bluuebunny/crossref_metadata_2025 huggingface dataset:
Induced Pulp Lesion Promotes Alteration of the Insulin Signal and Causes Insulin Resistance
Authors: Sumida Doris Hissako , Astolphi Rafael Dias , Curbete Mariane Machado , Chiba Fernando Yamamoto , de Melo Yamamoto Aline Pedro , Prieto Annelise Katrine Carrara , Cintra Luciano Tavares Angelo
Published: June 2021

The current study aimed to investigate if a pulp lesion, which is a local inflammatory disease, can promote alterations in insulin signal and insulin sensitivity (IS). Twenty‐eight two‐month‐old (230g) Wistar male rats were randomly divided into a control group (CN) and a pulp lesion group (PL). PL group was submitted to pulp lesion induction through the opening of the coronal pulp chamber of the upper right first molars by using a dental handpiece, followed by its exposure to oral cavity, promoting bacterial contamination and chronic inflammation. Thirty days after PL induction, the following evaluations were performed: 1) pp185 (IRS‐1/IRS‐2) tyrosine phosphorylation status (pp185 P‐Tyr), after insulin stimulation, in periepididimal white adipose tissue (WAT) and gastrocnemius muscle (G) by Western Blotting analysis; 2) IS determination by intravenous insulin tolerance test. The results showed that PL promoted: 1) a significant decrease (p<0.05) in the pp185 P‐Tyr in WAT (CN=152.9±12.7 vs PL=111.3±10.2 arbitrary units/ìg of protein, n=7) and in G (CN=133.1±9.1 vs PL=103.7±7.6 arbitrary units/ìg of protein, n=7); 2) a significant difference (p<0.05) in the IS (glucose disappearance speed rate: CN=3.04±0.26 vs PL=1.97±0.25 %min, n=7). These data suggest that PL is capable of changing insulin signal in both WAT and G, and causing insulin resistance in rats. Financial Support: FAPESP (2011/04255–8).
The role of nitric oxide in pericyte‐mediated regulation of vasa recta diameter
Authors: Crawford Carol , Pollock Jennifer S , Wildman Scott S P , Peppiatt‐Wildman Claire M
Published: June 2021

Pericytes are known to regulate vasa recta diameter in response to many different vasoactive agents endogenous to the kidney [1]. Nitric oxide (NO) signaling pathways are important in the renal medulla for regulation of blood flow and natriuresis. Here we use the live kidney slice model to characterize the mechanisms by which NO regulates pericyte‐mediated control of vasa recta diameter and thus medullary blood flow (MBF).

Kidney slices (200 μm thick) were obtained from adult, male rats as previously described [1]. Kidney slices were superfused with agents that manipulate endogenous NO signaling pathways: L‐NAME [an inhibitor of nitric oxide synthase (NOS)], or L‐VNIO (a NOS1‐specific inhibitor), or SNAP (a NO donor). DIC video imaging techniques were used to record real‐time changes in vasa recta diameter, in response to bath application of these compounds. SNAP (100 μM) caused a significant dilation of vasa recta specifically at pericyte sites (12.6±2.9% p<0.05 n=15). L‐NAME (1 mM) and L‐VNIO (1 μM) each caused vasoconstriction significantly greater at pericyte sites compared with non‐pericyte sites (8.6±1.1% p<0.05 n=13 and 10.8±2.0% p<0.05 n=6, respectively).

Here we demonstrate that NO derived from NOS1 specifically is key in pericyte‐mediated regulation of vasa recta diameter, and thus renal MBF.

Research supported by the Medical Research Council
Inhibition of NADPH oxidase improves renal autoregulation in rats fed a high salt diet
Authors: Fellner Robert C. , Giddens Matthew I. , Inscho Edward W.
Published: June 2021

Previous work shows that a high salt diet (HS) blunts afferent arteriolar (AA) autoregulatory behavior by 83% partly through reactive oxygen species (ROS). Thus, we tested the hypothesis that HS blunts autoregulation by enhancing NADPH‐dependent ROS production. Rats were fed a normal (NS 0.8% NaCl) or HS (8% NaCl) diet for 14 days. A subset of rats, received the NADPH oxidase inhibitor, apocynin (~25mg/kg/day) in the drinking water. Studies were performed in vivo to assess whole kidney autoregulation of renal blood flow (RBF) and glomerular filtration rate (GFR) when left kidney perfusion pressure (PP) was reduced to ~95mmHg using an adjustable aortic clamp. Decreasing PP significantly reduced RBF in HS rats by 18%, from a baseline of 12.6 ±0.6 to 10.4±0.6 ml/min (n=8) compared to 2% in NS rats (8.3±0.8 to 8.1±0.9 ml/min; n=10; P<.05 vs. HS). Apocynin treatment improved autoregulatory behavior in HS rats with RBF declining by just 8%, from a baseline of 12.5 ±0.5 to 11.6 ±0.5 ml/min (n=11). Similarly, left kidney GFR declined by 27% (457 ± 24 vs. 335 ± 31 μl/min/100gbw; n=8) in HS rats compared to just 5% (496 ±46 vs. 470± 39 μl/min/100gbw; n=10) in NS rats. Apocynin improved GFR autoregulation in HS rats with only a 12% reduction at 95 mmHg (472±39 vs. 417±21 μl/min/100gbw; n=11). These data suggest that NADPH oxidase‐derived ROS impair renal autoregulatory behavior in rats fed a HS diet.

