Search Results (137)

Background/Objectives: The hippocampus is an essential part of the central nervous system (CNS); it plays a significant role in social–cognitive memory processing. Prenatal exposure to valproic acid (VPA) can lead to impaired hippocampal functions. In this study, we evaluated the effect of plumbagin (PLB) as a natural product on spatial learning and memory, neuro-morphological changes, and inflammation levels in a VPA-induced autism model during adolescence. Methods: Pregnant Wistar rats received a single intraperitoneal (i.p.) injection of VPA (600 mg/kg) or saline on gestational day 12.5. The male offspring were then categorized and assigned to five groups: Saline+DMSO-, VPA+DMSO-, and VPA+PLB-treated groups at doses of 0.25, 0.5, or 1 mg/kg. Spatial learning and memory were evaluated using the Morris water maze. Histopathological evaluations of the hippocampus were performed using Nissl and hematoxylin–eosin staining, as well as immunofluorescence. The pro-inflammatory cytokine levels were also quantified by quantitative real-time PCR. Results: The findings revealed that a VPA injection on gestational day 12.5 is associated with cognitive impairments in male pups, including a longer escape latency and traveled distance, as well as decreased time spent in the target quadrant. Treatment with PLB significantly enhanced the cognitive function, reduced dark cells, and ameliorated neuronal–morphological alterations in the hippocampus of VPA-exposed rats. Moreover, PLB was found to reduce astrocyte activation and the expression levels of pro-inflammatory cytokines. Conclusions: These findings suggest that PLB partly mitigates VPA-induced cognitive deficits by ameliorating hippocampal inflammation levels. Full article

Fear is a critical welfare concern in laying hens. Fearful behaviors in domestic chickens are influenced by both genetic and environmental factors, contributing to individual differences in stress responses. Tonic immobility (TI) duration is widely recognized as a reliable indicator of fear levels. The hippocampus, a critical brain region for emotional states, plays a pivotal role in associating fearful experiences with specific stimuli, enabling adaptive behavioral responses. This study investigated hippocampal histological characteristics and transcriptomic profiles in laying hens with different fear responses categorized based on TI duration. A total of 80 native Lindian hens (75 weeks old) were individually housed in modified conventional cages. At 76 weeks of age, hens exhibiting the longest and shortest TI durations were classified into the high-fear (TH) and low-fear (TL) groups, respectively. Whole hemibrains were collected for histological and immunohistochemical analyses, while hippocampal tissues underwent transcriptome sequencing. The results showed a significant reduction in Nissl body counts in hippocampal neurons of high-fear hens (p < 0.05), suggesting potential neuronal damage or functional impairment. Transcriptomic analysis revealed 365 differentially expressed genes (DEGs) between two groups, with 277 upregulated and 88 downregulated genes in TH chickens. KEGG pathway enrichment analysis identified seven significantly associated pathways (p < 0.01), including retinol metabolism, vitamin B6 metabolism, and nicotinate and nicotinamide metabolism, all of which are crucial for neuronal function and immune regulation. In addition, a significant increase in DCX protein expression (p < 0.05) and a decrease in c-Fos protein expression (p < 0.05) was noted in in high-fear hens, whereas PCNA levels remained unchanged (p > 0.05) under immunohistochemical validation. The neuronal alterations observed in high fear individuals suggest neural damage, while transcriptomic variations point to potential disruptions in neurogenesis, synaptic signaling, and stress-related pathways. Collectively, these results provide novel insights into the neurobiological basis of fear regulation in laying hens and may have implications for poultry welfare and management strategies. Full article

