Bone marrow-derived mesenchymal stem cells drive lymphangiogenesis.
Maertens, Ludovic ; Erpicum, Charlotte ; Detry, Benoît et al
in PLoS ONE (2014), 9(9), 106976
It is now well accepted that multipotent Bone-Marrow Mesenchymal Stem Cells (BM-MSC) contribute to cancer progression through several mechanisms including angiogenesis. However, their involvement during ... [more ▼]
It is now well accepted that multipotent Bone-Marrow Mesenchymal Stem Cells (BM-MSC) contribute to cancer progression through several mechanisms including angiogenesis. However, their involvement during the lymphangiogenic process is poorly described. Using BM-MSC isolated from mice of two different backgrounds, we demonstrate a paracrine lymphangiogenic action of BM-MSC both in vivo and in vitro. Co-injection of BM-MSC and tumor cells in mice increased the in vivo tumor growth and intratumoral lymphatic vessel density. In addition, BM-MSC or their conditioned medium stimulated the recruitment of lymphatic vessels in vivo in an ear sponge assay, and ex vivo in the lymphatic ring assay (LRA). In vitro, MSC conditioned medium also increased the proliferation rate and the migration of both primary lymphatic endothelial cells (LEC) and an immortalized lymphatic endothelial cell line. Mechanistically, these pro-lymphangiogenic effects relied on the secretion of Vascular Endothelial Growth Factor (VEGF)-A by BM-MSC that activates VEGF Receptor (VEGFR)-2 pathway on LEC. Indeed, the trapping of VEGF-A in MSC conditioned medium by soluble VEGF Receptors (sVEGFR)-1, -2 or the inhibition of VEGFR-2 activity by a specific inhibitor (ZM 323881) both decreased LEC proliferation, migration and the phosphorylation of their main downstream target ERK1/2. This study provides direct unprecedented evidence for a paracrine lymphangiogenic action of BM-MSC via the production of VEGF-A which acts on LEC VEGFR-2. [less ▲]Detailed reference viewed: 23 (2 ULg)
Altered alpha-defensin 5 expression in cervical squamocolumnar junction: implication in the formation of a viral/tumour-permissive microenvironment.
Hubert, Pascale ; ; RONCARATI, Patrick et al
in Journal of Pathology (The) (2014), 234(4), 464-77
Human papillomavirus (HPV) infection, particularly type 16, is causally associated with cancer of the uterine cervix, which mainly develops at the squamocolumnar (SC) junction. The progression of cervical ... [more ▼]
Human papillomavirus (HPV) infection, particularly type 16, is causally associated with cancer of the uterine cervix, which mainly develops at the squamocolumnar (SC) junction. The progression of cervical HPV infections into (pre)neoplastic lesions suggests that viral antigens are not adequately recognized by innate immunity or presented to the adaptive immune system. Members of the defensin family have recently been found to inhibit viral and bacterial pathogens, to stimulate the migration of immune cells and to play a role in anticancer responses. In the present study, we focused on the poorly characterized human alpha-defensin 5 (HD-5) and its possible role in these processes. We showed that HD-5 was able to prevent HPV virion entry into cervical keratinocytes and to influence adaptive immunity. Indeed, this peptide specifically induced the chemoattraction and proliferation of both activated T lymphocytes and immature dendritic cells in a CCR2/CCR6-dependent manner and stimulated the infiltration of these professional antigen-presenting cells in a (pre)neoplastic epithelium transplanted in vivo in immunodeficient mice. No chemotactic effect was observed with plasmacytoid dendritic cells, macrophages or natural killer cells. Proliferative and angiogenic effects of HD-5 were also assessed in vitro and in vivo. However there was a striking regional disparity in expression of HD-5, being prominent in ectocervical, vaginal and vulvar neoplasia, while absent, or nearly so, in the cervical SC junction. Taken together, these results suggest one possible explanation for why the SC junction is uniquely vulnerable to both high-risk HPV infection (via reduced HD-5 expression and viral entry) and progression of neoplasia (via altered cell-mediated immune responses and altered microenvironment). Copyright (c) 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [less ▲]Detailed reference viewed: 41 (13 ULg)
A spiked tissue-based approach for quantification of phosphatidylcholines in brain section by MALDI mass spectrometry imaging.
