Repeated vaginal balloon dilatation for Chronic stress incontinence modeling in Sprague Dawley female rats

Repeated vaginal balloon dilatation for Chronic stress incontinence modeling in Sprague Dawley female rats | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Repeated vaginal balloon dilatation for Chronic stress incontinence modeling in Sprague Dawley female rats Basma Hamed, Mohamed Salem, Esam Mosbah, Ahmed El-Hefnawy, Sherry Khater, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7502745/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Objectives Vaginal distention (VD) serves as a validated model for studying birth-related trauma in rats. This study evaluated the effect of repeated vaginal balloon dilatation for chronic stress incontinence modeling in Sprague Dawley female rats. Methods Thirty mature female Sprague Dawley rats (200–250 g) were obtained from the Medical Experimental Research Center, Mansoura Faculty of Medicine. Rats were divided into three groups: (A) Control (no treatment), (B) Acute VD (single vaginal distension, euthanized after 2 weeks), and (C) Chronic VD (VD repeated at 0,2, and 4 weeks). Urodynamic was assessed by abdominal leak peritoneal pressure (ALPP) and urethral specimens were evaluated using hematoxylin/eosin, and Masson staining. Results The study established a chronic urinary stress incontinence (USI) model using repeated vaginal distension (VD), confirmed by significantly lower abdominal leak point pressure (ALPP) in both acute (0.25 ± 0.15 mmHg) and chronic (0.37 ± 0.13 mmHg) models compared to controls (1.41 ± 0.3 mmHg, P < 0.0001). Histopathological analysis revealed muscle fiber thinning, edema, and fibrosis, with urethral thickness progressively decreasing from controls (402.3 ± 41.54 µM) to acute (197.6 ± 5.318 µM) and chronic (166.6 ± 7.323 µM) models (P < 0.0001). These findings validate the model’s effectiveness in replicating chronic USI-associated structural deterioration. Conclusions The current data showed the validity of repeated vaginal balloon dilatation as a model to study chronic SUC better than the single dilatation model. This approach addresses the shortcomings of the conventional VD model by more accurately reflecting the progressive characteristics of pelvic floor weakening. This may enhance the accuracy of preclinical studies and improve therapeutic outcomes for patients with SUI. Chronic urinary stress incontinence (USI) Vaginal distension (VD) model Abdominal leak point pressure (ALPP) Urethral sphincter dysfunction Urethral thickness Animal model Figures Figure 1 Figure 2 Figure 3 Introduction When the detrusor muscle fails to contract, stress urinary incontinence (SUI) causes urine to leak. Incontinence is a serious health, social, and economic consequence caused by urethral sphincter and pelvic floor weakness [ 1 ]. Stress urinary incontinence (SUI) affects many women worldwide. SUI affects 26–49% of women, depending on study methodology and demographics. For instance, 59.4% of women receiving urodynamic tests for urine incontinence experienced stress urinary incontinence[ 2 , 3 ]. These numbers demonstrate the prevalence of SUI and the need for focused treatments. Researchers must use animal models to study stress urinary incontinence (SUI) mechanics and create therapies. Rodent models are popular because they're cheap and easy to handle. To assess post-menopausal pelvic floor degradation, ovariectomy, pudendal nerve injury, and vaginal balloon dilation are used [ 4 ]. These methods imitate nerve injury similarly. These models examine leak point pressure (LPP), histological anomalies, and bladder function to understand septic urinary insufficiency's physiological and biomechanical features [ 5 ]. Large animal models like rabbits, pigs, and lambs resemble humans anatomically and physiologically. Thus, these models effectively evaluate spinal cord injury surgical procedures, biomaterials, and regenerative therapies [ 6 , 7 ]. Tissue-engineered scaffolds are tested on rabbits, while reconstructive surgery and mesh investigations use pigs and sheep [ 8 ]. Mesh research uses rabbits [ 9 ]. Although non-human primates are anatomically closest to humans, ethical and budgetary constraints limit the use of non-human primates for complex interventions with translational potential. Stress urine incontinence research in rodents often uses the vaginal dilatation paradigm. It successfully simulates childbirth-related pelvic floor damage, a major risk factor for SUI [ 10 , 11 ]. This type inserts a balloon catheter into the vaginal canal. The balloon catheter is then inflated and held in place for several hours to cause nerve and mechanical damage similar to vaginal birth [ 11 ]. Thus, abdominal leak point pressure (ALPP) decreases, urethral sphincter malfunctions, and pelvic muscles atrophy. These symptoms resemble stress urinary incontinence (SUI) [ 12 ]. By showing inflammation, collagen remodeling, and nerve injury, histological tests can identify pelvic tissue changes after childbirth [ 13 ]. Model focuses on acute stress urine incontinence. This method is inadequate for long-term injury effects since it focuses on acute trauma. Chronic nerve dysfunction, muscle atrophy, fibrosis, and extracellular matrix remodeling deteriorate pelvic floor and sphincter mechanisms with time [ 4 ]. This study used Sprague Dawley female rats exposed to diverse stressors over long periods to produce a validated model for chronic stress urine incontinence (SUI). Materials and methods Animals: Experimental animals 30 mature Sprague Dawley female rats weighting 200–250 g b.wt, each, were used throughout this study, they were obtained from the medical experimental research center (XXXX); they were put in similar optimum housing conditions with free access to food and water. Animals were kept in cages in a room with a controlled temperature of 26 C and on a 12-h light-dark cycle. Experimental design: The rats was attained from the cage and kept in a metal container containing a piece of cotton drenched with 10 ml of halothane the rat left for 30 seconds and then maintained on a mixture of Ketamine®* 5% at a dose of (75 mg/kg) and Xylaject®** 2% at a dose of (10 mg/kg) Anesthetic doses administered with (75 mg/kg) Ketamine HCl and (10 mg/kg) Xylazine HCl respectively when given intraperitoneally. Experimental procedures: The vaginal distension model which simulates birth trauma, where the rats was weighed and anesthetized by intraperitoneal injection in the supine position with the limbs of the rat attached to the table and pulled towards the top of the cage then an 8-fr foley catheter was inserted into the vagina, and the balloons was inflated with 5 ml of sterile saline. The labia major was sutured in a figure–eight pattern with 5 − 0 surgical needles to prevent the ejection of the