bluuebunny/crossref_metadata_2025 huggingface dataset:
Structure‐Based Assessment of the Toxicity and Cytoprotective Effects of a Series of Small Molecule Polyphenolic Compounds
Authors: Lotomau Storm , Yavuz Ege , True Heather , Stamper Brendan
Published: June 2021

Objective

Polyphenols are widely known to possess antioxidant and anti‐inflammatory properties, but are also capable of eliciting toxicity at higher doses. This goal of this project was to investigate whether a structure‐toxicity relationship exists among five polyphenolic compounds: trans‐cinnamic acid (tCA), p‐coumaric acid (COU), caffeic acid (CAF), resveratrol (RES), and phloretin (PHL). Each compound is relatively similar in structure, but varies in the number of phenolic groups it contains (tCA=0, COU=1, CAF=2, RES=3, PHL=4).

Methods

Polyphenol toxicity was measured by determining the median lethal dose (LC50) and toxicant threshold concentration (LC1) for each compound using an in vitro mouse liver model (TAMH; transforming growth factor‐α transgenic mouse hepatocytes). TAMH cells were cultured and exposed to varying concentrations of each compound for 24‐hours followed by a test for cell viability using resazurin dye. Subsequent hepatoprotective studies based on LC1 values were then conducted by incubating TAMH cells with each polyphenol for 24 h prior to a hepatotoxic dose of diquat (18μM). Changes in cell viability were again measured using the resazurin‐based assay. Data analysis was performed using SPSS to calculate the LC50 and LC1 values for each compound. Significance was determined using one‐way analysis of variance (ANOVA) and post‐hoc Least Significant Difference tests with a fold cut‐off of p < 0.05.

Results

tCA and COU were relatively non‐toxic (LC50 > 5mM) compared to the compounds with multiple phenolic groups. In fact, addition of phenolic groups directly correlated with a decrease in the LC50 value indicating that as the number of phenolic groups increased so did the toxicity for this series of compounds. For the hepatoprotection studies, significant increases in cell viability were observed across only some of the polyphenolic compounds; however, this protection did not correlate with phenolic group number.

Conclusions

While greater toxicity was associated with an increase in the number of phenolic groups based on the LC50 determinations, no common trend was observed relating phenolic group number and hepatoprotective ability using an LC1‐based dosing regimen. Perhaps this suggests that subtle differences in the carbon skeleton among the polyphenols tested influences protective ability. Future work is currently underway to investigate the mechanistic features differentiating the degree of cytoprotection between the polyphenolic compounds.

Support or Funding Information

Source of Research Support: This work was supported by the Pacific University Research Incentive Grant Program and an ASPET Summer Undergraduate Research Fellow (SURF) Individual Award.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Amelioration of trichloroethene‐mediated autophagy, inflammasome activation, and hepatic immune dysregulation by N‐acetylcysteine
Authors: Wang Hui , Wang Gangduo , Liang Yuejin , Du Xiaotang , Sun Jiaren , Boor Paul J. , Khan M. Firoze
Published: June 2021

Trichloroethene (TCE) exposure can cause induction of autoimmune diseases (i.e., autoimmune hepatitis and systemic lupus erythematosus) potentially through increased oxidative stress (OS). Activation of autophagy has emerged as a pivotal pathogenic factor in various autoimmune diseases. However, role of OS in the regulation of autophagy leading to autoimmune diseases is largely unknown. Here, we tested the hypothesis that OS‐induced activation of autophagy and inflammasome contribute to induction and pathogenesis of TCE‐mediated autoimmune diseases. Female MRL +/+ mice were treated with TCE along with or without antioxidant N‐acetylcysteine (NAC) for 6 weeks (TCE, 10 mmol/kg, i.p., every 4th day; NAC, 250 mg/kg/day via drinking water). TCE‐treated mice had elevated anti‐nuclear antibodies (ANA) and anti‐4‐hydroxynonenal (HNE)‐specific circulating immune complexes, suggesting the association of TCE‐induced OS with autoimmune response. Moreover, TCE exposure led to hepatic autophagy activation as evident from increased beclin‐1 and autophagic light chain 3 conversation (LC3‐II/LC3‐I). The TCE‐mediated autophagy activation was associated with activation of hepatic inflammasome (NLRP3 and caspase‐1) and up‐regulated pro‐inflammatory cytokine IL‐1β. TCE exposure led to hepatic morphological changes, including sinusoid dilation, increased sinusoidal cellularity and proliferating cell nuclear antigen (PCNA) intensity. In addition, dysregulation of hepatic immune response was also observed as evident from markedly increased hepatic lymphocyte infiltration and imbalance between Tregs (decreased) and Th17 cells (increased). Remarkably, TCE‐mediated increased autoantibody formation, hepatic activation of autophagy and inflammasome, and immune dysregulation were effectively attenuated by NAC. Taken together, these results suggest that TCE mediated the hepatic immune dysregulation via OS and activating autophagy and inflammasome. Amelioration of these responses by NAC further supports the role of OS in TCE‐mediated autoimmunity. These novel findings could help in designing therapeutic strategies for such autoimmune diseases.

