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Objective: To explore the application value of combined detection of serum interleukin-6 (IL-6), stromal cell-derived factor-1 (SDF-1) and neutrophils CD64 (CD64) in the diagnosis of early postoperative infection after limb fractures., Methods: 419 patients with limb fractures un erwent surgical treatment in our hospital were selected as study subjects, and they were divided into infection group ( n  = 104) and non-infection group ( n  = 315) according to postoperative pathological results of puncture to detect the levels of serum IL-6, SDF-1 and CD64 in the two groups on the 1st, 3rd, 5th and 7th day after surgery, and to analyze the clinical diagnostic efficacy of single and combined detection of the three indexes in early postoperative infection after limb fractures by ROC curve., Results: The levels of serum IL-6, SDF-1 and CD64 in infection group were overtly higher than those in non-infection group at different time after surgery ( P  < 0.05), and the AUC value, specificity and sensitivity of combined detection were higher than those of single diagnosis. In this study, 14 patients in infection group underwent reoperation, 22 patients received conservative treatment in the later period, 6 patients had postoperative muscular dystrophy, and the remaining patients had a good prognosis., Conclusion: Serum IL-6, SDF-1 and CD64 are closely related to the occurrence of early postoperative infection after limb fractures, and their combination is helpful to improve diagnostic accuracy of early postoperative infection and provide effective reference value for the treatment of postoperative infection in orthopedics.

Background: A previous single-center study of patients with myocardial infarction (MI) showed that platelet FcγRIIa (pFCG) can distinguish patients at higher and lower risk of subsequent MI, stroke, and death., Objectives: The authors performed an 800-patient 25-center study to validate the prognostic implications of pFCG., Methods: Patients with type 1 MI (ST-segment elevation and non-ST-segment elevation) were enrolled in a prospective noninterventional trial during their index hospitalization. Enrolled patients had at least 2 of the following characteristics: age ≥65 years, multivessel coronary artery disease, previous MI, chronic kidney disease, or diabetes mellitus. Flow cytometry was used to quantify pFCG at a core laboratory. A predefined threshold was used to identify high and low pFCG. Patients were queried every 6 months by telephone with a standardized questionnaire. Events were confirmed by review of medical records., Results: Treatment with antithrombotic therapy (aspirin, P2Y 12 inhibitors, and anticoagulants) was similar in patients with high and low pFCG. The primary composite endpoint (MI, stroke, death) occurred more frequently in patients with high pFCG (HR: 2.09; 95% CI: 1.34-3.26; P = 0.001). Among individual components of the composite, both death (HR: 2.57; 95% CI: 1.50-4.40; P = 0.001) and MI (HR: 3.24; 95% CI: 1.64-6.37; P = 0.001) were more frequent in patients with high pFCG., Conclusions: Quantifying pFCG identifies patients at higher and lower risk of subsequent cardiovascular events. This prognostic information will be useful in clinical decisions regarding the intensity and duration of antiplatelet therapy. (Assessment of Individual Risk of Cardiovascular Events by Platelet FcγRIIa; NCT05175261)., Competing Interests: Funding Support and Author Disclosures This study was supported by Prolocor. Dr Schneider is named inventor on patents (U.S. 10,502,737, U.S. 11,747,335 B2) that propose the use of FcγRIIa for assaying platelet reactivity and treatment selection. Drs Schneider and DiBattiste are co-founders of Prolocor. Dr Angiolillo has received consulting fees or honoraria from Abbott, Amgen, AstraZeneca, Bayer, Biosensors, Boehringer Ingelheim, Bristol Myers Squibb, Chiesi, CSL Behring, Daiichi-Sankyo, Eli Lilly, Faraday, Haemonetics, Janssen, Merck, Novartis, Novo Nordisk, PhaseBio, PLx Pharma, Pfizer, Sanofi, and Vectura, outside of the present work; and his institution has received research grants from Amgen, AstraZeneca, Bayer, Biosensors, CeloNova, CSL Behring, Daiichi-Sankyo, Eisai, Eli Lilly, Faraday, Gilead, Janssen, Matsutani Chemical Industry Co, Merck, Novartis, Osprey Medical, Renal Guard Solutions, and the Scott R. MacKenzie Foundation. Dr Effron has received retirement income and equity from Eli Lilly. All other authors have no relationships relevant to the contents of this paper to disclose., (Copyright © 2024 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Objective: To study the distribution characteristics of natural killer (NK) cells and their subsets in normal peripheral blood in China, and to explore their normal value and significance., Methods: In this study, peripheral blood was collected from 200 healthy adults. Their age range was 18-87 years. All the subjects were divided into 6 age groups: 18-30, 31-40, 41-50, 51-60, 61-70, and 71-87 years. With CD16, CD56, CD4, CD19, as surface markers, fluid cytology detection techniques were used to detect NK cells and the relative and absolute counts. SPSS 27.0 was used for systematic analysis of the data, and the measurement data were expressed as mean±standard deviations. A t test, variance analysis or rank sum test were performed to compare the differences between the age groups and the sex groups. The significance level was set at α=0.05, and P < 0.05 was considered statistically significant., Results: The range of NK B cells in the 200 healthy adult subjects was (0.46±0.24)×10 6 /L, that of CD3 - CD56 + NK cells was (13.14±7.56)×10 6 /L, that of CD56 dim CD16 + NK cells was (5.23±3.12)×10 6 /L, that of CD56 bright NK cells was (85.61±7.40)×10 6 /L, and that of NK T cells was (4.16±3.34)×10 6 /L. There were no statistically significant differences in CD3 - CD56 + NK cells and NK T cells with respect to age ( P = 0.417, P =0.217). However, there was a decreasing trend in the number of NK B cells and CD56 dim CD16 + NK cells with increasing age ( r =0.234, P < 0.001; r =0.099, P < 0.001), particularly after the age of 50. Conversely, CD56 bright NK cells showed an increasing trend with age ( r =0.143, P < 0.001)., Conclusion: The detection of NK cells and their subsets has significant reference value for the diagnosis, treatment, and prognosis of autoimmune diseases, infectious diseases, and tumors. This study provides a preliminary reference range for clinical detection of NK cell subsets, but further research with a larger sample size and multi-center trials are needed to confirm these findings.

Esophageal cancer (EC) poses a significant global health burden, necessitating effective treatment strategies. Immune checkpoint inhibitors have emerged as a promising therapeutic option for EC, but the identification of predictive biomarkers remains crucial for optimizing patient outcomes. We conducted a retrospective analysis of medical records from advanced esophageal squamous cell carcinoma patients treated with first-line programmed death 1 inhibitors. Peripheral blood lymphocyte subpopulations were evaluated using flow cytometry, while hematological tests provided data on neutrophil, lymphocyte, and monocyte counts. Cox regression and logistic regression analyses were employed to explore the association between lymphocyte subpopulations, baseline characteristics, and progression-free survival (PFS). Among the 100 initially included patients, 70 met eligibility criteria. Multivariate Cox regression analysis revealed a significant association between high CD16+CD56+ lymphocyte proportions and longer PFS, independent of other clinical variables. Similarly, a high CD4+/CD8+ ratio was correlated with prolonged PFS. Kaplan-Meier survival curves supported these findings. Logistic regression analysis indicated no significant differences in the CD4+/CD8+ ratio and CD16+CD56+ lymphocytes concerning baseline characteristics, suggesting their potential as independent prognostic markers. Our study highlights the predictive value of peripheral blood CD16+CD56+ lymphocytes and the CD4+/CD8+ ratio for the efficacy of programmed death 1 inhibitors in advanced esophageal squamous cell carcinoma patients. These findings underscore the importance of peripheral blood biomarkers in guiding personalized immunotherapy strategies and improving outcomes for EC patients., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2024 the Author(s). Published by Wolters Kluwer Health, Inc.)

Globally, sepsis is the primary cause of death for children. While research conducted on adults has a significant impact on the diagnosis and treatment of sepsis in newborns and young children, there are significant factors that are pertinent to pediatrics as well. This prospective case-control study was conducted during the period from August 2020 to October 2022 after approval by the institutional ethical committee. The study included 48 critically ill children admitted at the Pediatric intensive care unit and 30 apparently healthy children as a control group. Laboratory investigations including complete blood picture, C-reactive protein (CRP), blood culture and sensitivity and plasma level of cluster of differentiation 64 (CD64) and myeloperoxidase (MPO) were investigated for all participants. A daily follow up for the signs of systemic inflammatory response syndrome (SIRS) or sepsis was done, and the patients were divided into SIRS and non-SIRS subgroups then patients were divided into two groups according to the presence of severe sepsis. A follow up CD64 and MPO sample were withdrawn from them to assess their prognostic value. SIRS was reported in 39.58 % of patients while severe sepsis was reported in 20.8%. CD64 and MPO were significantly higher in cases than controls (p= 0.003, p < 0.001, respectively) and, in patients with SIRs and severe sepsis CD64 was 1559.00± 367.09 pg/ml and 1547.9 4± 436.14 pg/ml, respectively. Also, MPO was significantly higher in patients with SIRS (113.58± 25.19 mU/ml) and severe sepsis (111.70± 26.50 mU/ml). CD64 and MPO significantly increased after development of sepsis in admitted patients. ROC was significantly higher for CD64 and MPO at admission than that for CRP at admission (p=0.123, p=0.014, respectively). In conclusion, plasma level of CD64 and MPO in peripheral blood can be considered an early sensitive marker for the detection and follow up of pediatric sepsis., (Copyright© by the Egyptian Association of Immunologists.)

