Glial responses to neonatal hypoxic–ischemic injury in the rat cerebral cortex

Neurogenesis is nearly completed after birth, whereas gliogenic activities remain intense during the postnatal period in the developing rat cortex. These include involution of radial glia, proliferation of astrocytes and oligodendrocytes and myelin formation. Little is known about the effects of hypoxic–ischemic (HI) injury on these critical postnatal processes. Here we explored the glial reactions to mild HI injury of the neonatal rat cerebral cortex at P3. We show that the HI lesion results in disruption of the normal radial glia architecture, which was paralleled by an increase in GFAP immunopositive reactive astrocytes. The morphology of these latter cells and the fact that they were immunolabelled for both nestin and GFAP suggest an accelerated transformation of radial glia into astrocytes. In addition, BrdU/GFAP immunostaining revealed a significant increase of double-labelled cells indicating an acute proliferation of astrocytes after HI. This enhanced proliferative activity of astrocytes persisted for several weeks. We found an elevated number and increased mitotic activity of both NG2-positive oligodendrocyte progenitors and RIP-positive oligodendrocytes after injury. These findings imply that glial responses are central to cortical tissue remodelling following neonatal ischemia and represent a potential target for therapeutic approaches.     

Late measures of microstructural alterations in severe neonatal hypoxic–ischemic encephalopathy by MR diffusion tensor imaging

Neonatal hypoxic–ischemic encephalopathy is a major cause of brain damage in infants, and is associated with periventricular white matter injury and chronic neurological dysfunctions. However, the mechanisms of the chronic white matter injury and reorganization are still unclear. In this study, in vivo diffusion tensor imaging (DTI) was employed to evaluate the late changes of white matter microstructural integrity in the rat brains at 10 weeks after severe neonatal hypoxic–ischemic insults at postnatal day 7. In the fractional anisotropy directionality map, qualitative evaluation showed that a dorsoventrally oriented fiber bundle extended from the corpus callosum into the cyst in the anterior brain, whilst the posterior peri-infarct areas had similar fiber orientations as the contralateral internal capsule, optic tract and fimbria of hippocampus. Compared to the contralateral hemisphere, significantly higher fractional anisotropy, axial diffusivity and diffusion trace value were observed quantitatively in the distal end of the extended fiber bundle connecting the anterior and posterior white matters rostrocaudally. A significantly lower fractional anisotropy but higher axial and radial diffusivities and trace were also found in the ipsilateral corpus callosum, proximal external capsule and anterior commissure, while slightly lower fractional anisotropy and axial diffusivity were noticed in the ipsilateral internal capsule and optic nerve. It was suggested that increased fractional anisotropy, axial diffusivity and trace characterize white matter reorganization in chronic neonatal hypoxic–ischemic insults, whereas reduction in fractional anisotropy appears to characterize two types of white matter lesions, with significantly higher axial and radial diffusivities and trace being primary and slightly lower axial diffusivity being secondary. Combined with fractional anisotropy directionality map, in vivo DTI provides important indices to differentiate the chronic effects of severe neonatal hypoxic–ischemic injury and recovery globally, quantitatively and non-invasively.     

Time course of NAA T2 and ADC(w) in ischaemic stroke patients: 1H MRS imaging and diffusion-weighted MRI

Background and purpose: Proton spectroscopy and quantitative diffusion-weighted imaging (DWI) were used to investigate the pertinence of N-acetyl aspartate (NAA) as a reliable marker of neuronal density in human stroke. Methods: The time courses of tissue water apparent diffusion coefficient (ADC_w) and metabolite T2 were investigated on a plane corresponding to the largest area of cerebral infarction, within and outside the site of infarction in 71 patients with a large cortical middle cerebral artery territory infarction. Results: Significant reductions are seen in NAA T2 deep within the infarction during the period comprised between 5 and 20 days postinfarction; the relaxation times appearing to normalise several months after stroke. After an acute reduction in ADC_w, the pseudonormalisation of ADC_w occurs at 8–12 days after the ischaemic insult. The minimum in N-acetyl aspartate T2 relaxation times and the pseudonormalisation of ADC_w appear to coincide. Conclusions: The data suggest that modifications in the behaviour of the observed proton metabolites occur during the period when the vasogenic oedema is formed and cell membrane integrity is lost. For this reason, NAA may not be a reliable marker of neuronal density during this period.     

