Carmelo Nieves

Arginine and immunity: a unique perspective

Arginine functions in the body as a free amino acid, a component of most proteins, and the substrate for several non-protein, nitrogen-containing compounds, many of which function in immunity. Although arginine is synthesized in the body, it is not made in sufficient quantities to support growth or meet metabolic requirements during periods of stress. Based on the biochemical and physiological role of arginine in maintaining health and immunity, arginine is being added at pharmacologic concentrations to enteral formulas to boost immune function. Unfortunately, animal and human studies that investigate enteral arginine supplementation as the single variable do not show clear immunologic benefit. The inconsistent effects of arginine supplementation on immune function are due to numerous factors, such as the amount and timing of arginine supplementation, the animal species or strain of species, and the experimental model. Systematic study is required to determine whether a basal dietary intake of arginine is required to maintain immune function during health and how much arginine is required to meet metabolic requirements during periods of growth or stress.

Immune Function Is Impaired With a Mini Nutritional Assessment Score Indicative of Malnutrition in Nursing Home Elders With Pressure Ulcers

BACKGROUND Malnutrition is prevalent in elders with pressure ulcers and is associated with increased morbidity and mortality. This study compared nutritional status, assessed by the Mini Nutrition Assessment (MNA), to immune function in nursing home elders with pressure ulcers. METHODS Nutritional status was assessed in nursing home residents (>65 years) with a stage II or more severe pressure ulcer. Subjects were classified as well nourished, at risk of malnutrition, or malnourished according to MNA score. Blood was drawn to assess whole blood mitogen-induced lymphocyte proliferation and neutrophil respiratory burst. Delayed-type hypersensitivity to 3 antigens was measured. MNA status was compared with immune parameters using the Kruskall-Wallis test. RESULTS Of the 24 subjects (23 men, 1 woman) who completed the study protocol, only 4 (17%) were classified as well nourished, whereas 7 (29%) were at risk and 13 (54%) were malnourished according to MNA score. Whole blood lymphocyte proliferation was significantly lower in the malnourished vs at risk subjects with both pokeweed (median [25th, 75th percentile], 0.6 [0.3, 0.9] vs 1.8 [1.2, 2.1] disintegrations per minute [dpm]/cell, p < .05); and concanavalin A (1.7 [0.9, 2.0] vs 2.8 [2.6, 3.9] dpm/cell, p < .05) mitogens. Neutrophil respiratory burst normalized to a young control was significantly lower in malnourished subjects vs well-nourished subjects (0.8 [0.5, 0.9] vs 1.4 [1.0, 1.7], p < .05). Total induration to 3 skin-test antigens was 13.4 +/- 4.6, 3.5 +/- 2.6, and 3.8 +/- 1.8 (mean +/- SEM) for well-nourished, at risk, and malnourished, respectively (p = .059). CONCLUSIONS Immune function is impaired with an MNA score indicative of malnutrition in nursing home elders with pressure ulcers.

Regular Consumption of Concord Grape Juice Benefits Human Immunity

γδ T cells are important immune surveillance cells residing in epithelial layers lining the intestine, lung, and reproductive tract. The main objective of this study was to test the hypothesis that consumption of dietary compounds from grapes would modify γδ T-cell function. Other factors related to immune function after grape juice consumption were also tested. A randomized, double-blind, placebo-controlled, parallel intervention was conducted: 100% grape juice made from Concord grapes or a placebo beverage was consumed by 85 individuals daily for 9 weeks. Subjects were asked not to consume other red, blue, and purple fruits during the study. Blood samples, taken at the beginning and the end, were analyzed for γδ T-cell numbers and proliferation, vitamin C, antioxidant capacity, and the ability to protect DNA from strand breaks. Those consuming the grape juice had significantly greater numbers of circulating γδ T cells and higher serum vitamin C levels compared to the placebo by two-way repeated-measure analysis of variance (P < .05). Individuals consuming the placebo had lower serum antioxidant activity, less γδ T-cell proliferation, and increased DNA strand breaks when challenged with H(2)O(2). Analysis of the data by structural equation modeling confirmed that 61% of the variance in biological functions at 9 weeks was due to grape juice consumption. Based on conventional statistical analyses, as well as on sophisticated modeling techniques, regular consumption of purple grape juice in the absence of other red, blue, or purple fruits benefited immunity in healthy, middle-aged human subjects.

