title: Smooth muscle cells differentiated from reprogrammed embryonic lung fibroblasts through DKK3 signaling are potent for tissue engineering of vascular grafts.
abstract: BACKGROUND
  Smooth muscle cells (SMCs) are a key component of tissue-engineered vessels. However, the sources by which they can be isolated are limited.
  
  
  OBJECTIVE
  We hypothesized that a large number of SMCs could be obtained by direct reprogramming of fibroblasts, that is, direct differentiation of specific cell lineages before the cells reaching the pluripotent state.
  
  
  RESULTS
  We designed a combined protocol of reprogramming and differentiation of human neonatal lung fibroblasts. Four reprogramming factors (OCT4, SOX2, KLF4, and cMYC) were overexpressed in fibroblasts under reprogramming conditions for 4 days with cells defined as partially-induced pluripotent stem (PiPS) cells. PiPS cells did not form tumors in vivo after subcutaneous transplantation in severe combined immunodeficiency mice and differentiated into SMCs when seeded on collagen IV and maintained in differentiation media. PiPS-SMCs expressed a panel of SMC markers at mRNA and protein levels. Furthermore, the gene dickkopf 3 was found to be involved in the mechanism of PiPS-SMC differentiation. It was revealed that dickkopf 3 transcriptionally regulated SM22 by potentiation of Wnt signaling and interaction with Kremen1. Finally, PiPS-SMCs repopulated decellularized vessel grafts and ultimately gave rise to functional tissue-engineered vessels when combined with previously established PiPS-endothelial cells, leading to increased survival of severe combined immunodeficiency mice after transplantation of the vessel as a vascular graft.
  
  
  CONCLUSIONS
  We developed a protocol to generate SMCs from PiPS cells through a dickkopf 3 signaling pathway, useful for generating tissue-engineered vessels. These findings provide a new insight into the mechanisms of SMC differentiation with vast therapeutic potential.
authors.affiliation: Cardiovascular Division, British Heart Foundation Centre, King's College London, UK.
authors.forename: Eirini
authors.initials: E
authors.lastname: Karamariti
authors.forename: Andriana
authors.initials: A
authors.lastname: Margariti
authors.forename: Bernhard
authors.initials: B
authors.lastname: Winkler
authors.forename: Xiaocong
authors.initials: X
authors.lastname: Wang
authors.forename: Xuechong
authors.initials: X
authors.lastname: Hong
authors.forename: Dilair
authors.initials: D
authors.lastname: Baban
authors.forename: Jiannis
authors.initials: J
authors.lastname: Ragoussis
authors.forename: Yi
authors.initials: Y
authors.lastname: Huang
authors.forename: Jing-Dong J
authors.initials: JD
authors.lastname: Han
authors.forename: Mei Mei
authors.initials: MM
authors.lastname: Wong
authors.forename: Can M
authors.initials: CM
authors.lastname: Sag
authors.forename: Ajay M
authors.initials: AM
authors.lastname: Shah
authors.forename: Yanhua
authors.initials: Y
authors.lastname: Hu
authors.forename: Qingbo
authors.initials: Q
authors.lastname: Xu
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D001807:Blood Vessel Prosthesis; D002454:Cell Differentiation / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism; D002469:Cell Separation / Q000379:methods; D018925:Chemokines; D005333:Fetus / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism*; D000090062:Kruppel-Like Factor 4; D008168:Lung / Q000166:cytology*; D008565:Membrane Proteins / Q000378:metabolism; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000166:cytology* / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology*; D015533:Transcriptional Activation / Q000502:physiology; D060449:Wnt Signaling Pathway / Q000502:physiology; D051176:beta Catenin / Q000378:metabolism
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C495270:CTNNB1 protein, human; D018925:Chemokines; C498624:DKK3 protein, human; D036341:Intercellular Signaling Peptides and Proteins; C000715467:KLF4 protein, human; C486075:KREMEN1 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D008565:Membrane Proteins; D008840:Microfilament Proteins; D009124:Muscle Proteins; D051176:beta Catenin; C051615:transgelin
keywords: animal models; smooth muscle cells; stem cells; vascular progenitors; vascular tissue engineering