Abstract
Specific heterodimers of α and β integrins are implicated in mediating adhesion and functional activation of mast cells to extracellular matrix (ECM) proteins, determining thus homing, secretion and tissue distribution of these cells. In the present study, we have examined integrin expression and associated morphological features of mast cells adhering to ECM, also depending on cell activation and under the influence of protein kinase C (PKC) inhibitors. Unstimulated and PMA-activated human leukaemic mast cells (HMC-1 line) were allowed to adhere to fibronectin or vitronectin-coated surfaces. Cells were specifically stained for actin, β1, α1–α6, αv and αvβ5 integrins and were evaluated by fluorescence microscopy and confocal laser scan microscopy. Spontaneously adhering cells rapidly assumed an oblong shape, with pronounced formation of filopodia, whereas PMA-stimulated cells were round in shape. Clustering of integrins on filopodia and on comma-like shapes at the cell circumference in rounded cells was noted only for α4, α5 and β1 chains in fibronectin-adhering cells, and for αv and αvβ5 chains in vitronectin-adhering cells. On double staining, clustered integrins co-localized with each other and with actin at the cell membrane and along intracellular tension lines of actin filaments. PKC inhibitors affected the shape of cells, but adhesion was maintained. These data provide a morphological correlate to previously reported functional studies, demonstrating clustering of selected integrins during ECM adhesion at the cell membrane. This was associated with alignment of integrins along actin filaments within the cytoplasm, PKC signalling and changes in shape and activation of mast cells.
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Butterfield JH, Weiler D, Dewald G, Gleich GJ (1988) Establishment of an immature mast cell line from a patient with mast cell leukemia. Leuk Res 12:345–355
Columbo M, Bochner BS (2001) Human skin mast cells adhere to vitronectin via the α vβ3 integrin receptor (CD51/CD61). J Allergy Clin Immunol 107:554
Columbo M, Bochner BS, Marone G (1995) Human skin mast cells express functional beta 1 integrins that mediate adhesion to extracellular matrix proteins. J Immunol 154:6058–6064
Dastych J, Wyczolkowska J, Metcalfe DD (1991) Mast cell adhesion to fibronektin. Immunology 73:478–484
Davis PD, Hill CH, Keech E, Lawton G, Nixon JS, Sedgwick AD, Wadsworth J, Westmacott D, Wilkinson SE (1989) Potent selective inhibitors of protein kinase C. FEBS Lett 259:61–63
Furitsu T, Tsujimura T, Tono T, Ikeda H, Kitayama H, Koshimizu U, Sugahara H, Butterfield JH, Ashman LK, Kanayama Y (1993) Identification of mutations in the coding sequence of the proto-oncogene c-kit in a human mast cell leukaemia cell line causing ligand-independent activation of c-kit product. J Clin Invest 92:1736–1744
Gascoigne MH, Holland K, Page CP, Shock A, Robinson M, Foulkes R, Gozzard N (2003) The effect of anti-integrin monoclonal antibodies on anitigen-induced pulmonary inflammation in allergic rabbits. Pulm Pharmacol Ther 16:279–285
Grützkau A, Kögel H, Krüger-Krasagakes S, Möller A, Henz BM (1997) Detection of intracellular interleukin-8 in human mast cells: flow cytometry as a guide for immunoelectron microscopy. J Histochem Cytochem 45:935–945
Gurish MF, Tao H, Arya A, Friend DS, Parker CM, Austen KF (2001) Intestinal mast cell progenitors require CD49dbeta7 (alpha4beta7 integrin) for tissue-specific homing. J Exp Med 194:1243–1252
Hamann K, Grabbe J, Welker P, Haas N, Algermissen B, Czarnetzki BM (1994) Phenotypic evaluation of cultured human mast and basophilic cells and of normal human skin mast cells. Arch Dermatol Res 286:380–385
Henz BM, Maurer M, Lippert U, Worm M, Babina M (2001) Mast cells as initiators of immunity and host defense. Exp Dermatol 10:1–10
