| 1. |
- Lundqvist, Carina, et al.
(författare)
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Cytokine profile and ultrastructure of intraepithelial gamma delta T cells in chronically inflamed human gingiva suggest a cytotoxic effector function
- 1994
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Ingår i: Journal of Immunology. - 0022-1767 .- 1550-6606. ; 153:5, s. 2302-2312
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Tidskriftsartikel (refereegranskat)abstract
- We have shown that gamma delta T cells in human gingiva have an intraepithelial location and, that in the chronic inflammatory disease periodontitis, the expression of CD45RO and CD8 or CD4 is induced on gamma delta T cells. To study the role of gamma delta T cells in local antibacterial responses, we determined the cytokine profiles of isolated human gingival cells. Different T cell subpopulations, isolated by positive selection with mAb-coated magnetic beads and macrophages, as well as epithelial cells, were analyzed for expression of mRNA for 15 cytokines by reverse transcriptase-PCR. The ultrastructure of gingival gamma delta T cells was also studied. The gamma delta T cells expressed mRNA for IFN-gamma, TNF-alpha, TGF-beta 1, and IL-6. Expression of IFN-gamma was a consequence of inflammation. CD4+ gamma delta T cells expressed IFN-gamma only, whereas CD8+ gamma delta T cells expressed all four cytokines. CD8+ cells expressing IFN-gamma, TNF-alpha, and IL-6 in combination suggest a cytotoxic effector function. Gingival gamma delta T cells contained cytoplasmic electron-dense membrane-bound granules and multivesicular bodies that are ultrastructural characteristics of cytotoxic cells. Epithelial cells from inflamed gingiva expressed HLA-DR, CD1a, CD1c, and heat shock protein 60 on the cell surface. They also expressed mRNA for IL-1 beta, IL-6, IL-8, TNF-alpha, and TGF-beta 1. Thus, epithelial cells may function as accessory cells in immune activation and, at the same time, be target cells for CD8+ gamma delta T cells reactive with CD1 Ag or heat shock protein. These results suggest that gamma delta T cells constitute a first line of defense in gingiva, preventing entrance of pathogens by cytotoxicity against infected and stressed epithelial cells, and by control of epithelial cell growth through secretion of regulatory cytokines.
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| 2. |
- Lundqvist, Carina
(författare)
-
Human intraepithelial lymphocytes : a comparative study of phenotype, morphology, and functional properties of intraepithelial lymphocytes in gut and oral mucosa
- 1995
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Human intraepithelial lymphocytes (IEL) constitute a unique cell population situated in the first line of defense of the alimentary tract. Here they are continuously exposed to a massive antigenic load of high complexity. However, different conditions prevail along the alimentary tract. In small intestine food antigens dominate whereas bacterial antigens are abundant in large intestine. The oral cavity is exposed to an enormous variety of antigens from the microflora as well as food constituents. The abundance and selective localization of lymphocytes in the surface epithelium of these challenged tissues implicate important roles for IEL in immune protection.IEL in normal human jejunum, ileum and colon as well as in normal and chronically inflamed gingiva were studied in situ and after isolation, with regard to phenotype, ultrastructure, cytokine mRNA expression and response to T-cell mitogens. Furthermore, an isolation technique was developed which yielded highly purified, functionally active IEL and enterocytes from the same sample.Intestinal IEL were situated in the basal part of the epithelium, often in small clusters and in close contact with adjacent lymphocytes and epithelial cells. They had an irregular shape with long processes and some had pseudopodium-like extensions penetrating the basement membrane. This indicates cell co-operation within the epithelium, as well as transmigration of IEL to underlying tissues. Freshly isolated IEL expressed several cytokines (IL-1β, IL-8, IL-2, TNF-α and IFN-γ) and in vitro activation induced expression of IL-2, IL-10, IFN-γ, TNF-α, TNF-β and TGF-β1, suggesting that IEL are involved in cell mediated cytotoxicity and suppressor cell activities.γδ T cells showed preferential homing to the epithelium both in gingiva and in intestine. They constituted the major lymphocyte population in normal gingiva and on average 30% of IEL at all levels of the intestine. Gingival as well as intestinal γδ IEL showed preferential usage of Vδ1Vγ8, suggesting common reactivity patterns along the alimentary tract. Intestinal γδ IEL and γδ IEL in normal gingiva were CD4-CD8-. In contrast, γδ IEL in chronically inflamed gingiva were predominantly CD8+ and showed induced expression of CD45RO. This indicates that γδ IEL participate in anti-bacterial immune responses in mucosa. Intestinal and gingival γδ IEL displayed ultrastructural features of cytolytic effector cells, e.g. electron-dense cytoplasmic granules and multivesicular bodies. They also expressed cytokines indicative of cell mediated-cytolytic effector functions. γδ IEL from inflamed gingiva expressed IFN-γ, TNF-α, TGF-β1 and IL-6 mRNA while intestinal γδ IEL expressed IL-2, IFN-γ and TNF-α.Intraepithelial αβ T cells were rare in gingiva while they constituted the major population of intestinal IEL. The phenotype of αβ IEL varied at different levels of the intestine. Thus, CD8+αβ IEL dominated in jejunum while cells with the unusual T-cell phenotype, CD4-CD8- TCR αβ+, constituted a major population of colonic IEL. CD4+ αβ IEL were equally represented, as a minor population, at all three levels of the gut. Intestinal αβ IEL had the same cytokine profile as γδ IEL. Taken together, these data suggest that αβ IEL are involved in immunoregulatory responses to luminal antigens.IEL with thymocyte-like phenotyped (CD2+TCR/CD3-, CD1+TCR/CD3-, CD1+TCRαβ+ and CD1+TCRγδ+) were present in jejunal epithelium. Furthermore, recombination activating gene-1 (RAG-1) mRNA was expressed in CD2+TCR/CD3- and CD3+/TCR- jejunal IEL. RAG-1 was not expressed in colonic IEL. Thus, the epithelium of small intestine is a site for extrathymic T cell maturation in humans.
