bluuebunny/crossref_metadata_2025 huggingface dataset:
B cells from lupus‐prone mice respond more vigorously to lipopolysaccharide stimulation (835.5)
Authors: Roberts Rebecca , Kane Andrew , Doyle Chris , Orlick Andrew , Thiele John , Thiele Karla , Zuber Mathew
Published: June 2021

Systemic Lupus Erythematosus (SLE) is an autoimmune disease characterized by its predominance in women and the production of autoantibodies by B cells. The causes of SLE are mostly unknown, but it has been linked to genetics, viruses and environmental factors. In order to determine how the immune system responds differently in an SLE context, this study investigated the response to lipopolysaccharide (LPS) by splenic B cells from two types of mice, control C57BL/6 and lupus‐prone NZB/WF1. Using fluorescence activated cell scanning (FACS) we found that NZB/WF1 mice have lower overall levels of splenic B cells than C57BL/6, making up 46% and 57.5%, respectively, of the total splenic lymphocyte cell population. However, those B cells are more responsive to LPS stimulation as indicated by a larger increase in blasting. Moreover, B cells from NZB/WF1 constitutively express lower surface levels of Major Histocompatibility Complex Class II (MHC‐II), yet again the relative increase upon LPS exposure is higher for NZB/WF1 (65%) compared to C57BL/6 (27%). A similar effect is seen for CD86, but not CD80. These findings may point to the importance of changse in B cell population in Lupus, where B cell overactivity is implicated in etiology.

Grant Funding Source: Supported by Van Sant and a Center for Science and the Common Good mini‐grant sponsored by HHMI
The action of photodynamic therapy on Candida species (836.10)
Authors: Doria Anelise Cristina , Bevilacqua Isabela , Santos Thaisa , Teodoro Guilherme , Oliveira Elizabeth , Khouri Sonia , Nicolau Renata
Published: June 2021

Candida yeasts are normal commensal of human microbiota, which can cause infection in patients with predisposing factors. Photodynamic antimicrobial therapy has been employed against several microbial infections. Light, when associated with dyes, promotes microbial death in a less invasive way, reducing the treatment time. The aim of the present study was to investigated the effect of Photodynamic Therapy (PDT) with MetyleneBlue (MB) and a light emitting diode (LED), on Candida species.Cultures ofC. albicans, C. glabrata, C. tropicalis, C. parapsilosis, C. kruseiand C. dubliniensisweresubmmited to photodynamic therapy employing LED and Methylen blue. The number of CFU/mL was reduced in 100% for Candida glabrata, 99.57% for Candida albicans, 99.41% for Candida krusei, 97.53% for Candida dubliniensis, 97.29% for Candida tropicalis, and 96.88% for Candida parapsilosis when compared with the control group. The results showed that the PDT significantly reduced the CFU of Candida. The result of this study allows us to conclude that all Candida species tested were susceptible to the PDT treatment.
Deletion of soluble epoxide hydrolase modulates coronary reactive hyperemia in isolated mouse hearts (837.10)
Authors: Hanif Ahmad , Zeldin Darryl , Nayeem Mohammed
Published: June 2021

Coronary reactive hyperemia (RH) is impaired in hypertrophic hearts and in hearts with metabolic syndrome. Epoxyeicosatrienoic acids (EETs) exert cardioprotective effects against ischemia / reperfusion injury. Soluble epoxide hydrolase (sEH) is the main enzyme responsible for EETs breakdown. We hypothesized that in the absence of sEH, isolated mouse hearts exhibit increased RH through increased generation of EETs in response to no‐flow ischemia compared to wild type (sEH+/+). Coronary flow (CF) in isolated sEH+/+ and sEH‐/‐ mouse hearts was measured using Langendorff system. Perfused isolated hearts were exposed to 15 second ischemia and RH parameters were assessed. Following ischemia, flow repayment (calculated as the area under the curve and normalized to heart weight (ml/g)) was significantly higher in sEH‐/‐ compared to sEH+/+ hearts (7.62 ± 0.47 in sEH‐/‐ vs. 5.86 ± 0.68 in sEH+/+, p<0.05). Moreover, sEH‐/‐ hearts exhibited significantly higher repayment to debt ratio when compared to sEH+/+ hearts (2.06 ± 0.17 in sEH‐/‐ vs. 1.48 ± 0.14 in sEH+/+, p<0.05). No significant differences were observed in baseline coronary flow, debt area, peak hyperemic flow and repayment duration between the two groups. Our results demonstrate that RH, as measured by flow repayment and repayment to debt ratio, is significantly higher in sEH‐/‐ compared to sEH+/+mice. These data suggest that targeting or silencing sEH could have a beneficial effect on myocardial recovery from ischemia. Supported by HL‐114559 to MAN and z01‐ES025034 to DCZ.

Grant Funding Source: Supported by HL‐114559 to MAN and z01‐ES025034 to DCZ
Acute increases of O‐GlcNAcylation reduces sepsis‐associated mortality (837.8)
Authors: Olivon Vania , Ferreira Raphael , Mestriner Fabíola , Alves‐Filho José , Cunha Fernando , Tostes Rita
Published: June 2021

The post‐translational modification of proteins by O‐GlcNAcylation (O‐GlcNAc) is highly dynamic and modulates cell‐signaling processes. Acute increases in O‐GlcNAc levels reduce migration of inflammatory cells and the release of pro‐inflammatory mediators, important events in sepsis caused by bacterial infection or multiple non‐infectious causes. This study tested the hypothesis that acute increases of O‐GlcNAc levels reduce mortality and inflammatory processes in experimental models of sepsis. C57/BL6 mice were used in experimental sepsis protocols cecal ligation and puncture (CLP) and lipopolysaccharide (LPS)‐induced severe and mild sepsis. Glucosamine treatment (300mg/Kg, i.v. 30 min. before the sepsis induction) acutely increased vascular and spleen O‐GlcNAc levels and increased survival of mice with LPS‐induced sepsis (50%) and CLP sepsis (40%). In mice with LPS‐induced sepsis, glucosamine also reduced neutrophil migration to the peritoneal cavity (severe LPS sepsis, p<0.05; mild LPS sepsis, p<0.01), myeloperoxidase (MPO) activity of lung neutrophils (severe LPS sepsis, p<0.01; mild LPS sepsis, p<0.001) and aortic IL‐1beta mRNA expression (mild LPS sepsis, p<0.01). In the CLP model, glucosamine reduced MPO activity of lung neutrophils (p<0.05). These results show that glucosamine‐induced acute increases of O‐GlcNAc levels increase survival in the CLP and LPS experimental models of sepsis. This study suggests that the O‐GlcNAc pathway represents a potential target in sepsis‐related systemic inflammatory responses.

