02305nas a2200277   4500000000100000000000100001008004100002260000900043100002000052700001700072700002200089700001500111700001600126700001800142700001500160700002200175700002300197700001900220700001700239245009900256856006300355300001400418490000700432520157400439022001402013       2023                            d  c20231 aAyşe J. Muñiz1 aTuğba Topal1 aMichael D. Brooks1 aAngela Sze1 aDo Hoon Kim1 aJacob Jordahl1 aJoe Nguyen1 aPaul H. Krebsbach1 aMasha G. Savelieff1 aEva L. Feldman1 aJoerg Lahann00aEngineered extracellular matrices facilitate brain organoids from human pluripotent stem cells  uhttps://onlinelibrary.wiley.com/doi/abs/10.1002/acn3.51820  a1239-12530 v103 aObjective Brain organoids are miniaturized in vitro brain models generated from pluripotent stem cells, which resemble full-sized brain more closely than conventional two-dimensional cell cultures. Although brain organoids mimic the human brain's cell-to-cell network interactions, they generally fail to faithfully recapitulate cell-to-matrix interactions. Here, an engineered framework, called an engineered extracellular matrix (EECM), was developed to provide support and cell-to-matrix interactions to developing brain organoids. Methods We generated brain organoids using EECMs comprised of human fibrillar fibronectin supported by a highly porous polymer scaffold. The resultant brain organoids were characterized by immunofluorescence microscopy, transcriptomics, and proteomics of the cerebrospinal fluid (CSF) compartment. Results The interstitial matrix-mimicking EECM enhanced neurogenesis, glial maturation, and neuronal diversity from human embryonic stem cells versus conventional protein matrix (Matrigel). Additionally, EECMs supported long-term culture, which promoted large-volume organoids containing over 250 μL of CSF. Proteomics analysis of the CSF found it superseded previous brain organoids in protein diversity, as indicated by 280 proteins spanning 500 gene ontology pathways shared with adult CSF. Interpretation Engineered EECM matrices represent a major advancement in neural engineering as they have the potential to significantly enhance the structural, cellular, and functional diversity that can be achieved in advanced brain models.  a2328-9503