Cytokine-induced apoptosis in epithelial HT-29 cells is independent of nitric oxide formation. Evidence for an interleukin-13-driven phosphatidylinositol 3-kinase-dependent survival mechanism.
A combination of the pro-inflammatory cytokines interleukin (IL)-1alpha, interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha induces nitric oxide synthase mRNA expression and nitric oxide (NO) generation in the human colon carcinoma cell line HT-29. This can be inhibited by pretreatment with IL-13 via a phosphatidylinositol (PI) 3-kinase-dependent mechanism (Wright, K., Ward, S. G., Kolios, G., and Westwick, J. (1997) J. Biol. Chem. 272, 12626-12633). Since NO has been implicated in regulating mechanisms leading to cell death, while activation of PI 3-kinase-dependent signaling cascades are thought to be involved with promoting cell survival events, we have investigated the outcome of these cytokine treatments on apoptosis and cell survival of HT-29 cells. Initiation of apoptosis can be achieved by the combinations of IFN-gamma/TNF-alpha, IFN-gamma/CD95, IL-1alpha/IFN-gamma, and IL-1alpha/IFN-gamma/TNF-alpha to varying extents. Induction of apoptotic markers by HT-29 cells in response to cytokine treatment is not dependent on NO production. Pretreatment with IL-13 protects against IL-1alpha/IFN-gamma/TNF-alpha- and IFN-gamma/TNF-alpha- as well as IFN-gamma/CD95-induced (but not IL-1alpha/IFN-gamma-induced) cell death. In addition, IFN-gamma/TNF-alpha and IL-1alpha/IFN-gamma/TNF-alpha stimulate activation of caspase-8 and caspase-3, which IL-13 pretreatment was able to partially inhibit and delay. IL-13 also stimulates activation of the major PI 3-kinase effector, protein kinase B. The PI 3-kinase inhibitors wortmannin and LY294002 inhibit IL-13 stimulation of protein kinase B as well as the cell survival effects of IL-13. These data demonstrate that cytokine-induced apoptosis of HT-29 cells is NO-independent and that the activation of a PI 3-kinase-dependent signaling cascade by IL-13 is a key signal responsible for the inhibition of apoptosis.
Wright
K
K
Department of Pharmacy and Pharmacology, Bath University, Claverton Down, Bath, BA2 7AY, United Kingdom.
Kolios
G
G
Westwick
J
J
Ward
S G
SG
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D017209:Apoptosis; D061945:Caspase Inhibitors; D002867:Chromones / Q000494:pharmacology; D016207:Cytokines / Q000494:pharmacology*; D004789:Enzyme Activation; D006146:Guanidines / Q000494:pharmacology; D019073:HT29 Cells; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D018793:Interleukin-13 / Q000494:pharmacology; D007413:Intestinal Mucosa / Q000187:drug effects*; D009025:Morpholines / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D052247:Nitric Oxide Synthase Type II; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011940:Receptor Aggregation; D015398:Signal Transduction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D000077191:Wortmannin; D019014:fas Receptor / Q000378:metabolism
D000590:Amino Acid Chloromethyl Ketones; D000730:Androstadienes; D061945:Caspase Inhibitors; D002867:Chromones; D016207:Cytokines; D006146:Guanidines; D007371:Interferon-gamma; D007375:Interleukin-1; D018793:Interleukin-13; D009025:Morpholines; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D014409:Tumor Necrosis Factor-alpha; D000077191:Wortmannin; D019014:fas Receptor; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C496319:NOS2 protein, human; C004479:pimagedine