bluuebunny/crossref_metadata_2025 huggingface dataset:
Decreased Telomerase Activity Converts the Mechanism of FMD from NO to H2O2 in Human and Mouse Arterioles
Authors: Beyer Andreas M. , Gutterman David D.
Published: June 2021

Reactive oxygen species (ROS) are important modulators of vascular reactivity. In disease, the mechanism of endothelium‐dependent vasodilation can shift from NO to hydrogen peroxide (H2O2). Reduced telomerase activity (TA) has been linked to increased ROS production. Animals lacking telomerase develop hypertension but the underlying effects on the vasculature are unknown. We hypothesized that decreases in TA are associated with elevated ROS and a change in the mechanism of endothelium‐dependent dilation from NO to H2O2. We postulate that this shift in dilator mechanism is conserved across species (mouse to human).

Murine mesenteric arteries (~200 μm) were isolated and flow‐mediated dilation (FMD) was studied ex‐vivo (pressure gradient 0–100 cm H2O). Inhibition of TA shifted the mediator of FMD from NO to H2O2 (% Max dilation at 100 cm H2O: Vehicle 72±9; +L‐NAME −8±8; +Catalase 92±6; TA inhibitor BIRB1532 10 μM 12–15 hours 84±7; +L‐NAME 81±10*; +Catalase −7±8* n≥4; * P <0.05 one way ANOVA vs. Vehicle). Resistance vessels (~200 μm) from the human adipose responded in a similar manner. Decreased TA shifted the mechanism of FMD from NO to H2O2 (% Max dilation at 100 cm H2O: Vehicle 88±3; +L‐NAME −39±2; +Catalase 83±3; BIRB1532 88.7±6; +L‐NAME 85±2*; +Catalase 34±5* n≥3; * P <0.05 one way ANOVA vs. Vehicle).

We conclude that telomerase may play a critical role in acute modulation of microvascular function across species.
PKC activation impairs endothelium‐dependent NO‐mediated dilation of coronary arterioles via enhanced superoxide production from xanthine oxidase
Authors: Thengchaisri Naris , Ren Yi , Hein Travis W. , Kuo Lih
Published: June 2021

We examined the role of PKC‐mediated pathway in eliciting endothelial dysfunction using isolated and pressurized porcine coronary arterioles. Diameter changes were recorded using videomicroscopic techniques. Superoxide production was detected with dihydroethidium (DHE) staining. Incubation of coronary arterioles with the PKC activator phorbol 12, 13‐dibutyrate (PDBu) (1 nM, 60 minutes) attenuated arteriolar dilation to the endothelium‐dependent, NO‐mediated agonists adenosine and serotonin, but not the NO donor sodium nitroprusside. In the presence of superoxide scavenger TEMPOL (1 mM), xanthine oxidase inhibitor allopurinol (10 μM), c‐Jun N‐terminal kinase (JNK) inhibitor SP600125 (5 μM) or Rho‐kinase inhibitor Y‐27632 (0.1 μM), the adverse effects of PDBu on adenosine‐ and serotonin‐induced dilations were prevented. Nevertheless, the inhibitory effects of PDBu were insensitive to H2O2 scavenger PEG‐catalase (500 units/ml), NAD(P)H oxidase inhibitor apocynin (100 μM) or p38 kinase inhibitor SB203580 (0.1 μM). DHE staining revealed that PDBu generated TEMPOL‐sensitive superoxide production in the coronary arteriolar smooth muscle. In conclusion, activation of vascular PKC with PDBu inhibits endothelium‐dependent, NO‐mediated dilation in coronary arterioles by producing superoxide from xanthine oxidase, which appears to be linked with JNK and RhoA/Rho kinase activation.
Stabilin‐1 protects against adverse inflammation and injury during viral myocarditis
Authors: Papageorgiou Anna-Pia , Carai Paolo , Verhesen Wouter , Velthuis Sebastiaan , Lutgens Ester , van Rooijen Nico , Swinnen Melissa , Kzhyshkowska Julia , Carmeliet Peter , Heymans Stephane
Published: June 2021

Stabilin‐1 is a transmembrane glycoprotein expressed on activated macrophages and endothelial cells of new capillaries during tissue remodeling. In the coxsackie B3 (CVB3) murine model of viral myocarditis, stabilin‐1 expression increased significantly and was primarily expressed in macrophages. This was also seen in endomyocardial biopsies from patients with fulminant myocarditis, where stabilin‐1 was expressed by CD‐68+ macrophages, forming a protective barrier around the healthy tissue. When challenged with the CVB3 virus, Stabilin‐1 null mice showed increased cardiac related mortality as compared to wild‐type mice (p<0,001, n=19). Histological analysis showed that the stabilin‐1 null mice had increased cardiac necrosis due to CD3+ T‐lymphocyte infiltration, and lower F4/80+ macrophages as compared to their wild‐type littermates, suggesting that stabilin‐1 may be important for monocytes recruitment. Thioglycolate induced peritonitis in stabilin‐1 WT and null mice also resulted in fewer monocytes recruited. Depletion of circulating monocytes with clodronate liposomes in wild‐type mice induced a similar phenotype after CVB3 infection. We identify stabilin‐1 as an important upregulated protein during VM in both mice and humans and that absence of stabilin‐1 results in increased cardiac inflammation, injury and mortality, demonstrating a novel cardio‐protective effect of stabilin‐1.
Reduced LDL oxidation is secondary to protection from in vivo thiol oxidation and hypocholesterolemia in Gilbert's syndrome
Authors: Bulmer Andrew Cameron , Boon Ai-Ching , Hawkins Clare , Bisht Kavita , Coombes Jeff , Wagner Karl-Heinz
Published: June 2021

