Polysaccharide of Alocasia cucullata Exerts Antitumor Effect by Regulating Bcl-2, Caspase-3 and ERK1/2 Expressions during Long-Time Administration / 中国结合医学杂志

OBJECTIVE@#To study the in vitro and in vivo antitumor effects of the polysaccharide of Alocasia cucullata (PAC) and the underlying mechanism.@*METHODS@#B16F10 and 4T1 cells were cultured with PAC of 40 µg/mL, and PAC was withdrawn after 40 days of administration. The cell viability was detected by cell counting kit-8. The expression of Bcl-2 and Caspase-3 proteins were detected by Western blot and the expressions of ERK1/2 mRNA were detected by quantitative real-time polymerase chain reaction (qRT-PCR). A mouse melanoma model was established to study the effect of PAC during long-time administration. Mice were divided into 3 treatment groups: control group treated with saline water, positive control group (LNT group) treated with lentinan at 100 mg/(kg·d), and PAC group treated with PAC at 120 mg/(kg·d). The pathological changes of tumor tissues were observed by hematoxylin-eosin staining. The apoptosis of tumor tissues was detected by TUNEL staining. Bcl-2 and Caspase-3 protein expressions were detected by immunohistochemistry, and the expressions of ERK1/2, JNK1 and p38 mRNA were detected by qRT-PCR.@*RESULTS@#In vitro, no strong inhibitory effects of PAC were found in various tumor cells after 48 or 72 h of administration. Interestingly however, after 40 days of cultivation under PAC, an inhibitory effect on B16F10 cells was found. Correspondingly, the long-time administration of PAC led to downregulation of Bcl-2 protein (P<0.05), up-regulation of Caspase-3 protein (P<0.05) and ERK1 mRNA (P<0.05) in B16F10 cells. The above results were verified by in vivo experiments. In addition, viability of B16F10 cells under long-time administration culture in vitro decreased after drug withdrawal, and similar results were also observed in 4T1 cells.@*CONCLUSIONS@#Long-time administration of PAC can significantly inhibit viability and promote apoptosis of tumor cells, and had obvious antitumor effect in tumor-bearing mice.

Yifei Jianpi recipe improves cigarette smoke-induced inflammatory injury and mucus hypersecretion in human bronchial epithelial cells by inhibiting the TLR4/NF-κB signaling pathway / 南方医科大学学报

OBJECTIVE@#To explore the mechanism of Yifei Jianpi recipe for improving cigarette smoke- induced inflammatory injury and mucus hypersecretion in cultured human bronchial epithelial cells.@*METHODS@#Serum samples were collected from 40 SD rats treated with Yifei Jianpi recipe (n=20) or normal saline (n=20) by gavage. Cultured human bronchial epithelial 16HBE cells were stimulated with an aqueous cigarette smoke extract (CSE), followed by treatment with the collected serum at different dilutions. The optimal concentration and treatment time of CSE and the medicated serum for cell treatment were determined with CCK-8 assay. The expressions of TLR4, NF-κB, MUC5AC, MUC7, and muc8 at both the mRNA and protein levels in the treated cells were examined with RT- qPCR and Western blotting, and the effects of TLR4 gene silencing and overexpression on their expressions were assessed. The expressions of TNF-α, IL-1 β, IL-6 and IL-8 in the cells were detected using ELISA.@*RESULTS@#At the optimal concentration of 20%, treatment with the medicated serum for 24 h significantly lowered the mRNA and protein expressions of TLR4, NF- κB, MUC5AC, MUC7, and MUC8 in CSE- exposed 16HBE cells, and these effects were further enhanced by TLR4 silencing in the cells. In 16HBE cells with TLR4 overexpression, the expressions of TLR4, NF-κB, MUC5AC, MUC7, and MUC8 were significantly increased after CSE exposure and were lowered following treatment with the medicated serum (P < 0.05). The medicated serum also significantly lowered the levels of TNF-α, IL-1β, IL-6 and IL-8 in CSE-exposed 16HBE cells (P < 0.05).@*CONCLUSIONS@#In the 16HBE cell model of chronic obstructive pulmonary disease (COPD), treatment with Yifei Jianpi recipe-medicated serum improves inflammation and mucus hypersecretion possibly by reducing MUC secretion and inhibiting the TLR4/NF-κB signaling pathway.

Total Saponin Fraction of Dioscorea Nipponica Makino Improves Gouty Arthritis Symptoms in Rats via M1/M2 Polarization of Monocytes and Macrophages Mediated by Arachidonic Acid Signaling / 中国结合医学杂志

OBJECTIVE@#To explore the mechanism of effects of total saponin fraction from Dioscorea Nipponica Makino (TSDN) on M1/M2 polarization of monocytes/macrophages and arachidonic acid (AA) pathway in rats with gouty arthritis (GA).@*METHODS@#Seventy-two Sprague Dawley rats were randomly divided into 4 groups (n=18 in each): normal, model, TSDN at 160 mg/kg, and celecoxib at 43.3 mg/kg. Monosodium urate crystal (MSU) was injected into the rats' ankle joints to induce an experimental GA model. Blood and tissue samples were collected on the 3rd, 5th, and 8th days of drug administration. Histopathological changes in the synovium of joints were observed via hematoxylin and eosin (HE) staining. The expression levels of arachidonic acid (AA) signaling pathway were assessed via real-time polymerase chain reaction (qPCR) and Western blot. Flow cytometry was used to determine the proportion of M1 and M2 macrophages in the peripheral blood. An enzyme-linked immunosorbent assay (ELISA) was used to detect interleukine (IL)-1 β, tumor necrosis factor-alpha (TNF-α), IL-4, IL-10, prostaglandin E2 (PGE2), and leukotriene B4 (LTB4).@*RESULTS@#HE staining showed that TSDN improved the synovial tissue. qPCR and Western blot showed that on the 3rd, 5th and 8th days of drug administration, TSDN reduced the mRNA and protein expressions of cyclooxygenase (COX)2, microsomal prostaglandin E synthase-1 derived eicosanoids (mPGES-1), 5-lipoxygenase (5-LOX), recombinant human mothers against decapentaplegic homolog 3 (Smad3), nucleotide-binding oligomerization domain-like receptor protein 3 (NALP3), and inducible nitric oxide synthase (iNOS) in rats' ankle synovial tissues (P<0.01). TSDN decreased COX1 mRNA and protein expression on 3rd and 5th day of drug administration and raised it on the 8th day (both P<0.01). It lowered CD68 protein expression on days 3 (P<0.01), as well as mRNA and protein expression on days 5 and 8 (P<0.01). On the 3rd, 5th, and 8th days of drug administration, TSDN elevated the mRNA and protein expression of Arg1 and CD163 (P<0.01). Flow cytometry results showed that TSDN decreased the percentage of M1 macrophages while increasing the percentage of M2 in peripheral blood (P<0.05 or P<0.01). ELISA results showed that on the 3rd, 5th, and 8th days of drug administration, TSDN decreased serum levels of IL-1 β, TNF-α, and LTB4 (P<0.01), as well as PGE2 levels on days 3rd and 8th days (P<0.05 or P<0.01); on day 8 of administration, TSDN increased IL-4 serum levels and enhanced IL-10 contents on days 5 and 8 (P<0.05 or P<0.01).@*CONCLUSION@#The anti-inflammatory effect of TSDN on rats with GA may be achieved by influencing M1/M2 polarization through AA signaling pathway.