Support or Funding Information

Supported by NIH R01 grants ES026887 and ES016302.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Impacts on Hepatic Ontogenic Expression of Drug‐metabolizing Enzymes by High‐fat Diet‐induced Obesity in Mice
Authors: Wang Pei , Shao Xueyan , Bao Yifan , Chen Liming , Zhang Lirong , Zhong Xiaobo
Published: June 2021

The prevalence of obesity‐associated conditions increases up to 30% of general populations globally. A few studies have shown that obesity may lead to altered drug metabolism. Although changed expression of drug‐metabolizing enzymes (DMEs) is reported, the impacts on DMEs due to different obese levels (overweight, obese, and morbidly obese) have not been elucidated. Especially, limited information is available on how obesity alters the expression of DMEs in younger ages. The aim of this study is to investigate impacts of high‐fat diet‐induced obesity (DIO) on the hepatic expression of DMEs in adult and the ontogenic expression in offspring with a focus on cytochrome P450s (CYPs) in a mouse model. The expression of CYPs, as well as constitutive androstane receptor (CAR), pregnane X‐receptor (PXR), hepatocyte nuclear factor (HNF4α), and peroxisome proliferator‐activated receptor α (PPARα) was examined. Adult mice consumed 60 kcal% fat diet (HFD) for 8 or 18 weeks showed a more rapid increase of body weight than 10 kcal% fat diet (LFD) control group. The body weight gain of males was >1.5 fold greater than females in HFD groups. Consistent with the weight gain, fat accumulation was found in the livers of HFD groups. Elevated expression of most selected DMEs and transcription factors was observed in HFD groups, expect for CYP3A16 in 8‐week HFD groups, and CYP1A2 and HNF4α in 18‐week HFD groups. Day 60 offspring derived from HFD‐fed adults showed heavier body weights compared with offspring from LFD‐fed adults. Altered ontogenic expression of CYP1A1, 1A2, 2E1, and UGT1A1 was also observed in HFD offspring. Notably, the changed ontogenic patterns were accompanied by the increased expression of DMEs after day 20 in HFD offspring related to LFD groups. Moreover, similar effects on CAR, PXR, HNF4α, and PPARα were found in HFD offspring, indicating that altered expression pattern of DMEs may due to changed expression of transcription factors. Taken together, our study suggested that obese level of DIO influenced the expression of DMEs in adults and offspring and transcription factors might play roles in the altered expression of DMEs.

Support or Funding Information

This work was supported by grant of NIH R01GM118367 and China Scholarship Council (201707040007).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Development of an inducible CRISPR/Cas9 system for temporally‐modulated gene editing
Authors: Olivas Micah , Fox Doug , Alvarez James
Published: June 2021

Despite its prolific track record of research into primary tumor dynamics, the field of cancer biology has yet to develop a reliable methodology for interrogating the genetic profiles of recurrent tumors. CRISPR/Cas9 gene editing has proven to be foundational for recent advancements in oncogenetic screening, due in part to its high degree of specificity in achieving on‐target gene knockout while minimizing off‐target effects. The primary limitation to its application in screening pro‐recurrence factors in cancer, however, is the constitutive nature of canonical Cas9 activity, which may not be remotely regulated after the point of Cas9 translation. To address this problem, we cultivated and transduced populations of a resected invasive mammary adenocarcinoma cell type with a selection of four recombinant Cas9 transgenes and characterized the relative inducibility of each system. These Cas9's are fused to peptides whose tertiary structure renders the endonuclease inactive, restoring catalytic activity only in the presence of a fusion‐specific ligand. Populations of EGFP‐expressing adenocarcinoma cells were transduced with a modified estrogen receptor‐Cas9 fusion (ERT2‐Cas9) and assayed for target knockout efficiency upon addition of small guide GFP (sgGFP) and a Cas9‐enabling estrogen analog. After three days exposure to the analogue, flow cytometry revealed GFP expression to be markedly reduced in cells expressing ERT2‐Cas9 and minimal change in those not exposed. Once the timing and treatment of this model are refined, ERT2‐Cas9‐expressing adenocarcinoma cells will be orthotopically‐injected into mice for an in vivo screening assay. By equipping neoplastic cells with the inducible‐Cas9 machinery before the point of tumor outgrowth, it becomes possible to systematically screen pro‐survival factors at late time points in cancer progression without drastically disturbing the microenvironment of the tumor.

Support or Funding Information

This work was supported by ah ASPET summer research award in partnership with SURPH at Duke University.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Effect of Cystathionine‐Gamma‐Lyase/Hydrogen Sulfide System Modulation on Vascular Dysfunction Induced by Insulin Resistance in Male Wistar Rat Thoracic Aorta
Authors: Silva‐Velasco Diana Laura , Hong Enrique , Beltran‐Ornelas Jesus Hernan , Sánchez‐López Araceli , Gomez Carolina Belen , de la Cruz Saul Huerta , Centurion David
Published: June 2021

