Crosstalk between TGF-beta and MAPK signaling during corneal wound healing.

PURPOSE: The aim of this study was to elucidate the mechanisms governing epithelial cell migration and proliferation during wound healing. METHODS: The authors used wound healing of mouse corneal epithelium to examine the role TGF-ß signaling plays during the healing process. To achieve this goal, they used transgenic mice in which the TGF-ß receptor type II (Tbr2) was conditionally ablated from the corneal epithelium. Epithelium debridement wounds were made, followed by the assessment of cell migration, proliferation, and immunostaining of various signaling pathway components. RESULTS: The authors showed that in the absence of TGF-ß signaling corneal epithelial wound healing is delayed by 48 hours; this corresponds to a delay in p38MAPK activation. Despite the delayed p38MAPK activation, ATF2, a substrate of p38MAPK, is still phosphorylated, leading to the suppression of cell proliferation at the leading edge of the wound. These data provide evidence that in the absence of TGF-ß signaling, the suppression of cell proliferation during the early stages of wound healing is maintained through the JNK activation of ATF2. CONCLUSIONS; Together the data presented here demonstrate the importance of the TGF-ß and MAPK signaling pathways in corneal epithelial wound healing.

The role of interleukin-33 in chronic allergic conjunctivitis.

PURPOSE: The authors discovered a genetic association between the ST2L gene and atopy. The ST2L gene encodes a membrane-bound functional marker for Th2 cells. Recently, a novel Th2 cytokine, interleukin-33 (IL-33), was discovered to be a specific ligand for ST2L. The authors investigated the role of IL-33 in chronic allergic conjunctivitis. METHODS: Immunohistochemical analysis was carried out using giant papillae samples obtained from patients with atopic keratoconjunctivitis. The authors used proinflammatory stimuli to clarify IL-33 mRNA/protein-inducing signals with cultured human conjunctival epithelial cells, fibroblasts, human umbilical vascular endothelial cells, and mast cells. These cells were also used to examine the expression of ST2L (IL-33R). Finally, cultured mast cells were stimulated with recombinant IL-33 (rIL-33) to examine the downstream signals. RESULTS: The authors found IL-33 protein expression in human vascular endothelial cells in the giant papillae and in the control conjunctivae. IL-33 expression was also observed in conjunctival epithelium of the giant papillae but not in the control conjunctivae. IL-1 beta stimulation upregulated IL-33 mRNA expression in conjunctival fibroblasts. The authors also confirmed mature IL-33 protein expression in ocular resident cells by Western blot analysis. Preferential ST2L expression was observed in human mast cells, and phosphorylation of p38 MAPK and IL-13 mRNA induction was observed in human cultured mast cells after rIL-33 stimulation. Phosphorylation of p38 MAPK was inhibited by soluble ST2 protein. CONCLUSIONS: The IL-33-ST2 signaling cascade plays some roles in the pathophysiology of chronic allergic conjunctivitis through the activation of mast cells.

Expression of keratin 12 and maturation of corneal epithelium during development and postnatal growth.

PURPOSE: To determine the kinetics of corneal epithelial maturation during embryonic development and postnatal growth. METHODS: Expression patterns of keratin (K)12 and K14 were determined in mouse embryos (embryonic days [E]15.5-19.5), corneas of postnatal day (P)0 to 10 months, and healing corneas after epithelial debridement in P30 and P90 mice. The expression of alkaline phosphatase (AP) was determined during postnatal growth and healing of epithelial debridement of Krt12(Cre/Cre)/ZAP bitransgenic mice. RESULTS: During embryonic development, K12 expression by corneal peridermal epithelium commenced at E15.5. In the period from E15.5 to P10, the expression of K12 was restricted to the suprabasal and/or superficial cells of the corneal epithelium, whereas the K14 expression was restricted to the basal cells. After P30, K12 expression was sporadically detected in the basal corneal epithelium, and the number of K12-positive basal cells increased as the mice grew older. The number of K14-positive cells that coexpressed K12 increased with age and reached a plateau after P180. Healing of the debrided epithelium facilitated the increase in K14-positive cells that coexpressed K12. Many basal cells of Krt12(Cre/Cre)/ZAP mice remained undifferentiated and expressed LacZ at P15, and they then differentiated to express Cre, which leads to excision of LacZ and AP expression. CONCLUSIONS: In the mouse, the corneal epithelium does not become fully mature until 3 to 6 months after birth, in that a significant number of corneal basal epithelial cells of young mice (

Characterization of tetracycline-inducible bitransgenic Krt12rtTA/+/tet-O-LacZ mice.

PURPOSE: To prepare binary transgenic mouse lines that overexpress reporter genes in a corneal-epithelium-specific manner when induced by doxycycline. METHODS: A gene-targeting construct containing an internal ribosomal entry site-reverse tetracycline transcription activator (IRES-rtTA) cassette was inserted into the Krt12 allele (keratin 12 gene) to produce a knock-in Krt12(rtTA/+) mouse line through gene-targeting techniques. The Krt12(rtTA/+) knock-in mice were bred with tet-O-LacZ reporter mice to obtain Krt12(rtTA/+)/tet-O-LacZ bitransgenic mice. The expression of the LacZ gene was induced in bitransgenic mice by administration of doxycycline in the drinking water and chow. RESULTS: Administration of doxycycline induced a 15-fold increase of beta-galactosidase activity in the cornea of adult bitransgenic mice (Krt12(rtTA/+)/tet-O-lacZ). Administration of doxycycline either to single transgenic Krt12(rtTA/+) or tet-O-LacZ mice as a control did not induce overexpression of LacZ as it did in the bitransgenic mice. The induction of beta-galactosidase enzyme activity by doxycycline in bitransgenic mice took place in 24 hours and reached a plateau by 2 days. Histochemical analysis also showed that beta-galactosidase induction was limited to the corneal epithelium of bitransgenic mice fed doxycycline. The increased beta-galactosidase activity in corneal epithelium caused by doxycycline returned to basal levels in 4 weeks after the antibiotics were omitted from the diet. CONCLUSIONS: A binary mouse model has been successfully established that conditionally overexpresses reporter genes in corneal epithelium. This mouse model will be useful in elucidating signaling pathways of various growth factors and cytokines and gene functions in the maintenance of homeostasis and pathogenesis in the adult mouse cornea.

[Case report revisited: review of the 6 cases].

Out of the cases we experienced in our 11-year service in sanatorium, 6 cases were selected to review the medical care and social environment that each patient was involved. Two cases were the residents in a sanatorium and 4 cases were in the community, including 2 cases having foreign nationality. The review of these cases drew the following conclusions. 1. We must be aware of our responsibility for early diagnosis and treatment of leprosy to prevent tragic disability. 2. The fixed duration of MDT/MB may not be enough for LL cases having high bacterial load before treatment. Enough duration of chemotherapy and follow-up is desired to prevent avoidable disability. 3. Basically, the treatment of leprosy should be carried on in outpatient clinic. The duration of hospitalization, if necessary, should be shortest to enhance patient's motive for treatment. 4. Intermittent administration of RFP must be done under direct observation. 5. For foreign patients not accustomed to the life in Japan or elderly patient living by oneself, various supports from community are greatly helpful to achieve the long course of leprosy treatment. Through these supports, we can expect community enlightening that may promote rehabilitation of the people once suffered from leprosy.