Is there a role for the IHH gene in Hirschsprung’s disease? M.-M. GARCIA-BARCELO ´ ,* W.-S. LEE,* M.-H. SHAM,  V. C.-H. LUI* & P. K.-H. TAM*à *Department of Surgery, Division of Paediatric Surgery, University of Hong Kong Medical Center, Queen Mary Hospital, Hong Kong SAR, China  Department of Biochemistry, The University of Hong Kong, Hong Kong SAR, China University of Hong Kong, Hong Kong SAR, China àGenome Research Centre, The University of Hong Kong, Hong Kong SAR, China University of Hong Kong, Hong Kong SAR, China Abstract Hirschsprung disease (HSCR) is character- ized by the absence of ganglion cells along a variable length of the intestine. HSCR has a complex genetic aetiology with 50% of the patients unexplained by mutations in the major HSCR genes. The Ihh gene is involved in the development of the enteric nervous system (ENS) and Ihh mutant mice present with a phenotype reminiscent of HSCR. The requirement of Ihh signalling for the proper development of the ENS, together with the evidence presented by the Ihh murine model, prompted us to investigate the involvement of the human IHH gene in HSCR. Sequence analysis revealed seven single nucleotide polymorphisms, six of which were new. Allele and haplotype frequencies were compared between cases and controls, and, among the cases, between genders, between different phenotypes, and between patients with different mutation status in the main HSCR genes. Despite the involvement of IHH in the devel- opment of the ENS, IHH is not a major gene in HSCR. Nevertheless, as the manifestation of the HSCR phe- notype is genetic background dependent, polymorphic loci should be tested simultaneously to characterize gene–gene interaction. The involvement of IHH in other intestinal anomalies should be investigated. Keywords development, enteric nervous system, Hirschsprung, Indian hedgehog, single nucleotide polymorphisms. INTRODUCTION Hirschsprung’s disease (HSCR) is a developmental disorder characterized by the absence of ganglion cells in the nerve plexuses of the lower digestive tract. HSCR phenotype is variable and can be classified into two groups: short segment aganglionosis (SSA) (80% of the cases), which includes patients with aganglionosis as far as the rectosigmoid junction, and long segment aganglionosis (LSA) (20% of the cases), which includes patients with aganglionosis beyond the rectosigmoid junction. 1 Aganglionosis is attributed to a disorder of the enteric nervous system (ENS) in which ganglion cells fail to innervate the lower gastrointestinal tract during embryonic development. The condition pre- sents in the neonatal period as a failure to pass meconium, chronic severe constipation, colonic dis- tention, secondary electrolyte disturbances and, some- times, enterocolitis and bowel perforation. 1 The estimated population incidence is 1/5000 live births although this is a representative value. The highest incidence is in Asian populations (2.8 per 10 000 live births) and the lowest in Hispanics (1 per 10 000 live births). 1 The male : female (M : F) ratio is approxi- mately 4 : 1 for SSA-HSCR patients and approximately 1 : 1 for LSA-HSCR patients. 1 Approximately 20% of HSCR cases are familial. The recurrence risks to sibs of SSA-HSCR probands ranges between 1.5 and 3.3% while risks to sibs of LSA-HSCR probands varies from 2.9 to 17.6%. 1 HSCR is frequently associated with chromosomal abnormalities, with other neurodevelop- mental disorders such as Waardenburg syndrome type 4, and with a variety of additional isolated anomalies and syndromes. 1,2 HSCR has a complex genetic aetiol- ogy with many studies indicating the receptor tyrosine kinase gene (RET) as the major susceptibility gene for HSCR. Mutations in the RET gene account for up to 50% of the familial cases and anywhere between 7 and 35% of the sporadic cases and lack genotype–phenotype Address for correspondence Professor Paul Kwong-Hang Tam, Department of Surgery, Division of Paediatric Surgery, University of Hong Kong Medical Center, Queen Mary Hospital, Hong Kong SAR, China. Tel.: +(852) 2855 4850; fax: +(852) 2817 3155; e-mail: paultam@hkucc.hku.hk Received: 2 May 2003 Accepted for publication: 17 June 2003 Merce ` GARCIA-BARCELO ´ and Wing-shan LEE contributed equally to this work. Neurogastroenterol Motil (2003) 15, 663–668 Ó 2003 Blackwell Publishing Ltd 663