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TREK‑TRAAK two‑pore domain potassium channels protect human retinal pigment epithelium cells from oxidative stress.

International journal of molecular medicine (2018-08-15)
Hao Huang, Han Li, Kangpei Shi, Lei Wang, Xiaotong Zhang, Xiaobo Zhu
RESUMEN

The aim of the current study was to explore the potential of TREK‑TRAAK two‑pore domain potassium (K2P) channels in protecting human retinal pigment epithelium (hRPE) cells against oxidative stress. hRPE cells were obtained from donors, and then cell identification and detection of the expression levels of TREK‑TRAAK K2P channels in hRPE cells were conducted. Subsequently, tert‑butyl hydroperoxide (t‑BH) was used to induce oxidative stress in hRPE cells. Docosahexaenoic acid (DHA) was used to stimulate and fluoxetine was used to inhibit the TREK‑TRAAK K2P channels. The survival rates of hRPE cells under oxidative stress were examined using flow cytometry. Apoptosis‑associated factors, including Bax, Bcl‑2, cleaved‑caspase‑3, αB‑crystallin and their mRNAs, were examined using immunofluorescence, western blot and reverse transcription‑polymerase chain reaction analyses. Variations in the cytoarchitecture were observed by immunofluorescence and electron microscopy. The cells examined in the present study were identified as hRPE cells. All members in the TREK‑TRAAK K2P channel family (including TREK‑1, TREK‑2 and TRAAK) were found to be expressed in hRPE cells. Stimulation of TREK‑TRAAK K2P channels increased the survival rates of hRPE cells under oxidative stress and the levels of intracellular protective factors, such as Bcl‑2 and αB‑crystallin. By contrast, inhibition of these channels decreased the cell survival rates and increased apoptosis enhancing factors, such as Bax and cleaved‑caspase‑3. Further examination of the cytoarchitecture revealed that TREK‑TRAAK K2P channels protected the integrity of the hRPE cell structure against oxidative stress. In conclusion, the present study suggested that the activated TREK‑TRAAK K2P channels serve a role in protecting hRPE cells against the oxidative stress induced by t‑BH, which indicated that these K2P channels are potential novel targets in retinal protection and provided a new direction for research and therapy in retinal degeneration diseases.

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D.E.R. 332, used as embedding medium
Sigma-Aldrich
ANTI-CLEAVED-CASP3 (ASP175) antibody produced in rabbit, affinity isolated antibody, buffered aqueous solution