Downregulation of miR-210 expression inhibits proliferation, induces apoptosis and enhances radiosensitivity in hypoxic human hepatoma cells in vitro

Hypoxia is a common feature of solid tumors and an important contributor to tumor radioresistance. miR-210 is the most consistently and robustly induced microRNA under hypoxia in different types of tumor cells and normal cells. In the present study, to explore the feasibility of miR-210 as an effective therapeutic target, lentiviral-mediated anti-sense miR-210 gene transfer technique was employed to downregulate miR-210 expression in hypoxic human hepatoma SMMC-7721, HepG2 and HuH7 cells, and phenotypic changes of which were analyzed. Hypoxia led to an increased hypoxia inducible factor-1α (HIF-1α) and miR-210 expression and cell arrest in the G(0)/G(1) phase in all cell lines. miR-210 downregulation significantly suppressed cell viability, induced cell arrest in the G(0)/G(1) phase, increased apoptotic rate and enhanced radiosensitivity in hypoxic human hepatoma cells. Moreover, apoptosis-inducing factor, mitochondrion-associated, 3 (AIFM3) was identified as a direct target gene of miR-210. AIFM3 downregulation by siRNA attenuated radiation induced apoptosis in miR-210 downregulated hypoxic human hepatoma cells. Taken together, these data suggest that miR-210 might be a potential therapeutic target and specific inhibition of miR-210 expression in combination with radiotherapy might be expected to exert strong anti-tumor effect on hypoxic human hepatoma cells.     

A novel porcine circovirus type 2a strain, 10JS-2, with eleven-nucleotide insertions in the origin of genome replication

Here, we report a novel porcine circovirus type 2a (PCV2a) strain with 11 nucleotides (nt) inserted in the origin of genome replication (Ori). This is the first report of a PCV2a strain with nucleotide insertion in Ori. Our study will help further epidemiological studies and extend our knowledge of evolutionary characteristics of PCV2.     

Complete Genome Sequence of a Porcine Orthoreovirus from Southern China

Porcine orthoreoviruses belong to the family Reoviridae and cause mainly mild enteritis in piglets. We present here the complete genome sequence of a novel porcine orthoreovirus strain (GD-1) isolated from a piglet in southern China. Our data will facilitate future investigations of the molecular characteristics and epidemiology of porcine orthoreoviruses.     

Serotype-specific neutralizing antibody epitopes of human adenovirus type 3 (HAdV-3) and HAdV-7 reside in multiple hexon hypervariable regions

Human adenovirus types 3 and 7 (HAdV-3 and HAdV-7) occur epidemically and contribute greatly to respiratory diseases, but there is no currently available licensed recombinant HAdV-3/HAdV-7 bivalent vaccine. Identification of serotype-specific neutralizing antibody (NAb) epitopes for HAdV-3 and HAdV-7 will be beneficial for development of recombinant HAdV-3/HAdV-7 bivalent vaccines. In this study, four NAb epitopes within hexon hypervariable regions (HVRs) were predicted for HAdV-3 and HAdV-7, respectively, by using bioinformatics. Eight hexon chimeric adenovirus vectors with the alternation of only one predicted neutralizing epitope were constructed. Further in vitro and in vivo neutralization assays indicated that E2 (residing in HVR2) and E3 (residing in HVR5) are NAb epitopes for HAdV-7, and E3 plays a more important role in generating NAb responses. Cross-neutralization assays indicated that all four predicted epitopes, R1 to R4, are NAb epitopes for HAdV-3, and R1 (residing in HVR1) plays the most important role in generating NAb responses. Humoral immune responses elicited by the recombinant rAdH7R1 (containing the R1 epitope) were significantly and durably suppressed by HAdV-3-specific NAbs. Surprisingly, the rAdΔE3GFP-specific neutralizing epitope responses induced by rAdMHE3 (R3 replaced by E3) and rAdMHE4 (R4 replaced by E4) were weaker than those of rAdMHE1 (R1 replaced by E1) or rAdMHE2 (R2 relaced by E2) in vitro and in vivo. Furthermore, rAdMHE4 replicated more slowly in HEp-2 cells, and the final yield was about 10-fold lower than that of rAdΔE3GFP. The current findings contribute not only to the development of new adenovirus vaccine candidates, but also to the construction of new gene delivery vectors.     

