Zinc alpha2 glycoprotein (ZAG) plays an important role in stimulating fat mobilization and lipolysis in adipose tissue, but its role in hepatic lipid metabolism remains unclear. Palmitic acid (PA) was used to stimulate HepG2 cells with ZAG overexpression or ZAG knock down (shRNA). Overexpression of ZAG significantly inhibited lipogenesis, promoted lipolysis and fatty acid β-oxidation, and attenuated PA-induced intracellular fat accumulation. Moreover, ZAG overexpression dramatically stimulated adiponectin expression in HepG2 cells. In contrast, knockdown of ZAG notably inhibited fatty acid β-oxidation, increased lipogenesis and lipid accumulation. Collectively, these data suggest that ZAG has the potential to alleviate hepatosteatosis, making it a promising therapeutic target for fatty liver.

Two ER membrane-resident transmembrane kinases, IRE1 and PERK, function as stress sensors in the unfolded protein response. IRE1 also has an endoribonuclease activity, which initiates a non-conventional mRNA splicing reaction, while PERK phosphorylates eIF2α. We engineered a potent small molecule, IPA, that binds to IRE1's ATP-binding pocket and predisposes the kinase domain to oligomerization, activating its RNase. IPA also inhibits PERK but, paradoxically, activates it at low concentrations, resulting in a bell-shaped activation profile. We reconstituted IPA-activation of PERK-mediated eIF2α phosphorylation from purified components. We estimate that under conditions of maximal activation less than 15% of PERK molecules in the reaction are occupied by IPA. We propose that IPA binding biases the PERK kinase towards its active conformation, which trans-activates apo-PERK molecules. The mechanism by which partial occupancy with an inhibitor can activate kinases may be wide-spread and carries major implications for design and therapeutic application of kinase inhibitors.

Nitrite-dependent anaerobic methane oxidation (n-damo) is performed by "Candidatus Methylomirabilis oxyfera" (M. oxyfera), which connects the carbon and nitrogen global nutrient cycles. In the present study, M. oxyfera-like bacteria sequences were successfully recovered from Yellow River Estuary sediments using specific primers for 16S rRNA and pmoA genes. A M. oxyfera-like sequences analysis based on the 16S rRNA gene revealed greater diversity compared with the pmoA gene; the 16S rRNA gene sequences retrieved from the Yellow River Estuary sediments belong to groups A as well as B and were mainly found in freshwater habitats. Quantitative PCR showed that 16S rRNA gene abundance varied from 9.28±0.11×10(3) to 2.10±0.13×10(5) copies g(-1) (dry weight), and the pmoA gene abundance ranged from 8.63±0.50×10(3) to 1.83±0.18×10(5) copies g(-1) (dry weight). A correlation analysis showed that the total organic carbon (TOC) and ammonium (NH4(+)) as well as the ratio of total phosphorus to total nitrogen (TP/TN) influenced the M. oxyfera-like bacteria distribution in the Yellow River Estuary sediments. These findings will aid in understanding the n-damo bacterial distribution pattern as well as their correlation with surrounding environmental factors in temperate estuarine ecosystems.

The kinase/endonuclease IRE1 is the most conserved signal transducer of the unfolded protein response (UPR), an intracellular signaling network that monitors and regulates the protein folding capacity of the endoplasmic reticulum (ER). Upon sensing protein folding perturbations in the ER, IRE1 initiates the unconventional splicing of XBP1 mRNA culminating in the production of the transcription factor XBP1s, which expands the ER's protein folding capacity. We show that an RNA-intrinsic conformational change causes the intron of XBP1 mRNA to be ejected and the exons to zipper up into an extended stem, juxtaposing the RNA ends for ligation. These conformational rearrangements are important for XBP1 mRNA splicing in vivo. The features that point to such active participation of XBP1 mRNA in the splicing reaction are highly conserved throughout metazoan evolution, supporting their importance in orchestrating XBP1 mRNA processing with efficiency and fidelity.

