Personne :
Laflamme, Karina

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Structures organisationnelles
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Département de biologie moléculaire, Faculté de médecine, Université Laval
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Voici les éléments 1 - 8 sur 8
  • Publication
    Recent optimization of a tissue engineered blood vessel : the LOEX experience
    (Excerpta Medica Foundation, 2004-05-01) Grenier, Guillaume.; Germain, Lucie; Auger, François A.; Rémy-Zolghadri, Murielle; Laflamme, Karina
    Creating a blood vessel by tissue engineering is one of the most demanding goals in tissue engineering. Our laboratory developed, using the self-assembly approach, the first completely biological tissue engineered blood vessel (TEBV) constituted of living human cells in the absence of any synthetic or exogenous material. The phenotypic and functional variations of smooth muscle cell (SMC) are of paramount importance in TEBV reconstruction. Thus, the phenotype and extracellular matrix (ECM) production of SMC were studied along the whole sequence of TEBV production. The functional and mechanical properties can be greatly enhanced by active cell orientation in the ECM. Accordingly, the method of preparing living tissue engineered sheets was modified to obtain an optimal alignment of SMC before rolling them into a tubular form. These results have allowed us to create a better TEBV.
  • Publication
    Tissue-engineered human vascular media produced in vitro by the self-assembly approach present functional properties similar to those of their native blood vessels
    (Mary Ann Liebert, Inc, 2006-09-12) Germain, Lucie; Pouliot, Stéphanie; Roberge, Charles; D’Orléans-Juste, Pedro; Auger, François A.; Laflamme, Karina
    We have developed a tissue-engineering approach for the production of a completely biological blood vessel from cultured human cells. In the present study, we took advantage of this tissue-engineering method to demonstrate that it can be used to reproduce the subtle differences in the expression of receptors present on the media of native human blood vessels. Indeed, a small percentage (3 of 18) of native human umbilical cord veins (HUCVs) responded to endothelin, the most powerful vasopressor agent known to date, via both endothelin A (ETA) and endothelin B (ETB) receptor activation. In contrast, most HUCVs tested responded toETviaETA receptor activation only. Tissue-engineered vascular media (TEVM) were next reconstructed by using vascular smooth muscle cells (VSMCs) isolated and cultured from HUCVs expressing both ETA and ETB receptors to determine the functional integrity of our TEVM model. The reconstructed TEVM presents an endothelin response similar to that of respective HUCVs from which VSMCs were isolated. Reverse transcriptase polymerase chain reaction on TEVM reconstructed in vitro correlated these vasocontractile profiles by showing the presence of messenger RNA for both ETA and ETB receptors. Taken together with recently published results on TEVM expressing only ETA receptor, these results show that our reconstructed TEVM present a similar ET response profile as the blood vessel from which the VSMCs were isolated and cultured. These findings indicate that subtle differences, such as receptor expression, are preserved in the reconstructed tissue. Therefore, our TEVM offers a valuable human in vitro model with which to study the functionality of human blood vessels, such as their vasoactive response, or to perform pharmacologic studies.
