Athens University thalassemia expertise unit: evolution, structure, perspectives and patients' expectations.

Thalassemia Expertise Centres (TECs), were first organized in developed countries with high thalassemia prevalence in the 70's to meet the increasing demands of the implementation of frequent transfusions in the treatment of thalassemia, and to consequently adopt, the rapid advances in the management of the disease. Recent evaluation of longitudinal implementation of the national programs for prevention and treatment, demonstrated their efficacy for patients and public health. The beneficial effects focused on clinical symptoms amelioration, reduction of incidence and severity of complications and considerable improvement in survival, quality of life and social adaptation.National programs leaded to the modification of the most common genetic, fatal pediatric disease with short survival, to a chronic long-lived disease for adults and a very rare disease for children. In the few developed countries new perspectives for pediatric TECs need to be considered.

Efficacy and tolerability of formoterol Turbuhaler in children.

A randomised, double-blind trial was undertaken to investigate the efficacy and tolerability of formoterol Turbuhaler in children with mild to moderate asthma. After a two-week run-in, 248 children aged 6-17 years were randomised to receive formoterol 4.5 and 9 pmicro b.i.d. or placebo for 12 weeks. Morning PEF (primary variable), was significantly improved versus placebo only in the formoterol 9 pmicro b.i.d. group (13 l/min, 95% CCI 1.9, 24.2%; p = 0 .02). Both formoterol 4.5 and 9 pmicro significantly increased the pre-bronchodilator FEV1 by 5.2-6.7% (p < 0 .05) and reduced use of daytime relief medication versus placebo (p < 0 .05). Formoterol 9 pmicro significantly reduced night-time reliever use and awakenings due to asthma versus placebo (p < 0.05). Both doses of formoterol were as well tolerated as placebo. In conclusion, formoterol 4.5 and 9 micro b.i.d. is effective and well tolerated as maintenance therapy in children with mild to moderate asthma.

Heart failure in beta thalassemia: a 5-year follow-up study.

PURPOSE: To evaluate the survival of patients with beta thalassemia and heart failure who were treated with iron chelation therapy. SUBJECTS AND METHODS: Fifty-two consecutive patients with beta thalassemia and heart failure were followed in a prospective 5-year study. All patients underwent a full clinical examination with chest radiograph, electrocardiogram, and echocardiographic investigation performed at 6-month intervals or when a new symptom developed. RESULTS: Of the 52 patients (mean [+/- SD] age, 24 +/- 5 years), 25 (48%) survived 5 years after the onset of heart failure. Forty-three patients had left-sided heart failure, and 9 had right-sided heart failure. Those with left-sided heart failure were younger at presentation with heart failure (22 +/- 4 years vs. 31 +/- 6 years; P <0.001), had lower ejection fractions (36% +/- 9% vs. 64% +/- 10%; P <0.001), and had a lower mean serum ferritin level (3355 +/- 1241 ng/mL vs. 6,397 +/- 1,613 ng/mL; P <0.001). CONCLUSION: The 5-year survival rate in patients with beta thalassemia with heart failure was greater than previously reported. There are clinical characteristics that may make patients more likely to develop left- or right-sided heart failure.

Reactogenicity of DTPa-HBV/Hib vaccine administered as a single injection vs DTPa-HBV and Hib vaccines administered simultaneously at separate sites, to infants at 2, 4 and 6 months of age.

An open, randomised, multicentre trial was performed to assess the reactogenicity and safety profile of the administration of a candidate Haemophilus influenzae type b (Hib) conjugate vaccine with a quadrivalent diphtheria-tetanus-acellular pertussis-hepatitis B (DTPa-HBV) vaccine as a single injection (Group 1) versus the simultaneous administration of the latter vaccine (DTPa-HBV) and an available Hib conjugate vaccine (Group 2) in opposite thighs, as a primary vaccination course to healthy infants at 2, 4 and 6 months of age. Eight hundred and eighty five infants (9.3+/-1.4 weeks old) were randomly allocated to Group 1 (n=665) and Group 2 (n=221). Oral polio vaccine was given concomitantly to all subjects. Blood samples (pre-vaccination and 1 month after the third dose) were obtained from a subset of infants (Group 1, 73; Group 2, 22) for serological determinations. Local and general symptoms were recorded by parents on diary cards. 2614 diary cards (Group 1, 1966; Group 2, 648) were collected. There were no statistically significant differences in the incidence of local and general symptoms between groups. Pain such that the infant cried when limb was moved was reported in 0.6 and 0.2% in groups 1 and 2, respectively. Redness and swelling (>20 mm in diameter) were recorded between 2.1 and 3% in both groups. Fussiness preventing normal activities was the most frequently reported general symptom in both groups (1.6 and 1.9% in groups 1 and 2, respectively). Fever (rectal temperature >39.5 degrees C) was reported in 0.4% (Group 1) and 0.3% (Group 2). All subjects included in the immunogenicity analysis had seroprotective or seropositive titres to the diphtheria, tetanus, hepatitis B and pertussis components of the vaccines. About 99 and 100% of infants had anti-PRP titres > or =0.15 mcg/ml in groups 1 and 2, respectively. This study indicates that DTPa-HBV vaccine given in a single injection with a candidate Hib conjugate vaccine has a similar reactogenicity profile to that of two commercially available vaccines (DTPa-HBV, Hib) given in two simultaneous injections to infants 2, 4 and 6 months of age.

