Hiroyuki Kiyokawa

Prevention of transmission of human T-lymphotropic virus type 1 (HTLV-1) through transfusion, by donor screening with antibody to the virus One-year experience

To prevent the transmission of human T‐lymphotropic virus, type 1 (HTLV‐1) during blood transfusion, a program was implemented to screen donors for antibodies to the virus, using a newly developed, passive agglutination (PA) method. During the period April 1986 to March 1987, 675 recipients of donor blood in whom the antibody to HTLV‐1 was not present before transfusion were followed for at least 50 days after transfusion. One of these 675 seroconverted despite the transfusion of screened blood, but this seroconversion rate (0.15%) represents a marked decrease from the rate of 8.3 percent prevalent before donor screening began. The rate in the Fukuoka area of donors seropositive for anti‐HTLV‐1 is 5.34 percent, as detected by the PA method and 1.80 percent, as assessed by the indirect immunofluorescence (IF) technique, with PA‐positive but IF‐negative blood units thus accounting for 3.5 percent (5.34–1.80) of the total blood donated. The seroconversion rate among recipients transfused with blood screened by IF (at Kyushu University Hospital only) from 1981 to 1985 was 0.41 percent, which was not significantly different from the rate of 0.15 percent observed after PA screening. The discrepancy between PA and IF in the rate of seropositivity was due, in part, to the higher sensitivity of PA in detecting anti‐HTLV‐1. It is proposed that all donor blood in areas where HTLV‐1 is endemic be screened by PA before transfusion. For reasons of expense and equipment unavailability, it is not possible for IF to be used on a general basis, but because of the possibility of false‐positive results with PA, PA‐positive blood units should be reexamined by IF or Western blot testing to reduce the amount of wasted blood, especially in areas highly endemic for HTLV‐1, such as Kyushu, Japan.

Simple standing test predicts and water ingestion prevents vasovagal reaction in the high-risk blood donors

BACKGROUND: One of the serious complications of blood donation is vasovagal reaction (VVR) with syncope. This study was performed to determine if the measurement of hemodynamic responses to standing before blood collection (BC) was useful to identify the high‐risk donors for VVR and also examined the effect of 300 mL of water ingestion in the prevention of VVR.

Performance Certification of Gelatin Particle Agglutination Assay for Anti-HTLV-1 Antibody: Inconclusive Positive Results

In order to test the performance of particle agglutination assay (PA), 800 preselected PA‐positive sera at 8 blood centers in the Kyushu area were tested in various assays. Most blood centers should improve their PA technique, since a third of the samples were PA‐negative in our hands. A third of our PA high‐titer sera were negative in indirect immunofluorescence and enzyme imiminoassay, the results of which were consistent with each other. Western blots did not detect every positive serum. PA inhibition positivity was not consistent with PA titer. Most IgM antibody‐positive sera also contained IgG antibody. PA should be used in combination with other methods before notifying the results to positive testees.

Preparation of white-cell-poor blood components using a quadruple bag system

A method for the preparation of white‐cell‐poor red cells from 400 ml of blood collected in a quadruple bag with one 80‐ml and two 300‐ml satellite bags is described. In this procedure, a platelet concentrate was prepared from the buffy coat fraction obtained by the first centrifugation of whole blood. After centrifugation of whole blood for 5 minutes at 3500 × g, the plasma was transferred into the 300‐ml bag until the interface of red cells and plasma reached a level 32 mm from the top of the bag; then approximately 70 g of plasma and buffy coat were collected into the 80‐ml bag. The buffy coat fraction was centrifuged further for 5 minutes at 170 × g, and the supernatant (concentrated platelets in plasma) was transferred into the second 300‐ml bag. In this blood processing, the recovery of red cells into the packed red cells and of platelets into the platelet concentrate was 93 ± 4 percent and 52 ± 13 percent, respectively, of the original value. White cells in the packed red cells were 70 ± 28 × 107 with recovery of 32 ± 9 percent of the original value, and the lymphocytes in the white cells were only 7 ± 5 × 107 (7 ± 4% of the original value). White cell contamination of platelet concentrate was below the threshold of white cell detection by the microcell counter (less than 300 cells per μl of concentrate).

Clinical evaluation of repeat apheresis donors in Japan.

AbstractBackground and Objectives: To ascertain the safety of repeat apheresis donation, hematological and biochemical tests were performed on 511 donors with a donation rate of over 6 times per year for a period of 12–19 months. Materials and Methods: Repeat donors who had apheresis more than 6 times in the previous year were chosen. Data for the repeat donors at the start of the experiments were compared with those at the end of the end of the study. Blood samples were taken prior to donation. Serum protein, albumin, immunoglobulin G, A, and M, serum ferritin levels were determined by biochemical tests. Results: When compared to prospective donors of 400 ml, WBC, lymphocytes, and serum ferritin levels were lower in a roughly frequency‐dependent manner in female and male donor groups at the beginning of the study. All the data for the male group remained almost constant with increasing frequency of apheresis donation. However, in the female group, ferritin levels significantly decreased with over 21 donations. Conclusions: The present data showed that the serum ferritin level of the female donors decreased the most with increasing frequency of apheresis donation. The cumulative RBC left in the collecting chamber and for the laboratory test is discussed in relation to a possible cause of iron deficiency in frequent apheresis donors.

