Inhibitor nc

sh-EZH2 or sh-NC (2.8 × 10⁸ TU/mL) was appointed to infect microglia. miR-29b-3p inhibitor or NC-inhibitor (RiboBio Co., Ltd, Guangzhou, Guangdong China) was transfected into microglia using Lipofectamine 2000 (Invitrogen Inc., Carlsbad, CA, USA) for further analysis. After the transfection, microglia were incubated in the medium with 100 ng/mL lipopolysaccharide (LPS) for 24 h [25 (link)].

Cells were seeded into 6-well plates at a density of 6 × 10⁴ cells/2 ml/well and maintained in an incubator at 37 °C, 5% CO₂. When the cells reached about 80% confluency, cell transfection was performed. The circ_ANRIL (hsa_circ_0008574) overexpression vector was synthesized by Sangon Biotech (Shanghai, China), and an empty vector (circ-NC) was used as a control. All miRNAs mimics (miR-622 mimic: 5′-ACAGUCUGCUGAGGUUGGAGC-3′, NC-mimic: 5′-UUCUCCGAACGUGUCACGUTT-3′), and miRNAs inhibitor (miR-622 inhibitor: 5′-GCUCCAACCUCAGCAGACUGU-3′, NC-inhibitor: 5′-CAGUACUUUUGUGUAGUACAA-3′) were purchased from RiboBio (Guangzhou, Guangdong, China). circ_ANRIL was knocked down using specific short interfering RNAs (siRNAs: si-circ_ANRIL: 5′-AGAATTTTGACAGTGTCCCTT-3′, si-NC: 5′TTCTCCGAACGTGTCACGT-3′) targeting the backsplice region. HBMECs were transfected with plasmids or oligonucleotides HBMECs using Lipofectamine™ 2000 (Life Technologies, Carlsbad, CA, USA), according to the manufacturer’s instructions. After 24 h of transfection, the efficiency of transfection was determined by qRT-PCR.

miR‐206 mimics and negative control (NC), miR‐206 inhibitor, and NC inhibitor were synthesized by RiboBio (Guangzhou, China). The sequences of the above RNA oligo were listed in Table S1. LDC000067 (CDK9 inhibitor) was purchased from Amquar Company (Shanghai, China). sh‐CDK9 plasmid was synthesized by GeneChem Company (Shanghai, China).