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Thrombosis, a prevalent condition, can provoke severe health issues like acute coronary syndrome (ACS), deep vein thrombosis (DVT), and pulmonary embolism (PE). The rising incidence of these diseases annually significantly impacts patient wellbeing and poses a substantial burden on healthcare systems. Recombinant neorudin is a developing anticoagulant drug for thrombotic diseases whose phase I clinical trials has been completed. The distribution pattern of it and its active metabolite, hirudin, in thrombi, blood surrounding the thrombus and peripheral blood remains uncertain. This study explored their distribution using a rat arteriovenous bypass thrombosis model, revealing higher neorudin levels in blood surrounding the thrombus and elevated hirudin concentrations in thrombus. Recombinant neorudin significantly increased Thrombin Time (TT) in both plasma surrounding the thrombus and peripheral blood, and reduced the wet weight of the thrombus. The results above demonstrated the anticoagulant and antithrombotic efficacy of recombinant neorudin in vivo. Give the distribution pattern of neorudin and hirudin, we hypothesized that neorudin was cleaved at the site of thrombus formation to produce hirudin, leading to the rapid accumulation of hirudin within local thrombi and resulting in a higher concentration inside the thrombus. This insight was crucial for understanding the action mechanisms of anticoagulants in thrombosis management and provided a valuable guidance for therapeutic strategies in treating thrombotic diseases. [ABSTRACT FROM AUTHOR]

Introduction: Endometriosis (EMs) and adenomyosis (AD) are common gynecological diseases that impact women's health, and they share symptoms such as dysmenorrhea, chronic pain, and infertility, which adversely affect women's quality of life. Current diagnostic approaches for EMs and AD involve invasive surgical procedures, and thus, methods of noninvasive differentiation between EMs and AD are needed. This retrospective cohort study introduces a novel, noninvasive classification methodology employing a stacked ensemble machine learning (ML) model that utilizes peripheral blood and coagulation markers to distinguish between EMs and AD. Methods: The study included a total of 558 patients (329 with EMs and 229 with AD), in whom key hematological and coagulation markers were analyzed to identify distinctive profiles. Feature selection was conducted through ML (logistic regression, support vector machine, and K-nearest neighbors) to determine significant hematological markers. Results: Red cell distribution width, mean corpuscular hemoglobin concentration, activated partial thromboplastin time, international normalized ratio, and antithrombin III were proved to be the key distinguishing indexes for disease differentiation. Among all the ML classification models developed, the stacked ensemble model demonstrated superior performance (area under the curve = 0.803, 95% credibility interval = 0.701-0.904). Our findings demonstrate the effectiveness of the stacked ensemble ML model for classifying EMs and AD. Discussion: Integrating biomarkers into this multi-algorithm framework offers a novel approach to noninvasive diagnosis. These results advocate for the application of stacked ensemble ML utilizing cost-effective and readily available peripheral blood and coagulation indicators for the early, rapid, and noninvasive differential diagnosis of EMs and AD, offering a potentially transformative approach for clinical decision-making and personalized treatment strategies. [ABSTRACT FROM AUTHOR]

Simple Summary: Peripheral blood analyses can offer a minimally invasive view into systemic immunity during cancer and can lead to the identification of biomarkers for cancer screening and therapeutic management. While a limited number of studies have reported blood transcriptome in breast cancer (BC) using RNA-seq analysis, our study is the first that aimed to identify potential BC biomarkers by analyzing transcriptome at an isoform level in peripheral blood mononuclear cells (PBMCs) from BC patients and healthy women. Our approach has led to the identification of an isoform of the RASGEF1A gene, the ENST00000374459 transcriptional variant, as a promising blood mRNA biomarker for distinguishing BC and healthy subjects. Additionally, our association analysis with clinicopathological characteristics revealed that lower ENST00000374459 expression in PBMCs of breast cancer patients was associated with higher proliferation and circulating tumor DNA (ctDNA) shedding, thereby linking expression of this isoform in blood immune cells to cancer progression and spreading. Background: Breast cancer (BC) comprises multiple subtypes with distinct molecular features, which differ in their interplay with host immunity, prognosis, and treatment. Non-invasive blood analyses can provide valuable insights into systemic immunity during cancer. The aim of this study was to analyze the expression of transcriptional isoforms in peripheral blood mononuclear cells (PBMCs) from BC patients and healthy women to identify potential BC immune biomarkers. Methods: RNA sequencing and isoform-level bioinformatics were performed on PBMCs from 12 triple-negative and 13 luminal A patients. Isoform expression validation by qRT-PCR and clinicopathological correlations were performed in a larger cohort (156 BC patients and 32 healthy women). Results: Transcriptional analyses showed a significant (p < 0.001) decrease in the ENST00000374459 RASGEF1A isoform in PBMCs of BC compared to healthy subjects, indicating disease-related expression changes. The decrease was associated with higher ctDNA and Ki-67 values. Conclusions: The levels of the RASGEF1A transcriptional isoform ENST00000374459 may have the potential to distinguish between BC and healthy subjects. The downregulation of ENST00000374459 in breast cancer is associated with higher proliferation and ctDNA shedding. Specialized bioinformatics analyses such as isoform analyses hold significant promise in the detection of biomarkers, since standard RNA sequencing analyses may overlook specific transcriptional changes that may be disease-associated and biologically important. [ABSTRACT FROM AUTHOR]

Endometriosis, a debilitating condition, affects one in ten women of reproductive age. Its pathophysiology remains unclear, though deficiencies in immune surveillance are thought to create an environment conducive to the evasion of ectopic endometrial cells from the immune system. Our research explores the immunological impact of endometriosis both locally and systemically, emphasizing natural killer (NK) and T cell subpopulations. We incorporated 62 female patients who underwent laparoscopic surgery; of those, 47 had endometriosis, and 15 were controls. We collected peritoneal fluid (PF) and peripheral blood (PB) samples which were tagged with monoclonal antibodies and subsequently scrutinized using flow cytometry. Our findings revealed significant differences in immunological profiles based on demographic factors and symptomatology. In the endometriosis cohort, there was an increase in PB CD56HiCD16dim and PF CD8+ CD56dimCD16Hi NK cells. CD16+ CD4 T cell levels were significantly lower in the PB of endometriosis patients who smoke. Individuals with more severe disease displayed significantly higher levels of PB CD16+ CD8 T cells, which also increased in those with non-menstrual pelvic pain. Dysmenorrhea severity correlated with a progressive increase in PF CD8+ CD56dimCD16Hi NK cells. These variations in specific lymphocyte subsets, namely, within NK and T cells, suggest potential immunological mechanisms in the evolution and clinical presentation of endometriosis. [ABSTRACT FROM AUTHOR]

Objective: This study assessed the diagnostic value of microRNA-200 (miR-200) expression in peripheral blood-derived extracellular vesicles (EVs) in early-stage non-small cell lung cancer (NSCLC). Methods: This study retrospectively analyzed 100 healthy volunteers (the control group) receiving physical examinations, 168 early-stage NSCLC patients (the NSCLC group), and 128 patients with benign lung nodules (the benign group). The basic and clinical data of participants were obtained, including age, sex, smoking history, carbohydrate antigen 242 (CA242), carcinoembryonic antigen (CEA), carbohydrate antigen 199 (CA199), forced expiratory volume in 1 s, maximal voluntary ventilation, forced vital capacity, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and miR-200 expression. The correlation of miR-200 expression in peripheral blood-derived EVs with CA242, CEA, and CA199 was analyzed, and the diagnostic value of peripheral blood-derived EV miR-200 for early-stage NSCLC was assessed. The risk factors of early-stage NSCLC development were also determined. Results: Age, the percentage of patients with smoking history, CA242, CEA, CA199, IL-6, and TNF-α levels, and miR-200 expression in peripheral blood-derived EVs were significantly higher in the NSCLC group than in the benign and control groups. Lung disease patients with high miR-200 expression in peripheral blood-derived EVs comprised a higher percentage of patients with smoking history and mixed lesions and had higher CA242, CEA, CA199, and TNF-α levels than those with low miR-200 expression in peripheral blood-derived EVs. In lung diseases, miR-200 expression in peripheral blood-derived EVs was significantly and positively correlated with CA242, CEA, and CA199. Peripheral blood-derived EV miR-200 combined with CA242, CEA and CA199 had higher diagnostic value (area under the curve = 0.942) than single detection, along with higher specificity, and high expression of peripheral blood-derived EV miR-200 was an independent risk factor for early-stage NSCLC. Conclusion: Peripheral blood-derived EV miR-200 expression in patients with lung diseases is closely correlated with CA242, CEA, and CA199, and high expression of peripheral blood-derived EV miR-200 is an independent risk factor for early-stage NSCLC and is of high clinical diagnostic value for early-stage NSCLC. [ABSTRACT FROM AUTHOR]