Objective: Biomarkers with high accuracy for identification of infection in decompensated chronic liver disease (DCLD) are urgently needed. We compared the accuracy of neutrophilic cluster of differentiation 64 (nCD64) with procalcitonin for diagnosis of bacterial infection in children with DCLD., Methods: Consecutive children admitted with DCLD were enrolled prospectively. nCD64 was assessed by flow cytometry and expressed in percentage. nCD64, procalcitonin and hemogram were measured at admission and 7-14 days after treatment in those with infection. Complete work-up for infection was done. Presence, site and severity of infection was classified as per guidelines., Results: 107 children [64 boys, age 97(18-168) months] were enrolled. 78(72.9%) had infection, 26(24%) had severe sepsis and 60(56%) had systemic inflammatory response syndrome. The commonest site of infection was ascitic fluid (n=37), followed by pneumonia (n=24), urinary tract (n=15), bacteraemia (n=10), cholangitis (n=8) and cellulitis (n=3). nCD64 (cut-off-51%, AUC-0.82) had a higher sensitivity (79.5%) and specificity (82.8%) than procalcitonin (cut-off ≥0.58ng/mL, AUC-0.74, sensitivity-76.9% and specificity-62.1%) for diagnosis of infection. nCD64 and procalcitonin correlated with infection severity, being highest in children with severe sepsis [88(71-97) %and 1.98(0.83-10.36) ng/mL], than in infection alone [72(45-84) % and 1.09(0.45-2.07) ng/mL], and no-infection [36(20.2-48) % and 0.42(0.19-1.08) ng/mL]. There was no difference in diagnostic utility of procalcitonin or nCD64 with different sites of infection. Elevation of all 3 parameters (nCD64, PCT and total leukocyte count) was uncommon but highly specific for presence of infection., Conclusion: nCD64 identifies infection better than procalcitonin and correlates well with infection severity in children with DCLD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Masson SAS.)

Sepsis is a life-threatening immune response to infection in the body, eventually resulting in fatal organ failure. Current methods utilize blood cultures and quick-Sequential-Organ-Failure-Assessment (qSOFA), but there is a need for more accurate and time-sensitive diagnostic methods to improve survival rates. We present a 3D-printed microfluidic chip that bioconjugates antibodies CD69, CD64, and CD25 to channel surfaces to capture sepsis cells in blood samples and validate it with clinical samples (n = 125 septic, n = 10 healthy). Other variables were taken such as healthy volunteer blood samples and patient demographics to validate and confirm our device's diagnostic ability. Statistical differences were found between healthy volunteer and sepsis patient antigen cell counts (CD69 p-value < 0.001, CD64 p-value < 0.004, CD25 p-value < 0.0009), and were confirmed using principal component analysis. Demographics such as length of stay, age, culture results, and need for surgery also factored into sepsis detection on a smaller scale than the antigen cell counts. The receiver operating characteristic (ROC) analysis showed an area under the curve (AUC) of 0.989, 0.988, and 0.992 for CD69, CD64, and CD25, respectively, and a combined biomarker panel of 0.997. Overall, the device performed within a shorter time frame of 4 h compared to standard blood culture tests and was validated for use in detecting sepsis in patients., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: D Pappas reports financial support was provided by National Institutes of Health. D Pappas reports a relationship with NaMi Diagnostics that includes: non-financial support. D Pappas has patent #10,761,093 licensed to NaMi Diagnostics. Dr. Pappas’s patent on this technology is licensed through NaMi Diagnostics, who have no competing interest in this work. Dr. Pappas has no financial position in NaMi Diagnostics. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2024 Elsevier B.V. All rights reserved.)

Objectives: This study was undertaken to assess CD91 expression on monocytes and changes in monocyte subset distribution during acute tissue damage and bloodstream infection (BSI)., Methods: We investigated blood specimens from healthy individuals, trauma and cardiac surgery patients as a model of tissue damage, and patients with BSI, by flow cytometry using a panel of antibodies comprising CD45, HLA-DR, CD14, CD16 and CD91 for the identification of monocyte subsets., Results: While infrequent in healthy subjects, CD91low/neg monocyte levels were markedly high in BSI, trauma and after cardiac surgery. This monocyte subset expanded up to 15-fold in both patient cohorts, whereas CD14+CD16+ inflammatory monocytes were multiplied by a factor of 5 only. CD14+CD91low monocytes displayed a significantly lower density of HLA-DR and markedly reduced expression of CD300e, compared to the other subsets. They also expressed high levels of myeloperoxidase and showed robust phagocytic and oxidative burst activity., Conclusions: Expansion of CD91low monocytes is a sensitive marker of acute inflammatory states of infectious and non-infectious etiology., (© 2024 Walter de Gruyter GmbH, Berlin/Boston.)

Background: We aimed to explore the prognostic utilities of C-reactive protein (CRP), procalcitonin (PCT), neutrophil CD64 (nCD64) index, in combination or alone, in septic patients., Methods: We retrospectively included 349 septic patients (based on Sepsis 3.0 definition). The primary outcome was 28-day all-cause mortality. Cox regression model, receiver-operating characteristic (ROC) curve, reclassification analysis, Kaplan-Meier survival curves were performed to evaluate the predictive efficacy of the above parameters., Results: CRP, nCD64 index were independent predictors of 28-day mortality for sepsis in the Cox regression model [CRP, HR 1.004 (95% CI 1.002-1.006), P < 0.001; nCD64 index, HR 1.263 (95% CI 1.187-1.345, P < 0.001]. Area under the ROC curve (AUC) of CRP, PCT, nCD64 index, nCD64 index plus PCT, nCD64 index plus CRP, were 0.798 (95% CI 0.752-0.839), 0.833 (95% CI 0.790-0.871), 0.906 (95% CI 0.870-0.935), 0.910 (95% CI 0.875-0.938), 0.916 (95% CI 0.881-0.943), respectively. nCD64 plus CRP performed best in prediction, discrimination, and reclassification of the 28-day mortality risk in sepsis. The risk of 28-day mortality increased stepwise as the number of data exceeding optimal cut-off values increased., Conclusions: nCD64 index combined with CRP was superior to CRP, PCT, nCD64 index and nCD64 index plus PCT in predicting 28-day mortality in sepsis. Multi-marker approach could improve the predictive accuracy and be beneficial for septic patients., (© 2022. The Author(s).)

Rationale: Sepsis is the leading cause of death in adult ICUs. At present, sepsis diagnosis relies on nonspecific clinical features. It could transform clinical care to have immune-cell biomarkers that could predict sepsis diagnosis and guide treatment. For decades, neutrophil phenotypes have been studied in sepsis, but a diagnostic cell subset has yet to be identified. Objectives: To identify an early, specific immune signature of sepsis severity that does not overlap with other inflammatory biomarkers and that distinguishes patients with sepsis from those with noninfectious inflammatory syndrome. Methods: Mass cytometry combined with computational high-dimensional data analysis was used to measure 42 markers on whole-blood immune cells from patients with sepsis and control subjects and to automatically and comprehensively characterize circulating immune cells, which enables identification of novel, disease-specific cellular signatures. Measurements and Main Results: Unsupervised analysis of high-dimensional mass cytometry data characterized previously unappreciated heterogeneity within the CD64 + immature neutrophils and revealed two new subsets distinguished by CD123 and PD-L1 (programmed death ligand 1) expression. These immature neutrophils exhibited diminished activation and phagocytosis functions. The proportion of CD123-expressing neutrophils correlated with clinical severity. Conclusions: This study showed that these two new neutrophil subsets were specific to sepsis and detectable through routine flow cytometry by using seven markers. The demonstration here that a simple blood test distinguishes sepsis from other inflammatory conditions represents a key biological milestone that can be immediately translated into improvements in patient care.

