bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of non‐clinical safety of Frutalin in adult Zebrafish (Danio rerio)
Authors: Monteiro‐Moreira Ana Cristina Oliveira , Vieira‐Neto Antônio Eufrásio , Ferreira Maria Kueirislene Amâncio , Mendes Francisco Rogênio Silva , Barros Adriana Rolim Campos , Magalhães Francisco Ernani Alves , Moreira Renato Azevedo
Published: June 2021

Our previous studies have shown several biotechnological and biomedical activities of Frutalin (FTL), a lectin from Artocarpus incisa seeds. Its multiple specificity for α‐D‐galactose and a‐D‐mannose promotes affinity for glycans present in various cell receptors, such as ion channels. FTL presents isoforms, according to its deconvolved mass spectrum, which increases the biotechnological potential of the molecule, allowing the prospection of new applications. This study was conducted to evaluate the nonclinical safety of FTL in adult Zebrafish (Danio rerio). The action of FTL (1.0 mg/mL; 20 μL; ip) on the locomotor activity of adult zebrafish (Danio rerio) (ZFa; n = 6/group) was evaluated in the open field test.1 As controls were used one group without treatment (Naive) and another with vehicle (Saline 0,9%, 20 μL, ip). After 24 h of treatments, mortality was analyzed and the concentration able to kill 50% (LC50) of the zebrafish was calculated.2 The experimental procedures were approved by the Ethics Committee for the Use of Animals (CEUA), under protocol no. 7210149/2016, of the State University of Ceará (UECE), Fortaleza, Ceará. Despite interacting with other cellular receptors and promoting other pharmacological activities, FTL, active in the physiological environment of Zebrafish, did not modify the locomotor activity nor did it prove to be toxic to the animals, suggesting that its lectin activity, capable of promoting pharmacological activities when recognizing glycans of several ion channels, and membrane protein receptors, can not be associated with toxicity or locomotor damage.

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.
OAT3 Mediated Interaction between Steviol Acyl Glucuronide and Probenecid: In Vitro and In Vivo Investigations
Authors: Chow Dandan
Published: June 2021

Steviol acyl glucuronide (SVAG) is a circulating metabolite formed after the ingestion of rebaudioside A, a naturally occurring sweetner with functional activities. Our previous studies have demonstrated that transporters particularly OAT3 played an important role in renal clearance of SVAG. The present study aimed to investigate the inhibitory effect of probenecid on OAT3 mediated SVAG transport using in vitro and in vivo systems. In the hOAT3 stably transfected HEK293 cell line, probenecid exhibited a dose‐dependent inhibition of SVAG uptake, with an IC50 value of 4.95 μM. In rat kidney slices, the uptake of SVAG was markedly inhibited by probenecid (67.5%), estrone‐3‐sulfate (41.9%), and para‐aminohippurate (PAH, 27.0%), respectively. In rats given an oral dose of 15 mg/kg of rebaudioside A, the Tmax, Cmax and AUC6–8 of SVAG were 6.25 hr, 176 ng/mL and 213.8 ng/mL*hr, respectively. In contrast, an intravenous dose of probenecid (20 mg/kg) led to an increase of Cmax and AUC by 2.15 and 1.71 folds, respectively, while Tmax remained relatively unchanged. Taken together, the inhibition effect on SVAG uptake in the presence of probenecid was mediated via OAT3. As OAT3 is a major uptake transporter in the kidney, the inhibition of OAT3 activity may alter SVAG's renal clearance. It is thus prudent to investigate transporter‐mediated food‐drug interactions and ensure safe and effective use of those products.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Molecular mechanisms of organic cation and anion transport at the blood‐CSF barrier
Authors: Hu Tao , Zha Weibin , Duan Haichuan , Wang Joanne
Published: June 2021

The blood‐cerebrospinal fluid (CSF) barrier, formed by epithelial cells of the choroid plexus (CP), plays an active role in removing neurotoxins, drugs, and metabolites from the brain. Organic cations and organic anions are known to be transported at the blood‐CSF barrier, but the underlying molecular mechanisms have not been fully understood. The goal of this study is to identify major organic cation and anion transporters expressed in human and mouse CP and to elucidate the functions of these transporters by live tissue imaging using fluorescent probe substrates. For organic cations, real‐time PCR analysis showed a predominant expression of plasma membrane monoamine transporter (PMAT) in both human and mouse CPs. In contrast, organic cation transporters 1–3 (OCT1–3) and the multidrug and toxin extrusion protein 1 and 2 (MATE1/2) are not or minimally expressed. In freshly isolated CP from wildtype mouse, live tissue imaging showed that IDT307, an analog of the organic cation MPP+, was transported into CP epithelial cells at the apical (CSF‐facing) membrane and sensitive to inhibition by the PMAT inhibitor quinine. Once inside CP epithelial cells, IDT307 was highly accumulated in the mitochondria with little efflux at the basolateral (blood‐facing) membrane. IDT307 uptake and intracellular accumulation was greatly attenuated by ~70% in CP tissue from the Pmat knockout mouse. For organic anions, real‐time PCR analysis showed the highest expression of the organic anion transporting polypeptides 1C1 (OATP1C1) and multidrug resistance‐associated protein 1 (MRP1). Organic anion transporter 3 (OAT3/Oat3), MRP4/Mrp4, Oatp1a4/5, OATP1A2 and OATP3A1 also showed significant expression in human or mouse CP. Unlike IDT307, the organic anion fluorescein‐methotrexate (FL‐MTX), which is a reported substrate of OATPs and MRPs, was rapidly taken up at the apical membrane and efficiently excreted into the capillary areas with little intracellular accumulation. Rifampicin, a pan‐inhibitor of OATPs, completely blocked FL‐MTX uptake into the CP tissue whereas MK571, a potent inhibitor of MRP1/4, abolished the basolateral efflux of FL‐MTX, resulting in its high accumulation within the CP epithelial cells. Taken together, our results established that organic cations are transported by PMAT from the CSF and highly accumulated in CP. Larger organic anions are removed from the CSF by OATP (e.g. OATP1C1)‐mediated apical uptake followed by MRP1/4‐mediated basolateral efflux into the blood. Our findings revealed the molecular mechanisms of organic cation and anion transport at the blood‐CSF barrier and suggested important roles of CP transporters in maintaining the homeostasis and normal function of the brain.

