ABSTRACT
OBJECTIVE：To study the effects of ferulic acid on t he proliferation ，invasion and apoptosis of HepG 2 hepatocelluar carcinoma cells. METHODS ：CCK-8 assay was used to screen the concentration of ferulic acid. Western blot assay was adopted to screen the optimal concentration of interleukin 6（IL-6）to induce HepG 2 cell model with high expression of phosphorylated signal transduction protein and activator 3（p-STAT3）protein. HepG 2 cell were divided into blank control group ， model group ，ferulic acid group （0.5 mmol/L）and positive control group （p-STAT3 inhibitor C 188-9，10 μmol/L）. Except for blank control group ，model group treated with IL- 6，while administration groups were treated with IL- 6 and relevant drugs. Cell survival rate ，invasion and apoptosis rate in early and late stage were detected by CCK- 8 assay，Transwell assay and Annexin V-FITC/PI double staining ，respectively. Western blot assay was used to detect the expression of p-STAT 3，caspase-3，ZBP-89 and vimentin proteins in each group. On the basis of the PDB protein database ，using 1BG1，a highly similar crystal structure of STAT3，as docking template ，using the region around Tyr 705 as the putative binding pocket ，the docking analysis of ferulic acid with STAT 3 protein was carried out. RESULTS ：It is selected to use 0.5 mmol/L ferulic acid intervention for 48 h as the follow-up experimental condition ；50 ng/mL IL- 6 was selected as the modeling condition. Compared with blank control group ，the number of cell invasion ，p-STAT3/STAT3 ratio and protein expression of vimentin were increased significantly in model group （P＜0.05 or P＜0.01），while late apoptosis rate and protein expression 20 of caspase- 3 were decreased significantly （P＜0.05 or P＜ 0.01）. Compared with model group ，cell survival rate ，the number of cell invasion ，p-STAT3/STAT3 ratio and protein expression of vimentin were d ecreased significantly in ferulic acid group and positive control group （P＜0.05 or P＜0.01）；early apoptotic rate （except for ferulic acid group ），late apoptotic rate，the protein expression of caspase- 3 and ZBP- 89（except for positive control group ）were increased significantly （P＜0.05 or P＜0.01）. The results of molecular docking showed that the carboxylic groups of ferulic acid could interact with 1.9 Å hydrogen bond of Asn 581 and 2.0 Å hydrogen bond of Lys 591，with binding energy of －4.4 kcal/mol. CONCLUSIONS ：Ferulic acid may inhibit the activity of p-STAT 3 by directly binding to the phosphorylation site of STAT 3；it may up-regulate the protein expression of caspase- 3 via STAT 3 dependent pathway ，or up-regulate the protein expression of ZBP- 89 via STAT 3 independent pathway and then down-regulate the protein expression of vimentin ，so as to inhibit the proliferation ，invasion and apoptosis of HepG 2 cells.
ABSTRACT
Cancer recurrence and severe side effects of currently being used chemotherapeutic agents reduce their clinical efficacy. Thus, there is a constant need to develop alternative anticancer drugs. Sustainable supply is an important challenge facing marine-based drug discovery. Primmorph, a 3D cell culture system, could provide a sustainable source to produce metabolites for anticancer drugs from marine sponges. In the present work, the anticancer activity of primmorph extracts and mesohyls of Negombata magnifica, Hemimycle arabica, Crella spinulata, and Stylissa carteri sponges was evaluated. Anti-proliferative activity was studied in terms of cytotoxicity, colony formation, cell cycle, and apoptosis. Migration was assessed by migration assay and matrix metalloproteinase activity. The expression of proliferation and migration-related genes was analyzed using real time PCR. Migration and proliferation activities of HepG2 cells were inhibited by treatment with primmorph extracts and mesohyls of N. magnifica, H. arabica, and C. spinulata. The mesohyl of S. carteri did not show any anticancer activity although the primmorph extract led to cell cycle arrest. Among the selected sponge species, the prim-morph extract of C. spinulata was the most promising anticancer agent regarding antiproliferative and antimigratory activities. In addition, primmorph extracts have the advantage of working under well-defined and controlled conditions, which allows the easy application as a bioreactor.
