DNA damage and repair in a model of rat vascular injury.
Restenosis rate following vascular interventions still limits their long-term success. Oxidative stress plays a relevant role in this pathophysiological phenomenon, but less attention has been devoted to its effects on DNA damage and to the subsequent mechanisms of repair. We analysed in a model of arteriotomy-induced stenosis in rat carotids the time-dependent expression of DNA damage markers and of DNA repair genes, together with the assessment of proliferation and apoptosis indexes. The expression of the oxidative DNA damage marker 7,8-dihydro-8-oxo-2'-deoxyguanosine was increased at 3 and 7 days after arteriotomy, with immunostaining distributed in the injured vascular wall and in perivascular tissue. The expression of the DNA damage marker phospho-H2A.X was less relevant but increasing from 4 hrs to 7 days after arteriotomy, with immunostaining prevalently present in the adventitia and, to a lesser extent, in medial smooth muscle cells at the injury site. RT-PCR indicated a decrease of 8 out of 12 genes of the DNA repair machinery we selected from 4 hrs to 7 days after arteriotomy with the exception of increased Muyth and Slk genes (p<0.05). Western Blot revealed a decrease of p53 and catalase at 3 days after arteriotomy (p<0.05). A maximal 7% of BrdU-positive cells in endothelium and media occurred at 7 days after arteriotomy, while the apoptotic index peaked at 3 days after injury (p<0.05). Our results highlight a persistent DNA damage presumably related to a temporary decreased expression of the DNA repair machinery and of the antioxidant enzyme catalase, playing a role in stenosis progression.
Department of Experimental Medicine, Naples, Italy.
Amalia
A
Forte
Mauto
M
Finicelli
Mario
M
Grossi
Mariano
M
Vicchio
Nicola
N
Alessio
Pasquale
P
Santé
Marisa
M
De Feo
Maurizio
M
Cotrufo
Liberato
L
Berrino
Francesco
F
Rossi
Umberto
U
Galderisi
Marilena
M
Cipollaro
D000080242:8-Hydroxy-2'-Deoxyguanosine; D000818:Animals; D017209:Apoptosis; D024682:BRCA2 Protein / Q000235:genetics; D015153:Blotting, Western; D002339:Carotid Arteries / Q000378:metabolism* / Q000473:pathology / Q000601:surgery; D002374:Catalase / Q000378:metabolism; D049109:Cell Proliferation; D004249:DNA Damage; D045647:DNA Glycosylases / Q000235:genetics; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000378:metabolism; D004195:Disease Models, Animal; D015870:Gene Expression; D006657:Histones / Q000037:antagonists & inhibitors / Q000378:metabolism; D007150:Immunohistochemistry; D008297:Male; D010750:Phosphoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D057772:Vascular System Injuries / Q000235:genetics* / Q000378:metabolism
D024682:BRCA2 Protein; D004268:DNA-Binding Proteins; D006657:Histones; D010750:Phosphoproteins; D016159:Tumor Suppressor Protein p53; C539916:XRCC4 protein, rat; C577575:gamma-H2AX protein, rat; D000080242:8-Hydroxy-2'-Deoxyguanosine; D002374:Catalase; D045647:DNA Glycosylases; C483741:OGG1 protein, rat; D003849:Deoxyguanosine