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Arthritis Research & Therapy Aug 2019The in vitro pharmacology of baricitinib, upadacitinib, and tofacitinib was evaluated to understand differences among these JAK inhibitors (JAKis) at the cellular level. (Comparative Study)
Comparative Study
BACKGROUND
The in vitro pharmacology of baricitinib, upadacitinib, and tofacitinib was evaluated to understand differences among these JAK inhibitors (JAKis) at the cellular level.
METHODS
Peripheral blood mononuclear cells from healthy donors were incubated with different JAKis, levels of phosphorylated signal transducer and activator of transcription (pSTAT) were measured following cytokine stimulation, and half maximum inhibitory concentration (IC) values were calculated in phenotypically gated leukocyte subpopulations. Therapeutic dose relevance of the in vitro analysis was assessed using calculated mean concentration-time profiles over 24 h obtained from JAKi-treated subjects. Time above IC and average daily percent inhibition of pSTAT formation were calculated for each JAKi, cytokine, and cell type.
RESULTS
Distinct JAKis displayed different in vitro pharmacologic profiles. For example, tofacitinib and upadacitinib were the most potent inhibitors of the JAK1/3-dependent cytokines tested (interleukin [IL]-2, IL-4, IL-15, and IL-21) with lower IC values and increased time above IC translating to a greater overall inhibition of STAT signaling during the dosing interval. All JAKis tested inhibited JAK1/2-dependent cytokines (e.g., IL-6 and interferon [IFN]-γ), the JAK1/tyrosine kinase 2 (TYK2)-dependent cytokines IL-10 and IFN-α, the JAK2/2-dependent cytokines IL-3 and granulocyte-macrophage colony-stimulating factor (GM-CSF), and the JAK2/TYK2-dependent cytokine granulocyte colony-stimulating factor (G-CSF), but often to significantly differing degrees.
CONCLUSIONS
Different JAKis modulated distinct cytokine pathways to varying degrees, and no agent potently or continuously inhibited an individual cytokine signaling pathway throughout the dosing interval. Notably, baricitinib inhibited JAK1/3 signaling to a lesser extent than upadacitinib and tofacitinib, while upadacitinib, baricitinib, and tofacitinib inhibited the signaling of JAK2/2-dependent cytokines, including GM-CSF and IL-3, as well as the signaling of the JAK2/TYK2-dependent cytokine G-CSF.
Topics: Arthritis, Rheumatoid; Azetidines; Biomarkers; Cytokines; Flow Cytometry; Heterocyclic Compounds, 3-Ring; Humans; Janus Kinase Inhibitors; Leukocytes, Mononuclear; Piperidines; Protein Kinase Inhibitors; Purines; Pyrazoles; Pyrimidines; Pyrroles; Signal Transduction; Sulfonamides
PubMed: 31375130
DOI: 10.1186/s13075-019-1964-1 -
Obstetrics and Gynecology Apr 2013To estimate the associations of change in immune response with preterm delivery, omega-3 supplementation, and fish diet. (Randomized Controlled Trial)
Randomized Controlled Trial
OBJECTIVE
To estimate the associations of change in immune response with preterm delivery, omega-3 supplementation, and fish diet.
METHODS
This was an ancillary study to a randomized trial of omega-3 fatty acid supplementation for the prevention of recurrent preterm birth. In vitro maternal peripheral blood mononuclear leukocyte production of the anti-inflammatory cytokine, interleukin-10, and the proinflammatory cytokine, tumor necrosis factor-α, in response to stimulation with lipopolysaccharide, was measured at 16-22 weeks of gestation (baseline) and again at 25-28 weeks of gestation (follow-up) among women with prior spontaneous preterm birth. Changes in concentrations from baseline to follow-up ([INCREMENT]) were compared separately among groups defined by gestational age category at delivery, fish diet history, and omega-3 compared with placebo treatment assignment with Kruskal-Wallis tests.
