TRPM2-dependent autophagy inhibition exacerbates oxidative stress-induced CXCL16 secretion by keratinocytes in vitiligo.

Vitiligo is a depigmented skin disease due to the destruction of melanocytes. Under oxidative stress, keratinocyte-derived chemokine C-X-C motif ligand 16 (CXCL16) plays a critical role in recruiting CD8+ T cells, which kill melanocytes. Autophagy serves as a protective cell survival mechanism and impairment of autophagy has been linked to increased secretion of the proinflammatory cytokines. However, the role of autophagy in the secretion of CXCL16 under oxidative stress has not been investigated. Herein, we initially found that autophagy was suppressed in both keratinocytes of vitiligo lesions and keratinocytes exposed to oxidative stress in vitro. Autophagy inhibition also promoted CXCL16 secretion. Furthermore, upregulated transient receptor potential cation channel subfamily M member 2 (TRPM2) functioned as an upstream oxidative stress sensor to inhibit autophagy. Moreover, TRPM2-mediated Ca2+ influx activated calpain to shear autophagy related 5 (Atg5) and Atg12-Atg5 conjugate formation was blocked to inhibit autophagy under oxidative stress. More importantly, Atg5 downregulation enhanced the binding of interferon regulatory factor 3 (IRF3) to the CXCL16 promoter region by activating Tank-binding kinase 1 (TBK1), thus promoting CXCL16 secretion. These findings suggested that TRPM2-restrained autophagy promotes CXCL16 secretion via the Atg5-TBK1-IRF3 signaling pathway under oxidative stress. Inhibition of TRPM2 may serve as a potential target for the treatment of vitiligo. © 2024 The Pathological Society of Great Britain and Ireland.

Identification of CXCL16 as a diagnostic biomarker for obesity and intervertebral disc degeneration based on machine learning.

Intervertebral disc degeneration (IDD) is the primary cause of neck and back pain. Obesity has been established as a significant risk factor for IDD. The objective of this study was to explore the molecular mechanisms affecting obesity and IDD by identifying the overlapping crosstalk genes associated with both conditions. The identification of specific diagnostic biomarkers for obesity and IDD would have crucial clinical implications. We obtained gene expression profiles of GSE70362 and GSE152991 from the Gene Expression Omnibus, followed by their analysis using two machine learning algorithms, least absolute shrinkage and selection operator and support vector machine-recursive feature elimination, which enabled the identification of C-X-C motif chemokine ligand 16 (CXCL16) as a shared diagnostic biomarker for obesity and IDD. Additionally, gene set variant analysis was used to explore the potential mechanism of CXCL16 in these diseases, and CXCL16 was found to affect IDD through its effect on fatty acid metabolism. Furthermore, correlation analysis between CXCL16 and immune cells demonstrated that CXCL16 negatively regulated T helper 17 cells to promote IDD. Finally, independent external datasets (GSE124272 and GSE59034) were used to verify the diagnostic efficacy of CXCL16. In conclusion, a common diagnostic biomarker for obesity and IDD, CXCL16, was identified using a machine learning algorithm. This study provides a new perspective for exploring the possible mechanisms by which obesity impacts the development of IDD.

ELK3-CXCL16 axis determines natural killer cell cytotoxicity via the chemotactic activity of CXCL16 in triple negative breast cancer.

Triple-negative breast cancer (TNBC) is the most challenging subtype of breast cancer because of its aggressive behavior and the limited therapeutic strategies available. In the last decade, immunotherapy has become a promising treatment to prolong survival in advanced solid cancers including TNBC. However, the efficacy of immunotherapy in solid cancers remains limited because solid tumors contain few tumor-infiltrating lymphocytes. Here, we show that targeting an ETS transcription factor ELK3 (ELK3) recruits immune cells including natural killer (NK) cells into tumors via the chemotactic activity of chemokine. ELK3 depletion increases CXCL16 expression level and promotes NK cell cytotoxicity through CXCL16-mediated NK cell recruitment in TNBC. In silico analysis showed that ELK3 is negatively correlated with CXCL16 expression in breast cancer patient samples. Low expression of ELK3 and high expression of CXCL16 were associated with a better prognosis. Low expression of ELK3 and high expression of CXCL16 were associated with increased expression of NK cell-related genes. Our findings demonstrate that the ELK3-CXCL16 axis modulates NK cell recruitment to increase NK cell cytotoxicity, suggesting that targeting the ELK3 gene could be an adjuvant strategy for increasing the efficacy of immunotherapy in TNBC.

