miR-199a-5p inhibits proliferation and induces apoptosis in hemangioma cells through targeting HIF1A.

MicroRNAs (miRNAs) exhibit a crucial role in the regulation of angiogenesis and tumor progression, of which miR-199a-5p (miR-199a) has been reported to function as a tumor suppressor in multiple malignancies. However, the precise mechanisms underlying miR-199a in hemangiomas (HAs) remain elusive. In this study, we found that miR-199a had low expression level, while proliferating cell nuclear antigen (PCNA) had high expression level in proliferating-phase HAs compared with the involuting-phase HAs and normal tissues. Spearman correlation analysis revealed the negative correlation of miR-199a with PCNA expression in proliferating-phase HAs. In vitro experiments showed that restoration of miR-199a suppressed cell proliferation capability and induced cell apoptosis in HA-derived endothelial cells (HDEC) and CRL-2586 EOMA cells, followed with decreased PCNA expression and increased cleaved caspase-3 expression, but miR-199a inhibitor reversed these effects. Furthermore, HIF1A was identified as a target of miR-199a and had negative correlation with miR-199a expression in proliferating-phase HAs. Overexpression of HIF1A attenuated the anti-proliferation effect of miR-199a mimic in HAs cells. Taken together, our findings demonstrate that miR-199a may inhibit proliferation and induce apoptosis in HAs cells via targeting HIF1A and provide a potential therapeutic target for HAs.

Effect of TALEN-mediated IL-6 knockout on cell proliferation, apoptosis, invasion and anti-cancer therapy in hepatocellular carcinoma (HCC-LM3) cells.

PURPOSE: To determine the exact effect of Interleukin-6 (IL-6) on tumor cell proliferation, apoptosis, invasion, and anti-cancer therapy in hepatocellular carcinoma (HCC). EXPERIMENTAL DESIGN: IL-6 was disrupted by transcription activator-like effector nucleases (TALEN) in HCCLM3 cells, and was used to evaluate the role of IL-6 on tumor cell proliferation, apoptosis, invasion and key signaling pathways involved in sorafenib and/or IFNα therapy. RESULTS: IL-6 has no direct effect on cell proliferation and invasion but promotes cell apoptosis and up-regulate IL-33 and VEGF-A expression. IL-6 could attenuate the anti-proliferation effect by sorafenib and combination therapy but facilitate the pro-apoptosis of the combination therapy and augment the pro-invasive effect induced by single treatment. IL-6 could down-regulate p-STAT3, however up-regulate the p-MEK/p-ERK and NF-kB/iNOS expression, and it also facilitated the promotion on p-JAK2 and p-MEK/p-ERK by either sorafenib or IFN-α. in vivo study, IL-6 significantly promotes tumor growth. The combination treatment showed the highest inhibition on tumor growth which is derived from HCCLM3-IL6(-) cells. CONCLUSIONS: IL-6 has no direct effect on cell proliferation and invasion but promotes tumor cell apoptosis in vitro study. Sorafenib and combination therapies are suitable for HCC cells with low or no IL-6 expression confirmed in vivo study.

Tim-3 inhibits low-density lipoprotein-induced atherogenic responses in human umbilical vein endothelial cells.

Endothelial injury and dysfunction followed by endothelial activation and inflammatory cell recruitment are factors contributing to the initiation and progression of atherosclerosis. Oxidized low-density lipoprotein (ox-LDL) promotes inflammation during atherogenesis and lipid deposition in the arterial wall. We observed that stimulation of human umbilical vein endothelial cells (HUVECs) with ox-LDL activated pro-inflammatory cytokine production and apoptosis, inhibited cell migration, and upregulated T-cell immunoglobulin and mucin domain 3 (Tim-3) expression. Tim-3, in turn, protected HUVECs from ox-LDL-induced apoptosis via the JNK pathway and reversed the inhibition of migration. Tim-3 also inhibited ox-LDL-induced inflammatory cytokine production by suppressing NF-κB activation. In addition, Tim-3 increased production of type 2 T helper cells (Th2) and regulatory T cell (Treg)-associated cytokines. Blocking Tim-3 reversed its effects on the inflammatory response to ox-LDL. Thus, Tim-3 signaling may be a "self-control" mechanism in ox-LDL-triggered inflammation in HUVECs. These results identify Tim-3 as a factor in HUVEC activity and suggest its potential in the treatment of atherosclerosis.

