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Byline: H. Kobayashi (1), K. Horikoshi (1), A. Yamataka (1), G. J. Lane (1), S. Watanabe (1), S. Futagawa (1), T. Miyano (1) Keywords: Key wordsaBiliary atresia; Bile lake; [alpha]-glutathione-s-transferase (GST); Prognosis Abstract: aEarly identification of patients likely to deteriorate post-hepatic portoenterostomy for biliary atresia (BA) would be beneficial. [alpha]-Glutathione-s-transferase ([alpha]-GST) is a serologic marker of reactive hepatocellular damage because of its low molecular weight, uniform hepatic distribution, high cytosol concentration, and short half-life. We evaluated whether serum [alpha]-GST in post-surgical BA patients correlates with liver function (LF) and investigated its potential as a medium- to long-term marker of prognosis. Postoperative BA patients (n=30 mean age: 11.8+-3.7 years) were divided into three groups based on average LF over the 3 months prior to this study. Group I (n=8) were jaundice-free and had normal LF. Group II (n=12) had moderate liver dysfunction, and group III (n=10) had severe liver dysfunction. Serum [alpha]-GST was determined using a specific ELISA. Tissue [alpha]-GST was determined immunohistochemically, using liver needle-biopsy specimens. Bile lakes were found in 5 group II patients and 5 group III patients. Serum [alpha]-GST was significantly higher in group II (20.7+-8.4ng/ml) than in groups I (4.7+-1.3ng/ml) and III (8.0+-1.2ng/ml) (P&lt 0.0001) and was highest in group II subjects with bile lakes. In control liver specimens [alpha]-GST distribution was weak but uniform throughout normal liver lobule hepatocytes. In group II there was strong staining in centrilobular hepatocytes, and in group III [alpha]-GST was only found in regenerative nodules. We conclude that [alpha]-GST may be a more sensitive indicator of hepatocellular damage in BA because its distribution is correlated to the proportion of functioning liver tissue present. This is the first report of this relationship, which has great implications for group II subjects because a sudden shift in concentration of [alpha]-GST may be a better predictor of impending hepatic dysfunction than conventional LF tests. Author Affiliation: (1) Department of Pediatric Surgery and Second Department of Surgery, Juntendo University School of Medicine, 2-1-1 Hongo, Bunko-ku, Tokyo 113-8421, Japan, JP

Purpose: Several studies have reported the herbal medicine Inchinko-to (ICKT) to have an antifibrotic effect which thus leads to an improvement of hepatic injury. However, there are still few reports of its use in the treatment of cholestatic liver disorder. The aim of this study was to clarify whether the administration of ICKT will ameliorate hepatic fibrosis and injury in cholestatic rats., Materials and Methods: We performed bile duct ligation on 7-week-old male cholestatic Wistar rats and assigned them to one of three groups according to the method of treatment: (1) the SHAM group, (2) the NT-group (non-treatment group), and (3) the T-group (treatment-group). Rats in the T-group were orally administered ICKT (TJ-135) at a dose of 1 g/kg/day and were killed on the 17th postoperative day. We subsequently investigated the levels of fibrosis and various clinical markers through measurement of the following: serum levels of AST and ALT; tissue transforming growth factor-beta 1 (TGF-beta1); tissue inhibitor metalloprotease-1 mRNA (TIMP-1 mRNA) through real-time PCR analysis; and Azan staining and immunohistochemical staining of alfa-smooth muscle actin (alfa-SMA) to evaluate the degree of fibrosis., Results: The levels of serum AST, serum ALT, and TGF-bata1 in the T-Group were significantly lower than those in the NT-Group. In addition, staining of Azan and alfa-SMA in the T-Group was significantly lower than those in the NT-Group., Conclusion: ICKT may help reduce hepatic fibrosis and injury by controlling stellate cell activation.

Purpose: Matrix metalloproteinases (MMPs) and their endogenous tissue inhibitors [tissue inhibitors of metalloproteinases (TIMPs)] have been implicated in tissue injury and remodeling in many organs. The objective of this study was to evaluate the expression of MMP-3 and -9, and TIMP-1, -2, and -3 and their relationship to liver fibrosis in infants with biliary atresia., Methods: The expression of MMP-3 and-9 and TIMP-1, -2 and -3 was investigated in liver tissue samples of nine patients with biliary atresia. In addition, the expression of CCR-4 and CCR-5 was analyzed to investigate the activation of Th1 and Th2 cells. The mRNA levels were measured by semiquantitative reverse transcriptase polymerase chain reaction., Results: The expression of MMP-3 was higher than that of MMP-9 in all samples (P < 0.01). The expression of TIMP-1 was higher than that of TIMP-2 and -3 in all samples (P < 0.01). The expression of CCR-5 was higher than that of CCR-4 (P < 0.05), which implied higher activation of Th1 cells relative to Th2 cells., Conclusion: Our findings suggest that MMP-3, possibly induced by Th1 cytokines, and its balance with TIMP-1, may be one of the factors involved in the pathogenesis of biliary atresia.

Background and Purpose: Many post-operative patients with biliary atresia (BA) suffer from liver dysfunction, such as chronic inflammation even without jaundice after a Kasai's hepatic portoenterostomy., Methods: The presence and degree of oxidative stress were evaluated in the post-operative patients with BA. Twelve outpatients who underwent a Kasai's hepatic portoenterostomy were evaluated. The active oxygen products, the rate of bioantioxidant, the markers of oxidative stress, and the degree of hepatic oxidative stress were examined by immunohistochemical staining of biopsied specimens., Results: All of the oxidative stress markers in the post-operative patients with BA increased in comparison to those in the controls. Moreover, 8-OHdG immunohistochemical staining was positive in 84+/-4.8% in hepatic cells in the portal area in the post-operative patients with BA., Conclusion: The post-operative patients with BA were under increased oxidative stress, even if their liver dysfunction was mild without jaundice. Antioxidant therapy might be necessary to decrease of oxidative stress in the post-operative patients with BA.

Split notochord syndrome (SNS) is an extremely rare anomaly. This report presents the case of a male infant with SNS associated with congenital ileal atresia and a dorsal enteric cyst communicating to the retroperitoneal isolated ceca with a vesical fistula. Dorsal fistulography and vesicography were useful and essential for the detailed study of the topology in this patient. The embryological mechanism and etiologic theories are discussed with a review of 19 cases reported in the literature.

