In Vivo Evaluation of Cervical Stiffness Evolution during Induced Ripening Using Shear Wave Elastography, Histology and 2 Photon Excitation Microscopy: Insight from an Animal Model

Abstract

Prematurity affects 11% of the births and is the main cause of infant mortality. On the opposite case, the failure of induction of parturition in the case of delayed spontaneous birth is associated with fetal suffering. Both conditions are associated with precocious and/or delayed cervical ripening. Quantitative and objective information about the temporal evolution of the cervical ripening may provide a complementary method to identify cases at risk of preterm delivery and to assess the likelihood of successful induction of labour. In this study, the cervical stiffness was measured in vivo in pregnant sheep by using Shear Wave Elastography (SWE). This technique assesses the stiffness of tissue through the measurement of shear waves speed (SWS). In the present study, 9 pregnant ewes were used. Cervical ripening was induced at 127 days of pregnancy (term: 145 days) by dexamethasone injection in 5 animals, while 4 animals were used as control. Elastographic images of the cervix were obtained by two independent operators every 4 hours during 24 hours after injection to monitor the cervical maturation induced by the dexamethasone. Based on the measurements of SWS during vaginal ultrasound examination, the stiffness in the second ring of the cervix was quantified over a circular region of interest of 5 mm diameter. SWS was found to decrease significantly in the first 4–8 hours after dexamethasone compared to controls, which was associated with cervical ripening induced by dexamethasone (from 1.779 m/s ± 0.548 m/s, p < 0.0005, to 1.291 m/s ± 0.516 m/s, p < 0.000). Consequently a drop in the cervical elasticity was quantified too (from 9.5 kPa ± 0.9 kPa, p < 0.0005, to 5.0 kPa ± 0.8 kPa, p < 0.000). Moreover, SWE measurements were highly reproducible between both operators at all times. Cervical ripening induced by dexamethasone was confirmed by the significant increase in maternal plasma Prostaglandin E2 (PGE2), as evidenced by the assay of its metabolite PGEM. Histological analyses and two-photon excitation microscopy, combining both Second Harmonic Generation (SHG) and Two-photon Fluorescence microscopy (2PF) contrasts, were used to investigate, at the microscopic scale, the structure of cervical tissue. Results show that both collagen and 2PF-active fibrillar structures could be closely related to the mechanical properties of cervical tissue that are perceptible in elastography. In conclusion, SWE may be a valuable method to objectively quantify the cervical stiffness and as a complementary diagnostic tool for preterm birth and for labour induction success.

Estimation of Shear Wave Speed in the Rhesus Macaques’ Uterine Cervix

“…2;31;49;50 Concordant with this, significant differences in SWS estimates between unripened and ripened tissue have been found in vivo and ex vivo in humans 4;6 and sheep. 13 While higher estimate uncertainty in the stiffer macaque cervix could mask small differences in stiffness, a more likely reason is that the misoprostol protocol we used is ineffective in the nonpregnant macaque. In contrast, the efficacy of misoprostol for cervical ripening in both pregnant and non-pregnant humans is well established, as is the efficacy of dexamethasone, the agent used in the study of pregnant ewes.…”

“…We and others have demonstrated its feasibility in humans and animal models. 4;6–8;13 This work represents our initial exploration of SWS estimation in the rhesus macaque model we are developing for comprehensive study of the pregnant cervix. The rhesus macaque carries several advantages with regard to homology with humans.…”

“…In contrast, the efficacy of misoprostol for cervical ripening in both pregnant and non-pregnant humans is well established, as is the efficacy of dexamethasone, the agent used in the study of pregnant ewes. 13 As discussed in the Methods, despite the absence of published reports of misoprostol alone for ripening the macaque cervix, particularly the nonpregnant macaque cervix, we chose this agent in order to harmonize this study with our human studies. This approached seemed reasonable because, it is known that macaques have decreased sensitivity to certain prostaglandins, 22;51 in pregnant macaques, intramuscular or intravenous prostaglandins induce preterm labor and abortion, 36;37 misoprostol is used in combination with other agents in contraceptive studies 14;23 and to induce labor in post-term pregnancy or promote expulsion of retained products of conception, 22;51;52 and intracervical application of a different prostaglandin (PGE2 gel) causes cervical ripening and decreased collagen fibers.…”

“…9–12 SWEI has been studied in vivo in a sheep model. 13 Unfortunately, sheep and rodents are far from ideal for comprehensive study. One issue is that they have bicornuate (two) uteri and they are obligate quadripeds (they walk on four feet, which governs biomechanical forces on the cervix).…”

Assessment of Structural Heterogeneity and Viscosity in the Cervix Using Shear Wave Elasticity Imaging: Initial Results from a Rhesus Macaque Model

“…The parameter used most commonly is shear wave speed group velocity ( c g , the propagation speed of the shear disturbance). Using c g , Peralta et al (2015a) reported an increase in cervical softness after labor induction in ewes. While compelling, mechanical and anatomical differences between sheep and humans may limit clinical applicability of the results.…”