Doxorubicin Induces a Senescent Phenotype in Murine and Human Astrocytes
Mariana Marques, Lívia de Sá Hayashide, Pedro Amorim, Beatriz Martins Fernandes, Ana Paula Bérgamo Araújo, Daniel Fernandes Messor, Vitor Emanuel Leocadio, Bruna V. Pessoa, João Bastos Lima Pacca Corrêa, Cristopher Villablanca, René L. Vidal, Christian González‐Billault, Isadora Matias, Flávia Carvalho Alcântara Gomes, Luan Pereira Diniz
Abstract
ABSTRACT Aging is a complex biological process that significantly impacts the central nervous system (CNS). Astrocytes, critical support cells in the brain, undergo senescence with age, contributing to neurodegenerative diseases. While previous studies have utilized murine models to investigate astrocyte senescence, human astrocytes offer a more physiologically relevant system to study age‐related neurodegenerative changes. This study presents a novel protocol for inducing senescence in both human primary and murine astrocytes using a combination of cellular stressors, such as lactacystin, H 2 O 2 , rotenone, and doxorubicin. Our results demonstrate that doxorubicin treatment effectively induces a robust senescent phenotype in both human and murine astrocytes, characterized by increased expression of senescence markers such as p21 and β‐galactosidase, along with activation of the DNA damage response (γ‐H2AX and 53BP1). Doxorubicin treatment increased nuclear size and induced cell cycle arrest in astrocytes, as revealed by reduced BrdU incorporation and decreased cell density, without inducing cytotoxic effects. This phenotype is accompanied by a pronounced pro‐inflammatory profile, with elevated expression of cytokines including MMP3, IL‐6, and IL‐1β, indicative of a strong senescence‐associated secretory phenotype (SASP). These findings provide a novel in vitro model for murine and human astrocyte senescence induced by doxorubicin, highlighting its relevance for studying mechanisms underlying age‐related neuroinflammation and neurodegeneration. By establishing a robust model of human astrocyte senescence, this study provides a valuable tool for exploring the molecular and cellular mechanisms driving age‐related neurodegenerative processes, serving as an alternative approach to traditional murine models. image