Funding: NIH‐HL098135, NIH‐1PO1HL095499
The α2‐Adrenoceptor Agonist, Dexmedetomidine, Elicits Pronounced Decreases in Arterial Blood Pressure and Renal Hemodynamics in Cyp1a1‐Ren2 Transgenic Rats with Angiotensin II‐Dependent Malignant Hypertension
Authors: Thorngren Christina K. , Patterson Matthew E. , Davis Porcha D. , Mitchell Kenneth D.
Published: June 2021

Previous studies have demonstrated that increased sympathetic nervous system activity contributes to the pathogenesis of various forms of hypertension. However, the contribution of increased sympathetic nervous system activity to the elevated arterial blood pressure in angiotensin (ANG) II‐dependent malignant hypertension remains unclear. The present study was performed to determine the effects of acute administration of the highly selective α2‐adrenoceptor agonist, dexmedetomidine, on mean arterial blood pressure (MAP) and renal hemodynamics in Cyp1a1‐Ren2 transgenic rats with ANG II‐dependent malignant hypertension. Male Cyp1a1‐Ren2 rats (n=5) were induced to develop malignant hypertension by dietary administration of indole‐3‐carbinol (I3C, 0.3% wt/wt) for 10 days. Mean arterial pressure (MAP) and renal hemodynamics were measured in pentobarbital‐anesthetized male Cyp1a1‐Ren2 rats during control conditions and during continuous infusion of dexmedetomidine (5 μg/kg/min, iv). Dexmedetomidine administration decreased MAP in the hypertensive rats (172±6 to 89±6 mmHg, P<0.001). The dexmedetomidine‐induced decrease in MAP was associated with marked reductions in renal plasma flow (2.22±0.37 to 0.80±0.34 ml/min.g, P<0.05) and glomerular filtration rate (0.70±0.07 to 0.23±0.09 ml/min.g, P<0.01). The present findings demonstrate that acute administration of an α2‐adrenoreceptor agonist elicits pronounced decreases in MAP and renal hemodynamic function in Cyp1a1‐Ren2 transgenic rats with ANG II‐dependent malignant hypertension. Thus, the data indicate that increased sympathetic nervous system activity plays an important role in maintaining the elevated MAP that occurs in ANG II‐dependent malignant hypertension.
Rapamycin, an inhibitor of mTOR signaling pathway, reverses lithium‐induced cell proliferation in renal collecting ducts
Authors: Gao Yang , Romero‐Aleshire Jill , Cai Qi , Price Ted J. , Brooks Heddwen L.
Published: June 2021

The most common side effect in patients undergoing lithium therapy is nephrogenic diabetes insipidus (NDI), in addition, lithium treatment can cause collecting duct cells to proliferate. The mammalian target of rapamycin (mTOR) signaling pathway is a key regulator of cell proliferation. We hypothesized that the mTOR signaling pathway may be playing a role in lithium‐induced cell proliferation of renal collecting duct. We fed mice lithium for 14d; AQP2 protein expression was significantly decreased (16 ± 4.0% vs control 100 ± 8.8%) and proliferating cell nuclear antigen (PCNA) protein expression was significantly increased (172 ± 8.6% vs control 100 ± 1.4%) in renal inner medulla (IM). We demonstrate that p‐mTOR (Ser 2448) was increased (154 ± 26.5%), as was phosphorylation of ribosomal S6 protein (p‐rS6), a downstream component of mTOR pathway (404 ± 151.4%) in renal IM of lithium‐treated mice. To test whether the inhibition of mTOR signaling pathway could reverse lithium‐induced cell proliferation, we treated mice with Rapamycin (Rapa), an inhibitor of mTOR. Rapa reversed lithium‐induced proliferation of IM collecting duct cells and decreased lithium‐induced p‐mTOR and p‐rS6 levels. Rapa had no effect on the upstream components of mTOR; p‐Akt and p‐TSC2 remained elevated by lithium. In conclusion, the mTOR signaling pathway is involved in lithium‐induced collecting duct cell proliferation.
Gene expression in the thin limbs of Henle of the waterrestricted rat
Authors: Nawata C. Michele , Dantzler William H. , Pannabecker Thomas L.
Published: June 2021

Sodium and water homeostasis and the formation of concentrated urine depend upon the generation of a steep osmotic gradient within the renal medulla. Although the mechanism of this process remains controversial, the descending thin limbs (DTLs) and ascending thin limbs (ATLs) of the loops of Henle play important roles. Our goal was to identify genes that may be involved in the urine concentrating mechanism and to quantify their transcript levels in the DTLs and ATLs. Using real‐time PCR, we analyzed transcripts in isolated DTLs and ATLs of water‐restricted rats. Claudin 4 (CLDN4), sodium‐dependent glucose transporter 1 (NaGLT1), sodium glucose cotransporter 1 (SGLT1), and sodium myo‐inositol cotransporter 1 (SMIT1) were present in both types of tubules and their levels increased in the ATLs after 72h of water restriction. Chloride channel (Clcnka) mRNA in the ATLs and urea transporter mRNA (UT‐A2) in the DTLs increased. We also identified a novel variant of SGLT1 and four variants of the arginine vasopressin receptor 2 (AVPR2). Our results indicate that in addition to the previously characterized aquaporins and urea transporters, several other genes and their products may be involved in the movement of ions and solutes in the thin limbs of Henle. Further studies will determine how these genes contribute to urine concentration. NIDDK DK083338
Protein carbamylation is a physiological post‐translational modification in the renal inner medulla
Authors: Claxton J. S. , Liu G. , Sandoval P. C. , Chou C‐L , Knepper M. A.
Published: June 2021

We hypothesize that the high urea concentration in the renal inner medulla (IM) is associated with physiological carbamylation of proteins. Using a carbamylation‐specific antibody, immunoblotting revealed multiple bands in the kidney of antidiuretic rats with a gradient of carbamylation from cortex to IM, suggestive of large‐scale protein carbamylation. To test the specificity of the antibody, we incubated BSA with 1 M urea in PBS or PBS alone for 4 days. The antibody recognized a band consistent with carbamylated BSA in the presence but not the absence of urea. To confirm widespread protein carbamylation in rat IM, we carried out protein mass spectrometry. Proteins, processed in the absence of urea, were separated via SDS‐PAGE. Successive gel blocks were subjected to in‐gel trypsin digestion. Using LC‐MS/MS (Orbitrap) and a high‐stringency spectral search (SEQUEST), we identified 686 carbamylation sites in 579 proteins. Many of the identified sites were present on lysine side chains. The results show that extensive protein carbamylation occurs in rat renal IM, which could have an impact on cell signaling by blocking post‐translational modifications of lysines, e.g. acetylation and ubiquitylation.
Sex does not impact asymmetric dimethylarginine (ADMA) or L‐arginine (L‐arg) levels in spontaneously hypertensive rats (SHR).
Authors: Brinson Krystal N. , Sharma Shruti , Romero Maritza J. , Sullivan Jennifer C.
Published: June 2021