Hydrogen sulfide (H2S) is a novel gasotransmitter synthetized from L‐Cysteine by three enzymatic pathways. In blood vessels, cystathionine‐gamma‐lyase (CSE) is mainly expressed and regulates vascular function. This enzyme may be affected by insulin resistance. Insulin resistance leads to vascular dysfunction and is the major mechanism underlying type 2 Diabetes Mellitus. The aim of this study was to determine the effect of chronic administration of sodium hydrosulfide (NaHS; inorganic H2S donor), L‐Cysteine (L‐Cys; substrate of H2S producing enzymes) and DL‐Propargylglycine (DL‐PAG; CSE inhibitor) on the vascular dysfunction induced by insulin resistance in male Wistar rat thoracic aorta. For that purpose, animals were divided into two main sets that received for 20 weeks: (1) tap water (control group; n=6); and (2) 15% p/v fructose in drinking water (insulin resistance group; n=30). Then, the insulin resistance group were divided into 5 subgroups (n=6 each) which received daily i.p. injections during 4 weeks of: (1) nothing (without pharmacological administration); (2) vehicle (phosphate buffer saline; PBS, 1 ml/kg); (3) NaHS (5.6 mg/kg); (4) L‐Cys (300 mg/kg); and (5) DL‐PAG (10 mg/kg). After 20 weeks, metabolic (oral glucose tolerance test, plasma insulin levels, and Matsuda‐index) and hemodynamics variables by tail‐cuff method as well as vascular function by in vitro experiments were determined. Fructose‐induced insulin resistance leads to: (1) hypertension (without affecting heart rate); (2) hyperinsulinemia; (3) a decrease in Matsuda‐index; and (4) a decrease in carbachol‐induced vasorelaxant responses with no effect on the contractile responses to noradrenaline. Interestingly, after 4 weeks of treatment, NaHS decreased blood pressure and restored vasorelaxant responses to carbachol with no effect on metabolic variables while L‐Cys improved the contractile and vasorelaxant responses to noradrenaline and carbachol, respectively, when compared to vehicle (PBS). On the other hand, DL‐PAG induced a slightly increase in systolic and median blood pressure with no effect on the contractile response to noradrenaline or relaxant responses to carbachol as well as metabolic variables compared to vehicle. Taken together, these results suggest that chronic treatment with NaHS and L‐Cys improved vascular dysfunction and hypertension induced by insulin resistance and may have a potential therapeutic application.

Support or Funding Information

The authors acknowledge to Conacyt (Grant No. 252702) for its financial support.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cardiovascular Effects Following a Single Inhalation Exposure to Crude Oil Vapor
Authors: Krajnak Kristine , Mckinney Walter , Waugh Stacey , Hong Kan , Kashon Michael , Fedan Jeffrey
Published: June 2021

Inhalation of crude oil vapor fumes has resulted in serious health effects in worker working in oil extraction, drilling, processing and refinement. These vapors can negatively affect functioning of the pulmonary, cardiovascular, central nervous, renal and reproductive systems. In this study, the effects of a single exposure to crude oil vapor on the peripheral and cardiovascular systems were assessed using a rat inhalation model. Rats received a single 6 hour long exposure to either filtered air or 300 PPM of total volatile organic compounds (TVOC) released from crude oil heated to 85°F, and the effects on the cardiovascular system were measured 1 and 28 days following the exposure. Inhalation of crude oil vapors resulted in reductions in diastolic and mean arterial blood pressures and baseline left ventricular diastolic pressure one day following the exposure. These changes in cardiovascular function were accompanied by a decreased vasoconstriction in response to the β‐adrenergic agonist, dobutamine, and an increase in hydrogen peroxide concentrations in the heart. There were also transient increases in the expression of the pro‐inflammatory factor, tumor necrosis factor‐α (TNFα) and hypoxia‐induced factor‐1 (HIF‐1), but reductions in the expression of interleukin‐6 and the apoptotic factor Bcl2 in the heart. There were no changes in the responsiveness of the ventral tail artery to phenylephrine‐induced vasoconstriction or acetylcholine induced re‐dilation. Twenty‐eight days following exposure, only the expression of TNFα was increased. The other inflammatory factors returned to baseline levels. Based on the results of this experiment, there are effects of crude oil inhalation on cardiovascular function that may be then result of transient hypoxia and longer‐term, low level of inflammation in the heart. Longer‐term exposures will determine whether the effects seen with an acute exposure are maintained, and lead to additional symptoms of cardiac dysfunction, and identify biomarkers that can be used to monitor these changes.

Support or Funding Information

This work was funded as part of the authors official duties as an employee at NIOSH

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The Effects of Metformin, Aminoguanidine, and Pyridoxamine on Methylglyoxal Induced Cardiac Myocytes Injury
Authors: Harvey Amanda , Ahmed Hafsa , Delgado Dean , Kim Alisa , Colangeli Julia , Sandu Robinderpal , Martorana Rose , Barsotti Robert , Young Lindon , Chen Qian
Published: June 2021

In diabetics, formation of advanced glycation end (AGE) products has been closely associated with cardiovascular complications. In order to mitigate the injury caused by AGEs, it is critical to understand the influence of the AGE precursor methylglyoxal. Methylglyoxal is a glycation intermediate and it has been found to be elevated in the blood of diabetic patients. The impact of methylglyoxal on cardiac cells is not well elucidated. In this study, the effects of methylglyoxal on H9C2 myoblast cell viability were evaluated. Thereafter, methylglyoxal induced cell injury was investigated by co‐treatment of methylglyoxal with metformin, aminoguanidine hydrochloride, or pyridoxamine dihydrochloride. Cell viability was evaluated after incubation of drugs for 24 hours by measuring absorbance at 450 nm by using tetrazolium to differentiate metabolically active and inactive cells (e.g., CCK‐8 kit). We found that methylglyoxal (1200 μM) significantly reduced cell viability by 72±6% when compared to the untreated control (n=4, p<0.05). By contrast, co‐treatment of metformin (1–80 mM) significantly reduced methylglyoxal‐induced cell injury and increased cell viability to 69% – 122± 6% (n=4, p<0.05 vs. methylglyoxal) when compared to the untreated control. Similarly, aminoguanidine hydrochloride (250–1000 μM) treated cells significantly increased cell viability to 77% – 109±10% (n=4, p<0.05 vs. methylglyoxal) when compared to the untreated control. Additionally, pyridoxamine dihydrochloride (0.1–15 μM, n=4) showed dose‐dependent protection to increase cell viability. Highest pyridoxamine (15 μM) treated cells completely abolished methylglyoxal induced cell injury by increasing cell viability to 99± 4% (p<0.05 vs. methylglyoxal) when compared to the untreated control. However, when the three drugs were washed out from the medium prior to the addition of the methylglyoxal, the protective effects were lost and cell viability was reduced similarly as methylglyoxal (n=1). Administration of all three drugs alone did not affect cell viability after incubation for 24 hours. The preliminary data suggests that metformin, aminoguanidine hydrochloride, or pyridoxamine dihydrochloride, when incubated concurrently with a high concentration of methylglyoxal, protect cardiac cells from methylglyoxal‐induced cell injury. The mechanism underlying the protective effects will be further investigated.

