Differential diagnostic value of simultaneous detection of CD69 and HLA-DR on host T and NK cells in QFT-TB assay for identifying active tuberculosis.

BACKGROUND: Interferon-gamma release assay (IGRA) for tuberculosis (TB) remains limited in its ability to discriminate between active TB (ATB) and latent TB infection (LTBI). Activation markers on host T and NK cells are currently considered to be promising markers in the diagnosis of ATB. METHODS: This prospective observational study enrolled 213 participants and the participants were divided into ATB, LTBI, other lung-related diseases (ORD), and health control (HC) groups. CD69 and HLA-DR on T and NK cells were detected in QFT-TB assay, and a composite scoring system (TB-Flow) was created for the diagnosis of ATB. RESULTS: The expression of activation markers (CD69 and HLA-DR) were significantly increased in ATB. HLA-DR on NK cells, CD69 on T cells, and QFT-TB in the differential diagnosis of ATB and HC were all of good diagnostic value (AUC>0.90). In addition, the TB-Flow greatly improved the efficiency of differential diagnosis between ATB and LTBI (AUC=0.90, 95%CI: 0.84-0.96), with sensitivity and specificity of 79.17 % (95%CI: 64.60%-89.04 %) and 88.68 % (95%CI: 76.28%-95.31 %). CONCLUSIONS: CD69 and HLA-DR on host T and NK cells are promising markers in distinguishing different TB infection status. Our blood-based TB-Flow scoring system can distinguish ATB from LTBI with good diagnostic efficacy.

Expression of HLA-DR and KLRG1 enhances the cytotoxic potential and cytokine secretion capacity of CD3+ T cells in tuberculosis patients.

BACKGROUND: Human T cells play an important role in immunity against tuberculosis (TB) infection. Activating receptor HLA-DR and inhibitory receptor KLRG1 are critical regulators of T cell function during viral infection and tumorigenesis, but they have been less studied in TB infection. METHODS: In this study, we explored the relationship between CD3+ T cell expression of HLA-DR and KLRG1 receptors and function against TB infection. Flow cytometry was conducted to assess the immunomodulatory effects of HLA-DR and KLRG1 receptors on CD3+ T cells in patients with different TB infection status. RESULTS: We found activating receptors HLA-DR, NKG2C, CD57 and NKP46, and inhibitory receptors KLRG1 and KIR on CD3+ T cells in different TB infection status showed different distribution patterns; the cytotoxic potential and cytokine secretion capacity of CD3+ T cells after Mtb-specific antigen stimulation were significantly enhanced in TB infection groups. Further studies revealed HLA-DR+ T and KLRG1+ T cells expressed higher activating and inhibitory receptors than the negative population. In addition, the expression of cytotoxic potential and cytokine secretion capacity of HLA-DR+ T and KLRG1+ T cells was significantly higher than that of HLA-DR- T and KLRG1- T cells. CONCLUSIONS: Expression of HLA-DR and KLRG1 enhances the cytotoxic potential and cytokine secretion capacity of CD3+ T cells in TB patients, suggesting CD3+ T cells expressing HLA-DR and KLRG1 are important effector cell phenotypes involved in the host anti-TB infection. HLA-DR and KLRG1 expressed by CD3+ T cells may be potential predictive markers of TB disease progression and clinical immune assessment.

Diagnostic Value of CD25, CD69, and CD134 on Tuberculosis-Specific Antigen-Stimulated CD4+ T Cells for Tuberculous Pleurisy.