balloon. The balloon will be positioned to avoid direct contact with the surface of the table and left to hang from the bottom of the cage, the balloon will be removed after 8 hours [ 14 ] The animals were allocated into three main groups: Group A : (Control) The animals received no treatment, Group B :(acute) The animals undergone single vaginal distention sacrificed 2 weeks after dilatation, Group C :(chronic) The rats underwent a VD then the VD was repeated at 0 week 2week.and 4week.The animals were euthanized by intraperitoneal injection of over dose of thiopental sodium at a dose of 800 mg/kg [ 15 ]. After necropsy all carcasses are to be placed in body bags and labeled with the following information: IACUC method used to ensure death, date, and initials of the person disposing of the carcass. After euthenesia, the animals will be sent to the incinerator. Methods of evaluation: A-Urodynamic study by power lab AD Instruments For doing the urodynamic study, AD Instruments PowerLab 4/30 equipment with Chart 7 software was used. A mixture of ketamine and xylazine were used to induce anesthesia according to the approved protocol. A three-way (24G) cannula that was placed into the urethra and coupled to a physiological pressure transducer (Cat # MLT844, ADInstruments, Australia) for measuring intravesical pressure (IVP) in millimeters of mercury was used for saline infusion and pressure monitoring. A polyethylene catheter was inserted into the bladder dome while under anesthesia, and a syringe pump was used to continuously inject sterile saline at a rate of 0.1 ml/min at 37°C in order to determine the maximum bladder capacity. The infusion was maintained until the external urethral meatus showed a discernible saline leak, which was seen as the endpoint signifying the maximum bladder capacity. Abdominal leak point pressure (ALPP) is the minimal intra-abdominal pressure resulting in urine leakage was determined by four trials per animal, and the average was derived. Abdominal pressure was given manually using two fingers to softly but firmly press the lower abdomen of the anesthetized rat in a downward manner toward the bladder. Gradually, the pressure was raised until the external urethral meatus showed signs of urine leakage. To standardized the precise measurements of intravesical pressure (IVP), the system was zeroed prior to the commencement of each trial and the sensors were attached to a known pressure source. b) Histopathological Examination: The whole bladder and urethra were harvested by removing the symphysis pubis thus preserving the entire urethral segment. The specimens were fixed in 10% neutral buffered formalin overnight and embedded in paraffin. Paraffin blocks were cut into 5-µm-thick sections. These were deparaffinized and hydrated with distilled water. Also, two sequential sections were stained with Masson's trichrome, Van Gieson solution, and silver stain to determine the distribution of smooth muscle and extracellular matrix as well as interstitial fibrosis. C-Statistical analysis: One-way ANOVA with post-comparison Tuckey’s test was used to determine the significance of difference (P 0.05) between the control, acute, and chronic model groups data. The software package used for statistical analysis is the IBM SPSS v.21 (SPSS, Chicago, IL, USA). Results Abdominal Leak Point Pressure (ALPP) Figure 1 presents a comparison of ALPP values among three groups: control, acute model, and chronic model. Key observations can be drawn from the statistical data. The distributional trends indicate that the control group consistently exhibits higher ALPP values than both the acute and chronic models ( p < 0.0001). The median ALPP for the control group is 14.1 ± 3.0mmHg (95% confidence intervals of 1.22 to 1.76), which is significantly higher than that of the acute SUI model rats at 2.5 ± 1.5 mmHg (95% confidence intervals of 0.09 to 0.38) and the chronic SUI model rats at 3.7 ± 1.3 mmHg (95% confidence intervals of 0.17 to 0.51). No significant difference was observed between the two models in the estimated ALPP. Histopathological evaluation Cross-sections were stained with hematoxylin-eosin for general assessment and further with Masson’s Trichrome (Bio-Optica, Milano, Italy) to evaluate urethral sphincter thickness and fibrosis. The evaluation of modeling samples revealed regions of thinned, circularly oriented muscle fibers and a blurring of sphincter structure, characterized by widely spaced fibers, particularly evident in the acute USI model. The submucosal area exhibits significant edema and hemorrhage in the acute model rats' urethra, whereas moderate edema is observed in the chronic model samples. Figure 2 . Regarding the Masson-stained slide estimated urethral muscle thickness, data illustrate the effects of vaginal dilation on urethral thickness for model establishment. The control group displayed the greatest urethral thickness, with a mean of 402.3 ± 41.54 µM. In contrast, the acute model group, exhibited a significant reduction ( P < 0.0001) in urethral thickness (mean: 197.6 ± 5.318 µM), indicative of acute structural compromise. The chronic group, subjected to repeated vaginal dilation, showed further thinning ( P < 0.0001) of the urethra, with the lowest mean thickness (166.6 ± 7.323 µM), suggesting progressive structural deterioration over time (Fig. 3 ). Discussion Animal models play a crucial role in enhancing our comprehension of stress urinary incontinence (SUI). These enable researchers to examine the pathophysiology of SUI and evaluate potential treatments within a controlled setting. Models simulating childbirth injuries have been employed to identify the mechanisms contributing to SUI, thereby aiding in the development of prophylactic treatments[ 16 ]. Animal models have played a crucial role in assessing new therapeutic interventions, such as regenerative medicine strategies and biomaterial applications, before their clinical implementation [ 4 ]. It is essential to recognize that although these models offer valuable insights, they cannot fully replicate the multifactorial nature of human SUI. The ongoing refinement and validation of these models are crucial for improving their translational relevance [ 14 ]. The single vaginal distension (VD) model serves as a prevalent animal model for investigating stress urinary incontinence (SUI). Although it has yielded important insights, various limitations impact its translational relevance. The single VD model primarily induces an acute injury to the pelvic floor, failing to fully replicate the chronic and multifactorial characteristics of SUI in humans. Human stress urinary incontinence (SUI) frequently arises from repetitive stressors, including multiple childbirths, aging, and chronic pelvic strain, which contribute to the progressive weakening of the pelvic