Crystallization and preliminary X-ray crystallographic analysis of SEDL

SEDL (known also as sedlin) is a 140 amino-acid protein with a putative role in endoplasmic reticulum-to-Golgi transport. Several missense mutations and deletion mutations in the SEDL gene, which result in protein truncation by frame shift, are responsible for spondyloepiphyseal dysplasia tarda, a progressive skeletal disorder. The protein is identical to MIP-2A, which was shown to interact physically with c-myc promotor-binding protein 1 (MBP-1) and relieve the regulatory role of MBP-1 as a general transcription repressor. In order to gain insights into the function of SEDL by structural analysis, the protein was overexpressed and crystallized as a first step. SEDL was overexpressed in Escherichia coli and crystallized using the hanging-drop vapour-diffusion method at 298 K. The crystals belong to the orthorhombic space group C2221, with unit-cell parameters a = 46.69, b = 101.30, c = 66.15 A. The unit cell is likely to contain one molecule of SEDL, with a crystal volume per protein mass (VM) of 2.36 A3 Da-1 and a solvent content of about 47.9% by volume. A native data set to 2.8 A resolution was obtained from a flash-cooled crystal using synchrotron radiation.open1

Cdk5 Phosphorylates Dopamine D2 Receptor and Attenuates Downstream Signaling

The dopamine D2 receptor (DRD2) is a key receptor that mediates dopamine-associated brain functions such as mood, reward, and emotion. Cyclin-dependent kinase 5 (Cdk5) is a proline-directed serine/threonine kinase whose function has been implicated in the brain reward circuit. In this study, we revealed that the serine 321 residue (S321) in the third intracellular loop of DRD2 (D2i3) is a novel regulatory site of Cdk5. Cdk5-dependent phosphorylation of S321 in the D2i3 was observed in in vitro and cell culture systems. We further observed that the phosphorylation of S321 impaired the agonist-stimulated surface expression of DRD2 and decreased G protein coupling to DRD2. Moreover, the downstream cAMP pathway was affected in the heterologous system and in primary neuronal cultures from p35 knockout embryos likely due to the reduced inhibitory activity of DRD2. These results indicate that Cdk5-mediated phosphorylation of S321 inhibits DRD2 function, providing a novel regulatory mechanism for dopamine signaling.X111111sciescopu

Toxicity assessment of modified Cry1Ac1 proteins and genetically modified insect-resistant Agb0101 rice

Insect-resistant Agb0101 rice was recently developed by modifying the cry1ac1 gene by changing codon usage changes relative to the native truncated cry1ac1 gene. To assess the toxicity of genetically modified Agb0101 rice, we conducted bioinfomational comparisons of the amino acid sequences that are not similar to known toxic proteins. Sufficient quantities of mCry1Ac1 protein were produced in Escherichia coli for in vitro evaluation and animal study. We compared the amino acid sequences and molecular mass. There have the same amino acid sequences and molecular masses after purifying the modified Cry1Ac1 (mCry1Ac1) protein from highly expressed bacteria and genetically modified rice were identical. We also investigated the acute and 90-days oral toxicities. No adverse effects were observed in mice following acute oral exposure to 2,000 mg/ kg body weight mCry1Ac1 protein of body weight and 90 days oral exposure to Agb0101. These results indicate that mCry1Ac1 proteins and Agb0101 rice demonstrate no adverse effects in these tests when applied via gavage and feed, respectively.Key words: Modified Cry1Ac1, food safety assessment, toxicity, insect- resistant rice Agb0101

Intravaginal Administration of Fc-Fused IL7 Suppresses the Cervicovaginal Tumor by Recruiting HPV DNA Vaccine-Induced CD8 T Cells

