Bovine casein peptides co‐stimulate naive macrophages with lipopolysaccharide for proinflammatory cytokine production and nitric oxide release

Three bovine casein peptides, ie LLY, PGPIPN and TTMPLW, were used to investigate their effects on cytokine (TNF‐α and IL‐6) production and nitric oxide (NO) release by murine bone marrow macrophages (BMMs). The results showed that these peptides alone were incapable of stimulating cytokine production or NO release in naive or IFN‐γ‐primed BMMs. However, when BMMs were co‐incubated with the peptides at a concentration of 1.0 µM and lipopolysaccharide (LPS; 100 ng ml⁻¹), an augmentative effect on TNF‐α, IL‐6 and NO production was observed. Of the three peptides, TTMPLW had the greatest augmentative effect on NO production by LPS‐stimulated BMMs and induced the highest amount of TNF‐α production at a concentration of 1.0 µM . All the peptides at a concentration of 1.0 µM stimulated IL‐6 production by BMMs. TNF‐α was neutralised by anti‐TNF‐α monoclonal antibody and the release of NO was reduced by about 33.3% (p < 0.01). These results demonstrate that bovine casein peptides can co‐stimulate naive macrophages with LPS for proinflammatory cytokine production and NO release and may play a role in host defence against pathogens. © 2000 Society of Chemical Industry     

Heat-killed lactic acid bacteria enhance immunomodulatory potential by skewing the immune response toward Th1 polarization

Abstract: Heat‐killed lactic acid bacteria not only possess immunomodulatory functions but also provide the advantages of longer product shelf life, easier storage, and more convenient transportation. To establish appropriate heat treatments for the industrial preparation of probiotics with immunomodulatory effects, 4 different heat treatments were used to kill 11 strains of lactic acid bacteria. Comparisons among the strains and with viable forms were carried out in terms of immunomodulatory activity and adhesion to Caco‐2 cells. Field‐emission scanning electron microscope (FE‐SEM) was employed to observe morphological changes in bacteria after heating. Among the 11 viable strains, Lactobacillus gasseri AI‐88 was the strongest inducer of interferon‐gamma (IFN)‐γ and interleukin (IL)‐12p70 production. However, after heat treatments its stimulatory ability was attenuated. Heat‐killed Enterococcus faecalis YM‐73 and Lactobacillus salivarius AP‐32 strains showed enhanced stimulation of IFN‐γ and IL‐12p70 secretion and coincidental decrease in IL‐13 production. The adhesion of lactic acid bacteria to Caco‐2 cells decreased with increases in temperature. However, heat exposure did not influence immunomodulatory activity. With rising temperature, roughness and unevenness of bacterial cell surfaces increased significantly. The results indicated that heat‐killed E. faecalis YM‐73 and L. salivarius AP‐32 have immunomodulatory ability via increased Th1‐associated cytokines and reduced Th2‐associated cytokines, switching the immune response from a Th2 toward a Th1 response. These 2 heat‐killed strains have the potential for development as commercial products.     

Partially hydrolyzed whey proteins prevent clinical symptoms in a cow's milk allergy mouse model and enhance regulatory T and B cell frequencies

1 Scope

Partially hydrolyzed cow's milk proteins are used to prevent cow's milk allergy in children. Here we studied the immunomodulatory mechanisms of partial cow's milk hydrolysates in vivo.

2 Methods and results

Mice were sensitized with whey or partially hydrolyzed whey using cholera toxin. Whey‐specific IgE levels were measured to determine sensitization and immune cell populations from spleen, mesenteric lymph nodes and Peyer's patches after oral whey administration were measured by flowcytometry. Whey‐specific IgE and IgG1 levels in partial whey hydrolysate sensitized animals were enhanced, but challenge did not induce clinical symptoms. This immunomodulatory effect of partial whey hydrolysate was associated with increased regulatory B and T cells in the spleen, together with a prevention of IgM‐IgA class switching in the mesenteric lymph nodes and an increased Th1 and activated Th17 in the Peyer's patches.

3 Conclusion

Partial hydrolysate sensitization did not induce whey‐induced clinical symptoms, even though sensitization was established. Increased regulatory cell populations in the systemic immune system and a prevention of increased total Th1 and activated Th17 in the intestinal immune organs could contribute to the suppression of allergic symptoms. This knowledge is important for a better understanding of the beneficial effects of hydrolysates.     

