Mauck R L, Wang C C-B, Oswald E S, Ateshian G A, Hung C T
Department of Biomedical Engineering, Columbia University, New York, NY 10027, USA.
Osteoarthritis Cartilage. 2003 Dec;11(12):879-90. doi: 10.1016/j.joca.2003.08.006.
Functional tissue engineering (FTE) of articular cartilage involves the use of physiologically relevant mechanical signals to encourage the growth of engineered constructs. The goal of this study was to determine the utility of deformational loading in enhancing the mechanical properties of chondrocyte-seeded agarose hydrogels, and to investigate the role of initial cell seeding density and nutrient supply in this process.
Chondrocyte-seeded agarose hydrogels were cultured in free-swelling conditions or with intermittent deformational loading (10% deformation, 1 Hz, 1 h on/ 1 h off, 3 h per day, five days per week) over a two-month culture period. Disks were seeded at lower (10 million cells/ml) and higher (60 million cells/ml) seeding densities in the context of a greater medium supply than previous studies (decreasing the number of cells/ml feed medium/day) and with an increasing concentration of fetal bovine serum (10 or 20% FBS).
Under these more optimal nutrient conditions, at higher seeding densities and high serum concentration (20% FBS), dynamically loaded constructs show >2-fold increases in material properties relative to free-swelling controls. After two months of culture, dynamically loaded constructs achieved a Young's modulus of approximately 185 kPa and a dynamic modulus (at 1 Hz) of approximately 1.6 MPa, with a frequency dependent response similar to that of the native tissue. These values represent approximately 3/4 and approximately 1/4 the values measured for the native tissue, respectively. While significant differences were found in mechanical properties, staining and bulk measurements of both proteoglycan and collagen content of higher seeding density constructs revealed no significant differences between free-swelling and loading groups. This finding indicates that deformational loading may act to increase material properties via differences in the structural organization, the production of small linker ECM molecules, or by modulating the size of macromolecular proteoglycan aggregates.
Taken together, these results point to the utility of dynamic deformational loading in the mechanical preconditioning of engineered articular cartilage constructs and the necessity for increasing feed media volume and serum supplementation with increasing cell seeding densities.
关节软骨的功能组织工程（FTE）涉及利用生理相关的机械信号来促进工程构建体的生长。本研究的目的是确定变形加载在增强接种软骨细胞的琼脂糖水凝胶力学性能方面的效用，并研究初始细胞接种密度和营养供应在此过程中的作用。
接种软骨细胞的琼脂糖水凝胶在自由膨胀条件下培养，或在为期两个月的培养期内进行间歇性变形加载（10%变形，1 Hz，开1小时/关1小时，每天3小时，每周5天）。与以往研究相比，在更大的培养基供应（降低每毫升培养基每天的细胞数量）以及胎牛血清浓度增加（10%或20%胎牛血清）的情况下，以较低（1000万个细胞/毫升）和较高（6000万个细胞/毫升）的接种密度接种圆盘。
在这些更优化的营养条件下，在较高接种密度和高血清浓度（20%胎牛血清）下，动态加载的构建体相对于自由膨胀对照显示出材料性能增加2倍以上。培养两个月后，动态加载的构建体实现了约185 kPa的杨氏模量和约1.6 MPa的动态模量（1 Hz时），其频率依赖性响应与天然组织相似。这些值分别约为天然组织测量值的3/4和约1/4。虽然在力学性能方面发现了显著差异，但对较高接种密度构建体的蛋白聚糖和胶原蛋白含量进行的染色及整体测量显示，自由膨胀组和加载组之间没有显著差异。这一发现表明，变形加载可能通过结构组织差异、小连接细胞外基质分子的产生或通过调节大分子蛋白聚糖聚集体的大小来增加材料性能。
综上所述，这些结果表明动态变形加载在工程化关节软骨构建体的机械预处理中的效用，以及随着细胞接种密度增加而增加培养基体积和血清补充的必要性。
