scholarly journals FRI0015 PHENOTYPE AND FUNCTION OF THE PERIPHERAL BLOOD DENDRITIC CELLS OF PSORIASIS PATIENTS WITH AND WITHOUT ARTHRITIS

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 578.1-579
Author(s):  
S. Schnitte ◽  
A. Fuchs ◽  
T. Funk ◽  
A. C. Pecher ◽  
D. Dörfel ◽  
...  
Keyword(s):  
Risk Factors ◽  
Flow Cytometry ◽  
Dendritic Cells ◽  
Psoriatic Arthritis ◽  
T Lymphocytes ◽  
Peripheral Blood ◽  
Surface Proteins ◽  
Research Support ◽  
Cross Sectional ◽  
Cell Counts

Background:Psoriasis is a frequent skin disease that can appear with an arthritic manifestation in approximately 30% of the cases [1]. The underlying excessive immune reaction caused by pro-inflammatory cytokines can be triggered by several risk factors [2]. Various subgroups of Dendritic cells (DCs) in the skin play a crucial role in the induction of the dermal inflammatory response [3].Objectives:As the role of peripheral blood DCs remains unknown and the cause of an arthritic manifestation is still not completely understood [4], this project aimed to detect differences in phenotype or function of peripheral blood DCs in psoriatic patients with or without arthritis.Methods:We analyzed peripheral blood cells of 60 psoriasis patients with and without arthritis. Different DC subpopulations were detected by flow cytometry. Monocyte-derived DCs were cultured with or without Lipopolysaccharides to gain immature (iDC) and mature (mDC) cells. The DC phenotype was determined by staining with CD80, CD83, CD86, CD206, CCR7, CD1a, HLA-DR, CD40, GPN-MB, DC209 and CD14. Their T-cell stimulatory capability was analyzed by co-incubation with Carboxyfluorescein succinimidyl ester stained lymphocytes and the quantification of CD4+ T-lymphocytes afterwards. To measure the migration capacity DCs were seated into transwell chambers with a semipermeable membrane and partly supplemented with Macrophage Inflammatory Protein 3 Beta (Mip3b). Migrated cells were detected by flow cytometry. Measured cell counts were normalized to cell counts without Mip3b stimulation.Results:Comparing the factor of increase of migrated mDC counts due to mip3b stimulation, we detected a significant lower rate in samples of patients with arthritis (PsA) compared to those of patients without (Ps). Assays of mDCs without mip3b stimulation showed a significant higher count of migrated cells in the samples of the arthritic group [Figure 1]. Cell counts with Mip3b stimulation did vary slightly in the groups. The DC subpopulations and the expression of analyzed cell surface proteins did not show significant differences. The amounts of stimulated T-Lymphocytes did not differ significantly.Figure 1.Migration essay showing mDCs following Mip3b (+miß3b) as multiples of mDCs without stimulation (-mip3b). The factor of increase is significantly lower in patients with arthritis (PsA) compared to patients without (Ps). Absolute counts of migrated mDCs without Mip3b are significantly higher in the arthritic group. Cell counts with stimulation do not differ significantly (data not shown). N=24, p<0.05Conclusion:CCL19 (Mip3b) is a potent ligand to the CCR7 receptor inducing migration of DCs towards the lymphatic node [5]. The CCR7 amounts on the DC surface did not differ significantly in the groups. The mDCs without CCL19 stimulation migrated in higher amounts in samples of arthritic patients. Cell counts of stimulated DCs showed only slight differences. These results could be generated by a different appearance of the DCs of arthritic patients that might facilitate migration. Further experiments focusing on this aspect should be performed. A possible effect of disruptive factors (age, sex, medication…) needs to be clarified.References:[1]Henes, J.C., et al.,High prevalence of psoriatic arthritis in dermatological patients with psoriasis: a cross-sectional study.Rheumatol Int, 2014.34(2): p. 227-34.[2]Lee, E.B., et al.,Psoriasis risk factors and triggers.Cutis, 2018.102(5s): p. 18-20.[3]Kim, T.G., S.H. Kim, and M.G. Lee,The Origin of Skin Dendritic Cell Network and Its Role in Psoriasis.Int J Mol Sci, 2017.19(1).[4]Veale, D.J. and U. Fearon,The pathogenesis of psoriatic arthritis.Lancet, 2018.391(10136): p. 2273-2284.[5]Ricart, B.G., et al.,Dendritic cells distinguish individual chemokine signals through CCR7 and CXCR4.J Immunol, 2011.186(1): p. 53-61.Acknowledgments:This project was financially supported by Novartis Pharma GmbH.Disclosure of Interests:Sarah Schnitte Grant/research support from: Reaserch grant by Novartis, Alexander Fuchs: None declared, Tanja Funk: None declared, Ann-Christin Pecher: None declared, Daniela Dörfel: None declared, Jörg Henes Grant/research support from: Novartis, Roche-Chugai, Consultant of: Novartis, Roche, Celgene, Pfizer, Abbvie, Sanofi, Boehringer-Ingelheim,