Support or Funding Information

This study is supported by National Institutes of Health Grant GM066233.

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 α‐Arrestin ARRDC3 Suppresses Breast Carcinoma Invasion by Regulating GPCR Lysosomal Sorting and Signaling
Authors: Arakaki Aleena K. , Pan Wen‐An , Trejo JoAnn
Published: June 2021

G protein‐coupled receptor (GPCR) signaling regulates cancer cell proliferation, invasion, migration and survival at metastatic sites. However, despite the success and promise of GPCRs as therapeutic targets, there are currently no FDA‐approved drugs targeting GPCRs for cancer. Protease‐activated receptor‐1 (PAR1) is a GPCR that promotes breast cancer progression. PAR1 is overexpressed in breast cancer patient tissue biopsies and in breast carcinoma cell lines, and correlates with increased rates of metastasis and poor prognosis and increased invasion and metastatic potential, respectively. One mechanism that leads to PAR1 aberrant over‐expression is defective lysosomal trafficking. The dysregulation of PAR1 trafficking leads to persistent G protein signaling and promotes breast cancer invasion. However, the mechanisms responsible for aberrant PAR1 trafficking remain unknown. Here we show that arrestin domain containing protein‐3 (ARRDC3), an adaptor protein for E3 ubiquitin ligases, functions as a tumor suppressor by controlling PAR1 trafficking in breast carcinoma cells. We used a tetracyclineinducible pSLIK lentiviral vector to restore expression of ARRDC3 in highly invasive, basal‐like MDA‐MB‐231 cells, which exhibit high PAR1 and low ARRDC3 expression. Re‐expression of ARRDC3 restored agonist‐induced PAR1 degradation, attenuated JNK and Hippo‐YAP signaling, and further inhibited PAR1‐mediated breast carcinoma cell invasion. Our studies suggest a crucial role for ARRDC3 in regulating normal PAR1 lysosomal degradation and signaling, with loss of ARRDC3 resulting in aberrant PAR1 signaling, leading to tumor progression in breast cancer. A better understanding of the mechanisms by which PAR1 contributes to breast cancer progression may lead to new potential therapeutic targets for the treatment or prevention of metastatic breast cancer.

Support or Funding Information

Funding for this project was provided by the UCSD Tribal Membership Initiative Graduate Fellowship, UCSD Graduate Training Program in Cellular and Molecular Pharmacology through NIH General Medical Sciences, T32 GM007752 (Aleena Arakaki) and by the National Institute of Health NIGMS R01 GM090689 (JoAnn Trejo).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Acarbose Delays Renal Cancer Progression in Mice: Implications for Obesity‐Induced Immunotherapeutic Failure
Authors: Orlandella Rachael M. , Boi Shannon , Smith Daniel L. , Norian Lyse A.
Published: June 2021

Obesity is a regarded as a major risk factor for developing renal cell carcinoma (RCC). Advanced‐stage RCC exhibits chemotherapeutic resistance, but is responsive to high‐dose IL‐2 and anti‐PD‐1 immunotherapies. Despite successes in these treatments, response rates remain low (20–30%). Currently, pre‐clinical evaluations of immunotherapeutic strategies typically use lean, young mice and do not account for the presence of patient comorbidities. This may partially explain the underwhelming success in clinical translation. Our laboratory previously demonstrated that a T cell‐based immunotherapy clears orthotopic renal tumors in lean mice, but fails in diet‐induced obese mice. This illustrates the importance of identifying methods to rescue obesity‐induced immunotherapeutic failure in this model. Calorie restriction (CR) delays the onset of chronic diseases, such as obesity and cancer. This attribute has prompted interest in investigating CR and its substitutes as potential cancer therapeutics. Recent murine research shows that diets containing calorie restriction mimetics (CRMs) improve anti‐tumor immunity and chemotherapeutic efficacy. Here we examined acarbose, an α‐glucosidase inhibitor with CRM properties, in a preclinical model of kidney cancer. BALB/c mice were fed an acarbose‐supplemented or control diet following orthotopic tumor challenge with syngeneic Renca renal carcinoma cells. Acarbose reduced endpoint tumor weights (p=0.02) and lung metastases (p=0.03). Flow cytometric analysis at endpoint revealed similar frequencies of splenic and intra‐tumoral CD8 T cells in mice treated with acarbose. These findings suggest that acarbose may exhibit anti‐tumor properties, and indicates a potential treatment opportunity for mice with diet induced obesity. Future experiments will focus on evaluating acarbose in combination with immunotherapy, in lean and obese mice.

