Role of puboperinealis and rectourethralis muscles as a urethral support system to maintain urinary continence after robot-assisted radical prostatectomy

The present study investigated the role of a urethral support system to maintain urinary continence after robot-assisted radical prostatectomy (RARP), with a focus on pelvic floor muscles, such as the puboperinealis muscle (PPM) and rectourethralis muscle (RUM). Finally, 323 patients who underwent RARP were analyzed in this study. All patients performed a one-hour pad test 1, 3, 6, 9, and 12 months after RARP to assess urinary incontinence and MRI before and 9 months after RARP to evaluate the pelvic anatomical structure. The preoperative cross-sectional area of PPM (2.21 ± 0.69 cm2) was significantly reduced by 19% after RARP (1.79 ± 0.60 cm2; p < 0.01). Positive correlations were observed between the amount of urinary leakage according to the 1-h pad test 1, 3, 6, 9, and 12 months after RARP and the change in the cross-sectional area of PPM by RARP (p < 0.01, < 0.001, < 0.001, < 0.001, and < 0.001, respectively). A positive correlation was also noted between the amount of urinary leakage 6 and 12 months after RARP and the preoperative RUM diameter (p < 0.05). The amount of urinary leakage 1, 3, 6, 9, and 12 months after RARP negatively correlated with the change in the antero-posterior diameter of the membranous urethra (MU diameter) from the static to dynamic phases during the Valsalva maneuver by cine MRI. Furthermore, the change in the MU diameter negatively correlated with the change in the cross-sectional area of PPM (p < 0.05). PPM and RUM play significant roles as a supportive mechanism to maintain urinary continence by functioning as a urethral support.

MRI examinations and imaging techniques.MRI was performed before and 9 months after RARP in all cases because the effects of postoperative inflammation in muscles needed to be excluded.Static and dynamic MRI were performed using a 1.5 T MR scanner (Magnetom Trio; Siemens Healthcare, Erlangen, Germany).Dynamic MRI consisted of a T2-weighted, single slice True Fast Imaging with Steady-state Procession (True-FISP) sequence in the sagittal, coronal, and transverse planes.The cross-sectional area of PPM and left-right (transverse) diameter of RUM (RUM diameter) were measured on T2-weighted fast spin echo (FSE) sequences.The left and right cross-sectional areas of PPM at the height of RUM on the dorsal side of the urethra were calculated using coronal sections, and the sum of the left and right cross-sectional areas was defined as the crosssectional area of PPM (Fig. 1A).The tissue between the urethra and rectum was considered to be RUM in the transverse plane, and the left-right diameter was measured (Fig. 1B).
Patients also underwent cine MRI to evaluate the antero-posterior MU diameter with a full bladder 9 months after RARP (Fig. 2).Cine MRI in static and dynamic phases was conducted during the Valsalva maneuver and T2-weighed sagittal sections were obtained.Cine acquisition was performed over a 30-s time period during the static and dynamic phases.A urologist provided patients with guidance in the examination room during MRI scanning.Changes in the MU diameter from the static (Fig. 2A) to dynamic phases (Fig. 2B) during the Valsalva maneuver by cine MRI were calculated.

Statistical analysis.
All data were analyzed using SPSS statistical package version 24.0 (SPSS Inc., Chicago, IL, USA).The significance of differences was considered when p < 0.05.Parametric comparisons between before and after RARP were performed using the paired t-test.Simple and multiple regression models were used to examine the relationship between two or more variables.Variables with p < 0.05 in the simple regression model were included in the multiple regression model.

