Association between overactive bladder and pelvic organ mobility as evaluated by dynamic magnetic resonance imaging

Overactive bladder (OAB) is a prevalent condition, which negatively impacts patients’ quality of life. Pelvic organ prolapse (POP), also prevalent in women, has been recognized as an important etiology of female OAB, although the pathophysiological mechanisms remain controversial. In this study, we reviewed findings of dynamic magnetic resonance imaging (dMRI) in 118 patients with POP and investigated the association between dMRI findings, including positions and mobilities of pelvic organs as well as parameters of pelvic organ support and bladder outlet obstruction (urethral kinking), and OAB in order to elucidate the pathophysiology of OAB in patients with POP. Our results showed that compared with non-OAB patients, OAB patients had a significantly higher body mass index, more severe pelvic floor muscle impairment, and more profound supportive defects in the uterine cervix (apical compartment). On the other hand, dMRI parameters showed hardly any significant difference between patients with mild and moderate to severe OAB. These findings may imply that levator ani impairment and defective supports of the apical compartment could be associated with the presence of OAB and that the severity of OAB could be affected by factors other than those related to pelvic organ mobility and support or urethral kinking.

Findings by presence or absence of OAB. Thirty-five (29.7%) patients had OAB, and patient characteristics by the presence or absence of OAB are shown in Table 1. Compared with non-OAB patients, OAB patients showed significantly higher BMI and a lower proportion of patients who smoke.
The findings of coordinate positions and POMs of pelvic organs as well as parameters of pelvic organ support and urethral kinking are shown in Table 2 and Figs. 1 and 2. In terms of coordinate points and POMs, OAB patients had significantly greater caudal uterine cervix (C) and anorectal angle (AR) at rest, and longer hiatal length at rest than non-OAB patients. During straining, C was located significantly more ventrally as well as obliquely downward in OAB patients than in non-OAB patients, which was reflected by the larger distance between the x coordinate positions (Cxx) as well as the larger diagonal distance (Cp) of C before and during straining. These movements of the apical compartment in OAB patients were accompanied by a significantly more extended imaginary uterosacral ligament (iUSL) as well as imaginary cardinal ligament (iCL) than in non-OAB patients. On the other hand, the degree of rotational descent of the urethra did not show a statistically significant difference between OAB patients and non-OAB patients. When the posterior urethrovesical angle (PUVA) was divided into groups below and above the upper limit of normal (115°) 21 , the proportion with OAB did not differ [29.9% (20/67) vs. 29.4% (15/51), respectively].
Of variables showing a statistically significant difference in Table 2, BMI, the y coordinate of AR (ARy) at rest, and strain on iUSL (iUSLε) were selected for multivariable logistic regression analysis. Odds ratios (ORs) Table 1. Characteristics of patients with or without overactive bladder (OAB). The p-value for numerical variables was calculated by Student's t-test except the p-values with a superscript (*), for which the Wilcoxon rank sum test was used. BMI, body mass index; CI, confidence interval; OABSS, overactive bladder symptom score; POP-Q, pelvic organ prolapse quantification system; SCIPP, sacrococcygeal inferior pubic point.