Polysialyated neural cell adhesion molecule (PSA-NCAM) is a marker for immature neurons and S100 beta is a known marker for enteric glia. The aim of this study was to determine the maturation of the enteric nervous system (ENS) in normoganglionic (Ng) bowel from rats with total colonic aganglionosis (TCA). Ng ileum was obtained from TCA rats (spotting lethal: mutant, sl/sl: n = 15) at 10, 19, and 24 days of age (n = 5 for each age). Normal control Ng ileum was obtained from wild type littermates (+/+, n = 25) at 10, 19, 24, 30, and 60 days of age (n = 5 for each age). Specimens were studied with immunohistochemistry for PSA-NCAM and S100 beta. On H-E staining, normal mature ganglion cells (GC) were identified in submucus and myenteric plexuses in all specimens from TCA rats and normal controls. For PSA-NCAM, submucus and myenteric GC in control ileum were strongly positive at 10 days old, weakly positive at 19 days old, and did not stain from 24 days old and after. However, in all ileum specimens from TCA rats, both submucus and myenteric GC were strongly positive for PSA-NCAM regardless of age. For S100 beta, submucus and myenteric glial cells in control ileum were negative at 10 and 19 days old, but positive from 24 days old and after. However, in all ileum specimens from TCA rats, submucus and myenteric glial cells were S100 beta negative regardless of age. Our results suggest that GC in the Ng segment of TCA rats remain immature beyond an age when they should be mature.

This study was undertaken to compare magnetic resonance angiography (MRA) and gastrointestinal tract endoscopy (ENDO) for detecting varices in postoperative biliary atresia (BA) patients. Thirty-four BA patients were divided into groups according to age and liver function; group 1: 1-4 years old; group 2: 5-9 years old; group 3: over 10 years old; group A: normal liver function; and group B: moderate liver dysfunction. All subjects had MRA and ENDO. ENDO showed esophageal or gastric varices in 16 of 34 subjects. MRA depicted gastroesophageal vessels in 12 of 16 cases with varices on ENDO (sensitivity: 75%). Two subjects appeared to have varices on MRA that could not be confirmed on ENDO. Sixteen subjects had no varices on ENDO and normal MRA (specificity: 88.9%). Sensitivities and specificities of MRA for detecting varices were 20 (1/5) and 100% (6/6) in group 1 (n = 11), 100 (6/6) and 66.7% (4/6) in group 2 (n = 12), 100 (5/5) and 100% (6/6) in group 3 (n = 11), 100 (8/8) and 100% (9/9) in group A (n = 17), 50 (4/8) and 77.8% (7/9) in group B (n = 17), respectively. From these findings, MRA was not accurate in BA patients less than 10 years old or with moderate liver dysfunction. Therefore, periodic endoscopic examination is recommended for the accurate assessment and follow-up of varices in postoperative BA patients.

Hepatic portoduodenostomy with interposition of the ileocecoappendix (HPI) can be used for the surgical treatment of biliary atresia (BA). The purpose of this study was to evaluate the long-term outcome of patients with HPI. The records of nine patients who had HPI performed for BA were reviewed. Mean age at portoenterostomy was 68.1 days (range 26-113 days). At the end of 2006, seven of the nine subjects were alive, although two required liver transplantation (LT). The two deaths occurred 94 days and 2 years after HPI due to varicella infection and variceal bleeding, respectively. Length of follow-up for the seven survivors ranged from 17 to 19 years (mean 18 years). Three subjects achieved normal liver function after HPI and have remained jaundice-free to date. Another three who were initially jaundice-free required Roux-en-Y jejunostomy (RYJ) to the ileocecum for severe obstructive cholestasis 6 months, 3 years, and 19 years after HPI secondary to stones in the cecum. RYJ was successful in these three cases, and liver function returned to normal within a few months. Two of these three have continued to have almost normal liver function, but one required LT 5 years after RYJ. The remaining case had LT because of liver dysfunction 14 months after HPI. There is a high risk for stone formation and obstruction with the HPI procedure because bile can stagnate in the ileocecum.

Conventional acetylcholinesterase (AChE) histochemistry is both time consuming and complicated and requires the mixing of reagents that are toxic to the human body. We developed a rapid technique for performing AChE histochemistry, which has already been published, and now present a kit for performing AChE histochemistry that is a further improvement. Rectal suction biopsy specimens taken from 20 constipated patients and three full thickness biopsy specimens taken from 4 Hirschsprung's disease (HD) patients during pull-through surgery from aganglionic, transitional, and ganglionic bowel segments were tested using our rapid technique and the new kit. Each specimen was incubated for only 6 min. All ganglion cells stained clearly for AchE in just 6 min using both techniques. However, the kit was able to stain AchE positive nerve fibers more clearly and did not detect endogenous peroxidase-containing histiocytes, as did the earlier rapid technique. The kit could also detect AchE positive nerve fibers in the circular and longitudinal muscle layers, unlike the earlier rapid technique. The kit allows AChE histochemistry to be performed rapidly with complete accuracy, without any risk for toxicity. Moreover, the kit provides more focused information on AchE distribution in the bowel itself without any extraneous staining and can be used for diagnosing HD and allied disorders as well as establishing the exact level of innervation for pull-through resection.

To measure serum monocyte chemotactic protein-1 (MCP-1) in patients with congenital diaphragmatic hernia (CDH) and investigate its relationship to the development of persistent pulmonary hypertension (PPH). Serum MCP-1 was measured in 13 neonates with high risk for CDH at the time of diagnosis and postoperatively, and in five age-matched controls using an ELISA system. The 13 CDH subjects were divided into four groups according to the presence of PPH and outcome. Group I (severe-pre group): subjects with severe PPH who died prior to surgery (n = 5); Group II (mild-pre group): subjects with mild PPH controlled by medications (n = 8); Group IIa (severe-post group): subjects who subsequently developed severe PPH postoperatively and died (n = 3); and Group IIb (mild-post group): subjects who continued to have mild PPH controlled by medications. We also examined nitrofen-induced hypoplastic lungs from five rat fetuses with CDH and five control lung specimens for MCP-1 using immunohistochemistry. Mean serum MCP-1 in Group I was (1038.0 +/- 95.8 pg/ml), which was significantly higher than Group II (444.9 +/- 39.7 pg/ml) (P < 0.0001) and controls (147.3 +/- 11.3 pg/ml) (P < 0.0001). Postoperatively, Group IIa was significantly higher than Group IIb from 24 to 120 h postoperatively (P < 0.001). In Group IIb serum MCP-1 did not rise at all between 24 and 120 h postoperatively. Hypoplastic fetal rat CDH lungs had strong expression of MCP-1 compared with control lungs. Up-regulated expression and high circulating levels of MCP-1 in CDH patients with PPH suggest that MCP-1 may play a role in the development of PPH in CDH.

