Postmortem Human Brain Tissue Research Articles

Dynamic expression of Hsp27 in the presence of mutant ataxin-3

Machado–Joseph disease (MJD)/spinocerebellar ataxia type 3 (SCA3) is an autosomal dominant spinocerebellar degeneration characterized by a wide range of clinical manifestations. The molecular mechanisms underlying the selective neuronal death typical of MJD/SCA3 are unknown. In this study, human SK-N-SH neuroblastoma cells stably transfected with full-length MJD with 78 CAG repeats were assayed for the dynamic expression of Hsp27, known as a suppressor of poly-Q mediated cell death, in the presence of mutant ataxin-3 in different passages of cultured cells. A dramatic decrease of Hsp27 expression was observed in the earlier passage of cultured SK-N-SH-MJD78 cells, however, the later passage of cells showed a significant increase of Hsp27 to almost the same level of the parental cells. Furthermore, immunohistochemical analysis of MJD transgenic mice and post-mortem human brain tissues showed increased expression of Hsp27 compared to normal control brain, suggesting an up-regulation of Hsp27 in the end stage of MJD. However, mutant cells of earlier passages were more susceptible to serum deprivation than mutant cells of later passages, indicating weak tolerance toward stress in cells with reduced Hsp27. While heat shock was used to assess the stress response, cells expressing mutant ataxin-3 displayed normal response upon heat shock stimuli when compared to the parental cells. Taken together, we proposed that during the early disease stage, the reduction of Hsp27 synthesis mitigated the ability of neuron cells to cope with cytotoxicity induced by mutant ataxin-3, triggering the cell death process during the disease progress. In the late stage of disease, after prolonged stressful conditions of polyglutamine cytotoxicity, the increased level of Hsp27 may reflect a dynamic process of the survived cells to unfold and remove mutant ataxin-3. However, this increased Hsp27 still cannot reverse the global dysfunction of cellular proteins due to accumulation of cytotoxic effects.

Binding of the amyloid-imaging PET tracer, Pittsburgh Compound-B, to post-mortem human brain tissue from the AN-1792 immunotherapy trial

Binding of the amyloid-imaging PET tracer, Pittsburgh Compound-B, to post-mortem human brain tissue from the AN-1792 immunotherapy trial

Biphenyls labeled with technetium 99m for imaging β-amyloid plaques in the brain

Formation and accumulation of excess aggregates of β-amyloid (Aβ) plaques in the brain are critical factors contributing to the development and progression of Alzheimer's disease (AD). There is an urgent need for in vivo imaging agents that can specifically demonstrate the location and density of Aβ plaques in the brain. The aim of this study was to develop potential technetium 99m ( 99mTc)-labeled diagnostic imaging agents specific for the detection of Aβ plaques. Based on previously obtained Aβ plaque-specific biphenyls containing a p- N, N-dimethylaminophenyl group, a series of 99mTc and Re–N 2S 2–biphenyl derivatives was prepared. The stable neutral and lipophilic 99mTc complexes, [ 99mTc] 19 and [ 99mTc] 23, A+B, were successfully obtained. As surrogates for the 99mTc complexes, the corresponding surrogates, Re complexes of 23, were also prepared. Surprisingly, it was found that the Re complexes showed distinctively different retention profiles as compared with the corresponding 99mTc complexes. Biodistribution studies indicated that [ 99mTc] 23A readily passed through the blood–brain barrier (1.18% dose/brain at 2 min) in contrast to the low brain penetration of [ 99mTc] 19 (0.29% dose/brain at 2 min). Initial results suggested that [ 99mTc] 23A showed selective binding to the Aβ plaque-like structures in the brain sections from transgenic mice but not in the postmortem human brain tissue of patients with confirmed AD. The results provide encouraging evidence that development of a 99mTc-labeled agent for imaging Aβ plaques in the brain may be feasible. Caution should be taken when comparing these 99mTc complexes with the corresponding surrogates — the Re complexes.