ABSTRACT
Objective To investigate the effect of CC chemokine receptor 4(CCR4) on sorafenib radiosensitivity and tumorigenesis of hepatocellular carcinoma cells in nude mouse models.Methods Western blot was used to detect the expression of CCR4 in hepatocellular carcinoma cell line.Lentivirus was utilized to construct PLC/PRF/5 and SMMC-7721 cell lines stably overexpressing and silencing CCR4,which were verified by Western blot.The influence of CCR4 on the radiosensitivity of hepatocellular carcinoma cells was assessed by plate clone formation assay.The effect of CCR4 on the tumorigenesis in hepatocellular carcinoma cells in vivo was evaluated by tumorigenesis assay in nude mice.Results CCR4 was highly expressed in highly-metastatic hepatocellular carcinoma cells and lowly expressed in hepatocellular carcinoma cells with low metastases.The PLC/PRF/5 and SMMC-7721 cells,which stably overexpressed and silenced CCR4,were successfully established.Overexpression of CCR4 reversed the inhibitory effect of sorafenib radiotherapy on PLC/PEF/5,whereas knockdown of CCR4 could increase the radiosensitivity of SMMC-7721 to sorafenib.Overexpressing CCR4 could promote the tumorigenicity of PLC/PEF/5,whereas knockdown of CCR4 could inhibit the tumorigenicity of SMMC-7721 in nude mice.Conclusion CCR4 overexpression significantly reduces the radiosensitivity of PLC/PRF/5 and increases the tumorigenicity in nude mice,whereas knockdown of CCR4 considerably increases the chemosensitivity and radiosensitivity of SMMC-7721 and suppresses the tumorigenicity in nude mice.
ABSTRACT
Vascular endothelial growth factor ( VEGF) is a mitogen that specifically acts on glyco-sylated cells of endothelial cells, enhances vascular permeability, induces angiogenesis, promotes the growth, invasion and metastasis of hepatocellular carcinoma cells. The prognosis of patients with high expres-sion of vascular endothelial growth factor is poor. Down-regulation of VEGF can significantly inhibit the ma-lignant proliferation of hepatocellular carcinoma ( HCC) and improve patients′ prognosis. This article sys-tematically reviewed the recent advances in VEGF expression and its role in promoting the invasion and me-tastasis of hepatocellular carcinoma cells as well as in VEGF-targeted anti-angiogenesis therapy for patients with HCC, aiming to provide some novel ideas to delay the disease progression in the patients.
ABSTRACT
Objective To investigate the influence of sorafenib on the expressions of B7-H3 and B7-H4 proteins in different human hepatocellular carcinoma cell lines,including HepG2,Hep3B,BEL-7402,BEL-7404,BEL-7405,QGY-7701,QGY-7703,SMMC-7721,MHCC97H,MHCC97L,HCCLM3 and HCCLM6.Methods Western blotting and MTT assay were used to check the influence of sorafenib on the expressions of B7-H3 and B7-H4 proteins in different human hepatocellular carcinoma cell lines and to test the inhibitory effect of sorafenib on different human hepatocellular carcinoma cell lines.Results Compared with normal human liver cells (HL-7702),the expressions of B7-H3 and B-H4 proteins in different human hepatocellular carcinoma cell lines were significantly up-regulated (P＜0.01).The cytotoxic activity IC50 values of sorafenib to Hep3B,BEL-7404,MHCC97H,HCCLM3 and HCCLM6 were 14.56,9.14,9.46,17.21 and 9.29 μmol/L respectively.After treating Hep3B,BEL-7404,MHCC97H,HCCLM3 and HCCLM6 with sorafenib at the doses of 5,10 and 20 μmol/L separately,the expressions of B7-H3 and B7-H4 proteins were strikingly down-regulated when compared with the control group (P＜0.01).Conclusion The overexpressions of B7-H3 and B7-H4 proteins in different human hepatocellular carcinoma cell lines are a common finding,which can influence tumor immune escape.It may be a new target for prevention and treatment of liver cancer in future.
