CD276 regulates the immune escape of esophageal squamous cell carcinoma through CXCL1-CXCR2 induced NETs.

BACKGROUND: CD276 (B7-H3), a pivotal immune checkpoint, facilitates tumorigenicity, invasiveness, and metastasis by escaping immune surveillance in a variety of tumors; however, the underlying mechanisms facilitating immune escape in esophageal squamous cell carcinoma (ESCC) remain enigmatic. METHODS: We investigated the expression of CD276 in ESCC tissues from patients by using immunohistochemistry (IHC) assays. In vivo, we established a 4-nitroquinoline 1-oxide (4NQO)-induced CD276 knockout (CD276wKO) and K14cre; CD276 conditional knockout (CD276cKO) mouse model of ESCC to study the functional role of CD276 in ESCC. Furthermore, we used the 4NQO-induced mouse model to evaluate the effects of anti-CXCL1 antibodies, anti-Ly6G antibodies, anti-NK1.1 antibodies, and GSK484 inhibitors on tumor growth. Moreover, IHC, flow cytometry, and immunofluorescence techniques were employed to measure immune cell proportions in ESCC. In addition, we conducted single-cell RNA sequencing analysis to examine the alterations in tumor microenvironment following CD276 depletion. RESULTS: In this study, we elucidate that CD276 is markedly upregulated in ESCC, correlating with poor prognosis. In vivo, our results indicate that depletion of CD276 inhibits tumorigenesis and progression of ESCC. Furthermore, conditional knockout of CD276 in epithelial cells engenders a significant downregulation of CXCL1, consequently reducing the formation of neutrophil extracellular trap networks (NETs) via the CXCL1-CXCR2 signaling axis, while simultaneously augmenting natural killer (NK) cells. In addition, overexpression of CD276 promotes tumorigenesis via increasing NETs' formation and reducing NK cells in vivo. CONCLUSIONS: This study successfully elucidates the functional role of CD276 in ESCC. Our comprehensive analysis uncovers the significant role of CD276 in modulating immune surveillance mechanisms in ESCC, thereby suggesting that targeting CD276 might serve as a potential therapeutic approach for ESCC treatment.

Gene expression associated with unfavorable vaginal bleeding in women using the etonogestrel subdermal contraceptive implant: a prospective study.

To evaluate gene expression associated with unfavorable vaginal bleeding in users of the Etonogestrel (ENG) contraceptive implant. Prospective study involving 100 women who intended to use the ENG implant. Exclusion criteria included abnormal uterine bleeding, inability to attend a 1-year follow-up, and implant removal for reasons unrelated to vaginal bleeding or loss of follow-up. We obtained endometrial biopsies before implant placement and assessed the expression of 20 selected genes. Users maintained a uterine bleeding diary for 12 months post-implant placement. For statistical analysis, we categorized women into those with or without favorable vaginal bleeding at 3 and 12 months. Women with lower CXCL1 expression had a 6.8-fold increased risk of unfavorable vaginal bleeding at 3 months (OR 6.8, 95% CI 2.21-20.79, p < 0.001), while those with higher BCL6 and BMP6 expression had 6- and 5.1-fold increased risks, respectively. By the 12-month follow-up, women with lower CXCL1 expression had a 5.37-fold increased risk of unfavorable vaginal bleeding (OR 5.37, 95% CI 1.63-17.73, p = 0.006). Women with CXCL1 expression < 0.0675, BCL6 > 0.65, and BMP6 > 3.4 had a higher likelihood of experiencing unfavorable vaginal bleeding at 3 months, and CXCL1 < 0.158 at 12 months. Users of ENG contraceptive implants with elevated BCL6 and BMP6 expression exhibited a higher risk of breakthrough bleeding at the 3-month follow-up. Conversely, reduced CXCL1 expression was associated with an elevated risk of bleeding at both the 3 and 12-month follow-ups.

The Clinical Significance and Involvement in Molecular Cancer Processes of Chemokine CXCL1 in Selected Tumors.

Chemokines play a key role in cancer processes, with CXCL1 being a well-studied example. Due to the lack of a complete summary of CXCL1's role in cancer in the literature, in this study, we examine the significance of CXCL1 in various cancers such as bladder, glioblastoma, hemangioendothelioma, leukemias, Kaposi's sarcoma, lung, osteosarcoma, renal, and skin cancers (malignant melanoma, basal cell carcinoma, and squamous cell carcinoma), along with thyroid cancer. We focus on understanding how CXCL1 is involved in the cancer processes of these specific types of tumors. We look at how CXCL1 affects cancer cells, including their proliferation, migration, EMT, and metastasis. We also explore how CXCL1 influences other cells connected to tumors, like promoting angiogenesis, recruiting neutrophils, and affecting immune cell functions. Additionally, we discuss the clinical aspects by exploring how CXCL1 levels relate to cancer staging, lymph node metastasis, patient outcomes, chemoresistance, and radioresistance.

