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Eur J Pediatr Surg 2018; 28(01): 109-114
DOI: 10.1055/s-0037-1604022
Original Article
Georg Thieme Verlag KG Stuttgart · New York

Decreased Expression of Integrin Subunits α3, α6, and α8 in the Branching Airway Mesenchyme of Nitrofen-Induced Hypoplastic Lungs

Toshiaki Takahashi
1   National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin, Ireland
,
Florian Friedmacher
1   National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin, Ireland
,
Julia Zimmer
1   National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin, Ireland
2   Department of Pediatric Surgery, Medizinische Hochschule Hannover Zentrum Chirurgie, Hannover, Germany
,
Prem Puri
1   National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin, Ireland
› Author Affiliations
Further Information

Publication History

19 April 2017

23 May 2017

Publication Date:
12 July 2017 (online)

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Abstract

Introduction Pulmonary hypoplasia (PH), characterized by smaller lung size and reduced airway branching, remains a major cause of neonatal mortality in newborns with congenital diaphragmatic hernia (CDH). Integrin-mediated cell–matrix interactions play an essential role in the fetal lung mesenchyme by stimulating branching morphogenesis. Mice lacking integrin subunits α3 (Itga3) and α6 (Itga6) exhibit severe PH. Furthermore, Itga8-knockout mice show defective airway branching, suggesting that Itga3, Itga6, and Itga8 are crucial for fetal lung development. We hypothesized that expression of Itga3, Itga6, and Itga8 is decreased in the branching airway mesenchyme of hypoplastic rat lungs in the nitrofen-induced CDH model.

Materials and Methods Time-mated rats received nitrofen or vehicle on gestational day 9 (D9). Fetuses were sacrificed on D15, D18, and D21, and dissected lungs were divided into control and nitrofen-exposed specimens (n = 12 per time-point and group, respectively). Pulmonary gene expression of Itga3, Itga6, and Itga8 was analyzed by quantitative real-time polymerase chain reaction. Immunofluorescence double-staining for Itga3, Itga6, and Itga8 was combined with the mesenchymal marker Fgf10 to evaluate protein expression and localization in branching airway tissue.

Results Relative mRNA expression of Itga3, Itga6, and Itga8 was significantly decreased in lungs of nitrofen-exposed fetuses on D15, D18, and D21 compared with controls. Confocal laser scanning microscopy showed markedly diminished immunofluorescence of Itga3, Itga6, and Itga8 mainly in mesenchymal cells surrounding branching airways of nitrofen-exposed fetuses on D15, D18, and D21 compared with controls.

Conclusion Decreased expression of Itga3, Itga6, and Itga8 in the pulmonary mesenchyme may lead to disruptions in airway branching morphogenesis, thus contributing to PH in the nitrofen-induced CDH model.

Keywords

integrin - lung branching - pulmonary hypoplasia - congenital diaphragmatic hernia - nitrofen
 

