Favourable effect of regular intake of fermented milk containing Lactobacillus johnsonii on Helicobacter pylori associated gastritis.

BACKGROUND: Lactobacillus johnsonii (Lj1) had an in vitro and in vivo inhibitory effect on Helicobacter pylori. Fermented milk containing Lj1 (LC1), coadministered with antibiotics had a favourable effect on H. pylori gastritis. AIM: Evaluate the effect of LC1 intake without antibiotics on H. pylori gastritis. METHODS: Fifty H. pylori positive healthy volunteers were randomised in a double-blind study to LC1 or placebo. Gastric biopsies from the antrum and corpus were obtained before, and after 3 and 16 weeks of treatment, for histology and quantitative cultures. RESULTS: Severity and activity of antral gastritis was reduced after 16-week LC1 intake (pretreatment and 16-week inflammatory cell score: 6.0 +/- 0.8 vs. 5.3 +/- 0.1; P=0.04). H. pylori density decreased in the antrum after LC1 intake (3-week: 4.4 +/- 0.6; 16-week: 4.3 +/- 0.5 log10 colony forming units (cfu) vs. pretreatment 4.5 +/- 0.4 log10 cfu; P=0.04, respectively). Mucus thickness increased after 16 weeks of LC1 consumption (change of mucus thickness with LC1 and placebo in the antrum: 0.6 +/- 1.3 vs. -0.2 +/- 1.0, P=0.01; in the corpus: 0.3 +/- 1.1 vs. -0.6 +/- 1.5, P=0.03). CONCLUSION: LC1 intake had a favourable, albeit weak, effect on H. pylori associated gastritis, particularly in the antrum. Regular ingestion of fermented milk containing L. johnsonii may reduce the risk of developing disorders associated with high degrees of gastric inflammation and mucus depletion.

Human lung dendritic cells have an immature phenotype with efficient mannose receptors.

Dendritic cells (DC) can be present at distinct stages of differentiation within the immune system. Sallusto and colleagues have recently described an in vitro culture system suitable for analyzing the maturation processes of DC (Sallusto and colleagues, J. Exp. Med. 1994;179:1109-1118). Monocytes cultured for 6 d in the presence of granulocyte macrophage colony-stimulating factor and interleukin-4 develop into immature DC with a high endocytic capacity but a low capacity to stimulate T cells. When challenged by lipopolysaccharide, these cells upregulate costimulatory molecules, express CD83, and become mature DC. CCR1 and CCR5 chemokine receptors are highly expressed on immature DC and downregulated on mature DC. This in vitro system was used to characterize human lung DC. Lung DC were shown to express some characteristics of in vitro immature DC. These are: (1) low expression of the costimulatory molecules CD40, CD80, and CD86; (2) poor expression of the differentiation marker CD83 and no CD1a; and (3) good capacity to incorporate dextran. Lung DC express moderate levels of CCR1 and CCR5. However, lung DC, like in vitro mature DC, express high levels of major histocompatibility complex Class II molecules, show low expression of CD14 and CD64, and are characterized by their high capacity to stimulate allogeneic T cells to proliferate during mixed leukocyte reactions (MLRs). Although lung DC express low levels of CD80 and CD86, the important role of these costimulatory molecules in inducing high MLR was demonstrated by using blocking antibodies. Therefore, while lung DC have overall a phenotype and an endocytic capacity close to in vitro immature DC, they share, like in vitro mature DC, a powerful capacity to stimulate T cells.

Differential regulation of tumor necrosing factor-alpha (TNF-alpha) and interleukin-10 (IL-10) secretion by protein kinase and phosphatase inhibitors in human alveolar macrophages.

