Dirk de Korte | Sanquin Bloodsupply (original) (raw)

Papers by Dirk de Korte

Research paper thumbnail of Composition of the additive solution affects red blood cell integrity after photodynamic treatment

Vox Sanguinis, 2003

Photodynamic treatment is a promising technique for pathogen inactivation of red blood cell conce... more Photodynamic treatment is a promising technique for pathogen inactivation of red blood cell concentrates. For protocol optimization, the influence of the composition of the storage solution on the integrity of phototreated red cells was studied. Red blood cells were resuspended in the storage solutions SAG-M or AS-3 to a haematocrit (Hct) of 30%. After addition of the photosensitizer, 1,9-dimethylmethylene blue (DMMB) (25 microm), the suspensions were illuminated with red light, and potassium leakage and delayed haemolysis were determined. In some experiments, the cells were washed after illumination and resuspended in modified storage solutions. Illumination of red cells in the presence of DMMB resulted in an immediate, light-dose-dependent increase in potassium leakage. The illumination conditions used induced no detectable haemolysis immediately after photodynamic treatment. Potassium leakage was higher when the illumination was performed in AS-3. In contrast, delayed haemolysis, measured after overnight storage, was considerably lower when cells were stored in AS-3. This protection was mainly a result of the presence of citrate in AS-3. In addition, other impermeant solutes protected against haemolysis. The additive solution strongly influences the integrity of red cells after photodynamic treatment. Whereas the solution in which the cells are illuminated has a small effect on red cell integrity, the main influence of the additive solution is during post-treatment storage. Red cell integrity is best maintained when illumination is performed in SAG-M followed by storage in AS-3. The presence of non-permeant solutes, such as citrate, in the solution used for storage, prevents haemolysis of the phototreated, cation-permeable cells by counterbalancing the osmotic activity of haemoglobin.

Research paper thumbnail of The age of stored red blood cell concentrates at the time of transfusion

Transfusion Medicine, 2005

Research paper thumbnail of The band III ligand dipyridamole protects human RBCs during photodynamic treatment while extracellular virus inactivation is not affected

Transfusion, 2002

BACKGROUND: Recently, the potential usefulness of dipyridamole (DIP) in protecting RBCs against t... more BACKGROUND: Recently, the potential usefulness of dipyridamole (DIP) in protecting RBCs against the harmful side effects of photodynamic sterilization was demonstrated. In the present study, the use of DIP for selective protection of RBCs was investigated under conditions more relevant for blood bank practice. STUDY DESIGN AND METHODS: WBC-reduced RBC suspensions (30% Hct) were treated with 1,9-dimethylmethylene blue and red light, and the influence of the inclusion of DIP on photohemolysis was assessed as a function of sensitizer concentration, light dose, and storage time. Furthermore, the possible interference of DIP with inactivation of extracellular virus by use of a panel of different viruses (HIV-1, pseudorabies virus [PRV], bovine viral diarrhea virus [BVDV], VSV, encephalomyocarditis, and canine parvovirus) was investigated. RESULTS: In WBC-reduced RBC suspensions (30% Hct), DIP exerted a clear protective effect against photohemolysis. Part of this protection was achieved with concentrations near the dissociation constant for band III binding. Importantly, efficiency of inactivation of extracellular HIV-1, PRV, BVDV, and VSV was not significantly impaired by the inclusion of DIP. Phototreatment conditions, resulting in a 4 to 5 log inactivation of extracellular HIV-1 and PRV, resulted in a high level of hemolysis after 28 days of storage. This long-term hemolysis could be decreased, but not completely prevented, by the inclusion of DIP. CONCLUSION: Photohemolysis in RBC concentrates can be reduced substantially by the application of DIP, while the efficacy of inactivation of HIV-1 and other viruses remains unchanged.

Research paper thumbnail of Should DEHP be eliminated in blood bags?

Vox Sanguinis, 2013

Whole blood for transfusion was initially collected in glass bottles, but these are fragile, heav... more Whole blood for transfusion was initially collected in glass bottles, but these are fragile, heavy to transport and prone to bacterial contamination. After the Second World War, Carl Walter experimented with plastics for collection and storage of blood, and found that di-ethylhexyl phthalate (DEHP) as plasticizer for polyvinyl chloride (PVC) had the most favourable properties [1, 2]. The breakage rate of frozen plasma in DEHP PVC is low, and the better storage properties of red cell concentrates in DEHP PVC are explained by leaching of plasticizer into the lipid bilayer of these cells, thereby improving membrane stability resulting in lower haemolysis rates [3]. However, ever since the 1970s, scientific and public concern has been raised on the toxicity of DEHP for transfused patients [4, 5]. DEHP and its metabolite mono ethylhexyl phthalate (MEHP) are associated with impairment of reproduction in animal models, resulting in testicular dysgenesis syndrome in (male) rodents, as reviewed in depth elsewhere [6], and particularly, neonates are susceptible to the risks of high exposure to DEHP. Due to the omnipresence of phthalates (not only DEHP), and some important differences in DEHP metabolism between animal models and humans [6], it is difficult to estimate the effect of DEHP toxicity in humans. A study of 234 young men showed no effect of MEHP in urine on their reproductive function [7], but others showed that DNA damage in sperm was associated with the MEHP concentration (after adjustment for other DEHP oxidative metabolites) [8]. In vitro human testis explants incubated with DEHP or MEHP showed that both significantly inhibited testosterone production [9], and a study of 881 men showed a 9% lower free androgen index between the lowest and the highest quartile of the proportion DEHP excreted as MEHP (P = 0.02) [10]. Also, an association was found between urinary MEHP concentrations and lower free T4 and lower total T3 thyroid hormones in adult men [11]. In addition to these effects on hormonal level in adult males, DEHP and its metabolites can have an effect on fetal development, with a shorter anogenital distance associated with higher metabolite concentrations in urine samples collected during pregnancy [12]. Others found no difference in T4 and testosterone levels, as well as no effect on phallic length in 13 male adolescents 14– 16 years of age that had underwent ECMO treatment in their neonatal period, as compared to ageand sexmatched controls [13]. Also, DEHP/MEHP exposure (as detected in cord blood) was significantly associated with a shorter pregnancy duration [14]. Despite these associations, they should be interpreted with care. DEHP is not the only phthalate that may cause these effects; studies that showed no association may not have been published; DEHP administered through the intestinal tract is converted to MEHP in a higher degree as by intravenously [15]; and because phthalates are everywhere in our environment, comparisons are less versus more, rather than absent versus present. DEHP-plasticized PVC has long been used as standard material for a wide range of applications, like packing foil (including food), building material, toys and plastic devices. DEHP is considered to be a ubiquitous environmental contaminant and is present in air, dust, water, soil and food. The latter is considered to be the main source of DEHP intake for the general population. The current daily intake is estimated to be between 2 and 5 lg/kg for adults and between 4 to 8 lg/kg for children [16, 17]. Various measures to reduce the use of DEHP-plasticized PVC have indeed resulted in a 2-times lower exposure to DEHP in the general population over the last two decades [18]. DEHP is applied in many medical devices, and depending on the procedure, patients can be exposed to high (peak) concentrations of DEHP, in the order of 1 mg per kg of bodyweight. One of the sources of DEHP exposure is the leaching from the blood bags into the content of the container, particularly into lipophilic solutions such as plasma [19], but also red cells. Therefore some patient groups, including pregnant or nursing women and children, are considered to be most at risk of possible harmful effects attributable to DEHP exposure by medical treatments. DEHP is broken down into various metabolites. First, it is hydrolysed to MEHP and then further oxidized to mono-(2-ethyl-5-hydroxyhexyl) phthalate (5OH-MEHP),

