The effect of substrate topography on direct reprogramming of fibroblasts to induced neurons.
Cellular reprogramming holds tremendous potential for cell therapy and regenerative medicine. Recently, fibroblasts have been directly converted into induced neurons (iNs) by overexpression of the neuronal transcription factors Ascl1, Brn2 and Myt1L. Hypothesizing that cell-topography interactions could influence the fibroblast-to-neuron reprogramming process, we investigated the effects of various topographies on iNs produced by direct reprogramming. Final iN purity and conversion efficiency were increased on micrograting substrates. Neurite branching was increased on microposts and decreased on microgratings, with a simplified dendritic arbor characterized by the reduction of MAP2(+) neurites. Neurite outgrowth increased significantly on various topographies. DNA microarray analysis detected 20 differentially expressed genes in iNs reprogrammed on smooth versus microgratings, and quantitative PCR (qPCR) confirmed the upregulation of Vip and downregulation of Thy1 and Bmp5 on microgratings. Electrophysiology and calcium imaging verified the functionality of these iNs. This study demonstrates the potential of applying topographical cues to optimize cellular reprogramming.
Department of Biomedical Engineering, Durham, NC 27708, USA.
Karina
K
Kulangara
Department of Biomedical Engineering, Durham, NC 27708, USA.
Andrew F
AF
Adler
Department of Neurobiology, Durham, NC 27708, USA.
Hong
H
Wang
Department of Biomedical Engineering, Durham, NC 27708, USA.
Malathi
M
Chellappan
Department of Biomedical Engineering, Durham, NC 27708, USA.
Ellen
E
Hammett
Department of Neurobiology, Durham, NC 27708, USA; Howard Hughes Medical Institute, Duke University, Durham, NC 27708, USA.
Ryohei
R
Yasuda
Department of Biomedical Engineering, Durham, NC 27708, USA. Electronic address: kam.leong@duke.edu.
Kam W
KW
Leong
D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D065150:Cellular Reprogramming; D005347:Fibroblasts / Q000166:cytology*; D015870:Gene Expression; D007150:Immunohistochemistry; D051379:Mice; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D016501:Neurites; D055495:Neurogenesis / Q000187:drug effects; D009474:Neurons / Q000166:cytology*; D044968:Regenerative Medicine; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D001672:Biocompatible Materials; D009419:Nerve Tissue Proteins; D014157:Transcription Factors
Biomaterials; Direct reprogramming; Induced Neurons; Topography