Optimizing the recovery of peripheral blood mononuclear cells trapped in leukoreduction filters - A comparison study

Abstract Introduction The isolation of captured peripheral blood mononuclear cells (PBMNCs) from leukoreduction filters (LRFs) can be of great importance in terms of bringing the lost cells back into use. Objective The aim of this study was to evaluate various methods based on their potential to recover the peripheral blood cells from LRFs with a focus on mononuclear cells (MNCs). Method For cell isolation from LRFs, three distinct methods (back-flushing, direct and vacuum pump) were compared through the calculation of the yield of isolated MNCs. The viability of extracted cells was determined by the flow cytometry technique. Moreover, the recovered MNCs were characterized regarding the presence of blood stem cell purification. The cell culture, microscopic observation, and immunophenotyping were employed to characterize the blood stem cells (hematopoietic, mesenchymal and progenitor endothelial stem cells). Results The yield of isolation obtained in the back-flushing, direct and vacuum pump methods were 17.7 ± 1.28, 17.3 ± 0.96 and 21.2 ± 0.90 percent, respectively. Although the highest potential for total blood cell recovery belonged to the vacuum pump method, the lowest cell viability (85.73 ± 4.84%) was observed in this method. However, the isolation process of the back-flushing and direct methods had less effect on cell viability. The characterization of the isolated MNCs displayed that the dominant positive phenotype was for CD34/CD45, indicating hematopoietic stem cells. In addition, the endothelial stem/progenitor cells were significantly detected as CD31/CD133 positive cells. Conclusion According to our results and considering the safety and efficiency potential of each of the applied methods, the back-flushing in comparison with the other methods can be considered a suitable procedure for MNC isolation from LRFs.

Application of leukoreduction therapy in severe pertussis with leukocytosis and pulmonary hyperten-sion:3 infant cases reports and literatures review / 中国小儿急救医学

Objective To investigate the efficacy and safety of leukoreduction therapy in severe per-tussis in infants. Methods Therapeutic processes of 3 cases of severe pertussis in PICU of Shanghai Children′s Medical Center were retrospectively studied from October 2017 to May 2018. We reviewed the related literatures and summarized the time and effectiveness of leukoreduction therapy in severe pertussis. Results All 3 cases had leukocytosis,respiratory faliure,pulmonary hypertension and right heart failure. One case had multiple organ failure before undergoing exchange transfusion therapy and eventually died. Two cases that had pulmonary hypertension during the period of WBC′s rising accepted leukopheresis therapy before multiple organ failure,and eventually survived. We reviewed the foreign literatures which was almost case reports,leukoreduction therapy might improve the prognosis of severe pertussis in infants,but the time of using it had no conclusion. Conclusion This is the first report of leukoreduction therapy for the severe per-tussis in infants in China. It provides a new method for the treatment of severe pertussis in infants. It is worth looking forward to use this method combined with continuous renal replacement therapy and extracorporeal membrane oxygenation technology. In the future,multicenter clinical research should be done to explore the effectiveness and safety of leukoreduction therapy in the severe pertussis in infants.

Proposal of Evaluation Method for Leukoreduction Blood Filter and Evaluation of Domestic Filter / 대한수혈학회지

BACKGROUND: A leukoreduction filter was recently developed in Korea to reduce various kinds of adverse transfusion reactions. The objective of this study was to propose a domestic evaluation system for leukoreduction filters and to apply this evaluation system to assess the newly developed leukoreduction filter. METHODS: We prepared packed red blood cells from 60 units of whole blood (400 mL) collected from 60 normal individuals and evaluated the efficacy of the newly developed filter (FINECELL, KOLON INDUSTRIES, Gumi, Korea) and a control filter (RCM1, Haemonetics, MA, USA). To verify the evaluation system, we assessed the filtration time, residual leukocyte count, RBC recovery, RBC hemolysis, hemoglobin concentration, and hematocrit using a control filter RCM1 and compared the results with those of an evaluation performed by the American Red Cross (ARC) in 2013. We then evaluated the efficacy of the test filter FINECELL using the methods established in this study and compared the results with those of the control filter RCM1. RESULTS: The results of the current study were similar to those of the ARC with the control filters. The test filters developed in Korea were not inferior to commonly used control filters regarding residual leukocyte count, RBC recovery, and RBC hemolysis at 35 days after filtration. All of the results in the evaluation satisfied the international standards. CONCLUSION: These results of this study showed that the efficacy of the newly developed domestic leukoreduction filter were satisfactory and will contribute to improvement of quality of blood components in Korea.

