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OBJECTIVE To explore the effect and mechanism of the alcoholic extract from Scabiosa comosa against hepatic fibrosis （HF）. METHODS Intragastrical administration of carbon tetrachloride was given to induce HF model. By observing the pathological changes in liver tissue， mRNA and protein expressions of HF indexes [α-smooth muscle actin （α-SMA）， collagen type Ⅰ] and phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt） pathway-related factors were detected， and the improvement effects and possible mechanism of low-dose， medium-dose and high-dose （50， 100， 200 mg/kg） of alcoholic extract from S. comosa on HF model rats were investigated. Drug-containing serum was prepared by intragastrical administration of alcoholic extract from S. comosa at a concentration of 1 800 mg/（kg·d） （calculated by the amount of raw material）. The effects of drug- containing serum of alcoholic extract from S. comosa on the expression of miRNA-21 were observed through the intervention of HSC-T6 cells with low， medium and high concentrations of drug-containing serum of alcoholic extract from S. comosa （diluted to 10%， 15%， 20%）. miRNA-21 mimics or inhibitors were used to transfect HSC-T6 cells， and the mRNA and protein expressions of factors related to the PI3K/Akt signaling pathway were detected. RESULTS The results of in vivo experiments showed that low， medium and high doses of alcoholic extract from S. comosa significantly ameliorated the histopathological changes in liver tissue of HF rats， and the percentage of collagen was significantly reduced （P＜0.01）； mRNA and protein expressions of the indicators related to HF as well as PI3K and Akt were significantly reduced （P＜0.01）， and mRNA and protein expressions of phosphatase and tensin homolog deleted on chromosome ten （PTEN） were increased in liver tissue of rats （P＜0.01）. The results of in vitro experiments showed that drug-containing serum of alcoholic extract from S. comosa significantly inhibited the expression of miRNA-21 at low， medium and high concentrations （P＜0.01）； whereas after transfection with miRNA-21 mimics， it was found that miRNA-21 mimics significantly increased mRNA and protein expressions of PI3K and Akt （P＜0.01）， while significantly decreased mRNA and protein expressions of PTEN （P＜0.01）； after transfection with miRNA-21 inhibitor， the changes of above indexes were opposite to the above results （P＜0.01）. CONCLUSIONS Alcoholic extracts of S. comosa may inhibit the PI3K/Akt signaling pathway by affecting the expression of miRNA-21， so as to achieve the effect of anti-hepatic fibrosis.
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Objective To investigate the role and mechanism of action of Scabiosa atropurea in inhibiting the proliferation of hepatic stellate cells using cell experiment. Methods A total of 20 Wistar rats were randomly divided into control group and administration group, with 10 rats in each group. The rats in the control group were given normal saline by gavage, and those in the administration group were given Scabiosa atropurea by gavage to prepare drug-containing serum. HSC-T6 cells were incubated with the serum from the control group (10%) or the low-, middle-, and high-dose serum containing Scabiosa atropurea (10%, 15%, and 20%, respectively). MTT assay was used to observe the effect of different drug concentrations on cells in different periods of time; flow cytometry was used to measure cell apoptosis; qRT-PCR and Western blot were used to measure the mRNA and protein expression levels of fibrosis markers (α-SMA, collagen Ⅰ) and PI3K/Akt signaling pathway-related factors in hepatic stellate cells (HSCs). A one-way analysis of variance was used for comparison between multiple groups, and the least significant difference t - test was used for further comparison between two groups. Results Compared with the control group, the low-, middle-, and high-dose serum containing Scabiosa atropurea groups had a significant reduction in the OD value of cells (all P < 0.05) and a significant increase in the overall apoptosis rate of cells (all P < 0.05). The results of qRT-PCR showed that compared with the control group, the low-, middle-, and high-dose serum containing Scabiosa atropurea groups had significant reductions in the mRNA expression levels of α-SMA, collagen Ⅰ, PI3K, and Akt and a significant increase in the mRNA expression level of PTEN (all P < 0.05); Western blot showed that compared with the control group, the low-, middle-, and high-dose serum containing Scabiosa atropurea groups had significant reductions in the protein expression levels of α-SMA, collagen Ⅰ, PI3K, Akt, and p-Akt and a significant increase in the protein expression level of PTEN (all P < 0.05). Conclusion The Mongolian medicine Scabiosa atropurea can inhibit the proliferation of HSC-T6 cells and promote their apoptosis, possibly by regulating fibrosis markers and the PI3K/Akt signaling pathway to exert an anti-liver fibrosis effect.
