MeSiFuN: A toolbox to map mesenchymal signaling networks in developing human tissues
June 25
Intercellular signaling correlations obtained by wide-field voltage-sensitive and Ca2+-sensitive optical imaging of epithelial sheets have yielded robust indicators of normal and pathological morphogenesis. Almost half of the embryonic body mass, however, is composed of loosely organized mesenchyme, and recent work has revealed rich, stimulus-locked bioelectric and second-messenger activity within this compartment. Yet current correlation analyses remain confined to epithelial regions, and optical data from mesenchyme are generally discarded. Here, we introduce the Mesenchymal Signaling Functional Networks (MeSiFuN) Toolbox, purpose-built for mesenchymal network analysis. The pipeline incorporates preprocessing routines that suppress epithelial bleed-through, optimized algorithms for isolating bona fide mesenchymal voltage and Ca2+ transients, and dedicated statistics and visualization modules for mesoscale network topology. MeSiFuN relies on the standard FIJI/ImgLib2 backbone, includes generalized linear-mixed modeling for cohort-level comparisons, and is wrapped in an intuitive graphical interface that executes every step from raw image stacks to final group statistics without requiring coding expertise. To showcase MeSiFuN’s capabilities, we analyzed time-lapse recordings from 98 sex-annotated human gastruloid specimens available in the HovOrg public repository. Significant sex-biased coupling emerged between the left mid-lateral plate mesoderm and a signaling module encompassing bilateral posterior paraxial mesoderm and dorsal pre-somitic tissue. MeSiFuN offers the developmental and regenerative medicine community an accessible platform to explore embryonic patterning as an integrated dialogue between epithelial and mesenchymal compartments.