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Resume L'interet de transplantation de pancreas vascularise est bien etabli. Cette transplantation s'adresse essentiellement aux patients atteints de diabete de type I. Les resultats etablis a partir du registre international sur plus de 17 000 cas rapportes font etat d'environ 80 % de survie du greffon a 1 an. L'indication la plus frequente est la greffe combinee renale et pancreatique, les deux greffons provenant du meme donneur, chez les patients diabetiques de type I candidats a une transplantation renale. Neanmoins, les progres, notamment dans le domaine de l'immunosuppression, ont permis d'ameliorer les resultats des greffes de pancreas chez des patients deja transplantes renaux ou de pancreas isole chez les diabetiques sans atteinte renale. Ces deux dernieres indications, encore peu frequentes en Europe, pourraient se developper. Les progres dans le domaine de la greffe d'ilots pancreatiques ont ete importants ces dernieres annees. D'excellents resultats ont ete publies par une equipe. Ils ont motive un essai multicentrique actuellement en cours. S'ils peuvent etre reproduits, la greffe d'ilots prendra une place de plus en plus importante dans le traitement du diabete de type I, meme si les difficultes techniques ne sont pas encore toutes resolues.

In order to shed light on the mechanisms of antifilarial protective immunity, we investigated the course of experimental loaiosis after vaccination in a nonhuman primate host, Mandrillus sphinx. Six vaccinated (V) mandrills received 50 irradiated L3 while six nonvaccinated (NV) received saline solution on days -60, -30 and -15. All animals were challenged with 100 intact L3 (day 0). Parasitological and immunological status were followed for 9 months. Vaccination delayed the appearance and mean peak of microfilaraemia. Five mandrills (Mf-) were never microfilaraemic (one V mandrill) or microfilaraemic on only one occasion (2 V and 2 NV), the other seven having stable microfilaraemia (Mf+). The cytokine response of peripheral blood mononuclear cells to L3 (L3 Ag) was Th2 dominated, while microfilariae (Mf Ag) elicited a Th0-like response. During vaccination, Th2 cytokine production significantly increased in V mandrills against L3 Ag, as well as Mf Ag, whereas Th1 cytokines decreased. On day 60 postinoculation, cellular proliferation was higher in V mandrills in response to L3 and Mf Ags and PHA-L mitogen. At the end of prepatency (on day 130), mandrills with delayed appearance of microfilaraemia exhibited a high, transient IL-2 and IL-4 secretion in response to L3 Ag. Finally, high anti-Mf Th2 cytokine levels characterized Mf-mandrills not only during prepatency, but also (for IL-5) before immunization. However, the presence of a balanced Th1 anti-L3 response during prepatency in the amicrofilaraemic mandrill suggests its importance in protective immunity. Taken together, our data suggest that Th2 cells and also Th1 components of the antifilarial response, especially to larval antigen, may contribute to parasite elimination.

In a previous analysis of HIV-1-specific CTLs in lymphoid organs from HIV-seropositive patients, we reported high frequencies of in vivo differentiated CTLs directed against two immunodominant regions in the central and in the C-terminal part of the HIV-1 Nef protein. The present study analyzes the epitopes recognized by CTLs in the carboxyl terminus of Nef (amino acids 182-205). In addition to several epitopes that are recognized in association with different HLA molecules (A1, A2, A25(10), B35, B52), we defined an optimal nonapeptide (190-198). This nonapeptide was recognized by CTLs down to nanomolar concentrations in the context of at least two HLA molecules, HLA-B52 and HLA-A2, including three HLA-A2 subtypes: HLA-A2.1, -A2.2, and -A2.4. We also determined the relative frequencies of effector CTLs directed against peptide 190-198 to be as high as 10(-4), as opposed to lower frequencies ranging between 5 x 10(-5) and 5 x 10(-6) observed for the other peptides recognized in the same region, thus confirming the optimal presentation of this nonapeptide in vivo. Molecular modeling of the interactions between HLA-A2.1 and Nef peptide 190-198 suggests the formation of a stable complex and allowed us to study sequence motifs that are important for the binding of the HIV-1 peptide in the pockets of the HLA-A2.1 molecule.

