Neurourology and Urodynamics 30:126–132 (2011)
Neuromuscular Morphometry of the Uterine Ligaments and Vaginal Wall in Women With Pelvic Organ Prolapse Petek Balkanlı Kaplan,1,∗ Ufuk Usta,2 Hasan Ali Inal,1 Tugba Tastekin,2 and Burcu Tokuc3 1
Department of Obstetric & Gynecology, Trakya University Medical Faculty, Edirne, Turkey 2 Department of Pathology, Trakya University Medical Faculty, Edirne, Turkey 3 Department of Public Health, Trakya University Medical Faculty, Edirne, Turkey
Aims: The aim of this study was to compare neuromuscular histomorphometry of the uterine ligaments and vaginal wall in women with and without pelvic organ prolapse. Methods: Biopsies were obtained from the round, uterosacral, and cardinal ligaments of the uterus and apical vaginal wall of women having pelvic organ prolapse repaired (stage ≥ II; prolapse group, 37) and the same location in patients with no prolapse (stage < II; control group, 47). Routine hematoxylin--eosin (H & E) staining and immunohistochemical staining for Protein Gene Product 9.5 (PGP 9.5) and smooth muscle ␣-actin were performed for all specimens. Results: Smooth muscle percentage of the uterosacral and cardinal ligaments were not significantly different in women with prolapse than in women without. In round ligament, mean smooth muscle percentage was lower than in women with normal support (81.63 ± 8.2 vs. 51.63 ± 16, P = 0.000). Mean distance of the smooth muscle fibers from surface epithelium of the vaginal epithelium of the women with prolapse were significantly higher than the control group (1.679 ± 0.34 vs. 2.240 ± 0.33, P = 0.000). PGP 9.5 stained area percentage of uterine ligaments and vaginal wall tissue samples were significantly lower in women with prolapse. Conclusions: Both total innervation of the anterior vaginal epithelium and uterine ligaments, and muscular percentage of the round ligament and vaginal wall were decreased in women with pelvic organ prolapse. Neurourol. Urodyn. 30:126–132, 2011. © 2010 Wiley-Liss, Inc. Key words: pelvic organ prolapse; PGP 9.5; uterine ligaments; vaginal wall; ␣-actin
INTRODUCTION
The fascial and muscular components within the pelvic floor and its neural pathways create a dynamic support mechanism that facilitates normal pelvic floor functions. The integrity of the vagina and its supportive connective tissues is essential for the maintenance of the pelvic organs in their normal anatomic position. Abnormalities in pelvic floor architecture at cellular or morphological levels lead to mechanical failure of pelvic support. Pelvic organ prolapse and pelvic floor dysfunction may occur if any or all of these tissues are adversely affected. Progressive pelvic floor denervation is thought to lead to sagging of the levators, widening of the levator hiatus with loss of vaginal support leading to genital prolapse. Evaluation of these tissues from a biochemical perspective enables us to better discern the complex interplay between structural composition and supportive capacity. The endopelvic fascia which in connection with the underlying levator ani muscles are mainly supporting the vaginal walls.1,2 Ligaments are sheets of connective tissue, which provide these structures with a high tensile strength. Although previous histochemical as well as electrophysiological investigations have suggested that partial denervation of the pelvic floor occurs both in patients with urinary stress incontinence and fecal incontinence,3,4 our knowledge concerning the neurogen impact on vaginal insufficiency is limited. There are only few studies investigated vaginal wall innervation pattern of women with prolapse. Only one study investigated the innervation process in anterior vaginal wall of women with pelvic organ prolapse showed decreased nerve bundles,5 and two studies investigated the posterior vaginal wall innervation showing decreased6 and increased7 nerve fiber density in the posterior vaginal wall in women with rectocele. To our knowledge, the © 2010 Wiley-Liss, Inc.
nerve fiber density in the uterine ligaments of women with pelvic organ prolapse has not been investigated. The present study was performed to evaluate if uterine ligaments and vaginal wall innervation measured by immunohistochemistry differed in patients with prolapse compared to control subjects. In addition, we evaluated the smooth muscle content in the uterine ligaments and vaginal wall, to identify possible morphologic changes contributing to the pathogenesis of POP.
