Regional Heterogeneity of Cerebral Microvessels and Brain Susceptibility to Oxidative Stress

The hippocampus is one of the earliest and most affected regions in Alzheimer’s disease (AD), followed by the cortex while the cerebellum is largely spared. Importantly, endothelial dysfunction is a common feature of cerebral blood vessels in AD. In this study, we sought to determine if regional heterogeneity of cerebral microvessels might help explain the susceptibility of the hippocampus and cortex as compared to the cerebellum. We isolated microvessels from wild type mice from the cerebellum, cortex, and hippocampus to characterize their vascular phenotype. Superoxide anion was significantly higher in microvessels isolated from the cortex and hippocampus as compared to the cerebellum. Importantly, protein levels of NADPH oxidase (NOX)-2 and NOX-4 were significantly higher in the cortical and hippocampal microvessels as compared to microvessels from the cerebellum. In addition, expression of manganese superoxide dismutase protein was significantly lower in microvessels from the cortex and hippocampus as compared to cerebellum while other antioxidant enzymes were unchanged. There was no difference in eNOS protein expression between the microvessels of the three brain regions; however, bioavailability of tetrahydrobiopterin (BH₄), an essential cofactor for eNOS activity, was significantly reduced in microvessels from the hippocampus and cortex as compared to the cerebellum. Higher levels of superoxide and reduced tetrahydrobiopterin bioavailability may help explain the vulnerability of the hippocampus and cortical microvessels to oxidative stress and development of endothelial dysfunction.     

Increased brain levels of 4-hydroxy-2-nonenal glutathione conjugates in severe Alzheimer's disease

In the last decade an important role for the progression of neuronal cell death in Alzheimer's disease (AD) has been ascribed to oxidative stress. trans-4-Hydroxy-2-nonenal, a product of lipid peroxidation, forms conjugates with a variety of nucleophilic groups such as thiols or amino moieties. Here we report for the first time the quantitation of glutathione conjugates of trans-4-hydroxy-2-nonenal (HNEGSH) in the human postmortem brain using the specific and very sensitive method of electrospray ionization triple quadrupole mass spectrometry (ESI-MS-MS). Levels of HNEGSH conjugates calculated as the sum of three chromatographically separated diastereomers were determined in hippocampus, entorhinal cortex, substantia innominata, frontal and temporal cortex, as well as cerebellum from patients with AD and controls matched for age, gender, postmortem delay and storage time. Neither age, nor postmortem delay, nor storage time did correlate with levels of HNEGSH conjugates which ranged between 1 and 500 pmol/g fresh weight in the brain areas examined. The brain specimen from patients with clinically and neuropathologically probable AD diagnosed according to criteria of the consortium to establish a registry for AD (CERAD) show increased levels of HNEGSH in the temporal and frontal cortex, as well as in the substantia innominata. Classification of disease severity according to Braak and Braak, which takes into consideration the amount of neurofibrillary tangles and neuritic plaques, revealed highest levels of HNEGSH in the substantia innominata and the hippocampus, two brain regions known to be preferentially affected in AD. These results substantiate the link between conjugates of glutathione with a product of lipid peroxidation and Alzheimer's disease and justify further studies to evaluate the role of HNE metabolites as potential biomarkers for disease progression in AD.     

Regional distribution and relative amounts of glutamate decarboxylase isoforms in rat and mouse brain.

The levels of the two isoforms of glutamate decarboxylase (GAD) were measured in 12 regions of adult rat brain and three regions of mouse brain by sodium dodecylsulfate-polyacrylamide gel electrophoresis and immunoblotting with an antiserum that recognizes the identical C-terminal sequence in both isoforms from both species. In rat brain the amount of smaller isoform, GAD65, was greater than that of the larger isoform, GAD67, in all twelve regions. GAD65 ranged from 77-89% of total GAD in frontal cortex, hippocampus, hypothalamus, midbrain, olfactory bulb, periaqueductal gray matter, substantia nigra, striatum, thalamus and the ventral tegmental area. The proportion of GAD65 was lower in amygdala and cerebellum but still greater than half of the total. There was a strong correlation between total GAD protein and GAD activity. In the three mouse brain regions analysed (cerebellum, cerebral cortex and hippocampus) the proportion of GAD65 (35,47, and 51% of total GAD) was significantly lower than in the corresponding rat-brain regions. The amount of GAD67 was greater than the amount of GAD65 in mouse cerebellum and was approximately equal to the amount of GAD65 in mouse cerebral cortex and hippocampus.     

