SAGE detects microRNA precursors

Background

MicroRNAs (miRNAs) have been shown to play important roles in regulating gene expression. Since miRNAs are often evolutionarily conserved and their precursors can be folded into stem-loop hairpins, many miRNAs have been predicted. Yet experimental confirmation is difficult since miRNA expression is often specific to particular tissues and developmental stages.

Results

Analysis of 29 human and 230 mouse longSAGE libraries revealed the expression of 22 known and 10 predicted mammalian miRNAs. Most were detected in embryonic tissues. Four SAGE tags detected in human embryonic stem cells specifically match a cluster of four human miRNAs (mir-302a, b, c&d) known to be expressed in embryonic stem cells. LongSAGE data also suggest the existence of a mouse homolog of human and rat mir-493.

Conclusion

The observation that some orphan longSAGE tags uniquely match miRNA precursors provides information about the expression of some known and predicted miRNAs.

     

Monoacylglycerol lipase promotes progression of hepatocellular carcinoma via NF-κB-mediated epithelial-mesenchymal transition

Background

Monoacylglycerol lipase (MAGL), a critical lipolytic enzyme, has emerged as a key regulator of tumor progression, yet its biological function and clinical significance in hepatocellular carcinoma (HCC) is still unknown.

Methods

In this study, we used a tissue microarray containing samples from 170 HCC patients to evaluate the expression of MAGL and its correlation with other clinicopathologic characteristics. In addition, we investigated the regulating effects of MAGL on various HCC lines. Finally, we identified the NF-κB signaling pathway participated in MAGL-mediated epithelial-mesenchymal transition (EMT) using HCC cell lines with different metastatic potentials.

Results

The expression of MAGL was significantly higher in HCC tumors than in matched peritumor tissues. Specifically, high MAGL expression was found in tumors with larger tumor size, microvascular invasion, poor differentiation, or advanced TNM stage. In addition, the clinical prognosis for the MAGL^high group was markedly poorer than that for the MAGL^low group in the 1-, 3-, and 5-year overall survival times and recurrence rates of HCC patients. MAGL expression was an independent prognostic factor for both survival and recurrence after curative resection. Furthermore, the upregulation of MAGL in HCC cells promoted cell growth and invasiveness abilities, and accompanied by EMT. In contrast, downregulation of MAGL obviously inhibited these characteristics. Moreover, further investigations verified that MAGL facilitates HCC progression via NF-κB-mediated EMT process.

Conclusions

Our findings demonstrate MAGL could promote HCC progression by the induction of EMT and suggest a potential therapeutic target, as well as a biomarker for prognosis, in patients with HCC.

     

Integrated analysis of microRNA-target interactions with clinical outcomes for cancers

Background

Clinical statement alone is not enough to predict the progression of disease. Instead, the gene expression profiles have been widely used to forecast clinical outcomes. Many genes related to survival have been identified, and recently miRNA expression signatures predicting patient survival have been also investigated for several cancers. However, miRNAs and their target genes associated with clinical outcomes have remained largely unexplored.

Methods

Here, we demonstrate a survival analysis based on the regulatory relationships of miRNAs and their target genes. The patient survivals for the two major cancers, ovarian cancer and glioblastoma multiforme (GBM), are investigated through the integrated analysis of miRNA-mRNA interaction pairs.

Results

We found that there is a larger survival difference between two patient groups with an inversely correlated expression profile of miRNA and mRNA. It supports the idea that signatures of miRNAs and their targets related to cancer progression can be detected via this approach.

Conclusions

This integrated analysis can help to discover coordinated expression signatures of miRNAs and their target mRNAs that can be employed for therapeutics in human cancers.

