CXC Chemokine Receptor 4 Expression Research Articles

Atorvastatin Pretreatment Ameliorates Mesenchymal Stem Cell Migration through miR-146a/CXCR4 Signaling.

We previously found that atorvastatin (ATV) enhanced mesenchymal stem cells (MSCs) migration, by a yet unknown mechanism. CXC chemokine receptor 4 (CXCR4) is critical to cell migration and regulated by microRNA-146a (miR-146a). Therefore, this study aimed to assess whether ATV ameliorates MSCs migration through miR-146a/CXCR4 signaling. Expression of CXCR4 was evaluated by flow cytometry. Expression of miR-146a was examined by reverse transcription-quantitative polymerase chain reaction. A transwell system was used to assess the migration ability of MSCs. Recruitment of systematically delivered MSCs to the infarcted heart was evaluated in Sprague-Dawley rats with acute myocardial infarction (AMI). Mimics of miR-146a were used in vitro, and miR-146a overexpression lentivirus was used in vivo, to assess the role of miR-146a in the migration ability of MSCs. The results showed that ATV pretreatment in vitro upregulated CXCR4 and induced MSCs migration. In addition, flow cytometry demonstrated that miR-146a mimics suppressed CXCR4, and ATV pretreatment no longer ameliorated MSCs migration because of decreased CXCR4. In the AMI model, miR-146a-overexpressing MSCs increased infarct size and fibrosis. The miR-146a/CXCR4 signaling pathway contributes to MSCs migration and homing induced by ATV pretreatment. miR-146a may be a novel therapeutic target for stimulating MSCs migration to the ischemic tissue for improved repair.

CXCR4 as a prognostic marker in Egyptian chronic lymphocytic leukemia patients

Chronic lymphocytic leukemia (CLL) has variable clinical presentations, and molecular and biological prognostic markers. The C-X-C chemokine receptor 4 (CXCR4) and its ligand stromal cell-derived factor-1 (SDF-1) play an important role in trafficking of lymphocytes and monocytes. The aim was to study lymphocyte expression of CXCR4 and its prognostic value in CLL. A case control study was carried out on 30 newly diagnosed CLL cases and 30 healthy controls. Fludarabine, cyclophosphamide, and rituximab (FCR) was the standard treatment. Flowcytometric measurement of CXCR4 expression on lymphocytes was done. CXCR4 was significantly higher in patients than controls (81.67 ± 17.95 vs. 11.78 ± 2.78; P< 0.001). CXCR4 was significantly higher (P<0.001) in high risk CLL (93.63 ± 6.78) vs. intermediate risk (82.50 ± 7.13) and low risk (75.84 ± 12.23). CXCR4 was significantly higher (P<0.001) in non-responders (91.63 ± 6.98) vs. partial responders (83.11 ± 5.55) and complete responders (70.11 ± 4.44). CXCR4 was significantly lower in survivors vs. non-survivors (80.89 ± 5.09 vs. 85.43 ± 5.51; P< 0.001. CXCR4 had significant positive correlation with WBCs (r=0.45, P=0.01) and lymphocytes (r=0.40, P=0.01) measured at diagnosis. In conclusions, expression of CXCR4 in newly diagnosed CLL is significantly high. CXCR4 increased expression is associated with poor prognosis and resistance to the therapy.so it can be used as prognostic tool.

High CXCR4 expression in pancreatic ductal adenocarcinoma as characterized by an inflammatory tumor phenotype with potential implications for an immunotherapeutic approach.

4021 Background: Immunotherapy is considered ineffective in the majority of patients with pancreatic ductal adenocarcinoma (PDAC), a consequence of a highly immunosuppressive tumor microenvironment (TME). However, treatment induced inhibition of CXC chemokine receptor 4 (CXCR4) and programmed cell death protein-1 (PD-1) in the COMBAT trial caused T cell infiltration and tumor regression in a subset of PDAC patients. Elucidating a phenotype that predicts response is clinically relevant. We performed a comprehensive molecular landscape study in PDAC evaluating CXCR4 RNA expression. Methods: 3,647 PDAC specimens were centrally analysed. NextGen DNA sequencing (NextSeq, 592 gene panel or NovaSeq, whole-exome sequencing), whole-transcriptome RNA sequencing (NovaSeq) and immunohistochemistry (Caris Life Sciences, Phoenix, AZ) were performed. Gene expression is reported as TPM (Transcripts per million). Pathway gene enrichment analyses were done using GSEA (Subramaniam 2015, PNAS). Immune cell fraction was calculated by QuantiSeq (Finotello 2019, Genome Medicine). The cohort was stratified in quartiles according to CXCR4 RNA expression status. Results: Overall, CXCR4 expression was higher in primary tumors compared to distant metastasis (38 vs. 28 TPM, p &lt; 0.0001). CXCR4-high (top quartile: &gt; 59 TPMs), when compared to CXCR4-low (bottom quartile: &lt; 17 TPM) PDACs, were characterized by a high prevalence of mutations in signal transduction pathway genes (e.g. GNAS: 3.6 vs. 0.5%), an increased infiltration of immune cells (e.g. CD8+ T cells, M1 macrophages), and a higher expression of HLA-DRA and HLA-E (all p &lt; 0.0001). We detected an upregulation of CXCL9, CXCL10, CXCL12, CCL5, IDO1 and LAG3 in CXCR4-high compared to CXCR4-low tumors. In contrast, lower PD-L1 expression (17.4 vs. 13.1%, p = 0.02), genomic loss of heterozygosity (17.4 vs. 10.8%), and a lower frequency of gene amplifications in ERBB2 (2.1 vs. 0.1%), TNFRSF14 (2.0 vs. 0.1%), and TP53 (82 vs. 73%, all p &lt; 0.0001) were observed. Moreover, CXCR4-high expression was associated with a better survival (HR: 1.417, 95% CI [1.168-1.72], p &lt; 0.001). Conclusions: This is the first study comprehensively investigating the molecular landscape of PDACs according to CXCR4 RNA expression. High CXCR4 expression is associated with an improved survival and a pro-inflammatory phenotype that may identify a subset of tumors with greater responsiveness to immunotherapeutic approaches.