Jadoul, Laure ; Longuespée, Rémi ; Noël, Agnès et al
in Analytical and Bioanalytical Chemistry (2014), sous presse
In the last few years, matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has been successfully used to study the distribution of lipids within tissue sections. However ... [more ▼]
In the last few years, matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has been successfully used to study the distribution of lipids within tissue sections. However, few efforts have been made to acquire reliable quantitative data regarding the localized concentrations of these molecules. Here we propose an approach based on brain homogenates for the quantification of phosphatidylcholines (PCs) in brain section by MALDI MSI. Homogenates were spiked with a range of PC(16:0 d31/18:1) concentrations. Sections from homogenates and intact brain were simultaneously prepared before being analyzed by MALDI MSI using a Fourier transform ion cyclotron resonance (FT-ICR) analyzer. Standard curves were generated from the signal intensity of the different PC(16:0 d31/18:1) ionic species ([M+H]+, [M+Na]+ and [M+K]+) detected from the homogenate sections. Localized quantitative data were finally extracted by correlating the standard curves with the signal intensities of endogenous PC (especially PC(16:0/18:1)) ionic species detected on different areas of the brain section. They were consistent with quantitative values found in the literature. This work introduces a new method to take directly into account biological matrix effects for the quantification of lipids as well as other endogenous compounds, in tissue sections by MALDI MSI. [less ▲]Detailed reference viewed: 18 (5 ULg)
Improved computer-assisted analysis of the global lymphatic network in human cervical tissues.
Balsat, Cédric ; ; GOFFIN, Frédéric et al
in Modern Pathology : An Official Journal of the United States & Canadian Academy of Pathology, Inc (2014), 27(6), 887-98
Lymphatic dissemination is a key event in cervical cancer progression and related tumor lymphatic markers are viewed as promising prognostic factor of nodal extension. However, validating such parameters ... [more ▼]
Lymphatic dissemination is a key event in cervical cancer progression and related tumor lymphatic markers are viewed as promising prognostic factor of nodal extension. However, validating such parameters requires an objective characterization of the lymphatic vasculature. Here, we performed a global analysis of the lymphatic network using a new computerized method applied on whole uterine cervical digital images. Sixty-eight cases of cervical neoplasia (12 CIN3, 10 FIGO stage 1A and 46 stage IB1) and 10 cases of normal cervical tissue were reacted with antibodies raised against D2-40, D2-40/p16 and D2-40/Ki67. Immunostained structures were automatically detected on whole slides. The lymphatic vessel density (D2-40), proliferating lymphatic vessel density (D2-40/ki67) and spatial lymphatic distribution in respect to the adjacent epithelium were assessed from normal cervix to early cervical cancer and correlated with lymphovascular space invasion and lymph node status. Prominent lymphatic vessel density and proliferating lymphatic vessel density are detected under the transformation zone of benign cervix and no further increase is noted during cancer progression. Notably, a shift of lymphatic vessel distribution toward the neoplastic edges is detected. In IB1 cervical cancer, although intra- and peritumoral lymphatic vessel density are neither correlated with lymphovascular space invasion nor with lymph node metastasis, a specific spatial distribution with more lymphatic vessels in the vicinity of tumor edges is predictive of lymphatic dissemination. Herein, we provide a new computerized method suitable for an innovative detailed analysis of the lymphatic network. We show that the transformation zone of the benign cervix acts as a baseline lymphangiogenic niche before the initiation of neoplastic process. During cancer progression, this specific microenvironment is maintained with lymphatic vessels even in closer vicinity to tumor cells.Modern Pathology advance online publication, 6 December 2013; doi:10.1038/modpathol.2013.195. [less ▲]Detailed reference viewed: 54 (20 ULg)
Blocking lipid synthesis overcomes tumor re-growth and metastasis after anti-angiogenic therapy withdrawal.
