In vitro and in vivo evaluation of organic solvent-free injectable melatonin nanoformulations.

Melatonin is a neurohormone with potenial therapeutic effects in many diseases including neonatal hypoxic-ischemic (HI) brain injury. Due to limited solubility in water there is currently no clinically available melatonin formulation for parenteral use. Clinical use of melatonin has thus relied on oral administration, which in many cases is hampered by low and variable bioavailability. In animal treatment studies of neonatal HI, this issue have been circumvented by using parenteral administration of melatonin dissolved in ethanol (EtOH) or dimethyl sulfoxide (DMSO), solvents that are potentially neurotoxic, especially to the newborn brain. Thus, there is an urgent need for a non-toxic injectable melatonin formulation. The aim of this study was to develop such a formulation comprised of melatonin and biocompatible lipid-based nanoparticles with improved melatonin bioavailability. We herein report the development and characterization of an injectable system composed of melatonin and liposomes (LP) or oil-in-water nanoemulsions (NE). Nanoparticle characterization confirmed physicochemical stability over a week and an improvement with respect to melatonin solubilization in water (2.6 mg/mL in our injectable system). Determination of the in vitro release kinetics showed a prolonged release when melatonin is solubilized in nanoparticles (T1/2: 81 min vs 50 min vs 26 min for melatonin-LP, melatonin-NE, and melatonin-EtOH respectively). The pharmacokinetic (PK) parameters were confirmed in vivo in adult rats as similar melatonin levels detected in blood and indicated higher bioavailability in brain after intravenous administration of melatonin nanoformulations (10 mg/kg) in comparison to the free-melatonin administration. In conclusion, we have developed an organic solvent-free injectable formulation for melatonin by utilizing FDA-approved components, as a safe alternative for facilitating the potential of melatonin against variety of pathological conditions.

Effects of Neural Stem Cell and Olfactory Ensheathing Cell Co-transplants on Tissue Remodelling After Transient Focal Cerebral Ischemia in the Adult Rat.

Effective transplant-mediated repair of ischemic brain lesions entails extensive tissue remodeling, especially in the ischemic core. Neural stem cells (NSCs) are promising reparative candidates for stroke induced lesions, however, their survival and integration with the host-tissue post-transplantation is poor. In this study, we address this challenge by testing whether co-grafting of NSCs with olfactory ensheathing cells (OECs), a special type of glia with proven neuroprotective, immunomodulatory, and angiogenic effects, can promote graft survival and host tissue remodelling. Transient focal cerebral ischemia was induced in adult rats by a 60-min middle cerebral artery occlusion (MCAo) followed by reperfusion. Ischemic lesions were verified by neurological testing and magnetic resonance imaging. Transplantation into the globus pallidus of NSCs alone or in combination with OECs was performed at two weeks post-MCAo, followed by histological analyses at three weeks post-transplantation. We found evidence of extensive vascular remodelling in the ischemic core as well as evidence of NSC motility away from the graft and into the infarct border in severely lesioned animals co-grafted with OECs. These findings support a possible role of OECs as part of an in situ tissue engineering paradigm for transplant mediated repair of ischemic brain lesions.

Brain development after neonatal intermittent hyperoxia-hypoxia in the rat studied by longitudinal MRI and immunohistochemistry.

BACKGROUND: Neonatal intermittent hyperoxia-hypoxia (IHH) is involved in the pathogenesis of retinopathy of prematurity. Whether similar oxygen fluctuations will create pathological changes in the grey and white matter of the brain is unknown. METHODS: From birth until postnatal day 14 (P14), two litters (total n = 22) were reared in IHH: hyperoxia (50% O2) interrupted by three consecutive two-minute episodes of hypoxia (12% O2) every sixth hour. Controls (n = 8) were reared in room-air (20.9% O2). Longitudinal MRI (Diffusion Tensor Imaging and T2-mapping) was performed on P14 and P28 and retinal and brain tissue were examined for histopathological changes. Long-term neurodevelopment was assessed on P20 and P27. RESULTS: Mean, radial and axial diffusivity were higher in white matter of IHH versus controls at P14 (p < 0.04), while fractional anisotropy (FA) was lower in the hippocampal fimbria and tended to be lower in corpus callosum (p = 0.08) and external capsule (p = 0.05). White matter diffusivity in IHH was similar to controls at P28. Higher cortical vessel density (p = 0.005) was observed at P14. Cortical and thalamic T2-relaxation time and mean diffusivity were higher in the IHH group at P14 (p ≤ 0.03), and albumin leakage was present at P28. Rats in the IHH group ran for a longer time on a Rotarod than the control group (p ≤ 0.005). Pups with lower bodyweight had more severe MRI alterations and albumin leakage. CONCLUSION: IHH led to subtle reversible changes in brain white matter diffusivity, grey matter water content and vascular density. However, alterations in blood-brain barrier permeability may point to long-term effects. The changes seen after IHH exposure were more severe in animals with lower bodyweight and future studies should aim at exploring possible interactions between IHH and growth restriction.