BACKGROUND To identify the role of survivin, a novel inhibitor of apoptosis (IAP) in colorectal tumorigenesis, the authors investigated tissue expression of survivin in human colorectal tumors including 43 hyperplastic polyps, 171 adenomas with low dysplasia, 42 adenomas with high dysplasia, and 60 carcinomas in adenoma, and examined whether the expression of survivin correlated with tumor cell apoptosis, proliferation, and angiogenesis, which is known to initiate the imbalance between cell proliferation and apoptosis. METHODS Immunohistochemical staining for the paraffin sections by using the monoclonal antibodies, survivin, p53, bcl‐2, Ki‐67, and CD34, was performed by the standard avidin‐biotin‐peroxidase technique. The apoptotic cells were detected by terminal deoxynucleotidyl transferase–mediated dUTP‐biotin nick end labeling method, using an Apop Tag in situ detection kit. RESULTS The immunoreactivity of survivin significantly increased in the transition from adenoma with low dysplasia to high dysplasia/carcinoma (P < 0.001). Similar changes in protein expression were observed for p53 but not for bcl‐2, which was expressed throughout the colorectal tumorigenesis. This transition was associated with a significant decrease in the apoptotic index (AI) and significant increases in the Ki‐67 labeling index (LI) and microvessel density (MVD; P < 0.001 for both). The expression of survivin inversely correlated with AI and was positively correlated with Ki‐67 LI and MVD (P < 0.001 for both). CONCLUSIONS These results suggest that, like p53, survivin plays an important role in transition from adenoma with low dysplasia to high dysplasia during human colorectal tumorigenesis. Cancer 2001;91:2026–32. © 2001 American Cancer Society.

We comparatively investigated the extent of apoptotic cell loss in human colorectal cancers evaluated by two methods, namely the conventional terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-biotin nick end-labeling (TUNEL) method and immunohistochemistry for single-stranded (ss) DNA. The apoptotic index (AI) obtained with the TUNEL method was higher than that shown by the immunohistochemistry for ssDNA. However, a significant correlation in AIs evaluated by these methods was found. The AIs obtained by both methods were significantly higher in the advanced cancers than in the early cancers. Cellular proliferation activity was assessed in terms of positivity rate (PR) for expression of proliferating cell nuclear antigen (PCNA). The PR of advanced cancers was significantly higher than that of early cancers. The present results indicate that immunohistochemistry for ssDNA is useful (as is the TUNEL method) for evaluation of apoptotic tumor cells in colorectal carcinomas. In addition, it was confirmed that there is a remarkable increase of not only proliferation activity, but also tumor cell apoptosis in the process of progression of colon cancer from early to advanced stages of the disease.Key words: Apoptosis -TUNEL -Immunohistochemistry -Single-stranded DNA -Colorectal cancer Apoptosis is characterized morphologically by large chromatin fragments, 1) by extensive margination and fragmentation of the chromatin viewed electromicroscopically,2) and by a characteristic "ladder formation" of DNA fragments of about 180-200 base pairs by endonuclease activation in gel electrophoresis.3)The terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-biotin nick endlabeling (TUNEL) method, which detects apoptosis and programmed cell death has been utilized to identify apoptotic cells at the single-cell level. 4) In contrast, Tomei has speculated that apoptosis involves the modification of chromatin, which results in a break-down of the supercoiling organization and the formation of individual super breaks. He has further proposed that single-stranded (ss) DNA modification in the nucleosomal linker region might constitute a critical early step in apoptosis.5) Naruse et al. showed that the antibody against ssDNA is a good marker of both drug-induced apoptosis and programmed cell death during embryogenesis. A polyclonal antibody against ssDNA has been used to immunohistochemically detect apoptotic cell death in epithelial cells. 6)There is, to our knowledge, no report on the relationship between apoptosis detected by immunohistochemistry for ssDNA and cellular proliferation activity assessed in terms of the positivity rate (PR) of proliferating cell nuclear antigen (PCNA) in colorectal carcinoma. In the present study, we investigated the extent of apoptotic cell loss in human colorectal cancers by the conventional TUNEL method and by immunohistochemistry for ssDNA. In addition, we compared the apoptotic indices (AIs) obtained by these two methods to determine whe...

Tumor cell apoptosis, proliferation and p27 expression using an in situ apoptosis detection kit, anti-Ki67 antibody and anti-p27 antibody were investigated in 171 colorectal adenocarcinoma specimens, together with the assessment of mutated p53 expression. No apparent association was observed among p27 expression, mutated p53 accumulation, the Ki67 labeling index and the apoptotic index (AI). In the multivariate analysis using the median values as the cut-off points (46.8% for p27 expression and 0.89% for AI), p27 expression was identified as an independent and significant predictor for overall survival. When the present series of patients were dichotomized by these cut-off points to categorize the cases into 4 subgroups, the patients in the group with low p27 expression and a low AI had the poorest prognosis. The assessment of p27 expression and AI therefore may prove valuable in identifying patients with colorectal adenocarcinoma who may have a poor prognosis.

SUMMARY A new concept for evaluating the pattern electroretinogram (PERG) in glaucoma and its clinical application are described. The PERG was elicited by two different stimulus fields, namely, the whole central retinal area with a radius of 180 and the paracentral ring area between the radius of 100 and 180. The amplitude of the PERG for each stimulus field and the ratio of them were analysed for 30 patients with glaucoma or ocular hypertension. Among these only a small number of eyes were detected as abnormal when we evaluated the amplitude itself either with the central stimulus or the paracentral ring stimulus, whereas the ratio of the two values was below normal in some cases of advanced glaucoma. We conclude that the ratio of the paracentral to central PERG is useful for detecting glaucoma.The pattern electroretinogram (PERG) is an electrical retinal response elicited, not by the luminance change, but by the pattern stimuli in the visual field. Since the response has been considered to originate from the ganglion cell layer in the retina,' many studies have been reported on the PERG's clinical application to glaucoma-7 as well as optic nerve disorders.=" However, most of the reports on the PERG and glaucoma studied only the amplitude of the PERG elicited by the central stimulus field and generally found that patients with glaucoma have statistically smaller amplitudes than do control groups.' In detecting the glaucomatous abnormality the amplitude of the PERG has not been proved to be sensitive, because it has large interindividual variation resulting in a wide overlap between glaucoma and normal control groups.We previously reported that the PERG can be recorded from the paracentral ring area as well as from the central area. We also found that the ratio of the amplitudes from these two areas is less variable and more reliable than the amplitudes of themselves.'2 Moreover, the ratio of the paracentral to central PERG is likely to be more sensitive in detecting glaucomatous changes, since the paraCorrespondence to

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