First report of expansion of CD4+/CD28 null T-helper lymphocytes in adult patients with idiopathic autoimmune hemolytic anemia

ABSTRACT CD28 null T helper (Th) cells are rare in healthy individuals, but they are increased in various inflammatory and immune-mediated diseases. In this study, we determined the size of the CD4+/CD28 null T lymphocyte compartment in the peripheral blood of 40 autoimmune hemolytic anemia (AIHA) patients (idiopathic and secondary) and 20 healthy control subjects, using tri-color flow cytometry. The frequency and absolute count of CD4+/CD28 null T helper (Th) cells was significantly higher in idiopathic AIHA patients, compared to healthy controls (p = 0.001 and 0.001, respectively) and to patients with secondary AIHA (p = 0.04 and 0.01, respectively). The percentage of CD4+/CD28 null Th cells was also negatively correlated to the hemoglobin (Hb) level (p = 0.03). These findings demonstrate, for the first time, the expansion of this phenotypically-defined population of T lymphocytes in patients with idiopathic AIHA and indicate that it likely plays an etiological role in the development of this disease. However, establishing the use of this marker for diagnosis or monitoring treatment of such patients needs further studies.

Erythroferrone Expression in Anemic Rheumatoid Arthritis Patients: Is It Disordered Iron Trafficking or Disease Activity?

PURPOSE: Erythroferrone (ERFE) is well acknowledged for its inhibitory function on hepcidin synthesis in the liver during stress erythropoiesis, thereby ensuring sufficient iron supply to bone marrow erythroblasts. Hepcidin plays an indispensable role in the pathogenesis of anemia of chronic disease (ACD). Thus, ERFE was suggested to protect against ACD in various diseases. Rheumatoid arthritis (RA) is commonly involved with ACD and high hepcidin levels, with a further increase of the latter in active states. The present study is a case-control study that aimed to determine the pattern of ERFE expression in RA patients with concomitant ACD and study its relationship with hepcidin, erythropoietin (EPO) and disease activity. PATIENTS AND METHODS: Fifty-five RA patients with ACD were categorized into active and inactive RA using the disease activity score (DAS28); 15 healthy subjects were included as control subjects. ERFE was measured for patients and control subjects using quantitative real-time polymerase chain reaction, in addition to testing for CBC, ESR, CRP, iron profile parameters and hepcidin. EPO was assessed for patients of both active and inactive RA groups. RESULTS: ERFE and hepcidin showed the highest levels in active RA; ERFE values were similar in control subjects and inactive RA patients, while hepcidin was significantly higher in inactive RA than control subjects. Patients with high ERFE levels had higher RBC, Hct, MCV, hepcidin and EPO levels. Stepwise regression analysis has identified DAS28 and disease duration as the best predictors of ERFE values, whereas ERFE and hepcidin were independent predictors of disease activity. CONCLUSION: We introduce ERFE as a novel marker of RA activity. Although the inhibitory effect of ERFE on hepcidin is not evident, our results still indicate that ERFE may have a beneficial erythropoietic effect in the context of ACD in RA disease activity.

First report of expansion of CD4+/CD28 null T-helper lymphocytes in adult patients with idiopathic autoimmune hemolytic anemia.

CD28 null T helper (Th) cells are rare in healthy individuals, but they are increased in various inflammatory and immune-mediated diseases. In this study, we determined the size of the CD4+/CD28 null T lymphocyte compartment in the peripheral blood of 40 autoimmune hemolytic anemia (AIHA) patients (idiopathic and secondary) and 20 healthy control subjects, using tri-color flow cytometry. The frequency and absolute count of CD4+/CD28 null T helper (Th) cells was significantly higher in idiopathic AIHA patients, compared to healthy controls (p = 0.001 and 0.001, respectively) and to patients with secondary AIHA (p = 0.04 and 0.01, respectively). The percentage of CD4+/CD28 null Th cells was also negatively correlated to the hemoglobin (Hb) level (p = 0.03). These findings demonstrate, for the first time, the expansion of this phenotypically-defined population of T lymphocytes in patients with idiopathic AIHA and indicate that it likely plays an etiological role in the development of this disease. However, establishing the use of this marker for diagnosis or monitoring treatment of such patients needs further studies.

Effect of intra-cellular trafficking on flow cytometric measurement of neutrophil's oxidative status in iron deficient pregnant females.

