bluuebunny/crossref_metadata_2025 huggingface dataset:
Expression of Arabidopsis LINEs from two promoters
Authors: Ohta Yoshizu , Noma Kenichi , Tsuchimoto Suguru , Ohtsubo Eiichi , Ohtsubo Hisako
Published: March 2003

Most Arabidopsis long interspersed elements (LINEs, called ATLNs) have two open reading frames, orf1 and orf2. In the 5′ untranslated regions (UTRs) located upstream of orf1, the most proximal segments of tens of base pairs long are not homologous even in two ATLN members with almost identical sequences. In this study, we first show that RT‐PCR products from ATLN39, a member of ATLN, can be detected only in total RNA from the hypomethylation mutant ddm1 or from suspension‐cultured cells treated with a DNA methylation inhibitor 5‐azacytidine, indicating that the expression of ATLN39 is negatively regulated by DNA methylation. We then show that orf1 fused in frame with the luciferase (luc) gene is expressed in suspension‐cultured cells of A. thaliana when the 5′ UTR is present in the region upstream of orf1. Analysis of deletion in the 5′ UTR revealed that the 5′ UTR has two promoters, designated here as P1 and P2. Analysis of transcripts by 5′ RACE showed that their 5′ ends were located at sites immediately upstream of the P1 region or at sites downstream of the P2 region. This observation and the fact that the P1 region contains no TATA sequence indicate that P1 is an internal promoter that initiates transcription from sites upstream of the promoter. A sequence containing GGCGA with a CpG methylatable site is conserved in the P1 regions in members closely related to ATLN39. The P2 region, however, contains the TATA sequence as well as another sequence with a CpG site. The TATA sequence is conserved in members closely related to ATLN39 but not in the other ATLN members, suggesting that P2 is the promoter uniquely present in the ATLN39‐related members. Transcripts from promoter P1 can be used as templates to give new copies proficient in retroposition, but those from promoter P2 cannot because of the lack of the proximal half region of the 5′ UTR sequence. Transcripts from promoter P2, as well as those from promoter P1 can, however, be used for the production of a sufficient amount of proteins for retroposition. Only a short sequence of the non‐homologous region is present at the 5′ ends of transcripts from promoter P1, thus suggesting that the non‐homologous regions seen in the most proximal regions in ATLN elements are not generated in transcription.
Touch modulates gravity sensing to regulate the growth of primary roots of Arabidopsis thaliana
Authors: Massa Gioia D. , Gilroy Simon
Published: March 2003

Plants must sense and respond to diverse stimuli to optimize the architecture of their root system for water and nutrient scavenging and anchorage. We have therefore analyzed how information from two of these stimuli, touch and gravity, are integrated to direct root growth. In Arabidopsis thaliana, touch stimulation provided by a glass barrier placed across the direction of growth caused the root to form a step‐like growth habit with bends forming in the central and later the distal elongation zones. This response led to the main root axis growing parallel to, but not touching the obstacle, whilst the root cap maintained contact with the barrier. Removal of the graviperceptive columella cells of the root cap using laser ablation reduced the bending response of the distal elongation zone. Similarly, although the roots of the gravisensing impaired pgm1–1 mutant grew along the barrier at the same average angle as wild‐type, this angle became more variable with time. These observations imply a constant gravitropic re‐setting of the root tip response to touch stimulation from the barrier. In wild‐type plants, transient touch stimulation of root cap cells, but not other regions of the root, inhibited both subsequent gravitropic growth and amyloplast sedimentation in the columella. Taken together, these results suggest that the cells of the root cap sense touch stimuli and their subsequent signaling acts on the columella cells to modulate their graviresponse. This interaction of touch and gravity signaling would then direct root growth to avoid obstacles in the soil while generally maintaining downward growth.
GFP‐tagging of cell components reveals the dynamics of subcellular re‐organization in response to infection of Arabidopsis by oomycete pathogens
Authors: Takemoto Daigo , Jones David A. , Hardham Adrienne R.
Published: March 2003

Cytoplasmic aggregation, the rapid translocation of cytoplasm and subcellular components to the site of pathogen penetration, is one of the earliest reactions of plant cells against attack by microorganisms. We have investigated cytoplasmic aggregation during Arabidopsis–oomycete interactions. Infection by non‐pathogenic Phytophthora sojae was prevented in the plant epidermal cell layer, whereas Peronospora parasitica isolates Cala2 (avirulent) and Noks1 (virulent) could both penetrate into the mesophyll cell layer. Epidermal cell responses to penetration by these oomycetes were examined cytologically with a range of transgenic Arabidopsis plants expressing Green Fluorescent Protein (GFP)‐tagged cell components. These included plants containing GFP‐TUA6 for visualizing microtubules, GFP‐hTalin for actin microfilaments, GFP‐tm‐KKXX for endoplasmic reticulum (ER), and STtmd‐GFP for the Golgi apparatus. In all interactions, actin microfilaments were actively re‐arranged and formed large bundles in cytoplasmic strands focused on the penetration site. Aggregation of ER membrane and accumulation of Golgi bodies at the infection site were observed, suggesting that production and secretion of plant materials were activated around the penetration site. Microtubules did not become focused on the penetration site. No difference was evident between the responses of epidermal cells in the non‐host, incompatible and compatible interactions. This result indicates that the induction of cytoplasmic aggregation in Arabidopsis epidermal cells was neither suppressed by the virulent strain of Peronospora, nor effective in stopping infection.
Inflammatory cytokines and their receptors in arterial and mixed venous blood before, during and after infusion of drained untreated blood
Authors: Krohn  , Reikerås  , Aasen
Published: March 2003

