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Biomedical subjects

S H Boyer

Publications and source records attributed to S H Boyer.

At least 37 records · Page 2Linked to original sources
PubMed1984

Production of F cells in sickle cell anemia: regulation by a genetic locus or loci separate from the beta-globin gene cluster.

Levels of fetal hemoglobin (HbF) bearing reticulocytes (F reticulocytes) range from 2% to 50% in patients with sickle cell (SS) anemia. To learn whether any portion of such variation in F cell production is regulated by loci genetically separable from the beta-globin gene cluster, percentages of F reticulocytes were compared in 59 sib pairs composed solely of SS members, including 40 pairs from Jamaica and 19 from the United States. We reasoned that differences in F reticulocyte levels might arise (1) from any of several kinds of artifact, (2) via half-sib status, or (3) because one or more genes regulating F cell production segregate separately from beta S. We minimized the role of artifact by assay of fresh samples from 84 SS individuals, including both members of 38 sib pairs. In 78 of the 84 subjects, serial values for percent F reticulocytes fell within 99.9% confidence limits or were alike by t test (P greater than or equal to .05). This left 32 sib pairs for which F reticulocyte levels in each member were reproducible. When sib-sib comparisons were limited to these 32 pairs, percentages of F reticulocytes were grossly dissimilar within 12 Jamaican and 3 American sibships. Within them, the probability that sibs were alike was always less than or equal to .005 and usually less than or equal to 10(-4). We next minimized the contribution of half-sibs among Jamaicans by a combination of paternity testing and sib-sib comparison of beta-globin region DNA restriction fragment length polymorphisms, especially among discordant pairs. We thereafter concluded that at least seven to eight Jamaican pairs were composed of reproducibly discordant full sibs. There is thus little doubt that there are genes regulating between-patient differences in F cell production that are separate from the beta-globin gene cluster. Still unanswered is (1) whether or not these genes are actually linked to beta S, (2) why F reticulocyte levels in Americans tend to be lower than in Jamaicans, and (3) whether or not differences in F cell production among SS patients are regulated by several major loci or by only one.

Adolescent↗
PubMed1983

Adjuvants to immunological methods for mRNA purification. Application to the isolation of mRNA for carbonic anhydrase I from rabbit reticulocytes.

In order to generate the molecular probes needed to investigate the seemingly coordinate expression of carbonic anhydrase (CA I) and beta-globin within erythrocytes during human development, CA I-containing polyribosomes have been isolated from rabbit reticulocytes by reaction with purified antibodies to CA I followed by immunoadsorption of immune complexes with formalin-fixed protein A-bearing bacteria. In the course of such isolation, a series of maneuvers were seen to have a markedly favorable influence on the level of purity attained. These maneuvers include the use of 5 mg/ml of heparin concentrations to attenuate nonspecific binding and entrapment of unwanted polyribosomes, the addition of 200 units/ml of placental ribonuclease inhibitor to augment recovery in reactions which by test already appeared RNase-free, and the preadsorption of polyribosomes with formalin-fixed bacteria prior to immunological reaction so as to remove a subset of polyribosomes seemingly predisposed to nonspecific binding. In the absence of all of the maneuvers, attained purity was no greater than a few per cent. When all were employed, CA I-mRNA derived from immunopurified polyribosomes was recovered with more than 80% purity and 20% yield, as evident from both immunoassays and electrophoresis of its cell-free products.

Animals↗
PubMed1983

Developmental changes in human erythrocyte carbonic anhydrase levels: coordinate expression with adult hemoglobin.

In order to bolster the argument that parallel developmental changes in erythrocyte adult hemoglobin (HbA) and carbonic anhydrase (CA) content provide a potentially suitable model for the dissection of coordinate gene expression, the magnitude of fetal vs adult differences in CA I and CA II levels was examined in human red cell subpopulations obtained after varying periods of exposure to CA-dependent, NH4Cl-HCO-3-mediated, acetazolamide-modulated hemolysis. When content of CA I and CA II was immunologically assessed in cohorts surviving successively longer periods of hemolysis, cord blood red cells were divisible into two populations. Fifteen to thirty percent are rapidly disrupted and have CA I and CA II concentrations similar to those in adult blood erythrocytes. The remaining 70 to 85% have CA I concentrations which are 100-fold less and CA II concentrations which are 5- to 20-fold less than those found in adults. Thus, contrary to past reports, the magnitude of the developmental change in CA I concentration closely resembles the magnitude of change in HbA levels.

