Expression pattern of the bone morphogenetic protein-4 and its relationship with inflammation, vascular injury in patients suffered the arterial occlusive diseases. / åŠ¨è„‰é˜»å¡žæ€§ç–¾ç— æ‚£è€ è¡€æµ†éª¨å½¢æ€å‘ç”Ÿè›‹ç™½-4çš„è¡¨è¾¾å˜åŒ–åŠå ¶ä¸Žç‚Žç—‡å’Œè¡€ç®¡æŸä¼¤çš„ç›¸å ³æ€§.

OBJECTIVES: Bone morphogenetic protein-4 (BMP4) has been proved to be an important regulatory factor for the pathological process of atherosclerosis (AS). However, there are few related clinical studies. This study aims to investigate the levels of plasma BMP4 in patients suffering from the arterial occlusive diseases (ACD) characterized by AS, and further to test the relationship between BMP4 and inflammation and vascular injury. METHODS: A total of 38 ACD patients (the ACD group) and 38 healthy people for the physical examination (the control group) were enrolled. The plasma in each subject from both groups was obtained to test the levels of BMP4, tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), IL-10, and vascular endothelial cadherin (VE-cadherin), and the relationship between BMP4 and the detected indicators above were further analyzed. RESULTS: Compared with the control group, the patients in the ACD group displayed significant elevations in the neutrophil to lymphocyte ratio [NLR, 1.63 (1.26, 1.91) vs 3.43 (2.16, 6.61)] and platelet to lymphocyte ratio [PLR, 6.37 (5.26, 7.74) vs 15.79 (7.97, 20.53)], while decrease in the lymphocyte to monocyte ratio [LMR, 5.67 (4.41, 7.14) vs 3.43 (2.07, 3.74)] (all P<0.05). Besides, the ACD patients displayed significant elevations in plasma BMP4 [581.26 (389.85, 735.64) pg/mL vs 653.97(510.95, 890.43) pg/mL], TNF-α [254.16 (182.96, 340.70) pg/mL vs 293.29(238.90, 383.44) pg/mL], and VE-cadherin [1.54 (1.08, 2.13) ng/mL vs 1.85 (1.30, 2.54) ng/mL], and decrease in IL-10 [175.89 (118.39, 219.25) pg/mL vs 135.92 (95.80, 178.04) pg/mL] (all P<0.05). While the levels of IL-1ß remained statistically comparable between the 2 groups (P=0.09). Furthermore, the plasma BMP4 levels were further revealed to be positively correlated with the levels of IL-1ß (r=0.35), TNF-α (r=0.31) and VE-cadherin (r=0.47), while they were negatively correlated with the levels of IL-10 (r=-0.37; all P<0.01). CONCLUSIONS: After ACD occurrence, the patients' plasma concentrations of BMP4 would be upregulated, which may serve as a candidate to indicate the levels of inflammation and vascular injury.

Natural killer cell subsets and their functional molecules in peripheral blood of the patients with breast cancer.

BACKGROUND: Natural killer (NK) cells, CD3- lymphocytes, are critical players in cancer immune surveillance. This study aimed to assess two types of CD3- NK cell classifications (subsets), that is, convectional subsets (based on CD56 and CD16 expression) and new subsets (based on CD56, CD27, and CD11b expression), and their functional molecules in the peripheral blood of patients with breast cancer (BC) in comparison with healthy donors (HDs). METHODS: Thirty untreated females with BC and 20 age-matched healthy women were enrolled. Peripheral blood samples were collected and directly incubated with fluorochrome-conjugated antibodies against CD3, CD56, CD16, CD27, CD11b, CD96, NKG2C, NKG2D, NKp44, CXCR3, perforin, and granzyme B. Red blood cells were then lysed using lysing solution, and the stained cells were acquired on four-color flow cytometer. RESULT: Our results indicated 15% of lymphocytes in peripheral blood of patients with BC and HDs had NK cells phenotype. However, the frequency of total NK cells (CD3-CD56+), and NK subsets (based on conventional and new classifications) was not significantly different between patients and HDs. We observed mean fluorescent intensity (MFI) of CXCR3 in total NK cells (p = .02) and the conventional cytotoxic (CD3-CD56dim CD16+) NK cells (p = .03) were significantly elevated in the patients with BC compared to HDs. Despite this, the MFI of granzyme B expression in conventional regulatory (CD3-CD56brightCD16- /+) NK cells and CD3-CD56-CD16+ NK cells (p = .03 and p = .004, respectively) in the patients was lower than healthy subjects. CONCLUSION: The higher expression of chemokine receptor CXCR3 on total NK cells in patients with BC may be associated with increased chemotaxis-related NK cell infiltration. However, lower expression of granzyme B in conventional regulatory NK cells and CD3-CD56-CD16+ NK cells in the patients compared to HDs suggests reduced cytotoxic activity of the NK cells in BC. These results might demonstrate accumulating NK subsets with a dysfunctional phenotype in the peripheral blood of patients with BC.

