Seeding decellularized nerve allografts with adipose-derived mesenchymal stromal cells: An in vitro analysis of the gene expression and growth factors produced

J Plast Reconstr Aesthet Surg. 2019 Aug;72(8):1316-1325. doi: 10.1016/j.bjps.2019.04.014. Epub 2019 May 9.

Abstract

Mesenchymal stromal cells (MSCs) secrete many soluble growth factors and have previously been shown to stimulate nerve regeneration. MSC-seeded processed nerve allografts could potentially be a promising method for large segmental motor nerve injuries. Further progress in our understanding of how the functions of MSCs can be leveraged for peripheral nerve repair is required before making clinical translation. The present study, therefore, investigated whether interactions of adipose-derived MSCs with decellularized nerve allografts can improve gene and protein expression of growth factors that may support nerve regeneration. Human nerve allografts (n = 30) were decellularized and seeded with undifferentiated human adipose-derived MSCs. Subsequently, the MSCs and MSC-seeded grafts were isolated on days 3, 7, 14, and 21 in culture for RNA expression analysis by qRT-PCR. Evaluated genes included NGF, BDNF, PTN, GAP43, MBP, PMP22, VEGF, and CD31. Growth factor production was evaluated and quantified using enzyme-linked immunosorbent assay (ELISA). On day 21, semi-quantitative RT-PCR analysis showed that adherence of MSCs to nerve allografts significantly enhances mRNA expression of neurotrophic, angiogenic, endothelial, and myelination markers (e.g., BDNF, VEGF, CD31, and MBP). ELISA results revealed an upregulation of BDNF and reduction of both VEGF and NGF protein levels. This study demonstrates that seeding of undifferentiated adipose-derived MSCs onto processed nerve allografts permits the secretion of neurotrophic and angiogenic factors that can stimulate nerve regeneration. These favorable molecular changes suggest that MSC supplementation of nerve allografts may have potential in improving nerve regeneration.

Keywords: Adipose-derived stem cells; Gene expression; MSCs; Peripheral nerve; Processed nerve allograft; qPCR.

MeSH terms

  • Adipose Tissue / cytology*
  • Allografts / cytology*
  • Allografts / innervation
  • Brain-Derived Neurotrophic Factor / genetics*
  • Cell Differentiation
  • Enzyme-Linked Immunosorbent Assay
  • Gene Expression*
  • Glial Cell Line-Derived Neurotrophic Factor / genetics
  • Humans
  • Mesenchymal Stem Cell Transplantation / methods*
  • Myelin Basic Protein / genetics
  • Nerve Growth Factor / genetics*
  • Nerve Regeneration / physiology*
  • Platelet Endothelial Cell Adhesion Molecule-1 / genetics
  • RNA, Messenger / genetics
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transplantation, Homologous
  • Vascular Endothelial Growth Factor A / genetics*

Substances

  • Brain-Derived Neurotrophic Factor
  • GDNF protein, human
  • Glial Cell Line-Derived Neurotrophic Factor
  • MBP protein, human
  • Myelin Basic Protein
  • NGF protein, human
  • Platelet Endothelial Cell Adhesion Molecule-1
  • RNA, Messenger
  • VEGFA protein, human
  • Vascular Endothelial Growth Factor A
  • BDNF protein, human
  • Nerve Growth Factor