scholarly journals Early Immune Profile Assessment By Flow Cytometry Predicts Severe COVID-19 Pneumonia

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4197-4197
Author(s):  
Abora Rial-Villavecchia ◽  
Maria Teresa Asensi ◽  
Carme Giménez-Argente ◽  
Joan Gómez-Junyent ◽  
Juan Jose Rodriguez-Sevilla ◽  
...  
Keyword(s):  
Risk Factors ◽  
Flow Cytometry ◽  
Dendritic Cells ◽  
Immunosuppressive Therapy ◽  
Peripheral Blood ◽  
Multivariable Analysis ◽  
High Flow ◽  
Respiratory Status ◽  
Group 1

Abstract Background and Objective The rapid spread of the COVID -19 pandemic and the high variability of the course of the disease make it essential to search for early predictors of outcome. The objective of our study is to predict severe SARS COV2 pneumonia using early cytometric profiles Material and Methods Prospective and observational study of adults with confirmed COVID-19 infection admitted on Emergency Department (ED). We collected epidemiological, clinical and laboratory data of every patient until they were discharged or died. Multiparametric flow cytometry (FC) analysis of T-lymphocytes (CD4, CD8, CD4 activated, CD8 activated, naïve (Tn), central-memory (Tcm), effector-memory (Tem), effector (Te) and Th17 subsets), B-lymphocytes (naïve, memory, transitional subsets, and assessment of clonality), NK cells, plasmablasts, p-DCs (plasmacytoid dendritic cells), m-DCs (myeloid dendritic cells), basophils, and monocytes (MO1, MO2, MO3, slan+ MO3) was performed on whole peripheral blood collected on EDTA, before immunosuppressive therapy was started. We designed a 7-tube 8-color experimental panel. Cell surface staining of 2 × 106 cells was performed and at least 500 000 total events were acquired for the assessment of plasmablasts, p-DCs, m-DCs, basophils, and the monocyte subsets; for the study of B, T and NK-lymphocyte populations we acquired at least 100 000 total events (FACS Canto II; BD Biosciences). Severity was assessed on the basis of World Health Organization´s (WHO) international 10 level ordinal scale (WHOs) and also according to 4 respiratory status, based on SpO2(peripheral blood oxygen saturation)/FIO2 (fraction of inspired oxygen) ratio (SpFi). SpFi group 1 &gt;452, SpFi2 2: 315-452; SpFi 3 236-315, SpFi 4: &lt;236 (respiratory distress) Results 53 patients were included: epidemiological and clinical data available on table 1. 23 patients (43.39%) arrived to SpFi4 status. WHOs &gt;6 (WHO 6:oxygen by non invasive ventilation or high flow) was achieved by 20 patients (37.7%) Good prognosis (meaning SpFi1 as the worse respiratory status in the follow -up) was associated to cytometric profiles: there was a significant increase in CD3, p-DCs, m-DCs, basophils, monocytes, Tcm, % of lymphocytes and CD3/CD19 ratio whereas there was a significant reduction in CD19, % of neutrophils and % Neutrophils/% lymphocyte ratio. In the SpFi4 group, there was a significant reduction of CD3, p-DCs, m-DCs, plasmablasts and CD3/NK ratio. In patients starting in SpFi group 1-2 in the ED but progressing to SpFi 3-4 during the follow up (27 patients), there was a statistically significant relation with Tn, Te and Tn/Te ratio (Tn/Te ratio &lt;0.717: OR 13.5 (p 0.002, [95% CI 2.552-71.403). Initial SpFi1 patients that evolved to SpFi 3-4 during follow up (10 patients), presented a Tn/Te ratio &lt; 0.717 with an OR 11.556 (p =0.005, [95% CI 2.059-64.853]). Plasmablasts &lt; 0.075 and CD3/NK &lt;5.71, were identified as independent risk factors for SpFI4 during follow up. After multivariable analysis, both variables kept their significance: CD3/NK (OR 11,247, p=0.005) and plasmablasts (OR 12,524 , p=0.004). About prediction of WHOs &gt;6, multivariable analysis showed CD3/NK &lt;5.71 (OR 22,240 [95%CI 2,340-211,342] p= 0.007) and plasmablasts&lt;0.075 (OR 28.635 [95% CI 3,187-257,301] p=0.003). A score (0,1,2) comprising both risk factors, was significantly predictive of SpFI4, regardless of the initial respiratory status, age or days from symptoms onset. In our cohort, only 1/15 (6.7%) patients with 0 points (neither plasmablasts nor CD3/NK score), arrived to SpFi4. However, 10/11 (90.9%) patients with 2 points, reached to SpFi4 respiratory status (C-index = 0.837) Same score was applied to predict WHOs &gt; 6: 90.9% with 2 points progressed to WHO&gt;6 and 0/15 patients with 0 points reached the same goal (C-index = 0.872) An incidental finding of 4 indolent B-lymphoproliferative disorders (2CLL-like MBLs and 2non -CLL-like MBL), was found, and they were associated with older age and progression to death. Conclusions Flow cytometry on whole peripheral blood samples of SARS-COV2 pneumonia patients, collected before corticosteroid or immunosuppressive therapy, could identify cytometric patterns associated to prognosis. Plasmablasts, mDCs and pDCs levels as well as CD3/NK ratio, are associated to a worse respiratory status, while Tn/Te ratio could detect non-severe patients who will require high-flow oxygen devices during follow up. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

