Intervention And Mechanism Study Of Xiaotan Tongfu Decoction On Colorectal Precancerous Lesions In Obese Mice

BackgroundColorectal cancer has become one of the most common malignancies in the world.With the development of our economy and the improvement of people’s living standard,the traditional lifestyle and eating habits have changed greatly,and the incidence of colorectal cancer in China has been increasing year by year.The occurrence of colorectal cancer is closely related to obesity,so colorectal cancer is an obesity-related tumor.The increase in morbidity may be associated with an increase in the incidence of obesity in recent years.The occurrence of colorectal cancer is a long process.If the early development of this process can be blocked,can reduce the incidence of colorectal cancer can be reduced.On the basis of summing up the theory of the previous people,professor Wei Pinkang put forward the "phlegm syndrome theory" to treat gastrointestinal tumors from phlegm.He believes that "sputum" is one of the key substances of tumor occurrence,development and metastasis,So he proposed Xiaotan Tongfu as the Core principles for the treatment of colorectal cancer,and the create " Xiaotan Tongfu Decoction" used in clinical,and achieved good results.Through the early part of our study,we found Xiaotan Tongfu Decoction recipe affected the occurrence and development and metastasis of colorectal cancer through multi-target and multi-channel,but in obesity colorectal cancer lesions have not yet in-depth study.Therefore,this subject on the basis of previous studies to observe the Xiaotan Tongfu Decoction of obesity colorectal cancer precancerous lesion model,for the treatment of colorectal cancer from the phlegm to provide experimental basis.PurposeTo observe the effect of obesity on precancerous lesions of colorectal cancer.Using Xiaotan Tongfu Decoction intervention to observe the changes of body weight,the incidence and the number of adenomas in obesity colorectal precancerous lesions,adiponectin and adiponectin receptor 1,adiponectin receptor 2 gene and protein expression changes.To investigate the effect of Xiaotan Tongfu Decoction on colorectal cancer precancerous lesion in obesity mice and its possible mechanism.MethodThe study is divided into three parts.The first part: the establish ofcolorectal precancerous lesion modelThe model of colorectal precancerous lesion was established by AOM combined with DSS composite modeling method.Pathological examination showed changes in intestinal tissue after modeling,confirming the success of the model.1、Copy the DSS "four-step" model with different concentrationsC57BL / 6 mice were divided into 4 groups by random number table method: group A(blank group),group B(AOM / DSS low dose group: 1.5% DSS),group C(AOM / DSS middle dose group: 2.5% DSS)Group D(AOM / DSS high dose group: 3.5% DSS).Blank group is not processed.In the three model groups,AOM(effective concentration 12.5mg / kg)was injected intraperitoneally with 1.5%,2.5% and 3.5% DSS drinking water for 1 week and followed by drinking water for 2 weeks free drinking,a total of 3 cycles(9 weeks).Used to determine the concentration of DSS.2、Establishment of three-cycle DSS model with different number of AOMAccording to the results of different concentrations of DSS "four-step" model,the concentration of DSS was chosen to be 3.5%.C57 BL / 6 mice were randomly divided into four groups: group A(blank group),group E(AOM × 1 / DSS group),group F(AOM × 2 / DSS group)and group G(AOM × 3 / DSS group).E group was injected intraperitoneally 12.5mg / kgAOM on the first day of the first week,F group was injected intraperitoneally 12.5mg / kgAOM on the first day of the first week and the fourth week,F group was injected intraperitoneally 12.5mg / kgAOM on the first day of the first week,the fourth week and the seventh week.Each group was given 3 weeks of 3.5% DSS drinking,the first week,the fourth week and the seventh week,the rest of the time to give ordinary drinking water.The blank group was injected intraperitoneally with the same dose of saline,given the ordinary drinking water.By the end of the eighteenth week,the experiment ended.Used to determine the number of AOM injection.The second part: effect of obesity on precancerous lesions of colorectal1、Preparation of mouse obesity modelThe mice were divided into normal diet group and high fat diet group.The high fat diet group was given high fat diet.When the average weight of the mice in the high fat diet group was more than 20% of the average weight of the mice in the normal diet group,obesity modeling success.2、To establish colorectal precancerous lesion modelUsing a random number table method,the normal diet group was divided into normal control group and common model group.The high fat diet group was divided into obesity control group and obesity model group according to the same method.Normal model group and obesity model group were treated with AOM 12.5mg / kg intraperitoneal injection combined three times with 3 cycles of 3.5% DSS free drinking method.The control group and the obesity control group were intraperitoneally injected with the same dose of saline,given the ordinary drinking water.3、To observe the effect of obesity on colorectal cancer18 weeks the experiment ends.To observe the changes of the basic condition,body weight and