Human alloreactive CTL interactions with gliomas and with those having upregulated HLA expression from exogenous IFN-gamma or IFN-gamma gene modification.

By flow cytometry, a panel of 18 primary glioma cell explants exhibited high expression of class I HLA-A, B, C, but class II HLA-DR expression was absent. Freshly isolated normal brain cells displayed little or no HLA antigens. Alloreactive cytotoxic T lymphocytes (aCTL), sensitized to the HLA of the patient, were generated in a one-way mixed lymphocyte response (MLR). The specificity of aCTL was confirmed to be to target cells (patient glioma cells or lymphoblasts) expressing the relevant HLA antigens. However, nontumor patient-specific aCTL did not lyse normal brain cells. Titration of antibodies to HLA class I into cytotoxicity assays blocked lysis of gliomas by aCTL, confirming aCTL T cell receptor (TCR) interactions with the class I antigen on gliomas. Furthermore, aCTL interactions with glioma cells caused their apoptosis. Coincubations of aCTL with gliomas resulted in upregulated cytokine secretion. Importantly, dexamethasone, an immunosuppressive steroid used for brain edema, did not affect aCTL lytic function against tumor, indicating that steroid-dependent patients may benefit from the immunotherapy. We also explored the use of interferon-gamma (IFN-gamma) to increase aCTL tumor recognition. Coincubation of gliomas with exogenous IFN-gamma (500 U/ml, 48 h) caused a 3-fold upregulation of HLA class I and a slight induction of class II antigen expression. Gene-modified glioma cells producing IFN-gamma similarly displayed upregulated HLA expression. Glioma cells incubated with exogenous IFN-gamma or IFN-gamma-transduced glioma cells were more susceptible to lysis by aCTL than their parental counterparts, thus supporting the concept of combining IFN-gamma cytokine gene therapy with adoptive aCTL immunotherapy for brain tumor treatment.     

Cytokine regulation of intercellular adhesion molecule-1 (ICAM-1) expression on human glioblastoma cells

Intercellular adhesion molecule-1 (ICAM-1) has recently been identified as one of the ligands for lymphocyte function-associated antigen-1 (LFA-1). Immunohistochemical staining of frozen tissue sections using the ICAM-1 antibody RR1/1 demonstrated significant levels of ICAM-1 expression on human glioblastoma cells and on intratumoural vascular endothelial cells. ICAM-1 was weakly expressed or absent from low grade gliomas and absent from normal and fetal brain. ICAM-1 expression was similar to that of MHC class II. HLA-DR antigens. Glioblastoma cell lines constitutively expressed ICAM-1 to a minimal or moderate extent. Surface antigen expression of ICAM-1 and ICAM-1-specific mRNA could be significantly increased by incubating glioblastoma cells with interleukin-1 beta (IL-1 beta), tumour necrosis factor-alpha (TNF-alpha), and interferon-gamma (IFN-gamma). IL-2, IL-4, IL-6 and transforming growth factor beta 2 (TGF-beta 2) had no significant effect on surface antigen expression. Significant enhancement of ICAM-1 expression was obtained using TNF-alpha and IL-1 beta at 1-10 U/ml and at 500 U/ml of IFN-gamma. Induction of ICAM-1 specific mRNA was observed 4 h after cytokine treatment and decreased by 24 h. Surface antigen expression of ICAM-1 increased for up to 48 h after treatment.     

Interferon-gamma increases susceptibility of murine pancreatic beta cells to lysis by allogeneic cytotoxic T lymphocytes

The selective loss of insulin-producing pancreatic beta cells which occurs in IDDM has been postulated to result from lysis by beta cell-specific cytotoxic T lymphocytes (CTL). CTL typically recognise antigen in the context of MHC class I molecules, which are normally present at low levels on beta cells. However, hyperexpression of class I antigens on islet cells has been observed in the early stages of beta cell destruction in IDDM. Since interferon-gamma (IFN-gamma) is known to increase class I expression on a number of cell types, we have investigated the responses of murine beta cells to this cytokine under various conditions. Two color immunostaining followed by FACS analysis showed that on average, only 14.9 +/- 3.1% of cultured beta cells were class I positive. However, a majority of beta cells could be induced to express class I after 24 hours of IFN-gamma treatment, and maximal induction (80-90% positive) occurred after 48 hours. Importantly, increased class I expression on beta cells could be achieved with very low concentrations of IFN-gamma (1-10 U/ml). Expression of class II MHC was never detected under any of the conditions employed to up-regulate class I. Interestingly, although islet cells were only moderately susceptible to lysis by allospecific CTL, this susceptibility was markedly enhanced by prior exposure of the islets to IFN-gamma. Taken together, these results suggest that beta cells are extremely susceptible to up-regulation of class I MHC molecules by IFN-gamma, and that this property may render these cells particularly susceptible to lysis by autologous class I-restricted CTL. Since enhanced expression of class I frequently accompanies inflammatory responses and viral infections, this property of beta cells may account in part for their selective destruction in IDDM.     