Rapid and accurate methods for the diagnosis of tuberculous pleurisy (TP) are urgently needed. Activation markers of tuberculosis (TB)-reactive T cells are considered promising for the diagnosis of active TB (ATB). Different activation indexes may play different roles in the progression of TB, but there are few reports on T cell activation indicators, except for HLA-DR. Hence, we evaluated the expression of early (CD25 and CD69) and late (CD134) activation markers on TB antigen-stimulated CD4+ T cells in populations with different TB infection status and investigated their diagnostic value for ATB, particularly, for TP. Moreover, we compared the differences in the diagnostic efficacy among the indexes from peripheral blood (PB) and pleural fluid (PF) for TP. The expression of each activation marker was significantly increased in TB-infected populations (patients with ATB and latent TB infection vs. healthy individuals; patients with TP vs. non-TP) and was significantly higher in the PF than in the PB of patients with TP. The diagnostic performance of the coexpressed activation markers was superior to that of single expression markers in the differential diagnosis of ATB and non-TB, with CD25+CD134+ showing the best diagnostic efficiency (AUC: 0.93, 95% CI, 0.87-0.99; sensitivity: 86.7%, 95% CI, 72.5%-94.5%; and specificity: 94.0%, 95% CI, 82.5%-98.4%). Except for TB-IGRA, the activation indexes were more accurate than conventional laboratory methods for ATB diagnosis. In addition, the expression of CD25+CD134+ in PB and PF was the best values for differential diagnosis of TP and NTP, with AUCs of 0.87 (95% CI, 0.77-0.96) and 0.95 (95% CI, 0.90-1.00), respectively. Our study provides information on the diagnostic value of different activation markers for TB and shows that the expression of CD25+CD134+ on CD4+ T cells in PF can serve as a potential marker for TP diagnosis.

Host biomarkers other than interferon gamma in QFT-TB supernatants for identifying active tuberculosis.

Interferon gamma release assays (IGRAs) for tuberculosis (TB) remain limited in their ability to discriminate between active TB (ATB) and latent TB infection (LTBI). The objective of our study was to evaluate the value of additional cytokines/chemokines other than interferon gamma (IFN-γ) as biomarkers to identify different TB infection status. A total of 128 subjects were enrolled to detect the quantification of IL-2, IP-10, MCP-1 and RANTES in the supernatants of QuantiFERON®-TB (QFT-TB). Area under the curve (AUC) was used to evaluate the diagnostic efficiency. Notably, Mycobacterium tuberculosis (Mtb) induced cytokines/chemokines of ATB patients were significantly higher than those of the LTBI, other lung related diseases (ORD) and healthy controls (HC). Moreover, ROC analysis indicated that all cytokine/chemokine parameters detected were more capable of distinguishing ATB from LTBI than IFN-γ, especially IL-2. The diagnostic model including TB specific IL-2 and RANTES improved the performance in distinguishing ATB from LTBI, which was superior to single cytokines/chemokines in QFT-TB supernatants. Our results suggest that the combination of Mtb specific cytokines/chemokines has great prospects in the diagnosis of ATB, and the diagnostic model based on IL-2 and RANTES can be used as an alternative to distinguish ATB from LTBI.

A Small-Molecule Inhibitor of the Anthranilyl-CoA Synthetase PqsA for the Treatment of Multidrug-Resistant Pseudomonas aeruginosa.

One of the challenges associated with the treatment of Pseudomonas aeruginosa infections is the high prevalence of multidrug resistance (MDR). Since conventional antibiotics are ineffective at treating such bacterial infections, innovative antibiotics acting upon novel targets or via mechanisms are urgently required. In this study, we identified a quorum sensing inhibitor (QSI), norharmane, that uniquely shows weak antibacterial activity but strongly inhibits pyocyanin production and biofilm formation of MDR P. aeruginosa. Biophysical experiments and molecular docking studies showed that norharmane competes with anthraniloyl-AMP for anthranilyl-CoA synthetase PqsA of P. aeruginosa at the ligand-binding pocket, which is not exploited by current inhibitors, thereby altering transcription regulatory activity. Moreover, norharmane exhibits synergy with polymyxin B. This synergism exhibits a high killing rate, low probability of resistance selection, and minimal cytotoxicity. Notably, norharmane can effectively boost polymyxin B activity against MDR P. aeruginosa-associated infections in animal models. Together, our findings provide novel insight critical to the design of improved PqsA inhibitors, and an effective combination strategy to overcome multiantibiotic bacterial resistance using conventional antibiotics and QSIs. IMPORTANCE Pseudomonas aeruginosa is a dominant hospital-acquired bacterial pathogen typically found in immunocompromised individuals. It is particularly dangerous for patients with chronic lung diseases and was identified as a serious threat for patients in the 2019 Antimicrobial Resistance Threats report (https://www.cdc.gov/drugresistance/biggest-threats.html). In this study, we used activity-based high-throughput screening to identify norharmane, a potent and selective inhibitor of P. aeruginosa PqsA, which is a well-conserved master quorum sensing (QS) regulator in multidrug resistant (MDR) P. aeruginosa. This compound competitively binds anthranilyl-CoA synthetase PqsA at the anthraniloyl-AMP binding domain, which has not been exploited by known inhibitors. Remarkably, norharmane can significantly block the production of the virulence factor, pyocyanin (87%), and biofilm formation (80%) in MDR P. aeruginosa. Furthermore, norharmane is capable of augmenting polymyxin B activity against MDR P. aeruginosa in cell cultures and animal models. Taken together, these results suggest that norharmane may be an effective adjuvant for combating multiantibiotic bacterial resistance.

