bluuebunny/crossref_metadata_2025 huggingface dataset:
Aβ transportation across blood brain barrier is mediated by two β‐strands of RAGE (651.5)
Authors: Kim Sun‐Jick , Ahna Jang‐Won , Kim Young Ho , Choia Cheol Yong
Published: June 2021

Receptors for advanced glycation end‐products (RAGE) is transmembrane receptor of the immunoglobulin superfamily with an extracellular domain that consists of a variable region as well as two constant regions. RAGE and its ligand interaction are often referred to result in pro‐inflamatory gene activation and effect in diabetic complications, Alzheimer's disease and even some tumors. V‐domain of RAGE is determinant for binding with Amyloid‐β (Aβ). RAGE‐mediated Aβ transport of circulating Aβ across blood brain barrier (BBB) is important in pathogenesis of cerebrovascular β‐amyloidosis. In this study, we show that two β‐strands of RAGE are required for interaction with Aβ. Serial deletion analysis and site‐directed mutagenesis of RAGE V domain show that the third and eighth β‐strands are important for interaction with Aβ. Wild‐type RAGE activates the NF‐κB signaling pathway in response to Aβ peptide treatment, while a RAGE mutant defective in Aβ binding does not. Furthermore, using peptide of third and eighth β‐strand of RAGE or monoclonal antibody targeting Aβ interaction interface of RAGE V domain, interaction of RAGE and Aβ is abolished and transportation of Aβ peptide across the blood brain barrier is blocked. These results provide information crucial to the development of RAGE‐derived therapeutic reagents for Alzheimer disease.
A2B adenosine receptors promote retention of cardiac stem cells in a myofibroblast‐like state induced by myocardial infarction (652.11)
Authors: Ryzhov Sergey , Sung Bong Hwan , Zhang Qinkun , Weaver Alissa , Gumina Richard , Biaggioni Italo , Feoktistov Igor
Published: June 2021

Adult cardiac stem cells can contribute to scar formation by acquiring myofibroblast phenotype in response to myocardial injury. Adenosine levels increase in ischemic hearts and modulate cardiac cell functions via binding to adenosine receptors (AR). Here, we tested the hypothesis that A2BAR regulate a reversible transition of cardiac Sca1+CD31‐ mesenchymal stem cells into a myofibroblast‐like state in a mouse model of myocardial infarction (MI). Using flow cytometry, we found that MI resulted in a rapid increase in the numbers of alpha smooth muscle actin (αSMA)‐positive Sca‐1+CD31‐ cells by day 5, followed by gradual decline. Genetic ablation of A2BAR had no effect on accumulation of αSMA‐positive Sca‐1+CD31‐ cells. However, their numbers remained significantly higher in wild‐type (WT) compared with A2BAR knockout (KO) hearts during their deactivation (by 1.2 and 2‐fold on post‐MI days 7 and 14, respectively; p<0.05) indicating that this process is modulated by A2BAR signaling. Furthermore, the numbers of αSMA/collagen I double positive Sca‐1+CD31‐ cells remained higher in WT compared with A2BAR KO hearts by 73%±10% and 54%±6% on post‐MI days 7 and 14, respectively. Our data suggest that A2BAR signaling can play a role in scar evolution by promoting retention of Sca‐1+CD31‐ cells in their myofibroblast‐like state.

Grant Funding Source: HL95787‐01A1:04
Differential temporal regulation of cardiac cytokine expression in response to chronic β‐adrenergic receptor stimulation (652.4)
Authors: Grisanti Laurel , Repas Ashley , Carter Rhonda , Talarico Jennifer , Gold Jessica , Koch Walter , Tilley Douglas
Published: June 2021

β‐adrenergic receptors (βAR) are critical regulators of cardiac function normally and during heart failure (HF). While alterations in cytokines contribute to HF pathogenesis, and βAR have been demonstrated to regulate cytokines in different models of HF, a comprehensive understanding of this relationship is lacking. Thus, we sought to characterize the impact of chronic βAR signaling on cardiac cytokine expression in vivo. C57BL/6 mice underwent infusion with isoproterenol (Iso; 3 mg/kg/day) via minipumps for 1 or 2 weeks, versus vehicle. Cardiac function was monitored by echocardiography and hearts were excised at study termination for biochemistry and immunohistology, which showed Iso‐induced hypertrophy, fibrosis and apoptosis. Expression of cardiac transcripts were assessed via whole transcriptome analysis, where 780 and 689 transcripts were significantly altered at 1 and 2 weeks of Iso, respectively, with only 115 transcripts regulated similarly between the two cohorts. Protein expression of 53 cytokines were assessed, revealing significant alterations in 42, including distinct changes in expression at the 1 versus 2 week timepoints. Differential alteration in cytokine expression following 1 versus 2 weeks of chronic Iso stimulation suggests distinct consequences of chronic βAR signaling on cytokine‐mediated processes at different stages in the development of HF.
Biphasic mechanisms of amphetamine action at the dopamine terminal (661.4)
Authors: Siciliano Cody , Calipari Erin , Ferris Mark , Chen Rong , Jones Sara
Published: June 2021

In light of recent studies suggesting that amphetamine (AMPH) increases spontaneous and electrically evoked dopamine release ([DA]o), we examined discrepancies between these findings and previous literature which has demonstrated marked decreases in [DA]o after AMPH. The current study has refined and expanded the mechanisms of AMPH actions by defining two mechanisms of AMPH effects at high and low doses, one dopamine transporter (DAT)‐independent and one DAT‐dependent, respectively. Using voltammetry in brain slices, we examined the effects of low, moderate, and high concentrations of AMPH across a range of tonic and phasic stimulation parameters (1 pulse; 5 pulse 20 Hz; 24 pulse 60 Hz) in wild‐type (WT) and DAT knockout (KO) mice. We discovered novel, biphasic, concentration‐dependent effects in WT mice, where AMPH increased [DA]o at low concentrations and decreased [DA]o at high concentrations. We then used DAT‐KO animals to examine the mechanisms by which AMPH exerts these effects. In DAT‐KO slices, [DA]o was decreased by all concentrations of AMPH, demonstrating that AMPH‐induced increases in [DA]o are entirely DAT‐dependent, while the decreases at high concentrations are DAT‐independent. We propose that when AMPH concentrations are low, they are insufficient to deplete dopamine vesicles, and therefore AMPH acts solely as a DAT blocker, increasing peak [DA]o. When AMPH concentrations are high, the added mechanism of vesicular depletion leads to reduced [DA]o. The biphasic mechanisms observed here support and extend the established actions of AMPH.

