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The objective of this study was to determine the effect of exogenous mitochondria obtained from granulosa cells on the development of bovine embryos in vitro. We classified cumulus oocyte complexes (COCs) as good (G)- and poor (P)-quality oocytes based on cytoplasmic appearance and cumulus characteristics, and assessed mtDNA copy numbers in the G and P oocytes with real-time polymerase chain reaction (PCR). The mitochondria were isolated by fractionation and suspended in mitochondria injection buffer (MIB). Part one of the experiment consisted of the following treatments: (1) G-oocytes + sperm, (2) P-oocytes + mitochondria + MIB + sperm, (3) P-oocytes + MIB + sperm, and (4) P-oocytes + sperm. In part 2, oocytes were parthenogenetically activated. The treatments were: (1) G-oocytes, (2) P-oocytes + mitochondria + MIB, (3) P-oocytes + MIB, and (4) P-oocytes alone. The results indicated a significant difference in mtDNA copy number between G (361 113 +/- 147 114) and P (198 293 +/- 174 178) oocytes (p < 0.01). The rates of morula, blastocyst, and hatched blastocysts derived from P-oocytes + mitochondria were similar to those of G-oocytes, but significantly higher than P-oocytes without exogenous mitochondria in both the ICSI and parthenogenetic activation experiments. We found no difference in blastomere numbers between G-oocytes and P-oocytes + mitochondria in either experiment, but blastomere numbers in these two groups were significantly higher than in P-oocyte groups without exogenous mitochondria. These data suggest that mtDNA content is very important for early embryo development. Furthermore, the transfer of mitochondria from the same breed may improve embryo quality during preimplantation development.

Curcumin, a natural polyphenol antioxidant extracted from the root of turmeric (Curcuma longa), can induce apoptosis and DNA demethylation in several types of cancer cells. However, the mechanism of its anticancer potentials and DNA demethylation effects and the potential relationships between these outcomes have not been clearly elucidated. In the present study, the effects of curcumin on the proliferation, colony formation, and migration of human gastric cancer cells (hGCCs) were explored. Reactive oxygen species (ROS) levels, mitochondrial damage, DNA damage, and apoptosis of curcumin-treated hGCCs were analyzed. Changes in the expression of several genes related to DNA damage repair, the p53 pathway, cell cycle, and DNA methylation following curcumin treatment were also evaluated. We observed that curcumin inhibited the proliferation, colony formation, and migration of hGCCs in a dose- and time-dependent fashion. A high concentration of curcumin elevated ROS levels and triggered mitochondrial damage, DNA damage, and apoptosis of hGCCs. Further, curcumin-induced DNA demethylation of hGCCs was mediated by the damaged DNA repair-p53-p21/GADD45A-cyclin/CDK-Rb/E2F-DNMT1 axis. We propose that the anticancer effect of curcumin could largely be attributed to its prooxidative effect at high concentrations and ROS elevation in cancer cells. Moreover, we present a novel mechanism by which curcumin induces DNA demethylation of hGCCs, suggesting the need to further investigate the demethylation mechanisms of other DNA hypomethylating drugs.

ABSTRACT. Bovine mammary epithelial stem cells (MESCs) are very important in agricultural production and bioengineering. In the present study, we compared different isolation and culture methods for MESCs and observed their growth and differentiation characteristics. MESCs have an extremely weak proliferation capacity, and it is very difficult to obtain and prolong subculture of a bovine mammary epithelial stem cell line. We obtained some multipotent MESC aggregates that looked like spherical colonies. These colonies were only derived from suspension culture and were induced to differentiate into epithelial-like cells, myoepithelial-like cells and secretory cells and to establish a ductal-like structure. In contrast, MESCs cultured in adherent culture displayed low morphogenetic competence and only differentiated into epithelial-like cells. MESCs are often identified by testing their differentiation in vivo; however, herein, we have demonstrated the in vitro differentiation potential of bovine MESCs. In our study, beta 1-integrin and alpha 6-integrin which are expressed by human epidermal stem cells, were found in bovine, which shows that bovine MESCs share the same molecular signature as human MESCs.