Vascular access for autologous peripheral blood stem cells collection by large volume leukapheresis: Single center experience.

INTRODUCTION: Peripheral blood stem cell (PBSC) harvesting requires reliable and safe vascular access. In our institution, a change of practice was implemented and the central venous catheter (CVC) placement for all autologous PBSC collections was abandoned in favor of a careful evaluation of peripheral venous access (PVA) for each individual patient. The aim of this prospective study was to evaluate the rate of patients with adequate peripheral veins for autologous PBSC collection and compare patient characteristics, collection efficacy, and complication rate between patients with PVA and CVC. METHOD: Peripheral veins were assessed by the apheresis nurse team in all patients referred between January 2020 and July 2021 to autologous PBSC collection. Only in case of difficult venous access, CVC was inserted. Large volume leukapheresis (LVL) procedures, which processed ≥3 total blood volumes, were performed. RESULTS: In 65 (57%) patients PVA was used, while 49 (43%) patients required placement of short-term CVC. Peripheral venous access was successfully used significantly more often in males (69.8%) (P = 0.010), and patients with multiple myeloma (71.0%) than in patients with non-Hodgkin's lymphoma (35.9%) and Hodgkin's lymphoma patients (33.3%) (P < 0.001). There was a significant difference in the type of prior administered chemotherapy; in the patients who received cytostatics free chemotherapy, PVA was used more often (75.0%) (P = 0.007). In terms of the efficacy and safety of LVLs, there were no differences between procedures performed using PVA and CVCs. CONCLUSION: Peripheral venous access is feasible for autologous PBSC collection in more than a half of patients, in particular in those with multiple myeloma. Changes in the treatment of multiple myeloma, using new proteasome inhibitors-based and immunomodulatory agents that do not adversely affect peripheral veins, have enabled the use of PVA even at the high blood flow rates required by LVL. Peripheral venous access is not associated with safety issues or with a lesser collection efficiency, and it is cost-effective as well. Each patient referred to autologous PBSC collection needs to be evaluated individually by the experienced apheresis team for the most appropriate venous access.

The impact of cryoprotectant exposure time on post-thaw viability of autologous and allogeneic hematopoietic stem cells and leukocyte subpopulations.

Although the use of cryoprotectant dimethyl sulfoxide (DMSO) is the gold standard in cryopreservation of hematopoietic stem cells, it is well known that it has a negative effect on cell viability. The aim of this prospective study was to examine how the length of post-thaw exposure to DMSO affects the cell viability and stability of peripheral blood stem cell (PBSC) samples. Additionally, the effects of donor type and pre-cryopreservation storage time on post-thaw viability during the stability study were evaluated. In 30 autologous and 30 allogeneic PBSC samples viable CD34+, CD14+, CD19+, CD16+/56+, and CD3+ cells were determined immediately after thawing, and one-and three-hours post-thaw. Analysis of the absolute count of viable cells in thawed samples showed a significant difference between all measurement points for CD34+ (p < 0.001), CD14+ (p < 0.001), and CD19+ cells (p < 0.001). No significant differences were observed for post-thaw stability of allogeneic samples analysed between products stored before cryopreservation ≥ 24 hours (N = 20), and those stored < 24 hours (N = 10), except for viable CD3+/CD4+ cells after three hours post-thaw (p = 0.028). In conclusion, DMSO had different effects on leukocyte subpopulations in cryopre-served PBSC samples. The type of donors and the length of storage before cryopreservation did not affect the post-thaw stability of cryopreserved PBSC samples.

P31-A150†SARS-CoV-2 real time polymerase chain reaction testing of corneas from post-mortem SARS-CoV-2 positive donors.

