Novel Fermentates Can Enhance Key Immune Responses Associated with Viral Immunity.

Fermented foods have long been known to have immunomodulatory capabilities, and fermentates derived from the lactic acid bacteria of dairy products can modulate the immune system. We have used skimmed milk powder to generate novel fermentates using Lb. helveticus strains SC234 and SC232 and we demonstrate here that these fermentates can enhance key immune mechanisms that are critical to the immune response to viruses. We show that our novel fermentates, SC234 and SC232, can positively impact on cytokine and chemokine secretion, nitric oxide (NO) production, cell surface marker expression, and phagocytosis in macrophage models. We demonstrate that the fermentates SC234 and SC232 increase the secretion of cytokines IL-1ß, IL-6, TNF-α, IL-27, and IL-10; promote an M1 pro-inflammatory phenotype for viral immunity via NO induction; decrease chemokine expression of Monocyte Chemoattractant Protein (MCP); increase cell surface marker expression; and enhance phagocytosis in comparison to their starting material. These data suggest that these novel fermentates have potential as novel functional food ingredients for the treatment, management, and control of viral infection.

Targeted Application of Functional Foods as Immune Fitness Boosters in the Defense against Viral Infection.

In recent times, the emergence of viral infections, including the SARS-CoV-2 virus, the monkeypox virus, and, most recently, the Langya virus, has highlighted the devastating effects of viral infection on human life. There has been significant progress in the development of efficacious vaccines for the prevention and control of viruses; however, the high rates of viral mutation and transmission necessitate the need for novel methods of control, management, and prevention. In recent years, there has been a shift in public awareness on health and wellbeing, with consumers making significant dietary changes to improve their immunity and overall health. This rising health awareness is driving a global increase in the consumption of functional foods. This review delves into the benefits of functional foods as potential natural means to modulate the host immune system to enhance defense against viral infections. We provide an overview of the functional food market in Europe and discuss the benefits of enhancing immune fitness in high-risk groups, including the elderly, those with obesity, and people with underlying chronic conditions. We also discuss the immunomodulatory mechanisms of key functional foods, including dairy proteins and hydrolysates, plant-based functional foods, fermentates, and foods enriched with vitamin D, zinc, and selenium. Our findings reveal four key immunity boosting mechanisms by functional foods, including inhibition of viral proliferation and binding to host cells, modulation of the innate immune response in macrophages and dendritic cells, enhancement of specific immune responses in T cells and B cells, and promotion of the intestinal barrier function. Overall, this review demonstrates that diet-derived nutrients and functional foods show immense potential to boost viral immunity in high-risk individuals and can be an important approach to improving overall immune health.

Autoantibodies - enemies, and/or potential allies?

Autoantibodies are well known as potentially highly harmful antibodies which attack the host via binding to self-antigens, thus causing severe associated diseases and symptoms (e.g. autoimmune diseases). However, detection of autoantibodies to a range of disease-associated antigens has enabled their successful usage as important tools in disease diagnosis, prognosis and treatment. There are several advantages of using such autoantibodies. These include the capacity to measure their presence very early in disease development, their stability, which is often much better than their related antigen, and the capacity to use an array of such autoantibodies for enhanced diagnostics and to better predict prognosis. They may also possess capacity for utilization in therapy, in vivo. In this review both the positive and negative aspects of autoantibodies are critically assessed, including their role in autoimmune diseases, cancers and the global pandemic caused by COVID-19. Important issues related to their detection are also highlighted.

Exopolysaccharides of Lactic Acid Bacteria: Production, Purification and Health Benefits towards Functional Food.

Lactic acid bacteria (LAB) are capable of synthesising metabolites known as exopolysaccharides (EPS) during fermentation. Traditionally, EPS plays an important role in fermented dairy products through their gelling and thickening properties, but they can also be beneficial to human health. This bioactivity has gained attention in applications for functional foods, which leads them to have prebiotic, immunomodulatory, antioxidant, anti-tumour, cholesterol-lowering and anti-obesity activity. Understanding the parameters and conditions is crucial to optimising the EPS yields from LAB for applications in the food industry. This review provides an overview of the functional food market together with the biosynthesis of EPS. Factors influencing the production of EPS as well as methods for isolation, characterisation and quantification are reviewed. Finally, the health benefits associated with EPS are discussed.

