Histological evaluations of mesenchymal stem cell therapy in a preterm IVH rabbit model.

Abstract: Human mesenchymal stem cell (MSC) therapy has shown neuroprotective effects and improvement on recovery from neonatal intraventricular hemorrhage (IVH). This study focused on histological evaluations of human amniotic fluid MSC therapy during early prenatal life in a preterm IVH rabbit model. IVH was diagnosed at 24 h of age with ultrasound and animals were randomized into subgroups with confirmed IVH and an IVH negative control group. Animals with confirmed IVH received vehicle only or MSCs at two different doses via intraperitoneal administration. The animals were sacrificed at 48 h post administration. The severity of IVH was histologically analyzed via staining of endogenous peroxidase activity in cryosections and the distribution of red blood cells and cell-free hemoglobin was scored. Primary antibodies targeting human epitopes were validated in IHC assays of frozen MSC pellet. An anti-human nuclear mitotic apparatus (hNuMA) antibody labeled the majority of cells in the MSC pellet and did not cross-react with rabbit NuMA when tested in the nontreated rabbit brain. Significant levels of red blood cells and cell-free hemoglobin were found in the IVH confirmed group, whereas the control group showed no hemorrhage. The MSC therapy groups showed similar scoring results as the IVH-vehicle group. Anti-hNuMA immunolabeling did not detect any cells in the brain of MSC treated rabbits. However, extracellular (nonnuclear) immunolabeling was detected, located in the midbrain of the animals that received MSCs, indicating the presence of MSC nuclear debris. The preterm rabbit model was proven successful for inducing IVH, whereas MSC treatment did not affect the degree of hemorrhage. To increase the possibility to detect the MSCs post administration, future studies should include prelabeling of the MSCs with a suitable cell tracker and analyses at time points closer to the administration of MSCs.