Related Experiment Video
Updated: Mar 30, 2026
Murine Aortic Crush Injury: An Efficient In Vivo Model of Smooth Muscle Cell Proliferation and Endothelial Function
Published on: June 11, 2017
MARCKS Signaling Differentially Regulates Vascular Smooth Muscle and Endothelial Cell Proliferation through a KIS-,
Dan Yu1, George Makkar2, Tuo Dong3
1Department of Surgery, Baltimore Veterans Affairs Medical Center, Baltimore, Maryland, United States of America; Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, Baltimore, Maryland, United States of America.
Background:
Overexpression of the myristolated alanine-rich C kinase substrate (MARCKS) occurs in vascular proliferative diseases such as restenosis after bypass surgery. MARCKS knockdown results in arrest of vascular smooth muscle cell (VSMC) proliferation with little effect on endothelial cell (EC) proliferation. We sought to identify the mechanism of differential regulation by MARCKS of VSMC and EC proliferation in vitro and in vivo.
Methods And Results:
siRNA-mediated MARCKS knockdown in VSMCs inhibited proliferation and prevented progression from phase G0/G1 to S. Protein expression of the cyclin-dependent kinase inhibitor p27kip1, but not p21cip1 was increased by MARCKS knockdown. MARCKS knockdown did not affect proliferation in VSMCs derived from p27kip1-/- mice indicating that the effect of MARCKS is p27kip1-dependent. MARCKS knockdown resulted in decreased phosphorylation of p27kip1 at threonine 187 and serine 10 as well as, kinase interacting with stathmin (KIS), cyclin D1, and Skp2 expression. Phosphorylation of p27kip1 at serine 10 by KIS is required for nuclear export and degradation of p27kip1. MARCKS knockdown caused nuclear trapping of p27kip1. Both p27kip1 nuclear trapping and cell cycle arrest were released by overexpression of KIS, but not catalytically inactive KIS. In ECs, MARCKS knockdown paradoxically increased KIS expression and cell proliferation. MARCKS knockdown in a murine aortic injury model resulted in decreased VSMC proliferation determined by bromodeoxyuridine (BrdU) integration assay, and inhibition of vascular wall thickening. MARCKS knockdown increased the rate of re-endothelialization.
Conclusions:
MARCKS knockdown arrested VSMC cell cycle by decreasing KIS expression. Decreased KIS expression resulted in nuclear trapping of p27kip1 in VSMCs. MARCKS knockdown paradoxically increased KIS expression in ECs resulting in increased EC proliferation. MARCKS knockdown significantly attenuated the VSMC proliferative response to vascular injury, but accelerated reestablishment of an intact endothelium. MARCKS is a novel translational target with beneficial cell type-specific effects on both ECs and VSMCs.
Insights
Myristolated alanine-rich C kinase substrate (MARCKS) knockdown halts vascular smooth muscle cell proliferation by trapping p27kip1, but paradoxically boosts endothelial cell growth. This suggests MARCKS is a therapeutic target for vascular diseases.
Area of Science:
- Vascular biology
- Cell cycle regulation
- Molecular mechanisms of disease
Background:
- Overexpression of myristolated alanine-rich C kinase substrate (MARCKS) is implicated in vascular proliferative diseases like restenosis.
- MARCKS knockdown inhibits vascular smooth muscle cell (VSMC) proliferation but has minimal impact on endothelial cell (EC) proliferation.
- The precise mechanism behind MARCKS's differential regulation of VSMC and EC proliferation requires elucidation.
Purpose of the Study:
- To investigate the mechanism by which MARCKS differentially regulates VSMC and EC proliferation.
- To identify the molecular players involved in MARCKS-mediated cell cycle control in VSMCs and ECs.
- To evaluate the therapeutic potential of targeting MARCKS in vascular injury models.
Main Methods:
- Utilized siRNA-mediated MARCKS knockdown in VSMCs and ECs in vitro.
- Assessed cell proliferation, cell cycle progression (G0/G1 to S phase), and protein expression levels (p27kip1, KIS, cyclin D1, Skp2).
- Employed a murine aortic injury model to study in vivo effects of MARCKS knockdown on vascular repair and proliferation.
Main Results:
- MARCKS knockdown in VSMCs inhibited proliferation by increasing p27kip1 levels and causing its nuclear trapping, dependent on p27kip1 expression.
- This arrest was linked to decreased kinase interacting with stathmin (KIS) expression and reduced p27kip1 phosphorylation.
- Conversely, MARCKS knockdown in ECs paradoxically increased KIS expression and proliferation; in vivo studies showed reduced VSMC proliferation and enhanced re-endothelialization post-injury.
Conclusions:
- MARCKS knockdown arrests VSMC cell cycle via KIS downregulation and p27kip1 nuclear trapping.
- MARCKS knockdown paradoxically promotes EC proliferation by increasing KIS expression.
- Targeting MARCKS offers a promising therapeutic strategy with cell-type-specific benefits for vascular proliferative diseases and injury repair.
More Related Videos
09:10Human Saphenous Vein Endothelial Cell Isolation and Exposure to Controlled Levels of Shear Stress and Stretch
Published on: April 21, 2023
11:38Visualization and Quantification of TGFβ/BMP/SMAD Signaling under Different Fluid Shear Stress Conditions using Proximity-Ligation-Assay
Published on: September 14, 2021
Related Concept Videos
01:24Regulation of Angiogenesis and Blood Supply
01:19Interactions Between Signaling Pathways
Convergence and divergence, and cross-talk between signaling pathways
Two distinct signaling pathways can converge on a single functional unit, which may either be a single protein or a complex of proteins. The response is either functionally distinct or synergistic between the two pathways but different from the response...
01:07MAPK Signaling Cascades
01:21Paracrine Signaling
01:12Role Of Notch Signalling In Intestinal Stem Cell Renewal
Direct cell-to-cell contact is needed for the activation of Notch signaling. The signal is initiated when a notch ligand binds to a receptor on an adjacent cell, also...
01:41Non-Canonical Wnt Signaling Pathways