Dual role of autophagy in stress-induced cell death in rheumatoid arthritis synovial fibroblasts

Masaru Kato, Caroline Ospelt, Renate E. Gay, Steffen Gay, Kerstin Klein*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

132 Scopus citations

Abstract

Objective: To investigate the role of autophagy in the regulation of cell death in rheumatoid arthritis synovial fibroblasts (RASFs). Methods: RASFs and osteoarthritis synovial fibroblasts (OASFs) were treated with thapsigargin (TG), an inducer of endoplasmic reticulum (ER) stress, and MG132, a proteasome inhibitor. Then, 3-methyladenine was used as an autophagy inhibitor and bafilomycin A1 as a lysosome inhibitor. Polyubiquitinated proteins, p62, and autophagy induction were evaluated by immunoblotting, immunofluorescence microscopy, and immunohistochemistry, respectively. OASFs were transfected with small interfering RNA targeting autophagy-linked FYVE protein (ALFY). Cell death was evaluated by flow cytometry and a caspase 3 activity assay. Results: In RASFs, the induction of autophagy by TG and MG132 was increased compared to that in OASFs. Whereas autophagy promoted a caspase 3-independent induction of cell death under ER stress, autophagy had a protective role in apoptosis induced by proteasome inhibition. Treatment of RASFs with 3-methyladenine blocked TG-induced cell death. ER stress induced a strong accumulation of p62-positive polyubiquitinated protein aggregates, accompanied by the formation of large vacuoles in RASFs but not OASFs. Furthermore, TG-induced p62 protein expression was increased, whereas TG-induced ALFY expression was reduced, in RASFs compared to OASFs. ALFY knockdown promoted the accumulation of p62, the formation of polyubiquitinated protein aggregates, and cell death. Conclusion: Our data provide the first evidence of a dual role of autophagy in the regulation of death pathways in RASFs. A reduced expression of ALFY and the formation of p62-positive polyubiquitinated protein aggregates promote cell death in RASFs under severe ER stress.

Original languageEnglish
Pages (from-to)40-48
Number of pages9
JournalArthritis and Rheumatology
Volume66
Issue number1
DOIs
StatePublished - 2014/01

ASJC Scopus subject areas

  • Immunology and Allergy
  • Rheumatology
  • Immunology

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