FcR Gamma Research Articles

TCR isoform containing the Fc receptor gamma chain exhibits structural and functional differences from isoform containing CD3 zeta.

The structure and function of the TCR-CD3 complex containing a homodimer of the gamma chain of the high affinity receptor for IgE (FcR gamma) (FcR gamma+ TCR) was investigated by transfecting the FcR gamma gene into a CD3 zeta-, CD3 eta-, FcR gamma- T cell line. Introduction of FcR gamma, as well as CD3 zeta, induced a high expression of the TCR-CD3 complex on the cell surface. Transfected FcR gamma formed a homodimer and associated firmly with the TCR alpha beta dimer but only weakly with the CD3 gamma delta epsilon. Stimulation of both FcR gamma and CD3 zeta transfectants by antibodies against TCR or CD3 induced accumulation of inositol phosphates, the Ca2+ response, IL-2 production, and growth inhibition. On the other hand, antigen stimulation of transfectants expressing FcR gamma as well as CD3 zeta induced IL-2 production, but only the latter exhibited the antigen-induced growth inhibition. In vitro kinase assay suggested that the CD3 zeta dimer but not the FcR gamma dimer associates with the Fyn kinase. These results indicate that the FcR gamma homodimer is able to form a functional TCR complex but that the mode of assembly and the signaling function of FcR gamma+ TCR, including its association with tyrosine kinase(s), may differ from the TCR-CD3 complex containing CD3 zeta homodimers (zeta+ TCR). This provides an example which illustrates that different TCR isoforms mediate distinct signals and functions.

Cellular cytotoxicity mediated by isotype-switch variants of a monoclonal antibody to human neuroblastoma

The biological property of an antibody is determined by its antigen binding characteristics and its isotype-related effector functions. We have established monoclonal antibodies of different isotypes by stepwise selection and cloning of the hybridoma CE7. The original CE7 secretes an IgG1/kappa (CE7 gamma 1) antibody that recognises a 185 kD cell surface glycoprotein expressed on all human sympatho-adrenomedullary cells. Isotype-switch variants were isolated in the following sequence: from the original CE7 gamma 1, CE7 gamma 2b variants were isolated, and from a CE7 gamma 2b variant CE7 gamma 2a variants were isolated. The antibodies of three different isotype variant cell lines possess identical antigen binding characteristics, but display distinct effector functions as demonstrated by antibody dependent cell-mediated cytotoxicity (ADCC). ADCC was performed with the neuroblastoma line IMR-32 as the target cells, and different FcR gamma positive cells were either freshly isolated from human peripheral blood leukocytes or cultured for 6-10 days and tested as potential effector cells. Tumour lysis mediated by monocyte-derived macrophages depended on the presence of CE7 gamma 2a antibodies; antibodies from the CE7 hybridomas of gamma 2b and gamma 1 isotypes were virtually inactive in ADCC assay. Pre-exposure of macrophages to rIFN-gamma enhanced their ADCC activity, a result that is compatible with the notion that the high affinity Fc IgG receptor (FcR gamma I/CD64) is involved in the triggering of ADCC in macrophages. In contrast to macrophages, mononuclear cells, nonadherent cells and monocytes displayed considerable non-specific lytic activity, which was little influenced by the presence of antibody regardless of the isotype added.

Differences in signal transduction between Fc gamma receptors (Fc gamma RII, Fc gamma RIII) and FMLP receptors in neutrophils. Effects of colchicine on pertussis toxin sensitivity and diacylglycerol formation.