Establishment of a reporter system for estimating activation of human hepatic stellate cells based on COL1A1 promoter and enhanced green fluorescent protein / 北京大学学报(医学版)

OBJECTIVE@#To establish a visual reporting system for evaluating the activity of collagen Ⅰ α 1 chain (COL1A1) gene promoter in immortalized human hepatic stellate cells, so as to estimate the activation status of the cells and provide a new cell model for the screening and study of anti-hepatic fibrosis drugs.@*METHODS@#The promoter sequence of human COL1A1 was amplified from the genomic DNA of human hepatocarcinoma cell line HepG2. Based on the pLVX-AcGFP1-N1 plasmid, the recombinant plasmid pLVX-COL1A1-enhanced green fluorescent protein (EGFP) was constructed, in which the enhanced green fluorescent protein gene expression was regulated by the COL1A1 promoter. The monoclonal cell line was acquired by stably transfecting pLVX-COL1A1-EGFP into the immortalized human hepatic stellate cell line LX-2 by the lentivirus packaging system and screening. The cell line was treated with transforming growth factor-β1 (TGF-β1) or co-treated with TGF-β1 and drugs with potential anti-hepatic fibrosis effects. The EGFP fluorescence intensity in cells was analyzed by the fluorescence microscope and ImageJ 1.49 software using a semi-quantitative method. The COL1A1 and EGFP mRNA were detected by reverse transcription real-time quantitative PCR (RT-qPCR), and corresponding proteins were detected by Western blot.@*RESULTS@#The recombinant plasmid pLVX-COL1A1-EGFP with the expression of EGFP regulated by COL1A1 promoter was successfully constructed. Kozak sequence was added to enhance the expression of EGFP, which was identified by double digestion and sequencing. The LX-2 monoclonal cell line LX-2-CE stably transfected with pLVX-COL1A1-EGFP was obtained. After co-treatment with TGF-β1 and 5 μmol/L dihydrotanshinone Ⅰ with potential anti-hepatic fibrosis effect for 24 h, the total fluorescence intensity and the average fluorescence intensity of LX-2-CE were lower than those in TGF-β1 single treatment group (P < 0.05), the intracellular mRNA and protein levels of COL1A1 and EGFP were also lower than those in the TGF-β1 single treatment group (P < 0.05).@*CONCLUSION@#A reporter system for estimating activation of hepatic stellate cells based on COL1A1 promoter regulated EGFP expression is successfully constructed, which could visually report the changes in COL1A1 expression, one of the activation-related markers of hepatic stellate cells, in vitro. It provides a new cell model for the screening and study of anti-hepatic fibrosis drugs.

Effect of miR-125b on T Cell Activation in Aplastic Anemia by Targetting B7-H4 / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of miR-125b on T cell activation in patients with aplastic anemia (AA) and its molecular mechanism.@*METHODS@#A total of 30 AA patients were enrolled in department of hematology, Binzhou Medical University Hospital from January 2018 to October 2021, as well as 15 healthy individuals as healthy control (HC) group. Peripheral blood mononuclear cells (PBMCs) were isolated, in which the levels of miR-125b and B7-H4 mRNA were detected by RT-qPCR. Immunomagnetic beads were used to separate naive T cells and non-naive T cells from AA patients and healthy people to detect the levels of miR-125b and B7-H4 mRNA. Lentivirus LV-NC inhibitor and LV-miR-125b inhibitor were transfected into cells, and T cell activation was detected by flow cytometry. The dual-luciferase reporter gene assay was used to detect the targetting relationship between miR-125b and B7-H4. RT-qPCR and Western blot were used to detect the levels of miR-125b, CD40L, ICOS, IL-10 mRNA and B7-H4 protein.@*RESULTS@#Compared with HC group, the expression of miR-125b was up-regulated but B7-H4 mRNA was down-regulated in PBMCs of AA patients (P <0.05), and the proportions of CD4+CD69+ T cells and CD8+CD69+ T cells in PBMCs of AA patients were higher (P <0.05). The expression of miR-125b was significantly up-regulated but B7-H4 mRNA was down-regulated in both naive T cells and non-naive T cells of AA patients (P <0.05), and non-naive T cells was more significant than naive T cells (P <0.05). Compared with NC inhibitor group, the expression of miR-125b was significantly decreased, the expression level of CD69 on CD4+ and CD8+ T cells in PBMCs was also significantly decreased, while the luciferase activity was significantly increased after co-transfection of miR-125b inhibitor and B7-H4-3'UTR-WT in the miR-125b inhibitor group (P <0.05). Compared with NC inhibitor group, the mRNA and protein levels of B7-H4 were significantly increased in the miR-125b inhibitor group (P <0.05). Compared with miR-125b inhibitor+shRNA group, the expression levels of CD69 on CD4+ and CD8+ T cells were significantly increased, and the levels of CD40L, ICOS and IL-10 mRNA were also significantly increased in the miR-125b inhibitor+sh-B7-H4 group (P <0.05).@*CONCLUSION@#MiR-125b may promote T cell activation by targetting B7-H4 in AA patients.

