Fecal continence and the puborectal continence reflex

Transcription

1 Fecal continence and the puborectal continence reflex A study about whether the puborectal continence reflex is regulated by the same nerve pathway as the conscious contraction of the puborectal muscle Jara Jonker s April 9, 2015 Research clerkship report Faculty supervisor: Dr. PMA Broens Department of Surgery, Anorectal Physiology Laboratory, University of Groningen, University Medical Center Groningen

2 Research clerkship Jara Jonker Preface Fecal continence or fecal incontinence is not a subject people talk about. When I tell someone that my research clerkship is about the mechanisms of fecal continence, they often laugh or feel uncomfortable. Fecal (in)continence is a taboo. People tend to avoid the subject and do not understand at first why I think it is an interesting subject for my research clerkship. However, when I enthusiastically tell them about my study, explain how many people suffer from different kinds of fecal incontinence and how badly the incontinence can influence a daily life then they realize how important it is to understand the mechanism of fecal continence, what makes people fecal incontinent and how can we help them. During my research clerkship I had the opportunity to attend several anorectal function test performed in the Anorectal Physiology Laboratory in the UMCG. This was my change of getting to know the patient behind the data and the patient s perspective about fecal incontinence. There was one patient I still remember and I would never forget her. She was 62 years old and had an active lifestyle. She underwent the anorectal function test and she could not hold her stool. Also the test revealed that she was incontinent. She was really shocked and stammered: This is exactly the same as what happened to me when I was on winter sport vacation last year. I just could not hold it anymore, right on the slope, everyone could see me and I felt so extremely embarrassed. That was the moment I realized what kind of impact fecal incontinence has on your life. That was the moment I realized that I would like to do something for that woman. And not only for that woman, but for a much more people who have the same problem turning their daily life to one big embarrassment. During my research clerkship I have expanded my competence in performing research. The literature research, the writing and eventually even the statistics became fun! Therefore, I would like to thank all the members of the research group of the Anorectal Physiology Laboratory. I would like to give special thanks to Dr. PMA Broens and Dr. M Trzpis for their shared research expertise and their valuable guidance in the developmental process of this research clerkship. I have learned a lot from both of you! Jara Jonker April 2015 Picture on title page: 2

3 Fecal continence and the puborectal continence reflex Summary Introduction It has been recently described that the conscious and unconscious contraction of the puborectal muscle is required for fecal continence. The conscious contraction is regulated with involvement of the brain and the unconscious contraction is regulated by the puborectal continence reflex. In this study we investigated whether the puborectal continence reflex is regulated by the same nerve pathway as the conscious contraction of puborectal muscle. Further we aimed to find whether the unconscious and conscious contractions are influenced by each other, age and gender. Methods We included all the 283 patients who underwent anorectal function tests between 2010 and 2014 at the UMCG. In total 189 patients were excluded because of possible generalized innervations damage or operations in the pelvic region after which 94 patients remained. The patients underwent three anorectal function test. First, to investigate sensory condition of the pudendal nerve, they underwent the anal electrosensitivity test. Second, to measure the conscious contraction, they underwent the anorectal pressure test. Finally, to measure the unconscious contraction, they underwent the balloon retention test. Results We found no correlation between the unconscious contraction and the anal electrosensitivity (P: 0.811). In contrast, we found a correlation between the conscious contraction and the anal electrosensitivity (P: 0.012). There was no correlation between the conscious and unconscious contraction (P: 0.634). Age had no influence on the unconscious, neither on the conscious contraction (P: and P: 0.344, respectively). Gender had no influence on the unconscious contraction (P: 0.673). However, gender did have a significantly influence the conscious contraction (P<0.001), since men had a significant stronger conscious contractions. Conclusion We conclude that the nerve pathway responsible for the puborectal continence reflex is different than that one responsible for the conscious contraction of the puborectal muscle. Further, the conscious and unconscious contraction work independently from each other. In addition, age and gender have no influence on the unconscious contraction. Further clinical studies are necessary to evaluate the exact nerve pathway responsible for the puborectal continence reflex. 3

4 Research clerkship Jara Jonker Table of contents PREFACE... 2 SUMMARY... 3 TABLE OF CONTENTS... 4 INTRODUCTION... 5 RESEARCH QUESTION AND HYPOTHESIS MATERIAL AND METHODS RESULTS DISCUSSION CONCLUSIONS DUTCH SUMMARY / SAMENVATTING REFERENCES APPENDIX

