Red Cells Of Individuals Research Articles

Immunology

Immunology

Familial distal renal tubular acidosis is associated with mutations in the red cell anion exchanger (Band 3, AE1) gene.

All affected patients in four families with autosomal dominant familial renal tubular acidosis (dRTA) were heterozygous for mutations in their red cell HCO3-/Cl- exchanger, band 3 (AE1, SLC4A1) genes, and these mutations were not found in any of the nine normal family members studied. The mutation Arg589--> His was present in two families, while Arg589--> Cys and Ser613--> Phe changes were found in the other families. Linkage studies confirmed the co-segregation of the disease with a genetic marker close to AE1. The affected individuals with the Arg589 mutations had reduced red cell sulfate transport and altered glycosylation of the red cell band 3 N-glycan chain. The red cells of individuals with the Ser613--> Phe mutation had markedly increased red cell sulfate transport but almost normal red cell iodide transport. The erythroid and kidney isoforms of the mutant band 3 proteins were expressed in Xenopus oocytes and all showed significant chloride transport activity. We conclude that dominantly inherited dRTA is associated with mutations in band 3; but both the disease and its autosomal dominant inheritance are not related simply to the anion transport activity of the mutant proteins.

Analysis of Knops blood group antigens on CR1 (CD35) by the MAIEA test and by immunoblotting.

Kna, McCa, Sla and Yka are red cell antigens of relatively high frequency, located on complement receptor 1 (CR1, CD35). Antibodies to these Knops system antigens are not uncommon. They are not haemolytic and do not reduce the survival of transfused incompatible red cells, but they are a nuisance in transfusion laboratories as they can cause an incompatible crossmatch and must be identified before they can be dismissed as clinically insignificant. Human red cell alloantibodies can be shown to be Knops system antibodies by the monoclonal-antibody-specific immobilization of erythrocyte antigens (MAIEA) test, using murine monoclonal anti-CR1. In addition to confirming that Kna, McCa, Sla and Yka are located on CR1, the MAIEA test was used to confirm that Csa is not on CR1. Red cells of the Helgeson phenotype, the null phenotype of the Knops system by conventional serological methods, have levels of Kna, McCa, Sla and Yka intermediate between those of alpha-chymotrypsin-treated cells (which lack Knops system antigens) and those of positive control cells. Level of expression of Knops system antigens is very variable and intensity of staining of immunoblots probed with monoclonal anti-CR1 correlated with strength of Knops system antigens, as determined by the MAIEA test. In individuals heterozygous for alleles producing different allotypes, separate bands representing each allotype on an immunoblot showed identical intensity of staining, suggesting that the quantity of CR1 on red cells is controlled, at least in part, by a locus independent of CR1. An analysis of CR1 on red cells of individuals who have made Knops system antibodies suggested that the Knops system antigens and the antibodies that detect them are complex and heterogeneous.

Protein-sequence studies on Rh-related polypeptides suggest the presence of at least two groups of proteins which associate in the human red-cell membrane.

The Rh D blood-group antigen forms part of a complex, involving several other polypeptides, that is deficient in the red cells of individuals who lack all the antigens of the Rh blood-group system (Rhnull red cells). These include components recognized by anti-(Rh D) antibodies and the murine monoclonal antibodies R6A and BRIC 125. We have carried out protein-sequence studies on the components immunoprecipitated by these antibodies. Anti-(Rh D) antibodies immunoprecipitate an Mr-30,000-32,000 polypeptide (the D30 polypeptide) and an Mr-45,000-100,000 glycoprotein (D50 polypeptide). Antibody R6A immunoprecipitates two glycoproteins of Mr 31,000-34,000 (R6A32 polypeptide) and Mr 35,000-52,000 (R6A45 polypeptide). The D30 and R6A32 polypeptides were found to have the same N-terminal amino acid sequences, showing that they are closely related proteins. The D50 polypeptide and the R6A45 polypeptide also had indistinguishable N-terminal amino acid sequences that differed from that of the D30 and R6A32 polypeptides. The putative N-terminal membrane-spanning segments of the two groups of proteins showed homology in their amino acid sequence, which may account for the association of each of the pairs of proteins during co-precipitation by the antibodies. Supplementary data related to the protein sequence have been deposited as Supplementary Publication SUP 50417 (6 pages) at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1988) 249, 5.

The A1B genotype expressed as A2B on the red cells of individuals with strong B gene-specific transferases. Results from two paternity cases.

Two paternity cases involving black persons are reported in which possible paternal exclusions existed if the A2B red cell phenotype represented the genotype A2B. ABH transferase assays were performed to determine if the A2B phenotypes arose from the genotype A1B or A2B. These assays confirmed that all group A members carried the A1 gene and not the A2 gene. Therefore, the alleged father was not excluded in either case. The B transferase showed increased activity in both AB individuals, possibly accounting for the observed A2B red cell phenotypes. Therefore, in the presence of a B gene, accurate assignment of A subtypes using standard serological methods may not be possible.

