Jonathan R. Powell

Influence of dietary supplementation with long-chain n−3 or n−6 polyunsaturated fatty acids on blood inflammatory cell populations and functions and on plasma soluble adhesion molecules in healthy adults

Greatly increasing the amounts of flaxseed oil [rich in α-linolenic acid (ALNA)] or fish oil (FO); [rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] in the diet can decrease inflammatory cell functions and so might impair host defense. The objective of this study was to determine the effect of dietary supplementation with moderate levels of ALNA, γ-linolenic acid (GLA), arachidonic acid (ARA), DHA, or FO on inflammatory cell numbers and functions and on circulating levels of soluble adhesion molecules. Healthy subjects aged 55 to 75 yr consumed nine capsules per day for 12 wk. The capsules contained placebo oil (an 80∶20 mix of palm and sunflowerseed oils) or blends of placebo oil with oils rich in ALNA, GLA, ARA, or DHA or FO. Subjects in these groups consumed 2 g ALNA; approximately 700 mg GLA, ARA, or DHA; or 1 g EPA plus DHA (720 mg EPA+280 mg DHA) daily from the capsules. Total fat intake from the capsules was 4 g per day. None of the treatments affected inflammatory cell numbers in the bloodstream; neutrophil and monocyte phagocytosis or respiratory burst in response to E. coli; production of tumor necrosis factor-α, interleukin-1β, and interleukin-6 in response to bacterial lipopolysaccharide; or plasma concentrations of soluble intercellular adhesion molecule-1. In contrast, the ALNA and FO treatments decreased the plasma concentrations of soluble vascular cell adhesion molecule-1 (16 and 28% decrease, respectively) and soluble E-selectin (23 and 17% decrease, respectively). It is concluded that, in contrast to previous reports using higher amounts of these fatty acids, a moderate increase in consumption of long-chain n−6 or n−3 polyunsaturated fatty acids does not significantly affect inflammatory cell numbers or neutrophil and monocyte responses in humans and so would not be expected to cause immune impairment. Furthermore, we conclude that moderate levels of ALNA and FO, which could be incorporated into the diet, can decrease some markers of endothelial activation and that this mechanism of action may contribute to the reported health benefits of n−3 fatty acids.

Antioxidants may contribute in the fight against ageing: an in vitro model.

Elderly humans have altered cellular redox levels and dysregulated immune responses, both of which are key events underlying the progression of chronic degenerative diseases of ageing, such as atherosclerosis and Alzeimers disease. Poorly maintained cellular redox levels lead to elevated activation of nuclear transcription factors such as NFkB and AP-1. These factors are co-ordinately responsible for a huge range of extracellular signalling molecules responsible for inflammation, tissue remodelling, oncogenesis and apoptosis, progessess that orchestrate many of the degenerative processess associated with ageing. It is now clear that levels of endogenous anti-oxidants such as GSH decrease with age. This study aimed to investigate the potential of exogenous anti-oxidants to influence inflammatory responses and the ageing process itself. We investigated the potential of the dietary antioxidant, quercetin, to reverse the age related influences of GSH depletion and oxidative stress using in vitro human umbilical vein endothelial cells (HUVEC) and human skin fibroblast (HSF) cell models. Oxidative stress-induced inflammatory responses were investigated in a GSH depletion and a Phorbol 12-myristate 13-acetate (PMA)-induced stress model. As measured with a sensitive HPLC fluorescence method, GSH in HUVEC was depleted by the addition of L-buthionine-[S,R]-sulfoxiniine (BSO), a gamma-glutamylcysteine synthetase inhibitor, to the culture medium at a concentration of 0.25 mM. Time course studies revealed that the GSH half-life was 4.6 h in HUVEC. GSH depletion by BSO for 24 h led to a slight increase in intracellular adhesion molecule - 1 (ICAM1) expression and prostaglandin E2 (PGE2) secretion in both types of cells. However, GSH depletion markedly enhanced PMA-induced ICAM and PGE2 production in HUVEC. Responses were progressively elevated following prolonged BSO treatment. Inhibition studies showed that 1-(5-Isoquinolinylsulfonyl)-2-methylpiperazine (H7), a protein kinase C (PKC) inhibitor, not only abolished most of PMA-induced ICAM-1 expression and PGE2, production, but also eliminated GSH depletion-enhanced PMA stimulation. This enhancement was also inhibited by supplementation with quercetin. The results clearly demonstrate that GSH depletion increased the susceptibility of vascular endothelial cells and fibroblasts to oxidative stress associated inflammatory stimuli. This increased in vitro susceptibility may be extrapolated to the in vivo situation of ageing, providing a useful model to study the influence of micronutrients on the ageing process. In conclusion, these data suggest that dietary antioxidants could play a significant role in the reduction of inflammatory responses.

