Lidocaine has been reported to induce apoptosis on rabbit corneal endothelial cells. However, the apoptotic effect and exact mechanism involved in cytotoxicity of lidocaine are not well-established in human corneal endothelial (HCE) cells. In this study, we investigated the apoptosis-inducing effect of lidocaine on HCE cells in vitro. After HCE cells were treated with lidocaine at concentrations of 0.15625-10.0 g/l, the morphology and ultrastructure of the cells were observed by inverted light microscope and transmission electron microscope (TEM). Cell viability was measured by MTT assay, and the apoptotic ratio was evaluated with flow cytometry and fluorescent microscopic counting after FITC-Annexin V/PI and AO/EB staining. DNA fragmentation was detected by electrophoresis, and the activation of caspases was evaluated by ELISA. In addition, changes in mitochondrial membrane potential were determined by JC-1 staining. Results suggest that lidocaine above 1.25 g/l reduced cellular viability and triggered apoptosis in HCE cells in a time-and dose-dependent manner. Diminishment of DΨm and the activation of caspases indicate that lidocaine-induced apoptosis was caspase dependent and may be related to mitochondrial pathway.Human corneal endothelium (HCE), a functional monolayer forming the demarcation between cornea and anterior chamber, plays pivotal roles in maintaining corneal transparency by regulating corneal stromal water content [1]. Notwithstanding the evidence of proliferation when deprived of their nature environment, HCE cells are trapped in the G1 phase of the cell cycle and have a limited, if not restricted, proliferative capacity in vivo [2,3], accompanied by an annual attrition rate of 0.3-0.6% in cell density during adulthood [4].In addition to ageing, excessive HCE cell loss may result from accidental injuries, surgical trauma and diseases [5][6][7] which consequently impair the physiology of cornea. Due to the inability of HCE to be repaired through cell number increase, wound healing can only be achieved by enlargement and migration of the neighbouring cells. However, if the decline of cell density transcends a threshold, the integrity of the corneal endothelium will be compromised, resulting in painful corneal oedema and ultimately loss of visual acuity. Recently, increased evidence has revealed corneal endothelium damage to be associated with abused administration of topical anaesthetics [8][9][10].Lidocaine is one of the most frequently used topical anaesthetic agents in ophthalmic surgeries for its inherent potency, rapid onset, tissue penetration and efficiency [11]. It can result in corneal thickening, opacification and significant corneal endothelial cell loss when applied intracamerally [12]. In addition, administration of lidocaine has been shown to cause an increase in apoptotic cells in corneal endothelium in rabbit models [13]. However, the side effects of lidocaine on HCE cells and their mechanism remain unknown. In this study, we aimed to investigate the apoptosis-inducing e...