OBJECTIVE: To identify the CD44-receptor-mediated effects of 5-7 x 10(5)MW hyaluronan (HA, Hyalgan) on cell viability in normal and damaged human chondrocyte primary cultures isolated from articular cartilage. DESIGN: Primary cultures of human chondrocytes were established from normal articular biopsies and expanded to the second culture passage. The dose-response effects of HA on the viability of normal cultures were identified. Chondrocytes were then treated with either hypoxanthine (2 mM) and xanthine oxidase (20-60 mU), or with activated polymorphonuclear leukocytes (PMNs) to induce injury. Damaged and control cells were then treated with 5-7 x 10(5)HA in the previously identified optimal dose of 0.05 mg/ml. Viability was assessed at specific time periods for the chemically and PMN-damaged cells. To identify if HA effects were mediated by the CD44 receptor, chondrocytes were incubated with anti-CD44 antibody at saturating concentrations (5 microg/ml for 100,000 cells) to produce a maximum inhibition of HA binding. Cells were evaluated using the MTT viability assay, histology, electron microscopy and immunohistochemistry. RESULTS: Direct addition of HA (optimal dose, 0.5 mg/ml) significantly increased cell survival in normal chondrocyte primary cultures (P<0.05). Similarly, addition of this same dose of HA to cultures of free radical-damaged chondrocytes, restored the viability to baseline conditions. Cell viability rates dropped significantly (P<0.05) when CD44 receptor binding was inhibited, indicating that cell growth was mediated by the CD44 receptor. CONCLUSIONS: HA (0.5 mg/ml of 5-7 x 10(5)) significantly increased the viability of normal human chondrocytes in primary culture and restored cell viability to near normal levels after oxidative cell injury.
The effect of hyaluronan on CD44- mediated proliferation of normal and hydroxyl radical-damaged chondrocytes.
BRUN, PAOLA;CORTIVO, ROBERTA;ABATANGELO, GIOVANNI
2003
Abstract
OBJECTIVE: To identify the CD44-receptor-mediated effects of 5-7 x 10(5)MW hyaluronan (HA, Hyalgan) on cell viability in normal and damaged human chondrocyte primary cultures isolated from articular cartilage. DESIGN: Primary cultures of human chondrocytes were established from normal articular biopsies and expanded to the second culture passage. The dose-response effects of HA on the viability of normal cultures were identified. Chondrocytes were then treated with either hypoxanthine (2 mM) and xanthine oxidase (20-60 mU), or with activated polymorphonuclear leukocytes (PMNs) to induce injury. Damaged and control cells were then treated with 5-7 x 10(5)HA in the previously identified optimal dose of 0.05 mg/ml. Viability was assessed at specific time periods for the chemically and PMN-damaged cells. To identify if HA effects were mediated by the CD44 receptor, chondrocytes were incubated with anti-CD44 antibody at saturating concentrations (5 microg/ml for 100,000 cells) to produce a maximum inhibition of HA binding. Cells were evaluated using the MTT viability assay, histology, electron microscopy and immunohistochemistry. RESULTS: Direct addition of HA (optimal dose, 0.5 mg/ml) significantly increased cell survival in normal chondrocyte primary cultures (P<0.05). Similarly, addition of this same dose of HA to cultures of free radical-damaged chondrocytes, restored the viability to baseline conditions. Cell viability rates dropped significantly (P<0.05) when CD44 receptor binding was inhibited, indicating that cell growth was mediated by the CD44 receptor. CONCLUSIONS: HA (0.5 mg/ml of 5-7 x 10(5)) significantly increased the viability of normal human chondrocytes in primary culture and restored cell viability to near normal levels after oxidative cell injury.Pubblicazioni consigliate
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