Chemotherapy-elicited extracellular vesicle CXCL1 from dying cells promotes triple-negative breast cancer metastasis by activating TAM/PD-L1 signaling.

BACKGROUND: Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, and chemotherapy still serves as the cornerstone treatment functioning by inducing cytotoxic cell death. Notably, emerging evidence suggests that dying cell-released signals may induce cancer progression and metastasis by modulating the surrounding microenvironment. However, the underlying molecular mechanisms and targeting strategies are yet to be explored. METHODS: Apoptotic TNBC cells induced by paclitaxel or adriamycin treatment were sorted and their released extracellular vesicles (EV-dead) were isolated from the cell supernatants. Chemokine array analysis was conducted to identify the crucial molecules in EV-dead. Zebrafish and mouse xenograft models were used to investigate the effect of EV-dead on TNBC progression in vivo. RESULTS: It was demonstrated that EV-dead were phagocytized by macrophages and induced TNBC metastasis by promoting the infiltration of immunosuppressive PD-L1+ TAMs. Chemokine array identified CXCL1 as a crucial component in EV-dead to activate TAM/PD-L1 signaling. CXCL1 knockdown in EV-dead or macrophage depletion significantly inhibited EV-dead-induced TNBC growth and metastasis. Mechanistic investigations revealed that CXCL1EV-dead enhanced TAM/PD-L1 signaling by transcriptionally activating EED-mediated PD-L1 promoter activity. More importantly, TPCA-1 (2-[(aminocarbonyl) amino]-5-(4-fluorophenyl)-3-thiophenecarboxamide) was screened as a promising inhibitor targeting CXCL1 signals in EVs to enhance paclitaxel chemosensitivity and limit TNBC metastasis without noticeable toxicities. CONCLUSIONS: Our results highlight CXCL1EV-dead as a novel dying cell-released signal and provide TPCA-1 as a targeting candidate to improve TNBC prognosis.

Acupuncture alleviates inflammatory pain by activating CXCL1/CXCR2 signaling in the primary somatosensory cortex in adjuvant induced arthritis rats. / S1脑区CXCL1/CXCR2å‚ä¸Žé’ˆåˆºæ”¹å–„ä½å‰‚æ€§å ³èŠ‚ç‚Žå¤§é¼ ç‚Žæ€§ç—›çš„ä½œç”¨æœºåˆ¶.

OBJECTIVES: To observe whether acupuncture up-regulates chemokine CXC ligand 1 (CXCL1) in the brain to play an analgesic role through CXCL1/chemokine CXC receptor 2 (CXCR2) signaling in adjuvant induced arthritis (AIA) rats, so as to reveal its neuro-immunological mechanism underlying improvement of AIA. METHODS: BALB/c mice with relatively stable thermal pain reaction were subjected to planta injection of complete Freund adjuvant (CFA) for establishing AIA model, followed by dividing the AIA mice into simple AF750 (fluorochrome) and AF750+CXCL1 groups (n=2 in each group). AF750 labeled CXCL1 recombinant protein was then injected into the mouse's tail vein to induce elevation of CXCL1 level in blood for simulating the effect of acupuncture stimulation which has been demonstrated by our past study. In vivo small animal imaging technology was used to observe the AF750 and AF750+CXCL1-labelled target regions. After thermal pain screening, the Wistar rats with stable pain reaction were subjected to AIA modeling by injecting CFA into the rat's right planta, then were randomized into model and manual acupuncture groups (n=12 in each group). Other 12 rats that received planta injection of saline were used as the control group. Manual acupuncture (uniform reinforcing and reducing manipulations) was applied to bilateral "Zusanli" (ST36) for 4×2 min, with an interval of 5 min between every 2 min, once daily for 7 days. The thermal pain threshold was assessed by detecting the paw withdrawal latency (PWL) using a thermal pain detector. The contents of CXCL1 in the primary somatosensory cortex (S1), medial prefrontal cortex, nucleus accumbens, amygdala, periaqueductal gray and rostroventromedial medulla regions were assayed by using ELISA, and the expression levels of CXCL1, CXCR2 and mu-opioid receptor (MOR) mRNA in the S1 region were detected using real time-quantitative polymerase chain reaction. The immune-fluorescence positive cellular rate of CXCL1 and CXCR2 in S1 region was observed after immunofluorescence stain. The immunofluorescence double-stain of CXCR2 and astrocyte marker glial fibrillary acidic protein (GFAP) or neuron marker NeuN or MOR was used to determine whether there is a co-expression between them. RESULTS: In AIA mice, results of in vivo experiments showed no obvious enrichment signal of AF750 or AF750+CXCL1 in any organ of the body, while in vitro experiments showed that there was a stronger fluorescence signal of CXCL1 recombinant protein in the brain. In rats, compared with the control group, the PWL from day 0 to day 7 was significantly decreased (P<0.01) and the expression of CXCR2 mRNA in the S1 region significantly increased in the model group (P<0.05), while in comparison with the model group, the PWL from day 2 to day 7, CXCL1 content, CXCR2 mRNA expression and CXCR2 content, and MOR mRNA expression in the S1 region were significantly increased in the manual acupuncture group (P<0.05, P<0.01). Immunofluorescence stain showed that CXCR2 co-stained with NeuN and MOR in the S1 region, indicating that CXCR2 exists in neurons and MOR-positive neurons but not in GFAP positive astrocytes. CONCLUSIONS: Acupuncture can increase the content of CXCL1 in S1 region, up-regulate CXCR2 on neurons in the S1 region and improve MOR expression in S1 region of AIA rats, which may contribute to its effect in alleviating inflammatory pain.