CD109 identified in circulating proteomics mitigates postoperative recurrence in chronic rhinosinusitis with nasal polyps by suppressing TGF-ß1-induced epithelial-mesenchymal transition.

BACKGROUND: Chronic rhinosinusitis with nasal polyps (CRSwNP) is a common inflammatory disorder with a high rate of recurrence. This study aimed to explore biomarkers for identifying patients with recurrent CRSwNP (rCRSwNP). METHODS: We recruited two independent cohorts. In the discovery cohort, rCRSwNP patients and non-recurrent CRSwNP (non-rCRSwNP) patients were recruited, and the serum proteomic profile was characterized. The top 5 upregulated and downregulated proteins were confirmed in the validation cohort by ELISA, WB, and qRT-PCR, and their predictive values for postoperative recurrence were assessed. In vitro, human nasal epithelial cells (HNEpCs) were employed to assess the ability of candidate proteins to induce epithelial-mesenchymal transition (EMT). RESULTS: Serum proteomics identified 53 different proteins, including 30 increased and 23 decreased, between the rCRSwNP and non-rCRSwNP groups. ELISA results revealed that serum levels of CD163 and TGF-ß1 were elevated, CD109 and PRDX2 were decreased in the rCRSwNP group compared to the non-rCRSwNP group, and serum CD163, TGF-ß1, and CD109 levels were proved to be associated with the risk of postoperative recurrence. In addition, qRT-PCR and WB revealed that tissue CD163, TGF-ß1, and CD109 expressions in rCRSwNP patients were enhanced compared to those non-rCRSwNP patients. Kaplan-Meier analysis showed that increased CD163 and TGF-ß1 expression and decreased CD109 expression are associated with the risk of recurrence in CRSwNP patients. Receiver operating characteristic curves showed that TGF-ß1 and CD109 had superior diagnostic performances for rCRSwNP. In vitro experiments showed that TGF-ß1 promoted EMT in HNEpCs, and overexpression of CD109 reversed this effect. Functional recovery experiments confirmed that CD109 could attenuate EMT in HNEpCs by inhibiting the TGF-ß1/Smad signaling pathway, attenuating EMT in epithelial cells. CONCLUSION: Our data suggested that TGF-ß1 and CD109 might serve as promising predictors of rCRSwNP. The TGF-ß1/Smad pathway was implicated in fostering EMT in epithelial cells, particularly those exhibiting low expression of CD109. Consequently, the absence of CD109 expression in epithelial cells could be a potential mechanism underlying rCRSwNP.

Lipopolysaccharide, VE-cadherin, HMGB1, and HIF-1α levels are elevated in the systemic circulation in chronic migraine patients with medication overuse headache: evidence of leaky gut and inflammation.

OBJECTIVE: Medication overuse headache (MOH) was recently shown to be associated with leaky gut in rodents. We aimed to investigate whether chronic migraine (CM) patients with MOH have elevated lipopolysaccharide levels and inflammatory molecules in blood circulation. MATERIALS AND METHODS: The study included women participants (40 CM patients with NSAID overuse headache, 35 episodic migraine (EM) patients, and 20 healthy non-headache sufferers). Migraine duration, monthly migraine headache days, MigSCog, HADS-D, HADS-A, and HIT-6 scores were recorded. Serum samples were collected to measure circulating LPS, LPS binding protein (LBP), tight junction protein occludin, adherens junction protein vascular endothelial cadherin (VE-cadherin), CGRP, HMGB1, HIF-1α, IL-6, and IL-17 levels. RESULTS: Serum LPS, VE-Cadherin, CGRP, HIF-1α, and IL-6 levels were significantly higher in the CM + MOH group compared to the EM group and healthy controls while serum LBP and HMGB1 were higher in the CM + MOH group compared to healthy controls. IL-17 and occludin levels were comparable between the three groups. Serum HMGB1 levels in EM patients were higher compared to the control group. Mig-SCog and HIT-6 scores were higher in the CM + MOH group compared to EM patients. HADS-A and HADS-D scores were significantly higher in the CM + MOH group compared to EM patients and healthy controls, and they were also higher in EM patients compared to healthy subjects. LPS levels were correlated with VE-cadherin and occludin levels. The number of monthly migraine headache days was positively correlated with serum LPS, HIF-1α, VE-cadherin, and IL-6 levels, HADS-A, HADS-D, HIT-6, and MigSCog scores. CONCLUSION: We have evidence for the first time that CM + MOH is associated with elevated serum LPS and LBP levels suggestive of LPS leak into the systemic circulation. Higher levels of nociceptive and/or pro-inflammatory molecules such as HMGB1, HIF-1α, IL-6, and CGRP may play a role in trigeminal sensitization and neurobiology of MOH. Intestinal hyperpermeability and consequent inflammatory response should be considered as a potential contributory factor in patients with MOH.

