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Sample records for aberrant cxcl12 expression

  1. CXCR4, CXCL12 and the relative CXCL12-CXCR4 expression as prognostic factors in colon cancer.

    PubMed

    Stanisavljević, Luka; Aßmus, Jörg; Storli, Kristian Eeg; Leh, Sabine Maria; Dahl, Olav; Myklebust, Mette Pernille

    2016-06-01

    The CXCL12-CXCR4 axis is proposed to mediate metastasis formation. In this study, we examined CXCL12, CXCR4 and the relative CXCL12-CXCR4 expression as prognostic factors in two cohorts of colon cancer patients. Immunohistochemistry (IHC) and in situ hybridization (ISH) were used to study CXCR4, CXCL12 and relative CXCL12-CXCR4 expression in tissue microarrays. Our study included totally 596 patients, 290 in cohort 1 and 306 in cohort 2. For tumour, node, metastasis (TNM) stage III, low nuclear expression of CXCR4 was a positive prognostic factor for 5-year disease-free survival (DFS) in cohort 1 (P = 0.007) and cohort 2 (P = 0.023). In multivariate analysis for stage III, nuclear expression of CXCR4 in cohort 1 was confirmed as a prognostic factor for DFS (hazard ratio (HR), 0.27; 95 % CI, 0.09 to 0.77). For TNM stage III, high cytoplasmic expression of CXCL12 was associated with better 5-year DFS in both cohorts (P = 0.006 and P = 0.006, respectively). We further validated the positive prognostic value of CXCL12 expression for 5-year DFS in stage III with ISH (P = 0.022). For TNM stage III, the relative CXCL12-CXCR4 expression (CXCL12 > CXCR4 vs CXCL12 = CXCR4 vs CXCL12 < CXCR4) was a prognostic factor for 5-year DFS in cohort 1 (92 % vs 46 % vs 31 %, respectively; P < 0.001) and cohort 2 (92 % vs 66 % vs 30 %, respectively; P = 0.006). In conclusion, CXCL12 and relative CXCL12-CXCR4 expression are independent prognostic factors for 5-year DFS in TNM stage III colon cancer.

  2. CXCL12 Mediates Aberrant Costimulation of B Lymphocytes in Warts, Hypogammaglobulinemia, Infections, Myelokathexis Immunodeficiency

    PubMed Central

    Roselli, Giuliana; Martini, Elisa; Lougaris, Vassilios; Badolato, Raffaele; Viola, Antonella; Kallikourdis, Marinos

    2017-01-01

    The Warts, Hypogammaglobulinemia, Infections, Myelokathexis (WHIM) syndrome is an immunodeficiency caused by mutations in chemokine receptor CXCR4. WHIM patient adaptive immunity defects remain largely unexplained. We have previously shown that WHIM-mutant T cells form unstable immunological synapses, affecting T cell activation. Here, we show that, in WHIM patients and WHIM CXCR4 knock-in mice, B cells are more apoptosis prone. Intriguingly, WHIM-mutant B cells were also characterized by spontaneous activation. Searching for a mechanistic explanation for these observations, we uncovered a novel costimulatory effect of CXCL12, the CXCR4 ligand, on WHIM-mutant but not wild-type B cells. The WHIM CXCR4-mediated costimulation led to increased B-cell activation, possibly involving mTOR, albeit without concurrently promoting survival. A reduction in antigenic load during immunization in the mouse was able to circumvent the adaptive immunity defects. These results suggest that WHIM-mutant CXCR4 may lead to spontaneous aberrant B-cell activation, via CXCL12-mediated costimulation, impairing B-cell survival and thus possibly contributing to the WHIM syndrome defects in adaptive immunity. PMID:28928741

  3. A meta-analysis of CXCL12 expression for cancer prognosis.

    PubMed

    Samarendra, Harsh; Jones, Keaton; Petrinic, Tatjana; Silva, Michael A; Reddy, Srikanth; Soonawalla, Zahir; Gordon-Weeks, Alex

    2017-06-27

    CXCL12 (SDF1) is reported to promote cancer progression in several preclinical models and this is corroborated by the analysis of human tissue specimens. However, the relationship between CXCL12 expression and cancer survival has not been systematically assessed. We conducted a systematic review and meta-analysis of studies that evaluated the association between CXCL12 expression and cancer survival. Thirty-eight studies inclusive of 5807 patients were included in the analysis of overall, recurrence-free or cancer-specific survival, the majority of which were retrospective. The pooled hazard ratios (HRs) for overall and recurrence-free survival in patients with high CXCL12 expression were 1.39 (95% CI: 1.17-1.65, P=0.0002) and 1.12 (95% CI: 0.82-1.53, P=0.48) respectively, but with significant heterogeneity between studies. On subgroup analysis by cancer type, high CXCL12 expression was associated with reduced overall survival in patients with oesophagogastric (HR 2.08; 95% CI: 1.31-3.33, P=0.002), pancreatic (HR 1.54; 95% CI: 1.21-1.97, P=0.0005) and lung cancer (HR 1.37; 95% CI: 1.08-1.75, P=0.01), whereas in breast cancer patients high CXCL12 expression conferred an overall survival advantage (HR 0.5; 95% CI: 0.38-0.66, P<0.00001). Determination of CXCL12 expression has the potential to be of use as a cancer biomarker and adds prognostic information in various cancer types. Prospective or prospective-retrospective analyses of CXCL12 expression in clearly defined cancer cohorts are now required to advance our understanding of the relationship between CXCL12 expression and cancer outcome.

  4. Placental development during early pregnancy: Effects of embryo origin on expression of chemokine ligand twelve (CXCL12).

    PubMed

    Quinn, K E; Reynolds, L P; Grazul-Bilska, A T; Borowicz, P P; Ashley, R L

    2016-07-01

    The aim was to localize chemokine ligand twelve (CXCL12) in sheep placental tissues during early gestation and after assisted reproductive technologies (ART). Uteri were collected from naturally (NAT) mated ewes and ewes receiving embryo transfer (ET), in vitro fertilization (IVF) or in vitro activation (IVA). CXCL12 was immunolocalized to endometrial stroma, glands, and trophoblast. Greater CXCL12 immunoreactivity was present in trophoblast on day 22 and 24 and in NAT ewes compared to IVF and IVA. Increased CXCL12 expression suggests CXCL12 promotes implantation and placentation. Decreased CXCL12 in IVF and IVA embryos, may compromise pregnancy establishment when utilizing ART methods.

  5. Differential Expression of CXCL12 and CXCR4 During Human Fetal Neural Progenitor Cell Differentiation

    PubMed Central

    Peng, Hui; Kolb, Ryan; Kennedy, J. E.

    2007-01-01

    Stromal cell-derived factor 1 alpha (SDF-1α, CXCL12) and its receptor CXCR4 play an important role in the central nervous system (CNS) development and adulthood by mediating cell migration, enhancing precursor cell proliferation, assisting in neuronal circuit formation, and possibly regulating migration during repair. The expression pattern of CXCR4 and CXCL12 during neurogenesis has not been thoroughly elucidated. In this study, we investigated the expression of CXCL12 and CXCR4 during neural progenitor cells (NPC) differentiation by microarray analysis and reverse transcriptase-polymerase chain reaction (RT-PCR) using human fetal NPC as a model system. The production of CXCL12 was measured by enzyme-linked immunosorbent assay (ELISA). CXCR4 expression was determined by florescence-activated cell sorting (FACS) analysis, immunocytochemical staining, and CXCR4-mediated inhibition of cyclic AMP (cAMP) accumulation. Our data demonstrated that CXCR4 expression is significantly upregulated when NPC are differentiated into neuronal precursors, whereas CXCL12 is upregulated when differentiated into astrocytes. We also provide evidence that CXCR4 localization changes as neurons mature. In neuronal precursors, CXCR4 is localized in both neuronal processes and the cell body, whereas in mature neurons, it is primarily expressed on axons and dendrites. This differential expression of CXCR4 and CXCL12 may be important for the temporal regulation of neuronal migration and circuit formation during development and possibly in adult neurogenesis and repair. PMID:18040858

  6. MicroRNA expression profiles and networks in CXCL12-stimulated human endometrial stromal cells

    PubMed Central

    Mei, Jie; Li, Ming-Qing; Li, Da-Jin; Sun, Hai-Xiang

    2016-01-01

    The chemokine stromal cell-derived factor-1 (C-X-C motif chemokine ligand 12; CXCL12) is important in the recruitment of leukocytes to the peritoneal cavity and the regulation of endometriotic tissue growth in endometriosis patients. However, the alterations in microRNA (miRNA) expression induced by CXCL12 remain to be fully elucidated. The present study evaluated key miRNAs in CXCL12-stimulated endometrial stromal cells (ESCs), and investigated the underlying cellular regulatory mechanisms of CXCL12 in endometriosis by building networks between miRNAs, genes and gene ontologies (GOs). Differential expression of miRNAs and mRNAs induced by CXCL12 stimulation in ESCs was measured using miRNA and gene chips, and it was observed that 35 miRNAs and 1,671 mRNAs were differentially expressed. Using potential target genes of the 35 miRNAs, intersections of these genes were examined and 63 intersection genes were identified. A total of 39 GOs were obtained for these intersection genes, based on information from GO databases, including immune cell chemoattractants, inflammatory and immune responses, and pathological processes of endometriotic lesions in endometriosis. In addition, miRNA-gene networks were built according to the GO and Kyoto Encyclopedia of Genes and Genomes databases. The present study, to the best of our knowledge, provides the most complete miRNAome and mRNAome profiles, and the most detailed investigation of the underlying cellular regulatory mechanisms, of the effects of CXCL12 in endometriosis. These results may facilitate the complete elucidation of the role of CXCL12 in endometriosis, and its underlying epigenetic mechanisms. PMID:27959395

  7. Expression of CXCL12 and its receptor CXCR4 in patients with adenomyosis.

    PubMed

    Li, Juan; Yin, Geping; Chen, Ming; Yang, Shujun; Wu, Aifang; Liang, Jing; Yuan, Zheng

    2017-04-01

    The aim of the present study was to investigate the role of chemokine (C-X-C motif) ligand 12 (CXCL12) and its receptor, chemokine (C-X-C motif) receptor 4 (CXCR4) in the pathogenesis of adenomyosis (AD). Immunohistochemistry and reverse transcription-quantitative polymerase chain reaction analysis were used to measure the protein and mRNA expression of CXCL12 and CXCR4 in eutopic endometrial and ectopic foci tissue samples. Samples from a total of 36 patients with AD (study group) were compared with endometrial tissue samples from 33 patients who underwent uterine fibroids surgery (control group) during the same period. All data are presented as the mean ± standard deviation and were analyzed with SPSS software (version 16.0). Analysis of variance was used for between group analysis and pairwise comparison was performed using Fisher's least significant difference post hoc test. The results of the present study revealed that CXCL12 and CXCR4 protein expression was significantly increased in ectopic foci tissue compared with eutopic endometrial tissue samples from patients with AD. CXCL12 and CXCR4 protein expression in ectopic foci and eutopic endometrial tissue samples were significantly increased compared with the control group (P<0.05 for between group comparisons). No significant differences were identified in CXCL12 and CXCR4 protein expression between the proliferative and secretory phases within each group. Furthermore, CXCL12 and CXCR4 mRNA expression was significantly increased in ectopic foci tissue and eutopic endometrial tissue compared with the control group (P<0.05 for between group comparisons). CXCL12 mRNA expression was markedly increased in ectopic foci tissue compared with eutopic endometrial tissue of patients with AD. The expression of CXCR4 mRNA was significantly increased in eutopic endometrial tissue compared with ectopic foci tissue and the control group (P<0.05 for between group comparisons). No significant differences were identified in

  8. Pathological Expression of CXCL12 at the Blood-Brain Barrier Correlates with Severity of Multiple Sclerosis

    PubMed Central

    McCandless, Erin E.; Piccio, Laura; Woerner, B. Mark; Schmidt, Robert E.; Rubin, Joshua B.; Cross, Anne H.; Klein, Robyn S.

    2008-01-01

    Dysregulation of blood-brain barrier (BBB) function and transendothelial migration of leukocytes are essential components of the development and propagation of active lesions in multiple sclerosis (MS). Animal studies indicate that polarized expression of the chemokine CXCL12 at the BBB prevents leukocyte extravasation into the central nervous system (CNS) and that disruption of CXCL12 polarity promotes entry of autoreactive leukocytes and inflammation. In the present study, we examined expression of CXCL12 and its receptor, CXCR4, within CNS tissues from MS and non-MS patients. Immunohistochemical analysis of CXCL12 expression at the BBB revealed basolateral localization in tissues derived from non-MS patients and at uninvolved sites in tissues from MS patients. In contrast, within active MS lesions, CXCL12 expression was redistributed toward vessel lumena and was associated with CXCR4 activation in infiltrating leukocytes, as revealed by phospho-CXCR4-specific antibodies. Quantitative assessment of CXCL12 expression by the CNS microvasculature established a positive correlation between CXCL12 redistribution, leukocyte infiltration, and severity of histological disease. These results suggest that CXCL12 normally functions to localize infiltrating leukocytes to perivascular spaces, preventing CNS parenchymal infiltration. In the patient cohort studied, altered patterns of CXCL12 expression at the BBB were specifically associated with MS, possibly facilitating trafficking of CXCR4-expressing mononuclear cells into and out of the perivascular space and leading to progression of disease. PMID:18276777

  9. Wnt/β-catenin pathway is required for epithelial to mesenchymal transition in CXCL12 over expressed breast cancer cells.

    PubMed

    Shan, Shumei; Lv, Qiang; Zhao, Yiling; Liu, Chunfeng; Sun, Yingyan; Xi, Kemin; Xiao, Jiayi; Li, Caijuan

    2015-01-01

    CXCL12 is positively associated with the metastasis and prognosis of various human malignancies. Cancer-associated fibroblasts (CAFs), the main cells secreting CXCL12, are capable of inducing epithelial to mesenchymal transition (EMT) of breast cancer cells. However, it has not been completely understood whether CXCL12 is involved in EMT of breast cancer cells and the underlying mechanisms. The present study aimed to investigate the effects of CXCL12 on the EMT and cancer stem cell (CSC)-like phenotypes formation by transfecting pEGFP-N1-CXCL12 plasmid into MCF-7 cells. Real time-PCR and Western blot analysis demonstrated the successful over expression of CXCL12 in MCF-7 cells. Cell counting kit-8 assay, wound healing assay and Transwell invasion analysis confirmed that over expression of CXCL12 significantly promoted the proliferation, migration and invasion in MCF-7 cells (P<0.05). In addition, ALDH activity was dramatically enhanced compared with parental (P<0.001), accompanied by the notably elevated mRNA and protein levels of OCT-4, Nanog, and SOX2 in CXCL12 overexpressed-MCF-7 cells (P<0.001). Furthermore, we observed the down regulation of E-cadherin and up regulation of vimentin, N-cadherin, and α-SMA in CXCL12 overexpressed-MCF-7 cells (P<0.01). Meanwhile, western blot and immunofluorescence assay showed that over expression of CXCL12 activated Wnt/β-catenin pathway to induce EMT of MCF-7 cells, as evidenced by the increased expression of E-cadherin after silencing β-catenin by siRNA interference (P<0.001). Collectively, our findings suggested that over expression of CXCL12 could trigger EMT by activating Wnt/β-catenin pathway and induce CSC-like phenotypes formation to promote the proliferation and metastasis in MCF-7. Hence, CXCL12 may become a promising candidate for breast cancer therapy.

  10. Expression of CXCL12 receptors in B cells from Mexican Mestizos patients with systemic lupus erythematosus

    PubMed Central

    2012-01-01

    Background Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disease characterized by B-cell hyper-reactivity and the production of pathogenic anti-nuclear-directed auto-antibodies (Abs). B-cell ontogeny is partly dependent on the CXCL12/CXCR4 axis for which the contribution to SLE pathogenesis remains unclear. CXCR7, the novel receptor for CXCL12, is differentially expressed among memory B-cell subsets. However, its biological role in SLE remains to be explored. Methods Relative CXCR4 and CXCR7 expression levels were compared by quantitative PCR in leukocytes from blood samples of 41 Mexican Mestizos patients with SLE and 45 ethnicity-matched healthy subjects. Intracellular and membrane expression of both receptors was analyzed by flow cytometry in naive and Ab-secreting B cells. B-cell responsiveness to CXCL12 was investigated using Transwell-based chemotaxis assays. Data were analyzed using the Kruskal-Wallis test for comparisons of values amongst healthy controls and patients with inactive or active SLE, and non-parametrically using the Mann–Whitney U-test for multiple comparisons and unpaired samples. Correlations were determined by Spearman’s ranking. Result SLE leukocytes displayed reduced levels of CXCR4 and CXCR7 transcripts. In SLE patients, a significant defect in CXCR4 expression was detected at the surface of naive and Ab-secreting B cells, associated with an abnormal intracellular localization of the receptor. CXCR7 predominantly localized in cytosolic compartments of B cells from healthy and SLE individuals. Disease activity did not impact on these expression patterns. Altered receptor compartmentalization correlated with an impaired CXCL12-promoted migration of SLE B cells. Conclusions Our data highlight a down-regulation of CXCL12 receptors on circulating B cells from SLE patients that likely influences their migratory behavior and distribution. PMID:23244336

  11. Altered CXCL12 expression reveals a dual role of CXCR4 in osteosarcoma primary tumor growth and metastasis.

    PubMed

    Neklyudova, Olga; Arlt, Matthias J E; Brennecke, Patrick; Thelen, Marcus; Gvozdenovic, Ana; Kuzmanov, Aleksandar; Robl, Bernhard; Botter, Sander M; Born, Walter; Fuchs, Bruno

    2016-08-01

    Better understanding of the molecular mechanisms of metastasis-the major cause of death in osteosarcoma (OS)-is a key for the development of more effective metastasis-suppressive therapy. Here, we investigated the biological relevance of the CXCL12/CXCR4 axis in OS. We interfered with CXCL12/CXCR4 signaling in CXCR4-expressing human 143-B OS cells through stable expression of CXCL12, of its competitive antagonist P2G, or of CXCL12-KDEL, designed to retain CXCR4 within the cell. Intratibial OS xenograft mouse model metastasizing to the lung was used to assess tumorigenic and metastatic potential of the manipulated cell lines. Constitutive expression of native CXCL12 promoted lung metastasis without affecting tumor growth. Stable expression of P2G or CXCL12-KDEL significantly accelerated tumor growth but diminished lung metastasis. Tumors grown from P2G- or CXCL12-KDEL-expressing cells contained higher levels of CXCR4-encoding mRNA going along with a higher percentage of CXCR4-expressing tumor cells. Lung metastases of all groups were predominantly enriched with CXCR4-expressing tumor cells. Higher abundance of CXCR4 possibly contributed to increased local retention of tumor cells by bone marrow-derived CXCL12, reflected in the increased primary tumor growth and decreased number of lung metastases in P2G and CXCL12-KDEL groups. Higher percentage of CXCR4-expressing lung metastatic cells compared to the corresponding primary tumors point to important functions of the CXCL12/CXCR4 axis in late steps of metastasis. In conclusion, based on the here reported results, local treatment of lung metastases with novel CXCR4-targeting therapeutics might be considered and favored over anti-CXCR4 systemic therapy.

  12. CXCL12 methylation-mediated epigenetic regulation of gene expression in papillary thyroid carcinoma

    PubMed Central

    Zhang, Sijia; Wang, Yihan; Chen, Meijun; Sun, Lulu; Han, Jun; Elena, V. Kazakova; Qiao, Hong

    2017-01-01

    Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer, and its incidence rate is rapidly growing. It is necessary to understand the pathogenesis of PTC to develop effective diagnosis methods. Promoter methylation has been recognized to contribute to the alterations in gene expression observed in tumorigenesis. Our RNA-seq data identified 1191 differentially expressed mRNAs and 147 differentially expressed lncRNAs in PTC. Next, promoter methylation of these genes was detected by reduced representation bisulfite sequencing (RRBS) technology and comprehensively analyzed to identify differential methylation. In total, 14 genes (13 mRNAs and 1 lncRNA), in which methylation was intimately involved in regulating gene expression, were proposed as novel diagnostic biomarkers. To gain insights into the relationships among these 14 genes, a core co-function network was constructed based on co-expression, co-function and co-methylation data. Notably, CXCL12 was identified as an essential gene in the network that was closely connected with the other genes. These data suggested that CXCL12 down-regulation in PTC may be caused by promoter hypermethylation. Our study was the first to perform an RRBS analysis for PTC and suggested that CXCL12 may contribute to PTC development by methylation-mediated epigenetic regulation of gene expression. PMID:28272462

  13. CXCL12 methylation-mediated epigenetic regulation of gene expression in papillary thyroid carcinoma.

    PubMed

    Zhang, Sijia; Wang, Yihan; Chen, Meijun; Sun, Lulu; Han, Jun; Elena, V Kazakova; Qiao, Hong

    2017-03-08

    Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer, and its incidence rate is rapidly growing. It is necessary to understand the pathogenesis of PTC to develop effective diagnosis methods. Promoter methylation has been recognized to contribute to the alterations in gene expression observed in tumorigenesis. Our RNA-seq data identified 1191 differentially expressed mRNAs and 147 differentially expressed lncRNAs in PTC. Next, promoter methylation of these genes was detected by reduced representation bisulfite sequencing (RRBS) technology and comprehensively analyzed to identify differential methylation. In total, 14 genes (13 mRNAs and 1 lncRNA), in which methylation was intimately involved in regulating gene expression, were proposed as novel diagnostic biomarkers. To gain insights into the relationships among these 14 genes, a core co-function network was constructed based on co-expression, co-function and co-methylation data. Notably, CXCL12 was identified as an essential gene in the network that was closely connected with the other genes. These data suggested that CXCL12 down-regulation in PTC may be caused by promoter hypermethylation. Our study was the first to perform an RRBS analysis for PTC and suggested that CXCL12 may contribute to PTC development by methylation-mediated epigenetic regulation of gene expression.

  14. PGE2-Induced CXCL12 Production and CXCR4 Expression Controls the Accumulation of Human MDSCs in Ovarian Cancer Environment

    PubMed Central

    Obermajer, Nataša; Muthuswamy, Ravikumar; Odunsi, Kunle; Edwards, Robert P.; Kalinski, Pawel

    2016-01-01

    Signals mediated by CXCL12 (SDF1) and its receptor CXCR4 are centrally involved in cancer progression, both directly by activating cancer cells and indirectly by inducing angiogenesis plus recruiting T regulatory and plasmacytoid dendritic immune cells. Here, we show that in ascites isolated from ovarian cancer patients, both CXCL12 and CXCR4 are controlled by the tumor-associated inflammatory mediator prostaglandin E2 (PGE2), which attracts myeloid-derived suppressor cells (MDSC) into the ascites microenvironment. In this setting, PGE2 was essential both for expression of functional CXCR4 in cancer-associated MDSCs and for production of its ligand CXCL12. Frequencies of CD11b+CD14+CD33+CXCR4+ MDSCs closely correlated with CXCL12 and PGE2 levels in patient ascites. MDSCs migrated toward ovarian cancer ascites in a CXCR4-dependent manner that required COX2 activity and autocrine PGE2 production. Inhibition of COX2 or the PGE2 receptors EP2/EP4 in MDSCs suppressed expression of CXCR4 and MDSC responsiveness to CXCL12 or ovarian cancer ascites. Similarly, COX2 inhibition also blocked CXCL12 production in the ovarian cancer environment and its ability to attract MDSCs. Together, our findings elucidate a central role for PGE2 in MDSC accumulation triggered by the CXCL12-CXCR4 pathway, providing a powerful rationale to target PGE2 signaling in ovarian cancer therapy. PMID:22025564

  15. SMAD signaling regulates CXCL12 expression in the bone marrow niche, affecting homing and mobilization of hematopoietic progenitors.

    PubMed

    Khurana, Satish; Melacarne, Alessia; Yadak, Rana; Schouteden, Sarah; Notelaers, Tineke; Pistoni, Mariaelena; Maes, Christa; Verfaillie, Catherine M

    2014-11-01

    We recently demonstrated that ex vivo activation of SMAD-independent bone morphogenetic protein 4 (BMP4) signaling in hematopoietic stem/progenitor cells (HSPCs) influences their homing into the bone marrow (BM). Here, we assessed whether alterations in BMP signaling in vivo affects adult hematopoiesis by affecting the BM niche. We demonstrate that systemic inhibition of SMAD-dependent BMP signaling by infusion of the BMP antagonist noggin (NGN) significantly increased CXCL12 levels in BM plasma leading to enhanced homing and engraftment of transplanted HSPCs. Conversely, the infusion of BMP7 but not BMP4, resulted in decreased HSPC homing. Using ST2 cells as an in vitro model of BM niche, we found that incubation with neutralizing anti-BMP4 antibodies, NGN, or dorsomorphin (DM) as well as knockdown of Smad1/5 and Bmp4, all enhanced CXCL12 production. Chromatin immunoprecipitation identified the SMAD-binding element in the CXCL12 promoter to which SMAD4 binds. When deleted, increased CXCL12 promoter activity was observed, and NGN or DM no longer affected Cxcl12 expression. Interestingly, BMP7 infusion resulted in mobilization of only short-term HSCs, likely because BMP7 affected CXCL12 expression only in osteoblasts but not in other niche components. Hence, we describe SMAD-dependent BMP signaling as a novel regulator of CXCL12 production in the BM niche, influencing HSPC homing, engraftment, and mobilization. © 2014 AlphaMed Press.

  16. SMAD signaling regulates CXCL12 expression in the bone marrow niche, affecting homing and mobilization of hematopoietic progenitors

    PubMed Central

    Khurana, Satish; Melacarne, Alessia; Yadak, Rana; Schouteden, Sarah; Notelaers, Tineke; Pistoni, Mariaelena; Maes, Christa; Verfaillie, Catherine M

    2014-01-01

    We recently demonstrated that ex vivo activation of SMAD-independent BMP4 signaling in hematopoietic stem/progenitor cells (HSPCs) influences their homing into the bone marrow (BM). We here assessed if alterations in BMP signaling in vivo affects adult hematopoiesis by affecting the BM niche. We demonstrate that systemic inhibition of SMAD-dependent BMP signaling by infusion of the BMP antagonist Noggin (NGN) significantly increased CXCL12 levels in BM plasma leading to enhanced homing and engraftment of transplanted HSPCs. Conversely, the infusion of BMP7 but not BMP4, resulted in decreased HSPC homing. Using ST2 cells as an in vitro model of BM niche, we found that incubation with neutralizing anti-BMP4 antibodies, NGN or dorsomorphin (DM) as well as knockdown of Smad1/5 and Bmp4, all enhanced CXCL12 production. Chromatin immunoprecipitation identified the SMAD-binding element in the CXCL12 promoter to which SMAD4 binds. When deleted, increased CXCL12 promoter activity was observed, and NGN or DM no longer affected Cxcl12 expression. Interestingly, BMP7 infusion resulted in mobilization of only short-term HSCs, likely because BMP7 affected CXCL12 expression only in osteoblasts but not in other niche components. Hence, we describe SMAD-dependent BMP signaling as a novel regulator of CXCL12 production in the BM niche, influencing HSPC homing, engraftment and mobilization. PMID:25069965

  17. Expression of the CXCL12/CXCR4 chemokine axis predicts regional control in head and neck squamous cell carcinoma.

    PubMed

    León, Xavier; Diez, Santiago; García, Jacinto; Lop, Joan; Sumarroca, Anna; Quer, Miquel; Camacho, Mercedes

    2016-12-01

    Expression of the CXCL12/CXCR4 chemokine axis has been related with the appearance of metastatic recurrence survival, including regional and distant recurrence, in patients with head and neck squamous cell carcinoma (HNSCC). RT-PCR was used to determine mRNA expression levels of CXCL12 and CXCR4 in biopsy tumor samples in 111 patients with HNSCC. Five-year regional recurrence-free survival for patients with low CXCR4 expression (n = 39, 31.5 %) was 97.4 %, for patients with high CXCR4/high CXCL12 expression (n = 22, 19.8 %) it was 94.7 %, and for patients with high CXCR4/low CXCL12 expression (n = 50, 45.0 %) it was 63.3 %. We found significant differences in the regional recurrence-free survival according to CXCR4/CXCL12 expression values (P = 0.001). HNSCC patients with high CXCR4 and low CXCL12 expression values had a significantly higher risk of regional recurrence and could benefit from a more intense treatment of lymph node areas in the neck.

  18. Expression of chemokine CXCL12 and its receptor CXCR4 in folliculostellate (FS) cells of the rat anterior pituitary gland: the CXCL12/CXCR4 axis induces interconnection of FS cells.

    PubMed

    Horiguchi, Kotaro; Ilmiawati, Cimi; Fujiwara, Ken; Tsukada, Takehiro; Kikuchi, Motoshi; Yashiro, Takashi

    2012-04-01

    The anterior pituitary gland is composed of five types of hormone-producing cells plus folliculostellate (FS) cells, which do not produce classical anterior pituitary hormones. FS cells are interconnected by cytoplasmic processes and encircle hormone-producing cells or aggregate homophilically. Using living-cell imaging of primary culture, we recently reported that some FS cells precisely extend their cytoplasmic processes toward other FS cells and form interconnections with them. These phenomena suggest the presence of a chemoattractant factor that facilitates the interconnection. In this study, we attempted to discover the factor that induces interconnection of FS cells and succeeded in identifying chemokine (CXC)-L12 and its receptor CXCR4 as potential candidate molecules. CXCL12 is a chemokine of the CXC subfamily. It exerts its effects via CXCR4, a G protein-coupled receptor. The CXCL12/CXCR4 axis is a potent chemoattractant for many types of neural cells. First, we revealed that CXCL12 and CXCR4 are expressed by FS cells in rat anterior pituitary gland. Next, to clarify the function of the CXCL12/CXCR4 axis in FS cells, we observed living anterior pituitary cells in primary culture with specific CXCL12 inhibitor or CXCR4 antagonist and noted that extension of cytoplasmic processes and interconnection of FS cells were inhibited. Finally, we examined FS cell migration and invasion by using Matrigel matrix assays. CXCL12 treatment resulted in markedly increased FS cell migration and invasion. These data suggest that FS cells express chemokine CXCL12 and its receptor CXCR4 and that the CXCL12/CXCR4 axis evokes interconnection of FS cells.

  19. Expression of CXCR4 and CXCL12 and their correlations to the cell proliferation and angiogenesis in mycosis fungoides

    PubMed Central

    Maj, Joanna; Hałoń, Agnieszka; Woźniak, Zbigniew; Plomer-Niezgoda, Ewa; Reich, Adam

    2015-01-01

    Introduction Chemokines play an important role in tumor growth, invasion and metastasis. The CXCR4/CXCL12 axis has been implicated in development of both solid tumors and hematological malignancies and is also relevant in the pathogenesis of the most common primary cutaneous T-cell lymphoma, mycosis fungoides (MF). Aim To evaluate the expression of CXCR4 and CXCL12 in MF and to examine their associations with cell proliferation and angiogenesis. Material and methods The material for the study consisted of skin samples obtained from 56 patients with MF and 20 healthy volunteers. The expression of CXCR4 and CXCL12 was assessed by immunohistochemistry on the paraffin blocks and compared to the expression of angiogenesis marker (CD34) and proliferation indicators (Ki-67, AgNORs). Results The expression of chemokine CXCL12 and its receptor CXCR4 was significantly higher in MF than in the healthy skin (p < 0.001). There was no significant difference between early and advanced stages of MF. Similarly, there was no statistically important correlation between the expression of CXCR4/CXCL12 and angiogenesis and proliferation markers, however a significant correlation between CD34 and AgNORs expression was found (p < 0.001). Conclusions The CXCR4/CXCL12 axis seems to play an important role in MF development in the early as well as in the advanced stages of the disease. Therefore, the CXCR4/CXCL12 axis seems to be an interesting potential target for the future strategies of new drug development, giving hope for more efficacious therapies for mycosis fungoides. PMID:26755907

  20. CXCL12 and CXCR4, but not CXCR7, are primarily expressed by the stroma in head and neck squamous cell carcinoma.

    PubMed

    Clatot, Florian; Cornic, Marie; Berghian, Anca; Marchand, Vinciane; Choussy, Olivier; El Ouakif, Faissal; François, Arnaud; Ruminy, Philippe; Laberge-Le-Couteulx, Sophie; Picquenot, Jean-Michel; Jardin, Fabrice

    2015-01-01

    The CXCL12/CXCR4 axis is involved in numerous models of metastatic dissemination, including head and neck squamous cell carcinoma (HNSCC). We assessed the relative expressions of CXCL12, CXCR4 and CXCR7 in the stroma and the tumour of HNSCC, and evaluated the methylation status of the CXCL12 promoter.Snap-frozen, HPV negative HNSCC samples were micro-dissected to isolate the tumoural and stromal compartments. The expression levels of CXCL12, CXCR4 and CXCR7 were assessed by qRT-PCR, and the methylation level of the CXCL12 promoter was evaluated by pyrosequencing.In total, 23 matched tumour/stroma samples were analysed. Higher expressions of CXCR4 and CXCL12 were observed in the stroma (p = 0.012 and p < 0.0001, respectively). No significant difference in expression was observed for CXCR7. A high methylation level (>40%) of the CXCL12 promoter was observed in only a few tumoural samples (5/23) and was associated with a lower expression of the gene (p = 0.03).Stromal cells, rather than the tumour itself, are mainly responsible for the expression of both CXCL12 and CXCR4 expression in HNSCC. CXCR7 expression did not differ between the two compartments and was not related to CXCL12 or CXCR4 expression. Finally, the methylation of the CXCL12 promoter could only explain the low intra-tumoural expression of this gene in 20% of cases.

  1. Expression analysis of angiogenic growth factors and biological axis CXCL12/CXCR4 axis in idiopathic pulmonary fibrosis.

    PubMed

    Antoniou, Katerina M; Soufla, Giannoula; Lymbouridou, Rena; Economidou, Foteini; Lasithiotaki, Ismini; Manousakis, Manolis; Drositis, Ioannis; Spandidos, Demetrios A; Siafakas, Nikolaos M

    2010-01-01

    Idiopathic pulmonary fibrosis (IPF) is associated with aberrant repair, persistence of collagen deposition, and the development of vascular remodeling. However, the role of angiogenesis in the pathogenesis of IPF is still undetermined. The aim of this study was to evaluate the combined mRNA expression of vascular endothelial growth factor A (VEGFA), fibroblast growth factor 2 (FGF2), insulin-like growth factor 1 (IGF1) epidermal growth factor (EGF), and its receptor (EGFR) in lung tissue obtained from IPF patients. We have also investigated the expression of chemokine CXCL12/stromal cell-derived factor-1 (SDF-1) and its receptor, CXCR4, to identify alterations that maybe implicated in the pathogenesis of IPF. The subjects studied consisted of two distinct groups: patients with IPF (n = 25) and subjects (control) undergoing thoracic surgery for reasons other than interstitial lung disease (n = 10). Expression analysis of the aforementioned growth factors and biological axis CXCL12/CXR4 analysis were performed using real-time RT-PCR. IGF-1, EGF, and FGF2 mRNA levels are significantly decreased in the patients compared to the controls (p = 0.028, p = 0.023 and p = 0.009, respectively). SDF1-TR1 and SDF1-TR2 transcript levels were significantly lower in patients compared to controls (p = 0.017 and p = 0.001). Significant coexpression of VEGF mRNA with IGF mRNA was observed in the group of the patients (p = 0.017). An additional coexpression of VEGF mRNA with SDF1-TR1 mRNA was demonstrated(p = 0.030). Our results show a downregulation in angiogenetic mechanisms in IPF. However, our results should be further verified by measuring other angiogenetic pathways in more samples.

  2. MicroRNA expression profiles and networks in CXCL12‑stimulated human endometrial stromal cells.

    PubMed

    Mei, Jie; Li, Ming-Qing; Li, Da-Jin; Sun, Hai-Xiang

    2017-01-01

    The chemokine stromal cell-derived factor-1 (C-X-C motif chemokine ligand 12; CXCL12) is important in the recruitment of leukocytes to the peritoneal cavity and the regulation of endometriotic tissue growth in endometriosis patients. However, the alterations in microRNA (miRNA) expression induced by CXCL12 remain to be fully elucidated. The present study evaluated key miRNAs in CXCL12‑stimulated endometrial stromal cells (ESCs), and investigated the underlying cellular regulatory mechanisms of CXCL12 in endometriosis by building networks between miRNAs, genes and gene ontologies (GOs). Differential expression of miRNAs and mRNAs induced by CXCL12 stimulation in ESCs was measured using miRNA and gene chips, and it was observed that 35 miRNAs and 1,671 mRNAs were differentially expressed. Using potential target genes of the 35 miRNAs, intersections of these genes were examined and 63 intersection genes were identified. A total of 39 GOs were obtained for these intersection genes, based on information from GO databases, including immune cell chemoattractants, inflammatory and immune responses, and pathological processes of endometriotic lesions in endometriosis. In addition, miRNA‑gene networks were built according to the GO and Kyoto Encyclopedia of Genes and Genomes databases. The present study, to the best of our knowledge, provides the most complete miRNAome and mRNAome profiles, and the most detailed investigation of the underlying cellular regulatory mechanisms, of the effects of CXCL12 in endometriosis. These results may facilitate the complete elucidation of the role of CXCL12 in endometriosis, and its underlying epigenetic mechanisms.

  3. CXCL12-mediated induction of plasminogen activator inhibitor-1 expression in human CXCR4 positive astroglioma cells.

    PubMed

    Oh, Jae-Wook; Olman, Mitchell; Benveniste, Etty Nadia

    2009-04-01

    Glioblastoma is the most malignant and common brain tumor. To promote their growth, these glioma cells secrete a variety of soluble factors including plasminogen activator inhibitor-1 (PAI-1), which functions as an inhibitor of plasminogen activators. We report here with the basis of microarray gene expression analysis that CXCR4 expressing glioma cells are capable of expressing PAI-1 mRNA and protein upon CXCL12 stimulation. Pretreatment with U0126, an inhibitor of mitogen activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase (MEK) 1/2, abrogated CXCL12-induced PAI-1 expression. Pertussis toxin (PTX), an inhibitor of Galpha(i) proteins, also had inhibitory effects, indicating that the activation of Galpha(i) and ERK MAPK are required for this response. Interestingly, CXCL12 showed additive effects with another PAI-1 inducers, tumor necrosis factor (TNF)-alpha and/or tumor growth factor (TGF)-beta1, in increasing PAI-1 expression. These results indicate that CXCL12/CXCR4 signaling in glioma cells may be another mechanism for these cells to express PAI-1, which may be involved in angiogenesis and tumor invasion in brain tumors.

  4. CXCR4 and CXCL12 Expression in Rectal Tumors of Stage IV Patients Before and After Local Radiotherapy and Systemic Neoadjuvant Treatment.

    PubMed

    Tamas, Karin; Domanska, Urszula M; van Dijk, Tonnis H; Timmer-Bosscha, Hetty; Havenga, Klaas; Karrenbeld, Arend; Sluiter, Wim J; Beukema, Jannet C; van Vugt, Marcel A T M; de Vries, Elisabeth G E; Hospers, Geke A P; Walenkamp, Annemiek M E

    2015-01-01

    Metastatic rectal cancer patients could benefit from novel therapeutic approaches. The signaling network formed by chemokines and their receptors can promote metastasis and resistance to current anticancer treatments. This study assessed the expression of chemokine receptor 4 (CXCR4) and its ligand CXCL12 immuhistochemically in stage IV rectal tumors. Paraffin-embedded primary tumor collected before and after local radiotherapy and systemic treatment with bevacizumab, oxaliplatin and capecitabine was analyzed. Receptor and ligand expression was assessed in the cytoplasm and nucleus of tumor, stromal and normal rectal crypt cells. Baseline expression of CXCR4 and CXCL12 was correlated with patients' pathologic response to treatment. At diagnosis (n=46), 89% of the rectal tumors expressed cytoplasmic CXCR4 and 81% CXCL12. Nuclear CXCR4 expression in tumor cells was present in 30% and nuclear CXCL12 expression in 35% of the tumors. After radiochemotherapy and administration of bevacizumab, nuclear CXCL12 expression was observed in 79% of residual tumors, as compared to 31% of the paired tumor samples expressing nuclear CXCL12 before treatment (P=0.001). There were no differences in CXCR4 or CXCL12 expression at baseline between the patients who had (n=9) and did not have (n=30) a pathologic complete response. Our results show that CXCR4 and CXCL12 are extensively expressed in primary rectal tumors of patients presenting with metastatic disease, while radiochemotherapy and bevacizumab further upregulate CXCL12 expression. These data indicate the importance of the CXCR4/CXCL12 axis in rectal tumor biology, and may suggest the CXCR4/CXCL12 receptor-ligand pair as a potential therapeutic target in metastatic rectal cancer.

  5. CXCR7 Is Highly Expressed in Acute Lymphoblastic Leukemia and Potentiates CXCR4 Response to CXCL12

    PubMed Central

    Melo, Rita de Cássia Carvalho; Longhini, Ana Leda; Bigarella, Carolina Louzão; Baratti, Mariana Ozello; Traina, Fabiola; Favaro, Patrícia; de Melo Campos, Paula; Saad, Sara Teresinha Olalla

    2014-01-01

    Recently, a novel CXCL12-binding receptor, has been identified. This CXCL12-binding receptor commonly known as CXCR7 (CXC chemokine receptor 7), has lately, based on a novel nomenclature, has received the name ACKR3 (atypical chemokine receptor 3). In this study, we aimed to investigate the expression of CXCR7 in leukemic cells, as well as its participation in CXCL12 response. Interesting, we clearly demonstrated that CXCR7 is highly expressed in acute lymphoid leukemic cells compared with myeloid or normal hematopoietic cells and that CXCR7 contributed to T-acute lymphoid leukemic cell migration induced by CXCL12. Moreover, we showed that the cellular location of CXCR7 varied among T-lymphoid cells and this finding may be related to their migration capacity. Finally, we hypothesized that CXCR7 potentiates CXCR4 response and may contribute to the maintenance of leukemia by initiating cell recruitment to bone marrow niches that were once occupied by normal hematopoietic stem cells. PMID:24497931

  6. Polarized CD163+ tumor-associated macrophages are associated with increased angiogenesis and CXCL12 expression in gastric cancer.

    PubMed

    Park, Jae-Young; Sung, Ji-Youn; Lee, Juhie; Park, Yong-Koo; Kim, Youn Wha; Kim, Gou Young; Won, Kyu Yeoun; Lim, Sung-Jig

    2016-06-01

    Tumor-associated macrophages (TAMs) play a significant role in tumor progression and angiogenesis. However, the prognostic value of TAMs in different histologic locations of gastric cancer (GC) is still unknown. We evaluated the distribution of TAMs in different histologic locations to investigate its importance in predicting prognosis and the relationship with angiogenesis and CXCL12 expression in GC. The distribution of TAMs and microvessel density (MVD) in 113 GC samples were evaluated by immunohistochemical staining of CD163 and CD105, respectively. The extent of TAM distribution in the tumor was categorized into three groups: infiltrated TAMs in the tumor nest (TN), tumor stroma (TS) and invasive tumor margin (TM). The expression of CXCL12 in GC were evaluated by immunohistochemical staining of tissues from 88 GC samples. The increased CD163+ TAMs in TS and TM were closely correlated with tumor size, depth of invasion, TNM stage, lymph node metastasis, and lymphovascular invasion. TAMs in TN was not related with any clinicopathologic characteristics except histologic differentiation. The high infiltration of CD163+ TAMs in TS and TM were significantly correlated with poor overall survival. Regardless of location, CD163+ TAMs were significantly correlated with increased MVD. CXCL12 expression was significantly associated with increased CD163+ TAMs in TS and TM. TAMs in different histologic locations in GC were related to distinct aspects of tumor progression. CD163+ TAMs in TS and TM are associated with tumor progression and CXCL12 expression in GC. TAMs may be involved in tumor progression through the angiogenesis. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  7. CXCR4 and CXCL12 are inversely expressed in colorectal cancer cells and modulate cancer cell migration, invasion and MMP-9 activation

    SciTech Connect

    Brand, Stephan . E-mail: stephan.brand@med.uni-muenchen.de; Dambacher, Julia; Beigel, Florian; Olszak, Torsten; Diebold, Joachim; Otte, Jan-Michel; Goeke, Burkhard; Eichhorst, Soeren T.

    2005-10-15

    Colorectal cancer (CRC) is characterized by a distinct metastatic pattern resembling chemokine-induced leukocyte trafficking. This prompted us to investigate expression, signal transduction and specific functions of the chemokine receptor CXCR4 in CRC cells and metastases. Using RT-PCR analysis and Western blotting, we demonstrated CXCR4 and CXCL12 expression in CRC and CRC metastases. Cell differentiation increases CXCL12 mRNA levels. Moreover, CXCR4 and its ligand are inversely expressed in CRC cell lines with high CXCR4 and low or not detectable CXCL12 expression. CXCL12 activates ERK-1/2, SAPK/JNK kinases, Akt and matrix metalloproteinase-9. These CXCL12-induced signals mediate reorganization of the actin cytoskeleton resulting in increased cancer cell migration and invasion. Moreover, CXCL12 increases vascular endothelial growth factor (VEGF) expression and cell proliferation but has no effect on CRC apoptosis. Therefore, the CXCL12/CXCR4 system is an important mediator of invasion and metastasis of CXCR4 expressing CRC cells.

  8. CXCL12 induces connective tissue growth factor expression in human lung fibroblasts through the Rac1/ERK, JNK, and AP-1 pathways.

    PubMed

    Lin, Chien-Huang; Shih, Chung-Huang; Tseng, Chih-Chieh; Yu, Chung-Chi; Tsai, Yuan-Jhih; Bien, Mauo-Ying; Chen, Bing-Chang

    2014-01-01

    CXCL12 (stromal cell-derived factor-1, SDF-1) is a potent chemokine for homing of CXCR4+ fibrocytes to injury sites of lung tissue, which contributes to pulmonary fibrosis. Overexpression of connective tissue growth factor (CTGF) plays a critical role in pulmonary fibrosis. In this study, we investigated the roles of Rac1, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and activator protein-1 (AP-1) in CXCL12-induced CTGF expression in human lung fibroblasts. CXCL12 caused concentration- and time-dependent increases in CTGF expression and CTGF-luciferase activity. CXCL12-induced CTGF expression was inhibited by a CXCR4 antagonist (AMD3100), small interfering RNA of CXCR4 (CXCR4 siRNA), a dominant negative mutant of Rac1 (RacN17), a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor (PD98059), a JNK inhibitor (SP600125), a p21-activated kinase inhibitor (PAK18), c-Jun siRNA, and an AP-1 inhibitor (curcumin). Treatment of cells with CXCL12 caused activations of Rac1, Rho, ERK, and c-Jun. The CXCL12-induced increase in ERK phosphorylation was inhibited by RacN17. Treatment of cells with PD98059 and SP600125 both inhibited CXCL12-induced c-Jun phosphorylation. CXCL12 caused the recruitment of c-Jun and c-Fos binding to the CTGF promoter. Furthermore, CXCL12 induced an increase in α-smooth muscle actin (α-SMA) expression, a myofibroblastic phenotype, and actin stress fiber formation. CXCL12-induced actin stress fiber formation and α-SMA expression were respectively inhibited by AMD3100 and CTGF siRNA. Taken together, our results suggest that CXCL12, acting through CXCR4, activates the Rac/ERK and JNK signaling pathways, which in turn initiates c-Jun phosphorylation, and recruits c-Jun and c-Fos to the CTGF promoter and ultimately induces CTGF expression in human lung fibroblasts. Moreover, overexpression of CTGF mediates CXCL12-induced α-SMA expression.

  9. CXCL12 Induces Connective Tissue Growth Factor Expression in Human Lung Fibroblasts through the Rac1/ERK, JNK, and AP-1 Pathways

    PubMed Central

    Tseng, Chih-Chieh; Yu, Chung-Chi; Tsai, Yuan-Jhih; Bien, Mauo-Ying; Chen, Bing-Chang

    2014-01-01

    CXCL12 (stromal cell-derived factor-1, SDF-1) is a potent chemokine for homing of CXCR4+ fibrocytes to injury sites of lung tissue, which contributes to pulmonary fibrosis. Overexpression of connective tissue growth factor (CTGF) plays a critical role in pulmonary fibrosis. In this study, we investigated the roles of Rac1, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and activator protein-1 (AP-1) in CXCL12-induced CTGF expression in human lung fibroblasts. CXCL12 caused concentration- and time-dependent increases in CTGF expression and CTGF-luciferase activity. CXCL12-induced CTGF expression was inhibited by a CXCR4 antagonist (AMD3100), small interfering RNA of CXCR4 (CXCR4 siRNA), a dominant negative mutant of Rac1 (RacN17), a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor (PD98059), a JNK inhibitor (SP600125), a p21-activated kinase inhibitor (PAK18), c-Jun siRNA, and an AP-1 inhibitor (curcumin). Treatment of cells with CXCL12 caused activations of Rac1, Rho, ERK, and c-Jun. The CXCL12-induced increase in ERK phosphorylation was inhibited by RacN17. Treatment of cells with PD98059 and SP600125 both inhibited CXCL12-induced c-Jun phosphorylation. CXCL12 caused the recruitment of c-Jun and c-Fos binding to the CTGF promoter. Furthermore, CXCL12 induced an increase in α-smooth muscle actin (α-SMA) expression, a myofibroblastic phenotype, and actin stress fiber formation. CXCL12-induced actin stress fiber formation and α-SMA expression were respectively inhibited by AMD3100 and CTGF siRNA. Taken together, our results suggest that CXCL12, acting through CXCR4, activates the Rac/ERK and JNK signaling pathways, which in turn initiates c-Jun phosphorylation, and recruits c-Jun and c-Fos to the CTGF promoter and ultimately induces CTGF expression in human lung fibroblasts. Moreover, overexpression of CTGF mediates CXCL12-induced α-SMA expression. PMID:25121739

  10. Association of rs1801157 single nucleotide polymorphism of CXCL12 gene in breast cancer in Pakistan and in-silico expression analysis of CXCL12-CXCR4 associated biological regulatory network.

    PubMed

    Khalid, Samra; Hanif, Rumeza

    2017-01-01

    C-X-C chemokine ligand 12 (CXCL12) has important implications in breast cancer (BC) pathogenesis. It is selectively expressed on B and T lymphocytes and is involved in hematopoiesis, thymocyte trafficking, stem cell motility, neovascularization, and tumorigenesis. The single nucleotide polymorphism (SNP) rs1801157 of CXCL12 gene has been found to be associated with higher risk of BC. Our study focuses on the genotypic and allelic distribution of SNP (rs1801157; G/A) in Pakistani population as well as its association with the clinico-pathological features. The association between rs1801157 genotypes (G/A) and BC risks was assessed by a multivariate logistic regression (MLR) analysis. Genotyping was performed in both healthy individuals and patients of BC using PCR-restriction fragment length polymorphism (PCR-RFLP) method. Furthermore, in-silico approaches were adapted to investigate the association of CXCL12 and its receptor CXCR4 with genes/proteins involved in BC signalling. Significant differences in allelic and genotypic distribution between BC patients and healthy individuals of genotype (G/G) and (A/G) (p < 0.05) were observed. The frequency of the allele G in the BC group (77%) was significantly higher as compared to control group (61%) (p = 0.01). The association of genotype GG with clinico-pathological features including age, stages of cancer and organ (lung, liver, bones and brain) metastasis (p > 0.05) was assessed. In a MLR analysis, a number of variables including age, weight of an individual, affected lymph nodes, hormonal status (estrogen and progesterone receptor), alcohol consumption and family history associated with the GG genotype (GG:AA, odds ratio (OR) = 1.30, 95% CI [1.06-1.60]) were found to be independent risk factors for BC. Our in-vitro results suggest that genotype GG is possibly increasing the risk of BC in Pakistani cohorts. in-silico analysis finds that CXCL12-CXCR4 is associated with an increased expression of PDZK1, PI3k and

  11. Down-Regulation of CXCL12/CXCR4 Expression Alleviates Ischemia-Reperfusion-Induced Inflammatory Pain via Inhibiting Glial TLR4 Activation in the Spinal Cord.

    PubMed

    Li, Xiao-Qian; Zhang, Zai-Li; Tan, Wen-Fei; Sun, Xi-Jia; Ma, Hong

    2016-01-01

    Toll-like receptor 4 (TLR4) is important for the pathogenesis of inflammatory reactions and the promotion of pain processing after ischemia/reperfusion (IR) in spinal cord. Recently, C-X-C chemokine ligand 12 (CXCL12) and its receptor, C-X-C chemokine receptor 4 (CXCR4), were demonstrated to be simultaneously critical for inflammatory reactions, thereby facilitating glial activation. However, whether CXCL12/CXCR4 expression can contribute to IR-induced inflammatory pain via spinal TLR4 remained unclear. A rat model was established by 8 min of aortic arch occlusion. The effects of CXCL12/CXCR4 expression and TLR4 activation on inflammatory hyperalgesia were investigated by pretreatments with CXCL12-neutralizing antibody, CXCR4 antagonist (AMD3100) and TLR4 antagonist (TAK-242) for 5 consecutive days before surgery. The results indicated that IR induced significant and sustained inflammatory pain, observed as decreases in paw withdrawal threshold (PWT) and paw withdrawal latency (PWL), throughout the post-injury period. The increased levels of TLR4 and proinflammatory chemokine CXCL12, as well as its receptor, CXCR4, were closely correlated with the PWT and PWL trends. Double immunostaining further suggested that TLR4, which is mainly expressed on astrocytes and microglia, was closely co-localized with CXCL12 and CXCR4 in spinal dorsal horn. As expected, intrathecal pretreatment with the TLR4 antagonist, TAK-242 markedly ameliorated pain by inhibiting astrocytic and microglial activation, as shown by decreases in TLR4 immunoreactivity and the percentage of double-labeled cells. These protective effects were likely due in part to the reduced production of the downstream cytokines IL-1β and TNF-α, as well as for the recruitment of CXCL12 and CXCR4. Additionally, intrathecal pretreatment with CXCL12-neutralizing antibody and AMD3100 resulted in similar analgesic and anti-inflammatory effects as those receiving TAK-242 pretreatment. These results suggest that

  12. Phenotypic and morphological properties of germinal center dark zone Cxcl12-expressing reticular cells (CRCs) 1,2

    PubMed Central

    Rodda, Lauren B.; Bannard, Oliver; Ludewig, Burkhard; Nagasawa, Takashi; Cyster, Jason G.

    2015-01-01

    The germinal center (GC) is divided into a dark zone (DZ) and a light zone (LZ). GC B cells must cycle between these zones to achieve efficient antibody affinity maturation. Follicular dendritic cells (FDCs) are well characterized for their role in supporting B cell antigen encounter in primary follicles and in the GC LZ. However, the properties of stromal cells supporting B cells in the DZ are relatively unexplored. Recent work identified a novel stromal population of Cxcl12-expressing reticular cells (CRCs) in murine GC DZs. Here we report that CRCs have diverse morphologies, appearing in ‘open’ and ‘closed’ networks, with variable distribution in lymphoid tissue GCs. CRCs are also present in splenic and peripheral lymph node primary follicles. Real-time two-photon microscopy of Peyer’s patch GCs demonstrates B cells moving in close association with CRC processes. CRCs are gp38+ with low to undetectable expression of FDC markers, but CRC-like cells in the DZ are lineage marked, along with FDCs and FRCs, by CD21-Cre and Ccl19-Cre directed fluorescent reporters. In contrast to FDCs, CRCs do not demonstrate dependence on lymphotoxin or TNF for chemokine expression or network morphology. CRC distribution in the DZ does require CXCR4 signaling, which is necessary for GC B cells to access the DZ and likely to interact with CRC processes. Our findings establish CRCs as a major stromal cell type in the GC DZ and suggest CRCs support critical activities of GC B cells in the DZ niche through Cxcl12 expression and direct cell-cell interactions. PMID:26453751

  13. CXC chemokine CXCL12 tissue expression and circulating levels in peptic ulcer patients with Helicobacter pylori infection.

    PubMed

    Bagheri, Vahid; Hassanshahi, Gholamhossein; Mirzaee, Vahid; Khorramdelazad, Hossein

    2016-09-01

    Helicobacter pylori (H. pylori) infection is among the most prevalent human infections. CXCL12 is a well-known CXC chemokine involved in inflammation and play major roles in angiogenesis. There is currently very limited data on the role of CXCL12 in peptic ulcer disease. Hence, we aimed to explore whether CXCL12 is involved in the pathogenesis of peptic ulcer induced by H. pylori. In this study, we enrolled 102 H. pylori-infected patients, including 51 with active ulcer (GA) and 51 with healing ulcer (GH). We also recruited 50 healthy subjects as control, which did not show any sign or symptoms of chronic inflammatory diseases, infection, or immune-related disorders. Endoscopy was performed to determine the stage of the disease. ELISA was used for detection of H. pylori infection and CXCL12 measurement. We also employed western blotting to detect CXCL12 in ulcerative lesions of H. pylori. Demographic data were also collected by questionnaire. Our results demonstrated that CXCL12 serum levels in GA group (151.8±18.31pg/mL) were significantly higher than those in GH (36.89±6.78pg/mL) and control groups (33.77±9.12pg/mL) (P<0.0001). However, we did not observe a significant difference between GH and control groups. Moreover, overexpression of CXCL12 in gastric lesions of patients in GA group was confirmed by Western blot analysis. According to the result of the present study, it could be concluded that CXCL12 is involved in the pathogenesis and healing of H. pylori-induced peptic ulcer. CXCL12 serum levels may also be used to distinguish between GA and GH phases of the disease.

  14. Loss of Twist1 in the Mesenchymal Compartment Promotes Increased Fibrosis in Experimental Lung Injury by Enhanced Expression of CXCL12

    PubMed Central

    Tan, Jiangning; Tedrow, John R.; Nouraie, Mehdi; Dutta, Justin A.; Miller, David T.; Li, Xiaoyun; Yu, Shibing; Chu, Yanxia; Juan-Guardela, Brenda; Kaminski, Naftali; Ramani, Kritika; Biswas, Partha S.; Zhang, Yingze

    2017-01-01

    Idiopathic pulmonary fibrosis (IPF) is a disease characterized by the accumulation of apoptosis-resistant fibroblasts in the lung. We have previously shown that high expression of the transcription factor Twist1 may explain this prosurvival phenotype in vitro. However, this observation has never been tested in vivo. We found that loss of Twist1 in COL1A2+ cells led to increased fibrosis characterized by very significant accumulation of T cells and bone marrow–derived matrix-producing cells. We found that Twist1-null cells expressed high levels of the T cell chemoattractant CXCL12. In vitro, we found that the loss of Twist1 in IPF lung fibroblasts increased expression of CXCL12 downstream of increased expression of the noncanonical NF-κB transcription factor RelB. Finally, blockade of CXCL12 with AMD3100 attenuated the exaggerated fibrosis observed in Twist1-null mice. Transcriptomic analysis of 134 IPF patients revealed that low expression of Twist1 was characterized by enrichment of T cell pathways. In conclusion, loss of Twist1 in collagen-producing cells led to increased bleomycin-induced pulmonary fibrosis, which is mediated by increased expression of CXCL12. Twist1 expression is associated with dysregulation of T cells in IPF patients. Twist1 may shape the IPF phenotype and regulate inflammation in fibrotic lung injury. PMID:28179498

  15. CXCL12 protects pancreatic β-cells from oxidative stress by a Nrf2-induced increase in catalase expression and activity

    PubMed Central

    DINIĆ, Svetlana; GRDOVIĆ, Nevena; USKOKOVIĆ, Aleksandra; ĐORĐEVIĆ, Miloš; MIHAILOVIĆ, Mirjana; JOVANOVIĆ, Jelena Arambašić; POZNANOVIĆ, Goran; VIDAKOVIĆ, Melita

    2016-01-01

    Due to intrinsically low levels of antioxidant enzyme expression and activity, insulin producing pancreatic β-cells are particularly susceptible to free radical attack. In diabetes mellitus, which is accompanied by high levels of oxidative stress, this feature of β-cells significantly contributes to their damage and dysfunction. In light of the documented pro-survival effect of chemokine C-X-C Ligand 12 (CXCL12) on pancreatic β-cells, we examined its potential role in antioxidant protection. We report that CXCL12 overexpression enhanced the resistance of rat insulinoma (Rin-5F) and primary pancreatic islet cells to hydrogen peroxide (H2O2). CXCL12 lowered the levels of DNA damage and lipid peroxidation and preserved insulin expression. This effect was mediated through an increase in catalase (CAT) activity. By activating downstream p38, Akt and ERK kinases, CXCL12 facilitated Nrf2 nuclear translocation and enhanced its binding to the CAT gene promoter, inducing constitutive CAT expression and activity that was essential for protecting β-cells from H2O2. PMID:27840391

  16. Internal Tandem Duplication Mutations in FLT3 Gene Augment Chemotaxis to Cxcl12 Protein by Blocking the Down-regulation of the Rho-associated Kinase via the Cxcl12/Cxcr4 Signaling Axis*

    PubMed Central

    Onish, Chie; Mori-Kimachi, Satomi; Hirade, Tomohiro; Abe, Mariko; Taketani, Takeshi; Suzumiya, Junji; Sugimoto, Toshitsugu; Yamaguchi, Seiji; Kapur, Reuben; Fukuda, Seiji

    2014-01-01

    Internal tandem duplication mutations in the Flt3 gene (ITD-FLT3) enhance cell migration toward the chemokine Cxcl12, which is highly expressed in the therapy-protective bone marrow niche, providing a potential mechanism underlying the poor prognosis of ITD-FLT3+ acute myeloid leukemia. We aimed to investigate the mechanisms linking ITD-FLT3 to increased cell migration toward Cxcl12. Classification of the expression of Cxcl12-regulated genes in ITD-FLT3+ cells demonstrated that the enhanced migration of ITD-FLT3+ cells toward Cxcl12 was associated with the differential expression of genes downstream of Cxcl12/Cxcr4, which are functionally distinct from those expressed in ITD-FLT3− cells but are independent of the Cxcr4 expression levels. Among these differentially regulated genes, the expression of Rock1 in the ITD-FLT3+ cells that migrated toward Cxcl12 was significantly higher than in ITD-FLT3− cells that migrated toward Cxcl12. In ITD-FLT3− cells, Rock1 expression and Mypt1 phosphorylation were transiently up-regulated but were subsequently down-regulated by Cxcl12. In contrast, the presence of ITD-FLT3 blocked the Cxcl12-induced down-regulation of Rock1 and early Mypt1 dephosphorylation. Likewise, the FLT3 ligand counteracted the Cxcl12-induced down-regulation of Rock1 in ITD-FLT3− cells, which coincided with enhanced cell migration toward Cxcl12. Rock1 antagonists or Rock1 shRNA abolished the enhanced migration of ITD-FLT3+ cells toward Cxcl12. Our findings demonstrate that ITD-FLT3 increases cell migration toward Cxcl12 by antagonizing the down-regulation of Rock1 expression. These findings suggest that the aberrant modulation of Rock1 expression and activity induced by ITD-FLT3 may enhance acute myeloid leukemia cell chemotaxis to the therapy-protective bone marrow niche, where Cxcl12 is abundantly expressed. PMID:25237195

  17. Activation of the A2B adenosine receptor in B16 melanomas induces CXCL12 expression in FAP-positive tumor stromal cells, enhancing tumor progression

    PubMed Central

    Sorrentino, Claudia; Miele, Lucio; Porta, Amalia; Pinto, Aldo; Morello, Silvana

    2016-01-01

    The A2B receptor (A2BR) can mediate adenosine-induced tumor proliferation, immunosuppression and angiogenesis. Targeting the A2BR has proved to be therapeutically effective in some murine tumor models, but the mechanisms of these effects are still incompletely understood. Here, we report that pharmacologic inhibition of A2BR with PSB1115, which inhibits tumor growth, decreased the number of fibroblast activation protein (FAP)-expressing cells in tumors in a mouse model of melanoma. This effect was associated with reduced expression of fibroblast growth factor (FGF)-2. Treatment of melanoma-associated fibroblasts with the A2BR agonist Bay60-6583 enhanced CXCL12 and FGF2 expression. This effect was abrogated by PSB1115. The A2AR agonist CGS21680 did not induce CXCL12 or FGF2 expression in tumor associated fibroblasts. Similar results were obtained under hypoxic conditions in skin-derived fibroblasts, which responded to Bay60-6583 in an A2BR-dependent manner, by stimulating pERK1/2. FGF2 produced by Bay60-6583-treated fibroblasts directly enhanced the proliferation of melanoma cells. This effect could be reversed by PSB1115 or an anti-FGF2 antibody. Interestingly, melanoma growth in mice receiving Bay60-6583 was attenuated by inhibition of the CXCL12/CXCR4 pathway with AMD3100. CXCL12 and its receptor CXCR4 are involved in angiogenesis and immune-suppression. Treatment of mice with AMD3100 reduced the number of CD31+ cells induced by Bay60-6583. Conversely, CXCR4 blockade did not affect the accumulation of tumor-infiltrating MDSCs or Tregs. Together, our data reveal an important role for A2BR in stimulating FGF2 and CXCL12 expression in melanoma-associated fibroblasts. These factors contribute to create a tumor-promoting microenvironment. Our findings support the therapeutic potential of PSB1115 for melanoma. PMID:27590504

  18. Secreted CXCL12 (SDF-1) Forms Dimers under Physiologic Conditions

    PubMed Central

    Ray, Paramita; Lewin, Sarah A.; Mihalko, Laura Anne; Lesher-Perez, Sasha Cai; Takayama, Shuichi; Luker, Kathryn E.; Luker, Gary D.

    2015-01-01

    Chemokine CXCL12 signaling through receptors CXCR4 and CXCR7 has essential functions in development and underlies diseases including cancer, atherosclerosis, and autoimmunity. Chemokines may form homodimers that regulate receptor binding and signaling, but previous studies with synthetic CXCL12 have produced conflicting evidence for homodimerization. We used bioluminescence imaging with Gaussia luciferase fusions to investigate dimerization of CXCL12 secreted from mammalian cells. By column chromatography and Gaussia luciferase complementation, we established that CXCL12 was secreted from mammalian cells as both monomers and dimers. Secreted CXCL12 also formed homodimers in the extracellular space. Monomeric CXCL12 preferentially activated CXCR4 signaling through Gαi and AKT, while dimeric CXCL12 more effectively promoted recruitment of β-arrestin 2 to CXCR4 and chemotaxis of CXCR4-expressing breast cancer cells. We also showed that CXCR7 preferentially sequestered monomeric CXCL12 from the extracellular space and had minimal effects on dimeric CXCL12 in cell-based assays and an orthotopic tumor xenograft model of human breast cancer. These studies establish that CXCL12 secreted from mammalian cells forms homodimers under physiologic conditions. Since monomeric and dimeric CXCL12 have distinct effects on cell signaling and function, our results have important implications for ongoing efforts to target CXCL12 pathways for therapy. PMID:22142194

  19. CXC Chemokine CXCL12 and Its Receptor CXCR4 in Tree Shrews (Tupaia belangeri): Structure, Expression and Function

    PubMed Central

    Meng, Shengke; Zhang, Lichao; Wang, Wenxue; Jiang, Zongmin; Yu, Min; Cui, Qinghua; Li, Meizhang

    2014-01-01

    Chemokines are small secreted proteins functionally involved in the immune system's regulation of lymphocyte migration across numerous mammalian species. Given its growing popularity in immunological models, we investigated the structure and function of chemokine CXCL12 protein in tree shrews. We found that CXCL12 and its receptor CXCR4 in tree shrew had structural similarities to their homologous human proteins. Phylogenetic analysis supports the view that tree shrew is evolutionarily-close to the primates. Our results also showed that the human recombinant CXCL12 protein directly enhanced the migration of tree shrew's lymphocytes in vitro, while AMD3100 enhanced the mobilization of hematopoietic progenitor cells (HPCs) from bone marrow into peripheral blood in tree shrew in vivo. Collectively, these findings suggested that chemokines in tree shrews may play the same or similar roles as those in humans, and that the tree shrew is a viable animal model for studying human immunological diseases. PMID:24858548

  20. CXC chemokine CXCL12 and its receptor CXCR4 in tree shrews (Tupaia belangeri): structure, expression and function.

    PubMed

    Chen, Guiyuan; Wang, Wei; Meng, Shengke; Zhang, Lichao; Wang, Wenxue; Jiang, Zongmin; Yu, Min; Cui, Qinghua; Li, Meizhang

    2014-01-01

    Chemokines are small secreted proteins functionally involved in the immune system's regulation of lymphocyte migration across numerous mammalian species. Given its growing popularity in immunological models, we investigated the structure and function of chemokine CXCL12 protein in tree shrews. We found that CXCL12 and its receptor CXCR4 in tree shrew had structural similarities to their homologous human proteins. Phylogenetic analysis supports the view that tree shrew is evolutionarily-close to the primates. Our results also showed that the human recombinant CXCL12 protein directly enhanced the migration of tree shrew's lymphocytes in vitro, while AMD3100 enhanced the mobilization of hematopoietic progenitor cells (HPCs) from bone marrow into peripheral blood in tree shrew in vivo. Collectively, these findings suggested that chemokines in tree shrews may play the same or similar roles as those in humans, and that the tree shrew is a viable animal model for studying human immunological diseases.

  1. Sdf-1 (CXCL12) induces CD9 expression in stem cells engaged in muscle regeneration.

    PubMed

    Brzoska, Edyta; Kowalski, Kamil; Markowska-Zagrajek, Agnieszka; Kowalewska, Magdalena; Archacki, Rafał; Plaskota, Izabela; Stremińska, Władysława; Jańczyk-Ilach, Katarzyna; Ciemerych, Maria A

    2015-03-24

    Understanding the mechanism of stem cell mobilization into injured skeletal muscles is a prerequisite step for the development of muscle disease therapies. Many of the currently studied stem cell types present myogenic potential; however, when introduced either into the blood stream or directly into the tissue, they are not able to efficiently engraft injured muscle. For this reason their use in therapy is still limited. Previously, we have shown that stromal-derived factor-1 (Sdf-1) caused the mobilization of endogenous (not transplanted) stem cells into injured skeletal muscle improving regeneration. Here, we demonstrate that the beneficial effect of Sdf-1 relies on the upregulation of the tetraspanin CD9 expression in stem cells. The expression pattern of adhesion proteins, including CD9, was analysed after Sdf-1 treatment during regeneration of rat skeletal muscles and mouse Pax7-/- skeletal muscles, that are characterized by the decreased number of satellite cells. Next, we examined the changes in CD9 level in satellite cells-derived myoblasts, bone marrow-derived mesenchymal stem cells, and embryonic stem cells after Sdf-1 treatment or silencing expression of CXCR4 and CXCR7. Finally, we examined the potential of stem cells to fuse with myoblasts after Sdf-1 treatment. In vivo analyses of Pax7-/- mice strongly suggest that Sdf-1-mediates increase in CD9 levels also in mobilized stem cells. In the absence of CXCR4 receptor the effect of Sdf-1 on CD9 expression is blocked. Next, in vitro studies show that Sdf-1 increases the level of CD9 not only in satellite cell-derived myoblasts but also in bone marrow derived mesenchymal stem cells, as well as embryonic stem cells. Importantly, the Sdf-1 treated cells migrate and fuse with myoblasts more effectively. We suggest that Sdf-1 binding CXCR4 receptor improves skeletal muscle regeneration by upregulating expression of CD9 and thus, impacting at stem cells mobilization to the injured muscles.

  2. The contribution of CXCL12-expressing radial glia cells to neuro-vascular patterning during human cerebral cortex development

    PubMed Central

    Errede, Mariella; Girolamo, Francesco; Rizzi, Marco; Bertossi, Mirella; Roncali, Luisa; Virgintino, Daniela

    2014-01-01

    This study was conducted on human developing brain by laser confocal and transmission electron microscopy (TEM) to make a detailed analysis of important features of blood-brain barrier (BBB) microvessels and possible control mechanisms of vessel growth and differentiation during cerebral cortex vascularization. The BBB status of cortex microvessels was examined at a defined stage of cortex development, at the end of neuroblast waves of migration, and before cortex lamination, with BBB-endothelial cell markers, namely tight junction (TJ) proteins (occludin and claudin-5) and influx and efflux transporters (Glut-1 and P-glycoprotein), the latter supporting evidence for functional effectiveness of the fetal BBB. According to the well-known roles of astroglia cells on microvessel growth and differentiation, the early composition of astroglia/endothelial cell relationships was analyzed by detecting the appropriate astroglia, endothelial, and pericyte markers. GFAP, chemokine CXCL12, and connexin 43 (Cx43) were utilized as markers of radial glia cells, CD105 (endoglin) as a marker of angiogenically activated endothelial cells (ECs), and proteoglycan NG2 as a marker of immature pericytes. Immunolabeling for CXCL12 showed the highest level of the ligand in radial glial (RG) fibers in contact with the growing cortex microvessels. These specialized contacts, recognizable on both perforating radial vessels and growing collaterals, appeared as CXCL12-reactive en passant, symmetrical and asymmetrical, vessel-specific RG fiber swellings. At the highest confocal resolution, these RG varicosities showed a CXCL12-reactive dot-like content whose microvesicular nature was confirmed by ultrastructural observations. A further analysis of RG varicosities reveals colocalization of CXCL12 with Cx43, which is possibly implicated in vessel-specific chemokine signaling. PMID:25360079

  3. Intra-Bone Marrow Transplantation of Endosteal Bone Marrow Cells Facilitates Allogeneic Hematopoietic and Stromal Cells Engraftment Dependent on Early Expression of CXCL-12.

    PubMed

    Chen, Chen; Su, Yingjun; Chen, Jianwu; Zhang, Dongliang; Song, Yajuan; Guo, Shuzhong

    2015-09-16

    Hematopoietic stem cell transplantation (HSCT) has been considered as an effective approach at inducing allogeneic hematopoietic reconstitution and immune tolerance. However, it remains critical to find the optimal HSCT delivery method and robust sources of hematopoietic stem cells (HSCs). We introduced a new method by infusing allogeneic endosteal bone marrow cells (BMCs) harvested from long bones endosteum through intra-bone marrow transplantation (IBBMT) into irradiated mice. Recipient mice that were transplanted with central BMCs or through intravenous bone marrow transplantation (IVBMT) were used as controls (n=6 per group). We compared the new method with each control group for allogeneic HSCs homing pattern, peripheral blood chimerism level, skin allograft survival time, and donor stromal cell percentage in recipient BM. AMD3100 was injected to determine whether chemokine stromal cell-derived factor-1 (CXCL-12) was critical for the new method. More allogeneic HSCs homed into spleen and bone marrow for the new method as compared to each control group. IBBMT of endosteal BMCs led to a higher peripheral blood chimerism and skin allograft survival. At 18 weeks, donor stromal cell percentage in recipient BMCs was higher for the new method than in each control group. By AMD3100 blockade at day 1, peripheral blood chimerism level and donor stromal cell percentage were significantly reduced as compared to the control group without AMD3100 blockade. Our study suggests that IBBMT of endosteal BMCs is an effective approach for HSCT in inducing allogeneic hematopoietic reconstitution. The advantage is dependent upon the early expression of CXCL-12 after bone marrow transplantation.

  4. A Comprehensive Analysis of CXCL12 Isoforms in Breast Cancer(1,2.)

    PubMed

    Zhao, Shuang; Chang, S Laura; Linderman, Jennifer J; Feng, Felix Y; Luker, Gary D

    2014-05-13

    CXCL12-CXCR4-CXCR7 signaling promotes tumor growth and metastasis in breast cancer. Alternative splicing of CXCL12 produces isoforms with distinct structural and biochemical properties, but little is known about isoform-specific differences in breast cancer subtypes and patient outcomes. We investigated global expression profiles of the six CXCL12 isoforms, CXCR4, and CXCR7 in The Cancer Genome Atlas breast cancer cohort using next-generation RNA sequencing in 948 breast cancer and benign samples and seven breast cancer cell lines. We compared expression levels with several clinical parameters, as well as metastasis, recurrence, and overall survival (OS). CXCL12-α, -β, and -γ are highly co-expressed, with low expression correlating with more aggressive subtypes, higher stage disease, and worse clinical outcomes. CXCL12-δ did not correlate with other isoforms but was prognostic for OS and showed the same trend for metastasis and recurrence-free survival. Effects of CXCL12-δ remained independently prognostic when taking into account expression of CXCL12,CXCR4, and CXCR7. These results were also reflected when comparing CXCL12-α, -β, and -γ in breast cancer cell lines. We summarized expression of all CXCL12 isoforms in an important chemokine signaling pathway in breast cancer in a large clinical cohort and common breast cancer cell lines, establishing differences among isoforms in multiple clinical, pathologic, and molecular subgroups. We identified for the first time the clinical importance of a previously unstudied isoform, CXCL12-δ.

  5. Central SDF-1/CXCL12 expression and its cardiovascular and sympathetic effects: the role of angiotensin II, TNF-α, and MAP kinase signaling.

    PubMed

    Wei, Shun-Guang; Zhang, Zhi-Hua; Yu, Yang; Felder, Robert B

    2014-12-01

    The chemokine stromal cell-derived factor-1 (SDF-1/CXCL12) and its receptors are expressed by neurons and glial cells in cardiovascular autonomic regions of the brain, including the hypothalamic paraventricular nucleus (PVN), and contribute to neurohumoral excitation in rats with ischemia-induced heart failure. The present study examined factors regulating the expression of SDF-1 in the PVN and mechanisms mediating its sympatho-excitatory effects. In urethane anesthetized rats, a 4-h intracerebroventricular (ICV) infusion of angiotensin II (ANG II) or tumor necrosis factor-α (TNF-α) in doses that increase mean blood pressure (MBP) and sympathetic drive increased the expression of SDF-1 in PVN. ICV administration of SDF-1 increased the phosphorylation of p44/42 mitogen-activated protein kinase (MAPK), JNK, and p38 MAPK in PVN, along with MBP, heart rate (HR), and renal sympathetic nerve activity (RSNA), but did not affect total p44/42 MAPK, JNK, and p38 MAPK levels. ICV pretreatment with the selective p44/42 MAPK inhibitor PD98059 prevented the SDF-1-induced increases in MBP, HR, and RSNA; ICV pretreatment with the selective JNK and p38 MAPK inhibitors attenuated but did not block these SDF-1-induced excitatory responses. ICV PD98059 also prevented the sympatho-excitatory response to bilateral PVN microinjections of SDF-1. ICV pretreatment with SDF-1 short-hairpin RNA significantly reduced ANG II- and TNF-α-induced phosphorylation of p44/42 MAPK in PVN. These findings identify TNF-α and ANG II as drivers of SDF-1 expression in PVN and suggest that the full expression of their cardiovascular and sympathetic effects depends upon SDF-1-mediated activation of p44/42 MAPK signaling.

  6. Central SDF-1/CXCL12 expression and its cardiovascular and sympathetic effects: the role of angiotensin II, TNF-α, and MAP kinase signaling

    PubMed Central

    Wei, Shun-Guang; Zhang, Zhi-Hua; Yu, Yang

    2014-01-01

    The chemokine stromal cell-derived factor-1 (SDF-1/CXCL12) and its receptors are expressed by neurons and glial cells in cardiovascular autonomic regions of the brain, including the hypothalamic paraventricular nucleus (PVN), and contribute to neurohumoral excitation in rats with ischemia-induced heart failure. The present study examined factors regulating the expression of SDF-1 in the PVN and mechanisms mediating its sympatho-excitatory effects. In urethane anesthetized rats, a 4-h intracerebroventricular (ICV) infusion of angiotensin II (ANG II) or tumor necrosis factor-α (TNF-α) in doses that increase mean blood pressure (MBP) and sympathetic drive increased the expression of SDF-1 in PVN. ICV administration of SDF-1 increased the phosphorylation of p44/42 mitogen-activated protein kinase (MAPK), JNK, and p38 MAPK in PVN, along with MBP, heart rate (HR), and renal sympathetic nerve activity (RSNA), but did not affect total p44/42 MAPK, JNK, and p38 MAPK levels. ICV pretreatment with the selective p44/42 MAPK inhibitor PD98059 prevented the SDF-1-induced increases in MBP, HR, and RSNA; ICV pretreatment with the selective JNK and p38 MAPK inhibitors attenuated but did not block these SDF-1-induced excitatory responses. ICV PD98059 also prevented the sympatho-excitatory response to bilateral PVN microinjections of SDF-1. ICV pretreatment with SDF-1 short-hairpin RNA significantly reduced ANG II- and TNF-α-induced phosphorylation of p44/42 MAPK in PVN. These findings identify TNF-α and ANG II as drivers of SDF-1 expression in PVN and suggest that the full expression of their cardiovascular and sympathetic effects depends upon SDF-1-mediated activation of p44/42 MAPK signaling. PMID:25260613

  7. Multiple functions of CXCL12 in a syngeneic model of breast cancer

    PubMed Central

    2010-01-01

    Background A growing body of work implicates chemokines, in particular CXCL12 and its receptors, in the progression and site-specific metastasis of various cancers, including breast cancer. Various agents have been used to block the CXCL12-CXCR4 interaction as a means of inhibiting cancer metastasis. However, as a potent chemotactic factor for leukocytes, CXCL12 also has the potential to enhance anti-cancer immunity. To further elucidate its role in breast cancer progression, CXCL12 and its antagonist CXCL12(P2G) were overexpressed in the syngeneic 4T1.2 mouse model of breast carcinoma. Results While expression of CXCL12(P2G) significantly inhibited metastasis, expression of wild-type CXCL12 potently inhibited both metastasis and primary tumor growth. The effects of wild-type CXCL12 were attributed to an immune response characterized by the induction of CD8+ T cell activity, enhanced cell-mediated cytotoxicity, increased numbers of CD11c+ cells in the tumor-draining lymph nodes and reduced accumulation of myeloid-derived suppressor cells in the spleen. Conclusions This study highlights the need to consider carefully therapeutic strategies that block CXCL12 signaling. Therapies that boost CXCL12 levels at the primary tumor site may prove more effective in the treatment of metastatic breast cancer. PMID:20849618

  8. CXCL12/SDF-1 facilitates optic nerve regeneration.

    PubMed

    Heskamp, Annemarie; Leibinger, Marco; Andreadaki, Anastasia; Gobrecht, Philipp; Diekmann, Heike; Fischer, Dietmar

    2013-07-01

    Mature retinal ganglion cells (RGCs) do not normally regenerate injured axons, but undergo apoptosis soon after axotomy. Besides the insufficient intrinsic capability of mature neurons to regrow axons inhibitory molecules located in myelin of the central nervous system as well as the glial scar forming at the site of injury strongly limit axon regeneration. Nevertheless, RGCs can be transformed into a regenerative state upon inflammatory stimulation (IS), enabling these neurons to grow axons into the injured optic nerve. The outcome of IS stimulated regeneration is, however, still limited by the inhibitory extracellular environment. Here, we report that the chemokine CXCL12/SDF-1 moderately stimulates neurite growth of mature RGCs on laminin in culture and, in contrast to CNTF, exerts potent disinhibitory effects towards myelin. Consistently, co-treatment of RGCs with CXCL12 facilitated CNTF stimulated neurite growth of RGCs on myelin. Mature RGCs express CXCR4, the cognate CXCL12 receptor. Furthermore, the neurite growth promoting and disinhibitory effects of CXCL12 were abrogated by a specific CXCR4 antagonist and by inhibition of the PI3K/AKT/mTOR-, but not the JAK/STAT3-pathway. In vivo, intravitreal application of CXCL12 sustained mTOR activity in RGCs upon optic nerve injury and moderately stimulated axon regeneration in the optic nerve without affecting the survival of RGCs. Importantly, intravitreal application of CXCL12 also significantly increased IS triggered axon regeneration in vivo. These data suggest that the disinhibitory effect of CXCL12 towards myelin may be a useful feature to facilitate optic nerve regeneration, particularly in combination with other axon growth stimulatory treatments.