Complete Genome Sequence of a Vero Cell-Adapted Isolate of Porcine Epidemic Diarrhea Virus in Eastern China

In early 2012, a widespread porcine epidemic diarrhea virus (PEDV) occurred in eastern China. A cell-adapted isolate, SD-M, was at the four-passage level of virulent field strain SD, which was isolated from a 2-day-old dead suckling piglet that had suffered from severe diarrhea in Shandong Province, China. We report here the complete genome sequence of SD-M. This sequence will promote a better understanding of the molecular pathogenesis of PEDV.     

Ouabain-stimulated trafficking regulation of the Na/K-ATPase and NHE3 in renal proximal tubule cells

We have demonstrated that ouabain regulates protein trafficking of the Na/K-ATPase α1 subunit and NHE3 (Na/H exchanger, isoform 3) via ouabain-activated Na/K-ATPase signaling in porcine LLC-PK1 cells. To investigate whether this mechanism is species-specific, ouabain-induced regulation of the α1 subunit and NHE3 as well as transcellular (22)Na(+) transport were compared in three renal proximal tubular cell lines (human HK-2, porcine LLC-PK1, and AAC-19 originated from LLC-PK1 in which the pig α1 was replaced by ouabain-resistant rat α1). Ouabain-induced inhibition of transcellular (22)Na(+) transport is due to an ouabain-induced redistribution of the α1 subunit and NHE3. In LLC-PK1 cells, ouabain also inhibited the endocytic recycling of internalized NHE3, but has no significant effect on recycling of endocytosed α1 subunit. These data indicated that the ouabain-induced redistribution of the α1 subunit and NHE3 is not a species-specific phenomenon, and ouabain-activated Na/K-ATPase signaling influences NHE3 regulation.     

The different expression of immune-related cytokine genes in response to velogenic and lentogenic Newcastle disease viruses infection in chicken peripheral blood

Newcastle disease virus (NDV) is an important pathogen hazardous to poultry industry, and the pathogenicity of NDV strains varies with different virulence. Peripheral blood serves as an important producer and carrier of viruses and cytokines in NDV infection. In order to explore the difference of cytokine expression in the peripheral blood between velogenic strain and lentogenic strain infection, NDV virulent strain F48E9 and vaccine strain Lasota were used to infect specific-pathogen-free (SPF) chickens separately, and peripheral blood was collected on 0, 3, 7, 10, 14, and 21 days post-infection (d.p.i.). Real-time PCR was then used to detect the expression of six kinds of immune-related cytokine genes. For the F48E9 group, a sharp increase of the expression of interferon-alpha (IFN-α), interferon-gamma (IFN-γ), interleukin-16 and IL-18 was observed on 3 d.p.i. before the NDV blood peak (7 d.p.i.), followed by a rapid decline to the level lower than that of control group, then the expression of IFN-α increased slowly and reached or exceeded the level of control group in the later phase of the infection, while the expression of IFN-γ, IL-16, and IL-18 fluctuated at the level of control group for the rest of study period. The increase of IL-2 expression was not obvious, and no increase of IL-15 expression was noted. For the Lasota (vaccine) group, the picture was quite different, a sharp increase of IFN-γ (but not IFN-α), IL-2 was observed on 7 d.p.i. before the NDV blood peak (10 d.p.i.). On the contrary, there was no dramatic increase of IL-16 and IL-18. Interestingly, in contrast to the F48E9 group, there was an increase of IL-15 on day 10 d.p.i., but it remained modest. There was also an increase of IFN-α on day 21 d.p.i. Our results revealed that infection with NDV strains of different virulence was associated with distinct cytokine expression patterns in peripheral blood, modulation of cytokine responses may play a key role in mediation of NDV pathogenesis.     

Identification of a new gene encoding 5-enolpyruvylshikimate-3-phosphate synthase using genomic library construction strategy

Applying the genomic library construction strategy and colony screening, a new aroA gene encoding 5-enolpyruvylshikimate-3-phosphate synthase has been identified, cloned and overexpressed in Escherichia coli, and the enzyme was purified to homogeneity. Kinetic analysis of the AroA _{P.fluorescens} indicated that the full-length enzyme exhibits 10-fold increased IC50 and an approximately 38-fold increased K _i for glyphosate compared to those of the AroA _E.coli , while retaining high affinity for the substrate phosphoenolpyruvate. Furthermore, we have transformed the new aroA _{P.fluorescens} gene into Arabidopsis thaliana via a floral dip method, and demonstrated that transgenic A. thaliana plants exhibit significant glyphosate resistance when compared with the wild type.