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Biomedical subjects

P Vassalli

Publications and source records attributed to P Vassalli.

At least 109 records · Page 6Linked to original sources
PubMed1986

L3T4+ T lymphocytes play a major role in the pathogenesis of murine cerebral malaria.

The pathogenic importance of L3T4+ T cells in the development of murine cerebral malaria was demonstrated by the following observations. First, in vivo administration of an anti-L3T4 monoclonal antibody protected Plasmodium berghei-infected CBA mice from the development of neurologic symptoms and acute death. In contrast, injection with an MAb directed against the Ly.2+ T cell subset had no protective effect. Second, thymectomized, irradiated, and bone marrow reconstituted (ATxBM) CBA mice did not develop acute cerebral malaria when infected by P. berghei, although parasitemia and anemia rose to the same extent as in normal P. berghei-infected CBA mice. The occurrence of lethal neurologic perturbations could be restored in ATxBM mice selectively reconstituted with L3T4+ Ly.2-lymphocytes but not with Ly.2+ L3T4- cells. Third, adoptive transfer of L3T4+ cells from mice dying of cerebral malaria into euthymic mice subsequently infected by P. berghei led to an acceleration of the disease. These results confirm that cerebral malaria in mice is the expression of immunopathologic reactions and outline the particular pathogenic importance of L3T4+ T cells.

Animals↗
PubMed1986

Administration of recombinant interleukin 2 to mice enhances production of hemopoietic and natural killer cells.

Seven days of continuous perfusion of mice with human recombinant interleukin 2 (rIL 2) (approximately 3 X 10(4) U/day) increased the percentage of large mononuclear leukocytes (LML) among bone marrow, spleen, lymph node cells and liver interstitial cells (LIC). An increase in the lymphokine-activated killer (LAK) activity was evident in these organs. The greatest increase in the number of LML and in the LAK activity was observed among the liver interstitial cells (about 500-fold increase). The LML were nonphagocytic, Thy-1+, sIg-, Ly 2+, L3T4- and asialo Gm1+. Perfusion of athymic nude mice, or of thymectomized, irradiated radiation chimera, showed that the Thy-1+, LAK+ LML were the thymus and T lymphocyte-independent progeny of Thy-1- marrow precursors. The LML had no T cell function in a graft-vs.-host reactivity assay, neither did they have an inhibitory effect on T lymphocyte function in vivo. rIL 2 perfusion did not significantly affect the medullary hemopoiesis but did strongly enhance the extramedullary hemopoiesis, particularly within the interstice of the liver: the number of erythroid and myeloid cell was increased as well as the number of colony-forming units per spleen and colony-forming units per culture for various lineages (20-50-fold increment). These results show that in vivo, rIL 2 has a global enhancing effect on hemopoiesis together with a more selective influence on the production of LML.

Animals↗
PubMed1986

Stimulation of hematopoiesis in vivo by recombinant bacterial murine interleukin 3.

Mouse interleukin 3 (IL-3) cDNA was cloned into a plasmid construction, allowing the synthesis of very high quantities of IL-3 in Escherichia coli. The recombinant (r) IL-3, purified to homogeneity, was active in vitro on the proliferation and differentiation of various hematopoietic progenitor cells at 1 pM. To maintain detectable blood levels of IL-3, osmotic pumps containing rIL-3 or control solutions were placed under the skin of normal and irradiated C3H/HeJ and (BALB X B10) F1 mice. The effect of IL-3 on hematopoietic progenitor cell numbers in spleen and bone marrow was evaluated 3 and 7 days later by using an in vitro clonal assay. The results demonstrated the following: (i) Doses of IL-3 infused at the rate of 2.5-5 ng per g of body weight per hr were sufficient to increase the numbers of hematopoietic progenitors in normal mice by at least 2-fold within 3 days. (ii) In mice with progenitor cell levels depressed by sublethal irradiation, 7-day treatment with IL-3 resulted in a 10-fold increase to near normal levels. (iii) The erythroid and myeloid lineages appeared to be enhanced to the same extent. (iv) Enhancement of hematopoiesis occurred primarily in spleen, but hematopoietic foci were also evident in the liver; in contrast, total cell and progenitor cell numbers were decreased in the bone marrow.

