Perforin and Fas induced by IFNgamma and TNFalpha mediate beta cell death by OT-I CTL.
Direct interaction between auto-reactive CTL and specific peptide-MHC class I complexes on pancreatic beta cells is critical in mediating beta cell destruction in type I diabetes. We used mice with genetic modifications in three major pathways used by CTL, perforin, Fas and pro-inflammatory cytokines to assess the relative contribution of these mechanisms to beta cell death. In vitro-activated ovalbumin (OVA)-specific CTL, from OT-I TCR-transgenic mice, specifically killed transgenic beta cells expressing OVA (from RIP-mOVA mice) in a 16-h cytotoxicity assay. Perforin-deficient CTL had a reduced ability to kill OVA-expressing islets in vitro (22.1 +/- 3.8%) compared with wild-type CTL (71.4 +/- 4.6%). Fas-deficient islets were only slightly protected from wild-type CTL but were completely protected from the residual killing observed with perforin-deficient CTL. Residual cytotoxicity in perforin-deficient CTL was also prevented by overexpression of SOCS-1, which blocks multiple cytokine signaling pathways. It was also prevented by pre-incubation with anti-tumor necrosis factor-alpha (anti-TNFalpha) antibody or by blocking IFNgamma responsiveness through expressing a dominant negative IFNgamma receptor. Perforin-deficient CTL produced IFNgamma and TNFalpha that was shown to directly induce islet Fas expression during the assays. This suggests that Fas-deficiency, SOCS-1 overexpression and blockade of IFNgamma and TNFalpha all protect beta cells from residual cytotoxicity of perforin-deficient CTL by blocking Fas upregulation. These findings indicate that wild-type CTL destroy antigen-expressing islets via a perforin-dependent mechanism. However, in the absence of perforin, the Fas/FasL pathway provides an alternative mechanism dependent on islet cell Fas upregulation by cytokines IFNgamma and TNFalpha.
McKenzie
Mark D
MD
St Vincent's Institute, Fitzroy, Melbourne, Australia.
Dudek
Nadine L
NL
Mariana
Lina
L
Chong
Mark Mw
MM
Trapani
Joseph A
JA
Kay
Thomas Wh
TW
Thomas
Helen E
HE
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D001323:Autoantibodies / Q000276:immunology; D002352:Carrier Proteins / Q000276:immunology; D016923:Cell Death / Q000187:drug effects / Q000276:immunology; D003922:Diabetes Mellitus, Type 1 / Q000276:immunology*; D053222:Fas Ligand Protein; D005805:Genes, MHC Class I / Q000276:immunology; D050417:Insulin-Secreting Cells / Q000276:immunology*; D007371:Interferon-gamma / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000172:deficiency / Q000276:immunology*; D051379:Mice; D008822:Mice, Transgenic; D010455:Peptides / Q000276:immunology; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D017471:Receptors, Interferon / Q000276:immunology; D012097:Repressor Proteins / Q000276:immunology; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D000071222:Suppressor of Cytokine Signaling 1 Protein; D050826:Suppressor of Cytokine Signaling Proteins / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology*; D048069:Tumor Necrosis Factors; D015854:Up-Regulation / Q000187:drug effects / Q000276:immunology; D019014:fas Receptor / Q000276:immunology*; D000096743:Interferon gamma Receptor
D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D002352:Carrier Proteins; D053222:Fas Ligand Protein; D007371:Interferon-gamma; D008562:Membrane Glycoproteins; D010455:Peptides; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D017471:Receptors, Interferon; D012097:Repressor Proteins; D000071222:Suppressor of Cytokine Signaling 1 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D014409:Tumor Necrosis Factor-alpha; D048069:Tumor Necrosis Factors; D019014:fas Receptor; D000096743:Interferon gamma Receptor; C505658:Fasl protein, mouse; C491590:Socs1 protein, mouse