Inhibition of necroptosis mitigates paclitaxel-induced neuronal damage and cognitive impairment

Abstract

Paclitaxel (PTX) is a first-line chemotherapy agent for treating many types of cancers, induces cognitive impairment and neuronal damage. However, PTX-induced a limited apoptosis of neurons is not consistent with a wide range of neuroinflammation. Here, we demonstrated that in addition to inducing apoptosis in hippocampal neurons (HT22 cells), PTX causes necroptosis, a programmed cell death, via activation of the RIPK1-RIPK3-MLKL signaling pathway. Annexin V/PI dual labeling, flow cytometric analysis, image-based PI staining, and western blot techniques were used to evaluate PTX-induced necroptosis. Cell viability was determined using the CCK8 assay, whereas Ca2+ levels were measured using the Fluo-4 AM fluorescent probe. The number of cells that were positive for both Annexin V and PI staining was considerably higher in PTX-treated HT22 cells compared to those treated with the vehicle. Additionally, the nuclei of PTX-treated cells showed more diffuse necrotic staining with PI compared to the vehicle-treated cells. The Western blot study demonstrated a considerable increase in the expression of necroptotic proteins, including RIPK1, RIPK3, MLKL, and p-MLKL, following PTX treatment. The compound Necrotatin-1 (Nec-1), which specifically inhibits the protein RIPK1, effectively decreased the occurrence of necroptosis in HT22 cells triggered by PTX by lowering the excessive accumulation of intracellular Ca2+ overload. In addition, administration of Nec-1 in vivo rescued cognitive impairments in novel object recognition and Morris Water Maze tests in PTX-treated mice. These data suggest that PTX induces cognitive impairments through RIPK1-mediated necroptosis. Inhibition of necroptosis provided a potential therapeutic approach to reduce PTX-induced cognitive deficits.