Figure 1.
The effect of S1P, SEW2871 and FTY720 on ovarian transplant revascularization and stromal cell population.
S1P induces ovarian angiogenesis (Left axis) and increases stromal cell population (Right axis) while its analogs, SEW2871 and FTY720 have the opposite effect, 10 days after grafting of ovarian transplantation. * Significantly different from control. # Significantly different from baseline ovarian tissue.
Figure 2.
Impact of S1P and its analogues on neo-angiogenesis, stromal cell proliferation and tissue hypoxia.
(A–F) Evaluation of angogenic effect of S1P and its analogs in ovarian transplants by anti-CD31 IHC. S1P induces angiogenesis of ovarian transplants (D) while FTY720 (E) or SEW2871 (F) treatment reduces it. Arrows show newly formed blood vessels positively stained for anti-CD31. (G–L) Evaluation of cell proliferation by anti-PCNA expression. S1P induces (J) where as FTY720 (K) reduces stromal cell proliferation. SEW2871 (L) treatment does not affect stromal cell proliferation compared to vehicle-treated control grafts. Arrows show proliferating stromal cell positively stained for PCNA. (M–R) Evaluation of ovarian tissue hypoxia by HIF1α.While S1P (P) treatment reduces tissue hypoxia compared to vehicle-treatment. FTY720 (Q) and SEW2871 (R) treatment is associated with massive tissue hypoxia 10 days post-grafting. (Q) Arrow shows hypoxic ovarian follicles in a FTY720-treated graft. (S) There is an inverse correlation between vascular density and hypoxic ovarian tissue surface area in the grafts, as determined by HIF1α staining. This indicates that enhanced vascular density reduces hypoxia in ovarian transplants.
Figure 3.
Impact of S1P on early angiogenesis after ovarian transplantation.
S1P-treated ovarian transplants show significantly higher density of microvasculature by as early as two days after transplantation. * Significantly different from matching vehicle-treated control. # Significantly different from baseline.
Figure 4.
Impact of S1P on follicle survival in frozen-thawed ovarian transplants.
Frozen-thawed ovarian tissue was transplanted to SCID mice and percentage of apoptotic follicles were compared between S1P-treated and vehicle-treated control grafts. (A–D) Histomorphological evaluation of apoptosis induced by cryopreservation and/or transplantation. The percentage of apoptotic follicles was significantly lower in fresh baseline ovarian tissue (A) compared to cryopreserved-untransplanted ovarian tissue (B) indicating that cryopreservation process itself induces apoptotic ovarian follicle death. S1P treatment results in significantly decrease in the density of apoptotic follicles (D) compared to the vehicle-treated group (C). Arrows indicate apoptotic ovarian follicles. (E) Inverse correlation of apoptotic follicle death with vascular density indicating that improved vascularization by S1P enhances follicle survival.