Megakaryocyte-Active Chemokines: Dysregulation in the SDF-1a/CXCR4 Axis in Patients with Essential Thrombocythemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4969-4969
Author(s):  
Juan P. Salim ◽  
Rosana F. Marta ◽  
Felisa C. Molinas
Keyword(s):  
Flow Cytometry ◽  
Fluorescence Intensity ◽  
Essential Thrombocythemia ◽  
Plasma Levels ◽  
Platelet Membrane ◽  
Rank Test ◽  
Control Group ◽  
Expression Levels ◽  
Normal Controls ◽  
Cxcr4 Axis

Abstract Chemokines belong to a large family of molecules that are implicated in the localization and production of blood cells. Some of them, such as Interleukin 8 (IL-8) and GRO-a, participate in the regulation of megakaryopoiesis, mostly by exerting an inhibitory action. Recently, the involvement of stromal derived factor 1 (SDF-1) in synergizing the stimulatory effect of thrombopoietin on megakaryopoiesis and its participation in megakaryocyte transendothelial migration has been described. The aim of the present study was the evaluation of the plasma levels of IL-8, GRO- a and SDF-1 in patients with essential thrombocythemia (ET), a myeloproliferative disorder characterized by megakaryocytic hyperproliferation with increased circulating platelet count. Besides, the corresponding chemokine receptors were assayed on platelet membrane from ET patients. A cohort of 27 patients diagnosed according to the Polycitemia Vera Study Group were enrolled in the study (mean age, 45; 21 women). Twenty-seven normal subjects matched by sex and age were taken as the control group. The Ethic Committee from IDIM A. Lanari approved the study and all patients and normal controls signed the informed consent. Plasma levels of the chemokines were measured by ELISA technique (R&D Systems) according to the manufacturer. Expression levels of IL-8 receptors (CXCR1 and CXCR2), GRO-a receptors (CXCR2) and SDF-1 receptors (CXCR4) on platelet membrane were evaluated by flow cytometry using specific MoAbs and the corresponding isotype controls (B-D Pharmingen). Flow cytometry results were expressed as relative fluorescence intensity (RFI, the relationship between the mean fluorescence intensity from the specific antibody and the isotype control). Results were expressed as median and range. Statistic analysis was carried out using Mann-Whitney Wilcoxon rank sum test and Wilcoxon signed rank test. Plasma levels of the chemokines measured in 19 ET patients were similar to that found in normal controls, IL-8 2.5, pg/ml (0.8–28.2) and 2.8 pg/ml (1.1–16.5), GRO-a, 30.0 pg/ml (7.4–463.1) and 23.9 pg/ml (9.6–148.0), SDF-1, 1895.0 pg/ml (1246.0–2719.0) and 1915.0 pg/ml (822–2424.0), respectively. The expression levels of CXCR4 receptor was found diminished in platelets from ET patients, RIF 16.94 (1.3–31.3) compared to normal controls, 27.4 (2.4–58.4); p=0.0059, n=10. However, the expression of CXCR1 and CXCR2 in platelets from ET patients was normal. In conclusion, although plasma levels of the chemoquines IL-8, GRO-a and SDF-1 were normal in these patients, the decreased level of CXCR4 on platelet membrane suggests a dysregulation in the SDF-1a/CXCR4 axis in patients with essential thrombocythemia.

Decreased CXCR4 mRNA Levels and Abnormal SDF-1 Response in Platelets from Essential Thrombocythemia.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3637-3637
Author(s):  
Juan P. Salim ◽  
Rosana F. Marta ◽  
Nora P. Goette ◽  
Carlos D. Chazarreta ◽  
Felisa C. Molinas
Keyword(s):  
Platelet Aggregation ◽  
Fluorescence Intensity ◽  
Essential Thrombocythemia ◽  
Mrna Levels ◽  
Platelet Response ◽  
Expression Levels ◽  
Functional Studies ◽  
Cxcr4 Mrna ◽  
Normal Controls ◽  
Cxcr4 Axis