IL-10, a cytokine first identified as a product of cloned Th2 lymphocytes, is also produced by monocytes/macrophages. By its ability to inhibit cytokine synthesis and the expression of surface antigens, IL-10 is able to temper inflammation. In contrast, TNF-alpha plays a key role in acute and chronic inflammation and has been implicated in several forms of lung injury. The objective of this study was to investigate whether activators or inhibitors of LPS-activated signalling pathways might be able to dissociate TNF-alpha from IL-10 secretion in alveolar macrophages (AM). The results show that PMA activates expression of TNF-alpha without inducing IL-10 expression. We further demonstrate that LPS-induced TNF-alpha secretion is independent of PKC activation and can be increased by inhibitors of the serine/threonine phosphatases PP1 and PP2A. In contrast, LPS-mediated IL-10 secretion is down-regulated by PMA and inhibitors of PP1 and PP2A. Addition of PKC inhibitors reverses the PMA-mediated down-regulation of LPS-induced IL-10 secretion, indicating that PKC, once activated in vivo, might play a prominent role in controlling the secretion of IL-10 by AM. This study provides evidence that the PKC activator PMA and the phosphatase inhibitor calyculin A are able to dissociate TNF-alpha from IL-10 secretion by AM. Our data further indicate that LPS-mediated activation of certain signalling molecules has different consequences on the secretion of TNF-alpha or IL-10 by AM, an observation which may be important for the modulation of immune and inflammatory processes.

Interleukin-12 production by human alveolar macrophages is controlled by the autocrine production of interleukin-10.

By releasing interleukin (IL)-12 in the lung, alveolar macrophages (AM) may profoundly modify an immune response. The autocrine regulation of the heterodimeric, biologically active form of IL-12 (IL-12 p70) by IL-10 was studied, as well as the expression of its subunits of 35 kD (p35) and 40 kD (p40). AM cultured in medium alone expressed only p35 mRNA. Both p35 and p40 mRNA levels were induced by lipopolysaccharide (LPS) and were further increased by interferon-gamma (IFN-gamma). LPS alone induced IL-12 p40 but not IL-12 p70 production in monocytes and in AM. However, IL-12 p70 was released when the autocrine production of IL-10 was neutralized by IL-10 blocking antibody, and IL-12 p40 production increased. Although IFN-gamma markedly decreased LPS-induced IL-10 production in AM, neutralizing IL-10 further enhanced the level of LPS and IFN-gamma-induced IL-12 p70 in AM. In contrast, neutralizing the trace amount of IL-10 released by AM stimulated by CD40 crosslinking and IFN-gamma did not increase IL-12 p70. Thus, IL-12 p70 production by AM appears to be tightly controlled by the autocrine release of IL-10 when stimulated by LPS, or by LPS and IFN-gamma, whereas CD40 crosslinking triggered IL-12 p70 production in the absence of autocrine regulation by IL-10.

Alveolar macrophages in sarcoidosis coexpress high levels of CD86 (B7.2), CD40, and CD30L.

Alveolar macrophages (AM) from sarcoid patients have been shown to be good antigen presenting cells (APC) unlike normal AM which are usually ineffective. We demonstrate in ten consecutive sarcoid patients that most of their AM, unlike normal AM, do coexpress high levels of CD86, CD40, and CD30L, all known to be important for T-cell activation. CD80 is also slightly more expressed on sarcoid AM than on normal AM, but is detected on only 26 +/- 6% (mean +/- SEM) of sarcoid AM. A good correlation is present between the percentage of sarcoid AM expressing CD86 and CD40 or CD86 and CD30L. However, no correlation is found between the percentage of CD80 and CD86 positive AM in these same patients. Blocking antibodies against CD86 were able to reduce by more than 80% allogeneic T-cell proliferation induced by the AM of sarcoid patients. This study provides evidence that AM can, in pathologic states such as sarcoidosis, express functional costimulatory molecules for T-cell activation such as CD86, thought to be rather specific for more professional APC such as dendritic cells.

Production of prostaglandin E2 and collagenase is inhibited by the recombinant soluble tumour necrosis factor receptor p55-human gamma 3 fusion protein at concentrations a hundred-fold lower than those decreasing T cell activation.