[Ascites CD100 levels and immunomodulation effects in the peripheral blood of patients with liver cirrhosis combined with spontaneous bacterial peritonitis].

Objective: To observe the level and detection of ascites CD100 on the activity of CD4(+) and CD8(+) T lymphocytes in vitro in the peripheral blood of patients with liver cirrhosis combined with spontaneous bacterial peritonitis. Methods: Peripheral blood and ascites were collected from 77 cases of liver cirrhosis (49 patients with liver cirrhosis combined with simple ascites and 28 patients with liver cirrhosis combined with SBP), and peripheral blood was collected from 22 controls. Soluble CD100 (sCD100) in peripheral blood and ascites was detected by an enzyme-linked immunosorbent assay. Flow cytometry was used to detect membrane-bound CD100 (mCD100) on the surface of CD4(+) and CD8(+)T lymphocytes. CD4(+) and CD8(+)T lymphocytes in ascites were sorted. CD4(+)T lymphocyte proliferation, key transcription factor mRNA, and secreted cytokine changes, as well as CD8(+)T lymphocyte proliferation, important toxic molecule mRNA, and secreted cytokine changes, were detected after CD100 stimulation. The killing activity of CD8(+)T cells was detected by direct contact and indirect contact culture systems. Data conforming to normality were compared using one-way ANOVA, a student's t-test, or a paired t-test. Data that did not conform to a normal distribution were compared using either the Krusal-Willis test or the Mann-Whitney test. Results: There was no statistically significant difference in plasma sCD100 level between patients with liver cirrhosis combined simple ascites (1 415 ± 434.1) pg/ml, patients with liver cirrhosis combined with SBP (1 465 ± 386.8) pg/ml, and controls (1 355 ± 428.0) pg/ml (P = 0.655). The ascites sCD100 level was lower in patients with liver cirrhosis combined with SBP than that of patients with simple ascites [(2 409 ± 743.0) pg/ml vs. (2825±664.2) pg/ml, P=0.014]. There was no statistically significant difference in the level of mCD100 in peripheral blood CD4(+) and CD8(+) T lymphocytes among the three groups (P > 0.05). The levels of mCD100 in ascites CD4(+) and CD8(+) T lymphocytes were higher in patients with liver cirrhosis combined with SBP than those in patients with simple ascites (P < 0.05). CD100 stimulation had no significant effect on the proliferation of CD4(+) and CD8(+)T lymphocytes in the ascites of patients with liver cirrhosis combined with SBP (P > 0.05). There were no significant effects on the expression of transcription factors in effector CD4(+)T lymphocytes (T-bet, retinoic acid associated solitary nuclear receptor γt, aromatic hydrocarbon receptor) or secretion of cytokines (interferon-γ, 17, and 22) (P > 0.05). CD100 stimulation had increased the relative expression of perforin, granzyme B, and granlysin mRNA and the levels of secreted interferon-γ and tumor necrosis factor-α, killing activity in ascites CD8+ T lymphocytes of patients with liver cirrhosis combined with SBP (P < 0.05). Conclusion: The active form of CD100 is sCD100 instead of mCD100. There is an imbalance between the expression of sCD100 and mCD100 in the ascites of patients with cirrhosis combined with SBP. sCD100 can enhance the function of CD8(+)T lymphocytes in the ascites of patients with cirrhosis combined with SBP and thus is one of the potential therapeutic targets.

The impact of probiotic yogurt versus ordinary yogurt on serum sTWEAK, sCD163, ADMA, LCAT and BUN in patients with chronic heart failure: a randomized, triple-blind, controlled trial.