Grant Funding Source: CNPq, CAPES and FAPESP
Animal model for assessment pro‐inflammatory response of fat tissue and liver induced by prenatal stress and diet high in sugar and fat (840.14)
Authors: Bezpalko Lyudmyla , Zayachkivska Oksana , Havryluk Elena
Published: June 2021

Life style diseases (LSD) recognized as one of the non‐inflectional epidemic worldwide and LSD‐related liver pathology is common distributed. Recent data had shown intrauterine (prenatal) programming of adult metabolic phenotype. No study has explored potential interaction between maternal diet high in sugar and fat and stress on adipocytokine secretion and changes of liver functions in adulthood.

The aim: To create a animal model of prenatal stress in rats to examine role of maternal stress and diet on activities of adipose tissue and hepatocellular organization in the offspring which can provide basis in treating hepatopathologies in preclinical and clinical settings.

We used rat offsprings of mothers which had during all period of pregnancy social stress with (1) ad libitum access to 30% saccharose (Kozar, 2009); (2) high‐fat diet (to 45% kcal, Lintermans, 2009); and (3) binary those factors vs control. Scoring lesion (LI), vascular indexes (VI) by fat, liver histomorphology, IL‐1β, GRO/CINC‐1, leptin, adiponectin by ELISA.

Relative to the control the most changes were determined in the 3d group: histologically ‐ both in the visceral fat tissue and liver appeared the cell reorganization with signs of inflammation, rise IL‐1β (217%), GRO/CINC‐1 (99%), leptin (79%) levels and decreased of adiponectin (41% ); rise of ratio Leptin/Adiponectin was in twice.

Conclusion: Taken together maternal stress and high sugar‐fat diet reveal both prenatal effect on prenatal programming of offspring adipocyte secretion with liver proinflammatory changes. This animal model can be useful for follow up pharmacology investigation of pro‐inflammatory and metabolic state in adulthood.
Cytotoxic effects of selective serotonin‐ and serotonin‐norepinephrine reuptake inhibitors on human metastatic breast cancer cell line, MCF‐7 (842.3)
Authors: Bavadekar Supriya , Panchal Palak , Hanbashi Ali , Vansal Sandeep
Published: June 2021

Current evidence suggests that some antidepressants possess growth‐inhibitory effects against a number of cancer cell lines. The present study evaluated seven of the most commonly prescribed antidepressants in the US for cytotoxic effects on MCF‐7 breast cancer cells. These included selective serotonin reuptake inhibitors (sertraline, fluoxetine, paroxetine, citalopram, escitalopram) and serotonin‐norepinephrine reuptake inhibitors (venlafaxine, duloxetine). Viability was determined after 24 h using the XTT cell proliferation assay. All test agents inhibited cell growth in a concentration‐dependent manner. Sertraline (IC50~16 µM) was the most potent, followed by equipotent paroxetine, duloxetine, and fluoxetine (IC50~31‐36 µM), while citalopram, escitalopram, and venlafaxine were significantly weaker. The lead compound, sertraline, induced apoptosis, as evidenced by morphological changes, phosphatidylserine externalization, and poly(ADP‐ribose) polymerase (PARP) cleavage. Future experiments are aimed towards identifying the binding site(s) for sertraline and further elucidating the signal transduction pathway(s) involved in the cytotoxic effect. In summary, the high sensitivity shown by the four most potent agents, and especially sertraline, in inhibiting MCF‐7 growth suggests that these agents should be evaluated further for potential use in anti‐breast cancer therapy.

Grant Funding Source: This work is supported by the Division of Pharmaceutical Sciences at Long Island University.
A systems pharmacology approach to model tyrosine kinase inhibitor‐induced cardiotoxicity gene interaction networks (844.17)
Authors: Sarntivijai Sirarat , Hur Junguk , Ozgur Arzucan , Burkhart Keith , He Yongqun , Omenn Gilbert , Athey Brian , Abernethy Darrell
Published: June 2021

This study implements an integrative bioinformatics workflow to predict gene networks underlying TKI‐induced cardiac adverse events (AEs). The Ontology of Adverse Events (OAE) assisted Natural Language Processing (NLP) algorithm analyzes PubMed abstracts for gene interactions based on the gene centrality in a network.The algorithm utilizes support vector machine (SVM) learning to determine gene pairs of interaction. The gene interactions were mined in association with TKI‐induced cardiac AEs identified by disproportionality analysis of the FDA Adverse Event Reporting System data. These TKIs include dasatinib, imatinib, lapatinib, cetuximab, and trastuzumab. The result gene interaction networks were classified and interpreted for their relevance to the established knowledge confirmed by published experiments. Significant findings include mediator genes linking JUN and ABL1 to JAK‐STAT pathway, and inflammatory response mechanism.
Identifying NMDA receptor subtypes in the ventral respiratory column (847.6)
Authors: Leavitt Lee , Curtice Kigen , Raghuraman Shrinivasan , Teichert Russ , Twede Vernon , Olivera Baldomero
Published: June 2021

The ventral respiratory column (VRC) is a region in the brainstem shown to control breathing patterns in mammals. Using a mouse model, cells from this region are dissociated, plated and incubated with a dye that indicates changes in cytoplasmic calcium levels. Hundreds of cells are monitored as a diverse set of pharmacological agents are applied. The spectrum of cellular responses provides a profile of the receptors found on each of these neurons. Previously, varieties of cell classes were shown to contain NMDA receptors. However, specific compositions of subunits within these receptors are not known. These receptors are ligand gated ion channels composed of four non‐covalently bound protein subunits. Each subunit has a different response profile determined by interactions of agonist and antagonists. Conantokins (peptides isolated from snail venom) and other compounds afford understanding which NMDA receptor subtypes are present in different cell‐classes. This project has continued to classify the subunit compositions of NMDA receptors with the ultimate goal of defining which NMDA receptor subunits are present in each class. The results indicate that different neuronal subclasses express different NMDA response subtypes. This will provide valuable information on the individual neurons in the networks of the VRC, and will allow for pharmacological interventions to change behavior in this region.