Sounni, Nor Eddine ; Cimino, Jonathan ; BLACHER, Silvia et al
in Cell Metabolism (2014), 20(2), 280-94
The molecular mechanisms responsible for the failure of antiangiogenic therapies and how tumors adapt to these therapies are unclear. Here, we applied transcriptomic, proteomic, and metabolomic approaches ... [more ▼]
The molecular mechanisms responsible for the failure of antiangiogenic therapies and how tumors adapt to these therapies are unclear. Here, we applied transcriptomic, proteomic, and metabolomic approaches to preclinical models and provide evidence for tumor adaptation to vascular endothelial growth factor blockade through a metabolic shift toward carbohydrate and lipid metabolism in tumors. During sunitinib or sorafenib treatment, tumor growth was inhibited and tumors were hypoxic and glycolytic. In sharp contrast, treatment withdrawal led to tumor regrowth, angiogenesis restoration, moderate lactate production, and enhanced lipid synthesis. This metabolic shift was associated with a drastic increase in metastatic dissemination. Interestingly, pharmacological lipogenesis inhibition with orlistat or fatty acid synthase downregulation with shRNA inhibited tumor regrowth and metastases after sunitinib treatment withdrawal. Our data shed light on metabolic alterations that result in cancer adaptation to antiangiogenic treatments and identify key molecules involved in lipid metabolism as putative therapeutic targets. [less ▲]Detailed reference viewed: 89 (31 ULg)
Estetrol and neuroprotection against perinatal ischemic insult
Tskitishvili, Ekaterine ; Nisolle, Michelle ; Noël, Agnès et al
in Estetrol attenuates neonatal hypoxic-ischemic brain injury (2014)Detailed reference viewed: 19 (5 ULg)
PAI-1 mediates the antiangiogenic and profibrinolytic effects of 16K prolactin.
Bajou, Khalid ; Herkenne, Stéphanie ; et al
in Nature Medicine (2014), sous presse
The N-terminal fragment of prolactin (16K PRL) inhibits tumor growth by impairing angiogenesis, but the underlying mechanisms are unknown. Here, we found that 16K PRL binds the fibrinolytic inhibitor ... [more ▼]
The N-terminal fragment of prolactin (16K PRL) inhibits tumor growth by impairing angiogenesis, but the underlying mechanisms are unknown. Here, we found that 16K PRL binds the fibrinolytic inhibitor plasminogen activator inhibitor-1 (PAI-1), which is known to contextually promote tumor angiogenesis and growth. Loss of PAI-1 abrogated the antitumoral and antiangiogenic effects of 16K PRL. PAI-1 bound the ternary complex PAI-1-urokinase-type plasminogen activator (uPA)-uPA receptor (uPAR), thereby exerting antiangiogenic effects. By inhibiting the antifibrinolytic activity of PAI-1, 16K PRL also protected mice against thromboembolism and promoted arterial clot lysis. Thus, by signaling through the PAI-1-uPA-uPAR complex, 16K PRL impairs tumor vascularization and growth and, by inhibiting the antifibrinolytic activity of PAI-1, promotes thrombolysis. [less ▲]Detailed reference viewed: 54 (20 ULg)
Cell invasion in the spheroid sprouting assay: a spatial organisation analysis adaptable to cell behaviour.
Blacher, Silvia ; Erpicum, Charlotte ; et al
in PLoS ONE (2014), 9(5), 97019
The endothelial cell spheroid assay provides a suitable in vitro model to study (lymph) angiogenesis and test pro- and anti-(lymph) angiogenic factors or drugs. Usually, the extent of cell invasion ... [more ▼]
The endothelial cell spheroid assay provides a suitable in vitro model to study (lymph) angiogenesis and test pro- and anti-(lymph) angiogenic factors or drugs. Usually, the extent of cell invasion, observed through optical microscopy, is measured. The present study proposes the spatial distribution of migrated cells as a new descriptor of the (lymph) angiogenic response. The utility of this novel method rests with its capacity to locally characterise spheroid structure, allowing not only the investigation of single and collective cell invasion but also the evolution of the spheroid core itself. Moreover, the proposed method can be applied to 2D-projected spheroid images obtained by optical microscopy, as well as to 3D images acquired by confocal microscopy. To validate the proposed methodology, endothelial cell invasion was evaluated under different experimental conditions. The results were compared with widely used global parameters. The comparison shows that our method prevents local spheroid modifications from being overlooked and leading to the possible misinterpretation of results. [less ▲]Detailed reference viewed: 33 (5 ULg)
Changes in elastin density in different locations of the vaginal wall in women with pelvic organ prolapse.
