Abstract:Scintillation proximity assays (SPA), developed from the original radioimmunoassay, is a homogeneous, sensitive, fast and simple scintillation bead-based assay platform. Due to the diversification of radio-labeled ligands and of affinity tags modified on the scintillation carrier’s surface, as well as the development of scintillation recorders and liquid handling technologies, SPA has been widely used as a high throughput method for lead compound screening and biomarker detection. In this review, after a brief introduction of the principle and operation of SPA, a detailed advance of its applications in life sciences were outlined, especially on membrane proteins analysis and at the cell level analysis with detailed classical reports. Beyond that, the methods on increasing the ratios of signal/noise were also analyzed. The wide spread of SPA, especially of its innovations at the cell level research will definitely promote our comprehensive understanding of cellular system biology.

Abstract:The mammalian target of rapamycin(mTOR) is a serine/threonine kinase that regulates cell growth and proliferation, which plays a significant role in the pathogenesis and progression of tumors, including adrenal tumors. Numerous studies have shown that the phosphorylation levels of Akt, mTOR, S6K1 and 4EB-P1, are obviously higher in adrenocortical carcinoma(ACC) and pheochromocytomas(PCC) than in normal adrenal glands, which suggest that PI3K/Akt/mTOR signal pathway is active in adrenal tumors and probably associated with the malignant biological properties. This pathway in adrenal gland can be activated by several factors, including the loss of heterozygosity of IGF2 gene, the germline mutation in PTEN gene and the abnormal expression of microRNA, resulting in the increasing expression of VEGF and cyclin D1, which will promote proliferation, apoptosis resistance, invasion and metastasis of tumors. At present, the treatment of ACC and PCC with mTOR inhibitors has shown satisfactory effect in vitro and in vivo. What's more, a combination of mTOR inhibitors and other anti-cancer drugs provide superior effectiveness, giving new hopes to patients suffering from adrenal tumors. This review summarizes recent advances in understanding the roles of mTOR signal pathway in the pathogenesis and progression of adrenal tumors.

Abstract:Long non-coding RNAs are defined as endogenous molecules with a length greater than 200 nucleotides and with no apparent coding potential. Recent studies showed that lncRNA could interact with the miRNA as a competing endogenous RNAs (ceRNAs) to participate in the expression regulation of target genes, which exert an importance role in the initiation and progression of tumor. In this review, based on lncRNA functioning as ceRNAs, we described the mechanism and function of some relevant lncRNA in the initiation and progression of tumor. A growing number of researches indicated that the interaction between lncRNA and miRNA in tumor genesis would provide new ideas in tumor diagnosis and treatment.

Abstract:LXRα could have the anti-proliferative effect on multiple cancer cells including breast cancer. However, the mechanisms of LXRα regulating the breast cancer cells remain unclear. This study is to investigate the different expression of LXRα, NF-κB p65 and cyclinD1 in the proliferation of human breast cancer cells. At first, LXRα, NF-κB p65 and cyclinD1 expression were detected by immunohistochemical staining in human breast cancer and paired adjacent breast tissues (n=60). As a result, the three kinds of protein were mainly expressed in cell nuclei. Among them, NF-κB p65 and cyclinD1 were higher expressed in breast cancer tissues than in adjacent tissues while LXRα was lower expressed. Then, MTT assay was used to detect the proliferation of MCF-7 cells and Western blot was used to examine the expression of the three kinds of protein. TO901317 (a kind of artificial agonists against LXRs especially for LXRα) could increase LXRα expression, but decrease NF-κB p65 and cyclinD1 expression and suppress the proliferation of MCF-7 cells in a dose- and time-dependent manner (P < 0.05). Finally, the effects of LXRα siRNA and pyrrolidinedithiocarbamic acid (PDTC, an inhibitory of NF-κB) on TO901317 were observed respectively. LXRα siRNA could significantly decrease the up-regulation of LXRα expression and reverse the inhibited effect of TO901317 on cyclinD1 and NF-κB p65 expression and MCF-7 cell proliferation (P < 0.05) while PDTC could strengthen the inhibition of cell proliferation and further down-regulate NF-κB p65 and cyclinD1 expression induced by TO901317 (P < 0.05), but have little effect on LXRα. In a conclusion, the expression of LXRα, NF-κB p65 and cyclinD1 plays an important role in the proliferation of human breast cancer cells, so as to provide a new method for the molecular targeting treatment of breast cancer in the future.

Abstract:T-box transcription factor Tbx18 (Tbx18) was expressed in epicardial cells and controls the differentiation of epicardial epithelial cells into myocardial lineage in the developing mouse embryo. Epithelial-to-mesenchymal transition (EMT) plays a crucial role in embryonic development. For the purpose of determine whether Tbx18 regulates downstream EMT-associated signaling molecules, such as the transcriptional factors Snail1, Slug, Smad, Twist and cell adhesion molecule E-cadherint in mouse heart development. In this study, we used the Tbx18-Cre/Rosa26R-EYFP double-heterozygous mice and Tbx18 mutants to investigate the spatiotemporal expressions patterns of the major EMT-associated signaling molecules within Tbx18 lineage cells. We observed that the major EMT-associated transcriptional factors co-localized with Tbx18 lineage cells in the epicedium and subepicardial mesenchyme. The expression patterns of the major EMT-associated signaling molecules within Tbx18 lineage cells were consistent with Tbx18. Most important, we further demonstrate that the four EMT-associate transcriptional factors are reduced and E-cadherin is significantly increased in the Tbx18 mutant hearts compared with the wild type hearts. Our data together indicated that Tbx18 regulated the major EMT-associated major signaling molecules in mouse heart development. Understanding the signaling pathways by which Tbx18 regulates mouse heart development may help to improve regeneration in adult heart disease.

