Formalin tissue fixation biases myelin‐sensitive MRI

Abstract

Purpose Chemical fixatives such as formalin form cross‐links between proteins and affect the relaxation times and diffusion properties of tissue. These fixation‐induced changes likely also affect myelin density measurements produced by quantitative magnetization transfer and myelin water imaging. In this work, we evaluate these myelin‐sensitive MRI methods for fixation‐induced biases. Methods We perform quantitative magnetization transfer, myelin water imaging, and deuterium oxide‐exchanged zero TE imaging on unfixed human spinal cord tissue at 9.4 Tesla and repeat these measurements after 1 day and 31 days of formalin fixation. Results The quantitative magnetization‐transfer bound pool fraction increased by 30.7% ± 21.1% after 1 day of fixation and by 42.6% ± 33.9% after 31 days of fixation. Myelin water fraction increased by 39.7% ± 15.5% and 37.0% ± 15.9% at these same time points, and mean T2 of the myelin water pool nearly doubled. Reference‐normalized deuterium oxide‐exchanged zero TE signal intensity increased by 8.17% ± 6.03% after 31 days of fixation but did not change significantly after 1 day of fixation. After fixation, specimen cross‐sectional area decreased by approximately 5%; after correction for shrinkage, changes in deuterium oxide‐exchanged zero TE intensity were nearly eliminated. Conclusion Bound pool fraction and myelin water fraction are significantly increased by formalin fixation, whereas deuterium oxide‐exchanged zero TE intensity is minimally affected. Changes in quantitative magnetization transfer and myelin water imaging may be due in part to delamination and formation of vacuoles in the myelin sheath. Deuterium oxide‐exchanged signal intensity may be altered by fixation‐induced changes in myelin lipid solid‐state 1H T1. We urge caution in the comparison of these measurements across subjects or specimens in different states, especially unfixed versus fixed tissue.

Can MRI measure myelin? Systematic review, qualitative assessment, and meta-analysis of studies validating microstructural imaging with myelin histology

“…Second, the variability in the type and duration of fixation can lead to variability in the MRI-based metrics. Longitudinal studies investigating tissue fixation report spatially and temporally varying effects on relaxation rates, MWF and also MP ( Dawe, Bennett, Schneider, Vasireddi, Arfanakis, 2009 , Seifert, 2019 , Shatl, Uddin, Matsuda, Figley, 2018 ). In the majority of studies we assessed, the fixation process ranged from a few days to a few weeks, which means the MR signal was likely very different between studies.…”

Improved MR fingerprinting for relaxation measurement in the presence of semisolid magnetization transfer

“…A recent publication showed that while comparing to that with conventional method, T 1 in WM measured in the same brains was 10% underestimated with MRF in which TR was from 15 to 55 ms longer than that (from 10 to 12 ms) in our experiment . As compared to the previous in vivo studies, another possible explanation for the increased sensitivity to MT effect in WM in this ex vivo experiment would be from the potential increase of the fractional water binding with myelin and thus the MT due to the delamination and formation of vacuoles in the myelin sheath in the brain tissue fixed with formalin . Although there would be adjustable options for attenuating the MT effect, a 2P model here was used to directly address the MT effect and the proposed framework can be combined with partial or single‐shot k‐space sampling techniques for further acquisition time reduction.…”

Comparative Neuroanatomy of the Lumbosacral Spinal Cord of the Rat, Cat, Pig, Monkey, and Human

“…The comparative atlas created in this study was based on images of perfused spinal cord specimen which may have experienced some shrinkage due to the perfusion process. Siefert et al [48] quantified the level of shrinkage in human spinal cords 1 day and 31 days post formalin fixation and found that the cross-sectional size of the spinal cords shrinks by an average of 4%, and 4.4%, respectively. Therefore, an appropriate correction factor should be taken into consideration when absolute measurement values are needed.…”