Association of a 70-kDa tyrosine phosphoprotein with the CD16:ζ:γ complex expressed in human natural killer cells

The CD16: ζ: γ receptor complex allows natural killer (NK) cells to recognize and eliminate antibody-coated target cells. Whereas the ectodomain of CD16 is the receptor for Fcγ domains of immunoglobulins, disulfide-linked homo- and heterodimers composed of ζ and γ are required for the cell surface expression, and signal transduction properties of the complex. Engagement of CD16 activates the tyrosine kinase pathway, which induces the tyrosine phosphorylation of several substrates, including the ζ subunit and the phospholipase C γ-1 and γ-2 isoforms. Here we show that CD 16 stimulation of either peripheral blood NK cells, leukemic NK cells, or Jurkat transformants expressing a CD16:ζ:γ receptor complex, results in the tyrosine phosphorylation of a 70 kDa ζ-associated protein (pp70). Similarly, a 70-kDa ζ-associated phosphoprotein in T cells has been shown to be a tyrosine kinase (ZAP-70). Peptide mapping analysis indicates that the 70-kDa ζ-associated phosphoproteins from T cells and NK cells are structurally indistinguishable. We conclude that the CD16:ζ:γ complex may use a ZAP-70-related non-receptor tyrosine kinase, in the CD16 signaling cascade leading to NK cell activation.     

Cyclosporine A alleviates severe anaemia associated with refractory large granular lymphocytic leukaemia and chronic natural killer cell lymphocytosis

Large granular lymphocytic (LGL) leukaemia and chronic natural killer cell lymphocytosis (CNKL) are chronic indolent disorders often associated with neutropenia and constitutional symptoms. Severe anaemia occurs in about 20% of patients and is currently treated with corticosteroids followed by oral cyclophosphamide in non-responders. 30% of patients fail initial measures, and salvage therapy is inadequate. We describe three transfusion-dependent patients (two with T-LGL leukaemia, one with CNKL) refractory to corticosteroids, cyclophosphamide, and in one case fludarabine. Cyclosporine A (CSA) initiation resulted in prompt transfusion-independence and was well tolerated in all patients, making it an attractive alternative therapy for this disorder.     

Comparison of the use of bulk to micro culture of cell preparations for lymphokine-activated cytotoxicity assays.

Different assay systems have been used to quantitate lymphokine-induced natural cytotoxic activity as a measure of immune status. This study compares the effects of inducing cytotoxicity in a bulk culture system, where effector cells are transferred to a micro culture well for assay, to a micro culture system where the effector cells are not transferred. The effector/target ratio for both the bulk and micro culture systems was calculated using the number of viable effector cells present at the time of target cell addition. After overnight incubation with interleukin-2 (IL-2), the lytic activity of murine spleen cells to targets using a micro culture system was increased two-fold over the bulk culture method. This increase was amplified further after 5 days of activation with IL-2, in that the micro culture system resulted in a four-fold increase in cytotoxic activity. The loss of some adherent cells in the bulk culture system did not explain the overall decrease in recovered cytotoxicity. The difference appeared to be related to cell loss during centrifugation. Therefore, the E/T ratios are different in the two systems if not corrected for the number of viable cells.     

乌苯美司体外对人单核细胞功能的活化作用

观察了国产乌苯美司（乌比美克）体外对人单核细胞功能的影响：①０．０１￣１００μｇ／ｍｌ乌苯美司能直接诱导人单核细胞生成ＩＬ－１；②经０．１μｇ／ｍｌ乌苯美司作用４ｈ后，人单核细胞即开始分泌ＩＬ－１，２４ｈ达到高峰，以后逐渐下降；③经１０￣１００μｇ／ｍｌ乌苯美司预处理，单核细胞可促进ＮＫ细胞活性，而经０．０１￣０．１μｇ／ｍｌ乌苯美司处理，单核细胞则抑制ＮＫ细胞活性。     

The Effects of Astragalus membranaceus and Tripterygium hypoglaucum on NK Activity in Systemic Lupus Erythematosus

ＴｈｅＥｆｆｅｃｔｓｏｆＡｓｔｒａｇａｌｕｓｍｅｍｂｒａｎａｃｅｕｓａｎｄＴｒｉｐｔｅｒｙｇｉｕｍｈｙｐｏｇｌａｕｃｕｍｏｎＮＫＡｃｔｉｖｉｔｙｉｎＳｙｓｔｅｍｉｃＬｕｐｕｓＥｒｙｔｈｅｍａｔｏｓｕｓＺｈａｏＸｉａｏ－ｚｈｏｎｇ（赵小忠）（Ｄｅｐａｒ...     

