DIFFERENTIAL DISPLAY OF GENES IN NORMAL AND HYPOPLASTIC FETAL MURINE LUNGS

To evaluate lung development at the level of gene expression, a compar ison was made between normal and hypoplastic murine fetal lungs by usi ng the mRNA differential display technique, We focused on altered gene expressions at gestational day (Gd) 19 in normal and hypoplastic muri ne lungs, Hypoplastic fetal lungs were created by gavaging pregnant mi ce at Gd8 with 25 mg of nitrofen (2,4-dichlorophenyl-p-nitrophenyl eth er) [1], Normal as well as gavaged mice were euthanized by an overdose of halothane at Gd19, and fetuses were removed by laparotomy, Lungs w ere excised and total RNA was extracted from normal and hypoplastic fe tal lungs. Differential display technique was carried out using the RN Aimage kit (GrenHunter Corp., TN). Each reverse transcription and poly merase chain reaction (RT-PCR) was performed using one specific anchor primer H-T11M ((5')HindIII-T(11)A/C/G(3')) and one arbitrary primer. We have used a total of 3 different anchor primers and 24 arbitrary pr imers for each sample, There were 20 differentially expressed cDNA clo nes, either induced, inhibited, enhanced, or reduced in hypoplastic fe tal lungs as compared to normal. Of these, one clone (NL2) with reduce d expression in Gd19 hypoplastic lungs had 100% homology with mouse nu cleosome assembly protein I gene. Another clone (NT5) with induced exp ression in hypoplastic lungs is an unknown gene. Further, analyses of Northern blots of lungs from various gestational ages showed that the expression of NT5 was induced in hypoplastic lungs at Gd18, whereas in normal lungs it was first expressed at the neonatal stage and was inc reasingly expressed into adulthood. There is a single hybridized band, approximately 400 bp long for NT5 message, Dexamethasone induced expr ession of NT5 in normal Gd14 pseudoglandular lungs cultured for 7 days ; however, different growth factors did not, Northern blot hybridizati on of multiple adult mice tissues showed NT5 expression in the lung, i ntestine, and spleen, The thyromimetic action of nitrofen and the inte ractive functional pathways of dexamethasone with T-3 are known. There fore, we suggest that the isolation and characterization of NT5 may pr ovide valuable information on the regulation of lung development, (C) 1998 Academic Press.