Effect of Lifelong Exposure to Dietary Plant and Marine Sources of n-3 Polyunsaturated Fatty Acids on Morphologic and Gene Expression Biomarkers of Intestinal Health in Early Life.

Altered intestinal health is also associated with the incidence and severity of many chronic inflammatory conditions, which could be attenuated via dietary n-3 PUFA interventions. However, little is known about the effect of lifelong exposure to n-3 PUFA from plant and marine sources (beginning in utero via the maternal diet) on early life biomarkers of intestinal health. Harems of C57Bl/6 mice were randomly assigned to one of three isocaloric AIN-93G modified diets differing in their fat sources consisting of the following: (i) 10% safflower oil (SO, enriched in n-6 PUFA), (ii) 3% flaxseed oil + 7% safflower oil (FX, plant-based n-3 PUFA-enriched diet), or (iii) 3% menhaden fish oil + 7% safflower oil (MO, marine-based n-3 PUFA-enriched diet). Mothers remained on these diets throughout pregnancy and offspring (n = 14/diet) continued on the same parental diet until termination at 3 weeks of age. In ileum, villi:crypt length ratios were increased in both the FX and MO dietary groups compared to SO (p < 0.05). Ileum mRNA expression of critical intestinal health biomarkers was increased by both n-3 PUFA-enriched diets including Relmß and REG3γ compared to SO (p < 0.05), whereas only the FX diet increased mRNA expression of TFF3 and Muc2 (p < 0.05) and only the MO diet increased mRNA expression of ZO-1 (p < 0.05). In the proximal colon, both the FX and MO diets increased crypt lengths compared to SO (p < 0.05), whereas only the MO diet increased goblet cell numbers compared to SO (p < 0.05). Further, the MO diet increased proximal colon mRNA expression of Relmß and REG3γ (p < 0.05) and both MO and FX increased mRNA expression of Muc2 compared to SO (p < 0.05). Collectively, these results demonstrate that lifelong exposure to dietary n-3 PUFA, beginning in utero, from both plant and marine sources, can support intestinal health development in early life. The differential effects between plant and marine sources warrants further investigation for optimizing health.

The Tolerance Model of Non-Inflammatory Immune Competence in Acute Pediatric Malnutrition: Origins, Evidence, Test of Fitness and Growth Potential.

The tolerance model rests on the thesis of a physiologically regulated, albeit unsustainable, systemic attempt to adapt to the catabolic challenge posed by acute prepubescent malnutrition even in its severe forms. The model centers on the immunological component of the attempt, positing reorientation toward a non-inflammatory form of competence in place of the classic paradigm of immunological attrition and exhaustion. The foundation of the model was laid in 1990, and sixteen years later it was articulated formally on the basis of a body of evidence centered on T cell cytokines and interventions with cytokine and hormonal mediators. The benefit originally suggested was a reduced risk of autoimmune pathologies consequent to the catabolic release of self-antigens, hence the designation highlighting immune tolerance. Herein, the emergence of the tolerance model is traced from its roots in the recognition that acute malnutrition elicits an endocrine-based systemic adaptive attempt. Thereafter, the growth of the evidence base supporting the model is outlined, and its potential to shed new light on existing information is tested by application to the findings of a published clinical study of acutely malnourished children. Finally, some knowledge gaps pertinent to the model are identified and its potential for growth consonant with evolving perceptions of immunobiology is illustrated.

N-3 Polyunsaturated Fatty Acids Ameliorate Neurobehavioral Outcomes Post-Mild Traumatic Brain Injury in the Fat-1 Mouse Model.

