bluuebunny/crossref_metadata_2025_split huggingface dataset:
Impact of Sex and GPER on the Cardiovascular Effects of the Environmental Estrogen Bisphenol A
Authors: Spooner Hallie Marie , Lindsey Sarah H. 
Published: November 2023

We previously found that the membrane‐bound G‐protein coupled estrogen receptor (GPER) attenuates blood pressure. Environmental estrogens, such as Bisphenol A (BPA) compete for binding at GPER, potentially reducing its beneficial vascular effects. Recent clinical data shows that exposure to BPA is associated with higher blood pressure and cardiovascular disease. The goal of the current study was to assess the effects of BPA on blood pressure in a mouse model and determine the impact of sex and GPER. We hypothesized that BPA administration in female mice would induce a larger increase in blood pressure in comparison to males because of its interference with estrogen's protective vascular effects. Further, we hypothesized that the response to BPA in GPER knockout mice would be diminished. Female (f) and Male (m) wild type (WT) and knockout (KO) mice were administered 5 mg/kg body weight/day of BPA via the drinking water for eight weeks. Blood pressures were measured via tail cuff by volume pressure recording biweekly. Two‐way ANOVA revealed a significant effect of sex (P<0.001) but not GPER deletion (P=0.19) on systolic blood pressure, with females having lower pressures (mWT 148 ± 2.8 vs. fWT 131± 4, P<0.01; mKO 142±3 vs. fKO 128± 3, P<0.05). Diastolic pressures were not statistically different by sex (P=0.08) or GPER deletion (P=0.82). Although the study is ongoing, two weeks after the administration of BPA, there was no significant impact of BPA on blood pressure in any group (P>0.05). In conclusion, since GPER mediates some of the beneficial cardiovascular actions of estrogen, environmental estrogens such as Bisphenol A may act as a “hormonal block” of GPER. Increasing amounts of environmental estrogens may promote cardiovascular problems, especially in aging postmenopausal women.Support or Funding InformationTulane University Newcomb College Lurcy GrantAmerican Heart Association 15GRNT25890021
Electrically‐Induced Cardioprotection via the Skin
Authors: Roessler Anne E , Luther Kristin , Ren Xiaoping , Wang Yang , Shen Zilong , Jones Walter Keith 
Published: November 2023

Ischemic preconditioning (IPC) is a well‐characterized cardioprotective phenomenon involving multiple brief, non‐lethal ischemic episodes to the heart. Remote ischemic preconditioning (RIPC) elicits cardioprotection via repetitive ischemia and reperfusion to a distant vascular bed, organ, or limb. A non‐ischemic, nociceptive stimulus in distant tissues can result in the most powerful cardioprotection reported – an 80% decrease in infarct size – called remote nociceptor‐induced cardioprotection (NIC). Electrically‐induced cardioprotection via electroacupuncture with needles (EA) or electrical stimulation with patches (ES) on the skin of the abdomen may be a clinically feasible way to induce cardioprotection.Our previous work shows that the transcription factor NF‐κB regulates many cardioprotective genes in the heart after IPC, including heat shock proteins (HSP), which act in unity to assist in protein folding, repair, and degradation following myocardial injury. We found that IPC requires NF‐κB‐dependent upregulation and post‐transcriptional regulation of HSP70.3. This study's objective is to determine the molecular mechanism of electrically‐induced NIC. Based upon our work in IPC, we hypothesized that electrical stimulation of the skin is cardioprotective via NF‐κB‐dependent upregulation of HSPs.Our methodology uses a 5‐volt electrical stimulus applied to the abdomen via EA or ES. This reduces infarct size in a mouse surgical model of myocardial infarction when applied as either a preconditioning or postconditioning stimulus. Western blotting and PCR revealed only small non‐significant changes in HSP mRNA and protein relative to sham, inconsistent with the large significant increases in HSP seen in ischemic preconditioning. Whether these small increases in HSP mRNA and protein are functionally significant is under investigation using pharmacological and genetic inhibition of the NF‐κB‐dependent HSPs, and deep sequencing studies are planned to delineate additional potential genes that may be involved. Future investigations will also address the neurobiology of electrically‐induced cardioprotection via skin stimulation.Support or Funding Informationgrant number: R01HL091478
Method to Assess Interactivity of Drugs with Nonparallel Concentration‐Effect Relationships
Authors: Zhao Liang , Au Jessie L.‐S. , Wientjes M. Guillaume 
Published: November 2023