Grant Funding Source: Supported by NIH grants DA021325, DA030161, DA006634, DA007246, DA031533, DA031791 and AA007565
Use of geranylgeraniol to rescue osteoblasts and periodontal ligament fibroblasts from zoledronate‐induced apoptosis (663.16)
Authors: Williams Calli , Johnson Brooke , Svrcina Haley , Kawsky Ben , Jones Dan
Published: June 2021

Bisphosphonate treatments can produce an unwanted side effect of bisphosphonate‐related osteonecrosis of the jaw (BRONJ). One of the most potent bisphosphonates, zolendronate (ZOL), is used to treat postmenopausal osteoporosis, Paget’s disease, and cancer metastasis to bone. Individuals affected with BRONJ are unable to heal from dental procedures, leading to painful oral lesions. The goal of the work presented here was to investigate the ability of GGOH to rescue both murine MC3T3E1 osteoblasts and human periodontal ligament fibroblasts from ZOL‐induced apoptosis. Apoptosis was measured using the APOPercentage® assay. In osteoblasts, 72 hours of 50 micromolar ZOL treatment induced apoptosis by 12.5‐fold but simultaneous exposure to 10 micromolar GGOH decreased apoptosis by 64% (p<0.05). In a separate set of experiments, a booster of GGOH was administered 24 hours after the initiation of treatment, dramatically reducing ZOL‐induced apoptosis by 98% (p<0.0001). In periodontal ligament fibroblasts, ZOL treatment induced apoptosis 10‐fold and GGOH reduced apoptosis by 82% (p<0.05). The booster of GGOH reduced ZOL‐induced apoptosis by 99%. Collectively the data demonstrates the ability of GGOH to potentially rescue both hard and soft tissues of the oral cavity from ZOL‐induced apoptosis with a supplemental dosage of GGOH 24 hours following initiation of treatment essentially eliminating apoptosis.

Grant Funding Source: Supported by grants from the Indiana Academy of Science and the Hodson Summer Research Institute
Loss of lymphocyte‐specific adaptor protein, LNK, aggravates endothelial dysfunction in angiotensin II‐induced hypertension (669.9)
Authors: McMaster William , Saleh Mohamed , Itani Hana , Harrison David , Madhur Meenakshi
Published: June 2021

LNK, a lymphocyte‐specific adaptor protein, is a negative regulator of hematopoiesis and endothelial cell signaling. LNK reduces cytokine production and cellular proliferation. Recent genome wide association studies have identified a non‐synonymous single nucleotide polymorphism in LNK that is associated with human hypertension (HTN). We previously found that in mice, loss of LNK is associated with a marked increase in blood pressure and enhanced aortic infiltration of T cells in response to angiotensin II (AngII) infusion. We hypothesized that this HTN is due in part to altered vascular reactivity. LNK‐/‐ and wild type (WT) mice were treated with AngII or vehicle for two weeks and vascular reactivity was assessed. Aortic rings were pre‐constricted with phenylephrine and subsequent relaxations to acetylcholine (Ach) and nitroglycerin (NTG) were examined. Interestingly, relaxation responses to Ach in LNK‐/‐ mice were shifted to the right, indicating impairment of endothelial‐dependent relaxation. The responses to NTG were not significantly altered between LNK‐/‐ and WT aortic rings. Therefore, loss of LNK aggravates endothelial function in the setting of hypertension and may provide one mechanism by which LNK contributes to hypertension and the associated end organ damage.

WT Sham
WT AngII
LNK ‐/‐ Sham
LNK ‐/‐ AngII

Ach‐Log(EC50)
6.87±0.12
6.60±0.10
6.33±0.11
5.88±0.10

LNK ‐/‐ vs WT p = 0.01; Ang vs Sham p = < 0.0001 Data are mean ± SEM (n = 4‐6)
Cardiac hypertrophy leads to coronary microvascular dysfunction and enhanced endothelin‐1/Rho‐kinase signaling in mice with transverse aortic coarctation (676.1)
Authors: Tsai Shu‐Huai , Lu Guangrong , Hein Travis , Kuo Lih
Published: June 2021

Hypertensive cardiac hypertrophy induced by transverse aortic coarctation (TAC) has been reported to reduce coronary flow reserve in mice, but its impact on oxygen delivery and microvascular function remains unknown. In the present study, we found that induction of TAC in mice for 3‐4 weeks increased heart‐to‐body weight ratio by 55% and decreased cardiac fractional shortening and left coronary arterial blood velocity (LCABV) by 51% and 21%, respectively. The myocardial lactate level was 43% higher in TAC than sham control mice. Isoproterenol (5 µg/g, i.p.) increased heart rate by 20% but the increased LCABV (23%) was observed only in the sham mice. In TAC mice, the superoxide levels in myocardium and coronary arterioles were increased by 1.7‐ and 2.3‐fold, respectively, and the myocardial endothelin‐1 (ET‐1) was increased by 59%. The endothelium‐dependent, nitric oxide (NO)‐mediated vasodilations to acetylcholine and adenosine, but not endothelium‐independent vasodilations to NO donor sodium nitroprusside, were impaired in TAC mice. The Rho kinase (ROCK)‐dependent vasoconstriction to ET‐1 was enhanced and the expressions of ROCK‐1 and ‐2 were increased in coronary arterioles of TAC mice. We conclude that myocardial pressure overload leads to cardiac deficiency, coronary microvascular dysfunction and ischemia possibly due to oxidative stress, elevated ET‐1/ROCK signaling and compromised NO function.