The physiological importance of endogenous bilirubin remains unknown. This study aimed to assess whether persons expressing Gilbert's syndrome (GS, i.e. unconjugated hyperbilirubinaemia) are protected from thiol oxidation and to determine if this and their improved lipoprotein profile, explained reduced oxidized lipoprotein (oxLDL) in them. Forty‐four matched GS and control subjects were recruited. Blood was assayed for lipid profile, multiple plasma antioxidants and measures of oxidative stress. Elevated plasma reduced thiols (8.03±1.09 vs. 6.75±1.39 nmol/mg protein; P<0.01) and reduced glutathione existed in GS subjects (12.7±2.39 vs. 9.44±2.45 ìM; P<0.001). Reduced:oxidized glutathione was significantly improved in GS (0.49±0.16 vs. 0.32±0.12; P<0.001). Protein carbonyl concentrations were negatively associated with bilirubin concentrations and were significantly lower in persons with >40 ìM bilirubin versus controls (P<0.05). Furthermore, oxLDL was lower in GS subjects. Forward stepwise regression analysis revealed that bilirubin augmented the GSH:GSSG ratio, which, in addition to reduced circulating LDL, decreased oxLDL concentrations within the cohort. These findings implicate bilirubin's physiological importance in reducing thiol and subsequent lipoprotein oxidation, in addition to reducing circulating LDL concentrations.
Local adiponectin production in skeletal muscle resistance arteries: effects of exercise and shear stress
Authors: Sapp Glenn Herod , Chen Bei , Gurovich Alvaro N. , Muller-Delp Judy M.
Published: June 2021

Adiponectin is as an adipose‐derived secretory protein with anti‐inflammatory and anti‐atherosclerotic effects in the endothelium and vascular smooth muscle. Paradoxically, despite being secreted from adipose tissue, plasma adiponectin correlates inversely with obesity and obesity‐associated cardiovascular disease. The purpose of the current study was to determine whether the endothelium of resistance arteries is a local source of adiponectin. We also investigated the possibility that exercise training upregulates adiponectin in rat skeletal muscle resistance arteries. Real‐time qRT‐PCR was used to assess mRNA for adiponectin in both endothelium‐intact and ‐denuded arteries, and in cultured endothelial cells exposed to shear stress. Adiponectin mRNA was also assessed in skeletal muscle resistance arteries isolated from sedentary and exercise trained rats. Adiponectin mRNA was detected in both intact and denuded arteries. Shear stress increased adiponectin mRNA in cultured endothelial cells, and exercise training increased adiponectin expression in skeletal muscle resistance arteries. These data indicate that both the endothelium and the vascular smooth muscle of skeletal muscle resistance arteries are local sources of adiponectin. Exercise training increases adiponectin expression in skeletal muscle resistance arteries, possibly through a shear stress‐sensitive mechanism.
Effect of aerobic exercise on capillary and vascular endothelial growth factor in rat soleus muscle
Authors: Murakami Shinichiro , Fujita Naoto , Kondo Hiroyo , Tanaka Masayuki , Sakita Masahiro , Fujino Hidemi
Published: June 2021

Aerobic exercise has been known an effective therapy against the muscle endurance fitness. And skeletal muscle is classified into 2 types, slow or fast muscle. Slow muscles have rich oxidative utilization and rich capillaries. And rat soleus muscle is a slow muscle.

PURPOSE

To clarify the effect of aerobic training exercise on capillary architecture in slow muscle, the 3‐D capillarity has been investigated.

METHODS

Wistar rat (Con) rats and age matched Wistar rat that performed aerobic exercise preconditioning for 3 weeks (EX) were used. Contrast medium‐injected capillary in soleus muscle was visualized using a confocal laser microscope. And vascular endothelial growth factor (VEGF) expression was measured.

RESULTS

VEGF expression in EX was significantly higher than that in Con. Number of capillaries and number of anastomoses in EX were increased than Con. Capillary diameter in soleus of EX rat was thicker than that of Con. And mean capillary volume in EX was significantly higher than that in Con.

CONCLUSION

Results of present study indicate that aerobic training builds capillarity architecture in soleus muscle more closely. This study was supported by Grantsin‐ Aid for Scientific Research from the Japanese Ministry of Education, Culture, Sports, Science and Technology no. 23650340, 23650324, 23700600, 23700932 and 22300189.
Hypertension development in ethanol treated rats could involve baroreflex activity
Authors: da Silva Andreia Lopes , Crestani Carlos C , Scopinho America A , Correa Fernando MA , Elias Lucila LK , Antunes-Rodrigues Jose , Resstel Leonardo BM
Published: June 2021

This study evaluated MAP, HR and baroreflex responses in unanesthetized male wistar rats maintained under ethanol treatment (1st: 5%, 2nd: 10%, 3rd and 4th: 20%). After this period water was replaced for one week. During the treatment rats had a progressive increase of MAP since the 1st week, without alteration of HR. This increase was sustained until the 4th week and returned to normal after withdrawal. The baroreflex analysis with linear regression indicated no difference between slopes of bradycardic and tachycardic responses on the 1st and 2nd weeks. However, after 3rd and 4th weeks it was observed alteration between slopes of bradycardic (3rd t=2.27, p<0.05; 4th t=4.76, p<0.01) and tachycardic responses (3rd t=2.41, p<0.05; 4th t=3.65, p<0.01). After withdrawal linear regression analysis indicated no difference between slopes of bradycardic and tachycardic responses. These results indicate that ethanol treatment promotes a mild hypertension being this effect dependent on ethanol consume. Moreover, the baroreflex analysis showed a possible increase of sympathetic activity after 3rd week, which could involve MAP increases in ethanol treated rats.