scholarly journals POS1015 ANTI-TNF DRUGS AND CARDIOVASCULAR EVENTS IN PATIENTS WITH SPONDYLOARTHRITIS

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 775.2-776
Author(s):  
C. W. S. Chan ◽  
P. H. LI ◽  
C. S. Lau ◽  
H. Y. Chung
Keyword(s):  
Risk Factors ◽  
Psoriatic Arthritis ◽  
Cardiovascular Events ◽  
Clinical Information ◽  
Incidence Rates ◽  
Major Adverse Cardiovascular Events ◽  
Cross Sectional ◽  
Cerebrovascular Events ◽  
Cardiovascular Morbidity And Mortality

Background:Cardiovascular (CVS) diseases are the leading cause of death worldwide and patients with rheumatic diseases have an increased CVS risk including stroke and myocardial infarction (MI) (1-3). CVS risk factors and CVS events are common in SpA (4). Delineating the CVS risk and the association with medications in patients with SpA would be useful.Objectives:The objective of this study was to delineate the CVS risk and the association with medications in patients with SpA.Methods:Patients with SpA and patients with non-specific back pain (NSBP) were identified in rheumatology and orthopedics clinics respectively. Clinical information and CVS events were retrieved. Incidence rates were calculated. Association analysis was performed to determine the CVS risk of SpA and other modifiable risk factors.Results:A total of 5046 patients (SpA 2616 and NSBP 2430) were included from eight centers. Over 56 484 person-years of follow-up, 160 strokes, 84 MI and 262 major adverse cardiovascular events (MACE) were identified. Hypercholesterolemia was more prevalent in SpA (SpA 34.2%, NSBP 28.7%, P<0.01). Crude incidence rates of stroke and MI were higher in SpA patients. SpA was associated with a higher risk of MACE (HR 1.66, 95%CI 1.22-2.27, P<0.01) and cerebrovascular events (HR 1.42, 95%CI 1.01-2.00, p=0.04). The use of anti-tumor necrosis factor (TNF) drugs was associated with a reduced risk of MACE (HR 0.37, 95%CI 0.17-0.80, P=0.01) and cerebrovascular events (HR 0.21, 95%CI 0.06-0.78, P=0.02).Conclusion:SpA is an independent CVS risk factor. Anti-TNF drugs were associated with a reduced CVS risk in these patients.References:[1]Crowson CS, Liao KP, Davis JM, 3rd, Solomon DH, Matteson EL, Knutson KL, et al. Rheumatoid arthritis and cardiovascular disease. Am Heart J. 2013;166(4):622-8 e1.[2]Verhoeven F, Prati C, Demougeot C, Wendling D. Cardiovascular risk in psoriatic arthritis, a narrative review. Joint Bone Spine. 2020;87(5):413-8.[3]Liew JW, Ramiro S, Gensler LS. Cardiovascular morbidity and mortality in ankylosing spondylitis and psoriatic arthritis. Best Pract Res Clin Rheumatol. 2018;32(3):369-89.[4]Molto A, Etcheto A, van der Heijde D, Landewe R, van den Bosch F, Bautista Molano W, et al. Prevalence of comorbidities and evaluation of their screening in spondyloarthritis: results of the international cross-sectional ASAS-COMOSPA study. Ann Rheum Dis. 2016;75(6):1016-23.Disclosure of Interests:None declared.

