Patchouli Alcohol Inhibits D-Gal Induced Oxidative Stress and Ameliorates the Quality of Aging Cartilage via Activating the Nrf2/HO-1 Pathway in Mice

Figure 4

Influences of patchouli alcohol (PA) on oxidative stress and expression of Nrf2/HO-1 signaling pathway of chondrocytes mediated by D-gal. (a) Effect of PA on gene expression of catalase (Cat), glutathione synthetase (Gss), and superoxide dismutase 1 (Sod1) in D-gal-caused senescent chondrocytes was analyzed by RT-qPCR. (b) PA’s effects upon antioxidant indices (CAT and SOD) generation by chondrocytes administrated with D-gal (5 mg/ml) and PA (0, 2.5, 5, and 10 μM) for 24 h were determined by biochemical kit. (c) Representative fluorescent images of ROS levels in D-gal (5 mg/ml) induced senescent chondrocytes treated with 10 μM PA for 24 h. Scale bar, 50 μm. (d) The RT-qPCR was employed to detect the gene expression of Nrf2/HO-1 pathway, including nuclear factor-erythroid 2-related factor-2 (Nrf2), heme oxygenase-1 (Hmox1), NAD(P)H quinone dehydrogenase 1 (Nqo1), and Kelch-like ECH-associated protein 1 (Keap1), mediated by D-gal in aging chondrocytes. (e) Representative immunofluorescence pictures of Nrf2 were captured in D-gal (5 mg/ml) induced senescent chondrocytes. Nrf2 was stained with Nrf2-specific antibody (red staining), while the nucleus was stained with DAPI (blue staining). Scale bar, 50 μm. (f) The expression and quantitative analysis of Nrf2 protein in the nucleus and HO-1 protein in the cytoplasm were assessed through western blotting. The Nrf2 and HO-1 protein expression levels are the gray value ratio of the target protein to lamin B and β-actin, respectively. . and vs. control group; ^# vs. D-gal group.

(a)

(b)

(c)

(d)

(e)

(f)