  9. PARP-1 and YY1 are important novel regulators of CXCL12 gene transcription in rat pancreatic beta cells.

    PubMed

    Marković, Jelena; Grdović, Nevena; Dinić, Svetlana; Karan-Djurašević, Teodora; Uskoković, Aleksandra; Arambašić, Jelena; Mihailović, Mirjana; Pavlović, Sonja; Poznanović, Goran; Vidaković, Melita

    2013-01-01

    Despite significant progress, the molecular mechanisms responsible for pancreatic beta cell depletion and development of diabetes remain poorly defined. At present, there is no preventive measure against diabetes. The positive impact of CXCL12 expression on the pancreatic beta cell prosurvival phenotype initiated this study. Our aim was to provide novel insight into the regulation of rat CXCL12 gene (Cxcl12) transcription. The roles of poly(ADP-ribose) polymerase-1 (PARP-1) and transcription factor Yin Yang 1 (YY1) in Cxcl12 transcription were studied by examining their in vitro and in vivo binding affinities for the Cxcl12 promoter in a pancreatic beta cell line by the electrophoretic mobility shift assay and chromatin immunoprecipitation. The regulatory activities of PARP-1 and YY1 were assessed in transfection experiments using a reporter vector with a Cxcl12 promoter sequence driving luciferase gene expression. Experimental evidence for PARP-1 and YY1 revealed their trans-acting potential, wherein PARP-1 displayed an inhibitory, and YY1 a strong activating effect on Cxcl12 transcription. Streptozotocin (STZ)-induced general toxicity in pancreatic beta cells was followed by changes in Cxcl12 promoter regulation. PARP-1 binding to the Cxcl12 promoter during basal and in STZ-compromised conditions led us to conclude that PARP-1 regulates constitutive Cxcl12 expression. During the early stage of oxidative stress, YY1 exhibited less affinity toward the Cxcl12 promoter while PARP-1 displayed strong binding. These interactions were accompanied by Cxcl12 downregulation. In the later stages of oxidative stress and intensive pancreatic beta cell injury, YY1 was highly expressed and firmly bound to Cxcl12 promoter in contrast to PARP-1. These interactions resulted in higher Cxcl12 expression. The observed ability of PARP-1 to downregulate, and of YY1 to upregulate Cxcl12 promoter activity anticipates corresponding effects in the natural context where the functional

  10. Interactions between CXCR4 and CXCL12 promote cell migration and invasion of canine hemangiosarcoma.

    PubMed

    Im, K S; Graef, A J; Breen, M; Lindblad-Toh, K; Modiano, J F; Kim, J-H

    2017-06-01

    The CXCR4/CXCL12 axis plays an important role in cell locomotion and metastasis in many cancers. In this study, we hypothesized that the CXCR4/CXCL12 axis promotes migration and invasion of canine hemangiosarcoma (HSA) cells. Transcriptomic analysis across 12 HSA cell lines and 58 HSA whole tumour tissues identified heterogeneous expression of CXCR4 and CXCL12, which was associated with cell movement. In vitro, CXCL12 promoted calcium mobilization, cell migration and invasion that were directly proportional to surface expression of CXCR4; furthermore, these responses proved sensitive to the CXCR4 antagonist, AMD3100, in HSA cell lines. These results indicate that CXCL12 potentiates migration and invasion of canine HSA cells through CXCR4 signalling. The direct relationship between these responses in HSA cells suggests that the CXCR4/CXCL12 axis contributes to HSA progression. © 2015 John Wiley & Sons Ltd.

  11. Targeting CXCL12/CXCR4 Axis in Tumor Immunotherapy.

    PubMed

    Zhou, Weiqiang; Guo, Shanchun; Liu, Mingli; Burow, Matthew; Wang, Guangdi

    2017-08-29

    Chemokines, which have chemotactic abilities, are comprised of over 50 family members. Through binding to the 7-transmembrane domain of G-protein-coupled receptors (GPCR), they function in immune cells by trafficking and regulating cell prolif¬eration, differentiation, activation, and migration, homing under both physiologic and pathologic conditions. The alpha-chemokine receptor CXCR4 for the alpha-chemokine stromal cell-derived-factor-1 (SDF-1) is most widely expressed by tumors. CXCL12/CXCR4 axis is a major culprit for human tumor because of its crucial role in tumor initiation and progression by activating a number of signaling pathways, such as ERK1/2, ras, PLC/ MAPK, p38 MAPK, and SAPK/ JNK, as well as regulating cancer stem cells. CXCL12/CXCR4 antagonists have been developed, which have shown promising results in both in vitro and in vivo anti-cancer activity in several tumor types. This review provides an evaluation of CXCL12/CXCR4 as a potential therapeutic target for human tumors; it also focuses on the synergistic effects of inhibition of CXCL12/CXCR4 axis and immunotherapy as well as chemotherapy. Together, CXCL12/CXCR4 axis can be a potential therapeutic target for tumors and used with immunotherapy for additive effects. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  12. Preconditioning-induced CXCL12 upregulation minimizes leukocyte infiltration after stroke in ischemia-tolerant mice.

    PubMed

    Selvaraj, Uma Maheswari; Ortega, Sterling B; Hu, Ruilong; Gilchrist, Robert; Kong, Xiangmei; Partin, Alexander; Plautz, Erik J; Klein, Robyn S; Gidday, Jeffrey M; Stowe, Ann M

    2017-03-01

    Repetitive hypoxic preconditioning creates long-lasting, endogenous protection in a mouse model of stroke, characterized by reductions in leukocyte-endothelial adherence, inflammation, and infarct volumes. The constitutively expressed chemokine CXCL12 can be upregulated by hypoxia and limits leukocyte entry into brain parenchyma during central nervous system inflammatory autoimmune disease. We therefore hypothesized that the sustained tolerance to stroke induced by repetitive hypoxic preconditioning is mediated, in part, by long-term CXCL12 upregulation at the blood-brain barrier (BBB). In male Swiss Webster mice, repetitive hypoxic preconditioning elevated cortical CXCL12 protein levels, and the number of cortical CXCL12+ microvessels, for at least two weeks after the last hypoxic exposure. Repetitive hypoxic preconditioning-treated mice maintained more CXCL12-positive vessels than untreated controls following transient focal stroke, despite cortical decreases in CXCL12 mRNA and protein. Continuous administration of the CXCL12 receptor (CXCR4) antagonist AMD3100 for two weeks following repetitive hypoxic preconditioning countered the increase in CXCL12-positive microvessels, both prior to and following stroke. AMD3100 blocked the protective post-stroke reductions in leukocyte diapedesis, including macrophages and NK cells, and blocked the protective effect of repetitive hypoxic preconditioning on lesion volume, but had no effect on blood-brain barrier dysfunction. These data suggest that CXCL12 upregulation prior to stroke onset, and its actions following stroke, contribute to the endogenous, anti-inflammatory phenotype induced by repetitive hypoxic preconditioning.

  13. c/EBPbeta is a major regulatory element driving transcriptional activation of the CXCL12 promoter.

    PubMed

    Calonge, E; Alonso-Lobo, J M; Escandón, C; González, N; Bermejo, M; Santiago, B; Mestre, L; Pablos, J L; Caruz, A; Alcamí, J

    2010-02-26

    CXCL12 is considered a constitutively expressed chemokine with homeostatic functions. However, induction of CXCL12 expression and its potential role in several pathologic conditions have been reported, suggesting that CXCL12 gene expression can be induced by different stimuli. To elucidate the molecular mechanisms involved in the regulation of CXCL12 gene expression, we aim to define the molecular factors that operate at the transcriptional level. Basal, constitutive expression of CXCL12 was dependent on basic helix-loop-helix factors. Transcriptional up-regulation of the CXCL12 gene was induced by cellular confluence or inflammatory stimuli such as interleukin-1 and interleukin-6, in a CCAAT/enhancer binding protein beta (c/EBPbeta)-dependent manner. Chromatin immunoprecipitation assays confirmed c/EBPbeta binding to a specific response element located at -1171 of the promoter region of CXCL12. Our data show that c/EBPbeta is a major regulatory element driving transcription of the CXCL12 gene in response to cytokines and cell confluence.

  14. Association of CXCL12 gene promoter methylation with periodontitis in patients with diabetes mellitus type 2.

    PubMed

    Grdović, Nevena; Rajić, Jovana; Petrović, Sanja Matić; Dinić, Svetlana; Uskoković, Aleksandra; Mihailović, Mirjana; Jovanović, Jelena Arambašić; Tolić, Anja; Pucar, Ana; Milašin, Jelena; Vidaković, Melita

    2016-12-01

    CXCL12 is widely expressed, constitutive chemokine involved in tissue repair and regeneration, while the extent of its expression is important in various chronic inflammatory conditions. Involvement of DNA methylation in CXCL12 gene suppression (CXCL12) has been shown in malignancy and some autoimmune diseases. The aim of this study was to investigate whether the alterations in DNA methylation of CXCL12 are also involved in progression of periodontitis in combination with diabetes, as these chronic inflammatory conditions are strongly interrelated. Study included 72 subjects divided in three groups: healthy control (C, n=21), periodontitis (P, n=29) and diabetes/periodontitis group (D/P, n=22). DNA extracted from epithelial cells obtained by sterile cotton swabs from buccal mucosa was subjected to methylation specific polymerase chain reaction (MSP) to obtain DNA methylation pattern of CXCL12 promoter. CXCL12 promoter was predominantly unmethylated in all groups. However, increase in the frequency of the methylated form and increase in percent of methylation of CXCL12 promoter in periodontitis and diabetes/periodontitis group compared to control group were found, although without statistical significance. However, statistically significant increase in Tm of MSP products in diabetes/periodontitis group was observed. Correlation analysis revealed statistically significant relationship between the extent of DNA methylation of the CXCL12 promoter and periodontal parameters, as well as between DNA methylation of CXCL12 and glycosylated hemoglobin. Presented results suggest that chronic inflammation contributes to the change of CXCL12 DNA methylation in buccal cells and that DNA methylation profile of CXCL12 promoter plays important role in development and progression of periodontal disease. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. CXC receptor-4 mRNA silencing abrogates CXCL12-induced migration of colorectal cancer cells

    PubMed Central

    2011-01-01

    Background Interactions between CXCR4 and its ligand CXCL12 have been shown to be involved in cancer progression in colorectal cancer (CRC). We performed a comparative CXCL12/CXCR4 expression analysis and assessed the effect of external CXCL12 stimulation on migration of CRC cells without and with CXCR4 inhibition. Methods Expression of CXCL12/CXCR4 was assessed by quantitative real-time PCR, ELISA and immunohistochemistry in resection specimens of 50 CRC patients as well as in the corresponding normal tissues and in three human CRC cell lines with different metastatic potential (Caco-2, SW480 and HT-29). Migration assays were performed after stimulation with CXCL12 and CXCR4 was inhibited by siRNA and neutralizing antibodies. Results In CRC tissues CXCL12 was significantly down-regulated and CXCR4 was significantly up-regulated compared to the corresponding normal tissues. In cell lines CXCR4 was predominantly expressed in SW480 and less pronounced in HT-29 cells. CXCL12 was only detectable in Caco-2 cells. CXCL12 stimulation had no impact on Caco-2 cells but significantly increased migration of CXCR4 bearing SW480 and HT-29 cells. This effect was significantly abrogated by neutralizing anti-CXCR4 antibody as well as by CXCR4 siRNAs (P < 0.05). Conclusions CXCR4 expression was up-regulated in CRC and CXCL12 stimulation increased migration in CXCR4 bearing cell lines. Migration was inhibited by both neutralizing CXCR4 antibodies and CXCR4 siRNAs. Thus, the expression and functionality of CXCR4 might be associated with the metastatic potential of CRC cells and CXCL12/CXCR4 interactions might therefore constitute a promising target for specific treatment interventions. PMID:21349176

  16. Microfluidic source-sink model reveals effects of biophysically distinct CXCL12 isoforms in breast cancer chemotaxis.

    PubMed

    Cavnar, S P; Ray, P; Moudgil, P; Chang, S L; Luker, K E; Linderman, J J; Takayama, S; Luker, G D

    2014-05-01

    Chemokines critically regulate chemotaxis in normal and pathologic states, but there is limited understanding of how multicellular interactions generate gradients needed for cell migration. Previous studies of chemotaxis of CXCR4+ cells toward chemokine CXCL12 suggest the requirement of cells expressing scavenger receptor CXCR7 in a source-sink system. We leveraged an established microfluidic device to discover that chemotaxis of CXCR4 cells toward distinct isoforms of CXCL12 required CXCR7 scavenging only under conditions with higher than optimal levels of CXCL12. Chemotaxis toward CXCL12-β and -γ isoforms, which have greater binding to extracellular molecules and have been largely overlooked, was less dependent on CXCR7 than the more commonly studied CXCL12-α. Chemotaxis of CXCR4+ cells toward even low levels of CXCL12-γ and CXCL12-β still occurred during treatment with a FDA-approved inhibitor of CXCR4. We also detected CXCL12-γ only in breast cancers from patients with advanced disease. Physiological gradient formation within the device facilitated interrogation of key differences in chemotaxis among CXCL12 isoforms and suggests CXCL12-γ as a biomarker for metastatic cancer.

  17. CXCL12 in early mesenchymal progenitors is required for haematopoietic stem-cell maintenance.

    PubMed

    Greenbaum, Adam; Hsu, Yen-Michael S; Day, Ryan B; Schuettpelz, Laura G; Christopher, Matthew J; Borgerding, Joshua N; Nagasawa, Takashi; Link, Daniel C

    2013-03-14

    Haematopoietic stem cells (HSCs) primarily reside in the bone marrow where signals generated by stromal cells regulate their self-renewal, proliferation and trafficking. Endosteal osteoblasts and perivascular stromal cells including endothelial cells, CXCL12-abundant reticular cells, leptin-receptor-positive stromal cells, and nestin-green fluorescent protein (GFP)-positive mesenchymal progenitors have all been implicated in HSC maintenance. However, it is unclear whether specific haematopoietic progenitor cell (HPC) subsets reside in distinct niches defined by the surrounding stromal cells and the regulatory molecules they produce. CXCL12 (chemokine (C-X-C motif) ligand 12) regulates both HSCs and lymphoid progenitors and is expressed by all of these stromal cell populations. Here we selectively deleted Cxcl12 from candidate niche stromal cell populations and characterized the effect on HPCs. Deletion of Cxcl12 from mineralizing osteoblasts has no effect on HSCs or lymphoid progenitors. Deletion of Cxcl12 from osterix-expressing stromal cells, which include CXCL12-abundant reticular cells and osteoblasts, results in constitutive HPC mobilization and a loss of B-lymphoid progenitors, but HSC function is normal. Cxcl12 deletion from endothelial cells results in a modest loss of long-term repopulating activity. Strikingly, deletion of Cxcl12 from nestin-negative mesenchymal progenitors using Prx1-cre (Prx1 also known as Prrx1) is associated with a marked loss of HSCs, long-term repopulating activity, HSC quiescence and common lymphoid progenitors. These data suggest that osterix-expressing stromal cells comprise a distinct niche that supports B-lymphoid progenitors and retains HPCs in the bone marrow, and that expression of CXCL12 from stromal cells in the perivascular region, including endothelial cells and mesenchymal progenitors, supports HSCs.

  18. Histone hypoacetylation contributes to CXCL12 downregulation in colon cancer: impact on tumor growth and cell migration.

    PubMed

    Romain, Benoît; Benbrika-Nehmar, Radhia; Marisa, Laetitia; Legrain, Michèle; Lobstein, Viviane; Oravecz, Attila; Poidevin, Laetitia; Bour, Cyril; Freund, Jean-Noël; Duluc, Isabelle; Guenot, Dominique; Pencreach, Erwan

    2017-06-13

    CXCL12 has been shown to be involved in colon cancer metastasis, but its expression level and molecular mechanisms regulating its expression remain controversial. We thus evaluated CXCL12 expression in a large cohort of colon adenomas and carcinomas, investigated for an epigenetic mechanism controlling its expression and evaluated the impact of CXCL12 levels on cell migration and tumor growth. CXCL12 expression was measured in human colon adenomas and carcinomas with transcriptome array and RT-qPCR. The promoter methylation was analyzed with whole-genome DNA methylation chips and protein expression by immunohistochemistry. We confirm a reduced expression of CXCL12 in 75% of MSS carcinomas and show that the decrease is an early event as already present in adenomas. The methylome analysis shows that the CXCL12 promoter is methylated in only 30% of microsatellite-stable tumors. In vitro, treatments with HDAC inhibitors, butyrate and valproate restored CXCL12 expression in three colon cell lines, increased acetylation of histone H3 within the CXCL12 promoter and inhibited cell migration. In vivo, valproate diminished (65%) the number of intestinal tumors in APC mutant mice, slowed down xenograft tumor growth concomitant to restored CXCL12 expression. Finally we identified loss of PCAF expression in tumor samples and showed that forced expression of PCAF in colon cancer cell lines restored CXCL12 expression. Thus, reduced PCAF expression may participate to CXCL12 promoter hypoacetylation and its subsequent loss of expression. Our study is of potential clinical interest because agents that promote or maintain histone acetylation through HDAC inhibition and/or HAT stimulation, may help to lower colon adenoma/carcinoma incidence, especially in high-risk families, or could be included in therapeutic protocols to treat advanced colon cancer.

  19. CXCL12-CXCR4 axis promotes the natural selection of breast cancer cell metastasis.

    PubMed

    Sun, Yanan; Mao, Xiaoyun; Fan, Chuifeng; Liu, Chong; Guo, Ayao; Guan, Shu; Jin, Quanxiu; Li, Bo; Yao, Fan; Jin, Feng

    2014-08-01

    CXCR4 and its ligand CXCL12 can promote the proliferation, survival, and invasion of cancer cells. They have been shown to play an important role in regulating metastasis of breast cancer to specific organs. High CXCR4 expression was also correlated to poor clinical outcome. Previous study also showed that tumor cells express a high level of CXCR4 and that tumor metastasis target tissues (lung, liver, and bone) express high levels of the ligand CXCL12, allowing tumor cells to directionally migrate to target organs via a CXCL12-CXCR4 chemotactic gradient. However, the exact mechanisms of how CXCR4 and CXCL12 enhance metastasis and/or tumor growth and their full implications on breast cancer progression are unknown. Yet it is likely that chemokine receptor signaling may provide more than just a migrational advantage by also helping the metastasized cells establish and survive in secondary environments. In this study, we investigated CXCR4 and CXCL12 expression in breast cancer and analyzed its association with clinicopathological factors by immunohistochemistry first. Then, we detected the mRNA and protein expression of CXCR4 and CXCL12 in breast cancer cell lines by Western blot and RT-PCR. The MDA-MB-231 has CXCR4 expression and very weak CXCL12 expression. So, we constructed the functional CXCL12 expression in MDA-MB-231 using a gene transfection technique. Further experiments were conducted to evaluate the effect of CXCL12 transfection on the biological behaviors of MDA-MB-231. The cell proliferation of MDA-MB-231-CXCL12 was accessed by MTT assay; the apoptosis was analyzed by an AnnexinV-FITC/propidium iodide double staining of flow cytometry method; and the cell invasive ability was examined by Matrigel invasion assay. Immunohistochemical analysis showed the co-expression of CXCR4 and CXCL12 correlated with lymph node metastasis and TNM stage (p < 0.01). It suggested that the chemokine CXCL12 and its sole ligand CXCR4 play important role in the malignance

  20. mTORC2 MEDIATES CXCL12-INDUCED ANGIOGENESIS

    PubMed Central

    Ziegler, Mary E.; Hatch, Michaela M.S.; Wu, Nan; Muawad, Steven A.; Hughes, Christopher C.W.

    2016-01-01

    The chemokine CXCL12, through its receptor CXCR4, positively regulates angiogenesis by promoting endothelial cell (EC) migration and tube formation. However, the relevant downstream signaling pathways in EC have not been defined. Similarly, the upstream activators of mTORC2 signaling in EC are also poorly defined. Here we demonstrate for the first time that CXCL12 regulation of angiogenesis requires mTORC2 but not mTORC1. We find that CXCR4 signaling activates mTORC2 as indicated by phosphorylation of serine 473 on Akt, and does so through a G-protein- and PI3K-dependent pathway. Significantly, independent disruption of the mTOR complexes by drugs or multiple independent siRNAs reveals that mTORC2, but not mTORC1, is required for microvascular sprouting in a 3D in vitro angiogenesis model. Importantly, in a mouse model both tumor angiogenesis and tumor volume are significantly reduced only when mTORC2 is inhibited. Finally, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), which is a key regulator of glycolytic flux, is required for microvascular sprouting in vitro, and its expression is reduced in vivo when mTORC2 is targeted. Taken together, these findings identify mTORC2 as a critical signaling nexus downstream of CXCL12/CXCR4 that represents a potential link between mTORC2, metabolic regulation and angiogenesis. PMID:27106789

  1. Evidence for the involvement of the CXCL12 system in the adaptation of skeletal muscles to physical exercise.

    PubMed

    Puchert, Malte; Adams, Volker; Linke, Axel; Engele, Jürgen

    2016-09-01

    The chemokine CXCL12 and its primary receptor, CXCR4, not only promote developmental myogenesis, but also muscle regeneration. CXCL12 chemoattracts CXCR4-positive satellite cells/blood-borne progenitors to the injured muscle, promotes myoblast fusion, partially with existing myofibers, and induces angiogenesis in regenerating muscles. Interestingly, the mechanisms underlying muscle regeneration are in part identical to those involved in muscular adaptation to intensive physical exercise. These similarities now prompted us to determine whether physical exercise would impact the CXCL12 system in skeletal muscle. We found that CXCL12 and CXCR4 are upregulated in the gastrocnemius muscle of rats that underwent a four-week period of constrained daily running exercise on a treadmill. Double-staining experiments confirmed that CXCL12 and CXCR4 are predominantly expressed in MyHC-positive muscle fibers. Moreover, these training-dependent increases in CXCL12 and CXCR4 expression also occurred in rats with surgical coronary artery occlusion, implying that the muscular CXCL12 system is still active in skeletal myopathy resulting from chronic heart failure. Expression of the second CXCL12 receptor, CXCR7, which presumably acts as a scavenger receptor in muscle, was not affected by training. Attempts to dissect the molecular events underlying the training-dependent effects of CXCL12 revealed that the CXCL12-CXCR4 axis activates anabolic mTOR-p70S6K signaling and prevents upregulation of the catabolic ubiquitin ligase MurF-1 in C2C12 myotubes, eventually increasing myotube diameters. Together, these findings point to a pivotal role of the CXCL12-CXCR4 axis in exercise-induced muscle maintenance and/or growth. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. MicroRNA-23a mediates post-transcriptional regulation of CXCL12 in bone marrow stromal cells

    PubMed Central

    Arabanian, Laleh S.; Fierro, Fernando A.; Stölzel, Friedrich; Heder, Carolin; Poitz, David M.; Strasser, Ruth H.; Wobus, Manja; Borhäuser, Martin; Ferrer, Ruben A.; Platzbecker, Uwe; Schieker, Matthias; Docheva, Denitsa; Ehninger, Gerhard; Illmer, Thomas

    2014-01-01

    The chemokine CXCL12 regulates the interaction between hematopoietic stem and progenitor cells and bone marrow stromal cells. Although its relevance in the bone marrow niche is well recognized, the regulation of CXCL12 by microRNA is not completely understood. We transfected a library of 486 microRNA in the bone marrow stromal cell line SCP-1 and studied the expression of CXCL12. Twenty-seven microRNA were shown to downregulate expression of CXCL12. Eight microRNA (miR-23a, 130b, 135, 200b, 200c, 216, 222, and 602) interacted directly with the 3′UTR of CXCL12. Next, we determined that only miR-23a is predicted to bind to the 3′UTR and is strongly expressed in primary bone marrow stromal cells. Modulation of miR-23a changes the migratory potential of hematopoietic progenitor cells in co-culture experiments. We discovered that TGFB1 mediates its inhibitory effect on CXCL12 levels by upregulation of miR-23a. This process was partly reversed by miR-23a molecules. Finally, we determined an inverse expression of CXCL12 and miR-23a in stromal cells from patients with myelodys-plastic syndrome indicating that the interaction has a pathophysiological role. Here, we show for the first time that CXCL12-targeting miR23a regulates the functional properties of the hematopoietic niche. PMID:24584347

  3. Abortifacient metapristone (RU486 derivative) interrupts CXCL12/CXCR4 axis for ovarian metastatic chemoprevention.

    PubMed

    Zheng, Ning; Chen, Jiahang; Li, Tao; Liu, Weiqun; Liu, Jian; Chen, Hongning; Wang, Jichuang; Jia, Lee

    2017-03-09

    Recent global epidemiological studies revealed the lower ovarian cancer death from long-term use of oral contraceptives. However, the underlying mechanism of action is not clear. Here, we use the abortifacient metapristone (RU486 derivative) to test the hypothesis that the contraceptives might interrupt CXCL12/CXCR4 chemokine axis to inhibit ovarian cancer metastasis. Metapristone at concentrations (CXCL12-induced CXCR4 expression on ovarian SKOV-3 and IGROV-1 cell lines, and down-regulates the CXCR4-related mRNAs and intracellular proteins. Metapristone concentration-dependently inhibits cancer cell proliferation via interfering the CXCL12-activated Akt and ERK signaling pathways. The effect of CXCL12 could be antagonized by AMD3100. Metapristone interrupts the cellular filamentous actin polymerization and the related chemotaxis. Metapristone also concentration-dependently inhibits the events of cellular invasion, migration, and adhesion. The inhibition is related to down-regulation of the invasive molecules MMP-2, MMP-9, COX-2, and VEGF without affecting the adhesion molecules ICAM-1, integrins α1, α3, α5, α6, and β1. Six-week in vivo xenograft study confirms the anti-metastatic effect of metapristone on mouse ascites and metastatic foci induced by intraperitoneal IGROV-1 and CXCL12. The present studies, for the first time, reveal the therapeutic effect of metapristone on ovarian metastasis and the CXCL12/CXCR4 axis-related mechanisms of actions.

  4. CXCL12 N-terminal end is sufficient to induce chemotaxis and proliferation of neural stem/progenitor cells.

    PubMed

    Filippo, Thais R M; Galindo, Layla T; Barnabe, Gabriela F; Ariza, Carolina B; Mello, Luiz E; Juliano, Maria A; Juliano, Luiz; Porcionatto, Marimélia A

    2013-09-01

    Neural stem/progenitor cells (NSC) respond to injury after brain injuries secreting IL-1, IL-6, TNF-α, IL-4 and IL-10, as well as chemokine members of the CC and CXC ligand families. CXCL12 is one of the chemokines secreted at an injury site and is known to attract NSC-derived neuroblasts, cells that express CXCL12 receptor, CXCR4. Activation of CXCR4 by CXCL12 depends on two domains located at the N-terminal of the chemokine. In the present work we aimed to investigate if the N-terminal end of CXCL12, where CXCR4 binding and activation domains are located, was sufficient to induce NSC-derived neuroblast chemotaxis. Our data show that a synthetic peptide analogous to the first 21 amino acids of the N-terminal end of CXCL12, named PepC-C (KPVSLSYRCPCRFFESHIARA), is able to promote chemotaxis of neuroblasts in vivo, and stimulate chemotaxis and proliferation of CXCR4+ cells in vitro, without affecting NSC fate. We also show that PepC-C upregulates CXCL12 expression in vivo and in vitro. We suggest the N-terminal end of CXCL12 is responsible for a positive feedback loop to maintain a gradient of CXCL12 that attracts neuroblasts from the subventricular zone into an injury site.

  5. SDF-1/CXCL12 modulates mitochondrial respiration of immature blood cells in a bi-phasic manner.

    PubMed

    Messina-Graham, Steven; Broxmeyer, Hal

    2016-05-01

    SDF-1/CXCL12 is a potent chemokine required for the homing and engraftment of hematopoietic stem and progenitor cells. Previous data from our group has shown that in an SDF-1/CXCL12 transgenic mouse model, lineage(-) Sca-1(+) c-Kit(+) (LSK) bone marrow cells have reduced mitochondrial membrane potential versus wild-type. These results suggested that SDF-1/CXCL12 may function to keep mitochondrial respiration low in immature blood cells in the bone marrow. Low mitochondrial metabolism helps to maintain low levels of reactive oxygen species (ROS), which can influence differentiation. To test whether SDF-1/CXCL12 regulates mitochondrial metabolism, we employed the human leukemia cell line HL-60, that expresses high levels of the SDF-1/CXCL12 receptor, CXCR4, as a model of hematopoietic progenitor cells in vitro. We treated HL-60 cells with SDF-1/CXCL12 for 2 and 24h. Oxygen consumption rates (OCR), mitochondrial-associated ATP production, mitochondrial mass, and mitochondrial membrane potential of HL-60 cells were significantly reduced at 2h and increased at 24h as compared to untreated control cells. These biphasic effects of SDF-1/CXCL12 were reproduced with lineage negative primary mouse bone marrow cells, suggesting a novel function of SDF-1/CXCL12 in modulating mitochondrial respiration by regulating mitochondrial oxidative phosphorylation, ATP production and mitochondrial content. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Downregulation of CXCL12 in mesenchymal stromal cells by TGFβ promotes breast cancer metastasis

    PubMed Central

    Yu, P F; Huang, Y; Xu, C L; Lin, L Y; Han, Y Y; Sun, W H; Hu, G H; Rabson, A B; Wang, Y; Shi, Y F

    2017-01-01

    Mesenchymal stromal cells (MSCs) are one of major components of the tumour microenvironment. Recent studies have shown that MSC tumour residence and their close interactions with inflammatory factors are important factors that affect tumour progression. Among tumour-associated inflammatory factors, transforming growth factor β (TGFβ) is regarded as a key determinant of malignancy. By employing a lung metastasis model of a murine breast cancer, we show here that the prometastatic effect of MSCs was dependent on their response to TGFβ. Interestingly, we found that MSC-produced CXCL12, an important chemokine in tumour metastasis, was markedly inhibited by TGFβ. Furthermore, silencing of CXCL12 in TGFβ-unresponsive MSCs restored their ability to promote tumour metastasis. We found that 4T1 breast cancer cells expressed high levels of CXCR7, but not of CXCR4, both of which are CXCL12 receptors. In presence of CXCL12, CXCR7 expression on tumour cells was decreased. Indeed, when CXCR7 was silenced in breast cancer cells, their metastatic ability was inhibited. Therefore, our data demonstrated that sustained expression of CXCL12 by MSCs in the primary tumour site inhibits metastasis through reduction of CXCR7, while, in the presence of TGFβ, this CXCL12 effect of MSCs on tumour cells is relieved. Importantly, elevated CXCR7 and depressed CXCL12 expression levels were prominent features of clinical breast cancer lesions and were related significantly with poor survival. Our findings reveal a novel mechanism of MSC effects on malignant cells through which crosstalk between MSCs and TGFβ regulates tumour metastasis. PMID:27669436

  7. Disturbed CXCR4/CXCL12 axis in paediatric precursor B-cell acute lymphoblastic leukaemia.

    PubMed

    van den Berk, Lieke C J; van der Veer, Arian; Willemse, Marieke E; Theeuwes, Myrte J G A; Luijendijk, Mirjam W; Tong, Wing H; van der Sluis, Inge M; Pieters, Rob; den Boer, Monique L

    2014-07-01

    Malignant cells infiltrating the bone marrow (BM) interfere with normal cellular behaviour of supporting cells, thereby creating a malignant niche. We found that CXCR4-receptor expression was increased in paediatric precursor B-cell acute lymphoblastic leukaemia (BCP-ALL) cells compared with normal mononuclear haematopoietic cells (P < 0·0001). Furthermore, high CXCR4-expression correlated with an unfavourable outcome in BCP-ALL (5-year cumulative incidence of relapse ± standard error: 38·4% ± 6·9% in CXCR4-high versus 12% ± 4·6% in CXCR4-low expressing cases, P < 0·0001). Interestingly, BM levels of the CXCR4-ligand (CXCL12) were 2·7-fold lower (P = 0·005) in diagnostic BCP-ALL samples compared with non-leukaemic controls. Induction chemotherapy restored CXCL12 levels to normal. Blocking the CXCR4-receptor with Plerixafor showed that the lower CXCL12 serum levels at diagnosis could not be explained by consumption by the leukaemic cells, nor did we observe an altered CXCL12-production capacity of BM-mesenchymal stromal cells (BM-MSC) at this time-point. We rather observed that a very high density of leukaemic cells negatively affected CXCL12-production by the BM-MSC while stimulating the secretion levels of granulocyte colony-stimulating factor (G-CSF). These results suggest that highly proliferative leukaemic cells are able to down-regulate secretion of cytokines involved in homing (CXCL12), while simultaneously up-regulating those involved in haematopoietic mobilization (G-CSF). Therefore, interference with the CXCR4/CXCL12 axis may be an effective way to mobilize BCP-ALL cells.

  8. The chemokine CXCL12 and its receptor CXCR4 are implicated in human seminoma metastasis.

    PubMed

    McIver, S C; Loveland, K L; Roman, S D; Nixon, B; Kitazawa, R; McLaughlin, E A

    2013-05-01

    Seminoma and non-seminoma tumours increasingly occur within the western population. These tumours originate from carcinoma in situ (CIS) cells, which arise from dysfunctional gonocytes. CXCL12 and its receptors, CXCR4 and CXCR7, have been implicated in migration, proliferation and survival of gonocytes and their precursors and progeny, primordial germ cells and spermatogonial stem cells respectively. We previously found evidence that several miRNA molecules predicted to modulate CXCR4 signalling are differentially expressed during the differentiation of gonocytes into spermatogonia in mice. Bioinformatic analysis predicted these miRNA to modulate CXCR4 signalling, leading us to hypothesize that CXCL12-mediated CXCR4 signalling is involved in the disrupted differentiation of gonocytes that underpins CIS formation. Indeed, we detected CXCL12 in Sertoli cells of normal human testis, and relatively high expression in tumour stroma with concomitant weak staining in dispersed tumour cells. In contrast, CXCR4 was expressed in spermatogonial and meiotic germ cells of normal testis and in the majority of tumour cells. Quantitative RT-PCR identified elevated CXCR4 transcript levels in seminoma compared with normal testis and to non-seminoma, potentially reflecting the higher proportion of dysfunctional germ cells within seminomas. In the normal testis, expression of CXCR4 downstream signalling molecules phospho-MEK1/2 and phospho-ERK1/2 correlated with CXCR4/CXCL12 expression. Strikingly, this correlation was absent in seminoma and non-seminoma samples, suggesting that CXCL12 signalling is disrupted. Proliferation rate and cell survival were not altered by CXCL12 in either seminoma (TCam-2) or non-seminoma (833ke) cell lines. However, CXCL12 exposure induced TCam-2 cell invasion though simulated basement membrane, while in contrast, we provide the novel evidence that CXCR4-expressing non-seminoma cell lines 833ke and NTera2/D1 do not invade in response to CXCL12. These

  9. Intermediate Progenitors Facilitate Intracortical Progression of Thalamocortical Axons and Interneurons through CXCL12 Chemokine Signaling.

    PubMed

    Abe, Philipp; Molnár, Zoltán; Tzeng, Yi-Shiuan; Lai, Dar-Ming; Arnold, Sebastian J; Stumm, Ralf

    2015-09-23

    Glutamatergic principal neurons, GABAergic interneurons and thalamocortical axons (TCAs) are essential elements of the cerebrocortical network. Principal neurons originate locally from radial glia and intermediate progenitors (IPCs), whereas interneurons and TCAs are of extrinsic origin. Little is known how the assembly of these elements is coordinated. C-X-C motif chemokine 12 (CXCL12), which is known to guide axons outside the neural tube and interneurons in the cortex, is expressed in the meninges and IPCs. Using mouse genetics, we dissected the influence of IPC-derived CXCL12 on TCAs and interneurons by showing that Cxcl12 ablation in IPCs, leaving meningeal Cxcl12 intact, attenuates intracortical TCA growth and disrupts tangential interneuron migration in the subventricular zone. In accordance with strong CXCR4 expression in the forming thalamus and TCAs, we identified a CXCR4-dependent growth-promoting effect of CXCL12 on TCAs in thalamus explants. Together, our findings indicate a cell-autonomous role of CXCR4 in promoting TCA growth. We propose that CXCL12 signals from IPCs link cortical neurogenesis to the progression of TCAs and interneurons spatially and temporally. Significance statement: The cerebral cortex exerts higher brain functions including perceptual and emotional processing. Evolutionary expansion of the mammalian cortex is mediated by intermediate progenitors, transient amplifying cells generating cortical excitatory neurons. During the peak period of cortical neurogenesis, migrating precursors of inhibitory interneurons originating in subcortical areas and thalamic axons invade the cortex. Although defects in the assembly of cortical network elements cause neurological and mental disorders, little is known how neurogenesis, interneuron recruitment, and axonal ingrowth are coordinated. We demonstrate that intermediate progenitors release the chemotactic cytokine CXCL12 to promote intracortical interneuron migration and growth of thalamic axons

  10. Expression of the CXCR4 ligand SDF-1/CXCL12 is prognostically important for adenocarcinoma and large cell carcinoma of the lung.

    PubMed

    Sterlacci, William; Saker, Shereen; Huber, Bettina; Fiegl, Michael; Tzankov, Alexandar

    2016-04-01

    The SDF-1/CXCR4 axis is associated with tumor progression and has been reported as a prognostic parameter, although with conflicting data for non-small cell lung cancer (NSCLC). This study examines a large cohort of clinically and pathologically well-characterized NSCLC patients and includes the activated form of CXCR4 (pCXCR4), which has not been studied in this context so far. SDF-1, CXCR4, and pCXCR4 were assessed immunohistochemically in 371 surgically resected NSCLC using a standardized tissue microarray platform. Extensive clinical and pathological data and a postoperative follow-up period of 17 years enabled detailed correlations. CXCR4 and pCXCR4 were frequently expressed on squamous cell carcinoma. Membranous expression of SDF-1 was a marker of poor prognosis and proved to be an independent prognostic parameter for the entire cohort and for patients with adenocarcinoma (ACA) and large cell carcinoma (LCC). Targeted cancer therapies blocking SDF-1/CXCR4 interaction already exist, and our data suggest that expression of SDF-1, especially on poorer prognosis subgroups of LCC and ACA, indicates patients that might benefit more than others. This should be taken into account when assessing the effectiveness of such targeted approaches for NSCLC patients and could lead to important implications.

  11. Role of metalloproteinases MMP-9 and MT1-MMP in CXCL12-promoted myeloma cell invasion across basement membranes.

    PubMed

    Parmo-Cabañas, Marisa; Molina-Ortiz, Isabel; Matías-Román, Salomón; García-Bernal, David; Carvajal-Vergara, Xonia; Valle, Inmaculada; Pandiella, Atanasio; Arroyo, Alicia G; Teixidó, Joaquin

    2006-01-01

    Malignant plasma cells in multiple myeloma home to the bone marrow (BM), accumulate in different niches and, in late disease, migrate from the BM into blood. These migratory events involve cell trafficking across extracellular matrix (ECM)-rich basement membranes and interstitial tissues. Metalloproteinases (MMP) degrade ECM and facilitate tumour cell invasion. The chemokine CXCL12 is expressed in the BM, and it was previously shown that it triggers myeloma cell migration and activation. In the present work we show that CXCL12 promotes myeloma cell invasion across Matrigel-reconstituted basement membranes and type I collagen gels. MMP-9 activity was required for invasion through Matrigel towards CXCL12, whereas TIMP-1, a MMP-9 inhibitor that we found to be expressed by myeloma and BM stromal cells, impaired the invasion. In addition, we show that the membrane-bound MT1-MMP metalloproteinase is expressed by myeloma cells and contributes to CXCL12-promoted myeloma cell invasion across Matrigel. Increase in MT1-MMP expression, as well as induction of its membrane polarization by CXCL12 in myeloma cells, might represent potential mechanisms contributing to this invasion. CXCL12-promoted invasion across type I collagen involved metalloproteinases different from MT1-MMP. These data indicate that CXCL12 could contribute to myeloma cell trafficking in the BM involving MMP-9 and MT1-MMP activities.

  12. CXCL12 induces hepatic stellate cell contraction through a calcium-independent pathway.

    PubMed

    Saiman, Yedidya; Agarwal, Ritu; Hickman, DaShawn A; Fausther, Michel; El-Shamy, Ahmed; Dranoff, Jonathan A; Friedman, Scott L; Bansal, Meena B

    2013-09-01

    Liver fibrosis, with subsequent development of cirrhosis and ultimately portal hypertension, results in the death of patients with end-stage liver disease if liver transplantation is not performed. Hepatic stellate cells (HSCs), central mediators of liver fibrosis, resemble tissue pericytes and regulate intrahepatic blood flow by modulating pericapillary resistance. Therefore, HSCs can contribute to portal hypertension in patients with chronic liver disease (CLD). We have previously demonstrated that activated HSCs express functional chemokine receptor, CXCR4, and that receptor engagement by its ligand, CXCL12, which is increased in patients with CLD, leads to further stellate cell activation in a CXCR4-specific manner. We therefore hypothesized that CXCL12 promotes HSC contraction in a CXCR4-dependent manner. Stimulation of HSCs on collagen gel lattices with CXCL12 led to gel contraction and myosin light chain (MLC) phosphorylation, which was blocked by addition of AMD3100, a CXCR4 small molecule inhibitor. These effects were further mediated by the Rho kinase pathway since both Rho kinase knockdown or Y-27632, a Rho kinase inhibitor, blocked CXCL12 induced phosphorylation of MLC and gel contraction. BAPTA-AM, a calcium chelator, had no effect, indicating that this pathway is calcium sensitive but not calcium dependent. In conclusion, CXCL12 promotes stellate cell contractility in a predominantly calcium-independent fashion. Our data demonstrates a novel role of CXCL12 in stellate cell contraction and the availability of small molecule inhibitors of the CXCL12/CXCR4 axis justifies further investigation into its potential as therapeutic target for portal hypertension.

  13. Emerging Targets in Pituitary Adenomas: Role of the CXCL12/CXCR4-R7 System.

    PubMed

    Barbieri, Federica; Thellung, Stefano; Würth, Roberto; Gatto, Federico; Corsaro, Alessandro; Villa, Valentina; Nizzari, Mario; Albertelli, Manuela; Ferone, Diego; Florio, Tullio

    2014-01-01

    Chemokines are chemotactic regulators of immune surveillance in physiological and pathological conditions such as inflammation, infection, and cancer. Several chemokines and cognate receptors are constitutively expressed in the central nervous system, not only in glial and endothelial cells but also in neurons, controlling neurogenesis, neurite outgrowth, and axonal guidance during development. In particular, the chemokine CXCL12 and its receptors, CXCR4 and CXCR7, form a functional network that controls plasticity in different brain areas, influencing neurotransmission, neuromodulation, and cell migration, and the dysregulation of this chemokinergic axis is involved in several neurodegenerative, neuroinflammatory, and malignant diseases. CXCR4 primarily mediates the transduction of proliferative signals, while CXCR7 seems to be mainly responsible for scavenging CXCL12. Importantly, the multiple intracellular signalling generated by CXCL12 interaction with its receptors influences hypothalamic modulation of neuroendocrine functions, although a direct modulation of pituitary functioning via autocrine/paracrine mechanisms was also reported. Both CXCL12 and CXCR4 are constitutively overexpressed in pituitary adenomas and their signalling induces cell survival and proliferation, as well as hormonal hypersecretion. In this review we focus on the physiological and pathological functions of immune-related cyto- and chemokines, mainly focusing on the CXCL12/CXCR4-7 axis, and their role in pituitary tumorigenesis. Accordingly, we discuss the potential targeting of CXCR4 as novel pharmacological approach for pituitary adenomas.

  14. Drug Design Targeting the CXCR4/CXCR7/CXCL12 Pathway.

    PubMed

    Xu, Dongsheng; Li, Rongshi; Wu, Jianguo; Jiang, Li; Zhong, Haizhen A

    2016-01-01

    Under physiological conditions, CXCL12 modulates cell proliferation, survival, angiogenesis, and migration mainly through CXCR4. Interestingly, the newly discovered receptor CXCR7 for CXCL12 is highly expressed in many tumor cells as well as tumor-associated blood vessels, although the level of CXCR7 in normal blood cells is low. Recently, many studies have suggested that CXCR7 promotes cell growth and metastasis in various cancers, including lymphoma and leukemia, hepatocecullar, ovarian, colorectal, breast and lung cancer. Compared to CXCR4, CXCR7 is a non-classical GPCR that is unable to activate G proteins. The function of CXCR7 is generally considered to be mediated by: (a) recruiting β-arrestin-2; (b) heterodimerizing with CXCR4; and (c) acting as a "scavenger" of CXCL12, thus lowering the level of CXCL12 to weaken the activity of CXCR4. However, the crosstalk between CXCL12/CXCR7/CXCR4 and other signaling pathways (such as the p38 MAPK pathway, the PI3K/mTOR pathway, the STAT3 signaling, and metalloproteinases MMP-9 and MMP-2) is more complicated. The function of CXCR7 is also involved in modulating tumor microenvironment, tumor cell migration and apoptosis. Understanding these complex interactions will provide insight in drug design targeting the CXCR7 as potential anticancer therapy.

  15. Emerging Targets in Pituitary Adenomas: Role of the CXCL12/CXCR4-R7 System

    PubMed Central

    Thellung, Stefano; Würth, Roberto; Gatto, Federico; Corsaro, Alessandro; Villa, Valentina; Nizzari, Mario; Albertelli, Manuela; Ferone, Diego; Florio, Tullio

    2014-01-01

    Chemokines are chemotactic regulators of immune surveillance in physiological and pathological conditions such as inflammation, infection, and cancer. Several chemokines and cognate receptors are constitutively expressed in the central nervous system, not only in glial and endothelial cells but also in neurons, controlling neurogenesis, neurite outgrowth, and axonal guidance during development. In particular, the chemokine CXCL12 and its receptors, CXCR4 and CXCR7, form a functional network that controls plasticity in different brain areas, influencing neurotransmission, neuromodulation, and cell migration, and the dysregulation of this chemokinergic axis is involved in several neurodegenerative, neuroinflammatory, and malignant diseases. CXCR4 primarily mediates the transduction of proliferative signals, while CXCR7 seems to be mainly responsible for scavenging CXCL12. Importantly, the multiple intracellular signalling generated by CXCL12 interaction with its receptors influences hypothalamic modulation of neuroendocrine functions, although a direct modulation of pituitary functioning via autocrine/paracrine mechanisms was also reported. Both CXCL12 and CXCR4 are constitutively overexpressed in pituitary adenomas and their signalling induces cell survival and proliferation, as well as hormonal hypersecretion. In this review we focus on the physiological and pathological functions of immune-related cyto- and chemokines, mainly focusing on the CXCL12/CXCR4-7 axis, and their role in pituitary tumorigenesis. Accordingly, we discuss the potential targeting of CXCR4 as novel pharmacological approach for pituitary adenomas. PMID:25484899

  16. PI3K{gamma} activation by CXCL12 regulates tumor cell adhesion and invasion

    SciTech Connect

    Monterrubio, Maria; Mellado, Mario; Carrera, Ana C.

    2009-10-16

    Tumor dissemination is a complex process, in which certain steps resemble those in leukocyte homing. Specific chemokine/chemokine receptor pairs have important roles in both processes. CXCL12/CXCR4 is the most commonly expressed chemokine/chemokine receptor pair in human cancers, in which it regulates cell adhesion, extravasation, metastatic colonization, angiogenesis, and proliferation. All of these processes require activation of signaling pathways that include G proteins, phosphatidylinositol-3 kinase (PI3K), JAK kinases, Rho GTPases, and focal adhesion-associated proteins. We analyzed these pathways in a human melanoma cell line in response to CXCL12 stimulation, and found that PI3K{gamma} regulates tumor cell adhesion through mechanisms different from those involved in cell invasion. Our data indicate that, following CXCR4 activation after CXCL12 binding, the invasion and adhesion processes are regulated differently by distinct downstream events in these signaling cascades.

  17. CXCL12-CXCR7 axis contributes to the invasive phenotype of pancreatic cancer

    PubMed Central

    Yang, Jian-Yu; Zhang, Zhi-Gang; Liang, Zhi-Yong; Zhou, Wei-Xun; You, Lei; Zhang, Tai-Ping; Zhao, Yu-Pei

    2016-01-01

    Chemokine (C-X-C motif) receptor 7 (CXCR7) and its ligand, chemokine (C-X-C motif) ligand 12 (CXCL12), were established to be involved in biological behaviors and associated with prognosis in many cancers. However, effects, underlying mechanisms of CXCL12-CXCR7 axis in invasive phenotype of pancreatic cancer (PC) and its clinicopathologic significances have not been comprehensively explored. In the present study, it was first found by tissue microarray-based immunohistochemistry that CXCL12 and CXCR7 staining scores were significantly associated with vessel invasion and overall survival in two independent cohorts of PC. Besides, co-expression of these proteins was an independent prognosticator in multivariate analysis in both cohorts. Then, migration and invasion, but not proliferation, were decreased in CXCR7-stably silenced PC cells, whereas opposite changes were observed in CXCR7-stably overexpressed cells, accompanied by alterations of mTOR and Rho/ROCK pathways. CXCL12 stimulated migration, invasion, CXCR7 expression and phosphorylation of key mTOR proteins. AMD3100 did not influence effects of CXCL12. Two mTOR inhibitors, rapamycin and Torin1, reversed enhanced invasive phenotypes and mTOR phosphorylation in CXCR7-overexpressed cells. Moreover, CXCR7 directly interacts with mTOR. Finally, liver metastasis, but not growth, was affected by CXCR7 status in orthotopically-implanted PC models in nude mice. Collectively, CXCL12-CXCR7 axis accelerates migration and invasion of PC cells through mTOR and Rho/ROCK pathways, and predicts poor prognosis of PC. PMID:27542220

  18. Decreased interstitial FOXP3(+) lymphocytes in usual interstitial pneumonia with discrepancy of CXCL12/CXCR4 axis.

    PubMed

    Shimizu, Y; Dobashi, K; Endou, K; Ono, A; Yanagitani, N; Utsugi, M; Sano, T; Ishizuka, T; Shimizu, K; Tanaka, S; Mori, M

    2010-01-01

    Regulatory T cells (Treg) play a critical role in immune homeostasis and expansion of Treg is controlled by chemokine receptors. The chemokine CXCL12 and its G-protein-coupled receptor (CXCR4) are involved in the development of idiopathic pulmonary fibrosis (IPF), but the association of Treg with the CXCL12/CXCR4 axis has not been documented. The aim of this study is to determine the distribution and extent of CXCL12/CXCR4 expression in idiopathic type of pulmonary fibrosis, and the relation of Treg expansion in the interstitium of pulmonary fibrosis patients to CXCL12/CXCR4 expression. CXCL12 expression was examined by immunostaining and ELISA in tissue specimens from patients with usual interstitial pneumonia (UIP, n=15), patients with fibrotic non-specific interstitial pneumonia (f-NSIP, n=4), and controls (n=6). CXCR4 expression was examined by in situ hybridization and immunoblotting. Expression of CD45, CD3, CD20, transcription factor forkhead box P3 (FOXP3), and CD25 was assessed by immunostaining. Fibrosis was evaluated by determining the established fibrosis (EF) score. The CXCL12/CXCR4 axis was upregulated in UIP and f-NSIP, and CXCL12 derived from lung tissue attracted CXCR4(+) cells. CXCR4(+) cells showed a CD3(+) cell distribution pattern. The interstitial FOXP3(+)/CD3(+) and CD25(+)/CD3(+) cell ratios were lower in UIP than f-NSIP, but the CXCR4(+)/CD3(+) cell ratio was not different. The FOXP3(+)/CD3(+) cell ratio and EF score were inversely correlated. These findings suggest that the CXCL12/CXCR4 axis contributes to inflammation in UIP and f-NSIP by promoting the accumulation CXCR4(+) lymphocytes, and a decrease of Treg is correlated with the severity of fibrosis in UIP.

  19. Involvement of the CXCL12 System in the Stimulatory Effects of Prenatal Exposure to High-Fat Diet on Hypothalamic Orexigenic Peptides and Behavior in Offspring

    PubMed Central

    Poon, Kinning; Barson, Jessica R.; Shi, Huanzhi; Chang, Guo Qing; Leibowitz, Sarah F.

    2017-01-01

    Exposure to a high fat diet (HFD) during gestation stimulates neurogenesis and expression of hypothalamic orexigenic neuropeptides that affect consummatory and emotional behaviors. With recent studies showing a HFD to increase inflammation, this report investigated the neuroinflammatory chemokine, CXCL12, and compared the effects of prenatal CXCL12 injection to those of prenatal HFD exposure, first, by testing whether the HFD affects circulating CXCL12 in the dam and the CXCL12 system in the offspring brain, and then by examining whether prenatal exposure to CXCL12 itself mimics the effects of a HFD on hypothalamic neuropeptides and emotional behaviors. Our results showed that prenatal exposure to a HFD significantly increased circulating levels of CXCL12 in the dam, and that daily injections of CXCL12 induced a similar increase in CXCL12 levels as the HFD. In addition, prenatal HFD exposure significantly increased the expression of CXCL12 and its receptors, CXCR4 and CXCR7, in the hypothalamic paraventricular nucleus (PVN) of the offspring. Finally, the results revealed strong similarities in the effects of prenatal HFD and CXCL12 administration, which both stimulated neurogenesis and enkephalin (ENK) expression in the PVN, while having inconsistent or no effect in other regions of the hypothalamus, and also increased anxiety as measured by several behavioral tests. These results focus attention specifically on the CXCL12 chemokine system in the PVN of the offspring as being possibly involved in the stimulatory effects of prenatal HFD exposure on ENK-expressing neurons in the PVN and their associated changes in emotional behavior. PMID:28567007

  20. CXCL12/CXCR4 axis regulates neovascularization and lymphangiogenesis in sutured corneas in mice

    PubMed Central

    DU, LING-LING; LIU, PING

    2016-01-01

    The present study aimed to determine the plausible functional role of chemokine (C-X-C motif) ligand 12 (CXCL12)/chemokine (C-X-C motif) receptor 4 (CXCR4) in inflammatory corneal hemangiogenesis and lymphangiogenesis in vivo. Corneal hemangiogenesis and lymphangiogenesis were induced by placing an 11-0 nylon suture in an intrastromal position. The expression levels of the vascular endothelial growth factor (VEGF) family, CXCL12 and CXCR4 in the corneas were investigated in the corneas using reverse transcription-quantitative polymerase chain reaction and immunohistochemistry. Corneal hemangiogenic and lymphangiogenic responses were assessed by immunofluorescence using specific antibodies against cluster of differentiation 31 and lymphatic vessel endothelial hyaluronan receptor-1. Subconjunctival injection of AMD3100 to the sutured corneas was also performed. CXCL12/CXCR4 mRNA and protein expression levels increased markedly in suture-induced corneal neovascularization (CNV) and decreased with AMD3100 treatment. Hemangiogenesis and lymphangiogenesis were captured in images using immunofluorescence and were shown to be markedly increased with suture placement and reduced with AMD3100 treatment. VEGF-A/VEGFR-1 and VEGF-C/VEGFR-3 mRNA expression levels were upregulated in the suture placement and control groups, whereas the expression levels of all the factors were downregulated in the AMD3100 treatment group. The results from the present study demonstrated that CXCL12/CXCR4 interactions regulate hemangiogenesis and lymphangiogenesis in suture-induced CNV. AMD3100 may be a novel therapeutic target for the prevention of blindness. PMID:27121088

  1. Misbalanced CXCL12 and CCL5 Chemotactic Signals in Vitiligo Onset and Progression.

    PubMed

    Rezk, Ahmed F; Kemp, Daria Marley; El-Domyati, Moetaz; El-Din, Wael Hosam; Lee, Jason B; Uitto, Jouni; Igoucheva, Olga; Alexeev, Vitali

    2017-05-01

    Generalized nonsegmental vitiligo is often associated with the activation of melanocyte-specific autoimmunity. Because chemokines play an important role in the maintenance of immune responses, we examined chemotactic signatures in cultured vitiligo melanocytes and skin samples of early (≤2 months) and advanced (≥6 months) vitiligo. Analysis showed that melanocytes in early lesions have altered expression of several chemotaxis-associated molecules, including elevated secretion of CXCL12 and CCL5. Higher levels of these chemokines coincided with prominent infiltration of the skin with antigen presenting cells (APCs) and T cells. Most of the intralesional APCs expressed the CD86 maturation marker and co-localized with T cells, particularly in early vitiligo lesions. These observations were confirmed by in vivo animal studies showing preferential recruitment of APCs and T cells to CXCL12- and CCL5-expressing transplanted melanocytes, immunotargeting of the chemokine-positive cells, continuous loss of the pigment-producing cells from the epidermis, and development of vitiligo-like lesions. Taken together, our studies show that melanocyte-derived CXCL12 and CCL5 support APC and T-cell recruitment, antigen acquisition, and T-cell activation in early vitiligo and reinforce the role of melanocyte-derived CXCL12 and CCL5 in activation of melanocyte-specific immunity and suggest inhibition of these chemotactic axes as a strategy for vitiligo stabilization. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  2. CXCR7 Participates in CXCL12-mediated Cell Cycle and Proliferation Regulation in Mouse Neural Progenitor Cells

    PubMed Central

    Wang, Y.; Xu, P.; Qiu, L.; Zhang, M.; Huang, Y.; Zheng, J.C.

    2016-01-01

    Background: Cell cycle regulation of neural progenitor cells (NPCs) is an essential process for neurogenesis, neural development, and repair after brain trauma. Stromal cell-derived factor-1 (SDF-1, CXCL12) and its receptors CXCR4 and CXCR7 are well known in regulating the migration and survival of NPCs. The effects of CXCL12 on NPCs proliferation, cell cycle regulation, and their associated signaling pathways remain unclear. Cyclin D1 is a protein required for progression through the G1 phase of the cell cycle and a known downstream target of β-catenin. Therefore, cyclin D1 plays critical roles of cell cycle regulation, proliferation, and survival in NPCs. Methods: Primary mouse NPCs (mNPCs) were derived from brain tissues of wild-type, Cxcr4 knockout, or Cxcr7 knockout mice at mouse embryonic day 13.5 (E13.5). Flow cytometry was used to perform cell cycle analysis by quantitation of DNA content. Real-time PCR and Western blot were used to evaluate mRNA and protein expressions, respectively. Ki67 immunostaining and TUNEL assay were used to assess the proliferation and survival of mNPCs, respectively. Results: CXCL12 pretreatment led to the shortening of G0/G1 phase and lengthening of S phase, suggesting that CXCL12 regulates cell cycle progression in mNPCs. Consistently, CXCL12 treatment increased the expression of CyclinD1 and β-catenin, and promoted proliferation and survival of mNPCs. Cxcr7 knockout of mNPCs blocked CXCL12-mediated mNPCs proliferation, whereas Cxcr4 knockout mNPC did not significantly effect CXCL12- mediated mNPCs proliferation. Conclusion: CXCR7 plays an important role in CXCL12-mediated mNPC cell cycle regulation and proliferation. PMID:27573194

  3. CXCR7 Participates in CXCL12-mediated Cell Cycle and Proliferation Regulation in Mouse Neural Progenitor Cells.

    PubMed

    Wang, Y; Xu, P; Qiu, L; Zhang, M; Huang, Y; Zheng, J C

    2016-01-01

    Cell cycle regulation of neural progenitor cells (NPCs) is an essential process for neurogenesis, neural development, and repair after brain trauma. Stromal cell-derived factor-1 (SDF-1, CXCL12) and its receptors CXCR4 and CXCR7 are well known in regulating the migration and survival of NPCs. The effects of CXCL12 on NPCs proliferation, cell cycle regulation, and their associated signaling pathways remain unclear. Cyclin D1 is a protein required for progression through the G1 phase of the cell cycle and a known downstream target of β -catenin. Therefore, cyclin D1 plays critical roles of cell cycle regulation, proliferation, and survival in NPCs. Primary mouse NPCs (mNPCs) were derived from brain tissues of wild-type, Cxcr4 knockout, or Cxcr7 knockout mice at mouse embryonic day 13.5 (E13.5). Flow cytometry was used to perform cell cycle analysis by quantitation of DNA content. Real-time PCR and Western blot were used to evaluate mRNA and protein expressions, respectively. Ki67 immunostaining and TUNEL assay were used to assess the proliferation and survival of mNPCs, respectively. CXCL12 pretreatment led to the shortening of G0/G1 phase and lengthening of S phase, suggesting that CXCL12 regulates cell cycle progression in mNPCs. Consistently, CXCL12 treatment increased the expression of CyclinD1 and β -catenin, and promoted proliferation and survival of mNPCs. Cxcr7 knockout of mNPCs blocked CXCL12-mediated mNPCs proliferation, whereas Cxcr4 knockout mNPC did not significantly effect CXCL12- mediated mNPCs proliferation. CXCR7 plays an important role in CXCL12-mediated mNPC cell cycle regulation and proliferation.

  4. Oncogenic roles and drug target of CXCR4/CXCL12 axis in lung cancer and cancer stem cell.

    PubMed

    Wang, Zhidong; Sun, Jian; Feng, Yeqian; Tian, Xiaocai; Wang, Bin; Zhou, Yong

    2016-07-01

    Although the great progress has been made in diagnosis and therapeutic in lung cancer, it induces the most cancer death worldwide in both males and females. Chemokines, which have chemotactic abilities, contain up to 50 family members. By binding to G protein-coupled receptors (GPCR), holding seven-transmembrane domain, they function in immune cell trafficking and regulation of cell proliferation, differentiation, activation, and migration, homing under both physiologic and pathologic conditions. The alpha-chemokine receptor CXCR4 for the alpha-chemokine stromal cell-derived-factor-1 (SDF-1) is most widely expressed by tumors. In addition to human tissues of the bone marrow, liver, adrenal glands, and brain, the CXC chemokine SDF-1 or CXCL12 is also highly expressed in lung cancer tissues and is associated with lung metastasis. Lung cancer cells have the capabilities to utilize and manipulate the CXCL12/CXCR system to benefit growth and distant spread. CXCL12/CXCR4 axis is a major culprit for lung cancer and has a crucial role in lung cancer initiation and progression by activating cancer stem cell. This review provides an evaluation of CXCL12/CXCR4 as the potential therapeutic target for lung cancers; it also focuses on the synergistic effects of inhibition of CXCL12/CXCR4 axis and immunotherapy as well as chemotherapy. Together, CXCL12/CXCR4 axis can be a potential therapeutic target for lung cancers and has additive effects with immunotherapy.

  5. Involvement of the CXCL12/CXCR4 pathway in the advanced liver disease that is associated with hepatitis C virus or hepatitis B virus.

    PubMed

    Wald, Ori; Pappo, Orit; Safadi, Rifaat; Dagan-Berger, Michal; Beider, Katia; Wald, Hanna; Franitza, Suzanna; Weiss, Ido; Avniel, Shani; Boaz, Pal; Hanna, Jacob; Zamir, Gidi; Eid, Ahmed; Mandelboim, Ofer; Spengler, Ulrich; Galun, Eithan; Peled, Amnon

    2004-04-01

    Chronic hepatitis C virus (HCV) and hepatitis B virus (HBV) infection is accompanied by inflammation and fibrosis eventually leading to cirrhosis. The chemokine CXCL12 is involved in chronic inflammatory conditions. The role of the CXCL12/CXCR4 pathway in HCV- and HBV-associated liver inflammation and fibrosis was therefore studied. The levels and tissue localization of CXCL12 in liver and plasma of HCV and HBV patients were tested using immunohistochemistry and ELISA. The expression and function of CXCR4 on liver-infiltrating lymphocytes (LIL) were tested by FACS and transwell migration assays. We found that CXCL12 is expressed by bile duct epithelial cells in normal liver tissue. Bile duct proliferation and liver fibrosis in chronic HCV and HBV infection result in the anatomical re-distribution of CXCL12 in the liver. Moreover, CXCL12 is up-regulated in the endothelium of neo-blood-vessels formed in active inflammatory foci and is significantly elevated, compared with controls, in the plasma of patients with advanced liver fibrosis. Complementing these observations were others indicating that over 50% of LIL express CXCR4 and, in response to CXCL12, migrated and adhered to fibronectin. These observations suggest an important role for the CXCL12/CXCR4 pathway in recruitment and retention of immune cells in the liver during chronic HCV and HBV infection.

  6. Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4

    PubMed Central

    Collins, Paul J.; McCully, Michelle L.; Martínez-Muñoz, Laura; Santiago, César; Wheeldon, James; Caucheteux, Stephan; Thelen, Sylvia; Cecchinato, Valentina; Laufer, Julia M.; Purvanov, Vladimir; Monneau, Yoan R.; Lortat-Jacob, Hugues; Legler, Daniel F.; Uguccioni, Mariagrazia; Thelen, Marcus; Piguet, Vincent; Mellado, Mario; Moser, Bernhard

    2017-01-01

    The chemokine receptor, CXC chemokine receptor 4 (CXCR4), is selective for CXC chemokine ligand 12 (CXCL12), is broadly expressed in blood and tissue cells, and is essential during embryogenesis and hematopoiesis. CXCL14 is a homeostatic chemokine with unknown receptor selectivity and preferential expression in peripheral tissues. Here, we demonstrate that CXCL14 synergized with CXCL12 in the induction of chemokine responses in primary human lymphoid cells and cell lines that express CXCR4. Combining subactive concentrations of CXCL12 with 100–300 nM CXCL14 resulted in chemotaxis responses that exceeded maximal responses that were obtained with CXCL12 alone. CXCL14 did not activate CXCR4-expressing cells (i.e., failed to trigger chemotaxis and Ca2+ mobilization, as well as signaling via ERK1/2 and the small GTPase Rac1); however, CXCL14 bound to CXCR4 with high affinity, induced redistribution of cell-surface CXCR4, and enhanced HIV-1 infection by >3-fold. We postulate that CXCL14 is a positive allosteric modulator of CXCR4 that enhances the potency of CXCR4 ligands. Our findings provide new insights that will inform the development of novel therapeutics that target CXCR4 in a range of diseases, including cancer, autoimmunity, and HIV.—Collins, P. J., McCully, M. L., Martínez-Muñoz, L., Santiago, C., Wheeldon, J., Caucheteux, S., Thelen, S., Cecchinato, V., Laufer, J. M., Purvanov, V., Monneau, Y. R., Lortat-Jacob, H., Legler, D. F., Uguccioni, M., Thelen, M., Piguet, V., Mellado, M., Moser, B. Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4. PMID:28360196

  7. Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4.

    PubMed

    Collins, Paul J; McCully, Michelle L; Martínez-Muñoz, Laura; Santiago, César; Wheeldon, James; Caucheteux, Stephan; Thelen, Sylvia; Cecchinato, Valentina; Laufer, Julia M; Purvanov, Vladimir; Monneau, Yoan R; Lortat-Jacob, Hugues; Legler, Daniel F; Uguccioni, Mariagrazia; Thelen, Marcus; Piguet, Vincent; Mellado, Mario; Moser, Bernhard

    2017-07-01

    The chemokine receptor, CXC chemokine receptor 4 (CXCR4), is selective for CXC chemokine ligand 12 (CXCL12), is broadly expressed in blood and tissue cells, and is essential during embryogenesis and hematopoiesis. CXCL14 is a homeostatic chemokine with unknown receptor selectivity and preferential expression in peripheral tissues. Here, we demonstrate that CXCL14 synergized with CXCL12 in the induction of chemokine responses in primary human lymphoid cells and cell lines that express CXCR4. Combining subactive concentrations of CXCL12 with 100-300 nM CXCL14 resulted in chemotaxis responses that exceeded maximal responses that were obtained with CXCL12 alone. CXCL14 did not activate CXCR4-expressing cells (i.e., failed to trigger chemotaxis and Ca(2+) mobilization, as well as signaling via ERK1/2 and the small GTPase Rac1); however, CXCL14 bound to CXCR4 with high affinity, induced redistribution of cell-surface CXCR4, and enhanced HIV-1 infection by >3-fold. We postulate that CXCL14 is a positive allosteric modulator of CXCR4 that enhances the potency of CXCR4 ligands. Our findings provide new insights that will inform the development of novel therapeutics that target CXCR4 in a range of diseases, including cancer, autoimmunity, and HIV.-Collins, P. J., McCully, M. L., Martínez-Muñoz, L., Santiago, C., Wheeldon, J., Caucheteux, S., Thelen, S., Cecchinato, V., Laufer, J. M., Purvanov, V., Monneau, Y. R., Lortat-Jacob, H., Legler, D. F., Uguccioni, M., Thelen, M., Piguet, V., Mellado, M., Moser, B. Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4. © The Author(s).

  8. Chemokine receptor CXCR7 is a functional receptor for CXCL12 in brain endothelial cells.

    PubMed

    Liu, Yang; Carson-Walter, Eleanor; Walter, Kevin A

    2014-01-01

    The chemokine CXCL12 regulates multiple cell functions through its receptor, CXCR4. However, recent studies have shown that CXCL12 also binds a second receptor, CXCR7, to potentiate signal transduction and cell activity. In contrast to CXCL12/CXCR4, few studies have focused on the role of CXCR7 in vascular biology and its role in human brain microvascular endothelial cells (HBMECs) remains unclear. In this report, we used complementary methods, including immunocytofluorescence, Western blot, and flow cytometry analyses, to demonstrate that CXCR7 was expressed on HBMECs. We then employed short hairpin RNA (shRNA) technology to knockdown CXCR7 in HBMECs. Knockdown of CXCR7 in HBMECs resulted in significantly reduced HBMEC proliferation, tube formation, and migration, as well as adhesion to matrigel and tumor cells. Blocking CXCR7 with a specific antibody or small molecule antagonist similarly disrupted HBMEC binding to matrigel or tumor cells. We found that tumor necrosis factor (TNF)-α induced CXCR7 in a time and dose-response manner and that this increase preceded an increase in vascular cell adhesion molecule-1 (VCAM-1). Knockdown of CXCR7 resulted in suppression of VCAM-1, suggesting that the reduced binding of CXCR7-knockdown HBMECs may result from suppression of VCAM-1. Collectively, CXCR7 acted as a functional receptor for CXCL12 in brain endothelial cells. Targeting CXCR7 in tumor vasculature may provide novel opportunities for improving brain tumor therapy.

  9. Targeting the CXCL12/CXCR4 axis in acute myeloid leukemia: from bench to bedside.

    PubMed

    Cho, Byung-Sik; Kim, Hee-Je; Konopleva, Marina

    2017-03-01

    The interactions between the cancerous cells of acute myeloid leukemia (AML) and the bone marrow (BM) microenvironment have been postulated to be important for resistance to chemotherapy and disease relapse in AML. The chemokine receptor CXC chemokine receptor 4 (CXCR4) and its ligand, CXC motif ligand 12 (CXCL12), also known as stromal cell-derived factor 1α, are key mediators of this interaction. CXCL12 is produced by the BM microenvironment, binds and activates its cognate receptor CXCR4 on leukemic cells, facilitates leukemia cell trafficking and homing in the BM microenvironment, and keeps leukemic cells in close contact with the stromal cells and extracellular matrix that constitutively generate growth-promoting and anti-apoptotic signals. Indeed, a high level of CXCR4 expression on AML blasts is known to be associated with poor prognosis. Recent preclinical and clinical studies have revealed the safety and potential clinical utility of targeting the CXCL12/CXCR4 axis in AML with different classes of drugs, including small molecules, peptides, and monoclonal antibodies. In this review, we describe recent evidence of targeting these leukemia-stroma interactions, focusing on the CXCL12/CXCR4 axis. Related early phase clinical studies will be also introduced.

  10. Targeting the CXCL12/CXCR4 axis in acute myeloid leukemia: from bench to bedside

    PubMed Central

    Cho, Byung-Sik; Kim, Hee-Je; Konopleva, Marina

    2017-01-01

    The interactions between the cancerous cells of acute myeloid leukemia (AML) and the bone marrow (BM) microenvironment have been postulated to be important for resistance to chemotherapy and disease relapse in AML. The chemokine receptor CXC chemokine receptor 4 (CXCR4) and its ligand, CXC motif ligand 12 (CXCL12), also known as stromal cell-derived factor 1α, are key mediators of this interaction. CXCL12 is produced by the BM microenvironment, binds and activates its cognate receptor CXCR4 on leukemic cells, facilitates leukemia cell trafficking and homing in the BM microenvironment, and keeps leukemic cells in close contact with the stromal cells and extracellular matrix that constitutively generate growth-promoting and anti-apoptotic signals. Indeed, a high level of CXCR4 expression on AML blasts is known to be associated with poor prognosis. Recent preclinical and clinical studies have revealed the safety and potential clinical utility of targeting the CXCL12/CXCR4 axis in AML with different classes of drugs, including small molecules, peptides, and monoclonal antibodies. In this review, we describe recent evidence of targeting these leukemia-stroma interactions, focusing on the CXCL12/CXCR4 axis. Related early phase clinical studies will be also introduced. PMID:28219003

  11. CXCL12 Gene Therapy Ameliorates Ischemia-Induced White Matter Injury in Mouse Brain.

    PubMed

    Li, Yaning; Tang, Guanghui; Liu, Yanqun; He, Xiaosong; Huang, Jun; Lin, Xiaojie; Zhang, Zhijun; Yang, Guo-Yuan; Wang, Yongting

    2015-10-01

    Remyelination is an important repair process after ischemic stroke-induced white matter injury. It often fails because of the insufficient recruitment of oligodendrocyte progenitor cells (OPCs) to the demyelinated site or the inefficient differentiation of OPCs to oligodendrocytes. We investigated whether CXCL12 gene therapy promoted remyelination after middle cerebral artery occlusion in adult mice. The results showed that CXCL12 gene therapy at 1 week after ischemia could protect myelin sheath integrity in the perifocal region, increase the number of platelet-derived growth factor receptor-α (PDGFRα)-positive and PDGFRα/bromodeoxyuridine-double positive OPCs in the subventricular zone, and further enhance their migration to the ischemic lesion area. Coadministration of AMD3100, the antagonist for CXCL12 receptor CXCR4, eliminated the beneficial effect of CXCL12 on myelin sheath integrity and negatively influenced OPC proliferation and migration. At 5 weeks after ischemia, CXCR4 was found on the PDGFRα- and/or neuron/glia type 2 (NG2)-positive OPCs but not on the myelin basic protein-positive mature myelin sheaths, and CXCR7 was only expressed on the mature myelin sheath in the ischemic mouse brain. Our data indicated that CXCL12 gene therapy effectively protected white matter and promoted its repair after ischemic injury. The treatment at 1 week after ischemia is effective, suggesting that this strategy has a longer therapeutic time window than the treatments currently available. This study has demonstrated for the first time that CXCL12 gene therapy significantly ameliorates brain ischemia-induced white matter injury and promotes oligodendrocyte progenitor cell proliferation in the subventricular zone and migration to the perifocal area in the ischemic mouse brain. Additional data showed that CXCR4 receptor plays an important role during the proliferation and migration of oligodendrocyte progenitor cells, and CXCR7 might play a role during maturation. In

  12. Activation of Vav/Rho GTPase Signaling by CXCL12 Controls Membrane-Type Matrix Metalloproteinase–Dependent Melanoma Cell Invasion

    PubMed Central

    Bartolomé, Rubén A.; Molina-Ortiz, Isabel; Samaniego, Rafael; Sánchez-Mateos, Paloma; Bustelo, Xosé R.; Teixidó, Joaquin

    2007-01-01

    Melanoma cells express the chemokine receptor CXCR4, which confers invasive signals on binding to its ligand CXCL12. We show here that knocking down membrane-type matrix metal-loproteinase (MT1-MMP) expression translates into a blockade of invasion across reconstituted basement membranes and type I collagen gels in response to CXCL12, which is the result of lack of MMP-2 activation. Interference with MMP-2 expression further confirms its important role during this invasion. Vav proteins are guanine-nucleotide exchange factors for Rho GTPases that regulate actin dynamics and gene expression. We show that melanoma cells express Vav1 and Vav2, which are activated by CXCL12 involving Jak activity. Blocking Vav expression by RNA interference results in impaired activation of Rac and Rho by CXCL12 and in a remarkable inhibition of CXCL12-promoted invasion. Importantly, up-regulation of MT1-MMP expression by CXCL12, a mechanism contributing to melanoma cell invasion, is blocked by knocking down Vav expression or by inhibiting Jak. Together, these data indicate that activation of Jak/Vav/Rho GTPase pathway by CXCL12 is a key signaling event for MT1-MMP/MMP-2–dependent melanoma cell invasion. PMID:16397238

  13. CXCL12 Mediates Trophic Interactions between Endothelial and Tumor Cells in Glioblastoma

    PubMed Central

    Choe, Eun Joo; Woerner, B. Mark; Jackson, Erin; Sun, Tao; Leonard, Jeffrey; Piwnica-Worms, David; Rubin, Joshua B.

    2012-01-01

    Emerging evidence suggests endothelial cells (EC) play a critical role in promoting Glioblastoma multiforme (GBM) cell proliferation and resistance to therapy. The molecular basis for GBM-EC interactions is incompletely understood. We hypothesized that the chemokine CXCL12 and its receptor CXCR4 could mediate direct interactions between GBM cells and tumor-associated endothelial cells and that disruption of this interaction might be the molecular basis for the anti-tumor effects of CXCR4 antagonists. We investigated this possibility in vivo and in an in vitro co-culture model that incorporated extracellular matrix, primary human brain microvascular ECs (HBMECs) and either an established GBM cell line or primary GBM specimens. Depletion of CXCR4 in U87 GBM cells blocked their growth as intracranial xenografts indicating that tumor cell CXCR4 is required for tumor growth in vivo. In vitro, co-culture of either U87 cells or primary GBM cells with HBMECs resulted in their co-localization and enhanced GBM cell growth. Genetic manipulation of CXCL12 expression and pharmacological inhibition of its receptors CXCR4 and CXCR7 revealed that the localizing and trophic effects of endothelial cells on GBM cells were dependent upon CXCL12 and CXCR4. These findings indicate that the CXCL12/CXCR4 pathway directly mediates endothelial cell trophic function in GBMs and that inhibition of CXCL12-CXCR4 signaling may uniquely target this activity. Therapeutic disruption of endothelial cell trophic functions could complement the structural disruption of anti-angiogenic regimens and, in combination, might also improve the efficacy of radiation and chemotherapy in treating GBMs. PMID:22427929

  14. Scavenging of CXCL12 by CXCR7 Promotes Tumor Growth and Metastasis of CXCR4-positive Breast Cancer Cells

    PubMed Central

    Luker, Kathryn E.; Lewin, Sarah A.; Mihalko, Laura Anne; Schmidt, Bradley T.; Winkler, Jessica S.; Coggins, Nathaniel L.; Thomas, Dafydd G.; Luker, Gary D.

