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Biomedical subjects

Clive Harper

Publications and source records attributed to Clive Harper.

12 recordsLinked to original sources
PubMed2006

Validation of endogenous controls for quantitative gene expression analysis: application on brain cortices of human chronic alcoholics.

Real-time PCR is frequently used for gene expression quantification due to its methodological sensitivity and reproducibility. The gene expression is quantified by normalization to one or more reference genes, usually beta-actin (ACTB), glyceraldehyde-3-phosphate dehydrogenase (GAPD) or to ribosomal RNA (18S). However, different environmental or pathological conditions might also influence the expression of normalizing genes, which could severely skew the interpretation of quantitative results. This study evaluates whether 16 genes frequently used as endogenous controls in expression studies, can serve as such for comparison of human brain tissues of chronic alcoholics and control subjects. The prefrontal and motor cortices that are affected differently by chronic alcohol consumption were analyzed. The reference genes that have no or small differences in expression in alcoholics and control subjects, were found to be specific for each region: beta-actin (ACTB) and ribosomal large P0 (RPLP0) for the prefrontal cortex while importin 8 (IPO8) and RNA polymerase II (POLR2A) for the motor cortex. Four out of sixteen analyzed genes demonstrated significant differences in expression between alcoholics and controls: phosphoglycerate kinase (PGK1), hypoxanthine phosphoribosyl transferase (HPRT1) and peptidylprolyl isomerase A (PPIA) in the motor cortex and beta-2-microglobulin (B2M) in the prefrontal cortex. Our study demonstrates the importance of validation of endogenous control genes prior to real-time PCR analysis of human brain tissues. Prescribed and non-prescribed drugs, pathological or environmental conditions along with alcohol abuse may differentially influence expression of reference genes.

Adult↗
PubMed2006

Differential protein expression in the corpus callosum (splenium) of human alcoholics: a proteomics study.

It is widely accepted that the chronic use of alcohol induces metabolic abnormalities and neuronal damage in the brain, which can lead to cognitive dysfunction. Neuroimaging studies reveal that alcohol-induced brain damage is region specific and prominent damage has been observed in both gray and white matter of the prefrontal cortex, and a wide range of white matter structures including the corpus callosum. Molecular mechanisms underlying these structural changes are largely unknown. Using proteomics we have analysed the changes in protein expression in the splenium of the corpus callosum in two different alcoholic groups. Protein extracts from splenium of 22 human brains (nine controls, seven uncomplicated alcoholics and six complicated alcoholics with hepatic cirrhosis-designated complicated) were separated using two-dimensional gel electrophorosis. Image analysis revealed that there were significant alterations in protein expression for 25 protein spots in the uncomplicated alcoholic group and 45 in the complicated group compared to control (P<0.05; ANOVA). In a total of 72 spots (identified as 36 proteins), 15 (identified as 14 proteins) spots overlapped between two alcoholic groups. Another 32 protein spots (26 different proteins) were identified only in the complicated alcoholics. It is therefore possible that these 26 proteins in the complicated group are likely to be the results of hepatic compromise. When compared with our previous data of white matter from the prefrontal cortex in alcoholics, large numbers of identified proteins in the splenium are different. This suggests that there may be different mechanisms causing alcohol-induced brain damage in different regions of the white matter. Our data also indicate the importance of other pathways including oxidative stress, lipid peroxidation and apoptosis as potential causes of alcohol-induced brain damage.

Aged↗
PubMed2006

Stable neuropsychological deficits in adult polyglucosan body disease.

We describe a 61-year-old woman who gradually developed deficits of balance, gait, and the ability to negotiate movement in space, together with an unusual pattern of cognitive deficits. A series of non-invasive investigations over three years including EEG, CT, MRI, PET and serial neuropsychological review had not provided a diagnosis. Significantly, the four neuropsychological assessments had revealed no progressive decline in cognition. Brain biopsy revealed an abundance of corpora amylacea, and a diagnosis of adult polyglucosan body disease (APBD) was made. This case contributes to the body of knowledge about the cognitive manifestations of this rare disease, and the stability of its functional impact over time.

Brain Diseases, Metabolic↗
PubMed2005

Ethanol and brain damage.

It is now well established that even uncomplicated alcoholics who have no specific neurological or hepatic problems show signs of regional brain damage and cognitive dysfunction. Improvements in neuroimaging technology, magnetic resonance imaging, magnetic resonance spectroscopy and positron emission tomography have contributed significantly, revealing alcoholic-specific changes in the CNS associated with neuropsychological abnormalities. Although greater efforts are needed, a human brain bank specifically targeting alcohol cases is now able to provide fresh and frozen tissue for alcohol researchers. These tissues can be used to test hypotheses developed using animal models and/or in vitro studies. The aim is to delineate mechanisms underlying alcohol-related brain damage in humans. The development of high-throughput, non-hypothesis-driven approaches using DNA microarrays and proteomics might also provide clues to this important problem.