Concussions and mild traumatic brain injury (m-TBI) have been identified as a consequential public health concern because of their potential to cause considerable impairments in physical, cognitive, behavioral, and social functions. Given their prominent structural and functional roles in the brain, n-3 polyunsaturated fatty acids (PUFA) have been identified as a potentially viable prophylactic agent that may ameliorate the deleterious effects of m-TBI on brain function. The purpose of the present pilot study was to investigate the effect of n-3 PUFA on neurologic function using a weight drop injury (WDI) model. Fat-1 mice, capable of synthesizing n-3 PUFA endogenously from n-6 PUFA, and their wild-type (WT) counterparts, were subjected to a mild low-impact WDI on the closed cranium, and recovery was evaluated using the neurological severity score (NSS) to assess the motor and neurobehavioral outcomes. In comparison to the WT mice, the fat-1 mice had a significantly (p ≤ 0.05) lower NSS at all time points post-WDI, and significantly greater neurological restoration measured as the time to first movement. Overall, these findings demonstrate the protective effect of n-3 PUFA against mild brain injury.

An Evaluation of Omega-3 Status and Intake in Canadian Elite Rugby 7s Players.

BACKGROUND: EPA and DHA n-3 FA play crucial roles in both neurological and cardiovascular health and high dietary intakes along with supplementation suggest potential neuroprotection and concussion recovery support. Rugby athletes have a high risk of repetitive sub-concussive head impacts which may lead to long-term neurological deficits, but there is a lack of research looking into n-3 FA status in rugby players. We examined the dietary n-3 FA intake through a FFQ and n-3 FA status by measuring the percentage of n-3 FA and O3I in elite Canadian Rugby 7s players to show distribution across O3I risk zones; high risk, <4%; intermediate risk, 4 to 8%; and low risk, >8%. METHODS: n-3 FA profile and dietary intake as per FFQ were collected at the beginning of the 2017-2018 Rugby 7s season in male (n = 19; 24.84 ± 2.32 years; 95.23 ± 6.93 kg) and female (n = 15; 23.45 ± 3.10 years; 71.21 ± 5.79 kg) athletes. RESULTS: O3I averaged 4.54% ± 1.77, with female athlete scores slightly higher, and higher O3I scores in supplemented athletes (4.82% vs. 3.94%, p = 0.183), with a greater proportion of non-supplemented athletes in the high-risk category (45.5% vs. 39.1%). Dietary intake in non-supplemented athletes did not meet daily dietary recommendations for ALA or EPA + DHA compared to supplemented athletes. CONCLUSIONS: Overall, despite supplementation, O3I score remained in the high-risk category in a proportion of athletes who met recommended n-3 FA dietary intakes, and non-supplemented athletes had a higher proportion of O3I scores in the high-risk category, suggesting that dietary intake alone may not be enough and athletes may require additional dietary and n-3 FA supplementation to reduce neurological and cardiovascular risk.

The effect of omega-3 fatty acids on a biomarker of head trauma in NCAA football athletes: a multi-site, non-randomized study.

BACKGROUND: American-style football (ASF) athletes are at risk for cardiovascular disease (CVD) and exhibit elevated levels of serum neurofilament light (Nf-L), a biomarker of axonal injury that is associated with repetitive head impact exposure over the course of a season of competition. Supplementation with the w-3 fatty acid (FA) docosahexaenoic acid (DHA) attenuates serum Nf-L elevations and improves aspects of CVD, such as the omega-3 index (O3I). However, the effect of combining the w-3 FA eicosapentaenoic acid (EPA) and docosapentaenoic acid (DPA) with DHA on, specifically, serum Nf-L in ASF athletes is unknown. Therefore, this study assessed the effect of supplemental w-3 FA (EPA+DPA+DHA) on serum Nf-L, plasma w-3 FAs, the O3I, and surrogate markers of inflammation over the course of a season. METHODS: A multi-site, non-randomized design, utilizing two American football teams was employed. One team (n = 3 1) received supplementation with a highly bioavailablew-3 FA formulation (2000mg DHA, 560mg EPA, 320mg DPA, Mindset®, Struct Nutrition, Missoula, MT) during pre-season and throughout the regular season, while the second team served as the control (n = 35) and did not undergo supplementation. Blood was sampled at specific times throughout pre- and regular season coincident w ith changes in intensity, physical contact, and changes in the incidence and severity of head impacts. Group differences were determined via a mixed-model between-within subjects ANOVA. Effect sizes were calculated using Cohen's dfor all between-group differences. Significance was set a priori at p< .05. RESULTS: Compared to the control group, ASF athletes in the treatment group experienced large increases in plasma EPA (p < .001, d = 1.71) and DHA (p < .001, d = 2.10) which contributed to increases in the O3I (p < .001, d = 2.16) and the EPA:AA ratio (p = .001, d = 0.83) and a reduction in the w-6: w-3 ratio (p < .001, d = 1.80). w-3 FA supplementation attenuated elevations in Nf-L (p = .024). The control group experienced a significant increase in Nf-L compared to baseline at several measurement time points (T2, T3, and T4 [p range < .001 - .005, drange = 0.59-0.85]). CONCLUSIONS: These findings suggest a cardio- and neuroprotective effect of combined EPA+DPA+DHA w-3 FA supplementation in American-style football athletes. TRIAL REGISTRATION: This trial was registered with the ISRCTN registry ( ISRCTN90306741 ).

