Yosuke Asada

Cell shape and growth of budding yeast cells in restrictive microenvironments

Effects of limited growth space on the cell morphology and cell growth are investigated by creating rigid outside environments. The cube‐shaped holes big enough for a single cell of the budding yeast Saccharomyces cerevisiae were prepared with a focused ion‐beam (FIB), commonly used for processing semiconductors. We demonstrated that the outline of the cells changes their ellipsoidal morphology into a cubic form when the daughter cells are grown in the holes, indicating that yeast cells change their shape in response to external limited space. The yeast cells grown in the microenvironments exhibit neither bud formation nor nuclear division. Although restricted growth caused by the physical barriers leads to the block of cell cycle progression in the wild‐type cells, swe1Δ cells defective in the morphogenesis checkpoint become binucleate after being grown in the microenvironments. These results suggest that yeast cells under spatial restriction arrest cell cycle progression in a Swelp‐dependent manner. Copyright

Roles of Epithelial Cell-Derived Type 2-Initiating Cytokines in Experimental Allergic Conjunctivitis.

PURPOSE To clarify the possible involvement of the type 2-initiating cytokines interleukin (IL)-25, IL-33, and thymic stromal lymphopoietin (TSLP) in the pathophysiology of allergic conjunctivitis, we evaluated ragweed (RW)-induced experimental allergic conjunctivitis (EAC) models by using IL-25 knockout (KO), IL-33 KO, and TSLP receptor (TSLPR) KO mice. METHODS Interleukin-25 KO, IL-33 KO, TSLPR KO, and BALB/c wild-type mice were sensitized twice with RW in alum and then challenged with RW in eye drops. Clinical scores and eosinophil infiltration were evaluated. Expression levels of serum immunoglobulin E (IgE) and cytokines in the conjunctival tissues were quantified and immunohistochemical analysis was carried out. RESULTS Significant reductions in clinical scores and numbers of infiltrating eosinophils were observed in the RW-EAC model using IL-33 KO mice. There were no significant differences in clinical scores and numbers of infiltrating eosinophils among IL-25KO, TSLPR KO, and wild-type mice. Serum IgE concentration was upregulated after RW challenges, and there were no differences among the mouse genotypes. Expression levels of of il4, il5, il13, and ccl5 mRNA were diminished in the conjunctivae of the RW-EAC model using IL-33 KO mice compared to those in wild-type mice. Interleukin-33 expression was upregulated as early as 1 hour after RW eye-drop challenge. The number of infiltrating basophils in the conjunctivae of the RW-EAC model using IL-33 KO mice was diminished compared to that in wild-type mice. CONCLUSIONS Among the type 2-initiating cytokines, IL-33 may play a major role in conjunctival inflammation in an RW-EAC model.

Interaction Between Conjunctival Epithelial Cells and Mast Cells Induces CCL2 Expression and Piecemeal Degranulation in Mast Cells

PURPOSE Intraepithelial mast cells are observed in giant papillae tissue samples obtained from patients with atopic keratoconjunctivitis (AKC)/vernal keratoconjunctivitis (VKC). We examined the roles of interaction between the conjunctival epithelial cells and mast cells. METHODS The interaction between human mast cells and conjunctival epithelial cells (HCjE) was investigated using a coculture model. Protein array analysis, ELISA, and real-time PCR were performed to test the interaction. Tissue samples (n = 6) from giant papillae were resected for therapeutic purposes, and subjected to immunohistological analysis of CCL2 expression. Recombinant CCL2 (10 ng/mL) was reacted with the cultured human mast cells and ultrastructural analysis was performed. A ragweed (RW)-induced mouse experimental allergic conjunctivitis model was used to examine ccl2 mRNA expression and mast cell morphology. RESULTS Protein array and real-time PCR analyses showed that CCL2 protein/mRNA expression was induced by mast cell-HCjE coculture. Upregulation of CCL2 mRNA was observed in mast cells, whereas in situ CCL2 expression was observed at the conjunctival epithelium of the giant papillae by immunohistochemistry. Ultrastructural analysis showed that recombinant CCL2 treatment induced piecemeal degranulation (PMD) in the mast cells. Ultrastructural analysis of tissues from the giant papillae showed PMD of mast cells within the conjunctival epithelial cells. The RW-induced experimental allergic conjunctivitis model showed increased ccl2 mRNA expression and PMD morphology in the conjunctivae. CONCLUSIONS Mast cell-conjunctival epithelial cell interaction induces CCL2 expression and subsequent PMD.

Contributions of Interleukin-33 and TSLP in a papain-soaked contact lens-induced mouse conjunctival inflammation model

Pathological changes of severe chronic allergic conjunctivitis are driven not only via acquired immunity but also via innate immunity. Type 2 immune response‐initiating cytokines may play some roles as innate immunity‐dependent components of the ocular surface inflammation. To investigate the involvement of type 2 immune response‐initiating cytokines in innate immunity‐dependent, papain‐induced conjunctival inflammation model using IL‐25‐, IL‐33‐, and TSLP receptor (TSLPR)‐knockout (KO) mice with reference to basophils and ILC2.

Filaggrin mutations are not associated with chronic allergic keratoconjunctivitis

Filaggrin gene mutations are predisposing genetic factors for atopic dermatitis (AD).1 In this study, we investigated the association between atopic keratoconjunctivitis (AKC)/vernal keratoconjunctivitis (VKC) and filaggrin gene mutations using a Japanese population. Detailed information on the diagnostic criteria for AKC and VKC was provided elsewhere.2 The mean age of VKC patients was 15.7±8.9 years (7–32 years), that of AKC patients 28.34±9.9 years (6–50 years) and for controls 42.7±24.3 years (23–67 years). The male:female ratios were 3.2:1.0 for VKC, 3.9:1.0 for AKC and 1:1 for controls. Four common filaggrin gene mutations in the Japanese population (S2554X, S2889X, S3296X, 3321delA)3 were genotyped using DNA samples obtained from 69 cases (AKC 44; VKC 25) enrolled in this study. Ninety-three DNA samples obtained from healthy controls (from Health Science Research Resources Japan, Osaka, Japan) were also …

Vernal keratoconjunctivitis with giant papillae on the inferior tarsal conjunctiva.

Purpose: In vernal keratoconjunctivitis (VKC), giant papillae are commonly observed on the superior tarsal conjunctiva. We found 3 cases of giant papillae on the inferior tarsal conjunctiva, and diagnosed them as being VKC based on their clinical and histopathological features. Methods: Three patients with inferior tarsal giant papillae were studied. In 2 patients, the giant papillae were resected for therapeutic purposes. Immunohistochemical analysis was carried out by indirect immunofluorescent staining using anti-CD3, anti-CD20, anti-CD35 antibodies. Results: In all 3 patients, giant papilla formation was observed on the inferior lid margin. Clusters of CD20+ B lymphocytes with CD35+ follicular dendritic cells, and CD3+ marginal zone T lymphocytes, common features of lymphoid neogenesis, were observed. In 2 patients, typical giant papillary formation was also observed on the superior tarsal conjunctiva. In all the patients, topical dexamethasone and tacrolimus treatments were found to be effective. Conclusions: The giant papillae of VKC can occur not only on the superior tarsal conjunctiva but also on the inferior tarsal conjunctiva. The possibility of the presence of giant papillae on the inferior tarsal conjunctiva should be considered in the clinical examination of patients with VKC.