Synthesis, taste characteristics and taste mechanism of N-lactoyl leucine from soy sauce using sensory analysis and UPLC-MS/MS.

Recently, amino acid derivatives gradually gained attention, but studies on N-lactoyl-leucine (Lac-Leu) and N-lactoyl-isoleucine (Lac-Ile) are limited. This study aims to explore the contributions of Lac-Leu and Lac-Ile to soy sauce. Lac-Leu and Lac-Ile were synthesized via enzymatic synthesis method catalyzed by Tgase. The mixed solutions containing Lac-Leu were found to have greater taste improvement than those containing Lac-Ile. Sensory evaluation indicated the sour, bitter, and astringent taste of Lac-Leu in water as well as its kokumi, astringent, and umami-enhancing taste in MSG solution. The taste threshold and umami-enhancing threshold of Lac-Leu measured by TDA and cTDA, respectively, were 0.08 mg/mL and 0.16 mg/mL. Molecular docking of Lac-Leu and Lac-Ile with the kokumi receptor CaSR and the umami receptors T1R1 and T1R3 indicated that Lac-Leu had higher affinities with receptors than Lac-Ile. These findings demonstrated the underlying contribution Lac-Leu made to soy sauce, indicating its potential to improve the flavor quality of soy sauce.

Circulating TIGIT±PD1+TPH, TIGIT ± PD1+TFH cells are elevated and their predicting role in systemic lupus erythematosus.

It is well established that increased peripheral helper T cells (TPH) and follicular helper T cells (TFH) was found in systemic lupus erythematosus (SLE) patients. However, the expression patterns and immunomodulatory roles of TIGIT and PD1 on TPH/TFH in SLE are poorly understood. The expression patterns of TIGIT and PD1 on TPH and TFH cells were examined using flow cytometry and their expression patterns in SLE patients were then further evaluated for their correlation with auto-antibodies, disease activity and severity, B cell differentiation. Logistic regression was used to analyze the risk factors. And the receiver operating characteristic curves and logistic regression model were created to evaluate the predicting role in SLE. TIGIT±PD1+TPH, TIGIT±PD1+TFH cells in the peripheral blood of SLE patients were upregulated, whereas TIGIT+PD1-TFH was downregulated. TIGIT ± PD1+TPH, TIGIT ± PD1+TFH cells positively correlated with auto-antibodies production, disease activity and severity, whereas TIGIT+PD1-TFH cells negatively correlated. TIGIT ± PD1+TPH, TIGIT-PD1+TFH were positively correlated with the frequency of plasmablasts. Furthermore, higher TIGIT+PD1+TPH and TIGIT+PD1+TFH were shown to be risk factors for SLE, whereas TIGIT+PD1-TFH was found to be a protective factor, according to logistic regression analysis. A further logistic regression model showed that combination of TPH/TFH and routine blood indicators may has potential predicting value for SLE, with AUC of 0.957. The increased TIGIT ± PD1+TPH, increased TIGIT ± PD1+TFH, decreased TIGIT+PD1-TFH correlates with disease severity and activity, may boost our comprehending of the role of TIGIT and PD1 on TPH/TFH in SLE, and a logistic regression model based on combination of TPH/TFH and routine blood indicators shows prominent value for predicting SLE.

Neutrophil lncRNA ZNF100-6:2 is a potential diagnostic marker for active pulmonary tuberculosis.

Active pulmonary tuberculosis (PTB) poses challenges in rapid diagnosis within complex clinical conditions. Given the close association between neutrophils and tuberculosis, we explored differentially expressed long non-coding RNAs (lncRNAs) in neutrophils as potential molecular markers for diagnosing active PTB. We employed a gene microarray to screen for lncRNA alterations in neutrophil samples from three patients with active PTB and three healthy controls. The results revealed differential expression of 1457 lncRNAs between the two groups, with 916 lncRNAs upregulated and 541 lncRNAs down-regulated in tuberculosis patients. Subsequent validation tests demonstrated down-regulation of lncRNA ZNF100-6:2 in patients with active PTB, which was restored following anti-tuberculosis treatment. Our findings further indicated a high diagnostic potential for lncRNA ZNF100-6:2, as evidenced by an area under the receiver operating characteristic (ROC) curve of 0.9796 (95% confidence interval: 0.9479 to 1.000; P < 0.0001). This study proposes lncRNA ZNF100-6:2 as a promising and novel diagnostic biomarker for active PTB.

Using combination of albumin to fibrinogen ratio and prognostic nutritional index model for predicting disease activity in patients with systemic lupus erythematosus.

