Dimitrios Margaritis

Global Impairment of CD4+CD25+FOXP3+ Regulatory T Cells in Idiopathic Pulmonary Fibrosis

RATIONALE The implication of T cells in the pathogenesis of idiopathic pulmonary fibrosis (IPF) is controversial. CD4(+)CD25(+)FOXP3(+) regulatory T cells (Tregs) are pivotal in maintaining immune homeostasis, but their role in IPF pathophysiology has not yet been studied. OBJECTIVES To explore Treg dynamics and function in IPF. METHODS Treg levels and dynamics were analyzed by flow cytometry in the peripheral blood (PB) and bronchoalveolar lavage (BAL) of 21 patients with IPF, 35 patients with lung diseases other than IPF (patients without IPF), 20 patients with collagen vascular diseases with pulmonary parenchymal involvement (CVD-IP), and 28 healthy volunteers. The suppression of autologous CD4(+)CD25(-) cell-proliferative responses and cytokine release by magnetic bead-isolated Tregs was evaluated by proliferation assays and cytometric bead array. Correlations of Treg function and levels with lung function parameters were also performed. MEASUREMENTS AND MAIN RESULTS In patients with IPF, both BAL and PB Tregs were reduced compared with those of healthy volunteers and patients without IPF, although not always significantly. Treg levels were not affected by the administration of low-dose prednisone in four nonresponding patients. The suppressor potential of BAL and PB Tregs was compromised in patients with IPF and patients with CVD-IP, compared with healthy volunteers and patients without IPF. Similarly, the Treg-induced suppression of helper T-cell type 1 and 2 cytokine secretion was impaired in the BAL of patients with IPF and patients with CVD-IP. Moreover, the defective function of BAL Tregs correlated highly with parameters of disease severity. CONCLUSIONS This study provides the first evidence of global Treg impairment in IPF that strongly correlates with disease severity, suggesting a role for Tregs in the fibrotic process.

Circulating angiopoietin‐1 to angiopoietin‐2 ratio is an independent prognostic factor for survival in newly diagnosed patients with multiple myeloma who received therapy with novel antimyeloma agents

The circulating levels of several angiogenic cytokines [angiopoietin‐1 (Ang‐1), angiopoietin‐2 (Ang‐2), vascular endothelial growth factor (VEGF), angiogenin and basic fibroblast growth factor (bFGF)] were evaluated in 174 consecutive patients with newly diagnosed, symptomatic, multiple myeloma (MM). Circulating levels of Ang‐1/Ang‐2 were reduced in myeloma patients compared to controls, whereas VEGF and angiogenin levels were increased. Reduced angiopoietin‐1/angiopoietin‐2 ratio correlated with advanced disease features including international staging system (ISS)‐3 stage, renal impairment and extensive bone disease. Based on immunohistochemical results in 20 patients (10 with the higher and 10 with the lower values of circulating angiopoietin‐2) we found that angiopoietin‐2 is expressed by myeloma cells and correlates with increased microvessel density in subsets of patients. Furthermore, Ang‐1/Ang‐2 ratio correlated with survival. Patients with circulating Ang‐1/Ang‐2 below or equal to the median value (6.03) had a median survival of 26.3 months compared to 53 months of all others (p = 0.002). Interestingly, this was mainly observed in patients who received first‐line therapy with novel agent‐based regimens (65% of our patients). Furthermore, a subset of ISS‐3 patients with serum Ang‐1/Ang‐2 above the median value had favourable prognosis (median survival: 45 months versus 17 months of all others; p = 0.0001). The multivariate analysis revealed that low Ang‐1/Ang‐2 ratio could independently predict for inferior survival in our cohort of patients (relative risk (RR) 2.07, 95% CI 1.50–2.42; p < 0.001). These results highlight the role of angiopoietins pathway in the biology of MM and reveal novel targets for the development of antimyeloma agents.

