| Objective:Diabetic gastroparesis(DGP)refers to gastrointestinal dysfunction on the basis of diabetes,which is characterized by abnormal esophageal pressure,decreased motor function of gastric smooth muscle,prolonged gastric cramps,and delayed gastric emptying.Clinically,it can cause manifestations such as nausea,vomiting,bloating,constipation,or diarrhoea.DGP belongs to the categories of"fullness","stomach pain","stomach pain","stomach slowness","vomiting" and"nausea" secondary to "thirst quenching disease" in Chinese medicine.Summarize and sort out the views of various doctors on the pathogenesis of DGP,and believe that DGP involves the spleen,stomach,liver,kidney and other internal organs,Qi stagnation,phlegm drinking,blood stasis,food accumulation and other pathological factors,but always based on spleen deficiency.Supervisor Professor Yang Xiaohui is one of the academic inheritors of Professor Lv Renhe,a master of traditional Chinese medicine,on the basis of inheriting Lv Lao’s academic theory,combined with clinical practice,Professor Yang Xiaohui believes that thirst is quenched for a long time,gas consumption hurts yin,resulting in weakened spleen and stomach function,qi machine lifting disorder and DGP.Spleen deficiency and qi stasis type accounted for a relatively high proportion of DGP TCM type.Professor Lv Renhe created Foxiang powder with the effect of rationalizing qi and strengthening the spleen,dissolving phlegm and reducing rebellion,which is composed of traditional Chinese medicines such as bergamor,citron,incense,swirling flower,substitute ochre,black medicine,tangerine peel,prince ginseng,ginger banxia,etc.,which have the effect of rationalizing qi and strengthening the spleen,dissolving phlegm and reducing rebellion.It has been clinically used for decades and has good clinical efficacy.In recent years,with the development of neurogastroenterology and neuroendocrinology,the role of various brain-gut peptides and neurotransmitter-mediated neuroendocrine in the pathogenesis of DGP has received more and more attention,but the biomolecular mechanism related to DGP neuroendocrine still needs to be further studied,which also limits the discovery of drug targets for the treatment of DGP and the development of new drugs.The activation of DA receptor-related signaling pathways may be the scientific basis for its efficacy,but there is a lack of in-depth research on the upstream and downstream factors and signaling pathways of dopamine-mediated neuroendocrine in the treatment of DGP by Foxiang powder.Current studies confirm that 5-HT/cAMP/PKA is an important signaling pathway for regulating DA production,regulating diabetic glycolipid metabolism and gastrointestinal motility,so this study explores the therapeutic effect and potential drug targets of Buddha Xiangshan on DGP from the neuroendocrine effect mediated by 5HT/cAMP/PKA,verifies the important role of neuroendocrine in DGP,explains the scientific connotation of related neurotransmitter pathways in DGP,and further enriches the theoretical system of traditional Chinese medicine for DGP.The full text includes three parts:literature review,clinical research and animal experiments.Methods:Clinical trial:In the clinical patient intervention study,53 subjects were divided into 27 cases in the traditional Chinese medicine group and 26 cases in the western medicine group,the traditional Chinese medicine group was treated with Foxiang powder(1 dose per day,100ml/time with water,oral before breakfast and dinner),and the western medicine group was given mosapride citrate tablets(5mg each time,3 times a day,oral before meals),and the treatment cycle of both groups was 4 weeks.The blood glucose,blood lipids,glycated hemoglobin,glycated albumin,fasting serum insulin,fasting C-peptide level,gastric emptying rate,GCSI score,and total score of ruffian certificate were evaluated in the Chinese medicine group and the western medicine group,and the protein expression of serum 5-HT,cAMP and PKA was detected by ELISA method.Animal experimentⅠ:The DGP rat model was established,and 58 male SD rats were randomly divided into 50 for modeling and 8 for normal group.The rats were fed with high-fat and high-sugar feed for 2 weeks,fasted and drank freely for 12 h and then intraperitoneally injected with 1%STZ 50 mg/kg(STZ dissolved in pH 4.50.1mol/L citrate buffer solution at a concentration of 10 mg/mL),and the blood glucose value of the tail vein was measured by glucose oxidase method 72 h after injection.The fasting blood glucose ≥of 13.9mmol/L in rats used for modeling indicated that the