Macrophages contribute to liver repair after monocrotaline-induced liver injury via SDF-1/CXCR4.

Monocrotaline (MCT) administration induces liver injury in rodents that mimics the pathology of human sinusoidal obstruction syndrome. MCT-induced SOS models are used to investigate the mechanism of injury and optimize treatment strategies. However, the processes underlying liver repair are largely unknown. Specifically, the role of macrophages, the key drivers of liver repair, has not been elucidated. The current study aimed to examine the role of macrophages in the repair of MCT-induced liver injury in male C57/BL6 mice. Maximal liver injury occurred at 48 h post-MCT treatment, followed by repair at 120 h post-treatment. Immunofluorescence analysis revealed that CD68+ macrophages were recruited to the injured regions after MCT treatment. This was associated with the decreased expression of genes related to a pro-inflammatory macrophage phenotype and the increased expression of those associated with a reparative macrophage phenotype during the repair phase. The results also revealed that stromal cell-derived factor-1 (SDF-1) and its receptor C-X-C chemokine receptor-4 (CXCR4) were upregulated, and CD68+ macrophages were co-localized with CXCR4 expression. Treatment of mice with AMD3100, a CXCR4 antagonist, delayed liver repair and increased the expression of genes related to a pro-inflammatory macrophage phenotype. In contrast, SDF-1 treatment stimulated liver repair and increased the expression of genes related to a reparative macrophage phenotype. The results suggested that macrophages accumulate in the liver and repair damaged tissue after MCT treatment, and that the SDF-1-CXCR4 axis is involved in this process.

The CXCL12/CXCR7 signalling axis promotes proliferation and metastasis in cervical cancer

C-X-C chemokine receptor 7 (CXCR7), a novel receptor of C-X-C motif chemokine ligand 12 (CXCL12), is associated with the occurrence and metastasis of various malignant tumours. However, the role, function and underlying mechanisms of CXCR7 expression in cervical cancer remain undefined. The expression level of CXCR7 was evaluated in cervical cancer samples by immunohistochemistry and real-time PCR analyses. Western blot analysis was used to examine the expression level of CXCR7 in cervical cancer cell lines. HeLa cells were genetically silenced or pharmacologically inhibited for CXCR7 or CXCR4. Transwell and CCK-8 assays were used to examine cell migration and proliferation. The expression levels of MMP2, MMP9, TIMP-1 and TIMP-2 in HeLa cells were assessed by western blot or real-time PCR. HeLa cells silenced for CXCR7 were subcutaneously injected into nude mice to form tumours. The expression of CXCR7 in nude mice was investigated by immunohistochemical staining. Tumour volumes and weights were measured. The in vivo expression levels of MMP2, MMP9, TIMP-1 and TIMP-2 were determined by western blot analysis and real-time PCR. CXCR7 was overexpressed in cervical cancer tissues and cell lines. CXCL12 was highly expressed in cervical cancer lines. CXCR7 silencing or CCX733 treatment rather than CXCR4 silencing or AMD3100 treatment suppressed the proliferation, migration and invasion of cervical cancer cells stimulated by CXCL12. In a xenograft tumour model, CXCR7 silencing or CCX733 treatment inhibited the volumes and weights of xenograft tumours. In addition, downregulation of CXCR7 decreased the expression levels of MMP2 and MMP9 but increased the expression levels of TIMP-1 and TIMP-2 in vivo. These data support the finding that the downregulation of CXCR7 suppresses the proliferation and metastasis of cervical cancer cells. Inhibition of CXCR7 may be a potential targeted therapy for cervical cancer.

Leptin Receptor q223r Polymorphism Influences Clostridioides difficile Infection-Induced Neutrophil CXCR2 Expression in an Interleukin-1β Dependent Manner.

