bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition of Sepiapterin Mediated Formation of Dihydrobiopterin and Chemical Redox Cycling by Sulfa Drugs
Authors: Yang Shaojun , Jan Yi‐Hua , Mishin Vladimir , Richardson Jason , Heindel Ned , Heck Diane , Laskin Debra , Laskin Jeffrey
Published: June 2021

Sepiapterin reductase (SPR) catalyzes the reduction of sepiapterin to dihydrobiopterin (BH2), a precursor for tetrahydrobiopterin (BH4), a cofactor critical for nitric oxide biosynthesis and alkylglycerol and aromatic amino acid metabolism. SPR also mediates chemical redox cycling, catalyzing one electron reduction of redox active chemicals including quinones; rapid reaction of these radicals with molecular oxygen generates reactive oxygen species (ROS). Using recombinant human SPR, sulfonamide and sulfonylurea based sulfa drugs were found to be potent non‐competitive inibitors of both sepiapterin reduction and redox cycling. The most active inhibitors of sepiapterin reduction (IC50's = 31‐180 nM) were sulfasalazine, sulfathiazole, sulfapyridine, sulfamethoxazole and chlorpropamide. Higher concentrations of the sulfa drugs (IC50's = 0.37‐19.4 µM) were required to inhibit redox cycling. In PC12 cells, which generate catecholamine and monoamine neurotransmitters via BH4‐dependent amino acid hydroxylases, sulfa drugs inhibited both BH2/BH4 biosynthesis and redox cycling. This resulted in decreased production of dopamine and dopamine metabolites, 3, 4‐dihydroxyphenylacetic acid and homovanillic acid, and 5‐hydroxytryptamine. This was reversed by BH4. These data suggest that SPR and BH4‐dependent enzymes, are ‘off‐targets' for sulfa drugs, which may underlie their side effects. AR055073, ES005022.
The Water Channel Aquaporin 1 is Associated with Human Atherosclerotic Lesions and Attenuates Progression of Atherosclerosis in Male Mice Independent of Blood Pressure
Authors: Stubbe Jane , Wintmo Pamela , Johansen Søren , Hansen Pernille , Lindholt Jes , Rasmussen Lars , Bie Peter , Jensen Boye
Published: June 2021

The water channel aquaporin 1 (AQP1) supports tumor growth by facilitating endothelial cell migration in tumor neovascularization. We tested the hypothesis that AQP1 promotes neovascularization of growing vascular lesions and aneurysms using human vascular tissue and AQP1‐/‐Apolipoprotein E (ApoE)‐/‐mice. AQP1 was localized in the neovasculature in human atherosclerotic lesions and in human abdominal aortic aneurysm wall, while in internal thoracic arteries AQP1 was restricted to endothelial cells and adventitial vasa vasorum. In female ApoE‐/‐ mice with lesion burden aortic AQP1 mRNA expression was augmented. AQP1 was localized in endothelial cells, vasa vasorum and intralesional cells at the aortic root. Lesion burden did not differ between AQP1‐/‐ApoE ‐/‐ and ApoE ‐/‐ mice after 8 and 16 weeks on western diet (WD, n =13‐15). Male AQP1‐/‐ApoE‐/‐ mice fed WD and treated with angiotensin II (ANGII) for 4 weeks had elevated atherosclerotic lesion burden compared to ApoE‐/‐ mice (11± 1 vs. 4 ± 2 %, n =10‐11, p<0.02). Freely moving AQP1‐/‐ApoE‐/‐ and ApoE‐/‐ mice showed similar blood pressure responses to ANGII i.v. infusion (60ng/kg*day for 7 d) measured by indwelling catheters (nighttime: 129 ± 4 and 128 ± 5 mmHg, respectively). It is concluded that AQP1 is associated with intralesional neovasculature in humans and mice. AQP1 accelerates ANGII‐induced atherosclerosis in male mice independent of blood pressure.

Supported by A. Andersen grant and J. & O. Madsens grant
Electrophilic Nitro‐Fatty Acids Exert Cardioprotection against Hypertrophic Remodeling and Fibrosis in Pressure Overloaded Mice
Authors: Villacorta Luis , Guo Yanhong , Zhang Ji , Converso‐Baran Kimber , Schopfer Francisco , Freeman Bruce , Chen Yuqing E.
Published: June 2021

Electrophilic nitro‐fatty acids, the nitration products of unsaturated fatty acids are generated in cardiomyocytes after ischemia/reperfusion injury and protects against myocardial infarction. Herein, we determine whether nitro‐oleic acid (OA‐NO2) is equally cardioprotective in hypertrophic remodeling. Cardiac hypertrophy were established in mice by transverse aortic constriction (TAC). Mice received either OA‐NO2, or OA as non‐nitrated fatty acid control. OA‐NO2 delivery reduces hypertrophic remodeling and significantly reduces cardiac fibrosis compared to OA. Overall cardiac function, e.g. ejection fraction, is significantly improved by OA‐NO2. Electrophilic signiling through Nrf2 is a plausible mechanism of cardioprotection by nitroalkenes. However, in Nrf2 knockout mice, which develop exacerbated cardiac hypertrophy after TAC, OA‐NO2 delivery persistently confer cardioprotection indicating an alternative pathway for the improved cardiac function mediated by OA‐NO2. Rather, OA‐NO2 primarily targets cardiac fibrosis and extracellular matrix gene expression profiles. Thus, protection against cardiac hypertrophic by nitro‐fatty acids via improving cardiac fibrosis supports further promise on nitro‐fatty acids derivatives as therapeutic agents for cardiovascular disease. Supported by the NIH (R01‐HL68878, R01‐HL058115, R01‐AT006822A), and the American Heart Association (10SDG4150085).
Impaired Compensatory Responses to Hypovolemic Circulatory Stress in Women with Vasovagal Syncope
Authors: Skoog Johan , Lindenberger Marcus , Zachrisson Helene , Länne Toste
Published: June 2021

Objective

A fall in cardiac output (CO) due to a reduced venous return has been suggested as an important factor for the hemodynamic instability in vasovagal syncope (VVS). The aim was to investigate compensatory mechanisms to defend central blood volume during hypovolemic stress in women with VVS.