DE LANDSHEERE, Laurent ; Blacher, Silvia ; Munaut, Carine et al
in International Urogynecology Journal & Pelvic Floor Dysfunction (2014)
INTRODUCTION AND HYPOTHESIS: The purpose of this study was to analyze the histomorphometric properties of the vaginal wall in women with pelvic organ prolapse (POP). METHODS: In 15 women undergoing ... [more ▼]
INTRODUCTION AND HYPOTHESIS: The purpose of this study was to analyze the histomorphometric properties of the vaginal wall in women with pelvic organ prolapse (POP). METHODS: In 15 women undergoing surgery for POP, full-thickness biopsies were collected at two different sites of location from the anterior and/or posterior vaginal wall. Properties of the precervical area (POP-Q point C/D) were compared with the most distal portion of the vaginal wall (POP-Q point Ba/Bp) using histological staining and immunohistochemistry. The densities of total collagen fibers, elastic fibers, smooth muscle cells, and blood vessels were determined by combining high-resolution virtual imaging and computer-assisted digital image analysis. RESULTS: The mean elastin density was significantly decreased in the lamina propria and muscularis layer of the vaginal wall from the most distal portion of the prolapsed vaginal wall compared with the precervical area. This difference was statistically significant in the lamina propria for both anterior (8.4 +/- 1.2 and 12.1 +/- 2.0, p = 0.048) and posterior (6.8 +/- 0.5 and 10.1 +/- 1.4, p = 0.040) locations, and in the muscularis for the anterior (5.2 +/- 0.4 and 8.4 +/- 1.2, p = 0.009) vaginal wall. There were no statistically significant differences in the mean densities of collagen fibers, smooth muscle cells or blood vessels between the two locations. CONCLUSIONS: In this study, we observed changes in elastin density in two different locations of the vaginal wall from women with POP. The histomorphometric properties of the vaginal wall can be variable from one place to another in the same patient. This result supports the existence of most vulnerable locations within the vaginal wall and the potential benefit of site-specific prolapse surgery. [less ▲]Detailed reference viewed: 55 (12 ULg)
Effects of adenosine on lymphangiogenesis.
Lenoir, Bénédicte ; ; Blacher, Silvia et al
in PloS one (2014), 9(3), 92715
BACKGROUND: The lymphatic system controls tissue homeostasis by draining protein-rich lymph to the vascular system. Lymphangiogenesis, the formation of lymphatic vessels, is a normal event in childhood ... [more ▼]
BACKGROUND: The lymphatic system controls tissue homeostasis by draining protein-rich lymph to the vascular system. Lymphangiogenesis, the formation of lymphatic vessels, is a normal event in childhood but promotes tumor spread and metastasis during adulthood. Blocking lymphangiogenesis may therefore be of therapeutic interest. Production of adenosine is enhanced in the tumor environment and contributes to tumor progression through stimulation of angiogenesis. In this study, we determined whether adenosine affects lymphangiogenesis. METHODS: Lymphatic endothelial cells (HMVEC-dLy) were cultured in presence of adenosine and their proliferation, migration and tube formation was assessed. Gelatin sponges embedded with the stable analogue of adenosine 2-chloro adenosine were implanted in mice ear and lymphangiogenesis was quantified. Mice were intravenously injected with adenoviruses containing expression vector for 5'-endonucleotidase, which plays a major role in the formation of adenosine. RESULTS: In vitro, we observed that adenosine decreased the proliferation of lymphatic endothelial cells, their migration and tube formation. However, in vivo, gelatin sponges containing 2-chloro adenosine and implanted in mice ear displayed an elevated level of lymphangiogenesis (2.5-fold, p<0.001). Adenovirus-mediated over-expression of cytosolic 5'-nucleotidase IA stimulated lymphangiogenesis and the recruitment of macrophages in mouse liver. Proliferation of lymphatic endothelial cells was enhanced (2-fold, p<0.001) when incubated in the presence of conditioned medium from murine macrophages. CONCLUSION: We have shown that adenosine stimulates lymphangiogenesis in vivo, presumably through a macrophage-mediated mechanism. This observation suggests that blockade of adenosine receptors may help in anti-cancer therapies. [less ▲]Detailed reference viewed: 44 (2 ULg)
Gene expression pattern of synovial cells from inflammatory and normal areas of osteoarthritis synovial membrane.