Recurrent pregnancy loss (RPL) is thought to be related to maternal-fetal immune tolerance disorders. Immune monitoring of RPL patients mainly involves two aspects: inflammatory factors and immune cells. However, most observational studies have reported controversial findings. This study aimed to confirm whether abnormal inflammatory factors and immune cells in peripheral blood may lead to RPL, and guide clinical immune monitoring. We demonstrated causality using two-sample Mendelian randomization. Sensitivity analysis, reverse Mendelian randomization and meta-analysis were used to enhance the effectiveness of the results. There was a causal relationship between the level of IL-12 (OR = 1.78, 95% CI = 1.25–2.55; P = 0.00149) and RPL for 41 inflammatory factors. We screened 5 groups of immune cell subtypes that were causally associated with RPL: switched memory B-cell absolute count (OR = 0.66, 95% CI = 0.49–0.87, P = 0.00406), IgD + CD24 + B-cell absolute count (OR = 0.69, 95% CI = 0.53–0.88, P = 0.00319), CD39 + resting CD4 regulatory T-cell %CD4 regulatory T-cell (OR = 0.86, 95% CI = 0.78–0.95, P = 0.00252), activated & resting CD4 regulatory T-cell %CD4 regulatory T-cell (OR = 0.89, 95% CI = 0.82–0.97, P = 0.00938) and CD45 RA + CD28-CD8 + T-cell %CD8 + T-cell (OR = 0.99, 95% CI = 0.98-1.00, P = 0.01231). In terms of inflammatory factors, a causal relationship between IL-12 and RPL in peripheral blood was confirmed. We also identified five immune cell phenotypes that play a protective role. This suggests that there may be protective B cells and CD8 + T-cell subsets in peripheral blood, and the protective effect of Tregs was proved again. Immune monitoring of peripheral blood in patients with RPL seems to be necessary and the foundation for precision medicine. [ABSTRACT FROM AUTHOR]

Purpose: Lung cancer remains a leading cause of cancer-related death and chemotherapy stands as a fundamental component in therapy. Chemotherapy-induced myelosuppression encompasses a spectrum of hematological declines, including not only neutrophils but also lymphocytes, hemoglobin levels and platelets. This retrospective cohort study investigates alterations in peripheral blood lymphocyte subsets. By uncovering these changes, our goal is to refine patient management strategies, ensuring that the benefits of chemotherapy are maximized while minimizing its detrimental effects. Patients and Methods: We retrospectively analyzed 159 lung cancer patients. Patients were categorized as "NT" (n=108, no previous anti-tumor therapy), and "PT" (n=51, prior therapy followed by at least a two-month treatment-free interval). Post-chemotherapy, patients were reassessed and grouped into "EarlyCycle" for those who underwent four or fewer cycles, and "LateCycle" for those who underwent more than four cycles. Results: The study focused on analyzing the percentages of lymphocyte subsets, including T cells (CD4+, CD8+), B cells, and natural killer (NK) cells, across these groups. For T cells, the EarlyCycle group exhibited a significant increase compared to NT (0.7783 vs 0.7271; p=0.0017) and PT (0.7783 vs 0.6804; p=1.6e-05). B cells showed a significant decrease from NT to LateCycle (0.1014 vs 0.0817; p=2.2e-05) and from PT to LateCycle (0.1317 vs 0.0817; p=6.2e-10). NK cells significantly decreased in the EarlyCycle group compared to NT (0.1109 vs 0.1462; p=0.00816) and PT (0.1109 vs 0.1513; p=0.00992), with no significant change in the LateCycle group compared to either NT or PT (p> 0.05). Conclusion: Chemotherapy significantly affects lymphocyte subsets in a treatment-specific manner. The EarlyCycle group experienced a reduction in NK cell and an increase in T cell, suggesting a damage of innate immunity and an early shift towards adaptive immunity. The LateCycle group showed a substantial decrease in B cell, indicating a delayed effect on humoral immunity components. [ABSTRACT FROM AUTHOR]

Background: The majority of patients with lung cancer exhibit drug resistance after anti-PD-1 immunotherapy, leading to shortened patient survival time. Previous studies have suggested an association between epigenetic abnormalities such as methylation and clinical response to anti-PD-1 immunotherapy, while the role of SUMOylation in resistance to anti-PD-1 antibody immunotherapy is still unclear. Methods: Here, the mRNA expression of 15 SUMO machinery components in PBMC from lung cancer patients receiving anti-PD-1 immunotherapy were analyzed using real-time PCR. Base on the percentage change in mRNA levels, the relationship between the expression of SUMO machinery components and outcomes of anti-PD-1 immunotherapy, and the influencing factors of SUMOylation were evaluated. PBMC was treated with different concentrations of 2-D08 (a specific inhibitor of SUMOylation) in vitro, and analyzed the activation and the death rates of lymphocyte subsets by flow cytometry analysis. Results: A predictive method, base on the gene expression of three SUMO machinery components (SUMO1, SUMO3 and UBE2I), were developed to distinguish non-responders to PD-1 inhibitors. Furthermore, the number of lymphocytes in peripheral blood significantly reduced in the dysregulated SUMOylation groups (the percentage change >100 or -50 ~ -100 groups). In vitro studies confirmed that lightly low SUMOylation level improved the activation status of T and NK lymphocytes, but extremely low SUMOylation level lead to the increased death rates of lymphocytes. Conclusion: Our findings implied that dysregulated gene expression of SUMO machinery components could induce the resistance of anti-PD-1 immunotherapy in lung cancer by upregulating the death of peripheral blood lymphocytes. These data might provide effective circulating biomarkers for predicting the efficacy of anti-PD-1 immunotherapy, and uncovered a novel regulatory mechanism of resistance to anti-PD-1 immunotherapy. [ABSTRACT FROM AUTHOR]

The use of flow cytometry in mice is constrained by several factors, including the limited availability of mouse-specific antibodies and the need to work with small volumes of peripheral blood. This is particularly challenging for longitudinal studies, as serial blood samples should not exceed 10% of the total blood volume in mice. To address this, we have developed two novel flow cytometry panels designed to extensively analyze immune cell populations in mice during longitudinal studies, using only 50 µL of peripheral blood per panel. Additionally, a third panel has been designed to conduct a more detailed analysis of cytotoxic and inhibitory markers at the end point. These panels have been validated on a lipopolysaccharide (LPS)-induced lung inflammation model. Two experiments were conducted to 1) validate the panels' sensitivity to immune challenges (n=12) and 2) to assess intrinsic variability of measurements (n=5). In both experiments, we collected 50 µL of peripheral blood for each cytometry panel from the maxillary venous sinus. All antibodies were titrated to identify the optimal concentration that maximized the signal from the positive population while minimizing the signal from the negative population. Samples were processed within 1 hour of collection using a MACSQuant Analyzer 16 cytometer. Our results demonstrate that these immunological panels are sensitive enough to detect changes in peripheral blood after LPS induction. Moreover, our findings help determine the sample size needed based on the immune population variability. In conclusion, the panels we have designed enable a comprehensive analysis of the murine immune system with a low blood volume requirement, enabling the measure of both absolute values and relative percentages effectively. This approach provides a robust platform for longitudinal studies in mice and can be used to uncover significant insights into immune responses. [ABSTRACT FROM AUTHOR]

Conventional CD4pos regulatory T (Treg) cells characterized by expression of the key transcription factor forkhead box P3 (FoxP3) are crucial to control immune responses, thereby maintaining homeostasis and self-tolerance. Within the substantial population of non-conventional T cell receptor (TCR)αβpos CD4negCD8αneg double-negative (dn) T cells of dogs, a novel FoxP3pos Treglike subset was described that, similar to conventional CD4pos Treg cells, is characterized by high expression of CD25. Noteworthy, human and murine TCRαβpos regulatory dn T cells lack FoxP3. Immunosuppressive capacity of canine dn T cells was hypothesized based on expression of inhibitory molecules (interleukin (IL)-10, cytotoxic T-lymphocyte associated protein 4, CTLA4). Here, to verify their regulatory function, the dnCD25pos (enriched for FoxP3pos Treg-like cells) and the dnCD25neg fraction, were isolated by fluorescence-activated cell sorting from peripheral blood mononuclear cells (PBMC) of Beagle dogs and analyzed in an in vitro suppression assay in comparison to conventional CD4posCD25pos Treg cells (positive control) and CD4posCD25neg T cells (negative control). Canine dnCD25pos T cells suppressed the Concanavalin A-driven proliferation of responder PBMC to a similar extent as conventional CD4posCD25pos Treg cells. Albeit to a lesser extent than FoxP3-enriched dn and CD4posCD25pos populations, even dnCD25neg T cells reduced the proliferation of responder cells. This is remarkable, as dnCD25neg T cells have a FoxP3neg phenotype comparable to non-suppressive CD4posCD25neg T cells. Both, CD25pos and CD25neg dn T cells, can mediate suppression independent of cell-cell contact and do not require additional signals from CD4posCD25neg T cells to secrete inhibitory factors in contrast to CD4posCD25pos T cells. Neutralization of IL-10 completely abrogated the suppression by dnCD25pos and CD4posCD25pos Treg cells in a Transwell™ system, while it only partially reduced suppression by dnCD25neg T cells. Taken together, unique canine nonconventional dnCD25pos FoxP3pos Treg-like cells are potent suppressor cells in vitro. Moreover, inhibition of proliferation of responder T cells by the dnCD25neg fraction indicates suppressive function of a subset of dn T cells even in the absence of FoxP3. The identification of unique immunoregulatory nonconventional dn T cell subpopulations of the dog in vitro is of high relevance, given the immunotherapeutic potential of manipulating regulatory T cell responses in vivo. [ABSTRACT FROM AUTHOR]

