293 EFFECT OF MEDIUM TYPE FOR CULTURE OF ADIPOSE-DERIVED MESENCHYMAL STEM CELLS ON PRE-IMPLANTATION DEVELOPMENT OF CLONED EMBRYOS
Notice bibliographique
Résumé
Mesenchymal stem cells (MSC) have been known as useful donor cells for somatic cell nuclear transfer (SCNT). It has been suggested that the culture condition of donor cells causes different results on preimplantation development of SCNT embryos. In this study, we investigated the patterns of gene expression of adipose-derived mesenchymal stem cells (ad-MSC) in different culture media (DMEM and RKME), and examined the effect of ad-MSC, with the gene expression changed, used as donor cells on the preimplantation development of cloned embryos. Canine ad-MSC were isolated from fat tissue of 3-year-old female beagle and were cultured in DMEM supplemented with 10% fetal bovine serum (MSC-DMEM) and RKME (MSC-MSC) provided from RNL Bio Corp. (Seoul, Korea). Total RNA was extracted from ad-MSC cultured in each culture medium. After synthesising cDNA of each sample, quantitative RT-PCR was done according to the Takara Bio Inc. guidelines and using the 7300 Real Time PCR Cycler System (Applied Biosystems, Carlsbad, CA, USA). The level of all tested gene transcription was normalized to ß-actin expression levels. The relative quantification of gene expression was analysed by the 2–??Ct method. The data from all experiments were analysed by Student’s t-test using a statistical analysis GraphPad Prism 4.02 (GraphPad Software Inc., San Diego, CA, USA). Significance was determined at P < 0.05. The stemness, the reprogramming-related gene expression level of donor cells of MSC-DMEM and MSC-MSC were compared. In order to confirm the effect of MSC cultured in 2 different culture media on somatic cell nuclear transfer, we performed interspecies somatic cell nuclear transfer (iSCNT). The enucleated bovine oocytes were injected, respectively, with donor cells of MSC-DMEM and MSC-MSC, and were fused by electrofusion. The iSCNT embryos were cultured in modified SOF at 38.5°C for 7 days in an atmosphere of 5% CO2 and 5% O2, and the developmental ability of iSCNT embryos was observed under the microscope. The MSC-MSC contained a significantly higher amount of Sox2, Nanog, Oct4, Stella, HDAC1, DNMT1, and MeCP2 than the MSC-DMEM, whereas the amount of Rex1 was not different in either MSC-MSC or MSC-DMEM. In the development ability of iSCNT embryos, MSC-DMEM embryos resulted in a 16-cell embryo formation rate that was higher than that of MSC-MSC embryos (9.09 and 5.30%, respectively; P < 0.05). However, the blastocyst formation rate was not different between MSC-DMEM embryos and MSC-MSC embryos (4.5 and 3.2%, respectively; P > 0.05). These results demonstrate that the gene expression of ad-MSC can be modified, by culture media, into a state where reprogramming is easily done. Even so, ad-MSC with gene expression changed by culture medium did not influence the developmental ability of blastocysts. In conclusion, the alteration of gene-related stemness and reprogramming in canine ad-MSC would not be able to effectively control reprogramming in SCNT. This study was supported by RDA (#PJ0089752012), RNL Bio (#550-20120006), IPET (#311062-04-1-SB010), Research Institute for Veterinary Science, and Nestlé Purina Korea.
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Comment cette classification a été obtenuedéplier
Prédiction distillée sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Apprise à partir de 10 348 étiquettes directes de Codex et de 10 348 étiquettes directes de Gemma. Le mode candidate est l'union des têtes enseignantes seuillées; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont ni des étiquettes humaines ni des étiquettes directes de modèles de pointe.
Scores Codex et Gemma par catégorie
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,002 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,001 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,000 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule tête enseignante, pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».