Probiotic fermented beverages based on acid whey

Acid whey is a byproduct of cheesemaking that is difficult to use because of its low pH and less-favorable processing properties compared with rennet whey. The aim of this study was to evaluate the qualities of fermented beverages made using acid whey. In manufacturing the beverages, we used probiotic cultures Lactobacillus acidophilus LA-5 or Bifidobacterium animalis ssp. lactis BB-12. The production process included combining pasteurized acid whey with UHT milk, unsweetened condensed milk, or skim milk powder. We introduced milk to enrich casein content and obtain a product with characteristics similar to that of fermented milk drinks. The products were stored under refrigerated conditions (5 ± 1°C) for 21 d. During storage, we assessed the beverages' physicochemical properties and organoleptic characteristics. The properties of the beverages depended on their composition, microbial culture, and storage time. Beverages containing L. acidophilus had higher acidity, which increased during storage; the acidity of samples containing B. animalis was more stable. Beverages made with skim milk powder (La1 and Bb1) had higher acetaldehyde content, but this parameter decreased in all samples during storage. The hardness of the samples did not change during storage and was highest in beverage La3, made from whey, condensed milk, and L. acidophilus. Beverage La2, made from whey, milk, condensed milk, and L. acidophilus, had the best sensory properties. The whey beverages we developed provided a good medium for the probiotic bacteria; bacteria count throughout the storage period exceeded 8 log cfu/mL, distinctly higher than the minimum therapeutic dose.     

Cheese whey fermented with kefir micro‐organisms: Antagonism against Salmonella and immunomodulatory capacity

The antagonistic effect against Salmonella spp. and the immunomodulatory capacity of whey fermented with kefir grains and with three strains isolated from them – Lactobacillus plantarum CIDCA 8327, Lactobacillus kefiri CIDCA 8348 and Kluyveromyces marxianus var. marxianus CIDCA 8154 – were evaluated. Both fermented products interfered with the capacity of Salmonella spp. to associate and invade Caco‐2/TC7 cells and reduced up to the basal level the expression of the CCL20 in response to a pro‐inflammatory stimulus on Caco‐2 CCL20:luc cells. The products with potential application as probiotics obtained by fermentation of whey with kefir micro‐organisms represent an alternative to increase whey economic value.     

Immunomodulatory activities and antioxidant properties of polysaccharides from Monascus-fermented products in vitro

BACKGROUND: Monascus ‐fermented products have featured in Chinese cuisine for thousands of years and are widely used as food colourants and dietary materials in many Asian countries. Rice and dioscorea fermented with Monascus purpureus NTU 568 have health‐promoting attributes in vitro and in vivo . The aim of this study was to investigate the immunomodulatory and antioxidant effects of polysaccharides from red mould rice (RMRP) and red mould dioscorea (RMDP) in Raw 264.7 cells. RESULTS: The results showed the antioxidant capabilities (including scavenging, chelating, inhibition of lipid peroxidation, and reducing power) of RMRP and RMDP at a concentration of 10 mg mL^?1. RMRP and RMDP also stimulated cell proliferation, nitric oxide production, phagocytosis and cytokine production (including IL1‐β, IL‐6 and TNF‐α) in Raw 264.7 cells. CONCLUSION: These findings demonstrate that RMRP and RMDP have antioxidant and immunomodulation potential to be developed as novel dietary supplements. Copyright © 2011 Society of Chemical Industry     

Antioxidative and Anti-Inflammatory Neuroprotective Effects of Astaxanthin and Canthaxanthin in Nerve Growth Factor Differentiated PC12 Cells

