Immature Platelet Fraction and Its Kinetics in Neonates.

Thrombocytopenia is a common abnormality encountered in the neonatal period, and immature platelet fraction (IPF) may be an informative indicator of thrombopoiesis; however, data on IPF in neonates are scarce. To define reference intervals (RIs) and factors affecting IPF in neonates, we measured the IPF of 533 consecutive neonates. With a multiple regression analysis of 330 newborns with normal platelet counts at birth, premature delivery, neonatal asphyxia, intrauterine infection, chromosomal abnormalities, and respiratory disorders were identified as independent factors for IPF%. The RIs of IPF% and absolute IPF value in neonates were determined to be 1.3% to 5.7% and 3.2 to 14.5×10 9 /L, respectively. On day 14 after birth, IPF% increased to twice the value at birth and thereafter returned to the previous value on day 28. Reticulocyte counts, in contrast, were the lowest at day 14. IPF% was increased in 16 thrombocytopenic patients with various clinical conditions, especially those with immune-mediated thrombocytopenia. IPF in neonates may be evaluated essentially based on the same RIs as in adults, although some precautions must be taken when evaluating IPF in neonates in the first 2 weeks of life. IPF may be useful for evaluating thrombopoiesis and thrombocytopenia in neonates.

Basophil activation test for allergic and febrile non-haemolytic transfusion reactions among paediatric patients with haematological or oncological disease.

BACKGROUND AND OBJECTIVES: Allergic transfusion reactions (ATRs) and febrile non-haemolytic transfusion reactions (FNHTRs) are common, although their mechanisms remain unclear. Immunoglobulin E (IgE)-mediated type I hypersensitivity may be involved in the pathogenesis of ATR. A basophil activation test (BAT) may help elucidate this process. MATERIALS AND METHODS: The BAT was based on peripheral blood samples from paediatric patients with a haematological or oncological disease and on samples of residual blood products transfused in each case. Dasatinib was used to evaluate whether basophil activation was mediated by an IgE-dependent pathway. RESULTS: Twenty-seven patients with and 19 patients without ATR/FNHTR were included in this study, respectively. The median BAT values associated with ATR- (n = 41) and FNHTR-causing (n = 5) blood products were 22.1% (range = 6.1%-77.0%) and 27.8% (range = 15.2%-47.8%), respectively, which were higher than the median value of 8.5% (range = 1.1%-40.9%) observed in blood products without a transfusion reaction. Dasatinib suppressed basophil activity. BAT values were comparable in patients with ATR regardless of severity. Meanwhile, BAT values analysed with blood products non-causal for ATR/FNHTR were higher in patients with ATR/FNHTR than in those without. CONCLUSION: The IgE-mediated type I hypersensitivity may be involved in the pathogenesis of ATR and FNHTR. BAT analyses may help elucidate the underlying mechanisms and identify patients at risk.

Relationship between allergic transfusion reactions and allergic predisposition among pediatric patients with hematological/oncological disease.

BACKGROUND: Allergic transfusion reactions (ATRs) manifest frequently as transfusion reactions, and their onset may be related to a patient's allergic predisposition. Moreover, although pediatric patients with hematological/oncological disease are more susceptible to ATRs, the relationship between allergic predisposition and ATRs remains to be fully clarified. STUDY DESIGN AND METHODS: Patients who were diagnosed with pediatric hematological/oncological disease and received transfusion at the study institutions were included. We determined patient background information related to their allergy history, measured the levels of allergen-specific immunoglobulin E (IgE) using sera obtained on diagnosis, and analyzed their associations with ATR onset. RESULTS: Of the 363 patients analyzed, 144 developed ATRs. Multivariate analysis identified cases with high basophils in the peripheral blood, and Dermatophagoides pteronyssinus- and egg white-specific IgEs were involved in the development of ATR in all age groups. Meanwhile, a history of food allergies, and positivity for Japanese cypress- and D. pteronyssinus-specific IgEs were risk factors for developing ATRs in the <5 years age group. Moreover, patients aged 5-<10 years with a history of asthma, allergic rhinitis, pollinosis, or atopic dermatitis, and those aged ≥10 years with positivity for dog dander-specific IgE were at risk for developing ATRs. CONCLUSION: The allergic constitution of patients plays a role in ATR onset even in pediatric hematological/oncological diseases. Therefore, advance confirmation of a patient's allergic constitution may partly predict the onset of ATRs. However, since multiple allergic predispositions within complex mechanisms may be involved in the onset of ATRs, further verification is required.