     

TGF-β1-induced epithelial–mesenchymal transition in lung cancer cells involves upregulation of miR-9 and downregulation of its target,E-cadherin

Background

TGF-β1 plays an important role in the epithelial–mesenchymal transition (EMT) of epithelial cancers, including non-small cell lung cancer (NSCLC). While the full underlying mechanism remains unclear, miR-9 is known to play a critical role in the regulation of NSCLC cell invasion. We tested whether miR-9 targets E-cadherin and thus affects TGF-β1-induced EMT in NSCLC cells by assessing the expression levels of miR-9 and E-cadherin for NSCLC patients and then verifying the targeting of E-cadherin by miR-9 using the dual luciferase reporter system.

Results

MiR-9 was significantly upregulated in NSCLC tissues compared with its level in adjacent normal tissues. The expression of E-cadherin in NSCLC tissues was significantly decreased. In addition, we found that TGF-β1 significantly upregulated the expression of miR-9 and downregulated the expression of E-cadherin. E-cadherin was confirmed as a direct target gene of miR-9. Using an miR-9 inhibitor reversed the TGF-β1-mediated inhibition of E-cadherin expression and upregulation of the mesenchymal marker α-SMA. TGF-β1 significantly induced cell invasion, and this effect was significantly inhibited by miR-9 inhibitors.

Conclusions

TGF-β1 induced EMT in NSCLC cells by upregulating miR-9 and downregulating miR-9’s target, E-cadherin.

     

Stimulation of mouse vibrissal follicle growth by recombinant human fibroblast growth factor 20

Objectives

To explore potential effects of recombinant human fibroblast growth factor 20 (rhFGF20) in the growth of cultured mouse vibrissal follicles.

Results

The growth of cultured mouse vibrissal follicles was significantly induced by rhFGF20 in a dose dependent pattern in the in vitro vibrissal follicle organ culture model. However, too high concentration of rhFGF20 could inhibit the growth of vibrissal follicles. We further demonstrated that rhFGF20 stimulated the proliferation of hair matrix cells and activated Wnt/β-catenin signaling pathway.

Conclusions

The rhFGF20 might be a potential therapeutic agent to treat hair loss disorders.

     

miR-1260b promotes cell migration and invasion of hepatocellular carcinoma by targeting the regulator of G-protein signaling 22

Objectives

To investigate whether miR-1260b can regulate migration and invasion of hepatocellular carcinoma (HCC) by targeting RGS22.

Results

miR-1260b was up-regulated in HCC tissues compared with their corresponding non-cancerous tissues. Over-expression of miR-1260b increased migration and invasion of HepG2 and SMMC-7721 cells associated with HCC. Regulator of G-protein signaling 22 (RGS22) was identified as a directly target of miR-1260b and was inhibited by miR-1260b. Knockdown of RGS22 increased proliferation of HCC cells.

Conclusions

The new identified miR-1260b/RGS22 axis provides useful therapeutic methods for treatment of HCC deepening on our understanding of underlying mechanisms of HCC tumorigenesis.

     

Association of mitochondrial dysfunction and lipid metabolism with type 2 diabetes mellitus: A review of literature

Background

Diabetes mellitus (DM) is one of the most prevalent chronic diseases, and its prevalence continues to increase globally. The impact of mitochondrial dysfunction and lipid metabolism on diabetes mellitus and insulin resistance (IR) has been implicated in several previous reports; however, the results of studies are confusing despite four decades of study.

Methods/Results

This review has evaluated updated understanding of the role of mitochondrial dysfunction and lipid metabolism on type 2 diabetes, and found that mitochondrial dysfunction and lipid metabolism disorder induce the dysregulation of liver and pancreatic beta cells, insulin resistance, and type 2 diabetes.

Conclusion

Mitochondrial dysfunction and lipid metabolism induce metabolic dysregulation and finally increasing the possibility of diabetes.

