Down-regulation of ATF1 leads to early neuroectoderm differentiation of human embryonic stem cells by increasing the expression level of SOX2.

We reveal by high-throughput screening that activating transcription factor 1 (ATF1) is a novel pluripotent regulator in human embryonic stem cells (hESCs). The knockdown of ATF1 expression significantly up-regulated neuroectoderm (NE) genes but not mesoderm, endoderm, and trophectoderm genes. Of note, down-regulation or knockout of ATF1 with short hairpin RNA (shRNA), small interfering RNA (siRNA), or clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) was sufficient to up-regulate sex-determining region Y-box (SOX)2 and paired box 6 (PAX6) expression under the undifferentiated or differentiated conditions, whereas overexpression of ATF1 suppressed NE differentiation. Endogenous ATF1 was spontaneously down-regulated after d 1-3 of neural induction. By double-knockdown experiments, up-regulation of SOX2 was critical for the increase of PAX6 and SOX1 expression in shRNA targeting Atf1 hESCs. Using the luciferase reporter assay, we identified ATF1 as a negative transcriptional regulator of Sox2 gene expression. A novel function of ATF1 was discovered, and these findings contribute to a broader understanding of the very first steps in regulating NE differentiation in hESCs.-Yang, S.-C., Liu, J.-J., Wang, C.-K., Lin, Y.-T., Tsai, S.-Y., Chen, W.-J., Huang, W.-K., Tu, P.-W. A., Lin, Y.-C., Chang, C.-F., Cheng, C.-L., Lin, H., Lai, C.-Y., Lin, C.-Y., Lee, Y.-H., Chiu, Y.-C., Hsu, C.-C., Hsu, S.-C., Hsiao, M., Schuyler, S. C., Lu, F. L., Lu, J. Down-regulation of ATF1 leads to early neuroectoderm differentiation of human embryonic stem cells by increasing the expression level of SOX2.

Antioxidant capacity, insecticidal ability and heat-oxidation stability of Tagetes lemmonii leaf extract.

The aim of this study was to examine the effect of process factors such as ethanol concentration, extraction time and temperature on the extraction yield and the bioactive contents of Tagetes lemmonii leaf extracts using response surface methodology (RSM). ANOVA results showed that the response variables were affected by the ethanol concentration to a very significant degree and by extraction temperature to a lesser degree. GC/MS characterization showed that the extract is rich in bioactive compounds and those present exhibited important biological activities such as antioxidant, insect repellence and insecticidal activities. The results from the toxicity assay demonstrate that the extract obtained from the leaves of Tagetes lemmonii was an effective insect toxin against Tribolium castaneum. The radical scavenging activity and p-anisidine test results of olive oil spiked with different concentrations of leaf extract showed that the phenolic compounds can retard lipid oxidation.

Investigation of the Antioxidant Capacity, Insecticidal Ability and Oxidation Stability of Chenopodium formosanum Seed Extract.

To maximize the extraction of antioxidants from Chenopodium formosanum seeds, the process factors, such as the ethanol concentration (0⁻100%), extraction time (30⁻180 min) and temperature (30⁻70 °C), for the extraction of the bioactive contents as well as the antioxidant capacity are evaluated using response surface methodology (RSM). The experimental results fit well with quadratic models. The extract was identified by GC/MS, and it was found that some active compounds had antioxidant, repellency and insecticidal activities. Various concentrations of the extract were prepared for the evaluation of the insecticidal activity against Tribolium castaneum, and the toxicity test results indicated that the extract was toxic to Tribolium castaneum, with an LC50 value of 354.61 ppm. The oxidative stability of the olive oil determined according to the radical scavenging activity and p-anisidine test demonstrates that the extract obtained from the Chenopodium formosanum seeds can retard lipid oxidation.

Inducible liver-specific overexpression of gankyrin in zebrafish results in spontaneous intrahepatic cholangiocarcinoma and hepatocellular carcinoma formation.

Liver cancer is the second leading cause of death worldwide. As such, establishing animal models of the disease is important for both basic and translational studies that move toward developing new therapies. Gankyrin is a critical oncoprotein in the genetic control of liver pathology. In order to evaluate the oncogenic role of gankyrin without cancer cell inoculation and drug treatment, we overexpressed gankyrin under the control of the fabp10a promoter. A Tet-Off system was used to drive expression in hepatocytes. At seven to twelve months of age, gankyrin transgenic fish spontaneously incurred persistent hepatocyte damage, steatosis, cholestasis, cholangitis, fibrosis and hepatic tumors. The tumors were both hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma (ICC). ICC is the second most frequent primary liver cancer in human patients and the first to develop in this tumor model. We further investigated the role of complement C3, a central molecule of the complement system, and found the expression levels of both in mRNA and protein are decreased during tumorigenesis. Together, these findings suggest that gankyrin can promote malignant transformation of liver cells in the context of persistent liver injury. This transformation may be related to compensatory proliferation and the inflammatory microenvironment. The observed decrease in complement C3 may allow transforming cells to escape coordinated induction of the immune response. Herein, we demonstrate an excellent zebrafish model for liver cancers that will be useful for studying the molecular mechanisms of tumorgenesis.

