Ischemic stroke is a major cause of death and disability worldwide. Necroptosis is known as a form of cell death, playing an essential role in regulating ischemia-induced brain injury. Triad3A is a ubiquitin ligase of the RING-in-between-RING family, and regulates necroptotic cell death under different pathological conditions, including neurodegenerative disorders. In the present study, the effects of Triad3A on experimental stroke were explored on a mouse model with middle cerebral artery occlusion (MCAO). The results indicated that Triad3A expression was markedly induced in the ischemic brain after MCAO operation. The neurons and microglia cells were the major cellular sources for Triad3A induction. Triad3A knockdown enhanced the infarction area, cell death, microglia activity, and the expression levels of pro-inflammatory markers including tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, inducible nitric oxide synthase (iNOS), CD32 and CD68 in MCAO mice. Triad3A and necroptosis were triggered in mouse microglia cells treated with oxygen and glucose deprivation (OGD), and in TNFα-incubated mouse hippocampal neuronal cells treated with Z-VAD-fmk, known as a pan-caspase inhibitor. Moreover, Triad3A knockdown accelerated cell death in microglial cells and neurons under these stresses. Furthermore, pre-treatment with necroptosis inhibitor markedly inhibited the cell death promoted by Triad3A silence in brain of mice with MCAO operation, demonstrating that Triad3A could regulate necroptosis to meditate the progression of cerebral I/R injury. Collectively, these finding illustrated that Triad3A could be served as a potential target for stroke therapy.