Background: Systemic lupus erythematosus (SLE) is chronic autoimmune disease with multiple organ damage and is associated with poor prognosis and high mortality. Identification of universal biomarkers to predict SLE activity is challenging due to the heterogeneity of the disease. This study aimed to identify the indicators that are sensitive and specific to predict activity of SLE.Methods: We retrospectively analyzed 108 patients with SLE. Patients were categorized into SLE with activity and without activity groups on the basis of SLE disease activity index. We analyzed the potential of routine and novel indicators in predicting the SLE activity using receiver operating characteristic curves and multivariate logistic regression. The Spearman method was used to understand the correlation between albumin to fibrinogen ratio (AFR), prognostic nutritional index (PNI), AFR-PNI model and disease activity.Results: SLE with activity group had higher ESR, CRP, D-dimer, fibrinogen, CRP to albumin ratio, positive rate of anti-dsDNA and ANUA, and lower C3, total bilirubin, total protein, albumin, albumin/globulin, creatinine, high density liptein cholesterol, hemoglobin, hematocrit, lymphocyte count, positive rate of anti-SSA, AFR, PNI than SLE without activity. A further established model based on combination of AFR and PNI (AFR-PNI model) showed prominent value in distinguishing SLE with activity patients from SLE without activity patients. In addition, the sensitivity and specificity of AFR-PNI model + anti-dsDNA combination model were superior to AFR-PNI model. AFR and PNI were risk factors for SLE activity. Moreover, AFR+PNI model correlated with disease activity and AFR-PNI model was associated with fever, pleurisy, pericarditis, renal involvement.Conclusion: These findings suggest that predictive model based on combination of AFR and PNI may be useful markers to identify active SLE in clinical practice.

Comprehensive Analysis of circRNA, lncRNA, miRNA and mRNA Expression Profiles and Their Competing Endogenous RNA Networks in Hepatitis B Virus-Related Hepatocellular Carcinoma.

Pursuing knowledge about circular RNA (circRNA), long non-coding RNA (lncRNA), microRNA (miRNA), and messenger RNA (mRNA) expression profiles and their competing endogenous RNA (ceRNA) networks in hepatitis B virus-related hepatocellular carcinoma (HBV-related HCC) was the focus of this research. Expression patterns of circRNAs, lncRNAs, miRNAs, and mRNAs were searched for in relation to HBV-related HCC using whole-transcriptome sequencing. The expression levels of chosen circRNA, lncRNA, miRNA, and mRNA were analyzed using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The potential connections and roles of ceRNA were deduced via bioinformatics research. The sum of 284 circRNAs, 2,927 lncRNAs, 693 miRNAs, and 5566 mRNAs were discovered to be expressed at considerably different levels in HBV-related HCC tissue and adjacent normal tissue. And the most significantly up- and down-regulated circRNAs, lncRNAs, miRNAs, and mRNAs were verified in HBV-related HCC by qRT-PCR. The circRNA/miRNA/mRNA and lncRNA/miRNA/mRNA networks of HBV-related HCC were established, and the ceRNA regulatory networks revealed the gene expression mechanisms controlled by ncRNAs. Collectively, we revealed the contribution of various circRNA, lncRNA, miRNA, and mRNA expression profiles and identified their ceRNA regulatory networks in HBV-related HCC, providing a theoretical basis for further exploration.

Decreased expression of NAT10 in peripheral blood mononuclear cells from new-onset ankylosing spondylitis and its clinical significance.

BACKGROUND: NAT10 is the firstly recognized RNA acetyltransferase that participates in multiple cellular biological processes and human disease. However, the role of N-acetyltransferase 10 (NAT10) in ankylosing spondylitis (AS) is still poorly elaborated. METHODS: Fifty-six patients with New-Onset AS, 52 healthy controls (HC), 20 patients with rheumatoid arthritis (RA) and 16 patients with systemic lupus erythematosus (SLE) were recruited from The First Afliated Hospital of Nanchang University, and their clinical characteristics were recorded. The expression level of NAT10 in peripheral blood mononuclear cell (PBMC) was examined using reverse transcription-quantitative PCR analysis. The correlations between the expression level of NAT10 in the New-Onset AS patients and disease activity of AS were examined, and receiver operating characteristic (ROC) curves were built to evaluate predictive value in AS. Univariate analysis and multivariate regression analysis were used to analyze the risk factors and construct predictive model. RESULTS: The mRNA expressions of NAT10 in PBMC from new-onset AS patients were significantly low and there were negative correlation between mRNA NAT10 and ASDAS-CRP, BASDIA in new-onset AS patients. ROC analysis suggested that mRNA NAT10 has value in distinguishing new-onset AS patients from HC, RA and SLE. Furthermore, a novel predictive model based on mRNA NAT10 and neutrophil percentages (N%) was constructed for distinguishing new-onset AS patients from HC (AUC = 0.880, sensitivity = 84.62%, specificity = 76.92%) and the predictive model correlated with the activity of new-onset AS. Furthermore, the predictive model could distinguish new-onset AS patients from RA and SLE (AUC = 0.661, sensitivity = 90.38%, specificity = 47.22%). Moreover, the potential predictive value of the combination of predictive model-HLA-B27 for AS vs. HC with a sensitivity of 92.86% (39/42), a specificity of 100.00% (52/52) and an accuracy of 96.81% (91/94) was superior to that of HLA-B27, which in turn had a sensitivity of 84.44% (38/45), a specificity of 100.00% (52/52) and an accuracy of 92.78% (90/97). CONCLUSION: The present study suggested that the decreased mRNA NAT10 may play a role in AS pathogenesis and predictive model based on mRNA NAT10 and N% act as bioindicator for forecast and progression of diseases.