Helicobacter pylori and 17ß-estradiol induce human intrahepatic biliary epithelial cell abnormal proliferation and oxidative DNA damage.

BACKGROUND: Biliary cancers are more common in females, and previous studies have suggested that Helicobacter pylori (H. pylori) exists in the biliary system. However, the effects of H. pylori infection and estrogen on the biological behaviors of human biliary epithelium mucosa remain unknown. The present study aimed to clarify their effects on the proliferation, apoptosis, migration and oxidative DNA damage of a human intrahepatic biliary epithelial cell (HIBEC) line in vitro. METHODS: HIBECs were co-cultured with 17ß-estradiol (at 10-9 mol/L, 10-7 mol/L, and 10-5 mol/L) and H. pylori (at MOI=0.5:1, 1:1, and 2:1) and continuously passaged until the 15th generation (approximately 45 days). Then, the following assays were performed. HIBEC proliferation was measured using the CCK-8 assay, plate clone-formation assay and by determining Ki-67 expression with immunocytochemistry; cell apoptosis and migration were investigated using Annexin-V/PI and transwell assays, respectively; and reactive oxygen species (ROS) and 8-hydroxy-2'-deoxyguanosine (8-OHdG) production were detected by flow cytometry and immunofluorescence staining combined with confocal laser scanning microscopy, respectively. The results were the basis for evaluating the level of oxidative stress and the related DNA damage in HIBECs. RESULTS: HIBECs maintained a normal morphology and vitality when treated with 17ß-estradiol (at 10-9 mol/L) and H. pylori (at MOI=0.5:1 and 1:1). 17ß-estradiol at 10-7 mol/L and 10-5 mol/L and H. pylori at MOI=2:1, by contrast, caused cell death. Compared with controls, HIBECs treated with 17ß-estradiol (10-9 mol/L) and H. pylori (MOI=1:1) had a higher up-regulation of proliferation, Ki-67 expression, clone formation, migration activity and the expression of ROS and 8-OHdG and exhibited a down-regulation of apoptosis. The above effects were further increased when 17ß-estradiol and H. pylori were combined (P<0.05). CONCLUSIONS: H. pylori and 17ß-estradiol, separately or in combination, promoted cell proliferation and suppressed apoptosis of HIBECs in vitro. The above phenomena might be related to oxidative stress and its subsequent DNA damage with H. pylori and 17ß-estradiol.

Long noncoding RNA UCA1 induced by SP1 promotes cell proliferation via recruiting EZH2 and activating AKT pathway in gastric cancer.

Long noncoding RNA UCA1 has emerged as a novel regulator in cancer initiation and progression of various cancers. However, function and underlying mechanism of UCA1 in the progression of gastric cancer (GC) remain unclear. In the present study, we report that UCA1 expressed highly in GC tissues and GC cells, which was partly induced by SP1. UCA1 promoted GC cell proliferation and G1/S transition in vitro and in vivo. Moreover, UCA1 exerted its function through interacting with EZH2, promoting direct interaction with cyclin D1 promoter to activate the translation of cyclin D1. Furthermore, AKT/GSK-3B/cyclin D1 axis was activated to upregulate cyclin D1 due to overexpression of UCA1. In addition, EZH2 and phosphorylated AKT induced by UCA1 could impact each other to form a positive feedback to promote cyclin D1 expression. This study demonstrated that UCA1 as a critical regulator involved in GC proliferation and cell cycle progression by promoting cyclin D1 expression, which indicates that it may be clinically a potential therapeutic target in GC.

ROCK inhibition as a potential therapeutic target involved in apoptosis in hemangioma.

Gene expression was examined in hemangiomas (HA), benign, birthmark-like tumors occurring in infancy, and confirmed in HA-derived endothelial cells (HDEC), for which cell proliferation and apoptosis were also assessed. Protein and mRNA accumulation of Rho-associated protein kinase (ROCK), vascular endothelial growth factor (VEGF), Ki-67 and proliferating cell nuclear antigen was significantly higher in proliferating phase HAs than in involuting phase HAs. In contrast, p53 and caspase-3 exhibited higher levels of accumulation in involuting than proliferating HAs. Cell apoptotic indexes were low in proliferating phase HAs and increased in involuting phase HAs. HDECs were treated with the ROCK inhibitor Y-27632. Y-27632 induced p53 expression and downregulated VEGF expression, significantly inhibited cell proliferation, and induced cell apoptosis in HA cells. The inhibitor effects were confirmed in HAs from HDEC-injected nude mice. These results indicated that ROCK is involved in p53-mediated apoptosis and VEGF expression in HA cells and suggested that such inhibition may be exploited for future HA therapies.