Wound blood for postoperative autologous transfusion is drained through an area of damaged tissue, the surgical wound, and contains inflammatory mediators. The inflammatory cytokines interleukin‐1‐beta (IL‐1β), interleukin‐6 (IL‐6), interleukin‐8 (IL‐8) and tumour necrosis factor‐alpha (TNF‐α) and their modulators interleukin‐1‐receptor antagonist (IL‐1Ra), interleukin 6 soluble receptor (IL‐6sR), soluble tumour necrosis factor receptor 1 (sTNF‐R1) and interleukin 10 (IL‐10), together with white cell count (WCC) and white cell differential count were measured in arterial and mixed venous blood before, during and after infusion of postoperatively drained untreated blood in nine patients operated for thoracic scoliosis. We found a transient increase in IL‐6, an increase in TNF‐RI, an increase in IL‐8 with granulocytosis and a decrease in IL‐10 in the systemic circulation. The increase in IL‐6 was higher in mixed venous than in arterial blood.
Prenatal treatment of severe fetomaternal alloimmune thrombocytopenia
Authors: Sainio  , Teramo  , Kekomäki
Published: March 2003

Prenatal treatment of fetomaternal alloimmune thrombocytopenia (FMAIT) in previously affected families is of great clinical importance. We report here our experience in the prenatal treatment of 15 severely thrombocytopenic fetuses. Thrombocytopenia was in 13 cases due to immunization to HPA‐1a, in one case to HPA‐5b, and in one case to HPA‐6b. Thirteen fetuses received altogether 34 intrauterine platelet transfusions, seven of them in combination with maternal‐administered intravenous gammaglobulin (IVIG) and two in combination with IVIG and prednisone. Six of the 13 fetuses had only one transfusion just prior to delivery.  In our experience, IVIG seemed to be less effective than reported; only two fetuses of eight treated initially with weekly maternal‐administered IVIG responded, and these were the mildest affected cases in the study. On the other hand, owing to the short survival time, weekly platelet transfusions could only partly maintain a safe platelet count in the four fetuses treated with serial intrauterine platelet transfusions. The number of transfusions needed to be limited because of the high cumulative risk associated with repeated procedures. Three of 34 intrauterine platelet transfusions were associated with near‐loss of three different fetuses due to prolonged fetal bradycardia after the transfusion.  In conclusion, overall neonatal outcome was good, with no mortality; among the study group there was no intracranial haemorrhage (evaluated by postnatal ultrasonography) compared with one case in their untreated siblings. However, the problem of the optimal treatment of FMAIT remains to be solved. For the moment, the treatment of choice is a combination of maternal IVIG and platelet transfusions in severely affected cases. Serial fetal blood samplings (FBS) are needed in order to monitor the fetus with sufficient care.
Differential leucocyte subpopulation analysis of leucodepleted red cell products
Authors: Rider  , Want  , Winter  , Turton  , Pamphilon  , Nobes
Published: March 2003

Donor leucocytes are responsible for many adverse transfusion effects. Clinical reactions may be attributed to specific leucocyte subsets. In this study leucocyte subpopulations were identified and quantified pre‐ and post‐leucodepletion by integral filtration using novel Optipac® configurations incorporating either WBF‐1 (Pall Medsep) or RS2000 (Asahi) whole blood filters. Leucocytes were analysed by flow cytometry using direct, four‐colour, membrane immunofluorescence with monoclonal antibodies specific for CD 3, 14, 16, 19 and 45.  Filtration reduced the leucocyte load by 3–4 log10, consistently giving products with < 2 cells μL−1. Subset distributions were also affected with the proportion of neutrophils and monocytes increased and the lymphocyte/monocyte ratio inverted. These effects were independent of the preprocessing hold conditions, filter used and buffy coat (BC) removal. All filtered red cell products contained 75–80% neutrophils, 16–20% monocytes and 2–7% lymphocytes. Results presented here demonstrate that whole blood filtration, and BC removal, significantly reduce the content and substantially alter the subpopulation distribution of the donor leucocytes remaining in leucodepleted red cell products.
Quality assurance of progenitor cell content of apheresis products: a comparison of clonogenic assays and CD34+ enumeration
Authors: Rock  , Chin‐Yee  , Cantin  , Giulivi  , Gluck  , Keating  , Keeny  , Klassen  , Sutherland  ,   the Canadian Apheresis Group and Canadian Bone Marrow Transplant Group
Published: March 2003

The most common methods used for evaluation of the haematopoietic stem cell content of peripheral blood apheresis products are the colony forming cell assay and the enumeration of CD34+ cells by flow cytometry. The Canadian Apheresis Group and the Canadian Bone Marrow Transplant Group established a multicentre study to compare the reproducibility of colony forming cell assays and CD34+ enumeration by flow cytometry in six transplant centres routinely performing haematopoietic stem cell apheresis.  Over a 5‐month period in 1996, 31 fresh apheresis samples were shipped by overnight courier for testing at six centres to perform CD34+ enumeration by flow cytometry and clonogenic assays. The mean coefficient of variation and range for the following assays were: cell count 36% (2.6–148%), CFU‐GM 82% (46–123%), CD34+ absolute/kg 60% (14–174%) and CD34+ per cent 42% (12–84%).  The wide variation in cell count in this pilot study highlights the difficulties related to provision of samples for quality assessment programmes. Results showed poor interinstitutional reproducibility even among selected samples with similar cell counts for both CFC and CD34+ assays demonstrating the need for development and implementation of an interinstitutional quality assurance programme for haematopoietic stem cell assessment. Provision of a reliable source of testing material will be a necessary next step.
Predicting a donor's likelihood of donating within a preselected time interval
Authors: Flegel  , Besenfelder  , Wagner
Published: March 2003

