Peripheral Blood Lymphocytes Research Articles

A Nanobody/Monoclonal Antibody "hybrid" sandwich technology offers an improved immunoassay strategy for detection of African trypanosome infections.

The scarcity of reliable devices for diagnosis of Animal African trypanosomiasis (AAT) presents a limitation to control of the disease. Existing high-sensitivity technologies such as PCR are costly, laborious, time-consuming, complex, and require skilled personnel. Hence, utilisation of most diagnostics for AAT is impracticable in rural areas, where the disease occurs. A more accessible point-of-care test (POCT) capable of detecting cryptic active infection, without relying on expensive equipment, would facilitate AAT detection. In turn, early management, would reduce disease incidence and severity. Today, several ongoing research projects aim at modifying complex immunoassays into POCTs. In this context, we report the development of an antigen (Ag) detection sandwich ELISA prototype for diagnosis of T. congolense infections, which is comprised of nanobody (Nb) and monoclonal antibody (mAb) reagents. The Nb474H used here, originated from a past study. Briefly, the Nb was engineered starting from mRNA of peripheral blood lymphocytes of an alpaca immunized with soluble lysate of Trypanosoma congolense (TC13). T. congolense glycosomal fructose-1,6-bisphosphate aldolase (TcoALD) was discovered as the cognate Ag of Nb474H. In this study, splenocytes were harvested from a mouse immunized with recombinant TcoALD and fused with NS01 cells to generate a hybridoma library. Random screening of the library on TcoALD retrieved a lone binder, designated IgM8A2. Using Nb474H as Ag-capture reagent in combination with the IgM8A2 monoclonal antibody Ag-detection reagent resulted in a tool that effectively detects native TcoALD released during infection by T. congolense parasites. Hitherto, development of POCT for detection of active trypanosome infection is elusive. The Nanobody/Monoclonal Antibody (Nb/mAb) "hybrid" sandwich technology offers prospects for exploration, using the unique specificity of Nb as a key determinant in Ag capturing, while using the versatility of monoclonal Ab to adapt to various detection conditions.

Study protocol for a randomised, phase II, double-blind, experimental medicine study of obinutuzumab versus rituximab in ANCA-associated vasculitis: ObiVas

IntroductionRelapses in ANCA-associated vasculitis (AAV) increase the incidence of end-organ damage and their prevention requires prolonged immunosuppressive therapy. Rituximab, a type I anti-CD20 B cell depleting monoclonal antibody, is the...

Peripheral inflammatory T cell subsets are effective predictive factors in the development of heterotopic ossification after posttraumatic elbow surgery

Heterotopic ossification refers to the pathological formation of extra-skeletal bone. It is a common complication of trauma or surgery that can cause disability and has no definitive cure. Furthermore, the mechanisms underlying chronic inflammation during ossification remain unclear. Therefore, this study aimed to elucidate the systemic immune microenvironment status of heterotopic ossification and identify biomarkers of therapeutic efficacy and recurrence. A combination of stereoarthrolysis with prophylactic radiotherapy and non-steroidal anti-inflammatory drugs was used to treat patients with heterotopic ossification. Changes were observed in peripheral blood lymphocyte levels after treatment. The number of IFNγ+CD8+T cells (3.753 % vs 12.90 %, P < 0.0001) and IL17+CD4+T cells (3.420 % vs 5.560 %, P = 0.0281) were was higher in the peripheral blood of relapsed patients with heterotopic ossification than in that of non-relapsed patients. Similarly, the number of these cells was elevated in patients who developed heterotopic ossification after posttraumatic elbow surgery. Peripheral CD8+T cells derived from patients with this pathology promoted osteogenesis through IFNγ expression in vitro. Our findings demonstrate that IFNγ+CD8+T cells and IL17+CD4+T cells are potential biomarkers of heterotopic ossification after posttraumatic elbow surgery. Furthermore, these cells can be used to predict therapeutic efficacy and relapse after combination therapy.

