INTRODUCTION: Neointima formation is closely linked to vascular stenosis and occurs after endothelial damage. Hydrogen sulfide is an endogenous pleiotropic mediator with numerous positive effects on the cardio vascular system. OBJECTIVE: This study evaluates the effect of the slow releasing hydrogen sulfide donor GYY4137 (GYY) on neointimal formation in vivo. METHODS: The effect of GYY on neointimal formation in the carotid artery was studied in the FeCl3 injury model in GYY- or vehicle-treated mice. The carotid arteries were studied at days 7 and 21 after treatment by means of histology and immunohistochemistry for proliferating cell nuclear antigen (PCNA) and alpha smooth muscle actin (α-SMA). RESULTS: GYY treatment significantly reduced the maximal diameter and the area of the newly formed neointima on both days 7 and 21 when compared to vehicle treatment. GYY additionally reduced the number of PCNA- and α-SMA-positive cells within the neointima on day 21 after FeCl3 injury of the carotid artery. CONCLUSIONS: Summarizing, single treatment with the slow releasing hydrogen sulfide donor GYY reduced the extent of the newly formed neointima by affecting the cellular proliferation at the site of vascular injury.