5.5Successful insilico discovery of novel hypoxia regulated genes in pre-eclampsia

IntroductionPre-eclampsia is a pregnancy specific disorder with still unknown aetiology. Placental hypoxia is one of the mechanisms suggested in the pathogenesis of pre-eclampsia. Here, we have used insilico techniques to identify differentially expressed hypoxia genes between normal and pre-eclamptic placenta and validated it through reverse transcription-PCR (rt-QPCR).MethodologyThe National Centre for Biotechnology's UniGene database was found to contain 20 placental cDNA libraries, out of which three are from normal term placenta and one from pre-eclamptic placenta. Each of these libraries contain thousands of ESTs and by using the data-mining tool digital differential display (DDD), genes with altered expression were identified. These genes were classified into major functional groups and relationship to hypoxia explored using Online Mendelian Inheritance in Man and PubMed. 16 women with pre-eclampsia and 16 normal pregnant women were recruited for this study. Placental tissue was collected immediately after delivery and stored at -80 degree C. The mRNA expression of hypoxia regulated genes identified through DDD was measured in the placenta of these women using rt-QPCR.Results and conclusionDDD identified a total of 32 gene clusters to be differentially expressed and six of them were found to be hypoxia regulated. Angiogenin inhibitor, apolipoprotein E, NADH dehydrogenase (ubiquinone) 1, alpha/beta subcomplex, 1 (NDUFAB1) and round about homolog 4 (ROBO4) were the upregulated genes and H19 and growth differentiating factor (GDF15) were the down regulated genes. Rt-QPCR studies showed similar results thus providing a proof that insilico techniques can be used as tools for novel gene discovery in gestational diseases like pre-eclampsia.