Transforming growth factor ?2, but not ?1 and ?3, is critical for early rat lung branching

Mesenchymal-epithelial tissue interactions are critical for lung branching morphogenesis, and polypeptide growth factors are likely involved in these tissue interactions. Transforming growth factorβs (TGFβs) have been implicated in lung development, but their involvement in early lung branching morphogenesis is unclear. In the present study, we investigated the role of the three mammalian TGFβ subtypes (β1, β2, and β3) and their receptors (type III (TβR-III), type II (TβR-II), and two types I (TβR-I), ALK-1 and ALK-5) in early rat lung organogenesis by using an embryonic rat lung explant culture. Transcripts and proteins for all three TGFβs and their receptors were detected during the embryonic period of fetal rat lung development. Inhibition of TGFβ2, but not β1 and β3, with antisense oligonucleotides and neutralizing antibodies resulted in significant inhibition of early lung branching in culture. Addition of minute amounts (≤1 ng/ml) of exogenous TGFβ2, but not β1 and β3, restored the branching of TGFβ2 antisense-treated explants. Higher concentrations of TGFβ2 were inhibitory. BrdU labeling of lung explants was not altered by antisense TGFβ2 treatment, but low concentrations of TGFβ2 increased thymidine uptake by isolated epithelial cells. Fibronectin and metallogelatinase activities of embryonic lung cells were not affected by any TGFβ isoform but TGFβ2 specifically decreased mesenchymal hyaluronan synthesis. Antisense inhibition of ALK-5 and TβR-II showed a similar reduction in early lung branching as observed with antisense TGFβ2. Incubation of lung explants with soluble TβR-II receptors also abrogated lung branching. ALK-1 antisense treatment did not affect early branching. Administration of neither activin A, which can act via ALK-1, nor follistatin, the natural inhibitor of activin, to the explants cultures had any significant effect on lung branching. Antisense inhibition of the activin receptor-II (Act-RII) also did not affect lung branching. These results are consistent with TGFβ2, but not β1 and β3, regulating pattern formation during early rat lung organogenesis. This TGFβ signaling in rat lung branching in vitro appears to be predominantly mediated via the TβR-I(ALK-5)/TβR-II heteromeric complex. Dev Den;217:343–360. © 2000 Wiley-Liss, Inc.