    2011-01-01

    Chemokine CXCL12 and receptor CXCR4 control multiple steps in primary tumor growth and metastasis in breast cancer and more than 20 other human malignancies. Mechanisms that regulate availability of CXCL12 in tumor microenvironments will substantially impact cancer progression and ongoing efforts to target the CXCL12-CXCR4 pathway for cancer chemotherapy. We used dual luciferase imaging to investigate CXCR7 dependent scavenging of CXCL12 in breast tumors in vivo and quantify effects of CXCR7 on tumor growth and metastasis of a separate population of CXCR4+ breast cancer cells. In a mouse xenograft model of human breast cancer, in vivo imaging showed that malignant cells expressing CXCR7 reduced bioluminescent CXCL12 secreted in the primary tumor microenvironment. Capitalizing on sensitive detection of bioluminescent CXCL12, we also demonstrated that CXCR7+ cells reduced amounts of chemokine released from orthotopic tumors into the circulation. Immunofluorescence staining of human primary breast cancers showed expression of CXCR4 and CXCR7 on malignant cells in ≈ 30% of cases. In most cases, CXCR4 and CXCR7 predominantly were expressed on separate populations of malignant cells in a tumor. We modeled these cases of human breast cancer by co-implanting tumor xenografts with CXCR4+ breast cancer cells, human mammary fibroblasts secreting CXCL12, and CXCR7+ or control breast cancer cells. Bioluminescence imaging showed that CXCR7+ breast cancer cells enhanced proliferation of CXCR4+ breast cancer cells in orthotopic tumors and spontaneous metastases. Treatment with a small molecule inhibitor of CXCR7 chemokine scavenging limited growth of CXCR4+ breast cancer cells in tumors that also contained malignant CXCR7+ cells. These studies establish a new in vivo imaging method to quantify chemokine scavenging by CXCR7 in the tumor microenvironment and identify that CXCR7+ cells promote growth and metastasis of CXCR4+ breast cancer cells. PMID:22266857

  15. Platelet-derived CXCL12 regulates monocyte function, survival, differentiation into macrophages and foam cells through differential involvement of CXCR4–CXCR7

    PubMed Central

    Chatterjee, M; von Ungern-Sternberg, S N I; Seizer, P; Schlegel, F; Büttcher, M; Sindhu, N A; Müller, S; Mack, A; Gawaz, M

    2015-01-01

    Platelets store and release CXCL12 (SDF-1), which governs differentiation of hematopoietic progenitors into either endothelial or macrophage-foam cells. CXCL12 ligates CXCR4 and CXCR7 and regulates monocyte/macrophage functions. This study deciphers the relative contribution of CXCR4–CXCR7 in mediating the effects of platelet-derived CXCL12 on monocyte function, survival, and differentiation. CXCL12 and macrophage migration inhibitory factor (MIF) that ligate CXCR4–CXCR7 induced a dynamic bidirectional trafficking of the receptors, causing CXCR4 internalization and CXCR7 externalization during chemotaxis, thereby influencing relative receptor availability, unlike MCP-1. In vivo we found enhanced accumulation of platelets and platelet-macrophage co-aggregates in peritoneal fluid following induction of peritonitis in mice. The relative surface expression of CXCL12, CXCR4, and CXCR7 among infiltrated monocytes was also enhanced as compared with peripheral blood. Platelet-derived CXCL12 from collagen-adherent platelets and recombinant CXCL12 induced monocyte chemotaxis specifically through CXCR4 engagement. Adhesion of monocytes to immobilized CXCL12 and CXCL12-enriched activated platelet surface under static and dynamic arterial flow conditions were mediated primarily through CXCR7 and were counter-regulated by neutralizing platelet-derived CXCL12. Monocytes and culture-derived-M1–M2 macrophages phagocytosed platelets, with the phagocytic potential of culture-derived-M1 macrophages higher than M2 involving CXCR4–CXCR7 participation. CXCR7 was the primary receptor in promoting monocyte survival as exerted by platelet-derived CXCL12 against BH3-mimetic induced apoptosis (phosphatidylserine exposure, caspase-3 activation, loss of mitochondrial transmembrane potential). In co-culture experiments with platelets, monocytes predominantly differentiated into CD163+ macrophages, which was attenuated upon CXCL12 neutralization and CXCR4/CXCR7 blocking antibodies

  16. CXCL8/IL-8 and CXCL12/SDF-1α Co-operatively Promote Invasiveness and Angiogenesis in Pancreatic Cancer

    PubMed Central

    Matsuo, Yoichi; Ochi, Nobuo; Sawai, Hirozumi; Yasuda, Akira; Takahashi, Hiroki; Funahashi, Hitoshi; Takeyama, Hiromitsu; Tong, Zhimin; Guha, Sushovan

    2009-01-01

    CXC-chemokines are involved in the chemotaxis of neutrophils, lymphocytes and monocytes. However, role of these chemokines in tumorigenesis, especially with regard to interaction between tumor and its microenvironment, has not been clearly elucidated. The purpose of this study was to analyze the co-operative role of CXCL8 and CXCL12 in the tumor-stromal interaction in pancreatic cancer (PaCa). Using ELISA and RT-PCR, we initially confirmed the expression of ligands and receptors, respectively, of CXC-chemokines in PaCa and stromal cells. We examined the co-operative role of CXCL8 and CXCL12 in proliferation/invasion of PaCa and human umbilical vein endothelial cells (HUVECs), and in HUVEC tube-formations through tumor-stromal interaction by MTS, Matrigel invasion, and angiogenesis assays, respectively. We detected expression of CXCR4, but not CXCR2, in all PaCa cells and fibroblasts. PaCa cells secreted CXCL8, and fibroblast cells secreted CXCL12. CXCL8 production in PaCa was significantly enhanced by CXCL12, and CXCL12 production in fibroblasts was significantly enhanced by co-culturing with PaCa. CXCL8 enhanced proliferation/invasion of HUVECs but did not promote proliferation/invasion of PaCa. Both recombinant and PaCa-derived CXCL8 enhanced tube formation of HUVECs that were co-cultured with fibroblast cells. CXCL12 enhanced the proliferation/invasion of HUVECs and the invasion of PaCa cells but had no effect on tube formation of HUVEC. We showed that PaCa-derived CXCL8 and fibroblast-derived CXCL12 cooperatively induced angiogenesis in vitro by promoting HUVEC proliferation, invasion, and tube formation. Thus, corresponding receptors CXCR2 and CXCR4 are potential antiangiogenic and antimetastatic therapeutic targets in PaCa. PMID:19035451

  17. CXCR4 and CXCL12 (SDF-1) in prostate cancer: inhibitory effects of human single chain Fv antibodies.

    PubMed

    Vaday, Gayle G; Hua, Shao-Bing; Peehl, Donna M; Pauling, Michelle H; Lin, Yu-Huei; Zhu, Li; Lawrence, Diana M; Foda, Hussein D; Zucker, Stanley

    2004-08-15

    Metastasis is a major cause of morbidity in prostate cancer (PCa). Several studies have shown that the chemokine receptor CXCR4 and its ligand, CXCL12 (stromal cell-derived factor-1), regulate tumor cell metastasis to specific organs. Recently, it was demonstrated that CXCL12 enhances PCa cell adhesion, migration, and invasion, implicating CXCR4 in PCa metastasis. In this study, we examined the inhibitory effects of anti-CXCR4 antibodies on CXCL12-mediated PCa cell activities. We developed fully human single chain Fv antibodies (scFv), Ab124 and Ab125, against CXCR4 using the yeast two-hybrid system. We performed immunofluorescent staining, flow cytometry, and ELISA-binding assays to measure scFv binding to PCa cells. We also examined the effects of scFv on CXCL12-mediated calcium mobilization, cell migration, and invasion. Our results confirmed that PCa cell lines express cell-surface CXCR4. Real-time quantitative reverse transcription-PCR and immunohistochemical staining also verified that CXCR4 is expressed in primary cultures of prostate epithelial cells from adenocarcinomas and in human prostate tissues. Ab124 and Ab125 demonstrated specific binding to PCa cell lines by flow cytometry and in binding assays. Preincubation with scFv resulted in significant reduction of CXCL12-induced calcium mobilization in PC-3 and LNCaP cells. Ab124 and Ab125 also inhibited PCa cell migration toward CXCL12, as well as invasion through extracellular matrix gels. Ab124 and Ab125 inhibit CXCL12-mediated cellular activities by binding the receptor CXCR4. Recombinant scFv are an efficient mode of targeting tumor antigens. Considering the high incidence of PCa, the development of fully human scFv may be a useful therapeutic approach in the prevention and treatment of PCa metastasis.

  18. CXCR4/CXCL12 signaling impacts enamel progenitor cell proliferation and motility in the dental stem cell niche

    PubMed Central

    Otsu, Keishi; Harada, Hidemitsu; Shibata, Shunichi; Obara, Nobuko; Irie, Kazuharu; Taniguchi, Akiyoshi; Nagasawa, Takashi; Aoki, Kazunari; Caliari, Steven R.; Weisgerber, Daniel W.

    2015-01-01

    Dental stem cells are located at the proximal ends of rodent incisors. These stem cells reside in the dental epithelial stem cell niche, termed the apical bud. We focused on identifying critical features of a chemotactic signal in the niche. Here, we report that CXCR4/CXCL12 signaling impacts enamel progenitor cell proliferation and motility in dental stem cell niche cells. We report cells in the apical bud express CXCR4 mRNA at high levels while expression is restricted in the basal epithelium (BE) and transit-amplifying (TA) cell regions. Furthermore, the CXCL12 ligand is present in mesenchymal cells adjacent to the apical bud. We then performed gain- and loss-of-function analyses to better elucidate the role of CXCR4 and CXCL12. CXCR4-deficient mice contain epithelial cell aggregates, while cell proliferation in mutant incisors was also significantly reduced. We demonstrate in vitro that dental epithelial cells migrate toward sources of CXCL12, whereas knocking down CXCR4 impaired motility and resulted in formation of dense cell colonies. These results suggest that CXCR4 expression may be critical for activation of enamel progenitor cell division and that CXCR4/CXCL12 signaling may control movement of epithelial progenitors from the dental stem cell niche. PMID:26246398

  19. Relationship of the Chemokine, CXCL12, to Effects of Dietary Fat on Feeding-Related Behaviors and Hypothalamic Neuropeptide Systems

    PubMed Central

    Poon, Kinning; Barson, Jessica R.; Ho, Hui T.; Leibowitz, Sarah F.

    2016-01-01

    The intake of a high fat diet (HFD), in addition to stimulating orexigenic neuropeptides in the hypothalamus while promoting overeating and reducing locomotor behavior, is known to increase inflammatory mediators that modulate neuronal systems in the brain. To understand the involvement of chemokines in the effects of a HFD, we examined in rats whether HFD intake affects a specific chemokine, CXCL12, and its receptors, CXCR4 and CXCR7, in the hypothalamus together with the neuropeptides and whether CXCL12 itself acts similarly to a HFD in stimulating the neuropeptides and altering ingestion and locomotor behavior. Compared to low-fat chow, a HFD for 5 days significantly increased the expression of CXCL12 and its receptors, in both the paraventricular nucleus (PVN) where the neuropeptides enkephalin (ENK) and galanin were also stimulated and the perifornical lateral hypothalamus (PFLH) where orexin (OX) and melanin-concentrating hormone (MCH) were increased. In contrast, the HFD had no impact on expression of CXCL12 or its receptors in the arcuate nucleus (ARC) where the carbohydrate-related peptide, neuropeptide Y (NPY), was suppressed. Analysis of protein levels revealed a similar stimulatory effect of a HFD on CXCL12 levels in the PVN and PFLH, as well as in blood, and an increase in the number of CXCR4-positive cells in the PVN. In the ARC, in contrast, levels of CXCL12 and number of CXCR4-positive cells were too low to measure. When centrally administered, CXCL12 was found to have similar effects to a HFD. Injection of CXCL12 into the third cerebral ventricle immediately anterior to the hypothalamus significantly stimulated the ingestion of a HFD, reduced novelty-induced locomotor activity, and increased expression of ENK in the PVN where the CXCR4 receptors were dense. It had no impact, however, on NPY in the ARC or on OX and MCH in the PFLH where the CXCR4 receptors were not detected. These results, showing CXCL12 in the hypothalamus to be stimulated by a HFD

  20. Thymoquinone Inhibits the CXCL12-Induced Chemotaxis of Multiple Myeloma Cells and Increases Their Susceptibility to Fas-Mediated Apoptosis

    PubMed Central

    Badr, Gamal; Lefevre, Eric A.; Mohany, Mohamed

    2011-01-01

    In multiple myeloma (MM), malignant plasma cells reside in the bone marrow, where they accumulate in close contact with stromal cells. The mechanisms responsible for the chemotaxis of malignant plasma cells are still poorly understood. Thus, we investigated the mechanisms involved in the chemotaxis of MDN and XG2 MM cell lines. Both cell lines strongly expressed CCR9, CXCR3 and CXCR4 chemokine receptors but only migrated toward CXCL12. Activation of CXCR4 by CXCL12 resulted in the association of CXCR4 with CD45 and activation of PLCβ3, AKT, RhoA, IκBα and ERK1/2. Using siRNA-silencing techniques, we showed CD45/CXCR4 association is essential for CXCL12-induced migration of MM cells. Thymoquinone (TQ), the major active component of the medicinal herb Nigella sativa Linn, has been described as a chemopreventive and chemotherapeutic compound. TQ treatment strongly inhibited CXCL12-mediated chemotaxis in MM cell lines as well as primary cells isolated from MM patients, but not normal PBMCs. Moreover, TQ significantly down-regulated CXCR4 expression and CXCL12-mediated CXCR4/CD45 association in MM cells. Finally, TQ also induced the relocalization of cytoplasmic Fas/CD95 to the membrane of MM cells and increased CD95-mediated apoptosis by 80%. In conclusion, we demonstrate the potent anti-myeloma activity of TQ, providing a rationale for further clinical evaluation. PMID:21912642

  1. CXCL12/CXCR4 chemokine signaling in spinal glia induces pain hypersensitivity through MAPKs-mediated neuroinflammation in bone cancer rats.

    PubMed

    Hu, Xue-Ming; Liu, Yan-Nan; Zhang, Hai-Long; Cao, Shou-Bin; Zhang, Ting; Chen, Li-Ping; Shen, Wen

    2015-02-01

    The activation of MAPK pathways in spinal cord and subsequent production of proinflammatory cytokines in glial cells contribute to the development of spinal central sensitization, the basic mechanism underlying bone cancer pain (BCP). Our previous study showed that spinal CXCL12 from astrocytes mediates BCP generation by binding to CXCR4 in both astrocyters and microglia. Here, we verified that CXCL12/CXCR4 signaling contributed to BCP through a MAPK-mediated mechanism. In naïve rats, a single intrathecal administration of CXCL12 considerably induced pain hyperalgesia and phosphorylation expression of spinal MAPK members (including extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase), which could be partially prevented by pre-treatment with CXCR4 inhibitor AMD3100. This CXCL12-induced hyperalgesia was also reduced by MAPK inhibitors. In bone cancer rats, tumor cell inoculation into the tibial cavity caused prominent and persistent pain hyperalgesia, and associated with up-regulation of CXCL12 and CXCR4, activation of glial cells, phosphorylation of MAPKs, and production of proinflammatory cytokines in the spinal cord. These tumor cell inoculation-induced behavioral and neurochemical alterations were all suppressed by blocking CXCL12/CXCR4 signaling or MAPK pathways. Taken together, these results demonstrate that spinal MAPK pathways mediated CXCL12/CXCR4-induced pain hypersensitivity in bone cancer rats, which could be druggable targets for alleviating BCP and glia-derived neuroinflammation. Following tumor cell inoculation, chemokine CXCL12 from astrocytes spreads around the spinal environment, resulting in functional activation of CXCR4-expressing astrocytes and microglia. Once glia are activated, they may initiate MAPK (mitogen-activated protein kinase) pathways, and subsequently produce proinflammatory cytokines and chemokines. Among them, CXCL12 could reinforce the astrocytic and microglial activation in autocrine and paracrine manners

  2. Targeting the CXCR4/CXCL12 axis in treating epithelial ovarian cancer.

    PubMed

    Mao, T L; Fan, K F; Liu, C L

    2017-09-07

    Ovarian carcinoma is the most crucial and difficult target for available therapeutic treatments among gynecological malignancies, and great efforts are required to find an effective solution. Molecular studies showed that the chemokine stromal cell-derived factor-1 (also known as CXCL12) and its receptor, CXCR4, are key determinants of tumor initiation, progression and metastasis in ovarian carcinomas. Hence, it is generally believed that blocking the CXCR4/CXCL12 pathway could serve as a potential therapy for patients with ovarian cancer. Herein, we investigated the role of the CXCR4/CXCL12 axis in regulating ovarian cancer progression. Using flow cytometry, a real-time PCR and western blot analyses, we showed that the chemokine receptor CXCR4 protein and mRNA were overexpressed in human epithelial ovarian cancer cell lines, and these were closely correlated with poor outcomes. Moreover, silencing CXCR4 by small hairpin RNA in HTB75 cells reduced cell proliferation, migration and invasion and significantly reduced RhoA and Rac-1/Cdc42 expressions, whereas overexpression of CXCR4 in SKOV3 cells significantly increased cell migration and markedly increased RhoA, Rac-1/Cdc42 levels. Silencing CXCR4 also led to decreased in vitro cytotoxicity of AMD3100, a specific antagonist of CXCR4, which exerts its effect upon CXCR4 expression. Remarkably, knockdown of CXCR4 in HTB75 cells led to a significantly decreased capability to form tumors in vivo, and the Ki67 proliferation index of xenograft tumors showed a dramatic reduction. Our results revealed that the CXCR4/CXCL12 pathway represents a promising therapeutic target for epithelial ovarian carcinoma.Gene Therapy advance online publication, 7 September 2017; doi:10.1038/gt.2017.69.

  3. C-terminal engineering of CXCL12 and CCL5 chemokines: functional characterization by electrophysiological recordings.

    PubMed

    Picciocchi, Antoine; Siaučiūnaiteė-Gaubard, Lina; Petit-Hartlein, Isabelle; Sadir, Rabia; Revilloud, Jean; Caro, Lydia; Vivaudou, Michel; Fieschi, Franck; Moreau, Christophe; Vivès, Corinne

    2014-01-01

    Chemokines are chemotactic cytokines comprised of 70-100 amino acids. The chemokines CXCL12 and CCL5 are the endogenous ligands of the CXCR4 and CCR5 G protein-coupled receptors that are also HIV co-receptors. Biochemical, structural and functional studies of receptors are ligand-consuming and the cost of commercial chemokines hinders their use in such studies. Here, we describe methods for the expression, refolding, purification, and functional characterization of CXCL12 and CCL5 constructs incorporating C-terminal epitope tags. The model tags used were hexahistidines and Strep-Tag for affinity purification, and the double lanthanoid binding tag for fluorescence imaging and crystal structure resolution. The ability of modified and purified chemokines to bind and activate CXCR4 and CCR5 receptors was tested in Xenopus oocytes expressing the receptors, together with a Kir3 G-protein activated K(+) channel that served as a reporter of receptor activation. Results demonstrate that tags greatly influence the biochemical properties of the recombinant chemokines. Besides, despite the absence of any evidence for CXCL12 or CCL5 C-terminus involvement in receptor binding and activation, we demonstrated unpredictable effects of tag insertion on the ligand apparent affinity and efficacy or on the ligand dissociation. These tagged chemokines should constitute useful tools for the selective purification of properly-folded chemokines receptors and the study of their native quaternary structures.

  4. CXCL12/CXCR4 Axis Improves Migration of Neuroblasts Along Corpus Callosum by Stimulating MMP-2 Secretion After Traumatic Brain Injury in Rats.

    PubMed

    Mao, Weifeng; Yi, Xin; Qin, Jianbing; Tian, Meiling; Jin, Guohua

    2016-06-01

    To investigate the effect of CXCL12 on migration of neural precursor cells after traumatic brain injury (TBI). We randomly divided 48 rats into four groups: (1) the sham group, rats were performed craniotomy only, (2) the control group, saline were injected into the ipsilateral cortex after TBI, (3) the CXCL12 group, CXCL12 were injected into the ipsilateral cortex after TBI, and (4) the CXCL12 + AMD3100 group, CXCL12 and AMD3100 were mixed together and injected into the ipsilateral cortex after TBI. At 7 days after TBI, the brain tissues were subjected to immunofluorescent double-labeled staining with the antibodies of CXCR4/DCX, MMP-2/DCX, MMP-2/GFAP, MMP-2/NeuN. Western blot assay was used to measure the protein levels of MMP-2. Compared with the control group, the number of CXCR4/DCX and MMP-2 positive cells around the injured corpus callosum area were significantly increased in the CXCL12 treatment group. The area occupied by these cells expanded and the shape changed from chain distribution to radial. CXCL12 + AMD3100 treatment significantly decreased the number and distribution area of CXCR4/DCX and MMP-2 positive cells compared with the CXCL12 treatment and control group. The DCX positive cells could not form chain or radial distribution. The protein expressions of MMP-2 had the similar change trends as the results of immunofluorescent staining. MMP-2 could be secreted by DCX, GFAP and NeuN positive cells. CXCL12/CXCR4 axis can improve the migration of the neuroblasts along the corpus callosum by stimulating the MMP-2 secretion of different types of cells.

  5. Src family kinases involved in CXCL12-induced loss of acute morphine analgesia.

    PubMed

    Rivat, Cyril; Sebaihi, Soumia; Van Steenwinckel, Juliette; Fouquet, Stéphane; Kitabgi, Patrick; Pohl, Michel; Melik Parsadaniantz, Stéphane; Reaux-Le Goazigo, Annabelle

    2014-05-01

    Functional interactions between the chemokine receptor CXCR4 and opioid receptors have been reported in the brain, leading to a decreased morphine analgesic activity. However the cellular mechanisms responsible for this loss of opioid analgesia are largely unknown. Here we examined whether Src family-kinases (SFK)-linked mechanisms induced by CXCR4 contributed to the loss of acute morphine analgesia and could represent a new physiological anti-opioid signaling pathway. In this way, we showed by immunohistochemistry and western blot that CXCL12 rapidly activated SFK phosphorylation in vitro in primary cultured lumbar rat dorsal root ganglia (DRG) but also in vivo in the DRG and the spinal cord. We showed that SFK activation occurred in a sub population of sensory neurons, in spinal microglia but also in spinal nerve terminals expressing mu-(MOR) and delta-opioid (DOR) receptor. In addition we described that CXCR4 is detected in MOR- and DOR-immunoreactive neurons in the DRG and spinal cord. In vivo, we demonstrated that an intrathecal administration of CXCL12 (1μg) significantly attenuated the subcutaneous morphine (4mg/kg) analgesia. Conversely, pretreatment with a potent CXCR4 antagonist (5μg) significantly enhanced morphine analgesia. Similar effects were obtained after an intrathecal injection of a specific SFK inhibitor, PP2 (10μg). Furthermore, PP2 abrogated CXCL12-induced decrease in morphine analgesia by suppressing SFK activation in the spinal cord. In conclusion, our data highlight that CXCL12-induced loss of acute morphine analgesia is linked to Src family kinases activation.

  6. Molecular Pathways: Targeting the CXCR4-CXCL12 Axis--Untapped Potential in the Tumor Microenvironment.

    PubMed

    Scala, Stefania

    2015-10-01

    Evidence suggests that the CXC-chemokine receptor-4 pathway plays a role in cancer cell homing and metastasis, and thus represents a potential target for cancer therapy. The homeostatic microenvironment chemokine CXCL12 binds the CXCR4 and CXCR7 receptors, activating divergent signals on multiple pathways, such as ERK1/2, p38, SAPK/JNK, AKT, mTOR, and the Bruton tyrosine kinase (BTK). An activating mutation in CXCR4 is responsible for a rare disease, WHIM syndrome (warts, hypogammaglobulinemia, infections, and myelokathexis), and dominant CXCR4 mutations have also been reported in Waldenstrom macroglobulinemia. The CXCR4-CXCL12 axis regulates the hematopoietic stem cell niche--a property that has led to the approval of the CXCR4 antagonist plerixafor (AMD3100) for mobilization of hematopoietic precursors. In preclinical models, plerixafor has shown antimetastatic potential in vivo, offering proof of concept. Other antagonists are in preclinical and clinical development. Recent evidence demonstrates that inhibiting CXCR4 signaling restores sensitivity to CTLA-4 and PD-1 checkpoint inhibitors, creating a new line for investigation. Targeting the CXCR4-CXCL12 axis thus offers the possibility of affecting CXCR4-expressing primary tumor cells, modulating the immune response, or synergizing with other targeted anticancer therapies. ©2015 American Association for Cancer Research.

  7. CXCL12/CXCR4 Axis Activation Mediates Prostate Myofibroblast Phenoconversion through Non-Canonical EGFR/MEK/ERK Signaling

    PubMed Central

    Rodríguez-Nieves, José A.; Patalano, Susan C.; Almanza, Diego; Gharaee-Kermani, Mehrnaz; Macoska, Jill A.

    2016-01-01

    Benign prostate hyperplasia (BPH), an enlargement of the prostate common in aging in men, is associated with urinary voiding dysfunction manifest as Lower Urinary Tract Symptoms (LUTS). Although inflammation and abnormal smooth muscle contractions are known to play key roles in the development of LUTS, tissue fibrosis may also be an important and previously unrecognized contributing factor. Tissue fibrosis arises from the unregulated differentiation of fibroblasts or other precursor cell types into myofibroblasts, which is usually accomplished by activation of the TGFβ/TGFβR axis. Previously we reported that the CXC-type chemokines, CXCL5, CXCL8 and CXCL12, which are up-regulated in the aging in the prostate, can drive this differentiation process as well in the absence of TGFβ. Based on this data we sought to elucidate the molecular mechanisms employed by CXCL12, and its receptor CXCR4, during prostate myofibroblast phenoconversion. The results of these studies suggest that CXCL12/CXCR4-mediated signaling events in prostate myofibroblast phenoconversion may proceed through non-canonical pathways that do not depend on TGFβ/TGFβR axis activation or Smad signaling. Here we report that CXCL12/CXCR4 axis activation promotes signaling through the EGFR and downstream MEK/ERK and PI3K/Akt pathways during myofibroblast phenoconversion, but not through TGFβ/TGFβR and downstream Smad signaling, in prostate fibroblasts undergoing myofibroblast phenoconversion. We document that EGFR transactivation is required for CXCL12-mediated signaling and expression of genes associate with myofibroblast phenoconversion (α-SMA, COL1a1). Our study successfully identified TGFβ/TGFβR-independent molecular mechanisms that promote CXCL12/CXCR4-induced myofibroblast phenoconversion. This information may be crucial for the development of novel therapies and potential biomarkers for prostatic fibrosis. PMID:27434301

  8. CXCL12/CXCR4 Axis Activation Mediates Prostate Myofibroblast Phenoconversion through Non-Canonical EGFR/MEK/ERK Signaling.

    PubMed

    Rodríguez-Nieves, José A; Patalano, Susan C; Almanza, Diego; Gharaee-Kermani, Mehrnaz; Macoska, Jill A

    2016-01-01

    Benign prostate hyperplasia (BPH), an enlargement of the prostate common in aging in men, is associated with urinary voiding dysfunction manifest as Lower Urinary Tract Symptoms (LUTS). Although inflammation and abnormal smooth muscle contractions are known to play key roles in the development of LUTS, tissue fibrosis may also be an important and previously unrecognized contributing factor. Tissue fibrosis arises from the unregulated differentiation of fibroblasts or other precursor cell types into myofibroblasts, which is usually accomplished by activation of the TGFβ/TGFβR axis. Previously we reported that the CXC-type chemokines, CXCL5, CXCL8 and CXCL12, which are up-regulated in the aging in the prostate, can drive this differentiation process as well in the absence of TGFβ. Based on this data we sought to elucidate the molecular mechanisms employed by CXCL12, and its receptor CXCR4, during prostate myofibroblast phenoconversion. The results of these studies suggest that CXCL12/CXCR4-mediated signaling events in prostate myofibroblast phenoconversion may proceed through non-canonical pathways that do not depend on TGFβ/TGFβR axis activation or Smad signaling. Here we report that CXCL12/CXCR4 axis activation promotes signaling through the EGFR and downstream MEK/ERK and PI3K/Akt pathways during myofibroblast phenoconversion, but not through TGFβ/TGFβR and downstream Smad signaling, in prostate fibroblasts undergoing myofibroblast phenoconversion. We document that EGFR transactivation is required for CXCL12-mediated signaling and expression of genes associate with myofibroblast phenoconversion (α-SMA, COL1a1). Our study successfully identified TGFβ/TGFβR-independent molecular mechanisms that promote CXCL12/CXCR4-induced myofibroblast phenoconversion. This information may be crucial for the development of novel therapies and potential biomarkers for prostatic fibrosis.

  9. Intersection of Chemokine and TSH Receptor Pathways in Human Fibrocytes: Emergence of CXCL-12/CXCR4 Cross Talk Potentially Relevant to Thyroid-Associated Ophthalmopathy.

    PubMed

    Fernando, Roshini; Atkins, Stephen J; Smith, Terry J

    2016-10-01

    Fibrocytes are monocyte progenitor cells that have been implicated in normal and pathological tissue remodeling. Among the prominent chemokine receptors expressed by these cells is CXC motif receptor 4 (CXCR4), which, with its cognate ligand CXCL motif ligand 12 (CXCL-12), directs fibrocytes to sites of fibrosis. Fibrocytes have been implicated in the pathogenesis of thyroid-associated ophthalmopathy, the ocular manifestation of Graves' disease (GD), by virtue of their unique accumulation as CD34(+) orbital fibroblasts (OFs). Fibrocytes also express high levels of functional TSH receptor (TSHR). Here, we determined CXCL-12 and CXCR4 expression in fibrocytes and GD-OF and whether that pathway interacts with TSHR. CXCL-12 is highly expressed in GD-OF, whereas CXCR4 levels are dramatically higher in fibrocytes. Levels of these proteins are differentially regulated by TSH in a cell type-specific manner. Further, CXCL-12 enhances the induction by TSH of IL-6 in fibrocytes but attenuates this induction in GD-OF. In contrast, in pure CD34(+) OF, the interplay between TSH and CXCL-12 reverts to that observed in fibrocytes. Our results indicate that CXCL-12 enhances TSH actions in fibrocytes but inhibits them in GD-OF, a dichotomy imposed by factors emanating from CD34(-) OF. They also suggest a potentially important modulatory role for CD34(-) OF in determining the factors that influence pathological TSHR signaling in the TAO orbit.

  10. A role for the CXCL12 receptor, CXCR7, in the pathogenesis of human pulmonary vascular disease.

    PubMed

    Costello, Christine M; McCullagh, Brian; Howell, Katherine; Sands, Michelle; Belperio, John A; Keane, Michael P; Gaine, Sean; McLoughlin, Paul

    2012-06-01

    Given the critical role that endothelial cell dysfunction plays in the pathogenesis of pulmonary hypertensive diseases, we set out to establish if CXCR7, a receptor for the pro-angiogenic ligand CXCL12, is expressed in the vasculature of human lung diseases and examine its role in mediating CXCL12-induced responses in primary pulmonary human microvascular endothelial cells. Receptor and ligand expression was examined in control and explanted human hypertensive lungs, in human plasma and in hypoxic rodent lungs, by ELISA and immunohistochemical studies. Functional in vitro experiments examined the role of CXCR7 in CXCL12-induced lung microvascular endothelial cell proliferation, migration, and wound regeneration and repair. CXCR7 is elevated in the endothelium of explanted human hypertensive lungs and circulating CXCL12 concentrations are significantly elevated in disease. We demonstrate that alveolar hypoxia similar to that found in lung disease increases CXCR7 expression in the pulmonary endothelium. Furthermore, CXCR7 is the receptor through which endothelial cell regeneration and repair, and proliferation, is mediated, whereas signalling via CXCR4 is essential for chemotactic cell migration. Our findings demonstrate that CXCR7 has a critical but previously unrecognised role to play in endothelial cell proliferation, suggesting that CXCR7-mediated signalling may be functionally important in pulmonary vascular diseases.

  11. Multiple roles of chemokine CXCL12 in the central nervous system: A migration from immunology to neurobiology

    PubMed Central

    Li, Meizhang; Ransohoff, Richard M.

    2008-01-01

    Chemotactic cytokines (chemokines) have been traditionally defined as small (10–14 kDa) secreted leukocyte chemoattractants. However, chemokines and their cognate receptors are constitutively expressed in the central nervous system (CNS) where immune activities are under stringent control. Why and how the CNS uses the chemokine system to carry out its complex physiological functions has intrigued neurobiologists. Here, we focus on chemokine CXCL12 and its receptor CXCR4 that have been widely characterized in peripheral tissues and delineate their main functions in the CNS. Extensive evidence supports CXCL12 as a key regulator for early development of the CNS. CXCR4 signaling is required for the migration of neuronal precursors, axon guidance/pathfinding and maintenance of neural progenitor cells (NPCs). In the mature CNS, CXCL12 modulates neurotransmission, neurotoxicity and neuroglial interactions. Thus, chemokines represent an inherent system that helps establish and maintain CNS homeostasis. In addition, growing evidence implicates altered expression of CXCL12 and CXCR4 in the pathogenesis of CNS disorders such as HIV-associated encephalopathy, brain tumor, stroke and multiple sclerosis (MS), making them the plausible targets for future pharmacological intervention. PMID:18177992

  12. The chemokine receptors CXCR4/CXCR7 and their primary heterodimeric ligands CXCL12 and CXCL12/high mobility group box 1 in pancreatic cancer growth and development: finding flow.

    PubMed

    Shakir, Murtaza; Tang, Daolin; Zeh, Herbert J; Tang, Siu Wah; Anderson, Carolyn J; Bahary, Nathan; Lotze, Michael T

    2015-05-01

    Novel therapies need to be developed for patients with pancreatic cancer because of the poor outcomes of current regimens. Pancreatic cancer cells respond to the C-X-C chemokine receptor type 4 (CXCR4)/C-X-C chemokine receptor type 7 (CXCR7)/C-X-C motif chemokine 12 (CXCL12)/high-mobility group box 1 signaling axis and this axis presents a novel target for therapy. C-X-C motif chemokine 12 stimulates CXCR4/CXCR7-bearing cells in a paracrine manner. C-X-C chemokine receptor type 4 and CXCR7 are transmembrane G protein-coupled receptors that, upon interaction with ligand CXCL12, activate downstream protein kinases that promote a more aggressive behavior. C-X-C chemokine receptor type 4 is expressed on most pancreatic cancer cells, whereas CXCR7 is primarily expressed on tumor-associated endothelium. High-mobility group box 1 promotes the CXCR4 and CXCL12 interaction, promoting angiogenesis and lymphangiogenesis. Hypoxia-inducible factor 1 is a potent stimulator of CXCR4 and CXCL12 expression, promoting more aggressive behavior. This receptor/ligand interaction can be disrupted by CXCR4 antagonists available and in clinical use to harvest bone marrow stem cells. Novel imaging strategies are now being developed at several centers to evaluate response to therapy and identify early recurrence. Thus, the CXCR4/CXCR7/CXCL12 interaction plays a critical role in cancer cell progression, proliferation, invasion, as well as metastasis and is a suitable target for therapy, imaging, as well as development of novel diagnostics.

  13. CXCL12 prolongs naive CD4+ T lymphocytes survival via activation of PKA, CREB and Bcl2 and BclXl up-regulation.

    PubMed

    Vitiello, Laura; Ferraro, Elisabetta; De Simone, Salvatore; Gatta, Lucia; Feraco, Alessandra; Racioppi, Luigi; Rosano, Giuseppe

    2016-12-01

    Naive T lymphocytes recirculate through the body, traveling from secondary lymphoid organs through tissues and via lymphatic vessels and peripheral blood into other secondary lymphoid organs and into the bone marrow. In these tissues, lymphocytes are exposed to the chemokine CXCL12 which is abundantly produced in bone marrow and in lymph nodes by stromal cells. CXCL12 is known to drive lymphocytes chemotaxis and, in cells types such as stem cells, an antiapopototic effect has been described. Here we analyzed the effect of CXCL12 exposure on naïve CD4+ T lymphocytes purified from peripheral blood by immunomagnetic negative isolation and cultured in a nutrient poor medium. We also studied, mainly by western blot analysis, the signaling pathways involved in CXCL12 action on naïve CD4+ T lymphocytes. We found that CXCL12-exposed cells survived longer than untreated ones and this prolonged lifespan was specific for resting naïve lymphocytes, while in vitro activated lymphoblasts died rapidly despite CXCL12 treatment. We demonstrated that the increased percentage of living cells observed upon CXCL12 administration was not due to induction of proliferation but to a prosurvival effect of this chemokine. Moreover, our data suggest that this prosurvival effect on naïve CD4+ T lymphocytes might likely be mediated by PKA-dependent CREB activation and consequent increased expression of the antiapoptotic factors Bcl2 and BclXl. This newly reported activity of CXCL12 might contribute to the maintenance of the naïve T lymphocytes pool in vivo, which is needed to ensure a proper immune response to new antigens. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  14. Impact of a CXCL12/CXCR4 Antagonist in Bleomycin (BLM) Induced Pulmonary Fibrosis and Carbon Tetrachloride (CCl4) Induced Hepatic Fibrosis in Mice

    PubMed Central

    Chow, Leola N.; Schreiner, Petra; Ng, Betina Y. Y.; Lo, Bernard; Hughes, Michael R.; Scott, R. Wilder; Gusti, Vionarica; Lecour, Samantha; Simonson, Eric; Manisali, Irina; Barta, Ingrid; McNagny, Kelly M.; Crawford, Jason; Webb, Murray; Underhill, T. Michael

    2016-01-01

    Modulation of chemokine CXCL12 and its receptor CXCR4 has been implicated in attenuation of bleomycin (BLM)-induced pulmonary fibrosis and carbon tetrachloride (CCl4)-induced hepatic injury. In pulmonary fibrosis, published reports suggest that collagen production in the injured lung is derived from fibrocytes recruited from the circulation in response to release of pulmonary CXCL12. Conversely, in hepatic fibrosis, resident hepatic stellate cells (HSC), the key cell type in progression of fibrosis, upregulate CXCR4 expression in response to activation. Further, CXCL12 induces HSC proliferation and subsequent production of collagen I. In the current study, we evaluated AMD070, an orally bioavailable inhibitor of CXCL12/CXCR4 in alleviating BLM-induced pulmonary and CCl4-induced hepatic fibrosis in mice. Similar to other CXCR4 antagonists, treatment with AMD070 significantly increased leukocyte mobilization. However, in these two models of fibrosis, AMD070 had a negligible impact on extracellular matrix deposition. Interestingly, our results indicated that CXCL12/CXCR4 signaling has a role in improving mortality associated with BLM induced pulmonary injury, likely through dampening an early inflammatory response and/or vascular leakage. Together, these findings indicate that the CXCL12-CXCR4 signaling axis is not an effective target for reducing fibrosis. PMID:26998906

  15. Impact of a CXCL12/CXCR4 Antagonist in Bleomycin (BLM) Induced Pulmonary Fibrosis and Carbon Tetrachloride (CCl4) Induced Hepatic Fibrosis in Mice.

    PubMed

    Chow, Leola N; Schreiner, Petra; Ng, Betina Y Y; Lo, Bernard; Hughes, Michael R; Scott, R Wilder; Gusti, Vionarica; Lecour, Samantha; Simonson, Eric; Manisali, Irina; Barta, Ingrid; McNagny, Kelly M; Crawford, Jason; Webb, Murray; Underhill, T Michael

    2016-01-01

    Modulation of chemokine CXCL12 and its receptor CXCR4 has been implicated in attenuation of bleomycin (BLM)-induced pulmonary fibrosis and carbon tetrachloride (CCl4)-induced hepatic injury. In pulmonary fibrosis, published reports suggest that collagen production in the injured lung is derived from fibrocytes recruited from the circulation in response to release of pulmonary CXCL12. Conversely, in hepatic fibrosis, resident hepatic stellate cells (HSC), the key cell type in progression of fibrosis, upregulate CXCR4 expression in response to activation. Further, CXCL12 induces HSC proliferation and subsequent production of collagen I. In the current study, we evaluated AMD070, an orally bioavailable inhibitor of CXCL12/CXCR4 in alleviating BLM-induced pulmonary and CCl4-induced hepatic fibrosis in mice. Similar to other CXCR4 antagonists, treatment with AMD070 significantly increased leukocyte mobilization. However, in these two models of fibrosis, AMD070 had a negligible impact on extracellular matrix deposition. Interestingly, our results indicated that CXCL12/CXCR4 signaling has a role in improving mortality associated with BLM induced pulmonary injury, likely through dampening an early inflammatory response and/or vascular leakage. Together, these findings indicate that the CXCL12-CXCR4 signaling axis is not an effective target for reducing fibrosis.

  16. The role of the CXCL12-CXCR4/CXCR7 axis in the progression and metastasis of bone sarcomas (Review).

    PubMed

    Liao, Yu-Xin; Zhou, Cheng-Hao; Zeng, Hui; Zuo, Dong-Qing; Wang, Zhuo-Ying; Yin, Fei; Hua, Ying-Qing; Cai, Zheng-Dong

    2013-12-01

    Bone sarcomas, which comprise less than 1% of all human malignancies, are a group of relatively rare mesenchymal-derived tumors. They are mainly composed of osteosarcoma, chondrosarcoma and Ewing's sarcoma. In spite of advances in adjuvant chemotherapy and wide surgical resection, prognosis remains poor due to the high propensity for lung metastasis, which is the leading cause of mortality in patients with bone sarcomas. Chemokines are a superfamily of small pro-inflammatory chemoattractant cytokines which can bind to specific G protein-coupled seven-span transmembrane receptors. Chemokine 12 (CXCL12), also designated as stromal cell-derived factor-1 (SDF-1), is able to bind to its cognate receptors, chemokine receptor 4 (CXCR4) and chemokine receptor 7 (CXCR7), with high affinity. The binding of CXCL12 to CXCR4/CXCR7 stimulates the activation of several downstream signaling pathways that regulate tumor progression and metastasis. In this review, the structure and function of CXCL12 and its receptors, CXCR4 and CXCR7, as well as many factors affecting their expression are discussed. Phosphoinositide 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways are the two most important downstream pathways regulated by the CXCL12-CXCR4/CXCR7 interaction. CXCR4 expression in bone sarcomas, including tumor cells and samples and the correlation between CXCR4/CXCR7 expression and the survival of patients with bone sarcomas are also discussed. In addition, we review the involvement of the CXCL12‑CXCR4/CXCR7 axis in the growth and metastasis of bone sarcomas and the targeting of this axis in preclinical studies.

  17. Fragment-Based Optimization of Small Molecule CXCL12 Inhibitors for Antagonizing the CXCL12/CXCR4 Interaction

    PubMed Central

    Ziarek, Joshua J.; Liu, Yan; Smith, Emmanuel; Zhang, Guolin; Peterson, Francis C.; Chen, Jun; Yu, Yongping; Chen, Yu; Volkman, Brian F.; Li, Rongshi

    2013-01-01

    The chemokine CXCL12 and its G protein-coupled receptor (GPCR) CXCR4 are high-priority clinical targets because of their involvement in metastatic cancers (also implicated in autoimmune disease and cardiovascular disease). Because chemokines interact with two distinct sites to bind and activate their receptors, both the GPCRs and chemokines are potential targets for small molecule inhibition. A number of chemokines have been validated as targets for drug development, but virtually all drug discovery efforts focus on the GPCRs. However, all CXCR4 receptor antagonists with the exception of MSX-122 have failed in clinical trials due to unmanageable toxicities, emphasizing the need for alternative strategies to interfere with CXCL12/CXCR4-guided metastatic homing. Although targeting the relatively featureless surface of CXCL12 was presumed to be challenging, focusing efforts at the sulfotyrosine (sY) binding pockets proved successful for procuring initial hits. Using a hybrid structure-based in silico/NMR screening strategy, we recently identified a ligand that occludes the receptor recognition site. From this initial hit, we designed a small fragment library containing only nine tetrazole derivatives using a fragment-based and bioisostere approach to target the sY binding sites of CXCL12. Compound binding modes and affinities were studied by 2D NMR spectroscopy, X-ray crystallography, molecular docking and cell-based functional assays. Our results demonstrate that the sY binding sites are conducive to the development of high affinity inhibitors with better ligand efficiency (LE) than typical protein-protein interaction inhibitors (LE ≤ 0.24). Our novel tetrazole-based fragment 18 was identified to bind the sY21 site with a Kd of 24 μM (LE = 0.30). Optimization of 18 yielded compound 25 which specifically inhibits CXCL12-induced migration with an improvement in potency over the initial hit 9. The fragment from this library that exhibited the highest affinity and

  18. Baclofen and other GABAB receptor agents are allosteric modulators of the CXCL12 chemokine receptor CXCR4.

    PubMed

    Guyon, Alice; Kussrow, Amanda; Olmsted, Ian Roys; Sandoz, Guillaume; Bornhop, Darryl J; Nahon, Jean-Louis

    2013-07-10

    CXCR4, a receptor for the chemokine CXCL12 (stromal-cell derived factor-1α), is a G-protein-coupled receptor (GPCR), expressed in the immune and CNS and integrally involved in various neurological disorders. The GABAB receptor is also a GPCR that mediates metabotropic action of the inhibitory neurotransmitter GABA and is located on neurons and immune cells as well. Using diverse approaches, we report novel interaction between GABAB receptor agents and CXCR4 and demonstrate allosteric binding of these agents to CXCR4. First, both GABAB antagonists and agonists block CXCL12-elicited chemotaxis in human breast cancer cells. Second, a GABAB antagonist blocks the potentiation by CXCL12 of high-threshold Ca(2+) channels in rat neurons. Third, electrophysiology in Xenopus oocytes and human embryonic kidney cell line 293 cells in which we coexpressed rat CXCR4 and the G-protein inward rectifier K(+) (GIRK) channel showed that GABAB antagonist and agonist modified CXCL12-evoked activation of GIRK channels. To investigate whether GABAB ligands bind to CXCR4, we expressed this receptor in heterologous systems lacking GABAB receptors and performed competition binding experiments. Our fluorescent resonance energy transfer experiments suggest that GABAB ligands do not bind CXCR4 at the CXCL12 binding pocket suggesting allosteric modulation, in accordance with our electrophysiology experiments. Finally, using backscattering interferometry and lipoparticles containing only the CXCR4 receptor, we quantified the binding affinity for the GABAB ligands, confirming a direct interaction with the CXCR4 receptor. The effect of GABAergic agents on CXCR4 suggests new therapeutic potentials for neurological and immune diseases.

  19. Survival and Proliferation of Neural Progenitor-Derived Glioblastomas Under Hypoxic Stress is Controlled by a CXCL12/CXCR4 Autocrine-Positive Feedback Mechanism.

    PubMed

    Calinescu, Anda-Alexandra; Yadav, Viveka Nand; Carballo, Erica; Kadiyala, Padma; Tran, Dustin; Zamler, Daniel B; Doherty, Robert; Srikanth, Maithreyi; Lowenstein, Pedro Ricardo; Castro, Maria Graciela

    2017-03-01

    Purpose: One likely cause of treatment failure in glioblastoma is the persistence of glioma stem-like cells (GSLCs) which are highly resistant to therapies currently employed. We found that CXCL12 has highest expression in glioma cells derived from neural progenitor cells (NPC). The development and molecular signature of NPC-derived glioblastomas were analyzed and the therapeutic effect of blocking CXCL12 was tested.Experimental Design: Tumors were induced by injecting DNA into the lateral ventricle of neonatal mice, using the Sleeping Beauty transposase method. Histology and expression of GSLC markers were analyzed during disease progression. Survival upon treatment with pharmacologic (plerixafor) or genetic inhibition of CXCR4 was analyzed. Primary neurospheres were generated and analyzed for proliferation, apoptosis, and expression of proteins regulating survival and cell-cycle progression.Results: Tumors induced from NPCs display histologic features of human glioblastoma and express markers of GSLC. In vivo, inhibiting the CXCL12/CXCR4 signaling axis results in increased survival of tumor-bearing animals. In vitro, CXCR4 blockade induces apoptosis and inhibits cell-cycle progression, downregulates molecules regulating survival and proliferation, and also blocks the hypoxic induction of HIF-1α and CXCL12. Exogenous administration of CXCL12 rescues the drug-induced decrease in proliferation.Conclusions: This study demonstrates that the CXCL12/CXCR4 axis operates in glioblastoma cells under hypoxic stress via an autocrine-positive feedback mechanism, which promotes survival and cell-cycle progression. Our study brings new mechanistic insight and encourages further exploration of the use of drugs blocking CXCL12 as adjuvant agents to target hypoxia-induced glioblastoma progression, prevent resistance to treatment, and recurrence of the disease. Clin Cancer Res; 23(5); 1250-62. ©2016 AACR. ©2016 American Association for Cancer Research.

  20. Phosphorylation of Collapsin Response Mediator Protein 2 on Tyr-479 Regulates CXCL12-induced T Lymphocyte Migration*S⃞

    PubMed Central

    Varrin-Doyer, Michel; Vincent, Peggy; Cavagna, Sylvie; Auvergnon, Nathalie; Noraz, Nelly; Rogemond, Véronique; Honnorat, Jérôme; Moradi-Améli, Mahnaz; Giraudon, Pascale

    2009-01-01

    In the central nervous system, collapsin response mediator protein 2 (CRMP2) is a transducer protein that supports the semaphorin-induced guidance of axons toward their cognate target. However, we previously showed that CRMP2 is also expressed in immune cells and plays a crucial role in T lymphocyte migration. Here we further investigated the molecular mechanisms underlying CRMP2 function in chemokine-directed T-cell motility. Examining Jurkat T-cells treated with the chemokine CXCL12, we found that 1) CXCL12 induces a dynamic re-localization of CRMP2 to uropod, the flexible structure of migrating lymphocyte, and increases its binding to the cytoskeletal protein vimentin; 2) CXCL12 decreases phosphorylation of the glycogen synthase kinase-3β-targeted residues CRMP2-Thr-509/514; and 3) tyrosine Tyr-479 is a new phosphorylation CRMP2 residue and a target for the Src-family kinase Yes. Moreover, phospho-Tyr-479 increased under CXCL12 signaling while phospho-Thr-509/514 decreased. The functional importance of this tyrosine phosphorylation was demonstrated by Y479F mutation that strongly reduced CXCL12-mediated T-cell polarization and motility as tested in a transmigration model and on neural tissue. We propose that differential phosphorylation by glycogen synthase kinase-3β and Yes modulates the contribution of CRMP2 to cytoskeletal reorganization during chemokine-directed T-cell migration. In addition to providing a novel mechanism for T lymphocyte motility, our findings reveal CRMP2 as a transducer of chemokine signaling. PMID:19276087

  1. Adult mouse subventricular zones stimulate glioblastoma stem cells specific invasion through CXCL12/CXCR4 signaling

    PubMed Central

    Goffart, Nicolas; Kroonen, Jérôme; Di Valentin, Emmanuel; Dedobbeleer, Matthias; Denne, Alexandre; Martinive, Philippe; Rogister, Bernard

    2015-01-01

    Background Patients with glioblastoma multiforme (GBM) have an overall median survival of 15 months. This catastrophic survival rate is the consequence of systematic relapses that could arise from remaining glioblastoma stem cells (GSCs) left behind after surgery. We previously demonstrated that GSCs are able to escape the tumor mass and specifically colonize the adult subventricular zones (SVZs) after transplantation. This specific localization, away from the initial injection site, therefore represents a high-quality model of a clinical obstacle to therapy and relapses because GSCs notably retain the ability to form secondary tumors. Method In this work, we questioned the role of the CXCL12/CXCR4 signaling in the GSC-specific invasion of the SVZs. Results We demonstrated that both receptor and ligand are respectively expressed by different GBM cell populations and by the SVZ itself. In vitro migration bio-assays highlighted that human U87MG GSCs isolated from the SVZs (U87MG-SVZ) display stronger migratory abilities in response to recombinant CXCL12 and/or SVZ-conditioned medium (SVZ-CM) compared with cancer cells isolated from the tumor mass (U87MG-TM). Moreover, in vitro inhibition of the CXCR4 signaling significantly decreased the U87MG-SVZ cell migration in response to the SVZ-CM. Very interestingly, treating U87MG-xenografted mice with daily doses of AMD3100, a specific CXCR4 antagonist, prevented the specific invasion of the SVZ. Another in vivo experiment, using CXCR4-invalidated GBM cells, displayed similar results. Conclusion Taken together, these data demonstrate the significant role of the CXCL12/CXCR4 signaling in this original model of brain cancer invasion. PMID:25085362

  2. Administration of a CXCL12 Analog in Endotoxemia Is Associated with Anti-Inflammatory, Anti-Oxidative and Cytoprotective Effects In Vivo

    PubMed Central

    2015-01-01

    Background The chemokine receptor CXCR4 is a multifunctional receptor which is activated by its natural ligand C-X-C motif chemokine 12 (CXCL12). As CXCR4 is part of the lipopolysaccharide sensing complex and CXCL12 analogs are not well characterized in inflammation, we aimed to uncover the systemic effects of a CXCL12 analog in severe systemic inflammation and to evaluate its impact on endotoxin induced organ damages by using a sublethal LPS dose. Methods The plasma stable CXCL12 analog CTCE-0214D was synthesized and administered subcutaneously shortly before LPS treatment. After 24 hours, mice were sacrificed and blood was obtained for TNF alpha, IFN gamma and blood glucose evaluation. Oxidative stress in the liver and spleen was assessed and liver biotransformation capacity was determined. Finally, CXCR4, CXCL12 and TLR4 expression patterns in liver, spleen and thymus tissue as well as the presence of different markers for apoptosis and oxidative stress were determined by means of immunohistochemistry. Results CTCE-0214D distinctly reduced the LPS mediated effects on TNF alpha, IFN gamma, ALAT and blood glucose levels. It attenuated oxidative stress in the liver and spleen tissue and enhanced liver biotransformation capacity unambiguously. Furthermore, in all three organs investigated, CTCE-0214D diminished the LPS induced expression of CXCR4, CXCL12, TLR4, NF-κB, cleaved caspase-3 and gp91 phox, whereas heme oxygenase 1 expression and activity was induced above average. Additionally, TUNEL staining revealed anti-apoptotic effects of CTCE-0214D. Conclusions In summary, CTCE-0214D displayed anti-inflammatory, anti-oxidative and cytoprotective features. It attenuated reactive oxygen species, induced heme oxygenase 1 activity and mitigated apoptosis. Thus, the CXCR4/CXCL12 axis seems to be a promising target in the treatment of acute systemic inflammation, especially when accompanied by a hepatic dysfunction and an excessive production of free radicals. PMID

  3. Complement C3a enhances CXCL12 (SDF-1)-mediated chemotaxis of bone marrow hematopoietic cells independently of C3a receptor.

    PubMed

    Honczarenko, Marek; Ratajczak, Mariusz Z; Nicholson-Weller, Anne; Silberstein, Leslie E

    2005-09-15

    Complement C3a promotes CXCL12-induced migration and engraftment of human and murine hemopoietic progenitor cells, suggesting a cross-influence between anaphylatoxin and chemokine axes. Here we have explored the underlying mechanism(s) of complement anaphylatoxin and chemokine cooperation. In addition to C3a, C3a-desArg and C4a but not C5a, are potent enhancers of CXCL12-induced chemotaxis of human and murine bone marrow (BM) stem/progenitor cells and B lineage cells. C3a enhancement of chemotaxis is chemokine specific because it is also observed for chemotaxis to CCL19 but not to CXCL13. The potentiating effect of C3a on CXCL12 is independent of the classical C3a receptor (C3aR). First, human BM CD34(+) and B lineage cells do not express C3aR by flow cytometry. Second, the competitive C3aR inhibitor SB290157 does not affect C3a-mediated enhancement of CXCL12-induced chemotaxis. Third, enhancement of chemotaxis of hemopoietic cells is also mediated by C3a-desArg, which does not bind to C3aR. Finally, C3a enhances CXCL12-induced chemotaxis of BM cells from C3aR knockout mice similar to BM cells from wild-type mice. Subsequent studies revealed that C3a increased the binding affinity of CXCL12 to human CXCR4(+)/C3aR(-), REH pro-B cells, which is compatible with a direct interaction between C3a and CXCL12. BM stromal cells were able to generate C3a, C3a-desArg, C4a, as well as CXCL12, suggesting that this pathway could function in vivo. Taken together, we demonstrate a C3a-CXCL12 interaction independent of the C3aR, which may provide a mechanism to modulate the function of CXCL12 in the BM microenvironment.

  4. Inhibition of CXCL12-mediated chemotaxis of Jurkat cells by direct immunotoxicants.

    PubMed

    Shao, Jia; Stout, Inge; Volger, Oscar L; Hendriksen, Peter J M; van Loveren, Henk; Peijnenburg, Ad A C M

    2016-07-01

    Directional migration of cells to specific locations is required in tissue development, wound healing, and immune responses. Immune cell migration plays a crucial role in both innate and adaptive immunity. Chemokines are small pro-inflammatory chemoattractants that control the migration of leukocytes. In addition, they are also involved in other immune processes such as lymphocyte development and immune pathology. In a previous toxicogenomics study using the Jurkat T cell line, we have shown that the model immunotoxicant TBTO inhibited chemotaxis toward the chemokine CXCL12. In the present work, we aimed at assessing a novel approach to detecting chemicals that affect the process of cell migration. For this, we first evaluated the effects of 31 chemicals on mRNA expression of genes that are known to be related to cell migration. With this analysis, seven immunotoxicants were identified as potential chemotaxis modulators, of which five (CoCl2 80 µM, MeHg 1 µM, ochratoxin A 10 µM, S9-treated ochratoxin A 10 µM, and TBTO 100 nM) were confirmed as chemotaxis inhibitor in an in vitro trans-well chemotaxis assay using the chemokine CXCL12. The transcriptome data of the five compounds together with previously obtained protein phosphorylation profiles for two out of five compounds (i.e., ochratoxin A and TBTO) revealed that the mechanisms behind the chemotaxis inhibition are different for these immunotoxicants. Moreover, the mTOR inhibitor rapamycin had no effect on the chemotaxis of Jurkat cells, indicating that the mTOR pathway is not involved in CXCL12-mediated chemotaxis of Jurkat cells, which is opposite to the findings on human primary T cells (Munk et al. in PLoS One 6(9):e24667, 2011). Thus, the results obtained from the chemotaxis assay conducted with Jurkat cells might not fully represent the results obtained with human primary T cells. Despite this difference, the present study indicated that some compounds may exert their immunotoxic effects through

  5. Modulating the interaction of CXCR4 and CXCL12 by low-molecular-weight heparin inhibits hepatic metastasis of colon cancer.

    PubMed

    Ma, Lixin; Qiao, Haiquan; He, Changjun; Yang, Qian; Cheung, Chun Hei Antonio; Kanwar, Jagat R; Sun, Xueying

    2012-04-01

    Liver metastasis is the major obstacle for prolonging the survival of colon cancer patients. Low-molecular-weight heparin (LMWH), a common drug for venous thromboembolism, has displayed beneficial effects in improving the survival of cancer patients, though the mechanism remains unclear. This study aimed to investigate the effects of LMWH on hepatic metastasis of colon cancer and its underlying molecular mechanism by targeting the interaction of the chemokine receptor CXCR4 and its ligand CXCL12 (formerly known as stromal cell-derived factor 1α, SDF-1α), as the CXCR4-CXCL12 axis has been shown to regulate the interaction of cancer cells and stroma. Experimental results revealed that LMWH (Enoxaparin, 3500-5500 Da) inhibited the CXCL12-stimulated proliferation, adhesion and colony formation of human colon cancer HCT-116 cells that highly expressed CXCR4. Interestingly, LMWH or an anti-CXCR4 blocking antibody diminished the migrating and invading abilities of HCT116 cells stimulated by the recombinant CXCL12 protein or liver homogenates which contained endogenous CXCL12 protein. Although LMWH did not significantly inhibit the growth of subcutaneous colon tumors, it significantly suppressed the formation of hepatic metastasis established by intrasplenic injection of colon cancer cells in nude Balb/c mice and also downregulated the expression of CXCL12 in hepatic sinusoidal endothelial cells. The results suggest that LMWH inhibits the formation of hepatic metastasis of colon cancer by disrupting the interaction of CXCR4 and CXCL12, supporting that perioperative administration of LMWH may help to prevent the seeding and subsequent growth of hepatic metastases of colon cancer cells.

  6. MicroRNA-135b suppresses extravillous trophoblast-derived HTR-8/SVneo cell invasion by directly down regulating CXCL12 under low oxygen conditions.

    PubMed

    Tamaru, Shunsuke; Mizuno, Yosuke; Tochigi, Hideno; Kajihara, Takeshi; Okazaki, Yasushi; Okagaki, Ryugo; Kamei, Yoshimasa; Ishihara, Osamu; Itakura, Atsuo

    2015-05-29

    The expression of numerous microRNAs (miRNAs) in the trophoblasts changes under low oxygen conditions. However, little is known regarding the regulation of the trophoblast invasion by miRNAs under low oxygen conditions. The aim of this study was to identify those miRNAs and their target genes associated with the trophoblast invasion under low oxygen conditions. Culturing the extravillous trophoblast (EVT) cell line, HTR-8/SVneo, at 2% oxygen as compared to 20% oxygen suppressed trophoblast invasion that correlated with increased expression of microRNA-135b (miR-135b) and decreased expression of the its predicted target gene CXCL12. Overexpression of miR-135b suppressed CXCL12 mRNA expression and invasion of HTR-8/SVneo cells. Adding a neutralizing antibody against CXCL12 to the culture medium suppressed HTR-8/SVneo cell invasion. Reporter assays showed that the 3'UTR sequence of CXCL12 was directly targeted by miR-135b. Our results suggest that miR-135b and CXCL12 play important roles in modulating the EVT invasion under low oxygen conditions.

  7. Reelin and CXCL12 regulate distinct migratory behaviors during the development of the dopaminergic system.

    PubMed

    Bodea, Gabriela Oana; Spille, Jan-Hendrik; Abe, Philipp; Andersson, Aycan Senturk; Acker-Palmer, Amparo; Stumm, Ralf; Kubitscheck, Ulrich; Blaess, Sandra

    2014-02-01

    The proper functioning of the dopaminergic system requires the coordinated formation of projections extending from dopaminergic neurons in the substantia nigra (SN), ventral tegmental area (VTA) and retrorubral field to a wide array of forebrain targets including the striatum, nucleus accumbens and prefrontal cortex. The mechanisms controlling the assembly of these distinct dopaminergic cell clusters are not well understood. Here, we have investigated in detail the migratory behavior of dopaminergic neurons giving rise to either the SN or the medial VTA using genetic inducible fate mapping, ultramicroscopy, time-lapse imaging, slice culture and analysis of mouse mutants. We demonstrate that neurons destined for the SN migrate first radially and then tangentially, whereas neurons destined for the medial VTA undergo primarily radial migration. We show that tangentially migrating dopaminergic neurons express the components of the reelin signaling pathway, whereas dopaminergic neurons in their initial, radial migration phase express CXC chemokine receptor 4 (CXCR4), the receptor for the chemokine CXC motif ligand 12 (CXCL12). Perturbation of reelin signaling interferes with the speed and orientation of tangentially, but not radially, migrating dopaminergic neurons and results in severe defects in the formation of the SN. By contrast, CXCR4/CXCL12 signaling modulates the initial migration of dopaminergic neurons. With this study, we provide the first molecular and functional characterization of the distinct migratory pathways taken by dopaminergic neurons destined for SN and VTA, and uncover mechanisms that regulate different migratory behaviors of dopaminergic neurons.

  8. Recurrence of glioblastoma after radio-chemotherapy is associated with an angiogenic switch to the CXCL12-CXCR4 pathway.

    PubMed

    Tabouret, Emeline; Tchoghandjian, Aurelie; Denicolai, Emilie; Delfino, Christine; Metellus, Philippe; Graillon, Thomas; Boucard, Celine; Nanni, Isabelle; Padovani, Laetitia; Ouafik, L'Houcine; Figarella-Branger, Dominique; Chinot, Olivier

    2015-05-10

    Angiogenesis is one of the key features of glioblastoma (GBM). Our objective was to explore the potential changes of angiogenic factors in GBM between initial diagnosis and recurrence after radiotherapy-temozolomide (RT/TMZ). Paired frozen tumors from both initial and recurrent surgery were available for 29 patients. Screening of genes expressions related to angiogenesis was performed using RT- PCR arrays on 10 first patients. Next, RNA expressions of the selected genes were analyzed on all samples. Protein expression was examined by immunohistochemistry. The anti-tumor effect of AMD3100 (anti-CXCR4) was tested in GBM explants. In the screening step, the initial-recurrence expression changes contributed to a selection of seven genes (VEGFA, VEGFR2, VEGFR1, CXCL12, CXCR4, uPA HIF1α). By quantitative RT-PCR, RNA expressions of CXCR4 (p = 0.029) and CXCL12 (p = 0.107) were increased while expressions of HIF1α (p = 0.009) and VEGFR2 (p = 0.081) were decreased at recurrence. Similarly, CXCL12 protein expression tended to increase (p = 0.096) while VEGFR2 staining was decreased (p = 0.004) at recurrence. An increase of anti-tumoral effect was observed with the combination of AMD3100 and RT/TMZ versus RT/TMZ alone in GB explants. Recurrence of GB after chemo-radiation could be associated with a switch of angiogenic pattern from VEGFR2-HIF1α to CXCL12-CXCR4 pathway, leading to new perspectives in angiogenic treatment.

  9. CXCL12 Regulates through JAK1 and JAK2 Formation of Productive Immunological Synapses.

    PubMed

    Cascio, Graciela; Martín-Cófreces, Noa B; Rodríguez-Frade, José Miguel; López-Cotarelo, Pilar; Criado, Gabriel; Pablos, José L; Rodríguez-Fernández, José Luis; Sánchez-Madrid, Francisco; Mellado, Mario

    2015-06-01

    The adaptive immune response requires interaction between T cells and APC to form a specialized structure termed the immune synapse (IS). Although the TCR is essential for IS organization, other factors such as chemokines participate in this process. In this study, we show that the chemokine CXCL12-mediated signaling contributes to correct IS organization and therefore influences T cell activation. CXCR4 downregulation or blockade on T cells caused defective actin polymerization at the contact site with APC, altered microtubule-organizing center polarization and the IS structure, and reduced T cell/APC contact duration. T cell activation was thus inhibited, as shown by reduced expression of CD25 and CD69 markers and of IL-2 mRNA levels. The results indicate that, through Gi and JAK1 and 2 kinases activation, CXCL12 signaling cooperates to build the IS and to maintain adhesive contacts between APC and T cells, required for continuous TCR signaling. Copyright © 2015 by The American Association of Immunologists, Inc.

  10. The CXCL12/CXCR4/CXCR7 ligand-receptor system regulates neuro-glio-vascular interactions and vessel growth during human brain development.

    PubMed

    Virgintino, Daniela; Errede, Mariella; Rizzi, Marco; Girolamo, Francesco; Strippoli, Maurizio; Wälchli, Thomas; Robertson, David; Frei, Karl; Roncali, Luisa

    2013-05-01

    This study investigates glio-vascular interactions in human fetal brain at midgestation, specifically examining the expression and immunolocalization of the CXCL12/CXCR4/CXCR7 ligand-receptor axis and its possible role in the vascular patterning of the developing brain. At midgestation, the telencephalic vesicles are characterized by well developed radial glia cells (RGCs), the first differentiated astrocytes and a basic vascular network mainly built of radial vessels. RGCs have been recognized to contribute to cerebral cortex neuro-vascular architecture and have also been demonstrated to act as a significant source of neural cells (Rakic, Brain Res 33:471-476, 1971; Malatesta et al, Development 127:5253-5263, 2000). According to our hypothesis CXCL12, a potent migration and differentiation chemokine released by RGCs, may act as a linking factor coordinating neuroblast migration with vessel growth and patterning through the activation of different ligand/receptor axes. The obtained results support this hypothesis showing that together with CXCR4/CXCR7-reactive neuroblasts, which migrate in close association with CXCL12 RGCs, layer-specific subsets of CXCL12 RGCs and astrocytes specifically contact the microvessel wall. Moreover, the CXCL12/CXCR4/CXCR7 system appears to be directly involved in microvessel growth, its members being differentially expressed in angiogenically activated microvessels and vascular sprouts.

  11. CXCL12/CXCR4 axis induced miR-125b promotes invasion and confers 5-fluorouracil resistance through enhancing autophagy in colorectal cancer

    PubMed Central

    Yu, Xinfeng; Shi, Wenna; Zhang, Yuhang; Wang, Xiaohui; Sun, Shiyue; Song, Zhiyu; Liu, Man; Zeng, Qiao; Cui, Shuxiang; Qu, Xianjun

    2017-01-01

    The activation of CXCL12/CXCR4 axis is associated with potential progression of cancer, such as invasion, metastasis and chemoresistance. However, the underlying mechanisms of CXCL12/CXCR4 axis and cancer progression have been poorly explored. We hypothesized that miRNAs might be critical downstream mediators of CXCL12/CXCR4 axis involved in cancer invasion and chemoresistance in CRC. In human CRC cells, we found that the activation of CXCL12/CXCR4 axis promoted epithelial-mesenchymal transition (EMT) and concurrent upregulation of miR-125b. Overexpression of miR-125b robustly triggered EMT and cancer invasion, which in turn enhanced the expression of CXCR4. Importantly, the reciprocal positive feedback loop between CXCR4 and miR-125b further activated the Wnt/β-catenin signaling by targeting Adenomatous polyposis coli (APC) gene. There was a negative correlation of the expression of miR-125b with APC mRNA in paired human colorectal tissue specimens. Further experiments indicated a role of miR-125b in conferring 5-fluorouracil (5-FU) resistance in CRC probably through increasing autophagy both in vitro and in vivo. MiR-125b functions as an important downstream mediator upon the activation of CXCL12/CXCR4 axis that involved in EMT, invasion and 5-FU resistance of CRC. These findings shed a new insight into the role of miR-125b and provide a potential therapeutic target in CRC. PMID:28176874

  12. CXCL12/CXCR4 axis induced miR-125b promotes invasion and confers 5-fluorouracil resistance through enhancing autophagy in colorectal cancer.

    PubMed

    Yu, Xinfeng; Shi, Wenna; Zhang, Yuhang; Wang, Xiaohui; Sun, Shiyue; Song, Zhiyu; Liu, Man; Zeng, Qiao; Cui, Shuxiang; Qu, Xianjun

    2017-02-08

    The activation of CXCL12/CXCR4 axis is associated with potential progression of cancer, such as invasion, metastasis and chemoresistance. However, the underlying mechanisms of CXCL12/CXCR4 axis and cancer progression have been poorly explored. We hypothesized that miRNAs might be critical downstream mediators of CXCL12/CXCR4 axis involved in cancer invasion and chemoresistance in CRC. In human CRC cells, we found that the activation of CXCL12/CXCR4 axis promoted epithelial-mesenchymal transition (EMT) and concurrent upregulation of miR-125b. Overexpression of miR-125b robustly triggered EMT and cancer invasion, which in turn enhanced the expression of CXCR4. Importantly, the reciprocal positive feedback loop between CXCR4 and miR-125b further activated the Wnt/β-catenin signaling by targeting Adenomatous polyposis coli (APC) gene. There was a negative correlation of the expression of miR-125b with APC mRNA in paired human colorectal tissue specimens. Further experiments indicated a role of miR-125b in conferring 5-fluorouracil (5-FU) resistance in CRC probably through increasing autophagy both in vitro and in vivo. MiR-125b functions as an important downstream mediator upon the activation of CXCL12/CXCR4 axis that involved in EMT, invasion and 5-FU resistance of CRC. These findings shed a new insight into the role of miR-125b and provide a potential therapeutic target in CRC.

  13. Multiple myeloma cells recruit tumor-supportive macrophages through the CXCR4/CXCL12 axis and promote their polarization toward the M2 phenotype.

    PubMed

    Beider, Katia; Bitner, Hanna; Leiba, Merav; Gutwein, Odit; Koren-Michowitz, Maya; Ostrovsky, Olga; Abraham, Michal; Wald, Hanna; Galun, Eithan; Peled, Amnon; Nagler, Arnon

    2014-11-30

    Multiple myeloma (MM) cells specifically attract peripheral-blood monocytes, while interaction of MM with bone marrow stromal cells (BMSCs) significantly increased monocyte recruitment (p<0.01). The CXCL12 chemokine, produced by both the MM and BMSCs, was found to be a critical regulator of monocyte migration. CXCL12 production was up-regulated under MM-BMSCs co-culture conditions, whereas blockage with anti-CXCR4 antibodies significantly abrogated monocyte recruitment toward a MM-derived conditioned medium (p<0.01). Furthermore, elevated levels of CXCL12 were detected in MM, but not in normal BM samples, whereas malignant MM cells often represented the source of increased CXCL12 in the BM. Blood-derived macrophages effectively supported MM cells proliferation and protected them from chemotherapy-induced apoptosis. Importantly, MM cells affected macrophage polarization, elevating the expression of M2-related scavenger receptor CD206 in macrophages and blocking LPS-induced TNFα secretion (a hallmark of M1 response). Of note, MM-educated macrophages suppressed T-cell proliferation and IFNγ production in response to activation. Finally, increased numbers of CXCR4-expressing CD163+CD206+ macrophages were detected in the BM of MM patients (n=25) in comparison to MGUS (n=11) and normal specimens (n=8). Taken together, these results identify macrophages as important players in MM tumorogenicity, and recognize the CXCR4/CXCL12 axis as a critical regulator of MM-stroma interactions and microenvironment formation.

  14. Multiple myeloma cells recruit tumor-supportive macrophages through the CXCR4/CXCL12 axis and promote their polarization toward the M2 phenotype

    PubMed Central

    Beider, Katia; Bitner, Hanna; Leiba, Merav; Gutwein, Odit; Koren-Michowitz, Maya; Ostrovsky, Olga; Abraham, Michal; Wald, Hanna; Galun, Eithan; Peled, Amnon; Nagler, Arnon

    2014-01-01

    Multiple myeloma (MM) cells specifically attract peripheral-blood monocytes, while interaction of MM with bone marrow stromal cells (BMSCs) significantly increased monocyte recruitment (p<0.01). The CXCL12 chemokine, produced by both the MM and BMSCs, was found to be a critical regulator of monocyte migration. CXCL12 production was up-regulated under MM-BMSCs co-culture conditions, whereas blockage with anti-CXCR4 antibodies significantly abrogated monocyte recruitment toward a MM-derived conditioned medium (p<0.01). Furthermore, elevated levels of CXCL12 were detected in MM, but not in normal BM samples, whereas malignant MM cells often represented the source of increased CXCL12 in the BM. Blood-derived macrophages effectively supported MM cells proliferation and protected them from chemotherapy-induced apoptosis. Importantly, MM cells affected macrophage polarization, elevating the expression of M2-related scavenger receptor CD206 in macrophages and blocking LPS-induced TNFα secretion (a hallmark of M1 response). Of note, MM-educated macrophages suppressed T-cell proliferation and IFNγ production in response to activation. Finally, increased numbers of CXCR4-expressing CD163+CD206+ macrophages were detected in the BM of MM patients (n=25) in comparison to MGUS (n=11) and normal specimens (n=8). Taken together, these results identify macrophages as important players in MM tumorogenicity, and recognize the CXCR4/CXCL12 axis as a critical regulator of MM-stroma interactions and microenvironment formation. PMID:25526031

  15. Cxcl12/Cxcr4 chemokine signaling is required for placode assembly and sensory axon pathfinding in the zebrafish olfactory system.

    PubMed

    Miyasaka, Nobuhiko; Knaut, Holger; Yoshihara, Yoshihiro

    2007-07-01

    Positioning neurons in the right places and wiring axons to the appropriate targets are essential events for establishment of neural circuits. In the zebrafish olfactory system, precursors of olfactory sensory neurons (OSNs) assemble into a compact cluster to form the olfactory placode. Subsequently, OSNs differentiate and extend their axons to the presumptive olfactory bulb with high precision. In this study, we aim to elucidate the molecular mechanism underlying these two developmental processes. cxcr4b, encoding a chemokine receptor, is expressed in the migrating olfactory placodal precursors, and cxcl12a (SDF-1a), encoding a ligand for Cxcr4b, is expressed in the abutting anterior neural plate. The expression of cxcr4b persists in the olfactory placode at the initial phase of OSN axon pathfinding. At this time, cxcl12a is expressed along the placode-telencephalon border and at the anterior tip of the telencephalon, prefiguring the route and target of OSN axons, respectively. Interfering with Cxcl12a/Cxcr4b signaling perturbs the assembly of the olfactory placode, resulting in the appearance of ventrally displaced olfactory neurons. Moreover, OSN axons frequently fail to exit the olfactory placode and accumulate near the placode-telencephalon border in the absence of Cxcr4b-mediated signaling. These data indicate that chemokine signaling contributes to both the olfactory placode assembly and the OSN axon pathfinding in zebrafish.

  16. The Sphingosine-1-Phosphate Modulator FTY720 Targets Multiple Myeloma via the CXCR4/CXCL12 Pathway.

    PubMed

    Beider, Katia; Rosenberg, Evgenia; Bitner, Hanna; Shimoni, Avichai; Leiba, Merav; Koren-Michowitz, Maya; Ribakovsky, Elena; Klein, Shiri; Olam, Devorah; Weiss, Lola; Wald, Hanna; Abraham, Michal; Galun, Eithan; Peled, Amnon; Nagler, Arnon

    2017-04-01

    Purpose: To explore the functional consequences of possible cross-talk between the CXCR4/CXCL12 and the sphingosine-1-phosphate (S1P) pathways in multiple myeloma (MM) cells and to evaluate the effect of S1P targeting with the FTY720 modulator as a potential anti-MM therapeutic strategy.Experimental Design and Results: S1P targeting with FTY720 induces MM cell apoptosis. The combination of FTY720 with the SPHK1 inhibitor SKI-II results in synergistic inhibition of MM growth. CXCR4/CXCL12-enhanced expression correlates with reduced MM cell sensitivity to both FTY720 and SKI-II inhibitors, and with SPHK1 coexpression in both cell lines and primary MM bone marrow (BM) samples, suggesting regulative cross-talk between the CXCR4/CXCL12 and SPHK1 pathways in MM cells. FTY720 was found to directly target CXCR4. FTY720 profoundly reduces CXCR4 cell-surface levels and abrogates the CXCR4-mediated functions of migration toward CXCL12 and signaling pathway activation. Moreover, FTY720 cooperates with bortezomib, inducing its cytotoxic activity and abrogating the bortezomib-mediated increase in CXCR4 expression. FTY720 effectively targets bortezomib-resistant cells and increases their sensitivity to bortezomib, promoting DNA damage. Finally, in a recently developed novel xenograft model of CXCR4-dependent systemic MM with BM involvement, FTY720 treatment effectively reduces tumor burden in the BM of MM-bearing mice. FTY720 in combination with bortezomib demonstrates superior tumor growth inhibition and abrogates bortezomib-induced CXCR4 increase on MM cells.Conclusions: Altogether, our work identifies a cross-talk between the S1P and CXCR4 pathways in MM cells and provides a preclinical rationale for the therapeutic application of FTY720 in combination with bortezomib in patients with MM. Clin Cancer Res; 23(7); 1733-47. ©2016 AACR. ©2016 American Association for Cancer Research.

  17. Targeting CXCL12/CXCR4 signaling with oncolytic virotherapy disrupts tumor vasculature and inhibits breast cancer metastases.

    PubMed

    Gil, Margaret; Seshadri, Mukund; Komorowski, Marcin P; Abrams, Scott I; Kozbor, Danuta

    2013-04-02

    Oncolytic viruses hold promise for the treatment of cancer, but their interaction with the tumor microenvironment needs to be elucidated for optimal tumor cell killing. Because the CXCR4 receptor for the stromal cell-derived factor-1 (SDF-1/CXCL12) chemokine is one of the key stimuli involved in signaling interactions between tumor cells and their stromal microenvironment, we used oncolytic virotherapy with a CXCR4 antagonist to target the CXCL12/CXCR4 signaling axis in a triple-negative 4T1 breast carcinoma in syngeneic mice. We show here that CXCR4 antagonist expression from an oncolytic vaccinia virus delivered intravenously to mice with orthotopic tumors attains higher intratumoral concentration than its soluble counterpart and exhibits increased efficacy over that mediated by oncolysis alone. A systemic delivery of the armed virus after resection of the primary tumor was efficacious in inhibiting the development of spontaneous metastasis and increased overall tumor-free survival. Inhibition of tumor growth with the armed virus was associated with destruction of tumor vasculature, reductions in expression of CXCL12 and VEGF, and decrease in intratumoral numbers of bone marrow-derived endothelial and myeloid cells. These changes led to induction of antitumor antibody responses and resistance to tumor rechallenge. Engineering an oncolytic virus armed with a CXCR4 antagonist represents an innovative strategy that targets multiple elements within the tumor microenvironment. As such, this approach could have a significant therapeutic impact against primary and metastatic breast cancer.

  18. Disrupted cardiac development but normal hematopoiesis in mice deficient in the second CXCL12/SDF-1 receptor, CXCR7

    PubMed Central

    Sierro, Frederic; Biben, Christine; Martínez-Muñoz, Laura; Mellado, Mario; Ransohoff, Richard M.; Li, Meizhang; Woehl, Blanche; Leung, Helen; Groom, Joanna; Batten, Marcel; Harvey, Richard P.; Martínez-A, Carlos; Mackay, Charles R.; Mackay, Fabienne

    2007-01-01

    Chemotactic cytokines (chemokines) attract immune cells, although their original evolutionary role may relate more closely with embryonic development. We noted differential expression of the chemokine receptor CXCR7 (RDC-1) on marginal zone B cells, a cell type associated with autoimmune diseases. We generated Cxcr7−/− mice but found that CXCR7 deficiency had little effect on B cell composition. However, most Cxcr7−/− mice died at birth with ventricular septal defects and semilunar heart valve malformation. Conditional deletion of Cxcr7 in endothelium, using Tie2-Cre transgenic mice, recapitulated this phenotype. Gene profiling of Cxcr7−/− heart valve leaflets revealed a defect in the expression of factors essential for valve formation, vessel protection, or endothelial cell growth and survival. We confirmed that the principal chemokine ligand for CXCR7 was CXCL12/SDF-1, which also binds CXCR4. CXCL12 did not induce signaling through CXCR7; however, CXCR7 formed functional heterodimers with CXCR4 and enhanced CXCL12-induced signaling. Our results reveal a specialized role for CXCR7 in endothelial biology and valve development and highlight the distinct developmental role of evolutionary conserved chemokine receptors such as CXCR7 and CXCR4. PMID:17804806

  19. In vitro regulation of CCL3 and CXCL12 by bacterial by-products is dependent on site of origin of human oral fibroblasts

    PubMed Central

    Sipert, Carla Renata; Morandini, Ana Carolina; Dionísio, Thiago José; de Andrade Moreira Machado, Maria Aparecida; de Oliveira, Sandra Helena Penha; Campanelli, Ana Paula; Kuo, Winston Patrick; Santos, Carlos Ferreira

    2015-01-01

    Introduction Production of chemokines by tissue resident cells is one of the main mechanisms involved in the inflammatory infiltrate formation during inflammation. The specific ability of fibroblasts from different oral tissues such as gingiva, periodontal ligament and dental pulp from permanent and deciduous teeth in producing the chemokines CCL3 and CXCL12 under stimulation by bacterial products commonly found in endodontic infections was investigated. Methods Cultures of fibroblasts from gingiva and periodontal ligament as well as from dental pulp from permanent and deciduous teeth were established by using an explant technique and stimulated with increasing concentrations of Escherichia coli lipopolysaccharide (EcLPS) and Enterococcus faecalis lipoteichoic acid (EfLTA) for 1, 6 and 24 hours. Supernatants were tested for CCL3 and CXCL12 by enzyme-linked immunosorbent assay. Results In general, CCL3 production was induced by EcLPS in the 4 fibroblast subtypes and by EfLTA in fibroblasts from gingiva and periodontal ligament. Constitutive CXCL12 synthesis decreased in all fibroblast subtypes especially under stimulation with EcLPS. Fibroblast from permanent deciduous teeth was the cell type presenting the most expressive reduction in CXCL12 release by both stimuli. On the basis of computational matching of CXCL12 mRNA with the microRNAs miR-141 and miR-200a, their expression was also investigated. Although detected in the fibroblasts, these molecules remained unaltered by bacterial by-product stimulation. Conclusions EcLPS and EfLTA induced the production of CCL3 and unbalanced the synthesis of CXCL12 in a manner dependent on the specific tissue origin of fibroblasts. PMID:24331998

  20. Mutational Analysis of Atypical Chemokine Receptor 3 (ACKR3/CXCR7) Interaction with Its Chemokine Ligands CXCL11 and CXCL12.

    PubMed

    Benredjem, Besma; Girard, Mélanie; Rhainds, David; St-Onge, Geneviève; Heveker, Nikolaus

    2017-01-06

    Atypical chemokine receptors do not mediate chemotaxis or G protein signaling, but they recruit arrestin. They also efficiently scavenge their chemokine ligands, thereby contributing to gradient maintenance and termination. ACKR3, also known as CXCR7, binds and degrades the constitutive chemokine CXCL12, which also binds the canonical receptor CXCR4, and CXCL11, which also binds CXCR3. Here we report comprehensive mutational analysis of the ACKR3 interaction with its chemokine ligands, using 30 substitution mutants. Readouts are radioligand binding competition, arrestin recruitment, and chemokine scavenging. Our results suggest different binding modes for both chemokines. CXCL11 depends on the ACKR3 N terminus and some extracellular loop (ECL) positions for primary binding, ECL residues mediate secondary binding and arrestin recruitment potency. CXCL12 binding required key residues Asp-179(4.60) and Asp-275(6.58) (residue numbering follows the Ballesteros-Weinstein scheme), with no evident involvement of N-terminal residues, suggesting an uncommon mode of receptor engagement. Mutation of residues corresponding to CRS2 in CXCR4 (positions Ser-103(2.63) and Gln-301(7.39)) increased CXCL11 binding, but reduced CXCL12 affinity. Mutant Q301E(7.39) did not recruit arrestin. Mutant K118A(3.26) in ECL1 showed moderate baseline arrestin recruitment with ablation of ligand-induced responses. Substitutions that affected CXCL11 binding also diminished scavenging. However, detection of reduced CXCL12 scavenging by mutants with impaired CXCL12 affinity required drastically reduced receptor expression levels, suggesting that scavenging pathways can be saturated and that CXCL12 binding exceeds scavenging at higher receptor expression levels. Arrestin recruitment did not correlate with scavenging; although Q301E(7.39) degraded chemokines in the absence of arrestin, S103D(2.63) had reduced CXCL11 scavenging despite intact arrestin responses. © 2017 by The American Society for

  1. Common structural interactions between the receptors CXCR3, CXCR4 and CXCR7 complexed with their natural ligands, CXCL11 and CXCL12, by a modeling approach.

