Regulation of iron metabolism in HEC-1A endometrium cells by macrophage-derived factors and fractalkine.

Macrophages in the endometrium promote receptivity and implantation by secreting proinflammatory cytokines and other factors like fractalkine (FKN). Macrophages are closely linked to regulating iron homeostasis and can modulate iron availability in the tissue microenvironment. It has been revealed that the iron metabolism of the mother is crucial in fertility. Iron metabolism is strictly controlled by hepcidin, the principal iron regulatory protein. The inflammatory cytokines can modulate hepcidin synthesis and, therefore, the iron metabolism of the endometrium. It was proven recently that FKN, a unique chemokine, is implicated in maternal-fetal communication and may contribute to endometrial receptivity and implantation. In the present study, we investigated the effect of activated THP-1 macrophages and FKN on the iron metabolism of the HEC-1A endometrial cells. We established a noncontact coculture with or without recombinant human FKN supplementation to study the impact of the macrophage-derived factors and FKN on the regulation of hepcidin synthesis and iron release and storage of endometrial cells. Based on our findings, the conditioned medium of the activated macrophages could modify hepcidin synthesis via the nuclear factor kappa-light-chain-enhancer of activated B cells, the signal transducer and activator of transcription 3, and the transferrin receptor 2/bone morphogenetic protein 6/suppressor of mothers against decapentaplegic 1/5/8 signaling pathways, and FKN could alter this effect on the endometrial cells. It was also revealed that the conditioned macrophage medium and FKN modulated the iron release and storage of HEC-1A cells. FKN signaling may be involved in the management of iron trafficking of the endometrium by the regulation of hepcidin. It can contribute to the iron supply for fetal development at the early stage of the pregnancy.

Fractalkine isoforms differentially regulate microglia-mediated inflammation and enhance visual function in the diabetic retina.

Diabetic retinopathy (DR) affects about 200 million people worldwide, causing leakage of blood components into retinal tissues, leading to activation of microglia, the resident phagocytes of the retina, promoting neuronal and vascular damage. The microglial receptor, CX3CR1, binds to fractalkine (FKN), an anti-inflammatory chemokine that is expressed on neuronal membranes (mFKN), and undergoes constitutive cleavage to release a soluble domain (sFKN). Deficiencies in CX3CR1 or FKN showed increased microglial activation, inflammation, vascular damage, and neuronal loss in experimental mouse models. To understand the mechanism that regulates microglia function, recombinant adeno-associated viral vectors (rAAV) expressing mFKN or sFKN were delivered to intact retinas prior to diabetes. High-resolution confocal imaging and mRNA-seq were used to analyze microglia morphology and markers of expression, neuronal and vascular health, and inflammatory mediators. We confirmed that prophylactic intra-vitreal administration of rAAV expressing sFKN (rAAV-sFKN), but not mFKN (rAAV-mFKN), in FKNKO retinas provided vasculo- and neuro-protection, reduced microgliosis, mitigated inflammation, improved overall optic nerve health by regulating microglia-mediated inflammation, and prevented fibrin(ogen) leakage at 4 weeks and 10 weeks of diabetes induction. Moreover, administration of sFKN improved visual acuity. Our results elucidated a novel intervention via sFKN gene therapy that provides an alternative pathway to implement translational and therapeutic approaches, preventing diabetes-associated blindness.

PVT1 promotes proliferation and macrophage immunosuppressive polarization through STAT1 and CX3CL1 regulation in glioblastoma multiforme.

AIMS: This study aimed to investigate the role of plasmacytoma variant translocation 1 (PVT1), a long non-coding RNA, in glioblastoma multiforme (GBM) and its impact on the tumor microenvironment (TME). METHODS: We assessed aberrant PVT1 expression in glioma tissues and its impact on GBM cell growth in vitro and in vivo. Additionally, we investigated PVT1's role in influencing glioma-associated macrophages. To understand PVT1's role in cell growth and the immunosuppressive TME, we performed a series of comprehensive experiments. RESULTS: PVT1 was overexpressed in GBM due to copy number amplification, correlating with poor prognosis. Elevated PVT1 promoted GBM cell proliferation, while its downregulation inhibited growth in vitro and in vivo. PVT1 inhibited type I interferon-stimulated genes (ISGs), with STAT1 as the central hub. PVT1 correlated with macrophage enrichment and regulated CX3CL1 expression, promoting recruitment and M2 phenotype polarization of macrophages. PVT1 localized to the cell nucleus and bound to DHX9, enriching at the promoter regions of STAT1 and CX3CL1, modulating ISGs and CX3CL1 expression. CONCLUSION: PVT1 plays a significant role in GBM, correlating with poor prognosis, promoting cell growth, and shaping an immunosuppressive TME via STAT1 and CX3CL1 regulation. Targeting PVT1 may hold therapeutic promise for GBM patients.