Surface layer proteins from virulent Clostridium difficile ribotypes exhibit signatures of positive selection with consequences for innate immune response.

BACKGROUND: Clostridium difficile is a nosocomial pathogen prevalent in hospitals worldwide and increasingly common in the community. Sequence differences have been shown to be present in the Surface Layer Proteins (SLPs) from different C. difficile ribotypes (RT) however whether these differences influence severity of infection is still not clear. RESULTS: We used a molecular evolutionary approach to analyse SLPs from twenty-six C. difficile RTs representing different slpA sequences. We demonstrate that SLPs from RT 027 and 078 exhibit evidence of positive selection (PS). We compared the effect of these SLPs to those purified from RT 001 and 014, which did not exhibit PS, and demonstrate that the presence of sites under positive selection correlates with ability to activate macrophages. SLPs from RTs 027 and 078 induced a more potent response in macrophages, with increased levels of IL-6, IL-12p40, IL-10, MIP-1α, MIP-2 production relative to RT 001 and 014. Furthermore, RTs 027 and 078 induced higher expression of CD40, CD80 and MHC II on macrophages with decreased ability to phagocytose relative to LPS. CONCLUSIONS: These results tightly link sequence differences in C. difficile SLPs to disease susceptibility and severity, and suggest that positively selected sites in the SLPs may play a role in driving the emergence of hyper-virulent strains.

The Limonoids TS3 and Rubescin E Induce Apoptosis in Human Hepatoma Cell Lines and Interfere with NF-κB Signaling.

Hepatocellular carcinoma (HCC) is extremely resistant towards pharmacological therapy. To date, the multi-kinase inhibitor Sorafenib is the only available therapeutic agent with the potential to prolong patient survival. Using the human hepatoma cell lines HepG2 and Huh7, we analyzed anti-cancer activities of 6 purified havanensin type limonoids isolated from the traditional African medicinal plant Trichilia rubescens Oliv. Our results show that two of the compounds, TR4 (TS3) and TR9 (Rubescin E) reduced hepatoma cell viability, but not primary hepatocyte viability, at TC50s of 5 to 10 µM. These were significantly lower than the TC50s for Sorafenib, the histone deacetylase inhibitor SAHA or 5-Fluoruracil. In comparison, TR3 (Rubescin D), a limonoid isolated in parallel and structurally highly similar to TR4 and TR9, did not interfere with hepatoma cell viability. Both, TR4 and TR9, but not TR3, induced apoptosis in hepatoma cells and interfered with NF-κB activation. TR4 as well as TR9 significantly supported anti-cancer activities of Sorafenib. In summary, the limonoids TR4 and TR9 exhibit anti-cancer activities and support Sorafenib effects in vitro, having the potential to support future HCC therapy.

Khaya grandifoliola C.DC: a potential source of active ingredients against hepatitis C virus in vitro.

In this study, we examined the antiviral properties of Khaya grandifoliola C.DC (Meliaceae) on the hepatitis C virus (HCV) life cycle in vitro and identified some of the chemical constituents contained in the fraction with the most antiviral activity. Dried bark powder was extracted by maceration in a methylene chloride/methanol (MCM) system (50:50; v/v) and separated on silica gel by flash chromatography. Infection and replication rates in Huh-7 cells were investigated by luciferase reporter assay and indirect immunofluorescence assay using subgenomic replicons, HCV pseudotyped particles, and cell-culture-derived HCV (HCVcc), respectively. Cell viability was assessed by MTT assay, and cellular gene expression was analysed by qRT-PCR. The chemical composition of the fraction with the most antiviral activity was analysed by coupled gas chromatography and mass spectrometry (GC-MS). Five fractions of different polarities (F0-F100) were obtained from the MCM extract. One fraction (KgF25) showed the strongest antiviral effect on LucUbiNeoET replicons at nontoxic concentrations. Tested at 100 µg/mL, KgF25 had a high inhibitory effect on HCV replication, comparable to that of 0.01 µM daclatasvir or 1 µM telaprevir. This fraction also inhibited HCVcc infection by mostly targeting the entry step. KgF25 inhibited HCV entry in a pan-genotypic manner by directly inactivating free viral particles. Its antiviral effects were mediated by the transcriptional upregulation of the haem oxygenase-1 gene and interferon antiviral response. Three constituents, namely, benzene, 1,1'-(oxydiethylidene)bis (1), carbamic acid, (4-methylphenyl)-, 1-phenyl (2), and 6-phenyl, 4-(1'-oxyethylphenyl) hexene (3), were identified from the active fraction KgF25 by GC-MS. Khaya grandifoliola contains ingredients capable of acting on different steps of the HCV life cycle.

