scRNA-seq of brain cells from control and 3xDR mouse brains

As important immune cells, microglia undergo a series of alterations during aging that increase the susceptibility to brain dysfunctions. However, the longitudinal characteristics of microglia remain elusive. In this study, we mapped the transcriptional and epigenetic profiles of microglia from 3- to 24-month-old mice. We observed unexpected sex differences and identified age-dependent microglia (ADEM) genes during the aging process. We then compared the characteristics of aging and reactivity in female microglia at the single-cell resolution and epigenetic level. To dissect the functions of aged microglia excluding the influence from other aged brain cells, we established an accelerated microglial turnover model without directly affecting other brain cells. By this model, we achieved aged microglia in non-aged brains and confirmed that aged microglia per se contribute to cognitive decline. Collectively, our work provides a comprehensive resource for decoding the aging process of microglia, shedding light on how microglia maintain brain functions. Colony-stimulating factor 1 receptor (CSF1R) inhibition by PLX5622 for 14 days results in the ablation of 99% of microglia in the brain without directly affecting other cells. Once this inhibition is removed, residual microglia (100% - 99% = 1%) rapidly proliferate, repopulate the whole brain and recover to a normal density (100%) within 2 weeks. Each microglial cell proliferates approximately log2(100%/1%) = 6.64 times during 1-round depletion-repopulation (1xDR). Thus, after 3xDR, each microglial cell has proliferated approximately 6.64 × 3 ≈ 20 times, close to the Hayflick limit (for some cells, the Hayflick Limits is 10~15 population doublings). Whole brain cells from control and 3-round microglia depletion-repopulation (3xDR) mice were collected for sequencing.