Epoxidized Soybean-Oils-Based Pressure-Sensitive Adhesives with Di-Hydroxylated Soybean-Oils Copolymerizing and Antioxidant Grafting.

Vegetable-oils-based pressure-sensitive adhesives (PSAs) are being developed as a substitute for petrochemical-based PSAs for application in daily life. However, vegetable-oils-based PSAs face the problems of unsatisfactory binding strengths and easy aging. In this work, the grafting of antioxidants (tea polyphenol palmitates, caffeic acid, ferulic acid, gallic acid, butylated hydroxytoluene, tertiary butylhydroquinone, butylated hydroxyanisole, propyl gallate (PG), tea polyphenols) was introduced into an epoxidized soybean oils (ESO)/di-hydroxylated soybean oils (DSO)-based PSA system to improve the binding strengths and aging-resistant properties. PG was screened out as the most suitable antioxidant in the ESO/DSO-based PSA system. Under optimal conditions (ESO/DSO mass ratio of 9/3, 0.8% PG, 55% rosin ester (RE), 8% phosphoric acid (PA), 50 °C, and 5 min), the peel adhesion, tack, and shear adhesion of the PG-grafted ESO/DSO-based PSA increased to 1.718 N/cm, 4.62 N, and >99 h, respectively, in comparison with the control (0.879 N/cm, 3.59 N, and 13.88 h), while peel adhesion residue reduced to 12.16% in comparison with the control (484.07%). The thermal stability of the ESO/DSO-based PSA was enhanced after PG grafting. PG, RE, PA, and DSO were partially crosslinked in the PSA system, with the rest being free in the network structures. Thus, antioxidant grafting is a feasible method for improving the binding strengths and aging-resistant properties of vegetable-oils-based PSAs.

In Situ Hypoiodite-Catalyzed Oxidative Rearrangement of Chalcones: Scope and Mechanistic Investigation.

It is highly desirable to avoid using rare or toxic metals for oxidative reactions in the synthesis of pharmaceuticals and fine chemicals. Hypervalent iodine compounds are environmentally benign alternatives, but their catalytic use has been quite limited. Herein, the protocol for in situ hypoiodite-catalyzed oxidative rearrangement of chalcones is first realized under mild and metal-free conditions, which provided a nontoxic, environmental-benign, and catalytic alternative to the thallium-based protocol. Also, the applicability and effectiveness of this catalytic protocol got well demonstrated via gram-scale synthesis and product derivatization. What is more, control and NMR tracking experiments were performed to figure out the possible catalytic species and intermediates.

CD47 blockade alleviates acute rejection of allogeneic mouse liver transplantation by reducing ischemia/reperfusion injury.

Despite advances in immunosuppressive therapies, acute rejection response is still a serious concern especially in the early phase after liver transplantation. This study aimed to evaluate whether blocking the TSP1-CD47 signaling pathway could attenuate the acute rejection after liver transplantation. An allogeneic mouse orthotopic liver transplantation model (Balb/c→C3H) with prolonged cold ischemic phase was used to induce severe IRI and lethal acute rejection. CD47mAb or isotype matched-control IgG2a was administered to donor liver during graft perfusion. Recipients were sacrificed at 1d, 3d, 5d and 7d after reperfusion. Blood samples were collected to evaluate serum alanine aminotransferase, total bilirubin, HMGB-1,TNF-α, IL-2 and INF-γ level. Flow cytometric analysis was used to detect the strength of innate and adaptive immune response. Liver tissue was obtained for HE, TUNEL staining and F4/80 immumohistochemical staining. Moreover, we conducted a mixed lymphocyte reaction treated with IgG2a or CD47mAb. Mice in CD47mAb-treated group demonstrated improved survival and significantly lower increase in Suzuki score, apoptosis index, acute rejection index, serum alanine aminotransferase, total bilirubin, HMGB-1, TNF-α, IL-2, INF-γ level and the degree of Kupffer cells' activation especially in the early phase of acute rejection. In addition, Pearson's correlation analysis confirmed significant correlation between Suzuki score/ALT and acute rejection index. The in vitro inhibition assay showed that CD47 blockade couldn't directly inhibit recipient lymphocyte proliferation. Based on the evidence that TSP1-CD47 signaling blockade with CD47mAb could alleviate acute rejection by reducing the extent of IRI after liver transplantation indirectly, this study provided a basis for new interventions and management methods to support better transplant outcomes.