Lack of effect of meal fatty acid composition on postprandial lipid, glucose and insulin responses in men and women aged 50-65 years consuming their habitual diets

The aim of the study was to determine the effect of consuming meals with different fatty acid compositions on the postprandial changes over 6 h in plasma triacylglycerol, NEFA, total cholesterol, glucose and insulin concentrations in middle-aged men and women. Men (n 11; 58 (5) years) and women (n 11; 56 (4) years) consumed four test meals with a similar macronutrient energy content in random order: a reference meal based on the habitual pattern of fatty acid intake in the UK, a meal with an increased (155 %) linoleic acid (LA) to alpha-linolenic acid (alphaLNA) ratio (high LA:alphaLNA), a meal with increased (23 %) MUFA content (high MUFA) and a meal with increased (583 %) EPA and DHA content (high EPA+DHA). The high-LA:alphaLNA and high-EPA+DHA meals selectively increased the ratio of LA to alphaLNA (men 341 %; women 310 %) and the EPA+DHA (men 414 %; women 438 %) concentration in plasma triacylglycerol. The high-MUFA meal did not alter the change in MUFA content of the plasma. Plasma triacylglycerol, NEFA, glucose and insulin, but not total cholesterol, concentrations changed significantly after each meal. There was no significant effect of meal fatty acid composition or gender on maximum change in concentration, time to maximum concentration or area under the curve of any of the metabolites measured in the blood. These results suggest that differences in meal fatty acid composition exert little or no effect on postprandial changes in plasma lipids, glucose and insulin concentrations.

Evidence for Anti-Inflammatory Effects of Combined Administration of Vitamin E and C in Older Persons with Impaired Fasting Glucose: Impact on Insulin Action

Objective: Vitamin E and C given separately improve insulin sensitivity due to an inhibitory effect on oxidative stress and inflammation, however their combined effect on glucose control and inflammation is unknown. To investigate combined effect of Vitamin E and C in elderly with Impaired Fasting Glucose (IFG) on insulin action and substrate oxidation. Design: Controlled-trial administration of Vitamin E (1000 mg/day) and Vitamin C (1000 UI/day) for four weeks. Hyperinsulinemic euglycemic glucose clamp was performed before and following supplementation. Setting: Out-patient clinic. Participants: Thirteen older men with IFG. Main Outcome Parameters: Variations in whole body glucose disposal (WBGD), anti-oxidant, and inflammatory cytokines plasma levels. Results: An increase in plasma Vitamin E (8.3 + 0.8 vs. 64.9 + 2.1 μmol/l; p < 0.001] and C (35.9 + 5.4 vs. 79.4 + 7.4 μmol/l; p < 0.001) was found. Vitamin administration reduced insulin, glucose, lipid, TNF-α and [8-]isoprostane levels. Increase in plasma vitamin E levels correlated with decline in both plasma [8-]isoprostane levels (r = −0.58; p = 0.048) and TNF-α levels (r = − 0.62; p = 0.025), while no correlations were found for Vitamin C. Whole body glucose disposal (WBGD) (22.7 + 0.6 vs. 30.4 + 0.8 mmol × kg-1 × min-1; p = 0.001) and non-oxidative glucose metabolism rose after supplementation. Rise in plasma levels of Vitamin C and E correlated with WBGD. Multivariate linear regression models showed independent associations among the change in Vitamin E and the decline in TNF-α and [8-]isoprostane levels. Conclusions: Combined administration of Vitamin E and C lowered inflammation and improved insulin action through a rise in non-oxidative glucose metabolism.