Neurogenesis as a potential therapeutic strategy for neurodegenerative disorders

The discovery that new neurons could be observed in post-natal rodent brain was reported in the early 1960s [1–3], but only gained wider acceptance when a potential source of this “neurogenesis” was identified. Since the 1990s, progenitors for the main neural cell types had been isolated from rodent [45,49,58,59] and human adult or fetal brain tissue [11,63]. These isolated progenitors, or stem cells, are unique in their ability to self-renew and to differentiate, in a multipotential manner, into astrocytes, oligodendrocytes and neurons. In mouse and human stem cells, single growth factor treatment allows for continued proliferation or for differentiation toward a particular neural fate [28, 33,60,64]. At the same time that investigators were beginning to understand the multipotential nature of the isolated progenitors, the localization of in vivo neurogenesis to discrete areas of the brain were described. In rodents [4,32] and in non-human [23] and human primates [16], immunohistological examination determined that the areas of greatest concentration of new neurons in the adult brain are located in the subventricular zone, where they migrate to the olfactory bulb and in the subgranular zone of dentate gyrus. Results from the isolation of post-mortem human brain tissue, suggest that new neurons can be formed in the dentate gyrus well into adulthood [46]. It is important to note, when discussing neurogenesis, that three apparent stages exist, first the point of neural stem cell proliferation in the subgranular zone or subventricular zone, then the migration, on differentiation, to the granule cell layers, and finally the maturation with generation of functional neural networks for surviving neurons. Mitogen, growth factor and cytokine control of each neurogenesis step in isolated fetal neural progenitors [9,19,28] imply that each stage is finely regulated and deviations in for instance growth factor levels, could have physiological consequences. Accordingly, increases in proliferation of stem cells or neuronal progenitors promoted by the mitogen, basic fibroblast growth factor (bFGF), followed by differentiation to neurons, subsequently requires addition of brain-derived neurotrophic factor (BDNF) to maintain survival and arborization of the new neurons. It appears that only 50% of new neurons in the granule layer of dentate gyrus or new neurons in the olfactory bulb survive to maturity, normally a 2–4 week process. This observation could be important if neurogenesis is going to be exploited for therapeutic purposes.

Regional distribution and cellular localization of the ETS-domain transcription factor, FEV, mRNA in the human postmortem brain.

The ETS transcription factor Fifth Ewing Variant (FEV) mRNA, a homologue of the rodent Pet-1 gene that is exclusively expressed in serotonin-containing neurons and is a critical determinant of serotonin neuronal differentiation and development, was examined in human postmortem brain tissue using in situ hybridization histochemistry. Studies revealed that FEV mRNA is robustly and exclusively expressed in the major serotonin-containing cell groups of the dorsal and median raphe nuclei located in the midbrain and pons of the human brainstem. The localization of transcription factor, FEV, mRNA in serotonin-containing neurons of the human brain raises questions regarding the functional significance of this transcription factor in regulating serotonin-related genes and its potential role in psychiatric illness.

Functional polymorphisms in dopamine and serotonin pathway genes

There is mounting evidence on the functional significance of single nucleotide and simple repeat sequence polymorphisms in both the coding and regulatory regions of genes in the monoamine neurotransmitter pathways. Many of these gene variants have been associated with human behavioral disorders and traits, and thus have important clinical relevance. This review summarizes the literature on the published functional studies from a molecular, cellular, and neurobiological perspective, and notes their possible behavioral consequences. Functional studies have adopted a variety of strategies. Pharmacological studies have focused on the effects of gene variation at the protein level in terms of binding to ligands or drugs. Other key investigations have determined effects on gene expression at the level of transcription in mammalian cell cultures, lymphoblasts, and/or human postmortem brain tissue. This has enabled the comparison of in vitro and in vivo data, and furthermore provides an improved perceptive of their respective advantages. Additionally, molecular biological approaches have identified transcription factors (DNA-binding proteins) that interact with the motifs within the polymorphisms themselves. Various neuroimaging studies have further determined the relationship of genotype with protein availability in the brain, and thus have contributed to our understanding of the in vivo functional significance of gene variants. Finally, there is growing evidence from both human and animal studies on the interaction of functional polymorphisms with the environment in determining a behavioral outcome. Taken together, these findings have contributed to a greater understanding of the plausible molecular mechanisms that underpin the functional significance of polymorphisms in monoamine neurotransmitter pathway genes, and how they may influence behavioral phenotypes.

