Activation of natural killer cells via the p75 interleukin 2 receptor

The IL-2R is composed of at least two subunits, the p55 (CD25/Tac) and p75 glycoproteins. The p75 IL-2R is expressed preferentially on resting human peripheral blood NK cells, but is not detected on substantial proportions of T and B lymphocytes, monocytes, or granulocytes. Anti-p75 IL-2R mAb substantially inhibits the early events associated with NK cell activation by IL-2, including inhibition of cytotoxic activity and induction of the CD69 early activation antigen. While anti-p55 IL-2R mAb alone failed to substantially inhibit the initial events of IL-2 stimulation, maximal inhibition of IL-2-induced cytotoxicity and proliferation was achieved by combining both anti-p55 IL-2R and anti-p75 IL-2R. Collectively, results from the present studies directly implicate the p75 IL-2R as the structure predominantly responsible for IL-2 activation of NK cells.     

Interleukin 2 induction of pore-forming protein gene expression in human peripheral blood CD8+ T cells

Our studies have analyzed pore-forming protein (PFP) mRNA expression in resting and stimulated human peripheral blood CD3- large granular lymphocytes (LGL), CD3+ T cells, and their CD4+ or CD8+ subsets. Signals that stimulate T cells to develop cytotoxic activity (i.e., IL- 2 or OKT-3 mAb) led to the induction of PFP mRNA in T cells. The data indicated that IL-2 directly increased PFP mRNA in the CD8+ subset of T cells, in the absence of new DNA or protein synthesis. Abrogation of IL- 2-induced PFP mRNA expression and cytotoxic potential of T cells by the anti-p75 IL-2 receptor mAb suggested that low numbers of p75 IL-2 receptors on CD8+ T cells were capable of transducing signals responsible for these IL-2-induced effects. The induction of T cell PFP mRNA via CD3, using OKT-3 mAb, was less rapid but greater than that caused by IL-2; however, a combination of PMA and ionomycin, which bypasses crosslinking of the TCR/CD3 complex, could not mimic this increase in PFP mRNA levels in T cells. The role of second messenger systems in regulating PFP mRNA expression remains to be determined. In contrast, high constitutive PFP mRNA expression was observed in CD3- LGL and these mRNA levels could not be enhanced by stimulation with IL- 2. The cytotoxic potential of peripheral blood T cells and LGL induced in response to IL-2 correlated with IL-2-induced PFP mRNA levels in these cells and was consistent with PFP being one of several important molecules involved in the effector function of cytotoxic lymphocytes.     

T cell receptor-mediated selection of functional rat CD8 T cells from defined immature thymocyte precursors in short-term suspension culture

Recent results have indicated that positive and negative repertoire selection act on the major population of CD4,8 double-positive (DP) thymocytes that express 5-10-fold less T cell receptor (TCR) than mature T cells (i.e., they are TCRlow). Since DP cells obtained ex vivo are heterogeneous with regard to their stage within thymic selection, a homogeneous population of virgin DP cells suitable for selection studies was generated in vitro from their immediate precursors, the CD8 single-positive (SP) immature blast cells. To mimic TCR-mediated selection signals, these virgin DP cells were then cultured for another 2 d in the presence of immobilized anti-TCR monoclonal antibodies with or without interleukin 2 (IL-2). Daily monitoring of recovery and phenotype showed that without TCR stimulation, the cells remained DP and became small, TCRlow cells that were lost with a half-life of 1 d, regardless of the presence of IL-2. TCR stimulation resulted in rapid downregulation of CD4 and CD8, maintenance of a larger cell size, and induction of the CD53 antigen that marks mature and CD4,8 double-negative rat thymocytes. In the absence of IL-2, viability decreased as rapidly as without TCR stimulation. Addition of IL-2 rescued TCR-stimulated virgin DP cells and prevented CD8 downregulation, so that 50-80% of input DP cells were recovered after 2 d as CD4-8+53+ cells. After release from modulation, these in vitro generated CD8 SP cells quantitatively upregulated the TCR to the TCRhigh phenotype and were readily induced to proliferate and exhibit cytotoxic T lymphocyte (CTL) activity in a polyclonal readout. Evidence is presented implicating an IL-2 receptor (IL-2R) not containing the p55 chain (i.e., most likely the p70 intermediate affinity IL-2R) in the TCR plus IL-2-driven in vitro differentiation of virgin DP cells towards the mature CD8 SP phenotype.     