Relationship of cognitive measures to mRNA levels in lymphocytes from patients with schizophrenia and controls

Patients with schizophrenia show substantial cognitive deficits and abnormalities in neurotransmitter-related levels of mRNA in brain or peripheral blood lymphocytes. However, the relationship of cognitive deficits as measured by the MATRICS battery and mRNA levels in brain or lymphocytes has not been sufficiently explored. We measured levels of methylation or neurotransmitter-related mRNAs in lymphocytes of 38 patients with chronic schizophrenia (CSZ) and 33 non-psychotic controls (controls) by qPCR using TaqMan probes. We assessed cognitive function in these patients and controls with the MATRICS battery. We used correlation analysis and scatter plots to assess the relationship of lymphocyte mRNA levels to MATRICS domain and composite scores. CSZ subjects had a consistently negative correlation between mRNA levels in lymphocytes and MATRICS cognitive variables of speed of processing, attention-vigilance, working memory, visual learning, and overall composite score. It is uncertain whether these negative correlations represent a causative relation between specific mRNA levels and cognitive deficits. Controls had either positive correlations or non-significant correlations between mRNA and most of the MATRICS variables. There were statistically significant differences in the correlations between mRNA and MATRICS variables between CSZ vs controls for several mRNAs (DNMT1, DNMT3A, BDNF, NR3C1, FPRF3, CNTNAP2). Our data show a different relationship between mRNA levels in peripheral blood lymphocytes and MATRICS cognitive variables in CSZ vs controls. The substantive significance of these differences needs further investigation.

Association between immune-inflammation-based prognostic index and depression: An exploratory cross-sectional analysis of NHANES data

BackgroundImmune-inflammatory mediators influence numerous immune and inflammatory pathways, elevating the likelihood of depression. The systemic immune-inflammation index (SII) emerges as an innovative prognostic indicator, integrating various peripheral blood immune cell subpopulations, specifically neutrophils, platelets, and lymphocytes. This exploratory study aims to examine the correlation between SII and depression. MethodsData from the 2005–2020 National Health and Nutrition Examination Survey (NHANES) were utilized. Depression was diagnosed with a Patient Health Questionnaire score of 10 or higher. The relationship between log2-SII and depression incidence was analyzed using a restricted cubic spline (RCS). Logistic regression was employed to calculate the odds ratio of depression concerning log2-SII. In cases of non-linearity, piecewise linear models with change points were applied to assess the associations in both the overall population and specific subgroups. Additionally, subgroup analyses were conducted to determine the applicability of the findings to particular populations. ResultsA total of 42,133 participants were included in the study, comprising 49.32 % men and 50.68 % women, with an average age of 47.02 ± 17.45 years. RCS analysis demonstrated a J-shaped non-linear relationship between log2-SII and depression incidence. When log2-SII was ≥8.50, SII showed a positive association with depression incidence, even after adjusting for covariates. Additionally, each unit increase in log2-SII corresponded to an 18 % rise in depression incidence (OR = 1.18, 95 % CI: 1.10–1.27). Subgroup analysis further revealed that the association between SII and depression incidence varied across different populations. LimitationsDue to the cross-sectional nature of NHANES, causality or long-term implications cannot be inferred. Further research is needed to ascertain if a longitudinal relationship exists between SII and depression. ConclusionOur findings suggest a significant and complex non-linear association between SII and depression. However, further basic and prospective studies are necessary to explore SII's impact on depression and clarify its underlying mechanisms. Additionally, these studies will provide a foundation for personalized interventions targeting the immune-inflammatory processes in patients with depression and elevated SII.

Prevalence of karyotype alterations in couples with recurrent pregnancy loss in a tertiary center in Brazil.

To assess the prevalence and type of chromosomal abnormalities in Brazilian couples with recurrent pregnancy loss (RPL) and compare the clinical characteristics of couples with and without chromosome abnormalities. We assessed the medical records of 127 couples with a history of two or more miscarriages, referred to a tertiary academic hospital in Belo Horizonte, Brazil, from January 2014 to May 2023. Karyotype was generated from peripheral blood lymphocyte cultures, and cytogenetic analysis was performed according to standard protocols by heat-denatured Giemsa (RHG) banding. Abnormal karyotypes were detected in 10 couples (7.8%). The prevalence of chromosomal abnormalities was higher among females (6.3%) compared to males (2.0%), but this difference was not statistically significant (p=0.192). The mean number of miscarriages was. 3.3 ± 1.1 in couples with chromosome abnormalities and 3.1 ± 1.5 in couples without chromosome abnormalities (p=0.681). Numerical chromosomal anomalies (6 cases) were more frequent than structural anomalies. Four women presented low-grade Turner mosaicism. No differences were found between couples with and without karyotype alterations, except for maternal age, which was higher in the group with chromosome alterations. The prevalence of parental chromosomal alterations in our study was higher than in most series described in the literature and was associated with increased maternal age. These findings suggest that karyotyping should be part of the investigation for Brazilian couples with RPL, as identifying the genetic etiology may have implications for subsequent pregnancies.

