Lactobacillus Rhamnosus CRL1505 Research Articles

Investigation on the Efficacy of Two Food Supplements Containing a Fixed Combination of Selected Probiotics and β-Glucans or Elderberry Extract for the Immune System: Modulation on Cytokines Expression in Human THP-1 and PBMC

Several herbal and other natural products are used as ingredients in food supplements to strengthen immunity even if, very often, marketed products are proposed without a clear rationale or experimental evidence. In this study, we aimed to investigate the effect on human monocytes (THP-1) and on ex vivo human peripheral blood mononuclear cells (PBMC) of two formulations, one containing Bifidobacterium animalis subsp. lactis Bl-04® with β-glucans (for adults) and one containing Lactobacillus rhamnosus CRL1505 with elderberry extract (for children). We compared formulations with single ingredients, with bacterial lipopolysaccharide (LPS) and the drug pidotimod; cytokines expression level was evaluated testing different concentrations of samples at two exposure times. As expected, LPS caused a non-specific huge upregulation of cytokines expression both in THP-1 and in PBMC, whereas pidotimod mainly upregulated IL-2 in PBMC and IL-8 in THP-1. The two formulations showed a difference between a pro-inflammatory stimulus such as LPS, and also from an immunostimulant drug, such as pidotimod, as they mainly upregulated the expression of IL-6 and IL-10 in PBMC but not in THP-1, in a concentration-dependent mode. Probiotics were shown to play a major role, but β-glucans and elderberry extract exerted a synergistic activity. This work demonstrated that combining selected probiotics with other natural products having immunomodulatory properties is an interesting strategy to develop innovative formulations in the sector of food supplements.

The effectiveness of metabiotics in the treatment of acute respiratory diseases

This article discusses the relevance and prospects of using metabiotics and targeted metabiotics in the complex therapy of acute respiratory diseases. The metabiotics (postbiotics) include bacterial metabolites and/or signaling molecules with a known chemical structure that optimize the composition and functions of the indigenous microbiota, including improving human immunity and metabolism. Bacterial metabolites of Bacillus subtilis SA44 and Lactobacillus rhamnosus CRL1505 strains demonstrate high efficiency in the treatment of patients with respiratory diseases. It is important to use targeted metabiotics, i.e. metabiotics that contribute to the restoration and maintenance of the microbiota of the respiratory organs and human immunity. Targeted metabiotics, as a rule, contain not only bacterial metabolites, but also additional substances with positive effects aimed at optimizing the functioning of a particular organ or system. An example of a targeted metabiotic for the treatment of acute respiratory diseases is a three-component Biocomplete® Bronchobiotic complex, which includes inactivated bacteria of the species Lactobacillus rhamnosus CRL1505 (the metabiotic, is an antagonist of opportunistic and pathogenic microorganisms, promotes immunity), inulin (the prebiotic, serves as nutrition for the indigenous flora) and thyme extract (has expectorant, anti-inflammatory and bactericidal, antioxidant effect).

Inorganic polyphosphate from the immunobiotic Lactobacillus rhamnosus CRL1505 prevents inflammatory response in the respiratory tract.

Lactobacillus (L.) rhamnosus CRL1505 accumulates inorganic polyphosphate (polyP) in its cytoplasm in response to environmental stress. The aim of this study was to evaluate the potential effects of polyP from the immunobiotic CRL1505 on an acute respiratory inflammation murine animal model induced by lipopolysaccharide (LPS).First, the presence of polyP granules in the cytoplasm of CRL1505 strain was evidenced by specific staining. Then, it was demonstrated in the intracellular extracts (ICE) of CRL1505 that polyP chain length is greater than 45 phosphate residues. In addition, the functionality of the genes involved in the polyP metabolism (ppk, ppx1 and ppx2) was corroborated by RT-PCR. Finally, the possible effect of the ICE of CRL1505 strain containing polyP and a synthetic polyP was evaluated in vivo using a murine model of acute lung inflammation. It was observed that the level of cytokines pro-inflammatory (IL-17, IL-6, IL-2, IL-4, INF-γ) in serum was normalized in mice treated with ICE, which would indicate that polyP prevents the local inflammatory response in the respiratory tract. The potential application of ICE from L. rhamnosus CRL1505 as a novel bioproduct for the treatment of respiratory diseases is one of the projections of this work.

Improvement of Myelopoiesis in Cyclophosphamide-Immunosuppressed Mice by Oral Administration of Viable or Non-Viable Lactobacillus Strains.

