Protective efficacy of anti-neuraminidase monoclonal antibodies against H7N9 influenza virus infection.

The H7N9 influenza virus has been circulating in China for more than six years. The neuraminidase (NA) has gained great concern for the development of antiviral drugs, therapeutic antibodies, and new vaccines. In this study, we screened seven mouse monoclonal antibodies (mAbs) and compared their protective effects against H7N9 influenza virus. The epitope mapping from escape mutants showed that all the seven mAbs could bind to the head region of the N9 NA close to the enzyme activity sites, and four key sites of N9 NA were reported for the first time. The mAbs D3 and 7H2 could simultaneously inhibit the cleavage of the sialic acid of fetuin protein with large molecular weight and NA-XTD with small molecule weight in the NA inhibition experiment, prevent the formation of virus plaque at a low concentration, and effectively protect the mice from the challenge of the lethal dose of H7N9 virus.

Enhancing Immune Response and Heterosubtypic Protection Ability of Inactivated H7N9 Vaccine by Using STING Agonist as a Mucosal Adjuvant.

Influenza vaccines for H7N9 subtype have shown low immunogenicity in human clinical trials. Using novel adjuvants might represent the optimal available option in vaccine development. In this study, we demonstrated that the using of the STING agonist cGAMP as a mucosal adjuvant is effective in enhancing humoral, cellular and mucosal immune responses of whole virus, inactivated H7N9 vaccine in mice. A single dose of immunization was able to completely protect mice against a high lethal doses of homologous virus challenge with an significant dose-sparing effect. We also found that intranasal co-administration of H7N9 vaccine with cGAMP could provide effective cross protection against H1N1, H3N2, and H9N2 influenza virus. Furthermore, cGAMP induced significantly higher nucleoprotein specific CD4+ and CD8+ T cells responses in immunized mice, as well as upregulated the IFN-γ and Granzyme B expression in the lung tissue of mice in the early stages post a heterosubtypic virus challenge. These results indicated that STING agonist cGAMP was expected to be an effective mucosal immune adjuvant for pre-pandemic vaccines such as H7N9 vaccines, and the cGAMP combined nasal inactivated influenza vaccine will also be a promising strategy for development of broad-spectrum influenza vaccines.

Flavonoid ingredients of Ginkgo biloba leaf extract regulate lipid metabolism through Sp1-mediated carnitine palmitoyltranferase 1A up-regulation.

BACKGROUND: Lipid accumulation is the primary evidence of non-alcoholic fatty liver disease (NAFLD). Ginkgo biloba extract (GBE) and its flavonoid ingredients (quercetin, kaempferol, and isorhamnetin) could lessen the lipid accumulation associated with up-regulation of the rate-limiting enzyme, carnitine palmitoyltransferase 1A (CPT1A), in the ß-oxidation of long-chain fatty acids. In this study, we investigated the mechanisms by which GBE and its flavonoids induced expression of CPT1A. RESULTS: CPT1A inhibition with RNAi resulted in triglyceride accumulation in HepG2 cells. Through deletion and mutation analysis of CPT1A's promoter combined with electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) experiments, the CPT1A promoter region (-50 to -5 nt) was determined to contain two putative Sp1 binding sites, namely Sp1a and Sp1b, which might act as the GBE regulation response DNA element. Sp1 might be induced to transfer from cytoplasma to nucleus to bind the promoter region of -50 to -5 nt by GBE. The regulatory effects of GBE on CPT1A were also verified on the flavonoid ingredients quercetin, kaempferol, and isorhamnetin. CONCLUSION: Sp1 was crucial in regulating CPT1A expression with GBE and its flavonoid ingredients, and the -50 to -5 nt region of CPT1A promoter played important roles in Sp1 binding.

A pipeline with multiplex reverse transcription polymerase chain reaction and microarray for screening of chromosomal translocations in leukemia.

Chromosome rearrangements and fusion genes present major portion of leukemogenesis and contribute to leukemic subtypes. It is practical and helpful to detect the fusion genes in clinic diagnosis of leukemia. Present application of reverse transcription polymerase chain reaction (RT-PCR) method to detect the fusion gene transcripts is effective, but time- and labor-consuming. To set up a simple and rapid system, we established a method that combined multiplex RT-PCR and microarray. We selected 15 clinically most frequently observed chromosomal rearrangements generating more than 50 fusion gene variants. Chimeric reverse primers and chimeric PCR primers containing both gene-specific and universal sequences were applied in the procedure of multiplex RT-PCR, and then the PCR products hybridized with a designed microarray. With this approach, among 200 clinic samples, 63 samples were detected to have gene rearrangements. All the detected fusion genes positive and negative were validated with RT-PCR and Sanger sequencing. Our data suggested that the RT-PCR-microarray pipeline could screen 15 partner gene pairs simultaneously at the same accuracy of the fusion gene detection with regular RT-PCR. The pipeline showed effectiveness in multiple fusion genes screening in clinic samples.