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Objectives: Sepsis-associated cognitive dysfunction is a common complication in patients with sepsis and lack of effective treatment. Its pathological mechanisms remain unclear. Salt-induced kinase (SIK) is an important molecule in the regulation of metabolism, immunity, and inflammatory response. It is associated with the development of many neurological diseases. This study aims to investigate the expression of SIK in the hippocampus of septic mice, and to evaluate the role and mechanism of the SIK inhibitor HG-9-91-01 in sepsis-associated cognitive dysfunction., Methods: Firstly, C57BL/6 mice were randomly divided into a control group (Con group) and a sepsis model group [lipopolysaccharide (LPS) group]. The model group was injected intraperitoneally with LPS at a dose of 8 mg/kg and the Con group was injected with an equal volume of normal saline. Hippocampal tissues were harvested at 1, 3, and 6 days after injection and the expressions of SIK1, SIK2, and SIK3 were detected by real-time fluorescence quantitative PCR (qPCR) and Western blotting. Secondly, C57BL/6 mice were randomly divided into a Con group, a LPS group, and a SIK inhibitor group (HG group). The LPS and HG groups were injected with LPS to establish a sepsis model; in the HG group, HG-9-91-01 (10 mg/kg) was injected intraperitoneally at 3-6 days after LPS injection, and the LPS group was injected with the same volume of vehicle. Cognitive function was assessed at 7-11 days after LPS injection using the Morris water maze (MWM). Hippocampal tissues were harvested after the behavioral tests, and the mRNA levels of inflammatory factors and microglial markers were assessed by qPCR. The protein levels of inducible nitric oxide synthase (iNOS), CD68, ionized calcium binding adaptor molecule 1 (Iba-1), N-methyl- D -aspartate (NMDA) receptor (NR) subunit, cAMP response element-binding protein (CREB)-regulated transcription coactivator 1 (CRTC1), and insulin-like growth factor 1 (IGF-1) were detected by Western blotting. Immunohistochemistry (IHC) was used to detect the expression of Iba-1 positive cells in the CA1, CA3 and dentate gyrus (DG) of the hippocampus, followed by Sholl analysis., Results: Compared with the Con group, the mRNA and protein levels of SIK1 , SIK2 , and SIK3 in the hippocampus were increased in the LPS group (all P <0.05). Compared with the Con group, mice in the LPS group had a significantly longer escape latency, a lower percentage of target quadrant dwell time and a reduced locomotor speed (all P <0.05); the HG group had a decreased escape latency and an increased percentage of time spent in the target quadrant in comparison with the LPS group (both P <0.05). The mRNA levels of inflammatory factors [tumor necrosis factor-α ( TNF-α ), interleukin-1β ( IL-1β ), interleukin-6 ( IL-6 )], and the M1-type microglial markers iNOS and CD68 in the hippocampus of the LPS group were increased in comparison with the Con group, while the M2-type microglial markers CD206 and arginase-1 ( Arg-1 ) were decreased. Compared with the LPS group, the mRNA levels of TNF-α , IL-1β , IL-6 , and iNOS were downregulated, while the levels of CD206 and Arg-1 were upregulated in the HG group (all P <0.05). The protein levels of iNOS, CD68, and Iba-1 in the hippocampus of the LPS group were increased in comparison with the Con group, but they were downregulated in the HG group in comparison with the LPS group (all P <0.05). The number of Iba-1 positive cells in CA1, CA3, and DG of the hippocampus was increased in the LPS group in comparison with the Con group, but they were decreased in the HG group in comparison with the LPS group (all P <0.05). Sholl analysis showed that the number of intersections at all radii between 8-38 µm from the microglial soma was decreased in the LPS group in comparison with the Con group (all P <0.05). Compared with the LPS group, the number of intersections at all radii between 14-20 µm was significantly increased in the HG group (all P <0.05). The protein levels of NR subunit NR1, NR2A, NR2B, and IGF-1 were downregulated in the hippocampus of the LPS group in comparison with the Con group, while the expression of phosphorylated CRTC1 (p-CRTC1) was increased. Compared with the LPS group, the levels of NR1, NR2A, NR2B, and IGF-1 were upregulated, while p-CRTC1 was downregulated in the HG group (all P <0.05)., Conclusions: SIK expression is upregulated in the hippocampus of septic mice. The SIK inhibitor HG-9-91-01 ameliorates sepsis-associated cognitive dysfunction in mice, and the mechanism may involve in the activation of the CRTC1/IGF-1 pathway, inhibition of neuroinflammation, and enhancement of synaptic plasticity.

Major depressive disorder is a frequently occurring neuropsychiatric disorder throughout the world. However, the limited and delayed therapeutic efficacy of monoaminergic medications has led to intensive research efforts to develop novel antidepressants. We have previously demonstrated that hippocampal salt-inducible kinase 2 (SIK2) plays a role in the pathogenesis of depression via regulating the downstream CREB-regulated transcription coactivator 1 (CRTC1)-cAMP response element-binding protein (CREB)-brain derived neurotrophic factor (BDNF) pathway. HG-9-91-01 is a potent and selective inhibitor of salt-inducible kinases (SIKs). The present study aims to explore whether HG-9-91-01 has antidepressant-like actions in male C57BL/6J mice. The chronic unpredictable mild stress (CUMS) model of depression, various behavioral tests, western blotting, co-immunoprecipitation, immunofluorescence, stereotactic infusion, and viral-mediated genetic knockdown were used together. It was found that hippocampal infusion of HG-9-91-01 induced significant antidepressant-like effects in the CUMS model, accompanied with preventing the enhancement of CUMS on the hippocampal SIK2 expression and cytoplasmic translocation of CRTC1. HG-9-91-01 treatment also reversed the decreasing effects of CUMS on the BDNF signaling cascade and adult neurogenesis in the hippocampus. Moreover, the antidepressant-like actions of HG-9-91-01 in mice required the hippocampal CRTC1-CREB-BDNF pathway. In conclusion, HG-9-91-01 has potential of being a novel antidepressant candidate. • HG-9-91-01 administration fully prevented the CUMS-induced depressive-like behaviors in mice. • HG-9-91-01 administration significantly antagonized the effects of CUMS on the hippocampal SIK2-CRTC1-CREB system. • HG-9-91-01 administration notably reversed the CUMS-decreased BDNF signaling and adult neurogenesis in the hippocampus. [ABSTRACT FROM AUTHOR]

