Incorporation of rat histocompatibility (AgB) antigens into liposomes, and their susceptibility to immune lysis

Willoughby, Eileen; Turner, Mervyn J.; Sanderson, Arnold R.

doi:10.1002/eji.1830080905

Cited by 18 publications

(6 citation statements)

References 24 publications

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“…3B) Thin-section electron microscopy of liposomes prepared from H-2Kk and lipid showed them to be spherical unilamellar vesicles of relatively homogeneous size (Fig. 4) (20)(21)(22).…”

Section: Methodsmentioning

confidence: 96%

“…Both these liposomes and liposomes made from H-2 and lipid appear to have most of the antigen on the surface. Preferential exposure of MHC antigen on the outer surface of vesicles prepared by dialysis has been found in a number ofstudies (20)(21)(22)(23)(24). Increased efficiency of uptake by adherent cells cannot account for the greater antigenicity of liposomes either.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Secondary cytolytic T lymphocyte stimulation by purified H-2Kk in liposomes.

Herrmann¹,

Mescher²

1981

Proc. Natl. Acad. Sci. U.S.A.

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Purified H-2Kk' incorporated into lipid vesicles induced a secondary allogeneic cytolytic T lymphocyte response. However, the level of the response was much less than that generated by using purified plasma membranes containing an equivalent amount of antigen. Similarly, reconstituted membranes stimulated less effectively than did intact plasma membranes. In both cases the stimulating activity of the antigen was increased by including a detergent-insoluble membrane matrix fraction during formation of the liposomes or reconstituted vesicles. Liposomes formed in the presence of the matrix were larger, were more irregular in shape, and had a higher density than those formed in its absence. Both the H-2 antigen and matrix proteins were incorporated into the same vesicles. The greater antigenicity of H-2 in vesicles containing the matrix protein might be due to either the larger size of the liposomes or interaction of the antigen with a' component(s) of the matrix.Generation ofa cytolytic T lymphocyte (CTL) response requires the recognition of cell surface antigens of the stimulator cells. The demonstration that purified membranes (1) and detergentsolubilized reconstituted membranes (2, 3) from stimulator cells are able to generate a specific in vitro secondary allogeneic CTL response has made it possible to-begin to examine the requirements for recognition at the molecular level. Purified HLA antigens (4) and partially purified'H-2 antigens (5) have been shown to stimulate a secondary CTL response when incorporated into liposomes. Similarly, liposomes containing the appropriate foreign antigens and H-2 will specifically stimulate secondary virus-specific (6-9) and tumor-specific (10,11) responses.Previous studies (2, 5) had demonstrated that reconstituted membranes could stimulate generation of an allogeneic CTL response, but the level of response was much lower than that obtained by using intact membranes. The results reported here demonstrate that activity comparable to that of intact membranes is obtained if the detergent-insoluble fraction of the membrane is not removed prior to reconstitution. Furthermore, including this isolated, detergent-insoluble fraction during incorporation of purified H-2K into liposomes results in liposomes that effectively stimulate an allogeneic response.Examination of the detergent-insoluble fraction of plasma membranes from murine tumor and lymphoid cells has shown it to consist of a relatively small number of membrane proteins which remain selectively insoluble (12). The major protein components are actin and proteins with molecular weights of70,000, 69,000, 38,000, and 36,000. '5'-Nucleotidase, a cell surface glycoprotein, also remains associated with the detergent-insoluble proteins. The proteins appear to form a matrix at the inner surface ofthe plasma membrane which is, in many respects, similar to the spectrin matrix of erythrocytes. Thin-section electron micrographs of the isolated detergent-insoluble matrix show it to be primarily in the form of closed structures si...

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“…3B) Thin-section electron microscopy of liposomes prepared from H-2Kk and lipid showed them to be spherical unilamellar vesicles of relatively homogeneous size (Fig. 4) (20)(21)(22).…”

Section: Methodsmentioning

confidence: 96%

Section: Methodsmentioning

confidence: 99%

Secondary cytolytic T lymphocyte stimulation by purified H-2Kk in liposomes.

Herrmann¹,

Mescher²

1981

Proc. Natl. Acad. Sci. U.S.A.

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“…The high protein content may also account for the low spontaneous radionuclide release we observed in our vesicle preparation, as opposed to the high background release in other vesicle systems (17). The diffusion of small molecules has been reported for proteoliposomes (18,19) and attributed to the displacement of lipid by hydrophobic protein.…”

mentioning

confidence: 47%

Allogeneic cytolysis of reconstituted membrane vesicles.