Neuroprotective Effects of Phenylbutyrate in the N171-82Q Transgenic Mouse Model of Huntington's Disease

Huntington's disease (HD) is caused by an expansion of exonic CAG triplet repeats in the gene encoding the huntingtin protein (Htt), however, the means by which neurodegeneration occurs remains obscure. There is evidence that mutant Htt interacts with transcription factors leading to reduced histone acetylation. We report that administration of the histone deacetylase inhibitor phenylbutyrate after onset of symptoms in a transgenic mouse model of HD significantly extends survival and attenuates both gross brain and neuronal atrophy. Administration of phenylbutyrate increased brain histone acetylation and decreased histone methylation levels as assessed by both immunocytochemistry and Western blots. Phenylbutyrate increased mRNA for components of the ubiquitin-proteosomal pathway and down-regulated caspases implicated in apoptotic cell death, and active caspase 3 immunoreactivity in the striatum. These results show that administration of phenylbutyrate, at doses that are well tolerated in man, exerts significant neuroprotective effects in a transgenic mouse model of HD, and therefore represents a very promising therapeutic approach for HD.

The Development of a Posttraumatic Stress Disorder Brain Collection

The Development of a Posttraumatic Stress Disorder Brain CollectionAmy Deep–Soboslay, Catherine E. Martin and Joel E. KleinmanAmy Deep–Soboslay1. Section on Neuropathology, Clinical Brain Disorders Branch, NIMH/NIH, in Bethesda, Maryland.Search for more papers by this author, Catherine E. Martin2. Section on Neuropathology, Clinical Brain Disorders Branch, NIMH/NIH, in Bethesda, Maryland.Search for more papers by this author and Joel E. Kleinman3. Section on Neuropathology, Clinical Brain Disorders Branch, NIMH/NIH, in Bethesda, Maryland.Search for more papers by this authorPublished Online:June 2005https://doi.org/10.1521/psyc.67.4.416.56568PDFPDF PLUS ShareShare onFacebookTwitterLinkedInRedditEmail ToolsAdd to favoritesDownload CitationsTrack Citations AboutReferencesAmerican Psychiatric Association. (1994). Diagnostic and statistical manual for mental disorders. 4th ed. Washington, DC: American Psychiatric Press. Google ScholarBallenger J.C., Davidson J.R.T., Lecrubier Y., Nutt D.J., Marshall R.D., Nemeroff C.B., Shalev A.Y., & Yehuda R. (2004). Consensus statement update on posttraumatic stress disorder from the international consensus group on depression and anxiety. Journal of Clinical Psychiatry, 65 (suppl. 1), 55–62. Medline, Google ScholarBrewin , C.R., Andrews, B., & Valentine, J.D. (2000). Meta–analysis of risk factors for posttraumatic stress disorder in trauma–exposed adults. Journal of Consulting and Clinical Psychology, 68(5), 748–766. Crossref Medline, Google ScholarCaspi , A., Sugden, K., Moffitt, T.E., Taylor, A., Craig, I.W., Harrington, H., McClay, J., Mill, J., Martin, J., Braithwaite, A., & Poulton, R. (2003). Influence of life stress on depression: Moderation by a polymorphism in the 5–HTT gene. Science, 301, 386–389. Crossref Medline, Google ScholarConnor , K.M., & Davidson, J.R. (1997). Familial risk factors in posttraumatic stress disorder. Annals of the New York Academy of Sciences, 21(821), 35–51. Crossref, Google ScholarKessler R.C. (2000). Posttraumatic stress disorder: The burden to the individual and to society. Journal of Clinical Psychiatry, 61(Suppl. 5), 4–12. Medline, Google ScholarLewis D.A. (2002). The human brain revisited: Opportunities and challenges in postmortem studies of psychiatric disorders. Neuropsychopharmacology, 26, 143–154. Crossref Medline, Google ScholarNutt D.J. & Malizia A.L. (2004). Structural and functional brain changes in posttraumatic stress disorder. Journal of Clinical Psychiatry, 65 (Suppl. 1), 11–17. Medline, Google ScholarRadant , A., Tsuang, D., Peskind, E.R., McFall, M., & Raskind, W. (2001). Biological markers and diagnostic accuracy in the genetics of posttraumatic stress disorder. Psychiatry Research, 102, 203–214. Crossref Medline, Google ScholarStein M.B., Jang, K.L., Taylor, S., Vernon, P.A., & Livesley, W.J. (2002). Genetic and environmental influences on trauma exposure and posttraumatic stress disorder symptoms: A twin study. American Journal of Psychiatry, 159(10), 1675–1681. Crossref Medline, Google ScholarTrue , W.R., Rice, J., Eisen, S.A., Heath, A.C., Goldberg, J., Lyons, M.J., & Nowak, J. (1993). A twin study of genetic and environmental contributions to liability for posttraumatic stress symptoms. Archives of General Psychiatry, 50, 257–264. Crossref Medline, Google Scholar Previous article Next article FiguresReferencesRelatedDetails Cited byCited by1. Matthew J. Friedman, M.D., Ph.D. and His Legacy of Leadership in the Field of Post-traumatic Stress DisorderOnline publication date: 19 May 2022. Go to citation Crossref Google Scholar2. Two Decades of Progress “Toward a National PTSD Brain Bank,” on the Occasion of Dr. Matthew J. Friedman’s RetirementOnline publication date: 19 May 2022. Go to citation Crossref Google Scholar3. Implementation and clinical characteristics of a posttraumatic stress disorder brain collectionOnline publication date: 13 June 2017. Go to citation Crossref Google Scholar4. VA’s National PTSD Brain Bank: a National Resource for ResearchOnline publication date: 25 August 2017. Go to citation Crossref Google Scholar Volume 67Issue 4Dec 2004 Information© Guilford Publications Inc.PDF download