Grant Funding Source: Supported by Kruse Endowed Research Fund from Baylor Scott & White Healthcare
Sympathetic reactivity in young women with a family history of hypertension (680.11)
Authors: Greaney Jody , Kaiser Christopher , Matthews Evan , Wenner Megan
Published: June 2021

Young adults with a family history of hypertension (+FH) have an increased risk of developing hypertension. Further, the blood pressure (BP) response to sympatho‐excitatory stimuli in young adults can predict the future development of hypertension. Therefore, we hypothesized young women +FH (n=7, 22±1 yrs, 21±1 kg/m2, 81±2 mmHg resting mean BP) will have exaggerated sympathetic reactivity compared to young women without a family history of hypertension (‐FH; n=9, 23±2 yrs, 22±1 kg/m2, 80±2 mmHg resting mean BP). BP and muscle sympathetic nerve activity (MSNA) were measured during three sympatho‐excitatory maneuvers: 2 min of isometric handgrip exercise at 30% of maximal voluntary contraction (EX), 3 min of post‐exercise ischemia (PEI), and a cold pressor test (CPT). The increase in BP was greater in +FH during the last minute of EX (ΔMean BP: +FH 19±3 vs. ‐FH 11±2 mmHg, P=0.03) and during PEI (ΔMean BP: +FH 14±3 vs. ‐FH 6±1 mmHg, P=0.03). The increase in MSNA was greater in +FH during EX (ΔMSNA: +FH 15±4 vs. ‐FH 8±2 burst/min, P=0.13) and PEI (ΔMSNA: +FH 15±2 vs. ‐FH 5±2 burst/min, P<0.01). In a subset of women (+FH n=6, ‐FH n=5), +FH tended to have a greater increase in BP during CPT (ΔMean BP: +FH 12±2 vs. ‐FH 8±2 mmHg, P=0.21), but ΔMSNA was not different (P>0.05). These preliminary data suggest that +FH women may have greater sympathetic reactivity compared to ‐FH women.
Protein kinase C alpha is a target for the meprin B metalloproteinase (690.14)
Authors: Boyd Sada , Newman Robert , Ongeri Elimelda
Published: June 2021

Meprin metalloproteinases have been shown to play a role in the progression of diabetic nephropathy (DN) in both mice and humans. However, the mechanisms involved are not fully understood. The objective of this study was to determine whether PKC, a serine/threonine kinase known to play a role in DN, is a meprin substrate. Activated forms of purified recombinant meprin A and B were incubated for 0‐4 h with cytosolic‐enriched proteins from meprin αβ double knockout mouse kidneys (which lack endogenous meprins) and protein lysates extracted from Mardin‐Darby canine kidney (MDCK) cells (which do not express meprins). The levels of PKCα were determined by Western blot analysis using anti‐PKCα specific antibodies coupled with optic densitometry. Incubation with meprin B significantly reduced the levels of PKCα present in the kidney proteins and MDCK lysates. This decrease was not observed in proteins incubated with meprin A or control reactions without meprins, suggesting isoform‐specific degradation of PKCα. To confirm the degradation, human PKCα proteins were expressed in yeast, purified by GST‐affinity chromatography, and incubated with the activated meprins. These data suggest that meprin B may impact kidney function via proteolysis of PKCα. Consistently, PKC isoforms are upregulated in DN and PKCα has been shown to play a role in the development of albuminuria in mice with streptozotocin‐induced type 1 diabetes.

Grant Funding Source: Supported by NIGMS (SC3GM102049)
Sustained Resveratrol infusion increases renal blood flow without changing glomerular filtration rate in normal rat (692.1)
Authors: Gordish Kevin , Beierwaltes Wiliam
Published: June 2021

The polyphenol resveratrol has cardiovascular benefits, but it’s effect on renal hemodynamics and glomerular filtration rate (GFR) remain unknown. Resveratrol vasodilates by increasing nitric oxide (NO), may inhibit reactive oxygen species (ROS), and prevent ROS depletion of NO. We found an acute bolus of resveratrol transiently increased renal blood flow (RBF) and decreased renal vascular resistance (RVR). We hypothesized a sustained infusion of resveratrol may increase RBF, and therefore GFR. We infused vehicle for 30 minutes followed by resveratrol infusion (30 minutes) at of 0, 0.5, or 1.0 mg/min. We measured RBF, calculated RVR and measured GFR with FITC‐inulin. 1.0 mg/min resveratrol infusion increased RBF by 15%, from 7.17±0.40 to 8.25±0.40 ml/min/gkw (p<0.01, n=7). RVR decreased from 13.61±0.85 to 11.81±1.01 ARU (p<0.04). GFR remained unchanged (1.61±0.17 vs. 1.68±0.20 mg/min/gkw, respectively). In separate experiments, control RBF was unchanged with 0.5 mg/min resveratrol (7.69±0.84 vs. 7.88±0.94 ml/min/gkw, n=9). RVR was also unaffected. Control GFR was 0.98±13 ml/min/gkw remained unchanged by resveratrol (1.12±0.22 ml/min/gkw). We conclude that a constant infusion of resveratrol induced a significant increase in RBF and decreased RVR while not changing GFR, suggesting uniform dilator effects on both afferent and efferent resistance vessels.

Grant Funding Source: supported by NIH 5P01HL090550‐04
Increased oxidative damage in the plasma of adult type 2 diabetics with poor glycemic control (693.7)
Authors: Platat Carine , Habib Hosam , Souka Usama , AlMaqbali Fatma , Kamal Hina , Ibrahim Wissam , Baynouna Latifa , Ali Habiba
Published: June 2021

Increased oxidative damage in the plasma of adult Type 2 diabetics with poor glycemic control Increasing evidence associates oxidative stress with the pathogenesis of Type 2 Diabetes (T2DM). Nonetheless, the available data related to oxidative stress and its relationships with glycemic status among T2DM patients are scarce and contradictory.

Our goal was to investigate the relationships of metabolic control of T2DM with oxidative damage and antioxidant status.

