title: Role of mitofusin-2 in high mobility group box-1 protein-mediated apoptosis of T cells in vitro.
abstract: BACKGROUND
  High mobility group box-1 protein (HMGB1), a ubiquitous nuclear protein, which is recognized as a danger-associated molecular pattern (DAMP) triggering activation of the innate immune system. Previous studies have shown that HMGB1 also plays a role in T cell-mediated immunity, but the effect of HMGB1 on apoptosis of T cells and its precise mechanism remain to be determined.
  
  
  METHODS
  Two kinds of apoptosis assay techniques were used, i.e., Annexin V-FITC conjunction with PI to identify early apoptotic cells, Hoechst 33342 staining for double-stranded DNA to observe nuclear fragmentation or apoptotic body. The activation status of caspase-3, caspase-8, as well as caspase-9 was examined by colorimetric assay. The dynamic changes in intracellular calcium concentration ([Ca(2+)]i) was monitored by flow cytometry. Overexpression of Mfn2 was preformed by lentiviral vector transfection. The mRNA and protein levels of Mfn2 were determined by RT-PCR and Western-blotting.
  
  
  RESULTS
  Treatment of Jurkat T cells with recombinant human HMGB1 (rhHMGB1) causes a significant dose-dependent increase in percentage of apoptotic cells. When T cells are incubated with HMGB1 they express decreased mitochondria fusion-related protein mitofusin-2 (Mfn2) and activate mitochondrial apoptotic pathway via elevation of [Ca(2+)]i, Bax insertion, and activation of caspase. Furthermore, overexpression of Mfn2 ameliorates the apoptosis of T cells induced by HMGB1. This occurs at least partly through Mfn2 keeps Ca(2+) homeostasis in T cells evidenced by monitoring [Ca(2+)]i dynamics.
  
  
  CONCLUSIONS
  HMGB1 can trigger apoptosis of T lymphocytes through mitochondrial death pathway associated with [Ca(2+)]i elevation. Mfn2 plays a pivotal role in this process, and it might be a novel therapeutic target in T cell apoptosis related disorders.
authors.affiliation: Emergency Center, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, P. R. China.
authors.forename: Zong-sheng
authors.initials: ZS
authors.lastname: Wu
authors.forename: Yong-ming
authors.initials: YM
authors.lastname: Yao
authors.forename: Guang-liang
authors.initials: GL
authors.lastname: Hong
authors.forename: Xiu-ping
authors.initials: XP
authors.lastname: Xu
authors.forename: Yao
authors.initials: Y
authors.lastname: Liu
authors.forename: Ning
authors.initials: N
authors.lastname: Dong
authors.forename: Jia-yi
authors.initials: JY
authors.lastname: Zheng
authors.forename: Zhong-qiu
authors.initials: ZQ
authors.lastname: Lu
authors.forename: Guang-ju
authors.initials: GJ
authors.lastname: Zhao
authors.forename: Xiao-mei
authors.initials: XM
authors.lastname: Zhu
authors.forename: Qing-hong
authors.initials: QH
authors.lastname: Zhang
authors.forename: Zhi-yong
authors.initials: ZY
authors.lastname: Sheng
mesh_terms: D017209:Apoptosis / Q000276:immunology*; D020013:Calcium Signaling / Q000276:immunology*; D020169:Caspases / Q000276:immunology; D020558:GTP Phosphohydrolases / Q000276:immunology*; D024243:HMGB1 Protein / Q000276:immunology*; D006801:Humans; D019169:Jurkat Cells; D024101:Mitochondrial Proteins / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology*
chemical_list: D024243:HMGB1 Protein; C585491:HMGB1 protein, human; D024101:Mitochondrial Proteins; D020169:Caspases; D020558:GTP Phosphohydrolases; C470018:MFN2 protein, human