Grant Funding Source: Supported by GM48677
Mechanisms of hypoxia‐induced immune escape in cancer and their regulation by nitric oxide (85.1)
Authors: Graham Charles , Barsoum Ivraym , Kim Judy , Black Madison , Macdonald‐Goodfellow Shannyn , Siemens D.
Published: June 2021

The acquired ability of tumour cells to avoid destruction by immune effector mechanisms (immune escape) is important to malignant progression. Also associated with malignant progression is tumour hypoxia, which induces aggressive phenotypes such as invasion, metastasis and drug resistance in cancer cells. Our studies revealed that hypoxia contributes to escape from innate immunity by increasing tumour cell expression of the metalloproteinase ADAM10 in a manner dependent on accumulation of the alpha subunit of the transcription factor hypoxia‐inducible factor‐1 (HIF‐1α). Increased ADAM10 expression leads to shedding of the NK cell‐activating ligand, MICA, from the surface of tumour cells, thereby resulting in resistance to NK cell‐mediated lysis. Our more recent studies demonstrated that hypoxia, also via HIF‐1α accumulation, increases the expression of the inhibitory co‐stimulatory ligand PD‐L1 on tumour cells. Elevated PD‐L1 expression leads to escape from adaptive immunity via increased apoptosis of CD8+ cytotoxic T lymphocytes. We have also shown that activation of nitric oxide signalling inhibits hypoxia‐induced escape from innate and adaptive immunity at least partly by interfering with HIF‐1α accumulation. These findings indicate that approaches designed to interfere with tumour cell adaptations to hypoxia may have useful applications as adjuvants to immunotherapy.
Differential role of microRNAs miR‐221/222 and miR‐103/107 in type 2 diabetes and effects of metformin (851.2)
Authors: Upadhyay Rohit , Ding Hong
Published: June 2021

Background and Aim: Type‐2 diabetes (T2D) is a highly prevalent, progressive disease constituting the major challenge to global health. MicroRNAs (miRNAs) play an important role in regulating physiological processes in diabetes through binding multiple target mRNAs, resulting in gene silencing. In the present study, the primary objective is to explore the role of miR221/222 and miR103/107 in diabetes.

Materials and Methods: Mouse microvascular endothelial cells (MMECs) were cultured for 48 hours under conditions designed to mimic the mileu of T2D mice: normal glucose (NG=11mM) and high glucose (HG=40mM). The levels of miRNA221/222 and mir103/107 expression were analysed by real‐time PCR. MMECs were transfected with specific miR inhibitors/ mimics and then cells were exposed to normal/ high glucose in the presence or absence of metformin. The effects of miR expressions on Akt, P‐Akt, c‐kit, eNOS, P‐eNOS and Cav1 protein levels were determined through western blotting.

Results: We found that exposure to high glucose increased the expression of miR‐221/222 but decreased the expression of miR‐103/107. Treatment with metformin partially restored HG‐induced expression of miRNAs towards that seen in control, NG, mileau. Furthermore, we observed that protein levels of total eNOS and P‐eNOS decreased in HG in comparison with NG. The eNOS/P‐eNOS protein levels were restored upon inhibition of miR221 but not with miR‐222, miR‐103 or miR‐107, thus indicating correlation of miR‐221 with eNOS signalling. Results were validated by using miR‐221 mimics (overexpression) and known target (c‐kit) of miR221.

Conclusion: These findings suggest that miR221 may offer a new therapeutic strategy for treatment of endothelial dysfunction in diabetic patients or may work as a therapeutic modulator.
New insights on endothelial CaMKII in angiotensin II‐induced hypertension (851.6)
Authors: Charbel Chimene , Toussaint Fanny , Blanchette Alexandre , Gillis Marc‐Antoine , Comtois Philippe , Ledoux Jonathan
Published: June 2021

Angiotensin II‐induced arterial hypertension (AngII‐AH) is associated with an endothelial dysfunction characterized by a Ca2+ dyshomeostasis. Alteration of the delicate balance between reactive oxygen species (ROS) and nitric oxide (NO) then arise. Recent evidences suggest that hypertension stimulates Ca2+ pulsars and CaMKII but the contribution of AngII and the concomitant ROS remains elusive. The aim of this study was to establish a relationship between endothelial Ca2+ pulsars, ROS and CaMKII in AngII‐AH. Acute exposure to high (> 10‐8 M) concentrations of AngII stimulated Ca2+ pulsars and ROS production in high‐speed confocal microscopy. However, lower AngII level (10‐9 M) only increased Ca2+ pulsars frequency, suggesting a role for ROS in the Ca2+ pulsars stimulation. All AngII concentrations induced endothelial CaMKII recruitment within myoendothelial projections. AngII also stimulated NO production, an effect blunted by CaMKII inhibition (KN‐93). Candesartan, an AT1 blocker, partially reduced Ca2+ pulsars stimulation, suggesting roles for both AT receptors. Moreover, an increased in Ca2+ pulsars frequency and ROS production, associated with CaMKII clustering, is also observed in AngII‐AH mice. Our findings shed a new light on hypertension and Ca2+ dyshomeostasis along with their relationships with CaMKII, a key regulator of endothelial function.

