Fibrillin-1 genetic deficiency leads to pathological ageing of arteries in mice

Abstract

Fibrillin-1, the major component of extracellular microfibrils that associate with insoluble elastin in elastic fibres, is mainly synthesized during development and postnatal growth and is believed to guide elastogenesis. Mutations in the fibrillin-1 gene cause Marfan syndrome, a multisystem disorder characterized by aortic aneurysms and dissections. The recent finding that early deficiency of elastin modifies vascular ageing has raised the possibility that fibrillin-1 deficiency could also contribute to late-onset pathology of vascular remodelling. To address this question, we examined cardiovascular function in 3-week-old, 6-month-old, and 24-month-old mice that are heterozygous for a hypomorphic structural mutation of fibrillin-1 (Fbn1{+/mgΔ} mice). Our results indicate that Fbn1{+/mgΔ} mice, particularly those that are 24 months old, are slightly more hypotensive than wild-type littermates. Additionally, aneurysm and aortic insufficiency were more frequently observed in ageing Fbn1{+/mgΔ}$ mice than in the wild-type counterparts. We also noted substantial fragmentation and decreased number of elastic lamellae in the aortic wall of Fbn1{+/mgΔ} mice, which were correlated with an increase in aortic stiffness, a decrease in vasoreactivity, altered expression of elastic fibre-related genes, including fibrillin-1 and elastin, and a decrease in the relative ratio between tissue elastin and collagen. Collectively, our findings suggest that the heterozygous mgΔ mutation accelerates some aspects of vascular ageing and eventually leads to aortic manifestations resembling those of Marfan syndrome. Importantly, our data also indicate that vascular abnormalities in Fbn1{+/mgΔ} mice are opposite to those induced by elastin haploinsufficiency during ageing that affect blood pressure, vascular dimensions, and number of elastic lamellae.

Figure 1

Effect of aging and genotype on the survival rate (A), aneurismal ascending aorta of 24-month-old Fbn1^+/mgΔ mouse (B) (the arrow indicate the saccular aneurysm at the vessel root), and histological examination of semi-thin sections of ascending aorta of 3-week-old, 6- and 24-month-old mice (C). Eosin/Hematoxylin staining of cross-sections of ascending aorta of Fbn1^+/+, non-pathological Fbn1^+/mgΔ, and pathological Fbn1^+/mgΔ (Patho) 24-month-old mice (D). n=45 for (A). Bar sizes: 1mm (B) and 500μm (D).

Figure 2

Histological examinations of paraffin-embedded cross-sections of the ascending aorta of 3-week-old, 6- and 24-month-old Fbn1^+/+ and Fbn1^+/mgΔ mice, and 24-month-old pathological Fbn1^+/mgΔ mice (Patho). The elastic lamellae were thinner and disorganized in adult and aged Fbn1^+/mgΔ mice. H/E: Hematoxylin/Eosin staining. Bar size: 50 μm.

Figure 3

Diameter-pressure curves and derived mechanical parameters of the ascending aorta of 3-week-old Fbn1^+/+ and Fbn1^+/mgΔ mice. *Significant difference between Fbn1^+/+ and Fbn1^+/mgΔ mice. n=4–7 per group.

Figure 4

Diameter-pressure curves and derived mechanical parameters of the ascending aorta of 6- and 24-month-old Fbn1^+/+ and Fbn1^+/mgΔ mice. *,+Significant difference between 6-month-old Fbn1^+/+ and Fbn1^+/mgΔ mice or between 24-month-old Fbn1^+/+ and Fbn1^+/mgΔ mice, respectively. □,ΔSignificant difference between 6–month-old and 24-month-old values for Fbn1^+/+ and Fbn1^+/mgΔ, respectively. n=5–6 per group.

Figure 5

Desmosine and hydroxyproline contents (A–F) and elastic fiber-related gene expression (G–K) of the ascending aorta of 3 weeks, 6 and 24-month-old Fbn1^+/+ and Fbn1^+/mgΔ mice. D,E present the desmosine and hydroxyproline ratios relative to total protein content. The general effect of genotype (G) and age (A) was assessed by two-way ANOVA separately in each condition (A to K), and followed by Fisher’s LSD test for paired comparisons: *,#Significant difference between Fbn1^+/+ and Fbn1^+/mgΔ mice of the same age, and between 6- and 24-months-old mice of the same genotype, respectively. §Significant difference between young (3-week-old) on one hand and 6- and 24-month old animals of the same genotype on the other hand. +Significant difference between young (3-week-old) and 6-month old animals of the same genotype on the other hand. For desmosine and hydroxyproline dosages: n=5–7 (6-month-old mice) and n=3–5 (24-month-old mice) in each group. For mRNA expression: n=6 (3-week-old), n=8 (6-month-old) and n=5–7 (24-month-old) in each group.

Figure 6

Reactivity to vasoactive agents of the ascending aorta of 6- and 24-month-old Fbn1^+/+ and Fbn1^+/mgΔ. (A,B) 10μmol/L phenylephrine (PE). 24-month-old Fbn1^+/mgΔ mice were pooled (A) or separated (B) in non-pathological and pathological group. Ach-induced vasorelaxation (10μmol/L phenylephrine + 10μmol/L acetylcholine) in non-pathological vessels is represented as the reversal of the PE-induced vasoconstriction, in percent (C). *Generally significant difference with the control value, independent of age. #Generally significant difference between Fbn1^+/+ and Fbn1^+/mgΔ mice, independent of age. n=3–6 per group.