Mild hypothermia pretreatment protects hepatocytes against ischemia reperfusion injury via down-regulating miR-122 and IGF-1R/AKT pathway.

BACKGROUND: Mild hypothermia has been well known as an effective way to reduce ischemia reperfusion injury (IRI), while the mechanisms are still unclear. More and more evidences have indicated that miRNAs should been involved in the regulation of IRI and expecially some miRNAs have shown temp-responsiveness for temperature variation. Therefore, the role of miR-122 in mild hypothermia pretreatment after IRI was investigated. METHODS: We established a LO2 cell anoxia-reoxygenation injury model to simulate liver IRI. Five groups of differently pretreated L02 cells were studied. ALT, AST and LDH as well as cell viability were measured. Flow cytometric analysis was used to evaluate the apoptosis. The expression of miR-122 was quantified by qRT-PCR. Insulin-like growth factor 1 receptor (IGF-1R), protein kinase B (p-AKT), AKT, forkhead box O3a (p-FOXO3a) and Caspase3 were examined using western blot analysis. RESULTS: We found that mild hypothermia pretreatment could reduce the hepatocellular injury and induce a significant down-regulation in miR-122 expression after IRI. However, those effects of protection were attenuated by overexpressed miR-122 blockade. We further demonstrated that down-regulation of miR-122 promoted IGF-1R translation and AKT activity, suppressed FOXO3a activity and Caspase3 expression after mild hypothermia pretreatment, which was abrogated by miR-122 mimic. CONCLUSION: Our data clearly demonstrate that mild hypothermia pretreatment can down-regulate miR-122 to protect hepatocytes against IRI through activation IGF-1R/AKT signaling pathway and inhibit cells apoptosis.

Intra-host dynamics of Ebola virus during 2014.

Since 2013, West Africa has encountered the largest Ebola virus (EBOV) disease outbreak on record, and Sierra Leone is the worst-affected country, with nearly half of the infections. By means of next-generation sequencing and phylogeographic analysis, the epidemiology and transmission of EBOV have been well elucidated. However, the intra-host dynamics that mainly reflect viral-host interactions still need to be studied. Here, we show a total of 710 intra-host single nucleotide variations (iSNVs) from deep-sequenced samples from EBOV-infected patients, through a well-tailored bioinformatics pipeline. We present a comprehensive distribution of iSNVs during this outbreak and along the EBOV genome. Analyses of iSNV and its allele frequency reveal that VP40 is the most conserved gene during this outbreak, and thus it would be an ideal therapeutic target. In the co-occurring iSNV network, varied iSNV sites present different selection features. Intriguingly, the T-to-C substitutions at the 3'-UTR of the nucleoprotein (NP; positions 3008 and 3011), observed in many patients, result in the upregulation of the transcription of NP through an Ebola mini-genome reporting system. Additionally, no iSNV enrichment within B-cell epitopes of GP has been observed.

Ebola Virus Outbreak Investigation, Sierra Leone, September 28-November 11, 2014.

During 2014-2015, an outbreak of Ebola virus disease (EVD) swept across parts of West Africa. The China Mobile Laboratory Testing Team was dispatched to support response efforts; during September 28-November 11, 2014, they conducted PCR testing on samples from 1,635 suspected EVD patients. Of those patients, 50.4% were positive, of whom 84.6% lived within a 3-km zone along main roads connecting rural towns and densely populated cities. The median time from symptom onset to testing was 5 days. At testing, 75.7% of the confirmed patients had fever, and 94.1% reported at least 1 gastrointestinal symptom; all symptoms, except rash and hemorrhage, were more frequent in confirmed than nonconfirmed patients. Virus loads were significantly higher in EVD patients with fever, diarrhea, fatigue, or headache. The case-fatality rate was lower among patients 15-44 years of age and with virus loads of <100,000 RNA copies/mL. These findings are key for optimizing EVD control and treatment measures.

Genetic diversity and evolutionary dynamics of Ebola virus in Sierra Leone.

A novel Ebola virus (EBOV) first identified in March 2014 has infected more than 25,000 people in West Africa, resulting in more than 10,000 deaths. Preliminary analyses of genome sequences of 81 EBOV collected from March to June 2014 from Guinea and Sierra Leone suggest that the 2014 EBOV originated from an independent transmission event from its natural reservoir followed by sustained human-to-human infections. It has been reported that the EBOV genome variation might have an effect on the efficacy of sequence-based virus detection and candidate therapeutics. However, only limited viral information has been available since July 2014, when the outbreak entered a rapid growth phase. Here we describe 175 full-length EBOV genome sequences from five severely stricken districts in Sierra Leone from 28 September to 11 November 2014. We found that the 2014 EBOV has become more phylogenetically and genetically diverse from July to November 2014, characterized by the emergence of multiple novel lineages. The substitution rate for the 2014 EBOV was estimated to be 1.23 × 10(-3) substitutions per site per year (95% highest posterior density interval, 1.04 × 10(-3) to 1.41 × 10(-3) substitutions per site per year), approximating to that observed between previous EBOV outbreaks. The sharp increase in genetic diversity of the 2014 EBOV warrants extensive EBOV surveillance in Sierra Leone, Guinea and Liberia to better understand the viral evolution and transmission dynamics of the ongoing outbreak. These data will facilitate the international efforts to develop vaccines and therapeutics.