Significance of Tim-3 and Its Ligand Galectin-9 in Th1/Th2 Imbalance in Patients with Multiple Myeloma / 中国实验血液学杂志

OBJECTIVE@#To investigate the significance of Tim-3 and Galectin-9 in Th1/Th2 imbalance in patients with multiple myeloma (MM).@*METHODS@#55 newly diagnosed MM patients and 20 healthy controls were included. Flow cytometry was used to detect the expression of Tim-3 on CD4+T cells, the proportion of Th1, Th2, Tim-3+Th1 and Tim-3+Th2 cells in peripheral blood. ELISA was used to detect the levels of cytokines IFN-γ and IL-4 in serum, and PCR was used to detect the level of Galectin-9 mRNA. Then the correlations between Galectin-9 mRNA expression and Th-cell subsets and related cytokine levels, as well as the relationship between Tim-3+Th1/Tim-3+Th2 ratio and corresponding clinical features were analyzed.@*RESULTS@#Compared with the control group, the expression of Tim-3 on CD4+T cells in peripheral blood of MM patients was significantly increased (P<0.05), the proportions of Tim-3+Th1 cells, Tim-3+Th2 cells and Tim-3+Th1/Tim-3+Th2 ratio in MM patients were also increased (P<0.05), while the proportion of Th1 cells and Th1/Th2 ratio in MM patients were significantly decreased (P<0.05). The level of cytokine IFN-γ and IFN-γ/IL-4 ratio in MM patients were significantly decreased (P<0.05), while the level of cytokine IL-4 was increased (P<0.05). The mRNA levels of Galectin-9 in MM patients were significantly increased (P<0.05). The levels of Galectin-9 mRNA were positively correlated with Tim-3+CD4+T cells (r=0.663), Tim-3+Th2 cells (r=0.492) and IL-4 (r=0.470), while negatively correlated with IFN-γ (r=-0.593). The ratios of Tim-3+Th1/Tim-3+Th2 in MM patients were positively correlated with ISS stage (r=0.511), osteolytic damage (r=0.556) and chromosome abnormality (r=0.632).@*CONCLUSION@#These results suggest that Tim-3 and Galectin-9 are involved in Th1/Th2 imbalance in MM patients, and the high ratio of Tim-3+Th1/Tim-3+Th2 is associated with poor clinical prognosis.

Effect of wheat-grain moxibustion on Wnt/β-catenin signaling pathway in bone marrow cell in mice with bone marrow inhibition / 中国针灸

OBJECTIVE@#To observe the effect of wheat-grain moxibustion at "Dazhui" (GV 14), "Zusanli" (ST 36) and "Sanyinjiao" (SP 6) on Wnt/β-catenin signaling pathway in bone marrow cell in mice with bone marrow inhibition, and to explore the possible mechanism of wheat-grain moxibustion in treating bone marrow inhibition.@*METHODS@#Forty-five SPF male CD1(ICR) mice were randomly divided into a blank group, a model group and a wheat-grain moxibustion group, 15 mice in each group. The bone marrow inhibition model was established by intraperitoneal injection of 80 mg/kg of cyclophosphamide (CTX). The mice in the wheat-grain moxibustion group were treated with wheat-grain moxibustion at "Dazhui" (GV 14), "Zusanli" (ST 36) and "Sanyinjiao" (SP 6), 3 moxa cones per acupoint, 30 s per moxa cone, once a day, for 7 consecutive days. The white blood cell count (WBC) was measured before modeling, before intervention and 3, 5 d and 7 d into intervention. After intervention, the general situation of mice was observed; the number of nucleated cells in bone marrow was detected; the serum levels of interleukin-3 (IL-3), interleukin-6 (IL-6) and granulocyte macrophage colony stimulating factor (GM-CSF) were measured by ELISA; the protein and mRNA expression of β-catenin, cyclinD1 and C-Myc in bone marrow cells was measured by Western blot and real-time PCR method.@*RESULTS@#Compared with the blank group, the mice in the model group showed sluggish reaction, unstable gait, decreased body weight, and the WBC, number of nucleated cells in bone marrow as well as serum levels of IL-3, IL-6, GM-CSF were decreased (P<0.01), and the protein and mRNA expression of β-catenin, cyclinD1 and C-Myc was decreased (P<0.01). Compared with the model group, the mice in the wheat-grain moxibustion group showed better general condition, and WBC, the number of nucleated cells in bone marrow as well as serum levels of IL-3, IL-6, GM-CSF were increased (P<0.01, P<0.05), and the protein and mRNA expression of β-catenin, cyclinD1 and C-Myc was increased (P<0.05).@*CONCLUSION@#Wheat-grain moxibustion shows therapeutic effect on bone marrow inhibition, and its mechanism may be related to activating Wnt/β-catenin signaling pathway in bone marrow cells, improving bone medullary hematopoiesis microenvironment and promoting bone marrow cell proliferation.

The expression and significance of protease activated receptor 2 in ovarian epithelial carcinoma / 中华肿瘤杂志

Objective: To investigate the expression and significance of protease activated receptor 2 (PAR2) in ovarian epithelial carcinoma. Methods: PAR2 mRNA expression levels in 410 cases of epithelial ovarian carcinoma and 88 cases of human normal ovary were analyzed from cancer Genome Atlas (TCGA) database and tissue genotypic expression database (GTEx). Immunohistochemical (IHC) staining of PAR2 protein was performed in 149 patients with ovarian cancer who underwent primary surgical treatment at Cancer Hospital of Chinese Academy of Medical Sciences. Then the relationship between mRNA/protein expression of PAR2 and clinicopathological features and prognosis was analyzed. Gene functions and related signaling pathways involved in PAR2 were studied by enrichment analysis. Results: The mRNA expression of PAR2 in epithelial ovarian carcinoma was significantly higher than that in normal ovarian tissue (3.05±0.72 vs. 0.33±0.16, P=0.004). There were 77 cases showing positive and 19 showing strong positive of PAR2 IHC staining among the 149 patients, accounting for 64.4% in total. PAR2 mRNA/protein expression was closely correlated with tumor reduction effect and initial therapeutic effect (P<0.05). Survival analysis showed that the progression free survival time (P=0.033) and overall survival time (P=0.011) in the group with high PAR2 mRNA expression was significantly lower than that in the low PAR2 mRNA group. Multivariate analysis showed tumor reduction effect, initial therapeutic effect were independent prognostic factors on both progression-free survival and overall survival (P<0.05). The progression-free survival (P=0.016) and overall survival (P=0.038) of the PAR2 protein high expression group was significantly lower than that of the low group. Multivariate analysis showed PAR2 expression, initial treatment effect and chemotherapy resistance were independent prognostic factors on both progression-free survival and overall survival (P<0.05). Based on Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG), PAR2 target genes were mainly enriched in function related to intercellular connection, accounting for 40%. Gene enrichment analysis (GSEA) showed that the Wnt/β-catenin signaling pathway (P=0.023), the MAPK signaling pathway (P=0.029) and glycolysis related pathway (P=0.018) were enriched in ovarian cancer patients with high PAR2 mRNA expression. Conclusions: PAR2 expression is closely related to tumor reduction effect, initial treatment effect and survival of ovarian cancer patients. PAR2 may be involved in Wnt/β-catenin signaling pathway and intercellular connection promoting ovarian cancer invasion and metastasis.

