Biomedical subjects
Publications and source records attributed to J Choppin.
T-cell receptors recognize foreign proteins as peptide fragments associated with major histocompatibility complex (MHC) molecules. Properties of antigenic peptides, methods for detecting peptide-MHC molecule combinations, and the characteristics of the interaction are reviewed. Possible explanations for graft rejection and autoimmune disease are suggested in the light of these data.
To identify HIV peptides containing HLA class I binding regions, different studies have been performed. These include the detection of interactions between HIV peptides and purified HLA molecules in solid-phase assays, the measurement of HLA molecule assembly in the presence of peptide added to cell lysates, and the detection of inhibition of CTL-mediated cytolysis by competition between peptides on target cells. To date, the HIV epitopes recognized by anti-HIV CTL are from the Env, Gag, Nef, and Pol proteins and they are identified using synthetic peptides of 12 to 20 amino acids. The search for a correlation between known HIV CTL epitopes and the results of HLA/peptide interaction assays reveals that: (1) most of the peptides that are positive in the assembly assay contain a HLA-A2 peptide motif but the correlation between these positive peptides and the CTL epitopes is not obvious; (2) a high proportion of HIV epitopes are included in the peptides positive in solid-phase binding and in inhibition of cytolysis assays, although these tests do not allow us to predict HLA restriction; (3) the HLA-A2 peptide motif is not systematically included in HLA-A2-restricted CTL epitopes, this observation raising the possibility that other sequences are involved in HLA binding.
The physical association of HLA class I or H-2 molecules with 36 HIV-1 Nef synthetic peptides was studied using a direct peptide binding assay (PBA) in solid phase. To assess the functional significance of the PBA results, the Nef peptides were also tested for their ability to inhibit the lytic activity of human or murine CTL. The PBA results showed that seven partly overlapping regions of the Nef protein contained MHC binding peptides (4-18, 46-67, 73-94, 100-128, 126-155, 182-198, and 192-206). Five of these seven regions included all the already described epitopes recognized by CD8+ human CTL. The two other regions, 4-18 and 46-67, are not yet described as antigenic for human CD8+ cells but they are located in the N-terminal part of Nef that was previously described as being stimulator for rat or chimpanzee CD4+ cells. Altogether, it can be concluded that 1) In virtually 100% of the cases, the PBA is capable to detect known antigenic peptides recognized by CTL. 2) The PBA and the functional inhibition assay provide similar results, supporting the functional significance of PBA results. 3) The PBA is easy to handle on a large scale, using multiple peptide and several MHC molecules, so that it can be used as a routine method for prevision of possibly epitopic sequences. 4) Systematic studies of peptides issued from the whole sequence of a given protein allow to map polyepitopic areas that are probably the most interesting parts of proteins for a vaccine purpose.
To detect HLA-binding peptides in 10 HIV-1 proteins (Rev, Tat, Vif, Vpr, Vpu, Gag p18, Gag p24, Gag p15, Env gp120 and Env gp41), the peptide binding assay (PBA) has been performed using three HLA class I molecules. Correlations have been searched between the PBA results and the peptide competitor activity in a functional test using HLA-A2-restricted CTL and target cells. A correlation between the data found in the PBA and well-defined CTL epitopes could be attempted only for the three Gag proteins. For these proteins, our results are in agreement with the known existence of epitopes reacting with human CD8+ CTL, with some exceptions. Together with the results reported with a panel of Nef peptides, these experiments showed that at least 18/20 of the already reported CTL epitopes from HIV-1 Gag, Nef, and Env proteins could be detected by the PBA, most (17/18) corresponding to strong reactivities. Perhaps more important, the regions of HIV-1 Gag p24 or Nef proteins that contain multiple associated CTL epitopes, with different HLA restrictions, were clearly identified by the reactivities in the PBA of several overlapping peptides and the major practical interest of the PBA might be the detection of such polyepitopic regions. Prediction are proposed in this report for 10 proteins, including several proteins for which CTL epitopes remain presently unknown.
Single-chain mouse Kd molecules (SC-Kd) were engineered by connecting residue 276 of Kd heavy chain to the first residue of beta 2-microglobulin through spacers of various lengths, and expressed intracellularly in monkey COS-1 cells. Labeled SC-Kd molecules were found to react with several monoclonal antibodies which recognize native Kd molecules. SC-Kd-15 (with a spacer of 15 residues) was studied in more details. It could be purified and shown to regain a native-like structure after treatment with denaturing agents. Purified SC-Kd-15 could bind certain peptides in a manner qualitatively similar to the Kd.