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| 3. |
- Lundqvist, Carina, et al.
(författare)
-
Intra-epithelial lymphocytes. Evidence for regional specialization and extrathymic T cell maturation in the human gut epithelium
- 1995
-
Ingår i: International Immunology. - : Oxford University Press (OUP). - 0953-8178 .- 1460-2377. ; 7:9, s. 1473-1487
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Tidskriftsartikel (refereegranskat)abstract
- The human gut epithelium is a unique immunological compartment, containing substantial amounts of intra-epithelial lymphocytes (IEL) with unknown functions. In this study we show that distinct and unusual subpopulations of IEL are present at different levels of human intestine. IEL phenotypes in normal jejunum, ileum and colon were compared using immunoflow cytometry and immunohistochemistry. The expression of mRNA for recombination-activating gene-1 (RAG-1) in IEL from all three levels was compared using reverse-transcription polymerase chain reaction, and the morphology of IEL in situ was determined using immunoelectron microscopy. Surface marker profiles of isolated intestinal epithelial cells at all three levels were also investigated. On average the proportion of TCR gamma delta IEL was comparable in jejunum than ileum and colon and varied in phenotype with gut level. CD4-CD8-TCR alpha beta IEL dominated in colon but were absent in jejunum. CD8+ TCR alpha beta IEL were present at all levels but only in jejunum did they constitute the majority of all IEL. CD4+ TCR alpha beta IEL were present in similar frequencies at all levels of the gut. In general, the majority of IEL had an activated phenotype (CD45RO+, alpha E beta 7+). Furthermore, IEL exhibited phenotypes which are rare in peripheral blood. The thymocyte markers CD1a and CD1c as well as the NK cell marker CD56 were expressed on a fraction of TCR alpha beta and TCR gamma delta IEL. A small population of 'null' cells (CD45+ TCR/CD#-CD20-CD14-CD15- cells) was also present at equal proportions along the gut. Jejunal but not colonic IEL expressed RAG-1 mRNA suggesting that extrathymic T cell maturation occurs in the epithelium of small intestine. RAG-1 was expressed in CD2+TCR/CD3- and CD3+/TCR-IEL. Ultrastructurally, IEL often formed small clusters and intimate contacts with epithelial cells, suggesting cell cooperation within the epithelium. Some IEL had pseudopodium-like extensions penetrating the epithelial basement membrane suggesting transmigration. Epithelial cells in small intestine but not colon expressed heat shock protein 60 and HLA-DR. CD1a, CD1b and CD1c were not expressed on intestinal epithelial cells at any level. The distinct surface marker profiles of IEL and epithelial cells along small and large intestine suggest functional regional specialization and are compatible with the hypothesis that TCR alpha beta IEL participate in immune reactions to lumenal antigens while TCR gamma delta IEL perform surveillance of the epithelium.
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| 4. |
- Lundqvist, C, et al.
(författare)
-
Intraepithelial lymphocytes in human gut have lytic potential and a cytokine profile that suggest T helper 1 and cytotoxic functions.
- 1996
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Ingår i: Journal of Immunology. - 0022-1767 .- 1550-6606. ; 157:5, s. 1926-34
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Tidskriftsartikel (refereegranskat)abstract
- The functional properties of intraepithelial lymphocytes (IEL) in normal human jejunum, ileum, and colon were investigated. Cytokine mRNA expression in IEL and enterocytes was determined by reverse transcriptase-PCR and IFN-gamma+ IEL by immunohistochemistry. Polyclonal activators were used to study proliferation and IFN-gamma secretion of IEL, and an anti-CD3-mediated redirected cytotoxicity assay was used to determine the lytic potential of IEL. Freshly isolated IEL at all three gut levels expressed mRNA for IL-1 beta, IL-2, IL-8, IFN-gamma, and TNF-alpha. Approximately 10% of IEL produced IFN-gamma, suggesting that IEL are immunologically active in vivo, performing similar functions along the intestine. IEL could be stimulated further in vitro to express IL-10, TNF-beta, and TGF-beta 1, while no Th2-type cytokines were induced, suggesting suppressive and cytolytic functions for IEL. All three jejunal IEL subpopulations (CD4-CD8-TCR-gamma delta+, CD4+TCR-alpha beta+, CD8+TCR-alpha beta+) expressed the same four cytokines, IL-2, IL-8, IFN-gamma, and TNF-alpha, indicating that CD4+TCR-alpha beta+ IEL are Th1 cells and that TCR-gamma delta+ IEL and CD8+TCR-alpha beta+ IEL include cytotoxic effector cells. Indeed, freshly isolated jejunal IEL displayed cytolytic activity. IEL were induced to proliferation by anti-CD3/TCR complex mAbs and leukoagglutinin, but not by Con A. There was no correlation between the magnitude of the proliferative response and the amounts of secreted IFN-gamma. Enterocytes expressed IL-1 beta and IL-8, and sometimes TNF-alpha. Although jejunal enterocytes express HLA-DR and hsp60, Ag presentation by these cells may induce anergy since their cytokine profile is different from that of classical APCs.