Financial Support: FAPESP.
Aerobic Training (T) Improves Cerebral Circulation of Normotensive (WKY) and Spontaneously Hypertensive Rats (SHR)
Authors: Jordão Maria Tereza , Cavalleri Marina Tuppy , Burgi Katia , Ceroni Alexandre , Michelini Lisete Compagno
Published: June 2021

We compared the effects of hypertension and T in brain macro and microcirculation. Rats were submitted to treadmill T (55% of maximum capacity, 1h/day, 5 d/week) or kept sedentary (S) for 3 months. After hemodynamic measurements at rest, rats were anesthetized for iv. administration of FITC‐dextran (capillary density) or 4% PFA perfusion (morphometry of cerebral arteries). Brains were removed, post‐fixed and sliced for analysis of the capillary density within the NTS (25 μm) and of the basilar (BA), middle (MCA) and posterior cerebral (PCA) arteries (10 μm) (Image ProPlus). Compared to WKY, SHR exhibited elevated MAP and HR (180±2 mmHg, 359±6 b/min), increased wall/lumen ratio (+40%, +100% and +80% for BA, MCA and PCA, respectively), but no NTS capillary rarefaction. T was equally effective to improve treadmill performance and to cause resting bradycardia (−9%) in both groups; pressure fall (−8%) was only observed in the SHR. T reduced wall/lumen ratio only in the SHR (BA= −40%, MCA= −47%; PCA= −37%) and caused marked increase in capillary density in medial, intermediate and commissural NTS in both groups (on average 7.05 ±0.18 and 7.07±0.88%, corresponding to changes of +55% and +43%, WKY and SHR, respectively). Data showed that T improves brain perfusion by causing eutrophic outward remodeling in SHR brain arteries and capillary angiogenesis in both groups. Financial Support: CAPES, FAPESP
The effects of a 5 second bend over maneuver on cerebral perfusion and autoregulation upon standing from squat
Authors: Bentley Robert F , Walsh Jeremy J , Kellawan J Mikhail , Tschakovsky Michael E
Published: June 2021

PURPOSE

To investigate the effects of maintaining a bent over posture (BOP) for 5 s upon rising from squat on cerebral autoregulation.

METHODS

8 healthy subjects (24.4 ± 1.9 years) completed 1 min of quiet standing, followed by 1 min of squat, followed by rising from squat over a 1–2 s period and then no maneuver (CON) or a BOP maneuver for 5 s before standing upright. Middle cerebral artery blood velocity (MCABV (cm/s); Transcranial Doppler) and mean arterial blood pressure at heart level (MAP (mmHg); finger photoplethysmography) were continuously measured. Cerebral vascular conductance (CVC) was calculated from MCA blood pressure (MCABP) and MCABV. MCABP was estimated by accounting for the hydrostatic column between the heart and the MCA.

RESULTS

Data are mean Δ from bsl ± SD. BOP attenuated the decrease in MCABV during the first 5 s upon rising from squat compared to CON (−6.5 ± 2.5 vs. −12.1 ± 3.1, P<0.05). MAP hypotension during the same time period was abolished in BOP (+3.8 ± 10.0 vs. −12.9 ± 4.5, P<0.05). Upon restoring upright posture in BOP, the MAP nadir ~10 s after rising was virtually identical but the drop in MCABV was attenuated in BOP, indicating CVC was functionally increased (+0.41 ± 0.3 vs. +0.2 ± 0.1, P=0.1).

CONCLUSIONS

The BOP maneuver blunts disturbances to cerebral perfusion following rising from squat during the 5 s that it is maintained and enhances cerebral vasodilation at the MAP nadir after rising. NSERC.
Interactions between sorbitol pathway of glucose metabolism and 12/15‐lipoxygenase activation in diabetic peripheral neuropathy
Authors: Stavniichuk Roman , Obrosova Irina G
Published: June 2021

Both sorbitol pathway of glucose metabolism and, recently, 12/15‐lipoxygenase (12/15‐LO) activation have been implicated in the pathogenesis of diabetic peripheral neuropathy (DPN). Activation of the key enzyme of the sorbitol pathway, aldose reductase, is known to lead to NF‐κB activation and accumulation of cytosolic Ca++ i.e., two events implicated in 12/15‐LO overexpression and activation. We, therefore, evaluated the relationship between the two mechanisms using a pharmacological approach. Male C57Bl6/J mice were made diabetic with streptozotocin, after which non‐diabetic and diabetic mice were treated with the aldose reductase inhibitor fidarestat, at 16 mgkg−1d−1, in the drinking water, for 12 weeks. Diabetic mice displayed accumulation of glucose, sorbitol pathway intermediates, as well as 12/15‐LO overexpression and 12(S)‐HETE accumulation (a sign of increased 12/15‐LO activity) in two tissue‐targets for DPN, sciatic nerve and spinal cord. Fidarestat treatment did not affect diabetes‐induced glucose accumulation and completely corrected sorbitol and fructose accumulation in both tissues. It did not affect sciatic nerve or spinal cord 12/15‐LO overexpression; however, 12(S)‐HETE concentrations were either completely (sciatic nerve) or partially (spinal cord) corrected. In conclusion, increased sorbitol pathway activity is responsible for activation, but not overexpression, of 12/15‐LO in the pathogenesis of DPN. Studies in dorsal root ganglia are in progress.
Estimation of an optimum intake level of epigallocatechin gallate for suppression of inflammation in type 2 diabetic GK rats
Authors: Goda Toshinao , Uchiyama Yumiko , Tan Yo , Mochizuki Kazuki
Published: June 2021

Previous studies have suggested that epigallocatechin gallate (EGCG) exerts anti‐oxidative and anti‐inflammatory actions in various tissues, which might be beneficial for reducing risks of development of diabetes. However, it is unknown whether there is an optimum intake level of EGCG. In this study, we have examined the effect of a diet containing various concentrations of EGCG on the expression of inflammation‐related genes in various tissues including visceral adipose tissue and the liver of non‐obese type‐2 diabetes animal model, Goto‐Kakizaki (GK) rats. GK rats at 5 weeks of age were fed a control high‐fat diet (45 energy% as fat) or the high‐fat diet containing 0.1%, 0.2% or 0.5% EGCG for 34 weeks. The mRNA levels of IL‐1β, IL‐18, TNF‐α and resistin were significantly reduced in the adipose tissue of rats fed a diet containing 0.1% EGCG, but not in those fed diets containing 0.2% or more EGCG, as compared with control. The mRNA levels of IL‐1β and IL‐18 in the liver of rats fed a diet containing 0.1% EGCG were also significantly lower than those in the control. These results suggest that there is an optimum range of intake of EGCG, which may suppress the expression of genes involved in inflammation in the adipose tissue and the liver.
Insulin resistance is associated with reduced levels of the age‐related protein SIRT2
Authors: Lemos Vera , Oliveira Rita Machado , Outeiro Tiago F. , Gomes Pedro
Published: June 2021