Exercise, Spinal Microglia and Neuropathic Pain: Potential Molecular Mechanisms.

As one of the most common neuropathic disorders, neuropathic pain often has a negative impact on patients with persistent pain, mood disorders and sleep disturbances. Currently, neuropathic pain is not treated with any specific drug, instead, drugs for other diseases are used as replacements in clinics, but most have adverse effects. In recent years, the role of spinal cord microglia in the pathogenesis of neuropathic pain has been widely recognized, and they are being explored as potential therapeutic targets. Spinal microglia are known to be involved in the pathogenic mechanisms of neuropathic pain through purine signaling, fractalkine signaling, and p38 MAPK signaling. Exercise is a safe and effective treatment, and numerous studies have demonstrated its effectiveness in improving neurological symptoms. Nevertheless, it remains unclear what the exact molecular mechanism is. This review summarized the specific molecular mechanisms of exercise in alleviating neuropathic pain by mediating the activity of spinal microglia and maintaining the phenotypic homeostasis of spinal microglia through purine signaling, fractalkine signaling and p38 MAPK signaling. In addition, it has been proposed that different intensities and types of exercise affect the regulation of the above-mentioned signaling pathways differently, providing a theoretical basis for the improvement of neuropathic pain through exercise.

Hypothalamic FTO promotes high-fat diet-induced leptin resistance in mice through increasing CX3CL1 expression.

Long-term consumption of a high-fat diet (HFD) disrupts energy homeostasis and leads to weight gain. The fat mass and obesity-associated (FTO) gene has been consistently identified to be associated with HFD-induced obesity. The hypothalamus is crucial for regulating energy balance, and HFD-induced hypothalamic leptin resistance contributes to obesity. FTO, an N6-methyladenosine (m6A) RNA methylation regulator, may be a key mediator of leptin resistance. However, the exact mechanisms remain unclear. Therefore, the present study aims to investigate the association between FTO and leptin resistance. After HFD or standard diet (SD) feeding in male mice for 22 weeks, m6A-sequencing and western blotting assays were used to identify target genes and assess protein level, and molecular interaction changes. CRISPR/Cas9 gene knockout system was employed to investigate the potential function of FTO in leptin resistance and obesity. Our data showed that chemokine (C-X3-C motif) ligand 1 (CX3CL1) was a direct downstream target of FTO-mediated m6A modification. Furthermore, upregulation of FTO/CX3CL1 and suppressor of cytokine signaling 3 (SOCS3) in the hypothalamus impaired leptin-signal transducer and activator of transcription 3 signaling, resulting in leptin resistance and obesity. Compared to wild-type (WT) mice, FTO deficiency in leptin receptor-expressing neurons of the hypothalamus significantly inhibited the upregulation of CX3CL1 and SOCS3, and partially ameliorating leptin resistance under HFD conditions. Our findings reveal that FTO involved in the hypothalamic leptin resistance and provides novel insight into the function of FTO in the contribution to hypothalamic leptin resistance and obesity.

Validating Fractalkine receptor as a target and identifying candidates for drug discovery against type 2 diabetes.

Type 2 diabetes mellitus (T2DM) is one of the most common chronic diseases employing abnormal levels of insulin. Enhancing the insulin production is greatly aided by the regulatory mechanisms of the Fractalkine receptor (CX3CR1) system in islet ß-cell function. However, elements including a high-fat diet, obesity, and ageing negatively impact the expression of CX3CR1 in islets. CX3CL1/CX3CR1 receptor-ligand complex is now recognized as a novel therapeutic target. It suggests that T2DM-related ß-cell dysfunction may result from lower amount of these proteins. We analyzed the differential expression of CX3CR1 gene samples taken from persons with T2DM using data obtained from the Gene Expression Omnibus database. Homology modeling enabled us to generate the three-dimensional structure of CX3CR1 and a possible binding pocket. The optimized CX3CR1 structure was subjected to rigorous screening against a massive library of 693 million drug-like molecules from the ZINC15 database. This screening process led to the identification of three compounds with strong binding affinity at the identified binding pocket of CX3CR1. To further evaluate the potential of these compounds, molecular dynamics simulations were conducted over a 50 ns time scale to assess the stability of the protein-ligand complexes. These simulations revealed that ZINC000032506419 emerged as the most promising drug-like compound among the three potent molecules. The discovery of ZINC000032506419 holds exciting promise as a potential therapeutic agent for T2D and other related metabolic disorders. These findings pave the way for the development of effective medications to address the complexities of T2DM and its associated metabolic diseases.