    PubMed

    Costantini, Susan; Raucci, Raffaele; De Vero, Teresa; Castello, Giuseppe; Colonna, Giovanni

    2013-10-01

    Chemokine receptor trio composed by CXCR3, CXCR4 and CXCR7 represents a hard and interesting challenge for cancer biology because these three receptors are found to be over-expressed in different cancers as well as to bind the same chemokines. In fact, CXCR4 interacts with CXCL12, CXCR7 not only with CXCL12 but also with CXCL11, that is a natural ligand for CXCR3. For these reasons, it seems necessary to define and to identify the structural determinants of CXCR3, CXCR4 and CXCR7 and their related physic-chemical properties that permit them to bind CXCL11 and CXCL12. Hence in this paper we show the modeling of CXCR7 and its complex with CXCL11 and CXCL12 compared to CXCR3/CXCL11 and CXCR4/CXCL12. Our results show that (i) CXCR3, CXCR4 and CXCR7 present similar trans-membrane helices and different conformations of N-terminal and C-terminal regions as well as of three extracellular loops, and (ii) the predominant interaction between the three receptors and the two chemokines are on hydrophobic and electrostatic basis. Moreover, our data confirm that CXCL12 binds to CXCR7 with higher affinity than to CXCR4. Methodologically, we can also conclude that our computational strategy is adequate to model correctly the interactions between these chemokines and their receptors; therefore, our models represent a good structural basis to design and develop peptides able to block contemporaneously CXCR3, CXCR4 and CXCR7 receptor trio.

  2. CXCL12-mediated murine neural progenitor cell movement requires PI3Kβ activation.

    PubMed

    Holgado, Borja L; Martínez-Muñoz, Laura; Sánchez-Alcañiz, Juan Antonio; Lucas, Pilar; Pérez-García, Vicente; Pérez, Gema; Rodríguez-Frade, José Miguel; Nieto, Marta; Marín, Oscar; Carrasco, Yolanda R; Carrera, Ana C; Alvarez-Dolado, Manuel; Mellado, Mario

    2013-08-01

    The migratory route of neural progenitor/precursor cells (NPC) has a central role in central nervous system development. Although the role of the chemokine CXCL12 in NPC migration has been described, the intracellular signaling cascade involved remains largely unclear. Here we studied the molecular mechanisms that promote murine NPC migration in response to CXCL12, in vitro and ex vivo. Migration was highly dependent on signaling by the CXCL12 receptor, CXCR4. Although the JAK/STAT pathway was activated following CXCL12 stimulation of NPC, JAK activity was not necessary for NPC migration in vitro. Whereas CXCL12 activated the PI3K catalytic subunits p110α and p110β in NPC, only p110β participated in CXCL12-mediated NPC migration. Ex vivo experiments using organotypic slice cultures showed that p110β blockade impaired NPC exit from the medial ganglionic eminence. In vivo experiments using in utero electroporation nonetheless showed that p110β is dispensable for radial migration of pyramidal neurons. We conclude that PI3K p110β is activated in NPC in response to CXCL12, and its activity is necessary for immature interneuron migration to the cerebral cortex.

  3. Cellular and subcellular localization of CXCL12 and CXCR4 in rat nociceptive structures: physiological relevance.

    PubMed

    Reaux-Le Goazigo, Annabelle; Rivat, Cyril; Kitabgi, Patrick; Pohl, Michel; Melik Parsadaniantz, Stéphane

    2012-09-01

    Initial studies implicated the chemokine CXC motif ligand 12 (CXCL12) and its cognate CXC motif receptor 4 (CXCR4) in pain modulation. However, there has been no description of the distribution, transport and axonal sorting of CXCL12 and CXCR4 in rat nociceptive structures, and their direct participation in nociception modulation has not been demonstrated. Here, we report that acute intrathecal administration of CXCL12 induced mechanical hypersensitivity in naive rats. This effect was prevented by a CXCR4-neutralizing antibody. To determine the morphological basis of this behavioural response, we used light and electron microscopic immunohistochemistry to map CXCL12- and CXCR4-immunoreactive elements in dorsal root ganglia, lumbar spinal cord, sciatic nerve and skin. Light microscopy analysis revealed CXCL12 and CXCR4 immunoreactivity in calcitonin gene related peptide-containing peptidergic primary sensory neurons, which were both conveyed to central and peripheral sensory nerve terminals. Electron microscopy clearly demonstrated CXCL12 and CXCR4 immunoreactivity in primary sensory nerve terminals in the dorsal horn; both were sorted into small clear vesicles and large dense-core vesicles. This suggests that CXCL12 and CXCR4 are trafficked from nerve cell bodies to the dorsal horn. Double immunogold labelling for CXCL12 and calcitonin gene related peptide revealed partial vesicular colocalization in axonal terminals. We report, for the first time, that CXCR4 receptors are mainly located on the neuronal plasma membrane, where they are present at pre-synaptic and post-synaptic sites of central terminals. Receptor inactivation experiments, behavioural studies and morphological analyses provide strong evidence that the CXCL12/CXCR4 system is involved in modulation of nociceptive signalling.

  4. Monomeric structure of the cardioprotective chemokine SDF-1/CXCL12

    PubMed Central

    Veldkamp, Christopher T; Ziarek, Joshua J; Su, Jidong; Basnet, Harihar; Lennertz, Richard; Weiner, Joshua J; Peterson, Francis C; Baker, John E; Volkman, Brian F

    2009-01-01

    The chemokine stromal cell-derived factor-1 (SDF-1/CXCL12) directs leukocyte migration, stem cell homing, and cancer metastasis through activation of CXCR4, which is also a coreceptor for T-tropic HIV-1. Recently, SDF-1 was shown to play a protective role after myocardial infarction, and the protein is a candidate for development of new anti-ischemic compounds. SDF-1 is monomeric at nanomolar concentrations but binding partners promote self-association at higher concentrations to form a typical CXC chemokine homodimer. Two NMR structures have been reported for the SDF-1 monomer, but only one matches the conformation observed in a series of dimeric crystal structures. In the other model, the C-terminal helix is tilted at an angle incompatible with SDF-1 dimerization. Using a rat heart explant model for ischemia/reperfusion injury, we found that dimeric SDF-1 exerts no cardioprotective effect, suggesting that the active species is monomeric. To resolve the discrepancy between existing models, we solved the NMR structure of the SDF-1 monomer in different solution conditions. Irrespective of pH and buffer composition, the C-terminal helix remains tilted at an angle with no evidence for the perpendicular arrangement. Furthermore, we find that phospholipid bicelles promote dimerization that necessarily shifts the helix to the perpendicular orientation, yielding dipolar couplings that are incompatible with the NOE distance constraints. We conclude that interactions with the alignment medium biased the previous structure, masking flexibility in the helix position that may be essential for the distinct functional properties of the SDF-1 monomer. PMID:19551879

  5. Elucidating a Key Component of Cancer Metastasis: CXCL12 (SDF-1α) Binding to CXCR4

    PubMed Central

    2015-01-01

    The chemotactic signaling induced by the binding of chemokine CXCL12 (SDF-1α) to chemokine receptor CXCR4 is of significant biological importance and is a potential therapeutic axis against HIV-1. However, as CXCR4 is overexpressed in certain cancer cells, the CXCL12:CXCR4 signaling is involved in tumor metastasis, progression, angiogenesis, and survival. Motivated by the pivotal role of the CXCL12:CXCR4 axis in cancer, we employed a comprehensive set of computational tools, predominantly based on free energy calculations and molecular dynamics simulations, to obtain insights into the molecular recognition of CXCR4 by CXCL12. We report, what is to our knowledge, the first computationally derived CXCL12:CXCR4 complex structure which is in remarkable agreement with experimental findings and sheds light into the functional role of CXCL12 and CXCR4 residues which are associated with binding and signaling. Our results reveal that the CXCL12 N-terminal domain is firmly bound within the CXCR4 transmembrane domain, and the central 24–50 residue domain of CXCL12 interacts with the upper N-terminal domain of CXCR4. The stability of the CXCL12:CXCR4 complex structure is attributed to an abundance of nonpolar and polar intermolecular interactions, including salt bridges formed between positively charged CXCL12 residues and negatively charged CXCR4 residues. The success of the computational protocol can mainly be attributed to the nearly exhaustive docking conformational search, as well as the heterogeneous dielectric implicit water-membrane-water model used to simulate and select the optimum conformations. We also recently utilized this protocol to elucidate the binding of an HIV-1 gp120 V3 loop in complex with CXCR4, and a comparison between the molecular recognition of CXCR4 by CXCL12 and the HIV-1 gp120 V3 loop shows that both CXCL12 and the HIV-1 gp120 V3 loop share the same CXCR4 binding pocket, as they mostly interact with the same CXCR4 residues. PMID:24660779

  6. Pro-inflammatory-Related Loss of CXCL12 Niche Promotes Acute Lymphoblastic Leukemic Progression at the Expense of Normal Lymphopoiesis

    PubMed Central

    Balandrán, Juan Carlos; Purizaca, Jessica; Enciso, Jennifer; Dozal, David; Sandoval, Antonio; Jiménez-Hernández, Elva; Alemán-Lazarini, Leticia; Perez-Koldenkova, Vadim; Quintela-Núñez del Prado, Henry; Rios de los Ríos, Jussara; Mayani, Héctor; Ortiz-Navarrete, Vianney; Guzman, Monica L.; Pelayo, Rosana

    2017-01-01

    Pediatric oncology, notably childhood acute lymphoblastic leukemia (ALL), is currently one of the health-leading concerns worldwide and a biomedical priority. Decreasing overall leukemia mortality in children requires a comprehensive understanding of its pathobiology. It is becoming clear that malignant cell-to-niche intercommunication and microenvironmental signals that control early cell fate decisions are critical for tumor progression. We show here that the mesenchymal stromal cell component of ALL bone marrow (BM) differ from its normal counterpart in a number of functional properties and may have a key role during leukemic development. A decreased proliferation potential, contrasting with the strong ability of producing pro-inflammatory cytokines and an aberrantly loss of CXCL12 and SCF, suggest that leukemic lymphoid niches in ALL BM are unique and may exclude normal hematopoiesis. Cell competence ex vivo assays within tridimensional coculture structures indicated a growth advantage of leukemic precursor cells and their niche remodeling ability by CXCL12 reduction, resulting in leukemic cell progression at the expense of normal niche-associated lymphopoiesis. PMID:28111575

  7. Pro-inflammatory-Related Loss of CXCL12 Niche Promotes Acute Lymphoblastic Leukemic Progression at the Expense of Normal Lymphopoiesis.

    PubMed

    Balandrán, Juan Carlos; Purizaca, Jessica; Enciso, Jennifer; Dozal, David; Sandoval, Antonio; Jiménez-Hernández, Elva; Alemán-Lazarini, Leticia; Perez-Koldenkova, Vadim; Quintela-Núñez Del Prado, Henry; Rios de Los Ríos, Jussara; Mayani, Héctor; Ortiz-Navarrete, Vianney; Guzman, Monica L; Pelayo, Rosana

    2016-01-01

    Pediatric oncology, notably childhood acute lymphoblastic leukemia (ALL), is currently one of the health-leading concerns worldwide and a biomedical priority. Decreasing overall leukemia mortality in children requires a comprehensive understanding of its pathobiology. It is becoming clear that malignant cell-to-niche intercommunication and microenvironmental signals that control early cell fate decisions are critical for tumor progression. We show here that the mesenchymal stromal cell component of ALL bone marrow (BM) differ from its normal counterpart in a number of functional properties and may have a key role during leukemic development. A decreased proliferation potential, contrasting with the strong ability of producing pro-inflammatory cytokines and an aberrantly loss of CXCL12 and SCF, suggest that leukemic lymphoid niches in ALL BM are unique and may exclude normal hematopoiesis. Cell competence ex vivo assays within tridimensional coculture structures indicated a growth advantage of leukemic precursor cells and their niche remodeling ability by CXCL12 reduction, resulting in leukemic cell progression at the expense of normal niche-associated lymphopoiesis.

  8. Effects of undenatured whey protein supplementation on CXCL12- and CCL21-mediated B and T cell chemotaxis in diabetic mice

    PubMed Central

    2011-01-01

    Background Long and persistent uncontrolled diabetes tends to degenerate the immune system and leads to an increased incidence of infection. Whey proteins (WPs) enhance immunity during early life and have a protective role in some immune disorders. In this study, the effects of camel WP on the chemotaxis of B and T cells to CXCL12 and CCL21 in diabetic mice were investigated. Results Flow cytometric analysis of the surface expressions of CXCR4 (CXCL12 receptor) and CCR7 (CCL21 receptor) on B and T cells revealed that the surface expressions of CXCR4 and CCR7 were not significantly altered in diabetic and WP-supplemented diabetic mice compared with control mice. Nevertheless, B and T lymphocytes from diabetic mice were found to be in a stunned state, with a marked and significant (P < 0.05) decrease in CXCL12- and CCL21-mediated actin polymerization and subsequently, a marked decrease in their chemotaxis. WP supplementation in the diabetes model was found to significantly increase CXCL12- and CCL21-mediated actin polymerization and chemotaxis in both B and T cells. Conclusion Our data revealed the benefits of WP supplementation in enhancing cytoskeletal rearrangement and chemotaxis in B and T cells, and subsequently improving the immune response in diabetic mice. PMID:22070978

  9. Deficits in microRNA-mediated Cxcr4/Cxcl12 signaling in neurodevelopmental deficits in a 22q11 deletion syndrome mouse model.

    PubMed

    Toritsuka, Michihiro; Kimoto, Sohei; Muraki, Kazue; Landek-Salgado, Melissa A; Yoshida, Atsuhiro; Yamamoto, Norio; Horiuchi, Yasue; Hiyama, Hideki; Tajinda, Katsunori; Keni, Ni; Illingworth, Elizabeth; Iwamoto, Takashi; Kishimoto, Toshifumi; Sawa, Akira; Tanigaki, Kenji

    2013-10-22

    22q11 deletion syndrome (22q11DS) frequently accompanies psychiatric conditions, some of which are classified as schizophrenia and bipolar disorder in the current diagnostic categorization. However, it remains elusive how the chromosomal microdeletion leads to the mental manifestation at the mechanistic level. Here we show that a 22q11DS mouse model with a deletion of 18 orthologous genes of human 22q11 (Df1/+ mice) has deficits in migration of cortical interneurons and hippocampal dentate precursor cells. Furthermore, Df1/+ mice show functional defects in Chemokine receptor 4/Chemokine ligand 12 (Cxcr4/Cxcl12; Sdf1) signaling, which reportedly underlie interneuron migration. Notably, the defects in interneuron progenitors are rescued by ectopic expression of Dgcr8, one of the genes in 22q11 microdeletion. Furthermore, heterozygous knockout mice for Dgcr8 show similar neurodevelopmental abnormalities as Df1/+ mice. Thus, Dgcr8-mediated regulation of microRNA is likely to underlie Cxcr4/Cxcl12 signaling and associated neurodevelopmental defects. Finally, we observe that expression of CXCL12 is decreased in olfactory neurons from sporadic cases with schizophrenia compared with normal controls. Given the increased risk of 22q11DS in schizophrenia that frequently shows interneuron abnormalities, the overall study suggests that CXCR4/CXCL12 signaling may represent a common downstream mediator in the pathophysiology of schizophrenia and related mental conditions.

  10. The CXCL12-3'A allele plays a favourable role in patients with multiple myeloma.

    PubMed

    Mazur, Grzegorz; Gębura, Katarzyna; Gieryng, Anna; Butrym, Aleksandra; Wróbel, Tomasz; Bogunia-Kubik, Katarzyna

    2013-10-01

    Multiple myeloma (MM) is a plasma cell malignancy characterized by bone marrow infiltration and the presence of a monoclonal protein in serum and/or urine. CXCR4 and its ligand CXCL12 are essential for neoplastic cell homing to bone marrow in haematological malignancies. The JAK2/STAT3 pathway, which is activated after CXCL12 binding to CXCR4, takes part in many signalling cascades which are linked to cell proliferation and cell survival. Constitutive activation of this pathway plays an important role in tumourigenesis and malignant transformation. The present study aimed to determine the association between the polymorphic features located within the CXCR4 (rs2228014) and CXCL12 (rs1801157) encoding genes and disease susceptibility and progression. For this purpose 172 individuals including 54 patients with MM and 118 healthy controls were typed for the CXCL12 (A/G) and CXCR4 (C/T) alleles using the PCR-RFLP technique. The CXCL12 alleles and genotypes segregated similarly among patients and controls while the CXCR4 T variant was less frequently represented among patients (OR=0.074, p<0.001). All patients with the CXCR4 T allele and 16 out of 48 with wild type genotype presented with grade III of MM according to the International Staging System (ISS) (p=0.047). The CXCL12-3'A variant was more frequently detected in patients with less advanced MM (9/17 vs. 7/38, p=0.012 for patients in stage IA or IIA vs. IIB, IIIA and IIIB, respectively). Moreover, patients lacking the CXCL12-3'A variant more frequently presented with ISS II-III (32/38 vs. 5/13, p=0.003 for patients lacking CXCL12-3'A with ISS>I vs. ISS=I). This favourable effect of the CXCL12-3'A allele was also seen in the analysis of patient survival (p<0.05). The impact of the CXCL12-3'A allele was confirmed by multivariate analyses. In conclusion, these results imply that the CXCL12-3'A allele plays a favourable role in patients with multiple myeloma. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Infection of Female BWF1 Lupus Mice with Malaria Parasite Attenuates B Cell Autoreactivity by Modulating the CXCL12/CXCR4 Axis and Its Downstream Signals PI3K/AKT, NFκB and ERK

    PubMed Central

    Badr, Gamal; Sayed, Ayat; Abdel-Maksoud, Mostafa A.; Mohamed, Amany O.; El-Amir, Azza; Abdel-Ghaffar, Fathy A.; Al-Quraishy, Saleh; Mahmoud, Mohamed H.

    2015-01-01

    Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by abnormal autoreactivity in B cells. Lymphocytes and their soluble mediators contribute to the disease pathogenesis. We recently demonstrated that infecting lupus mice with malaria confers protection against lupus nephritis by attenuating oxidative stress in both liver and kidney tissues. In the current study, we further investigated B cell autoreactivity in female BWF1 lupus mice after infection with either live or gamma-irradiated malaria, using ELISA, flow cytometry and Western blot analysis. The lupus mice exhibited a significant elevation in plasma levels of IL-4, IL-6, IL-7, IL-12, IL-17, IFN-α, IFN-γ, TGF-β, BAFF and APRIL and a marked elevation of IgG2a, IgG3 and ant-dsDNA autoantibodies compared with normal healthy mice. Infecting lupus mice with live but not gamma-irradiated malaria parasite partially and significantly restored the levels of the soluble mediators that contribute to the progression of lupus. Furthermore, the B cells of lupus mice exhibited an increased proliferative capacity; aberrant overexpression of the chemokine receptor CXCR4; and a marked elevation in responsiveness to their cognate ligand (CXCL12) via aberrant activation of the PI3K/AKT, NFκB and ERK signaling pathways. Interestingly, infecting lupus mice with live but not gamma-irradiated malaria parasite restored a normal proliferative capacity, surface expression of CXCR4 and B cell response to CXCL-12. Taken together, our data present interesting findings that clarify, for the first time, the molecular mechanisms of how infection of lupus mice with malaria parasite controls B cell autoreactivity and thus confers protection against lupus severity. PMID:25909640

  12. Infection of Female BWF1 Lupus Mice with Malaria Parasite Attenuates B Cell Autoreactivity by Modulating the CXCL12/CXCR4 Axis and Its Downstream Signals PI3K/AKT, NFκB and ERK.

    PubMed

    Badr, Gamal; Sayed, Ayat; Abdel-Maksoud, Mostafa A; Mohamed, Amany O; El-Amir, Azza; Abdel-Ghaffar, Fathy A; Al-Quraishy, Saleh; Mahmoud, Mohamed H

    2015-01-01

    Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease characterized by abnormal autoreactivity in B cells. Lymphocytes and their soluble mediators contribute to the disease pathogenesis. We recently demonstrated that infecting lupus mice with malaria confers protection against lupus nephritis by attenuating oxidative stress in both liver and kidney tissues. In the current study, we further investigated B cell autoreactivity in female BWF1 lupus mice after infection with either live or gamma-irradiated malaria, using ELISA, flow cytometry and Western blot analysis. The lupus mice exhibited a significant elevation in plasma levels of IL-4, IL-6, IL-7, IL-12, IL-17, IFN-α, IFN-γ, TGF-β, BAFF and APRIL and a marked elevation of IgG2a, IgG3 and ant-dsDNA autoantibodies compared with normal healthy mice. Infecting lupus mice with live but not gamma-irradiated malaria parasite partially and significantly restored the levels of the soluble mediators that contribute to the progression of lupus. Furthermore, the B cells of lupus mice exhibited an increased proliferative capacity; aberrant overexpression of the chemokine receptor CXCR4; and a marked elevation in responsiveness to their cognate ligand (CXCL12) via aberrant activation of the PI3K/AKT, NFκB and ERK signaling pathways. Interestingly, infecting lupus mice with live but not gamma-irradiated malaria parasite restored a normal proliferative capacity, surface expression of CXCR4 and B cell response to CXCL-12. Taken together, our data present interesting findings that clarify, for the first time, the molecular mechanisms of how infection of lupus mice with malaria parasite controls B cell autoreactivity and thus confers protection against lupus severity.

  13. The CXCL12/CXCR4 chemokine ligand/receptor axis in cardiovascular disease

    PubMed Central

    Döring, Yvonne; Pawig, Lukas; Weber, Christian; Noels, Heidi

    2014-01-01

    The chemokine receptor CXCR4 and its ligand CXCL12 play an important homeostatic function by mediating the homing of progenitor cells in the bone marrow and regulating their mobilization into peripheral tissues upon injury or stress. Although the CXCL12/CXCR4 interaction has long been regarded as a monogamous relation, the identification of the pro-inflammatory chemokine macrophage migration inhibitory factor (MIF) as an important second ligand for CXCR4, and of CXCR7 as an alternative receptor for CXCL12, has undermined this interpretation and has considerably complicated the understanding of CXCL12/CXCR4 signaling and associated biological functions. This review aims to provide insight into the current concept of the CXCL12/CXCR4 axis in myocardial infarction (MI) and its underlying pathologies such as atherosclerosis and injury-induced vascular restenosis. It will discuss main findings from in vitro studies, animal experiments and large-scale genome-wide association studies. The importance of the CXCL12/CXCR4 axis in progenitor cell homing and mobilization will be addressed, as will be the function of CXCR4 in different cell types involved in atherosclerosis. Finally, a potential translation of current knowledge on CXCR4 into future therapeutical application will be discussed. PMID:24966838

  14. Slit-2/Robo-1 modulates the CXCL12/CXCR4-induced chemotaxis of T cells.

    PubMed

    Prasad, Anil; Qamri, Zahida; Wu, Jane; Ganju, Ramesh K

    2007-09-01

    Slit, which mediates its function by binding to the Roundabout (Robo) receptor, has been shown to regulate neuronal, dendritic, and leukocyte migration. However, the molecular mechanism by which the Slit/Robo complex inhibits the migration of cells is not well defined. Here, we showed that Slit-2 can inhibit the CXCL12-induced chemotaxis and transendothelial migration of T cells and monocytes. We observed that CXCR4 associates with Robo-1 and that Slit-2 treatment enhances this association with the Robo-1 receptor. Robo-1 is a single-pass transmembrane receptor whose intracellular region contains four conserved motifs designated as CC0, CC1, CC2, and CC3. Structural and functional analyses of Robo receptors revealed that interaction of the CC3 motif with the CXCR4 receptor may regulate the CXCL12-induced chemotaxis of T cells. We further characterized Slit-2-mediated inhibition of the CXCL12/CXCR4 chemotactic pathway and found that Slit-2 can block the CXCL12-induced activation of the Src and Lck kinases but not Lyn kinase. Although Slit-2 did not inhibit the CXCL12-induced activation of MAPKs, it did inhibit the Akt phosphorylation and Rac activation induced by this chemokine. Altogether, our studies indicate a novel mechanism by which the Slit/Robo complex may inhibit the CXCR4/CXCL12-mediated chemotaxis of T cells.

  15. Stromal cell-derived CXCL12 and CCL8 cooperate to support increased development of regulatory dendritic cells following Leishmania infection.

    PubMed

    Nguyen Hoang, Anh Thu; Liu, Hao; Juaréz, Julius; Aziz, Naveed; Kaye, Paul M; Svensson, Mattias

    2010-08-15

    In the immune system, stromal cells provide specialized niches that control hematopoiesis by coordinating the production of chemokines, adhesion molecules, and growth factors. Stromal cells also have anti-inflammatory effects, including support for the differentiation of hematopoietic progenitors into dendritic cells (DCs) with immune regulatory properties. Together, these observations suggest that the alterations in hematopoiesis commonly seen in infectious disease models, such as experimental visceral leishmaniasis in mice, might result from altered stromal cell function. We report in this study that the stromal cell-derived chemokines CXCL12 and CCL8 cooperate to attract hematopoietic progenitors with the potential to differentiate into regulatory DCs. We also show that infection of murine bone marrow stromal cells by Leishmania donovani enhanced their capacity to support the development of regulatory DCs, as well as their capacity to produce CCL8. Likewise, in experimental visceral leishmaniasis, CCL8 production was induced in splenic stromal cells, leading to an enhanced capacity to attract hematopoietic progenitor cells. Thus, intracellular parasitism of stromal cells modifies their capacity to recruit and support hematopoietic progenitor differentiation into regulatory DCs, and aberrant expression of CCL8 by diseased stromal tissue may be involved in the switch from resolving to persistent infection.

  16. Inhibition of the CXCL12/CXCR4-axis as preventive therapy for radiation-induced pulmonary fibrosis.

    PubMed

    Shu, Hui-Kuo G; Yoon, Younghyoun; Hong, Samuel; Xu, Kaiming; Gao, Huiying; Hao, Chunhai; Torres-Gonzalez, Edilson; Nayra, Cardenes; Rojas, Mauricio; Shim, Hyunsuk

    2013-01-01

    A devastating late injury caused by radiation is pulmonary fibrosis. This risk may limit the volume of irradiation and compromise potentially curative therapy. Therefore, development of a therapy to prevent this toxicity can be of great benefit for this patient population. Activation of the chemokine receptor CXCR4 by its ligand stromal cell-derived factor 1 (SDF-1/CXCL12) may be important in the development of radiation-induced pulmonary fibrosis. Here, we tested whether MSX-122, a novel small molecule and partial CXCR4 antagonist, can block development of this fibrotic process. The radiation-induced lung fibrosis model used was C57BL/6 mice irradiated to the entire thorax or right hemithorax to 20 Gy. Our parabiotic model involved joining a transgenic C57BL/6 mouse expressing GFP with a wild-type mouse that was subsequently irradiated to assess for migration of GFP+ bone marrow-derived progenitor cells to the irradiated lung. CXCL12 levels in the bronchoalveolar lavage fluid (BALF) and serum after irradiation were determined by ELISA. CXCR4 and CXCL12 mRNA in the irradiated lung was determined by RNase protection assay. Irradiated mice were treated daily with AMD3100, an established CXCR4 antagonist; MSX-122; and their corresponding vehicles to determine impact of drug treatment on fibrosis development. Fibrosis was assessed by serial CTs and histology. After irradiation, CXCL12 levels increased in BALF and serum with a corresponding rise in CXCR4 mRNA within irradiated lungs consistent with recruitment of a CXCR4+ cell population. Using our parabiotic model, we demonstrated recruitment of CXCR4+ bone marrow-derived mesenchymal stem cells, identified based on marker expression, to irradiated lungs. Finally, irradiated mice that received MSX-122 had significant reductions in development of pulmonary fibrosis while AMD3100 did not significantly suppress this fibrotic process. CXCR4 inhibition by drugs such as MSX-122 may alleviate potential radiation-induced lung

  17. Pharmacological targeting of CXCL12/CXCR4 signaling in prostate cancer bone metastasis.

    PubMed

    Conley-LaComb, M Katie; Semaan, Louie; Singareddy, Rajareddy; Li, Yanfeng; Heath, Elisabeth I; Kim, Seongho; Cher, Michael L; Chinni, Sreenivasa R

    2016-11-03

    The CXCL12/CXCR4 axis transactivates HER2 and promotes intraosseous tumor growth. To further explore the transactivation of HER2 by CXCL12, we investigated the role of small GTP protein Gαi2 in Src and HER2 phosphorylation in lipid raft membrane microdomains and the significance of CXCR4 in prostate cancer bone tumor growth. We used a variety of methods such as lipid raft isolation, invasion assays, an in vivo model of intratibial tumor growth, bone histomorphometry, and immunohistochemistry to determine the role of CXCR4 signaling in lipid raft membrane microdomains and effects of targeting of CXCR4 for bone tumor growth. We determined that (a) CXCL12/CXCR4 transactivation of EGFR and HER2 is confined to lipid raft membrane microdomains, (b) CXCL12 activation of HER2 and Src is mediated by small GTP proteins in lipid rafts, (c) inhibition of the CXCL12/CXCR4 axis through plerixafor abrogates the initial establishment of tumor growth without affecting the growth of established bone tumors, and (d) inhibition of EGFR signaling through gefitinib leads to inhibition of established bone tumor growth. These data suggest that lipid raft membrane microdomains are key sites for CXCL12/CXCR4 transactivation of HER2 via small GTP binding protein Gαi2 and Src kinase. The initial establishment of prostate cancer is supported by the endosteal niche, and blocking the CXCL12/CXCR4 axis of this niche along with its downstream signaling severely compromises initial establishment of tumors in the bone microenvironment, whereas expanding bone tumors are sensitive only to the members of growth factor receptor inhibition.

  18. The CXCL12/CXCR4 Signaling Pathway: A New Susceptibility Factor in Human Papillomavirus Pathogenesis

    PubMed Central

    Meuris, Floriane; Carthagena, Laetitia; Cutolo, Pasquale; Xue, Yuezhen; Thierry, Françoise; Doorbar, John; Bachelerie, Françoise

    2016-01-01

    The productive human papillomavirus (HPV) life cycle is tightly linked to the differentiation and cycling of keratinocytes. Deregulation of these processes and stimulation of cell proliferation by the action of viral oncoproteins and host cell factors underlies HPV-mediated carcinogenesis. Severe HPV infections characterize the wart, hypogammaglobulinemia, infection, and myelokathexis (WHIM) immunodeficiency syndrome, which is caused by gain-of-function mutations in the CXCR4 receptor for the CXCL12 chemokine, one of which is CXCR41013. We investigated whether CXCR41013 interferes in the HPV18 life cycle in epithelial organotypic cultures. Expression of CXCR41013 promoted stabilization of HPV oncoproteins, thus disturbing cell cycle progression and proliferation at the expense of the ordered expression of the viral genes required for virus production. Conversely, blocking CXCR41013 function restored virus production and limited HPV-induced carcinogenesis. Thus, CXCR4 and its potential activation by genetic alterations in the course of the carcinogenic process can be considered as an important host factor for HPV carcinogenesis. PMID:27918748

  19. Natural nitration of CXCL12 reduces its signaling capacity and chemotactic activity in vitro and abrogates intra-articular lymphocyte recruitment in vivo

    PubMed Central

    Janssens, Rik; Mortier, Anneleen; Boff, Daiane; Vanheule, Vincent; Gouwy, Mieke; Franck, Charlotte; Larsen, Olav; Rosenkilde, Mette M.; Van Damme, Jo; Amaral, Flávio A.; Teixeira, Mauro M.; Struyf, Sofie; Proost, Paul

    2016-01-01

    The chemokine CXCL12/stromal cell-derived factor-1 is important for leukocyte migration to lymphoid organs and inflamed tissues and stimulates tumor development. In vitro, CXCL12 activity through CXCR4 is abolished by proteolytic processing. However, limited information is available on in vivo effects of posttranslationally modified CXCL12. Natural CXCL12 was purified from the coculture supernatant of stromal cells stimulated with leukocytes and inflammatory agents. In this conditioned medium, CXCL12 with a nitration on Tyr7, designated [3-NT7]CXCL12, was discovered via Edman degradation. CXCL12 and [3-NT7]CXCL12 were chemically synthesized to evaluate the biological effects of this modification. [3-NT7]CXCL12 recruited β-arrestin 2 and phosphorylated the Akt kinase similar to CXCL12 in receptor-transfected cells. Also the affinity of CXCL12 and [3-NT7]CXCL12 for glycosaminoglycans, the G protein-coupled chemokine receptor CXCR4 and the atypical chemokine receptor ACKR3 were comparable. However, [3-NT7]CXCL12 showed a reduced ability to enhance intracellular calcium concentrations, to generate inositol triphosphate, to phosphorylate ERK1/2 and to induce monocyte and lymphocyte chemotaxis in vitro. Moreover, nitrated CXCL12 failed to induce in vivo extravasation of lymphocytes to the joint. In summary, nitration on Tyr7 under inflammatory conditions is a novel natural posttranslational regulatory mechanism of CXCL12 which may downregulate the CXCR4-mediated inflammatory and tumor-promoting activities of CXCL12. PMID:27566567

  20. Essential role of immobilized chemokine CXCL12 in the regulation of the humoral immune response

    PubMed Central

    Barinov, Aleksandr; Luo, Lingjie; Gasse, Pamela; Meas-Yedid, Vannary; Donnadieu, Emmanuel; Arenzana-Seisdedos, Fernando; Vieira, Paulo

    2017-01-01

    Chemokines control the migration of a large array of cells by binding to specific receptors on cell surfaces. The biological function of chemokines also depends on interactions between nonreceptor binding domains and proteoglycans, which mediate chemokine immobilization on cellular or extracellular surfaces and formation of fixed gradients. Chemokine gradients regulate synchronous cell motility and integrin-dependent cell adhesion. Of the various chemokines, CXCL12 has a unique structure because its receptor-binding domain is distinct and does not overlap with the immobilization domains. Although CXCL12 is known to be essential for the germinal center (GC) response, the role of its immobilization in biological functions has never been addressed. In this work, we investigated the unexplored paradigm of CXCL12 immobilization during the germinal center reaction, a fundamental process where cellular traffic is crucial for the quality of humoral immune responses. We show that the structure of murine germinal centers and the localization of GC B cells are impaired when CXCL12 is unable to bind to cellular or extracellular surfaces. In such mice, B cells carry fewer somatic mutations in Ig genes and are impaired in affinity maturation. Therefore, immobilization of CXCL12 is necessary for proper trafficking of B cells during GC reaction and for optimal humoral immune responses. PMID:28193885

  1. CXCL12 retargeting of an adenovirus vector to cancer cells using a bispecific adapter.

    PubMed

    Bhatia, Shilpa; O'Bryan, Samia M; Rivera, Angel A; Curiel, David T; Mathis, J Michael

    2016-01-01

    Ad vectors are promising delivery vehicles for cancer therapeutic interventions. However, their application is limited by promiscuous tissue tropism and hepatotoxicity. This limitation can be avoided by altering the native tropism of Ads so that they can be redirected to the target cells through alternate cellular receptors. The CXCR4 chemokine receptor belongs to a large superfamily of G-protein-coupled receptors and is known to be upregulated in a wide variety of cancers, including breast cancer and melanoma. These receptors have been associated with cancer cell survival, progression, and metastasis. In the current study, an Ad to cancer cells overexpressing CXCR4 by using a bispecific adapter, sCAR-CXCL12, was retargeted. The sCAR-CXCL12 adapter contained the soluble ectodomain form of the native Ad5 receptor (sCAR), which was fused to a mature human chemokine ligand, CXCL12, through a short peptide linker. A dramatic increase in the infectivity of cancer cells using a targeted Ad vector compared with an untargeted vector was observed. Furthermore, sCAR-CXCL12 attenuated Ad infection of liver ex vivo and in vivo and enhanced Ad vector infection of xenograft tumors implanted in immunodeficient SCID-bg mice. Thus, the sCAR-CXCL12 adapter could be used to retarget Ad vectors to chemokine receptor-positive tumors.

  2. CXCL12 retargeting of an adenovirus vector to cancer cells using a bispecific adapter

    PubMed Central

    Bhatia, Shilpa; O’Bryan, Samia M; Rivera, Angel A; Curiel, David T; Mathis, J Michael

    2016-01-01

    Ad vectors are promising delivery vehicles for cancer therapeutic interventions. However, their application is limited by promiscuous tissue tropism and hepatotoxicity. This limitation can be avoided by altering the native tropism of Ads so that they can be redirected to the target cells through alternate cellular receptors. The CXCR4 chemokine receptor belongs to a large superfamily of G-protein-coupled receptors and is known to be upregulated in a wide variety of cancers, including breast cancer and melanoma. These receptors have been associated with cancer cell survival, progression, and metastasis. In the current study, an Ad to cancer cells overexpressing CXCR4 by using a bispecific adapter, sCAR-CXCL12, was retargeted. The sCAR-CXCL12 adapter contained the soluble ectodomain form of the native Ad5 receptor (sCAR), which was fused to a mature human chemokine ligand, CXCL12, through a short peptide linker. A dramatic increase in the infectivity of cancer cells using a targeted Ad vector compared with an untargeted vector was observed. Furthermore, sCAR-CXCL12 attenuated Ad infection of liver ex vivo and in vivo and enhanced Ad vector infection of xenograft tumors implanted in immunodeficient SCID-bg mice. Thus, the sCAR-CXCL12 adapter could be used to retarget Ad vectors to chemokine receptor-positive tumors. PMID:27957479

  3. Modulation of CXCR4, CXCL12, and Tumor Cell Invasion Potential In Vitro by Phytochemicals

    PubMed Central

    Hsu, Erin L.; Chen, Natalie; Westbrook, Aya; Wang, Feng; Zhang, Ruixue; Taylor, Robert T.; Hankinson, Oliver

    2009-01-01

    CXCR4 is a chemokine receptor frequently overexpressed on primary tumor cells. Organs to which these cancers metastasize secrete CXCL12, the unique ligand for CXCR4, which stimulates invasion and metastasis to these sites. Similar to our previous work with the chemoprotective phytochemical, 3,3′-diindolylmethane (DIM), we show here that genistein also downregulates CXCR4 and CXCL12 and subsequently lowers the migratory and invasive potentials of breast and ovarian cancer cells. Moreover, genistein and DIM elicit a significantly greater cumulative effect in lowering CXCR4 and CXCL12 levels than either compound alone. Our data suggest a novel mechanism for the protective effects of phytochemicals against cancer progression and indicate that in combination, these compounds may prove even more efficacious. PMID:19325924

  4. CXCL12 impairs the acquisition and extinction of auditory fear conditioning in rats via crosstalk with GABAergic system.

    PubMed

    Shi, Lijuan; Bi, Qiang; Li, Wai; Qin, Ling; Yang, Pingting

    2016-09-01

    Chemokines, such as CXCL12, are signaling molecules playing an important role in immune regulations. Chemokine upsurge has also been associated with neuroinflammatory conditions characterized with cognitive impairments. Recently, some in-vitro data suggests that CXCL12 is a potential neuromodulator and interacts with GABAergic system, but, so far, whether these effects translate into alterations in neural and behavioral functions has not been investigated. In the present study, we used auditory fear conditioning as a model to define the contribution of CXCL12/CXCR4 on fear-related cognitive disorders. We microinjected different dosages of CXCL12 into the bilateral amygdala of rats to investigate their behavioral effects on the acquisition and extinction of conditioned fear memory. Moreover, we pretreated the rats with the selective CXCR4 receptor antagonist (AMD3100), GABAA antagonist (bicuculline) and GABAB antagonist (CGP55845) to examine whether the CXCL12 induced changes could be reversed. We found that intra-amygdala infusion of CXCL12 impaired the acquisition and extinction of conditioned fear response. Pretreatment with AMD3100, rescued the CXCL12 induced impairments, indicating that CXCL12 produced the effects by activating CXCR4 receptors. Furthermore, both bicuculline and CGP55845 prevented CXCL12 from impairing the rat's ability of conditioned learning, indicating a crosstalk between CXCL12/CXCR4 and GABAergic system. Our data suggest that the chemokine CXCL12 is able to regulate neurotransmitter mechanisms involved in associative learning functions, and the effect of GABAergic agents on CXCL12/CXCR4 may be new therapeutic potentials for neuroinflammatory diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. CCR2, CCR5, and CXCL12 variation and HIV/AIDS in Papua New Guinea.

    PubMed

    Mehlotra, Rajeev K; Hall, Noemi B; Bruse, Shannon E; John, Bangan; Blood Zikursh, Melinda J; Stein, Catherine M; Siba, Peter M; Zimmerman, Peter A

    2015-12-01

    Polymorphisms in chemokine receptors, serving as HIV co-receptors, and their ligands are among the well-known host genetic factors associated with susceptibility to HIV infection and/or disease progression. Papua New Guinea (PNG) has one of the highest adult HIV prevalences in the Asia-Pacific region. However, information regarding the distribution of polymorphisms in chemokine receptor (CCR5, CCR2) and chemokine (CXCL12) genes in PNG is very limited. In this study, we genotyped a total of nine CCR2-CCR5 polymorphisms, including CCR2 190G >A, CCR5 -2459G >A and Δ32, and CXCL12 801G >A in PNG (n=258), North America (n=184), and five countries in West Africa (n=178). Using this data, we determined previously characterized CCR5 haplotypes. In addition, based on the previously reported associations of CCR2 190, CCR5 -2459, CCR5 open reading frame, and CXCL12 801 genotypes with HIV acquisition and/or disease progression, we calculated composite full risk scores, considering both protective as well as susceptibility effects of the CXCL12 801 AA genotype. We observed a very high frequency of the CCR5 -2459A allele (0.98) in the PNG population, which together with the absence of Δ32 resulted in a very high frequency of the HHE haplotype (0.92). These frequencies were significantly higher than in any other population (all P-values<0.001). Regardless of whether we considered the CXCL12 801 AA genotype protective or susceptible, the risk scores were significantly higher in the PNG population compared with any other population (all P-values<0.001). The results of this study provide new insights regarding CCR5 variation in the PNG population, and suggest that the collective variation in CCR2, CCR5, and CXCL12 may increase the risk of HIV/AIDS in a large majority of Papua New Guineans.

  6. Microfluidic investigation of BDNF-enhanced neural stem cell chemotaxis in CXCL12 gradients.

    PubMed

    Xu, Hui; Heilshorn, Sarah C

    2013-02-25

    In vivo studies have suggested that gradients of CXCL12 (aka stromal cell-derived factor 1α) may be critical for neural stem cell (NSC) migration during brain development and neural tissue regeneration. However, traditional in vitro chemotaxis tools are limited by unstable concentration gradients and the inability to decouple cell migration directionality and speed. These limitations have restricted the reproducible and quantitative analysis of neuronal migration, which is required for mechanism-based studies. Using a microfluidic gradient generator, nestin and Sox-2 positive human embryonic NSC chemotaxis is quantified within a linear and stable CXCL12 gradient. While untreated NSCs are not able to chemotax within CXCL12 gradients, pre-treatment of the cells with brain-derived neurotrophic factor (BDNF) results in significant chemotactic, directional migration. BDNF pre-treatment has no effect on cell migration speed, which averages about 1 μm min(-1). Quantitative analysis determines that CXCL12 concentrations above 9.0 nM are above the minimum activation threshold, while concentrations below 14.7 nM are below the saturation threshold. Interestingly, although inhibitor studies with AMD 3100 revealed that CXCL12 chemotaxis requires receptor CXCR4 activation, BDNF pre-treatment is found to have no profound effects on the mRNA levels or surface presentation of CXCR4 or the putative CXCR7 scavenger receptor. The microfluidic study of NSC migration within stable chemokine concentration profiles provides quantitative analysis as well as new insight into the migratory mechanism underlying BDNF-induced chemotaxis towards CXCL12. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. IL-2 augments the therapeutic efficacy of adoptively transferred B cells which directly kill tumor cells via the CXCR4/CXCL12 and perforin pathways

    PubMed Central

    Chen, Xin; Xia, Leiming; Zhou, Li; Wang, Yi; Bao, Yangyi; Huang, Shiang; Ren, Xiubao; Lundy, Steven K.; Dai, Fu; Li, Qiao; Chang, Alfred E.

    2016-01-01

    We previously reported that antitumor B cells directly kill tumor cells via the Fas/FasL pathway and are regulated by IL-10. In this study, we defined additional mechanisms involved in B cell antitumor immunity. Administration of IL-2 significantly augmented the therapeutic efficacy of adoptively transferred tumor-draining lymph node (TDLN) B cells which express IL- 2R. Culture supernatant of purified B splenocytes harvested from the mice that received adoptive transfer of 4T1 TDLN B cells plus IL-2 administration produced larger amounts of IgG which bound to 4T1, resulting in 4T1 lysis. Furthermore, we detected CXCR4 expression on 4T1 TDLN B cells, and 4T1 tumor cells produced its ligand CXCL12. Transwell experiments demonstrated the chemoattraction of CXCR4-expressing 4T1 TDLN B cells towards CXCL12- producing 4T1 cells. Blockade of CXCR4 using a CXCR4-specific inhibitor, AMD3100, significantly reduced the killing of 4T1 tumor cells by 4T1 TDLN B cells. Blockade of FasL and CXCR4 concurrently inhibited B cell-mediated direct killing of tumor cells in an additive manner, indicating that both Fas/FasL and CXCL12/CXCR4 pathways are involved in the direct killing of 4T1 cells by 4T1 TDLN B cells. TDLN B cells produced perforin. Additional transwell experiments showed that effector B cells could directly kill tumor cells in cell-cell contact via the Fas/FasL and CXCR4/CXCL12 pathways as well as perforin, while without cell contact, perforin secreted by B cells led to tumor cell cytotoxicity. These findings underscore the diversity of function by which B cells can play an important role in the host immune response to tumor. PMID:27528023

  8. IL-2 augments the therapeutic efficacy of adoptively transferred B cells which directly kill tumor cells via the CXCR4/CXCL12 and perforin pathways.

    PubMed

    Xia, Yang; Tao, Huimin; Hu, Yangyang; Chen, Quanning; Chen, Xin; Xia, Leiming; Zhou, Li; Wang, Yi; Bao, Yangyi; Huang, Shiang; Ren, Xiubao; Lundy, Steven K; Dai, Fu; Li, Qiao; Chang, Alfred E

    2016-09-13

    We previously reported that antitumor B cells directly kill tumor cells via the Fas/FasL pathway and are regulated by IL-10. In this study, we defined additional mechanisms involved in B cell antitumor immunity. Administration of IL-2 significantly augmented the therapeutic efficacy of adoptively transferred tumor-draining lymph node (TDLN) B cells which express IL- 2R. Culture supernatant of purified B splenocytes harvested from the mice that received adoptive transfer of 4T1 TDLN B cells plus IL-2 administration produced larger amounts of IgG which bound to 4T1, resulting in 4T1 lysis. Furthermore, we detected CXCR4 expression on 4T1 TDLN B cells, and 4T1 tumor cells produced its ligand CXCL12. Transwell experiments demonstrated the chemoattraction of CXCR4-expressing 4T1 TDLN B cells towards CXCL12- producing 4T1 cells. Blockade of CXCR4 using a CXCR4-specific inhibitor, AMD3100, significantly reduced the killing of 4T1 tumor cells by 4T1 TDLN B cells. Blockade of FasL and CXCR4 concurrently inhibited B cell-mediated direct killing of tumor cells in an additive manner, indicating that both Fas/FasL and CXCL12/CXCR4 pathways are involved in the direct killing of 4T1 cells by 4T1 TDLN B cells. TDLN B cells produced perforin. Additional transwell experiments showed that effector B cells could directly kill tumor cells in cell-cell contact via the Fas/FasL and CXCR4/CXCL12 pathways as well as perforin, while without cell contact, perforin secreted by B cells led to tumor cell cytotoxicity. These findings underscore the diversity of function by which B cells can play an important role in the host immune response to tumor.

  9. Regulator of G protein signaling 1 suppresses CXCL12-mediated migration and AKT activation in RPMI 8226 human plasmacytoma cells and plasmablasts.

    PubMed

    Pak, Hyo-Kyung; Gil, Minchan; Lee, Yoonkyung; Lee, Hyunji; Lee, A-Neum; Roh, Jin; Park, Chan-Sik

    2015-01-01

    Migration of plasma cells to the bone marrow is critical factor to humoral immunity and controlled by chemokines. Regulator of G protein signaling 1 (RGS1) is a GTPase-activating protein that controls various crucial functions such as migration. Here, we show that RGS1 controls the chemotactic migration of RPMI 8226 human plasmacytoma cells and human plasmablasts. LPS strongly increased RGS1 expression and retarded the migration of RPMI 8226 cells by suppressing CXCL12-mediated AKT activation. RGS1 knockdown by siRNA abolished the retardation of migration and AKT suppression by LPS. RGS1-dependent regulation of migration via AKT is also observed in cultured plasmablasts. We propose novel functions of RGS1 that suppress AKT activation and the migration of RPMI 8226 cells and plasmablasts in CXCL12-mediated chemotaxis.

  10. A Novel Combination of Calprotectin and CXCL12 for Predicting Malignancy in Patients with Exudative Pleural Effusion.

    PubMed

    Luo, Jian; Wang, Maoyun; Li, Chuntao; Liang, Binmiao; Liu, Dan; Shi, Chaoli; Jiang, Faming; Wang, Ting; Li, Peijun; Liang, Zongan

    2015-11-01

    Pleural effusion (PE) remains a significant challenge and public health problem, which needs novel noninvasive biomarkers for the precise diagnosis. The aim of this study was to further determine the clinical efficacy and diagnostic accuracy of a novel combination of calprotectin and CXCL12 for predicting malignancy in patients with exudative PE.Calprotectin and CXCL12 concentrations were measured in 95 individuals of exudative PE, with 39 malignant PE (MPE) and 56 benign PE (BPE). The accuracy of calprotectin and CXCL12 levels for discriminating MPE from BPE or tuberculous PE were evaluated using receiver-operating characteristic (ROC) curves. Univariate and multivariate logistic regression analyses were performed to test the association between calprotectin and CXCL12 levels and MPE.Calprotectin and CXCL12 levels of patients with MPE were significantly lower than that of BPE and tuberculous PE (P < 0.05). The area under the curve (AUC) of calprotectin and CXCL12 was 0.683 and 0.641 in MPE and BPE, and a combination of calprotectin ≤500.19 ng/mL and CXCL12 ≤6.11 ng/mL rendered a sensitivity and specificity of 48.72% and 78.57%, respectively. While in MPE and tuberculous PE, the AUC of calprotectin and CXCL12 was 0.696 and 0.690, and a combination of calprotectin ≤421.73 ng/mL and CXCL12 ≤3.71 ng/mL presented a sensitivity and specificity of 25.64% and 95.45%, respectively. Multivariate logistic regression demonstrated that both calprotectin and CXCL12 were independent predictors of MPE.Calprotectin and CXCL12 in pleural fluid are informative diagnostic biomarkers for predicting patients with MPE.

  11. CXCL12 sensitizes vago-vagal reflex neurons in the dorsal medulla.

    PubMed

    Rogers, Richard C; Viard, Edouard; Hermann, Gerlinda E

    2013-01-25

    Previous studies from our laboratory illustrated the potential for stromal cell-derived factor one [CXCL12; also referred to as SDF-1] to act on its receptor [CXCR4] within the dorsal vagal complex [DVC] of the hindbrain to suppress gastric motility (Hermann et al., 2008). While CXCR4 receptors are essential for normal brain development, they also play a critical role in the proliferation of the HIV virus and initiation of metastatic cell growth in the brain. Anorexia, nausea, and failed autonomic regulation of gastrointestinal function are significant causes of morbidity and are contributory factors in the mortality associated with these disease states. The implication of our previous study was that CXCL12 caused gastric stasis by acting on gastric reflex circuit elements in the DVC. This hindbrain complex includes vagal afferent terminations in the solitary nucleus, neurons in the solitary nucleus (NST) and visceral efferent motorneurons in the dorsal motor nucleus (DMN) that are responsible for the regulation of digestive functions from the oral cavity to the transverse colon. In the current study, in vivo single-unit neurophysiological recordings from physiologically-identified NST and DMN components of the gastric accommodation reflex show that while injection of femtomole doses of CXCL12 onto NST or DMN neurons has no effect on their basal activity, CXCL12 amplifies the effect of gastric vagal mechanosensory input to activate the NST and, in turn, inhibit DMN motor activity.

  12. CXCR4/CXCL12 axis counteracts hematopoietic stem cell exhaustion through selective protection against oxidative stress

    PubMed Central

    Zhang, Yanyan; Dépond, Mallorie; He, Liang; Foudi, Adlen; Kwarteng, Edward Owusu; Lauret, Evelyne; Plo, Isabelle; Desterke, Christophe; Dessen, Philippe; Fujii, Nobutaka; Opolon, Paule; Herault, Olivier; Solary, Eric; Vainchenker, William; Joulin, Virginie; Louache, Fawzia; Wittner, Monika

    2016-01-01

    Hematopoietic stem cells (HSCs) undergo self-renewal to maintain hematopoietic homeostasis for lifetime, which is regulated by the bone marrow (BM) microenvironment. The chemokine receptor CXCR4 and its ligand CXCL12 are critical factors supporting quiescence and BM retention of HSCs. Here, we report an unknown function of CXCR4/CXCL12 axis in the protection of HSCs against oxidative stress. Disruption of CXCR4 receptor in mice leads to increased endogenous production of reactive oxygen species (ROS), resulting in p38 MAPK activation, increased DNA double-strand breaks and apoptosis leading to marked reduction in HSC repopulating potential. Increased ROS levels are directly responsible for exhaustion of the HSC pool and are not linked to loss of quiescence of CXCR4-deficient HSCs. Furthermore, we report that CXCL12 has a direct rescue effect on oxidative stress-induced HSC damage at the mitochondrial level. These data highlight the importance of CXCR4/CXCL12 axis in the regulation of lifespan of HSCs by limiting ROS generation and genotoxic stress. PMID:27886253

  13. Real-time detection of the chemokine CXCL12 in urine samples by surface plasmon resonance.

    PubMed

    Vega, Beatriz; Calle, Ana; Sánchez, Alejandra; Lechuga, Laura M; Ortiz, Ana M; Armelles, Gaspar; Rodríguez-Frade, José Miguel; Mellado, Mario

    2013-05-15

    Surface plasmon resonance (SPR)-based biosensors are established tools for measuring biomolecular interactions between unlabeled analytes in real time, and are thus an ideal method to evaluate G protein-coupled receptor (GPCR) binding interactions. Using as a vehicle lentiviral particles bearing the chemokine receptor CXCR4 in its native plasma membrane context, SPR analysis can be performed using the particles as specific receptors to monitor the CXCR4 interaction with its ligand, CXCL12. The method shows linear correlation in the 5-40 nM range, with low intra- and inter-assay variation, a relative standard deviation <10%, chip-to-chip variation <12%, with stability of the sensor response for more than 150 measurements in the same chip over a four-week period. Our objective was to develop a method for rapid detection and quantification of analytes such as CXCL12 in biological samples, with no need for pretreatment. As a proof of concept, we tested for CXCL12 in urine samples from rheumatoid arthritis patients, who have elevated levels of this chemokine in plasma and synovial fluid. The biosensor method allowed sensitive, reproducible CXCL12 detection in the physiological range, suggesting its value for the diagnosis of autoimmune disorders. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Emerging targets in cancer management: role of the CXCL12/CXCR4 axis

    PubMed Central

    Cojoc, Monica; Peitzsch, Claudia; Trautmann, Franziska; Polishchuk, Leo; Telegeev, Gennady D; Dubrovska, Anna

    2013-01-01

    The chemokine CXCL12 (SDF-1) and its cell surface receptor CXCR4 were first identified as regulators of lymphocyte trafficking to the bone marrow. Soon after, the CXCL12/CXCR4 axis was proposed to regulate the trafficking of breast cancer cells to sites of metastasis. More recently, it was established that CXCR4 plays a central role in cancer cell proliferation, invasion, and dissemination in the majority of malignant diseases. The stem cell concept of cancer has revolutionized the understanding of tumorigenesis and cancer treatment. A growing body of evidence indicates that a subset of cancer cells, referred to as cancer stem cells (CSCs), plays a critical role in tumor initiation, metastatic colonization, and resistance to therapy. Although the signals generated by the metastatic niche that regulate CSCs are not yet fully understood, accumulating evidence suggests a key role of the CXCL12/CXCR4 axis. In this review we focus on physiological functions of the CXCL12/CXCR4 signaling pathway and its role in cancer and CSCs, and we discuss the potential for targeting this pathway in cancer management. PMID:24124379

  15. The hematopoietic chemokine CXCL12 promotes integration of human endothelial colony forming cell-derived cells into immature vessel networks.

    PubMed

    Newey, Sarah E; Tsaknakis, Grigorios; Khoo, Cheen P; Athanassopoulos, Thanassi; Camicia, Rosalba; Zhang, Youyi; Grabowska, Rita; Harris, Adrian L; Roubelakis, Maria G; Watt, Suzanne M

    2014-11-15

    Proangiogenic factors, vascular endothelial growth factor (VEGF), and fibroblast growth factor-2 (FGF-2) prime endothelial cells to respond to "hematopoietic" chemokines and cytokines by inducing/upregulating expression of the respective chemokine/cytokine receptors. Coculture of human endothelial colony forming cell (ECFC)-derived cells with human stromal cells in the presence of VEGF and FGF-2 for 14 days resulted in upregulation of the "hematopoietic" chemokine CXCL12 and its CXCR4 receptor by day 3 of coculture. Chronic exposure to the CXCR4 antagonist AMD3100 in this vasculo/angiogenesis assay significantly reduced vascular tubule formation, an observation recapitulated by delayed AMD3100 addition. While AMD3100 did not affect ECFC-derived cell proliferation, it did demonstrate a dual action. First, over the later stages of the 14-day cocultures, AMD3100 delayed tubule organization into maturing vessel networks, resulting in enhanced endothelial cell retraction and loss of complexity as defined by live cell imaging. Second, at earlier stages of cocultures, we observed that AMD3100 significantly inhibited the integration of exogenous ECFC-derived cells into established, but immature, vascular networks. Comparative proteome profiler array analyses of ECFC-derived cells treated with AMD3100 identified changes in expression of potential candidate molecules involved in adhesion and/or migration. Blocking antibodies to CD31, but not CD146 or CD166, reduced the ECFC-derived cell integration into these extant vascular networks. Thus, CXCL12 plays a key role not only in endothelial cell sensing and guidance, but also in promoting the integration of ECFC-derived cells into developing vascular networks.

  16. CXCL12 and CXCR7 are relevant targets to reverse cell adhesion-mediated drug resistance in multiple myeloma.

    PubMed

    Waldschmidt, Johannes M; Simon, Anna; Wider, Dagmar; Müller, Stefan J; Follo, Marie; Ihorst, Gabriele; Decker, Sarah; Lorenz, Joschka; Chatterjee, Manik; Azab, Abdel K; Duyster, Justus; Wäsch, Ralph; Engelhardt, Monika

    2017-10-01

    Cell adhesion-mediated drug resistance (CAM-DR) by the bone marrow (BM) is fundamental to multiple myeloma (MM) propagation and survival. Targeting BM protection to increase the efficacy of current anti-myeloma treatment has not been extensively pursued. To extend the understanding of CAM-DR, we hypothesized that the cytotoxic effects of novel anti-myeloma agents may be abrogated by the presence of BM stroma cells (BMSCs) and restored by addition of the CXCL12 antagonist NOX-A12 or the CXCR4 inhibitor plerixafor. Following this hypothesis, we evaluated different anti-myeloma agents alone, with BMSCs and when combined with plerixafor or NOX-A12. We verified CXCR4, CD49d (also termed ITGA4) and CD44 as essential mediators of BM adhesion on MM cells. Additionally, we show that CXCR7, the second receptor of stromal-derived-factor-1 (CXCL12), is highly expressed in active MM. Co-culture proved that co-treatment with plerixafor or NOX-A12, the latter inhibiting CXCR4 and CXCR7, functionally interfered with MM chemotaxis to the BM. This led to the resensitization of MM cells to the anti-myeloma agents vorinostat and pomalidomide and both proteasome inhibitors bortezomib and carfilzomib. Within a multicentre phase I/II study, NOX-A12 was tested in combination with bortezomib-dexamethasone, underlining the feasibility of NOX-A12 as an active add-on agent to antagonize myeloma CAM-DR. © 2017 John Wiley & Sons Ltd.

  17. Structural and Functional Basis of CXCL12 (stromal cell-derived factor-1 alpha) Binding to Heparin

    SciTech Connect

    Murphy,J.; Cho, Y.; Sachpatzidis, A.; Fan, C.; Hodsdon, M.; Lolis, E.

    2007-01-01

    CXCL12 (SDF-1a) and CXCR4 are critical for embryonic development and cellular migration in adults. These proteins are involved in HIV-1 infection, cancer metastasis, and WHIM disease. Sequestration and presentation of CXCL12 to CXCR4 by glycosaminoglycans (GAGs) is proposed to be important for receptor activation. Mutagenesis has identified CXCL12 residues that bind to heparin. However, the molecular details of this interaction have not yet been determined. Here we demonstrate that soluble heparin and heparan sulfate negatively affect CXCL12-mediated in vitro chemotaxis. We also show that a cluster of basic residues in the dimer interface is required for chemotaxis and is a target for inhibition by heparin. We present structural evidence for binding of an unsaturated heparin disaccharide to CXCL12 attained through solution NMR spectroscopy and x-ray crystallography. Increasing concentrations of the disaccharide altered the two-dimensional 1H-15N-HSQC spectra of CXCL12, which identified two clusters of residues. One cluster corresponds to {beta}-strands in the dimer interface. The second includes the amino-terminal loop and the a-helix. In the x-ray structure two unsaturated disaccharides are present. One is in the dimer interface with direct contacts between residues His25, Lys27, and Arg41 of CXCL12 and the heparin disaccharide. The second disaccharide contacts Ala20, Arg21, Asn30, and Lys64. This is the first x-ray structure of a CXC class chemokine in complex with glycosaminoglycans. Based on the observation of two heparin binding sites, we propose a mechanism in which GAGs bind around CXCL12 dimers as they sequester and present CXCL12 to CXCR4.

  18. Technical advance: Surface plasmon resonance-based analysis of CXCL12 binding using immobilized lentiviral particles.

    PubMed

    Vega, Beatriz; Muñoz, Laura Martínez; Holgado, Borja L; Lucas, Pilar; Rodríguez-Frade, J M; Calle, Ana; Rodríguez-Fernández, J L; Lechuga, Laura M; Rodríguez, José F; Gutiérrez-Gallego, Ricardo; Mellado, Mario

    2011-08-01

    Use of SPR-based biosensors is an established method for measuring molecular interactions. Their application to the study of GPCRs is nonetheless limited to detergent-solubilized receptors that can then be reconstituted into a lipid environment. Using the chemokine receptor CXCR4 and its specific ligand CXCL12, we outline here a highly reproducible biosensor method based on receptor presentation on the surface of lentiviral particles; the approach is simple and does not require the use of antibodies to achieve correct receptor orientation on the sensorchip surface. We measured the kinetic parameters of CXCR4/CXCL12 binding in a single step and in real time and evaluated the effect of GAG presentation of chemokines on this interaction. The data indicate that at low concentrations, soluble heparin modulates CXCR4/CXCL12 interaction and at high concentrations, abrogates binding. These observations suggest that in addition to their known role in modulating local chemokine availability, GAG affect the receptor/ligand interaction, although their influence on affinity parameters is very limited. The method will also be useful for quantifying these biomarkers in biological fluids and for the development of high-throughput screening for their antagonists.

  19. Differential changes in platelet VEGF, Tsp, CXCL12, and CXCL4 in patients with metastatic cancer.

    PubMed

    Wiesner, Tina; Bugl, Stefanie; Mayer, Frank; Hartmann, Jörg T; Kopp, Hans-Georg

    2010-03-01

    Data from animal studies indicate that platelets play a key role in tumor dissemination and metastasis. We therefore hypothesized that metastastic cancer patients may display a specific platelet phenotype. Percentage of activated, p-selectin positive platelets as well as platelet contents (i.e., plasma and platelet count-corrected serum levels of VEGF-A, CXCL12, CXCL4, and thrombospondin-1) were analyzed in 43 patients with newly diagnosed metastatic disease prior to treatment. Tumor patients had increased platelet counts and significantly elevated percentages of activated platelets. Moreover, the platelet content of VEGF-A in cancer patients was significantly increased compared to healthy controls, while thrombospondin-1, CXCL12 and CXCL4 were significantly decreased. Our data contain several unexpected results: firstly, CXCL12 was found in minute quantities in the serum as compared with murine studies. Secondly, CXCL4, which was found by mass spectrometry to be the single massively upregulated intraplatelet chemokine in mice after tumor xenotransplantation, was decreased in tumor patient platelets. While increased contents of VEGF-A have been attributed to platelet scavenger activity, the differential decrease of specific platelet contents may be due to differential secretion or altered megakaryopoiesis in metastatic cancer patients.

  20. Truncation of CXCL12 by CD26 reduces its CXC chemokine receptor 4- and atypical chemokine receptor 3-dependent activity on endothelial cells and lymphocytes.

    PubMed

    Janssens, Rik; Mortier, Anneleen; Boff, Daiane; Ruytinx, Pieter; Gouwy, Mieke; Vantilt, Bo; Larsen, Olav; Daugvilaite, Viktorija; Rosenkilde, Mette M; Parmentier, Marc; Noppen, Sam; Liekens, Sandra; Van Damme, Jo; Struyf, Sofie; Teixeira, Mauro M; Amaral, Flávio A; Proost, Paul

    2017-05-15

    The chemokine CXCL12 or stromal cell-derived factor 1/SDF-1 attracts hematopoietic progenitor cells and mature leukocytes through the G protein-coupled CXC chemokine receptor 4 (CXCR4). In addition, it interacts with atypical chemokine receptor 3 (ACKR3 or CXCR7) and glycosaminoglycans. CXCL12 activity is regulated through posttranslational cleavage by CD26/dipeptidyl peptidase 4 that removes two NH2-terminal amino acids. CD26-truncated CXCL12 does not induce calcium signaling or chemotaxis of mononuclear cells. CXCL12(3-68) was chemically synthesized de novo for detailed biological characterization. Compared to unmodified CXCL12, CXCL12(3-68) was no longer able to signal through CXCR4 via inositol trisphosphate (IP3), Akt or extracellular signal-regulated kinases 1 and 2 (ERK1/2). Interestingly, the recruitment of β-arrestin 2 to the cell membrane via CXCR4 by CXCL12(3-68) was abolished, whereas a weakened but significant β-arrestin recruitment remained via ACKR3. CXCL12-induced endothelial cell migration and signal transduction was completely abrogated by CD26. Intact CXCL12 hardly induced lymphocyte migration upon intra-articular injection in mice. In contrast, oral treatment of mice with the CD26 inhibitor sitagliptin reduced CD26 activity and CXCL12 cleavage in blood plasma. The potential of CXCL12 to induce intra-articular lymphocyte infiltration was significantly increased in sitagliptin-treated mice and CXCL12(3-68) failed to induce migration under both CD26-inhibiting and non-inhibiting conditions. In conclusion, CD26-cleavage skews CXCL12 towards β-arrestin dependent recruitment through ACKR3 and destroys the CXCR4-mediated lymphocyte chemoattractant properties of CXCL12 in vivo. Hence, pharmacological CD26-blockade in tissues may enhance CXCL12-induced inflammation.

  1. CXCR4/CXCL12/CXCR7 axis is functional in neuroendocrine tumors and signals on mTOR.

    PubMed

    Circelli, Luisa; Sciammarella, Concetta; Guadagno, Elia; Tafuto, Salvatore; del Basso de Caro, Marialaura; Botti, Giovanni; Pezzullo, Luciano; Aria, Massimo; Ramundo, Valeria; Tatangelo, Fabiana; Losito, Nunzia Simona; Ieranò, Caterina; D'Alterio, Crescenzo; Izzo, Francesco; Ciliberto, Gennaro; Colao, Annamaria; Faggiano, Antongiulio; Scala, Stefania

    2016-04-05

    To evaluate the possible crosstalk between C-X-C chemokine receptor 4 (CXCR4)/C-X-C motif chemokine 12 (CXCL12)/C-X-C chemokine receptor 7 (CXCR7) axis with the mammalian target of rapamycin (mTOR) pathway in neuroendocrine tumors (NETs). Sixty-one human NETs were included into the study. CXCR4/CXCL12/CXCR7 axis and mTOR pathway were assessed by qRT-PCR and immunohistochemistry (IHC). The effect of mTOR inhibitor, RAD001, was evaluated on CXCR4 pathway through proliferation and p-Erk and p-AKT induction. CXCR4/CXCL12/CXCR7 axis and p-mTOR were found to be active and correlated with grading, Ki67 index and tumor stage. mTOR pathway activation significantly correlated with poor prognosis. In human NET cells, CXCL12 induced mTOR signalling while AMD3100 (CXCR4-antagonist) impaired it. The mTOR-antagonist, RAD001, impaired the CXCL12-dependent induction of CXCR4 downstream effectors. Combination of AMD3100 and RAD001 potentiate cell growth inhibition. CXCR4/CXCL12/CXCR7 axis is active in NETs and signals on mTOR. CXCR4 might be considered a prognostic factor in NETs. Combined treatment with AMD3100 and RAD001 may provide clinical benefits in NET patients with drug-resistant.

  2. CXCR4/CXCL12/CXCR7 axis is functional in neuroendocrine tumors and signals on mTOR

    PubMed Central

    Guadagno, Elia; Tafuto, Salvatore; del Basso de Caro, Marialaura; Botti, Giovanni; Pezzullo, Luciano; Aria, Massimo; Ramundo, Valeria; Tatangelo, Fabiana; Losito, Nunzia Simona; Ieranò, Caterina; D'Alterio, Crescenzo; Izzo, Francesco; Ciliberto, Gennaro; Colao, Annamaria; Faggiano, Antongiulio; Scala, Stefania

    2016-01-01

    Objective To evaluate the possible crosstalk between C-X-C chemokine receptor 4 (CXCR4)/C-X-C motif chemokine 12 (CXCL12)/C-X-C chemokine receptor 7 (CXCR7) axis with the mammalian target of rapamycin (mTOR) pathway in neuroendocrine tumors (NETs). Methods Sixty-one human NETs were included into the study. CXCR4/CXCL12/CXCR7 axis and mTOR pathway were assessed by qRT-PCR and immunohistochemistry (IHC). The effect of mTOR inhibitor, RAD001, was evaluated on CXCR4 pathway through proliferation and p-Erk and p-AKT induction. Results: CXCR4/CXCL12/CXCR7 axis and p-mTOR were found to be active and correlated with grading, Ki67 index and tumor stage. mTOR pathway activation significantly correlated with poor prognosis. In human NET cells, CXCL12 induced mTOR signalling while AMD3100 (CXCR4-antagonist) impaired it. The mTOR-antagonist, RAD001, impaired the CXCL12-dependent induction of CXCR4 downstream effectors. Combination of AMD3100 and RAD001 potentiate cell growth inhibition. Conclusions CXCR4/CXCL12/CXCR7 axis is active in NETs and signals on mTOR. CXCR4 might be considered a prognostic factor in NETs. Combined treatment with AMD3100 and RAD001 may provide clinical benefits in NET patients with drug-resistant. PMID:26934559

  3. Heterologous Quaternary Structure of CXCL12 and its Relationship to the CC Chemokine Family

    SciTech Connect

    Murphy, J.; Yuan, H; Kong, Y; Xiong, Y; Lolis, E

    2010-01-01

    X-ray crystallographic studies reveal that CXCL12 is able to form multiple dimer types, a traditional CXC dimer and a 'CC-like' form. Phylogenetic analysis of all known human chemokines demonstrates CXCL12 is more closely related to the CC chemokine class than other CXC chemokines. These observations indicate that CXCL12 contains genomic and structural elements characteristic of both CXC and CC chemokines.Chemokines are members of a superfamily of proteins involved in the migration of cells to the proper anatomical position during embryonic development or in response to infection or stress during an immune response. There are two major (CC and CXC) and two minor (CX3C and XC) families based on the sequence around the first conserved cysteine. The topology of all structures is essentially identical with a flexible N-terminal region of 3-8 amino acids, a 10-20 residue N-terminal loop, a short 3{sub 10}-helix, three {beta}-strands, and a {alpha}-helix. The major consequence of the subtle difference between the families occurs at the oligomeric level. Monomers of the CC, CXC, and CX3C families form dimers in a family-specific manner. The XCL1 chemokine is a monomer that can interconvert between two folded states. All chemokines activate GPCRs according to family-specificity, however there are a few examples of chemokines crossing the family boundary to function as antagonists. A two-stage mechanism for chemokine activation of GPCRs has been proposed. The N-terminal region of the receptor interacts with the chemokine, followed by receptor activation by the chemokine N-terminal region. Monomeric chemokines have been demonstrated to be the active form for receptor function. There are numerous examples of both chemokines and their receptors forming dimers. While family-specific dimerization may be an attractive explanation for why specific chemokines only activate GPCRs within their own family, the role of dimers in the function of chemokines has not been resolved. Given

  4. The chemokine CXCL12 mediates the anti-amyloidogenic action of painless human nerve growth factor

    PubMed Central

    Capsoni, Simona; Malerba, Francesca; Carucci, Nicola Maria; Rizzi, Caterina; Criscuolo, Chiara; Origlia, Nicola; Calvello, Mariantonietta; Viegi, Alessandro; Meli, Giovanni

    2017-01-01

    Abstract Nerve growth factor is a therapeutic candidate for Alzheimer’s disease. Due to its pain-inducing activity, in current clinical trials nerve growth factor is delivered locally into the brain by neurosurgery, but data on the efficacy of local nerve growth factor delivery in decreasing amyloid-β deposition are not available. To reduce the nerve growth factor pain-inducing side effects, thus avoiding the need for local brain injection, we developed human painless nerve growth factor (hNGFp), inspired by the human genetic disease hereditary sensory and autonomic neuropathy type V. hNGFp has identical neurotrophic potency as wild-type human nerve growth factor, but a 10-fold lower pain sensitizing activity. In this study we first mimicked, in the 5xFAD mouse model, the intraparenchymal delivery of hNGFp used in clinical trials and found it to be ineffective in decreasing amyloid-β plaque load. On the contrary, the same dose of hNGFp delivered intranasally, which was widely biodistributed in the brain and did not induce pain, showed a potent anti-amyloidogenic action and rescued synaptic plasticity and memory deficits. We found that hNGFp acts on glial cells, modulating inflammatory proteins such as the soluble TNFα receptor II and the chemokine CXCL12. We further established that the rescuing effect by hNGFp is mediated by CXCL12, as pharmacological inhibition of CXCL12 receptor CXCR4 occludes most of hNGFp effects. These findings have significant therapeutic implications: (i) we established that a widespread exposure of the brain is required for nerve growth factor to fully exert its neuroprotective actions; and (ii) we have identified a new anti-neurodegenerative pathway as a broad target for new therapeutic opportunities for neurodegenerative diseases. PMID:28031222

  5. MUTZ-3 Langerhans cell maturation and CXCL12 independent migration in reconstructed human gingiva.

    PubMed

    Kosten, Ilona J; Spiekstra, Sander W; de Gruijl, Tanja D; Gibbs, Susan

    2016-01-01

    Here we describe a reconstructed full thickness human oral mucosa (gingiva) equivalent with integrated Langerhans Cells (GE-LC) and use it to compare LC activation and migration from oral versus skin epithelium. The physiologically representative models consist of differentiated reconstructed epithelium (keratinocytes and Langerhans-like cells derived from the MUTZ-3 cell line) on a fibroblast-populated collagen hydrogel which serves as a lamina propria for gingiva and dermis for skin. Topical exposure of GE-LC and the skin equivalent (SE-LC) to sub-toxic concentrations of the allergens cinnamaldehyde, resorcinol and nickel sulphate, resulted in LC migration out of the epithelia. Neutralizing antibody to CXCL12 blocked allergen-induced LC migration in SE-LC but not in GE-LC. Also, gingival fibroblasts secreted very low amounts of CXCL12 compared to skin fibroblasts even when stimulated with rhTNFα or rhIL-1α. Surprisingly, cinnamaldehyde exposure of GE-LC resulted in an increase in MUTZ-3 LC and CD83 mRNA in the hydrogel but did not result in an increase in CD1a+ cells in the collagen hydrogel (as was observed for SE-LC. These results indicate that in gingiva, upon allergen exposure, MUTZ-3 LC migrate in a CXCL12 independent manner from epithelium-to-lamina propria and in doing so mature become CD1a- and increase CD83+ mRNA. These physiologically relevant in vitro models which not only are human but which also resemble specific tissues, may aid in the identification of factors regulating immune stimulation which in turn will aid the development of therapeutic interventions for allergy and inflammation, anti-cancer vaccines as well as improving diagnostics for skin and oral allergy.

  6. The chemokine CXCL12 mediates the anti-amyloidogenic action of painless human nerve growth factor.

    PubMed

    Capsoni, Simona; Malerba, Francesca; Carucci, Nicola Maria; Rizzi, Caterina; Criscuolo, Chiara; Origlia, Nicola; Calvello, Mariantonietta; Viegi, Alessandro; Meli, Giovanni; Cattaneo, Antonino

    2017-01-01

    Nerve growth factor is a therapeutic candidate for Alzheimer's disease. Due to its pain-inducing activity, in current clinical trials nerve growth factor is delivered locally into the brain by neurosurgery, but data on the efficacy of local nerve growth factor delivery in decreasing amyloid-β deposition are not available. To reduce the nerve growth factor pain-inducing side effects, thus avoiding the need for local brain injection, we developed human painless nerve growth factor (hNGFp), inspired by the human genetic disease hereditary sensory and autonomic neuropathy type V. hNGFp has identical neurotrophic potency as wild-type human nerve growth factor, but a 10-fold lower pain sensitizing activity. In this study we first mimicked, in the 5xFAD mouse model, the intraparenchymal delivery of hNGFp used in clinical trials and found it to be ineffective in decreasing amyloid-β plaque load. On the contrary, the same dose of hNGFp delivered intranasally, which was widely biodistributed in the brain and did not induce pain, showed a potent anti-amyloidogenic action and rescued synaptic plasticity and memory deficits. We found that hNGFp acts on glial cells, modulating inflammatory proteins such as the soluble TNFα receptor II and the chemokine CXCL12. We further established that the rescuing effect by hNGFp is mediated by CXCL12, as pharmacological inhibition of CXCL12 receptor CXCR4 occludes most of hNGFp effects. These findings have significant therapeutic implications: (i) we established that a widespread exposure of the brain is required for nerve growth factor to fully exert its neuroprotective actions; and (ii) we have identified a new anti-neurodegenerative pathway as a broad target for new therapeutic opportunities for neurodegenerative diseases.

  7. Aberrant gene expression in deceased transgenic cloned calves.

    PubMed

    Zhang, L; Wang, S H; Dai, Y P; Li, N

    2009-05-01

    Several transgenic cloned species have been obtained; however, the efficiency of transgenic cloning remains very low, even lower than cloning. Many experiments have demonstrated abnormal growth and development, and inappropriate gene expression in cloned animals. In this study, we examined the expression of 19 development-related genes in lungs of three normal controls and three aberrant transgenic cloned calves. Results showed in transgenic cloned calves, 84.2% genes had decreased expression levels, however, 5.3% genes had increased levels. This study suggests transgenic cloning and the aberrant expression would cause abnormal growth and development in transgenic cloned calves. To our knowledge, this is the first time that gene expression was examined in transgenic cloned cattle. These findings may have some implications in understanding the low efficiency of the transgenic cloning.

  8. CXCL12 modulation of CXCR4 and CXCR7 activity in human glioblastoma stem-like cells and regulation of the tumor microenvironment

    PubMed Central

    Würth, Roberto; Bajetto, Adriana; Harrison, Jeffrey K.; Barbieri, Federica; Florio, Tullio

    2014-01-01

    Chemokines are crucial autocrine and paracrine players in tumor development. In particular, CXCL12, through its receptors CXCR4 and CXCR7, affects tumor progression by controlling cancer cell survival, proliferation and migration, and, indirectly, via angiogenesis or recruiting immune cells. Glioblastoma (GBM) is the most prevalent primary malignant brain tumor in adults and despite current multimodal therapies it remains almost incurable. The aggressive and recurrent phenotype of GBM is ascribed to high growth rate, invasiveness to normal brain, marked angiogenesis, ability to escape the immune system and resistance to standard of care therapies. Tumor molecular and cellular heterogeneity severely hinders GBM therapeutic improvement. In particular, a subpopulation of chemo- and radio-therapy resistant tumorigenic cancer stem–like cells (CSCs) is believed to be the main responsible for tumor cell dissemination to the brain. GBM cells display heterogeneous expression levels of CXCR4 and CXCR7 that are overexpressed in CSCs, representing a molecular correlate for the invasive potential of GBM. The microenvironment contribution in GBM development is increasingly emphasized. An interplay exists between CSCs, differentiated GBM cells, and the microenvironment, mainly through secreted chemokines (e.g., CXCL12) causing recruitment of fibroblasts, endothelial, mesenchymal and inflammatory cells to the tumor, via specific receptors such as CXCR4. This review covers recent developments on the role of CXCL12/CXCR4–CXCR7 networks in GBM progression and the potential translational impact of their targeting. The biological and molecular understanding of the heterogeneous GBM cell behavior, phenotype and signaling is still limited. Progress in the identification of chemokine-dependent mechanisms that affect GBM cell survival, trafficking and chemo-attractive functions, opens new perspectives for development of more specific therapeutic approaches that include chemokine

  9. CXCL12 modulation of CXCR4 and CXCR7 activity in human glioblastoma stem-like cells and regulation of the tumor microenvironment.