Insulin resistance is a key feature of type 2 diabetes and the metabolic syndrome. Insulin resistance and oxidative stress seem to be intimately intertwined. Thus, we sought to explore the possible role of sirtuin 2 (SIRT2), reportedly having antioxidant activity, on improving insulin sensitivity in ob/ob mice and human HepG2 cells. We found that livers of ob/ob mice had increased levels of oxidative stress assessed by elevated hydrogen peroxide (H2O2) production, activation of stress‐sensitive kinases and protein oxidative modifications and this was accompanied by downregulation of SIRT2 and SIRT2‐regulated antioxidant enzymes superoxide dismutase 2 (SOD2) and catalase. Treatment of HepG2 cells with glucosamine (GlcN) impaired insulin‐stimulated glycogen synthesis and induced increases in oxidative stress levels. Pre‐treatment with the antioxidant N‐acetyl cysteine decreased H2O2 levels and improved insulin sensitivity. Furthermore, GlcN induced a reduction in SIRT2 protein levels in a time‐dependent manner. Taken together, our results suggest a link between insulin resistance and diminished levels of SIRT2 protein. Ongoing studies will determine whether enhancement of SIRT2 expression and/or activity could be a strategy for improving insulin sensitivity and subsequently prevent complications associated with the metabolic syndrome. Supported by grant PEst‐OE/SAU/UI0038/2011 (FCT).
Acute dietary nitrate consumption improves glucose tolerance
Authors: Scalzo Rebecca Lynn , Peltonen Garrett L. , Schweder Melani M. , Szaller Steve E. , Binns Scott E. , Wood Lacey M. , Klochak Anna L. , Bell Christopher
Published: June 2021

Insulin resistance is characterized by low nitric oxide (NO) bioavailability. Consistent with this observation, insulin sensitivity is improved with stimulation of NO generating pathways. Consumption of dietary nitrate (e.g. beet juice) has been purported to increase NO formation, mediated in part through the reduction of nitrate (NO3−) to nitrite (NO2−) by oral bacteria, and subsequent NO2− reduction to NO in blood. We hypothesized that acute dietary nitrate consumption would improve glucose tolerance. Accordingly, in 8 sedentary adults (23 ± 0 years, body mass index: 26.0 ± 0.8 kg/m2 (mean ± SE)) we determined the blood glucose response to oral consumption of 500 ml organic beet juice, supplemented with 25 g of sucrose (estimated carbohydrate load ~ 75g), with and without prior use of an antibacterial mouthwash to inhibit NO3− reduction to NO2−. In the absence of inhibition of oral bacteria NO3− reductase activity, peak blood glucose concentration (106 ± 4 vs. 118 ± 6 mg/dL; P = 0.04) and the area under the blood glucose response curve (93 ± 4%; P = 0.07) were decreased. These preliminary data suggest dietary nitrate consumption may promote glucose tolerance, and this effect may be attenuated following inhibition of NO3− conversion to NO2−.

Support: America Diabetes Association 1‐09‐RA‐09
Anti‐diabetic effect and acute toxicity of the water extract of Cyperus rotundus L. in rats
Authors: Krisanapun Chutwadee , Wongkrajang Yuvadee , Temsiririrkkul Rungravi , Kongsaktrakoon Boontium , Peungvicha Penchom
Published: June 2021

Herbal medicine is a complementary and alternative medicine which spread in many conditions/diseases, including diabetes mellitus. Cyperus rotundus L. in Cyperaceae family is a traditional herb found in Thailand. Although, some activities such as diuresis, flatulence and skin diseases had the scientific support, anti‐diabetic effect is not yet exhibited. So, the aim of this study was to examine the anti‐diabetic, toxicity and 2 mechanisms of action of C. rotundus's rhizome. The extraction of the rhizome with water got the yield around 10% w/w of dry plant. The results of the oral glucose tolerance test revealed that the extract 0.5 g/kg could significantly reduce the plasma glucose levels of either type 1 or type 2 diabetic rats comparing with control diabetic group. The toxicity test could summarize the single oral LD50 was more than 5 g/kg. The mechanism testing demonstrated that the extract at a dose of 5 mg/ml could inhibit the intestinal glucose absorption significantly in everted sac model and the extract since 1 mg/ml could enhance the glucose utilization of muscle like insulin did. We can conclude that the C. rotundus extract has a hypoglycemic or anti‐diabetic effect by inhibiting intestinal glucose absorption and promoting glucose consumption. Thus, it is very interesting for developing the extract to, at least, adjuvant for diabetes patients in the future. This work was support by Mahidol University, Thailand.
Renal responses to severe haemorrhage in conscious lambs
Authors: Smith Francine Gabriel , Samhan Mohamed , Qi Wei
Published: June 2021

The role of the kidney in the physiological responses to severe degrees of haemorrhage beyond which compensatory mechanisms fail (>25% of vascular volume) has not been investigated in the newborn period. In this study, parameters of glomerular and tubular function were measured for 30 min before (Control, C) and 120 min after haemorrhage of 30% of vascular volume, in conscious, chronically instrumented lambs aged ~one (N=10) and ~six weeks (N=9). Renal plasma flow decreased 30 min after haemorrhage from 1.4±0.5 SD (C) to 0.7±0.2 mL.min−1.g−1 at one week and from 1.6±0.4 (C) to 0.9±0.4 mL.min−1.g−1 at six weeks and remained decreased at 120 min. Similarly, glomerular filtration rate decreased from 0.26±0.08 (C) to 0.18±0.14 mL.min−1.g−1 at one week and from 0.31±0.08 (C) to 0.17±0.08 mL.min−1.g−1 at six weeks, and remained below C at 120 min. Urinary flow rate also decreased from 3.8±1.6 (C) to 1.8±0.7 uL.min−1.g−1 by 30 min after haemorrhage at one week and from 3.9±1.5 (C) to 2.2±0.8 uL.min−1.g−1 at six weeks, and remained below C at 120 min. Clearances of Na+ and K+ also decreased after haemorrhage in both age groups. These data provide evidence that soon after birth, glomerular and tubular responses to severe blood loss are elicited, though they do not appear to be developmentally regulated.

