[Assay of glycosylated hemoglobin A1c in children. Determination of reference values].
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Adult↗
Biomedical subjects
J Rosa
Publications and source records attributed to J Rosa.
Clinical manifestations and erythrocyte adhesion to endothelium in sickle cell syndrome.
Painful vasocclusive episodes are one of the most prominent pathological features of sickle cell disease. In addition to abnormal shape and poor deformability, increased adhesion of red cells to endothelium has been reported. On several occasions, we have studied the adhesion of erythrocytes from 30 patients with mixed sickle cell syndromes to evaluate the influence of clinical conditions. The percentage of erythrocytes adhering was significantly higher when erythrocytes from sickle patients were compared with controls (p less than 0.01). Furthermore, adhesion was significantly higher when the patients were in crises (p less than 0.01), and the highest values of all were observed in patients with inflammatory conditions. To investigate the possibility that a limited population of red cells could be responsible for the increase in red cell adhesion, we have measured the HbS concentration in the different washes and found that the HbS concentration was higher in the last washes compared to the first washes. Sickle red cells capable of protein synthesis (young red cells) were labelled with [3H] leucine. The adhesion to endothelial cells of [3H] leucine-labelled red cells was higher than that of the 51Cr-labelled red cells from the same patient. On the other hand, the most dense sickle red cells separated by density gradient adhered to a greater extent than the light red cells. This apparent discrepancy could be partly explained by the presence of [3H] leucine-labelled red cells in the dense fractions of sickle red cells separated by stractan gradient.
Adolescent↗
Hemoglobin kenitra alpha 2 beta 2 69 (E13) Gly----Arg. A new beta variant of elevated expression associated with alpha-thalassemia, found in a Moroccan woman.
Hemoglobin Kenitra is a new variant of the beta chain alpha 2 beta 2 69 (E13) Gly----Arg which does not produce any clinical symptoms. It is a slow-moving hemoglobin with a distinctive pattern of electrophoretic mobilities. The stability test was negative. Oxygen affinity studies were not performed. It was found in association with alpha-thalassemia and microcytosis, but paradoxically a high expression of the variant (55%) was observed.
Amino Acids↗
A new case of hemoglobin Providence (alpha 2 beta 2 82 (EF6) Lys----Asn or Asp) discovered in a French Caucasian family. Structural and functional studies.
A new case of Hb Providence was discovered in a French caucasian family presenting a mild polycythemia. Structural and functional studies of the two abnormal fractions (Hb Providence Asn and Asp) have been performed. These confirm the abnormal characteristics of Hb Providence described previously. Red cells containing Hb Providence were fractionated with a Percoll Albumin density gradient. The respective amounts of the two components were determined in the youngest and oldest cells. We observed a slight increase of Hb Providence Asp from 34 to 37% during the life span of the erythrocytes which confirms that the deamidation of Hb Providence Asn to Hb Providence Asp is a fast process, already present and close to its maximum, in the reticulocytes. Both abnormal components are stable in the presence of isopropanol.
Adolescent↗
[Biochemical characterization and study of the sensitivity to Plasmodium falciparum infection. Apropos of a case of double CN Baltimore hemoglobinopathy discovered in Senegal].
The discovery of a double CNBalt hemoglobinopathy allowed the authors, after a family background investigation, to study the possible biological effects linked to that dual heterozygoty and to look for a protection against Plasmodium falciparum infections. It hasn't been possible to make evident any protection.
Adolescent↗
[Abnormal hemoglobins identified in Martinique].
Epidemiologic programs in Martinique during the last 10 years and particularly the last 5, have allowed the determination of the Hb S, Hb C, beta thalassemia traits frequencies. A number of rare variants have been detected during the course of these screening programs. Many of these Hb variants have been analysed at the structural level. For some of them a pathologic interaction with Hb S is observed (Hb D Punjab, Hb O Arab...), and the use of electrophoretic mobilities obtained with the reference samples provides the basis of a rapid, highly probable presumptive identification and then, a useful tool, when for example genetic counselling is necessary.
Epidemiologic Methods↗
Hb Marseille [alpha 2 beta 2 N methionyl-2 (NA2) His----Pro]: a new beta chain variant having an extended N-terminus.
A new abnormal hemoglobin was found in a diabetic Maltese woman by citrate agar electrophoresis. This variant was undetectable by isoelectric focusing. No hematological abnormalities were observed. The structural analysis included isolation of the abnormal beta chain, high pressure liquid chromatography of the corresponding tryptic peptides and then microsequencing of the abnormal T1. These procedures revealed a double abnormality: the presence of a methionyl residue extending the NH2 terminus and a histidine to proline substitution in position NA2.
