Development of Clinical Referral Score Model for Early Diagnosis of Hirschsprung’s Disease in Suspected Pediatric Patients

The diagnosis of Hirschsprung’s disease (HSCR) relies on history, physical examination, and investigations. Some of investigation modalities could not be done in primary hospital. This study was aimed to develop the clinical score model for diagnosing and early referrals of HSCR, especially in areas where investigations were not available. Overall 483 consecutive suspected HSCR patients who were under 15 years old from January 2006 to December 2020 were included in this study, with 207 (42.86%) patients diagnosed with HSCR and 276 (51.14%) patients in the non-HSCR group. Five clinical parameters were included in the prediction model. The AuROC of clinical parameters, which included having an age younger than one month, male gender, the term infant, history of delayed meconium passage, and history of enterocolitis, was 72%. The prediction score ranged from 0–7, with a score 0–3 meaning a low risk to be HSCR (LHR+ = 0.37). We concluded that patients with suspected HSCR who had clinical score 4–7 had a high probability to be HSCR and, thus, it was suggested that these patients have an early referral for further investigations, which were contrast enema and rectal suction biopsy. In the case of a low probability of HSCR, clinical observation is still warranted. This clinical scoring system can be used as a screening tool to prevent delay diagnosis and complications.

Increasing Trend of Calretinin-Positive Mucosal Innervation from Aganglionic Zone toward Transition Zone in Hirschsprung's Disease

Introduction Diagnosis of very short-segment Hirschsprung's disease (vsHD) by rectal suction biopsy is challenging as its aganglionic zone (AZ) overlaps with physiologic hypoganglionic zone and calretinin-positive mucosal nerves may extend from the transition zone (TZ) into AZ. We studied whether an increasing trend/gradient of calretinin-positive mucosal nerves along the distance from AZ toward TZ aids in diagnosis of HD. Materials and Methods In this study, 46 rectal suction biopsies from non-HD and HD, and 15 pull-through specimens from short-segment HD were evaluated by mucosal calretinin immunostain (CI) and image processing and analysis (IPA) to measure pixel count (PC, the percentage of calretinin stained pixels in the mucosa). Consecutive longitudinal sections of proximal AZ toward distal TZ in HD pull-through specimens were utilized as a vsHD surrogate model. First, we studied variability of mucosal CI in non-HD biopsies along the distance from dentate line. Second, we determined a cutoff point of mucosal CI by IPA that separated non-HD versus HD and applied this cutoff to longitudinal sections from proximal AZ to distal TZ segments in HD pull-through specimens. Third, we studied whether an increasing trend of mucosal CI was universally observed in HD pull-through. Results Our findings included a significant variability in PC along the biopsy distance in non-HD cases. Positive mucosal CI was found in proximal AZ in 6 (43%) of 14 HD pull-through, among which 1 case lacked submucosal nerve hypertrophy in the proximal AZ. All 14 HD pull-through cases showed an increasing trend/gradient of PC from AZ toward TZ. Conclusion Based on our findings, the presence or absence of mucosal CI positivity and submucosal nerve hypertrophy may not reliably diagnose vsHD in rectal suction biopsy. While we acknowledge that the density of mucosal innervation in variable contexts and anatomical locations is unknown and yet to be explored, our study suggests that an increasing trend of positive mucosal CI from AZ toward TZ by IPA might prove to be a useful tool for the diagnosis of vsHD in the future.

Constipation and dilated bowel: Hirschsprung’s disease is not always the case

A case of a 2-month-old boy with constipation and a localized abdominal distension in the right abdomen and hypogastrium is presented. Plain radiograph, ultrasound exam as well as a barium enema were suggestive of Hirschsprung’s disease (HD) (ultrashort segment disease). Nevertheless, rectal suction biopsy was negative for neuronal abnormalities and unexpectedly on plain radiograph vertebral abnormalities were noticed. Subsequently magnetic resonance imaging of the spine and abdomen was performed, to evaluate possible spinal cord lesions and potential abnormalities of the perineal region musculature. A syringe throughout medullary cone was noted. The well-known issue, that the diagnosis of HD does not depend on the imaging (radiological) findings, was confirmed. The step-bystep diagnostic approach from the initial thought of ultrashort segment HD to the later imaging-based diagnosis of syringomyelia is discussed in this present paper.

Calretinin versus Hematoxylin and Eosin Stain for Diagnosis of Hirschsprung’s Disease; Comparison in Ganglionic, Transitional, and Aganglionic Zones

Objective: To compare the results of calretinin stain with hematoxylin and eosin (H&E) stain for diagnosis of Hirschsprung’s disease. Materials and Methods: A prospective double-blind diagnostic study was done. Hirschsprung’s patients who underwent transanal endorectal pull through (TERPT) surgery between February 2015 and February 2017 were included. The pulled-through specimens were sampled at the ganglionic, transitional, and aganglionic zones. Each specimen was stained with calretinin and H&E. Agreement and kappa analysis were done. Results: Fifty-one Hirschsprung’s patients (153 specimens) were included for analysis. There were 37 males (72.5%) and 14 females (27.5%) with a median age at surgery of four months. Thirty-one specimens showed a negative stain for calretinin (aganglionic bowel) and 33 specimens showed no ganglion cells in the H&E stain. One hundred twenty-two specimens in calretinin stain were consistently positive with ganglionic bowel and 120 specimens in the H&E stain showed ganglion cells. Agreement and Cohen’s kappa coefficient were 97.4% and 0.921 (95% confidence interval 0.845 to 0.997), respectively. Disconcordance was found in four specimens. Three out of four were in the transitional zone. One was in the aganglionic zone, which has no muscular layer attached. (Thirty-six specimens had no muscular layer.) Conclusion: Calretinin stain was found to be comparable with the H&E stain and could be used for diagnosis of Hirschsprung’s disease. In rectal suction biopsy specimen in which the muscular layer was not included, both calretinin and H&E can be used. Keywords: Megacolon, Hematoxylin and eosin (H&E), Calretinin, Rectal suction biopsy (RSB), Full thickness biopsy

Diagnostic Accuracy of Combined Acetylcholinesterase Histochemistry and Calretinin Immunohistochemistry of Rectal Biopsy Specimens in Hirschsprung’s Disease

Background. Acetylcholinesterase (AchE) histochemistry has been established as an accurate diagnostic tool for Hirschsprung’s disease (HD). In addition, calretinin immunohistochemistry is also reported as a reliable and adjunctive method to diagnose HD. We investigated the diagnostic value of combined AchE histochemistry and calretinin immunohistochemistry in rectal suction biopsies from HD and non-HD patients. Methods. We retrospectively reviewed 99 rectal suction biopsy specimens including 4 repeat biopsies from 95 patients (34 HD and 61 non-HD). Each specimen was evaluated with hematoxylin-eosin, AchE histochemistry, and calretinin immunohistochemistry. Results. Of 95 patients, only 21 (22.1%) showed some ganglion cells. All 61 non-HD cases revealed no abnormal AchE-positive fibers. Of 34 HD patients, 32 exhibited abnormal AchE fibers, but 2 showed no stained fibers. None of the tissues from the HD patients exhibited calretinin immunoreactivity. Test sensitivity and specificity of AchE histochemistry alone were 93.5% and 100.0%, respectively, while calretinin immunohistochemistry were 100.0% and 85.2%, respectively. Conclusions. AchE histochemistry is a good diagnostic method for HD, if feasible, and a combination of AchE histochemistry and calretinin immunohistochemistry will help increase the accuracy of the diagnosis of HD.