Kazumasa Morita

Characterization of ZO-2 as a MAGUK Family Member Associated with Tight as well as Adherens Junctions with a Binding Affinity to Occludin and α Catenin

ZO-2, a member of the MAGUK family, was thought to be specific for tight junctions (TJs) in contrast to ZO-1, another MAGUK family member, which is localized at TJs and adherens junctions (AJs) in epithelial and nonepithelial cells, respectively. Mouse ZO-2 cDNA was isolated, and a specific polyclonal antibody was generated using corresponding synthetic peptides as antigens. Immunofluorescence microscopy with this polyclonal antibody revealed that, similarly to ZO-1, in addition to TJs in epithelial cells, ZO-2 was also concentrated at AJs in nonepithelial cells such as fibroblasts and cardiac muscle cells lacking TJs. When NH2-terminal dlg-like and COOH-terminal non-dlg-like domains of ZO-2 (N-ZO-2 and C-ZO-2, respectively) were separately introduced into cultured cells, N-ZO-2 was colocalized with endogenous ZO-1/ZO-2, i.e. at TJs in epithelial cells and at AJs in non-epithelial cells, whereas C-ZO-2 was distributed along actin filaments. Consistently, occludin as well as α catenin directly bound to N-ZO-2 as well as the NH2-terminal dlg-like portion of ZO-1 (N-ZO-1) in vitro. Furthermore, immunoprecipitation experiments revealed that the second PDZ domain of ZO-2 was directly associated with N-ZO-1. These findings indicated that ZO-2 forms a complex with ZO-1/occludin or ZO-1/α catenin to establish TJ or AJ domains, respectively.

Tight junctions in Schwann cells of peripheral myelinated axons: a lesson from claudin-19–deficient mice

Tight junction (TJ)–like structures have been reported in Schwann cells, but their molecular composition and physiological function remain elusive. We found that claudin-19, a novel member of the claudin family (TJ adhesion molecules in epithelia), constituted these structures. Claudin-19–deficient mice were generated, and they exhibited behavioral abnormalities that could be attributed to peripheral nervous system deficits. Electrophysiological analyses showed that the claudin-19 deficiency affected the nerve conduction of peripheral myelinated fibers. Interestingly, the overall morphology of Schwann cells lacking claudin-19 expression appeared to be normal not only in the internodal region but also at the node of Ranvier, except that TJs completely disappeared, at least from the outer/inner mesaxons. These findings have indicated that, similar to epithelial cells, Schwann cells also bear claudin-based TJs, and they have also suggested that these TJs are not involved in the polarized morphogenesis but are involved in the electrophysiological “sealing” function of Schwann cells.

Expression patterns of claudins, tight junction adhesion molecules, in the inner ear

Tight junctions (TJs) are indispensable for the establishment of compositionally distinct fluid compartments in the inner ear, but our knowledge of the claudins, TJ adhesion molecules, in the inner ear is still fragmentary. We examined the expression and distribution of claudin-1 to claudin-18 (except for claudin-7, -13 and -17) in the inner ear by immunofluorescence microscopy. In the cochlea, the organ of Corti expressed claudin-1, -2, -3, -9, -10, -12, -14 and -18. In the stria vascularis, claudin-1, -2, -3, -8, -9, -10, -12, -14 and -18 were expressed in the marginal cells, whereas the basal cells were positive only for claudin-11. In Reissners membrane and the spiral limbus, the expression of claudin-1, -2, -3, -8, -9, -10, -12, -14 and -18 was detected. Furthermore, in the vestibule, claudin-1, -3, -9, -12, -14 and -18 were expressed in the sensory epithelia, whereas in the dark cell area claudin-1, -3, -8, -9, -12, -14 and -18 were detectable. These findings, i.e., very complex expression patterns of claudin species in the inner ear, would reflect the importance and the complexity of the barrier function of TJs in the inner ear.

FRACTALKINE AND MACROPHAGE-DERIVED CHEMOKINE : T CELL-ATTRACTING CHEMOKINES EXPRESSED IN T CELL AREA DENDRITIC CELLS

Dendritic cells (DC) are a system of antigen‐presenting cells specialized in interaction with T cells. Recently it has been reported that DC can produce CC (β) chemokines that attract T cells. In this study we isolated mouse fractalkine and macrophage‐derived chemokine (MDC) belonging to CX3C (δ) and CC chemokine families, respectively, from bone marrow‐derived mature DC. While expression of fractalkine, which has so far been only examined in the brain and in vitro endothelial cells so far, was rather ubiquitous, MDC, which has been reported to be synthesized by macrophages and DC, was expressed specifically in the thymus and lymphnode. This is the first report that indicates fractalkine expression by DC. Expression of fractalkine and MDC mRNA increased with maturation of DC during in vitro culture of bone marrow cells. Spleen‐ and epidermis‐derived mature DC in culture also expressed these chemokines. Furthermore, their expression was detected selectively by Northern hybridization in CD11c+ B220– DC freshly purified from lymph nodes, and in large stellate cells in the lymph node T cell areas by in situ hybridization. Conditioned media of 293T cells transfected with these chemokine cDNA were chemotactic to Con A‐activated splenic T cells as well as the mouse T cell line EL4. In conclusion, while fractalkine and MDC belong to different families of chemokines, both may be involved in recruitment of T cells for interaction with mature DC in the immune response.