miR-302a inhibits human HepG2 and SMMC-7721 cells proliferation and promotes apoptosis by targeting MAP3K2 and PBX3.

Hepatocellular carcinoma (HCC) is the most common liver cancer and has a poor prognosis. miR-302a is an important regulator of tumor occurrence and deterioration, while MAP3K2 and PBX3 genes are involved in cancer cell proliferation and apoptosis. In this study, the expression of miR-302a and MAP3K2/PBX3 were evaluated by qPCR in liver cancer cell lines. Next, the target relationship between miR-302a and MAP3K2/PBX3 was verified using luciferase assays. Meanwhile, the expression correlation between miR-302a and target genes was analyzed in cancer tissue and para-cancerous tissue. In addition, an increased miR-302a level in HepG2 cells and SMMC-7721 cells were achieved through transfection with miR-302a mimics, and the effects on HepG2 cell and SMMC-7721 cell proliferation, apoptosis and MAPK pathways were determined using MTT, flow cytometry, qPCR and western blot assays. The results showed that liver cancer cell lines exhibited low miR-302a expression and MAP3K2 and PBX3 were confirmed to be the target genes of miR-302a. Meanwhile, the HE results showed that cells became enlarged with loose cytoplasm and formed balloon-like lesions in HCC specimens and we found a significant negative correlation between miR-302a and MAP3K2/PBX3 expression. In addition, treatment with miR-302a mimics inhibited HepG2 cells and SMMC-7721 cells proliferation and increased the apoptosis rate. Further research revealed that the MAPK key factors p-p38, p-ERK1/2 and p-JNK were significantly reduced in miR-302a transfected cells and MAP3K2/PBX3 silenced cells. Besides, MAP3K2 and PBX3 overexpression in miR-302a mimics-treated cells exerted the opposite effects. In conclusion, miR-302a inhibited proliferation and promoted apoptosis in human hepatoma cells by targeting MAP3K2 and PBX3.

Role of microRNA-141-3p in the progression and metastasis of hepatocellular carcinoma cell.

Hepatocellular carcinoma (HCC) is a leading cause of cancer related death worldwide. However, the mechanisms underlying HCC progression and metastasis are still in obscure. Here, we used bioinformatic analysis to identify miRNAs that regulate GP73, a specific marker for HCC diagnosis and prognosis. The correlations between miR-141-3p and clinic-pathological factors were analyzed in HCC patient samples; proliferation, migration, invasion, and colony formation were studied using established HCC cell lines. Expression levels of target genes (miR-141-3p, GP73, E-cadherin, N-cadherin, occludin, vimentin, and cytokeratin 18) were detected by either Western blot or qRT-PCR analysis. Xenograft models were established to evaluate tumor growth and metastasis. MiR-141-3p was significantly reduced in HCC tumors and cell lines, highly correlated with tumor progression. In contrast, GP73 was negatively correlated with miR-141-3p in HCC tumors. MiR-141-3p overexpression significantly decreased HCC cell proliferation, migration, and invasion by inhibiting epithelial-mesenchymal transition (EMT). GP73 overexpression partially restored the inhibitory effects of miR-141-3p, while miR-141-3p overexpression markedly inhibited tumor growth and pulmonary metastasis, which were partially reversed by GP73 overexpression. Our findings suggest that miR-141-3p targets GP73 to reverse EMT, subsequently inhibiting HCC progression and metastasis. Thus, overexpression of miR-141-3p could serve as a therapeutic strategy to arrest HCC.

