Recognition of a novel naturally processed, A2 restricted, HCV-NS4 epitope triggers IFN-gamma release in absence of detectable cytopathicity

Using short term CTL lines derived from HLA A2/K^b transgenic mice and IFN-gamma release assays we demonstrate that the NS4.1769 epitope, is generated from natural processing of the NS4 antigen, and presented in the context of the A2/K^b molecules. Interestingly, T cell recognition of the naturally processed form of the NS4.1769 epitope was associated with significant IFN-gamma release, but no direct cytolytic activity. Epitopes of this phenotype might be of interest, in terms of therapy of chronic HCV infection by associating the benefit of localized lymphokine release with low or absent direct cytopathicity.     

Analysis of the role of HLA-G in preeclampsia

Preeclampsia (PE) is a multisystem disorder of human pregnancy, occurring in 5%-10% of all population births and represents the leading cause of both fetal and maternal morbidity and mortality in pregnancy. Although the disorder only becomes clinically apparent late in pregnancy, the underlying pathology indicates that invasion of fetal trophoblasts into maternal spiral arteries during early pregnancy is shallow or absent in PE. A large number of epidemiologic studies have been carried out and they demonstrate that the disorder is highly heritable and occurs with a high incidence in all populations. Studies have shown that PE is largely under genetic control, but the mode of its inheritance remains unclear. Genetic studies have been carried out using both large scale linkage analysis and candidate gene approaches; however, the genetic mechanisms underlying the disorder have yet to be determined. We focus on the potential role of HLA-G, a nonclassical class I HLA located on chromosome 6, which appears to be a key component in trophoblast invasion. We examine the hypothesis that HLA-G may have a key role in both genetic susceptibility to, and pathogenesis of, PE.     

The expansion and selection of T cell receptor αβ intestinal intraepithelial T cell clones

In conventional mice, the T cell receptor (TCR)αβ⁺ CD8αα⁺ and CD8αβ⁺ subsets of the intestinal intraepithelial lymphocytes (IEL) constitute two subpopulations. Each comprise a few hundred clones expressing apparently random receptor repertoires which are different in individual genetically identical mice (Regnault, A., Cumano, A., Vassalli, P., Guy-Grand, D. and Kourilsky, P., J. Exp. Med. 1994. 180: 1345). We analyzed the repertoire diversity of sorted CD8αα and CD8αβ⁺ IEL populations from the small intestine of individual germ-free mice that contain ten times less TCRαβ⁺ T cells than conventional mice. The TCRβ repertoire of the CD8αα and the CD8αβ IEL populations of germ-free adult mice shows the same degree of oligoclonality as that of conventional mice. These results show that the intestinal microflora is not responsible for the repertoire oligoclonality of TCRαβ⁺ IEL. The presence of the microflora leads to an expansion of clones which arise independently of bacteria. To evaluate the degree of expansion of IEL clones in conventional mice, we went on to measure their clone sizes in vivo by quantitative PCR in the total and in adjacent sections of the small intestine of adult animals. We found that both the CD8αα and the CD8αβ TCRαβ IEL clones have a heterogeneous size pattern, with clones containing from 3 × 10³ cells up to 1.2 × 10⁶ cells, the clones being qualitatively and quantitatively different in individual mice. Cells from a given IEL clone are not evenly distributed throughout the length of the small intestine. The observation that the TCRαβ IEL populations comprise a few hundred clones of very heterogeneous size and distribution suggests that they arise from a limited number of precursors, which may be slowly but continuously renewed, and undergo extensive clonal expansion in the epithelium.     

Biology and functions of human leukocyte antigen-G in health and sickness

In 1998, the first International Conference on human leukocyte antigen-G (HLA-G) was held in Paris. At that time, HLA-G was still a new HLA class I molecule, few aspects of its immunological functions were known, and its expression by tumors was just being described. In 1998, tools to properly study HLA-G were lacking, especially monoclonal antibodies, and three conclusions were drawn after the congress: (i) animal models were needed, (ii) the biology of HLA-G isoforms had to be confirmed, and (iii) HLA-G expression by tumors required clarification. Five years later, these three issues have been addressed. HLA-G is now gaining pace and is investigated for its immuno-inhibitory functions in the context of multiple pathologies. Eighty five oral presentations were given this year for more than 200 investigators working on HLA-G by speakers from over 20 countries. The success of the 3rd International Conference on HLA-G reflects the interest and tremendous work of the many research teams which, over the years, contributed to the publication of more than 500 peer-review articles. We summarize the key points that were presented and discussed during this meeting.     

An immunochemical approach to investigating the mechanism of inhibition of cysteine proteinases by members of the cystatin superfamily.

