Induction and function of Fc epsilon RII on YT cells; possible role of ADF/thioredoxin in Fc epsilon RII expression.

The regulation of low-affinity Fc receptor for IgE (Fc epsilon RII) and the characteristics of both membrane and soluble forms of Fc epsilon RII were studied using YT cell line. We found that YT cells, a human NK like cell line, expressed Fc epsilon RII after IL-1 stimulation. Cross-linking of Fc epsilon RII on IL-1-stimulated YT cells as well as the transfectant of Fc epsilon RII-cDNA (YTSER) resulted in the up-regulation of IL-2R alpha (p55/Tac). A 59 kDa protein phosphorylated at tyrosine residues was co-immunoprecipitated with Fc epsilon RII from YTSER lysate using H107 anti-Fc epsilon RII mAb. YTSER not only expressed Fc epsilon RII on their surface but also secreted soluble form of Fc epsilon RII (sFc epsilon RII/sCD23; IgE binding factor). Affinity purification revealed that sFc epsilon RII released from YTSER is heterogeneous and consisted of several proteins differing in molecular weight. Both EBV+ B cells and HTLV-1+ T cells are high producers of ATL derived factor (ADF)/thioredoxin (TRX) and express Fc epsilon RII and IL-2R alpha respectively. To clarify the mechanism of Fc epsilon RII and IL-2R alpha induction by ADF/TRX, we examined the effect of ADF/TRX on the bindability of nuclear factor kappa B (NF-kappa B), which is known to regulate IL-2R alpha gene expression. In the gel shift assay, ADF/TRX was shown to enhance the bindability of NF-kappa B to its responsive element.     

Tumour necrosis factor-alpha augments the expression of Fc IgE receptor (Fc epsilon RII/CD23) on human monocytic cell lines and down-regulates interleukin-4-driven Fc epsilon RII expression on monocytes.

We investigated the expression of the low affinity Fc IgE receptor (Fc epsilon RII/CD23) on the human monocytic cell lines U937, THP-1, Mono-Mac-6, and cultured human peripheral blood monocytes under stimulation with human tumour necrosis factor-alpha (TNF-alpha) and other cytokines. Fc epsilon RII was demonstrated by flow cytometry analysis employing the anti-Fc epsilon RII monoclonal antibody 3-5. TNF-alpha alone had a weak but significant stimulating effect on the Fc epsilon RII expression on the cell lines U937 and THP-1, and very modestly on Mono-Mac-6 cells. TNF-alpha strongly synergized with interferon-gamma (IFN-gamma) and interleukin-4 (IL-4). IFN-alpha per se was ineffectual, but was able to increase the TNF-alpha effect. Furthermore, the action of TNF-alpha was slightly augmented by human IL-6. Similar effects were noted with TNF-beta alone or in combination with other cytokines. Interestingly, on human monocytes TNF-alpha weakly reduced the basal level of Fc epsilon RII, and markedly diminished the IL-4-induced Fc epsilon RII expression. Our results indicate that several cytokines may interact in a cytokine network to modulate Fc epsilon RII expression on monocytic cell lines. On human blood monocytes, TNF-alpha, like IFN-gamma or IL-6, counteracts the IL-4-induced Fc epsilon RII expression. These data suggest different regulatory pathways of Fc epsilon RII expression on blood monocytes and myelomonocytic cell lines.     

Induction of Fc epsilon RII/CD23 on PHA-activated human peripheral blood T lymphocytes and association of fyn tyrosine kinase with Fc epsilon RII/CD23.

