Optimum temperature for extracellular matrix production by articular chondrocytes
Optimum temperature for extracellular matrix production by articular chondrocytes
复制标题
DOI：
10.3109/02656736.2014.880857
复制
发表时间：
2014-01-01
影响因子：
3.1
通讯作者：
Kuroki, Hiroshi
中科院分区：
文献类型：
--
作者：
Ito, Akira;Aoyama, Tomoki;Kuroki, Hiroshi
目的：探讨温度对软骨细胞生成细胞外基质（ECM）能力的影响。材料和方法：从猪膝关节中分离关节软骨细胞。软骨细胞在三种不同的温度下培养：32℃、37℃和41℃。在颗粒培养系统中，通过基因表达分析、组织学和生化评估来评估产生ECM的能力。结果：低温培养21天后产生的微球湿重明显变重。用于评价胶原蛋白生成的Picrosirius红染色在较低温度下较高，用于评价硫酸糖胺聚糖（GAG）的safranin-O染色在32℃时比在37℃和41℃时低。胶原型IIA1 mRNA的表达在41℃时明显上调。然而，Picrosirius红染色在41℃时被抑制。分别。与32℃和37℃相比，41℃时每个球粒的GAG含量显著降低，低温时每个球粒的DNA含量较大。与37℃和41℃相比，32℃下的GAG含量与每个颗粒的DNA含量归一化显著降低。结论：我们的结果表明，大约41℃的培养温度通过降低DNA含量和可能通过胶原蛋白错误折叠来抑制ECM的产生。综上所述，关节软骨细胞产生ECM的最佳温度可能在32℃至37℃之间。
Purpose: The purpose of this study was to investigate the influence of temperature on the ability of the chondrocytes to produce extracellular matrix (ECM). Materials and methods: Articular chondrocytes were isolated from porcine knee joints. The chondrocytes were cultured at three different temperatures: 32 degrees C, 37 degrees C, and 41 degrees C. The ability to produce ECM was assessed by gene expression analysis, histological, and biochemical evaluation in a pellet culture system. Results: Wet weight of the pellets generated after 21 days, was significantly heavier when cultured at lower temperatures. Picrosirius red staining, employed to evaluate collagen production, was higher at lower temperatures, and safranin-O staining, used to evaluate sulphated glycosaminoglycan (GAG), was lower at 32 degrees C than at 37 degrees C and 41 degrees C. Collagen type IIA1 mRNA expression was markedly up-regulated at 41 degrees C. However, picrosirius red staining was inhibited at 41 degrees C. GAG and DNA content were measured by 1,9-dimethylmethylene blue (DMMB) assay and PicoGreen (R) assay, respectively. The GAG content per pellet was significantly low at 41 degrees C compared to that at 32 degrees C and 37 degrees C. The DNA content per pellet was larger at lower temperatures. The GAG content normalised with the DNA content per pellet was significantly lower at 32 degrees C compared to that at 37 degrees C and 41 degrees C. Conclusion: Our results suggest that a culture temperature of approximately 41 degrees C inhibits ECM production by decreasing DNA content and perhaps by collagen misfolding. Taken together, the optimum temperature for ECM production in articular chondrocytes may be between 32 degrees C and 37 degrees C.