[Supported by the Canadian Institutes of Health Research.]
Blood pressure and renal hemodynamics in hypertensive Cyp1a1‐Ren2 transgenic rats fed a sodium‐deficient diet
Authors: Collins Amy E , Howard Catherine G , Mitchell Kenneth D
Published: June 2021

The present study was performed to determine whether dietary sodium restriction attenuates the development of hypertension and the impairment in renal hemodynamics that occur during the induction of the Ren2 gene in Cyp1a1‐Ren2 rats. Male Cyp1a1‐ Ren2 rats (n=5–6/group) were fed either a normal salt (0.6%) or salt‐deficient diet (<0.01%) containing indole‐3‐carbinol (I3C; 0.15%, wt/wt) for 16 days to induce slowly progressive ANG II‐dependent hypertension. The rats were then anesthetized with pentobarbital sodium and surgically prepared for measurement of mean arterial pressure (MAP), renal hemodynamics and renal excretory function. There were no significant differences between the induced normal salt group and the induced salt‐deficient group regarding MAP (183±4 vs. 181±5 mmHg), RVR (33.7±2.4 vs. 31.1±3.8 mmHg/mL/min.g), RPF (2.78±0.20 vs. 2.91±0.35 mL/min.g), or GFR (0.90±0.06 vs. 0.91±0.08 mL/min.g). Both groups had similar urine flows (41.1±1.3 vs. 39.0±6.9 μL/min), urinary sodium excretions (2.9±0.5 vs. 2.4±0.9 μEq/min) and fractional sodium excretions (0.94±0.15 vs. 0.76±0.22%). These data demonstrate that a sodium‐deficient diet does not attenuate the increase in arterial blood pressure or the associated renal functional derangements in Cyp1a1‐Ren2 transgenic rats with slowly progressive ANG II‐dependent hypertension.

Support: NHLBI, Tulane COBRE in Hypertension and Renal Biology.
Pravastatin Stimulates Proliferation of Endogenous Cardiac Progenitor Cells in the Heart
Authors: SUZUKI GEN , Palka Beth A. , Canty John M.
Published: June 2021

Background

While the HMG‐CoA reductase inhibitor pravastatin mobilizes cKit+ bone marrow‐derived progenitor cells and improves regional function in hibernating myocardium, the effect of pravastatin on resident cardiac progenitor cells (CPCs; cKit+/CD45‐ cells) is unknown.

Methods

Swine with ischemic cardiomyopathy were treated with pravastatin (80mg/day, n=10) for 4 weeks and compared to untreated controls (n=6). CPCs differentiating into a myocyte lineage and proliferation were quantified by immunohistochemical colocalization with GATA4, cardiac troponin I (cTnI) and Ki67. Differentiation and proliferation pathways were assessed by western blot analysis for the Notch ligand Jagged1.

Results

Pravastatin increased regional function (LAD wall thickening: 29±3% to 45±5%, p<0.01). Pravastatin increased cKit+/CD45‐ cells two‐fold but did not alter the percentage of cKit+GATA4+ or cKit+cTnI+ myocyte precursors. In contrast, pravastatin stimulated CSC proliferation with a nine‐fold increase in cKit+/Ki67+ and a three‐fold increase in the expression of Jagged1.

Conclusions

These data indicate that pravastatin up‐regulates Jagged1 in the heart and stimulates CPC proliferation but not the differentiation of CPCs into cardiac myocytes.

Supported by NYSTEM and HL55324
Use of human platelet lysates in stem cell‐based alternative testing strategies
Authors: Gstraunthaler Gerhard , Rauch Caroline , Feifel Elisabeth
Published: June 2021

The supplementation of culture media with fetal bovine serum (FBS) is common practice and essential for cell growth and proliferation in vitro. The major function of serum is to provide hormones, cytokines, growth factors, etc. However, the use of FBS in cell culture bears a number of disadvantages: high lot‐to‐lot variations; presence of adverse factors; suffering of bovine fetuses during blood harvest. The most potent mitogenic factors present in serum are derived from activated thrombocytes. Recently, we proved the capacity of human platelet lysates (PL) to replace FBS in human and animal cell culture systems. Stem cells may become the alternative to animal testing and in vitro toxicology, and their performance under animal‐derived component free culture conditions has to be defined in validation studies. In order to accomplish these tasks, human bone marrow and adipose‐derived mesenchymal stem cells were expanded in media supplemented with PL. Proliferation assays confirmed the growth promoting effect of PL, comparable to high FBS. Furthermore, the cells kept their undifferentiated state as proofed by CD44 and CD105 expression. Next, it will be tested if the cells can be differentiated towards osteogenic, chondrogenic and adipogenic phenotypes under these culture conditions.
Microglial Kv1.3 Channels as a Potential Target for Alzheimer's Disease
Authors: Jenkins David Paul , Maezawa Izumi , Wulff Heike , Jin Lee-Way
Published: June 2021

Microglia, the major inflammatory cells of the brain, play a pivotal role in the initiation and progression of Alzheimer's Disease (AD) by either phagozytosing amyloid‐β deposits or by releasing cytotoxic and pro‐inflammatory substances in response to activation by amyloid‐β aggregates, including amyloid‐β oligomers (AβO). We here propose microglial Kv1.3 channels as a novel target for curbing the harmful effects of Aβ‐induced microglia activation. Microglia isolated from the brains of adult 5xFAD mice expressed higher levels of Kv1.3 than microglia from age‐matched control mice. We further observed strong Kv1.3 immunoreactivities in microglia associated with amyloid plaques in brains of 5xFAD mice. Proof for the functional importance of Kv1.3 in microglia comes from our observations that the Kv1.3 blocker PAP‐1 inhibits AβO‐stimulated NO production as well as microglia‐mediated neurotoxicity in dissociated cultures and organotypic brain slices. A 6‐week course of daily PAP‐1 injections also reduced the degree of microglia activation in 5xFAD mice. In contrast, Kv1.3 blockade with PAP‐1 does not affect phagocytosis of Aβ aggregates by microglia. These observations suggest that Kv1.3 blockers might preferentially inhibit microglia mediated neuronal killing without affecting beneficial functions such as scavenging of debris.

