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Objective: To investigate the effect of Clerodendrum bungei-containing serum on liver cancer MHCC97-H cells and its possible mechanism from the perspective of phosphatidylinositol 3-kinase(PI3K)/protein kinase (Akt) signaling pathway. Method: The medicinal serum of 15% C. bungei was used to treat MHCC97-H cells. The effect of serum containing C. bungei on cell proliferation was observed by cell counting kit-8(CCK-8) method, in order to select the best time and concentration. The apoptosis was detected by Annexin V-FITC/PI double staining method. Western blot was used to detect the posphatase and tensin homologous gene deleted from chromosome 10 in key proteins (PTEN), phosphoprotein kinase B (p-Akt) and phosphatidylinositol 3-kinase (PI3K)-related protein expression of PI3K/Akt signaling pathway. Real-time PCR was used to detect C. bungei-containing serum on cells for 72 h after activation of nuclear factor-activated B cell kappa light chain(NF-κB) and tumor necrosis factor-α (TNF-α) mRNA expression. Result: The results of CCK-8 showed an inhibitory effect of the C. bungei-containing serum on the proliferation of tumor cells in a dose and time-dependent manner. Among them, the high-dose group had the most obvious inhibitory effect, and the maximum inhibition rates at 24, 48,72 h were 28%, 32%, and 43%, respectively. The results of flow cytometry showed that with the increase of drug-containing serum concentration, the cell growth was observed. The inhibition rate of cells was increased to different degrees, and the inhibition effect was significantly increased in the 72 h intervention group (PC. bungei-containing serum group was 19.48% and 19.72%, compared with the blank group. The difference was significant (PC. bungei-containing serum (PPC. bungei-containing serum could down-regulate the expression of NF-κB and up-regulate the expression of TNF-α mRNA (PConclusion: The medicinal serum of C. bungei can effectively inhibit the proliferation of MHCC97-H hepatoma cells and promote its apoptosis, which may be related to the PI3K/Akt signaling pathway and its key factors.
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Objective: To detect the effects of dicentrine on the migration and invasion of human hepatocellular carcinoma MHCC97-H cells and potential mechanisms. Methods: MHCC97-H cells were cultured in vitro and cultured at logarithmic growth stage. The cells were treated with 5, 10, 25, 50, 100 μmol/L dicentrine for 24, 48, 72 h, and cell proliferation was determined by MTT assay. The cells were treated with 5, 10, 25 μmol/L dicentrine for 24 h, the cell adhesion, cell migration, cell invasion, gene expression of VEGF, MMP-2 and MMP-9, and protein expression of p-JAK2 and p-STAT3 were determined by MTT, scratch, Transwell, real-time qPCR, and western blot assay, respectively. Results: The cell viability of MHCC-97H cells treated with 25 μmol/L dicentrine at 48 and 72 h, and 50 and 100 μmol/L dicentrine at 24, 48, and 72 h was decreased significantly, which was significantly different from that of the control group (P < 0.05 or P < 0.01). Compared with control group, cell adhesion ability, wound healing ability, cell penetration modulus, gene expression of VEGF, MMP-2 and MMP-9, and protein expression of p-JAK2 and p-STAT3 of MHCC-97H cells treated with 5, 10 and 25 μmol/L dicentrine at 24 h were decreased significantly (P < 0.05 or P < 0.01). Conclusion: Dicentrine can inhibit the migration and invasion of human hepatocellular carcinoma MHCC97-H cells, which is related to the down-regulation of VEGF, MMP-2, and MMP-9 gene expression and inhibition of JAK2/STAT3 signaling pathway activation.
