Cloned cellular reagents to define antigens encoded between HLA-DR and glyoxylase

Primed lymphocyte typing reagents have been used to define antigens encoded by genes of a locus (loci) mapping between HLA-DR and glyoxalase I. This locus, which we shall refer to as the third locus of the HLA-D region, has been variously referred to as D beta, PL beta, PL3, and SB. Generating discriminatory primed lymphocyte typing reagents which can be used to define these antigens, however, has been extremely difficult. Donors of responding and stimulating cells for the priming combinations have usually been matched not only for the DR, D, and MB/MT antigens but also for the HLA-A, -B, and -C antigens. Even under these very restricted conditions, not all bulk primed lymphocyte typing reagents that are generated are discriminatory enough to be useful for antigen definition. We have derived "clones" from bulk priming combinations in which stimulator and responder differed for known antigens of this third locus. Even though the bulk reagents that were prepared did not provide discriminatory results, approximately 7-12% of the clones derived from the bulk priming combination proved to be highly discriminatory. We have been able to obtain these results with regard to all three antigens of the third locus so far evaluated. The very ease of screening clones and deriving discriminatory reagents, as compared with screening responder-stimulator combinations, allows the ready derivation of cellular reagents that define the antigens of this third locus.     

Detection of non-HLA-DR determinants by alloreactive lymphocyte clones

Using the soft-agar colony assay, we have generated three MT3-associated clones: HJ1, HJ13, and HJ39, from an MLR combination of two unrelated individuals. Another clone, HJ37, appeared to recognize a novel HLA-D determinant. PLT inhibition studies with monoclonal anti-Ia-like antibodies (Mab) were conducted on clones HJ1, HJ39, and HJ37. Five different anti-DR Mab had no significant inhibitory effect on these clones. On the other hand, two Mab SG171 and Q5/13 which appear to react with DR and MT3 (I-A like) molecules strongly inhibited the two MT3-specific PLT clones. While SG171 and Q5/13 had little effect on HJ37, it was observed that a polymorphic Mab 17.15 had a strong inhibitory effect. These results, in concordance with biochemical data on Ia molecules precipitated by these Mab, suggest that these alloreactive clones may recognize non-DR PLT determinants. They also provide further indirect support that MT3 molecules represent the human homologue of murine I-A molecules.     

Lack of shared lymphocyte-stimulating determinants between H-2I and HLA-D/DR using mouse primed lymphocyte typing cells

A number of anti-H-2 alloantisera containing antibody reactive with I region gene products (Ia) of the major histocompatibility complex cross-react with determinants expressed by human peripheral blood B lymphocytes. Such data have led to the conclusion that Ia and DR antigens share cross-reacting determinants. We have attempted to generate mouse primed T lymphocyte populations specific for defined I region gene product determinants which concomitantly recognize DR determinants on human peripheral blood leukocytes in primed lymphocyte typing (PLT) analysis. Mouse PLT cells were generated in primary MLC using strain combinations identical to those in which positive mouse/human cross-reacting antisera have been obtained. The resulting PLT cells exhibited strong, yet specific, secondary MLC responses against mouse cells expressing the Ia determinants used as the primary stimulus. In contrast, when examined on panels of human peripheral blood leukocytes, no reactivity was detected. This lack of cross-reactivity suggests that mouse T cells primed toward Ia determinants do not regularly recognize cross-reacting determinants of DR or D-associated antigens expressed on human PBLs. Consequently, mPLT cells are not a useful reagent in defining HLA-D region polymorphism.     