Research paper thumbnail of Storage of Leukocyte-Poor Red Cell Concentrates: Filtration in a Closed System Using a Sterile Connection Device

Vox Sanguinis, 1989

Storage of leukocyte-poor red cell concentrates (LP-RCC) was investigated after filtration in a c... more Storage of leukocyte-poor red cell concentrates (LP-RCC) was investigated after filtration in a closed system that was assembled using a Sterile Connection Device (SCD). The LP-RCC were stored for up to 6 weeks following filtration with either 0.9% saline solution (n = 14) or saline-adenine-glucose-mannitol (SAG M) solution (n = 15) to prime and rinse the cellulose acetate filter. The results were compared with the data of nonfiltered buffy-coat-poor red cell concentrates (BC-poor RCC) stored in SAG M solution (n = 10). All LP-RCC contained less than lo6 leukocytes whereas the nonfiltered BC-poor RCC contained 675+286 x lo6 leukocytes at day 1, decreasing to 83+49 X lo6 at day 42. Although glucose consumption, lactic acid production and decrease in pH was similar from day 7 through 28 in both groups of LP-RCC, a significantly steeper decline of ATP values as well as a higher hemolysis and LDH release was observed in the LP-RCC filtered with saline. During storage of the nonfiltered BC-poor RCC in SAG M, significantly higher glucose consumption (p<O.Ol), LDH release (p<O.OOl), rate of hemolysis (p<O.OOl) and a lower pH (p<O.OOl) were found, compared to the filtered units. It is postulated that the leukocytes present in the nonfiltered BC-poor RCC were responsible for these differences. The ATP values in the SAG-M-filtered and nonfiltered BC-poor RCC in SAG M were comparable. By comparing the ATP levels and values of the filtered RCC and the nonfiltered BC-poor RCC we conclude that the LP-RCC can be stored for 35 days if SAG M solution is used to prime and rinse the filter.

Research paper thumbnail of A novel flow cytometry–based platelet aggregation assay

Blood, 2013

Key PointsFCA is a novel flow cytometry–based platelet aggregation assay that allows single recep... more Key PointsFCA is a novel flow cytometry–based platelet aggregation assay that allows single receptor analysis in small volume/thrombocytopenic samples FCA facilitates platelet studies in experimental animal models even during gestation and allows kinetic measurements in individual animals

Research paper thumbnail of Inhalt, Vol. 18, Suplement 1, 1991

Transfusion Medicine and Hemotherapy, 1991

Research paper thumbnail of Clinical and in vitro evaluation of red blood cells collected and stored in a non‐DEHP plasticized bag system

Research paper thumbnail of Small but clear pro-hemostatic effects of blood donation-induced stress on donor’s hemostasis

Background: Donating blood induces stress responses in donors, e.g. increased anxiety and blood p... more Background: Donating blood induces stress responses in donors, e.g. increased anxiety and blood pressure. Whereas acute stress in a non-donation setting is associated with pro-hemostatic effects, such effects in blood donors have not been investigated, despite potential importance for the quality of blood products. Therefore, this study examines whether donation-induced stress has an effect on hemostatic parameters in whole-blood donors. Study design and methods: In 372 healthy whole-blood donors, three sets of stress-related parameters were assessed at needle insertion: 1) psychological stress (i.e. donation-stress and arousal on a visual analogue scale); 2) hormonal stress, (i.e. cortisol in saliva); and 3) physiological stress (i.e. blood pressure, pulse rate, and pulse rate variability). Hemostatic parameters (PT, aPTT, fibrinogen, factor VII, factor VIII, and von Willebrand factor (vWF)) were assessed using a 5-ml citrated blood sample taken from the diversion pouch directly af...

Research paper thumbnail of Quality of Platelets in Stored Whole Blood

Transfusion Medicine Reviews, 2020

Transfusion of whole blood rather than blood components is gaining popularity. It is easy to use,... more Transfusion of whole blood rather than blood components is gaining popularity. It is easy to use, with one transfusion product to administer rather than 3, and is held at one storage temperature. It only contains anticoagulant-preservative solution, while components contain various storage solutions, which in theory may induce dilution coagulopathy. In this review, the quality of platelets in stored whole blood is summarized. In cold-stored whole blood, the platelet count declines by 1% to 2% per day. The responsiveness to various agonists declines during the storage time, but this appears to have a limited impact on clotting time or on clot strength as measured with thromboelastography. Animal studies have confirmed that platelets from stored whole blood participate equally well in clot formation. The recovery of platelets in stored whole blood is acceptable during at least 15 days of storage. The survival of platelets after transfusion is only 1 day, but this is likely to be sufficient for the intended patient group requiring massive transfusions, as the platelets are rapidly consumed in the wound area. In addition to the logistic benefits, there are drawbacks, most importantly having a sufficiently large inventory with an acceptable outdating rate, particularly since massive transfusions are rare, while requiring a lot of whole blood. The positive experience of the United States military with whole blood transfusion is often brought forward for introduction in the civilian blood bank, but patients with trauma are only a small fraction of the civilian population requiring massive transfusions. It needs to be determined whether in the resourceful environment of the hospital, these patients benefit from whole blood transfusions. Optimization of whole blood storage, with focus on platelet quality, needs to be performed to allow extension of the storage time beyond 15 days to a point where the number of units in inventory and outdating can be balanced.