P4-367 Recombinant neprilysin digestion of beta-amyloid in post-mortem human brain tissue models with analysis by mass spectroscopy

An increasing number of studies conducted under in vitro and in vivo conditions, have concluded that polyphenols, compounds frequently occurring in many herbs with antioxidant properties, prevent and reverse amyloid fibril formation. However, the mechanisms by which these natural products modulate the protein aggregation process are poorly understood. Herein, a range of techniques including thioflavin T (ThT) and ANS fluorescence assays, electron microscopy and circular dichroism have been employed to determine the efficacy of rosmarinic acid (RA) and resveratrol (Res) on the inhibition/reversion of fibrillogenesis and hindering cytotoxicity induced by protofibrils and amyloid fibrils of hen egg white lysozyme (HEWL). Results demonstrated that both polyphenols effectively inhibit fibrillogenesis and destabilize preformed fibrils of HEWL in a concentration-dependent manner. Cytotoxicity protection on PC12 cells was also observed using the MTT assay, ROS production assay, and phase-contrast microscopy. It is suggested that the mechanism underlying the inhibitory effects of RA and Res is to prevent hydrophobic interactions between HEWL amyloidogenic prefibrillar species, although additional studies is needed to elucidate the detailed mechanisms involved. A combination of antioxidative and anti-amyloidogenic properties of these molecules may provide them with the described neuroprotective capacities.

Single-cell gene expression analysis: implications for neurodegenerative and neuropsychiatric disorders.

Technical and experimental advances in microaspiration techniques, RNA amplification, quantitative real-time polymerase chain reaction (qPCR), and cDNA microarray analysis have led to an increase in the number of studies of single-cell gene expression. In particular, the central nervous system (CNS) is an ideal structure to apply single-cell gene expression paradigms. Unlike an organ that is composed of one principal cell type, the brain contains a constellation of neuronal and noneuronal populations of cells. A goal is to sample gene expression from similar cell types within a defined region without potential contamination by expression profiles of adjacent neuronal subpopulations and noneuronal cells. The unprecedented resolution afforded by single-cell RNA analysis in combination with cDNA microarrays and qPCR-based analyses allows for relative gene expression level comparisons across cell types under different experimental conditions and disease states. The ability to analyze single cells is an important distinction from global and regional assessments of mRNA expression and can be applied to optimally prepared tissues from animal models as well as postmortem human brain tissues. This focused review illustrates the potential power of single-cell gene expression studies within the CNS in relation to neurodegenerative and neuropsychiatric disorders such as Alzheimer's disease (AD) and schizophrenia, respectively.

Gene expression profiles derived from single cells in human postmortem brain

The study of postmortem human brain tissue remains the basis for the understanding of many CNS disorders and to verify data obtained in experimental studies. So far, however, gene expression profiling in cellular sub-populations derived from human postmortem brain was hampered by several technical drawbacks. Here, we describe a method that allows the generation of mRNA expression profiles from single neurons. Dopaminergic neurons from different midbrain areas including substantia nigra, central gray substance and ventral tegmental area were identified and isolated by immuno-laser capture microscopy (LCM). Expression profiles were generated from microdissected cells using a modified RNA fingerprinting protocol. Using this approach, we were able to generate specific RNA fingerprints at a high resolution from phenotype-specific single neurons. Polymorphic fragments were isolated from gels and differential gene expression was confirmed by real-time PCR using gene-specific primer pairs and hybridization probes. The method described here is easy to use and reliable for profiling gene expression at the single cell level in human postmortem brain. It could therefore be valuable to open new insights into the molecular pathogenesis of CNS disorders.

