Pharmacological inhibition of STAT3 pathway ameliorates acute liver injury in vivo via inactivation of inflammatory macrophages and hepatic stellate cells

Abstract

Abbreviations: AEC, 3-amino-9-ethyl carbazole; Arg1, Arginase 1; CCL2/MCP1, chemokine (C-C motif) ligand 2/Macrophage chemotactic protein-1; CCl 4 , carbon tetrachloride; CCR2, chemokine (C-C motif) receptor 2; DAPI, 4',6-diamidino-2-phenylindole; DMSO, Dimethyl sulfoxide; ECM, extracellular matrix; HRP, horseradish peroxidase; HSCs, hepatic stellate cells; IL-1β, Interleukin 1 beta; IL-6, Interleukin 6; MHC II, major histocompatibility complex class II; MRC1, mannose receptor 1; mRNA, messenger RNAs; NaCl, sodium chloride; NASH, non-alcoholic steatohepatitis; NOS2/iNOS, nitric oxide synthase 2; qRT-PCR, quantitative real-time polymerase chain reaction; SDS, sodium dodecyl sulfate; TGFβ, transforming growth factor beta; Tris-HCl, Tris(hydroxymethyl)aminomethane hydrochloride; YM1, Beta-N-acetyl hexosaminidase or Chi3l3, Chitinase-3-like protein 3; α-SMA, alpha smooth muscle actin. AbstractLiver diseases represent a major health problem worldwide, in particular, acute liver injury is associated with high mortality and morbidity. Inflammatory macrophages and hepatic stellate cells (HSCs) are known to be involved in the pathogenesis of acute liver injury. In this study, we have investigated the implication of STAT3 inhibition in acute liver injury/early fibrogenesis. In fibrotic human livers, we found STAT3 mRNA expression was significantly upregulated and correlated with collagen I expression. In vitro, STAT3 signaling pathway was found to be activated in TGFβ-activated HSCs and inflammatory macrophages. STAT3 inhibitor, WP1066 significantly inhibited TGFβ-induced collagen I, vimentin and α-SMA expression, and contractility in human HSCs. In LPS-and IFNγ-induced pro-inflammatory macrophages, WP1066 strongly attenuated nitric-oxide release and expression of major inflammatory markers such as TNF-α, iNOS, CCL2, IL-1β, IL-6, and CCR2. In vivo in CCl 4 -induced acute liver injury mouse model, WP1066 significantly reduced collagen expression, HSCs activation, and intrahepatic inflammation. Finally, in LPSinduced human hepatic 3D spheroid model, WP1066 inhibited LPS-induced fibrotic and inflammatory parameters. In conclusion, our results demonstrate that the therapeutic inhibition of STAT3 pathway using WP1066 targeting HSCs and inflammatory macrophages suggests a potential pharmacological approach for the treatment of acute liver injury. K E Y W O R D Sacute liver injury, hepatic stellate cells, inflammation, macrophages, STAT3 signaling pathway, WP1066 78 | ÖZTÜRK AKCORA eT Al.

Stattic alleviates acute hepatic damage induced by LPS/d-galactosamine in mice

“… 10 WP1066, another STAT3 inhibitor, significantly reduced the level of TNF-α and IL-6 in LPS- or IFN-γ-stimulated mouse RAW264.7 macrophages. 11 In vivo , STAT3 inhibitor Nifuroxazide significantly decreased the mRNA and protein level of TNF-α and IL-18 and reduced infiltration of inflammatory cells in diabetes-associated renal injury. 23 LLL12, an allosteric STAT3 inhibitor, also attenuated the infiltration of inflammatory cells and the level of TNF-α and IL-1β in LPS-induced acute lung injury.…”

“…Recently, increasing evidence indicates that STAT3 plays crucial roles in the regulation of inflammation and apoptosis. 10 , 13 STAT3 has become a novel therapeutic target for intervention in inflammation-related disorders, 10 , 11 but it remains unclear whether STAT3 plays a part in acute hepatic damage. In the present study, the results showed that Stattic mitigated the hepatic morphologic abnormalities, such as hepatocellular necrosis and destruction of hepatic lobule, and decreased the level of aminotransferase in LPS/ D -GalN-insulted mice.…”

“… 8 , 9 A growing body of evidence shows that inhibition of STAT3 plays an anti-inflammatory role in inflammation-related experimental models. 10 , 11 For instance, STAT3 inhibitor dramatically reduced the level of TNF-α and IL-6 in LPS-stimulated mouse microglia BV-2 cells and mouse RAW264.7 macrophages. 10 , 11 However, it is still unclear whether STAT3 also plays a crucial role in LPS/ D -GalN-induced hepatic damage.…”

Niclosamide Targets Inflammatory and Profibrotic Pathways in Amyotrophic Lateral Sclerosis

“…STAT3 contributes to brosis by inducing the production of ECM, generally sustaining the differentiation of organ resident cells via canonical and non-canonical pathways. Indeed, several lines of evidence report the fundamental role of STAT3 in broblast plasticity in different tissues [6,18], where the inhibition of its signaling pathway attenuates brosis by decreasing several markers, among which α-SMA [34] and N-cadherin [27]. Since S100A4 is a known inducer of JAK/STAT pathway, it is possible that S100A4 knock-down can indirectly decrease the levels of the brotic molecules α-SMA and N-cadherin through the inhibition of STAT3.…”

Participation of ABCA1 Transporter in Pathogenesis of Chronic Obstructive Pulmonary Disease

“…Such multidirectional participation highlights the complexity of parallel processes and their insufficient investigation. STAT3 regulates a number of fundamental cellular processes, including inflammation, proliferation, differentiation, and cell migration [ 142 ]. STAT3 can also regulate apoptosis by inducing the expression of the apoptosis inhibitor Bcl-2 (B-cell lymphoma 2) [ 143 , 144 ].…”