tumor growth and pathological changes of the common model group and the obesity model group.The third part: effect of Xiaotan Tongfu Decoction on Colorectal precancerous lesions and Adiponectin and Its Receptor in Obesity Mice1、Preparation of obesity and colorectal precancerous lesions modelThe same method with the second part.2、To observe the effect of Xiaotan Tongfu Decoction on colorectal cancerprecancerous lesion and its effect on adiponectin and its receptorexpressionMice were randomly divided into four groups: obesity group,obesity model group,obesity prevention group and obesity treatment group.Obesity blank group made obesity model,the other three groups were prepared obesity and colorectal cancer lesions model.Obesity prevention group in the seventh week began to added the Xiaotan Tongfu Decoction,obesitytreatment group at the beginning of the fifteenth week to added the Xiaotan Tongfu Decoction,to the end of the eighteen week,end experiment.The general condition and weight change of mice were observed.Six mice were randomly sacrificed at the end of the sixth week,thefourteenth and the eighteen week.Enzyme-linked immunosorbent assay of serum interleukin-6 and serum adiponectin levels,observe the pathological changes of colorectal mucosa in mice,Real time PCR and Western Blot were used to detect the gene and protein expression of adiponectin and adiponectin receptor 1 and adiponectin receptor 2 in colorectal cancer.Result1、Successful establishment of mouse colon precancerous lesion model.AOM12.5% combined with DSS3.5% two-step three-cycle group was the best.At the end of the sixth week,mice intestinal mucosal congestion,edema,at the end of the ninth week,mucosal ulceration appeared,at the end of the fourteenth week,early adenoma appeared,the end of the eighteenth week,adenoma tumor formation rate of 100%.No mouse death occurred.2、Obesity model mice had colorectal cancer lesions earlier than in the normal model group,and the number of adenomas was more than that of the normal model group,P <0.05,there was significant difference between the two groups.Indicating that obesity on the occurrence of colorectal cancer precancerous lesions have a role in promoting.3、From the seventh week after the application of Xiaotan Tongfu Decoction intervention,at the end of fourteenth week and eighteenth week,the number of tumors in obesity prevention group mice were all less than the obesity model group,P <0.05,the difference was statistically significant.From the fifteenth week after the application of Xiaotan Tongfu Decoction intervention,at the end of eighteenth week,the number of tumors in obesity treatment group mice was less than the obesity model group,P <0.05,the difference was statistically significant.It is suggested that Xiaotan Tongfu Decoction can reduce the incidence of colorectal cancer in obesity mice.4、By detecting the levels of interleukin-6 and lipids in mice,we found that,at the end of the fourteenth week,the level of serum interleukin-6 in the group of obesity prevention group was lower than that in obesity model group,P <0.05,the difference was statistically significant,the level of serum adiponectin in the group of obesity prevention group was higher than that in obesity model group,P <0.05,the difference was statistically significant.At the end of the eighteenth week,the level of serum interleukin-6 in the group of obesity prevention group was lower than that in obesity model group,P <0.05,the difference was statistically significant,the level of serum adiponectin in obesity prevention group and treatment group were all higher than that in obesity model group,P<0.05,the difference was statistically significant.The results showed that Xiaotan Tongfu Decoction could decrease serum interleukin-6 level and increase serum adiponectin level in obesity mice,and the effect of early application was better.5、By detecting the gene expression of adiponectin and its receptor in colorectal tissue of mice,we found that,at the end of fourteenth week,the expression of adiponectin and its receptor in the obesity prevention group was lower than that in the obesity model group,P <0.05,the difference was statistically significant.At the end of eighteenth week,the expression of adiponectin and its receptor in obesity prevention group and treatment group were all higher than that in obesity model group,P<0.05,the difference was statistically significant.The results showed that Xiaotan Tongfu Decoction could increase the colorectal tissue adiponectin and its receptor gene expression.6、At the end of eighteenth week,compared with the obesity model group,the adiponectin and adiponectin receptor proteins in obesity prevention group mice were up-regulated,and adiponectin receptor protein expression was up-regulated in obesity treated group mice.7、At the end of eighteenth week,compared with obesity treatment gtoup,there was a decreasing trend in the number of colorectal adenomas in obesity prevention group,adiponectin and its receptor gene expression upregulated.Conclusion1、Xiaotan Tongfu Decoction can inhibit the occurrence of precancerous lesions of colorectal cancer,by regulating obese mice colorectal mucosa adiponectin and its receptor expression,change the intestinal local microenvironment.2、Early use of the Xiaotan Tongfu Decoction intervention may be more conducive to its role to play.