Cerebellar endothelial cells were isolated through enzymatic digestion, followed by separation using magnetic-activated cell sorting by anti-CD31–conjugated magnetic beads (MACS MS system, Miltenyi Biotec), as previously described (6). Isolated cerebellar endothelial cells were pelleted, and total RNA was extracted using the RNeasy micro kit (Qiagen 74004). PCR reactions were set up in triplicate, and data shown are from at least three independent repeats. Changes in gene expression were detected using the following mouse primers: Gapdh, 5′-GTCCCGTAGACAAAATGGTGA-3′ (forward) and 5′-TTTGATGTTAGTGGGGTCTCG-3′ (reverse); Klf2, 5′-CGCCTCGGGTTCATTTC-3′ (forward) and 5′-AGCCTATCTTGCCGTCCTTT-3′ (reverse); Klf4, 5′-GTGCCCCGACTAACCGTTG-3′ (forward) and 5′-GTCGTTGAACTCCTCGGTCT-3′ (reverse); eNos, 5′-CCTGTGCATGGATGAGTATGA-3′ (forward) and 5′-TGAGCAGGAGACACTGTTGAA-3′ (reverse); and Id1, 5′-ATCCTGCAGCATGTAATCGAC-3′ (forward) and 5′-GAGTCCATCTGGTCCCTCAGT-3′ (reverse).

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