NOS affects mitochondrial respiration.

Methods and Results

Mitochondria were isolated from rats by homogenization and gradient separation. Isolated mitochondria were divided into three groups: untreated control, neuronal NOS inhibitor‐treated (nNOS; ARL dihydrochloride or ARL), and endothelial NOS inhibitor‐treated (eNOS; N5‐(1‐Iminoethyl)‐L‐ornithine or NIO). Mitochondrial function was assessed by measuring oxygen consumption rates (OCR) using Seahorse XFe Analyzer. First, we observed that isolated rat heart mitochondria displayed respiratory control ratios above 2 suggestive of the acceptable tightness of the coupling between respiration and phosphorylation indicative of the good quality of the preparation. Second, both NOS inhibitors decreased the OCR related to ATP production and increased proton leak in the mitochondria although the basal and maximal respiration were unaffected.

Conclusions

Pharmacological inhibition of NOS impairs ATP production in isolated rat heart mitochondria that is likely to be related to increased proton leak. Thus, the physiological function of mitochondrial NOS activity involves modulation of inner mitochondrial membrane permeability to promote efficiency of ATP production

Support or Funding Information

American Heart Association (PVG: 14SDG20490359 and VNS: 16PRE31450006); National Institute of Health: National Institute of Neurological Disorders and Stroke and National Institute of General Medical Sciences (PVK: R01NS094834); and National Institute of Diabetes and Digestive and Kidney Diseases (RS: DK107694).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Changes In Some Liver Enzymes Activities Of Albino Rats After Prolonged Intake Of Antihypertensives; B‐Blockers And Moduretics
Authors: ANACLETUS FRANCIS C. , ONYEGEME‐OKERENTA BLESSING MINAOPUNYE , ONWUKA ADAORA PATIENCE
Published: June 2021

Antihypertensive drugs; beta‐blockers with moduretics are the classes of therapeutics used in the management of cardiac arrhythmias, and to protect the heart from a second heart attack after a first heart attack. They play important roles in the treatment of many cardiovascular diseases. This study investigated the possible effect of prolonged use of these beta‐blockers on liver function.

Twenty wister rats with average weight of 180g each were carefully examined and grouped into 4 (5 per cage). Group 1 served as the normal control. Group 2 was administered a normal dose of the drug combination (25 mg of the B‐blocker in 50·ml of distilled water and 25 mg of moduretic in 50 ml of distilled water respectively), Group 3 was administered with overdose of the drug combination (50 mg of the B‐blocker in 50 ml of distilled water and 50 mg of moduretic in 50 ml of distilled water). Administration of the drugs was done for four weeks for these two groups. Group 4 was administered a normal dose of the drug combination but for a prolonged period of six weeks. Biochemical markers used for this test to ascertain the integrity of the liver were, Alanine transaminase (ALT), Aspartate transaminase (AST), and Alkaline phosphatase (ALP), were measured. Results obtained after 4 weeks showed a significant increase (p<0.05) in ALT of group 3(28.00±3.60) when compared to the groups 1 (8.66±0.66) and 2(14.66±1.45). ALT activities after 6 weeks showed a significant increase (p<0.05) for group 4 (34.66±2.60) when compared to groups 1 and 2. Similarly ALP results for group 3 after 4 weeks of treatment showed a significant increase (p<0.05) in activities (155.66±12.98) when compared to groups 1 (96.16±12.53) and 2 (93.73±8.44) as well as in group 4 (99.96±10.35) after 6 weeks of treatment. Furthermore, AST activities obtained in group 3 (71.33±2.60) after 4 weeks and group 4 (58.66±4.91) after 6 weeks showed a significant increase (p<0.05) when compared to the groups 1 (37.66±4.97) and 2 (42.33±8.95). The result showed that there was an average change in liver markers in the group treated with B‐blockers in combination with moduretics when taken for a prolonged period as is always prescribed for known hypertensives