Fractalkine's dual role in inflammation and hard tissue formation in cultured human dental pulp cells.

This study aimed to explore the potential roles of fractalkine/CX3CR1, primarily expressed in vascular endothelial cells and has recently been identified in dental pulp cells at sites of pulp tissue inflammation, not only in inflammation but also in pulp hard tissue formation. To this end, cultured human dental pulp cells were grown in 10% FBS-supplemented α-MEM. Fractalkine was introduced to the culture, and COX-2 and dentin sialophosphoprotein (DSPP) expression levels were evaluated via western blotting. Real-time PCR was used to examine BMP-2 and Osterix mRNA expression. Calcified nodule formation was evaluated with Alizarin red staining. Results revealed that fractalkine increased COX-2 protein expression, calcified nodule formation, and BMP-2 and Osterix mRNA expression in a concentration- and time-dependent manner. DSPP protein expression also increased upon fractalkine addition. This effect of fractalkine on expression of DSPP protein was inhibited in the presence of the CX3CR1 inhibiter ADZ8797. In conclusion, our findings suggest a dual role for fractalkine in promoting pulp inflammation via COX-2 production and contributing to pulp hard tissue formation by stimulating the expression of hard tissue formation markers.

The CX3CL1-CX3CR1 chemokine axis can contribute to tumor immune evasion and blockade with a novel CX3CR1 monoclonal antibody enhances response to anti-PD-1 immunotherapy.

CX3CL1 secreted in the tumor microenvironment serves as a chemoattractant playing a critical role in metastasis of CX3CR1 expressing cancer cells. CX3CR1 can be expressed in both cancer and immune-inhibitory myeloid cells to facilitate their migration. We generated a novel monoclonal antibody against mouse CX3CR1 that binds to CX3CR1 and blocks the CX3CL1-CX3CR1 interaction. We next explored the immune evasion strategies implemented by the CX3CL1-CX3CR1 axis and find that it initiates a resistance program in cancer cells that results in 1) facilitation of tumor cell migration, 2) secretion of soluble mediators to generate a pro-metastatic niche, 3) secretion of soluble mediators to attract myeloid populations, and 4) generation of tumor-inflammasome. The CX3CR1 monoclonal antibody reduces migration of tumor cells and decreases secretion of immune suppressive soluble mediators by tumor cells. In combination with anti-PD-1 immunotherapy, this CX3CR1 monoclonal antibody enhances survival in an immunocompetent mouse colon carcinoma model through a decrease in tumor-promoting myeloid populations. Thus, this axis is involved in the mechanisms of resistance to anti-PD-1 immunotherapy and the combination therapy can overcome a portion of the resistance mechanisms to anti-PD-1.

Chronic hyperpalatable diet induces impairment of hippocampal-dependent memories and alters glutamatergic and fractalkine axis signaling.

Chronic consumption of hyperpalatable and hypercaloric foods has been pointed out as a factor associated with cognitive decline and memory impairment in obesity. In this context, the integration between peripheral and central inflammation may play a significant role in the negative effects of an obesogenic environment on memory. However, little is known about how obesity-related peripheral inflammation affects specific neurotransmission systems involved with memory regulation. Here, we test the hypothesis that chronic exposure to a highly palatable diet may cause neuroinflammation, glutamatergic dysfunction, and memory impairment. For that, we exposed C57BL/6J mice to a high sugar and butter diet (HSB) for 12 weeks, and we investigated its effects on behavior, glial reactivity, blood-brain barrier permeability, pro-inflammatory features, glutamatergic alterations, plasticity, and fractalkine-CX3CR1 axis. Our results revealed that HSB diet induced a decrease in memory reconsolidation and extinction, as well as an increase in hippocampal glutamate levels. Although our data indicated a peripheral pro-inflammatory profile, we did not observe hippocampal neuroinflammatory features. Furthermore, we also observed that the HSB diet increased hippocampal fractalkine levels, a key chemokine associated with neuroprotection and inflammatory regulation. Then, we hypothesized that the elevation on glutamate levels may saturate synaptic communication, partially limiting plasticity, whereas fractalkine levels increase as a strategy to decrease glutamatergic damage.