Specific caspase interactions and amplification are involved in selective neuronal vulnerability in Huntington's disease.

Huntington's disease (HD) is an autosomal dominant progressive neurodegenerative disorder resulting in selective neuronal loss and dysfunction in the striatum and cortex. The molecular pathways leading to the selectivity of neuronal cell death in HD are poorly understood. Proteolytic processing of full-length mutant huntingtin (Htt) and subsequent events may play an important role in the selective neuronal cell death found in this disease. Despite the identification of Htt as a substrate for caspases, it is not known which caspase(s) cleaves Htt in vivo or whether regional expression of caspases contribute to selective neuronal cells loss. Here, we evaluate whether specific caspases are involved in cell death induced by mutant Htt and if this correlates with our recent finding that Htt is cleaved in vivo at the caspase consensus site 552. We find that caspase-2 cleaves Htt selectively at amino acid 552. Further, Htt recruits caspase-2 into an apoptosome-like complex. Binding of caspase-2 to Htt is polyglutamine repeat-length dependent, and therefore may serve as a critical initiation step in HD cell death. This hypothesis is supported by the requirement of caspase-2 for the death of mouse primary striatal cells derived from HD transgenic mice expressing full-length Htt (YAC72). Expression of catalytically inactive (dominant-negative) forms of caspase-2, caspase-7, and to some extent caspase-6, reduced the cell death of YAC72 primary striatal cells, while the catalytically inactive forms of caspase-3, -8, and -9 did not. Histological analysis of post-mortem human brain tissue and YAC72 mice revealed activation of caspases and enhanced caspase-2 immunoreactivity in medium spiny neurons of the striatum and the cortical projection neurons when compared to controls. Further, upregulation of caspase-2 correlates directly with decreased levels of brain-derived neurotrophic factor in the cortex and striatum of 3-month YAC72 transgenic mice and therefore suggests that these changes are early events in HD pathogenesis. These data support the involvement of caspase-2 in the selective neuronal cell death associated with HD in the striatum and cortex.

Amplification of RNA transcripts using terminal continuation

A new methodology has been developed to amplify RNA from minute amounts of starting material. Specifically, an efficient means of second-strand (ss) cDNA synthesis using a sequence-specific ‘terminal continuation' (TC) method is demonstrated. An RNA synthesis promoter is attached to the 3′ and/or 5′ region of cDNA utilizing the TC mechanism. The orientation of amplified RNAs is ‘antisense' or a novel ‘sense' orientation. TC RNA amplification is utilized for many downstream applications including gene expression profiling, cDNA microarray analysis, and cDNA library/subtraction library construction. Synthesized sense TC-amplified RNA can also be used as a template for in vitro protein translations and downstream proteomic applications. The TC RNA amplification methodology offers high sensitivity, flexibility, and throughput capabilities. A likely mechanism is that the TC primer binds preferentially to GC-rich CpG islands flanking 5′ regions of DNA that contain promoter sequences. Following TC RNA amplification, a large proportion of genes can be assessed quantitatively as evidenced by bioanalysis and cDNA microarray analysis in mouse and human postmortem brain tissues.

Dephosphorylation of microtubule-associated protein tau by protein phosphatase 5.

Protein phosphatase 5 (PP5) is a 58-kDa novel phosphoseryl/phosphothreonyl protein phosphatase. It is ubiquitously expressed in all mammalian tissues examined, with a high level in the brain, but little is known about its physiological substrates. We found that this phosphatase dephosphorylated recombinant tau phosphorylated with cAMP-dependent protein kinase and glycogen synthase kinase-3beta, as well as abnormally hyperphosphorylated tau isolated from brains of patients with Alzheimer's disease. The specific activity of PP5 toward tau was comparable to those reported with other protein substrates examined to date. The PP5 activity toward tau was stimulated by arachidonic acid by 30- to 45-fold. Immunostaining demonstrated that PP5 was primarily cytoplasmic in PC12 cells and in neurons of postmortem human brain tissue. A small pool of PP5 associated with microtubules. Expression of active PP5 in PC12 cells resulted in reduced phosphorylation of tau, suggesting that PP5 can also dephosphorylate tau in cells. These results suggest that PP5 plays a role in the dephosphorylation of tau and might be involved in the molecular pathogenesis of Alzheimer's disease.

