Xianghui Fu

miR-194 is a marker of hepatic epithelial cells and suppresses metastasis of liver cancer cells in mice†

MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression by interacting with the 3′ untranslated region (3′‐UTR) of multiple mRNAs. Recent studies have linked miRNAs to the development of cancer metastasis. In this study, we show that miR‐194 is specifically expressed in the human gastrointestinal tract and kidney. Moreover, miR‐194 is highly expressed in hepatic epithelial cells, but not in Kupffer cells or hepatic stellate cells, two types of mesenchymal cells in the liver. miR‐194 expression was decreased in hepatocytes cultured in vitro, which had undergone a dedifferentiation process. Furthermore, expression of miR‐194 was low in liver mesenchymal‐like cancer cell lines. The overexpression of miR‐194 in liver mesenchymal‐like cancer cells reduced the expression of the mesenchymal cell marker N‐cadherin and suppressed invasion and migration of the mesenchymal‐like cancer cells both in vitro and in vivo. We further demonstrated that miR‐194 targeted the 3′‐UTRs of several genes that were involved in epithelial‐mesenchymal transition and cancer metastasis. Conclusion: These results support a role of miR‐194, which is specifically expressed in liver parenchymal cells, in preventing liver cancer cell metastasis. (HEPATOLOGY 2010;.)

MicroRNA-26a regulates insulin sensitivity and metabolism of glucose and lipids

Type 2 diabetes (T2D) is characterized by insulin resistance and increased hepatic glucose production, yet the molecular mechanisms underlying these abnormalities are poorly understood. MicroRNAs (miRs) are a class of small, noncoding RNAs that have been implicated in the regulation of human diseases, including T2D. miR-26a is known to play a critical role in tumorigenesis; however, its function in cellular metabolism remains unknown. Here, we determined that miR-26a regulates insulin signaling and metabolism of glucose and lipids. Compared with lean individuals, overweight humans had decreased expression of miR-26a in the liver. Moreover, miR-26 was downregulated in 2 obese mouse models compared with control animals. Global or liver-specific overexpression of miR-26a in mice fed a high-fat diet improved insulin sensitivity, decreased hepatic glucose production, and decreased fatty acid synthesis, thereby preventing obesity-induced metabolic complications. Conversely, silencing of endogenous miR-26a in conventional diet-fed mice impaired insulin sensitivity, enhanced glucose production, and increased fatty acid synthesis. miR-26a targeted several key regulators of hepatic metabolism and insulin signaling. These findings reveal miR-26a as a regulator of liver metabolism and suggest miR-26a should be further explored as a potential target for the treatment of T2D.

MicroRNA-26a targets ten eleven translocation enzymes and is regulated during pancreatic cell differentiation.

Significance Ten eleven translocation (TET) enzymes are recently identified proteins that mediate DNA demethylation, but their regulation is still unknown. This paper indicates that a microRNA, miR-26a, is able to directly target TETs and then modulate 5-hydroxymethylcytosine levels. Furthermore, regulation of TETs by miR-26a is shown to promote pancreatic cell differentiation both in vitro and in vivo. These results highlight a link between a miRNA, DNA demethylation, and stem/progenitor cell differentiation. It also suggests a potential approach to increase the efficiency of generating pancreatic endocrine cells in vitro through modulating miR-26a. Ten eleven translocation (TET) enzymes (TET1/TET2/TET3) and thymine DNA glycosylase (TDG) play crucial roles in early embryonic and germ cell development by mediating DNA demethylation. However, the molecular mechanisms that regulate TETs/TDG expression and their role in cellular differentiation, including that of the pancreas, are not known. Here, we report that (i) TET1/2/3 and TDG can be direct targets of the microRNA miR-26a, (ii) murine TETs, especially TET2 and TDG, are down-regulated in islets during postnatal differentiation, whereas miR-26a is up-regulated, (iii) changes in 5-hydroxymethylcytosine accompany changes in TET mRNA levels, (iv) these changes in mRNA and 5-hydroxymethylcytosine are also seen in an in vitro differentiation system initiated with FACS-sorted adult ductal progenitor-like cells, and (v) overexpression of miR-26a in mice increases postnatal islet cell number in vivo and endocrine/acinar colonies in vitro. These results establish a previously unknown link between miRNAs and TET expression levels, and suggest a potential role for miR-26a and TET family proteins in pancreatic cell differentiation.

miR-26a enhances miRNA biogenesis by targeting Lin28B and Zcchc11 to suppress tumor growth and metastasis.

