Recommendations for in vitro evaluation of blood components collected, prepared and stored in non-DEHP medical devices.

BACKGROUND AND OBJECTIVES: DEHP, di(2-ethylhexyl) phthalate, is the most common member of the class of ortho-phthalates, which are used as plasticizers. The Medical Device Regulation has restricted the use of phthalates in medical devices. Also DEHP has been added to the Annex XIV of REACH, "Registration, Evaluation, Authorisation and Restriction of Chemicals" due to its endocrine disrupting properties to the environment. As such, the sunset date for commercialisation of DEHP-containing blood bags is May 27th 2025. There are major concerns in meeting this deadline as these systems have not yet been fully validated and/or CE-marked. Also, since DEHP is known to affect red cell quality during storage, it is imperative to transit to non-DEHP without affecting blood product quality. Here, EBA members aim to establish common grounds on the evaluation and assessment of blood components collected, prepared and stored in non-DEHP devices. MATERIALS AND METHODS: Based on data as well as the input of relevant stakeholders a rationale for the validation of each component was composed. RESULTS: The red cell components will require the most extensive validation as their quality is directly affected by the absence of DEHP, as opposed to platelet and plasma components. CONCLUSION: Studies in the scope of evaluating the quality of blood products obtained with non-DEHP devices, under the condition that they are carried out according to these recommendations, could be used by all members of the EBA to serve as scientific support in the authorization process specific to their jurisdiction or for their internal validation use.

DEHT is a suitable plasticizer option for phthalate-free storage of irradiated red blood cells.

BACKGROUND AND OBJECTIVES: Due to increasing concerns about possible endocrine-disrupting properties, the use of the plasticizer di(2-ethylhexyl) phthalate (DEHP) will be banned in future blood storage. Di(2-ethylhexyl) terephthalate (DEHT) provides sufficient red blood cell (RBC) quality during conventional blood bank storage. It is important that a new plasticizer also maintains acceptable quality during exposure to high cell stress, such as irradiation, which is commonly used to prevent graft-versus-host disease. MATERIALS AND METHODS: A total of 59 RBC units were collected and processed in polyvinyl chloride (PVC)-DEHT or PVC-DEHP blood bags combined with either saline-adenine-glucose-mannitol (SAGM) or phosphate-adenine-glucose-guanosine-saline-mannitol (PAGGSM) additive solution. All units were X-ray irradiated on day 2 post-collection. Sampling for assessment of parameters of storage lesion was performed on day 2 pre-irradiation and day 14 and 28 post-irradiation. RESULTS: Though irradiation increased cell stress, DEHT/PAGGSM and current common European preference DEHP/SAGM were equally affected up to 14 days post-irradiation for all measured parameters. At day 28, haemolysis and microvesicle count were slightly increased in DEHT, whereas extracellular potassium ions, glucose, lactate, pH, mean corpuscular volume and microvesicle phosphatidylserine remained unaffected by plasticizer choice throughout storage. No individual unit exceeded 0.8% haemolysis, not even in DEHT/SAGM, the combination overall most affected by irradiation. Of the four combinations, membrane stability was least impacted in DEHP/PAGGSM. CONCLUSION: We demonstrate that DEHT is a suitable plasticizer for storage of RBCs after X-ray irradiation cell stress. This strengthens the option of DEHT as a viable non-phthalate substitute for DEHP.

Non-phthalate plasticizer DEHT preserves adequate blood component quality during storage in PVC blood bags.

