Fig 1.
(A) Structure of ectoine. (B) Ectoine improves the fluidity of the lipid head groups in the cell membrane and stabilizes macromolecules.
Table 1.
Primer sequences for RT‒qPCR.
Fig 2.
Effect of ectoine on trypsin resistance in chondrocytes.
With increasing Ectoine concentration, fewer chondrocytes contracted into spot-like cells after treatment with trypsin. (n≥3, scale bar: 10 μm).
Fig 3.
Cell viability of chondrocytes at 50°C determined by MTT assay.
As the ectoine concentration increased, the high-temperature tolerance of the chondrocytes gradually increased. (n≥6, one-way ANOVA, **P < 0.01, ***P < 0.001).
Fig 4.
ROS assay (A) Images of ROS in chondrocytes pretreated with ectoine or dexamethasone (DEX) and poststimulated with H2O2. (B) The mean fluorescence intensity in chondrocytes from the different concentrations of ectoine group was significantly lower than that in chondrocytes from the H2O2 group, and the fluorescence intensity in chondrocytes from the 1.5% ectoine group was significantly lower than that in chondrocytes from the DEX group. (n≥3, one-way ANOVA, scale bar: 10 μm, *P < 0.05, **P < 0.01, ***P < 0.001).
Fig 5.
COX-2, MMP-3, and MMP-9 relative expression in chondrocytes.
(A-C) Chondrocytes were pretreated with different concentrations of ectoine or DEX and then treated with IL-1β. COX-2, MMP-3, and MMP-9 were significantly downregulated in each ectoine group and were positively correlated with the ectoine concentration. (n≥3, one-way ANOVA, *P < 0.05, ***P < 0.001).
Fig 6.
Immunofluorescence staining and RT‒qPCR analysis of type II collagen.
(A). Immunofluorescence staining images of chondrocytes pretreated with ectoine or dexamethasone (DEX) and post-stimulated with IL-1β. (B) The fluorescence intensities of type II collagen in chondrocytes in the ectoine groups treated with different concentrations of ectoine were greater than those in the IL-1β group, but only the fluorescence intensity in the 1.5% ectoine group was significantly different from that in the DEX group. (C) The expression of Col2A1 in the different groups. (n≥3, one-way ANOVA, scale bar: 10 μm, *P < 0.05, **P < 0.01, ***P < 0.001).
Fig 7.
Effect of ectoine on OA development in the rat model.
(A) Typical H-E and S-O staining of cartilage and subchondral cortical bone from the different experimental groups at 8 w postsurgery. (B) Diagram showing the OARSI scores of the cartilage. (n = 10, one-way AVOVA, scale bar: 100 μm, ***P < 0.001).