Quality validation of platelets obtained from the Haemonetics and Trima Accel automated blood-collection systems

BackgroundPlatelet transfusion is required to treat haemo-oncology or trauma patients. Platelet apheresis (PA) performed with apheresis equipment has increased rapidly in recent years. Leucocyte-reduced platelet apheresis (LRPA) can reduce the risk of platelet refractoriness and febrile nonhemolytic transfusion reactions (FNHTRs) for transfusion. Accordingly, this study aimed to investigate and compare the platelet metabolic and functional responses between PA performed with Haemonetics and LRPA performed with Trima Accel cell separator.MethodsThe qualities of platelets collected through PA and LRPA were evaluated in terms of visual appearance, morphology, platelet-aggregation changes, metabolic activities, and bacterium-screening test during 5-day storage. Statistical analyses included two-sample t-test and generalised estimating equation(GEE) method.ResultsDuring 5-day storage in LRPA, residual leucocytes were all <1.0×10⁶, and the parameters of platelet function were as follows: platelet aggregated to agonists such as adenosine 5′-diphosphate (ADP) and collagen, and the extent of shape change and pO₂ showed no statistically significant difference between PA and LRPA. The hypotonic shock reaction (HSR) on days 0, 1, and 3 were significantly higher in LRPA than in PA (71.78±6.92 vs. 64.10±7.42; P=0.002; 71.53±8.98 vs. 62.96±9.84; P=0.007; 68.05±7.28 vs. 57.76±6.80; P<0.0001, respectively). Values of mean platelet volume (MPV) were statistically larger in PA than in LRPA on days 0, 1, and 3. On day 5, the swirling score was higher in LRPA than in PA. The mean lactate levels had no statistically significant difference between PA and LRPA. Moreover, no growth was observed through bacterium-screening test conducted on 40 samples.ConclusionComparison of LRPA and PA products collected from the Trima Accel and Haemonetics automated blood-collection systems, respectively, revealed that both products possessed good platelet qualities even though additional processes are needed to reduce leucocytes. Furthermore, investigating the outcomes of other apheresis instruments with focus on the safety of donors, products, and recipients is necessary.     

血液透析患者普通肝素与低分子肝素抗凝血小板参数变化

目的探讨维持血液透析患者应用不同分子量肝素抗凝血小板参数变化。方法测定26例应用普通肝素(UFH)及38例应用低分子量肝素(LMWH)抗凝的血液透析患者血小板数量(PLT)及血小板平均体积(MPV),并进行对比,分别与30例健康体检者PLT及MPV进行比较。测定并比较18例患者自UFH改为LMWH抗凝3个月PLT及MPV的变化。结果UFH组PLT及MPV低于对照组(P<0.01);LMWH组PLT及MPV低于对照组,差异有显著意义(P<0.05);在UFH组及LMWH组间,后者PLT及MPV高于前者,(P<0.05);自UFH改为LMWH抗凝后,PLT及MPV有升高,但差异无显著意义(P>0.05)。结论肝素抗凝可致PLT及MPV下降,UFH尤其显著;UFH抗凝致PLT及MPV下降后改用LMWH抗凝并不能使PLT及MPV有明显上升。     

血小板、蛋白激酶C、脂蛋白(a)检测在老年突发性聋诊断中的价值

目的研究血小板(PLT)、蛋白激酶C(PKC)及脂蛋白a[LP(a)]在老年性突发性聋中的变化及作用.方法老年性突发性耳聋和健康对照组各30例,分别检测脂蛋白(a)[LP(a)],总胆固醇(TC)、甘油三脂(TG)、高密度脂蛋白(HDL-C)、低密度脂蛋白(LDL-C)、载脂蛋白AL(apoAI)、载脂蛋白B(apoB)、血小板(PLT)、膜和浆蛋白激酶C(M-PKC和P-PKC),对检测结果做统计学分析.结果突发性耳聋组LP(a)、apoB、TG均明显升高,HDL-C则明显减低,与对照组比较,差异均有显著性(LP(a)t=2.33,P＜0.05.apoB:t=2.83,P＜0.晒.TG:t=2.81,P＜0.05.HDL-C:t=3.58,P＜0.01),PLT计数明显减低,与对照组比较,差异具有显著性t=2.25,P＜0.05.M-PKC活性增高、P-PKC活性减低,与对照组比较,前者差异具有非常显著性(t=3.26,P＜0.01),后者差异具有显著性(t=2.22,P＜0.05).结论老年性突发性耳聋时LP(a)、apoB及M-PKC增高,而PLT减低,在血栓形成过程中起重要的作用,对突发性耳聋的发病起促进作用.     

