| Background and objective:Peptic ulcer(PU)is one of the common diseases of digestive system in the world.The incidence of the disease ranges from 1.5%to 3%,affecting about 4 million peo-ple worldwide each year.Among them,about 10%patients have suffered from the disease in their lifetime.PU often occurs in the esophagus,stomach or duodenum,of which the stomach and duodenum are the most common.Gastrointestinal bleeding,perforation,pyloric obstruction and canceration are four common complications of PU.R-(+)-rabeprazole sodium(RRS)is a new generation agent of proton pump inhibi-tor(PPI),which is the dextral body of the optical isomer of rabeprazole sodium(RS).Compared with other PPIs,RRS has the advantages of small individual differences,faster onset and long-lasting efficacy.It has been reported that RRS plays a stronger role than S-(-)-rabeprazole sodium(SRS)in the models of gastric ulcer such as water stress,non-steroidal anti-inflammatory drugs(e.g.indomethacin,aspirin)and pyloric ligation.RRS is the main anti-ulcer isomer of RS.However,systematic quantitative evaluation method has not been reported for peptic ulcer modelsin rat.Besides,the pharmacodynamics of RRS has not been reported in the models of erosive esophagitis or duodenal ulcer in rats.It is necessary to develop a sterile powder injection of RRS for severe patients such as acute gastric hemorrhage,shock,gastrointestinal surgery and disease stress.In this paper,we use erosive esophagitis,gastric ulcer and duodenal ulcer models to systematically evaluate the therapeutic effects of RRS by comparing with SRS and RS.Using the comparative method,we elucidated the image quantitative evaluation methods including the discovery,establishment,application and discussion.The ad-vantages,applicability and scientificity of image quantitative evaluation method in pharmacodynamic evaluation of peptic ulcer model were found,by comparing with traditional evaluation methods.It is the first time to establisha digital image quantita-tive evaluation method both for model evaluation and efficacy evaluation and system-atically studythe anti-peptic ulcer effect of RRS,which provided a new scientific re-search method and evaluation basis for the systematic evaluation of anti-peptic ulcer drugs.Methods:Experimental models of reflux esophagitis,pyloric ligation gastric ulcer and acetic acid duodenal ulcer were reproduced,and rats were treated with low,medium and high doses of RRS(0.9,1.8,3.6 mg·kg-1),SRS(1.8 mg·kg-1)and RS(3.6 mg·kg-1)respectively.ImageJ image quantitative evaluation method was established to obtain key indicators including ulcer area and ulcer proportion through image processing processes such as calibration,binarization,background removal,threshold setting,data acquisition,etc.The anti-peptic ulcer effects of RRS,SRS and RS were evaluat-ed by observing the total gastric acid secretion,pH value,esophagitis injury area,esophagitis injury proportion,gastric ulcer area,gastric ulcer proportion and duodenal ulcer area.The differences between ImageJ image quantitative evaluation method and traditional grading method were discussed in experimental ulcer research by analogy and exploring the methods.1.Gastric juice secretion function testSeventy rats were randomly divided into 7 groups.The intervention groups were injected with corresponding solution via caudal vein once.The model control group and sham operation group were given saline in the same way.An hour later,the rats were actualized operation.After treatment for 5 hours,gastric juice was collected and pepsin activity was measured.Total gastric acid content was determined by acid-base titration.The absorbance of enzymatic reduction products was determined by colorim-etry.Total gastric acid secretion,pH value,inhibition rate of gastric acid secretion and pepsin activity were calculated.2.Erosive esophagitis testSeventy rats were randomly divided into 7 groups.After anesthesia,the rats were actualized operation.Each drug intervention group was injected with the correspond-ing solution via caudal vein once,and the model control group and sham operation group were given the same volume of physiological saline.After treatment for 24 h,the esophaguses werecut off and scored.The digital images of tissue mucosa were collected and processed for imagequantization evaluation.The area of esophageal in-jury and the proportion of esophageal injury were obtained.3.Gastric ulcer testSeventy rats were randomly divided into 7 groups.Each drug intervention group was injected with the corresponding solution via caudal vein once,and the model control group and sham operation group were given the same volume of physiological saline.After treatment for 18 h,the degree of gastric ulcer was measured and judged.The gastric ulcer index and inhibition rate of gastric ulcer were calculated.The digital images of tissue mucosa were collected and processed for image quantization evalua-tion.The area of esophageal injury and the proportion of esophageal injury were ob-tained.4.Duodenal ulcer testSeventy rats were randomly divided into 7 groups.After anesthesia,duodenal ulcer was duplicated by acetic acid.Rats in each group were injected with corresponding drug solution via tail vein,once a day for consequtive 7 days.The model control group and the sham operation group were given normal saline with the same method.After the last administration,the anesthetic was executed and the upper part of the stomach and duodenum were removed.Manual measurement and image quantifica-tion were used to measure the area of ulcer.Results:1.Effect of RRS on gastric secretion and pepsin activity in ratsCompared with model control group,RRS(1.8 and 3.6 mg kg-1)significantly in-hibited the total gastric acid secretion and gastric ulcer index,and increased the pH value of gastric