Prenatal diagnosis in congenital erythropoietic porphyria by metabolic measurement and DNA mutation analysis.

Identification of uroporphyrinogen III synthase (UROIIIS) gene mutations in patients with congenital erythropoietic porphyria (CEP) allows fast and reliable carrier detection and prenatal diagnosis. We describe here the first case of prenatal diagnosis by concomitant measurement of uroporphyrin I in amniotic fluid and direct detection of the gene mutation. A French couple, whose first child was diagnosed with CEP, requested prenatal diagnosis at 16 weeks of gestation. Uroporphyrin I was dramatically increased in amniotic fluid and the fetus was homozygous for the C73R mutation, the most common mutation in this disease. The pregnancy was then terminated.

Changes in immunological properties of neutrophil alkaline phosphatase in trisomy 21 pregnancies.

Immunoreactivity, cytochemical, immunocytochemical characteristics and subcellular distribution of neutrophil alkaline phosphatase (NAP) were investigated in blood and/or smear samples from 18 women aged 23-46 years (mean 32.5 years) with trisomy 21 fetuses (17-21 weeks) and 28 women aged 20-42 years (mean 31 years) with normal fetuses (17-22 weeks). Immunochemical NAP investigations were carried out in 8 pathological and 8 normal pregnancies; cytochemical and immunocytochemical procedures were carried out in 18 pregnant women with trisomy 21 fetuses and 28 controls. NAP from women with trisomy 21 fetuses is characterized by: (1) a significant decrease in reactivity with anti-liver-type alkaline phosphatase (AP) and anti-NAP antisera; (2) low or very slight reactivity with antiplacental or anti-intestinal antibodies; (3) marked dispersion of NAP lead citrate reaction products or anti-NAP antibody colloidal gold-labelling in neutrophil cytoplasms, as detected by electron microscopy. This subcellular AP distribution (extramembranous) is different from that of normal NAP sites associated with plasma membrane, nuclear membrane and secretory vesicles. The NAP immunochemical and cytochemical characteristics suggest that neutrophils of a woman with a trisomy 21 fetus contain two AP isoenzymes: the liver/bone type and an atypical AP.

Effects of H-7 and staurosporine on proliferation and self-renewal of acute myeloid leukemia progenitors.

In this study, we compared the impact of two protein kinase (PK) inhibitors, H-7 and staurosporine, on the normal myeloid progenitors (CFU-GM) and acute myeloid leukemia progenitors (AML-CFU) proliferation measured by in vitro clonogenic assay. H-7 and staurosporine displayed a biphasic dose-effect on both CFU-GM and AML-CFU recovery. At the lowest concentration range (0.1 microM to 20 microM for H-7 and 0.1 nM to 1 nM for staurosporine), we observed growth stimulation whereas higher concentrations induced dose-dependent growth inhibition. Moreover, AML-CFU proved to be significantly more sensitive to the inhibitory effect of both H-7 and staurosporine than CFU-GM (3.16- and 2.12-fold, respectively). These results were further confirmed with comparable murine cell line models (FDC-P1, a hematopoietic cell line generated from normal bone marrow and WEHI, a myelomonocytic leukemia cell line). Furthermore, we report that both H-7 and staurosporine present similar inhibitory effects on proliferation (PE1) as on self-renewal (PEs) of AML-CFU. In an attempt to understand more fully the mechanism of action of H-7 and staurosporine, we investigated their impact (when used at their D50) on the human myelogenous leukemia cell line, K562. H-7 and staurosporine induced a transient decrease of cell growth, between 0 and 24 hours, and produced a transient blockade of K562 cells in the S-phase, either 24 or 48 hours after the addition of staurosporine and H-7, respectively.

Prognostic significance of karyotype in a twelve-year follow-up in childhood acute lymphoblastic leukemia.

