An in vitro study of micromechanics, cellular proliferation and viability on both decellularized porcine dura grafts and native porcine dura grafts

Abstract

Damage to the dura mater may occur during intracranial or spinal surgeries, which can result in cerebrospinal fluid leakage and other potentially fatal physiological changes. As a result, biological and synthetic derived scaffolds are typically used to repair dura mater post intracranial or spinal surgeries. The extracellular matrix of xenogeneic dura scaffolds has been shown to exhibit increased cell infiltration and regeneration than synthetic dura materials. In this study, we investigated the biocompatibility of native and decellularized porcine dura by seeding rat fibroblast cells onto the constructs. Cell proliferation, cell viability, and the mechanical properties of these dural grafts were evaluated post-re-seeding on days 3,7 and 14. Live-dead staining and resazurin salts were used to quantify cell viability and cell proliferation, respectively. Micro indentation was conducted to quantify the mechanical integrity of the native and acellular dura graft. The findings indicate that the acellular porcine dura graft creates a beneficial setting for infiltrating rat fibroblast cells. Cell viability, proliferation, and micro indentation results on the acellular grafts are comparable with the native control porcine dura tissue. In conclusion, the porcine scaffold material showed increased cell viability at each time point evaluated. The sustained mechanical response and favorable viability of the cells on the decellularized grafts provide promising insight into the potential use of porcine dura in clinical cranial dura mater graft applications.

Keywords: Cranial dura mater; Decellularization; Native dura; Porcine dura.

Fig. 1.

H &E stained images of dura mater in the transverse direction at 20X magnification (scale Bar 100 μm) at 3, 7, and 14 days after re-seeding compared with decellularized and native tissue (red arrow represents the presence of cells).

Fig. 2.

The area fraction of collagen fibers of native dura, decellularized dura pre-seeding, and re-seeded dura tissue at prescribed time points (3 days, 7 days, and 14 days) (no significant differences between groups (‘NS’ represents no significant differences).

Fig. 3.

Bar graph of cell count of re-seeded dura mater at 3 days, 7 days, and 14 days post re-seeding compared with native tissue (Location for cell count: four corners and two central areas)(‘***’ represent significant difference $P<0.001$).

Fig. 4.

Live dead staining image of the fibroblast cells grown on the prepared sample after 3 days,7 days, and 14 days of incubation compared with native dura matter. Live cells are shown in green (scale Bar 300 μm).

Fig. 5.

Proliferation of fibroblast cells on the prepared sample using percentage reduction of resazurin assay using fluorescence (‘NS’ represent no significant differences, while ‘**’, represent significant differences($P<0.01$).

Fig. 6.

Bar Graph of effective modulus from micro indentation at 3, 7, and 14 days after re-seeding on native and decellularized dura (‘NS’ on all bar graphs represents no significant differences).