PURPOSE: Possible transmission of SARS-CoV-2 from donors to recipients via cornea grafts is still a concern of the transplantation community. Current recommendations are to avoid corneal transplants from donors with ongoing SARS-CoV-2 infection or those recently exposed to it. During pandemic period in Croatia 21/1113; (1,9%) corneas were procured from donors positive for SARS-CoV-2 by postmortem nasopharyngeal swab tests. That tissue was discarded. Due to the lack of knowledge about the infectivity of such corneas, we started prospective study of SARS-CoV-2 presence in cornea tissue. Here we show our first results. METHODS: In the study period we had four corneas procured from two post-mortem SARS-CoV-2 positive donors. For the presence of SARS-CoV-2, analysis is performed on donor serum, hypothermic storage medium and cornea tissue lysate. Corneas were stored in hypothermic condition for 8 to 10 days, after which tissue was macerated and washed with PBS. The intracellular content was released by incubation with lysis buffer, followed by centrifugation. Next, tissue lysate, serum and hypothermic storage medium were in parallel subjected to fully automated nucleic acid isolation and RNA expression was analyzed by qRT-PCR. During isolation, RNasaP was used as internal control for successful nucleic acids isolation. RESULTS: No SARS-CoV-2 RNA was detected in the donors serum, storage medium and cornea tissue from donors who were SARS-CoV-2-positive upon tissue procurement. In nasopharyngeal swabs of post mortem positive donors cycle threshold values of viral copies were high (CT>34), indicating that there was small number of viral particles in infected donors that could have impact on negative results in tested tissue. CONCLUSION: Our data suggested that corneas may not be SARS-CoV-2 permissive if the donor was postmortem positive. Further research is required to gain more coherent insight into SARS-CoV-2 transmission via corneal transplantation.

P39-A145†Analysis of cornea donation program in croatia.

PURPOSE: Croatian Tissue and Cell Bank (CTCB) regularly monitors the effectiveness of cornea donation program on the national level. All hospitals are required to have designated tissue donation coordinators in charge of detection, family interview and tissue procurement. If hospital has cornea donation program only from donors after brain death (DBD), tissue donation coordinator can be the same as for organs. Five collection centres have cornea donation program for donors after circulatory death (DCD) with designated cornea donation coordinators. METHODS: We retrospectively analyzed all monthly reports from tissue donation coordinators in the period from May 2019 to September 2022. Additional data was collected from national organ and tissue database Croatian National Transplantation Network (NTM). RESULTS: During the analyzed period, 25.753 deaths were recorded, from which 38,6% to 54,7% of DCD and 0,6% to 1,1% of DBD donors were considered for cornea donation, depending on the hospital. Out of all deceased, 2,4% to 5,2% of patients were realized as cornea donors, 0,4 to 0,5% of which were DBD and 2 to 4,7% were DCD. Cornea donations were realized in 18,2% to 38,9% cases of all DBD donors. As SARS-CoV-2 pandemic has strated in March 2020, the cumulative number of donations declined for 26,1% in 2020 and 12,1% in 2021, compared to the pre-pandemic 2019. Moreover, CTCB received 30,5% less DCD in 2020 and 21,9% less in 2021. Despite that, we recorded increase in DBD during 2020 and 2021, for 13,3% and 44,7%, respectively. The same trend continued throughout 2022, where only until September 16,1% more DBD were received than in the whole 2019. CONCLUSION: Hospitals involved in cornea donation program record high number of deaths, however only a small proportion of which are realized for cornea donation. This is particularly pronounced in DBD donors. SARS-CoV-2 pandemic left significant impact on donation program. However, CTCB recorded higher number of DBD donors during that period. The current situation leaves plenty of room for improvement of CTCB and corresponding donation hospitals, to increase disproportionately low rate of cornea procurement in respect to the total rate of deaths and considered donors.

Variable recovery of cryopreserved hematopoietic stem cells and leukocyte subpopulations in leukapheresis products.

