Transfusion of stored red blood cells in critically ill trauma patients: a retrospective study.

OBJECTIVE: The many published studies on the effects of the transfusion of stored red blood cells on clinical outcomes yielded discordant results. Therefore, we chose to study patients with severe trauma. The clinical outcomes considered included in-hospital mortality, the occurrence of sepsis, length of stay in intensive care unit and in hospital, and days of mechanical ventilation. PATIENTS AND METHODS: We selected all patients with traumatic injury, who received at least 2 red cell units in the first day of admission. Patients were divided into two groups: those who had received fresh red cells only (fresh group) and those who had received at least one "old" red cell unit (old group). The red cells were considered fresh if they had been stored <14 days. RESULTS: The fresh and old groups included 376 and 321 patients, respectively. Baseline demographic and clinical characteristics were comparable between the groups. However, old group received more red cell and plasma units during whole hospital stay (red cells: 11 ± 7 vs 6 ± 4, p < 0.001; plasma: 7 [0-9] vs 3 [0-6]). Among outcomes, only length of stay in intensive care unit (old vs fresh: 18 ± 9 vs 12 ± 8 days, p < 0.001) and in hospital (77 ± 35 vs 45 ± 30 days, p < 0.001) differed significantly between groups. The association remained statistically significant in a multivariate analysis including known confounding factors. CONCLUSIONS: Patients with major trauma transfused with old (≥14 days) red cells had a longer length of stay in intensive care unit and in hospital, without any difference in mortality, occurence of sepsis or days of mechanical ventilation.

[In vitro experimentation of a new model of radial flow bioreactor containing isolated hepatocytes]. / Sperimentazione in vitro di un nuovo modello di bioreattore a flusso radiale contenente epatociti isolati.

Hepatocyte based artificial liver support systems are under investigation to support acute liver failure patients. The main purpose of such systems is to serve as a bridge to liver transplantation, or to promote spontaneous liver recovery. Limitation in mass-transfer capacity makes hollow-fiber bioreactors unsuited for long-term functioning of hybrid devices. We developed a novel radial-flow bioreactor in which the fluid perfuses the module from the center to the periphery, after having diffused through a space occupied by a three-dimensional structure filled with the hepatocytes. Five grams of freshly isolated porcine hepatocytes were seeded into uncoated, woven-non woven, hydrophilic polyester fabric, overlaid by two polyethersulfone membranes. Liver cells were perfused with 37 degrees C-warm, oxygenated, serum-free tissue culture medium, in which NH4Cl and Lidocaine were added at the final concentration of 1 mM and 60 micrograms/ml, respectively. Ammonium chloride removal, urea synthesis, monoethylglycinexylide (MEGX), pO2, pCO2, and pH were measured throughout the 14 day duration of the study. In a separate set of experiments, a scaled-up version of the radial flow bioreactor containing 150 grams of cells was perfused for 7 h with recirculating human plasma and MEGX production was monitored. During the 2 weeks of the study, an increasing production of urea was paralleled by constant ammonium removal. MEGX concentration after Lidocaine addition increased throughout the 14 days of perfusion with tissue culture medium, as well as after 7 hour perfusion with human plasma. Under transmission and scanning electron microscopy cells appeared attached to the polyester and one to each other, displaying ultrastructural features typical of functioning hepatocytes. Our study showed that liver cells were metabolically active when perfused into the radial-flow bioreactor. This configuration allowed close contact between media, or plasma, and cells at a physiological flow rate, by equalizing the concentration of the perfusing components, including O2, throughout the module. Our results suggest a potential use of this system for temporary extracorporeal liver support in acute hepatic failure patients.

Assessment of distribution of CD34 epitope classes in fresh and cryopreserved peripheral blood progenitor cells and acute myeloid leukemic blasts.