T2DM Emirati adults (n=187), participating in the Skills for Change randomized intervention trial and recruited from seven different ambulatory health care centers in Al Ain, United Arab Emirates, were considered. 24 h‐recall was collected. Anthropometry and HbA1c were assessed. Aantioxidant status markers were measured: Vitamin C, ABTS, Ferric Reducing Ability of Plasma (FRAP) and Glutathione (GSH). After stratification on HbA1c and age, oxidative damage (Malonaldehyde (MDA), protein carbonyls) were assessed in a sub‐sample (n=47).

Means ± s.d. or percentages were calculated as appropriate. General linear model was used with adjustment for age, body mass index, food groups’ consumption (fruits, vegetables meat/beans, dairy), fat and total daily energy intakes.

In the whole sample, mean age was 53.46±9.36 years, mean HbA1c was 7.69±1.54 %. A majority (86.50%) of patients was taking oral medication. Mean values for vitamin C, ABTS, FRAP and GSH were 0.09±0.07 g/L, 0.10±0.04 g /L, 1.00±0.46 g/L and 0.13±0.03 g/L, respectively. In the sub‐sample, MDA and protein carbonyls were 12.67±3.44 µmol/L and 0.29 nmol/L, respectively. None of the markers of antioxidant status were significantly related to HbA1c. MDA and protein carbonyls were strongly positively associated with HbA1c (p=0.02, 0.01, respectively).

In this work, poor glycemic control is strongly positively associated with oxidative damage in adults with T2DM, suggesting that oxidative stress is closely intertwined with this disease.

Grant Funding Source: Supported by Sheikh Hamdan Award
Red light mediated vasodilation (698.12)
Authors: Lohr Nicole , Lindemer Brian , Gebremendhin Debebe , Harder David
Published: June 2021

Over 8 million Americans are treated for symptoms related to peripheral artery disease (PAD). Risk factors for PAD have been directly related to a reduction in NOS expression and NO production. Light irradiation at the far red/near infrared region (670 nm (R/NIR)) can stimulate angiogenesis by increasing NO independent of NOS through release of NO from nitrosyl‐heme stores. We hypothesized R/NIR can stimulate vasodilation, and serve as a potential therapy to improve endothelial function. Methods: Human umbilical vein endothelial cells (HUVEC) were loaded with DAF‐2 and stimulated with R/NIR at 25mW, 50mW, and 75mW up to 12J. NO was measured using HPLC of DAF‐2T. Facial arterioles from c57Bl/6 mice and human coronary arterioles from patients with and without atherosclerosis were pressurized to 60mmHg and pre‐constricted. Vessels were irradiated with 670 nm light (10mW/cm2) up to 5 min at a time with 10 min dark period between irradiation. Results: R/NIR stimulated a 26.83% (± 6.99, p<0.05) increase in NO over control. Preincubation with c‐PTIO, a NO scavenger, completely abolished R/NIR stimulated NO. In mice, vessel diameter increased 17.8% (±3.0, p<0.05) after R/NIR exposure. In vessels denuded prior to R/NIR dilation, vasodilation was abolished (1.2%, ± 2.21, p<0.001), suggesting R/NIR vasodilation was endothelial dependent. Pre‐incubation with 1mM L‐NAME did not significantly attenuate dilation (13.74% ±2.2), however c‐PTIO (100µM) could successfully block dilation (‐.51% ± 1.1, p<0.001). R/NIR significantly increased in coronary arterioles from patients with atherosclerosis 24.5% (±2.1%, p<0.01) and without atherosclerosis 44.8% (±6.2%, p<0.01). Conclusion: R/NIR can stimulate vasodilation in human and murine vessels. This vasodilation is endothelium and NO dependent, but does not require functional nitric oxide synthase. The exact source for the NO increase is unknown, but intracellular nitrosyl heme moieties should be considered as a previously unexplored source for NO.
Nitrite supplementation improves arterial mitochondrial health in old mice (698.5)
Authors: Sindler Amy , Johnson Lawrence , de Picciotto Natalie , Ahsan Sanjana , Weber Stewart , Seals Douglas , Gioscia‐Ryan Rachel
Published: June 2021

Age‐related declines in endothelial‐dependent dilation (EDD) are mediated in part by excessive mitochondrial (mito) reactive oxygen species (mtROS) production. Since nitrite improves vascular function with aging and enhances mito health, we hypothesized that nitrite supplementation (drinking water, 50 mg/L, 8 w) would improve arterial mito health in old mice. Old (O; 27 mo, n = 9) C57 male mice had lower ex vivo carotid artery EDD vs. young (Y; 6 mo, n = 9) (56 ± 6 vs. 84 ± 3%, p<0.05), which nitrite restored (ON; 91 ± 3%). Incubation with the mito specific antioxidant, MitoQ (1.0 mM), selectively improved EDD in O (85 ± 4%), but had no effect in Y or ON. Increased mtROS challenge with rotenone (0.5 uM) further impaired EDD in O (35 ± 6%) while Y and ON were protected. Consistent with mtROS‐mediated vascular dysfunction in aging, arteries from O had reduced expression (WB) of key markers of mito health, including mito maintenance proteins (AMP‐activated protein kinase, O: 0.5 ± 0.1 vs. Y: 1.0 ± 0.2; peroxisome proliferator‐activated receptor γ coactivator 1‐α, O: 0.7 ± 0.1 vs. Y: 1.0 ± 0.1) and mito mass (complex IV, O: 0.6 ± 0.1 vs. Y: 1.0 ± 0.2; voltage‐dependent anion channel, O: 0.7 ± 0.07 vs Y: 1.0 ± 0.05, AU all p<0.05), which nitrite restored (ON: 1.7 ± 0.5; 1.9 ± 0.5; 1.8 ± 0.4; .9 ± 0.04 AU, respectively, all p<0.05 vs. O). Nitrite supplementation may reverse age‐related endothelial dysfunction in part by improving vascular mitochondrial health.