Grant Funding Source: Supported by CIHR, FRQS, CFI, HSFC and SQHA.
Chronic hypoxia differentially modulates the response of fetal ovine middle cerebral arteries to endothelin‐1 (853.2)
Authors: Silpanisong Jinjutha , Kim Dahlim , Adeoye Olayemi , Thorpe Richard , Pearce William
Published: June 2021

To better understand how hypoxia‐induced vascular remodeling alters intracellular responses to ET‐1, we hypothesized that chronic hypoxia alters ET‐1 induced phenotypic transformation in arterial smooth muscle cells within the fetal cerebrovasculature. Endothelium denuded middle cerebral arteries (MCA) harvested from term fetuses of pregnant adult sheep kept at sea level or 3820m for 110 days were serum starved for 24h, then cultured 24h with ET‐1 and various kinase inhibitors. Fetal normoxic (FN) MCAs incubated with ET‐1 exhibited decreased colocalization between MLCK and α‐actin compared to control (starved), suggesting a shift of smooth muscle cells (SMCs) from a contractile to a more synthetic phenotype. This appeared to be mediated by multiple intracellular pathways, with PKC playing the largest role in normoxic arteries. Chronic hypoxia attenuated the ET‐1 induced decrease in MLCK and α‐actin colocalization, decreased the effects of PKC inhibitors, but increased the apparent effects of the CaMK‐dependent pathway, on colocalization. Given that plasma levels of ET‐1 were unaltered by chronic hypoxia, these results suggest that chronic hypoxia shifts the coupling of ET‐1 from PKC to CaMK to alter gene expression and thereby influence vascular remodeling.

Grant Funding Source: Supported by National Institutes of Health Grants HD‐31266, HL‐64867, and NS‐076945
NPY ő a regulator of hypoxic cerebrovascular remodeling in fetal lambs (853.4)
Authors: Latschaw Kimberly , Adeoye Olayemi , Silpanisong Jinjutha , Kim Dahlim , Williams James , Pearce William
Published: June 2021

Prenatal, intra‐partum or post partum exposure of a fetus to hypoxia results in structural and functional changes in cerebral arteries, known as remodeling. Sympathetic nerves have been suggested to contribute to this process. Whereas NE is suggested as the major mediator of adrenergic effects, our preliminary experiments show that there is a guanethidine‐resistant (norepinephrine‐independent) component. This study explores the hypothesis that NPY contributes to hypoxic cerebrovascular remodeling in the fetal cerebral circulation. Chronic hypoxia (CH) produced a 254% ↑ in K‐induced tone and a 30% ↓ in stretch‐induced myogenic tone in fresh nerve‐intact fetal cerebral arteries. CH also increased the magnitude of nerve stimulation induced guanethidine‐resistant contractile tone. Immunohistochemistry revealed that CH significantly ↑ cerebrovascular NPY whereas sympathectomy (SANX) ablated this effect. Likewise, Western blot quantification of NPY‐Y1 receptors revealed that CH increased Y1 receptors 73%. Whereas SANX in normoxic tissues had no significant effect, SANX produced a 30% ↓ in Y1 receptors in hypoxic arteries. Together these results suggest that NPY released from sympathetic cerebrovascular nerves contributes to hypoxic remodeling through increased NPY content/release and Y1 receptor density.

Grant Funding Source: Supported by NIH Grants HD‐31266 and NS‐076945
Myricetin alleviates postprandial hyperglycemia in rats with diabetes mellitus (854.1)
Authors: Kim Jung‐In , Choi Ha‐Neul , Lee Soo‐Jin , Kang Min‐Jung
Published: June 2021

Tight control of both fasting and postprandial blood glucose is essential in the treatment of diabetes mellitus. It has been suggested that postprandial hyperglycemia is more strongly associated with the risk of cardiovascular diabetic complications than is fasting hyperglycemia. Myricetin strongly inhibits α‐glucosidase in vitro, suggesting that it could have hypoglycemic activity in vivo. This study investigated the α‐glucosidase inhibitory activity of myricetin in vivo. Streptozotocin‐induced diabetic rat were administered a starch solution (1 g/kg) with and without myricetin (100 mg/kg) or acarbose (40 mg/kg) orally after an overnight fast, and postprandial plasma glucose levels were monitored. The administration of myricetin or acarbose significantly decreased the incremental plasma glucose levels 30‐180 min after a single oral dose of starch (p<0.05). Myricetin or acarbose treatment significantly reduced the incremental areas under the glucose response curve (AUC). The AUC of the myricetin‐treated group was not significantly different from that of the acarbose‐treated group. Therefore, we conclude that myricetin is effective for controlling postprandial hyperglycemia in an animal model of diabetes mellitus.

Grant Funding Source: This research was supported by NRF in Korea (2012015763)
Exploring off‐target binding of thiazolidinediones and a novel mechanism for restoring insulin sensitivity (854.2)
Authors: Exner Eric , Hoffmann Brian , Sem Daniel , Greene Andrew
Published: June 2021

The prevalence of type 2 diabetes mellitus (T2DM) is increasing. As insulin resistance is a major feature of T2DM, this will necessitate the increased use of drugs that restore insulin sensitivity. Thiazolidinediones (TDZs) comprise one such class of drugs and are thought to act through peroxisome proliferator‐activated receptor gamma (PPARγ); however, this mechanism is not completely understood, as neither antagonists nor transcriptional/translational inhibitors of PPARγ abolish the effects of these drugs. Moreover, TDZs have been reported to elicit serious side effects that could be mediated through interactions of TDZs with effectors other than PPARγ. We developed a novel method to determine off‐target binding interactions of drugs, which we applied to TDZs. In addition to interactions that could be related to side effects, we observed affinity for salt‐inducible kinase 2 (SIK‐2), which could represent a novel mechanism of action for the insulin sensitizing effects of TDZs. SIK‐2 impairs insulin receptor signal transduction during conditions of nutrient deprivation by phosphorylating insulin receptor substrate 1 (IRS‐1). This study tests the hypothesis that TDZs prevent SIK‐2 mediated phosphorylation of IRS‐1, thus restoring or preserving insulin sensitivity and outlines a method for utilization of existing, effective molecules to identify novel targets for the development of safer therapeutics.