Support or Funding Information

This study was supported by the Center for Chronic Disorders of Aging, the Division of Research and the Department of Bio‐Medical Sciences at Philadelphia College of Osteopathic Medicine.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Carvedilol Repurposing in Orofacial Pain: Preclinical Findings in Adult Zebrafish
Authors: Barros Adriana Rolim Campos , Santos Sacha Aubrey Alves Rodrigues , Matias Bruna Barbosa , Vieira‐Neto Antonio Eufrásio , Magalhães Francisco Ernani Alves , Júnior Otacilio Benvindo Deocleciano
Published: June 2021

Carvedilol is a non‐selective β‐adrenergic antagonist that is used in the treatment of heart diseases. β‐blockers are known to have an anti‐inflammatory effect that contributes to its analgesic effect. Here we show the acute orofacial antinociceptive effect of carvedilol in adult zebrafish (Danio rerio). Acute nociception was induced by capsaicin (4.93 μM; TRPV1 agonist), formalin (0.1 %; TRPA1 agonist) or menthol (1.2 μM; TRPM8 agonist) into in the upper lip (5.0 μL) of adult wild (short‐fin phenotype) zebrafish. Corneal nociception was induced with hypertonic saline (TRPV1 agonist; 5.0 M NaCl; 5.0 μL) applied into the surface of the right or left eye of the fishes. Then, animals (n=6/group) were placed in a glass Petri dish (10 × 15 cm), divided into quadrants, and the nociceptive response was registered after capsaicin (10 – 20 min), formalin (0 – 5 and 15 – 30 min), menthol (0 – 10 min) or hypertonic saline (0 – 5 min) application. Zebrafish were pretreated by gavage (20 μL) with vehicle (Control; 3 % Tween 80) or carvedilol (0.03; 0.1 or 0.3 mg/mL) 60 min before induction. The effect of carvedilol on zebrafish locomotor behavior was evaluated with the open field test (0 – 5 min). Naive groups (n=6) were included. The interaction between carvedilol and TRPV1 was analyzed using molecular docking. The HEX software was used for molecular docking calculations and the direct action of carvedilol on TRPV1 was viewed using PyMOL v1.4.7. The experimental protocols followed the ethical guidelines of CONCEA (Brazilian Council for the Control of Animal Experimentation) and were approved by the UECE Animal Research Ethics Committee (#7210149/2016). Pre‐treatment with carvedilol (0.03 and 0.1 mg/mL) was associated with a reduction in nociceptive behavior induced by capsaicin and hypertonic saline (TRPV1 agonists; (*p<0.05 vs Control), but not by formalin, or menthol. Carvedilol per se promoted no alteration on the locomotor activity of zebrafish. In the docking experiments, the interaction between carvedilol and the TRPV1 channel showed high reproducibility. It was possible to observe a direct action of carvedilol on TRPV1 amino acids (Asn687, Asn687, Asn687, Asn687, Ala680 and Gly683). The results indicate the potential clinical application of carvedilol an inhibitor of orofacial nociception and that this effect may be due to the modulation of TRPV1 channel.

Support or Funding Information

National Council for Scientific and Technological Development ‐ CNPq

Coordination for the Improvement of Higher Education Personnel ‐ Capes

Research Support Foundation of Ceará State ‐ Funcap

Edson Queiroz Foundation ‐ FEQ

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Treatment of aged mice with the mitochondria targeted antioxidative peptide SS‐31 protects against hypertension‐induced cerebral microhemorrhages
Authors: Tarantini Stefano , Kiss Tamas , Yabluchanskiy Andriy , Balasubramanian Priya , Csipo Tamas , Lipecz Agnes , Sonntag William Edmund , Csiszar Anna , Ungvari Zoltan
Published: June 2021

