Yogurt and Immunity

The Health Benefit of Fermented Milk Products That Contain Lactic Acid Bacteria

Chapter 5

Contents • 5.1 History and Culture Related to Fermented Food.• 5.2 Yogurt Production.• 5.3 The Effects of Fermentation on Milk.• 5.3.1 Effects on Chemistry, Nutrient Content, and Activities of Enzymes.• 5.3.2 Health Benefits of Fermented Milk Consumption.• 5.4 Fermented Milk and the Immune System• 5.4.1 Overview of the Immune System.• 5.4.2 The Immune System of the Gastrointestinal Tract.• 5.5 Effects of Lactic Acid Bacteria on the Gastrointestinal System• 5.5.1 Lactic Acid Bacteria and Mucosal Health• 5.5.2 Lactic Acid Bacteria and Protection Against Enteric Pathogens• 5.5.3 Lactic Acid Bacteria, Nutrient Digestion, and Antigen Uptake• 5.6 Gut-Associated Lymphoid Tissue and the Establishment of Immune Tolerance• 5.7 Intestinal Microflora and Oral Tolerance• 5.8 Cytokines and Fermented Milk• 5.9 Lactic Acid Bacteria and Immune Cell Function• 5.9.1 Innate Immune Responses• 5.9.2 Phagocytic Activity of Macrophages and Granulocytes• 5.9.3 Natural Killer Cell Activity• 5.9.4 Immunostimulating versus Immunosuppressive Effects.• 5.10 Conclusions

• In this chapter we will learn the compositional changes of milk during fermentation, and health effects of consumption of fermented milk in both animals and humans.

5.1 History and culture related to fermented food

The history of fermented milk is long and widely varied according to culture

Abraham- Fecundity and longevity

I the early 1500s King Francis I- goats’ milk

Elie metchnikoff hypothesis in the early 1900s

“ When people have learnt how to cultivate a suitable microflora in the intestine of children as soon as they are weaned from the breast, the normal life may extend to twice”

5.2 YOGURT PRODUCTION

Yogurt is a thick liquid food prepared from cream or milk fermented by added active cultures of bacteria

US FDA- Lactic acid bacteria (LAB) – Lactobacillus bulgaricus (Lb. bulgaricus) and Streptococcus thermophilus (S.thermophilus)

Milk Sugar (Lactose) lactic acid fermentation 40-400C for 4-5 h.

100 to 10,000-fold 109/ml complete coagulation at pH – 4.6 Lactococcus cremoris, Leuconostoc Viscosity or create a ropy texture

Acetaldehyde provide aroma (smell)

Diacetyl by Lc. Diacetylactis and Leuconostoc cremoris for buttery taste Flavor components- Acetoin, Acetone, Lactones and volatile acids by-products of bacterial

metabolism

“Protocooperation” between S. thermophilus (formic acid) and Lb. bulgaricus (AA,peptides)

5.3 THE EFFECTS OF FERMENTATION ON MILK 5.3.1 Effects on Chemistry, Nutrient Content, and Activities of

Enzymes

• Protein (Casein) Lb.bulgaricus peptides , free AA ( valine, histidine, serine & proline)

Pepetidase • Homofermentative one major product is lactic acid

5.3.2 Health benefits of fermented milk consumption

• Numerous reports published of health benefit but mechanism are still being investigated• Small scale studies on diarrhea have shown that probiotics are beneficial

• Probiotics influence intestinal physiology either directly or indirectly through modulation of endogenous ecosystem or Immune system

• survival rate are 20-40% for selective strains main obstacle is gastric acidity ad bile salt • Maximum probiotic effect can be achieved if the organism adhere to the intestinal mucosal

cells • Prebiotic ( food ingredient that stimulate the growth of probiotic)

• studies giving a proof that enable probiotics to be used as treatment of GI disturbances -The use of Saccharomyces boulardii, Lactobacillus, and Enterococcus faecium SF 68 to

prevent or shorten the duration of antibiotic-associated diarrhea

• Yogurt also enhances protective immunity against respiratory tract infections - Mice infected with Streptococcus pneumonia improved many ecological parameters

• Human study by Gluck et al. investigated the effects of a probiotic, fermented milk drink with Lb. GG (ATCC 53103), Bifidobacterium sp B420, Lb. acidophilus 145, and S.thermophilus on nasal colonization with pathogenic bacteria (Staphylococcus aureus, S. pneumoniae, and β-hemolytic streptococci).