Infection is one of the leading causes of death in burn patients. Many researchers regard neutrophil CD64 (nCD64) as a biomarker in the early diagnosis of burn patients with infection. Nevertheless, the conclusions are controversial. A comprehensive analysis of the diagnostic value of nCD64 for burn infection was performed in China using a meta-analysis method. Pubmed, Cochrane Library, Web of Science, Embase, China National Knowledge Infrastructure (CNKI), and China Wanfang databases were searched for studies on nCD64 as a diagnostic biomarker of burn patients with infection from the establishment of the databases to September 29, 2020. The data were analyzed by Stata 15.0 software. Six studies were identified. The results showed that the pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, and diagnostic odds ratio were 0.92 (95% confidence interval [CI]: 0.88~0.95), 0.82 (95% CI: 0.76~0.87), 5.10 (95% CI: 3.90~6.80), 0.10 (95% CI: 0.06~0.15), and 52 (95% CI: 29~94), respectively. The area under the curve was 0.94 (95% CI: 0.92~0.94). According to the analysis of the sepsis subgroup, it showed that nCD64 had good diagnostic value in the patients with burn sepsis in Chinese population. Neutrophil CD64 is highly efficient to diagnose burn infection in Chinese population. Therefore, nCD64 could be regarded as a valuable biomarker for the early diagnosis of burn infection in China, especially in patients with burn sepsis. Combined with other diagnostic indexes, nCD64 can be clinically used in the early diagnosis of burn infection to improve the sensitivity and specificity., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Burn Association. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Following therapy, breast cancer survivors (BCS) have an increased risk of infections because of age and cancer dysregulation of inflammation and neutrophil functions. Neutrophil functions may be improved by exercise training, although limited data exist on exercise and neutrophil functions in BCS.Sixteen BCS [mean age: 56 (SD 11) years old] completed 16 weeks of community-based exercise training and a 45-minute acute bout of cycling before (Base) and after (Final) the exercise training program. Exercise training consisted of 3 x 40 - 60 minute mixed mode aerobic exercises, comprising 10 - 30 minutes aerobic and 30 minutes resistance training. At Base and Final, we took BCS blood samples before (PRE), immediately after (POST), and 1 hour after (1Hr) acute exercise to determine neutrophil counts, phenotype, bacterial killing, IL-6, and IL-8 levels. Eleven healthy, age- and physical activity levels-matched women (Control) completed the acute bout of exercise once as a healthy response reference. Resting Responses. BCS and Controls had similar Base PRE absolute neutrophil counts [mean (SD): 3.3 (1.9) v 3.1 (1.2) x 10 9 /L, p=0.801], but BCS had lower bacterial phagocytosis [3991 (1233) v 4881 (417) MFI, p=0.035] and higher oxidative killing [6254 (1434) v 4709 (1220) MFI, p=0.005], lower CD16 [4159 (1785) v 7018 (1240) MFI, p<0.001], lower CXCR2 [4878 (1796) v 6330 (1299) MFI, p=0.032] and higher TLR2 [98 (32) v 72 (17) MFI, p=0.022] expression, while IL-6 [7.4 (5.4) v 4.0 (2.7) pg/mL, p=0.079] levels were marginally higher and IL-8 [6.0 (4.7) v 7.9 (5.0) pg/mL, p=0.316] levels similar. After 16 weeks of training, compared to Controls, BCS Final PRE phagocytosis [4510 (738) v 4881 (417) MFI, p=0.146] and TLR2 expression [114 (92) v 72 (17) MFI, p=0.148] were no longer different. Acute Exercise Responses. As compared to Controls, at Base, BCS phagocytic Pre-Post response was lower [mean difference, % (SD): 12% (26%), p=0.042], CD16 Pre-Post response was lower [12% (21%), p=0.016] while CD16 Pre-1Hr response was higher [13% (25%), p=0.022], TLR2 Pre-Post response was higher [15% (4%) p=0.002], while IL-8 Pre-Post response was higher [99% (48%), p=0.049]. As compared to Controls, following 16 weeks of training BCS phagocytic Pre-Post response [5% (5%), p=0.418], CD16 Pre-1Hr response [7% (7%), p=0.294], TLR2 Pre-Post response [6% (4%), p=0.092], and IL-8 Pre-Post response [1% (9%), p=0.087] were no longer different. Following cancer therapy, BCS may have impaired neutrophil functions in response to an acute bout of exercise that are partially restored by 16 weeks of exercise training. The improved phagocytosis of bacteria in BCS may represent an exercise-induced intrinsic improvement in neutrophil functions consistent with a reduced risk of infectious disease., Clinical Trial Registration: ClinicalTrials.gov, identifier NCT03760536., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Bartlett, Hanson, Lee, Wagoner, Harrell, Sullivan, Bates, Alzer, Amatuli, Deal, Jensen, MacDonald, Deal, Muss, Nyrop and Battaglini.)

The neonatal Fc receptor (FcRn) is a ubiquitously expressed protein historically involved in IgG and albumin recycling. Recent data suggest an involvement in the pathophysiology of antibody-mediated autoimmune diseases. Among them, systemic lupus erythematosus (SLE) implies clinical and biological abnormalities of innate and adaptive circulating immune cells, potentially involving newly described functions of FcRn. In this study, FcRn expression was assessed by flow cytometry in peripheral blood leukocytes of 41 SLE patients with either active or inactive disease and 32 healthy donors. FcRn expression in B cells, natural killer cells, and T cells of SLE patients was statistically lower as compared to healthy donors. Conversely, FcRn level was statistically higher in non-classical monocyte subpopulations (CD14+CD16+ monocytes) of SLE patients versus healthy donors providing an interesting perspective to further explore its role in SLE pathophysiology.

Standard-of-care infliximab dosing regimens were developed prior to the routine use of therapeutic drug monitoring and identification of target concentrations. Not surprisingly, subtherapeutic infliximab concentrations in pediatric Crohn's disease (CD) are common. The primary aim was to conduct a real-world pharmacokinetic (PK) evaluation to discover blood biomarkers of rapid clearance, identify exposure targets, and a secondary aim to translate PK modeling to the clinic. In a multicenter observational study, 671 peak and trough infliximab concentrations from 78 patients with CD were analyzed with a drug-tolerant assay (Esoterix; LabCorp, Calabasas, CA). Individual area under the curve (AUC) estimates were generated as a measure of drug exposure over time. Population PK modeling (nonlinear mixed-effect modeling) identified serum albumin, antibody to infliximab, erythrocyte sedimentation rate (ESR), and neutrophil CD64 as biomarkers for drug clearance. Week 14 and week 52 biochemical remitters (fecal calprotectin < 250 µg/g) had higher infliximab exposure (AUC) throughout induction. The optimal infliximab AUC target during induction for week 14 biochemical remission was 79,348 µg*h/mL (area under the receiver operating characteristic curve (AUROC) 0.77, [0.63-0.90], 85.7% sensitive, and 64.3% specific) with those exceeding the AUC target more likely to achieve a surgery-free week 52 biochemical remission (OR 4.3, [1.2-14.6]). Pretreatment predictors for subtherapeutic week 14 AUC included neutrophil CD64 > 6 (OR 4.5, [1.4-17.8]), ESR > 30 mm/h (OR 3.8, [1.4-11]), age < 10 years old (OR 4.2, [1.2-20]), and weight < 30 kg (OR 6.6, [2.1-25]). We created a decision-support PK dashboard with an iterative process and embedded the modeling program within the electronic health record. Model-informed precision dosing guided by real-world PKs is now available at the bedside in real-time., (© 2020 The Authors. Clinical Pharmacology & Therapeutics © 2020 American Society for Clinical Pharmacology and Therapeutics.)

SARS coronavirus 2 (SARS-CoV-2) is a novel viral pathogen that causes a clinical disease called coronavirus disease 2019 (COVID-19). Although most COVID-19 cases are asymptomatic or involve mild upper respiratory tract symptoms, a significant number of patients develop severe or critical disease. Patients with severe COVID-19 commonly present with viral pneumonia that may progress to life-threatening acute respiratory distress syndrome (ARDS). Patients with COVID-19 are also predisposed to venous and arterial thromboses that are associated with a poorer prognosis. The present study identified the emergence of a low-density inflammatory neutrophil (LDN) population expressing intermediate levels of CD16 (CD16Int) in patients with COVID-19. These cells demonstrated proinflammatory gene signatures, activated platelets, spontaneously formed neutrophil extracellular traps, and enhanced phagocytic capacity and cytokine production. Strikingly, CD16Int neutrophils were also the major immune cells within the bronchoalveolar lavage fluid, exhibiting increased CXCR3 but loss of CD44 and CD38 expression. The percentage of circulating CD16Int LDNs was associated with D-dimer, ferritin, and systemic IL-6 and TNF-α levels and changed over time with altered disease status. Our data suggest that the CD16Int LDN subset contributes to COVID-19-associated coagulopathy, systemic inflammation, and ARDS. The frequency of that LDN subset in the circulation could serve as an adjunct clinical marker to monitor disease status and progression.

