Beitrage zur Infusionstherapie und Transfusionsmedizin = Contributions to infusion therapy and transfusion medicine最新文献

Selection of CD34+ cells for autologous transplantation is increasingly being used to reduce potential tumor cell contamination of the autograft. Haematopoietic reconstitution in 40 patients after transplantation of CD34(+)-selected versus non-selected G-CSF-mobilized PBPC was compared and was almost identical in the two groups of patients. Delayed platelet engraftment was only observed in patients transplanted with a CD34+ cell dose of < 2.5 x 10(6)/kg body weight. It has to be shown whether the positive selection of CD34+ cells will improve the disease free survival after autologous PBPC transplantation.

Common features of all myeloproliferative diseases (CMPE) are a markedly increased number of platelets and restrictions in the subjective feeling of the patients, comprising symptoms like nausea, headache, sensory deficits and transient paresis. Meanwhile, it has been shown that platelet pheresis (mTD) does not only reduce platelet counts sufficiently but also results in an impressive relief of the subjective complaints. To test the part that rheological mechanisms play hereby, relevant rheological parameters in the blood of 22 CMPE patients and of 8 healthy platelet donors as controls were analyzed before and after treatment with an AS-104 cell separator. Concerning the viscosity of whole blood und the filterability of erythrocytes, there was seen neither a difference between the patients and the controls, nor before and after mTD. As a result of treatment of patients, the moderately raised plasma viscosity was normalized, whereas the aggregation of erythrocytes was significantly lowered. It is concluded that there is an influence of hemorheological conditions on the patient's subjective feeling. These would originate in neurological dysfunctions caused by CMPE-induced restrictions in cortical microcirculation. The erythrocyte aggregation, lowered by mTD, would cause an improvement of microcirculatory fluidity and would, in consequence, abolish the neurological symptoms.

The DiaMed-ID (D-ID) gel system is known to be a very sensitive and specific method for the detection of red cell antibodies. In various cases, we failed to find an antibody in the eluate in which red-cell-bound antibodies (IgG) where proven by a positive direct antiglobulin test (DAT). SPRCA (Capture-R, Ready Screen, Immucor; C-R) seems also to be very sensitive, in part due to the antihuman, IgG-coated indicator cells. Therefore, we compared 39 acid eluates from patients who had a positive DAT (monospecific rabbit antihuman IgG) both in the D-ID and in the C-R system. Patients (19 female, 20 male; mean age 62 years) were suspected either to have an autoimmune hemolytic anemia or an alloantibody. Identification of the antibodies was done with the system's own panel cells. Agglutination strength was scored from 1 to 4. Quantification of selected eluates was performed by titration, using the same cells in both systems. From 39 eluates, 31 were positive in the C-R and 25 in the D-ID. Nine eluates were negative in the C-R and 14 in the D-ID. If only eluates with a DAT reaction strength of 2 or lower were considered, obviously more negative results were found with the D-ID technique (p < 0.027) than with the C-R technique. In all eluates the degree of test reaction was stronger in the C-R system. However, titration endpoints of 6 quantified antibodies did not differ significantly. In 2 patients with slightly positive DAT, antibody typing was negative or not clear in the serum. In the corresponding eluates, an anti-K and an anti-JKa could be identified only by the C-R technique. In such instances we recommend to use the C-R technique to prevent transfusion complications.

A group of 28 children was investigated after hematopoietic stem cell transplantation for evidence of hepatitis G virus (HGV) infection. HGV RNA was detected in 14 of the 28 patients (50%) and persisted in 9 of 11 patients with follow-up samples for up to 32 months. Whereas thrombopoiesis was delayed in 2 of the 14 HGV-RNA-negative patients (14.3%), 6 out of 14 (42.9%) patients in the HGV-RNA-positive group had a delayed thrombopoiesis and 2 of the latter group had to be retransplanted because of complete graft failure. These were the only cases with hepatitis C virus (HCV) coinfection. Significant liver diseases were also found only in these 2 patients with HGV and HCV coinfection. These results suggest that HGV infection may significantly influence the engraftment in patients after hematopoietic stem cell transplantation, particularly if coinfection with HCV occurs.

The influence of 6 different cooling rates (1.4, 5, 10, 15, 20, 160 K/min) and 5 different compositions of the suspensions to be frozen (% DMSO/% HES: 10/0, 7.5/2.5, 5/6, 2.5/7.5, 0/10) were investigated in 30 aliquots from each of 12 peripheral blood progenitor cells (PBSC) products which had been obtained by leukapheresis from 11 patients with hematological malignancies and solid tumors and 1 healthy donor treated with 5-24 micrograms/kg body weight/day granulocyte colony stimulating factor (G-CSF) over 5 days. The MNC concentration in the products obtained amounted to 4.51 +/- 1.55 x 10(7)/ml (mean +/- SEM), range: 2.10-7.65 x 10(7)/ml). For freezing, cooling rates were generated by means of a liquid nitrogen(LN2)-operated, computer-controlled freezer, a mechanical -80 degrees C freezer, in the vapor phase over LN2, and by submerging into LN2. The statistical analysis of the results clearly indicates that optimum results compared with the prefreeze values for numerical recovery (80.6 +/- 20.1%, Coulter counter), viability (membrane integrity by Trypan blue exclusion 91.6 +/- 4.1%), and colony-forming potential (56.2 +/- 45.8% erythroid burst-forming units [BFU-E], 63.4 +/- 72.8% myeloid colonies [CFU-GM]) were achieved at a cooling rate of 1.4 K/min with 10% DMSO being present. The values obtained at a cooling rate of 5 K/min (-80 degrees C mechanical refrigerator) in the presence of 5% DMSO and 6% HES did not differ significantly (i.e., membrane integrity 91.8 +/- 3.9%, BFU-E 53.0 +/- 37.4%, CFU-GM 47.8 +/- 58.2%). At cooling rates above 5 K/min and DMSO concentrations lower than 5% (in spite of higher HES concentrations) there was a significant drop of CFU recovery (CFU-GM plus BFU-E) to almost 0%. In parallel, numerical recovery and viability dropped as well, but less pronounced, indicating that both methods are unsuitable for the prediction of CFU recovery when different freezing protocols are applied. We need at least 5% DMSO (in the presence of 6% HES) in spite of the undesirable histamine-releasing effect of this compound. The cooling rate is not critical in the range from 1 to 5 K/min and can easily be achieved by -80 degrees C freezers.