Commonly used methods for analyzing interactivity between drugs (e.g., synergy, antagonism) such as isobologram, combination index, and curve shift are based on the Loewe Additivity principle of dose equivalence and the inherent assumption of similar concentration‐effect (C‐E) including parallel curves and equal maximum effects (Emax), and therefore are not suitable for drugs with dissimilar C‐E. This study describes a new method that is without this limitation and has the additional advantage of enabling statistical analysis. The method comprises two steps. First, based on the dose equivalence principle, the experimentally obtained C‐E of one drug was used to calculate the equally effective C‐E of the other drug at no interactivity; the resulting two zero‐interactivity C‐E formed the upper and lower boundaries of Additivity Envelope. Next, 95% confidence intervals calculated from experimental data were added to Additivity Envelope to obtain Uncertainty Envelope (UE). Experimentally observed effects of drug combinations (C‐Ecomb,observed) located within UE indicate additivity whereas C‐Ecomb,observed located above or below UE indicate statistically significant (p<0.05) synergy or antagonism, respectively. Additional in silico studies demonstrated the shape and size of Additivity Envelope, which determine the ability to detect drug interactivity, depended on the Drug A‐to‐B concentration ratios and the ratios of their C‐E curve shape parameter. Analyses of experimental results of combinations of drugs with nonparallel C‐E and/or unequal Emax indicate UE was more versatile and provided more information, compared to earlier methods. UE is a broadly applicable method for analysis, including statistical significance assessment, of drug interactivity.Support or Funding InformationThis research is supported in part by the National Institutes of Health, National Institute of General Medical Sciences [Grant R01GM100487]; and National Cancer Institute [Grant R01CA163015].
Quantitative Pharmacology of Intercellular Drug Shuttle by Exosomes
Authors: Wang Jin , Cui Minjian , Yeung Bertrand Z. , Lu Ze , Wientjes Michael Guill , Woo Sukyung , Au Jessie L.‐S. 
Published: November 2023

BackgroundExosomes are extracellular vesicles (30–100 nm) that contain bioactive molecules, secreted by most cell types and play a role in intercellular communications. Tumor cells exosomes may affect tumor progression, metastasis, and therapeutic efficacy and resistance. The current study used in vitro experiments and in silico studies to investigate the roles of exosomes in intercellular drug transfer and the quantitative relationship between exosomes and pharmacodynamics of chemotherapy drugs in solid tumor cells.MethodsEffects of paclitaxel (PTX) and doxorubicin (DOX) on exocytosis were studied in human breast (MCF7 and a lung metastatic subline of MDA‐MB‐231 cells, LM2) and ovarian (A2780 and OVCAR4) cancer cells. Inhibition of exocytosis was studied using omeprazole (proton pump inhibitor) and GW4869 (ceramide synthesis inhibitor). Time‐lapse laser confocal microscopy was used together with RFP‐transfected cells to monitor the release of exosomes into extracellular fluid. Extracellular exosomes were collected and quantified based on their acetylcholinesterase activity. Drug concentrations were measured using LC‐MS/MS. Pharmacodynamics of exosomes were measured in recipient cells, using the sulforhodamine B assay for anti‐proliferation and the wound‐healing assay for anti‐migration.ResultsAll four cells in exponential growth phase maintained similar baseline exosome level (~220 exosomes/cell). PTX (2–1000 nM) or DOX (10–1000 nM) treatment for 24 or 48 h stimulated exosome production/secretion, in dose‐ and time‐dependent manner, by up to 10‐fold. After 24 h treatment, the amount of PTX in exosomes was 5–10% of that in donor cells, and these PTX‐containing exosomes inhibited cell viability and migration in vitro. Pretreatment for 24 h with omeprazole (10 μM) or GW4869 (10 μg/mL) reduced the production/secretion of, and the drug amount in, exosomes by >20%, and enhanced the exosome cytotoxicity in donor cells. Our quantitative pharmacology model, comprising 6 intracellular and intercellular compartments and 11 rate constants, captured and characterized the effect of exocytosis on intercellular transfer of drug‐containing exosomes and PTX pharmacodynamics in donor and recipient cells with good fitness (CV <19%).ConclusionOur results indicate (a) chemotherapeutic agents stimulate exosome production/secretion, (b) exocytosis is a mechanism of intercellular drug transfer responsible for conferring biological activities on neighboring recipient cells, and (c) successful application of quantitative pharmacology to depict the spatial and temporal effects of exosomes on chemotherapy pharmacodynamics.Support or Funding InformationSupported in part by RO1CA163015 (GW, JA) from National Cancer Institute and RO1EB015253 (JA, GW) from National Institute of Biomedical Imaging and Bioengineering, DHHS and RSG‐16‐006‐01‐CCE (SW) from American Cancer Society.
Influence of fan induced photoperiod on selected blood physiological variables of male broilers reared to 63 days of age
Authors: Olanrewaju Hammed A , Purswell Joseph L , Collier Stephanie D , Branton Scott L 
Published: November 2023