Support or Funding Information

T32HL105349

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cardiac autonomic neuropathy as a result of mild hyper‐caloric challenge in absence of signs of diabetes: Modulation by anti‐diabetic drugs
Authors: Bakkar Nour Mounira , Mroueh Ali , Ghali Rana , Al‐Assi Ola , Kaplan Abdullah , Mougharbil Nahed , Eid Ali , Zouein Fouad , El‐Yazbi Ahmed
Published: June 2021

Cardiac autonomic neuropathy (CAN) is an early cardiovascular complication of diabetes occurring before signs of metabolic derangement are evident. The cause of CAN remains elusive and cannot be directly linked to hyperglycemia since pre‐diabetic and metabolic syndrome patients demonstrate signs of cardiac autonomic dysfunction. Recent clinical data report cardioprotective effects of some anti‐diabetic drugs independent of their hypoglycemic action. Significantly, pioglitazone and metformin demonstrated protection in pre‐diabetic patients. Here, we used a rat model receiving limited daily increase in calories from fat (HC diet) to assess whether mild metabolic challenge led to CAN in absence of interfering effects of hyperglycemia, glucose intolerance, or obesity. Compared to controls, rats receiving HC diet for 12 weeks showed reduction in baroreceptor sensitivity and heart rate variability despite lack of change in baseline hemodynamic and cardiovascular structural parameters. Impairment of cardiac autonomic control was accompanied with perivascular adipose inflammation observed as an increased interleukin‐1b (IL‐1b) and tumor necrosis factor‐a (TNF‐a) expression, together with increased cardiac oxidative stress, IL‐1b expression and signaling derangements characteristic of diabetic cardiomyopathy including reduction of AMPK phosphorylation and activation of the TGF‐b/Erk1/2 pathway. At this stage, examination of the brainstem revealed no molecular disorders in support of an early, potentially reversible effect. Two‐week treatment with metformin or pioglitazone rectified the autonomic derangement and corrected the molecular changes in heart and adipose tissue. Switching rats on HC diet to normal chow but not to isocaloric amounts of HC for two weeks reversed CAN. As such, we conclude that adipose inflammation due to increased fat intake might underlie development of CAN and, hence, the beneficial effects of metformin and pioglitazone.

Support or Funding Information

Supported by AUB MPP fund #320148.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
VE‐PTP Functions as a Scaffold to Stabilize the AJ and Endothelial Barrier
Authors: Juettner Vanessa , Dan Arkaprava , Leckband Deborah , Komarova Yulia , Malik Asrar
Published: June 2021

Vascular Endothelial Protein Tyrosine Phosphatase (VE‐PTP) is an endothelial‐specific phosphatase that dephosphorylates multiple proteins including Vascular Endothelial (VE)‐cadherin, the main adhesive protein of adherens junctions (AJ). VE‐PTP‐mediated dephosphorylaiton of VE‐cadherin is required for reassembly of VE‐cadherin junctions and restoration of endothelial barrier function after humoral or mechanical stress. The role of VE‐PTP in regulation of VE‐cadherin adhesion complexes under basal conditions, however, remains unknown. VE‐cadherin establishes adhesion events through trans‐interaction with opposing VE‐cadherin molecules at AJs. In resting endothelial monolayers, VE‐cadherin undergoes continues exchange between junctional and cytosolic pools due to VE‐ cadherin recruitment to and internalization from AJs. Using VE‐cadherin tagged with the photoconvertable protein Dendra2, we have demonstrated that VE‐PTP controls stability of VE‐cadherin junctions in a phosphatase‐independent manner. Overexpression of wild‐type or phosphatase “dead” VE‐PTP mutant similarly decreased the VE‐cadherin internalization rate without having any effect on VE‐cadherin recruitment rate. Consistent with these data, depletion of VE‐PTP increased VE‐cadherin internalization rate from AJs as compared to control cells suggesting that VE‐PTP stabilizes AJs in resting endothelial monolayers. Interestingly, treatment of cells with VE‐PTP phosphatase inhibitor had no effect on VE‐cadherin kinetics, further indicating a specific role of VE‐PTP in regulating stability of AJs independent of Aniopoetin‐1/Tie2 signaling. The phosphatase‐independent role of VE‐PTP was further investigated by elucidating VE‐PTP binding partners through mass spectrometry analysis. We found that VE‐PTP binds to GEF‐H1, a RhoA GEF, and inhibits RhoA activity at AJs. Knockdown of GEF‐H1 reduced both RhoA activity at AJs and tension across VE‐cadherin adhesion in VE‐PTP knockdown cells. These events coincided with restoration of VE‐cadherin kinetics at AJs suggesting that VE‐PTP stabilizes VE‐cadherin junctions via inhibition of RhoA and intracellular tension. Our data, for the first time, describe the scaffold function of VE‐PTP at AJs in the mechanism of AJ stabilization.

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 function RGS2 mutations augment vascular contractility ex vivo
Authors: Phan Hoa Thi Nhu , Jackson William , Neubig Richard
Published: June 2021

Objective

Regulator of G protein signaling 2 (RGS2) plays a significant role in alleviating vascular contraction and promoting vascular relaxation due to its GTPase accelerating protein activity toward Gαq. Through a cell‐based Ca2+ mobilization assay, we identified 4 loss‐of‐function (LOF) mutations (Q2L, D40Y, R44H and R188H) out of 16 rare, missense mutations in RGS2 selected from various human exome sequencing projects. This study is to investigate whether these LOF RGS2 mutations disrupt the protein function in regulating vascular contractility.

Approach and result

We confirmed that RGS2‐deficient mice expressed a hypertensive phenotype by telemetric femoral artery blood pressure measurement in conscious mice (WT SBP 120 ± 3 vs RGS2−/−SBP 128 ± 2 mmHg, p<0.01). Isolated mesenteric arteries from RGS2−/− mice showed higher contractile response to 10 nM angiotensin II (AngII) stimulation in pressure myograph while isolated aortic rings from those animals generated more tension upon phenylephrine (PE) stimulation (Emax 238 mg vs 68 mg, p = 0.01) compared to aortic rings from RGS2+/+ mice. Reintroduction of wild‐type RGS2‐GFP plasmids into RGS2−/− mesenteric arteries by reversible permeabilization suppressed the vasoconstrictor response to 50 nM AngII (RGS2 WT plasmid −4% vs pcDNA control −35% of basal diameter, p = 0.01). Transfection with RGS2 LOF mutation plasmidss did not suppress Ang II constriction compared to those expressing WT‐RGS2 (Q2L −20%, D40Y, R44H and R188H −30% of basal diameter, p < 0.05).