The identification of a bacterial, viral, or even noninfectious cause is essential in the management of febrile syndrome in the emergency department (ED), especially in epidemic contexts such as flu or CoVID-19. The aim was to assess discriminative performances of two biomarkers, CD64 on neutrophils (nCD64) and CD169 on monocytes (mCD169), using a new flow cytometry procedure, in patients presenting with fever to the ED during epidemics. Eighty five adult patients presenting with potential infection were included during the 2019 flu season in the ED of La Timone Hospital. They were divided into four diagnostic outcomes according to their clinical records: no-infection, bacterial infection, viral infection and co-infection. Seventy six patients with confirmed SARS-CoV-2 infection were also compared to 48 healthy volunteers. For the first cohort, 38 (45%) patients were diagnosed with bacterial infections, 11 (13%) with viral infections and 29 (34%) with co-infections. mCD169 was elevated in patients with viral infections, with a majority of Flu A virus or Respiratory Syncytial Virus, while nCD64 was elevated in subjects with bacterial infections, with a majority of Streptococcus pneumoniae and Escherichia coli. nCD64 and mCD169 showed 90% and 80% sensitivity, and 78% and 91% specificity, respectively, for identifying patients with bacterial or viral infections. When studied in a second cohort, mCD169 was elevated in 95% of patients with SARS-CoV-2 infections and remained at normal level in 100% of healthy volunteers. nCD64 and mCD169 have potential for accurately distinguishing bacterial and acute viral infections. Combined in an easy and rapid flow cytometry procedure, they constitute a potential improvement for infection management in the ED, and could even help for triage of patients during emerging epidemics., (© 2021 International Society for Advancement of Cytometry.)

Background: The aim of the study was to conduct a meta-analysis to evaluate the accuracy of neutrophil CD64, procalcitonin (PCT), and interleukin-6 (IL-6) as markers for the diagnosis of sepsis in adult patients., Methods: Various databases were searched to collect published studies on the diagnosis of sepsis in adult patients using neutrophil CD64, PCT, and IL-6 levels. Utilizing the Stata SE 15.0 software, forest plots and the area under the summary receiver operating characteristic curves were drawn. The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, diagnostic odds ratio, and area under the curve (AUC) were calculated., Results: Fifty-four articles were included in the study. The pooled sensitivity, specificity, and AUC of neutrophil CD64 for the diagnosis of sepsis were 0.88 (95% confidence interval [CI], 0.81-0.92), 0.88 (95% CI, 0.83-0.91), and 0.94 (95% CI, 0.91-0.96), respectively. The pooled sensitivity, specificity, and AUC of PCT for the diagnosis of sepsis were 0.82 (95% CI, 0.78-0.85), 0.78 (95% CI, 0.74-0.82), and 0.87 (95% CI, 0.83-0.89), respectively. Subgroup analysis showed that the AUC for PCT diagnosis of intensive care unit (ICU) sepsis was 0.86 (95% CI, 0.83-0.89) and the AUC for PCT diagnosis of non-ICU sepsis was 0.82 (95% CI, 0.78-0.85). The pooled sensitivity, specificity, and AUC of IL-6 for the diagnosis of sepsis were 0.72 (95% CI, 0.65-0.78), 0.70 (95% CI, 0.62-0.76), and 0.77 (95% CI, 0.73-0.80), respectively., Conclusions: Of the three biomarkers studied, neutrophil CD64 showed the highest diagnostic value for sepsis, followed by PCT, and IL-6. On the other hand, PCT showed a better diagnostic potential for the diagnosis of sepsis in patients with severe conditions compared with that in patients with non-severe conditions.

Abstract: Sepsis occurs when an infection induces a dysregulated immune response, and is most commonly bacterial in origin. This condition requires rapid treatment for successful patient outcomes. However, the current method to confirm infection (blood culture) requires up to 48 h for a positive result and many true cases remain culture-negative. Therefore, new diagnostic tests are urgently needed. Recent clinical studies suggest that CD69, CD64, and CD25 may serve as useful biomarkers of sepsis. In this study, we evaluated the cecal ligation and puncture and cecal slurry mouse models as tools to study these biomarkers in young and aged mice, and elucidate the timeliness and specificity of sepsis diagnosis. Fluorescence-activated cell sorting analysis revealed that all three biomarkers were elevated on blood leukocytes during sepsis. CD69 was specifically upregulated during sepsis, while CD64 and CD25 were also transiently upregulated in response to sham surgery. The optimal biomarker, or combination of biomarkers, depended on the timing of detection, mouse age, and presence of surgery. CD69 demonstrated an excellent capacity to distinguish sepsis, and in some scenarios the diagnostic performance was enhanced by combining CD69 with CD64. We also analyzed biomarker expression levels on specific cell populations (lymphocytes, monocytes, and neutrophils) and determined the cell types that upregulate each biomarker. Elevations in blood biomarkers were also detected via microfluidic analyses; in this case CD64 distinguished septic mice from naive controls. Our results suggest that CD69 and CD64 are valuable biomarkers to rapidly detect sepsis, and that mouse models are useful to study and validate sepsis biomarkers., Competing Interests: The authors report no conflicts of interest., (Copyright © 2020 by the Shock Society.)

Recently, host whole blood gene expression signatures have been identified for diagnosis of tuberculosis (TB). Absolute quantification of the concentrations of signature transcripts in blood have not been reported, but would facilitate diagnostic test development. To identify minimal transcript signatures, we applied a transcript selection procedure to microarray data from African adults comprising 536 patients with TB, other diseases (OD) and latent TB (LTBI), divided into training and test sets. Signatures were further investigated using reverse transcriptase (RT)-digital PCR (dPCR). A four-transcript signature ( GBP6, TMCC1, PRDM1 , and ARG1 ) measured using RT-dPCR distinguished TB patients from those with OD (area under the curve (AUC) 93.8% (CI 95% 82.2-100%). A three-transcript signature ( FCGR1A, ZNF296, and C1QB ) differentiated TB from LTBI (AUC 97.3%, CI 95% : 93.3-100%), regardless of HIV. These signatures have been validated across platforms and across samples offering strong, quantitative support for their use as diagnostic biomarkers for TB., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Gliddon, Kaforou, Alikian, Habgood-Coote, Zhou, Oni, Anderson, Brent, Crampin, Eley, Heyderman, Kern, Langford, Ottenhoff, Hibberd, French, Wright, Dockrell, Coin, Wilkinson and Levin.)

Lower level of low-density lipoprotein cholesterol (LDL-C) is paradoxically associated with increased mortality in ST elevation myocardial infarction (STEMI) patients. The underlying mechanism remains unclear. In a cohort of 220 de novo STEMI patients receiving timely primary percutaneous coronary intervention, admission LDL-C was negatively associated with circulating CD14++CD16+ monocyte counts. Moreover, admission LDL-C < 85 mg/dL was associated with increased risk for major adverse cardiovascular events (MACE) during a median follow-up of 2.7 years. After categorizing the patients according to the cutoff values of 85 mg/dL for LDL-C and the median for CD14++CD16+ monocytes, low LDL-C-associated MACE risk was only observed in those with high CD14++CD16+ monocyte counts (low LDL-C/high CD14++CD16+ monocytes vs. low LDL-C/low CD14++CD16+ monocytes: hazard ratio 5.38, 95% confidence interval 1.52 to 19.06, P = 0.009). This work provided the proof-of-principle evidence indicating a role of CD14++CD16+ monocytes in risk stratification of STEMI patients presenting with low LDL-C level. Graphical abstract.

Inflammation has been suggested to play a key role in the pathogenesis of atrial fibrillation (AF). Our hypothesis was that this inflammation, mediated by intermediate monocytes and toll-like receptor 4 (TLR4), causes the formation and expansion of low-voltage zones (LVZs). Prior to ablation, the monocyte subsets of 78 AF patients and TLR4 expression of 66 AF patients were analyzed via a flow cytometric analysis. Based on the CD14/CD16 expression, the monocytes were divided into three subsets: classical, intermediate, and non-classical. At the beginning of the ablation session, voltage mapping was performed. LVZs were defined as all bipolar electrogram amplitudes of < 0.5 mV. Correlations between the flow cytometric analysis results and presence of LVZs, as well as the total area of the LVZ, were examined. Patients with LVZs clearly had a higher proportion of intermediate monocytes (10.0 ± 3.6% vs. 7.2 ± 2.7%, p < 0.001) than those without LVZs. TLR4 was much more frequently expressed in the intermediate monocytes than other two monocyte subsets (p < 0.001). Moreover, the TLR4 expression level in intermediate monocytes correlated positively with the total area of the LVZs (r = 0.267, p = 0.030), especially in patients with paroxysmal AF (r = 0.365, p = 0.015). The intermediate monocytes and TLR4 expression positively correlated with LVZs in AF patients.