BACKGROUND AND OBJECTIVE: So far several reports have described changes in the expression of surface antigens in progenitor cells and blasts following cryopreservation. However, there are no data on the effects of cryopreservation on the expression of the three CD34 epitope classes, and on their relationship with the clonogenic capacity of PBPC collected by leukapheresis. DESIGN AND METHODS: In order to analyze the effects of freezing/thawing procedures (Eth 80C storage for 3 months) and use of dimethylsulfoxide (DMSO) on the immunophenotype profile and colony production of peripheral blood progenitor cells (PBPC) in apheresis products derived from 20 patients with stage 0-III non-Hodgkin's lymphoma (nHL), a flow cytometry study was undertaken using different CD34 monoclonal antibodies (MoAbs) capable of recognizing the 3 epitope classes of CD34 molecule (class III: HPCA-2/FITC, HPCA-2/PE, 581/FITC, 581/PE; class II: Q-Bend 10/PE; class I: ICH3/PE, BI3C5-PE, Immu-133-PE). CD34 epitope expression was also analyzed in thawed CD34+ blasts obtained from 14 patients with acute myeloid leukemia (AML), who were analyzed using a larger number (#17) of CD34 epitope class I, II, and III reactive MoAbs. RESULTS: Under our experimental conditions it was found that class III and class II CD34 epitopes (differentially resistant to enzymatic cleavage with neuraminidase, chymopapain and glycoprotease) are better preserved than class I epitope Eth sensitive to degradation Eth after cell exposure to cryoprotectant DMSO and the freezing- thawing procedures. Results further showed a concomitant decrease in class I CD34+ counts and in BFU-E colony production. A significant increase in CD34 antigen expression levels (i.e. antibody binding capacity, ABC) by cryopreserved cells stained with CD34 epitope class III, and class II reactive MoAbs was also documented, while no changes after cryopreservation were noted using class I-reactive MoAbs. The slight increase in the percentage of CD34+ cells detected after frozen storage was correlated to a concomitant decrease in the number of more mature myeloid cells (CD15+, CD13+, CD33+). Compared to pre-cryopreservation values, a slight reduction in class I CD34 epitope expression was also found in thawed CD34+ AML blasts. INTERPRETATION AND CONCLUSIONS: As far as the reduction of class I CD34 epitope is concerned, it may be hypothesized that the freezing procedure, use of DMSO, and/or lysis methodology may either damage a CD34 subset, or induce distinct alterations of the CD34 glycoprotein, possibly determining a reduction in their immunoreactivity with some CD34 MoAbs. In conclusion, this study has shown that exposure to the cryoprotectant DMSO and the freezing/thawing procedures modifies the distribution of CD34 epitopes as well as the clonogenic capacity of PBPCs from nHL patients, and CD34+ blasts from AML. These findings need to considered when selecting CD34 MoAbs for enumeration and positive selection of stem/progenitor cells for research and clinical purposes.

Flow cytometric analysis of anti-platelet antibodies in idiopathic thrombocytopenic purpura.

Anti-platelet antibody measurement may be important in defining the pathogenesis of thrombocytopenic states. In this paper we compared three anti-human immunoglobulin reagents by using them to detect anti-platelet antibodies on the platelet surface and in the serum of 14 patients with chronic idiopathic thrombocytopenic purpura (ITP) and 22 thrombocytopenic disorders. Samples were analyzed by both flow cytometry and a fluorescence microscope. In ITP patients, the direct test was positive in 50% of the cases, while the indirect technique proved to be positive in a slightly higher number of those tested (56%). Furthermore, the number of positive cases was similar for the three reagents used in this study, although the mean percentage of positive platelets was higher for the kappa/lambda monoclonal reagent. These data further support the sensitivity and reproducibility of flow cytometry analysis, which was capable of detecting antiplatelet antibodies in all patients with transfused Cooley's disease (regarded as positive control), as well as in a significant number of patients with ITP or related diseases. On the basis of the data presented here, definitive proof regarding the presence of anti-platelet antibodies in patients with thrombocytopenia still has to be found, and further studies are needed in order to ascertain the autoimmune nature of these disorders.

Increased uptake of radio labelled white blood cells into abdomen of uremics on peritoneal dialysis.

Peritonitis and exit-site tunnel infection are frequent causes of CAPD drop out. We studied 9 patients, 8 treated by CAPD and 1 by IPD. These patients underwent sonographic and scintigraphic study of the abdomen. All scintigraphic examinations showed a visceral uptake. In two cases, sub-clinical bowel inflammation, demonstrated by scintigraphic study, preceded a gram negative peritonitis. The scintigraphic study with radiolabelled white blood cells may be useful in identifying chronic aseptic inflammations and some bowel and exit-site conditions which are possible risk factors in some cases of peritonitis.