Background/Objectives: Heparosan is a component of the capsular polysaccharide in Escherichia coli K5 and Pasteurella multocida Type D. It shares a similar glycan structure with heparin and can be enzymatically modified to produce bioactive heparin. Methods: In this study, the probiotic strain E. coli Nissle 1917 (EcN), which naturally produces heparosan, was genetically engineered to utilize sucrose as a carbon source for growth while achieving high-yield heparosan biosynthesis. Results: By expressing the sucrose hydrolase genes sacA (from Bacillus subtilis) or spI (from Bifidobacterium adolescentis), EcN was enabled to utilize sucrose, achieving heparosan titers of 131 mg/L and 179 mg/L, respectively. Further metabolic engineering was performed to block the glycolytic and pentose phosphate pathways, thereby redirecting sucrose-derived glucose-6-phosphate and fructose-6-phosphate toward heparosan biosynthesis, while glycerol was supplemented as an auxiliary carbon source to support cell growth. Finally, the key biosynthesis genes galU, kfiD, and glmM were overexpressed, resulting in an engineered strain with a heparosan titer of 622 mg/L. Conclusions: This study represents the first successful engineering of EcN to utilize sucrose as the carbon source for growth, while achieving enhanced heparosan production through synergistic carbon source utilization. These findings establish a foundational strategy for employing this strain in the sucrose-based biosynthesis of other glycosaminoglycans. Full article

Heparosan, a microbially synthesized capsular polysaccharide, possesses a polysaccharide backbone structurally analogous to heparin. Its biosynthesis holds significant importance for achieving the chemoenzymatic synthesis of heparin. Here, we developed a systematic metabolic engineering strategy in Escherichia coli Nissle 1917 to establish an efficient heparosan production platform. Through the systematic engineering of the glycolytic pathway involving the targeted knockout of zwf, pfkAB, pgi, and fruA (or alternatively fbaA) genes, we generated recombinant strains that lost the capacity to utilize glucose or fructose as sole carbon sources in a minimal medium. This metabolic reprogramming established glycerol as the exclusive carbon source for cell growth, thereby creating a tripartite carbon allocation system, including glycerol for biomass, glucose for UDP-glucuronic acid, and fructose for UDP-N-acetylglucosamine. Therefore, heparosan production was significantly improved from 137.68 mg/L in the wild type to 414.40 mg/L in the recombinant strain. Building upon this foundation, the overexpression of glmM, pgm, and galU genes in the biosynthetic pathway enabled a heparosan titer of 773.78 mg/L in shake-flask cultures. Temporal induction optimization further enhanced titers to 1049.96 mg/L, representing a 7.60-fold enhancement compared to the wild-type strain. This study establishes a triple-carbon-source co-utilization strategy, which holds promising implications for the biosynthesis of heparosan-like microbial polysaccharides. Full article

Depression is associated with bidirectional interactions between inflammatory responses and behavioral dysfunction. Paeoniflorin (PF), a monoterpene glycoside derived from Paeonia lactiflora, exhibits potent anti-inflammatory properties. This study investigates the therapeutic effects of PF on lipopolysaccharide (LPS)-induced depression-like behaviors in mice and neuroinflammation in BV2 microglial cells. Mice were co-administered PF (20, 40, or 80 mg/kg/day) and LPS (2 mg/kg) for 7 days. Behavioral tests; Nissl staining; and Golgi, Iba1, DLG4, and cytokine assays were conducted. Additionally, hippocampal NF-κB, Nrf2, and BDNF signaling pathways were analyzed using Western blots. In BV2 cells, oxidative stress and the Nrf2/HO-1 pathway were assessed using CCK-8, flow cytometry, and Western blotting after 24 h of LPS and PF treatment. PF significantly alleviated LPS-induced depression-like behaviors, increased hippocampal neuron and dendritic spine density, and upregulated synaptic proteins (PSD95, SNAP25, and BDNF). Mechanistically, PF suppressed NLRP3 inflammasome activation via the Akt/GSK3β pathway, reduced pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6), and enhanced the Nrf2/HO-1 antioxidant axis. In BV2 cells, PF restored mitochondrial membrane potential, inhibited apoptosis, and decreased cytokine levels (TNF-α, IL-1β, and IL-6) by inhibiting TLR4/NF-κB signaling. In conclusion, PF significantly improved LPS-induced depression-like behaviors and attenuated neuroinflammation in BV2 microglial cells, highlighting its potential as a therapeutic agent for inflammation-associated depression. Full article