Hynes RO (1992) Integrins: versatility, modulation, and signaling in cell adhesion. Cell 69:11–25
Issekutz TB, Palecanda A, Kadela-Storlarz U, Marshall JS (2001) Blockade of either alpha-4 or beta-7 integrins selectively inhibits intestinal mast cell hyperplasia and worm expulsion in response to Nippostrongylus brasiliensis infection. Eur J Immunol 31:860–868
Johnson GD, Davidson RS, McNamee KC, Russell G, Goodwin D, Holborow EJ. (1982) Fading of immunofluorescence during microscopy: a study of the phenomenon and its remedy. J Immunol Methods 55:231–242
Kinashi T, Asaoka T, Setoguchi R, Takatsu K (1999) Affinity modulation of very late antigen-5 through phosphatidylinositol 3-kinase in mast cells. J Immunol 162:2850–2857
Krüger-Krasagakes S, Grützkau A, Krasagakes K, Hoffmann S, Henz BM (1999) Adhesion of human mast cells to extracellular matrix provides a co-stimulatory signal for cytokine production. Immunology 98:253–257
Krüger-Krasagakes S, Grützkau A, Baghramian R, Czarnetzki BM (1996) Interactions of immature human mast cells with extracellular matrix: expression of specific adhesion receptors and their role in cell binding to matrix proteins. J Invest Dermatol 106:538–543
Longhurst CM, Jennings LK (1998) Integrin-mediated signal transduction. Cell Mol Life Sci 54:514–526
Lorentz A, Schuppan D, Gebert A, Manns MP, Bischoff SC (2002) Regulatory effects of stem cell factor and interleukin-4 on adhesion of human mast cells to extracellular matrix proteins. Blood 99:966–972
Metcalfe DD, Baram D, Mekori YA (1997) Mast cells. Physiol Rev 77:1033–1079
Nishizuka Y (1984) The role of protein kinase C in cell surface signal transduction and tumor promotion. Nature 308:693–698
Ruegg UT, Burgess GM (1989) Staurosporine, K-252 and UNC-01: potent but nonspecific inhibitors of protein kinases. Trends Pharmacol Sci 10:218–220
Springer TA (1990) Adhesion receptors of the immune system. Nature 346:425–34
Tachimoto H, Hudson SA, Bochner BS (2001) Acquisition and alteration of adhesion molecules during cultured human mast cell differentiation. J Allergy Clin Immunol 107:302–309
Tan BL, Yazicioglu MN, Ingram D, McCarthy J, Borneo J, Williams DA, Kapur R (2003) Genetic evidence for convergence of c-Kit- and alpha4 integrin-mediated signals on class IA PI-3kinase and the Rac pathway in regulating integrin-directed migration in mast cells. Blood 101:4725–4732
Vuori K, Ruoslahti E (1993) Activation of protein kinase C precedes α5β1 integrin-mediated cell spreading on fibronectin. J Biol Chem 268:21459–21462
Weber S, Babina M, Krüger-Krasagakes S, Grützkau A, Henz BM (1996) A subclone (5C6) of the human mast cell line HMC-1 represents a more mature phenotype than the original cell line. Arch Dermatol Res 288:778–782
Wimazal F, Ghannadan M, Muller MR, End A, Willheim M, Meidlinger P, Schernthaner GH, Jordan JH, Hagen W, Agis H, Sperr WR, Czerwenka K, Lechner K, Valent P (1999) Expression of homing receptors and related molecules on human mast cells and basophils: a comparative analysis using multi-color flow cytometry and toluidine blue/immunofluorescence staining techniques. Tissue Antigens 54:499–507
Wulf E, Deboben A, Bautz FA, Faulstich H, Wieland T (1979) Fluorescent phallotoxin, a tool for the visualization of cellular actin. Proc Natl Acad Sci USA 76:4498–4502
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This work was supported in part by the Pythagoras II program by the Greek Ministry of Education and the European Social Fund.
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Küchler, J., Grützkau, A., Henz, B.M. et al. Morphological analysis of integrin-mediated adhesion of immature human mast cells to extracellular matrix proteins. Arch Dermatol Res 298, 153–161 (2006). https://doi.org/10.1007/s00403-006-0676-y
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DOI: https://doi.org/10.1007/s00403-006-0676-y