5 Fecal continence and the puborectal continence reflex Introduction Fecal incontinence Definition and consequences Fecal incontinence is the involuntary loss of flatus, liquid or solid stool during a period of at least three months. (1-4) It is one of the most devastating diseases that significantly impairs the quality of patient s life. (1) People suffering from fecal incontinence feel very embarrassed, which can result in social isolation, anxiety and depression. Moreover, patients are often absent at work because of their fecal incontinence and consequently, they lose social independence and self-esteem. (2,5,6) Besides this emotional burden, individuals can also feel physical discomfort caused by, for instance, local reactions of the perianal skin and urinary tract while having fecal incontinence. (2) Prevalence The overall prevalence of fecal incontinence in the normal population varies between 0.4% and 15% in adults (7-9) and between 1.4% and 4.1% in children older than 4 years. (10-12) However, these values of prevalence are probably underestimated, especially in case of adults, because many patients feel embarrassed to admit and report the fecal incontinence. (9,13) Bharucha et al report that 90% of all women who have problems with incontinence do not report the problem to the physician. Even 50% the women with severe fecal incontinence are reluctant to report this. (7) Several studies indicate a strong association between fecal incontinence and age. (8,9,14,15) Whitehead et al show an augmentation in prevalence of fecal incontinence from 2.6% in the group of years old, up to 15.3% in the group of 70 years old and older. (8) However, among elderly patients living in nursing homes, fecal incontinence can reach up to 50%. (16) Risk factors The risk factors associated with fecal incontinence are increasing age, female gender, physical limitations, poor general health and increased body mass index. (1,6,9) According to Nelson et al and Landefeld et al, urinary incontinence is most frequently associated with fecal incontinence and vice versa. The high interdependence of fecal and urinary incontinence is probably caused by dysfunction of the same mechanism. (2,9) Many diseases are associated with fecal incontinence, for instance, diabetes, stroke, inflammatory bowel disease, inflammatory bowel syndrome, chronic pulmonary obstructive disease, multiple sclerosis, Parkinson s disease, imperforate anus, Hirschsprung s disease and diarrhea. (2,6,9) Various surgical interventions, including episiotomy, sphincterotomy, colectomy, fistulectomy and ureterosigmoidostomy carry the risk of fecal incontinence. Particularly, obstetric trauma was found to be the most frequent cause of fecal incontinence. (2,6,9) 5

6 Research clerkship Jara Jonker Anatomy of the pelvic floor To be able to understand the current knowledge considering the mechanism of fecal continence and incontinence, it is crucial to know the pelvic floor anatomy. Anatomical differences of the male and female pelvic floor are not significant. In this report, only the anatomy of the female pelvic floor will be described. Pelvic skeleton (17,18) At birth, there are five pelvic bones; ilium, ischium, pubis, sacrum and coccyx. The ilium, ischium and pubis fuse at the age of years and in adults they form one pelvic bone. The ilium is the superior bone of the pelvis and supports the lower abdomen. The ischium is the most posterior and inferior bone. It has one major projection, called the ischial spine, which is suitable for palpation in physical examination. The sacrum consists of five sacral vertebrae which are fused together and inferior the coccyx is attached. The coccyx consists of four fused coccygeal vertebrae. The last pelvic bone, the pubis, is the most anterior part of the boney pelvis. The pubis consists of two parts, the superior and the inferior pubic ramus, and when going more medial, these are fused together. In the midline they articulate as the pubic symphysis (figure 1). In the pelvic floor two ligaments are present: the sacrospinous ligament and the sacrotuberous ligament (figure 1). Both have a triangulated shape. One of the pelvic floor muscles, the coccygeus muscle, coincides with the sacrospinous ligament on the superior side. 6

7 Fecal continence and the puborectal continence reflex Figure 1. Pelvic bones and ligaments (F Netter, human anatomy) 7

8 Research clerkship Jara Jonker Muscles of the pelvic floor (19-21) There are two groups of muscles of the pelvic floor. One group forms the sidewalls of the pelvic floor; the piriformis muscle and the obturator internus muscle. The piriformis muscle is part of the posterolateral wall of the pelvis. The obturator internus muscle is part of the anterolateral wall of the pelvis. This muscle is covered with fascia, on which some muscles of the levator ani originate (arcus tendineus levator ani) (figure 2 and 3). Figure 2. Medial view of the female pelvic floor muscles. The puborectal forms a sling around the rectum and vagina and creates the anorectal angulation, which facilitates the fecal continence. (F Netter, human anatomy) 8

9 Fecal continence and the puborectal continence reflex Figure 3. Superior view of the female pelvic floor muscles. (F Netter, human anatomy) The second group, consisting of the coccygeus muscle and the levator ani muscle, forms the pelvic diaphragm. The coccygeus, also called the ischiococcygeus muscle, coincides with the sacrospinous ligament, as mentioned above. The levator ani muscle consists of three muscles, the puborectal, the pubococcygeus and the iliococcygeus (figure 2 and 3). The puborectal muscle is the most outstanding muscle of the levator ani muscle in preserving fecal continence. (22) The puborectal muscle originates from the pubic bone and forms a sling around the vagina and rectum. When the muscle contracts it creates a sharper anorectal angle, which helps to maintain continence. When the muscle relaxes, it increases the anorectal angle, which facilitates the defecation (figure 4). Therefore, people who cannot relax the puborectal muscle properly can suffer from constipation problems because of impaired defecation. (23) On the contrary, malfunctioning of the puborectal muscle can jeopardize the fecal continence. The name pubococcygeus implies that the muscle comes from the pubis and goes to the coccyx. However, this is incorrect, because the pubococcygeus originates from the posterior inferior pubic ramus and inserts on the midline visceral organs. Therefore, this muscle is also called the pubovisceral muscle. In addition, the pubococcygeus can be divided in three different components, the puboperinealis, the pubovaginalis and the puboanalis. (19) 9

