bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential expression and function of Arabidopsis thaliana NHX Na+/H+ antiporters in the salt stress response
Authors: Yokoi Shuji , Quintero Francisco J. , Cubero Beatriz , Ruiz Maria T. , Bressan Ray A. , Hasegawa Paul M. , Pardo Jose M.
Published: March 2003

The Arabidopsis thaliana vacuolar Na+/H+ antiporter AtNHX1 is a salt tolerance determinant. Predicted amino acid sequence similarity, protein topology and the presence of functional domains conserved in AtNHX1 and prototypical mammalian NHE Na+/H+ exchangers led to the identification of five additional AtNHX genes (AtNHX2–6). The AtNHX1 and AtNHX2 mRNAs are the most prevalent transcripts among this family of genes in seedling shoots and roots. A lower‐abundance AtNHX5 mRNA is present in both shoots and roots, whereas AtNHX3 transcript is expressed predominantly in roots. AtNHX4 and AtNHX6 mRNAs were detected only by RT–PCR. AtNHX1, 2 or 5 suppress, with differential efficacy, the Na+/Li+‐sensitive phenotype of a yeast mutant that is deficient in the endosomal/vacuolar Na+/H+ antiporter ScNHX1. Ion accumulation data indicate that these AtNHX proteins function to facilitate Na+ ion compartmentalization and maintain intracellular K+ status. Seedling steady‐state mRNA levels of AtNHX1 and AtNHX2 increase similarly after treatment with NaCl, an equi‐osmolar concentration of sorbitol, or ABA, whereas AtNHX5 transcript abundance increases only in response to salt treatment. Hyper‐osmotic up‐regulation of AtNHX1, 2 or 5 expression is not dependent on the SOS pathway that controls ion homeostasis. However, steady‐state AtNHX1, 2 and 5 transcript abundance is greater in sos1, sos2 and sos3 plants growing in medium that is not supplemented with sorbitol or NaCl, providing evidence that transcription of these genes is negatively affected by the SOS pathway in the absence of stress. AtNHX1 and AtNHX2 transcripts accumulate in response to ABA but not to NaCl in the aba2‐1, mutant indicating that the osmotic responsiveness of these genes is ABA‐dependent. An as yet undefined stress signal pathway that is ABA‐ and SOS‐independent apparently controls transcriptional up‐regulation of AtNHX5 expression by hyper‐saline shock. Similar to AtNHX1, AtNHX2 is localized to the tonoplast of plant cells. Together, these results implicate AtNHX2 and 5, together with AtNHX1, as salt tolerance determinants, and indicate that AtNHX2 has a major function in vacuolar compartmentalization of Na+.
KCl activates phospholipase D at two different concentration ranges: distinguishing between hyperosmotic stress and membrane depolarization
Authors: Meijer Harold J. G. , Ter Riet Bas , Van Himbergen John A. J. , Musgrave Alan , Munnik Teun
Published: March 2003

Hyperosmotic stress induces the rapid formation of phosphatidic acid (PA) in Chlamydomonas moewusii via the activation of two signalling pathways: phospholipase D (PLD) and phospholipase C (PLC), the latter in combination with diacylglycerol kinase (DGK) (Munnik et al., 2000). A concomitant increase in cell Ca2+ becomes manifest as deflagellation. When KCl was used as osmoticum we found that two concentration ranges activated deflagellation: one between 50 and 100 mm and another above 200 mm. Deflagellation in low KCl concentrations was complete within 30 sec whereas in high concentrations it took 5 min. PLC was not activated, as it was by high KCl concentrations that cause hyperosmotic stress. Moreover PLD was activated more strongly by low than by high KCl concentrations. Potassium was the most potent monovalent cation based on the induction of deflagellation and the formation of PA and PBut. During treatment, the external medium acidified, indicating an increase in H+‐ATPase activity in order to re‐establish the membrane potential. Activation of PLD and deflagellation at low KCl concentrations were abrogated by treatment with La3+, Gd3+ and EGTA, indicating the dependency on extracellular Ca2+. This suggests that low concentrations of KCl depolarize the plasma membrane, resulting in the activation of H+‐ATPases and opening voltage‐dependent Ca2+± channels, observed as deflagellation and an increase in PLD activity.
Femtosecond near‐infrared laser pulses as a versatile non‐invasive tool for intra‐tissue nanoprocessing in plants without compromising viability
Authors: Tirlapur Uday K. , König Karsten
Published: March 2003

In this report, we describe a highly reproducible femtosecond near‐infrared (NIR) laser‐based nanoprocessing technique that can be used both for non‐invasive intra‐tissue nanodissection of plant cell walls as well as selective destruction of a single plastid or part thereof without compromising the viability of the cells. The ultra‐precise intra‐tissue nanoprocessing is achieved by the generation of high light intensity (1012 W cm−2) by diffraction‐limited focusing of the radiation of an NIR (λ = 740 and 800 nm) femtosecond titanium‐sapphire laser to a sub‐femtolitre volume and subsequent highly localized instantaneous plasma formation. Following nanosurgery, electron microscopical analysis of the corresponding cellular target areas revealed clean non‐staggering lesions across the cell wall with a cut width measuring less than 400 nm. To our knowledge, this is the smallest cut made non‐invasively within a plant tissue. Further evidence, including two‐photon imaging of chlorophyll fluorescence, revealed that a single target chloroplast or part thereof can be completely knocked out using intense ultra‐fast NIR pulses without any visible deleterious effect on the adjacent plastids. The vitality of the cells after nanoprocessing has been ascertained by exclusion of propidium iodide from the cells as well as by the presence of cytoplasmic streaming. The potential applications of this technical advance include developmental biology applications, particularly studies addressing spatio‐temporal control of ontogenetic events and cell–cell interactions, and gravitational biology applications.
In planta protein–protein interactions assessed using a nanovirus‐based replication and expression system
Authors: Aronson Marie N. , Complainville Arnaud , Clérot Danielle , Alcalde Hélène , Katul Lina , Vetten H. Josef , Gronenborn Bruno , Timchenko Tatiana
Published: March 2003