ABSTRACT: Nerve growth factor differentiated PC12 cells were used to examine the antioxidative and anti‐inflammatory effects of astaxanthin (AX) and canthaxanthin (CX). PC12 cells were pretreated with AX or CX at 10 or 20 μM, and followed by exposure of hydrogen peroxide (H₂O₂) or 1‐methyl‐4‐phenylpyridinium ion (MPP⁺) to induce cell injury. H₂O₂ or MPP⁺ treatment significantly decreased cell viability, increased lactate dehydrogenase (LDH) release, enhanced DNA fragmentation, and lowered mitochondrial membrane potential (MMP) (P < 0.05). The pretreatments from AX or CX concentration‐dependently alleviated H₂O₂ or MPP⁺‐induced cell death, LDH release, DNA fragmentation, and MMP reduction (P < 0.05). Either H₂O₂ or MPP⁺ treatment significantly increased malonyldialdehyde (MDA) and reactive oxygen species (ROS) formations, decreased glutathione content, and lowered glutathione peroxidase (GPX) and catalase activities (P < 0.05). The pretreatments from AX or CX significantly retained GPX and catalase activities, and decreased MDA and ROS formations (P < 0.05). H₂O₂ or MPP⁺ treatment significantly decreased Na⁺‐K⁺‐ATPase activity, elevated caspase‐3 activity and levels of interleukin (IL)‐1, IL‐6, and tumor necrosis factor (TNF)‐α (P < 0.05); and the pretreatments from these agents significantly restored Na⁺‐K⁺‐ATPase activity, suppressed caspase‐3 activity and release of IL‐1, IL‐6, and TNF‐α (P < 0.05). Based on the observed antioxidative and anti‐inflammatory protection from AX and CX, these 2 compounds were potent agents against neurodegenerative disorder.     

Anti-inflammatory activities of aqueous extract of Mesona procumbens in experimental mice

BACKGROUND: Mesona procumbens is consumed as a herbal drink and jelly‐type dessert in Taiwan. The aim of this study was to determine the mechanism of anti‐inflammatory activities of the aqueous extract of M. procumbens (AMP) using the λ‐carrageenin (Carr)‐induced mouse paw oedema model. The fingerprint chromatogram of AMP was obtained by high‐performance liquid chromatography (HPLC) analysis. To investigate the anti‐inflammatory mechanism of AMP, the activities of catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GPx) and the level of malondialdehyde (MDA) in paw oedema were monitored. Serum nitric oxide (NO), tumour necrosis factor‐α (TNF‐α) and interleukin‐1β (IL‐1β) were also evaluated. RESULTS: The fingerprint chromatogram from HPLC indicated that AMP contained protocatechuic acid, chlorogenic acid, vanillic acid and caffeic acid. In the anti‐inflammatory test, AMP decreased paw oedema after Carr administration and increased the CAT, SOD and GPx activities and decreased the MDA level in paw oedema at 5 h after Carr injection. AMP also affected the serum NO, TNF‐α and IL‐1β levels at 5 h after Carr injection. Western blotting revealed that AMP decreased the expression of Carr‐induced inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2). CONCLUSION: Mesona procumbens has the potential to provide a therapeutic approach to inflammation‐associated disorders. Copyright © 2011 Society of Chemical Industry     

Effect of milk fat replacement by polyunsaturated fatty acids on the microbiological,rheological and sensorial properties of fermented milks

A modified milk (W3DD) where fat had been replaced by oils enriched in ω‐3 polyunsaturated fatty acids was used for the manufacture of a set‐type fermented product. In order to improve the organoleptic properties of the product, 30 g l^?1 whey protein concentrate (WPC) was added during the manufacturing process. Samples were fermented employing a commercial probiotic starter culture (ABT‐2), which contained Streptococcus thermophilus ST‐20Y, Lactobacillus acidophilus LA‐5 and Bifidobacterium lactis BB‐12. The acidification process was dependent on the WPC addition, which favoured the increase of viable counts, but fermentation was not influenced by the milk fat composition. The highest counts of the probiotic strains, L acidophilus LA‐5 (3.3 × 10⁵ cfu g^?1) and B lactis BB‐12 (5.5 × 10⁷ cfu g^?1), after 21 days of storage at 4 °C, were found in fermented products derived from W3DD supplemented with WPC. Addition of WPC also increased the firmness of the products and reduced syneresis. No apparent colour changes due to fat composition or WPC supplementation were observed in the products. Milk fat replacement by oils rich in ω‐3 polyunsaturated fatty acids had a negative influence on the product texture but did not affect the typical yoghurt flavour. These defects were overcome by the addition of 30 g l^?1 WPC, which improved the appearance, texture and general acceptability scores in the product. Copyright © 2004 Society of Chemical Industry     