Lymphokine activated killer cells in murine coxsackievirus B3 myocarditis

The purpose of the present study was to determine whether lymphokine activated killer (LAK) cells were involved in the development of coxsackievirus B3 (CB3) myocarditis in both the acute viremic (Experiment I) and the subacute aviremic (Experiment II) stages. To induce LAK cells, recombinant human interleukin-2 (IL-2) was administered to CB3-infected mice subcutaneously daily, starting on day 0 in Experiment I and on day 7 in Experiment II for 7 days, respectively. The treated groups were compared to infected controls. Splenic lymphocytes of IL-2 treated mice were further cultured in vitro in IL-2 containing medium for 7 days, and LAK cell activity, i.e., cytotoxic activity of the lymphocytes against EL-4 tumor cells and against cultured fetal myocytes, was assayed by⁵¹Cr-release method. In Experiment I, histologic scores, myocardial virus titers, and LAK cell activity did not differ significantly between IL-2 treated and untreated groups. In contrast, in Experiment II, there were more cellular infiltration associated with severe necrosis and higher LAK cell activity against EL-4 cells and cultured myocytes in IL-2 treated than in untreated groups. The presence of LAK cells was demonstrated in the subacute stage of murine CB3 myocarditis. Thus, the behavior of LAK cell activity may vary with the course of myocarditis, and enhanced LAK cell activity may be involved in the development of the disease.This work was supported by research grants from the Conference on Coronary Artery Disease, Japanese Education of Science and Walfare (Nos. 08877110 and 09470164), Kanae Shinyaku Foundation, and Japan Cardiovascular Research Foundation.     

肿瘤病人红细胞免疫功能与自然杀伤细胞活性的关系

①目的　探讨恶性肿瘤病人红细胞免疫功能与自然杀伤 (NK)细胞活性的关系。②方法　选择原发性肺癌、卵巢癌及白血病病人各 2 0例为研究对象 ,以 2 0名健康人为正常对照 ,采用免疫黏附法测定红细胞黏附肿瘤细胞的能力 ,采用MTT法测定NK细胞活性。③结果　恶性肿瘤病人红细胞黏附肿瘤细胞的能力和NK细胞活性明显低于正常对照组 (F =9.2 9、12 .85 ,q =5 .15～ 7.86 ,P <0 .0 1) ;正常人红细胞胞质液可以显著提高自身及病人NK细胞活性 (F =4 .32～ 5 .89,q =3.96～ 4 .4 9,P <0 .0 5、0 .0 1)。④结论　红细胞免疫功能与NK细胞活性密切相关。     

体外细胞培养检验“方证对应”关系的方法学研究

目的通过在体外细胞培养中加入中药复方粗提制剂对具有典型的中医证型的肺癌患者的外周血淋巴细胞NK活性的影响，探索能否在体外实验中反映“方证对应”关系。方法以水煎醇沉法制备两种中药复方（益气养阴和健脾化痰）的粗提制剂，取具有典型的中医证型（气阴两虚或脾虚痰湿证型）的肺癌患者的外周血淋巴细胞，将同一个患者的标本分为四个体外实验组，即空白对照组，中药5mg／ml组及10mg／ml组，自细胞介素2（250^u/ml，阳性对照）组。按“辨证论治”原则加入相应的中药复方制剂。体外培养22h或94h，以MTT法检测各组的NK活性。结果无论培养22h或94h，中药组的NK活性与对照组比较皆无显著性差异（P〉0．05），而白细胞介素2（阳性对照）组的NK括性均明显升高（P〈0．05）。结论两种中药复方制剂都未能在体外反应出“方证对应”的效果，说明中药粗提制剂直接用于体外细胞培养，其结果的可靠性差。     

Dynamic observations on NK cell activity and IL-2 and IFNγ productionin vitro in patients on maintenance hemodialysis

Summary In order to study the immune function of patients on maintenance hemodialysis (MHD), we assayed NK cell cytotoxicity against K⁵⁶² targets in 40 patients on MHD, and the production of IL-2 and IFN in peripheral blood mononuclear cells (PBMC) after PHA stimulation, in contrast to those in normal controls. The results showed that NK cell activity and IL-2 and IFN levels were markedly lower in the patients than in the controls. Afeter a single dialysis, NK cell activity as well as IL-2 and IFN levels were elevated to different extent. But there was no significant change in patients after long-term dialysis. There was a positive correlation between the NK cell activity and IL-2 and IFN activity in the controls, but no such correlation was found in the patients on MHD. There was a positive correlation between the NK cell activity and IL-2 activity in patients after dialysis, suggesting that immune function were impaired in the patients on MHD, with a decline in the activity of NK cell and IL-2 and IFN, and a disorder of immune regulation cycle. These abnormal immune impairments in the patients could be partly corrected by hemodialysis. However, long-term hemodialysis is not much helpful in the improvement of patient’s immune function.