Objective To investigate the inhibitory effect of L⁃carnitine on skeletal muscle fibrosis in aged mice with cancer cachexia via Runx2/COL1A1 pathway. Methods A cancer cachexia model was constructed by subcutaneously implanting colon cancer MC38 cells in the right inguinal groin of C57 aged mice. The mice were divided into non⁃tumor⁃bearing group (NTB group), tumor⁃bearing group (TB group) and L⁃carnitine group (LC group), with respect to corresponding interventions. After the cessation of the intervention, one side of the gastrocnemius muscle (GM) was weighed. Hematoxylin and eosin (HE) staining and Masson staining were then performed, and the cross⁃sectional area of the GM and the percent of collagen fiber area were measured respectively. The total protein from the opposite side of the GM was extracted, and the levels of Runx2 and COL1A1 protein were detected by Western blotting. NIH/3T3 cells were induced using transforming growth factor⁃β1 (TGF⁃β1) to establish fibrosis model in vitro. Total protein and mRNA were extracted after L⁃carnitine intervention, respectively. Runx2 and COL1A1 protein levels were detected by Western blotting, and COL1A1 mRNA levels were detected by qRT⁃PCR. cDNA⁃Runx2 was transfected to verify that COL1A1 was regulated by the L⁃carnitine through Runx2. Results By comparing the three groups, the weight of the GM in the TB group was significantly lower than that of the NTB group [(98.12±17.04)mg vs (122.18±6.91)mg] (P<0.05); the cross⁃sectional area of the GM in the TB group (207.46±54.55)µm2 was significantly lower than the NTB group (488.61±46.72) µm2 and the LC group (434.54±113.84)µm2 (P<0.05); the area of collagen fibers in the TB group (9.69±1.55)% was significantly higher than that of the LC group (5.48±1.19)% and the NTB group (3.88±0.86)% (P<0.05). The results of Western blotting showed that compared with the LC group and the NTB group, Runx2 and COL1A1 were highly expressed in the TB group(P<0.05). In vitro studies showed that L⁃carnitine reversed the TGF⁃β1 induced high expression of Runx2 protein and COL1A1 mRNA. Overexpression of Runx2 confirmed that L⁃carnitine down⁃regulated COL1A1 mRNA by inhibiting Runx2 protein expression. Conclusion L⁃carnitine ameliorates skeletal muscle fibrosis in aged mice with cancer cachexia partly due to its down⁃regulating effects of Runx2 / COL1A1.