ADMA and L‐arg are endogenous amino acids that impact nitric oxide synthase (NOS) activity. ADMA has been implicated in nitric oxide (NO) deficiency and hypertension. We have reported that female SHR have significantly greater NO bioavailability and NOS enzymatic activity in the renal inner medulla compared to age‐matched males, yet the mechanism responsible is unknown. We hypothesized that male SHR have greater ADMA levels and less L‐arg compared to females. ADMA and L‐arg levels in plasma (N=6–7) and renal inner medullas (N=10) were measured via HPLC from 12–13 week old male and female SHR. ADMA (Male: 20±6; Female: 31±8 nmols/ml, p=0.7), L‐arg (Male: 152±7; Female: 141±11 nmols/ml, p=0.7) levels and the L‐arg/ADMA ratio (p=0.2) in plasma were comparable between sexes. Renal inner medullary ADMA (Male: 172±55 Female: 192±58 grams wet weight (gww), p=0.8), L‐arg (Male: 507±107; Female: 552±127 gww, p=0.8) levels and L‐arg/ADMA ratio (p=0.8) were also comparable between groups. We conclude that while ADMA and L‐arg may be important in maintaining renal NOS activity in SHR, they do not contribute to sex differences in renal inner medullary NOS activity. Funded by the American Heart Association.
Role of the Angiotensin AT2 receptor in obesity‐linked inflammation and renal injury: Effect of gender
Authors: Dhande Isha , Khan Mohammed Azhar , Nag Sourashish , Hussain Tahir
Published: June 2021

The Angiotensin AT2 receptor (AT2R) is up‐regulated in response to tissue injury and is implicated in tissue protection and repair. We have recently demonstrated that the AT2R signaling may be differentially regulated in males and females in the setting of obesity. The present study was designed to investigate the effect of gender on the protective role of the AT2R in mediating obesity‐linked renal inflammation and injury. Young, 3–4 week old male and female AT2R‐deficient (AT2KO) mice were maintained on a normal (ND) or high‐fat diet (HFD) for 16 weeks. Female, but not male, AT2KO ND mice had 5‐fold higher levels of plasma and renal tumor necrosis factor‐α (TNF‐α). HFD significantly increased (Male: 4‐fold and Female: 5‐fold higher compared to ND controls) plasma and renal levels of TNF‐α. AT2KO males and females on HFD had 1.5‐fold higher TNF‐α compared to HFD controls. Renal histology revealed increased mesangial matrix expansion (MME) in all the HFD groups (both, control and AT2KO), while the glomeruli of the ND control and AT2KO male mice were normal. On the other hand, female AT2KO mice even on ND had greater extent of MME compared to controls (1.1±0.2 vs 0.3±0.1) and HFD further increased MME (2.9±0.1 vs 1.3±0.2). Our results suggest that AT2R is reno‐protective in obesity‐linked renal inflammation and injury. Further, the protective role of the AT2R appears to be more critical in females. Supported by NIH R01 DK‐61578.
Renal Intramedullary Infusion of Tempol Normalizes the Blood Pressure Response to Intrarenal Blockade of Heme Oxygenase‐1 in Angiotensin II‐Dependent Hypertension
Authors: Stec David , Gousset Monette
Published: June 2021

Previous studies have demonstrated that intramedullary inhibition of heme oxygenase‐1 (HO‐1) increases the blood pressure response to angiotensin II (Ang II) infusion. The increase in blood pressure was associated with an increase in renal medullary superoxide production. The current study was designed to test the hypothesis that increased renal medullary superoxode production contributes to the increase in blood pressure in response to blockade of renal medullary HO‐1 in Ang II hypertension. Male C57BL/6J mice (16–20 weeks of age) were implanted with chronic intrarenal medullary interstitial (IRMI) and infused with: saline, Tempol (T; 6mM), the HO‐1 inhibitor QC‐13 (25 μM), or a combination of T + QC‐13. T treatment was started 2 days before infusion of QC‐13 and IRMI infusions were switched 4 days prior to implantation of Ang II containing osmotic minipumps which infused Ang II at a rate of 1 μg/kg/min. Blood pressures on days 7–10 post Ang II infusion averaged 150 ± 3 mmHg in mice receiving IRMI infusion of saline. IRMI infusion of T alone decreased blood pressure to 136 ± 3 mmHg. IRMI infusion of QC‐13 alone increased blood pressure to 164 ± 2 (p<0.05) and IRMI infusion of T along with QC‐13 significantly attenuated the rise in blood pressure to 142 ± 2 mmHg (p<0.05). These results demonstrated that blockade of intrarenal medullary superoxide production with T attenuates the increase in blood pressure in response to intrarenal medullary inhibition of HO‐1. These results support the important role for the antioxidant actions of HO‐1 metabolites in the renal medulla in the regulation of blood pressure.
Enhanced urinary sodium and potassium excretion in VAMP3 knockout mice
Authors: Haque Mohammed Ziaul , Ortiz Pablo Alfredo
Published: June 2021

Several members of the vesicle associated membrane proteins (VAMP) family are expressed in the kidney. VAMP3 is expressed in the apical membrane of the thick ascending limb (TAL) where it mediates NKCC2 trafficking to the apical membrane. Other transporters along the nephron segment may also be regulated by VAMP3 thereby influencing renal function. We hypothesized that VAMP3 is involved in maintaining renal Na excretion and thus blood pressure. To test our hypothesis we obtained homozygous VAMP3 knockout mice (KO) and measured systolic blood pressure (SBP) and urinary parameters using c57BL6 mice (WT) as controls. Urinary Na excretion was higher (325±25 vs 248±20 μmoles/day) as was K excretion (785±76 vs 645±27 μmoles/day) and urine volume (2.56±0.16 vs 1.95±0.19 ml/day) without a change in urinary creatinine. Body weight tended to be higher but not significantly so (27.2±0.6 vs 26.3 ±0.2 gram). After decreasing Na in the diet from 0.4 to 0.02% for 24 hours, Na excretion decreased in both strains. However, it remained elevated in KO (61±14 vs 38±3 μmoles/day). Baseline SBP was lower in KO mice compared to WT (KO: 101±3 vs WT: 115±3 mmHg). Feeding mice a low salt diet for 7 days did not further decrease SBP in VAMP3 KO (96±4 mmHg) but it decreased in WT (99±3 mmHg). Taken together our data suggest that VAMP3 maintains normal renal Na excretion and is involved in blood pressure homeostasis. This is the first phenotype observed in VAMP3 KO mice.
Hypertonic NaCl intake induces renal TNF production by a pathway involving NKCC2A and NFAT5
Authors: Hao shoujin , Bellner Lars , Ferreri Nicholas R.
Published: June 2021