Lentivirus-mediated knockdown of P27RF-Rho inhibits hepatocellular carcinoma cell growth.

AIM OF THE STUDY: To investigate the effects of P27RF-Rho on hepatocellular carcinoma (HCC) cell growth and explore the possibility of using it as a novel therapeutic target for liver cancer treatment. MATERIAL AND METHODS: P27RF-Rho in HCC cells was silenced by lentivirus-mediated RNA interference, and the silencing effect was verified by RT-PCR. Cell proliferation was determined by MTT and clone formation assay. Cell cycle phase and apoptosis were detected through FACS. The expression level of cell growth, apoptosis, and metastasis associated genes was detected by quantitative PCR. RESULTS: Lentivirus-mediated P27RF-Rho knockdown inhibited HCC cell growth and clone formation. P27RF-Rho silence induced cell cycle arrest and apoptosis. The mRNA level of genes associated with cell cycle, apoptosis, and invasion also significantly altered after P27RF-Rho knockdown. Cyclin A, CDK2, BCL-2, and MMP-9 were down-regulated. P27 and Bax were up-regulated. CONCLUSIONS: P27RF-Rho knockdown inhibits HCC cell growth, and P27RF-Rho is probably a promising target for HCC treatment.

Overexpression of Aquaporin 1 on cysts of patients with polycystic liver disease.

BACKGROUND AND OBJECTIVE: Polycystic liver disease (PCLD) represents a group of genetic disorders that include autosomal dominant polycystic kidney disease (ADPKD) and isolated polycystic liver disease (iPCLD). There is currently no definitive treatment except for liver transplantation. The aim of this study was to assess the expression level of aquaporin 1 (AQP1) on the PCLD cysts with different sizes and provide the potential therapeutic target. METHODS: We collected 3 normal bile ducts, and recruited 8 patients with simple liver cyst disease, 24 patients with ADPKD, and 17 patients with iPCLD. AQP1 expression in different types of cyst walls and in normal bile ducts was detected using real time quantitative PCR, western blot and immunofluorescence staining. We also compared AQP1 expression levels in cysts of different sizes. Besides, ionic concentrations, pH and osmolality of cyst fluid were analyzed. RESULTS: The results showed that AQP1 expression in PCLD cysts was significantly higher than that in simple liver cysts and the normal bile ducts. In addition, a comparable increasing trend was found in cysts of smaller sizes to cysts of larger sizes. pH values, the sodium and chloride concentrations were higher in cyst fluid than that in the serum. CONCLUSIONS: AQP1 was overexpressed in cystic cholangiocytes. A tendency of increased AQP1 protein expression in correlation with the cyst size was also found. These observations offered a direction into the molecular mechanisms of cyst expansion and maybe provide new treatment strategies to reduce fluid secretion into liver cysts.

Overexpression of Aquaporin 1 on cysts of patients with polycystic liver disease

Background and objective: Polycystic liver disease (PCLD) represents a group of genetic disorders that include autosomal dominant polycystic kidney disease (ADPKD) and isolated polycystic liver disease (iPCLD). There is currently no definitive treatment except for liver transplantation. The aim of this study was to assess the expression level of aquaporin 1 (AQP1) on the PCLD cysts with different sizes and provide the potential therapeutic target. Methods: We collected 3 normal bile ducts, and recruited 8 patients with simple liver cyst disease, 24 patients with ADPKD, and 17 patients with iPCLD. AQP1 expression in different types of cyst walls and in normal bile ducts was detected using real time quantitative PCR, western blot and immunofluorescence staining. We also compared AQP1 expression levels in cysts of different sizes. Besides, ionic concentrations, pH and osmolality of cyst fluid were analyzed. Results: The results showed that AQP1 expression in PCLD cysts was significantly higher than that in simple liver cysts and the normal bile ducts. In addition, a comparable increasing trend was found in cysts of smaller sizes to cysts of larger sizes. pH values, the sodium and chloride concentrations were higher in cyst fluid than that in the serum. Conclusions: AQP1 was overexpressed in cystic cholangiocytes. A tendency of increased AQP1 protein expression in correlation with the cyst size was also found. These observations offered a direction into the molecular mechanisms of cyst expansion and maybe provide new treatment strategies to reduce fluid secretion into liver cysts (AU)

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Effects of Ras homolog gene family, member C gene silencing combined with rapamycin on hepatocellular carcinoma cell growth.

