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R Ducrocq

Publications and source records attributed to R Ducrocq.

At least 19 recordsLinked to original sources
PubMed2001

A novel delta beta fusion gene expresses hemoglobin A (HbA) not Hb Lepore: Senegalese delta(0)beta(+) thalassemia.

This study identified and characterized a novel delta beta fusion gene in which the delta-globin gene promoter is linked to intact beta-globin coding sequences in a Senegalese family. It results from a 7.4-kb deletion that removes the delta-globin coding sequences, the delta beta intergenic region as well as the beta-globin gene promoter and causes delta(0)beta(+) thalassemia with hemoglobin A expressed at the 11% to 15% range. The phenotype of this naturally occurring delta beta hybrid gene not only clarifies, in an in vivo context, the respective strength of delta- and beta-globin gene promoters, but also emphasizes the importance of beta-globin intragenic sequences in the expression of beta-globin chains. (Blood. 2001;98:1261-1263)

Adolescent↗
PubMed2001

[Neonatal screening for sickle cell anemia: evaluation of a five-year experience in an area of northern Paris].

UNLABELLED: We report a five-year experience of targeted neonatal screening for sickle cell disease in the northern part of the Paris area as well as the follow-up procedure of screened patients. POPULATION: This geographic area in France is characterized by a high frequency of populations at risk for sickle cell disease. RESULTS: Among 115,480 tested newborns, 250 patients were diagnosed (frequency: 1/462). The quality of the screening was attested by the high frequency (5.34%) of newborn carriers for a hemoglobin abnormality (n = 6168). We developed an optimized strategy which avoids the majority of pitfalls (false positive and false negative responses), except for S/HPFH. More than 95% of sickle cell disease was followed, the majority by medical sickle cell disease experts from hospitals. Only two deaths were recorded during this time period. CONCLUSION: We demonstrate the efficiency of targeting the proposed methodological strategy and the follow-up of affected newborns, a major argument demonstrating the importance of newborn screening.

Anemia, Sickle Cell↗
PubMed2001

Strategy linking several analytical methods of neonatal screening for sickle cell disease.

BACKGROUND: The French national programme for the neonatal screening of sickle cell disease (SCD) was set up in 1995. This screening is targeted at newborn infants at risk. Over 5 years, 115,480 newborn infants were tested from 80 maternity departments from the northern part of the Paris area. 250 Patients with SCD were identified--that is, one in 462 newborn infants tested. Carriers for a haemoglobin (Hb) variant are frequent (5.34%). Some uncommon Hb variants were also identified, which gave rise to pitfalls to the testing when associated with HbS: HbKorle-Bu, HbHope, HbBougardirey-Mali, and HbLadésirade (4% of SS-like profiles). OBJECTIVE: An effective screening strategy was developed to avoid these false positive and false negative responses. METHODS: Isoelectric focusing (IEF), the method of primary screening, is rapid and inexpensive. Cation exchange high performance liquid chromatography (CE-HPLC), which is automated, fast, and quantitative was selected as a secondary method. RESULTS: IEF diagnosed normal profiles in 89% of the tested samples from newborn infants. CE-HPLC identified most of the common Hb variants by their retention time and the measure of HbA/HbS ratio, important for the differential diagnosis between an asymptomatic HbS carrier and an HbS/beta+thal compound heterozygote. Furthermore, the high sensitivity of the CE-HPLC detected as little as 0.5% of a Hb variant. This avoided false negatives in samples from premature or transfused newborn infants. All samples with SS-like profiles were confirmed with a second CE-HPLC with another programme. A combination of these three methods confirmed the status of 99.7% of the samples from the tested newborn infants. Some cases required a reverse phase-HPLC method (for gamma-globin or alpha-globin chain variants). Finally, some exceptional samples required confirmation by testing DNA extracted with Güthrie paper for a precise diagnosis. CONCLUSIONS: This effective strategy combining several methods dramatically reduces the risk of errors. Many families are thus spared unnecessary worrying recalls. The only unavoidable cause of false positives remains the HbS/hereditary fetal Hb (HPFH).

Anemia, Sickle Cell↗
PubMed2000

Spectrum of beta thalassemia mutations and their linkage to beta-globin gene haplotypes in the Indo-Mauritians.

