The immunosuppressive potency of various steroids on peripheral blood lymphocytes, T cells, NK and K cells

The immunosuppressive effect of natural and synthetic steroids was tested in vitro on phytohemagglutinin (PHA) stimulated T lymphocytes and peripheral blood lymphocytes (PBL), as well as on NK and K cell activity. Three groups of steroids, significantly different in their immunosuppressive activity, were identified. Fluorohydrocortisone, and methylprednisolone were highly potent in suppressing PHA stimulation of T lymphocytes and PBL. Hydrocortisone was of intermediate potency, whereas cortisone, dihydrocortisol, and tetrahydrocortisol were of low potency. T lymphocytes were more sensitive to the suppressive effect of fluorohydrocortisone and methylprednisolone than were PBL cultures. NK and K cell activity was suppressed only by the high potency synthetic steroids and even then the suppression of K cell activity was not significant except at high in vitro steroid concentrations. The present findings support the conception that different lymphocyte subpopulations have different susceptibility to the effect of highly potent steroids. Thus, lymphocyte heterogeneity must be taken into account when the immunosuppressive potencies of different glucocorticoids are studied. Furthermore, the findings in different lymphocyte populations ranked the relative in vitro immunosuppressive potency of glucocorticoids different from the relative anti-inflammatory potencies reported in the literature.     

Targeting of peripheral blood T lymphocytes

Conclusions In summary, human single-chain gene transduced T cells were shown: to express the scFv on their surface, to recognize their relevant ligand (tumor-associated antigen) on tumor target cells, to produce cytokines and, to lyze tumor cells. In our earlier review on retargeting T lymphocyte specificity [11], we concluded that a number of questions needed to be answered: (1) Is triggering of cytolysis by CTL or lymphokine production most important to generate anti-cancer effects? Both are important, especially to eliminate bystander cells which do not express tumor-associated antigen [35, 75, 83, 106]. (2) Can targeted CTL traffick and home to the tumor site? Yes, they can. (3) Does humanization of mouse mAbs reduce HAMA responses? Our preliminary experiences suggest that this is the case (unpublished data).Significant progress has therefore been made in the laboratory and in clinical tests, and will continue to be made. We are now preparing for the clinical phase I/H testing of in vivo anti-tumor activity of T lymphocytes retargeted by transfer of chimeric receptor genes encoding Ab-type specificity.     

Comparison of NK activity in mouse spleen and peripheral blood lymphocytes

Natural killer (NK) cells originating in mouse peripheral blood were studied with regard to their lytic activity against YAC-1 target cells and to their expression of asialo-GM1 marker on their surface. In Balb/c, CBA/LAK and A/J mice, PBL were found to be approximately twice as effective as splenocytes. Splenic and peripheral NK cells were shown by flow cytometry to have similar lytic potential per cell; the difference in NK activity found in the spleen and in PBL was solely due to the differences in the size of the NK cell population found in the two sites. Strain distribution of NK activity in PBL followed the same pattern observed in splenocytes. The difference in NK activity between CBA and Balb/c mice was shown to be due to the fact that the lytic potential per NK cell was approximately twice as high in the former.     

人外周血T淋巴细胞胀亡现象的观察

目的观察人外周血T淋巴细胞的胀亡现象,探讨建立T细胞胀亡检测方法.方法密度梯度离心法及尼龙棉柱法分离健康成年人外周血T淋巴细胞,分空白组及地塞米松组,培养后观察细胞光镜、荧光镜及电镜形态学,并用流式细胞仪检测胀亡细胞比例变化.结果①人外周血T淋巴细胞经96小时体外培养,可自然出现典型细胞胀亡形态学改变.②经72小时培养,不同浓度地塞米松组(1×10-6、1×10-5、1×10-4、1×10-3 mol/L)T细胞的胀亡率分别为(3.49±0.42)%、(5.17±0.48)%、(8.44±0.72)%、17.93±1.50)%.③在1×10-5mol/L地塞米松作用下,不同培养时间(48、72、96、120小时)T淋巴细胞的胀亡率分别为(0.53±0.10)%、(6.36±0.80)%、(20.60±1.59)%、25.56±1.76)%.结论人外周血T淋巴细胞存在胀亡现象,地塞米松可诱导人外周血T淋巴细胞胀亡.     