Human brain nucleoside diphosphate kinase activity is decreased in Alzheimer's disease and Down syndrome

In brain, nucleoside diphosphate kinase (NDPK) and its coding gene, nm23, have been implicated to modulate neuronal cell proliferation, differentiation, and neurite outgrowth. However, a role of NDPK in neurodegenerative diseases has not been reported yet. Using proteomics techniques, we evaluated the protein levels of NDPK-A in seven brain regions from patients with Alzheimer's disease (AD) and Down syndrome (DS) showing AD-like neuropathology. NDPK-A was significantly decreased in brain regions (frontal, occipital, and parietal cortices) of both disorders. Due to the limitation of brain samples, the activity of NDPK was measured in three brain regions (frontal cortex, temporal cortex, and cerebellum). The specific activity of NDPK was significantly decreased in AD (frontal cortex) and DS (frontal and temporal cortices). Since NDPK-B could also drive the activity of NDPK, protein expression levels of both NDPK-A and NDPK-B were studied in frontal cortex by Western blot analysis. NDPK-A was significantly decreased in AD, which was consistent with the results of proteomics. However, NDPK-A was slightly decreased in DS and protein expression levels of NDPK-B in both DS and AD were moderately decreased, without reaching statistical significance. We propose that oxidative modification of NDPK could lead to the decreased activity of NDPK and, subsequently, influence several neuronal functions in neurodegenerative diseases as multifunctional enzyme through several mechanisms.     

Heat-Shock 70 Messenger RNA Levels in Human Brain: Correlation with Agonal Fever

Abstract: Systematic review of antemortem clinical information on randomly selected Alzheimer disease (AD) patients revealed that ∼40% of the patients had a recorded fever of ≥39.2°C at or near death. Using isolation and quantitation techniques appropriate for analysis of human brain mRNAs, we found that low levels of inducible heat-shock protein 70 (hsp70) mRNAs were present in cerebellum of afebrile AD patients and that mRNA levels were usually lower in two brain regions affected in AD, i.e., hippocampus and temporal cortex. Levels of hsp70 mRNAs were increased three- to 33-fold in cerebellum of febrile patients compared with levels in patients whose recorded temperatures were ≤37.5°C. Levels of hsp70 mRNAs were also increased in hippocampus and cortex of these febrile patients, but to a lesser extent than cerebellum. Heat-shock cognate 70 (hsc70) mRNAs were present at highest levels in afebrile cerebellum and were also present in the other brain regions. In cerebellum of patients with the highest temperatures, hsc70 mRNAs were induced severalfold over basal levels. Although there was a low and variable induction of hsc70 mRNAs in temporal cortex of these patients, there was no evidence for any induction in hippocampus. Increased heat-shock 70 mRNA levels did not correlate with hypoxia, coma, hypertension, hypoglycemia, seizures, or medication. These results indicate that a specific agonal stress, namely fever, can increase the levels of heat shock 70 mRNAs in AD brain; however, there is no evidence to suggest that affected regions of AD brain have higher overall levels of these mRNAs. Failure to obtain adequate agonal state information could result in inaccurately identifying short-term stress-related changes in postmortem brain as neuropathology characteristic of a chronic disease state.     