The oriental herbal formulation inchin-ko-to (ICKT) inhibits liver cell apoptosis induced by transforming growth factor-beta 1 (TGF-beta1). This study evaluated the effect of ICKT on serum markers of liver function and liver fibrosis in postoperative biliary atresia (BA) patients. Twenty-one postoperative BA patients with elevated GOT, GPT and gamma-GTP, but normal serum total bilirubin levels, were divided into two groups arbitrarily; an ICKT group (n = 12), and a no-ICKT group (n = 9). Serum markers of liver function [GOT, GPT, gamma-GTP, total bile acids (TBA)], and serum markers of liver fibrosis [hyaluronic acid (HA), type IV collagen (C-IV)], were measured in both groups at the beginning of the study, and at 1, and 3 years after the beginning of the study and the results compared statistically. All patients tolerated ICKT well, and there were no side effects. In the ICKT group, mean serum HA levels were significantly decreased at 1 year (P < 0.012), and at 3 years, both mean serum HA and C-IV were significantly decreased (P < 0.001 and P < 0.003, respectively). However, mean serum levels of GOT, GPT, gamma-GTP, and TBA did not change significantly following ICKT use for any length of time (all P > 0.05). Administration of ICKT in postoperative BA patients appears to lower the serum levels of markers of fibrosis in the medium-term. Whether this in fact correlates with prevention cannot be determined from this paper, but ICKT would appear to protect against liver fibrosis. Long-term studies are required to determine the exact role ICKT plays in prognosis of BA patients.

A review of 100 consecutive cases of congenital diaphragmatic hernia (CDH) treated at our institute focusing on the efficacy of protocolized management (PM) was conducted. Of the 100 cases, 14 who became symptomatic more than 24 h after birth, and seven with fatal anomalies (four cardiac and three chromosomal) were excluded, leaving 79 subjects for this study. Of these, 41 were diagnosed prenatally (PD). Subjects were divided into four groups. Group I: No PD, no PM (n = 34), Group II: No PD, PM (n = 4), Group III: PD, no PM (n = 21), and Group IV: PD, PM (n = 20). PM includes criteria for planned delivery, use of high frequency oxygenation, nitric oxide, echocardiography (EC), and a medication schedule. Overall survival rates for Groups I, II, III, and IV were 73.5% (25/34), 75% (3/4), 38.1% (8/21), and 70.0% (14/20), respectively. Survival rates were higher when PM was used: 70.8% (Groups II, IV) versus 60.0% (Groups I, III). Survival rates were significantly lower if diagnosed prenatally (PD+): 53.7% (Groups III, IV) versus 73.7% (Groups I, II) (P < 0.01). However, in PD+ groups, survival was significantly higher if PM was used (P < 0.05). PM significantly reduced length of hospital stay (35.5 vs. 52.0 days: P < 0.05). EC was found to be a predictor for survival while post-ductal AaDO(2) was not. In 17 cases with cardiac anomalies, PM did not affect survival. Our study suggests that use of PM for prenatally diagnosed CDH cases is associated with improved outcome, although the components of PM need to be tested in prospective trials to determine their true value.

Commonly, reduction of intussusception is performed by experienced radiologists. We review the performance of a pediatric surgical team for treating intussusception according to a standard protocol and present our findings. Three hundred and seventy eight patients with signs and symptoms of intussusception we treated from 1980 to 2005 were reviewed. Hydrostatic reduction (HR) was performed using a water-soluble contrast agent under fluoroscopy unless there was a serious condition clinically. Before 1998, HR was performed exclusively by pediatric surgical trainees (period A). In 1998, a standard protocol (double-balloon tube, maximum pressure of 120 cm H2O, repeated a maximum of five times, and HR performed by a pediatric surgical trainee under the supervision of a consultant pediatric surgeon) was adopted (period B). As part of the protocol, the operating room was notified of the HR procedure and placed on call for emergency surgery. Of the 378 patients, 21 required immediate laparotomy due to serious general condition, leaving 138 during period A and 219 during period B who had HR. Patient age, sex, and duration of symptoms (period A, 14.5 +/- 7.8 h; period B, 13.1 +/- 9.9 h) were not statistically significant. Success of HR during period A was 64.5%, and significantly improved for period B at 94.5% (P < 0.01). During period B, 128 of our patients had been referred from elsewhere for failed reduction attempted by radiologists or pediatricians. We were able to perform HR successfully in 118 of these (92.2%). During period A, it was significantly less at 54.0% (P < 0.01). Bowel perforation during HR occurred in two patients during period A (1.4%) and two patients during period B (0.9%), but the latter cases were transferred immediately for emergency surgery with good outcome. Reduction of intussusception by a pediatric surgical team would appear to be significantly safer with better outcome, and is thus more efficient.

To study the surgical treatment and outcome of hydronephrosis secondary to pelviureteric junction obstruction (PUJS) that is so massive that the renal pelvis crosses over the vertebral column [mega-hydronephrosis (MH)]. Of the 40 cases of PUJS we treated over the past 5 years, 6 cases had MH secondary to PUJS (MH-PUJS) in 6 renal units and were reviewed. Incidence of MH was 15.0%. All had Anderson-Hynes dismembered pyeloplasty to treat obstructive renal pattern on technetium-99m diethylenetriaminepentaacetic acid (DTPA) scans in five and gait disturbance due to MH in one. The contralateral kidney was normal in five and dysplastic in one. Three had nephrostomy before pyeloplasty. Preoperative DTPA scans showed good renal function in four and poor in two. Plication of the entire dilated, lax renal parenchyma ("nephroplication") was performed in three to decrease the intra-renal pelvic cavity. Prenatal diagnosis was performed in three cases. Mean age at pyeloplasty was 3.5 years and mean duration of follow-up was 1.5 years. Postoperatively, two of the three who did not have nephroplication had prolonged but not obstructed renal pattern on DTPA scans, and the other case who did not have nephroplication had nephrectomy for non-functioning kidney caused by recurrent pyelonephritis 2 years after pyeloplasty. In the three who had nephroplication, DTPA scans showed acceptable to relatively good passage in all cases. The surgical treatment of MH-PUJS is challenging with no fixed guidelines. However, nephroplication may be useful during pyeloplasty to decrease the size of MH-PUJS.