MATERIALS AND METHODS
The study was approved by the Ethics Committee at Trakya University, Edirne, Turkey. The institutional review board approved the investigation protocol and each patient signed a consent form prior to surgery, allowing the excision of tissue and its use for research purposes after receiving written and oral information. From July 2006 to June 2008, a total of 37 women with pelvic organ prolapse, who underwent vaginal hysterectomies at our hospital, were included in this study. Forty-seven women without uterine prolapse but with other benign gynecological disorders for abdominal hysterectomies were recruited for our control group. Contract grant sponsor: Trakya University Research Council; Contract grant number: TUBAP-800. Conflicts of interest: None. Linda Brubaker led the review process. ∗ Correspondence to: Petek Balkanlı Kaplan, Department of Obstetric & Gynecology, Trakya University Medical Faculty, 22030 Edirne, Turkey. E-mail:
[email protected] Received 13 March 2010; Accepted 7 June 2010 Published online 2 November 2010 in Wiley Online Library (wileyonlinelibrary.com). DOI 10.1002/nau.20972
Morphometry of the Prolapsed Uterine Ligaments Each participant completed a general questionnaire regarding their past medical, obstetrical, gynecological, and pelvic floor disorder history. Included in this questionnaire were focused questions pertaining to their menopausal and hormone status, number and type of delivery. Variables included age, BMI, gravidity and parity, maximal stage of prolapse, date of last menstrual period. A general diagnostic workup and formal neurologic assessment ruled out primary or secondary (due to diabetes mellitus or other endocrine or metabolic abnormalities) central and/or peripheral nervous system diseases in patients and controls. None of the women in the study gave a history of any known connective tissue disease. Patients were excluded if they had pelvic malignancy, fibroids, endometriosis, pelvic inflammatory disease, previous pelvic irradiation, or previous pelvic surgery. None of the patients took any hormonal drugs in the 3 months prior to surgery. None of them used a supportive vaginal pessary or ring prior to surgery. All patients were examined and their prolapse scored according to the International Continence Society Pelvic Organ Prolapse (POP-Q) Classification.8 The study group had POP-Q prolapse stage 2 or more and the control group had a POP-Q prolapse stage 1 or less.
Sampling of Tissues
Full-thickness vaginal specimens from the apical part of the vaginal wall and uterine ligaments (uterosacral, cardinal, round ligaments) were obtained from women having a pelvic organ prolapse repaired (prolapse group, 37) and from the same location in patients with no prolapse who underwent surgery for benign gynecologic conditions (control group, 47). All biopsies were obtained during gynecological operations in general anesthesia. Because the fraction of morphologic components of the vaginal wall may vary throughout the length of the vagina, the site of tissue collection was standardized at the apical portion of the vagina for all subjects. The site of the vaginal specimen was 1 cm from the cervix in the midline anterior portion of the pericervical cuff. Biopsies from the all uterine ligaments were obtained during abdominal or vaginal surgery at the level of their insertions to the uterus. Medial ends of the cardinal ligament were obtained from the part of the cervix above its portio vaginalis.