Investigation of patient factors associated with the number of transfusions required during chemotherapy for high-risk neuroblastoma.

BACKGROUND: Blood transfusion is an important supportive care for high-risk neuroblastoma. When the number of transfusions increases, transfusion-associated adverse reactions may be more problematic. However, the factors determining the degree of myelosuppression and the number of transfusions during chemotherapy for high-risk neuroblastoma remain unclear. MATERIALS AND METHODS: We investigated patient factors determining the number of required transfusions in 15 high-risk neuroblastoma patients who received five courses of chemotherapy. Clinical data, cytokine profile and colony-forming assay with bone marrow samples at diagnosis were analysed. RESULTS: The required number of transfusions of both platelets and erythrocytes decreased once in the second course and then increased as the course progressed. The variability among cases increased as the chemotherapy course progressed. In cases of low peripheral blood platelet count and lower fibrinogen level at diagnosis, the number of platelet transfusions was higher during chemotherapy. In contrast, there was a negative correlation between the forming ability of granulocyte-macrophage or erythroid colonies and the number of erythrocyte transfusions in the latter period. CONCLUSION: In the early stages of chemotherapy, bone marrow infiltration in neuroblastoma and/or coagulopathy complication may cause thrombocytopenia and requirement of platelet transfusion; conversely, in the later stages, the number of erythrocyte transfusions may be defined by the patient's inherent hematopoietic ability. These factors may be useful in predicting the required number of transfusions.

Relationship between allergic sensitisation-associated single-nucleotide polymorphisms and allergic transfusion reactions and febrile non-haemolytic transfusion reactions in paediatric cases.

BACKGROUND: Allergic transfusion reactions (ATR) and febrile non-haemolytic transfusion reactions (FNHTR) are common transfusion-related adverse reactions; however, their pathogenesis remains unclear and it is difficult to predict their occurrence. Single-nucleotide polymorphisms (SNP) are related to the onset of various diseases and therapy-related adverse events; therefore, identification of SNP related to transfusion-related adverse reactions may help to elucidate the underlying mechanism and predict the onset of these reactions. MATERIALS AND METHODS: We retrospectively analysed the association between the onset of ATR or FNHTR and 22 allergic sensitisation-related SNP in 219 children (aged ≤20 years) who had haematological and oncological diseases and who had received transfusions of platelets and/or red blood cell concentrates. RESULTS: Among the 219 children, 105 had developed an ATR and/or FNHTR at least once. The patients who developed ATR frequently had a risk allele in rs6473223, while the patients who developed FNHTR frequently had a risk allele in rs10893845. Furthermore, patients who developed ATR accompanied by febrile symptoms also frequently had a risk allele in rs10893845, similar to patients who developed FNHTR. DISCUSSION: The results suggested that allergic sensitisation is associated with the onset of ATR and/or FNHTR in some patients. Although further prospective evaluation is necessary, analysis of these SNP might help to provide safer transfusion therapy by predicting patients at higher risk of transfusion-related adverse reactions and further clarifying the pathogenic mechanism underlying such reactions.

Marked Changes in Serum Amyloid A Distribution and High-Density Lipoprotein Structure during Acute Inflammation.