Analysis of major fatty acids from matched plasma and serum samples reveals highly comparable absolute and relative levels.

Measuring fatty acid (FA) levels in blood as a risk factor for chronic disease has been studied extensively. Previous research has used either plasma or serum samples to examine these associations. However, whether results from plasma and serum samples can be compared remains unclear, as differences in methodology related to the separation of plasma and serum from whole blood may impact FA levels. This study analyzed the individual FA content of matched plasma and serum samples in both absolute (µg/mL) and relative percent (%) composition. Analyses were performed using archived fasted morning samples from the Florey Adelaide Male Ageing Study (FAMAS). Matched plasma and serum samples were available from 98 male subjects aged 40-85. Total FA were analyzed by gas-liquid chromatography equipped with a flame ionization detector (GLC-FID). Analyses comprised of over 60 FA including major FA such as Palmitic Acid (PA), Palmitoleic acid (POA), Stearic Acid (SA), Oleic Acid (OA), Linoleic Acid (LNA), alpha-linolenic acid (ALA), Eicosapentaenoic acid (EPA), Arachidonic Acid (ARA), and Docosahexaenoic acid (DHA). Differences between groups was determined by t-test. Correlation and Bland-Altman analyses were also performed to examine the relationship between plasma and serum samples. There were no significant differences between major plasma and serum fatty acids expressed in µg/mL and relative % composition. Correlation analysis determined a strong and significantly positive association (r ≥ 0.65, p < 0.05) between major plasma and serum FA in absolute and relative terms. Bland-Altman analysis further supported the strong agreement between plasma and serum values in both absolute and relative terms. These findings demonstrate that studies reporting plasma or serum fatty acid analyzed by GLC-FID can be compared with one another.

Her-2 Breast Cancer Outcomes Are Mitigated by Consuming n-3 Polyunsaturated, Saturated, and Monounsaturated Fatty Acids Compared to n-6 Polyunsaturated Fatty Acids.

Lifestyle habits, such as the consumption of a healthy diet, may prevent up to 30-50% of breast cancer (BC) cases. Dietary fats are of specific interest, as research provides strong evidence regarding the association of dietary fats and BC. However, there is limited research on the role of different types of fats including polyunsaturated (PUFA), monounsaturated (MUFA), and saturated fatty acids (SFA). The objective of this study was to determine the effects of lifelong exposure to various dietary fats on mammary tumour development over a 20-week period. Female heterozygous MMTV-neu (ndl) YD5 mouse models were fed five maternal diets containing (1) 10% safflower oil (n-6 PUFA, control), (2) 3% menhaden oil + 7% safflower oil (marine n-3 PUFA, control), (3) 3% flaxseed + 7% safflower oil (plant-based n-3 PUFA), (4) 10% olive oil (MUFA), or (5) 10% lard (SFA). The primary measures, tumour latency, volume, and multiplicity differed by diet treatment in the following general order, n-6 PUFA > plant n-3 PUFA, SFA, MUFA > marine n-3 PUFA. Overall, these findings show that the quality of the diet plays a significant role influencing mammary tumour outcomes.