Increasing broiler house size and ventilation capacity have resulted in increased light ingress through ventilation system components. Effectively of this may alter the photoperiod for broiler chickens and create local increases in light intensity, which may also impact broiler body homeostasis. The objective of this study was to determine the influence of fan‐induced photoperiod on selected blood plasma biochemistry, enzyme activities and electrolyte levels of male broilers reared to 63 day of age. A total of 960 male broiler chicks were randomly distributed to 16 environmentally‐controlled rooms at 60 chicks/room. Similar feed and water were provided ad libitum to all chickens. The lighting program to day 35 was identical for both treatments and treatments were initiated at day 36. The two treatments consisted of constant (C) light intensity at 2.5 lx at 16L: 8D based on prescriptive intensities in typical heavy broiler lighting programs and a variable (V) light intensity and photoperiod program that was based on field measurements near tunnel fans in a solid‐walled broiler house. The study was a completely randomized design with room serving as the experimental unit. Blood samples were collected from the wing brachial vein of 6 birds per room on day 35 (before initiation of the treatments) and also on day 63 and analyzed immediately. The results showed broilers exposed to a constant light intensity of 2.5 lx as compared with broilers exposed to variable lighting to mimic areas near tunnel fans had significantly lower level of pH, Ca2+, K+, and higher levels of angap. However, all these changes were still within the acid‐base homeostasis and physiological ranges. Also, blood glucose and plasma corticosterone concentrations were not affected by treatments, suggesting an absence of physiological stress. It was concluded that the data herein illustrate the need to mitigate light ingress through ventilation system components to improve live performance in broilers without compromising welfare.Support or Funding InformationUSDA/ARS
Laminar Shear Stress Reduces NADPH Oxidase II Expression but Does Not Influence Superoxide Dismutase Expression in Human Adipose Microvascular Endothelial Cells
Authors: Robinson Austin T , Cook Marc D , Brown Michael D , Phillips Shane A 
Published: November 2023

We have shown regular aerobic exercise reduces the expression of the pro‐oxidant enzyme NADPH Oxidase II (NOX II) and increases the expression of the anti‐oxidant enzyme superoxide dismutase (SOD) in ex vivo adipose resistance arteries. These adaptations contributed to preserved endothelial function following high intraluminal pressure‐ which elicits excess oxidative stress via the local renin angiotensin system. The purpose of this study was to determine if the primary mediator of exercise‐induced adaptations in NOX II and SOD in the adipose microcirculation is high physiological laminar shear stress (LSS).MethodsHuman adipose microvascular endothelial cells (HAMECs) were grown to 80–90% confluence and either 1) kept in the static condition; or 2) subjected to LSS (20 dyn/cm2) for 24 hours before cell harvesting. LSS was generated using a modified cone and plate design. Western blot was used to measure NOX II subunit and SOD isoform expression. In order to elucidate the role of Angiotensin II induced oxidative stress, HAMECs were either 1) untreated (control); or 2) treated with Ang II (400nm), and incubated with the superoxide (O2−) indicator DHE (5μM)‐with or without the Angiotensin Type I receptor blocker Losartan (50μM), the NOX II inhibitors VAS2870 (2μM) and NSC23766 (10 μM), or the SOD mimetic Tiron (100 mM).ResultsHAMECs exposed to LSS expressed significantly less of the NOX II subunits gp91phox and p47phox than static HAMECs. In contrast, there was no difference between static and LSS treated HAMECs in regards to SOD isoform expression. HAMECs exposed to Ang II produced significantly more O2− compared to control HAMECs. Pharmacological treatment with Losartan, VAS2870, NSC23766, or Tiron prevented the increased O2− production indicating NOX II and SOD play a role in mediating Ang II induced oxidative stress.ConclusionLaminar shear stress results in decreased NOX II, but not increased SOD expression in the adipose microvasculature. Both of these enzymes play a role in regulating Ang II induced O2− generation, thus shear stress may play a role in reducing Ang II mediated oxidative stress.Support or Funding InformationK23HL085614 (SAP), R01HL095701 (SAP), R01HL095701‐03S1 (support for ATR)
Spironolactone can act on pannexin1 channels in smooth muscle cells to regulate vasoconstriction and blood pressure
Authors: Good Miranda Elizabeth , Poon Ivan KH , Chiu Yu‐Hsin , Butcher Joshua T , DeLalio Leon J , Mendu Suresh , Jaffe Iris Z , Leitinger Norbert , Desai Bimal N , Bayliss Douglas A , Ravichandran Kodi S , Isakson Brant E 
Published: November 2023