Peripheral Th17/Treg imbalance in Chinese patients with untreated antisynthetase syndrome associated interstitial lung disease

Interstitial lung disease (ILD) is a common and fatal manifestation of antisynthetase syndrome (ASS). The aim of this study was to provide new insight into investigate peripheral blood lymphocytes, CD4+ T cells, cytokine levels and their relation to the clinical profile of untreated patients with ASS-ILD. The retrospective study population included thirty patients diagnosed with ASS-ILD and 30 healthy controls (HCs). Baseline clinical and laboratory data were collected for all subjects, including peripheral blood lymphocyte, CD4+ T cell subsets measured by flow cytometry, and serum cytokine levels measured by multiple microsphere flow immunofluorescence. Their correlations with clinical and laboratory findings were analyzed by Pearson’s or Spearman’s correlation analysis. In addition, the Benjamini-Hochberg method was used for multiple correction to adjust the p-values. Patients with ASS-ILD had lower CD8+ T cells, higher proportion of Th17 cells and Th17/Treg ratio than HCs. Serum cytokine levels (IL-1β, IL-6, IL-12, IL-17, IL-8, IL-2, IL-4, IL-10, TNF-α and IFN-γ) were higher in patients with ASS-ILD than HCs. Moreover, Th17/Treg ratio was negatively correlated with diffusing capacity of carbon monoxide (DLCO)%. Our study demonstrated abnormalities of immune disturbances in patients with ASS-ILD, characterized by decreased CD8+ T cells and an increased Th17/Treg ratio, due to an increase in the Th17 cells. These abnormalities may be the immunological mechanism underlying the development of ILD in ASS.

Short-term exposure to air pollution on peripheral white blood cells and inflammation biomarkers: a cross-sectional study on rural residents

Effects of short-term exposure to ambient air pollution on systemic immunological and inflammatory biomarkers in rural population have not been adequately characterized. From May to July 2021, 5816 participants in rural villages of northern Henan Province, China, participated in this cross-sectional study. Blood biomarkers of systemic inflammation were determined including peripheral white blood cells (WBC), eosinophils (EOS), basophils (BAS), monocytes (MON), lymphocytes (LYM), neutrophils (NEU), neutrophil-lymphocyte ratio (NLR), and serum high-sensitivity C-reactive protein (hs-CRP). The concentrations of ambient fine particulate matter (PM2.5), PM10, nitrogen dioxide (NO2), carbon monoxide (CO), and ozone (O3) were assessed up to 7 days prior to the blood draw. A generalized linear model was used to analyze the associations between air pollution exposure and the above-mentioned blood biomarkers. Significantly positive associations were revealed between PM2.5, CO and WBC; CO, O3 and LYM; PM2.5, PM10, SO2, CO and NEU; PM2.5, PM10, SO2, CO and NLR; PM2.5, PM10, SO2, NO2, CO, O3 and hs-CRP. Meanwhile, negative associations were found between SO2 and WBC; PM2.5, PM10, NO2, CO, or O3 and EOS; PM2.5, SO2, or CO and BAS; SO2, NO2 or O3 and MON; PM2.5, PM10, SO2, or NO2 and LYM. Moreover, men, individuals with normal body mass index (BMI), current smokers, and those older than 60 years were found vulnerable to air pollution effects. Taken together, short-term exposure to air pollution was associated with systemic inflammatory responses, providing insight into the potential mechanisms for air pollution-induced detrimental systemic effects in rural residents.

