Quantitative three‐dimensional imaging of live avian embryonic morphogenesis via micro‐computed tomography

Abstract

Many clinically relevant congenital malformations arise during mid to late embryonic stages. This period is challenging to image quantitatively in live embryos, necessitating the use of multiple specimens with increased experimental variability. Here we establish X-ray and blood-pool computed tomography (CT) contrast agent toxicity and teratogenesis thresholds for 3D Micro-CT imaging of live avian embryos. Day 4 chick embryos micro-injected with Visipaque™ (VP) developed for an additional 6 days without defect. X-ray radiation up to 798 mGy was nontoxic. Peak average contrast of 1,060 HU occurred within 1 hr of imaging at 50 μm resolution. VP-enhanced contrast persisted past 24 hr with delayed accumulation in the allantois. Regional volumes of VP-injected embryos were statistically identical to those of fixed embryos perfused with osmium tetroxide. We further quantified longitudinal volumetric morphogenesis of the allantois over 30 hr. These results demonstrate the safety and efficacy of contrast enhanced quantitative micro-CT imaging for live embryos.

3D Reconstruction of Chick Embryo Vascular Geometries Using Non-invasive High-Frequency Ultrasound for Computational Fluid Dynamics Studies

“…The same challenges were addressed by some other investigators, highlighting the recent attention placed on the development of a non-destructive high-resolution scanning method and its importance. For example, Henning et al 17 developed a technique for injecting non-toxic contrast agents through the vitelline vessels and for subsequent high resolution micro-CT imaging of the chick embryo. The authors found that their technique could similarly be used repeatedly on the same chick embryo without harming it, thus enabling longitudinal studies like our technique.…”

Micro/Nano-Computed Tomography Technology for Quantitative Dynamic, Multi-scale Imaging of Morphogenesis

“…Omnipaque is administered to a live specimen but it is lethal within 24 hours making it ineffective for longitudinal studies (Henning, Jiang et al 2011)…”

“…Visipaque is administered to a live specimen and has shown to be nonembryotoxic (Henning, Jiang et al 2011)…”

“…Henning and colleagues demonstrated the first in vivo live embryonic study using microCT imaging. Henning et al quantified the level of contrast, biodistribution and organ volume of developing chick embryos (Figure 5, (Henning, Jiang et al 2011)). Furthermore, Henning and colleagues demonstrated that radiation from the scan up to 798mGray did produce any morphological defects up to day 10 of chick development when the experiment was ended (Henning, Jiang et al 2011).…”

“…Visipaque™ (VP) and Omnipaque™ (OP) were used as exogenous soft tissue contrast. OP was found to be embryotoxic within 24 hours post-injection due to being hyperosmotic whereas iso-osmotic VP was non-toxic up to day 10 of development (Henning, Jiang et al 2011). Additionally, VP produced 1060 Hounsfield units of contrast with a 50µm resolution microCT scan (Henning, Jiang et al 2011).…”