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The probiotic Limosilactobacillus reuteri DSM 17938 produces anti-inflammatory effects in scurfy (SF) mice, a model characterized by immune dysregulation, polyendocrinopathy, enteropathy, and X-linked inheritance (called IPEX syndrome in humans), caused by regulatory T cell (Treg) deficiency and is due to a Foxp3 gene mutation. Considering the pivotal role of lipids in autoimmune inflammatory processes, we investigated alterations in the relative abundance of lipid profiles in SF mice (± treatment with DSM 17938) compared to normal WT mice. We also examined the correlation between plasma lipids and gut microbiota and circulating inflammatory markers. We noted a significant upregulation of plasma lipids associated with autoimmune disease in SF mice, many of which were downregulated by DSM 17938. The upregulated lipids in SF mice demonstrated a significant correlation with gut bacteria known to be implicated in the pathogenesis of various autoimmune diseases. Chronic hepatitis in SF livers responded to DSM 17938 treatment with a reduction in hepatic inflammation. Altered gene expression associated with lipid metabolism and the positive correlation between lipids and inflammatory cytokines together suggest that autoimmunity leads to dyslipidemia with impaired fatty acid oxidation in SF mice. Probiotics are presumed to contribute to the reduction of lipids by reducing inflammatory pathways.

When new-born mice are subjected to acute maternal separation stress, cow-milk based formula feeding, and brief recurrent hypoxia with cold stress, they develop gut inflammation similar to the phenotype of neonatal necrotizing enterocolitis, characterised by an increase in gut mucosal effector T (Teffs) and reduced Foxp3 + regulatory T (Tregs) cells. The imbalance can be prevented by probiotic Limosilactobacillus reuteri DSM 17938 (LR 17938). We hypothesised that LR 17938 could potentiate a tolerogenic function of Tregs. To analyse whether LR 17938 can educate Tregs to improve their tolerogenic potency during neonatal stress, we isolated T cells (Tregs and Teffs) from 'donor' mice fed with either LR 17938 (10 7 cfu) or control media. The cells were adoptively transferred (AT) by intraperitoneal injection (5 × 10 5 cells/mouse) to new-born (d5) recipient mice. Mice were then separated from their dams, fed formula by gavage, and exposed to hypoxia and cold stress (NeoStress) for 4 days. We analysed the percentage of Tregs in CD4 + T helper cells in the intestine (INT) and mesenteric lymph nodes (MLN) of recipient mice. We found that: (1) the percentage of Tregs in the INT and MLN following NeoStress were significantly reduced compared to dam-fed unstressed mice; (2) AT of either naïve Tregs or LR-educated Tregs to mice with Neostress increased the percentage of Tregs in the INT and MLN compared to the percentage in NeoStress mice without Treg treatment; however, LR-educated Tregs increased the Tregs significantly more than naïve Tregs; and (3) AT of LR-educated Tregs reduced pro-inflammatory CD44 + Foxp3 - NonTregs and inflammatory CX3CR1 + dendritic cells in the intestinal mucosa of NeoStress mice. In conclusion, adoptive transfer of Tregs promotes the generation of and/or migration of endogenous Tregs in the intestinal mucosa of recipient mice. Importantly, probiotic-educated Tregs are more potent than naïve Tregs to enhance immune tolerance following neonatal stress.