The inhibition of the viability of myeloma cells and the production of cytokine by two strains of Lactobacillus sakei from meat

Ten strains of lactic acid bacteria were isolated from a dry fermented sausage and tested for stimulation or inhibition of the viability of Vero and myeloma cells. They did not significantly affect the viability of Vero cells but two isolates (CBL/H and CBL/K) showed a strong and moderate inhibition of the myeloma cell viability (at 10⁸ CFU mL^?1, 17.6 and 33.2%, respectively, survival of myeloma cells). The isolates were identified as Lactobacillus sakei by DNA sequencing of the 16S rRNA products of a polymerase chain reaction. No protective effect was found, at the concentrations used, against cytotoxicity of N‐nitrosamines. To test the effect of L. sakei CBL/H and CBL/K on cytokine production [tumour necrosis factor alpha (TNF‐α), interleukin‐1β (IL‐1β) and interleukin‐8 (IL‐8)], the human macrophage cell line (THP‐1) was cultured in the presence and absence of lipopolysaccharide (LPS). Lactobacillus sakei CBL/H and CBL/K induced IL‐1β and IL‐8 release when cells were stimulated with and without LPS. However there was TNF‐α release only in the presence of the LPS.     

Effect of bovine colostrum on the growth inhibition and differentiation of human leukemic U937 cells

BACKGROUND: Colostrum is the breast milk of female mammals produced within in a short time after giving birth. It is thought to protect neonates from infection as well as to facilitate immune system maturation. In this study the effect of bovine colostrum on the proliferation and differentiation of human leukemic U937 cells was investigated to understand more about its immunomodulatory activity. RESULTS: Human mononuclear cells (MNCs) were stimulated with bovine colostrum (CS) and then filtered to obtain a conditioned medium (CM) (CS‐MNC‐CM). CS‐MNC‐CMs prepared with day 1 to day 4 colostrums inhibited U937 cells by 39.4–64.4%. The expressions of surface markers CD11b and CD14 on U937 cells in the treated groups were 30.6–33.5% and 40.0–42.6% respectively. High levels of cytokines IL‐1β, IFN‐γ and TNF‐α were detected in CS‐MNC‐CMs. CONCLUSION: Evidence indicates that colostrums stimulate human MNCs to secrete cytokines IL‐1β, IFN‐γ and TNF‐α which subsequently inhibit the growth of U937 cells and induce their differentiation into mature monocytes and macrophages. There is a possibility for bovine colostrum to be processed into an anti‐leukemia ingredient for use in health foods. Copyright © 2009 Society of Chemical Industry     

Conjugated linoleic acid content in fermented goat milk as affected by the starter culture and the presence of free linoleic acid

The effect of yoghurt starter cultures and probiotic Lactobacillus casei on the formation of conjugated linoleic acid (CLA) and microbial populations of fermented goat milk was investigated during 35 days of cold storage. The addition of hydrolysed sunflower oil as a source of free linoleic acid was investigated. The fermentation process enhanced the content of the cis9, trans11‐CLA isomer in milk, whereas the trans10, cis12‐CLA isomer was not detected in goat milk or control fermented milks. The use of both starters generated trans10, cis12‐CLA only when hydrolysed sunflower oil was supplemented. Populations of streptococci and lactobacilli were affected by the presence of hydrolysed sunflower oil.     

EFFECTS OF CONJUGATED LINOLEIC ACID ISOMERS ON SERUM TUMOR NECROSIS FACTOR-A CONCENTRATION IN MICE

In previously studies, we showed that tumor necrosis factor‐α (TNF‐α), a cytokine involved in a variety of biologically important events, is partially involved in the effects of conjugated linoleic acid (CLA). In this report, we further tested the effects of individual CLA isomers on serum TNF‐α concentration along with other biological markers in mice. The animals were fed experimental diets (control, 0.5% CLA‐mixed isomer, 0.25% cis‐9,trans‐11 CLA or 0.25% trans‐10,cis‐12 CLA) for 5 days and were then challenged with endotoxin. Both cis‐9,trans‐11 and trans‐10,cis‐12 CLA isomers reduced serum TNF‐α levels compared to control. CLA had no effect on the other biological markers examined. These results suggest possible early involvement of CLA in immune and/or inflammatory responses, followed by reduction of body fat. Its effect on TNF‐α helps explain in part how CLA modulates other biological functions such as immune response, insulin responses, atherosclerosis and cancer.     