Monoclonal antibody defining a molecule possibly identical to the p75 subunit of interleukin 2 receptor

A mouse hybridoma cell line, TU27, producing an mAb was established. TU27 mAb reacted with various human and Gibbon ape T cell lines bearing the IL-2R p75 (IL-2Rp75), but not with cell lines expressing only Tac antigen, IL-2Rp55, and numbers of its binding sites on cell surfaces were similar to those of high-affinity IL-2R. Radioimmunoprecipitation with TU27 mAb defined a molecule with a molecular mass of 75 kD on the surface of IL-2Rp75 bearing cells. TU27 mAb completely blocked IL-2 binding to IL-2Rp75 and to the high-affinity IL-2R but not to IL-2Rp55 composing the low-affinity IL-2R. The IL-2-dependent growth of a human T cell line, ILT-Mat, was significantly inhibited by TU27 mAb only at low concentrations of IL-2, and combination of TU27 mAb and H-31 mAb specific for IL-2Rp55 completely inhibited the cell growth even at high concentrations of IL-2. These data strongly suggest that TU27 mAb is specific for the human IL-2Rp75.     

Characterization of a novel subset of CD8(+) T cells that expands in patients receiving interleukin-12.

IL-12 has significant antitumor activity in mice that may be mediated by CD8(+) T cells. We show in this report that repeated subcutaneous injections of IL-12 in patients with cancer resulted in the selective expansion of a subset of peripheral blood CD8(+) T cells. This T cell subset expressed high levels of CD18 and upregulated IL-12 receptor expression after IL-12 treatment in vivo. In normal subjects, these CD3(+)CD8(+)CD18(bright) T cells expressed IL-12 and IL-2 receptors and adhesion/costimulatory molecules to a greater degree than other CD8(+) and CD4(+) T cells. They appeared morphologically as large granular lymphocytes, although they did not express NK cell markers such as CD56. In addition, CD8(+)CD18(bright) T cells were almost exclusively T cell receptor (TCR) alphabeta+, and exhibited a TCR Vbeta repertoire that was strikingly oligoclonal, whereas the Vbeta repertoire of CD18(dim) T cells was polyclonal. Although CD8+CD18(bright) T cells demonstrated little functional responsiveness to IL-12 or IL-2 alone in vitro, they responded to the combination of IL-12+IL-2 with strong IFN-gamma production and proliferation and enhanced non-MHC-restricted cytolytic activity. In contrast, CD18(dim) T cells were not activated by IL-12 or IL-2, alone or in combination. These findings demonstrate that CD8+CD18(bright) T cells are a unique population of peripheral blood lymphocytes with features of both memory and effector cells that are capable of TCR-independent activation through combined stimulation with IL-12+IL-2. As this activation results in IFN-gamma production and enhanced cytolytic activity, these T cells may play a role in innate as well as acquired immunity to tumors and infectious pathogens. Additional studies will be necessary to determine whether CD8+CD18(bright) T cells mediate the antitumor effect of IL-12 or IL-2 administered to cancer patients, and if so, whether maximal activation of these T cells with the combination of IL-12+IL-2 in vivo can augment the clinical effectiveness of these cytokines.     

Recombinant interferon-alpha selectively inhibits the production of interleukin-5 by human CD4+ T cells.

The effects of recombinant IFN-alpha on the production of IL-5 by human CD4+ T cells were first analyzed on resting CD4+ T cells purified from normal PBMC and stimulated either with a combination of PMA and anti-CD28 mAb or anti-CD3 mAb cross-linked on B7-1/CD32-transfected mouse fibroblasts. We found that IFN-alpha profoundly inhibited in a dose-dependent manner IL-5 production by resting CD4+ T cells whereas IL-10 was upregulated in both systems. The addition of a neutralizing anti-IL-10 mAb to PMA and anti-CD28 mAb upregulated IL-5 production by resting CD4+ T cells but did not prevent IFN-alpha-induced IL-5 inhibition. We then analyzed the effect of IFN-alpha on the production of cytokines by differentiated type 2 helper (Th2) CD4+CD3- cells isolated from peripheral blood of two patients with the hypereosinophilic syndrome. In both cases, IFN-alpha markedly inhibited IL-5 production while it induced mild upregulation of IL-4 and IL-10. Finally, the inhibitory effect of IFN-alpha on IL-5 production was confirmed on a panel of Th2 and Th0 clones generated in vitro. In 2 out of 6 clones, IL-5 inhibition was associated with upregulation of IL-4 and IL-10. We conclude that IFN-alpha selectively downregulates IL-5 synthesis by human CD4+ T cells.     