Conclusions

This study is the first to investigate the function of human mutant RGS2 in resistance arteries in a near physiological condition. It demonstrates that RGS2 attenuates vasoconstriction in the mesenteric arteries and that RGS2 mutations disrupt this effect. Although these mutations are rare, knowledge about their function may provide insights into pathophysiology and treatment.

Support or Funding Information

This work was supported by the American Heart Association predoctoral fellowship [15PRE24680004] to H.P).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Enhanced Arterial Medial Calcification in Mice with Smooth Muscle‐Specific Deletion of Lysosomal Acid Ceramidase.
Authors: Bhat Owais M. , Yuan Xinxu , Zhang Yang , Boini Krishna , Lohner Hannah , Li Pin‐Lan
Published: June 2021

Arterial medial calcification (AMC) is associated with transition of arterial smooth muscle cells (SMCs) to osteogenic phenotype. These arterial SMCs secrete matrix vesicles or exosomes to form the first nidus for mineralization. However, the mechanisms mediating exosome excretion in SMCs remain poorly understood. Whether SMC‐specific lysosomal Asah1 gene (acid ceramidase (AC) gene code in mice) expression and associated lysosomal ceramide signaling are required for osteogenic phenotype change and exosome excretion are unknown. In the present study, we used Asah1fl/fl/SMcre mice with SMC‐specific deletion of lysosomal AC and Smpd1trg/SMcre with SMC‐specific overexpression of acid sphingomyelinase (ASM) gene (mouse gene code: Smpd1) to demonstrate the role of lysosomal ceramide signaling pathway in AMC. Using high dose of vitamin D (500,000 IU/kg/day) to induce AMC, Asah1fl/fl/SMcre mice were found to have increased expression of RUNX2 and osteopontin, but downregulation of SM22‐α (SMC marker) with decreased ratio of α‐SMA vs. vimentin in coronary and aortic media compared to their littermates (Asah1fl/fl/WT and WT/WT mice). Asah1fl/fl/SMcre mice also had augmented AnX2 and ALP expression (exosome markers) and reduced co‐localization of lysosome marker (LAMP‐1) with multivesicular body marker (VPS16) in both coronary and aortic media. Morphological examinations confirmed that severe AMC occurred in both aorta and coronary arteries in Asah1fl/fl/SMcre, but not in their littermates. In Smpd1trg/SMcre mice, similar biochemical and morphological changes were found to Asah1fl/fl/SMcre mice. These data indicate that lysosomal AC in SMC plays a critical role in osteogenic phenotype change, exosome release and mineral deposition in the mouse model of AMC and that lysosomal ceramide signaling pathway may control exosome excretion and osteogenic phenotype transition in SMCs that determine the development of AMC.

Support or Funding Information

(Supported by NIH grants HL057244, HL075316 and HL122937).

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 Colonic Inflammation on Morphine Induced Antinociceptive Tolerance
Authors: Komla Essie , Dewey William L. , Akbarali Hamid I.
Published: June 2021

Opioids are one of the most widely prescribed classes of drugs for the treatment of moderate to severe pain. Opioids are prescribed for abdominal pain, a major symptom associated with Crohn's Disease. Recent studies have reported that continued opioid use in patients with Crohn's Disease is associated with increased mortality rate, duration and severity of disease, as well as infection. However, it is not clear if this association is causal. We have previously demonstrated that chronic use of morphine, in addition to the development of antinociceptive tolerance, also causes the disruption of the intestinal barrier function resulting in bacterial translocation from the gut lumen into the peritoneal organs, thus inducing an inflammatory response in the colon. Therefore, we tested if inflammation in the colon, as in Crohn's disease, affects the rate of antinociceptive tolerance development to morphine. To test this hypothesis, we induced colonic inflammation in Swiss Webster male mice using 2,4,6‐trinitro‐benzene sulfonic acid (TNBS) or vehicle intrarectally and then surgically implanted 25 mg, 50 mg (2 × 25 mg), or 75 mg slow release morphine pellets or placebo pellets subcutaneously on the dorsum. Tail immersion assay was performed on day 3, 4, and 5 post treatment and colons were extracted from the animals to measure mRNA expression of Interleukin 1 beta (IL‐1β) and tumor necrosis factor alpha (TNF‐α) by real‐time polymerase chain reaction (RT‐PCR). A morphine (10 mg/kg) challenge was used to test for the development of antinociceptive tolerance at various times in morphine pelleted with (n= 7) or without TNBS (n= 5). In TNBS + morphine pelleted mice, tolerance developed by day 5 in 25 mg morphine pelleted mice, by day 4 in 50 mg (2 × 25 mg) morphine pelleted mice, and as early as day 2 in 75 mg morphine pelleted mice. In the absence of TNBS‐induced inflammation, tolerance to morphine did not develop to either 25 mg or 50 mg morphine pellets up to day 5, and at day 4 in 75 mg morphine pellets. Furthermore, IL‐1β production was significantly increased in the morphine and placebo pelleted treated animals with TNBS from day 1 compared to the vehicle treated groups. In summary, chronic morphine administration in the presence of colonic inflammation leads to an increased rate and extent of antinociceptive tolerance development.

