Evaluation of histamine-induced conjunctival oedema in guinea pigs by means of image analysis.

BACKGROUND: Conjunctival oedema is commonly observed in patients with allergic conjunctivitis and can be induced by histamine. In animal models of allergic conjunctivitis, conjunctival oedema is generally evaluated by measuring the extravasation of Evans blue dye into the conjunctiva. A limitation of this method is that it only allows evaluation at a single time point. The aim of the present study was to investigate kinetic changes in histamine-induced bulbar oedema. METHODS: Evans blue dye was injected intravenously into male guinea pigs. Histamine eye-drops were administered 30 min later. One group of animals received levocabastine (an antihistamine) eye-drops 10 min before histamine challenge. A digital camera was used to obtain images of the bulbar conjunctiva at 1 min intervals until 30 min after histamine challenge. The conjunctivas were then harvested, and the concentration of Evans blue was measured. The ImageJ software was used to analyse the images by counting the number of absolute pixel values. RESULTS: The degree of conjunctival oedema increased progressively until 20 min after histamine challenge and then stabilised. Correspondingly, the number of absolute pixel values increased significantly until 5 min after histamine challenge, then increased gradually until the 20 min time point and finally plateaued. Pixel values were significantly lower in animals treated with levocabastine than in control animals. A significant correlation was observed between the pixel values of the conjunctival images and the concentration of Evans blue in the conjunctiva. CONCLUSIONS: This is the first study to have quantitatively evaluated kinetic changes in histamine-induced bulbar oedema by means of image analysis.

Multiple elements for negative regulation of the rat catalase gene expression in dedifferentiated hepatoma cells.

Like such hepatic genes as those for albumin and aldolase B, the rat catalase gene shows markedly reduced expression in carcinogenesis of hepatocytes. Strong silencer activity has been widely observed in the 5'-flanking region of the gene, downstream from the G-rich sequence identified in a previous study. In this study, we identified and characterized multiple elements involved in negative regulation of catalase gene expression by reporter assay and gel shift assay. One of the silencer elements is located 3 kb upstream of the gene and has GATATCCCGATATC as core sequence. The observation that protein binding to the element is abundantly expressed in dedifferentiated hepatoma cell lines, but scarcely in well-differentiated cell lines suggests that this element is involved in negative regulation of the catalase gene expression in hepatocarcinogenesis. This element was targeted by a novel 20-kDa nuclear protein, which is designated HNRF (hepatocarcinogenesis-related negative regulatory factor).