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Increasing studies have confirmed that abnormally expressed microRNAs (miRNAs) take part in the carcinogenesis as well as the aggravation of hepatocellular carcinoma (HCC). However, little information is currently available about miR‐1914 in HCC. Here, we first confirmed that miR‐1914 inhibition in HCC cell lines and tumour specimens correlates with tumour size and histological grade. In a series of functional experiments, miR‐1914 inhibited tumour proliferation and colony formation, resulting in cell cycle arrest and increased apoptosis. Moreover, miR‐1914 mediated its functional effects by directly targeting GPR39 in HCC cells, leading to PI3K/AKT/mTOR repression. Restoring GPR39 expression incompletely counteracted the physiological roles of miR‐1914 in HCC cells. In addition, down‐regulation of AKT phosphorylation inhibited the effects of miR‐1914 in HCC. Furthermore, the overexpression of lncRNA DUXAP10 negatively correlated with the expression of miR‐1914 in HCC; thus, lncRNA DUXAP10 regulated miR‐1914 expression and modulated the GPR39/PI3K/AKT‐mediated cellular behaviours. In summary, the present study demonstrated for the first time that lncRNA DUXAP10–regulated miR‐1914 plays a functional role in inhibiting HCC progression by targeting GPR39‐mediated PI3K/AKT/mTOR pathway, and this miRNA represents a novel therapeutic target for patients with HCC.

Local and systemic metastasis of hepatocellular carcinoma (HCC) causes the poor prognosis and increasing evidence confirms that aberrant miRNAs were involved in cancer progression. However, the expression and mechanisms of a specific miR-3194-3p in HCC remains unknown. In this research, we demonstrated that miR-3194-3p, significantly down-regulated in HCC tissues and cell lines, was associated with metastasis and recurrence of HCC. Notably, gain- and loss-of-function assays demonstrated that miR-3194-3p inhibited the migration, invasion and epithelial-mesenchymal transition (EMT) of HCC cells

The construction of roadways in permafrost regions modifies ground-surface conditions and consequently, negatively varies thermal stability of the underlying frozen soils. To avoid the thawing of the permafrost layer under the scenario of global warming, roadways are usually laid on a built-up embankment, which not only disperses the traffic loads to underlying layers but also minimize the thermal disturbance. In the embankment, duct ventilation, or called air duct, can be embedded to further cool the underlying permafrost. While the thermal performance of duct ventilations has been well documented, the long-term structural stability of duct ventilation remains unknown. This study examines the structural stress of ventilation ducts that are placed in harsh weather such as the Qinghai-Tibet Plateau. The ducts are currently buried in the embankment filler, with the wind-outlet and -inlet ends exposed and cantilevered out of the embankment. Field studies found that the exposed parts have plagued cracking and even failures, especially at the fixed end of the cantilevered part. Damages of these concrete ducts are attributed to cyclic freezing-thawing attack, thermally-induced stresses, moisture-induced stresses, and concrete swelling. These physical attacks are caused by the harsh weather in the Qinghai-Tibet plateau. It is recommended to insulate the exposed part of the ducts and to fabricate durable and dense concrete ducts.

Emerging evidence has indicated that abnormal microRNAs (miRNAs) participated in carcinogenesis and tumor progression in hepatocellular carcinoma (HCC). Better understanding the association between miRNAs and HCC may contribute to discover novel therapeutic approaches for diagnosis and treatments. In the current study, we have shown that miR-1204 level was elevated in HCC tissues and cell lines, which was associated with malignant clinical features, including large tumor size and advanced TNM stage. Furthermore, gain-or loss-of function assays demonstrated that miR-1204 promoted cell proliferation in vitro and tumor growth in vivo as well as inhibited apoptosis in vitro. Luciferase reporter gene assays confirmed that ZNF418 was a direct downstream target of miR-1204. Recuse assays showed that ZNF418 mediates the biological function of miR-1204 on HCC cells through regulating MAPK and c-Jun signaling. In conclusion, our results suggest that miR-1204 functions as an oncogene to promote proliferation and inhibit apoptosis through regulating MAPK and c-Jun signaling by targeting ZNF418, and potentially serves as a novel prognostic biomarker and therapeutic target for HCC.

Nuclear translocation of mothers against decapentaplegic homolog 2/3 (SMAD2/3), core transcription factors of transforming growth factor β (TGF-β) signaling, is critical for hepatic stellate cell (HSC) differentiation into metastasis-promoting myofibroblasts. SMAD2/3 have multiple coactivators, including WW domain-containing transcription regulator protein 1 (WWTR1 or TAZ) and p300 acetyltransferase. In the nucleus, TAZ binds to SMAD2/3 to prevent SMAD2/3 nuclear export. However, how TAZ and SMAD2/3 enter the nucleus remains poorly understood because neither contains a nuclear localization signal (NLS), an amino acid sequence tagging proteins for nuclear transport. p300 is an NLS-containing large scaffold protein, so we hypothesized that SMAD2/3 and TAZ may undergo nuclear import through complexing with p300. Coimmunoprecipitation, immunofluorescence, and nuclear fractionation assays revealed that TGF-β1 promoted binding of SMAD2/3 and TAZ to p300 and that p300 inactivation disrupted TGF-β1-mediated SMAD2/3 and TAZ nuclear accumulation. Deleting the p300 NLS blocked TGF-β1-induced SMAD2/3 and TAZ nuclear transport. Consistently, p300 inactivation suppressed TGF-β1-mediated HSC activation and transcription of genes encoding tumor-promoting factors, such as connective tissue growth factor, Tenascin C, Periostin, platelet-derived growth factor C, and fibroblast growth factor 2, as revealed by microarray analysis. Chromatin immunoprecipitation-real-time quantitative PCR showed that canonical p300-mediated acetylation of histones also facilitated transcription in response to TGF-β1 stimulation. Interestingly, although both TGF-β1-mediated and stiffness-mediated HSC activation require p300, comparison of gene expression data sets revealed that transcriptional targets of TGF-β1 were distinct from those of stiffness-p300 mechanosignaling. Lastly, in tumor/HSC coinjection and intrasplenic tumor injection models, targeting p300 of activated-HSC/myofibroblasts by C646, short hairpin RNA, or cre-mediated gene disruption reduced tumor and liver metastatic growth in mice. Conclusion: p300 facilitates TGF-β1-stimulated HSC activation by both noncanonical (cytoplasm-to-nucleus shuttle for SMAD2/3 and TAZ) and canonical (histone acetylation) mechanisms. p300 is an attractive target for inhibiting HSC activation and the prometastatic liver microenvironment.

Background Accumulating evidence has highlighted the potential role of long non-coding RNAs (lncRNAs) in the biological behaviors of hepatocellular carcinoma (HCC). Here, we elucidated the function and possible molecular mechanisms of the effect of lncRNA-AGAP2-AS1 on the biological behaviors of HCC. Methods EdU, Transwell and flow cytometry were used to determine proliferation, migration, invasion and apoptosis of HCC cells in vitro. The subcutaneous tumor model and lung metastasis mouse model in nude mice was established to detect tumor growth and metastasis of HCC in vivo. The direct binding of miR-16-5p to 3’UTR of ANXA11 was confirmed by luciferase reporter assay. The expression of AGAP2-AS1 and miR-16-5p in HCC specimens and cell lines were detected by real-time PCR. The correlation among AGAP2-AS1 and miR-16-5p were disclosed by a dual-luciferase reporter assay, RIP assay and biotin pull-down assay. Results Here, we demonstrated that AGAP2-AS1 expression was up-regulated in HCC tissues and cell lines, especially in metastatic and recurrent cases. Gain- and loss-of-function experiments indicated that AGAP2-AS1 promoted cell proliferation, migration, invasion, EMT progression and inhibited apoptosis of HCC cells in vitro and in vivo. Further studies demonstrated that AGAP2-AS1 could function as a competing endogenous RNA (ceRNA) by sponging miR-16-5p in HCC cells. Functionally, gain- and loss-of-function studies showed that miR-16-5p promoted HCC progression and alteration of miR-16-5p abolished the promotive effects of AGAP2-AS1 on HCC cells. Moreover, ANXA11 was identified as direct downstream targets of miR-16-5p in HCC cells, and mediated the functional effects of miR-16-5p and AGAP2-AS1 in HCC, resulting in AKT signaling activation. Clinically, AGAP2-AS1 and miR-16-5p expression were markedly correlated with adverse clinical features and poor prognosis of HCC patients. We showed that hypoxia was responsible for the overexpression of AGAP2-AS1 in HCC. And the promoting effects of hypoxia on metastasis and EMT of HCC cells were reversed by AGAP2-AS1 knockdown. Conclusions Taken together, this research supports the first evidence that AGAP2-AS1 plays an oncogenic role in HCC via AGAP2-AS1/miR-16-5p/ANXA11/AKT axis pathway and represents a promising therapeutic strategy for HCC patients.