TNF-alpha and lymphotoxin alpha (TNF-beta) are pleiotropic cytokines with regulatory functions in inflammatory reactions and T cell activation. Natural TNF inhibitors such as soluble TNF-binding proteins, i.e. TNFsR55 and TNFsR75, are shed from white blood cells and probably other cells. These naturally occurring inhibitors of TNF are shown to be 10 times less effective than the bivalent antagonist of TNF, recombinant soluble TNF receptor p55-human gamma 3 fusion protein (rsTNFR-p55h gamma 3), in controlling the release of prostaglandin E2 (PGE2) and collagenase by fibroblasts, as well as in controlling T cell proliferation. In order to block the action of rhTNF-alpha added to fibroblasts, a fivefold excess of rsTNFR-p55h gamma 3 was sufficient, but concentrations of a hundred to a thousand times higher were required to obtain a significant inhibition of T cell activation. This concentration appears to be required to block membrane-bound TNF-alpha on peripheral blood mononuclear cells as shown by Scatchard analysis. We additionally show that rsTNFR-p55h gamma 3 at high concentrations also blocks T cell activation by dendritic cells. In conclusion rsTNFR-p55h gamma 3 has a much higher anti-inflammatory effect than immunosuppressive effect.

[Adult form of GM2-gangliosidosis: a man and 2 sisters with hexosaminidase-A and -B deficiency (Sandhoff disease) and literature review]. / Adulte Form der GM2-Gangliosidose: drei Geschwister mit Hexosaminidase-A- und -B-Mangel (Morbus Sandhoff) und Literaturübersicht.

Three adult siblings had atypical progressive spinal muscular atrophy of the limb-girdle type, predominantly sensory polyneuropathy and cerebellar ataxia. Hexosaminidase A and B activity was profoundly decreased in serum, leukocytes and cultured fibroblasts. GM2-gangliosidosis, variant O (Sandhoff disease) was diagnosed. Mechano-allodynia was the presenting symptom in two of the patients. After 50 years of disease evolution, the patients led an independent life and were intellectually normal. The literature on the adult form of GM2-gangliosidosis is reviewed.

Contact-dependent stimulation of monocytic cells and neutrophils by stimulated human T-cell clones.

By means of direct cell-cell contact, fixed, stimulated T lymphocytes trigger the production of interleukin-1 beta (IL-1 beta) and tumour necrosis factor by monocytes and prime polymorphonuclear leucocytes (PMN) for the respiratory burst induced by N-formyl-methionyl-leucyl-phenylalanine (FMLP). In order to assess whether this activity is displayed by a particular subpopulation of T lymphocytes, 88 T-cell clones (TCC) were generated from healthy human peripheral blood. The clones were stimulated by Phaseolus vulgaris agglutinin (PHA) and phorbol 12-myristate acetate monocytic cell line THP-1. All fixed, stimulated TCC induced IL-1 beta production by THP-1 cells, although to varied degrees. The activity of TCC on THP-1 cells correlated with their activity on PMN (r2 = 0.84, P < 0.001), suggesting that this pro-inflammatory activity of TCC may similarly affect both types of infiltrating cells. The ability of TCC to modulate target cells did not correlate with either their phenotype (CD4 or CD8) or their cytokine production profile (interferon-gamma, IL-4 and IL-6), although it tended to correlate inversely with their capacity to produce IL-6 (P < 0.02). These observations suggest that a large proportion, if not all, of human peripheral blood T lymphocytes may potentially affect monocytes and PMN by direct cell-cell contact. This activity may be relevant to the maintenance of chronic inflammation.

Direct contact between T lymphocytes and monocytes is a major pathway for induction of metalloproteinase expression.

Monocytes and macrophages can modulate the turnover of extracellular matrix by producing metalloproteinases such as interstitial collagenase and 92-kDa gelatinase as well as tissue inhibitor of metalloproteinases. To study mechanisms of metalloproteinase induction in human mononuclear phagocytes, the effects of direct cell-cell contact between activated T lymphocytes and the human monocytic cell line THP-1 were determined. T cells were first activated with phorbol 12-myristate 13-acetate and phytohemagglutinin for 24 h, fixed with paraformaldehyde, and then exposed to THP-1 cells for 48 h. Upon contact with fixed activated T lymphocytes, a massive induction in the expression of both proteinases and tissue inhibitor of metalloproteinases was observed, whereas unstimulated T cells had no effect. Stimulation of metalloproteinase biosynthesis by THP-1 cells was mimicked by a membrane preparation derived from activated T cell lines, whereas cytosol and nuclear fractions of the T cells were ineffective. Furthermore, activated T lymphocytes exposed to trypsin, tunicamycin, or cycloheximide lost the capacity to stimulate THP-1 cells upon subsequent contact, implying the involvement of cell-surface glycoproteins. Similar induction of metalloproteinases by direct contact with activated T cells was also observed using normal blood monocytes as the target cells, and stimulation of monocyte metalloproteinases by T cell contact occurs at a pretranslational level. Consequently, cell-cell contact may represent an important biological mechanism for potentiating the inflammatory response that leads to extracellular matrix destruction.

