Binghua Jiao

Down-regulation of miR-23b may contribute to activation of the TGF-β1/Smad3 signalling pathway during the termination stage of liver regeneration

MicroRNAs (miRNAs) are known to play important roles in liver regeneration, although the role of miRNAs associated with the termination of liver regeneration is not as well studied. Here we reported the down‐regulation of miR‐23b in the termination stage of liver regeneration in rats. In addition, Smad3 was identified as a target of miR‐23b during liver regeneration. Up‐regulation of miR‐23b promoted BRL‐3A cell proliferation and partially inhibited transforming growth factor (TGF)‐β1‐induced apoptosis. Furthermore, TGF‐β1 transcriptionally inhibited miR‐23b expression. We conclude that down‐regulation of miR‐23b may contribute to activation of the TGF‐β1/Smad3 signalling pathway during the termination stage of liver regeneration.

Mir-34a Is Upregulated during Liver Regeneration in Rats and Is Associated with the Suppression of Hepatocyte Proliferation

Background MicroRNAs are a class of small regulatory RNAs that modulate a variety of biological processes, including cellular differentiation, apoptosis, metabolism and proliferation. This study aims to explore the effect of miR-34a in hepatocyte proliferation and its potential role in liver regeneration termination. Methodology/Principal Finding MiR-34a was highly induced after partial hepatectomy. Overexpression of miR-34a in BRL-3A cells could significantly inhibit cell proliferation and down-regulate the expression of inhibin βB (INHBB) and Met. In BRL-3A cells, INHBB was identified as a direct target of miR-34a by luciferase reporter assay. More importantly, INHBB siRNA significantly repressed cell proliferation. A decrease of INHBB and Met was detected in regenerating liver. Conclusion/Significance MiR-34a expression was upregulated during the late phase of liver regeneration. MiR-34a-mediated regulation of INHBB and Met may collectively contribute to the suppression of hepatocyte proliferation.

Down-Regulation of MiR-127 Facilitates Hepatocyte Proliferation during Rat Liver Regeneration

Liver regeneration (LR) after partial hepatectomy (PH) involves the proliferation and apoptosis of hepatocytes, and microRNAs have been shown to post-transcriptionally regulate genes involved in the regulation of these processes. To explore the role of miR-127 during LR, the expression patterns of miR-127 and its related proteins were investigated. MiR-127 was introduced into a rat liver cell line to examine its effects on the potential target genes Bcl6 and Setd8, and functional studies were undertaken. We discovered that miR-127 was down-regulated and inversely correlated with the expression of Bcl6 and Setd8 at 24 hours after PH, a time at which hypermethylation of the promoter region of the miR-127 gene was detected. Furthermore, in BRL-3A rat liver cells, we observed that overexpression of miR-127 significantly suppressed cell growth and directly inhibited the expression of Bcl6 and Setd8. The results suggest that down-regulation of miR-127 may be due to the rapid methylation of its promoter during the first 24 h after PH, and this event facilitates hepatocyte proliferation by releasing Bcl6 and Setd8. These findings support a miRNA-mediated negative regulation pattern in LR and implicate an anti-proliferative role for miR-127 in liver cells.

miR-376a suppresses proliferation and induces apoptosis in hepatocellular carcinoma

MicroRNAs are known to be involved in the pathogenesis of hepatocellular carcinoma (HCC). This study aims to explore the potential biological function of miR‐376a, which was found to be inhibited after partial hepatectomy, in HCC. We discovered that miR‐376a was frequently down‐regulated in HCC cell lines and HCC tissues, while higher relative level of miR‐376a was significantly associated with high serum AFP level. Over‐expression of miR‐376a not only inhibited proliferation but induced apoptosis in Huh7 cells. Additionally, p85α (PIK3R1) was identified as a direct and functional target of miR‐376a in Huh7 cells. Moreover, we confirmed that p85α and miR‐376a were inversely correlated in HCC. These findings suggest that down‐regulation of miR‐376a may contribute to the development of HCC by targeting p85α.