Amount of evidence demonstrate that aberrant microRNAs (miRNAs) are involved in tumorigenesis and progression in hepatocellular carcinoma (HCC). Among them, miR-645 is recently recognized as cancer-related miRNA and its significance in HCC remains largely unknown. In this study, we reported for the first that miR-645 expression was markedly elevated in HCC tissues and cell lines, and its up-regulation was associated with malignant clinical features, including tumor size and venous infiltration and poor prognosis. Our data revealed that miR-645 promoted cell proliferation, colony formation and inhibited apoptosis by gain- and loss-of function experiments in vitro. In vivo assays showed that miR-645 overexpression enhanced tumor growth. Moreover, miR-645 directly bound to the SOX30 3'-UTR and post-transcriptionally repressed SOX30 expression in HCC cells. Furthermore, miR-645 inversely correlated with SOX30 expression in HCC tissues. Restoration of SOX30 expression at least partially abolished the biological effects of miR-645 on HCC cells. SOX30 regulated HCC progression through aberrant activation of p53 by directly binding to its promoter. Taken together, this research supports the first evidence that miR-645 exerts an oncogenic role in HCC progression and may be a therapeutic target for HCC treatment.

Background Aberrant expression of miR-532-3p was involved in progression and development of multiple cancers, whereas miR-532-3p has not been reported in hepatocellular carcinoma (HCC). The aim of this study was to elucidate the functions of miR-532-3p in progression of HCC. Methods Real-time PCR in HCC tissues and cell lines and database analysis were conducted for detection of the expression of miR-532-3p in HCC. Then, the association of miR-532-3p with clinicopathological features and prognosis of HCC patients were statistically measured. Subsequently, we attempted to observe the effects of miR-532-3p on migration, invasion and proliferation of HCC cells by Wound healing assay, Transwell assays, MTT assay and EdU assay. Furthermore, bioinformatics tools, database analysis, luciferase reporter gene assay and rescue experiments were conducted to explore the target of miR-532-3p in HCC, and to explore whether the target mediated the effects of miR-532-3p on HCC cells. Results Our findings and data from databases consistently indicated that the miR-532-3p expression level was higher in HCC. In addition, high miR-532-3p expression was found to be closely related to larger tumor size (P = 0.0027), presence of vascular invasion (P = 0.015), and advanced TNM stage (P = 0.015). In addition, experiments in vitro revealed that miR-532-3p promotes migration, invasion and proliferation of HCC cells. Furthermore, receptor protein tyrosine phosphatase T (PTPRT) was identified as the target and mediator of miR-532-3p in HCC cells. Conclusion Our results demonstrate that miR-532-3p, which is frequently up-regulated in HCC, contributes to HCC cells mobility and proliferation through targeting PTPRT.

Accumulating evidence has demonstrated that aberrant microRNA (miRNA) expression is involved in hepatocellular carcinoma (HCC) progression. Previous findings suggested that miRNA (miR)-875-5p participates in the development of various types of cancer. However, the expression and function of miR-875-5p in HCC remains largely unclear. The analysis of clinical samples in the present study demonstrated that miR-875-5p expression was downregulated in HCC tissues compared to adjacent non-tumor tissues, which was associated with a large tumor size, venous infiltration, advanced tumor-node-metastasis stage and unfavorable overall survival. In vitro experiments revealed that ectopic expression of miR-875-5p suppressed, whereas inhibition of miR-875-5p promoted HCC cell proliferation, migration, invasion and epithelial-to-mesenchymal transition (EMT) progression. Overexpression of miR-875-5p restrained HCC tumor growth and metastasis in vivo. Mechanistically, eukaryotic translation initiation factor 3 subunit a (eIF3a) was identified as the downstream target of miR-875-5p in HCC. Further experiments demonstrated that the expression of eIF3a was upregulated and negatively correlated with that of miR-875-5p in HCC tissues. In addition, miR-875-5p negatively regulated the luciferase activity of wild-type, but not mutant 3′-untranslated region (3′UTR) of eIF3a mRNA. miR-875-5p suppressed eIF3a expression at the mRNA and protein level in HCC cells. Additionally, eIF3a exerted an oncogenic role, and knockdown of eIF3a inhibited the proliferation, motility and EMT of HCC cells. In addition, eIF3a overexpression abolished the inhibitory effects of miR-875-5p on the proliferation, motility and EMT in HCC cells. In conclusion, miR-875-5p, which was downregulated in HCC, may inhibit tumor growth and metastasis by eIF3a downregulation via targeting its 3′UTR and may be a promising prognostic and therapeutic strategy in HCC.

Background: Long non-coding RNAs (lncRNAs) have obtained growing attention due to their potential effects as novel regulators in various tumors. This study aimed to investigate the expression and roles of lncRNA ZFPM2-AS1 in the progression of hepatocellular carcinoma (HCC). Methods: Transwell was used to determine migration and invasion of HCC cells in vitro. The lung metastasis mouse model was established to detect tumor metastasis of HCC in vivo. The direct binding of miR-3612 to 3'UTR of DAM15 was confirmed by luciferase reporter assay. The expression of ZFPM2-AS1 and miR-3612 in HCC specimens and cell lines were detected by real-time PCR. The correlation among ZFPM2-AS1 and miR-3612 were disclosed by a dual-luciferase reporter assay, RIP assay and biotin pull-down assay.Results: In present study, we found that ZFPM2-AS1 was up-regulated in HCC tissues and cells and its upregulation was associated with TNM stage, vascular invasion, and poor prognosis of HCC patients. Functionally, gain- and loss-of-function experiments indicated that ZFPM2-AS1 promoted cell migration, invasion and EMT progress in vitro and in vivo. ZFPM2-AS1 could function as a competing endogenous RNA (ceRNA) by sponging miR-3612 in HCC cells. Mechanically, miR-3612 inhibited HCC metastasis and alternation of miR-3612 reversed the promotive effects of ZFPM2-AS1 on HCC cells. In addition, we confirmed that ADAM15 was a direct target of miR-3612 in HCC and mediated the biological effects of miR-3612 and ZFPM2-AS1 in HCC. Curcumin, an active derivative from turmeric, exerts its anticancer effects through ZFPM2-AS1/miR-3612/ADAM15 pathway. Our data identified ZFPM2-AS1 as a novel oncogenic lncRNA and correlated malignant clinical outcomes in HCC patients. Conclusions: ZFPM2-AS1 performed as oncogenic role via targeting miR-3612 and subsequently promoted ADAM15 expression in HCC. Our results revealed that ZFPM2-AS1 could be a potential prognostic biomarker and therapeutic target for HCC.