Preparing and Analyzing Methods of the Tissues
Specimens with dimensions of 0.3--1.1 cm and with dept of 0.2--0.6 cm were fixed in 10% buffered formaldehyde for 6--8 hr and processed in graded alcohols for 16 hr. Five-micrometer thick sections from each patient’s four specimens were obtained from the paraffin embedded tissues and stained with hematoxylin and eosin. Two sections from each specimen, totally 672 biopsy sections were put on poly-L-lysine coated slides for immunohistochemical staining of the tissues for ␣-smooth muscle actin (SMA) (Neomarkers, Freemont, CA) and Protein Gene Product 9.5 (PGP9.5; UC, Isle of Weight, UK). Stained sections were analyzed with under light microscope Nikon Eclipse E600 and Zeiss Axioplan 2 imaging microscope and image analysis program KS300 Software. All tissue specimens and slides were examined independently by two experienced pathologists (UU and ET). The examiners were blinded to the subject’s clinical history. Immunochemical stained sections for PGP 9.5 and smooth muscle actin of each patient’s four tissue samples (vaginal wall and three uterine ligaments) were analyzed with under light Neurourology and Urodynamics DOI: 10.1002/nau
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microscope Nikon Eclipse E600 with a magnification of 50×. Smooth muscle cells were identified by specific staining with antibodies to ␣-actin. Percentage of the SMA stained area was estimated for investigation of uterine ligaments smooth muscle content. In vaginal tissue sections, the distance of the muscle fibers to the surface epithelium of each case were calculated from the nearest point to the surface epithelium via Zeiss Axioplan 2 imaging microscope and image analysis program KS300 Software. Immunohistochemical quantification and mapping of peripheral nerves is a widely used technique when staining tissue samples for innervation and the nerve cytoplasmic protein gene product (PGP 9.5) antibody is commonly used as a neural marker due to its high sensitivity and selectivity.9,10 PGP 9.5 is a tissue-specific ubiquitin carboxyl terminal hydrolase isoenzyme and instead of other neural markers, its antibody stains only peripheral nerves and ganglia in tissues but not adipose tissue. This difference leads to an ease in evaluating the density of the peripheral nerves under light microscope by PGP 9.5 antibody.11 Statistical Analysis
Data analysis was performed using SPSS Version 16.0 computer statistical software package. Analysis of Covariance was used for comparison of the histomorphological parameters of the vaginal wall and uterine ligaments of women with or without prolapse. Pearson correlation analysis was performed for relations between demographics of all women studied and histomorphological parameters of the vaginal wall and uterine ligaments. P < 0.05 was considered statistically significant in all analysis.
RESULTS Demographics
The clinical characteristics of women from whom tissue samples were obtained are listed in Table I. Women with pelvic organ prolapse were significantly older and more parous than women without prolapse. Most of the women with prolapse were in menopausal period of life. Details of the POP-Q values of the all patients are also shown in Table I. Of the 47 women belongs to control group, 18 had stage 0, 29 had stage I pelvic organ prolapse according to POP-Q examination. None of them had more than stage II pelvic organ prolapse. Of the 37 patients with prolapse, 3 had stage II, 15 had stage III, and 19 had stage IV pelvic organ prolapse. Procedures included abdominal total hysterectomy (n = 47), bilateral salpyngoopherectomy (n = 43) and appendectomy (n = 1) for control subjects; and vaginal hysterectomy (n = 34), laparoscopy assisted vaginal hysterectomy (n = 3), McCall culdoplasty (n = 14), high uterosacral ligament suspension (n = 15), salpyngoopherectomy (n = 11), transobturator tape (n = 2), posterior colporrhaphy (n = 28), and anterior colporrhaphy (n = 30). Histomorphology
Hematoxylin--eosin stained sections of apical vaginal wall of both prolapsus group and the control group revealed vascular rich collagenous stroma and beneath dispersed fibers of smooth muscle under the nonkeratonized stratified squamous epithelium. All other specimens contain vascular fat, collagen, and muscle tissue components. Although, there was no prominent
