Results show that among CONT, fasting levels of proline, ornithine, lysine, alanine and threonine trended upward. In contrast, the nutrient bar INT significantly altered amino acid metabolism, such that concentrations of several gluconeogenic, sulfur redox and urea cycle intermediates decreased, notably fasting levels of serine, proline, aspartate, cystathionine, alanine, glutamine methionine, and citrulline. A small but significant increase in histidine concentration associated with favorable nitrogen balance, improved bioavailability of arginine, and an increase in the Fischer ratio the molar ratio of BCAA and aromatic amino acids , indicative of favorable vascular and liver functions respectively, were observed in the INT group only.Improved nutrition represents a lifestyle modification target to lower the growing global cardiometabolic disease burden, especially among youth and their families struggling with socioeconomic disparity and the many associated barriers to a healthy diet. Even without economic barriers, lifestyle adherence is challenging. Furthermore, strict dietary counseling and adherence for the management of hyperlipidemia has been associated with increased anxiety and depression in youth. The nutrient bar strategy utilized in this study was developed to enable research on the impact of nutrition on CMR, by facilitating daily delivery of nutrients from foods missing from a standard American diet, essential fibers, minerals, vitamins, strawberry gutter system omega-3 fatty acids, and polyphenols, in a readily consumed, low-calorie, palatable nutrient bar.
Results herein show that a 2 month nutrient bar intervention together with increased physical activity improved systolic blood pressure in Teens only without other significant changes observed in traditional biomarkers, but was associated with metabolomicshifts in the full INT PAC and Teen group that correlate with favorable changes in cardiometabolic disease risk biomarkers. Baseline metabolomic analysis identified ceramides as unique biomarkers of atherogenic dyslipidemia in both Teens and adults at heightened CMR . Although the metabolomic panel included BCAA and AAA previously implicated in CMR, only PC1, composed primarily of ceramide metabolites, associated with baseline TG levels and TG-rich VLDL subspecies, suggesting that ceramides may contribute to TG homeostasis. Fatty-acyl CoA is a substrate for both ceramide and TG biosynthesis and thus, excess ceramide and TG synthesis may occur under conditions of abundant fatty acid availability through both high fat diets or high refined carbohydrate diets and de novo lipogenesis. In contrast to TGs that serve a passive fatty acid storage function, ceramides have unique signaling roles in mediating insulin resistance and mitochondrial bioenergetics. Ceramides may therefore act as cellular energy sensors to regulate tissue fuel uptake and storage via insulin signaling. In obesity with chronic inflammation, excess ceramide, particularly C16:0 Cer, is associated with mitochondrial dysfunction, impaired sphingolipid cell signaling and insulin resistance. Decreasing tissue ceramide through pharmacological or genetic manipulations can reverse or attenuate insulin resistance, providing support for a direct role of ceramides as critical part of fuel sensing circuitry in the body. Ceramides may represent a sensitive biomarker of metabolic dysregulation when energetic supply chronically exceeds storage capacity. The current findings suggest that this tipping point may be influenced not only by absolute caloric intake but also by the dietary nutrient value. Intensive lifestyle modification has been shown to improve CMR biomarkers in the obese.