scholarly journals Immunostimulatory properties of bigroot geranium (Geranium macrorrhizum L.) extract

Medicina ◽  
2007 ◽  
Vol 43 (1) ◽  
pp. 60 ◽  
Author(s):  
Vilma Jurkštienė ◽  
Anatolijus Kondrotas ◽  
Egidijus Kėvelaitis
Keyword(s):  
T Lymphocytes ◽  
B Lymphocytes ◽  
Peripheral Blood ◽  
Leukocyte Count ◽  
Ethanol Extract ◽  
Food Supplement ◽  
Total Leukocyte Count ◽  
Control Group ◽  
Case Group ◽  
Cell Counts

The aim of the study was to investigate the immunostimulatory properties of bigroot geranium. Material and methods. Possible nonspecific characteristics of bigroot geranium were evaluated by the total leukocyte count in the peripheral blood, and qualitative changes of blood were assessed using Shilling’s formula by evaluating changes in lymphocyte counts. In addition, we also studied changes in the counts of Tcell precursors in the thymus and B lymphocytes in the spleen. Ethanol extract of the leaves of bigroot geranium was produced at the Department of Food Technology, Kaunas University of Technology. Studies were performed on mice Bl 57 (n=21). The control group (n=7) received distilled water at a dose of 1 mL/day. The second and third groups received 1% and 10% extract of bigroot geranium, respectively, as a food supplement. Changes in cell counts were investigated after 4 weeks following the initiation of the trial. Results. After a 4-week administration of 1% extract of bigroot geranium (1 mL/day) (mice group, n=7), leukocyte count in the peripheral blood increased to 6.1×109 cells/L, and lymphocyte count – to 70%, but changes were not statistically significant. The other case group of mice (n=7) received 10% extract of bigroot geranium for 4 weeks at a dose of 1 mL/day. In this group, leukocyte count in the peripheral blood increased statistically significantly from 4.4×109 cells/L to 7.2×109 cells/L (p<0.01), and lymphocyte percentage – from 52% to 80% (p<0.001), as compared to control. Thymocyte (T lymphocytes) counts in thymus and splenocyte (B lymphocytes) counts in the spleen showed a tendency to increase after the administration of 1% and 10% extracts. After a 4-week administration of 1% extract of bigroot geranium, thymocyte and splenocyte counts increased from 0.342×106 cells to 0.372×106 cells per mg of tissue and from 0.395×106 cells to 0.405×106 cells per mg of tissue, respectively, as compared to control group (p>0.1). After the administration of 10% extract of bigroot geranium, thymocyte count increased to 0.488×106 cells per mg of tissue (p<0.01), and splenocyte count – to 0.504×106 cells per mg of tissue (p<0.01). Conclusion. The extracts of the leaves of bigroot geranium increased leukocyte count and lymphocyte percentage in the peripheral blood, and after a 4-week administration of 10% extract of bigroot geranium, a statistically significant increase in the counts of T lymphocytes (in the thymus) and B lymphocytes (in the spleen) was observed. The immunostimulatory effect depends on the dose of the extract.

