LS Diet Research Articles

Microvascular function is reduced in normotensive salt‐sensitive individuals independent of dietary sodium intake

Impairments in microvascular (MV) function are thought to occur early in cardiovascular disease development. We sought to determine if MV function is altered in salt‐sensitive (SS) adults compared to salt‐resistant (SR) adults during low (LS) and high (HS) sodium diets. Seven healthy SS adults and 7 healthy SR adults were studied (SS: 2M, 5F; 48±4 yrs; SR: 2M, 5F; 45±3 yrs). After a run‐in diet, subjects were randomized to a 7 day LS (20 mmol/d) and 7 day HS (300–350 mmol/d) diet (controlled feeding study). Salt sensitivity was defined as a >; 5 mmHg change in 24‐hour mean arterial BP (MAP) from the LS to HS diet. MV function was assessed using laser Doppler flowmetry during local heating (42°C). Cutaneous vascular conductance (CVC) was calculated as red blood cell flux/MAP and all data were expressed as a percentage of the maximum CVC (28 mM SNP, 43°C). 24 hour MAP (mmHg) increased in the SS group between diets, but was unchanged in the SR group (SS: LS: 85±2, HS: 92±3, p<0.05; SR: LS:83±3, HS: 83±2, p>;0.05). Plateau %CVCmax was impaired in the SR group on the HS diet compared to the LS diet (80±2 vs. 90±2%; p<0.05). There were no differences in plateau %CVCmax in the SS group between LS and HS diets (80±4 vs. 82±3%; p>;0.05) but both were impaired (p<0.05) compared to the SR group on the LS diet. These data suggest that MV function is impaired in SS adults independent of dietary sodium intake. Supported by 2 P20 RR016472‐11 & R01 HL 104106.

Role of the collecting duct insulin receptor in fluid homeostasis in response to high‐ and low‐NaCl diets.

Recently we showed reduced ENaC activity and blood pressure (BP) in mice lacking the insulin receptor (IR) in collecting duct (CD) principal cells by aquaporin‐2 promoter targeted Cre‐recombinase knockout (KO). Here we test whether KO mice have impaired ability to maintain homeostasis in response to chronic high‐ (HS, 5% NaCl) or low‐ (LS, 0.02% Na+) sodium diets. After 1‐week feeding, LS KO mice had 20% reduced 24‐hr urine volume, as compared to WT, and significantly reduced ratios of Na+ to Cl− and K+ in urine. Likewise, the Na+ to Cl− ratio was reduced in HS KO (relative to WT). BP (by telemetry) did not change in either genotype with HS diet, but decreased slightly by LS in both genotypes. Four weeks of HS or LS diet resulted in KO mice being modestly, but significantly (p = 0.046) lighter than WT littermates (~10%). HS diet increased kidney weight in both genotypes (~17%). Western blotting was used to evaluate principal cell proteins involved in electrolyte homeostasis. LS diet led to an increase in the α‐subunit of the epithelial sodium channel (ENaC) and the lower (75‐kDa) band of γ‐ENaC; however, band densities did not differ between genotypes. Serum and glucorticoid‐regulated kinase (sgk‐1) was increased by HS, and to a significantly greater extent in the WT mice (p = 0.038). Overall, these studies support a role for the renal CD‐IR in electrolyte homeostasis with chronic adjustments in dietary NaCl. NIH DK082507.

Microbial ecosystem and fermentation traits in the caecum of growing rabbits given diets varying in neutral detergent soluble and insoluble fibre levels

The effect of the level of neutral detergent fibre (NDF: 0.35, LI and 0.42, HI) and neutral detergent soluble fibre (NDSF: 0.14, LS and 0.17, HS) in the caecal ecosystem was studied in 24 weaned (28 days of age) rabbits, weighing 630 ± 80.2 g in a 2 × 2 factorial design. After 22 days, rabbits were slaughtered and their caecal contents sampled. The caecal pH (on average 6.2) and molar volatile fatty acids (VFA) proportions were not affected by dietary treatments, but total VFA concentration tended to be lower with NDF (84.7 vs. 74.1 mmol/l; P = 0.095). The amount of total bacteria tended (P = 0.075) to increase with NDSF, but only in diets with 0.35 NDF. The caecal proportions of Ruminococcus albus and Fibrobacter succinogenes were not affected by type or level of fibre, but Butyrivibrio fibrisolvens decreased (P = 0.055) with the NDF proportion in LS diets. Denaturing gradient gel electrophoresis (DGGE) analysis showed that bacterial communities clustered according to each combination of NDF and NDSF, but did not greatly differ among diets (similarity indexes between 0.67 and 0.70), nor biodiversity was affected (average Shannon and richness indexes 3.50 and 33.1; P > 0.10). Archaeal population revealed changes in the amount and composition that were particularly evident in HS diets, decreasing in concentration (from 4.37 to 4.12 log10 gene copy number/g) and biodiversity (Shannon index from 3.14 to 2.52 and richness index from 23.7 to 13.9) compared to LS. The type and level of dietary fibre had a minor impact on caecal fermentation traits or caecal bacterial community. However, the increase in NDSF from 0.14 to 0.17 reduced concentration and diversity of methanogenic archaea.