ABSTRACT
Objective To explore the role and mechanism of miR-20a in the radiosensitivity of hepatocellular carcinoma (HCC). Methods The expression level of miR-20a in HCC cell lines and tissue specimens was detected by real-time fluorescent quantitative PCR.HCC cell line stably over-expressing miR-20a was constructed. The effect of miR-20a on HCC cell radiosensitivity was evaluate by cloning assay. The expression levels of Bcl-2,Caspase-3 and γ-H2AX proteins were quantitatively detected by Western blot. The target gene of the downstream regulation of miR-20a was predicted by bioinformatics analysis,which was further verified by dual luciferase reporter assay,real-time fluorescent quantitative PCR and Western blot. HCC cell line stably overexpressing miR-20a was transfected with pCDNA3. 0-PTEN to investigate the changes in the radiosensitivity of cells and to determine whether PTEN is a functional target gene for miR-20a-induced radioresistance of HCC. Results The expression levels of miR-20a was significantly up-regulated in HCC cell line and tissue specimens ( both P< 0. 05). After identical radiotherapy, the cell survival rate,the radioresistance was strengthened ( P< 0. 05),the expression of Bcl-2 was up-regulated, whereas the expression levels of Caspase-3 and γ-H2AX were down-regulated in the LV-miR-20a group compared with those in the blank and control groups ( WT and LV-con groups). Overexpression of PTEN could reverse the miR-20a-induced radioresistance. Conclusion miR-20a is up-regulated in the HCC cell lines and tissue specimens. Overexpression of miR-20a can promote the radioresistance of HCC cells. PTEN is a functional target gene for miR-20a-induced radioresistance of HCC,indicating that miR-20a/ PTEN site may be an effective molecular target associated with clinical radiotherapy for liver cancer.
ABSTRACT
[Objective]This study was conducted to examine the effects of dexmedetomidine on the proliferation and angiogenesis of MHCC97H and SMCC7721 human hepatocellular carcinoma(HCC)cell lines cultured in hypoxia condition in vitro,and investigated the possible mechanism involved.[Methods]MHCC97H and SMCC7721 human HCC cell lines under hypoxia culture condition were treated with presence or absence of dexmedetomidine(100 μmol/L). Cell viability,colony formation,vasculogenic mimicry(VM) formation were assessed. The effects of dexmedetomidine on α-2A adrenergic receptor(α2A),hypoxia induced factor-1a(HIF-1a),and vascular endothelial growth factor(VEGF)protein expression were evaluated with Western blot analysis.[Results]Cell proliferation assay and colony formation assay indicated that hypoxia obviously promoted the proliferation of MHCC 97H and SMCC7721 cells(CoCl2 group vs corresponding control group,the proliferation rate of MHCC97H and SMCC7721:Day 3,142.2%and 133.8%;Day 4,134.7%and 131.0%;Day 5,133.5%and 136.2%;all P<0.05),and VM formation assay suggested that hypoxia increased angiogenesis of MHCC97H and SMCC7721 cells. Whereas dexmedetomidine significantly inhibited the proliferation(Dex+CoCl2 group vs CoCl2 group,the proliferation rate of MHCC97H and SMCC7721:Day 3,55.7%vs 60.7%;Day 4,46.9%vs 58.1%;Day 5,46.4%vs 57.0%,all P<0.05)and angiogenesis of MHCC97H,SMCC7721 cells induced by hypoxia. Dexmedetomidine may exert these functions by activating α-2A adrenergic receptor,causing an decrease in HIF-1a and VEGF protein,while hypoxia activated HIF-1a and VEGF protein to promote the growth and angiogenesis of cells.[Conclusion]The findings provide evidence that hypoxia could promote the proliferation and angiogenesis of MHCC97H and SMCC7721 cells,while dexmedetomidine might inhibit these effects by down-regulating HIF-1a and VEGF protein expression through activatingα-2A adrenergic receptor.