Profiling Humoral Immune Responses to Clostridium difficile-Specific Antigens by Protein Microarray Analysis.

Clostridium difficile is an anaerobic, Gram-positive, and spore-forming bacterium that is the leading worldwide infective cause of hospital-acquired and antibiotic-associated diarrhea. Several studies have reported associations between humoral immunity and the clinical course of C. difficile infection (CDI). Host humoral immune responses are determined using conventional enzyme-linked immunosorbent assay (ELISA) techniques. Herein, we report the first use of a novel protein microarray assay to determine systemic IgG antibody responses against a panel of highly purified C. difficile-specific antigens, including native toxins A and B (TcdA and TcdB, respectively), recombinant fragments of toxins A and B (TxA4 and TxB4, respectively), ribotype-specific surface layer proteins (SLPs; 001, 002, 027), and control proteins (tetanus toxoid and Candida albicans). Microarrays were probed with sera from a total of 327 individuals with CDI, cystic fibrosis without diarrhea, and healthy controls. For all antigens, precision profiles demonstrated <10% coefficient of variation (CV). Significant correlation was observed between microarray and ELISA in the quantification of antitoxin A and antitoxin B IgG. These results indicate that microarray is a suitable assay for defining humoral immune responses to C. difficile protein antigens and may have potential advantages in throughput, convenience, and cost.

Soluble factors from colonic epithelial cells contribute to gut homeostasis by modulating macrophage phenotype.

Intestinal macrophages originate from inflammatory blood monocytes which migrate to the intestine, where they differentiate into anti-inflammatory macrophages through a number of transitional stages. These macrophages typically remain hypo-responsive to commensal bacteria and food Ags in the intestine, yet also retain the ability to react to invading pathogens. In this study we examined the role of epithelial cells in inducing this intestinal macrophage phenotype. Using an in vitro system we showed that, in two-dimensional culture, epithelial cell-derived factors from a murine cell line, CMT-93, are sufficient to induce phenotypic changes in macrophages. Exposure of monocyte-derived macrophages, J774A.1, to soluble factors derived from epithelial cells, induced an altered phenotype similar to that of intestinal macrophages with decreased production of IL-12p40, IL-6 and IL-23 and expression of MHC ІІ and CD80 following TLR ligation. Furthermore, these conditioned macrophages showed enhanced phagocytic activity in parallel with low respiratory burst and NO production, similar to the response seen in intestinal macrophages. Our findings suggest a role for colonic epithelial cells in modulation of macrophage phenotype for maintenance of gut homeostasis. Further understanding of the cell interactions that maintain homeostasis in the gut could reveal novel therapeutic strategies to restore the balance in disease.

Adult vascular smooth muscle cells in culture express neural stem cell markers typical of resident multipotent vascular stem cells.