Research paper thumbnail of The endosomal RIN2/Rab5C machinery prevents VEGFR2 degradation to control gene expression and tip cell identity during angiogenesis

Angiogenesis, 2021

Sprouting angiogenesis is key to many pathophysiological conditions, and is strongly regulated by... more Sprouting angiogenesis is key to many pathophysiological conditions, and is strongly regulated by vascular endothelial growth factor (VEGF) signaling through VEGF receptor 2 (VEGFR2). Here we report that the early endosomal GTPase Rab5C and its activator RIN2 prevent lysosomal routing and degradation of VEGF-bound, internalized VEGFR2 in human endothelial cells. Stabilization of endosomal VEGFR2 levels by RIN2/Rab5C is crucial for VEGF signaling through the ERK and PI3-K pathways, the expression of immediate VEGF target genes, as well as specification of angiogenic ‘tip’ and ‘stalk’ cell phenotypes and cell sprouting. Using overexpression of Rab mutants, knockdown and CRISPR/Cas9-mediated gene editing, and live-cell imaging in zebrafish, we further show that endosomal stabilization of VEGFR2 levels is required for developmental angiogenesis in vivo. In contrast, the premature degradation of internalized VEGFR2 disrupts VEGF signaling, gene expression, and tip cell formation and migr...

Research paper thumbnail of Metabolic effect of alkaline additives and guanosine/gluconate in storage solutions for red blood cells

Transfusion, 2018

Background-Over a century of advancements in the field of additive solutions for red blood cell s... more Background-Over a century of advancements in the field of additive solutions for red blood cell storage has made transfusion therapy a safe and effective practice for millions of recipients worldwide. Still, storage in the blood bank results in the progressive accumulation of metabolic alterations, a phenomenon that is mitigated by storage in novel storage additives, such as alkaline additive solutions. While novel alkaline additive formulations have been proposed, no metabolomics characterization has been performed to date. Study design and Methods-We performed UHPLC-MS metabolomics analyses of red blood cells stored in SAGM (standard additive in Europe), PAGGSM or alkaline additives SOLX, ESOL-5 and PAG3M for either 1, 21, 35 (end of shelf-life in The Netherlands) and 56 days. Results-Akaline additives (especially PAG3M) better preserved 2,3-diphosphoglycerate and adenosine triphosphate (ATP). Deaminated purines such as hypoxanthine were predictive of hemolysis and morphological alterations. Guanosine supplementation in PAGGSM and PAG3M fueled ATP generation by feeding into the non-oxidative pentose phosphate pathway via phosphoribolysis. Decreased urate to hypoxanthine ratios were observed in alkaline additives, suggestive of decreased generation of urate and hydrogen peroxide. Despite the many benefits observed in purine and redox metabolism, alkaline additives did not prevent accumulation of free fatty acids and oxidized byproducts, opening a window for future alkaline formulations including (lipophilic) antioxidants.

Research paper thumbnail of In vivo labelling of intermediates in the discontinuous synthesis of mRNAs in Trypanosoma brucei

The EMBO Journal, 1987

Discontinuous mRNA synthesis in trypanosomes is thought to involve a 140-nucleotide precursor, ca... more Discontinuous mRNA synthesis in trypanosomes is thought to involve a 140-nucleotide precursor, called the mini-exonderived RNA or medRNA, which contributes its 5' 35 nucleotides to the 5' end of nascent mRNAs. We used in vivo labelling of RNA to show that medRNA has a half-life of <6 min, whereas putative high mol. wt intermediates containing the 3' part of the medRNA have an average half-life of < 1 min. This eliminates priming of pre-mRNA synthesis by intact medRNA as the main mode of discontinuous mRNA synthesis. Potential intermediates of 35 and 105 nucleotides were labelled in parallel with medRNA, but their significance could not be assessed in RNA preparations containing med-RNA, as they are also produced by artefactual cleavage of medRNA. We show, however, that high mol. wt RNA, free of medRNA, can release medRNA segments upon a debranching treatment. These results are consistent with a trans splicing mechanism involving short-lived forked intermediates, analogous to lariats in cis splicing systems. Key words: discontinuous transcription/in vivo labelling/RNA turnover/trans splicing/trypanosomes 1984; Laird et al., 1985; Lenardo et al., 1985a). This mini-exonderived RNA (medRNA) is thought to transfer its mini-exon sequence to the 5' end of pre-mRNAs, but how is unknown. As shown in the schematic diagrams in Figure 1, the medRNA could prime transcription upstream of the main exons or, alternatively, the linkage could occur post-transcriptionally by RNA-RNA ligation or by trans splicing between the medRNA and main exon precursor transcripts. The intermediates are depicted by analogy with nuclear mRNA precursor splicing systems of other eukaryotes (see Padgett et al., 1986). It is likely that a splicing event is responsible for the final joining of the two RNA segments, since the mini-exon sequence is flanked at the 3' end by GT (De Lange et al., 1983) and all main exon sequences are preceded by AG (see Borst, 1986), in compliance with the conventional splice site consensus sequences (Mount, 1982). The pyrimidine-rich stretch upstream of splice acceptor sites in other eukaryotes is less prominent in trypanosomes, while a conserved sequence TTTCPy is found almost invariably in one or more copies within 30 bp upstream of trypanosome splice acceptor sites (P.W.Laird, unpublished results). No pre-mRNA transcription unit has been fully defined yet, but, in the few cases studied, these units appear to span several protein-coding regions, which lack conventional introns (see Borst, 1986). Studies of the mechanism of discontinuous mRNA synthesis have been hampered by the limited available methodology. We have used in vivo labelling of RNA to study the turnover of med-RNA, to track the fate of its 5' and 3' segments and to identify possible intermediates.

Research paper thumbnail of The Platelet Adhesion Capacity to Subendothelial Matrix and Collagen in a Flow Model during Storage of Platelet Concentrates for 7 Days

Thrombosis and Haemostasis, 1994

SummaryThe influence of storage of platelet concentrates (PC) on the adhesion capacity of platele... more SummaryThe influence of storage of platelet concentrates (PC) on the adhesion capacity of platelets was studied. Twenty-four PC, 12 prepared by the buffy coat (BC) method and 12 by the platelet-rich plasma (PRP) method, were stored for 7 days at room temperature. On days 1,3 and 7 of storage, the platelet adhesion capacity to subendothelial matrix (SEM) and collagen was studied in a rectangular perfusion system under flow conditions in conjunction with the platelet aggregation capacity after stimulation and the adenine nucleotide content. The platelet adhesion capacity to collagen was constant until day 3 of storage and decreased to about 80% of the starting value on day 7 of storage. The adhesion capacity to SEM, however, had already decreased on day 3 to about 75% of the value of day 1 and was even more decreased on day 7 to about 45% of the starting value. On day 1, platelets prepared by the BC method displayed a higher adhesion capacity to collagen and a higher aggregation capac...

