Pediatric Surgery International

, Volume 26, Issue 10, pp 1017–1023 | Cite as

HoxB2, HoxB4 and Alx4 genes are downregulated in the cadmium-induced omphalocele in the chick model

  • Takashi Doi
  • Prem Puri
  • John Bannigan
  • Jennifer Thompson
Original Article



In the chick embryo, administration of cadmium (Cd) induces omphalocele phenotype. HoxB2 and HoxB4, expressed in cell types that contribute to ventral body wall (VBW) formation, act together to mediate proper closure of the VBW, involving a key downstream transcription factor, Alx4. HoxB2 and HoxB4 knockout mice display VBW defects with specific downregulation of Alx4 gene expression, while homozygous Alx4 knockouts show omphalocele phenotype. Although the earliest histological changes in the Cd chick model occur commencing at 4H post treatment, the exact timing and molecular mechanism by which Cd acts is still unclear. We hypothesized that HoxB2, HoxB4 and Alx4 genes are downregulated during the critical timing of very early embryogenesis in the Cd-induced omphalocele chick model.


After 60H incubation, chick embryos were harvested at 1H, 4H and 8H after treatment with saline or Cd and divided into controls and Cd group (n = 24 for each group). RT–PCR was performed to investigate the gene expression of HoxB2, HoxB4 and Alx4 and statistically analyzed (significance was accepted at p < 0.05). Immunohistochemical staining was also performed to evaluate the protein expression/distribution of HoxB2, HoxB4 and Alx4 in the chick embryo.


The expression levels of HoxB2, HoxB4 and Alx4 gene at 4H were significantly downregulated in the Cd group as compared to controls, whereas there were no significant differences at the other time points. Immunoreactivity of HoxB2, HoxB4 and Alx4 at 4H is also markedly decreased in the ectoderm and the dermomyotome in the Cd chick model as compared to controls.


Downregulation of HoxB2, HoxB4 and Alx4 expression during the narrow window of early embryogenesis may cause omphalocele in the Cd chick model by interfering with molecular signaling required for proper VBW formation. Furthermore, these results support the concept that HoxB2, HoxB4 and Alx4 genes work together to mediate proper VBW formation.