floor. The single VD model simulates a singular traumatic event, omitting the progressive nature of human SUI pathophysiology [ 16 ]. Research indicates that animals undergoing a single VD procedure frequently demonstrate partial recovery of urethral function over time. Less than half of the rats exhibited stress incontinence four weeks after VD, suggesting notable spontaneous recovery. The transient nature of the model presents challenges in the study of long-term SUI mechanisms and the evaluation of sustained therapeutic interventions [ 17 ]. The single VD model primarily results in transient pelvic floor and nerve injury. Studies demonstrate that vaginal distension leads to reduced blood flow and hypoxia in urogenital tissues, resulting in acute damage. This model may fail to accurately represent the ongoing nerve damage and progressive muscle atrophy seen in chronic SUI patients, as the induced injuries do not persist over time [ 18 ]. Moreover, Human SUI typically entails a persistent weakening of the pelvic floor, while the single VD model results in acute over-distension. The model's design fails to consider the gradual deterioration of pelvic support structures observed in patients, thereby restricting its applicability in examining the progressive aspects of SUI [ 16 ]. Ultimately, the single VD model's acute and self-limiting characteristics may render it an unreliable platform for assessing the long-term efficacy of therapeutic interventions. Treatments demonstrating potential in this model may not be effectively applicable in clinical environments, where stress urinary incontinence arises from chronic and progressive tissue alterations. This discrepancy highlights the necessity for models that more accurately replicate the chronicity of human SUI to enhance preclinical testing [ 16 ]. Although the single VD model has significantly contributed to our comprehension of SUI, its limitations underscore the need for models that more precisely represent the chronic and multifactorial aspects of the condition in humans. The commonly employed vaginal distension (VD) model simulates birth trauma through acute pelvic floor stretching; however, it may not adequately represent the progressive characteristics of stress urinary incontinence (SUI) observed in humans [ 19 ]. A modified approach has been developed that incorporates repeated vaginal dilatation (RVD) every two weeks for three sessions. This discussion emphasizes the benefits of our model in improving the translational relevance of SUI research. In contrast to the single-event VD model, which mainly causes acute damage, our RVD model more precisely represents the cumulative impacts of multiple childbirths, aging, and chronic pelvic strain—significant risk factors for SUI in humans. The continuous mechanical stress on pelvic floor muscles and connective tissues reflects the gradual deterioration seen in patients over time, thereby enhancing the model's representation of the clinical progression of SUI [ 20 ]. Preliminary findings suggest that the RVD model results in a sustained decrease in leak point pressure (LPP), a recognized functional indicator of SUI severity. This extended dysfunction differs from the temporary impairment typically seen in the conventional VD model, which allows for potential partial recovery. Histological analysis indicates progressive remodeling of the extracellular matrix, and atrophy of muscle fibers, which correspond more closely with the pathophysiology seen in human SUI patients [ 21 ]. The standard VD model has a significant limitation in that it cannot adequately evaluate the long-term effectiveness of therapeutic interventions because of its acute focus. The RVD model offers a more robust testing platform for regenerative and pharmacological therapies by more accurately simulating the chronic degeneration of pelvic support structures. This model facilitates a more reliable assessment of treatment durability and effectiveness, thereby enhancing clinical translation. Conclusion Incorporating repeated vaginal dilatation at controlled intervals, our model provides a clinically relevant representation of the development of SUI. This approach addresses the shortcomings of the conventional VD model by more accurately reflecting the progressive characteristics of pelvic floor weakening. This may enhance the accuracy of preclinical studies and improve therapeutic outcomes for patients with SUI. Additional validation via urodynamic studies and histopathological analyses strengthens its applicability as an advanced model for the progression of SUI research. Declarations Authors’ contributions Basma Hamed, Mohamed Salem, and Ahmed S. El-Hefnawy performed the experiments and drafted the paper. Sherry Khater contributed to the pathological examination. Mahmoud El Tohamy performed the urodynamic study. Ekramy Elmorsy contributed to Data analysis and interpretation. Mohamed Salem, Esam Mosbah, and Gamal Karrouf analyzed the data and contributed to the final revision and submission. All authors read and approved the final manuscript. Funding Open access funding is provided by The Science, Technology & Innovation Funding Authority (STDF) in cooperation with The Egyptian Knowledge Bank (EKB). This study did not receive any funding. Availability of data and materials All data generated or analyzed during this study are included in this article. Ethics approval and consent to participate The Local Ethical Committee of the Mansoura University Faculty of Veterinary Medicine approved the study design (Ph.D./94). The National Research Center-Egypt Ethics Committee standards for the care of research animals were adhered to. MERC supplied the animals, and all studies were conducted under their supervision to ensure adherence to ethical standards concerning the experimental animals. Consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare no conflict of interest in the current research work. 