Accumulating evidence show that long non-coding RNAs (lncRNAs) are identified as regulators in tumor progression and development. Here, we elucidated the function and possible molecular mechanisms of the effect of lncRNA-PICSAR (p38 inhibited cutaneous squamous cell carcinoma associated lincRNA) on the biological behaviors of HCC. In present study, we demonstrated that PICSAR was up-regulated in HCC tissues and cells and correlated with progression and poor prognosis in HCC patients. Gain- and loss-of-function experiments indicated that PICSAR enhanced cell proliferation, colony formation, cell cycle progression and inhibited apoptosis of HCC cells. PICSAR could function as a competing endogenous RNA (ceRNA) by sponging miR-588 in HCC cells. Mechanically, miR-588 inhibited HCC progression and alternation of miR-588 reversed the promotive effects of PICSAR on HCC cells. In addition, we confirmed that eukaryotic initiation factor 6 (EIF6) was a direct target of miR-588 in HCC and mediated the biological effects of miR-588 and PICSAR in HCC, resulting in PI3K/AKT/mTOR pathway activation. Our data identified PICSAR as a novel oncogenic lncRNA and correlated malignant clinical outcomes in HCC patients. PICSAR performed as oncogenic role via targeting miR-588 and subsequently promoted EIF6 expression and PI3K/AKT/mTOR activation in HCC. Our results revealed that PICSAR could be a potential prognostic biomarker and therapeutic target for HCC.

Tuftelin1 (TUFT1), an acidic protein constituent of developing and mineralizing tooth tissues, is regulated by hypoxia and the Hedgehog signaling pathway. We investigated the role of TUFT1 in hepatocellular carcinoma (HCC). qRT-PCR, immunohistochemistry and western blot were employed to evaluate TUFT1 level in HCC. MTT, BrdU, 3D culture and Transwell assays were used to assess cell viability, proliferation, in vitro growth, migration, and invasion. Subcutaneous and tail vein injection models were established to investigate in vivo growth and metastasis. Chromatin immunoprecipitation was performed to assess binding of hypoxia-inducible factor 1α (HIF-1α) to TUFT1 promoter. A microRNA array was used to identify hypoxia-related microRNAs. TUFT1 was elevated in HCC, and correlated with unfavorable clinicopathologic characteristics and poor survival. TUFT1 promoted HCC cell growth, metastasis and epithelial-mesenchymal transition in vitro and in vivo via activation of Ca2+/PI3K/AKT pathway. Hypoxia induced TUFT1 expression in an HIF-1α dependent manner, and TUFT1 expression was positively correlated with HIF-1α level in HCC tissues. Hypoxiaenhanced TUFT1 expression by downregulating miR-671-5p rather than by directly promoting the binding of HIF-1α to TUFT1 promoter. MiR-671-5p interacted with the 3'-UTR of TUFT1 mRNA and subsequently inhibited TUFT1 expression. Consequently, knockdown of TUFT1 blocked the effects of hypoxia in promoting HCC progression. TUFT1 promoted the growth, metastasis and EMT of HCC cells through activating Ca2+/PI3K/AKT pathway. The hypoxic microenvironment increased the expression of TUFT1 via downregulation of miR-671-5p. TUFT1 may function as a potential therapeutic target for the intervention and treatment of HCC.

Rational: Patients with hepatocellular carcinoma (HCC) have a poor prognosis mostly due to intrahepatic as well as distal metastasis. Vasodilator-stimulated phosphoprotein (VASP), a regulator of actin cytoskeleton and cell migration, is overexpressed in HCC and correlated with its malignant features and poor prognosis. Very little is known about its function in HCC. Methods: qRT-PCR, Western blot and IHC were used to detect the VASP expression in tissues and cells. Transwell and wound healing assays were used to measure the migration and invasion of HCC cells. Immunoblotting and immunofluorescence were used for detection of epithelial-to-mesenchymal transition (EMT) progression in HCC cells. A lung metastasis mouse model was used to evaluate metastasis of HCC in vivo. The putative targets of miR-204 were disclosed by public databases and a dual-luciferase reporter assay. IP was used to show the interaction between VASP and CRKL. ChIP was used to analyze the binding of HIF-1α to VASP promoter region. Results: Our data involving both gain- and loss-of-function studies revealed that VASP activated AKT and ERK signaling and promoted HCC migration and invasion in vitro and in vivo by altering the EMT phenotype and expression of MMPs. We investigated the positive correlation between VASP and an adapter protein, CRKL. VASP dynamically co-localized at the SH3N domain of CRKL and mediated its function. Mechanistically, VASP overexpression at the transcriptional level was mediated by HIF-1α through direct binding to two hypoxia response elements (HRE) in the VASP promoter region. Furthermore, we identified hypoxia-induced down-regulation of miR-204, which functioned as the regulator of VASP overexpression at the post-transcriptional level. Also, hypoxia-activated p-Smad3 dependent TGF-β signaling indirectly promoted VASP expression. Conclusion: A variety of hypoxia-induced molecular mechanisms contributed to the upregulation of VASP at transcriptional and post-transcriptional levels. These mechanisms involved CRKL, HIF-1α, miR-204, and TGF-β activating the AKT and ERK signaling to promote EMT and expression of MMPs. Taken together, our results defined VASP as an oncogene of HCC pathogenesis and metastasis with the potential to serve as a prognostic biomarker.

Background Heat shock protein 90 (HSP90) functions as a well-known onco-protein to regulate protein conformation, stability and degradation. Pyruvate kinase M2 (PKM2), a critical regulator of the metabolism, growth and metastasis of cancer cells, has been confirmed to be overexpressed in various human cancer including hepatocellular carcinoma (HCC). However, the molecular mechanisms underlying the oncogenic functions of HSP90 and PKM2 overexpression in HCC remain unknown. Methods The expression of HSP90 and PKM2 in HCC specimens and cells were detected by immunoblotting and immunostaining. The interaction between HSP90 and PKM2 was confirmed by tandem affinity purification, co-immunoprecipitation and Glutathione S transferase (GST)-pulldown assay. Results In this study, we found that HSP90 could bind to PKM2 and subsequently increased PKM2 abundance in HCC cells. Immunohistochemistry (IHC) staining showed that HSP90 level was positively correlated with PKM2 level in HCC tissues. Mechanistically, HSP90 was found to increase the phosphorylation of PKM2 at Thr-328. Protein kinase glycogen synthase kinase-3β (GSK-3β) formed a protein complex with HSP90 and PKM2, and directly mediated Thr-328 phosphorylation of PKM2 induced by HSP90. Thr-328 phosphorylation was critical for maintaining PKM2 stability and its biological functions in regulating glycolysis, mitochondria respiration, proliferation and apoptosis. Functionally, we found that HSP90 promoted the glycolysis and proliferation and inhibited apoptosis of HCC cells in a PKM2 dependent manner. In vivo experiments disclosed that PKM2 was required for the promoting effects of HSP90 on the growth of HCC cells in mice. Furthermore, we demonstrated that positive expression of HSP90 and PKM2 was correlated with poor clinicopathological features including high alpha fetoprotein (AFP) level, large tumor size, portal vein tumor thrombus (PVTT) and advanced tumor-node-metastasis (TNM) stage. Furthermore, we demonstrated that positive expression of HSP90 and PKM2, and a combination of these proteins could strongly predict the poor prognosis of HCC patients. Conclusions We suggest that HSP90 potentiates the glycolysis and proliferation, reduces the apoptosis and thus enhances the growth of HCC cells through PKM2. Electronic supplementary material The online version of this article (10.1186/s12943-017-0748-y) contains supplementary material, which is available to authorized users.

