A display substrate includes a substrate, at least one protruding structure, a water-oxygen barrier pattern, and a display layer. The substrate has a display area, an opening located in the display area, and an isolation region disposed around the opening. The at least one protruding structure is disposed on the substrate and located in the isolation region. The water-oxygen barrier pattern covers the at least one protruding structure and located in the isolation region. The display layer is disposed on the substrate, a portion of the display layer is located in the display area, and a remaining portion of the display layer extends into the isolation region and is located at a side of the water-oxygen barrier pattern away from the substrate.