Despite the evidence for the expression of Fc epsilon RII/CD23, a glycoprotein that is a low-affinity Fc receptor for IgE, obtained on T cell lines and some pathological T cells, that of Fc epsilon RII/CD23 on normal human T cells is still unclear. We studied the emergence of T cells bearing Fc epsilon RII/CD23 in short-term culture of normal human peripheral blood mononuclear cells stimulated with 15 microliters/ml phytohemagglutinin (PHA). Using two-dimension flow cytometry, more than 10% of Fc epsilon RII/CD23(+) cells were shown to co-express CD3 antigen. Both CD4(+) and CD8(+) T cells expressed Fc epsilon RII/CD23. The expression of mRNA for Fc epsilon RII/CD23 on PHA and IL-4 stimulated PBMC was demonstrated by northern blotting and in-situ hybridization. The mechanism of signal transduction through Fc epsilon RII/CD23 was dissected by transfection of cDNA coding for Fc epsilon RII to the human natural killer-like cell line YT, activation of which was easily detected by the induction of interleukin-2 receptor/p55 (Tac). Cross-linking of Fc epsilon RII/CD23 with H107 anti-Fc epsilon RII monoclonal antibody enhanced IL-2R/p55 expression on YT cells transfected with Fc epsilon RII cDNA (YTSER). A possible involvement of protein-tyrosine kinase in the Fc epsilon RII-mediated signal transduction was studied using YTSER. Fc epsilon RII was physically associated with an src-family tyrosine kinase p59fyn and not with p56lck, which was also found in YT cells. Recently it was reported that p59fyn was associated with T-cell antigen receptor. Our results collectively suggest the multiple function of p59fyn which may be implicated in the Fc epsilon RII-mediated activation signal in YT cells.     

Mesangial alterations in steroid-responsive minimal change nephrotic syndrome

Summary Renal biopsies from 25 children with steroid-sensitive minimal change nephrotic syndrome were evaluated retrospectively to determine whether there is any relation between the morphological changes and the frequency of relapses. Biopsy material was examined by light-, immunofluorescence-, and electron microscopy, and by morphometric methods. The patients were divided in a group of 15 children with frequent relapses (FR) and another group of 10 children with an absence of, or only infrequent, relapses (NR/IR). Semiquantitative evaluation of biopsy specimens disclosed no significant differences between groups, but morphometric measurements performed on toluidine stained semithin sections showed a significant increase of mesangial nuclei in FR compared with NR/IR (P<0.01). Furthermore, the mean area of mesangial nuclei was decreased and the relative frequency of smaller nuclear profiles was higher in patients with FR compared to NR/IR (p<0.01). These findings suggest mesangial cell activation in FR which may be related to a longer course of the disease prior to renal biopsy (mean 4.0 years in FR vs. 1.4 years in NR/IR). In our opinion, morphometric assessment of discrete mesangial alterations is a promising method for exploring clinicopathological correlations in minimal change nephrotic syndrome.Presented in part at the 5th International Symposium of Pediatric Nephrology, Philadelphia, USA, October 6–10, 1980     

Quantitative and qualitative analysis of the Fc receptor for IgE (Fc epsilon RII) on human eosinophils

In order to characterize the Fc receptor for IgE (Fc epsilon RII) on human eosinophils, we have compared the binding of human IgE myeloma protein to that of a monoclonal antibody (mAb BB10) directed against a common antigenic determinant of the Fc epsilon RII present on eosinophils, platelets and macrophages. Scatchard analysis of the binding to human eosinophils of the BB10 mAb revealed a linear monophasic binding curve, with a binding affinity of 1.17 x 10(7) M-1 and a number of 10(5) binding sites per cell. Biochemical analysis of the human eosinophil Fc epsilon R, performed by immunosorbent chromatography with either BB10 mAb or IgE, showed under nonreducing conditions a major component of 200 kDa. Under reducing conditions, 3 peptide fragments were obtained, with molecular masses of 45-50, 23 and 15 kDa. Finally, comparative analysis suggested that the Fc epsilon RII of human eosinophils and of a human macrophage cell line (U937) are structurally related and differ from the high-affinity Fc epsilon RI present on basophilic granulocytes.     