// Meng Xu 1, * , Zhikui Liu 1, * , Cong Wang 1, * , Bowen Yao 1 and Xin Zheng 1 1 Department of Hepatobiliary Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi 710061, China * Co-first author Correspondence to: Xin Zheng, email: xin.zheng.xjtu@gmail.com Keywords: EDG2, HCC, post-surgical prognosis, PI3K/AKT/mTOR, EMT Received: April 28, 2017 Accepted: June 28, 2017 Published: August 02, 2017 ABSTRACT HCC is the leading type of the malignant liver tumors with the unsatisfied prognosis. Liver resection has been considered as the predominant curative therapy, however, the post-surgical prognostic evaluation remains an urgent problem and the mechanism of HCC metastases has not been understood completely. EDG2 has been found to accelerate tumor progression through mediating different cell pathways, however, it remains unclear about the role of EDG2 on hepatocarcinogenesis. Here, EDG2 expression was found increased notably in HCC tissues by immunohistochemistry compared with adjacent liver tissues and comparison of survival curves revealed that EDG2 upregulation in HCC tissues was associated with the worse prognosis after liver resection. The positive correlation between EDG2 up-regulation and EMT was observed in HCC samples. Furthermore, EDG2 over-expression in HCC cells brought the typical EMT characteristics including up-regulation of Vimentin, Fibronectin and N-cadherin, suppression of E-cadherin, and enhanced cell migration and invasion capacities. Knockdown of EDG2 reversed the EMT phenotype in HCC cells. The in vivo experiments also identified the oncogenic role of EDG2 on HCC growth. The mechanistic studies elucidated that EDG2 enhanced mTOR phosphorylation via PI3K/AKT signaling and consequently induced EMT of HCC cells. Moreover, EDG2 was found to promote cell viability and proliferation of HCC cell through PI3K/AKT/mTOR/Skp2/p27 Kip1 signaling. Taken together, the data here demonstrated EDG2 was a potential predictor for HCC patients receiving liver resection and accelerated HCC progression via regulating EMT driven by PI3K/AKT/mTOR signaling.

// Hang Tuo 1, * , Yufeng Wang 1, * , Liang Wang 1 , Bowen Yao 1 , Qing Li 1 , Cong Wang 1 , Zhikui Liu 1 , Shaoshan Han 1 , Guozhi Yin 1 , Kangsheng Tu 1 and Qingguang Liu 1 1 Department of Hepatobiliary Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi Province 710061, China * These authors contributed equally to this work Correspondence to: Qingguang Liu, email: liuqingguang@vip.sina.com Kangsheng Tu, email: tks0912@foxmail.com Keywords: hepatocellular carcinoma, miR-324-3p, DACT1, proliferation, Wnt/β-catenin pathway Received: June 22, 2017 Accepted: July 26, 2017 Published: August 07, 2017 ABSTRACT Recently, it has been reported that miR-324-3p participates in regulation of the carcinogenesis and tumor progression in various cancers. However, the expression and function of miR-324-3p in hepatocellular carcinoma (HCC) remain unclear. In the current study, miR-324-3p expression was significantly up-regulated in HCC tissues and cell lines. HCC patients with high miR-324-3p level showed poor prognostic features and shorter overall survival and disease-free survival. And in vitro and in vivo experiments revealed that miR-324-3p promoted cell viability, colony formation, proliferation and cell cycle progression of HCC cells. Further studies demonstrated that miR-324-3p could directly target DACT1 (dishevelled binding antagonist of beta catenin 1) and negatively regulated its expression in HCC cells. And rescue experiments revealed that DACT1 could reverse the effects of miR-324-3p on HCC cells. Furthermore, the accumulation of both cytoplasmic and nuclear β-catenin as well as its downstream targets including c-Myc and cyclin D1 could be positively regulated by miR-324-3p. The regulatory effects of miR-324-3p on β-catenin, c-Myc and cyclin D1 levels could be reversed by DACT1. Overall, we concluded that miR-324-3p could promote tumor growth through targeting DACT1 and activation of Wnt/β-catenin pathway in HCC. MiR-324-3p may be a ponderable and promising therapeutic target for HCC.

Lymphocyte-specific protein 1 (LSP1) has been reported to regulate cell biology in several human cancers including lymphoma and breast cancer. However, the functions of LSP1 in human hepatocellular carcinoma (HCC) are still unknown. In this study, we found that LSP1 expression was downregulated in HCC tissues and cell lines, and lower LSP1 expression was correlated with poor clinicopathological features including large tumor size, high Edmondson-Steiner grading and advanced tumor-node-metastasis (TNM) stage. Additionally, we demonstrated that patients with high LSP1 expression had significantly better overall survival and disease-free survival. Moreover, LSP1 was found to be an independent factor for predicting the prognosis of HCC patients. In vitro and in vivo assays showed that overexpressing LSP1 inhibited HCC growth by inducing both apoptosis and growth arrest. Mechanistically, we found that expression of phosphorylated extracellular regulated protein kinases 1 and 2 (ERK1/2) was downregulated after LSP1 overexpression, indicating LSP1 could suppress HCC growth by inhibiting the ERK pathway in HCC cells. Taken together, these results indicate that LSP1 may serve as a prognostic marker and a potential therapeutic target in human HCC.

The heterogeneity existing in tumours is responsible for the poor response to treatment. Therefore, elucidating the molecular mechanisms of intratumoural heterogeneity in hepatocellular carcinoma (HCC) is vital for the discovery of new therapeutic methods for improving the prognosis of patients. Of note, cancer stem cells (CSCs) existing in HCC may explain the pathological properties of heterogeneity and recurrence. An increasing number of studies have confirmed that abnormally expressed microRNAs (miRNAs) take part in the carcinogenesis as well as the aggravation of HCC. However, little information is currently available about the specific miR-4319 in HCC. Herein, we demonstrated that the level of miR-4319 was remarkably decreased in HCC specimens and cells compared to that in normal counterparts and that the depression of miR-4319 in tumour specimens correlates with tumour size, histological grade and venous invasion. Through a series of functional experiments, we illustrated that miR-4319 repressed cell proliferation, accelerated apoptosis, inhibited epithelial-mesenchymal transition (EMT) and prevented cancer stemness in HCC cells by targeting FOXQ1 (Forkhead box Q1). An in vivo tumourigenesis assay uncovered that depletion of miR-4319 in Hep3B cells increased tumour growth and elevated the expression of EMT and CSC markers in comparison to those of the control group. Restoration of FOXQ1 expression also partially reversed the miR-4319-induced biological effects on HCC cells. Thus, miR-4319, as a posttranscriptional regulator, plays a profound role in suppressing the malignant progression of HCC, and our study highlights the miR-4319/FOXQ1 cascade as a potential therapeutic target for conquering HCC.

Transarterial chemoembolization (TACE) has been considered the standard treatment for intermediate-stage hepatocellular carcinoma according to BCLC algorithm. However, it has been unclear about the TACE-related predictive bio-markers and underlying molecular mechanisms. This investigation revealed that HCCs with higher HIF-1α suffered from unfavorable OS after TACE. mRNA expression microarray revealed that HIF-1α was potential target of p-STAT3 which was verified by ChIP and immunoblotting assay. Activation of IL-6/STAT3/HIF-1α signaling was found to promote EMT and chemoresistance to Doxorubicin in vitro and in vivo by regulating SNAI1. Hypoxia did not enhance HIF-1α expression and influence cell growth and chemoresistence to Doxorubicin in HCC cells when STAT3 expression was abolished. Taken together, HIF-1α overexpression in HCC tissues predicted the unfavorable outcome of HCCs after TACE and IL-6/STAT3 pathway resulted in EMT induced-metastases and chemoresistance of HCC after TACE through HIF-1α/SNAI1 axis.

Buildings in Southern China widely use a double-skin roof to reduce heat entry through the roof to the building interior during summertime. Concrete roof tiles are preferably installed as the outmost layer of the double-skin roof due to their resistance to hail and wind damages and their attractive price. However, after construction, the tile’s top tends to be darkened by dust deposit and algae growth, increasing the heat entry through the roof to the building. Here, we show that this heat entry can be curtailed by lowering the emissivity at the tile’s underside. Temperatures and heat fluxes at different elevations of a double-skin roof with concrete tiles as the outmost layer of the roof are monitored. The underside of each concrete tile is coated with a specific paint to get a unique emissivity. Observations reveal that lowering the emissivity of concrete roof tiles could cut down the summer heat gain of buildings in tropical regions.

