Shikhman A R, Brinson D C, Lotz M
The Scripps Research Institute, and Scripps Clinic, La Jolla, California 92037, USA.
Arthritis Rheum. 2000 Jun;43(6):1307-14. doi: 10.1002/1529-0131(200006)43:6<1307::AID-ANR13>3.0.CO;2-3.
To determine enzymatic activities of the 8 key glycosaminoglycan-degrading glycosidases and glycoside sulfatases in cultured human articular chondrocytes and in synovial fluid from patients with osteoarthritis.
The following enzymes were analyzed: hexosaminidase and its isoenzyme A, N-acetyl-alpha-D-glucosaminidase, beta-galactosidase, beta-glucuronidase, alpha-L-iduronidase, aryl sulfatase, and galactose-6-sulfate sulfatase. Activity of the selected enzymes was analyzed by fluorometry with the aid of 4-methylumbelliferryl derivatives of the appropriate monosaccharides.
Hexosaminidase was found to be the dominant enzyme released by chondrocytes into the extracellular compartment. Stimulation of chondrocytes with interleukin-1beta resulted in a selective increase of the extracellular hexosaminidase activity and, to a lesser degree, of the extracellular beta-galactosidase activity, without significant changes in the activity of the other studied enzymes. Analysis of the pH dependency of the enzymatic activities revealed that even at neutral pH, hexosaminidase expressed a measurable activity, much higher than the activity of the other studied enzymes. Chondrocyte apoptosis did not result in increased extracellular glycosidase activities, including hexosaminidase activity. The spectrum of glycosidase and glycoside sulfatase activities in the synovial fluid from patients with osteoarthritis was similar to that in cultured human articular chondrocytes.
These data support the concept that lysosomal glycosidases, in particular hexosaminidase, represent a distinct subset of cartilage matrix-degrading enzymes that are activated by proinflammatory stimuli.
测定培养的人关节软骨细胞及骨关节炎患者滑液中8种关键的糖胺聚糖降解糖苷酶和硫酸酯酶的酶活性。
分析以下几种酶：己糖胺酶及其同工酶A、N - 乙酰 - α - D - 葡糖胺酶、β - 半乳糖苷酶、β - 葡糖醛酸酶、α - L - 艾杜糖醛酸酶、芳基硫酸酯酶和半乳糖 - 6 - 硫酸酯硫酸酯酶。借助适当单糖的4 - 甲基伞形酮衍生物，通过荧光法分析所选酶的活性。
发现己糖胺酶是软骨细胞释放到细胞外区室的主要酶。用白细胞介素 - 1β刺激软骨细胞导致细胞外己糖胺酶活性选择性增加，细胞外β - 半乳糖苷酶活性也有较小程度增加，而其他所研究酶的活性无显著变化。对酶活性的pH依赖性分析表明，即使在中性pH下，己糖胺酶仍表现出可测量的活性，远高于其他所研究酶的活性。软骨细胞凋亡并未导致包括己糖胺酶活性在内的细胞外糖苷酶活性增加。骨关节炎患者滑液中糖苷酶和硫酸酯酶活性谱与培养的人关节软骨细胞中的相似。
这些数据支持以下概念，即溶酶体糖苷酶，特别是己糖胺酶，代表了一类由促炎刺激激活的独特的软骨基质降解酶。