Effect of consumption of soy isoflavones on behavioural, somatic and affective symptoms in women with premenstrual syndrome

Up to 80 % of the Western female population experience premenstrual syndrome (PMS). Long-term pharmacological therapy is unacceptable to most women, and is not warranted for moderate symptoms. Nutritional therapies are popular, but lack a clear evidence base. Anecdotal evidence suggests beneficial effects of soy isoflavones because of their influence on endogenous oestrogen and actions on specific tissues. The effect of isolated soya protein (ISP) containing 68 mg/d (aglycone equivalents) soy isoflavones (IF) on premenstrual symptom severity was studied in a seven-menstrual cycle, double-blind, placebo-controlled, crossover intervention study in twenty-three women with prospectively confirmed PMS aged 18-35 years and BMI 19-30 kg/m(2). ISP containing IF or milk protein placebo was consumed for two complete menstrual cycles. ISP containing IF (genistein, daidzein, equol) were measured in 24 h urine samples. After two cycles of ISP containing IF intervention, total symptoms (F(2,36) 8.20, P=0.000) and physical symptoms (F(2,36) 8.18, P=0.000) were significantly reduced compared with baseline after both active and placebo treatments, although differences between active and placebo treatment were non-significant. Specific premenstrual symptoms, headache (F(2,32) 4.10, P=0.026) and breast tenderness (F(2,32) 4.59, P=0.018), were reduced from baseline after soy IF, but not milk protein placebo. Cramps (F(2,32) 4.15, P=0.025) and swelling (F(2,32) 4.64, P=0.017) were significantly lower after active treatment compared with placebo. Concentrations of genistein and daidzein were increased following soy IF consumption, but equol production did not enhance symptom reduction. The present study showed that ISP containing IF may have potential to reduce specific premenstrual symptoms via non-classical actions.

Whole-genome microarray analysis identifies up-regulation of Nr4a nuclear receptors in muscle and liver from diet-restricted rats

One of the most conserved methods to significantly increase lifespan in animals is through dietary restriction (DR). The mechanisms by which DR increases survival are controversial but are thought to include improvements in mitochondrial function concomitant with reductions in reactive oxygen species production and alterations in the insulin signalling pathway, resulting in global metabolic adaptation. In order to identify novel genes that may be important for lifespan extension of Brown Norway rats, we compared gene expression profiles from skeletal muscle of 28-month-old animals fed ad libitum or DR diets using whole-genome arrays. Following DR, 426 transcripts were significantly down-regulated whilst only 52 were up-regulated. Included in the up-regulated transcripts were three functionally related previously unidentified DR-regulated genes: Nr4a1, Nr4a2, and Nr4a3. Up-regulation of all three Nr4a receptors was also observed in liver - but not brain - of DR-fed animals. Furthermore, RT-PCR revealed up-regulation of several NR4A transcriptional targets (Ucp-3, Ampk-gamma3, Pgc-1alpha and Pgc-1beta) in skeletal muscle of DR animals. Due to the proposed roles of the NR4A nuclear receptors in sensing and responding to changes in the nutritional environment and in regulating glucose and lipid metabolism and insulin sensitivity, we hypothesise that these proteins may contribute to DR-induced metabolic adaptation.

Gene expression changes in long-term culture of T-cell clones: genomic effects of chronic antigenic stress in aging and immunosenescence.