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International Urogynecology Journal 32:501-552. https://doi.org/10.1007/s00192-020-04622-9 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7502745","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":513770844,"identity":"99291842-d8e8-4201-98db-9bd59bdbf840","order_by":0,"name":"Basma Hamed","email":"","orcid":"","institution":"Mansoura University","correspondingAuthor":false,"prefix":"","firstName":"Basma","middleName":"","lastName":"Hamed","suffix":""},{"id":513770846,"identity":"910ff9c2-2452-44c7-83a3-82b6569e9181","order_by":1,"name":"Mohamed Salem","email":"","orcid":"","institution":"Mansoura University","correspondingAuthor":false,"prefix":"","firstName":"Mohamed","middleName":"","lastName":"Salem","suffix":""},{"id":513770848,"identity":"b7c9a409-dbc6-4b80-97fb-e54129e8ac1b","order_by":2,"name":"Esam Mosbah","email":"","orcid":"","institution":"Mansoura University","correspondingAuthor":false,"prefix":"","firstName":"Esam","middleName":"","lastName":"Mosbah","suffix":""},{"id":513770850,"identity":"53d07c68-99f6-46e9-b21e-d974a0c4f5a3","order_by":3,"name":"Ahmed El-Hefnawy","email":"","orcid":"","institution":"Mansoura University","correspondingAuthor":false,"prefix":"","firstName":"Ahmed","middleName":"","lastName":"El-Hefnawy","suffix":""},{"id":513770851,"identity":"9834accd-d40e-4fd9-ab76-197db35bc01f","order_by":4,"name":"Sherry Khater","email":"","orcid":"","institution":"Mansoura University","correspondingAuthor":false,"prefix":"","firstName":"Sherry","middleName":"","lastName":"Khater","suffix":""},{"id":513770852,"identity":"0d273331-1cc9-4ea6-bec7-4a88d9a076e4","order_by":5,"name":"Ekramy Elmorsy","email":"","orcid":"","institution":"Northern Border University","correspondingAuthor":false,"prefix":"","firstName":"Ekramy","middleName":"","lastName":"Elmorsy","suffix":""},{"id":513770853,"identity":"8c5a0371-c27f-4c46-bc4e-3feef3dfeabf","order_by":6,"name":"Abdelaziz M. Hussein","email":"","orcid":"","institution":"Mansoura University","correspondingAuthor":false,"prefix":"","firstName":"Abdelaziz","middleName":"M.","lastName":"Hussein","suffix":""},{"id":513770855,"identity":"68e65fb2-ac35-4372-b03f-df1ee3fa2e29","order_by":7,"name":"Gamal Karrouf","email":"data:image/png;base64,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","orcid":"","institution":"Mansoura University","correspondingAuthor":true,"prefix":"","firstName":"Gamal","middleName":"","lastName":"Karrouf","suffix":""}],"badges":[],"createdAt":"2025-08-31 20:53:06","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7502745/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7502745/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":91372168,"identity":"715665a7-daad-4d59-918b-7fd23d023c89","added_by":"auto","created_at":"2025-09-15 18:55:55","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":689520,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eUrodynamic charts of Abdominal leak point pressure (ALPP) (mmHg) in controls and acute and chronic urinary stress incontinence (SUI) models rats’ groups.\u003c/strong\u003ea) control, b) 2 weeks after injury in after the single dilatation group (acute USI model), c) 6 weeks after injury in the rat tail vein PBS treated chronic SUI model group, d) The bar chart shows a statistical comparison of the estimated urethral thickness of the normal rats, acute urinary stress incontinence (SUI) and chronic SUI models rats’ groups. Data were presented as means and standard deviation (n=6). One way ANOVA and post hoc Tucky’s test were used to evaluate the significance. ***means \u003cem\u003eP\u003c/em\u003e\u0026lt;0.0001\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7502745/v1/cb179b86f491b27520e33e10.png"},{"id":91370933,"identity":"c91429d0-f69e-4059-9160-aa7c29652e78","added_by":"auto","created_at":"2025-09-15 18:47:55","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":331091,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSpecimen of urethra of rats by hematoxylin and eosin (H\u0026amp;E), (100x) for normal rats (A), acute stress urinary incontinence (SUI) (B) and chronic SUI models (C).\u003c/strong\u003e Evaluation of modeling samples showed areas of thinned circularly orientated muscle fibers and blurring of sphincter structure with widely spaced fibers with submucosal edema and hemorrhage especially 2 weeks after the single dilatation group, while submucosal area showed moderate edema in chronic model samples of 2 weeks spaced 3 reaped dilatations.\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7502745/v1/1f2ba69d437b6a7e100560b1.png"},{"id":91372614,"identity":"d2ffb54e-a5bd-4919-a955-dff367e321f5","added_by":"auto","created_at":"2025-09-15 19:03:55","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":628454,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSpecimen of urethra of rats by Masson trichrome, (200x) for normal rats, acute stress urinary incontinence (SUI) and chronic SUI models.\u003c/strong\u003e The submucosal area is free of fibrosis (Masson trichrome)(A) the submucosal area is masked by marked edema in acute model of control urethra (Masson trichrome)(B) the submucosal area showed marked fibrosis in chronic model of control urethra (Masson trichrome)(C)\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7502745/v1/cde215834f3fc270b6f188b8.png"},{"id":94932802,"identity":"efbfeb96-8aea-4ecc-b09d-a70bd5b6f18a","added_by":"auto","created_at":"2025-11-01 21:23:25","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2375663,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7502745/v1/9cda4343-9861-42f8-be35-7337211b2a99.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Repeated vaginal balloon dilatation for Chronic stress incontinence modeling in Sprague Dawley female rats","fulltext":[{"header":"Introduction","content":"\u003cp\u003eWhen the detrusor muscle fails to contract, stress urinary incontinence (SUI) causes urine to leak. Incontinence is a serious health, social, and economic consequence caused by urethral sphincter and pelvic floor weakness [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Stress urinary incontinence (SUI) affects many women worldwide. SUI affects 26\u0026ndash;49% of women, depending on study methodology and demographics. For instance, 59.4% of women receiving urodynamic tests for urine incontinence experienced stress urinary incontinence[\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. These numbers demonstrate the prevalence of SUI and the need for focused treatments.\u003c/p\u003e\u003cp\u003eResearchers must use animal models to study stress urinary incontinence (SUI) mechanics and create therapies. Rodent models are popular because they're cheap and easy to handle. To assess post-menopausal pelvic floor degradation, ovariectomy, pudendal nerve injury, and vaginal balloon dilation are used [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. These methods imitate nerve injury similarly. These models examine leak point pressure (LPP), histological anomalies, and bladder function to understand septic urinary insufficiency's physiological and biomechanical features [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eLarge animal models like rabbits, pigs, and lambs resemble humans anatomically and physiologically. Thus, these models effectively evaluate spinal cord injury surgical procedures, biomaterials, and regenerative therapies [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Tissue-engineered scaffolds are tested on rabbits, while reconstructive surgery and mesh investigations use pigs and sheep [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Mesh research uses rabbits [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Although non-human primates are anatomically closest to humans, ethical and budgetary constraints limit the use of non-human primates for complex interventions with translational potential.