Manabu Okawada

Distraction-induced intestinal enterogenesis: Preservation of intestinal function and lengthening after reimplantation into normal jejunum

Background: Significant bowel lengthening can occur in an isolated intestinal segment with the use of linearly directed distractive forces, resulting in increased surface area and epithelial cell proliferation. We hypothesized that reimplantation of this lengthened intestine into normal jejunum would preserve this gain in intestinal length and function similar to normal jejunum. Methods: An intestinal lengthening device was inserted into isolated jejunal segments in pigs, and fully expanded over 8 days. Lengthened segments were then reimplanted into normal intestinal continuity. Pigs were studied after another 28 days. Function was assessed by motility, mucosal enzyme activity, barrier function, and intestinal ion transport. Results: Lengthened segments were significantly longer than control segments and had nearly 2-fold greater surface area. Bowel lengthening was maintained 4 weeks after reimplantation. Motility after reimplantation was similar to nonoperated pigs. Barrier function, mucosal disaccharidase levels, and electrophysiologic measures declined immediately after lengthening but returned to nearly normal levels 28 days after reimplantation. Conclusion: Bowel lengthening results in a transient decline in mucosal absorptive function and smooth muscle contractility. However, function approaches that of normal bowel after reimplantation into enteric flow. These data may support the use of this technique as a potential new option for the treatment of patients with short bowel syndrome.

Administration of a dipeptidyl peptidase IV inhibitor enhances the intestinal adaptation in a mouse model of short bowel syndrome

BACKGROUND Glucagon-like peptide-2 induces small intestine mucosal epithelial cell proliferation and may have benefit for patients who suffer from short bowel syndrome. However, glucagon-like peptide-2 is inactivated rapidly in vivo by dipeptidyl peptidase IV. Therefore, we hypothesized that selectively inhibiting dipeptidyl peptidase IV would prolong the circulating life of glucagon-like peptide-2 and lead to increased intestinal adaptation after development of short bowel syndrome. METHODS Eight-week old C57BL/6J mice underwent a 50% proximal small bowel resection and were treated with either sitagliptin, a dipeptidyl peptidase IV-inhibitor, starting 1 day before surgery versus placebo. The efficacy of dipeptidyl peptidase IV-inhibitor was assessed 3 days after resection, including intestinal morphology, epithelial cell apoptosis, and epithelial cell proliferation. Adaptive mechanisms were assessed with quantitative real-time polymerase chain reaction, and plasma bioactive glucagon-like peptide-2 was measured by radioimmunoassay. RESULTS Body weight loss and peripheral blood glucose levels did not change compared with short bowel syndrome controls. Dipeptidyl peptidase IV-inhibitor treatment led to significant increases in villus height and crypt depth. Dipeptidyl peptidase IV-inhibitor treatment did not change EC apoptosis rates significantly, but it did increase crypt epithelial cell proliferation significantly versus placebo-short bowel syndrome controls. Dipeptidyl peptidase IV-inhibitor treatment markedly increased messenger RNA expression of β-catenin and c-myc in ileal mucosa. Plasma glucagon-like peptide-2 levels increased significantly (∼ 40.9%) in dipeptidyl peptidase IV-inhibitor short bowel syndrome mice. CONCLUSION Dipeptidyl peptidase IV-inhibitor treatment increased short bowel syndrome adaptation and might potentially be useful for short bowel syndrome patients.

Application of distractive forces to the small intestine: defining safe limits.