BACKGROUND AND OBJECTIVES: Commercial blood bags are predominantly made of polyvinyl chloride (PVC) plasticized with di(2-ethylhexyl) phthalate (DEHP). DEHP is favourable for storage of red blood cells (RBC). Historically, removal of DEHP from blood bags has been linked to unacceptable haemolysis levels. Oncoming regulatory restrictions for DEHP due to toxicity concerns increase the urgency to replace DEHP without compromising RBC quality. Di(2-ethylhexyl) terephthalate (DEHT) is one suggested substitute. The aim of this study was to compare PVC-DEHT to PVC-DEHP blood bags using additive solutions saline-adenine-glucose-mannitol (SAGM) and phosphate-adenine-glucose-guanosine-saline-mannitol (PAGGSM), to determine whether DEHT can maintain acceptable component quality. MATERIALS AND METHODS: RBC concentrates (N = 64), platelet concentrates (N = 16) and fresh frozen plasma (N = 32) were produced from whole blood collected into either DEHT or DEHP plasticized systems. Using a pool-and-split study design, pairs of identical RBC content were created within each plasticizer arm and assigned either SAGM or PAGGSM. Storage effects were assessed weekly for 49 days (RBC), 7 days (platelets) and before/after freezing (plasma). RESULTS: Though haemolysis was slightly higher in DEHT, all study arms remained below half of the European limit 0·8%. K+ was lower in DEHT than in DEHP independent of additive solution. The metabolic parameters were not influenced by choice of plasticizer. Platelet activation/metabolism and plasma content were similarly preserved. CONCLUSION: Our study demonstrates that the plasticizer DEHT provides adequate blood component quality. We propose DEHT as a strong future candidate for replacement of DEHP in blood bags.

A novel protocol for cryopreservation of paediatric red blood cell units allows increased availability of rare blood types.

BACKGROUND AND OBJECTIVES: There is a growing concern for shortage in future blood supply, caused by a predicted decrease in eligible blood donors and simultaneous increase in recipients. Cryopreservation of split red blood cell units could increase stock supply by reducing waste of rare blood. This would be particularly useful in paediatric care where very small volumes often are transfused. The aim of this study was to develop a cryopreservation protocol for split units using the closed-system automated cell processor ACP215, as such protocols are currently missing. MATERIALS AND METHODS: Using a pool-and-split design, red blood cell units (N = 8) were glycerolized and frozen, either as standard volume reference units, or further divided into three smaller split units each. After thawing/deglycerolization, the supernatant of the smaller splits was reduced by additional centrifugation, and new SAGM was added to 60% haematocrit. The units were analysed for storage lesion effects up to ten days post-thawing. RESULTS: Haemolysis and extracellular potassium ion levels were lower in the split units than in the whole units from day three onwards. The metabolic parameters pH, ATP, glucose and lactate were also lower, though likely caused by lower additive solution pH rather than storage. CONCLUSION: Split units of red blood cells can be successfully cryopreserved using the ACP215. In addition to favourably low haemolysis and potassium, they also have higher haematocrit than corresponding whole units and enable involvement of fewer donors at repeated transfusions. These characteristics are all desirable features in paediatric care.

Pathogen reduced red blood cells as an alternative to irradiated and washed components with potential for up to 42 days storage.

BACKGROUND: The urgency of maintaining a safe and adequate blood supply is increasing. One approach to ensure a sufficient supply is to limit the outdating frequency of blood components. Pathogen inactivation technology was developed primarily to increase safety by preventing transmission of infectious diseases. The Intercept Blood System for pathogen reduction of red blood cells (RBC) has additional benefits such as inactivation of leucocytes and removal of plasma and storage debris through centrifugation. Irradiation and automated washing are detrimental to the RBC membrane and often implicate shortened shelf-life. We aimed to assess whether pathogen inactivation can replace RBC irradiation and washing to avoid shelf-life reduction. MATERIALS AND METHODS: RBC concentrates (No.=48) were pooled-and-split into four study arms, which underwent pathogen inactivation treatment, irradiation, automated washing or no treatment (reference). RBC quality was evaluated during 42 days by assessment of storage lesion. Washing efficacy was defined by IgA and albumin reduction. RESULTS: Pathogen reduced RBCs had similar membrane preservation to reference RBCs (hemolysis, microvesicles and extracellular potassium ions), whereas the RBCs were negatively impacted by irradiation or automated washing. ATP increased substantially post-pathogen inactivation, while 2,3-DPG decreased. Pathogen inactivation considerably reduced albumin and IgA, though slightly less efficiently than automated washing. DISCUSSION: RBCs exhibit superior membrane preservation after pathogen inactivation treatment, compared to both irradiation and automated washing. This suggests that replacement is possible, even though the plasma reduction protocol could be further optimised.Replacement of irradiated and washed RBC concentrates with pathogen reduced RBC concentrates storable up to 42 days would be advantageous for both the blood supply and patient safety.