    PubMed

    Würth, Roberto; Bajetto, Adriana; Harrison, Jeffrey K; Barbieri, Federica; Florio, Tullio

    2014-01-01

    Chemokines are crucial autocrine and paracrine players in tumor development. In particular, CXCL12, through its receptors CXCR4 and CXCR7, affects tumor progression by controlling cancer cell survival, proliferation and migration, and, indirectly, via angiogenesis or recruiting immune cells. Glioblastoma (GBM) is the most prevalent primary malignant brain tumor in adults and despite current multimodal therapies it remains almost incurable. The aggressive and recurrent phenotype of GBM is ascribed to high growth rate, invasiveness to normal brain, marked angiogenesis, ability to escape the immune system and resistance to standard of care therapies. Tumor molecular and cellular heterogeneity severely hinders GBM therapeutic improvement. In particular, a subpopulation of chemo- and radio-therapy resistant tumorigenic cancer stem-like cells (CSCs) is believed to be the main responsible for tumor cell dissemination to the brain. GBM cells display heterogeneous expression levels of CXCR4 and CXCR7 that are overexpressed in CSCs, representing a molecular correlate for the invasive potential of GBM. The microenvironment contribution in GBM development is increasingly emphasized. An interplay exists between CSCs, differentiated GBM cells, and the microenvironment, mainly through secreted chemokines (e.g., CXCL12) causing recruitment of fibroblasts, endothelial, mesenchymal and inflammatory cells to the tumor, via specific receptors such as CXCR4. This review covers recent developments on the role of CXCL12/CXCR4-CXCR7 networks in GBM progression and the potential translational impact of their targeting. The biological and molecular understanding of the heterogeneous GBM cell behavior, phenotype and signaling is still limited. Progress in the identification of chemokine-dependent mechanisms that affect GBM cell survival, trafficking and chemo-attractive functions, opens new perspectives for development of more specific therapeutic approaches that include chemokine

  10. Circulating fibrocytes traffic to the lungs in response to CXCL12 and mediate fibrosis.

    PubMed

    Phillips, Roderick J; Burdick, Marie D; Hong, Kurt; Lutz, Marin A; Murray, Lynne A; Xue, Ying Ying; Belperio, John A; Keane, Michael P; Strieter, Robert M

    2004-08-01

    Previous reports have identified a circulating pool of CD45(+) collagen I(+) CXCR4(+) (CD45(+)Col I(+)CXCR4(+)) cells, termed fibrocytes, that traffic to areas of fibrosis. No studies have demonstrated that these cells actually contribute to fibrosis, however. Pulmonary fibrosis was originally thought to be mediated solely by resident lung fibroblasts. Here we show that a population of human CD45(+)Col I(+)CXCR4(+) circulating fibrocytes migrates in response to CXCL12 and traffics to the lungs in a murine model of bleomycin-induced pulmonary fibrosis. Next, we demonstrated that murine CD45(+)Col I(+)CXCR4(+) fibrocytes also traffic to the lungs in response to a bleomycin challenge. Maximal intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) fibrocytes directly correlated with increased collagen deposition in the lungs. Treatment of bleomycin-exposed animals with specific neutralizing anti-CXCL12 Ab's inhibited intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) circulating fibrocytes and attenuated lung fibrosis. Thus, our results demonstrate, we believe for the first time, that circulating fibrocytes contribute to the pathogenesis of pulmonary fibrosis.

  11. Stromal-derived factor-1α/CXCL12-CXCR4 chemotactic pathway promotes perineural invasion in pancreatic cancer.

    PubMed

    Xu, Qinhong; Wang, Zheng; Chen, Xin; Duan, Wanxing; Lei, Jianjun; Zong, Liang; Li, Xuqi; Sheng, Liang; Ma, Jiguang; Han, Liang; Li, Wei; Zhang, Lun; Guo, Kun; Ma, Zhenhua; Wu, Zheng; Wu, Erxi; Ma, Qingyong

    2015-03-10

    Perineural invasion (PNI) is considered as an alternative route for the metastatic spread of pancreatic cancer cells; however, the molecular changes leading to PNI are still poorly understood. In this study, we show that the CXCL12/CXCR4 axis plays a pivotal role in the neurotropism of pancreatic cancer cells to local peripheral nerves. Immunohistochemical staining results revealed that CXCR4 elevation correlated with PNI in 78 pancreatic cancer samples. Both in vitro and in vivo PNI models were applied to investigate the function of the CXCL12/CXCR4 signaling in PNI progression and pathogenesis. The results showed that the activation of the CXCL12/CXCR4 axis significantly increased pancreatic cancer cells invasion and promoted the outgrowth of the dorsal root ganglia. CXCL12 derived from the peripheral nerves stimulated the invasion and chemotactic migration of CXCR4-positive cancer cells in a paracrine manner, eventually leading to PNI. In vivo analyses revealed that the abrogation of the activated signaling inhibited tumor growth and invasion of the sciatic nerve toward the spinal cord. These data indicate that the CXCL12/CXCR4 axis may be a novel therapeutic target to prevent the perineural dissemination of pancreatic cancer.

  12. Combined Wnt/β-catenin, Met, and CXCL12/CXCR4 signals characterize basal breast cancer and predict disease outcome.

    PubMed

    Holland, Jane D; Györffy, Balázs; Vogel, Regina; Eckert, Klaus; Valenti, Giovanni; Fang, Liang; Lohneis, Philipp; Elezkurtaj, Sefer; Ziebold, Ulrike; Birchmeier, Walter

    2013-12-12

    Prognosis for patients with estrogen-receptor (ER)-negative basal breast cancer is poor, and chemotherapy is currently the best therapeutic option. We have generated a compound-mutant mouse model combining the activation of β-catenin and HGF (Wnt-Met signaling), which produced rapidly growing basal mammary gland tumors. We identified the chemokine system CXCL12/CXCR4 as a crucial driver of Wnt-Met tumors, given that compound-mutant mice also deficient in the CXCR4 gene were tumor resistant. Wnt-Met activation rapidly expanded a population of cancer-propagating cells, in which the two signaling systems control different functions, self-renewal and differentiation. Molecular therapy targeting Wnt, Met, and CXCR4 in mice significantly delayed tumor development. The expression of a Wnt-Met 322 gene signature was found to be predictive of poor survival of human patients with ER-negative breast cancers. Thus, targeting CXCR4 and its upstream activators, Wnt and Met, might provide an efficient strategy for breast cancer treatment.

  13. The miR-25-93-106b cluster regulates tumor metastasis and immune evasion via modulation of CXCL12 and PD-L1.

    PubMed

    Cioffi, Michele; Trabulo, Sara M; Vallespinos, Mireia; Raj, Deepak; Kheir, Tony Bou; Lin, Meng-Lay; Begum, Julfa; Baker, Ann-Marie; Amgheib, Ala; Saif, Jaimy; Perez, Manuel; Soriano, Joaquim; Desco, Manuel; Gomez-Gaviro, Maria Victoria; Cusso, Lorena; Megias, Diego; Aicher, Alexandra; Heeschen, Christopher

    2017-03-28

    The stromal microenvironment controls response to injury and inflammation, and is also an important determinant of cancer cell behavior. However, our understanding of its modulation by miRNA (miR) and their respective targets is still sparse. Here, we identified the miR-25-93-106b cluster and two new target genes as critical drivers for metastasis and immune evasion of cancer cells. Using miR-25-93-106b knockout mice or antagomiRs, we demonstrated regulation of the production of the chemoattractant CXCL12 controlling bone marrow metastasis. Moreover, we identified the immune checkpoint PD-L1 (CD274) as a novel miR-93/106b target playing a central role in diminishing tumor immunity. Eventually, upregulation of miR-93 and miR-106b via miR-mimics or treatment with an epigenetic reader domain (BET) inhibitor resulted in diminished expression of CXCL12 and PD-L1. These data suggest a potential new therapeutic rationale for use of BET inhibitors for dual targeting of cancers with strong immunosuppressive and metastatic phenotypes.

  14. Identification and characterization of circulating variants of CXCL12 from human plasma: effects on chemotaxis and mobilization of hematopoietic stem and progenitor cells.

    PubMed

    Richter, Rudolf; Jochheim-Richter, Andrea; Ciuculescu, Felicia; Kollar, Katarina; Seifried, Erhard; Forssmann, Ulf; Verzijl, Dennis; Smit, Martine J; Blanchet, Xavier; von Hundelshausen, Philipp; Weber, Christian; Forssmann, Wolf-Georg; Henschler, Reinhard

    2014-08-15

    Mobilization of hematopoietic stem and progenitor cells (HPCs) is induced by treatment with granulocyte-colony stimulating factor, chemotherapy, or irradiation. We observed that these treatments are accompanied by a release of chemotactic activity into the blood. This plasma activity is derived from the bone marrow, liver, and spleen and acts on HPCs via the chemokine receptor CXCR4. A human blood peptide library was used to characterize CXCR4-activating compounds. We identified CXCL12[22-88] and N-terminally truncated variants CXCL12[24-88], CXCL12[25-88], CXCL12[27-88], and CXCL12[29-88]. Only CXCL12[22-88] could effectively bind to CXCR4 and induce intracellular calcium flux and chemotactic migration of HPCs. CXCL12[25-88] and CXCL12[27-88] revealed neither agonistic nor antagonistic activities in vitro, whereas CXCL12[29-88] inhibited CXCL12[22-88]-induced chemotactic migration. Since binding to glycosaminoglycans (GAG) modulates the function of CXCL12, binding to heparin was analyzed. Surface plasmon resonance kinetic analysis showed that N-terminal truncation of Arg22-Pro23 increased the dissociation constant KD by one log10 stage ([22-88]: KD: 5.4 ± 2.6 μM; [24-88]: KD: 54 ± 22.4 μM). Further truncation of the N-terminus decreased the KD ([25-88] KD: 30 ± 4.8 μM; [27-88] KD: 23 ± 1.6 μM; [29-88] KD: 19 ± 5.4 μM), indicating increasing competition for heparin binding. Systemic in vivo application of CXCL12[22-88] as well as CXCL12[27-88] or CXCL12[29-88] induced a significant mobilization of HPCs in mice. Our findings indicate that plasma-derived CXCL12 variants may contribute to the regulation of HPC mobilization by modulating the binding of CXCL12[22-88] to GAGs rather than blocking the CXCR4 receptor and, therefore, may have a contributing role in HPC mobilization.

  15. Identification and Characterization of Circulating Variants of CXCL12 from Human Plasma: Effects on Chemotaxis and Mobilization of Hematopoietic Stem and Progenitor Cells

    PubMed Central

    Jochheim-Richter, Andrea; Ciuculescu, Felicia; Kollar, Katarina; Seifried, Erhard; Forssmann, Ulf; Verzijl, Dennis; Smit, Martine J.; Blanchet, Xavier; von Hundelshausen, Philipp; Weber, Christian; Forssmann, Wolf-Georg; Henschler, Reinhard

    2014-01-01

    Mobilization of hematopoietic stem and progenitor cells (HPCs) is induced by treatment with granulocyte-colony stimulating factor, chemotherapy, or irradiation. We observed that these treatments are accompanied by a release of chemotactic activity into the blood. This plasma activity is derived from the bone marrow, liver, and spleen and acts on HPCs via the chemokine receptor CXCR4. A human blood peptide library was used to characterize CXCR4-activating compounds. We identified CXCL12[22–88] and N-terminally truncated variants CXCL12[24–88], CXCL12[25–88], CXCL12[27–88], and CXCL12[29–88]. Only CXCL12[22–88] could effectively bind to CXCR4 and induce intracellular calcium flux and chemotactic migration of HPCs. CXCL12[25–88] and CXCL12[27–88] revealed neither agonistic nor antagonistic activities in vitro, whereas CXCL12[29–88] inhibited CXCL12[22–88]-induced chemotactic migration. Since binding to glycosaminoglycans (GAG) modulates the function of CXCL12, binding to heparin was analyzed. Surface plasmon resonance kinetic analysis showed that N-terminal truncation of Arg22-Pro23 increased the dissociation constant KD by one log10 stage ([22–88]: KD: 5.4±2.6 μM; [24–88]: KD: 54±22.4 μM). Further truncation of the N-terminus decreased the KD ([25–88] KD: 30±4.8 μM; [27–88] KD: 23±1.6 μM; [29–88] KD: 19±5.4 μM), indicating increasing competition for heparin binding. Systemic in vivo application of CXCL12[22–88] as well as CXCL12[27–88] or CXCL12[29–88] induced a significant mobilization of HPCs in mice. Our findings indicate that plasma-derived CXCL12 variants may contribute to the regulation of HPC mobilization by modulating the binding of CXCL12[22–88] to GAGs rather than blocking the CXCR4 receptor and, therefore, may have a contributing role in HPC mobilization. PMID:24708339

  16. SDF-1α/CXCL12 enhances GABA and glutamate synaptic activity at serotonin neurons in the rat dorsal raphe nucleus

    PubMed Central

    Heinisch, Silke; Kirby, Lynn G.

    2009-01-01

    Summary The serotonin (5-hydroxytryptamine; 5-HT) system has a well-characterized role in depression. Recent reports describe comorbidities of mood-immune disorders, suggesting an immunological component may contribute to the pathogenesis of depression as well. Chemokines, immune proteins which mediate leukocyte trafficking, and their receptors are widely distributed in the brain, mediate neuronal patterning, and modulate various neuropathologies. The purpose of this study was to investigate the neuroanatomical relationship and functional impact of the chemokine stromal cell-derived factor-1α/CXCL12 and its receptor, CXCR4, on the serotonin dorsal raphe nucleus (DRN) system in the rat using anatomical and electrophysiological techniques. Immunohistochemical analysis indicates that over 70% of 5-HT neurons colocalize with CXCL12 and CXCR4. At a subcellular level, CXCL12 localizes throughout the cytoplasm whereas CXCR4 concentrates to the outer membrane and processes of 5-HT neurons. CXCL12 and CXCR4 also colocalize on individual DRN cells. Furthermore, electrophysiological studies demonstrate CXCL12 depolarization of 5-HT neurons indirectly via glutamate synaptic inputs. CXCL12 also enhances the frequency of spontaneous inhibitory and excitatory postsynaptic currents (sIPSC and sEPSC). CXCL12 concentration-dependently increases evoked IPSC amplitude and decreases evoked IPSC paired-pulse ratio selectively in 5-HT neurons, effects blocked by the CXCR4 antagonist AMD3100. These data indicate presynaptic enhancement of GABA and glutamate release at 5-HT DRN neurons by CXCL12. Immunohistochemical analysis further shows CXCR4 localization to DRN GABA neurons, providing an anatomical basis for CXCL12 effects on GABA release. Thus, CXCL12 indirectly modulates 5-HT neurotransmission via GABA and glutamate synaptic afferents. Future therapies targeting CXCL12 and other chemokines may treat serotonin related mood disorders, particularly depression experienced by immune

  17. E-cadherin Is Critical for Collective Sheet Migration and Is Regulated by the Chemokine CXCL12 Protein During Restitution*

    PubMed Central

    Hwang, Soonyean; Zimmerman, Noah P.; Agle, Kimberle A.; Turner, Jerrold R.; Kumar, Suresh N.; Dwinell, Michael B.

    2012-01-01

    Chemokines and other immune mediators enhance epithelial barrier repair. The intestinal barrier is established by highly regulated cell-cell contacts between epithelial cells. The goal of these studies was to define the role for the chemokine CXCL12 in regulating E-cadherin during collective sheet migration during epithelial restitution. Mechanisms regulating E-cadherin were investigated using Caco2BBE and IEC-6 model epithelia. Genetic knockdown confirmed a critical role for E-cadherin in in vitro restitution and in vivo wound repair. During restitution, both CXCL12 and TGF-β1 tightened the monolayer by decreasing the paracellular space between migrating epithelial cells. However, CXCL12 differed from TGF-β1 by stimulating the significant increase in E-cadherin membrane localization during restitution. Chemokine-stimulated relocalization of E-cadherin was paralleled by an increase in barrier integrity of polarized epithelium during restitution. CXCL12 activation of its cognate receptor CXCR4 stimulated E-cadherin localization and monolayer tightening through Rho-associated protein kinase activation and F-actin reorganization. These data demonstrate a key role for E-cadherin in intestinal epithelial restitution. PMID:22549778

  18. Applications of stripe assay in the study of CXCL12-mediated neural progenitor cell migration and polarization.

    PubMed

    Zhang, Min; Song, Aihong; Lai, Siqiang; Qiu, Lisha; Huang, Yunlong; Chen, Qiang; Zhu, Bing; Xu, Dongsheng; Zheng, Jialin C

    2015-12-01

    The polarization and migration of neural progenitor cells (NPCs) are critical for embryonic brain development and neurogenesis after brain injury. Although stromal-derived factor-1α (SDF-1α, CXCL12) and its receptor CXCR4 are well-known to mediate the migration of NPCs in the developing brain, the dynamic cellular processes and structure-related molecular events remain elusive. Transwell and microfluidic-based assays are classical assays to effectively study cellular migration. However, both of them have limitations in the analysis of a single cell. In this study, we modified the stripe assay and extended its applications in the study of NPC polarization and intracellular molecular events associated with CXCL12-mediated migration. In response to localized CXCL12, NPCs formed lamellipodia in the stripe assay. Furthermore, CXCR4 and Rac1 quickly re-distributed to the area of lamellipodia, indicating their roles in NPC polarization upon CXCL12 stimulation. Although the chemokine stripes in the assay provided concentration gradients that can be best used to study cellular polarization and migration through immunocytochemistry, they can also generate live imaging data with comparable quality. In conclusion, stripe assay is a visual, dynamic and economical tool to study cellular mobility and its related molecule mechanisms. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. CXCL12/CXCR4 blockade by oncolytic virotherapy inhibits ovarian cancer growth by decreasing immunosuppression and targeting cancer-initiating cells.

    PubMed

    Gil, Margaret; Komorowski, Marcin P; Seshadri, Mukund; Rokita, Hanna; McGray, A J Robert; Opyrchal, Mateusz; Odunsi, Kunle O; Kozbor, Danuta

    2014-11-15

    Signals mediated by the chemokine CXCL12 and its receptor CXCR4 are involved in the progression of ovarian cancer through enhancement of tumor angiogenesis and immunosuppressive networks that regulate dissemination of peritoneal metastasis and development of cancer-initiating cells (CICs). In this study, we investigated the antitumor efficacy of a CXCR4 antagonist expressed by oncolytic vaccinia virus (OVV) against an invasive variant of the murine epithelial ovarian cancer cell line ID8-T. This variant harbors a high frequency of CICs that form multilayered spheroid cells and express the hyaluronan receptor CD44, as well as stem cell factor receptor CD117 (c-kit). Using an orthotopic ID8-T tumor model, we observed that i.p. delivery of a CXCR4 antagonist-expressing OVV led to reduced metastatic spread of tumors and improved overall survival compared with oncolysis alone. Inhibition of tumor growth with the armed virus was associated with efficient killing of CICs, reduced expression of ascitic CXCL12 and vascular endothelial growth factor, and decreases in i.p. numbers of endothelial and myeloid cells, as well as plasmacytoid dendritic cells. These changes, together with reduced recruitment of T regulatory cells, were associated with higher ratios of IFN-γ(+)/IL-10(+) tumor-infiltrating T lymphocytes, as well as induction of spontaneous humoral and cellular antitumor responses. Similarly, the CXCR4 antagonist released from virally infected human CAOV2 ovarian carcinoma cells inhibited peritoneal dissemination of tumors in SCID mice, leading to improved tumor-free survival in a xenograft model. Our findings demonstrate that OVV armed with a CXCR4 antagonist represents a potent therapy for ovarian CICs with a broad antitumor repertoire.

  20. CXCL12/CXCR4 Blockade by Oncolytic Virotherapy Inhibits Ovarian Cancer Growth by Decreasing Immunosuppression and Targeting Cancer Initiating Cells1

    PubMed Central

    Gil, Margaret; Komorowski, Marcin P.; Seshadri, Mukund; Rokita, Hanna; McGray, A. J Robert; Opyrchal, Mateusz; Odunsi, Kunle O.; Kozbor, Danuta

    2014-01-01

    Signals mediated by the chemokine CXCL12 and its receptor CXCR4 are involved in progression of ovarian cancer by enhancing tumor angiogenesis and immunosuppressive networks that regulate dissemination of peritoneal metastasis and development of cancer initiating cells (CICs). Here, we investigated the antitumor efficacy of a CXCR4 antagonist expressed by oncolytic vaccinia virus (OVV) against an invasive variant of the murine epithelial ovarian cancer cell line ID8-T. This variant harbors a high frequency of CICs that form multilayered spheroid cells and express the hyaluronan receptor CD44 as well as stem cell factor receptor CD117 (c-kit). Using an orthotopic ID8-T tumor model, we observed that intraperitoneal delivery of a CXCR4 antagonist-expressing OVV led to reduced metastatic spread of tumors and improved overall survival over that mediated by oncolysis alone. Inhibition of tumor growth with the armed virus was associated with efficient killing of CICs, reductions in expression of ascitic CXCL12 and VEGF, and decreases in intraperitoneal numbers of endothelial and myeloid cells as well as plasmacytoid dendritic cells (pDCs). These changes, together with reduced recruitment of T regulatory cells, were associated with higher ratios of IFN-γ+/IL-10+ tumor-infiltrating T lymphocytes as well as induction of spontaneous humoral and cellular antitumor responses. Similarly, the CXCR4 antagonist released from virally-infected human CAOV2 ovarian carcinoma cells inhibited peritoneal dissemination of tumors in SCID mice leading to improved tumor-free survival in a xenograft model. Our findings demonstrate that OVV armed with a CXCR4 antagonist represents a potent therapy for ovarian CICs with a broad antitumor repertoire. PMID:25320277

  1. Do truncated cyclins contribute to aberrant cyclin expression in cancer?

    PubMed

    Van Dross, Rukiyah; Browning, Philip J; Pelling, Jill C

    2006-03-01

    Cyclin overexpression is found in several types of cancer. Genetic events that place cyclin genes under the control of active promoters or that increase cyclin gene copy number account for most instances of cyclin overexpression. New paradigms for aberrant cyclin expression have been suggested by studies showing that truncated cyclins are expressed in specific subsets of cancer. The altered cyclins lack regulatory sequences (compared to the wild-type protein) that modulate their stability, subcellular localization or cdk-associated kinase activity. In this communication, we review the current literature and assess the role of truncated cyclins D, E, A, B, C and virus encoded-cyclin D (K-cyclin) in the development of cancer. We also report the molecular characteristics, expression patterns and if available, prognostic significance of these proteins.

  2. CXCL12-CXCR4 Axis Is Required for Contact-Mediated Human B Lymphoid and Plasmacytoid Dendritic Cell Differentiation but Not T Lymphoid Generation.

    PubMed

    Minami, Hirohito; Nagaharu, Keiki; Nakamori, Yoshiki; Ohishi, Kohshi; Shimojo, Naoshi; Kageyama, Yuki; Matsumoto, Takeshi; Sugimoto, Yuka; Tawara, Isao; Masuya, Masahiro; Miwa, Hiroshi; Katayama, Naoyuki

    2017-10-01

    We investigated the involvement of CXCL12-CXCR4 interactions in human lymphohematopoiesis by coculture with telomerized human stromal cells. CXCR4 expression was low in CD34(+)CD38(-)CD45RA(-)CD10(-)CD7(-)CD19(-) immature hematopoietic stem/precursor cells (HSPCs) but higher in CD34(+)CD38(-)CD45RA(+)CD10(+)CD7(+/-)CD19(-) early lymphoid precursors and even higher in CD34(+)CD38(+)CD45RA(+)CD10(+)CD7(-)CD19(+) pro-B cells. Inhibition of the effect of stromal cell-produced CXCL12 by an anti-CXCR4-blocking Ab suppressed the generation of CD45RA(+)CD10(-)CD7(+)CD19(-) early T lymphoid precursors (ETPs) and CD45RA(+)CD10(+)CD7(-)CD19(+/-) B lymphoid precursors on stromal cells, but it did not affect the generation of ETPs in conditioned medium of stromal cell cultures. Replating assays showed that contact with stromal cells was critical for HSPC-derived CD45RA(+)CD10(+)CD7(-)CD19(-) B lineage-biased precursors to differentiate into CD19(+) pro-B cells, which was suppressed by the anti-CXCR4 Ab. Conversely, HSPC-derived ETPs possessed T and B lymphoid and monocytic differentiation potential; stromal cell contact was not required for their growth but rather promoted B lymphoid differentiation. The anti-CXCR4 Ab did not affect the growth of ETPs in conditioned medium, but it suppressed their B lymphoid differentiation on stromal cells. CD14(-)CD11c(-)HLA-DR(+)CD123(high)CD303(+) plasmacytoid dendritic cells developed from HSPCs and ETPs exclusively in contact with stromal cells, which was suppressed by the anti-CXCR4 Ab. These data indicate that CXCL12 plays an essential role in stromal cell contact-mediated B lymphoid and plasmacytoid dendritic cell differentiation from immature hematopoietic and early T lymphoid precursors with a multilineage differentiation potential, but it does not participate in contact-independent generation of early T lymphoid precursors. Copyright © 2017 by The American Association of Immunologists, Inc.

  3. Aberrant expression of homeobox gene SIX1 in Hodgkin lymphoma

    PubMed Central

    Nagel, Stefan; Meyer, Corinna; Kaufmann, Maren; Drexler, Hans G.; MacLeod, Roderick A.F.

    2015-01-01

    In Hodgkin lymphoma (HL) we recently identified deregulated expression of homeobox genes MSX1 and OTX2 which are physiologically involved in development of the embryonal neural plate border region. Here, we examined in HL homeobox gene SIX1 an additional regulator of this embryonal region mediating differentiation of placodal precursors. SIX1 was aberrantly activated in 12 % of HL patient samples in silico, indicating a pathological role in a subset of this B-cell malignancy. In addition, SIX1 expression was detected in HL cell lines which were used as models to reveal upstream factors and target genes of this basic developmental regulator. We detected increased copy numbers of the SIX1 locus at chromosome 14q23 correlating with enhanced expression while chromosomal translocations were absent. Moreover, comparative expression profiling data and pertinent gene modulation experiments indicated that the WNT-signalling pathway and transcription factor MEF2C regulate SIX1 expression. Genes encoding the transcription factors GATA2, GATA3, MSX1 and SPIB – all basic lymphoid regulators - were identified as targets of SIX1 in HL. In addition, cofactors EYA1 and TLE4, respectively, contrastingly mediated activation and suppression of SIX1 target gene expression. Thus, the protein domain interfaces may represent therapeutic targets in SIX1-positive HL subsets. Collectively, our data reveal a gene regulatory network with SIX1 centrally deregulating lymphoid differentiation and support concordance of lymphopoiesis/lymphomagenesis and developmental processes in the neural plate border region. PMID:26473286

  4. Drug design strategies focusing on the CXCR4/CXCR7/CXCL12 pathway in leukemia and lymphoma.

    PubMed

    Barbieri, Federica; Bajetto, Adriana; Thellung, Stefano; Würth, Roberto; Florio, Tullio

    2016-11-01

    Chemokines control homing and trafficking of leukocytes in bone marrow and lymphoid organs. In particular, CXCL12 and its receptors CXCR4/CXCR7 control the homeostasis of multiple organs and systems. Their overexpression is linked to tumor development, both through a direct modulation of neoplastic cell proliferation, survival, and migration, and, indirectly, acting on the tumor microenvironment which sustains drug resistant tumor stem-like cells. Leukemia and lymphomas frequently display upregulation of CXCL12/CXCR4 in bone marrow that nurtures tumor cells, and confers resistance to conventional chemotherapy, increasing disease relapse. Areas covered: The authors review the molecular and cellular mechanisms by which the CXCL12/CXCR4-7 system supports leukemic bone marrow and how it contributes to leukemia development, and their potential pharmacological targeting. Besides receptor antagonists that directly inhibit leukemic cell proliferation, preclinical and clinical studies demonstrate that CXCR4 inhibition mobilizes leukemic-lymphoma cells from their niches, improving conventional chemotherapy efficacy. Clinically available and experimental pharmacological tools targeting CXCR4/CXCR7 are also described. Expert opinion: Studies have revealed the therapeutic efficacy of combining CXCR4 inhibitors and cytotoxic agents to sensitize leukemic cells, and overcome natural or acquired resistance. However, several issues are still to be unveiled (for example the role of CXCR7) to maximize therapeutic response and reduce potential toxicities.

  5. Improving chemotherapeutic efficiency in acute myeloid leukemia treatments by chemically synthesized peptide interfering with CXCR4/CXCL12 axis

    PubMed Central

    Li, Xiaojin; Guo, Hua; Duan, Hongyang; Yang, Yanlian; Meng, Jie; Liu, Jian; Wang, Chen; Xu, Haiyan

    2015-01-01

    Bone marrow stroma can protect acute myeloid leukemia (AML) cells against chemotherapeutic agents and provide anti-apoptosis and chemoresistance signals through secreting chemokine CXCL12 to activate its receptor CXCR4 on AML cells, resulting in minimal residual leukemia and relapse. Therefore disrupting the CXCR4/CXCL12 axis with antagonists is of great significance for improving chemosensitivity and decreasing relapse rate. In a previous study, we reported a novel synthetic peptide E5 with its remarkable effect on inhibiting CXCR4/CXCL12-mediated adhesion and migration of AML cells. Here we presented E5’s capacity of enhancing the therapeutic efficiency of various chemotherapeutics on AML in vitro and in vivo. Results showed that E5 can diminish bone marrow stromal cell-provided protection to leukemia cells, significantly increasing the apoptosis induced by various chemotherapeutics in multiple AML cell lines. In an AML mouse xenograft model, E5 induced 1.84-fold increase of circulating AML cells out of protective stroma niche. Combined with vincristine or cyclophosphamide, E5 inhibited infiltration of AML cells into bone marrow, liver and spleen, as well as prolonged the lifespan of AML mice compared with mice treated with chemotherapy alone. In addition, E5 presented no toxicity in vivo according to the histological analysis and routine clinical parameters of serum analysis. PMID:26538086

  6. Pharmacologically antagonizing the CXCR4-CXCL12 chemokine pathway with AMD3100 inhibits sunlight-induced skin cancer.

    PubMed

    Sarchio, Seri N E; Scolyer, Richard A; Beaugie, Clare; McDonald, David; Marsh-Wakefield, Felix; Halliday, Gary M; Byrne, Scott N

    2014-04-01

    One way sunlight causes skin cancer is by suppressing anti-tumor immunity. A major mechanism involves altering mast cell migration via the C-X-C motif chemokine receptor 4-C-X-C motif chemokine ligand 12 (CXCR4-CXCL12) chemokine pathway. We have discovered that pharmacologically blocking this pathway with the CXCR4 antagonist AMD3100 prevents both UV radiation-induced immune suppression and skin cancer. The majority of control mice receiving UV-only developed histopathologically confirmed squamous cell carcinomas. In contrast, skin tumor incidence and burden was significantly lower in AMD3100-treated mice. Perhaps most striking was that AMD3100 completely prevented the outgrowth of latent tumors that occurred once UV irradiation ceased. AMD3100 protection from UV immunosuppression and skin cancer was associated with reduced mast cell infiltration into the skin, draining lymph nodes, and the tumor itself. Thus a major target of CXCR4 antagonism was the mast cell. Our results indicate that interfering with UV-induced CXCL12 by antagonizing CXCR4 significantly inhibits skin tumor development by blocking UV-induced effects on mast cells. Hence, the CXCR4-CXCL12 chemokine pathway is a novel therapeutic target in the prevention of UV-induced skin cancer.

  7. Aberrant expression of RUNX3 in patients with immune thrombocytopenia.

    PubMed

    Qiao, Jianlin; Liu, Yun; Wu, Yulu; Li, Xiaoqian; Zhu, Feng; Xia, Yuan; Yao, Haina; Chu, Peipei; Li, Hongchun; Ma, Ping; Li, Depeng; Li, Zhenyu; Xu, Kailin; Zeng, Lingyu

    2015-09-01

    Immune thrombocytopenia (ITP) is an autoimmune disease, characterized by dysregulation of cellular immunity. Previous studies demonstrated that immune imbalance between Th1 and Th2 was associated with the pathogenesis of ITP. Runt-related transcription factor 3 (RUNX3) is a member of the runt domain-containing family of transcription factors and plays an important role in the regulation of T cell differentiation into Th1 cells. Whether RUNX3 was involved in the pathogenesis of ITP remains unclear. In this study, 47 active ITP patients, 18 ITP with remission and 26 age and gender matched healthy control were included. Peripheral blood mononuclear cells (PBMCs) were isolated from ITP and control for isolation of RNA and plasma which were used to measure mRNA level of RUNX3 and T-box transcription factor (T-bet) by quantitative real-time PCR and interferon γ (IFN-γ) plasma level by ELISA. Meanwhile, protein was also extracted from PBMCs for Western blot analysis of RUNX3 expression. Our results showed a significantly higher expression of RUNX3, T-bet and plasma level of IFN-γ in active ITP patients compared to control. No differences were observed between ITP with remission and control. Furthermore, a positive correlation of RUNX3 with T-bet was found in active ITP patients. In conclusion, aberrant expression of RUNX3 was associated with the pathogenesis of ITP and therapeutically targeting it might be a novel approach in ITP treatment.

  8. CXCR4-CXCL12-CXCR7, TLR2-TLR4, and PD-1/PD-L1 in colorectal cancer liver metastases from neoadjuvant-treated patients.

    PubMed

    D'Alterio, Crescenzo; Nasti, Guglielmo; Polimeno, Marianeve; Ottaiano, Alessandro; Conson, Manuel; Circelli, Luisa; Botti, Giovanni; Scognamiglio, Giosuè; Santagata, Sara; De Divitiis, Chiara; Nappi, Anna; Napolitano, Maria; Tatangelo, Fabiana; Pacelli, Roberto; Izzo, Francesco; Vuttariello, Emilia; Botti, Gerardo; Scala, Stefania

    2016-01-01

    A neoadjuvant clinical trial was previously conducted in patients with resectable colorectal cancer liver metastases (CRLM). At a median follow up of 28 months, 20/33 patients were dead of disease, 8 were alive with disease and 5 were alive with no evidence of disease. To shed further insight into biological features accounting for different outcomes, the expression of CXCR4-CXCL12-CXCR7, TLR2-TLR4, and the programmed death receptor-1 (PD-1)/programmed death-1 ligand (PD-L1) was evaluated in excised liver metastases. Expression profiles were assessed through qPCR in metastatic and unaffected liver tissue of 33 CRLM neoadjuvant-treated patients. CXCR4 and CXCR7, TLR2/TLR4, and PD-1/PD-L1 mRNA were significantly overexpressed in metastatic compared to unaffected liver tissues. CXCR4 protein was negative/low in 10/31, and high in 21/31, CXCR7 was negative/low in 16/31 and high in 15/31, CXCL12 was negative/low in 14/31 and high in 17/31 CRLM. PD-1 was negative in 19/30 and positive in 11/30, PD-L1 was negative/low in 24/30 and high in 6/30 CRLM. Stromal PD-L1 expression, affected the progression-free survival (PFS) in the CRLM population. Patients overexpressing CXCR4 experienced a worse PFS and cancer specific survival (CSS) (p = 0.001 and p = 0.0008); in these patients, KRAS mutation identified a subgroup with a significantly worse CSS (p < 0.01). Thus, CXCR4 and PD-L1 expression discriminate patients with the worse PFS within the CRLM evaluated patients. Within the CXCR4 high expressing patients carrying Mut-KRAS in CRLM identifies the worst prognostic group. Thus, CXCR4 targeting plus anti-PD-1 therapy should be explored to improve the prognosis of Mut-KRAS-high CXCR4-CRLMs.

  9. CXCR4/CXCL12 Participate in Extravasation of Metastasizing Breast Cancer Cells within the Liver in a Rat Model

    PubMed Central

    Krasnyanska, Julia; Mees, Sören Torge; Kochetkova, Marina; Stoeppeler, Sandra; Haier, Jörg

    2012-01-01

    Introduction Organ-specific composition of extracellular matrix proteins (ECM) is a determinant of metastatic host organ involvement. The chemokine CXCL12 and its receptor CXCR4 play important roles in the colonization of human breast cancer cells to their metastatic target organs. In this study, we investigated the effects of chemokine stimulation on adhesion and migration of different human breast cancer cell lines in vivo and in vitro with particular focus on the liver as a major metastatic site in breast cancer. Methods Time lapse microscopy, in vitro adhesion and migration assays were performed under CXCL12 stimulation. Activation of small GTPases showed chemokine receptor signalling dependence from ECM components. The initial events of hepatic colonisation of MDA-MB-231 and MDA-MB-468 cells were investigated by intravital microscopy of the liver in a rat model and under shRNA inhibition of CXCR4. Results In vitro, stimulation with CXCL12 induced increased chemotactic cell motility (p<0.05). This effect was dependent on adhesive substrates (type I collagen, fibronectin and laminin) and induced different responses in small GTPases, such as RhoA and Rac-1 activation, and changes in cell morphology. In addition, binding to various ECM components caused redistribution of chemokine receptors at tumour cell surfaces. In vivo, blocking CXCR4 decreased extravasation of highly metastatic MDA-MB-231 cells (p<0.05), but initial cell adhesion within the liver sinusoids was not affected. In contrast, the less metastatic MDA-MB-468 cells showed reduced cell adhesion but similar migration within the hepatic microcirculation. Conclusion: Chemokine-induced extravasation of breast cancer cells along specific ECM components appears to be an important regulator but not a rate-limiting factor of their metastatic organ colonization. PMID:22253872

  10. CXCL12/CXCR4 axis confers adriamycin resistance to human chronic myelogenous leukemia and oroxylin A improves the sensitivity of K562/ADM cells.

    PubMed

    Wang, Yu; Miao, Hanchi; Li, Wei; Yao, Jing; Sun, Yang; Li, Zhiyu; Zhao, Li; Guo, Qinglong

    2014-08-01

    This study was aimed at investigating the reversal effect of oroxylin A, a naturally bioactive monoflavonoid separated and purified from Scutellaria baicalensis Georgi, in human chronic myeloid leukemia (CML) and the underlying mechanisms. The results showed that CXCL12 could enhance the resistance of K562 cells to adriamycin (ADM) by increasing the expression of CXCR4, up-regulating the downstream PI3K/Akt pathway, and promoting translocation of NF-κB dimers into nucleus and subsequently decreasing the expression of apoptosis-related proteins in K562 cells. And we found that ADM resistance was partially reversed by CXCR4 siRNA transfection. Moreover, the sensitivity enhancement of oroxylin A was demonstrated by decreasing the expression of CXCR4 at both protein and mRNA levels, via PI3K/Akt/NF-κB pathway and triggering the apoptosis pathway in vitro. In addition, the in vivo study showed that oroxylin A increased apoptosis of leukemic cells with low systemic toxicity, and the mechanism was the same as in vitro study. In conclusion, all these results showed that oroxylin A improved the sensitivity of K562/ADM cells by increasing apoptosis in leukemic cells and decreasing the expression of CXCR4 and PI3K/Akt/NF-κB pathway, and probably served as a most promising agent for CML treatment. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. NF-κB2 Controls Migratory Activity of Memory T Cells by Regulating Expression of CXCR4 in A Mouse Model of Sjögren's Syndrome.

    PubMed

    Kurosawa, Mie; Arakaki, Rieko; Yamada, Akiko; Tsunematsu, Takaaki; Kudo, Yasusei; Sprent, Jonathan; Ishimaru, Naozumi

    2017-08-13

    Dysregulated chemokine signaling contributes to autoimmune diseases by facilitating aberrant T-cell infiltration into target tissues, but the specific chemokines, receptors, and T-cell populations remain largely unidentified. Role of the potent chemokine CXCL12 and its receptor CXCR4 in T-cell autoimmune response was examined using alymphoplasia (aly)/aly mice, a Sjögren's syndrome (SS) model. T-cell phenotypes in the salivary gland of aly/aly mice were evaluated using immunological analysis. In vitro migration assay was used to assess T-cell migratory activity toward several chemokines. Gene expression of chemokine receptors, and transforming growth factor (TGF)β receptors was measured with quantitative reverse transcription-polymerase chain reaction. The CXCR4 antagonist AMD3100 was administered to the aly/aly mice to evaluate its suppressive effect on autoimmune lesions. Effector memory T (TEM) cells derived from aly/aly mice demonstrated higher in vitro migratory activity toward CXCL12 than aly/+ TEM cells. CXCL12 expression was specifically upregulated in the SS target cells of aly/aly mice. TEM cells from RelB(-/-) mice, but not nuclear factor (NF)-κB1(-/-) mice, also showed high migratory activity toward CXCL12, implicating a nonclassical NF-κB2/RelB pathway in the regulation of TEM cell migration. TEM cells from aly/aly mice also overexpressed TGFβ receptors I and II. The CXCR4 antagonist AMD3100 suppressed autoimmune lesions in aly/aly mice by reducing TEM cell infiltration. Our results suggest that the NF-κB2/RelB pathway regulates T-cell migration to autoimmune targets through TGFβ/TGFβR-dependent regulation of CXCL12-CXCR4 signaling. This suggests that these signaling pathways are potential therapeutic targets for treating autoimmune diseases. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  12. Serum ENA78/CXCL5, SDF-1/CXCL12, and their combinations as potential biomarkers for prediction of the presence and distant metastasis of primary gastric cancer.

    PubMed

    Lim, Jong-Baeck; Chung, Hye Won

    2015-05-01

    Chemokines play important roles in cancer development and progression. Epithelial-derived neutrophil-activating peptide-78 (ENA78/CXCL5) and stromal cell-derived factor (SDF-1/CXCL12) supposedly contribute to gastric cancer (GC) development and progression. This study aims to evaluate serum levels of ENA78/CXCL5 and SDF-1/CXCL12 along the GC carcinogenesis, and analyze their clinical significance, and diagnostic potentials through human serum samples. A total of 300 subjects were enrolled in this study. Serum levels of ENA78/CXCL5 and SDF-1/CXCL12, measured by chemiluminescent immunoassay, were compared among 4 disease groups; normal, high-risk (intestinal metaplasia and adenoma), early GC (EGC), and advanced GC (AGC) groups in both training (n=25 per group) and validation dataset (n=70, 30, 50, 50, respectively) by ANOVA test (post hoc Bonferroni). Correlations between serum ENA78/CXCL5 or SDF-1/CXCL12 levels and clinicopathological parameters of GC patients were evaluated (Spearman's correlation; γs). To validate the diagnostic accuracy, receiver operating characteristic (ROC) curve and logistic regression analysis was performed. Serum ENA78/CXCL5 and SDF-1/CXCL12 levels were significantly higher in AGC groups than EGC, high-risk and normal groups in both training and validation dataset (Bonferroni, from p<0.01 to p<0.001). Clinicopathologically, serum ENA78/CXCL5 was correlated with T-stage (γs=0.231, p=0.021) and distant metastasis (γs=0.357, p<0.001), while serum SDF-1/CXCL12 was correlated with lymph node (γs=0.220, p=0.029) and distant (γs=0.425, p<0.001) metastasis. ROC curve and logistic regression demonstrated that serum ENA78/CXCL5 and SDF-1/CXCL12 showed higher diagnostic accuracy compared with carcinoembryonic antigen (CEA) in predicting GC. Serum ENA78/CXCL5 could predict both the presence of GC and distant metastasis, while serum SDF-1/CXCL12 could mainly predict its distant metastasis. All combination of serum ENA78/CXCL5, SDF-1/CXCL12, and CEA

  13. Alzheimer's disease shares gene expression aberrations with purinergic dysregulation of HPRT deficiency (Lesch-Nyhan disease).

    PubMed

    Kang, Tae Hyuk; Friedmann, Theodore

    2015-03-17

    Transcriptomic studies of murine D3 embryonic stem (ES) cells deficient in the purinergic biosynthetic function hypoxanthine guanine phosphoribosyltransferase (HPRT) and undergoing dopaminergic neuronal differentiation has demonstrated a marked shift from neuronal to glial gene expression and aberrant expression of multiple genes also known to be aberrantly expressed in Alzheimer's and other CNS disorders. Such genetic dysregulations may indicate some shared pathogenic metabolic mechanisms in diverse CNS diseases. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  14. Interaction of the chemokines I-TAC (CXCL11) and SDF-1 (CXCL12) in the regulation of tumor angiogenesis of colorectal cancer.

    PubMed

    Rupertus, Kathrin; Sinistra, Janine; Scheuer, Claudia; Nickels, Ruth M; Schilling, Martin K; Menger, Michael D; Kollmar, Otto

    2014-04-01

    The chemokine CXCL12 has a decisive role in tumor progression by mediating pro-angiogenic and pro-metastatic effects through its receptor CXCR4. The CXCL12 pathway is connected with another chemokine, CXCL11, through its second receptor CXCR7. CXCL11 also binds to the CXCR3 receptor. CXCL11 function in tumor angiogenesis is likely receptor dependent because CXCR3 predominantly mediates angiostatic signals whereas CXCR7 mediated signaling is rather angiogenic. We therefore studied the interaction of CXCL12 and CXCL11 in an in vivo model of colorectal cancer metastasis. GFP-transfected CT26.WT colorectal cancer cells were implanted into the dorsal skinfold chamber of syngeneic BALB/c mice. The animals received either peritumoral application of CXCL11 or intraperitoneal injections with neutralizing antibodies against CXCL11, CXCL12 or both. Tumor growth characteristics, angiogenesis, cell migration, invasive tumor growth, tumor cell proliferation and apoptosis were studied by intravital fluorescence microscopy and immunohistochemistry during an observation period of 14 days. Local exposure to CXCL11 significantly stimulated tumor growth compared to controls and enhanced invasive growth characteristics without affecting tumor angiogenesis and tumor cell migration. Neither CXCL11 nor CXCL12-blockade had a significant impact on tumor growth and angiogenesis, whereas the combined neutralization of CXCL11 and CXCL12 almost completely abrogated tumor vessel formation. As a consequence, tumor growth and invasive growth characteristics were reduced compared to the other groups. The results of the present study underline the interaction of CXCL12 and CXCL11 during tumor angiogenesis. The combined blockade of both signaling pathways may provide an interesting anti-angiogenic approach for anti-tumor therapy.

  15. Delayed Healing of Sickle Cell Ulcers Is due to Impaired Angiogenesis and CXCL12 Secretion in Skin Wounds.

    PubMed

    Nguyen, Van Tuan; Nassar, Dany; Batteux, Fréderic; Raymond, Karine; Tharaux, Pierre-Louis; Aractingi, Sélim

    2016-02-01

    Leg ulcers are a major complication of sickle cell disease that occur in 2.5-40% of patients. Leg ulcers are responsible for frequent complications because they are often long-lasting and are highly resistant to therapy. Although their occurrence is associated with hyperhemolysis, the mechanisms underlying sickle cell ulcers remain poorly understood. In this study, we show that skin wound healing is severely altered in old SAD sickle cell mice but is normal in young animals, consistent with reports in humans. Alterations of wound healing were associated with impaired blood and lymphatic angiogenesis in the wound beds and poor endothelial progenitor cell mobilization from the bone marrow. CXCL12 secretion by keratinocytes and inflammatory cells was low in the wounds of SAD mice. Local therapy with endothelial progenitor cells or recombinant CXCL12 injections restored wound angiogenesis and rescued the healing defect together with mobilization of circulating endothelial progenitor cells. To our knowledge, this is a previously unreported study of the cellular and molecular mechanisms of sickle cell ulcers in a murine model that provides promising therapeutic perspectives for clinical trials. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  16. Higher plasma CXCL12 levels predict incident myocardial infarction and death in chronic kidney disease: findings from the Chronic Renal Insufficiency Cohort study

    PubMed Central

    Mehta, Nehal N.; Matthews, Gregory J.; Krishnamoorthy, Parasuram; Shah, Rhia; McLaughlin, Catherine; Patel, Parth; Budoff, Matthew; Chen, Jing; Wolman, Melanie; Go, Alan; He, Jiang; Kanetsky, Peter A.; Master, Stephen R.; Rader, Daniel J.; Raj, Dominic; Gadegbeku, Crystal A.; Shah, Rachana; Schreiber, Marty; Fischer, Michael J.; Townsend, Raymond R.; Kusek, John; Feldman, Harold I.; Foulkes, Andrea S.; Reilly, Muredach P.; Appel, Lawrence J.; Feldman, Harold I.; Go, Alan S.; He, Jiang; Kusek, John W.; Lash, James P.; Ojo, Akinlolu; Rahman, Mahboob; Townsend, Raymond R.

    2014-01-01

    Aims Genome-wide association studies revealed an association between a locus at 10q11, downstream from CXCL12, and myocardial infarction (MI). However, the relationship among plasma CXCL12, cardiovascular disease (CVD) risk factors, incident MI, and death is unknown. Methods and results We analysed study-entry plasma CXCL12 levels in 3687 participants of the Chronic Renal Insufficiency Cohort (CRIC) Study, a prospective study of cardiovascular and kidney outcomes in chronic kidney disease (CKD) patients. Mean follow-up was 6 years for incident MI or death. Plasma CXCL12 levels were positively associated with several cardiovascular risk factors (age, hypertension, diabetes, hypercholesterolaemia), lower estimated glomerular filtration rate (eGFR), and higher inflammatory cytokine levels (P < 0.05). In fully adjusted models, higher study-entry CXCL12 was associated with increased odds of prevalent CVD (OR 1.23; 95% confidence interval 1.14, 1.33, P < 0.001) for one standard deviation (SD) increase in CXCL12. Similarly, one SD higher CXCL12 increased the hazard of incident MI (1.26; 1.09,1.45, P < 0.001), death (1.20; 1.09,1.33, P < 0.001), and combined MI/death (1.23; 1.13–1.34, P < 0.001) adjusting for demographic factors, known CVD risk factors, and inflammatory markers and remained significant for MI (1.19; 1.03,1.39, P = 0.01) and the combined MI/death (1.13; 1.03,1.24, P = 0.01) after further controlling for eGFR and urinary albumin:creatinine ratio. Conclusions In CKD, higher plasma CXCL12 was associated with CVD risk factors and prevalent CVD as well as the hazard of incident MI and death. Further studies are required to establish if plasma CXCL12 reflect causal actions at the vessel wall and is a tool for genomic and therapeutic trials. PMID:24306482

  17. White matter tracts for the trafficking of neural progenitor cells characterized by cellular MRI and immunohistology: the role of CXCL12/CXCR4 signaling.

    PubMed

    Chen, Chiao-Chi V; Hsu, Yi-Hua; Jayaseema, D M; Chen, Jeou-Yuan Joanne; Hueng, Dueng-Yuan; Chang, Chen

    2015-07-01

    White matter tracts are important for the trafficking of neural progenitor cells (NPCs) in both normal and pathological conditions, but the underlying mechanism is not clear. The directionality of white matter is advantageous for molecules or cells to distribute over a long distance, but this feature is unlikely solely responsible for efficient migration. The present study hypothesizes that the efficient migration of NPCs into white matter is under the influences of neurochemical attraction—CXCL12/CXCR4 signaling, a major mechanism underlying the targeted migration of NPCs. To test this view, the present study investigated the effects of CXCL12 administration into the corpus callosum (CC) on the migratory behavior of transplanted NPCs. A living animal tracking platform based on MRI and a magnetic cell labeling technique was employed. The NPCs were magnetically labeled and then transplanted at the right end of the CC. CXCL12 was infused continuously at the left end. Migration of NPCs was monitored repeatedly over a 7-day course using 3D gradient echo T2*-weighted imaging. It was found that, CXCL12 induced NPCs to migrate up to 1,881 μm from the graft whereas the spontaneous migration was mere 200 μm. CXCL12 induced migration that was nine times as efficient in the speed. The results indicate that the CXCL12/CXCR4 signaling may be a mechanism via which NPCs efficiently migrate along the white matter tracts. The study also presents a potential strategy for facilitating the targeted migration in NPC therapy for brain disorders.

  18. Engineering NK Cells Modified With an EGFRvIII-specific Chimeric Antigen Receptor to Overexpress CXCR4 Improves Immunotherapy of CXCL12/SDF-1α-secreting Glioblastoma.

    PubMed

    Müller, Nadja; Michen, Susanne; Tietze, Stefanie; Töpfer, Katrin; Schulte, Alexander; Lamszus, Katrin; Schmitz, Marc; Schackert, Gabriele; Pastan, Ira; Temme, Achim

    2015-06-01

    Natural killer (NK) cells are promising effector cells for adjuvant immunotherapy of cancer. So far, several preclinical studies have shown the feasibility of gene-engineered NK cells, which upon expression of chimeric antigen receptors (CARs) are redirected to otherwise NK cell-resistant tumors. Yet, we reasoned that the efficiency of an immunotherapy using CAR-modified NK cells critically relies on efficient migration to the tumor site and might be improved by the engraftment of a receptor specific for a chemokine released by the tumor. On the basis of the DNAX-activation protein 12 (DAP12), a signaling adapter molecule involved in signal transduction of activating NK cell receptors, we constructed an epidermal growth factor variant III (EGFRvIII)-CAR, designated MR1.1-DAP12 which confers specific cytotoxicity of NK cell towards EGFRvIII glioblastoma cells in vitro and to established subcutaneous U87-MG tumor xenografts. So far, infusion of NK cells with expression of MR1.1-DAP12 caused a moderate but significantly delayed tumor growth and increased median survival time when compared with NK cells transduced with an ITAM-defective CAR. Notably, the further genetic engineering of these EGFRvIII-specific NK cells with the chemokine receptor CXCR4 conferred a specific chemotaxis to CXCL12/SDF-1α secreting U87-MG glioblastoma cells. Moreover, the administration of such NK cells resulted in complete tumor remission in a number of mice and a significantly increased survival when compared with the treatment of xenografts with NK cells expressing only the EGFRvIII-specific CAR or mock control. We conclude that chemokine receptor-engineered NK cells with concomitant expression of a tumor-specific CAR are a promising tool to improve adoptive tumor immunotherapy.

  19. CCL3 and CXCL12 production in vitro by dental pulp fibroblasts from permanent and deciduous teeth stimulated by Porphyromonas gingivalis LPS

    PubMed Central

    SIPERT, Carla Renata; MORANDINI, Ana Carolina de Faria; MODENA, Karin Cristina da Silva; DIONÍSIO, Thiago José; MACHADO, Maria Aparecida Andrade Moreira; de OLIVEIRA, Sandra Helena Penha; CAMPANELLI, Ana Paula; SANTOS, Carlos Ferreira

    2013-01-01

    Objective: The aim of this study was to compare the production of the chemokines CCL3 and CXCL12 by cultured dental pulp fibroblasts from permanent (PDPF) and deciduous (DDPF) teeth under stimulation by Porphyromonas gingivalis LPS (PgLPS). Material and Methods: Primary culture of fibroblasts from permanent (n=3) and deciduous (n=2) teeth were established using an explant technique. After the fourth passage, fibroblasts were stimulated by increasing concentrations of PgLPS (0 - 10 µg/mL) at 1, 6 and 24 h. The cells were tested for viability through MTT assay, and production of the chemokines CCL3 and CXCL12 was determined through ELISA. Comparisons among samples were performed using One-way ANOVA for MTT assay and Two-way ANOVA for ELISA results. Results: Cell viability was not affected by the antigen after 24 h of stimulation. PgLPS induced the production of CCL3 by dental pulp fibroblasts at similar levels for both permanent and deciduous pulp fibroblasts. Production of CXCL12, however, was significantly higher for PDPF than DDPF at 1 and 6 h. PgLPS, in turn, downregulated the production of CXCL12 by PDPF but not by DDPF. Conclusion: These data suggest that dental pulp fibroblasts from permanent and deciduous teeth may present a differential behavior under PgLPS stimulation. PMID:23739851

  20. PERIPHERAL NERVE-DERIVED CXCL12 AND VEGF-A REGULATE THE PATTERNING OF ARTERIAL VESSEL BRANCHING IN DEVELOPING LIMB SKIN

    PubMed Central

    Li, Wenling; Kohara, Hiroshi; Uchida, Yutaka; James, Jennifer M.; Soneji, Kosha; Cronshaw, Darran G.; Zou, Yong-Rui; Nagasawa, Takashi; Mukouyama, Yoh-suke

    2013-01-01

    SUMMARY In developing limb skin, peripheral nerves provide a spatial template that controls the branching pattern and differentiation of arteries. Our previous studies indicate that nerve-derived VEGF-A is required for arterial differentiation but not for nerve-vessel alignment. In this study, we demonstrate that nerve-vessel alignment depends on the activity of Cxcl12-Cxcr4 chemokine signaling. Genetic inactivation of Cxcl12-Cxcr4 signaling perturbs nerve-vessel alignment, and abolishes arteriogenesis. Further in vitro assays allow us to uncouple nerve-vessel alignment and arteriogenesis, revealing that nerve-derived Cxcl12 stimulates endothelial cell migration, while nerve-derived VEGF-A is responsible for arterial differentiation. These findings suggest a coordinated sequential action in which nerve-Cxcl12 functions over a distance to recruit vessels to align with nerves and subsequent arterial differentiation presumably requires a local-action of nerve-VEGF-A in the nerve-associated vessels. PMID:23395391

  1. Interaction of chemokine receptor CXCR4 in monomeric and dimeric state with its endogenous ligand CXCL12: coarse-grained simulations identify differences.

    PubMed

    Cutolo, Pasquale; Basdevant, Nathalie; Bernadat, Guillaume; Bachelerie, Françoise; Ha-Duong, Tâp

    2017-02-01

    Despite the recent resolutions of the crystal structure of the chemokine receptor CXCR4 in complex with small antagonists or viral chemokine, a description at the molecular level of the interactions between the full-length CXCR4 and its endogenous ligand, the chemokine CXCL12, in relationship with the receptor recognition and activation, is not yet completely elucidated. Moreover, since CXCR4 is able to form dimers, the question of whether the CXCR4-CXCL12 complex has a 1:1 or 2:1 preferential stoichiometry is still an open question. We present here results of coarse-grained protein-protein docking and molecular dynamics simulations of CXCL12 in association with CXCR4 in monomeric and dimeric states. Our proposed models for the 1:1 and 2:1 CXCR4-CXCL12 quaternary structures are consistent with recognition and activation motifs of both partners provided by the available site-directed mutagenesis data. Notably, we observed that in the 2:1 complex, the chemokine N-terminus makes more steady contacts with the receptor residues critical for binding and activation than in the 1:1 structure, suggesting that the 2:1 stoichiometry would favor the receptor signaling activity with respect to the 1:1 association.

  2. HMGB1 promotes recruitment of inflammatory cells to damaged tissues by forming a complex with CXCL12 and signaling via CXCR4

    PubMed Central

    Schiraldi, Milena; Raucci, Angela; Muñoz, Laura Martínez; Livoti, Elsa; Celona, Barbara; Venereau, Emilie; Apuzzo, Tiziana; De Marchis, Francesco; Pedotti, Mattia; Bachi, Angela; Thelen, Marcus; Varani, Luca; Mellado, Mario; Proudfoot, Amanda; Bianchi, Marco Emilio

    2012-01-01

    After tissue damage, inflammatory cells infiltrate the tissue and release proinflammatory cytokines. HMGB1 (high mobility group box 1), a nuclear protein released by necrotic and severely stressed cells, promotes cytokine release via its interaction with the TLR4 (Toll-like receptor 4) receptor and cell migration via an unknown mechanism. We show that HMGB1-induced recruitment of inflammatory cells depends on CXCL12. HMGB1 and CXCL12 form a heterocomplex, which we characterized by nuclear magnetic resonance and surface plasmon resonance, that acts exclusively through CXCR4 and not through other HMGB1 receptors. Fluorescence resonance energy transfer data show that the HMGB1–CXCL12 heterocomplex promotes different conformational rearrangements of CXCR4 from that of CXCL12 alone. Mononuclear cell recruitment in vivo into air pouches and injured muscles depends on the heterocomplex and is inhibited by AMD3100 and glycyrrhizin. Thus, inflammatory cell recruitment and activation both depend on HMGB1 via different mechanisms. PMID:22370717

  3. Regulation of Cell Migration by Sphingomyelin Synthases: Sphingomyelin in Lipid Rafts Decreases Responsiveness to Signaling by the CXCL12/CXCR4 Pathway

    PubMed Central

    Asano, Satoshi; Kitatani, Kazuyuki; Taniguchi, Makoto; Hashimoto, Mayumi; Zama, Kota; Mitsutake, Susumu; Igarashi, Yasuyuki; Takeya, Hiroyuki; Kigawa, Junzo; Hayashi, Akira; Umehara, Hisanori

    2012-01-01

    Sphingomyelin synthase (SMS) catalyzes the formation of sphingomyelin, a major component of the plasma membrane and lipid rafts. To investigate the role of SMS in cell signaling and migration induced by binding of the chemokine CXCL12 to CXCR4, we used mouse embryonic fibroblasts deficient in SMS1 and/or SMS2 and examined the effects of SMS deficiency on cell migration. SMS deficiency promoted cell migration through a CXCL12/CXCR4-dependent signaling pathway involving extracellular signal-regulated kinase (ERK) activation. In addition, SMS1/SMS2 double-knockout cells had heightened sensitivity to CXCL12, which was significantly suppressed upon transfection with the SMS1 or SMS2 gene or when they were treated with exogenous sphingomyelin but not when they were treated with the SMS substrate ceramide. Notably, SMS deficiency facilitated relocalization of CXCR4 to lipid rafts, which form platforms for the regulation and transduction of receptor-mediated signaling. Furthermore, we found that SMS deficiency potentiated CXCR4 dimerization, which is required for signal transduction. This dimerization was significantly repressed by sphingomyelin treatment. Collectively, our data indicate that SMS-derived sphingomyelin lowers responsiveness to CXCL12, thereby reducing migration induced by this chemokine. Our findings provide the first direct evidence for an involvement of SMS-generated sphingomyelin in the regulation of cell migration. PMID:22688512

  4. Progesterone reduces the migration of mast cells toward the chemokine stromal cell-derived factor-1/CXCL12 with an accompanying decrease in CXCR4 receptors.

    PubMed

    Belot, Marie-Pierre; Abdennebi-Najar, Latifa; Gaudin, Françoise; Lieberherr, Michèle; Godot, Véronique; Taïeb, Joelle; Emilie, Dominique; Machelon, Véronique

    2007-05-01

    Mast cell recruitment is implicated in many physiological functions and several diseases. It depends on microenvironmental factors, including hormones. We have investigated the effect of progesterone on the migration of HMC-1(560) mast cells toward CXCL12, a chemokine that controls the migration of mast cells into tissues. HMC-1(560) mast cells were incubated with 1 nM to 1 microM progesterone for 24 h. Controls were run without progesterone. Cell migration toward CXCL12 was monitored with an in vitro assay, and statistical analysis of repeated experiments revealed that progesterone significantly reduced cell migration without increasing the number of apoptotic cells (P = 0.0084, n = 7). Differences between progesterone-treated and untreated cells were significant at 1 microM (P < 0.01, n = 7). Cells incubated with 1 microM progesterone showed no rearrangment of actin filaments in response to CXCL12. Progesterone also reduced the calcium response to CXCL12 and Akt phosphorylation. Cells incubated with progesterone had one-half the control concentrations of CXCR4 (mRNA, total protein, and membrane-bound protein). Progesterone also inhibited the migration of HMC-1(560) cells transfected with hPR-B-pSG5 plasmid, which contained 2.5 times as much PR-B as the control. These transfected cells responded differently (P < 0.05, n = 5) from untreated cells to 1 nM progesterone. We conclude that progesterone reduces mast cell migration toward CXCL12 and that CXCR4 may be a progesterone target in mast cells.

  5. The chemokine CXCL12 and the HIV-1 envelope protein gp120 regulate spontaneous activity of Cajal-Retzius cells in opposite directions.

    PubMed

    Marchionni, Ivan; Beaumont, Michael; Maccaferri, Gianmaria

    2012-07-01

    Activation of the CXC chemokine receptor 4 (CXCR4) in Cajal–Retzius cells by CXC chemokine ligand 12 (CXCL12) is important for controlling their excitability. CXCR4 is also a co-receptor for the glycoprotein 120 (gp120) of the envelope of the human immunodeficiency virus type 1 (HIV-1), and binding of gp120 to CXCR4 may produce pathological effects. In order to study CXCR4-dependent modulation of membrane excitability, we recorded in cell-attached configuration spontaneous action currents from hippocampal stratum lacunosum-moleculare Cajal–Retzius cells of the CXCR4-EGFP mouse. CXCL12 (50 nM) powerfully inhibited firing independently of synaptic transmission, suggesting that CXCR4 regulates an intrinsic conductance. This effect was prevented by conditioning slices with BAPTA-AM (200 μM), and by blockers of the BK calcium-dependent potassium channels (TEA (1 mM), paxilline (10 μM) and iberiotoxin (100 nM)). In contrast, exposure to gp120 (pico- to nanomolar range, alone or in combination with soluble cluster of differentiation 4 (CD4)), enhanced spontaneous firing frequency. This effect was prevented by the CXCR4 antagonist AMD3100 (1 μM) and was absent in EGFP-negative stratum lacunosum-moleculare interneurons. Increased excitability was prevented by treating slices with BAPTA-AM or bumetanide, suggesting that gp120 activates a mechanism that is both calcium- and chloride-dependent. In conclusion, our results demonstrate that CXCL12 and gp120 modulate the excitability of Cajal–Retzius cells in opposite directions. We propose that CXCL12 and gp120 either generate calcium responses of different strength or activate distinct pools of intracellular calcium, leading to agonist-specific responses, mediated by BK channels in the case of CXCL12, and by a chloride-dependent mechanism in the case of gp120.

  6. The chemokine CXCL12 and the HIV-1 envelope protein gp120 regulate spontaneous activity of Cajal–Retzius cells in opposite directions

    PubMed Central

    Marchionni, Ivan; Beaumont, Michael; Maccaferri, Gianmaria

    2012-01-01

    Activation of the CXC chemokine receptor 4 (CXCR4) in Cajal–Retzius cells by CXC chemokine ligand 12 (CXCL12) is important for controlling their excitability. CXCR4 is also a co-receptor for the glycoprotein 120 (gp120) of the envelope of the human immunodeficiency virus type 1 (HIV-1), and binding of gp120 to CXCR4 may produce pathological effects. In order to study CXCR4-dependent modulation of membrane excitability, we recorded in cell-attached configuration spontaneous action currents from hippocampal stratum lacunosum-moleculare Cajal–Retzius cells of the CXCR4-EGFP mouse. CXCL12 (50 nm) powerfully inhibited firing independently of synaptic transmission, suggesting that CXCR4 regulates an intrinsic conductance. This effect was prevented by conditioning slices with BAPTA-AM (200 μm), and by blockers of the BK calcium-dependent potassium channels (TEA (1 mm), paxilline (10 μm) and iberiotoxin (100 nm)). In contrast, exposure to gp120 (pico- to nanomolar range, alone or in combination with soluble cluster of differentiation 4 (CD4)), enhanced spontaneous firing frequency. This effect was prevented by the CXCR4 antagonist AMD3100 (1 μm) and was absent in EGFP-negative stratum lacunosum-moleculare interneurons. Increased excitability was prevented by treating slices with BAPTA-AM or bumetanide, suggesting that gp120 activates a mechanism that is both calcium- and chloride-dependent. In conclusion, our results demonstrate that CXCL12 and gp120 modulate the excitability of Cajal–Retzius cells in opposite directions. We propose that CXCL12 and gp120 either generate calcium responses of different strength or activate distinct pools of intracellular calcium, leading to agonist-specific responses, mediated by BK channels in the case of CXCL12, and by a chloride-dependent mechanism in the case of gp120. PMID:22473778

  7. Aberrant gene expression in organs of bovine clones that die within two days after birth.

    PubMed

    Li, Shijie; Li, Yanxin; Du, Weihua; Zhang, Lei; Yu, Shuyang; Dai, Yunping; Zhao, Chunjiang; Li, Ning

    2005-02-01

    Cloning by somatic nuclear transfer is an inefficient process in which some of the cloned animals die shortly after birth and display organ abnormalities. In an effort to determine the possible genetic causes of neonatal death and organ abnormalities, we used real-time quantitative reverse transcription-polymerase chain reaction to examine expression patterns of eight developmentally important genes (PCAF, Xist, FGFR2, PDGFRa, FGF10, BMP4, Hsp70.1, and VEGF) in six organs (heart, liver, spleen, lung, kidney, and brain) of both cloned bovines that died soon after birth (n=9) and normal control calves produced by artificial insemination. In somatic cloning of cattle, fibroblasts have often been used for doner nuclei, and the effect of the age of the fibroblast donor cells on gene expression profiles was investigated. Aberrant expressions of seven genes were found in these clones. The majority of aberrantly expressed genes were common in clones derived from adult fibroblast (AF) and in clones derived from fetal fibroblast (FF) compared to controls, whereas some genes were dysregulated either in AF cell-derived or in FF cell-derived clones. For the studied genes, kidney was the organ least affected by gene dysregulation, and heart was the organ most affected, in which five genes were aberrant. Most dysregulations (12 of 19) were up-regulation, but PDGFRa only showed down-regulation. VEGF, BMP-4, PCAF, and Hsp70.1 were extremely dysregulated, whereas the other four genes had a low level of gene dysregulation. Our results suggest that the aberrant gene expression occurred in most tissues of cloned bovines that died soon after birth. For each specific gene, aberrant expression resulting from nuclear transfer was tissue-specific. Because these genes play important roles in embryo development and organogenesis, the aberrant transcription patterns detected in these clones may contribute to the defects of organs reported in neonatal death of clones.

  8. Deciphering causal and statistical relations of molecular aberrations and gene expressions in NCI-60 cell lines

    PubMed Central

    2011-01-01

    Background Cancer cells harbor a large number of molecular alterations such as mutations, amplifications and deletions on DNA sequences and epigenetic changes on DNA methylations. These aberrations may dysregulate gene expressions, which in turn drive the malignancy of tumors. Deciphering the causal and statistical relations of molecular aberrations and gene expressions is critical for understanding the molecular mechanisms of clinical phenotypes. Results In this work, we proposed a computational method to reconstruct association modules containing driver aberrations, passenger mRNA or microRNA expressions, and putative regulators that mediate the effects from drivers to passengers. By applying the module-finding algorithm to the integrated datasets of NCI-60 cancer cell lines, we found that gene expressions were driven by diverse molecular aberrations including chromosomal segments' copy number variations, gene mutations and DNA methylations, microRNA expressions, and the expressions of transcription factors. In-silico validation indicated that passenger genes were enriched with the regulator binding motifs, functional categories or pathways where the drivers were involved, and co-citations with the driver/regulator genes. Moreover, 6 of 11 predicted MYB targets were down-regulated in an MYB-siRNA treated leukemia cell line. In addition, microRNA expressions were driven by distinct mechanisms from mRNA expressions. Conclusions The results provide rich mechanistic information regarding molecular aberrations and gene expressions in cancer genomes. This kind of integrative analysis will become an important tool for the diagnosis and treatment of cancer in the era of personalized medicine. PMID:22051105

  9. Aberrant expression of splicing factors in newly diagnosed acute myeloid leukemia.

    PubMed

    Liu, Jing; Huang, Bo; Xiao, Yun; Xiong, Huo-Mei; Li, Jing; Feng, Dan-Qin; Chen, Xi-Min; Zhang, Hai-Bin; Wang, Xiao-Zhong

    2012-01-01

    Acute myeloid leukemia (AML) is the most common type of blood cancer in adults. Emerging evidence is establishing a connection between AML and aberrant alternative splicing of pre-mRNA, which may result from aberrant expression of splicing factors, the mediators of splicing reactions. Using quantitative real-time polymerase chain reaction, we measured mRNA expression of 7 splicing factors belonging to the serine/arginine-rich (SR) protein family, SRSF1 (SF2/ASF), SRSF2 (SC35), SRSF3 (SRp20), SRSF4 (SRp75), SRSF5 (SRp40), SRSF6 (SRp55), and SRSF7 (9G8), and 1 non-SR factor, heterogeneous nuclear ribonucleoprotein A1 (HNRNPA1), in peripheral blood mononuclear cells of 26 patients with newly diagnosed AML and 26 healthy controls. In addition, the relationship between splicing factors and the mRNA splicing patterns of the caspase-8 gene (CASP8) was investigated. Compared to healthy controls, the expression of splicing factors was obviously aberrant in newly diagnosed AML patients. The expression of SRSF1, SRSF3 and SRSF4 mRNAs was significantly decreased. Moreover, a significant correlation was observed between several splicing factors and caspase-8 pre-mRNA splicing in AML patients, but not in control subjects. These data suggest that aberrant expression of splicing factors in AML may potentially connect with abnormal expression of oncogenes and be useful for early diagnosis, prognosis, and therapy of AML. Copyright © 2012 S. Karger AG, Basel.

  10. From DNA Copy Number to Gene Expression: Local aberrations, Trisomies and Monosomies

    NASA Astrophysics Data System (ADS)

    Shay, Tal

    The goal of my PhD research was to study the effect of DNA copy number changes on gene expression. DNA copy number aberrations may be local, encompassing several genes, or on the level of an entire chromosome, such as trisomy and monosomy. The main dataset I studied was of Glioblastoma, obtained in the framework of a collaboration, but I worked also with public datasets of cancer and Down's Syndrome. The molecular basis of expression changes in Glioblastoma. Glioblastoma is the most common and aggressive type of primary brain tumors in adults. In collaboration with Prof. Hegi (CHUV, Switzerland), we analyzed a rich Glioblastoma dataset including clinical information, DNA copy number (array CGH) and expression profiles. We explored the correlation between DNA copy number and gene expression at the level of chromosomal arms and local genomic aberrations. We detected known amplification and over expression of oncogenes, as well as deletion and down-regulation of tumor suppressor genes. We exploited that information to map alterations of pathways that are known to be disrupted in Glioblastoma, and tried to characterize samples that have no known alteration in any of the studied pathways. Identifying local DNA aberrations of biological significance. Many types of tumors exhibit chromosomal losses or gains and local amplifications and deletions. A region that is aberrant in many tumors, or whose copy number change is stronger, is more likely to be clinically relevant, and not just a by-product of genetic instability. We developed a novel method that defines and prioritizes aberrations by formalizing these intuitions. The method scores each aberration by the fraction of patients harboring it, its length and its amplitude, and assesses the significance of the score by comparing it to a null distribution obtained by permutations. This approach detects genetic locations that are significantly aberrant, generating a 'genomic aberration profile' for each sample. The 'genomic

  11. Aberrant ADAM10 expression correlates with osteosarcoma progression

    PubMed Central

    2014-01-01

    Background Osteosarcoma is the most common type of bone cancer and is notorious for its rapid progression. The Notch signaling pathway has recently been shown to be involved in osteosarcoma. As a major sheddase of Notch receptors, ADAM10 has been implicated in many types of cancers, but its role in osteosarcoma has not been investigated. Previous studies have shown that the expression of CD31 was significantly elevated in metastatic osteosarcoma; however, its expression in nonmetastatic groups is not known. In addition, the mysterious multinucleated giant cell in giant cell-rich osteosarcoma was previously regarded as an osteoclast-like cell, but its exact identity is unclear. Method Tissue chip samples from 40 cases of nonmetastatic osteosarcoma were stained for cytoplasmic ADAM10, activated Notch1 and CD31. Osteoclasts in tumor sections were also stained for tartrate-resistant acid phosphatase (TRAP). Results Immunofluorescence staining revealed that ADAM10 expression significantly increased with the progression of osteosarcoma as well as in osteoblastic osteosarcoma, whereas the expression of the Notch intracellular domain (NICD) and CD31 was not significantly altered between different pathological stages. In addition, multinucleated giant cells in giant cell-rich osteosarcoma were also found to coexpress CD31, ADAM10 and NICD, but were negative for TRAP staining. Conclusions Our results highlight the importance of ADAM10 in the progression of osteosarcoma and suggest that the protein might be a potential therapeutic target in osteosarcoma treatment. This study also demonstrates that the multinucleated giant cell is an angiogenic tumor cell, rather than an osteoclast, and involves ADAM10/Notch1 signaling activation. PMID:24548763

  12. Aberrant maspin expression in gallbladder epithelium is associated with intestinal metaplasia in patients with cholelithiasis

    PubMed Central

    Maesawa, C; Ogasawara, S; Yashima‐Abo, A; Kimura, T; Kotani, K; Masuda, S; Nagata, Y; Iwaya, T; Suzuki, K; Oyake, T; Akiyama, Y; Kawamura, H; Masuda, T

    2006-01-01

    Objective Aberrant expression of maspin protein related to DNA hypomethylation in the promoter region is frequently observed in gallbladder carcinomas, whereas the non‐tumorous gallbladder epithelium is maspin negative. We investigated maspin expression in non‐tumorous gallbladder epithelium in patients with cholelithiasis. Methods An immunohistochemical study of maspin expression was performed in 69 patients with cholelithiasis and 30 patients with gastric cancer without cholelithiasis. Results Immunoreactivity for maspin was observed in focal and patchy regions of the gallbladder epithelium. Positive immunoreactivity for maspin was significantly associated with the presence of intestinal metaplasia in patients with cholelithiasis (p<0.05). Conclusion The high incidence of aberrant maspin expression in both intestinal metaplasia and carcinoma of the gallbladder supports the assumption that intestinal metaplasia of the gallbladder may predispose to gallbladder carcinoma. PMID:16505288

  13. Aberrant microRNA expression and its implications in the pathogenesis of leukemias.

    PubMed

    Babashah, Sadegh; Sadeghizadeh, Majid; Tavirani, Mostafa Rezaei; Farivar, Shirin; Soleimani, Masoud

    2012-10-01

    MicroRNAs (miRNAs) are a class of non-coding, endogenous, small RNAs that negatively regulate gene expression by inducing degradation or translational inhibition of target mRNAs. Aberrant expression of miRNAs appears to be a common characteristic of hematological malignancies including leukemias. Here we review the available data supporting a role of aberrant expression of miRNAs in the pathogenesis of leukemias including acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), chronic myeloid leukemia (CML), and chronic lymphocytic leukemia (CLL). The expression signatures of miRNAs provide exciting opportunities in the diagnosis, prognosis, and therapy of leukemia. Since miRNAs can function as either oncogenes or tumor suppressor genes in leukemogenesis, the potential of using these small RNAs as therapeutic targets opens up new opportunities for leukemia therapy by either inhibiting or augmenting their activity.

  14. Fibronectin: How Its Aberrant Expression in Tumors May Improve Therapeutic Targeting

    PubMed Central

    Wang, Jennifer Peyling; Hielscher, Abigail

    2017-01-01

    Fibronectin is a matrix glycoprotein which has not only been found to be over-expressed in several cancers, but has been shown to participate in several steps of tumorigenesis. The purpose of this review is to illustrate how aberrant fibronectin expression influences tumor growth, invasion, metastasis and therapy resistance. In particular, this review will focus on the interactions between cell receptor ligands and fibronectin and how this interaction influences downstream signaling events that aid tumor progression. This review will further discuss the possible implications of therapeutic drugs directed against fibronectin and/or cellular interactions with fibronectin and will additionally discuss novel approaches by which to limit intra- and extra-tumoral fibronectin expression and the cellular events which lead to aberrant fibronectin expression. It is anticipated that these studies will set a basis for future research that will not only aid understanding of fibronectin and its prognostic significance, but will further elucidate novel targets for therapeutics. PMID:28367247

  15. Aberrant expression of cyclin D1 in cancer

    PubMed Central

    Inoue, Kazushi; Fry, Elizabeth A.

    2016-01-01

    Cyclin D1 binds and activates cyclin-dependent kinases 4/6 (Cdk4/6) to phosphorylate the retinoblastoma (RB) family proteins, relieving E2F/DPs from the negative restraint of RB proteins and histone deacetylases. The cyclin D-Cdk4/6 complexes activate cyclin E/Cdk2 through titration of the Cdk inhibitors p21Cip1/p27Kip1. Cyclin E/Cdk2 further phosphorylates RBs, thereby activating E2F/DPs, and cells enter the S phase of the cell cycle. Cyclin D-Cdk4/6 also phosphorylates MEP50 subunit of the protein arginine methyltransferase 5 (PRMT5), which cooperates with cyclin D1 to drive lymphomagenesis in vivo. Activated PRMPT5 causes arginine methylation of p53 to suppress expression of pro-apoptotic and anti-proliferative target genes, explaining the molecular mechanism for tumorigenesis. Cyclin D1 physically interacts with transcription factors such as estrogen receptor, androgen receptor, and Myb family proteins to regulate gene expression in Cdk-independent fashion. Dmp1 is a Myb-like protein that quenches the oncogenic signals from activated Ras or HER2 by inducing Arf/p53-dependent cell cycle arrest. Cyclin D1 binds to Dmp1α to activate both Arf and Ink4a promoters to induce cell cycle arrest or apoptosis in non-transformed cells to prevent them from neoplastic transformation. Dmp1-deficiency significantly accelerates mouse mammary tumorigenesis with reduced apoptosis and increased metastasis. Cyclin D1 interferes with ligand activation of PPARγ involved in cellular differentiation; it also physically interacts with histone deacetylases (HDACs) and p300 to repress gene expression. It has also been shown that cyclin D1 accelerates tumorigenesis through transcriptional activation of miR-17/20 and Dicer1 which, in turn, represses cyclin D1 expression. Identification of cyclin D1-binding proteins/promoters will be essential for further clarification of its biological activities. PMID:28090171

  16. The poor recovery of neuromyelitis optica spectrum disorder is associated with a lower level of CXCL12 in the human brain.

    PubMed

    Kang, Hao; Cao, Shanshan; Chen, Tingjun; Jiang, Zhaocai; Liu, Zihao; Li, Zhaohui; Wei, Yangang; Ai, Nanping; Xu, Quangang; Lin, Qing; Wei, Shihui

    2015-12-15

    Neuromyelitis optica spectrum disorders (NMOSDs) are blindness-causing neuritis. In NMOSD patients, NMO-IgG evokes astrocytopathy that in turn causes demyelination. While measurement of NMO-IgG titer will help neurologists make the diagnosis of NMOSDs, it is not sufficient to evaluate the severity of astrocytopathy. In this study, we compared the different levels of an astrocyte biomarker in cerebrospinal fluid of NMOSD patients with good or poor recovery, and then linked their differences to the changes in remyelinating promoter (CXCL12) levels. Our results indicate that NMO-IgG down-regulated CXCL12 and impaired the remyelinating process, this may be a mechanism contributing to the poor recovery of NMOSDs. Copyright © 2015. Published by Elsevier B.V.

