No abstract

While the extravasation cascade of lymphocytes is well characterized, data on their intraepithelial positioning and morphology are scant. However, the latter process is presumably crucial for many immune functions. Integrin ␣ E (CD103)␤ 7 has previously been implicated in epithelial retention of some T cells through binding to E-cadherin. Our current data suggest that ␣ E (CD103)␤ 7 also determines shape and motility of some lymphocytes. Time-lapse microscopy showed that wild-type ␣ E (CD103)␤ 7 conferred the ability to form cell protrusions/filopodia and to move in an amoeboid fashion on E-cadherin, an activity that was abrogated by ␣ E (CD103)␤ 7 -directed antibodies or cytochalasin D. The ␣ E -dependent motility was further increased (P < .001) when point-mutated ␣ E (CD103) locked in a constitutively active conformation was expressed. Moreover, different yellow fluorescent protein-coupled ␣ E (CD103) species demonstrated that the number and length of filopodia extended toward purified E-cadherin, cocultured keratinocytes, cryostat-cut skin sections, or epidermal sheets depended on functional ␣ E (CD103). The in vivo relevance of these findings was demonstrated by wild-type dendritic epidermal T cells (DETCs), which showed significantly more dendrites and spanned larger epidermal areas as compared with DETCs of ␣ E (CD103)-deficient mice (P < .001). Thus, integrin ␣ E (CD103)␤ 7 is not only involved in epithelial retention, but also in shaping and proper intraepithelial morphogenesis of some leukocytes. IntroductionPositioning and locomotion of leukocytes within tissues provide the basis for the molecular crosstalk with other cells and are prerequisites for a functional immune system. To date, the recruitment cascade of initial endothelial adhesion, activation, firm adhesion, transmigration, and subsequent localization into the connective tissue is one of the best established concepts in leukocyte biology. 1,2 However, many effector functions of immunocytes are exerted within parenchymatous organs, mostly epithelia. It is therefore somewhat surprising that we know relatively little about locomotion and function-determining morphogenesis of lymphocytes within epithelial tissues, such as the epidermis of the skin. 3 An adhesion receptor that is thought to mediate retention of lymphocytes within epithelial tissues is integrin ␣ E (CD103)␤ 7 . 4 First described 2 decades ago as a selective marker for intestinal intraepithelial lymphocytes, 5 ␣ E (CD103)␤ 7 has been implicated in epithelial T-cell retention through binding to E-cadherin. 6,7 Indeed, ␣ E (CD103)-deficient mice exhibited a reduced number of mucosal intraepithelial T cells. 8 However, ␣ E (CD103)␤ 7 has later been found to be also expressed by some lymphocytes within other epithelia, such as the epidermis of the skin, 9 where it presumably contributes to recruitment of T cells in inflamed human skin 10 as well as dendritic epidermal T cells (DETCs) in murine skin. 11 Thymic DETC precursor cells express integrin ␣ E (CD103)␤ 7 before their migration ...

Immortalization should overcome the problem of short lifespan and difficult culture of endothelial cells that limited their use in functional studies. We used four different immortalized endothelial cell lines to study dynamic interactions with lymphocytes. Surprisingly, tumor necrosis factor (TNF)alpha-stimulated human umbilical vein endothelial cells (HUVECs) or human dermal microvascular endothelial cells (HDMECs) readily supported rolling and binding of lymphocytes, whereas none of the immortalized cell lines did. As rolling interactions are primarily mediated by selectins and vascular cell adhesion molecule (VCAM)-1, the endothelial cells were analyzed regarding expression of selectins and other adhesion molecules. Interestingly, cell surface expression of E-selectin could only be detected on HUVEC and HDMEC. Immunocytochemistry showed that some immortalized endothelial cells expressed E-selectin intracellularly following TNFalpha stimulation, suggesting translation but defective post-translational processing or transport of the molecule. In contrast, other immortalized cell lines did not have detectable levels of E-selectin mRNA, suggesting impaired transcription. VCAM-1 could only be induced on normal and human placental microvascular endothelial cell-A2 endothelial cells, whereas all cell lines expressed intercellular adhesion molecule-1 following TNF stimulation. The immortalized endothelial cells tested here have lost functions that are required for dynamic interactions with immune cells and that are common to primary endothelial cells.

T-cells expressing a E (CD103), an integrin induced by TGFb on T-cells in vitro, accumulate within epithelia in inflammatory disorders, including psoriasis. However, it is unclear, if and how a E (CD103) contributes to skin inflammation. Using two complementary approaches, we have investigated a E (CD103) in psoriasis-like skin inflammation of mice with transgenic epidermal expression of human TGFb1: a E (CD103) was inhibited by function-blocking antibodies in vivo, and doublemutants with additional a E (CD103)-depletion were generated in two different genetic backgrounds. Epidermal hTGFb1 expression was associated with prominent expression of a E (CD103) on infiltrating cells. However, neither treatment with a E (CD103)-blocking antibodies nor deficiency of a E (CD103) in doublemutant mice altered the psoriasis-like phenotype. In addition, histopathological and flow cytometric analyses revealed similar pathological skin alterations and lymphocyte subgroups in the different mouse strains. Thus, while a E (CD103) expression is indeed associated with hTGFb1 in vivo, it has little, if any, influence on the course of the psoriasis-like phenotype in K5.hTGFb1 transgenic mice.