Research paper thumbnail of Apheresis causes complement deposition on red blood cells (RBCs) and RBC antigen alterations, possibly inducing enhanced clearance

Research paper thumbnail of A comprehensive proteomics study on platelet concentrates: Platelet proteome, storage time and Mirasol pathogen reduction technology

Platelets, Jan 19, 2018

Platelet concentrates (PCs) represent a blood transfusion product with a major concern for safety... more Platelet concentrates (PCs) represent a blood transfusion product with a major concern for safety as their storage temperature (20-24°C) allows bacterial growth, and their maximum storage time period (less than a week) precludes complete microbiological testing. Pathogen inactivation technologies (PITs) provide an additional layer of safety to the blood transfusion products from known and unknown pathogens such as bacteria, viruses, and parasites. In this context, PITs, such as Mirasol Pathogen Reduction Technology (PRT), have been developed and are implemented in many countries. However, several studies have shown in vitro that Mirasol PRT induces a certain level of platelet shape change, hyperactivation, basal degranulation, and increased oxidative damage during storage. It has been suggested that Mirasol PRT might accelerate what has been described as the platelet storage lesion (PSL), but supportive molecular signatures have not been obtained. We aimed at dissecting the influenc...

Research paper thumbnail of Vox Sanguinis International Forum on platelet cryopreservation: Summary

Vox Sanguinis, 2017

The demand for platelets continues to rise, outstripping supply worldwide. In higher-income count... more The demand for platelets continues to rise, outstripping supply worldwide. In higher-income countries, cancer patients remain platelet transfusion dependent for extended periods, which leads to sporadic and chronic regional shortages that are secondary to uncontrolled fluctuations in supply. In less-developed countries, the supply to demand ratio is critical for obstetric haemorrhage, trauma and medical patients. All countries, however, share the difficulty in providing platelets to rural areas or in support of troops in remote military theatres. Alternatives to conventional 5-to 7-day room temperature-stored platelets are needed. Cryopreservation of platelets is likely the best alternative to balance the supply and demand for platelets in cities and less accessible areas. Cryopreserved platelet units (CPP) can be shipped on dry ice and stored for 2 years or more in mechanical freezers or using liquid nitrogen [1]. This controlled supply may be used as a backup to liquid units in well-stocked hospitals, or as the sole source of platelets in remote or underserved settings. Cryopreserved units can be stockpiled for natural disasters, or special HLA-or HPA-selected units may be stored for patients who are refractory to platelet transfusions due to antibodies. In addition, cryopreservation reduces the risk of septic transfusion reactions, as bacteria will not proliferate under these conditions. Cryopreservation of platelets has been available since the 1970s, when Valeri published his original method for storing platelets in 6% dimethyl sulphoxide (DMSO) at-80°C [2]. Subsequent work led to an improved 'no-wash' method, which removed the DMSO prior to freezing, thus allowing for easier post-thaw utilization [1]. While other methods for cryopreservation have been published [3, 4], most data on CPPs in vitro characteristics, storage duration, autologous survival/recovery, clinical safety and efficacy have been developed using the 'no-wash' approach. Extensive in vitro characterization has been performed using CPP resuspended in plasma, saline or platelet additive solutions. These studies have shown that post-thaw platelets, when compared to liquid platelets, are partially activated, contain a distinct microparticle population and

Research paper thumbnail of Transfusion of 35-Day Stored RBCs in the Presence of Endotoxemia Does Not Result in Lung Injury in Humans

Critical care medicine, Jan 2, 2016

Transfusion-related acute lung injury is the leading cause of transfusion-related mortality. Prec... more Transfusion-related acute lung injury is the leading cause of transfusion-related mortality. Preclinical studies have shown that aged RBCs can induce transfusion-related acute lung injury in the presence of a "first hit" (e.g., sepsis). Clinical studies, however, show conflicting results on this matter. We tested whether maximally stored RBCs are able to induce lung injury in the presence of a "first hit" in humans (Dutch Trial Register: NTR4455). Open-label, randomized controlled trial. Healthy male volunteers. Eighteen healthy male volunteers donated one unit of autologous RBCs 2 or 35 days before the experiment. The experiment was started by infusion of 2 ng/kg lipopolysaccharide ("first hit"). After 2 hours, volunteers received normal saline (n = 6), 2-day stored transfusion (n = 6), or 35-day stored transfusion (n = 6) ("second hit"). Blood was sampled hourly. Six hours after transfusion, the diffusion capacity of the lungs for carbon mon...

Research paper thumbnail of Adhesion of anaerobic bacteria to platelet containers

Vox Sanguinis, 2014

Propionibacterium acnes and Staphylococcus saccharolyticus are frequently isolated during platele... more Propionibacterium acnes and Staphylococcus saccharolyticus are frequently isolated during platelet screening with anaerobic culture methods. Although neither P. acnes nor S. saccharolyticus proliferates during platelet storage, both species survive well in this environment. This study was aimed at determining whether strains of P. acnes and/or S. saccharolyticus form surfaceattached bacterial cell aggregates, known as biofilms, under platelet storage conditions. We report that these organisms are able to adhere to the inner surface of platelet containers in tight interaction with activated platelets.

Research paper thumbnail of Comparison of various dimethylsulphoxide-containing solutions for cryopreservation of leucoreduced platelet concentrates

Vox Sanguinis, 2003

Leucoreduced platelet concentrates (LR-PCs) can be stored at 20-24 degrees C for 5-7 days. When L... more Leucoreduced platelet concentrates (LR-PCs) can be stored at 20-24 degrees C for 5-7 days. When LR-PCs are cryopreserved they can be stored for several years. For cryopreservation to become applicable in blood-bank practice, an off-the-shelf cryoprotectant is needed that can be added to the LR-PC in a sterile manner. For this, we varied the composition of the cryopreservation medium and studied various parameters of cryopreserved LR-PCs for up to 24 h after thawing at room temperature. LR-PCs in plasma or Composol were concentrated and divided into 2 units. To each unit, an equal part of 10% dimethylsulfoxide (DMSO) in plasma, Composol with or without 5% albumin, or GPO (pasteurized plasma-protein solution) was added. Freezing occurred at 1 degrees C/min and LR-PCs were placed in the vapour phase of nitrogen. LR-PCs were thawed at 37 degrees C and stored at room temperature. LR-PCs were tested for morphology, platelet recovery, swirling effect, and activation antigens at various time-points thereafter. LR-PCs in 100%, 65% and 50% plasma supplemented with Composol showed good morphology scores (&amp;amp;amp;amp;gt;250), limited CD62P expression (&amp;amp;amp;amp;lt;35%), low CD63 expression (&amp;amp;amp;amp;lt;20%) and a swirling effect of about 2, at 24 h after thawing. At the same time-point, platelet recovery was &amp;amp;amp;amp;gt;80% under all conditions and CD42b expression varied between 70 and 85%. Results of LR-PCs in 15% plasma and Composol, with or without plasma substitutes, were not acceptable at 24 h after thawing, i.e. the morphology score was &amp;amp;amp;amp;lt;200 and the CD62P expression was &amp;amp;amp;amp;gt;40%. A minimum of 50% plasma in the cryopreserved LR-PC is necessary to maintain an acceptable in vitro quality of platelets up to 24 h after thawing. Composol is a good candidate for using to prepare an off-the-shelf cryoprotectant.