Activation of metabotropic glutamate receptor 1 inhibits chronic intermittent hypoxia-induced carotid body plasticity in rats / 生理学报

The purpose of the present study was to explore the role of carotid body metabotropic glutamate receptor 1 (mGluR1) in chronic intermittent hypoxia (CIH)-induced carotid body plasticity. Sprague Dawley (SD) rats were exposed to CIH (6%-21% O2, 4 min/cycle, 8 h/day) for 4 weeks. The blood pressure of rats was monitored non-invasively by tail-cuff method under consciousness. RT-qPCR was used to examine the mRNA expression level of mGluR1 in rat carotid body. Western blot was used to detect the protein expression level of mGluR1 in rat carotid body. The role of mGluR1 in CIH-induced carotid body sensory long-term facilitation (sLTF) was investigated by ex vivo carotid sinus nerve discharge recording, and the carotid body sLTF was evoked by a 10-episode of repetitive acute intermittent hypoxia (AIH: 1 min of 5% O2 interspersed with 5 min of 95% O2). The results showed that: 1) CIH increased the systolic blood pressure (P < 0.001), diastolic blood pressure (P < 0.005) and mean arterial blood pressure (P < 0.001) of rats; 2) CIH decreased the mRNA and protein levels of mGluR1 in the rat carotid body (P < 0.01); 3) 4 weeks of CIH induced carotid body sLTF significantly, exhibiting as an increasing baseline sensory activity during post-AIH, which was inhibited by application of an agonist of group I metabotropic glutamate receptors, (S)-3,5-dihydroxyphenylglycine (DHPG), during sLTF induction (P < 0.005). In summary, these results suggest that activation of mGluR1 inhibits CIH-induced carotid body plasticity in rats.

Role of group II and III mGluRs in carotid body plasticity induced by chronic intermittent hypoxia in rats / 生理学报

The aim of the present study was to explore the role of group II and III metabotropic glutamate receptors (mGluRs) in carotid body plasticity induced by chronic intermittent hypoxia (CIH) in rats. Sprague Dawley (SD) rats were treated with CIH in Oxycycler A84 hypoxic chamber for 4 weeks, and the tail artery blood pressure was measured at the end of model preparation. RT-qPCR was performed to examine the mRNA expression levels of mGluR2/3/8 in rat carotid body. Carotid sinus nerve activity was detected by ex vivo carotid sinus nerve discharge recording technique, and acute intermittent hypoxia (AIH) was administered to induce carotid body sensory long-term facilitation (sLTF), in order to observe the role of group II and group III mGluRs in carotid body plasticity induced by CIH. The results showed that: 1) After 4 weeks of CIH exposure, the blood pressure of rats increased significantly; 2) CIH down-regulated the mRNA levels of mGluR2/3, and up-regulated the mRNA level of mGluR8 in the carotid body; 3) AIH induced sLTF in carotid body of CIH group. In the CIH group, activation of group II mGluRs had no effect on sLTF of carotid body, while activation of group III mGluRs completely inhibited sLTF. These results suggest that CIH increases blood pressure in rats, and group III mGluRs play an inhibitory role in CIH-induced carotid body plasticity in rats.

Effects of acupuncture on pulmonary function and airway smooth muscle spasm in asthma rats / 中国针灸

OBJECTIVE@#To observe the effects of acupuncture at "Kongzui" (LU 6) and "Yuji" (LU 10) on the latent period of inducing asthma, pulmonary function and the expression of endothelin-1 (ET-1) and metallothionein-2 (MT-2) in asthma rats, and to explore the possible mechanism of acupuncture in alleviating airway smooth muscle spasm and improving the acute attack of asthma.@*METHODS@#A total of 40 male SD rats of SPF-grade were randomly divided into a normal group, a model group, a medication group and an acupuncture group, 10 rats in each group. Except for the normal group, ovalbumin sensitization method was used to establish the asthma model in the other 3 groups. Salbutamol nebulization was adopted in the medication group, while acupuncture was applied at bilateral "Kongzui" (LU 6) and "Yuji" (LU 10) in the acupuncture group. The intervention was given once a day for 14 days in the two groups. The latent period of inducing asthma and pulmonary function were observed, the levels of ET-1 and tumor necrosis factor (TNF)-α in serum and bronchoalveolar lavage fluid (BALF) were detected by ELISA method, the morphology of the airway was observed by Masson staining, the ultrastructure of the airway smooth muscle was observed by transmission electron microscopy, the mRNA and protein expression of ET-1 and MT-2 in lung tissue was detected by real-time PCR and Western blot methods.@*RESULTS@#Compared with the normal group, in the model group, the latent period of inducing asthma was shortened (P<0.01); the airway resistance (RL) was increased while the dynamic compliance (Cdyn) was decreased (P<0.01, P<0.05); the levels of ET-1 and TNF-α in serum and BALF were increased (P<0.01); collagen fibers and collagen depositions were found around the bronchi, airway smooth muscle was thickened, the cell damage was severe and mitochondria were swollen; the mRNA and protein expression of ET-1 was increased while the mRNA and protein expression of MT-2 was decreased (P<0.01). Compared with the model group, in the acupuncture group, the latent period of inducing asthma was prolonged (P<0.05), the RL was decreased while the Cdyn was increased (P<0.01, P<0.05). Compared with the model group, in the medication group and the acupuncture group, the levels of ET-1 and TNF-α in serum and BALF were decreased (P<0.01, P<0.05); collagen fibers and collagen depositions around the bronchi were reduced, the thickened airway smooth muscle was lightened, the cell damage was improved; the mRNA and protein expression of ET-1 was decreased while the mRNA and protein expression of MT-2 was increased (P<0.01). Compared with the medication group, the mRNA expression of MT-2 was increased in the acupuncture group (P<0.05).@*CONCLUSION@#Acupuncture at "Kongzui" (LU 6) and "Yuji" (LU 10) can improve the pulmonary function and alleviate the airway smooth muscle spasm in rats with asthma. Its mechanism may be related to the down-regulation of ET-1 expression and up-regulation of MT-2 expression.

