MicroRNAs in bovine milk exosomes are bioavailable in humans but do not elicit a robust pro-inflammatory cytokine response

Background: Bovine milk exosomes are studied for their roles as bioactive food compounds and as vehicles for drug delivery. Both lines of investigation converge on immune function, e.g., immune regulation by absorption of microRNAs encapsulated in milk exosomes across species boundaries, and the possibility of exosomes and their cargos triggering an immune response if used in drug delivery. This study assessed the bioavailability of immune-related microRNAs from bovine milk and changes in plasma cytokine concentrations after milk consumption in humans, and the secretion of cytokines by human peripheral blood mononuclear cells (PBMCs) cultured with milk exosomes transfected with immune-relevant microRNAs. Results: Human plasma samples were collected before and at timed intervals after a milk meal and analyzed for concentrations of six immune-relevant microRNAs and nine cytokines. The peak plasma concentrations of miR-15b-5p, miR- 21-5p, miR-106b-5p, and miR-223-3p were 60 ± 9.80% to 162 ± 31.80% higher after milk consumption (Ct values 23 ± 1.2 to 26 ± 1.1 cycles) compared to baseline values (P \u3c 0.05). Plasma concentrations of TNF-alpha were not significantly different before versus after milk consumption; eight other cytokines were below detection limit. PBMCs were collected before and six hours after milk consumption and cultured with or without concanavalin A (ConA). TNF-alpha, IL-1β, IL-6 and IL-10 were detectable in culture media, but concentrations did not depend on milk consumption prior to PBMC isolation (P \u3e 0.05). When PBMC cultures from fasted subjects were supplemented with milk exosomes that had been transfected with immune-relevant microRNAs, the concentrations of IL-1β, IL-6, IL-10 and TNF-alpha were 29 ± 12% to 220 ± 33% higher than controls cultured with non-transfected exosomes (P \u3c 0.05), but cytokine concentrations were not different compared with control exosomes transfected with scrambled microRNA (P \u3e 0.05). Conclusions: MicroRNAs in bovine milk exosomes are bioavailable. Milk exosomes do not elicit an increase of plasma cytokines following oral administration

IL-2 limits IL-12 enhanced lymphocyte proliferation during \u3ci\u3eLeishmania amazonensis\u3c/i\u3e infection

C3H mice infected with Leishmania amazonensis develop persistent, localized lesions with high parasite loads. During infection, memory/effector CD44hiCD4+ T cells proliferate and produce IL-2, but do not polarize to a known effector phenotype. Previous studies have demonstrated IL-12 is insufficient to skew these antigen-responsive T cells to a functional Th1 response. To determine the mechanism of this IL-12 unresponsiveness, we used an in vitro assay of repeated antigen activation. Memory/effector CD44hiCD4+ T cells did not increase proliferation in response to either IL-2 or IL-12, although these cytokines upregulated CD25 expression. Neutralization of IL-2 enhanced CD4+ T cell proliferation in response to IL-12. This cross-regulation of IL-12 responsiveness by IL-2 was confirmed in vivo by treatment with anti-IL-2 antibodies and IL-12 during antigen challenge of previously infected mice. These results suggest that during chronic infection with L. amazonensis, IL-2 plays a dominant, immunosuppressive role independent of identifiable conventional Treg cells

Immunomodulatory Role of Urolithin A on Metabolic Diseases

Urolithin A (UroA) is a gut metabolite produced from ellagic acid-containing foods such as pomegranates, berries, and walnuts. UroA is of growing interest due to its therapeutic potential for various metabolic diseases based on immunomodulatory properties. Recent advances in UroA research suggest that UroA administration attenuates inflammation in various tissues, including the brain, adipose, heart, and liver tissues, leading to the potential delay or prevention of the onset of Alzheimer’s disease, type 2 diabetes mellitus, and non-alcoholic fatty liver disease. In this review, we focus on recent updates of the anti-inflammatory function of UroA and summarize the potential mechanisms by which UroA may help attenuate the onset of diseases in a tissue-specific manner. Therefore, this review aims to shed new insights into UroA as a potent anti-inflammatory molecule to prevent immunometabolic diseases, either by dietary intervention with ellagic acid-rich food or by UroA administration as a new pharmaceutical drug

Amphiphilic Polyanhydride Nanoparticles Stabilize \u3ci\u3eBacillus anthracis\u3c/i\u3e Protective Antigen