Support or Funding Information

Self funding

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Reducing Disparities in the Treatment of Hypertension in African Americans Using Computational Modeling
Authors: Clemmer John S. , Pruett W. Andrew , Hester Robert L.
Published: June 2021

African Americans (AA) develop hypertension (HTN) at an earlier age, have a greater frequency and severity of HTN, and have poorer control of blood pressure (BP) as compared to the white population. The mechanisms responsible for these disparities are unknown. The multifactorial nature of HTN and the similar presentation of its etiologies complicate its treatment. Mathematical modeling provides the ability to analyze complicated interrelated effects across multiple systems. Our current model, HumMod, is a large physiological simulator that is comprised of 14 organ systems, and includes key systems that play an integral role in BP control such as neural, endocrine, circulatory, and renal systems. We have created tools that integrate HumMod to generate virtual populations that are calibrated from clinical data. Thiazide (THZ) diuretics generally reduce BP and improve cardiovascular outcomes in AA. Using de‐identified clinical data from the Genetic Epidemiology Network of Atherosclerosis (GENOA) study, we calibrated virtual populations to normotensive AA or hypertensive AA with or without THZ therapy. The clinical and virtual populations were statistically similar in multiple variables including mean arterial pressure (MAP), glomerular filtration rate (GFR), left ventricular mass index (LVMI), cardiac output, and total peripheral resistance (Figure 1). Interestingly, virtual patients that did not respond to 4 weeks of THZ therapy (ΔMAP = 0±5 mmHg) were associated with greater falls in renal hemodynamics, activation of the renin‐angiotensin system, and decreased plasma atrial natriuretic peptide (ANP) as compared to THZ responders (ΔMAP = −9±3 mmHg) (Figure 2). This suggests that differences in the hormonal response to diuretic therapy may be an important factor that prevents successful HTN treatment in AA. These data are both a proof of concept of established physiology and a tool to test existing and new hypotheses of the mechanisms of nonresponse to varying antihypertensive regimens. These techniques and the potential insights gleaned from these simulations may also have broad implications for improving BP control in other hypertensive populations.

Support or Funding Information

Supported by AHA 17POST33661071 and NIH PO1 HL51971

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Roles of Vascular Risk Factors In A Neurological Disease
Authors: Shim Joon W.
Published: June 2021

Aim

We have previously reported that angioneurins such as vascular endothelial growth factor that exert both vasculotrophic and neurotrophic activities can play a diverse role and transform into a pathogenic mediator of post‐hemorrhagic hydrocephalus in young brains. The objective of this study was to better understand the roles of vascular risk factors in the brain with an age‐related neurological condition.

Methods

Literatures reporting a relationship between vascular risk factors such as arterial stiffness and age‐related disorders like Alzheimer's disease (AD) and Parkinson's disease (PD) were surveyed. Elastin gene expression was quantified in the thoracic aorta of male mice with young and old age and of young mice fed with high fat high sucrose (HFHS) diet. Aortic stiffness was assessed in vivo using ultrasound and compared to age‐, diet‐, and gender‐matched mice with constitutive deletion of phospholipase A2 group 6 (PLA2g6). Gene expression analysis was conducted on aortic medial layer RNA and validated by elastic fiber staining. Cell‐specific effect on elastin gene expression and arterial stiffness was further tested in a mouse model with SMC‐specific impairment of PLA2g6.

Results

An inverse interrelationship between the PD and vascular risk factors including smoking and vascular stiffness were noted. Animal experiments revealed that aging and HFHS‐diet induced arterial stiffening in wild type (WT) animals but not in PLA2g6 knockout mice. The qRT‐PCR revealed that elastin gene expression was significantly elevated regardless of age in PLA2g6 impaired mice. The mRNA expression of Sp1 transcription factor was significantly increased in aged PLA2g6 knockout aorta. Mice with SMC‐specific deletion of PLA2g6 at young age showed significantly reduced arterial stiffness with elevated elastin during HFHS diet.

