Antibody-dependent Cell-mediated Cytotoxicity Reaction Research Articles

Antibody-induced NKG2D blockade in a rat model of intraportal islet transplantation leads to a deleterious reaction.

Intraportal islet transplantation is plagued by an acute destruction of transplanted islets. Amongst the first responders, NK cells and macrophages harbour an activating receptor, NKG2D, recognizing ligands expressed by stressed cells. We aimed to determine whether islet NKG2D ligand expression increases with culture time, and to analyse the impact of antibody-induced NKG2D blockade in islet transplantation. NKG2D-ligand expression was analysed in rat and human islets. Syngeneic marginal mass intraportal islet transplantations were performed in rats: control group, recipients transplanted with NKG2D-recombinant-treated islets (recombinant group), and recipients treated with a mouse anti-rat anti-NKG2D antibody and transplanted with recombinant-treated islets (antibody-recombinant group). Islets demonstrated increased gene expression of NKG2D ligands with culture time. Blockade of NKG2D on NK cells decreased in vitro cytotoxicity against islets. Recipients from the control and recombinant groups showed similar metabolic results; conversely, treatment with the antibody resulted in lower diabetes reversal. The antibody depleted circulating and liver NK cells in recipients, who displayed increased macrophage infiltration of recipient origin around the transplanted islets. In vitro blockade of NKG2D ligands had no impact on early graft function. Systemic treatment of recipients with an anti-NKG2D antibody was deleterious to the islet graft, possibly through an antibody-dependent cell-mediated cytotoxicity reaction.

Abstract 1182: New bioluminescent strategies to measure kinetics of immune cell-mediated cytotoxicity

Abstract The number of immunotherapeutic strategies for treating cancer have expanded in recent years, increasing the need for assays that can be used to assess their effectiveness. To facilitate studies of these immunotherapies, we developed bioluminescent methods that could rapidly and sensitively measure target cell cytotoxicity. Two approaches were developed. The first method utilizes a bioluminescent enzymatic activity assay for a well-accepted cytotoxicity marker, lactate dehydrogenase (LDH), which is rapidly released from dead cells. With the improved sensitivity of the bioluminescent assay, lower levels of LDH can be measured, resulting in the detection of fewer dead cells (LOD< 10 cells). Cytotoxicity in both 2D and 3D cell cultures can be monitored over time by removing samples of medium at different time points. Here we present examples in which this assay was used to test the effectiveness of immunotherapeutic treatments including antibody-drug conjugates (ADC) and antibody-dependent cell-mediated cytotoxicity (ADCC). In ADCC models, cell killing was detectable at early time points (by 2 hours). Increases in the cytotoxicity of individual samples could be followed over time, from early time points to 24 hours. In the second approach we developed a novel bioluminescent assay based on the specific labeling of target cells. With this method, intracellular proteins in live target cells are covalently labeled. Upon target cell death, the labeled proteins are released from the cell and the label is quantitated using bioluminescence detection reagents. Using this assay, we were able to quantitate the killing of target cells in both ADCC and CAR-T applications. By adding the detection reagents directly to the medium at the start of the ADCC reaction, cell death was monitored in real-time for up to 6 hours. Neither assay requires genetic engineering and both involve no or minimal target cell preparation. They are applicable to multiple cell types and the non-lytic nature of these approaches provides kinetic capabilities and opportunities for multiplexing. The ability to rapidly quantitate cytotoxicity will facilitate the functional analysis of immunotherapies. Citation Format: Maggie L. Bach, Natasha Karassina, Peter Hofsteen, Donna Leippe, James Cali, Jolanta Vidugiriene. New bioluminescent strategies to measure kinetics of immune cell-mediated cytotoxicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1182.

N-Acetyl- d-glucosamine-coated polyamidoamine dendrimer promotes tumor-specific B cell responses via natural killer cell activation