Advancements towards an improved vaccine against Bacillus anthracis, the causative agent of anthrax, have focused on formulations composed of the protective antigen (PA) adsorbed to aluminum hydroxide. However, due to the labile nature of PA, antigen stability is a primary concern for vaccine development. Thus, there is a need for a delivery system capable of preserving the immunogenicity of PA through all the steps of vaccine fabrication, storage, and administration. In this work, we demonstrate that biodegradable amphiphilic polyanhydride nanoparticles, which have previously been shown to provide controlled antigen delivery, antigen stability, immune modulation, and protection in a single dose against a pathogenic challenge, can stabilize and release functional PA. These nanoparticles demonstrated polymer hydrophobicity-dependent preservation of the biological function of PA upon encapsulation, storage (over extended times and elevated temperatures), and release. Specifically, fabrication of amphiphilic polyanhydride nanoparticles composed of 1,6-bis(p-carboxyphenoxy)hexane and 1,8-bis(p-carboxyphenoxy)-3,6- dioxaoctane best preserved PA functionality. These studies demonstrate the versatility and superiority of amphiphilic nanoparticles as vaccine delivery vehicles suitable for long-term storage

Urolithin A, a Gut Metabolite, Improves Insulin Sensitivity through Augmentation of Mitochondrial Function and Biogenesis

Objective: Urolithin A (UroA) is a major metabolite of ellagic acid produced following microbial catabolism in the gut. Emerging evidence has suggested that UroA modulates energy metabolism in various cells. However, UroA’s physiological functions related to obesity and insulin resistance remain unclear. Methods: Male mice were intraperitoneally administrated either UroA or dimethyl sulfoxide (vehicle) along with a high-fat diet for 12 weeks. Insulin sensitivity was evaluated via glucose and insulin tolerance tests and acute insulin signaling. The effects of UroA on hepatic triglyceride accumulation, adipocyte size, mitochondrial DNA content, and proinflammatory gene expressions were determined. The impact of UroA on macrophage polarization and mitochondrial respiration were assessed in bone marrow–derived macrophages. Results: Administration of UroA (1) improved systemic insulin sensitivity, (2) attenuated triglyceride accumulation and elevated mitochondrial biogenesis in the liver, (3) reduced adipocyte hypertrophy and macrophage infiltration into the adipose tissue, and (4) altered M1/M2 polarization in peritoneal macrophages. In addition, UroA favored macrophage M2 polarization and mitochondrial respiration in bone marrow–derived macrophages. Conclusions: UroA plays a direct role in improving systemic insulin sensitivity independent of its parental compounds. This work supports UroA’s role in the metabolic benefits of ellagic acid–rich foods and highlights the significance of its microbial transformation in the gut

Diet-microbiome-immune interplay in multiple sclerosis: Understanding the impact of phytoestrogen metabolizing gut bacteria

Multiple sclerosis (MS) is a chronic and progressive autoimmune disease of the central nervous system (CNS), with both genetic and environmental factors contributing to the pathobiology of the disease. Although HLA genes have emerged as the strongest genetic factor linked to MS, consensus on the environmental risk factors is lacking. Recently, the gut microbiota has garnered increasing attention as a potential environmental factor in MS, as mounting evidence suggests that individuals with MS exhibit microbial dysbiosis (changes in the gut microbiome). Thus, there has been a strong emphasis on understanding the role of the gut microbiome in the pathobiology of MS, specifically, factors regulating the gut microbiota and the mechanism(s) through which gut microbes may contribute to MS. Among all factors, diet has emerged to have the strongest influence on the composition and function of gut microbiota. As MS patients lack gut bacteria capable of metabolizing dietary phytoestrogen,we will specifically discuss the role of a phytoestrogen diet and phytoestrogen metabolizing gut bacteria in the pathobiology of MS. A better understanding of these mechanisms will help to harness the enormous potential of the gut microbiota as potential therapeutics to treat MS and other autoimmune diseases

Polyphenolic fractions isolated from red raspberry whole fruit, pulp, and seed differentially alter the gut microbiota of mice with diet-induced obesity

