Loeser Richard F, Todd Marque D, Seely B Lynn
Department of Medicine, Rush Medical College of Rush-Presbyterian-St. Luke's Medical Center, Chicago, Illinois 60612, USA.
J Rheumatol. 2003 Jul;30(7):1565-70.
To determine the level of anabolic response when chondrocytes isolated from human osteoarthritic cartilage are stimulated with 2 doses of insulin-like growth factor-I (IGF-I) for extended culture periods.
Human chondrocytes were isolated from knee cartilage removed at the time of joint replacement surgery for osteoarthritis (OA). The cells were cultured in alginate beads under serum-free conditions and treated with 100 ng/ml or 1000 ng/ml of human recombinant IGF-I. Response was measured during culture periods of 1 to 28 days by determining the level of radiolabeled sulfate incorporated into alcian blue precipitable material and by measuring the level of total proteoglycan accumulation using the dimethylmethylene blue (DMB) assay. For the latter assay, cultures treated with osteogenic protein-1 (OP-1) were used for comparison to IGF-I. Results were normalized to cell numbers using DNA measurements.
The level of IGF-I stimulated sulfate incorporation relative to untreated controls increased with time in culture, with a peak response occurring between days 7 and 14 of culture. There was no significant difference between the 2 IGF-I doses. Despite the stimulation of sulfate incorporation, the DMB assay did not reveal a significant accumulation of proteoglycans in the cell-associated and further-removed matrix with either dose of IGF-I in cultures carried out to 21 days. In contrast, compared to controls, OP-1 at 100 ng/ml stimulated a 3-fold increase in matrix proteoglycan at day 21 of culture.
Prolonged IGF-I treatment of human OA chondrocytes in serum-free alginate cultures stimulated sulfate incorporation without significant accumulation of a proteoglycan matrix in longterm cultures. However, significant proteoglycan accumulation was seen in cultures treated with OP-1, suggesting it is a better stimulator of proteoglycan production by OA chondrocytes.
确定从人骨关节炎软骨中分离出的软骨细胞在延长培养期内用2种剂量的胰岛素样生长因子-I（IGF-I）刺激时的合成代谢反应水平。
从骨关节炎（OA）关节置换手术时切除的膝关节软骨中分离出人软骨细胞。将细胞在无血清条件下培养于藻酸盐珠中，并用100 ng/ml或1000 ng/ml的人重组IGF-I处理。在1至28天的培养期内，通过测定掺入阿尔新蓝可沉淀物质中的放射性标记硫酸盐水平以及使用二甲基亚甲基蓝（DMB）测定法测量总蛋白聚糖积累水平来测量反应。对于后一种测定法，将用成骨蛋白-1（OP-1）处理的培养物用于与IGF-I进行比较。使用DNA测量将结果标准化为细胞数量。
相对于未处理的对照，IGF-I刺激的硫酸盐掺入水平随培养时间增加，在培养的第7天至第14天之间出现峰值反应。2种IGF-I剂量之间无显著差异。尽管刺激了硫酸盐掺入，但在培养至21天的培养物中，DMB测定法未显示两种剂量的IGF-I在细胞相关和更远距离的基质中蛋白聚糖有显著积累。相比之下，与对照相比，100 ng/ml的OP-1在培养第21天时刺激基质蛋白聚糖增加了3倍。
在无血清藻酸盐培养物中，对人OA软骨细胞进行长时间IGF-I处理可刺激硫酸盐掺入，但在长期培养中蛋白聚糖基质无显著积累。然而，在用OP-1处理的培养物中观察到显著的蛋白聚糖积累，表明它是OA软骨细胞产生蛋白聚糖的更好刺激物。