Macrophages acquire strikingly different properties that enable them to play key roles during the initiation, propagation, and resolution of inflammation. Classically activated (M1) macrophages produce proinflammatory mediators to combat invading pathogens and respond to tissue damage in the host, whereas regulatory macrophages (M2b) produce high levels of anti-inflammatory molecules, such as IL-10, and low levels of proinflammatory cytokines, like IL-12, and are important for the resolution of inflammatory responses. A central problem in this area is to understand how the formation of regulatory macrophages can be promoted at sites of inflammation to prevent and/or alleviate chronic inflammatory and autoimmune diseases. Here, we demonstrate that the salt-inducible kinases (SIKs) restrict the formation of regulatory macrophages and that their inhibition induces striking increases in many of the characteristic markers of regulatory macrophages, greatly stimulating the production of IL-10 and other anti-inflammatory molecules. We show that SIK inhibitors elevate IL-10 production by inducing the dephosphorylation of cAMP response element-binding protein (CREB)-regulated transcriptional coactivator (CRTC) 3, its dissociation from 14-3-3 proteins and its translocation to the nucleus where it enhances a gene transcription program controlled by CREB. Importantly, the effects of SIK inhibitors on IL-10 production are lost in macrophages that express a drug-resistant mutant of SIK2. These findings identify SIKs as a key molecular switch whose inhibition reprograms macrophages to an anti-inflammatory phenotype. The remarkable effects of SIK inhibitors on macrophage function suggest that drugs that target these protein kinases may have therapeutic potential for the treatment of inflammatory and autoimmune diseases [ABSTRACT FROM AUTHOR]

Background: Interleukin-10 (IL-10) is a potent immunoregulatory cytokine that plays a pivotal role in maintaining mucosal immune homeostasis. As a novel synthetic inhibitor of salt-inducible kinases (SIKs), HG-9-91-01 can effectively enhance IL-10 secretion at the cellular level, but its in vivo immunoregulatory effects remain unclear. In this study, we investigated the effects and underlying mechanism of HG-9-91-01 in murine colitis models., Methods: The anti-inflammatory effects of HG-9-91-01 were evaluated on 2, 4, 6-trinitrobenzene sulfonic acid (TNBS)-, dextran sulfate sodium-induced colitis mice, and IL-10 knockout chronic colitis mice. The in vivo effector cell of HG-9-91-01 was identified by fluorescence-activated cell sorting and quantitative real-time polymerase chain reaction. The underlying mechanism of HG-9-91-01 was investigated via overexpressing SIKs in ANA-1 macrophages and TNBS colitis mice., Results: Treatment with HG-9-91-01 showed favorable anticolitis effects in both TNBS- and DSS-treated mice through significantly promoting IL-10 expression in colonic macrophages but failed to protect against IL-10 KO murine colitis. Further study indicated that HG-9-91-01 markedly enhanced the nuclear level of cAMP response element-binding protein (CREB)-regulated transcription coactivator 3 (CRTC3), whereas treatment with lentiviruses encoding SIK protein markedly decreased the nuclear CRTC3 level in HG-9-91-01-treated ANA-1 macrophages. In addition, intracolonic administration with lentiviruses encoding SIK protein significantly decreased the nuclear CRTC3 level in the lamina propria mononuclear cells and ended the anti-inflammatory activities of HG-9-91-01., Conclusions: We found that HG-9-91-01 promoted the IL-10 expression of colonic macrophages and exhibited its anticolitis activity through the SIK/CRTC3 axis, and thus it may represent a promising strategy for inflammatory bowel disease therapy., (© 2021 Crohn’s & Colitis Foundation. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The salt-inducible kinases (SIKs) control a novel molecular switch regulating macrophage polarization. Pharmacological inhibition of the SIKs induces a macrophage phenotype characterized by the secretion of high levels of anti-inflammatory cytokines, including interleukin (IL)-10, and the secretion of very low levels of pro-inflammatory cytokines, such as tumour necrosis factor α. The SIKs, therefore, represent attractive new drug targets for the treatment of macrophage-driven diseases, but which of the three isoforms, SIK1, SIK2 or SIK3, would be appropriate to target remains unknown. To address this question, we developed knock-in (KI) mice for SIK1, SIK2 and SIK3, in which we introduced a mutation that renders the enzymes catalytically inactive. Characterization of primary macrophages from the single and double KI mice established that all three SIK isoforms, and in particular SIK2 and SIK3, contribute to macrophage polarization. Moreover, we discovered that inhibition of SIK2 and SIK3 during macrophage differentiation greatly enhanced the production of IL-10 compared with their inhibition in mature macrophages. Interestingly, macrophages differentiated in the presence of SIK inhibitors, MRT199665 and HG-9-91-01, still produced very large amounts of IL-10, but very low levels of pro-inflammatory cytokines, even after the SIKs had been reactivated by removal of the drugs. Our data highlight an integral role for SIK2 and SIK3 in innate immunity by preventing the differentiation of macrophages into a potent and stable anti-inflammatory phenotype., (© 2017 The Author(s).)