In this trial, despite excellent participation in weekly exercise sessions, self-report of increased exercise between sessions in all subjects and both subjective and objective evidenceof compliance with nutrient bar intake in the INT group, there were minimal changes in traditional parameters of dyslipidemia, inflammation, and insulin resistance . Changes in the plasma metabolome were however able to signify highly significant differences between CONT and INT . The nutrient supplement INT modified effects of physical activity on plasma ceramide metabolism. Contrary to the expectation that physical activity would lead to lower ceramide formation by improving lipid oxidation capacity, significant increases in C14:0, C16:0, C20:0 and C22:0 and overall increasing trends with all ceramide species were observed in CONT participants. A study by Bergman and co-workers similarly found that acute exercise transiently increases serum ceramides in obese untrained subjects. Significant increases in sphinganine in CONT may reflect increasesd de novo synthesis driven in part by a higher rate of lipolysis following exercise. Exercise increases lipolysis but given obesity related decreases in muscle fatty acid oxidation, a rise in free palmitate following exercise may, in the short-term, favor increased ceramide synthesis. The observed rise in ceramides in CONT may therefore reflect transient increases in fatty acid mobilization. Further training which would result in an increased FAO capacity would be expected to normalize these parameters. Effects of exercise on ceramides were largely blunted when subjects consumed daily nutrient bars, suggesting enhanced muscle FAO by the provision of nutrient substrates missing from a typical American diet. The nutrient bar is enriched in polyphenols and omega-3 fatty acids, known to increase muscle peroxisome proliferator activated receptor coactivator 1 alpha activity and mitochondrial biogenesis, processes which would be expected to improve amino acid and fatty acid catabolism.
In support of this mechanism, we also observed significant lowering of non-essential amino acids , serine, proline, aspartate, glutamine, and alanine . Lower NEAA’s may reflect improved substrate utilization and clearance of biomarkers associated with what has been termed the “metabolic gridlock” associated with obesity. Furthermore, because serine is a substrate for synthesis of sphingolipids, its decrease may also have contributed to the lack of a rise in ceramide detected among INT participants. Alternatively, serine mobilization from other sites into plasma may be lessened when ceramide synthesis is not triggered. Although clinical changes in TG, FBG, insulin and lipoproteins were not significant, we observed several significant correlations between changes in the plasma lipidome with changes in TG and physiologically interrelated pro-atherogenic small dense LDL particles . Plasma contains upwards of 200 distinct SPL species distributed across HDL, apoB-containing lipoproteins and albumin, but differential association with the most atherogenic lipoprotein subspecies has not been described. As with baseline correlations, the change to change correlations following the nutrient bar intervention were highly granular with specific ceramide species showing associations with changes in TG. Ceramides with C14:0, C16:0, C24:1 and C26:1 showed positive associations with changes in TG. C14:0 ceramide has been previously been implicated in nonalcoholic fatty liver disease in adolescent children. C16:0 ceramide inhibits mitochondrial FAO, oxidant production, and impairs Complex IV activity. C24:1 ceramide significantly decreased in the INT group. Inhibition of synthesis of very long chain ceramides by deletion of fatty acid elongase 6 protects against hepatic steatosis in high fat diet induced obesity. Very small LDL particles showed a highly specific correlation with C18:1 ceramide . Deletion of ceramide synthase 1 responsible for C18 ceramide synthesis has also been shown to increase mitochondrial FAO and protect against high fat diet induced NAFLD. Interaction between specific ceramide species and small LDL may be involved in the progression from NAFLD to nonalcoholic steatohepatitis . NASH is not only characterized by high concentrations of small LDL and a lowmean LDL diameter but post hoc analyses from the Pioglitazone vs Vitamin E vs Placebo for the Treatment of Nondiabetic Patients with NASH trial demonstrated that resolution of NASH was associated with an increase in the average LDL particle diameter and decreased small and very small LDL particles. These lipoprotein subspecies parameters worsened in the participants who did not resolve their NASH. Larger LDL particles have improved affinity for LDL receptors and decreased retention in the subendothelial glycoprotein matrix, grow strawberry in containers so would be expected to lead to decreased LDL and reduced atherogenesis. These findings collectively suggest that early changes in ceramide species among INT participants signal favorable shifts in lipid metabolism that may precede improvement in traditional clinical biomarkers. In both CONT and INT participants, sphingosine-1-phosphate levels increased, but with S1P levels rising higher among INT participants. A previous study showed that exercise increases plasma S1P levels and decreases erythrocyte ceramide levels and this effect was more evident in untrained subjects. S1P is degraded by sphingosine-1-phosphate lyase . SPL dependent degradation of S1P is terminal step in sphinogolipid degradation critical for regulating the pool size of all sphingolipids. In blood, ~50–70% of S1P is bound to apolipoprotein M within the HDL complex and is important for its reverse cholesterol transport. Decreased S1P bound to HDL is associated with higher risk for coronary artery disease. Although there were no absolute changes in HDL subspecies, quantification of HDL-bound S1P may provide further insights as to whether increases in plasma S1P in our study participants contribute to improved antiatherogenic HDL functions. Correlation analysis with changes in amino acids showed highly specific associations between changes in citrulline, arginine bio-availability and cystathionine and changes in measures of insulin resistance. Obesity and insulin resistance are associated with increased CRP, increased citrulline and decreased arginine bio-availability in Teens.
