Transcript Slide 1

Mesenchymal Stem Cells Adopt a Myofibroblastic Phenotype in Culture:
Implications for Cellular Cardiomyoplasty
Melanie A. Ngo, Ryan H. Cunnington, Sunil G. Rattan, Yun Li, Melanie M. Durston, Ian M.C. Dixon, Rakesh C. Arora, Darren H. Freed
Laboratory of Molecular Cardiology, Institute of Cardiovascular Sciences, St. Boniface Research Centre, University of Manitoba, Winnipeg, Manitoba, Canada
OBJECTIVE:
The ability of the heart to
regenerate is limited. Cell transplantation, in an
effort to repair wounded hearts has been
proposed and undertaken with variable success.
Various studies have shown an increase in heart
function with mesenchymal stem cell (MSC)
transplantation, but limited differentiation of
transplanted cells to mature, functional
cardiomyocytes.
One explanation for this
observation is that MSCs spontaneously
differentiate in culture, which limits their capacity
for further differentiation in vivo. We examined
MSC in vitro and compared their phenotypic and
physiological characteristics to cultured cardiac
fibroblasts. These observations were made on
human and rat cells.
RESULTS:
-SMA
SMemb
Collagen Gel Contraction After 24 hours
Control
METHODS:
Rat bone marrow cells were
isolated from the long bones using the Caplan
method ("Mesenchymal Stem Cells", Arnold I.
Caplan, Journal of Orthopaedic Research, pp. 641650, 1991). Rat cardiac fibroblasts were isolated
using collagen digestion. Human bone marrow
cells (from the sternum) and human cardiac
fibroblasts (atrial/ventricular tissue) were
obtained from patients undergoing open heart
surgery. All cells were cultured in standard DMEM
F12 with 10% fetal bovine serum conditions.
Phenotype
characterization
and
protein
expression
were
determined
by
immunofluorescent staining and western blotting.
Myofibroblasts (MF) exert sustained tonic
contraction on scar tissue, and this physiological
function was assessed with collagen gel
contraction assays on second passage cells (P2).
P1 Human BMSC Positive for Myofibroblast Markers
Collagen Gel Contraction Assessment
Vimentin
Rat and human bone marrow
cells initially expressed the stem cell marker ckit which decreased over serial passage. In
addition, these cells increasingly expressed the
myofibroblastic markers alpha smooth muscle
actin (α-SMA), the embryonic isoform of
smooth muscle myosin (SMemb), procollagen
1A2 and vimentin over serial passage. Collagen
gel contraction assays revealed that MSCs
contracted to a similar degree as cardiac MF
derived from either rats or humans. Human
cells treated with TGF (10 ng/mL), showed
increased contractile function compared to
non-treated cells (p < 0.05).
TGF treated
CONCLUSIONS:
Human BM
P1 Rat BMSC Positive for Myofibroblast Markers
Human MF
-SMA
Rat BM
SMemb
Rat MF
Vimentin
Both rat and human
MSC of bone marrow origin display phenotypic
and physiologic transformation to MF during
cell expansion in vitro. These cells
demonstrate similar contractile ability
between species as well with their respective
cardiac derived MF. These data suggest that
MSCs differentiate readily in vitro to a
myofibroblast phenotype that is unlikely to
support
further
differentiation
to
cardiomyocytes that will integrate into the
host myocardium to improve systolic function.
Additional studies are warranted to address
this spontaneous differentiation and improve
cardiomyocyte
differentiation
potential.
Furthermore, the contribution of MSCs to
cardiac fibrosis needs to be investigated.