Rapid Prediction of Peripheral Blood CD34 Counts Using a Multiple Regression Model Derived from Automated Hematology Analyzer Cell Population Data

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3099-3099 ◽  
Author(s):  
Thomas Porturas ◽  
Mary Sell ◽  
Leah Irwin ◽  
Una O'Doherty ◽  
Carlos Hipolito Villa
Keyword(s):  
Flow Cytometry ◽  
Stem Cell ◽  
Multiple Regression ◽  
Peripheral Blood ◽  
Population Data ◽  
Blood Samples ◽  
Peripheral Blood Cd34 ◽  
Cell Counts ◽  
Flow Cytometric ◽  
Hematology Analyzer

Abstract Background: Although peripheral blood CD34+ stem cell counts by flow cytometry correlate well with yields, the time, complexity, and cost associated with flow cytometry limits its utility. Rapid, cost-effective, surrogate predictors (with <1hr turnaround) would allow for same-visit analyses and alteration of collection and mobilization strategies, particularly for the optimal use of time-sensitive and costly agents such as plerixafor. We previously demonstrated that morphologic parameters of neutrophil-like cells measured by hematology analyzers correlated with CD34 counts. We aimed to improve these models by using multiple regression analyses on data from a common hematology analyzer. Methods: Patients undergoing stem cell apheresis were evaluated over a 6 month period. The day prior to initiation of apheresis, and on the morning of initial collection, peripheral blood samples were drawn into EDTA collection tubes and flow cytometric CD34 measurement and/or CBCs were performed on the Beckman Coulter DxH 800 hematology analyzer per standard protocol. CD34 cells were counted by flow cytometric ISHAGE protocols. Data from the DxH (48 variables per specimen) were exported into a data matrix with the corresponding flow cytometric data. Multiple regression analysis was performed using a step-wise method with log(peripheral CD34) as the dependent variable (SPSS, IBM). Data were randomly selected into a training-set of 70% of cases and a test-set of 30% of cases for validation. The derived model was further tested against peripheral blood data from the morning of collection to predict harvest yields. Further analyses were performed using Prism (GraphPad). Results: Tandem peripheral blood CD34 counts and CBC cell-population data were obtained from 69 blood samples in 64 patients. The population included patients with multiple myeloma (45), non-Hodgkin lymphoma (12), Hodgkin lymphoma (5), and amyloidosis (2). 41% of patients were female. In the test data set examining collection yields, 37 patients were mobilized with GCSF (+/- chemotherapy) alone, while 17 had plerixafor added to the regimen. 33 of these patients had same-day CBC data available for model prediction. The median processed volume was 15 L (range 5.9 to 19.7). The model to predict peripheral CD34 counts incorporated 3 variables from the hematology analyzer data (SD-V-EGC, SD-C-EGC, and NE#). Interestingly, the model included two variables descriptive of the morphology of early granulocytic cells. The model demonstrated an R value of 0.829 (adjusted R2 = 0.670, figure 1a). In testing the morning-of-collection model-predicted peripheral CD34, we found the model performed similarly to flow cytometry in predicting 1st collection yields. Furthermore, the CD34 prediction using the model (Figure 1 b) resulted in similar correlation with first-collection yields in patients treated with plerixafor versus patients not treated with plerixafor, in contrast to day-prior CD34 counts by flow-cytometry (Figure 1c). Two outliers for CD34 cell yield based on model predicted peripheral CD34 were identified. In one patient, the processed volume was very low (6.8 L, <5% percentile), while the second had a low mononuclear cell collection efficiency (35%) compared to the mean in this population (58.7%±23.3%). Threshold values for the model accurately identified patients appropriate for collection initiation (or plerixafor administration). Conclusion: Using data from a common, automated CBC analyzer, we developed a rapid, less-costly, and simple model to predict CD34 cell counts and 1st harvest yields. Because the measurement results can be obtained within the same clinic visit, and can be repeated with each CBC, the model is particularly useful to guide optimal use of plerixafor. We also envision that the model is useful for quality assurance of collection by identifying patients in whom cell yields were sub-optimal with respect to predicted CD34 cell counts. Additional studies to test the model in a larger population are ongoing. We propose that this model (and similarly derived models) can be implemented in clinical planning algorithms to improve the efficiency and cost of stem cell collection by apheresis. Acknowledgments: We would like to acknowledge and the nurses and staff of the apheresis unit and the stem cell and flow cytometry laboratories at the Hospital of the University of Pennsylvania for their contributions. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

Role of Carotid Ultrasound and Systematic Coronary Risk Evaluation Charts for the Cardiovascular Risk Stratification of Patients with Psoriatic Arthritis

2019 ◽  
Vol 47 (5) ◽  
pp. 682-689 ◽  
Author(s):  
María Paz Martínez-Vidal ◽  
Mariano Andrés ◽  
Vega Jovaní ◽  
Carlos Santos-Ramírez ◽  
Cintia Romera ◽  
...  
Keyword(s):  
Risk Factors ◽  
Psoriatic Arthritis ◽  
High Risk ◽  
Risk Evaluation ◽  
Intima Media Thickness ◽  
Inflammatory Rheumatic Diseases ◽  
Coronary Risk ◽  
Carotid Ultrasound ◽  
Cross Sectional ◽  
Cv Disease