Six categories of amino acid derivatives with potential taste contributions: a review of studies on soy sauce.

During the fermentation of soy sauce, the metabolism of microorganisms and the Maillard reaction produce a wide variety of metabolites that contribute to the unique and rich flavor characteristics of soy sauce, such as amino acids, organic acids and peptides. Amino acid derivatives, a relatively new taste compounds, formed by the reaction of enzymes or non-enzymes from sugars, amino acids, and organic acids released through metabolism by microorganisms during soy sauce fermentation, have begun to gain more and more attention in recent years. This review focused on our existing knowledge of the sources, taste characteristics and synthesis methods of the 6 categories of amino acid derivatives, including Amadori compounds, γ-glutamyl peptides, pyroglutamyl amino acids, N-lactoyl amino acids, N-acetyl amino acids and N-succinyl amino acids. Sixty-four amino acid derivatives were detected in soy sauce, of which 47 were confirmed to have potential contribution to the taste of soy sauce, especially umami and kokumi, and some of them also have the effect of reducing bitterness. Furthermore, some amino acid derivatives, like γ-glutamyl peptides and N-lactoyl amino acids, were found to be synthesized enzymatically in vitro, which laid the foundation for further study on their formation pathways in the future.

Hsa_circ_0044235 and hsa_circ_0001947 as novel biomarkers in plasma of patients with new-onset systemic lupus erythematosus.

Circular RNA (circRNA) are novel types of non-coding RNA that may be used as non-invasive noninvasive biomarkers in clinical plasma samples. However, the role of circRNA in plasma samples from patients with new-onset systemic lupus erythematosus (SLE) has not been extensively investigated. In the present study, reverse transcription-quantitative PCR was used to screen differentially-expressed circRNA (hsa_circ_0000175, hsa_circ_0044235, hsa_circ_0068367, hsa_circ_0002316, hsa_circ_0104871, hsa_circ_0001947, hsa_circ_0001481, hsa_circ_0008675, hsa_circ_0082689 and hsa_circ_0082688) in plasma samples isolated from 22 patients with new-onset SLE and 22 healthy control (HC). The results indicated hsa_circ_0000175, hsa_circ_ 0044235, hsa_circ_0068367, and hsa_circ_0001947 expression levels were significantly lower in plasma samples from new-onset SLE patients compared with corresponding levels in HC subjects and patients with new-onset rheumatoid arthritis (RA). Multivariate analysis indicated expression levels of hsa_circ_0044235 and hsa_circ_0001947 in plasma were independent risk factors for SLE. ROC curve analysis suggested that the combination of hsa_circ_0044235 and hsa_circ_0001947 indicated significant value in discriminating new-onset SLE from HC subjects and patients with RA. Moreover, the levels of hsa_circ_0044235 in plasma samples from patients with new-onset SLE were associated with platelet count, platelet-crit, and platelet distribution width; the expression of hsa_circ_0001947 in plasma from patients with SLE was associated with treatment. Thus, the present study demonstrated a promise for the combination of plasma hsa_circ_0044235 and hsa_circ_0001947 expression as potential diagnostic and prognostic biomarkers in patients with new-onset SLE.

Increased sIL-2Rα leads to obstruction of IL-2 biological function and Treg cells differentiation in SLE patients via binding to IL-2.

