P2D386: Comparative Effects on Fetal Hematopoiesis and Placental Inflammation from Mesenchymal and Hematopoietic Stem Cells as Agents of Transamniotic Stem Cell Therapy (TRASCET) in a Syngeneic Model of Intrauterine Growth Restriction

Background: Transamniotic stem cell therapy (TRASCET) with mesenchymal stem cells (MSCs) derived from the amniotic fluid or placenta has been shown experimentally to reverse or minimize fetal biometric, neurologic, and cardiopulmonary effects of intrauterine growth restriction (IUGR). Deficits of hematopoiesis are also relevant complications of IUGR as a result of placental insufficiency and inflammation driven by abnormal uterine natural killer cell (uNK) activity. We sought to determine the effects of TRASCET with either amniotic fluid-derived MSCs or amniotic fluid-derived hematopoietic stem cells (HSCs) on fetal hematopoiesis, as well as on downstream products of uNKs in a syngeneic model of IUGR.

Methods: After IACUC approval, eleven time-dated pregnant Lewis rat dams were exposed to alternating 12-hour hypoxia (10.5% O2) cycles during late gestation, from gestational day 15 (E15) until term (E21). Their fetuses (n=83) were divided in four groups. One group received no further interventions (untreated; n=9). Three groups underwent volume-matched (50μL) intra-amniotic injections on E17, via laparotomy, of suspensions of either HSCs (n=34), primed-MSCs (n=28), or saline (sham; n=12). Normal fetuses not exposed to hypoxia served as additional controls (n=18). All donor cells consisted of banked syngeneic Lewis rat cells isolated from the amniotic fluid. HSCs were positive for CD34, CD117, SSEA3, SSEA4, Sox2, Oct4, and alkaline phosphatase on flow cytometry. MSCs were positive for CD29, CD44, CD45, CD73, and CD90 on flow cytometry. MSC priming consisted of exposing cells to interferon-gamma (IFN-γ) and interleukin-1beta (IL-1β) for 24 hours prior to injection in order to enhance their anti-inflammatory capacity. Multiple analyses were performed at term, including peripheral blood counts and ELISA for placental levels of downstream products of uNKs, namely vascular endothelial growth factor (VEGF), basic fibroblast growth factor-2 (FGF-2), and transforming growth factor-alpha (TNF-α). Statistical analysis was by Wilcox-rank sum test with Bonferroni-adjustment (p < 0.05).

Results: Overall survival from hypoxia was 41% (34/83). Red blood count (RBC), hematocrit (Hct) and hemoglobin levels (Hb) were all significantly decreased from normal in all hypoxia groups (p < 0.001 for all). The primed-MSC group had significantly higher RBC, Hct, and Hb levels than sham (p=0.01-0.03; figure), though not higher than untreated (which had no blood loss from surgery). The HSC group had significantly higher Hb levels than sham (p=0.005), but no change in RBC or Hct. TRASCET with primed-MSC led to significantly lower placental levels of TNF-α than sham (p=0.04; figure), but not than untreated. No changes in any of the placental markers studied were noted in the HSC group.

Conclusion: Primed mesenchymal stem cells seem more effective than hematopoietic stem cells in improving hematopoiesis when used as donor cells for transamniotic stem cell therapy in a syngeneic model of IUGR. The same applies to their effects in decreasing placental inflammation in that model.