Lifelong n-3 Polyunsaturated Fatty Acid Exposure Modulates Size of Mammary Epithelial Cell Populations and Expression of Caveolae Resident Proteins in Fat-1 Mice.

Omega-3 polyunsaturated fatty acids (n-3 PUFA) have been associated with reduced breast cancer risk; however, the exact mechanism remains elusive. Female wildtype (WT) and fat-1 mice were fed a 10% safflower diet until 6 weeks of age. Mammary gland epithelial cells (EC) were isolated and EC populations were determined by CD24 surface expression. Fat-1 mice expressed 65%, 20%, and 15% while WT mice expressed 65%, 26% and 9% for non-, myo- and luminal ECs, respectively. The luminal EC population was significantly greater in fat-1 mice (p ≤ 0.05), while the total number of mammary ECs were similar between groups (p = 0.79). Caveolae was isolated from ECs and Her-2/neu, ER-α and cav-1 protein expression was determined by Western blotting. Fat-1 mice had a two-fold greater ER-α (p ≤ 0.05) and a 1.5-fold greater cav-1 (p ≤ 0.05) expression than WT with a similar amount of Her-2/neu protein (p = 0.990) between groups. Overall, this study provides novel mechanistic evidence by which n-3 PUFA modifies early mammary gland development that may potentially reduce breast cancer risk later in life.

Acutely malnourished weanling mice administered Flt3 ligand can support a cell-mediated inflammatory response.

The main objective of this investigation was to determine whether, despite acute (wasting) deficits of dietary nitrogen and energy, weanling mice could respond to the dendritic cell hematopoietin, Fms-like tyrosine kinase 3 ligand (Flt3L), in terms of an index of cell-mediated inflammatory competence. Male and female C57BL/6J weanlings were used, initially 19 days of age, and malnutrition was produced using a nitrogen-deficient diet. In preliminary work ten daily subcutaneous 1.0 µg doses of murine Flt3L, comparable to a protocol effective in humans, expanded the splenic conventional dendritic cell compartment (CD11c+F4/80-/low) of healthy weanlings without affecting the numbers of lymphocytes, macrophages, or recoverable mononuclear cells. Two subsequent experiments showed that, despite advancing malnutrition, exogenous Flt3L was able both to exert its classic influence on splenic conventional dendritic cell numbers and to invigorate the attenuated primary splenic cell-mediated inflammatory response to sheep erythrocytes. A final experiment showed that the cytokine intervention did not affect dendritic cell maturity according to several phenotypic indices. The findings provide new support for the proposition that dendritic cell numbers are the first limiting factor in the weak cell-mediated immune competence of acute pre-pubescent malnutrition. More substantially, intervention with Flt3L sustained an inflammatory systemic immune character despite progressive weanling malnutrition and weight loss. This outcome provides new support of fundamental character for the Tolerance Model which posits that the cell-mediated inflammatory incompetence of acute pre-pubescent protein and energy deficits is a regulated adaptive attempt, the antithesis of the classic paradigm of unregulated immunological attrition.

Novel Phospholipid Signature of Depressive Symptoms in Patients With Coronary Artery Disease.