diabetic rat model was successfully induced.After successful modeling,the western medicine group was given mosapride citrate 1.59mg/kg·d for gavage,and the high,medium and low dose groups of Buddha Xiangshan were gavaged by 2 times,1 times and 1/2 times the equivalent doses of human and animal drugs,respectively,with 11.4g/kg·d,5.7g/kg·d,and 2.85g/kg·d of Foxiang powder,and the normal group and model group were given equal volumes of normal saline for gavage,8 in each group,for a total of 8 weeks.At the end of the 8th week of the experiment,rats are sacrificed,abdominal aortic blood is collected,and the supernatant is taken by centrifugation at 3500 r/min for 15 min at 4℃.An appropriate auount of serum was taken for automatic biochemical analysis to detect fasting blood glucose and blood lipids,ELISA was used to detect the serum motilin level of rats,and the remaining serum was stored in α-20℃ refrigerator for later use.After the blood collection is completed,all the rats are taken out from the pylorus to the end of the rectum,without additional external pull,and naturally placed flat on the experimental operation platform,the distance from the front end of carbon ink to the pylorus is measured,and the length of the entire small intestine is measured.Calculate the small bowel propulsion rate%(charcoal leading to pyloric sphincter distance(cm)/pyloric sphincter to small bowel distal distance(cm)×100%).Leave an appropriate amount of antral tissue,divided into two parts,one part is stored in liquid nitrogen,and the other part is fixed in 4%paraformaldehyde for hematoxylin-eosin staining.Animal experiment Ⅱ:After successful modeling,the administration was started with a volume of 10ml/(Kg·d)per rat for a total of 8 weeks.The western drug control group was given mosapride citrate 1.59mg/kg·d for gavage,and the high,medium and low dose groups of Foxiang powder were gavaged with 11.4g/kg·d,5.7g/kg·d and 2.85g/kg·d respectively according to the equivalent dose of human and animal drugs,and 8 animals in each group.The normal group and the model group were given equal volumes of normal saline for gastric lavage.Intervention for 8 weeks of execution,specimens were taken.The protein content of cAMP in antral tissues was detected by ELISA method,the expression of 5-HT4,PKA and p-PKA proteins in antral tissues was detected by WB method,and the expression of 5-HT4 and PKA mRNA in antral tissues was detected by RT-qPCR.Results:Clinical trial:The study found that the levels of FBG,2hPG,HbA1c and FINS in the traditional Chinese medicine group were significantly lower than those before treatment(P<0.05).The levels of TC,TG and LDL-C in the traditional Chinese medicine group were significantly lower than those before treatment(P<0.05).The total score and GCSI score of the Chinese medicine group and the Western medicine group were significantly lower than those before treatment(P<0.05),and the gastric emptying rate was significantly higher than that before treatment(P<0.05).After treatment,the total score of the Chinese medicine group was significantly lower than that in the Western medicine group(P<0.001).After treatment,the GCSI score of Western medicine group was significantly lower than that in Chinese medicine group(P<0.001).After treatment,the gastric emptying rate in the western medicine group was significantly higher than that in the traditional Chinese medicine group(P<0.01).After treatment,the two groups of"stomach bloating/abdominal distention","stomach pain" and "less food and sluggishness" were significantly reduced compared with those before treatment(P<0.05).After treatment,the scores of "stomach bloating/abdominal distention"and "less food and sluggishness" in the Chinese medicine group were significantly lower than those in the Western medicine group(P<0.05).After treatment,the scores of "fatigue and weakness","noisy stomach","poor stool","heavy drowsiness" and "loose stool" in the Chinese medicine group were significantly lower than those in the Western medicine group(P<0.05).After treatment,the effective rate of ruffian syndrome in the Chinese medicine group was higher than that in the Western medicine group(P<0.05).Compared with the normal group,the expression of serum 5-HT,cAMP and PKA proteins in the two groups was downregulated before treatment and upregulated after treatment,and the Western medicine group was higher than that in the traditional Chinese medicine group.A nimal experiment Ⅰ:The study found that compared with the normal group,the fasting blood glucose and blood lipid levels of rats in the model group increased