N-Acetyl- d-glucosamine-coated polyamidoamine dendrimer (GN8P), exerting high binding affinity to rodent recombinant NKR-P1A and NKR-P1C activating proteins, was shown previously to delay the development of rat colorectal carcinoma as well as mouse B16F10 melanoma, and to potentiate antigen-specific antibody formation in healthy C57BL/6 mice via NK cell stimulation. In this study, we investigated whether GN8P also modulates tumor-specific B cell responses. Serum anti-B16F10 melanoma IgG levels, IgG2a mRNA expression, antibody dependent cell-mediated cytotoxicity (ADCC), and counts of plasma as well as antigen presenting B cells were evaluated in tumor-bearing C57BL/6 mice treated with GN8P and in respective controls. To reveal the mechanism of GN8P effects, the synthesis of interferon-gamma (IFN-γ) and interleukin-4 (IL-4), cytokines involved in regulation of immunoglobulin class switch, was determined. The GN8P treatment significantly elevated IgG, and particularly IgG2a, response against B16F10 melanoma, which led to augmented ADCC reaction. The significant increase in production of IFN-γ, which is known to support IgG2a secretion, was observed solely in NK1.1 expressing cell populations, predominantly in NK cells. Moreover, GN8P raised the number of plasma cells, and promoted antigen presenting capacity of I-A/I-E-positive B lymphocytes by up-regulation of their CD80 and CD86 co-stimulatory molecule expression. These results indicate that GN8P-induced enhancement of tumor-specific antibody formation is triggered by NK cell activation, and contributes to complexity of anticancer immune response involving lectin–saccharide interaction.

Serum containing ovine IgG2 antibody specific for maedi visna virus envelope glycoprotein mediates antibody dependent cellular cytotoxicity

Antibody-dependent cell-mediated cytotoxicity (ADCC) specific for maedi visna virus (MVV) has never been described. The IgG antibody response to MVV is restricted to an IgG1 response whilst MVV specific IgG2 is never seen in persistently infected sheep. To determine whether the isotypic restriction of the antibody response is responsible for the lack of ADCC, an ADCC assay was developed using polyclonal serum raised to recombinant MVV ENV protein. Sheep immunised with a recombinant GST:SUenv fusion protein in complete Freund's adjuvant produced an antibody response which contained IgG1 and IgG2 antibodies. The activity of this serum in an ADCC assay was compared to serum from persistently infected sheep. Serum from immunised sheep mediated ADCC reactions whilst no activity was ever seen in persistently infected sheep serum. IgG2 may therefore be the possible effector isotype for ADCC reactions against MVV. Failure of the IgG2 dependent ADCC system in vivo may contribute to the persistence of MVV-infected macrophages in vivo.

Heterogeneity of multiorgan metastases of human lung cancer cells genetically engineered to produce cytokines and reversal using chimeric monoclonal antibodies in natural killer cell-depleted severe combined immunodeficient mice.

Lung cancer is a major cause of cancer deaths, most of which can be attributed to distant multiorgan metastases. To examine the cellular and molecular mechanisms of lung cancer metastasis to distant organs, we have established novel models of human lung cancer (small cell and non-small cell lung cancer) metastasis in natural killer cell-depleted severe combined immunodeficient (SCID) mice. We investigated whether local production of the cytokines responsible for regulation of macrophage function at tumor growth sites affects the pattern of lung cancer metastasis in distant organs. Several lung cancer cell lines were genetically engineered to produce human macrophage colony-stimulating factor (M-CSF) and monocyte chemoattractant protein-1 (MCP-1), and their metastatic potentials were assessed. Interestingly, M-CSF gene transduction had an antimetastatic effect for the liver and lymph nodes, but not the kidneys. In contrast, MCP-1 gene-modified lung cancer cells and their parent cells had identical metastatic potentials. These findings indicate a possible role for cytokines and suggest that lung cancer has metastatic heterogeneity. Examining ways of controlling human lung cancer metastases, we investigated the antimetastatic effect of chimeric monoclonal antibodies (MAbs) against P-glycoprotein and ganglioside GM2 (MH162 and KM966, respectively). Both MAbs, when given on days 2 and 7, inhibited the development of distant metastases of lung cancer in a dose-dependent fashion. Combined use of anti-P-glycoprotein MAb with M-CSF or MCP-1 gene transduction caused complete inhibition of metastasis of H69/VP cells. The antimetastatic effect of these MAbs in vivo was mainly due to an antibody-dependent cell-mediated cytotoxicity reaction mediated by mouse macrophages. These findings suggest that the mouse-human chimeric MAb in combination with cytokine gene transduction may be useful for the eradication of lung cancer metastases in humans.

Evidence that antisulfatide autoantibodies from rats experimentally infected with Trypanosoma cruzi bind to homologous neural tissue.