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| 5. |
- Lundqvist, Carina, et al.
(författare)
-
Isolation of functionally active intraepithelial lymphocytes and enterocytes from human small and large intestine
- 1992
-
Ingår i: JIM - Journal of Immunological Methods. - 0022-1759 .- 1872-7905. ; 152:2, s. 253-263
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Tidskriftsartikel (refereegranskat)abstract
- A mild purification method has been developed for the isolation of human intraepithelial lymphocytes (IEL) and enterocytes from the same individual. The isolation procedure includes mechanical disruption of the mucosal layer, treatment with reducing agent and sedimentation followed by Percoll gradient centrifugation. Finally, epithelial cells are removed from the IEL fraction using magnetic beads coated with the anti-epithelial antigen monoclonal antibody (mAb) BerEP4. Leucocytes are removed from the enterocyte fraction using magnetic beads coated with mAbs directed against common leucocyte antigen (CD45). Using this procedure IEL and enterocytes have been isolated from apparently normal jejunal, ileal and colonic tissue specimens. Recoveries of IEL were 7 x 10(5), 4 x 10(5) and 1 x 10(5)/cm2 mucosa from jejunum, ileum and colon respectively. 1-2 x 10(6) enterocytes/cm2 mucosa were recovered from small intestine while the corresponding value for colonic biopsies was approximately 2 x 10(5) enterocytes/cm2. The IEL fraction was pure as judged by the low percentages of B cells, macrophages and BerEP4 positive cells (less than 4%) present in the purified fraction. The enterocyte fraction contained less than 2% CD45+ cells. The two cell fractions were viable and expanded in vitro. Enterocytes expanded spontaneously while IEL required initial stimulation with mitogens. The isolation procedure described here will make it possible to study the function of human IEL, interactions between IEL and enterocytes and the role of both cell types in local immunity.
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| 6. |
- Mincheva-Nilsson, Lucia, et al.
(författare)
-
Activated human gamma delta T lymphocytes express functional lactoferrin receptors.
- 1997
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Ingår i: Scandinavian Journal of Immunology. - 0300-9475 .- 1365-3083. ; 46:6, s. 609-18
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Tidskriftsartikel (refereegranskat)abstract
- Lactoferrin (Lf), an iron-binding protein in milk, mucosal secretions and neutrophil granules has bactericidal properties and is a source of iron for breast-fed infants. In this paper the authors show that most in vivo activated lymphocytes, i.e. freshly isolated lymphocytes from first trimester human decidua, and most in vitro activated human blood lymphocytes, express lactoferrin receptors (Lf-R), while unstimulated blood lymphocytes do not. All major lymphocyte subsets, i.e. alpha beta T cells, gamma delta T cells, CD8+ T cells, CD4+ T cells, B cells and NK cells, express Lf-R after activation. The proportion of Lf-R expressing activated gamma delta T cells is significantly larger than that of activated alpha beta T cells. Lf-R and transferrin receptors (Tr-R/CD71) show the same kinetics of appearance on activated blood lymphocytes and are, to a large extent, expressed on the same cells. However, 35% of decidual lymphocytes and 15% of activated blood lymphocytes express Lf-R only. Addition of Lf to cultures containing an optimal concentration of Tr augments the proliferative response to polyclonal T cell activators and alloantigens, suggesting that presently used standard culture conditions for in vitro activation are suboptimal in particular for gamma delta T cells. Lf-R on decidual lymphocytes contain bound Lf, which probably is produced locally. The results suggest that Lf is a growth-supporting factor, especially important in local immune responses in the mucosa.
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| 7. |
- Mincheva-Nilsson, Lucia, et al.
(författare)
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Gamma delta T cells of human early pregnancy decidua : evidence for local proliferation, phenotypic heterogeneity, and extrathymic differentiation.