非霍奇金淋巴瘤患者T细胞亚群、NK细胞检测的临床意义

目的：研究非霍奇金淋巴瘤（NHL）患者外周血T淋巴细胞亚群、NK细胞检测结果的变化与该病的关系及与慢性淋巴腺炎患者细胞免疫功能的不同变化。方法：采用流式细胞仪（FCM）检测非霍奇金淋巴瘤（NHL）患者、慢性淋巴腺炎及正常人外周血T淋巴细胞亚群比例、NK细胞的变化。结果：非雹奇金淋巴瘤患者与正常人比较总的T淋巴细胞、辅助性T淋巴细胞及CD4^＋／CD8^＋比值明显下降（P〈0．05），细胞毒性T淋巴细胞明显升高（P〈0．05），NK细胞则无明显变化（P〉0．05）。非霍奇金淋巴瘤患者与慢性淋巴腺炎患者比较，细胞毒性T淋巴细胞、NK细胞明显升高（P〈0.05），而总的T淋巴细胞、辅助性T淋巴细胞无明显改变（P〉0．05），CD4^＋／CD8^＋比值略有下降但无明显统计学意义。结论：非霍奇金淋巴瘤患者细胞免疫功能明显受到抑制，T细胞亚群及NK细胞的检测对NHL的诊断、治疗、预后判断有一定的临床价值。     

自然流产患者外周血与蜕膜NK细胞亚群数量变化的研究

目的：研究自然流产患者外周血与蜕膜组织自然杀伤细胞亚群数量的变化。方法：对36例自然流产患者及30例正常健康早孕者（对照）外周血及蜕膜组织中淋巴细胞进行单克隆抗体标记,应用流式细胞仪定性、定量分析技术对NK细胞（natural killer)亚群进行分析。结果：（1）流产组外周血及蜕膜组织NK细胞CD56＋CD16＋百分比较对照组明显升高。（P＜0．01）,而蜕膜组织中NK细胞CD56＋CD16－百分比明显低于对照组（P＜0．001）。（2）外周血与蜕膜组织NK细胞CD56＋CD16＋亚群存在相关性（γ＝0．516,P＜0．05）。结论：NK细胞亚群的数量变化可以引起蜕膜免疫微环境的失调,导致自然流产。外周血与蜕膜NK细胞CD56＋CD16＋亚群存在正相关关系。     

两种不同HLA-B分子对NK细胞表面受体表达的研究

目的：探讨两种HLA-B分子(HLA-B51和HLA—B39)对NK细胞表面活化性受体CDl6和抑制性受体KIR3DL1表达的调节。方法：采用流式细胞仪检测NK细胞分别与转染HLA-BSl和HLA-B39分子的K562细胞相互作用后，CD16和KIR3DL1的表达情况。结果：外周血淋巴细胞与K562细胞作用24小时后，CD56 CD16 细胞数、KIR3DL1 细胞数均明显增加。与表达HLA-B39的K562细胞相比，表达HLA-B51的K562细胞与外周血淋巴细胞作用后，CD56 CD16 细胞数、KIR3DL1 细胞数均明显下降。结论：NK细胞杀伤靶细胞时，活化性受体CD16表达上调后会伴有抑制性受体KIR3DL1的上调；HLA-B51分子表达在K562细胞后，表达外源性HLA-B51分子的K562细胞与NK细胞作用，NK细胞表面受体KIR3DL1的表达可下调，同时伴有CD16的表达下调。     

