SKI306X inhibition of glycosaminoglycan degradation in human cartilage involves down-regulation of cytokine-induced catabolic genes

Abstract

Background/aims: SKI306X, a mixed extract of three herbs, Clematis mandshurica (CM), Prunella vulgaris (PV), and Trichosanthes kirilowii (TK), is chondroprotective in animal models of osteoarthritis (OA). The objectives of this study were to investigate its effect on interleukin (IL)-1β-induced degradation of glycosaminoglycan (GAG) and the basis of its action in human OA cartilage, as well as to screen for the presence of inhibitors of matrix metalloproteinase (MMP)-13 and a disintegrin and metalloprotease with thrombospondin motifs (ADAMTS)-4 in SKI306X and its component herbs, as well as in fractions from SKI306X.

Methods: Human OA chondrocytes and cartilage explants were obtained during total knee replacements and incubated with IL-1β ± oncostatin M with or without SKI306X or its component herb extracts. GAG degradation was assayed in cartilage explants using a commercial kit. Expression of genes involved in cartilage destruction was measured by real-time polymerase chain reaction using chondrocyte RNA. SKI306X was fractionated by preparative liquid chromatography to test for the presence of inhibitors of MMP-13 and ADAMTS-4.

Results: SKI306X and PV inhibited IL-1β-induced GAG release from cartilage explants, and SKI306X, CM, PV, and TK inhibited IL-1β-induced MMP gene expression. Unexpectedly, SKI306X greatly stimulated IL-1β + oncostatin M-induced ADAMTS-4 gene expression, probably due to its TK component. Some fractions of SKI306X also inhibited ADAMTS-4 activity.

Conclusions: SKI306X and its herbal components inhibit GAG degradation and catabolic gene expression in human OA chondrocytes and cartilage explants. SKI306X likely also contains one or more ADAMTS-4 inhibitor.

Keywords: Aggrecanase; Cartilage; Matrix metalloproteinase; Osteoarthritis; SKI306X.

Figure 1

Glycosaminoglycan (GAG) release from human osteoarthritis (OA) cartilage explants with interleukin (IL)-1β (10 ng/mL) in the absence or presence of an IL-1 receptor antagonist (IL-1Ra, 500 µg/mL) and SKI306X (200 µg/mL) or its herbal components, Clematis mandshurica (CM), Prunella vulgaris (PV), and Trichosanthes kirilowii (TK) (50 µg/mL). ^ap < 0.05 vs. control by t test, ^bp < 0.05 vs. IL-1β by one-way analysis of variance with a post hoc test.

Figure 2

(A-E) Changes in matrix metalloproteinase (MMP) gene expression in human osteoarthritic cartilage after culture with interleukin (IL)-1β (10 ng/mL) in the absence or presence of SKI306X (200 µg/mL) or its herbal components, Clematis mandshurica (CM), Prunella vulgaris (PV), and Trichosanthes kirilowii (TK) (50 µg/mL). Y axis: relative expression compared to control (Ctrl). ^ap < 0.05 vs. control by Mann-Whitney rank-sum test, ^bp < 0.05 vs. IL-1β by one-way analysis of variance with a post hoc test.

Figure 3

(A-C) Change in aggrecanase (a disintegrin and metalloprotease with thrombospondin motifs, ADAMTS) gene expression in human osteoarthritic cartilage after culture with interleukin (IL)-1β (0.02 ng/mL) + oncostatin M (OSM; 10 ng/mL) in the absence or presence of SKI306X (200 µg/mL) or its herbal components Clematis mandshurica (CM), Prunella vulgaris (PV), and Trichosanthes kirilowii (TK) (50 µg/mL). Y axis: relative expression compared to control (Ctrl). ^ap < 0.05 vs. control by Mann-Whitney rank-sum test, ^bp < 0.05 vs. IL-1β by one-way analysis of variance with a post hoc test.

Figure 4

(A) Changes in aggrecan (AGC1) and (B) collagen type II (COL2A1) gene expression in human osteoarthritic cartilage after culture with interleukin (IL)-1β (10 ng/mL) in the absence or presence of SKI306X (200 µg/mL) or its herbal components, Clematis mandshurica (CM), Prunella vulgaris (PV), and Trichosanthes kirilowii (TK) (50 µg/mL). Y-axis: relative expression compared to the control (Ctrl). Data are expressed as means ± SD of 10 chondrocyte strains. ^ap < 0.05 vs. control by Mann-Whitney rank-sum test, ^bp < 0.05 vs. IL-1β by one-way analysis of variance with a post hoc test.

Figure 5

(A-C) Changes in tissue inhibitors of metalloproteinase (TIMP) gene expression in human osteoarthritic cartilage after culture with interleukin (IL)-1β (10 ng/mL) in the absence or presence of SKI306X (200 µg/mL) or its herbal components, Clematis mandshurica (CM), Prunella vulgaris (PV), and Trichosanthes kirilowii (TK) (50 µg/mL). Y-axis: relative expression compared to the control (Ctrl). ^ap < 0.05 vs. control by t test or Mann-Whitney rank-sum test (TIMP-3), ^bp < 0.05 vs. IL-1β by one-way analysis of variance with a post hoc test.