BACKGROUND Distraction enterogenesis is a novel method for increasing small bowel length by the application of linearly directed forces. However, the magnitude of distractive forces that human and animal small bowel can safely withstand is unknown. METHODS Acute ex vivo force-displacement curves for human (n = 5) and pig (n = 6) small intestine (with and without mesentery) were made by applying increasing amounts of distractive forces to bowel immersed in normal saline (39°C). Progressive load was applied until gross disruption of the tissue was detected, or the applied force reached 1000 gram-force (gf). Histology was used to detect evidence of load-induced damage. In vivo blood flow to pig bowel with distractive loads (30-200 gf) was measured by laser Doppler. RESULTS The relationship between the level of force and degree of displacement was linear. The presence of a mesentery increased stiffness of pig bowel, but did not affect human bowel. Gross tissue disruption in pig and human tissue was seen at forces between 235 and 295 gf, respectively. However, in grossly undamaged areas, histology was unchanged even after application of higher loads. With in vivo testing, mesenteric blood flow was present up to 200 gf; however, blood flow to the bowel wall was reduced to undetectable levels at loads exceeding 100 gf. CONCLUSIONS While whole bowel tissue may tolerate greater applied loads, blood flow to the bowel wall was compromised at loads over 100 gf, suggesting that any higher forces place the bowel at risk for ischemia. These measurements will help guide the clinical application of distraction enterogenesis.

Stimulation of intestinal growth and function with DPP4 inhibition in a mouse short bowel syndrome model

Glucagon-like peptide-2 (GLP-2) has been shown to be effective in patients with short bowel syndrome (SBS), but it is rapidly inactivated by dipeptidyl peptidase IV (DPP4). We used an orally active DPP4 inhibitor (DPP4-I), MK-0626, to determine the efficacy of this approach to promote adaptation after SBS, determined optimal dosing, and identified further functional actions in a mouse model of SBS. Ten-week-old mice underwent a 50% proximal small bowel resection. Dose optimization was determined over a 3-day post-small bowel resection period. The established optimal dose was given for 7, 30, and 90 days and for 7 days followed by a 23-day washout period. Adaptive response was assessed by morphology, intestinal epithelial cell (IEC) proliferation (proliferating cell nuclear antigen), epithelial barrier function (transepithelial resistance), RT-PCR for intestinal transport proteins and GLP-2 receptor, IGF type 1 receptor, and GLP-2 plasma levels. Glucose-stimulated sodium transport was assessed for intestinal absorptive function. Seven days of DPP4-I treatment facilitated an increase in GLP-2 receptor levels, intestinal growth, and IEC proliferation. Treatment led to differential effects over time, with greater absorptive function at early time points and enhanced proliferation at later time points. Interestingly, adaptation continued in the group treated for 7 days followed by a 23-day washout. DPP4-I enhanced IEC proliferative action up to 90 days postresection, but this action seemed to peak by 30 days, as did GLP-2 plasma levels. Thus DPP4-I treatment may prove to be a viable option for accelerating intestinal adaptation with SBS.

Distraction induced enterogenesis: a unique mouse model using polyethylene glycol.

BACKGROUND Recent studies have demonstrated that the small intestine can be lengthened by applying mechanical forces to the bowel lumen-distraction-induced enterogenesis. However, the mechanisms which account for this growth are unknown, and might be best examined using a mouse model. The purpose of this study is to establish the feasibility of developing distractive-induced small bowel growth in mouse. METHODS Twelve-week old C57BL/6J mice had a jejunal segment taken out of continuity, and distended with polyethylene glycol (PEG: 3350 KDa); this group was compared with a control group without stretching. Segment length and diameter were measured intra-operatively and after 5 d. Villus height, crypt depth, and muscle thickness in the isolated segment were assessed. Rate of epithelial cell proliferation (5-bromo-2-deoxyuridine: BrdU incorporation) in crypts were also examined. The mucosal mRNA expression of targeted factors was performed to investigate potential mechanisms which might lead to distraction-induced enterogenesis. RESULTS At harvest, the PEG-stretched group showed a significant increase in length and diameter versus controls. Villus height, crypt depth, and muscular layer thickness increased in the PEG group. The PEG group also showed significantly increased rates of epithelial cell proliferation versus controls. Real-time PCR showed a trend toward higher β-catenin and c-myc mRNA expression in the PEG-stretched group; however, this difference was not statistically significant. CONCLUSIONS Radial distraction-induced enterogenesis with PEG is a viable method for increasing small intestinal length and diameter. This model may provide a new method for studying the mechanisms leading to distraction-induced enterogenesis.