Conclusion

These results demonstrate that a vascular risk factor such as arterial stiffness declines with aging through modulation of age‐induced loss of elastin in arteries of mice with impaired PLA2g6 exhibiting PD symptoms. Molecular targets promoting expressions of vasculoprotective gene such as elastin along the cell‐specific pathway may provide novel therapies for the protection against an increase of vascular risk and the better understanding to the associated age‐related neurodegeneration.

Support or Funding Information

J.W.S is supported by the Fellowship from the Multidisciplinary Postdoctoral Training Program in Cardiovascular Research (NIH 5T32HL007224‐39).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
CD70 Modulates the Role of eNOS In Endothelial Cells
Authors: Pandey Arvind K. , Brown Jonathan D. , Harrison David G. , Itani Hana A.
Published: June 2021

CD70 has traditionally been identified on antigen presenting cells and interacts with CD27 on T cells, promoting proliferation and memory cell formation. We have recently shown that CD70 is important in the formation of effector memory T cells in hypertension. However, the precise cell types that express CD70 and the role of CD70 on non‐immune cells has not been investigated. Here we describe a new role of CD70 in endothelial function. We found that human umbillical vein endothelial cells express CD70 mRNA and that this is increased by > 30 fold by laminar (15 dynes/cm2) compared to oscillatory shear. Studies of mesenteric vascular reactivity showed that CD70−/− mice have markedly impaired endothelium‐dependent vasodilation to acetylcholine compared to WT mice (45± 5 vs 25 ± 4 %) at baseline. In contrast, there were no differences in relaxation responses to sodium nitroprusside. Pre‐incubating vessels with the endothelial nitric oxide synthase (eNOS) inhibitor L‐NAME blocked endothelium‐dependent vasodilatation to acetylcholine in WT mice nearly 75%; in contrast, L‐NAME had no effect on endothelial vessel relaxation in CD70−/− mice. This suggests that vessels of CD70−/− mice lack the capacity to produce nitric oxide (NO). Western blot for the endothelial nitric oxide synthase (eNOS) showed a marked reduction of this enzyme in the aortas of CD70−/− as compared to wild type mice. We found that CD70 and eNOS interact by co‐immunoprecipitation, however this association is abolished by siRNA against CD70. In additional experiments, aortas from CD70−/− did demonstrate expression of both neuronal NOS (nNOS) and inducible NOS (iNOS). In order to assess functional effects of this expression, nNOS NO production was blocked with LVNIO, resulting in further impairment in vasodilation of CD70−/− mesenteric vessels (−logEC50 of 5.4 after treatment, compared to 6.2 before treatment); however blockade of iNOS with 1400 W resulted in apparent recovery of acetylcholine‐induced vasodilation. These data indicate that loss of CD70−/− leads to destabilization of the eNOS protein in endothelial cells resulting in endothelial dysfunction. In the absence of eNOS in CD70−/− vessels, iNOS contributes to endothelial dysfunction, while nNOS has a compensatory role. These studies have identified a new role of the immune co‐stimulatory protein CD70 in modulating endothelial eNOS protein levels and vascular function.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Roles of MicroRNAs in Atherosclerosis and Neointimal Lesion Formation under Flow
Authors: Chiu Jeng‐Jiann , Chen Li‐Jing , Lee Ding‐Yu
Published: June 2021

Atherosclerosis, a vascular pathology responsible for most cardiovascular‐related morbidity and mortality, develops predictably in regions of the arterial tree in which wall shear stresses are generated by complex patterns of blood flow. It has been recognized that hemodynamic characteristics determine the location of lesions and contribute to the pathogenesis of atherosclerosis. The key cells involved in atherogenesis include vascular endothelial cells (ECs) and smooth muscle cells (SMCs). Recent evidence suggests that laminar blood flow in the straight part of the arterial tree and high shear stress modulate cellular signaling and EC function, and protect against atherogenesis. In contrast, disturbed flow in bifurcations of the arterial tree and the associated oscillatory low shear stress enhance leukocyte infiltration of the arterial wall and thus are atherogenic. However, little is known about the effect of disturbed flow on ECs, especially on their interactions with SMCs, whose phenotypic switching is significantly implicated in atherosclerosis and neointimal lesion formation. Our recent studies using microRNA (miR) assay, in vitro EC‐SMC co‐culture flow system, experimental animal models, and human specimens from patients with coronary artery disease (CAD) have identified several miRs to be involved in the formation and progression of atherosclerosis. Among these miRs, miR‐146a and 10a play atheroprotective roles in ECs against atherogenesis induced by disturbed flow. MiR‐451, by targeting Rab5a, inhibits vascular SMC proliferation and inflammation and suppresses injury‐induced neointimal lesion formation. Our findings help the discovery of new target biomarkers and elucidation of functional mechanisms underlying atherosclerosis, thereby facilitating the development of new approaches for therapeutic interventions.