The multipartite genome of the nanovirus Faba bean necrotic yellows virus, which consists of one gene on each DNA component, was exploited to construct a series of virus‐based episomal vectors designed for transient replication and gene expression in plants. This nanovirus based expression system yields high levels of protein which allows isolation of recombinant protein and protein complexes from plant tissues. As examples, we demonstrated in planta interaction between the nanovirus F‐box protein Clink and SKP1, a constituent of the ubiquitin‐dependent protein turnover pathway. Thus, replicative nanovirus vectors provide a simple and efficient means for in planta characterization of protein‐protein interaction.
Light‐signalling pathways leading to the co‐ordinated expression of HEMA1 and Lhcb during chloroplast development in Arabidopsis thaliana
Authors: McCormac Alex C. , Terry Matthew J.
Published: March 2003

During de‐etiolation, the co‐ordinated synthesis of chlorophyll and the chlorophyll a/b‐binding proteins is critical to the development of functional light‐harvesting complexes. To understand how this co‐ordination is achieved, we have made a detailed study of the light‐regulated signalling pathways mediating the expression of the HEMA1 and Lhcb genes encoding glutamyl‐tRNA reductase, the first committed enzyme of 5‐aminolaevulinic acid formation, and chlorophyll a/b‐binding proteins, respectively. To do this, we have screened 7 photoreceptor and 12 light‐signalling mutants of Arabidopsis thaliana L. for induction of HEMA1 and Lhcb expression in continuous red, far‐red and blue light and following a red pulse. We have categorised these mutants into two groups. The phyA, phyB, phyAphyB, cry1, cry2, cop1, det1, poc1, eid1, and far1 mutations lead to diverse effects on the light regulation of HEMA1, but affect Lhcb expression to a similar degree. The hy1, hy2, hy5, fin219, fhy1, fhy3, spa1, ndpk2, and pat1 mutants also affect light regulation of both HEMA1 and Lhcb expression, but with differences in the relative magnitude of the two responses. The fhy1 and fhy3 mutants show the most significant differences in light regulation between the two genes, with both showing a strong inhibition of HEMA1 expression under continuous red light. These results demonstrate that co‐ordinated regulation of HEMA1 and Lhcb is largely achieved through parallel light regulation mediated by shared phytochrome‐ and cryptochrome‐signalling pathways. However, glutamyl‐tRNA reductase is also required for the synthesis of other tetrapyrroles and this dual role may account for the observed differences in these light‐signalling pathways.
A genomics approach to the early stages of triterpene saponin biosynthesis in Medicago truncatula
Authors: Suzuki Hideyuki , Achnine Lahoucine , Xu Ran , Matsuda Seiichi P. T. , Dixon Richard A.
Published: March 2003

The saponins of the model legume Medicago truncatula are glycosides of at least five different triterpene aglycones: soyasapogenol B, soyasapogenol E, medicagenic acid, hederagenin and bayogenin. These aglycones are most likely derived from β‐amyrin, a product of the cyclization of 2,3‐oxidosqualene. Mining M. truncatula EST data sets led to the identification of sequences putatively encoding three early enzymes of triterpene aglycone formation: squalene synthase (SS), squalene epoxidase (SE), and β‐amyrin synthase (β‐AS). SS was functionally characterized by expression in Escherichia coli, two forms of SE by complementation of the yeast erg1 mutant, and β‐AS by expression in yeast. β‐Amyrin was the sole product of the cyclization of squalene epoxide by the recombinant M. truncatulaβ‐AS, as judged by GC–MS and NMR. Transcripts encoding β‐AS, SS and one form of SE were strongly and co‐ordinately induced, associated with accumulation of triterpenes, upon exposure of M. truncatula cell suspension cultures to methyl jasmonate. Sterol composition remained unaffected by jasmonate treatment. Molecular verification of induction of the triterpene pathway in a cell culture system provides a new tool for saponin pathway gene discovery by DNA array‐based approaches.
The chlorate‐resistant and photomorphogenesis‐defective mutant cr88 encodes a chloroplast‐targeted HSP90
Authors: Cao Dongsun , Froehlich John E. , Zhang Hui , Cheng Chi‐Lien
Published: March 2003

The cr88 mutant of Arabidopsis is a novel chlorate‐resistant mutant that displays long hypocotyls in red light, but not in far red or blue light, and is delayed in the greening process. In cotyledons and young leaves, plastids are less developed compared with those of the wild type. In addition, a subset of light‐regulated genes are under‐expressed in this mutant. To understand the pleiotropic phenotypes of cr88, we isolated the CR88 gene through map‐based cloning. We found that CR88 encodes a chloroplast‐targeted 90‐kDa heat shock protein (HSP90). The CR88 gene is expressed at highest levels during early post‐germination stages and in leaves and reproductive organs. It is constitutively expressed but is also light and heat shock inducible. Chloroplast import experiments showed that the protein is localized to the stroma compartment of the chloroplast. The possible function of an HSP90 in the chloroplast and a plausible explanation of the pleiotropic phenotypes observed in cr88 are discussed.
Alteration of GCN5 levels in maize reveals dynamic responses to manipulating histone acetylation
Authors: Bhat Riyaz A. , Riehl Marcus , Santandrea Geraldina , Velasco Riccardo , Slocombe Stephen , Donn Günter , Steinbiss Hans‐Henning , Thompson Richard D. , Becker Heinz‐Albert
Published: March 2003