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TABLE I. Clinical and Physical Characteristics of the Women With or Without Pelvic Organ Prolapse
Age (year)a BMI (kg/m2 )a Parityb Number of vaginal delivery per patientb Number of patients perform episiotomy Number of patients deliver via cesarean section Number of patients in menopausal period of life Menopausal period (year)b Anterior compartment stage Stage 0 Stage 1 Stage 2 Stage 3 Stage 4
Control group (stage < 2) (n = 47)
Prolapse group (stage ≥ 2) (n = 37)
P
46.40 ± 5.92 28.31 ± 4.47 2 (0--5) 2 (0--4) 12/47 (26%) 18/47 (38%) 6/47 (13%) 0 (0--25)
59 ± 1.09 26.01 ± 3.97 4 (1--9) 3 (1--9) 21/37 (57%) 2/37 (5%) 26/37 (70%) 6 (0--36)
0.000c 0.016c 0.000d 0.000d 0.000e 0.000e 0.000e 0.000c
18/47 29/47 — — —
— — 3/37 15/37 19/37
— — — — —
Values are mean ± standard deviation. Values are median (min--max). c Student’s t-test. d Mann--Whitney U-test. e Pearson Chi-square test. a
b
difference between the groups, vaginal specimen of the prolapse group showed thickening in the subepithelial collagen fibers. Although there was no statistically significant difference, the smooth muscle fibers of the vaginal wall of the prolapsed group were lesser and smaller than the control group. Immunohistochemistry
All histomorphological analyze results were summarized in Table II. In immunohistochemistry with antibodies to smooth muscle ␣-actin, smooth muscle fibers, perivascular smooth muscle fibers, and myofibroblasts showed positive reaction for SMA more densely. Mean distance of the smooth muscle fibers from surface epithelium of the vaginal epithelium of the women with prolapse were significantly higher than the control group, as our team demonstrated previously12 (Table II). We also demonstrated that percentage of the smooth muscle of the vaginal wall from women with prolapse was decreased (Fig. 1A,C). We showed that smooth muscle percentage of the uterosacral and cardinal ligaments of uterus were not significantly different in women with prolapse than in women without (P = 0.843, P = 0.942, respectively). But in round ligament of uterus, mean
smooth muscle percentage were statistically significant lower than in women with normal support (51.63 ± 16 vs. 81.63 ± 8.2, P = 0.000) (Fig. 1B,D). Immunohistochemical staining for PGP 9.5 revealed a strong cytoplasmic reaction with the peripheral nerve fibers and ganglion cells. Mean ± SD PGP 9.5 stained area percentage of all tissue samples (all three uterine ligaments and vaginal wall) in prolapsed women were significantly lower than the women without prolapse (P = 0.000 for cardinal ligament and vaginal wall, P = 0.045 for uterosacral ligament, P = 0.049 for round ligament, Table II). Neuronization was diminished in all of the uterine ligaments and vaginal wall sample in women with prolapse (Fig. 2). The unique difference in the histomorphometry of the uterosacral and cardinal ligaments of the uterus was significantly diminished PGP 9.5 stained area percentage. Instead of these ligaments, content of the mean smooth muscle and PGP 9.5 stained area percentage in round ligament tissue samples were significantly lower in patients with prolapse than in control subjects (Table II). There was no correlation between round ligament nerve content with age and parity in women with prolapse. In addition, neither number of vaginal delivery nor stage of prolapse showed correlation with the fraction of nerve density in the round
TABLE II. Comparison of the Histomorphological Parameters of the Vaginal Wall and Uterine Ligaments of Women With or Without Prolapse
Vaginal apex Muscular distance from superficial epithelium (mm) PGP 9.5 stained area (%) Uterosacral ligament Smooth muscle (%) PGP 9.5 stained area (%) Round ligament Smooth muscle (%) PGP 9.5 stained area (%) Cardinal ligament Smooth muscle (%) PGP 9.5 stained area (%)
Control group (stage < 2) (n = 47)
Prolapse group (stage ≥ 2) (n = 37)
P
1.679 ± 0.34 14.886 ± 8.03
2.240 ± 0.33 4.303 ± 1.44
0.000 0.000
35.544 ± 17.03 10.351 ± 5.05
34.459 ± 18.9 7.699 ± 2.45
0.843 0.045
81.627 ± 8.2 8.830 ± 5.02
51.625 ± 16.05 6.265 ± 2.6
0.000 0.049
48.882 ± 15.06 15.752 ± 7.2
49.228 ± 17.3 6.795 ± 4.6
0.942 0.000
Statistically significant differences are shown in bold. Parameters presented as mean ± SD, covariates appearing in the model are evaluated at the following values; age = 51.95, BMI = 27.25, parity = 2.92, no. of vaginal delivery = 2.49, postmenopausal period = 4.84.