Myelosuppression is the major dose-limiting toxicity of cancer chemotherapy. There have been many attempts to find new strategies that reduce myelosuppression. The dietary supplementation with lactic acid bacteria (LAB) improved respiratory innate immune response and the resistance against respiratory pathogens in immunosupressed hosts. Although LAB viability is an important factor in achieving optimal protective effects, non-viable LAB are capable of stimulating immunity. In this work, we studied the ability of oral preventive administration of viable and non-viable Lactobacillus rhamnosus CRL1505 or L. plantarum CRL1506 (Lr05, Lr05NV, Lp06V or Lp06NV, respectively) to minimize myelosuppressive and immunosuppressive effects derived from chemotherapy. Cyclophosphamide (Cy) impaired steady-state myelopoiesis in lactobacilli-treated and untreated control mice. Lr05V, Lr05NV and Lp06V treatments were the most effective to induce the early recovery of bone marrow (BM) tissue architecture, leukocytes, myeloid, pool mitotic and post-mitotic, peroxidase positive, and Gr-1Low/High cells in BM. We selected the CRL1505 strain for being the one capable of maintaining its myelopoiesis-enhancing properties in its non-viable form. Although the CRL1505 treatments do not modify the Cy ability to induce apoptosis, both increased the incorporation of BrdU in BM cells. Consequently, Lr05NV and Lr05V treatments were able to promote early recovery of LSK cells (Lin-Sca-1+c-Kit+ cells), multipotent progenitors (Lin-Sca-1+c-Kit+CD34+ cells), and myeloid cells (Gr-1+Ly6G+Ly6C- cells) with respect to the untreated Cy control. In addition, these treatments were able to increase the frequency of IL17A-producing innate lymphoid cells in the intestinal lamina propria (IL-17A+RORγt+CD4-NKp46+ cells) after Cy injection. These results were correlated with an increase in the IL-17A serum levels, a GM-CSF high expression and a CXCL12 lower expression in BM. Therefore, both Lr05V and Lr05NV treatments are able to activate beneficially the IL-17A/GM-CSF axis and accelerate the recovery of Cy-induced immunosuppression by increasing BM myeloid precursors. We demonstrated for the first time the beneficial effect of CRL1505 strain on myelopoiesis affected by a chemotherapeutic drug. Furthermore, Lr05NV could be a good and safe resource for reducing chemotherapy-induced leukopenia. The results are a starting point for future research and open up broad prospects for future applications of the immunobiotics.

The Role of Alveolar Macrophages in the Improved Protection against Respiratory Syncytial Virus and Pneumococcal Superinfection Induced by the Peptidoglycan of Lactobacillus rhamnosus CRL1505.

The nasal priming with nonviable Lactobacillus rhamnosus CRL1505 (NV1505) or its purified peptidoglycan (PG1505) differentially modulates the respiratory innate immune response in infant mice, improving their resistance to primary respiratory syncytial virus (RSV) infection and secondary pneumococcal pneumonia. In association with the protection against RSV-pneumococcal superinfection, it was found that NV1505 or PG1505 significantly enhance the numbers of CD11c+SiglecF+ alveolar macrophages (AMs) producing interferon (IFN)-β. In this work, we aimed to further advance in the characterization of the beneficial effects of NV1505 and PG1505 in the context of a respiratory superinfection by evaluating whether their immunomodulatory properties are dependent on AM functions. Macrophage depletion experiments and a detailed study of their production of cytokines and antiviral factors clearly demonstrated the key role of this immune cell population in the improvement of both the reduction of pathogens loads and the protection against lung tissue damage induced by the immunobiotic CRL1505 strain. Studies at basal conditions during primary RSV or S. pneumoniae infections, as well as during secondary pneumococcal pneumonia, brought the following five notable findings regarding the immunomodulatory effects of NV1505 and PG1505: (a) AMs play a key role in the beneficial modulation of the respiratory innate immune response and protection against RSV infection, (b) AMs are necessary for improved protection against primary and secondary pneumococcal pneumonia, (c) the generation of activated/trained AMs would be essential for the enhanced protection against respiratory pathogens, (d) other immune and nonimmune cell populations in the respiratory tract may contribute to the protection against bacterial and viral infections, and (e) the immunomodulatory properties of NV1505 and PG1505 are strain-specific. These findings significantly improve our knowledge about the immunological mechanisms involved in the modulation of respiratory immunity induced by beneficial microbes.

The Modulation of Mucosal Antiviral Immunity by Immunobiotics: Could They Offer Any Benefit in the SARS-CoV-2 Pandemic?