- 1997
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Ingår i: Journal of Immunology. - 0022-1767 .- 1550-6606. ; 159:7, s. 3266-77
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Tidskriftsartikel (refereegranskat)abstract
- The uterine mucosa in pregnancy, the decidua, allows placenta formation and survival of the fetus despite the fact that it is semiallogeneic. Decidua contains large numbers of lymphocytes, of which CD56+ cells dominate, followed by T cells expressing either alpha beta or gamma delta TCR. We have investigated the developmental relationship between the CD56- and TCR gamma delta-expressing cells in early pregnancy decidua using dual labeling immunoelectron microscopy, immunoflow cytometry, and cell fractionation. Lymphocyte subpopulations were, in addition, analyzed for expression of the cytokine receptor for IL-7 and c-kit and for mRNA expression of recombinase-activating genes 1 and 2. Four different cell populations could be distinguished: CD56+bright, CD56+dim/TCR gamma delta+low, CD56+dim/TCR gamma delta+high, and TCR gamma delta+low. Recombinase-activating genes 1 and 2 were expressed in the CD56+bright cells and to a limited degree in CD56+dim/TCR gamma delta+low cells. c-kit was preferentially expressed on the CD56+bright cells, while IL-7R was preferentially expressed on CD56+dim/TCR gamma delta+low and CD56+dim/TCR gamma delta+high cells. The CD56+dim TCR gamma delta+low and CD56+dim/TCR gamma delta+high cells displayed the characteristic morphology of large granular lymphocytes, while single positive TCR gamma delta+low cells were usually smaller and did not contain cytoplasmic granules. The gamma delta 1 gene segment was almost exclusively used in the TCR. Gamma delta T cells in mitosis were seen. We suggest that human early pregnancy decidua is a transient site for extrathymic maturation and that the progenitors of TCR gamma delta+ cells are bone marrow-derived immature cells expressing the CD56 (neural cell adhesion molecule) homing receptor.
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| 8. |
- Mincheva-Nilsson, Lucia, et al.
(författare)
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Human decidual leukocytes from early pregnancy contain high numbers of gamma delta+ cells and show selective down-regulation of alloreactivity.
- 1992
-
Ingår i: Journal of Immunology. - 0022-1767 .- 1550-6606. ; 149:6, s. 2203-11
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Tidskriftsartikel (refereegranskat)abstract
- The mononuclear lymphoid cell population in human pregnant uterus mucosa, decidua, from early normal pregnancies was studied phenotypically and functionally. The phenotype was determined in situ by immunohistochemistry, and in isolated decidual mononuclear cell preparations by immunofluorescence and flow cytometry. A mild isolation procedure of gentle mechanical disruption followed by density gradient centrifugation was used. Leukocytes comprised a large part of the decidual tissue. They were present in aggregates mainly situated adjacent to the glandular epithelium. In addition, individual leukocytes were present intraepithelially, as well as scattered between the stromal cells and around vessels and lacunes. Four lymphocyte populations of approximately the same size were identified: TCR gamma delta+/CD56+ cells, TCR gamma delta+/CD56- cells, TCR gamma delta-/CD56+ cells, and TCR alpha beta+/CD8+ cells. TCR gamma delta- expressing cells comprised about 60% of the T cells. They were CD4-/CD8-, and about half of the TCR gamma delta+ cells expressed the memory/activation marker CD45RO. The Kp 43 Ag, earlier described on activated CD56+ and TCR gamma delta+ cells in peripheral blood, was essentially only expressed on the TCR gamma delta-/CD56+ cell population in decidua. At least 50% of the TCR alpha beta+ cells were CD8+. The function(s) of either one of these populations might be to prevent immunologic reactions against the fetus, to protect the uterus from unwanted extensive invasion of trophoblasts, or to protect the uteroplacental unit from infection. Decidual T cells did not respond to stimulation by alloantigens or mitogenic anti-CD3 mAb but responded to the same extent as PBMC to mitogenic lectins. The surface density of the TCR/CD3 complex was low on freshly isolated decidual lymphocytes, but could be up-regulated upon stimulation with PMA/Ionomycin. Local selective down-regulation of surface expression of the TCR/CD3 complex and of activation involving this complex might be one of the mechanisms by which a maternal immunologic reaction against the semiallogeneic fetus is prevented.
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| 9. |
- Mincheva-Nilsson, Lucia, et al.
(författare)
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Human milk contains proteins that stimulate and suppress T lymphocyte proliferation
- 1990
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Ingår i: Clinical and Experimental Immunology. - : Oxford University Press. - 0009-9104 .- 1365-2249. ; 79:3, s. 463-469
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Tidskriftsartikel (refereegranskat)abstract
- The modulatory effect of human milk proteins from colostrum and late milk on the proliferative response of human T lymphocytes activated by mitogens (OKT3 and leucoagglutinin from Phaseolus vulgaris) and alloantigens was studied. High concentrations (10-100 micrograms/ml) of crude colostral milk proteins had an inhibitory effect on T cell growth while low concentrations (0.1-1 microgram/ml) enhanced T cells growth. In contrast, proteins from late milk did not inhibit T lymphocyte proliferation while the enhancing effect was retained. Colostrum was fractionated by ammonium sulphate precipitation and gel filtration on sepharose 6B. The inhibitory activity was recovered in a protein fraction containing lactoferrin as its major component. Lactoferrin was, however, not responsible for the observed inhibition. On the contrary, lactoferrin in most cases augmented the proliferative response induced by polyclonal activators. The inhibitory activity was found to bind concanavalin A-sepharose suggesting an association with glycoprotein. Inhibitory fractions contained glycoproteins of the following molecular sizes 26, 74/76 (doublet), 84, 145 and 160 kD under reducing conditions. The inhibitory effect appeared to be lymphocyte specific since the active fraction did not inhibit the growth of tissue culture cells (HeLa cells and human fibroblasts) or bacteria. Furthermore, the fraction was not toxic for lymphocytes. The inhibitory colostrum factor may prevent the newborn from overreacting immunologically against the environmental antigens encountered at birth.