Design and Methods

Single (30 mmHg) and graded (20 to 70 mmHg) lower body negative pressure (LBNP) were used to create hypovolemic stress in 15 young women with VVS and 15 matched controls. Peripheral mobilization of venous blood from the arm (capacitance response), net fluid absorption and lower limb blood pooling were evaluated with volumetric technique. Hemodynamic responses and plasma norepinephrine (P‐NE) were measured.

Results

Although CO decreased in both groups during graded LBNP (P < 0.05), VVS presented a significantly lower CO at LBNP of 40 mmHg (P = 0.045). Mobilization of capacitance blood from the peripheral circulation (ml/100 ml) was reduced in VVS (LBNP 40 mmHg, 0.194±0.02 vs. 0.299±0.02, P = 0.007; LBNP 50 mmHg, 0.172±0.01 vs. 0.314±0.04, P = 0.003), and net fluid absorption was ~40% lower (P = 0.019). VVS displayed a blunted percentage increase in P‐NE (P = 0.009) and a reduced maximal percentage increase in TPR (P = 0.041) during graded LBNP. Lower limb blood pooling was comparable in VVS and controls.

Conclusions

Important compensatory mechanisms to defend central blood volume during hypovolemic stress were impaired in VVS. This may seriously impede maintenance of cardiac output during hypovolemic stress and could contribute to the pathogenesis of VVS.
Coronary Blood Flow is Reduced by Doxorubicin: a Ultrasound Microscopic Study
Authors: Liu Shi
Published: June 2021

The use of doxorubicin (DOX), an anti‐cancer drug, in treatment for various malignancies is limited because of its cardiotoxicity; whether DOX causes vascular disorders is unknown. Ultrasound biomicroscopy (UBM) has been a valuable, non‐invasive technique in monitoring cardiac function but not commonly used in small animal vascular studies.This study aimed to 1) study the velocity of blood flow in the left coronary artery (LCA) under physiological conditions and vascular responsiveness to DOX treatment with UBM, and 2) examine the effect of an aqueous extract of TA bark (TAAqE), a cardiotonic, on the blood‐flow velocity of LCA after DOX treatment. Young adult mice were injected intrapleurally with a single dose of DOX (20 mg/kg) or saline in the presence or absence of TAAqE. Doppler UBM was performed the day before and 1 week after DOX treatment using Vevo®2100. The blood flow of LCA and left ventricle (LV) function were monitored in the parasternal short‐axis view of UBM. The peak blood‐flow velocity of LCA during cardiac diastole and systole was 909±35 and 284±13 mm/s, respectively, (n=20) without drug treatment. Abnormality of LV function could be detected in the 1st week and was increased rapidly in the 2nd week of DOX treatment. Meanwhile, the integrated blood flow of LCA was reduced about 42% (n=9) accompanied by an increase in the resistive index by 85%. Pretreatment of TAAqE offset the detrimental effects of DOX on LCA and LV function. In summary, 1) DOX caused reduction in LCA blood‐flow velocity besides cardiotoxicity, and 2) TAAqE offered some cardiovascular protective effects. In conclusion, Doppler UBM is very useful in detecting longitudinal hemodynamic changes of LCA. The DOX‐induced decrease in blood‐flow velocity of LCA could result from reduction in the blood flow and/or vasoconstriction.
Effect of Left Ventricular‐Central Artery Coupling on Cerebrovascular Hemodynamics: Insights from Endurance Training Intervention
Authors: Tomoto Tsubasa , Sugawara Jun , Nogami Yoshie , Aonuma Kazutaka , Maeda Seiji
Published: June 2021

Increased pulsatile fluctuation of cerebral blood flow may mechanically impact on vulnerable brain tissue. The aim of this study was to investigate participation of central artery damping function on pulsatile blood flow transmission from LV to the brain using the endurance training intervention study design. We measured maximal oxygen uptake (VO2max), LV ejection velocity (via ultrasound), carotid arterial compliance (via ultrasound and applanation tonometory), and middle cerebral arterial (MCA) hemodynamics (via transcranial Doppler) before and after 16‐week endurance training in 13 collegiate tennis players. After the intervention, VO2max was significantly increased (P<0.01). Despite a significant increase in LV ejection peak velocity (P<0.05), MCA blood flow (i.e., peak and mean flow velocity, pulsatility index) were remained unchanged (P=0.854). Carotid arterial compliance was improved after the intervention (P<0.01) and individual changes were correlated with the corresponding changes in MCA pulsatility index (r=‐0.584, P<0.05). These results suggest that because of the contribution of increased central arterial compliance, cerebral pulsatile flow fluctuation was not augmented even when LV ejection was increased by endurance training.
Reduced Sympathetic Tone by Overexpression of Kv4.3 in the Rostral Ventrolateral Medulla (RVLM) of Rats with Heart Failure
Authors: Becker Bryan , Wang Hanjun , Zucker Irving
Published: June 2021