A total of 40 T2DM patients and 50 healthy controls who underwent carotid endarterectomy in the Department of Neurointerventional Intervention, Xuanwu Hospital, Capital Medical University from December 2021 to December 2022 were selected for this study. Real‑time fluorescence quantitative polymerase chain reaction (RT‑qPCR) and RNA methylation immunoprecipitation quantitative polymerase chain reaction (MeRIP‑qPCR) techniques were applied to determine the level of FASN mRNA expression and m6 A modification of FASN mRNA in their adipose tissues. Subsequently, a nested case‑control study design was used in “Functional Community Cohort” from 2018 to 2023 at the Health Management Center of Xuanwu Hospital of Capital Medical University. With 2018 as the baseline, new‑onset T2DM patients after 5 years were used as the new‑onset T2DM group (87 cases), and healthy controls were used as the control group according to the 1∶ 1 individual matching method, and the results were validated in peripheral blood. The t‑test was used to compare the differences in the expression levels of FASN mRNA and the m6 A modification levels of FASN mRNA between different groups, and Spearman correlation was used to analyze the correlation between them. Multifactorial binary logistic regression models were used to assess the strength of the association between the m6 A modification level of FASN mRNA and T2DM. Receiver operating characteristic (ROC) curves were constructed to evaluate the ability of m6 A modification levels of FASN mRNA to predict T2DM. Results FASN mRNA was highly expressed in T2DM patients (1.30±0.38 vs. 0.70±0.30 for adipose tissue, P<0.001; 1.25±0.76 vs. 1.02±0.45 for peripheral blood, P<0.01), and the m6 A modification level of FASN mRNA was low in T2DM patients (0.53±0.24 vs. 0.84±0.53 for adipose tissue, P<0.001; 0.67±0.55 vs. 1.54±0.87 for peripheral blood, P<0.001) both in adipose tissue and peripheral blood. FASN mRNA and m6 A modification level of FASN mRNA were negatively correlated (adipose tissue r=-0.360, P<0.001; peripheral blood r=-0.192, P<0.05). Baseline low m6 A modification level of FASN mRNA was a risk factor of T2DM in peripheral blood (OR=5.71, 95%CI 1.73-18.85). The ROC curve for the T2DM prediction model showed that the m6 A modification level of FASN mRNA was a good predictor of T2DM (area under the ROC curve was 0.817, 95%CI 0.753-0.888). Conclusion Abnormal m6 A modification of FASN mRNA was linked to T2DM, and the m6 A modification level of FASN mRNA can predict the risk of developing T2DM. [ABSTRACT FROM AUTHOR]

Background: Gestational diabetes mellitus (GDM) poses significant health risks for both mothers and children, contributing to long-term complications such as type 2 diabetes and cardiovascular disease. This study explores the potential of microRNAs (miRNAs) as biomarkers for GDM by analyzing peripheral blood samples from GDM patients. Method: Ten samples, including peripheral blood from 5 GDM patients and 5 controls, were collected to perform the RNA sequencing analysis. Differentially expressed miRNAs were further validated by quantitative real-time polymerase chain reaction. Results: A total of 2287 miRNAs were identified, 229 of which showed differential expression. Validation by qRT-PCR confirmed significant up-regulation of miR-5193, miR-5003-3p, miR-3127-5p, novel-miR-96, miR-6734-5p, and miR-122-5p, while miR-10395-3p was down-regulated. Bioinformatics analyses revealed the involvement of these miRNAs in pathways associated with herpes simplex virus 1 infection. Conclusion: This study provides insights into the differential expression of miRNAs in GDM patients and their potential roles in disease pathogenesis. It suggests that the differentially expressed miRNAs could serve as potential biomarkers for GDM, shedding light on the complex molecular mechanisms involved. [ABSTRACT FROM AUTHOR]

Background: With the rapid adoption of immunotherapy for the treatment of cancer comes the pressing need for readily accessible biomarkers to guide immunotherapeutic strategies and offer insights into outcomes with specific treatments. Regular sampling of solid tumour tissues outside of melanoma for immune monitoring is not often feasible; conversely, routine, frequent interrogation of circulating immune biomarkers is entirely possible. As immunotherapies and immune checkpoint inhibitors, in particular, are more widely used in first‐line, neoadjuvant, and metastatic settings, the discovery and validation of peripheral immune biomarkers are urgently needed across solid tumour types for improved prediction and prognostication of clinical outcomes in response to immunotherapy, as well as elucidation of mechanistic underpinnings of the intervention. Careful experimental design, encompassing both retrospective and prospective studies, is required in such biomarker identification studies, and concerted efforts are essential for their advancement into clinical settings. Conclusion: In this review, we summarize shared immune features between the tumour microenvironment and systemic circulation, evaluate exploratory peripheral immune biomarker studies, and discuss associations between candidate biomarkers with clinical outcomes. We also consider integration of multiple peripheral immune parameters for better prediction and prognostication and discuss considerations in study design to further evaluate the clinical utility of candidate peripheral immune biomarkers for immunotherapy of solid tumours. Highlights: Peripheral immune biomarkers are critical for improved prediction and prognostication of clinical outcomes for patients with solid tumours treated with immune checkpoint inhibition.Candidate peripheral biomarkers, such as cytokines, soluble factors, and immune cells, have potential as biomarkers to guide immunotherapy of solid tumours.Multiple peripheral immune parameters may be integrated to improve prediction and prognostication.The potential of peripheral immune biomarkers to guide immunotherapy of solid tumours requires critical work in biomarker discovery, validation, and standardization. [ABSTRACT FROM AUTHOR]

Abstract Purpose This study aimed to investigate the clinical significance of combining peripheral blood miR-21 and miR-486 with CT for the early cancer diagnosis in pulmonary nodules. Methods A total of 215 patients diagnosed with isolated pulmonary nodules with a history of smoking were selected as researchsubjects. 30 healthy volunteers with a history of smoking were recruitedas the control group.The selection of subjectswas based on the presence of isolated pulmonary nodules detected on chest CT scans. The training set consisted of 65 patients with lung nodules and 30 healthy smokers, while the verification setincluded 150 patients with lung nodules. Results Compared with the control group, the plasma expression level of miR-210 was significantly higher in the group of patients with benign pulmonary nodules (P

Abstract Background Bone marrow (BM) evaluation is the de facto standard for diagnosis, molecular analysis, risk stratification, and therapy response assessment in acute myeloid leukemia (AML), but in patients with a high number of circulating blast cells, the peripheral blood (PB) sample could provide similar information as BM. However, there is no large‐scale molecular study comparing the two specimens in terms of their gene expression profiles, cellular heterogeneities, and ex‐vivo drug sensitivity. Methodology We used (i) the BEAT‐AML cohort each with detailed molecular data; (ii) cell‐type deconvolution to estimate leukemic and immune cell proportions between specimen types; (iii) differential expression (DE) and drug‐cell type association analysis; and (iv) logistic regression models to assess the association between induction therapy response, cell‐type composition and first‐line drug treatment. Results Results: We identified 207 patients having BM and 116 patients having PB samples. There was a total of 1271 DE genes (false discovery rate

Endah Wulandari,1 Rr Ayu Fitri Hapsari2 1Department of Biochemistry, Islamic State University Syarif Hidayatullah Jakarta of Medicine Faculty, South Tangerang, Banten, Indonesia; 2Department of Histology, Islamic State University Syarif Hidayatullah Jakarta of Medicine Faculty, South Tangerang, Banten, IndonesiaCorrespondence: Endah Wulandari, Department of Biochemistry, Islamic State University Syarif Hidayatullah Jakarta of Medicine Faculty, South Tangerang, Banten, Indonesia, Email endah.wulandari@uinjkt.ac.idPurpose: The main aim of this study was to compare and analyze hematological profiles using menstrual blood, as an alternative to peripheral blood.Patients and Methods: This study used menstrual and peripheral blood samples from women who were menstruating. The design of this research is analytical observational.Results: Menstrual blood can show an overall hematological profile similar to peripheral blood. Data shows the detection of blood component parameters, white blood cells and reticulocytes in MB with a range within and outside normal blood. Data on MB that show higher values (WBC, MCH, MCHC, PLT, RDW-CV, PDW, MPV, P-LCR, PCT, neutrophils, lymphocytes, monocytes, basophils, reticulocytes, LFR, Ret-He) and lower values lower (RBC, HGB, HCT, MVC, RDW-SD, Eosinophils, IRF, MFR, HFR) when compared with peripheral blood controls. The hematological profiles of Menstrual and peripheral blood showed significant differences (p < 0.01) for several parameters, while several other parameters did not show significant differences (p > 0.05) according to the Wilcoxon test.Conclusion: All hematological profile parameters were detected in menstrual blood. The new concept that menstrual blood can be used as a supporting medium for hematological examinations opens up opportunities for developing independent hematological detection tools in productive women.Keywords: hematological, menstrual blood, peripheral blood, WBC, RBC, reticulocyte