Brucella abortus regulates bovine macrophage-T-cell interaction by major histocompatibility complex class II and interleukin-1 expression

T-cell activation is dependent on nominal antigen associated with major histocompatibility complex (MHC) class II molecules and interleukin-1 (IL-1), both provided by antigen-presenting cells. We have studied the effects of Brucella abortus and recombinant bovine gamma interferon (IFN-gamma) on bovine macrophage expression of MHC class II and IL-1 molecules and subsequent T-cell proliferation in response to B. abortus. When peripheral blood mononuclear cells were cocultured with B. abortus and IFN-gamma, increasing amounts of IFN-gamma, from 1 to 100 U/ml, down regulated T-cell proliferation. Expression of MHC class II molecules on macrophages was increased independently by IFN-gamma or B. abortus treatment. Thus, class II molecule expression was not the cause of down regulation of T-cell proliferation as observed in other systems. However, B. abortus-IFN-gamma-treated macrophages obtained from overnight cultures had minimal membrane IL-1 compared with macrophages treated with B. abortus alone. Loss of membrane IL-1 required IFN-gamma and the o-polysaccharide of the lipopolysaccharide. IFN-gamma at 1 U/ml in combination with B. abortus produced a 32% decrease in T-cell response, while IFN-gamma at 100 U/ml added to B. abortus-treated cultures produced an 82% reduction in T-cell response. Membrane IL-1 levels were not altered when recombinant bovine IFN-alpha or the rough strain 45/20 of B. abortus, which lacks the o-polysaccharide, was used. Secreted IL-1 levels were unaffected by IFN-gamma and B. abortus treatment. The addition of recombinant bovine IL-1 beta (0.001 to 0.1 ng/ml) to B. abortus- and IFN-gamma-treated cultures failed to provide a signal necessary for T-cell proliferation. These data suggest that membrane IL-1 has a key role in T-cell activation in response to B. abortus. When the o-polysaccharide of B. abortus lipopolysaccharide is combined with IFN-gamma at an inappropriate time during an immune response, T-cell proliferation is prevented and cannot be restored by the addition of exogenous IL-1.     

Monophosphoryl lipid A (MPL) upregulates major histocompatibility complex (MHC) class I expression by increasing interferon-gamma (IFN-gamma)

Tumor immunity is primarily mediated by cells as CD8+ cytotoxic T lymphocytes (CTL) recognize tumor antigen by MHC class I molecules. But most tumors are associated with a decreased expression of MHC class I to escape the antitumor immunity of the host. Our previous data have demonstrated that MPL has an antitumor effect on metastatic lung cancer of B16 melanoma with enhancing cytotoxicity due to increase of IFN-gamma and IL-2, and decrease of IL-4, which indicates the stimulation of type 1 helper T cells (Th1). To determine the effects of MPL, IFN-gamma, TNF-alpha, and IL-1 alpha on MHC class I expression of B16 melanoma cells, we evaluated the expression of MHC class I molecules with treatments of MPL, IFN-gamma, TNF-alpha, and IL-1 alpha by flow cytometry. The supernatant of MPL-treated spleen cells in vitro upregulated the expression of MHC class I molecules of B16 melanoma cells compared to the control supernatant of spleen cells. The MHC class I expression of B16 melanoma cells treated with IFN-gamma, but not TNF-alpha or IL-1 alpha, increased in a time-dependent manner. In conclusion, MPL upregulated MHC class I expression of B16 melanoma cells by activating spleen cells via IFN-gamma. These data suggest that increased IFN-gamma by MPL is responsible for the upregulation of MHC class I expression to augment cytotoxicity. Therefore, we suggest that MPL could play an important role in immunotherapy.     

