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Increased synthesis of matrix metalloproteinases by aortic smooth muscle cells is implicated in the etiopathogenesis of abdominal aortic aneurysms  Manish.

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Presentation on theme: "Increased synthesis of matrix metalloproteinases by aortic smooth muscle cells is implicated in the etiopathogenesis of abdominal aortic aneurysms  Manish."— Presentation transcript:

1 Increased synthesis of matrix metalloproteinases by aortic smooth muscle cells is implicated in the etiopathogenesis of abdominal aortic aneurysms  Manish I. Patel, MBBS, James Melrose, PhD, Peter Ghosh, PhD, Michael Appleberg, FRACS  Journal of Vascular Surgery  Volume 24, Issue 1, Pages (July 1996) DOI: /S (96) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

2 Fig. 1 Demonstration of SMC monolayers from AAA specimen stained with SMC α-actin monoclonal antibody FITC conjugate. Magnification ×50. Control SMC slides were also examined by using mouse IgG of same subclass (IgG1) as a-actin monoclonal antibody but non reactive with SMCs to assess any autofluorescence by SMCs; because this was not significant, no data are shown. Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

3 Fig. 2 Assessment of relative elastolytic activity present in AAA SMC serum-free media as function of pH. Range of buffers were prepared to cover pH range 4 to 11 by using sodium acetate (open circles), tris-hydrochloride (closed circles), and CAPS (filled squares) buffers that were substituted into standard assay protocol by using SAAANA as substrate. Each point represents mean of triplicate samples, and error bars are standard deviations. Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

4 Fig. 3 Assessment of relative elastolytic activity (against SAAANA) present in serum-free SMC media samples from IMA, AOD, normal and AAA tissue groups. Mean values of relative elastolytic activity (PPE degradative units/ml of media) + SD are presented. One unit of PPE degradative activity was defined as that amount of enzyme causing increase in absorbance at 405 nm of in 48 hours by using SAAANA as substrate under our standard assay conditions. AAA SMCs degrade significantly more substrate than IMA (p < 0.005), AOD (p = 0.002), and normal SMCs (p = 0.03). Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

5 Fig. 4 Assessment of relative elastolytic activity (as dpm solubilized from 14C-insoluble elastin) present in serum-free SMC media samples from the IMA, AOD, normal, and AAA tissue groups. Mean values + SD are presented, and data were normalized on basis of equivalent cell numbers. AAA SMCs degrade more elastin than IMA SMC media (p < 0.005) and AOD (p = 0.01). No significant difference between normal and AAA SMC media (p = 0.198). Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

6 Fig. 5 Zymographic assessment of gelatinolytic activity present in serum-free AOD, IMA, normal, and AAA SMC media samples and of IL-1β stimulated human rheumatoid fibroblast media sample (Fib). The prestained molecular weight standards run at left of figure were bovine serum albumin (98 kDa), glutamic dehydrogenase (64 kDa), alcohol dehydrogenase (50 kDa), and carbonic anhydrase (36 kDa). Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

7 Fig. 6 Western blot analysis of same SMC media samples as electrophoresed in Fig. 5. Samples were subjected to 4% to 20% gradient SDS PAGE and electrophoretically transferred to nitrocellulose. After blocking, blots were probed with monoclonal antibodies to MMP-2 (A) and MMP-9 (B). The protein standards used are indicated at left of figure and were same as Fig. 5. Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

8 Fig. 6 Western blot analysis of same SMC media samples as electrophoresed in Fig. 5. Samples were subjected to 4% to 20% gradient SDS PAGE and electrophoretically transferred to nitrocellulose. After blocking, blots were probed with monoclonal antibodies to MMP-2 (A) and MMP-9 (B). The protein standards used are indicated at left of figure and were same as Fig. 5. Journal of Vascular Surgery  , 82-92DOI: ( /S (96) ) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

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