Convergent transcriptomic and genomic evidence supporting a dysregulation of CXCL16 and CCL5 in Alzheimer's disease.

BACKGROUND: Neuroinflammatory factors, especially chemokines, have been widely reported to be involved in the pathogenesis of Alzheimer's disease (AD). It is unclear how chemokines are altered in AD, and whether dysregulation of chemokines is the cause, or the consequence, of the disease. METHODS: We initially screened the transcriptomic profiles of chemokines from publicly available datasets of brain tissues of AD patients and mouse models. Expression alteration of chemokines in the blood from AD patients was also measured to explore whether any chemokine might be used as a potential biomarker for AD. We further analyzed the association between the coding variants of chemokine genes and genetic susceptibility of AD by targeted sequencing of a Han Chinese case-control cohort. Mendelian randomization (MR) was performed to infer the causal association of chemokine dysregulation with AD development. RESULTS: Three chemokine genes (CCL5, CXCL1, and CXCL16) were consistently upregulated in brain tissues from AD patients and the mouse models and were positively correlated with Aß and tau pathology in AD mice. Peripheral blood mRNA expression of CXCL16 was upregulated in mild cognitive impairment (MCI) and AD patients, indicating the potential of CXCL16 as a biomarker for AD development. None of the coding variants within any chemokine gene conferred a genetic risk to AD. MR analysis confirmed a causal role of CCL5 dysregulation in AD mediated by trans-regulatory variants. CONCLUSIONS: In summary, we have provided transcriptomic and genomic evidence supporting an active role of dysregulated CXCL16 and CCL5 during AD development.

CXCR6 Mediates Pressure Overload-Induced Aortic Stiffness by Increasing Macrophage Recruitment and Reducing Exosome-miRNA29b.

Aortic stiffness is an independent risk factor for aortic diseases such as aortic dissection which commonly occurred with aging and hypertension. Chemokine receptor CXCR6 is critically involved in vascular inflammation and remodeling. Here, we investigated whether and how CXCR6 plays a role in aortic stiffness caused by pressure overload. CXCR6-/- and WT mice underwent transverse aortic constriction (TAC) surgery for 8 weeks. CXCR6 deficiency significantly improved TAC-induced aortic remodeling and endothelial dysfunction by decreasing CD11c+ macrophage infiltration, suppressing VCAM-1 and ICAM-1, reducing collagen deposition, and downregulating MMP12 and osteopontin in the aorta. Consistently, blocking the CXCL16/CXCR6 axis also reduced aortic accumulation of CD11c+ macrophages and vascular stiffness but without affecting the release of TNF-α and IL-6 from the aorta. Furthermore, pressure overload inhibited aortic release of exosomes, which could be reversed by suppressing CXCR6 or CXCL16. Inhibition of exosome release by GW4869 significantly aggravated TAC-induced aortic calcification and stiffness. By exosomal microRNA microarray analysis, we found that microRNA-29b was significantly reduced in aortic endothelial cells (AECs) receiving TAC. Intriguingly, blocking the CXCL16/CXCR6 axis restored the expression of miR-29b in AECs. Finally, overexpression of miR-29b significantly increased eNOS and reduced MMPs and collagen in AECs. By contrast, antagonizing miR-29b in vivo further enhanced TAC-induced expressions of MMP12 and osteopontin, aggravated aortic fibrosis, calcification, and stiffness. Our study demonstrated a key role of the CXCL16/CXCR6 axis in macrophage recruitment and macrophage-mediated aortic stiffness under pressure overload through an exosome-miRNAs-dependent manner.

CXCL16 inhibits epithelial regeneration and promotes fibrosis during the progression of radiation enteritis.

