Correlation between serum cytokine levels and the effect of allogeneic serum-based eye drops.

INTRODUCTION: The Institute of Hematology and Transfusion Medicine (IHTM) in Warsaw has produced autologous serum eye drops (ASEDs) for the treatment of Dry Eye Syndrome (DES) since 1991. In 2019, IHTM introduced allogeneic tears (alloSEDs) for patients on long-term treatment. MATERIALS AND METHODS: 114 patients who applied alloSEDs were included in the study.They were asked to complete the OSDI questionnaire before and after using ASEDs and 100 units of alloSEDs drops from each donation. The OSDI index rates DES severity (0 no symptoms; 100 severe). We also compared the content of IL-1ß, IL-2, IL- 6, IL-10 and VEGF in ASEDs (38 samples) and alloSEDs (15 serum samples). The study data covered the 2019-2022 period. RESULTS: 114 patients participated in the study. We compared the the effectiveness of ASEDs and alloSEDs. The average, OSDI dropped from 68.42 ± 5,86 (before application) to 51.05 ± 19,06 (after application). Data from the questionnaires (prepared at IHTM) completed and returned (41/114) present the most common indications for the use of serum drops, including DES with no underlying disease, DES secondary to GvHD (Graft versus Host Disease), Sjögren's Syndrome (SS). The study reported higher cytokine levels associated with disease entities such as SS. After application of drops with high cytokine levels, patients reported adverse reactions such as sand under the eyelids, impaired visual acuity, and worse eye lubrication. CONCLUSIONS: AlloSEDs with acceptably low values of pro-inflammatory cytokines and sufficiently high levels of VEGF growth factor may contribute to alleviation of inflammatory eye symptoms.

Introduction of EUBIS standards to upgrade inspections of blood establishments to improve the safety of blood transfusion: The Polish experience.

BACKGROUND AND OBJECTIVES: The competent authority (CA) responsible for external inspections of Polish blood establishments (BEs) and supervision of the quality system is the Institute of Haematology and Transfusion Medicine (IHTM). Before the implementation of the European Blood Inspection System (EuBIS) classification of non-compliance, the IHTM inspections were conducted according to national guidelines and the non-compliance-related recommendations were based on the inspectors' own experience and interpretation of the observed problems. Since 2009, IHTM inspections were already performed according to EuBIS guidelines. The study assessed the impact of the EuBIS classification on the IHTM recommendations. We assumed that the implementation of consistent assessment criteria contributed to the upgrading of the quality of BE inspections. MATERIALS AND METHODS: BE-inspection protocols; 30 from 2009 to 2010 and 61 from 2016 to 2019. Non-compliance-related recommendations were classified according to the seriousness of non-compliances (critical, major, other significant, and observation) and also to the area of BE activity (documentation, organisation of work, qualification and validation, pathway from donor qualification to blood component-issue, quality control of blood components, adverse events and reactions). RESULTS: The recommendations mostly referred to document-keeping and work organisation and were distributed as follows: 2009-2 critical (others unclassified), 2010-1-13 major, 4-25 other significant and 1-7 suggestions, 2016-2019-3-9 critical, 90-196 major, and 157-297 other significant as well as 14-22 suggestions. CONCLUSION: Polish BEs still require: integrated document management, analysis of IHTM recommendations, implementation of corrective and preventive measures and personnel training in identifying similar non-compliances in other procedures.

Molecular and serological infection marker screening in blood donors indicates high endemicity of hepatitis E virus in Poland.

