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Copenhagen, Denmark

Jensen P.R.,University of Southern Denmark | Andersen T.L.,University of Southern Denmark | Pennypacker B.L.,Merck And Co. | Duong L.T.,Merck And Co. | And 2 more authors.
Biochemical and Biophysical Research Communications | Year: 2014

The bone matrix is maintained functional through the combined action of bone resorbing osteoclasts and bone forming osteoblasts, in so-called bone remodeling units. The coupling of these two activities is critical for securing bone replenishment and involves osteogenic factors released by the osteoclasts. However, the osteoclasts are separated from the mature bone forming osteoblasts in time and space. Therefore the target cell of these osteoclastic factors has remained unknown. Recent explorations of the physical microenvironment of osteoclasts revealed a cell layer lining the bone marrow and forming a canopy over the whole remodeling surface, spanning from the osteoclasts to the bone forming osteoblasts. Several observations show that these canopy cells are a source of osteoblast progenitors, and we hypothesized therefore that they are the likely cells targeted by the osteogenic factors of the osteoclasts. Here we provide evidence supporting this hypothesis, by comparing the osteoclast-canopy interface in response to two types of bone resorption inhibitors in rabbit lumbar vertebrae. The bisphosphonate alendronate, an inhibitor leading to low bone formation levels, reduces the extent of canopy coverage above osteoclasts. This effect is in accordance with its toxic action on periosteoclastic cells. In contrast, odanacatib, an inhibitor preserving bone formation, increases the extent of the osteoclast-canopy interface. Interestingly, these distinct effects correlate with how fast bone formation follows resorption during these respective treatments. Furthermore, canopy cells exhibit uPARAP/Endo180, a receptor able to bind the collagen made available by osteoclasts, and reported to mediate osteoblast recruitment. Overall these observations support a mechanism where the recruitment of bone forming osteoblasts from the canopy is induced by osteoclastic factors, thereby favoring initiation of bone formation. They lead to a model where the osteoclast-canopy interface is the physical site where coupling of bone resorption to bone formation occurs. © 2013 Elsevier Inc. All rights reserved.

Jensen T.O.,Aarhus University Hospital | Schmidt H.,Aarhus University Hospital | Moller H.J.,Aarhus University Hospital | Donskov F.,Aarhus University Hospital | And 4 more authors.
Cancer | Year: 2012

BACKGROUND: Tumor cell and host immune cell interaction plays a key role in carcinogenesis. Signal transducer and activator of transcription 3 (STAT3) is constitutively activated in cancer and believed to be an important mediator of tumor-induced immunosuppression. This paper aims to describe the prognostic impact of neutrophil and dendritic cell infiltration in primary melanoma and the association of this infiltration with activated STAT3 (pSTAT3) in primary melanoma cells. METHODS: Formalin-fixed, paraffin-embedded primary melanomas from 186 stage-I/II melanoma patients surgically resected from 1997 to 2000. Infiltrating neutrophils (CD66b), dendritic cells (CD123+ and DC-LAMP+), T-lymphocytes (CD8) and pSTAT3 melanoma cell expression were studied by immunohistochemistry and evaluated as present or absent. DC-LAMP+ cell infiltration was evaluated as absent/few versus dense. Study endpoints: relapse-free survival, melanoma-specific, and overall survival. RESULTS: The median observation time was 12.2 years (range, 10.4-14.2 years). Fifty-one deaths were observed of which 38 (20%) were melanoma-specific. In a multivariate Cox proportional hazards model including ulceration and melanoma thickness, neutrophil and CD123+ dendritic cell infiltration were independently associated with poor prognosis (CD66b: hazard ratio [HR] = 3.13; 95% confidence interval [CI], 1.43-6.83; P =.004; CD123: HR = 2.45; 95% CI, 1.22-4.92; P =.012). The association between melanoma cell pSTAT3 expression and immune infiltration (neutrophils and CD123+ cells) was strong. pSTAT3 expression, CD8 and DC-LAMP infiltration were not independently associated with poor prognosis. CONCLUSIONS: Neutrophil infiltration and CD123+ dendritic cell infiltration in primary melanoma are independently associated with poor prognosis. Melanoma cell expression of pSTAT3 is strongly associated with the surrounding immune infiltrate. Copyright © 2011 American Cancer Society.

