1. Academic Validation
  2. Expression and regulation of alarmin cytokine IL-1α in human retinal pigment epithelial cells

Expression and regulation of alarmin cytokine IL-1α in human retinal pigment epithelial cells

  • Exp Eye Res. 2018 Jul;172:10-20. doi: 10.1016/j.exer.2018.03.015.
Zong-Mei Bian 1 Matthew G Field 2 Susan G Elner 1 Victor M Elner 1
Affiliations

Affiliations

  • 1 Department of Ophthalmology, University of Michigan, Ann Arbor, MI, 48105, United States.
  • 2 Department of Ophthalmology, University of Michigan, Ann Arbor, MI, 48105, United States. Electronic address: [email protected].
Abstract

Human retinal pigment epithelial (hRPE) cells play important immune-regulatory roles in a variety of retinal pathologic processes, including the production of inflammatory cytokines that are essential mediators of the innate immune response within the ocular microenvironment. The pro-inflammatory "alarmin" cytokine IL-1α has been implicated in both infectious and non-infectious retinal diseases, but its regulation in the retina is poorly understood. The purpose of this study was to elucidate the expression and regulation of IL-1α within hRPE cells. To do this, IL-1α mRNA and protein in hRPE cells was assessed by RT-PCR, qPCR, ELISA, Western blot, and immunofluorescence following treatment with a variety of stimuli and inhibitors. ER stress, LPS, IL-1β, and TLR2 activation all significantly increased intracellular IL-1α protein. Increasing intracellular calcium synergized both LPS- and Pam3CSK4-induced IL-1α protein production. Accordingly, blocking calcium signaling and calpain activity strongly suppressed IL-1α protein expression. Significant but more moderate inhibition occurred following blockage of TLR4, caspase-4, or Caspase-1. Neutralizing Antibodies to IL-1β and TLR2 partially eliminated LPS- and TLR2 ligand Pam3CSK4-stimulated IL-1α protein production. IFN-β induced caspase-4 expression and activation, and also potentiated LPS-induced IL-1α expression, but IFN-β alone had no effect on IL-1α protein production. Interestingly, all inhibitors targeting the PI3K/Akt pathway, with the exception of Ly294002, strongly increased IL-1α protein expression. This study improves understanding of the complex mechanisms regulating IL-1α protein expression in hRPE cells by demonstrating that TLR4 and TLR2 stimulation and exposure to IL-1β, ER stress and intracellular calcium all induce hRPE cells to produce intracellular IL-1α, which is negatively regulated by the PI3K/Akt pathway. Additionally, the non-canonical inflammasome pathway was shown to be involved in LPS-induced hRPE IL-1α expression through caspase-4 signaling.

Keywords

Caspase-4; IL-1α; Interleukin-1α; Non-canonical inflammasome; RPE; Retinal pigment epithelium; TLR2; TLR4.

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