脐血来源间充质干细胞可抑制异体T细胞的反应

目的研究脐血间充质干细胞(HUCB-MSCs)对异体T细胞的抑制作用。方法体外培养HUCB-MSCs,流式细胞术测表面标记;取正常人外周血,免疫磁珠分离CD3+T细胞,将分离的CD3+T与HUCB-MSCs 1∶1混合培养5 d,PHA刺激或不刺激,采用3H-TdR掺入法观察T细胞增殖,ELISA方法检测细胞因子,流式细胞术观察细胞凋亡。结果 HUCB-MSCs呈纺锤样的细胞形态,不表达CD14、CD34、CD45、HLA-DR,而表达CD29、CD44、HLA-ABC。HUCB-MSCs抑制PHA引起的T细胞增殖(5 230±550 vs 10 500±800 counts/min,P<0.001);HUCB-MSCs还能抑制异体T细胞分泌IFN-γ(510±60 vs 1 580±100 pg/mL,P<0.001)和TNF-α(590±20 vs 1 180±30 pg/mL,P<0.001),上调IL-4(16.3±8.2 vs 4.1±1.8 pg/mL,P<0.001)和IL-10(105±5 vs 17±2 pg/mL,P<0.001)分泌;HUCB-MSCs不诱导T细胞的凋亡。结论 HUCB-...     

氨茶碱诱导T淋巴细胞自噬现象的观察

目的：观察氨茶碱对分离培养的人外周血T淋巴细胞自噬的影响。方法：密度梯度离心法及尼龙棉柱法分离健康成年人外周血T淋巴细胞,分对照组、氨茶碱组（10^-8～10^-3mol/L,即0.0018～180μg/ml）及地塞米松组（DXM）,培养后用流式细胞术检测自噬率和凋亡率变化。结果：①T淋巴细胞的分离纯度为81.3%～94.5%。②氨茶碱组干预T淋巴细胞培养72小时后,10^-5～10^-3mol/L氨茶碱组自噬率、10^-8～10^-3mol/L氨茶碱组凋亡率与阴性对照组比较差异均有显著性（P值均〈0.05）;各浓度组的自噬率和凋亡率之间均无相关性（P值均〉0.05）;在10^-5mol/L氨茶碱干预下,按不同细胞密度接种T淋巴细胞,观察0、24、48、72小时的自噬率,可见随着细胞密度的减少和时间的延长,其自噬率有增加趋势,但无显著性差异。③DXM干预T淋巴细胞72小时后,其凋亡率与阴性对照组比较有显著差异（P=0.000）,而自噬率无显著差异（P=0.481）。结论：10^-5～10^-3mol/L的氨茶碱可诱导人外周血T淋巴细胞自噬率增加,10^-8mol/L～10^-3mol/L的氨茶碱可诱导人外周血T淋巴细胞凋亡率增加,且自噬与凋亡间相互独立;DXM可诱导人外周血T淋巴细胞凋亡率增加,表明不同的药物可诱导T淋巴细胞启动不同的程序性死亡方式。     

T and B lymphocytes in peripheral blood of splenectomized subjects]

The report is a part of more extensive studies on the role of the spleen in immune processes. Assessing the ability of lymphocytes T to non-immunological binding of sheep erythrocytes and the ability of B cells to bind immunological complexes through the receptor for C3 component of complement, and the presence of immunoglobulins on the surface of lymphocytes B the number of these cells in peripheral blood was determined in 14 healthy individuals who had been splenectomized. In all these subjects the degree of blastic transformation of lymphocytes in PHA-stimulated 3 day-old white blood cell cultures was determined. It was found that the number of T and B lymphocytes in the peripheral blood of splenectomized and non-splenectomized patients was similiar. Lymphocytes obtained from splenectomized patients had however, an impairment of transformation ability after PHA stimulation. It is suggested that, apart from determination of T and B cell pool in the peripheral blood, an evaluation of their transformation ability after PHA stimulation is necessary for assessment of the immunological state in vitro investigations.     