significantly(P<0.05).The small intestine advancement rate and motilin level of rats in the model group were significantly reduced(P<0.001),the structure of the antrum in the model group showed poor clarity,the arrangement of mucosal glandular cells and muscular smooth muscle cells was disordered,the intercellular space became larger,the submucosal tissue structure was loose,the vascular hyperemia was dilated,and the cytoplasmic staining of the smooth muscle layer of the muscular layer became lighter with vacuolar changes.Compared with the rats in the model group,there was no significant difference in blood glucose level between the treatment groups(P>0.05).TC and TG levels decreased in Western medicine group,Foxiang powder high-dose group and Foxiang powder mediumdose group(P<0.05),and there were no significant differences in TC and TG levels in Foxiang powder’s low-dose group(P>0.05).Compared with the model group,motilin levels were significantly reduced in the western medicine group,the high-dose group of Foxiang powder and the medium-dose group of Foxiang powder(P<0.001),and the difference between the low-dose group of Foxiang powder was not statistically significant(P>0.05).Compared with the model group,the small intestine propulsion rate was significantly increased in the western medicine group,the high-dose group of Foxiang powder,the medium-dose group of Foxiang powder,and the low-dose group of Foxiang powder(P<0.01).Compared with the model group,the gastrontus tissue structure of rats in the western medicine group and Foxiang powder was clearer,the mucosal glandular cells and muscle smooth muscle cells were arranged neatly,the intercellular space was slightly larger,and the vacuolar changes of smooth muscle cells were reduced.The number of neurons in the cerebral cortex is increased,scattered in a small number of glial cells,and the cell morphology is close to normal.Neurons in the hippocampus are arranged neatly,the number of neurons increases,the cytoplasmic staining is deeper,and vacuolar changes are reduced.Animal experiment Ⅱ:ELISA results showed that the protein expression of cAMP in antral tissue of rats in the model group was significantly reduced compared with the normal group.Compared with the model group,the protein expression of cAMP in the western medicine group and the traditional Chinese medicine group was increased,and the expression of cAMP in antrum tissue was higher in the western medicine group and the high-dose group of Foxiang powder.The WB results showed that compared with the normal group,the expression of 5-HT4,PKA and p-PKA in the rat antrum tissue in the model group was low.Compared with the model group,the expressions of 5-HT4,PKA and p-PKA were all regulated in the traditional Chinese medicine group and the western medicine group.RT-qPCR results showed that the rat antral tissue 5-HT4 and PKA mRNA expression in the model group were low compared with the normal group.Compared with the model group,the expressions of 5-HT4 and PKAmRNA in the traditional Chinese medicine group and the western medicine group were up-regulated,among which the high-dose group and western medicine group were particularly obvious,and the expression was the lowest in the low-dose group of Foxiang powder.Conclusions:Foxiang powder can significantly reduce the glycolipid metabolism indexes such as FBG,2hPG,HbAlc,TC,TG,LDL-C,FINS and other glycolipid metabolism indicators in patients with Spleen deficiency and stagnation syndrome DGP,improve gastric emptying rate,reduce GCSI points and total points of ruffian,increase serum 5-HT,cAMP,PKA levels in patients with Spleen deficiency and stagnation syndrome DGP,and promote gastric emptying.Compared with mosabili citrate,Foxiang powder can significantly reduce the TCM symptom score in patients with Spleen deficiency and stagnation syndrome DGP,and increase the effectiveness of TCM symptoms.Intraperitoneal injection of SD rats with high sugar and high fat combined with streptozotocin can cause glucose and lipid metabolism disorders,decreased gastrointestinal motility and pathological changes of antral tissue,which can successfully construct DGP model rats,and this study successfully established DGP rat model.Foxiang powder can upregulate the expression of 5-HT4 in DGP rats,upregulate the levels of cAMP and PKA in the 5-HT signaling pathway,improve the glycolipid metabolism disorders in DGP rats,and promote gastrointestinal motility.At the same time,it can also reduce the pathological changes of antral tissue in DGP rats. |
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