The aim of the present study was to investigate the combined effects of inhibiting the Ras homolog gene family, member C (RhoC)/Rho kinase and phosphoinositide 3 kinase/Akt/mammalian target of rapamycin (mTOR) pathways on hepatocellular carcinoma cell growth. The RhoC gene was silenced by RNA interference (RNAi) and mTOR was inhibited by rapamycin (RAPA). Subsequently, an MTT assay for cell growth detection, western blot analysis for gene expression analysis, silver nitrate staining for cell proliferation, Wright's staining for analysis of the apoptotic rate analysis, soft agar clonogenic assay for the determination of cell growth characteristics and a Transwell assay for cell migration were performed. RhoC expression in hepatoma cell lines was lower than that in the HL7702 normal human liver cell line. The level of cell proliferation in the RNAi + RAPA group was lower than that in the RNAi, RAPA and Scramble groups. The levels of cyclin­dependent kinase 2 in the RNAi + RAPA group were lower than those in the other groups, while the levels of P16 in the RNAi + RAPA group were higher than those in the other experimental groups. No significant difference was found between the RNAi + RAPA and the normal HL7702 group. The number of silver nitrate­stained particles was reduced in the RNAi + RAPA group compared with that in the other groups. No significant difference was found between the RNAi + RAPA and HL7702 groups. Wright's staining for apoptosis demonstrated that apoptosis in the Scramble group was rare, while the RAPA and RNAi groups contained a large number of apoptotic cells, which displayed nuclear condensation, fragmentation, deepened staining, as well as a wrinkled membrane. B­cell lymphoma­2 (Bcl­2) expression in the RNAi + RAPA group was lower than that in the other groups, while the gene expression of Bcl­2­associated X protein in the RNAi + RAPA group was increased compared with that in the other groups. No cell colony formation was observed in the soft agar cloning experiment in the RNAi + RAPA and HL7702 group, while in the other groups, visible cell clones appeared. In the Transwell assay the number of migrated cells in the RNAi + RAPA group was lower than that in the other groups. The gene expression of matrix metalloproteinase (MMP)2, MMP­9 and vascular endothelial growth factor in the RNAi + RAPA group was lower than that in the other experimental groups. In conclusion, RhoC gene silencing combined with RAPA was able to significantly inhibit the growth of hepatocellular carcinoma cells.

In vitro regulation of hepatocellular carcinoma cell viability, apoptosis, invasion, and AEG-1 expression by LY294002.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common malignancies in the world, and is characterized by advanced clinical stages at diagnosis and very poor prognosis. SUBJECTS AND METHODS: This study investigated the effects of PI3K inhibitor, LY294002, on suppression of astrocyte elevated gene-1 (AEG-1) and regulation of HCC cell viability, apoptosis, and invasion in vitro. Cell lines derived from normal liver and HCC were treated with LY294002 and evaluated by RT-PCR, western blot, cell viability, migration, and invasion assays. RESULTS: The data showed that AEG-1 mRNA and protein were overexpressed in HCC cells, compared to the normal liver cells. LY294002 treatment of HCC cells significantly reduced tumor cell viability, but promoted apoptosis. Tumor cell migration and invasion assays showed that LY294002 treatment also decreased the capacity of HCC cell migration and invasion. Molecularly, LY294002 treatment down-regulated AEG-1 expression, AKT and GSK3ß phosphorylation, and expression of cyclinD1, CDK4, VEGF and Bcl2, but up-regulated Bax and c-Myc expression. CONCLUSION: The data from this study demonstrated usefulness of LY294002 for effective control of HCC. Future studies should investigate the effects of LY294002 on HCC cells in vivo before initiating clinical trials in HCC patients.