BACKGROUND: Until now, markers of hepatitis E virus (HEV) infection have not been studied in blood donors throughout Poland, and no acute case of HEV infection has been closely documented or confirmed by HEV RNA detection. The prevalence of HEV infection markers, including HEV RNA in Polish blood donors and virus genotypes was investigated. STUDY DESIGN AND METHODS: In total, 12,664 individual donations from 22 Polish blood transfusion centers were tested for HEV RNA by transcription-mediated amplification. In addition, 3079 first-time donors sampled throughout Poland also were screened for antibodies to HEV. HEV RNA and immunoglobulin M-positive donations were confirmed using real-time reverse transcription-polymerase chain reaction and Western blotting, respectively. RESULTS: Ten donors were identified as RNA initial reactive (one of 1266 donors), and six (one of 2109) were identified as repeat reactive and confirmed by real-time reverse transcription-polymerase chain reaction or seroconversion. Sequence analysis identified HEV Genotype 3c in one donor and Genotype 3i in two others. On average, 43.5% of donors were immunoglobulin G-positive. Immunoglobulin G seroprevalence ranged from 22.7% to 60.8% in group ages 18 to 27 years and 48 to 57 years, respectively and differed between administrative regions from 28.9% in Podlasie to 61.3% in Wielkopolska. Thirty-nine of the donors were immunoglobulin M-positive, and seven donors were IgM positive only (0.2%). Of 37 immunoglobulin M-reactive samples tested by Western blot, 24 (64.9%) were confirmed. CONCLUSIONS: The current results indicate a high level of HEV endemicity throughout Poland compared with other countries. There is an urgent need to consider the protection of recipients of blood components against transfusion-transmitted HEV infection.

Methods of freezing cord blood hematopoietic stem cells.

BACKGROUND: Cord blood (CB) is a valuable source of hematopoietic stem cells (HSCs). Extended storage of CB is possible provided that validated cryopreservation procedures are used. The study objective was to determine optimal methods of CB cryopreservation. STUDY DESIGN AND METHODS: In the study we 1) compared the effect of two-step cryopreservation and controlled-rate freezing method on the postthaw quality of CB (Study A) and 2) evaluated the postthaw quality of HSC fractions isolated from CB with various methods and frozen with controlled-rate freezing method (Study B). The same cryoprotectant mixture was used for 20 CB units (Study A) and 122 CB units (Study B). RESULTS: In Study A, 13.79 × 10(8) and 13.29 × 10(8) initial white blood cell (WBC) counts decreased to 6.38 × 10(8) and 6.02 × 10(8) after thaw for the two methods, respectively. The mononuclear cell (MNC) counts decreased from 5.90 × 10(8) to 3.71 × 10(8) and from 5.64 × 10(8) to 3.47 × 10(8) dependent on the method. MNC viability decreased from 99.0% to 97.4% for the former and from 98.5% to 97.2% for the latter method. The differences were insignificant. In Study B, postthaw WBC recovery in HSC fractions was 74.4% to 103.5%, MNC recovery 106.4% to 118.5%, CD34+ cell recovery 102.5% to 150.2%, and MNC viability 94.1% to 97.4%. CONCLUSION: Neither the cryopreservation procedure nor the freezing of isolated HSCs affected product quality, which may indicate that various freezing methods can be used for cell banking provided the they follow recommendations of good manufacturing practice and Directive 2004/33/EC.

Identification and follow-up of pregnant women with platelet-type human platelet antigen (HPA)-1bb alloimmunized with fetal HPA-1a.

INTRODUCTION: Pregnant women negative for human platelet antigen 1a (HPA-1a) are at risk of alloimmunization with fetal HPA-1a antigen inherited from the father, and their offspring may develop fetal and neonatal alloimmune thrombocytopenia (FNAIT). The aim of this study was to analyze the frequency of HPA-1a alloimmunization in pregnant Polish women, the feasibility of using maternal platelets for intrauterine transfusions in women subjected to diagnostic fetal blood sampling (FBS) and to discuss potential consequences of alloimmunization. MATERIAL AND METHODS: Fifteen thousand two hundred and four pregnant women were typed for HPA-1a; HPA-1a negative were screened for anti-HPA-1a. Alloimmunized women received specialist perinatology care; some of them were subjected to FBS, followed by transfusion of HPA-1a negative platelet concentrates (PC) prepared from maternal blood. RESULTS: Three hundred seventy-three (2.5%) women were HPA-1a negative, and 32 (8.6%) tested positively for anti-HPA-1a. Antibodies were detected in 22 women during pregnancy. Diagnostic FBS followed by PC transfusion was performed in 14 woman, who were platelet donors for their 16 unborn babies. Blood donations were tolerated well by the patients, and also intrauterine platelet transfusions were uneventful. Pharmacotherapy with intravenous immunoglobulins was implemented in 11/22 patients. CONCLUSIONS: HPA-1a negative women (ca. 2.5% of all pregnant patients) are at risk of alloimmunization with HPA-1a antigen and developing FNAIT. Alloimmunized women can be donors of platelets for their offspring providing removal of antibodies from PC. Owing to potential complications, special care should be taken if an alloimmunized woman was qualified as a blood or stem cell recipient.