Research paper thumbnail of Composition of the additive solution affects red blood cell integrity after photodynamic treatment

Vox Sanguinis, 2003

Photodynamic treatment is a promising technique for pathogen inactivation of red blood cell conce... more Photodynamic treatment is a promising technique for pathogen inactivation of red blood cell concentrates. For protocol optimization, the influence of the composition of the storage solution on the integrity of phototreated red cells was studied. Red blood cells were resuspended in the storage solutions SAG-M or AS-3 to a haematocrit (Hct) of 30%. After addition of the photosensitizer, 1,9-dimethylmethylene blue (DMMB) (25 microm), the suspensions were illuminated with red light, and potassium leakage and delayed haemolysis were determined. In some experiments, the cells were washed after illumination and resuspended in modified storage solutions. Illumination of red cells in the presence of DMMB resulted in an immediate, light-dose-dependent increase in potassium leakage. The illumination conditions used induced no detectable haemolysis immediately after photodynamic treatment. Potassium leakage was higher when the illumination was performed in AS-3. In contrast, delayed haemolysis, measured after overnight storage, was considerably lower when cells were stored in AS-3. This protection was mainly a result of the presence of citrate in AS-3. In addition, other impermeant solutes protected against haemolysis. The additive solution strongly influences the integrity of red cells after photodynamic treatment. Whereas the solution in which the cells are illuminated has a small effect on red cell integrity, the main influence of the additive solution is during post-treatment storage. Red cell integrity is best maintained when illumination is performed in SAG-M followed by storage in AS-3. The presence of non-permeant solutes, such as citrate, in the solution used for storage, prevents haemolysis of the phototreated, cation-permeable cells by counterbalancing the osmotic activity of haemoglobin.

Research paper thumbnail of The age of stored red blood cell concentrates at the time of transfusion

Transfusion Medicine, 2005

Research paper thumbnail of The band III ligand dipyridamole protects human RBCs during photodynamic treatment while extracellular virus inactivation is not affected

Transfusion, 2002

BACKGROUND: Recently, the potential usefulness of dipyridamole (DIP) in protecting RBCs against t... more BACKGROUND: Recently, the potential usefulness of dipyridamole (DIP) in protecting RBCs against the harmful side effects of photodynamic sterilization was demonstrated. In the present study, the use of DIP for selective protection of RBCs was investigated under conditions more relevant for blood bank practice. STUDY DESIGN AND METHODS: WBC-reduced RBC suspensions (30% Hct) were treated with 1,9-dimethylmethylene blue and red light, and the influence of the inclusion of DIP on photohemolysis was assessed as a function of sensitizer concentration, light dose, and storage time. Furthermore, the possible interference of DIP with inactivation of extracellular virus by use of a panel of different viruses (HIV-1, pseudorabies virus [PRV], bovine viral diarrhea virus [BVDV], VSV, encephalomyocarditis, and canine parvovirus) was investigated. RESULTS: In WBC-reduced RBC suspensions (30% Hct), DIP exerted a clear protective effect against photohemolysis. Part of this protection was achieved with concentrations near the dissociation constant for band III binding. Importantly, efficiency of inactivation of extracellular HIV-1, PRV, BVDV, and VSV was not significantly impaired by the inclusion of DIP. Phototreatment conditions, resulting in a 4 to 5 log inactivation of extracellular HIV-1 and PRV, resulted in a high level of hemolysis after 28 days of storage. This long-term hemolysis could be decreased, but not completely prevented, by the inclusion of DIP. CONCLUSION: Photohemolysis in RBC concentrates can be reduced substantially by the application of DIP, while the efficacy of inactivation of HIV-1 and other viruses remains unchanged.

Research paper thumbnail of Should DEHP be eliminated in blood bags?

Vox Sanguinis, 2013

Whole blood for transfusion was initially collected in glass bottles, but these are fragile, heav... more Whole blood for transfusion was initially collected in glass bottles, but these are fragile, heavy to transport and prone to bacterial contamination. After the Second World War, Carl Walter experimented with plastics for collection and storage of blood, and found that di-ethylhexyl phthalate (DEHP) as plasticizer for polyvinyl chloride (PVC) had the most favourable properties [1, 2]. The breakage rate of frozen plasma in DEHP PVC is low, and the better storage properties of red cell concentrates in DEHP PVC are explained by leaching of plasticizer into the lipid bilayer of these cells, thereby improving membrane stability resulting in lower haemolysis rates [3]. However, ever since the 1970s, scientific and public concern has been raised on the toxicity of DEHP for transfused patients [4, 5]. DEHP and its metabolite mono ethylhexyl phthalate (MEHP) are associated with impairment of reproduction in animal models, resulting in testicular dysgenesis syndrome in (male) rodents, as reviewed in depth elsewhere [6], and particularly, neonates are susceptible to the risks of high exposure to DEHP. Due to the omnipresence of phthalates (not only DEHP), and some important differences in DEHP metabolism between animal models and humans [6], it is difficult to estimate the effect of DEHP toxicity in humans. A study of 234 young men showed no effect of MEHP in urine on their reproductive function [7], but others showed that DNA damage in sperm was associated with the MEHP concentration (after adjustment for other DEHP oxidative metabolites) [8]. In vitro human testis explants incubated with DEHP or MEHP showed that both significantly inhibited testosterone production [9], and a study of 881 men showed a 9% lower free androgen index between the lowest and the highest quartile of the proportion DEHP excreted as MEHP (P = 0.02) [10]. Also, an association was found between urinary MEHP concentrations and lower free T4 and lower total T3 thyroid hormones in adult men [11]. In addition to these effects on hormonal level in adult males, DEHP and its metabolites can have an effect on fetal development, with a shorter anogenital distance associated with higher metabolite concentrations in urine samples collected during pregnancy [12]. Others found no difference in T4 and testosterone levels, as well as no effect on phallic length in 13 male adolescents 14– 16 years of age that had underwent ECMO treatment in their neonatal period, as compared to ageand sexmatched controls [13]. Also, DEHP/MEHP exposure (as detected in cord blood) was significantly associated with a shorter pregnancy duration [14]. Despite these associations, they should be interpreted with care. DEHP is not the only phthalate that may cause these effects; studies that showed no association may not have been published; DEHP administered through the intestinal tract is converted to MEHP in a higher degree as by intravenously [15]; and because phthalates are everywhere in our environment, comparisons are less versus more, rather than absent versus present. DEHP-plasticized PVC has long been used as standard material for a wide range of applications, like packing foil (including food), building material, toys and plastic devices. DEHP is considered to be a ubiquitous environmental contaminant and is present in air, dust, water, soil and food. The latter is considered to be the main source of DEHP intake for the general population. The current daily intake is estimated to be between 2 and 5 lg/kg for adults and between 4 to 8 lg/kg for children [16, 17]. Various measures to reduce the use of DEHP-plasticized PVC have indeed resulted in a 2-times lower exposure to DEHP in the general population over the last two decades [18]. DEHP is applied in many medical devices, and depending on the procedure, patients can be exposed to high (peak) concentrations of DEHP, in the order of 1 mg per kg of bodyweight. One of the sources of DEHP exposure is the leaching from the blood bags into the content of the container, particularly into lipophilic solutions such as plasma [19], but also red cells. Therefore some patient groups, including pregnant or nursing women and children, are considered to be most at risk of possible harmful effects attributable to DEHP exposure by medical treatments. DEHP is broken down into various metabolites. First, it is hydrolysed to MEHP and then further oxidized to mono-(2-ethyl-5-hydroxyhexyl) phthalate (5OH-MEHP),

