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Background Studies confirmed that hydroxycamptothecin cause the apoptosis of human Tenon capsule fibroblasts (HTFs) by protein kinase R-like endoplasmic reticulum stress kinase (PERK) single pathway. Autophagy and apoptosis are programmed cell death following stress reaction, so they remain a close association. However, the effect of hydroxycamptothecin on the autophagy of HTFs and its mechanism are still unclear. Objective This study was to explore the promoting effect of PERK signal pathway on hydroxycamptothecin inducing the autophagy of HTFs. Methods This study procedure was approval by Ethic Committee of Nanjing Medical University. Human Tenon capsule tissue was obtained from fresh adult donors. HTFs were cultured and passaged by explant-culture method and identified by immunofluorescence for vimentin and keratin. pLVX-PERK lentiviral packed by 293T cells was transfected into HTFs to obtain stable PERK-knockout cell line by puromycin selection. Then the HTFs were treated with 0.10 g/L of hydroxycamptothecin for 5 minutes and consecutively cultivated for 24 hours, and the untreated cells were used as the control group. Western blot assay was used to detect the expressions of autophagy specific proteins in the cells, including autophagy related gene 5 (ATG-5), Beclin-1, light chain 3 (LC-3). Cyto-ID staining was used to identify the autophagosome in the cells. The experimental results were analyzed and compared between different treating groups. Results The gray scales for the expressions of Beclin 1, ATG-5, LC-3-Ⅰ and LC-3-Ⅱ proteins in HTFs were 0.365±0.045, 0.765±0.055, 0.120±0.030 and 0.215 …