The procurement of some advanced blood components, like quarantined plasma units, depends critically on retesting the donor within a fixed time frame. For health care systems, such as that in Germany, with mandatory retesting of donors before plasma release, the reliable identification of donors who are more likely to return in time has an immense practical implication, because their blood components could be preferably selected for quarantine purposes. The donation histories of about 760 000 donors with 4910 000 donation attempts were analysed. We developed a logistic regression model to calculate a probability of donation, p(Dts–te), within a preselected time frame (ts–te). The donation history was compounded in a score and shown to be very useful for determining p(Dts–te). A logistic regression model was developed with score and donor status as parameters; different regression coefficients applied to first‐time‐donors (ftd) and to repeat donors (intercept, int, and score factor, scf ). This model allowed us to determine the probability of donation, p(Dts–te), within a preselected time interval, e.g. 6–9 months after an index donation. The p(Dts–te) can be calculated for any donor of blood services. The p(D170–275 days) ranged from about 22% to 86% for any index donation in 1996/97. First‐time donors had a p(D170–275 days) of 33% and were more likely to return within the time interval than certain subsets of repeat donors who can be defined by our model. We provided a technical procedure to increase the rate of plasma unit release after quarantine storage and showed the usefulness of our procedure for blood component management, if quarantine storage is required. By applying the model to our current plasma quarantine programme we could retrieve about 30% more units, which would represent about 30 000 units per year, without incurring additional costs. General implications for blood collection, like planning blood drives, were discussed. The whole demand of a health care system for single plasma units may be met by quarantine plasma and their cost‐efficiency can be improved.
Platelet alloantigen frequencies in Amazon Indians and Brazilian blood donors
Authors: Chiba  , Bordin  , Kuwano  , Figueiredo  , Carvalho  , Vieira‐Filho  , Kerbauy
Published: March 2003

The frequencies of human platelet‐specific alloantigens (HPAs) vary between different ethnic groups, and genotyping using DNA techniques has been preferred over immunophenotyping methods for population studies. Using a polymerase chain reaction with allele‐specific primers (PCR‐ASP) method, we determined the allelic polymorphisms of five HPA systems among 174 unrelated individuals of two different Brazilian ethnic groups including Amazon Indians (n = 95) and blood donors (n = 79). Comparison of the calculated gene frequencies of the two alleles of HPA‐1, ‐2, ‐3, ‐4 and ‐5 systems for Amazon Indians and Brazilian blood donors showed that gene frequencies obtained for the two alleles of HPA‐1 (P < 0.001), HPA‐2 (P = 0.001) and HPA‐5 (P < 0.001) were significantly different between the two groups of individuals. All natives tested carried the HPA‐2a and the HPA‐5a alleles, but the HPA‐1b and HPA‐4b alleles are absent from the Indian population. It was also observed that all blood donors carried the HPA‐1a, HPA‐4a and HPA‐5a alleles. In conclusion, the present data indicate differences in the frequency of the HPA systems between Amazon Indians and Brazilian subjects who present a high rate of racial admixture. While the frequencies of the HPA‐1 and HPA‐5 genes seen in Amazon Indians are similar to those reported for Oriental populations, the frequencies of the HPAs alleles in Brazilian blood donors are comparable to those reported for populations in North America and Europe.
Determination of red cell volume in infants needing blood transfusion
Authors: Fisher  , Matthes  , Wynn  , Al‐Ismail  , Hoy  , Wardrop  , Williams
Published: March 2003

The total circulating red cell volume (RCV) is a better guide to the oxygen‐carrying capacity of the blood in the whole circulation than is the haemoglobin concentration (Hb) or haematocrit in a blood sample. Pre‐ and post‐transfusion RCV (and blood volume (BV)) may be determined by flow cytometry by exploiting antigen differences between transfused donor red cells and the recipient's red cells. This paper describes the use of red cell antigen differences of Duffy, Kidd, MN and RhD between donor and recipient.

In 20 infants, transfused on 21 occasions, pretransfusion RCV ranged from 12 to 39 mL kg−1 body weight. Only at one transfusion could no usable donor–recipient antigen differences be exploited.

Measurement of RCV, used routinely, may determine the transfusion requirements of sick infants more accurately, with the aim of normalizing RCV and BV — securing euvolaemia — at the end of the transfusion. This may allow a complete correction of the RCV deficiency at the first occasion of transfusion. This approach may reduce donor exposures and also optimize oxygen transport and organ perfusion of the infant undergoing intensive management, perhaps leading ultimately to improved survival rates and fewer long‐term complications of neonatal intensive care.
Anti‐HNA‐1a antibodies detected in a FcγRIIIb ‘null’ male blood donor
Authors: Lucas G. F. , Evans R. G. , Griffiths R. E. , Rogers S. E.
Published: March 2003

FcγRIIIb ‘null’ individuals do not express the HNA‐1a, ‐1b & ‐1c alloantigens and the reported incidence of this condition in the Caucasian population is between 1 in 1000 and 2000. The majority of FcγRIIIb ‘null’ individuals have been identified following the investigation of neonatal alloimmune neutropenia due to maternally transferred anti‐FcγRIIIb antibodies.