ABSTRACT
Objective@#To investigate the effect of hepatitis B core antigen (HBcAg) in promoting the invasion of hepatitis B virus (HBV)-related hepatocellular carcinoma cell line HepG2.2.15 and the role of Toll-like receptor 4 (TLR4) in the mechanism.@*Methods@#TLR4 mRNA and protein expression in HepG2 cells and HepG2.2.15 cells was measured by reverse transcription real-time PCR and Western blot analysis, respectively. HepG2.2.15 cells were transfected with TLR4 specific small interfering RNA (siRNA) to silence TLR4 expression, and stimulated by recombinant HBcAg in culture. The invasion of cells was measured by Transwell invasion assay. The expression of TLR4 signaling pathway-related proteins in the cultured cells and proinflammatory cytokines in the supernatant was also determined. The student’s t-test, one-way ANOVA, and SNK-q test were used for statistical analysis.@*Results@#TLR4 mRNA and protein expression in HepG2.2.15 cells was significantly higher than that in HepG2 cells. TLR4 siRNA transfection remarkably down-regulated TLR4 expression in HepG2.2.15 cells. Inhibiting TLR4 expression and/or HBcAg stimulation did not affect the proliferation of HepG2.2.15 cells. However, HBcAg stimulation significantly increased the invasion ability of HepG2.2.15 cells and the secretion of proinflammatory cytokines [including interferon (IFN)-γ, interleukin (IL)-6, IL-8, and tumor necrosis factor (TNF)-α]. Inhibiting TLR4 expression significantly reduced HBcAg-induced cellular invasion. Meanwhile, HBcAg stimulation elevated the expression of MyD88 and TRIF in HepG2.2.15 cells. TLR4 silencing inhibited HBcAg-induced increase in the expression of MyD88, while it showed no significant impact on TRIF expression.@*Conclusion@#HBcAg can promote the invasion of HepG2.2.15 cells. The TLR4/MyD88 signaling pathway may be involved in this process by inducing the expression of proinflammatory cytokines.
ABSTRACT
Aim To investigate the effect of neferine on proliferation and invasion of human hepatocellular car-cinoma. Methods HepG2 and Bel-7402 cells were cultured in vitro with different concentrations of nefer-ine, then cell proliferation was observed by CCK-8 as-say; cell invasion was observed by transwell invasion assay; the protein expression of RhoA, RhoC and ROCK was detected by Western blot. Results CCK-8 results showed that neferine could significantly inhibit cell proliferation in a dose-dependent manner compared with the control group. Transwell invasion assay showed that cell invasion was significantly decreased with neferine 3 μmol · L-1 . Western blot results showed that RhoA, RhoC and ROCK protein expres-sion was decreased when neferine was co-incubated with hepatocellular carcinoma cells. Conclusion Nef-erine can inhibit proliferation and invasion of HepG2 and Bel-7402 cells, which is mediated mainly by the inhibition of RhoA, RhoC and ROCK protein expres-sion.
ABSTRACT
Objective To study the impact of different low concentrations of cerium oxide for hepatocellular carcinoma cell prolifera-tion.Methods Three different types of hepatoma cells (Huh7, HepG2,7721) were cultured,and added different concentrations of cerium oxide (0.005,0.01,0.05,0.1,1 μg/mL),of which the cell proliferation was detected by CCK8.The apoptosis-related genes was detected by qRT-PCR technology.The cell cycle was analyzed by flow cytometry.And the effect of low concentration cerium oxide on hepatocellular carci-noma cells tumorigenicity was confirmed by the nude mice experiments.Results CCK8 experiment showed that low concentrations of cerium oxide could promote proliferation of hepatocellular carcinoma cell, especially in concentration of 0.01μg/mL.The qRT-PCR showed that low concentration of cerium oxide could inhibit the apoptosis of hepatocellular carcinoma cell.The flow cytometry analysis had not found any effect of cerium oxide on cell cycle.The tumorigenicity experiments confirmed that low concentrations of cerium oxide could enhance the tumorigenic ability of hepatocellular carcinoma cell.Conclusion Low concentration of cerium oxide can significantly improve the proliferation of liver cancer cells.
ABSTRACT
Objective The cancer risk of patients with diabetes mellitus who are treated by metformin declines remarkably in comparison to patients receiving other drug therapies.The article was to investigate the relationship between antineopastic activity and fatty acid synthase (FASN) of metformin in human hepatocellular carcinoma cell(HCC) line HepG2. Methods HepG2 cells were treated with various concentrations of metformin( 0, 1, 5, 10, 15 mmol/L) for 24, 48 and 72 h respectively and cell growth was assessed by CCK-8 assay.Positive control(paclitaxel 10μg/mL) and negative control(metformin 0mmol/L) were set up simultaneously.After being treated with doses of metformin(0, 5, 10,15mmol/L) for 72h, protein expression levels of AMPKα、P-AMPKα、FASN、P-mTOR and P-Akt were measured by western blotting analysis and FASN mRNA expression levels were measured by RT-PCR. Results Being treated with vari-ous doses of metformin(1, 5, 10, 15 mmol/L) for 24, 48 and 72 h, the growth of HepG2 cells were inhibited by metformin in dose-dependent and time-dependent manner( P0.05) .FASN mRNA expression levels decreased significantly in all metformin-treated groups( P<0.05) . Conclusion Met-formin actitiviates AMPK, inhibits mTOR and downregulates FASN, which are implicated in its antineopastic activity on HCC.Although metformin inhibits mTOR activation, it is not involved in Akt upregulation through a negative loop.