Anti‐glycative and anti‐inflammatory effects of caffeic acid and ellagic acid in kidney of diabetic mice

Protective effects of caffeic acid (CA) and ellagic acid (EA) in kidney of diabetic mice were examined. CA or EA at 2.5 and 5% was mixed in diet and supplied to diabetic mice for 12 wk. Results showed that the intake of CA or EA increased renal content of these compounds, alleviated body weight loss, decreased urine output, increased plasma insulin and decreased blood glucose levels at weeks 6 and 12 (p<0.05). The intake of these compounds dose dependently reduced plasma blood urea nitrogen and elevated creatinine clearance (p<0.05). CA or EA at 5% significantly decreased the levels of plasma HbA1c, urinary glycated albumin, renal carboxymethyllysine, pentosidine, sorbitol and fructose (p<0.05), and significantly diminished renal activity of aldose reductase and sorbitol dehydrogenase, as well as suppressed renal aldose reductase mRNA expression (p<0.05). CA or EA dose dependently lowered renal levels of IL‐6, IL‐1β, tumor necrosis factor (TNF)‐α and monocyte chemoattractant protein 1 (MCP‐1) (p<0.05). Furthermore, CA or EA dose dependently down‐regulated tumor necrosis factor‐α and monocyte chemoattractant protein‐1 mRNA expression in kidney (p<0.05). Based on the observed anti‐glycative and anti‐inflammatory effects, the supplement of CA or EA might be helpful for the prevention or attenuation of diabetic kidney diseases.     

Development of a fermented goats' milk containing Lactobacillus rhamnosus: in vivo study of health benefits

BACKGROUND: Lactobacillus rhamnosus CRL1505, a strain of goats' milk origin, is able to stimulate mucosal immunity and protect immunocompetent mice from intestinal and respiratory infections. RESULTS: In this work we developed and characterized a fermented goats' milk containing L. rhamnosus CRL1505, and we demonstrated in a model of immunosuppression in mice that the final dairy product preserves the immunomodulatory properties of the strain. L. rhamnosus CRL1505 survived the manufacturing process of fermented milk and maintained a viability of 10⁶ cfu g^?1 during storage. The fermented goats' milk was accepted by 90.48% of the panelists and was considered as having an acid taste and pleasant aroma. We also demonstrated that the developed product, used as a supplement during the repletion of immunocompromised malnourished mice, was effective in accelerating the recovery of clinical parameters altered by malnutrition and to induce increased resistance against intestinal and respiratory infections. CONCLUSION: Goats' milk fermented with L. rhamnosus CRL1505 could be manufactured as an alternative probiotic dairy product since this new food has the ability to stimulate the common mucosal immune system and to improve defenses against respiratory and intestinal infections. Copyright © 2011 Society of Chemical Industry     

Effects of Lactobacillus kefiranofaciens M1 isolated from kefir grains on enterohemorrhagic Escherichia coli infection using mouse and intestinal cell models

A potential probiotic strain, Lactobacillus kefiranofaciens M1, was previously isolated from kefir grains, which are used to manufacture the traditional fermented drink kefir. The aim of this study was to investigate the effects of Lb. kefiranofaciens M1 on enterohemorrhagic Escherichia coli (EHEC) infection, using mice and intestinal cell models. BALB/c mice were daily administrated with either phosphate buffered saline or Lb. kefiranofaciens M1 at 2 × 10⁸ cfu/mouse per day intragastrically for 7 d. Intragastric challenges with EHEC (2 × 10⁹ cfu/mouse) were conducted on d 0, 4, and 7 after treatment. Administration of Lb. kefiranofaciens M1 was able to prevent EHEC infection-induced symptoms, intestinal damage, renal damage, bacterial translocation, and Shiga toxin penetration. Furthermore, the mucosal EHEC-specific IgA responses were increased after Lb. kefiranofaciens M1 administration in the EHEC-infected mouse system. Additionally, in vitro, Lb. kefiranofaciens M1 was shown to have a protective effect on Caco-2 intestinal epithelial cells and Caco-2 intestinal epithelial cell monolayers; the bacteria limited EHEC-induced cell death and reduced the loss of epithelial integrity. These findings support the potential of Lb. kefiranofaciens M1 treatment as an approach to preventing EHEC infection and its effects.     