Use of Enterally Delivered Angiotensin II Type Ia Receptor Antagonists to Reduce the Severity of Colitis

BackgroundRenin-angiotensin system blockade reduces inflammation in several organ systems. Having found a fourfold increase in angiotensin II type Ia receptor expression in a dextran sodium sulfate colitis model, we targeted blockade with angiotensin II type Ia receptor antagonists to prevent colitis development. Because hypotension is a major complication of angiotensin II type Ia receptor antagonists use, we hypothesized that use of angiotensin II type Ia receptor antagonists compounds which lack cell membrane permeability, and thus enteric absorption, would allow for direct enteral delivery at far higher concentrations than would be tolerated systemically, yet retain efficacy.MethodsBased on the structure of the angiotensin II type Ia receptor antagonist losartan, deschloro-losartan was synthesized, which has extremely poor cell membrane permeability. Angiotensin II type Ia receptor antagonist efficacy was evaluated by determining the ability to block NF-κB activation in vitro. Dextran sodium sulfate colitis was induced in mice and angiotensin II type Ia receptor antagonist efficacy delivered transanally was assessed.ResultsIn vitro, deschloro-losartan demonstrated near equal angiotensin II type Ia receptor blockade compared to losartan as well as another angiotensin II type Ia receptor antagonist, candesartan. In the dextran sodium sulfate model, each compound significantly improved clinical and histologic scores and epithelial cell apoptosis. Abundance of TNF-α, IL-1β, and IL6 mRNA were significantly decreased with each compound. In vitro and in vivo intestinal drug absorption, as well as measures of blood pressure and mucosal and colonic blood flow, showed significantly lower uptake of deschloro-losartan compared to losartan and candesartan.ConclusionsThis study demonstrated efficacy of high-dose angiotensin II type Ia receptor antagonists in this colitis model. We postulate that a specially designed angiotensin II type Ia receptor antagonist with poor oral absorption may have great potential as a new therapeutic agent for inflammatory bowel disease in the future.

Repair of hypospadias with severe chordee using a long, wide, U-shaped flap that preserves ventral penile tissues intact for second-stage urethroplasty.

PURPOSE The aim of the study was to report a new technique for repairing hypospadias with severe chordee (HSC). METHODS Our new technique involves making a long, wide, U-shaped incision on the ventral penis from the coronal sulcus to very distal to the meatus and dissecting to create a flap (U-flap). During dissection, the urethra is divided just proximal to the meatus. After release of chordee, the U-flap is returned to the ventral penile shaft and sutured in place. A buttonhole made distally in the U-flap is anastomosed to the cut end of the urethra to create a neomeatus. Snodgrass urethroplasty is performed 6 to 18 months later. We have treated 11 patients with HSC (mean age, 22.3 months) using this technique. RESULTS Postoperatively, all U-flaps were viable. The neomeatus appeared to be more proximal because the penis was straighter. Urethroplasty using the central part of the U-flap was uncomplicated by scar tissue and successful in all cases. After a mean follow-up of 15.7 months, all patients have satisfactory penises without stenosis or diverticulum, although 1 had fistula. CONCLUSION Our U-flap technique allows the ventral penis to be preserved intact without scarring for second-stage urethroplasty and as a result is well suited for treating HSC.

Rectal mucosal dissection commencing directly on the anorectal line versus commencing above the dentate line in laparoscopy-assisted transanal pull-through for Hirschsprung's disease: Prospective medium-term follow-up.