Grant Funding Source: Supported by NIH AG013038 and AG000279
Influences of gender, age, and ethnicity on nitric oxide and cGMP concentrations biocaptured over acupoint/meridians: analysis in fifty‐nine human subjects (698.7)
Authors: Ma Sheng‐Xing , Li Xi‐Yan , Jiang Isabelle , Hu Jay , Lee Paul
Published: June 2021

The purpose of this study was to determine the influence of gender, age, and race on nitric oxide (NO)‐cGMP concentrations over acupuncture points (acupoints), meridian lines between acupoints, and non‐meridian controls in healthy volunteers. A biocapture device was taped onto the skin surface along the Pericardium (PC) and Lung (LU) acupoints/meridian located on the arm, and the Bladder (BL) acupoints/meridian on the leg in 59 subjects. Total nitrate and nitrite (NOx¯) and cGMP levels in the biocaptured samples were measured using chemiluminescence and ELISA, respectively. Results show that cGMP concentrations over the PC acupoints and non‐meridian area are higher in females than males, but NO concentration over the region is modified by body weight, which contain more subjects with obesity in man. NO and cGMP levels along the BL meridian are increased in middle age/elder subjects compared to adolescent/adult subjects but not over PC regions. There was no detectable significance between NO nor cGMP concentrations among racial groups studied. Our findings suggest that different NO and cGMP levels between genders and age groups are influenced by sympathetic activity, body weight, hormones, sweat rates, and skin thickness. Enhanced NO and cGMP levels along the BL meridian and bladder dysfunction in aged subjects requires investigation.

Grant Funding Source: Supported by NIH AT002478 and Oppenheimer Seed Grant to S. Ma
Chronic hypoxia promotes preferential loss of MLCK not associated with MLC in fetal lamb arteries (700.7)
Authors: Williams James , Aguilar Luisa , Pearce William
Published: June 2021

As we have previously shown (PMID: 23325408), chronic hypoxia (CH) dramatically reduces MLCK abundance in fetal common carotid arteries without proportionate loss of contractility. The present study explores the hypothesis that CH preferentially decreases MLCK not associated with MLC and thereby preserves contractility. Common carotid arteries from term fetal sheep were mounted in a custom‐made organ bath apparatus that enabled computer controlled freezing of artery samples in 200 msec increments (PMID: 18835918). Artery segments were frozen in methacarn fixative and analyzed for MLC, S19 pMLC and MLCK by Westerns and confocal microscopy. CH reduced MLCK abundance 24‐fold, but did not significantly alter MLC abundance. Despite decreased MLCK abundance, rates of ser19 MLC phosphorylation after 1 sec electrical stimulation were unchanged in Calyculin A (10 nM) pre‐treated arteries (27.6±0.8% N, 22.4±4.9% H). Confocal microscopy revealed that colocalization of MLC with MLCK was increased more than 3‐fold in hypoxic arteries. These data indicate that hypoxic losses of MLCK are restricted to a fraction of MLCK not associated with MLC, such that the remaining MLCK is more tightly colocalized with MLC and is fully capable of phosphorylating MLC and sustaining contraction. The role of the large fraction of MLCK not colocalized with MLC is unclear, but may serve structural or other non‐enzymatic purposes.

Grant Funding Source: Supported by NIH Grants HD‐31266 and NS‐076945
A microRNA cluster miR‐23/24/27 is regulated by aldosterone to alter Na+ transport in the kidney distal nephron (711.5)
Authors: Butterworth Michael , Liu Xiaoning , Edinger Robert
Published: June 2021

The epithelial sodium channel (ENaC) is a major determinant of sodium (Na+) and water balance in the kidney. We previously reported that aldosterone (aldo) modifies expression of microRNAs (miRs) to alter target gene expression and regulate Na+ transport in principal cortical collecting duct (CCD) epithelial cells. These studies focus on 2 clusters of miRs that are significantly upregulated by aldo, and are members of the miR‐23/24/27 family. In cultured mCCD‐cl1 cells, aldo stimulation (50nM, 24hr) resulted in a ~1.7 fold increase in all family members as determined by microarray and qPCR. In vivo, these miRs were significantly increased (1.7‐3.5 fold) in isolated CCD cells from mice on low Na+ vs standard salt diets. Exogenous overexpression on the miRs, using oligonucleotide mimics, increased ENaC‐mediated Na+ transport in the mCCD cell line (44±5.9%, n=8, p<0.002), in the absence of aldo stimulation. Using our newly developed algorithm, ComiR, we predicted and verified regulation of intersectin 1&2 as targets of these miR clusters, and possible mediators of ENaC regulation. These data confirm a central role for aldo‐regulated miRs in the maintenance of Na+ transport in the distal kidney nephron.

Grant Funding Source: Supported by DK078917
Maturation of vascular anatomy in the zebrafish brain (726.1)
Authors: Rahmat Sulman , Gilland Edwin , Heinbockel Thomas
Published: June 2021

Although embryonic and larval vasculature in zebrafish have been described, few studies document brain vessels beyond two weeks of development. Transgenic zebrafish, between one and twelve weeks old, expressing GFP in cranial motor neurons were injected with fluorescent vascular dye and examined with confocal angiography to demonstrate maturation of cerebral vasculature. A segmental series of hindbrain central arteries (CAs) in early larvae extend from the basilar artery, penetrate the midline of rhombomeres (r) r3‐8 and branch sparsely within the neuroepithelium before draining into laterally located veins. While hindbrain CAs in younger stages were roughly similar along the series, the r3 and r8 stems became progressively larger than the others, with more elaborate branching and increased neurovascular territorial supply. By 75 days, the r3 CA was effectively the arterial supply of the entire rostral hindbrain, including branches that supply the migrated facial motor nucleus in r7. Since small r4‐7 CAs are present in mature brains, it is likely that the increased territory of the r3 CA resulted from angiogenic sprouting of existing branches rather than capture of neighboring arterial trees by vessel fusion. In contrast, the major cerebellar, midbrain and forebrain arterial vessels supplied roughly the same neuronal territories in mature animals as in early larvae, but with greatly increased branching.