Differentiation of resident multipotent vascular stem cells (MVSCs) or de-differentiation of vascular smooth muscle cells (vSMCs) might be responsible for the SMC phenotype that plays a major role in vascular diseases such as arteriosclerosis and restenosis. We examined vSMCs from three different species (rat, murine and bovine) to establish whether they exhibit neural stem cell characteristics typical of MVSCs. We determined their SMC differentiation, neural stem cell marker expression and multipotency following induction in vitro by using immunocytochemistry, confocal microscopy, fluorescence-activated cell sorting analysis and quantitative real-time polymerase chain reaction. MVSCs isolated from rat aortic explants, enzymatically dispersed rat SMCs and rat bone-marrow-derived mesenchymal stem cells served as controls. Murine carotid artery lysates and primary rat aortic vSMCs were both myosin-heavy-chain-positive but weakly expressed the neural crest stem cell marker, Sox10. Each vSMC line examined expressed SMC differentiation markers (smooth muscle α-actin, myosin heavy chain and calponin), neural crest stem cell markers (Sox10(+), Sox17(+)) and a glia marker (S100ß(+)). Serum deprivation significantly increased calponin and myosin heavy chain expression and decreased stem cell marker expression, when compared with serum-rich conditions. vSMCs did not differentiate to adipocytes or osteoblasts following adipogenic or osteogenic inductive stimulation, respectively, or respond to transforming growth factor-ß1 or Notch following γ-secretase inhibition. Thus, vascular SMCs in culture express neural stem cell markers typical of MVSCs, concomitant with SMC differentiation markers, but do not retain their multipotency. The ultimate origin of these cells might have important implications for their use in investigations of vascular proliferative disease in vitro.

Surface layer proteins isolated from Clostridium difficile induce clearance responses in macrophages.

Clostridium difficile is the leading cause of hospital-acquired diarrhoea worldwide, and if the bacterium is not cleared effectively it can pose a risk of recurrent infections and complications such as colitis, sepsis and death. In this study we demonstrate that surface layer proteins from the one of the most frequently acquired strains of C. difficile, activate mechanisms in murine macrophage in vitro that are associated with clearance of bacterial infection. Surface layer proteins (SLPs) isolated from C. difficile induced the production of pro-inflammatory cytokines and chemokines and increased macrophage migration and phagocytotic activity in vitro. Furthermore, we also observed up-regulation of a number of cell surface markers on the macrophage, which are important in pathogen recognition and antigen presentation. The effects of SLPs on macrophages were reversed in the presence of a p38 inhibitor, indicating the potential importance of this signalling protein in how SLP activates the immune system. In conclusion this study shows that surface layer proteins from a common strain of C. difficile can activate a clearance response in macrophage and suggests that these proteins are important in clearance of C. difficile infection. Understanding how the immune system clears C. difficile infection could offer important insights for new treatment strategies.

Activation of liver X receptor suppresses the production of the IL-12 family of cytokines by blocking nuclear translocation of NF-κBp50.

There is now convincing evidence that liver X receptor (LXR) is an important modulator of the inflammatory response; however, its mechanism of action remains unclear. This study aimed to examine the effect of LXR on the IL-12 family of cytokines and examined the mechanism by which LXR exerted this effect. We first demonstrated that activation of murine-derived dendritic cells (DC) with a specific agonist to LXR enhanced expression of LXR following activation with LPS, suggesting a role in inflammation. Furthermore, we showed LXR expression to be increased in vivo in dextrane sulphate sodium-induced colitis. LXR activation also suppressed production of IL-12p40, IL-12p70, IL-27 and IL-23 in murine-derived DC following stimulation with LPS, and specifically targeted the p35, p40 and EBI3 subunits of the IL-12 cytokine family, which are under the control of the NF-κB subunit p50 (NF-κBp50). Finally, we demonstrated that LXR can associate with NF-κBp50 in DC and that LXR activation prevents translocation of the p50 subunit into the nucleus. In summary, our study indicates that LXR can specifically suppress the IL-12 family of cytokines though its association with NF-κBp50 and highlights its potential as a therapeutic target for chronic inflammatory diseases.

Syntaxin-4 is essential for IgE secretion by plasma cells.