Heterozygous knockout of transforming growth factor-β1 protects Dahl S rats against high salt-induced renal injury

The present study employed a zinc-finger nuclease strategy to create heterozygous knockout (KO) rats for the transforming growth factor-β1 (Tgfb1) gene on the Dahl SS/Jr genetic background (TGF-β1(+/-) Dahl S). Intercrossing TGF-β1(+/-) rats did not produce any homozygous KO rats (66.4% +/-, 33.6% +/+), indicating that the mutation is embryonic lethal. Six-week-old wild-type (WT) littermates and TGF-β1(+/-) Dahl S rats were fed a 0.4% (low salt, LS) or 8% NaCl (high salt, HS) diet for 5 wk. Renal cortical expression of TGF-β1, urinary TGF-β1 excretion, proteinuria, glomerular injury and tubulointerstitial fibrosis, and systolic blood pressure were similar in WT and TGF-β1(+/-) Dahl S rats maintained on the LS diet. The expression and urinary excretion of TGF-β1 increased to a greater extent in WT than in TGF-β1(+/-)Dahl S rats fed an HS diet for 1 wk. Systolic blood pressure rose by the same extent to 235 ± 2 mmHg in WT and 239 ± 4 mmHg in TGF-β1(+/-) Dahl S rats fed a HS diet for 5 wk. However, urinary protein excretion was significantly lower in TGF-β1(+/-) Dahl S than in the WT animals. The degree of glomerular injury and renal cortical and outer medullary fibrosis was markedly less in TGF-β1(+/-) than in WT rats. These findings suggest that the loss of one copy of the TGF-β1 gene blunts the increase in renal TGF-β1 protein expression and slows the progression of proteinuria, glomerulosclerosis, and renal interstitial fibrosis in Dahl S rats fed an HS diet independently of changes in blood pressure.

Dietary sodium loading impairs microvascular function independent of blood pressure in humans: role of oxidative stress

Animal studies have reported dietary salt-induced reductions in vascular function independent of increases in blood pressure (BP). The purpose of this study was to determine if short-term dietary sodium loading impairs cutaneous microvascular function in normotensive adults with salt resistance. Following a control run-in diet, 12 normotensive adults (31 ± 2 years) were randomized to a 7 day low-sodium (LS; 20 mmol day(-1)) and 7 day high-sodium (HS; 350 mmol day(-1)) diet (controlled feeding study). Salt resistance, defined as a 5 mmHg change in 24 h mean BP determined while on the LS and HS diets, was confirmed in all subjects undergoing study (LS: 84 ± 1 mmHg vs. HS: 85 ± 2 mmHg; P > 0.05). On the last day of each diet, subjects were instrumented with two microdialysis fibres for the local delivery of Ringer solution and 20 mm ascorbic acid (AA). Laser Doppler flowmetry was used to measure red blood cell flux during local heating-induced vasodilatation (42°C). After the established plateau, 10 mm l-NAME was perfused to quantify NO-dependent vasodilatation. All data were expressed as a percentage of maximal cutaneous vascular conductance (CVC) at each site (28 mm sodium nitroprusside; 43°C). Sodium excretion increased during the HS diet (P < 0.05). The plateau % CVCmax was reduced during HS (LS: 93 ± 1 % CVCmax vs. HS: 80 ± 2 % CVCmax; P < 0.05). During the HS diet, AA improved the plateau % CVCmax (Ringer: 80 ± 2 % CVCmax vs. AA: 89 ± 3 % CVCmax; P < 0.05) and restored the NO contribution (Ringer: 44 ± 3 % CVCmax vs. AA: 59 ± 6 % CVCmax; P < 0.05). These data demonstrate that dietary sodium loading impairs cutaneous microvascular function independent of BP in normotensive adults and suggest a role for oxidative stress.