Adhesion molecules and cytokine production.

The exchange of cross-talks between cells relies on soluble factors or direct cell-cell contact. Soluble factors increase the expression of cell surface molecules that activate adjacent cells by direct contact to produce cytokines. In the lung, dendritic cells are potent inducers of T-cell proliferation, and the interaction between the two leads to the production of high amounts of TNF alpha and TNF beta. Of the molecules involved in these biologic functions, LFA-3, CD11c, and the combination of beta 1 and beta 2 integrins are the most efficient. However, blocking TNF alpha or TNF beta production does not affect the alloreaction. The interaction between activated T cells and monocytes resulted in a large production of IL-1 beta. In this reaction, CD69, CD2, and the beta 2 integrins (CD11a, b, c, and CD18) and also other molecules such as a 25- to 35-kD glycoprotein play an important part. Finally, interaction between monocytes and fibroblasts leads to the production of large amounts of collagenase and PGE2 by fibroblasts. Cell-associated IL-1, particularly IL-1 alpha and membrane-bound TNF alpha, can also play a crucial role in the process of cell-cell interaction. This interaction may be controlled by inhibitors to IL-1 and TNF.

Cell surface glycoproteins expressed on activated human T cells induce production of interleukin-1 beta by monocytic cells: a possible role of CD69.

In many immunoinflammatory diseases, macrophages, by producing interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF alpha), stimulate protease secretion in fibroblasts, thus contributing to tissue destruction. Monocyte/macrophage activation is prompted by soluble factors released by activated T cells as well as by cell-cell contact. Indeed, previous studies have shown that monocytes exposed to paraformaldehyde (PFA)-fixed, activated T cells produced high amounts of IL-1 beta. In this report, we used the T cell line HUT-78 to further characterize the T cell factor(s) responsible for monocyte activation by cell-cell contact. After subcellular fractionation, most of the activity was found in the cellular membrane fraction of PHA/PMA-stimulated HUT-78 cells, and proved to be due to glycoproteins, following trypsin digestion and tunicamycin treatment. HUT-78 cells acquired the capacity to stimulate monocytic cells after as little as 1h of stimulation. De novo protein synthesis was required for the expression of the IL-1 beta inducing factor, as shown by cycloheximide treatment. When membrane proteins of PHA/PMA-stimulated HUT-78 cells were separated on SDS-polyacrylamide gel, a peak of stimulatory activity was observed at Mr--25-35 x 10(3). By using specific cytokine inhibitors or blocking mAbs, we ascertained that cell-associated cytokines (IL-1, IL-2, IFN gamma and GM-CSF) were not involved in monocyte activation by cell contact. Anti-CD2 and -CD11a (LFA-1) mAbs partially blocked IL-1 beta production by -25% and -35%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

gp33-38, an early human T cell activation antigen.

We describe an activation Ag Me14/D12 that appears early after T cell activation and is absent in resting T lymphocytes. Me14/D12 is a nondisulfide-linked heterodimeric structure containing two polypeptide chains of 33,000 and 38,000 Da. The expression of Me14/D12 on resting T lymphocytes can be induced by different activation stimuli such as the lectins PHA and Con A, the phorbol ester PMA, and anti-CD3 mAb. The induction of mRNA for Me14/D12 (gp33-38) in PHA-activated T lymphocytes precedes that of IL-2R gene transcripts by more than 20 h. Me14/D12 mRNA was detectable as early as 2 h after the onset of activation and mRNA for the IL-2R only after 24 h. The surface expression of Me14/D12 was detectable between 12 and 24 h after activation and was maximal between 24 and 48 h. Several T leukemia cell lines express the Me14/D12 Ag. On Me14/D12- cell lines, PMA and IFN-gamma induced surface expression of Me14/D12. Once Me14/D12 Ag were expressed on Jurkat cells after stimulation with either PMA or IFN-gamma, the binding of mAb Me14/D12 induced the production of significant amounts of IL-2 and of Ca2+ mobilization from internal stores. Comparative biochemical studies clearly demonstrate that Me14/D12 (gp33-38) is different from the CD69 molecular complex defined by mAb MLR3 and AIM.