Support or Funding Information

Agency: NIDA Agency Number: 5R01DA036975‐04

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
SML0417 from Saussurea lappa ameliorates asthma features in murine allergic asthma model
Authors: Nam So‐Yeon , Lee Bo‐Kyung , Kang Saeromi , Park Soo‐Jin , Im Dong‐Soon
Published: June 2021

Background

The root of Saussurea lappa has traditionally been used as an ingredient in the treatment of asthma, rheumatism, abdominal pain, anorexia, nausea, and other conditions. Extracts of Saussurea lappa were reported to alleviate house dust mite‐induced atopic‐like dermatitis in Nc/Nga mice, and sesquiterpene lactones were isolated.

METHODS

To elucidate whether SML0417, a sesquiterpene lactone has beneficial effects on allergic asthma, anti‐asthma effects were studied using female Balb/c mice and rat RBL‐2H3 mast cells. Antigen‐induced degranulation was measured in vitro by measuring β‐hexosaminidase activity. A murine ovalbumin‐induced allergic asthma model was used to test the in vivo efficacy of SML0417.

RESULTS

Antigen‐induced degranulation was inhibited by SML0417. Sensitization and challenge of ovalbumin induced allergic asthma responses. Histologic studies showed administration of SML0417 reduced inflammatory signs and mucin production in lungs. Administration of SML0417 decreased numbers of inflammatory cells, especially eosinophils, and reduced the expression and secretion of Th2 cytokines, including IL‐4 and IL‐13, in lung tissues and bronchoalveolar lavage fluid.

CONCLUSION

The present findings provide evidence that SML0417 has a potential therapeutic use in allergic asthma.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Transport and Metabolism of Adenosine and its Metabolites in Capillary Endothelial Cells of Guinea Pig Hearts
Authors: Liu Aiping , Bassingthwaighte James B.
Published: June 2021

Adenosine (Ado) plays an important role in cardioprotection during ischemia by enhancing coronary flow through binding to purinoceptors on smooth muscle cells (SMC), and by preserving nucleotide pools through purine salvage in cardiomyocytes (MC). However, two questions impede our understanding of the mechanism by which Ado acts on MCs or SMCs: (1) how much Ado reaches cardiomyoctyes or SMC when EC avidly take‐up Ado from blood stream and interstitium? and (2) how much Ado degrades to uric acid (UA), or is salvaged back into cellular pools, mainly ATP? Our goal here is to quantify the fluxes of Ado and its metabolites in ECs. The multiple‐indicator‐dilution technique (MID) was used to measure the capillary transport and metabolism of Ado and metabolites in isolated, Krebs‐Ringer perfusated guinea pig hearts. Radiolabeled [14C]‐ adenosine, or inosine, or hypoxanthine, or xanthine was injected with [131I]‐albumin (intravascular reference solute) and [3H] AraH or L‐[3H] glucose (extracellular references) as a bolus into the coronary flow; venous outflow dilution curves of the injectate and its metabolites were quantified with HPLC and LS counting. The permeabilities of each solute (via interendothelial clefts, PSg, trans‐endothelial membrane, PSecl, trans‐cardiomyocytes PSpc) and the metabolism in ECs and MCs were estimated by fitting the MID curves with a general convection‐diffusion‐exchange‐reaction model, Gentex. Results are: Intracellular sequestration of injected Ado is high compared to its metabolites (~60% of Ado vs. ~50% of Ino). Similarly, PSecl is higher for Ado than metabolites, INo>Hx>Xa> UA (Ado:1.9±0.7 vs. Ino:1.7±0.5 vs. Hx: 0.5±0.1 ml·min−1·g−1). The low permeabilities help to retain purine in the cells. Plasma injectate Ado that enters ECs is estimated to be ~50% oxidized to UA, ~50% retained; Hx is ~67% oxidized, ~23% retained, both presumably incorporated into nucleotide pools. Thus ECs actively participate in purine salvage via both Ado and Hx. Uncertainty for EC parameters is much smaller than for MC. Having well‐defined parameters for permeabilities and metabolism for Ado and its metabolites in ECs is an essential step toward understanding the fate of endogenous Ado generated from cardiomyocytes during ischemia and reperfusion.

Support or Funding Information

Our study was supported by NIH research grant U01HL122199.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
RESVERATROL IMPROVES MICROVASCULAR FUNCTION IN ADULTS WHO REPORTED ADVERSE CHILDHOOD EVENTS
Authors: Miguelez Paula Rodriguez , Looney Jacob , Thomas Jeffrey , Pollock Jennifer , Harris Ryan
Published: June 2021

INTRODUCTION

Adversity early in life, referred to as adverse childhood events (ACEs), has been associated with the development of cardiovascular disease (CVDs) during adulthood. Microvascular dysfunction plays a pivotal role in the etiology of CVDs through a reduction in nitric oxide (NO)‐dependent vasodilation. Lower concentrations of sirtuin 1 (Sirt1), an NAD‐dependent protein that increases NO metabolism, have been described in multiple CVDs.

PURPOSE

This study sought to test the hypothesis that an acute dose of resveratrol (RES), a natural activator of Sirt1, will improve MVF in adults who reported ACEs.

METHODS

Ten adults (34 ± 3 yrs), with no evidence of overt CVD, participated in the study. Based on the ACE questionnaire, all participants had self‐reported an ACE score of 4 or greater. Two protocols were used to evaluate MVF using Laser Doppler flowmetry: 1) local thermal hyperemia (LTH), with a maximal plateau phase primarily mediated by NO, and 2) post‐occlusive reactive hyperemia (PORH), partly dependent on NO metabolism. MVF assessments were determined before and 90 min after an acute dose of either RES (500 mg) or Placebo (P), separated by at least 7 days and are expressed as cutaneous vascular conductance (CVC; cutaneous flux/mean arterial blood pressure). Maximal dilation capacity was evaluated through LTH. Skin resistance was also calculated for every individual and every treatment using Ohm's law and considered in all the statistical analysis.