Abstract 241: Influence of Dietary Salt and Renin-angiotensin System activity on plasma Retinol-binding Protein 4 in Healthy Human Subjects

Introduction: Retinol-binding protein 4 (RBP-4) is a circulating adipokine associated with insulin resistance, obesity and cardiovascular risk. Previous evidence suggests a negative impact on insulin sensitivity via an interaction with the insulin-stimulated glucose transporter (GLUT-4), specifically in adipocytes. Since angiotensin-2 (ANG-2) impairs GLUT-4 presentation and insulin sensitivity, we hypothesized that renin-angiotensin system (RAS) activity might affect RBP-4 expression as well. Furthermore, both RAS activity and insulin sensitivity are affected by dietary salt, but the relationship between these factors and RBP-4 expression is unknown. We hypothesized that liberal salt diet, via suppression of RAS activity, would be associated with reduced RBP-4 levels compared to salt restriction. METHODS: Healthy subjects (7 men, 7 women, mean age 24 yr, BMI 22.6) were studied on low salt (LS 10 mmol Na/day) diet and then liberal salt (HS 200 mmol Na/day) diet for 1 week each. Samples were drawn after overnight rest in the supine position for plasma renin activity (PRA), serum Aldosterone (Aldo) and plasma RBP-4 and again after ANG-2 infusion (3ng/kg/min x 45 mins). Results: Salt manipulation predictably modified RAS activity (HS v. LS Aldo, 4.5 ± 3.8 v. 19.5 ± 8.5 ng/dl [p&lt;0.01]; HS v. LS PRA, 0.5 ± 0.4 v. 2.7 ± 1.4 ng/ml/hr [p&lt;0.01]). RBP-4 was significantly lower on a HS compared to LS diet (35.4 ± 6.6 v. 38.6 ± 7.9 mg/L, p= 0.017). No significant change was observed in RBP-4 level after ANG-2 infusion on either diet. Fasting insulin levels trended lower on HS compared to LS (3.9 ± 6.8 v. 4.4 ± 1.2 μIU/ml, p=0.09), but there was no difference in fasting glucose (81.9 ± 6.8 v. 82.1 ± 4.7 mg/dl, p=0.92). Conclusion: RBP-4 levels are modified by dietary salt intake. RBP-4 levels were lower under HS conditions (suppressed RAS) as compared to LS conditions. Acute administration of ANG-2 does not increase RBP-4 levels in this healthy population. Insulin response was similar to prior reports suggesting that the connection between RAS activity, dietary salt and insulin sensitivity may include RBP-4. Future work exploring the relationship between RBP-4, GLUT-4 and insulin sensitivity should take into consideration the effect of dietary salt intake.

Effect of dietary supplementation with live-cell yeast at two dosages on lactation performance, ruminal fermentation, and total-tract nutrient digestibility in dairy cows

The experimental objective was to determine the effect of dietary supplementation with live-cell yeast (LCY; Procreatin-7, Lesaffre Feed Additives, Milwaukee, WI) at 2 dosages in high-starch (HS) diets [30% starch in dry matter (DM)] on lactation performance, ruminal fermentation, and total-tract nutrient digestibility in dairy cows compared with HS or low-starch (LS; 20% starch in DM) non-LCY diets. Sixty-four multiparous Holstein cows (114±37 d in milk and 726±74kg of body weight at trial initiation) were randomly assigned to 32 electronic gate feeders (2 cows per feeder), which were randomly assigned to 1 of 4 treatments in a completely randomized design. A 2-wk covariate adjustment period with cows fed a 50:50 mixture of the HS and LS diets was followed by a 12-wk treatment period with cows fed their assigned treatment diets. The HS diets were fed without (HS0) and with 2 (HS2) or 4 (HS4) g/cow per day of LCY. The LS diet did not contain LCY (LS0) and was formulated by partially replacing dry ground shelled corn with soy hulls. Cows fed LS0 consumed more DM than cows fed HS diets during wk 3, 10, 11, and 12. Yields of actual (44.5kg/d, on average), fat-, energy-, and solids-corrected milk were unaffected by treatment. Milk fat content tended to be greater for LS0 than for HS0 and HS2 but not different from HS4. Milk urea nitrogen contents were greater for cows fed LS0 than for cows fed the HS diets. Feed conversion (kg of milk/kg of DM intake) was numerically greater for HS diets than for LS0. Ruminal pH was unaffected by treatment. Ruminal molar proportion of acetate was greater, whereas that of propionate was lower, for LS0 compared with HS diets. Dry matter and organic matter digestibilities were greater for HS2 and HS4 than for HS0. Digestibility of neutral detergent fiber was greater for HS4 than for HS0 and HS2. Dry matter, organic matter, and neutral detergent fiber digestibilities were greater for LS0 than for HS diets; starch digestibility was greater for LS0 than for HS0 and HS4. Feeding LS0 increased DM intake and milk fat content, but reduced feed conversions. The addition of 4g/cow per day of LCY to HS diets tended to increase milk fat content and increased total-tract fiber digestibility in dairy cows.