Nielsen B.S.,Exiqon A S | Jorgensen S.,Exiqon A S | Fog J.U.,Exiqon A S | Sokilde R.,Exiqon A S | And 6 more authors.
Clinical and Experimental Metastasis | Year: 2011

Approximately 25% of all patients with stage II colorectal cancer will experience recurrent disease and subsequently die within 5 years. MicroRNA-21 (miR-21) is upregulated in several cancer types and has been associated with survival in colon cancer. In the present study we developed a robust in situ hybridization assay using high-affinity Locked Nucleic Acid (LNA) probes that specifically detect miR-21 in formalin-fixed paraffin embedded (FFPE) tissue samples. The expression of miR-21 was analyzed by in situ hybridization on 130 stage II colon and 67 stage II rectal cancer specimens. The miR-21 signal was revealed as a blue chromogenic reaction, predominantly observed in fibroblast-like cells located in the stromal compartment of the tumors. The expression levels were measured using image analysis. The miR-21 signal was determined as the total blue area (TB), or the area fraction relative to the nuclear density (TBR) obtained using a red nuclear stain. High TBR (and TB) estimates of miR-21 expression correlated significantly with shorter disease-free survival (p = 0.004, HR = 1.28, 95% CI: 1.06-1.55) in the stage II colon cancer patient group, whereas no significant correlation with disease-free survival was observed in the stage II rectal cancer group. In multivariate analysis both TB and TBR estimates were independent of other clinical parameters (age, gender, total leukocyte count, K-RAS mutational status and MSI). We conclude that miR-21 is primarily a stromal microRNA, which when measured by image analysis identifies a subgroup of stage II colon cancer patients with short diseasefree survival. © The Author(s) 2010.

Grum-Schwensen B.,Danish Cancer Society | Klingelhofer J.,Danish Cancer Society | Grigorian M.,Finsen Laboratory | Almholt K.,Finsen Laboratory | And 4 more authors.
Cancer Research | Year: 2010

Interactions between tumor and stroma cells are essential for the progression of cancer from its initial growth at a primary site to its metastasis to distant organs. The metastasis-stimulating protein S100A4 exerts its function as a stroma cell-derived factor. Genetic depletion of S100A4 significantly reduced the metastatic burden in lungs of PyMT-induced mammary tumors. In S100A4+/+ PyMT mice, massive leukocyte infiltration at the site of the growing tumor at the stage of malignant transition was associated with increased concentration of extracellular S100A4 in the tumor microenvironment. In contrast, in S100A4-/- PyMT tumors, a significant suppression of T-cell infiltration was documented at the transition period. In vitro, the S100A4 protein mediated the attraction of T cells. Moreover, S100A4+/+, but not S100A4-/-, fibroblasts stimulated the invasion of T lymphocytes into fibroblast monolayers. In vivo, the presence of S100A4+/+, but not S100A4-/-, fibroblasts significantly stimulated the attraction of T lymphocytes to the site of the growing tumor. Increased levels of T cells were also observed in the premetastatic lungs of tumor-bearing mice primed to metastasize by S100A4 +/+ fibroblasts. Treatment of T cells with the S100A4 protein stimulated production of cytokines, particularly granulocyte colony-stimulating factor and eotaxin-2. The same cytokines were detected in the fluid of S100A4+/+ PyMT tumors at the transition period. We suggest that release of S100A4 in the primary tumor stimulates infiltration of T cells and activates secretion of cytokines, thus triggering sequential events that fuel tumor cells to metastasize. Similar processes could occur in the premetastatic lungs, facilitating generation of inflammatory milieu favorable for metastasis formation. ©2010 AACR.

Beigneux A.P.,University of California at Los Angeles | Fong L.G.,University of California at Los Angeles | Bensadoun A.,Cornell University | Davies B.S.J.,University of Iowa | And 4 more authors.
Circulation Research | Year: 2015

Rationale: GPIHBP1, a GPI-anchored protein of capillary endothelial cells, binds lipoprotein lipase (LPL) in the subendothelial spaces and shuttles it to the capillary lumen. GPIHBP1 missense mutations that interfere with LPL binding cause familial chylomicronemia. Objective: We sought to understand mechanisms by which GPIHBP1 mutations prevent LPL binding and lead to chylomicronemia. Methods and Results: We expressed mutant forms of GPIHBP1 in Chinese hamster ovary cells, rat and human endothelial cells, and Drosophila S2 cells. In each expression system, mutation of cysteines in GPIHBP1's Ly6 domain (including mutants identified in patients with chylomicronemia) led to the formation of disulfide-linked dimers and multimers. GPIHBP1 dimerization/multimerization was not unique to cysteine mutations; mutations in other amino acid residues, including several associated with chylomicronemia, also led to protein dimerization/multimerization. The loss of GPIHBP1 monomers is relevant to the pathogenesis of chylomicronemia because only GPIHBP1 monomers - and not dimers or multimers - are capable of binding LPL. One GPIHBP1 mutant, GPIHBP1-W109S, had distinctive properties. GPIHBP1-W109S lacked the ability to bind LPL but had a reduced propensity for forming dimers or multimers, suggesting that W109 might play a more direct role in binding LPL. In support of that idea, replacing W109 with any of 8 other amino acids abolished LPL binding - and often did so without promoting the formation of dimers and multimers. Conclusions: Many amino acid substitutions in GPIHBP1's Ly6 domain that abolish LPL binding lead to protein dimerization/multimerization. Dimerization/multimerization is relevant to disease pathogenesis, given that only GPIHBP1 monomers are capable of binding LPL. © 2014 American Heart Association, Inc.

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