Cryopreserved platelets and amotosalen-treated plasma in an experimental clot formation set-up.

BACKGROUND: Amotosalen treatment of plasma and cryopreservation of platelets affect the quality and potentially the interplay between platelets and coagulation factors. We set up an experimental clot formation study to test the hypothesis that amotosalen treatment of plasma affects the interaction with different platelet preparations. MATERIALS AND METHODS: Pooled plasma units (n=16) were subjected to coagulation tests before and after pathogen inactivation with amotosalen treatment (PI) and aliquots were frozen at -80°C. Fresh and cryopreserved platelets were analyzed for phenotypic and activity markers. Finally, coagulation properties of different combinations of platelets and plasma, before and after PI, were analyzed by viscoelastography (ROTEM). RESULTS: PI of plasma reduced the concentration of several coagulation factors (p<0.01). Cryopreservation altered phenotypic expression and reduced the platelets' ability to respond to agonists (p<0.0001). The interplay between all plasma derivatives and cryopreserved platelets resulted in shortened coagulation time (p<0.0001) but prolonged clot formation time and reduced clot strength (p<0.0001) as compared to the interaction between fresh platelets with different plasma variants. PI of the plasma does not seem to have a major impact on coagulation time, clot formation time or clot strength. DISCUSSION: Our data show that the reduced concentration of coagulation factors after PI treatment of plasma are negligible measured by viscoelastography, with fresh and cryopreserved platelets in this experimental clot formation setup, and that platelets play a more pronounced role. Cryopreserved platelets are more activated and result in reduced clot stability.

Weight Development in Children After Gastric Bypass Surgery.

Objective: More and more young obese women get pregnant after undergoing gastric bypass surgery (GBP) but little is known about weight development in their offspring. The first aim of this study was to investigate weight development of children whose mothers have undergone GBP before pregnancy and compare them to age specific reference values in Sweden. Second aim was to study the frequency of small for gestational age (SGA) in this population. Materials and methods: Weight of offspring (38 male and 28 female) where the mother had undergone GBP before pregnancy were studied from birth up to 18 months of age and compared to age-specific reference values in Sweden. Results: The boys to mothers who had undergone GBP before pregnancy weighed more than Swedish reference values at 6 months, 8.44 ± 1.18 kilogram (kg) (n = 35) vs. 7.98 ± 0.81 kg (n = 1388; p = 0.001), and less at 18 months, 11.54 ± 0.93 kg (n = 19) vs. 12.27 ± 1.19 kg (n = 862; p < 0.001). The girls to mothers who had undergone GBP before pregnancy weighed more than Swedish reference values at 6 months, 7.84 ± 1.00 kg (n = 28) vs. 7.50 ± 0.77 kg (n = 1375; p = 0.020). Frequency of SGA was 3.0%. Conclusion: No clear pattern was found concerning the short-term weight development of the children. However, studies with larger material and more follow up time must be performed.

Ultra-high olfactory sensitivity for the human sperm-attractant aromatic aldehyde bourgeonal in CD-1 mice.

Recent studies have shown that certain aromatic aldehydes are ligands for olfactory receptors expressed in mammalian sperm cells and induce sperm chemotaxis. Using a conditioning paradigm, the olfactory sensitivity of five CD-1 mice for seven aromatic aldehydes was investigated. With all seven stimuli, the mice discriminated concentrations as low as 0.01 ppm (parts per million) from the solvent, and with bourgeonal the animals even detected concentrations as low as 0.1 ppq (parts per quadrillion) which constitutes the lowest olfactory detection threshold value reported in this species so far. The presence of a tertiary butyl group in para-position (relative to the functional aldehyde group) combined with a lack of an additional alkyl group next to the functional aldehyde group may be responsible for the extraordinary sensitivity of the mice for bourgeonal.

Digital speckle photography: visualization of mesoflow through clustered fiber networks.

Digital speckle photography (DSP) is used for velocity field measurements inside a fiber network. The width of the channels in which the flow is measured is typically less than 1 mm. Therefore a microscope is used to image the fiber network. When we sample 30 images/s and separate the moving parts of the images from the stationary parts, the velocity field can be deduced with DSP.