TNF is produced by the medullary thick ascending limb (mTAL), however, the effect of hypertonicity on production of this cytokine has not been determined. Mice given 1% NaCl in the drinking water for 3 days exhibited a two‐fold increase in urinary, but not plasma, TNF levels compared with mice given tap water. A concomitant 3‐fold increase in NKCC2A and 2‐fold increase in NFAT5 mRNA accumulation was observed in mTAL tubules from mice given 1% NaCl. Increases in urinary TNF levels were abolished after treatment with furosemide or intrarenal injection of a lentivirus shRNA construct that knocked down NKCC2A (EGFP‐N2A‐ex4); plasma levels of TNF did not change. NFAT5 mRNA abundance also decreased after injection of EGFP‐N2A‐ex4. TNF production by primary cultures of mTAL cells increased approximately 6‐fold in response to high NaCl concentration (400 mosmol/kg H2O) and was inhibited in cells treated with bumetanide or transiently transfected with a dnNFAT5 construct (3.0 ± 0.9 to 0.9±0.2 pg/μg protein; p<0.05). Transduction of mTAL cells with EGFP‐N2A‐ex4 prevented increases in TNF production in response to high NaCl concentration and reduced transcriptional activity of an NFAT5 promoter construct by more than 60%. Since NKCC2A expression is restricted to the TAL segment of the nephron, NKCC2A‐dependent activation of NFAT5 is part of a pathway contributing to TNF production by the TAL in response to hypertonic NaCl intake.
Augmented response of renal sympathetic nerve single units to chemoreceptor stimuli in rabbits with angiotensin induced hypertension
Authors: Head Geoffrey A , Burke Sandra L , Lukoshkova Elena V
Published: June 2021

We examined the effect of angiotensin (AngII) hypertension on the activity of post‐ganglionic renal sympathetic units to determine whether elevated whole renal nerve activity is due to recruitment or increased firing frequency. Rabbits were treated with AngII for 4 weeks (50ng/kg/min, high dose) or for 8 weeks (20ng/kg/min, low dose) and then chloralose‐urethane anesthetized to test unitary responses to altered chemoreceptor activity. Units were detected from multi‐unit recordings using a novel algorithm that separated units by action potential shape characteristics using templates that matched spikes within a prescribed SD. Resting firing frequency was similar in sham (1.0±0.04 spikes/s, n=144) and high AngII rabbits (1.1±0.04 spikes/s, n=112) but was lower with low AngII (0.65±0.04 spikes/s, n=149). Firing frequency increased markedly to hypoxia (8% O2) in the high AngII group (+103%) but only by +39% in low AngII and +82% in sham group. The increase in firing rate with hypercapnia (8% CO2) was also greatest in the high AngII group whilst 100% O2 suppressed unitary firing in high AngII and sham groups. This study suggests that increases in sympathetic outflow in low dose AngII hypertension are most likely due to recruitment of individual neurons but high dose AngII markedly increases firing frequency to chemoreceptor stimuli. Supported by grants from National Health and Medical Research Council of Australia.
A clinic‐academic partnership for recruitment using an electronic medical record (EMR) in a trial of diets for treating polycystic ovary syndrome (PCOS) in overweight and obese adolescents and young adults
Authors: Wong Julia M W , Gooding Holly , Gordon Catherine M , Emans S Jean , Ludwig David S , Ebbeling Cara B
Published: June 2021

BACKGROUND

The Androgen Excess and PCOS Society Position Statement specifies lifestyle modification as the primary therapy for obesity in PCOS. However, consensus on optimal diet is lacking. Routine clinical care often includes OCPs and/or insulin sensitizers to lower androgens.

OBJECTIVE

To recruit girls with PCOS in a tertiary care center for a 6‐mo pilot RCT of low‐fat vs. low‐glycemic load diets on bioavailable testosterone (primary outcome). Use of medications that mask dietary effects is exclusionary.

METHODS

A partnership between the clinic and research teams facilitated recruitment using a flagging protocol to identify potentially eligible patients based on age (13–21y), BMI (≥85th percentile), and PCOS‐related keywords pertaining to visit reason in an EMR. When appropriate, providers presented the RCT as a first‐line non‐pharmacologic treatment option.

RESULTS

Over a 2‐y period, 1026 girls have been flagged, with the majority identified from the Division of Adolescent Medicine at BCH and its Reproductive Endocrinology Clinic (>;40%). Primary visit reasons were irregular menses (46%) and PCOS (34%). Medication use was the most frequent reason for exclusion (26%).

CONCLUSION

Partnerships using an EMR offer a contemporary approach to recruitment for RCTs of non‐routine clinical therapies. The ongoing RCT will examine the effects of different macronutrient diets on hyperandrogenism in PCOS.

Grant Funding Source: Thrasher Research Fund
Effect of sugar‐sweetened soft drinks on serum uric acid and associated metabolic risk factors.
Authors: Crutchley Peter William , Morenga Lisa Te
Published: June 2021

Excess dietary fructose can lead to elevated serum uric acid (SUA) concentrations which could in turn increase the risk of metabolic syndrome. An important source of fructose worldwide is sugar or high fructose corn syrup sweetened beverages (SSBs). SSB intakes have increased in parallel with obesity and type 2 diabetes, and may have displaced milk beverages. Fruit juice, despite containing similar amounts of fructose has not been shown to elevate SUA, while milk may reduce SUA concentrations.