BACKGROUND: To date, no study has investigated the effects of probiotic yogurt as a functional food in patients with chronic heart failure (CHF). Therefore, the aim of this study was to compare the impact of probiotic yogurt versus ordinary yogurt on inflammatory, endothelial, lipid and renal indices in CHF patients. In this randomized, triple-blind clinical trial, 90 patients with CHF were randomly allocated into two groups to take either probiotic or ordinary yogurt for 10 weeks. Serum levels of soluble tumor necrosis factor-like weak inducer of apoptosis (sTWEAK), soluble cluster of differentiation 163 (sCD163), asymmetric dimethylarginine (ADMA), and lecithin cholesterol acyltransferase (LCAT) were measured by using ELISA kits, and blood urea nitrogen (BUN) was measured by calorimetry method at baseline and at the end of trial. The P-value <0.05 was defined as statistically significant. RESULTS: Seventy-eight patients completed the study. At the end of the intervention, the levels of sTWEAK in both groups increased significantly, and this increase was greater in the probiotic yogurt group [691.84 (335.60, 866.95)] compared to control group [581.96 (444.99, 929.40)], and the difference between the groups was statistically significant after adjusting for confounders (P-value: 0.257, adjusted P-value: 0.038). However, no significant differences were found between the groups in the cases of other study indices. CONCLUSION: Probiotic yogurt may be useful for improving the inflammatory status in patients with CHF through increasing sTWEAK levels, however, further studies are needed in this area. © 2022 Society of Chemical Industry.

Soluble Immune-Related Proteins as New Candidate Serum Biomarkers for the Diagnosis and Progression of Lymphangioleiomyomatosis.

Background: The goal of this study was to analyze serum from lymphangioleiomyomatosis (LAM) patients and healthy controls to identify novel biomarkers that could shed light on disease diagnosis and pathogenesis. Methods: From April 2017 to October 2019, qualified serum samples were obtained to explore differences in 59 immune proteins between 67 LAM patients and 49 healthy controls by the Luminex method. Results: We characterized 22 serum immune proteins that were differentially expressed in LAM patients compared with healthy people. Fifty-nine proteins were then classified into eight categories according to their biological function, and the results showed that LAM patients displayed significantly higher levels of growth factors (p = 0.006) and lower levels of costimulatory molecules (p = 0.008). LAG-3 was not only likely to have better predictive value than VEGF-D but also showed a significant difference between patients without elevated VEGF-D and healthy people. IL-18 was positively correlated with lung function and six-minute walk test (6MWT) distance and negatively correlated with St. George's Respiratory Questionnaire (SGRQ) score and pulmonary artery systolic pressure (PASP), which suggested that IL-18 was related to disease severity. PD-1 was significantly different between patients with pneumothorax and/or chylothorax and those without complications. Conclusion: We performed a large-scale serum immune factor analysis of LAM. Our study provides evidence that LAG-3 may be a novel candidate serum biomarker for the diagnosis of LAM. Future independent validation in prospective studies is warranted.

Dynamic personalized risk prediction in chronic heart failure patients: a longitudinal, clinical investigation of 92 biomarkers (Bio-SHiFT study).

The aim of our observational study was to derive a small set out of 92 repeatedly measured biomarkers with optimal predictive capacity for adverse clinical events in heart failure, which could be used for dynamic, individual risk assessment in clinical practice. In 250 chronic HFrEF (CHF) patients, we collected trimonthly blood samples during a median of 2.2 years. We selected 537 samples for repeated measurement of 92 biomarkers with the Cardiovascular Panel III (Olink Proteomics AB). We applied Least Absolute Shrinkage and Selection Operator (LASSO) penalization to select the optimal set of predictors of the primary endpoint (PE). The association between repeatedly measured levels of selected biomarkers and the PE was evaluated by multivariable joint models (mvJM) with stratified fivefold cross validation of the area under the curve (cvAUC). The PE occurred in 66(27%) patients. The optimal set of biomarkers selected by LASSO included 9 proteins: NT-proBNP, ST2, vWF, FABP4, IGFBP-1, PAI-1, PON-3, transferrin receptor protein-1, and chitotriosidase-1, that yielded a cvAUC of 0.88, outperforming the discriminative ability of models consisting of standard biomarkers (NT-proBNP, hs-TnT, eGFR clinically adjusted) - 0.82 and performing equally well as an extended literature-based set of acknowledged biomarkers (NT-proBNP, hs-TnT, hs-CRP, GDF-15, ST2, PAI-1, Galectin 3) - 0.88. Nine out of 92 serially measured circulating proteins provided a multivariable model for adverse clinical events in CHF patients with high discriminative ability. These proteins reflect wall stress, remodelling, endothelial dysfunction, iron deficiency, haemostasis/fibrinolysis and innate immunity activation. A panel containing these proteins could contribute to dynamic, personalized risk assessment.Clinical Trial Registration: 10/05/2013 https://clinicaltrials.gov/ct2/show/NCT01851538?term=nCT01851538&draw=2&rank=1 .