Support or Funding Information

This work was supported by grants from the Ministry of Science and Technology in Taiwan (grant numbers: MOST‐106‐2633‐B‐009‐001, 105‐2321‐B‐400‐007, 106‐3114‐Y‐043‐021)

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 Effect of Dietary Nitrate on Vascular Function in Chronic Kidney Disease
Authors: Ramick Meghan G. , Kirkman Danielle L. , Stock Joseph M. , Muth Bryce J. , Chirinos Julio A. , Edwards David G.
Published: June 2021

PURPOSE

The purpose of this study was to test the hypothesis that an acute dose of 12.6 mmol dietary nitrate in the form of concentrated beetroot juice (BRJ) would improve measures of microvascular and conduit artery endothelial function in patients with moderate to severe chronic kidney disease (CKD).

METHODS

Fifteen adults with moderate to severe CKD participated in this double blind, randomized, crossover study (61±4 yrs; 11 males; eGFR 49.2±3.4 ml•min−1•1.73 m2). Participants reported to the laboratory and baseline blood pressure (BP) was measured and a baseline blood sample obtained for determination of NO metabolites (NOm; Nitrate, nitrite, s‐nitrosothiols and metal bound NO). Participants were then randomized to ingest 12.6 mmol of BRJ or a nitrate depleted placebo (PLA). Vascular testing began 2.5 hours post beverage ingestion. A second blood sample was obtained and post‐ingestion BP was recorded. Microvascular function was assessed via the cutaneous response to local heating measured by laser Doppler flowmetry coupled with microdialysis, and conduit artery function was assessed via brachial artery flow‐mediated dilation (FMD). Arterial stiffness (via carotid‐femoral pulse wave velocity, PWV) was also measured. The entire protocol was repeated a minimum of 7 days later in the other condition.

RESULTS

Plasma NOm was not different between conditions at baseline but was significantly increased 2.5 hours post BRJ ingestion compared to pre‐ingestion values, as well as compared to post PLA values (PREPLA 53.0±11.6 POSTPLA 33.2±9.9 mM; PREBRJ 57.3±21.4 vs POSTBRJ1068.0±106.6 mM). Mean arterial pressure (MAP) was not different between conditions at baseline or 2.5 hours post, however the change in MAP from pre‐ingestion baseline to 2.5 hours post ingestion was significantly different between conditions (ΔMAPPLA 2.0±2.1 vs ΔMAPBRJ −3.2±2.6 mmHg, p>0.05). The plateau phase of the cutaneous response to local heating was significantly greater in the BRJ condition compared to PLA (PLA 83.4±2.2 vs BRJ 88.8±1.7%CVCmax; p<0.05). The NOS mediated contribution to the plateau phase as assessed via L‐NAME infusion was not different between conditions. We did not observe any improvements in FMD or PWV.

CONCLUSION

An acute dose of 12.6 mmol dietary nitrate significantly improves microvascular function in patients with moderate to severe CKD.