Neutrophils are key first-responders in the innate immune response to C. difficile infection (CDI) and play a central role in disease pathogenesis. Studies have clearly shown that tissue neutrophil numbers need to be tightly regulated for optimal CDI outcomes: while excessive colonic neutrophilia is associated with severe CDI, neutrophil depletion also results in worse outcomes. However, the biological mechanisms that control CDI-induced neutrophilia remain poorly defined. C-X-C chemokine receptor 2 (CXCR2) is a chemotactic receptor that is critical in neutrophil mobilization from bone marrow to blood and tissue sites. We have previously reported that a single nucleotide polymorphism (SNP) in leptin receptor (LEPR), present in up to 50% of people, influenced CDI-induced neutrophil CXCR2 expression and tissue neutrophilia. Homozygosity for mutant LEPR (i.e. RR genotype) was associated with higher CXCR2 expression and more tissue neutrophils. Here, we investigated the biological mechanisms that regulate neutrophil CXCR2 expression after CDI, and the influence of host genetics on this process. Our data reveal that: a) CXCR2 plays a key role in CDI-induced neutrophil extravasation from blood to colonic tissue; b) plasma from C. difficile-infected mice upregulated CXCR2 on bone marrow neutrophils; c) plasma from C. difficile-infected RR mice induced a higher magnitude of CXCR2 upregulation and had more IL-1β; and d) IL-1β neutralization reduced CXCR2 expression on bone marrow and blood neutrophils and their subsequent accrual to colonic tissue. In sum, our data indicate that IL-1β is a key molecular mediator that communicates between gastro-intestinal tract (i.e. site of CDI) and bone marrow (i.e. primary neutrophil reservoir) and regulates the intensity of CDI-induced tissue neutrophilia by modulating CXCR2 expression. Further, our studies highlight the importance of host genetics in affecting these innate immune responses and provide novel insights into the mechanisms by which a common SNP influences CDI-induced neutrophilia.

Oxygen Desaturation Is Associated With Fibrocyte Activation via Epidermal Growth Factor Receptor/ Hypoxia-Inducible Factor-1α Axis in Chronic Obstructive Pulmonary Disease

Background: Fibrocytes are bloodborne mesenchymal progenitors which accumulate and differentiate at disease site. We investigated whether hypoxemia activates fibrocytes, accelerating airflow limitation and exercise intolerance in chronic obstructive pulmonary disease (COPD) patients. Methods: Flow cytometry was used to determine collagen I+/CD45+ fibrocytes and α-smooth muscle actin+ differentiating fibrocytes within peripheral blood and cultured cells, as well as the expression of CXC chemokine receptor 4 (CXCR4), epidermal growth factor receptor (EGFR), connective tissue growth factor (CTGF) and hypoxia-inducible factor (HIF)-1α. Fibrocytes in lung specimens were identified by confocal microscopy. Findings: Compared to non-desaturators, COPD desaturators (peripheral blood oxygen saturation ≤88% during exercise) had greater number of fibrocytes in peripheral blood and lung specimens, paralleled with faster yearly lung function decline and 6-minute walk distance. Fibrocytes from desaturators expressed more EGFR, CXCR4, CTGF and HIF-1α, with a higher capacity to proliferation and myofibroblastic differentiation. Hypoxia (5% oxygen) increased the expression of EGFR, CXCR4, CTGF and HIF-1α, the number and differention in fibrocytes. These effects were attenuated by EGFR inhibitor gefitinib, HIF-1α gene silencing, and anti-CTGF antibody. Interpretation: These data elucidate that hypoxaemia triggers fibrocyte activation through the EGFR/HIF-1α axis, aggravating airflow obstruction in COPD. Funding Information: This work was supported by the National Medical Research Program (NMRP) grant 99-2314-B-182-043-MY3, 108-2314-B-182A-128-MY2 and the Chang Gung Medical Research Project (CMRP) grant CMRPG390041-2. Declaration of Interests: The authors have declared that no competing interest exists. Ethics Approval Statement: The study was approved by the Ethics Committee of Chang Gung Memorial Hospital (IRB: 98-3950B, 201801979A3). Written informed consents were obtained from all participants.

CXCR4 Promotes Growth and Metastasis of Esophageal Squamous Cell Carcinoma as a Critical Downstream Mediator of HIF-1α