Memory impairment is a defining characteristic of Alzheimer’s disease (AD), with amnesia often appearing as its earliest symptom. Given the multifactorial nature of AD pathogenesis, this study investigates the multi-target therapeutic potential of sobrerol (coded as NRM-331) in a scopolamine-induced amnesia mouse model, focusing specifically on its effects in ameliorating memory deficits and enhancing neuronal plasticity. Sixty male C57BL/6NCrljOri mice were divided into six groups (10 mice/group): vehicle control (CTL, saline), scopolamine (SPA, 10 mg/kg/day), Aricept (APT, 2 mg/kg/day), and three treatment groups receiving NRM-331 at doses of 40, 80, and 100 mg/kg/day. Several behavioral tests were conducted, including the Y-maze test, passive avoidance test, and Morris water maze test. Additionally, biochemical assays were performed in serum (to measure Aß 1-40 and Aß 1-42) and in the brain (to assess ACh and AChE levels), along with histopathological examination of the brain using Nissl staining and p-tau IHC. No significant change was observed in the Y-maze test or the acquisition trial of the passive avoidance test. However, improvements were noted in the retention trial of the passive avoidance test and the Morris water maze test (including escape latency, swim distance, and number of platform crossed) for the NRM-331 groups compared to the SPA group. Serum levels of Aß 1-40 and Aß 1-42 decreased in the NRM-331 groups compared to the SPA group. In the brain, levels of ACh significantly increased, while AChE levels significantly decreased compared to the SPA group. The number of neuronal cells improved in the CA1, CA3, and DG regions of the hippocampus, as indicated by Nissl staining. A significant reduction in p-tau accumulation was also observed in the NRM-331 groups. In conclusion, NRM-331 demonstrated an anti-amnesic effect by enhancing hippocampal cholinergic signaling, alongside exhibiting anti-tau and anti-Aβ synthesis properties. These therapeutic effects suggest that NRM-331 significantly mitigates memory impairment induced by SPA through a neuroprotective mechanism. Full article

Outer membrane vesicles (OMVs) are extracellular vesicles secreted by Gram-negative bacteria with diameters of 20–250 nm. OMVs contain various biologically active substances from their parent bacteria, such as proteins, lipids, and nucleic acids. Escherichia coli Nissle 1917 (EcN) is a Gram-negative probiotic that resides in the human intestine. EcN-derived OMVs are pivotal in modulating intestinal immune responses. However, few studies have addressed the heterogeneity of EcN-derived OMVs in terms of size, significantly limiting the research on their clinical applications. Currently, there are a lack of feasible methods for obtaining EcN-derived OMVs of different sizes. To address this knowledge gap, we developed a membrane filtration method to isolate EcN-derived OMVs of varying sizes. In this study, we first used gradient filtration to isolate high-purity EcN-derived OMVs and conducted a proteomic analysis. Subsequently, we used membrane filtration to separate the EcN-derived OMVs by size. We successfully obtained EcN-derived OMVs of three specific sizes: <50 nm, 50–100 nm, and 100–300 nm. We then performed proteomic analyses of these EcN-derived OMVs and compared their protein profiles. Finally, we compared the ability of each EcN-derived OMV type to induce RAW264.7 macrophages to secrete the pro-inflammatory factor interleukin (IL)-1β and the anti-inflammatory factor IL-10. The EcN-derived OMVs contained 646 different proteins overall; those of different sizes contained different protein types. Among them, the EcN-derived OMVs in the <50 nm group contained significantly fewer proteins (262 different types in total) than those in the 50–100 nm (1603 types) and 100–300 nm (1568 types) groups. Furthermore, the <50 nm group had fewer membrane proteins (40) than the 50–100 nm (215) and 100–300 nm (209) groups. We also found that RAW264.7 macrophages secreted different concentrations of IL-1β and IL-10 following co-incubation with the three EcN-derived OMV types. The 50–100 nm EcN-derived OMV group showed a stronger effect in terms of inducing inflammatory cytokine secretion compared to the other two groups. This study provides direct experimental evidence that EcN-derived OMVs of different sizes exhibit heterogeneous properties. Full article