The introduction of PCR-based HLA typing techniques has uncovered that the HLA system is much more variable than it has been expected from the conventional typing methods. More and more new alleles are detected which are not characterized by new sequence motifs, but by new combinations of already existing sequence motifs. This variability, reflecting the immunological necessity to have the greatest capacity for presenting antigenic peptides, is increasingly complicating DNA typing techniques based on the diversity of the coding region. We have determined the sequence of the 1st through 3rd intron of the majority of HLA-A and HLA-B alleles in 48 well-defined cell lines and 195 PCR-typed clinical samples. The few published sequences emerged to contain substantial errors. The introns turned out to be highly polymorphic. Besides extensive homologies, numerous locus- and group-specific sites could be identified. The most intriguing finding was that most of the polymorphic motifs were related to serological families. These sequence motifs were extremely beneficial for setting up PCR-based typing systems. In particular, sequencing-based typing strategies benefited from intron-restricted priming for amplification and sequencing by enabling complete analysis of the polymorphic exons. The determination of cis/trans linkages of sequence motifs was substantially facilitated. Apart from these advantages, the intron sequences were useful for evolutionary studies, delivering more insights into the genetic relationship between different alleles and the mechanisms involved in the development of the diversity of HLA. Moreover, the variability of the introns may provide a structural basis for the identification of regulatory elements acting on the level of transcription.

Qualitative and quantitative detection of contaminating malignant cells in stem cell apheresis products intended for supportive therapy after high-dose chemotherapy of sensitive solid tumors is a demanding task for the laboratory. Human breast cancer cells in differing concentrations among mononuclear cells were quantitated by multicolor flow cytometry and immunocytochemistry. Comparing specificity, sensitivity, and laboratory processing time, the immunocytochemical detection of the above-mentioned malignant cells of epithelial origin was demonstrated to be the superior method.

The in-line-filtration of red cell concentrates (RCC) is an effective method to reduce white-cell-induced reactions. SAG-M RCCs were prepared in quadruple top/bottom bag systems and filtered after storage at 4 degrees C (study: n = 12). The mean white blood cell (WBC) reduction was 99.9% with the number of residual WBCs at about 0.67 x 10(6) per unit and with a loss of Hb mass of 10.64%. No cell fragments of WBCs and monocytes were detected (MAIPA). After storage of 42 days, mean values of hemolysis level (0.64%) and ATP concentration (2.27 mumol/g Hb) were improved compared with unfiltered RCCs. Comparably good results of leukocyte depletion were achieved by preparation of about 3,000 in-line filtrated RCCs for clinical use, measured by routine quality control (n = 25). Our experience shows that the systems are easy to handle and to use. The Leucoflex LCR4 in-line T/B system (Maco Pharma) can be recommended in the clinical practice by virtue of the effective leukocyte elimination, particularly in those blood banks with platelet concentrate production from buffy coats beside the RCC in-line filtration.

Despite many published studies no parameter could be identified yet to acceptably and individually predict collection results in stem cell apheresis. We analyzed leukocyte counts and processed blood volume, absolute and relative CD34+ cell counts, and overall collection efficiency in 120 patients with hematological and solid malignancies (354 leukaphereses using the Cobe Spectra cell separator, a median of 3 per patient, span 1-9). Stem cells were mobilized into peripheral blood by conventional chemotherapy followed by daily doses of G-CSF. CD34+ progenitor cell counts were monitored through multiparametric flow cytometry. Blood and collection flows varied in the range of 45-90 ml/min and 0.7-1.5 ml/min, respectively. CD34+ progenitor cells were enriched 38-fold in the apheresis product as compared to peripheral blood at a processed blood volume lower than one total blood volume. Efficiency continuously declined, on to a 25-fold concentration at a processed blood volume above the 3-fold total blood volume. Total collection efficiency, calculated from the absolute content of CD34+ progenitor cells in peripheral blood and apheresis concentrate (a parameter for progenitor cell mobilization during the apheresis), reached a plateau at a processed blood volume above the 3-fold total blood volume. However, variation among individual patients was high. The concentration rate of CD34+ cells at a leukocyte count below 5,000/microliter averaged 50 and declined continuously to 8 at leukocyte counts between 45,000 and 50,000/microliter. To summarize, in 70% of patients with leukocyte counts below 5,000/microliter and CD34+ progenitor cell counts above 10,000/ml, more than 1.5 x 10(6) progenitors per kg body weight could be collected in a single leukapheresis. According to the presented data, the variation in overall collection efficiency is mainly due to: 1) varying mobilization of progenitors during the apheresis procedures itself and 2) dependence on peripheral leukocyte counts.