NIMA-related kinase 7 amplifies NLRP3 inflammasome pro-inflammatory signaling in microglia/macrophages and mice models of spinal cord injury.

BACKGROUND: NIMA-related kinase-7 (NEK7) is a serine/threonine kinase that drives cell-cycle dynamics by modulating mitotic spindle formation and cytokinesis. It is also a crucial modulator of the pro-inflammatory effects of NOD-like receptor 3 (NLRP3) inflammasome. However, the role of NEK7 in microglia/macrophages post-spinal cord injury (SCI) is not well defined. METHODS: In this study, we performed both in vivo and in vitro experiments. Using an in vivo mouse SCI model, NEK7 siRNAs were administered intraspinally. For in vitro analysis, BV-2 microglia cells with NEK7-siRNA were stimulated with 1 µg/ml lipopolysaccharide (LPS) and 2 mM Adenosine triphosphate (ATP). RESULTS: Here, we found that the mRNA and protein levels of NEK7 and NLRP3 inflammasomes were upregulated in spinal cord tissues of injured mice and BV-2 microglia cells exposed to Lipopolysaccharide (LPS) and Adenosine triphosphate (ATP). Further experiments established that NEK7 and NLRP3 interacted in BV-2 microglia cells, an effect that was eliminated following NEK7 ablation. Moreover, NEK7 ablation suppressed the activation of NLRP3 inflammasomes. Although NEK7 inhibition did not significantly improve motor function post-SCI in mice, it was found to attenuate local inflammatory response and inhibit the activation of NLRP3 inflammasome in microglia/macrophages of the injured spinal cord. CONCLUSION: NEK7 amplifies NLRP3 inflammasome pro-inflammatory signaling in BV-2 microglia cells and mice models of SCI. Therefore, agents targeting the NEK7/NLRP3 signaling offers great promise in the treatment of inflammatory response post-SCI.

Modelling gestational diabetes mellitus: large animals hold great promise.

Gestational diabetes mellitus (GDM) characterized by hyperglycemia during pregnancy is a risk factor for various maternal and fetal complications. The key pathophysiological mechanisms underlying its development have not been elucidated, largely due to the lack of a model that accurately simulates the major clinical and pathological features of human GDM. In this review, we discuss the refined criteria for an ideal animal model of GDM, focusing on the key clinical and pathophysiological characteristics of human GDM. We provide a comprehensive overview of different models and currently used species for GDM research. In general, insulin insufficiency consequent to pancreatic ß-cell death represents the current leading strategy to mimic human GDM-like hyperglycemia in animals. Nonetheless, these models have a limited capacity to mimic the natural history of GDM, the marked alteration in circulating estrogen/ progestogen, obesity and its related metabolic complications. We discuss emerging evidence of the increased susceptibility to GDM in rodents and large animals with genetic modifications in pregnancy-related hormones. An appraisal of current GDM models suggests that a combination strategy involving dietary stress, pregnancy-related hormones, insulin resistance and metabolic disorders might enable the development of better GDM models and expedite the translation of basic research findings to GDM treatment.

Resistant starch intake facilitates weight loss in humans by reshaping the gut microbiota.

Emerging evidence suggests that modulation of gut microbiota by dietary fibre may offer solutions for metabolic disorders. In a randomized placebo-controlled crossover design trial (ChiCTR-TTRCC-13003333) in 37 participants with overweight or obesity, we test whether resistant starch (RS) as a dietary supplement influences obesity-related outcomes. Here, we show that RS supplementation for 8 weeks can help to achieve weight loss (mean -2.8 kg) and improve insulin resistance in individuals with excess body weight. The benefits of RS are associated with changes in gut microbiota composition. Supplementation with Bifidobacterium adolescentis, a species that is markedly associated with the alleviation of obesity in the study participants, protects male mice from diet-induced obesity. Mechanistically, the RS-induced changes in the gut microbiota alter the bile acid profile, reduce inflammation by restoring the intestinal barrier and inhibit lipid absorption. We demonstrate that RS can facilitate weight loss at least partially through B. adolescentis and that the gut microbiota is essential for the action of RS.