Grant Funding Source: Supported in part by National Heart, Lung, and Blood Institute Grant HL‐082798 to A. S. Greene
Plasma irisin is associated with insulin resistance in men (854.4)
Authors: Huth Claire , Marette André , Tremblay Angelo , Weisnagel S. John , Mauriège Pascale , Lacaille Michel , Joanisse Denis
Published: June 2021

The hormone irisin has been described as increasing energy expenditure and improving glucose tolerance in mice, yet its potential role in humans remains controversial. Objective: We describe the association between circulating irisin and insulin resistance in men with wide‐ranging fitness levels and metabolic health. Methods: Fifty‐two men aged 34‐53 yrs from 4 groups were compared: sedentary non‐obese controls (BMI<25 kg/m2), sedentary obese (BMI>30 kg/m2), sedentary obese glucose intolerant, and non‐obese highly trained endurance active. All participants were evaluated for fitness (VO2max), anthropometric profile, circulating metabolic profile, insulin resistance (hyperinsulinemic‐euglycemic clamp), and baseline plasma irisin levels (EIA, Phoenix Pharamaceuticals, Inc.). Results: Baseline plasma irisin levels were significantly different between groups (ANOVA p=0.01), being lowest in trained men (1406±382 ng/mL) and highest in metabolically deteriorated glucose intolerant subjects (2040±505 ng/mL). Including all subjects, irisin levels were positively associated with percent body fat (r=0.445, p<0.01), and negatively associated with fitness (r=‐0.369, p<0.01) and insulin sensitivity (M/I, r=‐0.355, p<0.01). The strength of these associations was reduced and lost statistical significance when excluding the active individuals, except for that with insulin resistance, which was slightly increased (r=‐0.413, p<0.01). Conclusion: Plasma irisin levels are associated with decreased insulin sensitivity in men, but the association appears to be influenced by training status.

Grant Funding Source: Supported by the CIHR and CRIUCPQ.
Role of E2F transcriptional factor family in hydrogen peroxide regulation of dihydrofolate reductase in endothelial cells (855.5)
Authors: Li Hong , Li Qiang , Siu Kin , Cai Hua
Published: June 2021

Endothelial nitric oxide synthase (eNOS) uncoupling consequent to a deficiency in eNOS cofactor tetrahydrobiopterin (H4B) is an important mechanism of vascular dysfunction. Our previous work has shown that endothelial hydrogen peroxide (H2O2) overproduction specifically downregulates H4B salvage enzyme dihydrofolate reductase (DHFR), which mediates persistent H4B deficiency, eNOS uncoupling and elevated blood pressure in Angiotensin II‐infused mice. However, detailed molecular mechanisms underlying H2O2 regulation of DHFR in endothelial cells remain unclear. In this study, we found that hydrogen peroxide dose and time dependently downregulated DHFR protein expression and its enzyme activity in bovine aortic endothelial cells (BAECs). The DHFR mRNA levels were also decreased by H2O2, implicating transcriptional regulation. Based on the fact that DHFR promoter contains cis‐acting elements for Sp1 and E2F, we constructed wildtype DHFR promoter reporter construct (WT‐DHFR), and mutant DHFR promoter constructs that were deleted of either Sp1 or E2F binding sites. The dual‐luciferase reporter assay showed that while the promoter activities of both WT‐DHFR and Sp1 deletion mutant remained inhibited by H2O2, deletion of the E2F binding sites completely abolished H2O2 attenuation of DHFR promoter activity. RT‐PCR analyses indicated that H2O2 treatment resulted in marked downregulation of E2F1, E2F7 and E2F8 by 38%, 43% and 82% respectively. Intriguingly, overexpression of E2F1, E2F7 or E2F8 restored DHFR expression in response to H2O2, and cotransfection with E2F1 also significantly increased DHFR promoter activity. In conclusion, these results suggest that DHFR is downregulated by H2O2 in endothelial cells transcriptionally via a E2F‐dependent manner, and that restoration of E2F family‐transcription factor activity is effective in preserving DHFR function under oxidative stress.

Grant Funding Source: Supported by NHLBI awards HL077440 (HC), HL088975 (HC) and an AHA EI award 12EIA8990025 (HC)
Resveratrol attenuates endothelium‐dependent cyclooxygenase‐mediated contractile activity in the carotid artery of spontaneously hypertensive rats (855.6)
Authors: Rush James , Denniss Steven , Smith Christopher , Jeffery Andrew , Ford Rebecca
Published: June 2021

The objective of this study was to determine the chronic and acute effects of resveratrol (RSV) on endothelial function and endothelium‐derived contracting factor (EDCF) signaling using common carotid artery (CCA) isolated from spontaneously hypertensive rats (SHR) and Wistar Kyoto rats (WKY). Acetylcholine (Ach)‐stimulated endothelium‐dependent relaxation in pre‐contracted CCA was impaired in SHR (max. 53±6% vs. 86±6% in WKY) caused by competitive cyclooxygenase (COX)‐mediated contraction at high [Ach]. Chronic (28‐day) treatment in vivo (via drinking water) with a moderate RSV dose (Mod‐RSV, ~0.075 mg/kg/day) was without effect on Ach‐stimulated relaxation, or Ach‐stimulated contraction and associated prostaglandin (PG) production in quiescent CCA, in either strain. In contrast, in SHR but not WKY, a chronic high RSV dose (High‐RSV, ~7.5 mg/kg/day) improved Ach‐stimulated relaxation (max. 99±5% vs. 53±6% in No‐RSV SHR), and attenuated Ach‐stimulated contraction (55±5% vs. 68±5% in No‐RSV SHR) and associated PG production (189±30 vs. 519±93 pg/ml in No‐RSV SHR) (all p<0.05). High‐RSV also reduced mean arterial pressure in SHR (159±6 vs. 176±6 mmHg in No‐RSV SHR; p<0.05). In separate experiments, acute (30 min) in vitro treatment (in‐bath pre‐incubation) with 20µM RSV (RSV‐20µM) nearly abolished Ach‐stimulated contraction and greatly attenuated the associated PG production in SHR and WKY CCA (all p<0.05). Taken altogether, these experiments demonstrate that RSV can reverse endothelial dysfunction caused by EDCF activity by a signaling mechanism predominately involving the attenuation of COX‐derived PG production.