CXCL1-CXCR2 axis mediates inflammatory response after sciatic nerve injury by regulating macrophage infiltration.

Macrophages play a crucial role in the inflammatory response following sciatic nerve injury. Studies have demonstrated that C-X-C motif chemokine (CXCL) 1 recruit macrophages by binding to C-X-C chemokine receptor (CXCR) 2 and participates in the inflammatory response of various diseases. Based on these findings, we aimed to explore the role of the CXCL1-CXCR2 axis in the repair process after peripheral nerve injury. Initially, we simulated sciatic nerve injury and observed an increased expression of CXCL1 and CXCR2 in the nerves of the injury group. Both in vivo and in vitro experiments confirmed that the heightened CXCL1 expression occurs in Schwann cells and is secreted, while the elevated CXCR2 is expressed by recruited macrophages. In addition, in vitro experiments demonstrated that the binding of CXCL1 to CXCR2 can activate the NLRP3 inflammasome and promote the production of interleukin-1 beta (IL-1ß) in macrophages. However, after mice were subjected to sciatic nerve injury, the number of macrophages and the expression of inflammatory factors in the sciatic nerve were reduced following treatment with the CXCR2 inhibitor SB225002. Simultaneously, we evaluated the sciatic nerve function index, the expression of p75 neurotrophic factor receptor (p75NTR), and myelin proteins, and all of these results were improved with the use of SB225002. Thus, our results suggest that after sciatic nerve injury, the CXCL1-CXCR2 axis mediates the inflammatory response by promoting the recruitment and activation of macrophages, which is detrimental to the repair of the injured nerves. In contrast, treatment with SB225002 promotes the repair of injured sciatic nerves.

H3K18 lactylation-mediated VCAM1 expression promotes gastric cancer progression and metastasis via AKT-mTOR-CXCL1 axis.

The role of the Immunoglobulin Superfamily (IgSF) as adhesion molecules in orchestrating inflammation is pivotal, yet its specific involvement in gastric cancer (GC) remains unknown. We analyzed IgSF components and discerned conspicuously elevated VCAM1 expression in GC, correlating with a poor prognosis. Remarkably, VCAM1 enhances GC cell proliferation and migration by activating AKT-mTOR signaling. Moreover, lactate in the tumor microenvironment (TME) promotes dynamic lactylation of H3K18 (H3K18la), leading to transcriptional activation of VCAM1 in GC cells. Furthermore, VCAM1 actively mediates intercellular communication in the TME. AKT-mTOR-mediated CXCL1 expression is increased by VCAM1, facilitating the recruitment of human GC-derived mesenchymal stem cells (hGC-MSCs), thereby fostering immunesuppression and accelerating cancer progression. In summary, H3K18 lactylation upregulated VCAM1 transcription, which activated AKT-mTOR signaling, and promoted tumor cell proliferation, EMT Transition and tumor metastasis. VCAM1 upregulated CXCL1 expression by AKT-mTOR pathway, so as to facilitate hGC-MSCs and M2 macrophage recruitment and infiltration. These findings provide novel therapeutic targets for GC.