Earlier studies in Trypanosoma cruzi-infected rats revealed an increased antibody activity against sulfatide, a specific constituent of both myelin sheaths of peripheral nerves and T. cruzi epimastigotes. To investigate further the characteristics of such anti-sulfatide antibodies, we analyzed their IgG isotypes as well as their ability to bind to homologous neural host structures. Antisulfatide IgG-enriched fractions were obtained from rats acutely infected with T. cruzi. Immunoglobulin isotypes were determined by an enzyme-linked immunosorbent assay (ELISA) method to show that IgG2a and, more significantly, IgG2b were the predominant isotypes of antisulfatide autoantibodies. Further immunofluorescence studies carried out in coronal sections of the rat forebrain revealed, in turn, that antisulfatide antibodies were capable of reacting with homologous neural tissues. Specific binding of these rat autoantibodies to sulfocerebroside on cell surfaces in vivo may in theory play some detrimental role, given the reported ability of rat IgG2b to fix complement or to mediate antibody-dependent cell-mediated cytotoxicity reactions.

Interleukin 1 and tumour necrosis factor alpha may be responsible for the lytic mechanism during anti-tumour antibody-dependent cell-mediated cytotoxicity.

Antibodies are thought to bring about tumour cell lysis by antibody-dependent cell-mediated cytotoxicity (ADCC), but the exact mechanism is not well elucidated. Monocytes are known to be important mediators of anti-tumour ADCC and are also known to secrete the cytokines tumour necrosis factor alpha (TNF-alpha) and interleukin 1 beta (IL-1 beta), both of which have been shown to bring about tumour cell lysis. We examined the release of these cytokines during ADCC and attempted to elucidate which components of the ADCC reaction were necessary for cytokine production. We measured TNF-alpha and IL-1 beta in supernatants collected from a standard ADCC assay using each of the anti-colorectal antibodies m17-1A, c17-1A and cSF25. We found that there was significant TNF-alpha and IL-1 beta release during ADCC mediated by each of these three antibodies and that the magnitude of cytokine release seemed to reflect the degree of tumour cell lysis produced by each antibody. Furthermore, we found that effector cells, target cells and a specific anti-tumour antibody were necessary for this to occur. The presence of only some of the components of the reaction or of an irrelevant antibody produced little or no TNF-alpha or IL-1 beta. We conclude that TNF-alpha and IL-1 beta are released when an effector and tumour target cell are united by a specific tumour antibody and that these cytokines may be important in bringing about tumour cell lysis during the ADCC reaction.

The Epstein-Barr Virus-Encoded Glycoprotein gp 110 (BALF 4) Can Serve as a Target for Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC)

Antibody-dependent cell-mediated cytotoxicity (ADCC) is thought to play a major role in controlling the spread of the Epstein-Barr virus (EBV) in an infected individual. Recently, the viral membrane protein gp 350/220, which is also expressed at the surface of the virus producing cell, was identified as a target for ADCC reactions. Due to its glycoprotein nature, the EBV protein gp 110 is another possible ADCC target. It is one of the most abundant proteins found during the late phase of viral replication; until now, however, researchers have not been able to localize it on the surface of EBV positive cells. By means of recombinant vaccinia viruses containing the genes for gp 350/220 and gp 110, respectively, we expressed these proteins in lymphoblastoid cells, which were then used as targets in ADCC studies with sara from EBV-positive and -negative individuals. In these experiments we were able to demonstrate the feasibility of our approach for the investigation of EBV-specific ADCC reactions and could confirm the role of gp 350/220 as an ADCC target. Furthermore, we were able to show that gp 110 can also be recognized in an ADCC reaction, proving that at least some gp 110 molecules must be expressed at the cell surface.

Antibody-dependent cell-mediated cytotoxicity (ADCC) in Parkinson's disease

The possible role of a non-specific cell-mediated immune reaction in the pathogenesis of Parkinson's disease (PD) is discussed. It was found that the killer cell activity of PD patients below 60 years of age was significantly lower than in the older age groups or in the age-matched control group. On the other hand, it was also found that the killer cell activity of PD patients with severe symptoms (Hoehn-Yahr's IV, V stage) was significantly higher than that of the milder cases. These results support the hypothesis that an ADCC reaction — mediated by the killer cells — may play a role in the pathogenesis of PD.