Background: The decrease of IL-2 level is believed to play an important role in the disease occurrence and development of SLE, but the relevant mechanisms have not been fully clarified. Many studies have found that the level of soluble interleukin 2 receptor α (sIL-2Rα) in SLE patients is significantly increased. Considering the fact that sIL-2Rα has the ability to bind IL-2, we want to know whether the increased sIL-2Rα has some impact on the level and function of IL-2 in SLE patients. Methods: New onset SLE patients, treated SLE patients and healthy volunteers were recruited. The levels of serum IL-2, IL-2 mRNA in CD3+ T cells and serum sIL-2Rα were detected and compared in these subjects. Two mixed solid-phase sandwich ELISA system were designed to measure exclusively the heterodimers complex of sIL-2Rα/IL-2. The sera from SLE patients were pretreated with or without immune complex dissociation solution and detected for IL-2 levels. IL-2 standard or serum from HCs were used to co-incubate with recombinant sIL-2Rα or serum samples with high levels of sIL-2Rα and detected for IL-2 levels by ELISA. The inhibitory effect of sIL-2Rα on IL-2 biological activity was investigated by CTLL-2 cell proliferation assay. The frequencies and absolute counts of Treg cells were detected by flow cytometry before and after the addition of recombinant sIL-2Rα. Results: The levels of serum IL-2 in SLE patients were significantly decreased and negatively correlated with SLEDAI. However, there was no significant difference in IL-2 mRNA levels in CD3+ T cells between SLE patients and healthy controls. The levels of serum sIL-2Rα in SLE patients were significantly increased, positively correlated with the SLEDAI and negatively correlated with the levels of serum IL-2. sIL-2Rα was shown to bind to IL-2 to form immune complex, resulting in false reduction in the detection level of serum IL-2 and significant decrease in biological activity of IL-2. The increase of sIL-2Rα was demonstrated to be one of the important mechanisms for the obstruction of Treg cells differentiation in SLE patients. Conclusion: Increased serum sIL-2Rα can bind to IL-2, leading to obstruction of IL-2 activity and Treg cells differentiation.

Increased TIGIT+PD­1+CXCR5­CD4+T cells are associated with disease activity in rheumatoid arthritis.

It is well established that increased programmed cell death protein 1 (PD-1)+C-X-C chemokine receptor type 5 (CXCR5)- CD4+T cells are found in patients with rheumatoid arthritis (RA). T cell immunoreceptor with immunoglobulin and ITIM domain (TIGIT) is a co-inhibitory receptor that is expressed on T cells. However, the expression patterns and immunomodulatory roles of TIGIT on PD1+CXCR5-CD4+T cells in RA are poorly understood. Patients with RA were recruited and their clinical characteristics were recorded. The expression level of TIGIT on PD-1+CXCR5-CD4+T cells was examined using flow cytometry. Subsequently, the correlation between various parameters of TIGIT+PD-1+CXCR5-CD4+T cells [percentage of the cells, mean fluorescence intensity (MFI) of PD-1 and TIGIT in the cells] in patients with RA and clinical data, including autoantibodies, inflammatory indicators and RA activity, were analyzed. In addition, the risk factors of RA were assessed using univariate and multivariate regression analyses. The percentages of TIGIT+CXCR5-CD4+T, PD-1+CXCR5-CD4+T, TIGIT+PD-1+CXCR5-CD4+T, TIGIT-PD-1+CXCR5-CD4+T cells, the MFI of PD-1 in these cells and the MFI of TIGIT in TIGIT+PD-1+CXCR5-CD4+T cells were revealed to be significantly increased in patients with RA compared with those in healthy individuals. The parameters of TIGIT+PD-1+CXCR5-CD4+T cells (percentage and/or MFI of PD-1) in patients with RA were found to be associated with the levels of anti-cyclic citrullinated peptide antibodies, rheumatoid factor and inflammatory markers, such as lymphocyte count, lymphocyte percentage, neutrophil percentage, lymphocyte-to-monocyte ratio, neutrophil-to-lymphocyte ratio, systemic immune inflammation index, derived neutrophil-to-lymphocyte ratio, erythrocyte sedimentation rate and C-reactive protein. In addition, the MFI of PD-1 in TIGIT+PD-1+CXCR5-CD4+T cells was associated with a disease activity score of 28 and the patient visual analogue scale. Multivariate logistic regression analysis demonstrated that the percentage of TIGIT+PD-1+CXCR5-CD4+T cells and the MFI of PD-1 in TIGIT+PD-1+CXCR5-CD4+T cells were risk factors for RA. The present study suggests that the increase in TIGIT+PD-1+CXCR5-CD4+T cells is associated with the production of autoantibodies and RA activity and may serve a role in RA pathogenesis.

Platelets correlate with false negative T-SPOT.TB results by inhibiting interferon-γ production in T cells via degranulation.