Recently, authors have addressed the ability of human basophils to produce IL-4. We report here the detection of significant serum IL-4 levels in a case of acute transformation of chronic myelogenous leukemia with a predominant basophilic cell population. Leukemic basophils were isolated from patients' PBMC and assayed for their IL-4-mRNA expression and their ability to secrete this cytokine in vitro. Leukemic basophilic cells (> 90% toluidine blue positive) but not other PBMC expressed IL-4-mRNA, contained IL-4 protein, and secreted this cytokine. These cells had a spontaneous IL-4 secretion ability, without a need for an exogenous activator. Meanwhile, IL-4 release was significantly increased following leukemic cell activation through Fc epsilon RI-ligation or by Ca2+ ionophore. IL-4 and its mRNA were also detected in leukemic basophils from three other chronic myelogenous leukemia patients with moderate basophilia (13, 14, and 23% basophils in PBMC). To confirm these data in normal human cells, we have developed a method to obtain large numbers of purified basophils from human bone marrow cell cultures. In contrast to leukemic basophils, normal cells required in vitro activation through Fc epsilon RI ligation or by Ca2+ ionophore to express and secrete IL-4. Leukemic and normal basophils secreted histamine following in vitro activation, but were negative for tryptase. These data thus demonstrate the in vivo and in vitro ability of human basophils to produce IL-4.

A human CD4+ T lymphocyte clone synthesized platelet-activating factor (PAF) acether when stimulated via the CD2 pathway. PAF-acether was characterized by biochemical and biophysical properties and precursor-product relationships (alkyl-acyl-sn-glycero-3-phosphocholine (GPC)----alkyl-lyso-GPC (lyso-PAF)----PAF-acether) were demonstrated. The clone contained substantial amounts of alkyl-acyl-GPC. i) Hydrolysis of alkyl-acyl-GPC upon CD2 stimulation was evidenced: [3H]alkyl-lyso-GPC was formed from [3H]alkyl-acyl-GPC in [3H] alkyl-labeled cells; alkyl-lyso-GPC production was also bioassayed after CD2 triggering. ii) The rate of arachidonate transfer from diacyl-GPC to alkyl-acyl-GPC increased after CD2 stimulation of the [3H]arachidonate-labeled P28D T cells, demonstrating alkyl-lyso-GPC formation. iii) Comparison of the molecular species of the produced PAF-acether with those of arachidonate-containing alkyl-acyl-GPC raises the possibility that the produced PAF-acether is related to alkyl-arachidonoyl-GPC.

We previously demonstrated that long term treatment of the Ag-specific CD4+ T cell clone P28D with soluble HIV envelope glycoprotein gp120 results in a marked impairment of CD3/TCR-mediated responses. In this report, to further understand these inhibitory effects, the binding properties and internalization of gp120 have been investigated, in parallel with functional studies, in long term incubations of P28D cells with gp120. Immunofluorescence studies show that surface-bound gp120 level is maximal within 1 h of incubation at 37 degrees C and then gradually decreases. This decrease is accompanied by a progressive down-modulation of membrane CD4 (30-35% loss over a 18-h incubation period) without concomitant alteration of the CD4 mRNA steady-state level. Similar experiments performed with 125I-labeled gp120 demonstrate that the glycoprotein is progressively internalized (up to 35% internalized material after 18 h) and that it accumulates inside the cells. Confocal microscopy studies show that internalized gp120 is concentrated in localized intracellular compartments. CD4 also accumulates in compartments with a similar localization and is stained with mAb OKT4 but not with mAb OKT4a. Concomitantly to internalization of gp120 and disappearance of membrane CD4, a correlated loss of the CD4-associated tyrosine kinase p56lck is evidenced. Interestingly, a progressive impairment of the P28D responses to specific Ag or to anti-CD3 mAb is also observed. Inhibitions of T cell proliferation increase with the degree of both CD4 and p56lck down-modulation. Removal of exogenous gp120 results in a rapid and spontaneous release of internalized gp120 into a degraded form. A progressive restoration of CD4 and p56lck levels is also noticed. In parallel, CD3/TCR-mediated responses of clone P28D are fully recovered. Altogether, our results suggest that HIV-1 glycoprotein gp120 is able to down-modulate membrane CD4 presumably by a cointernalization process and to further down-modulate the associated p56lck. This dual phenomenon is presumably involved in the direct immunosuppressive effect of gp120 on the CD3/TCR-mediated activation pathway.