Grant Funding Source: Supported by NSERC Canada RGPIN 238342
The Dahl S rat on normal salt diet is a spontaneous model of preeclampsia (857.5)
Authors: Gillis Ellen , Mooney Jennifer , Williams Jan , Garrett Michael , Sasser Jennifer
Published: June 2021

Preeclampsia (PE), increased BP and proteinuria during pregnancy, is detrimental to mother and fetus. There is currently no cure, and the development of new animal models is necessary to further research in this field. Here we test the hypothesis that the Dahl S rat, a known genetic model of hypertension and kidney disease, is also a spontaneous model of preeclampsia. Female Dahl S rats were implanted with a telemetry unit via the femoral artery. After a 10 day recovery period, baseline BP was recorded. Rats were placed in metabolic cages for 24 hr urine collection at baseline, mid and late pregnancy (days 10‐11, 17‐18), and urinary protein, 8‐isoprostane, and TBARS concentrations were measured via commercial assays as increased oxidative stress is also associated with preeclampsia. There was no difference in baseline BP (152±1.33 mmHg vs 151±4.2 mmHg) or proteinuria (61±9.6 mg/d vs 60±16.8 mg/d) in the pregnant rats vs virgin rats (n=5‐7). Pregnant rats exhibited increases in BP, proteinuria (Table), urinary 8‐isoprostane excretion (13.46±2.34 vs 3.14±0.52 ng/d*) and urinary TBARS excretion (224.43±31.00 vs 140.47±7.70 μmol/d, p=0.06) during late pregnancy, but no changes were observed in age‐matched virgin rats. In addition, the Dahl S rat has a smaller litter size compared to the Sprague Dawley (6.6±2.3 vs 14.4±1.9*) and exhibits intrauterine growth restriction as measured by pup weight on day 20 (1.86±0.10 vs 2.43±0.03 g*). These data suggest that the Dahl S rat has increased oxidative stress during pregnancy, possibly contributing to the exacerbation of hypertension and proteinuria during pregnancy in this strain.

Mean Arterial Pressure (mmHg)
Urinary Protein Excretion (mg/d)

Baseline
152±1
61±10

Mid‐Pregnancy
161±2*
149±26*

Late‐Pregnancy
160±2
159±48

*p<0.05 vs baseline
Impaired renal NCC function and expression: a mechanism driving norepinephrine evoked salt‐sensitive hypertension? (857.6)
Authors: Walsh Kathryn , Wainford Richard
Published: June 2021

Aim: These studies test the hypothesis that impaired renal NCC regulation is a mechanism underlying the development of norepinephrine (NE) evoked salt‐sensitive hypertension in Sprague‐Dawley (SD) rats.

Methods: SD rats receiving a s.c. saline vehicle, NE (600ng/min), DMSO/saline vehicle (50:50), or hydrochlorothiazide (HCTZ 4mg/kg/d) and NE infusion were fed a 0.4% (NS) or high 8% NaCl (HS) diet for 14 days. On day 14, MAP, MAP response to i.v. hexamethonium (30mg/kg) and peak natriuresis to i.v. HCTZ (2mg/kg) infusion was assessed and NCC immunoblotting was performed on kidney cortex tissue.

Results: NE significantly increased MAP and vascular sympathetic tone. HS exacerbated NE induced hypertension, induced up‐regulation of NCC protein levels and prevented dietary sodium stimulated reduction in HCTZ evoked natriuresis. HCTZ + NE co‐infusion abolished the salt‐sensitive component of NE induced hypertension.

s.c. saline vehicle
s.c. NE
s.c. DMSO/saline
s.c. NE + HCTZ

NS
HS
NS
HS
HS
HS

MAP (mmHg)
124±2
124±1
151±3Ï•
171±4*â‰
134±4
152±3â‰ ÏˆÏ„

Peak Î”MAP to hexamethonium (mmHg)
‐61±4
‐66±3
‐77±6Ï•
‐81±8â‰
‐57±4.5
‐69±3Ï„

Peak Î”UNaV to HCTZ (Î¼eq/min)
8.7±0.3
7.2±0.7*
11.1±1.1 Ï•
10.8±0.4â‰
6.3±1.3
0.4±0.2 â‰ ÏˆÏ„

NCC expression(ODU/mm2 normalized to GAPDH)
TBD
TBD
0.15±0.02
0.27±0.06*
n.d.
n.d.

*p<0.05 vs respective NS gp; ϕp<0.05 vs vehicle NS gp; ≠p<0.05 vs vehicle HS gp; ψp<0.05 vs DSMO vehicle; τp<0.05 vs NE + HS gp; n=4‐6/gp.

Conclusion: These findings reveal that the combination of high salt intake and excess NE evokes the development of salt‐sensitive hypertension which consequently results in impaired NCC function and increased protein expression. These results contrast the established down‐regulation of NCC expression and circulating NE levels observed during high salt‐intake in salt‐resistant rat phenotypes and provides a new insight into the mechanisms underlying the influence of sympathoexcitation on the kidney in hypertension.

Grant Funding Source: R01HL107330 & K02HL112718A1
Post‐injury alcohol exposure exacerbates neuroinflammation and impairs neurological recovery following traumatic brain injury (859.6)
Authors: Teng Sophie , Maxi John , Katz Paige , Gilpin Nicholas , Molina Patricia
Published: June 2021

BACKGROUND: There are ~1.7 million reported traumatic brain injury (TBI) cases in the U.S. annually. Alcohol abuse contributes to 36% to 51% of all TBI incidents, and patients continue to use alcohol after TBI in up to 26% of cases. Limited preclinical studies have examined the impact of post‐injury alcohol exposure on TBI recovery and the underlying mechanisms. The aim of this study was to determine the effect of post‐injury alcohol exposure on TBI outcomes in terms of neuropathological and neurological recovery.

