Two independent replicons can support replication of the anthrax toxin-encoding plasmid pXO1 of Bacillus anthracis

The large pXO1 plasmid (181.6kb) of Bacillus anthracis encodes the anthrax toxin proteins. Previous studies have shown that two separate regions of pXO1 can support replication of pXO1 miniplasmids when introduced into plasmid-less strains of this organism. No information is currently available on the ability of the above two replicons, termed RepX and ORFs 14/16 replicons, to support replication of the full-length pXO1 plasmid. We generated mutants of the full-length pXO1 plasmid in which either the RepX or the ORFs 14/16 replicon was inactivated by TargeTron insertional mutagenesis. Plasmid pXO1 derivatives containing only the RepX or the ORFs 14/16 replicon were able to replicate when introduced into a plasmid-less B. anthracis strain. Plasmid copy number analysis showed that the ORFs 14/16 replicon is more efficient than the RepX replicon. Our studies demonstrate that both the RepX and ORFs 14/16 replicons can independently support the replication of the full-length pXO1 plasmid.     

Angiotensin-(1–7) Via the Mas Receptor Alleviates the Diabetes-Induced Decrease in GFAP and GAP-43 Immunoreactivity with Concomitant Reduction in the COX-2 in Hippocampal Formation: An Immunohistochemical Study

We have previously shown that chronic treatment with angiotensin-(1?C7) [Ang-(1?C7)] can prevent diabetes-induced cardiovascular dysfunction. However, effect of Ang-(1?C7) treatment on diabetes-induced alterations in the CNS is unknown. The aim of this study was to test the hypothesis that treatment with Ang-(1?C7) can produce protection against diabetes-induced CNS changes. We examined the effect of Ang-(1?C7) on the number of cyclooxygenase-2 (COX-2) immunoreactive neurons and the glial fibrillary acidic protein (GFAP)-immunoreactive astrocytes and assessed the changes in the neuronal growth-associated protein-43 (GAP-43) of the hippocampal formation in streptozotocin-induced diabetes in rats. Animals were sacrificed 30?days after induction of diabetes and/or treatment with Ang-(1?C7). Ang-(1?C7) treatment significantly prevented diabetes-induced decrease in the number of GFAP immunoreactive astrocytes and GAP-43 positive neurons in all hippocampal regions. Co-administration of A779, a selective Ang-(1?C7) receptor antagonist, inhibited Ang-(1?C7)-mediated protective effects indicating that Ang-(1?C7) produces its effects through activation of receptor Mas. Further, Ang-(1?C7) treatment through activation of Mas significantly prevented diabetes-induced increase in the number of the COX-2 immunolabeled neurons in all sub-regions of the hippocampus examined. These results show that Ang-(1?C7) has a protective role against diabetes-induced changes in the CNS.     

Bt rice expressing Cry1Ab does not stimulate an outbreak of its non-target herbivore, Nilaparvata lugens

In this study, the non-target effects of Bt rice “KMD2” expressing a Cry1Ab protein on the performance of the brown planthopper (BPH), Nilaparvata lugens, over multiple generations were evaluated under laboratory and field conditions. In the laboratory, BPH was reared to observe the impact of the Bt rice as compared to its parental non-Bt cultivar Xiushui 11, while the population dynamics and oviposition performance of BPH were investigated in the field. The survival of BPH nymphs fed Bt and non-Bt rice did not differ significantly. The nymph developmental duration of BPH was significantly delayed by the Bt rice by comparison with the non-Bt rice for the 1st and 2nd but not the 4th generation. Most importantly, the fecundity of BPH on the Bt rice was significantly decreased in every generation when compared with the non-Bt rice. In the field investigations, the population density of BPH nymphs was significantly lower in the Bt rice field. However, the temporal pattern of population dynamics of BPH adults was similar between the Bt and non-Bt rice, presumably due to migratory interference of the adults. In the Bt rice field, the percentage of tillers with eggs and the number of eggs per tiller were also significantly lower from tillering to mature stage. Additionally, Cry1Ab protein could not be detected in guts from single BPH adults. In general, our results suggest that the Bt rice “KMD2” could not stimulate an outbreak of BPH.     