Short communication: Physicochemical features and microbial community of milk kefir using a potential probiotic Saccharomyces cerevisiae KU200284

The aim of this study was to analyze the β-glucan contents, physicochemical features, and microbial communities in milk kefir prepared using Saccharomyces cerevisiae KU200284 isolated from cucumber jangajji, a fermented vegetable commonly eaten in Korean. Three types of milk kefir were manufactured, with (1) activated kefir grain, (2) activated kefir grain with commercial S. cerevisiae BOF, and (3) activated kefir grain with S. cerevisiae KU200284. β-Glucan contents of milk kefir using kefir grain and kefir grain with S. cerevisiae strains BOF and KU200284 were 8.29, 8.59, and 8.57%, respectively. The pH, titratable acidity, viscosity, Brix level, and alcohol contents of milk kefir using kefir grain with S. cerevisiae strains were acceptable compared with milk kefir using only kefir grain. In milk kefir produced using kefir grains and S. cerevisiae strains, 16S rRNA reads showed representative strains of Lactobacillus kefiranofaciens (>72% relative abundance) and Acetobacter fabarum (>16% relative abundance). In particular, milk kefir using kefir grain with S. cerevisiae KU200284 had the highest relative abundance of L. kefiranofaciens. In addition, the internal transcribed sequence (ITS) rRNA reads in tested milk kefir showed representative strains of Kluyveromyces marxianus (>52% relative abundance) and Saccharomyces cerevisiae (>16% relative abundance). In contrast, milk kefir using S. cerevisiae strains had higher relative abundance of S. cerevisiae (>37%). The β-glucan production, physicochemical properties, and microbial community profiling indicate that S. cerevisiae KU200284 could be used in functional dairy products as a starter culture.     

TRANSGLUTAMINASE IN DAIRY PRODUCTS: CHEMISTRY, PHYSICS, APPLICATIONS

Literature on the effects of microbial transglutaminase on various dairy‐based systems is discussed. Beginning with a short synopsis on the development of microbial transglutaminase as a functional tool for modifying foods, the principles of reactions catalyzed by transglutaminase and their structural implications, as well as the mechanisms of formation and cleavage of isopeptide bonds are reviewed. After summarizing the present knowledge on the specificity of microbial transglutaminase towards milk proteins, including reactions determined by individual lysine and glutamine residues, emphasis is placed on the effects of enzymatic cross‐linking on physicochemical properties in foods and, particularly, dairy‐based systems. Discussed are implications of cross‐linking on acidified milk gels including yogurt and effects on single milk protein fractions, with respect to several physicochemical properties including rheology and mechanical properties of these systems, but also syneresis, and emulsification behaviour.     

Effect of Flavourzyme® on Angiotensin‐Converting Enzyme Inhibitory Peptides Formed in Skim Milk and Whey Protein Concentrate during Fermentation by Lactobacillus helveticus

Angiotensin‐converting enzyme inhibitory (ACE‐I) activity as affected by Lactobacillus helveticus strains (881315, 881188, 880474, and 880953), and supplementation with a proteolytic enzyme was studied. Reconstituted skim milk (12% RSM) or whey protein concentrate (4% WPC), with and without Flavourzyme^® (0.14% w/w), were fermented with 4 different L. helveticus strains at 37 °C for 0, 4, 8, and 12 h. Proteolytic and in vitro ACE‐I activities, and growth were significantly affected (P < 0.05) by strains, media, and with enzyme supplementation. RSM supported higher growth and produced higher proteolysis and ACE‐I compared to WPC without enzyme supplementation. The strains L. helveticus 881315 and 881188 were able to increase ACE‐I to >80% after 8 h of fermentation when combined with Flavourzyme^® in RSM compared to the same strains without enzyme supplementation. Supplementation of media by Flavourzyme^® was beneficial in increasing ACE‐I peptides in both media. The best media to release more ACE‐I peptides was RSM with enzyme supplementation. The L. helveticus 881315 outperformed all strains as indicated by highest proteolytic and ACE‐I activities.