Limosilactobacillus fermentum NCU003089 and Lactiplantibacillus plantarum NCU001261, two probiotics with inhibition of Escherichia coli and Cronobacter sakazakii translocation in vitro.

The subject of this study was to screen lactic acid bacteria (LAB) with pathogen translocation inhibition and investigate the potential inhibition mechanism of it. Pathogens colonized in the intestine could cross the intestinal barrier to access blood circulation, causing severe complications. This study aimed to screen LAB with favorable inhibitory effects on the translocation of enterinvasive Escherichia coli CMCC44305 (E. coli) and Cronobacter sakazakii CMCC45401 (C. sakazakii), which were two common intestinal opportunistic pathogens. After an elaborate screening procedure including adhesion, antibacterial, and translocation assay, Limosilactobacillus fermentum NCU003089 (L. fermentum NCU3089) and Lactiplantibacillus plantarum NCU0011261 (L. plantarum NCU1261) were found to inhibit 58.38% and 66.85% of pathogen translocation, respectively. Subsequently, LAB pre-treatment suppressed the decline in TEER of Caco-2 monolayers caused by pathogens. Meanwhile, L. fermentum NCU3089 significantly inhibited claudin-1, ZO-1, and JAM-1 degradation caused by E. coli, and L. plantarum NCU1261 markedly reduced claudin-1 degradation caused by C. sakazakii. Also, the two LAB strains significantly decreased TNF-α level. In addition, L. fermentum NCU3089 but not L. plantarum NCU1261 tolerated well in the gastrointestinal fluids, and they were both sensitive or intermediate to nine common clinical antibiotics without hemolytic activity. In short, the two LAB strains could inhibit pathogen translocation by competing for adhesion sites, secreting antibacterial substances, reducing inflammatory cytokines levels, and maintaining intestinal barrier integrity. This study provided a feasible solution to prevent pathogen infection and translocation, and the two LAB strains were safe and had potential in food and pharmaceutical applications.

The lactic acid bacteria and yeast community of home-made sauerkraut from three provinces in Southwest China.

The aim of this study was to investigate the lactic acid bacteria (LAB) and yeast community from home-made sauerkraut collected from Southwest China through culture-dependent and culture-independent technology. Forty-eight samples of home-made sauerkraut were collected from households at three different locations in Southwest China. The pH, total acidity and salt contents among these fermented vegetables were 3.69 ± 0.42, 0.86 ± 0.43 g/100 ml, and 3.86 ± 2.55 g/100 ml, respectively. The number of lactic acid bacteria (LAB) and yeasts were 7.25 ± 1.05 log10 colony-forming units (CFU)/ml and 3.74 ± 1.01 log CFU/ml, respectively. A total of 182 LAB and 81 yeast isolates were identified. The dominant isolates were Lactobacillus plantarum, L. brevis, Pediococcus ethanolidurans, Pichia membranifaciens, P. fermentans and Kazachstania bulderi. Denaturing gradient gel electrophoresis (DGGE) showed that L. plantarum, uncultured Lactobacillus sp, P. ethanolidurans, and K. exigua were the predominant microflora. Our studies demonstrated that the DGGE technique combined with a culture-dependent method is very effective for studying the LAB and yeast community in Chinese traditional fermentation vegetables. The results will give us an understanding of LAB and yeast community of Chinese sauerkraut and improve the knowledge of LAB and yeast community of Chinese sauerkraut.

Stability of potential prophages in commercial strain Lactobacillus plantarum NCU116 under various stressors.