  17. The HMGB1–CXCL12 Complex Promotes Inflammatory Cell Infiltration in Uveitogenic T Cell-Induced Chronic Experimental Autoimmune Uveitis

    PubMed Central

    Yun, Juan; Jiang, Guomin; Wang, Yunsong; Xiao, Tong; Zhao, Yuan; Sun, Deming; Kaplan, Henry J.; Shao, Hui

    2017-01-01

    It is largely unknown how invading autoreactive T cells initiate the pathogenic process inside the diseased organ in organ-specific autoimmune diseases. In experimental autoimmune uveitis (EAU) induced by uveitogenic, interphotoreceptor retinoid-binding protein (IRBP)-specific T cells (tEAU) in mice, we have previously reported that high mobility group box 1 (HMGB1) released as a consequence of the direct interaction between IRBP-specific T cells and retinal parenchymal cells is an early and critical mediator in induction of intraocular inflammation. Our present study explored the roles of HMGB1 in intraocular inflammation, focusing on its role in recruiting inflammatory cells into the eye. Our results showed that supernatants from retinal explants either stimulated with HMGB1 or cocultured with IRBP-specific T cells attracted leukocytes. Notably, HMGB1 antagonists blocked supernatant-induced chemoattraction when present from the start of coculture, but not when added to the culture supernatants after coculture, indicating that molecules released by HMGB1-treated retinal cells are chemoattractive. Moreover, CXCL12 levels in the coculture supernatants were dependent on HMGB1, since they were increased in the cocultures and reduced when HMGB1 antagonists were added at the beginning of the coculture. When either anti-CXCL12 Ab was added to the supernatants after coculture or the responding lymphocytes were pretreated with Ab against CXCL12 specific receptor, CXCR4, chemoattraction by the coculture supernatants was decreased. Finally, induction of tEAU was significantly inhibited by a CXCR4 antagonist, AMD3100, at the time of autoreactive T cell transfer. Our study demonstrates that, at a very early stage of intraocular inflammation initiated by uveitogenic autoreactive T cells, synergism between HMGB1 and CXCL12 is crucial for the infiltration of inflammatory cells. PMID:28261206

  18. Heparin-induced Leukocytosis Requires 6-O-Sulfation and Is Caused by Blockade of Selectin- and CXCL12 Protein-mediated Leukocyte Trafficking in Mice*

    PubMed Central

    Zhang, Siyuan; Condac, Eduard; Qiu, Hong; Jiang, Junlin; Gutierrez-Sanchez, Gerardo; Bergmann, Carl; Handel, Tracy; Wang, Lianchun

    2012-01-01

    Leukocytosis refers to an increase in leukocyte count above the normal range in the blood and is a common laboratory finding in patients. In many cases, the mechanisms underlying leukocytosis are not known. In this study, we examined the effects, the structural determinants, and the underlying mechanisms of heparin-induced leukocytosis, a side effect occurring in 0.44% of patients receiving heparin. We observed that heparin induced both lymphocytosis and neutrophilia, and the effects required heparin to be 6-O-sulfated but did not require its anticoagulant activity. Cell mobilization studies revealed that the lymphocytosis was attributable to a combination of blockage of lymphocyte homing and the release of thymocytes from the thymus, whereas the neutrophilia was caused primarily by neutrophil release from the bone marrow and demargination in the vasculature. Mechanistic studies revealed that heparin inhibits L- and P-selectin, as well as the chemokine CXCL12, leading to leukocytosis. Heparin is known to require 6-O-sulfate to inhibit L- and P-selectin function, and in this study we observed that 6-O-sulfate is required for its interaction with CXCL12. We conclude that heparin-induced leukocytosis requires glucosamine 6-O-sulfation and is caused by blockade of L-selectin-, P-selectin-, and CXCL12-mediated leukocyte trafficking. PMID:22194593

  19. Pantethine Alters Lipid Composition and Cholesterol Content of Membrane Rafts, With Down-Regulation of CXCL12-Induced T Cell Migration.

    PubMed

    van Gijsel-Bonnello, Manuel; Acar, Niyazi; Molino, Yves; Bretillon, Lionel; Khrestchatisky, Michel; de Reggi, Max; Gharib, Bouchra

    2015-10-01

    Pantethine, a natural low-molecular-weight thiol, shows a broad activity in a large range of essential cellular pathways. It has been long known as a hypolipidemic and hypocholesterolemic agent. We have recently shown that it exerts a neuroprotective action in mouse models of cerebral malaria and Parkinson's disease through multiple mechanisms. In the present study, we looked at its effects on membrane lipid rafts that serve as platforms for molecules engaged in cell activity, therefore providing a target against inappropriate cell response leading to a chronic inflammation. We found that pantethine-treated cells showed a significant change in raft fatty acid composition and cholesterol content, with ultimate downregulation of cell adhesion, CXCL12-driven chemotaxis, and transendothelial migration of various T cell types, including human Jurkat cell line and circulating effector T cells. The mechanisms involved include the alteration of the following: (i) CXCL12 binding to its target cells; (ii) membrane dynamics of CXCR4 and CXCR7, the two CXCL12 receptors; and (iii) cell redox status, a crucial determinant in the regulation of the chemokine system. In addition, we considered the linker for activation of T cells molecule to show that pantethine effects were associated with the displacement from the rafts of the acylated signaling molecules which had their palmitoylation level reduced.. In conclusion, the results presented here, together with previously published findings, indicate that due to its pleiotropic action, pantethine can downregulate the multifaceted process leading to pathogenic T cell activation and migration.

  20. CXCL12 and TP53 genetic polymorphisms as markers of susceptibility in a Brazilian children population with acute lymphoblastic leukemia (ALL).

    PubMed

    de Lourdes Perim, Aparecida; Guembarovski, Roberta Losi; Oda, Julie Massayo Maeda; Lopes, Leandra Fiori; Ariza, Carolina Batista; Amarante, Marla Karine; Fungaro, Maria Helena Pelegrinelli; de Oliveira, Karen Brajão; Watanabe, Maria Angelica Ehara

    2013-07-01

    Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Genetic polymorphisms in the 3'UTR region of the CXCL12 (rs1801157) and TP53 codon 72 (rs1042522) genes may contribute to susceptibility to childhood ALL because they affect some important processes, such as metastasis regulation and tumor suppression. Thus the objective of the present study was to detect the frequency of two genetic polymorphisms in ALL patients and controls and to add information their impact on genetic susceptibility and prognosis. The CXCL12 and TP53 polymorphisms were tested in 54 ALL child patients and in 58 controls by restriction fragment length polymerase chain reaction and allelic specific chain reaction techniques, respectively. The frequencies of both allelic variants were higher in ALL patients than in the controls and indicated a positive association: OR = 2.44; 95 % CI 1.05-5.64 for CXCL12 and OR = 2.20; 95 % CI 1.03-4.70 for TP53. Furthermore, when the two genetic variants were analyzed together, they increased significantly more than fivefold the risk of this neoplasia development (OR = 5.24; 95 % CI 1.39-19.75), indicating their potential as susceptibility markers for ALL disease and the relevance of the allelic variant combination to increased risk of developing malignant tumors. Future studies may indicate a larger panel of genes involved in susceptibility of childhood ALL and other hematological neoplasias.

  1. Implication of IRF4 aberrant gene expression in the acute leukemias of childhood.

    PubMed

    Adamaki, Maria; Lambrou, George I; Athanasiadou, Anastasia; Tzanoudaki, Marianna; Vlahopoulos, Spiros; Moschovi, Maria

    2013-01-01

    The most frequent targets of genetic alterations in human leukemias are transcription factor genes with essential functions in normal blood cell development. The Interferon Regulatory Factor 4 (IRF4) gene encodes a transcription factor important for key developmental stages of hematopoiesis, with known oncogenic implications in multiple myeloma, adult leukemias and lymphomas. Very few studies have reported an association of IRF4 with childhood malignancy, whereas high transcript levels have been observed in the more mature immunophenotype of ALL. Our aim was to investigate the expression levels of IRF4 in the diagnostic samples of pediatric leukemias and compare them to those of healthy controls, in order to determine aberrant gene expression and whether it extends to leukemic subtypes other than the relatively mature ALL subpopulation. Quantitative real-time RT-PCR methodology was used to investigate IRF4 expression in 58 children with acute leukemias, 4 leukemic cell lines and 20 healthy children. We show that aberrant IRF4 gene expression is implicated in a variety of leukemic subtypes; higher transcript levels appear in the more immature B-common ALL subtype and in T-cell than in B-cell leukemias, with the highest expression levels appearing in the AML group. Interestingly, we show that childhood leukemia, irrespective of subtype or cell maturation stage, is characterised by a minimum of approximately twice the amount of IRF4 gene expression encountered in healthy children. A statistically significant correlation also appeared to exist between high IRF4 expression and relapse. Our results show that ectopic expression of IRF4 follows the reverse expression pattern of what is encountered in normal B-cell development and that there might be a dose-dependency of childhood leukemia for aberrantly expressed IRF4, a characteristic that could be explored therapeutically. It is also suggested that high IRF4 expression might be used as an additional prognostic marker of

  2. Aberrant expression of laminin-332 promotes cell proliferation and cyst growth in ARPKD

    PubMed Central

    Dang, Suparna; Marinkovich, M. Peter; Lazarova, Zelmira; Yoder, Bradley; Torres, Vicente E.; Wallace, Darren P.

    2013-01-01

    Basement membrane abnormalities have often been observed in kidney cysts of polycystic kidney disease (PKD) patients and animal models. There is an abnormal deposition of extracellular matrix molecules, including laminin-α3,β3,γ2 (laminin-332), in human autosomal dominant PKD (ADPKD). Knockdown of PKD1 paralogs in zebrafish leads to dysregulated synthesis of the extracellular matrix, suggesting that altered basement membrane assembly may be a primary defect in ADPKD. In this study, we demonstrate that laminin-332 is aberrantly expressed in cysts and precystic tubules of human autosomal recessive PKD (ARPKD) kidneys as well as in the kidneys of PCK rats, an orthologous ARPKD model. There was aberrant expression of laminin-γ2 as early as postnatal day 2 and elevated laminin-332 protein in postnatal day 30, coinciding with the formation and early growth of renal cysts in PCK rat kidneys. We also show that a kidney cell line derived from Oak Ridge polycystic kidney mice, another model of ARPKD, exhibited abnormal lumen-deficient and multilumen structures in Matrigel culture. These cells had increased proliferation rates and altered expression levels of laminin-332 compared with their rescued counterparts. A function-blocking polyclonal antibody to laminin-332 significantly inhibited their abnormal proliferation rates and rescued their aberrant phenotype in Matrigel culture. Furthermore, abnormal laminin-332 expression in cysts originating from collecting ducts and proximal tubules as well as in precystic tubules was observed in a human end-stage ADPKD kidney. Our results suggest that abnormal expression of laminin-332 contributes to the aberrant proliferation of cyst epithelial cells and cyst growth in genetic forms of PKD. PMID:24370592

  3. Aberrant Expression of MICO1 and MICO1OS in Deceased Somatic Cell Nuclear Transfer Calves.

    PubMed

    Wang, Guan-Nan; Yang, Wen-Zhi; Xu, Da; Li, Dong-Jie; Zhang, Cui; Chen, Wei-Na; Li, Shi-Jie

    2017-04-06

    Incomplete reprogramming of a donor nucleus following somatic cell nuclear transfer (SCNT) results in aberrant expression of developmentally important genes, and is the primary source of the phenotypic abnormalities observed in cloned animals. Expression of non-coding RNAs in the murine Dlk1-Dio3 imprinted domain was previously shown to correlate with the pluripotency of mouse induced pluripotent stem cells. In this study, we examined the transcription of the bovine orthologs from this locus, MICO1 (Maternal intergenic circadian oscillating 1) and MICO1OS (MICO1 opposite strand), in tissues from artificially inseminated and SCNT calves that died during the perinatal period. A single-nucleotide polymorphism (SNP), a T-to-C transition, was used to analyze the allelic transcription of MICO1. Our results indicate monoallelic expression of the MICO1 C allele among the six analyzed tissues (heart, liver, spleen, lung, kidney, and brain) of artificially inseminated calves, indicating that this gene locus may be imprinted in bovine. Conversely, we observed variable allelic transcription of MICO1 in SCNT calves. We asked if DNA methylation regulated the monoallelic expression of MICO1 and MICO1OS by evaluating the methylation levels of six regions within or around this locus in tissues with normal or aberrant MICO1 transcription; all of the samples from either artificially inseminated or SCNT calves exhibited hypermethylation, implying that DNA methylation may not be involved in regulating its monoallelic expression. Furthermore, three imprinted genes (GTL2, MEG9, and DIO3) nearby MICO1 showed monoallelic expression in SCNT calves with aberrant MICO1 transcription, indicating that not all of the genes in the bovine DLK1-DIO3 domain are mis-regulated. This article is protected by copyright. All rights reserved.

  4. Aberrant expression of HOXA9, DEK, CBL and CSF1R in acute myeloid leukemia.

    PubMed

    Casas, Sílvia; Nagy, Bálint; Elonen, Erkki; Aventín, Anna; Larramendy, Marcelo L; Sierra, Jorge; Ruutu, Tapani; Knuutila, Sakari

    2003-11-01

    Previous gene function analyses have indicated that HOXA9, DEK, CBL and CSF1R are aberrantly expressed in acute myeloid leukemia (AML). We analyzed the expression of these genes in a series of 41 adult patients with AML using quantitative real-time RT-PCR, and tested the association of the expression with the following hematologic and clinical parameters: age, FAB, immunophenotype and karyotype aberrations. A high proportion of the patients showed over- or underexpression of the analyzed genes. DEK was overexpressed in 98% of the cases, whereas CBL, CSF1R and HOXA9 were either overexpressed in 20%, 17% and 78% or underexpressed in 20%, 42% and 15% of the cases, respectively. Patients whose karyotype contained t(8;21)(q22;q22), showed lower relative expression of HOXA9 at a statistically significant level (p < 0.05). Bone marrow samples without expression of CD34 antigen were associated with either overexpression of DEK or HOXA9. Furthermore, an association was found between the AML-M2 subtype and lower expression of CBL, CSF1R or HOXA9, and between the AML-M5 subtype and CBL or CSF1R overexpression.

  5. Aberrant epigenetic changes and gene expression in cloned cattle dying around birth

    PubMed Central

    Lin, Li; Li, Qiang; Zhang, Lei; Zhao, Dingsheng; Dai, Yunping; Li, Ning

    2008-01-01

    Background Aberrant reprogramming of donor somatic cell nuclei may result in many severe problems in animal cloning. To assess the extent of abnormal epigenetic modifications and gene expression in clones, we simultaneously examined DNA methylation, histone H4 acetylation and expression of six genes (β-actin, VEGF, oct4, TERT, H19 and Igf2) and a repetitive sequence (art2) in five organs (heart, liver, spleen, lung and kidney) from two cloned cattle groups that had died at different stages. In the ED group (early death, n = 3), the cloned cattle died in the perinatal period. The cattle in the LD group (late death, n = 3) died after the perinatal period. Normally reproduced cattle served as a control group (n = 3). Results Aberrant DNA methylation, histone H4 acetylation and gene expression were observed in both cloned groups. The ED group showed relatively fewer severe DNA methylation abnormalities (p < 0.05) but more abnormal histone H4 acetylations (p < 0.05) and more abnormal expression (p < 0.05) of the selected genes compared to the LD group. However, our data also suggest no widespread gene expression abnormalities in the organs of the dead clones. Conclusion Deaths of clones may be ascribed to abnormal expression of a very limited number of genes. PMID:18261243

  6. Aberrant expression of laminin γ2 correlates with poor prognosis and promotes invasion in extrahepatic cholangiocarcinoma.

    PubMed

    Liu, Wei; Tian, Feng; Jiang, Peng; Zhao, Xin; Guo, Fei; Li, Xiaowu; Wang, Shuguang

    2014-01-01

    To investigate the potential role of laminin γ2 and its correlation with prognosis in patients with extrahepatic cholangiocarcinoma (CCA). Laminin γ2 expression was evaluated by immunohistochemistry in 72 extrahepatic CCA patients after surgical resection. Knockdown of laminin γ2 was achieved via small interfering RNA transfection in the extrahepatic CCA cell line QBC939. Thirty-six of 72 extrahepatic CCAs (50%) stained positive for laminin γ2 in two types of patterns: stromal staining (28/72, 39%) and cytoplasmic staining (24/72, 33%). All 16 paracancerous tissue samples showed negative staining. Both stromal and cytoplasmic laminin γ2 expressions correlated with lymph node metastasis. Kaplan-Meier analysis showed that aberrant expression of laminin γ2 correlated with poor overall survival and early recurrence. Cox regression analysis further demonstrated that laminin γ2 expression was a significant independent predictor of poor overall survival and early recurrence. Immunofluorescence staining revealed cytoplasmic expression of laminin γ2 in QBC939 cells. Knockdown of laminin γ2 significantly reduced QBC939 cell invasion and migration. Aberrant expression of laminin γ2 correlates with poor prognosis and promotes invasion in extrahepatic CCA. Crown Copyright © 2014. Published by Elsevier Inc. All rights reserved.

  7. Aberrant expression of intelectin-1 in gastric cancer: its relationship with clinicopathological features and prognosis.

    PubMed

    Zheng, Liduan; Weng, Mixia; Qi, Meng; Qi, Teng; Tong, Ling; Hou, Xiaohua; Tong, Qiangsong

    2012-01-01

    Human intelectin-1 (ITLN-1) is a novel identified galactose-binding lectin that is expressed in the colonic goblet cells. Since gastric adenocarcinomas can arise through a process of intestinalization, we speculate that ITLN-1 may be aberrantly expressed in gastric cancer. This study was undertaken to examine the ITLN-1 expression in gastric cancer and correlate it with clinical outcomes. One hundred and ninety-six gastric cancer patients were evaluated for the ITLN-1 expression by immunohistochemistry. The ITLN-1 transcripts were measured by real-time quantitative PCR. ITLN-1 expression was absent in normal gastric mucosa, whereas areas of intestinal metaplasia revealed ITLN-1 immunoreactivity. One hundred and forty-two gastric cancer patients (72.4%) were positive for ITLN-1 expression. In a subtotal of 20 patients, ITLN-1 transcripts were significantly enhanced in gastric cancer tissues than in normal gastric mucosa (P < 0.001). The expression rate of ITLN-1 was higher in intestinal-type carcinomas than in diffuse-type carcinomas (P = 0.003). ITLN-1 positivity in gastric cancer was positively correlated with tumor differentiation (P = 0.001) and CDX2 expression (P < 0.001), and inversely correlated with depth of invasion (P = 0.007), lymph node metastasis (P = 0.001), distant metastasis (P = 0.014), clinical stage (P = 0.006), Ki-67 expression (P = 0.001), and heparanase expression (P < 0.001), without correlation with age, gender, tumor location, or tumor size. In univariate and multivariate analyses, ITLN-1 was an independent prognostic factor for longer survival of gastric cancer patients (P = 0.001). The aberrant ITLN-1 expression in gastric cancer is correlated with clinicopathological features and may be a useful prognostic factor for predicting the outcomes of gastric cancer patients.

  8. Aberrant DNA methylation impacts gene expression and prognosis in breast cancer subtypes.

    PubMed

    Győrffy, Balázs; Bottai, Giulia; Fleischer, Thomas; Munkácsy, Gyöngyi; Budczies, Jan; Paladini, Laura; Børresen-Dale, Anne-Lise; Kristensen, Vessela N; Santarpia, Libero

    2016-01-01

    DNA methylation has a substantial impact on gene expression, affecting the prognosis of breast cancer (BC) patients dependent on molecular subtypes. In this study, we investigated the prognostic relevance of the expression of genes reported as aberrantly methylated, and the link between gene expression and DNA methylation in BC subtypes. The prognostic value of the expression of 144 aberrantly methylated genes was evaluated in ER+/HER2-, HER2+, and ER-/HER2- molecular BC subtypes, in a meta-analysis of two large transcriptomic cohorts of BC patients (n = 1,938 and n = 1,640). The correlation between gene expression and DNA methylation in distinct gene regions was also investigated in an independent dataset of 104 BCs. Survival and Pearson correlation analyses were computed for each gene separately. The expression of 48 genes was significantly associated with BC prognosis (p < 0.05), and 32 of these prognostic genes exhibited a direct expression-methylation correlation. The expression of several immune-related genes, including CD3D and HLA-A, was associated with both relapse-free survival (HR = 0.42, p = 3.5E-06; HR = 0.35, p = 1.7E-08) and overall survival (HR = 0.50, p = 5.5E-04; HR = 0.68, p = 4.5E-02) in ER-/HER2- BCs. On the overall, the distribution of both positive and negative expression-methylation correlation in distinct gene regions have different effects on gene expression and prognosis in BC subtypes. This large-scale meta-analysis allowed the identification of several genes consistently associated with prognosis, whose DNA methylation could represent a promising biomarker for prognostication and clinical stratification of patients with distinct BC subtypes.

  9. Elucidation of the Molecular Mechanisms for Aberrant Expression of Breast Cancer Specific Gene 1 in Invasive and Metastatic Breast Carcinomas

    DTIC Science & Technology

    2004-06-01

    cells in mitosis. Mutations in any of these genes result in failure to arrest Keywords: BCSG I: BubRl; mitotic checkpoint; yeast the cell cycle at G2...AD Award Number: DAMD17-02-1-0534 TITLE: Elucidation of the Molecular Mechanisms for Aberrant Expression of Breast Cancer Specific Gene 1 in Invasive...SUBTITLE 5. FUNDING NUMBERS Elucidation of the Molecular Mechanisms for Aberrant DAMD17-02-1-0534 Expression of Breast Cancer Specific Gene 1 in Invasive

  10. Migration of engrafted neural stem cells is mediated by CXCL12 signaling through CXCR4 in a viral model of multiple sclerosis.

    PubMed

    Carbajal, Kevin S; Schaumburg, Christopher; Strieter, Robert; Kane, Joy; Lane, Thomas E

    2010-06-15

    Multiple sclerosis (MS) is a human demyelinating disease characterized by multifocal regions of inflammation, progressive myelin loss within the central nervous system (CNS), and eventual failure to remyelinate damaged axons. These problems suggest deficiencies in recruiting and/or maturation of oligodendrocyte progentior cells (OPCs) and highlight cell replacement therapies to promote remyelination. We have used a model of viral-induced demyelination to characterize signaling cues associated with positional migration of transplanted remyelination-competent cells. Although successful transplantation of rodent-derived glial cell types into models of MS has been performed, the mechanisms by which these cells navigate within an inflammatory environment created by a persistent virus has not been defined. Infection of the mouse CNS with the neurotropic JHM strain of mouse hepatitis virus (JHMV) results in an immune-mediated demyelinating disease with clinical and histologic similarities to MS. Surgical engraftment of GFP+ neural stem cells (NSCs) into spinal cords of JHMV-infected mice with established demyelination results in migration, proliferation, and differentiation of the cells into OPCs and mature oligodendrocytes that is associated with increased axonal remyelination. Treatment with anti-CXCL12 [stromal derived factor-1alpha, (SDF-1alpha)] blocking serum resulted in a marked impairment in migration and proliferation of engrafted stem cells. Moreover, small molecule-mediated antagonism of CXCR4, but not CXCR7, impaired migration and proliferation, to an extent similar to that with anti-CXCL12 treatment. These data highlight the importance of the CXCL12:CXCR4 pathway in regulating homing of engrafted stem cells to sites of tissue damage within the CNS of mice persistently infected with a neurotropic virus undergoing immune-mediated demyelination.

  11. Early fibroblast progenitor cell migration to the AngII-exposed myocardium is not CXCL12 or CCL2 dependent as previously thought.

    PubMed

    Falkenham, Alec; Sopel, Mryanda; Rosin, Nicole; Lee, Tim D G; Issekutz, Thomas; Légaré, Jean-Francois

    2013-08-01

    Fibroblast progenitor cells (fibrocytes) are important to the development of myocardial fibrosis and are suggested to migrate to the heart via CXCL12 and chemokine ligand (CCL) 2. We hypothesized that if these chemokines are recruiting fibrocytes, disrupting their signaling will reduce early (3-day) fibrocyte infiltration and, consequently, fibrosis in the myocardium. C57/Bl6 and CCR2(-/-) mice were infused with saline or angiotensin (Ang) II, with or without CXC receptor 4 blockade (AMD3100). Hearts were assessed for chemokine up-regulation, immunofluorescence, and histological features. AngII caused early myocardial up-regulation of CXCL12 and CCL2, which corresponded to significant myocardial infiltration and fibrosis compared with controls. Animals receiving AMD3100 and/or with the genotype CCR2(-/-) failed to demonstrate reductions in infiltrate or fibrosis after 3 days of AngII, and AngII + AMD3100 animals showed exacerbated fibrocyte infiltration and fibrosis compared with AngII alone. CCR2(-/-) mice demonstrated significant reductions in myocardial fibrosis relative to wild type, but this was after 28 days of AngII infusion and was the result of reduced infiltrating cell proliferation. An alternative CCR2 ligand, CCL12, was found to be increasing infiltrating cell proliferation in the heart after AngII infusion, which we confirmed in vitro. In conclusion, early fibrocyte recruitment cannot be inhibited through modulating CXCL12 or CCL2, as previously thought. Ablating CCR2 signaling did confer myocardial fibrosis reductions, but these benefits were not observed until much later and were likely the result of modulated proliferation through ablating the CCL12-CCR2 interaction.

  12. Identification of aberrantly expressed glycans in gastric cancer by integrated lectin microarray and mass spectrometric analyses

    PubMed Central

    Li, Xiang; Guan, Feng; Li, Dongliang; Tan, Zengqi; Yang, Ganglong; Wu, Yanli; Huang, Zhaohui

    2016-01-01

    Cancer progression is usually associated with alterations of glycan expression patterns. Little is known regarding global glycomics in gastric cancer, the most common type of epithelial cancer. We integrated lectin microarray and mass spectrometry (MS) methods to profile glycan expression in three gastric cancer cell lines (SGC-7901, HGC-27, and MGC-803) and one normal gastric epithelial cell line (GES-1). Significantly altered glycans were confirmed by lectin staining and MALDI-TOF/TOF-MS. The three cancer cell lines showed increased levels of core-fucosylated N-glycans, GalNAcα-Ser/Thr (Tn antigen), and Sia2-6Galβ1-4GlcNAc N-glycans, but reduced levels of biantennary N-glycans, Galβ1-3GalNAcα-Ser/Thr (T antigen), and (GlcNAc)n N-glycans. Lectin histochemistry was used to validate aberrant expression of four representative glycans (core-fucosylation, Sia2-6Galβ1-4GlcNAc, biantennary N-glycans, T antigen, recognized respectively by lectins LCA, SNA, PHA-E+L, and ACA) in clinical gastric cancer samples. Lower binding capacity for ACA was correlated with significantly poorer patient prognosis. Our findings indicate for the first time that glycans recognized by LCA, ACA, and PHA-E+L are aberrantly expressed in gastric cancer, and suggest that ACA is a potential prognostic factor for gastric cancer. PMID:27895315

  13. Aberrant phenotypic expression of CD15 and CD56 identifies poor prognostic acute promyelocytic leukemia patients.

    PubMed

    Breccia, Massimo; De Propris, Maria Stefania; Minotti, Clara; Stefanizzi, Caterina; Raponi, Sara; Colafigli, Gioia; Latagliata, Roberto; Guarini, Anna; Foà, Robin

    2014-02-01

    Limited information is available on the relationship between expression of some additional aberrant phenotypic features and outcome of acute promyelocytic leukemia (APL) patients. Here, we set out to assess the frequency of CD15 and CD56 expression, and their prognostic value in a large series of APL patients. One hundred and fourteen adult patients consecutively diagnosed with PML/RARα-positive APL and homogeneously treated with the AIDA induction schedule at a single institution were included in the study. Twelve (10.5%) and 9 (8%) of the 114 patients expressed CD15 and CD56, respectively. CD15 expression identified a subset of patients with a classic morphologic subtype (92%), a prevalent association with a bcr1 expression (67%) with an unexpectedly higher frequency of relapses (42% vs 20% for the CD15- patients, p=0.03) and a low overall survival (OS) (median OS at 5 years 58% vs 85% for the CD15- patients, p=0.01). CD56 expression was detected only in patients with a classic morphologic subtype, a prevalent bcr3 expression (67%), high incidence of differentiation syndrome (55%), higher frequency of relapse (34% vs 20% for the CD56- population, p=0.04) and a low OS (60% vs 85% for the CD56- population p=0.02). We hereby confirm the negative prognostic value of CD56 and we show that the same applies also to cases expressing CD15. These aberrant markers may be considered for the refinement of risk-adapted therapeutic strategies in APL patients.

  14. Increased expression and aberrant localization of mucin 13 in metastatic colon cancer.

    PubMed

    Gupta, Brij K; Maher, Diane M; Ebeling, Mara C; Sundram, Vasudha; Koch, Michael D; Lynch, Douglas W; Bohlmeyer, Teresa; Watanabe, Akira; Aburatani, Hiroyuki; Puumala, Susan E; Jaggi, Meena; Chauhan, Subhash C

    2012-11-01

    MUC13 is a newly identified transmembrane mucin. Although MUC13 is known to be overexpressed in ovarian and gastric cancers, limited information is available regarding the expression of MUC13 in metastatic colon cancer. Herein, we investigated the expression profile of MUC13 in colon cancer using a novel anti-MUC13 monoclonal antibody (MAb, clone ppz0020) by immunohistochemical (IHC) analysis. A cohort of colon cancer samples and tissue microarrays containing adjacent normal, non-metastatic colon cancer, metastatic colon cancer, and liver metastasis tissues was used in this study to investigate the expression pattern of MUC13. IHC analysis revealed significantly higher (p<0.001) MUC13 expression in non-metastatic colon cancer samples compared with faint or very low expression in adjacent normal tissues. Interestingly, metastatic colon cancer and liver metastasis tissue samples demonstrated significantly (p<0.05) higher cytoplasmic and nuclear MUC13 expression compared with non-metastatic colon cancer and adjacent normal colon samples. Moreover, cytoplasmic and nuclear MUC13 expression correlated with larger and poorly differentiated tumors. Four of six tested colon cancer cell lines also expressed MUC13 at RNA and protein levels. These studies demonstrate a significant increase in MUC13 expression in metastatic colon cancer and suggest a correlation between aberrant MUC13 localization (cytoplasmic and nuclear expression) and metastatic colon cancer.

  15. Increased Expression and Aberrant Localization of Mucin 13 in Metastatic Colon Cancer

    PubMed Central

    Gupta, Brij K.; Maher, Diane M.; Ebeling, Mara C.; Sundram, Vasudha; Koch, Michael D.; Lynch, Douglas W.; Bohlmeyer, Teresa; Watanabe, Akira; Aburatani, Hiroyuki; Puumala, Susan E.; Jaggi, Meena

    2012-01-01

    MUC13 is a newly identified transmembrane mucin. Although MUC13 is known to be overexpressed in ovarian and gastric cancers, limited information is available regarding the expression of MUC13 in metastatic colon cancer. Herein, we investigated the expression profile of MUC13 in colon cancer using a novel anti-MUC13 monoclonal antibody (MAb, clone ppz0020) by immunohistochemical (IHC) analysis. A cohort of colon cancer samples and tissue microarrays containing adjacent normal, non-metastatic colon cancer, metastatic colon cancer, and liver metastasis tissues was used in this study to investigate the expression pattern of MUC13. IHC analysis revealed significantly higher (p<0.001) MUC13 expression in non-metastatic colon cancer samples compared with faint or very low expression in adjacent normal tissues. Interestingly, metastatic colon cancer and liver metastasis tissue samples demonstrated significantly (p<0.05) higher cytoplasmic and nuclear MUC13 expression compared with non-metastatic colon cancer and adjacent normal colon samples. Moreover, cytoplasmic and nuclear MUC13 expression correlated with larger and poorly differentiated tumors. Four of six tested colon cancer cell lines also expressed MUC13 at RNA and protein levels. These studies demonstrate a significant increase in MUC13 expression in metastatic colon cancer and suggest a correlation between aberrant MUC13 localization (cytoplasmic and nuclear expression) and metastatic colon cancer. PMID:22914648

  16. Role of SDF-1 (CXCL12) in regulating hematopoietic stem and progenitor cells traffic into the liver during extramedullary hematopoiesis induced by G-CSF, AMD3100 and PHZ.

    PubMed

    Mendt, Mayela; Cardier, Jose E

    2015-12-01

    The stromal cell derived factor 1 (SDF-1/CXCL12) plays an essential role in the homing of hematopoietic stem and progenitor cells (HSPCs) to bone marrow (BM). It is not known whether SDF-1 may also regulate the homing of HSPCs to the liver during extramedullary hematopoiesis (EMH). Here, we investigated the possible role of SDF-1 in attracting HSPCs to the liver during experimental EMH induced by the hematopoietic mobilizers G-CSF, AMD3100 and phenylhydrazine (PHZ). Mice treated with G-CSF, AMD3100 and PHZ showed a significant increase in the expression of SDF-1 in the liver sinusoidal endothelial cells (LSECs) microenvironments. Liver from mice treated with the hematopoietic mobilizers showed HSPCs located adjacent to the LSEC microenvironments, expressing high levels of SDF-1. An inverse relationship was found between the hepatic SDF-1 levels and those in the BM. In vitro, LSEC monolayers induced the migration of HSPCs, and this effect was significantly reduced by AMD3100. In conclusion, our results provide the first evidence showing that SDF-1 expressed by LSEC can be a major player in the recruitment of HSPCs to the liver during EMH induced by hematopoietic mobilizers.

  17. Pervasive transcription read-through promotes aberrant expression of oncogenes and RNA chimeras in renal carcinoma

    PubMed Central

    Grosso, Ana R; Leite, Ana P; Carvalho, Sílvia; Matos, Mafalda R; Martins, Filipa B; Vítor, Alexandra C; Desterro, Joana MP; Carmo-Fonseca, Maria; de Almeida, Sérgio F

    2015-01-01

    Aberrant expression of cancer genes and non-canonical RNA species is a hallmark of cancer. However, the mechanisms driving such atypical gene expression programs are incompletely understood. Here, our transcriptional profiling of a cohort of 50 primary clear cell renal cell carcinoma (ccRCC) samples from The Cancer Genome Atlas (TCGA) reveals that transcription read-through beyond the termination site is a source of transcriptome diversity in cancer cells. Amongst the genes most frequently mutated in ccRCC, we identified SETD2 inactivation as a potent enhancer of transcription read-through. We further show that invasion of neighbouring genes and generation of RNA chimeras are functional outcomes of transcription read-through. We identified the BCL2 oncogene as one of such invaded genes and detected a novel chimera, the CTSC-RAB38, in 20% of ccRCC samples. Collectively, our data highlight a novel link between transcription read-through and aberrant expression of oncogenes and chimeric transcripts that is prevalent in cancer. DOI: http://dx.doi.org/10.7554/eLife.09214.001 PMID:26575290

  18. AMPK Promotes Aberrant PGC1β Expression To Support Human Colon Tumor Cell Survival

    PubMed Central

    Fisher, Kurt W.; Das, Binita; Kim, Hyun Seok; Clymer, Beth K.; Gehring, Drew; Smith, Deandra R.; Costanzo-Garvey, Diane L.; Fernandez, Mario R.; Brattain, Michael G.; Kelly, David L.; MacMillan, John

    2015-01-01

    A major goal of cancer research is the identification of tumor-specific vulnerabilities that can be exploited for the development of therapies that are selectively toxic to the tumor. We show here that the transcriptional coactivators peroxisome proliferator-activated receptor gamma coactivator 1β (PGC1β) and estrogen-related receptor α (ERRα) are aberrantly expressed in human colon cell lines and tumors. With kinase suppressor of Ras 1 (KSR1) depletion as a reference standard, we used functional signature ontology (FUSION) analysis to identify the γ1 subunit of AMP-activated protein kinase (AMPK) as an essential contributor to PGC1β expression and colon tumor cell survival. Subsequent analysis revealed that a subunit composition of AMPK (α2β2γ1) is preferred for colorectal cancer cell survival, at least in part, by stabilizing the tumor-specific expression of PGC1β. In contrast, PGC1β and ERRα are not detectable in nontransformed human colon epithelial cells, and depletion of the AMPKγ1 subunit has no effect on their viability. These data indicate that Ras oncogenesis relies on the aberrant activation of a PGC1β-dependent transcriptional pathway via a specific AMPK isoform. PMID:26351140

  19. Aberrant P-cadherin expression is associated to aggressive feline mammary carcinomas.

    PubMed

    Figueira, Ana Catarina; Gomes, Catarina; de Oliveira, Joana Tavares; Vilhena, Hugo; Carvalheira, Júlio; de Matos, Augusto J F; Pereira, Patrícia Dias; Gärtner, Fátima

    2014-11-26

    Cadherins are calcium-dependent cell-to-cell adhesion glycoproteins playing a critical role in the formation and maintenance of normal tissue architecture. In normal mammary gland, E-cadherin is expressed by luminal epithelial cells, while P-cadherin is restricted to myoepithelial cells. Changes in the expression of classical E- and P-cadherins have been observed in mammary lesions and related to mammary carcinogenesis. P-cadherin and E-cadherin expressions were studied in a series of feline normal mammary glands, hyperplastic/dysplastic lesions, benign and malignant tumours by immunohistochemistry and double-label immunofluorescence. In normal tissue and in the majority of hyperplastic/dysplastic lesions and benign tumours, P-cadherin was restricted to myoepithelial cells, while 80% of the malignant tumours expressed P-cadherin in luminal epithelial cells. P-cadherin expression was significantly related to high histological grade of carcinomas (p <0.0001), tumour necrosis (p = 0.001), infiltrative growth (p = 0.0051), and presence of neoplastic emboli (p = 0.0401). Moreover, P-cadherin positive carcinomas had an eightfold likelihood of developing neoplastic emboli than negative tumours. Cadherins expression profile in high grade and in infiltrative tumours was similar, the majority expressing P-cadherin, regardless of E-cadherin expression status. The two cadherins were found to be co-expressed in carcinomas with aberrant P-cadherin expression and preserved E-cadherin. The results demonstrate a relationship between P-cadherin expression and aggressive biological behaviour of feline mammary carcinomas, suggesting that P-cadherin may be considered an indicator of poor prognosis in this animal species. Moreover, it indicates that, in queens, the aberrant expression of P-cadherin is a better marker of mammary carcinomas aggressive behaviour than the reduction of E-cadherin expression. Further investigation with follow-up studies in feline species should be conducted

  20. Gene expression and epigenetic aberrations in F1-placentas fathered by obese males.

    PubMed

    Mitchell, Megan; Strick, Reiner; Strissel, Pamela L; Dittrich, Ralf; McPherson, Nicole O; Lane, Michelle; Pliushch, Galyna; Potabattula, Ramya; Haaf, Thomas; El Hajj, Nady

    2017-02-10

    Gene expression and/or epigenetic deregulation may have consequences for sperm and blastocysts, as well as for the placenta, together potentially contributing to problems observed in offspring. We previously demonstrated specific perturbations of fertilization, blastocyst formation, implantation, as well as aberrant glucose metabolism and adiposity in offspring using a mouse model of paternal obesity. The current investigation analyzed gene expression and methylation of specific CpG residues in F1 placentas of pregnancies fathered by obese and normal-weight male mice, using real-time PCR and bisulfite pyrosequencing. Our aim was to determine if paternal obesity deregulated placental gene expression and DNA methylation when compared to normal-weight males. Gene methylation of sperm DNA was analyzed and compared to placentas to address epigenetic transmission. Of the 10 paternally expressed genes (Pegs), 11 genes important for development and transport of nutrients, and the long-terminal repeat Intracisternal A particle (IAP) elements, derived from a member of the class II endogenous retroviral gene family, we observed a significant effect of paternal diet-induced obesity on deregulated expression of Peg3, Peg9, Peg10, and the nutrient transporter gene Slc38a2, and aberrant DNA methylation of the Peg9 promoter in F1 placental tissue. Epigenetic changes in Peg9 were also found in sperm from obese fathers. We therefore propose that paternal obesity renders changes in gene expression and/or methylation throughout the placental genome, which could contribute to the reproductive problems related to fertility and to the metabolic, long-term health impact on offspring.

  1. Sonic Hedgehog Signaling Affected by Promoter Hypermethylation Induces Aberrant Gli2 Expression in Spina Bifida.

    PubMed

    Lu, Xiao-Lin; Wang, Li; Chang, Shao-Yan; Shangguan, Shao-Fang; Wang, Zhen; Wu, Li-Hua; Zou, Ji-Zhen; Xiao, Ping; Li, Rui; Bao, Yi-Hua; Qiu, Z-Y; Zhang, Ting

    2016-10-01

    GLI2 is a key mediator of the sonic hedgehog (Shh) signaling pathway and plays an important role in neural tube development during vertebrate embryogenesis; however, the role of gli2 in human folate-related neural tube defects remains unclear. In this study, we compared methylation status and polymorphisms of gli2 between spina bifida patients and a control group to explore the underlying mechanisms related to folate deficiency in spina bifida. No single nucleotide polymorphism was found to be significantly different between the two groups, although gli2 methylation levels were significantly increased in spina bifida samples, accompanied by aberrant GLI2 expression. Moreover, a prominent negative correlation was found between the folate level in brain tissue and the gli2 methylation status (r = -0.41, P = 0.014), and gli2 hypermethylation increased the risk of spina bifida with an odds ratio of 12.45 (95 % confidence interval: 2.71-57.22, P = 0.001). In addition, we established a cell model to illustrate the effect of gli2 expression and the accessibility of chromatin affected by methylation. High gli2 and gli1 mRNA expression was detected in 5-Aza-treated cells, while gli2 hypermethylation resulted in chromatin inaccessibility and a reduced association with nuclear proteins containing transcriptional factors. More meaningful to the pathway, the effect gene of the Shh pathway, gli1, was found to have a reduced level of expression along with a decreased expression of gli2 in our cell model. Aberrant high methylation resulted in the low expression of gli2 in spina bifida, which was affected by the change in chromatin status and the capacity of transcription factor binding.

  2. Aberrant expression of DNA damage response proteins is associated with breast cancer subtype and clinical features

    PubMed Central

    Guler, Gulnur; Himmetoglu, Cigdem; Jimenez, Rafael E.; Geyer, Susan M.; Wang, Wenle P.; Costinean, Stefan; Pilarski, Robert T.; Morrison, Carl; Suren, Dinc; Liu, Jianhua; Chen, Jingchun; Kamal, Jyoti; Shapiro, Charles L.

    2013-01-01

    Landmark studies of the status of DNA damage checkpoints and associated repair functions in preneoplastic and neoplastic cells has focused attention on importance of these pathways in cancer development, and inhibitors of repair pathways are in clinical trials for treatment of triple negative breast cancer. Cancer heterogeneity suggests that specific cancer subtypes will have distinct mechanisms of DNA damage survival, dependent on biological context. In this study, status of DNA damage response (DDR)-associated proteins was examined in breast cancer subtypes in association with clinical features; 479 breast cancers were examined for expression of DDR proteins γH2AX, BRCA1, pChk2, and p53, DNA damage-sensitive tumor suppressors Fhit and Wwox, and Wwox-interacting proteins Ap2α, Ap2γ, ErbB4, and correlations among proteins, tumor subtypes, and clinical features were assessed. In a multivariable model, triple negative cancers showed significantly reduced Fhit and Wwox, increased p53 and Ap2γ protein expression, and were significantly more likely than other subtype tumors to exhibit aberrant expression of two or more DDR-associated proteins. Disease-free survival was associated with subtype, Fhit and membrane ErbB4 expression level and aberrant expression of multiple DDR-associated proteins. These results suggest that definition of specific DNA repair and checkpoint defects in subgroups of triple negative cancer might identify new treatment targets. Expression of Wwox and its interactor, ErbB4, was highly significantly reduced in metastatic tissues vs. matched primary tissues, suggesting that Wwox signal pathway loss contributes to lymph node metastasis, perhaps by allowing survival of tumor cells that have detached from basement membranes, as proposed for the role of Wwox in ovarian cancer spread. PMID:21069451

  3. Aberrantly Expressed OTX Homeobox Genes Deregulate B-Cell Differentiation in Hodgkin Lymphoma

    PubMed Central

    Nagel, Stefan; Ehrentraut, Stefan; Meyer, Corinna; Kaufmann, Maren; Drexler, Hans G.; MacLeod, Roderick A. F.

    2015-01-01

    In Hodgkin lymphoma (HL) we recently reported that deregulated homeobox gene MSX1 mediates repression of the B-cell specific transcription factor ZHX2. In this study we investigated regulation of MSX1 in this B-cell malignancy. Accordingly, we analyzed expression and function of OTX homeobox genes which activate MSX1 transcription during embryonal development in the neural plate border region. Our data demonstrate that OTX1 and OTX2 are aberrantly expressed in both HL patients and cell lines. Moreover, both OTX loci are targeted by genomic gains in overexpressing cell lines. Comparative expression profiling and subsequent pathway modulations in HL cell lines indicated that aberrantly enhanced FGF2-signalling activates the expression of OTX2. Downstream analyses of OTX2 demonstrated transcriptional activation of genes encoding transcription factors MSX1, FOXC1 and ZHX1. Interestingly, examination of the physiological expression profile of ZHX1 in normal hematopoietic cells revealed elevated levels in T-cells and reduced expression in B-cells, indicating a discriminatory role in lymphopoiesis. Furthermore, two OTX-negative HL cell lines overexpressed ZHX1 in correlation with genomic amplification of its locus at chromosomal band 8q24, supporting the oncogenic potential of this gene in HL. Taken together, our data demonstrate that deregulated homeobox genes MSX1 and OTX2 respectively impact transcriptional inhibition of (B-cell specific) ZHX2 and activation of (T-cell specific) ZHX1. Thus, we show how reactivation of a specific embryonal gene regulatory network promotes disturbed B-cell differentiation in HL. PMID:26406991

  4. Aberrant expression of interferon regulatory factor 3 in human lung cancer

    SciTech Connect

    Tokunaga, Takayuki; Naruke, Yuki; Shigematsu, Sayuri; Kohno, Tomoko; Yasui, Kiyoshi; Ma, Yuhua; Chua, Koon Jiew; Katayama, Ikuo; Nakamura, Takashi; Hishikawa, Yoshitaka; Koji, Takehiko; Yatabe, Yasushi; Nagayasu, Takeshi; Fujita, Takashi; Matsuyama, Toshifumi; and others

    2010-06-25

    We analyzed the subcellular distributions and gene structures of interferon regulatory factor 3 (IRF3) transcription factor in 50 cases of human primary lung cancer. The immunohistochemical analyses revealed substantially aberrant IRF3 expression specific to the cancer lesions (2 and 6 tumors with nuclear staining, and 4 and 5 tumors with negative staining, in adenocarcinoma and squamous cell carcinoma, respectively), while the morphologically normal region around the tumors exhibited only cytoplasmic staining. In addition, we determined the sequence of the entire IRF3 coding region, and found two novel variants with the amino acid changes (S{sup 175}(AGC) {yields} R{sup 175}(CGC) and A{sup 208}(GCC) {yields} D{sup 208}(GAC)). The R{sup 175} variant was also detected in a morphologically normal region around the nuclear staining squamous cell carcinoma, and exhibited almost the same functions as the wild type IRF3. On the other hand, the D{sup 208} variant, found in the negative staining squamous cell carcinoma cases, reduced the nuclear translocation in response to I{kappa}B kinase {epsilon} stimulation, as compared to the wild type IRF3, but the same variant was detected in the surrounding morphologically normal region. The aberrant expression of IRF3 and the novel D{sup 208} variant may provide clues to elucidate the etiology of primary lung cancer.

  5. Aberrant LPL Expression, Driven by STAT3, Mediates Free Fatty Acid Metabolism in CLL Cells

    PubMed Central

    Rozovski, Uri; Grgurevic, Srdana; Bueso-Ramos, Carlos; Harris, David M.; Li, Ping; Liu, Zhiming; Wu, Ji Yuan; Jain, Preetesh; Wierda, William; Burger, Jan; O’Brien, Susan; Jain, Nitin; Ferrajoli, Alessandra; Keating, Michael J.; Estrov, Zeev

    2015-01-01

    While reviewing chronic lymphocytic leukemia (CLL) bone marrow slides we identified cytoplasmic lipid vacuoles in CLL cells but not in normal B cells. Because lipoprotein lipase (LPL), which catalyzes hydrolysis of triglycerides into free fatty acids (FFAs), is aberrantly expressed in CLL, we investigated whether LPL regulates the oxidative metabolic capacity of CLL cells. We found that unlike normal B cells, CLL cells metabolize FFAs. Because STAT3 is constitutively activated in CLL cells and because we identified putative STAT3 binding sites in the LPL promoter, we sought to determine whether STAT3 drives the aberrant expression of LPL. Transfection of luciferase reporter gene constructs driven by LPL promoter fragments into MM1 cells revealed that STAT3 activates the LPL promoter. In addition, chromatin immunoprecipitation (ChIP) confirmed that STAT3 binds to the LPL promoter. Furthermore, transfection of CLL cells with STAT3-shRNA downregulated LPL transcripts and protein levels, confirming that STAT3 activates the LPL gene. Finally, transfection of CLL cells with LPL-siRNAs decreased the capacity of CLL cells to oxidize FFAs and reduced cell viability. PMID:25733697

  6. Aberrant expression and hormonal regulation of Galectin-3 in endometriosis women with infertility.

    PubMed

    Yang, H; Yin, J; Ficarrotta, K; Hsu, S H; Zhang, W; Cheng, C

    2016-07-01

    To investigate the role and potential molecular mechanism of Galectin-3 (Gal-3) in the etiology of endometriosis-associated infertility. We detected Gal-3 expression in eutopic endometrium from women with endometriosis-associated infertility and healthy women without endometriosis or infertility. We then evaluated Gal-3 expression in endometrial glandular epithelial cells (EECs) and endometrial stromal cells (ESCs) and investigated its response to hormone stimulation in EECs and ESCs from both groups of women. Results of real-time PCR and western blot analysis showed Gal-3 expression in both proliferative and secretory stages of the menstrual cycle decreased significantly in women with endometriosis-associated infertility compared to healthy women. The changes in expression of Gal-3 were more dramatic in EECs than ESCs. Moreover, estrogen (E2) and progesterone (P4) induced Gal-3 expression in EECs of healthy groups, and P4 was more significant than E2 and combined E2 and P4 (E2P4). However, in the endometriosis group, P4 failed to induce a similar increase in Gal-3 expression. Our results suggest that aberrant expression of Gal-3 might contribute to infertility in patients with endometriosis due to progesterone resistance.

  7. Aberrant expression of Sonic hedgehog signaling in Peutz-Jeghers syndrome.

    PubMed

    Xu, Xiaoping; Su, Juan; Li, Ran; Wang, Yadong; Zeng, Di; Wu, Baoping

    2016-04-01

    The SHH signaling pathway is critical for gastrointestinal development and organic patterning, and dysregulation of SHH pathway molecules has been detected in multiple gastrointestinal neoplasms. This study investigated the role of the SHH signaling pathway in PJS. Expression of SHH, PTCH, and GLI1 was examined by real-time PCR and immunohistochemistry in 20 normal tissues and 75 colorectal lesions (25 PJPs, 25 adenomas, and 25 adenocarcinomas). Expression of SHH, PTCH, and GLI1 mRNA was higher in PJPs than in normal tissue (P < .05) and gradually increased along the PJP-adenoma-adenocarcinoma sequence (P < .05). Immunostaining indicated that SHH expression was present in 60% of PJPs, 72% of adenomas, and 84% of carcinomas, whereas 68% of PJPs, 72% of adenomas, and 88% of carcinomas exhibited cytoplasmic expression of PTCH. Moreover, high GLI1 expression was detected in 56% of PJPs, 64% of adenomas, and 80% of carcinomas; and high nuclear expression of GLI1 was observed in 8 adenomas with atypia and 15 carcinomas. Increased SHH, PTCH, and GLI1 protein correlated positively with tumor grade (P = .012, P = .003, and P = .007, respectively), tumor depth (P = .024, P = .007, and P = .01), and lymph node metastasis (P = .05, P = .015, and P = .005). This study identified aberrant expression of SHH pathway molecules in PJS, and the findings may supply a novel mechanism for the development of PJ polyps.

  8. HOXA11 gene is hypermethylation and aberrant expression in gastric cancer

    PubMed Central

    2014-01-01

    Background Aberrant DNA methylation is an acquired epigenetic alteration that serves as an alternative to genetic defects in the inactivation of tumor suppressor genes and other genes in diverse human cancers. Gastric carcinoma is one of the tumors with a high frequency of aberrant methylation in promoter region. Hence we investigated the promoter methylation status and expression level of HOXA11 gene which may involve in GC development. Methods Thirty-two surgical excised gastric cancer specimens, twelve paired adjacent non-cancerous specimens and seven normal gastric mucosas were examined. The methylation status and expression level of HOXA11 gene were determined by bisulfite sequencing polymerase chain reaction (BSP), real-time polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC) respectively. HOXA11 expression was knocked-down with siRNA to mimic HOXA11 gene hypermethylation and ability of cell proliferation and migration was determinate. In addition, we analyzed and correlated the findings with clinicopathological features. Results The methylation level of HOXA11 gene in gastric cancer tissues and adjacent non-cancerous tissues were higher than those in normal gastric mucosa (P < 0.05). The methylation level was higher in TNM III and IV patients of GC than those in TNM I and II patients (P < 0.05). The expression of HOXA11 mRNA and protein decreased in normal gastric mucosa, peri-cancer tissue and GC (P < 0.05). HOXA11 expression was inversely correlated with DNA methylation (P < 0.05). Knocked-down of HOXA11 expression with siRNA in BGC-823 cells enhanced cell proliferation compared with control, but no significant different was observed in migration ability. Conclusion Hypermethylation and decreased expression of HOXA11 gene may be involved in the carcinogenesis and development of GC and may provide useful information for the prediction of the malignant behaviors of GC. And the expression of HOXA11 is impaired by DNA methylation. However

  9. Identification of aberrant gene expression associated with aberrant promoter methylation in primordial germ cells between E13 and E16 rat F3 generation vinclozolin lineage

    PubMed Central

    2015-01-01

    Background Transgenerational epigenetics (TGE) are currently considered important in disease, but the mechanisms involved are not yet fully understood. TGE abnormalities expected to cause disease are likely to be initiated during development and to be mediated by aberrant gene expression associated with aberrant promoter methylation that is heritable between generations. However, because methylation is removed and then re-established during development, it is not easy to identify promoter methylation abnormalities by comparing normal lineages with those expected to exhibit TGE abnormalities. Methods This study applied the recently proposed principal component analysis (PCA)-based unsupervised feature extraction to previously reported and publically available gene expression/promoter methylation profiles of rat primordial germ cells, between E13 and E16 of the F3 generation vinclozolin lineage that are expected to exhibit TGE abnormalities, to identify multiple genes that exhibited aberrant gene expression/promoter methylation during development. Results The biological feasibility of the identified genes were tested via enrichment analyses of various biological concepts including pathway analysis, gene ontology terms and protein-protein interactions. All validations suggested superiority of the proposed method over three conventional and popular supervised methods that employed t test, limma and significance analysis of microarrays, respectively. The identified genes were globally related to tumors, the prostate, kidney, testis and the immune system and were previously reported to be related to various diseases caused by TGE. Conclusions Among the genes reported by PCA-based unsupervised feature extraction, we propose that chemokine signaling pathways and leucine rich repeat proteins are key factors that initiate transgenerational epigenetic-mediated diseases, because multiple genes included in these two categories were identified in this study. PMID:26677731

  10. Aberrant Expression Profile of Long Noncoding RNA in Human Sinonasal Squamous Cell Carcinoma by Microarray Analysis

    PubMed Central

    Meng, Ling-zhao; Sun, Jing-wu; Yang, Fan

    2016-01-01

    Objectives. This study aimed to identify aberrantly expressed long noncoding RNAs (lncRNAs) profile of sinonasal squamous cell carcinoma (SSCC) and explore their potential functions. Methods. We investigated lncRNA and mRNA expression in SSCC and paired adjacent noncancerous tissues obtained from 6 patients with microarrays. Gene ontology (GO) analysis and pathway analysis were utilized to investigate the gene function. Gene signal-network and lncRNA-mRNA network were depicted. Quantitative real-time polymerase chain reaction (qRT-PCR) was utilized to validate 5 lncRNAs in a second set of paired SSCC and adjacent noncancerous tissues obtained from 22 additional patients. Results. We identified significantly differentially expressed lncRNAs (n = 3146) and mRNAs (n = 2208) in SSCC relative to noncancerous tissues. The GO annotation indicated that there are some core gene products that may be attributed to the progress of SSCC. The pathway analysis identified many pathways associated with cancer. The results of lncRNA-mRNA network and gene signal-network implied some core lncRNAs/mRNAs might play important roles in SSCC pathogenesis. The results of qRT-PCR showed that all of the 5 lncRNAs were differentially expressed and consistent with the microarray results. Conclusion. Our study is the first screening and analysis of lncRNAs expression profile in SSCC and may offer new insights into pathogenesis of this disease. PMID:28044124

  11. Aberrant expression of epithelial and neuroendocrine markers in alveolar rhabdomyosarcoma: a potentially serious diagnostic pitfall.

    PubMed

    Bahrami, Armita; Gown, Allen M; Baird, Geoffrey S; Hicks, M John; Folpe, Andrew L

    2008-07-01

    Alveolar rhabdomyosarcoma may be extremely difficult to distinguish from other primitive round cell neoplasms without ancillary immunohistochemistry and/or genetic study. Particularly in adults and in the head and neck locations, the differential diagnosis of alveolar rhabdomyosarcoma includes small cell carcinoma and neuroepithelial tumors, such as esthesioneuroblastoma. We have recently seen cases of genetically confirmed alveolar rhabdomyosarcoma, which were misdiagnosed owing to expression of cytokeratins and neuroendocrine markers. We studied a large group of well-characterized alveolar rhabdomyosarcomas for expression of such markers. Forty-four alveolar rhabdomyosarcomas (18 genetically confirmed) were retrieved from our archives and immunostained for wide-spectrum cytokeratin (OSCAR), low molecular weight cytokeratin (Cam5.2), synaptophysin, chromogranin A, and CD56 using commercially available antibodies. Cases were scored as 'negative', 'rare' (<5% positive cells), '1+' (5-25%), '2+' (26-50%) and '3+' (>51%). The tumors occurred in 23 males and 21 females at a mean age of 18 years (range, <1-64 years), and involved many sites. Fifty percent of cases (22 of 44) expressed wide-spectrum cytokeratin, and scored almost equally as rare, 1+, and 2+, but rarely 3+. Cam5.2 was positive in 52% (14 of 27). Forty-three percent of cases (16 of 37) expressed at least one of the specific neuroendocrine markers, 32% (12 of 37) expressed synaptophysin, 22% (eight of 36) expressed chromogranin A, and 11% expressed both. Expression of synaptophysin and chromogranin A was typically confined to rare cells but could be more widespread. Thirty-two percent of cases (12 of 37) expressed the wide-spectrum cytokeratin and at least one of the neuroendocrine markers, and 8% (three of 36) expressed cytokeratin and both neuroendocrine markers. CD56 expression was nearly ubiquitous. Aberrant expression of epithelial and neuroendocrine markers is relatively common in alveolar

  12. Aberrant expression of posterior HOX genes in well differentiated histotypes of thyroid cancers.

    PubMed

    Cantile, Monica; Scognamiglio, Giosuè; La Sala, Lucia; La Mantia, Elvira; Scaramuzza, Veronica; Valentino, Elena; Tatangelo, Fabiana; Losito, Simona; Pezzullo, Luciano; Chiofalo, Maria Grazia; Fulciniti, Franco; Franco, Renato; Botti, Gerardo

    2013-11-01

    Molecular etiology of thyroid cancers has been widely studied, and several molecular alterations have been identified mainly associated with follicular and papillary histotypes. However, the molecular bases of the complex pathogenesis of thyroid carcinomas remain poorly understood. HOX genes regulate normal embryonic development, cell differentiation and other critical processes in eukaryotic cell life. Several studies have shown that HOX genes play a role in neoplastic transformation of several human tissues. In particular, the genes belonging to HOX paralogous group 13 seem to hold a relevant role in both tumor development and progression. We have identified a significant prognostic role of HOX D13 in pancreatic cancer and we have recently showed the strong and progressive over-expression of HOX C13 in melanoma metastases and deregulation of HOX B13 expression in bladder cancers. In this study we have investigated, by immunohistochemisty and quantitative Real Time PCR, the HOX paralogous group 13 genes/proteins expression in thyroid cancer evolution and progression, also evaluating its ability to discriminate between main histotypes. Our results showed an aberrant expression, both at gene and protein level, of all members belonging to paralogous group 13 (HOX A13, HOX B13, HOX C13 and HOX D13) in adenoma, papillary and follicular thyroid cancers samples. The data suggest a potential role of HOX paralogous group 13 genes in pathogenesis and differential diagnosis of thyroid cancers.

  13. Aberrant Expression and Secretion of Heat Shock Protein 90 in Patients with Bullous Pemphigoid

    PubMed Central

    Tukaj, Stefan; Kleszczyński, Konrad; Vafia, Katerina; Groth, Stephanie; Meyersburg, Damian; Trzonkowski, Piotr; Ludwig, Ralf J.; Zillikens, Detlef; Schmidt, Enno; Fischer, Tobias W.; Kasperkiewicz, Michael

    2013-01-01

    The cell stress chaperone heat shock protein 90 (Hsp90) has been implicated in inflammatory responses and its inhibition has proven successful in different mouse models of autoimmune diseases, including epidermolysis bullosa acquisita. Here, we investigated expression levels and secretory responses of Hsp90 in patients with bullous pemphigoid (BP), the most common subepidermal autoimmune blistering skin disease. In comparison to healthy controls, the following observations were made: (i) Hsp90 was highly expressed in the skin of BP patients, whereas its serum levels were decreased and inversely associated with IgG autoantibody levels against the NC16A immunodominant region of the BP180 autoantigen, (ii) in contrast, neither aberrant levels of circulating Hsp90 nor any correlation of this protein with serum autoantibodies was found in a control cohort of autoimmune bullous disease patients with pemphigus vulgaris, (iii) Hsp90 was highly expressed in and restrictedly released from peripheral blood mononuclear cells of BP patients, and (iv) Hsp90 was potently induced in and restrictedly secreted from human keratinocyte (HaCaT) cells by BP serum and isolated anti-BP180 NC16A IgG autoantibodies, respectively. Our results reveal an upregulated Hsp90 expression at the site of inflammation and an autoantibody-mediated dysregulation of the intracellular and extracellular distribution of this chaperone in BP patients. These findings suggest that Hsp90 may play a pathophysiological role and represent a novel potential treatment target in BP. PMID:23936217

  14. Aberrant expression of interleukin-7 (IL-7) and its signalling complex in human breast cancer.

    PubMed

    Al-Rawi, M A A; Rmali, K; Watkins, G; Mansel, R E; Jiang, W G

    2004-03-01

    Interleukin-7 (IL-7), a haematopoietic growth factor, is known to induce the differentiation and proliferation of some haematological malignancies including certain types of leukaemias and lymphomas. However, little is known about its role in solid tumours, including breast cancer. In this study, the expression level of IL-7, IL-7 receptor (IL-7R) and their downstream signalling molecules, including the Janus kinases (Jak-1 and Jak-3), phosphoinositide 3-kinase (PI3-K) and signal transducers and activators of transcription (Stat-5) were analysed using the reverse transcriptase-polymerase chain reaction (RT-PCR), real-time quantitative PCR and immunohistochemistry in a cohort of patients with breast cancer. The results were analysed in relation to tumour grade, TNM stage, patients' prognosis (using the Nottingham Prognostic Index (NPI)) and survival. The levels of expression of IL-7, IL-7R, Jak-1, Jak-3, PI3-K and Stat-5 were significantly higher in the most aggressive tumours. With the exception of Stat-5 expression, the transcript copies of IL-7 and all other signalling molecules were higher in patients with the worst prognoses (NPI3) and in patients who died from breast cancer after 72 months of follow-up. This aberrant expression of IL-7 and its signalling intermediates in invasive breast cancers could have significant diagnostic and prognostic implications. Measuring these molecules in breast cancer tissues may provide, for the first time, important molecular indicators of tumour differentiation, aggressiveness, nodal status, prognosis and patient survival.

  15. Analysis of genomic aberrations and gene expression profiling identifies novel lesions and pathways in myeloproliferative neoplasms

    PubMed Central

    Rice, K L; Lin, X; Wolniak, K; Ebert, B L; Berkofsky-Fessler, W; Buzzai, M; Sun, Y; Xi, C; Elkin, P; Levine, R; Golub, T; Gilliland, D G; Crispino, J D; Licht, J D; Zhang, W

    2011-01-01

    Polycythemia vera (PV), essential thrombocythemia and primary myelofibrosis, are myeloproliferative neoplasms (MPNs) with distinct clinical features and are associated with the JAK2V617F mutation. To identify genomic anomalies involved in the pathogenesis of these disorders, we profiled 87 MPN patients using Affymetrix 250K single-nucleotide polymorphism (SNP) arrays. Aberrations affecting chr9 were the most frequently observed and included 9pLOH (n=16), trisomy 9 (n=6) and amplifications of 9p13.3–23.3 (n=1), 9q33.1–34.13 (n=1) and 9q34.13 (n=6). Patients with trisomy 9 were associated with elevated JAK2V617F mutant allele burden, suggesting that gain of chr9 represents an alternative mechanism for increasing JAK2V617F dosage. Gene expression profiling of patients with and without chr9 abnormalities (+9, 9pLOH), identified genes potentially involved in disease pathogenesis including JAK2, STAT5B and MAPK14. We also observed recurrent gains of 1p36.31–36.33 (n=6), 17q21.2–q21.31 (n=5) and 17q25.1–25.3 (n=5) and deletions affecting 18p11.31–11.32 (n=8). Combined SNP and gene expression analysis identified aberrations affecting components of a non-canonical PRC2 complex (EZH1, SUZ12 and JARID2) and genes comprising a ‘HSC signature' (MLLT3, SMARCA2 and PBX1). We show that NFIB, which is amplified in 7/87 MPN patients and upregulated in PV CD34+ cells, protects cells from apoptosis induced by cytokine withdrawal. PMID:22829077

  16. Immunohistochemical expression of aberrant Notch-1 signaling in vitiligo: an implication for pathogenesis.

    PubMed

    Seleit, Iman; Bakry, Ola Ahmed; Abdou, Asmaa Gaber; Dawoud, Noha Mohammed

    2014-06-01

    The etiopathogenetic mechanisms leading to pigment loss in vitiligo are not fully understood. Notch signaling is required for development and maintenance of melanocyte lineage and acts as a key component among keratinocyte-melanocyte interactions. The current study aimed to investigate the possible role of Notch signaling and its effect on the whole melanocyte lineage in vitiligo and correlating it with the different clinicopathologic parameters. Using immunohistochemical technique, Notch-1 expression was evaluated in 50 lesional and 20 perilesional biopsies of patients with vitiligo in comparison with 20 normal skin biopsies as a control group. Lesional biopsies were stained with human melanoma black-45 and tyrosinase-related protein-2 to demonstrate the melanocyte lineage. Membranous and/or nuclear expression of Notch-1 was in favor of control and perilesional skin, whereas cytoplasmic expression appeared only in vitiliginous lesions (P < .05). Membranous and/or nuclear expression of Notch-1 was significantly associated with epidermal human melanoma black-45 positivity (P = .01) and percentage of expression in both epidermis (P = .02) and hair follicles (P = .03) of lesional skin. Cytoplasmic pattern of Notch-1 expression in epidermis was significantly found in lesions with white hair (P = .04) and in cases with marked keratinocyte vacuolization (P = .03). Segmental and acrofacial vitiligo were associated with mild to moderate Notch-1 intensity, whereas generalized vitiligo was associated with strong intensity of expression (P = .02). In conclusion, Notch-1 signaling is inactivated in vitiligo with consequent loss of epidermal and/or follicular active melanocytes. Aberrant Notch signaling in vitiliginous white hair and acral and segmental vitiligo may be the cause of their treatment resistance.

  17. Aberrant microRNA expression in patients with painful peripheral neuropathies.

    PubMed

    Leinders, Mathias; Üçeyler, Nurcan; Thomann, Anna; Sommer, Claudia

    2017-09-15

    Changes in the neuro-immune balance play a major role in the induction and maintenance of neuropathic pain. We recently reported pathophysiologically relevant alterations in skin and sural nerve cytokine expression in peripheral neuropathies of different etiologies. Immune processes and cytokine expression are under tight control of microRNAs (miRNAs). To identify potential master switches in the neuro-immune balance, we aimed at characterizing inflammation-regulating miRNA profiles in patients with peripheral neuropathies. In an unselected patient cohort with polyneuropathies of different etiologies seen at our neuromuscular center between 2014 and 2015, we determined the systemic and local relative expression of miR-21-5p, miR-146a, and miR-155. In white blood cells we found higher miR-21 (p<0.001) and miR-146a (p<0.001) expression and lower miR-155 (p<0.001) expression when compared to healthy controls. In sural nerve, miR-21 (p<0.02) was increased in painful compared to painless neuropathies. In painful neuropathies, skin biopsies from the lower leg had reduced miR-146a (p<0.001) and miR-155 (p<0.001) expression compared to the thigh. Thus, peripheral neuropathies are associated with aberrant miRNA expression in white blood cells, sural nerve, and skin. These miRNA patterns may help to identify factors that determine the painfulness of peripheral neuropathies and lead to druggable targets. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Chemokine receptor co-expression reveals aberrantly distributed TH effector memory cells in GPA patients.

    PubMed

    Lintermans, Lucas L; Rutgers, Abraham; Stegeman, Coen A; Heeringa, Peter; Abdulahad, Wayel H

    2017-06-14

    Persistent expansion of circulating CD4(+) effector memory T cells (TEM) in patients with granulomatosis with polyangiitis (GPA) suggests their fundamental role in disease pathogenesis. Recent studies have shown that distinct functional CD4(+) TEM cell subsets can be identified based on expression patterns of chemokine receptors. The current study aimed to determine different CD4(+) TEM cell subsets based on chemokine receptor expression in peripheral blood of GPA patients. Identification of particular circulating CD4(+) TEM cells subsets may reveal distinct contributions of specific CD4(+) TEM subsets to the disease pathogenesis in GPA. Peripheral blood of 63 GPA patients in remission and 42 age- and sex-matched healthy controls was stained immediately after blood withdrawal with fluorochrome-conjugated antibodies for cell surface markers (CD3, CD4, CD45RO) and chemokine receptors (CCR4, CCR6, CCR7, CRTh2, CXCR3) followed by flow cytometry analysis. CD4(+) TEM memory cells (CD3(+)CD4(+)CD45RO(+)CCR7(-)) were gated, and the expression patterns of chemokine receptors CXCR3(+)CCR4(-)CCR6(-)CRTh2(-), CXCR3(-)CCR4(+)CCR6(-)CRTh2(+), CXCR3(-)CCR4(+)CCR6(+)CRTh2(-), and CXCR3(+)CCR4(-)CCR6(+)CRTh2(-) were used to distinguish TEM1, TEM2, TEM17, and TEM17.1 cells, respectively. The percentage of CD4(+) TEM cells was significantly increased in GPA patients in remission compared to HCs. Chemokine receptor co-expression analysis within the CD4(+) TEM cell population demonstrated a significant increase in the proportion of TEM17 cells with a concomitant significant decrease in the TEM1 cells in GPA patients compared to HC. The percentage of TEM17 cells correlated negatively with TEM1 cells in GPA patients. Moreover, the circulating proportion of TEM17 cells showed a positive correlation with the number of organs involved and an association with the tendency to relapse in GPA patients. Interestingly, the aberrant distribution of TEM1 and TEM17 cells is modulated in CMV

  19. Aberrant neuronal activity-induced signaling and gene expression in a mouse model of RASopathy

    PubMed Central

    Nakhaei-Rad, Saeideh; Montenegro-Venegas, Carolina; Pina-Fernández, Eneko; Marini, Claudia; Santos, Monica; Ahmadian, Mohammad R.; Stork, Oliver; Zenker, Martin

    2017-01-01

    Noonan syndrome (NS) is characterized by reduced growth, craniofacial abnormalities, congenital heart defects, and variable cognitive deficits. NS belongs to the RASopathies, genetic conditions linked to mutations in components and regulators of the Ras signaling pathway. Approximately 50% of NS cases are caused by mutations in PTPN11. However, the molecular mechanisms underlying cognitive impairments in NS patients are still poorly understood. Here, we report the generation and characterization of a new conditional mouse strain that expresses the overactive Ptpn11D61Y allele only in the forebrain. Unlike mice with a global expression of this mutation, this strain is viable and without severe systemic phenotype, but shows lower exploratory activity and reduced memory specificity, which is in line with a causal role of disturbed neuronal Ptpn11 signaling in the development of NS-linked cognitive deficits. To explore the underlying mechanisms we investigated the neuronal activity-regulated Ras signaling in brains and neuronal cultures derived from this model. We observed an altered surface expression and trafficking of synaptic glutamate receptors, which are crucial for hippocampal neuronal plasticity. Furthermore, we show that the neuronal activity-induced ERK signaling, as well as the consecutive regulation of gene expression are strongly perturbed. Microarray-based hippocampal gene expression profiling revealed profound differences in the basal state and upon stimulation of neuronal activity. The neuronal activity-dependent gene regulation was strongly attenuated in Ptpn11D61Y neurons. In silico analysis of functional networks revealed changes in the cellular signaling beyond the dysregulation of Ras/MAPK signaling that is nearly exclusively discussed in the context of NS at present. Importantly, changes in PI3K/AKT/mTOR and JAK/STAT signaling were experimentally confirmed. In summary, this study uncovers aberrant neuronal activity-induced signaling and regulation

  20. Aberrant β-catenin expression in urothelial carcinomas in blackfoot disease-endemic areas.

    PubMed

    Chen, Yi-Ting; Wu, Chun-Chieh; Liu, Xuan-Ping; Chai, Chee-Yin

    2017-01-01

    Arsenic is a well-known toxic element and carcinogenic agent. The aim of this study was to investigate p63, E-cadherin, and β-catenin proteins in urothelial carcinoma (UC) in both arsenic contaminated areas [so-called blackfoot disease (BFD) area] and non-BFD areas. The expressions of p63, E-cadherin, and β-catenin proteins in 20 UC cases of blackfoot disease and 22 UC cases in non-BFD areas were detected using immunohistochemical methods. The results revealed a high p63 expression in 20 (47.6%) UC cases and high E-cadherin expression in six (14.3%) UC cases. Expressions of p63 and E-cadherin showed no significant correlations with clinicopathologic parameters. However, all 20 BFD cases and 12 of 22 (54.5%) non-BFD cases showed aberrant β-catenin expression. Ten out of 22 (45.5%) non-BFD cases also had normal membranous immunoreactivity. The β-catenin staining pattern significantly differed between cases in endemic and nonendemic areas of BFD (p=0.001). Tumor sites also significantly correlated with β-catenin expression (p=0.044). In addition, membranous localization of β-catenin was lower in UC from BFD-endemic areas compared with those from non-BFD endemic areas. In conclusion, it is suggested that relocalization of β-catenin from membrane to cytoplasm may be involved in the tumorigenesis of UC from BFD-endemic areas. Copyright © 2016. Published by Elsevier Taiwan.

  1. Aberrant RNA splicing in cancer; expression changes and driver mutations of splicing factor genes.

    PubMed

    Sveen, A; Kilpinen, S; Ruusulehto, A; Lothe, R A; Skotheim, R I

    2016-05-12

    Alternative splicing is a widespread process contributing to structural transcript variation and proteome diversity. In cancer, the splicing process is commonly disrupted, resulting in both functional and non-functional end-products. Cancer-specific splicing events are known to contribute to disease progression; however, the dysregulated splicing patterns found on a genome-wide scale have until recently been less well-studied. In this review, we provide an overview of aberrant RNA splicing and its regulation in cancer. We then focus on the executors of the splicing process. Based on a comprehensive catalog of splicing factor encoding genes and analyses of available gene expression and somatic mutation data, we identify cancer-associated patterns of dysregulation. Splicing factor genes are shown to be significantly differentially expressed between cancer and corresponding normal samples, and to have reduced inter-individual expression variation in cancer. Furthermore, we identify enrichment of predicted cancer-critical genes among the splicing factors. In addition to previously described oncogenic splicing factor genes, we propose 24 novel cancer-critical splicing factors predicted from somatic mutations.

  2. Aberrant Expression of Histone Deacetylases 4 in Cognitive Disorders: Molecular Mechanisms and a Potential Target

    PubMed Central

    Wu, Yili; Hou, Fei; Wang, Xin; Kong, Qingsheng; Han, Xiaolin; Bai, Bo

    2016-01-01

    Histone acetylation is a major mechanism of chromatin remodeling, contributing to epigenetic regulation of gene transcription. Histone deacetylases (HDACs) are involved in both physiological and pathological conditions by regulating the status of histone acetylation. Although histone deacetylase 4 (HDAC4), a member of the HDAC family, may lack HDAC activity, it is actively involved in regulating the transcription of genes involved in synaptic plasticity, neuronal survival, and neurodevelopment by interacting with transcription factors, signal transduction molecules and HDAC3, another member of the HDAC family. HDAC4 is highly expressed in brain and its homeostasis is crucial for the maintenance of cognitive function. Accumulated evidence shows that HDAC4 expression is dysregulated in several brain disorders, including neurodegenerative diseases and mental disorders. Moreover, cognitive impairment is a characteristic feature of these diseases. It indicates that aberrant HDAC4 expression plays a pivotal role in cognitive impairment of these disorders. This review aims to describe the current understanding of HDAC4’s role in the maintenance of cognitive function and its dysregulation in neurodegenerative diseases and mental disorders, discuss underlying molecular mechanisms, and provide an outlook into targeting HDAC4 as a potential therapeutic approach to rescue cognitive impairment in these diseases. PMID:27847464

  3. Aberrantly Expressed Long Non-Coding RNAs In CD8(+) T Cells Response to Active Tuberculosis.

    PubMed

    Fu, Yurong; Gao, Kunshan; Tao, Enxue; Li, Ruifang; Yi, Zhengjun

    2017-04-19

    Dysregulated expression of long noncoding RNAs (lncRNAs) has been demonstrated as being implicated in a variety of human diseases. In the study we aimed to determine lncRNA profile in CD8(+) T cells response to active tuberculosis (TB). We examined the lncRNA expression by microarray in circulating CD8(+) T cells isolated from patients with active TB and healthy controls. Change predictions to analysis was used to address functional roles of the deregulated mRNAs. Real-time quantitative PCR (RT-qPCR) was used to validate the microarray result. In total, 328 lncRNAs and 356 mRNAs were differentially expressed in TB CD8(+) T cells. Upregulated mRNAs were mainly enriched in cAMP signaling pathway, calcium signaling pathway, and TGF-beta signaling pathway, while downregulated mRNAs were enriched in antigen processing and presentation and natural killer cell mediated cytotoxicity in TB CD8(+) T cells. Interestingly, we found that heme oxygenase 1 (HMOX1) was decreased in active TB CD8(+) T cells, while its nearby lincRNA XLOC_014219 was upregulated. Subsequent RT-qPCR results confirmed the changes. This is the first research addressing lncRNA expression profiles in active TB CD8(+) T cells. The aberrantly expressed lncRNAs observed in the study may provide clues to the dysfunction of CD8(+) T cells and so to the pathophysiological properties of active TB. Further studies should focus on the function of lncRNAs involved in active TB. J. Cell. Biochem. 9999: 1-10, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  4. Association of arsenic-induced malignant transformation with DNA hypomethylation and aberrant gene expression

    PubMed Central

    Zhao, Christopher Q.; Young, Matthew R.; Diwan, Bhalchandra A.; Coogan, Timothy P.; Waalkes, Michael P.

    1997-01-01

    Inorganic arsenic, a human carcinogen, is enzymatically methylated for detoxication, consuming S-adenosyl-methionine (SAM) in the process. The fact that DNA methyltransferases (MeTases) require this same methyl donor suggests a role for methylation in arsenic carcinogenesis. Here we test the hypothesis that arsenic-induced initiation results from DNA hypomethylation caused by continuous methyl depletion. The hypothesis was tested by first inducing transformation in a rat liver epithelial cell line by chronic exposure to low levels of arsenic, as confirmed by the development of highly aggressive, malignant tumors after inoculation of cells into Nude mice. Global DNA hypomethylation occurred concurrently with malignant transformation and in the presence of depressed levels of S-adenosyl-methionine. Arsenic-induced DNA hypomethylation was a function of dose and exposure duration, and remained constant even after withdrawal of arsenic. Hyperexpressibility of the MT gene, a gene for which expression is clearly controlled by DNA methylation, was also detected in transformed cells. Acute arsenic or arsenic at nontransforming levels did not induce global hypomethylation of DNA. Whereas transcription of DNA MeTase was elevated, the MeTase enzymatic activity was reduced with arsenic transformation. Taken together, these results indicate arsenic can act as a carcinogen by inducing DNA hypomethylation, which in turn facilitates aberrant gene expression, and they constitute a tenable theory of mechanism in arsenic carcinogenesis. PMID:9380733

  5. Aberrant expression of cell cycle and material metabolism related genes contributes to hepatocellular carcinoma occurrence.

    PubMed

    Yan, Hongxian; Li, Zhaohui; Shen, Quan; Wang, Qian; Tian, Jianguo; Jiang, Qingfeng; Gao, Linbo

    2017-04-01

    This study aims to deepen our understanding of the molecular mechanism underlying the occurrence of hepatocellular carcinoma (HCC). We first downloaded a gene expression profile dataset GSE29721 (10 HCC and 10 control samples) from Gene Expression Omnibus database (http://www.ncbi.nlm.nih.gov/geo/). Differentially expressed genes (DEGs) were identified by the paired t-test using limma package. Pathway and functional enrichment analyses were performed with DAVID tools. Transcription factors were annotated with TRANSFAC database and tumor associated genes (TAGs) were annotated with TAG and TSGene databases. Protein-protein interaction (PPI) network was conducted using STRING online tool and function module was further identified with BioNet package. Totally, 527 up-regulated DEGs and 587 down-regulated DEGs were identified. GO functional and KEGG pathway enrichment analyses showed that the up-regulated DEGs were mainly related to cell division and cell cycle, while the down-regulated DEGs were largely related to material metabolism, especially secondary metabolism. Proteins encoded by DEGs CDK1, BUB1, CDC20, NCAPG, NDC80, CDCA8, MAD2L1, CCNB1, CCNA2 and BIRC5 were hub genes with high degrees in the PPI network; further module analysis detected a subnetwork consisting of 55 proteins, such as CYP2B6, ACAA1, BHMT and ALDH2. Taken together, aberrant expression of cell cycle related genes (e.g., CDK1, CCNA2, CCNB1, BUB1, MAD2L1 and CDC20) and material metabolism related genes (e.g., CYP2B6, ACAA1, BHMT and ALDH2) may contribute to HCC occurrence. Copyright © 2017 Elsevier GmbH. All rights reserved.

  6. The aberrant expression and localization of DNA methyltransferase 3B in endometriotic stromal cells

    PubMed Central

    Dyson, Matthew T.; Kakinuma, Toshiyuki; Pavone, Mary Ellen; Monsivais, Diana; Navarro, Antonia; Malpani, Saurabh S.; Ono, Masanori; Bulun, Serdar E.