Radiation enteritis (RE) is a prevalent complication of radiotherapy for pelvic malignant tumors, characterized by severe intestinal epithelial destruction and progressive submucosal fibrosis. However, little is known about the pathogenesis of this disease, and so far, there is no specific targeted therapy. Here, we report that CXCL16 is upregulated in the injured intestinal tissues of RE patients and in a mouse model. Genetic deletion of Cxcl16 mitigates fibrosis and promotes intestinal stem cell-mediated epithelial regeneration after radiation injury in mice. Mechanistically, CXCL16 functions on myofibroblasts through its receptor CXCR6 and activates JAK3/STAT3 signaling to promote fibrosis and, at the same time, to transcriptionally modulate the levels of BMP4 and hepatocyte growth factor (HGF) in myofibroblasts. Moreover, we find that CXCL16 and CXCR6 auto- and cross-regulate themselves in positive feedback loops. Treatment with CXCL16 neutralizing monoclonal antibody attenuates fibrosis and improves the epithelial repair in RE mouse model. Our findings emphasize the important role of CXCL16 in the progression of RE and suggest that CXCL16 signaling could be a potential therapeutic target for RE. © 2022 The Pathological Society of Great Britain and Ireland.

The CXCL16-CXCR6 axis in glioblastoma modulates T-cell activity in a spatiotemporal context.

Introduction: Glioblastoma multiforme (GBM) pathobiology is characterized by its significant induction of immunosuppression within the tumor microenvironment, predominantly mediated by immunosuppressive tumor-associated myeloid cells (TAMCs). Myeloid cells play a pivotal role in shaping the GBM microenvironment and influencing immune responses, with direct interactions with effector immune cells critically impacting these processes. Methods: Our study investigates the role of the CXCR6/CXCL16 axis in T-cell myeloid interactions within GBM tissues. We examined the surface expression of CXCL16, revealing its limitation to TAMCs, while microglia release CXCL16 as a cytokine. The study explores how these distinct expression patterns affect T-cell engagement, focusing on the consequences for T-cell function within the tumor environment. Additionally, we assessed the significance of CXCR6 expression in T-cell activation and the initial migration to tumor tissues. Results: Our data demonstrates that CXCL16 surface expression on TAMCs results in predominant T-cell engagement with these cells, leading to impaired T-cell function within the tumor environment. Conversely, our findings highlight the essential role of CXCR6 expression in facilitating T-cell activation and initial migration to tumor tissues. The CXCL16-CXCR6 axis exhibits dualistic characteristics, facilitating the early stages of the T-cell immune response and promoting T-cell infiltration into tumors. However, once inside the tumor, this axis contributes to immunosuppression. Discussion: The dual nature of the CXCL16-CXCR6 axis underscores its potential as a therapeutic target in GBM. However, our results emphasize the importance of carefully considering the timing and context of intervention. While targeting this axis holds promise in combating GBM, the complex interplay between TAMCs, microglia, and T cells suggests that intervention strategies need to be tailored to optimize the balance between promoting antitumor immunity and preventing immunosuppression within the dynamic tumor microenvironment.

Single-cell RNA transcriptome analysis of CNS immune cells reveals CXCL16/CXCR6 as maintenance factors for tissue-resident T cells that drive synapse elimination.

BACKGROUND: Emerging RNA viruses that target the central nervous system (CNS) lead to cognitive sequelae in survivors. Studies in humans and mice infected with West Nile virus (WNV), a re-emerging RNA virus associated with learning and memory deficits, revealed microglial-mediated synapse elimination within the hippocampus. Moreover, CNS-resident memory T (TRM) cells activate microglia, limiting synapse recovery and inducing spatial learning defects in WNV-recovered mice. The signals involved in T cell-microglia interactions are unknown. METHODS: Here, we examined immune cells within the murine WNV-recovered forebrain using single-cell RNA sequencing to identify putative ligand-receptor pairs involved in intercellular communication between T cells and microglia. Clustering and differential gene analyses were followed by protein validation and genetic and antibody-based approaches utilizing an established murine model of WNV recovery in which microglia and complement promote ongoing hippocampal synaptic loss. RESULTS: Profiling of host transcriptome immune cells at 25 days post-infection in mice revealed a shift in forebrain homeostatic microglia to activated subpopulations with transcriptional signatures that have previously been observed in studies of neurodegenerative diseases. Importantly, CXCL16/CXCR6, a chemokine signaling pathway involved in TRM cell biology, was identified as critically regulating CXCR6 expressing CD8+ TRM cell numbers within the WNV-recovered forebrain. We demonstrate that CXCL16 is highly expressed by all myeloid cells, and its unique receptor, CXCR6, is highly expressed on all CD8+ T cells. Using genetic and pharmacological approaches, we demonstrate that CXCL16/CXCR6 not only is required for the maintenance of WNV-specific CD8 TRM cells in the post-infectious CNS, but also contributes to their expression of TRM cell markers. Moreover, CXCR6+CD8+ T cells are required for glial activation and ongoing synapse elimination. CONCLUSIONS: We provide a comprehensive assessment of the role of CXCL16/CXCR6 as an interaction link between microglia and CD8+ T cells that maintains forebrain TRM cells, microglial and astrocyte activation, and ongoing synapse elimination in virally recovered animals. We also show that therapeutic targeting of CXCL16 in mice during recovery may reduce CNS CD8+ TRM cells.