Genetic stability of bacterium as a starter culture is vital for product quality in fermentation industry. The commercial strain Lactobacillus plantarum NCU116 widely used in fruit and vegetable fermentation was induced with various stressors to investigate the stability of potential prophages. PHAge Search Tool (PHAST) identified three potential prophages in bacterial genome. By spectrophotometric analysis, mitomycin C (MMC), lactic acid, and bile salt were found to inhibit the growth of L. plantarum NCU116 while ethanol and hydrogen peroxide had no notable impacts. Transcriptions of four phage-synthesizing genes (phaR, phacap, phaada, phatail) and four phage-resistant genes (cas116, helR, hsd1, hsd2) under stressors were investigated by quantitative reverse transcription PCR. MMC was found to most significantly upregulated transcriptions of phage-synthesizing genes, followed by lactic acid and bile salt. By transmission electron microscopy, no virus particles from the lysates of strain NCU116 treated by MMC were observed, corresponding to the result that no phage nucleic acids could be extracted from the supernatants of strain NCU116 treated by MMC. This study suggested that no prophages could be induced from L. plantarum NCU116 by strong inducer MMC, indicating its genetic stability, which supports the comprehensive application of strain NCU116 in industry without causing fermentation failure.

Prioritizing complex disease risk genes by integrating multiple data.

Complex diseases, such as obesity, type II diabetes and chronic obstructive pulmonary disease (COPD) as metabolic disorder-related diseases are major concern for worldwide public health in the 21st century. The identification of these disease risk genes has attracted increasing interest in computational systems biology. In this paper, a novel method was proposed to prioritize disease risk genes (PDRG) by integrating functional annotations, protein interactions and gene expression information to assess similarity between genes in a disease-related metabolic network. The gene prioritization method was successfully carried out for obesity and COPD, the effectiveness of which was superior to those of ToppGene and ToppNet in both literature validation and recall rate by LOOCV. Our method could be applied broadly to other metabolism-related diseases, helping to prioritize novel disease risk genes, and could shed light on diagnosis and effective therapies.

Probiotic-fermented tomato with hepatic lipid metabolism modulation effects: analysis of physicochemical properties, bioactivities, and potential bioactive compounds.

Lactiplantibacillus plantarum NCUH001046 (LP)-fermented tomatoes exhibited the potential to alleviate obesity in our previous study. This subsequent study further delves deeper into the effects of LP fermentation on the physicochemical properties, bioactivities, and hepatic lipid metabolism modulation of tomatoes, as well as the analysis of potential bioactive compounds exerting obesity-alleviating effects. Results showed that after LP fermentation, viable bacterial counts peaked at 9.11 log CFU mL-1 and sugar decreased, while organic acids, umami amino acids, total phenols, and total flavonoids increased. LP fermentation also improved the inhibition capacities of three digestive enzyme activities and Enterobacter cloacae growth, as well as antioxidant activities. Western blot results indicated that fermented tomatoes, especially live probiotic-fermented tomatoes (LFT), showed improved effects compared to unfermented tomatoes in reducing hepatic lipid accumulation by activating the AMPK signal pathway. UHPLC-Q-TOF/MS-based untargeted metabolomics analysis showed that chlorogenic acid, capsiate, tiliroside, irisflorentin, and homoeriodictyol levels increased after fermentation. Subsequent cell culture assays demonstrated that irisflorentin and homoeriodictyol reduced lipid accumulation via enhancing AMPK expression in oleic acid-induced hyperlipidemic HepG2 cells. Furthermore, Spearman's correlation analysis indicated that the five phenols were positively associated with hepatic AMPK pathway activation. Consequently, it could be inferred that the five phenols may be potential bioactive compounds in LFT to alleviate obesity and lipid metabolism disorders. In summary, these findings underscored the transformative potential of LP fermentation in enhancing the bioactive profile of tomatoes and augmenting its capacity to alleviate obesity and lipid metabolism disorders. This study furnished theoretical underpinnings for the functional investigation of probiotic-fermented plant-based foods.

Multi-dimensional characterization of apoptosis in the tumor microenvironment and therapeutic relevance in melanoma.