A comprehensive profiling of soluble immune checkpoints from the sera of patients with non-small cell lung cancer.

BACKGROUND: Immunotherapy was widely used for the treatment of non-small cell lung cancer (NSCLC). However, whether inhibition of immune checkpoints individually or simultaneously could improve the therapeutic efficacy of NSCLC remains to be investigated. Here, we explored the aberrant levels of several checkpoints and evaluated their potential diagnostic values for NSCLC. METHODS: Serum samples of 89 NSCLC patients and 57 healthy donors were collected from Nanjing Drum Tower Hospital between November 2019 and July 2020. Fourteen human immune checkpoints were quantified by Procarta-Plex Human Immuno-Oncology Checkpoint Panel. RESULTS: The expression levels of sTIM-3, sCD137, sCD27, sLAG-3, sIDO, sPD-L2, sCD152, sCD80, and sPD-1 were all significantly increased in serum of NSCLC patients. Especially, sLAG-3 was significantly elevated in serum of NSCLC patients at early-stage (stages I and II), TIM-3, CD137, and CD27 were significantly higher in the advanced NSCLC patients (stages III and IV) than in the early-stage groups. Receiver operating characteristics (ROC) results showed that except for PD-1, all the other immune checkpoint proteins had potential diagnostic values for NSCLC. sTIM-3 had the highest diagnostic accuracy, followed by sLAG-3. Combining sTIM-3, sLAG-3, and sCD137 could increase the accuracy to a higher level. Moreover, sCD27 was correlated with NSCLC cancer type, age, sex, and disease stage, while sCD137 was correlated with age and disease stage. sTIM-3 and sIDO were correlated with stage and age, respectively. CONCLUSIONS: TIM-3 and LAG-3 were independent biomarkers for the early diagnosis of NSCLC. The combination of TIM-3, LAG-3, and CD137 could increase the diagnostic accuracy.

Association between occupational lead exposure and immunotoxicity markers: A systematic review and meta-analysis.

Recent evidences suggest the role of chronic lead (Pb) exposure in altering immunological parameters. Present study aimed to systematically review existing literature and synthesize quantitative evidence on the association between chronic Pb exposure and changes in immunological markers. Observational studies reporting immunological markers such as leukocyte derivative counts (CD3+, CD4+, CD8+, CD45+, CD56+, lymphocyte, and total leukocyte), cytokine, Immunoglobulin (Igs), C-reactive protein (CRP) among Pb-exposed and unexposed controls were systematically searched from PubMed, Scopus and Embase digital databases from inception to January 2021. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines were adhered during systematic review. Mean differences in the immunological markers between Pb-exposed and control groups were pooled using random-effects model. The heterogeneity was assessed using Cochran-Q test and I2 statistic. The review included forty studies reporting immunological markers in Pb-exposed and unexposed control groups. The occupational Pb-exposed group exhibited significantly higher BLL, impaired immunological markers, characterized by a marginal lowering in lymphocyte count, lymphocyte subsets (CD3+, CD4+, CD4+/CD8+ ratio), IFN-γ and IgG levels, while CD8+, IgM, IgA, IgE, and cytokines (IL-4, IL-6, IL-10, and TNF-α) exhibited a trend of higher values in comparison to the control group. Further, inflammatory marker viz., total leukocyte count was significantly higher among Pb-exposed. The included studies exhibited high levels of heterogeneity. In conclusion, Occupational Pb exposure alters the immunological markers such as the circulating cytokines and leukocyte counts. However, high-quality, multicentered studies are required to strengthen present observations and further understand the Pb's role on the immune system. Prospero Registration ID: CRD42021228252.

Serum Carcinoembryonic Antigen-Related Cell Adhesion Molecule 1 Level in Patients with Hepatocellular Carcinoma.

OBJECTIVE: To evaluate the clinical value of carcinoembryonic-antigen-related cell-adhesion molecule 1 (CEACAM1) in predicting the severity of hepatocellular carcinoma(HCC). METHODS: We evaluated 40 healthy subjects and 40 HCC patients by collecting venous blood for the comparison. Serum CEACAM1 was detected using the Human CEACAM1 ELISA Kit. Other laboratory chemistries were analyzed by standard methods. RESULTS: The serum level of CEACAM1 was not different between HCC patients and healthy subjects (p=0.0069). There was a correlation between serum CEACAM1 level and total bilirubin, and direct bilirubin. There was also a statistically significant difference among serum CEACAM1 levels stratified by BCLC staging and MELD score at the cut-point of 18. Lower platelet count, higher levels of aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase were observed in HCC patients. CONCLUSION: An increase of serum CEACAM1 level was associated with cholestasis. The role of this molecule in HCC diagnosis was unclear. However, serum CAECAM1 may be useful to predict the severity in HCC patients.

