Regenerative Potential of Decellularized Porcine Nucleus Pulposus Hydrogel Scaffolds: Stem Cell Differentiation, Matrix Remodeling, and Biocompatibility Studies

Abstract

Nucleus pulposus (NP) tissue regeneration has been proposed as an early stage interventional therapy to combat intervertebral disc degeneration. We have previously reported on the development and characterization of a novel biomimetic acellular porcine NP (APNP) hydrogel. Herein, we aimed to evaluate this material for use as a suitable scaffold for NP tissue regeneration. Human-adipose-derived stem cells (hADSCs) were cultured for 14 days on APNP hydrogels in chemically defined differentiation media and were analyzed for an NP-cell-like mRNA expression profile, evidence of hydrogel remodeling including hydrogel contraction measurements, extracellular matrix production, and compressive dynamic mechanical properties. The innate capacity of the hydrogel itself to induce stem cell differentiation was also examined via culture in media lacking soluble differentiation factors. Additionally, the in vivo biocompatibility of non-crosslinked and ethyldimethylaminopropyl carbodiimide/N-hydroxysuccinimide and pentagalloyl glucose crosslinked hydrogels was evaluated in a rat subdermal model. Results indicated that hADSCs expressed putative NP-cell-positive gene transcript markers when cultured on APNP hydrogels. Additionally, glycosaminoglycan and collagen content of hADSC-seeded hydrogels was significantly greater than nonseeded controls and cell-seeded hydrogels exhibited evidence of contraction and tissue inhibitors of metalloproteinase-1 production. The dynamic mechanical properties of the hADSC-seeded hydrogels increased with time in culture in comparison to noncell-seeded controls and approached values reported for native NP tissue. Immunohistochemical analysis of explants illustrated the presence of mononuclear cells, including macrophages and fibroblasts, as well as blood vessel infiltration and collagen deposition within the implant interstices after 4 weeks of implantation. Taken together, these results suggest that APNP hydrogels, in concert with autologous ADSCs, may serve as a suitable scaffold for NP tissue regeneration.

Creation of an injectable in situ gelling native extracellular matrix for nucleus pulposus tissue engineering

“…An optimized decellularization process for nucleus pulposus needs to balance the removal of cells with the maintenance of sGAGs. The sGAGs are essential in maintaining the osmotic pressure of the nucleus pulposus [8], and they have the potential to play a role in driving cell phenotype [45][46][47]50,58] and preventing innervation [59][60][61][62]. The gentle decellularization protocol used in our laboratory for nerve [39,43,63] and lung tissue [38] was adapted for use in the nucleus pulposus by shortening the duration of wash cycles substantially.…”

“…Our laboratory has previously developed gentle decellularization techniques for nerve tissue to preserve essential proteins, proteoglycans, and microstructure [39,43], which would be ideal for the delicate nucleus pulposus. To date, some research has been performed to create decellularization protocols for the nucleus pulposus [44][45][46][47], anulus fibrosus [48,49], and the intact IVD [50]. These matrices have shown promise as viable scaffolds for IVD cells as well as drivers of stem cell fate [44][45][46][47][48][49][50], although they are not injectable or in situ gelling.…”

Pulmonary heart valve replacement using stabilized acellular xenogeneic scaffolds; effects of seeding with autologous stem cells

“…Valve-derived scaffolds are preferred because of easy accessibility, outstanding hemodynamic properties and the preservation of biological "niches" after decellularization (10)(11)(12). We also treat acellular scaffolds with penta-galloyl glucose (PGG) for tissue stabilization, to prevent premature scaffold degradation and to reduce calcification (13)(14)(15).…”

Generation of Bioartificial Salivary Gland Using Whole-Organ Decellularized Bioscaffold

“…Recent studies have shown that native organ-specific extracellular matrix (ECM) represents an ideal biochemical and biophysical scaffold material for such investigation [Gilbert et al, 2006;Badylak et al, 2009]. Native ECM consists of collagens, laminin, fibronectin and elastin, which play a central role in the physiological development, repair and regeneration of organs [Lu et al, 2013;Lynch and Ahearne, 2013;Mercuri et al, 2013;Naderi et al, 2013]. During the regenerative process, ECM not only provides the native spatial structure but also guides proper cellular interactions by releasing molecular cues [Song and Ott, 2011].…”