Hydroxycamptothecin Induces Apoptosis Of Human Tenon’s Capsule Fibroblasts By Activating The PERK Signaling Pathway

Purpose: Hydroxycamptothecin (HCPT) has been proven to induce apoptosis infibroblasts. In this study, we investigated whether the PRKR-like ER kinase (PERK)pathway is implicated in apoptotic signaling of human Tenon’s capsule fibroblasts(HTCFs) by HCPT.Methods: Normal and PERK-knockdown HTCFs were used in this study. Apoptosiswas determined by the cell viability assay, Annexin V/propidium iodide (PI)dual-staining, cell cycle analysis in HTCFs treated with HCPT in various doses andfor various durations. Endoplasmic reticulum (ER) stress markers and sensor proteinswere detected by Western blot. Mitochondrial dysfunction was measured by detectingthe mitochondrial membrane potential (ΔΨm) and measuring the expression ofcytochrome c (cyt c).Results:. HCPT induced apoptosis in the HTCFs, which was characterized asdecreased cell viability and sub-S fraction of the cell cycle and increased apoptosisrate by Annexin V/PI dual-staining. The activity levels of caspase-3and caspase-9 were significantly increased and were accompanied by cytosolic release of cyt c anddecreased ΔΨm in response to HCPT. Treatment with HCPT increased the expressionof glucose-regulated protein78(GRP78), phospho-PERK, activating transcriptionfactor6(ATF6), phospho-inositol-requiring kinase1(IRE1), C/EBP homologousprotein (CHOP), Bax, and phospho-c-Jun N-terminal kinase (JNK) and decreased theexpression of Bcl-2. Knockdown of PERK attenuates HCPT-induced apoptosis inHTCFs, dependent upon both ER stress and the mitochondrial apoptotic pathway.Conclusions: This study suggests that the endoplasmic reticulum stress response andmitochondrial dysfunction are involved in apoptosis induced by HCPT in fibroblasts,which might be mediated by PERK. Thus, this study offers new insight intopreventing postoperative scarring via treatment with HCPT.