Selective reduction of hydroperoxyeicosatetraenoic acids to their hydroxy derivatives by apolipoprotein D: implications for lipid antioxidant activity and Alzheimer's disease

ApoD (apolipoprotein D) is up-regulated in AD (Alzheimer's disease) and upon oxidative stress. ApoD inhibits brain lipid peroxidation in vivo, but the mechanism is unknown. Specific methionine residues may inhibit lipid peroxidation by reducing radical-propagating L-OOHs (lipid hydroperoxides) to non-reactive hydroxides via a reaction that generates MetSO (methionine sulfoxide). Since apoD has three conserved methionine residues (Met(49), Met(93) and Met(157)), we generated recombinant proteins with either one or all methionine residues replaced by alanine and assessed their capacity to reduce HpETEs (hydroperoxyeicosatetraenoic acids) to their HETE (hydroxyeicosatetraenoic acid) derivatives. ApoD, apoD(M49-A) and apoD(M157-A) all catalysed the reduction of HpETEs to their corresponding HETEs. Amino acid analysis of HpETE-treated apoD revealed a loss of one third of the methionine residues accompanied by the formation of MetSO. Additional studies using apoD(M93-A) indicated that Met(93) was required for HpETE reduction. We also assessed the impact that apoD MetSO formation has on protein aggregation by Western blotting of HpETE-treated apoD and human brain samples. ApoD methionine oxidation was associated with formation of apoD aggregates that were also detected in the hippocampus of AD patients. In conclusion, conversion of HpETE into HETE is mediated by apoD Met(93), a process that may contribute to apoD antioxidant function.     

Structure-Related Oxidative Damage in Rat Brain After Acute and Chronic Electroshock

The role of oxidative stress in electroconvulsive therapy-related effects is not well studied. The purpose of this study was to determine oxidative stress parameters in several brain structures after a single electroconvulsive seizure or multiple electroconvulsive seizures. Rats were given either a single electroconvulsive shock or a series of eight electroconvulsive shocks. Brain regions were isolated, and levels of oxidative stress in the brain tissue (cortex, hippocampus, striatum and cerebellum) were measured. We demonstrated a decrease in lipid peroxidation and protein carbonyls in the hippocampus, cerebellum, and striatum several times after a single electroconvulsive shock or multiple electroconvulsive shocks. In contrast, lipid peroxidation increases both after a single electroconvulsive shock or multiple electroconvulsive shocks in cortex. In conclusion, we demonstrate an increase in oxidative damage in cortex, in contrast to a reduction of oxidative damage in hippocampus, striatum, and cerebellum.     

Increased expression of the lysosomal cholesterol transporter NPC1 in Alzheimer's disease

The Niemann-Pick type C1 (NPC1) protein mediates the trafficking of cholesterol from lysosomes to other organelles. Mutations in the NPC1 gene lead to the retention of cholesterol and other lipids in the lysosomal compartment, and such defects are the basis of NPC disease. Several parallels exist between NPC disease and Alzheimer's disease (AD), including altered cholesterol homeostasis, changes in the lysosomal system, neurofibrillary tangles, and increased amyloid-beta generation. How the expression of NPC1 in the human brain is affected in AD has not been investigated so far. In the present study, we measured NPC1 mRNA and protein expression in three distinct regions of the human brain, and we revealed that NPC1 expression is upregulated at both mRNA and protein levels in the hippocampus and frontal cortex of AD patients compared to control individuals. In the cerebellum, a brain region that is relatively spared in AD, no difference in NPC1 expression was detected. Similarly, murine NPC1 mRNA levels were increased in the hippocampus of 12-month-old transgenic mice expressing a familial AD form of human amyloid-beta precursor protein (APP) and presenilin-1 (APP/PS1tg) compared to 12-month-old wild type mice, whereas no change in NPC1 was detected in mouse cerebellum. Immunohistochemical analysis of human hippocampus indicated that NPC1 expression was strongest in neurons. However, in vitro studies revealed that NPC1 expression was not induced by transfecting SK-N-SH neurons with human APP or by treating them with oligomeric amyloid-beta peptide. Total cholesterol levels were reduced in hippocampus from AD patients compared to control individuals, and it is therefore possible that the increased expression of NPC1 is linked to perturbed cholesterol homeostasis in AD.     