The genetics of platelet count and volume in humans

The last decade has witnessed an explosion in the depth, variety, and amount of human genetic data that can be generated. This revolution in technical and analytical capacities has enabled the genetic investigation of human traits and disease in thousands to now millions of participants. Investigators have taken advantage of these advancements to gain insight into platelet biology and the platelet’s role in human disease. To do so, large human genetics studies have examined the association of genetic variation with two quantitative traits measured in many population and patient based cohorts: platelet count (PLT) and mean platelet volume (MPV). This article will review the many human genetic strategies—ranging from genome-wide association study (GWAS), Exomechip, whole exome sequencing (WES), to whole genome sequencing (WGS)—employed to identify genes and variants that contribute to platelet traits. Additionally, we will discuss how these investigations have examined and interpreted the functional implications of these newly identified genetic factors and whether they also impart risk to human disease. The depth and size of genetic, phenotypic, and other -omic data are primed to continue their growth in the coming years and provide unprecedented opportunities to gain critical insights into platelet biology and how platelets contribute to disease.     

Toxicology of 3-epi-deoxynivalenol,a deoxynivalenol-transformation product by Devosia mutans 17-2-E-8

Microbial detoxification of deoxynivalenol (DON) represents a new approach to treating DON-contaminated grains. A bacterium Devosia mutans 17-2-E-8 was capable of completely transforming DON into a major product 3-epi-DON and a minor product 3-keto-DON. Evaluation of toxicities of these DON-transformation products is an important part of hazard characterization prior to commercialization of the biotransformation application. Cytotoxicities of the products were demonstrated by two assays: a MTT bioassay assessing cell viability and a BrdU assay assessing DNA synthesis. Compared with DON, the IC₅₀ values of 3-epi-DON and 3-keto-DON were respectively 357 and 3.03 times higher in the MTT bioassay, and were respectively 1181 and 4.54 times higher in the BrdU bioassay. Toxicological effects of 14-day oral exposure of the B6C3F₁ mouse to DON and 3-epi-DON were also investigated. Overall, there were no differences between the control (free of toxin) and the 25 mg/kg bw/day or 100 mg/kg bw/day 3-epi-DON treatments in body and organ weights, hematology and organ histopathology. However, in mice exposed to DON (2 mg/kg bw/day), white blood cell numbers and serum immunoglobulin levels were altered relative to controls, and lesions were observed in adrenals, thymus, stomach, spleen and colon. Taken together, in vitro and in vivo studies indicate that 3-epi-DON is substantially less toxic than DON.     

Hcy联合凝血功能检测在早孕先兆流产中的应用

目的：探讨同型半胱氨酸（ Hcy）联合凝血功能相关指标的检测在早孕先兆流产患者中的应用价值。方法选取59例先兆流产患者为研究组,另选取81例正常妊娠妇女为对照组。测定Hcy、纤维蛋白原（ Fg）、D－二聚体（ D－D）、血小板（PLT）,并对结果进行分析。结果研究组的Hcy（5．53±1．13μmol／L）、Fg（4．00±0．89g／L）较对照组的Hcy（4．63±1．26μmol／L）、Fg（3．45±0．73g／L）明显升高,差异均有统计学意义（t值分别为3．82、4．64,均P＜0．05）;同样,在研究组中D－D和PLT水平也高于对照组,差异均有统计学意义（t值分别为1．75、6．96,均P＜0．05）。经Spearman相关性分析得出,Hcy分别与Fg、D－D及PLT呈正相关关系（r值分别为0．721、0．447、0．368,均P＜0．05）。对59例先兆流产患者进行随访,按妊娠结局分为顺利分娩、难免流产、过期流产3组,单因素方差分析显示3组Hcy、D－D水平比较差异均有统计学意义（ F值分别为59．348、15．916,均P＜0．05）,Fg和PLT水平比较差异均无统计学意义（均P＞0．05）。结论联合检测血Hcy、Fg、D－D、PLT水平可以判断早期妊娠妇女体内的高凝状态,从而为分析早期先兆流产病因提供依据,更加合理地指导临床治疗,以阻止不良妊娠结局的发生。