Anti-inflammatory Effects of Empagliflozin and Gemigliptin on LPS-Stimulated Macrophage via the IKK/NF-κB, MKK7/JNK, and JAK2/STAT1 Signalling Pathways

Abstract

Background. Sodium-glucose cotransporter 2 (SGLT2) and dipeptidyl peptidase-4 (DPP-4) inhibitors are glucose-lowering drugs whose anti-inflammatory properties have recently become useful in tackling metabolic syndromes in chronic inflammatory diseases, including diabetes and obesity. We investigated whether empagliflozin (SGLT2 inhibitor) and gemigliptin (DPP-4 inhibitor) improve inflammatory responses in macrophages, identified signalling pathways responsible for these effects, and studied whether the effects can be augmented with dual empagliflozin and gemigliptin therapy. Methods. RAW 264.7 macrophages were first stimulated with lipopolysaccharide (LPS), then cotreated with empagliflozin, gemigliptin, or empagliflozin plus gemigliptin. We conducted quantitative RT-PCR (qRT-PCR) to determine the most effective anti-inflammatory doses without cytotoxicity. We performed ELISA and qRT-PCR for inflammatory cytokines and chemokines and flow cytometry for CD80, the M1 macrophage surface marker, to evaluate the anti-inflammatory effects of empagliflozin and gemigliptin. NF-κB, MAPK, and JAK2/STAT signalling pathways were examined via Western blotting to elucidate the molecular mechanisms of anti-inflammation. Results. LPS-stimulated CD80+ M1 macrophages were suppressed by coincubation with empagliflozin, gemigliptin, and empagliflozin plus gemigliptin, respectively. Empagliflozin and gemigliptin (individually and combined) inhibited prostaglandin E2 (PGE2) release and COX-2, iNOS gene expression in LPS-stimulated RAW 264.7 macrophages. These three treatments also attenuated the secretion and mRNA expression of proinflammatory cytokines, such as TNF-α, IL-1β, IL-6, and IFN-γ, and proinflammatory chemokines, such as CCL3, CCL4, CCL5, and CXCL10. All of them blocked NF-κB, JNK, and STAT1/3 phosphorylation through IKKα/β, MKK4/7, and JAK2 signalling. Conclusions. Our study demonstrated the anti-inflammatory effects of empagliflozin and gemigliptin via IKK/NF-κB, MKK7/JNK, and JAK2/STAT1 pathway downregulation in macrophages. In all cases, combined empagliflozin and gemigliptin treatment showed greater anti-inflammatory properties.

Empagliflozin attenuates cardiac microvascular ischemia/reperfusion through activating the AMPKα1/ULK1/FUNDC1/mitophagy pathway

“…It is well documented that the intracellular effectors of empagliflozin include extracellular signal-regulated kinase 1/2 [ 65 ], phosphoinositide 3-kinase/Akt [ 18 ], transforming growth factor β1 [ 66 ], nuclear factor κB [ 67 ], Janus kinase 2 [ 67 ], Sirtuin 1 [ 68 ], hypoxia inducible factor 1α [ 69 ], protein kinase G Iα [ 70 ], heme oxygenase 1 [ 71 ], peroxisome proliferator-activated receptor gamma coactivator 1α [ 72 ] and AMPKα1 [ 20 ]. Among them, AMPK has been identified as an indispensable inducer of mitophagy.…”

Neuroprotective Effect of SGLT2 Inhibitors

“…In a murine model, depletion of STAT-1 activation significantly reduced synaptic dysfunction and cognitive impairment associated with Tau accumulation [ 88 ]. Empagliflozin was proven to mitigate inflammation by downregulation of the JAK2/STAT1 pathway in macrophages [ 89 ]. Macrophages take part in cognitive impairment as perivascular macrophages (PVM) are the source of vascular oxidative stress by producing a large amount of free radicals near the neurovascular unit [ 90 ].…”

Interference With the AMPKα/mTOR/NLRP3 Signaling and the IL-23/IL-17 Axis Effectively Protects Against the Dextran Sulfate Sodium Intoxication in Rats: A New Paradigm in Empagliflozin and Metformin Reprofiling for the Management of Ulcerative Colitis

“…It was previously reported that empagliflozin could exhibit an anti-inflammatory effect through SGLT-2-independent mechanisms, including intracellular activation of AMPK, attenuation of superoxide production, and the suppression of oxidative stress ( Zhou et al, 2018 ). Additionally, it showed an anti-inflammatory effect via the inhibition of NF-κB/JNK/STAT signaling pathways ( Lee et al, 2021 ). Its suppressive effect on NFκB showed an anti-inflammatory cardioprotective effect in patients subjected to doxorubicin ( Quagliariello et al, 1894 ).…”