Grant Funding Source: N/A
Normal anatomy and variations of the lateral femoral cutaneous nerve in relation to the iliac crest: implications for anterior iliac crest bone grafts (726.10)
Authors: Klemensberg Jason , Johnson Marjorie , Merrifield Peter , Shimizu Michael , Galil Khadry
Published: June 2021

Anterior iliac crest bone grafting (AICBG) remains the gold standard for reconstructive procedures in orthopedic and oral‐maxillofacial surgeries. It has been estimated that 10% of patients undergoing AICBG are at risk of injury to their lateral femoral cutaneous nerve (LFCN). Injury to this nerve can result in numbness, burning, itching or pain sensations over the lateral aspect of the thigh, known as meralgia paresthetica. The course of the LFCN within the greater pelvis cannot be seen with imaging technologies. The objective of this study is to elucidate the 3D course and variability of the LFCN as it emerges from the psoas major muscle to the anterior superior iliac spine (ASIS). Using 25 embalmed cadavers, the length and depth of the LFCN is measured from the ASIS to 7 cm posterior to the ASIS at 1 cm intervals. Preliminary results (n=10 sides) show these measurements are highly variable between cadavers and that there are three ways the nerve courses: convex, concave or straight course. The first three dimensional danger‐zone of the LFCN relative to the iliac crest will be established and incorporated onto a 3D illustration of the pelvic bone using Amira software. A predictive model will be determined to approximate the location of the LFCN with respect to age, sex, height, bone width, and ASIS‐ASIS distance. A better understanding of the course of the LFCN is important to minimize graft site morbidity.
Examining the immunomodulatory effects of GM‐CSF and IL‐12 in a mouse model of Alzheimer’s disease (728.2)
Authors: Schettler Stephen , Klaidman Lori , Vinters Harry , Stark Elena
Published: June 2021

Granulocyte macrophage colony stimulating factor (GM‐CSF), a growth factor for microglia/macrophages, has been associated with protective functions in a mouse model of Alzheimer's disease (AD) in terms of Amyloid β (Aβ) plaque accumulation and cognitive function. We extended these studies by combining GM‐CSF and Interleukin‐12 (IL‐12), considered to be a differentiation factor of naive T‐cells into Th1 cells. A transgenic APPswe/PS1dE9 mouse model for AD was used and mice were grown to an advanced stage of brain Aβ plaque deposition. At 18 months old, AD mice were then injected daily for 20 days, subcutaneously with doses of GM‐CSF (1.7 μg/day) and/or IL‐12 once a week (20 ng) and sacrificed at day 21. Mice were divided into the following groups: 1) AD mice with GM‐CSF plus IL‐12, 2) AD mice with IL‐12 alone and 3) AD mice with no treatment. Mouse brains were perfused and brain slices were quantitated by immunohistochemical methods using Aperio software for Aβ 1‐40 or Aβ 1‐42 peptides. The results indicate a complex neuromodulatory effect. Aβ 1‐40 demonstrated a modest decrease in the AD, GM‐CSF, IL‐12 group compared to the control AD group. However, Aβ 1‐40 was highly variable in the AD, IL‐12 group compared to the control AD group. Aβ 1‐42 on the on the other hand, was significantly decreased in the AD, IL‐12 group, compared to the AD control group. However, there was no significant change in Aβ 1‐42 in the AD, GM‐CSF, IL‐12 group. Analysis of microglial activation in these samples is ongoing and final results will be presented.
The unique developmental and aging profile of tau protein in the long‐lived naked mole‐rat brain resists Aβ toxicity (728.8)
Authors: Orr Miranda , Salinas Angelica , Buffenstein Rochelle
Published: June 2021

Alzheimer’s disease (AD), the most prominent neurodegenerative disease, is characterized by Aß plaques, hyperphosphorylated tau, neurodegeneration and dementia. A prominent hypothesis suggests that elevated Aß peptide initiates AD pathogenesis including tau pathology. The tau protein, encoded by microtubule associated protein tau (MAPT) gene, is expressed predominantly in neuronal axons where it physiologically stabilizes microtubules (MTs). With over 20% of residues permissive to phosphorylation, tau function greatly depends on its phosphorylation state. Specifically, heavily phosphorylated tau loses MT binding affinity and coincides with neurodegeneration in AD. Unfortunately, genetically modified animal models overexpressing transgenic Aß have failed to recapitulate AD‐like tau misprocessing. To determine, instead, if endogenously expressed high levels of Aß alter tau and induce AD‐like pathology, we are studying the extremely long‐lived naked mole‐rat (NMR). NMRs, unlike mice, naturally contain measurable amounts of brain Aß. Using histological and biochemical assays, we have characterized tau protein from postnatal day 1 through 30 years of age. Remarkably, NMR brains display tau expression, phosphorylation, and splicing patterns distinct from mice and humans that seemingly enable them to cope with life‐long high levels of Aß.

Grant Funding Source: Supported by: Biology of Aging Training Program 5T32AG021890
In vitro skeletal myofiber disassembly and recovery with cardiotoxin (731.9)
Authors: Mattioli Patrisia , Hernanadez Elena , Bratton Stephaie , Bova Michael , DiLullo Camille
Published: June 2021

Injection of cobra venom cardiotoxin into skeletal and cardiac muscle In Vivo causes injury to the respective muscle myofibers. This cardiotoxin has been shown specifically to disassemble myofibers in the muscle while leaving the surrounding tissues unaffected. To determine if cardiotoxin would similarly affect skeletal muscle In Vitro, primary embryonic chick skeletal muscle cultures were exposed to low doses of cardiotoxin from Naja mossambica mossambica. Cells at 4 days in culture were exposed to toxin for 48 hours. Toxin was washed out after the treatment period. Cultures were fed daily, fixed on days 1‐3 after wash out and immunofluorescently labeled for muscle specific proteins. Immediately after wash‐out it was observed that myofibers had disassembled in the majority of myocytes. Preliminary data indicated that nuclear counts between treated and untreated cultures remained relatively constant suggesting limited cell death in treated cultures. Compared to control cultures, length and girth of treated myocytes as well as number of nuclei per myotube were negatively impacted by toxin exposure. After wash‐out toxin treated cultures exhibited thin, unstriated myofibers in mononucleated myocytes by day 2 of recovery. By day 3 of recovery myotubes with 2‐3 nuclei, slightly increased girth and small numbers of myofibers with alpha‐actinin striations were detected.
Alteration of metabolic activities by modulation of PI3K/AKT and mTOR pathways in ovarian cancer cells (766.10)
Authors: Calianese David , Cammarata Garrett , Best Charles , Wan Yinsheng
Published: June 2021