This report describes a healthy, male blood donor who was screened for the presence of granulocyte‐specific antibodies as part of a foetal concentration donor accreditation programme. The donor's serum was found to contain anti‐HNA‐1a IgM (and weak IgG) antibodies using an immunofluorescence test. Anti‐lymphocyte antibodies were not detected. Further investigation was undertaken to determine whether the antibodies were alloimmune or autoimmune in origin. Granulocyte membrane bound IgG and IgM was within the normal range and the granulocytes were unreactive both with monoclonal FcRIIIb and HNA‐1a antibodies in immunofluorescence tests. HNA‐1a and HNA‐1b PCR products could not be amplified from the donor's genomic DNA using an SSP technique. It was concluded that the donor was FcγRIIIb ‘null’. The donor denied any history of blood transfusion or surgical procedures; nor was there any evidence of such procedures in early life. This suggests either that HNA‐1a antibodies may, in some circumstances, be ‘naturally occurring’ or that the antibodies detected may be part of an autoimmune response to ‘HNA‐1a like’ structures on other tissues. This is the first time that anti‐HNA‐1a antibodies have been reported in an FcγRIIIb ‘null’ individual.

There was no history of transfusion reactions associated with the use of the donor's blood products. However, the donor was withdrawn from the donor panel because HNA‐1a IgM antibodies have been implicated in Transfusion Related Acute Lung Injury.
On investigation of markers that may influence alloantibody responses to HPA‐1a
Authors: Railton D. , Brown J. , Brown C. , Williamson L. M. , Navarrete C.
Published: March 2003

In > 99% of cases, HPA‐1a antibody production during pregnancy is associated with maternal DRB3*0101 positivity. However, only 35% of HPA‐1a neg/DRB3*0101 women produce antibodies (Ab). This study attempted to identify additional genetic marker(s) that may better predict anti‐HPA‐1a production in these women.

Seventy‐eight DRB3*0101 pos HPA‐1a neg, women (40 HPA‐1a Ab pos, 38 Ab neg) with HPA‐1a pos infants, were typed for HLA‐DRB1*, ‐B3*, ‐B4*, ‐B5*. Results were compared with those from 83 DRB3*0101 pos normal donors. SNaPshot™ was used to test for a polymorphism of the TNF‐α locus.

The frequency of DRB1*15 was significantly lower in Ab pos mothers (1/40) compared to controls (16/83; P = 0·03), but not compared to Ab neg mothers (6/38; P = 0·07). DRB1*12 was found only in Ab neg mothers (5/38) (controls 0/83; P = 0·02). A study of TNF‐α genotype (n = 30, Ab pos; Ab neg; controls) found no difference between the mothers' groups, or mothers and controls.

Presence of DRB1*15 or DRB1*12 may lower the likelihood of HPA‐1a Ab production. Raised frequency of DRB4* was seen in mothers with affected neonates (13/22) (unaffected, Ab pos 3/11; P = 0·02). DRB4* may increase the odds of HPA‐1a alloimmunization. Three NAITP cases due to anti‐HPA‐1a have been reported (2 local, 1 published), involving mothers DRB3*0101 neg, but DRB4* pos. TNF‐α genotype may not predict anti‐HPA‐1a production in these women. Studies with larger groups would establish the value of these markers in defining women at high risk of HPA‐1a alloimmunization.
Accelerated autoantibody clearance by IVIG therapy
Authors: Bleeker W. K. , Teeling J. L. , Hack C. E.
Published: March 2003

Recently, it has been postulated that the beneficial effect of intravenous immunoglobulins (IVIG) in antibody‐mediated autoimmune disorders is based on an accelerated catabolism of autoantibodies due to saturation of FcRn. In the present study, we determined the magnitude and time course of this effect in a computer simulation to see whether this mechanism may indeed explain clinical observations.

A pharmacokinetic model for IgG clearance was established in which concentration changes were calculated in small time steps. To validate the model we performed in vivo experiments with mice. A computational simulation accurately predicted the magnitude of the observed in vivo effect of IVIG administration on endogenous IgG levels.

For human patients our model predicted a gradual decrease in autoantibody levels after IVIG administration (2 g/kg) with a maximum reduction of about 25% after 3–4 weeks and a duration of several months. This corresponds well to the effects of IVIG therapy observed in two patient studies: one on calcium‐channel autoantibodies by Bain et al. (Neurology 1996; 47: 678–683) and another on ANCA by Jayne et al. (J Autoimmunity 1993; 6: 207–219).

In conclusion, accelerated IgG clearance can explain a modest decrease in autoantibody levels observed in several patient studies. However, in some other clinical studies larger or more rapid effects have been observed, which can not be explained by accelerated clearance, suggesting that IVIG can also reduce autoantibody levels via other mechanisms such as down‐regulation of antibody production or neutralization by anti‐idiotypic antibodies.
The FcγRIIIA‐158V allele is a risk factor for ITP
Authors: De Haas M. , Osborne J. , Kleijer M. , Von Dem Borne A. E. G. KR. , Bussel J.
Published: March 2003

In those patients with idiopatic thrombocytopenic purpura (ITP) that do not respond to standard therapy with corticosteroids and/or splenectomy, high‐dose intravenous IgG or low dose anti‐D is given to achieve transient increases in circulating platelet counts. The effect of these treatments is thought to be blockade of macrophage Fc receptors. Macrophages express FcγRIa, FcγRIIa and FcγRIIIa, all three classes may be involved in binding and phagocytosis of sensitized platelets. The FcγRIIa‐131R/H and FcγRIIIa‐158V/F polymorphisms influence the IgG‐binding affinity of the receptors. In literature, skewing of distribution of the FcγRIIA alleles towards the low‐affinity‐genotype: FcγRIIA‐131R/R has been described in ITP patients. Others did not confirm these findings.