Cholangiography is often crucial for establishing the definitive cause of neonatal jaundice. We present our protocol for using laparoscopy-assisted cholangiography in infants with prolonged jaundice and discuss its benefits. Firstly, a 5 mm supra-umbilical trocar is introduced to create a port for a 0 degrees laparoscope. A 5 mm trocar is then inserted through a right subcostal incision to allow the liver and gallbladder to be visualized. If the gallbladder is of good size, the fundus is exteriorized through the right subcostal trocar site and a catheter is inserted into the gallbladder for cholangiography. If the gallbladder is atretic, the fundus is not exteriorized and a laparotomy is performed for open intraoperative cholangiography because the lumen of an atretic gallbladder is usually not fully patent and cholangiography through its exteriorized fundus often fails. We reviewed 18 jaundiced infants thought to have biliary atresia (BA) who had laparoscopy-assisted cholangiography. At laparoscopy, four patients had good sized gallbladders and minimal to mild liver fibrosis. They underwent cholangiography via the exteriorized fundus, and BA in two cases and biliary hypoplasia in two cases were identified. The remaining 14 had atretic gallbladders and varying degrees of liver fibrosis. Cholangiography via the exteriorized fundus was performed in one patient, but failed and converted to open cholangiography. Open intraoperative cholangiography identified BA in all 14 cases. All BA cases progressed to Kasai portoenterostomy directly after diagnosis. Laparoscopy is used to determine the type of cholangiography to be performed based on the appearance of the gallbladder and this simple, accurate, and safe protocol allows the anatomical structure of the biliary tree to be obtained accurately with minimal surgical intervention.

Symptomatic hepatic duct diverticulum located outside the liver is extremely rare, with only one reported case. We encountered this anomaly and present our experience, focusing on surgical management. A 17-year-old-girl was referred to our institute because of recurrent right upper quadrant pain. At presentation, serum amylase, lipase, bilirubin, and transaminases were normal. White blood cell count and C-reactive protein were slightly raised. Ultrasonography and magnetic resonance cholangiopancreatography (MRCP) revealed a diverticulum-like lesion on the right side of the bile duct system appearing to overlap the gallbladder. The provisional diagnosis was type II choledochal cyst. Pancreaticobiliary malunion (PBMU) could not be confirmed on MRCP. The origin of the lesion could not be identified initially at laparotomy, but after repeated intraoperative cholangiography (IC) studies, the lesion was found to originate from a normal-sized right hepatic duct via a narrow duct. It was relatively easy to ligate the narrow duct and treat the lesion smoothly. IC also showed that the common bile duct was not dilated and that PBMU was absent. Histopathology showed the lesion to be a diverticulum of the bile duct epithelium. This is only the second report of a symptomatic hepatic duct diverticulum located outside the liver. Based on our experience, IC is particularly useful for the complete understanding of anomalous hepatobiliary duct anatomy and for planning surgical treatment.

In high jejunal atresia (HJA), discrepancy between the diameters of the proximal and distal jejunum can often be significant and cause major anastomotic complications. We developed 2 novel procedures to enhance conventional jejuno-jejunostomy (JJ): bilateral side-plication during JJ (BSP): plication of both lateral aspects of the dilated proximal jejunum, and jejunal plication before JJ (pre-P): plication of the jejunum proximal to the planned JJ site along the anti-mesenteric line. We used BSP in a neonate with a membranous stenosis 5 cm distal to the duodeno-jejunal flexure, and pre-P in 2 neonates, both with HJA and hugely dilated proximal jejunum. By using BSP, the jejunum could be plicated symmetrically near the closed jejunotomy, allowing the caliber change at the closed jejunotomy to be smooth without any kinking. If simple anti-mesenteric plication had been performed instead, proximal-to-distal kinking caused by closing the jejunotomy site transversely after the membrane was excised could have worsened. By using pre-P, the caliber change across the JJ could be made smooth and symmetric. If plication had been performed along the anti-mesenteric line after completing the JJ, the proximal jejunum near the JJ would have become asymmetrical causing the JJ to become kinked. Postoperatively, there were no JJ-related complications, and all subjects are well after a mean follow-up period of 3.0 years. Our novel procedures are simple and appear to enhance JJ by controlling kinking at the JJ site.

We experienced two cases with disseminated HP and IND occurring with normal bowel in between (disseminated mixed intestinal dysmotility--DMID) and postulate whether it could be classified as a new intestinal motility disorder. Our cases, both boys, died at 3 and 7 months, respectively. Both had irregular stool passage, and abdominal distention with bilious vomiting since birth. On barium enema, both had rigid distal ileum and colon with narrow lumens, with dilated and atonic proximal ileum and jejunum. An ileostomy was created on days 3 and 2 of life, respectively, however, they did not function and jejunostomies were created, which also did not function well. Both boys died after repeated episodes of severe enterocolitis. In each case, three 10 cm specimens were obtained randomly from the jejunum and ileum, and two 5 cm specimens were obtained randomly from each of the ascending colon, transverse colon, descending colon, and rectum and treated with hematoxylin and eosin (H & E) staining, acetylcholine esterase (AchE) histochemistry, and protein gene product 9.5 (PGP9.5) and neural cell adhesion molecule (NCAM) immunohistochemistry for histopathologic assessment. All specimens showed a mixture of disseminated IND and HP, with normal intestine in between. There was abnormal expression of NCAM activity in the intestinal smooth muscle layers in small and large intestine. This is the first report about disseminated IND and HP occurring with normal bowel in between (DMID) and we suggest it should be classified as a new intestinal motility disorder. The present findings demonstrate that patients with DMID have a complicated abnormality of NMJ that may directly influence bowel motility and prognosis according to the severity of the abnormality.

We used Acetylcholinesterase (AchE) staining and neural cell adhesion molecule (NCAM) immunoreactivity to examine such resected lengths to determine the complete innervation profile of resected bowel in Hirschsprung's disease (HD). Resected specimens of colon obtained at pull-through surgery from 15 patients with HD [short type (S-type: n=5), recto-sigmoid type (RS-type: n=5), long type (L-type: n=5)] were sectioned at 1.5 cm intervals and stained conventionally with AchE histochemistry and NCAM immunohistochemistry. The number of positive nerve fibers (PNFs) in the lamina propria and smooth muscle layers was assessed on a scale of 0 to 3 where 0 meant no PNF and 3 meant many PNFs. The three types of HD had different AchE and NCAM innervation profiles, especially the L-type. There were also different AchE and NCAM innervation patterns seen within the same aganglionic or transitional segments of bowel depending on the site of sampling. The mean proportion of transitional segment in resected specimens from RS-type HD was significantly larger than that of S-type HD (P<0.001) although the proportion of aganglionic segments from S- and RS-type HD were almost the same. Our results suggest that the etiology of L-type HD may be different from the etiology of S- and RS-type HD because of different innervation profiles. Because segments of the excised bowel would appear to have different innervation patterns depending on whether the specimen is sampled proximally or distally, it is important to record the exact site of sampling to allow valid comparisons between types of HD to be made.