INTRODUCTION: Due to the expansion of cell therapy using not only haematopoietic stem cells (HSC) but also other leukocyte subpopulations, the loss of these cells in cryopreserved apheresis products needs to be evaluated. Various factors that could negatively affect post-thaw recovery, such as leukapheresis product characteristics, storage time and cryopreservation protocols have been identified. METHODS: The post-thaw recovery of HSCs, lymphocytes, NK cells and monocytes, as well as the factors that could adversely affect it were analysed in autologous and allogeneic leukapheresis products. RESULTS: The lowest post-thaw recovery was observed in autologous and allogeneic CD34+ cells, with the median of 73.7% and 68.1%, respectively. In leukocyte subpopulation, the lowest post-thaw recovery was observed for CD14+ cells, both autologous and allogeneic. The highest post-thaw recovery was observed for CD3+/CD8+ cells in autologous, and for CD19+ cells in allogeneic samples. The statistically significant difference was observed between autologous and allogeneic PBSC products for CD3+ cell recovery (P = 0.031) and CD3+/CD8+ cell recovery (P = 0.009). The evaluation of factors that could adversely affect the post-thaw recovery in autologous samples showed weak negative correlations between platelet concentration and CD3+ recovery, as well as between storage time and CD3+CD8+ recovery. In allogeneic samples, a strong negative correlation was observed only between the percentage of granulocytes and CD3+, CD3+/CD8+ and CD3+/CD4+ cell recoveries. CONCLUSION: Since various post-thaw recoveries of leukocyte subpopulations were observed, the cell therapy manufacturing centers should evaluate how their cryopreservation method and other factors affect the recovery of cell population of interest in their settings.

Changes in the incidence of transfusion reactions in hematological patients over the past 30 years.

BACKGROUND: Patients with hematological diseases are polytransfused and often immunocompromised, therefore susceptible to transfusion reactions (TR). This study aims to document the incidence of TRs in adult hematological patients and assess the effect of changes in the production of blood components and transfusion practice on their occurrence. STUDY DESIGN AND METHODS: Retrospective observational analysis of TRs reported from 1993 to 2019 was performed. For the analysis of the effect of changes on the incidence of TRs, the evaluated time was divided into two periods: the 1st period before the introduction of changes in production, when leukoreduced blood components were used only selectively, and the 2nd period, when semi-automated method of production and universal leukoreduction was introduced. RESULTS: The decrease in the incidence of TRs was observed for both red blood cell (RBC) and platelet concentrate (PC) transfusions in the 2nd period. Since platelet additive solution has been used, a further decrease in the incidence was reported. The decrease in incidence was also observed for delayed hemolytic/serological transfusion reactions and for transfusion-transmitted bacterial infections. Four cases of incorrect blood transfusions were uniquely related to the hematological patients, caused by antigen loss and transfusion ordering after ABO-incompatible hematopoietic stem cell transplantation. DISCUSSION: Our results provided evidence that the introduction of tools offered by modern transfusion medicine: universal leukodepletion, plasma replacement with additive solutions, sensitive laboratory techniques, prophylactic antigen matching policy, informatization, and automatization, decreased the incidence of TRs and improved transfusion safety.

The role of serological and molecular testing in the diagnostics and transfusion treatment of autoimmune haemolytic anaemia.

Autoimmune haemolytic anaemia (AIHA) is a rare autoimmune disease characterised by haemolysis associated with the presence of immunoglobulins and/or components of the complement system on red blood cells (RBCs). It is classified into warm or cold antibody-mediated AIHA according to the temperature at which autoantibodies bind optimally to RBCs. Clinicians should be familiar with the procedural tests used for a complete laboratory investigation of AIHA. Good collaboration between clinicians and laboratory specialists with correct sample handling and an exact diagnostic work-up is extremely important for the correct classification and proper therapeutic management of AIHA. Specialised serological test procedures are very complex. Problems with serological testing may be excluded with the molecular testing, which has now become a gold standard to predict a patient's phenotype in order to secure the right antigen-matched blood for AIHA patients. More recently, genotyping has been used instead of serological typing and complex adsorption tests. This paper offers a description of various tests for differentiating between types of AIHA. A diagnostic algorithm and the problems of laboratory investigation are also presented, and an application of molecular methods for the blood group typing in AIHA is elaborated.

