Research On The Difference Of The Laser-induced Autofluorescence Spectra Of Rat Colorectal Atypical Hyperplasia And Different Stages Of Colorectal Carcinoma And Its Mechanism

Objective: To establish rat colorectal carcinoma model and analyze the characteristics of the laser-induced auto-fluorescence spectra of normal colorectal tissue, atypical hyperplasia tissue and different stages carcinoma colorectal tissue, investigate the method of identification and the mechanism in difference of spectra by studying on the expression of collagen IV and cathepsin B and their correlation.Methods: 60 young Sprague-Dawley rats (6~7weeks) were given injections of 25mg 1,2-dimethylhydrazine (DMH) per kg in abdominal cavity once a week for 18 weeks. 3 rats' colorectum were dissected every 4 weeks, the rest were dissected at the 20th week. Spectra both in vitro and in vivo of laser induced fluorescence excited at 370nm were collected and dealt with a special software. According to pathology diagnosis the specimens were divided into 4 groups: normal group, atypical hyperplasia group, early-stage carcinoma group and advancedcarcinoma group. The wavelength and intensity of peaks of spectra were detected and analyzed. At the same time, 25 specimens were collected for detecting the expression of collagen IV and cathepsin B in each group with immumohistochemical staining. The correlation between the expression of collagen IV and cathepsin B was studied. With B-actin gene as reference, semi-quantitative reverse transcriptase polymerase chain reaction(RT-PCR) was conducted to analyze cathepsin B mRNA expression in four groups.Results : The rat colorectal carcinoma models were established successfully after 20 weeks, The ratio of colorectal carcinoma induction was 82% (49/60) , multiple tumors were found in 92% (45/49) rats. Of 283 specimens detected by laser-induced autofluorescence, 64 were normal, 57 atypical hyperplasia, 78 early-stage colorectal carcinoma, and 84 advanced colorectal carcinoma. There were peaks at about 460nm, and 505nm in most spectra. 73 percent of spectra from colorectal carcinoma tissue , 69 percent of spectra from early-stage colorectal carcinoma tissue and 54 percent of spectra from colorectal atypical hyperplasia tissues showed a peak at about 635nm. The peak was not found in normal colorectal tissue. The mean wavelength of the peak ( about 460nm ) in normal group was 4.3nm larger, 10.2nm larger and 10.9nm larger than that in atypical hyperplasia group, early-stage and advanced colorectal carcinoma respectively. The wavelengths of the peak(about 460nm ) between in vitro and in vivo were different in every groups (p<0.05 ) .The value of I460 of spectra in early-stage carcinoma group and advanced carcinoma group was less than that of spectra in normal group and atypical hyperplasia group. The value of I635/I460 and I635/I600 of early-stage carcinoma group and advanced carcinoma group were larger than those of normal group and atypical hyperplasia group ( p<0.05). There was no significant difference in the value of I535/I460 and I635/I600 between early-stage carcinoma group and advanced carcinoma group. The value of I635/I460 and I635/I600 of atypical hyperplasia group was larger than that of normal group (p<0.05). A threshold of I635/I460 and I635/I600 between normal group and atypical hyperplasia group and that between atypical hyperplasia group and different stage colorectal carcinoma was used to diagnose colorectal lesions( atypical hyperplasia and different stage colorectal carcinoma ). Collagen IV expression existed in all specimens in different degrees(+~+++), there was no significant difference between collagen IV expression in normal group and that in atypical hyperplasia group. It decreased in early-stage carcinoma group and advanced carcinoma group, and there was significant difference compared with the normal group and atypical hyperplasia group. The expression of cathepsin B in early-stage carcinoma group and advanced carcinoma group increased compared with normal group and atypical hyperplasia group(p<0.05). There was anegative correlation in colorectal carcinoma tissue between collagen IV expression and cathepsin B expression (r=-0.721, p<0.05). Cathepsin B...