Identification of a novel signal transduction surface molecule on human cytotoxic lymphocytes

In this study, we have used a newly generated monoclonal antibody (mAb C1.7) to identify a novel 38-kD signal-transducing surface molecule (p38) expressed by lymphocyte subsets capable of cell-mediated cytotoxicity. Virtually all CD16+/CD56+ natural killer (NK) cells and approximately half of CD8+ (T cell receptor [TCR] alpha/beta+) T cells and TCR-gamma/delta+ T cells express the p38 surface molecule. Stimulation of p38 on NK cells with mAb C1.7 activated cytotoxicity, induced lymphokine production, and initiated polyphosphoinositol turnover and [Ca2+]i increases. Unlike other NK cell surface molecules that activate cytotoxicity, p38 stimulation did not result in the release of the granule enzyme N-carbobenzoxy-L-thiobenzyl ester- esterase even under conditions in which mAb C1.7 induced NK cell- mediated redirected lysis of Fc gamma R+ target cells. Activated (recombinant interleukin 2 [rIL-2], 5 d) CD8+ T cells mediated non- major histocompatibility complex (MHC)-restricted cytotoxicity, and the CD8+/p38+ subset contained the overwhelming majority of this activity. F(ab')2 fragments of mAb C1.7 inhibited non-MHC-restricted cytotoxicity mediated by resting NK cells and rIL-2-cultured T cells but did not affect spontaneous cytotoxicity mediated by activated, cultured NK cells. Taken as a whole, our results suggest that p38 may have a direct role in the recognition, signal transduction, and/or lytic mechanisms of non-MHC-restricted cytotoxicity.     

Induction of interleukin 2 receptor beta chain expression by self- recognition in the thymus

1-2% of adult mouse thymocytes express the T cell receptor alpha/beta (TCR-alpha/beta) together with the interleukin (IL) 2R beta (p70), but not the alpha (p 55) chain. We show that the previously described alpha/beta-TCR +CD4-8- and the partially overlapping Ly6C+ thymocytes are contained within this subset. Most IL-2R beta+ alpha/beta-TCR+ cells have a mature and activated (heat stable antigen [HSA]-, thymic shared antigen 1 [TSA-1]-, CD44high, CD69+) phenotype. Overrepresentation of V beta 8.2 in both CD4-8- and CD4 and/or CD8+ IL- 2R beta+ thymocytes suggests that IL-2R beta expression is induced by a TCR-mediated activation event. In mice transgenic for an H-2Kb-specific TCR, IL-2R beta+ cells were abundant under conditions of mainstream negative selection, i.e., in the presence of Kb, but absent under conditions of mainstream positive selection or in a nonselecting environment. Together, these results show that in addition to clonal deletion, self-recognition by immature thymocytes leads to phenotypic maturation of a small subset of thymocytes expressing IL-2R beta. IL-2- deficient mice contain normal numbers of IL-2R beta+ alpha/beta-TCR+ thymocytes, indicating that like mainstream T cell development, this minor pathway of positive selection does not depend on IL-2. However, in the absence of IL-2, the CD4/CD8 subset composition of IL-2R beta+ thymocytes is skewed towards CD4-8+, mostly at the expense of CD4-8-. A possible relevance of this finding for the development of the immune pathology of IL-2-deficient mice is discussed.     

A novel 80-kD cell surface structure identifies human circulating lymphocytes with natural killer activity