We report a follow-up of 49 children with acute lymphoblastic leukemia (ALL) diagnosed between 1972 and 1978 (follow-up 12-18 years). This series allowed us to analyze the predictive value of karyotype in a long-term follow-up. Karyotypes were abnormal in 33 cases (67.3%): pseudodiploidy in 11 (22.4%), hyperdiploidy > 50 chromosomes in 8 (16.3%), hyperdiploidy 47-50 chromosomes in 11 (22.4%), and hypodiploidy in 3 cases (6.1%). Event-free survival (EFS) and survival studies showed that the outcome of patients was determined only by treatment and karyotype. Eleven patients have survived, nine in first remission (6 years 5 months to 15 years 2 months), and two are in second remission (3 years 8 months and 8 years 2 months). All ploidy groups are represented in these patients. Late relapses can occur in the hyperdiploid > 50 group, thus accounting for shorter EFS than expected, but because of the unusually long second remission of one patient, the rate of surviving patients was higher for this ploidy group than for all other ploidy groups together. Conversely, patients with only numerical abnormalities (no matter which ploidy group they belonged to), had a better outcome than did patients with structural changes or normal karyotypes and no discrepancy between EFS and survival curves was observed in this chromosomal group. Thus, our results suggest that numerical changes only should be considered an indicator of low risk factor, but our results, based on partially banded karyotypes, need to be verified by a current method and therapy.

t(9;11)(p22;q23) translocation in blastic phase of chronic myeloid leukemia.

A patient with chronic myeloid leukemia showed clonal karyotypic evolution, with the appearance of an i(17q) and t(9;11)(p22;q23). This case sheds light upon leukemogenic events related to t(9;11)(p22;q23). The presence of t(9;22) and t(9;11) in the same clone showed that t(9;11) may affect a pluripotent stem cell, thus accounting for t(9;11) in both lymphoid and monocytic leukemias. In this patient, t(9;11) could not be related to a prior cytotoxic exposure and was instead the result of natural evolution of chronic myeloid leukemia. Furthermore, this led us to assume that the phenotype of blast cells may be determined by a chromosome abnormality. A phenotypic conversion from myeloblastic to undifferentiated morphologic aspect was observed when t(9;11) was detected, suggesting that t(9;11) may have induced a loss in differentiation of blast cells affected by this change. This assumption is in agreement with the putative presence of genes activated in pluripotent progenitors by 11q23 rearrangements.

[Role and contribution of karyotyping in male infertility]. / Place et apports du caryotype en infertilité masculine.

In the context of the aetiological investigation of male infertility, the authors stress the place and the contribution of blood karyotype testing in the light of their personal experience based on 1,612 subjects. This examination has an important place, as about 15% of azoospermic subjects and 6 to 7% of subjects with oligospermia less than 10 million spermatozoa per ml, either alone or in combination with other abnormalities of the semen examination, present a congenital chromosomal abnormality. A remarkable constancy of the results was observed according to identical recruitment criteria. The contribution of this examination is also important: the medical and psychological value of detecting the cause of azoospermia, genetic counselling and antenatal chromosomal diagnosis for non-azoospermic subjects with an equilibrated structural abnormality, in whom treatment allows a chance of procreation, genetic counselling for the family of these subjects in order to prevent the appearance of a chromosomally abnormal infant. In conclusion, the authors argue in favour of the routine use of this test in all infertile subjects with at least isolated oligospermia less than 10 million spermatozoa per ml.

Effect of melphalan against self-renewal capacity of leukemic progenitors in acute myeloblastic leukemia.

