Fluid shear stress-induced IL-8/CXCR signaling in human ovarian cancer cells

Lei Sun, Jirui Wen, Ling Wang, Qiao Wen, Jiang Wu, Mingjiang Bie


Background: Interleukin-8 (IL-8) released from mechanosensitive cancer cells plays a key role in the inflammation and metastasis of solid carcinomas. In this study, we have explored IL-8 and its receptors signal transduction process of human ovarian cancer cells under conditions of FSS.
Methods: After the fluid shear stress was loaded, LightCycler™ system and quantitative sandwich ELISA were employed to assay the IL-8 mRNA expression and protein production, respectively. IL-8 reporter gene pEGFP1-IL8USCS was constructed for determining IL-8 gene transcriptional activation through gene transfer and flow cytometric analysis. NF-κB nuclear translocation was observed by immunocytofluorescent staining. Western blot was used to examine IκB phosphorylation and degradation. RT-PCR, Northern blot and immunofluorescence were used to determine the expression of a cell-surface chemokine receptor CXCR2 at mRNA and protein levels.
Results: IL-8 mRNA expression and protein production had biphasic responses to low shear stress (1.5 dyne/cm2), with the peaks at 1 and 2 h respectively. There was an increase in enhanced green fluorescent protein expression in pEGFP1-IL8USCS-transfected SKOV3 cells subjected to a fluid shear stress of 1.5 dyne/cm2 for 2 h. Following the application of shear stress of 1.5 dyne/cm2, NF-κB p65 became detectable in the cell nuclei, and Phosphorylated IκB in cell lysates increased significantly. CXCR2, which was constitutively present on the surface of SKOV3 cells, increased following exposure to fluid shear stress for 60 min.
Conclusions: Fluid shear stress triggered IL-8/CXCR2 signaling of SKOV3 cells is an early gene activation, and the activation can be mediated through NF-κB. This observation suggested that fluid shear stress-induced IL-8 activation and the downstream signal pathways may have an important contribution to the pathogenesis and development of both inflammation and metastasis of ovarian carcinomas.