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Fig. 1. Immunohistochemistry of smooth muscle ␣-actin in the anterior vaginal wall and round ligament in the two study groups (SMA 50×). A,B: Muscular percentage of the tissue samples from vaginal wall and round ligament in women with genital prolapse, respectively. C,D: Same tissue samples from women without prolapse. Note, significant decrease of the vaginal wall and round ligament smooth muscle content in women with prolapse than women with normal vaginal support.
ligament of the uterus (Table III). But, there was a statistically significant negative correlations between the factors that acknowledged as contributing factors for prolapse (age, parity, the numbers of the vaginal deliveries, stage of prolapse) and the neuromuscular density of the sampled tissues of the vaginal wall, uterosacral, and cardinal ligaments from women with pelvic organ prolapse.
DISCUSSION
The major finding of this investigation is that the neuronization of the vaginal wall and uterine ligaments from women with pelvic organ prolapse is significantly diminished compared with the women with normal support. Abnormal histologic features consisted of diminished smooth muscle bundles and decreased in the PGP 9.5-immunoreactive nerve bundles in the round ligament of uterus and apical vaginal wall of women with pelvic organ prolapse. The other abnormal histologic feature of the prolapsed vaginal wall was an increase in the distance of the muscularis layer to the superficial epithelium. Vaginal wall support arise from the connective tissue attachments between the vagina and the pelvic sidewall.13,14 The connective tissue of the vagina and supportive tissues contains a fibrillar component (collagen and elastin), a nonfibrillar component (noncollagenous glycoprotiens, hyaluronan, and proteoglycans) and a significant amount of smooth muscle. Regarding the morphometric determination of the smooth muscle fraction in the pelvic floor, contradicting results can Neurourology and Urodynamics DOI: 10.1002/nau
be found in the literature. Histochemical studies of the pelvic floor muscle biopsies have demonstrated the muscle fiber damage in women with SUI or POP.15 By morphometry of SMA immunostained areas in sections from the anterior vaginal wall, a significant reduction in smooth muscle cell content was shown in women with POP compared to the control group independently of age or race was described.16 Badiou et al.17 confirmed this finding that showing decreased fraction of smooth muscle in the anterior vaginal wall in primary and also recurrent POP cases. Our previous12 and present study confirms these findings showing decreased muscular fraction of vaginal wall. We also demonstrated increased distance of the muscularis layer from the surface epithelium of the anterior vagina in our previous study.12 In this study we also attempted to investigate the neuromuscular changes in all ligaments of the uterus. The uterine ligaments are condensations of endopelvic fascia which provide the primary support, and they are important parts of the pelvic support system.18,19 There are only two studies which investigated the muscular content of the uterosacral ligament in women with prolapse.20,21 The authors observed no difference in the amount of smooth muscle in uterosacral ligament in women with or without pelvic organ prolapse. In the round ligament, a significantly decreased smooth muscle fraction was morphometrically measured in H&E and Masson’s trichrome slides from women with uterine prolapse.22 In parallel with the Ozdegirmenci et al. study,22 we showed that the mean smooth muscle percentage in the round ligament of the women with prolapse was significantly lower than in women with normal support. In addition to their study, we evaluated the muscle
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Fig. 2. Comparative illustration of PGP 9.5 staining nerve densities in the anterior vaginal wall (A1,B1) and the one of the uterine ligaments (A2,B2) of the women with prolapse (A) and without the prolapse (B), respectively (50×). Note, marked decrease of PGP 9.5 immunoreactive nerves in tissue specimens sampled from prolapsed women compared with specimens from control subjects.