Abstract:The morphology and elasticity of adhered cells is closely related to the structure of the cytoskeletal meshwork and the way that actomyosin fibers interact with each other to form the network, which is mediated by the activity of myosin Ⅱ. Therefore the interaction between minifilaments (assembled by non-muscle myosin Ⅱ) is important to the cell-level cytoskeletal remodeling through mechanosensing. However, at present it is still difficult to precisely obtain the experimental data of myosin Ⅱ in vivo because the invasive measurement would disturb the cellular local structure and physiological activities, and the non-invasive measurement couldn't provide information about myosin Ⅱ activity accurately. In this paper, we developed and applied new images analysis methods, such as protein fluorescent image tracking and image correlation spectroscopy, to quantify the kinetics of disassembly and reassembly of actomyosin networks and compared them to studies by other groups. This analysis suggested the following processes contribute to the assembly of cortical actomyosin and stress fibers: random myosin mini-filament assembly and disassembly along the cortex; myosin mini-filament aligning and contraction; stabilization of cortical myosin upon increasing contractile tension. We found that the number of myosin Ⅱ and focal adhesions are very important to the formation and stability of the typeⅠ, Ⅱ and Ⅲ actomyosin network in HeLa cells, and the activity of myosin Ⅱ, which determines the dynamics of the actomyosin network reorganization, can be quantified through STICS (spatial temporal image correlation spectroscopy). The formation of typeⅠ, Ⅱ, and Ⅲ actomyosin networks was explained through a mechanical model by adjusting the parameters of myosin Ⅱ activities and number density. The STICS method used in this study can be applied to evaluate the activity of other proteins in live cells.

Abstract:Analytical ultracentrifugation has been utilized to capture the motion of macromolecules in centrifugation field, and is widely applied in assessing protein properties in solution, especially sedimentation coefficient, diffusion coefficient, Stokes radius, molecular mass, and hydrodynamic and thermodynamic parameters, including binding affinity. Utilizing analytical ultracentrifugation, we analyzed the impact of poly acidic amino acids sequence on SnRK2.6's hydrodynamic properties. Chimeric proteins are engineered which combine full length PYL10 and c-terminal poly acidic amino acids sequence of SnRK2.6(333～362) or PDI(441～491). Simultaneously, molecular mass and aggregative state of these proteins were analyzed and calculated by size exclusion chromatography and static light scattering. We demonstrate addition of this sequence leads to increase of Stokes radius, axial ratio, friction ration, and decrease of elution volume on size exclusion chromatography.

Abstract:In Caenorhabditis elegans, the expression of all piRNAs, small of miRNAs were influenced by the mutation of prg-1. Especially, some ncRNA-like, a kinds of small RNA, were found in C. elegans. The ncRNA-like had the similar sequence with piRNAs or miRNAs. The results of the analysis in the gene sites between the ncRNA-like and piRNAs or between ncRNA-like and miRNAs, showed that ncRNA-like had the same gene sites with piRNAs or miRNAs. All of these showed that the ncRNA-like were a kind of small RNA and may be processed from the RNA precursor same to piRNAs or miRNAs.

Abstract:Konjac mannan oligosaccharide (KMOS) is a new food additive with prebiotic function in China. In this study, KMOS with degree of polymerization (DP) of 2～7 was prepared by enzymatic method followed by organic solvent precipitation. Then its subchronic toxicity and genotoxicity was investigated. In the subchronic toxicity test, KMOS was administered to rat orally for 90 days at dose of 0, 2.25, 5.25, 7.50 g/kg body weight (BW) daily, respectively. No significant toxicological manifestation in clinical examination as well as clinical pathology was noted. At terminal necropsy, histopathology changes in the liver and kidney were observed, which was considered to be spontaneous and incidental in nature and unrelated to KMOS-treatment. In addition, a battery of tests including the Ames test, micronucleus test and sperm abnormality test suggested no mutagenicity potential. In conclusion, the results of this study supported that ingestion of KMOS appeared to be safe as a food additive for oral consumption.

Abstract:Nucleic acid bases are an important part of nucleic acids, and Raman spectroscopy is an important technique in studying on molecular structure. Therefore, it has a great significance for the research on the structural changes of nucleic acid macro-molecules and the interactions between nucleic acid molecules and small molecule that the Raman spectroscopy of nucleic acid bases are characterized. In this study, the molecular structures of adenine, guanine, cytosine, thymine and uracil were optimized based on the density functional theory method (DFT). The intra-molecular bond vibration frequency of these five nucleic acid bases were predicted by quantitative calculation, and the Raman spectroscopy of nucleic acid bases was characterized by corresponding the relation between the results of nucleic acid bases Raman spectroscopy in experiment and in theory. Moreover，the characteristic Raman peaks in the Raman spectroscopy of each nucleic acid bases were explanation in detail. It laid the theoretical foundation for the further structural information of nucleic acid molecules by the means of Raman spectra.