Background: Recent studies have shown that C-X-C chemokine receptor 4(CXCR4) plays an important role in a variety of tumours. However, its role and regulated mechanism in esophageal squamous cell carcinoma (ESCC) remain unclear. Methods: Immunohistochemistry, Western blot and qPCR were used to detect the expression of CXCR4 and HIF-1α. HIF-1α downstream target gene was detected by RNA-seq. The invasion and metastasis ability of HIF-1α/CXCR4 axis to ESCC cells were detected by transwell and animal experiments. Dual-luciferase analysis and chromatin immunoprecipitation assay showed that HIF-1α directly binds to the HRE site in the CXCR4 promoter. Findings: We showed that CXCR4 was enriched in ESCC tissues compared with corresponding normal tissues and predicted lymph node metastasis and poor prognosis. CXCR4 overexpression promotes the proliferation, migration and invasion of ESCC. In addition, we revealed that hypoxia-inducible factor-1α (HIF-1α) regulated CXCR4 expression via binding to a hypoxia response element in its promoter, and the HIF-1α-enhanced metastasis ability of ESCC cells was reversed by CXCR4 knockdown or treatment with MSX-122, a CXCR4 antagonist. Interpretation: In showing that the dysregulated HIF-1α/CXCR4 axis promotes ESCC malignancy, the results suggested that CXCR4 is a potential therapeutic target for ESCC. Funding: This research was supported by the National Natural Science Foundation of China (81772619, 82002551), the Science and Technology Project of Tianjin Municipal Health Commission (RC20119), the Bethune Charitable Foundation-Excelsior Surgical Fund (HZB-20190528-18), and the Clinical Trial Project of Tianjin Medical University (2017kylc006). Declaration of Interest: The authors declare no conflict of interest. Ethical Approval: All operations for human subjects were approved by the ethics committee of our hospital, and all patients provided informed consent. Gene Expression Omnibus (GEO) data were downloaded from the website (https://www.ncbi.nlm.nih.gov/geo/).

High-Contrast CXCR4-Targeted 18F-PET Imaging Using a Potent and Selective Antagonist.

C-X-C chemokine receptor 4 (CXCR4) is highly expressed in cancers, contributing to proliferation, metastasis, and a poor prognosis. The noninvasive imaging of CXCR4 can enable the detection and characterization of aggressive cancers with poor outcomes. Currently, no 18F-labeled CXCR4 positron emission tomography (PET) radiotracer has demonstrated imaging contrast comparable to [68Ga]Ga-Pentixafor, a CXCR4-targeting radioligand. We, therefore, aimed to develop a high-contrast CXCR4-targeting radiotracer by incorporating a hydrophilic linker and trifluoroborate radioprosthesis to LY2510924, a known CXCR4 antagonist. A carboxy-ammoniomethyl-trifluoroborate (PepBF3) moiety was conjugated to the LY2510924-derived peptide possessing a triglutamate linker via amide bond formation to obtain BL08, whereas an alkyne ammoniomethyl-trifluoroborate (AMBF3) moiety was conjugated using the copper-catalyzed [3+2] cycloaddition click reaction to obtain BL09. BL08 and BL09 were radiolabeled with [18F]fluoride ion using 18F-19F isotope exchange. Pentixafor was radiolabeled with [68Ga]GaCl3. Side-by-side PET imaging and biodistribution studies were performed on immunocompromised mice bearing Daudi Burkitt lymphoma xenografts. The biodistribution of [18F]BL08 and [18F]BL09 showed tumor uptake at 2 h postinjection (p.i.) (5.67 ± 1.25%ID/g and 5.83 ± 0.92%ID/g, respectively), which were concordant with the results of PET imaging. [18F]BL08 had low background activity, providing tumor-to-blood, -muscle, and -liver ratios of 72 ± 20, 339 ± 81, and 14 ± 3 (2 h p.i.), respectively. [18F]BL09 behaved similarly, with ratios of 64 ± 20, 239 ± 72, and 17 ± 3 (2 h p.i.), respectively. This resulted in high-contrast visualization of tumors on PET imaging for both radiotracers. [18F]BL08 exhibited lower kidney uptake (2.2 ± 0.5%ID/g) compared to [18F]BL09 (7.6 ± 1.0%ID/g) at 2 h p.i. [18F]BL08 and [18F]BL09 demonstrated higher tumor-to-blood, -muscle, and -liver ratios compared to [68Ga]Ga-Pentixafor (18.9 ± 2.7, 95.4 ± 36.7, and 5.9 ± 0.7 at 2 h p.i., respectively). In conclusion, [18F]BL08 and [18F]BL09 enable high-contrast visualization of CXCR4 expression in Daudi xenografts. Based on high tumor-to-organ ratios, [18F]BL08 may prove a valuable new tool for CXCR4-targeted PET imaging with potential for translation. The use of a PepBF3 moiety is a new approach for the orthogonal conjugation of organotrifluoroborates for 18F-labeling of peptides.

Characterization of the Prognostic Values of the CXCR1-7 in Clear Cell Renal Cell Carcinoma (ccRCC) Microenvironment.

