In order to investigate the contribution of transfusion to CMV infection, 20 CMV-seropositive and 3 seronegative patients undergoing allogeneic bone marrow transplantation were studied for their CMV infection. All the patients received cellular blood component transfusion through a leukocyte depletion filter. CMV infection is defined as positive results with a nested double polymerase chain reaction (PCR) and/or IgM antibody testing. Blood samples of 10 healthy marrow donors and 10 pretransplant patients were available for PCR examination. No CMV-DNA was demonstrated. In contrast, among a total of 122 peripheral blood specimens from the 23 posttransplant patients, 51 from 14 patients were positive for CMV-DNA. Of 20 seropositive patients, 15 were transfused with CMV-seronegative blood which is consisted of 14.3% of blood donors and 5 were transfused with CMV-unscreened blood. Four (80%) of the latter 5 seropositive patients showed CMV infection. Of 15 seropositive patients transfused with CMV-seronegative blood, 11 (73.3%) exhibited CMV infection. There was no significant difference in the incidence of CMV infection between the two seropositive patient groups (Fisher's exact probability test). In most cases, CMV infection occurred after 30 days post BMT. Interstitial pneumonia developed in two seropositive patients who had received CMV-seronegative blood products, and had positive results on both PCR and IgM testing. Regardless of transfusing CMV-unscreened or seronegative blood, none of the seronegative patients were infected with CMV. Our present observations suggest that the CMV-seronegative blood component contributes little if any to prevent seropositive patients from CMV infection.

View full abstract

Human umbilical cord blood (HUC) contains as many hematopoietic stem/progenitor cells as bone marrow. However, this number may not be sufficient for allogenic transplantation in adults. In this study, mononuclear cells from umbilical cord blood were loaded into flasks of the AIS Micro CELLector™ to capture CD34⁺ cells. CD34⁺ cells were enriched 60 times by the treatment and the recoveries of CD34⁺ cells and colony forming cells was 37.3±19.3% and 14.5±11.2% (n=14, mean±SD), respectively. The enriched CD34⁺ cells were then cultured in vitro for 7 to 14 days in the presence of IL-3, IL-6 and stem cell factor. The total number of cells and the number of granulocyte-macrophage colony-forming cells increased 80.8±51.2 and 75.0±53.6 fold, respectively in 14 days. The number of CD34⁺ cells was retained up to 7 days and increased 3-fold after 14 days. Surface marker analysis showed that the percentage of CD34⁺ CD33^- cells decreased from 65.5% to 4.0% in 3 days and that of CD34^- CD33⁺ cells increased from 3.8% to 86.2% in 7 days. The percentage of CD34⁺ CD38^- cells decreased from 10.6% to 3.8% in 3 days and that of CD34^- CD38⁺ cells increased from 10.9 to 76.2% in 7 days. These results indicate that the cultured progenitor cells differentiated as the culture period was extended.
Thus it was shown that hematopoietic progenitor cells enriched from HUC using the Micro CELLector system can be efficiently expanded in vitro.

View full abstract

UV-B irradiation of platelet concentrates (PC) has been tried in several institutes to inactivate leukocytes in PC and prevent alloimmunization on platelet transfusion. However, the mechanism of inactivation of leukocytes contaminating PC has not been fully understood.
It is known that UV-B light is absorbed by photosensitizers in cells and produces active oxygen and radicals, such as singlet oxygen, superioxide anions and hydroxyl radicals. These active oxygen or radicals should injure cellular components and this could cause the suppression of cellular functions.
In this study, we investigated the relationships among UV-B irradiation, free radical generation and leukocyte inactivation. We found the evidence that active oxygen and radicals were produced in peripheral blood mononuclear cells by UV-B irradiation. UV-B irradiation suppressed the stimulatory function of leukocytes in a mixed lymphocyte reaction (MLR), and the suppression depended on the dosage of UV-B. Even a low dosage of UV-B, 10J/m², could inhibit the MLR if the irradiated cells were incubated at 37°C for 24 hours before co-culture with responder cells. Treatments of cells with the exogenous singlet oxygen or superoxide anions also caused suppression of the stimulatory function in the MLR, inhibition of capping formation of HLA-DR antigens, and an increase of intracellular free Ca²⁺ levels as did the UV-B treatment.
These results indicate that the active oxygen or radicals generated in UV-B-irradiated leukocytes could be one of the causes of leukocyte inactivation.