Long non-coding RNA TUG1 promotes cell proliferation and metastasis by negatively regulating miR-300 in gallbladder carcinoma.

BACKGROUND: As we all know, long non-coding RNAs (lncRNAs) have been reported to play vital roles in various human cancers. In this study, we aimed to explore the role of lncRNA TUG1 in gallbladder carcinoma (GBC) development. METHODS: Total RNA was extracted from the tissues of thirty GBC patients, four GBC cell lines. We detected the expression levels of TUG1 using quantitative real-time PCR. We performed CCK8, colony formation, transwell invasion and apoptosis assays to study the effects of TUG1 on GBC cell proliferation and invasion. Western blot assay was performed to assess to the expression level of epithelial-mesenchymal transition (EMT) markers in transforming growth factor-ß1 (TGF-ß1) treated and TUG1 knockdown GBC cell. Lastly, dual-luciferase reporter assay and quantitative real-time PCR were performed to verify the potential target microRNAs (miRNAs) of TUG1. RESULTS: TUG1 expression was significantly overexpressed in GBC tissues. Functionally, this study demonstrated that knockdown of TUG1 significantly inhibited GBC cell proliferation, metastasis. Mechanically, we found that TUG1 is upregulated by TGF-ß1, and knockdown of TUG1 inhibited GBC cell EMT. Furthermore, we identified that miR-300, which has been reported as a suppressor in other types of cancer, is negatively regulated by TUG1. CONCLUSIONS: LncRNA TUG1 promotes GBC cell proliferation, metastasis and EMT progression by functioning as a miRNA sponge to abrogate the endogenous effect of miR-300.

Upregulation of long non-coding RNA LINC00152 by SP1 contributes to gallbladder cancer cell growth and tumor metastasis via PI3K/AKT pathway.

Gallbladder cancer (GBC) is one of the most lethal cancers with poor prognosis. In this study, we report that the long non-coding RNA LINC00152 is significantly upregulated in GBC tissues and cell lines. The high LINC00152 levels correlated positively with tumor status progression, lymph node invasion and TNM stage advancement. Functionally, we revealed that LINC00152 dramatically promoted cell proliferation, metastasis and inhibited apoptosis in vitro. In vivo, LINC00152 overexpression significantly promoted tumor growth. Mechanistic analyses indicated that LINC00152 could participate in the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway, and transcription factor specificity protein 1 (SP1) induces its overexpression. In summary, our findings suggest that LINC00152 contributes to the oncogenic potential of GBC and SP1/LINC00152/PI3K/AKT may be a potential therapeutic target for GBC.

Long non-coding RNA H19 regulates FOXM1 expression by competitively binding endogenous miR-342-3p in gallbladder cancer.

BACKGROUND: Long non-coding RNA (lncRNA) H19 has been reported to involve in many kinds of human cancers and functions as an oncogene. Our previous study found that H19 was over-expressed in gallbladder cancer (GBC) and was shown to promote tumor development in GBC. However, the competing endogenous RNA (ceRNA) regulatory network involving H19 in GBC progression has not been fully elucidated. We aim to detect the role of H19 as a ceRNA in GBC. METHODS AND RESULTS: In this study, the expression of H19 and miR-342-3p were analyzed in 35 GBC tissues and matched normal tissues by using quantitative polymerase chain reaction (qRT-PCR). We demonstrated H19 was overexpressed and negatively correlated with miR-342-3p in GBC. By dual-luciferase reporter assays, RNA-binding protein immunoprecipitation (RIP) and RNA pull-down assays, we verified that H19 was identified as a direct target of miR-342-3p. QRT-PCR and Western-blotting assays demonstrated that H19 silencing down-regulated, whereas over-expression enhanced the expression of miR-342-3p targeting FOXM1 through competitively 'sponging' miR-342-3p. Furthermore, transwell invasion assays and cell cycle assays indicated that H19 knockdown inhibited both cells invasion and proliferation, but this effects was attenuated by co-transfection of siRNA-H19 and miR-342-3p inhibitor in GBC cells. In vivo, tumor volumes were decreased significantly in H19 silenced group compared to the control group, but was attenuated by co-transfection of shRNA-H19 and miR-342-3p inhibitor, which were stablely constructed through lenti-virus vector. CONCLUSION: Our results suggest a potential ceRNA regulatory network involving H19 regulates FOXM1 expression by competitively binding endogenous miR-342-3p in GBC. This mechanism may contribute to a better understanding of GBC pathogenesis and provides potential therapeutic strategy for GBC.