content by immunohistochemically by SMA and we also investigated the smooth muscle percentage of the uterosacral and cardinal ligaments of the uterus. We demonstrated that there was no significant difference in smooth muscle content of these two ligaments in women with prolapse than in women without. There is a difference in the effect of POP on the round ligaments when compared to the uterosacral/cardinal ligaments. This might mean that there is a fundamental difference in function of the round ligaments versus the uterosacral and cardinal ligaments on organ support. An other explanation could be that smooth muscle contents are not directly related to prolapse status. Decreased content of smooth muscle in these tissues of women with pelvic organ prolapse may cause of mechanical forces that are imposed on prolapsed tissues or to denervation of them. Nerve damage with resultant pelvic floor muscle weakness is a common suggestion for pelvic organ prolapse.15,23,24 Afflicted nerve endings may also interact with the connective tissues, blood vessels, and pelvic floor muscles.25 So, we decided to investigate the innervation of the uterine ligaments and vaginal tissue of the women with pelvic organ prolapse. In studies which investigated vaginal wall innervation, investigators reported different nerve densities in the vagina. But, probably due to the different methods used, no consistent picture of the innervation was obtained. Zhu et al. study5 is the first survey that investigated the anterior vaginal innervation pattern in control, POP, and SUI patients. The nerve fiber profiles in the SUI/POP groups were lower than those in the control group. This result is supported by the findings of Boreham et al.6 showing decreased nerve bundles and ganglia in the biopsies from the vaginal cuff in patients with posterior vaginal wall Neurourology and Urodynamics DOI: 10.1002/nau
prolapse compared to control subjects. In this study, we showed that nerve density was diminished in apical vaginal wall sample in women with prolapse. Contrary results were demonstrated by Smith et al.4 and Altman et al.7 showing increased nerve fiber density in vaginal wall in patients with POP compared to a control group. The contrary results may be explained by differences in the selection of patients and biopsy sampling, but also by differences in sensitivity of the neuronal markers. Most investigators studied only the connective tissue part20 and the smooth muscle content21,22 of the uterine ligaments. Some studies discovered the presence of nervous and vascular elements within the connective tissue of the uterosacral ligament26,27 and cardinal ligament.28 So, we have attempted to investigate the neuronization of the ligaments of the uterus. To our knowledge, this is the first morphometric study that investigated innervation pattern of the uterine ligaments in women with prolapse. Although there were no difference in smooth muscle percentage in uterosacral and cardinal ligaments, we showed that there was lower innervation in these two ligaments of the uterus in women with prolapse. We showed diminished innervation in all three uterine ligaments of uterus and apical vaginal wall in women with prolapse. Aging is associated with decreased elasticity and worsened innervation of the pelvic floor. Boreham16 reported that women with pelvic organ prolapse were more likely to be white, postmenopausal, and older. Our study subjects were also significantly older, postmenopausal, and more parous than the women with normal vaginal support. This obvious selection bias might have affected our results. We did not use an age-matched control group must be considered critically. Lots of biomechanical stress factors such as obstetric trauma and
r: −0.356, P: 0.001 r: 0.124, P: 0.260 r: −0.281, P: 0.010 r: −0.347, P: 0.001 r: −0.574, P: 0.000 r: −0.205, P: 0.061 r: 0.077, P: 0.485 r: −0.111, P: 0.316 r: −0.089, P: 0.424 r: −0.201, P: 0.068 r: 0.044, P: 0.690 r: −0.154, P: 0.162 r: −0.151, P: 0.174 r: −0.157, P: 0.157 r: 0.016, P: 0.885
CONCLUSIONS
Statistically significant correlations are shown in bold. a Partial correlation analysis was performed. b Kendall’s tau b correlation analysis used.