The role played by histone acetyltransferase (HAT), GCN5, in transcriptional co‐activation has been analysed in detail in yeast and mammals. Here, we present the cloning and expression pattern of Zmgcn5, the maize homologue. The enzymatic activity of the recombinant ZmGCN5 was analysed with histone and nucleosome substrates. In situ hybridisation of developing maize kernels using Zmgcn5 as probe shows that the transcript is concentrated in rapidly dividing cells. To investigate the role of ZmGCN5 in the transcription of specific plant genes, direct protein–protein interactions were tested. A cDNA clone encoding a putative interacting partner in GCN5‐adapter complexes, ZmADA2, was isolated and the interaction between ZmGCN5 and ZmADA2 was confirmed by a GST‐spin down experiment. Co‐immunoprecipitation of the plant transcriptional activator Opaque‐2 and ZmADA2 in nuclear extracts suggests ADA2/GCN5‐containing complexes to mediate transcriptional activation by binding of this bZIP factor. For a more general analysis of the effects of histone acetylation on plant gene expression, 2500 ESTs spotted on filters were hybridised with cDNA probes derived either from maize cell lines treated with Trichostatin A (TSA), or from a transgenic line expressing the ZmGCN5 antisense transcript. Several sequences showing marked changes in abundance were confirmed by RNA blot analysis. Inhibition of histone deacetylation with TSA is accompanied by a decrease in the abundance of ZmGCN5 acetylase protein, but by increases in mRNAs for histones H2A, H2B, H3 and H4. The elevated histone mRNA levels were not reflected in increasing histone protein concentrations, suggesting hyperacetylated histones arising from TSA treatment may be preferentially degraded and substituted by de novo synthesised histones. The ZmGCN5 antisense material showed suppression of the endogenous ZmGCN5 transcript and the profiling analysis revealed increased mRNA levels for H2A, H2B and H4. Furthermore, in the antisense line, a reduction in the amount of the RPD3‐type HD1B‐I histone deacetylase protein was observed. A model for linked regulation of histone acetylation and histone mRNA transcription is discussed.
ADP‐glucose pyrophosphorylase from potato tuber: site‐directed mutagenesis of homologous aspartic acid residues in the small and large subunits
Authors: Frueauf Jeremiah B. , Ballicora Miguel A. , Preiss Jack
Published: March 2003

Asp142in the homotetrameric ADP‐glucose pyrophosphorylase (ADP‐Glc PPase) enzyme fromEscherichia coliwas demonstrated to be involved in catalysis of this enzyme [Frueauf, J.B., Ballicora, M.A. and Preiss J. (2001)J. Biol. Chem., 276, 46319–46325]. The residue is highly conserved throughout the family of ADP‐Glc PPases, as well as throughout the super‐family of sugar–nucleotide pyrophosphorylases. In the heterotetrameric ADP‐Glc PPase from potato (Solanum tuberosumL.) tuber, the homologous residue is present in both the small (Asp145) and the large (Asp160) subunits. It has been proposed that the small subunit of plant ADP‐Glc PPases is catalytic, while the large subunit is modulatory; however, no catalytic residues have been identified. To investigate the function of these conserved Asp residues in the ADP‐Glc PPase from potato tuber, we used site‐directed mutagenesis to introduce either an Asn or a Glu. Kinetic analysis in the direction of synthesis or pyrophosphorolysis of ADP‐Glc showed a significant decrease (more than four orders of magnitude) in the specific activity of the SD145NLwt, SD145NLD160N, and SD145NLD160Emutants, while the effect was smaller (approximately two orders of magnitude) with the SD145ELwt, SD145ELD160N, and SD145ELD160Emutants. By contrast, mutation of the large subunit alone did not affect the specific activity but did alter the apparent affinity for the activator 3‐phosphoglycerate, showing two types of apparent roles for this residue in the different subunits. These results show that mutation of Asp160of the large subunit does not affect catalysis, thus the large subunit is not catalytic, and that the negative charge of Asp145in the small subunit is necessary for enzyme catalysis.
The Arabidopsis ABORTED MICROSPORES (AMS) gene encodes a MYC class transcription factor
Authors: Sorensen Anna‐Marie , Kröber Sandra , Unte Ulrike S. , Huijser Peter , Dekker Koen , Saedler Heinz
Published: March 2003

Visual screening of a T‐DNA mutagenised population of Arabidopsis thaliana for an absence of silique elongation lead to the isolation of the aborted microspores (ams) mutant that shows a sporophytic recessive male sterile phenotype. Homozygous mutant plants are completely devoid of mature pollen. Pollen degeneration occurs shortly after release of the microspores from the tetrad, prior to pollen mitosis I. Premature tapetum and microspore degeneration are the primary defects caused by this lesion, while a secondary effect is visualised in the stamen filaments, which are reduced in length and lie beneath the receptive stigma at flower opening. The disrupted gene was isolated and revealed a T‐DNA element to be inserted into the eighth exon of a basic helix‐loop‐helix (bHLH) gene located on chromosome II. This protein sequence contains a basic DNA binding domain and two alpha helices separated by a loop, typical of a transcription factor belonging to the MYC sub family of bHLH genes. Therefore, AMS plays a crucial role in tapetal cell development and the post‐meiotic transcriptional regulation of microspore development within the developing anther.
Nurse practitioner support for transfusion in patients with haematological disorders in hospital and at home
Authors: Craig  , Milligan  , Cairns  , McClelland  , Parker
Published: March 2003