Human cancers often exhibit attenuated microRNA (miRNA) biogenesis and global underexpression of miRNAs; thus, targeting the miRNA biogenesis pathway represents a novel strategy for cancer therapy. Here, we report that miR-26a enhances miRNA biogenesis, which acts as a common mechanism partially accounting for miR-26a function in diverse cancers including melanoma, prostate and liver cancer. miR-26a was broadly reduced in multiple cancers, and overexpression of miR-26a significantly suppressed tumor growth and metastasis both in vitro and in vivo, including melanoma, prostate and liver cancers. Notably, miR-26a overexpression was accompanied by global upregulation of miRNAs, especially let-7, and let-7 expression was concordant with miR-26a expression in cancer cell lines, xenograft tumors and normal human tissues, underscoring their biological relevance. We showed that miR-26a directly targeted Lin28B and Zcchc11—two critical repressors of let-7 maturation. Furthermore, we have demonstrated that Zcchc11 promoted tumor growth and metastasis, and it was prominently overexpressed in human cancers. Our findings thus provide a novel mechanism by which a miRNA acts as a modulator of miRNA biogenesis. These results also define a role of the miR-26a and Zcchc11 in tumorigenesis and metastasis and have implications to develop new strategies for cancer therapy.

Neonatal activation of the nuclear receptor CAR results in epigenetic memory and permanent change of drug metabolism in mouse liver

Aberrant epigenetic alterations during development may result in long‐term epigenetic memory and have a permanent effect on the health of subjects. Constitutive androstane receptor (CAR) is a central regulator of drug/xenobiotic metabolism. Here, we report that transient neonatal activation of CAR results in epigenetic memory and a permanent change of liver drug metabolism. CAR activation by neonatal exposure to the CAR‐specific ligand 1,4‐bis[2‐(3,5‐dichloropyridyloxy)] benzene (TCPOBOP) led to persistently induced expression of the CAR target genes Cyp2B10 and Cyp2C37 throughout the life of exposed mice. These mice showed a permanent reduction in sensitivity to zoxazolamine treatment as adults. Compared with control groups, the induction of Cyp2B10 and Cyp2C37 in hepatocytes isolated from these mice was more sensitive to low concentrations of the CAR agonist TCPOBOP. Accordingly, neonatal activation of CAR led to a permanent increase of histone 3 lysine 4 mono‐, di‐, and trimethylation and decrease of H3K9 trimethylation within the Cyp2B10 locus. Transcriptional coactivator activating signal cointegrator‐2 and histone demethylase JMJD2d participated in this CAR‐dependent epigenetic switch. Conclusion: Neonatal activation of CAR results in epigenetic memory and a permanent change of liver drug metabolism. (HEPATOLOGY 2012)

Insufficient bile acid signaling impairs liver repair in CYP27−/− mice

BACKGROUND & AIMS Previous studies indicate that bile acids (BAs) promote normal liver regeneration and repair after injury. However, the impact of insufficient BA signaling, which is observed in patients with BA sequestrant medication or cerebrotendinous xanthomatosis (CTX) disease, on liver injury is still unknown. Our aim is to determine the outcomes of reduced BA levels upon liver injury. METHODS Seventy percent partial hepatectomy (PH) and carbon tetrachloride (CCl(4)) treatment were performed using CYP27(-/-) mice, a genetic animal model with low BA levels. The liver repair of CYP27(-/-) mice after the treatments was characterized by histological staining, chemical analysis, and quantitative real-time PCR. RESULTS CYP27(-/-) mice exhibited enhanced CCl(4)-induce liver injury, and defective liver regeneration and prolonged steatosis after 70% PH. Due to the insufficient BA signaling, farnesoid X receptor (FXR) activities were significantly reduced in CYP27(-/-) livers after 70% PH. Activation of FXR by either 0.2% cholic acid feeding or oral infusion of an FXR agonist greatly promoted liver regeneration in CYP27(-/-) mice. CONCLUSIONS Normal physiological levels of BAs are required for liver repair. Patients with BA sequestrant medications or CTX disease due to CYP27 gene mutations may have an increased risk of liver failure, and treatment with FXR ligands can promote liver regeneration of patients with low BA levels.