Metabolic Syndrome in Psoriasis Is Associated With Upregulation of CXCL16 on Monocytes and a Dysbalance in Innate Lymphoid Cells.

Psoriasis is frequently associated with the metabolic syndrome and occurs more often in obese individuals. In order to understand innate immune mechanisms mediating this inflammatory pattern we investigated expression of the chemokine and lipid scavenger receptor CXCL16 in patients with psoriasis and associated comorbidities. CXCL16 expression was enhanced on all monocyte subsets in psoriatic patients compared with healthy controls and positively correlated with psoriasis activity and severity index, body mass index and the risk for cardiovascular disease indicated by PROCAM score. The intensity of CXCL16 expression on monocytes further correlated with their capability to phagocytose oxidized LDL indicating the possibility to transform into foam cells in atherosclerotic plaques. Patients with psoriasis and atherosclerosis or obesity displayed elevated numbers of innate lymphoid cells in blood with specific increase of the IFN-γ or IL-17 producing ILC1 and ILC3 subpopulations. The expression of the CXCL16 receptor, CXCR6, was increased in ILCs and co-expressed with CCR6 but not CCR7 indicating their migratory potential to psoriatic skin or adipose tissue that is characterized by strong CXCL16 and CCL20 expression. This hypothesis was supported by the finding that the percentage of CXCR6 expressing ILCs was alleviated in blood of psoriatic patients. Together these data link a strong expression of CXCL16 to metabolic syndrome in psoriasis and indicate a possible link to ILC activation and tissue distribution in obese psoriatic patients. These data contribute to the understanding of the complex interaction of innate immunity and metabolic state in psoriasis.

CXCL16/ERK1/2 pathway regulates human podocytes growth, migration, apoptosis and epithelial mesenchymal transition.

Primary nephrotic syndrome (PNS) is the commonest glomerular disease affecting children. Previous studies have confirmed that CXC motif chemokine ligand 16 (CXCL16) is involved in the pathogenesis of PNS. However, the exact mechanisms underlying the pathogenesis of PNS remain to be elucidated. Thus, the present study aimed to elucidate the role of CXCL16 in PNS. It was found that the expression of CXCL16 and extracellular signal­regulated kinases 1 and 2 (ERK1/2) were significantly increased in clinical PNS renal tissues using reverse transcription­quantitative PCR, western blot analysis and immunohistochemistry. Lentivirus overexpression or short hairpin RNA vector was used to induce the overexpression or knockdown of CXCL16 in podocytes, respectively. Overexpression of CXCL16 in podocytes could decrease the cell proliferation and increase the migration and apoptosis, whereas CXCL16 knockdown increased cell proliferation and decreased cell migration and apoptosis. Results of the present study further demonstrated that ERK2 protein expression was regulated by CXCL16. The knockdown of ERK2 expression reversed the effects of CXCL16 on the proliferation, apoptosis, migration and epithelial mesenchymal transition (EMT) of podocytes. Collectively, the findings of the present study highlighted that the CXCL16/ERK1/2 pathway regulates the growth, migration, apoptosis and EMT of human podocytes.

JNK and Jag1/Notch2 co-regulate CXCL16 to facilitate cypermethrin-induced kidney damage.

