Histopathology in the isolated chicken eye test and comparison of different stainings of the cornea.

The isolated chicken eye (ICE) test, developed at our Institute, is accepted by the OECD for identification of severe eye irritants. The OECD ICE Guideline (No. 438) encourages preservation of the treated eyes for possible histopathology of the cornea, which is believed to strengthen evidence of absence or presence of irritation and to help clarify borderline effects by assessment of the corneal Depth-of-Injury. Histopathology of the cornea in addition to the normal slit-lamp microscope assessment of corneal effects has already been performed routinely in ICE tests at our Institute, using two standard stainings (H&E and PAS). In this study, three other stainings (AZAN, EVG and Trichrome), more specific for collagen-rich membranes such as basement- and Bowman's membranes were examined with corneas exposed to four model compounds ranging from non- to severely irritating (corrosive). PAS appeared to be the superior staining method. Surprisingly, the well-known eye corrosive sodium hydroxide (NaOH, solid) did not visibly compromise the integrity of Bowman's or the basement membrane. Based on our experience, histopathology of the treated cornea is confirmative in relation to the standard assessment of eye irritation by slit-lamp observation in the ICE and in certain cases can help to evaluate borderline effects. Besides establishing the depth of injury, additional investigation of corneal limbal stem cell damage after chemical exposure might be appropriate to determine reversibility or irreversibility of eye effects.

Do aberrant crypt foci have predictive value for the occurrence of colorectal tumours? Potential of gene expression profiling in tumours.

The effects of different dietary compounds on the formation of aberrant crypt foci (ACF) and colorectal tumours and on the expression of a selection of genes were studied in rats. Azoxymethane-treated male F344 rats were fed either a control diet or a diet containing 10% wheat bran (WB), 0.2% curcumin (CUR), 4% rutin (RUT) or 0.04% benzyl isothiocyanate (BIT) for 8 months. ACF were counted after 7, 15 and 26 weeks. Tumours were scored after 26 weeks and 8 months. We found that the WB and CUR diets inhibited the development of colorectal tumours. In contrast, the RUT and BIT diets rather enhanced (although not statistically significantly) colorectal carcinogenesis. In addition, the various compounds caused different effects on the development of ACF. In most cases the number or size of ACF was not predictive for the ultimate tumour yield. The expression of some tumour-related genes was significantly different in tumours from the control group as compared to tumours from the treated groups. It was concluded that WB and CUR, as opposed to RUT and BIT, protects against colorectal cancer and that ACF are unsuitable as biomarker for colorectal cancer. Effects of the different dietary compounds on metalloproteinase 1 (TIMP-1) expression correlated well with the effects of the dietary compounds on the ultimate tumour yield.