Vascular tissue engineering - Biomaterials and Devices for the Circulatory System - page 335

As scientists have delved into the molecular mechanisms involved in
iPSC derivation, they have uncovered an alarming trend which would
hinder their clinical application. Chimeras and progeny mice derived from
iPSCs developed tumours which may be caused by reactivation of the
c-Myc oncogene, particularly when transferred using potentially harmful
retroviral vectors (Okita et al. , 2007). The fi rst iPSC clones that were gener-
ated had approximately 20 retroviral integrations, of which some were
thought to have fused with endogenous genes (Takahashi and Yamanaka,
2006). In order to avoid the development of tumours, iPSCs can be derived
without the retroviral-vector-mediated insertion of c-Myc (Nakagawa et al. ,
2008; Wernig et al. , 2008). Another approach to circumvent the develop-
ment of tumours is by avoiding retroviral vectors and generating iPSCs
using an adenovirus-mediated gene delivery system (Okita et al. , 2008;
Stadtfeld et al. , 2008). However, avoiding the use of retroviruses markedly
reduces the effi ciency of iPSC derivation. An adenovirus-mediated system
demonstrates that insertional mutagenesis is not required for in vitro repro-
gramming (Stadtfeld et al. , 2008).
Murine iPSCs have been shown, in vitro , to differentiate into SMCs
within 10 days through incubation in a two-dimensional environment with
retinoic acid. The derived SMCs, which expressed markers of contractility,
were capable of contracting when stimulated to do so (Xie et al. , 2009).
Human ECs and mural cells have also been derived from iPSCs (Narazaki
et al. , 2008). The mechanism the iPSCs use to differentiate into vascular
cells is thought to be similar to the mechanism employed by human embry-
onic stem cells (Narazaki et al. , 2008; Taura et al. , 2009). In particular, the
time course, potential and effi ciency of embryonic stem cell differentiation
were similar to iPSCs (Narazaki et al. , 2008). Flk1 is the accepted earliest
differentiation marker for ECs and blood cells. Flk1 is also a marker of the
lateral plate mesoderm from which, among others, red blood cells, bone
marrow, connective tissue and cardiac muscle develop (Kataoka et al. , 1997;
Yamaguchi et al. , 1993). Hence Flk1 positive progenitor cells are considered
to be the pluripotent common denominator from which cardiomyocytes,
ECs and VSMCs are generated (Kattman et al. , 2006). A subset of the Flk1
positive cells, which possess the islet 1 (isl1) and Nkx2.5 transcription
factors, has been identifi ed. This subset primarily generates cells inducing
cardiogenesis. Although cardiomyocytes are the predominant cells gener-
ated from these isl1 and Nkx2.5 positive cells, ECs and VSMCs have also
been derived from this subset of the Flk1 positive cells (Moretti et al. , 2006).
In a mouse model, iPSCs, which were positive for Flk1, were identifi ed
by differentiation at day 3.5. After a further three days, these differentiated
cells generated EC-type and SMC-type cells. When aggregates of these
Flk1 positive cells were cultured in a 3-dimensional environment, a vascu-
lar-like structure developed (Narazaki et al. , 2008). In the presence of

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