| 1. |
- Alattar, AG, et al.
(författare)
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Gene editing of CD34+ progenitor cells from single blood donor waste bags to create cultured early erythroid cells for study of blood group knock-outs
- 2020
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Ingår i: Vox Sanguinis. - : Wiley. - 1423-0410 .- 0042-9007. ; 115:Suppl. s1, s. 363-363
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Konferensbidrag (refereegranskat)abstract
- Background: Many blood group antigens are carried by red cell surface proteins, thefunctions of which are not yet fully characterized or in some cases completelyunknown. By investigating red cells with naturally-occurring blood group variants,much has been learnt about the underlying molecules. However, interindividualvariation affecting other molecules than the one(s) of interest may confound orotherwise hamper such studies. As an alternative, a reductionistic approach whereonly a single factor differs between test and control cells significantly facilitatesinterpretation of functional studies. Using various siRNA, shRNA and various gene-editing tools blood group expression can be manipulated and useful modelsdeveloped. Applying the latter on primary hematopoietic stem and progenitor cells(HSPCs) can be challenging.Aims: We evaluated a protocol for gene editing of CD44 using a CRISPR/Cas9hybrid system on HSPCs isolated from blood donation leukocyte waste bags fromsingle donors to develop a model for study of blood group molecular function inerythropoiesis.Methods: Peripheral blood mononuclear cells (PBMCs) were from anonymizedleucocyte waste bags obtained after whole blood unit processing in the Reveosautomated blood component system. Cells were harvested following Lymphoprepgradient separation and CD34+HSPCs enriched and collected using magnetic beads.CD34+cells were cultured in 2-phase culture medium to generate erythroid cellsfrom HSPCs (Vidovic, Vox Sang 2017). For CRISPR/Cas9 gene editing, a short guideRNA (sgRNA) targeting CD44 was designed and cloned into the lentiCRISPR v2vector (Addgene plasmid #52961). A non-targeting sgRNA cloned into the vectorwas used as control. Lentivirus particles were produced in the human 293T cell lineas described previously (Galeev, Methods Mol Biol 2017). Equal number of CD34+cells were transduced 24 hours after collection using RetroNectin following theRetroNectin-Bound Virus (RBV) Infection Method according to manufacturer’sprotocol. Cells were transduced at a multiplicity of infection (MOI) of 10 with atarget transduction efficiency of 20–30%. Cells were cultured at 37°C, 5% CO2 for72 hours in phase I culture medium and then electroporated with Cas9 mRNA usingthe ECM 830 Electroporation System as described previously for cord blood-derivedCD34+cells (Backstrom, Exp Hematol 2019). GFP+CD44-edited cell frequencieswere monitored by flow cytometry at day 7 of the HSPC expansion phase and atday 14 of the erythroid expansion-differentiation phase using antibodies againsterythroid-specific cell surface markers GPA and Band3 in addition to CD49d toassess the erythroid development stage.Results: We tested the above protocol and observed that the frequencies of editedcells lacking CD44 expression within the GFP+population at day 7 of culture were10–25% while the edited frequencies were increased at day 14 of culture to 60–80%within the GFP+cells. Whilst this stage corresponds to erythroblasts, earlier or laterstages can be tested.Summary/Conclusions: The previously established hybrid system for CRISPR/Cas9gene editing in cord blood-derived CD34+HSPCs, which combines lentiviraldelivery of the sgRNA with transient delivery of Cas9 mRNA by electroporation, isalso applicable to primary human adult HSPCs from Reveos-processed single-donorwhole blood donation, resulting in a traceable high-yield gene-editing system tostudy blood group function and erythroid development
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| 2. |
- Anderson, H, et al.