The aim of our 8 week, randomised controlled trial was to determine the effects on SUA and metabolic syndrome risk factors of replacing SSBs with non‐sugar sweetened beverages including diet soft drink, orange juice, or low‐fat milk in a relatively insulin resistant population with high habitual SSB intakes (≥500ml/d). Blood and anthropometric data were collected at week 0, 4, and 8. Dietary data and 24‐hour urine samples were collected at week 0 and 8. There were significant reductions in serum uric acid levels in the diet soft drinks, orange juice, and milk groups compared to SSBs (39, 44, & 44μmol/L; p<0.01). C‐reactive protein concentrations were significantly reduced in the juice group but not in the others (−1.8mg/L; p=0.025). No significant differences were found between interventions for fasting glucose, TAG, or HDL cholesterol concentrations, weight, BMI, blood pressure, or waist circumference. Our findings indicate that reducing SSB intakes by exchanging SSBs for non‐sugar sweetened beverages lowers serum uric acid concentrations, and improves some features of the metabolic syndrome independently of changes in adiposity.
Choroid plexus (CP) inorganic phosphate (Pi) transport: Stress‐induced alteration of PiT‐1 (Slc20A1) localization
Authors: Renfro J. Larry , Parker Sonda L. , Bataille Amy
Published: June 2021

The CP actively transports Pi from CSF to blood utilizing PiT‐2 (Slc20A2) Na‐dependent transporter. In rats, PiT‐2 is normally located in or near the CP microvilli, positioned for removal of Pi from CSF. PiT‐1 predominates in the vascular endothelium, but a faint, diffuse signal is seen in the CP cell cytosol. In human CP, PiT‐2 is localized as above; however, PiT‐1 predominates in the cytosol of CP epithelial cells with a very faint signal in the vascular endothelium. We hypothesized that the PiT‐1 location may be influenced by pre‐fixation cellular stress. Our objective was to examine the distribution of PiT‐1 in rat lateral CP before and after induction of the cellular stress response. Immunohistochemical localization of PiT‐1 followed three treatments: 1. Fixation immediately after sacrifice; 2. Fixation of explants after 7.5 h in DME/F12 at 37°C and 5%CO2 ; 3. Fixation of explants exposed to 100 μM ZnCl2 in DME/F12 for 6 h followed by Zn‐free medium for 1.5 h. After 7.5 h in control culture medium, PiT‐1 increased markedly in the CP epithelium and was membrane associated. Though fainter than non‐cultured CP, staining was present in the capillary endothelial cells. Following Zn‐induced stress, PiT‐1 was again predominately in the CP epithelial cell membranes, including the microvilli but was absent from the capillary endothelium. We conclude that Pi transport capacity of CP may change with cellular stress. Support by NSF.
Cannabinoid 1 Receptor Promotes Cardiac Dysfunction, Oxidative Stress, Inflammation, and Fibrosis in Diabetic Cardiomyopathy
Authors: PACHER PAL , Rajesh Mohanraj , Batkai Sandor , Mukhopadhyay Partha , Lee Wen‐Shin , Horvath Bela , Cinar Resat , Liaudet Lucas , Mackie Ken , Haskó György
Published: June 2021

Endocannabinoids and cannabinoid (CB)1 receptors have been implicated in cardiac dysfunction, inflammation, and cell death associated with various forms of shock, heart failure, and atherosclerosis. We explored the role of CB1 receptors in cardiac dysfunction, inflammation and cell death using a mouse model of type 1 diabetic cardiomyopathy. Diabetic cardiomyopathy was characterized by increased myocardial endocannabinoid anandamide levels, oxidative/nitrative stress, activation of p38/Jun kinase, mitogen‐activated protein kinases, enhanced inflammation, increased expression of CB1, advanced glycation end product and angiotensin II type 1 receptors[AT(1)R], p47(phox), β‐myosin heavy chain isozyme switch, accumulation of AGE, fibrosis, and decreased expression of sarcoplasmic/endoplasmic reticulum Ca(2+)‐ATPase. Pharmacological inhibition or genetic deletion of CB1 receptors attenuated the diabetes‐induced cardiac dysfunction and the above‐mentioned pathological alterations. Activation of CB1 receptors by endocannabinoids may play an important role in the pathogenesis of diabetic cardiomyopathy by facilitating MAPK activation, AT(1)R expression/signaling, AGE accumulation, oxidative/nitrative stress, inflammation, and fibrosis. Conversely, CB1 receptor inhibition may be beneficial in the treatment of diabetic cardiovascular complications.
Intravenous infusion of Bendavia during renal revascularization attenuates cardiac injury and dysfunction in swine renovascular hypertension (RVH)
Authors: Williams Barbara J , Eirin Alfonso , Ebrahimi Behzad , Zhang Xin , Woollard John R. , Tang Hui , Jordan Kyra L. , Crane John A. , Lerman Amir , Lerman Lilach O.
Published: June 2021

Percutaneous transluminal renal angioplasty and stenting (PTRS) for RVH may not revert the secondary cardiac dysfunction and remodeling. In experimental RVH, infusion of the tetrapeptide Bendavia (that inhibits mitochondrial permeability transition pore opening) during PTRS attenuated stenotic kidney inflammatory injury and improved its function. We hypothesized that adjunct Bendavia during PTRS would also attenuate cardiac remodeling after reversal of RVH. RVH was induced in 18 pigs, of which 6 were sham treated (RVH) and 12 underwent PTRS plus a 4h IV infusion of Bendavia (0.05 mg/kg) or vehicle (n=6, each); 6 normal pigs served as controls. Cardiac oxygenation and function were assessed 4 wks later by blood oxygen level‐dependent (BOLD)‐MRI and CT, and tissue studies performed. PTRS normalized blood pressure (Table). Systolic function was unchanged. Apoptotic activity (caspase‐3), hypoxia (BOLD‐R2*), inflammation (TNF‐α, IL‐6), and fibrosis (trichrome) remained elevated and diastolic function (E/A) impaired in PTRS+Vehicle, but all improved in PTRS+Bendavia (Figure). Thus, novel cardioprotective properties of Bendavia enhance cardiac recovery after reversal of experimental RVH, and may provide a novel therapeutic option.
PPARγ mediated Modulation of Acrolein‐Induced Oxidative Stress: Role of NAD(P)H Oxidase subunit gp91phox
Authors: Chug Neha , Marek Katarzyna , Yousefipour Zivar , Newaz Mohammad
Published: June 2021

