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Differential cyclin E expression in human in-stent stenosis smooth muscle cells identifies targets for selective
Michael O'Sullivan1, Stephen D Scott, Nicola McCarthy
1Division of Cardiovascular Medicine, Cambridge University, Box 110, Addenbrooke's Hospital, CB2 2QQ, Cambridge, UK.
Objective:
Cell cycle inhibitors are promising agents to prevent or treat human coronary in-stent stenosis (ISS). However, their lack of specificity for ISS vascular smooth muscle cells (VSMCs) may inhibit medial VSMC proliferation and suppress vessel healing.
Methods:
To identify inhibitor targets that differentially regulate proliferation of ISS vs. medial VSMCs, we examined cell cycle regulation in human VSMCs derived from (A) normal media, (B) ISS sites and (C) primary atherosclerotic plaques (P-VSMCs) using time-lapse videomicroscopy, flow cytometry, immunoblotting and immunohistochemistry.
Results:
ISS-VSMC proliferation was intermediate between P-VSMCs and medial VSMCs. Compared with medial cells, P-VSMCs expressed increased p16 and p21, reduced p27, reduced cyclins D(1) and E, and reduced pRb phosphorylation. In contrast, ISS-VSMCs expressed high levels of cyclins E and A with pRb hyperphosphorylation, both in vitro and in vivo, associated with increased and chronic cell proliferation in vivo. Roscovitine, a selective CDK2 inhibitor, inhibited VSMC proliferation by both pRb-dependent and independent pathways and more potently in ISS-VSMCs than medial VSMCs.
Conclusions:
Human ISS-VSMCs have marked differences in the stable expression of multiple cell cycle regulators, suggesting that ISS-VSMCs derive from P-VSMCs driven to proliferate through cyclin E overexpression. The critical role for cyclin E-CDK2 enables the identification of the first agent that selectively inhibits ISS-VSMC proliferation.
Insights
Targeting cell cycle regulators in in-stent stenosis (ISS) vascular smooth muscle cells (VSMCs) is key. Cyclin E overexpression drives ISS-VSMC proliferation, and a CDK2 inhibitor selectively reduced this proliferation.
Area of Science:
- Cardiovascular Biology
- Cell Cycle Regulation
- Vascular Smooth Muscle Cell Biology
Background:
- Cell cycle inhibitors show promise for treating in-stent stenosis (ISS).
- Non-specific inhibitors risk impairing medial vascular smooth muscle cell (VSMC) proliferation and vessel healing.
Purpose of the Study:
- Identify cell cycle targets that differentially regulate proliferation in ISS-VSMCs versus medial VSMCs.
- Investigate cell cycle dysregulation in human VSMCs from normal media, ISS sites, and atherosclerotic plaques.
Main Methods:
- Utilized time-lapse videomicroscopy, flow cytometry, immunoblotting, and immunohistochemistry.
- Examined cell cycle regulatory proteins (e.g., p16, p21, p27, cyclins, pRb) in different human VSMC populations.
- Assessed the effect of roscovitine, a CDK2 inhibitor, on VSMC proliferation.
Main Results:
- ISS-VSMC proliferation was intermediate between atherosclerotic plaque VSMCs and medial VSMCs.
- ISS-VSMCs exhibited high cyclin E and A levels with pRb hyperphosphorylation, driving chronic proliferation.
- Roscovitine potently inhibited ISS-VSMC proliferation via pRb-dependent and independent pathways.
Conclusions:
- Human ISS-VSMCs display distinct cell cycle regulator expression profiles, linked to cyclin E overexpression.
- Cyclin E-CDK2 activity is critical for ISS-VSMC proliferation.
- Identified a selective inhibitor targeting ISS-VSMC proliferation, offering therapeutic potential.
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