    2015-01-01

    Objective To define the expression and function of DNA methyltransferases (DNMTs) in response to decidualizing stimuli in endometriotic cells compared with healthy endometrial stroma. Design Basic science. Setting University research center. Patients Premenopausal women with or without endometriosis. Interventions Primary cultures of stromal cells from healthy endometrium (E-IUM) or endometriomas (E-OSIS) were subjected to in vitro decidualization (IVD) using 1 µM medroxyprogesterone acetate, 35 nM 17β-estradiol, and 0.05 mM 8-Br-cAMP. Main Outcome Measure(s) DNMT1, DNMT3A, and DNMT3B expression in E-IUM and E-OSIS were assessed by qRT-PCR and immunoblotting. DNMT3B recruitment to the promoters of steroidogenic factor 1 (SF-1) and estrogen receptor α (ESR1) was examined by chromatin immunoprecipitation Results IVD treatment reduced DNMT3B mRNA (74%) and protein levels (81%) only in E-IUM. DNMT1 and DNMT3A were unchanged in both cell types. Significantly more DNMT3B bound to the SF-1 promoter in E-IUM compared with E-OSIS, and IVD treatment reduced binding in E-IUM to levels similar to those in E-OSIS. DNMT3B enrichment across three ESR1 promoters was reduced in E-IUM after IVD, although the more distal promoter showed increased DNMT3B enrichment in E-OSIS after IVD. Conclusions The inability to downregulate DNMT3B expression in E-OSIS may contribute to an aberrant epigenetic fingerprint that misdirects gene expression in endometriosis and contributes to its altered response to steroid hormones. PMID:26239024

  7. Aberrant expression and localization of deoxyribonucleic acid methyltransferase 3B in endometriotic stromal cells.

    PubMed

    Dyson, Matthew T; Kakinuma, Toshiyuki; Pavone, Mary Ellen; Monsivais, Diana; Navarro, Antonia; Malpani, Saurabh S; Ono, Masanori; Bulun, Serdar E

    2015-10-01

    To define the expression and function of DNA methyltransferases (DNMTs) in response to decidualizing stimuli in endometriotic cells compared with healthy endometrial stroma. Basic science. University research center. Premenopausal women with or without endometriosis. Primary cultures of stromal cells from healthy endometrium (E-IUM) or endometriomas (E-OSIS) were subjected to in vitro decidualization (IVD) using 1 μM medroxyprogesterone acetate, 35 nM 17β-estradiol, and 0.05 mM 8-Br-cAMP. Expression of DNMT1, DNMT3A, and DNMT3B in E-IUM and E-OSIS were assessed by quantitative real-time polymerase chain reaction and immunoblotting. Recruitment of DNMT3B to the promoters of steroidogenic factor 1 (SF-1) and estrogen receptor α (ESR1) was examined by chromatin immunoprecipitation. IVD treatment reduced DNMT3B messenger RNA (74%) and protein levels (81%) only in E-IUM; DNMT1 and DNMT3A were unchanged in both cell types. Significantly more DNMT3B bound to the SF-1 promoter in E-IUM compared with E-OSIS, and IVD treatment reduced binding in E-IUM to levels similar to those in E-OSIS. Enrichment of DNMT3B across 3 ESR1 promoters was reduced in E-IUM after IVD, although the more-distal promoter showed increased DNMT3B enrichment in E-OSIS after IVD. The inability to downregulate DNMT3B expression in E-OSIS may contribute to an aberrant epigenetic fingerprint that misdirects gene expression in endometriosis and contributes to its altered response to steroid hormones. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  8. RDX induces aberrant expression of microRNAs in mouse brain and liver.

    PubMed

    Zhang, Baohong; Pan, Xiaoping

    2009-02-01

    Although microRNAs (miRNAs) have been found to play an important role in many biological and metabolic processes, their functions in animal response to environmental toxicant exposure are largely unknown. We used hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), a common environmental contaminant, as a toxicant stressor to investigate toxicant-induced changes in miRNA expression in B6C3F1 mice and the potential mechanism of RDX-induced toxic action. B6C3F1 mice were fed diets with or without 5 mg/kg RDX for 28 days. After the feeding trials, we isolated RNAs from both brain and liver tissues and analyzed the expression profiles of 567 known mouse miRNAs using microarray and quantitative real-time polymerase chain reaction technologies. RDX exposure induced significant changes in miRNA expression profiles. A total of 113 miRNAs, belonging to 75 families, showed significantly altered expression patterns after RDX exposure. Of the 113 miRNAs, 10 were significantly up-regulated and 3 were significantly down-regulated (p < 0.01) in both mouse brain and liver. Many miRNAs had tissue-specific responses to RDX exposure. Specifically, expression of seven miRNAs was up-regulated in the brain but down-regulated in the liver or up-regulated in the liver but down-regulated in the brain (p < 0.01). Many aberrantly expressed miRNAs were related to various cancers, toxicant-metabolizing enzymes, and neurotoxicity. We found a significant up-regulation of oncogenic miRNAs and a significant down-regulation of tumor-suppressing miRNAs, which included let-7, miR-17-92, miR-10b, miR-15, miR-16, miR-26, and miR-181. Environmental toxicant exposure alters the expression of a suite of miRNAs.

  9. RDX Induces Aberrant Expression of MicroRNAs in Mouse Brain and Liver

    PubMed Central

    Zhang, Baohong; Pan, Xiaoping

    2009-01-01

    Background Although microRNAs (miRNAs) have been found to play an important role in many biological and metabolic processes, their functions in animal response to environmental toxicant exposure are largely unknown. Objectives We used hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), a common environmental contaminant, as a toxicant stressor to investigate toxicant-induced changes in miRNA expression in B6C3F1 mice and the potential mechanism of RDX-induced toxic action. Methods B6C3F1 mice were fed diets with or without 5 mg/kg RDX for 28 days. After the feeding trials, we isolated RNAs from both brain and liver tissues and analyzed the expression profiles of 567 known mouse miRNAs using microarray and quantitative real-time polymerase chain reaction technologies. Results RDX exposure induced significant changes in miRNA expression profiles. A total of 113 miRNAs, belonging to 75 families, showed significantly altered expression patterns after RDX exposure. Of the 113 miRNAs, 10 were significantly up-regulated and 3 were significantly down-regulated (p < 0.01) in both mouse brain and liver. Many miRNAs had tissue-specific responses to RDX exposure. Specifically, expression of seven miRNAs was up-regulated in the brain but down-regulated in the liver or up-regulated in the liver but down-regulated in the brain (p < 0.01). Many aberrantly expressed miRNAs were related to various cancers, toxicant-metabolizing enzymes, and neurotoxicity. We found a significant up-regulation of oncogenic miRNAs and a significant down-regulation of tumor-suppressing miRNAs, which included let-7, miR-17-92, miR-10b, miR-15, miR-16, miR-26, and miR-181. Conclusions Environmental toxicant exposure alters the expression of a suite of miRNAs. PMID:19270793

  10. Aberrant expression of microRNAs in gastric cancer and biological significance of miR-574-3p.

    PubMed

    Su, Yingying; Ni, Zhaohui; Wang, Guoqing; Cui, Juan; Wei, Chengguo; Wang, Jihan; Yang, Qing; Xu, Ying; Li, Fan

    2012-08-01

    The discovery of microRNAs (miRNAs) provides a new and powerful tool for studying the mechanisms, diagnosis and treatments of cancer. In this study, we employed AFFX miRNA expression chips to search for miRNAs that may be aberrantly expressed in gastric cancer tissues and to investigate the potential roles that miRNAs may play in the development and progression of gastric cancer. 14 miRNAs were found to be down-regulated and 2 miRNAs up-regulated in gastric cancer tissues compared to the normal gastric tissues. Among the aberrantly expressed miRNAs, miR-574-3p was selected to further study its expression features and functional roles. Interestingly, the reduced expression of miR-574-3p occurred mainly in the early stages of gastric cancer or in cancers with high level of differentiation, suggesting that it can be used as a marker for a mild case of gastric cancer. Functional study revealed that cell proliferation, migration and invasion were significantly inhibited in miR-574-3p-transfected gastric cancer SGC7901 cells. Computational prediction and experimental validation suggest that Cullin2 may be one of the targets of miR-574-3p. Overall our study suggests that the aberrantly expressed miRNAs may play regulatory and functional roles in the development and progression of gastric cancer.

  11. Aberrant PD-L1 expression through 3'-UTR disruption in multiple cancers.

    PubMed

    Kataoka, Keisuke; Shiraishi, Yuichi; Takeda, Yohei; Sakata, Seiji; Matsumoto, Misako; Nagano, Seiji; Maeda, Takuya; Nagata, Yasunobu; Kitanaka, Akira; Mizuno, Seiya; Tanaka, Hiroko; Chiba, Kenichi; Ito, Satoshi; Watatani, Yosaku; Kakiuchi, Nobuyuki; Suzuki, Hiromichi; Yoshizato, Tetsuichi; Yoshida, Kenichi; Sanada, Masashi; Itonaga, Hidehiro; Imaizumi, Yoshitaka; Totoki, Yasushi; Munakata, Wataru; Nakamura, Hiromi; Hama, Natsuko; Shide, Kotaro; Kubuki, Yoko; Hidaka, Tomonori; Kameda, Takuro; Masuda, Kyoko; Minato, Nagahiro; Kashiwase, Koichi; Izutsu, Koji; Takaori-Kondo, Akifumi; Miyazaki, Yasushi; Takahashi, Satoru; Shibata, Tatsuhiro; Kawamoto, Hiroshi; Akatsuka, Yoshiki; Shimoda, Kazuya; Takeuchi, Kengo; Seya, Tsukasa; Miyano, Satoru; Ogawa, Seishi

    2016-06-16

    Successful treatment of many patients with advanced cancer using antibodies against programmed cell death 1 (PD-1; also known as PDCD1) and its ligand (PD-L1; also known as CD274) has highlighted the critical importance of PD-1/PD-L1-mediated immune escape in cancer development. However, the genetic basis for the immune escape has not been fully elucidated, with the exception of elevated PD-L1 expression by gene amplification and utilization of an ectopic promoter by translocation, as reported in Hodgkin and other B-cell lymphomas, as well as stomach adenocarcinoma. Here we show a unique genetic mechanism of immune escape caused by structural variations (SVs) commonly disrupting the 3' region of the PD-L1 gene. Widely affecting multiple common human cancer types, including adult T-cell leukaemia/lymphoma (27%), diffuse large B-cell lymphoma (8%), and stomach adenocarcinoma (2%), these SVs invariably lead to a marked elevation of aberrant PD-L1 transcripts that are stabilized by truncation of the 3'-untranslated region (UTR). Disruption of the Pd-l1 3'-UTR in mice enables immune evasion of EG7-OVA tumour cells with elevated Pd-l1 expression in vivo, which is effectively inhibited by Pd-1/Pd-l1 blockade, supporting the role of relevant SVs in clonal selection through immune evasion. Our findings not only unmask a novel regulatory mechanism of PD-L1 expression, but also suggest that PD-L1 3'-UTR disruption could serve as a genetic marker to identify cancers that actively evade anti-tumour immunity through PD-L1 overexpression.

  12. Breast cancer-associated high-order SNP-SNP interaction of CXCL12/CXCR4-related genes by an improved multifactor dimensionality reduction (MDR-ER).

    PubMed

    Fu, Ou-Yang; Chang, Hsueh-Wei; Lin, Yu-Da; Chuang, Li-Yeh; Hou, Ming-Feng; Yang, Cheng-Hong

    2016-09-01

    In association studies, the combined effects of single nucleotide polymorphism (SNP)-SNP interactions and the problem of imbalanced data between cases and controls are frequently ignored. In the present study, we used an improved multifactor dimensionality reduction (MDR) approach namely MDR-ER to detect the high order SNP‑SNP interaction in an imbalanced breast cancer data set containing seven SNPs of chemokine CXCL12/CXCR4 pathway genes. Most individual SNPs were not significantly associated with breast cancer. After MDR‑ER analysis, six significant SNP‑SNP interaction models with seven genes (highest cross‑validation consistency, 10; classification error rates, 41.3‑21.0; and prediction error rates, 47.4‑55.3) were identified. CD4 and VEGFA genes were associated in a 2‑loci interaction model (classification error rate, 41.3; prediction error rate, 47.5; odds ratio (OR), 2.069; 95% bootstrap CI, 1.40‑2.90; P=1.71E‑04) and it also appeared in all the best 2‑7‑loci models. When the loci number increased, the classification error rates and P‑values decreased. The powers in 2‑7‑loci in all models were >0.9. The minimum classification error rate of the MDR‑ER‑generated model was shown with the 7‑loci interaction model (classification error rate, 21.0; OR=15.282; 95% bootstrap CI, 9.54‑23.87; P=4.03E‑31). In the epistasis network analysis, the overall effect with breast cancer susceptibility was identified and the SNP order of impact on breast cancer was identified as follows: CD4 = VEGFA > KITLG > CXCL12 > CCR7 = MMP2 > CXCR4. In conclusion, the MDR‑ER can effectively and correctly identify the best SNP‑SNP interaction models in an imbalanced data set for breast cancer cases.

  13. Genome-wide gene expression profiling reveals aberrant MAPK and Wnt signaling pathways associated with early parthenogenesis.

    PubMed

    Liu, Na; Enkemann, Steven A; Liang, Ping; Hersmus, Remko; Zanazzi, Claudia; Huang, Junjiu; Wu, Chao; Chen, Zhisheng; Looijenga, Leendert H J; Keefe, David L; Liu, Lin

    2010-12-01

    Mammalian parthenogenesis could not survive but aborted during mid-gestation, presumably because of lack of paternal gene expression. To understand the molecular mechanisms underlying the failure of parthenogenesis at early stages of development, we performed global gene expression profiling and functional analysis of parthenogenetic blastocysts in comparison with those of blastocysts from normally fertilized embryos. Parthenogenetic blastocysts exhibited changes in the expression of 749 genes, of which 214 had lower expression and 535 showed higher expressions than fertilized embryos using a minimal 1.8-fold change as a cutoff. Genes important for placenta development were decreased in their expression in parthenote blastocysts. Some maternally expressed genes were up-regulated and paternal-related genes were down-regulated. Moreover, aberrantly increased Wnt signaling and reduced mitogen-activated protein kinase (MAPK) signaling were associated with early parthenogenesis. The protein level of extracellular signal-regulated kinase 2 (ERK2) was low in parthenogenetic blastocysts compared with that of fertilized blastocysts 120 h after fertilization. 6-Bromoindirubin-3'-oxime, a specific glycogen synthase kinase-3 (GSK-3) inhibitor, significantly decreased embryo hatching. The expression of several imprinted genes was altered in parthenote blastocysts. Gene expression also linked reduced expression of Xist to activation of X chromosome. Our findings suggest that failed X inactivation, aberrant imprinting, decreased ERK/MAPK signaling and possibly elevated Wnt signaling, and reduced expression of genes for placental development collectively may contribute to abnormal placenta formation and failed fetal development in parthenogenetic embryos.

  14. Expression of aberrant forms of AUXIN RESPONSE FACTOR8 stimulates parthenocarpy in Arabidopsis and tomato.

    PubMed

    Goetz, Marc; Hooper, Lauren C; Johnson, Susan D; Rodrigues, Julio Carlyle Macedo; Vivian-Smith, Adam; Koltunow, Anna M

    2007-10-01

    Fruit initiation in Arabidopsis (Arabidopsis thaliana) is generally repressed until fertilization occurs. However, mutations in AUXIN RESPONSE FACTOR8 (ARF8) uncouple fruit initiation from fertilization, resulting in the formation of seedless, parthenocarpic fruit. Here we induced parthenocarpy in wild-type Arabidopsis by introducing either the mutant genomic (g) Atarf8-4 sequence or gAtARF8:beta-glucuronidase translational fusion constructs by plant transformation. Silencing of endogenous AtARF8 transcription was not observed, indicating that the introduced, aberrant ARF8 transcripts were compromising the function of endogenous ARF8 and/or associated factors involved in suppressing fruit initiation. To analyze the role of ARF8 in tomato (Solanum lycopersicum) we initially emasculated 23 tomato cultivars to test for background parthenocarpy. Surprisingly, all had a predisposition to initiate fertilization-independent fruit growth. Expression of gAtarf8-4 in transgenic tomato ('Monalbo') resulted in a significant increase in the number and size of parthenocarpic fruit. Isolation of tomato ARF8 cDNA indicated significant sequence conservation with AtARF8. SlARF8 may therefore control tomato fruit initiation in a similar manner as AtARF8 does in Arabidopsis. Two SlARF8 cDNAs differing in size by 5 bp were found, both arising from the same gene. The smaller cDNA is a splice variant and is also present in Arabidopsis. We propose that low endogenous levels of the splice variant products might interfere with efficient formation/function of a complex repressing fruit initiation, thereby providing an explanation for the observed ovary expansion in tomato and also Arabidopsis after emasculation. Increasing the levels of aberrant Atarf8-4 transcripts may further destabilize formation/function of the complex in a dosage-dependent manner enhancing tomato parthenocarpic fruit initiation frequency and size and mimicking the parthenocarpic dehiscent silique phenotype found in

  15. PIGN gene expression aberration is associated with genomic instability and leukemic progression in acute myeloid leukemia with myelodysplastic features

    PubMed Central

    Teye, Emmanuel K.; Sido, Abigail; Xin, Ping; Finnberg, Niklas K.; Gokare, Prashanth; Kawasawa, Yuka I.; Salzberg, Anna C.; Shimko, Sara; Bayerl, Michael; Ehmann, W. Christopher; Claxton, David F.; Rybka, Witold B.; Drabick, Joseph J.; Wang, Hong-Gang; Abraham, Thomas; El-Deiry, Wafik S.; Brodsky, Robert A.; Hohl, Raymond J.; Pu, Jeffrey J.

    2017-01-01

    Previous studies have linked increased frequency of glycosylphosphatidylinositol-anchor protein (GPI-AP) deficiency with genomic instability and the risk of carcinogenesis. However, the underlying mechanism is still not clear. A randomForest analysis of the gene expression array data from 55 MDS patients (GSE4619) demonstrated a significant (p = 0.0007) correlation (Pearson r =-0.4068) between GPI-anchor biosynthesis gene expression and genomic instability, in which PIGN, a gene participating in GPI-AP biosynthesis, was ranked as the third most important in predicting risk of MDS progression. Furthermore, we observed that PIGN gene expression aberrations (increased transcriptional activity but diminished to no protein production) were associated with increased frequency of GPI-AP deficiency in leukemic cells during leukemic transformation/progression. PIGN gene expression aberrations were attributed to partial intron retentions between exons 14 and 15 resulting in frameshifts and premature termination which were confirmed by examining the RNA-seq data from a group of AML patients (phs001027.v1.p1). PIGN gene expression aberration correlated with the elevation of genomic instability marker expression that was independent of the TP53 regulatory pathway. Suppression/elimination of PIGN protein expression caused a similar pattern of genomic instability that was rescued by PIGN restoration. Finally, we found that PIGN bound to the spindle assembly checkpoint protein, MAD1, and regulated its expression during the cell cycle. In conclusion, PIGN gene is crucial in regulating mitotic integrity to maintain chromosomal stability and prevents leukemic transformation/progression. PMID:28187452

  16. Abnormal expression of mRNA, microRNA alteration and aberrant DNA methylation patterns in rectal adenocarcinoma

    PubMed Central

    Liu, Xianglong; Yuan, Xiangfei; Qin, Hai; Zhang, Xipeng

    2017-01-01

    Aim Rectal adenocarcinoma (READ) is a malignancy cancer with the high morbidity and motility worldwide. Our study aimed to explore the potential pathogenesis of READ through integrated analysis of gene expression profiling and DNA methylation data. Methods The miRNA, mRNA expression profiling and corresponding DNA methylation data were downloaded from The Cancer Genome Atlas (TCGA) database. Differentially expressed mRNAs/ miRNAs/methylated regions (DEmRNA/DEmiRNAs) were identified in READ. The negatively correlation of DEmiRNA-DEmRNAs and DNA methylation-DEmRNAs were obtained. DEmRNAs expression was validated through quantitative real-time polymerase chain reaction (qRT-PCR) and microarray expression profiling analyses. Results 1192 dysregulated DEmRNAs, 27 dysregulated DEmiRNAs and 6403 aberrant methylation CpG sites were screened in READ compared to normal controls. 1987 negative interaction pairs among 27 DEmiRNAs and 668 DEmRNAs were predicted. 446 genes with aberrant methylation were annotated. Eventually, 50 DEmRNAs (39 down- and 11 up-regulated DEmRNAs) with hypermethylation, synchronously negatively targeted by DEmiRNAs, were identified through the correlation analysis among 446 genes with aberrant methylation and 668 DEmRNAs. 50 DEmRNAs were significantly enriched in cAMP signaling pathway, circadian entrainment and glutamatergic synapse. The validation results of expression levels of DEmRNAs through qRT-PCR and microarray analyses were compatible with our study. Conclusion 7 genes of SORCS1, PDZRN4, LONRF2, CNGA3, HAND2, RSPO2 and GNAO1 with hypermethylation and negatively regulation by DEmiRNAs might contribute to the tumorigenesis of READ. Our work might provide valuable foundation for the READ in mechanism elucidation, early diagnosis and therapeutic target identification. PMID:28350845

  17. Aberrantly expressed LGR4 empowers Wnt signaling in multiple myeloma by hijacking osteoblast-derived R-spondins

    PubMed Central

    van Andel, Harmen; Ren, Zemin; Koopmans, Iris; Joosten, Sander P. J.; Kocemba, Kinga A.; de Lau, Wim; Kersten, Marie José; de Bruin, Alexander M.; Guikema, Jeroen E. J.; Clevers, Hans; Spaargaren, Marcel; Pals, Steven T.

    2017-01-01

    The unrestrained growth of tumor cells is generally attributed to mutations in essential growth control genes, but tumor cells are also affected by, or even addicted to, signals from the microenvironment. As therapeutic targets, these extrinsic signals may be equally significant as mutated oncogenes. In multiple myeloma (MM), a plasma cell malignancy, most tumors display hallmarks of active Wnt signaling but lack activating Wnt-pathway mutations, suggesting activation by autocrine Wnt ligands and/or paracrine Wnts emanating from the bone marrow (BM) niche. Here, we report a pivotal role for the R-spondin/leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) axis in driving aberrant Wnt/β-catenin signaling in MM. We show that LGR4 is expressed by MM plasma cells, but not by normal plasma cells or B cells. This aberrant LGR4 expression is driven by IL-6/STAT3 signaling and allows MM cells to hijack R-spondins produced by (pre)osteoblasts in the BM niche, resulting in Wnt (co)receptor stabilization and a dramatically increased sensitivity to auto- and paracrine Wnts. Our study identifies aberrant R-spondin/LGR4 signaling with consequent deregulation of Wnt (co)receptor turnover as a driver of oncogenic Wnt/β-catenin signaling in MM cells. These results advocate targeting of the LGR4/R-spondin interaction as a therapeutic strategy in MM. PMID:28028233

  18. Aberrantly expressed LGR4 empowers Wnt signaling in multiple myeloma by hijacking osteoblast-derived R-spondins.

    PubMed

    van Andel, Harmen; Ren, Zemin; Koopmans, Iris; Joosten, Sander P J; Kocemba, Kinga A; de Lau, Wim; Kersten, Marie José; de Bruin, Alexander M; Guikema, Jeroen E J; Clevers, Hans; Spaargaren, Marcel; Pals, Steven T

    2017-01-10

    The unrestrained growth of tumor cells is generally attributed to mutations in essential growth control genes, but tumor cells are also affected by, or even addicted to, signals from the microenvironment. As therapeutic targets, these extrinsic signals may be equally significant as mutated oncogenes. In multiple myeloma (MM), a plasma cell malignancy, most tumors display hallmarks of active Wnt signaling but lack activating Wnt-pathway mutations, suggesting activation by autocrine Wnt ligands and/or paracrine Wnts emanating from the bone marrow (BM) niche. Here, we report a pivotal role for the R-spondin/leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) axis in driving aberrant Wnt/β-catenin signaling in MM. We show that LGR4 is expressed by MM plasma cells, but not by normal plasma cells or B cells. This aberrant LGR4 expression is driven by IL-6/STAT3 signaling and allows MM cells to hijack R-spondins produced by (pre)osteoblasts in the BM niche, resulting in Wnt (co)receptor stabilization and a dramatically increased sensitivity to auto- and paracrine Wnts. Our study identifies aberrant R-spondin/LGR4 signaling with consequent deregulation of Wnt (co)receptor turnover as a driver of oncogenic Wnt/β-catenin signaling in MM cells. These results advocate targeting of the LGR4/R-spondin interaction as a therapeutic strategy in MM.

  19. Serum concentrations of chemokines (CCL-5 and CXCL-12), chemokine receptors (CCR-5 and CXCR-4), and IL-6 in patients with posttraumatic stress disorder and avoidant personality disorder.

    PubMed

    Ogłodek, Ewa A; Szota, Anna M; Moś, Danuta M; Araszkiewicz, Aleksander; Szromek, Adam R

    2015-12-01

    Posttraumatic stress disorder (PTSD) can be perceived as a psychoneuroimmunological disorder in which cytokines affecting the neurochemical and neuroendocrine functions of the body play an important role. Among cytokines, chemokines participating in activation of the inflammatory response are considered to be crucial. 220 men and women were enrolled in the study. 180 of them constituted the study group. The studied groups consisted of: 60 patients with a diagnosed avoidant personality disorders (APD), 60 patients with a diagnosed APD and with PTSD and of 60 patients with PTSD but without a APD. There were 30 women and 30 men in each group of 60 subjects. The control group consisted of 40 healthy individuals. The plasma levels of chemokines and their receptors (CCL-5, CXCR-5, CXCL-12 and CXCR-4), as well as IL-6, were assessed by ELISA. There was an increase in the CXCL-12 and CCL-5 levels in women and men with the PTSD versus the control group. Also, increased levels of IL-6 and the receptors CXCR-4, CCR-5 were observed in women and men with PTSD. The levels of CXCL-12 and CCL-5 chemokines, as well as CCR-5 and CXCR4 receptors were higher in women than in men. The results of this study indicate a need for assessment of the CCL-5 and CXCL-12 chemokine levels, as they are likely markers of PTSD. Measurement of the concentrations of chemokines, chemokine receptors and IL-6 in women and men with PTSD along with concomittant APD may be useful for early detection of mental disorders. Copyright © 2015 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  20. Enhancement of intracellular signaling associated with hematopoietic progenitor cell survival in response to SDF-1/CXCL12 in synergy with other cytokines.

    PubMed

    Lee, Younghee; Gotoh, Akihiko; Kwon, Hyung-Joo; You, Minute; Kohli, Lisa; Mantel, Charlie; Cooper, Scott; Hangoc, Giao; Miyazawa, Keisuke; Ohyashiki, Kazuma; Broxmeyer, Hal E

    2002-06-15

    Stromal cell-derived factor 1 (SDF-1/CXCL12) is a multifunctional cytokine. We previously reported that myelopoiesis was enhanced in SDF-1 alpha transgenic mice, probably due in part to SDF-1 alpha enhancement of myeloid progenitor cell (MPC) survival. To understand signaling pathways involved in this activity, we studied the effects on factor-dependent cell line MO7e cells incubated with SDF-1 alpha alone or in combination with other cytokines. SDF-1 alpha induced transient activation of extracellular stress-regulated kinase (ERK1/2), ribosomal S6 kinase (p90RSK) and Akt, molecules implicated in cell survival. Moreover, ERK1/2, p90RSK, and Akt were synergistically activated by SDF-1 alpha in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF), Steel factor (SLF), or thrombopoietin (TPO). Similar effects were seen after pretreatment of MO7e cells with SDF-1 alpha followed by stimulation with the other cytokines, suggesting a priming effect of SDF-1 alpha. Nuclear factor-kappa B (NF-kappa B) did not appear to be involved in SDF-1 alpha actions, alone or in combination with other cytokines. These intracellular effects were consistent with enhanced myeloid progenitor cell survival by SDF-1 alpha after delayed addition of growth factors. SDF-1 alpha alone supported survival of highly purified human cord blood CD34(+++) cells, less purified human cord blood, and MO7e cells; this effect was synergistically enhanced when SDF-1 alpha was combined with low amounts of other survival-promoting cytokines (GM-CSF, SLF, TPO, and FL). SDF-1 may contribute to maintenance of MPCs in bone marrow by enhancing cell survival alone and in combination with other cytokines.

  1. Protein expression profile of celiac disease patient with aberrant T cell by two-dimensional difference gel electrophoresis.

    PubMed

    De Re, Valli; Simula, Maria Paola; Caggiari, Laura; Ortz, Nicoletta; Spina, Michele; Da Ponte, Alessandro; De Appolonia, Leandro; Dolcetti, Riccardo; Canzonieri, Vincenzo; Cannizzaro, Renato

    2007-08-01

    One complication of celiac disease (CD) is refractory CD. These patients frequently show aberrant intraepithelial T cell clones and an increasing risk of evolution into enteropathy-associated T cell lymphoma (EATL). There is debate in the literature whether these cases are actually a smoldering lymphoma from the outset. The mechanism inducing T cell proliferation and prognosis remains unknown. Recently, alemtuzumab has been proposed as a promising new approach to treat these patients. Only few single cases have been tested presently, nevertheless, in all of them a clinical improvement has been observed, while intraepithelial lymphocytes (IELs) effectively targeted by alemtuzumab are still a debated issue. Using 2D-DIGE, we found hyperexpressed proteins specifically associated with aberrant T cell in a patient with CD by comparing the protein expression with that of patients with CD and polyclonal T cell or with that of control subjects (patients with polyclonal T cell and no CD). Proteins with a higher expression in duodenal biopsy of the patient with aberrant T cell were identified as IgM, apolipoprotein C-III, and Charcot-Leyden crystal proteins. These preliminary data allow hypothesizing different clinical effects of alemtuzumab in patients with CD, since besides the probable effect of alemtuzumab on T cell, it could effect inflammatory-associated CD52(+) IgM(+)B cell and eosinophils cells, known to produce IgM and Charcot-Leyden crystal proteins, which we demonstrated to be altered in this patient. Results also emphasize the possible association of apolipoprotein with aberrant T cell proliferation.

  2. Hematopoietic expression of oncogenic BRAF promotes aberrant growth of monocyte-lineage cells resistant to PLX4720

    PubMed Central

    Kamata, Tamihiro; Dankort, David; Kang, Jing; Giblett, Susan; Pritchard, Catrin A.; McMahon, Martin; Leavitt, Andrew D.

    2013-01-01

    Mutational activation of BRAF leading to expression of the BRAFV600E oncoprotein was recently identified in a high percentage of specific hematopoietic neoplasms in monocyte/histiocyte and mature B-cell lineages. Although BRAFV600E is a driver oncoprotein and pharmacological target in solid tumors such as melanoma, lung and thyroid cancer, it remains unknown whether BRAFV600E is an appropriate therapeutic target in hematopoietic neoplasms. To address this critical question, we generated a mouse model expressing inducible BRAFV600E in the hematopoietic system, and evaluated the efficacy of pathway-targeted therapeutics against primary hematopoietic cells. In this model, BRAFV600E expression conferred cytokine-independent growth to monocyte/macrophage-lineage progenitors leading to aberrant in vivo and in vitro monocyte/macrophage expansion. Furthermore, transplantation of BRAFV600E-expressing bone marrow cells promoted an in vivo pathology most notable for monocytosis in hematopoietic tissues and visceral organs. In vitro analysis revealed that MEK inhibition, but not RAF inhibition, effectively suppressed cytokine-independent clonal growth of monocyte/macrophage-lineage progenitors. However, combined RAF and PI3K inhibition effectively inhibited cytokine-independent colony formation, suggesting autocrine PI3K pathway activation. Taken together, these results provide evidence that constitutively activated BRAFV600E drives aberrant proliferation of monocyte-lineage cells. This study supports the development of pathway-targeted therapeutics in the treatment of BRAFV600E-expressing hematopoietic neoplasms in the monocyte/histiocyte lineage. PMID:24152792

  3. Cell cycle aberration in ameloblastoma and adenomatoid odontogenic tumor: As evidenced by the expression of p53 and survivin.

    PubMed

    Shaikh, Zulfin; Niranjan, K C

    2015-01-01

    p53 and survivin are involved in cell cycle progression and inhibition of apoptosis, respectively. Survivin is a unique protein which functions in progression of cell division and inhibits apoptosis leading to cell proliferation and cell survival. According to the literature, mutation of p53 leads to promotion of survivin function. Thus, the importance of cell cycle aberration and uncontrolled proliferation resulting from mutation of p53 and up-regulation of survivin is discussed. To assess the role of p53 and survivin in ameloblastoma and adenomatoid odontogenic tumor (AOT). The percentages of positive tumor cells were considered for statistical evaluation. Nuclear labeling index for p53 and nuclear, cytoplasmic and combined labeling index for survivin was obtained from the stained slides. Immunohistochemical expression of p53 and survivin was done qualitatively and quantitatively in 25 cases each of ameloblastoma and AOT. Mann-Whitney U-test, Wilcoxon signed ranks test and Pearson's correlation test. Quantitatively, p53 and survivin expression was statistically significant in AOT (P = 0.003) and qualitatively, in ameloblastoma (P = 0.004). Survivin expression was significant (P = 0.002) between the study groups unlike that of p53 (P = 0.554). There was no much difference in p53 expression in ameloblastoma and AOT suggestive of cell cycle aberration in both the odontogenic tumors, but significant difference in survivin expression in ameloblastoma and AOT with higher percentage of positive cells in ameloblastoma may be indicative of an aggressive behavior of ameloblastoma.

  4. The expression of CXCR4 is induced by the luteinizing hormone surge and mediated by progesterone receptors in human preovulatory granulosa cells.

    PubMed

    Choi, Yohan; Park, Ji Yeon; Wilson, Kalin; Rosewell, Katherine L; Brännström, Mats; Akin, James W; Curry, Thomas E; Jo, Misung

    2017-06-01

    The chemokine CXC motif ligand 12 (CXCL12) and its cognate receptor, CXCR4, have been implicated in the ovulatory process in various animal models. However, little is known about the expression and regulation of CXCL12 and CXCR4 and their functions during the ovulatory period in the human ovary. In this study, we characterized the expression patterns of CXCL12 and CXCR4 in preovulatory follicles collected before the luteinizing hormone (LH) surge and at defined hours after hCG administration in women with the regular menstrual cycle. The levels of mRNA and protein for CXCR4 were increased in granulosa cells of late ovulatory follicles, whereas CXCL12 expression was constant in follicles throughout the ovulatory period. Both CXCR4 and CXCL12 were localized to a subset of leukocytes around and inside the vasculature of human preovulatory follicles. Using a human granulosa cell culture model, the regulatory mechanisms and functions of CXCL12 and CXCR4 expression were investigated. Human chorionic gonadotropin (hCG) stimulated CXCR4 expression, whereas CXCL12 expression was not affected, mimicking in vivo expression patterns. Both RU486 (progesterone receptor antagonist) and CoCl2 (HIFs activator) blocked the hCG-induced increase in CXCR4 expression, whereas AG1478 (EGFR inhibitor) had no effect. The treatment with CXCL12 had no effect on granulosa cell viability but decreased hCG-stimulated CXCR4 expression. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. Aberrant light directly impairs mood and learning through melanopsin-expressing neurons.

    PubMed

    LeGates, Tara A; Altimus, Cara M; Wang, Hui; Lee, Hey-Kyoung; Yang, Sunggu; Zhao, Haiqing; Kirkwood, Alfredo; Weber, E Todd; Hattar, Samer

    2012-11-22

    The daily solar cycle allows organisms to synchronize their circadian rhythms and sleep-wake cycles to the correct temporal niche. Changes in day-length, shift-work, and transmeridian travel lead to mood alterations and cognitive function deficits. Sleep deprivation and circadian disruption underlie mood and cognitive disorders associated with irregular light schedules. Whether irregular light schedules directly affect mood and cognitive functions in the context of normal sleep and circadian rhythms remains unclear. Here we show, using an aberrant light cycle that neither changes the amount and architecture of sleep nor causes changes in the circadian timing system, that light directly regulates mood-related behaviours and cognitive functions in mice. Animals exposed to the aberrant light cycle maintain daily corticosterone rhythms, but the overall levels of corticosterone are increased. Despite normal circadian and sleep structures, these animals show increased depression-like behaviours and impaired hippocampal long-term potentiation and learning. Administration of the antidepressant drugs fluoxetine or desipramine restores learning in mice exposed to the aberrant light cycle, suggesting that the mood deficit precedes the learning impairments. To determine the retinal circuits underlying this impairment of mood and learning, we examined the behavioural consequences of this light cycle in animals that lack intrinsically photosensitive retinal ganglion cells. In these animals, the aberrant light cycle does not impair mood and learning, despite the presence of the conventional retinal ganglion cells and the ability of these animals to detect light for image formation. These findings demonstrate the ability of light to influence cognitive and mood functions directly through intrinsically photosensitive retinal ganglion cells.

  6. Aberrant methylation of the specific CpG island portion regulates cyclooxygenase-2 gene expression in human gastric carcinomas.

    PubMed

    Hur, Keun; Song, Sang Hyun; Lee, Hye Seung; Ho Kim, Woo; Bang, Yung-Jue; Yang, Han-Kwang

    2003-10-24

    Although it has been well established that overexpression of cyclooxygenase-2 (Cox-2) favors tumorigenesis and metastasis, the molecular mechanism that regulates Cox-2 expression has not been well defined in gastric carcinoma. Aberrant methylation of the CpG island is known to be one of the powerful mechanisms for the suppression of gene expression, and usually, CpG islands are very rich in promoter region and exon-1 region. But, it is controversial whether Cox-2 gene expression is regulated by methylation of promoter region or exon-1 region. In this study, we examined whether the hyper-methylation mediated transcriptional silencing of Cox-2 also occurred in human gastric carcinoma tissues and which portion of CpG island methylation is important in Cox-2 gene expression. Genomic DNAs from human gastric carcinoma tissues were treated with three methylation-sensitive restriction enzymes and then Southern blot analysis was performed. Out of 30 primary gastric tumor samples, 26 cases (86.6%) showed overexpression of Cox-2. Four cases (13.3%) with relatively decreased Cox-2 gene expression were associated with the presence of aberrant methylation of Cox-2 CpG island. We also found that methylation of promoter region and not exon-1 region is related with the transcriptional silencing of Cox-2 in gastric carcinoma cancer by detailed methylation mapping using bisulfite sequencing analysis. Our results suggest that the DNA methylation-mediated transcriptional silencing of Cox-2 is a predominant mechanism for the down-regulation of Cox-2 expression in human gastric carcinoma. Furthermore, the results suggest that methylation of not exon-1 region but promoter region is important to regulation of Cox-2 gene expression.

  7. A Novel, Non-canonical Splice Variant of the Ikaros Gene Is Aberrantly Expressed in B-cell Lymphoproliferative Disorders

    PubMed Central

    Mancarelli, Maria Michela; Verzella, Daniela; Fischietti, Mariafausta; Di Tommaso, Ambra; Maccarone, Rita; Plebani, Sara; Di Ianni, Mauro; Gulino, Alberto; Alesse, Edoardo

    2013-01-01

    The Ikaros gene encodes a Krüppel-like zinc-finger transcription factor involved in hematopoiesis regulation. Ikaros has been established as one of the most clinically relevant tumor suppressors in several hematological malignancies. In fact, expression of dominant negative Ikaros isoforms is associated with adult B-cell acute lymphoblastic leukemia, myelodysplastic syndrome, acute myeloid leukemia and adult and juvenile chronic myeloid leukemia. Here, we report the isolation of a novel, non-canonical Ikaros splice variant, called Ikaros 11 (Ik11). Ik11 is structurally related to known dominant negative Ikaros isoforms, due to the lack of a functional DNA-binding domain. Interestingly, Ik11 is the first Ikaros splice variant missing the transcriptional activation domain. Indeed, we demonstrated that Ik11 works as a dominant negative protein, being able to dimerize with Ikaros DNA-binding isoforms and inhibit their functions, at least in part by retaining them in the cytoplasm. Notably, we demonstrated that Ik11 is the first dominant negative Ikaros isoform to be aberrantly expressed in B-cell lymphoproliferative disorders, such as chronic lymphocytic leukemia. Aberrant expression of Ik11 interferes with both proliferation and apoptotic pathways, providing a mechanism for Ik11 involvement in tumor pathogenesis. Thus, Ik11 could represent a novel marker for B-cell lymphoproliferative disorders. PMID:23874502

  8. Cytoskeleton alterations in melanoma: aberrant expression of cortactin, an actin-binding adapter protein, correlates with melanocytic tumor progression

    PubMed Central

    Xu, Xu-Zhi; Garcia, Marileila Varella; Li, Tian-yu; Khor, Li-Yan; Gajapathy, R Sujatha; Spittle, Cindy; Weed, Scott; Lessin, Stuart R; Wu, Hong

    2010-01-01

    Cortactin is a multidomain actin-binding protein important for the functions of cytoskeleton by regulating cortical actin dynamics. It is involved in a diverse array of basic cellular functions. Tumorigenesis and tumor progression involves alterations in actin cytoskeleton proteins. We sought to study the role of cortactin in melanocytic tumor progression using immunohistochemistry on human tissues. The results reveal quantitative differences between benign and malignant lesions. Significantly higher cortactin expression is found in melanomas than in nevi (P<0.0001), with levels greater in metastatic than in invasive melanomas (P<0.05). Qualitatively, tumor tissues often show aberrant cortactin localization at the cell periphery, corresponding to its colocalization with filamentous actin in cell cortex of cultured melanoma cells. This suggests an additional level of protein dysregulation. Furthermore, in patients with metastatic disease, high-level cortactin expression correlates with poor disease-specific survival. Our data, in conjunction with outcome data on several other types of human cancers and experimental data from melanoma cell lines, supports a potential role of aberrant cortactin expression in melanoma tumor progression and a rational for targeting key elements of actin-signaling pathway for developmental therapeutics in melanomas. PMID:19898426

  9. Transcription factor LSF-DNMT1 complex dissociation by FQI1 leads to aberrant DNA methylation and gene expression

    PubMed Central

    Chin, Hang Gyeong; Ponnaluri, V.K. Chaithanya; Zhang, Guoqiang; Estève, Pierre-Olivier; Schaus, Scott E.; Hansen, Ulla; Pradhan, Sriharsa

    2016-01-01

    The transcription factor LSF is highly expressed in hepatocellular carcinoma (HCC) and promotes oncogenesis. Factor quinolinone inhibitor 1 (FQI1), inhibits LSF DNA-binding activity and exerts anti-proliferative activity. Here, we show that LSF binds directly to the maintenance DNA (cytosine-5) methyltransferase 1 (DNMT1) and its accessory protein UHRF1 both in vivo and in vitro. Binding of LSF to DNMT1 stimulated DNMT1 activity and FQI1 negated the methyltransferase activation. Addition of FQI1 to the cell culture disrupted LSF bound DNMT1 and UHRF1 complexes, resulting in global aberrant CpG methylation. Differentially methylated regions (DMR) containing at least 3 CpGs, were significantly altered by FQI1 compared to control cells. The DMRs were mostly concentrated in CpG islands, proximal to transcription start sites, and in introns and known genes. These DMRs represented both hypo and hypermethylation, correlating with altered gene expression. FQI1 treatment elicits a cascade of effects promoting altered cell cycle progression. These findings demonstrate a novel mechanism of FQI1 mediated alteration of the epigenome by DNMT1-LSF complex disruption, leading to aberrant DNA methylation and gene expression. PMID:27845898

  10. Involvement of aberrant DNA methylation on reduced expression of lysophosphatidic acid receptor-1 gene in rat tumor cell lines

    SciTech Connect

    Tsujiuchi, Toshifumi . E-mail: ttujiuch@life.kindai.ac.jp; Shimizu, Kyoko; Onishi, Mariko; Sugata, Eriko; Fujii, Hiromasa; Mori, Toshio; Honoki, Kanya; Fukushima, Nobuyuki

    2006-10-27

    Lysophosphatidic acid (LPA) is a bioactive phospholipid that stimulates cell proliferation, migration, and protects cells from apoptosis. It interacts with specific G protein-coupled transmembrane receptors. Recently, it has been reported that alterations of LPA receptor expression might be important in the malignant transformation of tumor cells. Therefore, to assess an involvement of DNA methylation in reduced expression of the LPA receptor-1 (lpa1) gene, we investigated the expression of the lpa1 gene and its DNA methylation patterns in rat tumor cell lines. Both rat brain-derived neuroblastoma B103 and liver-derived hepatoma RH7777 cells used in this study indicated no expression of lpa1. For the analysis of methylation status, bisulfite sequencing was performed with B103 and RH7777 cells, comparing with other lpa1 expressed cells and normal tissues of brain and liver. The lpa1 expressed cells and tissues were all unmethylated in this region of lpa1. In contrast, both B103 and RH7777 cells were highly methylated, correlating with reduced expression of the lpa1. Treatment with 5-aza 2'-deoxycytidine induced expression of lpa1 gene in B103 and RH7777 cells after 24 h. In RH7777 cells treated with 5-aza 2'-deoxycytidine, stress fiber formation was also observed in response to LPA in RH7777 cells, but not in untreated RH7777 cells. These results suggest that aberrant DNA methylation of the lpa1 gene may be involved in its reduced expression in rat tumor cells.

  11. Comparison of methods to identify aberrant expression patterns in individual patients: augmenting our toolkit for precision medicine

    PubMed Central

    2013-01-01

    Background Patient-specific aberrant expression patterns in conjunction with functional screening assays can guide elucidation of the cancer genome architecture and identification of therapeutic targets. Since most statistical methods for expression analysis are focused on differences between experimental groups, the performance of approaches for patient-specific expression analyses are currently less well characterized. A comparison of methods for the identification of genes that are dysregulated relative to a single sample in a given set of experimental samples, to our knowledge, has not been performed. Methods We systematically evaluated several methods including variations on the nearest neighbor based outlying degree method, as well as the Zscore and a robust variant for their suitability to detect patient-specific events. The methods were assessed using both simulations and expression data from a cohort of pediatric acute B lymphoblastic leukemia patients. Results We first assessed power and false discovery rates using simulations and found that even under optimal conditions, high effect sizes (>4 unit differences) were necessary to have acceptable power for any method (>0.9) though high false discovery rates (>0.1) were pervasive across simulation conditions. Next we introduced a technical factor into the simulation and found that performance was reduced for all methods and that using weights with the outlying degree could provide performance gains depending on the number of samples and genes affected by the technical factor. In our use case that highlights the integration of functional assays and aberrant expression in a patient cohort (the identification of gene dysregulation events associated with the targets from a siRNA screen), we demonstrated that both the outlying degree and the Zscore can successfully identify genes dysregulated in one patient sample. However, only the outlying degree can identify genes dysregulated across several patient samples

  12. Aberrant expression of Notch1, HES1, and DTX1 genes in glioblastoma formalin-fixed paraffin-embedded tissues.

    PubMed

    Narayanappa, Rajeswari; Rout, Pritilata; Aithal, Madhuri G S; Chand, Ashis Kumar

    2016-05-01

    Glioblastoma is the most common malignant brain tumor accounting for more than 54 % of all gliomas. Despite aggressive treatments, median survival remains less than 1 year. This might be due to the unavailability of effective molecular diagnostic markers and targeted therapy. Thus, it is essential to discover molecular mechanisms underlying disease by identifying dysregulated pathways involved in tumorigenesis. Notch signaling is one such pathway which plays an important role in determining cell fates. Since it is found to play a critical role in many cancers, we investigated the role of Notch genes in glioblastoma with an aim to identify biomarkers that can improve diagnosis. Using real-time PCR, we assessed the expression of Notch genes including receptors (Notch1, Notch2, Notch3, and Notch4), ligands (JAG1, JAG2, and DLL3), downstream targets (HES1 and HEY2), regulator Deltex1 (DTX1), inhibitor NUMB along with transcriptional co-activator MAML1, and a component of gamma-secretase complex APH1A in 15 formalin-fixed paraffin-embedded (FFPE) patient samples. Relative quantification was done by the 2(-ΔΔCt) method; the data are presented as fold change in gene expression normalized to an internal control gene and relative to the calibrator. The data revealed aberrant expression of Notch genes in glioblastoma compared to normal brain. More than 85 % of samples showed high Notch1 (P = 0.0397) gene expression and low HES1 (P = 0.011) and DTX1 (P = 0.0001) gene expression. Our results clearly show aberrant expression of Notch genes in glioblastoma which can be used as putative biomarkers together with histopathological observation to improve diagnosis, therapeutic strategies, and patient prognosis.

  13. Divergent cyclin B1 expression and Rb/p16/cyclin D1 pathway aberrations among pulmonary neuroendocrine tumors.

    PubMed

    Igarashi, Toru; Jiang, Shi-Xu; Kameya, Toru; Asamura, Hisao; Sato, Yuichi; Nagai, Kanji; Okayasu, Isao

    2004-10-01

    A total of 111 pulmonary neuroendocrine tumors comprising 13 typical carcinoids, five atypical carcinoids, 44 large-cell neuroendocrine carcinomas and 49 small-cell carcinomas were immunohistochemically studied for dysregulated cyclin B1 expression and disruption of the Rb/p16/cyclin D1 pathway (Rb pathway), and the results were correlated with tumor proliferation activity and clinical outcome. Overexpression of cyclins B1 and D1, respectively, was detected in no and 15% typical carcinoids, 20 and 20% atypical carcinoids, 84 and 32% large-cell neuroendocrine carcinomas, 84 and 10% small-cell carcinomas. Loss of Rb and p16 expression, respectively, was observed in no and 14% typical carcinoids, no and 40% atypical carcinoids, 49 and 18% large-cell neuroendocrine carcinomas, 84 and 8% small-cell carcinomas. In summary, 29% typical carcinoids, 20% atypical carcinoids, 78% large-cell neuroendocrine carcinomas and 93% small-cell carcinomas had Rb pathway aberrations. Rb pathway aberration was mostly attributed to Rb loss in small-cell carcinomas, while p16 loss and/or cyclin D1 overexpression besides Rb loss also played an important role in large-cell neuroendocrine carcinomas, while cyclin D1 overexpression was the only cause of Rb pathway aberration in carcinoid tumors. Thus, both cyclin B1-associated G2/M arrest and Rb-mediated G1 arrest are consistently compromised in high-grade large-cell neuroendocrine carcinoma and small-cell carcinoma, but are generally intact or occasionally altered in carcinoid tumor; the mechanisms involved in Rb pathway aberration among the tumor categories are different, reflecting a genetic divergence among the individual tumor categories. Cyclin B1 expression closely correlated with the Ki-67 labeling index either in the individual tumor categories or overall tumors (P < 0.0001, r = 0.742), suggesting that cyclin B1 is one of the key factors regulating cell proliferation in pulmonary neuroendocrine tumors. Neither cyclins B1 and D1, Rb, p

  14. Involvement of aberrantly expressed microRNAs in the pathogenesis of head and neck squamous cell carcinoma.

    PubMed

    Koshizuka, Keiichi; Hanazawa, Toyoyuki; Arai, Takayuki; Okato, Atsushi; Kikkawa, Naoko; Seki, Naohiko

    2017-08-23

    MicroRNAs (miRNAs) are small noncoding RNAs that act as fine-tuners of the post-transcriptional control of protein-coding or noncoding RNAs by repressing translation or cleaving RNA transcripts in a sequence-dependent manner in cells. Accumulating evidence have been indicated that aberrantly expressed miRNAs are deeply involved in human pathogenesis, including cancers. Surprisingly, these small, single-stranded RNAs (18-23 nucleotides) have been shown to function as antitumor or oncogenic RNAs in several types of cancer cells. A single miRNA has regulating hundreds or thousands of different mRNAs, and individual mRNA has been regulated by multiple different miRNAs in normal cells. Therefore, tightly controlled RNA networks can be disrupted by dysregulated of miRNAs in cancer cells. Investigation of novel miRNA-mediated RNA networks in cancer cells could provide new insights in the field of cancer research. In this review, we focus on head and neck squamous cell carcinoma (HNSCC) and discuss current findings of the involvement of aberrantly expressed miRNAs in the pathogenesis of HNSCC.

  15. Oligoamine analogues in combination with 2-difluoromethylornithine synergistically induce re-expression of aberrantly silenced tumour-suppressor genes

    PubMed Central

    Wu, Yu; Steinbergs, Nora; Murray-Stewart, Tracy; Marton, Laurence J.; Casero, Robert A.

    2011-01-01

    Epigenetic gene silencing is an important mechanism in the initiation and progression of cancer. Abnormal DNA CpG island hypermethylation and histone modifications are involved in aberrant silencing of tumour-suppressor genes. LSD1 (lysine-specific demethylase 1) was the first enzyme identified to specifically demethylate H3K4 (Lys4 of histone H3). Methylated H3K4 is an important mark associated with transcriptional activation. The flavin adenine dinucleotide-binding amine oxidase domain of LSD1 is homologous with two polyamine oxidases, SMO (spermine oxidase) and APAO (N1-acetylpolyamine oxidase). We have demonstrated previously that long-chain polyamine analogues, the oligoamines, are inhibitors of LSD1. In the present paper we report the synergistic effects of specific oligoamines in combination with DFMO (2-difluoromethylornithine), an inhibitor of ornithine decarboxylase, in human colorectal cancer cells. DFMO treatment depletes natural polyamines and increases the uptake of exogenous polyamines. The combination of oligoamines and DFMO results in a synergistic re-expression of aberrantly silenced tumour-suppressor genes, including SFRP2 (secreted frizzled-related protein 2), which encodes a Wnt signalling pathway antagonist and plays an anti-tumorigenic role in colorectal cancer. The treatment-induced re-expression of SFRP2 is associated with increased H3K4me2 (di-methyl H3K4) in the gene promoter. The combination of LSD1-inhibiting oligoamines and DFMO represents a novel approach to epigenetic therapy of cancer. PMID:22132744

  16. Oligoamine analogues in combination with 2-difluoromethylornithine synergistically induce re-expression of aberrantly silenced tumour-suppressor genes.

    PubMed

    Wu, Yu; Steinbergs, Nora; Murray-Stewart, Tracy; Marton, Laurence J; Casero, Robert A

    2012-03-15

    Epigenetic gene silencing is an important mechanism in the initiation and progression of cancer. Abnormal DNA CpG island hypermethylation and histone modifications are involved in aberrant silencing of tumour-suppressor genes. LSD1 (lysine-specific demethylase 1) was the first enzyme identified to specifically demethylate H3K4 (Lys(4) of histone H3). Methylated H3K4 is an important mark associated with transcriptional activation. The flavin adenine dinucleotide-binding amine oxidase domain of LSD1 is homologous with two polyamine oxidases, SMO (spermine oxidase) and APAO (N(1)-acetylpolyamine oxidase). We have demonstrated previously that long-chain polyamine analogues, the oligoamines, are inhibitors of LSD1. In the present paper we report the synergistic effects of specific oligoamines in combination with DFMO (2-difluoromethylornithine), an inhibitor of ornithine decarboxylase, in human colorectal cancer cells. DFMO treatment depletes natural polyamines and increases the uptake of exogenous polyamines. The combination of oligoamines and DFMO results in a synergistic re-expression of aberrantly silenced tumour-suppressor genes, including SFRP2 (secreted frizzled-related protein 2), which encodes a Wnt signalling pathway antagonist and plays an anti-tumorigenic role in colorectal cancer. The treatment-induced re-expression of SFRP2 is associated with increased H3K4me2 (di-methyl H3K4) in the gene promoter. The combination of LSD1-inhibiting oligoamines and DFMO represents a novel approach to epigenetic therapy of cancer.

  17. Enhanced expression of ADCY1 underlies aberrant neuronal signalling and behaviour in a syndromic autism model

    PubMed Central

    Sethna, Ferzin; Feng, Wei; Ding, Qi; Robison, Alfred J.; Feng, Yue; Wang, Hongbing

    2017-01-01

    Fragile X syndrome (FXS), caused by the loss of functional FMRP, is a leading cause of autism. Neurons lacking FMRP show aberrant mRNA translation and intracellular signalling. Here, we identify that, in Fmr1 knockout neurons, type 1 adenylyl cyclase (Adcy1) mRNA translation is enhanced, leading to excessive production of ADCY1 protein and insensitivity to neuronal stimulation. Genetic reduction of Adcy1 normalizes the aberrant ERK1/2- and PI3K-mediated signalling, attenuates excessive protein synthesis and corrects dendritic spine abnormality in Fmr1 knockout mice. Genetic reduction of Adcy1 also ameliorates autism-related symptoms including repetitive behaviour, defective social interaction and audiogenic seizures. Moreover, peripheral administration of NB001, an experimental compound that preferentially suppresses ADCY1 activity over other ADCY subtypes, attenuates the behavioural abnormalities in Fmr1 knockout mice. These results demonstrate a connection between the elevated Adcy1 translation and abnormal ERK1/2 signalling and behavioural symptoms in FXS. PMID:28218269

  18. TGF-{beta}-stimulated aberrant expression of class III {beta}-tubulin via the ERK signaling pathway in cultured retinal pigment epithelial cells

    SciTech Connect

    Chung, Eun Jee; Chun, Ji Na; Jung, Sun-Ah; Cho, Jin Won; Lee, Joon H.

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer TGF-{beta} induces aberrant expression of {beta}III in RPE cells via the ERK pathway. Black-Right-Pointing-Pointer TGF-{beta} increases O-GlcNAc modification of {beta}III in RPE cells. Black-Right-Pointing-Pointer Mature RPE cells have the capacity to express a neuron-associated gene by TGF-{beta}. -- Abstract: The class III {beta}-tubulin isotype ({beta}{sub III}) is expressed exclusively by neurons within the normal human retina and is not present in normal retinal pigment epithelial (RPE) cells in situ or in the early phase of primary cultures. However, aberrant expression of class III {beta}-tubulin has been observed in passaged RPE cells and RPE cells with dedifferentiated morphology in pathologic epiretinal membranes from idiopathic macular pucker, proliferative vitreoretinopathy (PVR) and proliferative diabetic retinopathy (PDR). Transforming growth factor-{beta} (TGF-{beta}) has been implicated in dedifferentiation of RPE cells and has a critical role in the development of proliferative vitreoretinal diseases. Here, we investigated the potential effects of TGF-{beta} on the aberrant expression of class III {beta}-tubulin and the intracellular signaling pathway mediating these changes. TGF-{beta}-induced aberrant expression and O-linked-{beta}-N-acetylglucosamine (O-GlcNac) modification of class III {beta}-tubulin in cultured RPE cells as determined using Western blotting, RT-PCR and immunocytochemistry. TGF-{beta} also stimulated phosphorylation of ERK. TGF-{beta}-induced aberrant expression of class III {beta}-tubulin was significantly reduced by pretreatment with U0126, an inhibitor of ERK phosphorylation. Our findings indicate that TGF-{beta} stimulated aberrant expression of class III {beta}-tubulin via activation of the ERK signaling pathway. These data demonstrate that mature RPE cells have the capacity to express a neuron-associated gene in response to TGF-{beta} stimulation and provide useful information

  19. CXC Chemokine Receptor 7 (CXCR7) Regulates CXCR4 Protein Expression and Capillary Tuft Development in Mouse Kidney

    PubMed Central

    Haege, Sammy; Mueller, Wiebke; Nietzsche, Sandor; Lupp, Amelie; Mackay, Fabienne; Schulz, Stefan; Stumm, Ralf

    2012-01-01

    Background The CXCL12/CXCR4 axis is involved in kidney development by regulating formation of the glomerular tuft. Recently, a second CXCL12 receptor was identified and designated CXCR7. Although it is established that CXCR7 regulates heart and brain development in conjunction with CXCL12 and CXCR4, little is known about the influence of CXCR7 on CXCL12 dependent kidney development. Methodology/Principal Findings We provided analysis of CXCR7 expression and function in the developing mouse kidney. Using in situ hybridization, we identified CXCR7 mRNA in epithelial cells including podocytes at all nephron stages up to the mature glomerulus. CXCL12 mRNA showed a striking overlap with CXCR7 mRNA in epithelial structures. In addition, CXCL12 was detected in stromal cells and the glomerular tuft. Expression of CXCR4 was complementary to that of CXCR7 as it occurred in mesenchymal cells, outgrowing ureteric buds and glomerular endothelial cells but not in podocytes. Kidney examination in CXCR7 null mice revealed ballooning of glomerular capillaries as described earlier for CXCR4 null mice. Moreover, we detected a severe reduction of CXCR4 protein but not CXCR4 mRNA within the glomerular tuft and in the condensed mesenchyme. Malformation of the glomerular tuft in CXCR7 null mice was associated with mesangial cell clumping. Conclusions/Significance We established that there is a similar glomerular pathology in CXCR7 and CXCR4 null embryos. Based on the phenotype and the anatomical organization of the CXCL12/CXCR4/CXCR7 system in the forming glomerulus, we propose that CXCR7 fine-tunes CXCL12/CXCR4 mediated signalling between podocytes and glomerular capillaries. PMID:22880115

  20. CXC chemokine receptor 7 (CXCR7) regulates CXCR4 protein expression and capillary tuft development in mouse kidney.

    PubMed

    Haege, Sammy; Einer, Claudia; Thiele, Stefanie; Mueller, Wiebke; Nietzsche, Sandor; Lupp, Amelie; Mackay, Fabienne; Schulz, Stefan; Stumm, Ralf

    2012-01-01

    The CXCL12/CXCR4 axis is involved in kidney development by regulating formation of the glomerular tuft. Recently, a second CXCL12 receptor was identified and designated CXCR7. Although it is established that CXCR7 regulates heart and brain development in conjunction with CXCL12 and CXCR4, little is known about the influence of CXCR7 on CXCL12 dependent kidney development. We provided analysis of CXCR7 expression and function in the developing mouse kidney. Using in situ hybridization, we identified CXCR7 mRNA in epithelial cells including podocytes at all nephron stages up to the mature glomerulus. CXCL12 mRNA showed a striking overlap with CXCR7 mRNA in epithelial structures. In addition, CXCL12 was detected in stromal cells and the glomerular tuft. Expression of CXCR4 was complementary to that of CXCR7 as it occurred in mesenchymal cells, outgrowing ureteric buds and glomerular endothelial cells but not in podocytes. Kidney examination in CXCR7 null mice revealed ballooning of glomerular capillaries as described earlier for CXCR4 null mice. Moreover, we detected a severe reduction of CXCR4 protein but not CXCR4 mRNA within the glomerular tuft and in the condensed mesenchyme. Malformation of the glomerular tuft in CXCR7 null mice was associated with mesangial cell clumping. We established that there is a similar glomerular pathology in CXCR7 and CXCR4 null embryos. Based on the phenotype and the anatomical organization of the CXCL12/CXCR4/CXCR7 system in the forming glomerulus, we propose that CXCR7 fine-tunes CXCL12/CXCR4 mediated signalling between podocytes and glomerular capillaries.

  1. Aberrant p53 protein expression is associated with an increased risk of neoplastic progression in patients with Barrett's oesophagus.

    PubMed

    Kastelein, Florine; Biermann, Katharina; Steyerberg, Ewout W; Verheij, Joanne; Kalisvaart, Marit; Looijenga, Leendert H J; Stoop, Hans A; Walter, Laurens; Kuipers, Ernst J; Spaander, Manon C W; Bruno, Marco J

    2013-12-01

    The value of surveillance for patients with Barrett's oesophagus (BO) is under discussion given the overall low incidence of neoplastic progression and lack of discriminative tests for risk stratification. Histological diagnosis of low-grade dysplasia (LGD) is the only accepted predictor for progression to date, but has a low predictive value. The aim of this study was therefore to evaluate the value of p53 immunohistochemistry for predicting neoplastic progression in patients with BO. We conducted a case-control study within a prospective cohort of 720 patients with BO. Patients who developed high-grade dysplasia (HGD) or oesophageal adenocarcinoma (OAC) were classified as cases and patients without neoplastic progression were classified as controls. P53 protein expression was determined by immunohistochemistry in more than 12 000 biopsies from 635 patients and was scored independently by two expert pathologists who were blinded to long-term outcome. During follow-up, 49 (8%) patients developed HGD or OAC. P53 overexpression was associated with an increased risk of neoplastic progression in patients with BO after adjusting for age, gender, Barrett length and oesophagitis (adjusted relative risks (RR(a)) 5.6; 95% CI 3.1 to 10.3), but the risk was even higher with loss of p53 expression (RR(a) 14.0; 95% CI 5.3 to 37.2). The positive predictive value for neoplastic progression increased from 15% with histological diagnosis of LGD to 33% with LGD and concurrent aberrant p53 expression. Aberrant p53 protein expression is associated with an increased risk of neoplastic progression in patients with BO and appears to be a more powerful predictor of neoplastic progression than histological diagnosis of LGD.

  2. Systems biology of myasthenia gravis, integration of aberrant lncRNA and mRNA expression changes.

    PubMed

    Luo, ZhaoHui; Li, Ye; Liu, XiaoFang; Luo, MengChuan; Xu, LiQun; Luo, YueBei; Xiao, Bo; Yang, Huan

    2015-03-18

    A novel class of transcripts, long non-coding RNAs (lncRNAs), has recently emerged as a key player in several biological processes, and important roles for these molecules have been reported in a number of complex human diseases, such as autoimmune diseases, neurological disorders, and various cancers. However, the aberrant lncRNAs implicated in myasthenia gravis (MG) remain unknown. The aim of the present study was to explore the abnormal expression of lncRNAs in peripheral blood mononuclear cells (PBMCs) and examine mRNA regulatory relationship networks among MG patients with or without thymoma. Microarray assays were performed, and the outstanding differences between lncRNAs or mRNA expression were verified through RT-PCR. The lncRNAs functions were annotated for the target genes using Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) biological pathway. The potential regulatory relationships between the lncRNAs and target genes were analyzed using the 'cis' and 'trans' model. Outstanding lncRNAs were organized to generate a TF-lncRNA-gene network using Cytoscape software. The lncRNA and mRNA expression profile analysis revealed subsets of differentially expressed genes in MG patients with or without thymoma. A total of 12 outstanding dysregulated expression lncRNAs, such as lncRNA oebiotech_11933, were verified through real-time PCR. Several GO terms including the cellular response to interferon-γ, platelet degranulation, chemokine receptor binding and cytokine interactions were very important in MG pathogenesis. The chromosome locations of some lncRNAs and associated co-expression genes were demonstrated using 'cis' analysis. The results of the 'trans' analysis revealed that some TFs (i.e., CTCF, TAF1and MYC) regulate lncRNA and gene expression. The outstanding lncRNAs in each group were implicated in the regulation of the TF-lncRNA-target gene network. The results of the present study provide a perspective on lncRNA expression in MG

  3. Aberrant expression of nuclear HDAC3 and cytoplasmic CDH1 predict a poor prognosis for patients with pancreatic cancer.

    PubMed

    Jiao, Feng; Hu, Hai; Han, Ting; Zhuo, Meng; Yuan, Cuncun; Yang, Haiyan; Wang, Lei; Wang, Liwei

    2016-03-29

    Previous studies showed that aberrant CDH1 or/and HDAC3 localization is essential for the progression of some human cancers. Here, we investigate the prognostic significance of aberrant CDH1 and HDAC3 localization in 84 pancreatic cancer patients. Our results show that increases in both membrane and cytoplasmic CDH1 correlate with lymph node metastasis (P = 0.026 and P < 0.001, respectively) and clinical stage (P = 0.020 and P < 0.001, respectively). Increased nuclear HDAC3 correlates with lymph node metastasis (P < 0.001) and advanced clinical stage (P < 0.001), but increased cytoplasmic HDAC3 does not (P > 0.05). Multivariate analysis showed that nuclear HDAC3 and cytoplasmic CDH1 (P = 0.001 and P = 0.010, respectively), as well as tumor differentiation (P = 0.009) are independent prognostic factors. Most importantly, patients with high co-expression of nuclear HDAC3 and cytoplasmic CDH1 had shorter survival times (P < 0.001), more frequent lymph node metastasis (P < 0.001), and advanced clinical stage (P < 0.001). Our studies provide convincing evidence that nuclear HDAC3 and cytoplasmic CDH1 have independent prognostic value in pancreatic cancer and provide novel targets for prognostic therapeutics.

  4. Expression of Leukemia-Associated Nup98 Fusion Proteins Generates an Aberrant Nuclear Envelope Phenotype.

    PubMed

    Fahrenkrog, Birthe; Martinelli, Valérie; Nilles, Nadine; Fruhmann, Gernot; Chatel, Guillaume; Juge, Sabine; Sauder, Ursula; Di Giacomo, Danika; Mecucci, Cristina; Schwaller, Jürg

    2016-01-01

    Chromosomal translocations involving the nucleoporin NUP98 have been described in several hematopoietic malignancies, in particular acute myeloid leukemia (AML). In the resulting chimeric proteins, Nup98's N-terminal region is fused to the C-terminal region of about 30 different partners, including homeodomain (HD) transcription factors. While transcriptional targets of distinct Nup98 chimeras related to immortalization are relatively well described, little is known about other potential cellular effects of these fusion proteins. By comparing the sub-nuclear localization of a large number of Nup98 fusions with HD and non-HD partners throughout the cell cycle we found that while all Nup98 chimeras were nuclear during interphase, only Nup98-HD fusion proteins exhibited a characteristic speckled appearance. During mitosis, only Nup98-HD fusions were concentrated on chromosomes. Despite the difference in localization, all tested Nup98 chimera provoked morphological alterations in the nuclear envelope (NE), in particular affecting the nuclear lamina and the lamina-associated polypeptide 2α (LAP2α). Importantly, such aberrations were not only observed in transiently transfected HeLa cells but also in mouse bone marrow cells immortalized by Nup98 fusions and in cells derived from leukemia patients harboring Nup98 fusions. Our findings unravel Nup98 fusion-associated NE alterations that may contribute to leukemogenesis.

  5. Expression of Leukemia-Associated Nup98 Fusion Proteins Generates an Aberrant Nuclear Envelope Phenotype

    PubMed Central

    Fahrenkrog, Birthe; Martinelli, Valérie; Nilles, Nadine; Fruhmann, Gernot; Chatel, Guillaume; Juge, Sabine; Sauder, Ursula; Di Giacomo, Danika; Mecucci, Cristina; Schwaller, Jürg

    2016-01-01

    Chromosomal translocations involving the nucleoporin NUP98 have been described in several hematopoietic malignancies, in particular acute myeloid leukemia (AML). In the resulting chimeric proteins, Nup98's N-terminal region is fused to the C-terminal region of about 30 different partners, including homeodomain (HD) transcription factors. While transcriptional targets of distinct Nup98 chimeras related to immortalization are relatively well described, little is known about other potential cellular effects of these fusion proteins. By comparing the sub-nuclear localization of a large number of Nup98 fusions with HD and non-HD partners throughout the cell cycle we found that while all Nup98 chimeras were nuclear during interphase, only Nup98-HD fusion proteins exhibited a characteristic speckled appearance. During mitosis, only Nup98-HD fusions were concentrated on chromosomes. Despite the difference in localization, all tested Nup98 chimera provoked morphological alterations in the nuclear envelope (NE), in particular affecting the nuclear lamina and the lamina-associated polypeptide 2α (LAP2α). Importantly, such aberrations were not only observed in transiently transfected HeLa cells but also in mouse bone marrow cells immortalized by Nup98 fusions and in cells derived from leukemia patients harboring Nup98 fusions. Our findings unravel Nup98 fusion-associated NE alterations that may contribute to leukemogenesis. PMID:27031510

  6. Aberrant expressions of c-KIT and DOG-1 in mucinous and nonmucinous colorectal carcinomas and relation to clinicopathologic features and prognosis.

    PubMed

    Foda, Abd Al-Rahman Mohammad; Mohamed, Mie Ali

    2015-10-01

    c-KIT and DOG-1 are 2 highly expressed proteins in gastrointestinal stromal tumors. Few studies had investigated c-KIT, but not DOG-1, expression in colorectal carcinoma (CRC). This study aims to investigate expressions of c-KIT and DOG-1 in colorectal mucinous carcinoma and nonmucinous carcinoma using manual tissue microarray technique. In this work, we studied tumor tissue specimens from 150 patients with colorectal mucinous (MA) and nonmucinous adenocarcinoma (NMA). High-density manual tissue microarrays were constructed using modified mechanical pencil tip technique, and immunohistochemistry for c-KIT and DOG-1 was done. We found that aberrant c-KIT expression was detected in 12 cases (8%); 6 cases (4%) showed strong expression. Aberrant DOG-1 expression was detected in 15 cases (10%); among them, only 4 cases (2.7%) showed strong expression. Nonmucinous adenocarcinoma showed a significantly high expression of c-KIT, but not DOG-1, than MA. Aberrant c-KIT and DOG-1 expressions were significantly unrelated but were associated with excessive microscopic abscess formation. Neither c-KIT nor DOG-1 expression showed a significant impact on disease-free survival or overall survival. In conclusion, aberrant c-KIT and DOG-1 expressions in CRC are rare events, either in NMA or MA. Nonmucinous adenocarcinoma showed a significantly higher expression of c-KIT, but not DOG-1, than MA. The expressions of both in CRC are significantly unrelated but are associated with microscopic abscess formation. Neither c-KIT nor DOG-1 expression showed a significant impact on disease-free survival or overall survival. So, c-KIT and DOG-1 immunostaining is not a cost-effective method of identifying patients with CRC who may benefit from treatment with tyrosine kinase inhibitors.

  7. The haematopoietic specific signal transducer Vav1 is aberrantly expressed in lung cancer and plays a role in tumourigenesis.

    PubMed

    Lazer, Galit; Idelchuk, Yulia; Schapira, Vered; Pikarsky, Eli; Katzav, Shulamit

    2009-09-01

    Lung cancer is the leading cause of cancer death worldwide. The spectrum of aberrations affecting signalling pathways in lung cancer pathogenesis has not been fully elucidated. Physiological expression of Vav1 is restricted to the haematopoietic system, where its best-known function is as a GDP/GTP nucleotide exchange factor for Rho/RacGTPases, an activity strictly controlled by tyrosine phosphorylation downstream of cell surface receptors. Here we find Vav1 expression in 42% of 78 lung cancer cell lines analysed. Moreover, immunohistochemical analysis of primary human lung cancer tissue samples revealed Vav1 expression in 26/59 malignant samples, including adenocarcinoma, squamous cell carcinoma and bronchioloalveolar carcinoma. Stronger Vav1 staining was associated with larger tumour size. siRNA-mediated knockdown of Vav1 in lung cancer cells reduced proliferation in agar and tumour growth in nude mice, while control siRNA had no effect, suggesting that Vav1 plays a critical role in the tumorigenicity of lung cancer cells. Vav1 is tyrosine-phosphorylated in lung cancer cells following activation by the growth factors EGF and TGFalpha, suggesting its participation in signalling events in these cells. Depletion of Vav1 reduced Rac-GTP activation and decreased expression of TGFalpha, an autocrine growth factor. These data suggest that Vav1 plays a role in the neoplastic process in lung cancer, identifying it as a potential therapeutic target for lung cancer therapy.

  8. Microarray-based genomic profiling reveals novel genomic aberrations in follicular lymphoma which associate with patient survival and gene expression status.

    PubMed

    Schwaenen, Carsten; Viardot, Andreas; Berger, Hilmar; Barth, Thomas F E; Bentink, Stefan; Döhner, Hartmut; Enz, Martina; Feller, Alfred C; Hansmann, Martin-Leo; Hummel, Michael; Kestler, Hans A; Klapper, Wolfram; Kreuz, Markus; Lenze, Dido; Loeffler, Markus; Möller, Peter; Müller-Hermelink, Hans-Konrad; Ott, German; Rosolowski, Maciej; Rosenwald, Andreas; Ruf, Sandra; Siebert, Reiner; Spang, Rainer; Stein, Harald; Truemper, Lorenz; Lichter, Peter; Bentz, Martin; Wessendorf, Swen

    2009-01-01

    Follicular lymphoma (FL) is characterized by a large number of chromosomal aberrations. However, their exact genomic extension and involved target genes remain to be determined. For this purpose, we used array-based intermediate-high resolution genomic profiling in combination with Affymetrix gene expression analysis. Tumor specimens from 128 FL patients were analyzed for the presence of genomic aberrations and the results were correlated to clinical data sets and mRNA expression levels. In 114 (89%) of the 128 analyzed cases, a total of 688 genomic aberrations (384 gains/amplifications and 304 losses) were detected. Frequent genomic aberrations were: -1p36 (18%), +2p15 (24%), -3q (14%), -6q (25%), +7p (19%), +7q (23%), +8q (14%), -9p (16%), -11q (15%), +12q (20%), -13q (11%), -17p (16%), +18p (18%), and +18q (28%). Critical segments of these imbalances were delineated to genomic fragments with a minimum size down to 0.2 Mb. By comparison of these with mRNA gene expression data, putative candidate genes were identified. Moreover, we found that deletions affecting the tumor suppressor gene CDKN2A/B on 9p21 were detected in nontransformed FL grade I-II. For this aberration as well as for -6q25 and -6q26, an association with inferior survival was observed.

  9. Aberrant expression of circulating Th17, Th1 and Tc1 cells in patients with active and inactive ulcerative colitis.

    PubMed

    Dong, Zhaogang; Du, Lutao; Xu, Xiaofei; Yang, Yongmei; Wang, Haiyan; Qu, Ailin; Qu, Xun; Wang, Chuanxin

    2013-04-01

    Ulcerative colitis (UC) is a chronic relapsing inflammatory bowel disease, yet its etiology and pathogenesis remain poorly understood. The aberrant expression of T lymphocytes plays an essential role in the progression of UC. This study aimed to evaluate the expression profile of circulating Th17, Th1 and Tc1 cells in patients with active and inactive UC. Our results revealed that the percentage of circulating Th17 cells (CD3+CD8-IL-17+) was significantly increased in patients with active UC when compared with the percentage in patients with inactive UC, Crohn's disease (CD) and healthy controls. The percentages of circulating Th1 (CD3+CD8-IFN-γ+) and Tc1 (CD3+CD8+IFN-γ+) cells were also higher in patients with active UC when compared with the percentages in patients with inactive UC and normal controls, although levels were lower than that in CD. Further analysis showed that Th17 cells were positively correlated with Th1 cells, but not with Tc1 cells. Notably, the three cells had a positive correlation with disease activity, extent of disease, detection of erythrocyte sedimentation rate and c-reactive protein in active UC. Moreover, plasma IL-17 was higher in patients with active UC, and a similar trend applied to the mRNA levels of RORγt and T-bet in peripheral blood mononuclear cells (PBMCs). The levels of p-STAT3 and p-STAT5 in PBMCs, as well as the ratio of p-STAT3/p-STAT5, were also elevated in active UC patients. Taken together, our findings revealed that elevated circulating Th17, Th1 and Tc1 cells and the aberrant activation of the STAT pathway may be implicated in the progression of UC. These findings may provide preliminary experimental clues for the development of new therapies for UC.

  10. Aberrant expression of interleukin-22 and its targeting microRNAs in oral lichen planus: a preliminary study.

    PubMed

    Shen, Zhengyu; Du, Guanhuan; Zhou, Zengtong; Liu, Wei; Shi, Linjun; Xu, Hui

    2016-08-01

    Oral lichen planus (OLP) is a T cell-mediated autoimmune disease involving oral mucosa. Interleukin-22 (IL-22) as the signature cytokine of T helper 22 cells is increasingly recognized as a key regulator in various autoimmune diseases. Our previous study reported that IL-22 immunoexpression in OLP was significantly increased compared with the normal controls. The objective of this preliminary study was to compare the IL-22 expression levels in oral biopsies from patients with OLP (n = 50) against normal oral mucosa (n = 19) using RT-qPCR and Western blot, identify the potential targeting miRNAs of IL-22, and examine the miRNA expression levels in OLP. Interleukin-22 expression level in OLP was significantly increased compared with the normal controls. The Dual-Luciferase reporter assay system in human embryonic kidney 293 (HEK293) cells demonstrated that miR-562 and miR-203 were the target miRNAs of IL-22, which was consistent with predictions from bioinformatics software analyses. Interestingly, miR-562 expression in OLP was significantly decreased, but miR-203 expression in OLP was significantly increased compared with the normal controls. This preliminary study for the first time reported that aberrant expression levels of miR-562 and miR-203 were associated with high expression of IL-22 and demonstrated the target relationship between miRNAs and IL-22 in HEK293 cells. Our data indicated that IL-22 and its targeting miRNAs contribute to the pathogenesis of OLP. Further studies are required to investigate the regulatory pathways of IL-22 and miR-562 and miR-203 in OLP. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. Aberrant DKK3 Expression in the Oral Leukoplakia and Oral Submucous Fibrosis: A Comparative Immunohistochemical Study

    PubMed Central

    Al-dhohrah, T.; Mashrah, M.; Yao, Z.; Huang, J.