PPARγ and its ability to influence oxidative stress (OS) has been the focus of inflammatory signaling. NAD(P)H oxidase enzyme, transcriptionally regulated by PPARγ provides oxidant in acrolein (Acr)‐induced OS. We explored the influence of PPARγ ligand GW1929 (GW) on OS induced by Acr in mice lacking gp91phox subunit of NAD(P)H oxidase. GW reduced tissue 8‐Isoprostane in both WT and KO mice by 35% and 43% (p<0.05) respectively, which was increased by Acr. Acr reduced Total Antioxidant Status (TAS) which is more marked in WT (61%) than the KO (54%) mice. GW enhances TAS in WT but did not affect KO mice. Compare to WT, KO mice exhibit a significantly higher xanthine oxidase (XO) activity (51%) but not cycloxygenase (COX). Acr increased both XO and COX in WT but not in KO mice. GW reduced this increase COX from 4.6±0.3 to 2.9±0.19 nmol/mg prt/min but did not affect XO. NFκB was significantly higher in Acr‐treated WT but not in KO mice. GW blunted this increase in WT and reduced basal NFκB in KO mice by 51% (p<0.05). On the other hand, Acr increased CD36 activity in KO mice (55%) but not in WT. GW blunted CD36 increase in KO mice but did not affect WT. These results suggest that other oxidant system complement in acrolein mediated toxicity in absence of functional NAD(P)H oxidase. We also suggest that beneficial effect of PPARγ ligand involves inhibition of NAD(P)H oxidase as in absence of NAD(P)H this effects are minimal.
ER Stress Regulator BiP and Mitochondrial Chaperone Mortalin Differentially Regulate RelA/p65 Activation and Endothelial Cell Inflammation
Authors: Leonard Antony , Rahman Arshad , Fazal Fabeha
Published: June 2021

ER stress and mitochondrial dysfunction is implicated in the pathogenesis of neurological disorders, metabolic diseases, and more recently inflammatory conditions. Endothelial cell (EC) inflammation is an important component of many inflammatory diseases including acute lung injury (ALI). Here, we show that endoplasmic reticulum (ER) stress regulator BiP and mitochondrial chaperone mortalin differentially regulate NF‐κB to cause EC inflammation. RNAi knockdown of BiP or mortalin, each impaired thrombin‐induced NFκB‐transcriptional activity and expression of ICAM‐1 and VCAM‐1 in EC. Further analysis revealed that the inhibitory effect of BiP depletion on these responses was due to blockade of phosphorylation/degradation of IκBα, serine 536 phosphorylation, nuclear translocation and DNA binding of RelA/p65. By contrast, depletion of mortalin had no effect on the above responses, raising the possibility that mortalin may mediate NFκB activity by regulating the nuclear events such as acetylation/methylation of RelA/p65 and/or chromatin remodeling. These results identify BiP and Mthsp75 as novel mediators of EC inflammation that may act in concert to regulate cytosolic and nuclear events to promote NF‐κB activity. Importantly, these data also reveal mechanistic links between ER stress, mitochondrial dysfunction and EC inflammation. (Supported by NIH HL096907)
Role of sphingolipids in fibrinogen‐induced cerebrovascular permeability
Authors: Muradashvili Nino , Khundmiri Syed Jalal , Tyagi Reeta , Lee Menq‐Jer , Lominadze David
Published: June 2021

We showed that at an elevated level fibrinogen (Fg), a marker of inflammation, compromises endothelial cell (EC) layer integrity and increases cerebrovascular permeability. Since we found that Fg increases level of sphingolipids in mouse brain ECs (MBECs), we tested the role of cholesterol and sphingolipids in Fg‐induced permeability of MBECs.

Role of cholesterol and sphingolipids in Fg‐induced albumin crossing of MBECs layer was evaluated with a dual‐tracer probing method (allows differentiation of paracellular and transcellular transports). We found that chelation of cholesterol by methyl‐beta‐cyclodextrin (MβCD) inhibited leakage of both low molecular weight (LMW) and high molecular weight (HMW) tracers through EC layer. However, inhibition of sphingolipid de‐novo synthesis by myriocin inhibited Fg‐induced overall leakage (assessed by crossing of HMW tracer through MBECs), but did not affect leakage of LMW tracer. In addition, myriocin inhibited cerebrovascular leakage of albumin in mice with elevated blood level of Fg.

The present study suggests that at high level Fg increases cerebrovascular permeability altering mainly the transendothelial transport by enhancing formation of functional caveolae via sphingolipid signaling. Thus, sphingolipids can be identified as new targets for therapeutic intervention that can modulate inflammation‐induced cerebrovascular permeability
Effects of Obesity and Exercise Training on Testosterone Levels in Male Rats
Authors: Arsenis Nicole , Disanzo Beth , You Tongjian
Published: June 2021

The purpose of this investigation was to study the effects of obesity and chronic exercise on sex organ testosterone levels in male rats. Lean and obese male Zucker rats were randomly assigned to a sedentary group and an exercise training group (lean sedentary: n=7, lean exercise: n=8, obese sedentary: n=7, obese exercise: n=7). The exercise group walked on a rat treadmill for 8 weeks. Testicular testosterone was extracted and total testosterone levels were measured by using radioimmunoassay. Compared to lean sedentary rats, obese sedentary rats had lower levels of testicular testosterone (concentration: 17.2±11.0 vs. 61.5±21.2 ng/g; total content: 50.8±29.4 vs. 191.5±80.3 ng; both P<0.001). There were no group differences between lean sedentary and lean exercise rats in testicular testosterone levels. However, compared to obese sedentary rats, obese exercise rats had higher testicular testosterone levels (concentration: 40.6±20.0 vs. 17.2±11.0 ng/g; total content: 119.6±52.4 vs. 50.8±29.4 ng, both P<0.05). These results support our previous findings that chronic exercise attenuated obesity‐related hypogonadism. Further studies are needed to clarify the underlying mechanisms.
Gluconeogenesis and hepatic glycogenolysis during exercise at the lactate threshold
Authors: Emhoff Chi‐An W , Messonnier Laurent A. , Horning Michael A. , Fattor Jill A. , Carlson Thomas J. , Brooks George A.
Published: June 2021

