Abstract 29: Fate-Tracing Demonstrates that Medial SMCs, but Not Mature ECs, Contribute to Neointimal Cell Population in Injured Arteries

It has been proposed that various groups of cells, such as medial SMCs (mSMCs), bone marrow progenitors, and mature ECs (mECs) hold the potential to become neointimal SMCs via phenotypic switching, differentiation, or endothelial-mesenchymal-transition (EMT). However, no definitive evidence has been provided to support these claims, which renders the in vivo significance of these events in question. Using an inducible Cre-loxP system, where the production of a reporter (e.g. β-gal) is driven by a highly definitive marker gene for mSMCs (e.g. SMC myosin heavy chain, SMMHC) or mECs (e.g. stem cell leukemia, SCL), we traced the fate of these cells during neointimal hyperplasia (NIH) in ligated common carotid and wire-injured femoral arteries. Tamoxifen induction resulted in a homogeneous labeling of mSMCs of the unmanipulated arteries of R26R;SMMHC-CreERtm mice. Following arterial injury, SMCs that were initially located in the medial layer presented in neointimal lesions; however, the fraction of these cells varied widely among vessels with either type of injury. This heterogeneity was not secondary to the random loss of the genetic label as evidenced by the stable β-gal activity in SMC culture. Addition of a second (4wks after surgery) induction did not change the pattern and fraction of the labeled SMCs in the neointima, indicating that few recruited progenitors transform into mature SMCs. This observation is further supported through examination of mice chimeric for R26R;SMMHC-CreERtm bone marrow, where β-gal activity was not detected in injured arteries though strongly induced in culture following PDGF treatment. Neointima formed in R26R;SCL-CreERtm arteries demonstrated a co-localization of β-gal activity to the repopulated ECs, with none being detected in cells underneath the EC monolayer, suggesting a role for mECs in re-endothelialization and indicating EMT as a rare event in NIH development. We have provided direct evidence for the contribution of mSMCs to NIH. The recruited vascular progenitors, particularly bone marrow cells, do not differentiate into fully mature SMCs in NIH lesion. EMT does not serve as a mechanism for populating neointimal cells following vascular injury while occurring under other vascular disease conditions.