The D phenotype distribution in G2 resembles that in Europeans while the frequencies in G1 account for the Amerindian and African genetic contribution. The genotyping strategy described here is suitable to study D variants in the overall population and could allow a better use of the few available D- units and a rational administration of anti-D immunoprophylaxis. The results also show that weak D Type 1 alleles do not exclusively segregate with a Ce allele, as assumed until present.

The prevalence of D-/RHD+ samples is higher than that observed in Europeans. More than 50% of the RHD alleles found were represented by RHDψ and RHD-CE-D(s) showing the African contribution to the genetic pool of the admixed population analyzed. Interestingly, three new alleles were found, two of them being hybrid structures between previously described RHD variants recombined with RHCE sequences. The knowledge of the RHD allele repertoire in our population allowed the implementation of reliable typing and transfusion strategies for a better management of patients and pregnant women.

No abstract

Human red blood cells (RBC) have a well-defined lifespan of 120 days affected by many cellular parameters. The aim of the present study was to investigate through a functional assay the effect of some factors in the interaction of erythrocytes with monocytes: heat rigidification, equilibration at different pH and desialyzation. We also studied the interaction between stored RBC and peripheral blood monocytes with this functional erythrophagocytosis assay. Blood samples from 30 volunteer donors were investigated. 1) Senescent (Se) and Young (Y) RBC were obtained by differential centrifugation. 2) Erythrocyte suspensions: Aliquots of each sample were subjected to the following treatments: a) Rigidification by heat (RRBC), b) Equilibration at different pH (5.34, 6.30, 7.33, 9.20) and c) Desialyzation with neuraminidase and trypsin. The functional assay was performed incubating monocytes obtained by glass adherence with these suspensions of RBC. Whole blood samples (n = 20) were stored during different periods of time (0, 7, 14, 21, 28, 35 and 42 days). The erythrophagocytosis assay was performed during six weeks incubating isologous monocytes with RBC from every unit. Negative and positive controls were performed using non sensitized (NSRBC) and sensitized with IgG anti-RhD (SRBC) red cells. The percentage of active monocytes (AM) obtained were: 1) YRBC: 2.8 +/- 0.9 and SeRBC: 17.5 +/- 2.1; 2a) RRBC: 3.0 +/- 0.9; 2b) 10.9 +/- 0.9, 15.5 +/- 0.8, 3.1 +/- 1.0, 4.0 +/- 1.1; 2c) 11.1 +/- 1.4 and 3.9 +/- 1.0; SRBC 32.1 +/- 1.7 and NSRBC: 2.8 +/- 1.5. The % of AM with SeRBC was higher (p < 0.001) than those obtained with NSRBC. The data of AM with RRBC were significantly lower (p < 0.001) than those obtained with SeRBC and SBRC, indicatingthat heat rigidification of RBC does not increase phagocytosis by monocytes. The values of AM obtained from the suspensions of erythrocytes equilibrated at different pH indicate that the acidification of RBC increases the interaction with monocytes. The % AM with neuraminidase treated RBC was higher than those observed with YRBC and NSRBC (p < 0.001). No modifications were observed with trypsin treated RBC. These results suggest that the loss of sialic acid may be involved in the physiological phagocytosis. The values of AM of stored whole blood were: 2.3 +/- 1.3, 2.7 +/- 1.3, 4.4 +/- 1.6, 6.7 +/- 1.2, 9.6 +/- 1.0, 11.7 +/- 0.8 and 13.0 +/- 1.2. The results showed a significant increase in the % of AM as a function of the preservation time from 2,3 +/- 1,3 for the first day to 13,0 +/- 1,2 for the 42nd day (p < 0.001). The data obtained in this ex vivo model show a significant increase (p<0.001) in the phagocytosis of RBC equilibrated at low pH, desialinized (greater than 80%) with neuraminidase and stored for over 28 days. These factors would be involved in erythrocyte removal via phagocytosis during tissular homeostasis.