Isolating Immune Cells from Mouse Brain and Skull.

Mounting evidence indicates that the immune response triggered by brain disorders (e.g., brain ischemia and autoimmune encephalomyelitis) occurs not only in the brain, but also in the skull. A key step toward analyzing changes in immune cell populations in both the brain and skull bone marrow after brain damage (e.g., stroke) is to obtain sufficient numbers of high-quality immune cells for downstream analyses. Here, two optimized protocols are provided for isolating immune cells from the brain and skull bone marrow. The advantages of both protocols are reflected in their simplicity, speed, and efficacy in yielding a large quantity of viable immune cells. These cells may be suitable for a range of downstream applications, such as cell sorting, flow cytometry, and transcriptomic analysis. To demonstrate the effectiveness of the protocols, immunophenotyping experiments were performed on stroke brains and normal brain skull bone marrow using flow cytometry analysis, and the results aligned with findings from published studies.

Lactobacillus rhamnosus Attenuates Cisplatin-Induced Intestinal Mucositis in Mice via Modulating the Gut Microbiota and Improving Intestinal Inflammation.

Lactobacillus rhamnosus (LBS) is a well-documented probiotic strain in oncology and has a pivotal role in clinical applications. Here, we have investigated the protective effect of Lactobacillus rhamnosus on intestinal mucositis induced by cisplatin (CP) and explored the underlying mechanisms targeting inflammatory proteins, as well as the histological changes in the intestinal tissue of mice, in addition, the bacterial strains that may be related to the health-enhancing properties. BALB/c mice were pre-treated with or without LBS via oral gavage, followed by mucositis induction with cisplatin. Our results revealed that the LBS-treated groups significantly attenuated proinflammatory cytokine levels (IL-1ß, IL-6, and TNF-α) compared to the CP group. Furthermore, LBS mitigated the damaged tight junction integrity caused by CP via up-regulating the levels of claudin, occludin, ZO-1, and mucin-2 protein (MUC-2). Finally, the 16S rRNA fecal microbiome genomic analysis showed that LBS administration enhanced the growth of beneficial bacteria, i.e., Firmicutes and Lachnospiraceae, while the relative abundance of the opportunistic bacteria Bacteroides and Proteobacteria decreased. Collectively, LBS was found to beneficially modulate microbial composition structure and functions and enrich the ecological diversity in the gut.

Sea Conch Peptides Hydrolysate Alleviates DSS-Induced Colitis in Mice through Immune Modulation and Gut Microbiota Restoration.

Inflammatory bowel disease (IBD) is a persistent, lifelong inflammation of the digestive system. Dextran sulfate sodium is commonly used to induce colitis in experimental animal models, which causes epithelial damage, intestinal inflammation, mucin depletion, and dysbiosis of the gut microbiota. Various prebiotics, polysaccharides, and polypeptides are used for IBD treatment. In this study, we used a murine model utilizing BALB/c mice, with 10 mice per group, to investigate the treatment effect of sea conch peptide hydrolysate (CPH) on DSS-induced colitis mice. Colitis was induced through the administration of 2.5% DSS in drinking water over a seven-days period. Furthermore, on the eighth day of the experiment, sea conch peptide hydrolysate (CPH) at low (100 mg/kg), medium (200 mg/kg), and high (400 mg/kg) doses, which were continued for 14 days, were assessed for medicinal purposes in DSS-induced colitis mice. Our results showed that CPH treatment significantly alleviated the severity and symptoms of colitis. The epithelial integrity and histological damage were improved. Intestinal inflammation and inflammatory cell infiltration were improved. Furthermore, the expression of pro-inflammatory cytokines was reduced, and intestinal barrier integrity was restored by elevating the tight junction proteins. Moreover, 16s RNA sequencing revealed dysbiosis of the gut microbiota was observed upon DSS treatment, which was reinstated after CPH treatment. An increased level of Firmicutes and Lactobacillus was observed in the treatment groups. Finally, our results suggest that CPH would be recommended as a functional food source and also have the potential to be used as a medicinal product for different gastrointestinal disorders.

Virulence factors and mechanisms of paediatric pneumonia caused by Enterococcus faecalis.