Antibodies were raised against a synthetic dodecameric peptide KGAGQVVAGPWK (K12K), encompassing sequences thought to be important for the function of the cysteine proteinase inhibitors of the cystatin superfamily. These antibodies specifically recognized molecules of family 3, i.e., kininogens, in the serum of seven mammalian species tested in this study. The only notable exception was that of rat thiostatin (T kininogen) which is structurally related to the kininogen family. The antibodies also discriminated between family 2 (cystatins) and family 3 (kininogens) of the cystatin superfamily, since neither chicken cystatin nor human and rat cystatins C and S, which all belong to family 2 were recognized. The cystatin-like inhibitory domains resulting from fragmentation of human low molecular weight kininogen by bovine trypsin, were still recognized by antibodies, indicating that discrimination does not require two neighbouring inhibitory sites on the kininogen heavy chain. The antibodies blocked the capacity of kininogens to inhibit papain, suggesting that they recognize a conformational epitope at or near the kininogen inhibitory sites. The inhibitory properties of family 2 cystatins remained unchanged, confirming that members of this family do not interact with anti K12K antibodies. These antibodies are thus a new tool able to discriminate functionally and structurally between the members of the cystatin superfamily.     

Ligand-induced autoregulation of IL-2 receptor {alpha} chain expression in murine T cell lines

The IL-2 receptor (IL-2R) is composed of three chains a, ßand . In mice, contrary to the human system, we have previouslydemonstrated that the IL-2Rß complex does not bindIL-2. Therefore, mouse IL-2 response is completely dependenton the expression of the IL-2R gene product. T cell clones expressingmouse IL-2Rß and the human IL-2R transgene have beenstudied. When cells are grown in IL-4, mouse IL-2R is not expressed.However, exposure to IL-2 leads to the expression of the endogenousmurine IL-2R subunit. The T cell line expressing mouse IL-2Rand human IL-2Rß can grow in IL-2 but does not expressendogenous murine IL-2 R. Transfection of these cells with thehuman IL-2R gene restores the capacity to induce murine IL-2R.This result demonstrates that IL-2-IL-2R interactions are requiredfor induction of IL-2R. The kinetics of induction and deinductionof murine IL-2R have been studied using clone 18.III. From negativecells, expression of murine IL-2R is a very slow phenomenon.From cells fully expressing IL-2R, deinduction is a two-stepprocess: after a rapid decrease of IL-2R the cells continueto express, for a long period of time, basal levels of murineIL-2R. When cells expressing basal levels of IL-2R are exposedto IL-2, induction of IL-2R is a very rapid phenomenon. Theautoregulatory loop formed by IL-2-IL-2R therefore displaysdifferent levels of functioning.     

Characterization of three monoclonal antibodies specifically directed against the ligand binding site area of the p55 subunit of the mouse interleukin-2 receptor

Three new rat monoclonal antibodies (MAbs) (5A2, 125A8 and 135D5) directed against the mouse interleukin-2 receptor (IL-2R) were isolated. They were obtained after immunization of LOU rats with 14.1.6 T helper cell clones. These three MAbs recognize the p55 subunit of the IL-2R and compete with the binding of previously characterized MAbs AMT13 and 3C7 specific for this p55 subunit [Moreau et al. (1987) Eur. J. Immun. 15, 723-727]. They recognize the same (or closely related epitopes) since they reciprocally compete with each other's binding. Scatchard plot analysis of the data from inhibition experiments clearly indicate that they recognize with very high affinity the ligand binding site area of the p55 subunit of the IL-2R. The properties of the Fab fragment prepared from 5A2 and 135D5 indicate that at saturation one intact IgG molecule binds two IL-2R molecules.     

Treatment of R-3327 prostate tumors with a somatostatin analogue (somatuline) as adjuvant therapy following surgical castration

This study addresses, in an animal tumor model, the clinical problem of "escape from castration inhibition." Somatuline (BIM-23014C), an octapeptide analogue of somatostatin with enhanced potency and longer duration of biological activity was administered as a therapeutic agent, over a period of 90 and 197 days, to male Copenhagen rats bearing syngeneic Dunning R-3327-H prostate tumors. Androgen sensitivity was confirmed by the response of tumors to castration and by the significant inhibition of tumor growth in intact animals by treatment with a luteinizing hormone-releasing hormone antagonist (BIM-21009). Inhibition of tumor growth resulting from castration persisted for 102 days, after which progressive regrowth occurred, indicating an escape from castration inhibition. When Somatuline treatment was initiated as an adjuvant therapy 5 days after castration, the rate of tumor regrowth during escape was significantly retarded. During the period of 197 days postcastration, tumors in the vehicle-treated, intact controls grew to an average diameter of 38.6 +/- 7.6 mm and tumors in vehicle-treated castrate controls grew to an average diameter of 23.3 +/- 4.1 mm (60% test/control). Treatment with the luteinizing hormone-releasing hormone antagonist induced no significant additional tumor inhibitory effects in castrated animals which developed tumors having an average diameter of 30.2 +/- 8.2 mm (78% test/control). Treatment of tumors in castrate animals with Somatuline, on the other hand, induced a significant (P less than 0.01) tumor-inhibitory effect that was greater than that produced by castration alone, developing an average tumor diameter of only 14.3 +/- 2.6 mm, (37% test/control). A growth inhibitory effect was also inducible in animals having tumors that had already escaped castration inhibition. The relative nontoxicity of a somatostatin analogue such as Somatuline suggests that chronic or maintenance therapy of slow-growing prostate cancers may be both feasible and acceptable in a clinical setting.