Immune complexes (ICs) in blood are efficiently removed mainly by liver reticuloendothelial systems consisting of sinusoidal endothelial cells and Kupffer cells expressing FcγR. The bone marrow (BM) also has sinusoidal vasculatures, and sinusoidal BM endothelial cells (BMECs) bear unique function, including hematopoietic niches and traffic regulation of hematopoietic cells. In this study, we found that sinusoidal BMECs express FcγRIIb2, which is markedly increased in anemic conditions or by the administration of erythropoietin (Epo) in healthy mice. BMECs expressed Epo receptor (EpoR), and the Epo-induced increase in FcγRIIb2 expression was abolished in Epor -/- :: HG1-Epor transgenic mice, which lack EpoR in BMECs except for BM erythroblasts, suggesting the effect was directly mediated via EpoR on BMECs. Further, although BMECs hardly captured i.v.-injected soluble ICs in healthy mice, Epo administration induced a remarkable increase in the uptake of ICs in a FcγRIIb-dependent manner. Enhancement of the IC incorporation capacity by Epo was also observed in cultured BMECs in vitro, suggesting the direct effect of Epo on BMECs. Moreover, we found that i.v.-injected ICs in Epo-treated mice were more rapidly removed from the circulation than in PBS-treated mice. These results reveal a novel function of BMECs to efficiently remove circulating blood-borne ICs in an FcγRIIb2-mediated manner., (Copyright © 2020 by The American Association of Immunologists, Inc.)

To investigate the role of classical (CLM, CD14 ++ CD16 - ), intermediate (INTM, CD14 ++ CD16 + ), and non-classical (Non-CLM, CD14 + CD16 ++ ) monocytes in scar formation after ST-elevation myocardial infarction (STEMI), evaluated with cardiac magnetic resonance (CMR). One hundred two patients with a first STEMI had serial blood analyses after 1, 3, and 7 days. A CMR was performed at 7 days and 6 months, depicting scar core (CO), border zone (BZ), and the presence of BZ channels. CLM and INTM levels progressively decreased, correlated with the scar mass, CO, and BZ at 7 days and 6 months (p < 0.05), and inversely with left ventricular ejection fraction (LVEF, p < 0.01). Non-CLM levels gradually increased, correlated with BZ mass and the presence of BZ channels at 7 days and 6 months (p < 0.001).CLM and INTM are associated with infarct size and inversely with LVEF, whereas Non-CLM are associated with BZ mass and the presence of potentially arrhythmogenic substrate.

Purpose: Human epidermal growth factor receptor 2 (HER2) is overexpressed in 15-20% of patients with breast cancer. HER2 overexpression is the result of a genetic alteration and this marker is associated with poor clinical outcomes. HER2-targeted therapy can significantly improve the prognosis of patients with either early or advanced HER2-positive breast cancer. One such therapy is the antibody drug conjugate (ADC) trastuzumab emtansine (T-DM1), a combination of trastuzumab and the cytotoxic antimicrotubule agent DM1. After T-DM1 binds HER2, DM1 is subsequently released into the cell. T-DM1 is generally well tolerated and has a relatively low incidence of adverse events. However, there are clinical concerns regarding T-DM1-induced high-grade thrombocytopenia., Methods: Here, we summarize the incidence of thrombocytopenia from several clinical trials and review experimental studies to explore the causes for T-DM1-induced thrombocytopenia. Progress in several other ADCs targeting HER2-positive breast cancer was also reviewed., Conclusions: We conclude that T-DM1 uptake by megakaryocytes occurs through either Fcγ receptor binding or through pinocytosis, and we suggest several methods through which these processes could be interrupted to potentially improve the clinical safety of T-DM1. More generally, we recommend that toxicity should be carefully addressed during the development of ADCs., (Copyright © 2020 The Author(s). Published by Elsevier Masson SAS.. All rights reserved.)

Background: Neonatal sepsis (NS) is a very critical medical situation associated with high morbidities and mortalities. There is an utmost need for a new tool helping in early diagnosis and proper management of sepsis neonates. Neutrophil CD64 (nCD64) shows a very promising value in this concerning issue., Aim: Evaluate the diagnostic, monitoring, and prognostic performances of nCD64 and highly sensitive CRP (hs-CRP) in NS as well as the possible best panel of biomarkers that can achieve the most desirable results., Methods: Patients were enrolled from three neonatal intensive care units (NICUs) ( n = 121 patients) and classified according to their initial sepsis evaluation into three groups: disease control group ( n = 30), proven sepsis group ( n = 17), and clinical sepsis group ( n = 74). Laboratory evaluation included hs-CRP, complete blood count (CBC), and blood culture in addition to nCD64 (done by flow cytometry technique). Besides the diagnostic evaluations, follow-up evaluations were done for 40 patients after five days from the first time; patients were reclassified according to their outcome into the improved sepsis neonates' group ( n = 26) and sepsis neonates without improvement ( n = 14)., Results: Significant increase in nCD64 and hs-CRP results were present in sepsis groups compared to the disease controls ( P < 0.001); nCD64 at 43% cutoff value could detect the presence of sepsis with 85.6% sensitivity and 93% specificity. Additionally, delta change percentage (dC%) between improved sepsis neonates and sepsis neonates without improvement showed a significant difference in the levels of both nCD64 ( P < 0.001) and hs-CRP ( P = 0.001)., Conclusion: Besides the promising diagnostic performance documented by nCD64 which is higher than the other laboratory sepsis biomarkers used routinely in NICUs, nCD64 has a valuable role in sepsis patients' monitoring and prognostic evaluation. hs-CRP was moderate in its diagnostic and monitoring results being less than that achieved by nCD64. Combined measurement of nCD64% and hs-CRP gives better diagnostic and monitoring performance than that achieved by any of them alone., Competing Interests: No conflict of interest was declared., (Copyright © 2020 Heba E. Hashem et al.)

Head and neck squamous cell carcinomas (HNSCC) are highly immune suppressive and aggressive malignancies. As part of the tumor microenvironment, exosomes contribute to this immune suppression. The Fc receptor CD16 is widely expressed on monocytes, neutrophils, and natural killer (NK) cells and is involved in antibody-dependent cell-mediated cytotoxicity (ADCC). Here, surface levels of CD16 on total exosomes and tumor-derived exosomes (TEX) from plasma of HNSCC patients were analyzed regarding their potential as liquid biomarkers for disease stage. Exosomes were isolated from plasma using mini size exclusion chromatography. TEX were enriched by immune affinity capture with CD44v3 antibodies. On-bead flow cytometry was used to measure CD16 levels on total exosomes and TEX. The results were correlated with clinicopathological parameters. Total exosomes from HNSCC patients had significantly higher CD16 levels compared to TEX. Further, CD16 surface levels of total exosomes, but not TEX, correlated with clinicopathological parameters. Patients with advanced tumor stages T3/4 and Union for International Cancer Control (UICC) stages III/IV had significantly higher CD16 levels on total exosomes compared to patients with early tumor stages T1/2 and UICC stages I/II, respectively. Overall, CD16 positive exosomes have the potential as liquid biomarkers for HNSCC tumor stage and aggressiveness., Competing Interests: The authors declare no conflict of interest.

Although AML-M3 (APL) and HLA-DR negative non-APL are characterized by negative HLA-DR antigen, they are different entities with similar morphology in some cases. The aim of this study is the precise, differential diagnosis of APL from HLA-DR negative non-APL by flow cytometry to narrow the diagnosis window. Bone marrow or blood samples of 580 AML patients were analyzed, and flow cytometry and molecular analysis were performed for the diagnosis of blood disorders. In 105 HLA-DR negative AML patients, expression of HLA-DR, CD33, CD117, CD11b, CD64, CD34, CD9 and myeloperoxidase staining pattern were evaluated. Fifty-six patients were diagnosed with APL, and 49 patients were diagnosed with HLA-DR negative non-APL. The APL blasts expressed CD33, CD117, CD64, and CD9 in 100%, 80.3%, 94.6%, and 100% of the cases, respectively. HLA-DR negative non-APL blasts expressed CD33, CD117, CD64 and CD9 in 75.5%, 59.1%, 32.6%, and 73.4% of the cases, respectively. APL cells were negative for HLA-DR, CD11b, and CD34 in 96.4%, 94.6%, and 91.0%, respectively. Blasts in HLA-DR negative non M3-AML were negative for CD11b, CD117, and CD34 in 77.5%, 40.9%, and 22.4%, respectively. We also investigated myeloperoxidase (MPO) staining pattern and found strong diffuse reaction in APL cells while HLA-DR negative non-APL cells showed focal positive reaction. In all of the APL patients, except for one, PML/RARA translocation was positive, and in another case with HLA-DR negative non-APL, PML/RARA and other translocations were not detected. The six-panel combination profile rapidly and specifically identifies APL from other HLA-DR negative AML.

Aim: In an Emergency Department (ED), the etiological identification of infected subjects is essential. 13 infection-related biomarkers were assessed using a new flow cytometry procedure. Materials & methods: If subjects presented with febrile symptoms at the ED, 13 biomarkers' levels, including CD64 on neutrophils (nCD64) and CD169 on monocytes (mCD169), were tested and compared with clinical records. Results: Among 50 subjects, 78% had bacterial infections and 8% had viral infections. nCD64 showed 82% sensitivity and 91% specificity for identifying subjects with bacterial infections. mCD169, HLA-ABC ratio and HLA-DR on monocytes had high values in subjects with viral infections. Conclusion: Biomarkers showed promising performances to improve the ED's infectious stratification.