Ovarian cancer accounts for three percent of all cancers that affect women world‐wide and it is regarded at the deadliest form of gynecological cancer. Alteration in cellular metabolism is a manifestation of cancer that typically results in a shift from oxidative phosphorylation to aerobic glycolysis. This change is defined as the Warburg Effect. In this study, we used cell culture techniques to investigate the metabolic behavior of an ovarian cancer cell line, CaOV3. We determined metabolic activity through XF Cell Mito Stress and XF Glycolysis stress tests using a Seahorse Bioscience XFe96 Analyzer. After titrations were completed for optimal cell number and concentration of Oligomycin, we pretreated CaOV3 cells with PI3K/AKT and mTOR inhibitors and repeated stress tests. We found that under the influence of Rapamycin (mTOR inhibitor) and LY294002 (PI3K/AKT inhibitor), the cells exhibited an increase in oxygen consumption by OCR (pMoles/min) and ECAR (mpH/min) analysis, suggesting an increase in capacity for cellular respiration and subsequent ATP production. Our results also indicated that Doxarubicin, a commonly applied chemotherapy agent, does not affect metabolic levels. Taken together, we conclude that in response to the inhibition of the PI3K/AKT and mTOR pathway that promotes protein synthesis and proliferation, the cancer cells attempt to combat the effects by increasing their metabolic capacity. Our data also suggest that modulation of PI3K/AKT and mTOR pathways and alteration of metabolic activities may reveal molecular targets for ovarian cancer treatment.

Grant Funding Source: Supported by INBRE
Killing cancer cells with nanotechnology: the effect of CaS nanostructures on the cell replication rate and survival of human mammary adenocarcinoma cell lines (780.9)
Authors: Rivera Daniel , Carrasco Nestor , Maldonado Louis , Suarez Edu , Poventud Carlos , Marrero Carolina , Castro Miguel
Published: June 2021

Calcium sulfide (CaS) nanostructures have the potential to play an important role as cadmium‐free nanoscaled semiconductors with applications in bioimaging, in vivo labeling, and sensing, and for drug delivery systems. The objective of this work is to establish the effect of CaS nanostructures on cell replication rate and survival of human mammary adenocarcinoma cell lines. Human adenocarcinoma cell lines are grown in eagle’s minimum essential medium (EMEM). Single doses of dispersions containing traces of CaS nanostructures were added to human mammary adenocarcinoma cell lines with a 70 % confluence. After a 24 hour period, 200,000 cancer cells were subculture in T‐75 treated flasks. The number of dead cells in the cell culture media supernatant was determined in intervals of 24 hours using a Biorad T10 cell counter. The number of dead cells in the supernatant is found to increase from 2,000 in control cell cultures to over 70,000 in cancer cell cultures exposed to a single dose of the CaS dispersion in the first 24 hours: this represents 35 % of the initial cell seeding. The number of dead cells remains in the range of 60,000 to 30,000 in measurements performed between 48 and 72 hours following the single dose of the dispersion containing the CaS nanostructures. The number of replicating cells is found to decrease in the first 72 hours and to increase thereafter. We conclude that the CaS nanostructures affect the replication rate and survival of human adenocarcinoma cells and demonstrate the role CaS nanostructures may play to treat localized cancers.

Grant Funding Source: NIH NIGMS‐R25GM088023 and NIGMS‐R25GM096955
Viewers vs. doers: how watching food television and cooking frequency relate to BMI (811.21)
Authors: Pope Lizzy , Latimer Lara , Wansink Brian
Published: June 2021

Objectives: Examine where nutritional gatekeepers obtain information about new foods, and whether information source impacts Body Mass Index (BMI), as well as whether any impact varied according to how often the participant cooked from scratch.

Methods: Five hundred and one young women completed a survey assessing where they obtained information on new foods, and how often they cooked from scratch. Linear regressions were run to determine associations between information source, cooking from scratch, and BMI.

Results: Obtaining information from cooking shows was positively correlated with BMI (p<0.05), whereas obtaining information from print, online, or in‐person sources was not significantly associated with BMI. An interaction between watching cooking shows and cooking from scratch was associated with higher BMI (p<0.05).

Conclusions: Obtaining information about new foods from TV cooking shows appears to have a unique impact on BMI, versus obtaining information from other sources. Furthermore, watching cooking shows may have a differential effect on BMI for those who are “viewers,” in that they only watch the show, versus those who are “doers,” meaning they view and cook.
New Year’s res‐Illusions: food shopping in the New Year competes with healthy intentions (811.22)
Authors: Pope Lizzy , Hanks Andrew , Just David , Wansink Brian
Published: June 2021

Objective: To assess holiday and post‐holiday purchasing patterns of unhealthy and healthy foods when compared to purchasing in early fall (baseline).

Methods: Grocery‐purchase data were collected for 207 households from July 2010 to March 2011. Calorie data for foods purchased were extracted from online nutritional databases. The cooperating grocer’s proprietary nutrient‐rating system was used to designate “healthy,” and “unhealthy” items. Average weekly (Sunday through Saturday) expenditures and per‐serving calories for the holiday period, (Thanksgiving‐New Year’s), and the post‐holiday period (New Year’s‐March), were compared to average weekly expenditures during the baseline period (July‐Thanksgiving). Regression analysis was used to compare expenditures and calories purchased in each of the three time periods.