High-density lipoprotein- (HDL-) cholesterol measurements are generally used in the diagnosis of cardiovascular diseases. However, HDL is a complicated heterogeneous lipoprotein, and furthermore, it can be converted into dysfunctional forms during pathological conditions including inflammation. Therefore, qualitative analysis of pathophysiologically diversified HDL forms is important. A recent study demonstrated that serum amyloid A (SAA) can remodel HDL and induce atherosclerosis not only over long periods of time, such as during chronic inflammation, but also over shorter periods. However, few studies have investigated rapid HDL remodeling. In this study, we analyzed HDL samples from patients undergoing orthopedic surgery inducing acute inflammation. We enrolled 13 otherwise healthy patients who underwent orthopedic surgery. Plasma samples were obtained on preoperative day and postoperative days (POD) 1-7. SAA, apolipoprotein A-I (apoA-I), and apolipoprotein A-II (apoA-II) levels in the isolated HDL were determined. HDL particle size, surface charge, and SAA and apoA-I distributions were also analyzed. In every patient, plasma SAA levels peaked on POD3. Consistently, the HDL apoA-I : apoA-II ratio markedly decreased at this timepoint. Native-polyacrylamide gel electrophoresis and high-performance liquid chromatography revealed the loss of small HDL particles during acute inflammation. Furthermore, HDL had a decreased negative surface charge on POD3 compared to the other timepoints. All changes observed were SAA-dependent. SAA-dependent rapid changes in HDL size and surface charge were observed after orthopedic surgery. These changes might affect the atheroprotective functions of HDL, and its analysis can be available for the qualitative HDL assessment.

Highly oxidized low-density lipoprotein does not facilitate platelet aggregation.

OBJECTIVE: This study aimed to examine whether oxidized low-density lipoprotein (oxLDL) facilitates platelet aggregation, which is one cause for development of cardiovascular disease. METHODS: The susceptibility of platelets to aggregation was monitored by light transmittance aggregometry and a laser light scattering method using low-density lipoprotein (LDL) and oxLDL as agonists. ß-thromboglobulin (ß-TG) levels released from platelets were also measured after incubation with or without oxLDL. RESULTS: Platelet aggregation was suppressed by oxLDL as estimated by maximum light transmission. Additionally, adenosine diphosphate-induced further aggregation was slightly reduced by the presence of oxLDL. Aggregation levels of a low number of platelets, which was determined by the laser light scattering method, were lower upon addition of oxLDL compared with unoxidized LDL. After a short time of incubation, oxLDL increased secreted ß-TG levels in platelet-rich plasma. However, further incubation with oxLDL caused relatively lower secreted ß-TG levels compared with incubation with unoxidized LDL. This fluctuation was not due to ß-TG degradation by oxLDL. CONCLUSIONS: Levels of oxLDL in vitro weakly activate platelets at an early stage, but then inhibit platelet function, such as aggregation and ß-TG secretion.

Cholesterol transport between red blood cells and lipoproteins contributes to cholesterol metabolism in blood.

Lipoproteins play a key role in transport of cholesterol to and from tissues. Recent studies have also demonstrated that red blood cells (RBCs), which carry large quantities of free cholesterol in their membrane, play an important role in reverse cholesterol transport. However, the exact role of RBCs in systemic cholesterol metabolism is poorly understood. RBCs were incubated with autologous plasma or isolated lipoproteins resulting in a significant net amount of cholesterol moved from RBCs to HDL, while cholesterol from LDL moved in the opposite direction. Furthermore, the bi-directional cholesterol transport between RBCs and plasma lipoproteins was saturable and temperature-, energy-, and time-dependent, consistent with an active process. We did not find LDLR, ABCG1, or scavenger receptor class B type 1 in RBCs but found a substantial amount of ABCA1 mRNA and protein. However, specific cholesterol efflux from RBCs to isolated apoA-I was negligible, and ABCA1 silencing with siRNA or inhibition with vanadate and Probucol did not inhibit the efflux to apoA-I, HDL, or plasma. Cholesterol efflux from and cholesterol uptake by RBCs from Abca1+/+ and Abca1-/- mice were similar, arguing against the role of ABCA1 in cholesterol flux between RBCs and lipoproteins. Bioinformatics analysis identified ABCA7, ABCG5, lipoprotein lipase, and mitochondrial translocator protein as possible candidates that may mediate the cholesterol flux. Together, these results suggest that RBCs actively participate in cholesterol transport in the blood, but the role of cholesterol transporters in RBCs remains uncertain.

