Ryan D. Sheldon

Impact of acute exposure to increased hydrostatic pressure and reduced shear rate on conduit artery endothelial function: a limb-specific response

Unlike quadrupeds, humans exhibit a larger hydrostatic pressure in the lower limbs compared with the upper limbs during a major part of the day. It is plausible that repeated episodes of elevated pressure in the legs may negatively impact the endothelium, hence contributing to the greater predisposition of atherosclerosis in the legs. We tested the hypothesis that an acute exposure to increased hydrostatic pressure would induce conduit artery endothelial dysfunction. In protocol 1, to mimic the hemodynamic environment of the leg, we subjected the brachial artery to a hydrostatic pressure gradient ( approximately 15 mmHg) by vertically hanging the arm for 3 h. Brachial artery flow-mediated dilation (FMD) was assessed in both arms before and following the intervention. In protocol 2, we directly evaluated popliteal artery FMD before and after a 3-h upright sitting (pressure gradient approximately 48 mmHg) and control (supine position) intervention. Our arm-hanging model effectively resembled the hemodynamic milieu (high pressure and low shear rate) present in the lower limbs during the seated position. Endothelium-dependent vasodilation at the brachial artery was attenuated following arm hanging (P < 0.05); however, contrary to our hypothesis, upright sitting did not have an impact on popliteal artery endothelial function (P > 0.05). These data suggest an intriguing vascular-specific response to increased hydrostatic pressure and reduced shear rate. Further efforts are needed to determine if this apparent protection of the leg vasculature against an acute hydrostatic challenge is attributable to posture-induced chronic adaptations.

Acute impact of intermittent pneumatic leg compression frequency on limb hemodynamics, vascular function, and skeletal muscle gene expression in humans

The mechanisms by which intermittent pneumatic leg compression (IPC) treatment effectively treats symptoms associated with peripheral artery disease remain speculative. With the aim of gaining mechanistic insight into IPC treatment, the purpose of this study was to investigate the effect of IPC frequency on limb hemodynamics, vascular function, and skeletal muscle gene expression. In this two study investigation, healthy male subjects underwent an hour of either high-frequency (HF; 2-s inflation/3-s deflation) or low-frequency (LF; 4-s inflation/16-s deflation) IPC treatment of the foot and calf. In study 1 (n = 11; 23.5 ± 4.7 yr), subjects underwent both HF and LF treatment on separate days. Doppler/ultrasonography was used to measure popliteal artery diameter and blood velocity at baseline and during IPC treatment. Flow-mediated dilation (FMD) and peak reactive hyperemia blood flow (RHBF) were determined before and after IPC treatment. In study 2 (n = 19; 22.0 ± 4.6 yr), skeletal muscle biopsies were taken from the lateral gastrocnemius of the treated and control limb at baseline and at 30- and 150-min posttreatment. Quantitative PCR was used to assess mRNA concentrations of genes associated with inflammation and vascular remodeling. No treatment effect on vascular function was observed. Cuff deflation resulted in increased blood flow (BF) and shear rate (SR) in both treatments at the onset of treatment compared with baseline (P < 0.01). BF and SR significantly diminished by 45 min of HF treatment only (P < 0.01). Both treatments reduced BF and SR and elevated oscillatory shear index compared with baseline (P < 0.01) during cuff inflation. IPC decreased the mRNA expression of cysteine-rich protein 61 from baseline and controls (P <0 .01) and connective tissue growth factor from baseline (P < 0.05) in a frequency-dependent manner. In conclusion, a single session of IPC acutely impacts limb hemodynamics and skeletal muscle gene expression in a frequency-dependent manner but does not impact vascular function.

Gestational exercise protects adult male offspring from high-fat diet-induced hepatic steatosis.

BACKGROUND & AIMS Mounting evidence indicates that maternal exercise confers protection to adult offspring against various diseases. Here we hypothesized that maternal exercise during gestation would reduce high-fat diet (HFD)-induced hepatic steatosis in adult rat offspring. METHODS Following conception, pregnant dams were divided into either voluntary wheel running exercise (GE) or wheel-locked sedentary (GS) groups throughout gestation (days 4-21). Post-weaning, offspring received either normal chow diet (CD; 10% fat, 70% carbohydrate, 20% protein) or HFD (45% fat, 35% carbohydrate, and 20% protein) until sacrificed at 4- or 8-months of age. RESULTS GE did not affect offspring birth weight or litter size. HFD feeding in offspring increased weight gain, body fat percentage, and glucose tolerance test area under the curve (GTT-AUC). Male offspring from GE dams had reduced body fat percentage across all ages (p<0.05). In addition, 8-month male offspring from GE dams were protected against HFD-induced hepatic steatosis, which was associated with increased markers of hepatic mitochondrial biogenesis (PGC-1α and TFAM), autophagic potential (ATG12:ATG5 conjugation) and hepatic triacylglycerol secretion (MTTP). CONCLUSIONS The current study provides the first evidence that gestational exercise can reduce susceptibility to HFD-induced hepatic steatosis in adult male offspring.