Cadmium Omphalocele HoxB2 HoxB4 Alx4 


  1. 1.
    Axt R, Quijano F, Boos R et al (1999) Omphalocele and gastroschisis; prenatal diagnosis and perinatal management. A case analysis of the years 1989–1997 at the Department of Obstetrics and Gynecology, University of Homburg/Saar. Eur J Obstet Gynecol Reprod Biol 87(1):47–54CrossRefPubMedGoogle Scholar
  2. 2.
    Barisic I, Clementi M, Hausler R et al (2001) Evaluation of prenatal ultrasound diagnosis of fetal abdominal wall defects by 19 European registries. Ultrasound Obstet Gynecol 18(4):309–316CrossRefPubMedGoogle Scholar
  3. 3.
    Thompson JM, Bannigan JG (2001) The effects of cadmium on formation of the ventral body wall in chick embryos and their prevention by zinc pre-treatment. Teratology 64(2):87–97CrossRefPubMedGoogle Scholar
  4. 4.
    Thompson J, Hipwell E, Loo HV et al (2005) Effects of cadmium on cell death and cell proliferation in chick embryos. Reprod Toxicol 20(4):539–548CrossRefPubMedGoogle Scholar
  5. 5.
    Thompson JM, Bannigan JG (2007) Omphalocele induction in the chick embryo by administration of cadmium. J Pediatr Surg 42(10):1703–1709CrossRefPubMedGoogle Scholar
  6. 6.
    Webster WS, Messerle K (1980) Changes in the mouse neuroepithelium associated with cadmium-induced neural tube defects. Teratology 21:79–88CrossRefPubMedGoogle Scholar
  7. 7.
    Messerle K, Webster WS (1982) The classification and development of cadmium-induced limb defects in mice. Teratology 25:61–70CrossRefPubMedGoogle Scholar
  8. 8.
    Sunderman FW Jr, Plowman MC, Hopfer SM (1992) Teratogenicity of cadmium chloride in the South African frog, Xenopus laevis. IARC Sci Publ 118:249–256PubMedGoogle Scholar
  9. 9.
    Doi T, Puri P, Bannigan J et al (2008) Downregulation of ROCK-I and ROCK-II gene expression in the cadmium-induced ventral body wall defect chick model. Pediatr Surg Int 24:1297–1301CrossRefPubMedGoogle Scholar
  10. 10.
    Doi T, Puri P, Bannigan J et al (2010) Disruption of calreticulin-mediated cellular adhesion signaling in the cadmium-induced omphalocele in the chick model. Pediatr Surg Int 26:91–95CrossRefPubMedGoogle Scholar
  11. 11.
    Doi T, Puri P, Bannigan J et al (2010) Disruption of non-canonical wnt/Ca2+ pathway in the cadmium-induced omphalocel in the chick model. J Pediatr Surg (in press)Google Scholar
  12. 12.
    Doi T, Puri P, Bannigan J et al (2010) Msx1 and Msx2 gene expression is downregulated in the cadmium-induced omphalocele in the chick model. J Pediatr Surg (in press)Google Scholar
  13. 13.
    Doi T, Puri P, Bannigan J et al (2010) The role of transforming growth factor-Beta 2 and 3 in formation of ventral body wall in the cadmium induced omphalocele chick model. J Pediatr Surg (in press)Google Scholar
  14. 14.
    Barrow JR, Capecchi MR (1996) Targeted disruption of the Hoxb-2 locus in mice interferes with expression of Hoxb-1 and Hoxb-4. Development 122:3817–3828PubMedGoogle Scholar
  15. 15.
    Manley NR, Barrow JR, Zhang T et al (2001) Hoxb2 and Hoxb4 act together to specify ventral body wall formation. Dev Biol 237:130–144CrossRefPubMedGoogle Scholar
  16. 16.
    Qu S, Niswender KD, Ji Q et al (1997) Polydactyly and ectopic ZPA formation in Alx-4 mutant mice. Development 124:3999–4008PubMedGoogle Scholar
  17. 17.
    Hamburger V, Hamilton HL (1951) A series of normal stages in the development of the chick embryo. J Morphol 88:49–92CrossRefGoogle Scholar
  18. 18.
    Dugan JD Jr, Lawton MT, Glaser B et al (1991) A new technique for explantation and in vitro cultivation of chicken embryos. Anat Rec 229:125–128CrossRefPubMedGoogle Scholar
  19. 19.
    Sadler TW (2005) Embryology of neural tube development. Am J Med Genet Part C Semin Med Genet 135C:2–8CrossRefPubMedGoogle Scholar
  20. 20.
    Brewer S, Williams T (2004) Finally, a sense of closure? Animal models of human ventral body wall defects. Bioessays 26:1307–1321CrossRefPubMedGoogle Scholar
  21. 21.
    Sadler TW, Feldkamp ML (2008) The embryology of body wall closure: relevance to gastroschisis and other ventral body wall defects. Am J Med Genet C Semin Med Genet 148C:180–185CrossRefPubMedGoogle Scholar
  22. 22.
    Burke AC, Nowicki JL (2003) A new view of patterning domains in the vertebrate mesoderm. Dev Cell 4:159–165CrossRefPubMedGoogle Scholar
  23. 23.
    Beverdam A, Brouwer A, Reijnen M et al (2001) Severe nasal clefting and abnormal embryonic apoptosis in Alx3/Alx4 double mutant mice. Development 128:3975–3986PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  • Takashi Doi
    • 1
    • 2
  • Prem Puri
    • 1
    • 2
  • John Bannigan
    • 2
  • Jennifer Thompson
    • 2
  1. 1.The Children’s Research CentreOur Lady’s Children’s HospitalDublin 12Ireland
  2. 2.School of Medicine and Medical Science, Conway Institute of Biomolecular and Biomedical ResearchUniversity College DublinDublinIreland

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