Cytokine dependent inverse regulation of CD54 (ICAM1) and major histocompatibility complex class I antigens by nuclear factor kappaB in HEp2 tumor cell line: effect on the function of natural killer cells

The aim of this study is to investigate the mechanisms by which elevated nuclear factor kappaB (NFkappaB) activity in HEp2 cells can modulate the function and survival of immune effector cells. Inhibition of NFkappaB functional activity by stable expression of IkappaB super-repressor rendered HEp2 cells (HEp2-IkappaB((S32AS36A))) susceptible to natural killer (NK) cell mediated cytotoxicity. Increase in surface ICAM1 expression was greater on HEp2-IkappaB((S32AS36A)) cells than on the surface of vector alone transfected HEp2 cells when these cells were treated with IFN-gamma. In contrast, tumor necrosis factor alpha (TNF-alpha) treatment augmented ICAM-1 expression on the surface of vector-alone transfected HEp2 cells and not on the HEp2-IkappaB((S32AS36A)) cells. Moreover, synergistic augmentation of ICAM-1 by a combination of TNF-alpha and interferon-gamma (IFN-gamma) treatment was completely abrogated on the surface of HEp2-IkappaB((S32AS36A)) cells. The addition of blocking antibody to ICAM-1 surface antigen partially inhibited the increased cytotxicity mediated by interleukin-2 treated NK cells against HEp2-IkappaB((S32AS36A)) cells. In contrast to ICAM-1, the expression of major histocompatibility complex (MHC) class I antigens were downregulated when the function of nuclear NFkappaB was inhibited in HEp2 cells. The addition of IFN-gamma to HEp2-kappaB((S32AS36A)) cells increased the expression of MHC class I antigen and rendered these cells less susceptible to NK cell mediated cytotoxicity. Secretion of IFN-gamma and granulocyte macrophage-colony-stimulating factor (GM-CSF) by NK cells was also significantly increased in the presence of HEp2-IkappaB((S32AS36A)) cells, and the treatment of these tumor cells with IFN-gamma prior to their addition to the cultures of NK cells decreased the released IFN-gamma and GM-CSF by NK cells. However, the levels of NK cell mediated cytotoxicity and IFN-gamma secretion remained significantly higher in the presence of both untreated and IFN-gamma treated HEp2-IkappaB((S32AS36A)) cells when compared with vector-alone transfected HEp2 cells. Thus, NFkappaB regulates inversely the expression of ICAM-1 and MHC class I antigens on HEp2 tumor cells and this may contribute to the resistance of these cells to NK cell mediated cytotoxicity.     

Role of interferon-gamma in T-cell responses to Semliki Forest virus-infected murine brain cells

Primary brain cell cultures prepared from newborn C3H mice were infected with Semliki Forest virus (SFV) or treated with a beta-propiolactone-inactivated preparation of SFV (BPL-SFV). The effects of recombinant interferon-gamma (IFN-gamma) treatment on SFV replication, SFV antigen display, major histocompatibility complex (MHC) class I and class II antigen expression, susceptibility to lysis by SFV-specific cytotoxic T lymphocytes (CTL) and the ability to stimulate SFV-specific T lymphocytes to release IFN-gamma were determined. The IFN-gamma treatment prevented replication of SFV, as determined by incorporation of [3H]uridine into SFV-RNA, and reduced expression of SFV antigens on the cell surface, as determined by lysis with antibody and complement or indirect immunofluorescence. BPL-SFV-treated brain cells expressed no SFV antigen detectable by lysis with antibody and complement or indirect immunofluorescence. IFN-gamma increased expression of MHC class I and class II antigens, measured by indirect immunofluorescence, susceptibility to killing by alloreactive T-cell lines and ability to stimulate an allogeneic mixed lymphocyte reaction (MLR). Brain cells infected with SFV or treated with BPL-SFV were susceptible to killing by the CTL. The killing was MHC restricted and neither uninfected nor untreated cells were killed. IFN-gamma treatment prior to SFV infection or BPL-SFV treatment resulted in an augmentation of lysis by the CTL, indicating that even where SFV antigen expression is reduced or present at very low levels, in the context of enhanced MHC class I expression cells remain susceptible to CTL killing. Brain cells treated with BPL-SFV stimulated SFV-specific T cells to release IFN-gamma. Pretreatment of brain cells with IFN-alpha beta or IFN-gamma prior to BPL-SFV treatment markedly increased the ability of the cells to stimulate the SFV-specific T cells to release IFN-gamma. Release of IFN-gamma was MHC restricted and brain cells untreated with BPL-SFV did not stimulate IFN-gamma release. IFN-gamma released by T cells stimulated with BPL-SFV-treated brain cells increased class II MHC expression by brain cells as assessed by indirect immunofluorescence.     

