L-Carnitine (LC) has protective effects on high glucose-induced oxidative stress in the retinal ganglion cells (RGCs). The aim of this study was to investigate the role of NF-E2-related factor 2 (Nrf2), Kelch likeECH-associated protein 1 (Keap1), haemoxygenase-1 (HO-1) and g-glutamyl cysteine synthetase (gGCS) in the protective effect of LC on RGCs. RGCs were first processed with high concentrations of glucose. LC treatment at three concentrations (50 mM, 100 mM and 200 mM) was applied to high glucose stimulated RGCs. The expression of Nrf2, Keap1, haemoxygenase-1 (HO-1) and g-glutamyl cysteine synthetase (g-GCS) was quantified by Western blot in the treatment and control (high glucose stimulation) groups. In the three LC groups (50 mM, 100 mM and 200 mM), Nrf-2 (0.71 0.04, 0.89 0.05, 1.24 0.05 vs 0.56 0.03, p < 0.05), HO-1 (0.58 0.04, 0.76 0.06, 0.89 0.07 vs 0.25 0.03, p < 0.01), and g-GCS protein expression (0.66 0.03, 0.79 0.05, 0.84 0.08 vs 0.84 0.08, p < 0.01) was higher than in the control group. The levels of Keap1 protein were in the LC groups were lower than in the control group (0.50 0.03, 0.45 0.02, 0.53 0.03 vs 0.86 0.05, p < 0.01). In conclusion, in high glucose stimulated RGCs, LC treatment was associated with an increased level of Nrf2, HO-1and g-GCS. LC treatment was also associated with a reduced expression of Keap1 protein. These results suggest that the protective effect of LC treatment on RGCs may be related to Nrf2-Keap1 pathway.