Objective.The assessment of the cardiovascular (CV) risk is recommended in patients with chronic inflammatory rheumatic diseases. The objectives of this study were to assess the CV risk profile in a cohort of patients with psoriatic arthritis (PsA), to determine the presence of subclinical cardiovascular disease by carotid ultrasound (US), and to study the association of CV disease to PsA characteristics.Methods.This was a cross-sectional multicentric descriptive study. The clinical CV risk was calculated with Systematic Coronary Risk Evaluation (SCORE) charts. Common carotid US was conducted to evaluate the carotid wall intima-media thickness and the presence of atheroma plaques. Patients were reclassified upon US results. Multivariate analyses were performed to identify associations of US carotid abnormalities with the classical CV risk factors and PsA characteristics.Results.The study included 176 patients with PsA. The SCORE-estimated CV risk was intermediate in 65.3% of the patients. In the US study, 32% of the patients had abnormalities, and 30.8% of the patients were upgraded and reclassified as very high risk owing to the presence of atheroma. Subclinical CV disease was associated with age and dyslipidemia but not with other risk factors. It was associated with axial disease in the subgroup with intermediate risk, and with C-reactive protein levels in patients with high risk.Conclusion.Many patients with PsA have clinical estimated intermediate or high risk of a fatal CV event. A carotid US study detects subclinical vascular disease and may be useful to depict the real risk. The presence of atheroma is only partially explained by the classic CV risk factors.

scholarly journals Platelet-bound C4d, low C3 and lupus anticoagulant associate with thrombosis in SLE

2019 ◽  
Vol 6 (1) ◽  
pp. e000318 ◽  
Author(s):  
Michelle A Petri ◽  
John Conklin ◽  
Tyler O'Malley ◽  
Thierry Dervieux
Keyword(s):  
Risk Factors ◽  
Flow Cytometry ◽  
Logistic Regression ◽  
Risk Score ◽  
Lupus Anticoagulant ◽  
Cross Sectional ◽  
The Past ◽  
Pt Complex ◽  
Composite Risk ◽  
Sectional Analysis

BackgroundLow C3 and lupus anticoagulant (LAC) are known risk factors for thrombosis in SLE. We evaluated the association between C4d products deposited on platelets (PC4d) and thrombosis in SLE. Antiphosphatidyl serine/prothrombin (PS/PT) complex antibody was also evaluated as an alternative to LAC.MethodsThis was a cross-sectional analysis of 149 consented patients with SLE (mean age: 47±1 years, 86% female) classified with (n=16) or without (n=133) thrombotic events in the past 5 years. Abnormal PC4d (≥20 units) was measured using flow cytometry. LAC and C3 were measured using dilute Russell’s viper venom time (>37 s) and immunoturbidimetry, respectively. Anti-PS/PT antibody status (IgG) was measured by immunoassay. Statistical analysis consisted of logistic regression and calculation of OR estimates with 95% CI.ResultsAbnormal PC4d (OR=8.4, 95% CI 2.8 to 24.8), low C3 (OR=9.5, 95% CI 3.0 to 30.3), LAC (OR=5.4, 95% CI 1.3 to 22.3) and anti-PS/PT IgG (OR=3.4, 95% CI 1.2 to 9.7) status associated with thrombosis (p<0.05). Cumulatively, the presence of PC4d, low C3 and LAC abnormalities as a composite risk score was higher in the presence of thrombosis (1.93±0.25) than in its absence (0.81±0.06) (p<0.01). Each unit of this composite risk score yielded an OR of 5.2 (95% CI 2.5 to 10.7) to have thrombosis (p<0.01). The composite risk score with anti-PS/PT antibody status instead of LAC also associated with thrombosis (p<0.01).ConclusionA composite risk score including PC4d, low C3 and LAC was associated with recent thrombosis and acknowledges the multifactorial nature of thrombosis in SLE.