Application of leukoreduction filter in removal of leukocytes in platelet concentrate / 国际检验医学杂志

Objective To investigate the application of leukoreduction filter in removal of leukocytes in platelet concentrate . Methods Platelet concentrate was prepared by using platelet‐rich plasma method .35 bags of the same type of prepared platelet concentrate were filtered by using leukoreduction filter .The changes of conventional indicators of platelet before and after filter were measured and recorded .The activating platelets indicators PAC‐1 and CD62p were detected by using flow cytometry .The platelet hypotonic shock was measured by biochemical analyzer .The platelet aggregation was measured by using platelet aggrega‐tion instrument .Results After platelet concentrate was filtered by using leukoreduction filter ,leukocyte removal rate was(98 .28 ± 0 .97)% ,platelet recovery rate was(86 .37 ± 2 .84)% .After filtration ,white blood cell count ,platelets ,red blood cells were reduced than before filtration(P 0 .05) .Before and after filtration ,the expression of platelet activation markers PAC‐1 and CD62p ,platelet aggregation and platelet hypotonic shock were not statistically different(P>0 .05) .Conclusion Platelet leukore‐duction filter could effectively filter leukocytes in platelet concentrate .It would not change the conventional indicators ,and not affect platelet activation ,aggregation and anti‐hypotonic shock capacity significantly .

Universal Leukoreduction Decreases the Incidence of Febrile Nonhemolytic Transfusion Reactions to Cellular Blood Components: A 5 Year Study.

Background: Febrile nonhemolytic transfusion reactions (FNHTRs) are common complications associated with allogenic transfusion and it is caused by the leucocytes and cytokines released by leucocytes during storage of blood/ blood components. These reactions are generally not life threatening, but they are expensive in their management, evaluation, and associated blood-product wastage. 1st log prestorage universal leukoreduction (ULR) i.e. removal of Buffy coat is a useful and effective procedure in developing countries to control FNHTRs significantly. Aims and Objects: To know the efficacy of pre-storage 1st log universal leuckoreduction in controlling febrile nonhemolytic transfusion reactions (FNHTRs). Place and Duration of Study: Study was carried out at Blood Bank, Department of Pathology, G. R. Medical College, Gwalior from January 2009 to December 2013 (5years). Materials and Methods: Study was divided into control group (Year: 2009) and study group (Years: 2010-13). 14,292 recipients in control group and 45,064 in study group were transfused with non-leukoreduced and prestorage 1st log leukoreduced blood/ blood components respectively. Usefulness of prestorage 1st log ULR over non-leukoreduced blood/ blood components was observed, compared and discussed. Result: In the control group 610 (4.26%) out of 14,292 (p=0.0003) and in study group 381(0.84%) out of 45,064 (p=0.0003) recipients were reported to have FNHTRs. The comparative study showed significant reduction in FNHTRs from 4.26% to 0.84% (↓ 3.42%) (p=0.000001). Conclusion: 1st log Universal Leukoreduction (ULR) is a better option over Selective Leukoreduction (SLR) to prevent FNHTRs and it also helps the transfusion services of under-resourced developing countries in many ways.

The Status of Use of Leukoreduced Blood Products in Korean Hospitals

BACKGROUND: Leukoreduced blood components are recommended for prevention of non-hemolytic febrile transfusion reactions, HLA alloimmunization, platelet transfusion refractoriness, and transfusion-transmissible diseases. In addition, prestorage leukoreduction may be advantageous to poststorage leukoreduction. The authors investigated the current status of usage of leukoreduced blood components in Korea. METHODS: We surveyed 2,373 medical facilities, where blood components were supplied from Korean Red Cross blood centers and/or Hanmaeum blood center during one year period between January and December 2009. The survey was conducted about the current situation of usage of leukoreduction by web-based program (http://bms.cdc.go.kr), and 743 facilities answered and were analyzed. RESULTS: The leukoreduced RBC components comprised 10.3% (prestorage leukoreduction, 91,066 units, 5.7%; poststorage leukoreduction 73,192 units, 4.6%) of the total 1,593,098 units of RBC components used in 743 medical facilities. The leukoreduced platelet concentrates comprised 33.1% (458,552 units) of the total 1,386,184 units of platelet concentrates used in 397 medical facilities. If 1 single donor platelet is counted as 6 platelet concentrates, 48.9% of the total platelet components used were leukoreduced. CONCLUSIONS: The proportion of leukoreduced blood components to the total blood components used in Korea was much lower than that in Unites States of America, especially lower in the use of prestorage leukoreduction of RBC components. Further studies are required for cost-effectiveness and demand-supply amounts of leukoreduced blood components, and appropriate prestorage leukoreduction has to be performed in Korea based on these studies.