Analysis of the clinical characteristics of pediatric patients who experience ifosfamide-induced encephalopathy.

BACKGROUND: Several kinds of pediatric hematological and/or malignant diseases are treated with chemotherapy regimens including ifosfamide (IFO). IFO-induced encephalopathy (IIE) is one of the serious side effects, but there is not enough evidence regarding the clinical features of IIE in children. PROCEDURE: We performed a retrospective study on pediatric patients treated with chemotherapy regimens, including IFO, at a single center. We recorded the clinical characteristics of all patients; we compared the clinical characteristics between patients who developed IIE and those who did not. RESULTS: In total, 88 patients received a chemotherapy regimen including IFO. IIE developed in seven patients (8.0%). The median age of patients at the time of IIE development was 4.3 (range 1.4-6.5) years in the younger population. Six of seven patients with IIE improved with supportive therapy only; however, one patient died due to heart failure. Overall survival was not different between the two groups. Multivariable analysis revealed that the co-administration of cisplatin (CDDP) or carboplatin (CBDCA) was a significant risk factor associated with IIE. Although there was no significant difference in laboratory data between the groups before chemotherapy, patients who developed IIE showed exacerbation in several laboratory tests, including those for renal and liver functions. CONCLUSIONS: Renal dysfunction caused by the combination of nephrotoxic agents (IFO and CDDP/CBDCA) seems to be important for the development of pediatric IIE. It was thought to be difficult to predict IIE onset based on laboratory data before the initiation of chemotherapy regimens; however, careful observation of laboratory data during IFO chemotherapy regimens may help predict IIE onset and facilitate early treatment.

Red blood cells participate in reverse cholesterol transport by mediating cholesterol efflux of high-density lipoprotein and apolipoprotein A-I from THP-1 macrophages.

High-density lipoprotein (HDL) plays a main role in reverse cholesterol transport (RCT), one of the most important functions for preventing atherosclerosis. Recent reports have shown that red blood cells (RBCs) can be associated with RCT, an interaction facilitated by albumin. However, the RCT function of RBCs has not been thoroughly elucidated. In this study, the RCT function of RBCs was assessed using cholesterol efflux capacity (CEC) assays, in which [3H]-labeled cholesterol-loaded human acute monocytic leukemia (THP-1) macrophages were incubated with RBCs as a cholesterol acceptor in the presence or absence of HDL or its main component protein apolipoprotein A-I (apoA-I). The CEC of RBCs was found to be dose dependent, enabling uptake of cholesterol from THP-1 macrophages through apoA-I and HDL, and directly from apoA-I and HDL in medium without the presence THP-1 macrophages. Moreover, RBCs could exchange cholesterol with HDL in a bidirectional manner but could only exchange cholesterol with apoA-I in a single direction. Although albumin promoted the movement of cholesterol, synergistic effects were not observed for both apoA-I and HDL, in contrast to previous findings. These results strongly suggested that RBCs may play important roles in RCT by mediating cholesterol efflux as temporary cholesterol storage.

Characterization of the cholesterol efflux of apolipoprotein E-containing high-density lipoprotein in THP-1 cells.

High-density lipoprotein (HDL), also known as antiatherogenic lipoprotein, consists of heterogeneous particles in terms of size, density and composition, suggesting differences among HDL subclasses in characteristics and functions. We investigated the role of apolipoprotein E (apoE)-containing HDL, a minor HDL subclass, in the cholesterol efflux capacity (CEC) of HDL, which is its predominant atheroprotective function. The CEC of apoE-containing HDL was similar to that of apoE-deficient HDL, but the former exhibited a greater rate increase (1.48-fold) compared to that of the latter (1.10-fold) by the stimulation of THP-1 macrophages with the Liver X Receptor (LXR) agonist. No difference in CEC was observed without the LXR agonist between apoA-I, the main apolipoprotein in HDL, and apoE, whereas the increase in CEC in response to treatment with the LXR agonist was greater for apoA-I (4.25-fold) than for apoE (2.22-fold). Furthermore, the increase in the CEC of apoE-containing HDL induced by the LXR agonist was significantly reduced by treatment with glyburide, an inhibitor of ATP-binding cassette transporter A1 (ABCA1). These results suggest that apoE-containing HDL, unlike apoE-deficient HDL, is involved in cholesterol efflux via ABCA1.