juice.Both RRS(3.6 mg kg-1)and rabeprazole sodium(3.6 mg kg-1)significantly decreased pepsin activity(P<0.05,or P<0.01,respectively).There was no significant difference between RRS(1.8 mg·kg-1)and RS(1.8 mg·kg-1).The inhibi-tion rates of gastric acid secretion and pepsin in the R-(+)-rabeprazole sodium group were higher than those in the SRS and RS groups.2.Effect of RRS on esophagitis in rats(a)Grading method:compared with model control group,the esophagitis indexes of 1.8 and 3.6 mg kg-1 RRS and 3.6 mg kg-1 RS were significantly decreased after 24 h by a single tail intravenous injection(P<0.05,or P<0.01.respectively).The esophagitis index of RRS(1.8 mg·kg-1)was significantly lower than that of SRS(P<0.05).There was no significant difference in esophagitis index between RRS(3.6 mg·kg-1)and RS(3.6 mg·kg-1)(P>0.05).The inhibition rates of esophagitis by 1.8 and 3.6 mg·kg-1 RRS were higher than that of the same dose of SRS or RS,respectively.(b)Image quantification evaluation method:compared with model control group,the area and proportion of esophagitis injury were significantly reduced in 0.9,1.8,3.6 mg·kg-1 RRS groups(P<0.05,or P<0.01,respectively).The area and proportion of esophagitis induced by RRS(1.8 mg·kg-1)was significantly lower than that of SRS(P<0.01).There was no significant difference between 3.6 mg·kg-1 RRS and 3.6 mg·kg-1 RS in the area and the proportion of esophagitis.3.Effect of RRS on gastric ulcer in rats(a)Grading method:compared with model control group,the ulcer index of 1.8 and 3.6 mg·kg-1 RRS and 3.6 mg·kg-1 RS decreased significantly after 5 h by a single tail intravenous administration(P<0.05,or P<0.01,respectively).There were no sig-nificant differences in gastric ulcer indexes between 1.8,3.6 mg·kg-1 RRS and 1.8 mg·kg-1 SRS and 3.6 mg·kg-1 RS respectively(P>0.05).The gastric ulcer inhibitory rates of 1.8,3.6 mg·kg-1 RRS were higher than the same dose of SRS and RS,respectively.(b)Image quantification evaluation method:compared with the model control group,the area of gastric ulcer and the proportion of ulceration decreased significant-ly in the 0.9,1.8,3.6 mg·kg-1 RRS groups(P<0.05,or P<0.01,respectively).The area and the proportion of gastric ulcer induced by RRS(1.8 mg·kg-1)was signifi-cantly lower than that of SRS(P<0.05,or P<0.01,respectively).There was no sig-nificant difference between 3.6 mg·kg-1 RRS and 3.6 mg·kg-1 RS the area and the pro-portion of gastric ulcer.(c)Pathological results of gastric ulcer:Gastric mucosa injury was the most serious in the model control group.10/10 the rats had denaturation and necrosis of gastric mucosa epithelium and formed deep ulcer.The lesion degree of SRS 1.8 mg·kg-1 group was the same as that of model control group.The epithelium of gastric mucosa degenerated and necrotized in 10/10 rats,and formed deep ulcer.The degree of lesion of RRS 0.9 mg·kg-1 was similar to that of the model control group.10/10 rats gastric mucosal epithelium degenerated,necrotized and ulcer formed,and deep ulcer formed in 7/10 rats.RRS 1.8 mg·kg-1 ulcer was slightly improved compared with the model control group.Gastric mucosal epithelial degeneration and necrosis occurred in 10/10 rats,and ulcer formation occurred in 4/1 0 rats.The mucosal epithelial lesions of 3.6 mg·kg-1 RRS and 3.6 mg·kg-1 RS were basically the same.The mucosal epithelial de-generation and necrosis were observed in 8/10 rats,and mucosal ulcer was observed in 7/10 rats.4.Effect of RRS on duodenal ulcer in rats(a)Direct measurement method:compared with model control group,the duodenal ulcer area of RRS(1.8,3.6 mg·kg-1)and RS(3.6mg·kg-1)decreased significantly after tail vein injections for 7 d(P<0.01).Compared with SRS,the duodenal ulcer area of RRS(1.8 mg·kg-1)decreased significantly(P<0.01).There was no significant differ-ence in the area of duodenal ulcer between RRS(3.6 mg·kg-1)and RS(3.6 mg·kg-1)(P>0.05).The duodenal ulcer inhibitory rates of 1.8,3.6 mg·kg-1 RRS were higher than that of the same dose of SRS or RS,respectively.(b)Image quantification evaluation method:compared with the model control group,the area of duodenal ulcer and the proportion of duodenal ulcer decreased sig-nificantly in the RRS(1.8,3.6 mg·kg-1)groups(P<0.01).The duodenal ulcer inhibi-tory rates of RRS(1.8,3.6 mg·kg-1)were higher than that of equal dose of SRS or RS,respectively.5.Comparisons in evaluation methodsThe data obtained from the image quantization evaluation method are generally ap-proximate to the traditional method,but slightly different.The difference was mainly manifested as follows:image quantitative evaluation method suggested that the effec-tive dose of reflux esophagitis and pyloric ligation gastric ulcer model was slightly lower than that of traditional method.ImageJ image quantitative evaluation method can read ulcer integral area and ulcer proportion quickly through calibration,binariza-tion,background removal,threshold setting and data acquisition.The process of im-age quantitative evaluation method is controllable,with good repeatability and tracea-bility,and can objectively and accurately reflect the degree of lesion.Conclusion:Under the experimental condition of this thesis,RRS had the effects of anti-esophagitis and anti-gastric ulcer at the doses of 0.9,1.8 and 3.6 mg·kg-1,and also had the anti-duodenal ulcer effect at the doses of 1.8 and 3.6 mg·kg-1.RRS(1.8mg·kg-1)has a significant advantage in comparison with the same dose of SRS,and is equivalent to RS at the dose of 3.6 mg·kg-1.ImageJ program platform is suitable for image data quantitative evaluation onthe model of esophagitis,pyloric ligation gastric ulcer and acetic acid duodenal ulcer.The area and proportion of injury were more scientificas quantitative indicators for evalu-ating lesion severity. |
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