| BackgroundIrritable bowel syndrome(IBS)is the common refractory gastrointestinal functional disease.The clinical manifestations are abdominal pain associated with changes in bowel characteristics or bowel habits,with a wide range of age-related illnesses,high incidence,Easy to relapse.the current global incidence of IBS is 10%to 20%.Repeated episodes of IBS symptoms aggravate the patient's physical,mental,and economic burdens and severely affect the quality of life of patients.The Rome IV diagnostic criteria for functional gastrointestinal disorders classify IBS as constipation,diarrhea,mixed,and indeterminate,with diarrhea predominant irritable bowel syndrome(IBS-D)having the highest clinical incidence.Moderm medicine believes that the pathogenesis of IBS-D is abnormal gastrointestinal motility and visceral hypersensitivity,so treatment is to improve gastrointestinal motility,regulate visceral sensation,diarrhea,antispasmodic,rapid,but the long-term efficacy is not obvious,with high recurrence rate.The Rome Committee in 2016 Conclues that IBS-D is essentially a brain-intestinal interaction disorder,highlighting the interaction between the central nervous system and the gastrointestinal tract.Affect the status in the disease.The formal proposal of the theory provides more directions for the clinical treatment of modern medicine,including the use of psychoactive drugs,However,non-professional physicians use psychotropic drugs due to lack of regulation and the drugs have many side effects.IBS-D is the dominant disease of traditional Chinese herbs,but The treatment mechanism is not clear.Visceral hypersensitivity is the core mechanism of the pathogenesis of IBS.Brain-intestinal neural plasticity is one of the most important factors that contribute to the visceral hypersensitivity.The effect of brain-intestinal synaptic plasticity on the visceral hypersensitivity of IBS-D is a hot topic in recent years.As a physiological basis for changes in neuroplasticity,structural and functional changes in synapses and neurons are direct evidence of altered neural plasticity.The activation of some immediate early genes(IEGs)(Arc,c-fos)is associated with changes in synaptic plasticity.Activation of Arc and c-fos may be the intrinsic mechanism of IBS-D visceral hypersensitivity.Brain derived neurotrophic factor(BDNF)is an important substance that promotes the growth,development and repair of neurons and exerts their physiological functions.Therefore,whether BDNF expression is normal is closely related to neural plasticity.CREB/BDNF/TrkB signaling pathway is an important pathway to regulate BDNF expression.In this pathway,early and immediate genes are also involved in some of these links,but the mechanism of action is not yet clear.Based on the theoretical nature of the brain-intestinal interaction,this experiment studied the changes of IEGs and CREB/BDNF/TrkB signaling pathways in IBS-D and their influence between each other on the brain-intestine level,thus providing theoretical basis for Treatment of IBS-D.PurposeBased on brain-gut interactions,we used immunohistochemistry,PCR,and Elisa technology to explore the mechanism of immediate early gene and CREB/BDNF/TrkB signaling pathway neuroplasticity changes in the mechanism induced by Wen Shen Jian Pi intervention in spleen and kidney yang deficiency IBS-D rats.MethodsStudy one Therapeutic effect of warming the kidney and strengthening the spleen intervention on spleen-kidney yang deficiency IBS-D rat modelIn the previous study group,spleen and kidney yang deficiency IBS-D rats were treated with 0.3-0.5g/ml senna leaflet perfusion(4w)combined with simultaneous rectal dilatation 2w+clamp tail stimulation 2w+fixed binding 2w+glacial acetic acid injected into the colon for 2w.This experiment improved the modeling method,using lg/ml senna leaflet gavage 4w + avoid stress 10d duplicate spleen and kidney yang deficiency IBS-D rat model.Animals were randomly divided into normal control group,blank model group,Wenshen Jianpi high,middle and low dose groups.In addition to the control group,the remaining groups were given intragastric administration of senna for 4 weeks to induce diarrhea of spleen-kidney yang deficiency,and the control group was given the same dose of physiological saline.After 4 weeks,all groups except the control group started to avoid water stress,and the control group underwent false stress avoidance.On the sametime the high,middle,and low dosages of the traditional Chinese medicine group were administered with high,middle,and low concentrations of Wenshen Jianpi for 10d before completion,while the control group was with same dosages of water.The IBS-D model of spleen-kidney yang deficiency was evaluated from the aspects of morphology,stool condition,body weight changes,rectal temperature,weight-bearing swimming,grip strength,AWR,and pathological tissue,and the efficacy of Wenshen Jianpi Fang on this model was evaluated.Study two Effect of warming kidney and strengthening spleen Chinese herb intervention on CREB/BDNF/TrkB signaling pathway in IBS-D rat model of spleen-kidney yang deficiency type70 wistar rats were randomly divided into normal control group,blank model group,Wenshen spleen high,middle and low dose group,with 14 in each group.The model and intervention methods were the same as the study one.After the intervention,the expression of pCREB,BDNF,TrkB,ARC and C-FOS in colonic mucosa was detected by immunohistochemistry.The expression of BNDF in hippocampus was detected by Elisa.The expression of TrkB mRNA,ARC mRNA and C-FOS mRNA in hippocampus was detected by PCR.ResultsStudy one Therapeutic effect of Warming the Kidney