10 Research clerkship Jara Jonker The iliococcygeus muscle originates from the arcus tendineus levator ani. The arcus tendineus levator ani is a linear thickening of fascia that forms a line from the ischial spine to the posterior surface of the ipsilateral superior pubic ramus (figure 2 and 3). The levator ani muscle can consciously contract, which causes the mid urethra, distal vagina and rectum pushed against the pubic bone. (17) As described above, the way of attachment of the different levator ani muscles to the pelvic skeleton are different, which determines different functions of those three levator ani muscles in the pelvic floor. (19,20) Consequently, an impaired function of those muscles separately of each other can lead to different kinds of pelvic floor dysfunction. For example, improper function of the puborectal muscle can result in fecal incontinence, while dysfunction of the pubovaginalis can affect the urethral support. (19) Figure 4. Schematic presentation of the rectum and the anorectal angle created by the puborectal muscle. When the muscle contracts it creates a more acute anorectal angle, which helps to maintain continence (A). When the muscle relaxes, it increases the angle, which facilitates the defecation (B). ( Innervation of pelvic floor muscles (21,24-28) There are two major nerves which innervate the muscles of the pelvic floor: the pudendal nerve and the levator ani nerve (figure 5). The pudendal nerve originates from sacral vertebrae S2 to S4 (with the largest contribution of S3) and it innervates the external anal sphincter muscle, perineal skin, perineal muscles and the dorsal nerve of the clitoris. The levator ani nerve originates from sacral vertebrae S3, S4 and/or S5 and it innervates the coccygeal muscle and the three levator ani muscles. The origin of the levator ani nerve may differ between individuals. Barber et al investigated the innervations in female cadavers and found that the levator ani nerve originated in 30% from S4 alone, in 40% from S3 and S4 and in 30% from S4 and S5. (26) They also found that in several cadavers, a separate nerve innervated the puborectal muscle directly. 10

11 Fecal continence and the puborectal continence reflex It has been under debate whether the levator ani muscles are also innervated by the pudendal nerve. Wallner et al found that in 50-60% of the patients the pudendal nerve did innervate the levator ani muscle. (25) Schraffordt et al and Shafik et al also found an additional branch of the pudendal nerve innervating the levator ani muscle. (24,29) On the contrary, Barber et al did not find any branch of the pudendal nerve that would innervate the pubococcygeus muscle, the iliococcygeus muscle nor the puborectal muscle. (26) However, they did find some vasculature branching from the pudendal artery in the levator ani muscles. Therefore, their findings, which did not confirm pudendal nerve innervation in the levator ani muscles, might be a coincidence resulting from variations between the individuals they had investigated. Figure 5. Schematic view of the sacral nerves. (clinical anatomy 6 th edition) 11

12 Research clerkship Jara Jonker Mechanism of fecal continence The classic theory of fecal continence describes that the pelvic floor and anal sphincter muscles help to regulate defecation and maintain continence. The muscles which contribute to this mechanism are the external anal sphincter muscle, the internal anal sphincter muscle and the puborectal muscle. (30,31) The anus is normally closed by the internal anal sphincter, which provides most of the tonic muscle tension. The classic theory of continence explains that besides the involuntary internal anal sphincter squeezing, the voluntary squeezing to close the anus is maintained by the external anal sphincter. Additionally, the anal mucosal folds and the anal vascular cushions support a tight closure of the anus. Finally, contractions of the puborectal muscle maintain the anorectal angulation. Distension of the rectum, caused for example by fecal mass, results in voluntary contraction of the external anal sphincter and the puborectal muscle, which allows maintaining of the continence. (23,31) In addition, Fernandez-Fraga et al showed that the levator ani muscle, and therefore the puborectal muscle, plays the most important function in fecal incontinence. (22) See Figure 6 for the classic theory of fecal continence. Figure 6. Classic theory of fecal continence. (34) Distension of the rectum, caused by increased amount of fecal mass, results in a signal to the brain. The brain sends a signal to the external anal sphincter and orders to contract. This voluntary squeezing of the external anal sphincter results in maintaining the fecal continence. 12