Impact of porcine maternal aerobic exercise training during pregnancy on endothelial cell function of offspring at birth.

The purpose of this investigation was to test the hypothesis that maternal exercise training during pregnancy enhances endothelial function in offspring at birth. Six-month-old gilts (n = 8) were artificially inseminated and randomized into exercise-trained (n = 4) and sedentary groups (n = 4). Exercise training consisted of 15 weeks of treadmill exercise. The thoracic aorta of offspring were harvested within 48 h after birth and vascular responsiveness to cumulative doses of endothelium-dependent (bradykinin: 10-11-10-6 M) and independent (sodium nitroprusside: 10-10-10-4 M) vasodilators were assessed using in vitro wire myography. Female offspring from the exercised-trained gilts had a significantly greater endothelium-dependent relaxation response in the thoracic aorta when compared with the male offspring and female offspring from the sedentary gilts. The results of this investigation demonstrate for the first time that maternal exercise during pregnancy produces an enhanced endothelium-dependent vasorelaxation response in the thoracic aortas of female offspring at birth.

New insights into the physiologic basis for intermittent pneumatic limb compression as a therapeutic strategy for peripheral artery disease

The capability for externally applied rhythmic limb compressions to improve the outcomes of patients with peripheral artery disease has been recognized for nearly a century. Modern technology has permitted the development of portable and cost-effective intermittent pneumatic compression (IPC) systems to be made readily available for affordable at-home use. Mounting clinical evidence attests to the effectiveness of this strategy, with improvements in claudication distance rivaling those seen with exercise training or pharmacologic interventions, or both. However, owing to a lack of mechanistic knowledge, whether current application protocols are optimized for clinical outcomes is unknown. Traditional thinking has suggested that IPC transiently elevates blood flow, which is purported to relieve ischemia, improve vascular function, and promote vascular remodeling. Surprisingly, much ambiguity exists regarding the physiologic stimuli and adaptations that are responsible for the clinical effectiveness of IPC treatment. This review presents and critically discusses emerging evidence that sheds new light on the physiologic and molecular responses to IPC therapy. These novel findings highlight the importance of characterizing the phasic changes in the hemodynamic profile during IPC application. Further, these studies indicate that factors other than the elevation in blood flow during this therapy should be taken into account when designing an optimal IPC device. Lastly, we advance the hypothesis that manipulation of IPC stimulation characteristics could potentially magnify the documented clinical benefits associated with this therapy. In conclusion, recent evidence challenges the physiologic basis on which current IPC systems were designed, and further research to elucidate the basic and clinical outcomes of alternate stimulation characteristics is necessary.

Differential regulation of adipose tissue and vascular inflammatory gene expression by chronic systemic inhibition of NOS in lean and obese rats

We tested the hypothesis that a decrease in bioavailability of nitric oxide (NO) would result in increased adipose tissue (AT) inflammation. In particular, we utilized the obese Otsuka Long Evans Tokushima Fatty rat model (n = 20) and lean Long Evans Tokushima Otsuka counterparts (n = 20) to determine the extent to which chronic inhibition of NO synthase (NOS) with Nω‐nitro‐l‐arginine methyl ester (L‐NAME) treatment (for 4 weeks) upregulates expression of inflammatory genes and markers of immune cell infiltration in retroperitoneal white AT, subscapular brown AT, periaortic AT as well as in its contiguous aorta free of perivascular AT. As expected, relative to lean rats (% body fat = 13.5 ± 0.7), obese rats (% body fat = 27.2 ± 0.8) were hyperlipidemic (total cholesterol 77.0 ± 2.1 vs. 101.0 ± 3.3 mg/dL), hyperleptinemic (5.3 ± 0.9 vs. 191.9 ± 59.9 pg/mL), and insulin‐resistant (higher HOMA IR index [3.9 ± 0.8 vs. 25.2 ± 4.1]). Obese rats also exhibited increased expression of proinflammatory genes in perivascular, visceral, and brown ATs. L‐NAME treatment produced a small but statistically significant decrease in percent body fat (24.6 ± 0.9 vs. 27.2 ± 0.8%) and HOMA IR index (16.9 ± 2.3 vs. 25.2 ± 4.1) in obese rats. Further, contrary to our hypothesis, we found that expression of inflammatory genes in all AT depots examined were generally unaltered with L‐NAME treatment in both lean and obese rats. This was in contrast with the observation that L‐NAME produced a significant upregulation of inflammatory and proatherogenic genes in the aorta. Collectively, these findings suggest that chronic NOS inhibition alters transcriptional regulation of proinflammatory genes to a greater extent in the aortic wall compared to its adjacent perivascular AT, or visceral white and subscapular brown AT depots.