BACKGROUND: Depression in patients with coronary artery disease (CAD) is associated with increased cardiovascular morbidity. Given the proinflammatory actions of phospholipids, aberrant phospholipid metabolism may be an etiological mechanism linking CAD and depression. Our primary objective was to identify a phospholipid biomarker panel that characterizes CAD patients with significant depressive symptoms from those without. METHODS AND RESULTS: We performed a targeted lipidomic analysis on CAD patients with significant depressive symptoms (n=37, Center for Epidemiologic Studies Depression score ≥16) and those without (n=49). Phospholipid species were selected using partial least-square discriminant analysis, and the ability of the resulting model to discriminate between groups was evaluated using receiver operator characteristic curves. Biosignature scores were calculated from this model, and analyses of covariance were performed to compare intergroup differences in biosignature scores, with adjustment for clinical differences between patients. Those with significant depressive symptoms had lower cardiopulmonary fitness, more prevalent history of depression, and a greater number of vascular risk factors. A model of 10 phospholipid species had an area under the curve value of 0.84 (95% confidence interval 0.72-0.95), sensitivity of 0.73, and specificity of 0.71. This model passed permutation testing (n=1000, P<0.001). Biosignature scores were higher in those with significant depressive symptoms after adjustment for potential confounders (F[1.86]=14.39, P<0.0005). CONCLUSIONS: The present findings support the role of proinflammatory phospholipid species in the presence of depression in CAD patients from the CAROTID trial (Coronary Artery Disease Randomized Omega-3 Trial in Depression). Future investigations should aim to replicate findings in larger data sets and clarify possible pathophysiological mechanisms. CLINICAL TRIAL REGISTRATION: URL: https://www.clinicaltrials.gov. Unique identifier: NCT00981383.

Marine fish oil is more potent than plant-based n-3 polyunsaturated fatty acids in the prevention of mammary tumors.

Marine-derived n-3 polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have been shown to inhibit mammary carcinogenesis. However, evidence regarding plant-based α-linolenic acid (ALA), the major n-3 PUFA in the Western diet, remains equivocal. The objective of this study was to examine the effect of lifelong exposure to plant- or marine-derived n-3 PUFAs on pubertal mammary gland and tumor development in MMTV-neu(ndl)-YD5 mice. It is hypothesized that lifelong exposure to n-3 PUFA reduces terminal end buds during puberty leading to delayed tumor onset, volume and multiplicity. It is further hypothesized that plant-derived n-3 PUFAs will exert dose-dependent effects. Harems of MMTV-FVB males were bred with wild-type females and fed either a (1) 10% safflower (10% SF, n-6 PUFA, control), (2) 10% flaxseed (10% FS), (3) 7% safflower plus 3% flaxseed (3% FS) or (4) 7% safflower plus 3% menhaden (3% FO) diet. Female offspring were maintained on parental diets. Compared to SF, 10% FS and 3% FO reduced (P<.05) terminal end buds at 6 weeks and tumor volume and multiplicity at 20 weeks. A dose-dependent reduction of tumor volume and multiplicity was observed in mice fed 3% and 10% FS. Antitumorigenic effects were associated with altered HER2, pHER-2, pAkt and Ki-67 protein expression. Compared to 10% SF, 3% FO significantly down-regulated expression of genes involved in eicosanoid synthesis and inflammation. From this, it can be estimated that ALA was 1/8 as potent as EPA+DHA. Thus, marine-derived n-3 PUFAs have greater potency versus plant-based n-3 PUFAs.

The delta 6 desaturase knock out mouse reveals that immunomodulatory effects of essential n-6 and n-3 polyunsaturated fatty acids are both independent of and dependent upon conversion.

