| Research background and purpose:Banxia Xiexin Decoction,the classic prescription of traditional Chinese medicine,is a representative prescription of Xinkaikujiang method.It is widely used to treat a variety of diseases,especially gastrointestinal diseases.According to analysis of biological database,it is found that Banxia Xiexin Decoction treating gastrointestinal diseases may be related to 5-hydroxytryptamine receptors(5-HTR)and bitter receptors(TAS2Rs).5-HTR and TAS2Rs are widely found in gastrointestinal tissues,which connect with gastrointestinal movement,secretion and visceral sensation.The prescription has multi-target and multi-link because of their complex and diverse components,thus it is proposed that 5-HTR and TAS2Rs play an important role in the mechanism of Xinkai Kujiang method of Banxia Xiexin Decoction.Therefore,the study is designed based on 5-HT4R/TAS2R38,the mechanism of Banxia Xiexin Decoction in the treatment of functional dyspepsia.Research contents:Part One:Researches of Chinese and Western Medicine on functional dyspepsia were summarized and meta-analysis was carried out to assess the efficacy and safety of Banxia Xiexin Decoction for functional dyspepsia and related symptoms.Part Two:Efficacy of low(4.03g/kg),medium(8.06g/kg)and high(16.12g/kg)dose of Banxia Xiexin Decoction was observed on normal mice.Gastric emptying rate and small intestinal propulsion rate were calculated.Motilin,ghrelin,somatostatin and vasoactive intestinal peptide in serum were recorded by Elisa.Part Three:(1)Appropriate modeling method of functional dyspepsia was evaluated to explore the efficacy of Banxia Xiexin Decoction on model mice.Firstly,the normal mice were divided into control group,iodoacetamide group,loperamide group,tail clamping group and vinegar group.The general situation of mice,gastrointestinal motility,gastrointestinal hormone and HE staining were detected.It was assessed that the iodoacetamide method was better for modeling characteristics of functional dyspepsia.(2)The mice were divided into control group,model group,Banxia Xiexin Decoction group,Xinkai group,Kujiang group and Xinkai Kujiang group.Except for the control group,the other groups were modeled by iodoacetamide method.After modeling,each treatment group was given Banxia Xiexin Decoction,Xinkai drug,Kujiang drug and Xinkai Kujiang drug respectively for 14 days.The general situation of mice,gastrointestinal motility,gastrointestinal hormones and HE staining were recorded.Part Four:(1)The mice were divided into control group,model group,whole formula group,Xinkai group and Xinkai+piperserol group.Except for the control group,the other groups were modeled by iodoacetamide method.After modeling,each treatment group was given Banxia Xiexin Decoction,Xinkai drugs,Xinkai+piperserol drugs respectively for 14 days.The general situation of mice,gastrointestinal motility,gastrointestinal hormone,Calcium concentration flow detection of receptor activation,WB,PCR,immunohistochemistry and immunofluorescence were recorded.The effective monomer components that may play a role in Xinkai group were found through molecular docking.The activation of monomer components on receptor was detected by Calcium concentration flow detection.The cell safe administration concentration was screened by MTT,and the 5-HT secretion of Xinkai group monomer was assessed by ELISA.(2)The mice were divided into control group,model group,all prescription group,Kujiang group and Kujiang+probenecid group.Except for the control group,the other groups were modeled by iodoacetamide method.After modeling,each treatment group was given Banxia Xiexin Decoction,Kujiang drug and Kujiang+probenecid respectively for 14 days.The general situation of mice,gastrointestinal motility,gastrointestinal hormone,calcium ion imaging,receptor activation,WB,PCR,immunohistochemistry,immunofluorescence were recorded.The possible effective monomer components in kujiangyao were found through molecular docking.The activation of receptor by kujiangzu group monomer components was detected by Calcium concentration flow detection.Part Five:The agonist of 5-HT4R/TAS2R38 was used to verify the relationship between the two receptors according to assessing gastrointestinal motility and gastrointestinal hormones.Results:Part One:It is found that Banxia Xiexin Decoction may be closely related to 5-HT4R/TAS2R38.Banxia Xiexin Decoction is effective for functional dyspepsia.Meta results showed effective rate:RR=1.25,95%Cl[1.19,1.32];Comprehensive symptom score:WMD=-1.90,95%CI[-2.53,-1.28];Abdominal pain:WMD=-1.45,95%CI[-2.20,-0.69];Abdominal distension:WMD=-0.793 95%Cl[-1.01,-0.57];Belching:WMD=-0.42,95%CI[-0.67,-0.16];Acid reflux:WMD=-0.40,95%CI[-0.75,-0.04];Gastric semi emptying time:MD=-10.19,95%Cl[-14.79,-5.59];Total gastric emptying time:MD=-25.00,95%CI[-29.31,-20.69];Gastric peristalsis amplitude(short diameter):MD=0.25,95%CI[0.16,0.34];Gastric peristalsis amplitude(long diameter):MD=0.97,95%CI[0.89,1.05];Gastric peristalsis