BACKGROUND: In recent years, there have been many studies on the mechanism of exosomal non-coding RNA in gestational diabetes mellitus, but there is a lack of the latest systematic review of exosomes from different sources, especially placental sources. OBJECTIVE: To summarize the changes and potential roles of microRNA (miRNA), long non-coding RNA (lncRNA), circular RNA (circRNA), and exosomes in gestational diabetes mellitus to provide potential targets for early screening and treatment of clinical gestational diabetes mellitus. METHODS: A literature search was conducted on PubMed, Web of Science, China National Knowledge Infrastructure, WanFang Data, and VIP databases to retrieve relevant articles on non-coding RNA or exosomal non-coding RNA in relation to gestational diabetes mellitus. A total of 74 articles were included for review. RESULTS AND CONCLUSION: (1) Non-coding RNAs play important pathological and physiological roles in the lifecycle activities, and increasing evidences suggest that non-coding RNAs are involved in the occurrence and development of gestational diabetes mellitus by regulating various physiological functions. This provides a new direction for the research of gestational diabetes mellitus. (2) Exosomes are widely present in the human body. Various cells can secrete exosomes, such as red blood cells, epithelial cells, and placental cells. Non-coding RNAs found in exosomes from different sources have been demonstrated to play a role in the pathogenesis, diagnosis, and treatment of gestational diabetes mellitus. (3) MiRNA and gestational diabetes mellitus: The role of peripheral blood miRNA in gestational diabetes mellitus is mainly to affect the functions of trophoblast cells, pancreatic beta cells and blood glucose levels in gestational diabetes mellitus; placental miRNA can reflect the severity of gestational diabetes and impair the function of trophoblast cells. (4) LncRNA and gestational diabetes mellitus: Peripheral blood lncRNA can induce insulin resistance through the phosphatidylinositol 3-kinase/protein kinase B pathway and may provide new insights for the diagnosis and treatment of gestational diabetes mellitus; placental lncRNA can regulate proliferation and migration of placental trophoblast cells, promoting the occurrence and development of gestational diabetes mellitus. (5) CircRNA and gestational diabetes mellitus: Peripheral blood and placental circRNA can induce the occurrence and development of gestational diabetes mellitus by impairing the proliferation, migration and metabolism of placental trophoblast cells. (6) Non-coding RNA in exosomes and gestational diabetes mellitus: Peripheral blood non-coding RNA in exosomes can affect gestational diabetes mellitus blood glucose levels and glucose homeostasis, and participate in the occurrence and development of gestational diabetes mellitus by influencing placental function. (7) Non-coding RNA has the potential to serve as biomarkers for early diagnosis of gestational diabetes mellitus. Additionally, engineered exosomes can better achieve targeted therapy for gestational diabetes mellitus. These latest findings provide a reference for both basic research and clinical translation of gestational diabetes mellitus. (8) In the future, improvements in the extraction and purification methods of peripheral blood exosomes should be improved, and factors such as race, diet and physical activity should be excluded to improve the reproducibility of results. Further prospective clinical studies are required to explore the clinical application of circulating non-coding RNA and exosomes in the prediction and diagnosis of gestational diabetes mellitus. [ABSTRACT FROM AUTHOR]

Background: Industrial radiography uses gamma or X-ray radionuclide sources to investigate the safety of industrial materials. Industrial radiation workers receive the highest occupational radiation doses. Objective: The present study investigates the relationship between Bax and Bcl-2 gene expression variables in industrial radiation workers. Material and Methods: In this case-control study, data was collected using blood sampling from 40 workers, including two groups of non-radiation and radiation workers employed at the location. Expression levels of Bax and Bcl-2 genes were assessed in the laboratory. The environmental and absorbed doses of workers were measured using environmental and pen dosimeters. Results: Statistical analysis showed that the radiation group’s Bcl-2 gene expression level was significantly higher. Findings also demonstrated a correlation between Bcl-2 gene expression and the number of workdays. Also, the Bax gene expression did not show a significant change, and the expression ratio of Bax/Bcl-2 was insignificant in the two groups. Conclusion: Exposure to low doses of radiation could promote an adaptive response in cells by increasing Bcl-2 gene expression.

Inflammatory markers in peripheral blood, such as neutrophil-lymphocyte ratio and platelet-lymphocyte ratio, can reflect the reactive hyperplasia of inflammatory cells in tumors. The metabolic parameters of 18F-FDG PET/CT are also correlated with the reactive hyperplasia of inflammatory cells in tumors. However, only a few reports exist on the relationship between tumor metabolic parameters and peripheral blood inflammatory markers. Therefore, this review starts from three aspects: tumor peripheral blood inflammatory markers, inflammatory cell reactive hyperplasia in tumors, and 18F-FDG PET/CT metabolic parameters. The correlation between 18F-FDG PET/CT metabolic parameters and peripheral blood inflammatory markers is reviewed.

Abstract Background Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder affecting women of reproductive ages. Our previous study has implicated a possible link between RNA editing and PCOS, yet the actual role of RNA editing, its association with clinical features, and the underlying mechanisms remain unclear. Methods Ten RNA-Seq datasets containing 269 samples of multiple tissue types, including granulosa cells, T helper cells, placenta, oocyte, endometrial stromal cells, endometrium, and adipose tissues, were retrieved from public databases. Peripheral blood samples were collected from twelve PCOS and ten controls and subjected to RNA-Seq. Transcriptome-wide RNA-Seq data analysis was conducted to identify differential RNA editing (DRE) between PCOS and controls. The functional significance of DRE was evaluated by luciferase reporter assays and overexpression in human HEK293T cells. Dehydroepiandrosterone and lipopolysaccharide were used to stimulate human KGN granulosa cells to evaluate gene expression. Results RNA editing dysregulations across multiple tissues were found to be associated with PCOS in public datasets. Peripheral blood transcriptome analysis revealed 798 DRE events associated with PCOS. Through weighted gene co-expression network analysis, our results revealed a set of hub DRE events in PCOS blood. A DRE event in the eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2:chr2:37,100,559) was associated with PCOS clinical features such as luteinizing hormone (LH) and the ratio of LH over follicle-stimulating hormone. Luciferase assays, overexpression, and knockout of RNA editing enzyme adenosine deaminase RNA specific (ADAR) showed that the ADAR-mediated editing cis-regulated EIF2AK2 expression. EIAF2AK2 showed a higher expression after dehydroepiandrosterone and lipopolysaccharide stimulation, triggering changes in the downstrean MAPK pathway. Conclusions Our study presented the first evidence of cross-tissue RNA editing dysregulation in PCOS and its clinical associations. The dysregulation of RNA editing mediated by ADAR and the disrupted target EIF2AK2 may contribute to PCOS development via the MPAK pathway, underlining such epigenetic mechanisms in the disease.

Abstract Background Periprosthetic joint infection (PJI) is a severe complication that can occur after total joint arthroplasty (TJA). The timely and accurate diagnosis of PJI is the key to treatment. This study investigated the diagnostic value of platelet to lymphocyte ratio (PLR), platelet count to mean platelet volume ratio (PVR), neutrophil to lymphocyte ratio (NLR) and monocyte to lymphocyte ratio (MLR) in PJI after total knee arthroplasty (TKA) and total hip arthroplasty (THA). Methods We performed a retrospective analysis of the patients who underwent revision hip or knee arthroplasty at our Institute between June 2015 and June 2020. Of the 187 patients reviewed, 168 were included in the study. According to the diagnostic criteria of the Musculoskeletal Infection Society (MSIS), 58 patients were in the PJI group, and 110 patients were in the aseptic loosening (AL) group. We recorded and compared the preoperative peripheral blood white blood cell (WBC) count, platelet count (PLT), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), PLR, PVR, NLR, and MLR in both groups. The diagnostic performance of the WBC, PLT, PLR, PVR, NLR, and MLR individually and in combination with the ESR and CRP for PJI diagnosis was evaluated by receiver operating characteristic (ROC) curves, and the sensitivity, specificity, positive predictive value, and negative predictive value were calculated. Results Compared to those in the AL group, the mean WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR in the peripheral blood of the PJI group were significantly greater (P

Abstract Background There is an unmet need for precise biomarkers for early non-invasive breast cancer detection. Here, we aimed to identify blood-based DNA methylation biomarkers that are associated with breast cancer. Methods DNA methylation profiling was performed for 524 Asian Chinese individuals, comprising 256 breast cancer patients and 268 age-matched healthy controls, using the Infinium MethylationEPIC array. Feature selection was applied to 649,688 CpG sites in the training set. Predictive models were built by training three machine learning models, with performance evaluated on an independent test set. Enrichment analysis to identify transcription factors binding to regions associated with the selected CpG sites and pathway analysis for genes located nearby were conducted. Results A methylation profile comprising 51 CpGs was identified that effectively distinguishes breast cancer patients from healthy controls achieving an AUC of 0.823 on an independent test set. Notably, it outperformed all four previously reported breast cancer-associated methylation profiles. Enrichment analysis revealed enrichment of genomic loci associated with the binding of immune modulating AP-1 transcription factors, while pathway analysis of nearby genes showed an overrepresentation of immune-related pathways. Conclusion This study has identified a breast cancer-associated methylation profile that is immune-related to potential for early cancer detection.

Vipul Vilas Wagh,1 Tanvi Kottat,1 Suchita Agrawal,2 Shruti Purohit,2 Tejaswini Arun Pachpor,3,4 Leelavati Narlikar,5 Vasudeo Paralikar,2 Satyajeet Pramod Khare1 1Symbiosis School of Biological Sciences, Symbiosis International (Deemed University), Pune, MH, India; 2Psychiatry Unit, KEM Hospital Research Centre, Pune, MH, India; 3Department of Biosciences and Technology, School of Science and Environment Studies, Dr. Vishwanath Karad MIT World Peace University, Pune, MH, India; 4Department of Biotechnology, MES Abasaheb Garware College, Pune, MH, India; 5Department of Data Science, Indian Institute of Science Education and Research, Pune, MH, IndiaCorrespondence: Vasudeo Paralikar; Satyajeet Pramod Khare, Email paralikarv2010@gmail.com; satyajeetkhare@gmail.comIntroduction: Stigma contributes to a significant part of the burden of schizophrenia (SCZ), therefore reducing false positives from the diagnosis would be liberating for the individuals with SCZ and desirable for the clinicians. The stigmatization associated with schizophrenia advocates the need for high-precision diagnosis. In this study, we present an ensemble learning-based approach for high-precision diagnosis of SCZ using peripheral blood gene expression profiles.Methodology: The machine learning (ML) models, support vector machines (SVM), and prediction analysis for microarrays (PAM) were developed using differentially expressed genes (DEGs) as features. The SCZ samples were classified based on a voting ensemble classifier of SVM and PAM. Further, microarray-based learning was used to classify RNA sequencing (RNA-Seq) samples from our case-control study (Pune-SCZ) to assess cross-platform compatibility.Results: Ensemble learning using ML models resulted in a significantly higher precision of 80.41% (SD: 0.04) when compared to the individual models (SVM-radial: 71.69%, SD: 0.04 and PAM 77.20%, SD: 0.02). The RNA sequencing samples from our case-control study (Pune-SCZ) resulted in a moderate precision (59.92%, SD: 0.05). The feature genes used for model building were enriched for biological processes such as response to stress, regulation of the immune system, and metabolism of organic nitrogen compounds. The network analysis identified RBX1, CUL4B, DDB1, PRPF19, and COPS4 as hub genes.Conclusion: In summary, this study developed robust models for higher diagnostic precision in psychiatric disorders. Future efforts will be directed towards multi-omic integration and developing “explainable” diagnostic models. Keywords: Schizophrenia, peripheral blood, gene expression, machine learning, ensemble learning