Median sternotomy (M-S) provides excellent exposure and allows dissection of the medial side of an anterior mediastinal tumor (AMT) with minimal risk. We report our experience of resecting AMT using M-S. Five children with AMT were treated using M-S between 1997 and 2004 at our institute. Ages at M-S ranged from 8 months (case 2) to 9 years (case 1) and AMT ranged in size from 5x6x7 cm3 (case 2) to large enough to occupy nearly the entire right thoracic cavity (cases 4 and 5). AMT were resected completely in all cases through M-S with the patient in the supine position. M-S alone was used in cases 1, 2, and 3, and cases 4 and 5 required additional incisions. There were dense adhesions between AMT and important mediastinal structures such as the anterior part of the pericardium (cases 1 and 2), left phrenic nerve (case 3), and pulmonary vessels (case 4, 5), but all were dissected safely under direct vision. There was no respiratory or cardiovascular compromise during M-S due to compression of the healthy lung and mediastinum. Histopathologic findings were mature teratoma in cases 1, 3, and 5, immature teratoma in case 2, and pleuropulmonary blastoma in case 4. Postoperative recovery was unremarkable. After mean follow-up of 3.4 years, cases 1, 2, 3, and 5 are currently well with no signs of recurrence, but case 4 died from disseminated intravascular coagulation during postoperative chemotherapy for massive local tumor recurrence 6 months after surgery. Based on our experience, M-S allows access to all aspects of AMT under direct vision, and provides excellent exposure, thus facilitating complete resection.

We report two cases of rare cystic dilatations of the ejaculatory system. In case 1, a 6-month-old boy was referred to us for the management of recurrent epididymo-orchitis (E-O) complicating open drainage and a colostomy was performed elsewhere for a purulent rectal discharge thought to be rectal duplication. Diagnostic imaging showed a retrovesical cyst. Urethrocystoscopy showed a swelling of the verumontanum. No fistula was seen between the cyst and rectum on colonoscopy. At laparotomy, both ejaculatory ducts and seminal vesicles were found to be fused into a mass with cystic dilatation of the ejaculatory duct. Intraoperative histopathology of the cyst identified a metaplastic epithelial lesion. The cyst was excised with bilateral vasoligation. Since surgery, 8 years ago, urination and defecation have been normal. In case 2, a 4-month-old boy presented with fever and a swollen right scrotum. Ultrasonography showed a retrovesical cyst. Right grade IV vesicoureteral reflux diagnosed on voiding cystourethrography was treated by ureter reimplantation (Cohen) but complicated by recurrent E-O. Urethrocystoscopy with retrograde contrast via the utriculus showed that the cyst opened on the verumontanum, that both ejaculatory ducts opened into the cyst, and there was reflux into the right vas deferens. Right vasoligation alone was performed through a scrotal approach. Although the cyst was not excised, there has been no recurrence of E-O nor enlargement of the cyst for 6 years. Cysts of the ejaculatory system should be considered in the etiology of recurrent E-O in prepubertal children and a high index of awareness is recommended.

Traditionally, choledochal cyst (CC) is classified into five types according to Todani's classification. Of 356 cases of CC treated at our institute since 1965, we experienced four cases of mixed type I and II choledochal cyst (MTCC). The incidence of MTCC was 1.1%. All four patients had a fusiform common bile duct (CBD) with a diverticulum originating from the mid-portion of the CBD. The cystic duct drained into the right side of the diverticulum. All had pancreaticobiliary malunion (PBMU). All presented with recurrent abdominal pain. MTCC was suggested on ultrasonography or computerized tomography and confirmed by endoscopic retrograde cholangiopancreatography (three cases) or magnetic resonance cholangiopancreatography (one case). Age at surgery ranged from 3 to 12 years. All had hepaticoenterostomy, and total excision of the CC/diverticulum/gallbladder. There were no complications and all cases are currently well. In all cases, histopathology showed that the wall of the diverticulum was identical to the CBD. Although the incidence of MTCC is very low, pediatric surgeons should be aware of it as a new subtype of CC. Based on our experiences and a review of the literature, we suggest that its etiology does not affect treatment because the surgical management of choice is the same as for type I CC.

The aim of this study was to analyze the short- to mid-term outcome of primary modified Georgeson's laparoscopy-assisted endorectal pull-through (PMGLEPT) for Hirschsprung's disease (HD). HD patients treated by PMGLEPT were evaluated prospectively by a single surgeon using a standard structured questionnaire to assess complications, incidence of enterocolitis, and evaluate continence (CE). CE involved scoring five parameters (frequency of motions, severity of staining/soiling, severity of perianal erosions, anal shape, and requirement for medications) on a 3-point scale (0, 1, and 2 for each parameter). Thus, scores for CE were: 10 = normal, 8-9 = good, 6-7 = fair, and 0-5 = poor. Our modifications include transanal rectal dissection starting below or on the dentate line, near total excision of the posterior rectal cuff, and intraoperative acetylcholinesterase staining to accurately identify normal colon. Patients with total colon aganglionosis or trisomy-21 were excluded, leaving 33 cases of PMGLEPT performed between 1997 and 2004. Mean operative age was 11.0 months. Follow-up ranged from 8 months to 7 years (mean 4.0 years). There were no intraoperative complications. Post-PMGLEP, bowel obstruction occurred in 1 subject who required middle colic division for pull-through (PT), and enterocolitis occurred in 3 (9.1%) of 33 patients. In 20 subjects aged over 3 years with a follow-up period of more than 12 months, final CE was normal in 5, good in 10, fair in 4, and poor in 1, despite staining/soiling being present in 12 (60%) of 20 subjects. None of the 33 had constipation. Our results suggest that PMGLEPT is safe with acceptable outcome in the short- to mid-term. However, careful long-term follow-up is mandatory as there appears to be a relatively high incidence of staining/soiling on short- to mid-term follow-up.

Surgical stress induces systemic endocrine-metabolic responses that influence the function of endothelial cells (EC) to cause various systemic reactions. Intercellular adhesion molecule (ICAM)-1 is an adhesion molecule that plays an important role in inflammation, and increased expression of ICAM-1 on EC is a reflection of EC activation. In this study, we investigated the ICAM-1 response to surgical stress in neonates undergoing major surgery. Fifteen neonates (mean age at surgery: 3.5 +/- 1.2 days) were divided into two groups according to indications for surgery: Group I: Congenital diaphragmatic hernia without persistent pulmonary hypertension (n = 5); Group II: Gastrointestinal surgery [n = 10: duodenal atresia (n = 3), intestinal atresia (n = 6), and esophageal atresia (n = 1)]. Serum samples were obtained preoperatively, immediately after completion of surgery (time zero), and 24, 48, 72, 96, and 120 h after surgery to measure ICAM-1 levels using an enzyme-linked immunosorbent assay, C-reactive protein (CRP), and white blood cell count (WBC). Postoperative recovery was uneventful in all cases. ICAM-1 levels in both groups increased significantly within 24 h of surgery (Group I: P = 0.0038, Group II: P = 0.0320). In Group I, ICAM-1 peaked 72 h postoperatively while in Group II it continued to rise until 96 h postoperatively. The difference between peak levels reached was not significant. CRP was first detected 24 h postoperatively in both groups and continued to increase until 48 h postoperatively. Again, the difference between peak levels reached was not significant. No significant changes in WBC were observed in either group. We found that ICAM-1 increases in response to surgical stress in neonates, although there was no significant difference in levels. However, surgical stress as represented by serum ICAM-1 would appear to last longer with intestinal surgery than with non-intestinal surgery. Further research is required to establish the usefulness of ICAM-1 as an easily detectable substance associated with endothelial damage that reflects the host's response to major surgical stress.