RESULTS
Interleukin-10 [INCREMENT] differed by gestational age category among 292 women with paired assays. Concentrations increased less in women delivering between 35 and 36 6/7 weeks of gestation (48.9 pg/mL) compared with women delivering at term (159.3 pg/mL) and decreased by 65.2 pg/mL in women delivering before 35 weeks of gestation (P=.01). Tumor necrosis factor-α Δ also differed by gestational age category among 319 women, but the pattern was inconsistent. Those delivering between 35 and 36 6/7 weeks of gestation exhibited decreased concentrations of tumor necrosis factor-α at follow-up compared with baseline (-356.0 pg/mL); concentrations increased among women delivering before 35 weeks of gestation and those delivering at term, 132.1 and 86.9 pg/mL (P=.03). Interleukin-10 Δ and tumor necrosis factor-α Δ were unaffected by either omega-3 supplementation or fish diet.
CONCLUSION
Recurrent preterm birth was associated with decreased peripheral blood mononuclear leukocyte production of interleukin-10 in response to a stimulus during the second trimester.
CLINICAL TRIAL REGISTRATION
ClinicalTrials.gov, www.clinicaltrials.gov, NCT00135902.
LEVEL OF EVIDENCE
II.
Topics: Adult; Animals; Dietary Supplements; Fatty Acids, Omega-3; Female; Fishes; Humans; Interleukin-6; Leukocytes, Mononuclear; Pregnancy; Pregnancy Trimester, Second; Premature Birth; Tumor Necrosis Factor-alpha; Young Adult
PubMed: 23635681
DOI: 10.1097/AOG.0b013e3182878a80 -
Redox Report : Communications in Free... Jul 2015Prolidase plays a major role in collagen turnover, matrix remodeling, and cell growth. Benign prostatic hyperplasia (BPH) may be associated with an increased...
OBJECTIVES
Prolidase plays a major role in collagen turnover, matrix remodeling, and cell growth. Benign prostatic hyperplasia (BPH) may be associated with an increased extracellular matrix deposition. Therefore, the present study was designed to investigate the plasma prolidase activity, oxidative status, and peripheral mononuclear leukocyte DNA damage in patients with BPH.
PATIENTS AND METHODS
Twenty-six male patients with BPH and 24 healthy male subjects were included in this study. Blood samples were collected from antecubital vein after an overnight fasting period, and the plasma was separated. Plasma prolidase activity, total antioxidant capacity (TAC), total oxidant status (TOS), and oxidative stress index (OSI) were determined. The peripheral lymphocyte oxidative DNA damage was determined using an alkaline single cell gel electrophoresis assay (comet assay).
RESULTS
The plasma prolidase activity, TOS levels, OSI values, and peripheral mononuclear leukocyte DNA damage were significantly higher (P < 0.001), while the TAC levels were significantly lower (P < 0.001) in patients with BPH than controls. In BPH patients, the prolidase activity was significantly associated with TAC levels (r = -0.366, P < 0.05), TOS levels (r = 0.573, P < 0.001), and OSI (r = 0.618, P < 0.001) and peripheral mononuclear leukocyte DNA damage (r = 0.461, P < 0.001).
CONCLUSIONS
Our results showed that BPH might be associated with an increased oxidative stress, and also an increased plasma prolidase activity. Increased prolidase activity might play an important role in the etiopathogenesis and/or progression of BPH.
Topics: Antioxidants; Collagen; Comet Assay; DNA Damage; Dipeptidases; Extracellular Matrix; Humans; Leukocytes, Mononuclear; Male; Middle Aged; Oxidants; Oxidative Stress; Prospective Studies; Prostatic Hyperplasia
PubMed: 25551736
DOI: 10.1179/1351000214Y.0000000121 -
American Journal of Nephrology 2012Increased apoptosis along with enhanced inflammation has been reported in hemodialysis and pre-dialysis patients. However, there is limited information at which stage...
BACKGROUND/AIM
Increased apoptosis along with enhanced inflammation has been reported in hemodialysis and pre-dialysis patients. However, there is limited information at which stage during the progression of chronic kidney disease (CKD) the balance between pro- and anti-apoptotic mechanisms is disturbed and inflammatory response is activated. The aim of this study was to investigate possible alterations in apoptotic and inflammatory markers during CKD (stages 1-4) progression and the probable interactions between them.
