SIRT1 exerts protective effects against paraquat-induced injury in mouse type II alveolar epithelial cells by deacetylating NRF2 in vitro

Abstract

Silent information regulator 2-related enzyme 1 (SIRT1), a protein deacetylase, is known to strongly protect cells against oxidative stress-induced injury. The nuclear factor E2-related factor 2 (NRF2)-antioxidant response element (ARE) antioxidant pathway plays important regulatory roles in the antioxidant therapy of paraquat (PQ) poisoning. In the present study, we investigated whether the SIRT1/NRF2/ARE signaling pathway plays an important role in lung injury induced by PQ. For this purpose, mouse type II alveolar epithelial cells (AECs‑II) were exposed to various concentrations of PQ. The overexpression or silencing of SIRT1 was induced by transfecting the cells with a SIRT1 overexpression vector or shRNA targeting SIRT1, respectively. The protein expression levels of SIRT1 and NRF2 were measured by western blot analysis. The superoxide dismutase (SOD) and catalase (CAT) activities, as well as the glutathione (GSH) and malondialdehyde (MDA) levels were measured using respective kits. Heme oxygenase-1 (HO-1) activity was also determined by ELISA. In addition, cell apoptosis was determined by flow cytometry. The protein stability of NRF2 was analyzed using cycloheximide and its acetylation in the cells was also determined. The following findings were obtained: i) SIRT1 overexpression markedly increased NRF2 protein expression; ii) SIRT1 promoted the transcriptional activity of NRF2 and upregulated the expression of the NRF2 downstream genes, SOD, CAT, GSH and HO-1, thus inhibiting the apoptosis of AECs‑II; iii) the inhibition of SIRT1 activity further induced the production of malondialdehyde (MDA), which resulted in increased oxidative damage; iv) SIRT1 promoted the stability of NRF2 by regulating the deacetylation and activation of the NRF2/ARE antioxidant pathway. The findings of this study demonstrate that the protective effects of SIRT1 are associated with the activation of the NRF2/ARE antioxidant pathway in lung injury induced by PQ poisoning.

Crosstalk between Mitochondrial Fission and Oxidative Stress in Paraquat-Induced Apoptosis in Mouse Alveolar Type II Cells

“…Mouse AT-II cells were from our own laboratory 23 . Cells were cultured in DMEM containing 10% fetal bovine serum and 100U/ml antibiotic (penicillin and streptomycin) solution in a humidified incubator with 5% CO 2 atmosphere at 37℃.…”

Nrf2 Activation Induced by Sirt1 Ameliorates Acute Lung Injury After Intestinal Ischemia/Reperfusion Through NOX4-Mediated Gene Regulation

“…They propose that IIR may activate a cascade resulting in ROS accumulation, Sirt1 reduction, FOXO3 acetylation and MnSOD downregulation. Activation of the Nrf2/Sirt1 signalling pathway is also reported in a paraquat-induced lung injury model, where it is proposed that Sirt1 promoted the stability of Nrf2 by regulating the deacetylation and activation of the Nrf2/ARE antioxidant pathway [28]. …”

“…Upregulated Sirt1 expression was found to downregulate FOXO3 acetylation to confer a protective role in IIR acute lung injury by modulating the effects of downstream anti-oxidative and anti-apoptotic factors [1]. Furthermore, upregulating levels of Sirt1 has also been found to increase the nuclear translocation of Nrf2 which in turn inhibits mitogen-activated protein kinase phosphorylation and offers protection against cellular damage caused by oxidative stress [16-18]. …”

SIRT1 exhibits antioxidative effects in HT22 cells induced by tert‐butyl alcohol

“…MTBE and its major metabolite TBA have acute neurological, including behavioral and toxicological, effects through blood‐brain barrier . During the process of neurological damage caused by MTBE oxidative stress was involved . Therefore, we would like to know whether TBA, the main metabolite of MTBE, also impaired neurons via oxidative stress and during the process whether SIRT1 was implicated.…”

“…Otherwise, Sirtuin 1 (SIRT1) has been reported to be involved in these survival pathways by inhibiting apoptosis or cellular senescence induced by stresses, including DNA damage and oxidative stress . SIRT1, a deacetylase, was showed to reduce oxidative stress by deacetylating its substrates, such as forkhead transcription factors of the Oclass (FoxOs), nuclear factor E2‐related factor 2 (Nrf2), 66‐kDa Src homology 2 domain‐containing protein (p66Shc), and form a complex that induces ROS detoxifying mechanisms, such as SOD and catalase in response to oxidative stress …”