Deciphering the effects of PYCR1 on cell function and its associated mechanism in hepatocellular carcinoma

Overexpression of pyrroline-5-carboxylate reductase 1 (PYCR1) has been associated with the development of certain cancers; however, no studies have specifically examined the role of PYCR1 in hepatocellular carcinoma (HCC). Based on The Cancer Genome Atlas expression array and meta-analysis conducted using the Gene Expression Omnibus database, we determined that PYCR1 was upregulated in HCC compared to adjacent nontumor tissues (P < 0.05). These data were verified using quantitative real-time polymerase chain reaction, western blotting, and immunohistochemistry analysis. Additionally, patients with low PYCR1 expression showed a higher overall survival rate than patients with high PYCR1 expression. Furthermore, PYCR1 overexpression was associated with the female sex, higher levels of alpha-fetoprotein, advanced clinical stages (III and IV), and a younger age (< 45 years old). Silencing of PYCR1 inhibited cell proliferation, invasive migration, epithelial-mesenchymal transition, and metastatic properties in HCC in vitro and in vivo. Using RNA sequencing and bioinformatics tools for data-dependent network analysis, we found binary relationships among PYCR1 and its interacting proteins in defined pathway modules. These findings indicated that PYCR1 played a multifunctional role in coordinating a variety of biological pathways involved in cell communication, cell proliferation and growth, cell migration, a mitogen-activated protein kinase cascade, ion binding, etc. The structural characteristics of key pathway components and PYCR1-interacting proteins were evaluated by molecular docking, and hotspot analysis showed that better affinities between PYCR1 and its interacting molecules were associated with the presence of arginine in the binding site. Finally, a candidate regulatory microRNA, miR-2355-5p, for PYCR1 mRNA was discovered in HCC. Overall, our study suggests that PYCR1 plays a vital role in HCC pathogenesis and may potentially serve as a molecular target for HCC treatment.     

Matrix homeostasis within the immature annulus fibrosus depends on the frequency of dynamic compression: a study based on the self-developed mechanically active bioreactor

Evidence suggests that mechanical load is related to structural destruction of disk annulus fibrosus (AF) either in adult disk degeneration or in child disk acute injury. Both biochemical and biomechanical properties are different between immature and mature disks. However, the effects of mechanical compression on immature AF are not fully clear. This study was to investigate the effects of a relatively wide range of dynamic compressive frequency on matrix homeostasis within the immature AF. Immature disks from pig (3–4 months) were randomly assigned into the control group (non-compression) and compression groups (0.1, 0.5, 1.0, 3.0 and 5.0 Hz). All disks were bioreactor-cultured for 7 days. AF matrix production was evaluated by histology, gene expression, glycosaminoglycan (GAG) content, hydroxyproline (HYP) content and immunohistochemistry. Generally, no obvious difference was found in HE staining between control group and compression groups. However, alcian blue staining indicated proteoglycan content in the 5.0-Hz group was decreased compared with the control group and other compression groups. Similarly, a catabolic remodeling gene expression profile with the down-regulated matrix genes (aggrecan, collagen I and collagen II) and tissue inhibitor of metalloproteinases (TIMP-1 and TIMP-3) and the up-regulated matrix catabolic enzymes (ADAMTS-4 and MMP-3) was found in the 5.0-Hz group. Further analysis indicated that GAG content, HYP content and aggrecan protein deposition were also decreased in the 5.0-Hz group. Hence, we concluded that matrix homeostasis within the immature AF was compressive frequency dependent, and the relatively higher frequency (5.0 Hz) is unfavorable for matrix production within the immature AF. These findings will contribute to further understanding of the relationship between mechanical compression and immature AF biosynthesis.     

A Novel Rat Model of Nonalcoholic Fatty Liver Disease Constructed Through CRISPR/Cas-Based Hydrodynamic Injection