STAT1 negatively regulates hepatocellular carcinoma cell proliferation.

Signal transducer and activator of transcription 1 (STAT1) regulates cell proliferation and survival. The present study aimed to investigate the role of STAT1 in the development and progression of human hepatocellular carcinoma (HCC). The levels of STAT1 expression in 36 HCC and 12 non-HCC liver tissues were examined by immunohistochemistry. The effect of STAT1 overexpression or silencing on the proliferation and apoptosis of HCC cells was determined by MTT and flow cytometric assays. The effect of STAT1 overexpression or silencing on the levels of p53 and cyclin E expression was determined by quantitative PCR and western blot assays. The level of STAT1 expression in the HCC tissues was significantly lower compared to the level in the non-HCC liver tissues and was negatively associated with the histological grade of HCC and serum HBsAg, anti-HCV and α-fetoprotein positivity in HCC patients. Induction of STAT1 overexpression significantly inhibited HepG2 cell proliferation and enhanced HCC cell apoptosis, accompanied by upregulation of p53 expression and STAT1 phosphorylation, but a reduction in cyclin E expression in HepG2 cells. In contrast, knockdown of STAT1 by introduction of STAT1-specific siRNA promoted HepG2 cell proliferation, but inhibited HCC cell apoptosis, accompanied by significant downregulation of p53 expression, but enhancement of cyclin E expression in vitro. Our data suggest that STAT1 may inhibit HCC growth by regulating p53-related cell cycling and apoptosis.

Overexpression of Ras homologous C (RhoC) induces malignant transformation of hepatocytes in vitro and in nude mouse xenografts.

Ras homologous C (RhoC) is expressed in various cancers, including hepatocellular carcinoma (HCC). In this study, we first analyzed RhoC expression in 46 HCC tissue specimens and found that RhoC expression was significantly increased in HCC tissues compared to the adjacent normal liver tissues. Next, we investigated the role of RhoC in malignant transformation of normal hepatocytes. The HL7702 cell line was stably transfected with a RhoC expression vector and then subjected to cell proliferation, differentiation, colony formation, migration and invasion assays, as well as nude mouse xenograft assays. Gene expressions in these cells were determined using RT-PCR and Western blot. Overexpression of RhoC significantly promoted proliferation and anchorage-independent growth of HL7702 cells, but suppressed cell differentiation, as compared with the parental cells and the empty vector-transfected control cells. Moreover, RhoC overexpression induced migration and invasion of HL7702 cells in vitro. Molecularly, RhoC increased the expression of cell cycle-related genes, matrix metalloprotease 2 (MMP2), MMP9 and vascular endothelial growth factor (VEGF). In addition, RhoC-transfected cells formed tumors in nude mice, whereas vector-transfected HL7702 cells did not form any tumors in nude mice. This study demonstrated the role of RhoC overexpression in malignant transformation of normal human hepatocytes, suggesting that RhoC may function as an oncogene in hepatocytes.

miRNA-22 suppresses colon cancer cell migration and invasion by inhibiting the expression of T-cell lymphoma invasion and metastasis 1 and matrix metalloproteinases 2 and 9.