Transgenic expression of omega-3 PUFA synthesis genes improves zebrafish survival during Vibrio vulnificus infection.

BACKGROUND: Highly desaturated n-3 polyunsaturated fatty acids (PUFAs), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are synthesized by desaturases and elongase. They exert hepatoprotective effects to prevent alcoholic fatty liver syndrome or cholestatic liver injury. However, it is unclear how n-3 PUFAs improve immune function in liver. Vibrio vulnificus, a gram-negative bacterial pathogen, causes high mortality of aquaculture fishes upon infection. Humans can become infected with V. vulnificus through open wounds or by eating raw seafood, and such infections may result in systemic septicemia. Moreover, patients with liver diseases are vulnerable to infection, and are more likely than healthy persons to present with liver inflammation following infection. This study quantified n-3 PUFAs and their anti-bacterial effects in Fadsd6 and Elvol5a transgenic zebrafish. RESULTS: Two transgenic zebrafish strains with strong liver specific expression of Fadsd6 and Elvol5a (driven by the zebrafish Fabp10 promoter) were established using the Tol2 system. Synthesis of n-3 PUFAs in these strains were increased by 2.5-fold as compared to wild type (Wt) fish. The survival rate in 24 h following challenge with V. vulnificus was 20 % in Wt, but 70 % in the transgenic strains. In addition, the bacteria counts in transgenic fish strains were significantly decreased. The expression levels of pro-inflammatory genes, such as TNF-α, IL-1ß, and NF-κB, were suppressed between 9 and 12 h after challenge. This study confirms the anti-bacterial function of n-3 PUFAs in a transgenic zebrafish model. CONCLUSIONS: Fadsd6 and Elvol5a transgenic zebrafish are more resistant to V. vulnificus infection, and enhance survival by diminishing the attendant inflammatory response.

CXCL14 Maintains hESC Self-Renewal through Binding to IGF-1R and Activation of the IGF-1R Pathway.

Human embryonic stem cells (hESCs) have important roles in regenerative medicine, but only a few studies have investigated the cytokines secreted by hESCs. We screened and identified chemokine (C-X-C motif) ligand 14 (CXCL14), which plays crucial roles in hESC renewal. CXCL14, a C-X-C motif chemokine, is also named as breast and kidney-expressed chemokine (BRAK), B cell and monocyte-activated chemokine (BMAC), and macrophage inflammatory protein-2γ (MIP-2γ). Knockdown of CXCL14 disrupted the hESC self-renewal, changed cell cycle distribution, and further increased the expression levels of mesoderm and endoderm differentiated markers. Interestingly, we demonstrated that CXCL14 is the ligand for the insulin-like growth factor 1 receptor (IGF-1R), and it can activate IGF-1R signal transduction to support hESC renewal. Currently published literature indicates that all receptors in the CXCL family are G protein-coupled receptors (GPCRs). This report is the first to demonstrate that a CXCL protein can bind to and activate a receptor tyrosine kinase (RTK), and also the first to show that IGF-1R has another ligand in addition to IGFs. These findings broaden our understanding of stem cell biology and signal transduction.

Hypoxic-state estimation of brain cells by using wireless near-infrared spectroscopy.

Near-infrared spectroscopy (NIRS) is a modern measuring technology in neuroscience. It can be used to noninvasively measure the relative concentrations of oxyhemoglobin (OxyHb) and deoxyhemoglobin (DeoHb), which can reflect information related to cerebral blood volume and cerebral oxygen saturation. Therefore, it has the potential for noninvasive monitoring of cerebral ischemia. However, there is still a lack of reliable physiological information on the relationship between the concentrations of OxyHb and DeoHb in cerebral blood and the exact hypoxic state of brain cells under cerebral ischemia. In this study, we describe a wireless multichannel NIRS system, which we designed to noninvasively monitor the relative concentrations of OxyHb and DeoHb in bilateral cerebral blood before, during, and after middle cerebral artery occlusion. By comparing the results with the lactate/pyruvate ratio measured by microdialysis, we investigated the correlation between the relative concentrations of OxyHb and DeoHb in cerebral blood and the hypoxic state of brain cells. The results showed that the relationship between the concentration changes of DeoHb in cerebral blood and the hypoxic state of brain cells was significant. Therefore, by monitoring the changes in concentrations of DeoHb, the wireless NIRS can be used to estimate the hypoxic state of brain cells indirectly.