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| 10. |
- Mincheva-Nilsson, Lucia, et al.
(författare)
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Immunomorphologic studies of human decidua-associated lymphoid cells in normal early pregnancy.
- 1994
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Ingår i: Journal of Immunology. - 0022-1767 .- 1550-6606. ; 152:4, s. 2020-32
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Tidskriftsartikel (refereegranskat)abstract
- Human decidual lymphocytes from early, normal pregnancy were characterized in situ with respect to ultrastructure and distribution of subsets. The ultrastructure of isolated decidual gamma delta T cells was also studied. CD45+ cells comprised 11 +/- 2% of all decidual cells. The majority were localized in large lymphoid cell clusters (LCC), near endometrial glands, or as intraepithelial lymphocytes (IEL) in glandular epithelium. The major cell populations in LCC were CD56+TCR-gamma delta+ cells, CD56+ cells, TCR-alpha beta+CD4+ cells, and TCR-alpha beta+CD8+ cells. All expressed activation markers (CD45RO, Kp43, and/or HML-1) and MHC class II Ag (HLA-DR, HLA-DP, and/or HLA-DQ). No B cells were found. Almost all IEL were activated TCR-gamma delta+ cells (CD56+ and CD56-). The glandular epithelial cells expressed heat shock protein 60 at the basolateral side facing the TCR-gamma delta+ IEL. Decidual lymphocytes displayed cytoplasmic processes, microvilli, characteristic cytoplasmic granules, and had intimate contact with neighboring cells. Lymphocytes in the outer rim of LCC and the stroma showed signs of cellular movement. Two main morphotypes of gamma delta T cells could be distinguished. One had single microvilli, membrane-bound granules, and nuclear inclusions. The other had many microvilli, nonmembrane-bound granules and cytoplasmic multivesicular bodies. Our data suggest that LCC are centers of immune reactivity where T and NK cells become activated. The activated cells may guard against infections and undue trophoblast invasion and/or be involved in modulating the local maternal immune system toward unresponsiveness against the semiallogeneic fetus.
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| 11. |
- Nap, M, et al.
(författare)
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Specificity and affinity of monoclonal antibodies against carcinoembryonic antigen.
- 1992
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Ingår i: Cancer Research. - 0008-5472 .- 1538-7445. ; 52:8, s. 2329-39
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Tidskriftsartikel (refereegranskat)abstract
- The binding specificities of 52 well-characterized monoclonal antibodies (Mabs) against carcinoembryonic antigen (CEA) from 12 different research groups were studied by immunohistochemistry and immuno flow cytometry. In addition, the binding constant for the interaction between Mab and CEA was determined by a solution-phase assay. Cryostat sections of colon carcinoma and normal colon, stomach, liver, pancreas, and spleen were studied by immunohistochemistry. Peripheral blood granulocytes, monocytes, and lymphocytes were assayed by immuno flow cytometry. The Mabs used here have previously been classified into five essentially nonoverlapping epitope groups (GOLD 1-5) (Cancer Res., 49: 4852-4858, 1989). Most Mabs cross-reacted with different normal tissues, ranging from highly cross-reactive Mabs (positive reaction with 8 of 9 discriminating tissues) to relatively specific Mabs (positive reaction with 1 of 9 discriminating tissues). Five Mabs (10%) were specific, reacting only with colon carcinoma, normal colon mucosa, and normal gastric foveola. There was a correlation between epitope group and binding specificity. Mabs with a high degree of CEA specificity almost exclusively belonged to epitope groups 1, 2, and 3, while highly cross-reactive Mabs belonged to epitope groups 4 and 5. There was no correlation between antibody specificity and affinity for CEA. Specific Mabs with high as well as low affinity were found.