Aged↗
PubMed2005

Alcoholic neurobiology: changes in dependence and recovery.

This article presents the proceedings of a symposium held at the meeting of the International Society for Biomedical Research on Alcoholism (ISBRA) in Mannheim, Germany, in October, 2004. Chronic alcoholism follows a fluctuating course, which provides a naturalistic experiment in vulnerability, resilience, and recovery of human neural systems in response to presence, absence, and history of the neurotoxic effects of alcoholism. Alcohol dependence is a progressive chronic disease that is associated with changes in neuroanatomy, neurophysiology, neural gene expression, psychology, and behavior. Specifically, alcohol dependence is characterized by a neuropsychological profile of mild to moderate impairment in executive functions, visuospatial abilities, and postural stability, together with relative sparing of declarative memory, language skills, and primary motor and perceptual abilities. Recovery from alcoholism is associated with a partial reversal of CNS deficits that occur in alcoholism. The reversal of deficits during recovery from alcoholism indicates that brain structure is capable of repair and restructuring in response to insult in adulthood. Indirect support of this repair model derives from studies of selective neuropsychological processes, structural and functional neuroimaging studies, and preclinical studies on degeneration and regeneration during the development of alcohol dependence and recovery form dependence. Genetics and brain regional specificity contribute to unique changes in neuropsychology and neuroanatomy in alcoholism and recovery. This symposium includes state-of-the-art presentations on changes that occur during active alcoholism as well as those that may occur during recovery-abstinence from alcohol dependence. Included are human neuroimaging and neuropsychological assessments, changes in human brain gene expression, allelic combinations of genes associated with alcohol dependence and preclinical studies investigating mechanisms of alcohol induced neurotoxicity, and neuroprogenetor cell expansion during recovery from alcohol dependence.

Adult↗
PubMed2004

Postmortem MR imaging of formalin-fixed human brain.

High-resolution postmortem neuroimaging of the brain can play a role in research programs by providing archival and reslicable images of brain specimens before permanent sectioning. These images can supplement evidence attained from both traditional neuropathological observations and in vivo neuroimaging. Differential brain tissue conspicuity, detectable with MRI, is determined by the density and mobility of water protons. Water content is about 70% in white matter, 80% in gray matter, and 99% in cerebrospinal fluid (CSF). To the extent that brain tissue contrast is determined by the number and microenvironment of water protons, timing parameters of MR image acquisition can interrogate this environment. Because the chemical environment of protons is different in living from dead tissue, optimal temporal imaging parameters, for example, for spin-echo imaging, commonly used for in vivo clinical and research study are different from those best for postmortem imaging. Here, we present a series of observations to identify relaxation times and optimal parameters for high-resolution structural imaging of formalin-fixed postmortem brain tissue using commercially available clinical scanners and protocols. Examples of high-resolution images and results from attempts at diffusion imaging are presented.

Aged↗
PubMed2003

Increased levels of apolipoprotein E in the frontal cortex of subjects with schizophrenia.

BACKGROUND: It is unclear whether altered expression of a specific isoform of apolipoprotein E (apoE) is associated with the pathology of schizophrenia. METHODS: To address whether apoE may be involved in the pathology of schizophrenia, we measured the genotypic and allelic frequency of polymorphisms in its gene and transcriptional regulatory region in DNA from Brodmann's area (BA) 9 obtained postmortem from schizophrenic and control subjects as well as its levels in the same tissue using Western blot analysis. RESULTS: The genotypic or allelic frequencies of any polymorphism studied did not vary between diagnostic cohorts. There was a significant increase in the levels of apoE protein in BA 9 from the schizophrenic subjects (Mean +/- SEM: 270 +/- 8.3 vs. 238 +/- 7.1 ng apoE/mg protein, p =.008) and a decrease in tissue from an analogous cortical region from rats treated with haloperidol compared with vehicle-treated animals (50 +/- 6.4 vs. 116 +/- 9.2 ng apoE/mg protein; p =.0002). CONCLUSIONS: These data support the hypothesis that increased levels of apoE may be associated with the pathology of schizophrenia and that antipsychotic drugs decrease apoE levels as part of their therapeutic actions.

Adult↗
PubMed2003

Apolipoprotein D levels are elevated in prefrontal cortex of subjects with Alzheimer's disease: no relation to apolipoprotein E expression or genotype.

