Tissus adipeux humains reconstruits par génie tissulaire : applications in vitro et in vivo
|Abstract:||White adipose tissue is one of the most abundant tissues of the human body and it represents its largest energy storage depot. It is also known as a secretory and endocrine organ acting remotely to influence many biological processes in addition to being a source of multipotent stromal/stem cells. Many patients suffer from adipose tissue deficits resulting from tumor resection, accidents, deep burns, congenital anomalies, etc. Tissue engineering represents a promising alternative to traditional approaches to answer the strong demand in adipose substitutes to counteract tissue loss using reconstructive surgery. In order to determine the in vivo performance of adipose tissue substitutes produced by tissue engineering and to maximize preclinical studies, it is crucial to develop biomedical imaging methodologies allowing graft delineation as well as volume and perfusion analyses. Therefore, human reconstructed adipose tissues (hrATs) containing mature functional adipocytes in addition to a natural extracellular matrix were produced according to the self-assembly approach of tissue engineering from adipose-derived stromal/stem cells and were next implanted into athymic mice. We hypothesized that magnetic resonance imaging (MRI) would allow an improved analysis of the in vivo performance of hrATs by promoting the non-invasive and longitudinal follow-up of graft volume and blood perfusion after implantation. Indeed, T1-weighted MRI allowed graft delineation and determination of their volume. Volume retention was evaluated up to six weeks after implantation, revealing a gradual resorption averaging at 44 % of initial substitute’s volume. Blood perfusion was confirmed 14 and 21 days after grafting. The persistence of numerous adipocytes within a rich extracellular matrix was also confirmed. This study described, for the first time, the optimal parameters necessary to achieve efficient MRI visualization of grafted tissue-engineered adipose substitutes. Ensuring a timely perfusion of engineered grafts is important to favor their survival. We hypothesized that the incorporation of an endothelial component into hrATs would be advantageous for the tissues and their global vascularization. Three days after the implantation into nude mice, an extended human capillary network connecting with the host network by anastomosis was detected within the grafts containing an endothelial component. The human capillary network persisted at least 14 days and murine red blood cells were detected within human structures. The global vascularization analysis also unveiled a tendency towards a beneficial impact of the endothelial component. Assessing the impact of this approach on the vascularization of thicker tissues will prove useful for the development of voluminous substitutes to counteract adipose defects in reconstructive surgery. Lastly, inflammation developing upon tissue grafting can have detrimental effects on engineered tissues. It is therefore important to study its impact on reconstructed tissues and their components. We used the hrAT model containing a preformed capillary network to study the impact of an inflammatory microenvironment on the capillary network within a three-dimensional adipose context. We hypothesized that a prolonged inflammatory context, created in vitro with tumor necrosis factor (TNF) and interleukin-1β (IL-1β), would impact hrAT’s secretory profile and mediate detrimental effects on the microvascular network. Analysis of conditioned media established tissue responsiveness. A disorganization of the capillary network was observed after a chronic exposure of three and six days to TNF and IL-1β and was characterized by a decrease of its extent and complexity. This disorganization was in part due to apoptosis. The hrAT model will allow the elaboration of strategies aiming at preserving the capillary network and the development of adipose tissue substitutes better suited to face inflammatory conditions in vivo. Overall, this work contributes to the advancement of the field of adipose tissue engineering through the optimization of MRI parameters for the follow-up of engineered adipose grafts but also through the development of hrATs that possess an advantageous in vivo performance in addition to being relevant three-dimensional in vitro models.|
|Document Type:||Thèse de doctorat|
|Open Access Date:||13 March 2020|
|Collection:||Thèses et mémoires|
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