Using an unbiased screen for potential inhibitors of pannexin1 (Panx1) channels, we identified spironolactone, a potent anti‐hypertensive, as being able to block Panx1 channels. This was especially interesting considering spironolactone has been considered a mineralcorticoid receptor (MR) antagonist. Initial confirmation of this result was obtained using electrophysiology, which showed that spironolactone could directly and effectively inhibit Panx1 channel activity without inhibiting other putative ATP release channels. Next, because we have previously demonstrated that Panx1 channels on smooth muscle cells (SMCs) can regulate α‐adrenergic vasoconstriction and blood pressure, we hypothesized that spironolactone would also inhibit α‐adrenergic vasoconstriction. Indeed, we found that spironolactone inhibited α‐adrenergic vasoconstriction from resistance arteries isolated from hypertensive humans. This effect was also seen in C57Bl6 mice, mice lacking SMC MR, or mice lacking Panx1 in endothelial cells. However, mice with SMC specific deletion of Panx1 (SMC‐Panx1 deficient) did not show any change in α‐adrenergic vasoconstriction when treated with spironolactone. TO‐PRO dye uptake on intact arteries further confirmed these functional results. Using radiotelemetry, we found that C57Bl6 mice and SMC MR deficient mice injected with spironolactone had in a significant drop in blood pressure acutely. In line with the vasoreactivity experiments, SMC‐Panx1 deficient mice did not have any change in blood pressure. Together these data suggest that Panx1 is a novel target of spironolactone and inhibition of Panx1 by spironolactone may contribute to this drug's anti‐hypertensive effects.Support or Funding InformationFunding: HL088554, HL120840, HL007284, HL131399.
Determining eNOS Binding and Circulation Modulating Effects of an Alpha Globin Mimetic Peptide (HbαX)
Authors: Keller TC Stevenson , Butcher Joshua T , Swope Nicole K , Columbus Linda , Isakson Brant E 
Published: November 2023

The ability of hemoglobin to scavenge the potent vasodilator nitric oxide (NO) in the blood has been well established as a mechanism of vascular tone homeostasis. In endothelial cells, the alpha chain of hemoglobin (hereafter, alpha globin) and endothelial NO synthase (eNOS) form a macromolecular complex, providing a sink for NO directly adjacent to the production source. We have developed an alpha globin mimetic peptide (named HbαX) that displaces endogenous alpha globin and increases bioavailable NO for vasodilation. Initially we performed structural analysis of HbαX via solution NMR in order to better understand the interaction of alpha globin and eNOS. Next we show that, in vivo, HbαX administration increases capillary oxygenation and blood flow in arterioles acutely and produces a sustained decrease in systolic blood pressure in normal and angiotensin II–induced hypertensive states. HbαX acts with high specificity and affinity to endothelial NO synthase, without toxicity to liver and kidney and no effect on p50 of O2 binding in red blood cells. In human vasculature, HbαX blunts vasoconstrictive response to cumulative doses of phenylephrine, a potent constricting agent. By binding to endothelial NO synthase and displacing endogenous alpha globin, HbαX modulates important metrics of vascular function, increasing vasodilation and flow in the resistance vasculature.Support or Funding InformationFunding: HL088554, HL120840.
Adipose Stromal Vascular Fraction Cells Improve Coronary and Cardiac Physiology in the Aged Female Heart
Authors: Beare Jason , Shofner Charles , Dale Jacob , Tinney Joseph , Keller Brad , Hoying James , LeBlanc Amanda 
Published: November 2023