36†Cornea microbiology contamination rate depending on post mortem time, retrospective analysis at croatian tissue and cell bank, UHC Zagreb.

BACKGROUND: Corneas procured post mortem are at risk of microbiology contamination, therefore decontamination procedures before storage, aseptic techniques during processing and antimicrobials used in the storage medium are routinely used. Despite that, corneas are discarded due to microbiology contamination. According to professional guidelines, corneas can be procured preferably within 24 hours after cardiac arrest but up to 48 hours. Our goal was to evaluate the risk of contamination depending on the post mortem time and the spectrum of microbes isolated. METHODS: Corneas were decontaminated before procurement using 0,5% povidone iodine and tobramycin, stored in the organ culture medium and microbiologically tested after four to seven days of storage. Ten millilitres of cornea preservation medium were inoculated in two blood bottles (aerobic, anaerobic/fungi, Biomeriex) and incubated for seven days.Microbiology testing results in the period of four years (2016-2020) were retrospectively analysed. Corneas were divided in four groups depending on the duration of post mortem interval: group A post mortem interval < 8 h, group B post mortem ranging from 8 to 16 h, group C post mortem ranging from 16 to 24 h and group D post mortem > 24 h. Contamination rate and spectrum of isolated microorganisms in all four groups were analysed. RESULTS: 1426 of 2019 procured corneas were stored in organ culture and microbiologically tested. 65/1426 of tested corneas were contaminated (4,6%). In total, 28 strains of bacteria and fungi were isolated.Contamination rate of post mortem groups are as following: group A 3,1% (14/455), group B 4,1% (23/561), group C 6,7% (27/402) and group D 12,5% (1/8).In the group A bacteria family Staphylococcaceae, Moraxellaceae, Morganellaceae were predominately isolated (64,3%). In the group B fungi Saccharomycetaceae, bacteria Moraxellacea, Staphylococcaceae, Morganellaceae and Enterococcaceae were predominately isolated (78,1%). In the group C, bacteria family Enterococcaceae, Moraxellaceae and fungi Saccharomycetaceae are most often isolated (70,3%). In the group D bacteria family Enterobacteriaceae was isolated (100%). CONCLUSION: Organ culture allows detection and discard of microbiology contaminated corneas. Our results show higher microbiology contamination rate for corneas with longer post mortem intervals, suggesting these contaminations can be rather related to donor post mortem changes and contamination than previous infection. In order to keep the best quality and safety of the donor cornea, all efforts should be directed in disinfection of the cornea and keeping post mortem interval shorter.

Cost Analysis of Transfusion Therapy in Coronary Artery Surgery.

BACKGROUND: In patients undergoing coronary artery bypass grafting (CABG), wide variability in transfusion rate (7.8% to 92.8%) raises the question of the amount of unnecessary transfusions. The aim of the study was (1) to identify CABG patients at low risk of bleeding to whom transfusion treatment should be avoided and (2) to calculate the amount of possible cost savings that would be achieved by avoiding transfusion in low bleeding risk patients. METHODS: This retrospective observational study enrolled patients undergoing isolated elective CABG from January 2010 to January 2018. Patients were divided with respect to the presence of excessive bleeding and transfusion costs were compared between the two groups. Predictors for postoperative excessive bleeding were defined and multivariable logistic regression analysis and risk modeling were performed. The use of a model to predict patients at low risk of bleeding allowed for the estimation of transfusion cost savings assuming the patients who were found to be at low risk of bleeding should not be transfused. RESULTS: A total of 1,426 patients were enrolled in the analysis. Of those, 28.3% had excessive postoperative bleeding. The multivariate logistic regression analysis model was developed to identify/predict patients without excessive bleeding (receiver operating characteristic curve analysis, area under the curve 72.3%, p < 0.001). When applied to the existing database, the use of the developed model identifying patients at low risk of bleeding may result in a 39.1% reduction of transfusions. Specifically, cost savings would be 48.2% for packed red blood cells, 38.9% for fresh frozen plasma, 10.9% for platelets concentrate, and 17.9% for fibrinogen concentrate. CONCLUSION: The clinical and economic burdens associated with unnecessary transfusions are significant. Avoiding transfusion in CABG patients found to be at low risk of bleeding may result in significant reduction of transfusion rate and transfusion-associated costs.