Chronic heart failure (CHF) is characterized by excess sympathetic tone that exacerbates cardiovascular damage. Our previous work demonstrated reduced Kv4.3 expression in the RVLM, a key cardiovascular control center regulating sympathetic tone. Kv4.3 channels contribute to polarizing, outward potassium currents, which desensitize and limit neuronal activity. Reduced expression of Kv4.3 leads to increased neuronal excitability; however, the physiological importance of Kv4.3 in the RVLM has not been investigated during CHF. We hypothesized that overexpression of Kv4.3 in the RVLM of rats with CHF would reduce sympathetic tone. To test this hypothesis, rats with confirmed CHF were implanted with radiotelemetry and were bilaterally injected with 250 nL of adenoviral‐associated Kv4.3 (adKv4.3; n = 4) or GFP (adGFP; n = 3) into the RVLM. Telemetry recordings were collected for 14 days post‐injection followed by terminal/anesthetized baroreflex renal sympathetic nerve activity (RSNA) experiments. Mean arterial pressure remained the same in both groups following injection; however, heart rate was reduced after adKv4.3 relative to baseline (‐16.4 ± 2.7 bpm) but not adGFP (+6.1 ± 6.1 bpm; p < 0.05 between groups). Both heart rate variability as measured by the standard deviation of beat to beat intervals (10.4 ± 0.8 vs. 8.0 ± 0.3 ms; p = 0.09) and frequency‐domain analysis of LF/HF ratio (0.90 ± 0.13 vs. 1.66 ± 0.37) trended toward improvement in adKv4.3 compared to adGFP. Baseline RSNA trended lower (22. 8 ± 0.4 vs. 42.4 ± 8.5 % of max; p = 0.15), and the baroreflex sensitivity was higher (3.9 ± 0.4 vs. 1.43 ± 0.22 Gmax; p < 0.005) in adKv4.3 compared to adGFP. These data support the hypothesis that overexpression of Kv4.3 in the RVLM reduces sympathetic tone in the CHF state.
Restoration of Oxytocin Neuron Activity Prevents Hypertension Caused by Chronic Intermittent Hypoxia/Hypercapnia
Authors: Jameson Heather , Cauley Edmund , Mendelowitz David
Published: June 2021

New promising work has indicated oxytocin (OXT) receptor activation is blunted in cardiovascular diseases (CVDs) and OXT receptor activation may be a novel target to increase parasympathetic activity to the heart and treat CVDs. This study tests the hypotheses that the release of OXT from hypothalamic paraventricular (PVN) neurons onto cardiac vagal neurons (CVNs) is diminished with chronic intermittent hypoxia‐hypercapnia (CIH/H), an animal model of OSA, and furthermore that selective activation of PVN OXT neurons can reduce adverse cardiovascular consequences that occur with CIH/H. Optogenetic stimulation of channelrhodopsin expressing PVN fibers evoked large transient increases in Ca2+ in CHO cells stably transfected to express the human recombinant OXT receptor and the red fluorescent calcium indicator, R‐GECO1, dispersed adjacent to CVNs. The release of OXT onto CVNs upon photoactivation of PVN fibers was blunted in animals exposed to 21 days of CIH/H. To examine if restoration of OXT neuron activity can prevent the adverse cardiovascular effects of CIH/H in vivo, we selectively expressed excitatory DREADDs in PVN OXT neurons, and implanted animals with telemetry devices to monitor blood pressure (BP) and EKG activity. Selective chronic activation of PVN OXT neurons decreased resting BP and HR, and perhaps more importantly chronic PVN OXT neuron activation prevented the elevations in BP and HR that occur with CIH/H. These results indicate excitation of parvocellular PVN fibers releases OXT at brainstem CVN targets, and selective activation of OXT neurons in the PVN decreases resting BP and HR, and prevents the elevations in BP and HR that occur after 21 days of CIH/H.
Effects of Exercise Training and Intermittent Hypoxia on Ventilatory Chemoreflex Sensitivity in Humans
Authors: Leuenberger Urs , Ross Amanda , Sauder Charity , Blaha Cheryl , Cauffman Aimee
Published: June 2021

Exercise training exerts profound beneficial effects on cardiovascular function, reduces cardiovascular risk and improves outcome in patients who suffered cardiovascular events. The underlying mechanisms are incompletely understood but altered autonomic neural regulation may be involved. We recently found that short‐term intermittent hypoxia induced by 30 serial hypoxic breathholds raises sympathetic activity and blood pressure, and enhances chemoreflex sensitivity to hypoxia. Because chemoreflex activity is a marker of cardiovascular risk, here we tested the hypothesis that chemoreflex sensitization induced by intermittent hypoxia is attenuated by exercise conditioning. Eleven young healthy subjects (5M/6F, age 25±1 yrs, body‐mass‐index 24±1 kg/m2) were studied before and following 2 months of endurance training (peak VO2 pre vs. post: 36.4±1.4 vs. 40.1±1.7 ml/min/kg; P<0.05). Chemoreflex sensitivity was determined by the ventilatory responses to a series of 2‐8 breaths of N2 inhalation before and after 30 min of intermittent hypoxia (1 episode/min; O2 saturation nadir ~80%). Exercise training did not affect basal blood pressure or chemoreflex sensitivity but lowered resting heart rate. Pre training, intermittent hypoxia raised chemoreflex sensitivity from 0.261±0.032 to 0.305±0.032 L/min/%O2 desaturation (P<0.06). In contrast, post training chemoreflex sensitivity was unchanged by intermittent hypoxia (from 0.290±0.044 to 0.262±0.036 L/min/%O2 desaturation; P=NS). These data suggest that endurance training attenuates chemoreflex sensitization induced by intermittent hypoxia. Whether this is due to an antioxidant effect of exercise training requires further study. (Supported by HL098379)
Hypoxic‐Induction of Arginase II is Associated with EGFR Activation and EGF Production
Authors: Pool Caitlyn , Jin Yi , Chen Bernadette , Liu Yusen , Nelin Leif
Published: June 2021