We studied the effects of training, exercise intensity, and [lactate] on gluconeogenesis (GNG) and hepatic glycogenolysis (GLY) in fasted men exercising at the lactate threshold (LT). Twelve men (6 untrained, 6 trained) completed 1 h of constant load exercise at power outputs (PO) corresponding to their LT. Trained men completed two additional 1‐h exercise sessions at 10% below the LT (LT‐10%), one of which included a lactate clamp (LT‐ 10%+LC). Relative exercise intensity was matched between the untrained at LT and the trained at LT‐10%, and [lactate] was matched in the LT and LT‐10%+LC trials. Flux rates were determined by primed continuous infusion of [6,6‐2H2]glucose and [3‐13C]lactate tracers. Exercise at LT corresponded to 67.6 ± 1.3 and 74.8 ± 1.7 %VO2peak in the untrained and trained men, respectively (P < 0.05). GNG was decreased in the trained men exercising at LT and LT‐10% compared to the untrained men, but increasing [lactate] in the LT‐10%+LC trial increased GNG (4.4 ± 0.9 mg·kg−1·min−1) compared to that in LT‐10% (1.7 ± 0.4 mg·kg− 1·min−1, P < 0.05). Hepatic GLY was higher in the trained than untrained men. We conclude that GNG contributes ~25–40 % of glucose output during exercise in fasted men. While trained men can achieve a higher PO at the LT, this increase in exercise intensity is accompanied by a decrease in GNG, which can be recovered by raising [lactate]. This research was supported by a gift from CytoSport.
Impact of Exercise Training on Cardiovascular Risk and Anti‐Risk Factors in Adolescent Boys
Authors: Koozehchian Majid , Nazem Farzad , Kreider Richard B
Published: June 2021

Adolescence is a critical period in the formation of cardiovascular risk factors (RF). Long‐term exercise training has been found to be reversely related to RF levels in adolescents.

PURPOSE

To investigate the effects of training over a 12‐week timeframe on cardiovascular RFs in adolescent boys.

METHODS

Forty two boys, including swimmers (SW, n=14), soccer players (SO, n=13), and a control group (C, n=15) (Age=11.8±1.38yrs; Ht=149±.8.38cm; Wt=40±7.8kg) volunteered for this study. Fasting blood samples were obtained prior to initiation of training, at weeks 4, 8, and 12. A 3×4 repeated measures ANOVA with LSD post hoc analysis was conducted to determine the difference between three groups in total cholesterol (TC), LDL, HDL, TC/HDL, HDL/TC, LDL/HDL and triglyceride (TG).

RESULTS

Results showed greater reduction in LDL/HDL and TG in SO compared to C group (p ≤ 0.05). No significant difference was observed in TC, LDL, HDL, TC/HDL, and HDL/TC across all three groups.

CONCLUSION

Exercise training had a significant effect to lower LDL/HDL and TG in adolescent athletes. Participation in regular training is recommended for adolescents to improve cardiovascular RF profiles. Significant effects of training on TC, LDL, HDL, TC/HDL, and HDL/TC levels were inconclusive for 12‐week training period.
Influence of sprint interval training on skeletal muscle mitochondria as determined by high‐resolution respirometry
Authors: Beals Joseph W , Le Catherine H , Scalzo Rebecca L , Binns Scott E , Giordano Gregory R , Klochak Anna L , Paris Hunter L , Sevits Kyle E , Bell Christopher , Chicco Adam J
Published: June 2021

Short‐term sprint interval training (SIT) improves skeletal muscle mitochondrial function, as reflected by increased maximal activity of several oxidative enzymes. To date, no studies have determined directly the influence of SIT on mitochondrial respiration. 6 sedentary, but healthy adults (3 males; 25±2 years, peak oxygen uptake: 39±2 ml/kg/min (mean±SE)) completed 9 sessions of 4–8, 30‐second sprints, over 21 days. Vastus lateralis samples were obtained prior to and 48‐hours following SIT. Mitochondrial respiratory function was determined in saponin permeabilized fibers using high‐resolution respirometry, combined with a variety of substrate, uncoupler and inhibitor combinations. Measures of maximal complex‐I+II (all substrates) supported mitochondrial respiration revealed 9–13% increases in respiratory efficiency following SIT (P<0.05). Total respiratory chain capacity per mg of muscle was unaltered, as was oxidative phosphorylation (OXPHOS) capacity and efficiency with palmitoylcarnitine + malate (fatty acid) as substrate. Conversely, OXPHOS capacity and efficiency were increased by 16–25% when pyruvate + malate (carbohydrate) was used as substrate, either in the presence or absence of fatty acid (P<0.05). These preliminary data indicate that 3 weeks of SIT selectively improves the efficiency of muscle carbohydrate oxidation, with no appreciable effect on fatty acid oxidation or total muscle respiratory capacity.
Determination of exercise modality employing serum metabolomics profiling in type 2 diabetes: relation to clinical outcomes
Authors: Yang Jaeun Jane , Palmnas Marie , Su Juliet , Kirtadi Lawrence , Vogel Hans , Sigal Ron , Shearer Jane
Published: June 2021