Allolymphocytotoxic antibodies in sera from HTLV-I-associated myelopathy/tropical spastic paraparesis patients — putative anti-HLA antibodies

Thirty-one HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) patients and 34 asymptomatic carriers were studied in order to investigate the presence of anti-lymphocyte antibodies in their sera. 32% of the patients produced allolymphocytotoxic antibodies reactive to peripheral blood lymphocytes (mainly T cells) as well as to B cells, which belonged to the IgM class, reacted under warm and cold conditions, and showed specificity to multiple HLA antigens, especially HLA A2, A26, A33, B7, B27, B35, B48, B61 and Cw3. Moreover, their reactivities could be absorbed by pooled platelets. Therefore, it may be suggested that either anti-HLA class I antibodies or antibodies cross-reactive to HLA were present. On Western blot analysis, the HAM/TSP cytotoxic sera reacted to the 110 kDa molecules of lymphocyte lysates. In the case of carriers, weak cytotoxic antibodies were detectable in 14% of the sera. No multispecificity to HLA class I antigens was observed and there was no reactivity to the 110 kDa band. The production of allolymphocytotoxic antibodies in HAM/TSP patients may therefore be explained by the presence of allogeneic-like stimuli in this disease. HTLV-I infected cells, expressing altered HLA antigens could possibly account for this stimulation and also contribute to the alteration of immune functions in HAM/TSP, and thus play an important role in its pathogenesis.

Analysis of HTLV-1 proviral load (PVL) and antibody detected with various kinds of tests in Japanese blood donors to understand the relationship between PVL and antibody level and to gain insights toward better antibody testing

Adult T‐cell leukemia/lymphoma (ATL) occurs in approximately 5% of individuals infected with human T‐cell leukemia virus type 1 (HTLV‐1). A high proviral load (PVL; more than four copies per 100 peripheral blood mononuclear cells (PBMCs) or 1.6 copies per 100 blood leukocytes) and being male are risk factors for ATL development. Whether anti‐HTLV‐1 antibody level is related to such risk is unknown. Here, PVL and antibody levels were examined using real‐time PCR and other tests in 600 HTLV‐1 positive screened Japanese blood donors to understand the relationship between PVL and antibody level in asymptomatic carriers and to gain insights toward better antibody testing for HTLV‐1 infection. The 430 donors in whom proviral DNA was detected were considered as true positives for HTLV‐1 infection. Among donors aged 40 years or older, more males than females had a PVL corresponding to more than 1.6% infected leukocytes, and an antibody titer below the median (P = 0.0018). In antibody tests using an HTLV‐1 positive cell line or Env antigens there was a large discrepancy in antibody titer among 13 provirus‐positive samples, probably suggesting that antibody‐based screening tests should incorporate multiple HTLV‐1 antigens, such as Gag and Env antigens.

17beta-estradiol (E2) abrogated osteolysis induced by HTLV-1 Env protein in vivo

HTLV-1 is the causative retrovirus of ATL. Eighty percent of ATL patients develop hypercalcemia, a severe complication resulting from bone resorption. PTHrP is known as the major pathogenic factor to hypercalcemia associated malignancies. Recently, we observed hypercalcemia in rabbits with antibody against central region of HTLV-1 Env Gp46 (gp46-197), which shows a homology with osteoprotegerin (OPG), an inhibitor to the maturation of osteoclasts. To observe the effect of gp46197 synthetic peptide on osteogenesis, the peptide was intraperitoneally administered to mice in the presence or absence of co-administration of E2. Seven days later, serum E2 level and bone mineral density (BMD) were measured. Both of males and females exhibited a clear reduction in E2 and BMD by the gp46-197 peptide. Histomorphological analyses revealed that the peptide induced the loss of Ob.S/BS and increase in ES/BS, N. Oc/B.Pm, and Oc.S/BS. These results suggest that the structural mimicry of gp46-197 inhibits the OPG function and promote the maturation and stimulation of osteoclasts in vivo. Co-administration of E2 abrogated the adverse effects by gp46-197. This is a novel mechanism of osteolysis directly induced by HTLV-1 structural protein. We presume the therapeutic potential of E2 for the treatment of hypercalcemia in ATL patients.

Automated device for preparation of white-cell-poor blood components.

We performed to prepare white-cell-poor blood components using automatic device, which was made up a optic sensor detecting the interface of red cells and plasma, a sensor weighing the plasma and huffy coat fraction, and three clamps connecting these sensors. This device was enable to separate 400ml whole blood into packed red cells, plasma and huffy coat fraction involved the platelets within about 3 minutes. White cell contamination in packed red cells was 59±19×107 with recovery of 24±4% of the original value, and the lymphocytes in the white cells were only 3±1×107 (2±1% of the original value).White cell contamination of platelet concentrate was below the threshold of white cell detection by the microscopic observation (less than 100 cells per μl of concentrate). These results suggest that this automatic device was enable to introduce the procedure for preparation of white-cell poor blood components as a routine operation in blood center.