Evaluation Of The Efficacy Of Shipi Xiaoshui Cream In The Treatment Of Malignant Ascites And Its Mechanism Of Actio

1 BackgroundMalignant ascites(MA),also known as cancerous ascites,is the abnormal increase of peritoneal fluid caused by the invasion of peritoneum by Malignant tumor cells.It is one of the common severe complications in the late stage of various malignant tumors.The recurrence of malignant ascites and the unbearable abdominal distension and abdominal pain caused by it have become important factors affecting the quality of life of patients with advanced malignant tumors.The recurrence of malignant ascites and the unbearable abdominal distension and abdominal pain caused by it have become the important reasons affecting the quality of life of patients with advanced malignant tumors.At present,the mechanism of malignant ascites has not been completely clear,and thougn the therapeutic methods are numerous,the effect is not ideal.The prescription of shipi xiaoshui cream is derived from the integrated traditional Chinese and western oncology department of china-japan friendship hospital.It is a prescription of clinical experience in the treatment of malignant ascites.The prescription is based on the treatment principle of invigorating qi and invigorating spleen,warming Yang and benefiting water,and activating qi and activating blood.The curative effect is confirmed by clinic.Experimental studies have also shown that it can significantly reduce the amount of ascite and promote the apoptosis of tumor cells.However,there has been no in-depth study on its pharmacological mechanism of "multi-component-multi-target-multi-pathway" in vivo.Therefore,this study used holistic pharmacodynamics,metabolomics,network pharmacology and other methods to explore the efficacy and possible mechanism of the treatment of malignant ascites with shenpi xiaoshui cream2 ObjectiveBased on the inheritance of the clinical classic prescription,the research on the development of the new medicine with the prescription of "shi pi xiao shui" as a cream type was carried out.The research on the efficacy evaluation of the external treatment of malignant ascites with"shi pi xiao shui" cream and the systematic research on metabolomics provided the experimental basis for the development of this preparation as a traditional Chinese medicine new medicine for the treatment of malignant ascites.3 Methods3.1 The pharmacodynamic evaluation of the external treatment of malignant ascites with Shipixiaoshui creamThe ascites model of H22 hepatocellular carcinoma and EAC were used and three kinds of concentration of shipixiaoshui cream were adopted.The abdominal circumference and weight of mice were measured,the abdominal water volume of mice was detected by b-ultrasound,the urine volume was measured.3.2 Study on the mechanism of external treatment of malignant ascites with shipixiaoshui creamUsing the ascites model of H22 liver cancer,the content of Na+,K+,Cl-in mouse urine were determined by biochemical analysis.ELISA was used to detect renin,Angll,antidiuretic hormone(AVP),ALD in serum renin-angiotensin-aldosterone system(RASS).By analyzing the adjustment effect of SPX on urine(Cl-Na+,K+)and plasma reninangiotensin system(RAS)Angll and vasopressin(AVP),immunohistochemical detection of kidney AQP1 expression,ELISA to detect the expression of AQP1 in ascites,system evaluation from the Angle of the liquid water regulating spleen away on the other hand cream play to the function mechanism of the treatment effect of malignant ascites.The mechanism of the effect of SPX cream on malignant ascites from the perspective of water regulation was systematically evaluated.The ascites model of H22 hepatocellular carcinoma was applied.The weights of organs in each group were compared,the peripheral blood images of mice were analyzed by automatic blood cell analyzer,the number of tumor cells in malignant ascites was measured.Flow cytometry was used to detect the number of CD4+T lymphocytes CD8+T lymphocytes Regulatory T cells(Treg)in peripheral blood.Intravenous injection of evens blue dye to detect peritoneal permeability.Serum levels of the cytokines interleukin-2(il-2)and il-10 were determined by ELISA.ELISA was used to detect the expression level of VEGF,mmp-2,IL-2.TNF-α and INF-γ,IL-6 in ascites.The mechanism of the effect of shipi xiaoshui cream on malignant ascites from the perspective of immune regulation was evaluated systematically.3.3 Biological effect of SPX cream for treating malignant ascites based on metabolomicsNon-target metabolomics based on