Serum LAG-3 Predicts Outcome and Treatment Response in Hepatocellular Carcinoma Patients With Transarterial Chemoembolization.

Background: Transarterial chemoembolization (TACE) stands for the most commonly utilized therapy for hepatocellular carcinoma (HCC) worldwide. This study was to explore the potential predictive and prognostic roles of LAG-3 and PD-L1 as serum biomarkers in HCC patients underwent TACE treatment. Methods: A total of 100 HCC patients receiving TACE as well as 30 healthy controls were enrolled in the study. Serum LAG-3 and PD-L1 levels were determined at baseline and 3 day after TACE using enzyme-linked immunosorbent assay (ELISA). Results: We found serum levels of LAG-3 and PD-L1 were significantly elevated in HCC patients compared with healthy controls. Interestingly, patients with low pre-TACE and post-TACE levels of LAG-3 but not PD-L1 had a high probability of achieving an objective response (OR) after TACE treatment. Additionally, high pre-TACE LAG-3 level was correlated with poor disease outcome, and the patients with both high serum LAG-3 and PD-L1 level had the shorter overall survival (OS) than patients who are either PD-L1 or LAG-3 high or both PD-L1 and LAG-3 low. High pre-TACE serum LAG-3 level was positively associated with more cirrhosis pattern, advanced BCLC stage, pre-TACE alanine aminotransferase (ALT) level, and pre-TACE aspartate aminotransferase (AST) level. Furthermore, in 50 patients who underwent TACE, the serum LAG-3 level was significantly decreased at 3 day after TACE. Conclusion: Both pre-TACE and post-TACE serum LAG-3 levels could serve as powerful predictors for tumor response of TACE, and high pre-TACE serum LAG-3 level was an indicator for poor prognosis in HCC.

Identification of Novel Low-Density Neutrophil Markers Through Unbiased High-Dimensional Flow Cytometry Screening in Non-Small Cell Lung Cancer Patients.

Neutrophils have been described as a phenotypically heterogeneous cell type that possess both pro- and anti-tumor properties. Recently, a subset of neutrophils isolated from the peripheral blood mononuclear cell (PBMC) fraction has been described in cancer patients. These low-density neutrophils (LDNs) show a heterogeneous maturation state and have been associated with pro-tumor properties in comparison to mature, high-density neutrophils (HDNs). However, additional studies are necessary to characterize this cell population. Here we show new surface markers that allow us to discriminate between LDNs and HDNs in non-small cell lung cancer (NSCLC) patients and assess their potential as diagnostic/prognostic tool. LDNs were highly enriched in NSCLC patients (median=20.4%, range 0.3-76.1%; n=26) but not in healthy individuals (median=0.3%, range 0.1-3.9%; n=14). Using a high-dimensional human cell surface marker screen, we identified 12 surface markers that were downregulated in LDNs when compared to HDNs, while 41 surface markers were upregulated in the LDN subset. Using flow cytometry, we confirmed overexpression of CD36, CD41, CD61 and CD226 in the LDN fraction. In summary, our data support the notion that LDNs are a unique neutrophil population and provide novel targets to clarify their role in tumor progression and their potential as diagnostic and therapeutic tool.

Potential value of circulatory microRNA10b gene expression and its target E-cadherin as a prognostic and metastatic prediction marker for breast cancer.

BACKGROUND: Breast cancer (BC) is the leading cause of cancer death in women worldwide. Most BC studies on candidate microRNAs were tissue specimen based. Recently, there has been a focus on the study of cell-free circulating miRNAs as promising biomarkers in (BC) diagnosis and prognosis. Therefore, we aimed to investigate the circulating levels of miR-10b and its target soluble E- cadherin as potentially easily accessible biomarkers for breast cancer. METHODS: Sixty-one breast cancer patients and forty-eight age- and sex-matched healthy volunteers serving as a control group were enrolled in the present study. Serum samples were used to assess miRNA10b expression by TaqMan miRNA assay technique. In addition, soluble E-cadherin expression level in serum was determined using ELISA technique. RESULT: Circulating miR-10b expression level and serum sE-cadherin was significantly upregulated in patients with BC compared to controls. Moreover, serum miR-10b displayed progressive up-regulation in advanced stages with higher level in metastatic compared to non-metastatic BC. Additionally, the combined use of both serum miR-10b and sE-cadherin revealed the highest sensitivity and specificity for detection of BC metastasis (92.9% and 97.9% respectively) with an area under curve (AUC) of 0.98, 95% CI (0.958-1.00). CONCLUSION: Our data suggest that circulating miR-10b could be utilized as a potential non-invasive serum biomarker for diagnosis and prognosis of breast cancer with better performance to predict BC metastasis achieved on measuring it simultaneously with serum sE-cadherin. Further studies with a large cohort of patients are warranted to validate the serum biomarker for breast cancer management.