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Objective:To analyze the mechanism of Qiwei Qinggan Powder in the treatment of hepatic fibrosis by animal experiment and network pharmacology.Methods:A total of 50 rats were randomly divided into blank group, model group and low, medium and high dose of Qiwei Qinggan Powder groups with 10 rats in each group. Except the blank group, other groups were gavaged with 50% carbon tetrachloride peanut oil solution to prepare liver fibrosis model. Rats of low, medium and high dose of Qiwei Qinggan Powder groups were gavaged with 135, 270 and 405 mg/kg Qiwei Qinggan Powder 0.5% CMC-Na solution once a day for 10 weeks. The contents of serum GPT and GOT were detected by Wright's method, the content of ALP was detected by visible light colorimetry method, and the liver structure was observed by HE staining and Masson staining. The mRNA and protein expressions of α-SMA and collagen Ⅰ were detected by Q-PCR and Western blot from hepatic stellate cells. Flow Cytometry was used to detect the effect of Qiwei Qinggan Powder on hepatic stellate cells apoptosis. By searching for Traditional Chinese Medicines Integrated Database, Traditional Chinese Medicine Systems Pharmacology Database as well as literature retrieval, the active chemical components and targets of Qiwei Qinggan Powder were obtained. The targets of hepatic fibrosis were obtainable through OMIM and PubMed. By using Cytoscape 3.7.2, the Medicine-active components-Gene-Disease network was constructed. Obtaining Protein-Protein Interaction Networks screens the central target by using STRING, and using R language to retrieve Bioconductor online GO function enrichment and KEGG pathway enrichment analysis were carried out on the platform.Results:Compared with the model group, the levels of GFT, GOT and ALP in high, medium and low dose groups were decreased ( P<0.01). Compared with the control group, the mRNA levels of α- SMA (0.24 ± 0.12, 0.25 ± 0.12, 0.41 ± 0.15 vs. 1.00 ± 0.00), collagenⅠ (0.64 ± 0.24, 0.33 ± 0.13, 0.28 ± 0.11 vs. 1.00 ± 0.00)was decreased ( P<0.05 or P<0.01) of serum containing low, medium and high dose groups of Qiwei Qinggan Powder, and α-SMA (0.03 ± 0.01, 0.01 ± 0.00, 0.01 ± 0.00 vs. 0.04 ± 0.00), collagenⅠ (0.08 ± 0.01, 0.10 ± 0.01, 0.13 ± 0.01 vs. 0.18 ± 0.01) mRNA levels was decreased of serum containing low, medium and high dose groups ( P<0.01). A total of 35 active components, 196 targets, 3 740 disease targets and 170 disease common targets were screened out. 159 items were obtained by GO enrichment analysis; 43 signal pathways were obtained by KEGG enrichment analysis. Conclusion:Qiwei Qinggan Powder can promote HSCs apoptosis and treat HF through multi-component, multi-target and multi-channel therapy.
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OBJECTIVE：To study the improv ement effects of Mongolian medici ne eligen- 7 on hepatic fibrosis （HF）and its mechanism. METHODS ：Taking rat hepatic stellate cells HSC-T 6 as research object ，the cells were divided into model group （blank serum ）and low-dose ，medium-dose and high-dose groups of eligen- 7 containing serum （10％，15％ and 20％ eligen-7 containing serum ）. Transforming growth factor β solution（0.2 mg/mL）was added into the cells for 48 h to induce liver fibrosis model，and then added into the corresponding blank or drug-contained serum. The optical density （OD）of cells in each group was measured and inhibition rate of cell proliferation was calculated （after treated for 24，48 and 72 h）. The apoptotic rate and cycle distribution of cells were detected ；mRNA and protein expression of type Ⅰ collagen（Collagen Ⅰ），α-smooth muscle actin （α-SMA），phosphatidylinositol-3-kinase and protein-serine-threonine kinase （PI3K/Akt）signaling pathway related factors （PI3K， Akt，PTEN）were also detected （after treated for 24 h）. Wistar rats were further divided into blank group ，model group and low-dose，medium-dose and high-dose groups of eligen- 7（135，270，405 mg/kg），with 10 rats in each group. Blank group and model group were given 0.5％ sodium carboxymethyl cellulose solution intragastrically ；other groups were given relevant medicine intragastrically，once a day ，for consecutive 10 weeks. After last intragastric administration ，the pathomorphological changes of liver tissue were observed ；mRNA and protein expression of Collagen Ⅰ，α-SMA，PI3K，Akt and PTEN were detected in liver tissue. RESULTS ：Compared with model group ，OD value （except for medium-dose and high-dose groups of eligen- 7 containing serum）and the proportion of cells at S phase in administration groups were decreased significantly （P＜0.01）；late apoptotic rate ， early apoptotic rate （except for low-dose group ），total apoptotic rate and the proportion of cells at G 2/M phase increased significantly（P＜0.01）；mRNA and protein expression of Collagen Ⅰ，α-SMA，PI3K and Akt in cells and liver tissue were decreased significantly （P＜0.05 or P＜0.01），those of PTEN were increased significantly （P＜0.05 or P＜0.01）. CONCLUSIONS ： Eligen-7 shows the effect of anti-hepatic fibrosis ， themechanism of which may be related to regulating the activity of PI 3K/Akt signaling pathway and promoting the apoptosis of hepatic stellate cells.
