A Novel Strategy for TNF-Alpha Production by 2-APB Induced Downregulated SOCE and Upregulated HSP70 in O. tsutsugamushi-Infected Human Macrophages

PLoS One. 2016 Jul 29;11(7):e0159299. doi: 10.1371/journal.pone.0159299. eCollection 2016.


Orientia (O.) tsutsugamushi-induced scrub typhus is endemic across many regions of Asia and the Western Pacific, where an estimated 1 million cases occur each year; the majority of patients infected with O. tsutsugamushi end up with a cytokine storm from a severe inflammatory response. Previous reports have indicated that blocking tumor necrosis factor (TNF)-α reduced cell injury from a cytokine storm. Since TNF-α production is known to be associated with intracellular Ca2+ elevation, we examined the effect of store-operated Ca2+ entry (SOCE) inhibitors on TNF-α production in O. tsutsugamushi-infected macrophages. We found that 2-aminoethoxydiphenyl borate (2-APB), but not SKF96365, facilitates the suppression of Ca2+ mobilization via the interruption of Orai1 expression in O. tsutsugamushi-infected macrophages. Due to the decrease of Ca2+ elevation, the expression of TNF-α and its release from macrophages was repressed by 2-APB. In addition, a novel role of 2-APB was found in macrophages that causes the upregulation of heat shock protein 70 (HSP70) expression associated with ERK activation; upregulated TNF-α production in the case of knockdown HSP70 was inhibited with 2-APB treatment. Furthermore, elevated HSP70 formation unexpectedly did not help the cell survival of O. tsutsugamushi-infected macrophages. In conclusion, the parallelism between downregulated Ca2+ mobilization via SOCE and upregulated HSP70 after treatment with 2-APB against TNF-α production was found to efficiently attenuate an O. tsutsugamushi-induced severe inflammatory response.

MeSH terms

  • Animals
  • Boron Compounds / pharmacology*
  • Down-Regulation / drug effects*
  • HSP70 Heat-Shock Proteins / metabolism*
  • Humans
  • Imidazoles / pharmacology
  • Intracellular Calcium-Sensing Proteins
  • Macrophages / drug effects*
  • Macrophages / metabolism
  • Macrophages / microbiology
  • Membrane Proteins / metabolism*
  • Mice
  • Orientia tsutsugamushi / pathogenicity*
  • Tumor Necrosis Factor-alpha / biosynthesis*


  • Boron Compounds
  • HSP70 Heat-Shock Proteins
  • Imidazoles
  • Intracellular Calcium-Sensing Proteins
  • Membrane Proteins
  • SARAF protein, human
  • Tumor Necrosis Factor-alpha
  • 2-aminoethoxydiphenyl borate
  • 1-(2-(3-(4-methoxyphenyl)propoxy)-4-methoxyphenylethyl)-1H-imidazole

Grants and funding

This work was supported by the slow aging program, “Aim for the Top Universities Grant” (KMU-TP104G00, KMU-TP104G01, and KMU-TP104G04 [Ming-Wei Lin received] & KMU-TP104D04 [Tohru Yoshioka received]) at Kaohsiung Medical University, and the Ministry of Science and Technology of Taiwan, MOST, 104-2314-B-037-003 (Yaw-Bin Huang received) and 104-2314-B-037-060 (Ching-Ying Wu received). A part of this funding was supported by Glyen-Po Chen (Tohru Yoshioka received).