Relevance of imidazoline receptors and agmatine to psychiatry: a decade of progress.

The cardiovascular relevance of imidazoline receptors (IR) has received tremendous attention since their discovery in 1984. However, evidence also has accumulated for the relevance of IR and an endogenous ligand, agmatine, to psychiatric disease. Emphasis has been placed on altered levels of the I(1)-imidazoline site on human platelets and in human postmortem brain tissue from depressed patients. Attempts at exploring the molecular nature of the I(1) protein have led to the cloning of a protein, IRAS. Based on transfection studies, IRAS seems to be involved in neuronal plasticity events. The I(2) site also appears linked to psychiatric research since some of these sites are localized to a specific domain on monoamine oxidases. Different peptides have been identified by means of an imidazoline-receptor-binding-protein (IRBP) antiserum, and these peptides, some of which appear to be fragments derived from IRAS, undergo changes in platelets and brain commensurate with altered mood states of the subject, notably depressive symptomatology. The search for an endogenous ligand for imidazoline receptor(s) also has led to agmatine, a decarboxylated derivative of arginine. Research on agmatine has mushroomed over the past several years and its measurement in the blood and brain has opened new research opportunities. This novel neurotransmitter interacts with a variety of receptors and has been implicated in mediation of stress responses, analgesia, drug addiction and withdrawal, convulsions, and neuroprotection. Given that IR and agmatine appear involved in a multitude of neurophysiologic and pathologic functions, the potential for new drug development is intriguing.

Isolation and characterization of neural progenitor cells from post-mortem human cortex.

Post-mortem human brain tissue represents a vast potential source of neural progenitor cells for use in basic research as well as therapeutic applications. Here we describe five human neural progenitor cell cultures derived from cortical tissue harvested from premature infants. Time-lapse videomicrography of the passaged cultures revealed them to be highly dynamic, with high motility and extensive, evanescent intercellular contacts. Karyotyping revealed normal chromosomal complements. Prior to differentiation, most of the cells were nestin, Sox2, vimentin, and/or GFAP positive, and a subpopulation was doublecortin positive. Multilineage potential of these cells was demonstrated after differentiation, with some subpopulations of cells expressing the neuronal markers beta-tubulin, MAP2ab, NeuN, FMRP, and Tau and others expressing the oligodendroglial marker O1. Still other cells expressed the classic glial marker glial fibrillary acidic protein (GFAP). RT-PCR confirmed nestin, SOX2, GFAP, and doublecortin expression and also showed epidermal growth factor receptor and nucleostemin expression during the expansion phase. Flow cytometry showed high levels of the neural stem cell markers CD133, CD44, CD81, CD184, CD90, and CD29. CD133 markedly decreased in high-passage, lineage-restricted cultures. Electrophysiological analysis after differentiation demonstrated that the majority of cells with neuronal morphology expressed voltage-gated sodium and potassium currents. These data suggest that post-mortem human brain tissue is an important source of neural progenitor cells that will be useful for analysis of neural differentiation and for transplantation studies.

Expression of BRI, the normal precursor of the amyloid protein of familial British dementia, in human brain.

Familial British dementia (FBD) is characterized neuropathologically by deposition of a unique amyloid-forming protein, ABri. It is a fragment of an abnormal form of a precursor protein, BRI. In FBD, BRI is elongated by 11 amino acids due to a point mutation that prevents recognition of the normal stop codon. We have investigated the expression of normal BRI in non-FBD cases. Three antibodies were raised against sequences of BRI and were used for immunoblotting and immunohistochemistry. Each of these antibodies detected a band at approximately 35 kDa by Western blotting. In postmortem human brain tissues, BRI was detected as fine granules in the neuronal cytoplasm. Pyramidal neurons in CA3 and CA4 of the hippocampus as well as Purkinje cells in the cerebellar cortex were most intensely stained for BRI. Such a distribution of neurons strongly expressing BRI parallels the reported occurrence of ABri deposits in patients with FBD. In pathological cases, BRI was detected in dystrophic neurites in senile plaques, around lesions in ischemic cases, in torpedo and glumose changes in the cerebellum, Lewy neurites, ballooned neurons, and neurons generally in hypoxic cases. These results suggest that BRI is transported in neuronal processes and is possibly involved in some role in nerve terminals. While a physiological role of BRI in brain remains to be determined, the behavior of BRI in diverse brain lesions appears to be somewhat analogous to that of amyloid precursor protein, which is the source of the beta-amyloid protein of Alzheimer's disease.