Effects of a high maternal sodium intake on offspring of genetically diverse (CCHS) mice

The present study sought to determine whether a high maternal salt intake during pregnancy and lactation affected offspring blood pressure (BP), heart rate (HR), or locomotor activity in a genetically diverse stock of mice (Collaborative Cross Heterozygous Stock, CCHS). Dams were fed a low or high salt diet (0.25% NaCl (LS) vs 5% NaCl (HS)) from before conception through weaning, and telemetry studies were performed in 10 month old male offspring raised on either the LS or HS diet. A maternal HS diet was associated with significantly smaller litter sizes (P=0.03, data from groups of 34 and 37 litters), but offspring weight was not affected at any time between the day of birth (data from 22 and 24 litters) and 10 months of age (group sizes ≥ 11). Telemetry studies revealed no effects of maternal HS on mean BP or HR. However, maternal HS was associated with increased circadian variation in BP, increased percentage of time spent active during the 12h light period, and a tendency towards increased pulse pressure (P=0.03, 0.04, 0.06, respectively, group sizes of 8 – 14). Our results indicate that a maternal high salt diet does not affect offspring mean BP level in 10 month old male CCHS mice, but does reduce litter size and has subtle effects on activity and blood pressure cycles. Supported by the Canadian Institutes of Health Research (#91547).

Development of diet induced obesity models in rats, comparison of metabolic and genetic effects

The development of dietary obesity models in rats mimicking human obesity may help understanding its pathology and to evaluate new therapeutic strategies for treatment. We fed male Wistar rats either with a control chow diet (C), a 45%Kcal lard diet + 5% sucrose in water (LS) a chow diet +50%Kcal lard + 5% sucrose in water (CLS) or a chow diet + 20% sucrose in water (CS) for 22 weeks (W). At the end of study LS and CS diets increased hepatic and serum triglycerides, and these differences were found in CS diet since w 4. Hepatic SREBP‐1 mRNA was higher in LS and CS but FAS mRNA only increased in CS group. The oral glucose tolerance test (OGTT) at 18 W revealed that rats fed LS, CLS and CS developed insulin resistance. Body weight was 29% higher in rats fed LS than the rest of the groups due to increased adipose tissue (AT) mass. Subcutaneous (S) and visceral (V) AT was increased 3 fold in LS with respect to C, 2 fold with respect to CLS and 75% with respect to CS. HSL, perilipin and PPAR gamma mRNA was higher in SAT of LS and CS rats, whether in VAT those genes increased only in CS and CLS. CS diet increased 3 fold the glomerular filtration rate. In conclusion this study demonstrates than the fat and carbohydrate content of the diet differentially induces obesity and comorbidities.Grant Funding Source: Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán

Histone demethylase LSD1 deficiency during high-salt diet is associated with enhanced vascular contraction, altered NO-cGMP relaxation pathway, and hypertension

Histone methylation, a determinant of chromatin structure and gene transcription, was thought to be irreversible, but recent evidence suggests that lysine-specific demethylase-1 (LSD1, Kdm1a) induces demethylation of histone H3 lysine 4 (H3K4) or H3K9 and thereby alters gene transcription. We previously demonstrated a human LSD1 phenotype associated with salt-sensitive hypertension. To test the hypothesis that LSD1 plays a role in the regulation of blood pressure (BP) via vascular mechanisms and gene transcription, we measured BP and examined vascular function and endothelial nitric oxide (NO) synthase (eNOS) expression in thoracic aorta of male wild-type (WT) and heterozygous LSD1 knockout mice (LSD1(+/-)) fed either a liberal salt (HS; 4% NaCl) or restricted salt diet (LS; 0.08% NaCl). BP was higher in LSD1(+/-) than WT mice on the HS diet but not different between LSD1(+/-) and WT mice on the LS diet. Further examination of the mechanisms of this salt-sensitive hypertension in LSD1(+/-) mice on the HS diet demonstrated that plasma renin activity and plasma levels and urinary excretion of aldosterone were less in LSD1(+/-) than WT, suggesting suppressed renin-angiotensin-aldosterone system. In contrast, phenylephrine (Phe)-induced aortic contraction was greater in LSD1(+/-) than WT mice on the HS diet. Treatment of aortic rings with 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ; a blocker of guanylate cyclase) enhanced Phe contraction in LSD1(+/-) compared with WT mice on the HS diet. Acetylcholine (Ach)-induced relaxation was less in LSD1(+/-) than WT mice on the HS diet. Endothelium removal or pretreatment with N(ω)-nitro-L-arginine methyl ester (blocker of NOS) or ODQ abolished Ach-induced relaxation in aorta of WT but had minimal effect in LSD1(+/-). Vascular relaxation to sodium nitroprusside, an exogenous NO donor and guanylate cyclase activator, was decreased in LSD1(+/-) vs. WT mice on the HS diet. RT-PCR and Western blots revealed decreased eNOS mRNA expression and eNOS and guanylate cyclase protein in the heart and aorta of LSD1(+/-) compared with WT mice on HS diet. Thus, during the HS diet, LSD1 deficiency is associated with hypertension, enhanced vascular contraction, and reduced relaxation via NO-cGMP pathway. The data support a role for LSD1-mediated histone demethylation in the regulation of NOS/guanylate cyclase gene expression, vascular function, and BP during the HS diet.