We prospectively studied the effects of dedicating a nurse to manage the provision of blood product support in a hospital haematology unit and at home to 45 patients with haematological disorders requiring regular transfusion. During the study 335 home blood tests, 65 home platelet transfusions and 155 hospital transfusions were managed by the nurse who organized the whole transfusion process, made home visits for blood tests and platelet transfusions and arranged hospital visits for red cell transfusions. Two hundred clinic visits and 65 day hospital attendances were avoided. The nurse‐led service resulted in a significant reduction in the waiting time from admission to transfusion and in the total length of in‐patient stay. The importance of and satisfaction with different aspects of the care of the transfusion process assessed by a ranking questionnaire showed improved satisfaction scores for all aspects of care. Preference for home blood sampling instead of hospital increased from 24% before to 100% after intervention. We have shown that a dedicated transfusion nurse provides a quality service between hospital and home that is greatly appreciated by patients requiring regular transfusions.
The efficient organization of blood donation: what determines the number of donors and donations?
Authors: Van Der Pol  , Cairns  , Galea
Published: March 2003

This study investigates the influence of socio‐economic characteristics on the number of donors in geographical areas in the North of Scotland as well as the influence of panel‐specific characteristics on the number of donations obtained from each panel in the North of Scotland. Econometric models are applied to establish the relationship between these variables. The main findings were that there was no scope for identifying new areas in which to locate new panels or extend the activities of existing panels because the number of donors in an area was closely correlated with population. There was therefore little scope for recruitment of donors by focusing on any particular area. It was found that annual donations per panel are significantly influenced by the number of donors invited, the number of opportunities to donate and the average length of sessions. Combined with cost information, increasing the length of the session appears to be the most cost‐effective means of collecting higher volumes.
Improvement in transfusion safety using a specially designed transfusion wristband
Authors: Lau  , Wong  , Chui  , Ng  , Cheng
Published: March 2003

Fatal haemolytic transfusion reaction due to ABO incompatibility occurs mainly as a result of clerical error. A blood sample drawn from the wrong patient and labelled as another patient's will not be detected by the blood bank unless there is a previous ABO grouping result. We report here the detection of such clerical error by the use of a specially designed transfusion wristband. The wristband has the following special features: (i) once attached, it cannot be removed except by cutting; (ii) it has a pocket containing a transfusion label; (iii) a unique transfusion barcode is printed on each transfusion label and the corresponding wristband simultaneously by computer technology; (iv) a transfusion label removed from the wristband after attachment to the patient has a characteristic tear‐mark distinguishing it from one removed prior to attachment. The blood bank only accepted those specimens bearing the tear‐marked transfusion labels. All blood units for this patient were labelled with this unique transfusion code together with the patient's details. The nurses counter‐checked the transfusion code on the blood units against the transfusion code on the patient's transfusion wristband prior to transfusion. If the blood sample for compatibility testing was drawn from the ‘wrong’ patient, the intended patient either did not carry a wristband or the transfusion codes did not match at all.  Pretransfusion compatibility tests were performed on 2189 patient samples using this procedure. It was well accepted by both ward and blood bank staff. Two potential mismatched transfusions were avoided. These two clerical errors would not have been detected because neither patient had previous ABO grouping results.
Comparison between a new PVC platelet storage container (UPX80) and a polyolefin container
Authors: Kostelijk  , Gouwerok  , Veldman  , De Korte
Published: March 2003

During storage of platelet concentrates (PCs), the quality of the platelets deteriorates gradually, partially dependent on gas exchange.  UPX80 (JMS, Japan) 1‐L platelet storage PVC containers with increased gas transport capacity were compared with 1‐ and 1.5‐L polyolefin (PO) containers (NPBI, the Netherlands) with filtered PCs stored either in GAC (gluconate–acetate–citrate, < 10% plasma) or in plasma, for 8 days. In total 32 PCs were made (260–330 × 109 platelets per concentrate), equally divided over different bags and storage media. During storage, gas exchange, metabolic, physical and activation parameters were measured.  No consistent differences for all parameters were observed between UPX80 and PO containers (1‐L or 1.5‐L). Blood gas parameters indicated better gas exchange for UPX80 containers compared with PO containers. Good morphology was observed in UPX80 and metabolic functions were not significantly different compared with PO containers. During prolonged storage (after day 6), some significant differences in CD62P and CD63 expression were found, indicating a higher degree of platelet activation in UPX80 containers, especially in GAC.  UPX80 PC containers are suitable for storage of PCs. Although in UPX80 better gas exchange is demonstrated, as compared with PO containers, this does not improve the platelet quality during storage for 6 days, indicating that gas exchange above the level of PO containers has no effect on the switch to aerobic metabolism in platelets.
Elucidation of the primary structure of NB1‐glycoprotein (HNA‐2a antigen)
Authors: Kissel K. , Santoso S. , Hofmann C. , Stroncek D. , Bux J.
Published: March 2003

