| Sanzifang,a national medicine,is an experienced prescription summed up by professor Tian Yaozhou after many years of clinical practice.The national prescription for treating Dafa,which is invigorating qi and nourishing yin,removing blood stasis and detoxifying,has remarkable curative effect in improving the clinical symptoms of colorectal adenoma.Based on the previous clinical observation,TCM diagnosis data and literature research results of the research group,this topic further analyzes the theoretical basis of traditional Chinese and Western medicine and explores its mechanism of action.OBJECTIVE:To explore the effects of Sanzifang,a national medicine,on the growth intervention of intestinal adenoma,the proliferation and apoptosis of adenoma cells and the key molecules of Wnt/β-catenin signaling pathway in APC(min/+)mice,to clarify the action target and mechanism,and to further elaborate the action mechanism of Sanzifang,so as to provide scientific and theoretical basis for clinical treatment of colorectal adenoma.METHODS:Five 4-week-old APC(min/+)mice were selected as the blank group by random number table method,and the remaining 25 4-week-old APC(min/+)mice were given 2%DSS for one week,during which they were weighed daily.Then they were divided into model group,aspirin group and national medicine Sanzifang low,medium and high dose group according to random number table method,with 5 mice in each group.After modeling,the blank group and model group were given equal volume distilled water every day,aspirin group was given 12.9 mg/(kg·d)suspension,national medicine Sanzifang granules were given 1.7 g/(kg·d)Chinese medicine granules,middle dose group was given 3.4 g/(kg·d),and high dose group was given 6.8 g/(kg·d).After administration for 12 weeks,the changes of mice’s mass during the experiment were observed and counted daily.At the end of the experiment,the mice in each group were dissected to take out all the intestinal tubes,count the number,location and size of adenomas;HE staining was used to observe the pathological condition of colorectal adenoma in each group under light microscope and determine the pathological grade of adenoma.Ki67 staining and TUNEL staining were used to detect the proliferation and apoptosis of colorectal adenoma in each group.At the same time,the total protein of colorectal adenoma in each group was extracted,and the changes of β-catenin,PCNA and Bcl-2 protein were detected by Western blot Immunohistochemistry was used to detect the expression and localization of Wnt signaling pathway related proteins β-catenin,c-Myc and cyclin D1 in colorectal adenomas.RESULTS:① APC(min/+)mice in each group did not die during the experiment,and black stool and rectocele appeared in the model group at the eighth week after modeling;There was no black stool and rectocele in the blank group and the administration group during the experiment.②After modeling,compared with Apc(min/+)mice in blank group,the weight of model mice decreased significantly(P<0.05).After 12 weeks’ intervention,compared with the blank group,the weight of the model group mice was significantly different(P<0.05),while compared with the model group,the weight of Apc(min/+)mice in each administration group had no significant difference(P>0.05).③Compared with Apc(min/+)mice in the blank group,the number of total intestinal adenomas,small intestinal adenomas and large intestine adenomas in the model group increased significantly(P<0.05).Compared with the model group,the number of adenomas in the total intestine and small intestine of mice in each administration group decreased significantly,and the number of adenomas in the low and middle dose groups of Sanzifang was statistically significant(P<0.05).④compared with Apc(min/+)mice in blank group,the expression of β-catenin,c-Myc and cyclin D1 protein in large intestine of model group mice increased(P<0.05).Compared with the model group,the expressions of β-catenin,c-Myc and cyclin D1 in the large intestine of mice in each treatment group were decreased,and there was statistical significance in the middle dose group of Sanzifang(P<0.05).⑤Compared with the blank group,ki67 protein expression in colorectal tissue of model group was significantly increased,while TUNEL staining expression was significantly decreased,with statistical significance(P<0.01).Compared with the model group,the expression level of ki67 protein decreased and TUNEL staining increased in colorectal adenoma tissues of mice receiving drug intervention in each group,among which aspirin group and Sanzifang medium dose group had statistical significance(P<0.05).In TUNEL staining expression,the dose group of Sanzifang was more prominent(P<0.01).Compared with aspirin group,the middle dose group had statistical significance(P<0.05).⑥Compared with Apc(min/+)mice in blank group,the protein expression levels of PCNA and Bcl-2 in large intestine of model group mice were significantly higher(P<0.01).Compared with the model group,the expression levels of PCNA and Bcl-2 in large intestine tissue of each receiving group were decreased,and the expression levels of aspirin group,national medicine Sanzifang low and middle dose groups were statistically significant(P<0.01).CONCLUSION:① Sanzifang,a national medicine,has good drug safety.Aspirin and Sanzifang,a national medicine,have no obvious influence on the weight of Apc(min/+)mice.②Sanzifang,a national medicine,can significantly inhibit the growth of intestinal adenomas in APC(min/+)mice,and its effect is better than aspirin.③Sanzi Prescription,a national medicine,can reduce the positive rate of PCNA and down-regulate the expression of Bcl-2 in colorectal adenoma model APC(min/+)mice,thus inhibiting the proliferation of colorectal adenoma cells and promoting their apoptosis.④Sanzifang,a national medicine,may block the downstream target genes of Wnt/β-catenin signaling pathway—c-Myc and cyclin D1,reduce the expression of c-Myc and cylin D1,inhibit the activation of Wnt/β-catenin signaling pathway,promote the degradation ofβ-catenin,thus inhibiting cell proliferation,canceration and adenoma... |
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