It is generally acknowledged that exercise training, both aerobic and resistance, is conducive to health and decreases chronic disease risk. However, little information exists on type of exercise that confers the greatest benefit. The purpose of this study is to use metabolomics strategy to i) interrogate the effects of exercise training modality on serum biochemistry and ii) compare changes in metabolomic profiles with clinical data (i.e. weight, BMI, HbA1c) to determine any correlation. Individuals with type 2 diabetes (n=40) were recruited as a part of the Diabetes Aerobic Resistance Exercise (DARE) study. After a 4‐week run‐in program, individuals were randomly assigned into one of four groups: control, aerobic, resistance or a combination of aerobic + resistance training for 22wk. Serum was collected at baseline and at 6mo. Serum samples were analyzed for metabolites using gas‐chromatography‐ mass spectrometry (GC‐MS) and Proton Nuclear Magnetic Resonance Spectroscopy (H1‐NMR). Changes in metabolism involved metabolites such as amino acids, organic acid, and carbohydrates. Aerobic exercise elevated the rate of tricarboxylic acid cycle and antioxidant activity, while resistance exercise lead to hypertrophy and urea markers. In summary, GC‐MS and NMR based metabolomics analysis proved to effectively reveal exercise modality and explain differential clinical outcomes between treatment groups.
Muscle perilipin 3 is reduced using in vitro and in vivo exercise models and negatively associated with exercise lipid oxidation
Authors: Covington Jeffrey D , Galgani Jose E. , Rustan Arild C , Zhang Zhengyu , Moro Cedric , Smith Steven R , Ravussin Eric , Bajpeyi Sudip
Published: June 2021

Perilipin 2 (PLIN2) and perilipin 3 (PLIN3) have been shown to play important roles in the regulation of lipid droplet dynamics. Our aim was to understand changes in PLIN2 and PLIN3 protein content with increased lipid utilization during both in vitro and in vivo models of exercise. Pharmacological treatment (30μM palmitate, 4μM forskolin, 0.5μM ionomycin), which mimics exercise in human myotubes in vitro, showed that PLIN2 protein was unaltered while PLIN3 was reduced (p=0.01). Likewise, using biopsies from the vastus lateralis before and after a long endurance exercise bout (50% VO2Max until 650 kcal energy expenditure obtained) in 20 healthy, young males (age: 24.0 ± 4.5yrs, BMI: 23.6 ± 1.8 kg/m2), we found no change in PLIN2 and a reduction in PLIN3 (p=0.03). Moreover, the change in PLIN3 during exercise was positively correlated with cumulative whole‐body lipid oxidation measured by indirect calorimetry (r = 0.53; p = 0.04), changes in ex vivo muscle palmitate oxidation (r = 0.63, p = 0.01), and negatively correlated with changes in intramyocelluar lipid (r = −0.47; p = 0.05). These data show that both in vitro and in vivo models of exercise do not change PLIN2 but decrease PLIN3 expression. The reduction in PLIN3 is associated with decreased whole body as well as muscle lipid oxidation.
Recruitment of serotonergic responses in cardiac vagal neurons during hypoxia and hypercapnia
Authors: Weigand Letitia A , Mendelowitz David
Published: June 2021

Parasympathetic activity to the heart increases following exposure to hypoxia and hypercapnia (H/H). The recruitment of excitatory 5‐HT receptor‐mediated responses in cardiac vagal neurons (CVNs) plays a role in the post‐H/H response, however the mechanisms underlying this recruitment remain unclear. As a first step toward understanding how H/H leads to increased 5‐HT signaling in rat CVNs we examined the responses of these neurons to 5‐HT in in vitro brain slices before, during, and for 20 minutes after H/H (15 min; 6% O2‐9% CO2). Recording of excitatory post‐synaptic currents evoked by photostimulation of caged 5‐HT (100 μM for 10 ms every 30 s) revealed an inward 5‐HT receptor‐mediated current that increased in magnitude during H/H. In CVNs exposed to H/H this inward current increased from a mean of −8 ± 1.3 pA during normoxia (95% O2‐5% CO2) to −40 ± 1.4 pA during H/H, and −75 ± 0.4 pA 20 minutes post‐H/H (n=7). Pretreatment with ondansetron (100 μM), a competitive 5‐HT3 receptor antagonist partially blocked the recruitment of 5‐HT responses (peak amplitude during H/H of −20 ± 0.9 pA, and −40 ± 1.2 pA 20 minutes post‐H/H), a respective reduction of 50% and 47% compared to responses in the absence of ondansetron.(n=3). These data suggest that 5‐HT3 receptor‐mediated responses are recruited during H/H and persist post‐H/H, possibly due to increases in 5‐HT receptor trafficking to the postsynaptic membrane in CVNs during H/H.
The Effect of L‐arginine Supplementation & Voluntary Wheel Running on Vascular Function in Rats with Chronic Kidney Disease
Authors: Martens Christopher R , Kuczmarski James M , Lennon‐Edwards Shannon , Edwards David G
Published: June 2021

Reduced nitric oxide (NO) synthesis contributes to risk for cardiovascular disease in chronic kidney disease (CKD). Vascular uptake of the NO precursor L‐arginine (ARG) is attenuated in rodents with CKD, resulting in reduced substrate availability for NO synthesis and impaired vascular function. We tested the effect of 4 weeks of voluntary wheel running (RUN) and/or ARG supplementation on vascular function in rats with CKD. 12 week old male Sprague Dawley rats underwent 5/6 ablation infarction surgery to induce CKD, or SHAM surgery as a control. CKD animals either remained sedentary (SED) or received one of the following interventions: supplemental ARG, RUN, or combined ARG+RUN. Interventions began 4 weeks after surgery to allow time for disease progression. Animals were sacrificed 8 weeks after surgery and endothelium dependent relaxation (EDR) was assessed by dose response to acetylcholine in aortic rings. EDR was significantly impaired in SED vs. SHAM animals after 8 weeks, demonstrated by an attenuated maximal response (Emax; 66.87% ± 6.43 vs 96.79% ± 1.27, p <0.05) and rightward shifted LogEC50 (−6.83 ± 0.08 vs −7.54 ± 0.04, p<0.05). EDR was not significantly improved by ARG or RUN alone, but was improved by combined ARG+RUN (p< 0.05 vs SED). The results suggest a synergistic effect between L‐arginine and exercise that may act as a novel therapy to reduce cardiovascular risk in CKD.

Supported by ACSM Foundation Grant