(författare)
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Blood transfusion at delivery and risk of subsequent malignant lymphoma in the mother
- 1998
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Ingår i: Vox Sanguinis. - 0042-9007. ; 75:2, s. 145-148
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND AND OBJECTIVES: Blood transfusion has been shown to be a risk factor for non-Hodgkin's lymphoma (NHL).MATERIALS AND METHODS: In a cohort of 77,928 women with bleeding complications at delivery in the period of 1973-1986, subsequent NHL cases were identified and the number was compared with the number expected from national incidence rates. In a case-control study the proportion of transfused NHL cases was compared with the proportion of transfused controls.RESULTS: The observed number of NHL in the cohort was 18 versus 22.0 expected. Information on transfusion was obtained for 15 of the NHL cases and none (0%) was transfused versus 32 out of 136 controls (23%).CONCLUSIONS: Blood transfusion at delivery is not a risk factor for NHL. The immune tolerance induced by pregnancy may reduce the risk of NHL associated with the transfusion of allogeneic blood cells.
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| 3. |
- Backteman, K, et al.
(författare)
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A rapid and reliable flow cytometric routine method for counting leucocytes in leucocyte-depleted platelet concentrates
- 2002
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Ingår i: Vox Sanguinis. - : Wiley. - 0042-9007 .- 1423-0410. ; 83:1, s. 29-34
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Tidskriftsartikel (refereegranskat)abstract
- Background and Objectives: To ensure a proper quality control it is important to use a reliable method to count low numbers of leucocytes in leucocyte-reduced platelet concentrates (PCs). Materials and Methods: A modified flow cytometric method for counting low numbers of leucocytes, based on a reference population contained in tubes with an exact number of fluorescent beads and staining with propidium iodide was used. To increase the number of events, the original sample volume was increased. Results: There was a good correlation in the number of leucocytes (r = 0.99) between the modified flow cytometric method and microscopy of samples from unfiltered and expected numbers from serially diluted PCs. Samples from leucocyte-reduced PCs obtained by apheresis or filtered buffy coats showed no correlation between results from the modified flow cytometric method and microscopy (Nageotte). Conclusion: Counting by microscopy gave a lower number of leucocytes than the modified flow cytometric method when counting a low number of cells. However, analysis of the serially diluted PCs proved that the modified flow cytometric method was reliable and rapid, making it suitable for clinical routine use.
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| 4. |
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| 5. |
- Berglund, Sofia, et al.
(författare)
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Granulocyte transfusions could benefit patients with severe oral mucositis after allogeneic hematopoietic stem cell transplantation
- 2019
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Ingår i: Vox Sanguinis. - : Wiley-Blackwell. - 0042-9007 .- 1423-0410. ; 114:7, s. 769-777
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Tidskriftsartikel (refereegranskat)abstract
- Background and objectives Mucositis is a common complication after allogeneic hematopoietic stem cell transplantation (HSCT), and is caused by a combination of conditioning-induced mucosal damage and severe neutropenia. The symptoms include oral and abdominal pain, inability to swallow food and fluids, and severe diarrhoea. Severe mucositis is associated with increased risk of Graft-versus-Host disease and infection. Granulocyte transfusions (GCX) could be a treatment option, and our objective was to study its feasibility and potential benefits. Material and methods This retrospective, single-centre study included 30 patients receiving GCX because of severe oral mucositis after HSCT during 2005-2017. Clinical outcome, response to GCX, change in opiate administration and adverse events were studied. Results Twenty-seven patients received GCX from donors pre-treated with steroids and G-CSF, and three from donors pre-treated with steroids only. Overall response was 83% (24/29 evaluable patients). Fifteen patients reached a complete response. In 14 of 24 responders, a reduction of the administration of opiate pain relief was seen. In eight patients this reduction was >= 50% of the dose. Adverse events (AEs) were reported in 14 cases, and were mild to moderate, and well manageable with symptomatic treatment. No life-threatening or fatal AEs were recorded. Conclusions These results indicate that GCX could be a safe and effective treatment for oral mucositis after HSCT with the potential to reduce the necessity of opiate analgesic treatment in this disorder. No severe AEs were seen in this study, but the risk for severe pulmonary AEs after GCX needs to be considered.
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| 6. |
- Berntorp, Erik
(författare)
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Other ongoing rFVIII PUP studies
- 1999
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Ingår i: Vox Sanguinis. - : Wiley. - 1423-0410 .- 0042-9007. ; 77:Suppl. 1, s. 10-12
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Tidskriftsartikel (refereegranskat)
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| 7. |
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| 8. |
- Birgegård, Gunnar, et al.