ABSTRACT
Objective To detect the expression of SSX and to correlate it with clinical indicators of primary hepatocellular carcinoma (HCC).Methods The expression of SSX1-5 mRNA and SSX1 protein were respectively detected by RT-PCR and Western blot and immunohistochemistry staining.The relation between the expression of SSX mRNA and SSX1 protein with clinical indicators were analysed.Results SSX1,SSX2,and SSX3 mRNA were expressed in hepatocellular carcinoma cell lines BEL-7404,Hep G2,and SMMC-7721.In 26 HCC samples,SSX1-SSX5 mRNA was detectable in 53.8％,42.3％,50.0％,46.2％ and 26.9％.The expression of SSX1 mRNA was not related to serum AFP levels (P ＞0.05).Specific expression was both found in the normal group and the high value group.The expression rate of SSX1 mRNA was 85.7％ in the older group,which was higher than in the younger group (16.7％,P ＜ 0.05).The expression rate of SSX1 protein was 50％ in HCC tissues,which was not seen in the caner-adjacent or cirrhosis tissues.In 49 HCC paraffin tissue section samples,the expression rate of SSX1 protein was higher than that in caner-adjacent tissues (46.9％ vs 18.4％,P ＜ 0.05).The expression rate of SSX1 protein was 68.3％ in the large hepatocellular carcinoma group,which was higher than in the small hepatocellular carcinoma group (29.6％),(P ＜ 0.05).Conclusions SSX1 mRNA is expressed with a high percentage and specificity in HCC and their products are new potential promising targets for antigen-specific immunotherapy of HCC.The detection of SSX1 expression has the potential value for auxiliary diagnosis of HCC.
ABSTRACT
Objective To study the effects and preliminary mechanism of Stattic (Y705),an inhibitor of the signal transducer and activator of transcription-3 (STAT3),on the growth,migration,invasion and radiosensitization of hepatocellular carcinoma cell line Bel-7402.Methods Bel-7402 cells were divided to four groups:blank control group,Stattic treatment group,radiation group,and Stattic combined with radiation group.The cell growth and proliferation were detected by using CCK8 kit.The influence of Stattic on radiation sensitivity of Bel-7402 cells was determined by clone formation assay.The cell migration and invasion ability were tested by scratch migration assay and transwell assay,respectively.The protein expressions of STAT3,p-STAT3,Bax,Bcl-2,Caspase-3,Cleaved Caspase-3,MMP-2 and MMP-9 were quantified by Western blotting assay.Results Stattic significantly inhibited the growth of human hepatocellular carcinoma Bel-7402 cells with a dose-depended manner.The IC50 of Stattic after 48 h treatment was 2.5 μ mol/L.When 1.0 μmol/L Stattic was combined with 8 Gy X-rays,there was a synergistic effect in inhibition of cell proliferation with a inhibition rate of (15.00 ± 1.87) ％ (F =63.30,P ＜ 0.05).Scratch migration assay and transwell invasion assay showed that the migration and invasion abilities of the combination group were significantly reduced.In addition,compared with the radiation group,the SF2,D0and Dq values obtained from survival curve were decreased (t =4.20,6.92,9.32,P ＜0.05),the protein expressions of p-STAT3,MMP-2,MMP-9 were reduced (t =5.32,6.02,13.26,P ＜0.05),the protein expressions of Bax and Cleaved Caspase-3 were increased in the combination group(t =-7.82,-14.09,P ＜ 0.05),meanwhile the protein expressions of Bcl-2 was decreased (t =18.43,P ＜ 0.05).When the concentration of Stattic was 0.5 μmol/L,the radiation sensitization ratio at 2 Gy (SERSF2) was 1.22.Conclusions By inhibiting the activation of the p-STAT3 in Bel-7402 cells,stattic could induce cell apoptosis and increase the radiosensitivity,down regulate MMP-2 and MMP-9 and thereby reduce the invasion and migration of tumor cells.
