Effects of Elastase Digestion on the Murine Vaginal Wall Biaxial Mechanical Response

Abstract

Although the underlying mechanisms of pelvic organ prolapse (POP) remain unknown, disruption of elastic fiber metabolism within the vaginal wall extracellular matrix (ECM) has been highly implicated. It has been hypothesized that elastic fiber fragmentation correlates to decreased structural integrity and increased risk of prolapse; however, the mechanisms by which elastic fiber damage may contribute to prolapse are poorly understood. Furthermore, the role of elastic fibers in normal vaginal wall mechanics has not been fully ascertained. Therefore, the objective of this study is to investigate the contribution of elastic fibers to murine vaginal wall mechanics. Vaginal tissue from C57BL/6 female mice was mechanically tested using biaxial extension–inflation protocols before and after intraluminal exposure to elastase. Elastase digestion induced marked changes in the vaginal geometry, and biaxial mechanical properties, suggesting that elastic fibers may play an important role in vaginal wall mechanical function. Additionally, a constitutive model that considered two diagonal families of collagen fibers with a slight preference toward the circumferential direction described the data reasonably well before and after digestion. The present findings may be important to determine the underlying structural and mechanical mechanisms of POP, and aid in the development of growth and remodeling models for improved assessment and prediction of changes in structure–function relationships with prolapse development.

Smooth muscle regional contribution to vaginal wall function

“…The unloaded configuration was defined as when the tissue was slightly buckled and not collapsed (3 mmHg) [33,34]. Post explant, the vaginal tissue retracted in length [33,34,38]; therefore, in order to quantify tissue properties under near physiologically relevant loads, the tissue was axially extended [33,34,37,38,41] to an estimated physiological length (1.15 times the unloaded length) [33,34]. Initial preconditioning was performed to minimize hysteresis with cyclic pressurization from 0 mmHg to the mean measured in vivo pressure (7 mmHg) over five cycles at 1.5 mmHg s 21 [33,42].…”

“…The physiological length under basal tone was identified where the transducer-measured forces held constant over a range of increasing physiological pressures [33,34,37,38,41]. Preconditioning with five cycles of pressurization (0-15 mmHg) at the physiological length, and axial extension (10 mm s 21 ) from 2% below to 2% above the physiological length at a constant pressure (5 mmHg) was performed [33,34]. The tissue equilibrated for 10 min at the physiological length under 5 mmHg, followed by re-establishment of the unloaded configuration [33].…”

“…The vaginal tissue was subjected to pressure-diameter testing (0-15 mmHg) over five cycles at and about (22%, þ2% royalsocietypublishing.org/journal/rsfs Interface Focus 9: 20190025 and þ4%) the physiologic length [33,34,37]. This was followed by force -length testing to confirm the physiological length [33,34,37,41]. The tissue was axially extended from 2% below to 2% above the physiological length under four constant pressures: a tare load (P ¼ 2 mmHg), one-third maximum pressure (P ¼ 5 mmHg), two-thirds maximum pressure (P ¼ 10 mmHg) and maximum pressure (P ¼ 15 mmHg).…”

“…Following smooth muscle preconditioning, royalsocietypublishing.org/journal/rsfs Interface Focus 9: 20190025 contraction as a function of axial length was assessed. Pressure was held constant at the mean measured in vivo pressure (7 mmHg), and the tissue was contracted at 4% and 2% below, above and at the physiological length [27,33,34]. After contractile assessment, the passive state was induced and tissue geometry and force values were recorded [27,44].…”

“…Brightfield images were taken using an Olympus BX51 microscope, an Olympus DP27 Digital Camera and cellSens TM software (Olympus Corporation, Center Valley, PA, USA) at 4Â and 20Â magnification. Area fraction quantification was performed on 4Â images displaying the full cross-section with Colour Deconvolution, an open source plug-in for ImageJ, and a custom protocol using GNU Image Manipulation Program as previously described [34,50,51]. The epithelium, which is suggested to be non-load bearing, was excluded from this calculation [34,46].…”