Neuropsychiatric systemic lupus erythematosus (NPSLE) is a disorder with a poor prognosis characterized by psychiatric and neurological manifestations directly associated with systemic lupus erythematosus (SLE). Neutrophil ferroptosis has been identified as a significant contributor to neutropenia and disease progression in SLE, but its role in NPSLE remains unclear. Female MRL/lpr and MRL/Mpj mice were used. The selective ferroptosis inhibitor liproxstatin-1 and the acyl-CoA synthetase long-chain family member 4 (ACSL4) inhibitor rosiglitazone were administered separately. Assessments included behavioral testing, transmission electron microscopy (TEM), ELISA, Western blotting, RT-PCR, and Nissl staining. Our data showed that neurons in the brain parenchyma undergo ferroptosis, with decreased glutathione peroxidase 4 (GPX4) expression and increased levels of lipid peroxidation indicators and have the typical morphology of ferroptosis confirmed by transmission electron microscopy. Selective ferroptosis inhibitor liproxstatin-1 attenuated the neuropsychiatric manifestations, including depression-like and impulsive behaviors, of MRL/lpr mice. ACSL4 is the main enzyme in lipid metabolism. Our study further found that the utilization of rosiglitazone by inhibiting ACSL4 could also significantly attenuate neuropsychiatric manifestations of MRL/lpr mice. Moreover, blocking ACSL4 might considerably boost GPX4 levels and decrease lipid peroxidation indicators in NPSLE, with reduced neuronal damage, as well as reduced neuroinflammation. This study concluded that inhibiting ACSL4 could facilitate the recuperation of behavioral deficits by suppression of ferroptosis in NPSLE, implying that ACSL4 might be a potential new therapeutic focus for NPSLE. Full article

Amebiasis is a globally prevalent infection that can lead to fatal outcomes if not adequately treated. Conventional treatment with imidazoles often fails due to side effects and resistance, emphasizing the need for alternative therapies. The probiotic Escherichia coli Nissle 1917 (EcN) has shown potential in combating intestinal pathogens. This study aimed to evaluate the amebicidal activity of EcN in vitro and its effect on the production of reactive oxygen species (ROS). Trophozoites of Entamoeba histolytica (2.5 × 10⁴ cells/mL) were cultured in 96-well plates and exposed to varying concentrations of EcN (10²–10⁹ cells/mL). Plates were incubated at 36 °C for 6, 12, and 18 h, after which trophozoite viability was assessed. Intracellular ROS production, including superoxide and hydrogen peroxide, was measured using fluorescent probes. The highest efficacy was observed after 18 h at a CFU concentration of 10⁹ cells/mL. Increased ROS production at all probiotic concentrations suggested a role in EcN’s amebicidal mechanism. Morphological changes in trophozoites, such as rounding, vacuolization, and size reduction, were noted after EcN exposure, indicating growth inhibition. These findings suggest EcN induces structural and morphological changes in E. histolytica, inhibiting its growth in vitro. The findings suggest the potential efficacy of EcN; however, definitive confirmation requires data from human clinical trials. Full article

Background and Objectives: The administration of oral vaccines offers a potential strategy for cancer immunotherapy; yet, the development of effective platforms continues to pose a difficulty. This study examines Escherichia coli Nissle 1917 (EcN) as a microbial vector for the precise delivery of Glypican-1 (GPC1), a tumor-associated antigen significantly overexpressed in pancreatic ductal adenocarcinoma (PDAC).To evaluate the effectiveness of EcN as a vector for the delivery of GPC1 and assess its potential as an oral vaccination platform for cancer immunotherapy. Materials and Methods: EcN was genetically modified to produce a GPC1-flagellin fusion protein (GPC1-FL) to augment antigen immunogenicity. The expression and stability of GPC1 were confirmed in modified PANC02 cells using Western blot and flow cytometry, indicating that GPC1 expression did not influence tumor cell growth. A mouse model was employed to test immunogenicity post-oral delivery, measuring systemic IgG, IL-10, IL-2, and IFN-γ levels to indicate immune activation. Results: Oral immunization with EcN GPC1-FL elicited a robust systemic immune response, demonstrated by markedly increased levels of IgG and IL-10. IL-2 and IFN-γ concentrations were elevated in vaccinated mice relative to controls; however, the differences lacked statistical significance. Western blot examination of fecal samples verified consistent antigen expression in the gastrointestinal tract, indicating effective bacterial colonization and antigen retention. No detrimental impacts were noted, hence substantiating the safety of this methodology. Conclusions: These findings confirm EcN as a feasible and patient-friendly oral vaccination platform for cancer immunotherapy. The effective production of GPC1 in tumor cells, along with continuous antigen delivery and immune activation, underscores the promise of this approach for PDAC and other cancers. This study promotes microbial-based antigen delivery as a scalable, non-invasive substitute for traditional vaccine platforms. Full article