The beta thalassemia alleles in 53 thalassemic Indo-Mauritian patients and their families consisting of 23 homozygous beta-thalassemia, 9 HbE/beta-thalassemia, 18 HbS/beta-thalassemia, 1 HbD/beta-thalassemia, 1 deltabeta/beta-thalassemia and 1 HbH/beta-thalassemia from the island of Mauritius were studied. Characterization by polymerase chain reaction-based reverse dot blot hybridization technique revealed that the IVS1-5 (G-->C) mutation accounted for 74% of the beta thalassemic alleles, while six other mutations occurred at much lower frequencies: HbE codon 26 (G-->A); 10.4%, codon 8/9 (+G); 3.5%, codon 30 (AGG-->ACG) also called IVSI (-1).G-->C; 3.5%, codon 15 (G-->A); 3.5%, codon 41/42 (-CTTT); 2.4% and -28 (A-->G); 2.4%. Association of these mutations to specific beta globin gene sequence framework and haplotype allowed to trace their ancestral link. These data are useful in future molecular screening of the population in view of implementing a thalassemia prevention and control program in Mauritius.

Alleles↗
PubMed2000

Acute clinical events in 299 homozygous sickle cell patients living in France. French Study Group on Sickle Cell Disease.

A subset of 299 patients with homozygous sickle cell anaemia, enrolled in the cohort of the French Study Group on sickle cell disease (SCD), was investigated in this study. The majority of patients were children (mean age 10.1 +/- 5.8 yr) of first generation immigrants from Western and Central Africa, the others originated from the French West Indies (20.2%). We report the frequency of the main clinical events (mean follow-up 4.2 +/- 2.2 yr). The prevalence of meningitis-septicaemia and osteomyelitis was, respectively, 11.4% and 12% acute chest syndrome was observed in 134 patients (44.8%). Twenty patients (6.7%) developed stroke with peak prevalence at 10-15 yr of age. One hundred and seventy-two patients (58%) suffered from one or more painful sickle cell crises, while the others (42.5%) never suffered from pain. The overall frequency of acute anaemic episodes was 50.5%, (acute aplastic anaemia 46%; acute splenic sequestration 26%). A group of 27 patients were asymptomatic (follow-up > 3 yr). Epistatic mechanisms influencing SCD were studied. Coinherited alpha-thalassemia strongly reduced the risk of stroke (p <0.001) and increased that of painful crises (p < 0.02). There was a low prevalence of Senegal and Bantu (CAR) betas-chromosomes in patients with meningitis (p <0.04) and osteomyelitis (p < 0.03). Prevalence of Senegal betas-chromosomes was lower in the asymptomatic group of 27 patients (p < 0.02). The patients come from a population of unmixed immigrants in whom the beta-globin gene haplotype strongly reflects the geographic origin and identifies subgroups with a homogenous genetic background. Thus the observed effects might result more from differences in as yet unidentified determinants in the genetic background than from the direct linkage with differences in the beta-globin gene locus.

Acute Disease↗
PubMed2000

Two new Ggamma chain variants: Hb F-clamart [gamma17(A14)Lys-->Asn] and Hb F-Ouled Rabah [gamma19(B1)Asn-->Lys].

Two new fetal hemoglobin variants affecting the Ggamma chain are reported. Hb F-Clamart was found during investigation of a French newborn who presented with a mild microcytemia. The second variant was found during neonatal screening for hemoglobinopathies of 30,000 babies from a population-at-risk living in the Paris region. It was named Hb F-Ouled Rabah because its structural modification and ethnic distribution is similar to that of Hb D-Ouled Rabah [beta19(B1)Asn-->Lys]. Hb F-Ouled Rabah is clinically silent and occurs at a frequency of ca. 0.1% in newborns originating from Maghreb. Structural characterization of both variants was done by protein chemistry methods, including amino acids analysis and mass spectrometry.

Africa, Northern↗
PubMed1999

Purification, amplification and characterization of a population of human erythroid progenitors.

In humans, studies of the erythroid cell lineage are hampered by difficulties in obtaining sufficient numbers of erythroid progenitors. In fact, these progenitors in bone marrow or peripheral blood are scarce and no specific antibodies are available. We describe a new method which allows proliferation in liquid culture of large numbers of pure normal human erythroid progenitors. CD34+ cells were cultured for 7 d in serum-free conditions with the cytokine mixture interleukin (IL)-3/IL-6/stem cell factor (SCF). This resulted in cell expansion and the appearance of a high proportion of CD36+ cells which were purified on day 7. Methylcellulose clones from these cells were composed of 96.6% late BFU-E and 3.4% CFU-GM. These CD36+ cells could be recultured with the same cytokine mixture plus or minus erythropoietin (Epo) for a further 2-7 d. In both conditions further amplification of CD36+ cells was observed, but Epo induced a more dramatic cell expansion. Glycophorin-positive mature cells appeared only in the presence of Epo, and terminal red cell differentiation was observed after 7 d of secondary culture. Cells obtained from adult CD34+ progenitors mostly contained adult haemoglobin, whereas cord blood-derived cells contained equal proportions of adult and fetal haemoglobin. Activation of STAT5 and tyrosine phosphorylation of the Epo receptor and JAK2 were observed after Epo stimulation of these cells. This new method represents a straightforward alternative to the procedures previously described for the purification of normal erythroid progenitors and is useful in the study of erythropoietic regulation.

