Human Cord Blood Derived Unrestricted Somatic Stem Cells Restore Aquaporin Channel Expression, Reduce Inflammation and Inhibit the Development of Hydrocephalus After Experimentally Induced Perinatal Intraventricular Hemorrhage.

Deepti Purohit,Furong Hu,Carl Thompson,Etlinger Joseph,George M Kleinman,Larisa Ivanova,Ana Malfa,Govindaiah Vinukonda,Michael S Wolin,Dina A Finkel,Edmund F La Gamma,Yanling Liao,Mitchell S Cairo,Shetal Shah

doi:10.3389/fncel.2021.633185

Abstract

Intraventricular hemorrhage (IVH) is a severe complication of preterm birth associated with cerebral palsy, intellectual disability, and commonly, accumulation of cerebrospinal fluid (CSF). Histologically, IVH leads to subependymal gliosis, fibrosis, and disruption of the ependymal wall. Importantly, expression of aquaporin channels 1 and 4 (AQP1 and AQP4) regulating respectively, secretion and absorption of cerebrospinal fluids is altered with IVH and are associated with development of post hemorrhagic hydrocephalus. Human cord blood derived unrestricted somatic stem cells (USSCs), which we previously demonstrated to reduce the magnitude of hydrocephalus, as having anti-inflammatory, and beneficial behavioral effects, were injected into the cerebral ventricles of rabbit pups 18 h after glycerol-induced IVH. USSC treated IVH pups showed a reduction in ventricular size when compared to control pups at 7 and 14 days (both, P < 0.05). Histologically, USSC treatment reduced cellular infiltration and ependymal wall disruption. In the region of the choroid plexus, immuno-reactivity for AQP1 and ependymal wall AQP4 expression were suppressed after IVH but were restored following USSC administration. Effects were confirmed by analysis of mRNA from dissected choroid plexus and ependymal tissue. Transforming growth factor beta (TGF-β) isoforms, connective tissue growth factor (CTGF) and matrix metalloprotease-9 (MMP-9) mRNA, as well as protein levels, were significantly increased following IVH and restored towards normal with USSC treatment (P < 0.05). The anti-inflammatory cytokine Interleukin-10 (IL-10) mRNA was reduced in IVH, but significantly recovered after USSC injection (P < 0.05). In conclusion, USSCs exerted anti-inflammatory effects by suppressing both TGF-β specific isoforms, CTGF and MMP-9, recovered IL-10, restored aquaporins expression towards baseline, and reduced hydrocephalus. These results support the possibility of the use of USSCs to reduce IVH consequences in prematurity.

Highlights

Intraventricular hemorrhage (IVH) arises from rupture of immature and developing blood vessels in the germinal matrix of premature infants
To confirm that the attenuated ventricular area was the functional effect of Unrestricted somatic stem cells (USSC) rather than the glycerol injected in the peritoneal cavity, we assessed ventricular cross-sectional area measurements among healthy controls vs. glycerol injected but no IVH rabbit pups on postnatal day 14 (Supplementary Figures 3A–D)
After Intracerebral Ventricular (ICV) USSC administration, there was a significant increase on postnatal day 3 above control and IVH groups (100 ± 11 in control, 24.59 ± 7 in IVH and 165 ± 41 IVH USSC; Figure 8E, P < 0.05). This is the first study evaluating the effects of USSCs on the density of aquaporin water channels 1 and 4 in the choroid plexus epithelium and lateral ventricular ependymal wall after IVH-induced hydrocephalus

Summary

Introduction

Intraventricular hemorrhage (IVH) arises from rupture of immature and developing blood vessels in the germinal matrix of premature infants. A recent NIH workshop on post-hemorrhagic hydrocephalus (PHH) reported that IVH affected neonates develop somatic growth impairment, white matter damage, motor dysfunction and neurocognitive deficiency (Koschnitzky et al, 2018). IVH is associated with prolonged neonatal hospitalization and increased lifelong medical care costs (Christian et al, 2016). Post hemorrhagic hydrocephalus (PHH) arises from either over secretion or impaired absorption of cerebrospinal fluid (CSF). The latter is due in part to blood obstructing arachnoid villi associated with fibro-proliferative responses, inflammation and sub-ependymal gliosis. AQP1 and AQP4 are associated with CSF hypersecretion and reduced absorption respectively (Verkman et al, 2017). AQP1 is expressed primarily along the choroid plexus epithelial lining and, in knockout studies using mice, the loss of AQP1 function reduced CSF production and intraventricular pressure (Oshio et al, 2005; Trillo-Contreras et al, 2019)

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