Performance prediction algorithm for autologous PBSC collection in adults and pediatric patients using large volume leukapheresis.

BACKGROUND AND OBJECTIVES: The number of CD34+ cells collected in apheresis procedures depends mainly on the collection efficiency of the device and the blood volume processed. Large volume leukapheresis (LVL) can improve CD34+ cell yield and has previously been investigated using the COBE Spectra device (Terumo BCT, USA). MATERIALS AND METHODS: This was a retrospective analysis of LVL performance in patients undergoing continuous mononuclear cell collection (CMNC) using the new Spectra Optia apheresis system (Terumo BCT, USA) at the University Hospital Center, Zagreb, from March 2016 to September 2016. CD34+ cell yield predictability, determined using a customized algorithm, was also assessed. RESULTS: In total, 67 procedures performed in 46 adults and 14 performed in 11 children were included in the analysis. In adults, 30 (65.2%) patients successfully reached their target preapheresis CD34+ cell count on day 1, with a median (interquartile range [IQR]) CD34+ collected cell dose of 4.8 × 106 /kg (2.3-10.6 × 106 /kg). In the pediatric group, 81.8% successfully collected the target CD34+ cell dose on the first day, with a median (IQR) CD34+ collected cell dose of 11.1 × 106 /kg (3.2-16.3 × 106 /kg). The customized algorithm showed a strong and significant linear correlation with actual CD34+ cell dose (P < 0.0001). CONCLUSION: The results of this study support the use of LVL and the customized prediction algorithm in apheresis procedures. The ability to tailor the procedure to meet the needs of the individual patient may help to minimize the blood volume processed, shorten the duration, reduce the volume of infused anticoagulants, and improve patient comfort.

Haemolysis, pure red cell aplasia and red cell antibody formation associated with major and bidirectional ABO incompatible haematopoietic stem cell transplantation.

BACKGROUND: Acute and delayed haemolysis, alloimmunisation and pure red cell aplasia (PRCA) are potential complications after ABO incompatible haematopoietic stem cell transplantation (HSCT). The aims of this study were to investigate acute and delayed red blood cell (RBC) antibody-associated complications, including haemolysis, PRCA and alloimmunisation in major and bidirectional ABO incompatible HSCT. MATERIALS AND METHODS: We retrospectively examined the transplant courses of 36 recipients of bone marrow or peripheral blood stem cells from ABO incompatible donors and evaluated the current practice of performing plasmapheresis in patients with higher isoagglutinin titres. We investigated the role of ABO incompatibility in haematopoietic recovery, transfusion requirements, alloimmunisation and PRCA. RESULTS: Laboratory signs of acute haemolysis were noted in five (14%) patients, one (3%) of whom had clinically overt haemolysis. Patients with haemolysis had IgM titres ≥1:8 and received >16 mL of RBC in the HSCT. In patients with higher titres, plasmapheresis performed prior to the transplant prevented acute haemolysis. Delayed haemolysis was not recorded in the follow up. Haematopoietic recovery and transfusion requirements did not differ notably between patients with and without haemolysis. De novo RBC antibodies were detected in two (5.5%) patients after HSCT, and PRCA was noted in one (3%) patient. DISCUSSION: Carried out with adequate graft processing, plasmapheresis and blood component support, haemolysis is not a common complication after HSCT. Our results confirm that the occurrence of haemolysis depends on larger RBC volumes and higher isoagglutinin titres. Despite the reduction of patients' isoagglutinin titres by plasmapheresis, we still noted a critical combination for the development of laboratory signs of haemolysis (IgM titre ≥1:8 and RBC volume >16 mL). De novo immunisation to RBC antigens and PRCA are rare events following ABO incompatible HSCT.

Large volume leukapheresis is efficient and safe even in small children up to 15 kg body weight.

