SQSTM1-dependent autophagic degradation of PKM2 inhibits the production of mature IL1B/IL-1β and contributes to LIPUS-mediated anti-inflammatory effect

Autophagy. 2020 Jul;16(7):1262-1278. doi: 10.1080/15548627.2019.1664705. Epub 2019 Sep 22.

Abstract

Synovitis is implicated in the pathology of osteoarthritis (OA) and significantly contributes to the development of OA. As a noninvasive physical therapy, low-intensity pulsed ultrasound (LIPUS) has been reported to possess anti-inflammatory effect in recent years. However, the role of LIPUS on synovitis of OA and the underlying mechanisms are little known. The present study showed that LIPUS ameliorated synovial inflammation in destabilization of the medial meniscus (DMM) mouse model and air pouch model, and alleviated pain gait patterns of DMM mouse. LIPUS dramatically inhibited the production of mature IL1B/IL-1β (interleukin 1 beta) in vitro and in vivo. In addition, LIPUS upregulated the macroautophagy/autophagy level as well as accelerated the formation of an SQSTM1 (sequestosome1)-PKM (pyruvate kinase, muscle) complex in the lipopolysaccharide (LPS)-adenosine triphosphate (ATP)-treated macrophages. Besides, LIPUS downregulated the level of PKM2 in LPS-ATP-treated macrophages, which could be reversed by SQSTM1 knockdown. In brief, the present study for the first time demonstrates that LIPUS inhibits the production of mature IL1B partially via SQSTM1-dependent autophagic degradation of PKM2 in LPS-ATP-treated macrophages, which may further ameliorate the synovial inflammation and gait patterns in animal models. Our data provide new clues for the treatments of synovitis and other inflammatory diseases using LIPUS.

Abbreviations: 3-MA: 3-methyladenene; ATG7: autophagy-related 7; ATP: adenosine triphosphate; BafA1: bafilomycin A1; BMDMs: bone marrow derived macrophages; CHX: cycloheximide; DMM: destabilization of the medial meniscus; ELISA: enzyme-linked immunosorbent assay; GFP: green fluorescent protein; IL1B/IL-1β: interleukin 1 beta; LIPUS: low-intensity pulsed ultrasound; LIR: LC3-interacting region; LPS: lipopolysaccharide; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MDP: muramyl dipeptide; NFKB/NF-κB: nuclear factor kappa B; NLRP3: NLR family, pyrin domain containing 3; OA: osteoarthritis; PKM/PKM2: pyruvate kinase M1/2; PMA: phorbol-12-myristate-13-acetate; PYCARD/ASC; PYD and CARD domain containing; RFP: red fluorescent protein; siRNAs: small interfering RNAs; SQSTM1: sequestosome 1; TEM: transmission electron microscopy.

Keywords: Autophagy; PKM/PKM2; SQSTM1; inflammasome; low-intensity pulsed ultrasound; macrophage; synovitis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / pharmacology
  • Animals
  • Anti-Inflammatory Agents / pharmacology*
  • Autophagy* / drug effects
  • Disease Models, Animal
  • Gait / drug effects
  • Humans
  • Inflammation / pathology
  • Interleukin-1beta / metabolism*
  • Lipopolysaccharides / pharmacology
  • Macrophages / drug effects
  • Macrophages / metabolism
  • Menisci, Tibial / pathology
  • Menisci, Tibial / physiopathology
  • Mice
  • Mice, Inbred C57BL
  • Models, Biological
  • Pain / pathology
  • Pain / physiopathology
  • Proteolysis* / drug effects
  • Pyruvate Kinase / metabolism*
  • RAW 264.7 Cells
  • Sequestosome-1 Protein / metabolism*
  • Synovial Membrane / pathology
  • THP-1 Cells
  • Ultrasonic Waves

Substances

  • Anti-Inflammatory Agents
  • Interleukin-1beta
  • Lipopolysaccharides
  • Sequestosome-1 Protein
  • Adenosine Triphosphate
  • Pkm protein, mouse
  • Pyruvate Kinase

Grants and funding

This work was supported by (1) Special Funds for Major State Basic Research Program of China (973 program) (No. 2014CB942900); (2) National Natural Science Foundation of China (No. 81530071, 81871817).