title: Cytotoxicity of atropine to human corneal endothelial cells by inducing mitochondrion-dependent apoptosis.
abstract: Atropine, a widely used topical anticholinergic drug, might have adverse effects on human corneas in vivo. However, its cytotoxic effect on human corneal endothelium (HCE) and its possible mechanisms are unclear. Here, we investigated the cytotoxicity of atropine and its underlying cellular and molecular mechanisms using an in vitro model of HCE cells and verified the cytotoxicity using cat corneal endothelium (CCE) in vivo. Our results showed that atropine at concentrations above 0.3125 g/L could induce abnormal morphology and viability decline in a dose- and time-dependent manner in vitro. The cytotoxicity of atropine was proven by the induced density decrease and abnormality of morphology and ultrastructure of CCE cells in vivo. Meanwhile, atropine could also induce dose- and time-dependent elevation of plasma membrane permeability, G1 phase arrest, phosphatidylserine externalization, DNA fragmentation, and apoptotic body formation of HCE cells. Moreover, 2.5 g/L atropine could also induce caspase-2/-3/-9 activation, mitochondrial transmembrane potential disruption, downregulation of anti-apoptotic Bcl-2 and Bcl-xL, upregulation of pro-apoptotic Bax and Bad, and upregulation of cytoplasmic cytochrome c and apoptosis-inducing factor. In conclusion, atropine above 1/128 of its clinical therapeutic dosage has a dose- and time-dependent cytotoxicity to HCE cells in vitro which is confirmed by CCE cells in vivo, and its cytotoxicity is achieved by inducing HCE cell apoptosis via a death receptor-mediated mitochondrion-dependent signaling pathway. Our findings provide new insights into the cytotoxicity and apoptosis-inducing effect of atropine which should be used with great caution in eye clinic.
authors.affiliation: Laboratory for Corneal Tissue Engineering, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, Shandong Province, China.
authors.forename: Qian
authors.initials: Q
authors.lastname: Wen
authors.affiliation: Laboratory for Corneal Tissue Engineering, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, Shandong Province, China tjfan@ouc.edu.cn.
authors.forename: Ting-Jun
authors.initials: TJ
authors.lastname: Fan
authors.affiliation: Laboratory for Corneal Tissue Engineering, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, Shandong Province, China.
authors.forename: Cheng-Lei
authors.initials: CL
authors.lastname: Tian
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D001285:Atropine / Q000009:adverse effects* / Q000633:toxicity; D002415:Cats; D002463:Cell Membrane Permeability / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D018680:Cholinergic Antagonists / Q000009:adverse effects* / Q000633:toxicity; D003315:Cornea / Q000187:drug effects* / Q000648:ultrastructure; D053938:DNA Fragmentation; D042783:Endothelial Cells / Q000187:drug effects* / Q000648:ultrastructure; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008297:Male; D008928:Mitochondria / Q000187:drug effects*; D009184:Mydriatics / Q000009:adverse effects* / Q000633:toxicity
chemical_list: D018680:Cholinergic Antagonists; D009184:Mydriatics; D001285:Atropine
keywords: Atropine; apoptosis; cat corneal endothelial cells; cytotoxicity; human corneal endothelial cells; mitochondrion