Fecal Lactic Acid Bacteria Increased in Adolescents Randomized to Whole-Grain but Not Refined-Grain Foods, whereas Inflammatory Cytokine Production Decreased Equally with Both Interventions

The intake of whole-grain (WG) foods by adolescents is reported to be approximately one-third the recommended intake of 48 g/d. This 6-wk randomized interventional study determined the effect of replacing grains within the diet with refined-grain (RG; n = 42) or WG (n = 41) foods/d on gastrointestinal and immune health in adolescents (aged 12.7 ± 0.1 y). A variety of grain-based foods were delivered weekly to participants and their families. Participants were encouraged to eat 3 different kinds of study foods (e.g., bread, cereals, snacks)/d with goals of 0 g/d (RG) and 80 g/d (WG). Stool samples were obtained during the prebaseline and final weeks to measure bifidobacteria and lactic acid bacteria (LAB) using qPCR. Stool frequency was recorded daily. Blood was drawn at baseline and at final visits for immune markers. Across groups, total-grain intake increased by one serving. The intake of WG was similar at baseline (18 ± 3 g) between groups but increased to 60 ± 5 g in the WG group and decreased to 4 ± 1 g in the RG group. Fecal bifidobacteria increased from baseline with both interventions, but LAB increased (P < 0.05) from baseline [2.4 ± 0.2 log(10) genome equivalents (eq)] to wk 6 (3.0 ± 0.2 log(10) genome eq) in the WG group but not in the RG group (baseline: 2.9 ± 0.2 log(10) genome eq; wk 6: 3.0 ± 0.1 log(10) genome eq). There was no difference in stool frequency, serum antioxidant potential, or in vitro LPS-stimulated mononuclear cell production of inflammatory cytokines between groups. However, across both groups the number of daily stools tended to increase (P = 0.08) by 0.0034 stools/g WG or by 0.2 stools with 60 g WG, mean antioxidant potential increased by 58%, and mean production of TNF-α, IL-1β, and IL-6 decreased by 24, 22, and 42%, respectively, between baseline and wk 6. Overall, incorporating either WG or RG foods increased serum antioxidant concentrations and decreased inflammatory cytokine production; however, WG study foods had more of an effect on aspects of gastrointestinal health.

Consuming Lentinula edodes (Shiitake) Mushrooms Daily Improves Human Immunity: A Randomized Dietary Intervention in Healthy Young Adults

Background: Mushrooms are widely cited for their medicinal qualities, yet very few human intervention studies have been done using contemporary guidelines. Objective: The aim of this study was to determine whether consumption of whole, dried Lentinula edodes (shiitake) mushrooms could improve human immune function. Primary objectives were to ascertain whether L. edodes consumption would improve γδ-T cell proliferation and activation responses, quantify a dose response, and elicit cytokine secretion patterns. Secondary objectives included determining changes in natural killer T (NK-T) cell proliferation and activation, secretory immunoglobulin A (sIgA) in saliva, and C-reactive protein (CRP) in serum. Design: Fifty-two healthy males and females, aged 21–41 years, participated in a 4-week parallel group study, consuming either 5 or 10 g of mushrooms daily. Each subject had blood drawn before and after 4 weeks of daily L. edodes consumption. Saliva and serum were also collected. Peripheral blood mononuclear cells were cultured in autologous serum for 24 hours or 6 days, stained, and examined by flow cytometry. Results: Eating L. edodes for 4 weeks resulted in increased ex vivo proliferation of γδ-T (60% more, p < 0.0001) and NK-T (2-fold more, p < 0.0001) cells. Both cell types also demonstrated a greater ability to express activation receptors, suggesting that consuming mushrooms improved cell effector function. The increase in sIgA implied improved gut immunity. The reduction in CRP suggested lower inflammation. The pattern of cytokines secreted before and after mushroom consumption was significantly different; consumption resulted in increased interleukin (IL)-4, IL-10, tumor necrosis factor (TNF)-α, and IL-1α levels, a decreased macrophage inflammatory protein-1α/chemokine C-C ligand 3 (MIP-1α/CCL3) level, and no change to IL-6, IL-1β, MIP-1β, IL-17 and interferon (IFN)-γ levels. Conclusions: Regular L. edodes consumption resulted in improved immunity, as seen by improved cell proliferation and activation and increased sIgA production. The changes observed in cytokine and serum CRP levels suggest that these improvements occurred under conditions that were less inflammatory than those that existed before consumption.