Evaluating the diversity of circulating natural killer cells between active tuberculosis and latent tuberculosis infection.

Tuberculosis (TB) is a leading global public health problem; however, the mechanisms underlying the immunopathology of TB progression are not well understood. It is currently believed that Mycobacterium tuberculosis (Mtb) infection can modify natural killer (NK) cell phenotypic signatures. Hence, our study was designed to investigate the diversity of circulating NK cells in patients with different TB infection status. NK subsets, as well as their expression of activating and inhibitory receptors between active TB (ATB) and latent TB infection (LTBI) were evaluated. There were significant differences in NK cell phenotypes between ATB, LTBI and healthy controls. Notably, the proportion of KLRG1 in NK cells (P = 0.036), as well as in their subsets CD56DimCD16+ (P = 0.046) and CD27+ (P = 0.027) NK cells, increased significantly in LTBI group than in ATB group; while Mtb specific IFN-γ+CD56BrightCD16Dim NK cells expressed higher KLRG1 in ATB than in LTBI (P = 0.027). However, the expression of activating receptor NKG2D in NK subsets showed no significant difference among the study groups. Our results suggest that different TB infection status are coupled with the diversity of NK cell compartments, and the expression of KLRG1 in NK cells may be a specific phenotype that modulates the progression of TB from latent to active.

Study on the value of small dense low-density lipoprotein in predicting cardiovascular and cerebrovascular events in the high-risk stroke population.

BACKGROUND: Lipid management in people at high risk of stroke is an important measurement to prevent the occurrence of stroke. The study aims to investigate the association between sdLDL and cardiovascular and cerebrovascular events in high-risk stroke populations. METHODS: This was a prospective study. Screened from 15,933 individuals aged >40 years in April 2013 and followed up at 3rd, 6th, 12th, and 24th months, 823 participants met the screening criteria and were investigated for clinical data and biochemical parameters. RESULTS: A total of 286 subjects had varying degrees of carotid stenosis, and 18 subjects experienced cardiovascular and cerebrovascular events during the two-year follow-up period. There was no positive correlation between sdLDL and carotid stenosis. Carotid stenosis and extent of carotid stenosis involvement did not predict cardiovascular and cerebrovascular events in patients with high-risk stroke, while sdLDL did. The sdLDL level in the events group was significantly higher than those in the no event group (p = 0.002). In the events group, the risk of events in the fourth quartile of sdLDL was 10.136 times higher than in the first quartile (HR = 10.136, 95% CI: 1.298-79.180, p = 0.027). CONCLUSIONS: sdLDL was positively correlated with the incidence of cardiovascular and cerebrovascular events, which can predict the occurrence of an event and provide a scientific basis for early prevention.

Serum Bile Acid Profiles in Latent Autoimmune Diabetes in Adults and Type 2 Diabetes Patients.

