Quantification of morphological, functional, and biochemical features of H9c2 rat cardiomyoblast retinoic acid differentiation
Notice bibliographique
Résumé
Cell culture models enable advancement in our understanding of heart development and heart disease. The H9c2 rat ventricular cardiomyoblast cell line can be differentiated with retinoic acid and low serum, leading to morphological, molecular, and functional changes that partially resemble aspects of cardiomyoblast-to-cardiomyocyte differentiation. However, morphological, functional, and biochemical changes are rarely investigated in parallel, thereby limiting fulsome understanding of how these processes are interlinked, and to what extent these model cardiomyoblasts can be differentiated. To provide a parallel analysis as a resource for future studies, we therefore characterized H9c2 cell morphology, Ca 2+ handling, and gene expression after five days (5 days- in-vitro, DIV5), and fourteen days (DIV14) of exposure to differentiation stimuli, consisting of retinoic acid and low serum. We observed statistically significant morphological changes during differentiation. We saw changes consistent with those already described in the context of cardiomyoblast differentiation. However, some of these were previously limited to qualitative observations, for example increased cell length. Notably, several of our morphological observations are completely novel, such as increases in eccentricity, perimeter length (aka cell boundary length), and in the density of actin clusters, were investigated de novo. Differentiation also resulted in the onset of spontaneous Ca 2+ transients – this is the first instance, to our knowledge, that this has been characterized in the absence of pharmacological stimulation. The mean frequency and synchronicity of Ca 2+ transients in differentiated H9c2 cells were much lower than those observed in primary cardiomyocytes, underscoring their relatively immature differentiation state from a functional perspective. Additionally, key cardiomyocyte cytoskeletal proteins and ion channel transcript and protein expression levels changed significantly with differentiation, including at early timepoints (DIV5 h and DIV3) which had not yet been investigated by others, in alignment with changes normally observed in cardiomyocyte development. Overall, our findings position differentiated H9c2 cells as a relatively high-throughput, model for studying cardiomyoblast differentiation, while also clarifying their limitations in recapitulating fully mature cardiomyocyte phenotypes and highlight reliable markers (e.g., Cacna1c, Myom2, cTnT, VCL, and Gja5) for experimental readouts. • We quantified morphology, Ca 2+ handling, and gene expression changes in rat ventricular cardiomyoblast H9c2 cells across retinoic acid differentiation. • We compared multiple differentiation time points (mainly DIV0, DIV5, and DIV14) and observed time-dependent changes in all 3 aspects. We observed several morphological changes with increasing time in differentiation, including increased mean length, area, eccentricity, and multinucleation, as well as an increase in actin clusters. We observed the onset of spontaneous Ca 2+ transients with differentiation, with further changes in subpopulations with increasing time in differentiation. Finally, we observed multiple changes in gene expression consistent with cardiomyoblast differentiation. • We summarize these parallel time-point dependent changes in morphology, Ca 2+ handing, and gene expression, thereby providing an invaluable resource for other researchers.
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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 | 0,000 | 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 ».