Cypermethrin (CYP), a widely-used composite pyrethroid pesticide, has underlying nephrotoxic effects. To elucidate potential roles of the MAPK pathway, the Jag/Notch pathway, and miRNAs in CYP-mediated kidney lesion, Sprague-Dawley rats and glomerular mesangial cells were used in this work. Results displayed that ß-CYP abnormally altered renal histomorphology and ultrastructures, induced renal DNA damage, and impaired renal functions, as evidenced by the increase in plasma levels of Cys-C and ß2-Mg. ß-CYP activated the JNK/c-Jun pathway by inducing ROS and oxidative stress. Meanwhile, ß-CYP changed the miRNA expression profile, miR-21-5p showing the most significant increase. Moreover, the Jag1/Notch2/Hes1 pathway was directly targeted by miR-21-5p, the mRNA and protein expression of Jag1, Notch2, and Hes1 being declined in vivo and in vitro. The chemokine CXCL16 was induced by ß-CYP, accompanied by the inflammatory factor production and inflammatory cell infiltration in kidneys. The specific JNK inhibitor, Jag1 overexpression, Hes1 overexpression, bidirectional Co-IP, ChIP, and CXCL16 silencing demonstrated that CXCL16 co-regulated by the JNK/c-Jun and Jag1/Notch2/Hes1 pathways elicited renal inflammation. Collectively, our findings indicate that ß-CYP is of nephrotoxicity and it not only directly changes renal histomorphology and ultrastructures, but induces CXCL16 to trigger renal inflammation via the JNK/c-Jun and Jag1/Notch2/Hes1 pathways, finally synergistically contributing to kidney damage.

Autonomous and intercellular chemokine signaling elicited from mesenchymal stem cells regulates migration of undifferentiated gastric cancer cells.

Accumulating evidence demonstrates that bone marrow (BM)-derived mesenchymal stem cells (MSCs) play critical roles in regulating progression of various types of cancer. We have previously shown that Wnt5a-Ror2 signaling in MSCs induces expression of CXCL16, and that CXCL16 secreted from MSCs then binds to its cognate receptor CXCR6 on the surface of an undifferentiated gastric cancer cell line MKN45 cells, eventually leading to proliferation and migration of MKN45 cells. However, it remains unclear about a possible involvement of another (other) cytokine(s) in regulating progression of gastric cancer. Here, we show that CXCL16-CXCR6 signaling is also activated in MSCs through cell-autonomous machinery, leading to upregulated expression of CCL5. We further show that CCR1 and CCR3, receptors of CCL5, are expressed on the surface of MKN45 cells, and that CCL5 secreted from MSCs promotes migration of MKN45 cells presumably via its binding to CCR1/CCR3. These data indicate that cell-autonomous CXCL16-CXCR6 signaling activated in MSCs upregulates expression of CCL5, and that subsequent activation of CCL5-CCR1/3 signaling in MKN45 cells through intercellular machinery can promote migration of MKN45 cells. Collectively, these findings postulate the presence of orchestrated chemokine signaling emanated from MSCs to regulate progression of undifferentiated gastric cancer cells.

Clinical significance and role of CXCL16 in anti-neutrophil cytoplasmic autoantibody-associated vasculitis.

Anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis (AAV) is a group of potentially life-threatening autoimmune diseases. The kidney and lung are the most common and most severely affected organs. Previous studies have shown that the chemokine ligand CXCL16 and its receptor CXCR6 play an important role in kidney disease. However, whether CXCL16/CXCR6 is involved in the pathogenesis of AAV remains elusive. In this study, the levels of CXCL16 and its specific receptor CXCR6 were investigated. According to kidney outcome, patients were divided into two groups, specifically one with high CXCL16 levels and one with low CXCL16 levels, by cut-off values using receiver operating characteristic (ROC) curves. The clinical parameters and histological features were further compared between the two groups. The ability of CXCL16 to induce neutrophil chemotaxis was analysed using a Transwell migration assay in a coculture system of conditional immortalized human glomerular endothelial cells (ciGEnCs) and neutrophils. We observed that the levels of CXCL16 were significantly increased in the circulation, along with the expression in renal tissue of AAV patients compared to healthy controls (HCs). CXCR6 expression on neutrophils was significantly higher in patients with AAV than in HCs. There were positive correlations between the levels of CXCL16 and serum creatinine, IL-6, CRP, and TNF-α and negative correlations with eGFR. The serum levels of CXCL16 could act as a predictive biomarker of renal outcome in AAV. CXCL16 secretion was upregulated in ciGEnCs treated with AAV serum. CXCL16 released from ciGEnCs contributed to neutrophil migration. Furthermore, neutrophil migration was attenuated by silencing CXCL16 expression via transfection with short hairpin RNA (shRNA) sequences and lentivirus. Taken together, these data suggest that the inhibition of the CXCL16/CXCR6 axis may provide new therapeutic strategies targeting AAV.