Background: As cancer immunotherapy has become a hot research topic, the values of CXC chemokine receptors (CXCRs) in tumor microenvironment have been increasingly realized. More and more evidence showed that the aberrant expression of CXCRs is closely related to the prognosis of various cancers. However, prognostic values and the exact roles of different CXCRs in clear cell renal cell carcinoma (ccRCC) have not yet been elucidated.Methods: To further evaluate the potential of seven CXCRs as prognostic biomarkers for ccRCC, multiple online analysis tools, including ONCOMINE, UALCAN (TCGA dataset), Kaplan–Meier Plotter, MethSurv, cBioPortal, GEPIA, Metascape, and TIMER databases, were utilized in our research.Results: The mRNA expression of CXCR4/6/7 was significantly increased in ccRCC patients, and all CXCRs are remarkably related to tumor stage or grade of ccRCC. Higher levels of CXCR3/4/5/6 expression were correlated with worse overall survival (OS) in patients with ccRCC, while higher expression of CXCR2 was associated with better OS. 23.14% mutation rate (118/510) of CXCR1-7 was observed in ccRCC patients, and the genetic alterations in CXCRs were related to worse OS and progression-free survival in ccRCC patients. Additionally, 53 CpGs of CXCR1-7 showed significant prognostic values. For functional enrichment, our results showed that CXCRs and their similar genes may be involved in cancer-associated pathways, immune process, and angiogenesis, etc. Besides, CXCRs were significantly correlated with multiple immune cells (e.g., CD8+ T cell, CD4+ cell, and dendritic cell).Conclusion: This study explored the potential prognostic values and roles of the CXCRs in ccRCC microenvironment. Our results suggested that CXCR4 and CXCR6 could be the prognostic biomarkers for the patients with ccRCC.

Significance of CXC chemokine receptor 4 (CXCR4) in Renal Cell Carcinoma

Background: Cancer stem cells (CSCs) represent a population with tumour initiating, self-renewal, and differentiation potential. The aim of the work is to evaluate the expression patterns and clinical significance of chemokine receptor type 4 (CXCR4) as a novel CSC marker in renal cell carcinoma (RCC). Material and Methods: the expression of CXCR4 was examined in 50 well-defined renal tumour tissues, including 28 clear cell renal cell carcinomas (ccRCCs), 11 papillary renal cell carcinomas (pRCCs), and 11 chromophobe renal cell carcinomas (ChRCCs) with immunohistochemistry staining, the association between expression of this marker and clinicopathologic parameters, was then analyzed. Results: CXCR4 expression & intensity was significantly correlated with RCC clinicopathological features as grade, stage, size of the tumor and microvascular invasion (MVI) but there was insignificant correlation between histopathological types and the pattern of CXCR4 expression and its intensity . Conclusion: increased CXCR4 expression was associated with more aggressive tumour behaviour in RCC patients, especially in pRCC and ccRCC subtypes due to their more metastatic behaviour. These findings suggest that CXCR4 can be considered as a novel prognostic and therapeutic marker for targeted therapy of renal carcinoma.

CXCR4 and RIF1 overexpression induces resistance of epithelial ovarian cancer to cisplatin-based chemotherapy.

The chemoresistance of epithelial ovarian cancer (EOC) is a major problem. Thus, the search for novel biomarkers associated with cisplatin sensitivity is overwhelming. Previous studies have shown that CXC chemokine receptor 4 (CXCR4) is associated with tumor growth, angiogenesis, and distant metastases, and replication timing regulatory factor 1 (RIF1) is responsible for the repair of double-strand DNA breaks. This study, thus, aimed to identify the correlation between CXCR4 and RIF1 overexpression and cisplatin sensitivity in EOC. Fifty-five EOC patients were recruited to assess the chemosensitivity of EOC to cisplatin-based chemotherapy at the Oncology Department in Tanta University Hospitals, Egypt. The results showed that patients with a higher CXCR4 and RIF1 expression exhibited a significantly lower chemosensitivity, worse overall survival, and poorer progression-free survival. The only prognostic associated with overall survival was CXCR4. Our study showed that CXCR4 and RIF1expression levels are not associated only with poor prognostic features of epithelial ovarian cancer but also with its chemoresistance to cisplatin, and consequently, with worse overall survival and progression free survival.

Positive Cross-Talk Between CXC Chemokine Receptor 4 (CXCR4) and Epidermal Growth Factor Receptor (EGFR) Promotes Gastric Cancer Metastasis via the Nuclear Factor kappa B&#x0D; (NF-kB)-Dependent Pathway

BackgroundPrevious studies have established cross-talk between CXC chemokine receptor 4 (CXCR4) and epidermal growth factor receptor (EGFR) in gastric cancer, however, the effect of dual CXCR4/EGFR tumor status on patient survival and mechanisms regulating expression has yet to be investigated.Material/MethodsA total of 56 gastric cancer patients were recruited to reveal the relationship between CXCR4 and EGFR expression, and the clinic-pathological features of samples were investigated by immunohistochemical staining. Two gastric cancer cell lines were treated with CXCL12 or EGF, and expression levels of CXCR4 and EGFR were detected by reverse-transcription-polymerase chain reaction and western blotting. Cells were treated with an NF-κB pathway inhibitor to investigate its role in the regulation of CXCL12 or EGF-mediated CXCR4 and EGFR expression and migration ability.ResultsThe results show that CXCL12 upregulated CXCR4 and EGFR. Similarly, EGF could induce the expression of CXCR4 and contribute to gastric cancer cell metastasis. In addition, both CXCL12 and EGF could induce the activation of IKKα/β and P65. Conversely, suppression of the NF-κB pathway remarkably decreased the expression of CXCR4/EGFR and migration ability induced by EGF or CXCL12. Furthermore, a significantly positive correlation between CXCR4 and EGFR expression was observed in gastric cancer patient tissues (r=0.372, P=0.005). Samples expressing both receptors had significantly poorer patient prognosis than other patient groups (P=0.002).ConclusionsOur results showed that the CXCL12/CXCR4 and EGF/EGFR axis can regulate the expression of each other through the NF-κB pathway to promote metastasis. These data suggested that simultaneous inhibition of EGFR and CXCR4 may be a potential therapeutic strategy in gastric cancer.

