論文 - 詳細
| RRC ID | 73112 |
|---|---|
| 著者 | Kanzaki Y, Fujita H, Sato K, Hosokawa M, Matsumae H, Morizane Y, Ohuchi H. |
| タイトル | Protrusion of KCNJ13 Gene Knockout Retinal Pigment Epithelium Due to Oxidative Stress-Induced Cell Death. |
| ジャーナル | Invest Ophthalmol Vis Sci |
| Abstract |
PURPOSE:This study was performed to elucidate the mechanisms of morphological abnormalities in a Leber congenital amaurosis 16 (LCA16) cell model using KCNJ13 knockout (KO) retinal pigment epithelial cells derived from human iPS cells (hiPSC-RPE). METHODS:In KCNJ13 KO and wild-type hiPSC-RPE cells, ZO-1 immunofluorescence was performed, and confocal images were captured. The area and perimeter of each cell were measured. To detect cell death, ethidium homodimer III (EthD-III) staining and LDH assay were used. Scanning electron microscopy (SEM) was used to observe the cell surface. The expression levels of oxidative stress-related genes were examined by quantitative PCR. To explore the effects of oxidative stress, tert-butyl hydroperoxide (t-BHP) was administered to the hiPSC-RPE cells. Cell viability was tested by MTS assay, whereas oxidative damage was monitored by oxidized (GSSG) and reduced glutathione levels. RESULTS:The area and perimeter of KCNJ13-KO hiPSC-RPE cells were enlarged. EthD-III-positive cells were increased with more dead cells in the protruded region. The KO RPE had significantly higher LDH levels in the medium. SEM observations revealed aggregated cells having broken cell surfaces on the KO RPE sheet. The KCNJ13-deficient RPE showed increased expression levels of oxidative stress-related genes and total glutathione levels. Furthermore, t-BHP induced a significant increase in cell death and GSSG levels in the KO RPE. CONCLUSIONS:We suggest that in the absence of the Kir.7.1 potassium channel, human RPE cells are vulnerable to oxidative stress and ultimately die. The dying/dead cells form aggregates and protrude from the surviving KCNJ13-deficient RPE sheet. |
| 巻・号 | 63(12) |
| ページ | 29 |
| 公開日 | 2022-11-1 |
| DOI | 10.1167/iovs.63.12.29 |
| PII | 2783877 |
| PMID | 36413373 |
| PMC | PMC9695160 |
| MeSH | Cell Death Gene Knockout Techniques Glutathione Disulfide / genetics Glutathione Disulfide / metabolism Humans Oxidative Stress* / physiology Retinal Pigment Epithelium* / metabolism |
| IF | 3.47 |
| リソース情報 | |
| ヒト・動物細胞 | 454E2(HPS0077) |