RNAseq based transcriptomics study of SMCs from carotid atherosclerotic plaque: BMP2 and IDs proteins are crucial regulators of plaque stability

Abstract

Carotid artery atherosclerosis is a risk factor to develop cerebrovascular disease. Atheroma plaque can become instable and provoke a cerebrovascular event or else remain stable as asymptomatic type. The exact mechanism involved in plaque destabilization is not known but includes among other events smooth muscle cell (SMC) differentiation. The goal of this study was to perform thorough analysis of gene expression differences in SMCs isolated from carotid symptomatic versus asymptomatic plaques. Comparative transcriptomics analysis of SMCs based on RNAseq technology identified 67 significant differentially expressed genes and 143 significant differentially expressed isoforms in symptomatic SMCs compared with asymptomatic. 37 of top-scoring genes were further validated by digital PCR. Enrichment and network analysis shows that the gene expression pattern of SMCs from stable asymptomatic plaques is suggestive for an osteogenic phenotype, while that of SMCs from unstable symptomatic plaque correlates with a senescence-like phenotype. Osteogenic-like phenotype SMCs may positively affect carotid atheroma plaque through participation in plaque stabilization via bone formation processes. On the other hand, plaques containing senescence-like phenotype SMCs may be more prone to rupture. Our results substantiate an important role of SMCs in carotid atheroma plaque disruption.

Characterization of Carotid Smooth Muscle Cells during Phenotypic Transition

“…In a recent study, VSMCs isolated from carotid atherosclerotic plaques were found to express a series of macrophage markers [ 14 ], concomitant with a macrophage-like phenotype [ 25 ]. Analysis of the expression levels of CD68 and LGALS3 showed that both macrophage markers were significantly augmented in PLQ compared to MIT-VSMCs ( Figure 3 A).…”

“…Cell density at harvest was 200,000 cells/5 mL in a T25 flask. All experiments were carried out with cells in passage zero in order to keep overall cell characteristics as similar as possible to those in the natural context, and to avoid extended cultivation periods which would influence the expression levels of analyzed genes [ 14 ].…”

“…In the context of atherosclerosis, VSMCs are believed to dedifferentiate, proliferate, and migrate, thus contributing to plaque formation [ 5 ]; however, VSMCs may also display beneficial roles by protecting against plaque rupture through promotion of plaque repair [ 8 , 9 , 10 , 11 ]. Therefore, VSMCs play an essential role during plaque development from moderate stage to advanced atheroma plaques [ 12 , 13 , 14 ]. This process of cellular physiology change is called “switching of VSMCs” and indicates a change in their phenotype from differentiated contractile-state to dedifferentiated synthetic-state, which is considered a pre-requisite for the first stages of plaque development [ 5 ].…”

Uncovering the Potential Differentially Expressed miRNAs and mRNAs in Ischemic Stroke Based on Integrated Analysis in the Gene Expression Omnibus Database

“…The upregulated expression of ITGAM is found in the middle cerebral occlusion brain of rat [40]. ID3 regulates many biological processes, including cell growth, apoptosis, senescence, and angiogenesis [41]. Doran et al [42] found that ID3 was a key atheroprotective transcription factor.…”

Endothelial prostaglandin D2 opposes angiotensin II contractions in mouse isolated perfused intracerebral microarterioles

“…Total RNA was extracted with an RNeasy Mini Kit (Qiagen Inc. Germantown, MD 20874) and analyzed by RNAseq. 32 …”