Plerixafor stimulates adhesive activity and endothelial regeneration of endothelial progenitor cells via elevating CXCR7 expression

AimsTo assess the effects of plerixafor on function and endothelial regeneration of endothelial progenitor cells (EPCs). MethodsThe proliferation and adhesion capacity of EPCs were evaluated in vitro. Furthermore, the expression levels of CXC chemokine receptor-7 (CXCR7) were detected before and after treatment with plerixafor. The CXCR7 expression of EPCs was knocked-down by RNA interference to evaluate the role of CXCR7 in regulating function of EPCs. A rat carotid artery injury model was established to assess the influences of plerixafor on endothelial regeneration. ResultsPlerixafor stimulated adhesion capacity of EPCs, associating with upregulation of CXCR7 and activation of LFA-1 and VLA-4 molecules. Knockdown of CXCR7 slightly impaired proliferation capacity but significantly attenuated adhesion capacity of EPCs. Plerixafor facilitated endothelial repair at 7 days, while reduced neointimal hyperplasia at 7 and 14 days via recruiting more EPCs participating in endothelial reparation. ConclusionsPlerixafor can positively regulate adhesion capacity of EPCs to HUVECs via elevating the expression level of CXCR7 and stimulating LFA-1 and VLA-4 molecules activation. Treatment with plerixafor accelerated re-endothelialization and inhibited neointimal hyperplasia after endoth elial injury, indicating that it can to be used for endothelial regeneration.

MiR-181a reduces radiosensitivity of non-small-cell lung cancer via inhibiting PTEN.

The aim of this study is to explore the effect of micro ribonucleic acid (miR)-181a on the radiosensitivity of non-small cell lung cancer (NSCLC) and its potential mechanism of action. The differentially expressed miRNAs were screened in lung cancer tissues of radiotherapy-resistant and non-radiotherapy-resistant NSCLC patients, and verified via reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Next, the effects of different miRNA expressions on patients' survival time were discussed, and target genes of miR-181a were predicted. The effect of miR-181a expression on radiosensitivity was determined using cell counting kit-8 (CCK-8) assay and flow cytometry. The direct target of miR-181a was verified via luciferase reporter assay. Phosphatase and tensin homolog deleted on chromosome ten (PTEN) was overexpressed using lentiviruses, and then whether miR-181a reduces radiosensitivity via targeting PTEN was detected via CCK-8 assay and flow cytometry. Finally, Western blotting was performed to detect the protein expression of PTEN. The screening results of microarray expression profile assay revealed that 15 miRNAs had significant differences in lung cancer tissues of radiotherapy-resistant NSCLC patients compared with those in non-radiotherapy-resistant NSCLC patients. The results of RT-qPCR showed that hsa-miR-181a, hsa-miR-199b, hsa-miR-489 and hsa-miR-589 were significantly up-regulated in the lung cancer tissues of radiotherapy-resistant NSCLC patients compared with those in non-radiotherapy-resistant NSCLC patients. In addition, it was found that the survival time of NSCLC patients was obviously prolonged in hsa-miR-181a low-expression group and hsa-miR-589 high-expression group, but hsa-miR-489 and hsa-miR-199b had no significant influence on the survival time of NSCLC patients. According to KEGG enrichment analysis, the target genes of miR-181a were evidently enriched in the phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway, NSCLC signaling pathway and other cancer signaling pathways. Under the radiation dose of 2, 4, 6 and 8 Gy, the survival rate of A549 cells rose in miR-181a mimic group, but declined in miR-181a inhibitor group. Moreover, compared with that in model group, the radiotherapy-induced apoptosis was markedly inhibited in miR-181a mimic group, but markedly promoted in miR-181a inhibitor group. It was also observed that the response of cells to radiotherapy-induced apoptosis was remarkably weakened in miR-181a mimic + PTEN overexpression group compared with that in miR-181a mimic group. Finally, miR-181a mimic group had a significantly lower protein expression of PTEN and significantly higher protein expressions of CXC chemokine receptor 4 (CXCR4), phosphorylated signal transducer and activator of transcription 3 (p-STAT3), p-AKT1 and p-mammalian target of rapamycin (mTOR) than model group, while miR-181a inhibitor group had the opposite protein expressions. The protein expressions of CXCR4, p-STAT3, p-AKT1 and p-mTOR were obviously lower in miR-181a mimic + PTEN overexpression group than those in miR-181a mimic group. MiR-181a reduces the radiosensitivity of NSCLC via inhibiting PTEN expression.

CXCR4-Targeted PET Imaging of Central Nervous System B-Cell Lymphoma.