We have previously shown that hypoxia induces arginase II through epidermal growth factor receptor (EGFR) activation in human pulmonary microvascular endothelial cells (hPMVEC). We hypothesized that hypoxia‐induced EGFR activation occurs via epidermal growth factor (EGF) release. Before studying hPVMEC, HeLa cells were used to dissect cellular mechanisms. Cells were grown in normoxia (21% oxygen) or hypoxia (1% oxygen) for 48 hours and mRNA harvested. RT‐PCR analysis showed substantially higher levels of EGF, EGFR, and arginase II mRNA in the cells incubated in hypoxia than in those incubated in normoxia. Cells were incubated in normoxia or hypoxia for 24 hours, the media harvested and placed on naive cells which were than incubated in normoxia for 48 hours. Cells incubated with hypoxia‐conditioned media had greater arginase II protein levels compared to cells incubated with normoxia‐conditioned media. Cells were then incubated in hypoxia or normoxia for 48 hours with either vehicle or the EGFR kinase inhibitor AG1478 added to the media, and AG1478 treatment prevented hypoxia‐induced arginase II protein expression. Finally, cells were incubated in hypoxia and normoxia for 48 hours with either IgG or an EGFR blocking antibody added to the media, the hypoxia‐induced arginase II expression was substantially lower in the cells incubated with the EGFR blocking antibody than in those incubated with IgG. These findings suggest that hypoxia‐induced arginase II expression may be dependent on EGF‐mediated activation of EGFR. We speculate that the EGFR pathway may be a novel therapeutic target for pulmonary hypertensive disorders associated with hypoxia.
Secreted Klotho Augments the Therapeutic Potential of Mesenchymal Stem Cells for Monocrotaline‐induced Pulmonary Arterial Hypertension
Authors: Varshney Rohan , Wu Chengxiang , Ali Quaisar , Sun Zhongjie
Published: June 2021

Objective

Klotho is an anti‐aging gene. The purpose of this study is to investigate if secreted Klotho (SKL) in synergy with mesenchymal stem cells (MSCs) improves monocrotaline (MCT)‐induced pulmonary vascular dysfunction and pulmonary arterial hypertension (PAH).

Methods & Results

We generated lentivirus carrying SKL‐GFP or GFP genes driven by a CMV promoter. MSCs were transfected with SKL‐GFP or GFP. Four groups of male rats were treated with MCT (60 mg/Kg, IP) while an additional group was given saline (control). Three days later, four MCT‐treated groups received IV delivery of MSCs, MSC‐GFP, MSC‐SKL‐GFP, and no treatment, respectively. MCT significantly increased right ventricular (RV) systolic blood pressure. Interestingly, MSC‐SKL‐GFP significantly attenuated the MCT‐induced increase in RV pressure (Saline 17.27 ± 4.86, MCT 31.88 ± 1.39, MCT + MSC‐SKL ‐GFP 23.35 ± 3.26). Ex vivo vascular relaxing responses to acetylcholine were decreased in small pulmonary arteries (PA) in MCT‐treated rats, indicating that MCT causes pulmonary vascular endothelial dysfunction. MSCs overexpressing SKL improved MCT‐induced PA endothelial dysfunction. MSC overexpressing SKL also significantly attenuated RV hypertrophy. MSCs alone, however, improved MCT‐induced PAH, PA endothelial dysfunction and RV hypertrophy but not to a significant level.

Conclusion

MSCs overexpressing SKL attenuated MCT‐induced PAH, PA endothelial dysfunction, and RV hypertrophy. SKL augments the therapeutic effect of MSCs in PAH.
Raptor and Rictor Both Contribute to the Development and Progression of Pulmonary Arterial Hypertension
Authors: Tang Haiyang , Chen Jiwang , Fraidenburg Dustin , Gu Yali , Garcia Joe , Makino Ayako , Yuan Jason
Published: June 2021

Pulmonary arterial hypertension (PAH) is a rare but progressive and fatal disease that results in increased pulmonary vascular resistance (PVR). The aim of this study is to examine whether mTOR complexes in pulmonary arterial smooth muscle cells (PASMCs) are involved in the pathogenesis of PAH.

Methods

The mice with smooth muscle cells conditional and inducible knockout of mTOR, Raptor or Rictor were used in hypoxia‐mediated Pulmonary Hypertension (PH) model. Right ventricle systolic pressures (RVSP) and right ventricular hypertrophy, by RV/ (LV+S) ratios, were measured. Animals were treated with dual inhibitor of mTORC1/2 for protective and reversal groups and PH will be assessed by RVSP and RVH measurements.

Results

mTOR conditional and inducible knockout mice had significantly lower RVSP, RV/LV+S ratios, and displayed less pulmonary vascular remodeling when exposed to hypoxia compared to their WT mice. Conditional deletion of Raptor or Rictor in smooth muscle cells was also found to prevent hypoxia‐induced PH and vascular remodeling. Treatment with dual inhibitor of mTORC1/2 attenuated the rise in RVSP and the development of RV hypertrophy in hypoxia‐induced PH.

Conclusion

Both Raptor and Rictor play significant roles in pulmonary vascular remodeling associated with PH. Dual inhibition of mTORC1/2 attenuated hypoxia‐induced PH. The PI3K‐Akt1‐mTOR signaling pathway will continue to be a focus of research aimed at determining factors which effect pulmonary vascular remodeling and focusing on both complexes of mTOR may lead to novel therapeutic targets for PAH.
Role for RUNX2/HDAC6 Axis in Pulmonary Arterial Hypertension
Authors: Ruffenach Grégoire , Chabot Sophie , Potus Francois , Tanguay Virginie , Vaillancourt Mylène , Tremblay Ève , Breuils‐Bonnet Sandra , Provencher Steeve , Bonnet Sébastien
Published: June 2021

Introduction

Pulmonary arterial hypertension (PAH) is a vascular remodeling disease characterized by enhanced pulmonary artery smooth muscle cells (PASMC) proliferation, migration, calcification and suppressed apoptosis. Numerous biological pathways have been implicated in this phenotype, including HIF‐1α. Recent studies have shown that miR‐204 downregulation upregulates the expression of RUNX2. RUNX2 is implicated in many features seen in PAH‐PASMC in part through the activation of HIF‐1α by a HDAC6‐depepdent mechanism. Thus we hypothesis that miR‐204‐dependent upregulation of RUNX2/HDAC6 axis promotes HIF‐1α and development of PAH.

Results

Using human lungs, distal PAs and isolated PASMC from both control and PAH patients, we demonstrated a significant upregulation of RUNX2 and HDAC6 in PAH. Gain and loss of functions experiments in PASMC showed that downregulation of miR‐204 in controls PASMC increase RUNX2 expression, while increases miR‐204 in PAH‐PASMC decrease it. In controls PASMC, RUNX2 upregulation increases calcification,migration, proliferation, resistance to apoptosis through an HDAC6 and HIF‐1α dependent mechanism, as both tubastatin A (HDAC6 inhibitor) and siHIF‐1α blocks RUNX2 effects. Similarly in PAH‐PASMC siRUNX2 has the opposite effects. Finally, in sugen/hypoxia rat model of PAH, nebulization of siRUNX2 decreased mean PA pressure, pulmonary vascular resistance and increased compliance and cardiac output.

