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Archives of cardiovascular diseases
Volume 109, n° 3
pages 188-198 (mars 2016)
Doi : 10.1016/j.acvd.2015.10.002
Received : 11 May 2015 ;  accepted : 5 October 2015
Krox20 heterozygous mice: A model of aortic regurgitation associated with decreased expression of fibrillar collagen genes
La souris Krox20 +/−  : un modèle d’insuffisance aortique associée à une diminution des collagènes fibrillaires
 

Figure 1




Figure 1 : 

Comparison of EGR2 /Krox20 and COL1A2/Col1a2 expression in aortic valves (AoVs) from patients and heterozygous mice. A, B. Ribonucleic acid was extracted from human AoVs to quantify transcriptional levels of (A) COL1A2 and (B) EGR2 by real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Control AoVs were obtained after heart transplant from three patients with end-stage heart failure with no evidence of valvular defects. Diseased tissues (Diseased) were isolated from the pathological AoVs of four patients with severe AoV dysfunction at the time of valve replacement. C, D. Real-time qRT-PCR was performed on isolated AoVs to quantify Krox20 and Col1a2 expression in Krox20+/− (n =6) and control (n =6) embryos at 3months of age. Experiments were performed in triplicate and normalized on expression from controls. (*P <0.05, Student's t -test). mRNA: messenger ribonucleic acid.


Figure 2






Figure 2 : 

Aortic valve (AoV) function in Krox20+/− adult mice. A. Pulse-wave Doppler analysis from the aorta of 3-month-old control mice demonstrated no AoV dysfunction. B. Pulse-wave Doppler record showing mild aortic regurgitation (AR) in Krox20+/− mice. C. Pulse-wave Doppler record of aortic flow displays severe aortic stenosis with measurement of maximum velocity at 3000mm/s. D. Histograms showing measurements of cardiac variables calculated from control (n =35) and mutant (n =11) mice with mild-to-moderate AR (n =6) and severe AR (n =5). Haemodynamic evaluation revealed increased mean velocity and pressure gradients through the aortic valve of Krox20+/− mice with severe AR. Histograms are expressed as mean±standard error of the mean (*P <0.05, Student's t -test). BPM: beats per minute; IVS: interventricular septum; LV: left ventricular; RR: respiratory rate.


Figure 3




Figure 3 : 

Incidence of aortic valve (AoV) dysfunction in Krox20+/− versus control adult mice. A. Percentage of mice in each subgroup of AoV dysfunction. B. Comparison of AoV dysfunction occurrence in mutant mice aged<7months and>7months. AR: aortic regurgitation.


Figure 4




Figure 4 : 

Krox20 +/− mice have enlarged aortic valve (AoV) leaflets. A–C. Quantification of the surface, length and thickness of the AoV leaflets from Krox20+/− and control mice. D–F. Quantification of the surface, length and thickness of Krox20+/− AoVs at age<7months and>7months. All measurements were calculated from six mice for each genotype. Histograms are expressed as mean±standard error of the mean (*P <0.05, Student's t -test).


Figure 5




Figure 5 : 

Histological analysis of the aortic valve (AoV) from adult Krox20+/− mice. Comparison of (A, E) Masson's trichrome, (B, F) alcian blue, (C, G) orcein and (D, H) Von Kossa staining of the AoV from (A–D) Krox20+/+ and (E–H) Krox20+/− at age 3months (mo). (A, E) Masson's trichrome staining reveals reduction of collagen fibres in Krox20+/− mice compared with control mice. (B, F) Comparison of alcian blue staining demonstrates an increase of proteoglycan in Krox20+/− mice. (C, G) Orcein staining shows no major difference between Krox20+/− and control mice. (D, H) Von Kossa staining shows no signs of calcification in Krox20+/− and control AoVs. * Indicates the presence of black melanocytes in control AoV. Scale bars: 100μm.

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