Western blotting
The ipsilateral cerebral hemisphere, rat leukocytes, and cultured cells
were homogenised in lysis buffer containing phosphatase and protease
inhibitors. We used the following primary antibodies:
anti‐Interleukin (IL)-1β
(Catalogue, AF-501-NA), anti-tumour necrosis factor
(TNF)-α (Abcam, ab205587),
anti‐Matrix
Metallopeptidase(MMP)-9 (Abcam, ab76003), anti‐zonula occludens(ZO)-1
(arigo, ARG55738), anti-occludin (Abcam, ab216327), anti‐caspase-3
(Abcam, ab13847; Affinity Biosciences, AF7022), anti‐C1QTNF6 (Abcam,
ab36900), and anti‐β‐actin (Abcam, ab20272). Signals were detected
through incubation with horseradish peroxidase-conjugated secondary
antibodies (Santa Cruz Biotechnology) for 60 min at room temperature and
using an enhanced chemiluminescence kit (Millipore, Darmstadt, Germany).
Grey values of the protein bands were analysed using ALPHAEASE FC
software (Alpha Innotech, San Leandro, CA, USA).