Tissue biomechanics during cranial neural tube closure measured by Brillouin microscopy and optical coherence tomography

Abstract

Background: Embryonic development involves the interplay of driving forces that shape the tissue and the mechanical resistance that the tissue offers in response. While increasing evidence has suggested the crucial role of physical mechanisms underlying embryo development, tissue biomechanics is not well understood because of the lack of techniques that can quantify the stiffness of tissue in situ with 3D high-resolution and in a noncontact manner.

Methods: We used two all-optical techniques, optical coherence tomography (OCT) and Brillouin microscopy, to map the longitudinal modulus of the tissue from mouse embryos in situ.

Results: We acquired 2D mechanical maps of the neural tube region of embryos at embryonic day (E) 8.5 (n = 2) and E9.5 (n = 2) with submicron spatial resolution. We found the modulus of tissue varied distinctly within the neural tube region of the same embryo and between embryos at different development stages, suggesting our technique has enough sensitivity and spatial resolution to monitor the tissue mechanics during embryonic development in a noncontact and noninvasive manner.

Conclusions: We demonstrated the capability of OCT-guided Brillouin microscopy to quantify tissue longitudinal modulus of mouse embryos in situ, and observed distinct change in the modulus during the closure of cranial neural tube. Although this preliminary work cannot provide definitive conclusions on biomechanics of neural tube closure yet as a result of the limited number of samples, it provides an approach of quantifying the tissue mechanics during embryo development in situ, thus could be helpful in investigating the role of tissue biomechanics in the regulation of embryonic development. Our next study involving more embryo samples will investigate systematic changes in tissue mechanics during embryonic development.

Keywords: Brillouin microscopy; OCT; development; embryo; tissue biomechanics.

Figure 1.

Representative 3D images of mouse embryos acquired by OCT. (a) E8.5, (b) E9.5. The red dashed line indicated the cross-section of the neural tube scanned by Brillouin microscope. Both scale bars are 100 μm.

Figure 2.

Tissue stiffening of the neural tube during embryonic development. (a) and (b) are OCT cross-sectional images of representative E8.5 and E9.5 embryos. Dashed yellow boxes indicate the imaged region by Brillouin microscope. (c) and (d) are corresponding Brillouin images at the same cross-sections; the red dashed lines indicate the neural folds. (e) averaged Brillouin shift of the neural tube tissues of E8.5 (n=2) and E9.5 (n=2) embryos. All scale bars are 100 μm.

Figure 3.

Tissue modulus of the neural folds shows a gradient along dorsal-ventral direction. (a)-(c) are OCT cross-sectional images of the neural tube of a representative E9.5 mouse embryo. Dashed yellow boxes indicate the imaged region by Brillouin microscopy. (d)-(f) are corresponding Brillouin images at the same cross-sections; the neural tube is artificially segmented into different sub-regions (red lines) to quantify averaged modulus locally, and the results are shown in (g)-(i). Red arrows indicate the region of neural tube fusion is distinctly softer than other part of neural folds; Green arrows indicate the curve trend. All scale bars are 100 μm.

Figure 4.

(a)-(c) are Brillouin images of neural tubes at different levels of a representative E9.5 embryo (same as Fig. 3d-3f); (d)-(f) are averaged Brillouin shifts (mean ± s.d.) of neural tube tissues (region indicated by red dashed line) and ectoderm layers (region indicated by white dashed line); NT(left) and NT(right) indicate the left and right part of the neural fold, respectively. All scale bars are 100 μm.