Support or Funding Information

This research was supported by an ACSM Foundation Research Grant from the American College of Sports Medicine Foundation and NIH grant HL113514.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Translational Traits of a Swine Model of CKD: Inflammation
Authors: Engel Jason Edward , Williams Maxx , Guise Erika , Drummond Heather , Chade Alejandro
Published: June 2021

The cardiovascular and renal anatomy and physiology of the pig are virtually identical to humans. We recently developed a model of CKD induced by dietary dyslipidemia and bilateral renal artery stenosis in pigs that develops a sustained decrease in renal function (comparable to CKD stage 2) and progressive parenchymal injury. Inflammation plays a role in the increased cardiovascular and all‐cause mortality in CKD and may precede renal fibrosis. Thus, we aim to characterize inflammation in CKD using markers shared by human and pigs to support the translational traits of the model.

Ten pigs were divided into either normal or CKD (n=5 each). CKD/sham was induced and pigs observed for 14 weeks. Blood pressure was continuously measured by telemetry. Total RBF and GFR were quantified in vivo using multi‐detector CT. Blood and urine were collected, then pigs were euthanized and ex vivo studies performed to quantify plasma creatinine, blood urea nitrogen (BUN), renal injury markers kidney injury molecule 1 (KIM‐1) and neutrophil gelatinase associated lipocalin (NGAL) (urine, ELISA), renal inflammation (total macrophages, pro‐inflammatory M1‐IDO‐ and anti‐inflammatory M2‐MRC1‐subpopulations) and fibrosis (trichrome)

Following fourteen weeks of CKD, pigs showed reduced RBF, GFR, hypertension, elevated plasma creatinine and BUN. These were associated with elevated urinary KIM‐1 and NGAL, inflammation (indicated by significant M1‐IDO+ macrophage infiltration over M2‐MRC1), and fibrosis (Figure).

The decline in renal function associated with significant macrophage infiltration and increased markers of renal injury. KIM‐1 and NGAL have been suggested to initially attenuate inflammation and promote a pro‐repair M2 macrophage phenotype. However, they may have been insufficient to prevent the shift towards pro‐inflammatory M1s in our model, suggesting prolonged imbalance between inflammation and repair beyond the acute phase of renal injury. The use of markers shared by pigs and humans underscored the translational potential of this model for CKD research as a feasible platform to study and possibly target inflammation.

Support or Funding Information

Support: AHA‐18490005 and intramural grant UMMC to ARC.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
A Boolean Model of Microvascular Rarefaction to Predict Renal Outcomes in Renovascular Disease
Authors: Guise Erika , Pruett W. Andrew , Chade Alejandro R.
Published: June 2021

Renal microvascular (MV) rarefaction, a hallmark of renovascular disease (RVD), associates with decreased renal vascular endothelial growth factor (VEGF) and progressive renal damage. We designed a novel bioengineered fusion of an elastin‐like polypeptide (ELP) with VEGF, and demonstrated that ELP‐VEGF therapy decreases MV rarefaction, improves renal function, and reduces renal injury in RVD. Renal angioplasty (PTRA) of a stenotic kidney is a frequently used treatment in RVD that often does not ameliorate renal injury. We thus aimed to create a Boolean model of MV rarefaction to predict the renal outcomes of an RVD kidney after ELP‐VEGF therapy following PTRA.

We first created a Boolean model with 20 factors known to be involved in MV rarefaction and progression of renal injury in RVD (assuming each to be either totally present/active or totally absent/inactive) to predict the possible pathophysiological states and outcomes after treatments. The model predicts that MV rarefaction and fibrosis remain active following PTRA, driven by a feed‐forward cycle of inflammation and oxidative stress leading to glomerulosclerosis and fibrosis (Figure). When ELP‐VEGF therapy followed PTRA in the simulation, MV rarefaction, glomerulosclerosis, and fibrosis were inactivated. The model predicted that injurious processes driving progressive renal deterioration are decreased only with ELP‐VEGF therapy following PTRA.