BACKGROUND: Impaired bile acid (BA) metabolism has been associated with the progression of type 2 diabetes (T2D). However, the contribution of BAs to the pathogenesis of latent autoimmune diabetes in adults (LADA) remains unclear. This study was aimed at investigating the association of serum BAs with different diabetes types and analyzing its correlation with main clinical and laboratory parameters. METHODS: Patients with LADA, patients with T2D, and healthy controls (HCs) were enrolled. Serum BA profiles and inflammatory cytokines were measured. The correlation of BA species with different indicators was assessed by Spearman's correlation method. RESULTS: Patients with diabetes (LADA and T2D) had significantly higher serum BAs, especially conjugated BAs, compared with those in HCs. Nevertheless, serum BA profiles had no special role in the progression of LADA, because no significant differences in BAs were observed between LADA and T2D patients. Interestingly, HbA1c levels and HOMA-ß were found to be correlated with a series of BA species. Proinflammatory cytokines (IL-1ß, IL-6, and TNF-α) and anti-inflammatory cytokine (IL-10) were all positively associated with several BA species, especially the conjugated secondary BAs. CONCLUSION: Serum BAs regulate glucose homeostasis, but have no special value in the pathogenesis of LADA patients. Our study adds further information about the potential value of serum BAs in different types of diabetes.

[Feasibility of Preheating at 41 ℃ to Correct Red Blood Cell Parameters in the Presence of High-titer Cold Agglutinins].

Objective To explore the feasibility of preheating in 41 ℃ water bath for 30 minutes to correct the red blood cell parameters in the specimens containing high-titer cold agglutinins(CAs). Methods Two specimens containing high-titer CAs were selected during work,and the parameters of complete blood count at room temperature or after preheating in 37 ℃ or 41 ℃ water bath were compared.The smears were stained,and the distribution of red blood cells was observed with a microscope.Further,74 specimens without CAs were collected for complete blood count,and then the test results at room temperature and after preheating at 41 ℃ were compared. Results At room temperature,the specimens containing high-titer CAs showed significantly reduced red blood cell count(RBC)and hematocrit(HCT),abnormally increased mean corpuscular hemoglobin(MCH)and mean cell hemoglobin concentration(MCHC),abnormal percents of hemoglobin(HGB)and RBC,and aggregation of a large number of red blood cells.After being preheated at 37 ℃ for a certain time,the specimens demonstrated obviously improved parameters while still aggregation of a small number of red blood cells.After being preheated at 41 ℃ for 30 minutes,the specimens showed significantly increased RBC,normal HCT,MCH,and MCHC,and evenly distributed red blood cells.The 74 specimens without CAs showed the comparability was ≥80% between room temperature and preheating at 41 ℃ for 30 minutes or 60 minutes. Conclusion We can preheat the specimens containing high-titer CAs in a water bath at 41 ℃ to obtain accurate red blood cell parameters.

Establishment of the reference intervals of whole blood neutrophil phagocytosis by flow cytometry.

OBJECTIVE: To investigate the reference intervals (RIs) of the whole blood neutrophil phagocytosis by flow cytometry (FCM) and to study the application value of neutrophil phagocytosis in infectious diseases. METHODS: Pathogens (Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923) cultured for 18-24 h were labeled by fluorescence probe carboxyfluorescein diacetate succinimidyl ester (CFDA-SE), and then incubated with whole blood at 37℃. The phagocytosis of pathogens by neutrophils was detected by flow cytometry, and a reference interval was established. RESULTS: In the healthy adults, the reference interval for the neutrophil phagocytosis to Escherichia coli was 46.91%-83.09% and to Staphylococcus aureus was 33.92%-69.48%. This method showed good reproducibility. Neutrophil phagocytosis was negatively correlated with the neutrophil count, neutrophil percentage, and neutrophil-to-lymphocyte ratio (NLR, p < 0.05). CONCLUSION: We have successfully established the RIs of neutrophil phagocytosis in whole blood in healthy adults by flow cytometry (FCM), which might be of important clinical value in the diagnosis, treatment, and prognosis of infectious diseases.

