Anti-Inflammatory Effects of the Novel PIM Kinase Inhibitor KMU-470 in RAW 264.7 Cells through the TLR4-NF-κB-NLRP3 Pathway

Abstract

PIM kinases, a small family of serine/threonine kinases, are important intermediates in the cytokine signaling pathway of inflammatory disease. In this study, we investigated whether the novel PIM kinase inhibitor KMU-470, a derivative of indolin-2-one, inhibits lipopolysaccharide (LPS)-induced inflammatory responses in RAW 264.7 cells. We demonstrated that KMU-470 suppressed the production of nitric oxide and inducible nitric oxide synthases that are induced by LPS in RAW 264.7 cells. Furthermore, KMU-470 inhibited LPS-induced up-regulation of TLR4 and MyD88, as well as the phosphorylation of IκB kinase and NF-κB in RAW 264.7 cells. Additionally, KMU-470 suppressed LPS-induced up-regulation at the transcriptional level of various pro-inflammatory cytokines such as IL-1β, TNF-α, and IL-6. Notably, KMU-470 inhibited LPS-induced up-regulation of a major component of the inflammasome complex, NLRP3, in RAW 264.7 cells. Importantly, PIM-1 siRNA transfection attenuated up-regulation of NLRP3 and pro-IL-1β in LPS-treated RAW 264.7 cells. Taken together, these findings indicate that PIM-1 plays a key role in inflammatory signaling and that KMU-470 is a potential anti-inflammatory agent.

Gryllus bimaculatus extract protects against lipopolysaccharide and palmitate-induced production of proinflammatory cytokines and inflammasome formation

“…Previous studies showed that, both palmitate and LPS induce NLRP3 inflammasome activation ( 7 , 27 ), we therefore examined the inflammasome components in this study- NLRP3, ASC, pro-caspase-1, cleaved-caspase1 (c-caspase-1), and IL-1β. LPS or palmitate treatment increased the expression of all the components while GB extract reduced them, suggesting that the GB extract can reduce inflammasome formation ( Fig 6A and B ).…”

“…RAW264.7 cells were grown in Dulbecco's modified Eagles medium (HyClone) supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin (HyClone). LPS (50 ng/ml) or 250 µM palmitate were used to induce inflammation as previously described ( 7 , 18 , 19 ) and 50 µg/ml LPS or 2 mM palmitate were used to induce cell death (20, 21).…”

“…IL-1β, a pro-inflammatory cytokine, plays an important role in inflammatory responses while IL-18 activates inflammatory responses as well as protects against invading various microorganisms ( 6 ). The inflammasome complex formation is induced in response to various stimuli such as LPS ( 7 ), palmitate ( 8 ), and anthrax toxin ( 9 ); it is also formed in case of many inflammatory disease such as alcoholic hepatitis ( 10 ), liver injury ( 11 ), lung injury ( 12 ), and several cancers ( 13 ). Thus, formation of the inflammasome complex is a novel process involved in regulating many inflammatory responses.…”

Therapeutic Potentials of Poly (ADP‐Ribose) Polymerase 1 (PARP1) Inhibition in Multiple Sclerosis and Animal Models: Concept Revisiting

“…[ 110 ] A growing number of studies have demonstrated a crucial pathophysiological role in MS and EAE for MMP2, [ 111 ] MMP9, [ 112 ] and P2Y12 receptor signaling [ 113 ] and revealed an anti‐inflammatory effect of PIM1/2 inhibition through suppressing NF‐ κ B signaling. [ 114 ] Corroborating this idea, all publications administering PJ34 in the presence of PARP1 KO observed PARP1‐independent effects. [ 109 ] The promiscuous inhibition of a growing number of off‐targets by PJ34 makes it difficult to interpret its biological effects on MS and EAE.…”

Dracocephalum moldavica Ethanol Extract Suppresses LPS-Induced Inflammatory Responses through Inhibition of the JNK/ERK/NF-κB Signaling Pathway and IL-6 Production in RAW 264.7 Macrophages and in Endotoxic-Treated Mice

“…For immunoblot analysis, LPS-stimulated cells were washed twice with ice-cold PBS. Total proteins were isolated from the cells using a lysis buffer with cocktails of protein inhibitors and then harvested with a cell scraper [33]. Total cellular protein was quantified using the Bradford assay.…”