Accelerated aneurysmal dilation associated with apoptosis and inflammation in a newly developed calcium phosphate rodent abdominal aortic aneurysm model

Dai Yamanouchi, Stephanie Morgan, Colin Stair, Stephen Seedial, Justin Lengfeld, K. Craig Kent, Bo Liu

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47 Scopus citations

Abstract

Objective: The calcium chloride (CaCl 2) model is a widely accepted rodent model for abdominal aortic aneurysms (AAAs). Calcium deposition, mainly consisting of calcium phosphate (CaPO 4) crystals, has been reported to exist in human and experimental aneurysms. CaPO 4 crystals have been used for in vitro DNA transfection by mixing CaCl 2 and phosphate-buffered saline (PBS). Here, we describe accelerated aneurysm formation resulting from a modification of the CaCl 2 model. Methods: A modified CaCl 2 model, the CaPO 4 model, was created by applying PBS onto the mouse infrarenal aorta after CaCl 2 treatment. Morphologic, histologic, and immunohistochemical analyses were performed on arteries treated with the CaPO 4 model and the conventional CaCl 2 model as the control. In vitro methods were performed using a mixture of CaCl 2 and PBS to create CaPO 4 crystals. CaPO 4- induced apoptosis of primary cultured mouse vascular smooth muscle cells (VSMCs) was measured by DNA fragmentation enzyme-linked immunosorbent assay. Results: The CaPO 4 model produces AAA, defined as an increase of <50% in the diameter of the aorta, faster than in the CaCl 2 model. The CaPO 4 model showed significantly larger aneurysmal dilation at 7, 28, and 42 days, as reflected by a maximum diameter (measured in mm) fold-change of 1.69 ± 0.07, 1.99 ± 0.14, and 2.13 ± 0.09 vs 1.22 ± 0.04, 1.48 ± 0.07, and 1.68 ± 0.06 in a CaCl 2 model, respectively (n = 6; P <.05). A semiquantitative grading analysis of elastin fiber integrity at 7 days revealed a significant increase in elastin degradation in the CaPO 4 model compared with the CaCl 2 model (2.7 ± 0.2 vs 1.5 ± 0.2; n = 6; P <.05). A significantly higher level of apoptosis occurred in the CaPO 4 model (apoptosis index at 1, 2, and 3 days postsurgery: 0.26 ± 0.14, 0.37 ± 0.14, and 0.33 ± 0.08 vs 0.012 ± 0.10, 0.15 ± 0.02, and 0.12 ± 0.05 in the conventional CaCl 2 model; n = 3; P <.05). An enhancement of macrophage infiltration and calcification was also observed at 3 and 7 days in the CaPO 4 model. CaPO 4 induced approximately 3.7 times more apoptosis in VSMCs than a mixture of CaCl 2 (n = 4; P <.0001) in vitro. Conclusions: The CaPO 4 model accelerates aneurysm formation with the enhancement of apoptosis, macrophage infiltration, and calcium deposition. This modified model, with its rapid and robust dilation, can be used as a new model for AAAs.

Original languageEnglish (US)
Pages (from-to)455-461
Number of pages7
JournalJournal of vascular surgery
Volume56
Issue number2
DOIs
StatePublished - Aug 2012

Bibliographical note

Funding Information:
This work was supported by the National Institutes of Health R01HL088447 (B.L. and K.K.), American Heart Association 10GRNT3020052 (B.L. and S.M.), and Howard Hughes Medical Institute MSN135276 (S.S.).

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