Human lymphocytes with natural killer (NK) activity, including most activated gamma/delta+ T lymphocytes, recognize and lyse tumor target cells without requiring recognition of major histocompatibility complex antigen. However, unlike gamma/delta+ T lymphocytes, NK cells do not express CD3/T cell receptor (TCR) molecules, and the receptors involved in cell-mediated cytotoxicity are unknown. To further delineate circulating NK cells, we developed monoclonal antibodies (mAbs) against the human NK leukemia YT2C2. We report the isolation of a mAb termed BY55, recognizing at the cell surface a novel 80-kD protein with restricted expression. In addition to the immunizing cell line, this mAb binds to circulating NK cells, gamma/delta+ cells, and a minor subset of alpha/beta+ T lymphocytes. Expression of the BY55 mAb- reactive epitope/molecule is regulated by activation, as short-term culture of peripheral blood lymphocytes (PBL) with phorbol ester induced its downmodulation. Furthermore, BY55 mAb reactivity was found neither with the NK nor with the TCR alpha/beta+ gamma/delta+ clones tested. Biochemical studies as well as phenotypic analysis revealed that this structure is different from all previously identified molecules on the lymphocyte cell surface. Interestingly, we found that BY55+ cells exert most NK activity obtained with fresh circulating lymphocytes. We report that within fresh E rosette-positive PBL only a subset of the CD16+, CD56+, and CD57+ cells coexpressed BY55 molecule, indicating that BY55 mAb defines a unique subset mediating NK activity of circulating PBL.     

The CD28 ligand B7/BB1 provides costimulatory signal for alloactivation of CD4+ T cells.

Activation via the T lymphocyte cell surface molecule CD28 provides a potent amplification signal for interleukin 2 (IL-2) production in several in vitro systems. The B lymphocyte activation antigen, B7/BB1, is a natural ligand for CD28. Here we investigate the role of CD28 and B7/BB1 in primary activation of CD4+ T lymphocytes stimulated with allogeneic B lymphoblastoid cell lines. A subset of peripheral CD4+ T cells that is unresponsive to crosslinking of CD3/T cell receptor (TCR) with CD3 monoclonal antibody (mAb) does proliferate in response to allogeneic B lymphoblasts. TCR binding to allogeneic major histocompatibility complex antigens was an absolute requirement for activation of these cells because mAbs to either CD3 or human histocompatibility leukocyte antigen (HLA) class II completely inhibited activation. CD28 and B7/BB1 antibodies inhibited T cell proliferation 90% and 84%, respectively. Similar results were obtained with the total CD4+ T lymphocyte population. Crosslinking of HLA-DR antigens on small, resting B cells induced rapid expression of B7/BB1, which peaked at 6 h and returned to baseline levels within 18 h. These data demonstrate that CD28-B7/BB1 binding provides an important early second signal for alloactivation of CD4+ T lymphocyte by B lymphoblasts. The results also suggest that T cells interacting with allogeneic resting B cells may induce B7/BB1 expression in the alloantigen-presenting cell as a consequence of interaction between the TCR and class II molecules.     

IL-6-dependent spontaneous proliferation is required for the induction of colitogenic IL-17-producing CD8+ T cells

We propose a novel role for interleukin (IL) 6 in inducing rapid spontaneous proliferation (SP) of naive CD8(+) T cells, which is a crucial step in the differentiation of colitogenic CD8(+) T cells. Homeostasis of T cells is regulated by two distinct modes of cell proliferation: major histocompatibility complex/antigen-driven rapid SP and IL-7/IL-15-dependent slow homeostatic proliferation. Using our novel model of CD8(+) T cell-dependent colitis, we found that SP of naive CD8(+) T cells is essential for inducing pathogenic cytokine-producing effector T cells. The rapid SP was predominantly induced in mesenteric lymph nodes (LNs) but not in peripheral LNs under the influence of intestinal flora and IL-6. Indeed, this SP was markedly inhibited by treatment with anti-IL-6 receptor monoclonal antibody (IL-6R mAb) or antibiotic-induced flora depletion, but not by anti-IL-7R mAb and/or in IL-15-deficient conditions. Concomitantly with the inhibition of SP, anti-IL-6R mAb significantly inhibited the induction of CD8(+) T cell-dependent autoimmune colitis. Notably, the transfer of naive CD8(+) T cells derived from IL-17(-/-) mice did not induce autoimmune colitis. Thus, we conclude that IL-6 signaling is crucial for SP under lymphopenic conditions, which subsequently caused severe IL-17-producing CD8(+) T cell-mediated autoimmune colitis. We suggest that anti-IL-6R mAb may become a promising strategy for the therapy of colitis.     