Effects of acute manganese chloride exposure on lipid peroxidation and alteration of trace metals in rat brain

Although manganese (Mn) is an essential element, exposure to excessive levels of Mn and its accumulation in the brain can cause neurotoxicity and extrapyramidal syndrome. We have investigated the differences in the accumulated levels of Mn, the degree of lipid peroxidation, and its effects on the levels of trace elements (Fe, Cu, and Zn) in various regions in the brain of rats having undergone acute Mn exposure. The rats in the dose—effect group were injected intraperitoneally (ip) with MnCl₂ (25, 50, or 100 mg MnCl₂/kg) once a day for 24 h. The Mn significantly accumulated (p<0.05) in the frontal cortex, corpus callosum, hippocampus, striatum, hypothalamus medulla, cerebellum, and spinal cord in each case. The rats in the timecourse group were ip injected with MnCl₂ (50 mg MnCl₂/kg) and then monitored 12, 24, 48, and 72 h after exposure. The Mn accumulated in the frontal cortex, corpus callosum, hippocampus, striatum hypothalamus, medulla, cerebellum, and spinal cord after these periods of time, In both the dose—effect and time-course studies, we observed that the concentration of malondialdehyde, an end product of lipid peroxidation, increased significantly in the frontal cortex, hippocampus, striatum, hypothalamus, medulla, and cerebellum. However, no relationship between the concentrations of Mn in the brain and the extent of lipid peroxidation was observed. In addition, we found that there was a significant increase (p<0.05) in the level of Fe in the hippocampus, striatum, hypothalamus, medulla, and cerebellum, but the Cu and Zn levels had not changed significantly. These findings indicated that Mn induces an increase in the iron level, which provides direct evidence for Fe-mediated lipid peroxidation in the rats' brains; these phenomena might play important roles in the mechanisms of Mn-induced neurotoxicology.     

Reduced Glucose Transporter at the Blood-Brain Barrier and in Cerebral Cortex in Alzheimer Disease

We studied the hexose transporter protein of the frontal and temporal neocortex, hippocampus, putamen, cerebellum, and cerebral microvessels (which constitute the blood-brain barrier) in Alzheimer disease and control subjects by reversible and covalent binding with [3H]cytochalasin B and by immunological reactivity. In Alzheimer disease subjects, we found a marked decrease in the hexose transporter in brain microvessels and in the cerebral neocortex and hippocampus, regions that are most affected in Alzheimer disease, but there were no abnormalities in the putamen or cerebellum. Hexose transporter reduction in cerebral microvessels of Alzheimer subjects is relatively specific because other enzyme markers of brain endothelium were not significantly altered. The low density of the hexose transporter at the blood-brain barrier and in the cerebral cortex in Alzheimer disease may be related to decreased in vivo measurements of cerebral oxidative metabolism.     

The Posttranslational Arginylation of Proteins in Different Regions of the Rat Brain

The posttranslational incorporation of arginine into proteins catalyzed by arginyl-tRNA protein transferase was determined in vitro in different rat brain regions. The incorporation was found in all the regions studied, although with different specific activities (pmol [14C]arginine incorporated/mg protein). Of the regions studied, hippocampus had the highest specific activity followed by striatum, medulla oblongata, cerebellum, and cerebral cortex. Electrophoretic analysis of the [14C]arginyl proteins from the different regions followed by autoradiography and scanner densitometry showed at least 13 polypeptide bands that were labeled with [14C]arginine. The radioactive bands were qualitatively coincident with protein bands revealed by Coomassie Blue. There were peaks that showed different proportions of labeling in comparison with peaks of similar molecular mass from total brain. Most notable because of their high proportions were those of molecular mass 125 kDa in hippocampus, striatum, and cerebral cortex; 112 and 98 kDa in striatum and cerebellum; and 33 kDa in hippocampus and striatum. In lower proportions than in total brain were the peaks of 33 kDa in medulla oblongata and cerebral cortex and of 125 kDa in medulla oblongata.     