Rectal atresia is a rare condition in which the anus and sphincter muscles are normally developed, with usually no fistulous communication with the urinary tract. An unusual case of rectal atresia associated with recto-bulbar urethral fistula treated by a combination of colo-anal anastomosis and mucosal proctectomy via a posterior sagittal approach is reported for the first time.

We reviewed our experience of Roux-en-Y hepaticojejunostomy (RYHJ) and hepaticoduodenostomy (HD) performed for the surgical repair of choledochal cyst (CC), with special emphasis on postoperative complications related to the type of biliary reconstruction performed. Eighty-six patients underwent primary cyst excision for CC from 1986 to 2002 at our institution. Forty-six cases with concurrent intrahepatic bile duct dilatation (IHBD) were excluded because HD was not used for biliary reconstruction if IHBD was present. Thus, 28 cases had RYHJ, and 12 had HD. Differences between the RYHJ and HD groups with respect to type of CC, age at cyst excision, and length of follow-up were not statistically significant. However, the incidences of postoperative complications related to biliary reconstruction, such as endoscopy-proven bilious gastritis due to duodenogastric bile reflux [4/12 (33.3%) of the HD group], and adhesive bowel obstruction/cholangitis [2/28 (7.1%) of the RYHJ group] were significantly different (p<.05). Our experience suggests that HD is not ideal for biliary reconstruction in CC because of a high incidence (33.3%) of complications due to duodenogastric bile reflux. Currently, RYHJ is our exclusive technique of choice for biliary reconstruction during the surgical repair of CC.

Treatment of "forme fruste" choledochal cyst (FFCC) where pancreaticobiliary malunion (PBMU) is associated with minimal dilatation of the common bile duct (CBD) remains controversial. PBMU allows pancreaticobiliary reflux to occur, which causes complications such as recurrent pancreatitis and gallbladder cancer. Therefore, some surgeons opt to treat FFCC by cholecystectomy alone in order to prevent gallbladder cancer, with the result that pancreaticobiliary reflux could still occur. Our treatment of choice-excision of both the CBD and the gallbladder, followed by Roux-en-Y hepatico-jejunostomy-can eliminate pancreaticobiliary reflux and prevent complications. Our case, a 2-year-old boy, initially presented to a hospital abroad with recurrent abdominal pain. Endoscopic retrograde cholangiopancreatography showed massive protein plugs impacted in the papilla of Vater and mild CBD dilatation, but PBMU was not identified. Intraoperative cholangiography performed during laparotomy 5 days later suggested PBMU with minimal CBD dilatation. Despite these findings, cholecystectomy with T-tube drainage was performed rather than CBD excision with biliary reconstruction. Postoperative T-tube cholangiography clearly showed PBMU. The T-tube was removed after 2 weeks, and 3 months later the boy was referred to us because of recurrent pancreatitis. We performed CBD excision and Roux-en-Y hepatico-jejunostomy. His postoperative course was uneventful, and he is well after 10 years of follow-up. This case provides strong evidence that CBD excision with biliary reconstruction is mandatory for treating FFCC and, conversely, that cholecystectomy alone is inadequate for treating children with FFCC.

Systemic inflammatory disorders causing renal tissue damage do so by the adherence of polymorphonuclear leukocytes to endothelium, a process that is mediated by cell surface adhesion molecules. We determined the circulating levels of serum vascular cell adhesion molecule-1 (VCAM-1) in vesicoureteric reflux (VUR) patients and investigated the relationship between serum VCAM-1, grade of VUR, and secondary renal scarring. Serum levels of VCAM-1 were measured in 53 children aged between 3 months and 15 years with VUR (13 had grade III, 29 had grade IV, and 11 had grade V) and 25 controls using ELISA. Radionuclide scanning was used to assess renal scarring. Renal scarring was found in 29 of the 53 subjects. Serum VCAM-1 was significantly higher in subjects with high grades of VUR without renal scarring (grade IV: 715.9+/-121.0 ng/ml; grade V: 778.5+/-33.2ng/ml) compared with subjects with grade III VUR without renal scarring (609.8+/-64.3ng/ml, p<0.01). Serum VCAM-1 was also significantly higher in subjects with high grades of VUR with renal scarring (grade IV: 791.2+/-131.9ng/ml; grade V: 1171.8+/-235.6 ng/ml) compared with subjects with grade III VUR with renal scarring (687.3+/-163.4 ng/ml, p<0.001).

We report two rare cases of botryoid Wilms' tumor (BWT) occupying the renal collecting system with no macroscopic parenchymal mass. In case 1, a 3-year-old boy presented with a mass in the right flank, low-grade fever, abdominal pain, and macrohematuria. Radiology revealed an enlarged right kidney with a heterogeneous mass occupying a large part of the dilated renal calyx, pelvis, and ureter. Radical right nephroureterectomy was performed. The histopathologic diagnosis was nephroblastoma, and the pedicle of the tumor was attached to the renal parenchyma near the pelvic wall. In case 2, a 2-year-old boy presented with macrohematuria, and the clinical course was almost the same as in case 1. No local recurrence or metastatic spread has been detected for 4 years postoperatively in case 1 and for 9 months postoperatively in case 2.