This study aimed to evaluate the effect of melphalan on both terminal divisions and self-renewal capacity of acute myeloblastic leukemia (AML) progenitors (colony-forming units, CFU-L) grown in methylcellulose. Terminal divisions and self-renewal were assayed by primary (PE1) and secondary (PE2) colony formation, respectively. Thirteen cases of AML, were tested. Melphalan induced a negative exponential dose-effect on CFU-L survival. Moreover, melphalan was equally effective in inhibiting CFU-L growth in both PE1 and PE2 assays, with D10 values of 1.53 +/- 0.17 micrograms/ml and 1.59 +/- 0.21 micrograms/ml for PE1 and PE2, respectively (p = 0.48). Cytotoxicity of melphalan on CFU-L did not differ significantly from that observed for normal hemopoietic granulocyte-macrophage colony-forming units, erythroid burst-forming units, and granulocyte-erythroid-macrophage-megakaryocyte progenitors. Mafosfamide-lysine, a stable cyclophosphamide congener, strongly inhibited primary colony formation (PE1) with a D10 value of 14.46 +/- 1.76 micrograms/ml, but was much less efficient in the PE2 assay. Our findings suggest that the self-renewal capacity of AML progenitors can be differentially affected by alkylating agents. Moreover, since it is now considered that chemotherapy should be preferentially directed against the self-renewal of leukemic progenitors, melphalan might offer a greater potential than cyclophosphamide or cyclophosphamide derivatives in the therapy of AML.

Alkaline phosphatase and phosphotyrosine phosphatase activities of cultured amniotic cells with trisomy 18.

In cultured amniotic cells from fetuses with Edward's syndrome (trisomy 18), the activities of two protein phosphatases, alkaline phosphatase and phosphotyrosine phosphatase, were measured. Comparison with normal fetal cells showed a different behavior for each enzyme. Alkaline phosphatase was significantly lowered while phosphotyrosine phosphatase remained at normal levels. The interest of these enzyme assays in the screening procedure of this severe chromosome defect is discussed.

Difference in activity properties and subcellular distribution of neutrophil alkaline phosphatase between normal individuals and patients with trisomy 21.

Biochemical, cytochemical characteristics and electron microscopy subcellular distribution of neutrophil alkaline phosphatase (NAP) were analysed in blood and/or smear samples from 39 trisomy 21 patients (Down's syndrome) aged 11.5-18 years (mean 15.5 years) and 55 normal subjects aged 12-20.5 years (mean 17 years). All patients were karyotyped. NAP cytochemical procedures were carried out on all subjects; biochemical NAP determinations were made in 10 patients and 20 controls; ultrastructural electron microscopy of AP was performed in three patients and four normal subjects. Neutrophil alkaline phosphatase from patients with trisomy 21 displayed the following changes: (1) a significant increase of enzyme activity, (2) a high thermal lability of enzyme. Electron microscope morphology exhibited large deposits of NAP reaction product associated with the plasma membrane and intracellular main organelles, like phosphasomes. The NAP biochemical and cytochemical characteristics suggest that trisomy 21 neutrophils contain a non-specific AP isoenzyme, closely related to the early placental form.

Translocation t(3;21)(q26;q22) in acute myeloblastic leukemia secondary to polycythemia vera.

A case of acute myeloblastic leukemia secondary to polycythemia vera suggests that the t(3;21) translocation is not restricted to blastic phases of chronic myelocytic leukemia (CML) but can be associated with blastic phases occurring after other myeloproliferative syndromes. All published cases were in myeloid crises. Furthermore, this translocation may have been induced by mutagenic effects of either 32P or various chemotherapies administered in this case. In the nine cases reported (including ours), hydroxyurea and busulfan were most frequently used (each drug was used separately in six cases and in association in three cases). Even if the t(3;21) translocation is partly therapy induced, this chromosomal abnormality appears to characterize myeloid crises of myeloproliferative syndromes (often CML, seldom polycythemia vera).

[Detection of triple X syndrome during a familial inquiry for hemophilia A]. / Découverte d'un syndrome de triplo X lors d'une enquête familiale pour hémophilie A.