The humoral immune system provides a crucial first defense against the invasion of microbial pathogens via the secretion of antigen specific immunoglobulins (Ig). The secretion of Ig is carried out by terminally differentiated B-lymphocytes called plasma cells. Despite the key role of plasma cells in the immune response, the mechanisms by which they constitutively traffic large volumes of Ig out of the cell is poorly understood. The involvement of Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins in the regulation of protein trafficking from cells has been well documented. Syntaxin-4, a member of the Qa SNARE syntaxin family has been implicated in fusion events at the plasma membrane in a number of cells in the immune system. In this work we show that knock-down of syntaxin-4 in the multiple myeloma U266 human plasma cell line results in a loss of IgE secretion and accumulation of IgE within the cells. Furthermore, we show that IgE co-localises with syntaxin-4 in U266 plasma cells suggesting direct involvement in secretion at the plasma membrane. This study demonstrates that syntaxin-4 plays a critical role in the secretion of IgE from plasma cells and sheds some light on the mechanisms by which these cells constitutively traffic vesicles to the surface for secretion. An understanding of this machinery may be beneficial in identifying potential therapeutic targets in multiple myeloma and autoimmune disease where over-production of Ig leads to severe pathology in patients.

Network analysis of adipose tissue gene expression highlights altered metabolic and regulatory transcriptomic activity in high-fat-diet-fed IL-1RI knockout mice.

A subacute inflammatory phenotype is implicated in the pathology of insulin resistance (IR) and type 2 diabetes mellitus. Interleukin (IL)-1α and IL-1ß are produced by innate immune cells, including macrophages, and mediate their inflammatory response through the IL-1 type I receptor (IL-IRI). This study sought to understand the transcriptomic signature of adipose tissue in obese IL-1RI(-/-) mice. Following dietary intervention, markers of insulin sensitivity and inflammation in adipose tissue were determined, and gene expression was assessed with microarrays. IL-1RI(-/-) mice fed a high-fat diet (HFD) had significantly lower plasma inflammatory cytokine concentrations than wild-type mice. Metabolic network analysis of transcriptomic effects identified up-regulation and co-expression of genes involved in lipolysis, lipogenesis and tricarboxylic acid (TCA) cycle. Further assessment of gene expression in a network of protein interactions related to innate immunity highlighted Stat3 as a potential transcriptional regulator of IL-1 signalling. The complex, downstream effects of IL-1 signalling through the IL-1RI receptor remain poorly defined. Using network-based analyses of transcriptomic signatures in IL-1RI(-/-) mice, we have identified expression changes in genes involved in lipid cycling and TCA cycle, which may be more broadly indicative of a restoration of mitochondrial function in the context of HFD. Our results also highlight a potential role for Stat3 in linking IL-1 signalling to adipogenesis and IR.

Colon cancer associated genes exhibit signatures of positive selection at functionally significant positions.

BACKGROUND: Cancer, much like most human disease, is routinely studied by utilizing model organisms. Of these model organisms, mice are often dominant. However, our assumptions of functional equivalence fail to consider the opportunity for divergence conferred by ~180 Million Years (MY) of independent evolution between these species. For a given set of human disease related genes, it is therefore important to determine if functional equivalency has been retained between species. In this study we test the hypothesis that cancer associated genes have different patterns of substitution akin to adaptive evolution in different mammal lineages. RESULTS: Our analysis of the current literature and colon cancer databases identified 22 genes exhibiting colon cancer associated germline mutations. We identified orthologs for these 22 genes across a set of high coverage (>6X) vertebrate genomes. Analysis of these orthologous datasets revealed significant levels of positive selection. Evidence of lineage-specific positive selection was identified in 14 genes in both ancestral and extant lineages. Lineage-specific positive selection was detected in the ancestral Euarchontoglires and Hominidae lineages for STK11, in the ancestral primate lineage for CDH1, in the ancestral Murinae lineage for both SDHC and MSH6 genes and the ancestral Muridae lineage for TSC1. CONCLUSION: Identifying positive selection in the Primate, Hominidae, Muridae and Murinae lineages suggests an ancestral functional shift in these genes between the rodent and primate lineages. Analyses such as this, combining evolutionary theory and predictions - along with medically relevant data, can thus provide us with important clues for modeling human diseases.

Aflatoxins B(1), B(2) and G(1) modulate cytokine secretion and cell surface marker expression in J774A.1 murine macrophages.

