MicroRNA-7 promotes neural differentiation of trabecular meshwork mesenchymal stem cell on nanofibrous scaffold

The purpose of this study was to investigate miR-7 overexpression effects on neural differentiation of mesenchymal stem cells (MSCs) on both two-dimensional (2D) and three-dimensional (3D) culture systems. We upregulated miR-7 through lentiviral vector in trabecular meshwork MSCs (TMMSCs) and polymers of poly l -lactic acid/polycaprolactone fibrous scaffold were fabricated by electrospinning and characterized using scanning electron microscopy (SEM) and Fourier transform infrared (FTIR). Neural markers expression was evaluated through quantitative-polymerase chain reaction (q-PCR) and immunostaining. The results showed that the high percentage of cell transduction (84.9%) and miR-7 expression (folds: 677.93 and 556.4) was detected in TMMSCs-miR-7(+). SEM and FTIR established the fabrication of the hybrid scaffold. q-PCR analysis showed that on days 14 and 21 of transduction, the expression level of Nestin and glial fibrillary acidic protein (GFAP) genes were significantly higher in the scaffold (3D) compared with tissue culture polystyrene (2D) culture. The expression of microtubule-associated protein-2 (MAP-2) and GFAP genes in TMMSCs-miR-7(+) cells were significantly higher than those miR-7(−) cells after 21 days of cell culture. Also, MAP-2 and Nestin proteins were detected in TMMSCs-miR-7(+) cells. Our results demonstrate that miR-7 is involved in neural differentiation of TMMSCs and scaffold can improve differentiate into glial and neural progenitor cells. These findings provided some information for future use of microRNAs and scaffold in tissue engineering and cell therapy for neurological diseases.     

Indirect regeneration of Ficus carica by the TCL technique and genetic fidelity evaluation of the regenerated plants using flow cytometry and ISSR

Plant Cell, Tissue and Organ Culture (PCTOC) - Calcium-dependent protein kinases (CDPKs), as an important calcium sensor in plants, are widely involved in the signal transmission process of growth...     

Toward a fine-scale population health monitoring system

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The Interaction of CRM1 and the Nuclear Pore Protein Tpr

While much has been devoted to the study of transport mechanisms through the nuclear pore complex (NPC), the specifics of interactions and binding between export transport receptors and the NPC periphery have remained elusive. Recent work has demonstrated a binding interaction between the exportin CRM1 and the unstructured carboxylic tail of Tpr, on the nuclear basket. Strong evidence suggests that this interaction is vital to the functions of CRM1. Using molecular dynamics simulations and a newly refined method for determining binding regions, we have identified nine candidate binding sites on CRM1 for C-Tpr. These include two adjacent to RanGTP – from which one is blocked in the absence of RanGTP – and three next to the binding region of the cargo Snurportin. We report two additional interaction sites between C-Tpr and Snurportin, suggesting a possible role for Tpr import into the nucleus. Using bioinformatics tools we have conducted conservation analysis and functional residue prediction investigations to identify which parts of the obtained binding sites are inherently more important and should be highlighted. Also, a novel measure based on the ratio of available solvent accessible surface (RASAS) is proposed for monitoring the ligand/receptor binding process.     

AiiA lactonase disrupts N-acylhomoserine lactone and attenuates quorum-sensing-related virulence in Pectobacterium carotovorum EMPCC

Enzymatic degradation of N-acyl homoserine lactone (NAHL) was found to interfere with the quorum sensing (QS) system and related functions in several soil bacteria. In this research, the NAHL lactonase gene aiiA was amplified using aiiA-7F/aiiA7R PCR primers from the quorum sensing inhibitor rhizobacterium Bacillus sp. strain DMS133, and cloned. The plasmid pME7075, carrying the DMS133 aiiA gene under the constitutive lac promoter, was introduced into the plant pathogen Pectobacterium carotovorum EMPCC, creating strain EMPCC/aiiA. Heterologous expression of the DMS133 aiiA gene in EMPCC severely reduced the accumulation of the NAHL throughout growth, and completely prevented pigmentation of the CV026 bioreporter strain. Virulence analysis revealed that the P. carotovorum strain EMPCC/aiiA expressing AiiA lactonase had drastically reduced tissue maceration activity compared with the wild type EMPCC strain. These results provide evidence that AiiA plays an important role in the quorum quenching ability of Bacillus sp. DMS133 whose AHL degradation capacity was investigated previously. In addition, the communication signal-inactivation approach represents a promising strategy for the prevention of diseases in which virulence is regulated by QS signal molecules.     

Brownian dynamics simulation of nucleocytoplasmic transport: a coarse-grained model for the functional state of the nuclear pore complex

The nuclear pore complex (NPC) regulates molecular traffic across the nuclear envelope (NE). Selective transport happens on the order of milliseconds and the length scale of tens of nanometers; however, the transport mechanism remains elusive. Central to the transport process is the hydrophobic interactions between karyopherins (kaps) and Phe-Gly (FG) repeat domains. Taking into account the polymeric nature of FG-repeats grafted on the elastic structure of the NPC, and the kap-FG hydrophobic affinity, we have established a coarse-grained model of the NPC structure that mimics nucleocytoplasmic transport. To establish a foundation for future works, the methodology and biophysical rationale behind the model is explained in details. The model predicts that the first-passage time of a 15 nm cargo-complex is about 2.6±0.13 ms with an inverse Gaussian distribution for statistically adequate number of independent Brownian dynamics simulations. Moreover, the cargo-complex is primarily attached to the channel wall where it interacts with the FG-layer as it passes through the central channel. The kap-FG hydrophobic interaction is highly dynamic and fast, which ensures an efficient translocation through the NPC. Further, almost all eight hydrophobic binding spots on kap-β are occupied simultaneously during transport. Finally, as opposed to intact NPCs, cytoplasmic filaments-deficient NPCs show a high degree of permeability to inert cargos, implying the defining role of cytoplasmic filaments in the selectivity barrier.     

