Peripheral nerve injuries (PNI) affect more than 2 million people in the United States. For injuries with long gap transections (>30 mm in human), various synthetic polymers and natural materials have been investigated as potential matrices for generating nerve grafts. Among these, tissue-specific decellularised nerve matrices (dNMs) have attracted much attention since they preserve the main bioactive molecules like extracellular matrix (ECM) proteins (matrisome), which benefits PNI treatment. Previous studies have reported that decellularised nerve hydrogels (dNHs) promote neurite remyelination and inhibit synaptogenesis. However, the underlying molecular mechanism remains unclear.

In this study, dNHs from rat sciatic nerves were prepared and their effects on the neurite behaviours of Schwann cell (SC) and dorsal root ganglion (DRG) were examined.  Additionally, proteomic profiles were assessed and compared to commercial Rat Tail Type I Collagen (Col) hydrogels. In general, dNHs significantly enhanced the proliferation of SCs and neurite outgrowth of SCs and DRG compared to their Col counterparts. Through proteomic analysis, it was found that 48 proteins, 34 extracellular and 14 intracellular proteins, were found by dNH and Col hydrogels. In dNHs, core matrisomes including collagen, glycoproteins, and proteoglycans and matrisome-associated proteins such as ECM-affiliated proteins and ECM regulators occupied similar proportions, while Col had the highest proportion of collagens relative to the number of identified proteins. Among the matrisome proteins, 31 and 12 were found in dNH and Col samples respectively, and 8 matrisomes were found in both. Interestingly, collagen type III alpha 1 (COL3A1) and collagen type V alpha 1 (COL5A1) were abundantly and exclusively found in dNHs. Glycoproteins, including laminin β2 (LAMB2), Nidogen 2(NID2), and fibronectin 1(FN1), were also only detected in dNHs. Further investigations revealed that the addition of exogenous COLV strengthened the Col hydrogel’s ability to promote the proliferation of SC, while LAM significantly improved the neurite outgrowth of SCs and DRG.

How was PEAKS used?

Raw data were collected on a Thermo Orbitrap Fusion Lumos Tribrid  and were analyzed using PEAKS Studio 10.5. Oxidation (M) and acetylation (NQ) were included as variable post translational modifications (PTM) and carbamidomethylation was set as a fixed PTM. The parameters were set with a precursor mass tolerance of 12 ppm, a fragment mass tolerance of 0.02 Da, and a maximum of 3 missed cleavages. Quantitative analysis was performed by label-free quantitation in PEAKS Studio.

Xue, Wen, et al. “Regulation of Schwann Cell and DRG Neurite Behaviors within Decellularized Peripheral Nerve Matrix.” ACS Applied Materials & Interfaces 14.7 (2022): 8693-8704. https://doi.org/10.1021/acsami.1c20320

Abstract

Decellularized nerve hydrogels (dNHs) containing bioactive molecules are promising biomaterials for peripheral nerve injury (PNI) treatment and have been extensively applied in clinical and preclinical practice. However, most previous research projects studied their influences on nerve-related cellular behaviors in two dimensions (2D) without taking hydrogel biomechanics into consideration. The molecular mechanisms underlying the beneficial microenvironment provided by dNHs also remain unclear. In this study, dNHs from rat sciatic nerves were prepared, and their effects on Schwann cell (SC) and dorsal root ganglion (DRG) neurite behaviors were evaluated and compared to commercial rat tail type I collagen (Col) hydrogels in three-dimensional (3D) environments. We found that dNHs could promote SC proliferation and neurite outgrowth, and both the hydrogel mechanics and components contributed to the dNH functionalization. Through proteomics analysis, we found that laminin (LAM) and type V collagen (COLV) exclusively and abundantly existed in dNHs. By adding exogenous LAM and COLV into Col hydrogels, we demonstrated that they regulated SC gene expression and that LAM could promote SC spreading and neurite outgrowth, while COLV improved SC proliferation. Lastly, dNHs were fabricated into paper-like, aligned nerve scaffolds through unidirectional freezing to expand the dNH applications in PNI treatment.