Evaluation of RF300 for Leukoreduction of Red Blood Cells / 대한수혈학회지

BACKGROUND: Use of universal leukoreduction for prevention of leukocyte associated transfusion reactions is common practice in many countries. This study was conducted in order to evaluate the performance of a newly developed leukoreduction filter for red blood cells (RBCs), the RF300 (Kolon Industries, Inc, Gumi, Korea). METHODS: Filtration time, RBC recovery, residual leukocyte count, and leukocyte removal rate were evaluated. To assess the quality of RBCs after filtration, percent hemolysis was monitored for a period of 21 days. Performance of the RF300 (N=78) was compared with that of the Bio-R O2 plus (Fresenius, Hamburg, Germany), the Pall Purecell RC (Pall Co., Washington, USA), and the Sepacell R-500N (Asahi, Tokyo, Japan). RESULTS: The shortest filtration time was observed using the RF300 (P<0.05). Using the RF300, recovery of RBC was 96.5%, which was higher than that of two filters (P<0.05). Mean residual leukocyte count was 0.26x10(6)/unit, with a leukocyte removal rate of 3 log. Using the RF300, mean percent hemolysis was 0.32% at day 21, which was comparable with that of two filters, but lower than that of one filter (P<0.05). CONCLUSION: The RF300 meets all established quality requirements for conduct of safe and effective leukoreduction of RBCs.

Reduction of Influenza A (H1N1) Virus Through a Leukoreduction Filter / 대한수혈학회지

BACKGROUND: Leukoreduction can reduce the risk of HLA alloimmunization, recurrent febrile nonhemolytic transfusion reactions, and several transfusion-transmitted infectious diseases, including cytomegalovirus infection. Transmission of the new influenza A (H1N1) virus through transfusion may be a concern. We evaluated the effect of filtration with a leukoreduction filter on H1N1 genomes. METHODS: To evaluate the effect of filtration by a leukoreduction filter on H1N1 genomes, we analyzed pre- and post-filtered samples from nasopharyngeal swabs and 10 positive plasma samples using real time RT-PCR. RESULTS: The 10 samples (nasopharyngeal swabs and plasma) contained H1N1 RNA, and filtration with a leukoreduction filter reduced these levels (threshold cycle values from 31.42+/-2.06 to 38.84+/-1.47 in nasopharyngeal swabs, from 35.63+/-2.19 to 39.38+/-2.65 in plasma samples). CONCLUSION: Filtration with a leukoreduction filter can reduce H1N1 genome levels, but may not be completely sufficient for total eradication of this pathogen.

Analysis of Characteristics of Mononuclear Cells Remaining in the Leukoreduction System Chamber of Trima Accel(R) and Their Differentiation Into Dendritic Cells / 대한진단검사의학회지

BACKGROUND: We investigated the characteristics of the mononuclear cells remaining in the leukoreduction system (LRS) chambers of Trima Accel(R) in comparison with those of standard buffy coat cells, and evaluated their potential for differentiation into dendritic cells. METHODS: Twenty-six LRS chambers of Trima Accel(R) were collected after platelet pheresis from healthy adults. Flow cytometric analysis for T, B, NK, and CD14+ cells was performed and the number of CD34+ cells was counted. Differentiation and maturation into dendritic cells were induced using CD14+ cells seperated via Magnetic cell sorting (MACS(R)) Seperation (Miltenyi Biotec Inc., USA). RESULTS: Total white blood cell (WBC) count in LRS chambers was 10.8x108 (range 7.7-18.0x108). The median values (range) of proportions of each cells were CD4+ T cell 29.6% (18.7-37.6), CD8+ T cell 27.7% (19.2-40.0), B cell 5.5% (2.2-12.1), NK cell 15.7% (13.7-19.9), and CD14+ cells 12.4% (8.6-32.3) respectively. Although total WBC count was significantly higher in the buffy coat (whole blood of 400 mL) than the LRS chambers, the numbers of lymphocytes and monocytes were not statistically different. The numbers of B cells and CD4+ cells were significantly higher in the buffy coat than the LRS chambers (P<0.05). The median value (range) of CD34+ cells obtained from the LRS chambers was 0.9x10(6) (0.2-2.6x10(6)). After 7 days of cytokine-supplemented culture, the CD14+ cells were successfully differentiated into dendritic cells. CONCLUSIONS: The mononuclear cells in LRS chambers of Trima Accel(R) are an excellent alternative source of viable and functional human blood cells, which can be used for research purposes.