Conclusion

Taken together, our study uncovers a new miR‐204 dependent up regulation of RUNX2/HDAC6 axis contributing to calcification and to normoxic activation of HIF‐1α leading to proliferation, migration and resistance to apoptosis in PAH‐PASMC.
Stimulated Lymphocytes from Patients with ARG1 Single Nucleotide Polymorphism rs2781666 have Augmented NO Production in Hyperoxia
Authors: Trittmann Jennifer , Jin Yi , Chen Bernadette , Nelin Leif
Published: June 2021

L‐arginine metabolism by arginase and NO synthase have opposing effects on vascular tone and remodeling, with arginase activity favoring vasoconstriction and vascular remodeling. Our group recently found that the arginase I (ARG1) single nucleotide polymorphism (SNP) rs2781666 is associated with a decreased risk of PH in infants with BPD. We hypothesized that the mechanism for protection against PH in BPD is due to lower levels of arginase I and thereby greater endogenous NO production. Lymphocytes from BPD patients were genotyped for the ARG1 SNP rs2781666 (GG‐wildtype, TT minor allele), stimulated with IL‐4, IL‐13, and phorbol myristate acetate (PMA), and exposed to either normoxia (21%O2) or hyperoxia (90%O2) for 24 hours. Cell lysates from lymphocytes were assayed for arginase I, arginase II, and cleaved caspase‐3 protein by western blot. The media was assayed for nitrites. Stimulated lymphocytes from patients homozygous for the ARG1 rs2781666 SNP exposed to hyperoxia have decreased levels of arginase I protein, no difference in arginase II protein, increased nitrite levels, and increased cleaved caspase‐3 protein, as compared to lymphocytes from patients homozygous for the wild‐type allele. These data are consistent with our hypothesis that BPD patients who have the ARG1 rs2781666 SNP are protected against PH during hyperoxic exposure by producing lower levels of arginase I and greater levels of endogenous NO. The increase in endogenous NO in these lymphocytes was associated with increased apoptosis as demonstrated by greater cleaved caspase‐3 activation.
Lung ACE is Not Involved in The Progression of Pulmonary Arterial Hypertension
Authors: Joshi Sachindra , Gupte Sachin
Published: June 2021

Rationale

Although ACE–Ang II–AT1R axis has been well documented in promoting vasoconstriction, proliferation, and fibrosis‐the hallmarks of pulmonary arterial hypertension (PAH), the role of lung ACE in the progression of PAH is not clear. Pharmacological inhibition of ACE to effectively attenuate the progression of PAH has been ambiguous. Therefore, we sought to determine if knocking out lung ACE would prevent the progression of PAH.

Methods

To test this hypothesis we evaluated development of PAH in ACE knock‐out (ACE.3) and wild type C57BL mice using Sugen 5416‐Hypoxia PAH model. Wild type and ACE knock‐out mice were exposed to 3 weeks of normoxia or chronic hypoxia and simultaneously administered the sugen 5416 at 0, 1, and 3 weeks. Pulmonary artery acceleration time (PAAT) was measured by transthoracic echocardiography on week 0 and week 3. Right ventricular hypertrophy was assessed by taking the RV/LV+S weight. Cardiac function was assessed by left heart catheterization using Millar 1.4 F catheter. Pulmonary arteriopathy was assessed from hematoxylin and eosin stained formalin fixed paraffin embedded lung sections.

Results

Significant decrease in PAAT in sugen hypoxia wild type and ACE knock‐out compared to vehicle or sugen normoxia control was similar. The degree of right ventricular hypertrophy in sugen hypoxia ACE knock‐out and wild type was not different. Decreased cardiac output between sugen hypoxia wild type and the ACE knock‐out was similar. The arteriopathy in sugen hypoxia wild type and ACE knock‐out were also similar, which included medial thickening, occlusive lesions and plexiform‐like lesions.

Conclusion

It appears that lung ACE may not be involved in the progression of the PAH.

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Use of AMP Protein Kinase as a Potential Molecular Marker to Investigate the Development of Obesity in Non‐Model Fish Species
Authors: Vides Melissa , Evans Paige , Kobayashi Yasuhiro
Published: June 2021

Objectives of this study were to identify the genes encoding subunits of AMP protein kinase (AMPK) in channel catfish and examine their tissue distribution. The AMPK is considered as the master regulator of tissue nutrient sensing and metabolism. Functions of AMPK have been investigated in association with obesity and diabetes development. In channel catfish, genetic selection toward growth causes development of obese‐like phenotype, suggesting channel catfish can be used as an alternative model to investigate development of obesity. However, little is known about the role of AMPK in channel catfish. In this study, genes encoding subunits of AMPK (alpha 1 and 2, beta 2, and gamma 1) were identified by comparing the channel catfish expressed sequence tag database with rainbow trout AMPK subunit mRNA. Expression of various AMPK subunit mRNA was examined in the cDNA of brain, liver, muscle, spleen, kidney and heart using RT‐PCR. Alpha 1 and 2 mRNA, as well as beta 2 mRNA, were detectable in all tissues examined. Gamma 1 mRNA expression was detectable in brain, muscle, spleen, kidney, and heart. The nucleotide sequence of amplicon corresponding to each subunit was highly similar to those of other fish (>70%). Currently, we are investigating changes in the expression of AMPK subunit mRNA in relation to changes in food intake and genetic selection toward growth in channel catfish. This study is supported by the Kansas IDeA Network of Biomedical Research Excellence.
Insulin and IGF‐1 exert distinct actions on the basolateral electrolyte fluxes in principal and intercalated cells of the collecting duct
Authors: Zaika Oleg , Mamenko Mykola , Boukelmoune Nabila , Pochynyuk Oleh
Published: June 2021