TCR stimulation with modified anti-CD3 mAb expands CD8+ T cell population and induces CD8+CD25+ Tregs

Modified anti-CD3 mAbs are emerging as a possible means of inducing immunologic tolerance in settings including transplantation and autoimmunity such as in type 1 diabetes. In a trial of a modified anti-CD3 mAb [hOKT3gamma1(Ala-Ala)] in patients with type 1 diabetes, we identified clinical responders by an increase in the number of peripheral blood CD8+ cells following treatment with the mAb. Here we show that the anti-CD3 mAb caused activation of CD8+ T cells that was similar in vitro and in vivo and induced regulatory CD8+CD25+ T cells. These cells inhibited the responses of CD4+ cells to the mAb itself and to antigen. The regulatory CD8+CD25+ cells were CTLA4 and Foxp3 and required contact for inhibition. Foxp3 was also induced on CD8+ T cells in patients during mAb treatment, which suggests a potential mechanism of the anti-CD3 mAb immune modulatory effects involving induction of a subset of regulatory CD8+ T cells.     

T cell receptor ligation induces interleukin (IL) 2R beta chain expression in rat CD4,8 double positive thymocytes, initiating an IL-2- dependent differentiation pathway of CD8 alpha+/beta- T cells

The role of interleukin (IL)2 in intrathymic T cell development is highly controversial, and nothing is known about IL-2R expression on thymocytes of the T cell receptor (TCR) alpha/beta lineage undergoing TCR-driven differentiation events. We analyze here IL-2R alpha and beta mRNA expression in an in vitro system where newly generated rat CD4,8 double positive (DP) thymocytes respond to TCR ligation plus IL-2 (but not to either stimulus alone) with rapid differentiation to functional CD8 single positive T cells (Hunig, T., and R. Mitnacht. 1991. J. Exp. Med. 173:561). TCR ligation induced expression of IL-2R beta (but not alpha) chain mRNA in DP thymocytes. Addition of IL-2 then lead to functional maturation and expression of the IL-2R alpha chain. To investigate if the CD8 T cells generated via this IL-2R beta-driven pathway in vitro correspond to the bulk of CD8 T cells seeding peripheral lymphoid organs in vivo, we compared their phenotype to that of lymph node CD8 T cells. Surprisingly, analysis of CD8 cell surface expression using a novel anti-CD8 monoclonal antibody specific for the alpha/beta heterodimeric isoform, and of CD8 alpha and beta chain mRNA revealed that T cells generated by TCR ligation plus IL-2 resemble thymus-independent rather than thymus-derived CD8 cells in that they express CD8 alpha without beta chains. These findings demonstrate that TCR crosslinking induces functional IL-2R on immature DP rat thymocytes. In addition, they show that at least in vitro, CD8 alpha/alpha T cells are generated from TCR-stimulated DP thymocytes (which express the CD8 alpha/beta in the heterodimeric isoform) along an IL-2-driven pathway of T cell differentiation.     

Large granular lymphocytic leukemia]

We investigated the surface markers, cell-function, clonality, and the association of IL-2 receptors and a second messenger of src family of tyrosine kinase p56lck in IL-2 signal transduction of the leukemic cells of 12 patients with large granular lymphocytic leukemia (LGL leukemia). The leukemic cells of 5 patients were CD3+ and 5 of them were CD3-. In three patients with CD3- leukemia examined, one showed karyotype abnormality of 46, XY, -10, +mar and the delta gene of TCR was rearranged in one patient. The TCR of the leukemic cells of a patient MH with CD3+, CD4 and CD8 (double positive marker: DP) recognised rabbit IgG presented by macrophages. The recognition was class II restricted. We examined the expression pattern of CD8 subunits and found that DP leukemic cells commonly expressed CD8 alpha+ beta-. These results suggested that DP leukemic cells were CD4+ T cells and expressed CD8 alpha secondarily. The p75 IL-2 receptors were detected, however, the modulation of p56lck in the process of IL-2 signal transduction were not found out. There was no association between p75 and p56lck when leukemic LGL cells proliferated on stimulation with IL-2.     

