Jiong Bi

Involvement of PI3K/PTEN/AKT/mTOR pathway in invasion and metastasis in hepatocellular carcinoma: Association with MMP‐9

Aim:  To investigate the status of Phosphatidylinositol 3‐kinase (PI3K)/PTEN/AKT/mammalian target of rapamycin (mTOR) pathway and its correlation with clinicopathological features and matrix metalloproteinase‐2, ‐9 (MMP‐2, 9) in human hepatocellular carcinoma (HCC).

Bead‐based microarray analysis of microRNA expression in hepatocellular carcinoma: miR‐338 is downregulated

Aim:  Recent studies have underlined causative links between microRNA (miRNA) deregulation and cancer development. However, the relevance of abnormally expressed miRNA to tumor biology has not been well understood in hepatocellular carcinoma (HCC).

Interleukin‐8 modulates growth and invasiveness of estrogen receptor‐negative breast cancer cells

Breast cancer, especially estrogen receptor (ER)‐negative breast cancer, remains hard to treat despite major advances in surgery and adjuvant therapies. The deletion of ER has been consistently associated with tumor progression, recurrence, metastasis and poor prognosis. Among other differences in biological features, ER‐negative breast cancers express high levels of interleukin‐8 (IL‐8), whereas their ER‐positive counterparts do not. IL‐8 is a multi‐functional cytokine with many important biological functions in tumor formation and development. We aimed to study the role(s) of IL‐8 in ER‐negative breast cancer progression by using RNA interference to specifically knockdown IL‐8 expression in ER‐negative breast cancer cell lines MDA‐MB‐231 and MDA‐MB‐468. In vitro, suppression of IL‐8 led to significant reductions in cell invasion (p < 0.001), but had no effects on cell proliferation or cell cycle. In vivo, suppression of IL‐8 significantly reduced the microvessel density (p < 0.05), and markedly reduced neutrophil infiltration into the tumors (p < 0.05). In contrast to in vitro observations, suppression of IL‐8 promoted tumor growth in nude mice (p < 0.05). Our results imply that the complex roles of IL‐8 in the regulation of ER‐negative breast cancer progression may in part be related to its potent chemotactic effects on neutrophils, which in turn mediates many of the biological functions of IL‐8.

miR-338-3p suppresses invasion of liver cancer cell by targeting smoothened†

MicroRNAs are involved in human carcinogenesis and cancer progression. Our previous study has shown that loss of miR‐338‐3p expression is associated with clinical aggressiveness of hepatocellular carcinoma (HCC). However, the exact roles and mechanisms of miR‐338‐3p remain unknown in HCC. To determine whether and how miR‐338‐3p influences liver cancer cell invasion, we studied miR‐338‐3p in the liver cancer cell lines, and we found that miR‐338‐3p is down‐regulated in treated cells. Forced expression of miR‐338‐3p in SK‐HEP‐1 cells suppressed cell migration and invasion, whereas inhibition of miR‐338‐3p in SMMC‐7721 cells induced cell migration and invasion. Furthermore, smoothened (SMO) was identified as a direct target of miR‐338‐3p. Forced expression of miR‐338‐3p down‐regulated SMO and matrix metalloproteinase (MMP)‐9 expression, but inhibition of miR‐338‐3p up‐regulated SMO and MMP9 expression. However, small interfering RNA targeted SMO reversed the effects induced by blockade of miR‐338‐3p. SMO and MMP9 were overexpressed and associated with invasion and metastasis in HCC tissues. These data indicate that miR‐338‐3p suppresses cell invasion by targeting the smoothened gene in liver cancer in vitro and miR‐338‐3p might be a novel potential strategy for liver cancer treatment. Copyright

