Effects Of Nesfatin-1 On Colonic Motility And PAR2/mTOR Signal Pathway In Rats With Irritable Bowel Syndrome

Objective:To explore the possible regulatory role of Nesfatin-1 on colonic motility and PAR2/m TOR signaling pathway in rats with irritable bowel syndrome（IBS）visceral hypersensitivity,and further explore the regulatory role of Nesfatin-1 in the formation of IBS visceral hypersensitivity.It provides a basis for finding a new entry point for effective regulation of visceral hypersensitivity and discovering a new treatment strategy for IBS.Method:1.Experimental animals and groupsTwenty-five 2-day-old male Spragne-Dawley rats were purchased,including 15PAR2^-/-rats and 5 PAR2^+/+rats,with the body weight of 5-6g per rat.Neonatal rats were raised together with their mothers.The experimental animals were divided into blank control group（group C）and IBS model group.After successful modeling,the model group was divided into PAR2^+/+IBS group,PAR2^-/-IBS group,PAR2^-/-IBS central intervention group and PAR2^-/-IBS peripheral intervention group,with 5 mice in each group.2.Preparation of visceral hypersensitivity modelIBS group(PAR2^+/+IBS group and PAR2^-/-IBS group):Neonatal rats were separated from their mothers for 3 hours in the morning from day 2 to day 21,during which lactation was stopped.On day 8 to 20,0.5%acetic acid 0.4ml enema was given in the afternoon every day.On day 21,all rats were weaned and separated from their mothers,and the experiment was started after 5 weeks（8 weeks of age）in standard environment.3.Validate IBS animal modelThere were 5 rats in group C and 5 rats in group IBS,and the model was verified by AWR score and colonic histological staining.4.Experimental intervention methodExperimental animals were treated with nesfatin-1 central and peripheral intervention,respectively.The central intervention was injected into the lateral ventricle,and the peripheral intervention was injected into the tail vein.5.Colonic motility testThe amplitudes and frequencies of fast and slow waves in colon were recorded by a functional experiment system.6.Immunohistochemistry（IHC）was used to detect the expression of m TOR in colon tissues of rats7.Statistical analysisStatistical software SPSS26.0 and Empower Stats3.0 were utilized for information handling and examination.The measurement data in accordance with normal distribution were expressed by mean±standard deviation.T-test was utilized for compare between the two gatherings,and one-way ANOVA was utilized for pairwise examination between different gatherings.Result:1.General conditions of experimental animals There was no significant difference in body weight among C group,PAR2^+/+IBS group and PAR2^-/-IBS group（F=1.326,P>0.05）.2.AWR score of rats in IBS group suggested that rats had visceral hypersensitivity,and colonic histological staining suggested that rats in model group had no intestinal mucosa inflammation and other organic pathological changes,suggesting that modeling was successful.3.Comparison of colonic motility in IBS models with different subgroups3.1 Comparison of colon fast wave index:there were significant differences in amplitude and frequency of colon fast wave in PAR2^+/+IBS group,PAR2^-/-IBS group,PAR2^-/-IBS central intervention group and PAR2^-/-IBS peripheral intervention group（F=53.855,F=9.794,P<0.05）.3.2 Comparison of colon slow wave index:there were significant differences in amplitude and frequency of colon slow wave in PAR2^+/+IBS group,PAR2^-/-IBS group,PAR2^-/-IBS central intervention group and PAR2^-/-IBS peripheral intervention group（F=135.580,F=20.731,P<0.05）.4.The expression of m TOR in colon tissue:The expression of m TOR in colon tissue of PAR2^-/-IBS central intervention group and PAR2^-/-IBS peripheral intervention group was higher than that of PAR2^+/+IBS group（P<0.05）.The expression of m TOR in PAR2^+/+IBS group was higher than that in PAR2^-/-IBS group（P<0.05）.Conclusion:1.Nesfatin-1 is involved in the regulation of colonic motility in visceral hypersensitive rats with IBS,and the regulation effect of central intervention pathway is more significant than that of peripheral pathway.2.Nesfatin-1 may be involved in the regulation of colonic motility in splanchnic hypersensitive rats through bidirectional regulation of PAR2/m TOR signaling pathway.