The physical association of 40 antigenic peptides and purified HLA class I and class II molecules was monitored using a direct peptide binding assay (PBA) in solid phase and an inhibition peptide binding assay (IPBA) in which the competing peptide was present in a soluble phase. We also examined the ability of different peptides to inhibit the lytic activity of human antiviral cytolytic T cells towards cells incubated with the corresponding target peptide. Our results showed that: (a) Binding of a given human T cell-recognized peptide to several HLA class I and class II molecules occurred frequently. Nevertheless, preferential binding of peptides to their respective restriction molecules was also observed. (b) Binding of HLA molecules to peptides recognized by murine T cells occurred less frequently. (c) 11 of 24 (46%) randomly selected HIV-1 peptides contained agretopic residues allowing their binding to HLA molecules. (d) The kinetics of HLA/peptide association depended on the peptide tested and were faster than or similar to those reported for Ia molecules. Dissociation of these complexes was very low. (e) Peptide/HLA molecule binding was dependent on length, number of positive charges, and presence of hydrophobic residue in the peptide. (f) A correlation was demonstrated between a peptide inhibitory effect in the IPBA and its blocking effect in the cytolytic test. Our data indicated that the restriction phenomenon observed in T cell responses was not strictly related to either an elective HLA/peptide association, or a high binding capacity of a peptide to HLA molecules. These data also showed that the PBA and IPBA are appropriate for the detection of agretopic residues within HIV-1 proteins.
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Histochemical and histoenzymological studies were carried out on liver slices from mice which had received alpha-amanitine two days previously, some of which had been treated with silybin, and from control mice. The toxin produced certain changes in the activity of the enzymes involved in different metabolic processes, and in the amounts of lipids and nucleic acids. Treatment with silybin, given 60 min before administering alpha-amanitine or 10 min later alike, prevents the appearance of these changes and gives results comparable to those in the control animals.
The human peripheral blood lymphocytes are characterized by different surface markers: The B lymphocytes by the EAC rosettes and surface immunoglobulin and the T lymphocytes by the E-rosettes. Each lymphoid population has been studied for cytochemical parameters (Acid phosphatase, beta-glucuronidase) by light microscopy and for ultrastructural characteristics by micromanipulation and immunoelectronmicroscopy. It has been shown that the studied enzymes are markers for T cells with high affinity for sheep red blood cells and that the separation of lymphocytes by the rosette techniques leads to homogenous cell populations at the ultrastructural level. A classification of peripheral lymphocytes according to their immunocytochemical profile is discussed.
The administration of amino-3 beta hydroxy-20 beta pregnene-5, to the male Wistar rat, per os, at doses of 100 and 200 mg/kg/24 h, induce the development of a chronic active hepatitis. The ultrastructural observation shows slight changes only in perilobular hepatocytes at the beginning of treatment; then hepatocellular alterations progressively increase and may be observed in the whole lobule after 40 and 80 days of treatment; the progression of hepatocellular damage is associated with collagen increase and bile duct proliferation. The interest of this experimental hepatitis, as a model analogous to human chronic active hepatitis, is discussed.
Histochemical and histoenzymological studies carried out on liver slices of the mouse show that silybin both as prophylactic and curative treatment, inhibits the disorders of metabolic activity brought about by phallodine and maintains enzymatic activities at levels comparable with those of the control animals. Silybin, when administered alone, produces only minor changes in activities. As with silymarin, the only notable effect is an increased activity of alkaline phosphatase. The mode of action of these two substances appears to be the same.
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The growth of L 1210 leukemia in DBA/2 strain mouse, provokes in the grafted animals intraleucocytic enzymatic modifications. As increase of acid phosphatase and a decrease of beta-glucuronidase were observed in the lymphocytes, the level of polymorphonuclear non-specific esterase being decreased. The implications of these modifications in the host response toward the tumor is discussed.
Holaphyllaminol, or Amino-3 beta-Hydroxy-20-beta Pregnene-5, was administered by gastric tube to male Wistar rats at dose of 50, 100 and 200 mg/kg/24 hr respectively, during 10, 20, 40 and 80 days. This substance provokes hepatic lesions beginning at the periportal region, the characteristics of which are a cholestasis, a canalar proliferation and a fibrosis. The interest of these alterations lies in the fact that they present the same features as certain human chronic hepatitis; their progression, a function of the dose of the toxin and the duration of treatment, may make possible to realisation of a model of experimental chronic hepatitis.
The hepatoprotective action of silymarin, the active principle extracted from the fruit of Silybum marianum (L.) Gaertn., in animals (dogs, rabbits, rats, mice) intoxicated with phalloidine is evident, both after protective and curative treatment. A dose of 15 mg/kg of silymarin protects every animal when given 60 min before the toxin. When injected 10 mim after phalloidine, a dose of 100 mg/kg of silymarin again provides total protection. However, as the time span between administration of the toxic substance and start of treatment increases, so the efficacy of silymarin decreases; after 30 min its curative effect is negligible. The histochemical and histoenzymological studies show that during intoxication of the mice by phalloidine, silymarin inhibits the effect of the toxic substance and regulates the functions of the hepatocyte, when given either 60 min before or 10 min after phalloidine.