13 Fecal continence and the puborectal continence reflex Mechanism of fecal incontinence There are different theories considering the mechanism of fecal incontinence. A neurogenic theory, proposed by Parks et al, claims that fecal incontinence can result from pudendal nerve damage, which can be caused by trauma (for example during child delivery), repeated troublesome defecation (as consequence rectal bulge with nerve stretching) and neuropathy. (32) A myogenic theory, described by Sultan et al, complements the neurogenic theory, suggesting that fecal incontinence results from mechanical sphincter disruption. However, there are also patients with a sphincter defect who do not suffer from fecal incontinence. (33) It has not been explained why these patients with severe sphincter defects maintain continence, whereas there are some patients with mild sphincter defects who can be severely incontinent. Furthermore, those theories do not explain why some patients are incontinent only during sleep or why some women with obstetric injury can be continent at a young age and become incontinent in their 50 s. (31) In summary, fecal incontinence seems to be influenced by various factors and dysfunctional mechanisms. The current theories, which try to explain the causes underlying fecal incontinence, are still incomplete and controversial. The role of the anal external sphincter continence reflex in fecal continence Recently, Broens et al published an article in which they question the classic theory of fecal continence. (34) They show that fecal continence is regulated by the anal-external sphincter continence reflex (AESCR). This is a spinal reflex, which functions independently of the brain, thus involuntary (figure 7). This is in contrast with the aforementioned classic theory of fecal continence, claiming that the external anal sphincter can only voluntary contract to preserve fecal continence (figure 6). Van Meegdenburg et al (a researcher from our research group of the Anorectal Physiology Laboratory, UMCG) found that the unconscious contraction of the external anal sphincter, mediated by AESCR is regulated by different nerve fibers than the conscious contraction (article submitted). Van Meegdenbrug et al compared conscious and unconscious contraction of the external anal sphincter in patients with and without pudendal neuropathy. They found a significant correlation between the conscious contraction and the amount of pudendal neuropathy. The higher degree of pudendal neuropathy, the weaker was conscious contraction of the external anal sphincter. On the contrary, there was no correlation found between the unconscious contraction, regulated by the AESCR, and pudendal neuropathy. The finding that there are different nerve pathways involved in the mechanism of fecal continence, can explain the different fecal incontinence conditions. Normally, healthy people do not need to think all the time about fecal continence to stay continent, because the AESCR is controlling the fecal continence by the unconscious contractions, until the urge sensation level. When healthy people reach urge sensation, they can consciously contract and in this way they can keep the stool until they find a toilet. In addition, during sleep the AESCR controls the fecal continence. 13

14 Research clerkship Jara Jonker Figure 7. The Anal External Sphincter Continence Reflex. (34) Distension of the anal canal results in activating the anal continence receptor. With a spinal reflex, without involving the brain, a signal is directed to the external anal sphincter to contract. This involuntary squeezing of the external anal sphincter results in maintaining the continence. In some patients the AESCR is damaged and the unconscious contraction does not work properly. There is still a conscious contraction which can control a part of the continence. However, the conscious contraction is prone to fatigue and cannot contract as long as the unconscious contraction. Thus, when these patients feel any rectal sensation, they cannot hold the feces for a long time, only for about 1 minute. Therefore, most of these patients have trained themselves to go to the toilet immediately after feeling any rectal sensation. So they seem to be continent, but they are actually pseudo-incontinent. In addition, during sleep these patients still have a non functioning AESCR, and therefore the unconscious mechanism to control fecal continence is not working. Thus, patients with a non functioning AESCR are incontinent during sleep and pseudo-continent during the day. The role of puborectal continence reflex in fecal continence As mentioned before, the puborectal muscle plays a major role in the mechanism of fecal continence. It forms a sling around the vagina and rectum. When the muscle contracts it creates a sharp anorectal angle, which helps to maintain the continence. When the muscle relaxes, it increases the anorectal angle, which facilitates defecation. The classic theory of the puborectal muscle tells that the puborectal muscle can consciously contract in preserving fecal continence. (23) This is just like the classic theory of the external anal sphincter. The theory of the AESCR, which shows the crucial role of AESCR in fecal continence, explains that besides the conscious contraction of the external anal sphincter, there is also an unconscious contraction. However, the AESCR theory cannot explain why sphincter defects not always result in incontinence. Apparently, there are more regulatory mechanisms, next to 14

15 Fecal continence and the puborectal continence reflex the AESCR, which are responsible for fecal continence. Indeed, our research group from the Anorectal Physiology Laboratory, UMCG, found that there is additionally a secondary fecal continence reflex (Broens et al, article in preparation). This secondary fecal continence reflex, the puborectal continence reflex shows resembles with the earlier described AESCR but involves contractions of a different muscle. While AESCR, the primary fecal continence reflex, regulates the unconscious contractions of external anal sphincter, the puborectal continence reflex leads to unconscious contractions of the puborectal muscle. Furthermore, the puborectal continence reflex is initiated by stretch receptors in contrast to AESCR, which is initiated by contact receptors. In addition, the puborectal continence reflex is only capable of controlling solid stool and not liquid stool. To fully understand the mechanism of fecal incontinence and explain the different fecal incontinence conditions it is necessary to investigate how the puborectal continence reflex is regulated. Summarizing, the puborectal muscle contraction can be regulated consciously and unconsciously. The unconscious contraction of puborectal muscle is regulated by a reflex which is called the puborectal continence reflex. This reflex is only capable of maintaining solid stool continence. In this study, we will investigate whether the puborectal continence reflex is regulated by the same nerve pathway as the conscious contraction of puborectal muscle and whether the contractions are influenced by age and gender. 15

16 Research clerkship Jara Jonker Research question and hypothesis Research questions 1. Is the puborectal continence reflex regulated by the same nerve pathway as the conscious contraction of puborectal muscle? 2. Are the puborectal continence reflex and the conscious contraction of puborectal muscle influenced by the same factors? Thesis: 1. The puborectal continence reflex is regulated by a different nerve pathway than the conscious contraction of the puborectal muscle. 2. The factors age and gender differently influence the puborectal continence reflex and the conscious contraction of puborectal muscle. Antithesis: 1. The puborectal continence reflex is regulated by the same nerve pathway as the conscious contraction of the puborectal muscle. 2. The factors age and gender influence the puborectal continence reflex in the same way as the conscious contraction of puborectal muscle. 16