STS â ¡A inhibited angiogenesis of lung adenocarcinoma by activating FOXO3 to inhibit CXCL1/STAT3/VEGF pathway.

BACKGROUND: Lung adenocarcinoma (LUAD) is the most popular type of lung cancer. Sulfotanshinone IIA sodium (STS IIA) has been proven to have an anticancer effect. However, its role in LUAD and its underlying mechanism remain unclear. OBJECTIVE: To investigate the role and mechanism of STS IIA in LUAD angiogenesis. METHODS: The mRNA levels of genes, including forkhead box O3 (FOXO3) and chemokine C-X-C motif ligand 1 (CXCL1), were detected by qRT-PCR. The levels of proteins, including FOXO3, CXCL1, and vascular endothelial growth factor (VEGF), were measured by Western blot. The proliferation and angiogenesis of human umbilical vein endothelial cells (HUVECs) were detected by the EdU assay and the tubule formation assay, respectively. The binding relationship between FOXO3 and CXCL1 was detected by dual-luciferase reporter assay. RESULTS: Our results illustrated that different concentrations of STS IIA inhibited the proliferation and angiogenesis of HUVECs. FOXO3 regulated the proliferation and angiogenesis of HUVECs inhibited by STS ⅡA via targeting CXCL1. Subsequently, we proved that exogenous CXCL1 alleviated the inhibition of proliferation and angiogenesis of HUVECs regulated by STS IIA via activating the STAT3/VEGF pathway. Finally, we found that STS IIA inhibited the angiogenesis of lung adenocarcinoma though FOXO3 to inhibit the CXCL1/STAT3/VEGF pathway. CONCLUSION: Our study finally elucidated the underlying molecular mechanism by which STS ⅡA inhibits LUAD angiogenesis.

Cxcl1 monomer-dimer equilibrium controls neutrophil extravasation.

The chemokine Cxcl1 plays a crucial role in recruiting neutrophils in response to infection. The early events in chemokine-mediated neutrophil extravasation involve a sequence of highly orchestrated steps including rolling, adhesion, arrest, and diapedesis. Cxcl1 function is determined by its properties of reversible monomer-dimer equilibrium and binding to Cxcr2 and glycosaminoglycans. Here, we characterized how these properties orchestrate extravasation using intravital microscopy of the cremaster. Compared to WT Cxcl1, which exists as both a monomer and a dimer, the trapped dimer caused faster rolling, less adhesion, and less extravasation. Whole-mount immunofluorescence of the cremaster and arrest assays confirmed these data. Moreover, the Cxcl1 dimer showed impaired LFA-1-mediated neutrophil arrest that could be attributed to impaired Cxcr2-mediated ERK signaling. We conclude that Cxcl1 monomer-dimer equilibrium and potent Cxcr2 activity of the monomer together coordinate the early events in neutrophil recruitment.

The Role and Mechanism of Spinal NF-κB-CXCL1/CXCR2 in Rats with Nucleus Pulposus-induced Radicular Pain.

STUDY DESIGN: Experimental study of the role and mechanism of spinal NFκB-CXCL1/CXCR2 in rats with nucleus pulposus-induced radicular pain. OBJECTIVE: This study investigated the role and mechanism of spinal NFκB-CXCL1/CXCR2 in autologous nucleus pulposus-induced pain behavior in rats and to clarify the involvement and regulation of spinal NFκB as an upstream molecule of CXCL1 in autologous nucleus pulposus-induced radicular pain in rats. SUMMARY OF BACKGROUND DATA: The inflammatory response of nerve roots is an important mechanism for the occurrence of chronic pain. NFκB-CXCL1/CXCR2 pathway plays an important role in the development of radicular pain, but its regulatory mechanism in the model of radicular pain induced by autologous nucleus pulposus is still unclear. MATERIALS AND METHODS: We established a rat model of autologous medullary nucleus transplantation. We observed and recorded the changes in 50% mechanical withdrawal threshold and thermal withdrawal latency before and after the administration of CXCL1-neutralizing antibodies, CXCR2 inhibitor, and NFκB inhibitor in each group of rats and evaluated the expression of NFκB, CXCL1, and CXCR2 in the spinal dorsal horn using immunofluorescence and Western blot. To compare differences between groups in behavioral testing, analysis of variance was employed. Dunnett's method was used to compare differences at different time points within a group and between different groups at the same time point. A comparison of the relative concentration of protein, relative concentration of mRNA, and semiquantitative data from immunofluorescence staining was conducted utilizing one-way ANOVA and Dunnett's pairwise comparison. RESULTS: Autologous nucleus pulposus transplantation can induce radicular pain in rats and upregulate the expression of CXCL1, CXCR2, and NFκB in the spinal cord. CXCL1 is co-expressed with astrocytes, CXCR2 with neurons, and NFκB with both astrocytes and neurons. The application of CXCL1 neutralizing antibodies, CXCR2 inhibitors, and NFκB inhibitors can alleviate pain hypersensitivity induced by autologous nucleus pulposus transplantation in rats. Inhibitors of NFκB could downregulate the expression of CXCL1 and CXCR2. CONCLUSIONS: We found that spinal NFκB is involved in NP-induced radicular pain in rats through the activation of CXCL1/CXCR2, enriching the mechanism of medullary-derived radicular pain and providing a possible new target and theoretical basis for the development of more effective anti-inflammatory and analgesic drugs for patients with chronic pain following LDH.