METHODS
In a cross-sectional study, 152 steady-state CKD outpatients (83 males, 55%) with mean estimated glomerular filtration rate 46 (29-76) ml/min/1.73 m(2) were studied. Apoptosis was assessed in peripheral blood mononuclear cells by estimating Bcl-2 expression, annexin V-propidium iodine staining and serum soluble Fas (sFas) and Fas-ligand. Serum levels of C-reactive protein, tumor necrosis factor-α (TNF-α), interleukin-6 and plasma levels of fibrinogen were measured as markers of inflammation.
RESULTS
Bcl-2 expression was found to decrease significantly in both lymphocytes and monocytes from CKD stage 1 to 4. In contrast, the activity of sFas increased significantly and so did the levels of TNF-α and fibrinogen. The majority of these alterations occurred as soon as patients entered stage 3 of CKD. A multivariate regression analysis demonstrated that CKD remained a significant predictor of the aggregate of the assessed markers.
CONCLUSIONS
Apoptosis appeared to increase across CKD stages 1-4, and this was associated with increased proinflammatory activity.
Topics: Adult; Aged; Aged, 80 and over; Apoptosis; Biomarkers; C-Reactive Protein; Cross-Sectional Studies; Disease Progression; Female; Fibrinogen; Glomerular Filtration Rate; Humans; Inflammation; Interleukin-6; Leukocytes, Mononuclear; Male; Middle Aged; Multivariate Analysis; Proto-Oncogene Proteins c-bcl-2; Renal Insufficiency, Chronic; Severity of Illness Index; Tumor Necrosis Factor-alpha; fas Receptor
PubMed: 23258075
DOI: 10.1159/000345352 -
In Vivo (Athens, Greece) 2022Although taurolidine is known to exert a wide spectrum of biological actions, its effects on immune cells have not been characterized in detail. In this study, we...
BACKGROUND/AIM
Although taurolidine is known to exert a wide spectrum of biological actions, its effects on immune cells have not been characterized in detail. In this study, we investigated the ex vivo effects of taurolidine on relevant innate and adaptive immune cell functions.
MATERIALS AND METHODS
Leukocyte functions in whole blood were assessed following treatment with various taurolidine concentrations. Viability of peripheral blood mononuclear cells (PBMCs) and granulocytes was measured using the WST-1 assay. PBMC function was assessed by measuring TNFα and IFNγ production after stimulation with lipopolysaccharide (LPS) or Candida, respectively. Reactive oxygen species (ROS) production by granulocytes was measured in whole blood using luminol-enhanced chemiluminescence. Granulocyte degranulation and activation were evaluated by membrane expression of degranulation (CD63, CD66B) and adhesion markers (CD62L, CD11b) using immunofluorescent staining followed by flow-cytometric analysis.
RESULTS
Taurolidine decreased viability of PBMCs and granulocytes: after 2 h, IC concentrations were 500 and 520 μg/ml, respectively. Following prolonged exposure (≥24 h) of PBMCs, the IC concentrations declined to 40 μg/ml. PBMC cytokine production significantly decreased at taurolidine concentrations below the cytotoxic threshold, whereas no changes in ROS production were observed. The expression of all granulocyte adhesion and degranulation markers increased at concentrations higher than 500 μg/ml (the cytotoxic level of taurolidine).
CONCLUSION
Taurolidine exhibits a dose- and time-dependent cytotoxicity toward PBMCs and granulocytes. The effects on PBMCs, as exemplified by a decrease in cytokine production, occurred below the toxic threshold, whereas granulocyte function (ROS production) remained unchanged at these taurolidine concentrations. Granulocyte activation and degranulation markers only increased at cytotoxic taurolidine concentrations.
Topics: Anti-Infective Agents, Local; Antineoplastic Agents; Cytokines; Leukocytes, Mononuclear; Reactive Oxygen Species; Taurine; Thiadiazines
PubMed: 36099103
DOI: 10.21873/invivo.12933 -
Frontiers in Immunology 2022Ankylosing spondylitis (AS) is a chronic inflammatory disease with serious consequences and a high rate of morbidity and mortality, In our previous work, we reveal the...
BACKGROUND
Ankylosing spondylitis (AS) is a chronic inflammatory disease with serious consequences and a high rate of morbidity and mortality, In our previous work, we reveal the key features of proteins in new-onset ankylosing spondylitis patients.