FAK is involved in invasion and metastasis of hepatocellular carcinoma

Studies have shown that focal adhesion kinase (FAK) is overexpressed in several human tumors and plays an important role in tumor progression. However, the role and underlying mechanisms of FAK in hepatocellular carcinoma (HCC) progression remains to be elucidated. In this study, we examined FAK and phosphorylated FAK Tyr397 expression in a large series of HCCs. We found that both FAK and phosphorylated FAK Tyr397 were overexpressed in HCC samples and HCC cell lines. Increased FAK and phosphorylated FAK Tyr397 expressions were correlated with tumor stage, vascular invasion and intrahepatic metastasis in HCC. Furthermore, HCC cell adhesion, migration and invasion were substantially impaired by siRNA-mediated knockdown of FAK expression, whereas cell growth, apoptosis and cell cycle distribution were not affected. In addition, depletion of FAK induced a significant reduction in expressions and activities of both MMP-2 and MMP-9. Taken together, FAK contributes to invasion and metastasis of HCC partly through regulating expressions and activations of both MMP-2 and MMP-9, suggesting FAK could be a promising therapeutic target for HCC.

Clinical significance of miR-221 and its inverse correlation with p27Kip1 in hepatocellular carcinoma

The aim of the present study is to explore possible role of miR-221 in the pathogenesis of HCC. Matched HCC and adjacent non-cancerous samples were assayed for the expression of miR-221 and three G1/S transition inhibitors: p27Kip1, p21WAF1/Cip1and TGF-β1 by in situ hybridization and immunohistochemistry respectively. p27Kip1 is one of miR-221’s proven targets. Real time qRT-PCR was used to investigate miR-221 and p27Kip1 transcripts in different clinical stages. Western blotting was used to analyze the expression levels of p27Kip1 protein in different clinical stages. In result, miR-221 and TGF-β1 are frequently up-regulated in HCC, while p27Kip1 and p21WAF1/Cip1 proteins are frequently down-regulated. Moreover, miR-221 and p27Kip1’s expression correlated with metastasis and miR-221’s expression also correlated with tumor size. Both of p21WAF1/Cip1and TGF-β1’s expression correlated with tumor differentiations. miR-221’s upregulation and p27Kip1’s downregulation were significantly associated with tumor stages and metastasis. In conclusion, miR-221 is important in tumorigenesis of HCC, possibly by specifically down-regulating p27Kip1, a cell-cycle inhibitor. These results indicate miR-221 as a new therapeutic target in HCC.

Upregulation of PTEN involved in rosiglitazone-induced apoptosis in human hepatocellular carcinoma cells.

AbstractAim:To investigate the effects of rosiglitazone, a peroxisome proliferator-activated receptor gamma (PPARγ) agonist, on the expression of the phosphatase and tensin homologue deleted on chromosome 10 gene (PTEN) and cell growth in hepatocellular carcinoma cells, as well as the underlying mechanisms of these effects.Methods:RT-PCR and Western blotting analyses were performed to detect transcription and the expression of PTEN in Hep3B cells treated with rosiglitazone. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was used to evaluate cell growth. Flow cytometry, DNA fragmentation analysis, caspase enzymatic assay, and Hoechst 33258 staining were used to determine cell apoptosis. Furthermore, small interfering RNA was used to suppress PTEN expression.Results:Rosiglitazone increased the expression of PTEN in a dose-and time-dependent manner through the PPARγ-dependent signal transduction pathway. PTEN upregulation was concomitant with a decreased level of Akt phosphorylation, subsequently resulting in cell growth inhibition and apoptosis in Hep3B cells. PTEN knockdown dramatically blocked these effects of rosiglitazone. Moreover, the exposure of cells to rosiglitazone activated caspases-9 and -3 during apoptotic proceeding.Conclusion:Thus, upregulation of PTEN is involved in the inhibition of cell growth and the induction of cell apoptosis by rosiglitazone, suggesting that rosiglitazone may be useful in liver cancer therapy via apoptosis.