Soluble forms of the low-affinity immunoglobulin receptor FcγRIIa (sFcγRIIa) lacking the cytoplasmic tail have been reported in plasma however the mechanism and functional consequences are unknown. This study aimed to evaluate mechanisms of FcγRIIa release compared to GPVI release from platelets, and examine whether genetic polymorphisms at positions 27 and 131 within FcγRIIa correlate with platelet FcγRIIa stability and function. Enzyme-linked immunosorbent assays (ELISAs) were used to measure plasma sFcγRIIa and sGPVI levels. FcγRIIa genotype at positions 27 and 131 was evaluated. sFcγRIIa levels were not significantly different between non-131HH and 131HH but were significantly lower in 27W than non-27W. Treatment of platelets with aggregated immunoglobulin (Ig) G induced release of FcγRIIa and GPVI, but only sGPVI release was statistically significant, required functional FcγRIIa, and was blocked by inhibitors of signaling pathways and metalloproteinases. This indicated that sFcγRIIa was not released from platelets by metalloproteolysis. sFcγRIIa levels were not correlated with sGPVI levels in healthy individuals however levels of sFcγRIIa and sGPVI in plasma from patients with rheumatoid arthritis (RA) were significantly elevated above levels found in healthy individuals. Elevated level of sFcγRIIa in RA patients may reflect active immune-based arthritis and be predictive of active inflammation.

Objective: To determine diagnostic value of neutrophil CD64 index (iCD64n) in the diagnosis of postoperative infectious complications after colorectal resections., Material and Methods: Seventy-three patients underwent colorectal surgery for the period from January to December 2018. These patients were included into a single-center study. Peripheral blood samples were taken on 3 and 6 postoperative days (POD) to check iCD64n level. We analyzed incidence of postoperative infectious complications, sensitivity (Se) and specificity (Sp) of postoperative iCD64n level on the 3 rd and 6 th POD., Results: Postoperative infectious complications developed in 10 (13.7%) patients. Median iCD64n was significantly higher ( p =0.0017 for POD 3; p =0.018 for POD 6) in patients with infectious complications (1.6 on POD 3; 1.3 on POD 6) compared to those without complications (1.1 on POD 3; 0.9 on POD 6). Area under curve (AUC) on the 3 rd POD was 0.8 with the cut-off value of 1.4, Se - 70%, Sp - 93.7% ( p =0.002). On the 6 th POD, AUC was 0.91 with cut-off value of 1.23, Se - 80%, Sp - 93.7% ( p <0.001)., Conclusion: Neutrophil CD64 index is a valuable predictor for the diagnosis of postoperative infectious complications after colorectal resections. It is a useful tool to ensure a safe early discharge., The study is registered on the website «clinictrials.gov» (registration number NCT03559335).

Purpose: To characterize neutrophils in obstructive airway disease by measuring their surface adhesion molecules and oxidative burst along with characterizing them into different subsets as per their adhesion molecule expression., Patients and Methods: Peripheral blood from adults with COPD (n=17), asthma (n=20), and healthy participants (n=19) was examined for expression of CD16, CD62L, CD11b, CD11c, and CD54, and analyzed by flow cytometry. For oxidative burst and CD62L shedding analysis, CD16 and CD62L stained leukocytes were loaded with Dihydrorhodamine-123 (DHR-123) and stimulated with N-Formylmethionine-leucyl-phenylalanine (fMLF). Neutrophil subsets were characterized based on CD16 and CD62L expression. Marker surface expression was recorded on CD16 + neutrophils as median fluorescence intensity (MFI)., Results: Neutrophil surface expression of CD62L was significantly reduced in COPD (median (IQR) MFI: 1156 (904, 1365)) compared with asthma (1865 (1157, 2408)) and healthy controls (2079 (1054, 2960)); p=0.028. COPD neutrophils also demonstrated a significant reduction in CD62L expression with and without fMLF stimulation. Asthma participants had a significantly increased proportion and number of CD62L bright /CD16 dim neutrophils (median: 5.4% and 0.14 × 10 9 /L, respectively), in comparison with healthy (3.54% and 0.12 × 10 9 /L, respectively); p<0.017., Conclusion: Reduced CD62L expression suggests blood neutrophils have undergone priming in COPD but not in asthma, which may be the result of systemic inflammation. The increased shedding of CD62L receptor by COPD blood neutrophils suggests a high sensitivity for activation., Competing Interests: Dr Michael Fricker reports grants from NHMRC, grants from Thoracic Society of Australia and New Zealand, during the conduct of the study. The authors report no other conflicts of interest in this work., (© 2019 Lokwani et al.)

Background: Monocytes in human peripheral blood are heterogeneous and can be divided into 2 groups, inflammatory and pro-inflammatory, according to the differential expression of CD14 and CD16. Pro-inflammatory monocytes (CD14 + CD16 + ) seem to contribute to the development of coronary artery disease. This study aimed to investigate the involvement of specific human peripheral monocyte subsets in the development of future coronary events.Methods and Results:We enrolled 271 patients who were suspected to have either stable angina pectoris or silent myocardial ischemia and underwent coronary angiography (CAG). Two monocyte subsets (CD14 + CD16 - and CD14 + CD16 + ) were measured by flow cytometry. Patients who did not undergo coronary artery revascularization at initial CAG were followed as the medical therapy group, which included 136 patients among whom 15 had future coronary events. The frequency of CD14 + CD16 + monocytes was significantly higher in patients who had future coronary events than in those who did not (P<0.01). Furthermore, the frequencies of CD14 + CD16 + monocyte were not significantly different between patients who had future coronary events and those who underwent coronary revascularization at initial CAG (P<0.33). Multivariate analysis revealed that the frequency of CD14 + CD16 + monocytes was an independent predictor for future coronary events (P<0.01)., Conclusions: An increase in the abundance of human peripheral pro-inflammatory monocytes is related to the development of future coronary events.

BACKGROUND The aim of this study was to determine the relationship between the expression of CD35 and CD64 from white blood cells (neutrophil, monocytes, and lymphocytes) and acute infectious diseases in children. MATERIAL AND METHODS The blood samples were collected from 104 children with infections (42 viral infections and 62 bacterial infections). Blood samples were stained with CD45-PC5, CD35-FITC, and CD64-PE, and the fluorescence intensities were measured by flow cytometer, and then the ratio of CD35 to CD64 was calculated. RESULTS The ratio of CD64/CD35 on neutrophils (NCD35/NCD64) was significantly different between the bacterial group, the virus group, and the healthy control group. According to receiver operating characteristic (ROC) analysis, a cutoff value of 7.256 (sensitivity: 90.0%, specificity: 93.7%) was determined for the NCD35/NCD64 ratio. CONCLUSIONS This study shows that NCD35/NCD64 is helpful in the differential diagnosis of acute viral infection and bacterial infection in children.

Cytomegalovirus (CMV) infection is a major complication after allogeneic hematopoietic stem cell transplantation but is suggested to exert a strong antileukemia effect in part due to alterations in the composition of natural killer (NK) cells. We evaluated the impact of early CMV reactivation and changes in NK cell subset recovery on relapse rate and survival after haploidentical stem cell transplantation (haploSCT) for acute leukemia. Fifty patients with acute leukemia who received haploSCT were analyzed. Expression of T cells and specific receptors (NKG2A, NKG2D, DNAM1, and CD57) on circulating NK cells (CD56 bright CD16 dim/ - or CD56 dim CD16 + cells) was serially measured using multiparametric flow cytometry. CMV reactivation during the first 100 days was observed in 41 patients (82%) at a median of 23 days after haploSCT. The incidence of acute graft-versus-host disease (GVHD) and chronic GVHD tended to be higher in patients with CMV reactivation, although this difference was not statistically significant. Multivariate analysis showed that CMV reactivation (P = .011) and a dose of infused T cells > 3.2 × 10 8 /kg (P = .027) were independent predictors of a reduced relapse risk and only CMV reactivation (P = .029) was an independent predictor of improved leukemia-free survival. CD56 bright CD16 dim/- DNAM1 + NK cell counts increased from day 30 to 90 in patients with CMV reactivation but decreased after day 30 in patients without CMV reactivation. An increase in CD56 bright CD16 dim/- DNAM1 + NK cells was not associated with the occurrence of chronic GVHD but was associated with a reduced cumulative relapse rate (16.4% versus 58.0%, P = .019). Multivariate analysis indicates that an increase in the CD56 bright CD16 dim/- DNAM1 + NK cell count was an independent predictor of reduced relapse risk. Our study demonstrates a significant correlation between low relapse rates and CMV reactivation as well as the recovery of CD56 bright CD16 dim/- DNAM1 + NK cells, providing valuable information for understanding the plausible immunologic mechanism of the graft-versus-leukemia effect., (Copyright © 2019 American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

