Transfection of gene regulation nanoparticles complexed with pDNA and shRNA controls multilineage differentiation of hMSCs

Biomaterials. 2018 Sep;177:1-13. doi: 10.1016/j.biomaterials.2018.05.035. Epub 2018 May 29.

Abstract

Overexpression and knockdown of specific proteins can control stem cell differentiation for therapeutic purposes. In this study, we fabricated RUNX2, SOX9, and C/EBPα plasmid DNAs (pDNAs) and ATF4-targeting shRNA (shATF4) to induce osteogenesis, chondrogenesis, and adipogenesis of human mesenchymal stem cells (hMSCs). The pDNAs and shATF4 were complexed with TRITC-gene regulation nanoparticles (GRN). Osteogenesis-related gene expression was reduced at early (12 h) and late (36 h) time points after co-delivery of shATF4 and SOX9 or C/EBPα pDNA, respectively, and osteogenesis was inhibited in these hMSCs. By contrast, osteogenesis-related genes were highly expressed upon co-delivery of RUNX2 and ATF4 pDNAs. DEX in GRN enhanced chondrogenic differentiation. Expression of osteogenesis-, chondrogenesis-, and adipogenesis-related genes was higher in hMSCs transfected with NPs complexed with RUNX2 and ATF4 pDNAs, shATF4 and SOX9 pDNA, and shATF4 and C/EBPα pDNA for 72 h than in control hMSCs, respectively. Moreover, delivery of these NPs also increased expression of osteogenesis-, chondrogenesis-, and adipogenesis-related proteins. These alterations in expression led to morphological changes, indicating that hMSCs differentiated into osteoblasts, chondrocytes, and adipose cells.

Keywords: Gene regulation nanoparticles; MSC; Multi-lineage; pDNA; shRNA.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Activating Transcription Factor 4 / genetics
  • Adipogenesis
  • CCAAT-Enhancer-Binding Protein-alpha / genetics
  • Cell Differentiation*
  • Cell Line
  • Chondrogenesis
  • Core Binding Factor Alpha 1 Subunit / genetics
  • DNA / genetics
  • Humans
  • Mesenchymal Stem Cells / cytology*
  • Mesenchymal Stem Cells / metabolism
  • Nanoparticles / chemistry
  • Osteogenesis
  • Plasmids / genetics
  • RNA Interference*
  • RNA, Small Interfering / genetics
  • SOX9 Transcription Factor / genetics
  • Transfection / methods*

Substances

  • ATF4 protein, human
  • CCAAT-Enhancer-Binding Protein-alpha
  • Core Binding Factor Alpha 1 Subunit
  • RNA, Small Interfering
  • RUNX2 protein, human
  • SOX9 Transcription Factor
  • SOX9 protein, human
  • Activating Transcription Factor 4
  • DNA