Recent studies demonstrate that aging exacerbates hypertension‐induced cognitive decline, but the specific age‐related mechanisms remain elusive. Cerebral microhemorrhages (CMHs) are associated with rupture of small intracerebral vessels and are thought to progressively impair neuronal function. To determine whether aging exacerbates hypertension‐induced CMHs young (3 mo) and aged (24 mo) mice were treated with angiotensin‐II plus L‐NAME. We found that the same level of hypertension led to significantly earlier onset and increased incidence of CMHs in aged mice than in young mice, as shown by neurological examination, gait analysis and histological assessment of CMHs in serial brain sections. Our data suggest that aging promotes CMHs in mice likely by exacerbating hypertension‐induced oxidative stress and MMP activation. We found that hypertension increased mitochondrial free radical production in the wall of cerebral arteries. Treatment of aged mice with the mitochondria‐targeted antioxidative tetrapeptide SS‐31 exerted significant vasoprotective effects and significantly delayed the onset and reduced the incidence of CMHs. Thus, therapeutic strategies that reduce mitochondrial oxidative stress may be useful for the prevention of CMHs in the aged brain.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Age‐related alterations in gait function in freely moving male C57BL/6 mice: translational relevance of decreased cadence and increased gait variability
Authors: Tarantini Stefano , Yabluchanskiy Andriy , Fulop Gabor Aaron , Kiss Tamas , Perz Aleksandra , O'Connor Daniel , Johnson Emily , Sorond Farzaneh , Ungvari Zoltan , Csiszar Anna
Published: June 2021

Age‐related gait dysfunction and balance disorders are a major cause of falls and injury in the elderly population. Epidemiological studies have shown that disturbances in gait coordination which manifest with age are associated with increased morbidity and mortality, impaired cognitive capacity, as well as reduced level of function and loss of independence. In geroscience mice are the most frequently used model system to test efficiency of anti‐aging interventions. Despite the clinical importance of age‐related gait abnormalities in older adults, the impact of aging on mouse gait coordination is not well documented in the literature. To characterize the effect of aging on mouse gait, we assessed gait function in young (3 month old) and aged (24 month old) freely moving C57BL/6 mice using the semi‐automated, highly sensitive, Catwalk XT system for quantitative assessment of footfall and motor performance. We found that aged mice exhibited significantly decreased cadence and increased stride time variability. Aging also tended to alter footfall patterns. In aged mice speed, swing speed, stride length, duty cycle, base of support, terminal dual stance, the regularity index and the gait symmetry index were unaltered. Thus, aging is associated with characteristic alterations in gait function in C57BL/6 mice, which could potentially be assessed as clinically relevant endpoints in geroscience studies testing the effects of anti‐aging interventions.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Mir‐1976 Downregulation and Phosphorylation of LARP6 are Associated with Insulin‐like Growth Factor‐1 (IGF1)‐induced Collagen Upregulation in Vascular Smooth Muscle Cells
Authors: Danchuk Svitlana , Delafontaine Patrice , Higashi Yusuke
Published: June 2021

Collagen content in atherosclerotic plaque is a hallmark of plaque stability. We have shown that insulin‐like growth factor‐1 (IGF1) increased collagen content in atherosclerotic plaques of Apoe−/− mice. We have identified LARP6 (La Ribonucleoprotein Domain Family, Member 6), a collagen mRNA binding protein, as a mediator of IGF1‐dependent collagen I upregulation and increased mature fibril formation in smooth muscle cells (SMCs), however specific mechanisms have not been elucidated yet. A total of six micro RNAs (miRs), including miR‐1976, have been shown to bind to LARP6 mRNA. In fact, miR‐1976 mimic (20 nM) decreased LARP6 by 67% and reduced procollagen I by 85% (both are P<0.05), indicating that miR‐1976 negatively regulates LARP6 expression. We found that IGF1 significantly downregulated miR‐1976 by 41±5% in SMCs after 3h of exposure. Moreover, miR‐1976 mimic inhibited IGF1‐induced LARP6 upregulation, suggesting that the downregulation of miR‐1976 mediates IGF1 upregulation of LARP6.

It has been reported that LARP6 phosphorylation is critical for regulating translation of collagen polypeptides. To investigate whether IGF1 induces LARP6 phosphorylation, we assessed LARP6 electrophoretic mobility shift by SDS‐PAGE and immunoblotting. SMCs were serum‐starved and treated for 0–48h either with 10 ng/ml IGF1, or 2 ng/ml IFNy, or 2 ng/ml IL‐1b, or 2 ng/ml TNFa, or 10 ng/ml IL‐6. LARP6 protein was detected on immunoblots as a single 65kDa band in untreated SMCs. However, additional 67kDa LARP6 band was evident after 6h of cell treatment with IGF1 or with IL‐6. The density of 67kDa LARP6 band was gradually increased up to 18h of treatment. IGF1 and IL‐6 also increased expression levels of procollagen I and collagen I in a time‐dependent manner, with a peak at 18h. IFNy, IL‐1b, and TNFa did not induce 67kDa LARP6 band or increase in expression level of collagen in SMCs.

Taken together, our data suggest that IGF1 promotes collagen expression by upregulating LARP6 levels via a downregulation of miR‐1976 and also by inducing phosphorylation of LARP6. Suggested mechanisms by which IGF1 induces collagen expression in vascular SMCs and thus potentially in atherosclerotic plaques would provide novel insights for a therapeutic approach increasing plaque stability.