Suppression of macrophage‐mediated antibody‐dependent killing of Schistosoma mansoni larvae by concanavalin A stimulated spleen cell‐derived factor(s)

Culture supernatants (sup) from rat spleen cells stimulated with concanavalin A (Con A) sepharose 4B suppressed schistosomulum killing by macrophages in a system of antibody-dependent cell-mediated cytotoxicity (ADCC). (a) The suppressive effect of the sup was strongest when they were added to cultures at the start of the ADCC assay, and it was decreased when sup were added to cultures in which the ADCC assay had begun more than 3 h previously. (b) From the results of various experiments in ADCC reaction, it was suggested that the suppressive factor(s) inhibited at least in part the interaction between macrophages and antibodies which had been bound to schistosomula, and that the suppressive factor(s) was bound to the macrophage side. (c) The suppressive activity was partially inactivated by treatment at 56 degrees C for 30 min, and it was lost almost completely by the treatment at 100 degrees C for 10 min. (d) The suppressive factor(s) differed from known cytokines such as interleukin 1 (IL-1), interleukin 2 (IL-2), granulocyte-macrophage-colony stimulating factor (GM-CSF), interferon-gamma (IFN-gamma), and tumour necrosis factor (TNF).

Isolated Adrenocorticotropin Deficiency Associated with an Autoantibody to a Corticotroph Antigen That Is not Adrenocorticotropin or Other Proopiomelanocortin-Derived Peptides*

A 44-yr-old man with hypocortisolism was shown to have an undetectable basal plasma ACTH level and absent or subnormal ACTH and beta-lipotropin responses to provocative testing with insulin, vasopressin, and CRH. Endocrine function after glucocorticoid replacement was otherwise normal, thus establishing the diagnosis of isolated ACTH deficiency. This patient's serum was tested immunohistochemically for the presence of an antipituitary antibody by indirect immunofluorescence of rat pituitary tissue. Positive immunostaining was observed in stellate-shaped cells in the anterior and intermediate lobes. Immunopositive cells were shown by immunoelectron microscopy to have ultrastructural characteristics of corticotrophs. Immunoreactivity was concentrated in secretory granules 120-170 nm in diameter. In a double immunolabeling procedure, staining by the patient's serum was shown to colocalize with rabbit antiserum to ACTH, but not with antisera to PRL, GH, beta TSH, or beta LH. Immunoabsorption of the patient's serum with ACTH-(1-24), ACTH-(1-39), gamma MSH, corticotropin-like intermediate lobe peptide, beta-endorphin, or beta-lipotropin failed to diminish immunolabeling in the pituitary. We conclude that the antipituitary antibody in this patient's serum shows immunohistochemical specificity for a rat corticotroph antigen located in secretory granules that is neither ACTH nor any of the proopiomelanocortin (POMC)-derived peptides tested. The autoantigen could be a cell-specific granular factor involved in the posttranslational processing of POMC or secretion of ACTH. We postulate that an autoimmune process may account for this patient's disease, and that his antipituitary antibody could play a pathogenic role by either inhibiting a POMC-processing enzyme or initiating an antibody-dependent cell-mediated cytotoxicity reaction, resulting in the selective destruction of corticotrophs.

Effects of recombinant interleukin 2 on immunological effector cells of the peripheral blood in patients with HBe antigen-positive chronic hepatitis.

Changes in peripheral blood immunological effector cells after systemic administration of recombinant interleukin 2 (rIL2) were studied in 14 patients with HBe antigen-positive chronic hepatitis. The patients were intravenously injected with rIL2 for 4 weeks, and the experiments were performed using mononuclear cells isolated from the peripheral blood during and after rIL2 administration. No significant changes in OKT4-positive cell population, OKT8-positive cell population, OKT4/OKT8 ratio and Leu7-positive cell population were detected during and after intravenous injection of rIL2. However, the natural killer (NK) cell and lymphokine-activated killer (LAK) cell activities significantly increased in the fourth week of rIL2 administration, while the K cell activity in the antibody-dependent cell-mediated cytotoxicity reaction, antibody response, lectin-induced lymphocyte transformation and IL1 production of LPS-stimulated monocytes were not affected. These results suggest that IL2 increases the NK cell and LAK cell activities in vivo.

Protection of liver cells against experimental damage by extract of cultured Lentinus edodes mycelia (LEM).