DHA exhibits synergistic therapeutic efficacy with cisplatin to induce ferroptosis in pancreatic ductal adenocarcinoma via modulation of iron metabolism.

Pancreatic ductal adenocarcinoma (PDAC) is an extremely lethal cancer with limited treatment options. Cisplatin (DDP) is used as a mainstay of chemotherapeutic agents in combination with other drugs or radiotherapy for PDAC therapy. However, DDP exhibits severe side-effects that can lead to discontinuation of therapy, and the acquired drug resistance of tumor cells presents serious clinical obstacles. Therefore, it is imperative to develop a more effective and less toxic therapeutic strategy. We and others have previously discovered that dihydroartemisinin (DHA) represents a safe and promising therapeutic agent to preferentially induce cancer cell ferroptosis. In the present study, we find that DHA could intensively strengthen the cytotoxicity of DDP and significantly reduce its effective concentrations both in vitro and in vivo. Combination of DHA and DDP synergistically inhibits the proliferation and induces DNA damage of PDAC cells. Mechanically, the combinative treatment impairs mitochondrial homeostasis, characterized by destroyed mitochondrial morphology, decreased respiratory capacity, reduced ATP production, and accumulated mitochondria-derived ROS. Further studies show that ferroptosis contributes to the cytotoxic effects in PDAC cells under the challenge of DHA and DDP, together with catastrophic accumulation of free iron and unrestricted lipid peroxidation. Moreover, pharmacologic depleting of the free iron reservoir or reconstituted expression of FTH contributes to the tolerance of DHA/DDP-induced ferroptosis, while iron addition accelerates the ferroptotic cell death. In summary, these results provide experimental evidence that DHA acts synergistically with DDP and renders PDAC cells vulnerable to ferroptosis, which may act as a promising therapeutic strategy.

Multi-function PtCo nanozymes/CdS nanocrystals@graphene oxide luminophores and K2S2O8/H2O2 coreactants-based dual amplified electrochemiluminescence immunosensor for ultrasensitive detection of anti-myeloperoxidase antibody.

BACKGROUND: Anti-myeloperoxidase antibody (anti-MPO) is an important biomarker for anti-neutrophil cytoplasm antibody (ANCA)-associated vasculitides (AAVs). However, the complicated operation procedures and insufficient sensitivity of conventional anti-MPO detection methods limit their application in monitoring efficacy of AAVs in clinical diagnosis. Herein, a dual amplified electrochemiluminescence (ECL) immunosensor based on multi-function PtCo nanozymes/CdS nanocrystals@graphene oxide (PtCo/CdS@GO) luminophores and K2S2O8/H2O2 coreactants has been fabricated for ultrasensitive detection of anti-MPO. RESULTS: PtCo/CdS@GO luminophores as novel signal amplification labels and nanocarriers to load rabbit anti-mouse IgG were synthesized by co-doping with Pt and Co nanozymes simultaneously with several considerable advantages, including astonishing peroxidase-like catalytic activity, high-efficiency luminescence performance and superior stability in aqueous solutions. Meanwhile, upon the K2S2O8/H2O2 coreactants system, benefiting from the efficient peroxidase-like activity of the PtCo/CdS@GO toward H2O2, massive of transient reactive intermediates could react with K2S2O8, thus obtaining higher ECL emission. Therefore, the developed ECL immunosensor for anti-MPO detection displayed good analytical performance with good concentration linearity in the range of 0.02 to 1000 pg/mL and low detection limit down to 7.39 fg/mL. CONCLUSIONS: The introduction of multi-function PtCo/CdS@GO luminophores into the established ECL immunoassay not only was successfully applied for specific detection of anti-MPO in clinical serum samples, but also provided a completely new concept to design other high-performance luminophores. Meaningfully, the ECL immunoassay strategy held wide potential for biomarkers detection in clinical diagnosis.

Establishment of a regression model of bone metabolism markers for the diagnosis of bone metastases in lung cancer.