17 Fecal continence and the puborectal continence reflex Material and methods Patients In this retrospective study all 283 adult patients who underwent anorectal function tests at the University Medical Center Groningen, the Netherlands, between January 2010 and November 2014, were included. All data was obtained from the patient health records. Any kind of innervation damage might affect contractions measured in this study, which would make our results unreliable. Therefore, patients who underwent any form of surgery in the pelvic floor, had any trauma in the pelvic floor, patients who were diagnosed with polyneuropathy, or other reasons which could cause innervation damage were excluded from the study. In total, 189 patients were excluded because of the following reasons: generalized neurologic disorders (e.g.multiple sclerosis, spinal cord injury, spina bifida or polyneuropathy, n=31), anal sphincter rupture during childbirth, episiotomy or sphincterotomy (n=28), surgery for prolapse or perianal fistula (n=27), hysterectomy (n=20), surgery for anorectal malformation or Hirschsprung s disease (n=18), technical measurement defect (n=9), recto-sigmoïd resection (n=6), patient underwent sacral nerve stimulation therapy (n=3), other (e.g. prostatectomy, ileo-anal pouch, sphincter repair, surgery for hemorrhoids, anal or prostate cancer, pelvic floor trauma, radiation injury or mental retardation, n=28) or a combination of the reasons above (n=19). After these exclusions, 94 patients remained in the study. This study has been approved by the Medical Ethical Committee of the University Medical Center Groningen. Measuring equipment The anorectal function tests were performed using solar, gastrointestinal, high-resolution manometry equipment (Medical Measurements Systems, Enschede, the Netherlands), version Three different catheters were used for the measurement. Tests Three different anorectal function tests were performed: the anal electrosensitivity test, anorectal pressure test and balloon retention test. Anal electrosensitivity test This test measures the sensitivity of the anal canal upon electrical stimulation, and therefore informs us about the sensory condition of the pudendal nerve. (35) This unisensor catheter had an outer diameter of 8F and included two circular electrodes of 2 mm; the distance between the two electrodes was 10 mm. The catheter was inserted into the anal canal of the patient who was lying in left lateral position. The generator was set to produce 0.1 ms square wave, at a constant of 5 Hz. Starting proximally, every centimeter of the anal canal was stimulated from 1 ma to 20 ma, with steps of 1 ma. The threshold reported by the patient from the minimum stimulus was recorded. Anorectal function test In this test an unisensor K12981 solid state (Boston type) catheter with an outer diameter of 12F was used. This circumferential catheter measures the pressure at every 8 mm into 6.8 cm 17

18 Research clerkship Jara Jonker of the rectum. While the patient was lying in the left lateral position, the catheter was inserted into the anal canal. To prevent slipping out of the anal canal, the catheter was fixed on the buttocks with adhesive tape. Basal pressures were registered at start of the measurements. Then the patient was asked to squeeze. The maximum anal and puborectal pressure were registered during squeeze. This test reflects the conscious contraction. Balloon retention test For this test we used two catheters: the Unisensor K12981 catheter (as described above in the anorectal function test) and Unisensor K14204 with an outer diameter of 14F. The Unisensor K14204 catheter connects the rectal balloon, inflates it and has two microtip sensors to register the pressure inside the balloon. The solar, gastrointestinal, high resolution manometry automatically corrects the pressure measured for its own resistance pressure, so that only the real pressure in the anal canal is given. During this test the patient sits upright on a commode, to mimic the physiological situation. Basal pressures were registered at start of the measurements. Then, we filled the balloon with water of 37 C (1.0 ml/second) and we recorded the rectal balloon pressure and the volume in the balloon. The patient was asked to hold the balloon as long as possible. Additionally the patient was asked to report first sensation, constant sensation and urge sensation. First sensation is when the patient first feels a sensation in the rectum. Constant sensation is when the patient constantly feels a sensation in the rectum and could go to the toilet. Urge sensation is when the patient feels the urgency to defecate. The test was stopped when the patient reached the maximum tolerable sensation; this is when the filling reached the limit of tolerance of a patient. Additionally, when the patient was unable to retain the balloon until maximum tolerable sensation, the test was stopped and this was recorded as the maximum retainable sensation. So, this balloon retention test was used to determine the rectal filling sensation in patients and the responds of the anal canal to the rectal filling. (36-38) Because this test consists of a rectal balloon filled with water, it mimics the solid stool. With this balloon retention test, a patient can hold the contraction much longer than during voluntary squeezing measured with the anorectal pressure test. The conscious contraction is prone to fatigue, while the contractions measured in this test are not and remain for much longer. Therefore, this test reflects the unconscious contraction, which is regulated by the puborectal continence reflex. Statistical analysis The data was analyzed with SPSS 22.0 for Windows (IBM SPSS Statistics, IBM Corporation, Armonk, NY). We displayed values as number (percentage) or as median (range). According to Razali et al, the Shapiro-Wilk test is the best test to determine whether a variable is normal distributed. (39) Therefore we used the Shapiro-Wilk test and the visual interpretation of the Q-Q plot to determine if a variable is normal distributed. Simple regression analysis was used to determine predictors of the puborectal muscle contraction. According to Royston et al the best level of statistical significance for a single predictor in a simple linear regression model for creating a multiple linear regression model is a probability of < (40) Therefore, we used a separate probability of < 0.15 as significance for the simple linear regression analysis. After this separate selection of the simple linear regression results, all significant parameters were used to create a model with multiple linear regression analysis. The level of statistical significance was set at a probability of <