9-cis-retinoic acid signaling in Sertoli cells regulates their immunomodulatory function to control lymphocyte physiology and Treg differentiation

BackgroundTestis is an immune privileged organ, which prevents the immune response against sperm antigens and inflammation. Testicular cells responsible for immune tolerance are mainly Sertoli cells, which form the blood-testis barrier and produce immunosuppressive factors. Sertoli cells prevent inflammation in the testis and maintain immune tolerance by inhibiting proliferation and inducing lymphocyte apoptosis. It has been shown that 9-cis-retinoic acid (9cRA) blocks ex vivo apoptosis of peripheral blood lymphocytes and promotes the differentiation of Treg cells in the gut. However, the role of retinoid signaling in regulating the immune privilege of the testes remains unknown.ObjectiveThe aim of this study was to determine whether 9cRA, acting via the retinoic acid receptors (RAR) and the retinoic X receptors (RXR), controls the immunomodulatory functions of Sertoli cells by influencing the secretion of anti-inflammatory/pro-inflammatory factors, lymphocyte physiology and Treg cell differentiation.MethodsExperiments were performed using in vitro model of co-cultures of murine Sertoli cells and T lymphocytes. Agonists and antagonists of retinoic acid receptors were used to inhibit/stimulate retinoid signaling in Sertoli cells.ResultsOur results have demonstrated that 9cRA inhibits the expression of immunosuppressive genes and enhances the expression of pro-inflammatory factors in Sertoli cells and lymphocytes, increases lymphocyte viability and decreases apoptosis rate. Moreover, we have found that 9cRA blocks lymphocyte apoptosis acting through both RAR and RXR and inhibiting FasL/Fas/Caspase 8 and Bax/Bcl-2/Caspase 9 pathways. Finally, we have shown that 9cRA signaling in Sertoli cells inhibits Treg differentiation.ConclusionCollectively, our results indicate that retinoid signaling negatively regulates immunologically privileged functions of Sertoli cells, crucial for ensuring male fertility. 9cRA inhibits lymphocyte apoptosis, which can be related to the development of autoimmunity, inflammation, and, in consequence, infertility.

Description of lymphocyte and cytokine profiles in individuals with acute myeloid leukemia associated with FLT3-ITD and NPM1 mutation status.

The pathogenesis of acute myeloid leukemia (AML) involves mutations in genes such as FLT3 and NPM1, which are also associated with the prognosis of the disease. The immune system influences disease progression, but the mechanisms underlying the interaction between the immune system and AML are not clear. In this study, the profiles of lymphocytes and cytokines were described in individuals with AML stratified by molecular changes associated with prognosis. The participants included in this study were newly diagnosed AML patients (n = 43) who were about to undergo chemotherapy. Subtypes of lymphocytes in peripheral blood, including B cells, T cells, and natural killer cells, and serum concentrations of cytokines, including Th1, Th2, and Th17, were studied by flow cytometry assays (BD FACSCanto II). The correlations between lymphocyte subsets, cytokines, and genetic/prognostic risk stratification (based on the FLT3 and NPM1 genes) were analyzed. The differences in B lymphocytes (%), T lymphocytes (%), plasmablasts (%), leukocytes (cells/µl), and tumor necrosis factor (pg/ml) were determined between groups with FLT3-ITD+ and FLT3-ITD- mutations. The presence of mutations in NPM1 and FLT3-ITD and age suggested changes in the lymphocyte and cytokine profile in individuals with AML.

A Broad-specificity Neutralizing Nanobody against Hepatitis E Virus Capsid Protein.

Hepatitis E virus (HEV) is a worldwide zoonotic and public health concern. The study of HEV biology is helpful for designing viral vaccines and drugs. Nanobodies have recently been considered appealing materials for viral biological research. In this study, a Bactrian camel was immunized with capsid proteins from different genotypes (1, 3, 4, and avian) of HEV. Then, a phage library (6.3 × 108 individual clones) was constructed using peripheral blood lymphocytes from the immunized camel, and 12 nanobodies against the truncated capsid protein of genotype 3 HEV (g3-p239) were screened. g3-p239-Nb55 can cross-react with different genotypes of HEV and block Kernow-C1/P6 HEV from infecting HepG2/C3A cells. To our knowledge, the epitope recognized by g3-p239-Nb55 was determined to be a novel conformational epitope located on the surface of viral particles and highly conserved among different mammalian HEV isolates. Next, to increase the affinity and half-life of the nanobody, it was displayed on the surface of ferritin, which can self-assemble into a 24-subunit nanocage, namely, fenobody-55. The affinities of fenobody-55 to g3-p239 were ∼20 times greater than those of g3-p239-Nb55. In addition, the half-life of fenobody-55 was nine times greater than that of g3-p239-Nb55. G3-p239-Nb55 and fenobody-55 can block p239 attachment and Kernow-C1/P6 infection of HepG2/C3A cells. Fenobody-55 can completely neutralize HEV infection in rabbits when it is preincubated with nonenveloped HEV particles. Our study reported a case in which a nanobody neutralized HEV infection by preincubation, identified a (to our knowledge) novel and conserved conformational epitope of HEV, and provided new material for researching HEV biology.