The Morphology and Cellular Characterization of Thrombocytes in Adult Xenopus laevis.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3947-3947
Author(s):  
Takako Ishida ◽  
Miyako Obuchi-Shimoji ◽  
Takeshi Kuribara ◽  
Nami Nogawa ◽  
Tomoyuki Tahara ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Xenopus Laevis ◽  
Peripheral Blood ◽  
Blood Cells ◽  
Structural Characteristics ◽  
Flow Cytometric Analysis ◽  
Peripheral Blood Cells ◽  
Morphological Aspect ◽  
Cell Counts

Abstract In primates and rodents, platelets originate from the bone marrow megakaryocytes through a unique differentiation process with nuclear polyploidization, cytoplasmic maturation and proplatelet formation. In contrast, circulating thrombocytes of most non-mammalian vertebrates are particularly distinctive; the cells are large and nucleated. Adult Xenopus laevis may be an useful non-mammalian model for analyzing dynamic hematopoiesis because they are individually tolerable for time lapse analysis in vivo with sequential blood sampling, whereas classification of cell types has not been established yet. Microstructures of Xenopus thrombocytes observed with electron microscope exhibited structural characteristics largely resembling zebrafish thrombocytes with nucleated spindle cellular features (Thattaliyath et al., Blood 2005), and they had lobulated nuclear chromatin, granules, microparticles and open canalicular system-like-structures as in mammalian megakaryocytes. Since thrombocyte identification based on the morphological aspect was not sufficient, chemical staining with acetylecholinesterase and thiazole orange were performed. Additionally, mice were immunized by Xenopus peripheral blood cells to generate monoclonal antibodies, and two hybridomas producing IgG, respectively T12 and T5, were screened. T12+ (T12 positive) cells were morphologically typical thrombocytes. Flow cytometric analysis revealed that T12+ cells were also positive to anti-human GpIIb/IIIa polyclonal antibodies, and approximately 2-3% of whole peripheral blood cells were T12+/GpIIb/IIIa+ that distributed in FSClow/SSClow fraction. When T12 was injected into Xenopus to deplete T12+ cells in vivo, the detectable level of T12 in the circulation lasted for more than several weeks. Peripheral thrombocyte counts predominantly began to decrease immediately and reached their nadir at day 3, but white blood cell counts were not changed. RNA-rich blood cells considered as younger cells were then increasingly appeared, and finally the cell counts recovered to normal levels at day 10–15, indicating that in vivo depletion of T12+ cells induced thrombopoiesis and/or release of mature thrombocytes from the pool. T5 recognizing cells were classified into two populations by immunostaining and flow cytometry; T5+/GpIIb/IIIa+ cells were morphologically thrombocytic as the cells recognized by T12, while T5+/GpIIb/IIIa− cells were spherical and similar appearance to lymphocytic cells. These observations raised some possibilities e.g.; antigen of T5 was a membrane protein common to both lymphocytes and thrombocytes, or T5+/GpIIb/IIIa− cells were thrombocyte progenitors at earlier development stage than T12+/GpIIb/IIIa+ cells. Nevertheless only a few percent of T12+ and T5+ cells resided in peripheral blood, immunostaining revealed that the proportions of T12+/T5+ and T5+ cells in spleen were 10% and 70%, and T12+/T5+ and T5+ cells in liver were 5% and 20%, respectively. These suggest that spleen is predominantly involved in thrombopoiesis and/or thrombocyte storage in adult Xenopus. As T12 and T5 can be used successfully in flow cytometry and magnetic cell sorting, they should contribute us directly to elucidate the origin of circulating Xenopus thrombocytes and their cellular development process.

Family-Associated Monoclonal B Lymphocytosis Shows Differences From CLL That Suggest An Indolent Biology.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1241-1241
Author(s):  
Mark C Lanasa ◽  
Sallie D Allgood ◽  
Susan L Slager ◽  
Nicola J Camp ◽  
Neil E. Kay ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Peripheral Blood ◽  
Conflicts Of Interest ◽  
Family Members ◽  
Surface Expression ◽  
Cellular Activation ◽  
Immunoglobulin Isotype ◽  
Cell Counts ◽  
Intracellular Expression ◽  
Trisomy 12