We genotyped a group of 83 patients with chronic ITP and 87 ethnically comparable hospital controls for the FcγRIIA, FcγRIIIA and FcγRIIIB polymorphisms, the latter Fc receptor is only expressed by neutrophils.

In the ITP patient population, the gene frequency of the FcγRIIIA‐158V allele was 0·55, vs. 0·43 in the control population (P = 0·001). The genotype distribution of the FcγRIIIA‐V/F polymorphism was also significantly different from that of a control population (P = 0·006). The allele and genotype distributions of the FcγRIIA‐131R/H or the FcγRIIIB‐NA1,2 polymorphisms was similar in patients and controls.

Our observations show that macrophage FcãRIIIa may be the most important receptor in clearance of sensitized platelets, or that FcγRIIIa‐bearing cells (NK cells or macrophages) are implicated in the pathogenesis of this autoimmune disease.
New approaches to antibody‐mediated disease – lessons from characterizing helper T‐cell responses to the RHD protein
Authors: Barker R. N. , Hall A. M. , Stott L. M. , Urbaniak S. J.
Published: March 2003

The key pathogenic contribution of antibodies in conditions such as haemolytic disease of the newborn (HDN) and autoimmune haemolytic anaemia (AIHA) has been recognized for many years. However, although the production of such antibodies is largely thymus‐dependent, offering a potential therapeutic target, little attention has been focused on the T helper‐(Th‐) cells responsible. The RhD protein provides an ideal opportunity to study Th‐cells that drive pathogenic antibody production, since it bears major antigens in both HDN and AIHA. Mapping studies have determined the fine specificity of Rh‐reactive Th‐cells that are activated to proliferate in donors who have been alloimmunized with RhD‐positive erythrocytes, and in AIHA patients. In addition to identifying antigenic peptides that could be used to modulate the responses, this work demonstrates that Th‐cells of different specificities proliferate in the two groups, even when restricted by the same MHC class II element. Our explanation is that the allo‐response targets abundantly presented peptides from the antigen, whereas the autoreactive Th‐cells activated in AIHA are specific for subdominant or cryptic epitopes, which allows them to escape the processes of self‐tolerance. Nonetheless, the respective peptides recognized by the proliferating allo‐or auto‐reactive Th‐cells could, if given by a tolerogenic mucosal route, form the basis of a novel prophylaxis or therapy for Rh antibody‐mediated disease. Alternatively, it may be possible to exploit in vivo the finding that the RhD protein carries regulatory epitopes that elicit Th cytokines such as IL‐10, which can inhibit the proliferative responses in vitro.
Analysis of megakaryocytopoiesis in children with congenital thrombocytopenia
Authors: Van Den Oudenrijn S. , Bruin M. , Folman C. C. , Bussel J. , De Haas M. , Von Dem Borne A. E. G. Kr
Published: March 2003

In this study, we analysed 15 children with congenital thrombocytopenia in order to unravel the underlying mechanisms. Plasma thrombopoietin (Tpo) and glycocalicin (GC, a marker for platelet turnover) levels were measured and megakaryocytopoiesis was studied with an in vitro liquid‐culture system. Based on clinical and laboratory data included patients were divided into three groups. The first group of seven patients was diagnosed with amegakaryocytic thrombocytopenia. In bone marrow from these patients hardly any megakaryocyte was found and also in in vitro cultures no megakaryocyte formation was observed. Plasma Tpo levels were severely increased and GC levels were decreased, both indicating a platelet production defect. Sequence analysis of the gene encoding the Tpo‐receptor, c‐mpl, revealed mutations in five of the six patients analysed. Some mutations lead to the formation of a stopcodon and thus to a direct loss of Mpl function. We are currently investigating whether the other mutations, leading to single amino‐acid replacements, which do not directly predict loss of Mpl function, are causing disruption of Mpl function or expression. The second group of three patients had normal numbers of atypical megakaryocytes in the bone marrow, showed megakaryocyte formation in vitro, normal Tpo plasma levels and severely decreased GC levels. A defect in megakaryocyte maturation and subsequent proplatelet formation of megakaryocytes is likely the cause of the thrombocytopenia in this group. The third group of five patients was diagnosed with Wiskott Aldrich Syndrome (WAS). They had normal Tpo‐, normal GC‐levels and megakaryocyte formation both in vivo and in vitro. Thrombocytopenia in this group is due to an increased platelet turnover, that is WAS related. Concluding, with three parameters, Tpo and GC plasma levels and in vitro megakaryocyte culture, a good distinction between different causes of congenital thrombocytopenia in children can be achieved.
Platelet antibodies in transfused patients
Authors: Kiefel V. , König C. , Kroll H. , Santoso S.
Published: March 2003

P atients and M
ethods  

We studied sera of 252 patients with haematological or oncological diseases, who received multiple transfusions of rbc and/or platelet concentrates. The aim of our study was to analyse the specificity of platelet reactive antibodies (platelet specific and HLA‐antibodies) in a large series of transfused patients. All sera were analysed by platelet immunofluorescence, platelet agglutination, MAIPA and the lymphocytotoxic test (LCT).

R esults 

Platelet reactive antibodies were detected in 113 patients. Among female patients with previous pregnancies 21/36 were alloimmunized but only 7/30 female patients who had not been pregnant (P = 0·0049). In 108 (42·9%) of all sera, we found HLA‐specific antibodies. In a subset of 40 sera, reactions of HLA class I antibodies were compared with an identical cell panel using LCT and MAIPA assay. Twenty‐nine of these sera showed discrepant results in both techniques. Sera from 20 patients contained platelet specific alloantibodies with defined specificity: HPA‐5b (10), HPA‐1b (4), HPA‐5a (2) HPA‐1a (1), HPA‐2b (1), HPA‐5b + HPA‐1b (1), HPA‐1b + HPA‐2b (1). All sera with anti‐HPA‐1b contained additional HLA antibodies, 4 sera with anti‐HPA‐5b apparently contained no HLA‐antibodies. Ten sera contained ‘panreactive’ antibodies with broad specificity: GP IIb/IIIa (4), GP IIb/IIIa + Ia/IIa (2), GPIIb/IIIa + Ib/IX (1), GP IIb/IIIa + Ia/IIa + Ib/IX (3).