r: −0.057, P: 0.605 r: −0.016, P: 0.883 r: −0.020, P: 0.859 r: −0.068, P: 0.544 r: −0.081, P: 0.468 r: −0.269, P: 0.013 r: 0.110, P: 0.320 r: −0.230, P: 0.037 r: −0.268, P: 0.014 r: −0.299, P: 0.006 r: 0.412, P: 0.000 r: −0.245, P: 0.025 r: 0.264, P: 0.016 r: 0.373, P: 0.001 r: 0.665, P: 0.000 r: −0.446, P: 0.000 r: 0.231, P: 0.034 r: −0.343, P: 0.002 r: −0.477, P: 0.000 r: −0.631, P: 0.000 Agea BMI (kg/m2 )a Paritea Number of vaginal deliverya Stage of prolapse (POP-Q)b
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increasing body weight can predispose the pelvic floor to dysfunction, these factors are to be acknowledged as contributing factors. Our findings demonstrated that age, parity, the numbers of the vaginal deliveries, stage of prolapse were directly related to change in histomorphometric features of prolapsed tissue. First, the factors that might be influence the organ prolapse were stabilized at the following values: age = 51.95, BMI = 27.25, parity = 2.92, number of vaginal delivery = 2.49, postmenopausal period = 4.84. Afterwards we compared the histomorphometric parameters of the tissues between the women with and without pelvic organ prolapse. Previously, Boreham et al.16 found that the decreased fraction of smooth muscle in the anterior vaginal wall was not related to the severity of prolapse. Instead of this study, we demonstrated that when the prolapse was increasing, innervation of the anterior vaginal wall and uterine ligaments were significantly decreasing. Pelvic neuromuscular dysfunction caused by childbirth has been well documented in neurophysiologic studies, and dysfunction increases with time and by vaginal deliveries.4,29,30 Zhu et al. study5 demonstrated no correlation between nerve fiber profiles and parity in SUI, POP, and control subjects. Contrary to this finding, Ozdegirmenci et al.22 have found a negative correlation between parity and smooth muscle content of the ligament in the study group. Our study showed that neuromuscular changes of the tissues are directly related to the parity, age and number of vaginal deliveries. Lack of correlation of round ligament nerve content with increasing age and parity in women with prolapse suggests that round ligament innervation is not related to age and parity. In addition, neither number of vaginal delivery nor stage of prolapse seemed to be a determinant of the fraction of nerve density in the round ligament of the uterus. The lack of correlation between these contributing factors and innervation of the round ligament may be due to the chronic progressive course of the disease. To our knowledge, this is the first immunohistochemical report that evaluates the difference in nerve density in the cardinal, uterosacral, and round ligaments of uterus in women with utero-vaginal prolapse versus those with no prolapse.
r: −0.350, P: 0.001 r: 0.162, P: 0.141 r: −0.331, P: 0.002 r: −0.382, P: 0.000 r: −0.505, P: 0.000
Smooth muscle content (%) Neuronal content (%) Smooth muscle content (%) Neuronal content (%) Smooth muscle content (%) Neuronal content (%) Muscle distance from epithelium (m) Neuronal content (%)
Uterosacral ligament Vaginal wall
TABLE III. Correlation Between the Demographic Factors and Histomorphometric Parameters
Cardinal ligament
Round ligament
Morphometry of the Prolapsed Uterine Ligaments
Neurourology and Urodynamics DOI: 10.1002/nau
The smooth muscle content of the vaginal wall and round ligament was decreased, nerve density was decreased in all three uterine ligaments and vaginal wall tissue samples investigated in women with pelvic organ prolapse. It is unknown whether changes in the neuromuscular content are a result of stretch and mechanical distention or if decreased amounts of nerve and smooth muscle in the vagina and uterine ligaments contribute to the pathogenesis of prolapse. As it is difficult to find an agematched control group of women with pelvic organ prolapse, further studies with larger numbers of samples from patients with a wider age range may be necessary to fully evaluate the association of neuromuscular fraction with uterine prolapse. It is logical that in vivo or in vitro prolapse models will be needed to test the cause and effect relationship between denervation and smooth muscle morphologic features.
ACKNOWLEDGMENTS
We would like to acknowledge Mr Muzaffer Tudan, the laboratory technician, for his technical support in conducting this experiment.
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Neurourology and Urodynamics DOI: 10.1002/nau
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