METHODS: Male Sprague‐Dawley rats (~300 g) were fitted with a female Luer‐lock over a 5 mm left lateral craniotomy 1 week prior to injury. TBI was induced by lateral fluid percussion (~30 PSI, ~25 ms) under isoflurane anesthesia. At day 4 post‐injury, animals were exposed to either intermittent alcohol vapor (95% ethanol 14h on /10h off) or room air for 10 days. RESULTS: TBI produced immediate apnea (17±5 s) and delayed righting reflex (419±30 s). Neurological severity score (NSS) was increased as a result of injury prior to alcohol exposure (2‐fold; p<0.05). TBI/Air group showed progressive recovery with no significant impairment in NSS detected at 14 and 18 days post‐TBI. In contrast, alcohol exposure impaired recovery and had higher NSS than time‐matched air‐exposed animals throughout the 10 day period of alcohol exposure and following a 4 day abstinence period (3.5‐fold; p<0.05). Alcohol exposure alone increased NSS score after 10 days of vapor, but returned to baseline after 4 days of abstinence. Immunofluorescence revealed increased activation of astrocytes, microglia, and other markers of neuroinflammation in alcohol exposed brains after TBI.

CONCLUSIONS: These findings indicate that alcohol exposure impairs neurological recovery, which may be caused by sustained neuroinflammation. The clinical implications of long‐term consequences in TBI patients exposed to alcohol during recovery warrant further investigation.

Grant Funding Source: Supported by NIAAA‐AA7577, F30AA022838
Progesterone supplementation improves blood pressure and uterine artery resistance and hypoxia‐stimulated placental cytokines during pregnancy (860.16)
Authors: Amaral Lorena , Kipprono Luissa , Wallace Kedra , Frazier Krystal , Moseley Janae , Martin James , LaMarca Babbette
Published: June 2021

Preeclampsia (PE), new onset hypertension at 20 weeks of gestation, is characterized by increase uterine artery resistance (UARI), elevated TNF‐α, IL‐6, IL‐17, and sFlt‐1. We have demonstrated that PE women have significantly lower circulating progesterone (P) than normal pregnant (NP). Therefore, we hypothesized that progesterone (17‐OHP) supplementation could improve blood pressure and decrease UARI and inflammatory cytokines during PE. To address this question we examined the effect of intraperitoneally administered 17‐OHP (3.32mg/kg) on blood pressure (MAP) and UARI in the RUPP rat model of PE. In addition, we examined hypoxia‐stimulated cytokines secreted from placenta explants collected from NP subjects treated at UMMC via ELISA. MAP in NP rats (n=13) was 92+2; 123+2 in RUPP (n=18), and 116 +1 mmHg in RUPP+17‐OHP (n=10), p <0.05. UARI was 0.78+0.03 in RUPP (n=4) and 0.63+0.038 in RUPP+17‐OHP (n=8), p <0.05. Hypoxic placental TNF‐α, IL‐6, IL‐17, and sFlt‐1 was 2.9+1.0, 53 +27, 7.3+4, 1038+362 which was blunted with 1 uM progesterone to 1.8+7, 33+7, 5+1.5 and 623+133 pg/ml. In conclusion, progesterone supplementation improves UARI and hypertension and hypoxia‐stimulated cytokines during pregnancy.

Grant Funding Source: Supported by Obstetrics and Gynecology, University of Mississippi Medical Center
Effect of electrical field stimulation on spontaneous Ca2+ transients in rat pulmonary vein cardiomyocytes (864.4)
Authors: Henry Alasdair , Rankin Andrew , Rowan Edward , Drummond Robert
Published: June 2021

The pulmonary veins (PVs) are widely recognised as an important source of ectopic electrical activity that can lead to atrial fibrillation. While the ectopic activity most likely originates in the cardiomyocytes that form an external sleeve around the PVs, the underlying mechanism remains to be elucidated. Recently, changes in the intracellular Ca2+ concentration have been proposed to play a role. Studies were carried out on intact PVs isolated from the rat, using fluo‐4 to monitor intracellular Ca2+ within individual cardiomyocytes. Under basal unstimulated conditions, spontaneous Ca2+ transients were observed that were asynchronous in neighbouring cells. The frequency of spontaneous Ca2+ transients increased from 0.45 ± 0.06 Hz to 0.59 ± 0.05 Hz (P<0.001) immediately following electrical field stimulation at 5 Hz. Increasing the external Ca2+ concentration to 4.5 mM enhanced this effect, with the frequency of spontaneous Ca2+ transients increasing from 0.45 ± 0.05 Hz under basal conditions to 0.91 ± 0.07 Hz after electrical stimulation (P<0.001). Thus, although electrical field stimulation significantly increased the frequency of the spontaneous Ca2+ transients, their asynchronous nature would mean that they are unlikely to be capable of triggering an action potential. Other interventions that could lead to entrainment of the Ca2+ transients are currently being investigated.
Effect of lactate on intermediary metabolites expression and mitochondrial biogenesis in perfused hearts (864.5)
Authors: Gabriel‐Costa Daniele , Cunha Telma , Bechara Luiz Roberto , Barreto‐Chaves Maria Luiza , Brum Patricia
Published: June 2021