Viral respiratory infections are of major importance because of their capacity to cause of a high degree of morbidity and mortality in high-risk populations, and to rapidly spread between countries. Perhaps the best example of this global threat is the infectious disease caused by the new SARS-CoV-2 virus, which has infected more than 4 million people worldwide, causing the death of 287,000 persons according to the WHO's situation report on May 13, 2020. The availability of therapeutic tools that would be used massively to prevent or mitigate the detrimental effects of emerging respiratory viruses on human health is therefore mandatory. In this regard, research from the last decade has reported the impact of the intestinal microbiota on the respiratory immunity. It was conclusively demonstrated how the variations in the intestinal microbiota affect the responses of respiratory epithelial cells and antigen presenting cells against respiratory virus attack. Moreover, the selection of specific microbial strains (immunobiotics) with the ability to modulate immunity in distal mucosal sites made possible the generation of nutritional interventions to strengthen respiratory antiviral defenses. In this article, the most important characteristics of the limited information available regarding the immune response against SARS-CoV-2 virus are revised briefly. In addition, this review summarizes the knowledge on the cellular and molecular mechanisms involved in the improvement of respiratory antiviral defenses by beneficial immunobiotic microorganisms such as Lactobacillus rhamnosus CRL1505. The ability of beneficial microorganisms to enhance type I interferons and antiviral factors in the respiratory tract, stimulate Th1 response and antibodies production, and regulate inflammation and coagulation activation during the course of viral infections reducing tissue damage and preserving lung functionally, clearly indicate the potential of immunobiotics to favorably influence the immune response against SARS-CoV-2 virus.

Efficient Selection of New Immunobiotic Strains With Antiviral Effects in Local and Distal Mucosal Sites by Using Porcine Intestinal Epitheliocytes

Previously, we evaluated the effect of the immunobiotic strain Lactobacillus rhamnosus CRL1505 on the transcriptomic response of porcine intestinal epithelial (PIE) cells triggered by the challenge with the Toll-like receptor 3 (TLR-3) agonist poly(I:C) and successfully identified a group of genes that can be used as prospective biomarkers for the screening of new antiviral immunobiotics. In this work, several strains of lactobacilli were evaluated according to their ability to modulate the expression of IFNα, IFNβ, RIG1, TLR3, OAS1, RNASEL, MX2, A20, CXCL5, CCL4, IL-15, SELL, SELE, EPCAM, PTGS2, PTEGES, and PTGER4 in PIE cells after the stimulation with poly(I:C). Comparative analysis of transcripts variations revealed that one of the studied bacteria, Lactobacillus plantarum MPL16, clustered together with the CRL1505 strain, indicating a similar immunomodulatory potential. Two sets of in vivo experiments in Balb/c mice were performed to evaluate L. plantarum MPL16 immunomodulatory activities. Orally administered MPL16 prior intraperitoneal injection of poly(I:C) significantly reduced the levels of the proinflammatory mediators tumor necrosis factor α (TNF-α), interleukin 6 (IL-6), and IL-15 in the intestinal mucosa. In addition, orally administered L. plantarum MPL16 prior nasal stimulation with poly(I:C) or respiratory syncytial virus infection significantly decreased the levels of the biochemical markers of lung tissue damage. In addition, reduced levels of the proinflammatory mediators TNF-α, IL-6, and IL-8 were found in MPL16-treated mice. Improved levels of IFN-β and IFN-γ in the respiratory mucosa were observed in mice treated with L. plantarum MPL16 when compared to control mice. The immunological changes induced by L. plantarum MPL16 were not different from those previously reported for the CRL1505 strain in in vitro and in vivo studies. The results of this work confirm that new immunobiotic strains with the ability of stimulating both local and distal antiviral immune responses can be efficiently selected by evaluating the expression of biomarkers in PIE cells.

Nasal priming with immunobiotic lactobacilli improves the adaptive immune response against influenza virus

The nasal priming with Lactobacillus rhamnosus CRL1505 modulates the respiratory antiviral innate immune response and improves protection against influenza virus (IFV) challenge in mice. However, the potential beneficial effect of the CRL1505 strain on the adaptive immune response triggered by IFV infection or vaccination was not evaluated before. In this work, we demonstrated that nasally administered L. rhamnosus CRL1505 is able to improve both the humoral and cellular adaptive immune responses induced by IFV infection or vaccination. Higher levels of IFV-specific IgA and IgG as well as IFN-γ were found in the serum and the respiratory tract of CRL1505-treated mice after IFV challenge. Lactobacilli treated mice also showed reduced concentrations of IL-17 and improved levels of IL-10 during IFV infection. The differential balance of inflammatory and regulatory cytokines induced by L. rhamnosus CRL1505 contributed to the protection against IFV by favoring an effective effector immune response without inducing inflammatory-mediated lung damage. The optimal immunomodulatory effect of the CRL1505 strain was achieved with viable bacteria. However, non-viable L. rhamnosus CRL1505 was also efficient in improving the adaptive immune responses generated by IFV challenges and therefore, emerged as an interesting alternative for vaccination of immunocompromised hosts. Similar to other immunomodulatory properties of lactobacilli, it was shown here that the adjuvant effect in the context of IFV vaccination was a strain dependent ability, since differences were found when L. rhamnosus CRL1505 and the immunomodulatory strain L. rhamnosus IBL027 were compared. This investigation represents a thorough exploration of the role of immunobiotic lactobacilli in improving humoral and cellular adaptive immune responses against IFV in the context of both infection and vaccination.