Abstract Chemokines are soluble mediators implicated in a wide range of physiologic and pathologic processes including megakaryocytic development and platelet production. Previously, we found decreased levels of CXCR4, the only known receptor for the chemokine SDF-1, on platelet membrane from essential thrombocythemia (ET) patients, a chronic myeloproliferative disease characterized by megakaryocytic hyperplasia and high platelet count. In the present work we sought to evaluate the expression levels of platelet CXCR4 mRNA, the functional consequences of CXCR4 decrease in ET platelets as well as whether the receptor reduction was also extended to other lineages. We studied the expression levels of CXCR4 mRNA in platelets from ET patients by semi-quantitative RT-PCR. For semi-quantification, co-amplification of a GAPDH internal control sequence was performed. Functional studies consisted in turbidimetric evaluation of platelet aggregation induced by SDF-1 and/or ADP. In order to evaluate CXCR4 expression on leukocyte membrane, white cell subpopulations were individualized by the addition of specific MoAbs directed against CD45 labeled with PerCP and CD3 and CD14 labeled with FITC. Data were expressed as relative fluorescence intensity (RFI, ratio between the mean fluorescence intensity of CXCR4 antibody and the corresponding isotype control). The ratio between CXCR4 mRNA and the housekeeping gene GADPH in platelets from patients, 0.16 (0.1–0.24) was significantly decreased compared to controls, 0.35 (0.18–0.86), n=11, p=0.0002 (Wilcoxon signed rank sum test). Functional studies in response to SDF-1 showed a biphasic, dose-dependent full platelet aggregation in PRP in normal controls. On the contrary, abnormalities in SDF-1 response were observed in all 8 patients evaluated. Our results showed that platelet response to SDF-1 in ET patients closely paralleled that of ADP. Three patients did not respond either to SDF-1 or to ADP. From the remaining five patients, three had a delay in the second wave of aggregation induced by SDF-1 and the other two displayed only a primary wave of aggregation. These patients had a normal response to ADP when standard concentrations were used (2 mmol/L), but displayed a decreased response compared to normal controls when challenged with suboptimal doses of this agonist (0.8 to 1.2 mmol/L). CXCR4 RFI in granulocytes from patients was found elevated, 11.1 (2.22–22.74) compared to normal controls, 7.54 (1.73–9.52), p=0.036 (n=7). Similar results were found in CD3-positive lymphocytes, 40.0 (6.78–67.91) from patients and 17.23 (5.22–29.97) from normal controls, p=0.036 (Mann-Whitney Wilcoxon Rank Sum Test). On the contrary, CXCR4 on monocyte membrane was normally expressed, RFI from patients, 49.81 (1.17–118.28), and normal controls, 74.23 (3.0–80.49), p=0.8. These results indicate that the decrease in CXCR4 membrane expression on ET platelets is related to a reduced CXCR4 mRNA, and also to the abnormal platelet response to SDF-1. Besides, flow cytometric studies on leukocyte subpopulations demonstrated that CXCR4 decrease is restricted to the megakaryocytic lineage. Deregulation of SDF-1/CXCR4 axis in ET patients could be implicated in the megakaryocytic alterations seen in this illness and its presence in a megakaryocyte-related disease highlights the important role of SDF-1/CXCR4 axis in platelet development.

Association of microRNA-125a with the clinical features, disease activity and inflammatory cytokines of juvenile-onset lupus patients

Lupus ◽  
2021 ◽  
pp. 096120332110103
Author(s):  
Eman Eissa ◽  
Botros Morcos ◽  
Rania Fawzy Mahmoud Abdelkawy ◽  
Hanan H Ahmed ◽  
Naglaa M Kholoussi
Keyword(s):  
Disease Activity ◽  
Inflammatory Cytokines ◽  
Lupus Erythematosus ◽  
Plasma Levels ◽  
Clinical Manifestations ◽  
Juvenile Onset ◽  
Expression Levels ◽  
Ifn Γ ◽  
Chronic Autoimmune Disease ◽  
Normal Controls

Background Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with marked variation in its clinical presentation. Juvenile-onset SLE (jSLE) exhibits an aggressive clinical phenotype and severe complications. Dysregulated expression of microRNAs (miRs) in immune cells from patients with SLE has been found. We aim to evaluate the association of miR-125a with the clinical and laboratory characteristics, disease activity and inflammatory cytokines of jSLE patients. Methods 60 jSLE patients and 25 normal controls were involved in the study. The expression pattern of miR-125a was determined in plasma of all subjects using qRT-PCR. In addition, plasma levels of IL-17 and IFN-γ were examined using ELISA. The correlation of miR-125a expression with the clinical manifestations and disease activity of jSLE patients was analyzed. Also, its association with the inflammatory cytokines was investigated in jSLE patients. Results Our findings showed that miR-125a expression levels were significantly reduced in jSLE patients compared to normal controls ( p < 0.01) and these expression levels differed based on the clinical variability of patients. In addition, plasma levels of IL-17 and IFN-γ in jSLE patients were significantly higher than healthy controls ( p < 0.01). Finally, miR-125a expression had significant negative associations with each of SLEDAI-2K ( p < 0.01), SLICC ( p < 0.01), ESR ( p < 0.05), proteinuria ( p < 0.01) and IL-17 levels ( p < 0.01) in jSLE patients. Conclusion Our findings postulate that miR-125a could act as a candidate therapeutic target for its possible regulation of inflammation in jSLE patients.