Dihydro-sphingosine 1-phosphate interacts with carrier proteins in a manner distinct from that of sphingosine 1-phosphate.

Dihydro-sphingosine 1-phosphate (DH-S1P) is an analog of sphingosine 1-phosphate (S1P), which is a potent lysophospholipid mediator. DH-S1P has been proposed to exert physiological properties similar to S1P. Although S1P is known to be carried on HDL via apolipoprotein M (apoM), the association between DH-S1P and HDL/apoM has not been fully elucidated. Therefore, in the present study, we aimed to elucidate this association and to compare it with that of S1P and HDL/apoM. First, we investigated the distributions of S1P and DH-S1P among lipoproteins and lipoprotein-depleted fractions in human serum and plasma samples and observed that both S1P and DH-S1P were detected on HDL; furthermore, elevated amounts of DH-S1P in serum samples were distributed to the lipoprotein-depleted fraction to a greater degree than to the HDL fraction. Concordantly, a preference for HDL over albumin was only observed for S1P, and not for DH-S1P, when the molecules were secreted from platelets. Regarding the association with HDL, although both S1P and DH-S1P prefer to bind to HDL, HDL preferentially accepts S1P over DH-S1P. For the association with apoM, S1P was not detected on HDL obtained from apoM knockout mice, while DH-S1P was detected. Moreover, apoM retarded the degradation of S1P, but not of DH-S1P. These results suggest that S1P binds to HDL via apoM, while DH-S1P binds to HDL in a non-specific manner. Thus, DH-S1P is not a mere analog of S1P and might possess unique clinical significance.

[Successful readministration of L-asparaginase after development of severe hypertriglyceridemia in a young adult with T-cell acute lymphoblastic leukemia].

A 24-year-old male patient with T-cell acute lymphoblastic leukemia was diagnosed with severe hypertriglyceridemia after the sixth administration of L-asparaginase during remission-induction therapy of the Japan Adult Leukemia Study Group (JALSG) -ALL 202-U protocol. Lipoprotein analysis revealed type IV hyperlipidemia, which is associated with a relatively low risk for pancreatitis. Hypertriglyceridemia immediately resolved after discontinuing L-asparaginase and beginning a lipid-restricted diet. The patient did not develop any severe complications of hypertriglyceridemia (e.g., pancreatitis and thrombosis) ; therefore, L-asparaginase could be readministered according to the treatment protocol. Four adult patients with L-asparaginase-induced severe hypertriglyceridemia have been reported to date; however, none of the reports indicated that L-asparaginase had been readministered. Thus, this is the first report of a patient receiving such readministeration. In order to evaluate the safety of continuing L-asparaginase, it is considered necessary to accumulate similar readministration cases.

Lipoprotein-X in cholestatic patients causes xanthomas and promotes foam cell formation in human macrophages.

BACKGROUND: Lipoprotein-X (Lp-X) is an abnormal phospholipid-rich lipoprotein found in patients with cholestatic liver disease. Some patients exhibit skin xanthomas and severe hyperlipidemia. OBJECTIVE: We investigated whether Lp-X induces foam cell formation in human-derived macrophages. METHODS: To compare the atherogenic properties of Lp-X and modified LDL, we isolated Lp-X from 2 patients who had drug-induced cholestasis and xanthoma striata in the interphalangeal folds. We prepared oxidized LDL and acetylated LDL from healthy volunteers for the positive control experiments. RESULTS: When human monocyte-derived macrophages were incubated with these lipoproteins, the isolated Lp-X induced more prominent lipid accumulation than oxidized LDL or acetylated LDL. One case underwent liver biopsy, with the bile ducts showing marked damage, fulfilling the criteria for vanishing bile duct syndrome. The other case was clinically diagnosed as drug-induced hypersensitivity syndrome. In both cases, Lp-X levels decreased markedly and the xanthomas disappeared completely after the improvement of cholestasis. CONCLUSION: This study indicates that Lp-X induces foam cell formation in human-derived macrophages. Our findings strongly suggest that persistently elevated Lp-X may cause xanthomas.