Paediatric pneumonia is a respiratory infection that affects infants and young children under the age of 3. This disease is the leading cause of infant and child mortality in developing countries because of the weak immune system of young children. The difficulty and length of time required to identify the pathogen and causative agent are the main reasons for this high mortality rate. In addition, the identification of certain causative agents is particularly important for the treatment of paediatric pneumonia. In this study, we explored the possible mechanisms by which pathogenic Enterococcus faecalis induced pneumonia in vivo. The potential virulence factors of bacteria isolated from the intestines of paediatric pneumonia patients were determined. Taken together, the results suggested that lysophosphatidic acid (LTA) from pathogenic E. faecalis decreases the expression of platelet-activating factor receptor (PAFR), which in turn disrupts the function of intestinal tight junctions (Occ and Ccldn1), leading to the entry of LE-LTA into the bloodstream because of the disruption of the intestinal barrier. Although LTA can enter circulation, it cannot directly infiltrate the lungs, which indicates that lung inflammation in mice is not caused by the direct entry of LE-LTA into the lungs. We further found that LTA activates immune cells, such as CD8 + T cells and type 2 innate lymphocytes, in vivo. Interleukin-6 and interleukin-17 can produce large amounts of inflammatory factors and thus promote the development of pneumonia. In conclusion, our findings demonstrate that the LTA of pathogenic E. faecalis in the intestine is a virulence factor that can cause paediatric pneumonia. This study found that intestinal bacterial virulence factors can induce immune responses in the lungs and blood. These findings could provide further insight into the mechanism of infectious diseases in the lung that are caused by bacteria in the intestine.

Facial Synthesis, Stability, and Interaction of Ti3C2Tx@PC Composites for High-Performance Biocathode Microbial Electrosynthesis Systems.

Developing high-performance biocathodes remain one of the most challenging aspects of the microbial electrosynthesis (MES) system and the primary factor limiting its output. Herein, a hollow porous carbon (PC) fabricated with MXenes coated over an electrode was developed for MES systems to facilitate the direct delivery of CO2 to microorganisms colonized. The result highlighted that MXene@PC (Ti3C2Tx@PC) has a surface area of 434 m2/g. The Ti3C2Tx@PC MES cycle shows that in cycle 4 and cycle 5, the values are -309.2 and -352.3. Cyclic voltammetry showed that the coated electrode current response (mA) increased from -4.5 to -20.2. The substantial redox peaks of Ti3C2Tx@PC biofilms are displayed at -741, -516, and -427 mV vs Ag/AgCl, suggesting an enhanced electron transfer owing to the Ti3C2Tx@PC complex coating. Additionally, more active sites enhanced mass transfer and microbial development, resulting in a 46% rise in butyrate compared to the uncoated control. These findings demonstrate the value of PC modification as a method for MES-based product selection.

Morchella esculenta mushroom polysaccharide attenuates diabetes and modulates intestinal permeability and gut microbiota in a type 2 diabetic mice model.

Type 2 diabetes mellitus (T2DM) is a health issue that causes serious worldwide economic problems. It has previously been reported that natural polysaccharides have been studied with regard to regulating the gut microbiota, which plays an important role in T2DM. Here, we investigate the effects of Morchella esculenta polysaccharide (MEP) on a high-fat diet (HFD) and streptozotocin (STZ)-induced T2DM in BALB/c mice. The administration of MEP effectively regulated hyperglycemia and hyperlipidemia and improved insulin sensitivity. We also determined an improvement in gut microbiota composition by 16sRNA pyrosequencing. Treatment with MEP showed an increase in beneficial bacteria, i.e., Lactobacillus and Firmicutes, while the proportion of the opportunistic bacteria Actinobacteria, Corynebacterium, and Facklamia decreased. Furthermore, the treatment of T2DM mice with MEP resulted in reduced endotoxemia and insulin resistance-related pro-inflammatory cytokines interleukin 1ß (IL-1ß), tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6). Moreover, MEP treatment improved intestinal permeability by modulating the expression of the colon tight-junction proteins zonula occludens-1 (ZO-1), occludin, claudin-1, and mucin-2 protein (MUC2). Additionally, MEP administration affects the metagenome of microbial communities in T2DM mice by altering the functional metabolic pathways. All these findings suggested that MEP is a beneficial prebiotic associated with ameliorating the gut microbiota and its metabolites in T2DM.

Morchella esculenta polysaccharide attenuate obesity, inflammation and modulate gut microbiota.

Edible mushrooms have now been suggested as promising sources of biological functional ingredients and are the subject of the most recent nutrition research and novel functional foods. Polysaccharides from mushrooms exhibit impressive biological effects, notably against obesity. Obesity is a chronic metabolic disorder characterized by chronic inflammation, gut dysbiosis, and hyperpermeability of the colon. Here, we prove that mushrooms Morchella esculenta polysaccharide (MEP) effects on HFD-induced obesity, colonic inflammation, and gut microbiota dysbiosis. Our findings demonstrate MEP supplementation attenuates obesity parameters and reduces inflammation in the colon via regulation of Toll-like receptor 4 (TLR4), nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and inactivation of nuclear factor kappa B (NF-κB). Furthermore, MEP administration restores gut microbiota dysregulation by ameliorating Firmicutes to Bacteroidetes proportion as well as enhancing beneficial bacteria, like Lactobacillus, and inhibiting pathogenic bacteria like Enterococcus. MEP improves gut integrity by increasing tight junction proteins (TJs) and reducing endotoxin levels by controlling Lipopolysaccharide (LPS) in HFD-induced obese mice. These results demonstrated the therapeutic efficacy of MEP in attenuating HFD-induced obesity via regulating inflammatory cascades, ameliorating the gut microbiome, and modulating gut integrity.