scholarly journals Expression of Salivary miRNA-31 in Oral Submucous Fibrosis

2021 ◽  
pp. 137-144
Author(s):  
Saba Khan ◽  
Saima Akram Butt ◽  
Sobia Hassan ◽  
Rizma Khan
Keyword(s):  
Early Stage ◽  
Oral Submucous Fibrosis ◽  
Significant Negative Correlation ◽  
Fold Change ◽  
Rank Test ◽  
Control Group ◽  
Clinical Parameters ◽  
Expression Levels ◽  
Group I ◽  
Submucous Fibrosis

Objective: The present study aimed to evaluate the salivary miRNA31 expression in controls and cases and associate miRNA31 levels with clinical parameters of oral submucous fibrosis. Methods: This case control study was conducted in a hospital setup. A total of 50 individuals participated in the study with 25 subjects in group I (healthy individuals) and 25 subjects in group II 25 diagnosed cases of (oral submucous fibrosis). The sample size was calculated with open Epi version 3.01.  A detailed assessment of clinical parameters of oral submucous fibrosis was made. Unstimulated saliva samples were collected from all study subjects meeting the inclusion criteria and analysis of saliva samples was done by qRTPCR. Results: The results showed high expression levels of miRNA31 in oral submucous fibrosis as compared to the control group. The study demonstrated significantly higher median fold change of miRNA-31 expression level in OSMF patients as compared to the participants in the control group. Correlation between age of patients and miRNA31 fold change was discerned using the Spearman rank test that demonstrated a non-significant negative correlation. Conclusion: Increased expression levels of miRNA 31 among oral submucous fibrosis as compared to the control group make it a promising salivary biomarker that detects oral submucous fibrosis at an early stage of the disease.

scholarly journals In vitro observation: the GFP-E. coli adhering to porcine erythrocytes can be removed by porcine alveolar macrophages

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6439 ◽  
Author(s):  
Wei Yin ◽  
Chun Wang ◽  
Kuohai Fan ◽  
Na Sun ◽  
Yaogui Sun ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Confocal Microscopy ◽  
Fluorescence Intensity ◽  
Alveolar Macrophages ◽  
Control Group ◽  
Laser Confocal Microscopy ◽  
Functional Protein ◽  
Blank Control Group ◽  
E Coli

Although the activation of pathogen phagocytosis via complement system has been studied, erythrocyte-phagocyte interactions in pigs are not clearly understood. Therefore, we sought to investigate the ability of porcine erythrocytes to clear immune complexes (ICs) by using laser confocal microscopy and flow cytometry to observe the immune adhesion of porcine erythrocytes to fluorescent bacilli and the immune presentation process of transferring fluorescent bacilli to macrophages. Isolated porcine alveolar macrophages (PAMs) had uniform morphology and size, and a survival rate of 97.2%. The phagocytosis rate was 98.8%. After WT E. coli was labeled with Fluorescein Isothiocyanate (FITC), the bacteria showed a bright green fluorescence, and the labeling rate was 92.3%. When laser confocal microscopy was utilized to observe the co-incubation system of porcine erythrocytes, PAM, and fluorescent E. coli, the fluorescence intensity of bacilli decreased with increasing observation time and even disappeared. Flow Cytometry examination showed that the average fluorescence intensity of PAMs co-incubated with porcine erythrocytes adhered to WT-E. coli-FITC, was significantly higher than that of normal PAMs. Furthermore, when porcine erythrocytes adhered to WT E. coli were incubated with PAMs, the surface mean fluorescence intensity of porcine erythrocytes was significantly higher than that of the blank control group. This shows that PAMs can competitively bind to the oposinized E. coli adhered to the surface of porcine erythrocytes, and these oposinized pathogens can enter macrophages by the process of phagocytosis, which promoting the internalization of ICs or pathogens. During this process, the physical morphology of porcine erythrocytes was not damaged, but the levels of its main functional protein CR1-like were reduced.