[Protective effect of grape seed proanthocyanidin on spermatogenesis following testicular torsion/detorsion in mice].

OBJECTIVE: To investigate the protective effect of grape seed proanthocyanidin (GSP) on spermatogenesis following testicular torsion/detorsion in mice. METHODS: Twenty-four healthy male Kunming mice, aged 8 weeks and weighing 25 - 27 g, were randomly divided into a control, a torsion and a treatment group, each containing 8 animals. The unilateral testicular torsion/detorsion model was established in the treatment and torsion groups. Thirty minutes before detorsion, the animals of the treatment group were injected intraperitoneally with 50 mg/kg GSP, and those of the torsion group with normal saline at the same dose, both for 3 days postoperatively. On the 4th day after surgery, ipsilateral orchiectomy were performed to detect histopathological changes, the levels of superoxide dismutase (SOD) and malondialdehyde (MDA), and the apoptotic index (AI) of germ cells in all the mice. RESULTS: Compared with the torsion group, the treated mice showed significantly increased Johnsen score (5.00 +/- 1.85 vs 7.38 +/- 0.92, P < 0.05), seminiferous tubule diameter ([176.50 +/- 1.60]microm vs [178.75 +/- 1.58] microm, P > 0.05), spermatogenic cell layers (3.75 +/- 1.03 vs 5.75 +/- 0.71, P < 0.05) and SOD activity ([29.04 +/- 4.46] U/mg prot vs [52.67 +/- 3.57] U/mg prot, P < 0.05), but remarkably reduced level of MDA ([4.63 +/- 0.05] nmol/mg prot vs [2.91 +/- 0.04] nmol/mg prot, P < 0.05) and AI of germ cells ([40.50 +/- 1.60]% vs [16.25 +/- 1.67] %, P < 0.05). CONCLUSION: Grape seed proanthocyanidin has a protective effect against spermatogenic injury in mice, the mechanisms of which may be related to its actions of scavenging oxygen free radicals, inhibiting lipid peroxidation and improving the antioxidant ability of the body.

Downregulation of cold-inducible RNA-binding protein activates mitogen-activated protein kinases and impairs spermatogenic function in mouse testes.

Cold-inducible RNA-binding protein (CIRP) is an RNA-binding protein that is expressed in normal testes and downregulated after heat stress caused by cryptorchidism, varicocele or environmental temperatures. The purpose of this study was to investigate the functions of CIRP in the testes. We employed RNAi technique to knock down the expression of CIRP in the testes, and performed haematoxylin and eosin staining to evaluate morphological changes following knockdown. Germ cell apoptosis was examined by terminal deoxynucleotidal transferase-mediated dUTP nick end labelling (TUNEL) assay, and mitogen-activated protein kinase (MAPK) signalling pathways were investigated by Western blotting to determine the possible mechanism of apoptosis. We found that using siRNA is a feasible and reliable method for knocking down gene expression in the testes. Compared to controls, the mean seminiferous tubule diameter (MSTD) and the thickness of the germ cell layers decreased following siRNA treatment, whereas the percentage of apoptotic seminiferous tubules increased. The p44/p42, p38 and SAPK/JNK MAPK pathways were activated after downregulation of CIRP. In conclusion, we discovered that downregulation of CIRP resulted in increased germ cell apoptosis, possibly via the activation of the p44/p42, p38 and SAPK/JNK MAPK pathways.

[Expression of AIV subtype H5HA, H7HA and H9HA hemagglutinin gene in Pichia pastoris].

The expression of the hemagglutinins of Avian influenza virus H5 H7and H9 subtypes was studied in this article by Pichia pastoris, one of the eukaryotis expression systems. Three reconstructed expression plasmids and engineering strains, named pPIC9K-H5HA, pPIC9K-H7HA, pPIC9K-H9HA and GS115/pPIC9K-H5HA, GS115/pPIC9K-H7HA, GS115/pPIC9K-H9HA repectively, were obtained. The reconstructed yeast engineering strains were identified by MD and MM plate selecting and PCR. The induced interests proteins were examined by SDS-PAGE and Western-bloting,the results showed that the interest genes were expressed exactly. And this will be helpful in the future study of antigen detection and antibody detection kit, as well in the subunit vaccines developing.