PURPOSE: Melanoma is widely utilized as a prominent model for the development of immunotherapy, thought an inadequate immune response can occur. Moreover, the development of apoptosis-related therapies and combinations with other therapeutic strategies is impeded by the limited understanding of apoptosis's role within diverse tumor immune microenvironments (TMEs). METHODS: Here, we constructed an apoptosis-related tumor microenvironment signature (ATM) and employ multi-dimensional analysis to understand the roles of apoptosis in tumor microenvironment. We further assessed the clinical applications of ATM in nine independent cohorts, and anticipated the impact of ATM on cellular drug response in cultured cells. RESULTS: Our ATM model exhibits robust performance in survival prediction in multiple melanoma cohorts. Different ATM groups exhibited distinct molecular signatures and biological processes. The low ATM group exhibited significant enrichment in B cell activation-related pathways. What's more, plasma cells showed the lowest ATM score, highlighting their role as pivotal contributors in the ATM model. Mechanistically, the analysis of the interplay between plasma cells and other immune cells elucidated their crucial role in orchestrating an effective anti-tumor immune response. Significantly, the ATM signature exhibited associations with therapeutic efficacy of immune checkpoint blockade and the drug sensitivity of various agents, including FDA-approved and clinically utilized drugs targeting the VEGF signaling pathway. Finally, ATM was associated with tertiary lymphoid structures (TLS), exhibiting stronger patient stratification ability compared to classical "hot tumors". CONCLUSION: Our findings indicate that ATM is a prognostic factor and is associated with the immune response and drug sensitivity in melanoma.

The Circadian Clock Component RORA Increases Immunosurveillance in Melanoma by Inhibiting PD-L1 Expression.

Circadian clock perturbation frequently occurs in cancer and facilitates tumor progression by regulating malignant growth and shaping the immune microenvironment. Emerging evidence has indicated that clock genes are disrupted in melanoma and linked to immune escape. Herein, we found that the expression of retinoic acid receptor-related orphan receptor-α (RORA) is downregulated in melanoma patients and that patients with higher RORA expression have a better prognosis after immunotherapy. Additionally, RORA was significantly positively correlated with T-cell infiltration and recruitment. Overexpression or activation of RORA stimulated cytotoxic T-cell-mediated antitumor responses. RORA bound to the CD274 promoter and formed an inhibitory complex with HDAC3 to suppress PD-L1 expression. In contrast, the DEAD-box helicase family member DDX3X competed with HDAC3 for binding to RORA, and DDX3X overexpression promoted RORA release from the suppressive complex and thereby increased PD-L1 expression to generate an inhibitory immune environment. The combination of a RORA agonist with an anti-CTLA4 antibody synergistically increased T-cell antitumor immunity in vivo. A score based on the combined expression of HDAC3, DDX3X, and RORA correlated with immunotherapy response in melanoma patients. Together, this study elucidates a mechanism of clock component-regulated antitumor immunity, which will help inform the use of immunotherapy and lead to improved outcomes for melanoma patients receiving combined therapeutic treatments. Significance: RORA forms a corepressor complex to inhibit PD-L1 expression and activate antitumor T-cell responses, indicating that RORA is a potential target and predictive biomarker to improve immunotherapy response in melanoma patients.

d-mannose blocks the interaction between keratinocytes and Th17 cells to alleviate psoriasis by inhibiting HIF-1α/CCL20 in mice.

Psoriasis is an autoimmune chronic inflammatory skin disease with an unclear pathogenesis that is difficult to cure, causing serious physical and mental burdens for patients. Previous research showed that a mutually reinforcing vicious cycle caused by keratinocytes (KC) and a variety of immune cells plays an important role in psoriatic inflammation. d-Mannose, a widely distributed metabolite in the body, has been found to treat several metabolic diseases, but its impact on psoriasis remains unknown. Our study aims to investigate the effects of d-mannose on psoriasis and its specific mechanism. Here, we found that d-mannose alleviates psoriasis in mice both as oral and topical agents. Specifically, d-mannose down-regulated the expression of hypoxia-inducible factor 1A(HIF-1α) and inhibited the expression of chemokine CCL20 in keratinocytes, thereby inhibiting the local infiltration of Th17 cells and breaking the cycle of keratinocytes-Th17 cells. Overall, our study indicates that d-mannose alleviates cutaneous inflammation in psoriasis by inhibiting the HIF-1α/CCL20/Th17 cells axis, and d-mannose has the potential to be used as an oral and topical agent in the treatment of psoriasis.

