Application of real-time RT-PCR to quantifying gene expression of matrix metalloproteinases and tissue inhibitors of metalloproteinases in human abdominal aortic aneurysm

Backgound: The relative expression levels of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), key regulators in remodeling of extracellular matrix, are considered to play a pivotal role in the development of abdominal aortic aneurysm (AAA). However, few data exist regarding quantitative assessment of their expression in clinical settings. Methods: In 22 patients with AAA who underwent graft replacement, tissue samples of the AAA and non-dilated aorta were obtained. Using a real-time RT-PCR method that enabled quantitative measurement of mRNA levels in small tissue samples, we determined gene expression levels of MMPs and TIMPs relative to that of glutaraldehyde 3-phosphate dehydrogenase in each sample. Results: The expression levels of the MMP-1 and -3 genes were significantly augmented in AAA compared with non-dilated regions (4.48 ± 2.01 versus 0.26 ± 0.12, P<0.01 and 1.89 ± 1.00 versus 5.01 ± 0.97, P< 0.05, respectively). Although genes for TIMP-1, -2 and -3 tended to be upregulated in AAA, relative expression levels of MMP-1 to TIMP-1, MMP-1 to TIMP-2, MMP-1 to TIMP-3, and MMP-3 to TIMP-2 were still higher in AAA than in non-dilated regions (1.12 ± 0.63 versus 0.10 ± 0.03, 4.13 ± 1.12 versus 0.43 ± 0.11, 1.61 ± 0.59 versus 0.14 ± 0.03, and 7.81 ± 1.60 versus 2.56 ± 0.76, respectively, P< 0.05). Conclusion: These results demonstrate that the present real-time RT-PCR method is reliable for the determination of mRNA levels in small samples of vascular tissue and that disproportional expression of both MMP-1 and MMP-3 relative to TIMPs relates pathologically to the evolution of AAA.