BACKGROUND: The collection of peripheral blood stem cells, although now a routine procedure, is still a challenge in low body weight children because of specific technical and clinical issues. For paediatric patients it is crucial to obtain an adequate number of CD34+ cells with the minimum number of procedures: this can be done using large volume leukapheresis (LVL). MATERIALS AND METHODS: We analysed the efficacy and safety of 54 autologous LVL performed in 50 children (33 [66%] males and 17 [34%] females), median age 2 years (range, 1-5) and median body weight 12 kg (range, 6-15). The procedures were performed with a COBE Spectra previously primed with red blood cells; ACD-A solution and heparin were used as anticoagulants. RESULTS: The target CD34+ cell dose (≥5×10/kg body weight) were collected with one LVL in 46 (92%) patients, while four (8%) patients needed another procedure. All our LVL were well tolerated. Side effects were observed in five (9.2%) patients and one procedure had to be discontinued because of catheter-related haemorrhage. The platelet count decreased significantly (p<0.001) after each procedure but without bleeding or need for transfusion support. DISCUSSION: Our experience confirms that LVL is efficient and safe even in small children, if the procedure is adjusted considering the weight and age of child. The most important factors are good venous access, adequate preparation of the child's electrolyte status, and surroundings in which the small child as well as parents feel comfortable, and can tolerate the procedure better. Although a median platelet loss of 50% can be expected, LVL is safe and reduces the overall number of procedures required. It can be recommended for peripheral blood stem cell collection even in small body weight children with malignant diseases, particularly those who mobilise low numbers of CD34+ cells.

[Extracorporeal photopheresis in treatment of chronic graft versus host disease]. / Ekstrakorporalna fotofereza u lijecenju kronicne reakcije transplantata protiv primatelja.

Extracorporeal photopheresis (ECP) is an immunomodulatory therapy which has been used in the treatment of chronic GVHD (cGVHD). ECP involves separation of the mononuclear cells with leukapheresis, followed by ex vivo administration of 8-methoxypsoralen and UV-A radiation and reinfusion to the patient. Aim of the study was to evaluate clinical and immunomodulatory effect of ECP procedures in patients with cGVHD. We analyzed 341 ECP procedures performed in 7 patients with cGVHD; median ECP per patient was 37 (range 13-131). All patients suffered from skin changes in combination with impaired joint mobility and symptoms of oral disease. ECP procedures were performed for two consecutive days: in initial phase weekly, followed by every two weeks and than monthly according to clinical response. Median of ECP treatment duration was 10 months (range 2-58). The effect of ECP in patients with cGVHD with skin andjoint involvement was mostly beneficial: 6 patients experienced either improvement or stabilization in skin changes and joint mobility. In 2 patients who suffered from oral disease, the total recovery was observed. Clinical response was typically delayed until 2 to 3 months, and reduction in glucocorticoid dose was observed. Adverse reactions were observed in 4.9% procedures. In patients who responded to ECP treatment, CD4+/CD8+ ratio and number of NK cells were normalized. ECP proved to be an efficient and safe procedure that may be recommended for patients with cGVHD who do not respond to conventional therapy.

[Mesenchymal stem cells: immunomodulatory properties and clinical application]. / Mezenhimske maticne stanice: imunomodulatorne znacajke i klinicka primjena.

Mesenchymal stem cells (MSCs) are multipotent nonhematopoietic cells that were first identified in bone marrow. Clinical interest for MSCs was initiated by the observation that MSCs are immunoprivileged cells that display immunomodulatory properties in vitro. Ex vivo expanded MSCs have therefore become a new type of cellular therapy in development with a wide range of potential clinical applications. So far many clinical studies confirmed safety of their use and showed that infused MSCs suppress graft versus host disease, support engraftment of transplanted allogeneic hematopoietic stem cells and stimulate growth in patients with osteogenesis imperfecta. Although underlying immunomodulatory mechanisms of action are not completely understood, potential benefit of MSC therapy justifies its clinical use in a broad range of disorders. In this report we give historical overview of MSC discovery and current scientific and clinical achievements in this field. Better insight into their biological properties and mechanisms of action are needed.