Allergic Rhinitis (AR), a prevalent condition in otorhinolaryngology, is mediated by Type 1 hypersensitivity through IgE, characterized by Type 2 inflammatory response and eosinophil infiltration in the nasal mucosa. Since AR disease exhibits significant heterogeneity in symptom severity, an objective assessment of AR severity may facilitate better individualized treatment. To explore the changes in peripheral blood IL-9, Th9, and BAFF levels of allergic rhinitis (AR) in patients and the clinical significance associated with it. A retrospective study selected 80 AR patients admitted from January 2022 to October 2022 as the case group, dividing them into mild and moderate-to-severe groups based on symptom scores. Concurrently, 50 patients without AR, who were treated for nasal bone fractures or underwent septoplasty, were selected as the group for comparison. Alterations in the expression levels of peripheral blood IL-9, Th9, and BAFF were analyzed and compared among the different groups. The diagnostic value of serum BAFF for the severity of AR was analyzed using the receiver operating characteristic (ROC) curve. Noticeable variations were observed in clinical variables among the three groups such as, total IgE levels, peripheral blood eosinophil count and proportion, TNSS, and VAS (P< 0.05), while no statistically significant differences were observed in other variables (P> 0.05). The comparison of IL-9, Th9, and BAFF among the three groups revealed statistically significant differences (P< 0.05). Analysis using multivariate logistic regression revealed that IL-9 (OR = 2.365), Th9 (OR = 2.186), BAFF (OR = 2.307) were influencing factors of moderate-to-severe AR (P< 0.05). The ROC curve indicated that the AUC for the diagnosis of moderate-to-severe AR by IL-9, Th9, BAFF were 0.770, 0.734, 0.761, respectively, and the combined detection AUC was 0.888, an area under the curve higher than individual testing. Changes in peripheral blood IL-9, Th9, and BAFF levels in AR patients may function as indicators to assess the level of severity in diagnostic procedures. [ABSTRACT FROM AUTHOR]

Background: Hematological abnormalities in COVID-19 infection included quantitative and qualitative changes and should be further characterized. Evaluation for myelodysplastic syndromes (MDS) is usually prompted by abnormal hematologic findings and the presence of dysplastic morphologies. Viral infections are considered to be the cause of dysplastic morphologies and should be considered by morphologists. There are few reports of dysplastic abnormal morphologies in patients with COVID-19 infection. However, such correlations still have to be clarified. Materials and Methods: In the present study, we examined the granulocyte lineage morphological abnormalities in symptomatic RT-PCR-confirmed COVID patients. Peripheral blood samples were collected from 82 patients with symptomatic COVID-19. Blood smears were prepared according to the standard Wright-Giemsa staining procedure. The morphological examination was carried out by two laboratory experts. Results: Blood smear examination revealed common myelodysplastic syndrome (MDS) type abnormalities including but not limited to pseudo-pelger nuclear lobulation (4.8%), hypogranulation (7.3%), Howell-Jolly-like bodies or detached nuclear segments (6.0%) and elongated and thin nuclear filaments (6.0%). One case of abnormal immature granulocyte and ring form nucleus is also evident. Conclusion: Our results accounted for the possibility of active COVID-19 infection in all subjects with granulocyte dysplasia. These results are of practical importance for patients suspected of having myelodysplastic syndromes or disease processes associated with myeloid malignancies. [ABSTRACT FROM AUTHOR]

In sexual assault cases, it is crucial to discriminate between peripheral blood and menstrual blood to provide evidence for vaginal intercourse with traumatic injury. In this study, the menstrual blood mRNA markers progestagen-associated endometrial protein (PAEP), matrix metallopeptidase 7 (MMP7), and left–right determination factor 2 (LEFTY2) were evaluated by quantitative RT-PCR (RT-qPCR) for the discrimination of menstrual blood from peripheral blood and vaginal fluid. As a result, all markers with cutoff delta cycle quantification (ΔCq) values were specifically determined in menstrual blood among forensically relevant body fluids. Even though the changes in the expression levels of each marker differed during the menstrual cycle, all markers were determined to be positive in most of the randomly collected menstrual blood samples that were analyzed. Additionally, the markers with proposed cutoff ΔCq values could discriminate between menstrual blood and peripheral blood-mixed vaginal fluid samples. The determination of positive markers was less affected by storage temperature under dry conditions than under wet conditions, while PAEP was detectable in samples stored below room temperature under wet conditions. The detectability of PAEP was considered to be the result of its higher expression level compared with MMP7 and LEFTY2. In conclusion, menstrual blood markers for the RT-qPCR procedure evaluated in this study were highly specific for menstrual blood. The proposed procedure could be useful for discriminating between menstruation and traumatic bleeding in the female genital tract. In particular, PAEP is expected to be applicable to forensic casework samples because of its high specificity and robustness. [ABSTRACT FROM AUTHOR]

Despite advances in the genomic classification of breast cancer, current clinical tests and treatment decisions are commonly based on protein-level information. Nowadays breast cancer clinical treatment selection is based on the immunohistochemical (IHC) determination of four protein biomarkers: Estrogen Receptor 1 (ESR1), Progesterone Receptor (PGR), Human Epidermal Growth Factor Receptor 2 (HER2), and proliferation marker Ki-67. The prognostic correlation of tumor-infiltrating T cells has been widely studied in breast cancer, but tumor-infiltrating B cells have not received so much attention. We aimed to find a correlation between immunohistochemical results and a proteomic approach in measuring the expression of proteins isolated from B-cell lymphocytes in peripheral blood samples. Shotgun proteomic analysis was chosen for its key advantage over other proteomic methods, which is its comprehensive and untargeted approach to analyzing proteins. This approach facilitates better characterization of disease-associated changes at the protein level. We identified 18 proteins in B cell lymphocytes with a significant fold change of more than 2, which have promising potential to serve as breast cancer biomarkers in the future. [ABSTRACT FROM AUTHOR]

Background: Prior studies report associations of maternal serum Lamin A, encoded by the LMNA gene, with fetal congenital heart disease (CHD). It is unknown whether DNA methylation (DNAm) of cytosine‐phosphate‐guanine (CpG) sites in LMNA impacts the CHD susceptibility. Methods: We investigated the associations of LMNA DNAm with CHD using publicly available data of CHD cases (n = 197) and controls (n = 134) from the Gene Expression Omnibus repository. Peripheral blood DNAm was measured using Illumina 850 K BeadChip for cases and 450 K BeadChip for controls. We tested 31 LMNA CpGs to identify differences in DNAm between cases and controls using linear regression correcting for multiple testing with false discovery rate (FDR). In a case‐only analysis, we tested the variations in LMNA DNAm between CHD subtypes. To identify the consistency of DNAm across tissue types we compared peripheral blood (n = 197) and heart tissue DNAm (n = 20) in CHD cases. Results: After adjusting for age, sex, and cell types there were significant differences in 17 of the 31 LMNA CpGs between CHD cases and controls (FDR p ≤.05). We identified lower DNAm of cg09820673 at 3′ UTR for hypoplastic left heart syndrome compared to other CHD subtypes. Three CpGs exhibited uniform DNAm in blood and heart tissues in cases. Eleven CpGs showed changes in the same direction in blood and heart tissues in cases compared to controls. Conclusion: We identify statistically significant differences in LMNA DNAm between CHD cases and controls. Future studies should investigate the role of maternal LMNA DNAm in CHD development. [ABSTRACT FROM AUTHOR]

Parkinson's disease (PD) is one of the most common human neurodegenerative diseases. Belated diagnoses of PD and late treatment are caused by its elongated prodromal phase. Thus, searching for new candidate genes participating in the development of the pathological process in the early stages of the disease in patients who have not yet received therapy is relevant. Changes in mRNA and protein levels have been described both in the peripheral blood and in the brain of patients with PD. Thus, analysis of changes in the mRNA expression in peripheral blood is of great importance in studying the early stages of PD. This work aimed to analyze the changes in MEF2C, SLC22A4, P2RY12, and LRRN3 gene expression in the peripheral blood of patients in the early stages of PD. We found a statistically relevant and PD-specific change in the expression of the LRRN3 gene, indicating a disruption in the processes of neuronal regeneration and the functioning of synapses. The data obtained during the study indicate that this gene can be considered a potential biomarker of the early stages of PD. [ABSTRACT FROM AUTHOR]

AbstractBackgroundMethodsResultsConclusionsRUNX3 is hypermethylated in multiple cancers. TIMP2 also functions as a regulator of tumors. However, there are only very few reports on the association of methylation of RUNX3 and TIMP2 with lung cancer (LC) in peripheral blood.426 LC patients and 428 age- and sex-matched healthy controls were recruited. DNA methylation in blood was semi-quantitively assessed by mass spectrometry. For the association analysis, binary logistic regression analysis adjusted covariant was applied, and ORs were presented as per +10% methylation.Hypermethylation of CpG_1, CpG_5 and CpG_8 in RUNX3 was significantly associated with LC (ORs = 1.45, 1.35 and 1.35, respectively, adjusted p < 0.05), and even stage I LC. The association between the three RUNX3 CpG sites and LC was enhanced by increased age (> 55 years, ORs ranged from 1.43 to 1.75, adjusted p < 0.05), male gender (ORs ranged from 1.47 to 1.59, adjusted p < 0.05) and tumor stage (stage II&III&IV, ORs ranged from 1.86 to 3.03, adjusted p < 0.05).This study suggests a significant association between blood-based RUNX3 hypermethylation and LC, especially in elder people, in males and in LC patients with advanced stage. [ABSTRACT FROM AUTHOR]