The aim of this study was to evaluate the changes in the three subsets of monocyte (classical, intermediate, and non-classical) and the expression of human leukocyte antigen-DR (HLA-DR) on monocyte subsets during MP pneumonia in children. Monocyte subsets were analyzed in the peripheral blood of healthy volunteers and MP pneumonia patients at the stages of admission and remission after clinical therapy. They were defined as classical (CD14 + CD16 - ), intermediate (CD14 bright CD16 + ), and non-classical (CD14 dim CD16 + ) using flow cytometry. Furthermore, three subsets of monocyte were analyzed for the expression of HLA-DR. Patients with MP pneumonia at admission had a higher proportion of intermediate and non-classical monocytes than healthy subjects (all P < 0.05). The proportion of intermediate subset and non-classical subset was lower in MP pneumonia patients at remission than at admission (all P < 0.05). In comparison with the other monocyte subsets, intermediate subset showed a significantly higher percentage of HLA-DR in MP pneumonia patients at admission (P < 0.05). Further analysis revealed that the expression of HLA-DR on intermediate subset was lower in severe patients than in non-severe patients (P < 0.05).Our data has shown for the first time that MP pneumonia is associated with the increased proportion of non-classical and intermediate monocytes, indicating the involvement of monocyte-related mechanisms in the pathogenesis of this disease. Additionally, the decreased expression of HLA-DR on CD14 bright CD16 + subset may be a potential indicator of the severity of MP pneumonia.

Objectives: The clinical significance of low-level viraemia (LLV) during antiretroviral therapy (ART) is debated. We retrospectively investigated longitudinal levels of plasma markers associated with inflammation, altered coagulation and cardiovascular disease in Swedish HIV-positive adults in relation to LLV or permanent virological suppression during long-term ART., Methods: Plasma levels of C-reactive protein (CRP), D-dimer, vascular cell adhesion molecule 1 (VCAM-1), suppression of tumorigenicity 2 (ST2), growth differentiation factor 15 (GDF-15), soluble CD14 (sCD14), soluble CD163 (sCD163), interferon-γ-induced protein 10 (IP-10) and β-2-microglobulin were measured in 34 individuals with LLV (viral load 50-999 HIV-1 RNA copies/mL) and in matched controls with persistent virological suppression. Biomarker levels were analysed in samples obtained during episodes of LLV and follow-up samples obtained 1 year later (with similar timing for controls). All biomarkers were analysed using an independent sample t-test and analysis of covariance (ANCOVA) after logarithmic transformation. Log-rank analysis was applied for markers with concentration values out of range., Results: Compared with controls, patients with LLV had significantly higher levels of GDF-15 [geometric mean 3416 (95% confidence interval (CI) 804-14 516) pg/mL versus 2002 (95% CI 355-11 295) pg/mL in controls; P = 0.026] and D-dimer [mean 1114 (95% CI 125-9917) ng/mL versus 756 (95% CI 157-3626) ng/mL; P = 0.038] after adjustment for age, CD4 count nadir and type of ART. In the unadjusted t-test, only GDF-15 was significantly higher and in the log-rank test, both GDF-15 and D-dimer were significantly elevated. No significant differences were observed for the other biomarkers analysed., Conclusions: Although levels of inflammation markers were similar in ART recipients with and without LLV, persons with LLV had significantly higher levels of GDF-15 and D-dimer. These findings suggest a potential link between LLV and cardiovascular outcomes., (© 2019 British HIV Association.)

Sepsis is a major cause of death in hospitalised patients accounting for mortality rates as high as 60% and, hence, is called 'a hidden public health disaster'. Sepsis is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. Sepsis is not a disease but is a clinical syndrome, where the initial features are nonspecific resulting in delayed diagnosis. Lack of specific laboratory tests to diagnose the syndrome adds to the diagnostic confusion. Failure to identify sepsis in the early stages itself delays effective treatment resulting in high morbidity and mortality. Various biomarkers and newer laboratory tests help to address these issues. However, to date there is no ideal test to diagnose sepsis. The most commonly used markers are C-reactive protein (CRP) and procalcitonin (PCT). There are around 180 biomarkers reported to be useful in sepsis. In addition to CRP and PCT, various emerging laboratory markers, such as like serum amyloid A, soluble triggering receptor expressed on myeloid cell-1, mannan and antimannan antibodies, and interferon γ inducible protein-10 etc., have been reviewed and their clinical usefulness discussed in this paper., Competing Interests: No conflict of interests declared

Background: Although early identification of sepsis improves outcome, prompt and correct diagnostic remains often challenging. The expression of the high affinity immunoglobulin-Fc fragment receptor I CD64 on neutrophils is upregulated during acute inflammation. We here aimed at determining the usefulness of its rapid measurement in diagnosing sepsis., Methods: Seventy-two consecutive patients were enrolled upon admission to Intensive Care Unit within a two-month period. Sequential determination of serum C-reactive protein (CRP) and procalcitonin (PCT) concentrations was obtained. The neutrophil CD64 index was measured using Accellix-CD64® device, an in vitro diagnosis system allowing for an automatic and standardized measure., Results: Serum concentrations of CRP and PCT as well as the neutrophil CD64 index were higher in septic patients compared to all others (P<0.05 for the three markers). Only CD64 index was an independent predictor of sepsis, though with modest sensitivity and specificity (78% and 70%, respectively). Repeat determination of CD64 index at day 2 correctly classified 85% of patients., Conclusions: This prospective study demonstrates the moderate performance of the neutrophil CD64 index, assessed through the Accellix-CD64® device, in diagnosing sepsis in the critically ill patient. However, repeat measurements improve its accuracy and may help to predict ICU-acquired infections.

Background: Sepsis is the leading cause of morbidity and mortality in newborns. CD64 combined with c-reactive protein (CRP) could improve the sensitivity and specificity of neonatal sepsis diagnosis, but the results were still controversial. Therefore, this meta-analysis was conducted to clarify the importance of CD64 combined with CRP in the diagnosis of neonatal sepsis., Methods: The researches published as of December 24, 2018 were comprehensively searched in PubMed, Embase (included Embase and Medline), the Cochrane Library and Web of Science. Totally, 8 articles were included, involving 1114 objects. Statistical calculations were performed using Stata14.0 and Review Manager 5.3., Results: The diagnostic accuracy of all included studies was pooled as follows: sensitivity, 0.95 (95% CI: 0.86-0.98); specificity, 0.86 (95% CI: 0.74-0.93); positive likelihood ratio (PLR), 6.8 (95% CI: 3.50-13.20); negative likelihood ratio (NLR), 0.06 (95% CI: 0.02-0.18); diagnostic odds ratio (DOR), 118.0 (95% CI: 25.00-549.00), and the area under the curve (AUC) was 0.96 (95% CI: 0.94-0.97). It was found that heterogeneity was not caused by threshold effect (P = 0.16), but the results of sensitivity (I 2  = 87.57%) and specificity (I 2  = 89.07%) analyses indicated significant heterogeneity between studies., Conclusions: The combined application of CD64 and CRP improved the accuracy of neonatal sepsis diagnosis., (Copyright © 2019. Published by Elsevier Inc.)

Aim: Patients with ascites are at a higher risk for associated of on top bacterial infections with subsequent life-threatening complications. We aimed to evaluate CD64, calprotectin, and microRNA-155 (miR-155) levels as diagnostic markers of spontaneous bacterial peritonitis (SBP) and the effect of using more than one use on the same spot over their diagnostic efficiency., Patients and Methods: An observational comparative study included 103 patients with ascites admitted to the Tropical Medicine Department, Mansoura University Hospital, Egypt, divided into two groups: case group (64 patients) with ascites with SBP and control group (39 patients) with decompensated cirrhotic non-SBP ascites. Twenty milliliters of ascetic fluid was obtained from all participants for bacterial culture, and assessment of calprotectin and miR-155, in addition to 2 ml blood for the CD64 marker expression assay by a flowcytometer., Results: The sensitivity and specificity of CD64 expression assay were 95.3 and 92.3%, respectively, area under the curve (AUC)=0.93, whereas those of ascetic fluid calprotectin and miR-155 were 87.5 and 82.1%, AUC=0.90 and 95.3 and 97.4%, with AUC of 0.95. Combined blood CD64 and ascetic fluid calprotectin had a diagnostic accuracy of 0.988 for blood CD64 and ascetic fluid miR-155, AUC=0.991, and that for ascetic fluid calprotectin and miR-155 was 0.988. On using the three studied markers together, the diagnostic accuracy was the best recorded, AUC=0.994. P values were less than 0.001., Conclusion: CD64, calprotectin, and miR-155 were good diagnostic markers of SBP and on using this combination, greater efficiency in diagnosis was achieved.