Emerging evidence has demonstrated the altered expression of mRNAs in cancer development and progression. In this study, the precise role of miRNA-22 (miR-22) in colon cancer cells was investigated. Upon transfection with a miR-22 expression vector, the viability of HCT-116 human colon cancer cells was significantly reduced and tumor cell migration and invasion capacity were also suppressed. Computational in silico analysis predicted that T-cell lymphoma invasion and metastasis 1 (TIAM1) is a target gene of miR-22. This was confirmed by qRT-PCR and western blotting, which showed that miR-22 expression inhibited TIAM1 mRNA and protein expression, respectively. In addition, the expression of pro-invasive gene matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9) and pro-angiogenic protein vascular endothelial growth factor (VEGF) were also reduced by miR-22 expression. Collectively, these data suggest that miR-22 may act as a tumor suppressor in colon cancer, most likely by targeting TIAM1 expression.

Recombinant VAA-I from Viscum album induces apoptotic cell death of hepatocellular carcinoma SMMC7721 cells.

Researchers have proposed that VAA-I, a specific plant lectin found in Viscum album, has therapeutic effects on cancer and autoimmune diseases. VAA-I has shown some promising treatment results in some types of tumor cell lines, especially SMMC-7721 cells (human hepatocellular carcinoma cells). However, few details are known about the mechanism and process of cell death induced by VAA-I in tumor cells. In this study, the cell morphology results showed that SMMC-7721 cells treated with VAA-I exhibited several features typical of apoptotic cell death, which was confirmed by the Caspase inhibition assay. Fluo-3-acetoxymethyl ester (AM) fluorescence imaging techniques showed that rVAA-I significantly elevated the intracellular calcium level ([Ca2+]i) in SMMC-7721 cells. These findings suggest that apoptosis may play the most important role in SMMC-7721 cell death induced by rVAA-I. Finally, in the SMMC-7721 cells treated with rVAA-I, a series of genes in the p38 mitogen-activated protein kinase (MAPK) signaling pathway were expressed differentially, and further found that PI 3-kinase pathway is involved in rVAA-I signal transduction in SMMC-7721 cells.

Effects of STAT3 gene silencing and rapamycin on apoptosis in hepatocarcinoma cells.

The PI3K/Akt/mTOR and JAK/STAT3 signaling pathways are important for regulating apoptosis, and are frequently activated in cancers. In this study, we targeted STAT3 and mTOR in human hepatocellular carcinoma Bel-7402 cells and examined the subsequent alterations in cellular apoptosis. The expression of STAT3 was silenced with small interfering RNA (siRNA)-expressing plasmid. The activity of mTOR was inhibited using rapamycin. Following treatment, Annexin V/propidium iodide staining followed by flow cytometry and Hoechst33258 immunofluorescence staining was used to examine cellular apoptosis. JC-1 staining was used to monitor depolarization of mitochondrial membrane (ΔΨm). Furthermore, the expression of activated caspase 3 protein was analyzed by Western blotting. Compared to non-treated or control siRNA-transfected cells, significantly higher levels of apoptosis were detected in siSTAT3-transfected or rapamycin-treated cells (P < 0.05), which was further enhanced in cells targeted for both molecules (P < 0.05). The pro-apoptotic effects were accompanied with concomitant depolarization of mitochondrial membrane and up-regulation of activated caspase 3. Combined treatments using rapamycin and STAT3 gene silencing significantly increases apoptosis in Bel-7402 cells, displaying more dramatic effect than any single treatment. This study provides evidence for targeting multiple molecules in cancer therapy.

The role of RhoC in the proliferation and apoptosis of hepatocellular carcinoma cells.