Studies on the role of microtubule integrity in stimulus-response coupling in neutrophils have generated contradictory data. To determine the role of microtubule integrity in stimulus-response coupling elicited by two different mechanisms, i.e., engagement of the Fc receptors (FcR gamma II, FcR gamma III) or engagement of the receptor for FMLP, we utilized colchicine (10 microM), which reduces pericentriolar microtubules to 29% of control, and compared its effect with that of nocodazole (50 microM) and lumicolchicine (10 microM). We now demonstrate that treatment of neutrophils with colchicine but not lumicolchicine, inhibits degranulation elicited by engagement of Fc receptors but augments degranulation in response to FMLP. In contrast to the ligand-specific effect of microtubule-disruption on degranulation, superoxide anion production (assembly of the NADPH oxidase) is unaffected by colchicine regardless of the ligand. To determine whether intact microtubules were required for responses elicited by ligation of Fc gamma RII(CD32) or Fc gamma RIII(CD16), mAb directed against these receptors were employed. Treatment of neutrophils with mAb KuFc79 directed against Fc gamma RII(CD32) or mAb 3G8 directed against Fc gamma RIII(CD16) inhibited degranulation of neutrophils elicited by immune complexes (IC). In contrast, removal of most of Fc gamma RIII by phosphatidylinositol-specific phospholipase C did not significantly alter degranulation in response to IC. We conclude that degranulation elicited by IC results from ligation of both Fc gamma RII and phosphatidylinositol-specific phospholipase C-insensitive Fc gamma RIII. The importance of microtubule integrity on the generation of intracellular signals was also examined. Degranulation of neutrophils proceeds via pertussis toxin-sensitive and insensitive pathways; treatment of cells with colchicine did not augment the action of pertussis toxin. Stimulation of neutrophils by chemoattractants results in a biphasic increase in 1,2-sn-diacylglycerol; a rapid increase ("triggering") secondary to the action of a phosphatidylinositol-specific phospholipase C, and a late increase ("activation") secondary to the action of a phosphatidylcholine-specific phospholipase C. Treatment of cells with colchicine altered the production of both [3H]-arachidonic acid-diacylglycerol and diacyl[14C]glycerol in parallel to its effect on degranulation. These studies indicate that the requirement of intact microtubules for degranulation is ligand-specific. Furthermore, assembly of the respiratory burst oxidase does not require intact microtubules. Microtubules most likely alter the cycling of specific receptors or the generation of specific intracellular signals required for stimulus-response coupling in the course of degranulation. Intact microtubules are not uniformly required for the discharge of granule contents during exocytosis.

Lymphokine‐activated killer cell cytotoxicity against human colon carcinomas enhanced by monoclonal antibody D612

The antibody-dependent cellular cytotoxicity (ADCC) properties of a murine monoclonal antibody (MAb), designated D612 (IgG2a), which reacts with human colon carcinomas, was studied using normal human peripheral blood lymphocytes (PBMNC). Although the level of ADCC of PBMNC with D612 varied among different donors, it was 20 to 30 times higher than the lytic activity of control cultures containing isotype-matched control MAb. Incubation of PBMNC with recombinant interleukin-2 (IL-2) resulted in a 2- to 5-fold augmentation in the cytotoxicity of effector cells exposed to MAb. This augmentation was apparent after subtracting nonspecific cellular cytotoxicity from the total cytotoxicity mediated by activated effector cells in the presence of D612. Optimal stimulation of specific ADCC with IL-2 appeared after 24 hr of culture in 500 U/ml of IL-2, resulting in a 3.8 +/- 1.7 fold increase in lytic units. However, stimulation of ADCC was also evident at 10 U/ml of IL-2. Furthermore, antibody dose titrations with untreated and IL-2 activated effectors showed that the threshold dose of MAb needed for efficient ADCC was reduced by 200-fold with IL-2. Depletion of FcR gamma III-positive lymphoid cells markedly reduced D612 ADCC, demonstrating the participation of NK/LAK cells in D612-mediated ADCC. Low levels of ADCC activity were found associated with adherent cells, either untreated or following their activation with gamma-interferon, while D612 was most active with non-adherent effectors. The specificity and ADCC properties of the D612 MAb suggest that it should be considered as a candidate for immunotherapy of colon cancer, particularly when used in combination with IL-2 plus LAK cell treatment.

