Defining the role of paracrine growth factors in pulmonary development is likely to elucidate the pathogenesis of neonatal lung disease and offer avenues of therapeutic manipulation. The three transforming growth factor beta (TGF-beta) isoforms are important mediators of epithelial- mesenchymal interactions in the developing lung. Of the two major classes of TGF-beta receptors, one, the Type II receptor (TGF-betaIIR), is known to directly transduce TGF-beta signalling via an intrinsic protein kinase activity. Subclasses of Type II receptors exhibit varying affinities for individual TGF-beta isoforms, and the cellular response to TGF-beta probably depends on the receptor affinities as well as the TGF-beta isoforms present. This project hypotheses that changes in the expression and affinity of TGF-betaIIRs mediate events in lung development. The study will focus on the saccular stage of lung development, which is when severely premature infants are delivered. The hypothesis will be addressed by the following aims:
Aim 1 : Determine the sites, times, and cell types in which TGF-betaIIRs are transcribed and expressed in the late gestation lung. To meet this goal, reverse transcriptase polymerase chain reaction RNAase protection, in situ hybridization, and immunocytochemistry will be performed on fetal mouse lungs of representative gestational ages.
Aim 2 : Determine if TGF-betaIIR affinity for TGF-beta isoforms varies in a time and/or tissue-specific manner. The affinities of cell-specific TGF-betaIIR will be determined by dissociating fetal mouse lungs at representative ages, culturing the epithelial and mesenchymal cell populations, and performing competitive binding assays on the isolated cell populations.
Aim 3 : Define the development events mediated by TGF-betaIIR. TGF- betaIIR expression will be inhibited using antisense oligonucleotides in saccular stage murine lung explant cultures. TGF-betaIIR cDNA will also be selectively overexpressed in Type II pneumocytes using the surfactant protein C (SP-C) promotor in transgenic mice. Resulting distortions in lung differentiation may be directly ascribed to the loss or the cell- specific overexpression of TGF-betaIIR function. This project will be conducted in a state-of-the-art facility at the USC Center of Craniofacial Molecular Biology under the sponsorship of Dr. David Warburton, a leading investigator of paracrine growth factors in the developing lung.
| Lee, M K; Zhao, J; Smith, S M et al. (1998) The Shc 66 and 46 kD isoforms are differentially downregulated at parturition in the fetal mouse lung. Pediatr Res 44:850-9 |
| Lee, M; Hwang, C; Lee, J et al. (1997) TGF-beta isoforms differentially attenuate EGF mitogenicity and receptor activity in fetal lung mesenchymal cells. Am J Physiol 273:L374-81 |
| Zhao, J; Bu, D; Lee, M et al. (1996) Abrogation of transforming growth factor-beta type II receptor stimulates embryonic mouse lung branching morphogenesis in culture. Dev Biol 180:242-57 |