  • References

  • 1 McGivern MR, Best KE, Rankin J. , et al. Epidemiology of congenital diaphragmatic hernia in Europe: a register-based study. Arch Dis Child Fetal Neonatal Ed 2015; 100 (02) F137-F144
    CrossrefPubMedGoogle Scholar
  • 2 McHoney M. Congenital diaphragmatic hernia, management in the newborn. Pediatr Surg Int 2015; 31 (11) 1005-1013
    Google Scholar
  • 3 Jeanty C, Kunisaki SM, MacKenzie TC. Novel non-surgical prenatal approaches to treating congenital diaphragmatic hernia. Semin Fetal Neonatal Med 2014; 19 (06) 349-356
    CrossrefPubMedGoogle Scholar
  • 4 Tovar JA. Congenital diaphragmatic hernia. Orphanet J Rare Dis 2012; 7: 1
    CrossrefPubMedGoogle Scholar
  • 5 Keijzer R, Puri P. Congenital diaphragmatic hernia. Semin Pediatr Surg 2010; 19 (03) 180-185
    CrossrefPubMedGoogle Scholar
  • 6 Kotecha S, Barbato A, Bush A. , et al. Congenital diaphragmatic hernia. Eur Respir J 2012; 39 (04) 820-829
    CrossrefPubMedGoogle Scholar
  • 7 Sabharwal AJ, Davis CF, Howatson AG. Post-mortem findings in fetal and neonatal congenital diaphragmatic hernia. Eur J Pediatr Surg 2000; 10 (02) 96-99
    Article in Thieme ConnectPubMedGoogle Scholar
  • 8 Maeda Y, Davé V, Whitsett JA. Transcriptional control of lung morphogenesis. Physiol Rev 2007; 87 (01) 219-244
    CrossrefPubMedGoogle Scholar
  • 9 Metzger RJ, Klein OD, Martin GR, Krasnow MA. The branching programme of mouse lung development. Nature 2008; 453 (7196): 745-750
    CrossrefPubMedGoogle Scholar
  • 10 Roth-Kleiner M, Post M. Similarities and dissimilarities of branching and septation during lung development. Pediatr Pulmonol 2005; 40 (02) 113-134
    CrossrefPubMedGoogle Scholar
  • 11 Warburton D, Bellusci S, De Langhe S. , et al. Molecular mechanisms of early lung specification and branching morphogenesis. Pediatr Res 2005; 57 (5 Pt 2): 26R-37R
    CrossrefPubMedGoogle Scholar
  • 12 Friedmacher F, Gosemann JH, Fujiwara N, Takahashi H, Hofmann A, Puri P. Expression of Sproutys and SPREDs is decreased during lung branching morphogenesis in nitrofen-induced pulmonary hypoplasia. Pediatr Surg Int 2013; 29 (11) 1193-1198
    CrossrefPubMedGoogle Scholar
  • 13 Friedmacher F, Fujiwara N, Hofmann AD, Takahashi H, Gosemann JH, Puri P. Expression of Eya1 and Six1 is decreased in distal airways of rats with experimental pulmonary hypoplasia. J Pediatr Surg 2014; 49 (02) 301-304
    CrossrefPubMedGoogle Scholar
  • 14 Hynes RO. Integrins: versatility, modulation, and signaling in cell adhesion. Cell 1992; 69 (01) 11-25
    CrossrefPubMedGoogle Scholar
  • 15 Schuger L, Skubitz AP, O'Shea KS, Chang JF, Varani J. Identification of laminin domains involved in branching morphogenesis: effects of anti-laminin monoclonal antibodies on mouse embryonic lung development. Dev Biol 1991; 146 (02) 531-541
    CrossrefPubMedGoogle Scholar
  • 16 Mollard R, Dziadek M. A correlation between epithelial proliferation rates, basement membrane component localization patterns, and morphogenetic potential in the embryonic mouse lung. Am J Respir Cell Mol Biol 1998; 19 (01) 71-82
    CrossrefPubMedGoogle Scholar
  • 17 De Arcangelis A, Mark M, Kreidberg J, Sorokin L, Georges-Labouesse E. Synergistic activities of alpha3 and alpha6 integrins are required during apical ectodermal ridge formation and organogenesis in the mouse. Development 1999; 126 (17) 3957-3968
    PubMedGoogle Scholar