Nasal Priming with Lactobacillus rhamnosus CRL1505 Stimulates Mononuclear Phagocytes of Immunocompromised Malnourished Mice: Improvement of Respiratory Immune Response.

The effect of Lactobacillus rhamnosus CRL1505 (Lr) on macrophages (Ma) and dendritic cells (DC) in the orchestration of anti-pneumococcal immunity was studied using malnutrition and pneumococcal infection mouse models. Monocytes (Mo), Ma, and DC in two groups of malnourished mice fed with balanced diet (BCD) were studied through flow cytometry; one group was nasally administered with Lr (BCD+Lr group), and the other group was not (BCD group). Well-nourished (WNC) and malnourished (MNC) mice were used as controls.Malnutrition affected the number of respiratory and splenic mononuclear phagocytes. The BCD+Lr treatment, unlike BCD, was able to increase and normalize lung Mo and Ma. The BCD+Lr mice were also able to upregulate the expression of the activation marker MHC II in lung DC and to improve this population showing a more significant effect on CD11b+ DC subpopulation. At post-infection, lung Mo values were higher in BCD+Lr mice than in BCD mice and similar to those obtained in WNC group. Although both repletion treatments showed similar values of lung Ma post-infection, the Ma activation state in BCD+Lr mice was higher than that in BCD mice. Furthermore, BCD+Lr treatment was able to normalize the number and activation of splenic Ma and DC after the challenge.Lr administration stimulates respiratory and systemic mononuclear phagocytes. Stimulation of Ma and DC populations would increase the microbicide activity and improve the adaptive immunity through its antigen-presenting capacity. Thus, Lr contributes to improved outcomes of pneumococcal infection in immunocompromised hosts.

Screening and Characterization of Immunobiotic Lactic Acid Bacteria with Porcine Immunoassay Systems.

Microorganisms with the ability to modulate the immune system (immunobiotics) have shown to interact with different pattern recognition receptors (PRRs) expressed in nonimmune and immune cells and exert beneficial effects on host's health maintenance and promotion. Suitable assay systems are necessary for an efficient and rapid screening of potential immunobiotic strains. More than a decade of research have allowed us to develop efficient in vitro models based on porcine receptors and cells (porcine immunoassay systems) to study the immunomodulatory effects of lactic acid bacteria (LAB). In addition, detailed studies of model immunobiotic LAB strains with proved abilities to improve immune health in humans (Lactobacillus rhamnosus CRL1505) or pigs (Lactobacillus jensenii TL2937) allowed us to select the most suitable biomarkers that have to be evaluated in those porcine immunoassay systems. Our in vitro models based on transfectant cells expressing porcine PRRs as well as an originally established porcine intestinal epitheliocyte (PIE) cell line have shown to be useful in vitro tools for the selection of immunobiotics and for obtaining information to elucidate the molecular mechanisms behind their beneficial effects.

Nasal immunization with recombinant chimeric pneumococcal protein and cell wall from immunobiotic bacteria improve resistance of infant mice to Streptococcus pneumoniae infection.

Respiratory tract infections and invasive disease caused by Streptococcus pneumoniae in high-risk groups are a major global health problem. Available human vaccines have reduced immunogenicity and low immunological memory in these populations, as well as high cost as a public health strategy in poor communities. In addition, no single pneumococcal protein antigen has been able to elicit protection comparable to that achieved using protein-polysaccharide conjugate vaccines. In this context, chimeric pneumococcal proteins raise as potential good vaccine candidates because of their simplicity of production and reduced cost. The aim of this work was to study whether the nasal immunization of infant mice with the recombinant chimeric pneumococcal protein (PSFP) was able to improve resistance to S. pneumoniae, and whether the immunomodulatory strain Lactobacillus rhamnosus CRL1505 or its cell wall (CW1505) could be used as effective mucosal adjuvants. Our results showed that the nasal immunization with PSPF improved pneumococcal-specific IgA and IgG levels in broncho-alveolar lavage (BAL), reduced lung bacterial counts, and avoided dissemination of pneumococci into the blood. Of interest, immunization with PSPF elicited cross-protective immunity against different pneumococcal serotypes. It was also observed that the nasal immunization of infant mice with PSPF+CW1505 significantly increased the production of pneumococcal-specific IgA and IgG in BAL, as well as IgM and IgG in serum when compared with PSPF alone. PSPF+CW1505 immunization also improved the reduction of pneumococcal lung colonization and its dissemination in to the bloodstream when compared to PSPF alone. Our results suggest that immunization with PSPF together with the cell wall of the immunomodulatory strain L. rhamnosus CRL1505 as a mucosal adjuvant could be an interesting alternative to improve protection against pneumococcal infection in children.