Large volume leukapheresis: Efficacy and safety of processing patient's total blood volume six times.

Large-volume leukapheresis (LVL) differs from standard leukapheresis by increased blood flow and an altered anticoagulation regimen. An open issue is to what degree a further increase in processed blood volume is reasonable in terms of higher yields and safety. In 30 LVL performed in patients with hematologic malignancies, 6 total blood volumes were processed. LVL resulted in a higher CD34+ cell yield without a change in graft quality. Although a marked platelet decrease can be expected, LVL is safe and can be recommended as the standard procedure for patients who mobilize low numbers of CD34+ cells and when high number of CD34+ cells are required.

[Collection of hematopoietic progenitor cells from healthy donors]. / Prikupljanje krvotvornih maticnih stanica zdravih davatelja.

Allogeneic hematopoietic progenitor cell (HPC) transplantation is an established therapy for many hematologic disorders. HPCs may be collected from bone marrow, peripheral blood, or umbilical cord blood. In order to minimize the risk for healthy HPC donors, thorough investigation is required before donation. The donor work-up should include medical history, physical examination, ECG, chest x-ray, blood count, coagulation screening, and testing for infectious disease markers. Donors should be fully informed on the donation procedure and sign an informed consent for donation. HPCs are traditionally collected from bone marrow with the donor in general anesthesia. The procedure includes multiple bone marrow aspirates from pelvic bones and at least overnight hospital stay. Although marrow donation is generally safe and well tolerated, minor complications like pain at the collection site, fatigue and pain on walking or sitting may occur in a relatively small proportion of donors (6%-20%). Major and life-threatening complications such as anesthesia-related events, mechanical injury to the bone, sacroiliac joint and sciatic nerve following marrow donation are relatively rare, being estimated to 0.1%-0.3% of cases. In the last decade, peripheral blood progenitor cells (PBPC) have become an increasingly used altemative to bone marrow. PBPC transplantation offers faster hematopoietic recovery and lower early transplant-related morbidity and mortality. The incidence of acute graft vs. host disease (GvHD) is no greater than in bone marrow transplants. However, there is evidence for increased chronic GvHD, which is in part related to the higher number of T and NK cells that are collected with PBPC and re-infused to the patient. Recombinant human granulocyte colony-stimulating factor (G-CSF) is used to mobilize PBPCs for collection by leukapheresis. Leukapheresis is usually perfomed after 4 to 5 days of G-CSF subcutaneous administration at a dose of 10 mg/kg b.w. Vascular access for apheresis may be accomplished by use of apheresis needle in antecubital vein. Placement of a double-lumen central apheresis catheter is rarely required in healthy donors. Citrate is the most commonly used anticoagulant for apheresis. One to three leukapheresis procedures are required to collect adequate graft. There is an interindividual variation in progenitor cell mobilization among healthy donors, with a subset of donors that do not exhibit effective CD34+ cell mobilization. Donor age and G-CSF schedule are the factors that significantly affect PBPC mobilization and collection in healthy donors. Procedures for mobilization and collection of PBPC from healthy donors are generally well tolerated. Common adverse reactions of G-CSF application include bone pain, myalgia, headache and fatigue. Beside these mild side effects, moderate to life-threatening complications are sporadically observed. Spontaneous splenic rupture, acute lung injury, acute iritis, severe pyogenic infections, and anaphylactoid reactions were reported in healthy donors after G-CSF administration. Adverse effects of apheresis for PBPC collection are the same as for other apheresis procedure and include complications related to venous access and citrate toxicity. Leukapheresis typically results in a lower platelet count, an effect that is exacerbated by the use of G-CSF, which has been documented to cause mild, reversible thrombocytopenia. Fewer side effects were noted in pediatric donors compared to adult donors. PBPC collection in pediatric donors is safe and desired PBPC yields are easily achieved. Theoretical concerns exist about the potentially increasing long-term risk of leukemia after G-CSF administration in healthy donors. Recently, a report of AML developing in a 62-year-old female donor 14 months after G-CSF-primed PBPC donation has been published. Whether G-CSF therapy contributed to the development of this cancer is unknown, but future studies should carefully follow the donors and report any similar event. According to currently available evidence, the risk of major late toxicities secondary to administration of G-CSF is minimal.