Results
Finally, 323 patients (mean age, 67.2 ± 6.0 years; age range, 48-76 years) were enrolled in the present study (Fig. 3).The baseline and perioperative characteristics of patients are shown in Table 1.The average amounts of urinary leakage in the one-hour pad test obtained by weighing the pads were 5.4, 46,1, 22.1, 18.5, 14.3, and 13.0 ml before and 1, 3, 6, 9, and 12 months after RARP, respectively.When urinary continence was defined as   www.nature.com/scientificreports/total urine leakage of < 2 g in the one-hour pad test 10 , continence rates were 85.1, 44.6, 75.5, 75.5, 76.5, and 78.0%before and 1, 3, 6, 9, and 12 months after RARP, respectively (Table 2).We compared the cross-sectional area of PPM and RUM diameter using MRI before and after RARP (Table 3).The preoperative cross-sectional area of PPM (2.21 ± 0.69 cm 2 ) was significantly reduced by 19% after RARP (1.79 ± 0.60 cm 2 ; p < 0.01).No significant change was observed in the RUM diameter between before and after RARP (1.33 ± 0.28 and 1.36 ± 0.29 cm, respectively).There was no significant correlation between the change in  www.nature.com/scientificreports/ the cross-sectional area of PPM and preoperative anti-androgen administration in the simple regression model (p = 0.49).There was also no significant correlation between the change in the cross-sectional area of PPM and neurovascular bundle (NVB) preservation in the simple regression model (p = 0.17).In addition, this correlation was not found regardless of unilateral or bilateral neurovascular bundle (NVB) preservation (p = 0.26 and 0.13, respectively).We performed a correlation analysis between the amount of urinary leakage according to the one-hour pad test after RARP and BMI, neurovascular bundle (NVB) preservation during RARP or prostate volume, which are considered to be factors which may affect urinary continence 11 .In the simple regression model, the amount of urinary leakage 1 or 3 months after RARP positively correlated with BMI (p < 0.01 and < 0.05, respectively).The amount of urinary leakage 1, 6, or 9 months after RARP negatively correlated with NVB preservation during RARP (p < 0.05, respectively).The amount of urinary leakage 6 months after RARP positively correlated with prostate volume (p < 0.05; Table 4).The multiple regression model revealed a correlation between the amount of urinary leakage 1 months after RARP and BMI or NVB preservation (p < 0.01 and p = 0.05, respectively; Table 5).
We performed a correlation analysis between the amount of urinary leakage according to the one-hour pad test after RARP and MRI parameters, such as the cross-sectional area of PPM or RUM diameter (Table 4).In the simple regression model, the amount of urinary leakage 1, 3, 6, 9, or 12 months after RARP negatively correlated with the postoperative PPM cross-sectional area (p < 0.05, < 0.001, < 0.001, < 0.001, and < 0.001, respectively).The amount of urinary leakage 1, 3, 6, 9, or 12 months after RARP positively correlated with the change in the cross-sectional area of PPM by RARP (p < 0.001) and the postoperative RUM diameter (p < 0.001, < 0.001, < 0.0 01, < 0.01, and < 0.001, respectively).Furthermore, positive correlations were observed between the amount of urinary leakage 3, 6, 9, or 12 months after RARP and the preoperative RUM diameter (p < 0.01, < 0.01, < 0.05, and < 0.001, respectively) as well as between the amount of urinary leakage 1 month after RARP and the change in the RUM diameter by RARP (p < 0.05).
The multiple regression model revealed a negative correlation between the amount of urinary leakage 3, 9, or 12 months after RARP and the postoperative cross-sectional area of PPM (p < 0.01; Table 5).The amount of urinary leakage 1, 3, 6, 9, or 12 months after RARP positively correlated with the change in the cross-sectional area of PPM by RARP (p < 0.01, < 0.001, < 0.001, < 0.001, and < 0.001, respectively).These results indicate that a decrease in the cross-sectional area of PPM by RARP induced urinary incontinence after RARP.Representative distribution chart to show the correlation between the amount of urine leakage 12 months after RARP and the change in the cross-sectional area of PPM is shown in Fig. 4. A positive correlation was noted between the www.nature.com/scientificreports/amount of urinary leakage 6 or 12 months after RARP and the preoperative RUM diameter (p < 0.05).Furthermore, the amount of urinary leakage 1 month after RARP positively correlated with the postoperative RUM diameter (p < 0.01).These results indicate that a large RUM diameter, which reflects a large distance between the left and right PPM, is a risk factor for the development of urinary incontinence after RARP.A correlation analysis of the amount of urinary leakage according to the one-hour pad test after RARP and the change in the MU diameter from the static to dynamic phases during the Valsalva maneuver was also performed (Table 6).The simple regression model revealed a negative correlation between the amount of urinary leakage 1, 3, 6, 9, or 12 months after RARP and the change in the MU diameter (p < 0.001).The multiple regression model also showed that the amount of urinary leakage 1, 3, 6, 9, or 12 months after RARP negatively correlated with the change in the MU diameter (p < 0.001).Although the amount of urinary leakage 1, 3, 6, 9 or 12 months after RARP negatively correlated with preoperative membranous urethral length (p < 0.05, p < 0.05, p < 0.05, p < 0.05 and p < 0.01, respectively) in the simple model, this correlation did not find in the multiple regression model (p = 0.78, p = 0.27, p = 0.33, p = 0.07 and p = 0.30, respectively).It did not correlate with postoperative membranous urethral length in the simple regression model, (p = 0.11, 0.13, 0.33, 0.30 and 0.30, respectively).
We also examined the relationship between the change in the MU diameter from the static to dynamic phases during the Valsalva maneuver and the cross-sectional area of PPM or the RUM diameter (Table 7).The simple Change in the cross-sectional area of PPM (cm 2 ) Amount of urinary leakage (g)