The PD-1/PD-L1 Gateway: Peripheral Immune Regulation in the Pathogenesis of Endometriosis.

Endometriosis is a chronic inflammatory disease characterized by the presence of endometrial-like tissue outside the uterine cavity, causing pain and infertility. Despite the rather unclear etiopathogenesis, recent studies suggest the involvement of the immune system in the development and progression of endometriosis. The role of the PD-1/PD-L1 axis in the modulation of the immune response in this disease seems to be particularly interesting. This preliminary study aimed to investigate the expression of PD-1 and PD-L1 on T and B lymphocytes in peripheral blood in patients with endometriosis to assess their potential impact on disease progression. Our study involved peripheral blood samples from 80 patients diagnosed with endometriosis and 20 healthy women as a control group were analyzed. Flow cytometry was used to assess the expression of PD-1 and PD-L1 on T and B lymphocytes, and enzyme-linked immunosorbent assays were used to assess their soluble forms in serum and peritoneal fluid.in our research we observe significantly higher expression of PD-1 and PD-L1 on T and B lymphocytes was found in patients with endometriosis compared to the control group. Higher expression of both tested molecules correlated with the stage of endometriosis. The results of our preliminary studies indicate a potential role of the PD-1/PD-L1 axis in the modulation of the immune response in endometriosis. Modified expression of these proteins may contribute to immune evasion by ectopic tissues, supporting their survival and proliferation. These findings suggest that targeting PD-1/PD-L1 could be explored as a therapeutic option for the treatment of endometriosis, though further research with larger sample sizes is necessary to confirm these results and clarify the role of PD-1/PD-L1 in the pathogenesis of the disease.

Clinical characteristics, immunological alteration and distinction of MOG-IgG-associated disorders and GFAP-IgG-associated disorders

The classification of autoimmune encephalitis (AE) is based on the presence of different types of antibodies. Currently, the clinical manifestations and treatment regimens of patients with all types of AE exhibit similarities. However, the presence of immunological distinctions among different types of AE remains uncertain. In this study, we prospectively collected clinical data, as well as blood and cerebrospinal fluid (CSF) samples from patients diagnosed with MOG antibody-associated disease (MOGAD) or GFAP astrocytopathy (GFAP-A), in order to assess changes in inflammatory biomarkers such as immunoglobulin oligoclonal bands, cytokines in serum and CSF, as well as peripheral blood lymphocyte subtypes within different subsets. To further distinguish the immune response in patients with MOGAD and GFAP-A from that of healthy individuals, we prospectively recruited 20 hospitalized patients diagnosed with AE. Among them, 15 (75%) tested positive for MOG antibodies, 4 (20%) tested positive for GFAP antibodies, and 1 (5%) tested positive for both MOG and GFAP antibodies. These patients were then followed up for a period of 18 months. Compared to healthy controls (HC), AE patients exhibited elevated levels of MIP-1beta, SDF-1alpha, IL-12p70, IL-5, IL-1RA, IL-8 and decreased levels of IL-23, IL-31, IFN-alpha, IL-7, TNF-beta and TNF-alpha in serum. The CSF of AE patients showed increased levels of IL-1RA, IL-6 and IL-2 while decreased levels of RANTES, IL-18,IL-7,TNF-beta,TNF-alpha,RANTES,Eotaxin,and IL-9. The level of MCP-1 in the CSF of GFAP-A patients was found to be lower compared to that of MOGAD patients, while RANTES levels were higher. And the levels of IL-17A, Eotaxin, GRO-alpha, IL-8, IL-1beta, MIP-1beta were higher in the CSF of patients with epilepsy. The presence of intrathecal immune responses is also observed in patients with spinal muscular atrophy (SMA). However, no biomarker was found to be associated with disease severity in patients with AE. Among the 17 patients, recovery was observed, while 2 patients experienced persistent symptoms after an 18-month follow-up period. Additionally, within one year of onset, 8 patients had a single recurrence. Therefore, the immunological profiles of MOGAD and GFAP-A patients differ from those of normal individuals, and the alterations in cytokine levels may also exhibit a causal association with the clinical presentations, such as seizure.