Overexpression of YKL-40 is an independent prognostic marker in gastric cancer ☆

YKL-40 is a growth factor for connective tissue cells and a migration factor for endothelial cells. Elevated serum level of YKL-40 has been associated with poor prognosis in many cancers. However, the status of YKL-40 expression and its clinical/prognostic significance in gastric cancer are unclear. In this study, the expression of YKL-40 was studied by immunohistochemistry in gastric cancer tissue microarray containing 172 primary gastric cancer cases and 70 adjacent nonneoplastic mucosa specimens. The correlations between YKL-40 expression and clinicopathologic features, as well as activation of PI3K/Akt pathways were addressed. Expression of YKL-40 was significantly higher in gastric cancer tissues than that in adjacent nonneoplastic tissues. Overexpression YKL-40 was found in 28.4% of gastric cancers and was significantly associated with tumor invasion (P = .007) and lymph node metastasis (P = .009). For survival study, overexpression of YKL-40 was significantly associated with worse outcome (P = .001). When known clinical variables were added to a multivariate analysis, TNM stage, tumor size, and overexpression of YKL-40 emerged as independent prognostic factors. Further study indicated that the oncogenic function of YKL-40 might be through the activation of Akt pathway. These results suggest that overexpression of YKL-40 is correlated with the aggressive behavior of tumor cells, which could be used as an independent molecular marker for the predicting poor prognosis of patients with gastric cancer.

Characterization of a candidate tumor suppressor gene uroplakin 1A in esophageal squamous cell carcinoma

Esophageal squamous cell carcinoma (ESCC) is increasing in incidence, but the knowledge of the genetic underpinnings of this disease remains limited. In this study, we identified the tetraspanin cell surface receptor uroplakin 1A (UPK1A) as a candidate tumor suppressor gene (TSG), and we investigated its function and mechanism in ESCC cells. UPK1A downregulation occurred in 68% of primary ESCCs examined, where it was correlated significantly with promoter hypermethylation (P < 0.05). Ectopic expression of UPK1A in ESCC cells inhibited cell proliferation, clonogenicity, cell motility, and tumor formation in nude mice. Mechanistic investigations suggested that these effects may be mediated by inhibiting nuclear translocation of β-catenin and inactivation of its downstream targets, including cyclin-D1, c-jun, c-myc, and matrix metalloproteinase 7 (MMP7). Cell cycle arrest elicited by UPK1A at the G(1)-S checkpoint was associated with downregulation of cyclin D1 and cyclin-dependent kinase 4, whereas metastasis suppression was associated with reduction of MMP7. These findings were consistent with evidence derived from clinical samples, where UPK1A downregulation was correlated with lymph node metastasis (P = 0.009), stage (P = 0.015), and overall survival (P < 0.0001). Indeed, multivariate cyclooxygenase regression analysis showed that UPK1A was an independent prognostic factor for overall survival. Taken together, our findings define a function for UPK1A as an important TSG in ESCC development.

MicroRNA-129-5p inhibits hepatocellular carcinoma cell metastasis and invasion via targeting ETS1

MiR-129-5p is deregulated in various human cancers and has been associated with hepatocellular carcinoma (HCC) progression. However, the underlying mechanisms of miR-129-5p involvement in the development and progression of HCC and the effects of miR-129-5p deregulation on the clinical characteristics observed in HCC patients remain poorly understood. We therefore investigated the correlation between low miR-129-5p expression and vascular invasion, intrahepatic metastasis, and poor patient survival. Ectopic restoration of miR-129-5p expression in HCC cells suppressed cellular migration and invasion and the expression of v-ets erythroblastosis virus E26 oncogene homolog 1 (ETS1), while inhibition of endogenous miR-129-5p caused an increase in these parameters. We identified the ETS1 gene as a novel direct target of miR-129-5p. SiRNA-mediated ETS1 knockdown rescued the effects of anti-miR-129-5p inhibitor in HCC cell lines, while the effects of miR-129-5p overexpression were partially phenocopied in the knockdown model. In addition, miR-129-5p levels inversely correlated with those of ETS1 in HCC cells and tissues. Taken together, our findings indicate an important role for miR-129-5p in the molecular etiology of invasive HCC and suggest that miR-129-5p could have potential therapeutic applications in HCC.