Discussion
The present study investigated the relationship between the anatomical pelvic structure around the urethra by MRI and urinary incontinence after RARP, and suggested that a decrease in the postoperative cross-sectional area of PPM after RARP and a large RUM diameter affected the postoperative urethral support system and consequently induced urinary incontinence.The levator ani muscle plays a significant role in maintaining urinary continence and its preservation improves urinary continence [4][5][6][7] .Retzius-sparing RARP, which preserves the levator ani and puboprostatic ligaments, has a higher continence rate than standard RARP [12][13][14][15] .PPM, which has components of the levator ani muscle, is responsible for the quick stop phenomenon of urination in males, and, thus, the weakening of PPM by transection, traction injury, or denervation may affect urinary continence after radical prostatectomy 16 .In other words, preserving the integrity and innervation of PPM is crucial for the maintenance of urinary continence 16 .The right and left PPM course posteriorly from the pubis to flank the urethra, and converge in the midline behind the urethra, anterior to the rectum 16 .RUM occupies a space between the right and left PPM 17 .The left and right PPM edges are consistently located in close proximity in an area immediately lateral to RUM 17 , suggesting that PPM and RUM function as a urethral sling to maintain urinary continence 17,18 (Fig. 5A).This urethral support mechanism by the connection between PPM and RUM is important for urinary continence in women 19,20 .In women, urethral hypermobility and a low urethral closure pressure due to damage to the urethral support mechanism causes stress urinary incontinence 21 .Mid-urethral sling surgery has become the gold standard treatment for stress urinary incontinence in women 22 , and the effectiveness of sling surgery depends on adequate postoperative urethral mobility and the urethral closure pressure 23 .The loss of the prostate may cause instability in the urethral support mechanism in men after RARP, as in women 11 .Therefore, we considered the urethral support mechanism by the connection between PPM and RUM to be important for urinary continence in men after RARP, and that the low ability or weak strength of inherent PPM and RUM and surgical damage to PPM and RUM after RARP may induce urinary incontinence after RARP.We herein focused on morphological changes in PPM and RUM before and after RARP.Sohn et al. examined the anatomy of men preoperatively with MRI, and found that men with a thicker puborectalis muscle at the anorectal angle regained continence earlier after prostatectomy 24 .We demonstrated that the cross-sectional area of PPM decreased by 19% after RARP.Moreover, the decrease in the cross-sectional area of PPM after RARP correlated with postoperative urinary incontinence and induced inadequate urethral compression.These results suggest that surgical interventions during RARP may damage PPM itself and nerve and/or blood supply around PPM, leading to the atrophy of PPM, the loss of its function as a urethral sling (Fig. 5B), and, as a result, urinary incontinence after RARP.Therefore, to avoid postoperative urinary incontinence by loss of PPM muscle volume, it may be important to minimize damage to nerves and blood supply by refraining from using electrocautery around PPM and minimize mechanical damage to PPM itself from surgical manipulation during RARP.However, NVB preservation may not be able to avoid loss of PPM muscle volume.