Lambert, Cécile ; ; et al
in Arthritis and Rheumatism (2014), sous presse
Objective: The aim of this study was to compare the gene expression pattern of synovial cells from inflammatory (I) or normal/reactive (N/R) areas of a synovial membrane harvested from the same ... [more ▼]
Objective: The aim of this study was to compare the gene expression pattern of synovial cells from inflammatory (I) or normal/reactive (N/R) areas of a synovial membrane harvested from the same osteoarthritis (OA) patient. Methods: Synovial tissues were obtained from 12 knee OA patients at the time of total knee replacement. The inflammatory status of the synovial membrane was characterized according to macroscopic criteria and sorted as N/R and I. Biopsies were cultured separately for 7 days. Microarray gene expression profiling between N/R and I areas was performed. Western blot and immunohistochemistry confirmed the identified genes that were differentially expressed. Results: 896 differentially expressed genes between N/R and I zones were identified. The key pathways were related to inflammation, cartilage metabolism, Wnt signaling and angiogenesis. In the inflammatory network, TREM1 and S100A9 were strongly up-regulated. MMP-3 and -9, cathepsin H and S were significantly up-regulated in the cartilage catabolism pathway, whereas the most up-regulated anabolism enzyme was HAS1. Wnt-5A and LRP5 were up-regulated whereas FZD2 and DKK3 were down-regulated in the Wnt signaling. Finally, STC1, a protein involved in angiogenesis was identified as the most up-regulated gene in I zones compared to N/R zones. Conclusion: This study is the first to identify different expression pattern between two areas of the synovial membrane in the same patient. These differences concern several key pathways involved in OA pathogenesis. This analysis also provides information regarding new genes and proteins as potential targets for the future therapeutic. (c) 2013 American College of Rheumatology. [less ▲]Detailed reference viewed: 23 (4 ULg)
MMP-Mediated Collagen Remodeling and Vessel Functions
Noël, Agnès ; Sounni, Nor Eddine
in BRIX, K.; STOCKER, W. (Eds.) Proteases: Structure and Function, (2014)Detailed reference viewed: 36 (5 ULg)
Molecular mechanisms of type I collagen-induced apoptosis in breast carcinoma cells
Maquoi, Erik ; Assent, Delphine ; Foidart, Jean-Michel et al
Poster (2013, September 27)
Objective: As invading breast carcinoma cells breach the underlying basement membrane, they become confronted with a dense three-dimensional reactive stroma dominated by type I collagen. To develop ... [more ▼]
Objective: As invading breast carcinoma cells breach the underlying basement membrane, they become confronted with a dense three-dimensional reactive stroma dominated by type I collagen. To develop metastatic capabilities, invading tumour cells must acquire the capacity to negotiate this hostile microenvironment. By enmeshing cells in a dense fibrillar network, type I collagen acts as a physical barrier for cell migration as well as an endogenous antigrowth signal, partly by inducing apoptosis in epithelial cells. Aberrant cell survival resulting from an acquired resistance toward apoptosis represents a prominent hallmark of cancers. However, the molecular mechanisms implicated in collagen-induced apoptosis remain poorly defined. Here, we investigate the molecular mechanisms by which type I collagen induces apoptosis in breast carcinoma cells and identify MMP-14, a membrane-anchored matrix metalloproteinase, as a key anti-apoptotic factor. Methods: To investigate the induction of apoptosis by collagen, human breast adenocarcinoma MCF-7 cells overexpressing or not MMP-14 were plated on plastic plates or embedded within three dimensional type I collagen gels (Col3D). Cell death was evaluated by measuring cytoplasmic histone-associated DNA fragments (Cell Death Detection ELISA). The percentage of cells with an apoptotic nuclear morphology was also determined. The interactions between cancer cells and Col3D were analyzed by confocal microscopy and the impact of Col3D on the transcriptome of cancer cells was investigated using Illumina HT-12 BeadArrays. Results: When cultured within Col3D gels, MCF-7 cells displayed a round morphology and a cell death characterized by a Z-VAD-FMK-dependent chromatin condensation, nuclear segmentation and oligonucleosomal DNA fragmentation was induced. Transfection of MCF-7 cells with MMP-14 cDNA promoted the interactions between cells and collagen and prevented apoptosis. A transcriptomic analysis revealed that culturing MCF-7 cells within Col3D altered the expression of about 700 genes, irrespective of MMP-14 expression. Col3D modulated the expression of several apoptosis-related genes. Interestingly, MMP-14 activity was sufficient to prevent the Col3D-dependent induction of Bcl2-Interacting Killer (BIK), a pro-apoptotic member of the Bcl-2 family. Conclusions: Our results shed light on the molecular mechanisms by which a collagen-rich microenvironment triggers apoptosis in invading breast cancer cells. Furthermore, we demonstrate that MMP-14 promotes tumour progression by circumventing apoptosis. [less ▲]Detailed reference viewed: 103 (7 ULg)