Background: T-SPOT.TB (T-SPOT) is widely used for the detection of Mycobacterium tuberculosis infection by detecting interferon-gamma (IFN-γ) release in T lymphocytes. This assay is performed on peripheral blood mononuclear cells (PBMCs) separated by Ficoll density gradient centrifugation, which often contain some residual platelets. Here, we investigated the impact of platelets on T-SPOT assay and related mechanisms. Methods: The correlation between platelet count, platelet-to-lymphocyte ratio (PLR), and the IFN-γ secreting T cells (ISCs) in positive control wells of T-SPOT assay were retrospectively analyzed. T-SPOT assay was performed with un-treated PBMCs, platelets-removed PBMCs, and platelets-enriched PBMCs to confirm the impact of platelets on T-SPOT assay. The activation of platelets and their impact on IFN-γ production in T cells were detected by flow cytometry (FCM). Platelets and T cells were cultured in a mixed culture system and co-culture system respectively, followed by detection of the frequencies of IFN-γ-producing T cells and the levels of intracellular IFN-γ in T cells by FCM. Moreover, the effect of platelet releasate on the T-SPOT assay was evaluated. Results: The ISCs in positive control wells of the T-SPOT assay showed a significant decrease with the increase in platelet count. The PLR of the peripheral blood were negatively correlated with the ISCs in positive control wells of the T-SPOT assay. Removal or enrichment of platelets significantly increased or decreased the ISCs and the positive rate of T-SPOT. Inhibition of platelet activation significantly increased the ISCs of T-SPOT. The frequencies of IFN-γ-producing T cells in PBMCs and the levels of intracellular IFN-γ were significantly reduced by the addition of platelets, both in the mixed culture system and the co-culture system. Platelet releasate upon thrombin activation significantly decreased the ISCs of T-SPOT. Conclusions: Platelets correlate with negative T-SPOT results by inhibiting IFN-γ production in T cells via degranulation.

Validation of Serotransferrin in the Serum as Candidate Biomarkers for the Diagnosis of Pulmonary Tuberculosis by Label-Free LC/MS.

This study aimed to identify secreted protein biomarkers in serum from the label-free LC/MS proteomics of neutrophils in pulmonary tuberculosis (TB) patients for the diagnosis biomarkers of TB label-free LC/MS. The proteomic profiles of neutrophils from 15 active TB patients and 15 healthy controls (HCs) were analyzed using label-free LC/MS. We identified 358 differentially expressed proteins preliminarily, including 279 up-regulated proteins and 79 down-regulated proteins. Thirty-eight differentially expressed secreted proteins involved in the progress of platelet degranulation between TB patients and HCs were focused. Of these, serotransferrin (TRF), alpha-2-macroglobulin (AMG), alpha-1-antitrypsin (AAT), alpha-1-acid glycoprotein 1 (AAG), alpha-1-acid glycoprotein 2 (AGP2), and alpha-1B-glycoprotein (A1BG) were selected for further verification in the serum of additional 134 TB patients and 138 HCs by nephelometry and ELISA in the training set. Statistically significant differences of TRF (P < 0.0001), AAT (P < 0.0001), AAG (P < 0.0001), AGP2 (P < 0.0001), and A1BG (P = 0.0003) were observed. The serum concentration of TRF was down-regulated in TB patients compared with healthy controls, which was coincident with the proteomics results. An additional validation of TRF was performed in an independent cohort of patients with active TB (n = 46), patients with lung cancer (n = 37), 20 HCs, and patients with pneumonia (n = 35) in the test set by nephelometry. The serum expression levels of TRF in the TB patients showed lower levels compared with those in patients with pneumonia (P = 0.0125), lung cancer (P = 0.0005), HCs (P < 0.0001), and the non-TB controls (P < 0.0001). Furthermore, the AUC value of TRF was 0.647 with 90.22% sensitivity and 42.86% specificity in discriminating the TB group from the pneumonia group, 0.702 with 93.48% sensitivity and 47.16% specificity in discriminating the TB group from the lung cancer group, 0.894 with 91.30% sensitivity and 71.62% specificity in discriminating the TB group from all HCs, and 0.792 with 91.30% sensitivity and 58.90% specificity in discriminating the TB group from the non-TB controls. This study obtained the proteomic profiles of neutrophils in the TB patients and HCs, which contribute to a better understanding of the pathogenesis molecules existing in the neutrophils of pulmonary tuberculosis and provide candidate biomarkers for the diagnosis of pulmonary tuberculosis.

Combined identification of three lncRNAs in serum as effective diagnostic and prognostic biomarkers for hepatitis B virus-related hepatocellular carcinoma.

Hepatitis B virus-related hepatocellular carcinoma (HBV-related HCC) is a common, highly invasive malignant tumor associated with a high mortality rate. This study aimed to identify the effective diagnostic and prognostic biomarkers for HBV-related HCC. With HBV-related HCC RNA-sequencing data of The Cancer Genome Atlas (TCGA) database, 159 differentially expressed long noncoding RNAs (lncRNAs) between HBV-related HCC and para-carcinoma normal samples were identified, and 12 lncRNAs were eventually assessed for deeper research. Classification analysis developed a three-lncRNA signature of AC005332.5, ELF3-AS1 and LINC00665, which was demonstrated to be the most discriminatory with an AUC (Area under the curve) value of 0.913 (95% CI: 0.8610-0.9665) and verified in validation patients. The expression levels of AC005332.5, ELF3-AS1 and LINC00665 were significantly changed with different tumor stages or grades. Survival analysis revealed that AC005332.5, ELF3-AS1 and LINC00665 were highly associated with the prognosis of overall survival. Additionally, the lncRNA signature yielded statistical significance to predict clinical outcomes independently from other clinical variables in validation patients, as suggested in the multivariate Cox hazards analysis. Conclusively, a three-lncRNA signature of AC005332.5, ELF3-AS1 and LINC00665 may serve as an excellent diagnostic biomarker for HBV-related HCC and potential prognostic significance for HBV-related HCC sufferers.