A positive signal is transduced via surface CD4 molecules.

We have previously reported on the identification of a monoclonal antibody (mAb) directed against the human CD4 antigen which is capable of activating CD4+ peripheral blood T cells in the absence of other stimuli. In the present study, we extended these findings by demonstrating that the mAb, termed B66, was able to induce the production of interleukin-2 in murine T-cell hybridoma transfectants expressing the human CD4 glycoprotein. Moreover, we found that incubation of Jurkat cells with mAb B66 resulted in the nearly complete disappearance of both CD4 and CD3 from the cell surface, whereas modulation of CD4, but not CD3, was observed after incubation with a non-stimulatory anti-CD4 mAb. Similar results were obtained in modulation experiments using human CD4-expressing murine transfectants. It is thus conceivable that the stimulatory activity of anti-CD4 mAb B66 may be associated with an effect on the CD3 molecular complex. While the biochemical basis for the unique stimulatory activity of mAb B66 has yet to be defined, these findings provide direct evidence that cross-linking of CD4 alone may cause T-cell activation, thus supporting the notion that this glycoprotein can transduce independent positive signals upon binding to class II major histocompatibility complex molecules.

Antigen-independent activation of T cells mediated by a novel cell surface heterodimer (Tp135-145).

A new heterodimeric structure, Tp135-145, which can mediate interleukin 2 (IL2) production and Ca2+ mobilization by Jurkat cells is described. This structure was identified by a monoclonal antibody, MX24, on the surface of either T3/TcR+ or T3/TcR- human T cell lines as well as on B cell lines. Biochemical studies showed that antibody MX24 precipitated two polypeptide chains of 135 and 145 kDa, respectively, in lysates from 125I-labeled T cells. After reduction the 135-kDa polypeptide chain shifted to 140 kDa, whereas the molecular mass of the other polypeptide remained unchanged. The apparent molecular masses of the desialylated polypeptides differed by 5 kDa. No common peptide fragments between the two polypeptide chains were found after limited proteolysis by Staphylococcus aureus V8 protease. The expression of Tp135-145 was independent of the expression of the T3/TcR molecular complex. Incubation of Jurkat cells with anti-TcR or anti-T3 monoclonal antibody induced complete modulation only of the T3/TcR complex but not of Tp135-145. Conversely complete modulation of Tp135-145 was observed after incubation of these cells with MX24 antibody. Functional studies showed that anti-Tp135-145 antibody MX24 induced high levels of IL2 production in Jurkat cells. In addition, incubation of these cells with MX24 resulted in Ca2+ mobilization from internal stores. In peripheral blood, Tp135-145 was found to be expressed by 39%-76% of resting T cells in individual donors. Two-color flow microfluorimetry showed that the Tp135-145+ cells were equally distributed on the CD4+ and CD8+ subsets. Incubation of peripheral blood T cells with antibody MX24 resulted in IL2 production and cell proliferation. Taken together these results suggest that Tp135-145 is a novel surface molecule involved in antigen-independent pathway of T cell activation.

A novel IFN-gamma regulated human melanoma associated antigen gp33-38 defined by monoclonal antibody Me14/D12. I. Identification and immunochemical characterization.