C onclusion 

(1) Platelet alloantibody specificities in transfused patients (predominantly anti‐HPA‐5b and − 1b) differ significantly from those observed in neonatal alloimmune thrombocytopenia and post‐transfusion purpura, where anti‐HPA‐1a is the most prevalent specificity. (2) Results for detection of HLA‐antibodies differ considerably in relation to serological technique and target cells.
The frequency of anti‐C + anti‐G in the absence of anti‐D in alloimmunized pregnancies
Authors: Palfi M. , Gunnarsson C.
Published: March 2003

. Anti‐D+C are often initially identified in sera from alloimmunized women. Anti‐G may be present in these samples, mimicking anti‐D+C, and therefore the differentiation of anti‐D, ‐C and ‐G may be important.

Sera from 27 alloimmunized women, initially identified as containing anti‐D + anti‐C, were analysed by adsorption/elution studies in the presence of polyethylene glycol (PEG) using R0r (D+C–G+) and r'r(D–C+G+) red blood cells (RBC). Additionally, 15/27 samples were tested by adsorption in the presence of PEG and subsequently warm elution, using rGr (D–C–G+) RBC. Anti‐G + anti‐C, without anti‐D, were identified in 4/27 samples (14·8%) and none of the newborn children needed postpartum treatment. The combination of D+G, D+C and D+C+G antibodies occurred in 25·9%, 11·1% and 48·1% of the women, respectively. Overall, anti‐G was detected in 24/27 samples (88·9%). Pregnant women shown to have anti‐G+C but not anti‐D should receive Rh immune globulin. Additionally, the finding of apparent anti‐D+C during pregnancy in D‐negative spouses may lead to paternity testing and therefore a correct antibody identification is necessary.
ABO‐incompatibility in solid organ transplantation
Authors: Rydberg L.
Published: March 2003

. The most important transplantation antigen system in solid organ transplantation is the ABO histo‐blood group system. Crossing the ABO barrier in solid organ transplantation is usually not done except for emergency liver transplantations. Early experiences of crossing the ABO barrier in renal transplantation were very disappointing. In the 1970s, clinical trials were started transplanting kidneys of subgroup A2 into blood group O recipients. The tissues of the A2 subgroup expresses reduced amount of A antigens compared to subgroup A1 and the recipients had no special pretreatment and standard immunosuppression. A number of early graft losses were experienced but the trial also resulted in several long time surviving grafts. A few centres have adapted the concept of A2 to non A kidney transplantations with successful results, when the recipient anti‐A titres are low or reduced prior to transplantation.

In the early 1980s one group successfully transplanted A1 and B kidneys from living related donors across the ABO‐barrier using an immunosuppressive protocol consisting of quadruple drugs and splenectomy and this protocol was adapted by a few other groups. In Japan, where cadaver donors are available in very limited number, the largest number of ABO‐incompatible transplantations have been performed. Altogether more than 300 ABO‐incompatible kidney transplantations have been performed in more than 40 centres since 1989.

ABO‐incompatible liver transplantations have been performed mainly in emergency cases and the results have generally been inferior to ABO‐compatible grafts. In children below the age of three years, liver transplantations across the ABO‐barrier have been quite successful especially with living related donors. Very few ABO‐incompatible heart/heart–lung/lung‐transplantations have been reported with a few successful cases, but the majority have been failures. Recently a series of ABO‐incompatible heart transplants performed in small children have been reported with a high success rate.
The risk of antibody formation against HNA1a and HNA1b granulocyte antigens during pregnancy and its relation to neonatal neutropenia
Authors: Żupańska B. , Uhrynowska M. , Guz K. , Maślanka K. , Brojer E. , Czestyńska M. , Radomska I.
Published: March 2003

. The evaluation of immunization by the HNA1a and 1b antigens during pregnancy was based on (i) their genotyping in 1038 unselected mothers and newborns of homozygous mothers, (ii) granulocyte counting in all born infants and (iii) examination of granulocyte antibodies in maternal sera if an HNA1 incompatibile child was born. A total of 548 (52·8%) mothers were heterozygous – thus further examinations were not done. Four hundred and ninety (47·2%) were homozygous, of whom 203 (41·3%) delivered an incompatible child, i.e. 19·6% of all the infants. Among available sera from 195 mothers with feto‐maternal incompatibility, the granulocyte‐specific antibodies were found in nine (4·5%); six of these (3%) were HNA1 (four anti‐1a, two anti‐1b), and in three others the specificity was not determined. In the remaining 28 sera, the only antibodies detected were HLA. Hence, six out of 1000 pregnant women can be expected to develop anti‐HNA1. In none of the newborns was the cord neutrophil count below 1·5 × 109 L−1 and signs of infection found, thus the incidence of NAIN seems to be lower than 1 per 1000 infants. A comparison with our previous, unpublished data suggests that the incidence of severe NAIN is roughly 1 per 6000 (four cases among 24101 newborns).
JAHK: a low frequency antigen associated with the rG complex of the Rh blood group system
Authors: Green C. , Coghlan G. , Bizot M. , Kasulke D. , Bombail‐Girard M. , Wallace M. , Lomas‐Francis C. , Daniels G.
Published: March 2003