Bile Acid Receptors and Liver Cancer

Liver cancer, particularly hepatocellular carcinoma (HCC), is the third leading cause of cancer death in the world. Bile acids (BAs) are liver-produced amphipathic molecules that are required to facilitate the absorption of cholesterol, fat-soluble vitamins, and lipids in the intestine. However, BAs are also known to act as potential carcinogens and deregulation of BA homeostasis has been linked to HCC formation. Two key BA receptors, farnesoid X receptor (FXR) and G protein-coupled bile acid receptor 1 (TGR5), were recently identified, which provides great insights into BAs’ normal physiological functions as well as their carcinogenic effects. In this review, we focus on the potential links among BAs, two BA receptors, and HCC. FXR and TGR5 not only play key roles in regulating BA homeostasis but also are essential in suppressing BAs’ carcinogenic effects on liver cancer.

GPBAR1/TGR5 Mediates Bile Acid-Induced Cytokine Expression in Murine Kupffer Cells

GPBAR1/TGR5 is a novel plasma membrane-bound G protein–coupled bile acid (BA) receptor. BAs are known to induce the expression of inflammatory cytokines in the liver with unknown mechanism. Here we show that without other external stimuli, TGR5 activation alone induced the expression of interleukin 1β (IL-1β) and tumor necrosis factor-α (TNF-α) in murine macrophage cell line RAW264.7 or murine Kupffer cells. The TGR5-mediated increase of pro-inflammatory cytokine expression was suppressed by JNK inhibition. Moreover, the induced pro-inflammatory cytokine expression in mouse liver by 1% cholic acid (CA) diet was blunted in JNK−/− mice. TGR5 activation by its ligands enhanced the phosphorylation levels, DNA-binding and trans-activities of c-Jun and ATF2 transcription factors. Finally, the induced pro-inflammatory cytokine expression in Kupffer cells by TGR5 activation correlated with the suppression of Cholesterol 7α-hydroxylase (Cyp7a1) expression in murine hepatocytes. These results suggest that TGR5 mediates the BA-induced pro-inflammatory cytokine production in murine Kupffer cells through JNK-dependent pathway. This novel role of TGR5 may correlate to the suppression of Cyp7a1 expression in hepatocytes and contribute to the delicate BA feedback regulation.

Molecular mechanism of liver cancer.

Liver cancer, hepatocellular carcinoma (HCC) in particular, is one of the most deadly cancers worldwide, and the incidence of HCC is increasing rapidly in the United States and other developed countries. Epidemiological studies have identified major risk factors for HCC, including infection with hepatitis B and C virus (HBV and HCV), exposure to certain chemicals, high intake of alcohol, as well as metabolic diseases such as obesity and diabetes that are rapidly rising in the US. Although the etiologies for liver carcinogenesis are relatively well defined, the exact mechanism and pathways leading to cancer development are still unclear. Chronic liver injury, followed by inflammation and irregular liver regeneration has been suggested as an important step in hepatocarcinogenesis. Therefore, the identification of key factors that protect the liver from injury and inflammation could provide valuable insight into the development of HCC. In this review, we will summarize the recent findings in HCC studies, mainly focusing on the new molecular link among inflammation, liver repair and HCC.

MiR-26a enhances autophagy to protect against ethanol-induced acute liver injury.

Autophagy is a process for the turnover of intracellular organelles and molecules during stress responses. microRNAs (miRNAs) are small, non-coding endogenous RNAs that may regulate almost every cellular process. However, the roles of miRNAs in autophagy are still poorly understood. In this study, we show that miR-26a enhances autophagy in both culture cells and the mouse liver. Hepatic overexpression of miR-26a in mice alleviated ethanol-induced hepatic steatosis and liver injury. Overexpression of miR-26a increased the expression of the autophagy mediator Beclin-1, which is regulated by mitogen-activated protein kinases (MAPKs). We identified DUSP4 and DUSP5, two MAPKs inhibitors, as direct targets of miR-26a. We further demonstrated that miR-26a targeted the 3′-UTRs of several other negative regulators of autophagy. Our results thus identify a novel miRNA-mediated mechanism that enhances cytoprotective autophagy in the liver.Key messages• miR-26a enhances autophagy in liver cells.• Hepatic overexpression of miR-26a in mice alleviates ethanol-induced liver injury.• Overexpression of miR-26a increases the expression of autophagy mediator Beclin-1.• DUSP4 and DUSP5, two MAPKs inhibitors, were identified as direct targets of miR-26a.