19 Fecal continence and the puborectal continence reflex Results Patient s characteristics and anorectal parameters In this study 94 patients were included, of which 27 (29%) were male and 67 (71%) female (table 1). The median age of the patients was 57 (range between 18 and 81 years). In these patients we observed a median anal electrosensitivity of 6 ma (range between 2 and 20 ma) at 2 cm of the anal canal. The median basal pressure of the puborectal muscle was 5 mmhg (range 5-50 mmhg) and the median of the maximum conscious contraction, measured during squeeze, was 38 mmhg (range mmhg). The median pressure of the puborectal muscle at start of the unconscious contraction was 25 mmhg (range mmhg) and the median pressure of the maximum unconscious contraction was 115 mmhg (range mmhg). Table 1. Patient s descriptives Patients (n=94) Male 27 (29%) Patient s characteristics Female 67 (71%) Age (years) 57 (18-81) Anal electrosensitivity Anal electrosensitivity at 2 cm (ma) 6 (2-20) Anorectal pressure test Balloon retention test Basal puborectal pressure (mm Hg) 5 (5-50) Maximum puborectal contraction (mm Hg) = maximum conscious contraction 38 (5-165) Pressure puborectal at start (mm Hg) 25 (5-185) Pressure puborectal at MTV/MRV (mm Hg) = maximum unconscious contraction Values are presented as number (percentage) and median (range). MTV = Maximal tolerable volume, MRV = Maximal retainable volume 115 (20-270) Statistical analysis of normal distribution For our study we used linear regression analysis. However, linear regression analysis is only allowed if the dependent variables, in our case the conscious and unconscious contractions of puborectal muscle, are normally distributed. We analyzed the normal distribution using the Shapiro-Wilk test and simultaneously by visual interpretation of the Q-Q-plot. The Shapiro-Wilk test revealed that the maximum unconscious puborectal muscle contraction did not significantly differ from a normal distribution (P: 0.169), which was confirmed by the Q-Q-plot (see appendix figure I). In contrast, the maximum conscious puborectal muscle contraction did significantly differ from a normal distribution (Shapiro-Wilk test: P: <0.001) (see appendix figure II for the Q- Q-plot). However, after logarithmic transformation of the data, the logarithmic value of maximum conscious contraction of the puborectal muscle did not differ significantly from a 19

20 Research clerkship Jara Jonker normal distribution (Shapiro-Wilk test, P: 0.109). Also the Q-Q-plot showed a normal distribution, see appendix figure III. Correlation between anal electrosensitivity and the puborectal continence reflex We investigated whether anal electrosensitivity had a correlation with the puborectal continence reflex, which regulates the unconscious contraction of the puborectal muscle. A scatter-dot of these two variables (figure 8A) shows that different ma-thresholds used to induce the anal electrosensitivity resulted in an unpredictable spectrum of the unconscious contraction of puborectal muscle. In addition, we performed simple linear regression analysis to confirm that there was no correlation. And indeed we found that the anal electrosensitivity and the unconscious puborectal contraction were not correlated (P: 0.811, table 2). Thus, the sensory condition of the pudendal nerve had no influence on the unconscious contraction of the puborectal muscle. Figure 8. The correlation of the unconscious contraction of puborectal muscle and anal electrosensitivity at 2 cm depth (A), and between the conscious contraction of puborectal muscle and anal electrosensitivity at 2 cm depth (B). Anorectal pressure measured at maximum puborectal contraction describe the conscious contraction, whereas contractions measured at maximal tolerable volume or maximal retainable volume describe the unconscious contraction. 20

21 Fecal continence and the puborectal continence reflex Table 2. Simple linear regression of pressure puborectal at MTV/MRV = maximum unconscious contraction. Regression Variable Constant Age (years) Constant B SE B Bèta P Simple regression Anal electrosensitivity at 2 cm (ma) Constant Gender Constant Log (Maximum puborectal contraction) (mm Hg) MTV = Maximal tolerable volume MRV = Maximal retainable volume Correlation between anal electrosensitivity and conscious contraction of the puborectal muscle Further we investigated whether anal electrosensitivity had a correlation with the maximum conscious contraction of the puborectal muscle. A scatter-dot of these two variables (figure 8B) shows that the more anal electrosensitivity was diminished, the weaker the conscious contraction of puborectal muscle. In addition, we performed simple linear regression analysis to investigate whether there is a significant correlation. We found a significant correlation between the anal electrosensitivity and the conscious contraction (P: 0.012, table 3). Thus, we found that patients who need high electro stimulation in the anal canal (malfunction of the sensory condition of the pudendal nerve) cannot consciously contract as strong as patients who respond to lower electro stimuli in anal canal. 21