FKN secreted by kidney epithelial cells regulates macrophage activation in lupus nephritis via the Hippo signaling pathway.

BACKGROUND: Lupus nephritis (LN) is a serious complication of systemic lupus erythematosus (SLE), and its pathogenesis is not fully understood. Previously, we showed that fractalkine (FKN) expression was positively correlated with the severity of LN. Here, we aimed to study the role of the Hippo signaling pathway (HSP) and its interaction with FKN in LN in an attempt to provide novel strategies for LN treatment. METHODS: In this study, lipopolysaccharide (LPS)/interferon-γ (IFN-γ)-stimulated THP-1 cells were co-cultured with FKN up-regulated or down-regulated kidney epithelial cells Hkb20. FKN-knockout (KO-FKN) mice were used to construct LN model. Flow cytometric analysis, quantitative real-time polymerase chain reaction (qRT-PCR), enzyme-linked immunosorbent assay (ELISA), pathological staining, Western blot, and immunofluorescence (IF) staining were employed to investigate the role of FKN and its interaction with the Hippo signaling pathway (HSP) in LN. RESULTS: Up-regulation of FKN in kidney epithelial cells was associated with increased macrophage activation. FKN overexpression in kidney epithelial cells suppressed apoptosis, inflammation levels, and M1 polarization of THP-1 cells and inhibited the HSP. Oppositely, FKN knockdown in kidney epithelial cells increased apoptosis, inflammation, and M1 polarization and activated the HSP. HSP inhibitor reversed the effect of FKN knockdown on THP-1 cells. In LN mice, FKN knockout and YAP inhibitor decreased the levels of renal function markers, alleviated kidney injury induced by LN, and inhibited macrophage activation in LN mice. CONCLUSIONS: FKN down-regulation reduced the activation of macrophages in renal tissue and alleviated kidney damage by activating HSP. The regulatory effect of FKN on HSP should be confirmed in patients with LN, and the mechanism of FKN in LN should be further explored.

Fractalkine Enhances Hematoma Resolution and Improves Neurological Function via CX3CR1/AMPK/PPARγ Pathway After GMH.

BACKGROUND: Hematoma clearance has been a proposed therapeutic strategy for hemorrhagic stroke. This study investigated the impact of CX3CR1 (CX3C chemokine receptor 1) activation mediated by r-FKN (recombinant fractalkine) on hematoma resolution, neuroinflammation, and the underlying mechanisms involving AMPK (AMP-activated protein kinase)/PPARγ (peroxisome proliferator-activated receptor gamma) pathway after experimental germinal matrix hemorrhage (GMH). METHODS: A total of 313 postnatal day 7 Sprague Dawley rat pups were used. GMH was induced using bacterial collagenase by a stereotactically guided infusion. r-FKN was administered intranasally at 1, 25, and 49 hours after GMH for short-term neurological evaluation. Long-term neurobehavioral tests (water maze, rotarod, and foot-fault test) were performed 24 to 28 days after GMH with the treatment of r-FKN once daily for 7 days. To elucidate the underlying mechanism, CX3CR1 CRISPR, or selective CX3CR1 inhibitor AZD8797, was administered intracerebroventricularly 24 hours preinduction of GMH. Selective inhibition of AMPK/PPARγ signaling in microglia via intracerebroventricularly delivery of liposome-encapsulated specific AMPK (Lipo-Dorsomorphin), PPARγ (Lipo-GW9662) inhibitor. Western blot, Immunofluorescence staining, Nissl staining, Hemoglobin assay, and ELISA assay were performed. RESULTS: The brain expression of FKN and CX3CR1 were elevated after GMH. FKN was expressed on both neurons and microglia, whereas CX3CR1 was mainly expressed on microglia after GMH. Intranasal administration of r-FKN improved the short- and long-term neurobehavioral deficits and promoted M2 microglia polarization, thereby attenuating neuroinflammation and enhancing hematoma clearance, which was accompanied by an increased ratio of p-AMPK (phosphorylation of AMPK)/AMPK, Nrf2 (nuclear factor erythroid 2-related factor 2), PPARγ, CD36 (cluster of differentiation 36), CD163 (hemoglobin scavenger receptor), CD206 (the mannose receptor), and IL (interleukin)-10 expression, and decreased CD68 (cluster of differentiation 68), IL-1ß, and TNF (tumor necrosis factor) α expression. The administration of CX3CR1 CRISPR or CX3CR1 inhibitor (AZD8797) abolished the protective effect of FKN. Furthermore, selective inhibition of microglial AMPK/PPARγ signaling abrogated the anti-inflammation effects of r-FKN after GMH. CONCLUSIONS: CX3CR1 activation by r-FKN promoted hematoma resolution, attenuated neuroinflammation, and neurological deficits partially through the AMPK/PPARγ signaling pathway, which promoted M1/M2 microglial polarization. Activating CX3CR1 by r-FKN may provide a promising therapeutic approach for treating patients with GMH.