Study of the clinical significance of ETAR mRNA expression in high-grade serous ovarian cancer and the inhibitory effect of ETAR derived fusion polypeptide on cancer progression / 中华妇产科杂志

Objective: To investigate the clinical significance of endothelin A receptor (ETAR) expression in high-grade serous ovarian carcinoma (HGSOC). To design ETAR carboxyl terminal (ETAR-C) amino acids derived polypeptide and to study the inhibitory effect on ovarian epithelial carcinoma cells in vitro. Methods: (1) A total of 126 patients who received surgical treatment and were diagnosed with HGSOC by postoperative pathological examination in Central Hospital of Xuzhou from January 1, 2007 to December 31, 2017 were selected. All patients had completed clinicopathological data and follow-up data. Cancer tissue samples were collected and ETAR mRNA expression in HGSOC tissues was detected by reverse transcript-PCR. The clinical significance was analyzed. (2) ETAR-C fusion polypeptide was designed based on the sequence of carboxyl terminal amino acids of ETAR, expressed and purified in vitro. The effects of ETAR-C fusion polypeptide on migration and invasion ability of ovarian cancer SKOV3 and CAOV3 cells were detected by scratch test and invasion test, respectively. The effect of ETAR-C fusion polypeptide on chemosensitivity of cisplatin-resistant ovarian cancer SKOV3/cDDP and CAOV3/cDDP cells was determined by methyl thiazolyl tetrazolium (MTT) colorimetric assay. The effect of ETAR-C fusion polypeptide on β-arrestin-1 expression in ovarian cancer SKOV3 and CAOV3 cells was detected by western blot. Results: (1) The relative expression level of ETAR mRNA in HGSOC tissues was 18.6±5.1. Patients with HGSOC were divided into high ETAR mRNA expression (n=76) and low ETAR mRNA expression (n=50) with 61.7% as cut-off value analyzed by X-Tile software. High expression of ETAR mRNA was significantly correlated with abdominal water volume, platinum drug resistance, and cancer antigen 125 (CA125) value in HGSOC patients (all P<0.05), but was not related to the age of patients with HGSOC and the size of postoperative residual lesions (all P>0.05). The 5-year progression free survival rates were 18.4% and 28.0%, and the 5-year overall survival rates were 38.2% and 52.0% in HGSOC patients with high and low ETAR mRNA expression respectively, there were statistically significant differences (P=0.046, P=0.034). (2) The results of scratch test and invasion test showed that the scratch healing rate and cell invasion rate of SKOV3 or CAOV3 cells treated with endothelin-1 (ET-1) and ET-1+ETAR-C were respectively compared, and the differences were statistically significant (all P<0.05). MTT assay showed that the inhibition rates of ETAR-C fusion polypeptide treated in SKOV3/cDDP and CAOV3/cDDP cells were significantly higher than those of control cells after the addition of 4, 6, 8, 10, 12, and 24 μg/ml cisplatin (all P<0.05). Western blot analysis showed that the relative expression levels of β-arrestin-1 in SKOV3 or CAOV3 cells treated with ET-1 and ET-1+ETAR-C were 1.85±0.09 and 1.13±0.09 (SKOV3 cells), 2.14±0.15 and 1.66±0.12 (CAOV3 cells), respectively. The differences were statistically significant (all P<0.05). Conclusions: The prognosis of HGSOC patients with high expression of ETAR mRNA is significantly worse than those with low expression of ETAR mRNA. ETAR might be a new target for HGSOC treatment. The ETAR-C fusion polypeptide that interferes with the interaction of ETAR and β-arrestin-1 has good inhibitory effect on ovarian cancer cells in vitro, and might have clinical application potential.

miR-509-3p promotes oxidized low-density lipoprotein-induced apoptosis in mouse aortic endothelial cells / 中华危重病急救医学

OBJECTIVE@#To investigate the effect of microRNA-509-3p (miR-509-3p) on the apoptosis of atherosclerotic vascular endothelial cells.@*METHODS@#Mouse aortic endothelial cells (MAECs) were divided into normal control group, oxidized low-density lipoprotein (ox-LDL) group, miR-509-3p overexpression group, miR-509-3p overexpression control group, miR-509-3p inhibitor + ox-LDL group, and miR-509-3p inhibitor control + ox-LDL group. MAEC were induced with 100 mg/L ox-LDL for 24 hours, and then transfected with miR-509-3p overexpression/inhibitor and corresponding control for 48 hours. The miR-509-3p expression in MAECs exposed to ox-LDL was detected using real-time fluorescence quantitative polymerase chain reaction (RT-qPCR). Flow cytometry was used to detect the level of apoptosis, and cell counting kit (CCK-8) was used to detect the proliferation activity of MAECs. The direct gene targets of miR-509-3p were predicted using bioinformatics analyses and confirmed using a dual luciferase reporter assay. The expression of Bcl-2 mRNA and protein was detected by RT-qPCR and Western blotting, respectively.@*RESULTS@#Compared with the normal control group, miR-509-3p was significantly upregulated in ox-LDL-stimulated MAECs (1.68±0.85 vs. 1.00±0.30, t = 2.398, P < 0.05). After transfection of MAECs with miR-509-3p overexpression, the luciferase activity of the BCL2 3'UTR WT reporter gene was significantly lower than that of miR-509-3p overexpression control group (0.83±0.06 vs. 1.00±0.07, t = 4.531, P = 0.001). The luciferase activity of the BCL2 3'-UTR mutant (MUT) reporter gene was not significantly different from that of miR-509-3p overexpression control group (0.94±0.05 vs. 1.00±0.08, t = 1.414, P = 0.188). Compared with the normal control group and miR-509-3p mimics control group, the cell proliferation activity was decreased [(0.60±0.06)% vs. (1.00±0.09)%, (0.89±0.04)%, both P < 0.01], the percentage of apoptotic cells were increased [(23.46±2.02)% vs. (7.66±1.52)%, (10.40±0.78)%, both P < 0.05], and the mRNA and protein expression of Bcl-2 were significantly downregulated (Bcl-2 mRNA: 0.52±0.13 vs. 1.00±0.36, 1.10±0.19, Bcl-2 protein: 0.42±0.07 vs. 1.00±0.11, 0.93±0.10, both P < 0.01) in miR-509-3p overexpression group. Compared with the ox-LDL group, inhibition of miR-509-3p expression could increase the proliferation activity of MAECs induced by ox-LDL [(0.64±0.35)% vs. (0.34±0.20%)%, P < 0.05], and reduce the apoptosis rate [(13.59±2.22)% vs. (29.84±5.19)%, P < 0.01], and up-regulated the expression of Bcl-2 mRNA and protein in MAECs induced by ox-LDL (Bcl-2 mRNA relative expression: 0.82±0.09 vs. 0.52±0.10, Bcl-2 protein relative expression: 0.83±0.17 vs. 0.40±0.07, both P < 0.05).@*CONCLUSIONS@#Bcl-2 was one of the target genes of miR-509-3p. miR-509-3p can reduce the proliferation activity of endothelial cells, reduce the expression of Bcl-2, and promote cell apoptosis, thereby promoting the occurrence and development of atherosclerosis. Inhibition of miR-509-3p expression may be a potential therapeutic target for atherosclerosis.

