Proceedings of The Physiological Society
University of Edinburgh (2007) Proc Physiol Soc 6, C3
Prokineticin-1 upregulates interleukin-8 expression in placenta.
F. C. Denison1, S. Battersby1, M. Szuber1, M. J. Evans2, H. N. Jabbour1
1. Centre for Reproductive Biology, University of Edinburgh, Edinburgh, Lothian, United Kingdom. 2. Department of Pathology, Royal Infirmary, Edinburgh, Lothian, United Kingdom.
Background: Prokineticin-1 (PK1) is a pleiotrophic peptide whose functions include tissue specific angiogenesis1, vascular permeability2 and haemopoiesis3. Although expressed in third trimester placenta, the cellular localisations of PK1 and prokineticin receptor 1 (PKR1) and signalling pathway of PK1 are not known. In addition, there is a paucity of data regarding gene up-regulation by PK-1. Aim: To characterise PK1 and PKR1 immunolocalisation, expression and signalling in human placentae. Methods: Placentae (n=20) were collected after elective caesarean section at term (>37 weeks gestation) from women with uncomplicated pregnancies. PK1 and PKR1 were immunolocalised by standard immunohistochemical techniques. Extracellular regulated signal kinase -1/2 (ERK1/2) phosphorlyation was detected by Western blotting with signalling pathways being dissected using various inhibitors including YM25480, PP2 and AG1478, specific inhibitors of Gq, c-src and epidermal growth factor receptor (EGFR) kinase, respectively. Placental explants (n=6) were treated with 40nM PK-1 and interleukin-8 (IL-8) mRNA expression detected by taqman PCR. PK1 or PKR1 and IL-8 were colocalised using immunofluorescence and confocal microscopy. Results: PK1 was immunolocalised to endothelium and macrophages in fetal vessels and Hofbauer cells in placental villi. In contrast, PKR1 was predominately localised in syncytial sprouts. ERK-1/2 phosphorylation in placenta was significantly upregulated (5-fold increase; p<0.05) following treatment with PK1 for 30 minutes. Dissection of the upstream signalling pathway by the chemical inhibitors demonstrated that PK1 induced phosphorylation of ERK-1/2 was mediated via c-src and EGFR transactivation. Treatment of placenta with PK1 for 4 hours induced a significant increase in IL-8 expression (3.26+0.45 fold increase above control; p<0.05). IL-8 expression in response to treatment with PK1 was inhibited following coincubation of the tissue with inhibitors of Gq, c-src, EGFR kinase or ERK1/2. Using double immunofluorescence, co-localisation/co-expression of PK-1 or PKR1 and IL-8 was demonstrated in various cellular compartments within the placenta including trophoblast and macrophages. Conclusions: The cellular immunolocalisation of PK1 and PKR1 within placenta and upregulation of IL-8 by PK1 is supportive of PK1 being involved in placental vascular physiology. In addition, expression of PKR1 in syncytial sprouts, which characterise areas of hypoxia and immature villous formation, suggest that another role of PK1 may in mediating trophoblast differentiation in response to hypoxia. More studies are required to establish the role of PK1 in normal placental physiology and in hypoxic pre-eclamptic placentae which are characterised by increased syncytial sprouts and vaso-occlusive lesions. Acknowledgements: This project was funded by a Moray Endowment Fellowship and Action Medical Research. Reference 1 : LeCouter J et al. (2001). Nature 412, 877-884. Reference 2 : Battersby S et al. (2004). J Clin Endocrinol Metab 89,2463-24692. Reference 3 : LeCouter J et al. (2004). Proc Natl Acad Sci U.S.A. 101, 16813-16818
Where applicable, experiments conform with Society ethical requirements