Experimental traumatic occlusion drives immune changes in trigeminal ganglion.

We previously demonstrated that experimental traumatic occlusion (ETO) induces a long-lasting nociceptive response. These findings were associated with altered neuronal patterns and suggestive satellite glial cell activation. This study aimed to elucidate the activation of satellite glial cells following ETO in the trigeminal ganglion. Moreover, we explored the involvement of resident and infiltrating cells in trigeminal ganglion in ETO. Finally, we investigated the overexpression of purinergic signaling and the CX3CL1/CX3CR1 axis. RT-qPCR and electrophoresis showed overexpression of GFAP in the trigeminal ganglion (TG), and immunohistochemistry corroborated these findings, demonstrating SGCs activation. ELISA reveals enhanced levels of TNF-α and IL-1ß in TG after 28 d of ETO. In trigeminal ganglia, ETO groups improved the release of CX3CL1, and immunohistochemistry showed higher CX3CR1+ -immunoreactive cells in ETO groups. Immunohistochemistry and electrophoresis of the P2X7 receptor were found in ETO groups. The mRNA levels of IBA1 are upregulated in the 0.7-mm ETO group, while immunohistochemistry showed higher IBA1+ -immunoreactive cells in both ETO groups. The expression of CD68 by electrophoresis and immunohistochemistry was observed in the ETO groups. For last, ELISA revealed increased levels of IL-6, IL-12, and CCL2 in the TG of ETO groups. Furthermore, the mRNA expression revealed augmented transcription factors and cytokines associated with lymphocyte activation, such as RORγt, IL-17, Tbet, IFNγ, FOXP3, and IL-10. The findings of this study suggested that ETO activates SGCs in TG, and purinergic signaling and CX3CL1/CX3CR1 axis were upregulated. We uncovered the involvement of a distinct subtype of macrophages, named sensory neuron-associated macrophage activation (sNMAs), and detected an expanded number of infiltrated macrophages onto TG. These findings indicate that ETO induces chronic/persistent immune response.

Spinal Cathepsin S promotes visceral hypersensitivity via FKN/CX3CR1/p38 MAPK signaling pathways.

BACKGROUND: Irritable bowel syndrome (IBS) is one of the typical representatives of chronic functional visceral pain that lacks effective treatment. Recently, attention has been given to the role of microglia in IBS, particularly the activation of spinal microglia and the subsequent release of Cathepsin S (Cat S), a proteolytic enzyme. However, the specific role of spinal Cat S in IBS remains to be elucidated. The purpose of this study is to investigate the mechanisms underlying the regulation of visceral hypersensitivity in IBS-like rats by Cat S. METHODS: An IBS-like rat model was developed, and visceral sensitivity was tested via the electromyographic (EMG) response to colorectal distention (CRD) and pain threshold. Western blot and immunofluorescence were used to examine the expressions of proteins. The effects of inhibitors or neutralizing antibodies on visceral pain and the downstream molecular expressions were detected. The open-field test was performed to evaluate locomotor activity and anxiety-like behaviors in rats. RESULTS: We discovered that spinal Cat S was upregulated and colocalized with microglia in IBS-like rats. Treatment with LY3000328, a selective inhibitor of Cat S, dose-dependently down-regulated EMG amplitude and Fractalkine (FKN) expression, indicating that Cat S regulated visceral hypersensitivity via activating FKN in IBS-like rats. Furthermore, the expressions of FKN, CX3CR1, and p-p38 MAPK were elevated in IBS-like rats whereas inhibition of these molecules could alleviate visceral pain. Moreover, pharmacological inhibitor experiments suggested the activation of CX3CR1 by FKN facilitated p38 MAPK phosphorylation, which in turn promoted Cat S expression in IBS-like rats. CONCLUSIONS: Neonatal adverse stimulation might enhance the expression of spinal microglial Cat S, thereby activating the FKN/CX3CR1/p38 MAPK pathway and lead to visceral hypersensitivity in IBS-like rats. As a selective inhibitor of Cat S, LY3000328 could become a potential therapeutic option for IBS.