  • Publication
    Accès libre
    Vaisseaux humains reconstitués par génie tissulaire
    (2004-06-15) Stoclet, Jean-Claude; Germain, Lucie; Auger, François A.; Laflamme, Karina
    Les progrès du génie tissulaire permettent maintenant de reconstituer des vaisseaux sanguins fonctionnels à partir de cellules humaines. Dans des conditions bien précises, ces vaisseaux possèdent une structure, des propriétés mécaniques et des propriétés fonctionnelles (notamment en termes de vasomotricité) qui permettent de les utiliser comme modèles pour contourner les difficultés d’obtention et d’utilisation de vaisseaux humains pour la recherche expérimentale. En effet, l’utilisation de vaisseaux humains à des fins expérimentales est limitée par des problèmes éthiques et par les difficultés d’interprétation des résultats liées à leur hétérogénéité. C’est pourquoi une grande partie des recherches en biologie et en pharmacologie vasculaires est réalisée soit sur des modèles animaux, soit sur des cellules en culture, qui ne sont pas toujours représentatifs des vaisseaux humains. Le génie tissulaire peut apporter une source alternative de vaisseaux humains pour pallier ces inconvénients
  • Publication
    Adventitia contribution in vascular tone : insights from adventitia-derived cells in a tissue-engineered human blood vessel
    (Federation of American Societies for Experimental Biology, 2006-04-12) Grenier, Guillaume.; Germain, Lucie; Pouliot, Stéphanie; Labbé, Raymond; Roberge, Charles; Auger, François A.; Baker, Kathleen; D’Orléans-Juste, Pedro; Rémy-Zolghadri, Murielle; Laflamme, Karina
    Whether the adventitia component of blood vessels directly participates in the regulation of vascular tone remains to be demonstrated. We have recently developed a human tissue-engineered blood vessel comprising the three tunicae of a native blood vessel using the self-assembly approach. To investigate the role of the adventitia in the modulation of vascular tone, this tissue-engineering method was used to produce three vascular constructs from cells explanted and proliferated from donor vessel tunicae 1) an adventitia + a media, or only 2) an adventitia, or 3) a media. The vasoconstriction responses of these 3 constructs to endothelin, the most potent vasopressor known up-to-date, as well as to nonselective and selective agonists and antagonists, were compared. The adventitia contracted to endothelin-1, -2, whereas the media and the media+adventitia contracted to all three endothelins. Endothelin-induced contraction of the adventitia was dependent on ETA receptors, whereas that of the media and the adventitia+media was ETA and ETB receptor-dependent. RT-PCR studies corroborated these results. SNP induced a dose-dependent relaxation of the three tissue constructs. We also demonstrated that the endothelin-converting enzyme, responsible for the formation of the active endothelin peptides, was present and functional in the adventitia. In conclusion, this is the first direct demonstration that the adventitia has the capacity to contract and relax in response to vasoactive factors. The present study suggests that the adventitia of a blood vessel could play a greater role than expected in the modulation of blood vessel tone.—Laflamme, K., Roberge, C. J., Grenier, G., Rémy-Zolghadri, M., Pouliot, S., Baker, K., Labbé, R., D’Orléans-Juste, P., Auger, F. A., Germain, L. Adventitia contribution in vascular tone: insights from adventitia-derived cells in a tissue-engineered human blood vessel. the wall of a blood vessel is composed of three tunicae: intima, media, and adventitia (1)⤻ . The innermost tunica, known as the intima, includes a single layer of endothelial cells lining the vessel lumen and the internal elastic lamina membrane. The middle tunica, termed media, is mainly composed of vascular smooth muscle cells (VSMCs) in an extracellular matrix (ECM) and corresponds to the muscular portion of the blood vessel, whereas the tunica adventitia is mainly composed of vascular fibroblasts (VFs) and ECM. It is well accepted that the media of a blood vessel is responsible for the vasomotor tone control by contracting and relaxing in response to different hormonal factors released, for example, by the endothelial cells of the intima (2)⤻ . The adventitia, on the other hand, has long been thought to mainly serve as a structural support for the media, its main contribution to vascular compliance being controlled by autonomous perivascular innervation (1)⤻ . Interestingly, recent studies suggest that the adventitia influences vascular function (3⤻ 4⤻ 5⤻ 6⤻ 7)⤻ . Nonetheless, whether the adventitia can directly participate in the regulation of vasomotor tone of blood vessels still remains to be demonstrated. The lack of appropriate technical procedures to separate the adventitia tunica from the other components of a native blood vessel (stripping) has prevented direct investigations on the possible role of that layer in