Are the immunomodulatory properties of Lactobacillus rhamnosus CRL1505 peptidoglycan common for all Lactobacilli during respiratory infection in malnourished mice?

Previously, we reported that Lactobacillus rhamnosus CRL1505 peptidoglycan (PG05) improves the innate immune response in immunocompromised-malnourished mice after Streptococcus pneumoniae infection. This study extends those previous findings by demonstrating that the dietary recovery of malnourished mice with nasal administration of PG05 improves not only the innate immune response but the respiratory and systemic adaptive humoral response as well. PG05 enhanced the Th2 response, the recovery of B cells, and the concentration and opsonophagocytic activity of anti-pneumococcal antibodies. In addition, by performing comparative studies with the peptidoglycans from lactobacilli of the same species (L. rhamnosus CRL534) or with similar immunomodulatory properties (L. plantarum CRL1506), we demonstrated here that PG05 has unique immunomodulatory properties that cannot be extended to peptidoglycans from other probiotic strains. However, the knowledge of the molecular characteristics of PG05 is indispensable to understand immunomodulatory abilities of L. rhamnosus CRL1505.

Peptidoglycan from Immunobiotic Lactobacillus rhamnosus Improves Resistance of Infant Mice to Respiratory Syncytial Viral Infection and Secondary Pneumococcal Pneumonia.

Several research works have demonstrated that beneficial microbes with the capacity to modulate the mucosal immune system (immunobiotics) are an interesting alternative to improve the outcome of bacterial and viral respiratory infections. Among the immunobiotic strains with the capacity to beneficially modulate respiratory immunity, Lactobacillus rhamnosus CRL1505 has outstanding properties. Although we have significantly advanced in demonstrating the capacity of L. rhamnosus CRL1505 to improve resistance against respiratory infections as well as in the cellular and molecular mechanisms involved in its beneficial activities, the potential protective ability of this strain or its immunomodulatory cellular fractions in the context of a secondary bacterial pneumonia has not been addressed before. In this work, we demonstrated that the nasal priming with non-viable L. rhamnosus CRL1505 or its purified peptidoglycan differentially modulated the respiratory innate antiviral immune response triggered by toll-like receptor 3 activation in infant mice, improving the resistance to primary respiratory syncytial virus (RSV) infection, and secondary pneumococcal pneumonia. In association with the protection against RSV-pneumococcal superinfection, we found that peptidoglycan from L. rhamnosus CRL1505 significantly improved lung CD3+CD4+IFN-γ+, and CD3+CD4+IL-10+ T cells as well as CD11c+SiglecF+IFN-β+ alveolar macrophages with the consequent increases of IFN-γ, IL-10, and IFN-β in the respiratory tract. Our results also showed that the increase of these three cytokines is necessary to achieve protection against respiratory superinfection since each of them are involved in different aspect of the secondary pneumococcal pneumonia that have to be controlled in order to reduce the severity of the infectious disease: lung pneumococcal colonization, bacteremia, and inflammatory-mediated lung tissue injury.

Lactobacillus rhamnosus CRL1505 nasal administration improves recovery of T-cell mediated immunity against pneumococcal infection in malnourished mice.