We studied the effect of feeding a single probiotic Limosilactobacillus reuteri DSM 17938 (LR 17938) on the luminal and plasma levels of amino acids and their derivatives in the suckling newborn mouse, using gas chromatography and high-performance liquid chromatography. We found that LR 17938 increased the relative abundance of many amino acids and their derivatives in stool, while it simultaneously significantly reduced the plasma levels of three amino acids (serine, citrulline, and taurine). Many peptides and dipeptides were increased in stool and plasma, notably gamma-glutamyl derivatives of amino acids, following ingestion of the LR 17938. Gamma-glutamyl transformation of amino acids facilitates their absorption. LR 17938 significantly upregulated N-acetylated amino acids, the levels of which could be useful biomarkers in plasma and warrant further investigation. Specific fecal microbiota were associated with higher levels of fecal amino acids and their derivatives. Changes in luminal and circulating levels of amino acid derivatives, polyamines, and tryptophan metabolites may be mechanistically related to probiotic efficacy., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Background: Probiotic Lactobacillus reuteri DSM 17938 (LR 17938) is beneficial to infants with colic. To understand its mechanism of action, we assessed ultrasonic vocalizations (USV) and brain pain/stress genes in newborn mice exposed to maternal separation stress., Methods: Pups were exposed to unpredictable maternal separation (MSU or SEP) or MSU combined with unpredictable maternal stress (MSU + MSUS or S + S), from postnatal days 5 to 14. USV calls and pain/stress/neuroinflammation-related genes in the brain were analyzed., Results: We defined 10 different neonatal call patterns, none of which increased after MSU. Stress reduced overall USV calls. Orally feeding LR 17938 also did not change USV calls after MSU. However, LR 17938 markedly increased vocalizations in mice allowed to stay with their dams. Even though LR 17938 did not change MSU-related calls, LR 17938 modulated brain genes related to stress and pain. Up-regulated genes following LR 17938 treatment were opioid peptides, kappa-opioid receptor 1 genes, and CD200, important in anti-inflammatory signaling. LR 17938 down-regulated CCR2 transcripts, a chemokine receptor, in the stressed neonatal brain., Conclusions: USV calls in newborn mice are interpreted as "physiological calls" instead of "cries." Feeding LR 17938 after MSU did not change USV calls but modulated cerebral genes favoring pain and stress reduction and anti-inflammatory signaling., Impact: We defined mouse ultrasonic vocalization (USV) call patterns in this study, which will be important in guiding future studies in other mouse strains. Newborn mice with maternal separation stress have reduced USVs, compared to newborn mice without stress, indicating USV calls may represent "physiological calling" instead of "crying." Oral feeding of probiotic Lactobacillus reuteri DSM 17938 raised the number of calls when newborn mice continued to suckle on their dams, but not when mice were under stress. The probiotic bacteria had a dampening effect on monocyte activation and on epinephrine and glutamate-related stress gene expression in the mouse brain., (© 2021. The Author(s), under exclusive licence to the International Pediatric Research Foundation, Inc.)

Early administration of Lactobacillus reuteri DSM 17938 (LR) prevents necrotizing enterocolitis and inhibits regulatory T-cell (Treg)-deficiency-associated autoimmunity in mice. In humans, LR reduces crying time in breastfed infants with colic, modifies severity in infants with acute diarrheal illnesses, and improves pain in children with functional bowel disorders. In healthy breastfed newborns with evolving microbial colonization, it is unclear if early administration of LR can modulate gut microbiota and their metabolites in such a way as to promote homeostasis. We gavaged LR (10 7 colony-forming units/day, daily) to C57BL/6J mice at age of day 8 for 2 wk. Both male and female mice were investigated in these experiments. We found that feeding LR did not affect clinical phenotype or inflammatory biomarkers in plasma and stool, but LR increased the proportion of Foxp3 + regulatory T cells (Tregs) in the intestine. LR also increased bacterial diversity and the relative abundance of p_Firmicutes, f_Lachnospiraceae , f_Ruminococcaceae , and genera Clostridium and Candidatus arthromitus , while decreasing the relative abundance of p_Bacteriodetes, f_Bacteroidaceae , f_Verrucomicrobiaceae , and genera Bacteroides , Ruminococcus, Akkermansia , and Sutterella . Finally, LR exerted a major impact on the plasma metabolome, upregulating amino acid metabolites formed via the urea, tricarboxylic acid, and methionine cycles and increasing tryptophan metabolism. In conclusion, early oral administration of LR to healthy breastfed mice led to microbial and metabolic changes which could be beneficial to general health. NEW & NOTEWORTHY Oral administration of Lactobacillus reuteri DSM 17938 (LR) to healthy breastfed mice promotes intestinal immune tolerance and is linked to proliferation of beneficial gut microbiota. LR upregulates plasma metabolites that are involved in the urea cycle, the TCA cycle, methionine methylation, and the polyamine pathway. Herein, we show that LR given to newborn mice specifically increases levels of tryptophan metabolites and the purine nucleoside adenosine that are known to enhance tolerance to inflammatory stimuli.