A novel melanoma-associated differentiation Ag whose surface expression can be enhanced or induced by IFN-gamma was identified by mAb Me14/D12. Testing of numerous tumor cell lines and tumor tissue sections showed that Me14/D12-defined Ag was present not only on melanoma but also on other tumor lines of neuroectodermal origin such as gliomas and neuroblastomas and on some lymphoblastic B cell lines, on monocytes and macrophages. Immunoprecipitation by mAb Me14/D12 of lysates from [35S]methionine-labeled melanoma cells analyzed by SDS-PAGE revealed two polypeptide chains of 33 and 38 KDa, both under reducing and nonreducing conditions. Cross-linking experiments indicated that the two chains were present at the cell surface as a dimeric structure. Two-dimensional gel electrophoresis showed that the two chains of 33 and 38 KDa had isoelectric points of 6.2 and 5.7, respectively. Treatment of the melanoma cells with tunicamycin, an inhibitor of N-linked glycosylation, resulted in a reduction of the Mr from 33 to 24 KDa and from 38 to 26 KDa. Peptide maps obtained after Staphylococcus aureus V8 protease digestion showed no shared peptides between the two chains. Although biochemical data indicate that Me14/D12 molecules do not correspond to any known MHC class II Ag, their dimeric structure, tissue distribution, and regulation of IFN-gamma suggest that they could represent a new member of the MHC class II family.

Stimulation and proliferation of CD4+ peripheral blood T lymphocytes induced by an anti-CD4 monoclonal antibody.

There is experimental evidence that the CD4 molecule participates in the antigen-driven activation of T cells expressing this surface glycoprotein. Whether CD4, a member of the immunoglobulin supergene family, acts as a ligand-binding molecule and/or is directly involved in the activation pathway has yet to be established. In this study, we show that human CD4+ lymphocytes can be activated by exposure to the anti-CD4 monoclonal antibody (mAb) B66. Normal peripheral blood CD4+ cells were induced to proliferate and to synthesize interleukin 2 (IL 2) by the antibody. The specificity of the antibody stimulatory activity was tested by using IL 2-producing clones bearing either CD4 or CD8 on their surface. IL 2 production was induced by mAb B66 in CD4+, but not CD8+, clones, whereas both types of clones responded to stimulation by the anti-CD3 mAb Leu-4. Despite its unique stimulatory activity, mAb B66 shared with other anti-CD4 antibodies the ability to inhibit the specific cytolytic activity of CD4+ effector cells. These results clearly indicate that cross-linking of surface CD4 molecules with appropriate antibodies can fully activate CD4+ lymphocytes. Whether the natural ligand for CD4 can trigger this activation pathway remains to be defined.

Identification of a novel 45-kDa cell surface molecule involved in activation of the human Jurkat T cell line.

This report describes a surface molecule, Tp45, which appears to be involved in interleukin 2 production and Ca2+ mobilization by Jurkat cells. The Tp45 molecule was identified by a monoclonal antibody, MX13, on the surface of either T3/TCR+ or T3/TCR- human T cell lines. Biochemical data showed that mAb MX13 precipitated a single polypeptide chain of 45 kDa both under reduced and nonreduced conditions from lysates of 125I-surface-labeled cells. Sequential immunodepletion experiments using lysates of 125I-labeled T3/TCR+ cells showed that Tp45 was distinct from the alpha chain of the TCR complex. However, incubation of such cells with either anti-T3 or anti-TCR monoclonal antibody induced complete modulation of both the T3/TCR complex and Tp45. Conversely, complete modulation of both Tp45 and the T3/TCR complex was observed after incubation with anti-Tp45 antibody. Functional studies showed that anti-Tp45 antibody induced high levels of interleukin 2 production in Jurkat cells. In addition, incubation of these cells with the antibody resulted in Ca2+ mobilization from internal stores. Anti-Tp45 antibody reacted with 3-19% peripheral blood (E-rosette-positive) T cells in individual donors. The magnitude of the proliferative response elicited by anti-Tp45 antibody for peripheral blood T cells was lower than that induced by an anti-T3 antibody. This observation is compatible with the idea that only a subpopulation of T cells is reactive with anti-Tp45. Multicolor flow cytometry analysis showed that the Tp45+ cells belong preferentially to the T8 subset.

Phorbol 12-myristate 13-acetate induces surface expression of T3 on human immature T cell lines with and without concomitant expression of the T cell antigen receptor complex.

