Brucella suis vaccine strain S2-infected immortalized caprine endometrial epithelial cell lines induce non-apoptotic ER-stress

Brucella, which is regarded as an intracellular pathogen responsible for a zoonotic disease called brucellosis, survives and proliferates within several types of phagocytic and non-phagocytic cells. Brucella infects not only their preferred hosts but also other domestic and wild animal species, inducing abortion and infertility. Therefore, the interaction between uterine cells and Brucella is important for understanding the pathogenesis of this disease. In this study, we describe the Brucella suis vaccine strain S2 (B.suis.S2) infection and replication in the immortalized caprine endometrial epithelial cell line hTERT-EECs and the induced cellular and molecular response modulation in vitro. We found that B.suis S2 was able to infect and replicate to high titers and inhibit the proliferation of EECs and induce non-apoptotic pathways, as determined by B.suis.S2 detection using MTT and acridine orange/ethidium bromide (AO/EB) staining and flow cytometry. We explored the evidence of non-apoptotic pathways using real-time quantitative RT-PCR and by western blot analysis. Finally, we discovered the over-expression of GRP78, ATF4, ATF6, PERK, eIF2α, CHOP, and cytochrome c (Cyt-c) but not IRE1, xbp-1, and caspase-3 in B.suis.S2 (HK)-attacked and B.suis.S2-infected cells, suggesting that the molecular mechanism of ER stress sensor activation by B.suis.S2 is basically concomitant with that by B.suis.S2 (HK) and that ER stress, especially the PERK pathway, plays an important role in the process of B.suis.S2 infecting EEC, which may, in part, explain the role of the uterus in the pathogenesis of B.suis.S2.

Electronic supplementary material

The online version of this article (doi:10.1007/s12192-014-0564-x) contains supplementary material, which is available to authorized users.     

Nanoporous PtAg and PtCu alloys with hollow ligaments for enhanced electrocatalysis and glucose biosensing

Nanoporous silver (NPS) and copper (NPC) obtained by dealloying AgAl and CuAl alloys, respectively, were used as both three-dimensional templates and reducing agents for the fabrication of nanoporous PtAg (NPS-Pt) and PtCu (NPC-Pt) alloys with hollow ligaments by a simple galvanic replacement reaction with H(2)PtCl(6). Electron microscopy and X-ray diffraction characterizations demonstrate that NPS and NPC with similar ligament sizes (30-50 nm) have different effects on the formed hollow nanostructures. For NPS-Pt, the shell of the hollow ligament is seamless. However, the shell of NPC-Pt is comprised of small pores and alloy nanoparticles with a size of ～3 nm. The as-prepared NPS-Pt and NPC-Pt exhibit remarkably improved electrocatalytic activities towards the oxidation of ethanol and H(2)O(2) compared with state-of-the-art Pt/C catalyst, and can be used for sensitive electrochemical sensing applications. The hierarchical nanoporous structure also provides a good microenvironment for enzymes. After immobilization of glucose oxidase (GOx), the enzyme modified nanoporous electrode can sensitively detect glucose in a wide linear range (0.6-20 mM).     

General allometric equations and biomass allocation of <Emphasis Type="Italic">Pinus massoniana</Emphasis> trees on a regional scale in southern China

Applying allometric equations in combination with forest inventory data is an effective approach to use when qualifying forest biomass and carbon storage on a regional scale. The objectives of this study were to (1) develop general allometric tree component biomass equations and (2) investigate tree biomass allocation patterns for Pinus massoniana, a principal tree species native to southern China, by applying 197 samples across 20 site locations. The additive allometric equations utilized to compute stem, branch, needle, root, aboveground, and total tree biomass were developed by nonlinear seemingly unrelated regression. Results show that the relative proportion of stem biomass to tree biomass increased while the contribution of canopy biomass to tree biomass decreased as trees continued to grow through time. Total root biomass was a large biomass pool in itself, and its relative proportion to tree biomass exhibited a slight increase with tree growth. Although equations employing stem diameter at breast height (dbh) alone as a predictor could accurately predict stem, aboveground, root, and total tree biomass, they were poorly fitted to predict the canopy biomass component. The inclusion of the tree height (H) variable either slightly improved or did not in any way increase model fitness. Validation results demonstrate that these equations are suitable to estimate stem, aboveground, and total tree biomass across a broad range of P. massoniana stands on a regional scale.     