Animals↗
PubMed1986

Gut injury in mouse graft-versus-host reaction. Study of its occurrence and mechanisms.

The occurrence, nature, and pathogenesis of intestinal lesions were studied in a number of graft vs. host reaction (GVHR) conditions in mice, combining variations in the nature of the following: the F1 hosts (newborn or adult, normal or lethally irradiated), the injected parental T cells (mixed or selected subsets of Lyt2+ or L3T4+ cells), and the antigenic stimulus (semi-allogeneic or restricted to class I or II MHC loci). The following conclusions were drawn: Three gut alterations are always associated: donor T cell infiltration, predominating in the crypt region; acceleration of the epithelium renewal; and increased epithelial Ia expression. The initial event is T-cell infiltration, which results from stimulation within the Peyer patches followed by cyclic traffic, i.e., migration into the thoracic duct and then seeding to the whole gut mucosa. Both Lyt2+ and L3T4+ cells can infiltrate the gut wall, the extent of the infiltration by a given subset depending upon the capacity of the donor blasts to circulate in the thoracic duct (higher for L3T4+) and then to home in the gut (much higher for Lyt2+ blasts) and the nature of the alloantigenic stimulation that governs the extent of each donor subset proliferation. Both donor T-cell subsets can induce gut epithelial damage, but for a comparable amount of infiltrating cells, L3T4+ cells induce more lesions. When the antigenic stimulation is restricted to class I or class I MHC loci, gut GVHR is much more easily elicited across class II MHC differences, which stimulate preferentially L3T4+ donor cells. The main mechanism of epithelial damage is not direct cytotoxicity, but more probably lymphokine(s) release.

Animals↗
PubMed1985

Bone marrow pre-B lymphocytes synthesize immunoglobulin mu chains of membrane type with different properties and intracellular pathways.

Mouse normal bone marrow pre-B lymphocytes synthesize only membrane mu chains (micron), as shown by mRNA studies and peptide analysis. The micron chains exist in two forms: free micron chains assembled into dimers, or L chain-bound micron chains present in IgM monomers (in the case of 'late pre-B cells', i.e., after productive L chain gene rearrangement). These two forms of molecules are very different in properties, fate and intracellular pathways. Free but not L chain-bound mu chains are highly susceptible to mild proteolysis, which degrades their entire Cmu 1 and VH domains. Free mu chains are rapidly degraded within the lysosomal compartment, which they reach via the cis, avoiding the trans, part of the Golgi complex. In contrast, as soon as mu chains bind to L chains, they are directed towards the 'trans' Golgi compartment, where they undergo terminal glycosylation, then to the cell surface, where they progressively accumulate. It is suggested that the conformation instability of the Cmu 1 and VH domains of the free mu chains plays a critical role in the intracellular targeting of these molecules, as compared with that of L chain-bound mu chains.

Animals↗
PubMed1985

Human endothelial cell cultures: phenotypic modulation by leukocyte interleukins.

We report here that soluble factors from activated mononuclear leukocytes have a dramatic effect on cultured endothelial cells. While human umbilical vein endothelial cells grown under standard conditions show a polygonal, epithelial-like morphology, cells exposed to culture media conditioned by lectin-activated human mononuclear leukocytes become extremely elongated and/or send out numerous cytoplasmic processes, assuming a dendritic configuration. This effect cannot be mimicked by exogenous cyclic AMP, is reversible upon interruption of the treatment, and appears specific for endothelial cells, since it has not been observed so far with other cell types. The shape changes are accompanied by a reorganization of the endothelial cell cytoskeleton: actin microfilament bundles tend to be disposed in parallel arrays, while intermediate filaments and microtubules penetrate up to the extremity of the cytoplasmic processes. Colchicine prevents endothelial cell elongation but only slightly impairs the formation of lateral cell processes ("dendritic configuration"). Purified interleukins were tested for their ability to induce these changes of cell shape. Escherichia coli-recombinant human interleukin 2 had no effect, and gamma-interferon only a slight effect on endothelial cell morphology. Interleukin 1 induced moderate cell elongation, while combined treatment with both interleukin 1 and gamma-interferon resulted in shape changes indistinguishable from those elicited by supernatants of activated mononuclear leukocytes. The possible relevance of the observed endothelial cell changes to the reported angiogenic activity of mononuclear cell products is discussed.