Reduced hepatic eNOS phosphorylation is associated with NAFLD and type 2 diabetes progression and is prevented by daily exercise in hyperphagic OLETF rats

We tested the hypothesis that nonalcoholic fatty liver disease (NAFLD) is associated with reduced hepatic endothelial nitric oxide synthase (eNOS) activation status via S1177 phosphorylation (p-eNOS) and is prevented by daily voluntary wheel running (VWR). Hyperphagic Otsuka Long-Evans Tokushima Fatty (OLETF) rats, an established model of obesity, type 2 diabetes (T2D) and NAFLD, and normophagic controls [Long-Evans Tokushima Otsuka (LETO)] were studied at 8, 20, and 40 wk of age. Basal hepatic eNOS phosphorylation (p-eNOS/eNOS) was similar between LETO and OLETFs with early hepatic steatosis (8 wk of age) and advanced steatosis, hyperinsulinemia, and hyperglycemia (20 wk of age). In contrast, hepatic p-eNOS/eNOS was significantly lower (P < 0.05) in OLETF rats with T2D advancement and the transition to more advanced NAFLD with inflammation and fibrosis [increased tumor necrosis factor-α (TNF-α), CD68, and CD163 mRNA expression; 40 wk of age]. Reduced hepatic eNOS activation status in 40-wk OLETF rats was significantly correlated with reduced p-Akt/Akt (r = 0.73, P < 0.05), reduced serum insulin (r = 0.59, P < 0.05), and elevated serum glucose (r = -0.78, P < 0.05), suggesting a link between impaired glycemic control and altered hepatic nitric oxide metabolism. VWR by OLETF rats, in conjunction with NAFLD and T2D prevention, normalized p-eNOS/eNOS and p-Akt/Akt to LETO levels. Basal activation of hepatic eNOS and Akt are maintained until advanced NAFLD and T2D development in obese OLETF rats. The prevention of this reduction by VWR may result from maintained insulin sensitivity and glycemic control.

Aerobic exercise training in the treatment of non-alcoholic fatty liver disease related fibrosis.

Physiologically relevant rodent models of non‐alcoholic steatohepatitis (NASH) that resemble the human condition are limited. Exercise training and energy restriction are first‐line recommendations for the treatment of NASH. Hyperphagic Otsuka Long–Evans Tokushima fatty rats fed a western diet high in fat, sucrose and cholesterol for 24 weeks developed a severe NASH with fibrosis phenotype. Moderate intensity exercise training and modest energy restriction provided some improvement in the histological features of NASH that coincided with alterations in markers of hepatic stellate cell activation and extracellular matrix remodelling. The present study highlights the importance of lifestyle modification, including exercise training and energy restriction, in the regulation of advanced liver disease.

Chronic NOS inhibition accelerates NAFLD progression in an obese rat model

The progression in nonalcoholic fatty liver disease (NAFLD) to nonalcoholic steatohepatitis is a serious health concern, but the underlying mechanisms remain unclear. We hypothesized that chronic inhibition of nitric oxide (NO) synthase (NOS) via N(ω)-nitro-L-arginine methyl ester (L-NAME) would intensify liver injury in a rat model of obesity, insulin resistance, and NAFLD. Obese Otsuka Long-Evans Tokushima fatty (OLETF) and lean Long-Evans Tokushima Otsuka (LETO) rats received control or L-NAME (65-70 mg·kg(-1)·day(-1))-containing drinking water for 4 wk. L-NAME treatment significantly (P < 0.05) reduced serum NO metabolites and food intake in both groups. Remarkably, despite no increase in body weight, L-NAME treatment increased hepatic triacylglycerol content (+40%, P < 0.05) vs. control OLETF rats. This increase was associated with impaired (P < 0.05) hepatic mitochondrial state 3 respiration. Interestingly, the opposite effect was found in LETO rats, where L-NAME increased (P < 0.05) hepatic mitochondrial state 3 respiration. In addition, L-NAME induced a shift toward proinflammatory M1 macrophage polarity, as indicated by elevated hepatic CD11c (P < 0.05) and IL-1β (P = 0.07) mRNA in OLETF rats and reduced expression of the anti-inflammatory M2 markers CD163 and CD206 (P < 0.05) in LETO rats. Markers of total macrophage content (CD68 and F4/80) mRNA were unaffected by L-NAME in either group. In conclusion, systemic NOS inhibition in the obese OLETF rats reduced hepatic mitochondrial respiration, increased hepatic triacylglycerol accumulation, and increased hepatic inflammation. These findings suggest an important role for proper NO metabolism in the hepatic adaptation to obesity.

Mother's exercise during pregnancy programmes vasomotor function in adult offspring

•  What is the central question of this study? Adverse maternal behaviour during pregnancy is well known to have lifelong health consequences for offspring. We asked whether a positive maternal health behaviour may have positive long‐term effects for offspring. Specifically, we investigated whether aerobic maternal exercise improves nitric oxide signalling in femoral arteries of adult swine. •  What is the main finding and its importance? For the first time, a fetal programming outcome due to gestational exercise is reported in adult offspring. Second, contrary to previous speculation regarding the impact of in utero alterations on nitric oxide signalling, our data suggest differential myosin phosphorylation in vascular smooth muscle.