Polyphenol extracts from red raspberry (RR) whole fruit or pulp, but not seed, attenuate high-fat (HF) diet-induced obesity in mice. Because host metabolism is linked to the microbiota, we investigated the effects of polyphenols from RR fruit, pulp, and seed on the microbiome. RR polyphenols significantly decreased the abundance of specific taxa that were increased during HF feeding relative to a low-fat diet, including Ruminococcus and an unclassified genus from Clostridiales. Compared to a HF diet, pulp and seed polyphenols increased Roseburia abundance and decreased levels of an unclassified genus from Mogibacteriaceae. RR seed polyphenols uniquely increased Bifidobacterium compared to a HF diet. The most notable taxon driving differential abundance among all diets was an unclassified genus from Coriobacteriaceae. Importantly, host metabolic markers improved by pulp polyphenols were strongly correlated with select microbiome features, indicating that specific gut bacteria may be involved in RR polyphenol catabolism and/or mediating health benefits

Dietary Depletion of Milk Exosomes and Their MicroRNA Cargos Elicits a Depletion of miR-200a-3p and Elevated Intestinal Inflammation and Chemokine (C-X-C Motif) Ligand 9 Expression in Mdr1a−/− Mice

Background: Exosomes transfer regulatory microRNAs (miRs) from donor cells to recipient cells. Exosomes and miRs originate from both endogenous synthesis and dietary sources such as milk. miR-200a-3p is a negative regulator of the proinflammatory chemokine (C-X-C motif) ligand 9 (CXCL9). Male Mdr1a−/− mice spontaneously develop clinical signs of inflammatory bowel disease (IBD). Objectives: We assessed whether dietary depletion of exosomes and miRs alters the severity of IBD in Mdr1a−/− mice owing to aberrant regulation of proinflammatory cytokines. Methods: Starting at 5 wk of age, 16 male Mdr1a−/− mice were fed either milk exosome– and RNA-sufficient (ERS) or milk exosome– and RNA-depleted (ERD) diets. The ERD diet is characterized by a near-complete depletion of miRs and a 60% loss of exosome bioavailability compared with ERS. Mice were killed when their weight loss exceeded 15% of peak body weight. Severity of IBD was assessed by histopathological evaluation of cecum. Serum cytokine and chemokine concentrations and mRNA and miR tissue expression were analyzed by multiplex ELISAs, RNA-sequencing analysis, and qRT-PCR, respectively. Results: Stromal collapse, gland hyperplasia, and additive microscopic disease scores were (mean ± SD) 56.7% ± 23.3%, 23.5% ± 11.8%, and 29.6% ± 8.2% lower, respectively, in ceca of ERS mice than of ERD mice (P \u3c 0.05). The serum concentration of CXCL9 was 35.0% ± 31.0% lower in ERS mice than in ERD mice (P \u3c 0.05). Eighty-seven mRNAs were differentially expressed in the ceca from ERS and ERD mice; 16 of these mRNAs are implicated in immune function. The concentrations of 4 and 1 out of 5 miRs assessed (including miR-200a-3p) were ≤63% lower in livers and ceca, respectively, from ERD mice than from ERS mice. Conclusions: Milk exosome and miR depletion exacerbates cecal inflammation in Mdr1a−/− mice

Differential Effects of Whole Red Raspberry Polyphenols and Their Gut Metabolite Urolithin A on Neuroinflammation in BV-2 Microglia

Whole red raspberry polyphenols (RRW), including ellagic acid, and their gut-derived metabolite, urolithin A (UroA), attenuate inflammation and confer health benefits. Although results from recent studies indicate that polyphenols and UroA also provide neuroprotective effects, these compounds differ in their bioavailability and may, therefore, have unique effects on limiting neuroinflammation. Accordingly, we aimed to compare the neuroprotective effects of RRW and UroA on BV-2 microglia under both 3 h and 12 and 24 h inflammatory conditions. In inflammation induced by lipopolysaccharide (LPS) and ATP stimulation after 3 h, RRW and UroA suppressed pro-inflammatory cytokine gene expression and regulated the JNK/c-Jun signaling pathway. UroA also reduced inducible nitric oxide synthase gene expression and promoted M2 microglial polarization. During inflammatory conditions induced by either 12 or 24 h stimulation with LPS, UroA—but not RRW—dampened pro-inflammatory cytokine gene expression and suppressed JNK/c-Jun signaling. Taken together, these results demonstrate that RRW and its gut-derived metabolite UroA differentially regulate neuroprotective responses in microglia during 3 h versus 12 and 24 h inflammatory conditions