Support or Funding Information

NIH/NHLBI R01HL070241 (PD)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Reduction of Vascular Reactivity with Dantrolene in Type 1 Diabetic Rats is Dose‐dependent
Authors: Roman Marie , Ramirez Jose , Torres Ricardo , Marrero Victor , Garcia Laura , Morales Myrna , Torres Hector , Crespo Maria J
Published: June 2021

Diabetics have a greater predisposition for developing cerebral vasospasms (CVSPs) than nondiabetics, even when under glycemic control. The etiology of CVSPs is associated with increased vascular tone, secondary to augmented intracellular Ca+2 in vascular smooth muscle. Current pharmacological treatments are ineffective in reducing CVSPs. The combination of the ryanodine receptor blocker dantrolene (50 μM) and the Ca+2 channel blocker nimodipine significantly decreases vascular reactivity in streptozotocin (STZ)‐induced diabetic rats. In order to determine if the beneficial effects observed at 50 μM are also maintained at a much lower concentration with less potentially harmful secondary effects, we evaluated the vascular effects of 10 μM dantrolene, alone and in combination with 50 nM nimodipine. Dose‐response curves for the phenylephrine (PHE)‐induced contraction (0.1 nM to 10 μM) and acetylcholine (ACh)‐induced relaxation (1 nM to 10 μM) were performed on aortic rings from STZ‐diabetic rats, both before and after a 30‐minute incubation period with dantrolene alone or in combination with nimodipine. Whereas 50 μM dantrolene reduced PHE‐induced contraction by 47% in diabetic rats and 29% in controls, 10 μM dantrolene failed to reduce this parameter in either group. Moreover, 50 μM dantrolene reduced PHE‐induced contraction by about 80% in both diabetic and controls when combined with nimodipine. The combination of 10 μM dantrolene and nimodipine, however, does not increase the reduction of 35% in diabetic rats and 47% in controls obtained with nimodipine alone. In addition, only 50 μM dantrolene increased ACh‐induced relaxation by 50% (N=9, P<0.05) in diabetic rats. These results suggest that both, improved endothelial‐dependent relaxation and reduced vascular contraction with dantrolene are dosedependent. Thus, careful titration should be performed in order to achieve a significant reduction in hyper‐reactivity in both diabetics and nondiabetics.

Support or Funding Information

This work was supported by Grants from the National Institute of Health (MBRS‐RISE Grant R25GM061838), the NIMHD‐RCMI Program (Grant G12‐MD007600), and the Anesthesiology Department of the University of Puerto Rico‐School of Medicine.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Blockade of Cardiac Hypertrophy and Fibrosis by TGF‐Beta 1 Receptor Antagonist in Npr1 Gene‐Knockout Mice
Authors: Samivel Ramachandran , Subramanian Umadevi , Oakes Joshua M , Chen Helene , Zhao Hanqing , Gardner Jason D , Pandey Kailash N
Published: June 2021

Genetic disruption of guanylyl cyclase/natriuretic peptide receptor‐A (GC‐A/NPRA) gene (Npr1) in mice triggers cardiac hypertrophy, fibrosis, and remodeling associated with heart failure. However, the specific mechanisms of cardiac defects in Npr1 null mutant mice is not well understood. The objective of this study was to determine the mechanisms regulating the development of cardiac hypertrophy and fibrosis in Npr1 gene‐knockout mice. The Npr1 null mutant (Npr1−/−, 0‐copy), heterozygous (Npr1+/−, 1‐copy), and wild‐type (Npr1+/+, 2‐copy) mice were orally administered with transforming growth factor‐beta 1 (TGF‐β1) receptor antagonist GW788388 (1 mg/kg/day) for 28 days, the hearts were isolated, weighted, and used for quantification of hypertrophic and fibrotic markers using real time quantitative reverse transcription‐polymerase chain reaction (qRT‐PCR) and Western blot analyses. Together, systolic blood pressure (SBP), heart weight‐to‐body weight (HW/BW) ratio, and cardiac fibrosis were analyzed. HW/BW ratio was significantly higher in Npr1 knockout mice than wild‐type control mice. The Npr1−/− mice showed significant induction of cardiac hypertrophy and fibrosis with markedly induced expression of hypertrophic marker collagen‐1α (3.5‐fold) and fibrotic markers monocyte chemoattractant protein (4‐fold), connective tissue growth factor (CTGF, 5‐fold), alpha‐smooth muscle actin (α‐SMA, 4‐fold), and SMAD proteins (SMAD‐2, 5‐fold; SMAD‐3, 3‐fold) compared with wild‐type control mice. In addition, the expression of phosphorylated extracellular‐regulated kinase 1/2 (pERK1/2) proteins were also increased in Npr1−/−mice. On the other hand, Npr1−/−mice treated with GW788388 showed significant inhibition of cardiac hypertrophy and fibrosis and reduced expression of fibrotic markers and SMAD proteins compared to vehicle‐treated wild‐type mice. In contrast, the expressions of pERK1/2 proteins were unaffected in drug‐treated animals. The results of the present study suggest that development of cardiac hypertrophy and fibrosis in Npr1−/− mice is regulated independently through TGF‐β1‐mediated SMAD‐dependent pathways.

Support or Funding Information

This work was supported by NIH grants (HL057531 and HL062147).

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
SGLT2 Inhibition Is Predicted to Reduce LV End Diastolic Pressure: A Mathematical Modeling Analysis
Authors: Basu Sanchita , Boulton David , Hallow Melissa Karen
Published: June 2021

In the recent cardiovascular outcomes trials, Sodium Glucose Cotransporter‐2 Inhibitors (SGLT2i) were shown to reduce heart failure hospitalization, but the mechanisms underlying this effect remain unclear. While SGLT2i's direct mechanism of action is in the kidney, the consequences of its renal mechanisms may indirectly induce changes that improve cardiac hemodynamics. We sought to use a mathematical modeling approach to predict the effect of SGLT2i on cardiac hemodynamics in healthy and diabetic virtual subjects.