Over 50% of adult women presenting with symptoms of ischemic heart disease have “clear” coronary artery angiograms. Additional testing indicates coronary microvascular dysfunction, contributing to the coronary perfusion deficits in these women. Current therapies for this condition, called microvascular angina, are relatively ineffective. Our previous research demonstrated that tissue‐resident macrophages within the therapeutic cell preparation derived from the stromal vascular fraction (SVF) of adipose tissue improved peripheral small artery function. Delivered intravascularly, these SVF‐derived macrophages migrate to the walls of small vessels and reset vasomotor tone. Therefore, we hypothesized that i.v. delivery of SVF cells can reverse coronary microvascular dysfunction. Using the aged female rat heart (22 mos. of age) as a model of coronary microvascular dysfunction without coronary artery involvement, combined with high‐resolution ultrasound and telemetry, we evaluated cardiac function and coronary flow reserve (CFR) in aged animals receiving fluorescently tagged GFP+ SVF cells from syngeneic young rats (6 × 106 cells/rat) via the tail vein at one‐week and four‐weeks post‐injection. At the time of explant, the location of injected SVF cells was determined via confocal microscopy, and isolated coronary arteriole preparations were used to evaluate vasoactivity. Injected GFP+ SVF cells had incorporated into the coronary vasculature one‐week post‐injection and remained there at 4 weeks. While coronary blood flow was improved in animals receiving the cell therapy, vasoreactivity of isolated arterioles to flow, pressure, bradykinin, and endothelin was unaffected. Measures of heart function such as maximal cardiac output, percent heart rate increase, and left ventricular diameter and volume during diastole ‐ which are all compromised in aged animals ‐ were significantly improved in response to a dobutamine challenge compared to pre‐SVF measurements. The cell therapy did not lead to arrhythmias or increased mortality. Our findings indicate that intravenously‐delivered adipose SVF cells disseminate to the aging heart and incorporate into the microvascular wall and perivascular spaces of the coronary vasculature. The presence of the therapeutic cells in the heart is associated with improved coronary perfusion and concomitant cardiac function.Support or Funding InformationJewish Heritage Fund for Excellence
Hypoxia Disturbs the Metformin‐Evoked Curtailment of Chemokine CX3CL1 (Fractalkine) Signaling in Human Umbilical Vein Endothelial Cells (HUVECs)
Authors: Szukiewicz Dariusz , Pyzlak Michal , Szewczyk Grzegorz , Wejman Jaroslaw , Kochanowski Jan 
Published: November 2023

Metformin (dimethylbiguanid) is an antihyperglycemic agent widely used for treating non‐insulin‐dependent diabetes mellitus (NIDDM). Moreover, it was reported that metformin may prevent nuclear factor kappa‐beta (NF‐κβ) activation in endothelial cells exposed to inflammatory cytokines. These anti‐inflammatory properties resulting from activation of AMP‐activated protein kinase (AMPK). Inflammation and hypoxia activate chemokine CX3CL1 receptor (CX3CR1)‐related signaling pathway. Thus, we hypothesized that lowering of cellular AMP levels during hypoxia may reduce ability of metformin to counteract the CX3CXL1‐CX3CR1‐axis‐dependent mechanism of endothelial dysfunction. In this study we examined comparatively the effect of metformin on lipopolysaccharide (LPS)‐induced CX3CL1 production as well as CX3CR1 expression in HUVECs cultured under normoxic vs. hypoxic conditions. Immediately after childbirth, HUVECs were isolated from a 12‐cm umbilical cord segment via collagenase digestion. The material obtained from primigravidas after term, normal‐course pregnancies was equally divided into two groups: group I (N = 24) – kept under hypoxia (5% O2), and the control normoxic (20% O2) group II (N = 24). After achieving the confluence, HUVECs were exposed to metformin (10 mmol/L; the subgroups IA and IIA) or cultured without metformin (the subgroups IB and IIB, respectively). Following pretreatment with LPS (1 μg/mL), the mean levels of CX3CL1 in the culture media supernatants were determined at predefined time points by ELISA. At the end of the culture period, HUVECs were formalin fixed, paraffin embedded, immunostained for CX3CR1 and cut into 5 μm sections. The expression of CX3CR1 was estimated by quantitative immunohistochemistry. The mean levels of CX3CL1 in group I were 5.9‐fold and 2.7‐fold higher than observed in normoxic group II (IA vs. IIA and IB vs. IIB, respectively). Initially, the mean levels of CX3CL1 were significantly (p < 0.05) lower in IA compared to IB, but these differences gradually disappeared, being non‐significant over the time. The mean expression of CX3CR1 in hypoxia did not differ statistically between IA and IB, but was significantly (p < 0.05) upregulated compared to the both normoxic subgroups IIA and IIB. In normoxia, exposition to metformin produced significant and stable decrease in CX3CL1 production in HUVECs, compared to metformin‐free subgroup IIB. A significant increase in CX3CR1 expression was observed in IIB compared to IIA. In conclusion, upregulation of CX3CR1 in response to hypoxia and/or metformin may serve as a compensatory mechanism to preserve or augment the level of CX3CL1/CX3CR1 signaling. Ineffectiveness of metformin in lowering of CX3CL1 production under hypoxia may suggest insufficient activation of AMPK in HUVECs.Support or Funding InformationWUM grant: 2M2‐W1‐16