Tert butylhydroquinone tbhq

The human proximal tubular cell lines (HK-2 cells) were treated with rhSTC1 (50 ng/ml or 100 ng/ml) or (and) TBHQ (40 ng/ml) or (and) ML385 (10 μM) at the same time of iohexol (GE Healthcare, Shanghai, China) intervention to illuminate the effects of rhSTC1 on mitochondrial damage, ROS, inflammation, and cell apoptosis. rhSTC1 (Cat. No. 9400-SO-050) was purchased from R&D Systems (Minneapolis, MN, USA). Diluted by using distilled water, rhSTC1 protein was produced at a final concentration of 100 μg/ml. Tert-butylhydroquinone (TBHQ) and ML385 were purchased from Selleck (State of Texas, USA). STC1 siRNA and negative siRNA control were transfected into the HK-2 cells through using the Lipofectamine 2000 reagent (Life Technologies, USA).

In order to establish an LPS-induced cell injury model, human tubular epithelial (HK-2) cells were treated with 10, 20, or 30 μg/ml LPS for 6 h to select an optimal dosage of LPS. Then, HK-2 cells were treated with the optimal dosage of LPS for 0 h, 4 h, 6 h, or 8 h. According to the expression of Nrf2, HO-1, p62, LC3-II, and cleaved caspase 3, the optimal model of LPS-induced cell injury in HK-2 cells was established.
To elaborate the protective mechanism of the Nrf2 pathway, tert-butylhydroquinone (TBHQ, Selleckchem, Houston, TX, USA; Cat# S4990) and 3-methyladenine (3-MA, Selleckchem, Houston, TX, USA; Cat# S2767) were used to activate Nrf2 and inhibit autophagy, respectively. HK-2 cells were divided into four groups: the control group, LPS group, LPS+TBHQ group, and LPS+TBHQ+3-MA group, which were treated with LPS or LPS plus TBHQ 40 ng/ml or (and) 3-MA 10 mM for indicated times.

Blasticidin (AppliChem GmbH), RSL3 (Selleckchem), ML210 (Tocris), ML162 (Caymann), Erastin (Biomol), Sulfasalazine (SAS) (MedChemExpress), Imidazole Ketone Erastin (IKE) (Sellekchem), Ferrostatin-1 (Sigma Aldrich), Liproxstatin-1 (Biozol), Necrostatin-1s (Abcam), zVAD (ENZO), iFSP1 (Cayman Chemicals), Tert-butylhydroquinone (TBHQ) (Sellekchem), AMG510 (MedChemExpress), ARS1620 (Chemgood), PD184352 (Sigma Aldrich), MK2206 (Sellekchem), DRAQ7 (Biolegend), BODIPY C11 (Invitrogen), H2DCFDA (Invitrogen), Dharmafect I (Dharmacon), Puromycin (Sigma), Doxycycline hydrochloride (Alfa Aesar), 4-hydroxy-tamoxifen (4OHT) (Sigma), Polybrene (Merck), CaCl2 (Sigma Aldrich), HBS (Sigma Aldrich), propidium iodide (Sigma).