22 Research clerkship Jara Jonker Table 3. Simple and multiple regression analysis of maximum puborectal contraction = maximum conscious contraction (after logtransformation) Regression Variable B SE B Bèta P Constant Age (years) Constant Simple regression Anal electrosensitivity at 2 cm (ma) Constant Gender <0.001 Constant Pressure puborectal at MTV/MRV (mm Hg) Multiple regression Constant Anal electrosensitivity at 2 cm (ma) Gender <0.001 MTV = Maximal tolerable volume MRV = Maximal retainable volume Unconscious and conscious puborectal muscle contractions work independently of each other To investigate whether the puborectal continence reflex and the conscious contraction work independently, we compared the correlation between the maximum pressure during the MTV/MRV and during squeeze. Figure 9 shows that changes of the anorectal pressure during the maximal sensory filling do not correlate with changes of pressure during squeeze. Also simple linear regression analysis showed no correlation between the two variables (P: 0.634, table 2, 3 and figure 9). This means that there is no functional correlation between the conscious and unconscious contraction of puborectal muscle and consequently, conscious and unconscious contractions work independently of each other. 22

23 Fecal continence and the puborectal continence reflex Figure 9. Conscious puborectal msucle contraction versus unconscious puborectal muscle contraction. MTV = maximal tolerable volume, MRV = maximal retainable volume Influence of age on the puborectal muscle contraction Besides the anal electrosensitivity, we investigated whether other parameters had influence on the puborectal continence reflex, measured in terms of the unconscious contraction, and whether it had influence on the conscious contraction. We investigated whether the parameter age had an influence. We performed simple linear regression analysis between the unconscious contraction and age. We found no significant correlation between the unconscious contraction and age (P: 0.080, table 2). Additionally, we performed simple linear regression between the conscious contraction and the parameter age. We found no significant correlation between the conscious contraction and age (P: 0.344, table 3). Influence of gender on the puborectal muscle contraction We investigated whether the parameter gender had an influence on the puborectal continence reflex, measured in terms of the unconscious contraction, and whether it had influence on the unconscious contraction. We performed simple linear regression analysis between the unconscious contraction and gender. We found no significant correlation between the unconscious contraction and gender (P: 0.673, table 2). Additionally, we performed simple linear regression analysis between the conscious contraction and gender. We did observe a significant correlation between the conscious contraction and gender (P: <0.001, table 3). We found that males have significant higher conscious contraction of the puborectal muscle, compared to females. Multiple linear regression analysis of conscious contraction of the puborectal muscle. Since we found that both factors, the anal electrosensitivity at 2 cm and gender, had a significant correlation (<0.15 set as probability as described in the methods) with the maximum conscious puborectal muscle contraction in simple linear regression analysis, we further performed a multiple linear regression analysis. Both factors together showed a significant correlation with the conscious contraction of the puborectal muscle in multiple regression analysis, which for anal electrosensitivity at 2 cm was P<0.05 and for gender P<0.001 (table 3). 23

24 Research clerkship Jara Jonker Discussion In this study we investigated whether the puborectal continence reflex, which regulates the unconscious contraction of the puborectal muscle, is regulated by the same nerve pathway as the conscious contraction of the puborectal muscle. We also investigated whether the puborectal continence reflex is influenced by the same factors age and gender as the conscious contraction of the puborectal muscle. Difference in the nerve pathway responsible for the unconscious and conscious puborectal muscle contraction To investigate whether the puborectal continence reflex is regulated by the same nerve pathway as the conscious contraction of the puborectal muscle, we first evaluated whether there is a correlation between anal electrosensitivity and the unconscious contraction as well as between anal electrosensitivity and the conscious contraction. Anal electrosensitivity measures the threshold of the stimulus and it gives us information about the sensory condition of the pudendal nerve. The normal values of anal electrosensitivity reported for healthy people are 3 or 4 ma at 1 and 2 cm of the anal canal. People who suffer from de pudendal nerve damage require higher threshold of electrosensitivity to consciously respond to the stimuli. (41-43) We found that anal electrosensitivity had no correlation with the unconscious contraction of the puborectal muscle. In contrast, we observed that anal electrosensitivity did have a correlation with the conscious contraction. So, we found that patients who need high electro stimulation of the anal canal, thus malfunction of the pudendal nerve, had no effect on the on the puborectal continence reflex, which regulates the unconscious contraction of the puborectal muscle. However, this malfunction of the pudendal nerve was significantly correlated with an impaired conscious contraction of the puborectal muscle. Hence, we showed that the puborectal continence reflex is regulated by a different nerve pathway than the conscious contraction of the puborectal muscle. In addition, the fact that there is no statistically significant correlation between the puborectal continence reflex and the conscious contraction of the puborectal muscle proves that these unconscious and conscious contractions work independently of each other. Nerve pathway responsible for the unconscious contraction Although from this study we know that the unconscious contraction, mediated by the puborectal continence reflex, is regulated by a different nerve pathway than the conscious contraction of the puborectal muscle, the question remains which nerve pathway is responsible for the puborectal continence reflex. Based on the anatomical distribution of innervation, which showed great inter-individual diversity as described in the introduction, we postulate three possible nerve pathways responsible for the puborectal continence reflex. Since the levator ani muscle is innervated by the levator ani nerve, there is a possibility that the levator ani nerve can be responsible for the puborectal continence reflex. Second, given that Barber et al found that some people have a separate nerve which innervates the puborectal muscle directly, we propose that this separate nerve might be the element of the puborectal continence reflex. (26) Finally, although we proved that the puborectal continence reflex is not regulated by the same nerve pathway as the conscious contraction, it does not necessarily mean that it is innervated by a different nerve. There is a possibility that the 24