Supported by NIH and Alzheimer's Association
Direct interaction of Bestrophin 3 channels with Hrc calcium‐handling protein
Authors: O'Driscoll Kate E , Hatton William J , John Steeven , Baring Martha , Britton Fiona C
Published: June 2021

Bestrophin 3 (Best3) is a calcium‐activated chloride channel expressed in heart and other tissues. We hypothesized that Best3 channels function in a macromolecule complex similar to other cardiac ion channels.

Yeast two‐hybrid assays performed on a mouse heart library, identified histidine‐rich calcium binding protein (Hrc) as a novel binding partner of Best3. Hrc is a regulator of sarcoplasmic reticulum (SR) calcium uptake, storage and release. This interaction was further determined to involve Best3 amino acids 541–669 in the cytosolic C‐terminus of the channel and the cysteine‐rich region in mouse Hrc (amino acids 665–738 aa). Specific binding between Best3 channels and Hrc was verified by co‐immunoprecipitation using mouse heart homogenates. Confocal immunofluorescence microscopy further demonstrated a strong co‐localization of Best3 and Hrc in isolated ventricular and atrial myocytes.

In this study we report that cardiac Best3 channels interact with Hrc. Calcium‐activated chloride channels play a critical role in excitation‐contraction coupling in the SR by balancing charge movement during calcium release and reuptake. The direct coupling of Best3 with Hrc suggests that Best3 channels may play a role in calcium handling in the heart by functioning as a counterion channel.
The gasotransmitter hydrogen sulphide decreases Na+ transport across lung epithelial cells
Authors: Althaus Mike , Urness Kevin Douglas , Clauss Wolfgang , Baines Deborah L , Fronius Martin
Published: June 2021

The transepithelial absorption of Na+ in the lung is crucial for the maintenance of the volume and composition of epithelial lining fluid. The regulation of Na+ transport is essential, since hypo‐ or hyperabsorption of Na+ is associated with lung diseases such as pulmonary oedema or cystic fibrosis. This study investigated the effects of the gaseous signalling molecule hydrogen sulphide (H2S) on Na+ absorption across human H441 pulmonary epithelial cells and native preparations of mouse and pig trachea. H2S rapidly and reversibly inhibited Na+ transport by all models when electrophysiologically measured in Ussing chambers. There was no effect on Na+ channels, whereas Na+/K+‐ATPase currents were decreased. Membrane abundance of the Na+/K+‐ATPase, as determined by surface biotinylation and western blot, or cellular ATP concentrations were not affected. H2S inhibited basolateral K+ channels, the inhibition of which decreased Na+/K+‐ATPase currents and Na+ absorption by H441 monolayers.

Thus, H2S impairs pulmonary transepithelial Na+ absorption, due to inhibition of both Na+/K+‐ATPase activity and basolateral K+ channels. These data suggest that the H2S signaling system might represent a novel pharmacological target for interference with pulmonary transepithelial Na+ transport. This work was funded by the German Research Foundation (AL 1453/1‐1) and the German Academic Exchange Service.
CENTRAL PROCESSING BY ELECTROACUPUNCTURE OF CARDIOVASCULAR REFLEX VASODEPRESSION
Authors: Tjen-A-Looi Stephanie CheeYee , Li Peng , Hsiao An-Fu , Longhurst John C
Published: June 2021

Electroacupuncture (EA) at P5–6 reduces sympathoexcitatory blood pressure (BP) reflex responses. Gamma‐aminobutyric acid (GABA) receptors in the rostral ventral lateral medulla (rVLM) contribute to modulation of sympathoexcitatory visceral reflexes during EA. Gastric distension (GD) in hypercapnic acidotic (HA) rats, by activating both sympathetic and vagal afferents, decreases BP and heart rate (HR) through a GABAA mechanism in the rVLM. This study investigated the hypothesis that EA modulates GD‐induced hemodynamic depressor responses through actions in nuclei that process both sympathetic and parasympathetic outflow. Anesthetized and HA‐induced rats showed repeatable decreases in BP and HR with distension of the stomcah every 10 min. Bilateral EA at P5–6 for 30 min reversed the hypotensive response from − 26±3 to −6±1 mmHg and the bradycardia from −35±11 to −10±3 beats/min for over 70 min. EA's action on decreased BP and HR, respectively, was inhibited by microinjection of gabazine into the caudal–VLM (cVLM) or the nucleus ambiguus (NAmb). Gabazine microinjected into the rVLM reversed EA action on both depressor and bradycardia responses. Thus, EA through GABAA receptor mechanisms modulates reflex sympathoinhibition and vagal excitation leading to cardiovascular depression through actions in the rVLM, cVLM and NAmb. These data indicate that EA can normalize elevated and depressed blood pressure.
Rats with ischemic heart failure (IHF) exhibit exaggerated conditioned fear and altered β‐adrenergic receptor expression in forebrain regions that govern autonomic responses to fear
Authors: Glasgow Jaimee , Cunha Jonathan , Tiniakov Ruslan , Samarel Allen , Koshman Yevgeniya , Neafsey Edward J , Battaglia George , Scrogin Karie
Published: June 2021