Cells, Cultured↗
PubMed1985

Functional analysis of T cell subsets from mice bearing the lpr gene.

The autosomal recessive lpr (lymphoproliferation) gene is responsible for a thymus-dependent massive lymphoproliferation associated with the development of lupus-like autoimmune disease. Phenotypic analysis of adult lpr/lpr lymph nodes has demonstrated accumulation of a dull Lyt-1+, Thy-1+ population that expresses neither Lyt-2 nor L3T4 antigens. With the use of a depletion method based on complement-mediated lysis with an anti-Lyt-2 monoclonal antibody (31 M) and a new anti-L3T4 monoclonal antibody (RL 172.4), we have purified the Lyt-2- L3T4- subset from lymph nodes or spleens of C57BL/6-lpr/lpr mice and determined whether they are immunologically functional in vitro. Production of neither interleukin 2 nor interferon-gamma was detected by the double-negative subset after stimulation with concanavalin A and/or phorbol myristate acetate. The frequencies of allospecific cytotoxic T lymphocyte (CTL) precursors and lectin-induced antigen-nonspecific CTL precursors were diminished to almost undetectable levels, whereas the Lyt-2+ population from lpr/lpr mice had CTL-precursor frequencies comparable with that of +/+ mice. These results show that the major cell subset of adult lpr/lpr lymph nodes or spleens is composed of lymphocytes with markedly limited potential for lymphokine production or antigenic stimulation.

Animals↗
PubMed1984

Gut mucosal mast cells. Origin, traffic, and differentiation.

Gut mucosal mast cells (MMC), which are nearly absent in normal mice are abundant during nematode infection. In normal mice, study of MMC precursors (MMC-P: cells giving rise to MMC colonies in the presence of IL-3) show that: (a) their frequency, judged by limiting dilution is very high in bone marrow (BM) and gut, and very low in most lymphoid organs and thoracic duct lymph (TDL); (b) gut MMC-P are Thy-1- Lyt-1-2- and are not rapidly replicating; (c) they are the progeny of less differentiated BM MMC-P which are attracted from the blood to the gut mucosa by local factor(s), other than antigen and T cell factors (since normal amounts of gut MMC-P are found in germ-free, nude, and newborn mice). In mice bearing the Wehi 3 tumor (which releases enough IL-3 to produce detectable blood levels) spleen and mesenteric lymph nodes (LN) show increased MMC-P frequency, the greatest increase being in the gut and BM, where numerous differentiated MMC are found. In Nippostrongylus brasiliensis (Nb)-infested mice (known to develop a large, T cell-dependent, gut MMC infiltration), gut MMC-P proliferation is induced by IL-3 released from gut mucosal Thy-1+ Lyt-2- cells, whose in vitro IL-3 release capability is much higher than that of similar cells from normal mice. Both Nb-stimulated T blasts and proliferating MMC-P undergo cyclic traffic, migrating into the TDL and then seeding the whole length of the gut (a process which allows a widespread immune defense after a local antigenic stimulus). Experiments using 2-d interruption of this traffic and fetal gut grafts, suggest that the continuous homing of T blasts back to the gut which leads to permanent Nb-stimulated IL-3 release, is essential for the full maturation of MMC. Transfer experiments in the rat show that TDL circulating MMC-P rapidly mature into MMC when they home back to the Nb-infested gut. It is proposed that gut MMC arise after several stages of progressive differentiation of MMC-P, influenced both by IL-3 and unidentified gut factor(s).