TGFß1 induces CXCL1 to promote stemness features in lung cancer.

Chemokines critically orchestrate the tumorigenesis, metastasis, and stemness features of cancer cells that lead to poor outcomes. High plasma levels of transforming growth factor-ß1 (TGFß1) correlate with poor prognostic features in advanced lung cancer patients, thus suggesting the importance of TGFß1 in the lung tumor microenvironment. However, the role of chemokines in TGFß1-induced tumor stemness features remains unclear. Here, we clarify the previously undocumented role of CXCL1 in TGFß1-induced lung cancer stemness features. CXCL1 and its receptor CXCR2 were significantly upregulated in TGFß1-induced lung cancer stem cells (CSCs). CXCL1 silencing (shCXCL1) suppressed stemness gene expression, tumorsphere formation, colony formation, drug resistance, and in vivo tumorigenicity in TGFß1-induced lung tumorspheres. Immunohistochemistry staining showed that patients with stage II/III lung cancer had higher expression levels of CXCL1. The levels of CXCL1 were positively associated with lymph node metastasis and correlated with the expression of the CSC transcription factor Oct-4. Furthermore, online database analysis revealed that CXCL1 expression was negatively correlated with lung cancer survival in patients. Patients with high TGFß1/CXCL1/CD44 co-expression had a worse survival rate. We suggest that CXCL1 serves as a crucial factor in TGFß1-induced stemness features of lung cancer.

Collagen 1-mediated CXCL1 secretion in tumor cells activates fibroblasts to promote radioresistance of esophageal cancer.

Esophageal squamous-cell carcinoma (ESCC) is commonly treated with radiotherapy; however, radioresistance hinders its clinical effectiveness, and the underlying mechanism remains elusive. Here, we develop patient-derived xenografts (PDXs) from 19 patients with ESCC to investigate the mechanisms driving radioresistance. Using RNA sequencing, cytokine arrays, and single-cell RNA sequencing, we reveal an enrichment of cancer-associated fibroblast (CAF)-derived collagen type 1 (Col1) and tumor-cell-derived CXCL1 in non-responsive PDXs. Col1 not only promotes radioresistance by augmenting DNA repair capacity but also induces CXCL1 secretion in tumor cells. Additionally, CXCL1 further activates CAFs via the CXCR2-STAT3 pathway, establishing a positive feedback loop. Directly interfering with tumor-cell-derived CXCL1 or inhibiting the CXCL1-CXCR2 pathway effectively restores the radiosensitivity of radioresistant xenografts in vivo. Collectively, our study provides a comprehensive understanding of the molecular mechanisms underlying radioresistance and identifies potential targets to improve the efficacy of radiotherapy for ESCC.

Biological function of C-X-C Motif Chemokine Ligand 1 gene (CXCL1) in ovarian malignant tumors.