Mesangial myofibroblastic transformation in steroid-dependent minimal change nephrotic syndrome

Patients with minimal change nephrotic syndrome (MCNS) occasionally show frequent relapses with proteinuria after cessation of steroid treatment, even though no significant pathological abnormalities are found in the glomeruli, compared with those in nonrelapsed and good-prognosis cases of MCNS. To resolve this contradiction, we immunohistochemically and ultrastructurally examined a biopsied renal tissue of a patient who showed glomerular features of MCNS and frequent clinical relapses. Immunohistochemistry demonstrated the overexpression of alpha-smooth muscle actin (ASMA) and vimentin in glomerular mesangial cells despite no mesangial cell proliferation, compared with nine nonrelapsed cases of MCNS. These facts may be an important clue to the investigation of the pathogenesis of steroid-dependent MCNS with frequent relapses. Furthermore, the immunohistochemical examination of ASMA and vimentin may be useful to detect mesangial myofibroblastic transformation that is not demonstrated in conventional light microscopy and immunofluorescence study.     

Immune status of children suffering from minimal change nephrotic syndrome.

Twenty five children suffering from minimal change nephrotic syndrome were studied for immunological alterations at different stages of this disease i.e., onset, relapse and remission. Changes were found mainly at onset and during relapse in the form of altered helper and suppressor cell ratio, depressed delayed cutaneous hypersensitivity reaction, decreased S-IgG bearing lymphocytes with low serum IgG concentration, and increased S-IgM bearing lymphocytes with high serum IgM concentration. Majority of these parameters returned to normal values during remission. Serum IgE was found high at all stages of this. These alterations suggest defects in cell mediated immunity resulting in secretion of some substance which modifies the glomerular anionic charges.     

Impaired immunoglobulin G production in minimal change nephrotic syndrome in adults.

Serum IgG, T and B cell subsets, cytoplasmic IgG positive cells (cB gamma) and IgG in the medium (cIgG) of a 5 day culture of peripheral lymphocytes in both stimulated and non-stimulated (spontaneous) conditions with pokeweed mitogen (PWM) were studied in 30 adult patients with minimal change nephrotic syndrome (MCNS). In the nephrotic phase (11 patients), surface IgG positive B cells (sB gamma) and spontaneous cB gamma increased (P less than 0.05), whereas PWM-stimulated cIgG did not increase, and serum IgG decreased significantly (P less than 0.05). The cB gamma/sB gamma ratio calculated as an index of IgG synthesis in B cells increased spontaneously (P less than 0.05), but did not increase under PWM-stimulation. The cIgG/cB gamma ratio as an index of IgG secretion from each matured B cell, reduced in both spontaneous and stimulated conditions (P less than 0.05, P less than 0.01, respectively). In the phase of unstable remission maintained by steroid therapy (10 patients), these parameters tended to normalize and the OKT4/OKT8 ratio decreased (P less than 0.05), while the ratio remained unchanged in the nephrotic phase. However, after discontinuation of steroid (nine patients), spontaneous cB gamma and the spontaneous cB gamma/sB gamma ratio were again increased, and the cIgG/cB gamma ratio decreased (P less than 0.05) as observed in the nephrotic phase. These results suggest that B cells in patients with MCNS both in the nephrotic state and stable remission after discontinuation of steroid are activated spontaneously, but the secretory process of IgG from the matured cells is impaired, and that steroid improves these abnormalities.     

Fc epsilon receptor II/CD23-positive lymphocytes in atopic dermatitis. I. The proportion of Fc epsilon RII+ lymphocytes correlates with the extent of skin lesion.

Cells expressing Fc receptors for IgE (Fc epsilon RII) were identified in the peripheral blood from patients with atopic dermatitis and with eczematous dermatitis, and normal non-atopic subjects by using monoclonal antibodies to human lymphocyte Fc epsilon RII, and to lymphoid cell-surface antigens by immunofluorescence staining. Based on the extent of the dermatitis patients were classified as severe (greater than 50% skin surface involved), moderate (50-10%) and mild (less than 10%). Patients with severe and moderate atopic dermatitis had 5.9% and 5.7% Fc epsilon RII+ peripheral blood mononuclear cells (PBMC), respectively, that were significantly higher than percentages in mild atopic dermatitis patients (2.6%), severe to moderate eczematous dermatitis patients (2.3%), mild eczematous dermatitis patients (2.2%) and normal individuals (1.7%)(0.05 greater than P). In severe and moderate atopic dermatitis patients, 10% of Fc epsilon RII+ PBMC were T cells that preferentially expressed CD8, and the remainder B cells and monocytes. Fc epsilon RII+ T cells comprised 1% of peripheral T cells, while half or more of peripheral B cells expressed Fc epsilon RII. In mild atopic dermatitis patients, eczematous dermatitis patients and normal subjects. Fc epsilon RII were expressed exclusively on 25-35% of peripheral B cells. Short-term treatment and long-term follow-up of atopic dermatitis patients revealed that changes in the skin condition were related closely to fluctuations in the proportion of Fc epsilon RII+ PBMC. Total serum IgE levels and atopic respiratory allergy did not influence the percentage of Fc epsilon RII+ PBMC. These findings suggest that the percentage of Fc epsilon RII+ PBMC reflects the extent of atopic dermatitis.     