We next performed in vivo studies. RVD was induced by renal artery stenosis in 12 pigs fed a high‐cholesterol diet. Six weeks later, single‐kidney blood flow (RBF) and filtration (GFR) were quantified in vivo using multi‐detector CT, then pigs were randomized to either PTRA (RVD+PTRA, n=6) or PTRA followed by single intra‐renal injection of ELP‐VEGF (RVD+PTRA+ELP‐VEGF, n=7). Pigs were observed for 4 additional weeks; in vivo CT studies were repeated, then pigs euthanized and ex vivo renal micro‐CT quantification and fibrosis determined. Administration of ELP‐VEGF following PTRA improved RBF, GFR, MV density, and fibrosis more efficiently than PTRA alone.

Conclusion

The Boolean model predicted the in vivo/ex vivo data and confirmed pathological mechanisms and the key role of MV damage in renal injury progression. Our results may support the use of Boolean simulations to predict renal outcomes after treatments. Furthermore, this study unravels a simple but useful tool that may help to design, confirm, or deny pre‐clinical studies and predict therapeutic effects.

Support or Funding Information

AHA 18490005 (ARC), Intramural Research Support Program (IRSP) grant from UMMC (ARC), and PO1 HL 51971 (WAP)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Obesity and PM2.5 Exposure Effects Cardiac Function During Exercise in Mice
Authors: Grimmer Jacob , Tanwar Vineeta , Katapadi Aashish , Youtz Dane , Schwieterman Neill , Baer Lisa , Stanford Kristin , Wold Loren
Published: June 2021

Objective

Studies have demonstrated that particulate matter with diameter <2.5μm (PM2.5) exposure is linked to cardiovascular disease (CVD). Risk factors, such as obesity, correlate with higher occurrences of CVD. We aimed to study PM2.5 effects on cardiac function of obese mice and whether exercise could alter function.

Methods

Obese mice (Ob/Ob) (leptin deficient, FVB background) were exposed to either filtered air (FA) or PM2.5 (within the annual average range of 15 μg/m3 according to National Ambient Air Quality Standards (NAAQS)) for 6 h/day, 7 days/week for 9 months. The mice were divided into four groups: (1) FA sedentary, (2) FA treadmill exercise, (3) PM2.5 sedentary, and (4) PM2.5 treadmill exercise. After 8 weeks of exercise, echocardiography was used to examine global cardiac function. Following sacrifice, cardiomyocytes were isolated for studying cardiomyocyte function. Separate groups of sedentary mice were sacrificed for studying baseline myocardial mRNA expression.

Results

Echocardiography showed increased left ventricular end systolic (LVESd) and diastolic (LVEDd) diameters and decreased posterior wall thickness during systole (PWTs) and diastole (PWTd) in ob/obPM2.5 sedentary mice compared to ob/ob FA sedentary mice. Morphological alterations were associated with lower systolic function as indicated by reduced percent fractional shortening (%FS) in ob/ob PM2.5 sedentary mice compared to ob/obFA sedentary mice. These functional results are suggestive of contractile dysfunction leading to left ventricular volume overload. Cardiomyocytes isolated from sedentary ob/obPM2.5 mice showed no change in peak shortening (%PS) but marked reduction in positive and negative contractile velocity (±dL/dT) and increased calcium transient amplitude (Δ340/380) suggesting cellular contractile dysfunction. Comparison of mice after 8 weeks of exercise training demonstrated significant decrease in %FS, increase in LVESd and no change in LVEDd in ob/obPM2.5 exercised mice compared to ob/obFA exercised mice, suggesting systolic dysfunction. On comparing ob/obPM2.5 exercise and ob/obFA exercise with their respective sedentary counterparts decrease in %FS, PWTs and PWTd and increase in LVESd was observed. qPCR data showed increased inflammatory markers ICAM‐1, VCAM and CRP in ob/obPM2.5 sedentary mice compared to ob/obFA mice. These results indicated exercise did not improve cardiac function following exposure to PM2.5.

Conclusions

PM2.5 exposure in combination with obesity induced cardiac dysfunction as seen through echocardiography. Exercise training, however, did not improve cardiac function in obese PM2.5 mice.

Support or Funding Information

This work is supported by funding from National Institutes of Health (RO1ES019923) to LEW.

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