We conjugated a mathematical model of cardiorenal hemodynamics and volume homeostasis with clinical measures of plasma and urinary sodium, water, and creatinine in healthy volunteers who were subjected to dapagliflozin for 7 days. We have shown in our previous study that accounting for direct SGLT2 inhibition, coupled NHE3 downregulation and osmotic diuresis, allowed the model to describe the renal response to SGLT2i. We then simulated the effect of SGLT2i on cardiac hemodynamics.

Our simulations predicted a considerable decrease in left ventricle end diastolic pressure (LV EDP) as well as a small decrease in mean arterial pressure (MAP) from the baseline in healthy virtual subjects. In diabetic virtual patients, the predicted decrease in LV EDP was greater than that of healthy volunteers. Ejection fraction (EF) and cardiac output (CO) were minimally changed. While experimental validation is needed, a decrease in LV EDP, without a commiserate decrease in EF and CO, would be expected to reduce fluid congestion and edema, and may help explain how SGLT2i prevent the development of congestion and edema in patients at risk for heart failure. We are currently evaluating the expected effect of SGLT2i on cardiac hemodynamics in virtual patients with existing heart failure. This model provides a means for quantitatively understanding the link between SGLT2i's renal mechanisms and their consequent effects on the failing heart.

Support or Funding Information

This research is funded by AstraZeneca, Pfizer and Merck.

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Differential Regulation of Cardiac Substrate Utilization in Response to Chronic Central Nervous System Administration of Leptin and Melanotan II in Rats with Myocardial Infarction
Authors: da Silva Alexandre A. , Harmancey Romain , Dai Xuemei , Moak Sydney P. , Roy Cary N. , Wang Zhen , Hall John E. , do Carmo Jussara M.
Published: June 2021

The brain leptin‐melanocortin system pathway plays an important role in regulating many physiological functions including energy homeostasis. We found that chronic intracerebroventricular (ICV) infusion of leptin or the melanocortin 4 receptor (MC4R) agonist Melanotan II, MTII, markedly attenuated cardiac dysfunction in rats with myocardial infarction (MI) induced by permanent left anterior descending coronary artery (LAD) ligation. In the present study we examined if activation of leptin‐MC4R axis in the central nervous system (CNS) alters cardiac substrate utilization in rats with MI. Male Sprague‐Dawley rats at 15 weeks of age were implanted with an ICV cannula into the lateral ventricle. After 10 days of recovery, the LAD was permanently ligated and saline vehicle (0.5 μL/hr, n=5), leptin (15 μg/day, n=5) or MTII (240 ng/day, n=3) was infused ICV via osmotic minipump for 14 consecutive days. On day 14 of treatment the hearts were quickly excised and perfused ex vivo in the working mode with glucose (5.5 mmol/L) and the fatty acid oleate (0.4 mmol/L). After a 5‐min baseline equilibration period the hearts were perfused for an additional 20 min and coronary flow samples were collected every 5 min to determine rates of substrate oxidation by quantitative measurement of [14C]O2 and [3H]2O using [U‐14C]glucose (0.08 μCi/ml) and [9,10‐3H]oleate (0.1 μCi/ml) as the radiolabeled tracers. Compared to vehicle treatment, chronic ICV leptin infusion for 2 weeks markedly increased glucose oxidation (0.60±0.04 vs. 0.27±0.02 μmol/min/g dry heart weight) without altering oleate oxidation (0.95±0.01 vs. 0.90±0.02 μmol/min/g dry heart weight). Chronic central MTII infusion did not alter glucose oxidation (0.25±0.03 vs. 0.27±0.02 μmol/min/g dry heart weight) but increased fatty acid oxidation compared to vehicle treatment (1.13±0.01 vs. 0.90±0.02 μmol/min/g dry heart weight). These results suggest the CNSmechanisms by which leptin and MC4R activation improve cardiac function post MI may involve increased myocardial bioenergetics by differential modulation of glucose and fatty acid utilization.

Support or Funding Information

NHLBI PO1HL51971, NIGMS P20GM104357 and NIGMS U54GM115428

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Chemotherapy acutely impairs neurovascular and hemodynamic responses in patients with breast cancer
Authors: Sales Allan Kluser , Negrao Marcelo , Testa Laura , Ferreira‐Santos Larissa , Groehs Raphaela , Carvalho Bruna , Toschi‐Dias Edgar , Rocha Natalia , Laurindo Francisco , Debbas Victor Kluser , Rondon Maria Kluser , Mano Max , Hajjar Ludhmila , Hoff Paulo , Filho Robero , Negrao Carlos
Published: June 2021

Purpose

The purpose of the present study was to test the hypothesis that doxorubicin (DX) and cyclophosphamide (CY) adjuvant chemotherapy (CHT) acutely impairs neurovascular and hemodynamic responses in patients with breast cancer.

Methods

Sixteen women (age: 47.0±2.0 years; body mass index: 24.2±1.5 Kg.m−2) with stage II–III breast cancer and indication for adjuvant CHT underwent two experimental sessions, saline (SL) and CHT. In the CHT session, DX (60 mg/m2) and CY (600mg/m2) were administered in 45 min. In the SL session, a matching SL volume was infused in 45 min. Muscle sympathetic nerve activity (MSNA, microneurography), calf blood flow (CBF, plethysmography) and calf vascular conductance (CVC), heart rate (HR, electrocardiography) and beat‐to‐beat blood pressure (BP) (Finger plethysmography) were measured before, during and after each session. Blood samples (5 mL) were collected before and after both sessions for assessment of circulating endothelial microparticles (EMPs, flow cytometry), a surrogate marker for endothelial damage.