     

Cell adhesion heterogeneity reinforces tumour cell dissemination: novel insights from a mathematical model

Background

Cancer cell invasion, dissemination, and metastasis have been linked to an epithelial-mesenchymal transition (EMT) of individual tumour cells. During EMT, adhesion molecules like E-cadherin are downregulated and the decrease of cell-cell adhesion allows tumour cells to dissociate from the primary tumour mass. This complex process depends on intracellular cues that are subject to genetic and epigenetic variability, as well as extrinsic cues from the local environment resulting in a spatial heterogeneity in the adhesive phenotype of individual tumour cells. Here, we use a novel mathematical model to study how adhesion heterogeneity, influenced by intrinsic and extrinsic factors, affects the dissemination of tumour cells from an epithelial cell population. The model is a multiscale cellular automaton that couples intracellular adhesion receptor regulation with cell-cell adhesion.

Results

Simulations of our mathematical model indicate profound effects of adhesion heterogeneity on tumour cell dissemination. In particular, we show that a large variation of intracellular adhesion receptor concentrations in a cell population reinforces cell dissemination, regardless of extrinsic cues mediated through the local cell density. However, additional control of adhesion receptor concentration through the local cell density, which can be assumed in healthy cells, weakens the effect. Furthermore, we provide evidence that adhesion heterogeneity can explain the remarkable differences in adhesion receptor concentrations of epithelial and mesenchymal phenotypes observed during EMT and might drive early dissemination of tumour cells.

Conclusions

Our results suggest that adhesion heterogeneity may be a universal trigger to reinforce cell dissemination in epithelial cell populations. This effect can be at least partially compensated by a control of adhesion receptor regulation through neighbouring cells. Accordingly, our findings explain how both an increase in intra-tumour adhesion heterogeneity and the loss of control through the local environment can promote tumour cell dissemination.

Reviewers

This article was reviewed by Hanspeter Herzel, Thomas Dandekar and Marek Kimmel.

     

Id-1 promotes migration and invasion of non-small cell lung cancer cells through activating NF-κB signaling pathway

Background

Numerous studies have shown that Id-1 (Inhibitor of differentiation 1) is upregulated in several cancers and associated with tumor malignant characters. However, the clinical significance and biological role of Id-1 in non-small cell lung cancer (NSCLC) remains unclear.

Methods

We used RT-PCR, Western blot and Immunohistochemistry to measure Id-1 expression in NSCLC tissues and matched adjacent noncancerous tissues. The expression pattern of Id-1 in NSCLC tissues was determined by scoring system of immunohistochemical analysis. The Kaplan-Meier method was used to calculate the survival curve, and log-rank test to determine statistical significance. The Id-1 gene was overexpressed or downreuglated with Lentiviral vectors in NSCLC cells. And, the migration ability of NSCLC cells was tested in a Transwell Boyden Chamber.

Results

We found that Id-1 is generally expressed higher in NSCLC tissues compared with matched adjacent noncancerous tissues. We also found that high Id-1 expression in tumor tissues is significantly correlated with tumor progression and poor survival in NSCLC patients. Furthermore, our experimental data revealed that knockdown of Id-1 significantly suppressed the proliferation, migration and invasion of NSCLC cells, whereas ectopic expression of Id-1 promoted the malignant phenotype of NSCLC cells. Mechanistic study showed that NF-κB signaling pathway contributed to the effects of Id-1 in NSCLC cells. Moreover, blocking the NF-κB pathway significantly inhibited the tumor-promoting actions of Id-1 in NSCLC cells.

Conclusions

We identified a tumorigenic role of Id-1 in NSCLC and provided a novel therapeutic target for NSCLC patients.

     

Optimization of fecal sample preparation for untargeted LC-HRMS based metabolomics

Introduction

Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.

Objectives

In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.

Methods

The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.

Results

A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.

Conclusion

The workflow generated repeatable and informative fingerprints for robust metabolome characterization.

     

Inhibition of integrin β3, a binding partner of kallistatin,leads to reduced viability,invasion and proliferation in NCI-H446 cells

Background

Kallistatin is a serine proteinase inhibitor and heparin-binding protein. It is considered an endogenous angiogenic inhibitor. In addition, multiple studies demonstrated that kallistatin directly inhibits cancer cell growth. However, the molecular mechanisms underlying these effects remain unclear.

