Survival and recovery of human platelets stored for five days in a non- plasma medium

Human blood platelets were stored for five days as concentrates in 60 mL of: (a) plasma; (b) non-plasma medium with anticoagulant; and (c) non-plasma medium without anticoagulant. All preparations were equally functional when tested for platelet aggregation and release reaction in response to single agonist or synergistic pairs of agonists in vitro. Platelets stored in non-plasma medium with anti-coagulant had lower kallikrein, fibrino(gen)peptide A, lactate, and beta-thromboglobulin than did plasma controls after five days. In vivo recovery and survival of platelets stored in non-plasma medium with anticoagulant were 51.2% +/- 4.3% and 8.7 +/- 0.3 days, respectively, which were not statistically different from plasma controls of 39.2% +/- 4.9% and 7.2 +/- 0.8 days, respectively. It is concluded that platelets can be stored for five days in a non-plasma medium and still have good in vivo recoveries and survivals.     

Investigation of the thrombin-generating capacity, evaluated by thrombogram, and clot formation evaluated by thrombelastography of platelets stored in the blood bank for up to 7 days

BACKGROUND AND OBJECTIVES: Transfusion based on the Thrombelastograph (TEG) results reduces transfusion requirements in cardiac surgery and in liver transplantation. Taking the pivotal role of thrombin generation in the coagulation process into consideration, the clinical utility of the TEG may, in part, depend on its reflection of the dynamics of thrombin generation. MATERIAL AND METHODS: The kinetics of thrombin generation of platelets stored for 2 and 7 days, respectively, was assessed by calibrated automated thrombogram (CAT) and the lag time (min), time to peak (ttPeak; min), peak (nm thrombin) and endogenous thrombin potential (ETP; nm thrombin*min) were registered. Clot formation was evaluated by TEG and the R time (min), maxial amplitude (MA; mm), time to maximum thrombus generation (TMG; min) and maximum thrombus generation (MTG; dynes cm(-2) s(-1)) and total thrombus generation (TTG; dyne cm(-2)) were registered. RESULTS: Platelets become more procoagulant, evaluated both by TEG and CAT during storage. The reduction in CAT lag time and the ttPeak correlated with a decrease in the TEG R time and TMG (P < 0.0001) as did the CAT peak thrombin generation and the TEG MTG (P = 0.0035). No correlation between ETP and TTG was found (P = 0.65). CONCLUSION: The kinetics of thrombin generation, as evaluated by CAT, correlates with the thrombus generation, as evaluated by thrombelastography and this may in part explain the clinical utility of the TEG in identifying clinically relevant coagulopathies, secondary to impaired thrombin generation.     

Granulocyte colony-stimulating factor alone at 12 microg/kg twice a day for 4 days for peripheral blood progenitor cell priming in pediatric patients

In children, the optimal mobilization schedule for harvesting peripheral blood progenitor cells (PBPC) is an issue of continuous research. We have studied a schedule based on high and daily divided doses of G-CSF (12 microg/kg body weight twice daily) for 4 days for PBPC priming. Toxicity related to G-CSF was observed in 13 patients (23%), mainly mild bone pain and myalgia. The median CD34(+)cell number collected was 4.4 (0.4-35 x 10(6)/kg body weight), with 46 patients achieving 2 x 10(6)/kg body weight (83.6%) after a single large volume leukapheresis. In conclusion, this mobilization schedule allows safe and efficient collection of the minimum target CD34(+) cell dose in most pediatric patients by only one procedure.     

A simplified screening for alpha-thalassemia 1 (SEA type) using a combination of a modified osmotic fragility test and a direct PCR on whole blood cell lysates

In order to provide a rapid method for identifying alpha-thalassemia 1 in a region with massive population and limited resources, we have tested a rapid screening strategy. Preliminary screening was done using a modified one tube osmotic fragility test (OF test) followed by RBC indices; Hb analysis and detection of alpha-thalassemia 1 with the Southeast Asian deletion (SEA type) were performed by PCR. One hundred and seventy-five adult Thai subjects were studied. Fifty-one of the 175 subjects (29.1%) were positive for a modified OF test. They all had significantly lower MCV and MCH but higher RDW-CV values as compared to the OF negative group. A successful identification of alpha-thalassemia 1 deletion using a direct PCR on cell lysates was demonstrated. Among the 51 OF-test-positive subjects, 7 were found to be alpha-thalassemia 1 carriers, 3 of whom were also carriers of Hb E. No alpha-thalassemia 1 was detected in the OF-test-negative group. A combination of a modified OF test and a direct PCR analysis on whole blood cell lysates would therefore provide an effective screening for alpha-thalassemia 1 in the regions where a program of prevention and control of the disease remains underserved.     