IL-12 directly up-regulates the expression of HLA class I, HLA class II and ICAM-1 on human melanoma cells: a mechanism for its antitumor activity?

IL-12 enhances cytolytic activity and proliferation of NK and T cells, and induces cytokines such as IFN-gamma. No direct effects on non-hematopoietic cells have been shown. This study investigates the effects of IL-12 on melanoma cells in vitro. We analyzed 15 melanoma cell cultures and 1 melanoma cell line. Out of 16 samples 13 expressed the beta chain of the IL-12 receptor (IL-12Rbeta). Preincubation with IL-12 increased the surface levels of human leukocyte antigen (HLA) class I, HLA class II and intercellular adhesion molecule (ICAM)-1 of those cultures with IL-12Rbeta expression. The effects of IL-12 on HLA class I could be blocked by an IL-12-neutralizing monoclonal antibody (mAb), but not by an mAb against IFN-gamma. Melanoma cells transduced with IL-12 expressed enhanced levels of HLA class I, HLA class II and ICAM-1 compared to controls. Co-incubation of the melanoma cells with allogeneic peripheral blood mononuclear cells (PBMC) resulted in enhanced proliferation and increased production of IL-2 and IFN-gamma after pretreatment with IL-12. IL-12 pretreatment increased the susceptibility of melanoma cells to lysis by prestimulated autologous PBMC. Since IL-12 induced immunocritical surface molecules on melanoma cells, it might be beneficial during immune interventions in melanoma patients.     

Cellular responses to human chondrocytes: absence of allogeneic responses in the presence of HLA-DR and ICAM-1.

To assess the accessory cell function of human articular chondrocytes, we assessed the ability of human chondrocytes to stimulate allogeneic peripheral blood mononuclear cells (PBMC) and to support phytohaemagglutinin (PHA)-induced proliferation of highly purified T cells. We also examined the surface expression of HLA-DR and ICAM-1 on the chondrocytes both unstimulated and stimulated with cytokines in vitro. Chondrocytes failed to stimulate allogeneic PBMC despite the constitutive expression of MHC class I molecules and the cytokine-induced expression of class II molecules but were able to support T cell proliferation to PHA, IFN-gamma and to a limited extent, IL-1 beta, induced class II expression on chondrocytes. ICAM-1 was present on 94-99% of freshly isolated cells; this declined with culture (17-59%; P < 0.005) but was readily induced by IFN-gamma, IL-1 beta, and tumour necrosis factor-alpha. Alloreactivity and, presumably, autoreactivity to chondrocytes requires factors in addition to the surface expression of DR and ICAM-1. However the presence of these molecules suggests a capacity for cell-cell interactions in inflammatory sites such as the cartilage pannus junction.     

Response of glioma cells to interferon-gamma: increase in class II RNA, protein and mixed lymphocyte reaction-stimulating ability

Previous results by ourselves and others demonstrated that brain cells and cell lines express major histocompatibility complex class II antigens. We examined interferon-gamma (IFN-gamma)-mediated induction of human class II antigen expression on the glioma cells. Purified IFN-gamma induced the expression of HLA-DR antigens on the surface of the glioma cell lines U-373 MG and U-105 MG. Concomitant increase of HLA-DR alpha- and HLA-DC beta-specific RNA in the cytoplasm was also observed after treatment with IFN-gamma. Increases of class II antigen paralleled the increased level of class II-specific RNA. The effect of IFN-gamma on the induction of human class II antigen expression was dose and time dependent. A marked induction of human class II antigen expression was observed when glioma cells were cultured with more than 100 U/ml of IFN-gamma. Little or no induction was observed with less than 50 U/ml of IFN-gamma. Compared to human blood monocytes, glioma cells needed higher concentrations of IFN-gamma for the induction of class II antigen expression. In allogenic mixed lymphocyte cultures, the glioma cell line U-373 MG stimulated a mixed lymphocyte response (MLR). MLR-stimulating capacity was augmented by IFN-gamma. The concomitant augmentation of class II antigen levels and MLR-stimulating capacity suggests that the most relevant factor for MLR stimulation may be antigen density. This is the first report of MLR stimulation by a glioma cell line.     

