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Background: Anal incontinence is a disabling condition that adversely affects the quality of life of a large number of patients, mainly with anal sphincter lesions. In a previous experimental work, in-vitro expanded bone marrow (BM)-derived mesenchymal stem cells (MSC) were demonstrated to enhance sphincter healing after injury and primary repair in a rat preclinical model. In the present article we investigated whether unexpanded BM mononuclear cells (MNC) may also be effective., Methods: Thirty-two rats, divided into groups, underwent sphincterotomy and repair (SR) with primary suture of anal sphincters plus intrasphincteric injection of saline (CTR), or of in-vitro expanded MSC, or of minimally manipulated MNC; moreover, the fourth group underwent sham operation. At day 30, histologic, morphometric, in-vitro contractility, and functional analysis were performed., Results: Treatment with both MSC and MNC improved muscle regeneration and increased contractile function of anal sphincters after SR compared with CTR (p < 0.05). No significant difference was observed between the two BM stem cell types used. GFP-positive cells (MSC and MNC) remained in the proximity of the lesion site up to 30 days post injection., Conclusions: In the present study we demonstrated in a preclinical model that minimally manipulated BM-MNC were as effective as in-vitro expanded MSC for the recovery of anal sphincter injury followed by primary sphincter repair. These results may serve as a basis for improving clinical applications of stem cell therapy in human anal incontinence treatment.

Background: Interstitial cells of Cajal, expressing the proto-oncogene c-kit, have been shown to regulate the spontaneous activity of the gastrointestinal tract. They have been described in the human internal anal sphincter; however, their function is still unclear., Objective: We examined the effects of the c-kit tyrosine kinase inhibitor imatinib mesylate on sphincter strips to investigate the function of the interstitial cells., Design: This was a case series study., Settigs: This was a single-center study conducted at the University of Oxford., Patients: Internal anal sphincter strips were collected from 10 patients undergoing abdominoperineal resection or proctectomy and mounted in organ bath. Responses to electrical field stimulation and chemical agents were monitored in the absence of drugs and after the administration of increasing doses of imatinib mesylate. Immunohistochemistry was performed to identify interstitial cells., Main Outcome Measures: The role of the interstitial cells in the internal anal sphincter was assessed., Results: Imatinib mesylate significantly reduced the tone and the spontaneous activity of the strips. In the absence of drugs, the tone generated was 147.7 ± 33.0 mg/mg of tissue. Administration of ≥5 μM of imatinib mesylate caused a dose-dependent reduction in the tone. Strips exhibited spontaneous activity characterized by intermittent low-amplitude contractions superimposed on basal tone (135.6 ± 4.6 contractions in 10 minutes). Imatinib mesylate significantly reduced the number of contractions at concentration >5 μM. No differences were observed in the responses to electrical field stimulation, carbachol, or phenylephrine. Immunohistochemistry showed c-kit-positive cells., Limitations: This study was limited by the relatively small number of patients enrolled and thus the difficulty of finding human tissue for laboratory studies., Conclusions: Our results suggest that the interstitial cells modulate the tone and the spontaneous activity of the internal anal sphincter. This provides a foundation for new approaches to preclinical and clinical research. Moreover, these cells may represent a target for drugs inhibiting the c-kit receptor and provide a new approach for treating anorectal diseases.