Genetic signature of CTLA-4, BTLA, TIM-3 and LAG-3 molecular expression in colorectal cancer patients: Implications in diagnosis and survival outcomes.

OBJECTIVE: Accumulating evidences suggest that immune checkpoints (ICs) inhibit immune response against cancerous cells and promote tumor cell survival. Up-regulation of ICs in tumor microenvironment is reported in patients with colorectal cancer (CRC). Thus, evaluating the peripheral blood expression of ICs may be used as non-invasive biomarkers for diagnosis and prognosis of CRC. METHODS: This study included 60 primary and treatment naïve CRC patients along with 15 age and sex matched healthy volunteers as a control group. Total RNA was extracted from peripheral blood samples and gene expression of cytotoxic T lymphocyte antigen-4 (CTLA-4), B and T lymphocyte attenuator (BTLA), T-cell immunoglobulin and mucin domain-containing protein 3 (TIM-3), and Lymphocyte activation gene-3 (LAG-3) was measured by quantitative real time polymerase chain reaction (qRT-PCR). All patients were followed for 12 months to correlate the measured ICs to patients' survival. RESULTS: The gene expression of CTLA-4, BTLA, TIM-3 and LAG-3 was significantly up-regulated in CRC patients compared to the control group (p < 0.001). Individually, CTLA-4 and BTLA showed 85% sensitivity in discriminating CRC patients from control group (p < 0.001). On the other hand, TIM-3 and LAG-3 expression showed higher sensitivity (93%) for diagnosis of CRC (p < 0.001). Conversely, CTLA-4 or BTLA strongly predicted CRC patients' survival (p < 0.001) compared to TIM-3 (p = 0.018) or LAG-3 (p = 0.035). CTLA-4, BTLA, TIM-3 and LAG-3 were independent prognostic factors of survival after adjustment for age and gender. CONCLUSION: The current study provided evidence that blood gene expression of ICs was up-regulated in CRC patients and associated with cancer stage and patients' survival, which highlights the diagnostic and prognostic values of ICs expression in CRC. Further investigations and validations in larger cohorts are required.

ST6GAL1 and α2-6 Sialylation Regulates IL-6 Expression and Secretion in Chronic Obstructive Pulmonary Disease.

Chronic obstructive pulmonary disease (COPD) is a systemic disease strongly associated with cigarette smoking, airway inflammation, and acute disease exacerbations. Changes in terminal sialylation and fucosylation of asparagine (N)-linked glycans have been documented in COPD, but the role that glycosyltransferases may play in the regulation of N-linked glycans in COPD has not been fully elucidated. Recent studies suggest that modulation of ST6GAL1 (ST6 beta-galactoside alpha-2,6-sialyltransferase-1), which catalyzes terminal α2-6 sialylation of cellular proteins, may regulate inflammation and contribute to COPD phenotype(s). Interestingly, it has been previously demonstrated that ST6GAL1, a Golgi resident protein, can be proteolytically processed by BACE1 (beta-site amyloid precursor protein cleaving enzyme-1) to a circulating form that retains activity. In this study, we showed that loss of ST6GAL1 expression increased interleukin (IL)-6 expression and secretion in human bronchial epithelial cells (HBECs). Furthermore, exposure to cigarette smoke medium/extract (CSE) or BACE1 inhibition resulted in decreased ST6GAL1 secretion, reduced α2-6 sialylation, and increased IL-6 production in HBECs. Analysis of plasma ST6GAL1 levels in a small COPD patient cohort demonstrated an inverse association with prospective acute exacerbations of COPD (AECOPD), while IL-6 was positively associated. Altogether, these results suggest that reduced ST6GAL1 and α2-6 sialylation augments IL-6 expression/secretion in HBECs and is associated with poor clinical outcomes in COPD.