  • 18 Benjamin JT, Gaston DC, Halloran BA, Schnapp LM, Zent R, Prince LS. The role of integrin alpha8beta1 in fetal lung morphogenesis and injury. Dev Biol 2009; 335 (02) 407-417
    CrossrefPubMedGoogle Scholar
  • 19 Noble BR, Babiuk RP, Clugston RD. , et al. Mechanisms of action of the congenital diaphragmatic hernia-inducing teratogen nitrofen. Am J Physiol Lung Cell Mol Physiol 2007; 293 (04) L1079 –L1087
    CrossrefPubMedGoogle Scholar
  • 20 Eastwood MP, Russo FM, Toelen J, Deprest J. Medical interventions to reverse pulmonary hypoplasia in the animal model of congenital diaphragmatic hernia: A systematic review. Pediatr Pulmonol 2015; 50 (08) 820-838
    CrossrefPubMedGoogle Scholar
  • 21 Montedonico S, Nakazawa N, Puri P. Congenital diaphragmatic hernia and retinoids: searching for an etiology. Pediatr Surg Int 2008; 24 (07) 755-761
    CrossrefPubMedGoogle Scholar
  • 22 Herriges M, Morrisey EE. Lung development: orchestrating the generation and regeneration of a complex organ. Development 2014; 141 (03) 502-513
    CrossrefPubMedGoogle Scholar
  • 23 Miura T. Modeling lung branching morphogenesis. Curr Top Dev Biol 2008; 81: 291-310
    PubMedGoogle Scholar
  • 24 Roth-Kleiner M, Post M. Genetic control of lung development. Biol Neonate 2003; 84 (01) 83-88
    CrossrefPubMedGoogle Scholar
  • 25 Short K, Hodson M, Smyth I. Spatial mapping and quantification of developmental branching morphogenesis. Development 2013; 140 (02) 471-478
    CrossrefPubMedGoogle Scholar
  • 26 Howe A, Aplin AE, Alahari SK, Juliano RL. Integrin signaling and cell growth control. Curr Opin Cell Biol 1998; 10 (02) 220-231
    CrossrefPubMedGoogle Scholar
  • 27 Mercurio AM, Shaw LM. Laminin binding proteins. BioEssays 1991; 13 (09) 469-473
    CrossrefPubMedGoogle Scholar
  • 28 Bellusci S, Grindley J, Emoto H, Itoh N, Hogan BL. Fibroblast growth factor 10 (FGF10) and branching morphogenesis in the embryonic mouse lung. Development 1997; 124 (23) 4867-4878
    PubMedGoogle Scholar
  • 29 Weaver M, Dunn NR, Hogan BL. Bmp4 and Fgf10 play opposing roles during lung bud morphogenesis. Development 2000; 127 (12) 2695-2704
    PubMedGoogle Scholar
  • 30 Teramoto H, Yoneda A, Puri P. Gene expression of fibroblast growth factors 10 and 7 is downregulated in the lung of nitrofen-induced diaphragmatic hernia in rats. J Pediatr Surg 2003; 38 (07) 1021-1024
    CrossrefPubMedGoogle Scholar
  • 31 De Langhe SP, Sala FG, Del Moral PM. , et al. Dickkopf-1 (DKK1) reveals that fibronectin is a major target of Wnt signaling in branching morphogenesis of the mouse embryonic lung. Dev Biol 2005; 277 (02) 316-331
    CrossrefPubMedGoogle Scholar
  • 32 Roman J. Fibronectin and fibronectin receptors in lung development. Exp Lung Res 1997; 23 (02) 147-159
    CrossrefPubMedGoogle Scholar
  • 33 Sakai T, Larsen M, Yamada KM. Fibronectin requirement in branching morphogenesis. Nature 2003; 423 (6942): 876-881
    CrossrefPubMedGoogle Scholar
  • 34 Miner JH, Cunningham J, Sanes JR. Roles for laminin in embryogenesis: exencephaly, syndactyly, and placentopathy in mice lacking the laminin alpha5 chain. J Cell Biol 1998; 143 (06) 1713-1723
    CrossrefPubMedGoogle Scholar
  • 35 Miner JH, Patton BL, Lentz SI. , et al. The laminin alpha chains: expression, developmental transitions, and chromosomal locations of alpha1-5, identification of heterotrimeric laminins 8-11, and cloning of a novel alpha3 isoform. J Cell Biol 1997; 137 (03) 685-701
    CrossrefPubMedGoogle Scholar