Immunobiotic lactic acid bacteria have become an interesting alternative for the prevention of respiratory infections. Previously, we demonstrated that the nasal administration of Lactobacillus rhamnosus CRL1505, during repletion of malnourished mice, resulted in diminished susceptibility to the challenge with the respiratory pathogen Streptococcus pneumoniae. Considering the known alterations induced by malnutrition on T lymphocytes and the importance of this cell population on the protection against respiratory pathogens, we aimed to study the effect of L. rhamnosus CRL1505 nasal administration on the recovery of T cell-mediated defences against pneumococcal infection in malnourished mice under nutritional recovery. Malnourished mice received a balanced conventional diet (BCD) for seven days or BCD for seven days with nasal L. rhamnosus CRL1505 supplementation during last two days of the treatment. After the treatments mice were infected with S. pneumoniae. Flow cytometry studies were carried out in bone marrow, thymus, spleen and lung to study T cells, and Th1/Th2 cytokine profiles were determined in broncho-alveolar lavages and serum. The administration of CRL1505 strain to malnourished mice under recovery reduced quantitative and qualitative alterations of CD4+ T cells in the bone marrow, thymus, spleen and lung induced by malnutrition. In addition, CRL1505 treatment augmented Th2-cytokines (interleukin 10 and 4) in respiratory and systemic compartments after pneumococcal infection. These results show that modulation of CD4+ T lymphocytes induced by L. rhamnosus CRL1505 has an important role in the beneficial effect induced by this strain on the recovery of malnourished mice. These data also indicate that nasally administered L. rhamnosus CRL1505 may represent a non-invasive alternative to modulate and improve the T cell-mediated immunity against respiratory pathogens in immunocompromised malnourished hosts.

Transcriptomic Analysis of the Innate Antiviral Immune Response in Porcine Intestinal Epithelial Cells: Influence of Immunobiotic Lactobacilli.

Lactobacillus rhamnosus CRL1505 and Lactobacillus plantarum CRL1506 are immunobiotic strains able to increase protection against viral intestinal infections as demonstrated in animal models and humans. To gain insight into the host–immunobiotic interaction, the transcriptomic response of porcine intestinal epithelial (PIE) cells to the challenge with viral molecular associated pattern poly(I:C) and the changes in the transcriptomic profile induced by the immunobiotics strains CRL1505 and CRL1506 were investigated in this work. By using microarray technology and reverse transcription PCR, we obtained a global overview of the immune genes involved in the innate antiviral immune response in PIE cells. Stimulation of PIE cells with poly(I:C) significantly increased the expression of IFN-α and IFN-β, several interferon-stimulated genes, cytokines, chemokines, adhesion molecules, and genes involved in prostaglandin biosynthesis. It was also determined that lactobacilli differently modulated immune gene expression in poly(I:C)-challenged PIE cells. Most notable changes were found in antiviral factors (IFN-α, IFN-β, NPLR3, OAS1, OASL, MX2, and RNASEL) and cytokines/chemokines (IL-1β, IL-6, CCL4, CCL5, and CXCL10) that were significantly increased in lactobacilli-treated PIE cells. Immunobiotics reduced the expression of IL-15 and RAE1 genes that mediate poly(I:C) inflammatory damage. In addition, lactobacilli treatments increased the expression PLA2G4A, PTGES, and PTGS2 that are involved in prostaglandin E2 biosynthesis. L. rhamnosus CRL1505 and L. plantarum CRL1506 showed quantitative and qualitative differences in their capacities to modulate the innate antiviral immune response in PIE cells, which would explain the higher capacity of the CRL1505 strain when compared to CRL1506 to protect against viral infection and inflammatory damage in vivo. These results provided valuable information for the deeper understanding of the host–immunobiotic interaction and their effect on antiviral immunity. The comprehensive transcriptomic analyses successfully identified a group of genes (IFN-β, RIG1, RNASEL, MX2, A20, IL27, CXCL5, CCL4, PTGES, and PTGER4), which can be used as prospective biomarkers for the screening of new antiviral immunobiotics in PIE cells and for the development of novel functional food and feeds, which may help to prevent viral infections.

Immunobiotic Lactobacillus strains reduce small intestinal injury induced by intraepithelial lymphocytes after Toll-like receptor 3 activation.

Intestinal intraepithelial lymphocytes (IELs) play critical roles in disrupting epithelial homeostasis after Toll-like receptor (TLR)-3 activation with genomic rotavirus dsRNA or the synthetic dsRNA analog poly(I:C). The capacity of immunobiotic Lactobacillus rhamnosus CRL1505 (Lr1505) or Lactobacillus plantarum CRL1506 (Lp1506) to beneficially modulate IELs response after TLR3 activation was investigated in vivo using a mice model. Intraperitoneal administration of poly(I:C) induced inflammatory-mediated intestinal tissue damage through the increase of inflammatory cells (CD3(+)NK1.1(+), CD3(+)CD8αα(+), CD8αα(+)NKG2D(+)) and pro-inflammatory mediators (TNF-α, IL-1β, IFN-γ, IL-15, RAE1, IL-8). Increased expression of intestinal TLR3, MDA5, and RIG-I was also observed after poly(I:C) challenge. Treatment with Lr1505 or Lp1506 prior to TLR3 activation significantly reduced the levels of TNF-α, IL-15, RAE1, and increased serum and intestinal IL-10. Moreover, CD3(+)NK1.1(+), CD3(+)CD8αα(+), and CD8αα(+)NKG2D(+) cells were lower in lactobacilli-treated mice when compared to controls. The immunomodulatory capacities of lactobacilli allowed a significant reduction of intestinal tissue damage. This work demonstrates the reduction of TLR3-mediated intestinal tissue injury by immunobiotic lactobacilli through the modulation of intraepithelial lymphocytes response. It is a step forward in the understanding of the cellular mechanisms involved in the antiviral capabilities of immunobiotic strains.