Dysregulation of the DNA Damage Response and KMT2A Rearrangement in Fetal Liver Hematopoietic Cells.

Etoposide, a topoisomerase 2 (TOP2) inhibitor, is associated with the development of KMT2A (MLL)-rearranged infant leukemia. An epidemiological study suggested that in utero exposure to TOP2 inhibitors may be involved in generation of KMT2A (MLL) rearrangement. The present study examined the mechanism underlying the development of KMT2A (MLL)-rearranged infant leukemia in response to in utero exposure to etoposide in a mouse model. Fetal liver hematopoietic stem cells were more susceptible to etoposide than maternal bone marrow mononuclear cells. Etoposide-induced Kmt2a breakage was detected in fetal liver hematopoietic stem cells using a newly developed chromatin immunoprecipitation (ChIP) assay. Assessment of etoposide-induced chromosomal translocation by next-generation RNA sequencing (RNA-seq) identified several chimeric fusion messenger RNAs that were generated by etoposide treatment. However, Kmt2a (Mll)-rearranged fusion mRNA was detected in Atm-knockout mice, which are defective in the DNA damage response, but not in wild-type mice. The present findings suggest that in utero exposure to TOP2 inhibitors induces Kmt2a rearrangement when the DNA damage response is defective.

The role of alloantibodies against human platelet antigen-15 in multiply platelet transfused patients.

BACKGROUND: Several studies have documented the role of antibodies against human platelet (PLT) antigen (HPA)-15 in alloimmune-mediated thrombocytopenia including neonatal alloimmune thrombocytopenia, PLT transfusion refractoriness (PTR), and posttransfusion purpura in Caucasian persons. However, the relevance of anti-HPA-15 in PTR among the Japanese population is still unclear. STUDY DESIGN AND METHODS: The sera of 305 multiply PLT transfused (MPT) patients, previously investigated for the presence of human leukocyte antigen (HLA) and HPA antibodies by mixed passive hemagglutination, were reexamined for the presence of HPA-15 alloantibodies, using the monoclonal antibody-specific immobilization of PLT antigens (MAIPA) technique. RESULTS: Among the 305 MPT samples, antibodies against HPA-15 alloantigen was detected in seven (2.3%), two (0.66%) being anti-HPA-15a and five (1.64%) being anti-HPA-15b. Additionally, one case of CD109 panreactive antibody was found (0.33%). Among them, one aplastic anemia patient with blood group O developed multispecific anti-HLA and anti-HPA-15b alloantibody after MPTs. However, transfusion with HLA-matched PLTs of blood group AB did not result in adequate PLT count increment. Analysis of the possible influence of immune anti-A and anti-B by the MAIPA assay resulted negative, indicating that anti-HPA-15b is responsible for the refractory state in this patient. CONCLUSION: In this study, we found alloimmunization against HPA-15a and -15b in Japanese populations and demonstrated the relevance of these antibodies in a patient with PTR.

Sphingosine 1-phosphate release from platelets during clot formation: close correlation between platelet count and serum sphingosine 1-phosphate concentration.

BACKGROUND: Sphingosine 1-phosphate (Sph-1-P), abundantly stored in platelets and released extracellularly upon activation, plays important roles as an extracellular mediator by interacting with specific cell surface receptors, especially in the area of vascular biology and immunology/hematology. Although the plasma Sph-1-P level is reportedly determined by red blood cells (RBCs), but not platelets, this may not be true in cases where the platelets have been substantially activated. METHODS AND RESULTS: We measured the Sph-1-P and dihydrosphingosine 1-phosphate (DHSph-1-P) levels in serum samples (in which the platelets had been fully activated) from subjects with (n = 21) and without (n = 33) hematological disorders. We found that patients with essential thrombocythemia exhibited higher serum Sph-1-P and DHSph-1-P concentrations. The serum Sph-1-P concentration was closely correlated with the platelet count but was very weakly correlated with the RBC count. Similar results were obtained for DHSph-1-P. The serum Sph-1-P and DHSph-1-P levels were inversely correlated with the level of autotaxin (ATX), a lysophosphatidic acid-producing enzyme. A multiple regression analysis also revealed that the platelet count had the greatest explanatory impact on the serum Sph-1-P level. CONCLUSIONS: Our present results showed close correlations between both the serum Sph-1-P and DHSph-1-P levels and the platelet count (but not the RBC count); these results suggest that high concentrations of these sphingoid base phosphates may be released from platelets and may mediate cross talk between platelet activation and the formation of atherosclerotic lesions.