Expression and clinical significance of the m6A reader YTHDF2 in peripheral blood mononuclear cells from rheumatoid arthritis patients.

As an important m6A reader, the YT521-B homology domain family 2 (YTHDF2) has been shown to regulate mRNA degradation and translation, and to be involved in inflammation. However, little is known about the role of YTHDF2 in the autoimmune-based inflammatory disease rheumatoid arthritis (RA). To begin to ascertain any role for this reader, 74 RA patients and 63 healthy controls (HC) were recruited for this study. Blood was collected from each subject and peripheral blood mononuclear cells (PBMC) isolated. Thereafter, mRNA expression of YTHDF2, interleukin (IL)-1ß, IL-6, IL-8, and tumor necrosis factor (TNF)-α in the cells was determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The harvested blood was also assessed for a variety of parameters, including levels of C-reactive protein (CRP), erythrocyte sedimentation rates (ESR), white blood cell counts (WBC), neutrophils counts (N)/neutrophils percentages (N%), and neutrophil:lymphocyte ratios (NLR) - each markers of inflammation during RA. The results showed that YTHDF2 mRNA expression in RA patient PBMC was decreased significantly vs that in healthy control subject cells. Further, YTHDF2 mRNA expression in RA patient PBMC negatively-correlated with ESR, CRP levels, WBC counts, as well as neutrophils counts, percentages, and NLR values. In addition, it was seen that YTHDF2 mRNA expression in RA patient PBMC was associated with host serum RF levels and treatment. Moreover, it was found that mRNA expression of IL-1ß, IL-6, IL-8, and TNFα was increased in PBMC from RA patients relative to in control subject cells; however, only the increased IL-1ß expression was seen to be negatively-correlated with decreased YTHDF2 mRNA expression. In conclusion, the present study illustrated that YTHDF2 expression might have some regulatory role in the underlying mechanisms associated with the autoimmune disease RA and that this m6A reader could at some point represent a potential target for regulating inflammatory responses that occur during RA.

[LRG47/EBP50 recombinant lentivirus-targeted vector vaccine enhances anti-tuberculosis immunity of RAW264.7 macrophages and its mechanism].

Objective To investigate the effect of the gene vaccine in anti-tuberculosis immunity by constructing immunity-related p47 GTPase/ezrin-radixin-moesin-binding phosphoprotein 50 (LRG47/EBP50) gene co-expression recombinant lentivirus targeting vector. Methods Recombinant lentiviral plasmid vector pLenti-EBP50-LRG47 was established by using molecular cloning and packaged as lentivirus LV-EBP50-LRG47 and H37Rv infect macrophages. Then their bactericidal ability was tested by colony-forming units while the cellular autophagy and apoptosis was detected by flow cytometry. iNOS protein was detected by Western blotting and the expression level of nitric oxide (NO) was detected by ultraviolet spectrophotometer. Results The recombinant lentivirus LV-EBP50-LRG47 successfully up-regulated the expression of EBP50 and LRG47 after infecting macrophages. Compared with the control group, LV-EBP50-LRG47 can significantly inhibit the growth of intracellular H37Rv. The autophagy and apoptosis levels of LV-EBP50-LRG47 infected macrophages increased significantly, and the expression levels of iNOS and NO were significantly up-regulated. Conclusion LRG47/EBP50 gene co-expression enhances macrophages autophagy and apoptosis, and increases generation of iNOS and NO, which significantly inhibites the growth of intracellular H37Rv.

Expression and Clinical Significance of the m6A RNA-Binding Proteins YTHDF2 in Peripheral Blood Mononuclear Cells From New-Onset Ankylosing Spondylitis.