Age-related Macular Degeneration (AMD): from Metabolomics approach to the inhibition of PDH Kinase as a new therapeutic target
Arslan, Deniz ; Dilly, Sébastien ; LAMBERT, Vincent et al
Poster (2013, June 06)Detailed reference viewed: 86 (34 ULg)
Age-related Macular Degeneration Study: A Metabolomics Approach
LAMBERT, Vincent ; Hansen, Sylvain ; et al
Conference (2013, May 23)
Age-related macular degeneration (AMD) is a leading cause of vision loss in the western world among people aged 50 or older. 90% of all vision loss due to AMD result from the exudative form, which is ... [more ▼]
Age-related macular degeneration (AMD) is a leading cause of vision loss in the western world among people aged 50 or older. 90% of all vision loss due to AMD result from the exudative form, which is characterized by choroidal neovascularization (CNV). Age-related changes that induce pathologic CNV are incompletely understood. A successful application of anti-VEGF approaches in the clinic is obviously a turning point in AMD treatment. Nevertheless, despite such important advances, critical issues remain to be addressed. To better understand the etiology of this pathology, we have used and improved a model of laser-induced murine choroidal neovascularization. As none is known about the metabolic changes in patients with AMD, we decided to apply a 1H NMR metabolomics approach on AMD patients and on a mice CNV experimental model. This technique is a relatively new branch of « omics » technologies focused on the analysis and measurement of endogenous metabolites. These experiments provide unique challenges to fulfill the goal of improving the current status of physiological information related to metabolome and in general to functional genomics. For this purpose, sera from control and AMD patients, induced and non-induced mice have been collected and the metabolic profiles of these samples were determined by 1H NMR. After post-processing treatments, the different spectra were analyzed by statistical discriminant methodologies (PCA, ICA, PLS-DA). This approach allows the differentiation between control and AMD patients and between laser-induced mice and the control mice group. Moreover, the same discriminating spectral zones, lactate and lipoproteins profil, have been identified in human and mice model, leading to the emergence of different putative biomarkers. In mice model of laser-induced CNV, normalization of circulating lactate by dichloroacetate, decreases CNV development. These first interesting results open new way in the study of the AMD etiology and validate the use of 1H NMR and CNV model for the understanding of Age-related Macular Degeneration. [less ▲]Detailed reference viewed: 49 (11 ULg)
Investigation of potential new targets for the diagnosis and/or the treatment of osteoarthritis
Lambert, Cécile ; ; et al
in Osteoarthritis and Cartilage (2013, April), 21(Supplement April 2013),
Purpose: Synovial inflammation plays a key role in the pathophysiology process of osteoarthritis (OA). We have previously compared the gene expression pattern of synovial cells isolated from inflammatory ... [more ▼]
Purpose: Synovial inflammation plays a key role in the pathophysiology process of osteoarthritis (OA). We have previously compared the gene expression pattern of synovial cells isolated from inflammatory (I) or normal/reactive (N/R) areas of a synovial membrane harvested from the same OA patient. We identified a large number of mediators belonging to key pathways involved in OA pathogenesis. The aim of this study was to validate different potential new targets for the diagnosis and/or the treatment OA. Methods: Synovial cells (SC) were isolated from synovial specimens obtained from OA patients undergoing knee replacement. The inflammatory status of the synovial membrane was characterized according to macroscopic criteria. The biopsies from N/R and I areas were cultured separately for a period of 7 days. Microarray gene expression profiling between N/R and I areas was performed. The biological relevance of up- and down-regulated genes was analyzed with Ingenuity Pathways Analysis. Western blot and immunohistochemistry confirmed the identified