Celecoxib does not alter cardiovascular and renal function during dietary salt loading

1. Cyclo-oxygenase-2 (COX-2)-derived prostaglandins are important in controlling sodium excretion and renin release. In the present study, we tested the hypothesis that a clinical dose of celecoxib would impair urinary sodium excretion and elevate blood pressure (BP) during dietary salt loading. 2. Twelve normotensive individuals (mean (± SEM) age 35 ± 2 years) completed two separate 17 day dietary perturbations, one taking 200 mg/day celecoxib (CX2) and the other taking placebo (PL), randomized with a 1 month wash out. The controlled 17 day diet consisted of a 3 day run-in diet, 7 days of a low-salt (LS, 20 mmol sodium/day) diet and 7 days of a high-salt diet (HS, 350 mmol sodium/day) diet. The order in which the diets were applied was randomized. Data were collected on the last day of the LS and HS diets. 3. Plasma and urinary prostaglandins were modestly lower during celecoxib (P < 0.05). Urinary sodium excretion was greater (P < 0.01) during the HS diet (253 ± 10 vs 281 ± 27 mmol/24 h for PL vs CX2, respectively) compared with the LS diet (14 ± 3 vs 17 ± 7 mmol/24 h for PL vs CX2, respectively; P(drug) = 0.26). Celecoxib did not alter creatinine clearance (P > 0.50). Twenty-four hour mean arterial BP was similar during PL (87 ± 2 vs 87 ± 2 mmHg for LS and HS, respectively) and CX2 (88 ± 2 vs 87 ± 2 mmHg for LS and HS, respectively; P = 0.85), with no effect of dietary salt (P > 0.80). Plasma renin activity, angiotensin II and aldosterone were all suppressed with dietary salt loading (P < 0.05), with no effect of drug (P > 0.35). 4. In conclusion, blood pressure and renal function were not adversely affected by celecoxib, even during dietary salt loading. These findings support current guidelines suggesting minimal cardiovascular risks associated with short-term, low-dose use of celecoxib in young to middle-aged adults.

The effect of estrogen and a high salt diet on arteriolar reactivity to increased tissue oxygen in the spinotrapezius muscle of the female SS rat

Estrogen may protect against salt induced microvascular dysfunction. Female SS (Dahl Salt Sensitive, SS/JrHsdMcwi) were divided into 2 subgroups: ovariectomized with (OVX +E) or without (OVX‐E) estrogen replacement. Three days prior to the acute experiment the diet of half of the rats was changed from low salt (LS, 0.4% NaCl) to high salt (HS, 8.0% NaCl). On the day of the acute experiment a transverse arteriole in the spinotrapezius muscle was selected, allowed to equilibrate with 0% O2 in the superfusion solution, and then changes in arteriolar diameter to 21% O2 in the superfusion solution before and after the topical application of HET‐0016 (10μM), a selective inhibitor of 20‐HETE production, were measured. Neither estrogen nor HS affected arteriolar constriction to 21% O2. HET‐0016 significantly attenuated arteriolar constriction to 21% O2 in the SS rat with or without estrogen and on a LS diet. However, the inhibition was greater in the SS rat without estrogen. HET‐0016 did not significantly inhibit the oxygen induced arteriolar constriction in the SS rat with or without estrogen and on a HS diet. These results suggest that a HS diet and estrogen reduce the contribution of 20‐HETE to oxygen induced arteriolar constriction in the spinotrapezius muscle. Funded by an internal UW‐M Research Growth Initiative grant.