Use of novel structural features to identify urinary biomarkers during acute kidney injury that predict progression to chronic kidney disease.

BACKGROUND: A significant barrier to biomarker development in the field of acute kidney injury (AKI) is the use of kidney function to identify candidates. Progress in imaging technology makes it possible to detect early structural changes prior to a decline in kidney function. Early identification of those who will advance to chronic kidney disease (CKD) would allow for the initiation of interventions to halt progression. The goal of this study was to use a structural phenotype defined by magnetic resonance imaging and histology to advance biomarker discovery during the transition from AKI to CKD. METHODS: Urine was collected and analyzed from adult C57Bl/6 male mice at four days and 12 weeks after folic acid-induced AKI. Mice were euthanized 12 weeks after AKI and structural metrics were obtained from cationic ferritin-enhanced-MRI (CFE-MRI) and histologic assessment. The fraction of proximal tubules, number of atubular glomeruli (ATG), and area of scarring were measured histologically. The correlation between the urinary biomarkers at the AKI or CKD and CFE-MRI derived features was determined, alone or in combination with the histologic features, using principal components. RESULTS: Using principal components derived from structural features, twelve urinary proteins were identified at the time of AKI that predicted structural changes 12 weeks after injury. The raw and normalized urinary concentrations of IGFBP-3 and TNFRII strongly correlated to the structural findings from histology and CFE-MRI. Urinary fractalkine concentration at the time of CKD correlated with structural findings of CKD. CONCLUSIONS: We have used structural features to identify several candidate urinary proteins that predict whole kidney pathologic features during the transition from AKI to CKD, including IGFBP-3, TNFRII, and fractalkine. In future work, these biomarkers must be corroborated in patient cohorts to determine their suitability to predict CKD after AKI.

The Role of Fractalkine in the Regulation of Endometrial Iron Metabolism in Iron Deficiency.

Iron is a crucial element in the human body. Endometrial iron metabolism is implicated in endometrium receptivity and embryo implantation. Disturbances of the maternal as well as the endometrial iron homeostasis, such as iron deficiency, can contribute to the reduced development of the fetus and could cause an increased risk of adverse pregnancy outcomes. Fractalkine is a unique chemokine that plays a role in the communication between the mother and the fetus. It has been demonstrated that FKN is involved in the development of endometrial receptivity and embryo implantation, and it functions as a regulator of iron metabolism. In the present study, we examined the effect of FKN on the iron metabolism of HEC-1A endometrial cells in a state of iron deficiency mediated by desferrioxamine treatment. Based on the findings, FKN enhances the expression of iron metabolism-related genes in iron deficiency and modifies the iron uptake via transferrin receptor 1 and divalent metal transporter-1, and iron release via ferroportin. FKN can activate the release of iron from heme-containing proteins by elevating the level of heme oxygenase-1, contributing to the redistribution of intracellular iron content. It was revealed that the endometrium cells express both mitoferrin-1 and 2 and that their levels are not dependent on the iron availability of the cells. FKN may also contribute to maintaining mitochondrial iron homeostasis. FKN can improve the deteriorating effect of iron deficiency in HEC-1A endometrium cells, which may contribute to the development of receptivity and/or provide iron delivery towards the embryo.

The Cytokine CX3CL1 and ADAMs/MMPs in Concerted Cross-Talk Influencing Neurodegenerative Diseases.