CD3+/CD4+ cells combined with myosteatosis predict the prognosis in patients who underwent gastric cancer surgery.

This study aimed to investigate the predictive capacity of lymphocyte subpopulations, sarcopenia and myosteatosis for clinical outcomes in patients who underwent gastric cancer surgery. Additionally, the prognostic significance of CD3+/CD4+ cells in conjunction with myosteatosis was explored. A cohort of 190 patients with gastric cancer who underwent surgery and received computed tomography scans between July 2016 and December 2017 at our institution was examined. Complete clinical information and peripheral lymphocyte subpopulations were available for all patients. A comprehensive array of statistical methodologies was employed to scrutinize variances in both clinical and pathological characteristics among patients, with the aim of identifying autonomous prognostic determinants requisite for the development of a nomogram. Subsequent assessment of the predictive efficacy of the nomogram was conducted via calibration curve analysis. The study comprised a cohort of 190 participants, encompassing 126 males (66.32%) and 64 females (33.68%), with a mean age of 58.47 (±11.37) years. Patients were stratified into three groups based on CD3+/CD4+ cells and myosteatosis, with 24 in Group 1, 87 in Group 2 and 79 in Group 3. Notably, patients in the third group exhibited significantly shorter progression-free survival (PFS) (hazard ratio [HR]=0.208, P<0.001) and overall survival (OS) (HR=0.193, P<0.001). The subset of peripheral blood lymphocytes exhibited elevated levels of CD3+/CD4+ cells (HR=2.485, P<0.001) and heightened CD4+/CD8+ ratios (HR=1.705, P=0.038), whereas diminished CD19+ cell counts (HR=0.210, P=0.032) correlated with improved OS in patients. The individuals presenting with sarcopenia (HR=4.089, P=0.023) and myosteatosis (HR=2.857, P<0.001) displayed reduced OS. The multivariate Cox regression analysis showed that pathological tumour-node-metastasis stage, CD19+ cells, sarcopenia and CD3+/CD4+ cell-myosteatosis were identified as independent prognostic factors for PFS and OS in patients. The constructed nomograms for PFS and OS yielded C-index values of 0.839 (95% confidence interval [CI]: 0.798-0.880) and 0.836 (95% CI: 0.792-0.879), respectively. The calibration analysis demonstrated that the nomograms accurately predicted the 3- and 5-year survival rates of PFS and OS in patients. Lymphocyte subsets, including CD3+/CD4+ cells, CD4+/CD8+ ratio and CD19+ cells, are indicative of clinical prognosis in gastric cancer surgery patients. Body composition parameters, such as sarcopenia and myosteatosis, are also associated with the patient's prognosis. The combination of CD3+/CD4+ cells with myosteatosis demonstrates enhanced prognostic value, enabling the identification of patients at high risk of post-operative metastasis and recurrence.

Paroxetine alleviates dendritic cell and T lymphocyte activation via GRK2-mediated PI3K-AKT signaling in rheumatoid arthritis.

G protein-coupled receptor kinase 2 (GRK2) could participate in the regulation of diverse cells via interacting with non-G-protein-coupled receptors. In the present work, we explored how paroxetine, a GRK2 inhibitor, modulates the differentiation and activation of immune cells in rheumatoid arthritis (RA). The blood samples of healthy individuals and RA patients were collected between July 2021 and March 2022 from the First Affiliated Hospital of Anhui Medical University. C57BL/6 mice were used to induce the collagen-induced arthritis (CIA) model. Flow cytometry analysis was used to characterize the differentiation and function of dendritic cells (DCs)/T cells. Co-immunoprecipitation was used to explore the specific molecular mechanism. In patients with RA, high expression of GRK2 in peripheral blood lymphocytes, accompanied by the increases of phosphatidylinositol 3 kinase (PI3K), protein kinase B (AKT), and mammalian target of rapamycin (mTOR). In animal model, a decrease in regulatory T cells (Tregs), an increase in the cluster of differentiation 8 positive (CD8+) T cells, and maturation of DCs were observed. Paroxetine, when used in vitro and in CIA mice, restrained the maturation of DCs and the differentiation of CD8+ T cells, and induced the proportion of Tregs. Paroxetine inhibited the secretion of pro-inflammatory cytokines, the expression of C-C motif chemokine receptor 7 in DCs and T cells. Simultaneously, paroxetine upregulated the expression of programmed death ligand 1, and anti-inflammatory cytokines. Additionally, paroxetine inhibited the PI3K-AKT-mTOR metabolic pathway in both DCs and T cells. This was associated with a reduction in mitochondrial membrane potential and changes in the utilization of glucose and lipids, particularly in DCs. Paroxetine reversed PI3K-AKT pathway activation induced by 740 Y-P (a PI3K agonist) through inhibiting the interaction between GRK2 and PI3K in DCs and T cells. Paroxetine exerts an immunosuppressive effect by targeting GRK2, which subsequently inhibits the metabolism-related PI3K-AKT-mTOR pathway of DCs and T cell in RA.