In this study, we examined the effects of RhoC expression on the growth and apoptosis of human hepatocellular carcinoma cells (HCCs) in vitro in order to gain more understanding of its potential as a therapeutic target gene. We knocked down the endogenous expression levels of RhoC in human HCCs, BEL-7402, using siRNA and ectopically expressed RhoC in untransformed hepatocytes, HL7702. Stable cell lines were established, and cell growth was examined by MTT and colony formation assays, cell proliferation examined by silver nitrate staining of AgNORs, and cell cycle distribution examined by flow cytometry. RT-PCR analysis was performed to determine the mRNA expression levels of RhoC and cell cycle-related genes. Finally, the effect of RhoC expression on apoptosis was also examined by flow cytometry, agarose gel electrophoresis of fragmented DNA, Wright staining, and RT-PCR analysis for genes regulating apoptosis. Compared to the parental and control siRNA (siCtrl)-transfected BEL-7402 cells, the siRhoC-transfected cells exhibited significantly reduced cell growth, cell proliferation, percentage of cells in the S-G2/M phase, and expression of Cyclin D1, CDK4, and Bcl2. Knockdown of RhoC expression in BEL-7402 cells also significantly increased the percentage of cells in the G0/G1 phase, cellular apoptosis, and expression of p21, p16, and Bax. Furthermore, ectopic expression of RhoC in HL7702 cells led to a significant increase in cell growth compared to parental or siCtrl-transfected cells. These data suggest that RhoC is a key regulator of cell growth and apoptosis in HCC cells, making it a potential target for gene therapy.

[Effect of RhoC on hepatocellular carcinoma cell growth and related molecular mechanisms].

OBJECTIVE: To clarify the role of RhoC in the growth of hepatocellular carcinoma cells and its molecular mechanism, so as to explore the molecular target of tumor cell growth. METHODS: siRNA-RhoC plasmid was constructed and RhoC gene silencing the cell-line of hepatocellular carcinoma was setup. Cell growth was assessed by MTT assay. AgNORs staining was applied to determine cell proliferation. Plate cell clone test was conducted to examine the capacity of cell clone formation. FACS was adopted to measure the course of cell cycle and semi-quantitative RT-PCR was used to determine the expression of cell cycle proteins. In order to further determine the effect of RhoC expression on cell growth, a RhoC over-expression human hepatocellular cell line was setup by PcDNA3-RhoC plasmid transfection. RESULTS: The inhibition rate of RhoC was 82.3%. From the fourth day of cell culture, the growth of cells in RNAi group was significantly slower than that in parental Bel7402 and negative control groups (0.41 ± 0.10 vs. 0.73 ± 0.11 and 0.71 ± 0.07 respectively, P < 0.05). AgNORs staining showed that average cell stained particles in RNAi group was significantly lower than that in parental Bel7402 and negative control(1.23 ± 0.35 vs. 3.47 ± 0.93 and 3.17 ± 0.78, P < 0.01). Plate clone formation test showed that clone formation efficiency in the RNAi group was notably lower than that in the control group [(20.33 ± 5.42)% vs. (70.58 ± 10.10)% and (69.83 ± 14.77)%, respectively, P < 0.01]. Cell cycle analysis by FACS showed that G(0)/G(1) cell percentage in the RNAi group was significantly higher than that in the control group [(73.14 ± 5.93)% vs. (57.05 ± 5.97)% and (52.99 ± 4.80)%, P < 0.05]. Compared with Bel7402 and negative control groups, the expression of following growth associated genes was significantly decreased: cyclin D1(0.45 ± 0.21 vs. 1.25 ± 0.24 and 1.12 ± 0.15, respectively, P < 0.05)and CDK4 (0.55 ± 0.08 vs. 1.18 ± 0.32 and 1.10 ± 0.29, respectively, P < 0.05); the following genes were notably increased: p16(1.07 ± 0.23 vs. 0.36 ± 0.12 and 0.35 ± 0.13, respectively, P < 0.01)and p21(0.42 ± 0.12 vs. 0.17 ± 0.06 and 0.19 ± 0.08, respectively, P < 0.05). RhoC was highly expressed in PcDNA3-RhoC transfected hepatocellular cell line. From the third day on of the cell culture, cell growth in PcDNA3-RhoC group was remarkably higher than that in the HL7702 and PcDNA3 groups (0.83 ± 0.10 vs. 0.54 ± 0.11 and 0.58 ± 0.55, respectively, P < 0.05). CONCLUSIONS: RhoC is the key molecule in promoting hepatocellular cell growth, and is a promising target for tumor cell growth controlling.