Dietary Supplementation with Probiotic Strain Improves Immune-Health in Aged Mice

Ageing is associated with several anatomical and physiological changes of the organism, and the increase in global elderly population promotes the research to develop strategies to improve their quality of life. In this work, we characterized the immunological alterations naturally produced during aging in a mice model, and evaluated the effect of probiotic Lactobacillus (L.) rhamnosus CRL1505 administration on those immunological parameters. We demonstrated that L. rhamosus CRL1505 was able to improve peritoneal macrophages phagocytic activity, and the number of intestinal IgA+ cells in aged mice, reaching values of those parameters similar to young adult mice. The results of this work indicate that is plausible that the immunobiotic CRL1505 strain may find applications as a beneficial immunomodulator in aging to reinforce the intestinal and systemic immunity. The immune modulation in aging induced by L. rhamnosus CRL1505 could lead to the development of new strategies for functional foods specifically tailored for the elderly.

Nasal priming with immunobiotic Lactobacillus rhamnosus modulates inflammation-coagulation interactions and reduces influenza virus-associated pulmonary damage.

To evaluate the effect of the nasal administration of live and heat-killed Lactobacillus rhamnosus CRL1505 (Lr1505) on immune-coagulative response during influenza virus (IFV) infection to improve survival and reduce lung injury. Six-week-old BALB/c mice were treated with live or heat-killed Lr1505 by the nasal route during two consecutive days. Treated and untreated control mice were then nasally challenged with IFV. Both viable and non-viable Lr1505 protected infected mice by reducing pulmonary injury and lung viral loads trough several mechanisms: (a) Inflammatory cytokines were efficiently regulated allowing higher clearance of virus and reduction of inflammatory lung tissue damage, associated to higher levels of the regulatory cytokine IL-10. (b) The antiviral immune response was enhanced with improved levels of type I interferons, CD4(+)IFN-γ(+) lymphocytes, and lung CD11c(+)CD11b(low)CD103(+) and CD11c(+)CD11b(high)CD103(-) dendritic cells. (c) The procoagulant state was reversed mainly by down-regulating tissue factor expression and restoring thrombomodulin levels in lung. The capacity of Lr1505 to improve the outcome of IFV infection would be related to its ability to beneficially modulate lung TLR3-triggered immune response. Our work is the first to demonstrate the ability of an immunobiotic strain to beneficially modulate inflammation-coagulation interactions during IFV infection. Interestingly, non-viable L. rhamnosus CRL1505 was as effective as the viable strain to beneficially modulate respiratory antiviral immune response.