Neuroinflammation plays a fundamental role in the development and progression of neurodegenerative diseases. It could therefore be said that neuroinflammation in neurodegenerative pathologies is not a consequence but a cause of them and could represent a therapeutic target of neuronal degeneration. CX3CL1 and several proteases (ADAMs/MMPs) are strongly involved in the inflammatory pathways of these neurodegenerative pathologies with multiple effects. On the one hand, ADAMs have neuroprotective and anti-apoptotic effects; on the other hand, they target cytokines and chemokines, thus causing inflammatory processes and, consequently, neurodegeneration. CX3CL1 itself is a cytokine substrate for the ADAM, ADAM17, which cleaves and releases it in a soluble isoform (sCX3CL1). CX3CL1, as an adhesion molecule, on the one hand, plays an inhibiting role in the pro-inflammatory response in the central nervous system (CNS) and shows neuroprotective effects by binding its membrane receptor (CX3CR1) present into microglia cells and maintaining them in a quiescent state; on the other hand, the sCX3CL1 isoform seems to promote neurodegeneration. In this review, the dual roles of CX3CL1 and ADAMs/MMPs in different neurodegenerative diseases, such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (MH), and multiple sclerosis (MS), are investigated.

Bujing Yishi tablets alleviate photoreceptor cells death via the P2X7R/CX3CL1/CX3CR1 pathway in Retinitis Pigmentosa rats.

BACKGROUND: Retinitis pigmentosa (RP) refers to a group of progressive photoreceptor degenerative diseases. The activation of microglia has been reported to play an important role in the photoreceptor degeneration in RP retinal. Bujing Yishi tablets (BJYS), a Chinese herbal medicine, has been used to treat retinal diseases in China with desirable effect in improving visual function. However, the mechanisms underlying the efficacy of BJYS treatment in RP are not yet fully understood. PURPOSE: Based on the preliminary experiments, this study aimed to investigate the therapeutic mechanism involved in treating N-Methyl-N-Nitrosourea (MNU)-induced retinal degeneration of RP with BJYS. METHODS: To explore the efficacy of BJYS, a rat experimental RP model was established through intraperitoneal injection of MNU (50 mg/kg). Two experiment was carried out. After the treatment, we conducted H&E, TUNEL, retinal cytokine levels and IBA-1 expression in microglia to confirm the impact on RP model. The specific mechanism of action of BJYS tablet was assessed by western blot, real-time polymerase chain reaction (RT-PCR), and immunofluorescence to determine the mRNA and protein expression levels involved in clarifying the effectiveness of BJYS exerted through P2X7R/CX3CL1/CX3CR1 pathway. RESULTS: Significant alleviation of retinal morphological structure and photoreceptor degeneration by BJYS treatment was observed in the retinal of MNU-induced RP rats, BJYS prevented the reduction of ONL thickness and decreased the level of apoptotic cells in ONL. It also inhibited microglia overactivation and reduced retinal levels of IL-1ß, IL-6, TNF-α. In addition, BJYS decreased the protein expression and mRNA expression of P2X7, CX3CL1 and CX3CR1 and reduced the phosphorylation of p38 MAPK. CONCLUSION: In summary, this study suggested that BJYS treatment could alleviate photoreceptors degeneration of RP by inhibiting microglia overactivation and inflammation through the P2X7R/CX3CL1/CX3CR1 pathway. These effects suggest that BJYS tablets may serve as a promising oral therapeutic agent for RP.

CX3CL1 induces cell migration and invasion through ICAM-1 expression in oral squamous cell carcinoma cells.

Human oral squamous cell carcinoma (OSCC) has been associated with a relatively low survival rate over the years and is characterized by a poor prognosis. C-X3-C motif chemokine ligand 1 (CX3CL1) has been involved in advanced migratory cells. Overexpressed CX3CL1 promotes several cellular responses related to cancer metastasis, including cell movement, migration and invasion in tumour cells. However, CX3CL1 controls the migration ability, and its molecular mechanism in OSCC remains unknown. The present study confirmed that CX3CL1 increased cell movement, migration and invasion. The CX3CL1-induced cell motility is upregulated through intercellular adhesion molecule-1 (ICAM-1) expression in OSCC cells. These effects were significantly suppressed when OSCC cells were pre-treated with CX3CR1 monoclonal antibody (mAb) and small-interfering RNA (siRNA). The CX3CL1-CX3CR1 axis activates promoted PLCß/PKCα/c-Src phosphorylation. Furthermore, CX3CL1 enhanced activator protein-1 (AP-1) activity. The CX3CR1 mAb and PLCß, PKCα, c-Src inhibitors reduced CX3CL1-induced c-Jun phosphorylation, c-Jun translocation into the nucleus and c-Jun binding to the ICAM-1 promoter. The present results reveal that CX3CL1 induces the migration of OSCC cells by promoting ICAM-1 expression through the CX3CR1 and the PLCß/PKCα/c-Src signal pathway, suggesting that CX3CL1-CX3CR1-mediated signalling is correlated with tumour motility and appealed to be a precursor for prognosis in human OSCC.