Discussion
To the best of our knowledge, no study has reported the association between parameters on dMRI and OABSS in patients with POP. In the present study, approximately 30% of the patients had OAB. We demonstrated that BMI, smoking status, levator ani impairment, and defective supports of the apical compartment were associated with the presence of OAB. On the other hand, all but one (Bx) of the parameters on dMRI showed no significant difference between patients with mild OAB and those with moderate to severe OAB. Considering the findings of AR and the length of the urogenital hiatus (H-line), levator ani impairment could predispose patients to OAB. In contrast to "at rest", the indexes on AR as well as the length of the H-line "during straining" did not show any statistical differences between OAB and non-OAB patients. This suggests that levator ani impairment in our patients was severe enough to cause OAB, so that any further caudal movement of AR as well as extension of the urogenital hiatal length no longer affected the presence or absence of OAB. The levator ani impairment potentially causes pelvic organ descent and vaginal wall protrusion below the line through the hiatus 22 . Consequently, a portion of the anterior vaginal wall is exposed to the pressure differential between high intra-abdominal pressure and low atmospheric pressure, which generates tension in the vaginal wall and acts as a downward force that drags the vaginal wall and apex downward with the force becoming stronger  Table 2. Parameters on dynamic magnetic resonance imaging between patients with or without overactive bladder (OAB). x and y indicate the x and y coordinates of each pelvic organ point (AR, B, BN, and C). xx and yy indicate the distance between coordinate positions of the pelvic organ points before and during straining in x and y directions, respectively. p indicates the distance derived from the Pythagorean theorem. Note that the effect of pelvic organ mobility (POM) on the X-axis is negative when moving in the ventral direction and that of POM on the Y-axis is negative when moving in the caudal direction. Also, note that all diagonal POMs are positive. www.nature.com/scientificreports/ in proportion to the exposed vaginal wall length 14 . Therefore, levator ani impairment could have a profound adverse effect on the apical supports. On the other hand, defects in apical supports are associated with larger cystocele 23 . Chen et al. 23 reported that an 80% impairment in apical support resulted in a 33% larger cystocele size. Both the uterosacral ligament (USL) and cardinal ligament (CL) play a critical role in apical supports 23,24 . When these structures are compromised, the resultant larger cystocele, that is, larger anterior vaginal wall protrusion, develops 23 . Moreover, Petros et al. hypothesized that debilitated suspensory function of pubourethral and uterosacral ligaments weakened the musculo-elastic stretching mechanism of the vagina which supported the bladder base stretch receptors 25 . The consequent lax vagina is not sufficient to prevent activation of the bladder base stretch receptors [25][26][27] . Eventually, stretching of the bladder wall might trigger the stretch receptors in the urothelium, which causes a prematurely activated micturition reflex, leading to OAB 2,25-28 . This implies that female OAB may be derived from a ligamentous-fascial disorder remote from the symptomatic organ 16 , which could explain why supports for an apical compartment, rather than the positions and POMs of the bladder neck (BN) or the most dependent position of the bladder (B), are significantly associated with the presence of OAB in the present study. In this respect, the present results may be considered to be in line with the previously reported posterior fornix syndrome attributed to defects in apical supports, which consists of urgency/frequency, nocturia, chronic pelvic pain, and abnormal emptying 5 . In fact, compared with controls, patients with POP showed an increase in the deep USL length from resting to straining of 15 ± 12 mm vs. 7 ± 4 mm, while CL was 30 ± 16 mm vs. 15 ± 9 mm 24 , which would be compatible with our findings on iUSL and iCL. The present study showed that compared with non-OAB patients, positions of C were significantly more ventral as well as caudal and POMs of C were significantly greater in OAB patients. Moreover, strain on iUSL and iCL (iUSLε and iCLε, respectively) was significantly larger in patients with OAB. Takazawa et al. 9 showed that the transvaginal surgery with mainly apical support significantly improved OABSS total score from 4 to 2 points, suggesting that the importance of providing support for level I in improving OAB. Liedl et al. 29 demonstrated that cardinal/uterosacral ligament repair using a Tissue Fixation System for symptomatic apical prolapse ≥ stage II cured urgency incontinence, frequency, and nocturia in 85%, 83%, and 68% of patients, respectively. Also, an Elevate technique, which uses mesh arms attached to the sacrospinous ligaments to recreate apical ligamentous support, significantly improved all OAB symptoms 5 . Taken together, the levator ani impairment and worse apical supports would be a fundamental factor that is associated with the presence of OAB in patients with POP. In addition to the abnormally triggered micturition reflex by anterior vaginal prolapse 2 , neurodegeneration in the bladder wall seen in OAB should be considered based on the findings on iUSL and iCL. The USL is situated in close proximity to the hypogastric nerve (HN) and inferior hypogastric plexus (IHP) 30 , and a deeper portion www.nature.com/scientificreports/ of USL as well as the neural part of CL involves autonomic fibers from HN and IHP, and a part of IHP itself 30 . Therefore, it is reasonable that peripheral autonomic fibers travelling to and from the bladder are assumed to be subjected to cyclic strain in POP, which might