Amino Acid Sequence↗
Evidence for the presence of a hybrid of phosphoglyceromutase/bisphosphoglyceromutase in the red cells: partial characterization of the hybrid.
Purified phosphoglyceromutase was hybridized in vitro with pure biphosphoglyceromutase . The hybrid showed an electrophoretic mobility identical to that of the intermediate band of red cell phosphoglyceromutase activity in the hemolysate patterns. Electrophoretic tests showed that the partially purified hybrid displayed both bisphosphoglyceromutase and phosphoglyceromutase activities and that the sample contained also a small portion of non-hybrid bisphosphoglyceromutase . By constrast to a non-hybrid mixture of the two purified mutases the hybrid exhibited heat instability of bisphosphoglyceromutase activity and neutralization of phosphoglyceromutase activity by anti- bisphosphoglyceromutase antibody.
Bisphosphoglycerate Mutase↗
Cell-free translation of messenger RNA for human bisphosphoglyceromutase.
mRNAs prepared from different human tissues were translated in a cell-free reticulocyte lysate system and, when present, the neosynthesized bisphosphoglyceromutase ( BPGM ) was specifically isolated by immunoprecipitation and analyzed by polyacrylamide gel electrophoresis. Analysis of the translation products showed that bisphosphoglyceromutase was synthesized in vitro with its mature molecular weight and messenger RNA specifying the synthesis of BPGM exhibited a sedimentation coefficient of 12 S in human reticulocytes. This synthesis seems to be highly tissue specific since we could not evidence any synthesis of this enzyme using mRNA obtained from non erythroid tissue. The BPGM synthesis represents 0.1% of the total neosynthesized non heme proteins in human reticulocyte and ten times less (0.01%) in human fetal liver.
Animals↗
Effects of the alpha 20 mutation on the polymerization of Hb S.
The contribution of the alpha 20 residues in intermolecular contacts present in hemoglobin S fibers was investigated with mixtures of Hb Le Lamentin alpha 2(20)His----Gln beta 2A and of hemoglobin S alpha 2A beta 2(6)Glu----Val and with artificial hybrids alpha 2(20)His----Gln beta 2(6)Glu----Val. This study showed an increased solubility and delay time of polymerization of Hb S in solution only when the mutation at the alpha 20 residue is cis to the beta 6 Val contact. No modification of the polymerization process occurs when the mutation is trans to this beta 6 Val contact. This result is in agreement with the crystal model of Wishner and Love, who showed that one of the two alpha 20 residues of the Hb S tetramer was involved in an axial contact between hemoglobin S molecules in the crystals of Hb S ( Wishner , B.C., Ward, K.B., Lattman , E.E. and Love, W.E. (1975) J. Mol. Biol. 98, 179-194). The present observation is a new illustration of the validity of the crystal model for the structure of the fibers based on pairs of double filaments.
Globins↗
Mutant hemoglobin stability depends upon location and nature of single point mutation.
The temperature dependence of the rates of heme release from the beta subunits of methemoglobin A and 5 beta mutant methemoglobins has been determined. The rates were largest for two hemoglobins with mutations distal to heme, previously known to be unstable. The other 3 mutants also released heme faster than A. These hemoglobins, with single point mutations at the alpha 1/beta 2 interface, were previously thought to be stable. The low reported yields of the 5 mutant proteins covaries with the relative rates of heme release from the met species.
Heme↗
Cell-free translation of human uroporphyrinogen decarboxylase mRNAs.
Uroporphyrinogen decarboxylase was synthesized in a reticulocyte lysate cell-free system under the direction of messenger RNAs isolated from human fetal liver and from human reticulocytes. The enzyme was specifically isolated by immuno affinity chromatography. Analysis of the translation products showed that uroporphyrinogen decarboxylase was synthesized in vitro with its mature molecular weight. This enzyme represented 0.04% of the total neosynthesized proteins under the direction of fetal liver mRNA and about ten times less (0.005%) with reticulocyte mRNA.
Anemia, Sickle Cell↗
Isolation and identification of a cDNA clone coding for rat uroporphyrinogen decarboxylase.
We have cloned and identified a DNA sequence complementary to the mRNA of uroporphyrinogen decarboxylase ( UroDCase ) from rat. This mRNA is a minor species (0.1%) of the total mRNA from anemic rat spleen. Poly(A)+ mRNA was enriched for UroDCase mRNA to 20% purity by a very efficient procedure involving two successive steps of preparative gel electrophoresis under various denaturing conditions. cDNA prepared from partially purified UroDCase mRNA (1% purity) was cloned in the Pst I site of pBR322 by using the homopolymeric G-C tailing method. Primary screening of 500 clones from this cDNA library was performed with a cDNA probe complementary to highly purified mRNA for UroDCase (20% purity) and UroDCase cDNA clones were finally identified by hybrid-selected translation. The rat cDNA clones obtained hybridize to human UroDCase mRNA. This will permit the isolation of the corresponding human gene and molecular analysis of porphyria cutanea tarda, the commonest type of porphyria.