Does Occurrence of Other Malignancy after Essential Thrombocythemia and Polycythemia Vera Influence Their Rate of Transformation to Myelofibrosis?

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 5189-5189
Author(s):  
Lucia Masarova ◽  
Naval Daver ◽  
Naveen Pemmaraju ◽  
Prithviraj Bose ◽  
Sherry Pierce ◽  
...  
Keyword(s):  
Polycythemia Vera ◽  
Essential Thrombocythemia ◽  
Research Funding ◽  
Hematologic Malignancy ◽  
Myeloproliferative Neoplasm ◽  
Rank Test ◽  
Control Group ◽  
Transformation Rate ◽  
Similar Time ◽  
Female Ratio

Abstract Introduction: Other malignancies (OM) were reported to be increased in patients with essential thrombocythemia "ET" and polycythemia vera "PV". However, the effect of coexistent OM on the rate of disease transformation to myelofibrosis "MF" is unknown. Objective: To determine the occurrence of OM in patients with ET and PV and their influence on transformation rate to overt MF. Methods: Fisher's exact test, and Mann-Whitney test were used to compare categorical and continuous variable, respectively. Control group without OM matched for age, MPN diagnosis type and year was created to compare transformation rates, expressed by Kaplan-Meier curve and Log rank test. Results: We perform a retrospective chart review for 783 patients referred to our institution between years 1960 - 2013 with diagnosis of Philadelphia-negative myeloproliferative neoplasm "MPN", of which 256 had ET, 165 PV and 362 post ET- or post PV- MF. Overall, 107 (13.7%) patients had OM prior to MF transformation, accounting for total of 125 cases. See table for OM types and distribution. In total, 9 patients had more than 1 OM with the most common being prostatic carcinoma (n=6), melanoma (n=4) and renal cell carcinoma (n=2). Relapsed/refractory OM were noticed in 15 patients, 3 of these had recurrence only prior MPN. None of alive patients with refractory OM experienced progression after MF transformation. Median age at diagnosis of MPN and OM (59 and 62 years), median age at the time of first event (OM or MPN; 51 and 53 years), and male to female ratio (1:1) were similar. Time to secondary OM was shorter than time to secondary MPN (6.5 vs 9 years, p>0.05). After median follow up of 30 months (range, 1-150), 49 (46%) patients had died. Eleven (23%) patients died because of OM progression, most commonly due to lung carcinoma and melanoma. No MPN related deaths were observed, however, 2 patients progressed to acute leukemia. Median overall survival "OS" for MPN was 160 months (range, 1-240) and was not different for patients with and without MF (212 and 173 months; p>0.05). No significant differences were detected between patients with and without MF in terms of age at the time of MPN diagnosis, gender, MPN characteristics (cytogenetic, molecular mutations) or time between OM and MPN. However, patients who transformed to MF were older at the time of OM (53 vs 59 years, p= 0.041, 95% CI: 1.2-11.7). Similarly, within a group of MF, patients with OM prior to MPN were younger at the time of OM (53 vs 62 years, p<0.001, 95%CI 5.0-23.5) and older at the time of MPN (67 vs 55 years, p=0.001, 95%CI 8.3-19.4) that patients who had OM after previous MPN. Overall time to transformation to MF was similar between ET and PV, and between studied cohort and control group (6 years, range 0.5-29, p>0.05, chi square 1.14). However, patients who developed OM after MPN had longer time to transformation (14 years, range 1-36, p=0.011, HR 1.9, 95%CI of 1.3 -4.4), which remained significant when control group was added, p=0.024 [Fig.] Conclusions: OM diagnosed after ET or PV does not seem to influence rate of disease transformation to MF. Table. ET/PV, n=421 ET/PV -> MF, n= 362 OM characteristics OM prior MPN, n OM after MPN, n OM prior MPN, n OM after MPN, n No. of patients with OM 29 26 19 33 No. of OM 30 26 26 43 Breast 5 4 ( 1*) 2 (1*) 3 Female reproductive 5 1 4 Prostate 7 1 6 (1*) 14 (1*) Gastrointestinal tract 2 4 6 3 Genitourinary tract 3 (1*) 4 2 Lungs 1 3 (3*) Melanoma 3 (1*) 2 4 9 (1*) Soft tissue 1 2 (1*) 1 Thyroid 3 2 (1*) 1 2 Head and neck 1 (1*) Hematologic malignancy 3 5 (1*) 6 (1*) (*) stands for relapsed or refractory disease Disclosures Pemmaraju: Stemline: Research Funding; Incyte: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding; LFB: Consultancy, Honoraria. Cortes:Pfizer: Consultancy, Research Funding; Teva: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; ARIAD Pharmaceuticals Inc.: Consultancy, Research Funding.