Insulin and insulin‐like growth factor‐1 (IGF‐1) are known to stimulate ENaC‐mediated sodium transport in the collecting duct (CD) cells. Yet, elevated insulin levels favor kaliuresis, whereas IGF‐1 causes renal NaCl retention. The mechanism causing this phenomenon remains unexplored. Electrogenic ENaC‐mediated Na+ transport in the collecting duct establishes driving force for Cl‐ reabsorption and K+ secretion. Basolaterally localized Kir4.1/5.1 channels facilitate sodium reabsorption and potassium secretion in principal cells. Chloride ClC‐K2 channel, expressed at the basolateral membrane of intercalated cells, mediates trans‐cellular Cl‐ transport in this tubular segment. Using patch‐clamp electrophysiology in freshly isolated CD, we found that insulin acutely stimulates basolateral potassium Kir4.1/5.1 channels in a PI3 kinase‐dependent manner in principal cells, but inhibits basolateral chloride ClC‐K2 channels in intercalated cells via a MAPK‐dependent pathway. On the contrary, IGF‐1 markedly activates ClC‐K2 channel in intercalated cells by activating PI3 kinase. Therefore, we conclude that insulin, by stimulating transport in principal cells only, favors coupling of Na+ reabsorption with K+ secretion at the apical membrane contributing to kaliuresis. On the other side, IGF‐1, by activating ENaC in principal cells and ClC‐K2 channels in intercalated cells, promotes net NaCl retention.
Cardiovascular, Metabolic and Renal T‐cell Profile of Obese C57Bl6 mice: Role of Renal Nerves
Authors: Asirvatham‐Jeyaraj Ninitha , Fiege Jessica , Han Ruijun , Foss Jason , Bartolomucci Alessandro , Razzoli Maria , Burbach Brandon , Shimizu Yoji , Osborn John
Published: June 2021

Hypertension (HTN) is a global epidemic and often occurs in concurrence with obesity and diabetes (i.e. metabolic syndrome). Recent clinical trials suggest that renal denervation (RDNX) decreases blood pressure (BP) and improves glucose metabolism in drug resistant hypertensive patients. Using a murine model we tested the hypothesis that obesity is associated with renal inflammation secondary to T cell infiltration, which activates renal sensory nerves and increases global sympathetic activity. 8‐week old C57Bl6 mice were fed either a low fat diet (LFD; 10 KCal% fat) or a high fat diet (HFD; 45 KCal% fat) for 10 weeks. Two parallel protocols were conducted. In a metabolic protocol, body weight, food intake, body composition and fasting blood glucose were measured. In a cardiovascular protocol, mice were implanted with radiotelemeters for measurement of BP. Mice on a HFD exhibited increased splenic T cells indicating peripheral inflammation, exhibited a metabolic syndrome phenotype including increased fat mass, HTN and hyperglycemia compared to LFD. RDNX but not Sham surgery after 12 weeks of HFD normalized BP (116 ± 4 in sham vs. 97 ± 6 mmHg in RDNX mice). RDNX had no effect on BP in LFD mice. Finally, RDNX had no effect on glucose metabolism as determined by glucose tolerance test. These results suggest that renal nerves play a role in obesity induced HTN but do not contribute to impaired glucose metabolism in this murine model. Future data analysis will test the hypothesis that renal inflammation drives renal afferent nerves and global sympathetic activity in obesity‐induced HTN. Supported by RO1HL116476.
Regulation of Sodium/bicarbonate Co‐transporter 1, NBCe1, in Murine Hippocampal and Cortical Astrocytes
Authors: Schroedl Magdalena , Roussa Eleni
Published: June 2021

The product of the Slc4a4 gene, the Na+/HCO3‐ cotransporter 1 (NBCe1), is a key pH regulating plasma membrane transporter and broadly expressed in several cell types. It is well established that depolarization‐induced alkalinization of glial cells is ascribed to uptake of bicarbonate via the electrogenic NBCe1, activated by the K+‐induced membrane depolarization. However, in contrast to epithelial cells the molecular pathways underlying NBCe1 transcriptional regulation and trafficking in neural cells are largely unknown. In the present study we examined regulation of astrocyte NBCe1 and elucidated the underlying signalling pathways. Expression, regulation, and activity of NBCe1 was examined in acute hippocampal slices and primary cultured hippocampal and cortical astrocytes using the 4AP (4‐aminopyridine) model and following extracellular pH changes in the presence or absence of inhibitors for Src, ERK, and JNK signalling pathways.

The results show upregulation of NBCe1 mRNA level, protein abundance, and surface expression in hippocampal slices after 4‐AP treatment, effects that were abolished following inhibition of JNK signalling pathway. Increased membrane expression of NBCe1 was also prevented after inhibition of Src/ERK signalling. In cultured hippocampal and cortical astrocytes, 4AP treatment and extracellular alkalosis caused activation of astrocytes and increased surface NBCe1 protein.

These data demonstrate regulation of NBCe1 transcript and surface expression, via Src/ERK and JNK signalling pathways.
Gas Exchange Estimate of Pulmonary Capacitance is Similar in Cystic Fibrosis Patients and Healthy Humans When Matched for Stroke Volume
Authors: Kasak Alexander , Wheatley Courtney , Snyder Eric , Baker Sarah
Published: June 2021