Modulation of the Microbiome-Fat-Liver Axis by Lactic Acid Bacteria: A Potential Alleviated Role in High-Fat-Diet-Induced Obese Mice.

The major characteristics of obesity are abnormal lipid metabolism, chronic inflammation, and imbalanced gut microbiota. It has been reported that lactic acid bacteria (LAB) possess potential for alleviating obesity, considering which the strain-specific functions and diverse mechanisms and the roles and mechanisms of various LAB are worthy of investigation. This study aimed to validate and investigate the alleviating effects and underlying mechanisms of three LAB strains, Lactiplantibacillus plantarum NCUH001046 (LP), Limosilactobacillus reuteri NCUH064003, and Limosilactobacillus fermentum NCUH003068 (LF), in high-fat-diet-induced obese mice. The findings demonstrated that the three strains, particularly LP, suppressed body weight gain and fat deposition; ameliorated lipid disorders, liver and adipocyte morphology, and chronic low-grade inflammation; and reduced lipid synthesis via activating the adenosine 5'-monophosphate-activated protein kinase (AMPK) signaling pathway. In addition, LP and LF decreased the enrichment of bacteria positively correlated with obesity, like Mucispirillum, Olsenella, and Streptococcus, but facilitated the growth of beneficial bacteria negatively correlated with obesity, like Roseburia, Coprococcus, and Bacteroides, along with increasing the short-chain fatty acid levels. It is deduced that the underlying alleviating mechanism of LP was to modulate the hepatic AMPK signaling pathway and gut microbiota by the microbiome-fat-liver axis to alleviate obesity development. In conclusion, as a diet supplement, LP has promising potential in obesity prevention and treatment.

Exploring the association and causal effect between white blood cells and psoriasis using large-scale population data.

Introduction: Psoriasis is a chronic inflammatory disease of the skin. A few studies have shown that psoriasis is an immune-mediated disease in which multiple immune cells play crucial roles. However, the association between circulating immune cells and psoriasis remains elusive. Methods: To explore the role of circulating immune cells in psoriasis, 361,322 individuals from the UK Biobank (UKB) and 3,971 patients with psoriasis from China were included to investigate the association between white blood cells and psoriasis via an observational study. Genome-wide association studies (GWAS) and Mendelian randomization (MR) were used to evaluate the causal relationship between circulating leukocytes and psoriasis. Results: The risk of psoriasis increased with high levels of monocytes, neutrophils, and eosinophils (relative risks and 95% confidence intervals, respectively: 1.430 (1.291-1.584) for monocytes, 1.527 (1.379-1.692) for neutrophils, and 1.417 (1.294-1.551) for eosinophils). Upon further MR analysis, eosinophils showed a definite causal relationship with psoriasis (odds ratio of inverse-variance weighted: 1.386, 95% confidence intervals: 1.092-1.759) and a positive correlation with the psoriasis area and severity index (PASI) score (P = 6.6 × 10-5). The roles of the neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR), and lymphocyte-monocyte ratio (LMR) in psoriasis were also assessed. More than 20,000 genetic variations associated with NLR, PLR, and LMR were discovered in a GWAS analysis using the UKB data. Following adjustment for covariates in the observational study, NLR and PLR were shown to be risk factors for psoriasis, whereas LMR was a protective factor. MR results indicated that there was no causal relationship between these three indicators and psoriasis; however, NLR, PLR, and LMR correlated with the PASI score (NLR: rho = 0.244, P = 2.1 × 10-21; PLR: rho = 0.113, P = 1.4 × 10-5; LMR: rho = -0.242, P = 3.5×10-21). Discussion: Our findings revealed an important association between circulating leukocytes and psoriasis, which is instructive for the clinical practice of psoriasis treatment.