Liver cell damage can be induced when isolated liver cells coated with specific antibodies against the liver cell membrane are cultured with peripheral blood mononuclear cells. Although this antibody-dependent cell-mediated cytotoxicity (ADCC) is dependent on the close contact of effector cells with target cells via specific antibodies, a cytotoxic factor or factors causing the inhibition of protein synthesis in liver cells has been detected in the culture supernatant from the ADCC reaction. Similarly, peritoneal exudate macrophages activated by endotoxin lipopolysaccharide also exert cytotoxic effects on isolated liver cells by production of a cytotoxic substance or substances. The liver cell damage caused by either the ADCC or activated macrophage culture supernatants were significantly reduced by pretreating the isolated liver cells with the extract of cultured Lentinus edodes mycelia (LEM). These results suggest that LEM may protect liver cells from immunological damage.

Cell-mediated cytotoxicity-supporting activity of various human gammaglobulin preparations.

Antibody activity, especially that involved in the reaction of antibody-dependent cell-mediated cytotoxicity (ADCC), of five commercially available human gammaglobulin preparations (standard, pepsin-treated, plasmin-treated, polyethylene glycol-fractionated and S-sulfonated gammaglobulin) was measured. All these gammaglobulin preparations had high titers of hemagglutination inhibition and neutralizing antibody against measles virus. In ADCC reaction, the pepsin-treated gammaglobulin preparation showed no antibody activity. The standard gammaglobulin preparation showed weak activity only when highly diluted. The remaining three preparations showed high activity. Though the S-sulfonated gammaglobulin preparation showed no activity in ADCC reaction, it showed high activity after reconversion by means of oxidation and reduction in vitro. The plasmin-treated gammaglobulin preparation showed greater activity than the polyethylene glycol-fractionated preparation of the optimal concentration. In ADCC tests using the plasmin-treated gammaglobulin preparation, K cell activity was strongly inhibited by Hg (thimerosal), while, in those using the standard gammaglobulin preparation, the activity was hardly influenced by Hg, suggesting that the low ADCC activity of the standard gammaglobulin preparation of high concentrations was due to the inhibitory effect of aggregated immunoglobulin G molecules.

Identification of immunosuppressive factors in normal pregnant serum affecting antibody dependent cell-mediated cytotoxicity.

AbstractThe antibody dependent cell‐mediated cytotoxicity (ADCC) activity of peripheral blood lymphocytes was found to be lower in women experiencing normal pregnancy than in nonpregnant females. This decrease was especially striking during the early stage of pregnancy. It was determined that this decrease is caused by two suppressive factors contained in the pregnant serum. One was a high‐molecular‐weight substance with strong suppressive activity, which was not reduced even when the effector cells were washed. This factor showed binding to IgG Fc‐receptors on the lymphocyte surface and to Protein A. Its ability to suppress ADCC activity was eliminated when precipitation was performed with 4% polyethylene glycol. On the basis of these characteristics, it was surmised that this suppressive factor is an IgG‐immune complexlike substance. The suppressive mechanism of this substance is speculated to be competitive binding to the IgG Fc‐receptors of effector cells, which thereby inhibits the binding of specific antibody to these cells. The second suppressive factor was of low molecular weight, and its activity was lost when the effector cells were washed. This factor did not bind to IgG Fc‐receptors, and it is speculated that the action mechanism of this substance must involve direct action against the ADCC reaction system, thereby inhibiting the cellular metabolism of the effector cells.Moreover, it was demonstrated that in normal pregnancy there is a positive correlation between the circulating immune complex concentration in the serum and the serum's suppressive effect on the ADCC activity. This finding suggests that the immune complex‐like substance is the principal factor in the suppression of the ADCC activity.

Development of specific antibodies mediating antibody-dependent cell-mediated cytotoxicity following experimental infection of cattle with bovine herpesvirus 1 and subsequent viral reactivation

Evolution of sensitizing antibodies involved in antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-complement cell lysis and complement-facilitated ADCC was followed in bulls after primary infection by a wild strain of infectious bovine rhinotracheitis virus (bovine herpesvirus 1; BHV-1) and after experimental reactivation of the virus. These antibodies were detected between the 4th and the 7th day after primary infection, reached a maximum level after 2 weeks and rose slightly after reactivation of the virus following dexamethasone treatment. The presence of endogenous complement slightly enhanced the ADCC reaction.

Protection of liver cells from experimentally induced liver cell injuries by tritoqualine.