Hepatitis in children hospitalized with measles: the experience acquired after a Greek epidemic.

OBJECTIVE: In childhood, hepatitis is an uncommon and ill-defined complication of measles. We studied prospectively the prevalence of hepatitis in 189 children with measles, admitted to hospital during a measles epidemic in Greece. METHODOLOGY: Diagnosis of measles was based on clinical features and a fourfold rise of the haemagglutination inhibiting antibody titre, while liver impairment was based on a twofold or greater increase in liver enzymes. RESULTS: Nine children (4.8%) had increased liver enzymes. Hepatitis was not related to the duration and severity of fever or the coexistence of other complications, and in all children but one, was subclinical and resolved rapidly. One child with mental retardation who was being treated with anti-epileptic therapy and had normal liver enzymes prior to measles, developed hepatic coma from which he recovered 1 month later. CONCLUSIONS: Liver involvement in childhood measles is rare and transient but it may be severe in children receiving hepatotoxic drugs.

Different geographic origins of Hb Constant Spring [alpha(2) codon 142 TAA-->CAA].

BACKGROUND AND OBJECTIVES: The occurrence of Hb CS is usually limited to the geographic area which includes Southern China and South East Asia. In 1968 Hb CS was also found to occur in the Mediterranean area where it was originally described as Hb Athens. We investigated the independent origin of these termination codon mutations of the alpha 2-globin gene by determining the alpha-cluster haplotype and comparing the hematologic data from Hb CS-Hb H patients and their family members. DESIGN AND METHODS: We studied one Hb CS-Hb H patient of Greek origin and a Sicilian family in which one individual was affected by Hb CS-Hb H. The haplotype of the Hb CS allele was determined and compared to the haplotype of an Hb CS-Hb H individual of Chinese origin. RESULTS: The haplotype found for the Greek and Sicilian Hb CS was the same but differed significantly from the Asiatic Hb CS mutation. INTERPRETATION AND CONCLUSIONS: The Hb CS mutation found in both Mediterranean patients arose independently in the Mediterranean area. The difference in clinical manifestation of the Hb CS-Hb H disease in both patients is less common but consistent with similar variation in the clinical expression of analogous Hb Icaria-Hb H disease patients.

Variations of ferritin levels over a period of 15 years as a compliance chelation index in thalassemic patients.

We studied the changing pattern of the distribution of ferritin levels in 430 regularly-transfused patients with thalassemia in an attempt to evaluate compliance of chelation with deferoxamine. The study covered 15 years and was divided in three periods: 1981-1985, 1986-1990, and 1991-1995. The patients were stratified in age-groups. The mean ferritin levels of each period were calculated for each patient individually. The study showed that: (i) When all the patients were compared as a group, there was a significant decrease in mean ferritin between 1981-1985 and 1991-1995, despite a significant change in the patients' mean age; (ii) When patients of same age were compared between periods, there was a decrease in mean ferritin between 1981-1985 and 1991-1995, as well as a decrease in the proportion of patients with ferritin >4000 microg/L, with a parallel increase in the proportion of patients who had ferritin <2000 microg/L; (iii) When the same patients were followed longitudinally, they showed a decrease in their ferritin levels in all age groups with the exception of the late adolescence period. The decrease in iron overload observed in patients on close follow up implies that compliance with chelation therapy has improved with time and therefore, a favourable influence in survival could be expected.

Phenotypic and molecular diversity of haemoglobin H disease: a Greek experience.

