Combination treatment of stroke with sub-therapeutic doses of Simvastatin and human umbilical cord blood cells enhances vascular remodeling and improves functional outcome.

Human umbilical cord blood cells (HUCBCs) have been employed as a restorative treatment for experimental stroke. In this study, we investigated whether transplantation of sub-therapeutic doses of HUCBCs and Simvastatin enhances cerebral vascular remodeling after stroke. Adult male Wistar rats (n=34) were subjected to transient middle cerebral artery occlusion (MCAo) and treated with: phosphate-buffered solution (PBS, gavaged daily for 7 days); Simvastatin (0.5mg/kg, gavaged daily for 7 days); HUCBCs (1×10(6), injected once via tail vein); and combination Simvasatin with HUCBCs, starting at 24h after MCAo. There was no significant difference between Simvastatin- or HUCBC-monotherapy and MCAo-alone group. Combination treatment 24h post-stroke significantly increased the perimeter of von Willebrand factor (vWF)-positive vessels, the diameter and density of alpha smooth muscle actin (αSMA)-positive arteries, and the percentage of 5-bromodeoxyuridine (BrdU)-positive endothelial cells (ECs) in the ischemic boundary zone (IBZ) compared with MCAo-alone or HUCBC-monotherapy 14 days after MCAo (p<0.05, n=8/group); Combination treatment significantly increased the densities of vWF-vessels and αSMA-arteries as well as the densities of BrdU-ECs and BrdU-positive smooth muscle cells (SMCs) in vascular walls in the IBZ compared with Simvastatin-monotherapy. Moreover, the increased BrdU-ECs and BrdU-SMCs were significantly correlated with neurological functional outcome 14 days after MCAo. Combination treatment also significantly increased the expression of Angiopoietin-1 (Ang1), Tie2 and Occludin in the IBZ (p<0.05, n=8/group). The in vitro experiments showed that combination treatment and Ang1 significantly increased capillary-like tube formation and arterial cell migration; anti-Ang1 significantly reduced combination treatment-induced tube-formation and artery cell migration (p<0.05, n=6/group). These findings indicated that a combination of sub-therapeutic doses of Simvastatin and HUCBCs treatment of stroke increases Ang1/Tie2 and Occludin expression in the ischemic brain, amplifies endogenous angiogenesis and arteriogenesis, and enhances vascular remodeling which in concert may contribute to functional outcome after stroke.

Mannitol facilitates neurotrophic factor up-regulation and behavioural recovery in neonatal hypoxic-ischaemic rats with human umbilical cord blood grafts.

We recently demonstrated that blood-brain barrier permeabilization using mannitol enhances the therapeutic efficacy of systemically administered human umbilical cord blood (HUCB) by facilitating the entry of neurotrophic factors from the periphery into the adult stroke brain. Here, we examined whether the same blood-brain barrier manipulation approach increases the therapeutic effects of intravenously delivered HUCB in a neonatal hypoxic-ischaemic (HI) injury model. Seven-day-old Sprague-Dawley rats were subjected to unilateral HI injury and then at day 7 after the insult, animals intravenously received vehicle alone, mannitol alone, HUCB cells (15k mononuclear fraction) alone or a combination of mannitol and HUCB cells. Behavioural tests at post-transplantation days 7 and 14 showed that HI animals that received HUCB cells alone or when combined with mannitol were significantly less impaired in motor asymmetry and motor coordination compared with those that received vehicle alone or mannitol alone. Brain tissues from a separate animal cohort from the four treatment conditions were processed for enzyme-linked immunosorbent assay at day 3 post-transplantation, and revealed elevated levels of GDNF, NGF and BDNF in those that received HUCB cells alone or when combined with mannitol compared with those that received vehicle or mannitol alone, with the combined HUCB cells and mannitol exhibiting the most robust neurotropic factor up-regulation. Histological assays revealed only sporadic detection of HUCB cells, suggesting that the trophic factor-mediated mechanism, rather than cell replacement per se, principally contributed to the behavioural improvement. These findings extend the utility of blood-brain barrier permeabilization in facilitating cell therapy for treating neonatal HI injury.