Effect of Zhenwu Decoction on electrical remodeling of cardiomyocytes in heart failure via I_(to)/Kv channels / 中国中药杂志

This study aimed to investigate the underlying mechanism of Zhenwu Decoction in the treatment of heart failure by regulating electrical remodeling through the transient outward potassium current(I_(to))/voltage-gated potassium(Kv) channels. Five normal SD rats were intragastrically administered with Zhenwu Decoction granules to prepare drug-containing serum, and another seven normal SD rats received an equal amount of distilled water to prepare blank serum. H9c2 cardiomyocytes underwent conventional passage and were treated with angiotensin Ⅱ(AngⅡ) for 24 h. Subsequently, 2%, 4%, and 8% drug-containing serum, simvastatin(SIM), and BaCl_2 were used to interfere in H9c2 cardiomyocytes for 24 h. The cells were divided into a control group [N, 10% blank serum + 90% high-glucose DMEM(DMEM-H)], a model group(M, AngⅡ + 10% blank serum + 90% DMEM-H), a low-dose Zhenwu Decoction-containing serum group(Z1, AngⅡ + 2% drug-containing serum of Zhenwu Decoction + 8% blank serum + 90% DMEM-H), a medium-dose Zhenwu Decoction-containing serum group(Z2, AngⅡ + 4% drug-containing serum of Zhenwu Decoc-tion + 6% blank serum + 90% DMEM-H), a high-dose Zhenwu Decoction-containing serum group(Z3, AngⅡ + 8% drug-containing serum of Zhenwu Decoction + 2% blank serum + 90% DMEM-H), an inducer group(YD, AngⅡ + SIM + 10% blank serum + 90% DMEM-H), and an inhibitor group(YZ, AngⅡ + BaCl_2 + 10% blank serum + 90% DMEM-H). The content of ANP in cell extracts of each group was detected by ELISA. The relative mRNA expression levels of ANP, Kv1.4, Kv4.2, Kv4.3, DPP6, and KChIP2 were detected by real-time quantitative PCR. The protein expression of Kv1.4, Kv4.2, Kv4.3, DPP6, and KChIP2 was detected by Western blot. I_(to) was detected by the whole cell patch-clamp technique. The results showed that Zhenwu Decoction at low, medium, and high doses could effectively reduce the surface area of cardiomyocytes. Compared with the M group, the Z1, Z2, Z3, and YD groups showed decreased ANP content and mRNA level, increased protein and mRNA expression of Kv4.2, Kv4.3, DPP6, and KChIP2, and decreased protein and mRNA expression of Kv1.4, and the aforementioned changes were the most notable in the Z3 group. Compared with the N group, the Z1, Z2, and Z3 groups showed significantly increased peak current and current density of I_(to). The results indicate that Zhenwu Decoction can regulate myocardial remodeling and electrical remodeling by improving the expression trend of Kv1.4, Kv4.2, Kv4.3, KChIP2, and DPP6 proteins and inducing I_(to) to regulate Kv channels, which may be one of the mechanisms of Zhenwu Decoction in treating heart failure and related arrhythmias.

Effect of diosgenin on mTOR/FASN/HIF-1α/VEGFA expression in rats with non-alcoholic fatty liver disease / 中国中药杂志

The present study aimed to investigate the effect of diosgenin on mammalian target of rapamycin(mTOR), fatty acid synthase(FASN), hypoxia inducible factor-1α(HIF-1α), and vascular endothelial growth factor A(VEGFA) expression in liver tissues of rats with non-alcoholic fatty liver disease(NAFLD) and explore the mechanism of diosgenin on lipogenesis and inflammation in NAFLD. Forty male SD rats were divided into a normal group(n=8) fed on the normal diet and an experimental group(n=32) fed on the high-fat diet(HFD) for the induction of the NAFLD model. After modeling, the rats in the experimental group were randomly divided into an HFD group, a low-dose diosgenin group(150 mg·kg~(-1)·d~(-1)), a high-dose diosgenin group(300 mg·kg~(-1)·d~(-1)), and a simvastatin group(4 mg·kg~(-1)·d~(-1)), with eight rats in each group. The drugs were continuously given by gavage for eight weeks. The levels of triglyceride(TG), total cholesterol(TC), low-density lipoprotein cholesterol(LDL-C), alanine transaminase(ALT), and aspartate transaminase(AST) in the serum were detected by the biochemical method. The content of TG and TC in the liver was detected by the enzyme method. Enzyme-linked immunosorbent assay(ELISA) was used to measure interleukin 1β(IL-1β) and tumor necrosis factor α(TNF-α) in the serum. Lipid accumulation in the liver was detected by oil red O staining. Pathological changes of liver tissues were detected by hematoxylin-eosin(HE) staining. The mRNA and protein expression levels of mTOR, FASN, HIF-1α, and VEGFA in the liver of rats were detected by real-time fluorescence-based quantitative polymerase chain reaction(PCR) and Western blot, respectively. Compared with the normal group, the HFD group showed elevated body weight and levels of TG, TC, LDL-C, ALT, AST, IL-1β, and TNF-α(P<0.01), increased lipid accumulation in the liver(P<0.01), obvious liver steatosis, up-regulated mRNA expression levels of mTOR, FASN, HIF-1α, and VEGFA(P<0.01), and increased protein expression levels of p-mTOR, FASN, HIF-1α, and VEGFA(P<0.01). Compared with the HFD group, the groups with drug treatment showed lowered body weight and levels of TG, TC, LDL-C, ALT, AST, IL-1β, and TNF-α(P<0.05, P<0.01), reduced lipid accumulation in the liver(P<0.01), improved liver steatosis, decreased mRNA expression levels of mTOR, FASN, HIF-1α, and VEGFA(P<0.05, P<0.01), and declining protein expression levels of p-mTOR, FASN, HIF-1α, and VEGFA(P<0.01). The therapeutic effect of the high-dose diosgenin group was superior to that of the low-dose diosgenin group and the simvastatin group. Diosgenin may reduce liver lipid synthesis and inflammation and potentiate by down-regulating the mTOR, FASN, HIF-1α, and VEGFA expression, playing an active role in preventing and treating NAFLD.