The T3 complex is known to be expressed on the cell surface of mature T cells together with either the alpha-beta heterodimeric T cell receptor (TCR) or the TCR gamma protein. In a number of immature T cell malignancies, however, T3 has been described exclusively in the cytoplasm. We have investigated five such T cell lines with cytoplasmic T3 and could demonstrate by biosynthetic labeling the presence of the alpha and beta chains of the TCR in the cytoplasm of two of them, CEM and Ichikawa. No surface TCR alpha-beta protein could be detected by staining with the WT31 antibody. These observations, therefore, argue against the concept that expression of the TCR alpha chain controls the surface expression of the T3/TCR complex. Interestingly, phorbol 12-myristate 13-acetate (PMA) induced cell surface expression of T3 protein in these two cell lines only. Moreover, on surface-iodinated CEM cells no association of T3 and TCR molecules could be demonstrated after treatment with PMA, and expression of TCR alpha and beta chains was limited to the cytoplasm. In Ichikawa cells, however, PMA induced surface expression of a mature T3/TCR complex. Our findings indicate that separate regulatory mechanisms may exist for the surface expression of the T3 proteins and for the assembly of the T3/TCR complex.

A novel 90-kDa polypeptide (Tp90) possibly involved in an antigen-independent pathway of T cell activation.

A novel surface molecule, Tp90, is described which appears to be involved in an antigen-independent pathway of human T lymphocyte activation. The Tp90 molecule was identified by a monoclonal antibody (mAb), MX20, obtained from a fusion using spleen cells of a mouse immunized with cells from two T cell leukemia lines, Jurkat and HPB-ALL. Biochemical data show that Tp90 is distinct and physically independent from the structures already known to be involved in T cell activation, namely T11, T44 or T3/TCR. These results were confirmed by antibody-induced antigen modulation experiments. Modulation of Tp90 had no effect on the expression of T3 and of the T cell receptor. Conversely, the expression of Tp90 was not affected by modulation of the T3/TCR molecular complex by either anti-T3 or anti-TCR antibody. Functional studies showed that anti-Tp90 mAb MX20 induced high levels of interleukin 2 production in Jurkat cells. Modulation of the T3/TCR complex significantly decreased the response of Jurkat cells to stimulation by antibody MX20, suggesting that the T3/TCR complex regulates the ability of the Tp90 molecule to induce IL 2 synthesis. In addition to its effect on Jurkat cells, anti-Tp90 mAb was found to be mitogenic for peripheral blood T cells. As the magnitude of the proliferative response elicited by anti-Tp90 mAb was lower than that induced by anti-T3 mAb, the possibility was considered that only a subpopulation of T cells is reactive with anti-Tp90. Indeed as determined by FACS analyses, only 3-14% of E-rosette-positive cells were stained with mAb MX20. In addition, multicolor flow cytometry analysis showed that the Tp90+ cells belong preferentially to the CD8 subset.

Monoclonal antibodies against idiotypic determinant(s) of the T cell receptor from HPB-ALL cells induce IL2 production in Jurkat cells without apparent evidence of binding.

Two monoclonal antibodies (mAb) directed against idiotypic determinants of the T cell receptor (anti-Ti) from HPB-ALL cells induce interleukin 2 (IL2) production in Jurkat T cells without evidence of binding to these cells as judged by fluorescence-activated cell sorter (FACS) analysis, indirect antibody-binding radioimmunoassay and direct binding studies with 125I-labeled mAb. The IL2 response induced by these mAb observed both in the presence and absence of phorbol myristate acetate was in the range of that obtained when Jurkat cells were stimulated with phytohemagglutinin or anti-T3 mAb (Leu 4). The idiotypic specificity of the two anti-HPB-ALL Ti mAb was demonstrated by several criteria. Both mAb bound specifically to HPB-ALL cells as determined by radioimmunoassay or FACS analysis but not with 8 other T cell lines. The anti-HPB-ALL Ti mAb precipitated a disulfide-linked heterodimer of 85 kDa only from 125I-labeled HPB-ALL cells and not from other cell lines tested. Incubation of HPB-ALL cells with anti-T3 abrogated the expression of T3 and induced co-modulation of the idiotypic structures detected by the two anti-HPB-ALL Ti mAb. Conversely, incubation of HPB-ALL cells with either one of the anti-Ti mAb abrogated the expression of T3 and of the idiotypic structures. Our results suggest that mAb with an apparent unique specificity for the receptor of the immunizing T cell line HPB-ALL can activate Jurkat cells by a very weak cross-reaction with these cells, which is not detectable by conventional binding tests.