5.4 fermented milk and the Immune system5.4.1 Overview of the immune system

5.4.2 The immune system of the gastrointestinal tract• Organ specific lymphoid tissues have unique property not found elsewhere

• GI tract contains a specialized MALT system that also include the lungs called GALT

5.5 Effects of Lactic acid bacteria on the Gastrointestinal system

• Yogurt supplementation to a standard renutrition diet improves the intestinal function of malnourished animals

• Yogurt improves the barrier function of the intestine

• Swiss albino mice fed fresh yogurt for 7 d, then challenged with 20 LD50 of Salmonella tyhpimurium, showed higher concentrations of S. typhimurium- specific IgA in their intestinal fluid

• Experiment with human volunteers vaccination with attenuated S. typhi Ty21a consumed fermented milk containing Lb. acidophilus La1 (now Lb. johnsonii La1) and bifidobacteria (3 × 125 g/d, 107–108 colony forming units [CFU] per gram) and similar study with Lactobacillus GG (ATCC 53103), Lactococcus lactis

5.5.1 Lactic acid bacteria and mucosal health

• Assays of Ig-secreting cells Enzyme-linked immnospot (ELISPOT) and specific antibody-secreting cells (sASC) among circulating lymphocytes showed that the nonspecific immune response was significantly greater in children receiving Lactobacillus GG than in those given a placebo

• a higher rate of rotavirus IgA was observed in infants given an oral rotavirus vaccine together with Lb. casei GG in those who received the vaccine along with a placebo

• In children with Crohn’s disease, a disease often characterized by a relative deficiency in mucosal IgA, administration of Lactobacillus GG (1010 CFU twice daily) for 10 d was associated with a significant increase in cells secreting IgA specific for β-lactoglobulin and casein

5.5.2 Lactic Acid Bacteria and protection against enteric pathogens

• most commonly found LAB in fermented milk products, S. thermophilus and Lb. bulgaricus, are normally isolated from green plant material and milk, respectively

• These bacteria are not highly acid- and bile-resistant, with only 15% surviving the passage through the stomach and about 1% reach the large intestine where they fail to colonize

• But they may exert an effect in vivo due to intracellular enzymes, cell surface antigenic receptors, or metabolites produced during fermentation.

• studies with mice that feeding yogurt resulted in an alteration in the intestinal flora, stimulating the growth of lactobacilli and bifidobacteria Changes such as these in the intestinal microflora are thought to affect the intestinal transit time and may have an impact on nutrient absorption

• Reports suggest us while the colonizing bacteria are low they do have an impact on health

5.5.3 Lactic Acid Bacteria, Nutrient Digestion, and Antigen Uptake

• Important function of LAB is help in the absorption of indigestible food

• Recent study of transcription of broad array of genes and Bacteroides thetaiotaomicron similar result obtained with Escherichia coli or Bifidobacterium infantis

• Study suggests us colonization can play an important role in increasing the efficiency of nutrient absorption and metabolism.

• higher nutrient efficiency has been reported in animals fed a yogurt-based diet as compared to animals fed a milkbased diet

• Experiment with suckling rats given cows milk the absorption rate of degraded HRP was greater in the group receiving cows milk with Lb. casei GG like this many reports suggest LAB can modulate antigen uptake

• Experiment with peripheral blood mononuclear cells (PBMC) suggest Lb. casei GG can digest casein, and possibly other macromolecules, into less antigenic and potentially immunomodulating peptides

5.6 Gut Associated Lymphoid Tissue and The Establishment of Immune Tolerance

• The GALT in addition to acting as a helper in mucosal defense also plays a role in establishment of systemic hyporesponsiveness

• Allergy- Skewing of an Immune response to an allergen towards a Th2-type phenotype and overproduction of IL-4

• “Hygiene hypothesis”- Partial scarcity of microbial stimuli due to increased hygiene

• Several results suggest intestinal microflora of allergic children(coliforms, staphylococcus aureus) differ from non allergic children(bifidobacteria, lactobacilli)

• study using Nc/Nga mice administered Lb. johnsonii NCC533 during the weaning period prevented the development of atopic dermatitis induced by mite antigens

5.7 Intestinal Microflora and Oral Tolerance

• Oral tolerance – IgG unresponsiveness to ovalbumin (OVA)

• The immune response to commensal bacteria is transient and is replaced by tolerance soon after the initial colonization, it has therefore been proposed that, for the intestinal microflora to play a continued role in the protection from atopic diseases, a high turnover of bacterial genera, species, and strains may be required

• Supplementation with LAB may provide such renewed stimuli for the maintenance of predominant Th1-type responses

• In human in vitro study using three LAB strains (Lb. gasseri ATCC no. 19992, Lb. johnsonii ATCC no. 33200, and Lb. reuteri ATCC no. 23272) indicated a shift towards a Th1-type immune response

• Morita et al. study have demonstrated that oral administration of fermented milk with Lb. gasseri TMC0356 for 4 weeks significantly lowered serum levels of IgE

• selective LAB strains (Lb. plantarum and Lb. lactis) have been successfully used as a mucosal vaccine in an animal model of allergy.

