PMID- 35641967 OWN - NLM STAT- MEDLINE DCOM- 20220602 LR - 20230907 IS - 1471-2415 (Electronic) IS - 1471-2415 (Linking) VI - 22 IP - 1 DP - 2022 May 31 TI - The effect of A2E on lysosome membrane permeability during blue light-induced human RPEs apoptosis. PG - 241 LID - 10.1186/s12886-022-02464-1 [doi] LID - 241 AB - BACKGROUND: To investigate the effect of N-retinyl-N-retinylidene ethanolamine (A2E) on lysosome membrane permeability (LMP) during blue light-induced human retinal pigment epithelium cells (RPEs) apoptosis. METHODS: By building an A2E and blue light irradiation inducing RPEs damage model, the CCK-8 assay was used to detect RPEs viability loaded with different concentrations of A2E after different culturing time to determine the optimum A2E loading concentration. And the RPEs fluorescence intensity changes were observed by fluorescence microscopy loaded with different concentration of A2E. The RPEs were divided into four groups randomly: control group, A2E-loaded group, blue light irradiation group, and A2E-loaded + blue light irradiation group. Annexin V-FITC/PI and TUNEL/DAPI methods were used to detect RPEs apoptotic rate. Laser scanning confocal microscopy (LSCM) was used to observe RPEs LMP changes stained by acridine orange (AO) method. RESULTS: The CCK-8 result showed a downward trend in cells viability of RPEs loaded with increasing concentration of A2E and extending culturing time. The optimum A2E loading concentration was determined at 25 mumol/L. With increasing A2E loading concentrations, the intensity of fluorescence in RPEs decreased gradually. The RPEs apoptotic rate in blue light irradiation + A2E-loaded group was significantly higher than those in other three groups detected by Annexin V-FITC/PI method, which was similar to TUNEL/DAPI's result. After AO staining, cytoplasmic and nucleolar RNAs emits green fluorescence; lysosomes emit red fluorescence. Through the interference of A2E and blue light on RPEs, red fluorescent leakage from the lysosomes (means LMP increasing) can be observed. The mean red fluorescence intensity was chosen as the statistics indicator to estimate LMP change in RPEs cultured in vitro. Compared with the control group, the red fluorescence intensity decreased in A2E-loaded group, blue light irradiation group, and blue light irradiation + A2E-loaded group. Meanwhile, the mean red fluorescence intensity in blue light irradiation + A2E-loaded group was the lowest. CONCLUSIONS: Both A2E-loaded and blue light irradiation could induce human RPEs apoptosis, and the two factors had a synergistic effect. In addition, both A2E and blue light can lead to LMP increasing, which indicated LMP change might be the upstream part in inducing mitochondrion-dependent apoptotic pathway. These data provided evidence that A2E as the most important auto-fluorescence substance in lipofuscin is an initiator of blue light-mediated damage of RPEs and participate in pathogenesis of retinal degenerative diseases in humans. CI - (c) 2022. The Author(s). FAU - Xu, Yan AU - Xu Y AD - Department of Ophthalmology, Affiliated Hospital of Zunyi Medical University, No.149 Dalian Road, Zunyi, 563000, Guizhou, China. AD - Department of Ophthalmology, HwaMei Hospital, University of Chinese Academy of Sciences, Ningbo No.2 Hospital, Ningbo, Zhejiang, China. FAU - Li, Dan AU - Li D AD - Department of Ophthalmology, Affiliated Hospital of Zunyi Medical University, No.149 Dalian Road, Zunyi, 563000, Guizhou, China. AD - Department of Ophthalmology, The People's Hospital of Longchang, Neijiang, Sichuan, China. FAU - Su, Gang AU - Su G AD - Department of Ophthalmology, Affiliated Hospital of Zunyi Medical University, No.149 Dalian Road, Zunyi, 563000, Guizhou, China. AD - Guizhou Eye Hospital, Zunyi, Guizhou, China. AD - Guizhou Provincial Branch of National Eye Disease Clinical Medicine Research Center, Zunyi, Guizhou, China. AD - Special Key Laboratory of Ocular Diseases of Guizhou Province, Zunyi Medical University, Zunyi, Guizhou, China. FAU - Cai, Shanjun AU - Cai S AD - Department of Ophthalmology, Affiliated Hospital of Zunyi Medical University, No.149 Dalian Road, Zunyi, 563000, Guizhou, China. caishanjun@163.com. AD - Guizhou Eye Hospital, Zunyi, Guizhou, China. caishanjun@163.com. AD - Guizhou Provincial Branch of National Eye Disease Clinical Medicine Research Center, Zunyi, Guizhou, China. caishanjun@163.com. AD - Special Key Laboratory of Ocular Diseases of Guizhou Province, Zunyi Medical University, Zunyi, Guizhou, China. caishanjun@163.com. LA - eng GR - 81760174/National Natural Science Foundation of China/ GR - 81760174/National Natural Science Foundation of China/ GR - 81760174/National Natural Science Foundation of China/ GR - (2018)5601/Project of Innovative Talent Team of Guizhou province/ GR - (2018)5601/Project of Innovative Talent Team of Guizhou province/ GR - (2018)5601/Project of Innovative Talent Team of Guizhou province/ GR - (2018)5601/Project of Innovative Talent Team of Guizhou province/ GR - (2014) NO.95/Joint Project Foundation of Zunyi Medical University/ PT - Journal Article DEP - 20220531 PL - England TA - BMC Ophthalmol JT - BMC ophthalmology JID - 100967802 RN - 0 (Retinoids) SB - IM MH - Apoptosis MH - Humans MH - Lysosomes/metabolism MH - Permeability MH - *Retinal Pigment Epithelium MH - Retinoids/pharmacology PMC - PMC9158258 OTO - NOTNLM OT - Apoptosis, Lysosomal membrane permeability (LMP), N-retinyl-N-retinylidene ethanolamine (A2E), Blue light, Retinal pigment epithelium cells (RPEs), Apoptosis, Age-related macular degeneration (AMD) COIS- The authors declare that they have no competing interests. EDAT- 2022/06/02 06:00 MHDA- 2022/06/03 06:00 PMCR- 2022/05/31 CRDT- 2022/06/01 00:17 PHST- 2021/11/05 00:00 [received] PHST- 2022/05/24 00:00 [accepted] PHST- 2022/06/01 00:17 [entrez] PHST- 2022/06/02 06:00 [pubmed] PHST- 2022/06/03 06:00 [medline] PHST- 2022/05/31 00:00 [pmc-release] AID - 10.1186/s12886-022-02464-1 [pii] AID - 2464 [pii] AID - 10.1186/s12886-022-02464-1 [doi] PST - epublish SO - BMC Ophthalmol. 2022 May 31;22(1):241. doi: 10.1186/s12886-022-02464-1.