Human breast milk (HBM) may have beneficial effects on Lactobacillus reuteri DSM 17938 (LR 17938) -mediated immunomodulation. We aimed to determine the effects of HBM on proliferation of LR 17938 in vitro and its associated proteins and metabolites in culture, in order to provide mechanistic insights into the health benefits of LR 17938. LR 17938 was cultured anaerobically in MRS bacterial culture media, HBM (from 6 mothers), and 2 types of cow-milk formula. The colony-forming unit (CFU) was calculated to evaluate LR 17938 growth. Sixteen-hour-fermented supernatants were used for metabolomics, and bacterial lysates were used for proteomics analysis. We found that growth of LR 17938 was 10 times better in HBM than in formula. We detected 261/452 metabolites upregulated when LR 17938 cultured in HBM compared to in formula, mainly participating in the glyoxylate cycle (succinate), urea cycle (citrulline), methionine methylation (N-acetylcysteine), and polyamine synthesis (spermidine). The significantly up-regulated enzymes were also involved in the formation of acetyl-CoA in the glyoxylate cycle and the antioxidant N-acetylcysteine. In conclusion, HBM enhances the growth of LR 17938 compared to formula and promotes LR 17938-associated metabolites that relate to energy and antioxidant status, which may be linked to the physiological effects of L. reuteri .

Probiotics have been used to ameliorate gastrointestinal symptoms since ancient times. Over the past 40 years, probiotics have been shown to impact the immune system, both in vivo and in vitro. This interaction is linked to gut microbes, their polysaccharide antigens, and key metabolites produced by these bacteria. At least four metabolic pathways have been implicated in mechanistic studies of probiotics, based on mechanistic studies in animal models. Microbial⁻immune system crosstalk has been linked to: short-chain fatty acid production and signaling, tryptophan metabolism and the activation of aryl hydrocarbon receptors, nucleoside signaling in the gut, and activation of the intestinal histamine-2 receptor. Several randomized controlled trials have now shown that microbial modification by probiotics may improve gastrointestinal symptoms and multiorgan inflammation in rheumatoid arthritis, ulcerative colitis, and multiple sclerosis. Future work will need to carefully assess safety issues, selection of optimal strains and combinations, and attempts to prolong the duration of colonization of beneficial microbes.

Few treatments for human diseases have received as much investigation in the past 20 years as probiotics. In 2017, English-language meta-analyses totaling 52 studies determined the effect of probiotics on conditions ranging from necrotizing enterocolitis and colic in infants to constipation, irritable bowel syndrome, and hepatic encephalopathy in adults. The strongest evidence in favor of probiotics lies in the prevention or treatment of 5 disorders: necrotizing enterocolitis, acute infectious diarrhea, acute respiratory tract infections, antibiotic-associated diarrhea, and infant colic. Probiotic mechanisms of action include the inhibition of bacterial adhesion; enhanced mucosal barrier function; modulation of the innate and adaptive immune systems (including induction of tolerogenic dendritic cells and regulatory T cells); secretion of bioactive metabolites; and regulation of the enteric and central nervous systems. Future research is needed to identify the optimal probiotic and dose for specific diseases, to address whether the addition of prebiotics (to form synbiotics) would enhance activity, and to determine if defined microbial communities would provide benefit exceeding that of single-species probiotics., (© 2018, The American College of Clinical Pharmacology.)