Objectives: Programmed cell death protein-1 (PD-1) maintains peripheral immune tolerance by preventing T cell continuous activation. Aiming to understand the extent of PD-1 expression in inflammatory arthritis beyond its involvement with T cells, we assess its presence on various circulating single cells. Methods: Mass cytometry analysis of patients with active seropositive/seronegative rheumatoid (RA; n=9/8) and psoriatic (PsA; n=9) arthritis versus healthy controls (HC; n=13), re-evaluating patients after 3 months of anti-rheumatic treatment. Results: PD-1 was expressed in all leukocyte subpopulations, with the highest PD-1+ cell frequencies in eosinophils (59-73%) and T cells (50–60%), and the lowest in natural-killer cells (1–3%). PD-1+ cell frequencies and PD-1 median expression were comparable between patient subgroups and HC, in the majority of cell subsets. Exceptions included increases in certain T cell/B cell subsets of seropositive RA and specific monocyte subsets and dendritic cells of PsA; an expanded PD-1+CD4+CD45RA+CD27+CD28+ T subset, denoting exhausted T cells, was common across patient subgroups. Strikingly, significant inverse correlations between individual biomarkers of systemic inflammation (ESR and/or serum CRP) and PD-1+ cell frequencies and/or median expression were evident in several innate and adaptive immunity cell subsets of RA and PsA patients. Furthermore, all inverse correlations noted in individuals with active arthritis were no longer discernible in those who attained remission/low disease activity post-treatment. Conclusion: PD-1 expression may be insufficient, relative to the magnitude of the concomitant systemic inflammatory response on distinct leukocyte subsets, varying between RA and PsA. Our results point to the potential therapeutic benefits of pharmacological PD-1 activation, to rebalance the autoimmune response and reduce inflammation. [ABSTRACT FROM AUTHOR]

Background: Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder affecting women of reproductive ages. Our previous study has implicated a possible link between RNA editing and PCOS, yet the actual role of RNA editing, its association with clinical features, and the underlying mechanisms remain unclear. Methods: Ten RNA-Seq datasets containing 269 samples of multiple tissue types, including granulosa cells, T helper cells, placenta, oocyte, endometrial stromal cells, endometrium, and adipose tissues, were retrieved from public databases. Peripheral blood samples were collected from twelve PCOS and ten controls and subjected to RNA-Seq. Transcriptome-wide RNA-Seq data analysis was conducted to identify differential RNA editing (DRE) between PCOS and controls. The functional significance of DRE was evaluated by luciferase reporter assays and overexpression in human HEK293T cells. Dehydroepiandrosterone and lipopolysaccharide were used to stimulate human KGN granulosa cells to evaluate gene expression. Results: RNA editing dysregulations across multiple tissues were found to be associated with PCOS in public datasets. Peripheral blood transcriptome analysis revealed 798 DRE events associated with PCOS. Through weighted gene co-expression network analysis, our results revealed a set of hub DRE events in PCOS blood. A DRE event in the eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2:chr2:37,100,559) was associated with PCOS clinical features such as luteinizing hormone (LH) and the ratio of LH over follicle-stimulating hormone. Luciferase assays, overexpression, and knockout of RNA editing enzyme adenosine deaminase RNA specific (ADAR) showed that the ADAR-mediated editing cis-regulated EIF2AK2 expression. EIAF2AK2 showed a higher expression after dehydroepiandrosterone and lipopolysaccharide stimulation, triggering changes in the downstrean MAPK pathway. Conclusions: Our study presented the first evidence of cross-tissue RNA editing dysregulation in PCOS and its clinical associations. The dysregulation of RNA editing mediated by ADAR and the disrupted target EIF2AK2 may contribute to PCOS development via the MPAK pathway, underlining such epigenetic mechanisms in the disease. [ABSTRACT FROM AUTHOR]

To investigate the effect of Suhuang Zhike capsule combined with thymosin in the treatment of elderly patients with COPD complicated with pulmonary infection and its influence on immune function and blood gas indexes. 80 elderly patients treated in hospital were studied. All patients were diagnosed with COPD disease and were complicated with pulmonary infection. The admission time of the patients was distributed from September 2020 to September 2023, and the patients were divided into groups according to different treatment methods, among which 40 patients were divided into matched group and thymosin treatment was added on the basis of conventional symptomatic treatment, and the other 40 patients were divided into observation group and Suhuang Zhike capsule was added on the basis of matched group. Compared the total therapeutic effective rate, inflammatory factors [C-reactive protein (CRP) and tumor necrosis factor-α (TNF-α)] of the two groups, immune function [CD3+, CD4+ /CD8+ , Natural killer cell, NK)], blood gas index [partial pressure of carbon dioxide (PaCO2), arterial partial pressure of oxygen oxygen, PaO2), pH], occurrence of adverse reactions [nausea, vomiting, dizziness, rash, diarrhea]. The total response rate in the observation group was higher than that in the matched group (P<0.05). Post-treatment, the CRP and TNF- α levels in the observation group were significantly lower than those in the matched group (P<0.05). Post-treatment, the levels of CD3+, CD4+ /CD8+, and NK were higher in the observation group than in the matched group (P<0.05). After receiving treatment, the PaCO2 index level in the observation group was lower than that in the matched group (P<0.05), and the level of PaO2 and pH index in the observation group was higher than that in the matched group (P<0.05). The total incidence of adverse reactions such as nausea, vomiting, dizziness, rash and diarrhea in the two groups was 2.50% in the observation group and 7.50% in the matched group. There was no difference in the total incidence of adverse reactions between the two groups (P>0.05). In elderly patients with COPD complicated with pulmonary infection, the addition of Suhuang Zhike capsule on the basis of conventional symptomatic treatment and thymosin treatment can significantly improve the total effective rate of clinical treatment, improve the level of inflammatory factors and blood gas indexes, and enhance immune function, without increasing the risk of adverse reactions [ABSTRACT FROM AUTHOR]

Inflammatory markers in peripheral blood, such as neutrophil-lymphocyte ratio and platelet- lymphocyte ratio, can reflect the reactive hyperplasia of inflammatory cells in tumors. The metabolic parameters of "F-FDG PET/CT are also correlated with the reactive hyperplasia of inflammatory cells in tumors. However, only a few reports exist on the relationship between tumor metabolic parameters and peripheral blood inflammatory markers. Therefore, this review starts from three aspects: tumor peripheral blood inflammatory markers, inflammatory cell reactive hyperplasia in tumors, and "F-FDG PET/CT metabolic parameters. The correlation between "F-FDG PET/CT metabolic parameters and peripheral blood inflammatory markers is reviewed. [ABSTRACT FROM AUTHOR]

Background: There is an unmet need for precise biomarkers for early non-invasive breast cancer detection. Here, we aimed to identify blood-based DNA methylation biomarkers that are associated with breast cancer. Methods: DNA methylation profiling was performed for 524 Asian Chinese individuals, comprising 256 breast cancer patients and 268 age-matched healthy controls, using the Infinium MethylationEPIC array. Feature selection was applied to 649,688 CpG sites in the training set. Predictive models were built by training three machine learning models, with performance evaluated on an independent test set. Enrichment analysis to identify transcription factors binding to regions associated with the selected CpG sites and pathway analysis for genes located nearby were conducted. Results: A methylation profile comprising 51 CpGs was identified that effectively distinguishes breast cancer patients from healthy controls achieving an AUC of 0.823 on an independent test set. Notably, it outperformed all four previously reported breast cancer-associated methylation profiles. Enrichment analysis revealed enrichment of genomic loci associated with the binding of immune modulating AP-1 transcription factors, while pathway analysis of nearby genes showed an overrepresentation of immune-related pathways. Conclusion: This study has identified a breast cancer-associated methylation profile that is immune-related to potential for early cancer detection. [ABSTRACT FROM AUTHOR]

Background: The presence of circulating plasma cells (CPCs) is an important laboratory indicator for the diagnosis, staging, risk stratification, and progression monitoring of multiple myeloma (MM). Early detection of CPCs in the peripheral blood (PB) followed by timely interventions can significantly improve MM prognosis and delay its progression. Although the conventional cell morphology examination remains the predominant method for CPC detection because of accessibility, its sensitivity and reproducibility are limited by technician expertise and cell quantity constraints. This study aims to develop an artificial intelligence (AI)-based automated system for a more sensitive and efficient CPC morphology detection. Methods: A total of 137 bone marrow smears and 72 PB smears from patients with at Zhongshan Hospital, Fudan University, were retrospectively reviewed. Using an AI-powered digital pathology platform, Morphogo, 305,019 cell images were collected for training. Morphogo's efficacy in CPC detection was evaluated with additional 184 PB smears (94 from patients with MM and 90 from those with other hematological malignancies) and compared with manual microscopy. Results: Morphogo achieved 99.64% accuracy, 89.03% sensitivity, and 99.68% specificity in classifying CPCs. At a 0.60 threshold, Morphogo achieved a sensitivity of 96.15%, which was approximately twice that of manual microscopy, with a specificity of 78.03%. Patients with CPCs detected by AI scanning had a significantly shorter median progression-free survival compared with those without CPC detection (18 months vs. 34 months, p< .01). Conclusions: Morphogo is a highly sensitive system for the automated detection of CPCs, with potential applications in initial screening, prognosis prediction, and posttreatment monitoring for MM patients. Plain Language Summary: Diagnosing and monitoring multiple myeloma (MM), a type of blood cancer, requires identifying and quantifying specific cells called circulating plasma cells (CPCs) in the blood. The conventional method for detecting CPCs is manual microscopic examination, which is time-consuming and lacks sensitivity. This study introduces a highly sensitive CPC detection method using an artificial intelligence-based system, Morphogo. It demonstrated remarkable sensitivity and accuracy, surpassing conventional microscopy. This advanced approach suggests that early and accurate CPC detection is achievable by morphology examination, making efficient CPC screening more accessible for patients with MM. This innovative system has the potential to be used in the diagnosis and risk assessment of MM. [ABSTRACT FROM AUTHOR]