In order to give a genetic counsel to the mother and the two twin-sisters of an Hemophilia A boy, a familial investigation has been carried out. The study with the restriction fragment length polymorphism probes together with the hemostasis screening enabled to specify the status of each member of the family with respect to hemophilia A gene. This investigation evidenced the presence of two maternal alleles and one paternal allele in one of the twin-sisters. This led us to assume that she had a chromosomal abnormality in spite of her normal phenotype. The 47, XX, +(X) result of her karyotype confirmed this assumption. Therefore an accurate genetic counsel was provided to the members of this family and more particularly to the triple X subject.

Heat resistance, immunological and quantitative changes of neutrophil alkaline phosphatase in trisomy 21 pregnancies.

Neutrophil alkaline phosphatase (NAP) was analysed in 25 pregnant women with trisomy 21 foetuses whose chromosomal aberration was recognized by cytogenetic study after amniocentesis. Enzyme investigation was performed at 20-22 weeks of gestation using cytochemical and biochemical techniques. Twenty-nine women at the same stage of normal pregnancies were selected as controls. In parallel, each mother was karyotyped. Ten subjects from each series underwent biochemical and immunological investigation: measurement of enzyme levels, thermostability study and immunological tests with alkaline phosphatase isoenzyme antibodies. NAP from pregnant women with trisomy 21 foetuses was characterized by: (1) a lower rate of enzyme activity, (2) a large amount of heat-stable enzyme (T = 56 degrees C for biochemical assays, T = 85 degrees C for cytochemical tests), and (3) a marked loss of liver antigenicity. These findings suggest the presence in trisomy 21 pregnancies of a non-specific alkaline phosphatase isoenzyme which appears as an "enzyme marker" in maternal circulating neutrophils.

Association of the Philadelphia chromosome and 5q- in secondary blood disorder.

A patient developed a secondary blood disorder 7 years after radiotherapy for a gastric lymphoma. The initial myelodysplastic syndrome evolved to a myeloproliferative phase with transient polycythemia, progressive thrombocythemia, and hyperleukocytosis. Chromosome analysis performed in the terminal phase showed del(5)(q13q31),t(9;22)(q34;q11), and a complex rearrangement involving chromosomes #2 and #3. A correlation between chromosomal abnormalities and hematologic findings could be established. In this case, we have assumed that the Philadelphia translocation is a late event, due to prior mutagen exposure, and its association with a common secondary abnormality (5q-), followed by a progressively developing myeloproliferative phase. Furthermore, the association of Ph and 5q- in a single clone seems to indicate that the same stem cell is affected by these two abnormalities.

An enzymatic marker in mothers of trisomy 21 children: neutrophil alkaline phosphatase.

Neutrophil alkaline phosphatase (NAP) from 12 mothers of normal children was investigated and the results compared to those of 7 mothers with trisomy 21 offsprings, in an attempt to determine a parental molecular change in this chromosomal abnormality. The biochemical properties of the enzyme were analyzed by the procedures of isoenzyme characterization, i.e. enzyme assays, thermostability, inhibition patterns and slab gel electrophoresis. Immunological properties were determined on 5 samples from normal mothers and on the same sample number of mothers with affected children. In these latter NAP showed characteristics that were to some extent different from the ones of normal controls. The following changes were observed: highly significant loading of membrane and nucleus pellets in NAP activity, poor effect of inhibitors on thermostable component and immunodepletion measured by a significant decrease of the normal affinity for antiliver and antiplacental alkaline phosphatase antisera. These findings are discussed in the light of our knowledge of alkaline phosphatase isoenzymes.

[Chromosome anomalies and male infertility. A study of 1,444 subjects]. / Anomalies chromosomiques et infertilité masculine. Etude de 1,444 sujets.

Chromosomal anomaly was detected in 7.6% of a cytogenetic survey of 1,444 infertile male azoospermia or oligozoospermia with a sperm count below 20 million/ml. Anomalies are especially of the sex-chromosome and of autosomal robertsonian translocation type. Distribution of these subjects on account of the sperm count has pointed out an increase of chromosomal anomalies in inverse ratio to the sperm count.