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| 12. |
- Yeung, Moorix Mo-Wai
(författare)
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Specific and nonspecific immune mechanisms in human gut : a comparative study of normal and ulcerative colitis intestine
- 1996
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The intestine, with its large mucosal surface area, digests and absorbs food nutrients and maintains a beneficial microbial flora in the colon. Local protective immune responses against intestinal pathogens ensure the survival of the individual. These immune reactions are both specific and non-specific in nature. The intestinal epithelium is single-layered and constantly renewed with differentiating epithelial cells moving from the crypt to the luminal surface. Intraepithelial lymphocytes (IEL) are interspersed between the epithelial cells. Ulcerative colitis (UC) is a life-threatening chronic inflammation affecting colon.In this study three molecules belonging to the carcinoembryonic antigen (CEA) family, namely CEA proper, nonspecific cross-reacting antigen 50/90 (NCA 50/90) and biliary glycoprotein (BGP), were found to be specifically localized to the apical surface of colonic epithelial cells. Full expression of these molecules occurs when the cells reach the upper 1/3 of the crypts and are maintained on the mature cells at the luminal surface. Ultrastructurally, CEA, NCA50/90 and BGP are localized to microvesicles and microfilaments of the fuzzy coat/glycocalyx as well as to the microvilli of the epithelial cells. Their unique localization and documented bacterial binding capacities suggest that they have a role in innate immunity.Functional analysis of IEL in normal jejunum, ileum and colon revealed that IEL are in vivo activated T-lymphocytes expressing mRNA for the cytokines IL-1β, IL-2, IL-8, IFNγ and TNFα and that jejunal IEL have T-cell receptor (TCR)/CD3 dependent cytolytic capacity. As many as 10% of IEL actively produce IFNγ. CD4+TCRαβ+IEL, CD8+TCRαβ+IEL and CD4-CD8-TCRαβ+IEL all had a TH1/cytotoxic cytokine profile. IEL could be further stimulated in vitro to express IL-10, TNFβ and TGFβ1, to proliferate and to secrete IFNγ. Thus, active protection and/or regulation of the epithelium via cell-mediated immune reactions are prominent in the gut.UC colon was characterized by a marked lymphocyte infiltration in the lamina propria, 10-50 times the normal level. Most lymphocytes were present in follicle-like cell aggregates containing both T- and B-cells. An unexpected finding was that γδT-cells constituted about 15% of the cells in the aggregates. Such cells are only found intraepithelially in normal gut. γδT-cells of both the intestinal- (TCR-Vδ1/Vγ8) and blood type (TCR-Vδ2/Vγ9) were seen. T-cells in UC colon were activated but nonproliferating and had a down-regulated TCR/CD3 complex. RT-PCR and quantitative immunohistochemistry for cytokine mRNA (n=11) and protein respectively, revealed that the T-cells in UC colon did not produce IL-2, in marked contrast to T-cells in normal colon and to ileal T-cells from UC patients. This was a selective defect since TNFα, IFNγ, IL-1β, IL-8 and TGFβ1 were similarly expressed in normal colon. No TH2 cytokines were seen. Lamina propria leukocytes in UC colon expressed IL-6, a cytokine not found in normal colon. The epithelial cells of UC colon were activated expressing MHC class II antigens, heat shock proteins and the co-stimulatory molecule B7.1/CD80.Our study demonstrates that UC is an immunological disease. The immunopathological picture seen in UC colon probably reflects an inappropriate down-regulation of local immune responses perhaps due to a selective loss of the key cytokine IL-2 in a situation of extreme antigenic stress.
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| 13. |
- Baranov, Vladimir, et al.
(författare)
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Expression of carcinoembryonic antigen and nonspecific cross-reacting 50-kDa antigen in human normal and cancerous colon mucosa : comparative ultrastructural study with monoclonal antibodies
- 1994
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Ingår i: Cancer Research. - 0008-5472 .- 1538-7445. ; 54:12, s. 3305-3314
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Tidskriftsartikel (refereegranskat)abstract
- The precise localization of carcinoembryonic antigen (CEA) and non-specific cross-reacting 50-kDa antigen (NCA 50) in normal colon mucosa and colon adenocarcinoma was investigated by using an indirect immunoperoxidase electron microscopic technique with specific monoclonal antibodies. In normal adult colon both antigens were localized to microvesicles and filaments of the "fuzzy coat" on the apical surface of the epithelial cells. In addition, NCA 50 was found in the narrow spaces between adjoining microvilli. Mature columnar cells at the free luminal surface contained most of the antigen positive material. CEA and NCA 50 were also detected as intracellular components of goblet cells. In multilayered tumor glands, the cell surface expression of the antigens was dependent on the position of the tumor cell in the gland. The neoplastic cells showed either a predominant apical labeling or a positive staining of almost the entire cell surface. Some of the neoplastic cells contained CEA in so-called "intracellular lumina." In contrast to normal colon epithelial cells most tumor cells synthesized NCA 50 actively. In normal colonic mucosa, unlike in cancerous tissue, CEA and NCA 50 appear to be released via vesicles formed from the microvillous membrane of mature columnar cells. These results are consistent with the hypothesis that CEA and NCA play a role in the nonspecific defense against microorganisms in the large intestine.
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| 14. |
- Frängsmyr, Lars, et al.
