Long-term health and germline transmission in transgenic cattle following transposon-mediated gene transfer.

BACKGROUND: Transposon-mediated, non-viral gene delivery is a powerful tool for generating stable cell lines and transgenic animals. However, as multi-copy insertion is the preferred integration pattern, there is the potential for uncontrolled changes in endogenous gene expression and detrimental effects in cells or animals. Our group has previously reported on the generation of several transgenic cattle by using microinjection of the Sleeping Beauty (SB) and PiggyBac (PB) transposons and seeks to explore the long-term effects of this technology on cattle. RESULTS: Transgenic cattle, one female (SNU-SB-1) and one male (SNU-PB-1), reached over 36 months of age with no significant health issues and normal blood parameters. The detection of transgene integration and fluorescent signal in oocytes and sperm suggested the capacity for germline transmission in both of the founder animals. After natural breeding, the founder transgenic cow delivered a male calf and secreted milk containing fluorescent transgenic proteins. The calf expressed green fluorescent protein in primary cells from ear skin, with no significant change in overall genomic stability and blood parameters. Three sites of transgene integration were identified by next-generation sequencing of the calf's genome. CONCLUSIONS: Overall, these data demonstrate that transposon-mediated transgenesis can be applied to cattle without being detrimental to their long-term genomic stability or general health. We further suggest that this technology may be usefully applied in other fields, such as the generation of transgenic animal models.

The result of surgical treatment of medial epicondylitis: analysis with more than a 5-year follow-up.

BACKGROUND: Angiofibroblastic changes of a musculotendinous origin at the medial epicondyle characterize medial epicondylitis of the elbow. Although nonsurgical treatment is the primary approach for medial epicondylitis, surgical treatment should be considered when conservative therapy fails. This study reports the results of surgical treatment of medial epicondylitis monitored for more than 5 years. METHODS: This study included 55 patients with 63 cases of medial epicondylitis between 2000 and 2010. The conservative treatment periods lasted for a minimum of 1 year, and steroid injections were administered more than twice before surgery. One surgeon conducted the surgical procedures. The Nirschl and Pettrone grades, visual analog scale (VAS) scores, Disabilities of the Arm, Shoulder and Hand (DASH) scores, Mayo Elbow Performance scores, and grip strengths were analyzed. Statistical analyses were performed using paired t tests. RESULTS: The mean VAS score improved from 8.5 to 2.4 (P <.001). Nirschl and Pettrone grades rated 43% (27 elbows) as excellent and 51% (32 elbows) as good. The Mayo Elbow Performance scores improved from 72 to 88 (P <.001) and DASH scores from 57 to 23 (P <.001). The mean grip strength of the affected side improved from 30 to 43 lb (P <.001). The mean time required to return to work and exercise was 2.8 months and 4.8 months, respectively. One case of heterotrophic ossification, which had no functional instability afterward, was seen. CONCLUSION: The results indicate that surgical treatment of medial epicondylitis could be an effective and safe treatment when conservative treatment fails.

Efficient generation of transgenic cattle using the DNA transposon and their analysis by next-generation sequencing.

Here, we efficiently generated transgenic cattle using two transposon systems (Sleeping Beauty and Piggybac) and their genomes were analyzed by next-generation sequencing (NGS). Blastocysts derived from microinjection of DNA transposons were selected and transferred into recipient cows. Nine transgenic cattle have been generated and grown-up to date without any health issues except two. Some of them expressed strong fluorescence and the transgene in the oocytes from a superovulating one were detected by PCR and sequencing. To investigate genomic variants by the transgene transposition, whole genomic DNA were analyzed by NGS. We found that preferred transposable integration (TA or TTAA) was identified in their genome. Even though multi-copies (i.e. fifteen) were confirmed, there was no significant difference in genome instabilities. In conclusion, we demonstrated that transgenic cattle using the DNA transposon system could be efficiently generated, and all those animals could be a valuable resource for agriculture and veterinary science.