Functional properties of a unique subset of cytotoxic CD3+ T lymphocytes that express Fc receptors for IgG (CD16/Leu-11 antigen)

A subset of peripheral blood T lymphocytes coexpressing CD3 and IgG Fc receptors (FcR) (CD16/Leu-11 antigen) have been identified, isolated, and functionally characterized. The CD3+, CD16+ cells were established in short-term culture using growth medium containing interleukin 2 (IL- 2). Both the freshly isolated cells and the cultured cell line stably expressed the CD3+, CD16+ phenotype. Furthermore, a majority of these T cells lacked either CD4 or CD8 expression. Like in vitro-activated cytotoxic T lymphocytes and natural killer (NK) cells, the CD3+, CD16+ cells showed numerous azurophilic granules. Although these cells failed to mediate significant levels of NK cell-mediated cytotoxicity even after stimulation with IL-2, they efficiently functioned as effectors of antibody-dependent cellular cytotoxicity (ADCC). The Ig isotype specificity of the ADCC was analyzed using an isotype switch-variant family of a murine anti-HLA monoclonal antibody (mAb). Similar to the CD3-, CD16+ NK cell population, the CD3+, CD16+ T cells preferentially used the IgG2a antibody to mediate ADCC. The CD3+, CD16+ cells demonstrated a proliferative response when cocultured with either a NK- sensitive tumor cell line, K562, or a NK-insensitive B lymphoblastoid cell line, CCRF-SB. The response against CCRF-SB was significantly inhibited by anti-IL-2 receptor antibody, whereas the response against K562 was only partially diminished. Cytotoxicity was also induced in the CD3+, CD16+ population by the presence of anti-CD3 mAb, indicating that cytotoxicity can be triggered by stimulation via the CD3-T cell antigen receptor complex. By isolating these CD3+, CD16+ cells from the peripheral blood of a normal, healthy individual, it has been possible to extensively study the morphology, antigenic phenotype, and functional behavior of this unique subset of T lymphocytes expressing IgG FcR.     

Identification of novel lymphoid tissues in murine intestinal mucosa where clusters of c-kit+ IL-7R+ Thy1+ lympho-hemopoietic progenitors develop

We have revealed that about one and a half thousand tiny clusters, filled with one thousand closely packed lymphocytes, can be found throughout the murine small and large intestinal mucosa. They are located in crypt lamina propria (cryptopatches; CP) and can be first detected at 14-17 d after birth. A large fraction of lymphocytes in CP expresses c-kit, IL-7R, Thy1 and a lymphocyte function-associated antigen, LFA-1, whereas most of them remain CD3-, TCR alpha beta-, TCR gamma delta-, sIgM-, and B220-. The population size of IL-2R alpha+, HSA+ and Pgp-1+ subsets is variable (20-50%) and the composition of CD8+, Ly-1+, and CD4+ subsets is smaller but also variable (3-20%). In the small intestine, CP do not contain cells undergoing apoptosis nor cells bearing RAG-1 molecules, but do contain dendritic stromal cells bearing CD11c/CD18 molecules. The frequency of DNA replicating cells in CP is higher than that in Peyer's patches (PP), is lower than that in the thymic cortex and is almost comparable with that in the thymic medulla. The numbers of CP remain the same in aged mice (> 114 wk) but double after estrogen treatment even though the thymi are attenuated sharply in both conditions. Thus, with respect to histogenesis, lymphocyte composition and tissue level of cellular behavior, neither PP, isolated lymphoid follicles, peripheral LNs, nor thymus are identical with CP. Finally, CP are virtually absent in lamina propria of IL-7R-deficient mice that display a profound reduction in thymic and peripheral lymphoid cellularity. By contrast, CP are present in germ- free mice and in athymic (nu/nu), SCID, TCR beta x delta-/-, RAG-2-/-, PP-deficient (aly/aly), stem cell factor (Sl/Sld) and c-kit (W/Wv) mutant mice. Taking all of these results together, CP are the first identification of gut-associated murine lymphoid tissues where the generation of IL-7-dependent lympho-hematopoietic progenitors for T and/or B cell descendants may start to take place at the age of commencement of weaning.     

IL-17和IFN-γ在红内期间日疟原虫感染者外周血T细胞亚群中的表达

目的：观察分析间日疟原虫感染患者外周血T细胞亚群中分泌IL-17和IFN-γ的细胞的表达水平。方法采用流式细胞术检测分析患者组（PV）和健康对照组（HD）外周血CD3＋、CD3＋CD8＋、CD3＋CD8-和γδT细胞中分泌胞内因子IL-17、IFN-γ的细胞的百分比。结果在所有检测的细胞亚群中,分泌IFN-γ的CD3＋CD8-和γδT细胞在其所属亚群中所占比例明显高于健康对照组,分泌IL-17的γδT细胞的比例明显高于健康对照组。结论本研究对于理解机体抗疟原虫的细胞免疫的功能状态具有一定意义。     