Circadian rhythm disruption is a modifiable risk factor for Alzheimer’s disease (AD) progression, marked by neuroinflammation, oxidative stress, and amyloid-β (Aβ) accumulation. Hypoxia-inducible factor 3α (Hif3α) has emerged as a key regulator of inflammatory and oxidative pathways. To evaluate the impacts of circadian disruption on AD progression and investigate the therapeutic potential of bamboo leaf flavonoids (BLFs), C57BL/6N mice (normal mice) and APP/PS1 transgenic mice (AD mice) were exposed to circadian disruption via randomized light exposure and stress, as the in vivo model. Then, BLFs were administered to assess effects on neuroinflammation, oxidative stress, and organ damage. Next, Nissl body staining and Aβ protein immunohistochemistry were performed to evaluate the effects of BLFs on brain pathology. Through transcriptome sequencing, key factors and the related pathway were screened out. In vitro, molecular mechanisms were explored in PC12 cells treated with Aβ42 and Hif3α siRNA fragments. Results demonstrated that circadian disruption increased oxidative stress and early liver and kidney damage degrees, with greater severity in AD mice. BLFs partially reversed oxidative damage and reduced Aβ deposition. Transcriptome analysis revealed upregulation of Hif3α in circadian-disrupted mice, linked to inflammation and oxidative stress. In vitro, the knockdown of Hif3α reduced inflammation and normalized protein expression, which could be regulated by BLFs and suppressed AD progression. In conclusion, circadian disruption exacerbated AD progression via regulating Hif3α/Rab7/TNFα/IL1β pathway. BLFs offered neuroprotection roles by mitigating inflammation and oxidative damage, highlighting Hif3α as a promising target for AD therapy and biomarker development. Full article

Background: Microplastics (MPs) are small plastic fragments with diameters less than 5 mm in size and are prevalent in everyday essentials and consumables. Large global plastic production has now led to a flooding of MPs in our natural environment. Due to their detrimental impacts on the planet’s ecosystems and potentially our health, MPs have emerged as a significant public health concern. In this pilot study, we hypothesize that MPs exposure will negatively affect gut microbiota composition and function, in which metabolic reprogramming plays an important role. Methods: Using in vitro experiments, three bacterial strains (Escherichia coli MG1655, Nissle 1917, and Lactobacillus rhamnosus) were selected to investigate the impacts of MPs exposure. The bacterial strains were individually cultured in an anaerobic chamber and exposed to 1 µm polystyrene MPs at various concentrations (0, 10, 20, 50, 100, and 500 µg/mL) in the culture medium. Results: MPs exposure reduced the growth of all three bacterial strains in a dose-dependent manner. Liquid chromatography mass spectrometry (LC-MS)-based untargeted metabolomics revealed significant differences in multiple metabolic pathways, such as sulfur metabolism and amino sugar and nucleotide sugar metabolism. In addition, we extracted gut microbiota from C57BL/6 mice, and 16S rRNA sequencing results showed a significant upregulation of Lactobacillales and a significant reduction in Erysipelotrichales due to MPs exposure. Furthermore, targeted and untargeted metabolomics corroborated the in vitro results and revealed alterations in microbial tryptophan metabolism and energy producing pathways, such as glycolysis/gluconeogenesis and the pentose phosphate pathway. Conclusions: These findings provide evidence that MPs exposure causes comprehensive changes to healthy gut microbiota, which may also provide insights into the mechanistic effects of MPs exposure in humans. Full article

