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Martin G.,LOEX Inc | Guerard S.,LOEX Inc | Fortin M.-M.R.,LOEX Inc | Rusu D.,Laval University | And 3 more authors.
Laboratory Investigation | Year: 2012

Psoriasis, a chronic autoimmune-related skin disease, involves both immune and non-immune cells like T cells and keratinocytes. This study investigates the regulatory role of T cells-keratinocyte interactions during psoriasis on immune factors production. Cytokines and chemokines were evaluated by multiplex and ELISA assays in an in vitro model of co-culture of keratinocytes with T lymphocytes. Keratinocytes were from psoriatic skin lesions or healthy skin. T lymphocytes were from healthy volunteers. Psoriatic keratinocytes (PKs) alone generated concentrations of tumor necrosis factor (TNF)-α, interleukin (IL)-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-1Β, IL-8, monocyte chemotactic protein (MCP)-1, interferon-γ-induced protein 10 kDa (IP-10) and vascular endothelial growth factor (VEGF) higher than those produced by healthy keratinocytes (HKs). In contrast, IL-1α and IL-Ra production was reduced in PKs. Normal T cells, which had no effect on HKs, increased the production of TNF-α, IL-6, GM-CSF, IL-8, MCP-1 and IP-10 by PKs, but did not influence PK production of IL-1Β, IL-1α, IL-Ra and VEGF. The most striking effects were obtained with PK-and IL-2-stimulated T lymphocytes: most of the above cytokines and chemokines were greatly upregulated, except IL-1Β and VEGF that were decreased or unchanged, respectively. In addition, fractalkine was overproduced in this latter condition only. Our results indicate (1) a functional interaction between keratinocytes and T lymphocytes that requires a direct cellular contact, and (2) a reciprocal influence that depends on cytokine and chemokine types. In conclusion, lesional keratinocytes from psoriasis vulgaris alter functions of normal T lymphocytes that conversely modulate these keratinocytes. © 2012 USCAP, Inc All rights reserved.


Guerard S.,LOEX Inc | Guerard S.,Laval University | Allaeys I.,Laval University | Martin G.,LOEX Inc | And 3 more authors.
Archives of Dermatological Research | Year: 2013

Psoriatic plaques result from an abnormal proliferation of keratinocytes associated with the local presence of T lymphocytes and neutrophils. The exact role of neutrophils in psoriatic lesions remains unclear. The present investigation was aimed at deciphering the capacity of psoriatic keratinocytes to alter in vitro functions of neutrophils. Blood neutrophils from healthy donors were incubated with psoriatic (PK) or healthy keratinocytes (HK) with and without IL-2-activated healthy T lymphocytes. The study was focussed on neutrophil capacity of adherence, viability and superoxide anion production. PK or HK with or without T lymphocytes similarly augmented neutrophil viability after 48 h of co-incubation. PK or HK did not directly activate the superoxide production by neutrophils. However, they both primed neutrophils for an increased fMLF-induced production of superoxide, an effect enhanced by the presence of T lymphocytes. PK were 1.5-fold more efficient than HK to augment this superoxide production. PK cultured with T lymphocytes induced the adhesion of neutrophils 4.7 times more efficiently than HK. The adherence of neutrophils was mediated through ICAM-1, LFA-1 and Mac-1, independently of bioactive lipids. The effects of PK and HK on neutrophil viability and priming were independent of direct cellular contact. In conclusion, keratinocytes can impact neutrophils by increasing their lifespan, and by priming them to overproduce superoxide. PK are more efficient than HK in priming neutrophils, an effect enhanced by T lymphocytes. These results indicate that neutrophils could contribute to psoriasis pathogenesis partly through their pathological interactions with PK. © Springer-Verlag Berlin Heidelberg 2013.


Imbeault A.,Laval University | Bernard G.,LOEX Inc | Ouellet G.,LOEX Inc | Bouhout S.,LOEX Inc | And 3 more authors.
Journal of Sexual Medicine | Year: 2011