Purpose: The induction of tissue-localized virus-specific CD8 T-cell response is essential for the development of an effective therapeutic vaccine against genital diseases, such as cervical cancer and genital herpes. Here, we aimed to elucidate the immunologic role of IL7 in the induction of mucosal cellular immunity. Experimental Design: IL7 was engineered through Fc fusion to enhance mucosal delivery across the genital epithelial barrier. The immunomodulatory role of IL7 was evaluated by monitoring the kinetics of various immune cells and measuring the expression of chemokines and cytokines after intravaginal administration of Fc-fused IL7 (IL7-Fc). The antitumor effects of intramuscular human papillomavirus (HPV) DNA vaccine or topical IL7-Fc alone or in a combinational regimen on mice survival were compared using a orthotopic cervical cancer model. Results: Intravaginal treatment of IL7-Fc, but not native IL7, induces upregulation of chemokines (CXCL10, CCL3, CCL4, and CCL5), cytokines (IFN-gamma, TNF alpha, IL6, and IL1 beta), and an adhesion molecule (VCAM-1) in the genital tract, leading to the recruitment of several leukocytes, including CD4, CD8, gamma delta T cells, and dendritic cells. Importantly, in this murine cervical cancer model, topical administration of IL7-Fc after intramuscular HPV DNA vaccination increases the number of HPV-specific CD8 T cells in the genital mucosa, but not in the spleen, leading to stronger antitumor activity than the HPV DNA vaccine alone. Conclusions: Our findings provide an important insight into the immunomodulatory role of IL7-Fc via topical application and the design of therapeutic vaccine regimen that induces effective genital-mucosal CD8 T-cell responses.1110Ysciescopu

Wnt5a stimulates chemotactic migration and chemokine production in human neutrophils

Wnt5a is a ligand that activates the noncanonical Wnt signaling pathways (??-catenin-independent pathways). Human neutrophils expressed several Wnt5a receptors, such as Frizzled 2, 5 and 8. Stimulation of human neutrophils with Wnt5a caused chemotactic migration and the production of two important chemokines, CXCL8 and CCL2. CCL2 production by Wnt5a was mediated by a pertussis toxin-sensitive G-protein-dependent pathway. Wnt5a also stimulated the phosphorylation of three mitogen-activated protein kinases (MAPKs: ERK, p38 MAPK and JNK) and Akt. Inhibition of ERK, p38 MAPK or JNK by specific inhibitors induced a dramatic reduction in Wnt5a-induced CCL2 production. Supernatant collected from lipopolysaccharide-stimulated macrophages induced neutrophil chemotaxis, which was significantly inhibited by anti-Wnt5a antibody. Our results suggested that Wnt5a may contribute to neutrophil recruitment, mediating the inflammation response.open4

Preferential Paths of Air-water Two-phase Flow in Porous Structures with Special Consideration of Channel Thickness Effects.

Accurate understanding and predicting the flow paths of immiscible two-phase flow in rocky porous structures are of critical importance for the evaluation of oil or gas recovery and prediction of rock slides caused by gas-liquid flow. A 2D phase field model was established for compressible air-water two-phase flow in heterogenous porous structures. The dynamic characteristics of air-water two-phase interface and preferential paths in porous structures were simulated. The factors affecting the path selection of two-phase flow in porous structures were analyzed. Transparent physical models of complex porous structures were prepared using 3D printing technology. Tracer dye was used to visually observe the flow characteristics and path selection in air-water two-phase displacement experiments. The experimental observations agree with the numerical results used to validate the accuracy of phase field model. The effects of channel thickness on the air-water two-phase flow behavior and paths in porous structures were also analyzed. The results indicate that thick channels can induce secondary air flow paths due to the increase in flow resistance; consequently, the flow distribution is different from that in narrow channels. This study provides a new reference for quantitatively analyzing multi-phase flow and predicting the preferential paths of immiscible fluids in porous structures

Platinum Assisted Vapor–Liquid–Solid Growth of Er–Si Nanowires and Their Optical Properties

We report the optical activation of erbium coated silicon nanowires (Er–SiNWs) grown with the assist of platinum (Pt) and gold (Au), respectively. The NWs were grown on Si substrates by using a chemical vapor transport process using SiCl4 and ErCl4 as precursors. Pt as well as Au worked successfully as vapor–liquid–solid (VLS) catalysts for growing SiNWs with diameters of ~100 nm and length of several micrometers, respectively. The SiNWs have core–shell structures where the Er-crystalline layer is sandwiched between silica layers. Photoluminescence spectra analyses showed the optical activity of SiNWs from both Pt and Au. A stronger Er3+ luminescence of 1,534 nm was observed from the SiNWs with Pt at room- and low-temperature (25 K) using the 488- and/or 477-nm line of an Ar laser that may be due to the uniform incorporation of more Er ions into NWs with the exclusion of the formation of catalyst-induced deep levels in the band-gap. Pt would be used as a VLS catalyst for high performance optically active Er–SiNWs