Three B-cell-surface Molecules Associating with Membrane Immunoglobulin

Three cell-surface molecules associated with surface Ig on murine B cells have been identified. Surface-iodinated resting B cells were lysed in digitonin or Nonidet P-40. The lysates were then examined by immunocoprecipitation with anti-Ig reagents and SDS-PAGE. By this procedure, a 73,000 MW heterodimer, composed of 35,000 and 38,000 MW disulphide-linked subunits, was found to be associated with both mIgM and mIgD. This association was detectable in the detergent digitonin, but not Nonidet P-40. As an alternative approach, resting B cell mIg was capped and internalized by anti-Ig induced mIg cross-linking. The resulting 'modulated' cells were then screened for surface reactivity with a panel of monoclonal antibodies. Most antibodies tested (anti-class I, -class II, -LFA, -FcR gamma, -FcR epsilon) reacted equally well with control and modulated B cells. With two, however, there was a significant reduction in surface representation on modulated B cells. One determinant corresponds to a previously undescribed 90,000 MW murine surface marker, and the other was a polydisperse glycoprotein recognized by the mAb J11D.

IFN-gamma plus glucocorticoids stimulate the expression of a newly identified human mononuclear phagocyte-specific antigen.

Glucocorticoid hormones, although able to exert profound immunosuppressive effects, do not suppress mononuclear phagocyte activation by IFN-gamma and may even enhance it. For example, expression and functional activity of the high affinity FcR for IgG on human mononuclear phagocytes (FcR gamma I) is increased by IFN-gamma and is maximal after co-treatment with IFN-gamma plus the glucocorticoid dexamethasone (DEX). To determine whether there are other mononuclear phagocyte surface Ag that are regulated in this manner, hybridomas were prepared using IFN-gamma-plus-DEX-treated human monocytes as immunogen. Five IgG1 mAb (Mac 2-8, 2-38, 2-48, 2-49, and 2-158) were developed that recognize a trypsin-sensitive mononuclear phagocyte-specific surface Ag of Mr 155,000. There was no detectable reactivity of these mAb to lymphocytes or granulocytes or to several cell lines, including U-937 and HL-60. The p155 Ag was detected on monocytes and increased significantly with time of culture or after treatment with DEX. Expression was maximal after co-treatment with rIFN-gamma plus DEX, but was inhibited or unaffected by treatment with IFN-gamma alone. For freshly isolated cells, expression of the p155 Ag was highest on peritoneal macrophages. Our results indicate that the p155 Ag is a newly identified Ag of the human mononuclear phagocyte lineage and may represent, in the least, a phenotypic marker of monocyte differentiation or maturation.

Generation of continuous large granular lymphocyte lines by interleukin 2 from the spleen cells of mice infected with Moloney leukemia virus. Involvement of interleukin 3.

Continuous cell lines could be reproducibly established by culturing spleen cells from adult mice injected with MLV-producer cells or directly infected with Mo-MLV with rIL-2, whereas the culture of normal splenic cells with rIL-2 induced only transient and limited proliferation resulting in no such lines. All of the lines showed morphological characteristics as LGL with Thy-1+,Lyt-1-,L3T4-,Lyt-2-,AsGM1+,FcR gamma+ phenotype without exception, and most of them exhibited typical NK-patterned cytotoxicity. Analysis of reverse transcriptase activity of the culture supernatants as well as Southern hybridization of the DNA from the lines using an Mo-MLV-specific cDNA probe indicated no evidence of retroviral replication or proviral integration, suggesting that the generation of cell lines reflected a reactive process and viral infection was not directly responsible. It was subsequently revealed that Thy-1+,Lyt-1+,Lyt-2- spleen cells from mice infected with Mo-MLV in vivo spontaneously produced surprising amounts of IL-3 in vitro, leading to the possibility that IL-3 was responsible for the generation of lines. The possibility was directly supported by the observation that continuous lines with identical characteristics could be generated completely in vitro by sequential stimulation with rIL-3 and rIL-2 from normal spleen cells without any involvement of Mo-MLV. The C beta gene of TCR was shown to be rearranged in all the lines examined, indicating the LGL lines were all genetically committed to T cell lineage. Unlike the situation in normal splenic populations expanded by rIL-2, where the expression of IL-2-R was progressively lost, constitutive expression of high-affinity-IL-2-R was observed in all the lines and thus, this was considered to explain the unlimited proliferation of them in response to rIL-2 alone. These results suggested the probable role of IL-3 in the regulation of growth and differentiation of a set of LGL committed to T cell lineage. The possible implications of the phenomenon in the regulation of hematopoiesis as well as in the control of Mo-MLV-induced leukemogenesis were discussed.