cause partial denervation of the bladder wall. In fact, the density as well as the diameters of nerves in USL and CL in women with POP were significantly reduced compared with those without POP 2,31 . To the best of our knowledge, it remains to be determined whether cyclic strain of USL and CL in POP affects autonomic nervous fibers morphologically and functionally. A rapid recovery of the peripheral somatic nervous function is generally shown after 4% to 21% stretching of peripheral nerves 32 , although 5-25% stretching can cause nerve rupture 33 . Assuming that the autonomic nerve fibers are strained to the same degree as those ligaments, the findings on iUSLε and iCLε suggest that the strain on the nerve fibers would exceed these proposed ranges. Taken together, the extension of USL and CL might develop partial denervation of the bladder, which induces OAB. As far as PUVA and AUI are concerned, BOO would not be strongly associated with the presence or absence and the severity of OAB. BOO is often regarded as an important mechanism for developing OAB in patients with POP 2 , although the exact mechanism of BOO in POP is not fully understood 34 . The mechanisms of BOO include the urethral kinking or urethral and/or BN compression by the prolapsed organs, which has been diverted from the urinary continence mechanism in POP 34,35 . A rapid upswing in urethral pressure in any segment along the entire urethral length, the downward movement of the bladder and posterior urethra against the immobile distal urethra, or an intact PUVA with significant BN descent during straining in patients with POP not only provide continence but also cause BOO [36][37][38] . However, there have been no reports to date that have clearly demonstrated the exact site of urethral kinking or urethral compression during the voiding phase 34 . In the present study, the exact site of urethral kinking could not be identified on dMRI because the entire urethral morphology was difficult to evaluate on the midline sagittal plane during straining. Moreover, the intact PUVA, which is supposed to be one of the indicators of the urethral kinking in POP 38 , was not associated with the presence of OAB. Therefore, mechanical BOO due to the urethral kinking or urethral and/or BN compression would not fully explain a mechanism of OAB in patients with POP. In this regard, we may have to pay attention to a role of the pelvic floor in opening and closing of BN as proposed by Petros et al. 39,40 , which illustrates that selective and specific directional contractions of the pelvic floor muscles stretch the vagina against intact pubourethral and USLs to facilitate opening and closure of the urethra and BN. Later, Bush et al. 41 used a finite element model and demonstrated that backward/downward-acting pelvic floor muscles enabled normal micturition. Damage in the ligamentous structures could diminish the action of pelvic floor muscles, which impairs adequate opening of the urethra and BN, so-called functional BOO 26 . In fact, apical sling operations improved abnormal bladder emptying as well as OAB in patients with POP 42,43 . Taken together, the present dMRI findings suggest that the levator ani impairment and supportive defects in the apical compartment potentially cause functional BOO, which might be more likely to be involved in OAB in patients with POP, rather than mechanical BOO. www.nature.com/scientificreports/ Although parameters evaluated on dMRI would be associated with OAB, none of the parameters except Bx at rest was associated with the severity of OAB. On the other hand, hypertension tended to be associated with it, and an association between the severity of OAB and BMI as well as age was also suggested, which would indicate that the severity of OAB would be more affected by systemic factors such as neuroendocrine, vascular, or inflammatory processes than by POP itself 44,45 . Caution should be exercised with regard to interpreting the results because the number of patients with OAB was small in this study. While levator ani impairment and defective apical supports could be associated with OAB, the severity of OAB would be more affected by systemic factors than by POP itself, which might partly explain the inconsistent results of relationship between OAB and POP in the literature 4,5 . Further studies are needed to determine the relationship between effects of systemic factors and parameters evaluated in dMRI on the occurrence and severity of OAB.
This study has several limitations. First, dMRI was performed in a supine, rather than a standing, position because our magnetic resonance imaging (MRI) equipment did not allow upright examination. Also, we did not measure intra-abdominal pressure during the Valsalva maneuver. However, dMRI was conducted by one specialized and experienced technician with a consistent protocol, and we included only patients who were able to reproduce POP ≥ stage II during straining. Second, we measured POMs with coordinates of selected landmarks at rest and during straining. Strictly speaking, the coordinates at rest would not correspond to the "normal positions" of the patients. However, we do not have data on the "normal positions" because we have not performed dMRI on nulliparous women without any symptoms, and a definition of the "normal positions" on dMRI of Asian women has not been established. Moreover, POMs should be adjusted to the patient's height or pelvic bone geometry 13,46 . However, use of these corrected methods is not widespread yet. Third, we included patients with POP ≤ stage II at rest and POP ≥ stage II during straining. Therefore, we did not evaluate the association between mild (< stage II during straining) POP and OAB as well as between severe (≥ III at rest) POP and OAB. Fourth, we have not taken MRIs of patients after POP repair. Therefore, we did not investigate whether decreased POMs after POP repair correlated with the improvement of OAB.