BACKGROUND: The aim of this study was to establish a regression equation model of serum bone metabolism markers. We analyzed the diagnostic value of bone metastases in lung cancer and provided laboratory evidence for the early clinical treatment of bone metastases in lung cancer. METHODS: A total of 339 patients with non-metastatic lung cancer, patients with lung cancer with bone metastasis, and patients with benign lung disease who were treated in our hospital from July 2012 to October 2015 were included. A total of 103 patients with lung cancer in the non-metastatic group, 128 patients with lung cancer combined with bone metastasis group, and 108 patients with benign lung diseases who had nontumor and nonbone metabolism-related diseases were selected as the control group. Detection and analysis of type I collagen carboxyl terminal peptide ß-special sequence (ß-CTX), total type I procollagen amino terminal propeptide (TPINP), N-terminal-mid fragment of osteocalcin (N-MID), parathyroid hormone (PTH), vitamin D (VitD3), alkaline phosphatase (ALP), calcium (CA), phosphorus (P), cytokeratin 19 fragment (F211), and other indicators were performed. Four multiple regression models were established to determine the best diagnostic model for lung cancer with bone metastasis. RESULTS: Analysis of single indicators of bone metabolism markers in lung cancer was performed, among which F211, ß-CTX, TPINP, and ALP were significantly different (P < 0.05). The ROC curve of each indicator was less than 0.712. Based on the multiple regression models, the fourth model was the best and was much better than a single indicator with an AUC of 0.856, a sensitivity of 70.0%, a specificity of 91.0%, a positive predictive value of 82.5%, and a negative predictive value of 72.0%. CONCLUSION: Multiple regression models of bone metabolism markers were established. These models can be used to evaluate the progression of lung cancer and provide a basis for the early treatment of bone metastases.

Lp-PLA2 evaluates the severity of carotid artery stenosis and predicts the occurrence of cerebrovascular events in high stroke-risk populations.

BACKGROUND: Lipoprotein-associated phospholipase A2 (Lp-PLA2) is an independent risk factor for cardiovascular disease. However, relationship between carotid artery stenosis and cerebrovascular events in high stroke-risk populations is still unclear. METHODS: A total of 835 people at a high risk of stroke were screened from 15,933 people aged >40 years in April 2013 and followed at 3, 6, 12, and 24 months. Finally, 823 participants met the screening criteria, and the clinical data and biochemical parameters were investigated. RESULTS: Among the 823 participants, 286 had varying degrees of carotid artery stenosis and 18 had cerebrovascular events. The level of Lp-PLA2 in the carotid artery stenosis group was higher than that in the no stenosis group, and the level in the event group was higher than that in the no event group (p < 0.05). Spearman correlation analysis showed that Lp-PLA2 was positively correlated with the degree of carotid artery stenosis (r = 0.093, p = 0.07) and stenosis involvement (r = 0.094, p = 0.07). The correlation coefficient between Lp-PLA2 and lipoprotein was the highest on the levels of sdLDL (r = 0.555, p < 0.001), followed by non-HDL, LDL, TC, and TG. Cox multivariate regression analysis revealed that, compared with the first quantile of Lp-PLA2 level (Q1, low level), the risk of cerebrovascular events in the fourth quantile of Lp-PLA2 was 10.170 times that of the first quantile (OR = 10.170, 95% CI 1.302-79.448, p = 0.027). CONCLUSIONS: Lp-PLA2 levels can evaluate carotid artery stenosis and predict the occurrence of cerebrovascular events in high stroke-risk populations and provide scientific guidance for risk stratification management.

Penicillium janthinellum Pneumonia in an SLE Patient: A Case Study.

