Long Rebecca A, Nagatomi Jiro, Chancellor Michael B, Sacks Michael S
Department of Bioengineering, University of Pittsburgh, Pittsburgh, PA 15219, USA.
Biomaterials. 2006 Apr;27(11):2398-404. doi: 10.1016/j.biomaterials.2005.10.011. Epub 2005 Dec 7.
We have previously observed that muscle-derived stem cells (MDSC) seeded onto porcine small intestinal submucosa (SIS) increase the mechanical compliance of the engineered tissue construct [Lu SH, Sacks MS, Chung SY, Gloeckner DC, Pruchnic R, Huard J, et al. Biaxial mechanical properties of muscle-derived cell seeded small intestinal submucosa for bladder wall reconstitution. Biomaterials 2005;26(4):443-9]. To date, however, the initial remodeling events which occur when MDSC are seeded onto SIS have yet to be elucidated. One potential mechanism responsible for the observed increase in mechanical compliance is the release of matrix metalloproteinase-I (MMP-I). To investigate this finding, MDSC ( approximately 1x10(6)) were cultured on single-layer SIS cell culture inserts (4.7 cm2) for 1-10 days. MDSC MMP-I activity on SIS in the supernatant at 1, 3, 5, 7, and 10 days was determined using a collagenase assay kit. MMP-I activity of the MDSC/SIS was significantly higher (p<0.0025) after one day in culture compared to specimens collected from subsequent time points and the unseeded control. To further study the initial remodeling events, the impact of MMP-I on mechanical compliance was examined. SIS was incubated with 0.16 U/mL collagenase-I for 3, 4.5, 5, and 24h, then biaxial mechanical testing was performed. After 5h of digestion with collagenase-I, mechanical compliance under 1 MPa peak stress was increased by 7% in the circumferential direction, compared to control SIS. These findings suggest that the release of MMP-I in response to initial seeding on SIS and subsequent breakdown of collagen fibers is the mechanism responsible for an increase in mechanical compliance.
我们之前观察到，接种在猪小肠黏膜下层（SIS）上的肌肉衍生干细胞（MDSC）可提高工程化组织构建体的机械顺应性[Lu SH, Sacks MS, Chung SY, Gloeckner DC, Pruchnic R, Huard J, 等人。用于膀胱壁重建的接种肌肉衍生细胞的小肠黏膜下层的双轴力学性能。生物材料2005;26(4):443 - 9]。然而，迄今为止，当MDSC接种到SIS上时发生的初始重塑事件尚未阐明。观察到的机械顺应性增加的一个潜在机制是基质金属蛋白酶 - I（MMP - I）的释放。为了研究这一发现，将MDSC（约1×10⁶个）接种在单层SIS细胞培养插入物（4.7 cm²）上培养1 - 10天。使用胶原酶检测试剂盒测定第1、3、5、7和10天培养上清液中SIS上MDSC的MMP - I活性。与后续时间点收集的标本和未接种的对照相比，培养一天后MDSC/SIS的MMP - I活性显著更高（p < 0.0025）。为了进一步研究初始重塑事件，研究了MMP - I对机械顺应性的影响。将SIS与0.16 U/mL胶原酶 - I孵育3、4.5、5和24小时，然后进行双轴力学测试。与对照SIS相比，用胶原酶 - I消化5小时后，在1 MPa峰值应力下圆周方向的机械顺应性增加了7%。这些发现表明，响应于最初接种到SIS上以及随后胶原纤维的分解而释放的MMP - I是机械顺应性增加的机制。