Up to 26% of patients with IHF develop anxiety disorders. β‐adrenergic receptors (βARs) in the basolateral amygdala (BLA) mediate fear expression, while βARs in the infralimbic region of the medial prefrontal cortex (IL) contribute to extinction of learned fear. We found that stimulation of the locus ceoruleus (the main source of adrenergic input to the forebrain) reduced metabolic activity in the IL and increased arrhythmias in IHF rats, but not in controls. Here, we determined if rats with IHF exhibit increased conditioned fear, deficits in fear extinction and altered βAR receptor density in the forebrain. Rats trained to associate foot shock (0.5 mA, 0.5 s) with a 10 s tone were subjected to coronary artery ligation or sham surgery. Rats with IHF (fractional shortening < 25%) showed increased freezing behavior during the first 3 extinction tones (3.2±0.6 vs 6.2±0.5 s, P<0.05), and delayed fear extinction (4.1±0.7 vs 10.5±1.3 trials, P<0.01) compared to controls 7 weeks after surgery. β1AR density was reduced in the IL (15%) and rostral BLA (29%) of IHF rats (P<0.05). Inhibition of βAR antagonist binding by a βAR agonist was increased two‐fold in the BLA of IHF rats (P<0.05), suggesting a greater density of G protein‐coupled βAR receptors. These data implicate altered forebrain adrenergic neurotransmission in the anxiety and increased arrhythmias that develop with IHF. Supported by AHA 09PRE2260787.
Progression of heart failure after myocardial infarction in the conscious rat
Authors: Barrett Carolyn J , Pyner Susan , Malpas Simon C , Guild Sarah-Jane
Published: June 2021

The rat model of heart failure after myocardial infarction (MI), induced by coronary ligation, is commonly utilized in the study of heart failure. Typically animals are studied at one time point, with the progression of the changes in both ventricular function and regulation of the sympathetic system poorly understood. The aim of this study was to characterize the time course of the changes in left ventricular function and sympathetic activation following MI in the rat. In separate groups of animals left ventricular pressure and renal sympathetic nerve activity (RSNA) were measured continuously via telemetry both before and following MI or sham surgery. Despite echocardiography showing fractional shortening had halved, left ventricular pressures were initially well conserved, with the most obvious change 3 weeks post MI being a decrease in the maximum rate of pressure change (dP/dt max) at night. At the same time period RSNA had increased by more than 100% and neuronal excitation was observed within the PVN, as determined by Fra‐protein expression. In contrast similar levels of nNOS positive cells were observed in PVN of the sham and MI rats. These experiments suggest sympathetic activation occurs early in the progression of heart failure, independent of changes in nNOS regulation in the PVN. Funded by the Health Research Council of New Zealand and British Heart Foundation.
Effects of thyroid hormone supplementation on ventilation and metabolism in a rat model of Type 2 Diabetes
Authors: Schlenker Evelyn Heymann , Bollinger Stephen , Weltman Nathan , Gerdes A. Martin
Published: June 2021

Thyroid hormone supplementation can have beneficial effects in type 2 diabetes (T2D). In this study T2D was induced in thirty young female rats by an intraperitoneal (ip) injection of nicotinamide (200 mg/kg) followed 15 minutes later by an ip injection of streptozotocin (65 mg/kg, STZ/N). Control rats (CR, n=10) received vehicle injections. After 2 months, subgroups of STZ/N rats received either 4 (STZ/N4) or 8ug/kg/day (STZ/N8) of triiodothyronine (T3) in drinking water for one month. Subsequent metabolic and ventilatory studies (during exposure to air or hypoxia, 10% O2 in N2) indicated little effects in the STZ/N group compared to CR. In contrast, STZ/N8 group was clearly hyperthyroid with lower body weights, elevated body temperatures, metabolic rates, and ventilation during exposure to air and hypoxia. Except for changes in inspiratory (increased) and expiratory (decreased) times during air exposure, the ventilation and metabolism of the STZ/N4 group was similar to the CR and STZ/N groups. Importantly, the lower, but not higher dose of T3 attenuated the hyperglycemia observed in both the STZ/N and STZ/N8 groups. Thus, low levels of T3 supplementation may have beneficial effects in T2D, but have subtle effects of ventilatory timing parameters during air exposure. Funded by R01 7R01HL093160‐03 (from NHLBI) and a Clinical Scientist Training Award from the American Diabetes Association (ADA 7‐10‐CST‐01).
Spinal VEGF induced phrenic motor facilitation is unaffected by pretreatment with repetitive acute intermittent hypoxia
Authors: Dale Erica Arden , Satriotomo Irawan , Mitchell Gordon S
Published: June 2021

Vascular endothelial growth factor (VEGF) and VEGF receptor‐2 (VEGFR‐2) are expressed in identified phrenic motor neurons, and cervical spinal VEGF receptor activation elicits long‐lasting phrenic motor facilitation (pMF), demonstrating the potential of VEGFA to elicit respiratory plasticity. Since VEGF and VEGFR‐2 are regulated by hypoxia, we tested the hypotheses that: 1) VEGF and VEGFR‐2 are upregulated in phrenic motor neurons after repetitive exposure to acute intermittent hypoxia (3× per wk, 10 wks; 3xwAIH); and 2) 3xwAIH (4 wks) enhances VEGF‐induced pMF due to VEGF receptor up‐regulation. 3xwAIH increases VEGF (2.9±0.2 vs. 1.0±0.3; p<0.05) and VEGFR‐2 expression vs normoxic controls (8.7±0.07 vs. 1.9±0.5; p<0.05). Cervical spinal VEGFA administration elicited similar pMF in normoxic and 3xwAIH treated rats. Thus, although repetitive intermittent hypoxia up‐regulates VEGF and VEGR‐R expression in phrenic motor neurons, there was no evidence for any functional effect. Further exploration of this issue is warranted since: 1) there may be a “ceiling effect” as VEGF‐induced pMF may elicit maximal phrenic output, or 2) 4 weeks of 3xwAIH may not be sufficient time to up‐regulate the relevant proteins.