Toll-Like Receptor Activation in Immunity vs. Tolerance

After the discovery of Toll-like receptors (TLR) in the late 1990s, initial investigations were focused on understanding their role, stimulating immune responses against infectious agents. Yet, the human body is home to a myriad of TLR agonistic bacteria that have not only established symbiosis with the immune system but also likely contribute to the induction and maintenance of immune homeostasis by dampening immune responses. In fact, stimulation of TLRs might be critically involved in this process and ultimately contribute to preventing development of inflammatory and autoimmune diseases. Sander De Kivit and colleagues present an overview of those aspects, notably outlining the role of gut epithelial TLRs in the induction of immunity and the maintenance of tolerance (1). In addition, the authors highlight the mechanisms through which the gut microbiota regulates intestinal immune responses through interaction with TLRs. The capacity of probiotic microorganisms to modulate immunity via specific TLRs is further discussed in two articles by Julio Villena and Haruki Kitazawa. The authors explore the role of TLR interaction with immunobiotics for the regulation of intestinal inflammation in pigs, with a focus on TLR4 and Lactobacillus jensenii TL2937 (2). Interaction of probiotics with TLR3 to promote beneficial immunity beyond the gut in the respiratory tract is also examined (3). Specifically, inflammatory and immunoregulatory mechanisms conferred by Lactobacillus rhamnosus CRL1505 are presented along with their proposed effects increasing resistance to RSV infection while limiting immunopathology. While TLRs stimulate and regulate immunity against infectious agents, they are also highly pursued targets for therapy of cancer due to their strong ability to activate multiple arms of the immune system, and in particular to stimulate those specific cellular and cytokine responses critical to anti-tumor immunity. However, the immune potentiating, anti-tumor effects of TLR agonists may be restrained by their parallel ability to trigger immune regulation. Dampening of immune responses may enable control of inflammatory damage or immunopathology, but the downside is a limiting effect on efficacy. The impact of this dichotomy on the use of TLR agonists for immunotherapy of cancer is discussed by Hailing Lu in a review article where key immune regulatory facets of TLR agonists are presented, which may impair their efficacy (4). The dual role of TLRs in cancer likely extends beyond the dichotomy between immune stimulation and regulation. Inflammatory processes triggered by TLR engagement may notably constitute a double-edged sword in elimination vs. development of tumors. In that regard, Erin Burns and Nabiha Yusuf contribute two opinion articles discussing TLR targeting for the treatment of cancer. The use of TLR agonists for skin cancer treatment is presented in the context of their impact on skin carcinogenesis, and the question of TLR tolerance is also discussed (5). Similarly, the dual effect of TLR agonists in breast cancer is examined, where TLR stimulation might beneficially activate the immune system but inflammatory processes may also promote tumor development, while TLR-conferred immune regulation may further curb anti-tumor immunity (6). The mechanisms that underlie the immune regulatory properties of TLRs are not well understood. It is possible that the “default” response induced by TLR stimulation may vary between cell types, or depending on the microenvironment/anatomical location of TLR-expressing cells (as exemplified in gut mucosal tissue). Alexandra Zanin-Zhorov and colleagues described an immune regulatory mechanism conferred by TLR expression on T cells, which the authors review herein and discuss in the context of TLR-induced T cell effector functions (7). The role played by endogenous TLR ligands in conferring immune regulatory mechanisms is also presented. Nobuhiro Nakamoto and Takanori Kanai focus on a key organ, the liver, where various types of TLR-expressing cells are faced with continuous exposure to foreign antigens (8). The authors review immune stimulatory and regulatory effects of TLR signaling that coexist in the liver and influence liver health and disease. Elke Gulden and Li Wen concentrate on another organ, the pancreas, where TLR engagement also ultimately controls health and disease (9). Here, the beneficial and detrimental effects of TLR stimulation on type-1 diabetes are discussed, and notably the authors explore how endogenous TLR agonists can confer immune activation vs. regulation of autoreactive T cells. To close on the question of immune regulation conferred by TLRs, Himanshu Singh Chandel and colleagues contribute an opinion article investigating an alternative immune modulatory aspect that may influence the outcome of parasitic infection (10). The authors review mechanisms that underlie cross-talk between TLRs and CD40, and discuss how this interaction may determine the nature of anti-leishmanial immune responses and ultimately parasite elimination.

Non-viable immunobiotic Lactobacillus rhamnosus CRL1505 and its peptidoglycan improve systemic and respiratory innate immune response during recovery of immunocompromised-malnourished mice.

The effect of non-viable Lactobacillus rhamnosus CRL1505 and its cell wall and peptidoglycan on respiratory immunity in malnourished mice was studied. Weaned mice were malnourished with a protein-free diet for 21d and received BCD during 7d (BCD) or BCD with nasal non-viable L. rhamnosus CRL1505 (BCD+UV) or its cell wall (BCD+CW) or peptidoglycan (BCD+PG) supplementation during last 2d of the treatment. Malnourished mice without treatment (MNC) and well-nourished mice (WNC) were used as controls. Mice were infected nasally with Streptococcus pneumoniae after treatments. Resistance against pneumococci was reduced in MNC mice. Repletion with BCD reduced lung and blood bacterial cell counts when compared to MNC mice but the counts did not reach the levels of the WNC group. However, when malnourished mice received BCD+UV, BCD+CW or BCD+PG, pneumococci was not detected in lung or blood samples. Pneumococcal infection increased the levels of TNF-α, IL-1β, IL-6, and IL-10 in the respiratory tract, however the values were lower in MNC than in WNC mice. BCD+UV and BCD+PG groups showed values of phagocytes, IL-1β and IL-6 that were similar to WNC mice, while TNF-α was significantly higher in those groups when compared to WNC mice. Moreover, BCD+UV and BCD+PG treatments improved levels of respiratory IL-10, reaching values that were superior to those observed in WNC mice. The work demonstrates for the first time that non-viable probiotic bacteria or their cellular fractions could be an interesting alternative as mucosal immunomodulators, especially in immunocompromised hosts in which the use of live bacteria might be dangerous.