The role of primary myogenic regulatory factors in the developing diaphragmatic muscle in the nitrofen-induced diaphragmatic hernia.
OBJECTIVE
The nitrofen model of congenital diaphragmatic hernia (CDH) is widely used to investigate the pathogenesis of CDH. However, the exact pathomechanism of the diaphragmatic defect is still unclear. Diaphragmatic muscularization represents the last stage of diaphragmatic development. Myogenic differentiation 1 (MyoD) and myogenic factor 5 (Myf5) play a crucial role in muscularization. MyoD(-/-) : Myf5(+/-) mutant mice show reduced diaphragmatic size, whereas MyoD(+/-) : Myf5(-/-) mutants have normal diaphragms. We designed this study to investigate diaphragmatic gene expression of MyoD and Myf5 in the nitrofen CDH model.


METHODS
Pregnant rats received nitrofen or vehicle on day 9 of gestation (D9), followed by cesarean section on D18 and D21. Fetal diaphragms (n = 40) were micro-dissected and divided into CDH group and controls. MyoD and Myf5 mRNA-expression were determined using Real-time PCR. Immunohistochemistry was performed to evaluate protein expression of MyoD and Myf5.


RESULTS
Relative diaphragmatic mRNA expression levels and immunoreactivity of MyoD were decreased in the CDH group on D18 and D21. Myf 5 mRNA and protein expression were not altered in the CDH group.


CONCLUSIONS
This is the first study showing that MyoD expression is selectively decreased in the diaphragm muscle in the nitrofen model of CDH.
National Children's Research Centre, Our Lady's Children's Hospital, Dublin 12, Ireland.
Jens
J
Dingemann
Takashi
T
Doi
Elke
E
Ruttenstock
Prem
P
Puri
D000818:Animals; D000831:Animals, Newborn; D003964:Diaphragm / Q000196:embryology* / Q000378:metabolism; D004195:Disease Models, Animal; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006548:Hernia, Diaphragmatic / Q000139:chemically induced / Q000235:genetics / Q000378:metabolism; D065630:Hernias, Diaphragmatic, Congenital; D007150:Immunohistochemistry; D017570:MyoD Protein / Q000096:biosynthesis / Q000235:genetics*; D051787:Myogenic Regulatory Factor 5 / Q000096:biosynthesis / Q000235:genetics*; D010647:Phenyl Ethers / Q000633:toxicity; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D011270:Pregnancy, Animal; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley
D017570:MyoD Protein; D051787:Myogenic Regulatory Factor 5; D010647:Phenyl Ethers; D012333:RNA, Messenger; C007350:nitrofen
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