Integrin α3

Mice were sacrificed on the last treatment day with rTMS. The peri‐infract cortex tissues (approximately 20 mg) were homogenized in 200 μL of RIPA lysis buffer (P0013B, Beyotime, Shanghai, China). Protein quantification was performed using a BCA protein assay kit (Melone Pharmaceutical, Dalian, China). Samples with 40 μg protein were separated by size using 10% sodium dodecyl sulfate–polyacrylamide gels and transferred onto nitrocellulose (NC) membranes. Membranes were blocked with 5% nonfat milk for 1 h at room temperature and then incubated overnight at 4°C with the following primary antibodies: integrin α3 (1:1000, 66,070–1, Proteintech), GluN2B (1:1000, 21,920‐1‐AP, Proteintech), β‐actin (1:1000, SD0034, SIMUWU), GluN2A (1:1000, 19,953‐1‐AP, Proteintech), GluA2/3/4 (1:1000, 2460, Cell Signaling), GluA1 (1:1000, 67,642‐1‐Ig, Proteintech), GAD65 (1:1000, 20,746‐1‐AP, Proteintech), and vGlut1 (1:1000, ab227805, Abcam). Membranes were washed with TBST and incubated with HRP‐conjugated goat anti‐mouse and goat anti‐rabbit antibody for 1 h at room temperature. ImageJ software was used to analyze protein bands.

To evaluate the cytocompatibility, adhesion and proliferation, PDO, PDGA, PDAG, PGAG membranes with thickness of 40 µm were prepared. Then, the membranes were plated in 6‐well plates. A total of 1.5 × 10⁵ HUVECs were seeded on each membrane, and were cultured in endothelial cell medium supplied with 10% FBS. After incubation for 24 h and 48 h, the membranes were washed with PBS for 3 times, stained with crystal violet solution (0.5%) for 30 min and the cells on membranes were counted.
For immunofluorescent staining, after incubation for 24 h, the cells seeded on membranes were incubated with Integrin α3 (Proteintech, 66070‐1) and FAK (Abcam, ab40794) overnight, and were stained with 488 conjugate (Cell Signaling, #4408), 594 conjugate (Cell Signaling, #8889) and DAPI (Solarbio, C0065) according to the manufacture's guidelines and were observed by confocal laser scanning microscopy (TCS SP5II, Leica, Ernst‐Leitz‐Strasse, Germany).
The interior morphology of HUVECs on PDO, PDGA, PDAG, and PGAG membranes was investigated by SEM (S‐4800, Hitachi, Japan). Samples were fixed in 2.5% glutaraldehyde, dehydrated by CO2 critical point drying (K850X, Emitech), quick‐frozen in liquid nitrogen, lyophilized, and then coated with gold particles.