PLoS ONE (Jan 2021)

IL-10 and class 1 histone deacetylases act synergistically and independently on the secretion of proinflammatory mediators in alveolar macrophages.

  • Brent A Stanfield,
  • Todd Purves,
  • Scott Palmer,
  • Bruce Sullenger,
  • Karen Welty-Wolf,
  • Krista Haines,
  • Suresh Agarwal,
  • George Kasotakis

DOI
https://doi.org/10.1371/journal.pone.0245169
Journal volume & issue
Vol. 16, no. 1
p. e0245169

Abstract

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IntroductionAnti-inflammatory cytokine IL-10 suppresses pro-inflammatory IL-12b expression after Lipopolysaccharide (LPS) stimulation in colonic macrophages, as part of the innate immunity Toll-Like Receptor (TLR)-NF-κB activation system. This homeostatic mechanism limits excess inflammation in the intestinal mucosa, as it constantly interacts with the gut flora. This effect is reversed with Histone Deacetylase 3 (HDAC3), a class I HDAC, siRNA, suggesting it is mediated through HDAC3. Given alveolar macrophages' prominent role in Acute Lung Injury (ALI), we aim to determine whether a similar regulatory mechanism exists in the typically sterile pulmonary microenvironment.MethodsLevels of mRNA and protein for IL-10, and IL-12b were determined by qPCR and ELISA/Western Blot respectively in naïve and LPS-stimulated alveolar macrophages. Expression of the NF-κB intermediaries was also similarly assessed. Experiments were repeated with AS101 (an IL-10 protein synthesis inhibitor), MS-275 (a selective class 1 HDAC inhibitor), or both.ResultsLPS stimulation upregulated all proinflammatory mediators assayed in this study. In the presence of LPS, inhibition of IL-10 and/or class 1 HDACs resulted in both synergistic and independent effects on these signaling molecules. Quantitative reverse-transcriptase PCR on key components of the TLR4 signaling cascade demonstrated significant diversity in IL-10 and related gene expression in the presence of LPS. Inhibition of IL-10 secretion and/or class 1 HDACs in the presence of LPS independently affected the transcription of MyD88, IRAK1, Rela and the NF-κB p50 subunit. Interestingly, by quantitative ELISA inhibition of IL-10 secretion and/or class 1 HDACs in the presence of LPS independently affected the secretion of not only IL-10, IL-12b, and TNFα, but also proinflammatory mediators CXCL2, IL-6, and MIF. These results suggest that IL-10 and class 1 HDAC activity regulate both independent and synergistic mechanisms of proinflammatory cytokine/chemokine signaling.ConclusionsAlveolar macrophages after inflammatory stimulation upregulate both IL-10 and IL-12b production, in a highly class 1 HDAC-dependent manner. Class 1 HDACs appear to help maintain the balance between the pro- and anti-inflammatory IL-12b and IL-10 respectively. Class 1 HDACs may be considered as targets for the macrophage-initiated pulmonary inflammation in ALI in a preclinical setting.