Differential regulation of the superoxide dismutase family in experimental aortic aneurysms and rat aortic explants.
OBJECTIVE
Oxidative stress has been implicated in abdominal aortic aneurysm pathogenesis. This study sought to characterize the relevance of superoxide dismutases (SOD), a family of reactive oxygen catalyzing metalloenzymes, including manganese SOD (MnSOD), copper-zinc SOD (CuZnSOD), and extracellular SOD (EcSOD), in a rodent aortic aneurysm model.


METHODS
Male rat infrarenal abdominal aortas were perfused with either saline (control) or porcine pancreatic elastase (6 U/mL). Aortic diameter was measured and aortas harvested on post-operation days 1, 2, and 7 (N=5-6 per treatment group per day). MnSOD, CuZnSOD, EcSOD, catalase, MMP-2, MMP-9, and beta-actin expression in aortic tissue was determined by quantitative real-time PCR. MnSOD protein levels were measured using western immunoblotting and immunohistochemistry. In subsequent experiments, aimed at understanding the mechanism by which SOD is involved in AAA pathogenesis, rat aortic explants (RAEs) were incubated in media for 24 h in the presence of interleukin-1beta (IL-1beta, 2 ng/mL) and TEMPOL (SOD mimetic), catalase, or a combined SOD and catalase mimetic. Media MMP-2 and MMP-9 activity was determined by zymography. Data were analyzed by Student's t-tests and ANOVA.


RESULTS
Elastase-perfused aortic diameters were significantly increased compared to control aortas by post-perfusion day 7 (P=0.016). MnSOD mRNA levels in elastase perfused aortas were 6.0 (P=0.05) and 7.5 times (P<0.01) greater than control aortas at post-perfusion days 1 and 2, respectively. EcSOD, CuZnSOD, catalase, and MMP-2 mRNA expression did not statistically vary between the two groups. MMP-9 expression was 3.5-fold higher in the elastase group on post-perfusion day 2 (P=0.04). Western immunoblotting confirmed MnSOD protein was up-regulated on day 4 in the elastase-perfused group compared to controls (P=0.02). Immunohistrochemistry demonstrated increased MnSOD staining in the elastase group on day 4. In RAE experiments, TEMPOL increased both MMP-9 and MMP-2 activity 2 (P=0.09) and 3-fold (P=0.05), respectively, whereas catalase and the combined SOD/catalase mimetic failed to increase MMP-2 or MMP-9 activity.


CONCLUSIONS
Experimental abdominal aortic aneurysm formation is associated with early increases in MnSOD expression and an increase in MMP-9 activity. Strategies aimed at inhibiting oxidative stress during AAA formation should focus on MnSOD.
Sinha
Indranil
I
Jobst Vascular Research Laboratories, Section of Vascular Surgery, Department of Surgery, University of Michigan, Ann Arbor, Michigan, USA.
Pearce
Charles G
CG
Cho
Brenda S
BS
Hannawa
Kevin K
KK
Roelofs
Karen J
KJ
Stanley
James C
JC
Henke
Peter K
PK
Upchurch
Gilbert R
GR
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001012:Aorta, Abdominal / Q000201:enzymology; D017544:Aortic Aneurysm, Abdominal / Q000378:metabolism* / Q000503:physiopathology*; D003497:Cyclic N-Oxides / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology; D006861:Hydrogen Peroxide / Q000378:metabolism; D053583:Interleukin-1beta / Q000494:pharmacology; D008297:Male; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D009924:Organ Culture Techniques; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology*; D010196:Pancreatic Elastase / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013113:Spin Labels; D013482:Superoxide Dismutase / Q000235:genetics* / Q000378:metabolism*; D000072105:Superoxide Dismutase-1; D015854:Up-Regulation / Q000502:physiology; D000099156:Superoxide Dismutase 2
D000975:Antioxidants; D003497:Cyclic N-Oxides; D006861:Hydrogen Peroxide; D053583:Interleukin-1beta; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D010196:Pancreatic Elastase; D013113:Spin Labels; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D000099156:Superoxide Dismutase 2; C000606292:Sod1 protein, rat; C499089:Sod3 protein, rat; C001803:tempol