the regulation of vasomotor tone. For example, the stripping method used in these procedures can result in the injury of the media tunica and does not permit us to obtain functional adventitia isolated from a native blood vessel (6)⤻ . We have recently developed, using the self-assembly technique, a human tissue-engineered blood vessel (TEBV) composed of the layers representing the three tunicae found in a native blood vessel (8)⤻ . In the present study, we took advantage of the self-assembly method to produce three independent vascular constructs from amplified VSMCs and VFs isolated from the same human saphenous vein biopsy. The first vascular construct was composed of only an adventitia (TEVA), a second vascular construct contained only a media (TEVM), and the third contained a media and an adventitia (TEVMA). These three vascular models (TEVA, TEVM, and TEVMA) were reconstructed to investigate the role of the adventitia in the modulation of vascular tone by comparing each of these vascular construct responses to endothelin, the most powerful vasopressor agent known to date (9)⤻ . Studies in humans have demonstrated the importance of endothelin in the maintenance of vascular tone (10)⤻ and blood pressure (11)⤻ . Three endogenous isoforms of endothelin have been discovered, endothelin-1 (ET-1), endothelin-2 (ET-2) and endothelin-3 (ET-3) (12)⤻ . ET binds two different receptor subtypes: endothelin A (ETA) receptors, which have a higher affinity for ET-1 and ET-2 than ET-3, and endothelin B (ETB) receptors, which have equal affinity for ET-1, ET-2, and ET-3 (13)⤻ . The endothelin receptors (ETA and ETB) implicated in the observed responses to the peptide were also investigated in our three different vascular constructs. In the present study, all of the vascular constructs tested responded to endothelin, although a heterogeneity in the response was observed. Indeed, all three vascular constructs tested contracted to ET-1 and ET-2, but only TEVMA and TEVM responded to ET-3. Furthermore, endothelin-induced contraction of TEVA was found to be dependent on the presence of ETA receptors, while both ETA and ETB receptors were present and functional on TEVMA and TEVM. Finally, the three types of vascular constructs tested also had the capacity of vasodilating in response to a relaxing agent such as sodium nitroprusside (SNP). Our results show that the adventitia may play a greater role than expected in the maintenance of vascular tone and compliance.
  • Publication
    Tissue-Engineered Blood Vessels and the Future of Tissue Substitutes
    (Springer, 2005-01-01) Germain, Lucie; Auger, François A.; Laflamme, Karina
    It is seldom acknowledged that the very future of tissue engineering hinges upon the successful reproduction of the cardiovascular system. Since the tissue engineering of the heart is discussed elsewhere in this volume, the focus of the present chapter will be on the vascular system. The previous axiom has been clearly extolled in many physiology textbook, but has unfortunately received only recently all the scrutiny it deserves from tissue engineers. Thus, if the brain is the command centre, the human body must have an extremely efficient import/export system for the nutrition of all its tissues with the accompanying disposal of waste or toxic by-products of the normal metabolism. Our own group (LOEX) has been made keenly aware of the paramount role of vasculature by our clinicians. This has been so because every new project we put in place within our research team involve: biomedical biologists, bioengineers, and physicians specialised in the targeted organ. Thus the importance of reproducing functional vascular substitutes was frequently raised in many projects. These discussions were the impetus for our ongoing interest in tissue engineered blood vessels (TEBV) since 1989.
  • Publication
    Tissue-engineered human vascular media with a functional endothelin system.
    (American Heart Association etc, 2005-02-01) Labonté, Julie; Germain, Lucie; Pouliot, Stéphanie; Roberge, Charles; D’Orléans-Juste, Pedro; Auger, François A.; Laflamme, Karina
    Background— Cardiovascular diseases remain a major cause of death and disability in the Western world. Among the various approaches adopted to counteract the morbidity associated with these diseases, surgical procedures and cardiac and vascular xenotransplantations or allotransplantations are routinely performed. The suitable vascular graft would be as close as possible to the native and healthy vessel composed exclusively of human components provided by the patient and would adapt to the donor’s hemodynamics. We have developed such a tissue-engineered human blood vessel reconstructed with human cells. Because endothelin is the most potent vasopressor known to date, we were interested in investigating the functionality of the endothelinergic system in our reconstructed human blood vessel. Methods and Results— Vasoconstriction