RESULTS

No differences in resting CVC were observed between days (0.11 ± 0.08 vs. 0.11 ± 0.03 PU/mm Hg; p=0.297) or with the change from either treatment (ΔRES: −0.03 ± 0.03 vs. ΔP: −0.01 ± 0.01 PU/mm Hg; p=0.653). In addition, resveratrol significantly (p=0.005) increased max LTH when compared with P (ΔRES: 0.12 ± 0.24 vs. ΔP: −0.06 ± 0.26, PU/mm Hg). Moreover, RES significantly (p≤0.001) prevented the decrease in PORH relative to the maximal dilation capacity when compared with placebo (ΔRES: −6.4 ± 1.4 vs. ΔP: −18.1 ± 17.8 %CVC max). In addition, the acute dose of RES increased concentrations of Sirt1 (RES: 1.3 ± 0.1 vs. ΔP: −0.4 ± 0.3 ng/ml; p=0.075) which was significantly associated with the max LTH response (r=0.760; p=0.011).

CONCLUSION

These findings document an improvement in MVF after an acute oral dose of resveratrol in adults who have reported ACEs. Future studies are warranted to investigate the long term effects of resveratrol on MVF in adults who have reported ACEs.

Support or Funding Information

Supported in part by NIH/NHLBI P01HL069999 (JSP, RAH) and 16POST31080031 (PRM).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cerebrovascular Reactivity in Obstructive Sleep Apnea: Impact of Physical Activity
Authors: Sauder Christina J. , Carter Katrina J. , Ward Aaron T. , Morgan Barbara J. , Hagen Erika W. , Peppard Paul , Schrage William G.
Published: June 2021

Background

Obstructive sleep apnea (OSA) is associated with increased risk of cerebrovascular disease and stroke. This risk may be partially due to reduced cerebrovascular function. Conversely, regular physical activity (PA) reduces the risk for cerebrovascular disease. Whether PA levels can predict the OSA‐related reductions in cerebrovascular function remains unknown. Therefore, we tested the hypothesis that the level of PA alters the relationship between OSA‐severity and cerebrovascular dysfunction.

Methods

245 healthy adults (113 females; mean age ~60 years; smoking, continuous positive airway pressure (CPAP) treatment, and diabetes were exclusionary) performed overnight polysomnography to determine the presence and severity of OSA. Next, cerebrovascular function was tested in the middle cerebral artery (MCA) in response to CO2 rebreathing (10 mmHg increase), expressed as cerebrovascular reactivity (CVR; cm/sec/mmHg PETCO2). PA was estimated using the Paffenbarger Physical Activity Questionnaire and expressed as Metabolic Equivalent (MET) hours per week. Data were analyzed by linear regression analysis using CVR as the outcome variable and mean nighttime arterial oxygen saturation (SaO2) as the main independent variable. Model adjustments included age, sex, number of metabolic syndrome factors, number of caffeinated beverages per day, and mean arterial pressure (MAP) slope response to CO2. We also included a model that had an additional adjustment for PA.

Results

In an unadjusted model, higher nighttime SaO2 was a predictor of higher CVR (p < 0.05). When adjusted with the aforementioned variables, both SaO2 and MAP slope were predictors of CVR. Adding PA to the adjusted model did not change the overall relationship between SaO2 and CVR.

Conclusions

These data fail to demonstrate that PA level has a substantial impact on CVR in subjects with OSA.

Support or Funding Information

R01HL062252, R01HL075035, and UL1RR025011

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 role of adiponectin in exercise training‐induced vascular adaptation
Authors: Sepulveda Jaime Luis , Ghosh Payal , Delp Michael , Pritchard Emily , Maraj Joshua , Wahl Jake , Restrepo Alexis , Morgan Hannah , Ulrich Brody , Abeln Clayton , Nolan Jacob , Kenney Mary , Patterson Nicolas , Rosenthal Shaianne , Vega‐Figueroa Leilanie , Sadowsky Dylan , McKinley‐Caspanello Carly , Schwartz Kaley , Muller‐Delp Judy
Published: June 2021

We have previously reported that in old rats, reversal of age‐related vascular dysfunction by exercise training correlates with an increase in circulating adiponectin and its signaling within coronary vascular smooth muscle. In the current study, we investigated the effect of deletion of adiponectin on exercise training‐induced vascular adiponectin. C57BL/6 wild‐type (WT) or homozygous adiponectin knockout (AdipoKO) mice were obtained at 10–12 wks of age and underwent treadmill exercise training (EX) (12 m/min, 5° incline, 1 hr/day, 5 days/wk for 8 wks) or remained sedentary (SED) in cages. Arterioles isolated from cardiac and soleus muscle were assessed for contractile and vasodilatory function, and capillarity of the soleus muscle was evaluated. Exercise training increased flow‐induced dilation significantly in coronary arterioles of WT mice (P<0.01 EX vs. SED), but decreased flow‐induced dilation in coronary arterioles from AdipoKO mice. ACh‐induced dilation was reduced in coronary arterioles from AdipoKO mice as compared to those from WT mice (P<0.05 AdipoKO vs. WT). Exercise training reduced ACh‐induced dilation in arterioles from WT mice, but increased ACh‐induced dilation in arterioles from AdipoKO mice. Baseline capillarity increased in the soleus muscle of AdipoKO mice as compared to WT mice, but an exercise training‐induced increase in capillarity, detected in WT mice, was absent in AdipoKO mice. Contractile responsiveness to phenylephrine was increased in arterioles from the soleus muscle of both WT and AdipoKO mice (P<0.01 EX vs. SED in both WT and AdipoKO). These data indicate that adiponectin is a critical contributor to exercise training‐induced vascular adaptations; however, locally produced adiponectin may be more critical than circulating adiponectin in mediating these adaptations.