genes most differentially expressed in the key pathways. The production of the triggering receptor expressed on myeloid cells-1 (TREM1), the alarmin S100 calcium binding protein A9 (S100A9), the wingless-type MMTV integration site family, member 5A (Wnt-5A) and the stanniocalcin 1 (STC1) were evaluated by Western blot. S100A9, hyaluronan synthase-1 (HAS1) and STC1 expression and localization were investigated by immunohistochemistry. Results: 896 genes differentially expressed in N/R and I areas were identified. The key pathways were related to inflammation, cartilage metabolism, Wnt signaling and angiogenesis. In the inflammatory gene pattern, TREM1 and S100A9 were strongly upregulated. We validated the production of these proteins in OA synovial biopsies by Western blot. TREM1 and S100A9 were increased in I compared to N/R synovial cells culture. S100A9 was observed in the perivascular area and in sublining cells in I synovial biopsies, but not in N/R biopsies. An increased staining was also observed in the intima lining layer of I when compared to N/R biopsies. The most upregulated anabolism enzyme in I synovial biopsies was HAS1. Using immunohistochemistry, we observed in I areas an increase of the HAS1-positive cells mainly in the intima lining. We also studied the protein production of Wnt-5A, the most upregulated intermediate of Wnt signaling pathway. The protein level was increased in I compared to N/R areas. Finally, in the angiogenesis pathway, one the most u-regulated gene was STC1. A significant increase of STC1 production was observed in I areas compared to N/R areas by Western blot. This result was also supported by the immunohistochemical analysis. In I area, the staining for STC1 was more intense in perivascular and sublining cells. Conclusions: Synovial membrane inflammation is a key target for OA treatments. In this work, we have identified proteins involved in the synovitis pathways like angiogenesis, cells infiltration and matrix remodeling. These proteins could be targeted by drugs and used as companion biomarkers for evaluating their efficacy. Although qualitative, our results could also yield to the identification of markers of the disease. This investigation has to be further pursued. [less ▲]Detailed reference viewed: 86 (5 ULg)
Lymphangiogenesis and extracellular matrix remodeling
Erpicum, Charlotte ; Detry, Benoît ; Paupert, Jenny et al
Conference (2013, January 28)Detailed reference viewed: 40 (11 ULg)
Isoform 111 of vascular endothelial growth factor (VEGF111) improves angiogenesis of ovarian tissue xenotransplantation
Labied, Soraya ; Delforge, Yves ; Munaut, Carine et al
in Transplantation (2013), 95(3), 426-433
Background: Cryopreservation of cortex ovarian tissue before anti-cancer therapy is a promising technique for fertility preservation mainly in children and young women. Ischemia in the early stage after ... [more ▼]
Background: Cryopreservation of cortex ovarian tissue before anti-cancer therapy is a promising technique for fertility preservation mainly in children and young women. Ischemia in the early stage after ovarian graft causes massive follicle loss by apoptosis. VEGF111 is a recently described VEGF isoform that does not bind to the extracellular matrix, diffuse extensively and is resistant to proteolysis. These properties confer a significantly higher angiogenic potential to VEGF111 in comparison to the other VEGF isoforms. Methods: We evaluated the morphology of cryopreserved sheep ovarian cortex, grafted in the presence or absence of VEGF111. Ovarian cortex biopsies were embedded in type I collagen with or without VEGF111 addition before transplantation to SCID mice ovaries. Transplants were retrieved 3 days or 3 weeks later. Follicular density, vasculature network, haemoglobin content and cell proliferation were analysed. Results: Addition of VEGF111 increased density of functional capillaries (p=0.01) 3 days after grafting. By double immunostaining of Ki-67 and von Willebrand Factor (vWF) we demonstrated that proliferating endothelial cells were found in 83% of the VEGF111 group when compared to 33% in the control group (p=0.001). This angio-stimulation was associated with a significant enhancement of haemoglobin content (p=0.03). Three weeks after transplantation, the number of primary follicles was significantly higher in VEGF111 grafts (p=0.02). Conclusion: VEGF111 accelerates blood vessels recruitment, functional angiogenesis and improves the viability of ovarian cortex by limiting ischemia and ovarian cortex damage. [less ▲]Detailed reference viewed: 50 (19 ULg)
Targeting a single function of the multifunctional matrix metalloprotease MT1-MMP. Impact on lymphangiogenesis.