Table S1. A list of the utilized primary antibodies (DOCX 18 kb)

Table S2. Primers sequences for real-time PCR analysis (DOCX 18 kb)

Rocky desertification induced by soil erosion is a serious geological hazard in the karst regions of southwestern China. This paper made a research on small watersheds of Chengjiang River in Gongcheng County, Guilin City, Guangxi Province, analyzed the causes and trends of soil erosion in the regions by comparing rocky desertification areas in 1991, 2006 and 2015, and proposed countermeasures against soil erosion based on the above analysis. This research provides a basis for restoring ecological environments and preventing soil erosion in small watersheds of karst regions.

Meng Xu, Yuli Jia, Zhikui Liu, Linglong Ding, Run Tian, Hua Gu, Yufeng Wang, Hongyong Zhang, Kangsheng Tu, Qingguang Liu Department of Hepatobiliary Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, People’s Republic of China Abstract: Several studies have revealed that the abnormal expression of chromatin assembly factor 1, subunit A (P150) (CHAF1A) was involved in the development of some types of malignant tumors. However, CHAF1A expression and its role in hepatocellular carcinoma (HCC) remain poorly characterized. In this study, we first investigated CHAF1A expression in six cell lines and 116 pairs of HCC and matched normal tumor-adjacent tissues to evaluate the clinicopathological characteristics of CHAF1A in HCC. Then, we detected the proliferation and apoptosis in HCC cells. In addition, a subcutaneous tumor model in nude mice was performed to evaluate tumor growth in vivo. We found that the expression of CHAF1A was significantly higher in HCC tissues than that in adjacent nontumor tissues (P

// Zhikui Liu 1, * , Changwei Dou 1, * , Bowen Yao 1 , Meng Xu 1 , Linglong Ding 1 , Yufeng Wang 1 , Yuli Jia 1 , Qing Li 1 , Hongyong Zhang 1 , Kangsheng Tu 1 , Tao Song 1 , Qingguang Liu 1 1 Department of Hepatobiliary Surgery, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi, China * These authors contributed equally to this work Correspondence to: Kangsheng Tu, email: tks0912@foxmail.com Tao Song, email: 13572431619@163.com Qingguang Liu, email: liuqingguang@vip.sina.com Keywords: miR-129-2, methylation, high mobility group box 1, invasion, hepatocellular carcinoma Received: November 22, 2015 Accepted: April 16, 2016 Published: May 15, 2016 ABSTRACT Aberrant expression of microRNAs (miRNAs) and its dysfunction have been revealed as crucial modulators of cancer initiation and progression. MiR-129-2 has been reported to play a tumor suppressive role in different human malignancies. Here, we demonstrated that miR-129-2 was significantly decreased in hepatocellular carcinoma (HCC) tissues and cell lines. Furthermore, miR-129-2 was expressed at significant lower levels in aggressive and recurrent tumor tissues. Clinical analysis indicated that miR-129-2 expression was inversely correlated with venous infiltration, high Edmondson-Steiner grading and advanced tumor-node-metastasis (TNM) stage in HCC. Notably, miR-129-2 was an independent prognostic factor for indicating overall survival (OS) and disease-free survival (DFS) of HCC patients. Ectopic expression of miR-129-2 inhibited cell migration and invasion in vitro and in vivo . Furthermore, we confirmed that high mobility group box 1 (HMGB1) was a direct target of miR-129-2, and it abrogated the function of miR-129-2 in HCC. Mechanistic investigations showed that miR-129-2 overexpression inhibited AKT phosphorylation at Ser473 and decreased the expression of matrix metalloproteinase2/9 (MMP2/9). Upregulation of p-AKT abolished the decreased cell migration and invasion induced by miR-129-2 in HCC. Whereas inhibition of Akt phosphorylation significantly decreased HMGB1-enhanced HCC cell migration and invasion. Moreover, we found that miR-129-2 was downregulated by DNA methylation, and demethylation of miR-129-2 increased miR-129-2 expression in HCC cells and resulted in significant inhibitory effects on cell migration and invasion. In conclusion, miR-129-2 may serve as a prognostic indicator for HCC patients and exerts tumor suppressive role, at least in part, by inhibiting HMGB1.

The p300-CBP-associated factor (PCAF), other than its histone acetyltransferase (HAT) activity, possesses an intrinsic ubiquitination activity that is involved in various transcriptional regulators, including the transcription factor glioma-associated oncogene 1 (Gli1), a well-known regulator of epithelial-mesenchymal transition (EMT) in cancer. In present research, we detected that PCAF was down-regulated in hepatocellular carcinoma (HCC) tissues compared with the adjacent non-tumor tissues and significantly associated with malignant portal vein invasion (p 0.05) and poor survival (p 0.05) of HCC patients. Moreover, functional study demonstrated that downregulation of PCAF facilitated tumor cell migration, invasion via EMT. Further study found that Gli1 as a direct target of PCAF induced EMT and promoted tumor metastasis and invasion.PCAF is an anti-oncogene that plays an important role in the development of HCC by suppressing HCC cell metastasis and EMT by targeting Gli1, which indicates the potential therapeutic value of PCAF for suppression of metastasis of HCC.

Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. Accumulating studies have demonstrated that aberrant expression of several lncRNAs was found to be involved in the hepatocarcinogenesis. In this study, a lncRNA Ftx was chosen to investigate its effects on HCC cells, and clarify the possible mechanism. We demonstrated that the lncRNA Ftx and Ftx-derived miR-545 were up-regulated in both HCC tissues and cells. MiR-545 was positively correlated with lncRNA Ftx expression. Notably, clinical association analysis revealed that the high expression of lncRNA Ftx and miR-545 was associated with poor prognostic features, and conferred a reduced 5-year overall survival (OS) and disease-free survival (DFS) of HCC patients. We found that miR-545 was a pivotal mediator in Ftx-induced promotion of HCC cell growth. Subsequently, we identified RIG-I as a direct target of miR-545. The expression of RIG-I was downregulated in HCC tissues and was inversely correlated with miR-545 expression. Our data revealed that ectopic expression of RIG-I abrogated the effects of lncRNA Ftx or miR-545 on HCC cells. LncRNA Ftx/miR-545-mediated downregulation of RIG-I led to increased Akt phosphorylation in vitro and in vivo. Inhibition of Akt phosphorylation abolished the effects of lncRNA Ftx/miR-545 on HCC cells. In conclusion, our study demonstrates that the novel pathway lncRNA Ftx/miR-545/RIG-I promotes HCC development by activating PI3K/Akt signaling, and it may serve as a novel prognostic biomarker and therapeutic target for HCC.

MicroRNAs (miRNAs) have been found to play fundamental roles in the pathogenesis of hepatocellular carcinoma (HCC). Previous miRNA array data showed that miR-519a was upregulated in HCC tissues compared to adjacent non-tumor tissues. However, the functional role of miR-519a in HCC remains unexplored. In this study, we demonstrated that the expression of miR-519a was elevated in both HCC tissues and cell lines. Clinical association analysis revealed that high expression of miR-519a was correlated with adverse clinicopathological characteristics including large tumor size, high Edmondson-Steiner grading, advanced tumor-node-metastasis (TNM) tumor stage and venous infiltration. Furthermore, high expression of miR-519a conferred a reduced 5-year overall survival and disease-free survival of HCC patients. Moreover, we disclosed that miR-519a overexpression promoted, but miR-519a silencing reduced, HCC cell proliferation and cell cycle progression in vitro. Notably, we identified phosphatase and tensin homolog (PTEN) as a direct downstream target and functional mediator of miR-519a in HCC cells. Mechanistically, phosphatidylinositol-3-OH kinase (PI3K)/AKT pathway, downstream of PTEN, is essential for the functional roles of miR-519a in HCC cells. In conclusion, our results indicate that miR-519a promotes tumor growth of HCC by targeting PTEN-mediated PI3K/AKT pathway, and potentially serves as a novel prognostic biomarker and therapeutic target for HCC.