Methods

Pull-down, immunoprecipitation, and immunoblotting were used for binding experiments. To elucidate the mechanisms, integrin β3 knockdown (siRNA) or blockage (antibody treatment) on the cell surface of small the cell lung cancer NCI-H446 cell line was used.

Results

Interestingly, kallistatin was capable of binding integrin β3 on the cell surface of NCI-H446 cells. Meanwhile, integrin β3 knockdown or blockage resulted in loss of antitumor activities induced by kallistatin. Furthermore, kallistatin suppressed tyrosine phosphorylation of integrin β3 and its downstream signaling pathways, including FAK/-Src, AKT and Erk/MAPK. Viability, proliferation and migration of NCI-H446 cells were inhibited by kallistatin, with Bcl-2 and Grb2 downregulation, and Bax, cleaved caspase-9 and caspase 3 upregulation.

Conclusions

These findings reveal a novel role for kallistatin in preventing small cell lung cancer growth and mobility, by direct interaction with integrin β3, leading to blockade of the related signaling pathway.

     

Generalized logical model based on network topology to capture the dynamical trends of cellular signaling pathways

Background

Cellular responses to extracellular perturbations require signaling pathways to capture and transmit the signals. However, the underlying molecular mechanisms of signal transduction are not yet fully understood, thus detailed and comprehensive models may not be available for all the signaling pathways. In particular, insufficient knowledge of parameters, which is a long-standing hindrance for quantitative kinetic modeling necessitates the use of parameter-free methods for modeling and simulation to capture dynamic properties of signaling pathways.

Results

We present a computational model that is able to simulate the graded responses to degradations, the sigmoidal biological relationships between signaling molecules and the effects of scheduled perturbations to the cells. The simulation results are validated using experimental data of protein phosphorylation, demonstrating that the proposed model is capable of capturing the main trend of protein activities during the process of signal transduction. Compared with existing simulators, our model has better performance on predicting the state transitions of signaling networks.

Conclusion

The proposed simulation tool provides a valuable resource for modeling cellular signaling pathways using a knowledge-based method.

     

Overexpression of EMMPRIN is associated with lymph node metastasis and advanced stage of non-small cell lung cancer: a retrospective study

Background

Previous studies show that overexpression of EMMPRIN involved in the malignant biological behavior of tumors. This investigation was to disclose the expression status of EMMPRIN in non-small cell lung cancer (NSCLC) and its clinical value for the diagnosis of NSCLC.

Methods

The expression of EMMPRIN was examined using immunohistochemistry and enzyme-linked immunosorbent assay. The clinical value of EMMPRIN was evaluated by drawing a receiver operating characteristic (ROC) curve.

Results

NSCLC tissues and serum exhibited higher expression levels of EMMPRIN than the normal control (p?<?0.05), and the expression of the EMMPRIN was significantly associated with lymphatic invasion and advanced stage of NSCLC (p?<?0.05). ROC curve suggested that the threshold level of serum EMMPRIN for distinguishing NSCLC from control group was 80.3 pg/mL, and displayed a sensitivity of 97.22% and a specificity of 95%. And higher EMMPRIN expression in serum and tissues appeared to be risk factors for NSCLC development (risk ratio =1.56 and 1.1).

Conclusion

Overexpression of EMMPRIN was associated with lymphatic metastasis and advanced stage of NSCLC and test of serum EMMPRIN contributes to the NSCLC diagnosis.

     

A novel trichosanthin fusion protein with increased cytotoxicity to tumor cells

Objective

To evaluate the anti-tumor effects of trichosanthin after fusion with a cell penetrating peptide, heparin-binding peptide (HBP), derived from human heparin-binding EGF-like growth factor (HB-EGF).