Summary We have investigated a ‘new’ low frequency antigen JAHK, which is a marker for the rare Rh gene complex rG. The rG haplotype does not produce any D, c or E antigens, but does produce a strong G antigen. The rG haplotype [d(C)(e)G] is associated with weak C and weak e antigens. Three unrelated rG/dce individuals and one rG/rG propositus were JAHK+. In addition, three propositi whose red cells had a typical expression of C and/or e antigen, which could not be shown to be rG because of a normal D antigen produced by the haplotype in trans, were also JAHK+. Families of three of the propositi demonstrate the inheritance of JAHK as a Mendelian dominant character. It is likely that the JAHK antigen results from conformational changes in an RhCcEe protein that has the amino acid characteristic of c antigen at position 16 and the amino acid residues characteristic of C antigen at positions 60, 68, and 103. JAHK has been assigned the number RH53.
Analysis of human platelet antigen systems in a Moroccan Berber population
Authors: Ferrer G. , Muñiz‐Diaz E. , Aluja M. P. , Arilla M. , Martinez C. , Nogués R. , Servin A. , Baali A.
Published: March 2003

Summary The human platelet alloantigen systems HPA‐1, ‐2, ‐3, ‐4, ‐5 and ‐6 in a Moroccan Berber population from the Amizmiz region were determined by polymerase chain reaction using sequence‐specific primers (PCR‐SSP). The gene frequencies obtained from these unrelated Berbers were 0·747 and 0·252 for HPA‐1a and ‐1b; 0·817 and 0·182 for HPA‐2a and ‐2b; 0·682 and 0·317 for HPA‐3a and ‐3b; 1 and 0·0 for HPA‐4a and ‐4b; 0·8616 and 0·1383 for HPA‐5a and ‐5b; 1 and 0·0 for HPA‐6a and ‐6b. The Berbers have the highest frequency for the 1b, 2b and 5b alleles of all the populations reported to date and also the lowest frequency for the 3b allele.
The United Kingdom National External Quality Assessment Scheme (Blood Transfusion Laboratory Practice): trends in proficiency and practice between 1985 and 2000
Authors: Knowles S. M. , Milkins C. E. , Chapman J. F. , Scott M.
Published: March 2003

Summary Proficiency in blood transfusion laboratory practice has improved over the last 15 years (1985–2000). Error rates for ABO grouping have fallen from 0·19 to 0·02% (P = 0·003). A similar trend is evident for antibody screening, with error rates for false negative antibody screens falling from 3·2 to 0·5% (P < 0·001), and for antibody identification, for sera containing a single alloantibody, with error rates falling from 8·8% to 0·9% over the last 10 years (P < 0·001). Proficiency in serological crossmatching to detect clinically significant non‐ABO incompatibilities (other than Kidd), has also improved (P < 0·001). However, error rates for RhD grouping have not changed and there has been a recent decline in proficiency in detecting weak ABO incompatibilities and Kidd antibodies with heterozygous cells.

Procedures for pretransfusion testing have also been rationalized over this time. The indirect antiglobulin test (IAT) is used in isolation by 73% of laboratories for antibody screening, and 10% rely on the direct room temperature (DRT), immediate spin (IS) crossmatch in the absence (current and/or historical) of clinically significant antibodies.

Despite the improvements in error rates that have occurred alongside the rationalizations, there is still evidence of noncompliance with current published BCSH guidelines and manufacturers' instructions.
Clinical findings in female genital schistosomiasis in Madagascar
Authors: Leutscher P. , Ravaoalimalala V. E. , Raharisolo C. , Ramarokoto C. E. , Rasendramino M. , Raobelison A. , Vennervald B. , Esterre P. , Feldmeier H.
Published: March 2003

To assess the morbidity of S. haematobium infection in women of reproductive age (15–49 years) in the western part of Madagascar, the village of Betalatala with a prevalence of urinary schistosomiasis in women of 75.6% (95% confidence limit 69.3 to 81.9%) was compared with a neighbouring village with similar socio‐economic characteristics and a prevalence of 5.0% (95% confidence limit 0 to 11.75%). The women were questioned in Malagasy about obstetrical history and urogynecological symptoms. They were examined gynaecologically, parasitologically and by ultrasonography. Important STDs were excluded by appropiate diagnostics. In Betalatala significantly more women reported a history of spontaneous abortion (P < 0.01), complaints of irregular menstruation (P < 0.001), pelvic pain (<0.05), vaginal discharge (P < 0.0001), dysuria (P < 0.05) and haematuria (P < 0.01) than in the control village. Biopsies were obtained from the cervix of 36 women with macroscopical lesions, and in 12 cases S. haematobium eggs were found by histological sectioning (33.3%). In the control village no eggs were detected in the histological sections of biopsies taken from 14 women. (P < 0.05). Infections with Candida albicans, Trichomonas vaginalis, Gardnerella vaginalis and Treponema pallidum were found in similar frequencies in both villages. In 9.8% of the women in Betalatala abnormalities of the upper reproductive tract were revealed by ultrasonography versus none in the women from the control village (P < 0.05). Echographic abnormalities of the urinary tract were present in 24% and 3% of the women in the study village and in the control village, respectively (P < 0.0001). These findings were accompanied by an elevated frequency of haematuria (55% versus 20%) and proteinuria (70.4% versus 25%) in the study population (P < 0.0001). Our study indicates that S. haematobium infection in women may not only cause symptoms in the urinary tract, but also frequently in the lower and upper reproductive tract.
Financing changes of schistosomiasis control programmes in China 1980–1995: a case study in Songzi county
Authors: Xiang Huiyun , Yu Songli , Sun Yi  , Dai Yuhai
Published: March 2003