Peripheral Blood CD8+ T-Lymphocyte Immune Response in Benign and Subpopulations of Breast Cancer Patients.

Peripheral blood CD8+ T lymphocytes play a crucial role in cell-mediated immunity and tumor-related immune responses in breast cancer. In this study, label-free quantification analysis and gene set enrichment analysis (GSEA) of CD8+ T lymphocytes in the peripheral blood of benign patients and patients with different breast cancer (BC) subtypes, i.e., luminal A, luminal B, and triple-negative breast cancer (TNBC), were performed using nano-UHPLC and Orbitrap mass spectrometry. Differential protein expression in CD8+ T lymphocytes revealed significant downregulation (log2 FC ≥ 0.38 or ≤-0.38, adj. p < 0.05), particularly in proteins involved in cytotoxicity, cytolysis, and proteolysis, such as granzymes (GZMs) and perforin 1 (PRF1). This downregulation was observed in the benign group (GZMH, GZMM, and PRF1) and luminal B (GZMA, GZMH) subtypes, whereas granzyme K (GZMK) was upregulated in TNBC in comparison to healthy controls. The RNA degradation pathway was significantly downregulated (p < 0.05, normalized enrichment score (NES) from -1.47 to -1.80) across all BC subtypes, suggesting a potential mechanism for regulating gene expression during T cell activation. Also, the Sm-like proteins (LSM2, LSM3, and LSM5) were significantly downregulated in the RNA degradation pathway. Proteomic analysis of CD8+ T lymphocytes in peripheral blood across different breast cancer subtypes provides a comprehensive view of the molecular mechanisms of the systemic immune response that can significantly contribute to advancements in the diagnosis, treatment, and prognosis of this disease.

Exposure assessment and cytogenetic biomonitoring study of workers occupationally exposed to extremely low-frequency magnetic fields.

Human cytogenetic biomonitoring (HCB) has long been used to evaluate the potential effects of work environments on the DNA integrity of workers. However, HCB studies on the genotoxic effects of occupational exposure to extremely low-frequency electromagnetic fields (ELF-MFs) were limited by the quality of the exposure assessment. More specifically, concerns were raised regarding the method of exposure assessment, the selection of exposure metrics, and the definition of exposure group. In this study, genotoxic effects of occupational exposure to ELF-MFs were assessed on peripheral blood lymphocytes of 88 workers from the electrical sector using the comet and cytokinesis-block micronucleus assay, considering workers' actual exposure over three consecutive days. Different methods were applied to define exposure groups. Overall, the summarized ELF-MF data indicated a low exposure level in the whole study population. It also showed that relying solely on job titles might misclassify 12 workers into exposure groups. We proposed combining hierarchical agglomerative clustering on personal exposure data and job titles to define exposure groups. The final results showed that occupational MF exposure did not significantly induce more genetic damage. Other factors such as age or past smoking rather than ELF-MF exposure could affect the cytogenetic test outcomes.

GENETIC ABLATION OF THE C-TYPE LECTIN RECEPTOR CLEC2D INCREASES PERITONITIS MORTALITY, INFLAMMATION, AND PHYSIOLOGY WITHOUT DIMINISHING ORGAN INJURY.