Akt/Aquaporin-4 Signaling Aggravates Neuropathic Pain by Activating Astrocytes after Spinal Nerve Ligation in Rats.

Aquaporins (AQPs) play critical physiological roles in water balance in the central nervous system (CNS). Aquaporin-4 (AQP4), the principal aquaporin expressed in the CNS, has been implicated in the processing of sensory and pain transmission. Akt signaling is also involved in pain mediation, such as neuroinflammatory pain and bone cancer pain. Previously, we found that expression of AQP4 and p-Akt was altered in the rat spinal cord after spinal nerve ligation (SNL). Here, we further investigated the effects of the AQP4 and Akt pathways in the spinal dorsal horn (SDH) on the pathogenesis of neuropathic pain (NP). Spinal AQP4 was significantly upregulated after SNL and was primarily expressed in astrocytes in the SDH. Inhibition of AQP4 with TGN-020 attenuated the development and maintenance of NP by inhibiting glial activation and anti-neuroinflammatory mechanisms. Moreover, inhibition of AQP4 suppressed astrocyte activation both in the SDH and in primary cultures. Similar to AQP4, we found that p-Akt was also significantly elevated after SNL. Inhibition of Akt with MK2206 suppressed AQP4 upregulation and astrocyte activation both in vivo and in vitro. Furthermore, Akt blockade with MK2206 alleviated NP in the early and late phases after SNL. These results elucidate the mechanisms involved in the roles of Akt/AQP4 signaling in the development and maintenance of NP. AQP4 is likely to be a novel therapeutic target for NP management.

The resistant effect of SIRT1 in oxidative stress-induced senescence of rat nucleus pulposus cell is regulated by Akt-FoxO1 pathway.

Objective: The senescence of nucleus pulposus (NP) cells induced by oxidative stress is one of the important causes of intervertebral disc degeneration (IDD). Herein, we investigated the role and action mechanism of silent information regulator 1 (SIRT1) in oxidative stress-induced senescence of rat NP cell.Methods: Premature senescence of rat NP cells was induced by sublethal concentration of hydrogen peroxide (H2O2) (100 µM). SIRT1 was activated with SRT1720 (5 µM) to explore its effect on NP cells senescence. FoxO1 and Akt were inhibited by AS1842856 (0.2 µM) and MK-2206 (5 µM), respectively, to explore the role of Akt-FoxO1-SIRT1 axis in rat NP cells. Pretreatment with the resveratrol (20 µM), a common antioxidant and indirect activator of SIRT1, was done to investigate its role in senescent rat NP cells.Results: The mRNA and protein levels of SIRT1 were decreased in H2O2-induced senescent rat NP cells, and that specific activation of SIRT1 suppresses senescence. And the Akt-FoxO1 pathway, as the upstream of SIRT1, might be involved in the regulation of H2O2-induced senescence of rat NP cells by affecting the expression of SIRT1. In addition, the resveratrol played an anti-senescence role in rat NP cells, which might affect the Akt-FoxO1-SIRT1 axis.Conclusion: SIRT1 ameliorated oxidative stress-induced senescence of rat NP cell which was regulated by Akt-FoxO1 pathway, and resveratrol exerted anti-senescence effects by affecting this signaling axis.

Expression of BDNF, TrkB, VEGF and CD105 is associated with pelvic lymph node metastasis and prognosis in IB2-stage squamous cell carcinoma.

Brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B (TrkB), vascular endothelial growth factor (VEGF) and CD105 are highly expressed in several types of cancer. The present study aimed to determine whether BDNF, TrkB, VEGF and CD105 are associated with the prognosis and metastasis of patients with cervical squamous cell carcinoma (SCC) at the IB2 stage. A total of 79 patients with IB2-stage SCC were enrolled in the present study. The expression levels of BDNF, TrkB, VEGF and CD105 in IB2-stage cervical cancer tissue were detected by immunohistochemistry and their association with clinicopathological indexes or prognostic factors was statistically analyzed. Reverse transcription quantitative PCR was used to detect whether the expression of VEGF was affected in SiHa cells co-cultured with BDNF. In addition, BDNF-induced SiHa cell migration and invasion were examined. BDNF expression in the cervical cancer samples was significantly associated with positive lymphovascular space invasion (P<0.001) and pelvic lymph node metastasis (P<0.05). In addition, microvessel density was verified as an independent prognostic factor for overall survival (P<0.05). In vitro analysis indicated that BDNF significantly induced cellular migration and invasion of SiHa cells in a dose-dependent manner (P<0.001). BDNF induced the expression of VEGF in SiHa cells, which was inhibited by BDNF antibodies or an inhibitor of TrkB receptor (P<0.05). BDNF may be considered a useful indicator of pelvic metastasis, which is involved in the aggressive spread of IB2-stage SCC. BDNF-induced upregulation of VEGF was revealed to act as a pro-angiogenic factor in SCC (Trial registration no. http://apps.who.int/trialsearch/; ChiCTR1800017778).

CXCL12/CXCR4 signaling contributes to neuropathic pain via central sensitization mechanisms in a rat spinal nerve ligation model.

BACKGROUND: Previous studies have demonstrated that the CXCL12/CXCR4 signaling axis is involved in the regulation of neuropathic pain (NP). Here, we performed experiments to test whether the CXCL12/CXCR4 signaling pathway contributes to the pathogenesis of neuropathic pain after spinal nerve ligation (SNL) via central sensitization mechanisms. METHODS: Neuropathic pain was induced and assessed in a SNL rat model. The expression and distribution of CXCL12 or CXCR4 were examined by immunofluorescence staining and western blot. The effects of CXCL12 rat peptide, CXCL12 neutralizing antibody, CXCR4 antagonist, and astrocyte metabolic inhibitor on pain hypersensitivity were explored by behavioral tests in naive or SNL rats. We measured the expression level of c-Fos and CGRP to evaluate the sensitization of neurons by RT-PCR. The activation of astrocyte and microglia was analyzed by measuring the level of GFAP and iba-1. The mRNA levels of the pro-inflammatory cytokines such as TNF-α, IL-1ß, and IL-6 and Connexin 30, Connexin 43, EAAT 1, EAAT 2 were also detected by RT-PCR. RESULTS: First, we found that the expression of CXCL12 and CXCR4 was upregulated after SNL. CXCL12 was mainly expressed in the neurons while CXCR4 was expressed both in astrocytes and neurons in the spinal dorsal horn after SNL. Moreover, intrathecal administration of rat peptide, CXCL12, induced hypersensitivity in naive rats, which was partly reversed by fluorocitrate. In addition, the CXCL12 rat peptide increased mRNA levels of c-Fos, GFAP, and iba-1. A single intrathecal injection of CXCL12 neutralizing antibody transiently reversed neuropathic pain in the SNL rat model. Consecutive use of CXCL12 neutralizing antibody led to significant delay in the induction of neuropathic pain, and reduced the expression of GFAP and iba-1 in the spinal dorsal horn. Finally, repeated intrathecal administration of the CXCR4 antagonist, AMD3100, significantly suppressed the initiation and duration of neuropathic pain. The mRNA levels of c-Fos, CGRP, GFAP, iba-1, and pro-inflammatory cytokines, also including Connexin 30 and Connexin 43 were decreased after injection of AMD3100, while EAAT 1 and EAAT 2 mRNAs were increased. CONCLUSION: We demonstrate that the CXCL12/CXCR4 signaling pathway contributes to the development and maintenance of neuropathic pain via central sensitization mechanisms. Importantly, intervening with CXCL12/CXCR4 presents an effective therapeutic approach to treat the neuropathic pain.