Differential Regulation of IL-1β and IL-6 Release in Murine Macrophages

Our plain-English summary of Differential Regulation of IL-1β and IL-6 Release in Murine Macrophages, by S. Hirano, Q. Zhou, A. Furuyama, et al (Inflammation, 2017).

The study

• Authors: S. Hirano, Q. Zhou, A. Furuyama, S. Kanno
• Journal: Inflammation
• Published: 2017;40(6):1933-1943
• Cited by: 68 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Asbestos and silica (exogenous danger) and adenosine triphosphate (ATP, endogenous danger-signaling molecule) synergistically increase IL-1β release from endotoxin-primed macrophage, which is mediated by NOD-like receptor protein 3 (NLRP3) inflammasome. However, the conversion of pro-IL-1β to its active form seems to depend on the macrophage cell types. In the present study, bone marrow-derived macrophages (BMM) and three murine macrophage cell lines, J774.1, J774A.1, and RAW264.7 were exposed to ATP or fibrous titanium dioxide (FTiO2) in the presence or absence of lipopolysaccharide (LPS), and the concentrations of IL-1β and IL-6 in both cell lysates and in the culture media were measured by immunoblotting to differentiate active form of IL-1β from pro-IL-1β. IL-1β release was synergistically increased when the cells were exposed to both LPS and ATP or FTiO2, while IL-6 was readily released by LPS alone. IL-1β released into the culture medium was pro-IL-1β in J774.1 and RAW264.7, and most of the pro-IL-1β remained inside the cells. In contrast, the active form of IL-1β was released together with pro-IL-1β from J774A.1 and BMM after the co-stimulation. J774A.1 and BMM express apoptosis-associated speck-like protein contains a carboxyl-terminal CARD (ASC) while J774.1 and RAW264.7 do not or only faintly express ASC, and accordingly, caspase-1, which converts pro-IL-1β to its active form, is activated only in J774A.1 and BMM. Collectively, the canonical inflammasome pathway is not activated in J774.1 and RAW264.7, and the apparent synergistical increase of IL-1β in the culture medium mostly reflects the leakage of pro-IL-1β from these cells.

In plain English

This was a laboratory study of mouse immune cells (macrophages) by researchers in Japan. No asbestos was tested; it is mentioned only as background about earlier research. The authors exposed bone marrow macrophages from five male mice, and three mouse macrophage cell lines, to fibrous titanium dioxide or ATP, with or without a bacterial substance called LPS. They measured two inflammatory signaling proteins, IL-1β and IL-6, inside the cells and in the surrounding fluid, separating active IL-1β from its inactive precursor. Bone marrow cells and J774A.1 cells given LPS plus ATP released active IL-1β; the bone marrow cells also appeared to do so when fibrous titanium dioxide replaced ATP. In J774.1 and RAW264.7 cells, the precursor was not converted to the active form. These two lines made little or none of a protein (ASC) needed for that conversion. In J774.1 cells given both LPS and fibrous titanium dioxide, a large IL-1β release was seen only at a fibrous titanium dioxide concentration toxic to the cells. IL-6 was released after LPS alone. The authors warned that common IL-1β tests (ELISA and antibody arrays) are not appropriate for measuring the active form.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (in vitro) experiment on cultured mouse macrophages: three cell lines and bone marrow-derived macrophages from mice
• Who was studied: Three mouse macrophage cell lines (J774.1, J774A.1, RAW264.7), and bone marrow-derived macrophages from five male C57BL/6-J mice (one right femur from each), studied at the National Institute for Environmental Studies, Japan
• Exposure: Fibrous titanium dioxide (FTiO2; nominal average length 2.1 μm, width 0.14 μm), mainly at 100 μg/mL, or 5 mM ATP, with or without 100 ng/mL LPS. Tricalcium phosphate was also tested. No asbestos was tested.
• Main results:
  • In J774A.1 cells given LPS plus ATP, pro-caspase-1 was converted to its active form and active IL-1β (17 kDa) was released into the culture medium
  • In bone marrow macrophages first given LPS and then stimulated with ATP, both active caspase-1 and active IL-1β were released into the culture medium
  • In bone marrow macrophages, the inflammasome pathway appeared to be activated when fibrous titanium dioxide was used in place of ATP
  • In J774.1 and RAW264.7 cells, neither pro-caspase-1 nor pro-IL-1β was cleaved, and most of both stayed inside the cells
  • ASC was expressed in bone marrow macrophages and J774A.1 cells, was not expressed in RAW264.7, and was only faintly expressed in J774.1. NLRP3 was expressed about equally in all four cell types.
  • In J774.1 cells co-stimulated with LPS and fibrous titanium dioxide, a large IL-1β release was seen only at a concentration of fibrous titanium dioxide that was toxic to the cells (100 μg/mL)
  • IL-6 was released in response to LPS alone. ATP appeared to decrease IL-6 production in all three cell lines.
  • In J774.1 cells, caspase-1 activity inside the cells slightly decreased after fibrous titanium dioxide, LPS, or both. However, caspase inhibitors reduced IL-1β in the medium.
  • ATP was toxic to these macrophages at 2 mM or higher
• The authors' conclusion: The authors concluded that active IL-1β was released from J774A.1 cells and bone marrow macrophages through the inflammasome pathway. Only pro-IL-1β was released from J774.1 and RAW264.7 cells. They warned that ELISA and membrane array measurements of IL-1β are not appropriate for evaluating the active form.

Topics

Medical terms in this study, explained in our Lexicon: Macrophage, Apoptosis.

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