The transcriptional regulator CcpE is an important citrate-sensing regulator that modulates metabolic state, virulence factor expression, and bacterial virulence of Staphylococcus aureus (Ding et al., 2014 [1]). In this article, we report detailed methods for genome-wide transcriptional profiling of CcpE-regulated genes generated for the research article "Metabolic sensor governing bacterial virulence in Staphylococcus aureus" (Ding et al., 2014 [1]). All transcriptional profiling data was deposited to Gene Expression Omnibus (GEO) database under accession number GSE57260.

The contemporary use of nanomedicines for cancer treatment has been largely limited to serving as carriers for existing therapeutic agents. Here, we provide definitive evidence that, the metallofullerenol nanomaterial Gd@C82(OH)22, while essentially not toxic to normal mammary epithelial cells, possesses intrinsic inhibitory activity against triple-negative breast cancer cells. Gd@C82(OH)22 blocks epithelial-to-mesenchymal transition with resultant efficient elimination of breast cancer stem cells (CSCs) resulting in abrogation of tumour initiation and metastasis. In normoxic conditions, Gd@C82(OH)22 mediates these effects by blocking TGF-β signalling. Moreover, under hypoxic conditions found in the tumour microenvironment, cellular uptake of Gd@C82(OH)22 is facilitated where it functions as a bi-potent inhibitor of HIF-1α and TGF-β activities, enhancing CSC elimination. These studies indicate that nanomaterials can be engineered to directly target CSCs. Thus, Gd-metallofullerenol is identified as a kind of non-toxic CSC specific inhibitors with significant therapeutic potential.

Previously, we identified ISRIB as a potent inhibitor of the integrated stress response (ISR) and showed that ISRIB makes cells resistant to the effects of eIF2α phosphorylation and enhances long-term memory in rodents (Sidrauski et al., 2013). Here, we show by genome-wide in vivo ribosome profiling that translation of a restricted subset of mRNAs is induced upon ISR activation. ISRIB substantially reversed the translational effects elicited by phosphorylation of eIF2α and induced no major changes in translation or mRNA levels in unstressed cells. eIF2α phosphorylation-induced stress granule (SG) formation was blocked by ISRIB. Strikingly, ISRIB addition to stressed cells with pre-formed SGs induced their rapid disassembly, liberating mRNAs into the actively translating pool. Restoration of mRNA translation and modulation of SG dynamics may be an effective treatment of neurodegenerative diseases characterized by eIF2α phosphorylation, SG formation, and cognitive loss.

Transcatheter arterial embolization (TAE) plays an important role in clinical tumor therapy by accomplishing vessel-casting embolization of tumor arteries at all levels and suppressing tumor collateral circulation and vascular re-canalization. In this study, we describe smart blood-vessel-embolic nanogels for improving the anti-tumor efficacy of TAE therapy on hepatocellular carcinoma (HCC). Methods: In this study, an in vitro model composed of two microfluidic chips was used for simulating the tumor capillary network and analyzing artery-embolization properties. Also, blood-vessel-casting embolization of renal arteries was evaluated in normal rabbits. Using a VX2 tumor-bearing rabbit model, the therapeutic efficacy of TAE on HCC was investigated for tumor growth, necrosis, and proliferation. Neovascularization and collateral circulation were evaluated by immunofluorescent detection of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF), and CD31 following the TAE therapy of VX2 tumor-bearing rabbits. Results: Sufficient embolization of all eight levels of micro-channels was achieved in a tumor-vessel-mimetic model with two microfluidic chips using PIBI-2240, and was further confirmed in renal arteries of normal rabbit. Effective inhibition of tumor collateral circulation and vascular re-canalization was observed in VX2 tumor-bearing rabbits due to the reduced expression levels of HIF-1α, VEGF, and CD31. Conclusions: The exceptional anti-tumor effect of PIBI-2240 observed in this study suggested that it is an excellent blood-vessel-embolic material for tumor TAE therapy.