The human granulocyte antigen HNA‐2a is expressed on a subpopulation of granulocytes in 97% of Caucasians. Alloimmunization against this antigen can cause alloimmune neonatal neutropenia, transfusion‐related acute lung injury and immune neutropenia after bone marrow transplantation. Immunochemical studies demonstrated that NB1 allogenic determinants reside on a GPI‐linked glycoprotein with apparent molecular weight of 56–64 kDa. To elucidate the primary structure of NB1 protein, we isolated NB1 protein using NB1‐specific mab 7D8 by affinity chromatography. Identity of the purified glycoprotein was confirmed by immunoblot using a panel of seven human and two monoclonal NB1‐specific antibodies. Determination of molecular mass using MALDI yielded a 50·6 kDa glycoprotein. After removal of N‐linked carbohydrates, the molecular mass was reduced to 43·1 kDa. N‐terminal amino acid sequencing by Edman degradation yielded a 20 aa‐peptide. Rapid amplification of cDNA ends derived from an HPA‐2a (+) individual using primers based on N‐terminal amino acid sequence resulted in a 1614‐bp cDNA, consisting of a 27‐bp 5′ untranslated region, a 1311‐bp open reading frame and a 276‐bp 3′ untranslated region. The 1311 bp open reading frame encoded for 437 amino acids, of which the first 21 were found to form the signalpeptide. The identity of this transkript was confirmed by the expression of full lengh NB1 cDNA (bp 22–1394) in COS‐7 cells.
Prion protein release in apheresis platelet concentrates is associated with platelet activation and lysis
Authors: Bessos H. , Drummond O. , Prowse C. , Turner M. , MacGregor I.
Published: March 2003

Studies using a time‐resolved fluoroimmuno‐assay (DELFIA®) have shown that platelets and plasma are the main compartments of the normal isoform of prion protein (PrPc) in human blood. The aim of the present study was to monitor PrPc levels in various fractions of apheresis platelet concentrates (PC) during storage and to assess the association of this release with α granule protein β‐thrombo‐globulin (BTG) and cytoplasmic lactate dehydrogenase (LDH) as cell activation and cell lysis markers, respectively. Six apheresis PC were obtained from volunteer donors using Haemonetics MCS and stored up to 10 days. 7–9 mL samples were aseptically collected from each PC on days 1, 2, 3, 4, 5, 8, and 10 of storage. Platelet poor plasma (PPP) and platelets (P) were prepared and the PPP split into two equal fractions, one of which was centrifuged at 40,000g for 2 h at 4 °C to remove microparticles (MP) [high spun plasma (HSP)]. The results showed that the mean overall levels of PrPc during storage remained within 15% of day 1 levels. In contrast, the mean levels in P significantly decreased to 46% day 1 levels by day 10 of storage (P < 0·01), while the corresponding levels in plasma significantly rose by up to 329% (P < 0·01). Moreover, although MP‐bound PrPc was released during storage, it was increasingly superceded by soluble prion protein. PrPc and BTG release exhibited very similar patterns, both showing a significant rise in PPP and HSP (P < 0·01). In contrast, LDH showed a significant rise in HSP only towards the end of the storage period (P < 0·01). These results indicate that PrPc is released from platelets during storage of PCs and that this release is probably due mainly to platelet activation and α‐granule release in the first few days of storage. The release of PrPc is apparently reinforced through cell lysis, and is increasingly comprised of soluble prion proteins, at longer storage periods.
Anti‐idiotype interactions are not required for the protective effect of intravenous immunoglobulin in a murine model of passively transferred immune thrombocytopenia
Authors: Lazarus A. H. , Song S. , Freedman J. , Crow A. R.
Published: March 2003

The use of intravenous immunoglobulin (IVIg) as a treatment for autoimmune disease was first realized in the reversal of thrombocytopenia in Immune Thrombocytopenic Purpura (ITP) and, based upon this finding, it is currently used to treat an increasing number of autoimmune states and to prevent graft rejection. Previous reports, as well as much recent literature, have implied that the reactivity of variable region‐reactive (anti‐idiotypic) antibodies predominantly accounts for the beneficial effects of IVIg. Anti‐idiotypic antibodies reactive with endogenous Ig, autoantibodies, and those anti‐idiotype antibodies that modulate immune function have been implicated. Herein, we demonstrate that IVIg ameliorates thrombocytopenia in a passively transferred model of ITP (P‐ITP) and that anti‐idiotype antibodies present in IVIg ameliorates thrombocytopenia in a passively transferred model of ITP (P‐ITP) and that anti‐idiotype antibodies present in IVIg are not required for its protective effect. Both IVIg and an anti‐idiotype‐depleted IVIg protected equally against thrombocytopenia. As well, immunocompromised (SCID) mice, which lack endogenous IgM/G/A/D necessary for any idiotype‐anti–idiotype interactions, were protected by IVIg against thrombocytopenia to the same degree as normal mice.
A transfection model for weak rhesus D expression
Authors: Hemker M. B. , Berger L. , Van Der Schoot C. E. , Van Rhenen D. J. , Maaskant‐van wijk P. A.
Published: March 2003

Weak D is recently reported to be encoded by 14 different weak D genotypes. The amount of RHD mRNA in donors expressing weak D was measured in previous studies. By semiquantitative RT‐PCR analysis Rouillac et al. (Blood 1996) found lower RHD mRNA levels in donors expressing weak D compared to normal RhD donors, whereas Beckers et al. (Transfusion 1997) described equal mRNA levels.