MicroRNA-127 Post-Transcriptionally Downregulates Sept7 and Suppresses Cell Growth in Hepatocellular Carcinoma Cells

Background/Aims: Hepatocellular carcinoma is one of the most common cancers worldwide. It has been suggested that microRNAs, a class of small regulatory RNAs, are associated with tumorigenesis by targeting the mRNAs of hundreds of genes that modulate a variety of biological processes, including cellular differentiation, apoptosis, metabolism, and proliferation. Methods/Results: we analyzed the expression levels of mir-127 in 33 HCC and non-cancerous tissues using qRT-PCR. MiR-127 is downregulated in 69.7% of HCC tissues compared with adjacent normal tissues, but its expression level is not correlated with the TNM stage, AFP level, or age. In vitro, miR-127 can arrest Huh7 at the G2/M phase and inhibit Huh7 cell proliferation. In an in vivo xenograft model, the overexpression of miR-127 can inhibit Huh7 cell tumorigenicity. The luciferase reporter and western blot results confirm that miR-127 downregulates Sept7 expression by targeting its 3UTR. Furthermore, the knockdown of Sept7 has the same effect on cell proliferation as the overexpression of miR-127 in Huh7 cells. Conclusion: miR-127 plays a tumor-suppressor role and can serve as a potential diagnostic biomarker for HCC.

SD118-Xanthocillin X (1), a Novel Marine Agent Extracted from Penicillium commune, Induces Autophagy through the Inhibition of the MEK/ERK Pathway

A compound named SD118-xanthocillin X (1) (C18H12N2O2), isolated from Penicillium commune in a deep-sea sediment sample, has been shown to inhibit the growth of several cancer cell lines in vitro. In the present study, we employed a growth inhibition assay and apoptotic analysis to identify the biological effect and detailed mechanism of SD118-xanthocillin X (1) in human hepatocellular carcinoma (HepG2) cells. SD118-xanthocillin X (1) demonstrated a concentration-dependent inhibitory effect on the growth of HepG2 cells and caused slight cellular apoptosis and significantly induced autophagy. Autophagy was detected as early as 12 h by the conversion of microtubule-associated protein 1 light chain 3 (LC3-I) to LC3-II, following cleavage and lipid addition to LC3-I. The pharmacological autophagy inhibitor 3-methyladenine largely attenuates the growth inhibition and autophagic effect of SD118-xanthocillin X (1) in HepG2 cells. Our data also indicated that the autophagic effect of SD118-xanthocillin X (1) occurs via the down-regulation of the MEK/ERK signaling pathway and the up-regulated class III PI3K/Beclin 1 signaling pathway.

Expression of soluble, biologically active recombinant human tumstatin in Escherichia coli

Tumstatin, a 28-kDa C-terminal fragment of collagen IV, is a potent anti-angiogenic protein and inhibitor of tumour growth. Recombinant tumstatin was prepared from Escherichia coli deposited as insoluble, inactive inclusion bodies. In the present study, we produced soluble and biologically active recombinant human tumstatin in E. coli by the coding region of tumstatin being linked to the 3′-end of the maltose-binding protein (MBP) gene. The fusion protein was expressed as the soluble form after induction by isopropylthio-β-D-galactoside (IPTG). MBP-tumstatin was purified by amylose affinity chromatography. MBP can be removed by digestion with factor Xa. Expression could represent 20% of the total soluble protein in E. coli, allowing approximately 8.6 mg of highly purified protein to be obtained per litre of bacterial culture. The purified tumstatin specifically inhibited the proliferation of endothelial cells in a dose-dependent manner. Annexin V-FITC apoptotic assay showed that recombinant tumstatin induced significant increase of apoptotic endothelial cells after 20 h of exposure to 20 μg/ml tumstatin, and when tumstatin was incubated on the chicken embryo, chorioallantoic membrane at doses of 1–15 μg, there was a dramatic decrease in microvasculaturethe allantoids of chicken embryos neovascular vessel test in vivo demonstrated that tumstatin treatment at doses of 1–15 μg gives rise to dramatically decrease the number of neovascular vessel. Our study provides a feasible and convenient approach to produce soluble and biologically active tumstatin.

The noncoding RNA HOXD-AS1 is a critical regulator of the metastasis and apoptosis phenotype in human hepatocellular carcinoma.