OBJECTIVE: To determine the function of the chemokine (C-X-C motif) ligand 1 (CXCL1) gene in ovarian cancer cells and to investigate the relationship between CXCL1 gene mRNA expression and ovarian tumor clinical pathology. METHODS: Using bioinformatics methods to identify common differentially expressed genes associated with ovarian cancer in the GEO database. Growth curves of A2780 cells with or without CXCL1 expression were plotted by MTT assay. Cell cycles were measured by flow cytometry. Cell colony formation was enumerated in Transwell chambers. Migration and invasion in vitro were investigated using Cell Counting Kit-8 (CCK8), wound healing and Transwell, respectively. The relationship between CXCL1 gene mRNA expression and ovarian tumor clinical pathology was analyzed. RESULTS: CXCL1 was found to be one of the co-upregulated differentially expressed genes in the GEO database. The migration of A2780 cells expressing CXCL1 was significantly higher than that of A2780 cells without CXCL1 expression. CXCL1 mRNA expression in ovarian malignancy was significantly higher than those in benign lesions and the normal control (p < .01). In advanced ovarian cancer (Stages III-IV), CXCL1 mRNA expression was also significantly higher than that in patients with early-stage ovarian cancer (Stages I-II) (p = .005). Kaplan-Meier survival curve showed no correlation between CXCL1 mRNA expression and ovarian cancer prognosis. A Cox proportional hazard model also showed that CXCL1 expression was not an independent prognostic factor for ovarian cancer patients. CONCLUSIONS: CXCL1 gene could promotes ovarian cancer A2780 cell proliferation and invasion in vitro, and contributed theoretical knowledge for the target selection in molecular targeted therapy. CXCL1 mRNA over-expression may be correlated with the occurrence and development of ovarian malignancy. Level of plasma CXCL1 might serve as a biomarker for prognosis in ovarian carcinoma patients.

CXCL1 enhances COX-II expression in rheumatoid arthritis synovial fibroblasts by CXCR2, PLC, PKC, and NF-κB signal pathway.

Rheumatoid arthritis (RA) is the most common autoimmune disease, affecting the joints of the hands and feet. Several chemokines and their receptors are crucial in RA pathogenesis through immune cell recruitment. C-X-C Motif Chemokine Ligand 1 (CXCL1), a chemokine for the recruitment of various immune cells, can be upregulated in patients with RA. However, the discussion on the role of CXCL1 in RA pathogenesis is insufficient. Here, we found that CXCL1 promoted cyclooxygenase-2 (COX-II) expression in a dose- and time-dependent manner in rheumatoid arthritis synovial fibroblasts (RASFs). CXCL1 overexpression in RASFs led to a significant increase in COX-II expression, while the transfection of RASFs with the shRNA plasmid resulted in a noticeable decrease in COX-II expression. Next, we delineated the molecular mechanism underlying CXCL1-promoted COX-II expression and noted that CXC chemokine receptor 2 (CXCR2), phospholipase C (PLC), and protein kinase C (PKC) signal transduction were responsible for COX-II expression after CXCL1 incubation for RASFs. Finally, we confirmed the transcriptional activation of nuclear factor κB (NF-κB) in RASFs after incubation with CXCL1. In conclusion, the current study provided a novel insight into the role of CXCL1 in RA pathogenesis.

Chemokine Cxcl1-Cxcl2 heterodimer is a potent neutrophil chemoattractant.

Microbial infection is characterized by release of multiple proinflammatory chemokines that direct neutrophils to the insult site. How collective function of these chemokines orchestrates neutrophil recruitment is not known. Here, we characterized the role for heterodimer and show that the Cxcl1-Cxcl2 heterodimer is a potent neutrophil chemoattractant in mice and can recruit more neutrophils than the individual chemokines. Chemokine-mediated neutrophil recruitment is determined by Cxcr2 receptor signaling, Cxcr2 endocytosis, and binding to glycosaminoglycans. We have now determined heterodimer's Cxcr2 activity using cellular assays and Cxcr2 density in blood and recruited neutrophils in heterodimer-treated mice. We have shown that the heterodimer binds glycosaminoglycans with higher affinity and more efficiently than Cxcl1 or Cxcl2. These data collectively indicate that optimal glycosaminoglycan interactions and dampened receptor activity acting in concert in a dynamic fashion promote heterodimer-mediated robust neutrophil recruitment. We propose that this could play a critical role in combating infection.

TNF-α promotes CXCL-1/8 production in keratinocytes by downregulating galectin-3 through NF-κB and hsa-miR-27a-3p pathway to contribute psoriasis development.