Potentiating effect and mechanism of extract of Jingfang Granules on activation of macrophages / 中国中药杂志

This study aimed to explore the potentiating effect and mechanism of the extract of Jingfang Granules(JFG) on the activation of macrophages. The RAW264.7 cells were treated with JFG extract and then stimulated by multiple agents. Subsequently, mRNA was extracted, and reverse transcription-polymerase chain reaction(RT-PCR) was used to measure the mRNA transcription of multiple cytokines in RAW264.7 cells. The levels of cytokines in the cell supernatant were detected by enzyme-linked immunosorbent assay(ELISA). In addition, the intracellular proteins were extracted and the activation of signaling pathways was determined by Western blot. The results showed that JFG extract alone could not promote or slightly promote the mRNA transcription of TNF-α, IL-6, IL-1β, MIP-1α, MCP-1, CCL5, IP-10, and IFN-β, and significantly enhance the mRNA transcription of these cytokines in RAW264.7 cells induced by R848 and CpG in a dose-dependent manner. Furthermore, JFG extract also potentiated the secretion of TNF-α, IL-6, MCP-1, and IFN-β by RAW264.7 cells stimulated with R848 and CpG. As revealed by mechanism analysis, JFG extract enhanced the phosphorylation of p38, ERK1/2, IRF3, STAT1, and STAT3 in RAW264.7 cells induced by CpG. The findings of this study indicate that JFG extract can selectively potentiate the activation of macrophages induced by R848 and CpG, which may be attributed to the promotion of the activation of MAPKs, IRF3, and STAT1/3 signaling pathways.

Effect of multi-glycosides of Tripterygium wilfordii on renal injury in diabetic kidney disease rats through NLRP3/caspase-1/GSDMD pyroptosis pathway / 中国中药杂志

This study investigated the effect of multi-glycosides of Tripterygium wilfordii(GTW) on renal injury in diabetic kidney disease(DKD) rats through Nod-like receptor protein 3(NLRP3)/cysteine-aspartic acid protease-1(caspase-1)/gsdermin D(GSDMD) pyroptosis pathway and the mechanism. To be specific, a total of 40 male SD rats were randomized into the normal group(n=8) and modeling group(n=34). In the modeling group, a high-sugar and high-fat diet and one-time intraperitoneal injection of streptozotocin(STZ) were used to induce DKD in rats. After successful modeling, they were randomly classified into model group, valsartan(Diovan) group, and GTW group. Normal group and model group were given normal saline, and the valsartan group and GTW group received(ig) valsartan and GTW, respectively, for 6 weeks. Blood urea nitrogen(BUN), serum creatinine(Scr), alanine ami-notransferase(ALT), albumin(ALB), and 24 hours urinary total protein(24 h-UTP) were determined by biochemical tests. The pathological changes of renal tissue were observed based on hematoxylin and eosin(HE) staining. Serum levels of interleukin-1β(IL-1β) and interleukin-18(IL-18) were detected by enzyme-linked immunosorbent assay(ELISA). Western blot was used to detect the expression of pyroptosis pathway-related proteins in renal tissue, and RT-PCR to determine the expression of pyroptosis pathway-related genes in renal tissue. Compared with the normal group, the model group showed high levels of BUN, Scr, ALT, and 24 h-UTP and serum levels of IL-1β and IL-18(P<0.01), low level of ALB(P<0.01), severe pathological damage to kidney, and high protein and mRNA levels of NLRP3, caspase-1, and GSDMD in renal tissue(P<0.01). Compared with the model group, valsartan group and GTW group had low levels of BUN, Scr, ALT, and 24 h-UTP and serum levels of IL-1β and IL-18(P<0.01), high level of ALB(P<0.01), alleviation of the pathological damage to the kidney, and low protein and mRNA levels of NLRP3, caspase-1, and GSDMD in renal tissue(P<0.01 or P<0.05). GTW may inhibit pyroptosis by decreasing the expression of NLRP3/caspase-1/GSDMD in renal tissue, thereby relieving the inflammatory response of DKD rats and the pathological injury of kidney.

Effect of Erxian Decoction-containing serum on H_2O_2-induced proliferation and osteogenic differentiation of MC3T3-E1 cells via BK channels / 中国中药杂志

This study aimed to investigate the effects of Erxian Decoction(EXD)-containing serum on the proliferation and osteogenic differentiation of MC3T3-E1 cells under oxidative stress through BK channels. The oxidative stress model was induced in MC3T3-E1 cells by H_2O_2, and 3 mmol·L~(-1) tetraethylammonium(TEA) chloride was used to block the BK channels in MC3T3-E1 cells. MC3T3-E1 cells were divided into a control group, a model group, an EXD group, a TEA group, and a TEA+EXD group. After MC3T3-E1 cells were treated with corresponding drugs for 2 days, 700 μmol·L~(-1) H_2O_2 was added for treatment for another 2 hours. CCK-8 assay was used to detect cell proliferation activity. The alkaline phosphatase(ALP) assay kit was used to detect the ALP activity of cells. Western blot and real-time fluorescence-based quantitative PCR(RT-qPCR) were used to detect protein and mRNA expression, respectively. Alizarin red staining was used to detect the mineralization area of osteoblasts. The results showed that compared with the control group, the model group showed significantly blunted cell proliferation activity and ALP activity, reduced expression of BK channel α subunit(BKα), collagen Ⅰ(COL1), bone morphogenetic protein 2(BMP2), osteoprotegerin(OPG), and phosphorylated Akt, decreased mRNA expression levels of Runt-related transcription factor 2(RUNX2), BMP2, and OPG, and declining area of calcium nodules. EXD-containing serum could significantly potentiate the cell proliferation activity and ALP activity, up-regulate the protein expression of BKα, COL1, BMP2, OPG, and phosphorylated Akt, and forkhead box protein O1(FoxO1), promote the mRNA expression of RUNX2, BMP2, and OPG, and enlarge the area of calcium nodules. However, BK channel blockage by TEA reversed the effects of EXD-containing serum in promoting the protein expression of BKα, COL1, BMP2, OPG, and phosphorylated Akt and FoxO1, increasing the mRNA expression of RUNX2, BMP2, and OPG, and enlarging the area of calcium nodules. EXD-containing serum could improve the proliferation activity, osteogenic differentiation, and mineralization ability of MC3T3-E1 cells under oxidative stress, which might be related to the regulation of BK channels and downstream Akt/FoxO1 signaling pathway.

