Abstract
Periodic myometrial contraction is one of the important uterine functions to achieve embryo implantation and parturition. Although it is well-known that the mammalian myometrium is composed of longitudinal (outer) and circular (inner) layers, the precise mechanisms that coordinate both muscular contractions to produce peristaltic movements remain unclear. Recently, by treatment with our modified Clear Unobstructed Brain Imaging Cocktails and Computational analysis (CUBIC) tissue-clearing method, we obtained well-contrasted three-dimensional images of the transparent murine ovary using enhanced green fluorescent protein (EGFP) transgenic mice and light-sheet microscopy. Consequently, to investigate accurate anatomical connections between outer and inner myometrial fibers, we observed whole structures of the myometrium using a transparent murine uterus. By this method, we identified a novel muscle layer, a middle layer of the myometrium, which anatomically connects the conventional outer longitudinal and inner circular muscles. This new layer was visualized as a mesh-like structure and this structure was observed throughout the whole uterus from proximal to distal sites. In this area, CD31-positive vessels were abundantly localized around the mesh-like muscle fibers. In addition, CD34-positive uterine telocytes and tubulin β-3-positive nerve fibers were closely located in this middle layer. These findings indicate the presence of a novel mesh-like stratum that connects longitudinal and circular muscle layers, and suggest its coordinating role in myometrial contractions.
Highlights
Periodic myometrial contraction is one of the important uterine functions to achieve embryo implantation and parturition
Using a modified CUBIC tissue-clearing method, we succeeded in making the pregnant murine uterus transparent and analyzing the specific distribution of the embryo-derived trophoblast that had invaded toward maternal uterine muscle layer[17]
Our modified CUBIC method combined with light-sheet microscopy successfully provided clear 3D images of the murine uterus
Summary
Periodic myometrial contraction is one of the important uterine functions to achieve embryo implantation and parturition. We could obtain well-contrasted and 3D images of the whole ovary under light-sheet microscopy using a tissue-clearing technique and EGFP transgenic mice[18] Based on this advantage, we applied our modified CUBIC tissue-clearing method to the murine uterus to analyze the 3D structure of myometrium and elucidate the physiological mechanism of uterine contraction. We identified a novel mesh-like muscle structure, a middle layer of myometrium, which anatomically connects the conventional outer longitudinal and inner circular muscle layers Since this structure is one of the candidates to explain the mechanisms coordinating uterine peristaltic contractions, we further analyzed the mesh-like muscle region by immunohistochemical study together with 3D imaging under light-sheet microscopy
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