Long Noncoding RNA GCASPC, a Target of miR-17-3p, Negatively Regulates Pyruvate Carboxylase-Dependent Cell Proliferation in Gallbladder Cancer.

Long noncoding RNAs (lncRNA) are being implicated in the development of many cancers. Here, we report the discovery of a critical role for the lncRNA GCASPC in determining the progression of gallbladder cancer. Differentially expressed lncRNAs and mRNAs between gallbladder cancer specimens and paired adjacent nontumor tissues from five patients were identified and validated by an expression microarray analysis. Quantitative real-time PCR was used to measure GCASPC levels in tissues from 42 gallbladder cancer patients, and levels of GCASPC were confirmed further in a separate cohort of 89 gallbladder cancer patients. GCASPC was overexpressed or silenced in several gallbladder cancer cell lines where molecular and biological analyses were performed. GCASPC levels were significantly lower in gallbladder cancer than adjacent nontumor tissues and were associated with tumor size, American Joint Committee on Cancer tumor stage, and patient outcomes. GCASPC overexpression suppressed cell proliferation in vitro and in vivo, whereas GCASPC silencing had opposite effects. By RNA pull-down and mass spectrometry, we identified pyruvate carboxylase as an RNA-binding protein that associated with GCASPC. Because GCASPC is a target of miR-17-3p, we confirmed that both miR-17-3p and GCASPC downregulated pyruvate carboxylase level and activity by limiting protein stability. Taken together, our results defined a novel mechanism of lncRNA-regulated cell proliferation in gallbladder cancer, illuminating a new basis for understanding its pathogenicity. Cancer Res; 76(18); 5361-71. ©2016 AACR.

Overexpression of LncRNA-ROR predicts a poor outcome in gallbladder cancer patients and promotes the tumor cells proliferation, migration, and invasion.

LncRNA-ROR has been reported to be involved in many kinds of human cancers. However, whether LncRNA-ROR is involved in gallbladder cancer progression remains largely unknown. The objective of this study is to investigate the role of LncRNA-ROR in gallbladder cancer. We found that LncRNA-ROR expression level was upregulated in gallbladder cancer tissues (P < 0.05) and was significantly associated with tumor sizes (P < 0.05) and lymph node metastasis (P < 0.05). High expression of LncRNA-ROR was significantly associated with poor prognosis in gallbladder cancer patients (P < 0.05). Moreover, knockdown of LncRNA-ROR inhibited cell proliferation, migration, and invasion. The epithelial-mesenchymal transition (EMT) phenotype induced by TGF-ß1 was reversed after LncRNA-ROR knocking down in SGC-996 and Noz cells. LncRNA-ROR plays an important role in the development of gallbladder cancer and mediates the EMT in gallbladder cancer. LncRNA-ROR might act as a marker of prognosis and therapeutic target for gallbladder cancer.

The lncRNA MALAT1 functions as a competing endogenous RNA to regulate MCL-1 expression by sponging miR-363-3p in gallbladder cancer.

Gallbladder carcinoma (GBC) is an aggressive neoplasm, and the treatment options for advanced GBC are limited. Recently, long non-coding RNAs (lncRNAs) have emerged as new gene regulators and prognostic markers in several cancers. In this study, we found that metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) expression was up-regulated in GBC tissues (P < 0.05). Luciferase reporter assays and RNA pull down assays showed that MALAT1 is a target of miR-363-3p. Real-time quantitative PCR and Western blot analysis indicated that MALAT1 regulated Myeloid cell leukaemia-1 (MCL-1) expression as a competing endogenous RNA (ceRNA) for miR-363-3p in GBC cells. Furthermore, MALAT1 silencing decreased GBC cell proliferation and the S phase cell population and induced apoptosis in vitro. In vivo, tumour volumes were significantly decreased in the MALAT1 silencing group compared with those in the control group. These data demonstrated that the MALAT1/miR-363-3p/MCL-1 regulatory pathway controls the progression of GBC. Inhibition of MALAT1 expression may be to a novel therapeutic strategy for gallbladder cancer.

Downregulation of Notch-regulated Ankyrin Repeat Protein Exerts Antitumor Activities against Growth of Thyroid Cancer.