Aflatoxins are fungal products which occur in food and feed. They are potent hepatocarcinogens, and are known to cause immunosuppression. We investigated the effect of aflatoxin B(1) (AFB(1)), aflatoxin B(2) (AFB(2)) and aflatoxin G(1) (AFG(1)) exposure, alone and in combination, on the secretion of key pro- and anti-inflammatory cytokines from the murine macrophage cell line, J774A.1. Exposure of macrophages to low doses of aflatoxin (0.01 or 0.1ng/mL) resulted in a statistically significant change in the secretion of a number of cytokines following stimulation with lipopolysaccharide (LPS), a component of Gram-negative bacterial cell walls. Specifically, treatment with AFB(1) or AFB(2) alone significantly decreased (P<0.01) the secretion of the anti-inflammatory cytokine interleukin (IL) 10 (IL-10), while the secretion of the pro-inflammatory cytokine IL-6 was significantly increased (P<0.01). In addition, aflatoxin exposure affected expression levels of key cell surface markers involved in the inflammatory response. Toll-like receptor 2 (TLR2) and Cluster of Differentiation 14 (CD14) expression levels decreased significantly (P<0.01), but Toll-like receptor 4 (TLR4) expression was unaffected. This data provides further insight into the mechanisms by which aflatoxins modulate the host immune response to exert their immunosuppressive activity.

Omega-3 fatty acids attenuate dendritic cell function via NF-κB independent of PPARγ.

Long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) have been shown to modulate the immune response and have therapeutic effects in inflammatory disorders. PUFA are also peroxisome proliferators-activator receptor-gamma (PPARγ) ligands; a family of ligand-activated transcription factors, which when activated antagonise the pro-inflammatory capability of nuclear factor κB (NF-κB). PPARγ plays a role in dendritic cell (DC) maturation and n-3 PUFA have been shown to affect DC maturation by decreasing activation of NF-κB. While n-3 PUFA can function as PPAR ligands, it is not known whether the NF-κB-mediated immunomodulatory properties of n-3 PUFA are PPARγ-dependent. In this study we examined whether the immunomodulatory effects of n-3 PUFA on DC activation were mediated through activation of PPARγ. Treatment of murine bone marrow derived DCs with docosahexaenoic acid (DHA; 25 µM) and eicosapentaenoic acid (EPA; 25 µM) attenuated LPS-induced DC maturation. This was characterised by suppression of IL-12 production and expression of CD40, CD80, CD86 and MHC II and enhanced production of IL-10 and expression of IL-10R. This was coincident with enhanced PPARγ expression, suppressed NF-κB activity and increased the physical interaction and cellular colocalization between NF-κB with PPARγ. To understand the functional implication of the physical association of PPARγ with NF-κB, we determined whether the specific PPARγ inhibitor, GW9662 could abolish the anti-inflammatory effect of n-3 PUFA Inhibiting PPARγ did not impede the NF-κB-mediated anti-inflammatory cytokine profile induced by EPA and DHA alone. Thus n-3 PUFA activate PPARγ and interact with NF-κB in DC. However, the anti-inflammatory effects of EPA and DHA on DCs are independent of PPARγ.

Anti-inflammatory effects of EPA and DHA are dependent upon time and dose-response elements associated with LPS stimulation in THP-1-derived macrophages.

The long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFA) of fish oil, eicosapentanoic (EPA) and docosahexanoic (DHA) acids are considered cardioprotective. Inflammation elicited by macrophages is increasingly recognised in the aetiology of metabolic syndrome. This study investigated the differential anti-inflammatory potential of EPA and DHA through cytokine production and nuclear factor (NF)-kappaB signalling in a human macrophage model. We investigated the dependency of LC n-3 PUFA immune-modulation on concentration and duration of lipopolysaccharide (LPS) activation. Interleukin (IL)-1beta, IL-6 and tumor necrosis factor-alpha secretion from EPA, DHA and control cells were differentially limited by LPS concentration. In all cases, there was no benefit in activating cells with >0.1 microg/ml LPS. LC n-3 PUFA decreased proinflammatory cytokines production, an effect modulated by LPS concentration. Expression of the transcription factor NF-kappaB p65 was significantly reduced in the nucleus and retained in the cytoplasm of EPA- and/or DHA-treated macrophages during 5-h activation with 0.1 microg/ml LPS. Nuclear binding of p65 was significantly reduced in EPA- and DHA-treated cells at 2-h LPS activation. Over the time course, expression of nuclear IkappaBalpha was significantly reduced, cytoplasmic NF-kappaB p50 significantly increased and cytoplasmic cleaving enzyme IkappaB inhibitor complex significantly reduced in LC n-3 PUFA-treated cells. EPA and DHA down-regulated the production of proinflammatory cytokines associated with the aetiology of metabolic syndrome, NF-kappaB transcriptional activity and upstream cytoplasmic signalling events. Immune responses are dynamic, and the present study suggests a nutrient sensitive window of LPS activation at which EPA and DHA are strongly anti-inflammatory.