MATERIAL AND METHODS
Ankylosing spondylitis (AS) is a chronic inflammatory condition that affects the spine, and inflammation plays an essential role in AS pathogenesis. The inflammatory process in AS, however, is still poorly understood due to its intricacy. Systematic proteomic and phosphorylation analyses of peripheral blood mononuclear cells (PBMCs) were used to investigate potential pathways involved in AS pathogenesis.
RESULTS
Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis was performed and discovered 782 differentially expressed proteins (DEPs) and 122 differentially phosphorylated proteins (DPPs) between 9 new-onset AS patients and 9 healthy controls. The DEPs were further verified using parallel reaction monitoring (PRM) analysis. PRM analysis verified that 3 proteins (HSP90AB1, HSP90AA1 and HSPA8) in the antigen processing and presentation pathway, 6 proteins (including ITPR1, MYLK and STIM1) in the platelet activation pathway and 10 proteins (including MYL12A, MYL9 and ROCK2) in the leukocyte transendothelial migration pathway were highly expressed in the PBMCs of AS patients.
CONCLUSION
The key proteins involved in antigen processing and presentation, platelet activation and leukocyte transendothelial migration revealed abnormal immune regulation in patients with new-onset AS. These proteins might be used as candidate markers for AS diagnosis and new therapeutic targets, as well as elucidating the pathophysiology of AS.
Topics: Chromatography, Liquid; Humans; Leukocytes, Mononuclear; Proteomics; Spondylitis, Ankylosing; Tandem Mass Spectrometry
PubMed: 35371008
DOI: 10.3389/fimmu.2022.838891 -
Clinical Rheumatology Dec 2022To seek significant features of systemic lupus erythematosus (SLE) by utilizing bioinformatics analysis.
INTRODUCTION/OBJECTIVES
To seek significant features of systemic lupus erythematosus (SLE) by utilizing bioinformatics analysis.
METHOD
Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to quantify lysine crotonylation (Kcr) and lysine 2-hydroxyisobutyrylation (Khib) in peripheral blood mononuclear cells (PBMCs) of systemic lupus erythematosus (SLE) patients and normal controls.
RESULTS
Seventy-six differentially modified proteins (DMPs) dually modified by Kcr and Khib were identified between SLE patients and healthy people. GO enrichment analysis prompted significant enrichment of seventy-six DMPs in MHC class II protein complex binding and leukocyte migration. KEGG pathways were enriched in antigen processing and presentation pathway and leukocyte transendothelial migration pathway. Six DMPs (CLTC, HSPA1B, HSPA8, HSP90AB1, HSPD1, and PDIA3) were identified in antigen processing and presentation pathway, of which HSPA8 was the core protein. Significant changes of Kcr and Khib in HSPA8 may increase ATP hydrolysis and promote antigen binding to MHC II molecule. In leukocyte transendothelial migration pathway, 7 DMPs (ACTN1, ACTN4, EZR, MSN, RAC1, RHOA, and VCL) were identified. MSN was the protein with the most modification sites in this pathway. In amino terminal ferm region of MSN, Kcr and Khib expression change may lead to the adhesion between leukocytes and endothelial cells, which was an important step of leukocyte migration.
CONCLUSION
Kcr and Khib may promote the antigen presentation and jointly regulate the tissue damage mediated by leukocyte migration in SLE patients, which may play key roles in the pathogenesis of SLE probably. Key Points • Antigen processing and presentation and leukocyte transendothelial migration may play key roles in the pathogenesis of SLE.
Topics: Humans; Lysine; Chromatography, Liquid; Protein Processing, Post-Translational; Proteomics; Leukocytes, Mononuclear; Endothelial Cells; Tandem Mass Spectrometry; Lupus Erythematosus, Systemic
PubMed: 35941338
DOI: 10.1007/s10067-022-06254-4 -
Veterinary Immunology and... May 2019In mammals, the immune system is undergoing significant changes during development, which has many impacts on the individual's capacity to cope with infectious diseases... (Review)
Review
In mammals, the immune system is undergoing significant changes during development, which has many impacts on the individual's capacity to cope with infectious diseases or other pathologic conditions, where the immune system is involved. Especially in livestock, it is important to know in detail about these changes, including shifts in the composition of systemic leukocyte populations, as this knowledge may help to focus on relevant cell populations when developing novel vaccines for use in juvenile versus adult animals. In this mini-review, a synoptic comparison of published PBMC populations, which were analysed in healthy weaned piglets as well as multiparous non-gestating sows, shows remarkable shifts within leukocyte populations. γδ T cells increase by factor 1.5, plasmacytoid dendritic cells and T helper cells more than double, and cytotoxic T cells as well as regulatory T cells increase more than four fold, whereas NK cells as well as B cells in adult sows comprise only 40% and monocytes 70% of the relative population sizes in weaned piglets. In summary, these insights into age-dependent shifts of porcine leukocyte populations indicate a principal increase of acquired immunity-associated leukocyte populations, whereas primarily innate immunity-associated cell types (NK cells, monocytes) are diminished.
