Retinoic acid, midkine, and defects of secondary neurulation

Background: Retinoic acid (RA) is necessary for normal differentiation of t he tail bud into the secondary neural tube. Excess RA, however, is teratoge nic and causes neural tube defects (NTDs). The way in which RA modulates se condary neurulation is unclear but probably involves RA-regulated downstrea m genes such midkine (MK), which encodes a growth factor implicated in tail bud mesenchymal-neuroepithelial conversion. Our objective was to determine whether RA-deficiency would produce similar defects and if MK is involved. Methods: Citral, a drug that blocks endogenous RA formation, as well as a n eutralizing antibody, were used to block RA activity in chick embryos. Immu nohistochemistry and in situ hybridization were used to localize RA and MK in the tail bud. Competitive RT-PCR was used to examine the effects of exce ss RA and RA deficiency due to citral on the expression of MK mRNA. Results: Citral-induced NTDs displayed a morphological resemblance to those caused by excess RA. However, citral treatment did not significantly incre ase embryonic mortality, and RA rescue of citral-treated embryos proved uns uccessful. MK mRNA was detected in the differentiating tail bud by in situ hybridization. Competitive RT-PCR showed that excess RA decreased MK expres sion by 60%. Doses of citral that caused a comparable incidence of defects, however, caused only a 25% decrease. Conclusions: The results show that excess RA and RA deficiency both cause d efects of secondary neurulation. While excess RA decreased MK expression, R A deficiency had minimal effects. However, whether or not MK is an intermed iary in the developmental phenomena regulated physiologically or pathologic ally by RA remains to be elucidated. Teratology 62:123-133, 2000. (C) 2000 Wiley-Liss, Inc.