| Objective:1.To establish gastric ulcer rat model with liver depression and spleen deficiencyby methods of muti-factors and acetic acid injection.2.To explain the occurrence mechanism of gastric ulcer rat model with liver depression and spleen deficiency and the therapeutic effect of Chaihuangweikuining decotion(CHWKN)in the whole viewing from immunology and molecular biology,so as to provide more advantageous and reliable experimental evidence for clinical application.Methods:1.Based on TCM etiology,to establish gastric ulcer with liver depression and spleen deficiency rat model applied methods of multi-factors combining acetic acid injection.2.The male SPF SD rats were randomly divided into 5 groups of 10 animals each:normal control group,gastric ulcer model group,gastric ulcer with liver depression and spleen deficiency group,CHWKN high-dosage group and CHWKN middle-dosage group.After the intervention for 14 days continuously,the drug groups underwent treatment for 15 days.On the last day,all the laboratory rats were sacrificed and their stomach and blood were immediately collected for testing through various methods.The enzyme linked immunosorbent assay(ELISA)was applied to collect the expression ofserumamylase activity(AMY),stomach gastrin-releasing(GAS),plasma serotonin(5-HT),and norepineohrine(NE);The expression ofHIF-1α,PHD2 and LDH-Ain the gastric tissue were detected by Western blot and the expression of miRNA-20b,miRNA-23a,miRNA-24,miRNA-26b,miRNA-199,miRNA-107,miRNA-210andmiRNA-210 were collected by the real-time PCR.Results:1.Compared with the normal control group,the level of serum AMY,serum GAS and plasma 5-HT and plasma NE in model group of gastric ulcer rats with liver depression and spleen deficiency syndromeand gastric ulcer model groupwere conspicuous decreased(P<0.05),and the plasma NE level was arresting increased(P<0.05).And the four detection paramrters of blood were in accordance with the tendency changes of liver depression and spleen deficiency syndrome,which meant succucessful models were established.2.Compared with the normal control group,The expression of PHD2in both gastric ulcer model group,gastric ulcer with liver depression and spleen deficiency group were down-reglulaing;and the expression of HIF-1α,LDH-A in both groups were up-reglulaing(p<0.05).Compared with thegastric ulcer with liver depression and spleen deficiency groupthe expression of PHD2 protein was tending to be up-regulating;and the expression ofHIF-1α,LDH-αprotein was tending to be down-regulating(p<0.05).3.Compared with the normal control group,the expression ofmiR-20b,miR-23 a,miR-210,miR-107 were up-regulating;and the expression of miR-24,miR-26b,miR-199,miR-424were down-reglulaing(p<0.05),Compared with thegastric ulcer with liver depression and spleen deficiency groupthe expression of miR-20b,miR-23a,miR-210,miR-107 in both CHWKN high-dosage group and CHWKN middle-dosage group were down-reglulaing,and the expression of miR-24,miR-26b,miR-199,miR-424were up-reglulaing(p<0.05).Conclusions:1.Model rats with gastric ulcer of liver depression and spleen deficiency syndrome were established successfully.and multi-factors combined acetic acidinjection applied in the research to establish gastric ulcer with liver depression and spleen deficiency rat model were proved to be stable and reliable.2.The content changes of HIF-1α,PHD2 and LDH-A in gastric ulcer of liver depression and spleen deficiency syndrome group were a hint of mircoenvironment,which means mircoenvironment was a pathogenic factor when gastric ulcer of liver depression and spleen deficiency syndrome occured,and CHWKN could correct the mircoenvironment change.3.when gastric ulcer of liver depression and spleen deficiency syndrome occured,the content of miRs showed related changes,and miRNAs could regulate their targeted protein,like HIF-1α,PHD2 and LDH-A,and our exprienced decoctionCHWKN couldregulate miRs and further affect proteins,this could be the working mechanism of CHWKN. |
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