Dynamics of telomere's length and telomerase activity in Philadelphia chromosome negative myeloproliferative neoplasms

Telomere exhaustion and increased telomerase activity are associated with the acquisition of aggressive molecular events in a variety of haematological malignancies. In Philadelphia chromosome negative myeloproliferative neoplasms (Ph(neg)MPNs), telomere dynamics during clonal evolution of these diseases have not yet been fully elucidated. Herein we demonstrated that telomere shortening is a global phenomenon in Ph(neg)MPNs, irrespective of disease phenotype, treatment administration and JAK2V617F mutational status but the presence of additional cytogenetic abnormalities further affects them. Consistent with the above finding, TA was upregulated in CD34+ haemopoietic progenitors from almost all Ph(neg)MPN subgroups compared to healthy donors. Moreover, TL below the cut-off value of 27% could predict disease progression in Ph(neg)MPN patients (PFS at 5 years 39% vs 81%). Thus, TL emerges as a new prognostic marker in Ph(neg)MPN, reflecting probably the genetic instability of highly proliferating MPN clones.

Cardiovascular involvement in patients with β-thalassemia major without cardiac iron overload

BACKGROUND Cardiovascular complications are common in beta-thalassemia major (beta-TM), mainly attributed to increased cardiac iron depositions. Early cardiovascular involvement in patients without cardiac symptoms and without cardiac iron overload has not been adequately investigated. METHODS Twenty six patients (11 males) with beta-TM, on chelation therapy, age 23+/-4 years without cardiac iron overload (measured by magnetic resonance imaging), and 30 age and gender matched healthy controls were included in the study. Carotid-femoral and carotid-radial pulse wave velocity (PWVc-f and PWVc-r) and augmentation index (AI) were measured by SphygmoCor device; carotid intima-media thickness; left ventricular (LV) dimensions and function; left atrial (LA) volume and function were assessed by echocardiography. RESULTS Patients with beta-TM had higher PWVc-f (8.4+/-1.4 vs 7.2+/-1.1 m/s, p=0.002) and augmentation index (21.7+/-10.9 vs 14.7+/-9.7%, p=0.04) indicating decreased aortic elastic properties; greater LV mass index (72.0+/-13.3 vs 63.8+/-11.5 g/m(2), p=0.04) and greater LA volumes. Multivariate logistic regression analysis revealed that higher PWVc-f was independently associated with higher LV mass [OR 1.74 95%CI (1.09-2.88), p=0.026]; and greater LA dimensions [OR 1.68 95%CI (1.04-2.72), p=0.035]. CONCLUSIONS In the absence of cardiac iron overload, asymptomatic patients with beta-TM demonstrated aortic stiffening associated with increased LV mass and LA enlargement. These alterations may represent signs of early cardiovascular involvement.

Hydroxyurea (HU) is effective in reducing JAK2V617F mutated clone size in the peripheral blood of essential thrombocythemia (ET) and polycythemia vera (PV) patients

Ph-negative chronic myeloproliferative disorders (PhnegcMPD) are treated according to the estimated vascular risk. The recent discovery of V617F point mutation of the JAK2 kinase, which frequently occurs in these diseases, has not changed their management so far. However, emerging data tend to support a prothrombotic role for the mutation, along with a better response of JAK2V617F mutated patients to hydroxyurea treatment. Our data further support this notion.

CD1d expression as a prognostic marker for chronic lymphocytic leukemia

Abstract We analyzed the expression of CD1d, an antigen-presenting molecule, on peripheral blood leukemic cells of cases of chronic lymphocytic leukemia (CLL) by flow cytometry. We demonstrated variable expression of CD1d on leukemic lymphocytes and an association between high expression of CD1d with shorter time to treatment and overall survival of patients. CD1d was positively associated with CD38 expression, but not with unmutated heavy chain variable (VH) mutational status or adverse cytogenetics of leukemic lymphocytes. Our findings support that CD1d expression is a prognostic marker for CLL.

Oxidised low‐density lipoprotein and arterial function in β‐thalassemia major

Objectives:  Vascular abnormalities such as endothelial dysfunction and arterial stiffness have been described in patients with β‐thalassemia major (β‐TM). Increased concentrations of oxidised low‐density lipoprotein cholesterol (oxLDL) have been observed in those patients, but possible associations between oxLDL and arterial function in β‐TM have not been defined.

