Phenotypic diversity within clones of human normal t cells

Human normal T cells were selected for in vittro cloning according to the expression of T4, T8 or T10 antigens on individual cells. Clones were produced from each of these cells irrespective of the antigenic phenotype of the parental cell. The cloned progeny manifested, in many cases, shifts in antigen expression. Thus, T4⁺T8^? cells gave clones expressing predominantly T4^?T8⁺ and vice versa. The clonal expression of T4 and T8 seemed to be mutually exclusive. Antigenic shifts were recorded also in clones derived from T4^?T8^?T10^? cells, resulting in T10⁺ clones which were also either T4⁺ or T8⁺ and from T4⁺T8^?T10⁺ cloned cells yielding clones of either T4⁺ or T8⁺ cells. Testing functional properties we found that NK activity was mediated not only by T10⁺ cells but also, in some cases, by T4⁺ and T8⁺ cells. Moreover, TCGF production, which may reflect helper activity, was mediated not only by T4⁺ cells. Only the cytotoxic (CTL) activity seems to be confined to the T8 phenotype. Thus, it appears that T antigens, which seemed to be molecular markers of differentiation, are not markers for terminal differentiation and do not always reflect defined functional properties. These conclusions are drawn from cloning of normal T cells which manifest properties different from those of T-cell lines or T hybridomas.     

2010美国心脏病学会基金会/美国心脏学会关于氯吡格雷的应用警告概述

2010年8月,美国心脏病学会基金会（ACCF）、美国心脏学会（AHA）共同发布了美国食品药品管理局（FDA）关于氯吡格雷的＂盒装警告＂,主要针对医师和患者提出建议,其内容包括：通过检测药物基因型以明确患者氯吡格雷的代谢变化,患者不良反应的风险,基因多态性对氯吡格雷的代谢及临床影响。     

L一粘附素单抗在缺血再灌注损伤中的作用

目的：中性粒细胞粘附在缺血再灌注损伤中有非常重要的作用。本文用ＳＤ大鼠趾长屈肌缺血再灌注损伤模型,观察Ｌ一粘附素单抗ＬＡＭ１—１１６在缺血再灌注损伤中的作用。方法：３０只ＳＤ大鼠被均分为２组：ＬＡＭ１—１１６组和生理盐水对照组。每只大鼠的一侧趾长屈肌作为正常对照,另外一侧进行 ３ ｈ缺血 ４ ｈ再灌注。结果：ＬＡＭ１— １１６组实验侧的髓过氧化物酶为正常的２倍（２．３±２．２）,生理盐水对照组则为正常的２８倍（２７．５±１１．７）（Ｐ＜０．００１）;ＬＡＭ１—１１６组的湿重比（１．１０± ０．１０）、疲劳肌力（７７． １％±１２．１％）与对照组相比（分别为 １． ２３± ０． １０和 ４９． ７％± ９ ．３％）明显改善（Ｐ＜ ０．０５）;组织学上,ＬＡＭ１—１１６组的中性粒细胞局部浸润显著减少,水肿减轻。结论：通过 Ｌ－粘附素单克隆抗体 ＬＡＭ１— １１６阻断 Ｌ－粘附素的功能,可以有效地降低中性粒细胞在再灌注肌肉中的浸润,防止组织水肿,从而改善肌肉的功能。     

Ex vivo expansion with stem cell factor and interleukin-11 augments both short-term recovery posttransplant and the ability to serially transplant marrow

The characterization of many cytokines involved in the control of hematopoiesis has led to intense investigation into their potential use in ex vivo culture to expand progenitor numbers. We have established the optimum ex vivo culture conditions that allow substantial amplification of transient engrafting murine stem cells and which, simultaneously, augment the ability to sustain serial bone marrow transplantation (BMT). Short-term incubation of unfractionated BM cells in liquid culture with stem cell factor (SCF) and interleukin-11 (IL- 11) produced a 50-fold amplification of clonogenic multipotential progenitors (CFU-A). Following such ex vivo expansion, substantially fewer cells were required to rescue lethally irradiated mice. When transplanted in cell doses above threshold for engraftment, BM cells expanded ex vivo resulted in significantly more rapid hematopoietic recovery. In a serial transplantation model, unmanipulated BM was only able to consistently sustain secondary BMT recipients, but BM expanded ex vivo has sustained quaternary BMT recipients that remain alive and well more than 140 days after 4th degree BMT. These results show augmentation of both short-term recovery posttransplant and the ability to serially transplant marrow by preincubation in culture with SCF and IL-11.     