Support or Funding Information

Florida State University College of Medicine

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Early Exercise Training Postpones Hypertension Progression via Activation of Central ACE2‐Ang(1–7)‐Mas Axis in Prehypertensive Rats
Authors: Pan YanXia , Peng WenWen , Zhao XiaoLin , Gao Lie , Lin Cheng , Cheng XiuYun
Published: June 2021

Objectives

To investigate the effects of exercise training on the development of hypertension in spontaneously hypertensive rats (SHR) and explore the underlying mechanisms by focusing on the central ACE2‐Ang (1–7)‐MAS axis.

Methods

The male SHR (n=20, 5 weeks old) and 20 normotensive Wistar Kyoto rats (WKY) were randomly assigned to sedentary (Sed) group or exercise training (ExT) group. The trained rats run on a treadmill in 20 m/min, 30 min/day, 5 day/week for 20 weeks. Systolic blood pressure (SBP) were measured by a tail‐cuff method. The baroreflex sensitivity (BRS) was assessed by intravenous injection of phenylephrine under anesthesia. The expression of ACE2 and MAS receptors mRNA and protein in the cardiovascular centers was determined by real‐time PCR and Western blot. Alterations in the BRS were evaluated before and after intracerebroventricular injection of MAS receptor agonist Ang (1–7) and its antagonist A779, respectively.

Results

(1) Exercise training significantly postponed the development of hypertension in SHR and decreased SBP in both SHR and WKY as compared with Sed groups (P<0.01 and P<0.05). (2) Exercise training enhanced blood pressure regulation and improved the BRS in the SHR (P<0.01). (3) The expression of ACE2 and MAS receptor mRNA and protein in rostral ventrolateral medulla (RVLM), nucleus tract solitarius (NTS), and paraventricular nucleus (PVN) was up‐regulated in SHR+ExT group (P<0.05, P<0.01). (4) Central administration of A779 abolished the benefits of exercise‐induced improvement of BRS in the SHR+ExT rats (P<0.01). In contrast, Ang (1–7) improved the BRS in both SHR+Sed group and SHR +ExT group (P<0.05).

Conclusion

Exercise training postpones hypertension progression and improves blood pressure regulation, which is associated with the enhancement of central ACE2‐Ang(1–7)‐Mas axis.

Support or Funding Information

National Natural Science Foundation of China (No. 81372111) and Natural Science Foundation of Fujian Province of China (No. 2014J01339)

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Deciphering the Neural Signature of Human Blood Pressure Control
Authors: Manuel Jorge , Nazarenko Nataliia , Heusser Karsten , Tank Jens , Jordan Jens , Beissner Florian
Published: June 2021

The aim of this study was to measure the activity of hypothalamic and brainstem centers regulating blood pressure in humans

The human body has several systems to regulate blood pressure, one of which is the baroreflex. It is mediated by a number of nuclei in the brainstem and hypothalamus that via autonomic efferents adjust the mean arterial pressure by altering both the force and speed of the heart's contractions, as well as systemic vascular resistance. While cortical centers influencing the baroreflex have been studied in humans using functional magnetic resonance imaging (fMRI), the brainstem region suffers from strong physiological noise that makes detection more difficult. Here, we combined high‐resolution fMRI with lower body negative pressure (LBNP) and concomitant autonomic recordings hypothesizing that such an approach would make it possible to detect hypothalamic and brainstem nuclei controlling the baroreflex in humans.

15 healthy subjects were scanned using a 3T MR scanner. The protocol involved SMS‐EPI functional scans (voxel size=2×2×2 mm3) as well as T1‐weighted structural scans (voxel size=1*1*1.2 mm3). LBNP stimulation was delivered using a custom‐made MR‐compatible pressure chamber and a vacuum cleaner that was controlled by a digital pressure gauge. FMRI data were minimally preprocessed using tools from FMRIB Software Library (FSL v5.0) including motion correction, unwarping, temporal high‐pass filtering and normalization to a study template. The data were masked to retain only the brainstem and hypothalamus, excluding the adjacent areas with high physiological noise. Statistical analysis was conducted using masked independent component analysis (mICA), spectral analysis, and network modelling.

We found activations related to the LBNP paradigm in multiple nuclei known to be involved in baroreflex regulation. These included the nucleus of the solitary tract and the caudal ventrolateral medulla in the lower brainstem as well as the paraventricular hypothalamic nucleus and lateral hypothalamus. We further observed significant activity changes on the lower ventral medullary surface. Frequency analysis revealed that the BOLD signal in the rostroventrolateral medulla showed spectral changes in the Mayer band (0.1 ± 0.035 Hz) during LBNP.

Being able to measure baroreflex nuclei in vivo is an important step towards the understanding of this system in humans. Our results show the importance of selected nuclei in the hypothalamus and brainstem.

Support or Funding Information

Funding: Horst Goertz Foundation

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
MRI Characterization of Age‐Associated Changes to the Cerebral Arteries in a Rodent Model of Aging and Hypertension
Authors: McPherson Rebecca L. , Naraine Akshay , Lee Diana , Rejimon Abinand , Fishbein Kenneth W. , Spencer Richard G. , Lakatta Edward G. , Fedorova Olga V.
Published: June 2021

Cerebral blood vessel remodeling occurs in response to aging and hypertension. Of importance are structural changes that occur to the circle of Willis and the impact of these changes on blood flow within the brain, especially in regions associated with learning and memory, such as the hippocampus. In the present study, we apply MRI to a rodent model of aging and hypertension to elucidate changes in the circle of Willis during aging and age‐associated hypertension.