Background Cartilage oligomeric matrix protein (COMP) is known to promote fibrosis in skin, lung and liver. Emerging evidence shows that COMP plays critical roles in tumor development, including breast cancer, colon cancer and hepatocellular carcinoma (HCC). Nevertheless, the role of COMP in HCC proliferation and metastasis and its underlying mechanisms remain fully unclear. Methods Serum COMP was determined by ELISA. Cell Counting Kit-8 and plate colony formation were performed to evaluate cell proliferation. Wound healing and transwell assays were used to determine migration and invasion of HCC cells. Western blotting and immunofluorescence were carried out for detection of epithelial-to-mesenchymal transition (EMT) markers and MMPs in HCC cells. The in vivo role of COMP was evaluated using mouse models. We also measured effects of hepatic stellate cells (HSCs)-conditioned medium (CM) on HCC progression using transwell coculture system. Results Here, we found that serum COMP levels in HCC patients were significantly higher than those in healthy controls. Accordingly, high serum COMP levels in HCC patients significantly correlated with malignant clinical characteristics and poor clinical outcomes. Next, we investigated that recombinant human COMP protein (rCOMP) treatment resulted in increased abilities of proliferation, invasion and migration of HCC cells. Furthermore, rCOMP treatment enhanced proliferative and metastatic colonization of HCC cells in vivo. Mechanistically, CD36 receptor played an essential role in COMP-mediated HCC cell proliferation and metastasis. Functionally, COMP/CD36 signaling caused phosphorylation of ERK and AKT, resulting in the upregulation of tumor-progressive genes such as EMT markers, MMP-2/9, Slug and Twist in HCC cells. Interestingly, we revealed that COMP was secreted by HSCs. CM of LX2 cells with COMP knockdown showed weaker effects on the activation of MEK/ERK and PI3K/AKT signaling pathways in HCC cells compared to control CM. Conclusions Our findings indicated that HSCs-derived COMP collaborated with CD36 and subsequently played an essential role in MEK/ERK and PI3K/AKT-mediated HCC progression. COMP might act as a promising target for the diagnosis and treatment of aggressive HCC.

Transforming growth factor β (TGFβ) potently activates hepatic stellate cells (HSCs), which promotes production and secretion of extracellular matrix (ECM) proteins and hepatic fibrogenesis. Increased ECM synthesis and secretion in response to TGFβ is associated with endoplasmic reticulum (ER) stress and the unfolded protein response (UPR). TGFβ and UPR signaling pathways are tightly intertwined during HSC activation, but the regulatory mechanism that connects these two pathways is poorly understood. Here, we found that TGFβ treatment of immortalized HSCs (i.e. LX-2 cells) induces phosphorylation of the UPR sensor inositol-requiring enzyme 1α (IRE1α) in a SMAD2/3-procollagen I–dependent manner. We further show that IRE1α mediates HSC activation downstream of TGFβ and that its role depends on activation of a signaling cascade involving apoptosis signaling kinase 1 (ASK1) and c-Jun N-terminal kinase (JNK). ASK1–JNK signaling promoted phosphorylation of the UPR-associated transcription factor CCAAT/enhancer binding protein β (C/EBPβ), which is crucial for TGFβ- or IRE1α-mediated LX-2 activation. Pharmacological inhibition of C/EBPβ expression with the antiviral drug adefovir dipivoxil attenuated TGFβ-mediated activation of LX-2 or primary rat HSCs in vitro and hepatic fibrogenesis in vivo. Finally, we identified a critical relationship between C/EBPβ and the transcriptional regulator p300 during HSC activation. p300 knockdown disrupted TGFβ- or UPR-induced HSC activation, and pharmacological inhibition of the C/EBPβ–p300 complex decreased TGFβ-induced HSC activation. These results indicate that TGFβ-induced IRE1α signaling is critical for HSC activation through a C/EBPβ-p300–dependent mechanism and suggest C/EBPβ as a druggable target for managing fibrosis.

Previous evidences reveal that long non-coding RNA (lncRNA) down syndrome critical region 8 (DSCR8) involves in the progression of multiple cancers. However, the exact expression, function, and mechanism of DSCR8 in hepatocellular carcinoma (HCC) remain uncovered. In this study, real-time PCR in HCC tissues and cell lines indicated that DSCR8 expression was upregulated, while miR-485-5p was downregulated. MTT assay, plate clone formation, Edu assay, flow cytometry, and in vivo experiments indicated that DSCR8 promoted HCC cell proliferation and cycle, whereas accelerated cell apoptosis. Luciferase reporter gene assay, RIP assay, and rescue experiments demonstrated that DSCR8 functioned as a competing endogenous RNA (ceRNA) by sponging miR-485-5p in HCC cells. Furthermore, gain- and loss-of-function studies showed that miR-485-5p activated Wnt/β-catenin signal pathway by targeting Frizzled-7 (FZD7). Moreover, DSCR8 activated Wnt/β-catenin signal pathway to promote HCC progression by DSCR8/miR-485-5p/FZD7 axis. Statistical analysis revealed that DSCR8 and miR-485-5p were closely related to some malignant clinicopathological features and 5-year survival rates of HCC patients. Taken together, the present study reports for the first time that DSCR8 activates Wnt/β-catenin signal pathway to promote HCC progression by DSCR8/miR-485-5p/FZD7 axis. The findings provide promising and valuable strategies for targeted therapy of HCC.

Research has confirmed that abnormally expressed miRNAs are involved in the occurrence and development of hepatocellular carcinoma (HCC). In the present study, we confirmed that miR‑577 expression both in HCC tissues and cell lines was markedly downregulated. Clinically, downregulated expression of miR‑577 is notably related to malignant clinicopathological features, such as venous invasion and advanced TNM stage. Additionally, miR‑577 may act as a valuable tumor marker to predict the prognosis of HCC patients. Through knockdown and overexpression of miR‑577, miR‑577 was identified as an inhibitor of cell metastatic ability and EMT progress in HCC. Furthermore, miR‑577 was able to directly bind to the 3'‑UTR of homeobox A1 (HOXA1) to regulate the expression of HOXA1. In addition, there existed a negative correlation between the expression of miR‑577 and HOXA1 in HCC specimens. Rescue experiments revealed that the influence of miR‑577 on the migration, invasion and EMT of HCC cells was reversed by HOXA1. Taken together, our findings demonstrated that miR‑577 functions as an anti‑oncogene to suppress the migration, invasion and EMT of HCC cells through direct interaction with HOXA1. miR‑577 may act as a valuable target for the molecular‑targeted therapy of HCC.

Background Accumulating evidence confirm that aberrant microRNAs (miRNAs) expression contributes to hepatocellular carcinoma (HCC) development and progression. Previous study reported that miR-1468 showed an up-regulated tendency and might be a potential prognostic biomarker in HCC samples derived from TCGA database. However, the role of miR-1468 and its underlying mechanisms involved in the growth and metastasis of HCC remain poorly investigated. Methods CCK-8, EdU, colony formation and flow cytometry were used to determine proliferation, cell cycle progression and apoptosis of HCC cells in vitro. The subcutaneous tumor model in nude mice was established to detect tumor growth of HCC in vivo. The direct binding of miR-1468 to 3’UTR of Cbp/p300 interacting transactivator with Glu/Asp rich carboxy-terminal domain 2 (CITED2) and Up-frameshift protein 1 (UPF1) was confirmed by luciferase reporter assay. Results Here, we demonstrated that miR-1468 expression was up-regulated in HCC tissues and cell lines. Clinical analysis revealed that increased miR-1468 level was significantly correlated with malignant prognostic features and shorter survival. Gain- and loss-of-function experiments indicated that miR-1468 promoted cell proliferation, colony formation, cell cycle progression and induced apoptosis of HCC cells in vitro and in vivo. Moreover, CITED2 and UPF1 were identified as direct downstream targets of miR-1468 in HCC cells, and mediated the functional effects of miR-1468 in HCC, resulting in peroxisome proliferator-activated receptor-γ (PPAR-γ)/AKT signaling activation. In clinical samples of HCC, miR-1468 inversely correlated with the levels of CITED2 and UPF1, which were confirmed to be down-regulated in HCC. Restoration of CITED2 or UPF1 expression at least partially abolished the biological effects of miR-1468 on HCC cells. Moreover, alteration of PPAR-γ or AKT phosphorylation could reverse the function of miR-1468 in HCC. Conclusions Taken together, this research supports the first evidence that miR-1468 plays an oncogenic role in HCC via activating PPAR-γ/AKT pathway by targeting CITED2 and UPF1, and represents a promising therapeutic strategy for HCC patients.