In conclusion, considering the parameters derived from dMRI, levator ani impairment and defective supports of the apical compartment were associated with the presence of OAB, while almost all parameters derived from dMRI were not associated with the severity of OAB, which could be affected by systemic factors.

Patients and methods
This retrospective study was conducted under the approval of the Clinical Research Ethics Committee at Yotsuya Medical Cube (ID: YMCIRB-20R010) and also approved by the Treatise Certificate Committee at Faculty of Medicine, Toho University (ID: 2020-157), by which the Declaration of Helsinki adopted by the World Medical Association, Ethical Guidelines for Medical and Health Research Involving Human Subjects established by the Ministry of Health, Labour and Welfare, and other relevant acts were followed accordingly. According to the Ethical Guidelines, because the present study is a retrospective observational study that did not utilize human biological specimens, it is not necessarily required to obtain written or verbal informed consent from individual patients, but it is necessary to disclose the contents of the study and provide an opportunity for opt-out, which was followed by the Clinical Research Ethics Committee at Yotsuya Medical Cube in approving the present study.
Patients. Patients referred to Yotsuya Medical Cube for management of POP have routinely undergone dMRI to investigate the severity as well as the compartment of POP. Of 334 patients undergoing dMRI between August 2015 and July 2016, we included 148 patients who had POP ≤ stage II at rest and POP ≥ stage II during dMRI because POMs in patients with POP ≥ stage III both at rest and during straining paradoxically reveal small values. We also excluded patients taking OAB medication, with only rectocele and the following co-morbidities or previous histories: neurogenic lower urinary tract dysfunctions, anti-incontinence surgery, POP surgery, intrapelvic gynecologic, gastrointestinal and urologic surgeries, pelvic radiotherapy, and gynecological diseases, e.g., endometriosis, large uterine myoma, and large ovarian cysts. Of 148 patients, we excluded 30 patients due to unevaluable dMRI, which resulted from vigorous body motion during a forced Valsalva maneuver, so that we finally included 118 patients in this study.

Protocols of MRI.
We referred to the MRI protocol described in the study by Larson et al. 12 . The dMRI (1.5T, GE Signa Excite) was conducted by one experienced technician specializing in evaluation of urogynecological patients and was performed in the late afternoon because POP was more easily reproduced than in the morning or early afternoon. Prior to the MRI, the technician instructed patients on how to strain effectively and sufficiently during dMRI. Patients were instructed to drink enough fluid to be conscious of a normal desire to void at dMRI. First, axial as well as sagittal static T2 images were obtained to evaluate the integrity of pelvic floor musculature and connective tissue and the presence or absence of incidental lesions of pelvic organs. Then, dMRI on the midline sagittal plane was performed with consecutive acquisition of MRI images, at rest (2 to 3 images, repetition time [TR] range, 3.2; echo time [TE], minimum; 10 mm slice thickness, 0 mm gap; 1 number of excitations [NEX]) and during the maximal Valsalva maneuver for 10 to 15 s (10 to 15 images TR range, 3.2; TE, minimum; 10 mm slice thickness, 0 mm gap; 1 NEX), to generate cinematic images in the supine position with knees elevated on a high pillow. Dynamic sequences were repeated several times while subjectively monitoring the sufficiency of the Valsalva maneuver. At dMRI, incontinence pads were used to decrease fear of or embarrassment about leakage to allow patients to strain enough.