CRISPR/Cas technology has been widely used in generating conventional and conditional gene knockout animals through germline mutation. A recent study has demonstrated that CRISPR/Cas system also worked on nongermline mutation in mice liver via hydrodynamic injection of vector plasmid to blood circulation. However, whether this is also applied to rat and what is the optimal concentration of vector plasmid still need to be studied. Here, we attempted to use pX330-Pten plasmid to knock down the expression of Pten in rat liver for generation of nonalcoholic fatty liver disease (NAFLD) rat model. A range of pX330-Pten dosages (75, 150 and 300 μg/100 g) in 10 ml of saline water was injected into the blood circulation of SD rats within 30 s by tail vein injection. The data showed that, 9 weeks after injection, no differences of TC, AST and ALT were found between control and the treatment groups (low-dose, mid-dose, high-dose and fat-diet groups). However, the high-dose group revealed severe lipid deposition and significant knockdown of Pten, which was validated by western blot and real-time PCR. Notably, lipid deposition and Pten knockdown were detected in the liver of animals from the high-dose group as early as 6 weeks after injection. Taken together, our findings demonstrated that hydrodynamic injection of pX330-Pten plasmid at the dosage of 300 μg/100 g can knock down the expression of Pten in rat liver efficiently, resulting in lipid deposition as well as NAFLD. Taken together, this study presents a novel and efficient method to edit genome in rat liver.     

Copalic acid analogs down-regulate androgen receptor and inhibit small chaperone protein

Copalic acid, one of the diterpenoid acids in copaiba oil, inhibited the chaperone function of α-crystallin and heat shock protein 27 kD (HSP27). It also showed potent activity in decreasing an HSP27 client protein, androgen receptor (AR), which makes it useful in prostate cancer treatment or prevention. To develop potent drug candidates to decrease the AR level in prostate cancer cells, more copalic acid analogs were synthesized. Using the level of AR as the readout, 15 of the copalic acid analogs were screened and two compounds were much more potent than copalic acid. The compounds also dose-dependently inhibited AR positive prostate cancer cell growth. Furthermore, they inhibited the chaperone activity of α-crystallin as well.     

Higher-order oligomerization targets plasma membrane proteins and HIV gag to exosomes

Exosomes are secreted organelles that have the same topology as the cell and bud outward (outward is defined as away from the cytoplasm) from endosome membranes or endosome-like domains of plasma membrane. Here we describe an exosomal protein-sorting pathway in Jurkat T cells that selects cargo proteins on the basis of both higher-order oligomerization (the oligomerization of oligomers) and plasma membrane association, acts on proteins seemingly without regard to their function, sequence, topology, or mechanism of membrane association, and appears to operate independently of class E vacuolar protein-sorting (VPS) function. We also show that higher-order oligomerization is sufficient to target plasma membrane proteins to HIV virus-like particles, that diverse Gag proteins possess exosomal-sorting information, and that higher-order oligomerization is a primary determinant of HIV Gag budding/exosomal sorting. In addition, we provide evidence that both the HIV late domain and class E VPS function promote HIV budding by unexpectedly complex, seemingly indirect mechanisms. These results support the hypothesis that HIV and other retroviruses are generated by a normal, nonviral pathway of exosome biogenesis.     

C／EBPβ作为LIF的中介维持小鼠胚干细胞于未分化状态

C／EBPβ（CCAAT／增强子结合蛋白β,又称NF—IL6）是一个多功能的转录因子,其主要功能之一是促进细胞分化。白血病抑制因子（LIF）是一个细胞因子．其在不同类型的细胞中具有不同的效应：它诱导前脂肪细胞分化,却抑制小鼠胚多能干细胞（mES）的分化。在mES中,C／EBPβ究竟起什么作用．尚未有报道。本文首次报告在mES中。在LIF蛋白存在下,C／EBPβ的作用是维持mES的未分化状态．即C／EBPβ是LIF的调控对象。主要事实如下。在mES细胞中．内源C／EBPβ蛋白的表达量与加到培养基中的LIF蛋白的量呈正相关。而在未分化的mES细胞中人工高表达的外源C／EBPβ蛋白和其截短形式LIP蛋白。在LIF存在下．也不但不促进反而抑制mES细胞分化,C／EBPβ的大分子异型蛋白还显著促进mES细胞的增殖：而且,在LIF去除后,这种促进mES细胞增殖的效应还能持续一段短时间。当LIF不存在时。C／EBPβ和LIP才如所预期的那样．诱导分化相关基因表达并促进细胞分化。C／EBPβ和LIP所调控的某些分化相关的基因的表达水平．当LIF存在时．也比没有LIF时显著降低。因此,在mES细胞中C／EBPβ是受LIF的调控而作为LIF的中介．维持mES于未分化状态。     

Biodiversity variability of macrobenthic in the Yellow Sea and East China Sea between 2001 and 2011

In this study, we compared the biodiversity and ecological status of macrobenthos between 2001 and 2011 in terms of species composition, dominant species, abundance, biomass, diversity, and secondary productivity of macrobenthos in the Yellow Sea and East China Sea. The results indicate that the species abundances, biomass, and diversity in spring 2011 were all lower than those in autumn 2000 and spring of 2001. Most of the dominant species have changed from 2001 to 2011, with the exception of the echinoderm Ophiura sarsii vadicola that was dominant in these years. Small-sized animal species have increased possibly due to gradual environment deterioration such as the pollution and the climate warmer.     

