Expression of the TGF-β signaling machinery in lungs of neonatal and adult rats.

<p>Changes in the expression of genes encoding components of the TGF-β signaling machinery as assessed by real-time RT-PCR at days P1 and P70; n = 8–15 for each bar. The significance for each bar is indicated by p values, IUGR vs. CO; two-tailed Mann-Whitney test. A: Expression of genes encoding the TGF-β receptors <i>tgfbr1</i>, <i>tgfbr2</i> and <i>tgfbr3</i> at P1 (white bar) and P70 (striped bar). B: Expression of genes encoding the regulatory <i>smad2</i>, <i>smad3</i> and <i>smad4</i> at days P1 (white bar) and P70 (striped bar). C: Expression of genes encoding the inhibitory <i>smad7</i>, <i>smurf2</i> and smad anchor for receptor activation (<i>sara</i>) at days P1 (white bar) and P70 (striped bar).</p

Effect of IUGR on the expression of extracellular matrix (ECM) proteins and modulators of the ECM in neonatal rat lungs.

<p>Expression of TGF-β-regulated genes encoding elastin (Eln), tenascin N (Ten), collagens (Coll) I, Coll III, and Fibrillin (A), and genes of ECM modulators including matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitor of metalloproteinases (TIMP)-1, and TIMP-2 (B) in lungs extracted at day P1 from neonatal rats after IUGR and control rats. The mRNA expression, illustrated as relative fold induction, was assessed by real-time PCR. The control group was normalized to 1 as indicated by a scattered line; n = 15 for each group. The significance for each bar is indicated by p values, IUGR vs. Co; two-tailed Mann-Whitney test.</p

Primer pairs and TaqMan probes used.

<p>*TIMP-1, TIMP-2 and MMP-2 were detected by TaqMan realtime-PCR analysis.</p

Effect of IUGR on apoptosis in lungs of rats at days P1 and P70.

<p>Apoptosis is assessed by cleaved caspase-3 and cleaved fragment of Poly (ADP-ribose) polymerase (PARP). A: Representative immunoblots illustrating the expression of cleaved and total caspase-3, fragments of PARP and total PARP in lung homogenates of rats with IUGR and without IUGR (Co) at <i>day</i> P1 (A) and P70 (B). The β-actin served as loading control; n = 4–6 for each bar.</p

Body weight, respiration and physiological lung parameters of IUGR rats.

<p>A: Body weight (g) and relative lung weight (lung weight/body weight) of rats after IUGR (white bars) and control rats (black bars) at days P1 and P70; n = 8–15 for each bar. B: Architectural changes in lung structure were evident in hematoxylin and eosin-stained lung sections from IUGR rats and control rats at day P70. Measurement of septal thickness (µm) and mean linear intercept (MLI; µm) in IUGR rats and control rats (CO) at day P70; n = 6–10 for each bar. C: Assessment of respiratory system compliance by whole body plethysmography in IUGR rats (white bars) and in the control group (CO; black bars) at P70. n = 15–17 for each bar. D: Expression pattern of genes encoding surfactant protein A (SP-A), SP-C, and SPD. IUGR rats (white bars) and control group (black bars). n = 6–15 for each bar. The significance for each bar is indicated by p values, IUGR vs. Co, n.s. = not significant; two-tailed Mann-Whitney test.</p

Effect of IUGR on the expression of TGF-β1 in lungs of neonatal and adult rats.

<p>A: Expression of genes encoding TGF-β1 and TGF-β-inducible plasminogen activator inhibitor-1 (PAI-1) during late lung development (day P1). The mRNA expression was assessed by quantitative real-time PCR. The control group was normalized to 1 as indicated by a scattered line; n = 15 for each group. The significance for each bar is indicated by p values, IUGR vs. Co; two-tailed Mann-Whitney test. B: representative immunoblots illustrating expression of TGF-β1 in lungs extracted at day P1 and P70 from rats with and without IUGR. β-actin served as loading control. Immunoblot data were quantified for both day P1 and P70; n = 6 for each bar. The significance for each bar is indicated by p values, IUGR vs. Co; two-tailed Mann-Whitney test.</p

IUGR alters expression and phosphorylation of Smad proteins in rats.

<p>A: Representative immunoblots illustrating the expression of TGF-β-specific Smad2, Smad3, the co-Smad, Smad4, and the inhibitory Smad, Smad7, in lungs extracted at days P1 and P70 from rats with and without IUGR. β-actin served as loading control. Immunoblot data were quantified for Smad4 and Smad7 for both days P1 and P70 (Co as black bar, and IUGR as white bar); n = 4–6 for each bar. The significance for each bar is indicated by p values, IUGR vs. CO; two-tailed Mann-Whitney test. B: The expression of active TGF-β signaling components in lung homogenates of rats with and without IUGR was analyzed by immunoblotting of phosphorylated (p) and total Smad2 and Smad3. β-actin served as loading control. Immunoblot data were quantified for pSmad2 and pSmad3 for both days P1 and P70 (Co as black bar, and IUGR as white bar); n = 4–6 for each bar. The significance for each bar is indicated by p values, IUGR vs. CO; two-tailed Mann-Whitney test. C: Immunhistochemical localization and expression pattern of pSmad2 and pSmad3 in lungs of rats with IUGR (right column) and without IUGR (left column). A–F: representative fields illustrating the expression and localization of pSmad2 in bronchi (A–D) and in the alveoli (E–F) of lungs extracted on day P70. G–L: representative fields illustrating the expression and localization of pSmad3 in bronchi (G–J) and in the alveoli (K–L) of lungs extracted on day P70. M–N: negative control.</p

The role of transforming growth factor (TGF)-β signaling in lung disease subsequent to intrauterine growth restriction (IUGR).

<p>A proposed model depicting the effects of decreased TGF-β signaling during the development of IUGR-associated lung disease is shown. ECM - extracellular matrix.</p