The risk of opportunistic fungal infections is high in immunocompromised patients. The Penicillium genus is common and diverse in nature. However, it rarely causes infection in humans. Here, we reported a case of Penicillium janthinellum pneumonia in a systemic lupus erythematosus (SLE) patient, and the morphological characteristics of P. janthinellum were also described. The patient was a 64-year-old female. She had been diagnosed with SLE and membranous lupus nephritis 10 months previously. Her medications included methylprednisolone, cyclosporine, and hydroxychloroquine. She was admitted because of fever and diagnosed with pneumonia. P. janthinellum was isolated from sputum and bronchoalveolar lavage (BAL) samples. BAL fluid stained with multiple stains showed the presence of somewhat dichotomously branching septate fungal hyphae. P. janthinellum was identified, and its morphological features were described. Antibiotic susceptibility profiles showed that this strain had higher minimum inhibitory concentration (MIC) values in response to multiple antifungal drugs. The patient died 10 days after diagnosis. To the best of our knowledge, this report is the second to demonstrate that P. janthinellum causes infection and is the first to present an infection (pneumonia) caused by P. janthinellumi in an SLE patient. Clinical and laboratory personnel should be aware that the Penicillium genus also contains pathogenic bacteria that cannot simply be treated as contaminants, especially in immunosuppressed patients.

Decreased levels of regulatory B cells in patients with acute pancreatitis: association with the severity of the disease.

Early stratification of the severity of acute pancreatitis (AP) is clinically important. Regulatory B cells have been found to be associated with disease activity of autoimmune diseases. However, the role of Regulatory B cells in AP remains unknown. We investigate the dynamic longitudinal changes in circulating IL-10-producing B cells (B10) and memory CD19+CD24hiCD27hi cells in patients with AP to evaluate their prediction utility for AP severity. B10, CD19+CD24hiCD27hi cells, inflammatory markers and cytokines were detected in patients with AP immediately after admission to the hospital (day 1), then on the third and seventh days. We observed decreases in lymphocytes, CD19+, B10, CD19+CD24hiCD27hi cells and lower mean fluorescence intensity (MFI) of CD80 and CD86 on B10 or CD19+CD24hiCD27hi cells in patients with AP, especially in those with severe acute pancreatitis (SAP). CD19+CD24hiCD27hi cells from patients with AP suppressed the cytokine productions of CD4+ T cells and CD14+ monocytes, but had impaired ability to induce regulatory T cells response. B10 and CD19+CD24hiCD27hi cells significantly increased in patients with mild acute pancreatitis (MAP) from day 1 to day 7, whereas these indexes remained stable in patients with SAP. B10 or CD19+CD24hiCD27hi cells were negatively correlated with the severity index (APACHE II score), inflammatory markers (C-reactive protein, CD64 index), and cytokines (IL-6, IL-17, TNF-α). Furthermore, receiver operating characteristic (ROC) curve analysis revealed that B10 and CD19+CD24hiCD27hi cells could predict the development of SAP. Thus, the detection of B10 and CD19+CD24hiCD27hi cells may be a practical way to improve the early assessment of AP severity.

Functionally impaired follicular helper T cells induce regulatory B cells and CD14+ human leukocyte antigen-DR- cell differentiation in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) represents one of the most common and aggressive cancers worldwide, as it typically displays irreversible progression and poor prognosis. Interaction between programmed death 1 (PD-1) and its ligand, PD-L1, plays important roles in tumor immunology. Follicular helper T (Tfh) cells have characteristically high PD-1 expression; thus, in the present study, we investigated the role of circulating Tfh cells and their correlation with disease-free survival after tumor resection in NSCLC. We found significantly higher number of Tfh cells but lower serum interleukin (IL)-21 levels in NSCLC patients, especially in those with advanced stage (III and IV), indicating that the function of Tfh cells to produce IL-21 was impaired. Further analysis showed that the increase in Tfh cells was attributable to an expansion of the PD-1+ -Tfh2 and PD-1+ -Tfh17 subtypes. Functional analysis showed that Tfh cells from NSCLC patients induced the differentiation of regulatory B cells and CD14+ human leukocyte antigen (HLA)-DR- cells. Interestingly, the number of Tfh1 subtypes in NSCLC patients was negatively correlated with disease-free survival after tumor resection. In short, the high number and abnormal function of Tfh cells could cause further immunosuppression and lead to tumor development in NSCLC. Rescuing Tfh functions therefore represents a potential therapeutic strategy in NSCLC.

Increased circulating Tfh17 and PD-1+Tfh cells are associated with autoantibodies in Hashimoto's thyroiditis.