Relation of infarct site to 15 year prognosis in patients who survived for 28 days after a first myocardial infarction

Six hundred and eighty four patients (629 men), all aged under 60 years, who had survived for 28 days after a first acute myocardial infarction were studied to determine the influence of the site of infarction on long term prognosis. The infarct site was not significantly related to age nor to extent of infarct at the time of the acute episode. Mechanical complications were more common in patients with anteroseptal infarctions, while atrioventricular conduction disturbances were more commonly found in those with inferior infarction. The site of infarction was not related to smoking habits or angina before the infarction or at 2 year follow up. Life table methods did not show any relation between infarction site and morbidity or mortality either two years or 15 years after the initial infarction.     

Interaction of indomethacin with cytokine production in whole blood. Potential mechanism for a brain-protective effect

Both Alzheimer's disease and vascular dementia are featured by inflammatory responses and it is known that non-steroidal anti-inflammatory drugs (NSAIDs) decrease the risk and severity of these diseases.To study the effect of NSAIDs on PGE2 levels and pro- and anti-inflammatory cytokine levels in the whole blood assay, blood samples from 23 elderly persons aged 85 years were stimulated with thrombin or LPS as primary stimulus. Indomethacin was added in concentrations ranging from 0.4 to 16 microg/ml and acetylsalicylic acid was added to in concentrations ranging from 0.5 to 8.0 microg/ml.Indomethacin abrogated thrombin- and LPS-induced PGE2 production at all concentrations tested. In addition, indomethacin reduced the production of thrombin-induced IL-6 and IL-10 (p<0.05) at physiological concentrations. Indomethacin reduced the production of LPS-induced IL-6, IL-1 beta and IL-10 (p<0.05) at the highest indomethacin concentration tested. Similar results were obtained upon incubation with acetylsalicylic acid.It is concluded that indomethacin may reduce the thrombin-induced inflammatory reaction by decreasing IL-6 through inhibition of PGE2 synthesis. This IL-6 reduction may be relevant for the ability of indomethacin to reduce the risk of Alzheimer's disease. However, the decrease in IL-10 production due to indomethacin suggests a more inflammatory state.     

Characterization of clonogenic progenitors in autologous peripheral blood stem cell grafts: evaluation of a simple in vitro assay suitable for routine clinical use

Autologous peripheral blood stem cell (PBSC) transplantation has a low treatment-related morbidity and mortality when using appropriate criteria for patient selection and graft quality evaluation. It will be important to use simple and standardised procedures for evaluation of progenitor cell numbers when considering autografting in patients with malignant or non-malignant disorders and increased risk of prolonged posttransplant cytopenia. We determined the number of clonogenic cells in PBSC autografts after 7 days of in vitro culture, and these results were compared with both the total number of colonies and the numbers of colony subsets in conventional 14 days colony assays (colony-forming unit granulocyte-erythrocyte-macrophage-megakaryocyte, CFU-GEMM; CFU-E, CFU-GM; CFU-megakaryocyte). The total colony number after 7 days of culture correlated significantly with (i) the CD34+ cell number; (ii) the total colony number as well as the numbers of erythroid, nonerythroid and mixed colonies in a conventional assay using 14 days of culture; (iii) the number of megakaryocyte colonies. The total colony number after 7 days of in vitro culture is a simple in vitro parameter that seems to reflect the proliferative capacity of various progenitor subsets in PBSC autografts. This simple analysis may be used in combination with other in vitro techniques (e.g. estimation of stem cell viability and CD34+ cell subset analysis) for pretransplant evaluation of autografts. However, the possible clinical use of this parameter has to be examined in prospective clinical studies.     

Relation of infarct site to 15 year prognosis in patients who survived for 28 days after a first myocardial infarction.

Six hundred and eighty four patients (629 men), all aged under 60 years, who had survived for 28 days after a first acute myocardial infarction were studied to determine the influence of the site of infarction on long term prognosis. The infarct site was not significantly related to age nor to extent of infarct at the time of the acute episode. Mechanical complications were more common in patients with anteroseptal infarctions, while atrioventricular conduction disturbances were more commonly found in those with inferior infarction. The site of infarction was not related to smoking habits or angina before the infarction or at 2 year follow up. Life table methods did not show any relation between infarction site and morbidity or mortality either two years or 15 years after the initial infarction.     