A conjugated linoleic acid-enriched beef diet attenuates lipopolysaccharide-induced inflammation in mice in part through PPARgamma-mediated suppression of toll-like receptor 4.

Conjugated linoleic acid (CLA) is a PUFA found in beef and dairy products that has immunoregulatory properties. The level of CLA in beef can be enhanced by feeding cattle fresh grass rather than concentrates. This study determined the effect of feeding a high-CLA beef diet on inflammation in an in vivo model of septic shock. Mice were fed a high-CLA beef (4.3% total fatty acid composition) or low-CLA beef diet (0.84% total fatty acid composition) for 6 wk. Lipopolysaccharide (LPS; 3 microg) or sterile PBS was injected i.v. and serum was harvested 6 h after injection. Serum interleukin (IL)-1beta, IL-12p70, IL-12p40, and interferon-gamma concentrations were significantly reduced in response to the LPS challenge in the high-CLA beef diet group. Bone marrow-derived dendritic cells (BMDC) from the high-CLA beef diet group had significantly less IL-12 and more IL-10 in response to ex vivo LPS stimulation. Furthermore, toll-like receptor 4 (TLR4) and CD14 protein and mRNA expression on BMDC was significantly attenuated in the high-CLA compared with the low-CLA beef diet group. Complimentary in vitro experiments to determine the specificity of the effect showed that synthetic cis9, trans11-CLA suppressed surface expression of CD14 and TLR4 on BMDC. Treatment with the PPARgamma inhibitor GW9662 partially reversed TLR4 expression in immature BMDC. The results of this study demonstrate that feeding a diet enriched in high-beef CLA exerts profound antiinflammatory effects in vivo within the context of LPS-induced sepsis. In addition, downregulation of BMDC TLR4 is mediated through induction of PPARgamma.

Attenuation of inflammation and cellular stress-related pathways maintains insulin sensitivity in obese type I interleukin-1 receptor knockout mice on a high-fat diet.

The development of insulin resistance in the obese is associated with chronic, low-grade inflammation. We aimed to identify novel links between obesity, insulin resistance and the inflammatory response by comparing C57BL/6 with type I interleukin-1 receptor knockout (IL-1RI(-/-)) mice, which are protected against diet-induced insulin resistance. Mice were fed a high-fat diet for 16 wk. Insulin sensitivity was measured and proteomic analysis was performed on adipose, hepatic and skeletal muscle tissues. Despite an equal weight gain, IL-1RI(-/-) mice had lower plasma glucose, insulin and triacylglycerol concentrations, compared with controls, following dietary treatment. The higher insulin sensitivity in IL-1RI(-/-) mice was associated with down-regulation of antioxidant proteins and proteasomes in adipose tissue and hepatic soluble epoxide hydrolase, consistent with a compromised inflammatory response as well as increased glycolysis and decreased fatty acid beta-oxidation in their muscle. Their lower hepatic triacylglycerol concentrations may reflect decreased flux of free fatty acids to the liver, decreased hepatic fatty acid-binding protein expression and decreased lipogenesis. Correlation analysis revealed down-regulation of classical biomarkers of ER stress in their adipose tissue, suggesting that disruption of the IL-1RI-mediated inflammatory response may attenuate cellular stress, which was associated with significant protection from diet-induced insulin resistance, independent of obesity.