Spinal Cord Stimulation Increases Chemoefficacy and Prevents Paclitaxel-Induced Pain via CX3CL1.

INTRODUCTION: Despite increasing utilization of spinal cord stimulation (SCS), its effects on chemoefficacy, cancer progression, and chemotherapy-induced peripheral neuropathy (CIPN) pain remain unclear. Up to 30% of adults who are cancer survivors may suffer from CIPN, and there are currently no effective preventative treatments. MATERIALS AND METHODS: Through a combination of bioluminescent imaging, behavioral, biochemical, and immunohistochemical approaches, we investigated the role of SCS and paclitaxel (PTX) on tumor growth and PTX-induced peripheral neuropathy (PIPN) pain development in T-cell-deficient male rats (Crl:NIH-Foxn1rnu) with xenograft human non-small cell lung cancer. We hypothesized that SCS can prevent CIPN pain and enhance chemoefficacy partially by modulating macrophages, fractalkine (CX3CL1), and inflammatory cytokines. RESULTS: We show that preemptive SCS enhanced the antitumor efficacy of PTX and prevented PIPN pain. Without SCS, rats with and without tumors developed robust PIPN pain-related mechanical hypersensitivity, but only those with tumors developed cold hypersensitivity, suggesting T-cell dependence for different PIPN pain modalities. SCS increased soluble CX3CL1 and macrophages and decreased neuronal and nonneuronal insoluble CX3CL1 expression and inflammation in dorsal root ganglia. CONCLUSION: Collectively, our findings suggest that preemptive SCS is a promising strategy to increase chemoefficacy and prevent PIPN pain via CX3CL1-macrophage modulation.

Effects of CX3CR1 and CXCR2 antagonists on running-dependent intramuscular neutrophil recruitments and myokine upregulation.

Muscle contractile activity stimulates intramuscular recruitment of immune cells including neutrophils emerging to serve as a prerequisite for exerting proper muscular performance, although the underlying mechanisms and their contributions to myokine upregulation remain ill-defined. We previously reported that pharmacological inhibition of CX3CR1, a fractalkine receptor, dampens gnawing-dependent neutrophil recruitment into masseter muscles along with compromising their masticatory activity. By using a running exercise model, we herein demonstrated that hindlimb muscles require collaborative actions of both CX3CR1- and CXCR2-mediated signals for achieving neutrophil recruitment, upregulation of myokines including interleukin (IL)-6, enhanced GLUT4 translocation, and adequate endurance capability. Mechanistically, we revealed that a combination of CX3CR1 and CXCR2 antagonists, i.e., AZD8797 and SB2205002, inhibits exercise-inducible ICAM-1 and fractalkine upregulations in the area of the endothelium and muscle-derived CXCL1 upregulation, both of which apparently contribute to the intramuscular neutrophil accumulation in working muscles. Intriguingly, we also observed that 2 h of running results in intramuscular augmentation of innate lymphoid type 2 cells (ILC2s) markers, i.e., Bcl11b mRNA levels and anti-GATA-3-antibody-positive signals, and that these effects are completely abolished by administration of the combination of CX3CR1 and CXCR2 antagonists. Taken together, our findings strongly suggest that the exercise-evoked regional interplay among working myofibers, the adjacent endothelium, and recruited immune cells including neutrophils and possibly ILC2s, mediated through these local factors, plays a key role in the organization of the intramuscular microenvironment supporting the performance of hindlimb muscles during running.NEW & NOTEWORTHY This study provides compelling evidence that running-dependent intramuscular neutrophil recruitment requires both CX3CR1- and CXCR2-mediated signals that prime not only myofiber-derived myokine upregulations but also endothelium ICAM-1 and fractalkine expressions. The results revealed the importance of the exercise-evoked regional interplay among working myofibers, the adjacent endothelium, and recruited immune cells, including neutrophils and possibly ILC2s, which plays a key role in the organization of the intramuscular microenvironment supporting the performance of hindlimb muscles during running.