J Korean Med Sci.  2014 Mar;29(3):363-369. 10.3346/jkms.2014.29.3.363.

Intimal Hyperplasia in Loop-Injured Carotid Arteries Is Attenuated in Transglutaminase 2-Null Mice

• Min SK
• Min SI
• Jeong EM
• Cho SY
• Ha J
• Kim SJ
• Kim IG

• KMID: 1734923
• DOI: http://doi.org/10.3346/jkms.2014.29.3.363

Abstract

Arterial restenosis frequently develops after open or endovascular surgery due to intimal hyperplasia. Since tissue transglutaminase (TG2) is known to involve in fibrosis, wound healing, and extracellular matrix remodeling, we examined the role of TG2 in the process of intimal hyperplasia using TG2-null mice. The neointimal formation was compared between TG2-null and wild-type (C57BL/6) mice by two different injury models; carotid ligation and carotid loop injury. In ligation model, there was no difference in intimal thickness between two groups. In loop injury model, intimal hyperplasia developed in both groups and the intimal/medial area ratio was significantly reduced in TG2-null mice (P = 0.007). TG2 was intensely stained in neointimal cells in 2 weeks. In situ activity of TG2 in the injured arteries steadily increased until 4 weeks compared to uninjured arteries. Taken together, intimal hyperplasia was significantly reduced in TG2-null mice, indicating that TG2 has an important role in the development of intimal hyperplasia. This suggests that TG2 may be a novel target to prevent the arterial restenosis after vascular surgery.

Keyword

Tissue Transglutaminase; Intimal Hyperplasia; Carotid Artery Injury

MeSH Terms

Animals
Carotid Arteries/pathology/*surgery
Disease Models, Animal
GTP-Binding Proteins/deficiency/genetics/*metabolism
Hyperplasia
Mice
Mice, Inbred C57BL
Transglutaminases/deficiency/genetics/*metabolism
Tunica Intima/*pathology
Transglutaminases
GTP-Binding Proteins

Figure

• 

  Fig. 1 Representative photographs of carotid arteries from mice after ligation. (A, B) Left carotid arteries of wild-type (A) and TG2-null mice (B) were ligated for 2 or 4 weeks and harvested (n = 8 for each group). Sections were stained with H&E (×400 magnification).

• 

  Fig. 2 Intimal hyperplasia is significantly reduced in TG2-null mice after loop injury. (A, B) Representative photographs of loop-injured carotid arteries from wild-type (A) and TG2-null mice (B). The carotid artery was opened, and intimal injury was induced by twisting and traction of prolene loop. After 2 or 4 weeks, arteries were harvested (n = 20 for each group). Sections were stained with H&E (×400 magnification). (C, D) Comparison of intimal (C) and medial (D) area of carotid arteries. The area of arterial structure was measured by morphometric analysis using an Image J program 1.42q. Neointimal area is indicated by the arrows in A and B. (E) A ratio of intimal/medial area for each carotid artery. Data represent mean±S.D.

• 

  Fig. 3 The expression of TG2 in intimal and medial cells is increased by loop-injury. (A, B) Immunohistochemical staining for TG2 in carotid artery from wild-type mice after 2 (A) or 4 weeks (B) of loop injury. The intimal cells are densely stained after 2 weeks and intimal and medial cells are diffusely stained after 4 weeks. Adventitia was not stained at both time points.

• 

  Fig. 4 Intracellular TG2 is activated by loop-injury. (A, B) Intimal injury of carotid arteries (A) and aorta (B) were induced by traction and twisting of a loop. After 2, 4, or 5 hr, specimens were harvested and incubated with biotinylated pentylamine (BP). In situ TG activity was assessed by visualizing the BP-incorporated proteins using Western blot analysis with streptavidin-HRP (SA). Graph shows the densitometry analysis of TG2 activity of representative Western blot. (C) In situ TG2 activity of carotid arteries from wild-type and TG2-null mice was determined at day 4, 7, 14, and 28 after intimal injury using well-plate assay method (n = 20 for each group). In situ TG2 activity is expressed as a relative value to that of the uninjured artery. Data were analyzed by one-way ANOVA with test for linear trend using GraphPad Prism.

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