Treatment of advanced atherosclerotic mice with ABT-263 reduced indices of plaque stability and increased mortality
Santosh Karnewar, Vaishnavi Karnewar, Laura S. Shankman, Gary K. Owens
Santosh Karnewar, Vaishnavi Karnewar, Laura S. Shankman, Gary K. Owens
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Research Article Vascular biology

Treatment of advanced atherosclerotic mice with ABT-263 reduced indices of plaque stability and increased mortality

Abstract

The use of senolytic agents to remove senescent cells from atherosclerotic lesions is controversial. A common limitation of previous studies is the failure to rigorously define the effects of senolytic agent ABT-263 (Navitoclax) on smooth muscle cells (SMC) despite studies claiming that these cells are the major source of senescent cells. Moreover, there are no studies on the effect of ABT-263 on endothelial cells (EC), which — along with SMC — comprise 90% of α-smooth muscle actin+ (α-SMA+) myofibroblast-like cells in the protective fibrous cap. Here we tested the hypothesis that treatment of advanced atherosclerotic mice with ABT-263 will reduce lesion size and increase plaque stability. SMC (Myh11-CreERT2-eYFP) and EC (Cdh5-CreERT2-eYFP) lineage tracing Apoe–/– mice were fed a western diet (WD) for 18 weeks, followed by ABT-263 at 100 mg/kg/bw for 6 weeks or 50 mg/kg/bw for 9 weeks. ABT-263 treatment did not change lesion size or lumen area of the brachiocephalic artery (BCA). However, ABT-263 treatment reduced SMC by 90% and increased EC contributions to lesions via EC-to-mesenchymal transition (EndoMT) by 60%. ABT-263 treatment also reduced α-SMA+ fibrous cap thickness by 60% and was associated with a > 50% mortality rate. Taken together, ABT-263 treatment of WD-fed Apoe–/– mice with advanced lesions resulted in multiple detrimental changes, including reduced indices of stability and increased mortality.

Authors

Santosh Karnewar, Vaishnavi Karnewar, Laura S. Shankman, Gary K. Owens

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Figure 4

Treatment of Apoe–/– mice with advanced atherosclerotic lesions with the senolytic agent ABT-263 was associated with a marked reduction in the number of SMC within BCA lesions but an increase in EC-derived cells undergoing EndoMT.

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Treatment of Apoe–/– mice with advanced atherosclerotic lesions with the...
ABT-263 treatment (100 mg/kg/bw) of SMC (Myh11-CreERT2-eYFP) and EC (Cdh5-CreERT2-eYFP) lineage tracing Apoe–/– mice with advanced atherosclerotic lesions was associated with a marked reduction in the number of SMC within BCA lesions but an increase in EC-derived cells undergoing EndoMT. However, the latter did not result in increased investment of EC-derived α-SMA+ cells into the fibrous cap. (A) Experimental design for B–E. SMC lineage tracing Apoe–/– mice were fed a WD for 18 weeks followed by 100 mg/kg/bw ABT-263 treatment on WD for 6 weeks. (B) Representative confocal images of costaining for eYFP (for detecting SMC), α-SMA+, and DAPI in advanced BCA lesions from A. The confocal images show a maximum intensity projection ×20 zoom. Scale bar: 100 μm. (C) α-SMA+ cap area normalized to lesion area (α-SMA+ cap area/lesion area). (D) Quantification of the percentage of SMC-derived (Myh11-eYFP+/DAPI+) cells in the fibrous cap. (E) Quantification of the percentage of SMC-derived α-SMA+ (Myh11-eYFP+ α-SMA+/α-SMA+) cells in the fibrous cap. (F) The experimental design for G–J, EC-lineage tracing Apoe–/– mice were fed a WD for 18 weeks followed by 100 mg/kg/bw ABT-263 treatment on WD for 6 weeks. (G) Representative confocal images of costaining for eYFP (for detecting EC), α-SMA+ and DAPI in advanced BCA lesions from F. (H) α-SMA+ Cap area normalized to lesion area. (I) Quantification of the percentage of EC-derived Cdh5-eYFP+DAPI+ cells in the fibrous cap. (J) Quantification of the percentage of EC-derived α-SMA+ (Cdh5-eYFP+α-SMA+/α-SMA+) cells in the fibrous cap. The two-way ANOVA method was used 2 statistical analyses in C–E and H–J. Biologically independent animals are indicated as individual dots. Data are shown as mean ± SEM . The P values are indicated on the figures.