Animals↗
PubMed1984

Leukocyte interleukins induce cultured endothelial cells to produce a highly organized, glycosaminoglycan-rich pericellular matrix.

We report here that interleukins have a dramatic effect on extracellular matrix production by cultured endothelial cells. Human umbilical vein endothelial cells incubated with growth media conditioned by lectin-activated human peripheral blood mononuclear leukocytes undergo marked changes in cell shape and elaborate a highly organized extracellular material that is not detectable in untreated cultures. This material has the following characteristics: (a) it is not recognizable by electron microscopy unless the cationic dye, Alcian blue, is added to the fixative; (b) it is visualized as a network of branching and anastomosing fibrils of various thickness that can be resolved into bundles of fine filaments; (c) it is associated with the cell surface, extends between contiguous cells, and coats the culture substrate; (d) it is removed by digestion with glycosaminoglycan-degrading enzymes, such as crude heparinase and chondroitinase ABC. These results demonstrate that soluble factors released by activated peripheral blood mononuclear leukocytes (interleukins) stimulate cultured human umbilical vein endothelial cells to produce a highly structured pericellular matrix containing glycosaminoglycans (probably chondroitin sulfate and/or hyaluronic acid) as a major constituent. We speculate that this phenomenon corresponds to an early step of angiogenesis as observed in vivo as a consequence of interleukin release.

Alcian Blue↗
PubMed1984

Characterization of cytoplasmically oriented Golgi proteins with a monoclonal antibody.

BALB/c mice were repeatedly immunized with a galactosyl transferase-rich microsomal fraction of rat myeloma cells. Spleen cells were subsequently fused with Sp2/0 mouse myeloma cells, the resulting hybridomas were cloned, and their secreted Ig was screened for reactivity with antigens belonging to the Golgi complex. One such monoclonal antibody, 6F4C5, gave especially intense immunofluorescent staining of the Golgi area of myeloma cells and fibroblasts. It recognized two proteins bands on immunoblots of gel-fractionated cell lysates: a major one with an estimated Mr of 54,000 and a minor one at 86,000. Both proteins were concentrated in microsomal fractions isolated at low ionic strength. They were hydrophilic judging from partitioning of a Triton X-114 cell lysate. Both were cytoplasmically oriented as demonstrated by protease and high KCl treatments of postmitochondrial supernatants and microsomal fractions. Neither was retained by columns of insolubilized wheat germ agglutinin or concanavalin A, which suggests that they are not glycoproteins. Their more detailed location in the Golgi complex was studied by immunoelectron microscopy, using a saponin permeabilization procedure and peroxidase-conjugated reagents. The observed staining was restricted to two or three cisternae in the medial part of the stack. Nevertheless, differential centrifugation experiments indicated that the two antigens may be recovered in distinct subcellular fractions: this may be related to the unexpected observation that rather low salt concentrations strip the antigens from microsomal fraction.

Animals↗
PubMed1984

Production of interleukin 2, interleukin 3, and interferon by mouse T lymphocyte clones of Lyt-2+ and -2- phenotype.

T lymphocyte clones were derived by stimulation of positively selected Lyt-2+ and Lyt-2- lymphocytes with Con A in an Interleukin 2 (IL 2)-enriched medium under conditions of limiting dilution. Forty clones were expanded and tested, after activation by Con A, for the production of IL 2, IL 3, and interferon (IFN). Thirteen Lyt-2- clones were all co-producers of IL 2 and IL 3, and 10/13 were also producers of IFN. Twenty-seven Lyt-2+ clones were much more heterogeneous, 13 being IL 2 and IL 3 nonproducers, whereas 14 produced variable and poorly correlated amounts of IL 2 and IL 3. Three Lyt-2+ clones were observed to produce IL 2 or IL 3 alone. The majority of the Lyt-2- (10/13) and of the Lyt 2+ (21/27) clones were also producers of IFN. Exogenous IL 2 added during the activation of the Lyt-2+ by Con A did not enhance IL 3 production, whereas it did enhance IFN production by some but not all Lyt-2+ clones. Thus, among the T lymphocytes of the Lyt-2+ and -2- phenotypes there are cells capable of releasing IL 2, IL 3, and IFN. This supports the concept that these phenotypes are associated with antigen recognition rather than with cell functions, but it is apparent that the functional capacities of individual T cells, if accurately represented by their clonal progeny, are far from uniform.