Diet quality improves for parents and children when almonds are incorporated into their daily diet: a randomized, crossover study

The health benefits of nuts may, in part, be due to the fiber that provides substrate for the maintenance of a healthy and diverse microbiota. We hypothesized that consuming almonds would benefit immune status through improving diet quality and modulation of microbiota composition in parents and their children, while improving gastrointestinal function. In a crossover trial, 29 parents (35 ± 0.6 years) and their children (n = 29; 4 ± 0.2 years; pairs) consumed 1.5 and 0.5 oz, respectively, of almonds and/or almond butter or control (no almonds) for 3 weeks followed by 4-week washouts. Parents completed daily questionnaires of stool frequency and compliance with nut intake. The Gastrointestinal Symptom Response Scale was administered weekly. Participants provided stools for microbiota analysis and saliva for secretory immunoglobulin A. Serum antioxidant/proinflammatory balance was determined in parents. From weekly dietary recalls (Automated Self-Administered 24-Hour Dietary Recall), nutrient and energy intake were assessed and Healthy Eating Index-2010 scores were calculated. Consuming almonds increased total Healthy Eating Index score from 53.7 ± 1.8 to 61.4 ± 1.4 (parents) and 53.7 ± 2.6 to 61.4 ± 2.2 (children; P < .001). Minimal changes in gastrointestinal symptoms and no change in stool frequency were noted with the almond intervention. Microbiota was stable at the phylum and family level, but genus-level changes occurred with nut intake, especially in children. No differences were observed for immune markers. Although higher intakes of almonds or longer interventions may be needed to demonstrate effects on immune status, a moderate intake of almonds improves diet quality in adults and their young children and modulates microbiota composition.

Lactobacillus gasseri KS-13, Bifidobacterium bifidum G9-1, and Bifidobacterium longum MM-2 Ingestion Induces a Less Inflammatory Cytokine Profile and a Potentially Beneficial Shift in Gut Microbiota in Older Adults: A Randomized, Double-Blind, Placebo-Controlled, Crossover Study

Objective: This study determined whether older adults who consumed a probiotic mixture would have a greater proportion of circulating CD4+ lymphocytes, altered cytokine production, and a shift in intestinal microbiota toward a healthier microbial community. Methods: Participants (70 ± 1 years [mean ± SEM]; n = 32) consumed a probiotic (Lactobacillus gasseri KS-13, Bifidobacterium bifidum G9-1, and Bifidobacterium longum MM2) or a placebo twice daily for 3 weeks with a 5-week washout period between intervention periods. Blood and stools were collected before and after each intervention. The percentage of circulating CD4+ lymphocytes and ex vivo mitogen-stimulated cell cytokine production were measured. In stools, specific bacterial targets were quantified via quantitative polymerase chain reaction (qPCR) and community composition was determined via pyrosequencing. Results: During the first period of the crossover the percentage of CD4+ cells decreased with the placebo (48% ± 3% to 31% ± 3%, p < 0.01) but did not change with the probiotic (44% ± 3% to 42% ± 3%) and log-transformed concentrations of interleukin-10 increased with the probiotic (1.7 ± 0.2 to 3.4 ± 0.2, p < 0.0001) but not the placebo (1.7 ± 0.2 to 2.1 ± 0.2). With the probiotic versus the placebo a higher percentage of participants had an increase in fecal bifidobacteria (48% versus 30%, p < 0.05) and lactic acid bacteria (55% versus 43%, p < 0.05) and a decrease in Escherichia coli (52% versus 27%, p < 0.05). Several bacterial groups matching Faeacalibactierium prausnitzii were more prevalent in stool samples with the probiotic versus placebo. Conclusions: The probiotic maintained CD4+ lymphocytes and produced a less inflammatory cytokine profile possibly due to the changes in the microbial communities, which more closely resembled those reported in healthy younger populations.

Bifidobacterium bifidum R0071 decreases stress-associated diarrhoea-related symptoms and self-reported stress: a secondary analysis of a randomised trial

Psychological stress is associated with gastrointestinal (GI) distress. This secondary analysis from a randomised, double-blind, placebo-controlled study examined whether three different probiotics could normalise self-reported stress-associated GI discomfort and reduce overall self-reported stress. Undergraduate students (n=581) received Lactobacillus helveticus R0052, Bifidobacterium longum ssp. infantis R0033, Bifidobacterium bifidum R0071, or placebo. Participants self-reported 2 outcomes for a 6-week period, which included final academic exams: daily level of stress (0=no stress to 10=extremely stressed) and weekly three diarrhoea-related symptoms (DS, 1=no discomfort to 7=severe discomfort) using the GI Symptom Rating Scale. Self-reported stress was positively related to DS (P=0.0068). Mean DS scores were lower with B. bifidum versus placebo at week 2 at the average level of stress and the average body mass index (BMI). DS scores were lower with B. bifidum at week 5 versus week 0 and 1 and with B. infantis R0033 at week 6 versus week 0. DS scores were higher when antibiotics were used in the prior week with placebo (P=0.0092). DS were not different with or without antibiotic use with the probiotics. Only B. bifidum had an effect on self-reported stress scores (P=0.0086). The self-reported stress score was also dependent on hours of sleep per day where it decreased by 0.13 for each additional hour of sleep. During a stressful period, B. bifidum R0071 decreases DS and self-reported stress scores. This trial was registered at clinicaltrials.gov as NCT01709825.

