High-resolution diffusion MRI of rectal cancer ex vivo surgical specimens reveals post-treatment microstructural characteristics of tissue components

Abstract Advances in organ-preserving rectal cancer treatment require accurate restaging after neoadjuvant therapy (NAT). However, standard tools - endoscopy and T 2 -weighted pelvic MRI - struggle to differentiate residual tumour from normal tissue and treatment-induced changes, yielding high specificity but low sensitivity for complete response. Diffusion MRI (dMRI) techniques inform about tissue microstructure, such as directional organisation and cellular packing, yet their utility in post-NAT evaluation remains unexplored. This study performed high-resolution ex vivo dMRI on post-NAT total mesorectal excision (TME) specimens to map diffusion properties of tissue components. Five formalin-fixed samples were scanned at 9.4 T using diffusion-tensor (DTI) and diffusion-kurtosis (DKI) alongside multi-echo T 2 mapping, co-registered to histology via morphological landmarks and analysed using a linear mixed-effects model. Muscularis propria showed the highest fractional anisotropy (FA), allowing direction-encoded colour FA maps to distinguish muscular layers and identify histologically confirmed tumour invasion. While T 2 mapping only distinguished fibrosis ( p < 0.05), diffusion metrics provided broader differentiation: tumour exhibited the lowest mean diffusivity (0.0006 ± 0.0001 mm²/s) and elevated mean kurtosis (0.87 ± 0.06), whereas fibrosis showed higher diffusivity (0.001 ± 0.0002 mm²/s) compared to normal wall. Ultimately, advanced dMRI distinguishes tissue components beyond conventional contrast by providing microstructural information, requiring validation in larger cohorts as biomarkers for treatment response and ypT staging.