Sepsis is a life-threatening disease that affects millions of people every year. Rapid detection of sepsis assists clinicians to initiate timely antibiotic therapy and to reduce mortality. At the same time, accurate point-of-care detection is needed to reduce unnecessary use of antibiotics. One of the principal challenges in sepsis diagnosis is that many sepsis cases do not result in positive blood cultures. These so-called culture-negative cases present a significant health threat. In this work, we present a microfluidic cells separation system for the detection of sepsis in both culture-positive and culture-negative cases. Leukocytes were captured in several affinity separation zones of a microchip based on CD64, CD69, and CD25 expression. To validate this assay 40 septic patients and 10 healthy volunteers were enrolled in this study. Septic patients were divided into culture-positive (n = 12) and culture-negative cases (n = 21). CD64  +  cell capture demonstrated excellent accuracy for sepsis detection with an area under the receiver operating characteristic curves (AUC) of 0.962. A combined panel of CD64  +  and CD69  +  cell counts was constructed, and the new panel outperformed each of these two biomarkers alone with the AUC of 0.978. Our affinity microfluidic devices were validated by conventional flow cytometry analysis. Results showed that the cell capture number of specific affinity region increased along with the increase of its corresponding antigen expression. This clinical validation confirms that CD64 and CD69 cell separations are a powerful sepsis assay with the potential for point-of-care analysis in culture-positive and culture-negative cases., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Intermediate (CD14 ++ CD16 + ) monocytes have important pro-inflammatory and atherogenic features and are increased in patients with chronic kidney disease (CKD). The present study aims to elucidate the role of the uremic milieu and of platelet activation in monocyte differentiation. Monocyte subtypes were analyzed in CKD patients (n = 193) and healthy controls (n = 27). Blood from healthy controls (Ctrl; n = 8) and hemodialysis patients (HD; n = 8) was centrifuged, and plasma (pl) was exchanged between Ctrl and HD (Ctrlcells/HDpl and HDcells/Ctrlpl) or reconstituted as original (Ctrlsham and HDsham) and incubated for 24 h (T24). Monocyte differentiation and platelet aggregation to monocytes (MPA) was assessed by flow cytometry. Especially, a higher proportion of CD14 ++ CD16 + monocytes was found in hemodialysis (HD) patients (p < 0.01). In plasma exchange experiments, Ctrl cells/HD pl T24 showed an increased percentage of CD14 ++ CD16 + monocytes versus Ctrl sham (33.7% ± 15 vs. 15.7% ± 9.6; P < 0.005), comparable to the level of CD14 ++ CD16 + monocytes in the HD sham condition. The percentage of CD14 ++ CD16 + monocytes was lowered by suspending HD cells in Ctrl pl (18.4% ± 7.8 vs. 36.7% ± 15 in HD sham; P < 0.005) reaching the level of the Ctrl sham condition (15.7% ± 9.6). A mixture of uremic sulfates increased CD14 ++ CD16 + monocytes compared to control (19.8 ± 9.6% vs. 15.8 ± 10.9%; P < 0.05), paralleled by a rise MPA. Blocking MPA by abciximab, a potential therapeutic strategy, or anti-CD62P did not inhibit differentiation towards the CD14 ++ CD16 + monocytes. In conclusion, in the present cohort, CD14 ++ CD16 + monocytes are especially increased in HD patients and this can at least in part be attributed to the presence of the uremic milieu, with uremic sulfates inducing a reversible shift towards pro-inflammatory CD14 ++ CD16 + monocytes.

Sepsis, a life-threatening syndrome that contributes to millions of deaths annually worldwide, represents a moral and economic burden to the healthcare system. Although no single, or even a combination of biomarkers has been validated for the diagnosis of sepsis, multiple studies have shown the high specificity of CD64 expression on neutrophils (nCD64) to sepsis. The analysis of elevated nCD64 in the first 2-6 hours after infection during the pro-inflammatory stage could significantly contribute to early sepsis diagnosis. Therefore, a rapid and automated device to periodically measure nCD64 expression at the point-of-care (POC) could lead to timely medical intervention and reduced mortality rates. Current accepted technologies for measuring nCD64 expression, such as flow cytometry, require manual sample preparation and long incubation times. For POC applications, however, the technology should be able to measure nCD64 expression with little to no sample preparation. In this paper, we demonstrate a smartphone-imaged microfluidic biochip for detecting nCD64 expression in under 50 min. In our assay, first unprocessed whole blood is injected into a capture chamber to immunologically capture nCD64 along a staggered array of pillars, which were previously functionalized with an antibody against CD64. Then, an image of the capture channel is taken using a smartphone-based microscope. This image is used to measure the cumulative fraction of captured cells (γ) as a function of length in the channel. During the image analysis, a statistical model is fitted to γ in order to extract the probability of capture of neutrophils per collision with a pillar (ε). The fitting shows a strong correlation with nCD64 expression measured using flow cytometry (R 2 = 0.82). Finally, the applicability of the device to sepsis was demonstrated by analyzing nCD64 from 8 patients (37 blood samples analyzed) along the time they were admitted to the hospital. Results from this analysis, obtained using the smartphone-imaged microfluidic biochip were compared with flow cytometry. Again, a correlation coefficient R 2 = 0.82 (slope = 0.99) was obtained demonstrating a good linear correlation between the two techniques. Deployment of this technology in ICU could significantly enhance patient care worldwide.

Background: Graves' disease (GD) is a common autoimmune disease characterized by genetic and environmental factors. Fcγ receptors (FcγRs) are involved in several autoimmune disorders through recognizing immunoglobulin (Ig) G antibodies and mediating immune response. The study on the expression of FcγRs in GD patients is scarce. The purpose of this study was to evaluate the expression of three different types of FcγRs in patients with active and remissive GD., Methods: Blood samples of patients and healthy subjects were collected to analyze the percentage of FcγRI (CD64), FcγRII (CD32), and FcγRIII (CD16) on peripheral blood mononuclear cells (PBMCs) and monocytes by flow cytometry and Western blotting. CD32 isotypes were also examined in cases and controls by real-time PCR., Results: The cell percentages expressed CD32 and protein expressions of CD32 on PBMCs, and monocytes from patients with active GD were significantly reduced compared to controls and patients with remissive GD. In particular, the expression of CD32B on PBMC was also decreased in active GD patients. However, the cell percentages expressed CD16 and CD64 from PBMCs and monocytes were comparable between three groups. Besides, the percentages of CD14 + CD32 + cells were negatively correlated with TRAb titers in active GD patients (r = -0.5825, P ＜ 0.001)., Conclusion: These results suggested that CD32 may act as a novel marker for active GDs. The expression of monocytic CD32, in particular CD32B, in GD patients might play a crucial role in maintaining FcγRs function and be a therapeutic target in GD patients., (© 2019 The Authors. Journal of Clinical Laboratory Analysis Published by Wiley Periodicals, Inc.)

Background: Flow cytometry is a powerful analytical technique. However, it requires time-consuming, multi-step sample procedure. A new protocol was developed to perform extracellular staining and red blood cell lysis in one step, using dry antibodies. Common markers of white blood cells as well as sepsis biomarkers were tested as a model for modulated antigen expression., Methods: Peripheral blood was stained using the reference and the one-step methods. Recruitment and staining of CD3-, CD4-, CD8-, CD14-, and CD15-positive cells were analyzed. Then, protocol modifications were tested for optimization. Finally, the one-step method was evaluated on subjects in septic conditions, by measuring expressions of CD64 and of HLA-DR., Results: No statistical differences were observed between methods when comparing the proportions of cells. The procedure was optimized by decreasing blood volume from 100 μL to 5 μL, lysis from 1 mL to 500 μL, and time from 30 to 15 min. In the blood samples from septic subjects, an increase of CD64 on neutrophils and a decrease of HLA-DR on monocytes were observed., Conclusions: The one-step method, described here-in, enables an accurate, streamlined flow cytometry sample preparation protocol. The simplified phenotyping procedure reduces training requirements and could help overcome logistic constraints in many flow cytometry applications., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: Neonatal pneumonia (NP) is one of the major causes of neonatal death. Current NP diagnosis depends on a detailed history, physical examination, and radiographic and laboratory findings. There is no specific biomarker or diagnostic indicator of NP., Methods: In this study, we tried to find a reliable biomarker for quick NP diagnosis by collecting peripheral blood from neonates with NP and transient tachypnea of the newborn (TTN), and subsequently tested the expression of CD64 on white blood cells using flow cytometry. The cellularity of each blood cell population was also quantified. Furthermore, procalcitonin (PCT) and C-reactive protein (CRP) levels were evaluated in the blood sera., Results: We found that NP patients had moderately increased polymorphonuclear cells (PMNs), as well as elevated PCT and CRP levels in the blood sera. Importantly, the expression of CD64 on PMNs was profoundly increased in NP patients but not TTN patients. The receiver operating characteristic (ROC) curve of PMN CD64 index suggests that PMN CD64 index is sensitive and specific for NP diagnosis., Conclusions: Our study reveals that PMN CD64 could be a fast and reliable biomarker for NP diagnosis.