Animals↗
Molecular cloning of a cDNA sequence complementary to porphobilinogen deaminase mRNA from rat.
A cDNA clone containing sequences complementary to the mRNA coding for anemic rat spleen porphobilinogen deaminase (EC 4.3.1.8) has been isolated. A cDNA library was prepared from partially purified mRNA (1% purity). This library was then screened by colony hybridization, using a cDNA probe derived from porphobilinogen deaminase mRNA further enriched (10-20% purity) by gel electrophoresis in the presence of methylmercury hydroxide. Colonies hybridizing with the probe were analyzed by hybrid-selected translation using anemic rat spleen mRNA. Four recombinant plasmids containing porphobilinogen deaminase cDNA sequences were identified by specific immunoprecipitation of the translational product from hybrid-selected mRNA. Porphobilinogen deaminase mRNA was shown to contain 1800 bases by blot hybridization analysis. The cloned cDNA sequence consists of 1500 bases. Hybridization analysis of poly(A)+ RNA from uninduced and induced mouse erythroleukemic cells indicated that induction to erythroid differentiation by dimethyl sulfoxide results in a 10-fold increase of porphobilinogen deaminase mRNA. The rat cDNA clones hybridize to the corresponding sequences encoding human porphobilinogen deaminase. This property will be useful for isolation of human gene(s) and further characterization of the molecular lesion(s) responsible for acute intermittent porphyria.
Ammonia-Lyases↗
Prenatal diagnosis of sickle-cell anemia in the first trimester of pregnancy.
To investigate the usefulness of chorionic biopsy for prenatal diagnosis of sickle-cell anemia by restriction-endonuclease analysis of fetal DNA, we studied 30 pregnancies before elective abortion. When the reproducibility of the technique for obtaining adequate DNA samples was established, we successfully applied the test to five pregnancies at risk for sickle-cell anemia. In two cases, sickle-cell disease of the fetus led to a decision to terminate the pregnancy. In three other cases, a normal or AS genotype was demonstrated. One normal infant has been born, and one other pregnancy is continuing normally. In one case in which fetal death was observed three weeks after sampling, placental abnormalities found on histologic examination were compatible with a chromosomal aberration. Our study shows that chorionic biopsy is feasible for the prenatal diagnosis of sickle-cell disease before the 10th gestational week. If subsequent experience demonstrates this technique to be safe enough for mother and fetus, the ability to test in early pregnancy may make prenatal diagnosis acceptable to more couples at risk for serious genetic disorders.
Abortion, Therapeutic↗
An automatic method for determination of glycosylated hemoglobins using low-pressure liquid chromatography.
Several studies have revealed a correlation between blood levels of glucose and hemoglobin A1c (HbA1c), a minor form of hemoglobin (Hb) present at elevated concentrations in patients with diabetes mellitus. To facilitate a clinical study of the level of circulating HbA1c we have developed an automatic chromatographic system. An efficient separation of HbA1c from HbA0 and other rapid hemoglobins (HbA1a, HbA1b) was achieved on Bio-Rex-70 columns using three buffers. This system allows the daily analysis of 40 samples. The mean level of HbA1c in normal subjects was 5.4 +/- 0.4%. The method also detects the presence of elevated levels of HbF and the most frequent forms of abnormal hemoglobin (HbS, HbC).
Adult↗
Sickle cell hemoglobin fiber formation strongly inhibited by the Stanleyville II mutation (alpha 78 Asn leads to Lys).
A double mutant hemoglobin possessing both the Hb S (beta 6 Glu leads to Val) and the Hb Stanleyville II (alpha 78 Asn leads to Lys) mutations has been purified from the blood of a donor heterozygous for both of the mutations. The purification required two chromatography steps, with the second permitting resolution of the double mutant from Hb A2 remaining after the first step. Measurement of the competence for fiber formation by the double mutant hemoglobin was carried out by the centrifugation of gels to obtain Csat. The double mutant was found to have a greatly elevated Csat, 26.4 gm/dl, compared to 15.2 gm/dl for the Hb S control. The concentration of the pellet of the centrifuged gel for the double mutant was in the range 48-50 gm/dl, suggesting that no major rearrangement in the structure of the fibers had been induced by the Stanleyville II mutation. Electron microscopic observations on the fibers of the double mutant confirmed that a normal appearance was maintained.
Chromatography, DEAE-Cellulose↗
Hb Knossos, beta 27 Ala leads to Ser (B 9): a new hemoglobinopathy presenting as a silent beta-thalassemia.
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Amino Acid Sequence↗