Supported by: NIH NS05777 & HL080209.
Spinal respiratory plasticity is impaired by two models of systemic inflammation, lipopolysaccharide and severe intermittent hypoxia
Authors: Huxtable Adrianne G , Peterson Timothy , Smith Stephanie M , Watters Jyoti J , Mitchell Gordon S
Published: June 2021

Although inflammation is associated with many clinical disorders that challenge ventilatory control, we know little about the influence of inflammation on mechanisms of ventilatory control. Here, we show that two distinct models of systemic inflammation (lipopolysaccharide, LPS) and severe intermittent hypoxia (sIH: 2 min hypoxia, 2 min normoxia, 8–12 hrs) impair phrenic long‐term facilitation (pLTF), a model of spinal respiratory plasticity induced by acute intermittent hypoxia (AIH; 3X5min hypoxia; 5 min interval). Both LPS (100 μg/kg, i.p.) and sIH impaired pLTF (LPS: 4±4%, n=9; sIH: 0.3±1.3%, n=2; control: 58±17%, n=5). The NSAID ketoprofen (12.5 mg/kg, i.p.) restored pLTF following both LPS and sIH (LPS: 55.1±12.3%, n = 6; sIH: 45%; n=1). In our working model, multiple inflammatory molecules converge on and inhibit pLTF by activating p38 MAP kinase. Preliminary data suggest that spinal p38 MAP kinase inhibtion restores pLTF (31%) after LPS. Additionally, immunohistochemistry suggests increased cervical spinal p38 phsophorylation after LPS. Understanding the impact of inflammation on respiratory plasticity is fundamental since it may diminish natural, compensatory responses during pathological states. Supported by HL80209, HL69064, T32 HL007654, NS 049033.
Breathlessness descriptors during constant load cycling in obese men with and without dyspnea on exertion
Authors: Bernhardt Vipa , Haller Sarah F , Moran Raksa B , Babb Tony G
Published: June 2021

During constant load exercise, some otherwise healthy obese men experience dyspnea on exertion (+DOE), while others do not (−DOE). Their subjective respiratory sensations may explain why the intensity of breathlessness is greater in +DOE.

18 obese men participated and were divided based on their Ratings of Perceived Breathlessness (RPB, Borg 0–10 scale) during minute 6 of constant load (90 W) cycle exercise; ten +DOE (RPB ≥ 4) (36 yr, 39 BMI, 3.1 L/min peak V̇ O2, 104% predV̇O2) and eight −DOE (RPB ≤ 2) (34 yr, 35 BMI, 3.0 L/min peak V̇O2, 104% predV̇ O2). After the exercise, subjects filled out a questionnaire asking them to pick three of fifteen descriptors that best described the respiratory sensations they felt. Data were analyzed using z‐test proportion comparisons and clusters.

Both +DOE and −DOE groups chose descriptors with similar frequency, only “I feel that I am breathing more” (+DOE 80%, −DOE 50%) and “I feel that my breathing is rapid” (+DOE 20%, −DOE 50%) differed slightly but not significantly between the groups (p=0.37). The top four clusters were ‘Breathing more’ (67% of all participants), ‘Heavy’ (44%), ‘Work/Effort’ (44%), and ‘Rapid” (33%).

We conclude that obese men with and without DOE describe their respiratory sensations similarly during submaximal exercise, although their intensity of breathlessness is different.

Supported by HL096782, King Charitable Foundation Trust, ALA, AHA.
Upregulation of Orexin Receptor 1 Contributes to Increased Sympathetic Output in Obese Zucker Rats
Authors: Li De-Pei , Pan Hui-Lin
Published: June 2021

Increased sympathetic outflow is involved in the pathogenesis of metabolic syndrome. Orexins are important in the control of autonomic function and energy metabolism. We determined the role of orexins in the control of the paraventricular (PVN) presympathetic neurons in Obese Zucker Rats (OZRs) and Lean Zucker Rats (LZRs). The PVN‐spinal cord output neurons were retrogradely labeled by fluorescent microspheres injected into the rat thoracic spinal cord. The basal firing rate in the labeled PVN neurons was higher in OZRs than in LZRs. Orexin A produced a greater dose‐dependent increase in the firing rate in PVN neurons in OZRs than in LZRs. Orexin A caused a greater increase in NMDA currents induced by puff application of NMDA in OZRs than in LZRs. Orexin A increased the frequency of spontaneous excitatory postsynaptic currents in labeled PVN neurons. The orexin receptor 1 (OX1R) expression level in the PVN was higher in OZRs than in LZRs. OX1R immunoreactivity was colocalized with microsphere‐labeled PVN‐spinal neurons. In addition, microinjection of orexin A into the PVN produced greater increases in arterial blood pressure (ABP) and lumbar sympathetic nerve activity (LSNA) in anesthetized OZRs than in LZRs. Collectively, these data suggest that upregulated OX1R in the PVN contributes to increased sympathetic outflow and hyperactivity of PVN presympathetic neurons through enhancing glutamatergic transmission.
ABC Transporter Expression, Function and Regulation at the Blood‐Spinal Cord Barrier
Authors: Miller David S. , Schroeter Christian , Wang Xueqian , Campos Christopher R.
Published: June 2021

The blood‐spinal cord barrier (BSCB), which resides within spinal cord capillaries, is a major impediment to the delivery of drugs used to treat, e.g., traumatic spinal cord injury and amyotrophic lateral sclerosis. These capillaries appear structurally similar to brain capillaries, the site of the blood‐brain barrier (BBB), but we have little information about the molecular basis of BSCB function. Here we demonstrate protein expression (western blots) and transport function (confocal‐based assay) for three ATP‐driven, drug efflux pumps: P‐glycoprotein, multidrug resistance protein 2 (Mrp2) and breast cancer related protein (Bcrp) in capillaries isolated from rat and mouse spinal cords. We show upregulation of transporter function and expression in capillaries exposed to pregnenolone‐16α‐carbonitrile (activates pregnane X receptor), phenobarbital (activates constitutive androstane receptor) and dioxin (activates aryl hydrocarbon receptor). We also show increased P‐glycoprotein expression and transport function in brain capillaries from rats dosed with 5μg/kg TCDD. These results indicate that the BBB and BSCB are remarkably similar with regard to efflux transporter function and regulation.