Ginkgolide B (GB) is a small molecule from Ginkgo biloba and implicates the reduction of certain aging-related morbidities; however, whether GB improves overall healthspan and longevity remains unknown. By single-nucleus RNA sequencing (snRNA-seq), we revealed that GB partially restored the aging-related dysregulation in cell-type composition, intracellular signaling pathways, and cell-cell communication in skeletal muscle. Notably, GB reduced the quantity of aging-induced novel Runx1+ type 2B myonuclei, which are particularly associated with an apoptotic burden and aging-related signatures. Overall design: 3- (young), and 20-month-old (aged) female C57BL/6 mice were administered 12 mg/kg body weight GB or vehicle daily by oral gavage for two months. Gastrocnemius (GA) muscle of mice were minced, resuspended and labelled with DAPI. Labelled nuclei were sorted by FACS Aria II Cell Sorter and collected. Nuclei from 2 mice were pooled together for each sample and processed using Chromium Single Cell 3’ kit for downstream single-nuclei RNA sequencing.