Content of T and B lymphocytes in healthy human peripheral blood

It was shown by the rosette-formation method that the number of T and B lymphocytes in human blood varies with the time of day and season of the year. The number of T cells reaches a maximum in the morning and the number of B cells in the evening. The relative percentage of B cells is higher in the fall than in winter.Presented by Academician of the Academy of Medical Sciences of the USSR V. P. Kaznacheev.Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 7, pp. 872–874, July, 1976.     

T and B lymphocytes in peripheral blood in anesthesiologic personnel.

T and B lymphocytes were determined quantitatively in anesthesiologic personnel by the rosette technique. Percentages of T lymphocytes were diminished, while the lymphatic system of the peripheral blood showed signs of stimulation. The drop in percentages of T lymphocytes was greater in physicians compared with anesthesiologic nurses.     

Immunophenotype of peripheral T lymphocytes, NK cells and expression of CD69 activation marker in patients with recurrent spontaneous abortions, during the mid-luteal phase

PROBLEM: Determination of subpopulations of T lymphocytes, natural killer (NK) and activation status, in peripheral blood during the mid-luteal phase from patients with unexplained recurrent spontaneous abortion (RSA). METHOD OF STUDY: Peripheral blood samples from non-pregnant women with RSA and normal multiparous were taken and evaluated for subpopulations of T lymphocytes: CD4, CD8, ('naive-like' and 'memory-like'), TCR receptor (alphabeta and gammadelta), activation status by CD69(+surface or intracellular)/CD3(+), and NK cells (CD16(+)/CD56(dim)/CD3(-), CD16(+)/CD56 (bright)/CD3(-), CD69(+surface or intracellular)/CD56(+)/CD3(-) cells). RESULTS: The evaluation of T lymphocytes only showed an increase in the expression of CD69 (surface and intracellular) in the RSA group. Additionally, we observed an increase in the total NK cells, CD56(+) NK cells percentages, CD56(dim) NK cells and CD69 NK cells in RSA group. CONCLUSION: These observations support the concept that immunological activation of T lymphocytes and NK cells could be involved in peripheral blood during the mid-luteal phase in patients with unexplained RSA.     

中国恒河猴(Macaca mulatta)外周血CD4+CD25+T淋巴细胞的研究

目的:研究中国恒河猴外周血中CD4 CD25 T淋巴细胞亚群及其分布频率。方法:利用流式细胞术对50只中国恒河猴外周血CD4 CD25 T淋巴细胞进行了分析。结果:发现所有被检测的恒河猴个体中均存在明显的CD4 CD25 T淋巴细胞亚群;CD4 CD25 T淋巴细胞大约占CD4 T淋巴细胞的9.1%(变化范围为2.6%～18.1%);其中CD4 CD25highT淋巴细胞约占2.5%(0.3%～5.5%)。对不同年龄和性别个体中CD4 CD25 T淋巴细胞频率的初步分析未发现统计学上有年龄或性别差异。结论:中国恒河猴可用于与CD4 CD25 T细胞相关的人类疾病的研究中。     

Depletion of circulating T lymphocytes in pregnancy.

Changes were assessed in lymphocyte sub-populations in various stages of human pregnancy. The percentage and absolute number of E-RFC decreased during pregnancy. There was a concomitant rise in the percentages of EAC-RFC and cells bearing SmIg with little change in their absolute numbers. EAC-RFC continued to rise post-natally.     

T and B lymphocytes exert distinct effects on the homeostasis of NK cells

There is growing evidence that lymphocytes impact the development and/or function of other lymphocyte populations. Based on such observations we have tested whether the NK cell compartment was phenotypically and functionally altered in the absence of B and/or T cells. Here we show that T cell deficiency significantly accelerates BM NK cell production and the subsequent seeding of splenic and liver NK cell compartments. In contrast, B cell deficiency reduces splenic NK cell survival. In the absence of T and B cells, the size of the NK cell compartments is determined by the combination of these positive and negative effects. Even though NK cell homeostasis is significantly altered, NK cells from T and/or B cell-deficient mice show a normal capacity to kill a susceptible target cell line and to produce IFN. Nevertheless, we noted that the usage of MHC class I-specific Ly49 family receptors was significantly altered in the absence of T and/or B cells. In general, B cell deficiency expanded Ly49 receptor usage, while T cell deficiency exerted both positive and negative effects. These findings show that B and T cells significantly and differentially influence the homeostasis and the phenotype of NK cells.     