Antigens, CD34↗
PubMed1999

A novel C-->A transversion within the distal CCAAT motif of the Ggamma-globin gene in the Algerian Ggammabeta+-hereditary persistence of fetal hemoglobin.

Hereditary persistence of fetal hemoglobin (HPFH) is a group of genetically heterogeneous conditions characterized by the continued expression of fetal hemoglobin in adulthood. These constitute natural models for understanding the mechanism(s) of the hemoglobin switch. Many large deletions in the beta-globin gene cluster and point mutations in one of the fetal globin gene promoters have been described before. In this study we describe a novel C-->A transversion (-114) in the distal CCAAT box of the Ggamma-globin gene promoter associated with the Ggammabeta+-HPFH phenotype in an Algerian family. Individuals heterozygous for this mutation exhibit moderate raise in Hb F levels (0.6-3.5%). Much higher Hb F levels (3.8-11.2%) are observed when a beta(o)-thalassemia allele is present in trans to the hereditary persistence of fetal hemoglobin allele. This novel Algerian HPFH mutation further stresses the importance of the distal CCAAT box in the postnatal regulation of gamma-globin gene expression.

Adolescent↗
PubMed1998

New method for quantitative determination of fetal hemoglobin-containing red blood cells by flow cytometry: application to sickle-cell disease.

A new method to quantify red blood cells (RBCs) containing fetal hemoglobin (HbF), or F cells, by flow cytometry was developed. The use of formaldehyde as fixative agent and sodium dodecyl sulfate (SDS) to permeabilize fixed RBCs resulted in a low staining background, a negligible Hb leakage, and minimal cell clumping. HbF was detected by immunofluorescence with a murine monoclonal antibody directed to the gamma chain of human Hb. The accuracy of the F-cell count was evaluated on mixed-field populations of adult and cord RBCs. The results of flow cytometry were highly correlated with HbF dosage in the hemolysate by high-pressure liquid chromatography (HPLC) (R > 0.99). The sensitivity of the method allowed the study of HbF distribution at the single-cell level in normal controls and patients with sickle-cell disease (SCD). All patients exhibited a heterocellular distribution of HbF, with highly variable percentages of F cells and non-F cells. As an additional and valuable biological parameter in SCD, the mean HbF content per F cell could be deduced from the measurement of HbF level by HPLC and F-cell count by flow cytometry. Results were unchanged after storing the cells for several weeks in an optimized resuspending solution. Since it does not need uncommon reagents or devices and allows the simultaneous detection of membrane antigens by two-color flow cytometry, this method is convenient for routine laboratories as well as for research purposes.

Adult↗
PubMed1998

Fetal hemoglobin and F-cell responses to long-term hydroxyurea treatment in young sickle cell patients. The French Study Group on Sickle Cell Disease.

We have studied the cellular and molecular responses to long-term hydroxyurea (HU) treatment in 29 severely affected young patients with sickle cell disease (mean age, 10.9 +/- 4.1 years). Patients received HU at 20 mg/kg/d on 4 consecutive days per week initially, with a monthly escalated dose avoiding marrow-toxicity (mean steady-state dose, 34.2 +/- 4.6 mg/kg/d) for 12 to 36 months (mean duration, 22 months). The studied parameters were hemoglobin F (HbF), F reticulocytes (F retics), F cells, the amount of HbF per F cell (F/F cell), polymer tendency at 40% and 70% oxygen saturation, and hemolysis. Initial HbF (Fi) was dispersed (from 0.85% to 13.9%). HbF increased in all patients but 1. HbF at maximal response (Fmax) reached a sustained level varying from a 1.5-fold to a 16-fold Fi after a variable delay (6 to 24 months). Fmax was not related to HU dosage, but triangle upF (Fmax - Fi) was strongly correlated to triangle upMCV (MCVmax - MCVi). HbF increase resulted from the increase of both F cells and F/F cell. In this rather short series, Fi and Fmax were not significantly associated with age, gender, or beta-globin haplotype. Neither Fmax nor triangle upF was related to bone marrow reserve, as measured by baseline reticulocyte or neutrophil counts. However, Fmax was highly dependent on Fi. When patients are individualized into three groups according to Fmax (group 1, Fmax >20% [12 patients]; group 2, 10% < Fmax < 20% [11 patients]; group 3, Fmax <10% [5 patients]), Fi is significantly different between groups, being the highest in group 1. In addition, the best responders (group 1) were significantly different from patients in the two other groups with higher levels of total hemoglobin, decreased bilirubin, and decreased polymer tendency.