Beneficial effects of PCSK9 inhibition with alirocumab in familial hypercholesterolemia involve modulation of new immune players.

Familial hypercholesterolemia (FH) is associated with low-grade systemic inflammation, a key driver of premature atherosclerosis. We investigated the effects of inhibiting proprotein convertase subtilisin/kexin type 9 (PCSK9) function on inflammatory state, endothelial dysfunction and cardiovascular outcomes in patients with FH. Fourteen patients with FH were evaluated before and 8 weeks after administration of a PCSK9 blocking monoclonal antibody (alirocumab, 150 mg/subcutaneous/14 days). In vivo and ex vivo analysis revealed that alirocumab blunted the attachment of leukocytes to TNFα-stimulated human umbilical arterial endothelial cells (HUAEC) and suppressed the activation of platelets and most leukocyte subsets, which was accompanied by the diminished expression of CX3CR1, CXCR6 and CCR2 on several leukocyte subpopulations. By contrast, T-regulatory cell activation was enhanced by alirocumab treatment, which also elevated anti-inflammatory IL-10 plasma levels and lowered circulating pro-inflammatory cytokines. Plasma levels of IFNγ positively correlated with levels of total and LDL-cholesterol, whereas circulating IL-10 levels negatively correlated with these key lipid parameters. In vitro analysis revealed that TNFα stimulation of HUAEC increased the expression of PCSK9, whereas endothelial PCSK9 silencing reduced TNFα-induced mononuclear cell adhesion mediated by Nox5 up-regulation and p38-MAPK/NFκB activation, concomitant with reduced SREBP2 expression. PCSK9 silencing also decreased endothelial CX3CL1 and CXCL16 expression and chemokine generation. In conclusion, PCSK9 inhibition impairs systemic inflammation and endothelial dysfunction by constraining leukocyte-endothelium interactions. PCSK9 blockade may constitute a new therapeutic approach to control the inflammatory state associated with FH, preventing further cardiovascular events in this cardiometabolic disorder.

CXCL16/CXCR6 Axis in Adipocytes Differentiated from Human Adipose Derived Mesenchymal Stem Cells Regulates Macrophage Polarization.

Adipocytes interact with adipose tissue macrophages (ATMs) that exist as a form of M2 macrophage in healthy adipose tissue and are polarized into M1 macrophages upon cellular stress. ATMs regulate adipose tissue inflammation by secreting cytokines, adipokines, and chemokines. CXC-motif receptor 6 (CXCR6) is the chemokine receptor and interactions with its specific ligand CXC-motif chemokine ligand 16 (CXCL16) modulate the migratory capacities of human adipose-derived mesenchymal stem cells (hADMSCs). CXCR6 is highly expressed on differentiated adipocytes that are non-migratory cells. To evaluate the underlying mechanisms of CXCR6 in adipocytes, THP-1 human monocytes that can be polarized into M1 or M2 macrophages were co-cultured with adipocytes. As results, expression levels of the M1 polarization-inducing factor were decreased, while those of the M2 polarization-inducing factor were significantly increased in differentiated adipocytes in a co-cultured environment with additional CXCL16 treatment. After CXCL16 treatment, the anti-inflammatory factors, including p38 MAPK ad ERK1/2, were upregulated, while the pro-inflammatory pathway mediated by Akt and NF-κB was downregulated in adipocytes in a co-cultured environment. These results revealed that the CXCL16/CXCR6 axis in adipocytes regulates M1 or M2 polarization and displays an immunosuppressive effect by modulating pro-inflammatory or anti-inflammatory pathways. Our results may provide an insight into a potential target as a regulator of the immune response via the CXCL16/CXCR6 axis in adipocytes.

Interleukin-18 accelerates cardiac inflammation and dysfunction during ischemia/reperfusion injury by transcriptional activation of CXCL16.