The aim of the present study is to explore whether the pointmutations found in donors carrying the weak D phenotype can be responsible for weak D expression. Real‐time quantitative PCR analysis showed that levels of RHD mRNA from donors expressing different weak D genotypes (type 1, 2 and 3) did not differ from RHD mRNA levels from normal heterozygous RhD donors. This indicates that the mutations found in weak D genotypes do not affect RhD expression on transcription level.

To investigate whether the described mutations influence translation of mRNA into RhD protein or the configuration of the RhD protein, weak D type 1 cDNA (T809G) and weak D type 3 cDNA (C8G) was transfected in K562 cells. Preliminary results indicate that the ratios between RhD expression and RHD mRNA level (RhD expression/RHD mRNA level) do not differ between weak D type 1 and normal D K562 cells, but are reduced in weak D type 3 transfectants compared to normal D K562 cells. This suggests that in K562 neither the translation nor the configuration of RhD is influenced by the mutation T809G. There might be a missing or limited factor in the K562 model involved in the expression of RhD, hampering the interpretation of the weak D type 1. However, the lower expression of RhD with the mutation C8G might be caused by changed configuration or changed intracellular transport to the membrane.
The changes of cardiac ejection fraction up to 72 h after transfusion in patients with chronic anaemia
Authors: Ho C.‐H.
Published: March 2003

. The objective was to assess the serial change of the cardiac function after blood transfusion in patients with chronic anaemia. According to a previous study, transfusion would adjust the ejection fraction to the advantage of cardiac function in chronic anaemia patients, and thus the serial change of the cardiac function in a certain period should be evaluated after transfusion in patients with chronic anaemia.

In a prospective study, ejection fraction (EF) was determined before and ≤ 6 h, 24 h and 72 h after transfusion in 32 patients with chronic anaemia, and the results were compared and analysed.

In the left ventricle (LV), the four EFs noted before and ≤ 6 h, 24 h and 72 h after transfusion did not significantly differ between one another. In the right ventricle (RV), the EF 72 h after transfusion significantly increased in comparison with the pretransfusion value (P = 0·043).

Transfusion of small amounts of RBC did not significantly change the LVEF up to 72 h in patients with transfusion‐dependent chronic anaemia; whether it might change the RVEF 72 h after transfusion deserves further investigation.
Comparison of two platelet additive solutions
Authors: Van Der Meer P. F. , Pietersz R. N. I. , Reesink H. W.
Published: March 2003

. The use of an additive solution for substitution of plasma for storage of leukodepleted platelet concentrates can have many advantages. In this study, a comparison was made between two platelet additive solutions: one containing citrate and acetate (PAS‐II), the other also supplemented with additional salts such as magnesium, and with gluconate (Composol‐PS). Donor‐dependent differences were avoided by applying a paired experimental design (n = 10). The platelet concentrates were prepared by pooling five buffy coats and the additive solution, and prestorage filtration was utilized to remove leucocytes to well below 1 × 106. Storage of platelet concentrates up to 9 days after blood collection revealed that platelet concentrates in Composol‐PS maintained an almost constant pH of on average 6·93 from day 2 through day 7, and at 6·90 at day 9. This was in contrast to PAS‐II, which showed a gradually decreasing pH from on average 6·97 at day 1 to 6·86 at day 9. In all units stored in both solutions the swirling effect was present during 9 days of storage. In conclusion, both additive solutions allow storage of platelets, derived from pooled buffy coats, for up to 9 days after collection of the whole blood, with maintenance of good quality in vitro. Composol‐PS has a slightly better buffering capacity, reflected as a more constant pH throughout the storage period.
Harvesting of CD34 antigen‐expressing cells with a new programme for the collection of mononuclear cells with use of the Amicus (Baxter) blood cell separator
Authors: Moog R.
Published: March 2003

Summary A study was performed to evaluate a new programme for peripheral blood stem cell (PBSC) collection with the use of the Amicus (Baxter) blood cell separator. Healthy donors (n = 9) and oncology patients (n = 21) scheduled for PBSC transplant were studied. Ten PBSC harvests were performed in the donors and 30 in the patients. A median of 6·37 × 106 CD34+ cells per kg recipient body weight (range 3·08–11·06 × 106) were collected from the donors in a product weight of 169·5 g (118–186). From the patients, 6·26 × 106 CD34+ cells per kg body weight

(range 0·2–53·6 × 106) were harvested in a product weighing 121·5 g (range 92–190). The median platelet contamination was 0·93 × 1011 (range 0·45–1·23 × 1011) per donor product and 0·2 × 1011 (range 0·05–0·86 × 1011) per patient product. No severe side effects were observed during or after the PBSC collection procedures.
Anti‐Ce complicating two consecutive pregnancies with increasing severity of haemolytic disease of the newborn
Authors: Ranasinghe E. , Goodyear E. , Burgess G.
Published: March 2003

Summary. The Ce antigen is expressed on red cells of individuals with Rh haplotypes, R1 or CDe and r′ or Cde. However, anti‐Ce is rare, and only two cases of severe haemolytic disease of the newborn (HDN) caused by this antibody have been described (Malde et al., 2000; Wagner et al., 2000). We describe a woman who was found to have anti‐Ce in her second pregnancy, resulting in a neonate with HDN that required exchange transfusion. She subsequently had a twin pregnancy, where both the twins were affected by severe haemolytic disease (HD) of the fetus because of anti‐Ce and required repeated fetal transfusions, followed by exchange transfusions after birth. This is the first reported case of HD caused by anti‐Ce requiring fetal transfusions.
Survey of autologous blood transfusion activity in England (2001)
Authors: Hill J. , James V.
Published: March 2003

Summary. ‘ Better Blood Transfusion’ (Department of Health (1998) HSC 1998/224) stressed the need for National Health Service Trusts to explore the feasibility of autologous blood transfusions and raise patient awareness of these techniques. The development of an appropriate strategy to promote autologous transfusion requires an understanding of current use, along with constraints to increasing practice. Past surveys have shown unchanging attitudes to autologous transfusion (Torrella et al. (2001) Transfusion Medicine, 11, 15–19), but detailed information regarding the extent of autologous transfusion in England and North Wales was not available.