Aging↗
PubMed1983

5-Azacytidine increases fetal hemoglobin production in a patient with sickle cell disease.

5-Azacytidine, given to one sickle cell patient as 9 doses of 30 mg/m2 each at intervals of 8 hr, rapidly increased the number of red cells containing HbF (F cells), decreased the amount of HbS in F cells, and decreased the MCHC of F cells. Although a significant rise in the peripheral blood hemoglobin from 8 gm/dl to 10-12 gm/dl was obtained, painful vaso-occlusive crises continued to occur. Therapy was associated with no marrow toxicity, although erythroblastosis appeared after each course of therapy. Increased HbF production was associated with demethylation of two restriction enzyme sites 5' to the two gamma-globin genes. Five other restriction sites around the gamma-globin genes were unchanged. 5-Azacytidine is considered potentially mutagenic and carcinogenic in man and, therefore, further experimental treatment should be limited only to severely affected sickle cell patients.

Adult↗
PubMed1981

F-cell production in sickle cell anemia: regulation by genes linked to beta-hemoglobin locus.

Strong correlation of F-reticulocyte levels within sib pairs with sickle cell (SS) anemia suggests that the wide-ranging levels found in the SS population are governed by genes linked to the beta S-site. Correlations between F-cell levels in parents and F-reticulocyte levels in their children indicates that these same genes regulate F-cell production in nonanemic persons. Comparison of outcrossed and inbred SS populations suggests that relative well-being arises from homozygosity for alleles dictating high F-reticulocyte response to anemia.

Anemia, Sickle Cell↗
PubMed1980

Quantitation of hemoglobins within individual red cells: asynchronous biosynthesis of fetal and adult hemoglobin during erythroid maturation in normal subjects.

We outline a method for estimating either HbF or HbA content in single erythrocytes and their precursors. Our method depends on microphotometric assay of darkfield reflectance arising from individual pericellular immunoprecipitates developed with anti-HbF or anti-HbA. When uniform-diameter latex microspheres were used to normalize comparisons between preparations, mean coefficient of variation for HbF reflectance among separate preparations of the same sample was < 3%. Reflectance is a faithful (r = 0.99) linear function of the logarithm of picograms per cell in samples with known HbF or HbA content. The following features emerged from such analyses. First, despite the use of antigenically-specific antihemoglobins from different sources, the least detectable quantity of HbF (3.2 pg) and HbA (14.8 pg) remained invariant. Second, these detection thresholds depends on antihemoglobin affinity constants but are little influenced by antibody concentration. Third, our procedure is equally valid for persons with normal HbF constant (mean +/- SD = 4.4 +/- 0.3 pg per cell, 15 subjects) and for those with much higher levels. Fourth, like the percentage of HbF-bearing cells, HbF content is usually unchanging in serial samples. Fifth, the utility of the method is evidenced in bone marrow analyses of five hematologically normal persons in whom HbF content, unlike HbA content, remained constant throughout maturation from erythroblasts to erythrocytes. In vivo HbF biosynthesis is thus normally completed long before HbA production.

Animals↗
PubMed1979

Organization and heterogeneity of sequences within a repeating unit of human Y chromosome deoxyribonucleic acid.

Fragments of 3.4 kilobases (kb) are released from DNA of human males, but not DNA of human females, by cleavage with restriction endonucleases HaeIII, EcoRI, or EcoRII. Most, if not all, reiterated DNA which is specific for the Y chromosome (it-Y DNA) is present within these male-specific 3.4-kb molecules. Although such 3.4-kb molecules are themselves localized to the Y chromosome, this is not true for all sequences within them. At least two distinguishable types of reiterated sequences are found within each 3.4-kb molecule. One type consists of at least two families which are highly reiterated and are not confined to the Y chromosome. The other type is composed of an estimated minimum of 39 families, each moderately reiterated and localized to the Y chromosome. Y-specific and non-Y-specific sequences are interspersed with one another in the same 3.4-kb molecule. In the average 3.4-kb molecule, three 800 nucleotide lengths of Y-specific sequences alternate with four 250 nucleotide lengths of non-Y-specific sequences. Since the total number of families of Y-specific sequences, calculated on the basis of reiteration frequency and total abundance in a male genome, greatly exceeds the number of Y -specific sequences present in a single 3.4-kb molecule, it necessarily follows that the population of these 3.4-kb molecules is heterogeneous.