and Strengthening the Spleen intervention on spleen-kidney yang deficiency IBS-D rat modelMorphologically,the model group showed symptoms of spleen-kidney yang deficiency,which were improved after intervention of traditional Chinese herb.There was significant difference in the number of defecation within 1 hour after treatment in the five groups(P<0.01).The number of defecation in the model group was higher in the lh group than in the normal group(P<0.01).There was a significant difference between High-dose Wenshen Jianspleen group and the model group(P<0.05)after treatment.Compared with the changes of the five groups after the 4w as well as after the treatment in the weight gain of the two groups at different periods,there was a significant difference(P<0.01).The model group,the high dose group,the medium dose group,and the low dose group weight,all of which were significantly lower than the normal group.Compared with the control group,the weight gain of the model group was slower.The weight gain of the high-dose Warming the Kidney and Strengthening the Spleen with the model group was significantly different(P<0.01).The high dose of Warming the Kidney and Strengthening the Spleen Fang intervention After the model,the weight of the rats increased significantly.As the results of rectal temperature,at 4w,there was a statistically significant difference between the five groups in rectal temperature(P<0.05).The rectal temperature in the normal group was significantly higher than that in the model group,with a significant difference(P<0.05).After treatment,there was a statistically significant difference between the five groups of rectal temperature(P<0.05).Further multiple comparisons showed that the rectal temperature in the normal group was significantly higher than that in the model group,Wenshen Jianpi Recipe middle dose group,Wenshen Jianpi low side In the dose group,the difference was statistically significant(P<0.05).There was no significant difference between the normal group and the high-dose Wenshen Jianpi group(P>0.05).The model group was significantly lower than the Wenshen Jianpi high dose group.The difference was statistically significant(P<0.05).At 4w,there was a statistically significant difference in the weight-bearing swimming between the five groups(P<0.05).Further multiple comparisons revealed that the difference between the normal group and the model group was statistically significant(P<0.05).After treatment,there was a significant difference in weight-bearing swimming between the five groups(P<0.05).There was no significant difference between the normal group and the high-dose Wenshen Jianpi group(P>0.05).The model group was significantly lower than the high-dose Wenshen Jianpi group,the difference was significant(P<0.05).At 4w,there was a significant difference in the grip strength between the five groups(P<0.05).Further multiple comparisons revealed that the difference between the normal group and the model group was statistically significant(P<0.05).After treatment,the difference in grip strength between the five groups was statistically significant(P<0.05).Further multiple comparisons revealed that the grip strength of the normal group was significantly higher than that of the Wenshen Jianpi Fang model group and Wenshen Jianpi Fang middle dose group.Wenshenjian There was significant difference between the low-dose group and the spleen group(P<0.05).There was no significant difference between the normal group and the high-dose Wenshen Jianpi group(P>0.05).The model group was significantly lower than the Wenshen Jianpi high-dose group.The difference was statistically significant(P<0.05).When assessing visceral hypersensitivity,the balloon was expanded to 60 mmHg,there was a significant difference between the model group,the low-dose group and the normal group(P<0.01),and there was a significant difference between the middle-dose group and the normal group(P<0.05).There was no difference between the high dose group and the normal group.When the balloon was expanded to 80mmHg,there was a significant difference between the model group,the middle dose group,the low dose group and the normal group(P<0.01).The difference between the high dose group and the model group was statistically significant(P<0.05).Histopathological findings did not show intestinal organic disease.Study two Effect of warming kidney and strengthening spleen decoction on the signals of arc and c-fos in the brain-intestinal CREB/BDNF/TrkB pathway of IBS-D rats with spleen-kidney yang deficiency odel of spleen and kidney yang deficiency.Immunohistochemical expression of CREB/BDNF/TrkB signaling in colonic tissueThe difference of pCREB protein in each group was statistically significant(P<0.05).The control group was significantly lower than the model group,low dose group and middle dose group,the difference was statistically significant(P<0,05).The difference between the control group and the high dose group was not The statistical significance(P>0.05)the model group was significantly higher than the low,middle,and high dose groups(P<0.05).There was no significant difference between the low dose group and the middle dose group(P>0.05).The difference between the low dose group and the middle dose group was significantly higher than that of the high dose group(P<0.05).The difference in BDNF between the five groups was statistically significant(P<0.05).The comparison between the two groups showed that the difference between the control group and the high-dose group was not statistically significant(P>0.05).The control group and the high-dose group were significantly lower than the model,the