Serum lysophospholipase D/autotaxin may be a new nutritional assessment marker: study on prostate cancer patients.

BACKGROUND: It is now established that the bioactive lipid lysophosphatidic acid (LPA) contributes to cancer initiation, progression and metastasis, including those of prostate cancer. LPA is produced in the serum and plasma mainly by conversion from lysophospholipids through the action of lysophospholipase D (lysoPLD), which is identical to the soluble form of autotaxin (ATX) originally isolated as a tumour cell motility-stimulating factor. In this study, we evaluated the usefulness of lysoPLD/ATX activity as a diagnostic marker. METHODS: The serum lysoPLD activity, assessed by measuring choline liberation from the substrate lysophosphatidylcholine, was measured in patients with prostate cancer and compared with the concentrations of prostate-specific antigen (PSA) and conventional nutritional assessment markers. RESULTS: The serum lysoPLD activity in prostate cancer patients was not statistically different from that in the controls. Consistent with this, there was no correlation between the serum lysoPLD activity and the serum PSA concentrations. However, the lysoPLD/ATX activity did decrease after operation in the prostate cancer patients and seemed to reflect the postoperative damage or the nutritional status. CONCLUSIONS: We postulate that while the serum lysoPLD/ATX may not be a marker of prostate cancer, it promises to instead be a new marker of nutritional status.

Suppression of lysophosphatidic acid and lysophosphatidylcholine formation in the plasma in vitro: proposal of a plasma sample preparation method for laboratory testing of these lipids.

It is now established that lysophosphatidic acid (LPA) and lysophosphatidylcholine (LPC) play important roles in a variety of biological responses, especially in the area of vascular biology, and determination of their concentrations in the plasma is believed to be clinically relevant. Preparation of the measurement samples is a difficult task, however, because the blood levels of these lipids can be easily increased by in vitro manipulation after venepuncture. In this study, we examined the optimal conditions for the preparation of plasma samples for the measurement of LPA and LPC. It appears that regulation of platelet activation and the enzymatic activity of lysophospholipase D/autotaxin and lecithin-cholesterol acyltransferase is important to suppress the undesirable formation of LPA and LPC after venepuncture. We found that in vitro formation of LPA and LPC was negligible when whole blood samples were mixed with 7.5 mM EDTA plus 10% (v/v) citrate-theophylline-adenosine-dipyridamole (CTAD) and when all of the procedures, including the plasma preparation and preservation until measurement, were performed at 4 degrees C. Thus, although the plasma levels of LPA and LPC can be easily altered, laboratory testing of these important bioactive lipids for clinical purposes may be conducted reliably if the samples are prepared under stringent conditions.

Platelet release of beta-thromboglobulin and platelet factor 4 and serotonin in plasma samples.

OBJECTIVES: Platelet release of alpha granule-derived CXC chemokines and dense granule-derived serotonin in plasma samples was evaluated. METHODS: Concentrations of the CXC chemokines beta-TG and PF4 were assayed by an enzyme immunoassay; serotonin was measured by an HPLC method. RESULTS: Beta-TG and PF4 were more easily released than serotonin by in vitro procedures. Use of the anti-platelet cocktail CTAD and preservation of the samples at 4 degrees C were necessary to accurately measure beta-TG and PF4, but not serotonin. CONCLUSIONS: Assaying serotonin may be useful for assessing platelet activation in vivo as a laboratory test because of facile preparation of plasma samples.