The tight junction protein ZO-2 mediates proliferation of vascular smooth muscle cells via regulation of Stat1.
OBJECTIVE
Recent evidence suggests that the zonula occludens protein 2 (ZO-2) might have additional cellular functions, beyond regulation of paracellular permeability of epithelial and endothelial cells. Deregulation of ZO-2 in response to ischaemia, hypertensive stress, and vascular injury implies its involvement in cardiovascular disorders, most likely via regulating the functional behaviour of vascular smooth muscle cells (VSMC). However, a role of ZO-2 in VSMC biology has yet to be established. Our study was designed to understand the specific functions of ZO-2 in human VSMC.


RESULTS
The expression of ZO-2 and Stat1 upon vascular injury was studied using ex vivo organ culture of coronary arteries combined with immunohistochemistry. ZO-2 silencing in human primary VSMC was achieved by means of lentiviral gene transfer. Cell proliferation was assessed by analysing DNA synthesis and by cell counting. Stat1 expression was examined using immunoblotting, immunocytochemistry, TaqMan, and fluorescence activated cell sorting (FACS) analysis. Functional relevance of Stat1 up-regulation was studied using a Stat1 promoter-luciferase reporter assay and intracellular microinjections of a Stat1 specific antibody. ZO-2 was highly expressed in the media and neointima of dilated but not of control arteries, whereas expression of the transcription factor Stat1 was inversely regulated upon injury. Analysis of VSMC with down-regulated ZO-2 revealed increased expression of Stat1 in these cells, whereas Stat1 phosphorylation was not affected. Stat1 up-regulation in VSMC with ZO-2 silencing resulted in a coordinate activation of Stat1-specific genes and consequently led to inhibition of cell proliferation. This effect was restored by microinjection of a Stat1 neutralising antibody.


CONCLUSIONS
Our data suggest that the tight junction protein ZO-2 is involved in regulation of VSMC growth control upon vascular injury that is mediated by the transcription factor Stat1. Our findings point to a novel function of ZO-2 in VSMC and implicate ZO-2 as a novel important molecular target in pathological states of vascular remodelling in cardiovascular diseases.
Kusch
Angelika
A
Medical Faculty of the Charité, Franz Volhard Klinik/Experimental and Clinical Research Center-ECRC, Max Delbrück Center, Berlin, Germany. angelika.kusch@charite.de
Tkachuk
Sergey
S
Tkachuk
Natalia
N
Patecki
Margret
M
Park
Joon-Keun
JK
Dietz
Rainer
R
Haller
Hermann
H
Dumler
Inna
I
D000818:Animals; D049109:Cell Proliferation; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology / Q000378:metabolism; D005786:Gene Expression Regulation; D020868:Gene Silencing; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000378:metabolism*; D050794:STAT1 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction; D013552:Swine; D062827:Zonula Occludens-2 Protein
D008565:Membrane Proteins; D050794:STAT1 Transcription Factor; C569046:TJP2 protein, human; D062827:Zonula Occludens-2 Protein