CD23/Fc epsilon RII and its soluble fragments can form oligomers on the cell surface and in solution.

Human CD23 (also known as Fc epsilon RII) is a 45,000 MW glycoprotein with homology to C-type animal lectins. It is involved in B-cell differentiation and IgE regulation, and is naturally cleaved to give soluble products of 37,000, 33,000, 29,000, 25,000 and 16,000 MW. Previous work has suggested that the region between the transmembrane sequence and the extracellular lectin head is capable of forming an alpha-helical coiled coil, one of the main consequences of which would be formation of dimers or trimers. Here we present protein-protein cross-linking data showing that CD23 forms trimers on the cell surface and hexamers in solution, and we use several different fragments to determine the regions of the protein involved in this self-association. The region of the putative coiled coil is indeed responsible for trimerization, with additional interactions between the lectin heads resulting in the formation of hexamers observed in solution.     

Intestinal epithelial cells express the CD23/Fc epsilon RII molecule: enhanced expression in enteropathies.

Immunohistochemical analysis of normal human intestine revealed that two anti-CD23 monoclonal antibodies (mAb), EBVCS 1 and EBVCS 2, reacted with human intestinal epithelial cells. Both mAb exhibited an exclusive reactivity with epithelial cells of the small and large bowels. Staining with both EBVCS 1 and EBVCS 2 was localized on the apical and basal sides of enterocytes. Enhanced expression of CD23 on gut epithelial cells was found in inflammatory bowel diseases, in children with food intolerance to cows' milk proteins and in a young infant with severe autoimmune enteropathy. Western blot analysis of anti-CD23 mAb reactivity with gut epithelial cell extracts showed the presence of a non-reducible 42,000-45,000 M(r) polypeptide compatible with the membrane form of the intact CD23 molecule. These data show that CD23 is constitutively expressed by intestinal epithelial cells and that its expression is enhanced in enteropathies.     

Inhibition of human interleukin 4-induced IgE synthesis by a subset of anti-CD23/Fc epsilon RII monoclonal antibodies.

Specific monoclonal antibodies (mAb) directed against the CD23 antigen were used to study human interleukin 4 (hIL4)-induced IgE production by blood and tonsillar mononuclear cells. Both peripheral blood and tonsillar mononuclear cells stimulated by hIL4 expressed membrane CD23 as detected by the binding of all anti-CD23 mAb. Nevertheless, two sets of anti-CD23 mAb could be distinguished. The first set, including mAb 25, was able to decrease significantly hIL4-induced IgE synthesis by mononuclear cells. The second set, including EBVCS#1, did not affect hIL4-induced IgE synthesis. All the anti-CD23 mAb were able to bind specifically to a human B cell line expressing recombinant CD23. Inhibition experiments revealed that the two sets of anti-CD23 mAb did not recognize the same epitope on the CD23 antigen. In fact, all the anti-CD23 mAb, except EBVCS#1, were able to inhibit IgE binding to CD23 on RPMI 8866 cells. Moreover, the first set of antibodies, which decreased IgE production, was able to up-regulate membrane CD23 expression on hIL4-stimulated tonsillar mononuclear cells. Conversely, EBVCS#1, which had no effect on IgE production, did not affect hIL4-induced CD23 expression. These results indicate that CD23 plays a key role in human IgE synthesis.