\u003c/p\u003e\u003cp\u003eStress urine incontinence research in rodents often uses the vaginal dilatation paradigm. It successfully simulates childbirth-related pelvic floor damage, a major risk factor for SUI [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. This type inserts a balloon catheter into the vaginal canal. The balloon catheter is then inflated and held in place for several hours to cause nerve and mechanical damage similar to vaginal birth [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. Thus, abdominal leak point pressure (ALPP) decreases, urethral sphincter malfunctions, and pelvic muscles atrophy. These symptoms resemble stress urinary incontinence (SUI) [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. By showing inflammation, collagen remodeling, and nerve injury, histological tests can identify pelvic tissue changes after childbirth [\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. Model focuses on acute stress urine incontinence. This method is inadequate for long-term injury effects since it focuses on acute trauma. Chronic nerve dysfunction, muscle atrophy, fibrosis, and extracellular matrix remodeling deteriorate pelvic floor and sphincter mechanisms with time [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. This study used Sprague Dawley female rats exposed to diverse stressors over long periods to produce a validated model for chronic stress urine incontinence (SUI).\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eAnimals:\u003c/h2\u003e\u003cdiv id=\"Sec4\" class=\"Section3\"\u003e\u003ch2\u003eExperimental animals\u003c/h2\u003e\u003cp\u003e30 mature Sprague Dawley female rats weighting 200\u0026ndash;250 g b.wt, each, were used throughout this study, they were obtained from the medical experimental research center (XXXX); they were put in similar optimum housing conditions with free access to food and water. Animals were kept in cages in a room with a controlled temperature of 26 C and on a 12-h light-dark cycle.\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e\n\u003ch3\u003eExperimental design:\u003c/h3\u003e\n\u003cp\u003eThe rats was attained from the cage and kept in a metal container containing a piece of cotton drenched with 10 ml of halothane the rat left for 30 seconds and then maintained on a mixture of Ketamine\u0026reg;* 5% at a dose of (75 mg/kg) and Xylaject\u0026reg;** 2% at a dose of (10 mg/kg) Anesthetic doses administered with (75 mg/kg) Ketamine HCl and (10 mg/kg) Xylazine HCl respectively when given intraperitoneally.\u003c/p\u003e\n\u003ch3\u003eExperimental procedures:\u003c/h3\u003e\n\u003cp\u003eThe vaginal distension model which simulates birth trauma, where the rats was weighed and anesthetized by intraperitoneal injection in the supine position with the limbs of the rat attached to the table and pulled towards the top of the cage then an 8-fr foley catheter was inserted into the vagina, and the balloons was inflated with 5 ml of sterile saline. The labia major was sutured in a figure\u0026ndash;eight pattern with 5\u0026thinsp;\u0026minus;\u0026thinsp;0 surgical needles to prevent the ejection of the balloon. The balloon will be positioned to avoid direct contact with the surface of the table and left to hang from the bottom of the cage, the balloon will be removed after 8 hours [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]\u003c/p\u003e\u003cp\u003e\u003cb\u003eThe animals were allocated into three main groups: Group A\u003c/b\u003e: (Control) The animals received no treatment, \u003cb\u003eGroup B\u003c/b\u003e:(acute) The animals undergone single vaginal distention sacrificed 2 weeks after dilatation, \u003cb\u003eGroup C\u003c/b\u003e:(chronic) The rats underwent a VD then the VD was repeated at 0 week 2week.and 4week.The animals were euthanized by intraperitoneal injection of over dose of thiopental sodium at a dose of 800 mg/kg [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAfter necropsy all carcasses are to be placed in body bags and labeled with the following information: IACUC method used to ensure death, date, and initials of the person disposing of the carcass. After euthenesia, the animals will be sent to the incinerator.\u003c/p\u003e\n\u003ch3\u003eMethods of evaluation:\u003c/h3\u003e\n\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\u003ch2\u003eA-Urodynamic study by power lab AD Instruments\u003c/h2\u003e\u003cp\u003eFor doing the urodynamic study, AD Instruments PowerLab 4/30 equipment with Chart 7 software was used. A mixture of ketamine and xylazine were used to induce anesthesia according to the approved protocol. A three-way (24G) cannula that was placed into the urethra and coupled to a physiological pressure transducer (Cat # MLT844, ADInstruments, Australia) for measuring intravesical pressure (IVP) in millimeters of mercury was used for saline infusion and pressure monitoring. A polyethylene catheter was inserted into the bladder dome while under anesthesia, and a syringe pump was used to continuously inject sterile saline at a rate of 0.1 ml/min at 37\u0026deg;C in order to determine the maximum bladder capacity. The infusion was maintained until the external urethral meatus showed a discernible saline leak, which was seen as the endpoint signifying the maximum bladder capacity. Abdominal leak point pressure (ALPP) is the minimal intra-abdominal pressure resulting in urine leakage was determined by four trials per animal, and the average was derived. Abdominal pressure was given manually using two fingers to softly but firmly press the lower abdomen of the anesthetized rat in a downward manner toward the bladder. Gradually, the pressure was raised until the external urethral meatus showed signs of urine leakage. To standardized the precise measurements of intravesical pressure (IVP), the system was zeroed prior to the commencement of each trial and the sensors were attached to a known pressure source.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eb) Histopathological Examination:\u003c/h3\u003e\n\u003cp\u003eThe whole bladder and urethra were harvested by removing the symphysis pubis thus preserving the entire urethral segment. The specimens were fixed in 10% neutral buffered formalin overnight and embedded in paraffin. Paraffin blocks were cut into 5-\u0026micro;m-thick sections. These were deparaffinized and hydrated with distilled water. Also, two sequential sections were stained with Masson's trichrome, Van Gieson solution, and silver stain to determine the distribution of smooth muscle and extracellular matrix as well as interstitial fibrosis.