Haemoglobin H (Hb H) disease is the severest form of alpha-thalassaemia compatible with post-natal life and occurs when alpha-thalassaemia mutations interact to reduce alpha-globin synthesis to levels approximately equivalent to the output of a single alpha-globin gene. Hb H disease has variable clinical expression, mainly related to underlying genotypes. The spectrum of alpha-thalassaemia determinants in Greece appears greater than in any other population studied and, in 75 Greek Hb H disease patients, we found 12 alpha-thalassaemia mutations interacting to produce 15 Hb H disease genotypes. Evaluation of haematological, biochemical and clinical findings, and correlation with genotypes, defined genetic predictors of disease severity and factors involved in disease progression. In accordance with previous reports, patients with non-deletion alpha-thalassaemia mutations had more severe clinical expression. Additionally, we found that all patients with the most severe phenotypes had alpha-thalassaemic globin variants. Phenotypic severity was not simply related to the degree of alpha-globin deficiency: high Hb H levels were found to exacerbate anaemia by negatively influencing tissue oxygenation, and both Hb H and alpha-thalassaemic haemoglobin variants appear to reduce red cell survival within the bone marrow and circulation. Together with the long-term follow-up in many patients, this report provides comprehensive information for management of Hb H disease and appropriate family counselling.

Molecular studies of beta-thalassemia heterozygotes with raised Hb F levels.

Hb F levels in beta-thalassemia heterozygotes are usually less than 2%, but amongst 1,059 patients studied, 73 (7%) had Hb F levels above 2.5% (2.6-14.0%). To investigate factors that may influence the increase of Hb F levels in these heterozygotes, we characterized the beta-thalassemia mutations and their chromosomal background, gamma-globin gene promoter variations, and alpha-globin genotypes. All 73 beta-thalassemia heterozygotes carried beta-thalassemia point mutations previously observed in the Greek population; gene mapping excluded b gene cluster deletions; only two cases had an additional gamma-globin gene (gammagammagamma/gammagamma). Five alpha-globin genes (alphaalphaalpha/alphaalpha) were detected in 17/73 cases (23%) as compared to a carrier rate of 1.76% in the general population. Molecular, hematological, and biosynthetic findings in these compound heterozygotes indicated that the raised Hb F levels were caused by cell selection due to ineffective erythropoiesis. In the remaining 56 simple beta-thalassemia heterozygotes, 11 beta-thalassemia mutations were observed, each on the expected haplotype(s), and analysis of the gamma gene promoters revealed three known polymorphisms (in linkage disequilibrium), with minimal influence on gamma-globin levels. However, the overall distribution of beta-thalassemia mutations in the 56 simple beta-thalassemia heterozygotes was significantly different (P<0.0002) compared to that in 986 simple beta-thalassemia heterozygotes with <2.5% Hb F, implicating an association between beta-thalassemia mutations and moderately increased Hb F levels, most notably codon 39 (C-->T), IVS-II-1 (G-->A), codon 6 (-A), and codon 8 (-AA), which accounted for 41/56 (73%) cases with >2.5% Hb F. In the remaining 15/56 (27%) cases, no common underlying globin genotypes could explain the raised Hb F levels. Overall, this study indicates that the control of Hb F levels in beta-thalassemia heterozygotes is heterogeneous and multi-factorial.

Non-invasive myocardial iron assessment in thalassaemic patients. T2 relaxometry and magnetization transfer ratio measurements.

PURPOSE: To compare T2 relaxometry and magnetization transfer ratio (MTR) measurements of myocardial tissue in normal volunteers and thalassaemic patients for assessment of the myocardial iron levels. MATERIAL AND METHODS: All examinations were done on a 1 T MR system using a multi-echo spin-echo sequence with 8 echoes for T2 measurements and a gradient echo sequence for MTR measurements. Diastolic cardiac triggering was used in both sequences. Ten patients and 10 normal subjects were included in the study. T2 and MTR measurements were correlated with serum ferritin levels. RESULTS: Regression analysis between T2 and MTR measurements and ferritin demonstrated a reversed linear relationship, (r=-0.932, p<0.05) and (r= -0.824, p<0.05), respectively. Mean T2 relaxation time and mean MTR of the normal subjects (57.95+/-4.9 ms and 43.70+/-3.3%) was significantly higher than that of the thalassaemic patients (38.8+/-6.2 ms and 26.40+/-6.1%) (p<0.01), respectively. CONCLUSION: MTR measurements can be used to complement T2 measurements for non-invasive myocardial iron assessment.

Mucoepidermoid carcinoma of the bronchus.

Bronchoscopy in a 4.5-year-old girl with recurrent pneumonia showed an exophytic endobronchial mass. Biopsy disclosed microscopic and ultrastructural features of a low-grade mucoepidermoid carcinoma. Complete cure was accomplished by surgical removal of the tumor and right lower lobe.