Animals↗
PubMed1983

Specialization of the macrophage plasma membrane at sites of interaction with opsonized erythrocytes.

We incubated mouse peritoneal macrophages for 3-8 min at 37 degrees C with antibody-coated sheep erythrocytes and examined regions of close interaction between the two cell types by electron microscopy. At sites of focal macrophage-erythrocyte contact we observed a distinctive specialization of the macrophage plasma membrane consisting of a prominent subplasmalemmal band of electron-dense material, approximately 25-35 nm in thickness. In many instances, this band showed a periodic substructure similar to that seen in clathrin coats. Moreover, many slender erythrocyte processes penetrated into invaginations of the macrophage surface which were bristle-coated at their blind extremity. As previously shown for clathrin-coated pits, the segments of the macrophage plasma membrane beneath which the defense material was found were selectively resistant to the membrane-perturbing effect of the antibiotic, filipin. This structural specialization of the macrophage plasma membrane at sites of ligand-receptor interaction during immune phagocytosis of antibody-coated erythrocytes may represent the morphological counterpart of the zipper mechanism of phagocytosis previously demonstrated by functional studies.

Animals↗
PubMed1983

Lectin-binding sites as markers of Golgi subcompartments: proximal-to-distal maturation of oligosaccharides.

We investigated the subcellular sites of glycoprotein oligosaccharide maturation by using lectin conjugates to stain lightly-fixed, saponin-permeabilized myeloma cells. At the electron microscopic level, concanavalin A-peroxidase stains the cisternal space of the nuclear envelope, the rough endoplasmic reticulum, and cisternae along the proximal face of the Golgi stack. Conversely, wheat germ agglutinin-peroxidase stains cisternae along the distal face of the Golgi stack, associated vesicles, and the cell surface. These observations confirm the existence of two qualitatively distinct Golgi subcompartments, show that the lectin conjugates can be employed as relatively proximal or distal Golgi markers under conditions of excellent ultrastructural preservation, suggest that the asymmetric distribution of qualitatively distinct oligosaccharides is a property of underlying cellular components and not simply of the principal secretory product, and suggest that the oligosaccharide structure recognized by wheat germ agglutinin is attained during transport from the proximal toward the distal face of the Golgi stack.

Animals↗
PubMed1983

In vitro rapid organization of endothelial cells into capillary-like networks is promoted by collagen matrices.

We have studied the behavior of cloned capillary endothelial cells grown inside a three dimensional collagen matrix. Cell monolayers established on the surface of collagen gels were covered with a second layer of collagen. This induced the monolayers of endothelial cells to reorganize into a network of branching and anastomosing capillary-like tubes. As seen by electron microscopy, the tubes were formed by at least two cells (in transverse sections) delimiting a narrow lumen. In addition, distinct basal lamina material was present between the abluminal face of the endothelial cells and the collagen matrix. These results showed that capillary endothelial cells have the capacity to form vessel-like structures with well-oriented cell polarity in vitro. They also suggest that an appropriate topological relationship of endothelial cells with collagen matrices, similar to that occurring in vivo, has an inducive role on the expression of this potential. This culture system provides a simple in vitro model for studying the factors involved in the formation of new blood vessels (angiogenesis).

Animals↗
PubMed1983

Fate of T-lymphocyte injected into immunodeficient allogeneic nude or semi-allogeneic F1 mice: correlation with manifestations of graft-versus-host reaction.