BackgroundDespite accumulating evidence that long noncoding RNAs (lncRNAs) are associated with cancer development in multiple types of cancer, the biological roles of many lncRNAs in human hepatocellular carcinoma (HCC) metastasis have not been well characterized.MethodsA lncRNA+ mRNA human gene expression microarray analysis was used to identify differentially expressed lncRNAs in metastatic HCC tissues compared to non-metastatic tissue.ResultsWe observed remarkable overexpression of HOXD-AS1 in metastatic cancer tissues. In vitro and in vivo gain- or loss-of-function studies re-affirmed that HOXD-AS1 is able to facilitate cancer metastasis and inhibit apoptosis. Moreover, we identified that HOXD-AS1 upregulated the Rho GTPase activating protein 11A (ARHGAP11A) by competitively binding to microRNA-19a (miR19a), resulting in induced metastasis. Interestingly, the regulator of G-protein signaling 3 (RGS3), a potential inhibitor of the MEK-ERK1/2 signaling axis, was also found to be downregulated by ectopic HOXD-AS1 overexpression, leading to a remarkably reduced apoptotic effect.ConclusionsThe present investigation strongly indicates that HOXD-AS1 is an oncogenic lncRNA that promotes HCC metastasis and that its pro-metastatic phenotype can partially be attributed to the HOXD-AS1/miR19a/ARHGAP11A signaling axis.

Lipid metabolism and peroxisome proliferator-activated receptor signaling pathways participate in late-phase liver regeneration.

Liver regeneration (LR) is of great clinical significance in various liver-associated diseases. LR proceeds along a sequence of three distinct phases: priming/initiation, proliferation, and termination. Compared with the recognition of the first two phases, little is known about LR termination and structure/function reorganization. A combination of omics techniques, along with bioinformatics, may provide new insights into the molecular mechanism of the late-phase LR. Gene, protein, and metabolite profiles of the rat liver were determined by cDNA microarray, two-dimensional electrophoresis, and HPLC-MS analysis. Pathway enrichment analysis was performed to identify the pathways: 427 differentially expressed genes extracted from the microarray experiment revealed two expression patterns representing the early and late phase of LR. Functionally, the genes expressing at a higher level at the early phase than at the late phase were mainly involved in the response to stress, proliferation, and resistance to apoptosis, while those expressing at a lower level at the early phase than at the late phase were mainly engaged in lipid metabolism. Compared with the sham-operation control (SH) group, 5 proteins in the 70% partial hepatectomy (70%PHx) group were upregulated at the protein level, and 3 proteins were downregulated at 168 h after the 70%PHx. E-FABP, an upregulated fatty acid binding protein, was found to be involved in the peroxisome proliferator-activated receptor (PPAR) signaling pathway. The metabolomic data confirmed the enhancement of lipid metabolism by the detection of the intermediate and final metabolites. Weve concluded that increased lipid metabolism and activated PPAR signaling pathways play important roles in late-phase LR.

p53-insensitive PUMA down-regulation is essential in the early phase of liver regeneration after partial hepatectomy in mice

BACKGROUND & AIMSnLiver regeneration after partial hepatectomy involve proliferation and apoptosis of hepatocytes. PUMA, the well-known proapoptotic member of the Bcl-2 family, can respond to distinct stimuli. This study explores the role of PUMA and its relationship with other Bcl-2 family members in this process.nnnMETHODSnThe expression patterns of PUMA and its related proteins were investigated in livers after 70% hepatectomies. The contributions of PUMA to liver regeneration were assessed by manipulating its expression levels using adenovirus vectors. The differences in PUMA expression levels in human normal livers and hepatitis, as well as hepatoma tissues were characterised.nnnRESULTSnDuring the first 72h after hepatectomy, PUMA was highly down-regulated transcriptionally, while the levels of p53, Slug, Bax, and Bcl-X(L) proteins increased continuously. Highly induced expression of PUMA in the liver by Ad-PUMA caused lethal fulminant hepatitis 48h after treatment. Slightly induced expression was enough to impair liver regeneration, with an elevation of post-hepatectomy mortality, an increase of apoptosis, a decrease of proliferation, an up-regulation of Bax levels, an induction of inflammatory chemokines (KC and macrophage inflammatory protein-2), and an increase in the neutrophil infiltration relative to the control. In contrast to the results from the regenerating liver, PUMA expression showed an increased trend in human hepatitis and hepatoma tissues.nnnCONCLUSIONSnSharply p53-insensitive down-regulation of PUMA, coupled with Bcl-X(L) up-regulation, may play a cytoprotective role in liver regeneration after hepatectomy. Furthermore, the increased expression of PUMA in hepatitis and hepatoma may indicate misregulation of the apoptotic network in these diseases.