Background: Since sphingosine-1-phosphate (Sph-1-P) plays an important role as an extracellular mediator through interaction with specific cell surface receptors, especially in the area of vascular biology and immunology/haematology, determination of its plasma concentration may become important from the clinical viewpoint. Thus, we attempted to develop a method of measuring the plasma Sph-1-P concentration for use in the clinical laboratory setting. Methods: After two-step lipid extraction, Sph-1-P was coupled with o-phthaldialdehyde, and the resultant fluorescent derivative was separated by high-performance liquid chromatography. C 17 -Sph-1-P was used as the internal standard, instead of dihydrosphingosine-1-phosphate, which had been used previously for the same purpose but was actually detected in plasma. Results: Our procedures for preparing the plasma samples and assay Sph-1-P were found to be satisfactory for clinical laboratory testing. The plasma Sph-1-P concentrations were significantly higher in men (413.1 + 52.0 nmol/L; mean + SD) than in women (352.4 + 39.7 nmol/L). Unexpectedly, strong positive correlations were found between the plasma Sph-1-P concentration and red blood cell (RBC)-related parameters, rather than platelet-related parameters. Conclusions: Our present study confirmed the possibility of the clinical introduction of plasma Sph-1-P measurement, and in addition, suggested that RBCs may be involved in the regulation of plasma Sph-1-P concentrations.

Human neutrophil antigens (HNAs) play an important role in a variety of clinical conditions including immune‐mediated neutropenia, non‐hemolytic transfusion reactions, and transfusion‐related acute lung injury. The aim of this study was to investigate the frequency distribution of HNAs‐1 to ‐5 among the Japanese population. We analyzed samples from 570 healthy Japanese by molecular and serologic techniques to estimate the gene frequencies of HNAs‐1 to ‐5. DNA samples were obtained and typed for the HNA‐1 (n = 523), ‐3 (n = 570), ‐4 (n = 570), and ‐5 (n = 508), by molecular techniques. The HNA‐1 genotype was determined by using a commercial polymerase chain reaction‐reverse sequence‐specific oligonucleotide probes (PCR‐rSSOP) kit. The HNA‐3 to ‐5 genotypes were determined by the PCR‐sequence specific primer (PCR‐SSP), previously described, with a small modification. The HNA‐2a phenotype was determined in 301 donors by granulocyte immunofluorescence test. In Japanese, the gene frequencies of HNA‐1a, ‐1b, and ‐1c were 0.623, 0.377, and 0.000, respectively. The frequency of HNA‐2a phenotype was 0.987, and the gene frequencies of HNA‐3a and ‐3b were 0.654 and 0.346, respectively. HNA‐4a and ‐4b were found at 1.000 and 0.000, respectively, and HNA‐5a and ‐5b at 0.840 and 0.160, respectively. We describe, for the first time, the frequencies of all HNAs (HNA‐1 to ‐5) among the Japanese population. This study will be helpful for the prediction of the risk of alloimmunization to HNA, especially to determine the risk of HNA alloantibody production by transfusion of HNA incompatible blood and feto‐maternal incompatibility.

No abstract

No abstract

The liver plays a major role in the metabolism of plasma high-density lipoprotein (HDL). Several groups have postulated, but others refuted, the existence of a classical membrane receptor which recognizes HDL. In the present study, we identified and purified two liver HDL-binding proteins of 120 kDa (HB1) and 100 kDa (HB2), with apparent specificity for HDL3 devoid of E apolipoprotein. The plasma membrane was the richest source of the HDL-binding protein. Both proteins bound A-I and A-II apolipoproteins and retained HDL-binding activity after final purification. HB1 activity, but not that of HB2, was lost after treatment with beta-mercaptoethanol, but reduction did not change the apparent molecular mass of either protein. Antibodies against HB1 or HB2 did not cross-react, and preliminary structural investigations provide evidence to suggest that HB1 and HB2 are not structurally related. We thus provide evidence for at least two liver plasma-membrane proteins which bind HDL apolipoproteins, suggesting that protein-protein interaction participates to some degree in the mechanism of HDL recognition by cells.