Photothermal Nano-Vaccine Promoting Antigen Presentation and Dendritic Cells Infiltration for Enhanced Immunotherapy of Melanoma via Transdermal Microneedles Delivery.

Immunotherapy has demonstrated the potential to cure melanoma, while the current response rate is still unsatisfactory in clinics. Extensive evidence indicates the correlation between the efficacy and pre-existing T-cell in tumors, whereas the baseline T-cell infiltration is lacking in low-response melanoma patients. Herein, we demonstrated the critical contribution of dendritic cells (DCs) on melanoma survival and baseline T-cell level, as well as the efficacy of immunotherapy. Capitalized on this fact, we developed a photothermal nano-vaccine to simultaneously promote tumor antigens presentation and DCs infiltration for enhanced immunotherapy. The nano-vaccine was composed of polyserotonin (PST) core and tannic acid (TA)/Mn2+ coordination-based metal-organic-framework (MOF) shell for ß-catenin silencing DNAzyme loading, which was further integrated into dissolving microneedles to allow noninvasive and transdermal administration at melanoma skin. The nano-vaccine could rapidly penetrate skin upon microneedles insertion and exert a synergistically amplified photothermal effect to induce immunogenic cell death (ICD). The MOF shell then dissociated and released Mn2+ as a cofactor to self-activate DNAzyme for ß-catenin suppression, which in turn caused a persistent CCL4 excretion to promote the infiltration of DCs into the tumor. Meanwhile, the liberated PST core could effectively capture and facilitate tumor antigens presentation to DCs. As a result, potent antitumor efficacies were achieved for both primary and distal tumors without any extra treatment, indicating the great promise of such a nano-vaccine for on-demand personalized immunotherapy of melanoma.

Integration of SNP Disease Association, eQTL, and Enrichment Analyses to Identify Risk SNPs and Susceptibility Genes in Chronic Obstructive Pulmonary Disease.

Chronic obstructive pulmonary disease (COPD) is a complex disease caused by the disturbance of genetic and environmental factors. Single-nucleotide polymorphisms (SNPs) play a vital role in the genetic dissection of complex diseases. In-depth analysis of SNP-related information could recognize disease-associated biomarkers and further uncover the genetic mechanism of complex diseases. Risk-related variants might act on the disease by affecting gene expression and gene function. Through integrating SNP disease association study and expression quantitative trait loci (eQTL) analysis, as well as functional enrichment of containing known causal genes, four risk SNPs and four corresponding susceptibility genes were identified utilizing next-generation sequencing (NGS) data of COPD. Of the four risk SNPs, one could be found in the SNPedia database that stored disease-related SNPs and has been linked to a disease in the literature. Four genes showed significant differences from the perspective of normal/disease or variant/nonvariant samples, as well as the high performance of sample classification. It is speculated that the four susceptibility genes could be used as biomarkers of COPD. Furthermore, three of our susceptibility genes have been confirmed in the literature to be associated with COPD. Among them, two genes had an impact on the significance of expression correlation of known causal genes they interact with, respectively. Overall, this research may present novel insights into the diagnosis and pathogenesis of COPD and susceptibility gene identification of other complex diseases.

Screening, Safety Evaluation, and Mechanism of Two Lactobacillus fermentum Strains in Reducing the Translocation of Staphylococcus aureus in the Caco-2 Monolayer Model.

