| ObjectiveIn this study,we used high-fat diet combined with low-dose streptozotocin(STZ)injection to establish diabetic gastroparesis(DGP)rat model,and observed the differences of body weight,food intake,blood glucose,blood fat and gastric motility regulation of electroacupuncture at Zusanli and Tianshu in DGP rats,and explored some neural mechanisms of Electroacupuncture on gastric motility regulation of DGP rats from the perspective of the enteric nervous system(ENS).MethodsHealthy male Sprague Dawley(SD)rats(180-200g)were divided into normal group and model group.The normal group was fed with normal diet,and the model group was fed with high-fat diet.Two weeks later,the model rats were injected with 35mg/kg STZ to induce type 2 diabetes mellitus(T2DM).Six weeks later,the diabetic rats were divided into model group,Zusanli group and Tianshu group.The Zusanli group and Tianshu group were given electroacupuncture intervention for 5 weeks.During the whole experiment,the body weight and non fasting blood glucose of rats were measured every week.At the end of the electroacupuncture intervention,glucose tolerance test was performed to check the degree of glucose tolerance,blood samples were collected to detect the levels of glycated serum protein,insulin and lipids,gastric emptying rate was detected by ultrasound and phenol red test,and gastric antrum contraction was detected by balloon manometry.The gastric antrum tissues were collected,and the protein levels of neuronal nitric oxide synthase(nNOS),choline acetyltransferase(ChAT),protein gene product 9.5(PGP9.5),glial fibrillary acidic protein(GFAP),glial cell line-derived neurotrophic factor(GDNF)and phospho-Akt(P-Akt)were detected by western blot The mRNA levels of nNOS and ChAT were detected by real-time quantitative polymerase chain reaction(qPCR).The expressions of PGP9.5 and S100 were detected by immunofluorescence.The contents of nerve growth factor(NGF)and neurotrophin-3(NT-3)were detected by enzyme linked immunosorbent assay(ELISA).Results1.Compared with normal rats,the weight,food intake and blood glucose(including fasting blood glucose,non fasting blood glucose,glycosylated serum protein)of model rats increased;glucose tolerance decreased;serum insulin decreased;homeostatic model assessment of insulin resistance(HOMA-IR)increased;blood lipid(including triglyceride,total cholesterol,low-density lipoprotein and free fatty acid)increased;semi-solid and liquid gastric emptying rate decreased;The results showed that T2DM rat model with hyperlipidemia,insulin resistance and gastroparesis was successfully established by high-fat diet combined with STZ injection.Electroacupuncture at Zusanli or Tianshu significantly decreased the body weight,food intake,blood glucose and HOMA-IR of model rats,and increased glucose tolerance,among which Tianshu was better than Zusanli in regulating blood glucose,glucose tolerance and insulin resistance;for the regulation of blood lipid,electroacupuncture at both Zusanli and Tianshu can down regulate the levels of triglyceride,total cholesterol,low density lipoprotein and free fatty acid,and up regulate the level of high density lipoprotein.Among them,Tianshu has better regulation of triglyceride than Zusanli;for the regulation of gastric motility,electroacupuncture at Zusanli or Tianshu could increase gastric semi-solid and liquid emptying rate,gastric antral pressure and the amplitude of antral motility,and reduce the frequency of antral motility.Among them,electroacupuncture at Zusanli is better than Tianshu in regulating gastric emptying rate,gastric antral pressure and the amplitude of antral motility.2.The protein and mRNA levels of nNOS and ChAT in gastric antrum of DGP rats were significantly decreased.Electroacupuncture at Zusanli or Tianshu could up regulate the protein and mRNA levels of nNOS in gastric antrum of DGP rats.Electroacupuncture at Zusanli could raise the protein and mRNA levels of ChAT in gastric antrum of DGP rats.Tianshu had no significant effect on the regulation of ChAT.The protein ratio of ChAT/nNOS in model rats increased,and electroacupuncture at Zusanli or Tianshu could decrease the ratio of ChAT/nNOS.3.The results of ELISA showed that the contents of NGF and NT-3 were decreased in gastric antrum of DGP rats;the expressions of PGP9.5 and S100 were decreased by immunofluorescence;the expressions of PGP9.5,GFAP,GDNF and p-Akt were decreased by Western blot;Electroacupuncture at Zusanli or Tianshu could improve their levels,and the regulation of Zusanli on NGF,NT-3,PGP9.5 and p-Akt was better than Tianshu.Conclusion1.Electroacupuncture at Zusanli or Tianshu can regulate DGP comprehensively by regulating body weight,food intake,blood glucose,blood lipid and gastric motility.Tianshu is better than Zusanli in improving glucose metabolism and partial lipid metabolism,and Zusanli is better than Tianshu in improving gastric emptying and antral motility.2.Electroacupuncture at Zusanli or Tianshu can correct the imbalance of nNOS and ChAT in gastric antrum caused by DGP.But Zusanli can raise the expression of ChAT,while Tianshu had no significant effect.3.Electroacupuncture at Zusanli or Tianshu can reduce the injury of enteric neurons and enteric glial cells(EGCs),promote the secretion of neurotrophic factor and the activation of GDNF downstream pathway in EGCs.The protective effect of Zusanli on enteric neurons,the secretion of NGF and NT-3 and the activation of GDNF downstream pathway are better than Tianshu.In conclusion,electroacupuncture at Zusanli and Tianshu can treat DGP comprehensively,and its neural mechanism may include correcting the imbalance of excitatory and inhibitory neurotransmitters in ENS and promoting glial cells to secrete neurotrophic factors to promote the survival of enteric neurons.From the difference of acupoint effect,Tianshu may focus on improving blood glucose,blood lipid and insulin resistance,and Zusanli may focus on improving gastric motility. |
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