interacted with ECs for ciBEC specification (Figure 4C). Some reports have shown that the Wnt canonical pathway is crucial for proper neurovascular development. For example, the simultaneous ablation of Wnt7a/7b is embryonic lethal for neurovascular phenotypes (Daneman et al., 2009, Stenman et al., 2008). Our results showed that Wnt7a was expressed in neurons and Wnt7b was expressed in pericytes and astrocytes (Figure S6B), but treatment with Wnt inhibitors in our co-culture system did not cause significant differences in the expression of BBB-specific transporters (Figure 3A). Moreover, we found that Wnt7a was highly expressed in early-stage neural progenitors (Figure S6C), indicating that Wnt7a may be necessary for early vascularization in the brain. Overall, our technology could help elucidate the underlying molecular machinery for finely regulated BBB formation, which should offer new therapeutic strategies to repair the BBB following stroke or other neurological impairments.