Prestorage leukoreduced RBCs: Evaluation for two different quadruple collection bags with an inline RBC filter / 대한수혈학회지

BACKGROUND: The Korean Red Cross(KRC) are considering the introduction of prestorage leukoreduced RBCs, as it is thought one of best way to prevent WBC-associated adverse transfusion reactions. We evaluated two different quadruple top-and-bottom collection bags, specially designed for this purpose by two manufacturers(Boin Medica Co.,Ltd and Green Cross Medical Corporation, Korea), consisted of a CPD bag, a SAG-M bag and an inline RBC filter, either RCM1 (Pall Medical, U.S.A.) or BioR 01 1N (Fresenius Hemocare, Germany) each. METHODS: Fifty one bags from Boin and sixty bags from Green Cross and consequent blood components were studied in twice over a period of two years. As an inline filter, Boin used RCM1 and Green Cross used RCM1(1st) and BioR 01 1N(2nd). They were divided into group A(filtration performed after processing of RBCs) and B(filtration performed after storage of overnight at 4 degrees C). WBC-reduced RBCs were tested for pH, plasma K+, plasma Hb and blood culture every 7 days to investigate storage time-dependent changes up to 42 days. RESULTS: The results of pH, plasma K+ and plasma Hb from CPD added whole blood, RBCs, SAG-M added RBCs and WBC-reduced RBCs did not show significant differences. We observed both of adding SAG-M solution and filtration procedure did not effect on the prestorage leukoreduced RBCs. WBCs were removed effectively by prestorage filtration. But in the case of failed units, the design or method to integrate an RBC filter may be a cause for the failure. CONCLUSION: We concluded that two different quadruple collection bags with integral RCM1 filter from Boin(2nd) and Green Cross(1st) are useful for production of prestorage leukoreduced RBCs.

The Effect of Leukoreduction Transfusion on Improving the Medical Quality / 中国医师杂志

Objective To study the effect of leukoreduction transfusion on improving medical quality. Methods The cases in our hospital which had RBC transfusion records in the past 3 years were collected and divided into two groups: leukoreduction group(LR) and non-leukoreduction group(NLR). Through statistical analysis, the items related to leukoreduction were found out and the differences in these items between the two groups were compared. Results Operation complication, non-operation complication, infectious rate, transfusion reactions and mortality rate in LR group were significantly lower than those in NLR group. The transfusion of single components-RBC had less clinical side-effect than the transfusion of complex components including RBC. Conclusions Leukoreduction transfusion can effectively improve clinical therapy and promote the medical quality.

Experimental Research of Efficacy of Oxigenated Leukodepletion Filter in Battlefield Trauma Application / 中国医师杂志

Objective To study the effect of oxigenated leukodepletion filter(OLF), in order to evaluate its potential value in battlefield trauma application.Methods 12 bags of whole blood were filtered by OLFs, while the other 12 by normal filters,some indexes including ,partial pressure of oxygen(PO 2),O 2 saturation of oxygen(SO 2),HbO 2 WBC,RBC counter and volume of RBC supernatant were measured before and after leukocyte filtration.Results PO 2,SO 2 and HbO 2 increased 57%,51% and 53% respectively after filtration by OLFs, the residual leukocyte count decreased to (3 37?0 54)?10 7 each bag,and the leukocyte clearance rate was (97 52?0 74)%.Conclusions OLF can improve the effect of blood transfusion and reduce some acute transfusion adverse reactions.Thereby,application of the OLF has binefits significantly to heal the wounded patients in battlefield trauma.

The effect of leukoreduction on the IL-2 and IL-8 leveIs in blood storage / 中国免疫学杂志

Investigate the effect of prestorage white cell filtration on the cytokines content in blood. Methods: Ten units of whole blood was divided into two parts just after blood collection:one was used as control, the other was filtrated by leukoreduction filters. All units were stored at 4℃ for 6 weeks. Immediately after filtration and every week subsequently, samples for analses of IL-2, IL-8 were obtained. Results:The amounts of IL-2 and IL-8 increased during the storage period. In filtered units, the amounts of IL-8 did not increase during the study period, IL-2 still increased but was still lower than that of unfiltered units at the end of storage. Conclusion: Many cytokines, such as IL-2 and IL-8, were generated and accumulated in blood during blood storage. Leukocyte filtration may prevent this effect to some extend. Prestorage filtration may decrease FNHTRs by preventing the accumulation of cytokines in blood.