RUM also may play a significant role as a urethral support system by its connection with left and right PPM to maintain urinary continence.Individual variations have been reported in the development of RUM 17,18 .Soga et al. showed that RUM influenced MU stabilization with the rhabdosphincter, and also that it was one of the important structures forming a urethral sphincteric complex 18 .In the present study, pre-and postoperative RUM diameters positively correlated with urinary incontinence; however, the change in the MU diameter from the static to dynamic phases during the Valsalva maneuver did not correlate with the pre-or postoperative RUM diameter.Therefore, although the longer pre-and postoperative distances between left and right PPM were not directly proven to indicate weaker pre-and postoperative function as a urethral sling in the present study, these patients may originally have had weak urethral support from before RARP, which may easily induce postoperative urinary incontinence (Fig. 5C).Therefore, the preoperative measurement of RUM by MRI may predict urinary incontinence after RARP.
Preoperative pelvic floor muscle training (PFMT) enhances the postoperative function of pelvic muscles, including the levator ani muscle, reduces urinary incontinence after prostatectomy, and improves QOL outcomes related to urinary continence [25][26][27] .PFMT with a strength training protocol to hypertrophy the levator ani muscle may yield higher continence rates 25 .PFMT recuperates strength and lost tone in the sling mechanism after prostatectomy, and potentially enhances the pelvic floor support of vesicourethral anastomosis and the bladder base by strengthening PPM 16 .Additionally, the development of surgical procedures to reinforce the urethral sling system may promote urinary continence after RARP.Vladimir et al. reported that the advanced reconstruction of the vesicourethral support technique using the levator ani muscle attenuated urinary incontinence after RARP 28 .Previous studies demonstrated the effectiveness of urethral sling or bladder neck suspension techniques during RARP for urinary continence [29][30][31] .We showed that bladder neck sling suspension during RARP promoted the early return of urinary continence 29 .Cestari et al. found that a proper sling procedure during RARP allowed for the restoration of the sphincteric apparatus capability to its presurgical status; however, its capability decreased after prostate removal and subsequent ureterovesical anastomosis 30 .The efficacy of the male sling to treat postprostatectomy incontinence has been reported 32 .Our anatomical data on the relationship between pelvic muscles and urinary incontinence after RARP appear to support the effectiveness of PFMT, surgical procedures, such as the urethral sling or bladder neck suspension techniques, and postoperative male sling procedures to maintain urinary continence after RARP.There are several limitations that need to be addressed.First, this was a single-center analysis.Furthermore, postoperative MRI was only performed 9 months after RARP in all cases because the effects of postoperative inflammation in muscles need to be excluded.Second, we conducted pre-and postoperative anatomical analyses of the structure around the urethra using MRI, but not a functional analysis, such as the urethral pressure profile.Third, Valsalva effort is variable and subjective.The degree of dynamic movement is influenced by this effort.Therefore, further detailed studies are warranted.