Tight junction‐associated proteins (occludin, ZO‐1, claudin‐1, claudin‐4) in squamous cell carcinoma and Bowen's disease

Background  The epidermis, which is a typical stratified epithelium, has tight junctions (TJs) in the granular layer, as do simple epithelia. So far, abnormalities of TJs and involvement of claudin‐1 have been reported in tumours of simple epithelia.

Altered expression of occludin and tight junction formation in psoriasis.

Abstract In simple epithelia, tight junctions are well developed and have barrier and fence functions. On the other hand, tight junctions are less developed in stratified epithelia. In the rodent epidermis, only maculae occludentes (i.e. focal strands or spot tight junctions) are observed in the most superficial zone of the granular cell layer. Occludin is an integral membrane protein, and is localized at tight junctions in simple epithelia. In normal epidermis, occludin is expressed at the maculae occludentes in the granular cell layer, indicating that it is associated with keratinocyte differentiation. Thus, we examined occludin expression in psoriasis, in which differentiation of keratinocytes is impaired. In psoriasis, occludin was expressed more broadly in the upper epidermis than in normal epidermis. In addition, immunoelectron microscopy showed occludin to be concentrated on the maculae occludentes in the spinous layer of psoriatic skin. These findings indicate that occludin and the formation of tight junctions are related to the proliferation and differentiation of keratinocytes, and to the pathogenesis of psoriasis.

Tight junction proteins in keratinocytes: localization and contribution to barrier function

Abstract:  Recent research suggests that tight junctions (TJs) are located in the stratum granulosum, where they contribute to the barrier function of the epidermis. In this study, we investigated the formation of functional TJs in cultured normal human epidermal keratinocytes. We observed the development of permeability barrier function through the process of Ca2+‐induced differentiation. Immunofluorescence analyses at 96 h after Ca2+‐induced differentiation revealed concentrated portions of occludin, a TJ‐specific marker, arranged as continuous lines circumscribing individual flattened suprabasal cells in areas with high concentrations of claudin‐1 and ‐4. Transient Ca2+ depletion reversibly disrupted the continuous network of TJ proteins and the permeability barrier. We also found that the addition of ochratoxin A weakened the permeability barrier and the expression of claudin‐4. Our findings suggest that TJ proteins contribute to the permeability barrier in epidermal keratinocytes.

Expression of claudin‐5 in dermal vascular endothelia

Abstract: Claudins and occludin are integral membrane proteins at tight junctions (TJs). We examined subcellular localization of claudin‐5 and occludin in dermal vascular endothelia. Immunofluorescence staining showed that claudin‐5 was expressed at the cell–cell border of dermal vascular endothelia in mouse skin. However, in some dermal vessels, claudin‐5 expression was markedly decreased or absent in amount by double‐immunofluorescence stainings with PECAM‐1 and PAL‐E. In contrast, occludin was not detected in dermal vessels. Freeze‐fracture and immunoreplica electron microscopy on primary‐cultured human dermal endothelial cells showed that claudin‐5 was localized at tight junctions. These findings confirmed that TJs in dermal vascular endothelial cells are composed of claudin‐5.

Tight junctions in the skin.

Tight junctions (=zonulae occludentes, TJs) function as an effective barrier in simple epithelia. Recent developments in the molecular biology of TJs revealed that TJs also exist in the stratum granulosum and contribute to barrier function in epidermis. Furthermore, several TJ-related junctions were identified in epidermis. In this review article, the history of the investigation into TJs in epidermis and the perspectives of investigation into TJs in dermatology are described.

Atopic retinal detachment. Report of four cases and a review of the literature

Summary Ocular complications of atopic dermatitis include cataract, blepharitis, keratoconjunctivitis, keratoconus, iritis and retinal detachment. The aim of this study was to evaluate the characteristics of retinal detachment in atopic dermatitis patients. We examined four patients with atopic dermatitis and retinal detachment, and performed an extensive review of the literature. There have been about 130 reported cases of retinal detachment in patients with atopic dermatitis from Japan, in comparison with only a few reports from Europe and the U.S.A. An extensive review of the literature revealed that retinal detachment occurs at a young age in atopic dermatitis patients, and that often both eyes are involved. As retinal detachment is not a rare complication of atopic dermatitis, we propose that this type of retinal detachment is designated ‘atopic retinal detachment’. Dermatologists should be aware of this potential complication of atopic dermatitis.