BACKGROUND: Iron deficiency and iron deficiency anemia are prevalent among pregnant women particularly in developing countries. This study aimed to evaluate the iron status among Egyptian pregnant women and its impact on their neutrophil's count and antimicrobial functions. METHOD: Ninety pregnant females underwent complete blood count, iron profile, flow cytometric studies for neutrophil myeloperoxidase expression & oxidative burst using dihydrorhodamine 123 (DHR) after phorbol-12-myristate-13-acetate (PMA) stimulation as well as neutrophil phagocytic and lytic indices. RESULTS: According to percent saturation 54/90 women (60%) were iron deficient (<15% saturation) (cases) and 36/90 (40%) were iron sufficient (controls). A higher proportion of iron deficient pregnant women were in their third trimester compared to controls. No significant difference was found between the iron deficient & sufficient groups as regards anemia despite a positive correlation between haemoglobin level and percent saturation (P=.02). Both the phagocytic and lytic indices were significantly lower among the cases compared to controls (P=.014 & .002 respectively). Cases and controls were comparable as regards flow cytometric studies of neutrophils' myeloperoxidase and oxidative burst (P>.05). No significant correlation was found between any of the iron profile parameters and the oxidative burst by flow cytometry. CONCLUSION: Functional microphage assay (phagocytic and lytic indices) may be more relevant and cost effective than flow cytometry assays of myeloperoxidase and oxidative burst in reflecting either iron status or cellular immunity in pregnancy.

Seroprevalence of anti-WNV IgG antibodies and WNV-RNA in Egyptian blood donors.

Transmission of West Nile virus (WNV) from asymptomatic donors has been reported during blood transfusions and organ transplants in humans. In this work, we aimed to investigate the presence of WNV antibody and WNV RNA in blood donors to evaluate the sero-prevalence of WNV and risk for WNV transmission. One hundred and sixty blood donors were tested for the presence of anti-WNV IgG by ELISA and for WNVs 1 and 2 RNA by RT-PCR. About 55% of blood donors were seropositive for WNV IgG antibodies, with significantly higher percentage of positive donors coming from rural areas and Nile Delta region compared to other donors. Using RT-PCR all donors were negative for viral RNA of both WNV lineages 1 and 2. High sero-prevelance of WNV antibodies in asymptomatic blood donors denotes endemicity of the WNV in Egypt and points to the importance of routine screening of blood donors for WNV RNA. On the other hand the absence of WNV RNA by RT-PCR indicates apparent low risk of the blood products as regards WNV transmission. Further studies into significance of WNV seronegativity among Rh negative donors and into the use of WNV seropositive blood in prophylaxis or treatment of WNV neuroinvasive disease are recommended. J. Med. Virol. 89:1323-1329, 2017. © 2017 Wiley Periodicals, Inc.

Clinical to Molecular Screening Paradigm for ß-Thalassemia Carriers.

ß-Thalassemia (ß-thal) represents a major health problem worldwide and particularly in Egypt. Its prevention, compared to treatment, is cost-effective, possible and practical. In this study we evaluate a proposed paradigm for detection of the ß-thal carrier state. The present study included 1627 children and adolescents of both sexes, presenting as outpatients to clinics of Ain-Shams University Hospitals, Cairo, Egypt, from November 1 2009 to June 30 2010. In the first phase, after performing a complete blood count (CBC), 280 microcytic hypochromic patients were selected. These cases were further analyzed by iron profile and high performance liquid chromatography (HPLC); in the second phase, hybridization detected 22 common ß-globin mutations in 74.0% of the suspected cases. Thus, by HPLC, the Hb A2 level of >3.5% provided 100.0% sensitivity, 70.0% specificity, 75.0% positive predictive value (PPV), 100.0% negative predictive value (NPV) and accuracy of 70.0% to identify ß-thal trait and at a cut-off of 4.0%, it provided 97.4% sensitivity, 72.7% specificity, 92.6% PPV, 88.8% NPV and a diagnostic accuracy of 92%. High performance liquid chromatography is a reliable and cost effective primary screening tool for ß-thal trait at a Hb A2 level of ≥4.0%, while molecular testing is mandatory only for selected cases with borderline Hb A2 values between 3.5 and 4.0%.

Detection of 14q32 rearrangements in multiple myeloma, using simultaneous FISH analysis combined with immunofluorescence.