C-X-C chemokine receptor 4 (CXCR4) is a transmembrane chemokine receptor involved in growth, survival, and dissemination of cancer, including aggressive B-cell lymphoma. MRI is the standard imaging technology for central nervous system (CNS) involvement of B-cell lymphoma and provides high sensitivity but moderate specificity. Therefore, novel molecular and functional imaging strategies are urgently required. Methods: In this proof-of-concept study, 11 patients with lymphoma of the CNS (8 primary and 3 secondary involvement) were imaged with the CXCR4-directed PET tracer 68Ga-pentixafor. To evaluate the predictive value of this imaging modality, treatment response, as determined by MRI, was correlated with quantification of CXCR4 expression by 68Ga-pentixafor PET in vivo before initiation of treatment in 7 of 11 patients. Results:68Ga-pentixafor PET showed excellent contrast with the surrounding brain parenchyma in all patients with active disease. Furthermore, initial CXCR4 uptake determined by PET correlated with subsequent treatment response as assessed by MRI. Conclusion:68Ga-pentixafor PET represents a novel diagnostic tool for CNS lymphoma with potential implications for theranostic approaches as well as response and risk assessment.

Stromal cell-derived factor-1 (SDF-1) expression in very preterm human lungs: potential relevance for stem cell therapy for bronchopulmonary dysplasia

Background: The axis formed by CXC chemokine receptor 4 (CXCR4), expressed on mesenchymal stromal cells (MSCs), and stromal cell-derived factor-1 (SDF-1), expressed in recipient organs, is a critical mediator of MSC migration in non-pulmonary injury models. The role and regulation of SDF-1 expression in preterm lungs, of potential relevance for MSC-based cell therapy for bronchopulmonary dysplasia (BPD), is unknown. The aim of this study was to determine the spatiotemporal pattern of CXCR4/SDF-1 expression in lungs of extremely preterm infants at risk for BPD.Methods: Postmortem lung samples were collected from ventilated extremely preterm infants who died between 23 and 29 wks (“short-term ventilated”) or between 36 and 39 wks (“long-term ventilated”) corrected postmenstrual age. Results were compared with age-matched infants who had lived <12 h or stillborn infants (“early” and “late” controls). CXCR4 and SDF-1 expression was studied by immunohistochemistry, immunofluorescence/confocal microscopy, and qRT-PCR analysis.Results: Compared with age-matched controls without antenatal infection, lungs of early control infants with evidence of intrauterine infection/inflammation showed significant upregulation of SDF-1 expression, localized to the respiratory epithelium, and of CXCR4 expression, localized to stromal cells. Similarly, pulmonary SDF-1 mRNA levels were significantly higher in long-term ventilated ex-premature infants with established BPD than in age-matched controls. The pulmonary vasculature was devoid of SDF-1 expression at all time points. Endogenous CXCR4-positive stromal cells were preferentially localized along the basal aspect of SDF-1-positive bronchial and respiratory epithelial cells, suggestive of functionality of the CXCR4/SDF-1 axis.Conclusions: Incipient and established neonatal lung injury is associated with upregulation of SDF-1 expression, restricted to the respiratory epithelium. Knowledge of the clinical associations, time-course and localization of pulmonary SDF-1 expression may guide decisions about the optimal timing and delivery route of MSC-based cell therapy for BPD.

CXCL16 silencing alleviates hepatic ischemia reperfusion injury during liver transplantation by inhibiting p38 phosphorylation

CXC chemokine ligand 16 (CXCL16) has been reported to exacerbate acute kidney injury induced by ischemia-reperfusion (IR). This study aimed to investigate the probable role of CXCL16 in hepatic IR injury during liver transplantation. The expression patterns of CXCL16 and its receptor CXC chemokine receptor 6 (CXCR6) were detected in mouse models of IR injury during liver transplantation and cell models of oxygen-glucose deprivation and reoxygenation (OGD/R)-induced hepatocyte injury using RT-qPCR, Western blot analysis and ELISA. CXCL16 expression was silenced in AML12 cells exposed to OGD/R conditions to determine the role of CXCL16 in cell apoptosis and injury. After treatment with CXCL16 and a p38 phosphorylation inhibitor, SB203580, we examined whether CXCL16 regulated p38 phosphorylation to impact hepatocyte injury. To verify the effects of CXCL16 and p38 phosphorylation in vivo, CXCL16 was silenced and p38 phosphorylation was activated in IR-treated mice. CXCL16 and CXCR6 were highly expressed in mouse models of IR injury and cell models of OGD/R-induced hepatocyte injury. Silencing of CXCL16 in AML12 cells resulted in diminished CXCR6 expression and alleviated OGD/R-exposed cell injury. CXCL16 treatment in AML12 cells brought about increased protein expressions of CXCR6 and p38 phosphorylation and elevated apoptosis rate of hepatocytes. Inhibition of p38 phosphorylation neutralized CXCL16-induced apoptosis of AML12 cells. Furthermore, CXCL16 knockdown in vivo relieved hepatic injury, which was reversed by activation of p38 phosphorylation. Taken together, silencing of CXCL16 might protect against IR injury during liver transplantation by reducing p38 phosphorylation, highlighting the potential of CXCL16 as a promising target for treatment against hepatic IR injury.