Research paper thumbnail of Storage of Leukocyte-Poor Red Cell Concentrates: Filtration in a Closed System Using a Sterile Connection Device

Vox Sanguinis, 1989

Storage of leukocyte-poor red cell concentrates (LP-RCC) was investigated after filtration in a c... more Storage of leukocyte-poor red cell concentrates (LP-RCC) was investigated after filtration in a closed system that was assembled using a Sterile Connection Device (SCD). The LP-RCC were stored for up to 6 weeks following filtration with either 0.9% saline solution (n = 14) or saline-adenine-glucose-mannitol (SAG M) solution (n = 15) to prime and rinse the cellulose acetate filter. The results were compared with the data of nonfiltered buffy-coat-poor red cell concentrates (BC-poor RCC) stored in SAG M solution (n = 10). All LP-RCC contained less than lo6 leukocytes whereas the nonfiltered BC-poor RCC contained 675+286 x lo6 leukocytes at day 1, decreasing to 83+49 X lo6 at day 42. Although glucose consumption, lactic acid production and decrease in pH was similar from day 7 through 28 in both groups of LP-RCC, a significantly steeper decline of ATP values as well as a higher hemolysis and LDH release was observed in the LP-RCC filtered with saline. During storage of the nonfiltered BC-poor RCC in SAG M, significantly higher glucose consumption (p<O.Ol), LDH release (p<O.OOl), rate of hemolysis (p<O.OOl) and a lower pH (p<O.OOl) were found, compared to the filtered units. It is postulated that the leukocytes present in the nonfiltered BC-poor RCC were responsible for these differences. The ATP values in the SAG-M-filtered and nonfiltered BC-poor RCC in SAG M were comparable. By comparing the ATP levels and values of the filtered RCC and the nonfiltered BC-poor RCC we conclude that the LP-RCC can be stored for 35 days if SAG M solution is used to prime and rinse the filter.

Research paper thumbnail of A novel flow cytometry–based platelet aggregation assay

Blood, 2013

Key PointsFCA is a novel flow cytometry–based platelet aggregation assay that allows single recep... more Key PointsFCA is a novel flow cytometry–based platelet aggregation assay that allows single receptor analysis in small volume/thrombocytopenic samples FCA facilitates platelet studies in experimental animal models even during gestation and allows kinetic measurements in individual animals

Research paper thumbnail of Inhalt, Vol. 18, Suplement 1, 1991

Transfusion Medicine and Hemotherapy, 1991

Research paper thumbnail of Clinical and in vitro evaluation of red blood cells collected and stored in a non‐DEHP plasticized bag system

Research paper thumbnail of Small but clear pro-hemostatic effects of blood donation-induced stress on donor’s hemostasis

Background: Donating blood induces stress responses in donors, e.g. increased anxiety and blood p... more Background: Donating blood induces stress responses in donors, e.g. increased anxiety and blood pressure. Whereas acute stress in a non-donation setting is associated with pro-hemostatic effects, such effects in blood donors have not been investigated, despite potential importance for the quality of blood products. Therefore, this study examines whether donation-induced stress has an effect on hemostatic parameters in whole-blood donors. Study design and methods: In 372 healthy whole-blood donors, three sets of stress-related parameters were assessed at needle insertion: 1) psychological stress (i.e. donation-stress and arousal on a visual analogue scale); 2) hormonal stress, (i.e. cortisol in saliva); and 3) physiological stress (i.e. blood pressure, pulse rate, and pulse rate variability). Hemostatic parameters (PT, aPTT, fibrinogen, factor VII, factor VIII, and von Willebrand factor (vWF)) were assessed using a 5-ml citrated blood sample taken from the diversion pouch directly af...

Research paper thumbnail of Quality of Platelets in Stored Whole Blood

Transfusion Medicine Reviews, 2020

Transfusion of whole blood rather than blood components is gaining popularity. It is easy to use,... more Transfusion of whole blood rather than blood components is gaining popularity. It is easy to use, with one transfusion product to administer rather than 3, and is held at one storage temperature. It only contains anticoagulant-preservative solution, while components contain various storage solutions, which in theory may induce dilution coagulopathy. In this review, the quality of platelets in stored whole blood is summarized. In cold-stored whole blood, the platelet count declines by 1% to 2% per day. The responsiveness to various agonists declines during the storage time, but this appears to have a limited impact on clotting time or on clot strength as measured with thromboelastography. Animal studies have confirmed that platelets from stored whole blood participate equally well in clot formation. The recovery of platelets in stored whole blood is acceptable during at least 15 days of storage. The survival of platelets after transfusion is only 1 day, but this is likely to be sufficient for the intended patient group requiring massive transfusions, as the platelets are rapidly consumed in the wound area. In addition to the logistic benefits, there are drawbacks, most importantly having a sufficiently large inventory with an acceptable outdating rate, particularly since massive transfusions are rare, while requiring a lot of whole blood. The positive experience of the United States military with whole blood transfusion is often brought forward for introduction in the civilian blood bank, but patients with trauma are only a small fraction of the civilian population requiring massive transfusions. It needs to be determined whether in the resourceful environment of the hospital, these patients benefit from whole blood transfusions. Optimization of whole blood storage, with focus on platelet quality, needs to be performed to allow extension of the storage time beyond 15 days to a point where the number of units in inventory and outdating can be balanced.

Research paper thumbnail of The endosomal RIN2/Rab5C machinery prevents VEGFR2 degradation to control gene expression and tip cell identity during angiogenesis

Angiogenesis, 2021

Sprouting angiogenesis is key to many pathophysiological conditions, and is strongly regulated by... more Sprouting angiogenesis is key to many pathophysiological conditions, and is strongly regulated by vascular endothelial growth factor (VEGF) signaling through VEGF receptor 2 (VEGFR2). Here we report that the early endosomal GTPase Rab5C and its activator RIN2 prevent lysosomal routing and degradation of VEGF-bound, internalized VEGFR2 in human endothelial cells. Stabilization of endosomal VEGFR2 levels by RIN2/Rab5C is crucial for VEGF signaling through the ERK and PI3-K pathways, the expression of immediate VEGF target genes, as well as specification of angiogenic ‘tip’ and ‘stalk’ cell phenotypes and cell sprouting. Using overexpression of Rab mutants, knockdown and CRISPR/Cas9-mediated gene editing, and live-cell imaging in zebrafish, we further show that endosomal stabilization of VEGFR2 levels is required for developmental angiogenesis in vivo. In contrast, the premature degradation of internalized VEGFR2 disrupts VEGF signaling, gene expression, and tip cell formation and migr...