Effect and mechanism of Bovis Calculus on ulcerative colitis by inhibiting IL-17/IL-17RA/Act1 signaling pathway / 中国中药杂志

This study aimed to elucidate the effect and underlying mechanism of Bovis Calculus in the treatment of ulcerative colitis(UC) through network pharmacological prediction and animal experimental verification. Databases such as BATMAN-TCM were used to mine the potential targets of Bovis Calculus against UC, and the pathway enrichment analysis was conducted. Seventy healthy C57BL/6J mice were randomly divided into a blank group, a model group, a solvent model(2% polysorbate 80) group, a salazosulfapyridine(SASP, 0.40 g·kg~(-1)) group, and high-, medium-, and low-dose Bovis Calculus Sativus(BCS, 0.20, 0.10, and 0.05 g·kg~(-1)) groups according to the body weight. The UC model was established in mice by drinking 3% dextran sulfate sodium(DSS) solution for 7 days. The mice in the groups with drug intervention received corresponding drugs for 3 days before modeling by gavage, and continued to take drugs for 7 days while modeling(continuous administration for 10 days). During the experiment, the body weight of mice was observed, and the disease activity index(DAI) score was recorded. After 7 days of modeling, the colon length was mea-sured, and the pathological changes in colon tissues were observed by hematoxylin-eosin(HE) staining. The levels of tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), interleukin-6(IL-6), and interleukin-17(IL-17) in colon tissues of mice were detected by enzyme-linked immunosorbent assay(ELISA). The mRNA expression of IL-17, IL-17RA, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1β, CXCL1, CXCL2, and CXCL10 was evaluated by real-time polymerase chain reaction(RT-PCR). The protein expression of IL-17, IL-17RA, Act1, p-p38 MAPK, and p-ERK1/2 was investigated by Western blot. The results of network pharmacological prediction showed that Bovis Calculus might play a therapeutic role through the IL-17 signaling pathway and the TNF signaling pathway. As revealed by the results of animal experiments, on the 10th day of drug administration, compared with the solvent model group, all the BCS groups showed significantly increased body weight, decreased DAI score, increased colon length, improved pathological damage of colon mucosa, and significantly inhibited expression of TNF-α,IL-6,IL-1β, and IL-17 in colon tissues. The high-dose BCS(0.20 g·kg~(-1)) could significantly reduce the mRNA expression levels of IL-17, Act1, TRAF2, TRAF5, TNF-α, IL-6, IL-1β, CXCL1, and CXCL2 in colon tissues of UC model mice, tend to down-regulate mRNA expression levels of IL-17RA and CXCL10, significantly inhibit the protein expression of IL-17RA,Act1,and p-ERK1/2, and tend to decrease the protein expression of IL-17 and p-p38 MAPK. This study, for the first time from the whole-organ-tissue-molecular level, reveals that BCS may reduce the expression of pro-inflammatory cytokines and chemokines by inhibiting the IL-17/IL-17RA/Act1 signaling pathway, thereby improving the inflammatory injury of colon tissues in DSS-induced UC mice and exerting the effect of clearing heat and removing toxins.

Acteoside promotes autophagy and apoptosis of hepatoma cells by regulating JNK signaling pathway / 中国中药杂志

This study explored the molecular mechanism of acteoside against hepatoma 22(H22) tumor in mice through c-Jun N-terminal kinase(JNK) signaling pathway. H22 cells were subcutaneously inoculated in 50 male BALB/c mice, and then the model mice were classified into model group, low-dose, medium-dose, and high-dose acteoside groups, and cisplatin group. The administration lasted 2 weeks for each group(5 consecutive days/week). The general conditions of mice in each group, such as mental status, diet intake, water intake, activity, and fur were observed. The body weight, tumor volume, tumor weight, and tumor-inhibiting rate were compared before and after administration. Morphological changes of liver cancer tissues were observed based on hematoxylin and eosin(HE) staining, and the expression of phosphorylated(p)-JNK, JNK, B-cell lymphoma-2(Bcl-2), Beclin-1, and light chain 3(LC3) in each tissue was detected by immunohistochemistry and Western blot. qRT-PCR was performed to detect the mRNA expression of JNK, Bcl-2, Beclin-1, and LC3. The general conditions of mice in model and low-dose acteoside groups were poor, while the general conditions of mice in the remaining three groups were improved. The body weight of mice in medium-dose acteoside group, high-dose acteoside group, and cisplatin group was smaller than that in model group(P<0.01). The tumor volume in model group was insignificantly different from that in low-dose acteoside group, and the volume in cisplatin group showed no significant difference from that in high-dose acteoside group. Tumor volume and weight in medium-dose and high-dose acteoside groups and cisplatin group were lower than those in the model group(P<0.001). The tumor-inhibiting rates were 10.72%, 40.32%, 53.79%, and 56.44% in the low-dose, medium-dose, and high-dose acteoside groups and cisplatin group, respectively. HE staining showed gradual decrease in the count of hepatoma cells and increasing sign of cell necrosis in the acteoside and cisplatin groups, and the necrosis was particularly obvious in the high-dose acteoside group and cisplatin group. Immunohistochemical results suggested that the expression of Beclin-1, LC3, p-JNK, and JNK was up-regulated in acteoside and cisplatin groups(P<0.05). The results of immunohistochemistry, Western blot, and qRT-PCR indicated that the expression of Bcl-2 was down-regulated in the medium-dose and high-dose acteoside groups and cisplatin group(P<0.01). Western blot showed that the expression of Beclin-1, LC3, and p-JNK was up-regulated in acteoside and cisplatin groups(P<0.01), and there was no difference in the expression of JNK among groups. qRT-PCR results showed that the levels of Beclin-1 and LC3 mRNA were up-regulated in the acteoside and cisplatin groups(P<0.05), and the level of JNK mRNA was up-regulated in medium-dose and high-dose acteoside groups and cisplatin group(P<0.001). Acteoside promotes apoptosis and autophagy of H22 cells in mice hepatoma cells by up-regulating the JNK signaling pathway, thus inhibiting tumor growth.