Staphylococcus aureus is a common commensal of humans, and its translocation from gastrointestine to peripheral organs and tissues could cause severe diseases and complications. This study focuses on the screening and characterization of Lactobacillus strains with significant inhibitory effect on the translocation of S. aureus through Caco-2 monolayers. First, strains with strong affinity for mucin and Caco-2 cells were obtained, via microtiter plate assay and adhesion assay, respectively. Obtained bacteria were further tested for their inhibitory effects on the growth of S. aureus by well diffusion assay. Subsequently, two strains preincubated with Caco-2 monolayers were found to inhibit the translocation of S. aureus CMCC26003 by 80.95 and 43.96%, respectively, via the transcellular translocation assay. These two strains were then identified to be Lactobacillus fermentum NCU3087 and L. fermentum NCU3088. Second, the mechanism of inhibition was investigated by analyzing the relative concentration of tight junction proteins and proinflammatory cytokines of Caco-2 cells, by Western blot and enzyme-linked immunosorbent assay, respectively. Results showed that both NCU3087 and NCU3088 significantly attenuated the degradation of occludin, claudin-1, ZO-1, and JAM-1 and suppressed the secretion of interleukin 6 and tumor necrosis factor-α induced by S. aureus, to different extent. Moreover, two Lactobacillus strains could barely translocate the Caco-2 monolayers, had no hemolytic activity, displayed strong resistance to gastrointestinal fluids, and were sensitive or moderate sensitive to nine clinically relevant antibiotics. Collectively, this study identified two Lactobacillus strains with significant inhibitory effect on the translocation of S. aureus, and their safeness for application was evaluated, thereby providing potential solutions for prevention of S. aureus and prophylaxis of related diseases.

A susceptibility biomarker identification strategy based on significantly differentially expressed ceRNA triplets for ischemic cardiomyopathy.

Ischemic cardiomyopathy (ICM) is a common human heart disease that causes death. No effective biomarkers for ICM could be found in existing databases, which is detrimental to the in-depth study of this disease. In the present study, ICM susceptibility biomarkers were identified using a proposed strategy based on RNA-Seq and miRNA-Seq data of ICM and normal samples. Significantly differentially expressed competing endogenous RNA (ceRNA) triplets were constructed using permutation tests and differentially expressed mRNAs, miRNAs and lncRNAs. Candidate ICM susceptible genes were screened out as differentially expressed genes in significantly differentially expressed ceRNA triplets enriched in ICM-related functional classes. Finally, eight ICM susceptibility genes and their significantly correlated lncRNAs with high classification accuracy were identified as ICM susceptibility biomarkers. These biomarkers would contribute to the diagnosis and treatment of ICM. The proposed strategy could be extended to other complex diseases without disease biomarkers in public databases.

Bacterial community and composition in Jiang-shui and Suan-cai revealed by high-throughput sequencing of 16S rRNA.

Fermented vegetables have a long history in many cultures. Jiang-shui and Suan-cai are two of the most well-known instances in North China. They are made by a process of natural lactic acid fermentation. However, they have the different characteristics, i.e. acidity, taste and flavor, which are influenced by the specific bacterial community. Therefore, we used high-throughput sequencing methods to identify the bacterial community structure of Jiang-shui and Suan-cai in this study. Subsequently, we figured out the bacterial differences of these two products using the statistical analysis. Firmicutes and Proteobacteria were the dominant phyla in both Jiang-shui and Suan-cai. However, Lactobacillus was the main genus in Jiang-shui samples, whereas both Lactobacillus and Pediococcus were the major genera in the Suan-cai samples. At the species level, Lactobacillus amylolyticus, Lactobacillus fermentum and Lactobacillus pontis were the major species in Jiang-shui samples, while Pediococcus parvulus, Lactobacillus coryniformis, Lactobacillus pentosus and Lactobacillus parabrevis were the dominant species in the Suan-cai samples. These results suggested that Jiang-shui and Suan-cai had their own unique bacterial community, leading to the specific characteristics. Furthermore, the bacterial communities of both fermented vegetables varied at different locations. This study revealed the flora present in the Jiang-shui and the Suan-cai, providing a deep insight of the microbial species of Chinese fermented vegetables and guidance for the production of the Jiang-shui and the Suan-cai.