S100A9/CD163 expression profiles in classical monocytes as biomarkers to discriminate idiopathic pulmonary fibrosis from idiopathic nonspecific interstitial pneumonia.

Circulating monocytes have pathogenic relevance in idiopathic pulmonary fibrosis (IPF). Here, we determined whether the cell surface levels of two markers, pro-inflammatory-related S100A9 and anti-inflammatory-related CD163, expressed on CD14strongCD16- classical monocytes by flow cytometry could discriminate IPF from idiopathic nonspecific interstitial pneumonia (iNSIP). Twenty-five patients with IPF, 25 with iNSIP, and 20 healthy volunteers were prospectively enrolled in this study. The S100A9+CD163- cell percentages in classical monocytes showed a pronounced decrease on monocytes in iNSIP compared to that in IPF. In contrast, the percentages of S100A9-CD163+ cells were significantly higher in iNSIP patients than in IPF patients and healthy volunteers. In IPF patients, there was a trend toward a correlation between the percentage of S100A9+CD163- monocytes and the surfactant protein-D (SP-D) serum levels (r = 0.4158, [95% confidence interval (CI) - 0.02042-0.7191], p = 0.051). The individual percentages of S100A9+CD163- and S100A9-CD163+ cells were also independently associated with IPF through multivariate regression analysis. The unadjusted area under the receiver operating characteristic curve (ROC-AUC) to discriminate IPF from iNSIP was (ROC-AUC 0.802, 95% CI [0.687-0.928]), suggesting that these are better biomarkers than serum SP-D (p < 0.05). This preliminary study reports the first comparative characterization of monocyte phenotypes between IPF and iNSIP.

Assessment of the relationship between semaphorin4D level and recurrence after catheter ablation in paroxysmal atrial fibrillation.

OBJECTIVE: Semaphorin4D (Sema4D), a novel integral membrane glycoprotein, plays a role in atherosclerosis, angiogenesis and chronic inflammation. Elevated levels of sema4D were presented in myocardial infarction, heart failure and atrial fibrillation. Aim of the study was to investigate the relation between sema4D and recurrence after catheter ablation (CA) in paroxysmal AF. METHODS: The present study included 161 paroxysmal AF patients (PAF) (101 patients undergone CA) and 60 healthy subjects. Serum levels of sema4D were measured and study participants were followed-up for 3 months and 1 year since CA in terms of recurrence respectively. RESULTS: Sema4D levels were significantly elevated in the recurrent group compared to the non-recurrent PAF patients (p < 0.001). Sema4D was importantly positively correlated with both left atrial volume index (r = 0.51, p < 0.013) and high sensitive C-reactive protein (r = 0.38), p < 0.011). In multivariate analysis, sema4D [odds ratio (OR) = 1.23, 95% CI 1.11-1.42; p < 0.001] and left atrial diameter (OR = 1.13, 95% CI 1.02-1.23; p = 0.012) were found to be significant independent risk parameters for recurrence. CONCLUSIONS: Sema4D is a novel biomarker that may help to identify individuals with recurrence after CA procedure in long term period in PAF.

Combining Magnetic Resonance Imaging with Systemic Monocyte Evaluation for the Implementation of GBM Management.

Magnetic resonance imaging (MRI) is the gold standard for glioblastoma (GBM) patient evaluation. Additional non-invasive diagnostic modalities are needed. GBM is heavily infiltrated with tumor-associated macrophages (TAMs) that can be found in peripheral blood. FKBP51s supports alternative-macrophage polarization. Herein, we assessed FKBP51s expression in circulating monocytes from 14 GBM patients. The M2 monocyte phenotype was investigated by qPCR and flow cytometry using antibodies against PD-L1, CD163, FKBP51s, and CD14. MRI assessed morphologic features of the tumors that were aligned to flow cytometry data. PD-L1 expression on circulating monocytes correlated with MRI tumor necrosis score. A wider expansion in circulating CD163/monocytes was measured. These monocytes resulted in a dramatic decrease in patients with an MRI diagnosis of complete but not partial surgical removal of the tumor. Importantly, in patients with residual tumor, most of the peripheral monocytes that in the preoperative stage were CD163/FKBP51s- had turned into CD163/FKBP51s+. After Stupp therapy, CD163/FKBP51s+ monocytes were almost absent in a case of pseudoprogression, while two patients with stable or true disease progression showed sustained levels in such circulating monocytes. Our work provides preliminary but meaningful and novel results that deserve to be confirmed in a larger patient cohort, in support of potential usefulness in GBM monitoring of CD163/FKBP51s/CD14 immunophenotype in adjunct to MRI.