Increased Expression of Apolipoprotein D Following Experimental Traumatic Brain Injury

Increasing evidence suggests that apolipoprotein D (apoD) could play a major role in mediating neuronal degeneration and regeneration in the CNS and the PNS. To investigate further the temporal pattern of apoD expression after experimental traumatic brain injury in the rat, male Sprague-Dawley rats were subjected to unilateral cortical impact injury. The animals were killed and examined for apoD mRNA and protein expression and for immunohistological analysis at intervals from 15 min to 14 days after injury. Increased apoD mRNA and protein levels were seen in the cortex and hippocampus ipsilateral to the injury site from 48 h to 14 days after the trauma. Immunohistological investigation demonstrated a differential pattern of apoD expression in the cortex and hippocampus, respectively: Increased apoD immunoreactivity in glial cells was detected from 2 to 3 days after the injury in cortex and hippocampus. In contrast, increased expression of apoD was seen in cortical and hippocampal neurons at later time points following impact injury. Concurrent histopathological examination using hematoxylin and eosin demonstrated dark, shrunken neurons in the cortex ipsilateral to the injury site. In contrast, no evidence of cell death was observed in the hippocampus ipsilateral to the injury site up to 14 days after the trauma. No evidence of increased apoD mRNA or protein expression or neuronal pathology by hematoxylin and eosin staining was detected in the contralateral cortex and hippocampus. Our results reveal induction of apoD expression in the cortex and hippocampus following traumatic brain injury in the rat. Our data also suggest that increased apoD expression may play an important role in cortical neuronal degeneration after brain injury in vivo. However, increased expression of apoD in the hippocampus may not necessarily be indicative of neuronal death.     

Postnatal Development of Cholecystokinin-Like Immunoreactivity and Its mRNA Level in Rat Brain Regions

Developmental changes of preprocholecystokinin mRNA (CCK mRNA) and cholecystokinin-like immunoreactivity (CCK-LI) were examined in rat brain regions (frontal cortex, colliculi, hippocampus, striatum, and cerebellum) using RNA dot blot assays with cholecystokinin (CCK) cDNA and radioimmunoassay, respectively. The CCK-LI levels in all regions examined were very low at birth. Excluding the cerebellum, the levels in these regions increased postnatally and reached adult values at 28 days of age. In contrast to CCK-LI, CCK mRNA levels changed dramatically during development. A considerable amount of CCK mRNA was detected in the frontal cortex and hippocampus at birth. The changes in the level of CCK mRNA in the frontal cortex and colliculi paralleled those of CCK-LI, including a rapid increase from 7 to 14 days of age. The synthesis of CCK mRNA preceded the appearance of CCK-LI. CCK mRNA levels in the hippocampus and striatum exhibited a transient increase, with a peak at 14 days of age. In the adult brain, the CCK mRNA levels were high in the frontal cortex, moderate in the hippocampus and colliculi, and low in the striatum. The cerebellum contained only a negligible amount of CCK mRNA during development. The relatively high level of CCK-LI compared with the low level of CCK mRNA in the striatum supports the idea that most of the striatal CCK-LI is supplied from extrastriatal regions.(ABSTRACT TRUNCATED AT 250 WORDS)     

Novel Kv3 glycoforms differentially expressed in adult mammalian brain contain sialylated N-glycans.

The N-glycan pool of mammalian brain contains remarkably high levels of sialylated N-glycans. This study provides the first evidence that voltage-gated K+ channels Kv3.1, Kv3.3, and Kv3.4, possess distinct sialylated N-glycan structures throughout the central nervous system of the adult rat. Electrophoretic migration patterns of Kv3.1, Kv3.3, and Kv3.4 glycoproteins from spinal cord, hypothalamus, thalamus, cerebral cortex, hippocampus, and cerebellum membranes digested with glycosidases were used to identify the various glycoforms. Differences in the migration of Kv3 proteins were attributed to the desialylated N-glycans. Expression levels of the Kv3 proteins were highest in cerebellum, whereas those of Kv3.1 and Kv3.3 were much lower in the other 5 regions. The lowest level of Kv3.1 was expressed in the hypothalamus, whereas the lowest levels of Kv3.3 were expressed in both thalamus and hypothalamus. The other regions expressed intermediate levels of Kv3.3, with spinal cord expressing the highest. The expression level of Kv3.4 in the hippocampus was slightly lower than that in cerebellum, and was closely followed by the other 4 regions, with spinal cord expressing the lowest level. We suggest that novel Kv3 glycoforms may endow differences in channel function and expression among regions throughout the central nervous system.     