BACKGROUND In 2007, we began using the anorectal line (ARL) as the landmark for commencing rectal mucosal dissection (RMD) instead of the dentate line (DL) during laparoscopy-assisted transanal pull-through (L-TAPT) for Hirschsprungs disease (HD). We conducted a medium-term prospective comparison of postoperative fecal continence (POFC) between DL and ARL cases to follow our short-term study. METHODS POFC is assessed by scoring frequency of motions, severity of staining, severity of perianal erosions, anal shape, requirement for medications, sensation of rectal fullness, and ability to distinguish flatus from stool on a scale of 0 to 2 (maximum: 14). RESULTS Patient demographics were similar for ARL (2007-2014: n=33) and DL (1997-2006: n=41). There were no intraoperative complications and 2 cases of postoperative colitis in both ARL (6.1%) and DL (4.9%). Mean annual medium-term POFC scores for the 4-7 term of this study were consistently better in ARL: 9.7±1.4*, 10.1±1.6*, 10.6±1.6, and 11.3±1.4* in ARL and 8.6±1.5, 9.1±1.6, 9.8±1.9, 10.0±1.6 in DL (*: p<0.05). CONCLUSIONS Medium-term POFC is better when the ARL is used as the landmark for RMD during L-TAPT for HD.

Comparison of outcomes between laparoscopy-assisted and posterior sagittal anorectoplasties for male imperforate anus with recto-bulbar fistula

PURPOSE All reports comparing laparoscopy-assisted anorectoplasty (LAARP) with posterior sagittal anorectoplasty (PSARP) in male high-type imperforate anus include a mix of recto-vesical, recto-prostatic, recto-bulbar, and absent fistula cases without focusing on recto-bulbar fistula (RBF), the most challenging type to treat laparoscopically. We compared LAARP with PSARP for treating only RBF. METHOD We used our fecal continence evaluation questionnaire (FCE; maximum score=10), scoring of magnetic resonance imaging (MRI) findings (MRI scores), and the angle between the rectum and the anal canal (RAA) to assess 20 RBF cases (LAARP=12, PSARP=8) treated from 2000 to 2013 prospectively. RESULTS Mean ages at surgery, MRI scores, mean RAA, and duration of raised C-reactive protein (6.6 vs. 6.7days; p=NS) were similar. In all cases, postoperative MRI showed no residual fistula and normal urination. LAARP had consistently higher FCE (7.9 vs. 7.8 at 3years; 8.6 vs. 8.3 at 5years; 8.9 vs 8.6 at 7years; p=NS, respectively), less wound infections (0 vs. 37.5%; p<0.05), higher incidence of rectal mucosal prolapse (50.0 vs. 0%; p<0.05), and required less analgesia (p<0.05). CONCLUSION Although LAARP and PSARP are comparable for treating RBF, LAARP is associated with less wound infections and higher incidence of rectal mucosal prolapse.

Vaginal anomalies and atresia associated with imperforate anus: Diagnosis and surgical management

BACKGROUND The association of vaginal atresia (or Mayer-Rokitansky-Kuster-Hauser Syndrome) with imperforate anus is rare and can present significant diagnostic and therapeutic challenges. This study describes clinical characteristics, surgical treatment and outcomes in this group of complex children. METHODS Records of 20 patients were retrospectively analyzed from two pediatric surgical centers. RESULTS Five patients were excluded from the long-term analysis due to inadequate information, leaving long-term follow-up in 15 patients. Mean follow-up was 10 years (range 1-31.1 years). The diagnosis of vaginal atresia was made pre-operatively in 12 out of 15 patients, and in three patients it was identified during the anoplasty. The anorectal malformations were rectoperineal (N=2), rectovestibular (N=6), recto-bladder neck (N=1) and imperforate anus without fistula (N=6). Satisfactory surgical repair was performed in 13 patients, while one continues to stool through a low perineal fistula awaiting definitive surgery and another underwent a colostomy and mucous fistula. Delayed vaginal reconstruction was due to a failure to identify the problem prior to anoplasty (N=3). Long-term results demonstrated that anorectal continence was much worse than initially appreciated, and many had associated urinary incontinence. Overall stooling score was far lower than in a separate group of children with imperforate anus without vaginal atresia (Levitt and Peña, 2007). CONCLUSIONS Vaginal atresia with imperforate anus is a rare and an extensive pre-operative workup of females with imperforate anus must include assessment of vagina patency. Vaginal reconstruction and anorectal continuity can be performed in a variety of approaches, but long-term continence is often not optimal. We propose a pathway for management of this difficult genito-anorectal disorder.