ABSTRACT
Objective To investigate the molecular mechanism of epithelial-mesenchymal transition (EMT) induced by activated vascular endothelial growth factor receptor-1 ( VEGFR-I ) in cell line MHCC97-H.Methods MHCC97-H cells were cultured in DMEM with 1％ fetal bovine serum (control group),10 μmol/L PP2 (PP2 group),10 μmol/L PBS (PBS group),50 μmol/L VEGF-B (VEGF-B group),l0μmol/L PP2 and 50 μmol/L VEGF-B (PP2 +VEGF group),10 μmol/L PBS and 50 μmol/L VEGF-B (PBS + VEGF-B group),respectively.Protein expressions of epithelial marker E-cadherin,α-catenin and mesenchymal marker vimentin and N-cadherin were detected by Western blot.The expression sites of E-cadherin,α-catenin and mesenchymal marker vimentin and N-cadherin were detected by cell immunofluorescence.The ability of invasion and migration of cell line MHCC97-H were assessed by cell invasion and migration test.All data were analyzed by the t test.Results The expressions of E-cadherin,α-catenin,vimentin and N-cadherin were 3.23 +0.76,3.01 ±0.25,3.01 +0.22 and 2.63 +0.40 in the control group,4.18 +0.32,3.29 +0.11,4.85 +0.36 and 3.02 +0.52 in the PP2 group,2.83 +0.65,3.03 +0.27,1.37 ±0.24 and 2.98 ±0.36 in the PBS group,2.06 ±0.15,2.84 ±0.76,5.79 ± 0.38 and 5.54 ± 0.28 in the VEGF-B group,6.12 ± 0.08,5.45 ± 0.37,3.36 ± 0.42 and 3.26 ±0.13 in the PP2 + VEGF-B group and 1.36 ±0.54,1.26 ±0.45,4.05 ±0.17 and 1.05 ±0.33 in the PBS +VEGF-B group.There was a significant difference in the expressions of E-cadherin and α-catenin between the PP2 +VEGF-B group and the VEGF-B group (t =7.625,9.931,P ＜ 0.05 ).The expressions of vimentin and N-cadherin in the PP2 + VEGF-B group were significantly lower than those in the VEGF-B group (t =12.001,11.910,P ＜ 0.05).Six hours after the treatment with VEGF-B,the numbers of MHCC97-H migrated were 19 ± 1,5 ± 2and 16 ± 1 in the VEGF-B group,PP2 + VEGF-B group and PBS + VEGF-B group,respectively.The number of MHCC97-H cells migrated in the VEGF-B group was greater than that in the PP2 ± VEGF-B group ( t =13.566,P ＜ 0.05 ).The number of MHCC97-H cells passed through the Boyden chamber was 4 + 2,which was significantly less than 16 ± 1 of the VEGF-B group (t =12.350,P ＜0.05).Conclusion EMT induced by activated VEGFR-1 was mediated via c-Src kinase signal transduction in MHCC91-H cell line,and c-Src may be a potential target to interfere the invasion and migration of hepatic cancer cells.
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Objective To explore the effect of RNA interference on the bionomics of Cathepsin L of hepatoma carcinoma cells. Methods In this study, the experimental group was the Cathepsin L RNA interference group. Control groups were the normal blank hepatoma carcinoma cell group (the blank group) and the Cathepsin LRNA interference blank group (the fluorescence control group). Observing times were ld,3d and 6d after RNA interference. Transfection efficiency in each group was observed. Expression of Cathepsin L of hepatoma cells was detected by immunofluorescence, RT-PCR and WB. Cell vigor was detected by MTT assay. Changes in the cell cycle and apoptosis were observed by flow cytometry. Transwell cabin was used to detect the changes of cell invasive power.Results In the experimental group, Cathepsin L mRNA level and protein level significantly decreased, the proliferation index significantly decreased and apoptosis index significantly increased. The invasive power also decreased. Conclusion RNAi interference can inhibit Cathepsin L expression, cell proliferation and cell invasive power efficiently.
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Objective To investigate the effects of vascular endothelial growth factor receptor-1(VEGFR-1) in the invasion and metastasis of hepatocellular carcinoma(HCC)by detecting the expression of VEGFR-1 in HCC tissues.Methods Paraffin-embedded tissue which containing HCC tissues and adjacent tissues were prepared from patients(n=82)with HCC,then were analyzed by immunohistochemical technique for the expression of VEGFR-1,epithelial marker E-cadherin and mesenchymal marker Vimentin.The relationship between the expression of VEGFR-1 and pathological parameters,and the correlation between VEGFR-1 and E-cadherin,VEGFR-1 and Vimentin were analyzed.Results The expression rates of VEGFR-1 in HCC tissues and adjacent tissues were 89%(73/82)and 0,respectively.The difference of VEGFR-1 expression in portal vein tumor thrombus,tumor capsule,stage,size and differentiation grade of tumor had statistical significance(x2=22.192,15.934,16.751,20.154,6.487,P＜0.05).The expression of VEGFR-1 had effect on the 1-,2-year recurrence rate and 2-year survival rate (x2=0.983,0.958,0.847,P＜0.05),but not on 1-year survival rate(x2=0.359,P＞0.05).The expression of VEGFR-1 was negatively correlated with that of E-cadherin(r=0.765,P＜0.01)but positively with that of Vimentin (r=1.469,P＞0.05).Conclusions VEGFR-1 may play an important role in invasion and metastasis of HCC.Epithelial-mesenehymal transition that induced by VEGFR-1 may contribute to the invasion and metastasis of HCC.
ABSTRACT
Objective To investigate the auxiliary effects of huaier granule on hepatoeellular carcinoma (HCC)patients after liver transplantation.Methods Sixty HCC patients who had undergone liver transplantation from Julv 2004 to June 2006 and met the standard of UCSF were involved in this study.All patients were divided into huaier granule group(n=20),chemotherapy group(n=15),huaier granule+chemotherapy group (n=15)and control group(n=10).The white blood cell count,liver function,cell immunity and immunologieal reiection were detected.The 1-year tumor recurrence rate wag calculated.Results The white blood cell counts in chemotherapy group 1,3,and 6 months after treatment were significantly lower than that before treatment (F=62.053,58.472,49.807,P＜0.05).The changes of white blood cell counts of the other 3 groups before and aftertreatment were small.The difference on the white blood cell counts of the 4 groups had no statistical 8ignincanee(F=102.361,113.412,87.572,P＜0.05).The NK activity,CD4+/CD8+ ratio,IL-2 level in huaier granule group and huaier granule+chemotherapy group 1,3,6 months after treatment were significantly higher than those before treatment,and were significantly higher than those in chemotherapy group and control group(P＜0.05).No immunological rejection occurred in all the groups.Two patients in each group had recurrence and metastasis of HCC within 1 year after the treatment.and the incidence in control group was higher than the other 3 groups(P＜0.05). Conclusions Humer granule can increase the white blood cell count which is decreased after chemotherapy,impmve cellular immunity,and effectively suppress the recurrence and metastasis of HCC at the first year after operation.