Generation of non MHC restricted killing in non-cytotoxic T-lymphocytes from leukaemia patients.

Experiments were undertaken to determine whether the depression of natural killer (NK) activity previously observed in the peripheral blood lymphocytes (PBL) of leukaemia patients in remission extended to NK-like precursors in the blood. T-lymphocytes (E+) from leukaemia patients and normal subjects were depleted of IgG Fc-receptor-bearing (gamma FcR) fresh NK cells by passage over immune complex-coated monolayers. gamma FcR - E + PBL were cultured alone or with DAUDI cells. On day 5 of culture, cytotoxicity toward the NK-sensitive cell lines K562 and MOLT-4 was evaluated in the responder lymphocytes of leukaemia patients and controls. Negligible NK-like cytotoxicity was found in both FcR - E + PBL responder populations cultured alone. By contrast, stimulation with DAUDI induced high levels of K562 and MOLT-4 cytotoxicity in leukaemia as well as in normal responder cells. Complement-mediated cytotoxicity experiments using various McAb demonstrated that in both normal and leukaemia cultures NK-like effectors react with the pan-T OKT3 McAb and with the OKT11 McAB directed to the SRBC receptor, but not with Leu 1 1b and OKM1 McAbs, directed against antigens expressed on peripheral blood NK cells. Fractionation of the responder cells on discontinuous Percoll gradients showed that most of this activity was present in the highly dividing blast cell fraction.

Murine lymphokine-activated killer (LAK) cells: phenotypic characterization of the precursor and effector cells.

Abstract Murine and human lymphocytes incubated in recombinant interleukin 2 (RIL 2) generate a population of cytotoxic cells (lymphokine-activated killer cells [LAK]), which are able to lyse a wide array of fresh tumor cells but do not lyse fresh normal cells. Intravenous administration of these cells with the concomitant administration of RIL 2 can eliminate established pulmonary and hepatic metastases in mice. To characterize the cell that has in vitro LAK activity, we subdivided murine lymphocytes by lysing select subpopulations with the use of complement and antibodies against lymphocyte surface markers or by fluorescence-activated cell sorting. Thy-1.2-negative splenocytes were found to generate near normal amounts of LAK activity after RIL 2 incubation. Small and inconsistent LAK cell activity was generated from Thy-1.2-positive splenocytes. Ia-positive and surface immunoglobulin-positive splenocytes had little or no LAK precursor capability and did not appear to be necessary for LAK activation. Treatment of splenocytes with anti-asialo GM1 (anti-ASGM1) heterosera and complement markedly decreased their ability to generate LAK activity. At the effector stage, cytotoxic cells were of the Thy-1.2-positive, Ia-negative phenotype. Ia-depleted cells were separated into subpopulations bearing or not bearing the gamma Fc receptor (gamma FcR). The majority of cytotoxicity resided in gamma FcR-positive cells. Thus the precursors of murine LAK cells are "null" lymphocytes bearing neither T nor B cell surface markers but develop the Thy-1.2 cell surface marker in vitro, in association with the development of lytic activity for fresh tumor cells after stimulation by RIL 2.

Prognostically significant classification of immune changes in AIDS with Kaposi's sarcoma

Sixteen immunological parameters were assessed quantitatively for their value in providing an immunologically-based and prognostically significant classification of the immune alteration in 97 patients with AIDS and Kaposi's sarcoma (AIDS-KS). The dimensions of reductions in the T4 (T helper-inducer cells) subpopulation of lymphoid cells in the T4-T8 ratio were found to correlate most closely with prognosis. Most other immunological changes did not relate to clinical course. T4 lymphocyte levels greater than 300/microL and a T4-T8 ratio greater than 0.5 indicated a relatively good prognosis, eg, 85% to 95% survival at 12 months. T4 levels less than 100/microL and/or a T4-T8 ratio less than 0.2 had a very poor prognosis, eg, less than 25% survival at 12 months. Intermediate T4 levels and T4-T8 ratios had intermediate prognosis. These immunological findings were found to have independent prognostic value for survival when compared with disease classifications based on tumor stage (I through IV) or on clinical status A (without) or B (with fever, night sweats, or weight loss). Reduced proliferative capacity, increased OKT10 antigen expression, elevated levels of serum IgA, and immune complexes also correlated with prognosis. Elevated levels of serum IgG, cellular HLA-DR expression, and skin test anergy occurred frequently in AIDS-KS but did not have prognostic significance. Variations in level of total lymphocyte, T8 (T suppressor/cytotoxic) cell, gamma FcR receptor-positive cell number, NK activity, or level of serum IgM were less common in AIDS-KS and did not correlate with prognosis.