Background: Sepsis accounts for substantial morbidity and mortality motivating investigators to continue the search for pathways and molecules driving the pathogenesis of the disease. The current study examined if the novel C-type lectin receptor (CLR), Clec2d, plays a significant role in the pathogenesis of sepsis. Methods: Clec2d knockout (KO) mice were fully backcrossed onto the C57/BL6 background. Acute endotoxemia was induced with an intraperitoneal injection of lipopolysaccharide (LPS). Sepsis was induced in two different models, cecal ligation and puncture (CLP) and Pseudomonas aeruginosa pneumonia. Both models were treated with antibiotics and fluid resuscitation. In the sepsis models, physiologic and hematologic measurements were measured at 24 h by collecting a small sample of peripheral blood. Mortality was followed for 14 days. Results : A total of 197 mice were studied, 58 wild type (WT) and 54 knock-out (KO) in the LPS model; 27 wild type and 21 KO mice in the CLP model; and 22 WT and 15 KO mice in the pneumonia model. Clec2d KO mice had greater mortality in the LPS and CLP studies but not the pneumonia model. There were significant differences in multiple parameters determined 24 h post sepsis between mice who subsequently died and those lived. Consistent with previous reports in the CLP model, higher concentrations of IL-6, increased numbers of peripheral blood lymphocytes and greater renal injury were found in the dying mice. In contrast, in the pneumonia model, IL-6 was higher in the surviving mice; however, the IL-6 levels in the pneumonia model (0.6 ± 0.3 ng/mL mean ± SEM) were less than 2% of the IL-6 levels of mice that died in the CLP model (41 ± 9 ng/mL, mean ± SEM). There were no differences in the lymphocyte count or renal injury between living and dying mice in the pneumonia model. In both sepsis models, dying mice had lower heart rates, respiratory rates, and body temperatures. These values were also lower in the KO mice compared to the WT in CLP, but the breath rate and body temperature were increased in the KO pneumonia mice. Conclusion: The C-type lectin receptor Clec2d plays a complicated role in the pathogenesis of sepsis, which varies with source of infection as demonstrated in the models used to study the disease. These data highlight the heterogeneity of the responses to sepsis and provide further evidence that a single common pathway driving sepsis organ injury and death likely does not exist.

Genotoxic effects of ablative treatment with I-131 determined through the analysis of dicentric chromosomes in peripheral blood lymphocytes. Can it influence the clinical management of patients with differentiated thyroid carcinoma?

Study including 61 differentiated thyroid cancer (DTC) patients who received ablative doses of I-131 after total thyroidectomy. Biodosimetry was performed by dicentric chromosomes analysis before and after treatment, determining dicentric yield and blood absorbed dose. An increased yield of dicentrics after treatment was demonstrated. With an average administered activity of 4.32 ± 0.879 GBq, the absorbed dose in bone marrow was 0.28 Gy (0.25–0.30 95% CI). These data were compared with the administered I-131 activity and body mass index. Increased genotoxic effects were observed, with a slightly higher dicentric yield rising along I-131 activity. No correlation was found between administered activity and BMI. No significant risk of myelotoxicity due to dose regimens of I-131 intended for ablative purposes in “de novo” treated patients was found among the ranges of activities used in clinical practice. Biodosimetric data was included in patients’ Clinical History, which translates into a more detailed dosimetric report. With this information, physicians can take a more patient-specific approach. Personalized administration would be of special consideration in the event of re-treatments, be it due to recurrent disease or I-131 refractory DTC patients.

Monoclonal antibodies for equine CD25 improve detection of regulatory T cells in horses

CD25, the interleukin-2 receptor α-chain, is expressed on cell surfaces of different immune cells and is commonly used for phenotyping of regulatory T cells (Tregs). CD25 has essential roles in the maintenance of hemostasis and immune tolerance and Treg cell involvement has been shown in human diseases and murine models for allergy, autoimmunity, cancer, chronic inflammation, and many others. In horses, a cross-reactive anti-human CD25 antibody has previously been used for characterizing Tregs. Here, we developed monoclonal antibodies (mAbs) to equine CD25 and compared their staining pattern with the anti-human CD25 antibody by flow cytometry. The comparison of the two reagents was performed by two separate analyses in independent laboratories. Overall, similar staining patterns for equine peripheral blood lymphocytes were obtained with the anti-human CD25 antibody and equine CD25 mAb 15–1 in both laboratories. Both reagents identified comparable CD4+CD25+ and CD4+CD25+FOXP3+ percentages after stimulation of peripheral blood mononuclear cells (PBMC) with pokeweed mitogen. However, when compared to the anti-human CD25 antibody, the equine CD25 mAb 15–1 resulted in a better staining intensity of the equine CD25+ cells and increased the percentages of Tregs and other CD25+ cells ex vivo and after culturing of PBMC without stimulation. In summary, the equine CD25 mAbs provide new, improved reagents for Tregs and CD25+ cell phenotyping in horses.