The effect of a battery of CD1 mAb on intracellular free Ca2+ concentration and IL-2 production has been examined on different T cell lines in this study. Both 0249F and NU-T2 two CD1b specific mAb tested, induced a rapid increase in the intracellular Ca2+ concentration on HPBALL T cells whereas only one (L161) among three different CD1c mAb (L161, 10C3, and M241) produced a similar effect. In contrast the addition of four different CD1a mAb directed against two different epitopic groups of this molecule were uneffective in modifying the intracellular Ca2+. Both L161 and 0249F also induced a comparable increase in the intracellular Ca2+ concentration on MOLT 4 and JURKAT, two other T cell lines of similar phenotype. The effect of L161 mAb on the IL-2 production of the IL-2 producing T cell line JURKAT was also examined. The association of the latter with PMA strongly induced the production of IL-2 on this cell model while either L161 or PMA alone had no effect. Although the natural ligand and the function of CD1 molecules are still unknown, the accumulation of these data strongly suggest that CD1b and CD1c might represent two activatory pathways for immature T cells operating before the classical CD2 and CD3 activation pathways.

In previous studies we demonstrated the triggering of the phospholipase C (PLC) pathway during the activation of an Ag-specific human CD4+ T lymphocyte clone by a mitogenic pair of CD2 (X11,D66) mAb. Similar conditions were applied to investigate a possible involvement of a phospholipase A2 (PLA2) acting as an additional alternative pathway during human T cell activation. Our results show that arachidonic acid or its derivatives are released after CD2 triggering. This release is largely independent of PLC activation and is mediated by a PLA2 because: 1) phosphatidylcholine is the preferential source of [3H]arachidonate release; 2) [3H]arachidonic acid release and phosphatidylcholine hydrolysis are blocked by two inhibitors of solubilized PLA2, mepacrine, and 4-p-bromophenacylbromide; and 3) we evidenced a PLA2 activity in cell homogenates. Extracellular calcium appears to play a critical role because the effects of CD2 mAb were inhibited in a Ca2(+)-depleted medium. In contrast, protein kinase C is not implicated since PMA, a protein kinase C activator, neither stimulated arachidonic acid release nor modulated CD2-induced arachidonic acid release. Cyclic AMP which has been proved to regulate the activity of the PLC in T lymphocytes does not appear to play an important role in the regulation of PLA2 activity since PGE2 has only a minimal effect on [3H]-arachidonate release. Altogether, these findings suggest that CD2 triggering stimulates a PLA2 activity in T lymphocytes via an extracellular Ca2(+)-dependent PLC protein kinase C independent mechanism.

In this study we present evidence for reactivity of pregnancy lymphocytes, but not nonpregnancy lymphocytes, with the progesterone receptor-specific monoclonal antibody mPRI. Using an avidin-biotin peroxidase detection system, we found a nuclear staining in 14.6 +/- 3.7% (mean +/- SEM, N = 27) of pregnancy lymphocytes, while only 0.47 +/- 0.33% (mean +/- SEM, N = 15) of nonpregnancy lymphocytes reacted with the antibody. To characterize the receptor-bearing subset, CD8+ and CD4+ cells were depleted by complement-dependent lysis. Depletion of CD8+ cells was accompanied by 62 +/- 18% loss of progesterone receptor-bearing cells, while depletion of CD4+ cells resulted in a twofold increase in the number of positively staining lymphocytes. In nonpregnancy lymphocytes a 3-day PHA treatment, as well as allogeneic stimulation, resulted in a significant increase in the number of receptor-containing cells. These results suggest that pregnancy, but not nonpregnancy, lymphocytes contain progesterone binding structures, and that these are inducible by mitogenic or alloantigenic stimuli.