25 Fecal continence and the puborectal continence reflex puborectal continence reflex is regulated by the pudendal nerve after all, just like the conscious contraction. However, then this would be a high branch of the pudendal nerve, so high that damage of the distal pudendal nerve, which regulates the conscious contraction, does not influences this high branch responsible for the unconscious contraction. Therefore, the third possibility of a nerve pathway responsible for the puborectal continence reflex can be a high branch of the pudendal nerve. In summary, we postulate three possible nerve pathways responsible for the puborectal continence reflex: the levator ani nerve; a separate nerve innervating the puborectal muscle directly or a high branch of the pudendal nerve. However, to specifically identify the nerves and nerve pathways responsible for the puborectal continence reflex, clinical studies that involve different nerve blockages procedures are required. Influence of age on puborectal muscle contraction We showed that the age of the patient did not influence the unconscious contraction of the puborectal muscle. Also we found that the age of the patient neither influenced the conscious contraction. This is surprising, because Broens et al found that the maximum anal basal and squeeze pressures, were significant weaker in the older age group. (44) This anal basal squeeze pressures are parameters of the external anal sphincter contraction. It is unclear why this differs between the puborectal muscle as we have shown in our study that both pressures, conscious and unconscious, are not significantly influenced by age. Apparently, the puborectal muscle plays such an important role in the continence processes that its condition is preserved as long as possible and therefore not influenced by age. Nevertheless, further studies are necessary to fully explain this phenomenon. Influence of gender on puborectal muscle contraction Our study showed that gender does not influence the puborectal continence reflex. The fact that the unconscious contraction works similarly in male and female is not really surprising, considering that there are no significant anatomic differences in the pelvic floor between males and females. In contrast, we found that gender does have significant influence on the conscious contraction. Men have significantly stronger conscious contraction than women. Allen et al demonstrated that in 80% of the women after childbirth were having evidence of damage and reinnervation of the pudendal nerve, seen on the electromyography. (45) Since damage of the pudendal nerve has a negative effect on the conscious contraction of the puborectal muscle, it is not surprising that women have weaker conscious contraction of the puborectal muscle than men. Importantly, the fact that gender does not influence the puborectal continence reflex but does influence the conscious contractions again points into the conclusion that the different contractions of puborectal muscle are under control of different nerve pathways. Clinical implications The human organism has two important systems to maintain fecal continence. The external anal sphincter and the puborectal muscle. As showed by Broens et al, additionally to the conscious contraction of the external anal sphincter, there is also an unconscious contraction of the external anal sphincter which is regulated by the AESCR. (34) Van Meegdenburg et al showed that these contraction are regulated by different nerve pathways. (article submitted) As showed by Broens et al, additionally to the conscious contraction of the puborectal muscle, there is also an unconscious contraction of the puborectal muscle, which is regulated 25

26 Research clerkship Jara Jonker by the puborectal continence reflex. (Broens et al, article in preparation) Our study showed that these unconscious and conscious contractions are regulated by different nerve pathways. Apparently, continence plays such an important role in evolution and the human life, that it has a fourfold system for controlling the continence. First the external anal sphincter, which is regulated by two different nerve pathways. Second the puborectal muscle, also regulated by two different nerve pathways. This means that anal sphincter dysfunction alone not necessarily leads to complete incontinence. Because of the fourfold system, the puborectal muscle can preserve continence, when the external anal sphincter cannot. However, as mentioned before, the puborectal continence reflex is only capable of controlling solid stool and not liquids. When the external anal sphincter does not work properly, the puborectal muscle can maintain solid stool continence. Thus, dysfunction of the external anal sphincter does not necessarily leads to complete incontinence; solid stool continence can be maintained. Research implications Our study showed that the puborectal continence reflex is regulated by a different nerve pathway than the conscious contraction of the puborectal muscle. The question remains which nerve is responsible for the puborectal continence reflex. Further clinical studies are required to identify the nerves responsible for the puborectal continence reflex. When the location of the nerve is identified, surgical procedures can pay separate attention to this nerve and therefore not accidently damage it since we now know that it has an important function. Conclusive summary For a clear overview, a conclusive summary of all different factors influencing the unconscious and the conscious contraction of the puborectal muscle and whether they had a correlation is shown in table 4. Table 4. Summary of our findings and their correlations. Unconscious contraction (puborectal continence reflex) Conscious contraction of the puborectal muscle Anal electrosensitivity No correlation Correlation Conscious contraction No correlation n/a Age No correlation No correlation Gender No correlation Correlation 26