Effect of crude polysaccharide from seaweed, Dictyopteris divaricata (CDDP) on gut microbiota restoration and anti-diabetic activity in streptozotocin (STZ)-induced T1DM mice.

Type-1 Diabetes Mellitus (T1DM) is regarded as a multifunctional, immune-related disease which causes massive destruction of islet ß-cells in pancreas resulting in hyperglycemic, hypoinsulinemia and hyperlipidimic conditions. The aim of the present study, was to investigate the hypothesis that streptozotocin (STZ)-induced T1DM in Balb/c mice when treated with crude polysaccharide from seaweed, Dictyopteris divaricata (CDDP) depicts improvement in diabetes-related symptoms. Treatment with CDDP resulted in decreased body weight loss, improved food consumption and water intake disbalances. The CDDP effectively improved fasting blood glucose, oral glucose tolerance (OGTT), serum insulin, insulin secretion, rejuvenation of ß-cells mass, serum lipid profile and pro-inflammatory cytokines levels. Additionally, treatment with CDDP increased the population of beneficial bacteria such as Firmicutes, Bacteroidetes and Lactobacillus at phylum, family and genus levels by 16S rRNA sequencing. Furthermore, immunohistological examination confirmed that CDDP reduces the inflammation and restored the structural morphology of colon and upraised the levels of insulin receptor substrate-1 (IRS-1), Mucin-2 (MUC-2) and tight-junction proteins (TJs) whereby maintaining the gut structures and barrier permeability. Thus, the above presented data, highlights the safe and therapeutic effects of crude polysaccharide (CDDP) from D. divaricata in the treatment and restoration of T1DM disorders and can be used as a food supplement alternative to diabetes medicine.

Analysis of Particle Size and Concentration in Die Sinking Electric Discharge Machining.

Electric discharge machining with a powder mix dielectric is a promising technique to harden a work piece's surface using electricity with a high energy density. The quality of the electrical discharge-machined surface is related to its surface integrity in which the surface's roughness, residual stresses, micro hardness and surface micro cracks are some of the major factors. In this research, graphite powder was mixed in a dielectric with a particle size of 20 µm, 30 µm, and 40 µm, with the concentration of the graphite powder ranging from 2 g/L to 4 g/L. Moreover, the peak current and pulse time on were also coupled with an additive of graphite powder to investigate the effect on the surface quality, i.e., the recast layer thickness, micro hardness and crater depth as well as the material removal rate (MRR) and tool wear rate (TWR). A Box-Behnken design was employed to design the experiments and the experimental results revealed that the graphite powder size and concentration coupled with the electrical parameters (peak current and pulse time on) significantly influenced the recast layer thickness, micro hardness, crater size, MRR and TWR. The crater depth and micro hardness were maximized at a higher concentration and particle size, while the recast layer thickness was reduced with a higher gain size.

Shrimp peptide hydrolysate modulates the immune response in cyclophosphamide immunosuppressed mice model.

Bioactive peptides are naturally found in various foods and were shown to have various distinct physiological as well as medicinal benefits. In this study shrimp peptide hydrolysate (SPH) was prepared to investigate its immunomodulatory effect against cyclophosphamide (CTX) induced immunosuppressed mice. The SPH effect was also analyzed on murine macrophage (RAW264.7 cells). The findings show that SPH stimulates macrophages to form multiple pseudopodia, has no cytotoxic effect, and increases phagocytic activity in RAW264.7 cells. Furthermore, the immunosuppressed in-vivo model illustrates the improvement in various aspects, that is body weight, escalation in immune organ index, and ameliorates histopathological transformation of thymus along with the spleen. SPH enhances cell-mediated immunity by facilitating splenocyte proliferation and inhibit excessive apoptosis. Moreover, the significant outcome had been observed with the upregulation of cytokines interferon-gamma (IFN-ϒ), interleukin-2 (IL-2) level and simultaneously downregulate certain genes include interleukin-4 (IL-4) and interleukin-10 (IL-10). Additionally, SPH expedites cellular immunity by enhancing the regulation of immunoglobulin A (IgA) and immunoglobulin M (IgM). However, these findings support the hypothesis that SPH is an effective immunomodulatory agent capable of preventing immune system hypofunction. It is necessary to investigate the detailed mechanism to rule out any unforeseen effects of SPH in future research. PRACTICAL APPLICATIONS: Chemotherapy medications, despite their dominating detrimental effects of damaging immunological organs such as the spleen and thymus, extend the treatment process as well as the destruction of the self-immune system. This study found that SPH is an effective immunomodulatory agent capable of avoiding immune organ hypofunction and improving cell mediate immunity by enhancing macrophage activation, phagocytosis, spleenocyte proliferation, suppressing apoptosis, and elevating cytokines and antibodies. As a result, SPH can be utilized as a nutritional and functional dietary supplement to boost immunological modulation in combination with chemotherapy medications in order to lessen their adverse effects.