One third of alloantibodies in patients with sickle cell disease transfused with African blood are missed by the standard red blood cell test panel

Studies on red blood cell (RBC) antibodies in Africa routinely use standard test cells from donors of Caucasian descent. There are no systematic data on alloimmunization against antigens that are almost exclusively present in Africans. We studied the prevalence of antibodies in transfused Ghanaian patients with sickle cell disease (SCD) using standard test cells (representing predominantly antigens more common in Caucasians (Caucasian antigens) and cells expressing antigens more common among Africans (African antigens). Antibodies were present in 16% of 221 patients; 31% of these were directed against African antigens that were not detected with standard test cells. Our findings are not only relevant for an African setting, but also for Western blood banks that are developing strategies to recruit more African donors

Comparison of introns in a cdc2-homologous gene within a number of Plasmodium species

The first three introns of CRK2, a cdc2-homologous gene, have been compared in a total of seven Plasmodium species. The introns were located at conserved sites, suggesting an ancestral origin. Interspecific comparison of intron sequences agreed with the previously inferred evolutionary relationships of the malaria parasites. Unlike the introns in the rodent malaria species, the similarity of the CRK2 introns was regionalized between the human parasite P. vivax and the simian parasite P. knowlesi: the central region of all three introns showed markedly less interspecific similarity than the 5′ and 3′ regions. This was also in contrast with the organisation and composition of homologous intron pairs from other genes of the same two species. No conservation at the level of secondary strucure could be detected, even between highly similar introns. A database search for intron-containing Plasmodium genes was performed. All introns obtained in this way plus the additional CRK2 introns were scanned for the presence of putative branching site consensus sequences. For P. falciparum, we present an update of the 5′ and 3′ splice-site consensus

Hippocampal CARP over-expression solidifies consolidation of contextual fear memories

The Doublecortin-Like Kinase (DCLK) gene is involved in neuronal migration during development. Through alternative splicing the DCLK gene also produces a transcript called Ca2+/calmodulin dependent protein kinase (CaMK)-related peptide (CARP) that is expressed exclusively during adulthood in response to neuronal activity. The function of CARP, however, is poorly understood. To study CARP function, we have generated transgenic mice with over-expression of the CARP transcript in, amongst other brain areas, the hippocampus. We aimed to characterize possible behavioral adaptations of these mice by using a Pavlovian fear conditioning approach. This type of fear conditioning, in which both the hippocampus and amygdala are critically involved, allows studying the formation and extinction of fear related memories. We here report on the behavioral adaptations of two distinct transgenic lines: one with high levels of CARP in the hippocampus and amygdala, whilst the other has high levels of CARP in the hippocampal formation, but not in the amygdala. We tested both mouse lines separately by comparing them to their wild-type littermate controls. We provide evidence suggesting consolidation of contextual fear memories is strengthened in mice of both transgenic lines. © 2010 Elsevier Inc

Comprehensive genotyping for 18 blood group systems using a multiplex ligation-dependent probe amplification assay shows a high degree of accuracy

In recent years genotyping methods have been implemented in blood banks as alternative to comprehensive serologic typing. We evaluated a newly developed assay for convenient and comprehensive genotyping of blood group alleles based on multiplex ligation-dependent probe amplification (MLPA) technology. We analyzed 103 random and 150 selected samples to validate the specificity of the blood-MLPA assay that is able to determine the presence, absence, and copy number of 48 blood group and 112 variant alleles of 18 blood group systems. A total of 4038 serologic typing results, including 52 different antigens, were available for these samples. In 4018 (99.5%) of the 4038 serologic typing results the predicted phenotypes by the blood-MLPA were in concordance with serologic typing. Twenty discordant results were due to false-positive serologic results (n = 2), false-negative serologic results (n = 1), inability of routine serologic typing to detect variant antigens (n = 14), or false-positive prediction from the blood-MLPA due to the presence of a null allele (n = 3). The blood-MLPA reliably predicts the presence or absence of blood group antigens, including almost all clinically relevant blood group antigens, except ABO, in patients and donors. Furthermore, it is the first assay that determines copy numbers of blood group alleles in the same test. It even provides more detailed and accurate information than serologic typing, because most variant alleles are immediately recognized. Since only standard laboratory equipment is needed, this assay finally offers the possibility to comprehensively type recipients and makes extensive matching for selected patients groups more feasibl

Novel variants in Krueppel like factor 1 that cause persistence of fetal hemoglobin in In(Lu) individuals

Beta-hemoglobinopathies become prominent after birth due to a switch from γ-globin to the mutated β-globin. Haploinsufficiency for the erythroid specific indispensable transcription factor Krueppel-like factor 1 (KLF1) is associated with high persistence of fetal hemoglobin (HPFH). The In(Lu) phenotype, characterized by low to undetectable Lutheran blood group expression is caused by mutations within KLF1 gene. Here we screened a blood donor cohort of 55 Lutheran weak or negative donors for KLF1 variants and evaluated their effect on KLF1 target gene expression. To discriminate between weak and negative Lutheran expression, a flow cytometry (FCM) assay was developed to detect Lu antigen expression. The Lu(a−b−) (negative) donor group, showing a significant decreased CD44 (Indian blood group) expression, also showed increased HbF and HbA2 levels, with one individual expressing HbF as high as 5%. KLF1 exons and promoter sequencing revealed variants in 80% of the Lutheran negative donors. Thirteen different variants plus one high frequency SNP (c.304 T > C) were identified of which 6 were novel. In primary erythroblasts, knockdown of endogenous KLF1 resulted in decreased CD44, Lu and increased HbF expression, while KLF1 over-expressing cells were comparable to wild type (WT). In line with the pleiotropic effects of KLF1 during erythropoiesis, distinct KLF1 mutants expressed in erythroblasts display different abilities to rescue CD44 and Lu expression and/or to affect fetal (HbF) or adult (HbA) hemoglobin expression. With this study we identified novel KLF1 variants to be include into blood group typing analysis. In addition, we provide further insights into the regulation of genes by KLF1

Molecular analysis of the York antigen of the Knops blood group system

Antigens of the Knops blood group system are present on complement component (3b/4b) receptor 1 (CR1/CD35), which is a transmembrane glycoprotein encoded by the CR1 gene. Eight of the nine known antigens of this system are linked to polymorphisms in Exon 29. The molecular background of one antigen, York (Yk(a)), has not yet been described. We aimed to identify a polymorphism associated with the absence of Yk(a) to enable molecular typing. Yk(a)-negative individuals were identified by serologic typing. Their CR1 gene was partially sequenced and compared to that of Yk(a)-positive individuals. Loss of Yk(a) antigen was investigated by expressing the SCR22/23 domain of both wild-type and mutated CR1 as a GPI-linked protein on HEK293 cells. We observed that absence of the Yk(a) antigen is caused by a mutation in Exon 26 of the CR1 gene. This 4223C>T mutation results in a 1408T>M change at the protein level. Ten of 117 donors (8.5%) were homozygous TT, confirming the Caucasian frequency of 8% Yk(a)-negative individuals. Serologically, these TT donors showed a Yk(a)-negative phenotype, while CC/CT individuals were Yk(a)-positive. While the Yk(a) antigen was present on HEK293 cells expressing wild-type constructs, cells expressing the 4223C>T variant were Yk(a) negative. We identified a 4223C>T sequence variation in the CR1 gene causing absence of the Yk(a) antigen of the Knops blood group system. With this finding, all polymorphisms of the known Knops blood group antigens have been revealed, enabling molecular testing to contribute to red blood cell alloantibody identification procedure