Adolescent↗
PubMed1998

Combined effect of two different polymorphic sequences within the beta globin gene cluster on the level of HbF.

Beta thalassemia and Hb Lepore heterozygotes included in this study exhibit fetal hemoglobin levels varying from trace quantities to 14% (1.74 g/dl) of total hemoglobin in the adult. In this work, we have examined the correlation of DNA sequence polymorphisms with the observed HbF level. The analysis of polymorphic markers within the beta globin cluster in 39 individuals heterozygous for beta thalassemia or Hb Lepore confirms the previous findings for homozygous beta thalassemia: the presence of both an (AT)9 T5 sequence configuration at position -540 of the beta globin gene and a (C --> T) variation at -158 of the Ggamma globin gene is associated with elevated expression of HbF. However, at least one defective beta globin gene is required to reveal this association. The best evidence is from the study of individuals heterozygous for Hb Lepore with various levels of HbF. In these individuals it was possible to explore the effect of a single (AT)x Ty motif (the other being absent from the rearranged Lepore chromosome) on HbF expression. The presence of the (AT)9 T5 configuration increases HbF level from a median of 0.515 g/dl observed in (AT)7 T7 subjects, to 1.39 g/dl. We confirm the existence of linkage disequilibrium between the (C --> T) variation at -158 of Ggamma gene and the (TG)13 configuration at the second intervening sequence (IVS-2) of Agamma gene and identify two new polymorphisms in this region: (TG)7 (CG)5 (TG)8 linked to haplotype V and (TG)8 (CG)5 (TG)10 linked to haplotype II. This study suggests that two distinct regions of the beta cluster, whether in cis or in trans to each other, can interact to enhance HbF expression when a beta thalassemic determinant is present in heterozigosity.

Adolescent↗
PubMed1998

Compound heterozygosity Hb S/Hb Hope (beta 136 Gly-->Asp): a pitfall in the newborn screening for sickle cell disease.

The presence of Hb Hope associated with Hb S may represent a pitfall (false positive) in the neonatal detection of sickle cell disease by two of the most widely used analytical methods in screening programmes-isoelectric focusing (IEF) and high performance liquid chromatography (HPLC). This example illustrates the need to improve analytical strategies to avoid unnecessary anxiety and summoning of families often from a cultural background in which testing of the father is difficult to obtain. It is suggested that using two independent HPLC procedures might improve the specificity of the screening strategies. Additionally, simple procedures for detection of the most common mutations of the beta globin gene of DNA extracted from dried blood specimens could be easily developed for the control of abnormal samples. These procedures could be introduced into the analytical strategy.

Anemia, Sickle Cell↗
PubMed1997

Effect of alpha-thalassemia on sickle-cell anemia linked to the Arab-Indian haplotype in India.

Two population groups from Western India with a high prevalence of the beta(S) gene, one tribal (Valsad) and the other nontribal (Nagpur), were studied. The beta(S) gene frequency in both populations was similar (0.22 vs. 0.23), but not the clinical expression of sickle-cell anemia (SS): the sickle homozygotes in the tribal group appeared to have a mild clinical course, whereas the majority in the nontribal group exhibited a more severe clinical phenotype. Both tribal and nontribal SS patients had a similarly high mean hemoglobin (Hb)F expression (18.5% vs. 15.5%) and a high number of F cells (72.3% vs. 66.6%). DNA analysis of the beta-globin gene cluster region revealed that in these two populations, this portion of DNA was identical with and corresponded to the typical Arab-Indian haplotype. Nevertheless, in heterozygotes, the mean beta(S) expression was lower (27.9%) in the tribal as compared to the nontribal group (35.5%). The major epistatic factor distinguishing the milder presentation in tribals vs. a more severe manifestation in nontribals was the very high frequency (0.97) of the alpha-thalassemia gene in the former as compared to the latter (0.24). We conclude that the phenotypic expression of sickle-cell anemia, linked to the Arab-India haplotype and expressing similar levels of HbF and F cells, is not uniformly mild in India and that alpha-thalassemia is a powerful and additional epistatic factor in the Indian subcontinent.

Anemia, Sickle Cell↗