Tuftelin1 (TUFT1), an acidic protein constituent of developing and mineralizing tooth tissues, is regulated by hypoxia and the Hedgehog signaling pathway. We investigated the role of TUFT1 in hepatocellular carcinoma (HCC). qRT-PCR, immunohistochemistry and western blot were employed to evaluate TUFT1 level in HCC. MTT, BrdU, 3D culture and Transwell assays were used to assess cell viability, proliferation, in vitro growth, migration, and invasion. Subcutaneous and tail vein injection models were established to investigate in vivo growth and metastasis. Chromatin immunoprecipitation was performed to assess binding of hypoxia-inducible factor 1α (HIF-1α) to TUFT1 promoter. A microRNA array was used to identify hypoxia-related microRNAs. TUFT1 was elevated in HCC, and correlated with unfavorable clinicopathologic characteristics and poor survival. TUFT1 promoted HCC cell growth, metastasis and epithelial-mesenchymal transition in vitro and in vivo via activation of Ca

Table S1. Association between clinicopathological parameters and serum COMP level in primary hepatocellular carcinoma. (DOCX 17 kb)

High‑mobility group box 1 (HMGB1), a member of the high‑mobility group protein family, was originally characterized as a non‑histone, nuclear DNA‑binding protein. While the roles of HMGB1 in inflammation and cell differentiation have been previously reported, its role in tumor cell migration and invasion, particularly in hepatocellular carcinoma (HCC), has remained elusive. The present study reported that the expression of HMGB1 in HCC tissues was significantly higher than that in matched tumor‑adjacent tissues (P

MicroRNA-212 (miR-212) has been reported to play oncogenic or tumor suppressive role in different human malignancies. Here, we demonstrated that the mean level of miR-212 in hepatocellular carcinoma (HCC) tissues was significantly lower than that in matched tumor-adjacent tissues. Similarly, the expression of miR-212 was obviously reduced in HCC cell lines as compared with a nontransformed hepatic cell line. Ectopic expression of miR-212 inhibited cell viability and proliferation, and induced apoptosis in HepG2 cells. In contrast, down-regulation of miR-212 increased cell viability and proliferation, and suppressed apoptosis in Bel-7402 cells. In vivo studies showed that miR-212 inhibited tumor growth of HCC via suppressing proliferation and inducing apoptosis. Furthermore, we confirmed that Forkhead box protein A1 (FOXA1) was a direct target of miR-212, and it abrogated the function of miR-212 in HCC. Finally, we disclosed that the aberrant expression of miR-212 and FOXA1 was evidently correlated with poor prognostic features of HCC. MiR-212, FOXA1 and their combination were valuable prognostic markers for predicting survival of HCC patients. In conclusion, miR-212 may serve as a prognostic indicator for HCC patients and exerts tumor suppressive role, at least in part, by inhibiting FOXA1.

Background Recently, it has been reported that long non-coding RNA (lncRNA) cancer susceptibility candidate 2 (CASC2), a novel tumor suppressor, participates in regulating the carcinogenesis and suppresses tumor progression by sponging microRNAs (miRNAs). However, the expression and function of CASC2 in hepatocellular carcinoma (HCC) remain unclear. Methods The expression of CASC2 and miR-367 in HCC specimens and cell lines were detected by real-time PCR. Western blotting and immunohistochemistry were carried out for detection of epithelial-to-mesenchymal transition (EMT) markers in HCC. Transwell assays were used to determine migration and invasion of HCC cells. A mouse model for lung metastasis was established to evaluated HCC metastasis in vivo. The correlation among CASC2, miR-367 and F-box and WD repeat domain containing 7 (FBXW7) were disclosed by a dual-luciferase reporter assay, RIP assay and biotin pull-down assay. Results Here, CASC2 expression was significantly downregulated in HCC tissues, especially in aggressive and recurrent cases. In accordance, CASC2 underexpression was observed in HCC cell lines compared to LO2. In vitro and in vivo experiments revealed that CASC2 inhibited migration and invasion of HCC cells. Additionally, CASC2 repressed EMT process of HCC cells. Further studies demonstrated that CASC2 could function as a competing endogenous RNA (ceRNA) by sponging miR-367 in HCC cells. Functionally, gain- and loss-of-function studies showed that miR-367 promoted migration, invasion and EMT progression of HCC cells. Moreover, further investigations disclosed that FBXW7 was a downstream target of miR-367 and CASC2 prohibited EMT progression and subsequently exerted its anti-metastatic effects via CASC2/miR-367/FBXW7 axis in HCC cells. Clinically, CASC2 underexpression and miR-367 overexpression were closely correlated with the metastasis-associated clinicopathologic features. Notably, CASC2 low-expressing and miR-367 high-expressing HCC patients showed the poorest clinical outcome. Conclusions Overall, we conclude that the CASC2/miR-367/FBXW7 axis may be a ponderable and promising therapeutic target for HCC. Electronic supplementary material The online version of this article (doi:10.1186/s12943-017-0702-z) contains supplementary material, which is available to authorized users.

Background Several investigations have demonstrated that follistatin-like 1 (FSTL1) is implicated in the initiation and progression of diverse cancers. It remains unclear whether FSTL1 acted as a cancer-promoting gene through its overexpression in HCC. Patients and methods We detected FSTL1 protein expression in 210 consecutive HCC cases curatively resected in our hospital between 2004 and 2007. The correlation between FSTL1 expression in HCC tissues and post-surgical prognosis of HCCs was analyzed. The in vitro experiments including apoptosis assessment, MTT, BrdU incorporation ELISA assay, Western immunoblotting, and qRT-PCR were performed to determine the impact of FSTL1 on apoptosis and proliferation abilities of HCC cells and the relevant mechanisms. Results FSTL1 protein was found aberrantly increased in 172 of 210 HCC tissues (81.9%) compared to adjacent liver tissues. FSTL1 overexpression was apparently associated with larger tumor size, advanced TNM staging, portal vein invasion, intra-hepatic metastases. Patients with higher FSTL1 expression in tumors suffered from the worse overall survival rate as assessed by comparison of Kaplan-Meier survival curves. Higher FSTL1 expression in HCC tissues was identified as a independent poor post-surgical prognostic predictor for HCC. Silencing FSTL1 by siRNA promoted cell apoptosis and leaded to suppression of cell viability and proliferation in MHCC97h cells. Furthermore, enforced expression of FSTL1 obtained the opposite results in Huh7 cells. Mechanistic investigation showed that FSTL1 repressed HCC cell apoptosis through AKT/GSK-3β/Bcl2/BAX/Bim signaling. Conclusion These data proved that FSTL1 contributed to unfavorable post-surgical outcome of HCC patients via inhibiting cell apoptosis.

