Musy, M. | Search bioRxiv

bioRxiv2021

4D reconstruction of developmental trajectories using spherical harmonics

Although the full embryonic development of species such as Drosophila and zebrafish can be 3D imaged in real time, this is not true for mammalian organs, as normal organogenesis cannot be recapitulated in vitro. Currently available 3D data is therefore ex vivo images which provide only a snap shot of development at discrete moments in time. Here we propose a computer-based approach to recreate the continuous evolution in time and space of developmental stages from 3D volumetric images. Our method uses the mathematical approach of spherical harmonics to re-map discrete shape data into a space in which facilitates a smooth interpolation over time. We tested our approach on mouse limb buds (from E10 to E12.5) and embryonic hearts (from 10 to 29 somites). A key advantage of the method is that the resulting 4D trajectory takes advantage of all the available data (i.e. it is not dominated by the choice of a few "ideal" images), while also being able to interpolate well through time intervals for which there is little or no data. This method not only provides a quantitative basis for validating predictive models, but it also increases our understanding of morphogenetic processes. We believe this is the first data-driven quantitative 4D description of limb morphogenesis.

developmental biology↗

bioRxiv2020

Cellular mechanisms of chick limb bud morphogenesis

Although some of the molecular pathways involved in limb bud morphogenesis have been identified, the cellular basis of the process is not yet understood. Proposed cell behaviours include active cell migration and oriented cell division, but ultimately, these questions can only be resolved by watching individual mesenchymal cells within a completely normal developmental context. We developed a minimally-invasive in ovo two-photon technique, to capture high quality time-lapse sequences up to 100 microns deep in the unperturbed growing chick limb bud. Using this technique, we characterized cell shapes and other oriented behaviours throughout the limb bud, and found that cell intercalation drives tissue movements, rather than oriented cell divisions or migration. We then developed a 3D cell-based computer simulation of morphogenesis, in which cellular extensions physically pull cells towards each other, with directional bias controlled by molecular gradients from the ectoderm (Wnts) and the Apical Ectodermal Ridge (FGFs). We defined the initial and target shapes of the chick limb bud in 3D by OPT scanning, and explored which orientations of mesenchymal intercalation correctly explain limb morphogenesis. The model made a couple of predictions: Firstly, that elongation can only be explained when cells intercalate along the direction towards the nearest ectoderm. This produces a general convergence of tissue towards the central proximo-distal (PD) axis of the limb, and a resultant extension of the tissue along the PD axis. Secondly, the correct in silico morphology can only be achieved if the contractile forces of mesenchymal cells in the very distal region (under the Apical Ectodermal Ridge) have shorter life times than in the rest of the limb bud, effectively making the tissue more fluid by augmenting the rate of cell rearrangement. We argue that this less-organised region of mesenchyme is necessary to prevent PD-oriented intercalation events in the distal tip that would otherwise inhibit outgrowth.

developmental biology↗