Prognostically Significant Classification of Immune Changes in AIDS With Kaposi's Sarcoma

Sixteen immunological parameters were assessed quantitatively for their value in providing an immunologically-based and prognostically significant classification of the immune alteration in 97 patients with AIDS and Kaposi's sarcoma (AIDS-KS). The dimensions of reductions in the T4 (T helper-inducer cells) subpopulation of lymphoid cells in the T4-T8 ratio were found to correlate most closely with prognosis. Most other immunological changes did not relate to clinical course. T4 lymphocyte levels >300/μL and a T4-T8 ratio >0.5 indicated a relatively good prognosis, eg, 85% to 95% survival at 12 months. T4 levels <100/μL and/or a T4-T8 ratio <0.2 had a very poor prognosis, eg, <25% survival at 12 months. Intermediate T4 levels and T4-T8 ratios had intermediate prognosis. These immunological findings were found to have independent prognostic value for survival when compared with disease classifications based on tumor stage (I through IV) or on clinical status A (without) or B (with fever, night sweats, or weight loss). Reduced proliferative capacity, increased OKT10 antigen expression, elevated levels of serum IgA, and immune complexes also correlated with prognosis. Elevated levels of serum IgG, cellular HLA-DR expression, and skin test anergy occurred frequently in AIDS-KS but did not have prognostic significance. Variations in level of total lymphocyte, T8 (T suppressor/cytotoxic) cell, gamma FcR receptor-positive cell number, NK activity, or level of serum IgM were less common in AIDS-KS and did not correlate with prognosis.

The IgE antibody system is coordinately regulated by FcR epsilon-positive lymphoid cells and IgE-selective soluble factors.

Recent studies in mice have demonstrated that exposure of lymphocytes to appropriate levels of IgE initiates a cascade of cellular and molecular interactions which function as a network to control IgE synthesis. A key manifestation of these events is the expression of Fc receptors for IgE (FcR epsilon) on both B and T lymphocytes, and the fact that such expression of FcR epsilon can be selectively modulated by the isotype-specific regulatory mediators, suppressive factor of allergy (SFA) and enhancing factor of allergy (EFA). In humans, we have previously shown that the in vitro induction by pokeweed mitogen (PWM) of IgE biosynthesis by peripheral blood lymphocytes (PBL) can also be selectively suppressed by SFA. Herein we show that PWM also induces expression of FcR epsilon+ and FcR gamma+ cells among human PBL by day 2 or 3 in culture, and this early development of FcR epsilon+ lymphocytes appears to be a coordinate event with the ultimate de novo synthesis of IgE in this system. Moreover, as previously documented for IgE synthesis, the presence of SFA causes a 50% reduction of FcR epsilon+ cells induced by PWM. This inhibition is selective, since FcR+ cells for IgG are not affected by exposure to human SFA derived from a recently constructed human T cell hybridoma line which constitutively secretes large quantities of biologically active human SFA. These findings further support the regulatory role that FcR epsilon+ lymphocytes must play in the development of IgE responses by human cells in vitro, and suggest a mechanism by which SFA can selectively inhibit IgE synthesis in PWM-stimulated cultures of human PBL.

Role for mouse macrophage IgG Fc receptor as ligand-dependent ion channel.