BACKGROUND: 14q32 rearrangement has been identified as a recurrent hotspot of translocations in multiple myeloma (MM). The Fluorescence Immunophenotyping and Interphase Cytogenetics as a tool for the Investigation of Neoplasms (known as FICTION technique) for evaluation of chromosomal changes in MM. The aim of this work is to detect 14q32 rearrangement, using FICTION technique, on archival bone marrow (BM) slides of MM patients, and to study its prognostic value. METHOD: This study was conducted at Ain Shams University Hospital. The FICTION technique, which uses CD138 and dual color, and the break-apart 14q32 rearrangement probe, was performed on archived smears of BM slides for 50 MM patients at the time of diagnosis. RESULTS: A significantly higher percentage of cases were positive for 14q32 rearrangement by FICTION (32%) compared to fluorescence in situ hybridization (FISH) (12%) (p=0.04). Cases positive by FICTION for the rearrangement were designated as Group A, while negative cases were designated as Group B. Significantly lower Hb and CRP levels were found among Group B when compared to Group A patients (p=0.001 and 0.01, respectively). Serum albumin level and Bence Jones protein (BJP) significantly affect overall survival (OS) (p=0.01, 0.007, respectively). However, a statistically non-significant shorter mean survival time was found in positive cases through FICTION versus negative cases. CONCLUSION: FICTION technique provides a sensitive tool for establishing clonal plasma cells (PC) infiltration of BM aspirates, and is amenable for use on archived as well as fresh smears.

Effect of passive smoking on blood lymphocyte apoptosis in children.

BACKGROUND: Passive smoking is a well-known risk factor for both recurrent respiratory infections and disturbed lipid profile. Whether passive smoking problems are related to altered lymphocyte survival and its relation to altered lipid profile are the points of concern in this work. MATERIALS AND METHODS: Urinary cotinine and creatinine levels as well as lipid profile and flow cytometric assessment of apoptosis of peripheral blood lymphocytes (PBL) were assessed in 26 children with history of indoor exposure to cigarette smokers in comparison with 14 matched children with no such history. RESULTS: Lipid profile showed significantly higher mean levels of triglycerides, cholesterol and low-density lipoprotein (LDL) and significantly lower mean levels of high-density lipoprotein (HDL) in passive smoking children compared to nonpassive-smoking ones. Furthermore, cotinine parameters were positively correlated with triglycerides and LDL and negatively correlated with HDL. Early apoptosis of PBL was significantly higher in exposed vs nonexposed ones. CONCLUSIONS: Passive smoking in children could be a risk factor for enhanced lymphocytic apoptosis. It is possible that altered lipid profile may play a role in the increased risk. The impact of this lymphocytic derangement on increased frequency of infections is noticeable.

The prognostic potential of bone marrow cyclin E and P27, in Egyptian patients with acute myeloid leukemia.

Cyclin E and the cyclin dependent kinase inhibitor P27 are two important regulators of the G1-S transition modulating the activity of cyclin-dependent kinases. Aberrations in the cell cycle control are often observed in tumors and might even be mandatory in tumor development. There are few molecular biologic determinants that may be prognostic for patients with acute myeloid leukemia (AML). To investigate the importance of cell cycle defects in AML, the cellular levels of cyclin E and the cyclin-dependent kinase inhibitor P27 (Kip 1) were evaluated in thirty AML patients (11 males and 19 females) diagnosed by standard clinical, morphological and immunophenotypic criteria and staged according to the FAB classification. Using immunoblot analysis, cyclin E and P27 were detected in blast cells of AML patients who were then treated by the standard AML chemotherapeutic protocol and were followed up. With respect to cyclin E, it was detected in 9/30(30%) AML cases among them 13.3% (4/30 cases) exhibited very strong bands while 16.6% (5/30 cases) showed faint bands. Cyclin E was high among M4/M5 cases and low among M3 cases and showed a statistically significant positive correlation with percentage of blast cells, aberrant phenotype and abnormal karyotype at diagnosis. It also showed a significant negative correlation with complete remission (CR) rates. All our AML cases exhibited P27 at low and high levels as seen in 19/30 (63.4%) and 11/30 (36.6%) cases, respectively. P27 showed a statistically significant negative correlation to the percentage of blasts at diagnosis and a significant positive correlation with achievement of CR. A significant negative correlation between P27 and cyclin E (P < 0.004) was observed as well. The present study suggested that levels of cell cycle regulators cyclin E and P27 can be used as a useful prognostic molecular markers in AML patients.