; ; Erpicum, Charlotte et al
in Journal of Biological Chemistry (2013), sous presse
The group of matrix metalloproteases (MMPs) is responsible for multiple processes of extracellular matrix remodeling in the healthy body but also for matrix and tissue destruction during cancer invasion ... [more ▼]
The group of matrix metalloproteases (MMPs) is responsible for multiple processes of extracellular matrix remodeling in the healthy body but also for matrix and tissue destruction during cancer invasion and metastasis. The understanding of the contributions from each individual MMP, both in healthy and pathological events, has been complicated by the lack of specific inhibitors and the fact that some of the potent MMPs are multifunctional enzymes. These factors have also hampered the setup of therapeutic strategies targeting MMP activity. A tempting target is the membrane-associated MT1-MMP which has well-documented importance in matrix degradation but which takes part in more than one pathway in this regard. In this report, we describe the selective targeting of a single function of this enzyme by means of a specific monoclonal antibody against MT1-MMP, raised in an MT1-MMP knock-out mouse. The antibody blocks the enzyme ability to activate proMMP-2 without interfering with the collagenolytic function or the general proteolytic activity of MT1-MMP. Using this antibody, we have shown that the MT1-MMP-catalyzed activation of proMMP- 2 is involved in the outgrowth of cultured lymphatic endothelial cells in a collagen matrix in vitro, as well as in lymphatic vessel sprouting assayed ex vivo. This is the first example of the complete inactivation of a single function of a multifunctional MMP and the use of this strategy to pursue its role. [less ▲]Detailed reference viewed: 22 (8 ULg)
Sunitinib inhibits inflammatory corneal lymphangiogenesis.
Detry, Benoît ; Blacher, Silvia ; Erpicum, Charlotte et al
in Investigative Ophthalmology & Visual Science (2013), 54(5), 3082-93
PURPOSE: To evaluate the antilymphangiogenic potential of multi-target tyrosine kinase inhibitor sunitinib in corneal neovascularization (NV). METHODS: Inflammatory corneal NV was induced by thermal ... [more ▼]
PURPOSE: To evaluate the antilymphangiogenic potential of multi-target tyrosine kinase inhibitor sunitinib in corneal neovascularization (NV). METHODS: Inflammatory corneal NV was induced by thermal cauterization applied in the central cornea of mice, to which sunitinib malate was daily administered by gavage or not. At days 6, 11, or 17 post cauterization, lymphatic and blood vessels, as well as inflammatory cells were immunostained and quantified in whole-mounted corneas. RT-PCRs were performed to evidence VEGF-A, VEGF-C, VEGF-D, placental growth factor (PlGF), and soluble vascular endothelial growth factor receptor (VEGFR)-1 and -2 (sVEGFR-1, sVEGFR-2) expressions. Macrophages were isolated from mice peritoneal cavity following thioglycollate injection to produce conditioned medium. The effects of sunitinib were evaluated in vitro in the aortic and lymphatic ring assays in the presence or not of macrophage conditioned medium. RESULTS: Sunitinib treatment drastically reduced pathologic corneal lymphangiogenesis and angiogenesis. Reduced F4/80+ cell infiltration was evidenced in sunitinib-treated mice and was associated to decreased VEGF-A (by 50%, P < 0.01) and VEGF-C (by 35%, P < 0.01) expressions, while VEGF-D and sVEGFR-2 expressions were not affected. In vitro, sunitinib dose-dependently inhibited aortic ring outgrowth, but failed to affect lymphangiogenesis in the lymphatic ring assay. However, macrophage conditioned medium-enhanced angiogenesis and lymphangiogenesis were both strongly counteracted by sunitinib treatment. Mechanistically, sunitinib blocked VEGFR-2 phosphorylation induced by VEGF-A released by macrophages. CONCLUSIONS: Sunitinib exerts antihemangiogenic and antilymphangiogenic effects in vivo by reducing F4/80+ cell recruitment and interacting with their released factors. [less ▲]Detailed reference viewed: 32 (9 ULg)