Objective: To assess the safety of probiotic Lactobacillus reuteri strain Deutsche Sammlung von Mikroorganismen (DSM) 17938 with daily administration to healthy infants with colic and to determine the effect of L reuteri strain DSM 17938 on crying, fussing, inflammatory, immune, and microbiome variables., Study Design: We performed a controlled, double-blinded, phase 1 safety and tolerability trial in healthy breast-fed infants with colic, aged 3 weeks to 3 months, randomly assigned to L reuteri strain DSM 17938 (5 × 10 8 colony-forming units daily) or placebo for 42 days and followed for 134 days., Results: Of 117 screened infants, 20 were randomized to L reuteri strain DSM 17938 or placebo (sunflower oil) (in a 2:1 ratio) with 80% retention. Eleven of the 20 (55%) presented with low absolute neutrophil counts (<1500/mm 3 ), which resolved in all subjects by day 176. L reuteri strain DSM 17938 produced no severe adverse events and did not significantly change crying time, plasma bicarbonate, or inflammatory biomarkers. Fecal calprotectin decreased rapidly in both groups. In the infants with dominant fecal gram negatives (Klebsiella, Proteus, and Veillonella), resolution of colic was associated with marked decreases in these organisms., Conclusions: Daily administration of L reuteri strain DSM 17938 appears to be safe in newborn infants with colic, including those with neutropenia, which frequently coexists. A placebo response of 66% suggests that many infants with colic will have resolution within 3 weeks., Trial Registration: ClinicalTrials.gov: NCT01849991., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Necrotizing enterocolitis (NEC) is an inflammatory disease with evidence of increased production of proinflammatory cytokines in the intestinal mucosa. Lactobacillus reuteri DSM 17938 (LR17938) has been shown to have anti-inflammatory activities in an experimental model of NEC. Activated effector lymphocyte recruitment to sites of inflammation requires the sequential engagement of adhesion molecules such as CD44. The phenotype of CD44(+)CD45RB(lo) separates T effector/memory (Tem) cells from naive (CD44(-)CD45RB(hi)) cells. It is unknown whether these Tem cells participate in the inflammation associated with NEC and can be altered by LR17938. NEC was induced in 8- to 10-day-old C57BL/6J mice by gavage feeding with formula and exposure to hypoxia and cold stress for 4 days. Survival curves and histological scores were analyzed. Lymphocytes isolated from mesenteric lymph nodes and ileum were labeled for CD4, CD44, CD45RB, intracellular Foxp3, and Helios and subsequently analyzed by flow cytometry. LR17938 decreased mortality and the incidence and severity of NEC. The percentage of Tem cells in the ileum and mesenteric lymph nodes was increased in NEC but decreased by LR17938. Conversely, the percentage of CD4(+)Foxp3(+) regulatory T (Treg) cells in the intestine decreased during NEC and was restored to normal by LR17938. The majority of the Treg cells preserved by LR17938 were Helios+ subsets, possibly of thymic origin. In conclusion, LR17938 may represent a useful treatment to prevent NEC. The mechanism of protection by LR17938 involves modulation of the balance between Tem and Treg cells. These T cell subsets might be potential biomarkers and therapeutic targets during intestinal inflammation., (Copyright © 2014 the American Physiological Society.)

Background: There are few carefully-designed studies investigating the safety of individual probiotics approved under Investigational New Drug policies., Objectives: The primary aim of this prospective, double-blind placebo-controlled trial was to investigate if daily treatment of adults with Lactobacillus reuteri DSM 17938 (LR) for 2 months is safe and well-tolerated. Our secondary aim was to determine if LR treatment has immune effects as determined by regulatory T cell percentages, expression of toll-like receptors (TLR)-2 and -4 on circulating peripheral blood mononuclear cells (PMBCs), cytokine expression by stimulated PBMC, and intestinal inflammation as measured by fecal calprotectin., Methods: Forty healthy adults were randomized to a daily dose of 5 × 10(8) CFUs of LR (n = 30) or placebo (n = 10) for 2 months. Participants completed a daily diary card and had 7 clinic visits during treatment and observation., Results: There were no severe adverse events (SAEs) and no significant differences in adverse events (AEs). There were no differences in PBMC subclasses, TLRs, or cytokine expression after treatment. The probiotic-treated group had a significantly higher fecal calprotectin level than the placebo group after 2 months of treatment: 50 µg/g (IQR 24-127 µg/g) vs. 17 µg/g (IQR 11-26 µg/g), p = 0.03, although values remained in the normal clinical range (0-162.9 µg/g). LR vials retained >10(8) CFUs viable organisms/ml., Conclusions: LR is safe and well tolerated in adults, without significant changes in immunologic markers. There was a small but significant increase in fecal calprotectin, perhaps indicating some element of immune recognition at the intestinal level., Trial Registration: Clinical Trials.gov NCT00922727.