Base Sequence↗
PubMed1979

SV40 recombinants carrying rabbit beta-globin gene coding sequences.

We have constructed and propagated two SV40 recombinants in which different portions of the viral late gene region are replaced by a rabbit beta-globin coding sequence derived from a cloned cDNA (Maniatis et al., 1976). One of these recombinants, which retains the promoter, leader and intervening sequences for a viral late mRNA, directs the synthesis of a stable SV40-globin hybrid transcript. Using a sensitive radioimmunoassay, we have shown that this globin RNA is translated in infected monkey cells. A second recombinant, which retains the late region promoter but lacks the RNA splicing region, produces neither a stable globin transcript nor any detectable beta-globin. These experiments indicate that some sequence within a 500 bp SV40 segment, presumably the splicing region, is required for the accumulation of stable mRNA. They also demonstrate how hybrid transducing viruses can be used both to characterize viral regulatory sequences and to produce new gene products in cultured cells.

Animals↗
PubMed1979

Production of erythrocytes that contain fetal hemoglobin in anemia. Transient in vivo changes.

Serial microscopic immunodiffusion assays of F cells, i.e., erythrocytes that contain fetal hemoglobin (HbF), in four individuals recovering from anemia demonstrate initial increases in the percentage of circulating reticulocytes that contain HbF (F reticulocytes) and subsequent increases in the percentage of mature erythrocytes that contain HbF (F erythrocytes). In one individual responding to therapy for iron-deficiency anemia, the average percentage of F reticulocytes increased from 4.8+/-1.1 to 16.0+/-2.8% (mean+/-SD), while the mean level of F erythrocytes increased from 3.5+/-0.7 to 7.2+/-0.6%. Two normal children with transient erythroblastopenia exhibited F reticulocyte percentages of 71.3+/-6.7 and 41.5+/-1.5%, respectively, when erythropoiesis resumed. With recovery these values fell to finally measured values of 33.7+/-4.7 and 12.6+/-1.1%, respectively. In an adolescent with sickle cell anemia, F-reticulocyte percentages fluctuated between 0.6+/-1.1 and 34.0+/-2.8% and paralleled the rise and fall of total reticulocytes associated with therapy for a nasopharyngeal carcinoma. Such findings suggest that first, the production of F cells and non-F cells are separately regulated. Second, F-cell production is preferentially stimulated during escape from erythropoietic suppression and selectively depressed at the start of suppression. Third, during escape from erythropoietic suppression, F-cell production in vivo resembles that reported for in vitro cultures of erythroid stem cells. Fourth, individuals with sickle cell anemia, like individuals without hemoglobinopathies, can change their relative level of F-cell production.

Adolescent↗
PubMed1979

Robert A. Robinson.

Explore the source record for details and available documents.

History, 20th Century↗
PubMed1978

Individual variation in the production and survival of F cells in sickle-cell disease.

The protective role and underlying sources of the elevated levels of fetal hemoglobin associated with sickle-cell anemia were reassessed by microscopical immunodiffusion assays. Three variables that contribute to levels of fetal hemoglobin were examined: the percentage of fetal-hemoglobin-containing reticulocytes produced; the quantity of fetal hemoglobin synthesized within such cells; and the extent to which the fraction of fetal-hemoglobin-bearing erythrocytes is enriched beyond the level produced. Four general findings emerged from analysis of 29 patients: each variable is separately regulated; the expression of each is often distinctly different between individual patients; contrary to prior speculation, production of fetal hemoglobin may be as great in the absence of heterocellular hereditary persistence of the hemoglobin as in its presence; and fetal hemoglobin does not, as often supposed, guarantee preferential cell survival. We conclude that the differences encountered among patients must reflect heterogeneity among factors that modify production and survival of cells bearing fetal hemoglobin.

Adult↗
PubMed1978

The proportion of all point mutations which are unacceptable: an estimate based on hemoglobin amino acid and nucleotide sequences.