The normopotassic solution SCOT (Macopharma, France) used for the isolation of the islets of Langerhans may improve both graft function and survival. We believe that this is due to the immunoprotective properties of polyethyleneglycol (PEG) (20 kDa; 1.5 mM/L), which is contained in this solution. However, the optimal PEG chain length remains to be determined. Three extracellular type solutions (SCOT without PEG) containing various PEG-8 kDa, 20 kDa, or 35 kDa- at 1.5 mM/L were compared in vitro for viscosity and osmolarity as well as in vivo using a murine model of pancreatic islet allotransplantation. We compared the effects of the various solutions on functional cell recovery (primary nonfunction rate, PNF) and immunoprotection (allograft survival time). We showed that the viscosity of PEG 35 kDa solutions was too high for physiological use. PEG 20 kDa solution provided the best graft function (0% PNF, P < .05). PEG 8 kda and 20 kDa solutions significantly increased allograft survival time compared to the PEG 35 kDa solution (P < .05). Graft survival was similar with PEG 20 kDa and PEG 8 kDa solutions: 27.50 +/- 3.70 days versus 23.13 +/- 4.39 days (NS). However, the number of PNF with PEG 8 kDa solution (50%) was significantly higher (P < .01) than that with the PEG 20 kDa solution (0%). These preliminary results indicated that the optimal chain length at 1.5 mM/L of PEG is 20 kDa.

There is a special interest to investigate genetic peculiarities in the populations with a low HIV seroprevalence. Despite of presence of high-risk conditions for rapid spread of HIV/AIDS epidemics in Georgia, the prevalence of this infection in the country remains very low. We studied polymorphisms of CCR5 gene in Georgians. Blood samples from 190 women randomly selected from the cohort of pregnant women involved in the program of prevention of mother-to-child HIV transmission in Georgia have been investigated. Two-step PCR was used to amplify the whole CCR5 genetic sequence. Detection of mutations and polymorphisms was done by dHPLC. All samples showing specific patterns by dHPLC, were sequenced to identify the exact nature of the mutation. It was shown that CCR5-delta32 mutation is a predominant alteration of CCR5 gene among Georgians. All subjects bearing this mutation were heterozygotes. Frequency of delta32 CCR5 allele in the population of Georgia was equal to 5%. Only one case of R223Q mutation and two cases of mutations in the non-coding region of CCR5 gene were also found. Our findings differ from the existing data showing the absence of the CCR5-delta32 mutation among Georgians and provide further support to the hypothesis on a Northeastern European origin of this mutation and North to South gradient of its distribution.

The p21Ras GTPase activating protein-associated 62-kDa protein, p62Dok1, is an early substrate of various tyrosine phosphorylation pathways. Its recent cloning in human myeloid cells and in murine pre-B cells revealed an N-terminal pleckstrin-homology domain and a tyrosine- and proline-rich C-terminal tail in its sequence. Here, we characterized a new 1261-bp cDNA identical to that of p62Dok1, but with a central 185-bp deletion (bp 456-640). This induced a frameshift leading to a premature stop codon. The deduced protein, designated p22Dok(del), corresponded to a truncated p62Dok1 isoform of 177 amino acids that can be expressed both in vitro and in vivo with an apparent molecular mass of 22 kDa. This newly identified molecule was composed of the N-terminal PH domain of p62Dok1 followed by a new 25-amino acid C-terminal sequence containing a typical class II proline-rich motif, suggesting a specific role for p22Dok(del) in signal transduction pathways.

To define the medical characteristics of intravascular drug users in Ho Chi Minh City, Vietnam, we examined 280 men, of whom 235 were infected with human immunodeficiency virus (HIV), being treated in a rehabilitation center. The patients used mainly opium, often in shooting galleries (50%). The prevalence of oral candidiasis (58%) and zoster infection (20%) was high in HIV-seropositive patients, whereas oral hairy leukoplasia and Kaposi's sarcoma were absent. The prevalence of acquired immunodeficiency syndrome was 24%. More than 80% of the patients had infections with hepatitis C virus, hepatitis B virus, cytomegalovirus, or human T cell lym- photropic virus type-1. The CD4 1 cell counts correlated well with viral load. Only HIV-1 subtype E was detected in the 30 patients tested. A cohort study of HIV-infected subjects in this population seems feasible, and would permit introduction of anti-retroviral therapy The large number of HIV-seronegative subjects sharing the same at-risk practices as the HIV-infected subjects raises the possibility of natural protection in this population.