Research paper thumbnail of Metabolic effect of alkaline additives and guanosine/gluconate in storage solutions for red blood cells

Transfusion, 2018

Background-Over a century of advancements in the field of additive solutions for red blood cell s... more Background-Over a century of advancements in the field of additive solutions for red blood cell storage has made transfusion therapy a safe and effective practice for millions of recipients worldwide. Still, storage in the blood bank results in the progressive accumulation of metabolic alterations, a phenomenon that is mitigated by storage in novel storage additives, such as alkaline additive solutions. While novel alkaline additive formulations have been proposed, no metabolomics characterization has been performed to date. Study design and Methods-We performed UHPLC-MS metabolomics analyses of red blood cells stored in SAGM (standard additive in Europe), PAGGSM or alkaline additives SOLX, ESOL-5 and PAG3M for either 1, 21, 35 (end of shelf-life in The Netherlands) and 56 days. Results-Akaline additives (especially PAG3M) better preserved 2,3-diphosphoglycerate and adenosine triphosphate (ATP). Deaminated purines such as hypoxanthine were predictive of hemolysis and morphological alterations. Guanosine supplementation in PAGGSM and PAG3M fueled ATP generation by feeding into the non-oxidative pentose phosphate pathway via phosphoribolysis. Decreased urate to hypoxanthine ratios were observed in alkaline additives, suggestive of decreased generation of urate and hydrogen peroxide. Despite the many benefits observed in purine and redox metabolism, alkaline additives did not prevent accumulation of free fatty acids and oxidized byproducts, opening a window for future alkaline formulations including (lipophilic) antioxidants.

Research paper thumbnail of In vivo labelling of intermediates in the discontinuous synthesis of mRNAs in Trypanosoma brucei

The EMBO Journal, 1987

Discontinuous mRNA synthesis in trypanosomes is thought to involve a 140-nucleotide precursor, ca... more Discontinuous mRNA synthesis in trypanosomes is thought to involve a 140-nucleotide precursor, called the mini-exonderived RNA or medRNA, which contributes its 5' 35 nucleotides to the 5' end of nascent mRNAs. We used in vivo labelling of RNA to show that medRNA has a half-life of <6 min, whereas putative high mol. wt intermediates containing the 3' part of the medRNA have an average half-life of < 1 min. This eliminates priming of pre-mRNA synthesis by intact medRNA as the main mode of discontinuous mRNA synthesis. Potential intermediates of 35 and 105 nucleotides were labelled in parallel with medRNA, but their significance could not be assessed in RNA preparations containing med-RNA, as they are also produced by artefactual cleavage of medRNA. We show, however, that high mol. wt RNA, free of medRNA, can release medRNA segments upon a debranching treatment. These results are consistent with a trans splicing mechanism involving short-lived forked intermediates, analogous to lariats in cis splicing systems. Key words: discontinuous transcription/in vivo labelling/RNA turnover/trans splicing/trypanosomes 1984; Laird et al., 1985; Lenardo et al., 1985a). This mini-exonderived RNA (medRNA) is thought to transfer its mini-exon sequence to the 5' end of pre-mRNAs, but how is unknown. As shown in the schematic diagrams in Figure 1, the medRNA could prime transcription upstream of the main exons or, alternatively, the linkage could occur post-transcriptionally by RNA-RNA ligation or by trans splicing between the medRNA and main exon precursor transcripts. The intermediates are depicted by analogy with nuclear mRNA precursor splicing systems of other eukaryotes (see Padgett et al., 1986). It is likely that a splicing event is responsible for the final joining of the two RNA segments, since the mini-exon sequence is flanked at the 3' end by GT (De Lange et al., 1983) and all main exon sequences are preceded by AG (see Borst, 1986), in compliance with the conventional splice site consensus sequences (Mount, 1982). The pyrimidine-rich stretch upstream of splice acceptor sites in other eukaryotes is less prominent in trypanosomes, while a conserved sequence TTTCPy is found almost invariably in one or more copies within 30 bp upstream of trypanosome splice acceptor sites (P.W.Laird, unpublished results). No pre-mRNA transcription unit has been fully defined yet, but, in the few cases studied, these units appear to span several protein-coding regions, which lack conventional introns (see Borst, 1986). Studies of the mechanism of discontinuous mRNA synthesis have been hampered by the limited available methodology. We have used in vivo labelling of RNA to study the turnover of med-RNA, to track the fate of its 5' and 3' segments and to identify possible intermediates.

Research paper thumbnail of The Platelet Adhesion Capacity to Subendothelial Matrix and Collagen in a Flow Model during Storage of Platelet Concentrates for 7 Days

Thrombosis and Haemostasis, 1994

SummaryThe influence of storage of platelet concentrates (PC) on the adhesion capacity of platele... more SummaryThe influence of storage of platelet concentrates (PC) on the adhesion capacity of platelets was studied. Twenty-four PC, 12 prepared by the buffy coat (BC) method and 12 by the platelet-rich plasma (PRP) method, were stored for 7 days at room temperature. On days 1,3 and 7 of storage, the platelet adhesion capacity to subendothelial matrix (SEM) and collagen was studied in a rectangular perfusion system under flow conditions in conjunction with the platelet aggregation capacity after stimulation and the adenine nucleotide content. The platelet adhesion capacity to collagen was constant until day 3 of storage and decreased to about 80% of the starting value on day 7 of storage. The adhesion capacity to SEM, however, had already decreased on day 3 to about 75% of the value of day 1 and was even more decreased on day 7 to about 45% of the starting value. On day 1, platelets prepared by the BC method displayed a higher adhesion capacity to collagen and a higher aggregation capac...