The thin prep rat aortic ring assay: a modified method for the characterization of angiogenesis in whole mounts

The rat aortic ring model has gained broad acceptance as an angiogenic assay. This system can be used to study the activity of angiogenic and anti-angiogenic factors, and investigate the molecular mechanisms of the angiogenic process. We describe here a thin prep modification of the aortic ring model, which significantly simplifies the procedure and allows staining of aortic outgrowths as whole mounts. Using this procedure, intact preparations of angiogenic outgrowths are successfully and reproducibly stained with endothelial cell (anti-CD-31 and -Tie2 antibodies, Griffonia Simplicifolia isolectin-B4) and smooth muscle cell (anti--smooth muscle actin antibody) markers. Combined use of double immunostaining and confocal microscopy allows concurrent visualization of endothelial and mural cells in the same cultures. Whole mount immunostains of rat aorta cultures are an effective way to rapidly characterize the cellular composition of the angiogenic outgrowths, and localize proteins implicated in the regulation of angiogenesis. This method should facilitate the work of the many vascular biologists that have adopted the rat aorta model as a tool to study angiogenesis and its mechanisms.     

Comparison between a whole blood interferon-gamma release assay and tuberculin skin testing for the detection of tuberculosis infection among patients at risk for tuberculosis exposure

A new test that measures interferon-gamma (IFN-gamma) release in whole blood following stimulation with tuberculin has the potential to detect tuberculosis infection using a single blood draw. The IFN-gamma release assay was compared with the standard tuberculin skin test (TST) among 467 intravenous drug users at risk for tuberculosis in urban Baltimore. Among 300 human immunodeficiency virus (HIV)-seronegative patients, the IFN-gamma release assay was positive in 177 (59%), whereas the TST was positive in 71 (24%), for a percent agreement of 59% (kappa=26%). Among 167 HIV-seropositive subjects, the IFN-gamma release assay identified 32 reactors (19%); the TST identified 16 reactors (9.6%), for a percent agreement of 82% (kappa=28%). The IFN-gamma release assay detected more reactors than did the TST, but its agreement with TST was weak. As the TST is an imperfect standard, further evaluation of the IFN-gamma release assay among uninfected persons and persons with culture-confirmed tuberculosis will be useful.     

Evaluation of the validity of a rapid method for measuring high and low haemoglobin levels in whole blood donors

Background

Haemoglobin screening methods need to be highly sensitive to detect both low and high haemoglobin levels and avoid unnecessary rejection of potential blood donors. The aim of this study was to evaluate the accuracy of measurements by HemoCue in blood donors.

Materials and methods

Three hundred and fourteen randomly selected, prospective blood donors were studied. Single fingerstick blood samples were obtained to determine the donors'' haemoglobin levels by HemoCue, while venous blood samples were drawn for measurement of the haemoglobin level by both HemoCue and an automated haematology analyser as the reference method. The sensitivity, specificity, predictive values and correlation between the reference method and HemoCue were assessed. Cases with a haemoglobin concentration in the range of 12.5–17.9 g/dL were accepted for blood donation.

Results

Analysis of paired results showed that haemoglobin levels measured by HemoCue were higher than those measured by the reference method. There was a significant correlation between the reference method and HemoCue for haemoglobin levels less than 12.5 g/dL. The correlation was less strong for increasing haemoglobin levels. Linear correlation was poor for haemoglobin levels over 18 g/dL. Thirteen percent of donors, who had haemoglobin levels close to the upper limit, were unnecessarily rejected.

Discussion

HemoCue is suitable for screening for anaemia in blood donors. Most donors at Yazd are males and a significant percentage of them have haemoglobin values close to the upper limit for acceptance as a blood donor; since these subjects could be unnecessarily rejected on the basis of HemoCue results and testing with this method is expensive, it is recommended that qualitative methods are used for primary screening and accurate quantitative methods used in clinically suspicious cases or when qualitative methods fail.     

Quantitation of Hb a2 with DE-52 microchromatography in whole blood as screening test for beta-thalassemia heterozygotes.

HB A2 was assayed by means of DE-52 microchromatography in hemolysates from 285 normal subjects and 223 beta-thalassemia heterozygotes. No overlap was found between both groups. Comparable results were observed analyzing whole blood samples collected in capillary tubes from 550 normal subjects and 295 beta-thalassemia heterozygotes. Our results demonstrate that this technique is useful in a screening program for beta-thalassemia trait.