BACKGROUND: The Notch-regulated ankyrin repeat protein (NRARP) is recently found to promote proliferation of breast cancer cells. The role of NRARP in carcinogenesis deserves extensive investigations. This study attempted to investigate the expression of NRARP in thyroid cancer tissues and assess the influence of NRARP on cell proliferation, apoptosis, cell cycle, and invasion in thyroid cancer. METHODS: Thirty-four cases with thyroid cancer were collected from the Department of General Surgery, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine between 2011 and 2012. Immunohistochemistry was used to detect the level of NRARP in cancer tissues. Lentivirus carrying NRARP-shRNA (Lenti-NRARP-shRNA) was applied to down-regulate NRARP expression. Cell viability was tested after treatment with Lenti-NRARP-shRNA using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptosis and cell cycle distribution were determined by flow cytometry. Cell invasion was tested using Transwell invasion assay. In addition, expressions of several cell cycle-associated and apoptosis-associated proteins were examined using Western blotting after transfection. Student's t-test, one-way analysis of variance (ANOVA), or Kaplan-Meier were used to analyze the differences between two group or three groups. RESULTS: NRARP was highly expressed in thyroid cancer tissues. Lenti-NRARP-shRNA showed significantly inhibitory activities against cell growth at a multiplicity of infection of 10 or higher (P < 0.05). Lenti-NRARP-shRNA-induced G1 arrest (BHT101: 72.57% ± 5.32%; 8305C: 75.45% ± 5.26%) by promoting p21 expression, induced apoptosis by promoting bax expression and suppressing bcl-2 expression, and inhibited cell invasion by suppressing matrix metalloproteinase-9 expression. CONCLUSION: Downregulation of NRARP expression exerts significant antitumor activities against cell growth and invasion of thyroid cancer, that suggests a potential role of NRARP in thyroid cancer targeted therapy.

Long non-coding RNA MINCR promotes gallbladder cancer progression through stimulating EZH2 expression.

The regulation of MYC-regulated long non-coding RNAs has been reported to contribute to certain types of cancers. However, the role of MYC-induced long non-coding RNA (MINCR) in the tumorigenesis of gallbladder cancer (GBC) is still largely unknown. In this study, we discovered that MINCR was markedly upregulated in GBC tissues compared with adjacent normal tissues. High MINCR expression levels in GBC were positively associated with tumor volume and lymph node metastasis and were negatively correlated with overall survival (OS). Upregulation of MINCR and enhancer of zeste homolog 2 (EZH2) in GBC coincided with the downregulation of miR-26a-5p in GBC. Mechanistically, MINCR/miR-26a-5p/EZH2 axis was found to be involved in cell proliferation, cell invasive and apoptosis in GBC cells. Moreover, knockdown of MINCR suppressed cell proliferation, decreased S-phase cell numbers, increased cell apoptosis, and inhibited cell invasion by inhibiting the epithelial-mesenchymal transition (EMT) phenomenon in GBC cells. In vivo, tumor volumes were significantly decreased in the MINCR silencing group compared with those in the control group. These results demonstrated that MINCR could potentially be a therapeutic target as well as a prognostic marker in GBC.

Erratum: Upregulation of H19 indicates a poor prognosis in gallbladder carcinoma and promotes epithelial-mesenchymal transition.

[This corrects the article on p. 15 in vol. 6, PMID: 27073719.].

Long non-coding RNA Malat1 promotes gallbladder cancer development by acting as a molecular sponge to regulate miR-206.

Long non-coding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (Malat1) functions as an oncogene in many types of human cancer. In this study, we show that Malat1 is overexpressed in gallbladder cancer (GBC) tissue and cells. The high Malat1 levels correlated positively with tumor size and lymphatic metastasis, and correlated negatively with overall survival. We also show that Malat1 functions as a competing endogenous RNA (ceRNA) for miR-206. Because miR-206 directly suppresses expression of ANXA2 and KRAS, which are thought to promote GBC progression, Malat1 binding of miR-206 in GBC tissue and cells has an oncogenic effect. Conversely, Malat1 knockdown inhibits proliferation and invasion by GBC cells while increasing apoptosis. In vivo, silencing Malat1 decreases tumor volume. These results suggest Malat1 could potentially serve as a therapeutic target and prognostic marker for GBC.

Upregulation of H19 indicates a poor prognosis in gallbladder carcinoma and promotes epithelial-mesenchymal transition.