Results

The fusion protein of trichosanthin-HBP was expressed in Escherichia coli BL21 and purified by Ni–NTA affinity chromatography. The HBP domain had no influence on the topological inactivation activity and N-glycosidase activity of trichosanthin. Trichosanthin-HBP significantly inhibited the growth of tested cancer cells which are impervious to trichosanthin. Tumor cell apoptosis and both the mitochondrial- and death receptor-mediated apoptotic signaling pathways induced by trichosanthin-HBP were more significant than those induced by trichosanthin in HeLa cells.

Conclusion

HBP is an efficient intracellular delivery vehicle for trichosanthin and makes trichosanthin-HBP become a promising agent for cancer therapy.

     

MicroRNA-190b confers radio-sensitivity through negative regulation of Bcl-2 in gastric cancer cells

Objectives

To determine the role of miR-190b in radio-sensitivity of gastric cancer (GC).

Results

In radio-resistant GC cells, down-regulation of miR-190b and up-regulation of Bcl-2 were observed. The protein expression of Bcl-2 was negatively regulated by miR-190b. Overexpression of miR-190b significantly decreased cell viability and enhanced radio-sensitivity of GC cells. Of note, these effects of miR-190b on GC cells radio-sensitivity were abolished by Bcl-2.

Conclusion

miR-190b confers radio-sensitivity of GC cells, possibly via negative regulation of Bcl-2.

     

The miRNAome of ramie (<Emphasis Type="Italic">Boehmeria nivea</Emphasis> L.): identification,expression, and potential roles of novel microRNAs in regulation of cadmium stress response

Background

MicroRNAs (miRNAs) regulate numerous crucial abiotic stress processes in plants. However, information is limited on their involvement in cadmium (Cd) stress response and tolerance mechanisms in plants, including ramie (Boehmeria nivea L.) that produces a number of economic valuable as an important natural fibre crop and an ideal crop for Cd pollution remediation.

Results

Four small RNA libraries of Cd-stressed and non-stressed leaves and roots of ramie were constructed. Using small RNA-sequencing, 73 novel miRNAs were identified. Genome-wide expression analysis revealed that a set of miRNAs was differentially regulated in response to Cd stress. In silico target prediction identified 426 potential miRNA targets that include several uptake or transport factors for heavy metal ions. The reliability of small RNA sequencing and the relationship between the expression levels of miRNAs and their target genes were confirmed by quantitative PCR (q-PCR). We showed that the expression patterns of miRNAs obtained by q-PCR were consistent with those obtained from small RNA sequencing. Moreover, we demonstrated that the expression of six randomly selected target genes was inversely related to that of their corresponding miRNAs, indicating that the miRNAs regulate Cd stress response in ramie.

Conclusions

This study enriches the number of Cd-responsive miRNAs and lays a foundation for the elucidation of the miRNA-mediated regulatory mechanism in ramie during Cd stress.

     

Parthenolide induces apoptosis and autophagy through the suppression of PI3K/Akt signaling pathway in cervical cancer

Objective

To investigate the effect of parthenolide on apoptosis and autophagy and to study the role of the PI3K/Akt signaling pathway in cervical cancer.

Results

Parthenolide inhibits HeLa cell viability in a dose dependent-manner and was confirmed by MTT assay. Parthenolide (6 µM) induces mitochondrial-mediated apoptosis and autophagy by activation of caspase-3, upregulation of Bax, Beclin-1, ATG5, ATG3 and down-regulation of Bcl-2 and mTOR. Parthenolide also inhibits PI3K and Akt expression through activation of PTEN expression. Moreover, parthenolide induces generation of reactive oxygen species that leads to the loss of mitochondrial membrane potential.

Conclusion

Parthenolide induces apoptosis and autophagy-mediated growth inhibition in HeLa cells by suppressing the PI3K/Akt signaling pathway and mitochondrial membrane depolarization and ROS generation. Parthenolide may be a potential therapeutic agent for the treatment of cervical cancer.