(författare)
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Cell- and region-specific expression of biliary glycoprotein and its messenger RNA in normal human colonic mucosa
- 1995
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Ingår i: Cancer Research. - 0008-5472 .- 1538-7445. ; 55:14, s. 2963-2967
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Tidskriftsartikel (refereegranskat)abstract
- The localization of biliary glycoprotein (BGP) and its mRNA in normal colonic mucosa was studied by immunohistochemistry and in situ hybridization. BGP mRNA was confined to columnar epithelial cells and expressed abundantly in the superficial mature cells and at low levels in differentiating cells in the upper crypts. Epithelial expression of BGP coincided with that of BGP mRNA. Ultrastructurally, BGP was localized to microfilaments of the fuzzy coat of the columnar cells at the luminal surface and the upper crypts. Additionally, BGP was found in cryptal caveolated cells. The results are consistent with primary transcriptional regulation of BGP production and suggest that BGP synthesis is controlled by the degree of cytodifferentiation. The fuzzy-coat localization of BGP implies a role in nonspecific defense mechanisms against pathogens.
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| 15. |
- Goldsteins, Gundars, et al.
(författare)
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Characterisation of two highly amyloidogenic mutants of transthyretin
- 1997
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 36:18, s. 5346-5352
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Tidskriftsartikel (refereegranskat)abstract
- The plasma protein transthyretin (TTR) has the potential to form amyloid under certain conditions. More than 50 different point mutations have been associated with amyloid formation that occurs only in adults. It is not known what structural changes are introduced into the structure of this otherwise stable molecule that results in its aggregation into insoluble amyloid fibrils. On the basis of calculations of the frequency of known mutations over the polypeptide, we have constructed two mutants in the D-strand of the polypeptide. These molecules, containing either a deletion or a substitution at amino acid positions 53−55, were unstable and spontaneously formed aggregates upon storage in TBS (pH 7.6). The precipitates were shown to be amyloid by staining with thioflavin T and Congo Red. Their ultrastructure was very similar to that of amyloid fibrils deposited in the vitreous body of patients with familial amyloidotic polyneuropathy type 1 with an amino acid replacement in position 30 (TTRmet30). Like amyloid isolated from the vitreous body of the eye, the amyloid precipitates generated from the TTR mutants exposed a trypsin cleavage site between amino acid residues 48 and 49, while plasma TTRmet30 isolated from amyloidosis patients as well as wild-type TTR only showed minor trypsin sensitivity. Our data indicate that the mutants we have constructed are similar to amyloid precursors or may share structural properties with intermediates on a pathway leading to amyloid deposits of plasma TTR.
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| 16. |
- Mincheva-Nilsson, Lucia
(författare)
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Immune cells in pregnant uterine mucosa : functional properties, cellular composition and tissue organization
- 1993
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The pregnant uterus mucosa - decidua - is an "immunologically privileged" site. A semiallogeneic embryo is allowed to survive, develop, and grow while the same tissue implanted outside the uterus will be rejected. The decidua basalis, which participates in the placenta formation, is a tissue rich in lymphoid cells. We have studied decidua associated mononuclear cells (DMC) from normal early pregnancies in humans. The cells were investigated with respect to surface marker profiles, ultrastructure, organization in the tissue, and functional properties. In addition, we have studied the expression of receptors for the iron-binding protein lactoferrin on these cells, and characterized the receptor (Lf-R).Ten to fiftee percent of all cells in decidua belong to the lymphoid cell lineage. They are present in aggregates [lymphoid cell clusters (LCC) mainly located in the vicinity of decidual/endometrial glands] and as individual cells, intra- or subepithelially along the glands (IEL) , and in the stroma. The LCC appear to be centres of immune reactivity. They occur at a frequency of 0.40.2/mm2 tissue and are composed of different population of activated T cells and NK cells in close contact with each other. Interestingly, B cells are not present in the LCC. DMC consist of four major lymphocyte subpopulation of similar sizes: TCRγδ+/CD56+cells, TCRγδ+/CD56-cells, TCRγδ-/CD56+cells and TCRαβ+/CD8+cells. TCRαβ+/CD4+ cells and monocytes are also present. Most DMC have long, thick processes, microvilli, and cytoplasmic granules. They are in intimate contact with surrounding lymphoid, epithelial and stromal cells. Signs of cellular movement and excretion of granules are also seen.About half of the T cells are TCRαβ cells. These cells lack CD4 and CD8. A large fraction of them are CD56+, a rare phenotype at other sites. Most of the TCRγδ+ cells express activation/memory markers (CD45RO, the Kp43 antigen, transferrin receptor, and MHC class II antigens), and many cells express the mucosa homing receptor HML-1. Morphologically these cells either display features characteristic for cytotoxic cells or contain unique nuclear inclusions.More than half of TCRαβ cells are CD8+, but CD4+ cells are also found. These cells also display activation markers.DMC use both transferrin and lactoferrin for their iron supply. The Lf-R on activated lymphocytes appears to be made of two peptides of 47 and 65 kD MW.Freshly isolated DMC respond poorly or not at all to activation through the TCR/CD3 complex, probably due to the low surface density of the complex. However, the TCR/CD3 complex can be up-regulated by stimulation with PMA/Ionomycin in vitro, suggesting that the lymphocytes are suppressed in vivo. Glandular epithelial cells produce immunosuppressive factor(s) that act on CD8+, TCRγδ+, and CD56+ cells. The proximity between the LCC and the glands indicates that this factor(s) may play a role in local immunosuppression. The identity of the factor(s) is presently unknown. The cytokine mRNA profile of DMC, as determined by RT-PCR, reveals IFN-γ, IL-8 and TGF-β1 mRNA in all samples, and IL-1β, IL-2, IL-10, TNF-α and GM-CSF mRNA in some samples. The cytokine profile is compatible with down-regulation of CTL activity.The demands on the immune system in pregnant uterus mucosa are unique. On one hand, a genetically incompatible fetus must be accepted, the development of the placenta must be allowed, and the uteral mucosal tissue must be remodelled. On the other hand, the invasiveness of the trophoblast must be controlled, and the fetomaternal unit must be protected against infections. Our studies indicate that this is achieved through a highly regulated process involving different types of activated lymphoid cells interacting with each other and with glandular epithelial cells.