Results

MSNA and BP responses were increased (P<0.001), whereas CBF and CVC responses were decreased (P<0.001), during and after CHT session when compared to SL session. Interestingly, the vascular alterations were also observed at the molecular level through an increased EMPs response to CHT (P=0.03 vs. SL session). No difference in HR response was observed (p>0.05).

Conclusions

Adjuvant CHT with DX and CY in patients treated for breast cancer leads to acute sympathetic hyperactivity and endothelial alterations, which results in reduced muscle vascular conductance and elevated systemic BP. These responses may anticipate susceptibility to CHT‐induced longterm cardiovascular alterations.

Support or Funding Information

São Paulo Research Foundation ‐ FAPESP

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Epidural spinal cord stimulation for neuropathic pain reduces blood pressure in patients with hypertension independent of pain relief: A retrospective study
Authors: Holwerda Seth W. , Holland Marshall T. , Green Alex L. , Collins Michael T. , Pearson Amy C. , Pierce Gary L.
Published: June 2021

Evidence suggests that epidural spinal cord stimulation (SCS) reduces peripheral sympathetic outflow in patients treated for peripheral artery disease and neuropathic pain. However, the extent to which SCS reduces arterial blood pressure (BP) has received little attention. We tested the hypothesis that systolic BP (SBP) would be reduced in patients with hypertension (HTN) receiving surgically implanted SCS for management of neuropathic pain, and that SBP reduction would be independent of reduced neuropathic pain. Retrospective analysis of daytime SBP from patient medical records at the University of Iowa was performed from 2008 to 2018, and event‐triggered averaging was used to determine 2‐year SBP averages prior to SCS implant (pre‐SCS) and 2‐year SBP averages after SCS implant (SCS) in stage 1 HTN (n=170) and stage 2 HTN (n=141) patients. SCS SBP was significantly lower compared with pre‐SCS SBP (Pre‐SCS: 140±1 vs. SCS: 134±1 mmHg, P<0.001), and the reduction was greater in stage 2 HTN compared with stage 1 HTN (Stage 2: −11±1 vs. Stage 1: −2±1 mmHg, P<0.001), while reduction in neuropathic pain (scale 1–10) was not significantly different between groups (Stage 2: −1.4±0.3 vs. Stage 1: −1.1±0.2, P=0.37). Reduction in SBP following SCS was significantly greater in HTN patients on anti‐HTN medications compared with no anti‐HTN medications (Meds: −9±2 vs. No meds: −5±1 mmHg, P=0.036). The reduction in SBP following SCS was significantly correlated with higher Pre‐SCS SBP after controlling for sex, age, number of anti‐HTN medications, and reduction in neuropathic pain (R=−0.42, P<0.001). These findings suggest that SCS treatment for neuropathic pain in patients with HTN reduces BP independent of reductions in pain, and that the reduction in BP may be potentiated by anti‐HTN medications.

Support or Funding Information

AHA (17POST33440101), T32 HL07121, NHLBI (P01HL014388), U54 TR001356 and AHA (13DG143400012)

This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Dynamic and Static Handgrip Exercise Increase Wave Reflection in Healthy Young Adults
Authors: Stock Joseph M , Chouramanis Nicholas V , Chirinos Julio A , Edwards David G
Published: June 2021

Increased wave reflection augments pulsatile load, wastes left ventricular effort, and is associated with cardiovascular events. Acute handgrip (HG) exercise increases central blood pressure and surrogate measure of wave reflection, augmentation index. However, surrogate measures of wave reflection do not provide a comprehensive assessment of pulsatile load and ventricular‐arterial interactions. Wave separation analysis can determine the relative contributions of forward (Pf) and backward (Pb) pressure wave amplitudes and reflection magnitude (RM=Pb/Pf) as well as the timing of apparent wave reflection return.

PURPOSE

To determine the effect of acute dynamic and static HG exercise on wave reflection magnitude and timing with wave separation analysis in young healthy adults.

METHODS

Applanation tonometry was used to record radial artery pressure waveforms in 31 adults (25±4 years) at baseline and during dynamic and static HG exercise in randomized order. Radial waves were calibrated to respective brachial mean and diastolic pressures and synthesized to central pressure waves using a generalized transfer function. Wave separation analysis was used to derive Pf, Pb, RM, and reflected wave transit time which was performed offline using a synthesized flow waveform.

RESULTS

Central blood pressure and augmentation index increased with HG exercise as expected (p<0.05). Compared to baseline, both HG exercises increased wave reflection measures Pb (baseline 11.4±0.4 mmHg; dynamic 12.6±0.5 mmHg; static 12.7±0.6 mmHg, p<0.05) and RM (baseline 40±1%; dynamic 43±1%; static 44±1%, p<0.05), but not Pf (baseline 28.4±1.1 mmHg; dynamic 29.1±1.2 mmHg; static 28.8±1.2 mmHg, p>0.05). Reflected wave transit time was shortened with static but not dynamic HG exercise compared to baseline (baseline 95±4 ms; dynamic 90±3 ms; static 83±3 ms, p<0.05)

CONCLUSION

Both HG exercises increased measures of wave reflection (Pb and RM) suggesting greater wave reflection during HG exercise.

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