Organotypic slice culture is a living cell research technique which blends features of both in vivo and in vitro techniques. While organotypic brain slice culture techniques have been well established in rodents, there are few reports on the study of organotypic slice culture, especially of the central nervous system (CNS), in chicken embryos. We established a combined in ovo electroporation and organotypic slice culture method to study exogenous genes functions in the CNS during chicken embryo development. We performed in ovo electroporation in the spinal cord or optic tectum prior to slice culture. When embryonic development reached a specific stage, green fluorescent protein (GFP)-positive embryos were selected and fluorescent expression sites were cut under stereo fluorescence microscopy. Selected tissues were embedded in 4% agar. Tissues were sectioned on a vibratory microtome and 300 μm thick sections were mounted on a membrane of millicell cell culture insert. The insert was placed in a 30-mm culture dish and 1 ml of slice culture media was added. We show that during serum-free medium culture, the slice loses its original structure and propensity to be strictly regulated, which are the characteristics of the CNS. However, after adding serum, the histological structure of cultured-tissue slices was able to be well maintained and neuronal axons were significantly longer than that those of serum-free medium cultured-tissue slices. As the structure of a complete single neuron can be observed from a slice culture, this is a suitable way of studying single neuronal dynamics. As such, we present an effective method to study axon formation and migration of single neurons in vitro.

Ovarian cancer is the most lethal gynecological malignancy because of its poor prognosis. The Warburg effect is one of the key mechanisms mediating cancer progression. Molecules targeting the Warburg effect are therefore of significant therapeutic value for the treatment of cancers. Many microRNAs (miR) are dysregulated in cancers, and aberrant miR expression patterns have been suggested to correlate with the Warburg effect in cancer cells. In our study, we found that miR-145 negatively correlated with DNA methyltransferase (DNMT)3A expression at cellular/histological levels. miR-145 inhibited the Warburg effect by targeting HK2. Luciferase reporter assays confirmed that miR-145-mediated downregulation of DNMT3A occurred through direct targeting of its mRNA 3'-UTRs, whereas methylation-specific PCR (MSP) assays found that knockdown of DNMT3A increased mRNA level of miR-145 and decreased methylation levels of promoter regions in the miR-145 precursor gene, thus suggesting a crucial crosstalk between miR-145 and DNMT3A by a double-negative feedback loop. DNMT3A promoted the Warburg effect through miR-145. Coimmunoprecipitation assays confirmed no direct binding between DNMT3A and HK2. In conclusion, a feedback loop between miR-145 and DNMT3A is a potent signature for the Warburg effect in ovarian cancer, promising a potential target for improved anticancer treatment.

The aim of the current study was to explore the potential of TREK‑TRAAK two‑pore domain potassium (K2P) channels in protecting human retinal pigment epithelium (hRPE) cells against oxidative stress. hRPE cells were obtained from donors, and then cell identification and detection of the expression levels of TREK‑TRAAK K2P channels in hRPE cells were conducted. Subsequently, tert‑butyl hydroperoxide (t‑BH) was used to induce oxidative stress in hRPE cells. Docosahexaenoic acid (DHA) was used to stimulate and fluoxetine was used to inhibit the TREK‑TRAAK K2P channels. The survival rates of hRPE cells under oxidative stress were examined using flow cytometry. Apoptosis‑associated factors, including Bax, Bcl‑2, cleaved‑caspase‑3, αB‑crystallin and their mRNAs, were examined using immunofluorescence, western blot and reverse transcription‑polymerase chain reaction analyses. Variations in the cytoarchitecture were observed by immunofluorescence and electron microscopy. The cells examined in the present study were identified as hRPE cells. All members in the TREK‑TRAAK K2P channel family (including TREK‑1, TREK‑2 and TRAAK) were found to be expressed in hRPE cells. Stimulation of TREK‑TRAAK K2P channels increased the survival rates of hRPE cells under oxidative stress and the levels of intracellular protective factors, such as Bcl‑2 and αB‑crystallin. By contrast, inhibition of these channels decreased the cell survival rates and increased apoptosis enhancing factors, such as Bax and cleaved‑caspase‑3. Further examination of the cytoarchitecture revealed that TREK‑TRAAK K2P channels protected the integrity of the hRPE cell structure against oxidative stress. In conclusion, the present study suggested that the activated TREK‑TRAAK K2P channels serve a role in protecting hRPE cells against the oxidative stress induced by t‑BH, which indicated that these K2P channels are potential novel targets in retinal protection and provided a new direction for research and therapy in retinal degeneration diseases.

