Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Raffler, G.
Impact of intra-amniotically administered recombinant VEGF on rodent lung development in case of nitrofen induced congenital diaphragmatic hernia
Humanmedizin; [ Diplomarbeit ] ; 2014. pp. 69
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- Autor*innen der Med Uni Graz:
- Betreuer*innen:
- Klaritsch Philipp
- Ruttenstock Elke Maria
- Altmetrics:
- Abstract:
- Aim: Children with congenital diaphragmatic hernia (CDH) are subject to a significant morbidity and mortality due to lung hypoplasia and pulmonary hypertension. Vascular endothelial growth factor (VEGF-a) is a permeable, signal protein that has mitogenic effects and especially targets endothelial cells. Several studies have shown that VEGF-a plays an important role for angiogenesis and vasculogenesis as well as lung development in general. The aim of this study was to evaluate the effect of intra-amniotically injected VEGF-a on the development of hypoplastic lungs in the nitrofen model of CDH. Methods: On day 9 pregnant rats were exposed to 100mg nitrofen dissolved in olive oil. Distilled water (Placebo) or 50 ?l (microliter) VEGF-a (treatment group) was injected into the amniotic fluid on day 19 and day 20. On day 21 a caesarean section was performed to deliver the fetal rats. Fetuses with confirmed CDH were weighed and divided into VEGF-a (n=10) and Placebo (n=6) groups. Right and left lungs in each group were harvested for further analysis. Real time reverse transcription polymerase chain reaction (qRT-PCR) was performed to analyze the gene expression of the proliferation marker Ki67, the tissue markers insulin-like growth factor receptor 1 und 2 (IGF-1r und IGF-2r), podoplanin (PDPN), as well as the organogenesis markers bone morphogenetic protein-4 (BMP-4), fibroblast growth factor (FGF-10), thyroid transcription factor (TTF-1). Markers for vasculogenesis (VEGF-a, kinase insert domain-containing receptor (KDR), fetal liver tyrosinase (Flt-1), platelet endothelial cell adhesion molecule-1 (Pecam-1), plasminogen activator tissue (Plat)) were also used to perform qRT-PCR in the right fetal lungs. Left fetal lungs were analyzed by immunochemistry (IHC) for Ki67, FGF-10, PDPN, TTF-1, a-smooth muscle actin (SMA) and VEGF-a. Data are expressed as median ± standard deviation of fold changes and analyzed using parametric or non-parametric tests. Ethical approval number: 66.010/0011II/10b/2010. Results: When compared with the Placebo group, VEGF-a treated fetuses had significantly down-regulated gene expression levels of FGF-10 (0.9864 ± 1.685 vs. - 0.5917 ± 0.2979; p< 0.05), IGF-2r (0.2089 ± 1.385 vs. - 1.236 ± 0.4043; p< 0.05) and Plat (- 0.8012 ± 2.045 vs. - 2.859 ± 0.4052; p< 0.05) gene expression levels compared to placebo group. The gene expression levels of Ttf1 were highly significantly down regulated in the VEGF-a treated fetuses (- 1.444 ± 1.334, - 3.233 ± 0.3874, p< 0.01). Protein expression of Ki67, FGF-10, podoplanin, TTF-1, SMA and VEGF-a showed no significant difference in comparison to the placebo group. Conclusion: Intra-amniotic administration of VEGF-a appears to have a suppressive effect on lung branching, proliferation and lung growth of hypoplastic lungs in the nitrofen model of CDH. The clinical application of VEGF-a cannot be recommended, at least in the used dose and route of administration