Plastics are indispensable to modern life, but their widespread use has created an environmental crisis due to inefficient waste management. Mixed plastic waste, comprising diverse polymers, presents significant recycling challenges due to the high costs of sorting and processing, leading to ecosystem accumulation and harmful by-product generation. This study addresses this issue by engineering a synthetic bacterial consortium (SBC) designed to degrade mixed plastic monomers. The consortium pairs Escherichia coli Nissle 1917, which uses ethylene glycol (EG), a monomer derived from polyethylene terephthalate (PET), as a carbon source, with Pseudomonas putida KT2440, which metabolizes hexamethylenediamine (HD), a monomer from nylon-6,6, as a nitrogen source. Adaptive evolution of the SBC revealed a novel metabolic interaction where P. putida developed the ability to degrade both EG and HD, while E. coli played a critical role in degrading glycolate, mitigating its by-product toxicity. The evolved cross-feeding pattern enhanced biomass production, metabolic efficiency, and community stability compared to monocultures. The consortium’s performance was validated through flux balance analysis (FBA), high-performance liquid chromatography (HPLC), and growth assays. These findings highlight the potential of cross-feeding SBCs in addressing complex plastic waste, offering a promising avenue for sustainable bioremediation and advancing future polymer degradation strategies. Full article

Resveratrol, a plant-derived polyphenol, exhibits significant antidepressant effects and notably enhances neuroplasticity in neurological diseases. However, whether the antidepressant function of resveratrol is related to neuroplasticity remains uncertain, and the underlying mechanisms is poorly understood. This study aims to investigate the role and mechanism of resveratrol in neuroplasticity in depression. Here, we adopted the chronic unpredictable mild stress (CUMS) model and resveratrol intervention by oral gavage. Thereafter, behavioral tests confirmed resveratrol’s antidepressant effect, and Nissl staining, Golgi staining, and Western blotting (WB) were employed to assess the neuronal plasticity. Moreover, proteomic analysis and WB were used to screen and identify the key proteins. To investigate the downstream target of ELAV-like RNA-binding protein 4 (ELAVL4) (one of candidate genes), the RNA Interactome Database and the National Center for Biotechnology Information databases were utilized to predict the targets of ELAVL4. Finally, Quantitative PCR, WB, and Immunofluorescence were used to verify the prediction. Our results indicate that resveratrol alleviates CUMS-induced depressive-like behaviors accompanied by the restoration of impaired hippocampal neuroplasticity. Then, proteomic analysis shows that 351 differentially expressed proteins (DEPs) decrease after CUMS, while 24 DEPs increase remarkably with the resveratrol treatment. Among which, ELAVL4 is downregulated by CUMS, simultaneously increasing after resveratrol intervention, which acts as a protective protein in this process. Finally, brain-derived neurotrophic factor (Bdnf) mRNA is predicted to be the potential target of ELAVL4 and validated by molecular technologies. In conclusion, our findings demonstrate that resveratrol’s antidepressant efficacy is closely associated with ELAVL4, an RNA-binding protein, a mediated neuroplasticity pathway, potentially intersecting with the Bdnf mRNA. Overall, this research sheds light on the role of the ELAVL4-Bdnf mRNA pathway through neuroplasticity in resveratrol’s antidepressant action, which provides an mRNA regulation perspective for the development of novel antidepressants and understanding depression pathology. Full article

Probiotic bacteria are normal inhabitants of a healthy human gut, conferring multiple beneficial effects on the gut and beyond. Under various disease states, the abundance and diversity of beneficial bacteria are significantly decreased, a process called dysbiosis. Among the intra- and extracellular components of probiotics, the extracellular vesicles (EVs) secreted by them have recently garnered significant attention as potential mediators of probiotics’ effects on host health. Further, these nanosized particles that encapsulate a wide range of bioactive molecules (proteins, lipids, RNA, and DNA) are standing out as key factors that could mediate gut microbiota–host communication and confer ameliorating effects in experimental inflammatory, metabolic, and cardiovascular disease models. However, a standard protocol of EV isolation from probiotic bacteria, not varying from lab to lab, must be established to achieve consistency in the experimental results in these pre-clinical models. Our current study compared two commonly used methods for EV isolation from biological samples, ultracentrifugation and precipitation, to develop a standard protocol for isolating EVs from the probiotics Lactobacillus acidophilus (LA), a Gram-positive bacterium, and Escherichia coli Nissle (EcN), a Gram-negative bacterium. The ultracentrifugation method gave ~1.5-fold higher EV yield for both LA and EcN compared to the precipitation method. Further, EcN released a higher level of EVs compared to LA. EVs were quantified and characterized by nanoparticle-tracking analysis (NTA) and by measuring the specific surface biomarkers using Western blot. Here, we describe our standardized step-by-step protocol for isolating EVs from probiotic bacteria and their characterization. Full article