Pulmonary artery capacitance (PaCAP) is the ability of the pulmonary artery to accept a volume of blood at a given pressure. Measured invasively with heart catheterization; PaCAP can be estimated non‐invasively through the measurement of end‐tidal CO2 (PETCO2) and stroke volume (SV), known as gas‐exchange capacitance (GXCAP). We have shown that GXCAP is decreased at rest as well as during peak exercise in patients with cystic fibrosis (CF), when compared to healthy subjects. To determine whether GXCAP is different in CF and controls at a workload that matches for SV. 9 CF patients (age=22±3yrs, BMI=23±1kg/m2, VO2=19±1mL/kg/min, FEV1=81±8%pred.) and 13 healthy subjects (age=28±3yrs, BMI=25±1kg/m2, VO2=23±2L/min, FEV1=93±3%pred.) were studied. Exercise testing was performed on a cycle ergometer. PETCO2 was assessed using a metabolic cart, SV was calculated from heart rate and cardiac output (Q) using the acetylene rebreathe technique and GXCAP was calculated as the product of PETCO2 and SV. When matched for workload which sought to match SV, there was a significant difference in CF and healthy subjects in workload (Watts=72±6 vs. 90±4W), but no difference in percent of max watts (%MaxW=63±4 vs. 66±5W), Q (11±1 vs. 13±1L/min), or SV (89±4 vs. 90±4ml/beat). When matched, the previous difference in GXCAP in healthy and CF was abolished (GXCAP=32±2 vs. 32±2mL/BEAT • min). PETCO2 was similar between CF and healthy subjects (PETCO2 36±1 and 35±1mmHg). When matched for SV, GXCAP was statistically similar in CF. In relatively healthy CF there is no evidence of pulmonary vasculature dysfunction. NIH RO1 HL108962‐02
Crosstalk Between Retinoic Acid and Calcium Signaling Pathways Regulates Granulosa Cell Proliferation
Authors: Kipp Jingjing , Demczuk Michael , Huang Huiya , White Carl
Published: June 2021

Normal development of ovarian follicles is critical for female reproduction and endocrine function. We have identified retinoic acid (RA) and the RA‐degrading enzyme CYP26B1 as novel regulators of ovarian follicle development. The current study was designed to examine the role of intracellular Ca2+ signaling in mediating the effects of RA/CYP26B1 on primary mouse granulosa cell proliferation. In Ca2+ imaging experiments, treatment of cultured granulosa cells with RA was found to increase the steady‐state Ca2+ content of the endoplasmic reticulum (ER) stores. This correlated with increased store‐operated Ca2+ entry and enhanced IP3‐dependent Ca2+ release in response to purinergic stimulation. In proliferation assays, RA treatment or Cyp26b1 knockdown stimulated proliferation while Cyp26b1 overexpression or incubation with 2‐APB, a blocker of ER Ca2+ release and store‐operated Ca2+ entry, inhibited proliferation. These data support a model in which the RA regulates ovarian follicle development by stimulating granulosa cell proliferation that is at least partly driven by the modulation of Ca2+ signals mediated by increased ER Ca2+ store filling. Support for this work was provided by a DePaul‐RFUMS Collaborative Research Grant.
Collaborative Learning in the Undergraduate A&P Classroom
Authors: Mathis Karen , Schauer Daniel
Published: June 2021

The Collaborative Learning Assessment through Dialogue (CLAD) method has been shown to increase test scores in various types of courses. The purpose of this method is to promote active learning and social interaction, two factors that are seen as key in creating high achieving classrooms. Our hypothesis is that utilization of the CLAD method will increase scores on “traditional” tests taken during the A&P course thereby indicating greater retention of material and higher achievement. Students were randomly placed into groups. Students individually completed anticipation guides to foster preparation outside of class, took a solo quiz prior to lecture in class, and then took a group quiz, which was pre‐coded with the correct answer utilizing color changing markers, after lecture. Statistical analysis indicated that except for one test given during the entire A&P I and II course sequence, there was no difference in test scores in classes where CLAD was used compared to classes in previous years where it was not used. However, a survey of the students indicated that 75% felt the anticipation guide helped them to be better prepared for lecture, 90% liked the group discussion associated with the group quiz, 64% felt they learned more than they would have if CLAD had not been used and 71% recommended that the CLAD method be utilized in future A&P classes. Although the CLAD method does not appear to cause any improvement in “traditional” test scores, students believed that they were better prepared, more engaged and had an improved performance when this method was used.
Microbiological Investigation of Waters of Lake Përlepnica durnig Winter Season 2013
Authors: Ismaili Muharrem , Rashiti Fahrie , Kurteshi Yll , Sahiti Naim
Published: June 2021

Physicochemical and bacteriological analyses were carried out on lake water, used for drinking and swimming purposes in Përlepnica nearby Gjilani city, Kosovo.

The objective of this study is to assess the quality of water, of the Lake Përlepnica during winter season, 2013 year, through the microbiological analysis.

None of the samples complied with bacteriological standards as Total coliform counts generally exceeded 1,600 MPN/ml, and pathogen count such as Salmonella‐Shigella counts counts were very high.

Based on achieving results led us to conclude: The waters of Llake Përlepnica are high polluted by bacteria at all investigated localities(3).
Improving Longevity of Gelatin Ultrasound Trainers
Authors: Iceton Blake , Royer Danielle
Published: June 2021

Medical schools are increasingly integrating ultrasound training into their curricula as ultrasound utility expands in medical practice. Homemade ballistic gelatin ultrasound trainers have been developed as teaching tools that bypass the logistical complications of human test subjects and expense of commercial phantoms. This study explored methods to impede mold growth and extend the longevity of homemade trainers through chemical additives and physical barriers. Thirty‐five 500 mL forms with a base formula of gelatin (10% by weight) and Metamucil (2.5% by weight) in water were created in 750 mL Tupperware containers. Five had no additives or surface coverings; 8 contained the preservative Carolina's Perfect Solution; 7 contained the antiseptic surgical scrub Betasept; and 5 each had the exposed surface covered in plastic wrap, Duoderm bandages, or Tegaderm bandages. All containers were lidded, refrigerated at 4⁰ C and checked weekly for number and diameter of growing mold colonies. By week 6, all gelatin forms except those treated with Betasept or covered with Duoderm bandages had developed mold, with colony number ranging from 1‐4 and diameters from 1‐16 mm. While 2 of 5 Duoderm trials developed mold in week 10, all Betasept trials remained mold free through 25 weeks. Adding 1% Betasept by weight during gelatin trainer creation is recommended as a low cost, low hazard method for preventing mold growth.
Analysis of the Stapes of Krapina Neandertals
Authors: Hill Cheryl , Radovcic Jakov , Frayer David , McGechie Faye
Published: June 2021

Auditory ossicles are found rarely in the fossil hominid record, but the available ossicles allow for some inferences about the hearing abilities of fossil hominids. Previously three Neandertal stapes in varying degrees of preservation have been described. Two complete stapes have been identified in computed tomography scans of the temporal bones of Neandertals from the Krapina site in Croatia. The purpose of this project is to complete a descriptive and quantitative analysis of the stapes from Krapina to compare them to the stapes of other extant and fossil hominids.