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Objective To investigate the inhibitory effect of Glycyrrhizin in MHCC97-H cell line in vitro and explore the relevant mechanism.Methods MHCC97-H cells were cultured in vitro and treated with Glycyrrhizin in different concentrations and then cell viability was assayed at different time points.The concentration and time were selected with 50％ cell viability.MHCC97-H cell plate clone formation assay and invasion-migration experiment were also performed to study the tumor-suppressor efficacy of Glycyrrhizin.Acridine orange staining was used to evaluate the formation of autophagic vacuoles.Meanwhile,3-MA and Atg7-siRNA were both employed to avoid the autophagy activation in MHCC97-H cells and cell viability was reassessed.Western-blot was carried out to study the expression of autophagic proteins of LC3B,p-mTOR and p-ERK1/2.Results It showed Glycyrrhizin significantly inhibited MHCC97-H cell viability and the concentration and time at 50％ cell viability were 2 mmol/L and 48 h respectively.Clone number in Glycyrrhizin group was significantly smaller than that in the control group (176.7 ± 14.5 vs.410.0 ± 32.1).Invasion-migration rate was also lower in Glycyrrhizin group compared with the control group (41.0％ ±3.8％ vs.100％).Autophagic vacuoles was increased in MHCC97-H cells when treated with Glycyrrhizin and expression of LC3B-Ⅱ was enhanced and LC3B-Ⅱ/I Ratio was increased,at the same time degradation of P62 was accelerated.Reduced p-mTOR in concurrence with upregulated p-ERK1/2 could be observed in MHCC97-H cells administered with Glycyrrhi-zin.Cell groups additionally treated with 3-MA or Atg7-siRNA exerted higher cell viability (64.3％ vs.45.9％ and 67.7％ vs.47.1％,respectively).Conclusion Glycyrrhizin can induce excessive autophagy in hepatocellular carcinoma cells to cause autophagic cell death and exhibit great potential in clinical application.
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Objective To study the effects of Yiqi Huayu Jiedu Prescription on the migration ability of MHCC97-H and the expressions of CXCL12, CXCR4 and CXCR7; To discuss its relevant mechanism of action. Methods Setting Sorafenib as a positive control, CCK-8 method was used for determining the effects of Yiqi Huayu Jiedu Prescription on the cell proliferation of MHCC97-H and the optimum concentration. Scratch assay was used to observe the migration ability of MHCC97-H. The protein expressions of CXCL12, CXCR4 and CXCR7 were detected by Western blot after 24 h of medicine intervention. Results Yiqi Huayu Jiedu Prescription and Sorafenib can inhibit the cell proliferation of MHCC97-H , and the inhibitory concentration was 0.095 g/mL and 10 μmol/mL at 24 hours. Yiqi Huayu Jiedu Prescription can inhibit migration ability of MHCC97-H. The protein expressions of CXCL12, CXCR4 and CXCR7 in hepatocellular carcinoma cells decreased after the action of Yiqi Huayu Jiedu Prescription. Conclusion Yiqi Huayu Jiedu Prescription can inhibit MHCC97-H cell proliferation and migration, which may be realized by down-regulating chemokine axis of CXCL12/CXCR4/CXCR7.
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ABSTRACT:Objective To investigate the effects of total flavones of oldenlandia diffusa (FOD)on epithelial-mesenchymal transition in hepatocellular cancer cell line MHCC97-H.Methods TGF-β1 induced EMT in routinely cultured liver cancer cell line MHCC97-H;then MHCC97-H cell was divided into 5 groups:normal control group, TGF-β1 group,TGF-β1 + FOD group,TGF-β1 + 5-FU group,and TGF-β1 + FOD + 5-FU group.After 48 h of treatment,the invasion ability of MHCC97-H cell was detected by Transwell;the proteins of E-cadherin and vimentin were determined by Western blot.Results Compared with the normal form of MHCC97-H cell line,the cell had obvious long fusiform after TGF-β1 induction,and the invasion ability enhanced (P = 0.02 ).But after treatment,the invasion ability of MHCC97-H cell decreased in FOD group and 5-FU group compared with that in TGF-β1 group (P = 0.03,P = 0.02 ),and decreased more significantly in FOD + 5-FU group (P = 0.01 ).The expression of E-cadherin at the protein level decreased significantly (P = 0.01 )in TGF-β1 group,which was abolished in FOD group (P =0.03 )and 5-FU group (P = 0.02 ).The expression of vimentin at the protein level increased significantly (P =0.01)in TGF-β1 group,which was abolished in FOD group (P =0.04)and 5-FU group (P =0.03)and more obviously in FOD+5-FU group (P =0.01).Conclusion FOD can reverse the invasion of MHCC97-H cells in EMT induced by TGF-β1 through decreasing the expression of E-cadherin protein and inhibiting the epithelial-mesenchymal transition of MHCC97-H cell.