1.Protective Effect Of Diammonium Glycyrrhizin On Con A-induced Liver Injury In Mice 2.Antiplatelet Drugs Inhibit Tumor Metastasis By Interfering Platelet MiRNA In Tumor Microenvironment

Background and PurposeThe liver is the largest gland in the body,and it plays a very important role in human metabolism,blood coagulation,and bile production.Liver injury is a kind of disease,harmful to human health seriously.It is a pathological state existing in a variety of liver diseases.When the damage is over its compensatory ability,clinical and biochemical characteristic will change.The long-term existence of liver injury can lead to hepatic fibrosis,cirrhosis,and even liver cancer,so the prevention and treatment of liver injury is one of the main stages of clinical liver disease treatment.Diammonium glycyrrhizinate(DG)is a component extracted from licorice root,which has been used in China for thousands of years.Due to its better stability,higher solubility and stronger biological activity,it has been widely used in clinic.DG has a wide range of pharmacological activities,including anti-inflammatory,biooxidation,membrane protection and immunoregulation.Based on the above background,this project aims to study the liver protection of DG,and preliminarily discuss the liver protection mechanism of DG.Research MethodsFirstly,DG at the dose of 58.5 mg/kg equivalent to clinical dosage was orally administered to mice once daily for 7 consecutive days before Con A challenge.After that,blood samples were collected for AST and ALT detection,and histological analysis was carried out by hematoxylin-eosin staining to observe the histopathological features of liver tissue.Secondly,to test the direct effect of DG on AST and ALT enzyme activity,the co-incubation experiment of 18α-GA,18β-GA and serum with abnormal AST,ALT was conducted.Thirdly,bioinformatics analysis was used to predict the liver protection mechanism of DG.In the end,we used TUNEL staining,immunohistochemistry and western blot to verify the results of bioinformatics analysis and to further discuss the protective mechanism from the two aspects of apoptosis and inflammation.Research ResultsHematoxylin-eosin staining showed that DG pre-treatment prevented Con A-induced liver structural damage in ICR mice.We also observed the reduced serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)activities.Meanwhile the serum-drug incubation assay indicated that DG cannot directly attenuate ALT and AST levels.So we speculated that DG reduce the enzyme level by protecting liver cells.The protective mechanism of DG was explored.Bioinformatics analysis revealed that the protective effect of DG was closely related to apoptosis and inflammatory response.TUNEL staining,immunohistochemistry and western blot results showed that DG can alleviate double-stranded DNA rupture,immunological liver injury caused by liver cell apoptosis(reduce the expression of cleaved Caspase 3,cleaved PARP and BAX/BCL-2)and inflammatory reaction(reduce the expression of inflammatory factor,such as TGF-β1,COX-2,IL-6,TNF-α and INF-y).ConclusionsTaken together,DG pre-treatment can prevent Con A-induced liver injury in mice,and reduce ALT and AST activities,but cannot directly attenuate ALT and AST levels.And we found that DG can inhibit Con A-induced hepatic injury by inhibition of apoptosis and inflammation.Background and PurposeIn tumor microenvironment,tumor cells release the exosomes containing miRNA into the blood circulation,which can be swallowed by platelets.So the miRNA levels change in platelet,on the one hand,the changed miRNA in the platelet can adjust the translation of mRNA in platelets,which affect the biological function of platelet,such as inflammation,angiogenesis,tissue regeneration,etc.On the other hand,miRNA was transferred to the remote through the release of platelet particles,and play a role in receptor cells,such as influent endothelial cell function,promote angiogenesis and promote tumor cell invasion.At the same time,antiplatelet drugs have good antiplatelet activity and are used frequently in antitumor drugs.There are also studies showing that aspirin and salvia miltiorrhiza can play a role by regulating miRNA.Therefore,this topic aims to clarify that if the antiplatelet drugs such as aspirin and salvia miltiorrhiza of tanshinone and Dan phenolic acids B with antiplatelet activity could play the role of anti-tumor by adjusting the platelet miRNA.Research MethodsClinical data analysis;Detection of platelet releasing function(PF4,PPBP)screening drug concentration that can affect platelet releasing ability;Tumor cell exosomes and platelets were incubated in vitro,and drug intervention was given to detect RNA levels in platelets and platelet particles.Platelet RNA will be extracted from tumor-bearing colon cancer mice and colon cancer mice treated with tanshinol.q-RT-PCR technology will be used to verify differential miRNA of extracted platelet RNA.The targets miRNA will be identified by luciferase reporter gene.In vitro real-time dynamic observation system was used to observe the differential effects of platelets co-incubated with exosomes on tumor angiogenesis,invasion and proliferation.Platelets co-incubated with exosomes were incubated with tumor cells again,and tumor cells were centrifuged to prepare a tumor bearing mouse model of colon cancer.In vivo imaging of small animals was used to observe the effect on tumor metastasis.RNA knockout and overexpression confirmed its function.Research ResultsBy searching the database,it was found that patients with high expression of platelet production and activity-related regulatory genes in colon cancer cells had a shorter life span.By ultra high speed centrifugal extraction secrete body outside,appraisal results showed that secrete body outside the particle size of about 100 nm,presents the circular or elliptic,accord with literature describing secrete body form,and secrete body marker protein enrichment.ConclusionsThe expression of genes to regulate platelet production and activity differ between colon cancer patients and normal people.These differentially expressed genes are correlated with metastasis and survival in colon cancer patients significantly.The tumor cell-derived exosomes were successfully extracted by super-high-speed centrifugation.