This study attempted to determine whether the CCR5 Delta32 deletion affected progression to certain first AIDS-defining illnesses in human immunodeficiency virus type 1-infected patients enrolled in the French SEROCO/HEMOCO/SEROGEST cohorts. Toxoplasmosis onset as a first AIDS-defining illness was significantly delayed in 253 heterozygous patients, compared with 1404 wild type patients. The relative risk of toxoplasmosis associated with heterozygosity was 0. 39 (95% confidence interval, 0.16-0.96) after adjustment for age, CD4 cell count, and primary specific prophylaxis. A nonsignificant protective trend was observed with regard to the onset of mycobacterial, cytomegalovirus, and herpesvirus diseases, but these events were less frequent than toxoplasmosis. Progression to other conditions (e.g., wasting, non-Hodgkin's lymphoma, Kaposi's sarcoma) was similar in the 2 groups as was the frequency of toxoplasmosis as a subsequent AIDS-defining illness. As chemokines are involved in numerous infectious processes, the Delta32 deletion could delay progression to certain opportunistic infections such as toxoplasmosis.

To determine the influence of heterozygosity for the delta 32 mutant CCR-5 allele on HIV-1 disease progression.HIV-1 disease progression and serum viral load were analysed according to the C-C chemokine receptor (CCR)-5 genotype in 412 Caucasian patients (319 men and 93 women) with a known date of seroconversion, who were enrolled in the SEROCO cohort (median follow-up, 74 months).The frequency of heterozygosity for the mutant allele was 17% and did not differ according to sex or risk factor of HIV infection. Heterozygotes were significantly less likely than patients with two functional alleles to have symptomatic primary infection. Their serum viral load was lower during the 6- to 24-month plateau phase after seroconversion. This difference persisted afterwards, although the rate of decline in CD4+ cells was similar. Kaplan-Meier survival curves showed slower progression to clinical AIDS in heterozygotes during the first 7 years following infection (P0.02), the two curves tending to join thereafter (overall log-rank test, P = 0.17). However, the interaction term with time did not reach significance in a Cox model. The overall relative risk of progression was 0.67 (95% confidence interval, 0.38-1.18) and was not influenced by adjustment for age at seroconversion or symptomatic primary infection. After adjustment for early viral load the relative risk was 0.83. Pneumocystis carinii pneumonia and toxoplasmosis were less likely to be the first AIDS-defining illness in heterozygotes than in the other patients (0 versus 24.7% of AIDS cases, P = 0.04), despite similar management.Deletion of one CCR-5 gene allele appears to protect against HIV-1 disease progression, mainly during the early years of the infection. Heterozygosity for the deletion leads to persistently lower viral load, and also seems to protect against some opportunistic infections.

Apoptosis induced in different cell lines can be detected and quantified in one step. Direct labeling of apoptotic cells (DLAC) was performed by incorporation of fluorescein-dUTP (F-dUTP) in DNA strand breaks by terminal deoxynucleotidyl transferase (TdT). Nick-end-labeling using F-dUTP obviates the need for second-step revelation reagents without reducing the specificity of detection. Cells were analyzed by microscopy or flow cytometry for objective quantification of apoptotic cells in controlled dilution samples. Furthermore, we demonstrate that DLAC is compatible with surface labeling by monoclonal antibodies, allowing dual-color analysis. The data presented here illustrate the simplicity and potential of this method, which allows the preservation of fragile cells and the possibility of combining apoptosis detection with immunostaining.

The thymus is the primary lymphoid organ for differentiation of CD4+ and CD8+ lymphoid T celles. Bone marrow T cell precursors colonize the cortex where they differentiate into immature CD1+CD4+CD8+ and CD3+ T celles representing 80% of thymocytes, then migrate to the medulla where they loss CD1 and either CD4 or CD8 expression, and definitively leave the thymus as mature CD3+CD4+ or CD3+CD8+ lymphocytes for peripheral compartment.