Methods for the identification of differentially expressed genes in human post-mortem brain

Microarrays can be used to monitor the expression of thousands of genes simultaneously. This technique requires high-quality RNA which can be extracted from a variety of tissues and cells including post-mortem human brain. Given the vast amount of information obtained from microarray studies, it is critical to establish valid analysis techniques to identify differentially expressed genes. This technical report describes the basic methodology and analyses used to identify such genes in human post-mortem brain tissue.

Mitochondria and vascular lesions as a central target for the development of Alzheimer's disease and Alzheimer disease-like pathology in transgenic mice

Accumulating evidence strongly suggests that the AD brain is characterized by impairments in energy metabolism, and vascular hypoperfusion, whereby oxidative stress appears to be an especially important contributor to neuronal death and development of AD pathology. We hypothesized that mitochondria play a key role in the generation of reactive oxygen species, resulting in oxidative damage to neuronal cell bodies, as well as other cellular compartments in the AD brain. All of these changes have been found to accompany AD pathology. In this review we have outlined recent evidence from the literature and our own original studies concerning the role of mitochondrial abnormalities and vascular damage in the pathogenesis of AD and AD-like pathology in transgenic mice (as a model for human AD). We examined ultrastructural features of vascular lesions and mitochondria from vascular wall cells in human AD brain biopsies, in human short post-mortem brain tissues and in yeast artificial chromosome (YAC) and C57B6/ SJL transgenic positive (Tg+) mice overexpressing amyloid beta precursor protein (AßPP). In situ hybridization using mitochondrial DNA (mtDNA) probes for human wild type, 5kb deleted and mouse mtDNA was performed along with immunocytochemistry using antibodies against amyloid beta precursor protein (A β PP), 8-hydroxy-2′-guanosine (8OHG) and cytochrome C oxidase (COX) were studied at the electron microscopic levels. There was a higher degree of amyloid deposition in the vascular walls of the human AD, YAC and C57B6/SJL Tg (+) mice compared to aged-matched controls. In addition, vessels with more severe lesions showed immunopositive staining for APP and possessed large, lipid-laden vacuoles in the cytoplasm of endothelial cells (EC). Significantly more mitochondrial abnormalities were seen in human AD, YAC and C57B6/SJL Tg (+) mouse microvessels where lesions occurred. In situ hybridization using wild and chimera (5 kB) mtDNA probes revealed positive signals in damaged mitochondria from the vascular endothelium and in perivascular cells of lesioned microvessels close to regions of large amyloid deposition. These features were absent in undamaged regions of human AD tissues, YAC and C57B6/SJL Tg (+) mouse tissues and in aged-matched control subjects. In addition, vessels with atherosclerotic lesions revealed endothelium and perivascular cells possessing clusters of wild and deleted mtDNA positive probes. These mtDNA deletions were accompanied by increased amounts of immunoreactive APP, 8OHG and COX in the same cellular compartment. Our observations first time demonstrate that vascular wall cells, especially their mitochondria, appear to be a central target for oxidative stress induced damage.

Reactivation of atrophic neurons in Alzheimer's disease

Decreased metabolic rate may precede cognitive impairment in Alzheimer's disease (AD) and is thus an early occurring hallmark. Several observations in post-mortem brain indicate that activated neurons are better able to withstand aging and AD, a phenomenon paraphrased by us as 'use it or lose it'. Moreover, a number of pharmacological and nonpharmacological studies support the concept that activation of the brain has beneficial effects and may to a certain degree restore several aspects of cognition and other central functions. For instance, the circadian system may be restimulated in Alzheimer patients by exposing them to more light or transcutaneous nerve stimulation. A procedure has been developed to culture human post-mortem brain tissue that allows testing of the efficacy of putative stimulatory compounds such as neurotrophins.