Community-initiated screening programme for beta-thalassaemia trait.

Seven hundred and ninety-five people, (96% of whom were of Greek origin) were screened for thalassaemia trait. A prevalence rate of 7.1% for beta-thalassaemia carriers was found. The red cells of individuals with the alpha-1-thalassaemia, beta-thalassaemia, and haemoglobin Lepore traits all had a mean corpuscular volume (MCV) of less than 76 fL and a mean corpuscular haemoglobin (MCH) of less than 25 pg, thus confirming the usefulness of these indices as a preliminary thalassaemia screening test. However, in three of the six people provisionally diagnosed as having delta-beta-thalassaemia trait, an overlap of MCV and MCH values with the normal range occurred. Community attitudes to the survey and its implications are discussed.

Occurrence of G gamma Hb F in Greek HPFH: analysis of heterozygotes and compound heterozygotes with beta thalassaemia.

Haemoglobin F has been isolated from the red cells of individuals with the Greek form of hereditary persistence of fetal haemoglobin (HPFH), and the glycine/alanine composition of the gamma CB3 peptides determined. In contrast to previous reports we have shown that the Hb F of the Greek HPFH heterozygotes contains significant amounts of G gamma chains and circumstantial evidence indicates that these are the products of the same chromosome that carries the Greek HPFH determinant. Hence this chromosome must be directing the synthesis of G gamma, A gamma and (probably) beta and delta chains, thus implying that the Greek form of HPFH does not result from a deletion involving the globin chain structural genes. Analysis of the levels and structure of Hb F from the Greek HPFH heterozygotes and from separated cell populations from the Greek HPFH/beta thalassaemia compound heterozygotes indicate that the Greek HPFH determinant, while allowing an overall increase in gamma chain synthesis, is not the sole factor determining the absolute amount of Hb F production on a cellular basis.

Modification of the acid elution technique for quantitation of fetal hemoglobin in individual erythrocytes.

A modification of the acid elution procedure for the demonstration of fetal hemoglobin in red cells using fast green stain is described. This technique not only permits improved visual estimation of the amount of fetal hemoglobin in red cells, but allows for the direct quantitation of fetal hemoglobin content of individual cells, using a scanning and integrating microdensitometer. As an application of this method, the distribution of fetal hemoglobin in populations of red cells of individuals was determined. Distributions were examined in sickle cell anemia patients with different proportions of fetal hemoglobin.

Studies on the quantitative variation of human red cell peptidase A activity.

SUMMARY1. A new method for the assay of peptidase A activity in human red cells and leucocytes is described.2. Wide variation in the level of red cell peptidase A activity between different individuals with the common electrophoretic phenotype Pep A1 has been demonstrated.3. Family studies show highly significant parent‐child and sib‐sib correlations, but no significant parent‐parent correlation. The findings indicate that the differences in level of activity between individuals are to a large extent genetically determined.4. Peptidase A activities in leucocytes show much less variation than in red cells. Only a small correlation, which was not statistically significant was found between red cell and leucocyte activity. In particular the levels of leucocyte peptidase A activity in individuals with very weak red cell activities were found to be very similar to those seen in individuals with strong red cell activity.5. Weak peptidase A activity in red cells was not found to be associated with weak activity of other red cell peptidases.6. No evidence for the differential occurrence of inhibitors or activators of peptidase A in red cells of individuals with different levels of activity was obtained in various experiments designed to test this possibility.7. The electrophoretic patterns seen in individuals heterozygous for certain rare alleles (Pep A5 and Pep A6) which determine electrophoretic variants of peptidase A suggest that two different alleles (provisionally called Pep A1S and Pep A1W) determining different forms of the enzyme with the usual electrophoretic mobility (Pep Al), but with different activities in red cells occur. Individuals presumed to be heterozygous for Pep A5 (or Pep A6) and for Pep A1S show a symmetrical triple‐banded electrophoretic pattern, which suggests that the activity attributable to Pep A5 or Pep A6 is about the same as that attributable to Pep A.1S. Individuals presumed to be heterozygous for the rare allele Pep A5 or Pep A6 and for Pep A1W show in red cells an asymmetrical electrophoretic pattern with a marked deficiency of activity attributable to Pep A1W. In leucocytes, however, they show a symmetrical electrophoretic pattern indistinguishable from the patterns observed in individuals with the same rare allele in heterozygous combination with the common allele Pep AS.8. It is suggested that the wide variation in activity seen in red cells of individuals with the common electrophoretic phenotype Pep A1, is at least in part attributable to alleles (Pep A1S and Pep A1W) occurring at the same locus as the alleles which determine previously described electrophoretic variants. However, it remains possible that allelic differences at other gene loci and producing their effects in different ways may also contribute to the genetically determined variation in activity.