Molecular, haematological and clinical studies of the -101 C --> T substitution of the beta-globin gene promoter in 25 beta-thalassaemia intermedia patients and 45 heterozygotes.

We report the clinical, haematological, biosynthetic and molecular data of 25 double heterozygote beta-thalassaemia intermedia patients and 45 beta-thalassaemia heterozygotes with the C --> T substitution at nucleotide position -101 from the Cap site, in the distal CACCC box of the beta-globin gene promoter. This mutation is considered the most common amongst the silent beta-thalassaemia mutations in Mediterranean populations. Of the 25 compound heterozygotes for the beta -101 C --> T and common severe beta-thalassaemia mutations, all but one had mild thalassaemia intermedia preserving haemoglobin levels around 9.5 g/dl and haemoglobin F levels < 25%. The only transfused patient was characterized to have an additional alpha-globin gene. Strict assessment of haematological and biosynthetic findings in the heterozygotes for the beta -101 C --> T mutation (excluding six cases with an alpha-globin gene defect) demonstrated that less than half of them had completely normal (silent) haematology; the remainder had either high haemoglobin A2 values (in the range of 3.7-5.1%) and/or low red cells indices and/or raised haemoglobin F values. The alpha/non-alpha-globin chain synthesis ratios were generally raised, with mean 1.44 (1.07-2.10). Amongst the parents of the compound heterozygotes, who were not selected for molecular analysis following haematological screening, half of the cases were completely silent. Interaction with severe beta-thalassaemia mutations always resulted in the clinical phenotype of mild non-transfusion-dependent thalassaemia intermedia.

Hb Aghia Sophia [alpha62(E11)Val-->0 (alpha1)], an "in-frame" deletion causing alpha-thalassemia.

In this report we describe a case of Hb H disease due to the interaction of the --(MED 1) deletion with a new alpha(+)-thalassemia determinant. The molecular analysis of the proband's genomic DNA was carried out by polymerase chain reaction amplification and sequencing of both alpha genes of the alpha(+)-thalassemia chromosome and revealed a deletion of codon 62 of the alpha1 gene. This DNA triplet codes for a valine residue at the E11 alpha helix, which is located in the interior of the heme pocket. Substitutions of valine E11 with other amino acid residues in the alpha as well as beta polypeptide chains lead, in the heterozygous carrier, either to Hb M disease or to congenital non-spherocytic hemolytic anemia. We assume that the deletion of valine at alpha62(E11) disrupts the conformation of the alpha chain to such an extent that the mutated subunit is rapidly removed by proteolysis. The final result is an alpha-thalassemia phenotype rather than an unstable hemoglobin syndrome. This conclusion is supported by the apparent absence of an abnormal alpha chain in the peripheral blood of the patient.

Tissue immunodetection of c100 hepatitis C virus antigen in major thalassemic patients.

BACKGROUND/AIMS: Hepatitis C Virus (HCV) detection in the livers of chronically infected patients remains a debatable issue. To determine the significance of hepatic expression of hepatitis C viral antigen c100, an immunohistochemical assay was performed in 113 young thalassemics with chronic HCV infection. METHODOLOGY: One hundred and thirteen patients were seropositive for antibody to HCV by second-generation testing. The monoclonal antibody TORDJI-22 was used in an alkaline phosphatase 3-step staining method, and any possible association between the results of HCV immunodetection and various clinicopathologic variables was investigated by univariate and multivariate statistical analysis. In 36 cases, post-therapy liver biopsy specimens were also studied. RESULTS: HCV c100 antigen was detected in 62% of all pretherapy samples, exclusively in the cytoplasm of rather few hepatocytes. Its expression was positively associated with male gender (p = 0.02) as well as with rather advanced age (p = 0.03) and was frequently accompanied by low necroinflammatory scores (according to the modified HAI grading). At the end of interferon-alpha (IFN-alpha) therapy, the immunoreactive prevalence of c100 antigen decreased significantly (pF = 0.002). CONCLUSIONS: We conclude that hepatic expression of c100 antigen is detected in a considerable percentage of thalassemics but it is not likely to provide information concerning the viral load in the infected liver. IFN therapy appears to reduce the hepatic expression of this viral antigen in thalassemic patients.

Quantification of liver iron overload by T2 quantitative magnetic resonance imaging in thalassemia: impact of chronic hepatitis C on measurements.