low-dose group and the middle-dose group.There was a significant difference between the groups(P<0.05).There was no significant difference between the model group and the low-dose group(P>0.05).The model group and the low-dose group were significantly higher than the middle-dose group.The difference was statistically significant(P<0.05).There were significant differences in the five groups of TrkB receptors(P<0.05),and the comparison between the two groups showed that the control group was significantly lower than the model group,the low-dose group and the middle-dose group,and the difference was statistically significant(P<0.05).There was no significant difference between the control group and the high-dose group(P>0.05).The model group was significantly higher than the low-dose groups,middle-dose groups,and high-dose groups.The difference was statistically significant(P<0.05).The difference was not statistically significant(P>0.05)between the low-dose group and the middle-dose group.The low-dose group and middle-dose group were significantly higher than the high-dose group.The difference was statistically significant(P<0.05).Expression of CREB/BDNF/TrkB pathway signals in hippocampusThe difference of BDNF protein concentration in the five groups was statistically significant(P<0.05).The comparison between the two groups showed that there was no significant difference between the control group and the high dose group(P>0.05).The control group and high dose group were significantly higher than the model group.The difference between the low-dose group and the middle-dose group was statistically significant(P<0.05).There was no significant difference between the model group and the low-dose group and the middle-dose group(P>0.05).The expression of TrkB mRNA in the five groups was significantly different(P<0.05).The control group was significantly lower than the model group,the low-dose group and the middle-dose group,the difference was statistically significant(P<0.05).The difference between the control group and the high-dose group was not statistical significance(P>0.05),the model group was significantly higher than the low,medium and high dose groups,the difference was statistically significant(P<0.05),the low dose group,the middle dose group was significantly higher than the high dose group,the difference was Statistical significance(P<0.05).Immunohistochemical expression of early immediate genes at the colon levelThe expression of Arc protein in the five groups was statistically significant(P<0.05).The control group was significantly lower than the other four groups(P<0.05).The model group and low dose group were significantly higher than the middle dose group and In the high-dose group,the difference was statistically significant(P<0.05).The middle-dose group was higher than the high-dose group,and the difference was statistically significant(P<0.05).The difference in c-fos expression between the five groups was statistically significant(P<0.05).The control group was significantly lower than the other four groups(P<0.05).The difference between the model group and the low-dose group was not statistically significant(P<0.05).the model group and the low-dose group were significantly higher than the middle-dose group and the high-dose group,the difference was statistically significant(P<0.05),the middle-dose group was higher than the high-dose group,the difference was statistically significant(P<0.05).Early expression of mRNA in the hippocampus at the early stageThe difference in arc between the five groups was statistically significant(P<0.05).There was no significant difference between the control group and the high-dose group(P>0.05).The control group and the high-dose group were significantly lower than the model group,the low-dose group,and the middle-dose In the group,the difference was statistically significant(P<0.05).The model group was significantly higher than the middle-dose group(P<0.05).There was no statistical difference between the model group and the low-dose group,the low-dose group and the middle-dose group.Significance(P>0.05).The difference of c-fos in each group was statistically significant(P<0.05):The control group was significantly lower than the other four groups(P<0.05):The model group was significantly higher than the low,medium,and high dose groups.There was statistical significance(P<0.05).The low-dose group and the middle-dose group were significantly higher than the high-dose group.The difference was statistically significant(P<0.05).Conclusion1 This study was reliable in replicating spleen and kidney yang deficiency IBS-D animal models.Warming the Kidney and Strengthening the Spleen can reduce the visceral hypersensitivity of IBS-D rats.Warming the Kidney and Strengthening the Spleen treatment effect is superior to high doses.2 IBS-D with spleen-kidney yang deficiency has activation of CREB/BDNF/TrkB signaling pathways on different levels of the brain and intestine,while high-dose Wenshen Jianpi can regulate abnormal expression of signaling molecules on the pathway and reduce visceral hypersensitivity,initially changing the neuroplasticity of spleen-kidney yang deficiency IBS-D to achieve therapeutic effects.3 IBS-D with spleen-kidney yang deficiency has early and immediate gene-mediated neuroplasticity at different levels of the brain and intestine,while high-dose Wenshen Jianpi can reduce visceral hypersensitivity by down-regulating the expression of early and immediate genes,initially changing the neuroplasticity of spleen-kidney yang deficiency IBS-D to achieve therapeutic effects. |
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