\u003c/p\u003e\n\u003ch3\u003eC-Statistical analysis:\u003c/h3\u003e\n\u003cp\u003eOne-way ANOVA with post-comparison Tuckey\u0026rsquo;s test was used to determine the significance of difference (P 0.05) between the control, acute, and chronic model groups data. The software package used for statistical analysis is the IBM SPSS v.21 (SPSS, Chicago, IL, USA).\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\u003ch2\u003eAbdominal Leak Point Pressure (ALPP)\u003c/h2\u003e\u003cp\u003eFigure \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e presents a comparison of ALPP values among three groups: control, acute model, and chronic model. Key observations can be drawn from the statistical data. The distributional trends indicate that the control group consistently exhibits higher ALPP values than both the acute and chronic models (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). The median ALPP for the control group is 14.1\u0026thinsp;\u0026plusmn;\u0026thinsp;3.0mmHg (95% confidence intervals of 1.22 to 1.76), which is significantly higher than that of the acute SUI model rats at 2.5\u0026thinsp;\u0026plusmn;\u0026thinsp;1.5 mmHg (95% confidence intervals of 0.09 to 0.38) and the chronic SUI model rats at 3.7\u0026thinsp;\u0026plusmn;\u0026thinsp;1.3 mmHg (95% confidence intervals of 0.17 to 0.51). No significant difference was observed between the two models in the estimated ALPP.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e\u003ch2\u003eHistopathological evaluation\u003c/h2\u003e\u003cp\u003eCross-sections were stained with hematoxylin-eosin for general assessment and further with Masson\u0026rsquo;s Trichrome (Bio-Optica, Milano, Italy) to evaluate urethral sphincter thickness and fibrosis. The evaluation of modeling samples revealed regions of thinned, circularly oriented muscle fibers and a blurring of sphincter structure, characterized by widely spaced fibers, particularly evident in the acute USI model. The submucosal area exhibits significant edema and hemorrhage in the acute model rats' urethra, whereas moderate edema is observed in the chronic model samples. Figure\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eRegarding the Masson-stained slide estimated urethral muscle thickness, data illustrate the effects of vaginal dilation on urethral thickness for model establishment. The control group displayed the greatest urethral thickness, with a mean of 402.3\u0026thinsp;\u0026plusmn;\u0026thinsp;41.54 \u0026micro;M. In contrast, the acute model group, exhibited a significant reduction (\u003cb\u003eP\u003c/b\u003e\u0026thinsp;\u003cb\u003e\u0026lt;\u0026thinsp;0.0001)\u003c/b\u003e in urethral thickness (mean: 197.6\u0026thinsp;\u0026plusmn;\u0026thinsp;5.318 \u0026micro;M), indicative of acute structural compromise. The chronic group, subjected to repeated vaginal dilation, showed further thinning (\u003cb\u003eP\u003c/b\u003e\u0026thinsp;\u003cb\u003e\u0026lt;\u0026thinsp;0.0001)\u003c/b\u003e of the urethra, with the lowest mean thickness (166.6\u0026thinsp;\u0026plusmn;\u0026thinsp;7.323 \u0026micro;M), suggesting progressive structural deterioration over time (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eAnimal models play a crucial role in enhancing our comprehension of stress urinary incontinence (SUI). These enable researchers to examine the pathophysiology of SUI and evaluate potential treatments within a controlled setting. Models simulating childbirth injuries have been employed to identify the mechanisms contributing to SUI, thereby aiding in the development of prophylactic treatments[\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Animal models have played a crucial role in assessing new therapeutic interventions, such as regenerative medicine strategies and biomaterial applications, before their clinical implementation [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. It is essential to recognize that although these models offer valuable insights, they cannot fully replicate the multifactorial nature of human SUI. The ongoing refinement and validation of these models are crucial for improving their translational relevance [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe single vaginal distension (VD) model serves as a prevalent animal model for investigating stress urinary incontinence (SUI). Although it has yielded important insights, various limitations impact its translational relevance. The single VD model primarily induces an acute injury to the pelvic floor, failing to fully replicate the chronic and multifactorial characteristics of SUI in humans. Human stress urinary incontinence (SUI) frequently arises from repetitive stressors, including multiple childbirths, aging, and chronic pelvic strain, which contribute to the progressive weakening of the pelvic floor. The single VD model simulates a singular traumatic event, omitting the progressive nature of human SUI pathophysiology [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eResearch indicates that animals undergoing a single VD procedure frequently demonstrate partial recovery of urethral function over time. Less than half of the rats exhibited stress incontinence four weeks after VD, suggesting notable spontaneous recovery. The transient nature of the model presents challenges in the study of long-term SUI mechanisms and the evaluation of sustained therapeutic interventions [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe single VD model primarily results in transient pelvic floor and nerve injury. Studies demonstrate that vaginal distension leads to reduced blood flow and hypoxia in urogenital tissues, resulting in acute damage. This model may fail to accurately represent the ongoing nerve damage and progressive muscle atrophy seen in chronic SUI patients, as the induced injuries do not persist over time [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eMoreover, Human SUI typically entails a persistent weakening of the pelvic floor, while the single VD model results in acute over-distension. The model's design fails to consider the gradual deterioration of pelvic support structures observed in patients, thereby restricting its applicability in examining the progressive aspects of SUI [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eUltimately, the single VD model's acute and self-limiting characteristics may render it an unreliable platform for assessing the long-term efficacy of therapeutic interventions. Treatments demonstrating potential in this model may not be effectively applicable in clinical environments, where stress urinary incontinence arises from chronic and progressive tissue alterations. This discrepancy highlights the necessity for models that more accurately replicate the chronicity of human SUI to enhance preclinical testing [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAlthough the single VD model has significantly contributed to our comprehension of SUI, its limitations underscore the need for models that more precisely represent the chronic and multifactorial aspects of the condition in humans. The commonly employed vaginal distension (VD) model simulates birth trauma through acute pelvic floor stretching; however, it may not adequately represent the progressive characteristics of stress urinary incontinence (SUI) observed in humans [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eA modified approach has been developed that incorporates repeated vaginal dilatation (RVD) every two weeks for three sessions. This discussion emphasizes the benefits of our model in improving the translational relevance of SUI research. In contrast to the single-event VD model, which mainly causes acute damage, our RVD model more precisely represents the cumulative impacts of multiple childbirths, aging, and chronic pelvic strain\u0026mdash;significant risk factors for SUI in humans. The continuous mechanical stress on pelvic floor muscles and connective tissues reflects the gradual deterioration seen in patients over time, thereby enhancing the model's representation of the clinical progression of SUI [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e].\u003c/p\u003e\u003cp\u003ePreliminary findings suggest that the RVD model results in a sustained decrease in leak point pressure (LPP), a recognized functional indicator of SUI severity. This extended dysfunction differs from the temporary impairment typically seen in the conventional VD model, which allows for potential partial recovery. Histological analysis indicates progressive remodeling of the extracellular matrix, and atrophy of muscle fibers, which correspond more closely with the pathophysiology seen in human SUI patients [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThe standard VD model has a significant limitation in that it cannot adequately evaluate the long-term effectiveness of therapeutic interventions because of its acute focus. The RVD model offers a more robust testing platform for regenerative and pharmacological therapies by more accurately simulating the chronic degeneration of pelvic support structures. This model facilitates a more reliable assessment of treatment durability and effectiveness, thereby enhancing clinical translation.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIncorporating repeated vaginal dilatation at controlled intervals, our model provides a clinically relevant representation of the development of SUI. This approach addresses the shortcomings of the conventional VD model by more accurately reflecting the progressive characteristics of pelvic floor weakening. This may enhance the accuracy of preclinical studies and improve therapeutic outcomes for patients with SUI. Additional validation via urodynamic studies and histopathological analyses strengthens its applicability as an advanced model for the progression of SUI research.\u003c/p\u003e"},{"header":"Declarations","content":"\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026rsquo; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBasma Hamed, Mohamed Salem, and Ahmed S. El-Hefnawy performed the experiments and drafted the paper. Sherry Khater contributed to the pathological examination. Mahmoud El Tohamy performed the urodynamic study. Ekramy Elmorsy contributed to Data analysis and interpretation. Mohamed Salem, Esam Mosbah, and Gamal Karrouf analyzed the data and contributed to the final revision and submission. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eOpen access funding is provided by The Science, Technology \u0026amp; Innovation\u0026nbsp;Funding Authority (STDF) in cooperation with The Egyptian Knowledge Bank\u0026nbsp;(EKB). This study did not receive any funding.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll data generated or analyzed during this study are included in this article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe Local Ethical Committee of the Mansoura University Faculty of Veterinary Medicine approved the study design (Ph.D./94). The National Research Center-Egypt Ethics Committee standards for the care of research animals were adhered to. MERC supplied the animals, and all studies were conducted under their supervision to ensure adherence to ethical standards concerning the experimental animals.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no conflict of interest in the current research work.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors thank the Department of Veterinary Surgery, Anesthesiology, and Radiology-Mansoura University staff for their assistance.\u003c/p\u003e\n"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eWu JM (2021) Stress incontinence in women. New England Journal of Medicine 384 (25):2428-2436. DOI: 10.1056/NEJMcp1914037\u003c/li\u003e\n\u003cli\u003eZehra K, Aslan E (2021) The burden and cost in urinary incontinence. The New Journal of Urology 16 (1):79-88. DOI : 10.33719/yud.746448\u003c/li\u003e\n\u003cli\u003eRubilotta E, Balzarro M, D\u0026rsquo;Amico A, Cerruto MA, Bassi S, Bovo C, Iacovelli V, Bianchi D, Artibani W, Finazzi Agr\u0026ograve; E (2019) Pure stress urinary incontinence: analysis of prevalence, estimation of costs, and financial impact. BMC urology 19:1-5. https://doi.org/10.1186/s12894-019-0468-2\u003c/li\u003e\n\u003cli\u003eTan X, Li G, Li C, Kong C, Li H, Wu S (2024) Animal models, treatment options, and biomaterials for female stress urinary incontinence. Frontiers in Bioengineering and Biotechnology 12:1414323. https://doi.org/10.3389/fbioe.2024.1414323\u003c/li\u003e\n\u003cli\u003eShen J-D, Chen S-J, Chen H-Y, Chiu K-Y, Chen Y-H, Chen W-C (2021) Review of animal models to study urinary bladder function. Biology 10 (12):1316. https://doi.org/10.3390/biology10121316\u003c/li\u003e\n\u003cli\u003eAmend B, Harland N, Knoll J, Stenzl A, Aicher WK (2021) Large animal models for investigating cell therapies of stress urinary incontinence. International Journal of Molecular Sciences 22 (11):6092. https://doi.org/10.3390/ijms22116092\u003c/li\u003e\n\u003cli\u003eZambon JP, Williams KJ, Bennington J, Badlani GH (2019) Applicability of regenerative medicine and tissue engineering for the treatment of stress urinary incontinence in female patients. Neurourology and Urodynamics 38:S76-S83. \u003cstrong\u003ehttps://doi.org/10.1002/nau.24033\u003c/strong\u003e\u003c/li\u003e\n\u003cli\u003eJin Y, Yang M, Zhao W, Liu M, Fang W, Wang Y, Gao G, Wang Y, Fu Q (2024) Scaffold-based tissue engineering strategies for urethral repair and reconstruction. Biofabrication 17 (1):012003. \u003cstrong\u003eDOI\u003c/strong\u003e 10.1088/1758-5090/ad8965\u003c/li\u003e\n\u003cli\u003eDias FGF, Almeida SHMd, F\u0026aacute;varo W, Latuf Filho