200 TELOMERASE ACTIVITY MODIFICATION WITH RESVERATROL IN CANINE ADIPOSE-DERIVED MESENCHYMAL STEM CELLS

Abstract

It has been established that the telomerase activity of donor cells is causally linked to the reprogramming efficiency, which includes somatic cell nuclear transfer (SCNT). Among the many cell source as donor cells of SCNT, canine adipose-derived stem cells (Ad-MSCs), a form of adult stem cells, are donor cells that have been recently used. Resveratrol (trans-3,5,4′-trihydroxystilbene), a polyphenolphytoalexin, possesses diverse biochemical and physiological action, including antiplatelet and reducing cellular senescence via telomerase activity modification. However, appropriate dose and time of resveratrol for telomerase activity modification in Ad-MSCs have not been investigated. In addition, unlike most normal somatic cells which have negative telomerase activities; low to moderate levels of the enzyme in mesenchymal stem cells have been described. In the present study, we compared the cellular telomerase activity of Ad-MSCs according to the dose of resveratrol and passages of Ad-MSCs. Cells were isolated from collected adipose derived tissues of beagle at age 7 and cultured in RCME-P provided from RNL Bio incorporation. For identification of telomerase activity in ad-MSCs, adult fibroblasts derived from same dog were used as negative control. After 2 days of cultivation, Ad-MSCs were treated with 2 μM, 10 μM, or 25 μM resveratrol or without resveratrol at 39°C for 24 h in a humidified atmosphere of 5% CO₂ in air. Ad-MSCs with passage at 1, 4, and 7 were used for analysis. Telomerase activity was measured by the telomeric repeat amplification protocol assay. Statistical analysis was performed by the two-way ANOVA. P < 0.05 was considered as showing a statistically significant difference between means. It revealed that Ad-MSCs have telomerase activity significantly higher than those shown in fibroblasts (negative control). Resveratrol increased telomerase activity, with maximal increase at 10, 25 μM at passage of 1, 4 (P < 0.05). However, increase of telomerase activity in Ad-MSCs treated with 10, 25 μM at passage 7 was not shown and the telomerase activities of Ad-MSCs at passage 7 were lower than those of Ad-MSCs at passage 1 and 4 (P < 0.05). In conclusion, the telomerase activity was detectable in canine Ad-MSCs, which may suggest that canine Ad-MSCs have similar telomere biology compared to that of other adult stem cells. Furthermore, resveratrol can enhance activation of telomerase activity with dose dependent increase. Further studies are warranted on efficiency of establishing a stable donor cells for SCNT using Ad-MSCs treated with resveratrol.

This study was supported by R&E (#550–20130027), IPET (#311062–04–2-SB010), RNL Bio (#550–20130013), RDA (PJ008975022013), the Research Institute for Veterinary Science, and TS Corporation.