Typically fatty acids (FA) exert differential immunomodulatory effects with n-3 [α-linolenic acid (ALA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)] and n-6 [linoleic acid (LA) and arachidonic acid (AA)] exerting anti- and pro-inflammatory effects, respectively. This over-simplified interpretation is confounded by a failure to account for conversion of the parent FA (LA and ALA) to longer-chain bioactive products (AA and EPA/DHA, respectively), thereby precluding discernment of the immunomodulatory potential of specific FA. Therefore, we utilized the Δ6-desaturase model, wherein knockout mice (D6KO) lack the Fads2 gene encoding for the rate-limiting enzyme that initiates FA metabolism, thereby providing a model to determine specific FA immunomodulatory effects. Wild-type (WT) and D6KO mice were fed one of four isocaloric diets differing in FA source (9weeks): corn oil (LA-enriched), arachidonic acid single cell oil (AA-enriched), flaxseed oil (ALA-enriched) or menhaden fish oil (EPA/DHA-enriched). Splenic mononuclear cell cytokine production in response to lipopolysaccharide (LPS), T-cell receptor (TCR) and anti-CD40 stimulation was determined. Following LPS stimulation, AA was more bioactive compared to LA, by increasing inflammatory cytokine production of IL-6 (1.2-fold) and TNFα (1.3-fold). Further, LPS-stimulated IFNγ production in LA-fed D6KO mice was reduced 5-fold compared to LA-fed WT mice, indicating that conversion of LA to AA was necessary for cytokine production. Conversely, ALA exerted an independent immunomodulatory effect from EPA/DHA and all n-3 FA increased LPS-stimulated IL-10 production versus LA and AA. These data definitively identify specific immunomodulatory effects of individual FA and challenge the simplified view of the immunomodulatory effects of n-3 and n-6 FA.

Omega-3/omega-6 fatty acid ratios in different phospholipid classes and depressive symptoms in coronary artery disease patients.

Depressive symptoms are highly incident among coronary artery disease (CAD) patients and increase mortality. Reduced ratios of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (omega-3 fatty acids) to arachidonic acid (AA, omega-6 fatty acid) concentrations have been linked with depressive symptoms in CAD. It remains unclear whether depressive symptoms are differentially associated with that ratio in different phospholipid classes, and this may have mechanistic implications. This study investigated associations between depressive symptoms in CAD patients and the EPA+DHA to AA ratio in the major phospholipid classes. This was a cross-sectional study of stable CAD patients. Sociodemographic, medical, medication, and cardiopulmonary fitness data were collected from each patient. Each patient was assessed for depressive symptoms using the 17-item Hamilton Depression Rating Scale (HAM-D). The percentage of EPA, DHA, and AA in each erythrocyte phospholipid class was determined using gas chromatography from fasting blood. Relationships between EPA+DHA to AA ratios and depressive symptoms were assessed using linear regression and were corrected for multiple comparisons. Seventy-six CAD patients were included (age=61.9 ± 8.5, 74% male, HAM-D=7.2 ± 5.9). In a backward elimination linear regression model, lower EPA+DHA to AA in erythrocyte phosphatidylinositol (B=-12.71, ß=-0.33, p<.01) and sphingomyelin (B=-2.52, ß=-0.37, p<.01) was associated with greater depressive symptom severity, independently of other known predictors. Other phospholipid classes were not associated with depressive symptoms. In conclusion, the relationship between EPA+DHA to AA ratios and depressive symptoms in CAD may not be consistent across phospholipid classes. Continued investigation of these potentially differential relationships may clarify underlying disease mechanisms.

Lifelong exposure to n-3 PUFA affects pubertal mammary gland development.

There is growing evidence that early developmental periods may importantly influence future breast cancer risk. Also, there is great interest in the role of dietary fat in breast cancer risk, but the role of dietary fat during pubertal mammary gland development remains poorly understood. This study investigated the effect of n-3 polyunsaturated fatty acids (PUFA) using complementary dietary and genetic approaches to examine the effect of lifelong exposure of n-3 PUFA or n-6 PUFA (control) on mammary gland development and fatty acid composition. n-3 PUFA from both diet and genetics were enriched in mammary glands as early as 3 weeks of age. Parameters related to mammary gland development, including number of terminal end buds (TEB), percent coverage of ductal tree, and infiltration of TEB, were influenced by n-3 PUFA at 3 and 4 weeks of age. Overall, findings suggest that n-3 PUFA incorporation into the mammary gland early in life plays a role in the morphological development of the mammary gland during puberty.

Carcinogenesis alters fatty acid profile in breast tissue.

