Process-induced extracellular matrix alterations affect the mechanisms of soft tissue repair and regeneration

Abstract

Extracellular matrices derived from animal tissues for human tissue repairs are processed by various methods of physical, chemical, or enzymatic decellularization, viral inactivation, and terminal sterilization. The mechanisms of action in tissue repair vary among bioscaffolds and are suggested to be associated with process-induced extracellular matrix modifications. We compared three non-cross-linked, commercially available extracellular matrix scaffolds (Strattice, Veritas, and XenMatrix), and correlated extracellular matrix alterations to in vivo biological responses upon implantation in non-human primates. Structural evaluation showed significant differences in retaining native tissue extracellular matrix histology and ultrastructural features among bioscaffolds. Tissue processing may cause both the condensation of collagen fibers and fragmentation or separation of collagen bundles. Calorimetric analysis showed significant differences in the stability of bioscaffolds. The intrinsic denaturation temperature was measured to be 51°C, 38°C, and 44°C for Strattice, Veritas, and XenMatrix, respectively, demonstrating more extracellular matrix modifications in the Veritas and XenMatrix scaffolds. Consequently, the susceptibility to collagenase degradation was increased in Veritas and XenMatrix when compared to their respective source tissues. Using a non-human primate model, three bioscaffolds were found to elicit different biological responses, have distinct mechanisms of action, and yield various outcomes of tissue repair. Strattice permitted cell repopulation and was remodeled over 6 months. Veritas was unstable at body temperature, resulting in rapid absorption with moderate inflammation. XenMatrix caused severe inflammation and sustained immune reactions. This study demonstrates that extracellular matrix alterations significantly affect biological responses in soft tissue repair and regeneration. The data offer useful insights into the rational design of extracellular matrix products and bioscaffolds of tissue engineering.

Keywords: Biological scaffold; bovine pericardium; decellularization; extracellular matrix; matrix remodeling; porcine dermis; soft tissue regeneration; tissue engineering.

Figure 1.

Histological micrographs of bioscaffolds. Histological micrographs of three commercially available bioscaffolds (cross sections with hematoxylin and eosin stain): (a) Strattice, (b) Veritas, and (c) XenMatrix.

Figure 2.

Micro- and ultrastructural features of bioscaffolds. Scanning electron micrographs of three commercially available bioscaffolds: (a–d) Strattice, (e–h): Veritas, and (i–l) XenMatrix. (a, b, e, f, i, and j) Cross sections and (c, d, g, h, k, and l) surface views.

Figure 3.

Thermograms of unprocessed animal tissues. Calorimetric thermograms of fresh bovine pericardium and porcine dermis at the scan rate of 3°C/min. Three independent test curves are superimposed for each material type. T_d denotes the onset denaturation temperature.

Figure 4.

Thermograms of bioscaffold denaturation. Calorimetric thermograms of three bioscaffolds at the scan rate of 3°C/min. Note that significant modifications of extracellular matrices are observed in finished bioscaffold products when compared to unprocessed fresh tissues (see Figure 3).

Figure 5.

Effect of scan rates on bioscaffold denaturation temperatures: (a) Dependence of the onset denaturation temperatures (T_d) of bioscaffolds on the scan rate. Arrows indicate the extrapolated intrinsic T_d. (b) Thermal stability of bioscaffolds at different temperatures as measured by the duration of time needed to denature ~11% ECM (between the onset and peak temperature). The inset shows the method of calculating the duration of denaturation time. ECM: extracellular matrix; pd-ECM: minimally processed porcine dermis ECM; bp-ECM: minimally processed bovine pericardium ECM; DSC: differential heating calorimetry.

Figure 6.

Susceptibility of bioscaffolds to collagenase. Susceptibility of three bioscaffolds to type I collagenase at 37°C: (a) Strattice and XenMatrix are compared with pd-ECM and (b) Veritas is compared with bp-ECM. Data are mean ± standard deviation. ECM: extracellular matrix; pd-ECM: minimally processed porcine dermis ECM; bp-ECM: minimally processed bovine pericardium ECM.

Figure 7.

Contraction of implanted bioscaffolds. Changes of the size of implanted scaffolds after implantation in African green monkeys. All scaffold grafts were trimmed to 7 cm × 3 cm (i.e. 21 cm²) at the time of implantation. Explanted grafts were smaller than the originally implanted grafts due to graft contraction and/or resorption. Data are mean ± standard deviation.

Figure 8.

Non-human primate cellular responses. Histological micrographs (cross sections with hematoxylin and eosin stain) of explanted bioscaffolds after implantation in African green monkey for (a, d, and g) 1, (b, e, and h) 3 and (c, f, and i) 6 months. (a–c) Strattice, (d–f): Veritas, and (g–i): XenMatrix. The insets are representative micrographs at a high magnification showing the cell types in grafted materials. The bracket points to the implanted material.

Figure 9.

Non-human primate immunological responses. Elevation of systemic antibody (IgG) titer in the serum of animals after implantation of bioscaffolds over 6 months. The fold increase in antibody titer as compared with serum obtained prior to implantation was plotted on a logarithmic scale. Data are mean ± standard deviation.