Methods to evaluate POM (Fig. 3A,B). Using the mid-sagittal dMRI images on the monitor, we measured representative points of the pelvic organs, that is, coordinate positions of the BN, B, C, and AR at rest and during intense straining 10 www.nature.com/scientificreports/ To measure each coordinate of these landmarks, we used a sacrococcygeal inferior pubic point (SCIPP) line connecting the inferior pubic point and the sacrococcygeal junction 15 as an X-axis with the inferior margin of the pubic symphysis as an origin because its identification is more reproducible than the pubococcygeal line (PCL) on dMRI, and a line perpendicular to the SCIPP line at the origin as a Y-axis. The x and y coordinates of each representative point (e.g., BNx, and BNy, respectively) were measured before and during straining.
POM was evaluated by the distance between coordinate positions of the representative points before and during straining in x (e.g., BNxx = BNx during straining -BNx at rest), y directions (e.g., BNyy = BNy during straining -BNy at rest), and the distance derived from the Pythagorean theorem (e.g., BNp = [(BNxx) 2 + (BNyy) 2 ] 1/2 ). Note that POM on the X-axis is negative when moving in the ventral direction and POM on the Y-axis is negative when moving in the caudal direction. Also note that all diagonal POMs are positive.
Methods to evaluate parameters of pelvic organ support and bladder outlet obstruction (Fig. 4A,B). We attempted to measure the length of the USL as well as the CL on dMRI. However, because it was difficult to precisely delineate the courses of these ligaments on the mid-sagittal images of dMRI, the distance from the anterior surface between the fourth and fifth sacral bones (S4/5) to C was measured as a surrogate of the USL as an iUSL 16 , while the distance from the anterior surface between the second and third sacral bones (S2/3) to C was measured as a surrogate of the CL as an iCL 17,47 . The lengths of those imaginary lines were measured at rest and during straining, from which we calculated the strain on them (e.g., iUSL ε = [the length of iUSL during straining-the length of iUSL at rest]/the length of iUSL at rest). We also measured the H-line and of the hiatal descent at rest and during straining 10 . Because we used the SCIPP line rather than PCL as a reference line, the hiatal descent (Mʹ-line) extended perpendicularly from the SCIPP line to the posterior end of the H-line. Moreover, the anterior vaginal wall length (AVWL) at rest and during straining, which was traced from the anterior vaginal fornix to the external urinary meatus 48 , was also measured. The morphology of the entire urethra could not be precisely delineated in dMRI due to its movement during straining. Therefore, using the following parameters, we evaluated urethral kinking as well as mobility: the PUVA, which is the angle between the urethral axis and the posterior border of the bladder base or trigone, and the AUI, which is the angle of the urethral axis in relation to the vertical plane 21 . From these variables, changes in AVWL, PUVA, and AUI were also calculated.
Methods to evaluate OAB. The Japanese Clinical Guidelines for Overactive Bladder Syndrome strongly recommend the use of OABSS for diagnosis as well as evaluation of the severity and treatment outcomes of OAB 49 . In fact, several recent large epidemiological and clinical trials used OABSS as inclusion criteria, assessment of severity, and primary or secondary outcome measures 44,50,51 . The score is the simple sum of four symptom scores, which addresses daytime voiding, nighttime voiding, urgency, and urgency incontinence, with sum www.nature.com/scientificreports/ scores (OABSS total score) ranging from 0 to 15 points 20 . Diagnosing OAB requires urgency scores ≥ 2 and sum scores ≥ 3 on OABSS 20 . In addition, the OABSS total score is classified by a total score of 3-5 as mild, 6-11 as moderate, and 12-15 as severe 20 .
Statistical analysis. We used version 15 of Jmp (SAS Institute Japan Inc., Tokyo) for statistical analysis.
Student's t-test for normally distributed variables, the Wilcoxon signed-rank test for non-normally distributed variables, and the chi-square test for categorical variables were used to evaluate the association between the presence (OABSS ≥ 3) or absence (OABSS < 3) of OAB. Then, using multivariable logistic analysis, we evaluated the OR of the selected variables for the presence of OAB. Finally, variables of patients with mild OAB and those with moderate to severe OAB were compared. Data are shown as means with 95% CI, unless otherwise specified. P < 0.05 was considered to be statistically significant.

Data availability
The datasets generated during and/or analyzed during the current study are not publicly available due to regulations by the Clinical Research Ethics Committee at Yotsuya Medical Cube but are available from the corresponding author on reasonable request. www.nature.com/scientificreports/