Autologous tumor-specific cytotoxic T lymphocytes in the infiltrate of human metastatic melanomas. Activation by interleukin 2 and autologous tumor cells, and involvement of the T cell receptor

TIL from metastatic melanoma proliferated by greater than 1,000-fold (840-3,675, mean 1,543) after 6 wk in culture of mixtures of TIL and tumor cells with rIL-2 alone. Cytolysis was restricted to autologous tumor cells. CD8+ T cells were the predominant population of TIL before and after expansion, and were primarily responsible for autologous tumor-specific CTL activity. No other rIL-2-activated lymphocytes from peripheral blood, lymph nodes with melanoma metastasis, or TIL from sarcoma or renal cell carcinoma had autologous tumor-specific CTL activity. There were few or no CD16+ NK cells in TIL from metastatic melanoma before or after incubation with rIL-2, respectively. However, TIL from sarcoma or renal cell carcinoma contained a substantial proportion of CD3-CD16+ NK cells, which increased in number in culture with rIL-2. Purified CD16+ NK cells as well as CD3+CD16- T cells from rIL-2-activated TIL of renal cell carcinoma displayed MHC-nonrestricted cytotoxicity. At the clonal level as determined by limiting dilution, 8 of 10 clones from melanoma TIL displayed cytotoxicity restricted to autologous tumor cells, while all 13 clones from renal cancer TIL equally lysed autologous and allogeneic tumor cells. Anti-T cell receptor (TCR)-alpha/beta(WT31) mAb as well as anti-CD3 mAb inhibited autologous melanoma cell-specific CTL activity mediated by rIL-2-activated TIL at the effector phase. These two mAbs also inhibited rIL-2-dependent proliferation of these TIL when added to the culture. Pretreatment of fresh melanoma cells with mAb to MHC antigens followed by washing inhibited specific CTL activity. These results suggest that both TCR-alpha/beta on effector TIL and MHC antigens on fresh tumor cells are involved in the specific immune-recognition. After reaching maximum propagation, TIL from metastatic melanoma responded poorly to rIL-2 alone. However, stimulation with fresh autologous melanoma cells restored both CTL activity and proliferation in response to rIL-2. The latter is associated with IL-2 receptor (Tac antigen) expression on the surface. These results indicate that TIL from metastatic melanomas may have unique characteristics different from lymphocytes obtained from the other sources, and may contain precursor CTL sensitized in vivo to autologous tumor cells, and thus can be propagated in larger numbers with rIL-2 alone while retaining autologous tumor-specific CTL activity.     

Functional consequences of interleukin 2 receptor expression on resting human lymphocytes. Identification of a novel natural killer cell subset with high affinity receptors

In this study, we have used radiolabeled IL-2 binding assays, Northern blot analysis, immunofluorescent flow cytometry and cell sorting, as well as proliferation and cytotoxicity assays to perform an extensive phenotypic and functional characterization of the IL-2 receptor in normal resting human peripheral blood lymphocytes. Our results indicate that almost all T cells (greater than 98%) express neither the high affinity IL-2 receptor nor the functional intermediate affinity p75 chain of the IL-2 receptor without prior activation. In contrast, most NK cells constitutively express the isolated intermediate affinity p75 IL-2 receptor. In addition, a subpopulation of NK cells, distinguished by high density expression of the NKH1 antigen, constitutively express the high affinity IL-2 receptor, in addition to an excess of the isolated intermediate affinity p75 IL-2 receptor. These NKH1bright+ cells exhibit a brisk proliferative response to IL-2, similar to that seen with antigen-activated T cells, yet do so in the absence of any known antigenic stimuli. No other resting peripheral blood lymphocyte population, including CD4+, CD8+, and CD20 cells, exhibits this property. The intermediate affinity p75 IL-2 receptor, as it exists in its isolated form on resting NK cells, does not transduce a growth signal equivalent to that seen in NK cells expressing the high affinity IL-2 receptor, despite doses of IL-2 that are known to fully saturate the isolated p75 chain. This strongly suggests that additional structural or functional components are involved in generating the proliferative response following the binding of IL-2 to the high affinity heterodimeric form of the IL-2 receptor. The constitutive expression of this functional high affinity IL-2 receptor on a small population of resting NK cells provides further evidence in support of a role for these cells in the host's early defense against viral infection or malignant transformation, before the more delayed but specific T cell response.