TY - JOUR
T1 - KLF4-dependent phenotypic modulation of smooth muscle cells has a key role in atherosclerotic plaque pathogenesis
AU - Shankman, Laura S.
AU - Gomez, Delphine
AU - Cherepanova, Olga A.
AU - Salmon, Morgan
AU - Alencar, Gabriel F.
AU - Haskins, Ryan M.
AU - Swiatlowska, Pamela
AU - Newman, Alexandra A.C.
AU - Greene, Elizabeth S.
AU - Straub, Adam C.
AU - Isakson, Brant
AU - Randolph, Gwendalyn J.
AU - Owens, Gary K.
N1 - Funding Information:
M. Solga from the Flow Cytometry Core at the University of Virginia for their help designing flow cytometry panels and help running the cytometers; S. Guilot at the Advanced Microscopy Facility at University of Virginia for her help running the transmission electron microscope; J. Roithmayr, N. Hendley, M. Quetsch and M. Goodwin for their assistance in immunofluorescence image analysis; Y. Babiy for designing the cartoon in Supplementary Figure 6, and W. Evans for assistance in processing statistical analyses. We would also like to thank N. McGinn from the Department of Pathology, University of Virginia Hospital for the de-identified coronary arteries specimens; C. Murray from the University of Washington for the coronary artery heart transplant specimen; S. Offermanns from the Max Planck Institute for the Myh11-CreERT2 mice; K. Kaestner for the Klf4fl/fl mice; G. Randolph for the LysMCre/Cre mice; and A. Berns from the Netherlands Cancer Institute, Amsterdam, Netherlands, for the transgenic tamoxifen-inducible Cre (ERT-Cre) recombinase mice. This work was supported by US National Institutes of Health R01 grants HL057353, HL098538 and HL087867 to G.K.O., HL112904 to A.C.S., as well as a pilot grant from AstraZeneca as part of a University of Virginia–AstraZeneca Research Alliance to G.K.O.; and Mid-Atlantic American Heart Association fellowship grants 11PRE7170008 and 13POST17080043 to L.S.S. and D.G., respectively.
Publisher Copyright:
© 2015 Nature America, Inc. All rights reserved.
PY - 2015/6/9
Y1 - 2015/6/9
N2 - Previous studies investigating the role of smooth muscle cells (SMCs) and macrophages in the pathogenesis of atherosclerosis have provided controversial results owing to the use of unreliable methods for clearly identifying each of these cell types. Here, using Myh11-CreERT2 ROSA floxed STOP eYFP Apoe-/- mice to perform SMC lineage tracing, we find that traditional methods for detecting SMCs based on immunostaining for SMC markers fail to detect >80% of SMC-derived cells within advanced atherosclerotic lesions. These unidentified SMC-derived cells exhibit phenotypes of other cell lineages, including macrophages and mesenchymal stem cells (MSCs). SMC-specific conditional knockout of Krüppel-like factor 4 (Klf4) resulted in reduced numbers of SMC-derived MSC- and macrophage-like cells, a marked reduction in lesion size, and increases in multiple indices of plaque stability, including an increase in fibrous cap thickness as compared to wild-type controls. On the basis of in vivo KLF4 chromatin immunoprecipitation-sequencing (ChIP-seq) analyses and studies of cholesterol-treated cultured SMCs, we identified >800 KLF4 target genes, including many that regulate pro-inflammatory responses of SMCs. Our findings indicate that the contribution of SMCs to atherosclerotic plaques has been greatly underestimated, and that KLF4-dependent transitions in SMC phenotype are critical in lesion pathogenesis.
AB - Previous studies investigating the role of smooth muscle cells (SMCs) and macrophages in the pathogenesis of atherosclerosis have provided controversial results owing to the use of unreliable methods for clearly identifying each of these cell types. Here, using Myh11-CreERT2 ROSA floxed STOP eYFP Apoe-/- mice to perform SMC lineage tracing, we find that traditional methods for detecting SMCs based on immunostaining for SMC markers fail to detect >80% of SMC-derived cells within advanced atherosclerotic lesions. These unidentified SMC-derived cells exhibit phenotypes of other cell lineages, including macrophages and mesenchymal stem cells (MSCs). SMC-specific conditional knockout of Krüppel-like factor 4 (Klf4) resulted in reduced numbers of SMC-derived MSC- and macrophage-like cells, a marked reduction in lesion size, and increases in multiple indices of plaque stability, including an increase in fibrous cap thickness as compared to wild-type controls. On the basis of in vivo KLF4 chromatin immunoprecipitation-sequencing (ChIP-seq) analyses and studies of cholesterol-treated cultured SMCs, we identified >800 KLF4 target genes, including many that regulate pro-inflammatory responses of SMCs. Our findings indicate that the contribution of SMCs to atherosclerotic plaques has been greatly underestimated, and that KLF4-dependent transitions in SMC phenotype are critical in lesion pathogenesis.
UR - http://www.scopus.com/inward/record.url?scp=84930758301&partnerID=8YFLogxK
U2 - 10.1038/nm.3866
DO - 10.1038/nm.3866
M3 - Article
C2 - 25985364
AN - SCOPUS:84930758301
SN - 1078-8956
VL - 21
SP - 628
EP - 637
JO - Nature Medicine
JF - Nature Medicine
IS - 6
ER -