Hollander¹,

Mehdi²,

Il³

et al. 1979

Proc. Natl. Acad. Sci. U.S.A.

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The successful use of lipid bilayer model membranes as targets for cytotoxic lymphocytes is described.Lipid vesicles were made from a mixture of dipalmitoyl lecithin, dimyristoyl lecithin, and cholesterol. Membrane proteins of LSTRA or EL4 tumor cells (as source of H-2 antigens), human eye muscle membrane proteins (as supporting proteins), and 51Cr marker were inserted into the lipid vesicles. Incubation of the reconstituted vesicles with lymphocytes sensitized in mixed lymphocyte cultures against allogeneic cells resulted in the specific release of intravesicular 5tCr. Vesicle damage was mediated by thymus-derived lymphocytes. H-2 antigens could be incorporated into vesicles without eye muscle proteins. However, immune damage of the vesicles could not be demonstrated when vesicles inserted with H-2 antigens in the absence of eye muscle proteins were used as targets. Thymus-derived (T) lymphocyte-mediated cytotoxic reactions to alloantigens, virus antigens, minor histocompatibility antigens, and haptens all involve direct or associated recognition of gene products of the major histocompatibility complex (MHC) (1-4). Thus, T cell receptors recognize on target cells both MHC and other antigenic ligands. The mechanism whereby T cell reactivity is restricted by target cell MHC phenotype is not yet understood. Model membranes should be useful for the study of allogeneic T cell cytotoxicity as well as the MHC restriction phenomenon, because they involve welldefined systems in which lipid vesicles may be reconstituted with purified MHC gene products in the presence or absence of other cell membrane proteins or exogenous antigens. Although murine MHC components have been incorporated into synthetic liposomes (5), and reconstituted vesicles have been utilized in the in vitro restimulation of cytotoxic T cells involved in MHC-restricted Sendai virus immunity (6), such reconstituted vesicles have not yet been reported as targets for cytotoxic T cells.In a different system designed to study the mechanism of eye muscle injury in human Graves ophthalmopathy, lipid vesicles inserted with human eye muscle membrane proteins (EMP) were damaged when incubated with lymphocytes from patients with Graves ophthalmopathy in the presence of thyroglobulin (Tg) or thyroglobulin-anti thyroglobulin complexes (TgA), whereas lymphocytes from control subjects damaged such vesicles only in the presence of TgA (7). Lipid vesicles lacking EMP or containing other tissue proteins in equivalent amounts were not lysed even if TgA complexes were present, indicating that EMP have some characteristic that renders lipid vesicles susceptible to lysis by immune cells.The first step in our approach of using model membrane systems as targets in cell-mediated cytolytic reactions was therefore the use of lipid vesicles into which both murine cellThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact...

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“…However, ihis procedure does not yield intact AgB antigens in contrast to the method outlined by Willoughby et ul. [33].…”

Section: Discussionmentioning

confidence: 99%

“…Katagiri ('/ (//. [9] have described the partial purilicatlon of papain-solubilized AgB antigens, and recetitly Willoughby et al [33] published an isolation scheme that yields detergent-solubilized AgB antigens contaminated with only a lew proteins.…”

mentioning

confidence: 99%

Properties of Purified Papain‐solubilized Rat AgB Antigens and Reactivity of a Xenoantiserum against the Isolated Antigens

1980

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Rat AgB transplantation antigens were isolated after papain digestion of spleens from the inbred strain Hooded Lister. Both subunits of the AgB antigens were present in the purified material. Some physical characteristics of the antigens have been determined. An antiserum, raised in a rabbit, against the purified material reacted exclusively with AgB antigens on splenocytes but detected novel structures on both adult and embryonic fibroblasts. These structures, antigenically related to AgB antigens, were not detected on plasmacytoma or hepatoma cells, nor did they display any antigenic similarity with rat beta 2-microglobulin. Radioimmunoassays specific for the AgB antigen heavy chain and for beta 2-microglobulin, respectively, were used to estimate the contents of these antigens in several tissues. Spleen and thymus exhibit the largest density, while brain is almost devoid of these antigens.

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Incorporation of rat histocompatibility (AgB) antigens into liposomes, and their susceptibility to immune lysis

Cited by 18 publications

References 24 publications

Secondary cytolytic T lymphocyte stimulation by purified H-2Kk in liposomes.

Secondary cytolytic T lymphocyte stimulation by purified H-2Kk in liposomes.

Allogeneic cytolysis of reconstituted membrane vesicles.

Properties of Purified Papain‐solubilized Rat AgB Antigens and Reactivity of a Xenoantiserum against the Isolated Antigens

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