Background: Periprosthetic joint infection (PJI) is a severe complication that can occur after total joint arthroplasty (TJA). The timely and accurate diagnosis of PJI is the key to treatment. This study investigated the diagnostic value of platelet to lymphocyte ratio (PLR), platelet count to mean platelet volume ratio (PVR), neutrophil to lymphocyte ratio (NLR) and monocyte to lymphocyte ratio (MLR) in PJI after total knee arthroplasty (TKA) and total hip arthroplasty (THA). Methods: We performed a retrospective analysis of the patients who underwent revision hip or knee arthroplasty at our Institute between June 2015 and June 2020. Of the 187 patients reviewed, 168 were included in the study. According to the diagnostic criteria of the Musculoskeletal Infection Society (MSIS), 58 patients were in the PJI group, and 110 patients were in the aseptic loosening (AL) group. We recorded and compared the preoperative peripheral blood white blood cell (WBC) count, platelet count (PLT), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), PLR, PVR, NLR, and MLR in both groups. The diagnostic performance of the WBC, PLT, PLR, PVR, NLR, and MLR individually and in combination with the ESR and CRP for PJI diagnosis was evaluated by receiver operating characteristic (ROC) curves, and the sensitivity, specificity, positive predictive value, and negative predictive value were calculated. Results: Compared to those in the AL group, the mean WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR in the peripheral blood of the PJI group were significantly greater (P < 0.05). The analysis of the ROC curve revealed that the ESR, CRP, PLR, PVR, NLR, and MLR in peripheral blood had moderate effectiveness in diagnosing PJI, with area under the curve (AUC) values of 0.760 (95% CI: 0.688–0.823), 0.758 (95% CI: 0.687–0.821), 0.714 (95% CI: 0.639–0.781), 0.709 (95% CI: 0.634–0.777), 0.723 (95% CI: 0.649–0.789), and 0.728 (95% CI: 0.654–0.793), respectively. Conversely, the WBC and PLT counts demonstrated poor diagnostic value for PJI, with AUC values of 0.578 (95% CI: 0.499–0.653) and 0.694 (95% CI: 0.619–0.763), respectively. The results of the prediction model calculations revealed that the combined AUC of the WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR was the highest at 0.853 (95% CI, 0.790–0.909), indicating good value in the diagnosis of PJI, with a sensitivity of 82.8% and a specificity of 72.7%. Moreover, the novel composite of parameters improved the accuracy and reliability in diagnosing PJI compared to the traditional biomarkers ESR and CRP (P = 0.015). Conclusion: Our study suggested that the diagnostic value of the peripheral blood biomarkers PLR, PVR, NLR, and MLR for diagnosing PJI is limited and not superior to that of the ESR or CRP. However, when the WBC, PLT, ESR, CRP, PLR, PVR, NLR, and MLR are combined, the diagnostic performance of PJI in TJA patients can be improved. [ABSTRACT FROM AUTHOR]

This study aimed to explore the distribution, characteristics and prognostic value of baseline peripheral blood lymphocyte subsets in patients with extranodal NK/T-cell lymphoma (NKTCL). We conducted this cross-sectional study of 205 newly-diagnosed NKTCL patients receiving first-line chemotherapy and radiation at our institute between 2010 and 2020. Baseline peripheral blood lymphocytes were detected using flow cytometry, and the clinical value was analyzed. Compared with healthy controls, patients with NKTCL presented with a distinct peripheral immunity with higher levels of cytotoxic CD8+ T cells (33.230 ± 12.090% vs. 27.060 ± 4.010%, p < 0.001) and NKT cells (7.697 ± 7.219% vs. 3.550 ± 2.088%, p < 0.001) but lower proportions of suppressive regulatory T cells (Treg, 2.999 ± 1.949% vs. 3.420 ± 1.051%, p = 0.003) and CD4+ helper T cells (Th, 33.084 ± 11.361% vs. 37.650 ± 3.153%, p < 0.001). Peripheral lymphocytes were differentially distributed according to age, stage, and primary site in patients with NKTCL. The proportion of Th cells/lymphocytes was associated with tumor burden reflected by stage (p = 0.037), serum lactate dehydrogenase (p = 0.0420), primary tumor invasion (p = 0.025), and prognostic index for NK/T-cell lymphoma (PINK) score (p = 0.041). Furthermore, elevated proportions of T cells (58.9% vs. 76.4%, p = 0.005), Th cells (56.3% vs. 68.8%, p = 0.047), or Treg cells (49.5% vs. 68.9%, p = 0.040) were associated with inferior 5-year progression-free survivals (PFS) via univariable survival analysis. Multivariate cox regression revealed elevated Th cells as an independent predictor for unfavorable PFS (HR = 2.333, 95% CI, 1.030–5.288, p = 0.042) in NKTCL. These results suggested the proportion of Th cells positively correlated with tumor burden and was a potential non-invasive biomarker for inferior survival for patients with NKTCL. [ABSTRACT FROM AUTHOR]

Background: Industrial radiography uses gamma or X-ray radionuclide sources to investigate the safety of industrial materials. Industrial radiation workers receive the highest occupational radiation doses. Objective: The present study investigates the relationship between Bax and Bcl-2 gene expression variables in industrial radiation workers. Material and Methods: In this case-control study, data was collected using blood sampling from 40 workers, including two groups of non-radiation and radiation workers employed at the location. Expression levels of Bax and Bcl-2 genes were assessed in the laboratory. The environmental and absorbed doses of workers were measured using environmental and pen dosimeters. Results: Statistical analysis showed that the radiation group's Bcl-2 gene expression level was significantly higher. Findings also demonstrated a correlation between Bcl-2 gene expression and the number of workdays. Also, the Bax gene expression did not show a significant change, and the expression ratio of Bax/Bcl-2 was insignificant in the two groups. Conclusion: Exposure to low doses of radiation could promote an adaptive response in cells by increasing Bcl-2 gene expression. [ABSTRACT FROM AUTHOR]

Background: Infection with Treponema pallidum instigates complex immune responses. Prior research has suggested that persistent Treponema pallidum infection can manipulate host immune responses and circumvent host defenses. However, the precise role of immune cells in Treponema pallidum infection across different stages remains a contentious issue. Methods: Utilizing summary data from genome-wide association studies, we employed a two-sample Mendelian randomization method to investigate the association between 731 immunophenotypes and syphilis. Syphilis was categorized into early and late stages in this study to establish a more robust correlation and minimize bias in database sources. Results: Our findings revealed that 33, 36, and 27 immunophenotypes of peripheral blood were associated with syphilis (regardless of disease stage), early syphilis and late syphilis, respectively. Subsequent analysis demonstrated significant variations between early and late syphilis in terms of immunophenotypes. Specifically, early syphilis showcased activated, secreting, and resting regulatory T cells, whereas late syphilis was characterized by resting Treg cells. More B cells subtypes emerged in late syphilis. Monocytes in early syphilis exhibited an intermediate and non-classical phenotype, transitioning to classical in late syphilis. Early syphilis featured naive T cells, effector memory T cells, and terminally differentiated T cells, while late syphilis predominantly presented terminally differentiated T cells. Immature myeloid-derived suppressor cells were evident in early syphilis, whereas the dendritic cell immunophenotype was exclusive to late syphilis. Conclusion: Multiple immunophenotypes demonstrated associations with syphilis, showcasing substantial disparities between the early and late stages of the disease. These findings hold promise for informing immunologically oriented treatment strategies, paving the way for more effective and efficient syphilis interventions. [ABSTRACT FROM AUTHOR]

The underlying mechanism of fluorosis has not been fully elucidated. The purpose of this study was to explore the mechanism of fluorosis induced by sodium fluoride (NaF) using proteomics. Six offspring rats exposed to fluoride without dental fluorosis were defined as group A, 8 offspring rats without fluoride exposure were defined as control group B, and 6 offspring rats exposed to fluoride with dental fluorosis were defined as group C. Total proteins from the peripheral blood were extracted and then separated using liquid chromatography–tandem mass spectrometry. The identified criteria for differentially expressed proteins were fold change > 1.2 or < 0.83 and P < 0.05. Gene Ontology function annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed using the oeCloud tool. The 177 upregulated and 22 downregulated proteins were identified in the A + C vs. B group. KEGG pathway enrichment analysis revealed that transforming growth factor-β (TGF-β) signaling pathway significantly enriched. PPI network constructed using Cytoscape confirmed RhoA may play a crucial role. The KEGG results of genes associated with fluoride and genes associated with both fluoride and inflammation in the GeneCards database also showed that TGF-β signaling pathway was significantly enriched. The immunofluorescence in HPA database showed that the main expression sites of RhoA are plasma membrane and cytosol, while the main expression site of Fbn1 is the Golgi apparatus. In conclusion, long-term NaF intake may cause inflammatory response in the peripheral blood of rats by upregulating TGF-β signaling pathway, in which RhoA may play a key role. [ABSTRACT FROM AUTHOR]

The complexity and heterogeneity of PD necessitate advanced diagnostic and prognostic tools to elucidate its molecular mechanisms accurately. In this study, we addressed this challenge by conducting a pilot phospho-proteomic analysis of peripheral blood mononuclear cells (PBMCs) from idiopathic PD patients at varying disease stages to delineate the functional alterations occurring in these cells throughout the disease course and identify key molecules and pathways contributing to PD progression. By integrating clinical data with phospho-proteomic profiles across various PD stages, we identify potential stage-specific molecular signatures indicative of disease progression. This integrative approach allows for the discernment of distinct disease states and enhances our understanding of PD heterogeneity.

