AbstractThe mechanisms of meniscus fibrosis and novel ways to enhance fibrosis is unclear. This work reveals human meniscus fibrosis initiated at E24 weeks. Smooth muscle cell cluster is identified in embryonic meniscus, and the combined analysis with previous data suggests smooth muscle cell in embryonic meniscus as precursors of progenitor cells in the mature meniscus. NOTCH3 is constantly expressed in smooth muscle cells throughout embryogenesis to adulthood. Inhibition of NOTCH3 signaling in vivo inhibits meniscus fibrosis and exacerbates degeneration. Continuous histological sections show that HEYL, NOTCH3 downstream target gene, is expressed consistently with NOTCH3. HEYL knockdown in meniscus cells attenuated the COL1A1 upregulation by CTGF and TGF‐βstimulation. Thus, this study discovers the existence of smooth muscle cells and fibers in the meniscus. Inhibition of NOTCH3 signaling in meniscus smooth muscle cells in a HEYL‐dependent manner prevented meniscus fibrosis and exacerbated degeneration. Therefore, NOTCH3/HEYL signaling might be a potential therapeutic target for meniscus fibrosis.