CARP Is a Potential Tumor Suppressor in Gastric Carcinoma and a Single-Nucleotide Polymorphism in CARP Gene Might Increase the Risk of Gastric Carcinoma

Background

The caspase-associated recruitment domain-containing protein (CARP) is expressed in almost all tissues. Recently, the tumor-suppressive function of CARP was discovered and attracted increasing attention. This study aimed to investigate the role of CARP in the carcinogenesis of human gastric carcinoma.

Methodology/Principal Findings

Compared with normal gastric tissue, the downregulation of CARP expression was observed in gastric carcinoma tissue by cDNA array and tissue microarray assay. In vitro, the gastric carcinoma cell line (BGC-823) was stably transfected with pcDNA3.1B-CARP or plus CARP siRNA, and we used MTT, flow cytometry, cell migration on type I collagen, cell-matrix adhesion assay and western blot analysis to investigate the potential anti-tumor effects of CARP. The data showed that overexpressing CARP suppressed the malignancy of gastric carcinoma BGC-823 cell line, including significant increases in apoptosis, as well as obvious decreases in cell proliferation, migration, adhesion ability, and tumor growth. The tumor-suppressive effects of CARP were almost restored by siRNA-directed CARP silence. In addition, overexpression of CARP induced G1 arrest, decreased the expressions of cyclin E and CDK2, and increased the expressions of p27, p53 and p21. In vivo, the tumor-suppressive effect of CARP was also verified. A single-nucleotide polymorphism (SNP) genotype of CARP (rs2297882) was located in the Kozak sequence of the CARP gene. The reporter gene assay showed that rs2297882 TT caused an obvious downregulation of activity of CARP gene promoter in BGC-823 cells. Furthermore, the association between rs2297882 and human gastric carcinoma susceptibility was analyzed in 352 cases and 889 controls. It displayed that the TT genotype of rs2297882 in the CARP gene was associated with an increased risk of gastric carcinoma.

Conclusions/Significance

CARP is a potential tumor suppressor of gastric carcinoma and the rs2297882 C>T phenotype of CARP may serve as a predictor of gastric carcinoma.     

Monitoring Urban Greenness Dynamics Using Multiple Endmember Spectral Mixture Analysis

Urban greenness is increasingly recognized as an essential constituent of the urban environment and can provide a range of services and enhance residents’ quality of life. Understanding the pattern of urban greenness and exploring its spatiotemporal dynamics would contribute valuable information for urban planning. In this paper, we investigated the pattern of urban greenness in Hangzhou, China, over the past two decades using time series Landsat-5 TM data obtained in 1990, 2002, and 2010. Multiple endmember spectral mixture analysis was used to derive vegetation cover fractions at the subpixel level. An RGB-vegetation fraction model, change intensity analysis and the concentric technique were integrated to reveal the detailed, spatial characteristics and the overall pattern of change in the vegetation cover fraction. Our results demonstrated the ability of multiple endmember spectral mixture analysis to accurately model the vegetation cover fraction in pixels despite the complex spectral confusion of different land cover types. The integration of multiple techniques revealed various changing patterns in urban greenness in this region. The overall vegetation cover has exhibited a drastic decrease over the past two decades, while no significant change occurred in the scenic spots that were studied. Meanwhile, a remarkable recovery of greenness was observed in the existing urban area. The increasing coverage of small green patches has played a vital role in the recovery of urban greenness. These changing patterns were more obvious during the period from 2002 to 2010 than from 1990 to 2002, and they revealed the combined effects of rapid urbanization and greening policies. This work demonstrates the usefulness of time series of vegetation cover fractions for conducting accurate and in-depth studies of the long-term trajectories of urban greenness to obtain meaningful information for sustainable urban development.     

The Structure of Remsamabilinin B

A new cucurbitacin glycoside, named hemsamabilinin B (2), was isolat-ed from Hemsleya amabilis Diels. 2 was hydrolyzed by enzyme on acid to produce 23,24-dihydrocucurbitacin F (3) or D-glucose respectively. On the basis of the spectrosco-pic data of 2 and its acetate (4) (1H NMR, 13CNMR and MS), the structure of 2 wasassigned as 2-0-β-D-glucopyranoside of 3.