P, Riccetto CL (2021) Can platelet-rich plasma coating improve polypropylene mesh integration? An immunohistochemical analysis in rabbits. International braz j urol 47 (2):287-294. https://doi.org/10.1590/S1677-5538.IBJU.2020.0017 \u003c/li\u003e\n\u003cli\u003eLin YH, Liu G, Daneshgari F (2008) A mouse model of simulated birth trauma induced stress urinary incontinence. Neurourology and Urodynamics: Official Journal of the International Continence Society 27 (4):353-358. \u003cstrong\u003ehttps://doi.org/10.1002/nau.20509\u003c/strong\u003e\u003c/li\u003e\n\u003cli\u003eHuang J, Cheng M, Ding Y, Chen L, Hua K (2013) Modified vaginal dilation rat model for postpartum stress urinary incontinence. Journal of Obstetrics and Gynaecology Research 39 (1):256-263. \u003cstrong\u003ehttps://doi.org/10.1111/j.1447-0756.2012.01959.x\u003c/strong\u003e\u003c/li\u003e\n\u003cli\u003eHall R, Ward K (2022) Basic understanding of urodynamics. Obstetrics, Gynaecology \u0026amp; Reproductive Medicine 32 (6):110-119. https://doi.org/10.1016/j.ogrm.2022.04.003\u003c/li\u003e\n\u003cli\u003eQu Z, Chen B, Yang M, Chen Y, Ming S, Hou W (2023) Comparative study of two different rat models of stress urinary incontinence. International Urogynecology Journal 34 (12):2867-2872. https://doi.org/10.1007/s00192-023-05593-3\u003c/li\u003e\n\u003cli\u003eHijaz A, Daneshgari F, Sievert K-D, Damaser MS (2008) Animal models of female stress urinary incontinence. The Journal of urology 179 (6):2103-2110. https://doi.org/10.1016/j.juro.2008.01.096\u003c/li\u003e\n\u003cli\u003eZatroch KK, Knight CG, Reimer JN, Pang DS (2016) Refinement of intraperitoneal injection of sodium pentobarbital for euthanasia in laboratory rats (Rattus norvegicus). BMC veterinary research 13:1-7. DOI 10.1186/s12917-017-0982-y\u003c/li\u003e\n\u003cli\u003eJiang H-H, Damaser MS (2011) Animal models of stress urinary incontinence. Urinary tract:45-67\u003c/li\u003e\n\u003cli\u003eCannon T, Wojcik E, Ferguson C, Saraga S, Thomas C, Damaser M (2002) Effects of vaginal distension on urethral anatomy and function. BJU international 90 (4):403-407. \u003cstrong\u003ehttps://doi.org/10.1046/j.1464-410X.2002.02918.x\u003c/strong\u003e\u003c/li\u003e\n\u003cli\u003eDamaser MS, Whitbeck C, Chichester P, Levin RM (2005) Effect of vaginal distension on blood flow and hypoxia of urogenital organs of the female rat. Journal of Applied Physiology 98 (5):1884-1890. https://doi.org/10.1152/japplphysiol.01071.2004\u003c/li\u003e\n\u003cli\u003eGill BC, Moore C, Damaser MS (2010) Postpartum stress urinary incontinence: lessons from animal models. Expert review of obstetrics \u0026amp; gynecology 5 (5):567-580. https://doi.org/10.1586/eog.10.48.\u003c/li\u003e\n\u003cli\u003eWang X-x, Zhang L, Lu Y (2023) Advances in the molecular pathogenesis and cell therapy of stress urinary incontinence. Frontiers in cell and Developmental Biology 11:1090386. https://doi.org/10.3389/fcell.2023.1090386.\u003c/li\u003e\n\u003cli\u003eFalah-Hassani K, Reeves J, Shiri R, Hickling D, McLean L (2021) The pathophysiology of stress urinary incontinence: a systematic review and meta-analysis. International Urogynecology Journal 32:501-552. https://doi.org/10.1007/s00192-020-04622-9\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Chronic urinary stress incontinence (USI), Vaginal distension (VD) model, Abdominal leak point pressure (ALPP), Urethral sphincter dysfunction, Urethral thickness, Animal model","lastPublishedDoi":"10.21203/rs.3.rs-7502745/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7502745/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cb\u003eObjectives\u003c/b\u003e\u003c/p\u003e\u003cp\u003eVaginal distention (VD) serves as a validated model for studying birth-related trauma in rats. This study evaluated the effect of repeated vaginal balloon dilatation for chronic stress incontinence modeling in Sprague Dawley female rats.\u003c/p\u003e\u003cp\u003e\u003cb\u003eMethods\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThirty mature female Sprague Dawley rats (200\u0026ndash;250 g) were obtained from the Medical Experimental Research Center, Mansoura Faculty of Medicine. Rats were divided into three groups: (A) Control (no treatment), (B) Acute VD (single vaginal distension, euthanized after 2 weeks), and (C) Chronic VD (VD repeated at 0,2, and 4 weeks). Urodynamic was assessed by abdominal leak peritoneal pressure (ALPP) and urethral specimens were evaluated using hematoxylin/eosin, and Masson staining.\u003c/p\u003e\u003cp\u003e\u003cb\u003eResults\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe study established a chronic urinary stress incontinence (USI) model using repeated vaginal distension (VD), confirmed by significantly lower abdominal leak point pressure (ALPP) in both acute (0.25\u0026thinsp;\u0026plusmn;\u0026thinsp;0.15 mmHg) and chronic (0.37\u0026thinsp;\u0026plusmn;\u0026thinsp;0.13 mmHg) models compared to controls (1.41\u0026thinsp;\u0026plusmn;\u0026thinsp;0.3 mmHg, P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). Histopathological analysis revealed muscle fiber thinning, edema, and fibrosis, with urethral thickness progressively decreasing from controls (402.3\u0026thinsp;\u0026plusmn;\u0026thinsp;41.54 \u0026micro;M) to acute (197.6\u0026thinsp;\u0026plusmn;\u0026thinsp;5.318 \u0026micro;M) and chronic (166.6\u0026thinsp;\u0026plusmn;\u0026thinsp;7.323 \u0026micro;M) models (P\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). These findings validate the model\u0026rsquo;s effectiveness in replicating chronic USI-associated structural deterioration.\u003c/p\u003e\u003cp\u003e\u003cb\u003eConclusions\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe current data showed the validity of repeated vaginal balloon dilatation as a model to study chronic SUC better than the single dilatation model. This approach addresses the shortcomings of the conventional VD model by more accurately reflecting the progressive characteristics of pelvic floor weakening. This may enhance the accuracy of preclinical studies and improve therapeutic outcomes for patients with SUI.\u003c/p\u003e","manuscriptTitle":"Repeated vaginal balloon dilatation for Chronic stress incontinence modeling in Sprague Dawley female rats","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-15 18:47:50","doi":"10.21203/rs.3.rs-7502745/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"57667d97-92a9-4b50-883c-70a56710afed","owner":[],"postedDate":"September 15th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-11-21T11:23:31+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-15 18:47:50","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7502745","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7502745","identity":"rs-7502745","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}