Conclusions
PPM and RUM play significant roles in the supportive mechanism to maintain urinary continence as a urethral support.Surgical interventions during RARP lead to the atrophy of PPM, the loss of its function as a urethral sling, and, ultimately, urinary incontinence after RARP.The preoperative measurement of PPM by MRI may predict urinary incontinence after RARP.The further development of surgical procedures to prevent or minimize damage to and reinforce this urethral support system may contribute to the mitigation of urinary incontinence after RARP.

Figure 1 .
Figure 1.Anatomical analysis of the structure around the urethra using MRI before and after RARP.(A) The left and right cross-sectional areas of PPM (dotted circles) at the height of RUM on the dorsal side of the urethra were calculated using coronal sections.(B) The tissue between the urethra and rectum was considered to be RUM (double arrow) in the transverse plane, and the left-right diameter was measured.PPM puboperinealis muscle, RUM rectourethral muscle, U urethra, R rectum.

Figure 2 .Figure 3 .
Figure 2. Antero-posterior diameter of the membranous urethra (MU diameter) in static (A) and dynamic phases (B) during the Valsalva maneuver by cine MRI.

Figure 4 .
Figure 4. Distribution chart to show the correlation between the change in the cross-sectional area of PPM (before and 12 months after RARP) and amount of urine leakage.Change in PPM before and 12 months after RARP: preoperative PPM − postoperative PPM.

Figure 5 .
Figure 5. Anatomical structure around the urethra before and after RARP and the mechanism underlying postoperative urinary incontinence.(A) PPM and RUM function as a urethral sling to maintain urinary continence.(B) Surgical interventions during RARP damage and may lead to the atrophy of PPM and the loss of its function as a urethral sling.(C) The longer preoperative distance between left and right PPM indicate looser function as a urethral sling, and these patients may originally have had weak urethral support.PPM puboperinealis muscle, RUM rectourethral muscle, U urethra, PB; pubic body, PPL puboprostatic ligament, DVC dorsal vein complex.

Table 2 .
Amount of urinary leakage according to the 1-h pad test and the rate of urinary continence before and 1, 3, 6, 9, and 12 months after RARP.

Table 3 .
Comparison of the PPM cross-sectional area and left-right diameter of RUM before and after RARP.PPM puboperinealis muscle, RUM rectourethral muscle, RARP robot-assisted radical prostatectomy.

Table 4 .
Comparison between the amount of urinary leakage by the 1-h pad test and the PPM cross-sectional area or left-right diameter of RUM before and after RARP: simple regression analysis.BMI body mass index, NV neurovascular bundle, PPM puboperinealis muscle, RUM rectourethral muscle, RARP robot-assisted radical prostatectomy.Change in PPM before and after RARP: preoperative PPM − postoperative PPM.Change in RUM before and after RARP: preoperative RUM − postoperative RUM.

Table 5 .
Comparison between the amount of urinary leakage by the 1-h pad test and the PPM cross-sectional area or left-right diameter of RUM before and after RARP: multiple regression analysis.BMI body mass index, NV neurovascular bundle, PPM puboperinealis muscle, RUM rectourethral muscle, RARP robot-assisted radical prostatectomy.Change in PPM before and after RARP: preoperative PPM − postoperative PPM.Change in RUM before and after RARP: preoperative RUM − postoperative RUM.

Table 6 .
Correlation between the amount of urinary leakage by the 1-h pad test and pre-or post-operative membranous urethral length or the change in MU diameter on cine MRI parameters after RARP.PPM puboperinealis muscle, RUM rectourethral muscle, MU membranous urethra, MUL membranous urethral length, RARP robot-assisted radical prostatectomy.Change in MU diameter: Changes in the MU diameter from the static to dynamic phases during the Valsalva maneuver by cine MRI.

Table 7 .
Correlation between the change in MU diameter on cine MRI at 9 months after RARP and the PPM cross-sectional area and left-right diameter of RUM before and after RARP.PPM puboperinealis muscle, RUM rectourethral muscle, MU membranous urethra.Change in PPM before and after RARP: preoperative PPM − postoperative PPM.Change in RUM before and after RARP: preoperative RUM − postoperative RUM.Change in MU diameter: Changes in the MU diameter from the static to dynamic phases during the Valsalva maneuver by cine MRI.