frequency:MD=1.07,95%CI[0.29,1.85];Number of gastric mast cells:MD=-5.15,95%CI[-7.69,-2.61];Cholecystokinin:MD=18.25,95%Cl[10.61,25.89];Motilin:MD=39.01,95%CI[30.35,47.66].Banxia Xiexin Decoction is effective for functional dyspepsia.It can improve abdominal pain,abdominal distension,belching and other related symptoms.It also can improve gastrointestinal motility and gastrointestinal hormone level.Banxia Xiexin Decoction is a safe treatment for functional dyspepsia.Part Two:Different doses of Banxia Xiexin Decoction had no significant effects on the weight,food intake,activity,gastric emptying rate and small intestinal propulsion rate.Banxia Xiexin Decoction in medium and high dose groups decreased the level of motilin and ghrelin,however,it had no significant effect on somatostatin and vasoactive intestinal peptide.Part Three:(1)Mice in iodoacetamide group,loperamide group,tail clamping group and vinegar group has disordered hair and reduced activity.The gastric emptying rate and small intestinal propulsion rate of iodoacetamide and vinegar group decreased compared with the control group.Tail clamping increased motilin and ghrelin in serum.Vinegar group increased somatostatin in serum.The four modeling methods had no significant effect on the level of vasoactive intestinal peptide.HE staining showed that there were no significant inflammatory changes in each group.Therefore,iodoacetamide method,which reduces gastrointestinal motility and has no effect on gastrointestinal hormones,was better for Functional dyspepsia modeling.(2)After modeling with different components of Banxia Xiexin Decoctiongiven,it was found that each group had no significant effects on food intake and body weight.Xinkai group,Banxia Xiexin Decoction group,Xinkai group and Xinkai+Kujiang group improved the gastric emptying rate and small intestinal propulsion rate of functional dyspepsia mice.They also increased motilin in serum.Banxia Xiexin Decoction group,Kujiang group and Xinkai+Kujiang group increased the level of ghrelin in serum.Each group had no significant effect on somatostatin and vasoactive intestinal peptide in serum.HE staining showed that the administration did not cause inflammatory changes in gastrointestinal tract of mice.Part Four:The relationship between Xinkai group/Kujiang group and 5-HT4R/TAS2R38 was evaluated through inhibitors.After using 5-HT4R inhibitor pipercerol,compared with Xinkai group,Xinkai+pipercerol group reduced gastric emptying rate,intestinal propulsion rate and motilin in serum.Xinkai group drugs could activate 5-HT4R according Calcium concentration flow detection.It had no significant effect on the expression of 5-HT4R in stomach and small intestine.Through molecular docking,monomers with high binding scores(trigonelline,6-gingerol and protocatechuic aldehyde)could not activate 5-HT4R.It is speculated that other components in Xinkai group may activate 5-HT4R.In addition,trigonelline can increase 5-HT secretion in GES-1 cells.(2)After using TAS2R38 inhibitor probenecid,compared with Kujiang group,Kujiang+probenecid group had no significant effect on gastrointestinal motility and it reduced ghrelin level in serum.It was found that Kujiang group could activate TAS2R38 through Calcium concentration flow detection.Kujiang group increased the expression of TAS2R38 protein level in small intestine.However,it could not increase expression of gastric both protein level mRNA level as well as small intestinal mRNA level.The monomers with high binding scores(berberine,epiberberine,magnolia alkaloid,tetrandrine,scutellaria baicalensis,baicalin and scutellarin)were screened by molecular docking,in which berberine can activate TAS2R38 receptor.Part Five:The relationship between 5-HT4R/TAS2R38 and gastrointestinal motility and gastrointestinal hormones was verified by using the agonists of 5-HT4R and TAS2R38.It was found that the 5-HT4R agonist Mosapride could improve the rate of gastric emptying and intestinal propulsion.TAS2R38 agonist benzodina ammonium had no significant effect on gastrointestinal motility.Mosapride promoted the release of motilin in serum,while benzodina ammonium promoted the release of ghrelin in serum.Conclusion and significance:We confirmed that Banxia Xiexin Decoction is effective for functional dyspepsia.It can improve related symptoms,gastrointestinal motility and gastrointestinal hormone level without significant adverse reactions.Banxia Xiexin Decoction is a prescription for regulating body balance,which has no obvious effect on normal mice.Banxia Xiexin Decoction Combined with Xinkai Kujiang method can reconcile the gastrointestinal motility and gastrointestinal hormones of functional dyspepsia mice induced by iodoacetamide.The Xinkai group can improve gastrointestinal motility and promote the level of motilin by activating 5-HT4R.Kujiang group can promote the level of ghrelin by activating TAS2R38.It explains the scientific connotation of Xinkai and Kujiang of Banxia Xiexin Decoction. |
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