Prognostic and Predictive Role of CXC Chemokine Receptor 4 in Metastatic Colorectal Cancer Patients.

Colorectal cancer (CRC) is the third leading cause of cancer-related death worldwide. About 30% of patients present with metastatic disease involving predominantly the liver and a similar percentage will develop distant metastases later after removal of the primary tumor. In metastatic CRC, chemotherapies and biological drugs have prolonged survival for up to 30 months. However, there is a great need for biomarkers predictive of response and prognosis to optimize treatments. CXC chemokine receptor 4 (CXCR4) is a chemokine receptor; it binds to CXCL12 and plays a central role in colon cancer cells' growth and dissemination. CXCR4 was evaluated in CRC primary tissues by immunohistochemistry. Formalin-fixed, paraffin-embedded 4-μm tissue sections were immunostained using a biotin-streptavidin-peroxidase method and categorized into 2 semiquantitative classes: (i) absence of staining, ≤50% positive cells (negative/low) and (ii) >50% positive cells (high). Associations between clinic-pathologic variables and CXCR4 expression were evaluated using the χ test. The Kaplan-Meier product-limit method was applied to graph overall survival (OS). OS was defined as the time elapsed from diagnosis to death from any cause. Univariate analysis was carried out using the log-rank test. Cox proportional hazards regression was used to analyze the effect of several risk factors on OS. Seventy-eight primary adenocarcinomas were analyzed; 26 were categorized as negative/low and 52 as high. Age, sex, performance status, site of metastases, KRAS mutational status, type of first-line therapy, and a number of therapy lines did not correlate with CXCR4 expression. Although not significant (P=0.0533), high CXCR4 expression was more frequently localized on the right side of the colon. Significant correlations were detected with grading (P=0.0041) and response to first-line anti-epidermal growth factor receptors agents (P<0.0001), bevacizumab (P=0.0029), and chemotherapy alone (P=0.0260). At a median follow-up of 53 months, 77 deaths have been registered. Grading [hazard ratio (HR): 1.42; confidence interval (CI): 0.89-2.28; P<0.0001], KRAS mutational status (HR: 1.73; CI: 1.03-290; P=0.0133), response to first-line chemotherapy (HR: 3.39; CI: 2.10-5.48; P<0.0001), and CXCR4 expression (HR: 3.18; CI: 2.01-5.02; P<0.0001) showed prognostic power at univariate and multivariate analyses. In the present report, we show that CXCR4 expression on the primary tumor is an independent prognostic factor and correlates with response to first-line chemotherapy in metastatic CRC patients.

Reduction of endoglin receptor impairs mononuclear cell-migration.

Aim:To test if the impairment of mononuclear cell (MNC) migration in patients with hereditary hemorrhagic telangiectasia (HHT) is due to the reduction of the endoglin (ENG) receptor on the cell surface and oxidative stress.Methods:MNCs of HHT patients and normal controls were subjected to migration assay. Fractions of MNCs were pre-incubated with antibodies specific to HHT causative genes ENG [hereditary hemorrhagic telangiectasia type 1 (HHT1)] or activin receptor-like kinase 1 [ALK1, hereditary hemorrhagic telangiectasia type 2 (HHT2)], AMD3100 or Diprotin-A to block ENG, ALK1 C-X-C chemokine receptor 4 (CXCR4) or CD26 (increased in HHT1 MNCs) before migration assay. The MNCs were allowed to migrate toward stromal cell-derived factor-1α (SDF-1α) for 18 h. The expression of CXCR4, CD26, superoxide dismutase 1 (SOD1) and glutathione peroxidase 1 (GPX1) in MNCs and nitric oxide levels in the plasma were analyzed.Results:Compared to the controls, fewer HHT1 MNCs and similar number of HHT2 MNCs migrated toward SDF-1α. Diprotin-A pre-treatment improved HHT1 MNC-migration, but had no effect on normal and HHT2 MNCs. Pre-incubation with an anti-ENG antibody reduced the migration of normal MNCs. Diprotin-A did not improve the migration of ENG antibody pre-treated MNCs. Anti-ALK1 antibody had no effect on MNC-migration. AMD3100 treatment reduced normal and HHT MNC-migration. ENG mRNA level was reduced in HHT1 and HHT2 MNCs. ALK1 mRNA was reduced in HHT2 MNCs only. CD26 expression was higher in HHT1 MNCs. Pre-treatment of MNCs with anti-ENG or anti-ALK1 antibody had no effect on CD26 and CXCR4 expression. The expression of antioxidant enzymes, SOD1, was reduced in HHT1 MNCs, which was accompanied with an increase of ROS in HHT MNCs and nitric oxide in HHT1 plasma.Conclusions:Reduction of ENG receptor on MNC surface reduced monocyte migration toward SDF-1α independent of CD26 expression. Increased oxidative stress could alter HHT MNC migration behavior.