Cancerogenesis is associated with cell membrane changes. The aim of this study was to investigate whether breast tissues with different degrees of cancer involvement have different fatty acid profiles. Fourteen breast cancer patients with a mean age of 61 years were recruited. Morphological features of the tumoral specimens were characterized. Approximately 60 % of patients had invasive ductal carcinoma, and 80 % were ER positive; 65 % were PR positive; and 65 % were HER2 negative. The segments with confirmed cancer had significantly less amounts of total lipids as compared with the corresponding grossly normal or interface tissues. The fatty acid profile in cancer tissue was significantly different from that in other tissues. Fatty acid composition of five classes of phospholipids revealed the variations between cancer tissue and the other two segments. A transition of changes in fatty acid composition in these fractions of phospholipids was observed. The interface tissue had intermediate amounts of several fatty acids including palmitic acid, stearic acid, and arachidonic acid. Interestingly, we observed significantly higher amounts of the n-3 fatty acid DHA in cancer tissue as compared to the other two tissues. Data from this study will provide evidence that biochemical changes particularly phospholipid composition may take place well in advance prior to morphological changes. Should this theory be confirmed by larger studies, deviation of phospholipid composition from normal values can be used as markers of susceptibility of tissue to cancer development.

Vaccenic acid in serum triglycerides is associated with markers of insulin resistance in men.

Serum triglyceride levels are associated with metabolic disorders; however, it remains unclear whether the fatty acid (FA) composition of triglycerides is also changed. Although there were no differences in circulating triglyceride levels between normoglycaemic-normoinsulinaemic and hyperglycaemic-hyperinsulinaemic men, inspection of individual FA revealed that vaccenic acid was enriched with hyperglycaemia-hyperinsulinaemia. Moreover, vaccenic acid levels were positively correlated with insulin and HOMA-IR. This reinforces that examination of individual FA in the context of insulin resistance is warranted.

Constitutive, but not challenge-induced, interleukin-10 production is robust in acute pre-pubescent protein and energy deficits: new support for the tolerance hypothesis of malnutrition-associated immune depression based on cytokine production in vivo.

The tolerance model of acute (i.e., wasting) pre-pubescent protein and energy deficits proposes that the immune depression characteristic of these pathologies reflects an intact anti-inflammatory form of immune competence that reduces the risk of autoimmune reactions to catabolically released self antigens. A cornerstone of this proposition is the finding that constitutive (first-tier) interleukin(IL)-10 production is sustained even into the advanced stages of acute malnutrition. The IL-10 response to inflammatory challenge constitutes a second tier of anti-inflammatory regulation and was the focus of this investigation. Weanling mice consumed a complete diet ad libitum, a low-protein diet ad libitum (mimicking incipient kwashiorkor), or the complete diet in restricted daily quantities (mimicking marasmus), and their second-tier IL-10 production was determined both in vitro and in vivo using lipopolysaccharide (LPS) and anti-CD3 as stimulants of innate and adaptive defences, respectively. Both early (3 days) and advanced (14 days) stages of wasting pathology were examined and three main outcomes emerged. First, classic in vitro systems are unreliable for discerning cytokine production in vivo. Secondly, in diverse forms of acute malnutrition declining challenge-induced IL-10 production may provide an early sign that anti-inflammatory control over immune competence is failing. Thirdly, and most fundamentally, the investigation provides new support for the tolerance model of malnutrition-associated inflammatory immune depression.

Effector/memory T cells of the weanling mouse exhibit Type 2 cytokine polarization in vitro and in vivo in the advanced stages of acute energy deficit.