Impaired urinary nitrates plus nitrites (UNOx) in high‐NaCl fed Ksp‐cadherin‐targeted insulin receptor knockout (KO) mice is insensitive to Tempol

Previously, we showed significantly reduced UNOx in mice lacking the insulin receptor in renal tubule (primarily distal) by targeted knockout. To address the possibility that increased destruction of NO by superoxide in the KO might explain these findings, 3-month-old male KO and WT littermates were fed serially medium- (0.5% NaCl, MS), low- (0.08% NaCl, LS), and high-NaCl (5% NaCl, HS) diets for one week each followed by a week of HS plus 3 mM Tempol, a superoxide dismutase mimetic, in drinking water. On day 5 each week, 24-hr urine was collected. Starting body weights (BW) were not different. However, final BWs were significantly higher in KO mice (g): 30.5 ± 0.7, WT versus 33.1 ± 0.4, KO, p = 0.03. Urine volumes, sodium or potassium were not significantly different between genotypes under any condition. No genotypic differences in UNOx were observed when mice were fed the LS diet (μmol/kg·bw/d): 28.6 ± 4.0, WT and 27.4 ± 5.7, KO, p = 0.87. HS feeding led to a small decrease in WT, but decreased UNOx in KO by over 50% (20.2 ± 4.3, WT and 11.3 ± 0.5, KO, p = 0.057). Tempol did not correct this difference, nor significantly affect WT levels of UNOx (21.1 ± 2.3, WT and 9.2 ± 2.7, KO, p = 0.036). Our results suggest that KO mice are more sensitive to HS feeding with regard to UNOX and weight gain. This deficit in UNOx does not appear to be due to relatively increased superoxide activity. Thus genotypic differences may be the result of reduced NO production.

Effects of Synchronization of Carbohydrate and Protein Supply on Ruminal Fermentation, Nitrogen Metabolism and Microbial Protein Synthesis in Holstein Steers

Three rumen-cannulated Holstein steers were fed three diets, each with a different synchrony index (SI) (LS: 0.77, MS: 0.81, and HS: 0.83), in order to examine the effect of diet on rumen fermentation, nitrogen balance, and microbial protein synthesis. Synchrony index was calculated based on the carbohydrate and crude protein fractions of each ingredient and their degradation rates. Feeding the steers diets with different SIs did not influence dry matter, crude protein, NDF, or ADF digestibility. The concentrations of total and individual VFA in the rumens of steers that were fed the two higher-SI diets were higher than in those fed the low-SI diet (p<0.05), but there was no significant difference between the two higher-SI diets. One hour after feeding, steers on the LS diet had lower ruminal pHs than did those fed the MS or HS diets (p<0.05), and animals on the LS diet generally showed higher ruminal NH3-N levels than did animals on the other diets, with the 4-h post-feeding difference being significant (p<0.05). Steers receiving the LS diet excreted more nitrogen (N) in their urine than did those on the two higher-SI diets (p<0.05), and the total N excretion of those on the LS diet was also higher (p<0.05). Microbial N levels calculated from the concentration of urinary purine derivatives were generally higher when the SI was higher, with the highest microbial protein synthesis being produced by steers on the HS diet (p<0.05). In conclusion, in the current study, ingestion of a synchronous diet by Holstein steers improved microbial protein synthesis and VFA production and

Modulation by Cytochrome P450-4A ω-Hydroxylase Enzymes of Adrenergic Vasoconstriction and Response to Reduced PO2 in Mesenteric Resistance Arteries of Dahl Salt-Sensitive Rats

This study evaluated the contribution of the 20-HETE/cytochrome P450-4A ω-hydroxylase (CYP4A) system to the early development of salt-induced vascular changes in Dahl salt-sensitive (SS) rats. CYP4A expression and 20-HETE production were evaluated and responses to norepinephrine, endothelin, and reduced PO₂ were determined by video microscopy in isolated mesenteric resistance arteries from SS rats fed high salt (HS; 4% NaCl) diet for three days vs. low salt (LS; 0.4% NaCl) controls. CYP4A enzyme inhibition with dibromododecenyl methylsulfimide (DDMS) selectively reduced norepinephrine sensitivity and restored impaired vasodilation in response to reduced PO₂ in SS rats fed HS diet. In the presence of DDMS, vasodilatation to reduced PO₂ was eliminated by indomethacin and unaffected by l-NAME in rats fed LS diet, and eliminated by l-NAME and unaffected by indomethacin in rats fed HS diet. The 20-HETE agonist WIT003 restored norepinephrine sensitivity in DDMS-treated arteries of HS-fed rats. HS diet increased vascular 20-HETE production and CYP4A protein levels by ∼24% and ∼31%, respectively, although these differences were not significant. These findings support the hypothesis that the 20-HETE/CYP4A system modulates vessel responses to norepinephrine and vascular relaxation to reduced PO₂ in mesenteric resistance arteries of SS rats fed HS diet.