The Amino Terminus of Herpes Simplex Virus 1 Glycoprotein K Is Required for Virion Entry via the Paired Immunoglobulin-Like Type-2 Receptor Alpha

The herpes simplex virus 1 (HSV-1) glycoprotein K (gK)/UL20 protein complex is incorporated into virion envelopes and cellular membranes and functions during virus entry and cell-to-cell spread. To investigate the role of gK/UL20 in the context of a highly neurovirulent virus strain, the HSV-1(McKrae) genome was cloned into a bacterial artificial chromosome plasmid (McKbac) and utilized to construct the mutant virus McK(gKΔ31-68), carrying a 37-amino-acid deletion within the gK amino terminus. The McKbac virus entered efficiently into Chinese hamster ovary (CHO) cells constitutively expressing HSV-1 human receptors, nectin-1, herpesvirus entry mediator (HVEM), or paired immunoglobulin-like type-2 receptor alpha (PILRα). In contrast, the McK(gKΔ31-68) virus failed to enter into CHO-PILRα cells, while it entered CHO cells expressing HVEM and nectin-1 more efficiently than the McKbac virus. Both McKbac and McK(gKΔ31-68) viruses entered all CHO cells expressing HSV-1 receptors via a pH-independent pathway. The HSV-1(F) gBΔ28syn mutant virus, encoding a carboxyl-terminal truncated gB, causes extensive cell fusion. Previously, we showed that the gKΔ31-68 amino acid deletion abrogated gBΔ28syn virus-induced cell fusion, indicating that the amino terminus of gK is required for gB-mediated virus-induced cell fusion (V. N. Chouljenko, A. V. Iyer, S. Chowdhury, D. V. Chouljenko, and K. G. J. Kousoulas, Virology 83:12301–12313, 2009). Surprisingly, the gKΔ31-68/gBΔ28syn virus caused extensive fusion of CHO-nectin-1 cells but limited cell fusion of CHO-PILRα cells. Coimmunoprecipitation experiments revealed that both gK and PILRα bound gB in infected cells. Collectively, these results indicate that the amino terminus of gK is functionally and physically associated with the gB-PILRα protein complex and regulates membrane fusion of the viral envelope with cellular membranes during virus entry as well as virus-induced cell-to-cell fusion.     

Mechanical vulnerability of lower second premolar utilising visco-elastic dynamic stress analysis

Stress analysis determines vulnerability of dental tissues to external loads. Stress values depend on loading conditions, mechanical properties and constrains of structural components. The critical stress levels lead to tissue damage. The aim of this study is to analyse dynamic stress distribution of lower second premolar due to physiological cyclic loading, and dependency of pulsatile stress characteristics to visco-elastic property of dental components by finite element modelling. Results show that visco-elastic property markedly influences stress determinants in major anatomical sites including dentin, cementum–enamel and dentin–enamel junctions. Reduction of visco-elastic parameter leads to mechanical vulnerability through elevation of stress pulse amplitude, maximum stress value; and reduction of stress phase shift as a determinant of stress wave propagation. The results may be applied in situations in which visco-elasticity is reduced such as root canal therapy and post and core restoration in which teeth are more vulnerable to fracture.     

Phenotypic characterization of a tobacco mutant impaired in auxin polar transport

A mutation in tobacco (Nicotiana tabacum L. cv `Xanthi') called lat (low auxin transport) that changes many morphogenic features throughout the life of the plant has been isolated. Abnormalities were observed in seed development, embryogenesis, cotyledon formation, leaf initiation and development, leaf veination pattern, and flower development. Selfed R₂ lat mutant plants set between 60% and 90% fewer seeds than wild-type tobacco, and about 10% of these seeds did not germinate. Non-germinating seeds contained either abnormal embryos or abnormal endosperm tissues. There was no uniformity in the stage at which embryonic development ceased in the aberrant seeds. Seedlings often revealed abnormal and highly varied phenotypes after germination. In some of these cases, cotyledons were heart-shaped, fused, cup-shaped, or cylindrical. Leaf morphology ranged from normal to cup-shaped, and some leaves occasionally produced shoots from the leaf midvein. Flowers ranged from normal to compound with occasional fused floral parts or split petals. Stamens were sometimes petal-like. This unusual assortment of phenotypic changes suggested that the mutation might affect a basic component of plant metabolism. We found that polar transport of indole-3-acetic acid (IAA) was reduced to about 9–19% of the wild-type level in the inflorescence axis of selfed R₂ lat mutants. In addition, supplementation of 1-naphthaleneacetic acid (NAA) to sterile media suppressed some of the abnormalities of the lat mutation so long as the plants grew there. Similarities in the phenotype of embryos, cotyledon and leaf shapes, translocation of labeled IAA, and response to applied NAA indicate that the lat locus of tobacco may be analogous to the pin locus of Arabidopsis, or produce a protein that functions in the same auxin-transport pathway. Received: 18 March 1997 / Revision received: 1 May 1997 / Accepted: 17 June 1997