The interaction of ligands with the mouse macrophage Fc receptor which binds IgG2b and IgG1 immune complexes (FcR gamma 2b/gamma 1) triggers phagocytosis and secretion of various mediators of inflammation. FcR gamma 2b/gamma 1 has been purified using a monoclonal anti-FcR antibody, 2.4G2, and seems to be an integral membrane glycoprotein of molecular weight (Mr) 47,000-60,000 (ref. 6). Monoclonal antibody 2.4G2 is suitable as a tool for functional studies of FcR because it binds to a functional site of the receptor and induces cellular responses that are normally associated with the occupied receptor. We reported previously that binding of ligands to the macrophage FcR resulted in Na+/K+ ion fluxes through the plasma membrane, and that similar ion fluxes were observed in proteoliposomes containing reconstituted FcR. We have now incorporated FcR into planar lipid bilayers and report here that FcR gamma 2b/gamma 1 forms ligand-dependent cation-selective ion channels, with a conductance of 60 pS in 1 M KCl and an average open channel lifetime of 250 ms. The conductance decays to baseline levels within a few minutes. These results suggest a receptor-ionophore model for the signalling of phagocytosis and inflammatory responses.

Recombinant immune interferon increases immunoglobulin G Fc receptors on cultured human mononuclear phagocytes.

Although recent studies suggest that interferons can increase the number of IgG Fc receptor (FcR gamma) sites on mouse macrophages, direct assessment of similar effects on human mononuclear phagocytes is lacking. We therefore measured the specific binding of 125I- and fluorescein-labeled IgG1 to human monocytes and leukemic cell lines after culture in vitro with highly purified human interferons. We report that natural and recombinant human gamma-interferon causes a dramatic (nearly 10-fold) increase in the number of FcR gamma on normal human monocytes and on the human cell lines HL-60 and U-937. Alpha and beta-interferons cause a modest but significant increase in these receptors. This report demonstrates that gamma-interferon acts directly on human mononuclear phagocytes to increase FcR gamma sites, it identifies a qualitative difference in the physiologic actions of human type I and type II interferons, and it suggests that HL-60 and U-937 cells will be important models for further study of the molecular mechanisms of interferon action. The results reported here could also be the basis for a bioassay to assess the pharmacokinetics and variability of gamma-interferon action on monocytes of individual patients during treatment in vitro and in vivo.

Alterations in the phenotype of hairy cells during culture in the presence of PHA: requirement for T cells

Culture studies of peripheral blood mononuclear cells from 7 entirely typical cases of hairy cell leukemia showed that after culture in the presence of PHA for 2--5 days, the predominant cell type changed from E- SIg+ CIg+ gamma FcR+ muFcR+ hairy cells to an E+ SIg- CIg- gamma FcR- muFcR- population of transformed cells derived from hairy cells. Depletion and readdition experiments demonstrated that cell-to-cell contact with T cells was necessary for the phenotypic change, while several observations indicated that the E+ population was not derived from T cells present before culture. The E positivity of the cultured cells was shown to be due to the possession of E receptor not acquired from the culture fluid, but the cells differed from true T cells in lacking both mature and immature T-cell antigens. The relevance of these in vitro observations to the continuing controversy concerning the nature of the hairy cell and to the in vivo fluctuations in immunologic phenotype not infrequently observed in hairy cell leukemia is briefly discussed.

Alterations in the Phenotype of Hairy Cells During Culture in the Presence of PHA: Requirement for T Cells

Culture studies of peripheral blood mononuclear cells from 7 entirely typical cases of hairy cell leukemia showed that after culture in the presence of PHA for 2--5 days, the predominant cell type changed from E- SIg+ CIg+ gamma FcR+ muFcR+ hairy cells to an E+ SIg- CIg- gamma FcR-muFcR- population of transformed cells derived from hairy cells. Depletion and readdition experiments demonstrated that cell-to-cell contact with T cells was necessary for the phenotypic change, while several observations indicated that the E+ population was not derived from T cells present before culture. The E positivity of the cultured cells was shown to be due to the possession of E receptor not acquired from the culture fluid, but the cells differed from true T cells in lacking both mature and immature T-cell antigens. The relevance of these in vitro observations to the continuing controversy concerning the nature of the hairy cell and to the in vivo fluctuations in immunologic phenotype not infrequently observed in hairy cell leukemia is briefly discussed.