The sample consists of three‐dimensional reconstructions of the stapes from the Krapina site (N=2). Height of the stapes, length of the crura, and the dimensions of the stapedal foot plate were acquired for each stapes reconstruction using Avizo software. These dimensions were compared to previously published values for extant and fossil hominids.

The analysis indicates that the stapes from the Krapina site show asymmetry of the crura. The measured height of the stapes and the size of the stapedal footplate are smaller than recent H. sapiens. These results suggest that Krapina Neandertals may have had increased low frequency sensitivity.
Association Between The Shapes of Hindlimb Bone Articular Faces And Locomotor Behaviors in Artiodactyls
Authors: Dewar Eric , Grocott Olivia , Leclerc Mari
Published: June 2021

Habitat plays a role in selection for successful predator‐escape behaviors. Mammals that live in open habitats can use long‐distance running to escape predators, but in more closed habitats they would encounter barriers that require them to trade off speed for maneuverability. We examined differences in limb proportions of 31 living species of artiodactyls from various habitats (n = 94)—including species from grasslands, forests, forest‐edge habitats, and montane areas—along with some Eocene‐age artiodactyls. We examined articular faces of hindlimb bones and compared them with these species' habitats. We interpreted differences in limb morphology to be related to each species' habitat, due to selection for divergent predator‐escape behaviors. Visual differences in the shape of the proximal end of the tibia were recognized to be distinct for each of the habitats studied, but the shape of the distal end of the femur was not noticeably related to habitat. Species that lived in more open habitats had a broader tibial tuberosity and a more rounded articular margin for the femoral condyles. In contrast, species in closed habitats tended to have more pointed tuberosities with a jagged condylar margin. Mountain species were variable in the shape of the tibial tuberosity with a distinctly posterior orientation to the condyles. Closed/edge habitats had the shortest tuberosities and the most rounded condyles. Our Eocene artiodactyls showed morphologies that reflected closed and closed‐edge habitats.
Delineating the Cellular Hierarchy of Glioblastoma and Gliosarcoma for Identification of Therapeutic Targets
Authors: Hassiotou Foteini , Jackson Melanie , Seymour Tracy , Nowak Anna
Published: June 2021

Glioblastoma (GBM) and its more aggressive variant gliosarcoma (GSM) are the most lethal primary brain tumours, containing glioma stem cells (GSCs) which are resistant to current treatments. We compared the cellular hierarchy and molecular heterogeneity of GBM and GSM to identify genetic drivers of tumour growth and potential therapeutic targets. Fresh frozen human GBM and GSM specimens and 6 cell lines were interrogated using qRT‐PCR for expression of genes controlling self‐renewal and multilineage differentiation. SOX2 was overexpressed, whereas its core partners OCT4 and NANOG showed 90‐180 fold lower expression. Downstream targets ESRRB, PROX1, NESTIN and PAX6 had variable expression. GSC‐associated marker CD133 was highly expressed in both frozen GBM and GSM, but less so in the cell lines. Neural genes MAP2 and GAP43 and glial genes GFAP and S100B showed variable expression. Epithelial signatures (cytokeratin 18) were detected in GSM and to a lesser extent in GBM. Expression of E‐cadherin was significantly higher in GBM than in GSM (P<0.05), suggesting greater cell motility in GSM. Endodermal signatures (GATA4) were more highly expressed in GBM. Overall, GSM showed greater variation in gene expression than GBM. The presence of neural, epithelial and endodermal signatures suggests multipotential of GSCs. The stronger presence of different lineages in GSM reflects its greater heterogeneity, providing further evidence that GSM is a different disease to GBM and highlighting the need for patient‐tailored treatments.
Knockdown of Cardiolipin Synthase in Human Brain Microvessel Endothelial Cells Modulates Blood Brain Barrier Transport Properties
Authors: Nguyen Hieu , Wang Ying , Miller Donald , Hatch Grant
Published: June 2021

The blood brain barrier (BBB) is formed by microvessel endothelial cells and is a restrictively permeable interface that allows transport of only specific compounds into the brain. Cardiolipin (CL) a mitochondrial (MITO) lipid is required for activity/integrity of the respiratory chain. We examined the role of CL in maintaining barrier properties of brain microvessel endothelial cells. We show that human brain capillary endothelial cells (hCMEC/D3) produce a significant proportion of their basal ATP requirement through MITO oxidative phosphorylation. siRNA knockdown (KD) of the CL biosynthetic enzyme CL synthase (CLS) significantly reduced these cells ability to increase MITO function under conditions of increased metabolic demand compared to controls. In addition, KD of CLS acted as a metabolic switch causing hCMEC/D3 cells to become more dependent on glycolysis. KD of CLS increased [3H]‐2‐deoxyglucose ([3H]‐2DG) uptake compared to controls, and this may thus serve as a mechanism to compensate for the cells reduced MITO energy production efficiency. Finally, KD of CLS did not affect movement of the permeability marker fluorescent dextran across confluent hCMEC/D3 monolayers grown on Transwell®plates, an in vitro model for the human BBB, but caused in an increase in [3H]‐2DG transport across the monolayer compared to controls. The results suggest that brain endothelial MITO energy status play a role in regulating glucose transport across the human BBB. These observations may, in part, explain why brain capillary endothelial cells possess higher MITO content than endothelial cells derived from non‐BBB regions of the body. (Supported by NSERC and the CRC Program)