Cardiac phenotyping in ex vivo murine embryos using µMRI

Abstract

Microscopic MRI (microMRI) is an emerging technique for high-throughput phenotyping of transgenic mouse embryos, and is capable of visualising abnormalities in cardiac development. To identify cardiac defects in embryos, we have optimised embryo preparation and MR acquisition parameters to maximise image quality and assess the phenotypic changes in chromodomain helicase DNA-binding protein 7 (Chd7) transgenic mice. microMRI methods rely on tissue penetration with a gadolinium chelate contrast agent to reduce tissue T(1), thus improving signal-to-noise ratio (SNR) in rapid gradient echo sequences. We investigated 15.5 days post coitum (dpc) wild-type CD-1 embryos fixed in gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA) solutions for either 3 days (2 and 4 mM) or 2 weeks (2, 4, 8 and 16 mM). To assess penetration of the contrast agent into heart tissue and enable image contrast simulations, T(1) and T(*) (2) were measured in heart and background agarose. Compared to 3-day, 2-week fixation showed reduced mean T(1) in the heart at both 2 and 4 mM concentrations (p < 0.0001), resulting in calculated signal gains of 23% (2 mM) and 29% (4 mM). Using T(1) and T(*) (2) values from 2-week concentrations, computer simulation of heart and background signal, and ex vivo 3D gradient echo imaging, we demonstrated that 2-week fixed embryos in 8 mM Gd-DTPA in combination with optimised parameters (TE/TR/alpha/number of averages: 9 ms/20 ms/60 degrees /7) produced the largest SNR in the heart (23.2 +/- 1.0) and heart chamber contrast-to-noise ratio (CNR) (27.1 +/- 1.6). These optimised parameters were then applied to an MRI screen of embryos heterozygous for the gene Chd7, implicated in coloboma of the eye, heart defects, atresia of the choanae, retardation of growth, genital/urinary abnormalities, ear abnormalities and deafness (CHARGE) syndrome (a condition partly characterised by cardiovascular birth defects in humans). A ventricular septal defect was readily identified in the screen, consistent with the human phenotype.

The Cardiac Phenotype in Patients With a CHD7 Mutation

“…For clinical practice, this means that one should always be aware of other features of CHARGE syndrome, like coloboma, choanal atresia, cranial nerve dysfunction, balance problems, characteristic ear anomalies (triangular conchae or cup ear), or hypogonatropic hypogonadism, in patients with congenital heart defects especially in patients with AVSD and conotruncal defects. 5,24 Several types of heart defects are found in animal models with heterozygous CHD7 mutations: ventricular septal defects in adult mice, 25,26 ventricular septal defects and defects of the pharyngeal arch arteries in mouse embryos, 27 and abnormal positioning of the truncus arteriosus and cardiac outflow tract in Xenopus laevis (African clawed frog). 28 This finding suggests that CHD7 must be involved in several steps of the cardiac embryogenesis, and especially in the formation of the outflow tract and the atrioventricular cushion.…”

Imaging modalities to assess structural birth defects in mutant mouse models

“…The MRI system detects changes in the magnetic field of hydrogen atoms within the sample, converting this information into 2D or 3D images of the sample. For small animal imaging, MRI systems with high-magnetic field strengths (micro-MRI) with 7 to 14-Tesla magnets are commercially available (Schneider et al, 2004;Yelbuz et al, 2004;Cleary et al, 2009;Pallares et al, 2009;Parnell et al, 2009;French et al, 2010;Yamada et al, 2010;Zouagui et al, 2010). These systems have imaging resolution of 20 lm, similar to micro-CT and better than ultrasound biomicroscopy.…”

Dimensions of the Great Intrathoracic Arteries of Early Mouse Fetuses of the C57BL/6 Strain

“…This includes optical projection tomography (OPT) [16,17], micro-Magnetic resonance imaging ( MRI) [18,19], and micro-computed tomography ( CT) [6,20,21].…”