• A gel-free quantitative proteomics analysis of factors released from hypoxic-conditioned placentae.

      Blankley, R T; Robinson, N J; Aplin, J D; Crocker, I P; Gaskell, S J; Whetton, Anthony D; Baker, P N; Myers, J E; Maternal and Fetal Health Research Group, St Mary's Hospital, Manchester, UK. richard.blankley-2@manchester.ac.uk (2010-03)
      Characterizing the protein factors released from placentae during pathogenesis remains a key objective toward understanding preeclampsia and related pregnancy disorders. Gel-free proteomics technologies applied to placental explant-conditioned media offers the potential of identifying these factors. Relative quantification mass spectrometry using isobaric tagging for relative and absolute quantification (iTRAQ) labeling was employed to compare the ''secretome'' between healthy term placental tissue cultured under both normoxic and hypoxic oxygen tensions. Of the 499 proteins identified, 45 were differentially expressed (P < .01 level), including interleukin 8 (IL-8) which was significantly upregulated under hypoxia. Global protein level changes are suggestive of decreased extracellular matrix remodeling under the same conditions. A significant enrichment of soluble liberated placental factors is achieved using this model system. Identifying these changes resulting from hypoxic conditioning is hypothesis generating and may provide new mechanistic insights into preeclampsia.
    • A proof-of-principle gel-free proteomics strategy for the identification of predictive biomarkers for the onset of pre-eclampsia.

      Blankley, R T; Gaskell, S J; Whetton, Anthony D; Dive, Caroline; Baker, P N; Myers, J E; Maternal and Fetal Health Research Group, St Mary's Hospital, University of Manchester, Manchester, UK. richard.blankley-2@manchester.ac.uk (2009-10)
      OBJECTIVE: Progress in the prevention and treatment of women at risk of pre-eclampsia (PE) still remains hindered by the lack of clinical screening tools that can accurately predict which mothers are at risk. The identification and validation of predictive biomarkers is therefore seen as a critical milestone towards improved healthcare provision and the clinical testing of new therapeutic strategies. Gel-free proteomic technologies offer the capability of analysing hundreds of plasma proteins simultaneously, but as yet these methods have not been applied to pregnancy complications. To assess the feasibility of such an approach to plasma biomarker research in pregnancy we have applied the technique to samples from women with PE to gestation-matched controls. SAMPLE: Pooled plasma samples taken at time of disease from women with PE (n = 23) and gestation-matched controls (n = 23). METHODS: Proteomics strategy for relative quantification of proteins using mass spectrometry. RESULTS: We identified several differences, including elevated levels of endoglin, PAPP-A and PSG1 in PE plasma. Increased levels of endoglin were validated using immunoassay analysis of individual plasma samples. CONCLUSIONS: Although at a relatively early stage, this mass spectrometry-based approach shows promise as a tool to identify global protein changes in plasma. The application of these methods to pre-disease samples is the next step in the identification of clinically useful biomarkers.