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OBJECTIVE：To investigate the anti- hepatic fibrosis （HF）effects of Qiwei qinggan powder and explore its possible mechanism. METHODS ：Male Wistar rats were randomly divided into blank group ，HF model group ，Qiwei qinggan powder low-dose，medium-dose and high-dose groups [ 135，270，405 mg/（kg·d），by total amount of crude drugs] ，with 12 rats in each group. Except for blank group ，other groups were given 50％ CCl4-peanut oil solution intragastrically （2 mL/kg，twice a week ，for consecutive 8 weeks） to induce HF model. At same time ， blank group and model group were given constant volume of 0.5％ CMC-Na solution intragastrically ；administration groups were given relevant medicine intragastrically ，once a day ，for consecutive 8 weeks. General situation of rats were observedand liver morphology was observed after last administration and hepatic indexes were detected. The contents of liverfunction indexes （ALT，AST，ALP，HYP）in serum and the expression of α-SMA in hepatic tissue were determined ， and HE and Masson staining were performed to observe the histopathology. Using the difference multiple of expression quantity as the index ，TMT technology was used to screen the differentially expressed protein in medicine group （combining the liver tissue samples of Qiwei qinggan powder groups ）and HF model group. Uniprot-GOA database and KAAS ，KEGG mapper online tools were used to analyze GO and KEGG pathway enrichment. RESULTS ：The rats in the blank group were in good health ；the liver was bright red and smooth ，the liver lobules were intact ，no degeneration and necrosis ，inflammatory cell infiltration or fibrous tissue proliferation was found. Compared with blank group ，the rats in HF model group had poor diet ，depressed spirit ，disordered and lusterless fur ；the liver was dark red or yellow with rough surface ，hard texture ，inflammatory cell infiltration ，fiber tissue destruction ，bridge connection and so on ；the hepatic index ，the contents of liver function indexes and the expression of α-SMA were increased significantly （P＜0.05）. Compared with HF model group ，above symptoms of rats were improved to different extent in different dose groups of Qiwei qinggan powder ；hepatic index in Qiwei qinggan powder low-dose group ，the content of ALP in high-dose group ，the contents of ALT，AST and HYP and the expression of α-SMA in different dose groups were decreased significantly （P＜0.05）. A total of 42 differentially expressed proteins related to HF were screened ，of which 15 were up-regulated and 27 were down-regulated in expression，including fatty acid binding protein 4（FABP4），cholesterol 7α-hydroxylase（CYP7A1）. The results of enrichment analysis showed that the differentially expressed proteins were mainly enriched in extracellular space ，blood particles and other cell parts，involving the molecular functions of oxidoreductase activity and fatty acid binding ，the biological processes of the regulation of heterotypic cell adhesion ，protein activation cascade ，as well as retinol metabolism ，arachidonic acid metabolism ，PPAR and other signal pathway. CONCLUSIONS ：Qiwei qinggan powder can reduce the hepatic index ，ALT，AST，ALP and HYP contents in serum ，down-regulate the expression of α-SMA，improve the degree of inflammation and fibrosis of liver tissue ，and have a certain protective effect on rats. The anti-HF mechanism of it involves multiple targets and signal pathways ，such as FABP 4， CYP7A1 and PPAR.
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Objective:To study the protective effects of Danhong injection ( DH) on myocardial damage induced by doxorubicin ( DOX) in Lewis tumor bearing mice. Methods:The model of Lewis lung cancer in mice was established by underarm injecting tumor cells, and then randomly divided into four groups:the model control group, DOX group, DH group and DH+DOX group. After the experiment, myocardial and tumor tissue were separated from Lewis tumor bearing mice, and the excised tumors were weighted. The activities of lactate dehydrogenase ( LDH) , creatine kinase ( CK) , manganese superoxide dismutase ( SOD) , catalase ( CAT) and glu-tathione peroxidase ( GPx) , and the content of malondialdehyde ( MDA) were determined by a colorimetric method. Flow cytometry was used to determine the levels of apoptosis, reactive oxygen species (ROS) and mitochondrial membrane potential (△Ψm). Re-sults:Compared with that in the model control group, a significant decrease of tumor weight was shown in both DOX group and DH+DOX group (P<0. 01). DH had no significant influence on the anticancer function of DOX. The activity of LDH and CK, and the ap-optosis in myocardium cells significantly increased (P<0. 01). Compared with DOX group, the activities of LDH and CK, and the ap-optosis significantly decreased in DH+DOX group (P<0. 01). The activities of △Ψm, SOD, CAT and GPx significantly increased (P<0.05orP<0.01). ThecontentofMDAandROSgenerationbothdecreased(P<0.01).Conclusion:DHhasnosignificantin-fluence on the antitumor effect of DOX. The combination of DH and DOX shows cadioprotective effect on the myocardial damage through improving mitochondrial antioxidant defense capacity, ameliorating oxidative stress and maintaining △Ψm homeostasis.