Effect of HbS in the determination of HbA2 with the Menarini HA-8160 analyzer and comparison with other instruments

It is known that the presence of hemoglobin S (HbS) affects the determination of hemoglobin A2 (HbA2) levels in clinical samples. We quantitated this effect using the Menarini HA‐8160 analyzer and compared with other instruments (HELENA beta‐thal quik column, TOSOH HLC‐723G7 and BIORAD Variant II) using the HELENA SAS‐MX alkaline gel electrophoresis kit as the reference method. The %HbA2 values from the HA‐8160 analyzer and the alkaline gel electrophoresis show a good linear correlation in the absence of HbS. A strong positive bias in the %HbA2 values from the HA‐8160 is apparent in the presence of HbS in the samples, when compared with the alkaline electrophoresis. The analytical imprecision and bias of the three HPLC instruments are comparable both in the presence and absence of HbS. The manual column method shows a lower bias in the absence of HbS but is more affected when HbS is present in the samples.

Successful treatment of lymph node extramedullary plasmacytoma with bortezomib

Dear editor, The extramedullary plasmacytomas (EMPs) are localized mainly in the upper respiratory tract. Lymph node plasmacytoma is defined as a lymph node tumor composed of monoclonal proliferation of plasma cells. The diagnosis of lymph node EMP is based not only on the finding of a plasma cell tumor at an extramedullary site (lymph node) but also on the exclusion of multiple myeloma (MM). Radiotherapy with or without local resection is the current treatment of choice because these tumors are usually radiosensitive. With radiation doses of 4,000–5,000 cGy, local recurrence develops in 5%, and systemic progression develops in less than 30% of the patients [1]. The size of EMP has been reported to be a bad prognostic factor regarding the likelihood of control [2]. Therefore, the treatment of large EMP presented in other sites than the upper respiratory system remains a challenge. Chemotherapy is considered as a treatment option in systemic progression to MM or recurrent relapses. The proteasome inhibitor bortezomib has recently been evaluated in the setting of relapsed and therapy–refractory plasmacytic malignancies with some evidence of effectiveness in MM with extramedullary manifestations of the disease [3, 4]. We report here a patient suffering from refractory EMP who received bortezomib and achieved complete remission. A 63-year-old male patient presented with retroperitoneal mass, measuring 8×10 cm, and after total surgical excision, histological examination of the mass, and a bone marrow biopsy, as well as a detailed biochemical and radiological evaluation to exclude MM, the patient was diagnosed as having retroperitoneal lymph node EMP (previously reported by Pantelidou et al. 2005 [5]). Despite radiotherapy to a total dose of 4,400 cGy, he relapsed 20 months later. Computed tomography (CT) disclosed seven enlarged paraaortic lymph nodes with diameters of 1.0– 3.8 cm on axial images. Bone marrow aspiration and biopsy, detailed biochemical evaluation, and skeletal survey did not reveal any evidence of systemic MM. Four courses of vincristine, adriamycin, and dexamethasone (VAD) followed by high-dose melphalan with autologous stem cell transplantation were given, and partial remission was achieved. Unfortunately, the patient relapsed again after 4 months. The CT demonstrated enlargement of celiac axis, retropancreatic, pericaval, and periaortic lymph nodes; a total of 22 enlarged lymph nodes were identified, measuring 1.0–4.2 cm in diameter on axial images. The involved nodes exhibited homogeneous soft tissue densities and mild enhancement after iodinated contrast media administration (Figs. 1a, 2a). Based on the early relapse after high-dose melphalan supported by autologous stem cell transplantation and the extended lymph node involvement, treatment with the proteasome inhibitor bortezomib was considered. After obtaining the patient’s informed consent, he received bortezomib (1.3 mg/m) as intravenous bolus injections D. Pantelidou (*) . C. Tsatalas . D. Margaritis . A. G. Anastasiadis . G. Bourikas Hematology Department, School of Medicine, Democritus University of Thrace, Alexandroupolis, Greece e-mail: dpantelidou@yahoo.gr Tel.: +30-25510-74511 Fax: +30-25510-76154

Pregnancy in β-thalassemia trait carriers: an uneventful journey

Abstract Normochromic normocytic anemia during pregnancy reflects the significant increase in plasma volume, which disproportionately exceeds the increase in the red cell volume. In β-thalassemia (β-thal) trait carriers who become pregnant the plasma volume expansion may cause more pronounced anemia because the anemia of pregnancy is added to the pre-existed hypochromic microcytic anemia. In β-thal women, pregnancy outcome and obstetric complications do not differ from the general population. Anemia in β-thal carriers is generally not severe enough to warrant anxiety. No specific therapy is indicated and pregnant women generally require only supportive care with an anticipated favorable pregnancy outcome.