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2 protocols using mn sod

1

Evaluating Signaling Pathways in Cell Viability

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LIQ was purchased from Shang Hai Winherb Medical Science CO., Ltd. (White crystalline powder, purity >98%). LPS was purchased from Sigma-Aldrich (St. Louis, MO, United States). Compound C (S7840) was purchased from Selleck (Shanghai, China). Primary antibodies including anti-mTOR (T-mTOR, 2983), anti-phospho-mTOR (p-mTOR, 2971), anti-GAPDH (2118), anti-Bax (2772), anti-Bcl-2 (2870), anti-phospho-JNK (p-JNK, 4468), anti-JNK (T-JNK, 9258), anti-phospho-ERK (p-ERK, 4370), anti-ERK (T-ERK, 4695), anti-phospho-p38 (p-p38, 4511), anti-p38 (T-p38, 9212), anti-phospho-AKT (p-AKT, 4691), anti-AKT (T-AKT, 4060), anti-phospho-GSK3β (p-GSK3β, 9323), anti-GSK3β (T-GSK3β, 9315), anti-phospho-STAT3 (p-STAT3, 9136), and anti-STAT3 (T-STAT3, 9139) were purchased from Cell Signaling Technology. Primary antibodies, including anti-phospho-AMPKα2 (p-AMPKα2, ab109402), anti-AMPKα2 (T-AMPKα2, ab3760), anti-phospho-IκBα (ab133462), anti-IκBα (ab7217), anti-phospho-p65 (ab194726), and anti-p65 (ab16502) were obtained from Abcam (Cambridge, United Kingdom). Secondary antibodies were obtained from LI-COR Biosciences. The cell counting kit 8 (CCK-8) was purchased from Dojindo Molecular Technologies (Rockville, MD, United States), and the Cu/Zn-SOD, Mn-SOD and NADP+/NADPH assay kits were purchased from Beyotime (Shanghai, China).
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2

TBI-Induced Oxidative Stress Assay

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At 24 h after TBI, mice were sacrificed, their tissues obtained, lysed using a lysis buffer (Beyotime, Jiangsu, China) and homogenized for 30 min on ice. Supernatants were obtained after centrifugation (12,000 rpm) of samples for 10 min at 4 °C. Malondialdehyde (MDA) (Beyotime, Jiangsu, China), ATP (Beyotime, Jiangsu, China) levels and activities of oxidative stress-related enzymes, MnSOD (Beyotime, Jiangsu, China) and GSH-Px (Beyotime, Jiangsu, China) were assessed as instructed by the manufacturers’ corresponding commercial kits.
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