scholarly journals NETs Lead to Sympathetic Hyperactivity After Traumatic Brain Injury Through the LL37-Hippo/MST1 Pathway

2021 ◽  
Vol 15 ◽  
Author(s):  
Kaixin Zhu ◽  
Yibai Zhu ◽  
Xiaoxiang Hou ◽  
Wen Chen ◽  
Xiaolin Qu ◽  
...  
Keyword(s):  
Traumatic Brain Injury ◽  
Flow Cytometry ◽  
Brain Injury ◽  
Diffuse Axonal Injury ◽  
Arginine Deiminase ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Mortality And Morbidity ◽  
Sympathetic Hyperactivity ◽  
Expression Levels

Background: Paroxysmal sympathetic hyperactivity (PSH) is one of the important reasons for the high mortality and morbidity of traumatic brain injury (TBI). We aim to explore the role of the neutrophil extracellular traps (NETs) in the pathogenesis of sympathetic hyperexcitability after TBI and the underlying mechanisms, providing evidence for clinical treatment.Methods: Enzyme-linked immunosorbent assay was used to assess the plasma metanephrine and normetanephrine levels which represented the variation of the sympathetic system after TBI with rat diffuse axonal injury (DAI) model. NETs in the paraventricular nucleus (PVN) and circulating blood were examined using immunofluorescence and flow cytometry. Neutrophils-microglia co-culture system was established to further explore the effect of NETs on PSH and its mechanisms.Results: After TBI, metanephrine and normetanephrine levels began to increase at 9 h and peaked at 72 h. After the injury, the level of NETs kept increasing at 24 and 72 h in the PVN. A positive correlation was found between the concentration of the PVN NETs and blood catecholamine. Flow cytometry of peripheral blood cells revealed that NETs level in the injury group was higher than that in the control group. Immunofluorescence results confirmed the presence of NETs in the PVN after TBI. The positive result of immunoprecipitation suggested a correlation effect between LL37 and P2 × 7. Peptidyl arginine deiminase-4 (PAD4) inhibitor could inhibit the expression levels of MST1, YAP, and IL-1β. The hippo/MST1 pathway inhibitor could inhibit the expression levels of YAP and IL-1β.Conclusion: NETs formation in the PVN might be associated with sympathetic hyperactivity after TBI, which might relate to the activation of microglia cells and increased secretion of IL-1β via the hippo/MST1 pathway.

scholarly journals Cytological observations of transfer of immune complexes via porcine erythrocytes

2018 ◽  
Author(s):  
Wei Yin ◽  
Chun Wang ◽  
Kuohai Fan ◽  
Na Sun ◽  
Yaogui Sun ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Confocal Microscopy ◽  
Fluorescence Intensity ◽  
Immune Complexes ◽  
Observation Time ◽  
Control Group ◽  
Laser Confocal Microscopy ◽  
Functional Protein ◽  
Blank Control Group ◽  
E Coli

ABSTRACT Although the activation of pathogen phagocytosis via complement system has been studied, erythrocyte-phagocyte interactions in pigs are not clearly understood. Therefore, we sought to investigate the ability of porcine erythrocytes to clear immune complexes by using laser confocal microscopy and flow cytometry to observe the immune adhesion of porcine erythrocytes to fluorescent bacilli and the immune presentation process of transferring fluorescent bacilli to macrophages. Isolated porcine alveolar macrophages (PAMs) had uniform morphology and size, and a survival rate of 97.2%. The phagocytosis rate was 98.8%. After WT E. coli was labeled with FITC, the bacteria showed a bright green fluorescence, and the labeling rate was 92.3%. When laser confocal microscopy was utilized to observe the co-incubation system of porcine erythrocytes, PAM, and fluorescent E. coli, the fluorescence intensity of bacilli decreased with increasing observation time and even disappeared. Flow Cytometry examination showed that the average fluorescence intensity of PAMs co-incubated with porcine erythrocytes adhered to WT-E. coli-FITC, was significantly higher than that of normal PAMs. Furthermore, when porcine erythrocytes adhered to WT E. coli were incubated with PAMs, the surface mean fluorescence intensity of porcine erythrocytes was significantly higher than that of the blank control group. This shows that PAMs can competitively bind to the oposinized E. coli adhered to the surface of porcine erythrocytes, and these oposinized pathogens can enter macrophages by the process of phagocytosis，which promoting the internalization of immune complexes or pathogens. During this process, the physical morphology of porcine erythrocytes was not damaged, but the levels of its main functional protein CR1-like were decreased.