Probiotics (Lactobacillus gasseri KS-13, Bifidobacterium bifidum G9-1, and Bifidobacterium longum MM-2) improve rhinoconjunctivitis-specific quality of life in individuals with seasonal allergies: a double-blind, placebo-controlled, randomized trial

Background: Rhinoconjunctivitis-specific quality of life is often reduced during seasonal allergies. The Mini Rhinoconjunctivitis Quality of Life Questionnaire (MRQLQ) is a validated tool used to measure quality of life in people experiencing allergies (0 = not troubled to 6 = extremely troubled). Probiotics may improve quality of life during allergy season by increasing the percentage of regulatory T cells (Tregs) and inducing tolerance.Objective: The objective of this study was to determine whether consuming Lactobacillus gasseri KS-13, Bifidobacterium bifidum G9-1, and B. longum MM-2 compared with placebo would result in beneficial effects on MRQLQ scores throughout allergy season in individuals who typically experience seasonal allergies. Secondary outcomes included changes in immune markers as part of a potential mechanism for changes in MRQLQ scores.Design: In this double-blind, placebo-controlled, parallel, randomized clinical trial, 173 participants (mean ± SEM: age 27 ± 1 y) who self-identified as having seasonal allergies received either a probiotic (2 capsules/d, 1.5 billion colony-forming units/capsule) or placebo during spring allergy season for 8 wk. MRQLQ scores were collected weekly throughout the study. Fasting blood samples were taken from a subgroup (placebo, n = 37; probiotic, n = 35) at baseline and week 6 (predicted peak of pollen) to determine serum immunoglobulin (Ig) E concentrations and Treg percentages.Results: The probiotic group reported an improvement in the MRQLQ global score from baseline to pollen peak (-0.68 ± 0.13) when compared with the placebo group (-0.19 ± 0.14; P = 0.0092). Both serum total IgE and the percentage of Tregs increased from baseline to week 6, but changes were not different between groups.Conclusions: This combination probiotic improved rhinoconjunctivitis-specific quality of life during allergy season for healthy individuals with self-reported seasonal allergies; however, the associated mechanism is still unclear. This trial was registered at clinicaltrials.gov as NCT02349711.

Pharmacologic levels of dietary arginine in CB6F1 mice increase serum ammonia in the healthy state and serum nitrite in endotoxemia

BACKGROUND Therapeutic or pharmacologic doses of arginine are used to enhance blood flow and immune function despite the lack of dose-response studies and the potential for adverse effects. This study determined the optimal level of oral arginine supplementation required to elevate serum arginine concentrations yet limit adverse effects in healthy and endotoxemic mice. METHODS Male CB6F1 mice were fed one of the following diets: The standard AIN93G (3 g arginine/100 g of protein) or this diet modified to provide 10 g, 20 g, or 30 g arginine/100 g of protein. On day 14, mice were injected with lipopolysaccharide (endotoxemic) or saline (healthy) and 4 hours later were exsanguinated. RESULTS Weight gain was reduced 50% in the group fed the 30 g arginine vs standard diet. Serum arginine, ornithine, citrulline, histidine, lysine, serine, threonine, tyrosine, and phenylalanine were greater and glutamate levels were lower in healthy supplemented mice; lipopolysaccharide treatment negated these changes. Serum ammonia concentration was 52% greater in healthy mice fed the 30 g arginine vs standard diet. Serum nitrite and urea were unaffected by supplementation in healthy mice. Serum nitrite was 37% greater in endotoxemic mice fed 30 g vs 10 g arginine, and serum urea was 27% greater in mice fed 20 g or 30 g vs 10 g arginine. CONCLUSIONS Changes in serum arginine or its metabolites were observed with all of the modified diets; however, a 30-g arginine diet was associated with an initial impairment of growth and potential adverse effects.