To preserve the superior genetic resources and restore the endangered species, Somatic cell nuclear transfer (SCNT) has been used widely. In Korea, the research of dog cloning has made outstanding achievements including the production of the world`s first cloned dog. Sapsaree (Sapsalgae), the representative dog of Gyeongsan-si was designated as a Korea natural monument (No. 368). This male dog used in this study has azoospermia due to unknown cause. In this study, the aim was to confirm the cause of infertility in the cell donor dog and to evaluate the reproduction potential of dog cloning using infertile male dog by SCNT. First, to confirm the infertility of the cell donor dog, the reproductive history and the testis were evaluated. The breeding histology was not recorded in individual document. In histopathology, the Sertoli cell tumor was confirmed in biopsy of the cell donor dog after death. But, these tumors are predominantly in older dogs. Second, we produced the cloned dogs with the somatic cells of the infertile dog and the appearance was similar with the cell donor dog. Also, microsatellite analysis confirmed the genetic relationship between the cell donor and clone dogs. Third, the potential breeding capacity of the cloned dog was confirmed. In T4 assay, the normal dog (same age with cloned dogs), cell donor dog, and cloned dogs was investigated. The cell donor dog with azoospermia had very low T4 level, and cloned dogs showed higher level of T4 than normal dogs. In CASA, There was no significant difference in sperm motor ability between normal dogs and cloned dogs. As a result, cloned dogs produced by SCNT had no problem regarding the reproductive function of the testis. In AI experiment, the semen of clone dogs was used to fertilize a natural female bitch and was diagnosed pregnancy by ultrasonography. In total, 7 puppies were born by normal delivery (male: 3, female: 4). In conclusion, this study confirmed that the reproduction problem of non-genetic infertility can generate a normal descendant by SCNT. Also, the first successful research to restore infertile dogs was completed. Furthermore, SCNT would be useful for the restoration of endangered species and application of superior traits.

Cell transplantation therapy using adult stem cells has recently been identified as a potential treatment for spinal cord injury (SCI). But, recovery after traumatic SCI is very limited. As dogs are physiologically much more similar to human compared with other traditional mammalian models in disease presentation and clinical responses, a number of researches demonstrated canis familiaris is a suitable model for human diseases. This study investigated the effect of transplantation of canine Mesenchymal Stem Cells (cMSC) and neural-induced cMSC (nMSC) to understand how these cells improve neurological function in canine SCI model. The differentiation of cMSC into neural precursor cells was induced in dulbecco’s modified eagle’s medium supplemented with N2-supplement, dibutyryl cyclic adenosine monophosphate, and butylated hydroxyanisole. SCI was induced between T1 and T2 by surgical hemi-section in adult dogs, and then assigned to two groups according to the applied cell types (cMSC vs nMSC). Pelleted cMSC or nMSC were transplanted directly into the injured site after SCI, respectively. Analysis of motor function after transplantation was evaluated by modified Olby score. Magnetic resonance imaging (MRI), histological and immunohistichemical analysis were also performed. Functional recovery in group of cMSC was increasing gradually after transplantation and was higher than nMSC. In MRI, we could not confirm any difference between the cMSC and nMSC experimental groups. Immunohistochemically, beta3-tubuline and nestin were observed in injury site of two experimental groups with the expression level close to non-injured groups. Transplantation of mesenchymal stem cells could promote neuronal reconstruction and repair motor function in SCI. These showed mesenchymal stem cells could be a great candidate as a therapeutic tools in degeneration disease, and dogs could be used to explore human regenerative medicine as a promising animal model. This research was supported by iPET (Grants 110056032CG000), Ministry for Food, Agriculture, Forestry and Fisheries, Republic of Korea.

Acteoside (verbascoside) is a typical phenylethanoid glycoside, extracted from various plants. It has various biological functions such as anti-oxidant, anti-inflammation, and anti-hypertension. Specially, it was powerful anti-oxidants either by direct scavenging of reactive oxygen and nitrogen species, or by acting as chain-breaking peroxyl radical scavengers. We examined the role of acteoside in IVM medium on the morphological progress of meiosis, developmental competence, and ROS in porcine oocytes. And we investigated effect of acteoside on the oocytes condition represented by cytoplasmic maturation by homogeneous distribution and formation of cytoplasmic organelles and regulation of apoptosis-related genes. The selected COCs were cultured in TCM-199 with various concentration of acteoside: 0 (control), 10, 30, and 50 μM. After 22 h of maturation with hormones, the oocytes were washed twice in a fresh maturation medium before being cultured in hormone-free medium for additional 22 h. The oocytes maturation rates of supplemented with acteoside were no significantly different compared with control group (71.13, 75.96, 72.95 and 73.68%, respectively). Level of ROS was significantly decreased in acteoside treated group. Furthermore, the parthenogenetic blastocyst rate was significantly improved in 10 μM acteoside treated group compared with control group (40.03 vs. 22.95%). During IVM, 10 μM acteoside treated oocytes showed that the mitochondria and lipid droplet were smaller and homogeneous distribution in cytoplasm compare with non-treated control oocytes. And reverse transcription polymerase chain reaction (RT-PCR) witarthenogenetic blstocysts revealed that acteoside increased the anti-apoptoticgenes, otherwise reibued pro-apoptotic genes. In conclusion, our results represents that addition of acteoside to the IVM medium has a beneficial effect in physiology of porcine oocytes such as viability and activation, providing a improved method for porcine oocytes in vitro.