Hypothesis

We hypothesized that arteries comprising the circle of Willis in hypertensive Dahl‐salt sensitive rats (Dahl‐S) will exhibit age‐related volume changes, these changes will impact metabolite levels in the hippocampus and the effects will differ between Dahl‐S and age‐matched Sprague‐Dawley rats (SD).

Methods

Whole brain time‐of‐flight (TOF) 3D MRI and MRS (spectroscopy) of the hippocampus were acquired using a 7 Tesla 30 cm Bruker Biospec scanner in SD and Dahl‐S male rats on normal 0.5% NaCl intake at 3, 6 and 12‐mo of age (n=5/group). Cerebral arteries comprising the circle of Willis (cerebral arterial circle (CAC), caudal communicating artery (CCA) and rostral communicating artery (RCA)) were manually segmented using AMIRA. MRS data were processed using LCmodel. Systolic blood pressure (SBP) was measured via the tail‐cuff method at each time point before MRI scans. Data were analyzed using 2‐way ANOVA followed by a Tukey's posthoc test, and are presented as mean ± standard deviation

Results

At 3‐mo, SBP did not differ between Dahl‐S vs SD (134.6±14.1 vs 131.4±8.8 mmHg). However, Dahl‐S had higher SBP vs SD at 6‐mo (165.8±20.6 vs 123.7±8.8 mmHg) and 12‐mo (166.1±16.9 vs 142.2±12.8 mmHg). The CAC volume was greater in SD vs Dahl‐S at 3 and 12‐mo but did not differ at 6‐mo (Table). Importantly, the CAC was not visible by MRI in Dahl‐S at 12‐mo of age, except in one animal. The CCA volume was greater in SD vs Dahl‐S at 12‐mo and did not differ at 3 or 6‐mo. The RCA volume did not differ between strains at any time point. No arterial volume differed with age in SD. In Dahl‐S, the CCA and RCA volumes were larger at 6‐mo compared to 3 and 12‐mo (Table). Total N‐acetylaspartate (NAA) in the hippocampus was lowest at 6‐mo in both strains, compared to 3 and 12‐mo, but did not differ between strains at any time point (Table).

Conclusion

We observed variations with strain and age in the volume of arteries in the circle of Willis in normotensive SD and hypertensive Dahl‐S. Hypertensive Dahl‐S at 6‐mo of age may experience increased metabolic demand and blood volume requirements in the brain.

Support or Funding Information

This work was supported solely by the NIH/NIA Intramural Research Program.

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 Alters MicroRNA Signature Profile in Diabetic Rabbit following Myocardial Ischemia Reperfusion Injury: A Preclinical Approach for Cardioprotection.
Authors: Samidurai Arun , Ockaili Ramzi , Roh Sean K. , Kukreja Rakesh C. , Das Anindita
Published: June 2021

Background

Myocardial ischemia reperfusion (I/R) injury associated with cardiovascular risk factors and comorbidities, including Diabetes (DM), is a major complication for development of heart failure. Mammalian target of Rapamycin (mTOR) is robustly active in DM and further exacerbated during I/R. mTOR inhibition with rapamycin at reperfusion preserves cardiac function with reduction of myocardial infarction in diabetic mice. Recent advances in non‐coding RNA analysis demonstrates microRNA (miR) plays an important role in cardiovascular diseases. We examined the effect of rapamycin treatment at the onset of reperfusion in a conscious diabetic rabbit model of I/R and identified differentially regulated miRs.

Methods and Results

To induce diabetes, alloxan monohydrate (125 mg/kg, i.v.) was administered in New Zealand male rabbits (n=26; average 3 kg, 4 month old). Blood glucose levels increased to 345±66 mg/dL (n=10) following 2 weeks of alloxan administration. The animals were categorized to three groups: Diabetes (DM), Diabetes subjected to I/R (DM+I/R) and Diabetes underwent I/R and treated with Rapamycin (DM+I/R+Rapa). Post 7 days of implantation of balloon occluders on the top of the coronary artery, the conscious rabbits were subjected to 45‐min ischemia and 3 days of reperfusion by inflating/deflating the balloon. Rapamycin (0.25 mg/kg, i.v.) or DMSO (vehicle) was infused 5 min before reperfusion. Plasma was collected at baseline, 1, 2, 4 and 24 hours during the reperfusion and cardiac Troponin I (cTnI) release was measured by ELISA. The results show that rapamycin significantly reduced myocardial infarction in DM rabbits following I/R (Figure A) as assessed by triphenyltetrazolium chloride staining. Rapamycin also reduced cTnI at 1 hr and 2 hr of reperfusion as compared to vehicle alone (Figure B). RNA isolated from risk area of left ventricle heart tissue from different groups (n=3) was subjected to miRNA‐Array analysis using μParaflo® Microfluidic Biochip Technology (Figure C). The results show 34 out of 761 miRs were differentially regulated with Rapamycin treatment (p<0.01) (Figure D). Enrichment Analysis and Annotation of selected miRs were performed using miEAA and DAVID tools to identify potential targets and its interaction with mTOR signaling pathway.

Conclusion

This Preclinical translational study demonstrates that rapamycin could be a promising drug in attenuation of reperfusion injury in diabetic patients following acute MI by altering novel miRs.

Support or Funding Information

NIH RO1HL134366 (AD & RCK) & CCTR Endowment Fund UL1RR031990 (AD)

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