Association of TLR7 Variants with AIDS-Like Disease and AIDS Vaccine Efficacy in Rhesus Macaques

In HIV infection, TLR7-triggered IFN-α production exerts a direct antiviral effect through the inhibition of viral replication, but may also be involved in immune pathogenesis leading to AIDS. TLR7 could also be an important mediator of vaccine efficacy. In this study, we analyzed polymorphisms in the X-linked TLR7 gene in the rhesus macaque model of AIDS. Upon resequencing of the TLR7 gene in 36 rhesus macaques of Indian origin, 12 polymorphic sites were detected. Next, we identified three tightly linked single nucleotide polymorphisms (SNP) as being associated with survival time. Genotyping of 119 untreated, simian immunodeficiency virus (SIV)-infected male rhesus macaques, including an ‘MHC adjusted’ subset, revealed that the three TLR7 SNPs are also significantly associated with set-point viral load. Surprisingly, this effect was not observed in 72 immunized SIV-infected male monkeys. We hypothesize (i) that SNP c.13G>A in the leader peptide is causative for the observed genotype-phenotype association and that (ii) the underlying mechanism is related to RNA secondary structure formation. Therefore, we investigated a fourth SNP (c.-17C>T), located 17 bp upstream of the ATG translation initiation codon, that is also potentially capable of influencing RNA structure. In c.13A carriers, neither set-point viral load nor survival time were related to the c.-17C>T genotype. In c.13G carriers, by contrast, the c.-17C allele was significantly associated with prolonged survival. Again, no such association was detected among immunized SIV-infected macaques. Our results highlight the dual role of TLR7 in immunodeficiency virus infection and vaccination and imply that it may be important to control human AIDS vaccine trials, not only for MHC genotype, but also for TLR7 genotype

Ultrasonic reflection coefficient and surface roughness index of OA articular cartilage: relation to pathological assessment

<p>Abstract</p> <p>Background</p> <p>Early diagnosis of Osteoarthritis (OA) is essential for preventing further cartilage destruction and decreasing severe complications. The aims of this study are to explore the relationship between OA pathological grades and quantitative acoustic parameters and to provide more objective criteria for ultrasonic microscopic evaluation of the OA cartilage.</p> <p>Methods</p> <p>Articular cartilage samples were prepared from rabbit knees and scanned using ultrasound biomicroscopy (UBM). Three quantitative parameters, including the roughness index of the cartilage surface (URI), the reflection coefficients from the cartilage surface (R) and from the cartilage-bone interface (R_bone) were extracted. The osteoarthritis grades of these cartilage samples were qualitatively assessed by histology according to the grading standards of International Osteoarthritis Institute (OARSI). The relationship between these quantitative parameters and the osteoarthritis grades was explored.</p> <p>Results</p> <p>The results showed that URI increased with the OA grade. URI of the normal cartilage samples was significantly lower than the one of the OA cartilage samples. There was no significant difference in URI between the grade 1 cartilage samples and the grade 2 cartilage samples. The reflection coefficient of the cartilage surface reduced significantly with the development of OA (p < 0.05), while the reflection coefficient of the cartilage-bone interface increased with the increase of grade.</p> <p>Conclusion</p> <p>High frequency ultrasound measurements can reflect the changes in the surface roughness index and the ultrasound reflection coefficients of the cartilage samples with different OA grades. This study may provide useful information for the quantitative ultrasonic diagnosis of early OA.</p

Quantitative permeability imaging of plant tissues

A method for mapping tissue permeability based on time-dependent diffusion measurements is presented. A pulsed field gradient sequence to measure the diffusion encoding time dependence of the diffusion coefficients based on the detection of stimulated spin echoes to enable long diffusion times is combined with a turbo spin echo sequence for fast NMR imaging (MRI). A fitting function is suggested to describe the time dependence of the apparent diffusion constant in porous (bio-)materials, even if the time range of the apparent diffusion coefficient is limited due to relaxation of the magnetization. The method is demonstrated by characterizing anisotropic cell dimensions and permeability on a subpixel level of different tissues of a carrot (Daucus carota) taproot in the radial and axial directions