O-Acetyl GD3 ganglioside in human peripheral blood T lymphocytes

O-acetyl GD3 ganglioside is a cell surface molecule of someneural, neural crest and renal cells. Here we show, by usingmAbs specific for O-acetyl GD3 (clone 27A) and flow-cytomotric,biochemical or immunological techniques, that it is also expressedat high intensity level on the surface of 49.6% (median) ofthe CD3⁺ cells (T lymphocytes), at medium level in 16.2% ofthe CD16⁺ (natural killer) cells, at very low level in 51.9%of CD14⁺ cells (monocytes) and in 6.9% of CD20⁺ cells (B lymphocytes),but not in other human blood cells. Of the CD4⁺or CD8^* cells,52.6 or 36.5% respectively were 27A⁺. Furthermore, 81.6% ofthe CD45RO⁺ lymphocytes carried the O-acetyl GD3 ganglloslde.It was not detected in the thymus, although its immediate precursor,the GD3 ganglioside, was present in the meduilary thymocytes,suggesting that O-acetyltransferases are regulated by maturationevents taking place in the periphery. The anti-O-acetyl GD3antibodies induced a strong mitogenic response in cultured peripheralblood mononuclear cells, but not in purified T cells. However,in combination with phorbol myristate acetate the antibodiesinduced proliferation also in purified T cells, suggesting thatprotein kinase C priming is needed for this effect. This andthe restricted expression of O-acetyl GD3 suggest a functionalrole for this ganglioside in T cell subpopulations.     

人外周血T淋巴细胞表型改变与衰老相关性研究

目的：探讨人T淋巴细胞表型随年龄变化的规律，寻找敏感的免疫衰老生物标志。方法：取年轻组(20—35岁)和年老组(50—75岁)志愿者的新鲜肝素抗凝静脉血，进行三色或四色直接免疫荧光染色，获取有核细胞后以流式细胞仪分析T细胞亚群的表型。结果：2个年龄组的总T细胞(CD3^ )、辅助T细胞(CD4^ )以及细胞毒T细胞(CD8^ )亚群的相对百分率没有明显差别，但年老组T细胞上CD3分子的密度(MFI)明显下降；粘附分子CD44与CD62L在3群T细胞上阳性率的差别均无统计意义；而2组间总T细胞、辅助T细胞以及细胞毒T细胞亚群的CD95阳性率都有显著差别，表现为随年龄增加，CD95表达率上升。结论：在所研究的年龄组间，T细胞及其亚群的相对数量没有显著改变，而CD3表达密度随年龄增加而下调；T细胞的凋亡诱导分子CD95阳性率随年龄增加而上升，提示CD95可能是评价免疫衰老的潜在生物标志。     

粘附性NK细胞的制备及免疫学特性初探

目的：初步探讨粘附NK细胞的制备方法及免疫学特性。方法：首先经淋巴细胞分离液，贴壁粘附及尼龙毛柱粘附对外周血NK细胞进行初步分离，然后联合使用Percoll不连续密度梯度离心法和T细胞Panning法进一步纯化NK细胞，应用流式细胞仪鉴定NK细胞并检测其纯度，rhIL-2(6000IU/ml)诱导纯化的NK细胞（R－NK）转变为粘附NK（A－NK）细胞，然后利用细胞计数法，MTT法和RT－PCR方法检测A－NK细胞的增殖和杀伤等免疫学特性。结果：经过纯化，诱导获得较高纯度，较高活性的A－NK细胞。结论：初步建立了利用人外周血NK细胞制备A－NK细胞的方法。