Background: The germinal matrix (GM) is a site of proliferating neuronal & glial precursor cells & the origin of intraventricular hemorrhage (IVH) - a common complication of extreme prematurity leading to white matter injury & cerebral palsy. We previously showed that USSC administration resulted in recovery of white matter injury & improved locomotor function in a premature rabbit model of GMH-IVH (Vinukonda et al, Stem Cell Trans Med, 8:1157, 2019). Intrinsic signals from the GM & extrinsic signals from meninges, blood vessels and cerebrospinal fluid regulate cell proliferation & differentiation during normal brain development (Lehtinen, et al, 2011; Johansson, et al, 2010). We hypothesize that intracerebroventricular (ICV) USSC administration elicits regenerative potential by modulating expression of TGFβ1, FGF, and IGF1-2, enhancing progenitor cell proliferation & differentiation after IVH.
Objective: To test whether USSC administration alters the endogenous expression of growth factors: TGFβ1, FGF and IGF1-2 in IVH. Furthermore, to determine whether a change in growth factor expression after USSC infusion correlates with specific progenitor cell proliferation & differentiation, as well as recovery after white matter injury caused by IVH.
Design/Methods: A single dose of ICV USSCs (2x106) was injected in premature rabbits 1d after IP glycerol-induced IVH; naïve & non-treated IVH animals served as comparison groups. USSC effects were established from dissected ventricular ependyma, coronal slices and cerebrospinal fluid at postnatal days 3, 7 & 14.
Results: USSC survival and migration in vivo was confirmed using live animal bioluminescence imaging followed by immunostaining. We quantified total proliferating cells using Ki67-DAPI in the GM zone (GM- ventricular zone [VZ] & sub-VZ) of the lateral ventricles at 3d & observed decreased cell density in IVH (p<0.05), whereas with USSC infusion there was a trend towards recovery showing increased total proliferation. After USSC infusion, early oligodendrocyte precursor (PDGFR-Ki67 in SVZ) proliferation showed increased cell density (p<0.05) on day 3. Late oligodendrocyte precursor (Ki67-Olig2 in the corpus callosum & corona) showed increased cell density (p<0.05) on day 3 & 7. Known growth factors that regulate proliferation and maturation of progenitor cells were assessed (mRNA in SVZ dissected tissue & protein in CSF): TGFβ1, FGF & IGF1-2. mRNA expression was 37% lower for TGFβ1 & 47% for FGF in 3d IVH pups, whereas increased levels of mRNA for both occurred after USSC infusion (p<0.05). TGFβ1 protein levels in the CSF at 3d decreased in IVH pups (p<0.05) & increased after USSC administration (p<0.05). IGF1-2 levels were comparable between the groups at all timepoints.
Conclusion(s): USSCs removed the blockade to oligodendrocyte precursor cell division & favorably modulated TGFβ1 & FGF toward normal expression correlating with regenerative improvement after IVH.