The imprinted oncofetal long non-coding RNA H19 has been reported to be involved in many kinds of human cancers. However, whether lncRNA H19 implicate in oncogenesis and cancer progression in gallbladder cancer remain largely unknown. In the present study, compared with adjacent normal tissues, the level of H19 was significantly upregulated in gallbladder cancer tissues and was positively associated with lymphatic metastasis and tumor size. The overall survival is shorter in those who had higher H19 expression among GBC patients. In vitro, both TGF-ß1 and IL-6 treatment induced upregulation of H19, downregulated the protein level of E-cadherin while increased Vimentin, indicating an epithelial-mesenchymal transition (EMT) phenotype in GBC. The overexpression of H19 in GBC cells enhanced tumor invasion and promoted EMT by upregulated transcription factor Twist1. On the contrary, Loss of function studies indicated that H19 interference in GBC suppressed tumor cell invasion and promoted mesenchymal-epithelial transition (MET) via suppressing Twist expression. In vivo, the volume of the tumors in H19-inteference group was significantly decreased compared to those in the control group of nude mice. Both western-blot and immunohistochemistry confirmed that a MET phenotype existed in the H19 interference group when compared to control group. These results defined H19 as a novel prognostic factor for GBC, and indicated that it might play important regulatory roles in the EMT process.

Individualized nomogram improves diagnostic accuracy of stage I-II gallbladder cancer in chronic cholecystitis patients with gallbladder wall thickening.

BACKGROUND: Early diagnosis of gallbladder cancer (GBC) can remarkably improve the prognosis of patients. This study aimed to develop a nomogram for individualized diagnosis of stage I-II GBC in chronic cholecystitis patients with gallbladder wall thickening. METHODS: The nomogram was developed using logistic regression analyses based on a retrospective cohort consisting of 89 consecutive patients with stage I-II GBC and 1240 patients with gallbladder wall thickening treated at one biliary surgery center in Shanghai between January 2009 and December 2011. The accuracy of the nomogram was validated by discrimination, calibration and a prospective cohort treated at another center between January 2012 and December 2014 (n=928). RESULTS: Factors included in the nomogram were advanced age, hazardous alcohol consumption, long-standing diagnosed gallstones, atrophic gallbladder, gallbladder wall calcification, intraluminal polypoid lesion, higher wall thickness ratio and mucosal line disruption. The nomogram had concordance indices of 0.889 and 0.856 for the two cohorts, respectively. Internal and external calibration curves fitted well. The area under the receiver-operating characteristic curves of the nomogram was higher than that of multidetector row computed tomography in diagnosis of stage I-II GBC (P<0.001). CONCLUSION: The proposed nomogram improves individualized diagnosis of stage I-II GBC in chronic cholecystitis patients with gallbladder wall thickening, especially for those the imaging features alone do not allow to confirm the diagnosis.

Long noncoding RNA H19 contributes to gallbladder cancer cell proliferation by modulated miR-194-5p targeting AKT2.

Gallbladder cancer (GBC) is a highly malignant cancer with poor prognosis. Although long noncoding RNA (lncRNA) H19 has been reported to play vital role in many human cancers, whether it is involved in GBC proliferation is still unknown. This study was designed to explore the effect of H19 in GBC cell proliferation. The expression of H19 and AKT2 were significantly elevated in GBC tissues, and the level of miR-194-5p is markedly decreased. Moreover, the RNA levels of H19 and AKT2 were positively correlated, and H19 elevation was significantly associated with tumor size. Cell proliferation decreased significantly after knockdown of H19 in GBC-SD and NOZ cells and after knockdown of AKT2 in NOZ cells. Results from cell cycle studies indicated that the S phase were significantly decreased after knockdown of H19 in NOZ cells but significantly elevated after overexpression of H19 in GBC-SD cells. Furthermore, knockdown of H19 upregulated miR-194-5p levels, yet significantly decreased miR-194-5p targeting AKT2 gene expression in NOZ cells. Inhibitor against miR-194-5p reversed these effects. In addition, overexpression of H19 in GBC-SD cells downregulated miR-194-5p and markedly increased AKT2 expression, and miR-194-5p mimic reversed these effects. Eventually, GBC cells were arrested in G0/G1-phase after H19 knockdown, inhibition of miR-194-5p markedly promoted cells into S-phase and co-transfection of siH19, and miR-194-5p inhibitor exerted mutually counter-regulated effects on cell cycle. These results suggested that H19/miR-194-5p/AKT2 axis regulatory network might modulate cell proliferation in GBC.