ABSTRACT
BACKGROUND: The epididymis is important for sperm maturation and without its proper development, male infertility will result. Biomechanical properties of tissues/organs play key roles during their morphogenesis, including the Wolffian duct. It is hypothesized that structural/bulk stiffness of the capsule and mesenchyme/extracellular matrix that surround the duct is a major biomechanical property that regulates Wolffian duct morphogenesis. These data will provide key information as to the mechanisms that regulate the development of this important organ. OBJECTIVES: To measure the structural/bulk stiffness in Pascals (force/area) of the capsule and the capsule and mesenchyme together that surrounds the Wolffian duct during the development. To examine the relative membrane tension of mesenchymal cells during the Wolffian duct development. Since Ptk7 was previously shown to regulate ECM integrity and Wolffian duct elongation and coiling, the hypothesis that Ptk7 regulates structural/bulk stiffness and mesenchymal cell membrane tension was tested. MATERIALS AND METHODS: Atomic force microscopy and a microsquisher compression apparatus were used to measure the structural stiffness. Biomechanical properties within the membranes of cells within the capsule and mesenchyme were examined using a membrane-tension fluorescent probe. RESULTS AND DISCUSSION: The structural stiffness (Pascals) of the capsule and underlying mesenchyme was relatively constant during development, with a significant increase in the capsule at the later stages. However, this increase may reflect the ECM and associated mesenchyme being close to the capsule because the coiling of the duct pushed or compressed them into that space. Keeping the capsule and mesenchyme/ECM at constant stiffness would ensure that the duct will continue to coil under similar biomechanical forces throughout the development. Cells within the capsule and mesenchyme at different Wolffian duct regions during the development had varying degrees of membrane lipid tension. It is hypothesized that the dynamic changes ensure the duct is kept at a constant stiffness regardless of any external forces. Loss of Ptk7 resulted in an increase in stiffness at E18.5, which was presumable due to the loss of integrity of the ECM within the mesenchyme. CONCLUSION: Biomechanical properties of the capsule and the mesenchyme/extracellular matrix that surround the Wolffian duct play an important role toward Wolffian duct morphogenesis, thereby allowing for the proper development of the epididymis and subsequent male fertility.