Quality control of riboflavin-treated platelet concentrates using Mirasol® PRT system: Polish experience.

BACKGROUND: The quality of platelet concentrates (PCs) is affected by preparation, storage, the type of container, and pathogen reduction technology (PRT). The Mirasol® Pathogen Reduction Technology (PRT) system (Terumo BCT Inc., Lakewood, USA), which uses riboflavin and ultraviolet (UV) light, has recently been proven effective against bacteria, viruses, parasites, and leukocytes. OBJECTIVES: The aim of the study was to evaluate the effect of the Mirasol® PRT system, based on riboflavin and UV light exposure, on the most common in vitro platelet quality parameters of PCs prepared from whole blood-derived buffy coats. MATERIAL AND METHODS: The study included 15 trials (n = 15). For each trial, 2 PCs were used: 1 for treatment with the Mirasol® PRT system (M) and 1 for a control (C). In the M group, PCs were illuminated. In the C group, saline solution was added. PCs from groups M and C were stored at 20-24°C, with agitation. Samples were collected on days 1, 3 and 5 to determine platelet concentration, total platelet count/unit, mean platelet volume (MPV), power of hydrogen (pH), glucose and beta-thromboglobulin concentration (BTG), hypotonic shock response (HSR), aggregation, CD42b and CD62P expression, pCO2, and pO2. RESULTS: No significant differences in HSR or CD42b expression were observed between groups M and C. All pH values were stable during the whole storage period (7.1-7.5). On storage day 1, CD62P expression in group C was significantly higher than in group M. In the Mirasol® group, significantly higher glucose consumption was noted on storage days 3 and 5. On day 5, a 2-3-fold increase in BTG was observed in both groups as compared to day 1; on day 5, BTG concentration was 32% higher in group M than in group C. On all storage days, pCO2 was comparable in groups M and C; lower pO2 values were reported for group M. CONCLUSIONS: In vitro results demonstrated that pH, HSR, aggregation, CD42b antigen expression, and MPV and platelet count parameters were comparable in groups M and C.

Study of CD69 antigen expression and integrity of leukocyte cellular membrane in stored platelet concentrates following irradiation and treatment with Mirasol® PRT System.

BACKGROUND: Leukocytes in transfused blood components, particularly residual lymphocytes, have been shown to contribute to the occurrence of various adverse reactions. One of the most severe is transfusionassociated graft versus host disease (TA-GvHD) following transfusion of blood components contaminated with immunocompetent T lymphocytes. Irradiation is a routine method for protection against TA-GvHD. According to the literature, some pathogen reduction methods have also been proven effective for the inactivation of T lymphocytes, and so they may be considered as an alternative to irradiation. OBJECTIVES: Comparison of CD69 antigen expression and the integrity of the leukocyte cellular membrane in stored platelet concentrates (PCs) following irradiation with the Gammacell 3000 Elan (Nordion Inc., Ottawa, Canada) and treatment with the Mirasol® Pathogen Reduction Technology (PRT) System (Terumo BCT, Lakewood, USA). MATERIAL AND METHODS: The study included seven experiments. For each experiment we used 3 PCs, for Mirasol® PRT System treatment (M), for Gammacell 3000 Elan irradiation (R), and for the control (C). 7-amino-actinomycin D (7-AAD, Becton Dickinson, Franklin Lakes, USA) permeability was used to determine lymphocyte viability while CD69 antigen expression was the marker of lymphocyte activation. Analyses of 7-AAD and CD69 antigen expression were performed in a FACS Canto I flow cytometer (Becton Dickinson, USA). RESULTS: During 6 storage days, viable lymphocyte count decreased to 28% (p = 0.001) in the Mirasol® PRT System treated PCs and to 65% (p = 0.004) in the irradiated PCs. A statistically significant increase in CD69 expression in the irradiated PCs was observed; 1.3-fold on day 3 and 1.5-fold on day 6. In the Mirasol ® PRT System treated PCs, no statistically significant increase was observed. CONCLUSIONS: The in vitro results suggest that the Mirasol® PRT System is as effective as irradiation due to donor leukocyte inactivation capacity.