Statistical analysis of the distribution of 156 kinds of human hemoglobin beta (Hbbeta) chain variants suggests that mutations are essentially random in their location. Thus differential fitness, not differential mutability, is the principal source of nonrandom distribution of interspecies differences in Hbbeta amino acid sequence. Similar analyses of both the location and the kind of interspecies differences detected among primates support this viewpoint and lead us to estimate that at least 95% of all amino acid subsitutions,i.e., nonsynonymous mutations, in Hbbeta are functionally unacceptable in homozygous state. Through the combined use of this estimate and the number of nonsynonymous and synonymous substitutions per nucleotide site inferred from comparisons of entire human and rabbit HbbetamRNA nucleotide sequences, we calculate (a) approximately 70% of synonymous Hbbeta mutations are adaptively undersirable and (b) the mutation rate underlying all changes is lesser than or equal to 10(-8) nucleotide substitutions per nucleotide site per year. Apart from such calculations, analyses of nucleotide patterns in HbbetamRNA as well as in rat preproinsulin mRNA reinforce the notion that a large portion of synonymous mutations are functionally unacceptable and rendered so by selective constraint, at a pretranslational level, of the abundance of particular nucleotide doublets such as CpG.

Amino Acid Sequence↗
PubMed1978

Microscopic method for assaying F cell production: illustrative changes during infancy and in aplastic anemia.

Fetal hemoglobin (HbF)-bearing reticulocytes (F reticulocytes) can be detected in peripheral blood by a modification of the microscopic single-cell radial immunodiffusion method. Thereby otherwise inappreciable changes in HbF production can readily be recognized. F reticulocyte frequencies are reporducibly measurable whenever the product of whole blood HbF-bearing red cell (F cell) frequency and reticulocyte frequency is approximately 5 X 10(-4) or greater. Serial analyses of F reticulocytes and nonreticulocyte F cells (F erythrocytes) illustrate that (1) levels of F reticulocytes and F erythrocytes are persistently similar in normal adults with more than 6% F cells and thus cell survival times of F and non-F cells must be essentially the same, (2) changing levels of F reticulocytes can be sensitive predictors of later changes in mature F cell frequencies during infancy and in adults recovering from aplastic anemia, and (3) alterations in F reticulocyte frequency and the amount of HbF per F reticulocyte are discordant in some settings but concordant in others.

Adult↗
PubMed1977

Analysis of human Y-chromosome-specific reiterated DNA in chromosome variants.

A number of individuals with aberrant Y chromosomes have been tested for the presence of Y-chromosome-specific reiterated DNA. These studies locate Y-chromosome-specific reiterated sequences on the long arm of the Y chromosome. Correlation with phenotype and other known Y chromosome markers establish that the Y-chromosome-specific reiterated DNA discussed here has no evident role in male determination.

Base Sequence↗
PubMed1977

Changing erythrocyte populations in juvenile chronic myelocytic leukemia: evidence for disordered regulation.

Fetal and adult erythrocyte characteristics were studied serially in a 30-mo-old female with juvenile chronic myelocytic leukemia. On presentation the erythrocytes exhibited predominantly fetal characteristics as indicated by 69% hemoglobin F (HbF), 1.1% hemoglobin A2 (HbA2), absent I antigen, and fetal levels of the erythrocyte enzymes, carbonic anhydrase I and II, glucose-6-phosphate dehydrogenase, hexokinase, pyruvate kinase, and lactate dehydrogenase; 100% of the erythrocytes present contained HbF. However, Orskov-Jacobs-Stewart hemolysis demonstrated that at least one adult characteristic was present. Seven months later HbF was 17%; I antigen and carbonic anhydrase I had increased to adult levels. The number of cells containing HbF had decreased to 30%. Further studies indicated that at least three new populations of red cells were present after 7 mo which had not previously been detected. Two of these populations exhibited a mixture of both fetal and adult characteristics. Such findings suggested that an ongoing disturbance of regulatory mechanisms was responsible for the variable expression of fetal versus adult erythrocyte characteristics.

Child, Preschool↗
PubMed1977

Inheritance of F cell frequency in heterocellular hereditary persistence of fetal hemoglobin: an example of allelic exclusion.

Two kindreds are described in which F cell frequency is inherited. These families differ in ethnic origin, the mean quantity of HbF per F cell, and in G gamma: A gamma ratios. Heterozygotes have approximately 50% F cells while homozygotes have close to 100%. Semiquantitative single cell immunodiffusion assays establish that F cells contain all of the HbF found in heterozygotes. Our finding that the gene for this heterocellular form of hereditary persistence of fetal hemoglobin is expressed in only half the cells provides the first example of allelic exclusion known apart from immunoglobulin expression.

Black People↗