The effect of glycosaminoglycan loss on chondrocyte viability - A study on porcine cartilage explants
The effect of glycosaminoglycan loss on chondrocyte viability - A study on porcine cartilage explants
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DOI：
10.1002/art.23381
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发表时间：
2008-04-01
影响因子：
--
通讯作者：
Lotz, Martin
中科院分区：
文献类型：
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作者：
Otsuki, Shuhei;Brinson, Diana C.;Lotz, Martin
客观的。糖胺聚糖 (GAG) 损失是骨关节炎的早期事件。最近的研究结果表明，关节炎软骨中的细胞死亡增加，并与细胞外基质降解有关。本研究的目的是分析机械损伤后 GAG 丢失对软骨细胞存活和细胞死亡的直接影响。在猪膝关节的全层软骨外植体中，通过软骨素酶 ABC 消化，GAG 被耗尽。外植体受到单次冲击机械损伤。通过活/死细胞测定、活性 caspase 3 染色以及对 caspase 抑制剂的敏感性来分析细胞活力和细胞死亡类型。结果。 GAG 消耗并不会直接导致细胞死亡增加。在软骨素酶 ABC 处理的外植体中，机械损伤导致细胞活力立即降低（从 84.6% 降至 71.0%），但在对照外植体中则不然。减少在表层区域显着。这种立即细胞死亡并未被泛半胱天冬酶抑制剂 Z-VAD-FMK 抑制，表明细胞坏死。在随后的培养过程中，这些外植体的活力进一步下降，第 3 天降至 50.5%。在软骨素酶 ABC 处理的外植体中添加 Z-VAD-FMK 可以减少第二波细胞死亡，并且还与 caspase 3 的激活相关，表明细胞死亡的凋亡机制。结论。这些结果表明，单独的 GAG 丢失并不会直接导致软骨细胞死亡。为了响应机械损伤，会立即诱导坏死性细胞死亡，这种情况仅在 GAG 耗尽的外植体中可见，并且主要在浅表区域。在随后的培养过程中，细胞死亡通过细胞凋亡机制传播。
Objective. Loss of glycosaminoglycan (GAG) is an early event in osteoarthritis. Recent findings showed increased cell death in arthritic cartilage and linkage with extracellular matrix degradation. The aim of this study was to analyze the direct effect of GAG loss on chondrocyte survival and cell death following mechanical injury.Methods. In full-thickness cartilage explants from porcine knee joints, GAG was depleted by digestion with chondroitinase ABC. Explants were subjected to single-impact mechanical injury. Cell viability and the types of cell death were analyzed by Live/Dead cell assay, staining for active caspase 3, and sensitivity to caspase inhibitor.Results. GAG depletion did not directly lead to increased cell death. In chondroitinase ABC-treated explants, but not in control explants, mechanical injury caused an immediate reduction in cell viability (from 84.6% to 71.0%); the reduction was prominent in the superficial zone. This immediate cell death was not inhibited by the pancaspase inhibitor Z-VAD-FMK suggesting cell necrosis. During subsequent culture, viability in these explants decreased further, to 50.5% on day 3. The second wave of cell death was reduced by the addition of Z-VAD-FMK in chondroitinase ABC-treated explants and was also associated with activation of caspase 3, suggesting apoptotic mechanisms of cell death.Conclusion. These results indicate that GAG loss alone does not directly lead to chondrocyte death. In response to mechanical injury, there is an immediate induction of necrotic cell death that is seen only in GAG-depleted explants and primarily in the superficial zone. During subsequent culture, cell death spreads via apoptotic mechanisms.