Parallel increases in lipid and protein oxidative markers in several mouse brain regions after methamphetamine treatment

The neurotoxic actions of methamphetamine (METH) may be mediated in part by reactive oxygen species (ROS). Methamphetamine administration leads to increases in ROS formation and lipid peroxidation in rodent brain; however, the extent to which proteins may be modified or whether affected brain regions exhibit similar elevations of lipid and protein oxidative markers have not been investigated. In this study we measured concentrations of TBARs, protein carbonyls and monoamines in various mouse brain regions at 4 h and 24 h after the last of four injections of METH (10 mg/kg/injection q 2 h). Substantial increases in TBARs and protein carbonyls were observed in the striatum and hippocampus but not the frontal cortex nor the cerebellum of METH-treated mice. Furthermore, lipid and protein oxidative markers were highly correlated within each brain region. In the hippocampus and striatum elevations in oxidative markers were significantly greater at 24 h than at 4 h. Monoamine levels were maximally reduced within 4 h (striatal dopamine [DA] by 95% and serotonin [5-HT] in striatum, cortex and hippocampus by 60-90%). These decrements persisted for 7 days after METH, indicating effects reflective of nerve terminal damage. Interestingly, NE was only transiently depleted in the brain regions investigated (hippocampus and cortex), suggesting a pharmacological and non-toxic action of METH on the noradrenergic nerve terminals. This study provides the first evidence for concurrent formation of lipid and protein markers of oxidative stress in several brain regions of mice that are severely affected by large neurotoxic doses of METH. Moreover, the differential time course for monoamine depletion and the elevations in oxidative markers indicate that the source of oxidative stress is not derived directly from DA or 5HT oxidation.     

An Assessment of Oxidative Damage to Proteins, Lipids, and DNA in Brain from Patients with Alzheimer's Disease

Abstract: Oxidative stress may contribute to neuronal loss in Alzheimer's disease (AD). The present study compares the levels of oxidative damage to proteins, lipids, and DNA bases from seven different brain areas of AD and matched control tissues by using a range of techniques. No differences in levels of lipid peroxidation were found in any of the brain regions by using two different assay systems. Overall, there was a trend for protein carbonyl levels to be increased in AD in frontal, occipital, parietal, and temporal lobe, middle temporal gyrus, and hippocampus, but a significant difference was found only in the parietal lobe. Gas chromatography-mass spectrometry was used to measure products of damage to all four DNA bases. Increased levels of some (8-hydroxyadenine, 8-hydroxyguanine, thymine glycol, Fapy-guanine, 5-hydroxyuracil, and Fapy-adenine), but not all, oxidized DNA bases were observed in parietal, temporal, occipital, and frontal lobe, superior temporal gyrus, and hippocampus. The baseline level of oxidative DNA damage in the temporal lobe was higher than in other brain regions in both control and AD brain. The finding of increased oxidative damage to protein and DNA strengthens the possibility that oxidative damage may play a role in the pathogenesis of AD in at least some key brain regions.     

Calpastatin levels affect calpain activation and calpain proteolytic activity in APP transgenic mouse model of Alzheimer's disease

The intracellular Ca(2+)-dependent protease calpain and the specific calpain endogenous inhibitor calpastatin are widely distributed, with the calpastatin/calpain ratio varying among tissues and species. Increased Ca(2+) and calpain activation have been implicated in Alzheimer's disease (AD), with scant data available on calpastatin/calpain ratio in AD. Information is lacking on calpain activation and calpastatin levels in transgenic mice that exhibit AD-like pathology. We studied calpain and calpastatin in Tg2576 mice and in their wild type littermates (control mice). We found that in control mice calpastatin level varies among brain regions; it is significantly higher in the cerebellum than in the hippocampus, frontal and temporal cortex, whereas calpain levels are similar in all these regions. In the Tg2576 mice, calpain is activated, calpastatin is diminished, and calpain-dependent proteolysis is observed in brain regions affected in AD and in transgenic mice (especially hippocampus). In contrast, no differences are observed between the Tg2576 and the control mice in the cerebellum, which does not exhibit AD-like pathology. The results are consistent with the notion that a high level of calpastatin in the cerebellum renders the calpain in this brain region less liable to be activated; in the other brain parts, in which calpastatin is low, calpain is more easily activated in the presence of increased Ca(2+), and in turn the activated calpain leads to further diminution in calpastatin (a known calpain substrate). The results indicate that calpastatin is an important factor in the regulation of calpain-induced protein degradation in the brains of the affected mice, and imply a role for calpastatin in attenuating AD pathology. Promoting calpastatin expression may be used to ameliorate some manifestations of AD.     