Ku70 and Ku80 form a heterodimer called Ku that forms a holoenzyme with DNA dependent-protein kinase catalytic subunit (DNA-PKCS) to repair DNA double strand breaks (DSBs) through the nonhomologous end joining (NHEJ) pathway. As expected mutating these genes in mice caused a similar DSB repair-defective phenotype. However, ku70(-/-) cells and ku80(-/-) cells also appeared to have a defect in base excision repair (BER). BER corrects base lesions, apurinic/apyrimidinic (AP) sites and single stand breaks (SSBs) utilizing a variety of proteins including glycosylases, AP endonuclease 1 (APE1) and DNA Polymerase β (Pol β). In addition, deleting Ku70 was not equivalent to deleting Ku80 in cells and mice. Therefore, we hypothesized that free Ku70 (not bound to Ku80) and/or free Ku80 (not bound to Ku70) possessed activity that influenced BER. To further test this hypothesis we performed two general sets of experiments. The first set showed that deleting either Ku70 or Ku80 caused an NHEJ-independent defect. We found ku80(-/-) mice had a shorter life span than dna-pkcs(-/-) mice demonstrating a phenotype that was greater than deleting the holoenzyme. We also found Ku70-deletion induced a p53 response that reduced the level of small mutations in the brain suggesting defective BER. We further confirmed that Ku80-deletion impaired BER via a mechanism that was not epistatic to Pol β. The second set of experiments showed that free Ku70 and free Ku80 could influence BER. We observed that deletion of either Ku70 or Ku80, but not both, increased sensitivity of cells to CRT0044876 (CRT), an agent that interferes with APE1. In addition, free Ku70 and free Ku80 bound to AP sites and in the case of Ku70 inhibited APE1 activity. These observations support a novel role for free Ku70 and free Ku80 in altering BER.

Based upon LX2421, a previously identified antiplatelet aggregation agent, a series of novel 2-amino-3-(naphth-2-yl)propanoic acid derivatives were designed, synthesized and evaluated. Among them, compounds LX14 and LX25 were identified as promising antiplatelet aggregation agents. The in vitro biologic study demonstrated that LX14 can block platelet aggregation induced by four different inducers and displays comparable potency in inhibiting GPIIb/IIIa receptor in comparison with Tirofiban. In addition, LX14 has much lower risk of bleeding than Tirofiban and shows significant antithrombotic activity in vivo. Taking together, the results indicated that LX14 is a promising GPIIb/IIIa receptor antagonist against platelet aggregation worthy of further evaluation.

There is an abnormal expression of nuclear receptor subfamily 2 group F member 6 (NR2F6) in human cancers such as breast cancer, colon cancer, and acute myelogenous leukemia. However, its clinical significance in cervical cancer has not been established. We explored NR2F6 expression and its clinicopathological significance in early-stage cervical cancer.

Phosphorylation of the α-subunit of initiation factor 2 (eIF2) controls protein synthesis by a conserved mechanism. In metazoa, distinct stress conditions activate different eIF2α kinases (PERK, PKR, GCN2, and HRI) that converge on phosphorylating a unique serine in eIF2α. This collection of signaling pathways is termed the 'integrated stress response' (ISR). eIF2α phosphorylation diminishes protein synthesis, while allowing preferential translation of some mRNAs. Starting with a cell-based screen for inhibitors of PERK signaling, we identified a small molecule, named ISRIB, that potently (IC50 = 5 nM) reverses the effects of eIF2α phosphorylation. ISRIB reduces the viability of cells subjected to PERK-activation by chronic endoplasmic reticulum stress. eIF2α phosphorylation is implicated in memory consolidation. Remarkably, ISRIB-treated mice display significant enhancement in spatial and fear-associated learning. Thus, memory consolidation is inherently limited by the ISR, and ISRIB releases this brake. As such, ISRIB promises to contribute to our understanding and treatment of cognitive disorders. DOI:http://dx.doi.org/10.7554/eLife.00498.001.