Cathelicidins are a family of antibacterial peptides. Human cathelicidin LL-37 inhibits the binding of endotoxin lipopolysaccharide (LPS) to CD14-positive cells and could ameliorate sepsis. The aim of this study was to observe the effect of LL-37 on sepsis in neonatal rats. Intraperitoneal injection (IPI) of LPS was used to create sepsis in suckling rats. Group 1 rats were given LPS with LL-37, group 2 rats were given LL-37 2 h after LPS, and group 3 rats were given LPS without LL-37. Control group rats were given isovolemic normal saline by IPI. Rats given LL-37 IPI were divided into seven subgroups. Following IPI, an overall assessment score (OAS) and rectal temperature (RT) were assessed hourly. Serum C-reactive protein (CRP) was also assessed at death or at sacrifice 10 h after IPI. All rats in group 3 died. For rats receiving lower doses of LL-37 in groups 1 and 2, mortality was decreased. No deaths occurred among those receiving higher doses of LL-37 in group 1; however, mortality increased in group 2. In group 1, OAS and RT deteriorated initially for those receiving lower doses of LL-37, then improved. OAS and RT did not deteriorate throughout the study in rats given higher doses of LL-37. In group 2 rats given higher doses of LL-37, OAS and RT were not significantly different from rats in group 3. CRP was significantly decreased in group 1 compared with group 3, and decreased in group 2 for lower doses only. We conclude that LL-37 may prevent sepsis and be useful in lower doses for treating sepsis. However, LL-37 appears to have adverse effects when used at higher doses for treating sepsis.

Connective tissue growth factor (CTGF) is a newly described protein that stimulates transforming growth factor-beta1 (TGF-beta1). We evaluated the expression of CTGF mRNA in operative biopsy specimens from biliary atresia (BA) patients and normal controls to assess the role of CTGF in BA. Liver biopsy specimens were taken from BA patients at the time of portoenterostomy (n=22) and compared with specimens taken from normal controls (n=6). In situ hybridization was used to stain CTGF mRNA in all specimens. The distribution of collagen type IV (C-IV) was also assessed in the same specimens as an indicator of the severity of fibrosis present at the time of biopsy. Results were analyzed to determine whether there was any correlation between CTGF and C-IV and outcome. Of the 22 postoperative BA patients, eventual outcome was good in 17 (group I), and five (group II) subsequently required liver transplantation. Control specimens (n=6) had no apparent CTGF mRNA expression, and median C-IV positive immunoreactivity was 1.23+/-0.25%. CTGF mRNA was weakly expressed in hepatic stellate cells (HSC) and hepatocytes in specimens from group I, and median C-IV-positive immunoreactivity was 3.18+/-0.86%. However, in specimens from group II, there was increased CTGF mRNA in HSC and hepatocytes. Median C-IV-positive immunoreactivity was 6.31+/-0.96%. There was a significant correlation between CTGF mRNA intensity and the amount of C-IV, which implies that CTGF expression reflects prognosis. This study provides the first evidence that CTGF is strongly expressed in BA, in particular in HSC and hepatocytes, suggesting that they may be a source of CTGF. The strong correlation with C-IV indicates that CTGF plays a major role in the pathogenesis of progressive fibrosis in BA.

Currently, laparoscopic-assisted colon pull-through (LACPT) is the treatment of choice for male patients with high-type imperforate anus and rectourethral fistula. Since laparoscopy was introduced for treating this condition, reports concerning post-LACPT complications are rare. Here we discuss the case of a boy, now 3.5 years old, born at 37 weeks' gestation weighing 2,300 g, who was diagnosed with rectobulbar urethral fistula (RUF) at birth. LACPT was performed when the boy was 11 months old and weighed 7.2 kg. No intraoperative complications occurred, and the initial post-LACPT course was uneventful. When he was 2 years old, he developed dysuria requiring urethral catheterization. Diagnostic radiology confirmed a large cystic mass behind the bladder, suggestive of a posterior urethral diverticulum (PUD). Histopathology of the excised mucosa of the cyst showed colonic mucosa, confirming that the cyst was indeed an enlarged residual RUF. We discuss our treatment and our approach to prevention.

The aim of this study was to determine circulating levels of adhesion molecules in serum from patients with congenital diaphragmatic hernia (CDH) to investigate the relationship between soluble ICAM-1, ELAM-1, and VCAM-1 liberated by activated vascular endothelium and the development of persistent pulmonary hypertension (PPH) in patients with CDH. We measured serum levels of ICAM-1, ELAM-1, and VCAM-1 in 20 high-risk neonates with CDH at the time of diagnosis (11 with PPH and 9 without PPH) and 7 age-matched controls using ELISA system. We further examined the lungs of 5 patients with CDH complicated by PPH who died during resuscitation and stabilization, and three control lung specimens for the expression of adhesion molecules using immunohistochemistry. The mean serum ICAM-1 levels in CDH patients with PPH (227.0+/-98.9 ng/ml) were increased compared with levels in CDH patients without PPH (140.29+/-37.4 ng/ml; p<0.05) and controls (130.0+/-23.8 ng/ml; p<0.05). Mean serum ELAM-1 levels in CDH patients with PPH (116.5+/-19.2 ng/ml) were significantly increased compared with levels in CDH patients without PPH (79.3+/-27.9 ng/ml; p<0.01) and controls (58.4+/-14.5 ng/ml; p<0.001). Mean serum VCAM-1 levels in CDH patients with PPH (1596.9+/-460.4 ng/ml) were significantly higher compared with levels in CDH patients without PPH (1069.3+/-444.6 ng/ml; p<0.01) and controls (838.0+/-171.2 ng/ml; p<0.001). But serum adhesion molecule levels in CDH patients without PPH were no different from controls statistically. Pulmonary vascular endothelial cells from CDH lung with PPH had strong expression of adhesion molecules compared with controls. Up-regulated expression of adhesion molecules on the endothelium of pulmonary vessels and high circulating levels of adhesion molecules in CDH patients with PPH suggest that adhesion molecules may play a role in the development of PPH in CDH.

An 18-year-old girl who had undergone excision of a choledochal cyst and Roux-en-Y hepatico-jejunostomy at another hospital when 23 months old was referred to our department because of recurrent cholangitis. Radiological investigations showed stones lying in minimally dilated, right posterior intrahepatic bile ducts (IHBD). At laparotomy, the hepatico-jejunostomy site was incised, and a flexible endoscope inserted into the IHBD. Multiple stones packed in the IHBD were easily fragmented using an electro-hydraulic lithotripsy (EHL) device inserted through the endoscope, and removed. There were no EHL-related complications, and her postoperative progress was uneventful. She is currently well with no episodes of cholangitis after a follow-up period of 3 years. EHL is a simple, effective alternative method for removing IHBD stones after choledochal cyst excision. To the best of our knowledge, this is the first report of EHL being used to remove stones that developed in the IHBD after choledochal cyst excision.

There has been no report describing suspected prenatal pyriform sinus (PS) cyst and fistula. We report a case suspected by prenatal ultrasonography and fetal MRI. A large cystic mass was found in the left neck of the fetus. After the baby was born, preoperative laryngoscopic catheterization of the fistula tract was used to confirm the diagnosis and greatly facilitated the identification and excision of the PS cyst and fistula.