Results: Household expenditures increased in the holiday period from baseline, with 75% of additional expenditures accounted for by less‐healthy items. Furthermore, expenditures increased by 53% in the post‐holiday period compared to the holiday period, and households purchased twice as many additional per‐serving calories in the post‐holiday period compared to the holiday period. Specifically, additional per‐serving calories from healthier foods increased 510% in the post‐holiday period compared to the holiday period, while purchasing of less healthy foods remained constant.

Conclusions: Post‐holiday purchases of healthy items may have provided a “health halo” justifying purchasing of unhealthy items. Despite resolutions to eat more healthfully after New Year’s, consumers may adjust to a new “status quo” of increased unhealthy‐food purchasing during the holidays, and vicariously fulfill their New Year’s resolutions by spending more on healthy foods.

Grant Funding Source: Supported By the National Institutes of Health (NIH), grant 1RC1HD063370‐01
Soy protein isolate and estradiol differ in their effects on the mammary gland of weanling male and female rats (818.1)
Authors: Ronis Martin , Miousse Isabelle , Gomez‐Acevedo Horacio , Hennings Leah , Shankar Kartik , Cleves Mario , Badger Thomas
Published: June 2021

Isoflavones are phytochemical components of soy diets that bind weakly to estrogen receptors (ERs). To study potential estrogen‐like actions of soy in the mammary gland, we fed weanling male and female Sprague‐Dawley rats a casein diet from PND21 to PND33, the same diet substituting soy protein isolate (SPI) for casein, or the casein diet supplemented with estradiol (E2) at 10 μg/kg/day. In contrast to E2, the SPI diet induced no significant change in mammary morphology. In males, there were 34 genes for which expression changed 蠅2‐fold in the SPI group versus 509 changed significantly by E2, and 8 versus 174 genes in females. Nearly half of SPI‐responsive genes in males were also E2‐responsive, including adipogenic genes. Serum insulin was found to be decreased by SPI in males. SPI and E2 both down‐regulated the expression of ERα (Esr1) in males and females, and ERβ (Esr2) only in males. Chromatin immunoprecipitation (ChIP) revealed an increased binding of ERα to the promoter of the progesterone receptor (Pgr) and Esr1 in both SPI and E2 treated males compared to the casein group but differential recruitment of ERβ. ER promoter binding did not correlate with differences in Pgr mRNA expression. This suggests that SPI fails to recruit appropriate co‐activators at E2‐inducible genes. Our results indicate that SPI behaves like a selective estrogen receptor modulator rather than a weak estrogen in the developing mammary gland.

Grant Funding Source: Supported by USDA‐ARS CRIS 6251‐51999‐007‐04S
Reduced insulin sensitivity limits reductions in small dense LDL due to lean beef consumption (821.14)
Authors: Wu Xiuzhi , Roussell Michael , Smith Stephen , Kris‐Etherton Penny , Walzem Rosemary
Published: June 2021

Atherogenicity of small dense LDL (e.g. LDL‐4) is attributed to their ready penetration of the artery wall & reduced oxidative stability. We tested the effect of beef consumption on lipoprotein (Lp) density distribution in two beef feeding trials. Trial1, 27 subjects consumed in random order: Healthy American diet (HAD), Dietary Approaches to Stop Hypertension (DASH), Beef in an Optimal Lean Diet (BOLD) and BOLD plus extra protein (BOLD+) (20, 28, 113, 153g lean beef/d, respectively, all diets ~30 EN% fat) for 5wks. Trial 2, 25 men consumed in random order: 5‐114g, 24% fat beef patty/wk for 5wks of both low and high MUFA beef patties(Patty, 62 g lean beef/d, 40 EN% fat diet). Density profiles of NBD‐ceramide labeled Lp were imaged following isopycnic ultracentrifugation. Compared with HAD, DASH and BOLD diets decreased HDL‐2b (p<0.05), while BOLD+ only reduced LDL‐4 (p<0.05). Trial 2 found no general diet effect on Lp subclass compared to baseline. Stepwise multiple linear regression of data from both trials identified diet treatment (p<0.0001) and baseline HOMA‐IR [(fasting glu/fasting insulin)/22.5] (p<0.0001) as predictors of LDL‐4 change from baseline. Lower baseline HOMA‐IR & consumption of DASH, BOLD or BOLD+ diets corresponded to a greater reduction in LDL‐4 (p<0.0001). In summary, a heart healthy diet rich in lean beef decreases CVD risk (in part by decreasing LDL‐4) in insulin sensitive individuals.

Grant Funding Source: National Cattleman's Beef Association
Impact of dietary fiber‐based SCFA mixtures on colon epithelial barrier function (822.1)
Authors: Chen Tingting , Kim Choon Young , Hamaker Bruce
Published: June 2021

Patients with HIV, obesity, and inflammatory bowel disease often have impaired gut barrier function. Short chain fatty acids (SCFAs), particularly butyrate, provide energy to the intestinal epithelium and improve barrier function. Our lab reported various fibers have different fermentation profiles, generating SCFAs varying in total concentrations and proportions of acetate, propionate, and butyrate. To study the impact of SCFA profiles of various fibers on gut barrier function, SCFA mixtures with 5, 20, and 50% butyrate proportion were designed to correspond to SCFA profiles of tested fiber fermentations, and were subjected to healthy and LPS/TNF‐α treated Caco‐2 cell monolayers. Integrity of monolayers was indicated by transepithelial electric resistance, dextran flux, and tight junction (TJ)‐related protein levels. Pro‐inflammatory genes were determined by qPCR. Total concentration of 40‐60 mM SCFA with 20% butyrate improved barrier function without causing damage to the monolayer. Within this defined concentration range of SCFAs, butyrate proportion up to 50% further promoted TJ function. Applying this result to dietary fiber selection, certain rapidly fermenting fibers may generate a high amount of SCFA that may harm gut barrier. Slow fermenting fibers that deliver appropriate concentrations of SCFAs to the epithelium with high proportion of butyrate might improve barrier function.