Yang Ya-li, Kaufman Laura J
Department of Chemistry, Columbia University, New York, New York 10027, USA.
Biophys J. 2009 Feb 18;96(4):1566-85. doi: 10.1016/j.bpj.2008.10.063.
In this work, the gelation of three-dimensional collagen and collagen/hyaluronan (HA) composites is studied by time sweep rheology and time lapse confocal reflectance microscopy (CRM). To investigate the complementary nature of these techniques, first collagen gel formation is investigated at concentrations of 0.5, 1.0, and 1.5 mg/mL at 37 degrees C and 32 degrees C. The following parameters are used to describe the self-assembly process in all gels: the crossover time (t(c)), the slope of the growth phase (k(g)), and the arrest time (t(a)). The first two measures are determined by rheology, and the third by CRM. A frequency-independent rheological measure of gelation, t(g), is also measured at 37 degrees C. However, this quantity cannot be straightforwardly determined for gels formed at 32 degrees C, indicating that percolation theory does not fully capture the dynamics of collagen network formation. The effects of collagen concentration and gelation temperature on k(g), t(c), and t(a) as well as on the mechanical properties and structure of these gels both during gelation and at equilibrium are elucidated. Composite collagen/HA gels are also prepared, and their properties are monitored at equilibrium and during gelation at 37 degrees C and 32 degrees C. We show that addition of HA subtly alters mechanical properties and structure of these systems both during the gelation process and at equilibrium. This occurs in a temperature-dependent manner, with the ratio of HA deposited on collagen fibers versus that distributed homogeneously between fibers increasing with decreasing gelation temperature. In addition to providing information on collagen and collagen/HA structure and mechanical properties during gelation, this work shows new ways in which rheology and microscopy can be used complementarily to reveal details of gelation processes.
在本研究中，通过时间扫描流变学和延时共聚焦反射显微镜（CRM）研究了三维胶原蛋白和胶原蛋白/透明质酸（HA）复合材料的凝胶化过程。为了研究这些技术的互补性，首先在37℃和32℃下，对浓度为0.5、1.0和1.5mg/mL的胶原蛋白凝胶形成过程进行了研究。以下参数用于描述所有凝胶中的自组装过程：交叉时间（t(c)）、生长阶段的斜率（k(g)）和停滞时间（t(a)）。前两个测量值通过流变学确定，第三个通过CRM确定。还在37℃下测量了凝胶化的频率无关流变学测量值t(g)。然而，对于在32℃下形成的凝胶，无法直接确定该量，这表明渗流理论不能完全捕捉胶原蛋白网络形成的动力学。阐明了胶原蛋白浓度和凝胶化温度对k(g)、t(c)和t(a)的影响，以及对这些凝胶在凝胶化过程中和平衡时的力学性能和结构的影响。还制备了复合胶原蛋白/HA凝胶，并在37℃和32℃下对其在平衡状态和凝胶化过程中的性能进行了监测。我们表明，添加HA会在凝胶化过程中和平衡状态下微妙地改变这些系统的力学性能和结构。这种情况以温度依赖的方式发生，随着凝胶化温度的降低，沉积在胶原纤维上的HA与均匀分布在纤维之间的HA的比例增加。除了提供有关凝胶化过程中胶原蛋白和胶原蛋白/HA结构及力学性能的信息外，这项工作还展示了流变学和显微镜可以互补使用以揭示凝胶化过程细节的新方法。