Feedback regulation of immune responses by immune complexes; possible involvement of a suppressive lymphokine by FcR gamma-bearing B cell.

Inoculations of antigen-antibody complexes (immune complexes) with the intact Fc portion generates suppressor cells in vivo by binding to FcR gamma on B cells via Fc portions. The cell type responsible for the suppression appears to be B cells bearing FcR gamma. Neither T cells nor macrophages participate in both the inductive and effective phases of this type of regulation. The suppression caused by splenic B cells, previously stimulated with immune complexes in vivo, is mediated by humoral factor(s) released from them. The suppressive factor(s) have H-2 gene product(s) coded by the right-hand side of the H-2 gene complex, but not for FcR gamma themselves or immunoglobulins. It has shared component(s) with suppressive B cell factor (SBF) released from FcR gamma + B cells stimulated with immune complexes in vitro, and it resembles SBF in its mode of action. These findings indicate that immune complexes, the final products of antibody responses, control the immune responses by stimulating surface FcR gamma on B cells. It is of interest that this type of regulation functions in vivo.

Membrane expression of Fc-receptors in cultured leukemic cell lines. I. Induction of Fc-receptor in undifferentiated types of cells after passive modulation of lipid viscosity.

The role of lipid environment of plasma membrane for the expression of Fc receptor (gamma) (FcR gamma) were studied by physicochemical modification of the membranes of myeloid leukemia cell line cells, M1. Sterol binding polyene antibiotics such as amphotericine B or filipin inhibit the expression of FcR gamma during the differentiation of M1- cells by the CM obtained from the culture of mouse embryonic fibroblasts, suggesting the possible involvement of cholesterol molecules in the induction mechanism of FcR gamma. Low-temperature incubation at either 20 degrees C or 4 degrees C results in the increment of their membrane microviscosity, when tested by the fluorescence depolarization method, in company with the appearance of FcR gamma on approximately 60% of M1 cells when assayed by the EA-rosetting method. FcR gamma is also induced on approximately 40% of M1- cells by incubating cells with cholesterol-phosphatidyl-choline liposomes with the concomitant increase of their membrane microviscosity. Nevertheless, no phagocytosis of EA is observed in these cells treated with such physicochemical procedures. These findings imply that FcR gamma is embedded in the membrane of M1- cells and that physicochemical modification of the membrane lipid bilayers can induce the FcR gamma independent on their functional differentiation marked by the expression of phagocytic activity.

Immunological properties of Fc receptor on lymphocytes. 9. Functional analysis of FcR gamma+ and FcR gamma- lymphocytes in IgE antibody responses.

The present study was undertaken to clarify the function of FcR gamma+ and FcR gamma- lymphocytes in anti-DNP IgE antibody responses. The helper activity was observed predominantly in FcR gamma- T cells, but poorly in FcR gamma+ T cells, as in the case of IgG antibody responses, indicating that the absence of FcR gamma on T cell surface is a common surface characteristic to helper T cells, and that there is no close relationship between the determination of class specificity of Ig and T cell subsets divided by the presence or absence of FcR gamma. Furthermore, IgE antibody responses were more dependent upon helper T cells than IgG antibody responses. The data presented in this paper also demonstrated that the precursors of anti-DNP IgE antibody-forming cells were equally contained in both FcR gamma+ and FcR gamma- B cells, when they were transferred into recipients with helper T cells. This is a striking contrast to IgM/IgG antibody responses, in which only FcR gamma- B cells could differentiate into antibody-forming cells. Moreover, the precursor activity of FcR gamma+ and FcR gamma- B cells for IgE antibody-forming cells was not affected by the treatment of cells with suppressive B cell factor released from FcR gamma+ B cells after binding of antigen-antibody (IgG) complexes and known to suppress the proliferation of FcR gamma- IgM/IgG antibody-forming cell precursors. Thus, IgE B cells seem to be more heterogeneous than IgM/IgG B cells with respect to the FcR gamma marker and insensitive to the regulatory effect of FcR gamma+ B cells. IgE B cells are regulated differently from IgG B cells in B-B cell interactions.