Summary To assess the financing changes of schistosomiasis control programmes in China and estimate the impact of these changes on patients' treatment‐seeking behaviour and control of schistosomiasis, a survey was conducted in five schistosomiasis‐endemic areas of the lake regions, Hubei province, in 1996. This paper reports financing changes and their impact on the incidence and prevalence of schistosomiasis from one of the five areas as a case study. By examining the surveillance and financial data from 1980 to 1995, and through focus group discussions we found that the schistosomiasis control programmes in People's Republic of China have gone through dramatic financing changes from 1980 to 1995, when the transitions of China's social, economic, and political systems happened. The proportions of funding to schistosomiasis control programmes from high level governmental agencies, county budgets, and services revenue changed from 60%, 23%, and 17%, respectively, in 1980–1987 to 0.7%, 22.3%, and 72% in 1995. The percentageages of expenditure of schistosomiasis control activities, salaries and bonuses, and other activities unrelated to schistosomiasis control, were 53.5%, 14.4, and 17.2% in 1980. These percentageages changed to 7.7%, 33.3%, and 53.3%, respectively, in 1995. The preponderant role of the state in organizing, financing, and delivery of the services was replaced with the new system which is more influenced by the market economy. The incidence and the prevalence of schistosomiasis in the study area have increased year by year from 1980 to 1990, although there has been a tendency to decrease after 1991 but not to the low pre1980 levels. The collapse of the community‐based medical system in rural areas and the dramatic financing changes of schistosomiasis control programmes have created major difficulties for schistosomiasis control in China.
Polymorphisms in the dihydrofolate reductase (DHFR) and dihydropteroate synthetase (DHPS) genes of Plasmodium falciparum and in vivo resistance to sulphadoxine/pyrimethamine in isolates from Tanzania
Authors: Jelinek T. , Jelinek T. , Rønn A. M. , Lemnge M. M. , Curtis J. , Mhina J. , Duraisingh M. T. , Bygbjerg I. C. , Warhurst D. C.
Published: March 2003

The efficacy of sulphadoxine/pyrimethamine (S/P) in treatment of uncomplicated falciparum malaria in Africa is increasingly compromised by development of resistance. The occurrence of mutations associated with the active site sequence in the Plasmodium falciparum genes coding for dihydrofolate reductase (DHFR) and dihydropteroate synthetase (DHPS) is associated with in vitro resistance to pyrimethamine and sulphadoxine. This study investigates the occurrence of these mutations in infected blood samples taken from Tanzanian children before treatment with S/P and their relationship to parasite breakthrough by day 7. The results show that alleles of DHPS (436‐alanine, 437‐alanine and 540‐lysine) were significantly reduced in prevalence on day 7 after S/P treatment. In this area, a DHPS with 436‐serine, 437‐glycine and 540‐glutamate appears to play a major role in resistance to S/P in vivo. Evidence for the influence of mutations in the DHFR gene in this investigation is not clear, probably because of the high prevalence of ‘resistance‐related’ mutations at day 0 in the local parasite population. For apparently the same reason, it was not possible to show a statistical association between S/P resistance and the presence of particular polymorphisms in the DHFR and DHPS genes before treatment.
Isolate resistance of Blastocystis hominis to metronidazole
Authors: Haresh K. , Suresh K. , Anuar A. Khairul , Saminathan S.
Published: March 2003

Summary Isolates of Blastocystis hominis from infected immigrant workers from Indonesia, Bangladesh and infected individuals from Singapore and Malaysia were assessed for growth pattern and degree of resistance to different concentrations of metronidazole. Viability of the cells was assessed using eosin‐brilliant cresyl blue which stained viable cells green and nonviable cells red. The Bangladeshi and Singaporean isolates were nonviable even at the lowest concentration of metronidazole at 0.01 mg/ml. The parasites of the Malaysian isolate were resistant to a concentration of 0.01 mg/ml, whereas 40% of the initial inoculum of parasites from the Indonesian isolate at day one were still viable in cultures with 1.0 mg/ml metronidazole. The study shows that isolates of B. hominis of different geographical origin have different levels of resistance to metronidazole. The search for more effective drugs to eliminate the parasite appears inevitable, especially since surviving parasites from metronidazole cultures show greater ability to multiply in subcultures than controls.
Screening of army soldiers for Wuchereria bancrofti infection in the metropolitan Recife region, Brazil: implications for epidemiological surveillance
Authors: Medeiros Zulma , Gomes Judilson , Béliz Fátima , Coutinho Amaury , Dreyer Patricia , Dreyer Gerusa
Published: March 2003

SummaryBetween 1989 and 1995, blood surveys were performed for Wuchereria bancrofti infection in several barracks of the Brazilian army in the metropolitan Recife region. For initial screening, 60 μl of capillary blood were examined for microfilaria. All men who tested positive had microfilaria quantified by filtration of venous blood through a polycarbonate membrane. Of 23773 men screened, 585 (2.5%) had microfilaria (mf). Microfilarial density ranged from < 1–8706 mf/ml of blood. Thirteen individuals had ultra‐low microfilarial densities (1 mf/11 ml of blood). Characterization of 174 autochthonous cases made it possible to map 8 new districts in 4 cities within metropolitan Recife region where transmission of W. bancrofti was previously unknown. Routine screening of soldiers in the military may provide important surveillance data for national programmes to eliminate transmission of W. bancrofti.