Lipopolysaccharide promotes Drp1‐dependent mitochondrial fission and associated inflammatory responses in macrophages

Abstract

Mitochondria have a multitude of functions, including energy generation and cell signaling. Recent evidence suggests that mitochondrial dynamics (i.e. the balance between mitochondrial fission and fusion) also regulate immune functions. Here, we reveal that lipopolysaccharide (LPS) stimulation increases mitochondrial numbers in mouse bone marrow‐derived macrophages (BMMs) and human monocyte‐derived macrophages. In BMMs, this response requires Toll‐like receptor 4 (Tlr4) and the TLR adaptor protein myeloid differentiation primary response 88 (MyD88) but is independent of mitochondrial biogenesis. Consistent with this phenomenon being a consequence of mitochondrial fission, the dynamin‐related protein 1 (Drp1) GTPase that promotes mitochondrial fission is enriched on mitochondria in LPS‐activated macrophages and is required for the LPS‐mediated increase in mitochondrial numbers in both BMMs and mouse embryonic fibroblasts. Pharmacological agents that skew toward mitochondrial fusion also abrogated this response. LPS triggered acute Drp1 phosphorylation at serine 635 (S635), followed by sustained Drp1 dephosphorylation at serine 656 (S656), in BMMs. LPS‐induced S656 dephosphorylation was abrogated in MyD88 ‐deficient BMMs, suggesting that this post‐translational modification is particularly important for Tlr4‐inducible fission. Pharmacological or genetic targeting of Tlr4‐inducible fission had selective effects on inflammatory mediator production, with LPS‐inducible mitochondrial fission promoting the expression and/or secretion of a subset of inflammatory mediators in BMMs and mouse embryonic fibroblasts. Thus, triggering of Tlr4 results in MyD88‐dependent activation of Drp1, leading to inducible mitochondrial fission and subsequent inflammatory responses in macrophages.

The Mitochondrial Fission Regulator DRP1 Controls Post-Transcriptional Regulation of TNF-α

“…To investigate whether LPS-induced changes in mitochondrial morphology were due to DRP1 post-translational modification, we tested the effect of LPS stimulation on DRP1 phosphorylation at its activating site (Ser635) and inhibitory site (Ser656) during LPS stimulation. Similar to a recent report, we observed a trend in increased DRP1 Ser635 phosphorylation, yet no change in DRP1 Ser656 phosphorylation during 4 h LPS stimulation ( Figures S1B, C ) ( Kapetanovic et al., 2020 ). Furthermore, we examined the effect of LPS stimulation on expression of the key DRP1 adaptor protein Mitochondrial Fission Factor (MFF) as well as the well-characterized fusion regulators Mitofusin-2 (MFN2) and OPA1 ( Figures S1B, C ).…”

Selonsertib, a potential drug for liver failure therapy by rescuing the mitochondrial dysfunction of macrophage via ASK1–JNK–DRP1 pathway

“…Macrophages play a pivotal role in modulating the hepatic immune microenvironment and inflammatory response in ALF [ 19 ]. As previous studies showed that LPS exposure usually induced mitochondrial dysfunction in macrophages [ 20 , 21 ], we further investigated the effect of selonsertib in protecting the mitochondrial function and dynamics of hepatic macrophages. As shown in Fig.…”

“…Several studies have proven that the danger of ALF is dependent on liver immune homeostasis and the inflammatory microenvironment, which is mainly modulated by macrophages [ 19 , 37 ]. As LPS exposure is known to induce macrophage mitochondrial dysfunction in acute tissue injury [ 21 ], we then investigated the effect of selonsertib in protecting the mitochondrial function and dynamics of hepatic macrophages and also used LPS-primed macrophages as an in vitro model to determine this issue. We found that selonsertib pretreatment effectively prevented LPS-induced activation of JNK and DRP1 and mitochondrial dysfunction in primary hepatic macrophages and macrophagic cell line and also suppressed the release of the proinflammatory cytokines TNF-α and IL-1α, which are known as major cytokines causing liver injury and inflammation [ 38 , 39 ].…”

Selonsertib, a Potential Drug for Liver Failure Therapy by Rescuing the Mitochondrial Dysfunction of Macrophage via ASK1-JNK-DRP1 Pathway

“…Several studies proved that the danger of ALF was dependent on the liver immune homeostasis and in ammation microenvironment mainly modulated by macrophages [17,35]. As LPS exposure was known to induce macrophage mitochondrial dysfunction in acute tissue injury [19], we then used LPS-primed macrophages as in vitro model to determine whether selonsertib could protect the mitochondrial function of macrophages in ALF. We found that selonsertib pretreatment effectively prevented the LPS-induced mitochondrial dysfunction of macrophage and the release of pro-in ammatory cytokines TNF-α and IL-1α, which were known as major cytokines causing liver injury and in ammation [36,37].…”

“…Macrophages play a pivotal role in modulating hepatic immune microenvironment and in ammatory response in ALF [17]. As previous studies showed that LPS exposure usually induced mitochondrial dysfunction in macrophages [18,19], we further determine the role of selonsertib in protecting mitochondrial function in the LPS-primed mouse macrophage cell line, RAW264.7. As we expected, LPS exposure induced a marked increase in the mitochondrial oxidative stress in macrophages as indicating by MitoSox, while selonsertib treatment signi cantly weakened the red uorescence intensity in the LPSprimed macrophages ( Figure 4A-left).…”