Topics: Aging; Animals; Leukocytes, Mononuclear; Swine
PubMed: 31084891
DOI: 10.1016/j.vetimm.2019.04.002 -
American Journal of Epidemiology May 2013Large numbers of polymorphonuclear leukocytes in the amnion and chorion define histological chorioamnionitis (HCA), a condition linked to spontaneous preterm delivery... (Comparative Study)
Comparative Study
Large numbers of polymorphonuclear leukocytes in the amnion and chorion define histological chorioamnionitis (HCA), a condition linked to spontaneous preterm delivery (PTD). Less is known about placental patterns of mononuclear leukocyte (MNL) density and PTD. In this prospective study (1998-2004), women were sampled from 52 clinics in 5 Michigan communities and enrolled at 16-27 weeks' gestation. HCA and MNL distributions in delivered placentas were evaluated microscopically in a subcohort (290 preterm, 823 term). Midpregnancy biomarkers from maternal blood (i.e., C-reactive protein (CRP), corticotropin-releasing hormone, and cytokines) were compared among term and PTD subjects grouped by presence/absence of HCA and high MNL density. A density of more than 10 MNLs per high-power field in the chorion of the membrane roll, referred to as MNL-CMR, was associated with medically indicated PTD (odds ratio = 2.2, 95% confidence interval: 1.3, 3.6) and spontaneous PTD (odds ratio = 2.5, 95% confidence interval: 1.7, 3.7). Associations persisted after removal of women with HCA-positive placentas, abruption, hypertensive disorders, or obesity. HCA-associated PTD showed higher CRP and cytokine levels. MNL-CMR-associated PTD showed higher CRP and corticotropin-releasing hormone levels. These data suggest that an MNL infiltrate in the chorion of the membrane roll marks PTD pathways that are distinct from HCA and not entirely explained by pregnancy complications.
Topics: Adult; Biomarkers; Chorioamnionitis; Female; Humans; Leukocytes, Mononuclear; Placenta; Pregnancy; Premature Birth; Prospective Studies; Young Adult
PubMed: 23429723
DOI: 10.1093/aje/kws351 -
Cytometry. Part a : the Journal of the... Apr 2023This multiplex staining panel was developed to differentiate cattle T cells into conventional (CD4 and CD8) and unconventional (γδ-TCR) subsets as well as their stage...
This multiplex staining panel was developed to differentiate cattle T cells into conventional (CD4 and CD8) and unconventional (γδ-TCR) subsets as well as their stage of differentiation and activation. The combination of CD45RO and CD62L allows the identification of naïve (T ), central memory (T ), effector memory (T ) and terminal effector (T ) T cells. Activated cattle T cells (T ) can be identified by the cell surface expression of CD25. This panel was developed using cryopreserved cattle peripheral blood mononuclear cells (PBMCs) and tested on fresh as well as stimulated PBMCs. Therefore, this 8-color, 10-parameter flow cytometry panel simultaneously identifies cattle T , T , T , T , T and γδ-TCR cells. This panel will improve our ability to examine T-cell response to pathogens and vaccines in cattle including the potential to identify previously undescribed subpopulations. Furthermore, this panel can be readily optimized for other bovid species as many of these reagents are likely to cross react.
Topics: Cattle; Animals; T-Lymphocytes; Leukocytes, Mononuclear; Leukocyte Common Antigens; Flow Cytometry; Receptors, Antigen, T-Cell; T-Lymphocyte Subsets; Immunologic Memory; CD4-Positive T-Lymphocytes
PubMed: 36734489
DOI: 10.1002/cyto.a.24718