[Cord blood banking]. / Banke krvi iz pupkovine.

Transplantation of cord blood stem cells is a new and rapidly developing area. It has been used as a treatment for many diseases such as hematologic malignancies, primary immune deficiencies and metabolic diseases. Recently, stem cells have been used in regenerative medicine, particularly in neurodegenerative and cardiovascular diseases. For these reasons interest has been growing in banking cord blood. To be able to find an acceptable donor for any recipient in need, it is necessary to have on stock a great diversity of cells with different genetic types from different populations. Networks of banks and registries have been created around the world in order to share and exchange transplants. Public banks organize collection for altruistic donor of cord blood for unrelated hematopoietic stem cell transplantation and for directed donation in families at risk. But there are increasing numbers of families that are requesting storage of cord blood for possible future therapeutic use in the family. Establishment of cord blood banks has raised a number of important scientific, legal, ethical and political issues, which are discussed in this paper.

[Peripheral blood hematopoietic stem cell collection]. / Prikupljanje krvotvornih maticnih stanica iz periferne krvi.

Summary. Peripheral blood hematopoietic stem cells (PBSC) have numerous advatages in comparison with traditionally used bone marrow. PBSC collection by leukapheresis procedure is simpler and better tolerated than bone marrow harvest. PBCS are mobilized by myelosupressive chemotherapy or/and hematopoietic growth factors. Leukapheresis product contains PBSC along with lineage commited progenitors and precursors which contribute to faster hematopoietic recovery. In "poor mobilizers" options are large-volume leukapheresis (LVL) procedure or second generation of mobilising agents (pegfilgrastim, CXCR4 receptor antagonists). Total blood volume is processed 2-3 times in standard procedure compared to more than 3 times in LVL. LVL yields significantly higher numbers of CD34+ cells. Adverse effects of leukapheresis are electrolyte disbalance (hypocalcemia) caused by citrat administration and risk of bleeding due to trobocytopenia and heparin administration. PBSC collection and product quality control are regulated by national and international standards and recommendations.

Three new high-prevalence antigens in the Cromer blood group system.

BACKGROUND: The Cromer blood group system consists of nine high-prevalence and three low-prevalence antigens carried on decay-accelerating factor (DAF). This report describes three new Cromer high-prevalence antigens, named ZENA, CROV, and CRAM. STUDY DESIGN AND METHODS: Sequence analyses were performed on DNA from three probands whose serum samples each contained an alloantibody to a high-prevalence antigen in the Cromer blood group system. Polymerase chain reaction-restriction fragment length polymorphism analysis to detect the mutation encoding the CROV- phenotype was performed on 100 Croatian donors. To map the respective epitopes, DAF deletion mutants were tested by immunoblotting with eluates containing the antibodies. RESULTS: In each proband, sequence analysis revealed a single-nucleotide substitution in DAF: ZENA, 726T>G mutation, predicted change His242Gln; CROV, 466G>A mutation, predicted change Glu156Lys; and CRAM, 740A>G mutation, predicted change Gln247Arg. By analysis of DAF deletion mutants, the CROV antigenic determinant mapped to the complement control protein (CCP) domain 2, which is encoded by exon 3, whereas ZENA and CRAM mapped to CCP4, which is encoded by exon 6. CONCLUSION: This study describes three novel high-prevalence antigens in the Cromer blood group system each characterized by a predicted single-amino-acid substitution. The antigens have been assigned the following International Society of Blood Transfusion (ISBT) numbers: ZENA is CROM13, CROV is CROM14, and CRAM is CROM15.