OBJECTIVE: Hashimoto's thyroiditis (HT) is characterized by autoantibodies targeting the thyroid. Abnormal CD4+CXCR5+T cell levels were previously shown to be associated with HT. However, Tfh cells consist of heterogeneous subpopulations, and which T follicular helper (Tfh) cell subpopulation participates in the pathogenesis of HT remains poorly understood. METHODS: Thirty healthy controls (HCs) and 52 HT patients were enrolled in the study. The percentages of Tfh, ICOS+Tfh, PD1+Tfh, Tfh1, Tfh2, Tfh17, effector Tfh, resting Tfh, effector memory Tfh, central memory Tfh, and naïve Tfh cells in the peripheral blood were all determined via flow cytometry, and the associations between the percentages of these cells and thyroid function indices were also investigated. RESULTS: The percentage of Tfh cells was significantly higher in HT patients than in HCs. Examination of the Tfh cell subsets revealed that the percentages of Tfh1, Tfh2, and resting Tfh cells were significantly decreased, while those of the ICOS+Tfh, PD1+Tfh, Tfh17, and effector Tfh cells were significantly increased in HT patients. No significant differences in effector memory, central memory or naïve Tfh cell percentages were noted between the HC and HT groups. Furthermore, the percentage of PD1+Tfh cells was positively correlated with anti-thyroglobulin antibody levels. Most importantly, only Tfh17 cell percentages were positively correlated with anti-thyroglobulin and anti-thyroid peroxidase antibody levels and were negatively correlated serum free T3 and free T4 levels in HT patients. CONCLUSIONS: Increased circulating Tfh17 cell and PD1+Tfh percentages are associated with higher autoantibody levels in HT patients, which imply that Tfh17 or PD1+Tfh cells may play a pathogenic role in the development of HT.

Dissemination of blaNDM-5 gene via an IncX3-type plasmid among non-clonal Escherichia coli in China.

Background: The emergence and spread of New Delhi metallo-ß-lactamase-producing Enterobacteriaceae has been a serious challenge to manage in the clinic due to its rapid dissemination of multi-drug resistance worldwide. As one main type of carbapenemases, New Delhi metallo-ß-lactamase (NDM)is able to confer resistance to almost all ß-lactams, including carbapenems, in Enterobacteriaceae. Recently, New Delhi metallo-ß-lactamase-5 attracted extensive attention because of increased resistance to carbapenems and widespread dissemination. However, the dissemination mechanism of blaNDM-5 gene remains unclear. Methods: A total of 224 carbapenem-resistant Enterobacteriaceae isolates (CRE) were collected from different hospitals in Zhejiang province. NDM-5-positive isolates were identified and subjected to genotyping, susceptibility testing, and clinical data analysis. We established the genetic location of blaNDM-5 with southern blot hybridisation, and analysed plasmids containing blaNDM-5 with filter mating and DNA sequencing. Results: Eleven New Delhi metallo-ß-lactamase-5 (NDM-5)-producing strains were identified, including 9 Escherichia coli strains, 1 Klebsiella pneumoniae strain, and 1 Citrobacter freundii strain. No epidemiological links for E. coli isolates were identified by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). S1-PFGE and southern blot suggested that the blaNDM-5 gene was located on a 46-kb IncX3-type plasmid in all isolates. Nine of the 11 isolates (81.8%) tested could successfully transfer their carbapenem-resistant phenotype to E. coli strain C600. Moreover, sequence analysis further showed that this plasmid possessed high sequence similarity to most of previously reported blaNDM-5-habouring plasmids in China. Conclusion: The present data in this study showed the IncX3 type plasmid played an important role in the dissemination of blaNDM-5 in Enterobacteriaceae. In addition, to the best of our knowledge, this report is the first to isolate both E. coli and C. freundii strains carrying blaNDM-5 from one single patient, which further indicated the possibility of blaNDM-5 transmission among diverse species. Close surveillance is urgently needed to monitor the further dissemination of NDM-5-producing isolates.