The pathology of respiratory syncytial virus (RSV) infection remains unclear. An unbalanced immune response to RSV infection can lead to immunopathology, causing airway damage and impaired exchange of oxygen and carbon dioxide between the air and the bloodstream. We aimed to evaluate the association of the expression of inflammatory and antiviral genes in peripheral blood with severe hypoxemia in children with RSV infection seen in the hospital emergency room. We conducted a cross-sectional study on 121 RSV-infected children seen in hospital emergency rooms between 2015 and 2023. Total RNA was extracted from whole blood samples, and gene expression (IL-6, TNFα, CXCL8, ISG15, IFIT1, RIGI, IFNβ, CCL5, and CXCL10) was quantified using quantitative RT-PCR. The outcome variable was having severe hypoxemia (SpO2 ≤ 90%). The association analysis was performed using a volcano plot, adjusted logistic regression, and orthogonal partial least squares discriminant analysis (OPLS-DA). We found that 26 of 121 children had severe hypoxemia (SpO2 ≤ 90%). CXCL8 was overexpressed [fold changes (FC) > 2; q-value < 0.05], and ISG15, IFIT1, RIGI, IFNβ, CCL5, and CXCL10 were underexpressed (FC

ObjectiveThis study aims to explore the association of methyltransferase-like protein 3 (METTL3) expression with severity of coronary artery stenosis in patients with coronary heart disease (CHD).MethodsA total of 100 patients administrated in the Fourth Affiliated Hospital of China Medical University between October 2022 and June 2023 with primary symptoms of chest pain or tightness, or cardiac discomfort, and who underwent coronary angiography for a definitive diagnosis, were included in the study. The baseline characteristics, including TG, TC, LDL-C, HDL-C, uric acid and past history were recorded. Peripheral blood samples were collected to assess the expression levels of METTL3, YT521-B homology domains 1 (YTHDF1), YT521-B homology domains 2 (YTHDF2), and YT521-B homology domains 3 (YTHDF3) using the PCR method. Relative expression levels of METTL3 protein were determined by Western blotting. Correlation analysis were conducted to evaluate the relationship between METTL3/YTHDF1 gene expression and clinical data. Receiver operating characteristic (ROC) curve analysis was employed to assess the predictive value of METTL3 and YTHDF1 for CHD. Binary logistic regression was used to determine whether the expression of METTL3 and YTHDF1 in peripheral blood were risk factors for CHD.ResultsThe study found no significant differences in baseline characteristics between CHD patients and controls, except for length of stay, Lymphocytes, Neutrophils, AST, HDL-C and modified Gensini score. The gene expression levels of METTL3 and YTHDF1 were significantly higher in CHD patients compared to controls (p 0.052. The AUC for YTHDF1 in CHD was 0.623, with a sensitivity of 47.4% and a specificity of 74.4% at a cut-off value of >0.027. Binary logistic regression revealed that only increased METTL3 expression in peripheral blood was an independent risk factor for CHD (p

Abstract Background With the rapid adoption of immunotherapy for the treatment of cancer comes the pressing need for readily accessible biomarkers to guide immunotherapeutic strategies and offer insights into outcomes with specific treatments. Regular sampling of solid tumour tissues outside of melanoma for immune monitoring is not often feasible; conversely, routine, frequent interrogation of circulating immune biomarkers is entirely possible. As immunotherapies and immune checkpoint inhibitors, in particular, are more widely used in first‐line, neoadjuvant, and metastatic settings, the discovery and validation of peripheral immune biomarkers are urgently needed across solid tumour types for improved prediction and prognostication of clinical outcomes in response to immunotherapy, as well as elucidation of mechanistic underpinnings of the intervention. Careful experimental design, encompassing both retrospective and prospective studies, is required in such biomarker identification studies, and concerted efforts are essential for their advancement into clinical settings. Conclusion In this review, we summarize shared immune features between the tumour microenvironment and systemic circulation, evaluate exploratory peripheral immune biomarker studies, and discuss associations between candidate biomarkers with clinical outcomes. We also consider integration of multiple peripheral immune parameters for better prediction and prognostication and discuss considerations in study design to further evaluate the clinical utility of candidate peripheral immune biomarkers for immunotherapy of solid tumours. Highlights Peripheral immune biomarkers are critical for improved prediction and prognostication of clinical outcomes for patients with solid tumours treated with immune checkpoint inhibition. Candidate peripheral biomarkers, such as cytokines, soluble factors, and immune cells, have potential as biomarkers to guide immunotherapy of solid tumours. Multiple peripheral immune parameters may be integrated to improve prediction and prognostication. The potential of peripheral immune biomarkers to guide immunotherapy of solid tumours requires critical work in biomarker discovery, validation, and standardization.

The use of flow cytometry in mice is constrained by several factors, including the limited availability of mouse-specific antibodies and the need to work with small volumes of peripheral blood. This is particularly challenging for longitudinal studies, as serial blood samples should not exceed 10% of the total blood volume in mice. To address this, we have developed two novel flow cytometry panels designed to extensively analyze immune cell populations in mice during longitudinal studies, using only 50 µL of peripheral blood per panel. Additionally, a third panel has been designed to conduct a more detailed analysis of cytotoxic and inhibitory markers at the end point. These panels have been validated on a lipopolysaccharide (LPS)-induced lung inflammation model. Two experiments were conducted to 1) validate the panels’ sensitivity to immune challenges (n=12) and 2) to assess intrinsic variability of measurements (n=5). In both experiments, we collected 50 µL of peripheral blood for each cytometry panel from the maxillary venous sinus. All antibodies were titrated to identify the optimal concentration that maximized the signal from the positive population while minimizing the signal from the negative population. Samples were processed within 1 hour of collection using a MACSQuant Analyzer 16 cytometer. Our results demonstrate that these immunological panels are sensitive enough to detect changes in peripheral blood after LPS induction. Moreover, our findings help determine the sample size needed based on the immune population variability. In conclusion, the panels we have designed enable a comprehensive analysis of the murine immune system with a low blood volume requirement, enabling the measure of both absolute values and relative percentages effectively. This approach provides a robust platform for longitudinal studies in mice and can be used to uncover significant insights into immune responses.

Conventional CD4pos regulatory T (Treg) cells characterized by expression of the key transcription factor forkhead box P3 (FoxP3) are crucial to control immune responses, thereby maintaining homeostasis and self-tolerance. Within the substantial population of non-conventional T cell receptor (TCR)αβpos CD4negCD8αneg double-negative (dn) T cells of dogs, a novel FoxP3pos Treg-like subset was described that, similar to conventional CD4pos Treg cells, is characterized by high expression of CD25. Noteworthy, human and murine TCRαβpos regulatory dn T cells lack FoxP3. Immunosuppressive capacity of canine dn T cells was hypothesized based on expression of inhibitory molecules (interleukin (IL)-10, cytotoxic T-lymphocyte associated protein 4, CTLA4). Here, to verify their regulatory function, the dnCD25pos (enriched for FoxP3pos Treg-like cells) and the dnCD25neg fraction, were isolated by fluorescence-activated cell sorting from peripheral blood mononuclear cells (PBMC) of Beagle dogs and analyzed in an in vitro suppression assay in comparison to conventional CD4posCD25pos Treg cells (positive control) and CD4posCD25neg T cells (negative control). Canine dnCD25pos T cells suppressed the Concanavalin A-driven proliferation of responder PBMC to a similar extent as conventional CD4posCD25pos Treg cells. Albeit to a lesser extent than FoxP3-enriched dn and CD4posCD25pos populations, even dnCD25neg T cells reduced the proliferation of responder cells. This is remarkable, as dnCD25neg T cells have a FoxP3neg phenotype comparable to non-suppressive CD4posCD25neg T cells. Both, CD25pos and CD25neg dn T cells, can mediate suppression independent of cell-cell contact and do not require additional signals from CD4posCD25neg T cells to secrete inhibitory factors in contrast to CD4posCD25pos T cells. Neutralization of IL-10 completely abrogated the suppression by dnCD25pos and CD4posCD25pos Treg cells in a Transwell™ system, while it only partially reduced suppression by dnCD25neg T cells. Taken together, unique canine non-conventional dnCD25pos FoxP3pos Treg-like cells are potent suppressor cells in vitro. Moreover, inhibition of proliferation of responder T cells by the dnCD25neg fraction indicates suppressive function of a subset of dn T cells even in the absence of FoxP3. The identification of unique immunoregulatory non-conventional dn T cell subpopulations of the dog in vitro is of high relevance, given the immunotherapeutic potential of manipulating regulatory T cell responses in vivo.

We aimed to analyze the correlations of the expressions of Toll-like receptor 4 (TLR4) and Janus kinase 3 (JAK3) in peripheral blood mononuclear cells (PBMCs) with T helper 17 cell (Th17)/regulatory T cell (Treg) imbalance in patients with ankylosing spondylitis (AS).