201 SUPPRESSION OF Suz12 IN BOVINE PREIMPLANTATION EMBRYOS VIA CYTOPLASMIC SMALL INTERFERING RNA INJECTION
Notice bibliographique
Résumé
The proper removal of gametic epigenetic marks and coordinated reestablishment of the epigenome are critical to mammalian embryonic development. Suz12 is a member of Polycomb repressive complex 2, known to catalyse trimethylation of lysine 27 on histone 3 (H3K27me3; Pasini et al. 2004); Suz12 has also been shown to interact with Suv39h1 for proper trimethylation of H3K9 (de la Cruz et al. 2007). Our objective in this study was to suppress expression of Suz12 via cytoplasmic injection of small interfering RNA (siRNA) targeted at this gene, and to evaluate the effect on embryo development rates. Bovine zygotes were produced in vitro via standard laboratory procedures. Nineteen hours post-fertilization, presumptive zygotes (n = 3979) were divided into 3 treatment groups: noninjected control (CTL), or injected with a fluorescent dextran marker combined with either a nontargeting siRNA (NULL) or an Suz12-targeting siRNA (SUZ). Embryos were cultured in Bovine Evolve (Zenith Biotech, Canada) with 4 mg mL–1 of BSA (Probumin, Millipore, Billerica, MA) and collected at the 4-cell, 8-cell, morula, and blastocyst stages. Ribonucleic acid was isolated with an RNeasy® Mini Kit (Qiagen, Valencia, CA) from 3 replicates of pooled embryos at each stage (4-cell, n = 15; 8-cell, n = 20; morula, n = 10), except for the blastocyst stage [2 samples (n = 10) collected from the CTL and NULL groups, and 1 sample (n = 3) from the SUZ group]. Each RNA sample was reverse-transcribed into cDNA and diluted for use by quantitative real-time PCR. Relative gene expression levels from each sample were calculated in triplicate using the SYBR Green comparative Ct method (Applied Biosystems, Foster City, CA), adjusted according to individual PCR efficiencies for each primer pair (R2 > 0.95) and normalized to the geometric mean Ct of 3 endogenous controls (GAPDH, YWHAZ, and SDHA), to account for differences in both cell number and amount of total mRNA present in each sample (Goossens et al. 2005). Our data indicate that Suz12 expression was suppressed to undetectable levels in SUZ-treated zygotes at all embryo stages analysed. The blastocyst rate of the SUZ group was extremely low (0.88 ± 0.16% SEM) compared with the CTL (19.87 ± 0.36% SEM) and NULL (5.09 ± 0.36% SEM) groups. Morphologically, SUZ morulae appeared fragmented with fewer larger cells than expected, whereas the NULL and CTL morulae seemed to develop normally. We presume the loss of Suz12 expression during this important developmental time is detrimental to embryo morphology and results in a decreased rate of blastocyst formation. Because of this decrease, we were able to collect only 3 SUZ blastocysts from a total of 227 injected. The microinjection procedure also contributed to significant (P < 0.05) decreases in blastocyst rates of the injected groups as compared with the CTL. Future experiments will explore potential alterations in histone methylation, as well as other epigenetic modifiers in bovine preimplantation embryos, to further elucidate the role of the epigenome in early embryonic development.
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
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 | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 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 ».