scholarly journals Increased Lipid Peroxidation May Be Linked to Ferritin Levels Elevation in Adult-Onset Still’s Disease

Biomedicines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1508
Author(s):  
Po-Ku Chen ◽  
Kai-Jieh Yeo ◽  
Po-Hao Huang ◽  
Shih-Hsin Chang ◽  
Ching-Kun Chang ◽  
...  
Keyword(s):  
Lipid Peroxidation ◽  
Flow Cytometry ◽  
Fluorescence Intensity ◽  
Plasma Levels ◽  
Mean Fluorescence Intensity ◽  
Inflammatory Responses ◽  
Flow Cytometry Analysis ◽  
Inflammatory Parameters ◽  
Healthy Control ◽  
The Relationship

Lipid peroxidation (LPO) and hyper-ferritinemia are involved in inflammatory responses. Although hyper-ferritinemia is a characteristic of AOSD, its link to LPO remains unclear. We investigated the association between LPO and ferritin expression, and evaluated the relationship between LPO-related metabolites and inflammatory parameters. Mean fluorescence intensity (MFI) of LPO (C11-Biodipy581/591)-expressing PBMCs/monocytes in AOSD patients and healthy control (HC) subjects was determined by flow-cytometry analysis. Expression of ferritin and cytokines on PBMCs/macrophages was examined by immunoblotting. Plasma levels of LPO-related metabolites and cytokines were determined by ELISA and the MULTIPLEX platform, respectively. LPO MFI on PBMCs/monocytes were significantly higher in patients (median 4456 and 9091, respectively) compared with HC (1900, p < 0.05, and 4551, p < 0.01, respectively). Patients had higher ferritin expression on PBMCs (mean fold, 1.02) than HC (0.55, p < 0.05). Their ferritin expression levels on PBMCs stimulated with LPO inducers erastin or RSL3 (2.47 or 1.61, respectively) were higher than HC (0.84, p < 0.05, or 0.74, p < 0.01). Ferritin expression on erastin-treated/IL-1β-treated macrophages from patients were higher than those from HC (p < 0.001). The elevated levels of LPO-related metabolites, including malondialdehyde and 4-hydroxyalkenals, were positively correlated with disease activity scores, suggesting LPO involvement in AOSD pathogenesis. Increased ferritin expression on PBMCs/macrophages stimulated with LPO inducers indicates a link between LPO and elevated ferritin.

Low Levels of Zinc and Anti-Oxidant Vitamines (A, C & E) in Patients with Sickle Cell Anemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3781-3781
Author(s):  
Rana M. Hasanato
Keyword(s):  
Serum Level ◽  
Sickle Cell ◽  
Sickle Cell Anemia ◽  
Plasma Levels ◽  
Absorption Spectrometry ◽  
Copper Level ◽  
Control Group ◽  
P Value ◽  
Anti Oxidant ◽  
Normal Controls

Abstract Background : Patients with sickle cell anemia have a higher potential for oxidative damage due to chronic redox imbalance in their red blood cells that often leads to hemolysis, endothelial injury and recurrent vaso-occlusive episodes. In this study, we evaluated plasma levels of the anti-oxidant vitamins A, C,& E and serum level of Zinc along with serum level of Copper (pro-oxidant ). Patients and methods: 25 adult patients with documented severe sickle cell anemia and frequent painful episodes (12 males & 13 females, aged 29.27 +/− 12.94 years) and 25 matched normal controls were studied. Plasma levels of Vitamins A, C, & E were measured by HPLC technique and serum levels of zinc & copper were measured by atomic absorption spectrometry. Results: (Table ): Conclusion These results indicate that levels of Vitamines A, C, and E and Zinc are significantly low in patients with sickle cell anemia in comparison with matched normal controls. Besides, Copper level, which is a well known pro-oxidant, is significantly higher in sickle cell anemia patients. Therefore, supplementing sickle cell anemia patients with anti-oxidant vitamins and trace elements may contribute to the amilioration of symptoms and severity of sickle cell anemia Levels of Vitamine A, C, & E , Zinc & Copper in patients with Sickle cell anemia and normal controls Tested Items SCS patients Control group P-value Results are shown as mean +/− standard deviation., SCA = Sickle cell anemia Vitamin A level mcg/l 0.13 +/− 0.03 0.33 +/−0.05 <0.0001 Vitamin C level mg/l 9.00 +/− 3.43 20.32 +/−3.74 <0.0001 Vitamin E level, mcg/l 1.95 +/− .0.69 4.30 +/−0.80 <0.0001 Zinc level mmol/l 9.00 +/− 1.89 11.81 +/−1.93 <0.0001 Copper level mmol/l 24.58 +/− 4.87 18.42 +/−3.06 <0.0001