BACKGROUND: Apolipoprotein E (apoE) has been implicated in the pathology of AD ever since inheritance of the epsilon4 allele was shown to be an important risk factor for the development of AD. Apolipoprotein D (apoD) is elevated in association with several central nervous system disorders, including Alzheimer's disease (AD), and has been proposed to be an especially robust marker for brain regions specifically affected by particular neuropathologies. Progressive cognitive decline is the core clinical feature of AD and is associated with disturbances in the prefrontal cortex. METHODS: We measured apoD levels in prefrontal cortex samples obtained postmortem from 20 autopsy-confirmed AD subjects and 40 control subjects. RESULTS: Enzyme-linked immunosorbent assay analysis revealed a significant increase in apoD expression in AD subjects compared with control subjects (.218+/-.029 microg/mg protein vs.117+/-.011 microg/mg protein; p=0003). There was no significant difference in apoD expression between early-onset and late-onset Alzheimer's subjects. Apolipoprotein D expression levels were not correlated with apoE levels, nor were they correlated with inheritance of the APOE epsilon4 allele. CONCLUSIONS: These findings suggest that apoD may be related to the cognitive decline observed in AD patients and that apoD and apoE likely play different roles in the pathogenesis of AD.

Aged↗
PubMed2003

Neuropathological alterations in alcoholic brains. Studies arising from the New South Wales Tissue Resource Centre.

Alcohol dependence and abuse are among the most costly health problems in the world from both social and economic points of view. Patterns of drinking appear to be changing throughout the world with more women and young people drinking heavily. Excessive drinking can lead to impairment of cognitive function and structural brain changes--some permanent, some reversible. Patterns of damage appear to relate to lifetime alcohol consumption but, more importantly, to associated medical complications. The most significant of these is the alcohol-related vitamin deficient state, the Wernicke-Korsakoff syndrome (WKS), which is caused by thiamin deficiency but is seen most commonly in alcoholics. Careful selection and classification of alcoholic cases into those with and without these complications, together with detailed quantitative neuropathological analyses has provided data that gives clues to the most vulnerable regions and cells in the brain. Brain shrinkage is largely accounted for by loss of white matter. Some of this damage appears to be reversible. Alcohol-related neuronal loss has been documented in specific regions of the cerebral cortex (superior frontal association cortex), hypothalamus and cerebellum. No change is found in basal ganglia, nucleus basalis, or serotonergic raphe nuclei. Many of these regions which are normal in uncomplicated alcoholics are damaged in those with the WKS. Dendritic and synaptic changes have been documented in alcoholics and these, together with receptor and transmitter changes, may explain functional changes and cognitive deficits, which precede more severe structural neuronal changes. A resource to provide human brain tissues for these types of studies has been developed at the University of Sydney--the New South Wales Tissue Resource Centre. The aim of this facility is to provide research groups throughout the world with fresh and/or frozen tissues from well-characterized cases of alcohol-related brain damage and matched controls. The development of new technologies in pathology and molecular biology means that many more questions can be addressed using appropriately stored human brain tissues. Examples of the application of some of these techniques, involving neurochemical, neuropharmacological, neuroimaging and gene expression studies are included in this paper. Important public health outcomes have arisen from some of these studies including the enrichment of bread flour with thiamin for the whole of Australia. Researchers with an interest in alcohol studies can access tissues from this brain bank.

Alcoholism↗
PubMed2003

How important are brain banks for alcohol research?

This article contains the proceedings of a symposium at the 2002 RSA/ISBRA Meeting in San Francisco, organized and chaired by Clive Harper and co-chaired by Izuru Matsumoto. The presentations were (1) Introduction, by Clive Harper; (2) The quality of tissue-a critical issue, by Therese Garrick; (3) The first systematic brain tissue donor program in Japan, by Izuru Matsumoto; (4) Brain scans after death-really! by Adolf Pfefferbaum, Elfar Adalsteinsson, and Edith Sullivan; (5) Capture that (genial) expression, by Joanne Lewohl and Peter Dodd; and (6) Neurochemical/pharmacological studies: experimental design and limitations, by Roger Butterworth.

Alcohol-Induced Disorders, Nervous System↗
PubMedDate not supplied

The expression and significance of metallothioneins in murine organs and tissues following mercury vapour exposure.

The fate of inspired mercury vapour (Hg0) is critical in the central nervous system (CNS) where it can circumvent the blood-brain barrier (BBB) at the neuromuscular junction (NMJ) and accumulate indefinitely in motor neurons by retrograde transport. The detoxification of systemic Hg0 by lung and liver requires investigation. We exposed 129/Sv wild-type (Wt) and 129/Sv MT-I, II double knockout (KO) mice to 500 microg Hg0/m3 for 4 hours to investigate the expression of MT in the lung, liver, and spinal cord following Hg0 exposure using unexposed groups as controls. There were congestive changes in liver and lung of both Wt and MT-KO groups of Hg0-treated mice; these changes appeared more pronounced in the MT-KO group. Motor neurons in the spinal cord did not show any pathological changes. Based on expression of MT, liver appears to have a major role in trapping and stabilising mercury. In the spinal cord, MT was expressed in all white matter astrocytes and in some grey matter astrocytes. Notably, motor neurons did not express MT, and the presence of MT could not be demonstrated in the axons of the ventral root. The absence of MT expression in motor neurons and their axons suggests the dependence of the motor system on the detoxifying capacity of liver MTs.

Administration, Inhalation↗