View full abstract

Histamine and anaphylatoxin (C3a) levels in plasma of concentrated red cells (CRC) and red cell concentrates with MAP solution (RC-M·A·P) were examined.
Plasma histamine levels were increased to 20nmol/2U in CRC (WBC, 2.7×10⁹/2U), and 2nmol/2U in RC-M·A·P (WBC, 0.7×10⁹/2U) at 21 days of storage. Good linear correlation (r=0.952) was found between the initial WBC content in blood and the histamine levels at 21 days of storage. The high plasma histamine levels in CRC may be a contributing factor to the rashes, wheezeing and flushing of nonhemolytic transfusion reactions.
Anaphylatoxin (C3a and C3a-desArg) levels increased to approximately 200μg/2U in CRC and RC-M·A·P, a level reportedly sufficient to induce platelet activation, suppression of NK activity and induce IL-1 release from macrophages.
These findings suggested that some nonhemolytic transfusion reaction might be due to the infusion of plasma with high histamine and anaphylatoxin levels and not always result from an antigen-antibody reaction.

View full abstract

We measured the plasma levels of histamine, cytokines (IL-1β, IL-6, TNF-α, and IL-8) and anaphylatoxin (C3a) in platelet concentrates (PCs) during storage.
Increased TNF-α levels were found in PCs after 120 hours of storage. However, the PCs stored for 120 hours contained TNF-α in amounts well below those which induce fever, chills and other symptoms in most patients treated with recombinant TNF-α.
Increased IL-8 levels were also found in stored PCs. Thus, it is important to note that a critical IL-8 threshold might be reached in a patient upon infusion of stored PCs containing high levels of IL-8.
Plasma histamine levels were below 1nmol/2U PCs after 120 hours of storage.
Anaphylatoxin (C3a and C3a-desArg) levels were increased to 56μg/2U (1.6×10^-7M) after 72 hours of storage, a level reportedly sufficient to induce platelet activation, suppress NK activity and induce IL-1 release from macrophages.
These results may help explain some of the unexpected reactions in patients receiving PCs transfusions.

View full abstract

By adding adenine (CPDA-1) to citric soda, phosphate, and dextrose solution (CPD), the preservative period of whole blood can be lengthened for two weeks. To measure the effectiveness of CPDA-1 in autologous blood transfusion, we compared it with CPD in the autologous blood of 34 patients stored before elective surgery (CPDA-1 n=9; CPD n=25).
The volume of autologous blood collected, not using the leap-frog method, was 943ml in the CPDA-1 group and 854ml in the CPD group (p<0.05), and storage periods averaged 24.4 days and 18.3 days (p<0.05), respectively.
There was no functional disorder of the kidney and liver indicated in the laboratory data either before or after receiving autologous blood units. Of the patients, seven (78%) in the CPDA-1 group and 19 (76%) in the CPD group completed their operations using only autologous blood (n. s.). The recovery of in vivo autologous blood measured one day after transfusion was evaluated as effective in 8 (89%) of the 9 patients in the CPDA-1 group and in 12 (86%) of 14 (data available) patients in the CPD group (n. s.).
Our observations confirm the usefulness of CPDA-1 in the preservation of autologous blood, and we feel that the solution can also be used in allogenic blood transfusion.

View full abstract

This report described (1) a case of Streptococcus pneumoniae bacteremia whose erythrocytes showed Th and T polyagglutination in different conditions, and (2) a further investigation indicating that Th transformation was the “prestage” of T activation caused by desialylation from red cell membrane with bacterial enzyme at least in the case of infection.
A screening test with Arachis hypogaea happened to find erythrocyte-polyagglutination in a sample from a 55-year-old man with pneumonia and septicemia of the bacteria. An immunohematological study demonstrated Th activation (Bird's classification) on the red cells obtained directly from the patient. This contrasted with T activation of the patient's red cells taken from a culture bottle.
Erythrocytes from a healthy donor revealed Th transformation, when incubated with the patient's serum at 37°C only for more than 5 hours, and T transformation, easily when incubated with the undiluted supernatant of the blood culture. Interestingly, 10 times dilution of the supernatant brought about Th activity on the red cells after 30 minutes incubation, followed by T activation after 60 minutes or more. The 100 times dilution induced solely Th phenomenon.
The experiments using bacterial N-acetylneuraminic acid successfully transformed normal erythrocytes into either Th or T red cells, showing that the difference depended on the amount of sialic acid hydrolyzed: 4-21μg/10¹⁰ RBC for Th and >21μg/10¹⁰ RBC for T transformation.