Ammonia oxidation is a critical process of estuarine nitrogen cycling involving ammonia-oxidizing archaea (AOA) and bacteria (AOB). However, the distribution patterns of ammonia-oxidizing microorganisms (AOMs) between different habitats in the same area remain unclear. The present study investigated the AOMs' abundance and community compositions in both sediment and water habitats of the Yellow River estuary. Quantitative PCR (qPCR) revealed that AOA showed significant higher abundance than AOB both in sediment and water samples. AOA and AOB abundance distribution trends were consistent in sediment but distinct in water along the sampling sites. Clone library-based analyses showed that AOA sequences were affiliated with Nitrososphaera, Nitrosopumilus and Nitrosotalea clusters. Generally, Nitrososphaera was predominant in sediment, while Nitrosopumilus and Nitrosotalea dominated in water column. AOB sequences were classified into genera Nitrosospira and Nitrosomonas, and Nitrosospira dominated in both habitats. Principal coordinate analysis (PCoA) also indicated AOA community structures exhibited significant differences between two habitats, while AOB were not. Ammonium and carbon contents were the potential key factors to influence AOMs' abundance and compositions in sediment, while no measured variables were determined to have major influences on communities in water habitat. These findings increase the understanding of the AOMs' distribution patterns in estuarine ecosystems.

The numerous increasing use of carbon nanotubes (CNTs) derived from nanotechnology has raised concerns about their biosafety and potential toxicity. CNTs cause immunologic dysfunction and limit the application of CNTs in biomedicine. The immunological responses induced by pristine multi-walled carbon nanotubes (p-MWCNTs) and PEGylated multi-walled carbon nanotubes (MWCNTs-PEG) on BALB/c mice via an intravenous administration were investigated. The results reflect that the p-MWCNTs induced significant increases in spleen, thymus, and lung weight. Mice treated with p-MWCNTs showed altered lymphocyte populations (CD3+, CD4+, CD8+, and CD19+) in peripheral blood and increased serum IgM and IgG levels, and splenic macrophage ultrastructure indicated mitochondria swelling. p-MWCNTs inhibited humoral and cellular immunity function and were associated with decreased immune responses against sheep erythrocytes and serum hemolysis level. Natural killer (NK) activity was not modified by two types of MWCNTs. In comparison with two types of MWCNTs, for a same dose, p-MWCNTs caused higher levels of inflammation and immunosuppression than MWCNTs-PEG. The results of immunological function suggested that after intravenous administration with p-MWCNTs caused more damage to systemic immunity than MWCNTs-PEG. Here, we demonstrated that a surface functional modification on MWCNTs reduces their immune perturbations in vivo. The chemistry-modified MWCNTs change their preferred immune response in vivo and reduce the immunotoxicity of p-MWCNTs.

As uncontrolled urban growth has increasingly challenged the sustainable use of urban land, it is critically important to model urban growth from different perspectives. Using the SLEUTH (Slope, Land use, Exclusion, Urban, Transportation, and Hill-shade) model, the historical data of Hefei in 2000, 2005, 2010, and 2015 were collected and input to simulate urban growth from 2015 to 2040. Three different urban growth scenarios were considered, namely a historical growth scenario, an urban planning growth scenario, and a land suitability growth scenario. Prediction results show that by 2040 urban built-up land would increase to 1434 km2 in the historical growth scenario, to 1190 km2 in the urban planning growth scenario, and to 1217 km2 in the land suitability growth scenario. We conclude that (1) exclusion layers without effective limits might result in unreasonable prediction of future built-up land; (2) based on the general land use map, the urban growth prediction took the governmental policies into account and could reveal the development hotspots in urban planning; and (3) the land suitability scenario prediction was the result of the trade-off between ecological land and built-up land as it used the MCR -based (minimum cumulative resistance model) land suitability assessment result. It would help to form a compact urban space and avoid excessive protection of farmland and ecological land. Findings derived from this study may provide urban planners with interesting insights on formulating urban planning strategies.

Transcatheter aortic valve implantation (TAVI) is an effective treatment to aortic stenosis in patients with advanced age. However, age is recognized as one of the most important risk factors. The aim of our study is to compare the outcome of TAVI between octogenarian patients and young patients.

Although simplified clinicopathological features and serum tumour markers (STMs) were reported to be associated with the status of mismatch repair (MMR) in colorectal cancer (CRC) patients, their predictive value alone or in combination for MMR status remains unknown.