Vimentin and PSF Act in Concert to Regulate IbeA+ E. coli K1 Induced Activation and Nuclear Translocation of NF-κB in Human Brain Endothelial Cells

IbeA-induced NF-κB signaling through its primary receptor vimentin as well as its co-receptor PSF is required for meningitic E. coli K1 penetration and leukocyte transmigration across the blood-brain barrier (BBB), which are the hallmarks of bacterial meningitis. However, it is unknown how vimentin and PSF cooperatively contribute to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB, which are required for bacteria-mediated pathogenicities.IbeA-induced E. coli K1 invasion, polymorphonuclear leukocyte (PMN) transmigration and IKK/NF-κB activation are blocked by Caffeic acid phenethyl ester (CAPE), an inhibitor of NF-κB. IKKα/β phosphorylation is blocked by ERK inhibitors. Co-immunoprecipitation analysis shows that vimentin forms a complex with IκB, NF-κB and tubulins in the resting cells. A dissociation of this complex and a simultaneous association of PSF with NF-κB could be induced by IbeA in a time-dependent manner. The head domain of vimentin is required for the complex formation. Two cytoskeletal components, vimentin filaments and microtubules, contribute to the regulation of NF-κB. SiRNA-mediated knockdown studies demonstrate that IKKα/β phosphorylation is completely abolished in HBMECs lacking vimentin and PSF. Phosphorylation of ERK and nuclear translocation of NF-κB are entirely dependent on PSF. These findings suggest that vimentin and PSF cooperatively contribute to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB activation. PSF is essential for translocation of NF-κB and ERK to the nucleus.These findings reveal previously unappreciated facets of the IbeA-binding proteins. Cooperative contributions of vimentin and PSF to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB may represent a new paradigm in pathogen-induced signal transduction and lead to the development of novel strategies for the prevention and treatment of bacterial meningitis

Zinc protection of HepG2 cells from sporidesmin toxicity does not require de novo gene transcription

Sporidesmin is an epidithiodioxopiperazine mycotoxin secreted by the saprophytic fungus Pithomyces chartarum. Ingestion of sporidesmin by ruminants grazing on the saprophyte infested pasture causes severe liver and bile duct damage leading to secondary photosensitisation. Zinc supplementation is used as an effective prophylaxis against sporidesmin toxicity in ruminants, however, the mechanism by which zinc protects is unknown. This study used the human hepatoma cell line, HepG2, as a model to examine the mechanism of zinc protection against sporidesmin toxicity. Treatment of cells with various concentrations of sporidesmin (0–10 μg/ml) resulted in a sigmoidal dose response curve with an LC50 of 5 μg/ml. Cells were protected from sporidesmin toxicity by pre-treatment for 2 h or 16 h with zinc sulphate in a concentration dependent manner, with significant protection at 50 μM zinc and maximal protection at 200 μM zinc. To determine whether zinc protection required de novo gene transcription, cells were treated with the transcriptional inhibitor actinomycin D for one hour prior to and throughout the zinc pre-treatment. The presence of actinomycin D did not significantly reduce the zinc protection against sporidesmin cytotoxicity (80% protection without actinomycin D versus 71% protection with actinomycin D). Therefore, de novo gene transcription does not play a major role in the mechanism of zinc protection against sporidesmin toxicity in HepG2 cells

Cloning, mapping and association studies of the ovine ABCG2 gene with facial eczema disease in sheep

Facial eczema (FE) is a hepatogenous mycotoxicosis in sheep caused by the fungal toxin sporidesmin. Resistance to FE is a multigenic trait. To identify QTL associated with this trait, a scan of ovine chromosomes was implemented. In addition, ABCG2 was investigated as a possible positional candidate gene because of its sequence homology to the yeast PDR5 protein and its functional role as a xenobiotic transporter. The sequence of ovine ABCG2 cDNA was obtained from liver mRNA by RT‐PCR and 5′ and 3′ RACE. The predicted protein sequence shares >80% identity with other mammalian ABCG2 proteins. SNPs were identified within exon 6, exon 9 and intron 4. The intron 4 SNP was used to map ABCG2 to ovine chromosome 6 (OAR6), about 2 cM distal to microsatellite marker OarAE101. Interestingly, this chromosomal region contains weak evidence for a FE QTL detected in a previous genome‐scan experiment. To further investigate the association of ABCG2 with FE, allele frequencies for the three SNPs plus three neighbouring microsatellite markers were tested for differences in sheep selected for and against FE. Significant differences were detected in the allele frequencies of the intronic SNP marker among the resistant, susceptible and control lines. No difference in the levels of ABCG2 expression between the resistant and susceptible animals was detected by Northern hybridisation of liver RNA samples. However, significantly higher expression was observed in sporidesmin‐dosed sheep compared with naïve animals. Our inference is that the ABCG2 gene may play a minor role in FE sensitivity in sheep, at least within these selection lines

Rotavirus vaccine RIX4414 efficacy sustained during the third year of life: A randomized clinical trial in an Asian population

RIX4414 (Rotarix™), has shown high efficacy during the first 2-years of life. A 2-year randomized, double-blind, placebo-controlled trial in Singapore, Hong Kong, and Taiwan was extended for another year. Infants (6-17. weeks) received 2-doses (1-2. months apart) of RIX4414 (n= 5359) or placebo (n= 5349). During the third-year follow-up, 4359 (RIX4414) and 4328 (placebo) infants were monitored. 64 (1.2%) and 2 (0.04%) infants in the placebo and RIX4414 groups, respectively, reported severe rotavirus-gastroenteritis (RVGE), resulting in a vaccine efficacy of 96.9% (95% CI [88.3-99.6]). Efficacy was 100% (67.5-100) in the third-year. RIX4414 was efficacious against G1 (100.0% [84.8-100]) and pooled non-G1 RV types (94.9% [80.2-99.4]). This study shows that the vaccine is highly efficacious, regardless of circulating RV-types, up to the first 3. years of life in affluent Asian urban populations. © 2012 Elsevier Ltd.link_to_subscribed_fulltex

Evaluation of RIX4414, a live, attenuated rotavirus vaccine, in a randomized, double-blind, placebo-controlled phase 2 trial involving 2464 Singaporean infants

10.1086/431511Journal of Infectious Diseases192SUPPL.