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Background: Adipose stem cell‐derived exosomes (ADSC‐EXO) and botulinum toxin type A (BTX‐A) individually showed a therapeutic effect on skin wound repair. Aims: This study investigated their synergistic effect on promoting skin wound healing in vitro and in vivo and the underlying molecular events. Methods: ADSCs were isolated from Sprague–Dawley (SD) rats to obtain ADSC‐EXO by ultrafiltration and ultracentrifugation and were confirmed using nanoparticle tracking analysis and transmission electron microscopy. Human skin fibroblasts (HSF) were cultured and treated with or without ADSC‐EXO, BTX‐A, or their combination. Changes in cell phenotypes and protein expression were analyzed using different in vitro assays, and a rat skin wound model was used to assess their in vivo effects. Results: The isolated ADSC‐EXO from primarily cultured ADSCs had a circular vesicle shape with a 30–180 nm diameter. Treatment of HSF with ADSC‐EXO and/or BTX‐A significantly accelerated HSF migration in vitro and skin wound healing in a rat model. Moreover, ADSC‐EXO plus BTX‐A treatment dramatically induced VEGFA expression but reduced COL III and COL I levels in vivo. ADSC‐EXO and/or BTX‐A treatment significantly upregulated TGF‐β3 expression on Day 16 after surgery but downregulated TGF‐β1 expression, suggesting that ADSC‐EXO plus BTX‐A promoted skin wound healing and reduced inflammatory cell infiltration. Conclusions: The ADSC‐EXO plus BTX‐A treatment demonstrated a synergistic effect on skin wound healing through upregulation of VEGF expression and the TGF‐β3/TGF‐β1 and COL III/COL I ratio. [ABSTRACT FROM AUTHOR]

RNA polymerase III is an essential enzyme in eukaryotes for synthesis of tRNA, 5S rRNA, and other small nuclear and cytoplasmic RNAs. Thus, RNA polymerase III promoters are often used in small hairpin RNA (shRNA) expression. In this study, the porcine H1, U6, and 7SK RNA polymerase III type promoters were cloned into a pcDNA3.1(+) expression vector containing a shRNA sequence targeting enhanced green fluorescent protein (EGFP). PK and DF-1 cells were cotransfected with the construction of recombinant interference expression vector and the EGFP expression vector, pEGFP-N1. The average fluorescence intensity of EGFP in transfected cells was measured by fluorescence microscopy and flow cytometry. Real-time PCR was used to detect expressed shRNAs and the relative expression of EGFP, to confirm the activity of the promoters. The results showed that the activity of porcine 7SK promoter is stronger than the U6 promoter, which is in turn stronger than porcine H1. While the high levels of expression of the U6 and 7SK promoters saturate the shRNAs level in the host cell, which can cause cytotoxicity and tissue damage. Therefore, porcine H1 promoter is effective for expression of shRNA, and may be an excellent tool to knockdown gene expression in pigs for functional genomics studies. The results also lay a foundation for the development of porcine RNAi technology and genetically modified porcine research. [ABSTRACT FROM AUTHOR]

The generation of transgenic chickens is of both biomedical and agricultural significance, and recently chicken transgenesis technology has been greatly advanced. However, major issues still exist in the efficient production of transgenic chickens. This study was designed to optimize the production of enhanced green fluorescence protein (EGFP)-transgenic broilers, including egg windowing at the blunt end (air cell) of egg, and the direct transfection of circulating primordial germ cells by microinjection of the Tol2 plasmid-liposome complex into the early embryonic dorsal aorta. For egg windowing, we discovered that proper manipulation of the inner shell membrane at the blunt end could improve the rate of producing G0 transgenic roosters. From 27 G0 roosters, we successfully collected semen with EGFP-positive sperms from 16 and 19 roosters after direct fluorescence observation and fluorescence-activated cell sorting analyses (13 detected by both methods), respectively. After artificial insemination using the G0 rooster with the highest number of EGFP fluorescent sperm, one G1 EGFP transgenic broiler (1/81, 1.23%) was generated. Our results indicate that appropriate egg windowing and screening of potentially transgene-positive roosters can improve the production of germline-transmitted transgenic birds. [ABSTRACT FROM AUTHOR]

Background: Id (inhibitor of differentiation/DNA binding)-1 and -3 are involved in neoangiogenesis; they antagonize basic helix-loop-helix proteins, inhibit differentiation, and enhance cell proliferation. The aim of this study was to investigate Id-1 and -3 expression in gastric tumors and their clinical relevance in gastric cancer. Materials and Methods: We investigated Id-1 and Id-3 expression in gastric cancer samples by immunohistochemistry and Western blotting, and further analyzed the relationship between expression of Id-1 and Id-3 and clinicopathologic characteristics. Results: Expression of Id-1 and -3 was found significantly more often in gastric cancers than in matched adjacent nonmalignant tissues. Cancer samples with poor or moderate histologic differentiation showed significantly stronger Id-1 and -3 expression than cancer samples with high differentiation. In cancer samples, strong or moderate expression of Id-3, but not Id-1, was a strong independent predictor for shorter overall survival in multivariate analysis. Conclusions: The level of Id-1 and -3 protein expression was associated with the malignant potential of gastric tumors. In cancer samples, stronger Id-1 and -3 expression is associated with poor differentiation and more aggressive behavior of tumor cells, resulting in poor clinical outcome. Consequently, Id-3 might be used to independently predict survival of patients with gastric cancer. [Copyright &y& Elsevier]

Inactivated whole avian influenza virus (AIV) vaccine provides protection against homologous haemagglutinin (HA) subtype virus, but poor protection against a heterologous HA virus. Moreover, it induces chickens to produce antibodies to cross-reactive antigens, especially nucleoprotein, which is limits AIV serological surveillance. In this study, a recombinant fowlpox virus co-expressing HA (H5 subtype) and NA (N1 subtype) genes of AIV was evaluated for its ability to protect chickens against intramuscular challenge with a lethal dose of highly pathogenic (HP) AIV. Susceptible chickens were also vaccinated by wing-web puncture with the parent fowlpox vaccine virus. Following challenge 4 weeks later with HPAIV, all chickens vaccinated with recombinant virus were protected, while the chickens vaccinated with either the unaltered parent fowlpox vaccine virus or unvaccinated controls experienced 100% mortality following challenge. This protection was accompanied by the high levels of specific antibody to the respective components of the recombinant vaccine. The above results showed that rFPV-HA-NA could be a potential vaccine to replace current inactivated vaccines for preventing AI. [ABSTRACT FROM AUTHOR]