Treg deficiency causes a lethal, CD4+ T cell-driven autoimmune disease called IPEX syndrome (immunodysregulation, polyendocrinopathy, and enteropathy, with X-linked inheritance) in humans and in the scurfy (SF) mouse, a mouse model of the disease. Feeding Limosilactobacillus reuteri DSM 17938 (LR 17938, LR) to SF mice reprograms the gut microbiota, reduces disease progression, and prolongs lifespan. However, the efficacy and mechanism of LR, compared with other probiotics, in producing these effects is unknown. We compared LR with Lacticaseibacillus rhamnosus GG (LGG), an extensively investigated probiotic. LR was more effective than LGG in prolonging survival. Both probiotics restored the fecal microbial alpha diversity, but they produced distinct fecal bacterial clusters and differentially modulated microbial relative abundance (RA). LR increased the RA of phylum_Firmicutes, genus_Oscillospira whereas LR reduced phylum_Bacteroidetes, genus_Bacteroides and genus_Parabacteroides, reversing changes attributed to the SF phenotype. LGG primarily reduced the RA of genus_Bacteroides. Both LR and LGG reduced the potentially pathogenic taxon class_cproteobacteria. Plasma metabolomics revealed substantial differences among 696 metabolites. We observed similar changes of many clusters of metabolites in SF mice associated with treatment with either LR or LGG. However, a unique effect of LR was to increase the abundance of plasma adenosine metabolites such as inosine, which we previously showed had immune modulatory effects. In conclusion: 1) different probiotics produce distinct signatures in the fecal microbial community in mice with Treg deficiency; and 2) when comparing different probiotics, there are strain-specific microbial products with different anti-inflammatory properties, reinforcing the concept that “one size does not fit all” in the treatment of autoimmune disease. [ABSTRACT FROM AUTHOR]

Necrotizing enterocolitis (NEC) is an inflammatory disease with evidence of increased production of proinflammatory cytokines in the intestinal mucosa. Lactobacillus reuteri DSM 17938 (LR17938) has been shown to have anti-inflammatory activities in an experimental model of NEC. Activated effector lymphocyte recruitment to sites of inflammation requires the sequential engagement of adhesion molecules such as CD44. The phenotype of CD44+CD45RBlo separates T effector/memory (Tem) cells from naive (CD44-CD45RBhi) cells. It is unknown whether these Tem cells participate in the inflammation associated with NEC and can be altered by LR17938. NEC was induced in 8- to 10-day-old C57BL/6J mice by gavage feeding with formula and exposure to hypoxia and cold stress for 4 days. Survival curves and histological scores were analyzed. Lymphocytes isolated from mesenteric lymph nodes and ileum were labeled for CD4, CD44, CD45RB, intracellular Foxp3, and Helios and subsequently analyzed by flow cytometry. LR17938 decreased mortality and the incidence and severity of NEC. The percentage of Tem cells in the ileum and mesenteric lymph nodes was increased in NEC but decreased by LR17938. Conversely, the percentage of CD4+Foxp3+ regulatory T (Treg) cells in the intestine decreased during NEC and was restored to normal by LR17938. The majority of the Treg cells preserved by LR17938 were Helios+ subsets, possibly of thymic origin. In conclusion, LR17938 may represent a useful treatment to prevent NEC. The mechanism of protection by LR17938 involves modulation of the balance between Tem and Treg cells. These T cell subsets might be potential biomarkers and therapeutic targets during intestinal inflammation. [ABSTRACT FROM AUTHOR]