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Cathepsin G deficiency reduces periaortic calcium chloride injury-induced abdominal aortic aneurysms in mice  Jing Wang, MD, PhD, Galina K. Sukhova, PhD,

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Presentation on theme: "Cathepsin G deficiency reduces periaortic calcium chloride injury-induced abdominal aortic aneurysms in mice  Jing Wang, MD, PhD, Galina K. Sukhova, PhD,"— Presentation transcript:

1 Cathepsin G deficiency reduces periaortic calcium chloride injury-induced abdominal aortic aneurysms in mice  Jing Wang, MD, PhD, Galina K. Sukhova, PhD, Jian Liu, PhD, Keith Ozaki, MD, Adam Lesner, MD, Peter Libby, MD, Petri T. Kovanen, MD, Guo-Ping Shi, DSc  Journal of Vascular Surgery  Volume 62, Issue 6, Pages (December 2015) DOI: /j.jvs Copyright © 2015 Society for Vascular Surgery Terms and Conditions

2 Fig 1 Cathepsin G (CatG) expression in a human abdominal aortic aneurysm (AAA) lesion and AAA-free aorta. Immunofluorescent double staining with anti-α-actin and anti-human CatG antibodies showed negligible CatG expression in AAA-free aorta but localized CatG expression in smooth muscle cell (SMC; α-actin, from the media), macrophage (CD68, from the intima), and endothelial cell (EC; CD31, from the media) in human AAA lesions. Insets with higher magnifications are shown in the bottom row. Magnifications are indicated. Journal of Vascular Surgery  , DOI: ( /j.jvs ) Copyright © 2015 Society for Vascular Surgery Terms and Conditions

3 Fig 2 Inflammatory cytokine- and high glucose concentration-induced cathepsin G (CatG) expression and activity in angiotensin II (Ang-II) and angiotensin-converting enzyme (ACE) production from human aortic SMCs (huSMCs). A, Immunoblot-determined CatG expression in huSMCs treated with different inflammatory cytokines for 48 hours. b-FGF, Basic fibroblast growth factor; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; IFN-γ, interferon-γ; IL6, interleukin 6; TNF-α, tumor necrosis factor-α. B, Enzyme-linked immunosorbent assay (ELISA)–determined ACE and Ang-II levels in lysates and culture medium from huSMCs cultured in low-glucose or high-glucose DMEM with and without CatG inhibitor for 24 hours. C, Immunoblot-determined CatG expression in huSMCs from low-glucose and high-glucose medium with and without CatG inhibitor for 24 hours. D, CatG immunoblot in huSMCs transfected with CatG small interfering RNA (siRNA) or control siRNA and treated with low or high concentrations of glucose (glu). E, ELISA-determined Ang-II and ACE levels in culture medium and cell lysates from huSMCs from panel D. GAPDH or β-actin blots were used to ensure equal protein loading for immunoblots. Bar figures are mean ± standard error of the mean from three to six independent experiments. Journal of Vascular Surgery  , DOI: ( /j.jvs ) Copyright © 2015 Society for Vascular Surgery Terms and Conditions

4 Fig 3 Aortic diameter and lesion areas from periaortic CaCl2 injury-induced AAAs in Ldlr−/−Ctsg−/− and Ldlr−/−Ctsg+/+ mice. Data are mean ± standard error. Number of mice per experimental group is indicated in each bar. AAA, Abdominal aortic aneurysm. Journal of Vascular Surgery  , DOI: ( /j.jvs ) Copyright © 2015 Society for Vascular Surgery Terms and Conditions

5 Fig 4 Characterization of periaortic CaCl2 injury-induced abdominal aortic aneurysm (AAA) between Ldlr−/−Ctsg−/− and Ldlr−/−Ctsg+/+ mice. A, Plasma angiotensin II (Ang-II) and angiotensin-converting enzyme (ACE) levels. B, Systolic and diastolic blood pressures before and after AAA production. C, Lesion macrophages, CD4+ T cells, and major histocompatibility complex class II (MHC class-II). D, Lesion smooth muscle cells (SMCs), CD31+ microvessels, Ki67+ proliferating cells, and TUNEL-positive apoptotic cells. Data are mean ± standard error. Number of mice per experimental group is indicated in each bar. Journal of Vascular Surgery  , DOI: ( /j.jvs ) Copyright © 2015 Society for Vascular Surgery Terms and Conditions

6 Fig 5 Abdominal aortic aneurysm (AAA) lesion elastin fragmentation grades (A) and collagen content grades (B) in Ldlr−/−Ctsg−/− and Ldlr−/−Ctsg+/+ mice at 4 weeks after periaortic CaCl2 injury. Representative elastin fragmentation (with magnifications indicated) and collagen grading keys are illustrated to the right of each panel. Data are mean ± standard error. Number of mice per experimental group is indicated in each bar. Journal of Vascular Surgery  , DOI: ( /j.jvs ) Copyright © 2015 Society for Vascular Surgery Terms and Conditions

7 Fig 6 In vitro elastin and type I collagen degradation assays. A, DQ elastin degradation by different amounts of purified human neutrophil cathepsin G (CatG). B, DQ type I collagen degradation by different amounts of purified CatG. Data are mean ± standard error of four independent experiments. C, DQ elastin degradation by abdominal aortic aneurysm (AAA) lesion tissue extracts from Ldlr−/−Ctsg−/− mice and Ldlr−/−Ctsg+/+ mice at 4 weeks after periaortic CaCl2 injury. D, Gelatin gel zymogram assay tested matrix metalloproteinase (MMP) 2, pro-MMP-2, and MMP-9 (indicated) activities in AAA lesions from Ldlr−/−Ctsg−/− mice and Ldlr−/−Ctsg+/+ mice. Representative zymograph is shown to the right panel. Data are mean ± standard error of four experiments. Number of mice per experimental group is indicated in each bar. Journal of Vascular Surgery  , DOI: ( /j.jvs ) Copyright © 2015 Society for Vascular Surgery Terms and Conditions

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