Covalently linking the Escherichia coli global anaerobic regulator FNR in tandem allows it to function as an oxygen stable dimer

Glutathione (GSH) serves as an important anti-oxidant in the brain by scavenging harmful reactive oxygen species that are generated during different molecular processes. The GSH level in the brain provides indirect information on oxidative stress of the brain. We report in vivo detection of GSH non-invasively from various brain regions (frontal cortex, parietal cortex, hippocampus and cerebellum) in bilateral hemispheres of healthy male and female subjects and from bi-lateral frontal cortices in patients with mild cognitive impairment (MCI) and Alzheimer's disease (AD). All AD patients who participated in this study were on medication with cholinesterase inhibitors. Healthy young male (age 26.4±3.0) and healthy young female (age 23.6±2.1) subjects have higher amount of GSH in the parietal cortical region and a specific GSH distribution pattern (parietal cortex>frontal cortex>hippocampus ~ cerebellum) has been found. Overall mean GSH content is higher in healthy young female compared to healthy young male subjects and GSH is distributed differently in two hemispheres among male and female subjects. In both young female and male subjects, statistically significant (p=0.02 for young female and p=0.001 for young male) difference in mean GSH content is found when compared between left frontal cortex (LFC) and right frontal cortex (RFC). In healthy young female subjects, we report statistically significant positive correlation of GSH content between RFC and LFC (r=0.641, p=0.004) as well as right parietal cortex (RPC) and left parietal cortex (LPC) (r=0.797, p=0.000) regions. In healthy young male subjects, statistically significant positive correlation of GSH content was observed between LFC and LPC (r=0.481, p=0.032) regions. This statistical analysis implicates that in case of a high GSH content in LPC of a young male, his LFC region would also contain high GSH and vice versa. The difference in mean of GSH content between healthy young female control and female AD patients in RFC region (p=0.003) and difference in mean of GSH content between healthy young male control and male AD patients (p=0.05) in LFC region is found to be statistically significant. It is the first scientific report correlating alteration (in selective brain regions) of GSH level with clinical status of male and female subjects using non-invasive imaging technique.     

Apolipoprotein-D expression is increased during development and maturation of the human prefrontal cortex

Apolipoprotein D (apoD) is a lipid binding protein expressed in the brain where its function is largely unknown. Based on changes in lipid metabolism and deposition that occur in the human brain during postnatal development, we investigated changes in apoD expression in the prefrontal cortex in 69 normal cases ranging in age from 40 days to 49 years utilizing gene microarray, quantitative PCR and western blotting methods. In contrast to the high expression of apolipoprotein E ( APOE ), low-density lipoprotein receptor-related protein 8 ( LRP8 ) and 3-hydroxy-3-methyl-glutaryl-CoA reducatase ( HMGCR ) (genes that play a role in lipid-related pathways in brain development) early in life, apoD expression was low in neonates and increased in expression throughout life resulting in six- to eight-fold higher levels at the mRNA and protein levels in adults. Recent studies suggest that apoD has a novel antioxidant function in the brain and we found that the increased apoD expression throughout development and into adulthood was correlated with the expression of antioxidant genes superoxide dismutase 1 ( SOD1 ) and glutathione peroxidase 3 ( GPX3 ) as well as proteins that were modified by the lipid peroxidation end-product 4-hydroxynonenal. These studies reveal that apoD expression is increased throughout life in the human prefrontal cortex and that this is correlated with genetic and biochemical markers of oxidative stress.