This study has focused on determining the association of m6A methyltransferase [methyltransferase-like 3 (METTL3), methyltransferase-like 14 (METTL14), and Wilms tumor 1-associating protein (WTAP)], demethylase [fat mass and obesity-associated protein (FTO) and alkylation repair homolog protein 5 (ALKBH5)], RNA-binding proteins [YT521-B homology domains 2 (YTHDF2)], and ankylosing spondylitis (AS). A total of 154 specimens, containing 79 patients with new-onset AS and 75 healthy controls (HCs), participated in the study. The mRNA expressions of these m6A methyltransferase, demethylase, and RNA-binding protein in peripheral blood mononuclear cells (PBMCs) were detected by quantitative real-time PCR (qRT-PCR). The data showed that the mRNA expressions of YTHDF2 and ALKBH5 in PBMC from patients with new-onset AS were significantly decreased, and there was a positive correlation between RNA-binding proteins (YTHDF2) and demethylase (ALKBH5) in patients with new-onset AS. Logistic regression analysis demonstrated that the expression of YTHDF2 mRNA in PBMC is a risk factor of AS. Receiver operating characteristic (ROC) analysis of the area under the curve (AUC) for mRNA YTHDF2 in new-onset AS and HC was 0.692, with a cutoff value of <0.8724, a sensitivity of 67%, and a specificity of 63%. Moreover, we constructed a novel predictive model based on a combination of mRNA YTHDF2 and systemic immune-inflammation index (SII) for AS diagnosis (AUC = 0.865, sensitivity = 79.45%, specificity = 84.00%), and the predictive model correlated with the activity and severity of AS. This study indicates that the mRNA expression of YTHDF2 in PBMC may be involved in AS pathogenesis and a predictive model based on a combination of mRNA YTHDF2 and SII acts as a marker for diagnosis and progression of diseases.

PPARγ Ameliorates Mycobacterium tuberculosis H37Ra-Induced Foamy Macrophage Formation via the ABCG1-Dependent Cholesterol Efflux Pathway in THP-1 Macrophages.

Foamy macrophages are present during the course of Mycobacterium tuberculosis (Mtb) infection and seems to be nutrient-rich reservoir and secure reservoir for the bacilli, which leads to bacterial persistence and infection transmission. Peroxisome proliferator activated receptor γ (PPARγ) is a key transcription factor for cholesterol metabolism in macrophages and its role in regulating atherosclerosis related foamy macrophages (FMs) formation has been well-studied. However, knowledge about the mechanism of PPARγ regulating Mtb infection induced FM formation remains very limited. In this study, we investigate the functional role of PPARγ in Mtb H37Ra infection-induced foamy macrophages formation. H37Ra infection induced a time-dependent decreased expression of PPARγ that paralleled the augmented lipid body formation in THP1-derived macrophages. PPARγ antagonist GW9662 significantly potentiate H37Ra induced lipid body formation and inhibit ABCG1 expression, overexpression of ABCG1 by transduced macrophages with lentivirus significantly reversed the promotion effect of GW9662 on FM formation. Moreover, Treatment with a TLR2 neutralizing antibody ameliorated the activation of ABCG1 by Mtb H37Ra without significantly effecting the suppression of PPARγ, suggesting a greater role for TLR2 to regulate ABCG1 compared to PPARγ. Overall, this study showed that PPARγ is involved in ameliorating FM formation by regulating ABCG1 expression, these observations expose a novel role of PPARγ in the Mtb infection induced FM formation.

Increased TIM-3+PD-1+ NK cells are associated with the disease activity and severity of systemic lupus erythematosus.

It is well established that natural killer (NK) cells are dysregulated in systemic lupus erythematosus (SLE) patients. However, the functions of NK cells and the mechanisms regulated by them in SLE remain incompletely understood. Patients with SLE were recruited from The First Affiliated Hospital of Nanchang University, and their clinical characteristics and treatments were recorded. The expression levels of T cell immunoglobulin mucin-3 (TIM-3) and programmed cell death protein 1 (PD-1) on NK cells were examined using flow cytometry. The correlations between the increase in TIM-3+PD-1+ NK cells in the SLE patients and clinical traits, including inflammatory markers, auto-antibodies, disease activity and severity of SLE, were examined. The TIM-3+NK cells, PD-1+NK cells and TIM-3+PD-1+ NK cells were significantly increased in the SLE patients. The increase in TIM-3+PD-1+ NK cells in the patients with SLE was associated with erythrocyte sedimentation rate, C-reactive protein, anti-double stranded DNA, anti-ribosomal P, SLE disease activity index and clinical features. The frequency of TIM-3+PD-1+NK cells in SLE patients with a cardiovascular disease (CVD) was significantly lower than that in SLE patients without a CVD. Moreover, the increased TIM-3+PD-1+ NK cells were significantly decreased in SLE patients following treatment. The present study suggested that the increased TIM-3+PD-1+ NK cells were associated with the disease activity and severity of SLE and may play a role in SLE pathogenesis.

Deregulation of circ_003912 contributes to pathogenesis of erosive oral lichen planus by via sponging microRNA-123, -647 and -31 and upregulating FOXP3.