Amperometric biosensor based on tyrosinase-conjugated polysaccharide hybrid film: selective determination of nanomolar neurotransmitters metabolite of 3,4-dihydroxyphenylacetic acid (DOPAC) in biological fluid

The amperometric detection of neurotransmitters metabolite of 3,4-dihydroxyphenylacetic acid (DOPAC) was achieved at a tyrosinase-chitosan composite film-modified glassy carbon (GC) electrode. The optimal conditions for the preparation of the biosensor were established. This bio-composite film was characterized by scanning electron microscopy (SEM) and Fourier transformed infrared (FT-IR) spectra, suggesting that chitosan covalently connected to chitosan chains. Electrochemical characterization of the bio-hybrid membrane-covered electrodes were also performed in 0.05 M phosphate buffer solution (pH 6.52) containing neurotransmitters or their derivatives by using cyclic voltammetry (CV), linear sweep voltammetry (LSV), square wave voltammetry (SWV) and amperometry. This simply-prepared protein-polysaccharide hybrid film provides a microenvironment friendly for enzyme loading. The sensor was operated at -0.15 V with a short response time. The current linearly increased with the increasing concentration of DOPAC over the concentration of 6 nM-0.2 mM. The lower detection limit for DOPAC is 3 nM (S/N=3). The sensitivity of the sensor is 40 microA mM(-1). A physiological level of neurotransmitters and their derivatives including dopamine, l-dopa, adrenaline, noradrenaline and homovanillic acid as well as ascorbic acid, uric acid and acetaminophen do not affect the determination of DOPAC.     

Semiconductor nanocrystals for biological imaging

Conventional organic fluorophores suffer from poor photo stability, narrow absorption spectra and broad emission spectra. Semiconductor nanocrystals, however, are highly photo-stable with broad absorption spectra and narrow size-tunable emission spectra. Recent advances in the synthesis of these materials have resulted in the generation of bright, sensitive, extremely photo-stable and biocompatible semiconductor fluorophores. Commercial availability facilitates their application in a variety of unprecedented biological experiments, including multiplexed cellular imaging, long-term in vitro and in vivo labeling, deep tissue structure mapping and single particle investigation of dynamic cellular processes. Semiconductor nanocrystals are one of the first examples of nanotechnology enabling a new class of biomedical applications.     

Activation of p21-activated kinase 6 by MAP kinase kinase 6 and p38 MAP kinase

The p21-activated kinases (PAKs) contain an N-terminal Cdc42/Rac interactive binding domain, which in the group 1 PAKs (PAK1, 2, and 3) regulates the activity of an adjacent conserved autoinhibitory domain. In contrast, the group 2 PAKs (PAK4, 5, and 6) lack this autoinhibitory domain and are not activated by Cdc42/Rac binding, and the mechanisms that regulate their kinase activity have been unclear. This study found that basal PAK6 kinase activity was repressed by a p38 mitogen-activated protein (MAP) kinase antagonist and could be strongly stimulated by constitutively active MAP kinase kinase 6 (MKK6), an upstream activator of p38 MAP kinases. Mutation of a consensus p38 MAP kinase target site at serine 165 decreased PAK6 kinase activity. Moreover, PAK6 was directly activated by MKK6, and mutation of tyrosine 566 in a consensus MKK6 site (threonine-proline-tyrosine, TPY) in the activation loop of the PAK6 kinase domain prevented activation by MKK6. PAK6 activation by MKK6 was also blocked by mutation of an autophosphorylated serine (serine 560) in the PAK6 activation loop, indicating that phosphorylation of this site is necessary for MKK6-mediated activation. PAK4 and PAK5 were similarly activated by MKK6, consistent with a conserved TPY motif in their activation domains. The activation of PAK6 by both p38 MAP kinase and MKK6 suggests that PAK6 plays a role in the cellular response to stress-related signals.