We investigate the incidence, treatment, and implications of intrahepatic bile duct (IHBD) dilatation debris in 42 patients with choledochal cyst treated over a recent 8-year period. Mean age at cyst excision was 6.9 years, and mean follow-up period was 5.4 years. Intraoperative endoscopy (IOE) was performed at the time of cyst excision using a pediatric cystoscope, which identified IHBD debris in 11/42 (26.2%), and massive debris in 4/42 (9.5%). In all cases, IHBD debris was successfully washed out with normal saline through the cystoscope. There was no postoperative IHBD stone formation or cholangitis except for one case of cholangitis in a patient undergoing chemotherapy for acute myelocytic leukemia. Debris left in the IHBD during cyst excision is probably one of the primary causes of postoperative IHBD stone formation and can be prevented by washing during IOE.

We examined whether non-hematopoietic BM cells can migrate into the intestinal graft after fetal small intestinal transplantation (FSITx). Fetal small intestine from donor C57BL/6 mice was transplanted into the rectus abdominis of recipient C57BL/6 mice with only green fluorescent protein (GFP) BM cells (syngeneic FSITx). Intestinal grafts were harvested on days 5, 10, and 30 after FSITx and stained immunohistochemically using anti-CD45 antibody (a marker for hematopoietic BM cells). Although there were no GFP-positive cells identified in the epithelium of the graft intestinal villi, there were a few cells positive for both GFP and CD45 in the lamina propria on day 5 after FSITx, and many present on days 10 and 30. In some grafts there were only cells that were GFP positive/CD45 negative (i.e., non-hematopoietic BM cells) found in the lamina propria on days 10 and 30. These data indicate that non-hematopoietic BM cells as well as hematopoietic BM cells can migrate from the recipient's bone marrow, suggesting that recipient mesenchymal stem cells may be strongly implicated in graft regeneration and development after FSITx.

Background: Hypoganglionosis (HP) is characterized histologically by a decreased number of ganglion cells in intestinal myenteric plexuses and functionally by severely impaired gut motility. The purpose of the present study was to investigate the neuromuscular junction (NMJ) in small- and large-intestine biopsy specimens from patients with HP., Methods: Fresh frozen sections of small and large intestine from three patients with HP and five age/site-matched controls were examined. All specimens were labeled with neural cell adhesion molecule (NCAM) antibodies and synaptophysin (SY) antibodies to delineate the NMJ., Results: Normal small- and large-intestine specimens had a moderate number of NCAM-positive NMJ in association with nerve fibers in the lamina propria, muscularis mucosa, and many NCAM-positive NMJ in association with nerve fibers in the circular and longitudinal muscle layers. In HP, there was lack of expression of NCAM-positive NMJ in association with nerve fibers in the lamina propria, muscularis mucosa, and circular and longitudinal muscle layers in both small- and large-intestine specimens, and a marked increase in NCAM expression in the muscularis mucosa and the inner border of the circular muscle layer that was only seen in HP specimens. SY-positive synaptic vesicles were observed throughout the gut in all normal specimens, but there was a lack of SY expression in HP specimens. Expression of NCAM and SY in ganglion cells was the same in all specimens., Conclusion: These findings suggest there is a complicated abnormality of the NMJ in HP that may prove to contribute to the disturbances in gut motility seen in this condition.

Rectal suction biopsy (RSB) is a well-known diagnostic procedure for disorders of bowel motility such as Hirschsprung's disease (HD). However, there are few reports about the optimal method of obtaining rectal tissue. We introduce a new technique using Gruenwald's nasal cutting forceps (NCF). From 1986 to 1999, we performed 130 sets of rectal biopsies in patients suspected of having HD. In group I (1986 to 1994), 68 sets of three-site biopsies (2, 3, and 5 cm above the dentate line) were performed using a conventional blind RSB technique. In group II (1995 to 1999), 62 sets of one-site biopsies (2 cm above the dentate line) were performed using Gruenwald's NCF after anal dilatation during general anesthesia. Hematoxylin-eosin staining and acetylcholinesterase histochemistry were used to examine all specimens. Biopsy specimens in group II (4.39 +/- 1.07 mm(2)) were larger than in group I (1.59 +/- 0.39 mm(2)) ( P < 0.01). In 18 cases (26 %) in group I, normal and HD bowel could not be differentiated because the specimens were too small to detect ganglion cells (i.e., only lamina propria [9 cases] or a small area of submucosa [9 cases] was present). These cases required repeat biopsy. All cases of HD diagnosed in group I (n = 20) were based on the findings of biopsies taken at 2 cm; biopsies from 3 and 5 cm did not provide additional information. There were 2 cases of post-biopsy hemorrhage in group I. In group II, 18 subjects were diagnosed with HD and 39 were confirmed to have normal bowel. There were no complications and repeating the biopsy was unnecessary. Three cases of hypoganglionosis (1 in group I and 2 in group II) were missed because the myenteric plexus abnormalities could not be detected by RSB. Intestinal neuronal dysplasia (IND) was diagnosed in 5 cases (2 in group I by repeat full-thickness biopsy and 3 in group II by rectal biopsy). We conclude that our new technique is advantageous and safe to differentiate between normal bowel, HD, and even IND on the basis of a single biopsy taken 2 cm above the dentate line. The biopsy can be taken under direct vision and is histopathologically accurate.

To describe manifestations of acetylcholinesterase (AchE) activity in the bowel of patients presenting with refractory constipation and correlate them with outcome, rectal biopsy specimens (RBS) from 165 patients who presented with refractory constipation between 1988 and 1999 were examined. Age at biopsy ranged from 4 days to 17 years; 45 subjects were excluded because they satisfied diagnostic criteria for Hirschsprung's disease, intestinal neuronal dysplasia, or hypoganglionosis. Thirty-five autopsy subjects were used as controls. All RBS were compared and AchE activity was assessed in the lamina propria (LP), muscularis mucosae (MM), and around the submucosal vessels (V). Variations in AchE distribution were classified as grade I (no AchE-positive nerve fibers in the LP or MM), grade II (some positive fibers in the LP or MM), grade III (moderate positive fibers in the LP or MM), grade IV (many positive fibers in the LP, MM, or V), or grade V (fibrillar, foamy, or amorphous staining for AchE). All grade I (11/120) and V (12/120) subjects achieved normal bowel control with laxatives alone and all grade II subjects (58/120) did with laxatives and enemas. Grade III subjects (34/120) required addition of cisapride. All grade IV subjects (5/120) were unresponsive to conservative management and 4/5 were found to have a megarectum, which was treated surgically. AchE distribution correlated well with eventual outcome and requirement for surgery. AchE distribution could also be used to classify bowel motility disorders, and we suggest the term AchE-positive disease be used to describe them.