Diacylglycerol-containing oleic acid induces increases in [Ca]i via TRPC3/6 channels in human T-cells

Though most of the studies have focused on the effects of free fatty acids on T-cell activation, fatty acids incorporated into plasma membrane phospholipids may also affect cell signaling via diacylglycerol (DAG), generally produced by phospholipid hydrolysis. In the present study, we have synthesized a DAG-containing oleic acid and studied its implication in the modulation of calcium signaling in human Jurkat T-cells. 1-palmitoyl-2-oleoyl-sn-glycerol (POG) induced a dose-dependent increase in [Ca²⁺]_i. This effect was due to the presence of oleic acid at the sn-2 position as no differences were observed between POG and 1-stearoly-2-oleoyl-sn-glycerol (SOG). However, the substitution of oleic acid with arachidonic acid at the sn-2 position of the DAG moiety exerted a different response on the increases in [Ca²⁺]_i in these cells. POG-evoked increases in [Ca²⁺]_i were not due to its metabolites. Furthermore, POG-induced increases in [Ca²⁺]_i were due to the opening of TRPC3/TRPC6 channels as silencing of TRPC3 and TRPC6 genes by shRNA abolished calcium entry. Moreover, disruption of lipid rafts with methyl-β-cyclodextrin completely abolished POG-evoked increases in [Ca²⁺]_i. In conclusion, our results demonstrate that oleic acid can influence T-lymphocyte functions, in the conjugated form of DAG, via opening TRPC3/6 channels.     

Mesenchymal stem cells conditioned with glucose depletion augments their ability to repair-infarcted myocardium

Mesenchymal stem cells (MSCs) are an attractive candidate for autologous cell therapy, but their ability to repair damaged myocardium is severely compromised with advanced age. Development of viable autologous cell therapy for treatment of heart failure in the elderly requires the need to address MSC ageing. In this study, MSCs from young (2 months) and aged (24 months) C57BL/6 mice were characterized for gene expression of IGF‐1, FGF‐2, VEGF, SIRT‐1, AKT, p16^INK4a, p21 and p53 along with measurements of population doubling (PD), superoxide dismutase (SOD) activity and apoptosis. Aged MSCs displayed senescent features compared with cells isolated from young animals and therefore were pre‐conditioned with glucose depletion to enhance age affected function. Pre‐conditioning of aged MSCs led to an increase in expression of IGF‐1, AKT and SIRT‐1 concomitant with enhanced viability, proliferation and delayed senescence. To determine the myocardial repair capability of pre‐conditioned aged MSCs, myocardial infarction (MI) was induced in 24 months old C57BL/6 wild type mice and GFP expressing untreated and pre‐conditioned aged MSCs were transplanted. Hearts transplanted with pre‐conditioned aged MSCs showed increased expression of paracrine factors, such as IGF‐1, FGF‐2, VEGF and SDF‐1α. This was associated with significantly improved cardiac performance as measured by dp/dt_max, dp/dt_min, LVEDP and LVDP, declined left ventricle (LV) fibrosis and apoptosis as measured by Masson's Trichrome and TUNEL assays, respectively, after 30 days of transplantation. In conclusion, pre‐conditioning of aged MSCs with glucose depletion can enhance proliferation, delay senescence and restore the ability of aged cells to repair senescent infarcted myocardium.     

Differential fipronil susceptibility and metabolism in two rice stem borers from China

The susceptibilities of larvae of two rice stem borers, namely, Chilo suppressalis (Walker) (Lepidoptera: Crambidae) and Sesamia inferens (Walker) (Lepidoptera: Nocutidae) to fipronil and its metabolites were investigated, and then the activities of microsomal O-demethylase, and glutathione transferase (GST) in two species were measured. The metabolism of fipronil in both stem borers was determined in vivo and in vitro. The LD50 value of fipronil to S. inferens was 118.5-fold higher than that of C. suppressalis. The bioassay results offipronil metabolites showed that the toxicities of sulfone and sulfide were higher than fipronil for both species, and the differential toxicity between sulfone and fipronil was remarkable. Alternatively, the activities of microsomal O-demethylase and GST of C. suppressalis were 1.35- and 2.06-fold higher than S. inferens, respectively. The in vivo and in vitro studies on metabolism of fipronil showed that all of fipronil, sulfone, and sulfide were detected and the content of sulfone was higher than sulfide in both stem borers. The residue of sulfone in C. suppressalis was significantly higher than that in S. inferens. These results suggest that the higher activity of mixed function oxidases may cause the higher capacity of C. suppressalis to produce fipronil-sulfone, which is more toxic than fipronil leading to the higher susceptibility of this species.