Signalling pathways involved in the direct effects of IGFBP-5 on breast epithelial cell attachment and survival.
We have demonstrated previously that IGFBP-5 can confer survival against apoptosis induced by ceramide, C2, or a small synthetic arginine-glycine-aspartic acid (RGD)-containing peptide in a direct manner. The endogenous ceramide-induced pathway is normally counter-balanced by survival signals mediated by sphingosine kinase (SK) and protein kinase C (PKC). In order to investigate whether these pathways are involved in the IGFBP-5 survival effect, we have used inhibitors of SK (N, N-di-methyl sphingosine, DMS) and PKC (chelerythrine chloride, CC). The effect of pre-incubating Hs578T breast cancer cells with IGFBP-5 on cell adhesion or on subsequent cell death induced by C2 or RGD was investigated with and without the presence of DMS or CC. Cell death was determined by trypan blue cell counts and apoptosis confirmed by morphological assessment and flow cytometry. Cell attachment was determined by a cell adhesion assay. The presence of IGFBP-5 significantly inhibited cell death induced by C2 or RGD, compared to the triggers of apoptosis alone (P<0.01 in both cases). In the presence of either IGFBP-5, CC or DMS, there was no significant effect on cell death compared to the control. IGFBP-5 in the presence of either inhibitor resulted in a significant increase in cell death; IGFBP-5 also lost its ability to confer survival on C2 and RGD-induced apoptosis and in contrast significantly increased cell death. In the cell adhesion assay, IGFBP-5 significantly increased cell attachment over basal levels. In the presence of either inhibitor the IGFBP-5 effect on cell adhesion was reversed and cell attachment was reduced to below basal levels. These data suggest that IGFBP-5 promotes the attachment and survival of Hs578T cells by modulating the balance between ceramide and opposing survival signals.
McCaig
Catherine
C
Division of Surgery, Department of Hospital Medicine, Bristol Royal Infirmary, Bristol, BS2 8HW, England, United Kingdom. c_mccaig@hotmail.com
Perks
Claire M
CM
Holly
Jeff M P
JM
D000470:Alkaloids; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D053119:Benzophenanthridines; D001943:Breast Neoplasms; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002518:Ceramides / Q000494:pharmacology; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology; D006801:Humans; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000494:pharmacology*; D009842:Oligopeptides / Q000494:pharmacology; D010617:Phenanthridines / Q000494:pharmacology; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D000099157:Sphingosine Kinase
D000470:Alkaloids; D053119:Benzophenanthridines; D002518:Ceramides; D004791:Enzyme Inhibitors; D018975:Insulin-Like Growth Factor Binding Protein 5; D009842:Oligopeptides; D010617:Phenanthridines; D017853:Phosphotransferases (Alcohol Group Acceptor); D011493:Protein Kinase C; D013110:Sphingosine; D000099157:Sphingosine Kinase; C047981:arginyl-glycyl-aspartic acid; C016299:chelerythrine; C061800:N,N-dimethylsphingosine