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| 17. |
- Olofsson, Katarina
(författare)
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Immune cells in human pharyngeal and palatine tonsils and in the uvula : tissue distribution, cellular composition and functional properties
- 1999
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The adenoid (pharyngeal tonsil), the palatine tonsils and the uvula are strategically located at the entrance of the upper airodigestive tract. By virtue of their location they are incessantly exposed to inhaled and ingested antigens. Severe nasal obstruction, otitis media with effusion, recurrent tinsillitis and obstructive airways during sleep are conditions often due to diseases in these organs.To advance our knowledge about the etiology and pathogenesis of these diseases, we have compared immune cell composition, cytokine expression and microbial colonisation in adenoids from children with hypertrophic obstructive adenoid (HOA) and chronically infected adenoid (CIA). Similarly, we compared immune cell composition in palatine tonsils from children with idiopathic tonsillar hypertrophy and recurrent tonsillitis. Finally, we wanted to characterise the human uvula from an immunological point of view, which had previously not been done. Its composition and distribution of immune cells, its cytokine profile, connective tissue elements and ultrastructure was studied.When comparing adenoids from children with HOA and CIA, the most striking finding was their similarity. A cytokine profile that was independent of diagnosis but seemed characteristic for the adenoid emerged. T cell expression of IL-5 and TGF-β1 but not IL-4 suggested an ongoing humoral response driven by a "mucosal TH2" cell. αβ T cells also expressed TNF-α, IFN-γ and IL-2, indicating a concomitant cell mediated response. Cell mediated immune responses often reflect viral infection. In line with this, adenovirus DNA was found in 80% of the adenoid samples. Furthermore, IL-6, IL-8 and TNF-α expressed in the non-T cell fraction suggested that the tissue macrophages were activated. TNF-α, IFN-γ and TGF-β1 were expressed by γδ T cells. The following differences between HOA and CIA were however, noted: i) most intraepithelial lymphocytes were CD8+ γδ T cells in HOA, while CD8+ αβ T cells dominated intraepithelially in CIA; ii) the number of follicles was twice as high in CIA as in HOA; iii) there were signifacantly more granulocytes in the interfollicular area in CIA than in HOA; iv)IL-6 mRNA expressing γδ T cells were only found in HOA and v) there was a tendency of higher TNF-α mRNA levels in non-T cells of CIA compared to HOA. The following scenarios emerge: in CIA there appears to be an inadequate first line of defence, with a low frequency of intraepithelial γδ T cells and a high frequency of cytotoxic CD8+ αβ T cells eliminating infected epithelial cells. Togehter, these two conditions cause a "leaky" epithelium, allowing infiltration of microbes into the underlying tissue and subsequent recruitment of granulocytes and follicle formation initiated by activated macrophages. In HOA, activated intraepithelial γδ T cells appear to be involved in antimicrobial defence reactions and surveillance of the epithelium.The difference in leukocyte profiles between tonsils from patients subjected to surgery due to idiopathic tonsillar hypertrophy or recurrent tonsillitis was limited to the surface epithelium. CD8+ γδ T cells utilising the unusual combination Vδ1/Vγ9 in their T cell receptor constituted the majority of intraepithelial lymphocytes in both groups. However, the frequency of these cells was significantly higher in recurrent tonsillitis. These results suggest that CD8+ Vδ1/Vγ9+ γδ T cells are characteristic of palatine tonsils and selectively expanded in recurrent tonsillitis. These γδ T cells may be involved in clearing infectious bacteria at the surface of the tonsil.Tissue macrophages, αβ T cells, γδ T cells, mast cells and B cells constituted, in declining order, the immune cell populations in the uvula. No fillicle-like structures were present. Most T cells had a CD8+ CD28-TCR-αβ+ phenotype, suggesting a down-regulatory function. Production of the down-regulatory cytokine TGF-β was also noted. This is consistent with the hypothesis that the uvula contributes to the development of mucosal tolerance. Furthermore, the uvula seems to be protected from pathogens penetrating the internal milieu by a subepithelial barrier of γδ T cells and macrophages. TNF-α secreting immune cells were found at this location. TNF-α and TGF-β may cause tissue fibrosis, TNF-α indirectly by stimulating mast cells to release histamine. Tissue fibrosis in conjunction with water binding to hyaluronan present in the connective tissue is the most likely explanation for the observed enlargement of the uvula in patients with sleeping disorders.
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