5.8 Cytokines and Fermented Milk• Effect of fermented milk product on immune system involves the production of

various cytokines

• Many LAB (S. thermophilus, Lb. acidophilus, Lb. bulgaricus, and bifidobacteria) have been reported to induce the synthesis of IFN-γ in vitro

• Experiment with B6C3F1 mice on cytokine expression in spleen, Mesenteric lymph nodes (MSN) or Payers patches yogurt fermented with Lb. bulgaricus and S. thermophilus; in addition, some contained Lb. acidophilus and/or Bifidobacterium sp. The effect depended on the type of yogurt fed.

• IL-4 and IFN-γ cytokines maintain the balance between the populations Th2 and Th1 respectively

• Mouse study preformed by Perdigon et al., studied IL-4 and IFN-γ release by immunohistochemistry author determined in these studies that LAB were able to induce a diversity of responses mediated by the cytokines released

• In a more recent study, the authors also demonstrated that a homologous strain, B. animalis, was able to adhere and interact with mucosal epithelial cells of both small and large intestine

• Several human studies also indicate that yogurt consumption may be associated with an increase in IFN-γ synthesis

5.9 Lactic Acid bacteria and Immune Cell Function• Highly variable results were obtained both in vitro and ex vivo experiment

assessing the effects of LAB on lymphocyte proliferation

• incubation of human PBMC with live Lb. johnsonii or Lb. sakei for 5 d resulted in a strong proliferative response while Homogenates of Lb. rhamnosus GG, B. lactis, Lb. acidophilus, Lb. delbrueckii ssp. bulgaricus, and S. thermophilus all suppressed the proliferation

• Several studies have reported that the immunomodulatory effects of LAB are mainly on the innate immune response

• Vinderola et al. study Lb. casei CRL 431 interacted with epithelial cells through toll-like receptor 2 (TLR2), a pattern recognition receptor, and this interaction induced IL-6 secretion

5.9.1 Innate Immune Responses

5.9.2 Phagocytic activity of macrophages and Granulocytes

• Mice Peritoneal macrophages phagocytic activity significantly increased when supplemented with milk fermented with Lb. acidophilus, Lb. casei, for 8 d compared to non supplemented mice

• In human study provided fermented milk with 7 × 1010 CFU Lb. acidophilus La1 or 1 × 1010 B. bifidum strain Bb 12 for 3 weeks, there was a highly significant increase in leukocyte phagocytic activity

• From this and other experiment using different strain author concluded that minimal effective dose for modulating granulocyte or monocyte activities was 109 CFU/d

• Numerous LAB have a stimulatory effect on macrophage activities

5.9.3 Natural Killer cell activity

• Natural killer (NK) cells play an important role in inhibiting carcinogenesis and IL-12 is a strong inducer of NK cells

• LAB including LcS have been shown to stimulate IL-12 production

• In mice with Lb. rhamnosus and with human volunteer Daily intake for 3 weeks of B. lactis in low-fat milk was associated with significantly higher NK cytotoxicity

• Perdigon et al. studied the mechanisms by which yogurt was able to inhibit the growth of a chemically induced intestinal tumor. They demonstrated that yogurt can down regulate the inflammatory response

5.9.4 Immunostimulating versus Immunosuppressive Effects

• Production stimulation of TNF−α and IL-1β proinflammatory cytokines suggest that LAB may have proinflammatory activities

• LAB therapy has been examined in several of autoimmune diseases conditions, and to date, no worse condition of inflammation has been observed

• immunosuppressive effect have been shown by certain strains of LAB by inducing IL-10-producing regulatory T cells

• Immunomodulation is depend on the strain and species of LAB thorough assessment of LAB in vitro and in animal models will be important before they can be used to modulate diseases characterized by immune deviation

5.10 Conclusions

• More than a century research has surrounded the association between fermented product yogurt and health and Important finding of the study is direct effect of yogurt on immune system

• Even though human and mice studied a significant amount of research is still required to address the fundamental basis for the mechanisms of yogurt’s biological consequences

• Limits to rapid progress in this area due to equally complexity of immune system and microbial physiology in vivo

• Despite the limitations, the yogurt-immunity connection remains as an exciting and attractive area of research for a variety of disciplines

• new methodology, procedures, microbial strains, and research findings will make stronger the variation in studies present today, and drive the development of improved fermented milk products

Questions ……..?