This review emphasizes some of the differences between the fate of allogeneic H2-incompatible T-cells, when injected into immunodeficient, T-depleted mice, or as parental cells injected into F1 hybrids. In both conditions, allogeneic cells are eliminated, a rejection which cannot be predicted from the fate of a skin graft. While in both situations a phase of GVHR is observed, it is of short duration in allogeneic T-B reactions, and it is obvious that the production of allo-antibodies by the host, which is enhanced by the GVHR, is the major mechanism of allogeneic T-cell elimination, resulting in a rapid self-cure of the GVHR. In T-vs-B GVHR, the enhancement of the humoral response is strikingly restricted to the allo-antigens of the T-cells, and correspond therefore to a peculiar form of "allogeneic effect", in fact an "allogeneic suicide" which implies a linked T-B collaboration, while in P-F1 combinations, there is a enhancement of the humoral response to unrelated antigens (i.e. unlinked T-B collaboration or the classical allogeneic effect) and a polyclonal B-cell activation, which can result from the much more protracted course of the GVHR. The parental cells surviving in the spleen of an F1 semi-allogeneic host, undergoing a lethal GVHR were characterized at the clonal level: they were exclusively of the Lyt 2- MLR responder and polyvalent helper variety. This indicates that the Lyt-2- and polyvalent helper subset is selected during an allo-reaction in vivo (either because it is more rapidly replicating and/or less negatively selected by the host resistance) and that it is capable of inflicting a lethal GVHR.

Animals↗
PubMed1983

Positively selected Lyt-2+ and Lyt-2- mouse T lymphocytes are comparable, after Con A stimulation, in release of IL 2 and of lymphokines acting on B cells, macrophages, and mast cells, but differ in interferon production.

Purified mouse T lymphocytes were separated into Lyt-2+ and Lyt-2- populations by the procedure of panning, in which a monoclonal rat anti-Lyt-2 antibody and dishes coated with affinity-purified mouse anti-rat Ig antibodies were used. The populations obtained were 95 to 99% pure as determined by immunofluorescence. Graded doses of these T cells were cultured with optimal mitogenic doses of concanavalin A and the 0 to 24 and 24 to 48-hr culture supernatants were collected. The dose-curve assays of the supernatants of Lyt-2+ and Lyt-2- cells showed comparable activity in interleukin 2 (IL 2) and T cell-replacing factor (TRF), assayed on antigen-stimulated culture of T-depleted spleen cells. Limiting dilution assays of IL 2-secreting precursor cells stimulated by Con A showed a high frequency of precursors in both populations, slightly higher among Lyt-2- cells. The supernatants also contained comparable levels of IPA (inducer of plasminogen activator production by the macrophages), MAF (macrophage-activating factor, assayed by induction of their cytolytic function), and MCGF (mast cells growth factor, assayed on a mast cell line). IPA and MAF were not produced with the same kinetics and in the same T cell concentration conditions as IL 2 and TRF. In contrast, interferon was principally produced by the Lyt-2+ cells.

Animals↗
PubMed1982

The area of attachment of cytotoxic T lymphocytes to their target cells shows high motility and polarization of actin, but not myosin.

Conjugates of cytotoxic T lymphocytes attached to their target cells were studied by double immunofluorescence on fixed smears to detect simultaneously the localization of actin and myosin within the cells. Actin was found to be polarized in the area of attachment of the lymphocytes (but not of the target cells), whereas myosin remained evenly distributed in the cytoplasm. When living conjugates were brought to 37 degrees C to induce cytotoxicity, this pattern remained unchanged, but observation by interference reflexion microcinematography revealed a high motility of the lymphocytes in the contact area. This localized motility in the area of attachment associated with a peculiar actin polarization, which has no equivalent in any type of cell contact presently known, could represent a necessary step in the sequence of events leading to target cell killing by cytotoxic T lymphocytes.

Actins↗