Proteomic analysis of the effects of caffeine in a neonatal rat model of hypoxic‐ischemic white matter damage

AimWhite matter damage (WMD) is the main cause of cerebral palsy and cognitive impairment in premature infants. Although caffeine has been shown to possess neuroprotective effects in neonatal rats with hypoxic‐ischemic WMD, the mechanisms underlying these protective effects are unclear. Herein, proteins modulated by caffeine in neonatal rats with hypoxic‐ischemic WMD were evaluated.MethodsWe identified differential proteins and performed functional enrichment analyses between the Sham, hypoxic‐ischemic WMD (HI), and HI+caffeine‐treated WMD (Caffeine) groups. Confirmed the changes and effect of proteins in animal models and determined cognitive impairment via water maze experiments.ResultsIn paraventricular tissue, 47 differential proteins were identified between the Sham, HI, and Caffeine groups. Functional enrichment analyses showed that these proteins were related to myelination and axon formation. In particular, the myelin basic protein (MBP), proteolipid protein, myelin‐associated glycoprotein precursor, and sirtiun 2 (SIRT2) levels were reduced in the hypoxic‐ischemic WMD group, and this effect could be prevented by caffeine. Caffeine alleviated the hypoxic‐ischemic WMD‐induced cognitive impairment and improved MBP, synaptophysin, and postsynaptic density protein 95 protein levels after hypoxic‐ischemic WMD by preventing the HI‐induced downregulation of SIRT2; these effects were subsequently attenuated by the SIRT2 inhibitor AK‐7.ConclusionCaffeine may have clinical applications in the management of prophylactic hypoxic‐ischemic WMD; its effects may be mediated by proteins related to myelin development and synapse formation through SIRT2.     

A method to remove the influence of fixative concentration on postmortem T2 maps using a kinetic tensor model

Formalin fixation has been shown to substantially reduce T₂ estimates, primarily driven by the presence of fixative in tissue. Prior to scanning, post‐mortem samples are often placed into a fluid that has more favourable imaging properties. This study investigates whether there is evidence for a change in T₂ in regions close to the tissue surface due to fixative outflux into this surrounding fluid. Furthermore, we investigate whether a simulated spatial map of fixative concentration can be used as a confound regressor to reduce T₂ inhomogeneity. To achieve this, T₂ maps and diffusion tensor estimates were obtained in 14 whole, formalin‐fixed post‐mortem brains placed in Fluorinert approximately 48 hr prior to scanning. Seven brains were fixed with 10% formalin and seven brains were fixed with 10% neutral buffered formalin (NBF). Fixative outflux was modelled using a proposed kinetic tensor (KT) model, which incorporates voxelwise diffusion tensor estimates to account for diffusion anisotropy and tissue‐specific diffusion coefficients. Brains fixed with 10% NBF revealed a spatial T₂ pattern consistent with modelled fixative outflux. Confound regression of fixative concentration reduced T₂ inhomogeneity across both white and grey matter, with the greatest reduction attributed to the KT model versus simpler models of fixative outflux. No such effect was observed in brains fixed with 10% formalin. Correlations between the transverse relaxation rate R ₂ and ferritin/myelin proteolipid protein (PLP) histology lead to an increased similarity for the relationship between R ₂ and PLP for the two fixative types after KT correction.     

Aluminum chloride does not facilitate deposition of human synthetic amyloid β1–42 peptide in the rat ventricular system of a short‐term infusion model

Recent data have demonstrated that a 39–43 amino acid peptide called β‐amyloid peptide (Aβ), which predominantly contains 42 residues (Aβ1–42), has a strong tendency to form insoluble aggregates and that the toxic effects of Aβ are based on its aggregation. In a previous study, we reported that infusion of 100 µg of human synthetic Aβ1–42 (sAβ1–42), which is a main component of diffuse plaques, into the lateral ventricle of the rat brain of a short‐term infusion model resulted in almost complete disappearance of sAβ1–42 aggregates from the ventricles by 28 days. In addition, aluminum is considered a potential etiological factor in Alzheimer's disease (AD). Neurotoxicity from excess brain exposure to aluminum has been documented from both clinical observations and animal experiments, although a direct relationship between aluminum and AD has yet to be clearly established. We therefore investigated the effects of sAβ1–42 aggregates with aluminum chloride (AlCl₃) in the ventricular system of the rat brain of a short‐term infusion model. At either 2 or 7 days following infusion, sAβ1–42 formed aggregates with AlCl₃ that spread throughout the entire ventricular system. However, sAβ1–42 aggregates with AlCl₃ had almost disappeared from the ventricles by 28 days, resulting in similarities with respect to the time‐course and the neuropathological changes observed in sAβ1–42 aggregation without AlCl₃. We herein report for the first time that considerable amounts of sAβ1–42 aggregates with AlCl₃ almost disappear from the rat ventricular system by 28 days post‐infusion.