Allografting of murine ovarian follicles in a fibrin matrix: influence of follicle stage on graft outcome.
(2015) 4th International Conference “Strategies in Tissue Engineering”2015 — Location: Wurzburg (10.June.2015)
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- Abstract
- Introduction:For women diagnosed with a cancer at high risk of developing ovarian metastasis, transplantation of cryopreserved ovarian tissue is not recommended after disease remission. To restore fertility in these patients, a biodegradable artificial ovary containing isolated follicles and ovarian cells (OCs) could be a safer option. Materials and Methods:14 NMRI mice were ovariectomized in order to isolate murine ovarian follicles and OCs. Groups of 50 Primordial Primary (PP) and 50 Secondary (S) follicles were embedded in a fibrin matrix with 50,000 OCs and grafted to SCID mice for 2 and 7 days. For control purpose, follicles and cells were processed for live/dead assays to evaluate their viability before grafting. The follicle recovery rate was evaluated by histology. Follicle survival (TUNEL) and development (KI67) and graft vascularization (CD34) were analysed by immunohistochemical analysis after grafting. Follicle ultrastructure was assessed by transmission electron microscopy. Results: Before grafting the total percentage of viable follicles was 55.4% (<10% dead granulosa cells). All 28 fibrin clots were recovered after grafting. The follicle recovery rate was 16.1% for PP follicles and 39.2% for S follicles on day 2, and 5.9% for PP follicles and 26.8% for S follicles on day 7. Compared to the PP group, the S group showed a significantly higher recovery rate. On day 2, percentages of growing follicles were 81.8% and 100%, and on day 7, 100% and 96.7%, in the PP and S groups respectively. On day 2, new capillaries were found in the tissue surrounding the clot but not inside. On day 7, capillaries were observed around and inside the fibrin clot. Vessel area corresponded to 0.8% of the graft surface area in the PP group and 3.6% in the S group. Moreover, in the histological analysis, S follicles showed larger and functional vessels, as demonstrated by the presence of erythrocytes, than PP follicles. The preliminary results of transmission electron microscopy showed that follicles from PP and S groups had a normal aspect. Discussion Despite the healthy status of remaining follicles in both groups, our results suggest that secondary follicles are more likely to survive and develop after isolation, encapsulation in fibrin clots and grafting than primordial and primary follicles. These results lead us to hypothesize that the higher recovery rate of the larger follicles can be correlated with the mechanical properties of the fibrin matrix. Indeed in the mouse ovary, as well as in other animal species, the primordial follicles are located in the ovarian cortex, which composition and stiffness differ from the medullar region, where growing follicles tend to develop. Despite the preliminary results regarding the ultrastructure of the both populations of follicles further insights are necessary to assert with certainty that fibrin matrix is an optimal candidate to develop an artificial ovary.
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Chiti, M. C., & et al. (2015). Allografting of murine ovarian follicles in a fibrin matrix: influence of follicle stage on graft outcome. 4th International Conference “Strategies in Tissue Engineering”2015, Wurzburg.