studies were performed with nonselective and selective agonists and antagonists to demonstrate that ETA receptors were present and functional in tissue-engineered human vascular media constructed with the self-assembly method. Reverse-transcriptase polymerase chain reaction studies demonstrated that mRNA of the ETA but not the ETB receptor was present in these human tissue–engineered blood vessels. Furthermore, we demonstrated that the endothelin-converting enzyme, the main enzyme responsible for the formation of the biologically active endothelin peptides, was present and functional in these same bioengineered vascular media. Conclusions— Our results suggest that the media component of our tissue-engineered blood vessel has the potential of controlling vascular resistance via the presence of functional endothelin ETA receptors and endothelin-converting enzyme. endothelinmuscle contractionmyocytes, smooth m
  • Publication
    Polyphenols modulate calcium-independent mechanisms in human arterial tissue-engineered vascular media
    (Mosby, 2007-09-30) Germain, Lucie; Diebolt, Myriam; Labbé, Raymond; Auger, François A.; Laflamme, Karina; Andriantsitohaina, Ramaroson
    Background: In the present study, an arterial tissue-engineered vascular media (TEVM) was produced from cultured human smooth muscle cells of the umbilical artery and we took advantage of this model to evaluate the regulation of contraction and the signalling pathways of polyphenols in arteries. Methods: Cultured human smooth muscle cells of the umbilical artery were used to produce arterial TEVMs. Contraction experiments were performed to determine intracellular targets involved in the modulation of contraction by polyphenols extract from red wine, Provinols (SEPPIC Groupe Air Liquide, Paris, France). Results: Smooth muscle cells in arterial TEVM displayed a differentiated phenotype as demonstrated by the expression of alpha-smooth muscle actin, a vascular smooth muscle-specific marker, and tissue contraction in response to vasoconstrictor and vasodilator agents. Contractions caused by histamine were associated with an increase in [Ca(2+)](i) and a Ca(2+)-independent signalling pathway. The latter pathway involved mechanisms sensitive to protein kinase C, myosin light chain kinase, and Rho-associated protein kinase inhibitors. The regulation of contraction induced by Provinols shows that treatment of arterial TEVM with this compound significantly decreased histamine-induced contraction. This effect was associated with the inhibition of the Rho-associated protein kinase pathway and the decrease in alpha-smooth muscle actin expression. Conclusion: The use of arterial TEVM, brings new insights into the mechanisms by which polyphenols regulate vascular contraction in the human artery.
  • Publication
    Accès libre
    Réponse vasocontractile des endothélines sur les différentes tuniques (media et adventice) d'un vaisseau sanguin humain reconstruit par génie tissulaire
    (2005) Laflamme, Karina; Germain, Lucie
    La méthode d'auto-assemblage développée dans notre laboratoire a permis de mettre au point une prothèse vasculaire de faible diamètre composée uniquement de cellules humaines. Cette prothèse possède plusieurs caractéristiques propres aux vaisseaux sanguins natifs. Un des rôles principaux des vaisseaux sanguins est la régulation du tonus musculaire par la contraction et la relaxation suite à l'action de différentes hormones. La technique d'auto-assemblage permet la reconstruction de chacune des différentes tuniques du vaisseau indépendamment les unes des autres. La modulation du tonus vasculaire médiée par les cellules musculaires lisses de la média a pu tout d'abord être évaluée. L'endothéline est le vasopresseur le plus puissant connu à ce jour et est impliquée dans la régulation du tonus musculaire et de la résistance périphérique. La présence d'un système endothélinergique fonctionnel sur la média de la prothèse vasculaire a été étudiée. Les résultats montrent que la média de cette prothèse possède un système endothélinergique fonctionnel et identique à celui retrouvé dans le tissu d'origine des cellules prélevées pour reconstruire la prothèse vasculaire. De plus, suite à ces résultats, différentes prothèses vasculaires possédant une expression différentielle des récepteurs aux endothélines ont été créées. Le rôle de l'adventice dans la régulation du tonus vasculaire n'est pas bien établi. Par la technique d'auto-assemblage, une média, une adventice et une media+une adventice ont pu être reconstruites afin d'étudier l'implication de l'adventice dans la vasoréactivité du vaisseau sanguin. Les résultats montrent une implication directe de l'adventice dans la réponse vasoactive du vaisseau. De plus, un nouveau modèle pharmacologique d'adventice a été créé. Ainsi, la technique d'auto-assemblage constitue un substitut vasculaire fonctionnel de choix pour la recherche fondamentale et pharmacologique.