    2016-01-01

    We aimed to assess and compare the expression of Dickkopf homolog 3 (DKK3), a possible tumor suppressor gene (TSG), in oral leukoplakia (OLK) and oral submucous fibrosis (OSF) using immunohistochemistry. Seventy-five cases of normal oral mucosa (NOM), OLK, OSF, and squamous cell carcinoma (OSCC) were studied. DKK3 was expressed in all cases of NOM, OLK and OSCC. There was steady increases in the percentage of the positive cells progressing toward OSCC. The expression was localized in the cytoplasm and cell membrane of cell affected by OLK with mild dysplasia and OLK with severe dysplasia. No significant association was observed between DKK3 expression and dysplastic status of OLK. Loss of DKK3 expression was observed in 15 of 30 cases in the OSF group, which was significantly associated with histological grade of OSF (P<0.0001). The percentage of positive cells gradually declined with the increasing severity of epithelial atrophy. A significant difference (P<0.01) was observed when comparing DKK3 expression among different groups of OLK and OSF cases. DKK3 may have diverse expressions in oral premalignant lesions. Loss of DKK3 expression in dysplastic/advanced stage of OSF may imply a high risk of progression to oral cancer. PMID:27349317

  12. Aberrant Calreticulin Expression in Articular Cartilage of Dio2 Deficient Mice

    PubMed Central

    Bomer, Nils; Cornelis, Frederique M. F.; Ramos, Yolande F. M.; den Hollander, Wouter; Lakenberg, Nico; van der Breggen, Ruud; Storms, Lies; Slagboom, P. Eline; Lories, Rik J. U.; Meulenbelt, Ingrid

    2016-01-01

    Objective To identify intrinsic differences in cartilage gene expression profiles between wild-type- and Dio2-/--mice, as a mechanism to investigate factors that contribute to prolonged healthy tissue homeostasis. Methods Previously generated microarray-data (Illumina MouseWG-6 v2) of knee cartilage of wild-type and Dio2 -/- -mice were re-analyzed to identify differential expressed genes independent of mechanical loading conditions by forced treadmill-running. RT-qPCR and western blot analyses of overexpression and knockdown of Calr in mouse chondro-progenitor cells (ATDC5) were applied to assess the direct effect of differential Calr expression on cartilage deposition. Results Differential expression analyses of articular cartilage of Dio2-/- (N = 9) and wild-type-mice (N = 11) while applying a cutoff threshold (P < 0.05 (FDR) and FC > |1,5|) resulted in 1 probe located in Calreticulin (Calr) that was found significantly downregulated in Dio2-/- mice (FC = -1.731; P = 0.044). Furthermore, overexpression of Calr during early chondrogenesis in ATDC5 cells leads to decreased proteoglycan deposition and corresponding lower Aggrecan expression, whereas knocking down Calr expression does not lead to histological differences of matrix composition. Conclusion We here demonstrate that the beneficial homeostatic state of articular cartilage in Dio2-/- mice is accompanied with significant lower expression of Calr. Functional analyses further showed that upregulation of Calr expression could act as an initiator of cartilage destruction. The consistent association between Calr and Dio2 expression suggests that enhanced expression of these genes facilitate detrimental effects on cartilage integrity. PMID:27163789

  13. Aberrant DNA methyltransferase expression in pancreatic ductal adenocarcinoma development and progression

    PubMed Central

    2013-01-01

    Background Altered gene methylation, regulated by DNA methyltransferases (DNMT) 1, 3a and 3b, contributes to tumorigenesis. However, the role of DNMT in pancreatic ductal adenocarcinoma (PDAC) remains unknown. Methods Expression of DNMT 1, 3a and 3b was detected in 88 Pancreatic ductal adenocarcinoma (PDAC) and 10 normal tissue samples by immunohistochemistry. Changes in cell viability, cell cycle distribution, and apoptosis of PDAC cell lines (Panc-1 and SW1990) were assessed after transfection with DNMT1 and 3b siRNA. Levels of CDKN1A, Bcl-2 and Bax mRNA were assessed by qRT-PCR, and methylation of the Bax gene promoter was assayed by methylation-specific PCR (MSP). Results DNMT1, 3a and 3b proteins were expressed in 46.6%, 23.9%, and 77.3% of PDAC tissues, respectively, but were not expressed in normal pancreatic tissues. There was a co-presence of DNMT3a and DNMT3b expression and an association of DNMT1 expression with alcohol consumption and poor overall survival. Moreover, knockdown of DNMT1 and DNMT3b expression significantly inhibited PDAC cell viability, decreased S-phase but increased G1-phase of the cell cycle, and induced apoptosis. Molecularly, expression of CDKN1A and Bax mRNA was upregulated, and the Bax gene promoter was demethylated. However, a synergistic effect of combined DNMT1 and 3b knockdown was not observed. Conclusion Expression of DNMT1, 3a and 3b proteins is increased in PDAC tissues, and DNMT1 expression is associated with poor prognosis of patients. Knockdown of DNMT1 and 3b expression arrests tumor cells at the G1 phase of the cell cycle and induces apoptosis. The data suggest that DNMT knockdown may be a novel treatment strategy for PDAC. PMID:24423239

  14. Aberrant EphB/ephrin-B expression in experimental gastric lesions and tumor cells

    PubMed Central

    Uchiyama, Shintaro; Saeki, Noritaka; Ogawa, Kazushige

    2015-01-01

    AIM: To determine whether the expression profiles of EphB receptor and ephrin-B ligand can be used as markers for dysplastic/oncogenic transformation in gastric mucosa. METHODS: The protein expression and localization of EphB and ephrin-B in normal, ulcerated regenerating, and dysplastic gastric mucosa were examined in a rat experimental model by immunolabeling, and mRNA expression was assessed in four human gastric carcinoma cell lines by reverse transcription-polymerase chain reaction. RESULTS: Ephrin-B- and EphB-expressing regions were divided along the pit-gland axis in normal gastric units. EphB2 was transiently upregulated in the experimental ulcer, and its expression domain extended to gastric pits and/or the luminal surface where ephrin-B-expressing pit cells reside. EphB2, B3, and B4 and ephrin-B1 were coexpressed in the experimental gastric dysplasia, and more than one ligand-receptor pair was highly expressed in each of the gastric carcinoma cell lines. CONCLUSION: Robust and stable coexpression of EphB and ephrin-B is a feature common to experimentally induced gastric dysplasia and human gastric carcinoma cell lines as compared to normal gastric and ulcerated regenerating epithelia. Thus, EphB/ephrin-B may be a useful marker combination for dysplastic/oncogenic transformation in gastric cancer. PMID:25593460

  15. Aberrant expression of tumor-associated carbohydrate antigen Globo H in thyroid carcinoma.

    PubMed

    Cheng, Shih-Ping; Yang, Po-Sheng; Chien, Ming-Nan; Chen, Ming-Jen; Lee, Jie-Jen; Liu, Chien-Liang

    2016-12-01

    The induction of tumor-associated carbohydrate antigen results from altered glycosylation in transformed cells. Globo H is a hexasaccharide glycosphingolipid overexpressed on malignancies of epithelial origin and has become an attractive vaccine target. We aimed to investigate the expression patterns and prognostic value of Globo H in thyroid neoplasms. Globo H expression was examined by immunohistochemical analysis using commercial and in-house tissue microarrays. The expression was correlated with clinicopathologic characteristics in papillary thyroid cancer. Normal or benign thyroid lesions were negative for Globo H expression. Globo H was positive in 33% medullary, 24% papillary, 11% undifferentiated, and 8% follicular thyroid cancer. Globo H expression in papillary thyroid cancer was associated with extrathyroidal invasion (P = 0.017), BRAF mutation (P = 0.010), AMES high risk (P = 0.045), and increased ATA risk of recurrence (P = 0.022). Globo H is specifically expressed in a subset of thyroid malignancies. In papillary thyroid cancer, Globo H expression is associated with invasiveness and BRAF mutation. Immunotherapy targeting Globo H may have potential applications in thyroid cancer. J. Surg. Oncol. 2016;114:853-858. © 2016 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Developmental aberrations of liver gene expression in bovine fetuses derived from somatic cell nuclear transplantation.

    PubMed

    Herath, Chandana B; Ishiwata, Hiroko; Shiojima, Satoshi; Kadowaki, Tadashi; Katsuma, Susumu; Ushizawa, Koichi; Imai, Kei; Takahashi, Toru; Hirasawa, Akira; Takahashi, Seiya; Izaike, Yoshiaki; Tsujimoto, Gozoh; Hashizume, Kazuyoshi

    2006-01-01

    Cloning by somatic cell nuclear transfer (NT) has been accomplished. However, the process itself is inefficient since most clones die before birth and survivors often display various anomalies. In an effort to determine global expression profiles of developmentally regulated liver genes in NT bovine fetuses, we employed a custom-made bovine liver complementary DNA (cDNA) microarray. The NT fetuses in early pregnancy were derived from cumulus cells as the nuclear donor cells. Normal fetuses were derived from in vitro fertilization (IVF) and artificial insemination (AI). Gene expression levels in NT, IVF, and AI fetal livers were obtained by comparing individual fetal liver samples with that of adult liver of nonpregnant cycling cows. Statistical analyses of the expression data showed widespread dysregulation of developmentally important genes in the three NT fetuses examined. It was found that the number of dysregulated genes was within a range of 3.5-7.7% of the tested genes in the NT fetal livers. The analyses revealed that one NT fetus was markedly different in liver gene expression profile from the other two NT fetal livers in which the expression profiles were highly correlated. Thus, our findings demonstrate that widespread dysregulation of liver genes occurs in the developing liver of NT bovine fetuses. It is possible that inappropriate genomic reprogramming after NT is a key factor associated with abnormal gene expressions in the livers of NT fetuses, whereas distinct expression patterns between the fellow cloned fetuses likely have resulted from variable epigenetic status of the donor nuclei.

  17. Aberrant expression of N-methylpurine-DNA glycosylase influences patient survival in malignant gliomas.

    PubMed

    Liu, Ce; Tu, Yanyang; Yuan, Jun; Mao, Xinggang; He, Shiming; Wang, Liang; Fu, Guoqiang; Zong, Jianhai; Zhang, Yongsheng

    2012-01-01

    To examine the expression of N-methylpurine-DNA glycosylase (MPG) gene and protein in glioma samples with different WHO grades and its association with patients' survival. Immunohistochemistry assay, quantitative real-time PCR and Western blot analysis were carried out to investigate the expression of MPG gene and protein in 128 glioma and 10 non-neoplastic brain tissues. MPG gene expression level in glioma tissues was significantly higher than that in non-neoplastic brain tissues (P < 0.001). Immunohistochemistry also showed that MPG protein was over-expressed in glioma tissues, which was consistent with the resutls of Western blot analysis. Additionally, the expression levels of MPG gene and protein both increase from grade I to grade IV glioma according to the results of real-time PCR, immunohistochemistry and western blot analysis. Moreover, the survival rate of MPG-positive patients was significantly lower than that of MPG-negative patients (P < 0.001). We further confirmed that the over-expression of MPG was a significant and independent prognostic indicator in glioma by multivariate analysis (P < 0.001). Our data showed the over-expression of MPG gene and protein in human gliomas, and also suggested for the first time that MPG be an unfavorable independent prognostic indicator for glioma patients.

  18. Aberrant Expression of N-Methylpurine-DNA Glycosylase Influences Patient Survival in Malignant Gliomas

    PubMed Central

    Liu, Ce; Tu, Yanyang; Yuan, Jun; Mao, Xinggang; He, Shiming; Wang, Liang; Fu, Guoqiang; Zong, Jianhai; Zhang, Yongsheng

    2012-01-01

    Aim. To examine the expression of N-methylpurine-DNA glycosylase (MPG) gene and protein in glioma samples with different WHO grades and its association with patients' survival. Methods. Immunohistochemistry assay, quantitative real-time PCR and Western blot analysis were carried out to investigate the expression of MPG gene and protein in 128 glioma and 10 non-neoplastic brain tissues. Results. MPG gene expression level in glioma tissues was significantly higher than that in non-neoplastic brain tissues (P < 0.001). Immunohistochemistry also showed that MPG protein was over-expressed in glioma tissues, which was consistent with the resutls of Western blot analysis. Additionally, the expression levels of MPG gene and protein both increase from grade I to grade IV glioma according to the results of real-time PCR, immunohistochemistry and western blot analysis. Moreover, the survival rate of MPG-positive patients was significantly lower than that of MPG-negative patients (P < 0.001). We further confirmed that the over-expression of MPG was a significant and independent prognostic indicator in glioma by multivariate analysis (P < 0.001). Conclusions. Our data showed the over-expression of MPG gene and protein in human gliomas, and also suggested for the first time that MPG be an unfavorable independent prognostic indicator for glioma patients. PMID:22496614

  19. Aberrant large tumor suppressor 2 (LATS2) gene expression correlates with EGFR mutation and survival in lung adenocarcinomas

    PubMed Central

    Luo, Susan Y.; Sit, Ko-Yung; Sihoe, Alan D.L.; Suen, Wai-Sing; Au, Wing-Kuk; Tang, Ximing; Ma, Edmond S.K.; Chan, Wai-Kong; Wistuba, Ignacio I.; Minna, John D.; Tsao, George S.W.; Lam, David C.L.

    2015-01-01

    Background Large tumor suppressor 2 (LATS2) gene is a putative tumor suppressor gene with potential roles in regulation of cell proliferation and apoptosis in lung cancer. The aim of this study is to explore the association of aberrant LATS2 expression with EGFR mutation and survival in lung adenocarcinoma (AD), and the effects of LATS2 silencing in both lung AD cell lines. Methods LATS2 mRNA and protein expression in resected lung AD were correlated with demographic characteristics, EGFR mutation and survival. LATS2-specific siRNA was transfected into four EGFR wild-type (WT) and three EGFR mutant AD cell lines and the changes in LATS2 expression and relevant signaling molecules before and after LATS2 knockdown were assayed. Results Fifty resected lung AD were included (M:F = 23:27, smokers:non-smokers = 19:31, EGFR mutant:wild-type = 21:29) with LATS2 mRNA levels showed no significant difference between gender, age, smoking and pathological stages while LATS2 immunohistochemical staining on an independent set of 79 lung AD showed similar trend. LATS2 mRNA level was found to be a significant independent predictor for survival status (disease-free survival RR = 0.217; p = 0.003; Overall survival RR = 0.238; p = 0.036). siRNA-mediated suppression of LATS2 expression resulted in augmentation of ERK phosphorylation in EGFR wild-type AD cell lines with high basal LATS2 expression, discriminatory modulation of Akt signaling between EGFR wild-type and mutant cells, and induction of p53 accumulation in AD cell lines with low baseline p53 levels. Conclusions LATS2 expression level is predictive of survival in patients with resected lung AD. LATS2 may modulate and contribute to tumor growth via different signaling pathways in EGFR mutant and wild-type tumors. PMID:24976335

  20. Aberrant MHC class II expression in mouse joints leads to arthritis with extraarticular manifestations similar to rheumatoid arthritis

    PubMed Central

    Kanazawa, Satoshi; Ota, Shusuke; Sekine, Chiyoko; Tada, Toyohiro; Otsuka, Takanobu; Okamoto, Takashi; Sønderstrup, Grete; Peterlin, B. Matija

    2006-01-01

    Genetic susceptibility to rheumatoid arthritis (RA) is associated with certain MHC class II molecules. To clarify the role of these determinants in RA, we generated the D1CC transgenic mouse that expressed genes involved in antigen processing and presentation by the MHC class II pathway in joints. The class II transactivator, which was transcribed from the rat collagen type II promoter and enhancer, directed the expression of these genes. In D1CC mice congenic for the H-2q (DBA/1) background, small amounts of bovine collagen type II in adjuvant induced reproducibly an inflammatory arthritis resembling RA. Importantly, these stimuli had no effect in DBA/1 mice. Eighty-nine percent of D1CC mice developed chronic disease with joint swelling, redness, and heat in association with synovial proliferation as well as pannus formation and mononuclear infiltration of synovial membranes. Granulomatous lesions resembling rheumatoid nodules and interstitial pneumonitis also were observed. As in patients with RA, anticyclic citrullinated peptide antibodies were detected during the inflammatory stage. Finally, joints in D1CC mice displayed juxtaarticular demineralization, severe joint space narrowing, and erosions, which led to ankylosis, but without the appearance of osteophytes. Thus, aberrant expression of MHC class II in joints facilitates the development of severe erosive inflammatory polyarthritis, which is very similar to RA. PMID:16980409

  1. Aberrant expression of the neuronal-specific protein DCDC2 promotes malignant phenotypes and is associated with prostate cancer progression.

    PubMed

    Longoni, N; Kunderfranco, P; Pellini, S; Albino, D; Mello-Grand, M; Pinton, S; D'Ambrosio, G; Sarti, M; Sessa, F; Chiorino, G; Catapano, C V; Carbone, G M

    2013-05-02

    By integrating gene profiling and immunohistochemical data with functional experiments in cell lines in this study we show for the first time that doublecortin (DCX) domain containing 2 (DCDC2), a protein belonging to the DCX family and involved in neuronal cell migration, is aberrantly expressed in prostate tumors whereas absent in normal prostate. Furthermore, in patients treated with radical prostatectomy, high levels of DCDC2 RNA were significantly associated with increased biochemical relapse (LogRank Mantel-Cox=0.012). Mechanistically, we found that the ETS transcription factor ESE3/EHF, which is expressed in normal prostate and frequently lost in prostate tumors, maintained DCDC2 repressed by binding to a novel identified ETS binding site in the gene promoter. Consistently, in prostate tumors and in cellular models of gain and loss of ESE3/EHF, the expression of DCDC2 and ESE3/EHF were inversely correlated. In prostate cancer cells, DCDC2 colocalized with microtubules and promoted cell migration and resistance to the microtubule-targeting drug taxol. Collectively, this study establishes DCDC2 as a novel ESE3/EHF oncogenic target in prostate cancer. These findings may be relevant for the clinical management of prostate cancer as DCDC2 may signal tumors more prone to relapse and resistant to taxol treatment.

  2. Clinicopathological and prognostic significance of aberrant Arpin expression in gastric cancer

    PubMed Central

    Li, Tao; Zheng, Hong-Mei; Deng, Nai-Mei; Jiang, Ying-Jian; Wang, Jiang; Zhang, Dian-Liang

    2017-01-01

    AIM To detect the expression of Arpin, and determine its correlation with clinicopathological characteristics and the prognosis of gastric cancer (GC) patients. METHODS A total of 176 GC patients were enrolled as study subjects and classified into groups according to different clinicopathological variables. GC mucosal tissues were obtained via surgery. Another 43 paraffin-embedded tissue blocks of normal gastric epithelium (> 5 cm away from the edge of the tumor) were included in the control group. Immunohistochemistry (IHC) for the Arpin and Arp3 proteins was performed on the formalin-fixed, paraffin-embedded GC tissues. Additionally, expression of the Arpin protein in 43 normal gastric tissues was also determined using IHC. RESULTS Expression of the Arpin protein in GC was lower than that in normal gastric mucosa (30.68% vs 60.47%, P < 0.001). A χ2 test of the 176 GC samples used for IHC showed that decreased Arpin expression was associated with advanced TNM stage (P < 0.01) and the presence or absence of lymph node metastasis (80.92% vs 35.56%, P < 0.001). Additionally, a significant correlation was observed between the expression of Arpin and the presence of the Arp2/3 complex in GC tissues (χ2 = 30.535, P < 0.001). Moreover, a multivariate Cox regression analysis revealed that Arpin expression [hazard ratio (HR) = 0.551, P = 0.029] and TNM stage (HR = 5.344, P = 0.001) were independent prognostic markers for overall survival of GC patients. Regarding the 3-year disease-free survival (DFS), the recurrence rate of GC patients with low Arpin expression levels (median DFS 19 mo) was higher than that in the high-Arpin-expression group (median DFS 34 mo, P = 0.022). CONCLUSION Low Arpin levels are associated with clinicopathological variables and a poor prognosis in GC patients. Arpin may be regarded as a potential prognostic indicator in GC. PMID:28293092

  3. Aberrant Gene Expression Profile of Unaffected Colon Mucosa from Patients with Unifocal Colon Polyp

    PubMed Central

    Lian, Jingjing; Ma, Lili; Yang, Jiayin; Xu, Lili

    2015-01-01

    Background The aim of this study was to evaluate gene expression profiles in unaffected colon mucosa and polyp tissue from patients with unifocal colon polyp to investigate the potential mucosa impairment in normal-appearing colon mucosa from these patients. Material/Methods Colon polyp patients were prospectively recruited. We obtained colon biopsies from the normal-appearing sites and polyp tissue through colonoscopy. Gene expression analysis was performed using microarrays. Gene ontology and clustering were evaluated by bioinformatics. Results We detected a total of 711 genes (274 up-regulated and 437 down-regulated) in polyp tissue and 256 genes (170 up-regulated and 86 down-regulated) in normal-appearing colon mucosa, with at least a 3-fold of change compared to healthy controls. Heatmapping of the gene expression showed similar gene alteration patterns between unaffected colon mucosa and polyp tissue. Gene ontology analyses confirmed the overlapped molecular functions and pathways of altered gene expression between unaffected colon mucosa and polyp tissue from patients with unifocal colon polyp. The most significantly altered genes in normal-appearing tissues in polyp patients include immune response, external side of plasma membrane, nucleus, and cellular response to zinc ion. Conclusions Significant gene expression alterations exist in unaffected colon mucosa from patients with unifocal colon polyp. Unaffected colon mucosa and polyp tissue share great similarity and overlapping of altered gene expression profiles, indicating the potential possibility of recurrence of colon polyps due to underlying molecular abnormalities of colon mucosa in these patients. PMID:26675397

  4. Aberrant expression pattern and location of cullin 1 are associated with the development of papillary carcinoma in thyroid and cyclin D1 expression.

    PubMed

    Do, Sung-Im; Kim, Kyungeun; Lee, Hyunjoo; Kim, Hyun-Soo; Do, Tae Gu; Yun, Jisup; Kim, Dong-Hoon; Chae, Seoung Wan; Park, Yong Lai; Park, Chan Heun; Sohn, Jin Hee; Min, Kyueng-Whan; Pyo, Jung-Soo

    2014-09-01

    Cullin 1 (Cul1) is a rigid scaffold protein of a major class of E3 ubiquitin ligase, also known as the Skp1/cullin1/F-box (SCF) complex, which is involved in cell-cycle progression. The aberrant expression of Cul1 is involved in the dysfunction of SCF E3 ligase. Previous studies have revealed an association between increased Cul1 expression and tumor progression and poor outcome in several different tumors. We constructed a tissue microarray containing 103 papillary carcinoma tissues of the thyroid and 66 normal thyroid tissues. Cul1 expression and Cyclin D1 expression were evaluated by immunohistochemistry staining, and the relationship between their expression and clinicopathological parameters were analyzed. Cytoplasmic expression of Cul1 was correlated with tumor occurrence (p < 0.001), N stage (p = 0.027), and Cyclin D1 expression (p < 0.001). Cyclin D1 expression showed a correlation with tumor occurrence (p < 0.001) and T stage (p = 0.009). On the other hand, nuclear expression of Cul1 showed a negative correlation with tumor occurrence (p < 0.001) and Cyclin D1 expression (p < 0.001). Cytoplasmic Cul1 expression was associated with tumor development and higher nodal metastasis status, supporting the idea that the SCF complex is involved in cell-cycle regulation and promotes cell proliferation. Nuclear expression of Cul1 showed inverse relationship between tumor aggressiveness factors. Our data suggest that the expression site of Cul1 may affect the function of the SFC complex and play a role in tumor progression.

  5. Chemokine CXCL12 uses CXCR4 and a signaling core formed by bifunctional Akt, extracellular signal-regulated kinase (ERK)1/2, and mammalian target of rapamycin complex 1 (mTORC1) proteins to control chemotaxis and survival simultaneously in mature dendritic cells.

    PubMed

    Delgado-Martín, Cristina; Escribano, Cristina; Pablos, José Luis; Riol-Blanco, Lorena; Rodríguez-Fernández, José Luis

    2011-10-28

    Chemokines control several cell functions in addition to chemotaxis. Although much information is available on the involvement of specific signaling molecules in the control of single functions controlled by chemokines, especially chemotaxis, the mechanisms used by these ligands to regulate several cell functions simultaneously are completely unknown. Mature dendritic cells (maDCs) migrate through the afferent lymphatic vessels to the lymph nodes, where they regulate the initiation of the immune response. As maDCs are exposed to chemokine CXCL12 (receptors CXCR4 and CXCR7) during their migration, its functions are amenable to be regulated by this ligand. We have used maDCs as a model system to analyze the mechanisms whereby CXCL12 simultaneously controls chemotaxis and survival in maDCs. We show that CXCL12 uses CXCR4, but not CXCR7, and the components of a signaling core that includes G(i)/Gβγ, PI3K-α/-δ/-γ, Akt, ERK1/2 and mammalian target of rapamycin complex 1 (mTORC1), which organize hierarchically to control both functions. Downstream of Akt, Forkhead box class O (FOXO) regulates CXCL12-dependent survival, but not chemotaxis, suggesting that downstream of the aforementioned signaling core, additional signaling molecules may control more selectively CXCL12-dependent chemotaxis or survival. Finally, the data obtained also show that CXCR4 uses a signaling signature that is different from that used by CCR7 to control similar functions.

  6. Aberration of miRNAs Expression in Leukocytes from Sporadic Amyotrophic Lateral Sclerosis

    PubMed Central

    Chen, YongPing; Wei, QianQian; Chen, XuePing; Li, ChunYu; Cao, Bei; Ou, RuWei; Hadano, Shinji; Shang, Hui-Fang

    2016-01-01

    Background: Accumulating evidence indicates that miRNAs play an important role in the development of amyotrophic lateral sclerosis (ALS). Most of previous studies on miRNA dysregulation in ALS focused on the alterative expression in ALS animal model or in limited samples from European patients with ALS. In the present study, the miRNA expression profiles were investigated in Chinese ALS patients to explore leukocytes miRNAs as a potential biomarker for the diagnosis of ALS. Methods: We analyzed the expression profiles of 1733 human mature miRNAs using microarray technology in leukocytes obtained from 5 patients with sporadic ALS (SALS) and 5 healthy controls. An independent group of 83 SALS patients, 24 Parkinson's disease (PD) patients and 61 controls was used for validation by real-time polymerase chain reaction assay. Area under the receiver operating characteristic curve (AUC) was used to evaluate diagnostic accuracy. In addition, target genes and signaling information of validated differential expression miRNAs were predicted using Bioinformatics. Results: Eleven miRNAs, including four over-expressed and seven under-expressed miRNAs detected in SALS patients compared to healthy controls were selected for validation. Four under-expressed microRNAs, including hsa-miR-183, hsa-miR-193b, hsa-miR-451, and hsa-miR-3935, were confirmed in validation stage by comparison of 83 SALS patients and 61 HCs. Moreover, we identified a miRNA panel (hsa-miR-183, hsa-miR-193b, hsa-miR-451, and hsa-miR-3935) having a high diagnostic accuracy of SALS (AUC 0.857 for the validation group). However, only hsa-miR-183 was significantly lower in SALS patients than that in PD patients and in HCs, while no differences were found between PD patients and HCs. By bioinformatics analysis, we obtained a large number of target genes and signaling information that are linked to neurodegeneration. Conclusion: This study provided evidence of abnormal miRNA expression patterns in the peripheral

  7. Aberrant gene expression profile in a mouse model of endometriosis mirrors that observed in women

    PubMed Central

    Pelch, Katherine E.; Schroder, Amy L.; Kimball, Paul A.; Sharpe-Timms, Kathy L.; Davis, J. W.; Nagel, Susan C.

    2010-01-01

    Objective To define the altered gene expression profile of endometriotic lesions in a mouse model of surgically-induced endometriosis Design Autologous experimental mouse model. Setting Medical school department. Animals Adult C57Bl6 mice. Intervention(s) Endometriosis was surgically-induced by auto-transplantation of uterine tissue to the intestinal mesentery. Endometriotic lesions and eutopic uteri were recovered at 3 or 29 days post-induction. Main Outcome Measure(s) Altered gene expression was measured in the endometriotic lesion relative to the eutopic uterus by genome wide cDNA microarray analysis and was confirmed by real time RT-PCR for six genes. Relevant categories of altered genes were identified using gene ontology analysis to determine groups of genes enriched for altered expression. Result(s) The expression of 479 and 114 genes was altered in the endometriotic lesion compared to the eutopic uterus at 3 or 29 days post-induction, respectively. Gene ontology enrichment analysis revealed that genes associated with the extracellular matrix, cell adhesions, immune function, cell growth, and angiogenesis were altered in the endometriotic lesion compared to the eutopic uterus. Conclusion(s) Based on gene expression analysis, the mouse model of surgically-induced endometriosis appears to be a good model for studying the pathophysiology and treatment of endometriosis. PMID:19473656

  8. Heterogeneous nuclear ribonucleoprotein K is over expressed, aberrantly localised and is associated with poor prognosis in colorectal cancer.

    PubMed

    Carpenter, B; McKay, M; Dundas, S R; Lawrie, L C; Telfer, C; Murray, G I

    2006-10-09

    Heterogeneous ribonucleoprotein K (hnRNP K) is a member of the hnRNP family which has several different cellular roles including transcription, mRNA shuttling, RNA editing and translation. Several reports implicate hnRNP K having a role in tumorigenesis, for instance hnRNP K increases transcription of the oncogene c-myc and hnRNP K expression is regulated by the p53/MDM 2 pathway. In this study comparing normal colon to colorectal cancer by proteomics, hnRNP K was identified as being overexpressed in this type of cancer. Immunohistochemistry with a monoclonal antibody to hnRNP K (which we developed) on colorectal cancer tissue microarray, confirmed that hnRNP K was overexpressed in colorectal cancer (P<0.001) and also showed that hnRNP K had an aberrant subcellular localisation in cancer cells. In normal colon hnRNP K was exclusively nuclear whereas in colorectal cancer the protein localised both in the cytoplasm and the nucleus. There were significant increases in both nuclear (P=0.007) and cytoplasmic (P=0.001) expression of hnRNP K in Dukes C tumours compared with early stage tumours. In Dukes C patient's good survival was associated with increased hnRNP K nuclear expression (P=0.0093). To elaborate on the recent observation that hnRNP K is regulated by p53, the expression profiles of these two proteins were also analysed. There was no correlation between hnRNP K and p53 expression, however, patients who presented tumours that were positive for hnRNP K and p53 had a poorer survival outcome (P=0.045).

  9. Aberrant cytoplasmic expression of the p16 protein in breast cancer is associated with accelerated tumour proliferation.

    PubMed Central

    Emig, R.; Magener, A.; Ehemann, V.; Meyer, A.; Stilgenbauer, F.; Volkmann, M.; Wallwiener, D.; Sinn, H. P.

    1998-01-01

    The p16 protein plays an important role in the transition of cells into the G1 phase of the cell cycle. We have studied the prevalence of p16 protein expression in breast carcinomas in a prospective series of 368 invasive and 52 non-invasive malignancies, as well as in 88 locally recurring tumours and three tumour cell lines. p16 protein expression was evaluated immunohistochemically on paraffin sections using monoclonal and polyclonal anti-p16 antibodies, and by immunoblotting of tumour cell suspensions. Tumour cell lines were also subjected to polymerase chain reaction-single strand polymorphism (PCR-SSCP) analysis and direct DNA sequencing. The results were compared with established prognostic parameters, DNA flow cytometry and p53 protein expression. In 33 (9%) invasive and two (4%) intraductal carcinomas, a cytoplasmic accumulation of the p16 protein was seen. These cases were characterized by poor histological grade of differentiation, loss of of oestrogen receptors and progesterone receptors and frequent overexpression of the p53 protein. In addition, breast carcinomas with aberrant p16 expression demonstrated a high proliferative activity, with median S-phase fractions 74% higher than in the control group and the median Ki67 fractions elevated to 75%. A genetic alteration of the p16 gene was not detectable in three analysed cell lines with cytoplasmic p16 expression applying PCR-SSCP and direct DNA sequencing. These results indicate that cytoplasmic accumulation of the p16 protein identifies a subset of highly malignant breast carcinomas with accelerated tumour proliferation and other unfavourable parameters in breast cancer. The described protein accumulation is apparently not caused by an alteration of the p16 gene. Images Figure 1 Figure 4 PMID:9862580

  10. Caveolin-3 is aberrantly expressed in skeletal muscle cells in myasthenia gravis.

    PubMed

    Iwasa, Kazuo; Furukawa, Yutaka; Yoshikawa, Hiroaki; Yamada, Masahito

    2016-12-15

    Caveolin-3 is a muscle-specific membrane protein that localizes to the sarcolemma and T-tubule system. Caveolin-3 is needed for muscle repair and skeletal muscle development. The objective of this study was to compare caveolin-3 expression in myasthenia gravis (MG) and control muscles. Caveolin-3 was abnormally expressed in the MG muscle membrane, with partial loss of expression and overexpression in 5/15 and 10/15 patients, respectively. Caveolin-3 mRNA and protein levels were higher in MG than in control muscles, suggesting that partial deficiency of caveolin-3 is occasionally detected in MG muscle and that caveolin-3 overexpression may be required after MG muscle damage. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Aberrant Expression of proPTPRN2 in Cancer Cells Confers Resistance to Apoptosis.

    PubMed

    Sorokin, Alexey V; Nair, Binoj C; Wei, Yongkun; Aziz, Kathryn E; Evdokimova, Valentina; Hung, Mien-Chie; Chen, Junjie

    2015-05-01

    The protein tyrosine phosphatase receptor PTPRN2 is expressed predominantly in endocrine and neuronal cells, where it functions in exocytosis. We found that its immature isoform proPTPRN2 is overexpressed in various cancers, including breast cancer. High proPTPRN2 expression was associated strongly with lymph node-positive breast cancer and poor clinical outcome. Loss of proPTPRN2 in breast cancer cells promoted apoptosis and blocked tumor formation in mice, whereas enforced expression of proPTPRN2 in nontransformed human mammary epithelial cells exerted a converse effect. Mechanistic investigations suggested that ProPTPRN2 elicited these effects through direct interaction with TRAF2, a hub scaffold protein for multiple kinase cascades, including ones that activate NF-κB. Overall, our results suggest PTPRN2 as a novel candidate biomarker and therapeutic target in breast cancer. ©2015 American Association for Cancer Research.

  12. Aberrant expression of NF-κB in liver fluke associated cholangiocarcinoma: implications for targeted therapy.

    PubMed

    Seubwai, Wunchana; Wongkham, Chaisiri; Puapairoj, Anucha; Khuntikeo, Narong; Pugkhem, Ake; Hahnvajanawong, Chariya; Chaiyagool, Jariya; Umezawa, Kazuo; Okada, Seiji; Wongkham, Sopit

    2014-01-01

    Up-regulation and association of nuclear factor kappa B (NF-κB) with carcinogenesis and tumor progression has been reported in several malignancies. In the current study, expression of NF-κB in cholangiocarcinoma (CCA) patient tissues and its clinical significance were determined. The possibility of using NF-κB as the therapeutic target of CCA was demonstrated. Expression of NF-κB in CCA patient tissues was determined using immunohistochemistry. Dehydroxymethylepoxyquinomicin (DHMEQ), a specific NF-κB inhibitor, was used to inhibit NF-κB action. Cell growth was determined using an MTT assay, and cell apoptosis was shown by DNA fragmentation, flow cytometry and immunocytofluorescent staining. Effects of DHMEQ on growth and apoptosis were demonstrated in CCA cell lines and CCA-inoculated mice. DHMEQ-induced apoptosis in patient tissues using a histoculture drug response assay was quantified by TUNEL assay. Normal bile duct epithelia rarely expressed NF-κB (subunits p50, p52 and p65), whereas all CCA patient tissues (n  =  48) over-expressed all NF-κB subunits. Inhibiting NF-κB action by DHMEQ significantly inhibited growth of human CCA cell lines in a dose- and time-dependent manner. DHMEQ increased cell apoptosis by decreasing the anti-apoptotic protein expressions-Bcl-2, XIAP-and activating caspase pathway. DHMEQ effectively reduced tumor size in CCA-inoculated mice and induced cell apoptosis in primary histocultures of CCA patient tissues. NF-κB was over-expressed in CCA tissues. Inhibition of NF-κB action significantly reduced cell growth and enhanced cell apoptosis. This study highlights NF-κB as a molecular target for CCA therapy.

  13. Targeting aberrant expression of Notch-1 in ALDH(+) cancer stem cells in breast cancer.

    PubMed

    Pal, Deeksha; Kolluru, Venkatesh; Chandrasekaran, Balaji; Baby, Becca V; Aman, Masarath; Suman, Suman; Sirimulla, Suman; Sanders, Mary Ann; Alatassi, Houda; Ankem, Murali K; Damodaran, Chendil

    2017-03-01

    We have previously reported that high aldehyde dehydrogenase (ALDH) enzyme activity in breast cancer cells results in breast cancer stem cell (BCSC) properties by upregualting Notch-1 and epithelial mesenchymal markers. This results in chemoresistance in breast cancer. Here, we examined the functional and clinical significance of ALDH expression by measuring the ALDH levels in breast cancer tissues by immunohistochemistry. There was a significantly higher ALDH expression in higher grade breast cancer tumor tissues (Grade- II and III) versus normal breast tissues. Injection of BCSC (ALDH(+) and CD44(+) /CD22(-) ) cells resulted in aggressive tumor growth in athymic mice versus ALDH(-) cells. The ALDH(+) and CD44(+) /CD22(-) tumors grow rapidly and are larger than ALDH(-) tumors which were slow growing and smaller. Molecularly, ALDH(+) tumors expressed higher expression of Notch-1 and EMT markers than ALDH(-) tumors. Oral administration of the naturally occurring Psoralidin (Pso, 25 mg/kg of body weight) significantly inhibited the growth in ALDH(+) and ALDH(-) tumors as well. Psoralidin inhibited Notch-1 mediated EMT activation in ALDH(+) and ALDH(-) tumors-this confirms our in vitro findings. Our results suggest that Notch-1 could be an attractive target and inhibition of Notch-1 by Psoralidin may prevent pathogenesis of breast cancer as well as metastasis. © 2016 Wiley Periodicals, Inc.

  14. Hyperacetylation in prostate cancer induces cell cycle aberrations, chromatin reorganization and altered gene expression profiles

    PubMed Central

    Watson, Jenny A; McKenna, Declan J; Maxwell, Perry; Diamond, James; Arthur, Ken; McKelvey-Martin, Valerie J; Hamilton, Peter W

    2010-01-01

    Abstract Histone acetylation is a fundamental mechanism in the regulation of local chromatin conformation and gene expression. Research has focused on the impact of altered epigenetic environments on the expression of specific genes and their pathways. However, changes in histone acetylation also have a global impact on the cell. In this study we used digital texture analysis to assess global chromatin patterns following treatment with trichostatin A (TSA) and have observed significant alterations in the condensation and distribution of higher-order chromatin, which were associated with altered gene expression profiles in both immortalised normal PNT1A prostate cell line and androgen-dependent prostate cancer cell line LNCaP. Furthermore, the extent of TSA-induced disruption was both cell cycle and cell line dependent. This was illustrated by the identification of sub-populations of prostate cancer cells expressing high levels of H3K9 acetylation in the G2/M phase of the cell cycle that were absent in normal cell populations. In addition, the analysis of enriched populations of G1 cells showed a global decondensation of chromatin exclusively in normal cells. PMID:19583812

  15. Aberrant TIMELESS expression is associated with poor clinical survival and lymph node metastasis in early-stage cervical carcinoma.

    PubMed

    Zhang, Weijing; He, Weiling; Shi, Yongjie; Zhao, Jing; Liu, Sailan; Zhang, Fengxiang; Yang, Jiarui; Xie, Chuanmiao; Zhang, Yanna

    2017-01-01

    TIMELESS is a highly conserved protein required for the maintenance of normal mammalian circadian oscillations and for controlling cellular metabolism and proliferation. Recently, TIMELESS was implicated in the tumorigenesis of certain cancers. However, little is known on TIMELESS protein expression and its potential as a prognostic factor in cervical cancer. Here, we investigate TIMELESS expression pattern and its clinicopathological significance in early-stage cervical carcinoma. TIMELESS mRNA and protein expression was evaluated by real-time PCR and western blot analysis in cervical cancer cell lines, a normal cervical cell line, as well as in six pairs of surgically removed cervical cancer and adjacent normal cervical tissues. A total of 189 paraffin-embedded cervical carcinoma specimens were detected and diagnosed by immunohistochemistry (IHC), and the clinical significance of TIMELESS expression was further analyzed. Aberrant TIMELESS mRNA and protein expression were demonstrated in cervical cancer cell lines compared with the normal cervical cell line. TIMELESS mRNA and protein expression were significantly increased in cervical cancer specimens compared with adjacent non-cancerous cervical specimens. TIMELESS protein expression was significantly associated with the age (P=0.011), clinical stage (P<0.001), pelvic lymph node metastasis (P<0.001), squamous cell carcinoma antigen (P=0.003), tumor recurrence (P=0.015), vital status (P<0.001), tumor differentiation grade (P<0.001), property of the surgical margin (P=0.036) and lymphovascular space involvement (P=0.001). Patients with increased TIMELESS protein expression showed strong tendencies to receive postoperative radiotherapy (P=0.002). Upregulation of TIMELESS correlated with poorer overall survival (OS) and disease-free survival (DFS). Univariate and multivariate Cox-regression analyses showed that TIMELESS can be regarded as an independent predictive biomarker for poor clinical outcome for early

  16. Aberrant expression of the neuronal transcription factor FOXP2 in neoplastic plasma cells.

    PubMed

    Campbell, Andrew J; Lyne, Linden; Brown, Philip J; Launchbury, Rosalind J; Bignone, Paola; Chi, Jianxiang; Roncador, Giovanna; Lawrie, Charles H; Gatter, Kevin C; Kusec, Rajko; Banham, Alison H

    2010-04-01

    FOXP2 mutation causes a severe inherited speech and language defect, while the related transcription factors FOXP1, FOXP3 and FOXP4 are implicated in cancer. FOXP2 mRNA and protein expression were characterised in normal human tissues, haematological cell lines and multiple myeloma (MM) patients' samples. FOXP2 mRNA and protein were absent in mononuclear cells from different anatomical sites, lineages and stages of differentiation. However, FOXP2 mRNA and protein was detected in several lymphoma (8/20) and all MM-derived cell lines (n = 4). FOXP2 mRNA was expressed in bone marrow samples from 96% of MM patients (24/25), 66.7% of patients with the pre-neoplastic plasma cell proliferation monoclonal gammopathy of undetermined significance (MGUS) (6/9), but not in reactive plasma cells. The frequency of FOXP2 protein expression in CD138(+) plasma cells was significantly higher in MGUS (P = 0.0005; mean 46.4%) and MM patients (P < or = 0.0001; mean 57.3%) than in reactive marrows (mean 2.5%). FOXP2 (>10% nuclear positivity) was detectable in 90.2% of MM (55/61) and 90.9% of MGUS (10/11) patients, showing more frequent expression than CD56 and labelling 75% of CD56-negative MM (9/12). FOXP2 represents the first transcription factor whose expression consistently differentiates normal and abnormal plasma cells and FOXP2 target genes are implicated in MM pathogenesis.

  17. Aberrant Cosmc genes result in Tn antigen expression in human colorectal carcinoma cell line HT-29

    PubMed Central

    Yu, Xiaofeng; Du, Zhenzhen; Sun, Xuhong; Shi, Chuanqin; Zhang, Huaixiang; Hu, Tao

    2015-01-01

    The Tn antigen, which arises from mutation in the Cosmc gene is one of the most common tumor associated carbohydrate antigens. Cosmc resides in X24 encoded by a single gene and functions as a specific molecular chaperone for T-synthase. While the Tn antigen cannot be detected in normal cells, Cosmc mutations inactivate T-synthase and consequently result in Tn antigen expression within certain cancers. In addition to this Cosmc mutation-induced expression, the Tn antigen is also expressed in such cell lines as Jurkat T, LSC and LS174T. Whether the Cosmc mutation is present in the colon cancer cell line HT-29 is still unclear. Here, we isolate HT-29-Tn+ cells from HT-29 cells derived from a female colon cancer patient. These HT-29-Tn+ cells show a loss of the Cosmc gene coding sequence (CDS) leading to an absence of T-synthase activity and Tn antigen expression. Additionally, almost no methylation of Cosmc CpG islands was detected in HT-29-Tn+ as well as in HT-29-Tn- and Tn- tumor cells from male patients. In contrast, the methylation frequency of CpG island of Cosmc in normal female cells was ~50%. Only one active allele of Cosmc existed in HT-29-Tn+ and HT-29-Tn- cells as based upon detection of SNP sites. These results indicate that Tn antigens expression and T-synthase inactivity in HT-29-Tn+ cells can be related to the absence of CDS in Cosmc active alleles, while an inactive allele deletion of Cosmc in HT-29 cells has no influence on Cosmc function. PMID:26045765

  18. Aberrant expression of miR-9 in benign and malignant breast tumors.

    PubMed

    Hasanzadeh, Aisan; Mesrian Tanha, Hamzeh; Ghaedi, Kamran; Madani, Mahboobeh

    2016-10-01

    MicroRNAs (miRNAs) are involved in the progression of breast cancer (BC). miR-9 has been reported to be correlated with either favorable or unfavorable events in BC. This study was aimed to evaluate the expression level of miR-9 in human breast tissues, including benign and malignant tumor samples and also healthy tissue. The expression level of miR-9 was analyzed in 10 normal breast tissues, 30 malignant, and 30 benign breast tumor tissue samples using RT-PCR and qPCR. In addition, bioinformatics assessment upon miR-9 functionality in BC cells was performed. The miR-9 expression level was downregulated in tumor tissues, including benign and malignant compared to the healthy tissue was observed (P value, < 0.0001; fold change, -1.37). In addition, miR-9 expression level was reduced in benign tumors compared with malignant tumors (P value, < 0.0001; fold change, -1.35). Moreover, according to the AUCs (area under curve) of receiver operating characteristic (ROC) curves, miR-9 showed significant capability for distinguishing benign from healthy, malignant from healthy, benign from malignant, and tumor from health tissues. Furthermore, pathways in cancer, p53 signaling pathway, and focal adhesion were manifested by computational analysis as miR-9 related signaling pathways which have logical association with experimental observations. In conclusion, downregulation of miR-9 in benign tumors vs healthy tissue and its overexpression in malignant tumors vs benign tumors suggest paradoxical functionality for this miRNA. Our results shed additional information on controversial expression pattern of miR-9 depending on different progression level of BC. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Passenger mutations and aberrant gene expression in congenic tissue plasminogen activator-deficient mouse strains

    PubMed Central

    Szabo, R.; Samson, A. L.; Lawrence, D. A.; Medcalf, R. L.; Bugge, T. H.

    2017-01-01

    Summary Background The ability to generate defined null mutations in mice revolutionized the analysis of gene function in mammals. However, gene-deficient mice generated by using 129-derived embryonic stem cells may carry large segments of 129 DNA, even when extensively backcrossed to reference strains, such as C57BL/6J, and this may confound interpretation of experiments performed in these mice. Tissue plasminogen activator (tPA), encoded by the PLAT gene, is a fibrinolytic serine protease that is widely expressed in the brain. A number of neurological abnormalities have been reported in tPA-deficient mice. Objectives To study genetic contamination of tPA-deficient mice. Materials and methods Whole genome expression array analysis, RNAseq expression profiling, low- and high-density SNP analysis, bioinformatics, and genome editing was used to analyze gene expression in tPA-deficient mouse brains. Results and conclusions Genes differentially expressed in the brain of Plat−/− mice from two independent colonies highly backcrossed onto the C57BL/6J strain clustered near Plat on chromosome 8. SNP analysis attributed this anomaly to about 20 Mbp of DNA flanking Plat being of 129 origin in both strains. Bioinformatic analysis of these 129-derived chromosomal segments identified a significant number of mutations in genes co-segregating with the targeted Plat allele, including several potential null mutations. Using zinc finger nuclease technology, we generated novel “passenger mutation”-free isogenic C57BL/6J-Plat−/− and FVB/NJ-Plat−/− mouse strains by introducing an 11 bp deletion in the exon encoding the signal peptide. These novel mouse strains will be a useful community resource for further exploration of tPA function in physiological and pathological processes. PMID:27079292

  20. Aberrant Expression of NF-κB in Liver Fluke Associated Cholangiocarcinoma: Implications for Targeted Therapy

    PubMed Central

    Seubwai, Wunchana; Wongkham, Chaisiri; Puapairoj, Anucha; Khuntikeo, Narong; Pugkhem, Ake; Hahnvajanawong, Chariya; Chaiyagool, Jariya; Umezawa, Kazuo; Okada, Seiji; Wongkham, Sopit

    2014-01-01

    Background Up-regulation and association of nuclear factor kappa B (NF-κB) with carcinogenesis and tumor progression has been reported in several malignancies. In the current study, expression of NF-κB in cholangiocarcinoma (CCA) patient tissues and its clinical significance were determined. The possibility of using NF-κB as the therapeutic target of CCA was demonstrated. Methodology Expression of NF-κB in CCA patient tissues was determined using immunohistochemistry. Dehydroxymethylepoxyquinomicin (DHMEQ), a specific NF-κB inhibitor, was used to inhibit NF-κB action. Cell growth was determined using an MTT assay, and cell apoptosis was shown by DNA fragmentation, flow cytometry and immunocytofluorescent staining. Effects of DHMEQ on growth and apoptosis were demonstrated in CCA cell lines and CCA-inoculated mice. DHMEQ-induced apoptosis in patient tissues using a histoculture drug response assay was quantified by TUNEL assay. Principal Findings Normal bile duct epithelia rarely expressed NF-κB (subunits p50, p52 and p65), whereas all CCA patient tissues (n  =  48) over-expressed all NF-κB subunits. Inhibiting NF-κB action by DHMEQ significantly inhibited growth of human CCA cell lines in a dose- and time-dependent manner. DHMEQ increased cell apoptosis by decreasing the anti-apoptotic protein expressions–Bcl-2, XIAP–and activating caspase pathway. DHMEQ effectively reduced tumor size in CCA-inoculated mice and induced cell apoptosis in primary histocultures of CCA patient tissues. Conclusions NF-κB was over-expressed in CCA tissues. Inhibition of NF-κB action significantly reduced cell growth and enhanced cell apoptosis. This study highlights NF-κB as a molecular target for CCA therapy. PMID:25170898

  1. Benzene-Induced Aberrant miRNA Expression Profile in Hematopoietic Progenitor Cells in C57BL/6 Mice.

    PubMed

    Wei, Haiyan; Zhang, Juan; Tan, Kehong; Sun, Rongli; Yin, Lihong; Pu, Yuepu

    2015-11-12

    Benzene is a common environmental pollutant that causes hematological alterations. MicroRNAs (miRNAs) may play a role in benzene-induced hematotoxicity. In this study, C57BL/6 mice showed significant hematotoxicity after exposure to 150 mg/kg benzene for 4 weeks. Benzene exposure decreased not only the number of cells in peripheral blood but also hematopoietic progenitor cells in the bone marrow. Meanwhile, RNA from Lin(-) cells sorted from the bone marrow was applied to aberrant miRNA expression profile using Illumina sequencing. We found that 5 miRNAs were overexpressed and 45 miRNAs were downregulated in the benzene exposure group. Sequencing results were confirmed through qRT-PCR. Furthermore, we also identified five miRNAs which significantly altered in Lin(-)c-Kit⁺ cells obtained from benzene-exposed mice, including mmu-miR-34a-5p; mmu-miR-342-3p; mmu-miR-100-5p; mmu-miR-181a-5p; and mmu-miR-196b-5p. In summary, we successfully established a classical animal model to induce significant hematotoxicity by benzene injection. Benzene exposure may cause severe hematotoxicity not only to blood cells in peripheral circulation but also to hematopoietic cells in bone marrow. Benzene exposure also alters miRNA expression in hematopoietic progenitor cells. This study suggests that benzene induces alteration in hematopoiesis and hematopoiesis-associated miRNAs.

  2. CD34⁺/CD38⁻ acute myelogenous leukemia cells aberrantly express Aurora kinase A.

    PubMed

    Yang, Jing; Ikezoe, Takayuki; Nishioka, Chie; Nobumoto, Atsuya; Udaka, Keiko; Yokoyama, Akihito

    2013-12-01

    We previously showed that Aurora kinase A (AURKA) is aberrantly expressed in acute myelogenous leukemia (AML) cells when compared to bone marrow mononuclear cells isolated from healthy volunteers. We have also shown that CD34(+) /CD38(-) AML cells, one of compartments enriched for leukemia stem cells in most leukemia subgroups, were relatively resistant to cytarabine-mediated growth inhibition when compared to their CD34(+) /CD38(+) counterparts. Our study attempted to identify therapeutic targets in CD34(+) /CD38(-) AML cells and found that CD34(+) /CD38(-) AML cells isolated from patients (n = 26) expressed larger amounts of AURKA than their CD34(+) /CD38(+) counterparts and CD34(+) normal hematopoietic stem/progenitor cells isolated from healthy volunteers (n = 6), as measured by real-time reverse-transcriptase polymerase chain reaction. Blockade of AURKA by the specific inhibitor MLN8237 or a short hairpin RNA (shRNA) against AURKA significantly inhibited proliferation, impaired self-renewal capability and induced apoptosis of CD34(+) /CD38(-) AML cells, in association with modulation of levels of Bcl-2 family member proteins. Importantly, inhibition of AURKA in CD34(+) /CD38(-) AML cells by MLN8237 or an shRNA significantly impaired engraftment of these cells in severely immunocompromised mice and appeared to prolong their survival. These results suggest that AURKA is a promising molecular target to eliminate chemotherapy-resistant CD34(+) /CD38(-) AML cells.

  3. Subgroup J avian leukosis virus infection of chicken dendritic cells induces apoptosis via the aberrant expression of microRNAs.

    PubMed

    Liu, Di; Dai, Manman; Zhang, Xu; Cao, Weisheng; Liao, Ming

    2016-02-01

    Subgroup J avian leukosis virus (ALV-J) is an oncogenic retrovirus that causes immunosuppression and enhances susceptibility to secondary infection. The innate immune system is the first line of defense in preventing bacterial and viral infections, and dendritic cells (DCs) play important roles in innate immunity. Because bone marrow is an organ that is susceptible to ALV-J, the virus may influence the generation of bone marrow-derived DCs. In this study, DCs cultured in vitro were used to investigate the effects of ALV infection. The results revealed that ALV-J could infect these cells during the early stages of differentiation, and infection of DCs with ALV-J resulted in apoptosis. miRNA sequencing data of uninfected and infected DCs revealed 122 differentially expressed miRNAs, with 115 demonstrating upregulation after ALV-J infection and the other 7 showing significant downregulation. The miRNAs that exhibited the highest levels of upregulation may suppress nutrient processing and metabolic function. These results indicated that ALV-J infection of chicken DCs could induce apoptosis via aberrant microRNA expression. These results provide a solid foundation for the further study of epigenetic influences on ALV-J-induced immunosuppression.

  4. Mice expressing aberrant sperm-specific protein PMIS2 produce normal-looking but fertilization-incompetent spermatozoa.

    PubMed

    Yamaguchi, Ryo; Fujihara, Yoshitaka; Ikawa, Masahito; Okabe, Masaru

    2012-07-01

    Eight kinds of gene-disrupted mice (Clgn, Calr3, Pdilt, Tpst2, Ace, Adam1a, Adam2, and Adam3) show impaired sperm transition into the oviducts and defective sperm binding to the zona pellucida. All of these knockout strains are reported to lack or show aberrant expression of a disintegrin and metallopeptidase domain 3 (ADAM3) on the sperm membrane. We performed proteomic analyses of the proteins of these infertile spermatozoa to clarify whether the abnormal function is caused exclusively by a deficiency in ADAM3 expression. Two proteins, named PMIS1 and PMIS2, were missing in spermatozoa from Clgn-disrupted mice. To study their roles, we generated two gene-disrupted mouse lines. Pmis1-knockout mice were fertile, but Pmis2-knockout males were sterile because of a failure of sperm transport into the oviducts. Pmis2-deficient spermatozoa also failed to bind to the zona pellucida. However, they showed normal fertilizing ability when eggs surrounded with cumulus cells were used for in vitro fertilization. Further analysis revealed that these spermatozoa lacked the ADAM3 protein, but the amount of PMIS2 was also severely reduced in Adam3-deficient spermatozoa. These results suggest that PMIS2 might function both as the ultimate factor regulating sperm transport into the oviducts and in modulating sperm-zona binding.

  5. Aberrant AML1 gene expression in the diagnosis of childhood leukemias not characterized by AML1-involved cytogenetic abnormalities.

    PubMed

    Adamaki, Maria; Vlahopoulos, Spiros; Lambrou, George I; Papavassiliou, Athanasios G; Moschovi, Maria

    2017-03-01

    The AML1 ( acute myeloid leukemia 1) gene, a necessary prerequisite of embryonic hematopoiesis and a critical regulator of normal hematopoietic development, is one of the most frequently mutated genes in human leukemia, involving over 50 chromosome translocations and over 20 partner genes. In the few existing studies investigating AML1 gene expression in childhood leukemias, aberrant upregulation seems to specifically associate with AML1 translocations and amplifications. The aim of this study was to determine whether overexpression also extends to other leukemic subtypes than the ones karyotypically involving AML1. We use quantitative real-time polymerase chain reaction methodology to investigate gene expression in 100 children with acute leukemias and compare them to those of healthy controls. We show that in childhood acute lymphoblastic leukemia, AML1 gene overexpression is associated with a variety of leukemic subtypes, both immunophenotypically and cytogenetically. Statistically significantly higher transcripts of the gene were detected in the acute lymphoblastic leukemia group as compared to the acute myeloid leukemia group, where AML1 overexpression appeared to associate with cytogenetic abnormalities additional to those that engage the AML1 gene, or that are reported as showing a "normal" karyotype. Collectively, our study shows that AML1 gene overexpression characterizes a broader range of leukemic subtypes than previously thought, including various maturation stages of B-cell acute lymphoblastic leukemia and cytogenetic types additional to those involving the AML1 gene.

  6. Benzene-Induced Aberrant miRNA Expression Profile in Hematopoietic Progenitor Cells in C57BL/6 Mice

    PubMed Central

    Wei, Haiyan; Zhang, Juan; Tan, Kehong; Sun, Rongli; Yin, Lihong; Pu, Yuepu

    2015-01-01

    Benzene is a common environmental pollutant that causes hematological alterations. MicroRNAs (miRNAs) may play a role in benzene-induced hematotoxicity. In this study, C57BL/6 mice showed significant hematotoxicity after exposure to 150 mg/kg benzene for 4 weeks. Benzene exposure decreased not only the number of cells in peripheral blood but also hematopoietic progenitor cells in the bone marrow. Meanwhile, RNA from Lin− cells sorted from the bone marrow was applied to aberrant miRNA expression profile using Illumina sequencing. We found that 5 miRNAs were overexpressed and 45 miRNAs were downregulated in the benzene exposure group. Sequencing results were confirmed through qRT-PCR. Furthermore, we also identified five miRNAs which significantly altered in Lin−c-Kit+ cells obtained from benzene-exposed mice, including mmu-miR-34a-5p; mmu-miR-342-3p; mmu-miR-100-5p; mmu-miR-181a-5p; and mmu-miR-196b-5p. In summary, we successfully established a classical animal model to induce significant hematotoxicity by benzene injection. Benzene exposure may cause severe hematotoxicity not only to blood cells in peripheral circulation but also to hematopoietic cells in bone marrow. Benzene exposure also alters miRNA expression in hematopoietic progenitor cells. This study suggests that benzene induces alteration in hematopoiesis and hematopoiesis-associated miRNAs. PMID:26569237

  7. Aberrant alternative splicing and extracellular matrix gene expression in mouse models of myotonic dystrophy

    PubMed Central

    Du, Hongqing; Cline, Melissa S.; Osborne, Robert J.; Tuttle, Daniel L.; Clark, Tyson A.; Donohue, John Paul; Hall, Megan P.; Shiue, Lily; Swanson, Maurice S.; Thornton, Charles A.; Ares, Manuel

    2009-01-01

    Myotonic dystrophy (DM1) is associated with expression of expanded CTG DNA repeats as RNA (CUGexp RNA). To test whether CUGexp RNA creates a global splicing defect, we compared skeletal muscle of two mouse DM1 models, one expressing a CTGexp transgene, and another homozygous for a defective Mbnl1 gene. Strong correlation in splicing changes for ~100 new Mbnl1-regulated exons indicates loss of Mbnl1 explains >80% of the splicing pathology due to CUGexp RNA. In contrast, only about half of mRNA level changes can be attributed to loss of Mbnl1, indicating CUGexp RNA has Mbnl1-independent effects, particularly on mRNAs for extracellular matrix (ECM) proteins. We propose that CUGexp RNA causes two separate effects: loss of Mbnl1 function, disrupting splicing, and loss of another function that disrupts ECM mRNA regulation, possibly mediated by MBNL2. These findings reveal unanticipated similarities between DM1 and other muscular dystrophies. PMID:20098426

  8. Identification of aberrant tRNA-halves expression patterns in clear cell renal cell carcinoma

    PubMed Central

    Nientiedt, Malin; Deng, Mario; Schmidt, Doris; Perner, Sven; Müller, Stefan C.; Ellinger, Jörg

    2016-01-01

    Small non-coding RNAs (sncRNA; <200 nt) regulate various cellular processes and modify gene expression. Under nutritional, biological or physiochemical stress some mature sncRNAs (e.g. tRNAs) are cleaved into halves (30–50 nt) and smaller fragments (18–22 nt); the significance and functional role of these tRNA fragments is unknown, but their existence has been linked to carcinogenesis. We used small RNA sequencing to determine the expression of sncRNAs. Subsequently the findings were validated for miR-122-5p, miR-142-3p and 5'tRNA4-Val-AAC using qPCR. We identified differential expression of 132 miRNAs (upregulated: 61, downregulated: 71) and 32 tRNAs (upregulated: 13, downregulated: 19). Read length analysis showed that miRNAs mapped in the 20–24 nt fraction, whereas tRNA reads mapped in the 30–36 nt fraction instead the expected size of 73–95 nt thereby indicating cleavage of tRNAs. Overexpression of miR-122-5p and miR-142-3p as well as downregulation of 5'tRNA4-Val-AAC was validated in an independent cohort of 118 ccRCC and 74 normal renal tissues. Furthermore, staging and grading was inversely correlated with the 5'tRNA4-Val-AAC expression. Serum levels of miR-122-5p, miR-142-3p and 5'tRNA4-Val-AAC did not differ in ccRCC and control subjects. In conclusion, 5′ cleavage of tRNAs occurs in ccRCC, but the exact functional implication of tRNA-halve deregulation remains to be clarified. PMID:27883021

  9. Aberrant microRNA expression in Chinese patients with chronic lymphocytic leukemia.

    PubMed

    Zhu, Dan-Xia; Miao, Kou-Rong; Fang, Cheng; Fan, Lei; Zhu, Wei; Zhu, Hua-Yuan; Zhuang, Yun; Hong, Ming; Liu, Peng; Xu, Wei; Li, Jian-Yong

    2011-06-01

    MicroRNAs (miRNAs) are a class of small endogenous RNAs that play important regulatory roles by targeting mRNAs for cleavage or translational repression. Many reports have indicated that miRNAs play a critical role in malignancies, and regulations in the progression of leukemia. However, the miRNAs expression level in Chinese patients with chronic lymphocytic leukemia (CLL), and its prognostic value remain elusive. We identified various degrees of down-regulation of miR-15a, miR-16-1, miR-29b, miR-181a and miR-181b in CLL mononuclear cells. Moreover, we have identified miR-29b and miR-181a/b expression significantly correlated with IGHV mutational status. Transcript levels of predicted target genes BCL-2 and TCL-1 were also determined, and the expression levels were significantly upregulated in CLL patients compared with normal controls (P<0.001). Higher expression of TCL-1 was significantly correlated with aggressive disease features. In addition, an inverse correlation was observed in the CLL samples we examined between miRNAs (miR-16-1, miR-181a, miR-181b) and BCL-2 level; furthermore, an inverse correlation was also observed between miRNAs (miR-16-1, miR-181a, miR-181b) and TCL-1, which suggest that these miRNAs may implicate in negatively regulating target mRNA at transcriptional level. These different miRNAs may play an important role in the pathogenesis of CLL and might be applied for the assessment of prognosis in patients with CLL.

  10. Loss of keratin K2 expression causes aberrant aggregation of K10, hyperkeratosis, and inflammation.

    PubMed

    Fischer, Heinz; Langbein, Lutz; Reichelt, Julia; Praetzel-Wunder, Silke; Buchberger, Maria; Ghannadan, Minoo; Tschachler, Erwin; Eckhart, Leopold

    2014-10-01

    Keratin K2 is one of the most abundant structural proteins of the epidermis; however, its biological significance has remained elusive. Here we show that suprabasal type II keratins, K1 and K2, are expressed in a mutually exclusive manner at different body sites of the mouse, with K2 being confined to the ear, sole, and tail skin. Deletion of K2 caused acanthosis and hyperkeratosis of the ear and the tail epidermis, corneocyte fragility, increased transepidermal water loss, and local inflammation in the ear skin. The loss of K2 was partially compensated by upregulation of K1 expression. However, a significant portion of K2-deficient suprabasal keratinocytes lacked a regular cytoskeleton and developed massive aggregates of the type I keratin, K10. Aggregate formation, but not hyperkeratosis, was suppressed by the deletion of both K2 and K10, whereas deletion of K10 alone caused clumping of K2 in ear skin. Taken together, this study demonstrates that K2 is a necessary and sufficient binding partner of K10 at distinct body sites of the mouse and that unbalanced expression of these keratins results in aggregate formation.

  11. Local and transient gene expression primes the liver to resist cancer metastasis.

    PubMed

    Goodwin, Tyler J; Zhou, Yingqiu; Musetti, Sara N; Liu, Rihe; Huang, Leaf

    2016-11-09

    The liver is the primary site of metastasis for gastrointestinal cancers and is a location highly susceptible to the establishment of metastasis in numerous other primary cancers, including breast, lung, and pancreatic cancers. The current standard of care typically consists of primary tumor resection and systemic administration of potent but toxic chemotherapeutics, yielding a minimal improvement in the median survival rate. CXCL12, a chemokine, is a key factor for activating the migration/survival pathways of CXCR4(+) cancer cells and for recruiting immunosuppressive cells to areas of inflammation. Therefore, reducing CXCL12 concentrations within the liver has the potential to decrease tumor and immunosuppressive cell activation/migration within the liver. However, because of off-target toxicities associated with systemic administration of anti-CXCL12 therapies, transient and liver-specific expression of a CXCL12 trap is necessary. To address this challenge, we developed a lipid calcium phosphate nanoparticle optimized for delivering plasmid DNA, encoding an engineered CXCL12 protein trap, to the nucleus of liver hepatocytes. This pCXCL12-trap formulation yielded transient (4 days) liver-specific expression, which greatly decreased the occurrence of liver metastasis in two aggressive liver metastasis models, including colorectal [CT-26(FL3)] and breast (4T1) cancers. Subsequent studies in an aggressive human colorectal liver metastasis model (HT-29) decreased the establishment of liver metastasis more effectively than did systemic administration of the CXCL12 protein trap and to a level comparable to a high-dose regimen of a potent CXCR4 antagonist (AMD3100). Copyright © 2016, American Association for the Advancement of Science.

  12. Reduced cortical BDNF expression and aberrant memory in Carf knockout mice

    PubMed Central

    McDowell, Kelli A.; Hutchinson, Ashley N.; Wong-Goodrich, Sarah J.E.; Presby, Matthew M.; Su, Dan; Rodriguiz, Ramona M.; Law, Krystal C.; Williams, Christina L.; Wetsel, William C.; West, Anne E.

    2010-01-01

    Transcription factors are a key point of convergence between the cell-intrinsic and extracellular signals that guide synaptic development and brain plasticity. Calcium-Response Factor (CaRF) is a unique transcription factor first identified as a binding protein for a calcium-response element in the gene encoding Brain-Derived Neurotrophic Factor (Bdnf). We have now generated Carf knockout (KO) mice to characterize the function of this factor in vivo. Intriguingly, Carf KO mice have selectively reduced expression of Bdnf exon IV-containing mRNA transcripts and BDNF protein in the cerebral cortex while BDNF levels in the hippocampus and striatum remain unchanged, implicating CaRF as a brain region-selective regulator of BDNF expression. At the cellular level, Carf KO mice show altered expression of GABAergic proteins at striatal synapses, raising the possibility that CaRF may contribute to aspects of inhibitory synapse development. Carf KO mice show normal spatial learning in the Morris water maze and normal context-dependent fear conditioning. However they have an enhanced ability to find a new platform location on the first day of reversal training in the water maze and they extinguish conditioned fear more slowly than their wildtype (WT) littermates. Finally, Carf KO mice show normal short-term and long-term memory in a novel object recognition task, but exhibit impairments during the remote memory phase of testing. Taken together these data reveal novel roles for CaRF in the organization and/or function of neural circuits that underlie essential aspects of learning and memory. PMID:20519520

  13. Reduced cortical BDNF expression and aberrant memory in Carf knock-out mice.

    PubMed

    McDowell, Kelli A; Hutchinson, Ashley N; Wong-Goodrich, Sarah J E; Presby, Matthew M; Su, Dan; Rodriguiz, Ramona M; Law, Krystal C; Williams, Christina L; Wetsel, William C; West, Anne E

    2010-06-02

    Transcription factors are a key point of convergence between the cell-intrinsic and extracellular signals that guide synaptic development and brain plasticity. Calcium-response factor (CaRF) is a unique transcription factor first identified as a binding protein for a calcium-response element in the gene encoding brain-derived neurotrophic factor (Bdnf). We have now generated Carf knock-out (KO) mice to characterize the function of this factor in vivo. Intriguingly, Carf KO mice have selectively reduced expression of Bdnf exon IV-containing mRNA transcripts and BDNF protein in the cerebral cortex, whereas BDNF levels in the hippocampus and striatum remain unchanged, implicating CaRF as a brain region-selective regulator of BDNF expression. At the cellular level, Carf KO mice show altered expression of GABAergic proteins at striatal synapses, raising the possibility that CaRF may contribute to aspects of inhibitory synapse development. Carf KO mice show normal spatial learning in the Morris water maze and normal context-dependent fear conditioning. However they have an enhanced ability to find a new platform location on the first day of reversal training in the water maze and they extinguish conditioned fear more slowly than their wild-type littermates. Finally, Carf KO mice show normal short-term (STM) and long-term memory (LTM) in a novel object recognition task, but exhibit impairments during the remote memory phase of testing. Together, these data reveal novel roles for CaRF in the organization and/or function of neural circuits that underlie essential aspects of learning and memory.

  14. An amelogenin mutation leads to disruption of the odontogenic apparatus and aberrant expression of Notch I

    PubMed Central

    Chen, Xu; Li, Yong; Alawi, Faizan; Bouchard, Jessica R.; Kulkarni, Ashok B.; Gibson, Carolyn W.

    2012-01-01

    BACKGROUND Amelogenins are highly conserved proteins secreted by ameloblasts in the dental organ of developing teeth. These proteins regulate dental enamel thickness and structure in humans and mice. Mice that express an amelogenin transgene with a P70T mutation (TgP70T) develop abnormal epithelial proliferation in an amelogenin null (KO) background. Some of these cellular masses have the appearance of proliferating stratum intermedium, which is the layer adjacent to the ameloblasts in unerupted teeth. As Notch proteins are thought to constitute the developmental switch that separates ameloblasts from stratum intermedium, these signaling proteins were evaluated in normal and proliferating tissues. METHODS Mandibles were dissected for histology and immunohistochemistry using Notch I antibodies. Molar teeth were dissected for western blotting and RT-PCR for evaluation of Notch levels through imaging and statistical analyses. RESULTS Notch I was immunolocalized to ameloblasts of TgP70TKO mice, KO ameloblasts stained, but less strongly, and wild-type teeth had minimal staining. Cells within the proliferating epithelial cell masses were positive for Notch I and had an appearance reminiscent of calcifying epithelial odontogenic tumor with amyloid-like deposits. Notch I protein and mRNA were elevated in molar teeth from TgP70TKO mice. CONCLUSION Expression of TgP70T leads to abnormal structures in mandibles and maxillae of mice with the KO genetic background and these mice have elevated levels of Notch I in developing molars. As cells within the masses also express transgenic amelogenins, development of the abnormal proliferations suggests communication between amelogenin producing cells and the proliferating cells, dependent on the presence of the mutated amelogenin protein. PMID:20923441

  15. Aberrant Expression of Long Non-Coding RNAs in Schizophrenia Patients

    PubMed Central

    Chen, Shengdong; Sun, Xinyang; Niu, Wei; Kong, Lingming; He, Mingjun; Li, Wanshuai; Zhong, Aifang; Lu, Jim; Zhang, Liyi

    2016-01-01

    Background Dysfunction of long non-coding RNAs (lncRNAs) has been demonstrated to be involved in psychiatric diseases. However, the expression patterns and functions of the regulatory lncRNAs in schizophrenia (SZ) patients have rarely been systematically reported. Material/Methods The lncRNAs in peripheral blood mononuclear cells (PBMCs) were screened and compared between the SZ patients and demographically-matched healthy controls using microarray analysis, and then were validated by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) method. Three verified significantly dysregulated lncRNAs of PBMCs were selected and then measured in SZ patients before and after the antipsychotic treatment. SZ symptomatology improvement was measured by Positive And Negative Syndrome Scale (PANSS) scores. Results One hundred and twenty-five lncRNAs were significantly differentially expressed in SZ patients compared with healthy controls, of which 62 were up-regulated and 63 were down-regulated. Concurrent with the significant decrease of the PANSS scores of patients after the treatment, the PBMC levels of lncRNA NONHSAT089447 and NONHSAT041499 were strikingly decreased (P<0.05). Down-regulation of PBMC expression of NONHSAT041499 was significantly correlated to the improvement of positive and activity symptoms of patients (r=−0.444 and −0.423, respectively, P<0.05, accounting for 16.9% and 15.1%, respectively), and was also significantly associated with better outcomes (odds ratio 2.325 for positive symptom and 12.340 for activity symptom). Conclusions LncRNA NONHSAT089447 and NONHSAT041499 might be involved in the pathogenesis and development of SZ, and the PBMC level of NONHSAT041499 is significantly associated with the treatment outcomes of SZ. PMID:27650396

  16. Dysregulation of the epigenome in triple-negative breast cancers: basal-like and claudin-low breast cancers express aberrant DNA hypermethylation.

    PubMed

    Roll, J Devon; Rivenbark, Ashley G; Sandhu, Rupninder; Parker, Joel S; Jones, Wendell D; Carey, Lisa A; Livasy, Chad A; Coleman, William B

    2013-12-01

    A subset of human breast cancer cell lines exhibits aberrant DNA hypermethylation that is characterized by hyperactivity of the DNA methyltransferase enzymes, overexpression of DNMT3b, and concurrent methylation-dependent silencing of numerous epigenetic biomarker genes. The objective of this study was to determine if this aberrant DNA hypermethylation (i) is found in primary breast cancers, (ii) is associated with specific breast cancer molecular subtypes, and (iii) influences patient outcomes. Analysis of epigenetic biomarker genes (CDH1, CEACAM6, CST6, ESR1, GNA11, MUC1, MYB, SCNN1A, and TFF3) identified a gene expression signature characterized by reduced expression levels or loss of expression among a cohort of primary breast cancers. The breast cancers that express this gene expression signature are enriched for triple-negative subtypes - basal-like and claudin-low breast cancers. Methylation analysis of primary breast cancers showed extensive promoter hypermethylation of epigenetic biomarker genes among triple-negative breast cancers, compared to other breast cancer subclasses where promoter hypermethylation events were less frequent. Furthermore, triple-negative breast cancers either did not express or expressed significantly reduced levels of protein corresponding to methylation-sensitive biomarker gene products. Together, these findings suggest strongly that loss of epigenetic biomarker gene expression is frequently associated with gene promoter hypermethylation events. We propose that aberrant DNA hypermethylation is a common characteristic of triple-negative breast cancers and may represent a fundamental biological property of basal-like and claudin-low breast cancers. Kaplan-Meier analysis of relapse-free survival revealed a survival disadvantage for patients with breast cancers that exhibit aberrant DNA hypermethylation. Identification of this distinguishing trait among triple-negative breast cancers forms the basis for development of new rational

  17. Aberrant Expression of TIMP-2 and PBEF Genes in the Placentae of Cloned Mice Due to Epigenetic Reprogramming Error

    PubMed Central

    Kim, Hong Rye; Lee, Jae Eun; Oqani, Reza Kheirkhahi; Kim, So Yeon; Wakayama, Teruhiko; Li, Chong; Sa, Su Jin; Woo, Je Seok; Jin, Dong Il

    2016-01-01

    Cloned mice derived from somatic or ES cells show placental overgrowth (placentomegaly) at term. We had previously analyzed cloned and normal mouse placentae by using two-dimensional gel electrophoresis and mass spectrometry to identify differential protein expression patterns. Cloned placentae showed upregulation of tissue inhibitor of metalloproteinase-2 (TIMP-2), which is involved in extracellular matrix degradation and tissue remodeling, and downregulation of pre-B cell colony enhancing factor 1 (PBEF), which inhibits apoptosis and induces spontaneous labor. Here, we used Western blotting to further analyze the protein expression levels of TIMP-2 and PBEF in cloned placentae derived from cumulus cells, TSA-treated cumulus cells, intracytoplasmic sperm injection (ICSI), and natural mating (NM control). Cloned and TSA-treated cloned placentae had higher expression levels of TIMP-2 compared with NM control and ICSI-derived placentae, and there was a positive association between TIMP-2 expression and the placental weight of cloned mouse concepti. Conversely, PBEF protein expression was significantly lower in cloned and ICSI placentae compared to NM controls. To examine whether the observed differences were due to abnormal gene expression caused by faulty epigenetic reprogramming in clones, we investigated DNA methylation and histone modification in the promoter regions of the genes encoding TIMP-2 and PBEF. Sodium bisulfite sequencing did not reveal any difference in DNA methylation between cloned and NM control placentae. However, ChIP assays revealed that the level of H3-K9/K14 acetylation at the TIMP-2 locus was higher in cloned placentae than in NM controls, whereas acetylation of the PBEF promoter was lower in cloned and ICSI placenta versus NM controls. These results suggest that cloned placentae appear to suffer from failure of histone modification-based reprogramming in these (and potentially other) developmentally important genes, leading to aberrant

  18. CXCR7 Controls Competition for Recruitment of β-Arrestin 2 in Cells Expressing Both CXCR4 and CXCR7

    PubMed Central

    Coggins, Nathaniel L.; Trakimas, Danielle; Chang, S. Laura; Ehrlich, Anna; Ray, Paramita; Luker, Kathryn E.

    2014-01-01

    Chemokine CXCL12 promotes growth and metastasis of more than 20 different human cancers, as well as pathogenesis of other common diseases. CXCL12 binds two different receptors, CXCR4 and CXCR7, both of which recruit and signal through the cytosolic adapter protein β-arrestin 2. Differences in CXCL12-dependent recruitment of β-arrestin 2 in cells expressing one or both receptors remain poorly defined. To quantitatively investigate parameters controlling association of β-arrestin 2 with CXCR4 or CXCR7 in cells co-expressing both receptors, we used a systems biology approach combining real-time, multi-spectral luciferase complementation imaging with computational modeling. Cells expressing only CXCR4 maintain low basal association with β-arrestin 2, and CXCL12 induces a rapid, transient increase in this interaction. In contrast, cells expressing only CXCR7 have higher basal association with β-arrestin 2 and exhibit more gradual, prolonged recruitment of β-arrestin 2 in response to CXCL12. We developed and fit a data-driven computational model for association of either CXCR4 or CXCR7 with β-arrestin 2 in cells expressing only one type of receptor. We then experimentally validated model predictions that co-expression of CXCR4 and CXCR7 on the same cell substantially decreases both the magnitude and duration of CXCL12-regulated recruitment of β-arrestin 2 to CXCR4. Co-expression of both receptors on the same cell only minimally alters recruitment of β-arrestin 2 to CXCR7. In silico experiments also identified β-arrestin 2 as a limiting factor in cells expressing both receptors, establishing that CXCR7 wins the “competition” with CXCR4 for CXCL12 and recruitment of β-arrestin 2. These results reveal how competition for β-arrestin 2 controls integrated responses to CXCL12 in cells expressing both CXCR4 and CXCR7. These results advance understanding of normal and pathologic functions of CXCL12, which is critical for developing effective strategies to target

  19. Aberrant splicing and expression of the non muscle myosin heavy-chain gene MYH14 in DM1 muscle tissues.

    PubMed

    Rinaldi, F; Terracciano, C; Pisani, V; Massa, R; Loro, E; Vergani, L; Di Girolamo, S; Angelini, C; Gourdon, G; Novelli, G; Botta, A

    2012-01-01

    Myotonic dystrophy type 1 (DM1) is a complex multisystemic disorder caused by an expansion of a CTG repeat located at the 3' untranslated region (UTR) of DMPK on chromosome 19q13.3. Aberrant messenger RNA (mRNA) splicing of several genes has been reported to explain some of the symptoms of DM1 including insulin resistance, muscle wasting and myotonia. In this paper we analyzed the expression of the MYH14 mRNA and protein in the muscle of DM1 patients (n=12) with different expansion lengths and normal subjects (n=7). The MYH14 gene is located on chromosome 19q13.3 and encodes for one of the heavy chains of the so called class II "nonmuscle" myosins (NMHCII). MYH14 has two alternative spliced isoforms: the inserted isoform (NMHCII-C1) which includes 8 amino acids located in the globular head of the protein, not encoded by the non inserted isoform (NMHCII-C0). Results showed a splicing unbalance of the MYH14 gene in DM1 muscle, with a prevalent expression of the NMHCII-C0 isoform more marked in DM1 patients harboring large CTG expansions. Minigene assay indicated that levels of the MBNL1 protein positively regulates the inclusion of the MYH14 exon 6. Quantitative analysis of the MYH14 expression revealed a significant reduction in the DM1 muscle samples, both at mRNA and protein level. No differences were found between DM1 and controls in the skeletal muscle localization of MYH14, obtained through immunofluorescence analysis. In line with the thesis of an "RNA gain of function" hypothesis described for the CTG mutation, we conclude that the alterations of the MYH14 gene may contribute to the DM1 molecular pathogenesis.

  20. Toll-Like Receptor 9 Stimulation Induces Aberrant Expression of a Proliferation-Inducing Ligand by Tonsillar Germinal Center B Cells in IgA Nephropathy.

    PubMed

    Muto, Masahiro; Manfroi, Benoit; Suzuki, Hitoshi; Joh, Kensuke; Nagai, Masaaki; Wakai, Sachiko; Righini, Christian; Maiguma, Masayuki; Izui, Shozo; Tomino, Yasuhiko; Huard, Bertrand; Suzuki, Yusuke

    2017-04-01

    The TNF family member a proliferation-inducing ligand (APRIL; also known as TNFSF13), produced by myeloid cells, participates in the generation and survival of antibody-producing plasma cells. We studied the potential role of APRIL in the pathogenesis of IgA nephropathy (IgAN). We found that a significant proportion of germinal centers (GCs) in tonsils of patients with IgAN contained cells aberrantly producing APRIL, contributing to an overall upregulation of tonsillar APRIL expression compared with that in tonsils of control patients with tonsillitis. In IgAN GC, antigen-experienced IgD(-)CD38(+/-)CD19(+) B cells expressing a switched IgG/IgA B cell receptor produced APRIL. Notably, these GC B cells expressed mRNA encoding the common cleavable APRIL-α but also, the less frequent APRIL-δ/ζ mRNA, which encodes a protein that lacks a furin cleavage site and is, thus, the uncleavable membrane-bound form. Significant correlation between TLR9 and APRIL expression levels existed in tonsils from patients with IgAN. In vitro, repeated TLR9 stimulation induced APRIL expression in tonsillar B cells from control patients with tonsillitis. Clinically, aberrant APRIL expression in tonsillar GC correlated with greater proteinuria, and patients with IgAN and aberrant APRIL overexpression in tonsillar GC responded well to tonsillectomy, with parallel decreases in serum levels of galactose-deficient IgA1. Taken together, our data indicate that antibody disorders in IgAN associate with TLR9-induced aberrant expression of APRIL in tonsillar GC B cells.

  1. Novel mechanism of aberrant ZIP4 expression with zinc supplementation in oral tumorigenesis.

    PubMed

    Ishida, Sho; Kasamatsu, Atsushi; Endo-Sakamoto, Yosuke; Nakashima, Dai; Koide, Nao; Takahara, Toshikazu; Shimizu, Toshihiro; Iyoda, Manabu; Shiiba, Masashi; Tanzawa, Hideki; Uzawa, Katsuhiro

    2017-01-29

    Zrt-Irt-like protein 4 (ZIP4) is critical molecule for proper mammalian development and releasing zinc from vesicular compartments. Recent studies suggested that ZIP4 plays an important role of tumor progression in pancreatic, prostate, and hepatocellular cancers, however, little is known about the detail mechanism of ZIP4 in their cancers. In the present study, we examined the possibility of ZIP4 as a new molecular target for oral squamous cell carcinoma (OSCC). We evaluated ZIP4 expression in OSCC-derived cell lines and primary OSCC samples by quantitative RT-PCR, immunoblotting, and immunohistochemistry (IHC). We also analyzed the clinical correlation between ZIP4 status and clinical behaviors in patients with OSCC. In addition, ZIP4 knockdown cells (shZIP4 cells) and ZnCl2 treatment were used for functional experiments, including cellular proliferation assay, zinc uptake assay, and cell-cycle analysis. ZIP4 mRNA and protein were up-regulated significantly in OSCCs compared with normal counterparts in vitro and in vivo. IHC showed that ZIP4 expression in the primary OSCC was positively correlated with primary tumoral size. The shZIP4 cells showed decrease accumulation of intercellular zinc and decreased cellular growth by cell-cycle arrest at the G1 phase, resulting from up-regulation of cyclin-dependent kinase inhibitors and down-regulation of cyclins and cyclin-dependent kinases. Since cellular growth of OSCC cells after treatment with zinc was significantly greater than control cells, we speculated that intercellular ZnCl2 accumulation is an important factor for cellular growth. Consistent with our hypothesis, not only decreased zinc uptake by ZIP4 knockdown but also chelating agent, N,N,N',N'-tetrakis(2-pyridylmethyl) ethylenediamine (TPEN), showed inhibitory effects of cellular proliferation. Therefore, our data provide evidence for an essential role of ZIP4 and intracellular zinc for tumoral growth in OSCC, suggesting that zinc uptake might be a