Objective-Recent studies of bone marrow (BM)-transplanted apoE knockout (apoE Ϫ/Ϫ ) mice have concluded that a substantial fraction of smooth muscle cells (SMCs) in atherosclerosis arise from circulating progenitor cells of hematopoietic origin. This pathway, however, remains controversial. In the present study, we reexamined the origin of plaque SMCs in apoE Ϫ/Ϫ mice by a series of BM transplantations and in a novel model of atherosclerosis induced in surgically transferred arterial segments. Methods and Results-We analyzed plaques in lethally irradiated apoE Ϫ/Ϫ mice reconstituted with sex-mismatched BM cells from eGFP ϩ apoE Ϫ/Ϫ mice, which ubiquitously express enhanced green fluorescent protein (eGFP), but did not find a single SMC of donor BM origin among Ϸ10 000 SMC profiles analyzed. We then transplanted arterial segments between eGFP ϩ apoE Ϫ/Ϫ and apoE Ϫ/Ϫ mice (isotransplantation except for the eGFP transgene) and induced atherosclerosis focally within the graft by a recently invented collar technique. No eGFP ϩ SMCs were found in plaques that developed in apoE Ϫ/Ϫ artery segments grafted into eGFP ϩ apoE Ϫ/Ϫ mice. Concordantly, 96% of SMCs were eGFP ϩ in plaques induced in eGFP ϩ apoE Ϫ/Ϫ artery segments grafted into apoE Ϫ/Ϫ mice. Conclusions-These experiments show that SMCs in atherosclerotic plaques See page 2579 and coverUntil recently, the only source of SMCs in atherosclerotic lesions was considered to be the local vessel wall. According to this hypothesis, local SMCs in the arterial media and intima modulate into a synthetic and migratory phenotype (aka, phenotypic modulation 3 ) and form the fibrous component of the plaque. This theory was essentially inferred from a line of suggestive observations in arterial injury models in the 70s and 80s. 4,5 More to the point, Cre/lox fate mapping in the apoE knockout (apoE Ϫ/Ϫ ) mouse model of atherosclerosis recently confirmed that preexisting SMCs, presumably from the local media, contribute to plaque SMCs during atherogenesis, but the existence of other sources could not be excluded by this technique. 6 In 2002, an alternative and major source of SMCs in atherosclerosis was reported, 7 and this new paradigm has great impact on current research in this area. 8,9 Based on observations in bone marrow (BM)-transplanted apoE Ϫ/Ϫ mice, it was concluded that a substantial fraction of plaque SMCs arise from circulating progenitor cells of hematopoietic origin. 7 This pathway holds promise for the development of novel therapeutic means of controlling the recruitment and accumulation of SMCs in plaques. However, it remains questionable whether the quality of the histological documentation in that study can support a definitive conclusion, especially because the seeming use of unfixed tissue for detection of enhanced green fluorescent protein (eGFP) can lead to diffusion of the tracer marker from sectioned cells. 10 Furthermore, reconstitution of apoE Ϫ/Ϫ mice with apoE To address these concerns, we repeated the BM transplantation experiment in apoE Ϫ/Ϫ mice w...