PURPOSE: Measurement of liver T2 values seems to be an accurate and sensitive magnetic resonance imaging (MRI) method for the quantification of liver hemosiderosis in multiple transfused patients with thalassemia. Because many of these patients have coexistent chronic hepatitis C virus (HCV) infection, the effect of inflammatory changes on liver T2 values was assessed. MATERIALS AND METHODS: Liver MRI studies of 35 HCV+ and 17 HCV- patients with beta-thalassemia, 9 HCV+ patients without thalassemia, and 10 healthy controls of the same age range (13 to 32 years) were reviewed. Iron status was assessed by serum ferritin in all patients, and determination of liver iron concentration (LIC) was available in 16 HCV+ patients with thalassemia. Histologic activity index (HAI) and grades of siderosis were evaluated in all HCV+ patients with thalassemia. RESULTS: Patients with thalassemia had significantly lower T2 values (P < 0.0001) than subjects without thalassemia, whereas no difference existed between HCV+ patients without thalassemia and healthy controls. In HCV+ patients, LIC correlated more nearly with T2 values (r = 0.93) than with serum ferritin (r = 0.73). T2 values were not influenced by HAI score or fibrosis. CONCLUSION: Liver T2 values were found to be more accurate than serum ferritin in predicting liver iron overload and were not influenced by the presence of chronic hepatitis C. Therefore, MRI could serve as a noninvasive alternative to liver biopsy for the quantification of hemosiderosis in HCV+ patients with thalassemia.

Prevalence of yersinia plasmid-encoded outer protein (Yop) class-specific antibodies in multitransfused Greek patients with thalassemic syndromes.

OBJECTIVE: To evaluate the prevalence of class-specific antibodies (G, A, M) to Yersinia enterocolitica plasmid-encoded outer proteins (Yops), in a closely followed multitransfused population of patients with thalassemia. METHODS: Sera from 408 beta-thalassemic patients and 386 healthy blood donors used as controls were analyzed with the enzyme-linked immunosorbent assay (ELISA) for IgG, IgA and IgM antibodies to yersinia outer proteins. The Yop antigen for the ELISA was prepared using a plasmid-bearing wild-type strain of Y. enterocolitica of serotype O:8. RESULTS: Anti-Yop IgG antibodies were detected in 84 out of 408 beta-thalassemic patients (20.6%) compared with only eight out of 386 (2.1%) healthy blood donors. None of the sera of either group was positive for anti-Yop IgA or IgM antibodies. On evaluating patients with registered clinical and laboratory signs of a previous yersinia infection in the period from 1978 to 1996, we found that those with a positive agglutination test for Y. enterocolitica infection at the time of manifestation showed a higher rate of persisting IgG seropositivity to Yops than those with positive culture and clinical signs only. A significant percentage (9.49%) of the seropositive patients had no registered data of a past Y. enterocolitica infection. There was remarkable persistence of anti-Yop IgG antibodies in the thalassemic population, even in patients infected during the early years of our study period (1978--80). CONCLUSIONS: The results suggest that the determination of class-specific antibodies to Yops, which are specific antigens for the pathogenic yersiniae (Y. enterocolitica, Y. pseudotuberculosis and Y. pestis), in addition to its usefulness in the diagnosis of infection, will be a very sensitive and specific index for epidemiologic studies.

Erythroid marrow activity and hemoglobin H levels in hemoglobin H disease.

PURPOSE: To determine serum immunoreactive erythropoietin (Epo) and soluble transferrin receptors (sTfR) levels in patients with hemoglobin H (HbH) disease and the correlation with HbH levels and alpha-globin genotype. PATIENTS AND METHODS: Twenty patients with HbH disease were studied. Methods applied included cation-exchange high pressure liquid chromatography for HbH levels, chemoluminescence for Epo concentration, immunoassay for sTfR concentration, and DNA analysis for alpha-globin genotype characterization. RESULTS: Serum Epo and sTfR levels were significantly elevated (46.6+/-26.8 IU/l and 5.6+/-1.8 mg/l, respectively) in patients with HbH disease compared to controls (9.2+/-3.3 IU/l and 1.8+/-0.7 mg/l, respectively). Epo and sTfR levels correlated positively with HbH concentration (r = 0.93 and 0.80, respectively). The highest Epo and sTfR values were observed in three patients with the highest HbH levels who all had nondeletion alpha-thalassemia mutations. CONCLUSION: Epo and sTfR levels are increased in patients with HbH disease; this increase is directly related to the HbH concentration that usually reflects the degree of globin polypeptide imbalance. The correlation of Epo, sTfR, and reticulocyte production index in these patients indicates that anemia in HbH disease mainly is caused by ineffective erythropoiesis and a mild degree of peripheral hemolysis.