Quality control of multidrug resistance assays in adult acute leukemia: correlation between assays for P-glycoprotein expression and activity

We have compared multiple assays for the P-glycoprotein (Pgp/MDR1) phenotype in fresh and thawed adult acute leukemia to validate and quantitate measures for the expression and function of Pgp. The results are related to the Pgp-expressing KB8 and KB8-5 call lines. The most sensitive assay was the measurement of modulation of the rhodamine 123 (R123) fluorescence by 2 micromol/L PSC833, followed by the modulation of the probe calcein-AM. We also found a good intralaboratory and interlaboratory correlation between the values of the R123/PSC833 assay for fresh as well as thawed samples. In addition, the affects of PSC833 on 3H-daunorubicin (DNR) accumulation, DNR fluorescence, and 3H- vincristine accumulation were very similar. The correlation between the DNR/PSC833 and R123/PSC833 test was r = .86 (N = 51). The modulation of drug accumulation by 8 micromol/L verapamil was the some as the PSC833 effect for DNR (117%, N = 21), but was higher for vincristine in every single case (161% v 121%, N = 22; P< .001), indicating additional verapamil effects, not related to Pgp. The correlation of the staining of viable cells for Pgp with the monoclonal antibody MRK16 was r = .77 (N = 52) for the R123/PSC833 functional test and r = .84 (N = 50) for the DNR/PSC833 test. From these results it could be calculated that a maximal increase of the mean DNR accumulation of about 50% can be achieved by blocking Pgp pump activity with PSC833 in leukemic blast samples with the highest mean Pgp expression. Subpopulations of blast calls with higher Pgp activity are likely to be present. Their relevance has to be studied further. The methods outlined here allow the reliable, quantitative monitoring of the Pgp/MDR1 phenotype in leukemias in multicentered, clinical Pgp modulation studies.     

Analysis of clonal rearrangements of the Ig heavy chain locus in acute leukemia

Clonal rearrangements of the Ig heavy chain (IGH) locus occur in nearly all cases of B-cell precursor acute leukemia (BCP-ALL). Some of these rearrangements may be detected by polymerase chain reaction (PCR) using VH gene framework III (FRIII) and JH consensus primers. However, about 20% of BCP-ALLs fail to amplify with this technique. To determine the causes of these PCR failures and to investigate any possible association with specific subgroups of disease, we analyzed 72 acute leukemias of defined immunophenotype and cytogenetics, comparing FRIII with VH-family leader-specific PCR methods and Southern blotting. Of 37 BCP-ALL cases, 6 (16.2%) failed totally to amplify with FRIII and JH primers. None of these cases amplified with VH leader primers. Additionally, all cases retained germline VH6 genes and 5 of 11 rearranged alleles amplified with a consensus DH primer, indicating that these rearrangements represented biallelic DH-JH recombinations. Among the 6 FRIII and VH leader PCR-negative BCP-ALL cases, there was no common immunophenotype or consistent cytogenetic abnormality, although all showed structural chromosomal abnormalities and 3 of 5 successfully karyotyped had abnormalities of chromosome 12p. 13 cases with t(9;22)(q34;q11) Philadelphia chromosome-positive [Ph+]) and IGH rearrangements (9 BCP-ALL and 4 biphenotypic cases) were also analyzed. Of 23 rearranged IGH alleles, 19 (82%) were positive by FRIII PCR, and all 4 remaining alleles were amplified by VH leader primers. Use of the leader primers in these Ph+ cases also detected 3 additional clonal rearrangements that were not anticipated from Southern blotting; such unexpected bands were not observed in 21 other Ph- cases. The additional bands represented "new" and unrelated VH rearrangements rather than VH-VH replacement events. We conclude that biallelic DHJH rearrangements occur in a subgroup of BCP-ALL; in these cases, the activation of the full VHDHJH recombination mechanism had not occurred. Therefore, these cases of BCP-ALL were arrested at an early stage of B- cell differentiation. In contrast, all Ph+ BCP-ALLs and biphenotypic acute leukemias, which may represent the transformation of multipotent hemopoietic stem cells, had undergone VHDHJH recombination. Of 9 Ph+ BCP-ALL cases, 3 also showed ongoing VHDHJH rearrangement, reflecting the persistent expression of the VHDHJH recombinase. Finally, sequence analysis of 33 rearranged VHDHJH genes showed that only 3 including 2 Ph+ BCP-ALL maintained an intact open-reading frame. Loss of the open- reading frame occurred not only because of out-of-frame VHDH and DHJH joining, but also because of VH gene mutation and deletion. These data show that most BCP-ALLs may represent the neoplastic transformation of BCPs destined to die in the bone marrow.