This may reflect increased nitric oxide demand associated with the inflammatory metabolic stress of insulin resistance. Interestingly in this cohort with high baseline CRP, only change in ornithine correlated with change in CRP. Arginase expression and increased Ornithine/Arginine ratio has been shown to associate with vasculopathies in humans and in animals fed a high fat, high sugar Western diet. The favorable pairwise difference in arginine bioavailabity in the INT vs CONT groups may have contributed to subgroup specific improvement in systolic BP among INT Teens. Insulin sensitizing treatment with metformin plus pioglitazone for 3 months also improves arginine bioavailability in obese subjects with impaired glucose tolerance. Change in Cystathionine was positively associated with change in glucose, insulin, and HOMA. Cysth is part of an alternate redox pathway to NOS, synthesized by cystathionine-beta-synthase via the transsulfuration pathway. The expression of CBS in the liver observed in mice on a high fat diet has been proposed to be a defense mechanism against increased oxidative stress. Decreased Cysth among INT participants may therefore reflect a lower burden of oxidative stress that could lead to improved insulin regulation. Our study is limited by its relatively short observation period and the small sample size that precluded detailed analysis of data stratified by age. However, covariate analysis showed no significant effects of age with either the baseline correlations or with metabolic responses to the bar suggesting the metabolic impact of the nutrient bar intervention is conserved between the age groupings considered. The sample size was insufficient to consider hereditary factors. Furthermore, race and ethnicity, BMI and CRP differed by chance between INT and CONT groups although both groups exhibited comparable and considerable baseline CMR risk. The fact that lower total plasma Cers and specifically lower Cer 24:1 are described in persons of African American ancestry who by chance represented a larger proportion of the CONT group, cannot explain either the differential rise in total Cers among controls, nor the significant lowering of the Cer24:1 species in the INT group only. A larger study will be required to stratify analyses by race, ethnicity and other hereditary factors that may affect sphingolipid metabolism to determine if this may have influenced results. In summary, our study has uncovered several granular and unique associations between plasma ceramides and amino acid metabolites with clinical parameters of dyslipidemia, inflammation and insulin resistance. These findings suggest that remediation of essential nutrients typically lacking in western style diets should be considered an essential component of preventive interventions directed towards the alleviation of CMR. In this regard, the metabolomic changes as monitored herein are more sensitive indicators of favorable, but subtle shifts in metabolism. A lack of response in traditional clinical CMR biomarkers does not necessarily reflect a lack of compliance with behavioral interventions. The positive associations between changes in established traditional risk factors and changes in metabolomic biomarkers support the hypothesis that metabolomic biomarkers are sensitive prognostic indicators at the leading edge of response to lifestyle therapy, possibly mediated at the level of mitochondrial function. Further larger and longer studies are needed to validate whether the changes observed in these analyses will predict and precede future favorable changes in traditional CMR biomarkers and the benefits to weight regulation that can be anticipated with improved metabolism.Vineyard soil microorganisms are affected by wine growing region, climate and topography, as mediated in part by their suite of impacts on soil properties like pH and soil organic matter pools . These same soil properties are directly influenced by vineyard management practices. Soil microorganisms also influence their local environment through pathogen suppression; decomposition processes that affect soil organic matter mineralization, contribution and preservation of SOM and aggregate stability; and availability of nitrogen and other mineral nutrients .