Introduction. Surgical treatment is indicated in severe cases of Peyronie's disease. Incision of the plaque with subsequent graft material implantation is the option of choice. Ideal graft tissue is not yet available. Aim. To evaluate the use of an autologous tissue-engineered endothelialized graft by the self-assembly method, for tunica albuginea (TA) reconstruction in Peyronie's disease. Methods. Two TA models were created. Human fibroblasts were isolated from a skin biopsy and cultured in vitro until formation of fibroblast sheets. After 4 weeks of maturation, human umbilical vein endothelial cells (HUVEC) were seeded on fibroblasts sheets and wrapped around a tubular support to form a cylinder of about 10 layers. After 21 days of tube maturation, HUVEC were seeded into the lumen of the fibroblast tubes for the endothelialized tunica albuginea (ETA). No HUVEC were seeded into the lumen for the TA model. Both constructs were placed under perfusion in a bioreactor for 1 week. Main Outcome Measures. Histology, immunohistochemistry, and burst pressure were performed to characterize mature tubular graft. Animal manipulations were also performed to demonstrate the impact of endothelial cells in vivo. Results. Histology showed uniform multilayered fibroblasts. Extracellular matrix, produced entirely by fibroblasts, presented a good staining for collagen 1. Some elastin fibers were also present. For the TA model, anti-human von Willebrand antibody revealed the endothelial cells forming capillary-like structures. TA model reached a burst pressure of 584mmHg and ETA model obtained a burst pressure of 719mmHg. Conclusions. This tissue-engineered endothelialized tubular graft is structurally similar to normal TA and presents an adequate mechanical resistance. The self-assembly method used and the autologous property of this model could represent an advantage comparatively to other available grafts. Further evaluation including functional testing will be necessary to characterize in vivo implantation and behavior of the graft. © 2011 International Society for Sexual Medicine.


Gibot L.,LOEX Inc | Galbraith T.,LOEX Inc | Huot J.,Laval University | Auger F.A.,LOEX Inc | Auger F.A.,Laval University
Clinical and Experimental Metastasis | Year: 2013

Cutaneous malignant melanomas represent an important clinical problem because they are highly invasive, they can metastasize to distant sites and are typically resistant to available therapy. The precise molecular determinants responsible for melanoma progression and chemo-resistance are not yet known, in part due to lack of pertinent experimental models that mimic human melanoma progression. Accordingly, we developed a complex human microvascularized reconstructed skin substitute in which the organized three-dimensional (3D) architecture of the native skin is reproduced. Human melanoma cell lines derived from primary and metastatic sites were added to this 3D model. Our results demonstrate that histological features and behavior of melanoma cells applied in our skin substitute model are specific to their site of origin. In particular, the ability of melanoma cells to cross the dermal-epidermal junction correlates with their metastatic potential. In addition, a potent angiogenic effect was detected for an aggressive metastatic cell line that produces VEGF. The presence of a microvascular network within this model will allow studying a crucial step of the metastatic process. We conclude that such an in vitro human tumor microvascularized reconstructed skin substitute promises to be a versatile and efficient model to investigate skin cancer progression and to screen new anticancer drugs to improve currents clinical treatments. © 2012 Springer Science+Business Media B.V.


Landreville S.,University of Washington | Vigneault F.,Harvard University | Bergeron M.-A.,LOEX Inc | Leclerc S.,LOEX Inc | And 7 more authors.
Pigment Cell and Melanoma Research | Year: 2011

Cancer aggressiveness is related to the ability of cancer cells to escape the anchorage dependency toward the extracellular matrix, a process regulated by the integrin α5β1 and its ligand fibronectin. Here, we characterized the expression of the α5 gene in human uveal melanoma cell lines with distinct tumorigenic properties and investigated some of the mechanisms underlying the variations of their malignancy. Strong and weak expression of α5 was observed in cells with no (T108/T115) and high (T97/T98) tumorigenic properties, respectively. Expression and DNA binding of the transcription factors Sp1, activator protein 1 (AP-1) (both acting as activators), and nuclear factor I (NFI) (a strong repressor) to the α5 promoter were demonstrated in all cell lines. A reduced expression of AP-1 combined with a dramatic increase in NFI correlated with the suppression of α5 expression in T97 and T98 cells. Restoring α5 expression in T97 cells entirely abolished their tumorigenicity in immunodeficient mice. These uveal melanoma cell lines might therefore prove particularly useful as cellular models to investigate α5β1 function in the pathogenesis of invasive uveal melanoma. © 2011 John Wiley & Sons A/S.