OBJECTIVE: Treatment with TNF-α inhibitors improve psoriasis with minimize/minor neutrophils infiltration and CXCL-1/8 expression in psoriatic lesions. However, the fine mechanism of TNF-α initiating psoriatic inflammation by tuning keratinocytes is unclear. Our previous research identified the deficiency of intracellular galectin-3 was sufficient to promote psoriasis inflammation characterized by neutrophil accumulation. This study aims to investigate whether TNF-α participated in psoriasis development through dysregulating galectin-3 expression. METHODS: mRNA levels were assessed through quantitative real-time PCR. Flow cytometry was used to detect cell cycle/apoptosis. Western blot was used to evaluate the activation of the NF-κB signaling pathway. HE staining and immunochemistry were used to detect epidermal thickness and MPO expression, respectively. Specific small interfering RNA (siRNA) was used to knock down hsa-miR-27a-3p while plasmids transfection was used to overexpress galectin-3. Further, the multiMiR R package was utilized to predict microRNA-target interaction. RESULTS AND DISCUSSION: We found that TNF-α stimulation altered cell proliferation and differentiation and promoted the production of psoriasis-related inflammatory mediators along with the inhibition of galectin-3 expression in keratinocytes. Supplement of galectin-3 could counteract the rise of CXCL-1/8 but not the other phenotypes of keratinocytes induced by TNF-α. Mechanistically, inhibition of the NF-κB signaling pathway could counteract the decrease of galectin-3 and the increase of hsa-miR-27a-3p expression whereas silence of hsa-miR-27a-3p could counteract the decrease of galectin-3 expression induced by TNF-α treatment in keratinocytes. Intradermal injection of murine anti-CXCL-2 antibody greatly alleviated imiquimod-induced psoriasis-like dermatitis. CONCLUSION: TNF-α initiates psoriatic inflammation by increasing CXCL-1/8 in keratinocytes mediated by the axis of NF-κB-hsa-miR-27a-3p-galectin-3 pathway.

The role of CXCL1/CXCR2 axis in neurological diseases.

The C-X-C chemokine ligand (CXCL) 1 and its receptor C-X-C chemokine receptor (CXCR) 2 are widely expressed in the peripheral nervous systems (PNS) and central nervous systems (CNS) and are involved in the development of inflammation and pain after various nerve injuries. Once a nerve is damaged, it affects not only the neuron itself but also lesions elsewhere in its dominant site. After the CXCL1/CXCR2 axis is activated, multiple downstream pathways can be activated, such as c-Raf/MAPK/AP-1, p-PKC-µ/p-ILK/NLRP3, JAK2/STAT3, TAK1/NF-κB, etc. These pathways in turn mediate cellular motility state or cell migration. CXCR2 is expressed on the surface of neutrophils and monocytes/macrophages. These cells can be recruited to the lesion through the CXCL1/CXCR2 axis to participate in the inflammatory response. The expression of CXCR2 in neurons can activate some pathways in neurons through the CXCL1/CXCR2 axis, thereby causing damage to neurons. CXCR2 is also expressed in astrocytes, and when CXCR2 activated, it increases the number of astrocytes but impairs their function. Since inflammation can occur at almost any site of injury, elucidating the mechanism of CXCL1/CXCR2 axis' influence on inflammation may provide a favorable target for clinical treatment. Therefore, this article reviews the research progress of the CXCL1/CXCR2 axis in neurological diseases, aiming to provide a more meaningful theoretical basis for the treatment of neurological diseases.

Vascular cell-adhesion molecule 1 (VCAM-1) regulates JunB-mediated IL-8/CXCL1 expression and pathological neovascularization.

Vascular adhesion molecules play an important role in various immunological disorders, particularly in cancers. However, little is known regarding the role of these adhesion molecules in proliferative retinopathies. We observed that IL-33 regulates VCAM-1 expression in human retinal endothelial cells and that genetic deletion of IL-33 reduces hypoxia-induced VCAM-1 expression and retinal neovascularization in C57BL/6 mice. We found that VCAM-1 via JunB regulates IL-8 promoter activity and expression in human retinal endothelial cells. In addition, our study outlines the regulatory role of VCAM-1-JunB-IL-8 signaling on retinal endothelial cell sprouting and angiogenesis. Our RNA sequencing results show an induced expression of CXCL1 (a murine functional homolog of IL-8) in the hypoxic retina, and intravitreal injection of VCAM-1 siRNA not only decreases hypoxia-induced VCAM-1-JunB-CXCL1 signaling but also reduces OIR-induced sprouting and retinal neovascularization. These findings suggest that VCAM-1-JunB-IL-8 signaling plays a crucial role in retinal neovascularization, and its antagonism might provide an advanced treatment option for proliferative retinopathies.

CX3CL1 Pathway as a Molecular Target for Treatment Strategies in Alzheimer's Disease.

