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THU0026 Osm is more effective than il-6 at inducing endomt of human dermal microvascular cells
  1. K Nevin1,
  2. L Stawski2,
  3. M Feeney1,
  4. N Wisniacki1,
  5. M Trojanowska2
  1. 1Immuno-Inflammation Therapeutic Area Unit, GlaxoSmithKline, Stevenage, United Kingdom
  2. 2Arthritis Centre, Boston University, Boston, United States

Abstract

Background Oncostatin-M (OSM) and interleukin-6 (IL-6) are members of the IL-6 superfamily and signal via glycoprotein 130 (gp130). OSM signals with either the type I receptor complex, gp130/ Leukemia Inhibitory Factor Receptor (LIFR), or the type II receptor complex, gp130/OSM receptor (OSMR), whilst IL-6 signals via gp130 with the IL-6 receptor (IL-6R) or by trans signalling with soluble IL-6R (sIL6-R). Endothelial cells (ECs) express both gp130 and OSMR [Brown TJ et al 1991], however it is unclear whether ECs express IL-6R [Romano M et al 1997; Nilsson MB et al 2005]. Endothelial to mesenchymal transition (EndoMT) is the phenotypic transition of ECs into mesenchymal cells where ECs lose their specific EC markers, detach from the endothelial layer and initiate the expression of mesenchymal cell products. EndoMT is associated with vascular dysfunction, one of the early manifestations of systemic sclerosis. The role of OSM and IL-6 in EndoMT has not yet been fully elucidated.

Objectives To determine the effect of OSM and IL-6/sIL-6R on microvascular EC migration, proliferation and EndoMT.

Methods Human dermal microvascular ECs (HDMECs) were treated with human recombinant proteins OSM (1–100 ng/mL), IL-6 (10–100 ng/mL) and sIL-6R (10–100 ng/mL). Cell migration and proliferation were measured with Live-Cell imaging system over 50 hours and analysed using a two-way ANOVA. Secretion of Collagen type I protein was measured at 48 hours by western blot analysis of media supernatant from HDMEC cultures. Changes in VE-Cadherin and F-actin expression were examined by immunofluorescence over 72 hours. Gene expression was measured at 3 hours using quantitative RT-PCR analysis and analysed by Student's paired T-Test.

Results OSM and IL-6, with or without IL-6R, significantly increased (P<0.001, n=3 donors) HDMEC migration and proliferation and secretion of extracellular matrix (ECM) protein Collagen I compared to the control group. OSM, but not IL-6 with or without sIL-6R, reduced expression of EC marker VE-Cadherin and increased expression of elongated F-actin stress fibres (n=2 donors). OSM significantly affected (P<0.05, n=3 donors) expression of EndoMT genes SNAIL1, SNAIL2, SNAIL3 and TWIST [Figure 1] and ECM genes MMP1, MMP2 TIMP1 and TIMP2 compared to the control. Many of the gene changes in response to OSM were further augmented by co-stimulation with sIL-6R. IL-6 with or without sIL-6R only significantly affected (P<0.05, n=3 donors) EndoMT genes SNAIL1 and SNAIL3 and ECM gene TIMP1.

Conclusions OSM induced a stronger EndoMT phenotype in HDMECs in comparison to IL-6, suggesting that OSM is capable of initiating EndoMT activity in microvascular cells. The augmented effects observed for OSM with sIL-6R also suggests that OSM is capable of binding the sIL-6R and initiating signalling in HDMECs in vitro.

References

  1. Brown TJ, Rowe JM, Liu JW and Shoyab M. Regulation of IL-6 expression by oncostatin M. J Immunol 1991:147:2175–2180.

  2. Nilsson MB, Langley RR and Fidler IJ. Interleukin-6, secreted by human ovarian carcinoma cells, is a potent proangiogenic cytokine. Cancer Res 2005:65:10794–10800.

  3. Romano M, Sironi M, Toniatti C, Polentarutti N, Fruscella P, Ghezzi P, Faggioni R, Luini W, van Hinsbergh V, Sozzani S, Bussolino F, Poli V, Ciliberto G, and Mantovani A. Role of IL-6 and its soluble receptor in induction of chemokines and leukocyte recruitment. Immunity 1997:6:315–325.

References

Disclosure of Interest None declared

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