Background Increasing evidences demonstrate that miRNAs contribute to development and progression of hepatocellular carcinoma (HCC). Underexpression of miR-1296 is recently reported to promote growth and metastasis of human cancers. However, the expression and role of miR-1296 in HCC remain unknown. Methods The levels of miR-1296 in HCC tissues and cells were detected by qRT-PCR. Immunoblotting and immunofluorescence were used for detection of epithelial-to-mesenchymal transition (EMT) progression in HCC cells. Transwell assays were performed to determine migration and invasion of HCC cells. A lung metastasis mouse model was used to evaluated metastasis of HCC in vivo. The putative targets of miR-1296 were disclosed by public databases and a dual-luciferase reporter assay. Results We found that the expression of miR-1296 was reduced in HCC tissues and cell lines, and it was associated with metastasis and recurrence of HCC. Notably, miR-1296 overexpression inhibited migration, invasion and EMT progress of HCCLM3 cells, while miR-1296 loss facilitated these biological behaviors of Hep3B cells in vitro and in vivo. In addition, miR-1296 inversely regulated SRPK1 abundance by directly binding to its 3′-UTR, which subsequently resulted in suppression of p-AKT. Either SRPK1 re-expression or PI3K/AKT pathway activation, at least partially, abolished the effects of miR-1296 on migration, invasion and EMT progress of HCC cells. Furthermore, miR-1296 and SRPK1 expression were markedly correlated with adverse clinical features and poor prognosis of HCC patients. We showed that hypoxia was responsible for the underexpression of miR-1296 in HCC. And the promoting effects of hypoxia on metastasis and EMT of HCC cells were reversed by miR-1296. Conclusions Underexpression of miR-1296 potentially serves as a prognostic biomarker in HCC. Hypoxia-induced miR-1296 loss promotes metastasis and EMT of HCC cells probably by targeting SRPK1/AKT pathway. Electronic supplementary material The online version of this article (doi:10.1186/s12943-017-0675-y) contains supplementary material, which is available to authorized users.

Accumulating evidence demonstrates that aberrant miRNAs contribute to hepatocellular carcinoma (HCC) development and progression. However, the roles of various miRNAs in HCC remain to be determined. In present research, we confirmed that a reduced miR-542-3p expression was present in HCC tissues and cell lines. Our clinical analysis revealed that the down-regulated miR-542-3p expression was significantly correlated with poor prognostic features including advanced TNM stage and venous infiltration. Moreover, we confirmed that miR-542-3p was a novel independent prognostic marker for predicting 5-year survival of HCC patients. The ectopic overexpression of miR-542-3p inhibited cell migration, invasion and EMT progress, while down-regulated miR-542-3p reversed the effect. In addition, miR-542-3p could regulate UBE3C by directly binding to its 3'-UTR. In clinical samples of HCC, miR-542-3p inversely correlated with UBE3C, which was upregulated in HCC. Alternation of UBE3C expression at least partially abolished the migration, invasion and EMT progress effects of miR-542-3p on HCC cells. In conclusion, our results indicated that miR-542-3p functioned as a tumor suppressor gene in regulating the EMT and metastasis of HCC via targeting UBE3C, and may represent a novel potential therapeutic target and prognostic marker for HCC.

Increasing evidence confirms that aberrant miRNA expression contributes to breast cancer (BC) development and progression. However, the roles of different miRNAs in BC remain to be explored. In the present study, we demonstrated that miR-1297 expression was increased in BC tissues and cell lines. Our clinical analysis revealed that the upregulated miR-1297 expression was significantly correlated with poor prognostic features including advanced TNM stage and larger tumor size. Moreover, we found that miR-1297 was a novel independent prognostic marker for predicting 5-year survival of BC patients. The ectopic overexpression of miR-1297 promoted cell proliferation, cell cycle progression and inhibited apoptosis while miR-1297 knockdown reversed the effect. In addition, miR-1297 modulated PTEN by directly binding to its 3'-UTR, resulting in activation of AKT signaling. In clinical samples of BC, miR-1297 inversely correlated with PTEN, which was downregulated in BC. Alternation of PTEN expression or AKT inhibitor at least partially abolished the biological effects of miR-1297 on BC cells. In conclusion, our results indicated that miR-1297 functioned as an oncogene in regulating the proliferation, cell cycle and apoptosis of BC via targeting PTEN/PI3K/AKT signaling, and may represent a novel potential therapeutic target and prognostic marker for BC.

The primer sequences used in this study (DOCX 16Â kb)

Objective To observe the expression level of miR-141 in tumor tissues of human hepatocellular carcinoma (HCC) and determine the effect of miR-141 level on cell proliferation, invasion and migration of MHCC-97H cells by upregulation of miR-141. Methods We checked the miR-141 expression level in HCC by real-time quantitative PCR and analyzed the relationship between the expression level of miR-141 and clinical pathological indicators as well as survival rate. MHCC-97H cells were transiently transfected with miR-141 mimics which were artificially synthesized. The proliferation of MHCC-97H cells was detected by MTT assay. Transwell(TM) assay was performed to examine the invasion and migration of MHCC-97H cells. The expression of erythropoietin-producing hepatocellular receptor A2 (EphA2), which was the potential downstream target, was determined by Western blotting and immunohistochemistry. Results The expression level of miR-141 in HCC tissues was significantly lower than that in the adjacent normal tissues, and it was obviously associated with TNM stage, portal vein infiltration and Edmondson degree. Patients in the lower miR-141 group had a worse 3-year survival than those in higher miR-141 group. Overexpression of miR-141 in MHCC-97H cells significantly suppressed cell proliferation, invasion and migration, and inhibited the protein expression of EphA2. Correlation analysis showed that miR-141 level was negatively correlated with EphA2 expression level. Conclusion miR-141 is down-regulated in HCC tissues and it is negatively correlated with EphA2 expression. Its low expression is correlated with the malignant clinical pathological features. miR-141 overexpression down-regulates EphA2 expression and subsequently inhibits the proliferation, invasion and migration of HCC cells.

miR-187-3p, a novel cancer-related microRNA, was previously reported to play promoting or suppressive roles in different malignancies. However, the expression level, biological function, and underlying mechanisms of miR-187-3p in hepatocellular carcinoma (HCC) remain unknown. This study demonstrated that miR-187-3p was significantly down-regulated in HCC tissues and cell lines, and was associated with advanced TNM stage and metastasis in HCC. Functional studies confirmed that miR-187-3p could inhibit the metastasis of HCC both in vitro and in vivo. Moreover, we proved that miR-187-3p could prevent the epithelial-mesenchymal transition (EMT) of HCC cells. Mechanically, S100A4 was a direct downstream target of miR-187-3p, and mediated the functional influence of miR-187-3p in HCC. Furthermore, miR-187-3p and S100A4 expression was evidently correlated with adverse clinical features and poor prognosis of HCC. Lastly, we showed that hypoxia was responsible for the significantly decreased level of miR-187-3p in HCC, and miR-187-3p was involved in the promoting effects of hypoxia on the metastasis and EMT of HCC cells. Taken together, miR-187-3p inhibits the metastasis and EMT in HCC by targeting S100A4. miR-187-3p can serve as a prognostic indicator and a promising therapeutic target for HCC patients.

To investigate the level and clinical significance of miR-491 expression in human hepatocellular carcinoma (HCC) and the underlying mechanisms.Real-time quantitative PCR was applied to detect the expression of miR-491 in HCC and the corresponding tumor-adjacent tissues. The proliferation of MHCC-97H cells was tested by MTT assay. TranswellTM assay was performed to measure the invasion and migration of MHCC-97H cells. The expression of targeting protein for Xklp2 (TPX2) was determined by Western blotting and immunohistochemistry.The expression level of miR-491 in HCC tissues was significantly lower than that in the adjacent normal tissues, and it was obviously associated with tumor size, TNM stage, portal vein infiltration and Edmondson degree. Patients in the lower miR-491 group had a worse 3-year survival than those in higher miR-491 group. Overexpression of miR-491 in MHCC-97H cells markedly suppressed cell proliferation, invasion and migration, and downregulated the protein expression of TPX2. Correlation analysis showed that miR-491 level was negatively correlated with TPX2 expression level in HCC tissues.The expression of miR-491 in HCC tissue is significantly lower than that in tumor-adjacent tissues and correlated with the malignant clinical pathological features. miR-491 inhibits HCC cell proliferation, invasion and migration by downregulating the expression of TPX2.