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Personne :
Pouliot, Stéphanie

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Pouliot

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Stéphanie

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Département chirurgie, Faculté de médecine, Université Laval

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ncf11908659

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  • PublicationRestreint
    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.
  • PublicationRestreint
    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.
  • PublicationRestreint
    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