Targeting Keap1 by miR-626 protects retinal pigment epithelium cells from oxidative injury by activating Nrf2 signaling

Xu, Xiangzhong; Tang, Yu; Cheng, Libo; Yao, Jin; Jiang, Qin; Li, Ke-ran; Zhen, Yunfang

doi:10.1016/j.freeradbiomed.2019.08.024

Cited by 36 publications

(47 citation statements)

References 37 publications

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“…Keap1 3′-UTR reporter plasmid (containing the miR-941-binding sites, at position of 276-283) was generated using the same protocol described previously [31], which was transfected to human endometrial cells using the Lipofectamine 2000 protocol. Afterwards, cells were subjected to the applied genetic modifications, with the Keap1 3′-UTR luciferase activity tested through the Promega kit [40].…”

Section: Keap1 3′-utr Activitymentioning

confidence: 99%

“…Following the applied OGDR stimulation, the superoxide colorimetric assay kit (BioVision, Shanghai, China) was applied to examine the cellular superoxide contents. In brief, the superoxide detection reagent (50 μL/well) was added for 15 min under the dark, with the superoxide's absorbance tested at the test-wavelength of 450 nm [31].…”

Section: Superoxide Detectionmentioning

confidence: 99%

“…As reported [31] endometrial cells with the applied genetic treatments were initially seeded onto the six-well tissue-culturing plates (at 1 × 10 5 cells per well). Following the applied OGDR stimulation, the lipid peroxidation assay kit (Abcam, Shanghai, China) was utilized to examine cellular lipid peroxidation levels, via the malondialdehyde method.…”

Section: Lipid Peroxidation Assaymentioning

confidence: 99%

“…We have previously reported that KGF-induced Nrf2 activation efficiently protected endometrial cells from OGDR [4]. MicroRNAs (miRNAs) are essential and novel players in regulating activation of the Keap1-Nrf2 pathway [31][32][33][34][35][36]. These2 2-nt long single-strand non-coding RNAs (ncRNAs) bind to the 3′-untranslated region (3′-UTR) of the targeted mRNAs, causing degradation and/or translation repression [37,38].…”

Section: Introductionmentioning

confidence: 99%

“…These2 2-nt long single-strand non-coding RNAs (ncRNAs) bind to the 3′-untranslated region (3′-UTR) of the targeted mRNAs, causing degradation and/or translation repression [37,38]. A potential strategy to activate the Nrf2 signaling cascade is to reduce Keap1 protein levels by expressing Keap1-targeting miRNAs [31,35,36]. Here we show that microRNA-941 ("miR-941") targets Keap1 to activate Nrf2 and protects endometrial cells from OGDR-induced cytotoxicity.…”

Section: Introductionmentioning

confidence: 99%

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Keap1-targeting microRNA-941 protects endometrial cells from oxygen and glucose deprivation-re-oxygenation via activation of Nrf2 signaling

Cheng

et al. 2020

Cell Commun Signal

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Background: Mimicking ischemia-reperfusion injury, oxygen and glucose deprivation (OGD)-re-oxygenation (OGDR) applied to endometrial cells produces significant oxidative stress and programmed necrosis, which can be inhibited by nuclear-factor-E2-related factor 2 (Nrf2) signaling. MicroRNA (miRNA)-induced repression of Keap1, a Nrf2 suppressor protein that facilitates Nrf2 degradation, is novel strategy to activate Nrf2 cascade. Methods: MicroRNA-941 (miR-941) was exogenously expressed in HESC and primary human endometrial cells, and the Nrf2 pathway examined by Western blotting and real-time quantitative PCR analysis. The endometrial cells were treated with OGDR, cell programmed necrosis and apoptosis were tested. Results: MiR-941 is a novel Keap1-targeting miRNA that regulates Nrf2 activity. In T-HESC cells and primary human endometrial cells, ectopic overexpression of miR-941 suppressed Keap1 3′-UTR (untranslated region) expression and downregulated its mRNA/protein expression, leading to activation of the Nrf2 cascade. Conversely, inhibition of miR-941 elevated Keap1 expression and activity in endometrial cells, resulting in suppression of Nrf2 activation. MiR-941 overexpression in endometrial cells attenuated OGDR-induced oxidative stress and programmed necrosis, whereas miR-941 inhibition enhanced oxidative stress and programmed necrosis. MiR-941 overexpression and inhibition were completely ineffective in Keap1−/Nrf2-KO T-HESC cells (using CRISPR/Cas9 strategy). Restoring Keap1 expression, using an UTR-depleted Keap1 construct, abolished miR-941-induced anti-OGDR activity in T-HESC cells. Thus Keap1-Nrf2 cascade activation is required for miR-941-induced endometrial cell protection. Conclusions: Targeting Keap1 by miR-941 activates Nrf2 cascade to protect human endometrial cells from OGDRinduced oxidative stress and programmed necrosis.

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Section: Keap1 3′-utr Activitymentioning

confidence: 99%

Section: Superoxide Detectionmentioning

confidence: 99%

Section: Lipid Peroxidation Assaymentioning

confidence: 99%