Alzheimer's disease (AD) is a scourge for patients, caregivers and healthcare professionals due to the progressive character of the disease and the lack of effective treatments. AD is considered a proteinopathy, which means that aetiological and clinical features of AD have been linked to the deposition of amyloid ß (Aß) and hyperphosphorylated tau protein aggregates throughout the brain, with Aß and hyperphosphorylated tau representing classical AD hallmarks. However, some other putative mechanisms underlying the pathogenesis of the disease have been proposed, including inflammation in the brain, microglia activation, impaired hippocampus neurogenesis and alterations in the production and release of neurotrophic factors. Among all, microglia activation and chronic inflammation in the brain gained some attention, with researchers worldwide wondering whether it is possible to prevent and stop, respectively, the onset and progression of the disease by modulating microglia phenotypes. The following key points have been established so far: (i) Aß deposition in brain parenchyma represents repeated stimulus determining chronic activation of microglia; (ii) chronic activation and priming of microglia make these cells lose neuroprotective functions and favour damage and loss of neurons; (iii) quiescent status of microglia at baseline prevents chronic activation and priming, meaning that the more microglia are quiescent, the less they become neurotoxic. Many molecules are known to modulate the quiescent baseline state of microglia, attracting huge interest among scientists as to whether these molecules could be used as valuable targets in AD treatment. The downside of the coin came early with the observation that quiescent microglia do not display phagocytic ability, being unable to clear Aß deposits since phagocytosis is crucial for Aß clearance efficacy. A possible solution for this issue could be found in the modulation of microglia status at baseline, which could help maintain both neuroprotective features and phagocytic ability at the same time. Among the molecules known to influence the baseline status of microglia, C-X3-chemokine Ligand 1 (CX3CL1), also known as Fractalkine (FKN), is one of the most investigated. FKN and its microglial receptor CX3CR1 are crucial players in the interplay between neurons and microglia, modulating the operation of some neural circuits and the efficacy and persistence of immune response against injury. In addition, CX3CL1 regulates synaptic pruning and plasticity in the developmental age and in adulthood, when it strongly impacts the hippocampus neurogenesis of the adult. CX3CL1 has an effect on Aß clearance and tau phosphorylation, as well as in microglia activation and priming. For all the above, CX3CL1/CX3CR1 signalling has been widely studied in relation to AD pathogenesis, and its biochemical pathway could hide molecular targets for novel treatment strategies in AD. This review summarizes the possible role of CX3CL1 in AD pathogenesis and its use as a potential target for AD treatment.

Daphnetin alleviates neuropathic pain in chronic constrictive injury rats via regulating the NF-κB dependent CXCL1/CXCR2 signaling pathway.

CONTEXT: Daphnetin is a natural product with anti-inflammatory, antioxidant, and neuroprotective properties. Reports have found that it has a strong analgesic effect; however, its analgesic mechanism is unknown. OBJECTIVE: We explored the effect and mechanism of daphnetin on neuropathic pain (NP). MATERIALS AND METHODS: The rat model of NP was established by ligation of the sciatic nerve. Male Sprague-Dawley rats were divided into six groups: Control, Model, Sham, morphine (0.375 mg/kg), and daphnetin (0.0625 and 0.025 mg/kg). Rats were intrathecally injected with drugs or normal saline once daily for three days. Hyperalgesia was evaluated by mechanical withdrawal threshold (MWT) and thermal withdrawal threshold (TWT). Protein levels were detected using ELISA, immunofluorescence, and western blotting. RESULTS: Compared to the Model group, daphnetin improved TWT (46.70 °C vs. 42.20 °C) and MWT (45.60 g vs. 23.60 g), reduced the expression of interleukin-1ß (0.99 ng/g vs. 1.42 ng/g), interleukin-6 (0.90 ng/g vs. 1.52 ng/g), and tumor necrosis factor-α (0.93 ng/g vs. 1.52 ng/g) in the sciatic nerve. Daphnetin decreased the expression of toll-like receptor 4 (TLR4) (0.47-fold), phosphorylated inhibitor of NF-κB (p-IKBα) (0.29-fold), nuclear factor kappaB (NF-κB) (0.48-fold), glial fibrillary acidic protein (GFAP) (0.42-fold), CXC chemokine ligand type 1 (CXCL1) (0.84-fold), CXC chemokine receptor type 2 (CXCR2) (0.78-fold) in the spinal cord. DISCUSSION AND CONCLUSIONS: Daphnetin alleviates NP by inhibiting inflammation and astrocyte activation in the spinal cord, providing theoretical support for the extensive clinical treatment of NP.