Effect Of Shaoyao Gancao Decoction On Oddi Sphincter Nervous And C-kit Signal Transduction Pathway In Hypercholesterolemic Rabbits

Background:Sphincter of Oddi dysfunction（SOD）refers to the loss of normal physiological function of sphincter of Oddi（SO）,recurrent episodes of upper abdominal pain,jaundice,pancreatic enzymes or elevated liver enzymes for clinical manifestations of biliary and pancreatic diseases,and is also a common Sick,frequently-occurring disease.At present,the understanding of the pathogenesis of SOD is still in the initial stage,and there is no effective treatment for Western medicine.Shaoyao Gancao Decoction is one of the four classics of Chinese medicine^[1].The prescription in the Treatise on Febrile Diseases consists of white peony and licorice.White glutinous acid,licorice sweet,sour yin and yin can support the body meridians,alleviate the pain caused by various causes.Modern pharmacological research found that the main components of Radix Paeoniae Alba and Radix Glycyrrhizae are Paeoniflorin and Glycyrrhizin,which have antispasmodic and anti-inflammatory effects respectively.Therefore,traditional medicine uses Peony Medicago Decoction as a common prescription for relaxing smooth muscle and relieving its phlegm.Clinical use of Shaoyao Gancao Decoction for the treatment of SOD has achieved good results^[2],but the exact mechanism has not been studied clearly,and further research is needed.Objectives:The rabbit model of hypercholesterolemia was established by feeding cholesterol to purebred New Zealand rabbits.The neurological and c-kit signaling pathways in SO were used to study the damage of SOD in order to study the pathogenesis of SOD.Then,the drug was treated with Shaoyao Gancao Decoction,and the repairing effect of Shaoyao Gancao Decoction on SO nerve and c-kit signaling pathway was observed to study the mechanism of Shaozhi Gancao Decoction in treating SOD.Methods:Purebred New Zealand big ear rabbits are used,2 to 3 months old,weighing 1.8 to2.0 kg,regardless of gender.They were randomly divided into normal control group,hypercholesterolemia model group and Shaoyao Gancaotang treatment group.The control group was fed standard diet,and the model group and treatment group were fed standard diet+cholesterol.After successful modeling,the treatment group was treated with Shaoyao Gancao Decoction,and the control group and the model group were administered with normal saline.The feeding method was the same as before.After the end of the gavage treatment,the rabbits were sacrificed and the SO tissue was removed by laparotomy for experimental study.HE staining was performed to observe the pathological changes of SO tissue,and the nerve fibers were labeled by immunofluorescence.The expression of c-kit gene m RNA was detected by RT-PCR.Finally,the expression of c-kit protein and gap junction protein Cx43was detected by Western Blotting.the amount.In order to observe the pathogenesis of SOD and the repairing effect of the drug Gancao Decoction on the nerve and c-kit signaling pathway in SO.Results:1、The mucosal epithelial cells in the model group were disordered,the epithelial cells were severely deficient or shedding,the submucosal myometrial edema,and a large number of inflammatory cells infiltrated;the control epithelial cells were intact,no defect or shedding,no inflammatory muscles in the mucous membrane,no inflammatory cells Infiltration;the treatment group had a small amount of epithelial cell shedding and inflammatory cell infiltration,and there was no obvious edema in the submucosal muscle layer.2、The IOD value of smooth muscle surface markers in rabbit SO was the model group<treatment group<control group（P<0.05）.The IOD values of nerve fibers labeled with PGP9.5 were in the model group<treatment group<control group（P<0.05）.The SP-labeled SP neuron IOD values associated with smooth muscle contraction were model group>treatment group>control group（P<0.05）,and NO-NOS-labeled rabbit SO neuron IOD values were model group<treatment group<Control group（P<0.05）.3、In the control group,the expression of c-kit gene m RNA was the highest in SO,followed by the treatment group,and the model group was the least（P<0.05）.4、The expression levels of c-kit protein and Cx43 protein in each group had the same trend,and the expressions of the model group were the least,and the control group was the most,and the treatment group was between the two（P<0.05）.Conclusion:1、The results of HE staining showed that the morphological structure of SOD rabbit Oddi sphincter was obviously damaged,and the peony licorice soup had obvious repairing effect.2、The total number of nerve fibers in SOD rabbit Oddi sphincter was significantly reduced,the nerve fibers associated with smooth muscle relaxation were also significantly reduced,the number of smooth muscles was also reduced,and the morphology and structure of nerve fibers and smooth muscles were severely damaged.Peony licorice soup Can play a significant repair role.In addition,the number of SP nerve fibers in SOD rabbit Oddi sphincter increased,and the drug licorice can inhibit the proliferation of SP nerve fibers to some extent.3、The expression levels of protein and m RNA related to c-kit signaling pathway in SOD rabbit Oddi sphincter were significantly reduced.Paeonia lactose soup can significantly repair the signal pathway by enhancing the expression of related proteins.4、The expression of Cx43 protein in SOD rabbit Oddi sphincter was significantly reduced,indicating that the pathway of information transmission from Cajal interstitial cells to smooth muscle cells was destroyed.Paeonia sylvestris can enhance the relationship between cells by enhancing the expression of Cx43 protein.