The aim of this study was to investigate whether lactate would modulates the expression of genes and proteins related to intermediary metabolism and mitochondrial biogenesis, and its impact on cardiac hemodynamic function. The hearts excised from male healthy rats were perfused with or without lactate (20mM) in Krebs‐Henseleit solution along 120 min in a Langendorff apparatus. Hemodynamic values including of perfusion pressure (PP) were acquired during the experiment. PPARδ, PGC1‐α, COXIV, MCT1, MCT4 and LDH mRNA levels were quantified by real time RT‐PCR. Mitochondrial DNA replication was evaluated, comparing COXII mRNA levels, a specific mitochondrial gene.Protein expression was assessed by Western Blot. The activity of hexokinase, lactate dehydrogenase, pyruvate dehydrogenase, citrate synthase, superoxide dismutase (SOD) and catalase were analyzed by colorimetry and spectrophotometry. Significant differences were accepted when P□0.05. The lactate concentration in the heart tissue was elevated in lactate group at 120 min (1.5 ± 1.2 vs. 4.8 ± 0.7 mM). This response was paralleled by a significant increase in mRNA levels of PPARδ (2.3x), PGC1‐α (4.5x), MCT1 (8.0x), MCT4 (4.0x) and LDH (13.7x) when compared to control group. No differences were observed in mitochondrial DNA replication, COX IV mRNA levels and activity of all enzymes, excepted for SOD that was significantly increased in lactate (26.6 ± 12.1 vs 92.3 ± 25.7 µmol/mg/min). PP value reduced after lactate perfusion (74.2 ± 10.3 vs 27.8 ± 6.8 mmHg). Our results provide evidence that lactate up‐regulates the expression of genes involved in cardiac bioenergetics and mitochondrial biogenesis, which might favor cardiac oxygenation.

Grant Funding Source: FAPESP
Early life exposure to air pollution induces adult cardiovascular dysfunction in mice (864.9)
Authors: Velten Markus , Gorr Matthew , Nelin Timothy , Wold Loren
Published: June 2021

The developing fetus is affected by exposure to air pollutants. We hypothesized that intrauterine and postnatal exposure to air pollutants results in cardiac dysfunction. FVB mice were exposed to ambient particles <2.5 μM (PM2.5) or filtered air (FA) beginning at plug confirmation till weaning. Echocardiography, PV‐loops, cardiomyocyte contractility, and molecular analyses were performed at 10 wks of age. Birth weight was lower in PM2.5 exposed pups (1.36 g ± 0.12 FA vs. 1.30 ± 0.16 PM2.5, P=0.03). Echocardiography revealed reduced FS (43.6 % ± 2.1 FA vs. 33.2 % ± 1.6 PM2.5, P=0.001) and PV‐loops a reduced EF (79.1 % ± 3.5 FA vs. 35.5 % ± 9.5 PM2.5, P=0.005), increased ESV (10.4 µl ± 2.5 FA vs. 39.5 µl ± 3.8 µl PM2.5, P=0.001), reduced dp/dt max (11605 ± 200 FA vs. 9569 ± 800 PM2.5, P=0.05) and min (‐9203 ± 235 FA vs. ‐7045 ± 189 PM2.5, P=0.0005) in PM2.5 exposed mice. Isolated cardiomyocytes had reduced peak shortening (%PS, 8.53 ± 2.82 FA vs. 6.82 ± 2.04 PM2.5, P= 0.003) and lengthened calcium reuptake (0.22 s ± 0.09 FA vs. 0.26 ± 0.07 PM2.5, P= 0.048), coupled with reduced contractile reserve. Histological analyses revealed lower PicroSirius‐red stained areas in the PM2.5 group (0.55 % ± 0.09 FA, 0.55 % ± 0.09 PM2.5, P=0.0003). Our study indicated that developmental exposure to PM2.5 results in a heart failure phenotype at adulthood. This study supports the hypothesis that perinatal insults contribute to cardiac disease during adulthood.

Grant Funding Source: Supported by NIH
Cardiac regenerative capacity of the American alligator (87.5)
Authors: Yang Johnny , Owerkowicz Tomasz , Elsey Ruth , Lien Ching‐Ling
Published: June 2021

Human hearts lack regenerative capacity and form scars after myocardial infarction. While teleost fish and urodele amphibians are able replace lost or damaged cardiomyocytes even as adults, mice are only able do so as neonates. Whether regenerative capacity exists in other vertebrate clades (reptiles and birds) has not yet been described. In order to determine if reptile hearts have regenerative potential following tissue damage, we developed injury models similar to those used in zebrafish and mice—ventricular resection and cryoinjury—and tested them in a non‐traditional animal model, the American alligator.

Hatchling alligators were injured or sham operated, and allowed to recover over periods of time up to 4 months. Preliminary data from hatchlings 1, 3, and 14 days post‐injury show appearance of regenerative cardiomyocytes around the injury area. Whether this is a significant increase over background proliferation of sham operated hearts, as well as the ability to replace scar tissue with cardiomyocytes, remains to be determined. Results from this study will demonstrate whether the alligator— with its four‐chambered heart, and independent systemic and pulmonary pressure generation—can be a useful model in studies of cardiac regeneration, and serve to further elucidate the phylogenetic and evolutionary relationship between the aforementioned clades with regard to regenerative capacity.

Grant Funding Source: Supported by California Institute for Regenerative Medicine
The effects of developmental nicotine exposure on hypoglossal motoneuron morphology and electrophysiology (871.2)
Authors: Powell Gregory , Levine Richard , Fregosi Ralph
Published: June 2021

Developmental nicotine exposure (DNE) is known to have significant effects on motoneuron electrophysiology and morphology. Previous work in hypoglossal motoneurons (XIIMN) of neonatal rats exposed to nicotine in utero has shown decreased excitatory synaptic input, desensitized nicotinic acetylcholine receptors, increased input resistance, a left‐shifted frequency‐current curve as well as increased gain, and preliminary evidence for a decreased soma size. To further examine the effects that DNE has on XIIMNs, we have used whole‐cell patch clamp techniques to record responses to injected sine‐wave currents while neurobiotin passively fills the neurons. Filled cells were visualized using a DAB reaction followed by a detailed morphological reconstruction using the Neurolucida system. Response to sinusoidal current injection was used to record spike firing resonance, which is indicative of spike‐timing precision and variability. These measurements are useful for examining potential changes of the input‐output efficiency of XIIMNs by DNE. Preliminary data indicate significantly increased size of dendritic surface area in DNE cells (n=8 control; 2 DNE). Nicotine exposed cells also appear to be less responsive to sine wave inputs, and more variable in their response as well (n=4 control; 7 DNE).

Grant Funding Source: Supported by NIH R01HD071302, AHA 12GRXIT120S0345