Background: Neoadjuvant chemoradiotherapy is recommended in the treatment of locally advanced rectal cancer. Studies have suggested that chemoradiotherapy adversely affects anorectal function. However, the functional implication and the underlying neuromyogenic changes involved in radiation-induced damage are poorly understood., Objective: This study evaluated the functional changes following chemoradiotherapy on the internal anal sphincter., Design and Patients: This article describes an in vitro study on the internal anal sphincter collected from patients undergoing abdominoperineal resection or proctectomy. Five patients were treated by surgery alone (control group), and 6 received preoperative chemoradiotherapy (treatment group). Sphincter strips were mounted in organ bath, and the responses to electrical field stimulation and drugs were monitored., Settings: The study was performed at the University of Oxford., Main Outcome Measures: The end points of this study were to investigate whether chemoradiotherapy has any significant effects on internal anal sphincter function and, subsequently, to establish the type of injury induced., Results: Chemoradiotherapy strips developed similar tone, but significantly lower spontaneous activity (p = 0.001) than controls. Electrical field stimulation induced relaxation, followed by contraction. At 50 Hz, electrical field stimulation produced 25.6 ± 4.9% (mean ± SE) of maximum relaxation followed by a contraction of 5.5 ± 0.9% of basal tone in chemoradiotherapy strips i9n comparison with 47.0 ± 6.2% (p = 0.009) and 17.7 ± 4.0% (p = 0.007) in controls. Relaxation was significantly attenuated by N-nitro-L-arginine. Significant differences were found in responses to carbachol (p = 0.018) and phenylephrine (p = 0.022), but not to sodium nitroprusside., Limitations: This work was limited by the relatively small number of patients enrolled, because of the difficulty of finding human tissue for laboratory studies, and the lack of long-term results., Conclusions: Chemoradiotherapy significantly impairs internal anal sphincter function and intrinsic nerves seem more susceptible than smooth muscle. The exclusion of anal canal from the radiation field is recommended, when oncologically safe.

Purpose: Sphincter injury is a common cause of anal incontinence. Surgical repair remains the operation of choice; however, the outcome often is poor. We investigated the ability of injected bone marrow-derived mesenchymal stem cells to enhance sphincter healing after injury and primary repair in a preclinical model., Methods: Twenty-four inbred Wistar Furth rats were divided into three groups. As a control, Group A underwent sham operation. Group B had sphincterotomy and repair of both anal sphincters plus saline injections. The study group (Group C) underwent sphincterotomy and repair followed by intrasphincteric injections of syngenic bone marrow-derived mesenchymal stem cells. A further group (Group D) of outbred Wistar rats treated with mesenchymal stem cells and immunosuppressive therapy also was evaluated. At 30 days, histologic and morphometric analysis and in vitro contractility testing was performed., Results: A significant decrease of muscle tissue was observed at the site of repair after sphincter injury. However, in Groups C and D, histologic examination demonstrated new muscle fibers and morphometric analysis revealed a significantly greater muscle area fraction than in Group B (P < 0.05). Moreover, mesenchymal stem cells injection improved contractility of sphincters strips compared with Group B (P < 0.05). No significant differences were found between Groups C and D., Conclusions: In our experimental model, bone marrow-derived mesenchymal stem cells injection improved muscle regeneration and increased contractile function of anal sphincters after injury and repair. Therefore, mesenchymal stem cells may represent an attractive tool for treating anal sphincter lesions in humans. Investigations into the biologic basis of this phenomenon should increase our knowledge on underlying mechanisms involved in sphincter repair.

Background: Selective re-creation of a new internal anal sphincter could be indicated when the natural one is irreversibly damaged or excised., Methods: In this preliminary experimental work, surgical techniques of internal anal sphincter replacement in pigs were investigated. After preoperative anorectal manometry, surgical procedure was done in two phases: abdominal, mobilization of the colon-rectum to the pelvic floor; and perianal, dissection of the anal canal from the external anal sphincter through the intersphincteric space. The fully mobilized anorectal segment, including the internal anal sphincter, was pulled down through the anus and resected. The distal colonic stump was then demucosated and two types of plications of the demucosated segment were accomplished, each type in three animals. The plicated segment was then returned into the anal canal, inside the external sphincter. Short-term follow-up with clinical and manometric evaluations was performed and, subsequently, histological analysis of the plicated segment, after the animals were sacrificed., Results: None of the animals became incontinent. Anal manometry identified a high-pressure zone and relaxation reflex in the new anal canal. Histologic studies showed hypertrophy of smooth muscle layers without degenerative changes., Conclusion: This study indicates that a plication of colonic smooth muscle wall can re-create a high-pressure zone in the anal canal after the internal anal sphincter has been excised.