Diagnostic Application of Flow Cytometry in Phosphotyrosine Level Detection and Disease Status Evaluation in Patients with Bcr-abl Positive Leukemia.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4431-4431
Author(s):  
Xue-mei Sun ◽  
Jun-hao Chen ◽  
Lei-lei Chen ◽  
Jing-yan Xu ◽  
Yong-gong Yang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Statistical Analysis ◽  
Fusion Gene ◽  
Disease Status ◽  
Leukemia Cell Line ◽  
Control Group ◽  
Relative Value ◽  
Significant Difference ◽  
Normal Controls

Abstract Currently, karyotype analysis for chromosome translocation of 9 and 22 and FISH or gene amplification-based technique for bcr-abl fusion gene have been the major strategies for the diagnosis and disease status evaluation of bcr-abl positive leukemias. However, the method for aberranttyrosine kinase level, which is the reflection of cellular function, is limited to the detection of the level of phosphorylated tyrosine by Western blot. This method is technically laborious, costly and sometimes leads to feigned results. We thus tried to set up a flow cytometry based technique to evaluate its possible utilization in the diagnosis and the evaluation of the disease status in bcr-abl positive leukemias. Mononuclear cells from heparinized bone marrow of patients were collected. After treatment with hemolysin, 106 cells were fixated, permeabilized and then incubated with fluorescein isothiocyanate (FITC) conjugated anti-phosphotyrosine monoclonal antibody (PY20). Fluorescence conjugated microspheres were put in the system as a calibration factor in order to provide a quantitive value for the sake of comparison with different detections. The geomean value of fluorescent microsphere was adjusted to 20 in each case and the ratio of the geomean value of each detected sample to 20 was read as relative value of phosphotyrosine level. This relative value was used in statistical analysis. The human leukemia cell line Mo7e and its derivation Mo7e P210 expressing human bcr-abl fusion gene were employed as negative and positive samples to optimize the experimental conditions. Phosphotyrosine protein in fresh bone marrow cells from patients with chronic myelogenous leukemia (CML) in different phases, bcr-abl positive acute lymphoblastic leukemia (ALL) and normal controls were evaluated by semi-quantitative analysis. A total of 32 CML, including 22 in chronic phase(CP), 3 in accelerated phase(AP) and 7 in blast crisis(BC) were enrolled in this study. Four patients with bcr-abl positive ALL were also included. It was found that there was a significantly different geomean value between Mo7e and Mo7e P210. The relative values of phosphotyrosine levels were different between bcr-abl positive samples and normal controls. The value was 756±144, 865±96, 847±105 and 819±225 with CML patients in CP,AP,BC groups and ALL group respectively, while it was only 18±7 in control group. Statistical analysis showed a significant difference among these five groups. The comparison between each two groups showed a significant difference between each bcr-abl positive group and control group (P< 0.01). There were also significant differences between the AP/CML, BC/CML or ALL group and CP/CML group(P< 0.01). We observed a trend that the value of a given patient escalated with disease progression. In a patient who was on imatinib treatment for one month, there existed two peaks of cells with different fluorescent intensity, each representing a bcr-abl positive or negative cell group, coincident with the FISH result of 158 bcr-abl positive cells in 1000 detected cells. We concluded that flow cytometry analysis of the phosphotyrosine level could be used as an adjuvant technique for the diagnosis and differential diagnosis of bcr-abl positive leukemias. It may also play an important role in the evaluation of disease status or in the judgment of therapeutic efficacy during patient follow-up.

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