This survey, carried out in 2001, considered autologous transfusion under four headings: predeposit autologous donation, acute normovolaemic haemodilution, intra‐operative cell salvage and postoperative cell salvage. Questionnaires were returned from 265/310 (85·4%) hospitals served by the National Blood Service.

Sixty per cent of respondents performed at least one autologous technique, although practice was limited to a few clinicians within each hospital. Predeposit donation was the most widespread technique but involved very few patients. Intra‐ and postoperative cell salvage techniques were practised in fewer hospitals but on many more patients. The main constraints to increasing use of autologous techniques were perceived to be logistical, but removal of constraints may only result in small increases in individual practice. More clinicians need to be encouraged to utilize appropriate, evidence‐based autologous techniques.
Quantitative assessment of circulating antigens in human lymphatic filariasis: a field evaluation of monoclonal antibody‐based ELISA using blood collected on filter strips
Authors: Lalitha P. , Ravichandran M. , Suba S. , Kaliraj P. , Narayanan R. B. , Jayaraman Kunthala
Published: March 2003

objective
 
 To quantify circulating antigens in individuals with lymphatic filariasis by means of an ELISA using blood on filter strips.method  Circulating antigens in filarial patients and normal individuals living in an area endemic for W. bancrofti infection in Madras, India were estimated using a monoclonal antibody‐based ELISA. results  All microfilaraemics showed positivity to circulating antigens whereas people with chronic pathology and 80% of the endemic normals tested negative. The antigen levels in the blood collected in the night and during day time showed positivity and there was no difference in the antigen concentration. The results of the antigen levels collected onto filter strips correlated with their corresponding plasma antigen levels (r= 0.83). In microfilaraemics, DEC treatment did not alter the levels of circulating antigens for up to one month. conclusions  We conclude that this monoclonal antibody‐based ELISA using filter strips may be used in day time and replace the existing routine night blood surveys in our endemic area in India.
Distinction of recrudescences from new infections by pcr‐rflp analysis in a comparative trial of cgp 56 697 and chloroquine in Tanzanian children.
Authors: Irion Andrea , Felger Ingrid , Abdulla Salim , Smith Tom , Mull Robert , Tanner Marcel , Christoph Hatz  , Beck Hans‐Peter
Published: March 2003

Summary
objective To test the efficacy of a new compound drug (CGP 56 697) against acute, uncomplicated falciparum malaria.

method Reappearing parasites were analysed by PCR‐RFLP within a randomized controlled trial. 130 patients received chloroquine and 130 patients were treated with CGP 56 697. Samples from 96 patients with parasitological failure were tested by PCR‐RFLP for MSP2 of Plasmodium falciparum. Seven days after treatment 32 patients of the chloroquine control group with reappearing parasites were tested by PCR and one infection was unequivocally determined as a new infection. After 7 days, in the CGP 56 697 group, 6 samples were tested in which one new infection was identified. Similar observations were made one and three weeks later in both groups.

results Although a high multiplicity of infections on admission was observed, there was no significant correlation between multiplicity and either recrudescence or new infection. Patients in both treatment groups with subsequent recrudescent parasites had higher initial mean parasite densities than patients who cleared. Those of the patients with recrudescent parasites who were treated with CGP 56 697 had higher initial parasite densities than those treated with chloroquine. The rate of re‐infection increased with time as expected in holoendemic areas and appeared to be higher in chloroquine patients. Generally, CGP 56 697 showed a superior clearance rate, successfully cleared higher parasite densities and suppressed new infections over a longer period of time.

conclusion The PCR analysis confirmed that reinfections beyond day 7 are significant in areas highly endemic for malaria and showed the necessity of excluding these when estimating 14 day clearance rates. Provided new infections are excluded, the 28‐day clearance rate can also be used to determine the efficacy of antimalarial drugs in highly endemic areas, and adds to our knowledge of drug resistance and dynamics of infections in people living in such areas.
Rapid risk household screening by neonatal arm circumference: results from a cohort study in rural Burkina Faso
Authors: Benzler Justus , Sauerborn Rainer
Published: March 2003