Liver cell damages were inducible, when isolated liver cells coated with specific antibody against the liver cell membrane were cultured with peripheral blood mononuclear cells. Although this antibody-dependent cell-mediated cytotoxicity (ADCC) was induced by the closed contact of effector cells to targets via the specific antibody, a cytotoxic factor or factors causing the inhibition of protein synthesis in the liver cells was detected in the culture supernatant of the ADCC reaction. Similarly, peritoneal exudate macrophages activated by endotoxin lipopolysaccharide also exerted cytotoxic effects on the isolated liver cells by producing a cytotoxic substance or substances. These liver cell injuries induced by either the ADCC or activated macrophage culture supernatant were significantly reduced, when the isolated liver cells were pretreated with tritoqualine before adding the cytotoxic culture supernatants. These results suggest that tritoqualine may protect liver cells from immunological injuries.

Protection of liver cells from experimentally induced liver cell injury by glycyrrhizin.

Liver cell damage is induced when isolated liver cells coated with specific antibody against the liver cell membrane are cultured with peripheral blood mononuclear cells. Although this antibody-dependent cell-mediated cytotoxicity (ADCC) was induced by closed contact of effector cells with targets via specific antibody, a cytotoxic factor or factors causing inhibition of protein synthesis in liver cells was detected in the culture supernatant of the ADCC reaction. Similarly, peritoneal exudate macrophages activated by endotoxin lipopolysaccharide (LPS) also had cytotoxic effects on isolated liver cells by producing a cytotoxic substance or substances. These liver cell injuries caused by either ADCC or activated macrophage culture supernatants were significantly reduced by pretreatment of the isolated liver cells with glycyrrhizin before the addition of the cytotoxic culture supernatants. These results suggest that glycyrrhizin may protect liver cells from immunological injuries.

Role of IgE in fascioliasis

A prominent phenomenon of helminth infections is their stimulation of a substantial IgE response, both serum total IgE and parasite specific IgE, and there is now considerable evidence that the latter is involved in protective immunity to helminths: Parasite specific IgE has been shown as an essential factor in antibody dependent cell-mediated cytotoxicity reactions (ADCC) of various trematodes and nematodes in vitro. In Fasciola hepatica infections however, there is only limited information about the mechanisms of ADCC and the possible role of specific IgE in the immunity to this parasite. Rat eosinophils have been shown to adhere and to degranulate onto newly excysted F. hepatica in vitro in the presence of immune serum, while no bound complement was detectable on the worms. Also the killing capacity of eosinophil major basic protein was demonstrated but the particular involvement of reaginic antibodies has not yet been cleared. Time course studies in experimentally F. hepatica infected calves showed that the highest levels of homocytotropic antibodies were reached between 20 and 28 weeks of infection which coincides with the period of the parasite expulsion from the bile ducts of the host. This finding, together with the massive accumulation of mast cells at the bile ducts and the liberation of histamine from blood-cells of infected cattle when contacted with fluke antigen, further suggests that IgE might play a role in the defence mechanism of the host against F. hepatica infections. Using radioimmunoassay techniques in the rat model the increase of F. hepatica ES-specifc IgE is coincident with the increase of total serum IgE and the response shows a biphasic pattern. The first peak reflects the migratory phase of young flukes through the liver parenchyma, whereas the second increase appears when adult flukes establish in the bile ducts. However, the significance of these reactions is not yet understood and further investigations might possibly show wether circulating IgE can perform some regulatory mechanisms in immunity to liver fluke infections. In addition, it has been found that F. hepatica infections in rats — similarly to other helminth infections — were able to potentiate an unrelated IgE response. They also induced a prolonged elevation of serum total IgE, which declined after anthelmintic treatment. Other experiments suggested that the serum total IgE in F. hepatica infections is parasite-dose dependent. However, these observations are still not cleared and need to be further investigated.

Antibody-dependent cell-mediated cytotoxicity against HLA-A, HLA-B and HLA-DR specificities.

HLA specificity of alloantibodies involved in antibody-dependent cell-mediated cytotoxicity (ADCC) was investigated using PHA-induced lymphoblasts ( PLB ) from peripheral blood and Epstein-Barr virus-induced human lymphoblastoid cell lines (LCL). The ADCC activity with PLB sensitized by HLA-A or HLA-B antibodies was relatively low, but higher than that of control. DR antibodies completely failed to sensitize PLB for ADCC. LCL sensitized by corresponding DR antibodies gave strong ADCC reactions, independent from their D phenotypes. The ADCC reactions against LCL were abolished by absorption of the DR antibodies from the alloantisera. Thus the ADCC with LCL would provide a procedure to study the role of DR antibodies in chronic rejection of long-term renal allografts.