Myocardial ischemia/reperfusion(I/R) injury elicits an inflammatory response that drives tissue damage and cardiac remodeling. The trafficking and recruitment of inflammatory cells are controlled by C-X-C motif chemokine ligands and their receptors. CXCL16, a hallmark of acute coronary syndromes, is responsible for the recruitment of macrophages, monocytes and T lymphocytes. However, its role in cardiac I/R injury remains poorly characterized. Here we reported that CXCL16-mediated cardiac infiltration of CD11b+Ly6C+ cells played a crucial role in IL-18-induced myocardial inflammation, apoptosis and left ventricular(LV) dysfunction during I/R. Treatment with CXCL16 shRNA attenuated I/R-induced cardiac injury, LV remodeling and cardiac inflammation by reducing the recruitment of inflammatory cells and the release of TNFα, IL-17 and IFN-γ in the heart. We found that I/R-mediated NLRP3/IL-18 signaling pathway triggered CXCL16 transcription in cardiac vascular endothelial cells(VECs). Two binding sites of FOXO3 were found at the promoter region of CXCL16. By luciferase report assay and ChIP analysis, we confirmed that FOXO3 was responsible for endothelial CXCL16 transcription. A pronounced reduction of CXCL16 was observed in FOXO3 siRNA pretreated-VECs. Further experiments revealed that IL-18 activated FOXO3 by promoting the phosphorylation of STAT3 but not STAT4. An interaction between FOXO3 and STAT3 enhanced the transcription of CXCL16 induced by FOXO3. Treatment with Anakinra or Stattic either effectively inhibited IL-18-mediated nuclear import of FOXO3 and CXCL16 transcription. Our findings suggested that IL-18 accelerated I/R-induced cardiac damage and dysfunction through activating CXCL-16 and CXCL16-mediated cardiac infiltration of the CD11b+Ly6C+ cells. CXCL16 might be a novel therapeutic target for the treatment of I/R-related ischemic heart diseases.

CXCL16 Promotes Gastric Cancer Tumorigenesis via ADAM10-Dependent CXCL16/CXCR6 Axis and Activates Akt and MAPK Signaling Pathways.

Abnormal expression of CXC motif chemokine ligand 16 (CXCL16) has been demonstrated to be associated with tumor progression and metastasis, served as a prognostic factor in many cancers, with higher relative expression behaving as a marker of tumor progression. However, its role and mechanisms underlying progression and metastasis of gastric cancer (GC) are yet to be elucidated. In our investigation, public datasets and human GC tissue samples were used to determine the CXCL16 expression levels. Our results revealed that CXCL16 was upregulated in GC. The high expression CXCL16 in GC was significantly associated with histologic poor differentiation and pTNM staging. And high CXCL16 was positively correlated with the poor survival of GC patients. Gain-and loss-of-function experiments were employed to investigate the biological role of CXCL16 in proliferation and migration both in vitro and in vivo. Mechanically, Gene set enrichment analysis (GSEA) revealed that the epithelial­mesenchymal transition (EMT), Akt and MAPK signal pathway related genes were significantly enriched in the high CXCL16 group, which was confirmed by western blot. Moreover, overexpression CXCL16 promoted the disintegrin and metalloproteases (ADAM10) and the CXC motif chemokine receptor 6 (CXCR6) expression, which mediated the CXCL16/CXCR6 positive feedback loop in GC, with activating Akt and MAPK signaling pathways. Knocking down ADAM10 would interrupted the CXCL16/CXCR6 axis in the carcinogenesis and progression of GC. In conclusion, our findings offered insights into that CXCL16 promoted GC tumorigenesis by enhancing ADAM10-dependent CXCL16/CXCR6 axis activation.

CXCL16 may be a predisposing factor to atherosclerosis: An animal study.

Atherosclerosis (AS) is a chronic inflammatory process initiated when lipoprotein is retained in the arterial wall. Leukocyte recruitment accelerates this process. CXC chemokine ligand 16 (CXCL16) acts as a chemokine to attract immune cells and also facilitates the phagocytosis process of modified low­density lipoprotein. Whether CXCL16 promotes or inhibits the pathological process of AS remains to be elucidated. To clarify this, CXCL16 gene was introduced into C57BL/6J wild­type mice to establish a stable CXCL16 overexpression mouse model. The initial changes of AS in mice were induced by high­fat diet (HFD). To study how the interaction of HFD and CXCL16 affected fatty acid metabolism and deposition, body weight and plasma lipid profile were assessed. Soluble CXCL16, matrix metalloproteinase­9, monocyte chemoattractant protein­1 and intercellular adhesion molecule­1 were detected by immunohistochemistry and ELISA to identify how CXCL16 affects AS lesion formation. The present study suggested that overexpression of CXCL16 combined with HFD lead to atherogenesis by upregulating the aforementioned inflammatory related genes at a protein level. The present study was the first, to the best of the authors' knowledge, to build a CXCL16 homozygous transgenic mice model to study how overexpressed CXCL16 is associated with AS for intervening in the occurrence and development of AS.