Treatment of cerebral malaria by erythrocyte exchange.

A 33-years-old male presented with a severe malaria, caused by a chloroquine resistant strain of Plasmodium falciparum. The number of parasitized erythrocytes reached 20% and the patient had cerebral complication. During the second hospital day, an erythrocyte exchange was performed as an in addition to chemotherapy. The patient's clinical condition improved and the parasitemia disappeared. The erythrocyte exchange is recommended in severe malaria, when parasitemia greater than 10%, with or without cerebral, renal or blood coagulation complications.

Clinical efficacy of five filters specific for leukocyte removal.

The clinical efficacy of 5 filters specific for leukocyte depletion was evaluated retrospectively. The patient population was composed of 191 thalassemics who received about 15,000 units of filtered blood in 28 months. The frequencies of reactions to filtered blood varied significantly (p less than 0.0001) according to the type of filter and were as follows: Erypur, 0.7%; Imugard, 2.7%; Leucostop, 0.7%; Miropore, 2.1%; Sepacell, 0.6%. There was a notable concordance between the efficacy in the leukocyte removal and the clinical performance of the filters. The 2 polyester filters examined combined good clinical and laboratory results with superior flow properties.

Prevention of non-hemolytic transfusion reactions with leucocyte-poor blood: a prospective study.

Twenty-five patients with homozygous beta-thalassemia, who complained of occasional nonhemolytic transfusion reactions (NHTR), were alternately transfused eight times with normal packed red cells (PRC) and leucocyte-poor blood prepared by three methods: cotton wool filtration (CWF), buffy coat removal after dilution with saline and upright spin (BCR), and microaggregate filtration (MAF). On overall, CWF-treated RBC concentrates (CWF-RBC) contained 0.02 x 10(9) leucocytes; BCR-RBC contained 0.4 x 10(9), MAF-RBC 1.3 x 10(9) and PRC 1.7 x 10(9). The frequencies of NHTR were: CWF, 0.02; BCR, 0.04; MAF, 0.20; PRC, 0.28. Compared to PRC, both CWF and BCR significantly reduced the frequency of NHTR (p less than 0.01), whilst MAF did not (p greater than 0.1). On the other hand, in one patient CWF caused the two most severe NHTR recorded in this study, both of which were characterized by anaphylaxis.

[Antiglobulin test using an immunoenzyme method]. / Il test dell'antiglobulina con metodica immunoenzimatica.

An enzyme-linked antiglobulin test (ELAT) is described. All reagents are commercially available. The antiglobulin reagent is a monoclonal anti-IgG conjugated with alkaline phosphatase. The substrate (p-nitrophenylphosphate) is stable and innocuous. The procedure requires long incubations, particularly between red cells and the enzyme-conjugated antibody. However most of the time actively spent by the technician is consumed by washing procedures (LAV). These may be avoided by using density gradient centrifugation (CSG). ELAT is significantly more sensitive than agglutination, using either washing or gradient centrifugation (P = 0.002 and P = 0.0005, respectively). Moreover ELAT-CSG is significantly more sensitive than ELAT-LAV (P = 0.03). CSG yields better reproducibility and probably avoids some of the spontaneous elution of the primary and/or secondary antibody which may occur during washing.

[Hematological variables in marathon and ultramarathon runners]. / Variabili ematologiche in maratoneti ed ultramaratoneti.

The haemoglobin concentration of runners has been reported to be often below normal (1). In the present investigation the haemoglobin and iron concentrations and the haematocrit have been determined in 45 marathon runners examined before and after the 1982 Italian Marathon Championship and in 79 runners examined before and after the 1982 Firenze-Faenza race (107 km). The results obtained suggest that the training programs followed by the marathon runners (up to 260 km per week in the months preceding the race) are accompanied by a significant decrease of the haemoglobin and iron levels and of the haematocrit. Similar results have been observed in the participants to the Firenze-Faenza race. Nevertheless in these ultramarathoners the decrease of the haemoglobin and iron concentration and of the haematocrit are less marked, possibly because their training programs are less intense than those of the marathon runners. The findings obtained are in favour of the hypothesis that the degree of "sport anaemia" might be related to the amount of training of the athletes.