The adaptive immune response requires waves of T‐cell clonal expansion on contact with altered self and contraction after elimination of antigen. In the case of persisting antigen, as occurs for example in cytomegalovirus or Epstein–Barr virus infection, this critical process can become dysregulated and responding T‐cells enter into a dysfunctional senescent state. Longitudinal studies suggest that the presence of increased numbers of such T‐cells is a poor prognostic factor for survival in the very elderly. Understanding the nature of the defects in these T‐cells might facilitate intervention to improve immunity in the elderly. The process of clonal expansion under chronic antigenic stress can be modelled in vitro using continuously cultured T‐cells. Here, we have used cDNA array technology to investigate differences in gene expression in a set of five different T‐cell clones at early, middle and late passage in culture. Differentially expressed genes were confirmed by real‐time polymerase chain reaction, and relationships between these assessed using Ingenuity Systems evidence‐based association analysis. Several genes and chemokines related to induction of apoptosis and signal transduction pathways regulated by transforming growth factor β (TGFβ), epidermal growth factor (EGF), fos and β‐catenin were altered in late compared to early passage cells. These pathways and affected genes may play a significant role in driving the cellular senescent phenotype and warrant further investigation as potential biomarkers of aging and senescence. These genes may additionally provide targets for intervention.

SELDI-TOF-MS ProteinChip array profiling of T-cell clones propagated in long-term culture identifies human profilin-1 as a potential bio-marker of immunosenescence

BackgroundThe adaptive immune response requires waves of T-cell clonal expansion on contact with pathogen and elimination after clearance of the source of antigen. However, lifelong persistent infections with common viruses cause chronic antigenic stimulation which takes its toll on adaptive immunity in late life. Chronic antigenic stress results in deregulation of the T-cell response and accumulation of anergic cells. Longitudinal studies of the elderly show that this impacts on survival. Identifying the nature of the defects in chronically-stimulated T-cells and protein bio-markers of these dysfunctional cells would help to understand age-associated compromised T-cell function (immunosenescence) and facilitate the development of targeted intervention strategies.The purpose of this work was to use surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) to analyse proteins associated with T-cell senescence in order to identify potential bio-markers. Clonal populations of T-cells isolated from elderly octogenarian and centenarian donors were grown in vitro until senescence, and early passage and late passage (pre-senescent) cells were analysed using SELDI-TOF-MS ProteinChip arrays.ResultsDiscriminant analysis identified several protein or peptide peaks in the region of 14.5–16.5 kDa that were associated with T-cell clone senescence. Human profilin-1, a ubiquitous protein associated with actin remodelling and cellular motility was unambiguously identified. Altered expression of profilin-1 in senescent T-cell clones was confirmed by Western blot analysis.ConclusionDue to the proposed roles of profilin-1 in cellular survival, cytoskeleton remodelling, motility, and proliferation, it is hypothesised that differential expression of profilin-1 in ageing may contribute directly to immunosenescence.

Insulin Resistance, Chronic Inflammation and the Link with Immunosenescence

Ageing is associated with an activation of the innate immune system which manifests in a chronic, low-grade, inflammatory status common in elderly individuals. Age-related inflammatory activity, as measured by increased serum levels of proinflammatory cytokines and activation of inflammatory signalling pathways, leads to long-term tissue damage and is thought to contribute to—and occur as a consequence of—immunosenescence. In addition to immune system deregulation, this elevated inflammatory status is associated with a number of age-related diseases and conditions, including neurodegeneration, atherosclerosis, sarcopenia, and diabetes, and is a main contributor to the age-related decline in physical function and vitality known as frailty. Inflammation is also an important component of the insulin resistance syndrome. In addition to age, a major risk factor for the development of the insulin resistance syndrome is obesity. Obesity is associated with increased proinflammatory cytokine production and altered regulation of both pro and antiinflammatory molecules, including a class of adipose-derived signalling molecules termed adipocytokines. The increased production of inflammatory mol- ecules in obese and nonobese insulin resistant elderly individuals may contribute to age-related decline in health, including dysfunction of the immune system. Antiinflammatory strategies for the treatment of the insulin resistance syndrome may promote remodelling of the immune system thereby contributing to remediation of immunity and prevention of frailty in the elderly population.

SELDI Proteomics Approach to Identify Proteins Associated with T-Cell Clone Senescence

The immune system undergoes many complex changes as a result of the aging process. Elderly humans have altered cellular redox levels and deregulated immune responses, both key events underlying the progression of chronic degenerative diseases of aging, such as atherosclerosis and Alzheimer’s disease. T-cells are one of the major cell types affected by aging. As such, indentifying bio-markers of T-cell aging and senescence would aid in identifying and developing novel intervention strategies.