Angiotensin II Increases JNK and IRS‐1ser307 Activity via NAD(P)H and Inhibits Insulin Signaling in Rats with Insulin Resistance Induced by Low Salt Diet

Low salt diet (LS) increases angiotensin II (AII), which may enhance JNK and IRS1ser307 phosphorylation (PP) decreasing insulin signaling (INS‐S). AII stimulate ROS production through activation of NAD(P)H oxidase (NAD). However, whether AII, oxidative stress (OX) and inflammatory signaling (IS) are involved in INS resistance induced by LS is not established.ObjectiveEvaluate if losartan (LOS) improves INS resistance by decreasing OX, JNK and IRS1ser307 PP.MethodsRats were fed a LS (0.15% NaCl) or normal salt diet (NS: 1.3%) since weaning. Groups: NS, NS+LOS, LS and LS+LOS. NAD activity (subunits p47 and gp91PHOX), tissue AII, blood glucose (G), plasma INS, HOMA, INS‐S and IS, AII type 1 (AT1) receptor gene and protein expression were evaluated.Results(P&lt;0.05, n=8): INS level, HOMA index, muscle AII and NAD subunits PP were greater in LS (p47: 10,066±975, gp91: 740±106) than in NS (p47: 5,783±801, gp91: 416±70 AU). G and NAD activity were lower in LS+LOS than in LS. INS receptor and Akt PP were greater and JNK (58±3) and IRS1ser307 PP (47±3 AU) were lower in the muscle of LS+LOS than in LS. IkB‐α PP was lower in LS (50±3) than in NS (100±3 AU) suggesting an activation of IS in LS. AT1 did not differ among groups.ConclusionsINS resistance induced by LS is associated with alterations in regulation of INS‐S, activated IS (JNK and IkB‐α), and higher tissue AII and NAD activity. In LS, LOS improves INS‐S by decreasing OX, JNK and IRS1ser307 PP, indicating that LOS may has anti‐inflammatory properties. AII may be a pathway that connects IS, OX and INS resistance in LS.Funded by FAPESP

Health and performance of Omani sheep fed salt-tolerant sorghum ( Sorghum bicolor) forage or Rhodes grass ( Chloris gayana)

Sorghum variety Super Dan was planted and irrigated with water containing three different concentrations of salt: 3, 6 and 9 dS/m. The sorghum fodder was termed low salinity sorghum (LSS), medium salinity sorghum (MSS) and high salinity sorghum (HSS). The sorghum was manually harvested, dried and chopped before feeding. Thirty-two, 3-month-old Omani male lambs were randomly distributed into four groups of eight lambs each. The first group was fed a control diet of Rhodes grass hay (RGH) plus a commercial concentrate. The other groups were given one of three sorghum hays irrigated with water containing one of the three different concentrations of salt plus the commercial concentrate. Daily feed intakes and weekly BW were determined throughout the experimental period of 11 weeks of which the first two were regarded as an adaptation period. A digestibility trial was carried out using 12 animals (3 sheep per diet) consisting of 10 days of adaptation and a subsequent 10 days collection period for faeces and urine. Blood samples were drawn three times during the experiment and analyzed for haematological and serum biochemistry levels. At the end of the trial the animals were slaughtered. The RGH had higher mineral content than sorghum forage grown under various levels of salinity. Animals fed sorghum-based diets did not show any signs of ill health. There were no differences ( P > 0.05) in digestibility coefficients of acid detergent fibre, neutral detergent fibre, and ether extract between RGH, LSS, MSS and HSS diets. However, the LS diet had lower DM, Ca, CP, P and energy digestibilities but higher ash digestibility. There were no treatment effects on hay, concentrate or total feed intake; total body weight gain or gain per kg/body weight of experimental animals. Sheep fed the RGH, LSS, MSS and HSS diets had average daily body weight gains of 96, 84, 82 and 68 g/day, respectively. There was no diets effect on rumen condition except that RGH-fed animals had lower N-ammonia and butyric acid concentration. This study indicated that sorghum forage grown under high salinity levels may be used for feeding Omani sheep without adverse effects on health or performance.