BACKGROUND: The FOXP3/miR-146a/NF-κB axis was previously reported to modulate the induction and function of CD4+ Treg cells to alleviate oral lichen planus. Also, other signaling pathways including microRNA-155-IFN-γ loop and FOXP3/miR-146a/TRAF6 pathways were reported to be involved in the pathogenesis of oral lichen planus. In this study, we aimed to investigate the molecular mechanism underlying the pathogenesis of EOLP. METHOD: CircRNA microarray was used to observe the expression of candidate circRNAs in CD4+ T-cells collected from different groups. Real-time PCR and Western blot were conducted to observe the changes in the expression of different miRNAs, mRNAs and proteins. Flow cytometry was performed to compare the counts of Treg cells in the HC and EOLP groups, and ELISA was performed to evaluate the changes in the expression of inflammatory cytokines. RESULT: No obvious differences were seen between the HC and EOLP groups in terms of age and gender. Among all candidate circRNAs, the expression of circ_003912 was most dramatically elevated in CD4+ T-cells collected from the EOLP group. The levels of miR-1231, miR-31, miR-647, FOXP3 mRNA and miR-146a were decreased while the expression of TRAF6 mRNA was increased in CD4+ T-cells collected from the EOLP group. The count of Treg cells in the EOLP group was dramatically increased. The levels of inflammatory cytokines including IL-4 IFN-γ, IL-10 and IL-2 were influenced by the presence of circ_003912. In CD4+ T-cells in the EOLP group, the levels of IL-4 and IL-10 were decreased while the levels of IFN-γ and IL-2 were increased. The presence of miR-1231, miR-31 and miR-647 all obviously inhibited the expression of circ_003912, which was validated to sponge the expression of above miRNAs. Also, FOXP3 mRNA was proved to be targeted by miR-1231, miR-31 and miR-647. Transfection of circ_003912 up-regulated the expression of circ_003912, miR-146a and FOXP3 mRNA/protein while down-regulating the expression of miR-1231, miR-31, miR-647, and TRAF6 mRNA/protein. The levels of inflammatory cytokines including IL-4 IFN-γ, IL-10 and IL-2 as well as the speed of cell proliferation were influenced by circ_003912. CONCLUSION: In this study, we investigated the molecular mechanisms underlying the pathogenesis of EOLP which involved the functioning of circ_003912. We first demonstrated that circ_003912 was up-regulated in CD4+ T-cells of the EOLP group. And miRNAs including miR-1231, miR-31 and miR-647 were sponged by circ_003912 and down-regulated in CD4+ T cells of the EOLP group, which subsequently up-regulated the expression of FOXP3 and miR-146a, and resulted in the inhibition of NF-kB.

Circular RNAs hsa-circ0000175 and hsa-circ0044235 in plasma are novel biomarkers for new-onset rheumatoid arthritis.

Circular RNAs (circRNAs) are a class of non-coding RNAs that could serve as potential molecular markers for disease diagnosis. However, the role of circRNAs in plasma from new-onset rheumatoid arthritis (RA) has not been extensively investigated. In this study, the expression of hsa-circ0000175 and hsa-circ0044235 in plasma from RA patients, healthy controls (HCs), systemic lupus erythematosus (SLE) patients, osteoarthritis (OA), and undiagnosed arthritis (UA) patients were determined by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Correlation analysis was used to assess the correlation of the two circRNAs and clinical variables of RA. The receiver operating characteristic (ROC) curves were created to evaluate the diagnostic value and multivariate analysis (logistic regression) was performed to analyse the risk factors. We confirmed that hsa-circ0000175 was significantly elevated in plasma from patients with new-onset RA compared with HC and patients with new-onset SLE, but significantly was reduced when compared with OA + UA patients. Hsa-circ0044235 was found to be significantly decreased in plasma from patients with new-onset RA compared with HC and OA + UA patients, but was significantly increased compared with SLE patients. The expression of plasma hsa-circ0000175 in new-onset RA patients was associated with platelet count (PLT), plateletcrit (PCT), and platelet large cell ratio (PLR), the expression of plasma hsa-circ0044235 new-onset RA patients was associated with swollen joint count (SJC), painful joint count (PJC), and disease activity score 28 (DAS28). ROC curve analysis suggested that the combination of hsa-circ0000175 and hsa-circ0044235 has some value in the diagnosis of new-onset RA from HC, patients with SLE and patients with OA + UA. The logistic regression analysis revealed that the expression of hsa-circ0000175 and hsa-circ0044235 in plasma were risk factors for RA. This study suggests that the combination of plasma hsa-circ0000175 and hsa-circ0044235 improves the diagnostic accuracy for new-onset RA. Moreover, the expression levels of plasma hsa-circ0000175 and hsa-circ0044235 were associated with disease activity and severity of RA.