ABSTRACT
Objective To investigate the antagonistic effect of wogonin in combination with 5-fluorouracil (5-FU) on the proliferation of human Hep-G2 hepatocellular carcinoma cells .Methods Human hepatocellular Hep-G2 cells were divided into experimental group (wogonin group ,5-FU group ,wogonin + 5-FU group) and control group .MTT method was used to eval-uate tumor cell proliferation in vitro ,flow cytometry analysis was used to evaluate tumor cell apoptosis .Results The results showed that the wogonin inhibited the proliferation of tumor cells at the concentrations of 5 ,10 ,20 and 40 μmol/L after 24 h and 48 h treatment respectively (P<0.05);5-FU also inhibited the proliferation of tumor cells at the concentrations of 5 ,10 , 20 and 40 mg/L after 24 h and 48 h treatment respectively (P< 0.05) .However when wogonin was combined with 5-FU (wogonin+5-FU group) ,an antagonistic effect was observed on tumor cell proliferation (P<0.05) .When cells were treated by wogonin+5-FU for 48 h ,the combined index (CI) value slowed a dose-dependent antagonistic effect (P<0.05) .Conclusion Wogonin has anti-tumor effect .However when wogonin was combined with 5-FU ,an obvious antagonistic effect on 5-FU′s anti-tumor action was observed .The underlying mechanism deserves further study .
ABSTRACT
Background and purpose: Primary liver cancer is the malignant tumor of liver cells or intrahepatic bile duct epithelium with familiar metastasis and postsurgical recurrence. The purpose of this study was to investigate the effects of miR-148a on the invasion and migration of hepatocellular carcinoma cells and the underlying mechanisms. Methods: The supernatant containing LV-miR-148a lentivirus particles was used to infect SMMC-7721 cells. The expression of miR-148a was determined by RT-PCR. Wound healing assay and transwell assay were performed to detect the effects of miR-148a on the invasion of hepatocellular carcinoma cells. Gelatin zymography assay was used to detect the effects of miR-148a on the enzyme activities of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9). The expression of MMP-2, MMP-9, E-cadherin and vimentin proteins was detected by Western blot assay. Results:RT-PCR showed the expression of miR-148a was upregulated in the infected SMMC-7721 cells. Transwell assay and wound healing assay showed ectopic expression of miR-148a suppressed cell migration and invasion abilities. miR-148a overexpression led to the decrease of the enzyme activities of MMP-2 and MMP-9 (P<0.05). Western blot assay showed that the protein expression of MMP-2, MMP-9 and vimentin proteins was signiifcantly decreased, the expression of E-cadherin had no changes. Conclusion:miR-148a is able to inhibit the migration and invasion of human SMMC-7721 cells in vitro, and the possible mechanisms may be related to decrease the enzyme activities of the MMP-2 and MMP-9 and the down regulation expression of MMP-2, MMP-9 and vimentin.
ABSTRACT
Objective To explore the inhibition effect and the possible mechanism of resveratrol on human hepatocellular carcinoma cell line Bel-7402 both in vitro and vivo.Methods Four Res drugs in the experiment group,the final concentrations were 12.5,25,50,100μmol/L,the control group at the same time set not containing Res drugs,MTT assay was used to measure the inhibition of resveratrol on Bel-7402.The expression of Bcl-2 was detected by RT-PCR and Western Blot.The levels of IL-2,IL-6,IL-12 and TNF-αwere detected by ELISA.Results Resveratrol inhibited Bel-7402 cell proliferation in dose and time manner,and influenced the expression of Bcl-2 mRNA and protein.At the same time,resveratrol inhibited the growth of tumor and improved the levels of IL-2,IL-6,IL-12 and TNF-α.Conclusion Resveratrol could inhibit Bel-7402 cell proliferation both in vitro and vivo, the possible mechanism may be that resveratrol could low down the expression of Bcl-2 and improve the levels of IL-2,IL-6,IL-12 and TNF-α.