Role of postnatal dietary sodium in prenatally programmed hypertension

In this study we examined the short- and long-term impact of early life dietary sodium (Na) on prenatally programmed hypertension. Hypertension was induced in rat offspring by a maternal low protein (LP) diet. Control and LP offspring were randomized to a high (HS), standard (SS), or low (LS) Na diet after weaning. On the SS diet, the LP pups developed hypertension by 6 weeks of age. The development of hypertension was prevented by the LS diet and exacerbated by the HS diet. Kidney nitrotyrosine content, a measure of oxidative stress, was reduced by the LS diet compared with the HS diet. The modified diets had no effect on control pups. A group of animals on the SS diet was followed up to 51 weeks of age after an early life 3-week exposure to the HS or LS diet. This brief early exposure of LP animals to the LS diet prevented the later development of hypertension and ameliorated the nephrosclerosis observed after early exposure to the HS diet. The LP offspring with early exposure to LS diet had lost their salt-sensitivity when challenged with the HS diet at the age of 43-49 weeks. No effect of early life dietary Na was observed in control animals. These results show that hypertension in this model is salt sensitive and may, in part, be mediated by salt-induced renal oxidative stress and that there may exist a developmental window which allows postnatal "reprogramming" of the hypertension.

Angiotensin II maintains cerebral vascular relaxation via EGF receptor transactivation and ERK1/2

This study identified, on the integrative level, two components of the ANG II signaling pathway that lay downstream from the ANG II type 1 (AT(1)) receptor and are critically involved in maintaining vascular relaxation in cerebral resistance arteries. In these experiments, the relaxation of isolated middle cerebral arteries (MCA) in response to ACh (10(-9)-10(-5) M), iloprost (10(-16)-10(-11) g/ml), and reduced PO(2) was lost and the ratio of phospho-ERK/ERK1/2 was significantly reduced in aortas of male Sprague-Dawley rats fed a high-salt (HS; 4% NaCl) diet to suppress plasma ANG II levels. In salt-fed rats, relaxation of MCA in response to these vasodilator stimuli was restored by chronic (3 days) intravenous infusion of either ANG II (5 ngxkg(-1)xmin(-1)) or epidermal growth factor (EGF; 2 microg/h). The protective effect of ANG II infusion to restore vascular relaxation was eliminated by coinfusion of either the EGF receptor kinase inhibitor AG-1478 (20 microg/h), the ERK1/2 inhibitor PD-98059 (10 microg/h), or the protein synthesis inhibitor cycloheximide (5 microg/h). In rats fed a low-salt (0.4% NaCl) diet, MCA relaxation in response to ACh, reduced PO(2), and iloprost was eliminated by intravenous infusion of AG-1478, PD-98059, or cycloheximide. In ANG II-infused rats fed HS diet, and in rats fed LS diet, vasodilator responses to reduced PO(2) and iloprost were unaffected by the p38 MAP kinase inhibitor SB-203580 and the phosphatidylinositol 3-kinase inhibitor wortmannin. These findings indicate that maintenance of normal vascular relaxation mechanisms by ANG II in rat MCA requires activation of the EGF receptor kinase and ERK1/2.

Angiotensin II and Oxidative Stress Increases JNK and IRS‐1ser307 Activity and Inhibits Insulin Signal Transduction in Rats with Insulin Resistance Induced by Low Salt Intake

Low salt diet (LS) increases angiotensin II (AII) and oxidative stress (TBARS), which may enhance JNK and IRS1ser307 phosphorylation (PP) decreasing insulin signaling (INS‐S).ObjectiveTo evaluate if losartan (LOS) improves INS resistance by decreasing JNK and IRS1ser307 PP.MethodsTwelve‐week‐old rats were fed a LS (0.15% NaCl) or normal salt diet (NS: 1.3%) since weaning. Groups: LS+LOS (30mg/kg/day), LS+vehicle, NS+LOS and NS+vehicle. Blood pressure (BP), blood glucose (G), plasma INS and TBARS, HOMA, and tissue AII were evaluated. The INS‐S and inflammatory signaling (IS) were determined by Western blot and immunoprecipitation. AT1 receptor gene expression was determined by RT‐PCR.Results (mean±SEM, P&lt;0.05, n=8)BP and AT1 did not differ among groups. INS, HOMA and TBARS were higher in LS (INS: 712±60) than in NS (257±40 pmol/l). G and TBARS were lower and INS was higher in LS+LOS (TBARS: 4±0.5) than in LS (6±1 nmol/ml). Liver AII was higher in LS than in NS. INS receptor and Akt PP were higher and JNK (120±3) and IRS1ser307 PP (100±6 AU) were lower in the liver of LS+LOS compared to LS. IkBα PP was lower in LS (20±2) than in NS (100±4 AU) suggesting an activation of IS in LS. Similar results were observed in the muscle.ConclusionsINS resistance induced by LS is associated with alterations in regulation of INS‐S, activated IS, and higher tissue AII and TBARS. In LS, LOS improves INS‐S by decreasing JNK and IRS1ser307 PP with no change in BP.FAPESP