title: Pro-inflammatory cytokines enhance ERAD and ATF6α pathway activity in salivary glands of Sjögren's syndrome patients.
abstract: Salivary gland (SG) acinar-cells are susceptible to endoplasmic reticulum (ER) stress related to their secretory activity and the complexity of synthesized secretory products. SGs of Sjögren's syndrome patients (SS)-patients show signs of inflammation and altered proteostasis, associated with low IRE1α/XBP-1 pathway activity without avert increases in apoptosis. Acinar-cells may avoid apoptosis by activation of the ATF6α pathway and ER-associated protein degradation (ERAD). The aim of this study was to evaluate the role of pro-inflammatory cytokines in ATF6α pathway/ERAD activation and cell viability in labial salivary glands (LSG) of SS-patients. In biopsies from SS-patients increased ATF6α signaling pathway activity, as evidenced by generation of the ATF6f cleavage fragment, and increased expression of ERAD machinery components, such as EDEM1, p97, SEL1L, gp78, UBE2J1, UBE2G2, HERP and DERLIN1, were observed compared to controls. Alternatively, for pro- (active-caspase-3) and anti-apoptotic (cIAP2) markers no significant difference between the two experimental groups was detected. Increased presence of ATF6f and ERAD molecules correlated significantly with increased expression of pro-inflammatory cytokines. These observations were corroborated in vitro in 3D-acini treated with TNF-α and/or IFN-γ, where an increase in the expression and activation of the ATF6α sensor and ERAD machinery components was detected under ER stress conditions, while changes in cell viability and caspase-3 activation were not observed. Cytokine stimulation protected cells from death when co-incubated with an ERAD machinery inhibitor. Alternatively, when cytokines were eliminated from the medium prior to ERAD inhibition, cell death increased, suggesting that the presence of pro-inflammatory cytokines in the medium is essential to maintain cell viability. In conclusion, the ATF6α pathway and the ERAD machinery are active in LSG of SS-patients. Both were also activated by TNF-α and IFN-γ in vitro in 3D-acini and aided in preventing apoptosis. IFN-γ levels were elevated in SS-patients and UPR responses triggered in vitro by this cytokine closely matched those observed in LSG from SS-patients, suggesting that cytokines may induce ER stress.
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: María-José
authors.initials: MJ
authors.lastname: Barrera
authors.affiliation: Departamento de Reumatología, Clínica INDISA, Santiago, Chile.
authors.forename: Sergio
authors.initials: S
authors.lastname: Aguilera
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Isabel
authors.initials: I
authors.lastname: Castro
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Juan
authors.initials: J
authors.lastname: Cortés
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Verónica
authors.initials: V
authors.lastname: Bahamondes
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile; Center for Molecular Studies of the Cell (CEMC), Advanced Center for Chronic Diseases (ACCDiS), Faculty of Medicine, Universidad de Chile, Santiago, Chile.
authors.forename: Andrew F G
authors.initials: AFG
authors.lastname: Quest
authors.affiliation: Escuela Dental, Facultad de Ciencias, Universidad Mayor, Santiago, Chile.
authors.forename: Claudio
authors.initials: C
authors.lastname: Molina
authors.affiliation: Escuela Dental, Facultad de Ciencias, Universidad Mayor, Santiago, Chile.
authors.forename: Sergio
authors.initials: S
authors.lastname: González
authors.affiliation: Programa de Inmunología, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Marcela
authors.initials: M
authors.lastname: Hermoso
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Ulises
authors.initials: U
authors.lastname: Urzúa
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile.
authors.forename: Cecilia
authors.initials: C
authors.lastname: Leyton
authors.affiliation: Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas (ICBM), Facultad de Medicina, Universidad de Chile, Santiago, Chile. Electronic address: jgonzale@med.uchile.cl.
authors.forename: María-Julieta
authors.initials: MJ
authors.lastname: González
mesh_terms: D051702:Activating Transcription Factor 6 / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000293:Adolescent; D000328:Adult; D017209:Apoptosis / Q000276:immunology / Q000528:radiation effects; D015153:Blotting, Western; D053148:Caspase 3 / Q000276:immunology / Q000378:metabolism; D016207:Cytokines / Q000276:immunology* / Q000378:metabolism / Q000494:pharmacology; D060746:Endoplasmic Reticulum-Associated Degradation / Q000235:genetics / Q000276:immunology*; D004789:Enzyme Activation / Q000187:drug effects / Q000276:immunology; D005260:Female; D015870:Gene Expression / Q000276:immunology; D006801:Humans; D007150:Immunohistochemistry; D018836:Inflammation Mediators / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008875:Middle Aged; D011506:Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012469:Salivary Glands / Q000276:immunology* / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D012859:Sjogren's Syndrome / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D055815:Young Adult
chemical_list: C497111:ATF6 protein, human; D051702:Activating Transcription Factor 6; D016207:Cytokines; C486654:EDEM1 protein, human; D018836:Inflammation Mediators; D008565:Membrane Proteins; D011506:Proteins; C117384:SEL1L protein, human; D053148:Caspase 3
keywords: Cytokines; ERAD; Endoplasmic reticulum stress; Salivary glands; Sjögren's syndrome; UPR