ABSTRACT
Objective To explore the effects of hypoxia (1% O2) on the ability of cell invasiveness and expression of KAI1/CD82 in SMMC7721 hepatocellular carcinoma cells. Methods SMMC7721 hepatocellular carcinoma cells were cultured by hypoxia ( 1% O2) in vitro, and the ability of cell invasiveness was analyzed by cell invasion assay.Immunohistochemistry staining technique was used to evaluate the protein expression of KAI1/CD82. Results Cell invasion assay revealed that hypoxia enhanced the ability of invasiveness of hepatocellular carcinoma cells. In addition,KAI1/CD82 protein expression was positive in cultured SMMC7721 hepatocellular carcinoma cells, and it was located diffusedly in the cytoplasm and on the membrane. KAI1/CD82 protein expression was down-regulated when mediated by hypoxia; at the same time, it showed a time-effect relationship. Conclusion Hypoxia can enhance invasiveness of hepatocellular carcinoma cells. The down-regulation of KAI1/CD82 expression may play a certain role in those courses.
ABSTRACT
Objective: To explore the effects of hypoxia (1% O2) on the ability of cell invasiveness and expression of KAI1/CD82 in SMMC7721 hepatocellular carcinoma cells. Methods: SMMC7721 hepatocellular carcinoma cells were cultured by hypoxia (1% O2) in vitro, and the ability of cell invasiveness was analyzed by cell invasion assay. Immunohistochemistry staining technique was used to evaluate the protein expression of KAI1/CD82. Results: Cell invasion assay revealed that hypoxia enhanced the ability of invasiveness of hepatocellular carcinoma cells. In addition, KAI1/CD82 protein expression was positive in cultured SMMC7721 hepatocellular carcinoma cells, and it was located diffusedly in the cytoplasm and on the membrane. KAI1/CD82 protein expression was down-regulated when mediated by hypoxia; at the same time, it showed a time-effect relationship. Conclusion: Hypoxia can enhance invasiveness of hepatocellular carcinoma cells. The down-regulation of KAI1/CD82 expression may play a certain role in those courses.
ABSTRACT
Background: In a tumor, hypoxia-inducible factor-1α (HIF-1α) and vascular endothelial cell growth factor (VEGF) are induced to promote angiogenesis for the growth and metastasis of cells. There have been very few studies to examine in vivo relation between HIF-1α and VEGF during tumor progression. Objective: To study the relationship between HIF-1α and VEGF expressions under neovascularization induced by hepatocellular carcinoma cells (HepG2) implanted in nude mice. Methods: Male BALB/c-nude mice 8-10 weeks of age were used. A chamber was prepared on the dorsal skin in which HepG2 was transplanted to induce a tumor. On the day of the experiment, and on days 2, 7, and 14, microcirculation within the chamber was observed using fluorescence videomicroscopy. Based on the recorded video images, capillary vascularity (CV) was measured to examine tumor neovascularization. VEGF expression was measured in blood (serum) withdrawn, using enzyme immunoassay, while HIF-1α expression was measured on samples isolated from tumor tissue, using immunohistochemistry. Results: The measured CV significantly increased on day 7 and 14 compared to the aged-matched controls (p \< 0.05). HIF-1α markedly expressed on day 2, and the expression declined on day 7 and 14 post-inoculation. VEGF expression in serum increased more on day 7 and 14 than on day 2. HIF-1α expression decreased with the increase in VEGF expression from 2 to 14 days after HepG2 implantation, showing a reverse correlation. Conclusion: HIF-1α expression existed prior to both VEGF expression and neovascularization in the tumor. An inhibitor of HIF-1α might be a therapeutic agent for reducing neovascularization via adaptation to hypoxia in tumors.
ABSTRACT
Objective:To establish proteomic technique system of human hepatocellular carcinoma cell line QCY-7701. Method:Two different lysis buffer were taken to extract cell proteins. After two-dimensional gel electrophuresis (2-DE) and PDQuest analysis, 10 good-matched protein spots were chosen to be identified by MALDI-TOF-MS. Results:A steady 2-DE electrophregram of hepatocellular carcinoma cell line QGY-7701 was established. Compared with lysis bufferⅠ,protein quantities extracted from lysis bufferⅡincreased by 25%;protein spots in 2-DE gel increased by 32%. 9 out of 10 candidate protein spots were successfully identified (P