Our objective was to determine whether the polarizing cytokine profile of the effector/memory T-cell compartment reflects the profound decline of cell-mediated inflammatory competence that characterizes acute prepubescent malnutrition. Weanling C57BL/6J mice were permitted free access to a complete purified diet, free access to an isocaloric low-protein diet or restricted intake of the complete diet for 14 days. First, interleukin (IL)-4 and interferon (IFN)-gamma concentrations generated in vitro by splenic and nodal effector/memory T cells were assessed following exposure to plate-bound anti-CD3. Second, net systemic production of IFN-gamma and IL-4 by the effector/memory T-cell compartment was assessed by the in vivo cytokine capture assay following anti-CD3 stimulation. In vitro stimulation generated less IFN-gamma (P=.002) but more IL-4 (P=.05) by T cells from the restricted-intake group relative to the age-matched control group. Similarly, in vivo stimulation generated low serum levels of antibody-captured IFN-gamma in the restricted-intake group vis-à-vis the age-matched control group (P=.01), while the IL-4 response was sustained (P=.39). By contrast, the 14-day low-protein model exhibited no change in T-cell cytokine signature either in vitro or in vivo. However, following extended consumption of the low-protein diet (26 days), carcass energy losses exceeded those of the 14-day protocol and serum levels of in vivo antibody-captured IFN-gamma were low after anti-CD3 challenge relative to the age-matched control group (P=.02), while levels of captured IL-4 remained unaffected (P=.07). Acute weanling malnutrition elicits a Type 2 polarizing cytokine character on the part of the effector/memory T-cell compartment, but only in the most advanced stages of energy decrement.

Blood serum interferon-gamma bioactivity is low in weanling mice subjected to acute deficits of energy or both protein and energy.

The main objective of the present study was to determine the influence of acute deficits of protein and energy on the blood serum level of interferon-gamma, a signature type 1 polarising inflammatory cytokine. In two 14 d experiments, male and female C57BL/6J mice, initial age 19 d, consumed a complete purified diet ad libitum or in restricted daily quantities, or had free access to an isoenergetic purified low-protein diet. A zero-time control group (age 19 d) was included in the second experiment. Serum interferon-gamma was assessed in both experiments by sandwich ELISA and, in the second experiment, also by a bioassay based on inhibition of proliferation by WEHI-279 B lymphoma cells. The immunoassay detected interferon-gamma inconsistently in all groups (range 0-14 pg/ml; detection limits 1 x 5 and 0 x 7 pg/ml in experiments 1 and 2, respectively). By contrast, interferon-gamma bioactivity was found in all animals of each group (means 339, 499, 124 and 200 pg/ml in zero-time controls, age-matched controls, low-protein and restricted intake groups, respectively; detection limit, 12 pg/ml), and the mean serum bioactivity of each malnourished group was low compared with the age-matched control (P

The capacity of noninflammatory (steady-state) dendritic cells to present antigen in the primary response is preserved in acutely protein- or energy-deficient weanling mice.

The objective of this investigation was to determine the influence of wasting protein and/or energy deficits on the capacity of dendritic cells to initiate primary responses. Weanling male and female C57BL/6J mice were permitted free access to a complete purified diet, free access to an isocaloric low protein purified diet (combined deficiencies of protein and energy) or restricted intake of the complete diet (energy deficiency) for up to 14 d; a 19-d-old zero-time control group was also included. Malnourished mice lost 1.5-2% of initial body weight daily. Antigen presentation by dendritic cells from spleen and lymph nodes was assessed in vitro by the primary one-way allogeneic mixed lymphocyte reaction using CBA/J mononuclear or CD4(+) T cells as responders. This function was sustained despite advanced weight loss and, remarkably, was increased in cell suspensions from 14-d energy-deficient mice. Antigen presentation by dendritic cells in mononuclear suspensions was examined in vivo using the host-vs.-graft response in CBA/J recipients, and an ontogeny-related increase was sustained in both malnourished groups through 14 d of weight loss. Neither wasting protocol influenced the proportion of mononuclear cells (1-2%) exhibiting dendritic cell phenotype (CD11c(+)F4/80(-/low)) in the cellular suspensions used to study antigen-presenting activity. Consequently, these functional studies are interpretable on a per dendritic cell basis. In the absence of infectious or inflammatory pressure, the dendritic cell retains antigen-presenting capacity despite acute (wasting) deficiencies of protein and/or energy. The results are relevant to presentation of both foreign (adjuvant role) and self (tolerizing role) antigens by the dendritic cell.