For studying cellular communications ex-vivo, a two-dimensional (2D) cell culture model is currently used as the “gold standard”. 2D culture models are also widely used in the study of RNA expression profiles from tumor cells secreted extracellular vesicles (EVs) for tumor biomarker discovery. Although the 2D culture system is simple and easily accessible, the culture environment is unable to represent in vivo extracellular matrix (ECM) microenvironment. Our study observed that 2D- culture derived EVs showed significantly different profiles in terms of secretion dynamics and essential signaling molecular contents (RNAs and DNAs), when compared to the three-dimensional (3D) culture derived EVs. By performing small RNA next-generation sequencing (NGS) analysis of cervical cancer cells and their EVs compared with cervical cancer patient plasma EV-derived small RNAs, we observed that 3D- culture derived EV small RNAs differ from their parent cell small RNA profile which may indicate a specific sorting process. Most importantly, the 3D- culture derived EV small RNA profile exhibited a much higher similarity (~96%) to in vivo circulating EVs derived from cervical cancer patient plasma. However, 2D- culture derived EV small RNA profile correlated better with only their parent cells cultured in 2D. On the other hand, DNA sequencing analysis suggests that culture and growth conditions do not affect the genomic information carried by EV secretion. This work also suggests that tackling EV molecular alterations secreted into interstitial fluids can provide an alternative, non-invasive approach for investigating 3D tissue behaviors at the molecular precision. This work could serve as a foundation for building precise models employed in mimicking in vivo tissue system with EVs as the molecular indicators or transporters. Such models could be used for investigating tumor biomarkers, drug screening, and understanding tumor progression and metastasis. [ Lire plus… ]
Ce document explore la production de vésicules extracellulaires (EVs) dans des cultures cellulaires en 2D et 3D. Les auteurs constatent que les EVs produits en culture 3D ressemblent davantage aux EVs circulant in vivo, en termes de dynamique de sécrétion et de contenu moléculaire.
Ils observent que les EVs issus de cultures 2D présentent des profils de sécrétion et des contenus moléculaires essentiels (ARN et ADN) significativement différents de ceux issus de cultures 3D. Les EVs de culture 3D contiennent des petits ARN qui reflètent mieux l’état in vivo, avec une similarité d’environ 96% avec les EVs circulants de patients atteints de cancer du col de l’utérus. En revanche, l’analyse de l’ADN suggère que les conditions de culture n’affectent pas l’information génomique portée par la sécrétion des EVs.
Les auteurs soulignent que les cultures 3D sont plus pertinentes pour reproduire le comportement cellulaire in vivo, suggérant leur utilité dans la recherche sur les biomarqueurs tumoraux, le criblage de médicaments et la compréhension de la progression et de la métastase des tumeurs. Cette étude fournit des informations importantes sur la manière dont les EVs peuvent être utilisés pour étudier les comportements tissulaires en 3D et offre une approche alternative et non invasive pour examiner ces comportements au niveau moléculaire.