Effects of matrigel on growth and development of cerebral organoids

Abstract

Extracellular matrix (ECM) is very important in cerebral organoids culture because it affects organoids growth and development. The most common ECM is Matrigel. To study the effect of Matrigel on cerebral organoids, diameter analysis, live/dead staining and RNA sequencing were performed on cerebral organoids using two common Matrigel, Matrigel 354277 and Matrigel 356230 (low growth factors). The morphology and diameter of the organoids were analyzed, which revealed that the average size and the relative area of the neuroepithelium of 356230-group were slightly larger than that of 354277-group. Besides, it showed that there was essentially no difference in the activity of organoids between the two groups by live-dead staining at day 15. Finally, RNA sequencing was used to analyze gene expression at day 10, day 20 and day 30. There were 738, 2190 and 689 differentially expressed genes (DEGs) respectively. 14, 14 and 19 KEGG biological pathways involved in the DEGs were selected to study. At day 10, signaling pathways regulating pluripotency of stem cells was important and the downregulation of it in 354277-group affected ectodermal development. At day 20, PI3K-Akt signaling pathway and MAPK signaling pathway were the key pathways and the upregulation of key genes indicated that 354277-group might have more apoptosis than 356230-group. Besides, at day 30, retrograde endocannabinoid signaling pathway and GABAergic synapse were critical and downregulation of related genes affected synaptic signaling. Above all, types of Matrigel affected ectoderm differentiation, cell apoptosis and neurological function and Matrigel 356230 is a better choice than Matrigel 354277 for generating cerebral organoids.