View full abstract

Human umbilical cord blood (HUC) has been considered as a possible source of hematopoietic stem and progenitor cells for transplantation. In this study, we characterized mononuclear cells and hematopoietic stem/progenitor cells in cord blood and compared them to those in human peripheral blood (HPB). The concentration of mononuclear cells in HUC was twice as high as that in HPB. The percentage of CD34-positive cells in HUC was 4.0 times that in HPB and the result of subset analysis suggested that HUC involves many immature cells. The plating efficiency of colony forming unit in mononuclear cells in HUC was 0.446±0.295%, 14.4 times higher than in HPB. Thus, HUC contains a large number of hematopoietic stem cells. In addition the rate of CFU-GM and CFU-mix in HUC was more than in HPB. It is suggested that HUC is better source of stem/progenitor cells. The limitation of HUC as a potential source of transplantable hematopoietic stem cells is its small quantity. It is necessary to develop a method to expand the number of hematopoietic stem/progenitor cells in HUC to obtain enough cells for allogeneic transplantation in adults.

View full abstract

We tried to make an appropriate standard condition for UV-B irradiation of platelets concentrates (PC), which is useful for prophylaxis against post-transfusion graft versus host disease (PT-GVHD) as well as prevention against alloimmunization.
Agitation of PC bags during UV-B irradiation is necessary to irradiate evenly cells in the bag, becaue a lot of UV-B ray should be absorbed by bag membrane and plasma. Amounts of UV-B that each lymphocyte or platelet would actually receive on an average (UVavg) was calculated by the equation as below.
UV*avg=K₁×(K₂^L-1)×UV/(log_eK₂×L), K₁ and K₂ are permiability index of bag membrane and that in plasma, respectively; while L and UV stands for depth of PC bag and emitting dose of UV-B, respectively. We irradiated PC bags with UV-B in a dose of 541-13, 525J/m² of UV*avg, and examined lymphocytes in the bags about the responder and stimulator activities in mixed lymphocytes culutre (MLR), as well as platelet function in the bags. Irradiation more than 5, 000J/m² of UV*avg is needed to suppress responder and stimulator activities, and platelet function is maintained up to 13, 525J/m² of UV*avg.
In conclusion, UV-irradiation in a range of 6, 000-13, 000J/m² of UV*avg is considered appropriate to prevent from PT-GVHD or alloimmunization.

View full abstract

We surveyed the frequecy of autologous blood transfusion (AT) using a questionnaire sent to 80 hospitals, which use blood transfusion frequently in Aichi Prefecture. Answers were obtained from 65 hospitals (81%). In 36 hospitals (55%) AT was performed. The methods of transfusion were as follows; predonating AT in 86%, salvaging AT in 33%, and hemodilutional AT in 6%. AT was employed by orthopedic surgery in 23 hospitals, cardiovascular and thoracic surgery in 17 hospitals, and hematology and abdominal surgery in 7 hospitals, respectively.
The use of AT is still increasing in Aichi Prefecture and more hospitals are expected to employ it to prevent serious complications of homologous blood transfusion.

View full abstract

Blood program in Thailand will continue to develope, with regards to safety and adequacy of blood supply. The safety of blood is our primary concerm. Establishment of regional laboratory processing centers throughout the country is being implemented which will later transform into comprehensive regional transfusion center. Standardization of quality of blood throughout the country will hopefully be achieved. Heat treated lyophylized cryoprecipitates for home treatment program of hemophilia are now being prepared. PCR laboratory will also established.
Specialized blood products are being developed to accommodate the advanced medical care; e. g. peripheral stem cell collection and marrow donor program are being considered.

View full abstract