Normalisation of surfactant protein -A and -B expression in the lungs of low birth weight lambs by 21 days old

Abstract

Intrauterine growth restriction (IUGR) induced by placental restriction (PR) in the sheep negatively impacts lung and pulmonary surfactant development during fetal life. Using a sheep model of low birth weight (LBW), we found that there was an increase in mRNA expression of surfactant protein (SP)-A, -B and -C in the lung of LBW lambs but no difference in the protein expression of SP-A or -B. LBW also resulted in increased lysosome-associated membrane glycoprotein (LAMP)-3 mRNA expression, which may indicate an increase in either the density of type II Alveolar epithelial cells (AEC) or maturity of type II AECs. Although there was an increase in glucocorticoid receptor (GR) and 11β-hydroxysteroid dehydrogenase (11βHSD)-1 mRNA expression in the lung of LBW lambs, we found no change in the protein expression of these factors, suggesting that the increase in SP mRNA expression is not mediated by increased GC signalling in the lung. The increase in SP mRNA expression may, in part, be mediated by persistent alterations in hypoxia signalling as there was an increase in lung HIF-2α mRNA expression in the LBW lamb. The changes in the hypoxia signalling pathway that persist within the lung after birth may be involved in maintaining SP production in the LBW lamb.

Fig 1

There was higher mRNA expression of SP-A (A), -B (B) and -C (C), but not SP-D (D), in the lungs of the LBW lambs (n = 6; Closed bars) compared with ABW lambs (n = 9; Open bars). There was no difference in the protein expression of SP-A (E), -B (F) and SP-D (G) in the lungs of the LBW lambs (n = 6; Closed bars) compared with ABW lambs (n = 7; Open bars). Representative Western blot images from animals in each treatment group for SP-A (G),–B (H) and–D (J) are presented. MNE, mean normalised expression; ABW, Average birth weight; LBW, Low birth weight; *P<0.05 from ABW.

Fig 2

There was higher mRNA expression of 11βHSD-1 (A) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 8; Open bars). There was no difference in the mRNA expression of 11βHSD-2 (B) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 8; Open bars). There was no difference in the protein expression of 11βHSD-1 (C) and 11βHSD-2 (D) between the lungs of the LBW lambs (n = 6; Closed bars) and ABW lambs (n = 7; Open bars). Representative Western blot images from animals in each treatment group for 11βHSD-1 (E) and 11βHSD-2 (F) are presented. MNE, mean normalised expression; ABW, Average birth weight; LBW, Low birth weight; *P<0.05 from ABW.

Fig 3

There was higher mRNA expression of the GR (A) in the lungs of the LBW lambs (n = 7; Closed bars) compared with the ABW lambs (n = 6; Open bars). However, there was no difference in the protein expression of 94kDa GR-α (B), 91kDa GR-β (C) between the lungs of the LBW lambs (n = 6; Closed bars) and ABW lambs (n = 7; Open bars). There was no difference in the ratio of 91kDa GR-β to 94kDA GR-α protein expression (D) between the lungs of the LBW lambs (Closed bars) and ABW lambs (Open bars). A representative Western blot image from animals in each treatment group for GR (E) is presented. There was no difference in the mRNA expression of MR between the lungs of the LBW lambs (n = 6; Closed bars) and ABW lambs (n = 9; Open bars) (F). MNE, mean normalised expression; ABW, Average birth weight; LBW, Low birth weight; *P<0.05 from ABW.

Fig 4

There was lower mRNA expression of GATA-6 (A) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 9; Open bars). There was no difference in the mRNA expression of TTF-1 (B) in the lungs of the LBW lambs (n = 5; Closed bars) compared with the ABW lambs (n = 8; Open bars). There was no difference in the protein expression of GATA-6 (C) and TTF-1(D) between the lungs of the LBW lambs (n = 6; Closed bars) and ABW lambs (n = 7; Open bars). Representative Western blot images from animals in each treatment group for GATA-6 (E) and TTF-1 (F) are presented. MNE, mean normalised expression; ABW, Average birth weight; LBW, Low birth weight; *P<0.05 from ABW.

Fig 5

There was no difference in the mRNA expression of PHD-1 (A) and PHD-3 (C) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 9; Open bars). There was lower mRNA expression of PHD-2 (B) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 8; Open bars). There was no difference in the protein expression of PHD-1 (D) and PHD-2 (E) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 7; Open bars). Representative Western blot images from animals in each treatment group for PHD-1 (F) and PHD-2 (G) are presented. MNE, mean normalised expression; ABW, Average birth weight; LBW, Low birth weight; *P<0.05 from ABW.

Fig 6

There was no difference in the mRNA expression of HIF-1α (A) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 9; Open bars). There was higher mRNA expression of HIF-2α (B) in the lungs of the LBW lambs (n = 6; Closed bars) compared with the ABW lambs (n = 8; Open bars). MNE, mean normalised expression; ABW, Average birth weight; LBW, Low birth weight; *P<0.05 from ABW.