Application effect of tirofiban on percutaneous coronary intervention in patients with acute coronary syndrome and its postoperative effect on C-X-C motif chemokine ligand 16 level and myocardial perfusion.

This study aimed to investigate the application effect of tirofiban on percutaneous coronary intervention (PCI) in patients with acute coronary syndrome (ACS) and its postoperative effect on C-X-C motif chemokine ligand 16 (CXCL16) level and myocardial perfusion. A total of 50 cases of patients diagnosed with acute coronary syndrome and treated in Sunshine Union Hospital (Weifang, China) were included in group A and 30 cases of healthy subjects underwent physical examination in our hospital during the same period were enrolled in group B. Tirofiban was used in group A patients during PCI. Clinical efficacy evaluation criteria were used to evaluate the efficacy after treatment. The level of CXCL16 in serum before and after treatment was detected by qRT-PCR. Receiver operating characteristic (ROC) curve was drawn to analyze the value of C-X-C Motif Chemokine Ligand in diagnosing ACS. Before treatment, CXCL16 level in group A was significantly higher than that in group B (p<0.001). After treatment, patients in TMPG grade 3 in group A were significantly increased (p<0.001). Tirofiban could improve myocardial perfusion in patients with ACS after PCI, reduce adverse events and CXCL16 levels. Serum CXCL16 is expected to be a potential diagnostic and therapeutic predictor of ACS.

CXCR6 deficiency impairs cancer vaccine efficacy and CD8+ resident memory T-cell recruitment in head and neck and lung tumors.

BACKGROUND: Resident memory T lymphocytes (TRM) are located in tissues and play an important role in immunosurveillance against tumors. The presence of TRM prior to treatment or their induction is associated to the response to anti-Programmed cell death protein 1 (PD-1)/Programmed death-ligand 1 (PD-L1) immunotherapy and the efficacy of cancer vaccines. Previous work by our group and others has shown that the intranasal route of vaccination allows more efficient induction of these cells in head and neck and lung mucosa, resulting in better tumor protection. The mechanisms of in vivo migration of these cells remains largely unknown, apart from the fact that they express the chemokine receptor CXCR6. METHODS: We used CXCR6-deficient mice and an intranasal tumor vaccination model targeting the Human Papillomavirus (HPV) E7 protein expressed by the TC-1 lung cancer epithelial cell line. The role of CXCR6 and its ligand, CXCL16, was analyzed using multiparametric cytometric techniques and Luminex assays.Human biopsies obtained from patients with lung cancer were also included in this study. RESULTS: We showed that CXCR6 was preferentially expressed by CD8+ TRM after vaccination in mice and also on intratumoral CD8+ TRM derived from human lung cancer. We also demonstrate that vaccination of Cxcr6-deficient mice induces a defect in the lung recruitment of antigen-specific CD8+ T cells, preferentially in the TRM subsets. In addition, we found that intranasal vaccination with a cancer vaccine is less effective in these Cxcr6-deficient mice compared with wild-type mice, and this loss of efficacy is associated with decreased recruitment of local antitumor CD8+ TRM. Interestingly, intranasal, but not intramuscular vaccination induced higher and more sustained concentrations of CXCL16, compared with other chemokines, in the bronchoalveolar lavage fluid and pulmonary parenchyma. CONCLUSIONS: This work demonstrates the in vivo role of CXCR6-CXCL16 axis in the migration of CD8+ resident memory T cells in lung mucosa after vaccination, resulting in the control of tumor growth. This work reinforces and explains why the intranasal route of vaccination is the most appropriate strategy for inducing these cells in the head and neck and pulmonary mucosa, which remains a major objective to overcome resistance to anti-PD-1/PD-L1, especially in cold tumors.