Neonatal arm circumference (NAC) and other attributes of the newborn and its household were analysed as potential predictors of child death in a cohort of 1367 newborn children representing the majority of births in a rural area of Burkina Faso from 1992 to 1994. During 3872 person years observed 264 children died, resulting in an average mortality rate of 6.8% per year. 90 mm was chosen as the best cut‐off to differentiate low NAC associated with high mortality from normal NAC. The hazard ratio of children with low NAC (15.7%) compared to others was 1.7 (P < 0.001) in Cox regression. Kaplan‐Meier curves of cumulative survival showed that this higher risk lasted throughout the first two years of life. Multivariate Cox regression comparing NAC with other variables known or suspected to influence child survival yielded a model including mother's death, twin birth, affiliation to a particular health centre, home delivery and birth during the rainy or harvest season as other significant risk factors beside NAC. Protective factors were mother's participation in antenatal care despite considerable distance to the health centre, medium household size (5–7 members) and household cash crop production. We propose a simple risk score for rapid household screening in rural Burkina Faso and comparable settings elsewhere for identifying households at risk of experiencing child death. As much of the other variables’ contribution to the explanation of survival pattern is absorbed by NAC in more parsimonious models, even simpler screening strategies based on NAC make sense. In the study area risk households will be offered periodical home visits by the local nurse promoting immunization, treatment of illness and strengthening the mothers’ competence to recognize and manage frequent health problems of their children as part of a ‘Shared Care’ concept.
Multi‐centre evaluation of repeatability and reproducibility of the direct agglutination test for visceral leishmaniasis
Authors: Boelaert M. , El Safi S. , Mousa H. , Mbati J. , Gurubacharya V. L. , Shrestha J. , Jacquet D. , De Muynck A. , Le Ray D. , Boelaert M. , Van der Stuyft P.
Published: March 2003

Summary objective  To evaluate the repeatability and reproducibility of the serological direct agglutination test (DAT) for visceral leishmaniasis (VL) with aqueous antigen in a multi‐centre study in VL‐endemic areas in Sudan, Kenya and Nepal.

methods  Repeatability within each centre and reproducibility between the centres' results and an external reference laboratory (Belgium) was assessed on 1596 triplicate plain blood samples collected on filter paper.

results  High kappa values (range 0.86–0.97) indicated excellent DAT repeatability within the centres. The means of the titre differences between the reference laboratory and the centres in Sudan, Kenya and Nepal (2.3, 2.4 and 1.1, respectively, all significantly different from 0) showed weak reproducibility across centres. 95% of the titre differences between the reference laboratory and the respective centres were accounted for by large intervals: 0.6–9 fold titre variation for Sudan, 0.7–8 fold for Kenya and 0.26–4 fold for Nepal.

conclusion  High repeatability of DAT confirms its potential, but reproducibility problems remain an obstacle to its routine use in the field. Reproducibility was hindered by alteration of the antigen through temperature and shaking, especially in Kenya and Sudan, and by nonstandardization of the test reading. DAT handling procedures and antigen quality must be carefully standardized and monitored when introducing this test into routine practice.
Anti‐filarial IgG4 in men and women living in Brugia malayi‐endemic areas
Authors: Haarbrink Melang , Terhell Annemarie J. , Abadi Kunar , Asri Muhaammad , De Medeiros Filomena , Yazdanbakhsh Maria
Published: March 2003

Summary To assess whether antifilarial IgG4 can be used to study various epidemiological facets of filarial infections, we studied this isotype in 238 individuals resident in areas endemic for brugian filariasis, focusing on the differences between men and women. In the study area, the prevalence of microfilariae was 6.7% and the prevalence of antifilarial IgG4 was 49.2%. All microfilariae carriers were positive for antifilarial IgG4, whereas a proportion of the endemic normals (94/208) and elephantiasis patients (7/14) had IgG4 antibodies to filarial antigens. Data were analysed as a function of gender in distinct clinical groups and stratified for age. The prevalence of microfilariae was higher in males in all age groups, as reflected in significantly higher antifilarial IgG4 antibody levels compared to females. The prevalence of IgG4 increased to reach a plateau at the age of 30 years in both males and females. These results indicate that antifilarial IgG4 antibodies can reflect the differences in the extent of infection in males and females as measured by microfilarial counts, and that this parameter can be used for epidemiological assessments of filarial infection.
Identification of two species within the Anopheles minimus complex in northern Vietnam and their behavioural divergences
Authors: Bortel W. , Trung H. D. , Manh N. D. , Roelants P. , Verlé P. , Coosemans M.
Published: March 2003

Summary Elucidating the complex taxonomic status of the major malaria vector taxa and characterising the individual species within each complex is important for understanding the complexity of the vector system in the south‐east Asian region and will allow to estimate the impact of vector control measures. This applies to countries such as Laos, Cambodia and Vietnam that spend about 60% of their malaria control budget on implementing vector control activities. We used isozyme electrophoresis to clarify the Anopheles minimus s.l. species composition in northern Vietnam and identify behavioural divergences of individual species. Using different collection methods, adult mosquitoes were caught at monthly intervals from June to November 1995 in four villages. An. minimus s.l. could be distinguished from closely related species, An. aconitus and An. jeyporiensis, at the Octanol dehydrogenase (Odh) enzyme locus. Significant positive Fis values gave clear evidence of nonrandom mating within the An. minimus s.l. population. The highest heterozygote deficiency was observed at locus Odh, which was diagnostic for 2 sympatric An. minimus species in Vietnam similar to the An. minimus A and C species known from Thailand. We found no evidence for restricted gene flow between monthly samples, villages, or collection methods in either of the two An. minimus species. They occurred in sympatry, but in different proportions depending on the collection site, and had dissimilar resting and biting behaviours. Thus a vector control strategy will have a nonuniform effect on the various components of this diverse vector system.
Case report: French West Indies – a tourist destination at risk for Plasmodium falciparum transmission?
Authors: Poinsignon Yves , Arfi Catherine , Sarfati Claudine , Farge‐Bancel Dominique , Raccurt Christian P.
Published: March 2003

Summary We report a case of P. falciparum infection observed in Paris and presumably acquired in Guadeloupe, a French Caribbean island where malaria has been considered to be eradicated since 1970.