uhplc-qtof-ms was used to scan and detect the components of endogenous metabolites in mice urine in positive and negative ion mode.OPLS-DA(orthogonal partial least square method-discriminant analysis)was used to detect the effect of SPX cream on the contents of potential biomarkers in the urine of mice with malignant ascites,and the mechanism of SPX cream intervention in the treatment of malignant ascites was discussed from the perspective of metabolic pathway.4 Results4.1 Pharmacodynamic evaluation of the external treatment of malignant ascites with SPX creamBoth the ascites model and the ascites tumor model of H22 hepatocellular carcinoma were used,the results showed that the body weight and abdominal circumference of mice in the model group increased rapidly and ultrasound showed a large amount of effusion in the abdominal cavity,and decreased urine volume in model mice,which showed the model was built.Compared with mice in the model group,the medium dose and high dose of SPX ointment could slow down the weight and ascites growth trend of the mice,reduce the amount of abdominal water,and increase the amount of urine,the difference was statistically significant(P<0.05,P<0.01).The effect of reducing ascites was not observed with low dose of SPX cream.The effect of medium-and high-dose SPX cream on reducing abdominal water volume and increasing urine volume is similar to that of diuretic hydrochlorothiazide.4.2 Study on the mechanism of treating malignant ascites with SPX creamCompared with the normal group,the concentrations of Na+K+Cl-ions in the urine of H22 mouse hepatoma ascites model were all increased,the serum levels of PRA,Ang Ⅱ,ALD ADH were all increased,and the expression of AQP1 in the kidney was increased.The SPX could reduce the concentration of Na+K+cl-ion in the urine of mice(P<0.05),reduce the concentration of PRA Ang II ALD ADH in the serum(P<0.05),and reduce the expression of AQP1 in the kidney.Hydrochlorothiazide can reduce the concentration of ALD ADH and the expression of AQP1 in the kidney.No significant difference was found in the expression of AQP1 in ascites superscript.The results of tumor cell count in ascites showed that there was no significant difference in the total number of tumor cells and the survival rate in ascites in each group.No significant difference was found in peritoneal permeability detected by tail vein injection of evens blue dye.No significant difference was found in peritoneal permeability detected by tail vein injection of evens blue dye.The detection of VEGF content in ascites showed that hydrochlorothiazide and SPX cream could reduce the VEGF content in ascites of H22 mouse hepatoma ascites model compared with the model group4.3 The effect of SPX on metabolism in H22 mouse hepatoma ascites modelIn the model of malignant ascites of H22 liver cancer,53 metabolites were found to be related to the incidence of malignant ascites,mainly involving amino acid metabolism,vitamin B6 metabolism,purine metabolism and other related substances.It has a callback effect on 37 kinds of substances,mainly through regulating the metabolic pathway of amino acids,increasing the excretion of ammonia in urine,improving the metabolic disorder caused by malignant ascites,and playing the role of treating ascites.5 ConclusionThe medium dose and high dose of the SPX cream can reduce the amount of ascite in H22 mouse hepatoma ascites model and ehrlich ascites carcinoma bearing mice.Medium dose was the best,comparable to diuretic hydrochlorothiazide.It can reduce the Na+ K+ cl-ion content in the urine of mice with H22 mouse hepatoma ascites model,reduce the concentration of PRA Ang Ⅱ ALD ADH in the serum,and reduce the overexpression of AQP1 in the kidney,indicating that it plays a diuretic effect by regulating the RAAS system,thus promoting the excretion of ascites,reducing the amount of abdominal water and correcting electrolyte disorders.The SPX cream can increase the weight of thymus and spleen of mice,increase the percentage of lymphocytes,increase the proportion of CD4+T lymphocytes,and significantly increase the level of il-2 in serum and ascites,indicating that it plays a certain anti-tumor role by positively regulating the immune function,thus slowing down the formation of ascites.It can alleviate the symptoms of malignant ascites to some extent,and can change the metabolic disorder caused by malignant ascites,which may be achieved by regulating the metabolic disorder of amino acid metabolic pathway,and its therapeutic mechanism can be revealed from the perspective of metabolic pathway.