Research paper thumbnail of Apheresis causes complement deposition on red blood cells (RBCs) and RBC antigen alterations, possibly inducing enhanced clearance

Research paper thumbnail of A comprehensive proteomics study on platelet concentrates: Platelet proteome, storage time and Mirasol pathogen reduction technology

Platelets, Jan 19, 2018

Platelet concentrates (PCs) represent a blood transfusion product with a major concern for safety... more Platelet concentrates (PCs) represent a blood transfusion product with a major concern for safety as their storage temperature (20-24°C) allows bacterial growth, and their maximum storage time period (less than a week) precludes complete microbiological testing. Pathogen inactivation technologies (PITs) provide an additional layer of safety to the blood transfusion products from known and unknown pathogens such as bacteria, viruses, and parasites. In this context, PITs, such as Mirasol Pathogen Reduction Technology (PRT), have been developed and are implemented in many countries. However, several studies have shown in vitro that Mirasol PRT induces a certain level of platelet shape change, hyperactivation, basal degranulation, and increased oxidative damage during storage. It has been suggested that Mirasol PRT might accelerate what has been described as the platelet storage lesion (PSL), but supportive molecular signatures have not been obtained. We aimed at dissecting the influenc...

Research paper thumbnail of Vox Sanguinis International Forum on platelet cryopreservation: Summary

Vox Sanguinis, 2017

The demand for platelets continues to rise, outstripping supply worldwide. In higher-income count... more The demand for platelets continues to rise, outstripping supply worldwide. In higher-income countries, cancer patients remain platelet transfusion dependent for extended periods, which leads to sporadic and chronic regional shortages that are secondary to uncontrolled fluctuations in supply. In less-developed countries, the supply to demand ratio is critical for obstetric haemorrhage, trauma and medical patients. All countries, however, share the difficulty in providing platelets to rural areas or in support of troops in remote military theatres. Alternatives to conventional 5-to 7-day room temperature-stored platelets are needed. Cryopreservation of platelets is likely the best alternative to balance the supply and demand for platelets in cities and less accessible areas. Cryopreserved platelet units (CPP) can be shipped on dry ice and stored for 2 years or more in mechanical freezers or using liquid nitrogen [1]. This controlled supply may be used as a backup to liquid units in well-stocked hospitals, or as the sole source of platelets in remote or underserved settings. Cryopreserved units can be stockpiled for natural disasters, or special HLA-or HPA-selected units may be stored for patients who are refractory to platelet transfusions due to antibodies. In addition, cryopreservation reduces the risk of septic transfusion reactions, as bacteria will not proliferate under these conditions. Cryopreservation of platelets has been available since the 1970s, when Valeri published his original method for storing platelets in 6% dimethyl sulphoxide (DMSO) at-80°C [2]. Subsequent work led to an improved 'no-wash' method, which removed the DMSO prior to freezing, thus allowing for easier post-thaw utilization [1]. While other methods for cryopreservation have been published [3, 4], most data on CPPs in vitro characteristics, storage duration, autologous survival/recovery, clinical safety and efficacy have been developed using the 'no-wash' approach. Extensive in vitro characterization has been performed using CPP resuspended in plasma, saline or platelet additive solutions. These studies have shown that post-thaw platelets, when compared to liquid platelets, are partially activated, contain a distinct microparticle population and

Research paper thumbnail of Transfusion of 35-Day Stored RBCs in the Presence of Endotoxemia Does Not Result in Lung Injury in Humans

Critical care medicine, Jan 2, 2016

Transfusion-related acute lung injury is the leading cause of transfusion-related mortality. Prec... more Transfusion-related acute lung injury is the leading cause of transfusion-related mortality. Preclinical studies have shown that aged RBCs can induce transfusion-related acute lung injury in the presence of a "first hit" (e.g., sepsis). Clinical studies, however, show conflicting results on this matter. We tested whether maximally stored RBCs are able to induce lung injury in the presence of a "first hit" in humans (Dutch Trial Register: NTR4455). Open-label, randomized controlled trial. Healthy male volunteers. Eighteen healthy male volunteers donated one unit of autologous RBCs 2 or 35 days before the experiment. The experiment was started by infusion of 2 ng/kg lipopolysaccharide ("first hit"). After 2 hours, volunteers received normal saline (n = 6), 2-day stored transfusion (n = 6), or 35-day stored transfusion (n = 6) ("second hit"). Blood was sampled hourly. Six hours after transfusion, the diffusion capacity of the lungs for carbon mon...

Research paper thumbnail of Adhesion of anaerobic bacteria to platelet containers

Vox Sanguinis, 2014

Propionibacterium acnes and Staphylococcus saccharolyticus are frequently isolated during platele... more Propionibacterium acnes and Staphylococcus saccharolyticus are frequently isolated during platelet screening with anaerobic culture methods. Although neither P. acnes nor S. saccharolyticus proliferates during platelet storage, both species survive well in this environment. This study was aimed at determining whether strains of P. acnes and/or S. saccharolyticus form surfaceattached bacterial cell aggregates, known as biofilms, under platelet storage conditions. We report that these organisms are able to adhere to the inner surface of platelet containers in tight interaction with activated platelets.

Research paper thumbnail of Comparison of various dimethylsulphoxide-containing solutions for cryopreservation of leucoreduced platelet concentrates

Vox Sanguinis, 2003

Leucoreduced platelet concentrates (LR-PCs) can be stored at 20-24 degrees C for 5-7 days. When L... more Leucoreduced platelet concentrates (LR-PCs) can be stored at 20-24 degrees C for 5-7 days. When LR-PCs are cryopreserved they can be stored for several years. For cryopreservation to become applicable in blood-bank practice, an off-the-shelf cryoprotectant is needed that can be added to the LR-PC in a sterile manner. For this, we varied the composition of the cryopreservation medium and studied various parameters of cryopreserved LR-PCs for up to 24 h after thawing at room temperature. LR-PCs in plasma or Composol were concentrated and divided into 2 units. To each unit, an equal part of 10% dimethylsulfoxide (DMSO) in plasma, Composol with or without 5% albumin, or GPO (pasteurized plasma-protein solution) was added. Freezing occurred at 1 degrees C/min and LR-PCs were placed in the vapour phase of nitrogen. LR-PCs were thawed at 37 degrees C and stored at room temperature. LR-PCs were tested for morphology, platelet recovery, swirling effect, and activation antigens at various time-points thereafter. LR-PCs in 100%, 65% and 50% plasma supplemented with Composol showed good morphology scores (&amp;amp;amp;amp;gt;250), limited CD62P expression (&amp;amp;amp;amp;lt;35%), low CD63 expression (&amp;amp;amp;amp;lt;20%) and a swirling effect of about 2, at 24 h after thawing. At the same time-point, platelet recovery was &amp;amp;amp;amp;gt;80% under all conditions and CD42b expression varied between 70 and 85%. Results of LR-PCs in 15% plasma and Composol, with or without plasma substitutes, were not acceptable at 24 h after thawing, i.e. the morphology score was &amp;amp;amp;amp;lt;200 and the CD62P expression was &amp;amp;amp;amp;gt;40%. A minimum of 50% plasma in the cryopreserved LR-PC is necessary to maintain an acceptable in vitro quality of platelets up to 24 h after thawing. Composol is a good candidate for using to prepare an off-the-shelf cryoprotectant.