Elastase-induced matrix degradation in arterial organ cultures: an in vitro model of aneurysmal disease.
OBJECTIVE
Abdominal aortic aneurysms are characterized by degradation of the extracellular matrix, induction of endogenous metalloproteinases (MMPs), and development of a chronic inflammatory infiltrate. Despite intensive analysis of end-stage tissue, aneurysm pathogenesis remains obscure. The aim of this study was to develop an in vitro model of aneurysmal disease.


METHODS
Porcine aortic organ cultures were preincubated with pancreatic elastase before culture in standard conditions for up to 14 days. The extent of matrix degradation at various time points was determined by quantitative histologic estimation of collagen and elastin concentration. Endogenous metalloproteinase production within the tissue was quantified by gel enzymography and immunoblotting. A separate series of experiments was performed to investigate the effect of incorporating autologous leukocytes into the culture system.


RESULTS
Although exogenous elastase was removed after 24 hours, substantial degradation of the aortic extracellular matrix occurred in the subsequent 13 days in tissue culture. Analysis of samples preincubated with elastase (100 U/ml) for 24 hours before tissue culture demonstrated that elastin degradation occurred in a time-dependent manner (p < 0.001) and was not confined to the initial phase of exogenous elastase activity. Gelatin gel enzymography revealed a time-related production of metalloproteinases (55 to 250 kDa) within the aortic tissue. The presence of MMPs-1, 2, 3, and 9 was determined by immunoblotting. Immunohistochemistry identified the vascular smooth-muscle cell as the source of MMPs-1, 2, and 3. Addition of autogenous leukocytes to elastase-pretreated tissue initiated an inflammatory infiltrate within the aortic wall, which further enhanced both matrix degradation and MMP production (p < 0.001).


CONCLUSIONS
These data demonstrate that aortic samples pretreated with elastase before tissue culture undergo matrix degradation with MMP production and the development of an inflammatory infiltrate. These changes mirror the pathophysiological events within established aneurysms. It is suggested that this model may be useful in understanding early pathogenic events within aneurysmal tissue.
Department of Surgery, Leicester University, United Kingdom.
A
A
Wills
M M
MM
Thompson
M
M
Crowther
N P
NP
Brindle
A
A
Nasim
R D
RD
Sayers
P R
PR
Bell
D000818:Animals; D001014:Aortic Aneurysm / Q000378:metabolism / Q000473:pathology*; D003094:Collagen / Q000378:metabolism; D004549:Elastin / Q000378:metabolism; D005109:Extracellular Matrix / Q000473:pathology*; D015151:Immunoblotting; D007962:Leukocytes / Q000502:physiology; D008666:Metalloendopeptidases / Q000096:biosynthesis; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D009924:Organ Culture Techniques; D010196:Pancreatic Elastase / Q000494:pharmacology*; D013552:Swine
D003094:Collagen; D004549:Elastin; D010196:Pancreatic Elastase; D008666:Metalloendopeptidases