Abstract Abstract 1241 Poster Board I-263 Background and Significance Monoclonal B lymphocytosis (MBL) is a hematologic syndrome characterized by accumulations of clonal B lymphocytes in the peripheral blood. Most MBL have a CLL-like immunophenotype, though other, less common phenotypes are observed. Although some MBL, particularly those with MBL cell counts >1.9 × 109 / L, progress to CLL, “low count” MBL (<0.2 × 109 MBL / μL) have little potential to progress to MBL and may have different biologic characteristics from MBL with lymphocytosis or Rai Stage 0 CLL. Our hypothesis was that detailed characterization of family-associated MBL may also provide biologic insights into the pathogenesis of CLL. Methods Individuals with MBL were identified by flow cytometry screening of fresh and cyropreserved peripheral blood from unaffected members of CLL kindreds ascertained by Genetic Epidemiology of CLL Consortium (GEC) sites. We defined MBL as populations of CD5+, CD19+, CD20lo, CD23+ B cells that comprised at least 0.02% of the PBMC and did not exceed 5.0 × 109 MBL cells / L. Flow cytometry was used to determine the surface immunophenotype including prognostic parameters of CD38, intracellular ZAP-70, immunoglobulin isotype, CD49d, and the ratio of CD69:71. mRNA and genomic DNA from single MBL cells isolated by flow cytometric sorting were analyzed using PCR to determine immunoglobulin heavy and light chain sequences (IGVH status). MBL cells were sorted in bulk for FISH determination of genetic loci associated with clinical CLL. Results Fifty-four unaffected family members were found to have MBL, of these 48 (89%) showed a typical CLL immunophenotype (CLL-like MBL). We observed significant variability in the size of the MBL clone as a percentage of the CD19+ B cell compartment (mean 30%, median 16%; range 2% - 97%). CD38 positive (defined as CD38 surface expression in ≥ 30% of MBL cells) was observed in 6 of 38 (16%) subjects tested. ZAP-70 positive (defined as intracellular expression in ≥ 20% of MBL cells) was expressed in 5 of 28 (18%) participants. CD49d positive (defined as surface expression in ≥ 45% of MBL cells) was observed in 4 of 17 (24%) subjects. The ratio of surface expression of CD69:CD71, a measure of cellular activation that also correlates with an unmutated IGVH, was ≥2.0 in 3 of 34 (9%) subjects tested. Among 36 subjects tested, 9 (25%) MBL expressed both surface IgD and IgM (defined as surface expression ≥ 40% for both IgM and IgD), 12 (33%) expressed IgD only, 2 (6%) expressed IgM only, and 13 did not express IgD or IgM (36%). Analysis of IGVH status has been completed in 10 individuals. Both immunoglobulin heavy chain variable (IGVH) region mutated (n = 14) and unmutated (n = 5) sequences were observed. Six of 10 individuals had 2 or more unrelated MBL clones (range 2 - 4), including two individuals with both unmutated and mutated clones. Among the 19 MBL clones identified in these 8 subjects, VH3 or VH4 rearrangements were observed in all MBL clones. The most commonly rearranged IGVH genes were 3-07 (3 MBL clones), 3-15 (3), and 4-34 (3). No VH1 family gene rearrangements were observed. MBL cells were bulk sorted for FISH from 14 subjects. Mono or biallelic deletion of 13q14.3 was observed in 9 subjects, 4 were normal, and one showed trisomy 12. Conclusions Our data affirms that CLL-like MBL are commonly observed among the unaffected family members from CLL kindreds. We found that family associated MBL clones (most of which were small clones) express ZAP-70, CD38, and CD49d at an apparently lower frequency than observed in CLL. Unlike in CLL, the surface immunoglobulin isotype showed co-expression of IgM and IgD in only one third of cases. Moreover, small MBL clones are commonly oligoclonal and predominantly express mutated IGVH genes with an IGVH usage that also appears different from CLL. Taken together, these findings suggest that small MBL clones, though phenotypically similar to CLL, have important biologic differences from CLL that may explain the limited potential of these clones to progress to CLL. The clinical outcome of these MBL clones in relation to our baseline prognostic characterization will be of interest. In addition the further investigation of family associated MBL is being conducted and may clarify the genetics and immunobiology of familial CLL. Disclosures No relevant conflicts of interest to declare.

scholarly journals Simplified quantitation of myeloid dendritic cells in peripheral blood using flow cytometry

Cytometry ◽  
2000 ◽  
Vol 40 (1) ◽  
pp. 50-59 ◽  
Author(s):  
John W. Upham ◽  
Joachim Lundahl ◽  
Hong Liang ◽  
Judah A. Denburg ◽  
Paul M. O'Byrne ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Dendritic Cells ◽  
Peripheral Blood ◽  
Myeloid Dendritic Cells
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