Induction of intercellular adhesion molecule-1 but not of lymphocyte function-associated antigen-3 in thyroid follicular cells.

We investigated the expression of intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-3 (LFA-3) by thyrocytes and their regulation by cytokines. Immunofluorescence studies on cryostat sections and on freshly dispersed cell preparations showed that ICAM-1 and LFA-3 are barely detectable in non-autoimmune thyrocytes. However, thyrocytes acquired ICAM-1 expression in culture. IFN-gamma, IL-1 beta and TNF-alpha produced a clear enhancement of ICAM-1 expression. When tested in combination, IL-1 beta and TNF-alpha were additive to the IFN-gamma effect. LFA-3 expression was not modulated by these cytokines. In the HT93 thyroid cell line generated by transfection with SV40, ICAM-1 and LFA-3 were both constitutively expressed at high levels. Cytokines modulated ICAM-1 expression similarly, but to a greater extent than in normal thyrocytes. LFA-3 remained unmodified. These results support the notion that normal thyrocytes are immunologically silent cells. The capability of cytokines to induce ICAM-1 together with HLA class I and class II-expression on thyrocytes suggests that under their influence, these cells may express all the surface molecules required for antigen presentation and/or for being recognized as target cells in the context of thyroid autoimmune disease.     

Activation by interferon-gamma of expression of ICAM-1 and MHC class II antigens in tumour cells from colorectal carcinomas

Aims-To determine whether lack of MHC class II antigen and intercellular adhesion molecule-1 (ICAM-1) expression in some tumours is due to the inability of the tumour cells to respond to the cytokine interferon-gamma (IFN-gamma), an important activator of these surface molecules.Methods-Cells from 40 colorectal tumours which did not constitutively express class II MHC antigens or ICAM-1 were kept in short term culture after disaggregation for a few days to two weeks without significant loss of viability. These were treated with IFN-gamma. Expression of class II MHC antigens and ICAM-1 was determined using immunohistological techniques.Results-There was clear induction in vitro of both MHC class II antigens and ICAM-1 in cells from eight of the tumours, with between 50 and 80% of the tumour cells in the cultures staining positively. The staining was apparent within 24 hours, appeared maximal at about three days, and declined thereafter. There were no obvious differences in cell morphology or viability between the cultures which were inducible and those which were not, nor were there obvious differences between the tumours from which they were derived.Conclusions-Expression of MHC class II antigens and ICAM-1 may be induced by IFN-gamma in a small proportion of colorectal tumours which do not constitutively express these antigens, showing that only a minority of tumours are capable of responding to this cytokine.     

Dual role of TNF-alpha in NK/LAK cell-mediated lysis of chronically HIV-infected U1 cells. Concomitant enhancement of HIV expression and sensitization of cell-mediated lysis

The U937-derived chronically HIV-infected U1 cell line and uninfected U937 cell clones were efficiently lysed by both unstimulated (NK) and IL-2-stimulated (lymphokine-activated killer; LAK) peripheral blood mononuclear cells (PBMC) of healthy HIV-seronegative donors. Pretreatment of target cells with IFN-gamma down-modulated killing of both U1 cells and two U937 cell clones, and up-regulated MHC class I expression. In contrast, TNF-alpha enhanced the sensitivity of infected U1 cells, but not of U937 cell clones to NK / LAK cell lysis. Co-cultivation of IL-2-stimulated PBMC with U1 cells triggered expression and replication of HIV by cell-cell contact, and this effect was inhibited by anti-TNF-alpha antibodies (Ab); virus production was partially inhibited by zidovudine. Of interest, anti-TNF-alpha Ab protected U1 cells from LAK cell activity. Thus, TNF-alpha can induce HIV expression from chronically infected U1 cells, but also plays an important role in sensitizing these cells to lysis.     

Stimulatory effect of interleukin-1 beta on the interferon-gamma-dependent HLA-DR production.

Expression of major histocompatibility complex (MHC) class II genes is induced by interferon-gamma (IFN-gamma) in antigen-presenting cells. When human leukaemia THP-1 and PL-21 cells were pretreated with phorbol ester, IFN-gamma-dependent induction of MHC class II gene was markedly enhanced. To elucidate the mechanism of the phenomenon, we examined the effect of cytokines which were secreted from THP-1 cells by the treatment of phorbol ester. Among those cytokines, interleukin-1 beta (IL-1 beta) had a positive effect on IFN-gamma-dependent induction of MHC class II genes. In order to rule out the possibility that different clones of THP-1 cells respond independently to IFN-gamma or phorbol ester, we showed that all independent single clones expressed MHC class II and produced IL-1 beta by IFN-gamma or phorbol ester, respectively.     