Gendron S.P.,LOEX Inc | Bastien N.,LOEX Inc | Mallet J.D.,LOEX Inc | Rochette P.J.,LOEX Inc
Mutagenesis | Year: 2013

In human skin, the 3895-bp deletion of mitochondrial DNA (mtDNA 3895) is catalysed by ultraviolet (UV) light through the generation of reactive oxygen species. Given its function in vision, the human eye is exposed to oxidising UV and blue light in its anterior (cornea, iris) and posterior (retina) structures. In this study, we employed a highly sensitive quantitative PCR technique to determine mtDNA3895 occurrence in human eye. Our analysis shows that the mtDNA3895 is concentrated in both the cornea and the retina. Within the cornea, the highest mtDNA3895 level is found in the stroma, the cellular layer conferring transparency and rigidity to the human cornea. Moreover, mtDNA3895 accumulates with age in the stroma, suggesting a role of this deletion in corneal ageing. Within the retina, mtDNA3895 is concentrated in the macular region of both the neural retina and the retinal pigment epithelium, supporting the hypothesis that this deletion is implicated in retinal pathologies such as age-related macular degenerescence. Taken together, our results imply that UV and blue light catalyse mtDNA3895 induction in the human eye. © The Author 2013. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved.


Robitaille H.,LOEX Inc | Robitaille H.,Laval University | Simard-Bisson C.,LOEX Inc | Simard-Bisson C.,Laval University | And 6 more authors.
Journal of Investigative Dermatology | Year: 2010

Hsp27, a small heat-shock protein, has important roles in many cellular processes, including cytoskeleton dynamics, cell differentiation, and apoptosis. Its expression in normal epidermis correlates with differentiation; however, little is known about the regulatory mechanisms involved. In this study, we report that Hsp27 undergoes upregulation, phosphorylation, and redistribution to the cytoskeleton during the late phase of epidermal keratinocyte differentiation. Our results also show that the expression of the dual leucine zipper-bearing kinase (DLK), an upstream activator of the MAP kinase pathways, is sufficient by itself to induce Hsp27 phosphorylation, cell periphery localization, and redistribution to the insoluble protein fraction (cytoskeleton) in poorly differentiated keratinocytes. This redistribution correlates with the insolubilization of cornified envelope-associated proteins such as involucrin. Interestingly, the effects of DLK on Hsp27 were blocked by PD98059, a selective inhibitor of the extracellular signal-regulated protein kinase (ERK) pathway. Moreover, downregulation of Hsp27 by small interfering RNA in epithelial cells expressing DLK was accompanied by attenuated expression of involucrin in the cytoskeleton. Thus, these observations suggest that the DLK-ERK signaling pathway may act as a regulator of the interaction that occurs between Hsp27 and the cytoskeleton during the formation of the cornified cell envelope, a process conferring to the skin its crucial barrier function. © 2010 The Society for Investigative Dermatology.


Mayrand D.,Laval University | Laforce-Lavoie A.,Laval University | Larochelle S.,Laval University | Langlois A.,Laval University | And 4 more authors.
Angiogenesis | Year: 2012

During wound healing, angiogenesis plays a crucial role in inducing adequate perfusion of the new tissue, thereby allowing its survival. This angiogenic process contributes to the formation of granulation tissue, alongside myofibroblasts. Myofibroblasts are cells specialized in wound contraction and synthesis of new extracellular matrix. Fibroblasts, considered by some to be at the origin of myofibroblasts, have already been shown to promote neovascularization. Thus, we hypothesized that myofibroblasts play a key role during angiogenic development in wound healing. We isolated myofibroblasts from normal human skin wounds and dermal microvascular endothelial cells (HDMVEC) and fibroblasts from skin. Using an in vitro fibrin-based model, we compared the proangiogenic activity of wound myofibroblasts to that of fibroblasts in the presence of HDMVEC. By immunostaining with collagen IV antibodies, we observed the formation of a capillary network significantly more developed when HDMVEC were cultured with myofibroblasts compared to the network formed in the presence of fibroblasts. The differences between these cell types did not result from a differential secretion of Vascular Endothelial Growth Factor or basic Fibroblast Growth Factor. However, in the presence of myofibroblasts, a significant decrease in matrix metalloproteinase activity was observed. This finding was correlated with a significant increase in Tissue Inhibitor of MetalloProteinase (TIMP)-1 and TIMP-3. Furthermore, inhibition of TIMP-1 secretion using shRNA significantly decreased myofibroblasts induced angiogenesis. These results led to the hypothesis that normal wound myofibroblasts contribute to the vascular network development during wound healing. Our data emphasize the critical role of wound myofibroblasts during healing. © Springer Science+Business Media B.V. 2012.


PubMed | LOEX Inc, Montreal Neurological Institute, Umeå University, University of Ulm and 3 more.
Type: | Journal: Human molecular genetics | Year: 2016

Recent genetic studies yielded conflicting results regarding a role for the variant chromogranin B (CHGB)


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