(författare)
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High incidence of iron depletion and restless leg syndrome (RLS) in regular blood donors : intravenous iron sucrose substitution more effective than oral iron
- 2010
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Ingår i: Vox Sanguinis. - : Wiley. - 0042-9007 .- 1423-0410. ; 99:4, s. 354-361
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Tidskriftsartikel (refereegranskat)abstract
- Background and objectives Iron depletion is common in regular blood donors. The objective of the study was to investigate the frequency and severity of iron depletion in regular blood donors and whether IV iron is more effective than oral to avoid iron depletion and symptoms thereof, especially restless legs syndrome (RLS). Method One hundred and twenty blood donors with at least five previous whole blood donations were randomized to receive either IV iron sucrose (Venofer (R), RenaPharma/Vifor, Uppsala, Sweden), 200 mg, or to 20 x 100 mg of oral iron sulphate (Duroferon (R), GlaxoSmithKline, Stockholm, Sweden), after each blood donation during 1 year. Iron status and RLS incidence and severity were investigated. Results Iron status was generally poor among regular blood donors, especially in women, with a high incidence of iron depletion (> 20%) and RLS (18%). The IV iron group increased storage iron to a greater extent than the oral iron group after 12 months (P = 0 center dot 0043). Female donors were more responsive to IV iron sucrose compared to oral iron sulphate, particularly female donors below 50 years of age. RLS severity scores were significantly lower in the IV iron group. The two treatments were safe. Conclusion Iron status is poor in regular blood donors, restless legs syndrome is common, and the routine iron supplementation is insufficient. IV iron sucrose substitutes iron loss in blood donors more efficiently compared with oral iron sulphate, especially in women. Iron substitution to blood donors should be individualized and based on P-ferritin monitoring.
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| 9. |
- Birgegård, Gunnar, 1944-, et al.
(författare)
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Serum ferritin in the regulation of iron therapy in blood donors
- 1980
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Ingår i: Vox Sanguinis. - 0042-9007 .- 1423-0410. ; 38:1, s. 29-35
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Tidskriftsartikel (refereegranskat)abstract
- 12 regular blood donors were selected on the basis of subnormal serum ferritin levels as a criterion for iron deficiency. It was found that all had high transferrin levels but only 5 had subnormal serum iron or transferrin saturation. The donors were given oral iron therapy in a dose of 2,800 mg between each phlebotomy, and the donation interval was standardized to 8 weeks. Test samples were collected every 4th week. After an initial rise in ferritin during the first 2 months, 6 of the donors again had subnormal serum ferritin levels, and the iron dose was therefore doubled after 32 weeks. Following this, all subjects taking the higher dose had normal ferritin values and stainable marrow iron was found at the end of the study, after 92 weeks. 3 subjects did not take the higher dose, had no raised serum ferritin level or stainable hemosiderin. It is concluded that serum ferritin estimation can be used to monitor the therapy in blood donors so that a satisfactory amount of iron is stored.
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| 10. |
- Birgegård, Gunnar, et al.
(författare)
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Serum ferritin levels in male blood donors : relation to number of phlebotomies and iron supplementation
- 1978
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Ingår i: Vox Sanguinis. - 0042-9007 .- 1423-0410. ; 34:2, s. 65-70
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Tidskriftsartikel (refereegranskat)abstract
- Serum ferritin estimation has been shown to be a reliable test to reveal iron deficiency. Such estimations have been made in groups of male blood donors with a varying number of previous phlebotomies and a mean interval between donations of 9.9 +/- 1.7 SD weeks. It was found that the mean ferritin level was significantly (p less than 0.001) lower in the blood donors than in nondonors. After 6-8 phlebotomies it was about 40% lower. Subnormal ferritin values were found in 10% of the donors, almost exclusively among those who had taken less than 1,000 mg of iron supplementation since the last donation. It is concluded that with a donation interval of about 10 weeks, there is a considerable risk for iron deficiency after about 6 donations. This risk is far less if more than 1,000 mg of iron supplementation is taken between phlebotomies. A role for serum ferritin estimation in monitoring donation intervals and/or iron therapy is suggested.
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