Cytokine regulation of HLA on thyroid epithelial cells

The regulation of class I and class II HLA expression in human thyroid follicular cells was studied in vitro. Tumour necrosis factor-alpha (TNF-alpha) enhanced the expression of class I antigen on thyrocytes, but these cytokines had little effect on the expression of class II antigen. Interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6) did not affect class I and class II antigen expression. The combination of interferon-gamma (IFN-gamma) with TNF-alpha or IL-1 beta enhanced the induction of class I and class II antigens, compared with the effect of IFN-gamma alone. Neither class I nor class II expression was induced by IL-6 alone or in combination with IFN-gamma. These findings suggest that TNF-alpha and IL-1 beta may have an important role in inappropriate expression of HLA antigens on thyrocytes in thyroid gland.     

Inhibition of natural killer cell-mediated cytotoxicity by Kaposi's sarcoma-associated herpesvirus K5 protein

Kaposi's sarcoma-associated herpesvirus (KSHV) K3 and K5 proteins dramatically downregulate MHC class I molecules. However, although MHC class I downregulation may protect KSHV-infected cells from cytotoxic T lymphocyte recognition, these cells become potential targets for natural killer (NK) cell-mediated lysis. We now show that K5 also downregulates ICAM-1 and B7-2, which are ligands for NK cell-mediated cytotoxicity receptors. As a consequence, K5 expression drastically inhibits NK cell-mediated cytotoxicity. Conversely, de novo expression of B7-2 and ICAM-1 resensitizes the K5-expressing cells to NK cell-mediated cytotoxicity. This is a novel viral immune evasion strategy where KSHV K5 achieves immune avoidance by downregulation of cellular ligands for NK cell-mediated cytotoxicity receptors.     

B cell chronic lymphocytic leukaemia cells have reduced capacity to upregulate expression of MHC class I in response to interferon-gamma

AIMS: An important consideration in the design of a tumour vaccine is the ability of tumour-specific cytotoxic T lymphocytes (CTL) to recognise unmanipulated tumour cells in vivo. To determine whether B-CLL might use an escape strategy, the current studies compared B-CLL and normal B cell MHC class I expression. METHODS: Flow cytometry, TAP allele PCR and MHC class I PCR were used. RESULTS: While baseline expression of MHC class I did not differ, upregulation of MHC class I expression by B-CLL cells in response to IFN-gamma was reduced. No deletions or mutations of TAP 1 or 2 genes were detected. B-CLL cells upregulated TAP protein expression in response to IFN-gamma. Responsiveness of B-CLL MHC class I mRNA to IFN-gamma was not impaired. CONCLUSIONS: The data suggest that MHC class I molecules might be less stable at the cell surface in B-CLL than normal B cells, as a result of the described release of beta(2)m and beta(2)m-free class I heavy chains from the membrane. This relative MHC class I expression defect of B-CLL cells may reduce their susceptibility to CTL lysis in response to immunotherapeutic approaches.     

Target Cell Lysis by Natural Killer Cells is Influenced by β2-Microglobulin Expression

Natural killer (NK) cells form part of the vertebrate defence against viruses and tumours, but show only limited specificity. The molecule(s) recognized by NK cells on target cells are at present unknown. Major histocompatibility complex (MHC) class I antigen concentration on target cells is inversely correlated with NK cell lysis. Here we show that MHC class I-unassociated beta 2-microglobulin (beta 2-m) expression is involved in NK cell-target cell interaction. Two human MHC class I negative cell lines, Daudi and K562, are differentially susceptible to NK cell lysis. Daudi cells are beta 2-m-negative and resistant to NK lysis, K562 are beta 2-m-positive and highly susceptible to lysis by NK cells. Interferon (IFN) treatment augments beta 2-m expression and NK lysis of K562, but not in Daudi cells. NK cell lysis of K562, but not YAC-1 cells, can be inhibited by monoclonal anti-human beta 2-m antibody. Furthermore, susceptibility of mouse embryo fibroblasts (MEF) to NK lysis can be increased by infection with recombinant vaccinia virus expressing the human beta 2-m gene.