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Alterations in brain metal ion homeostasis have been reported with aging and are implicated in the pathogenesis of neurodegenerative diseases. To assess whether age-related changes in hypothalamic-pituitary-gonadal (HPG) hormones might be involved in modulating brain metal ion homeostasis, we treated 7.5-month intact, sham-ovariecomized and ovariectomized C57B6SJL mice with vehicle or leuprolide acetate (for 9-months) to differentiate between whether sex steroids or gonadotropins might modulate brain metal ion concentrations. Unlike other aging mammals, there was no increase in plasma luteinizing hormone (LH) and follicle-stimulating hormone (FSH) concentrations following estropause in mice, suggesting there was sufficient residual production by the follicle depleted ovary, of sex steroids like estrogens and protein hormones like the inhibins, in order to suppress pituitary LH/FSH production. Castration on the other hand induced significant increases in circulating LH and FSH. Modulation of plasma sex steroid and gonadotropin levels did not significantly alter the concentrations of brain metals tested (Fe, Zn, Cu, Mn, Co, Ni, Al, Li), although there was a tendency for a decrease in all brain metals following ovariectomy (low estrogens and progesterone, high gonadotropins), a response that was reversed with leuprolide acetate treatment (low sex steroids, low gonadotropins). Brain Cu concentration was the only metal correlated with plasma LH (−0.37, n = 30, p < 0.05) and FSH (−0.42, n = 29, p < 0.01). This study demonstrates that sex hormones do not markedly alter brain metal ion homeostasis, unlike previously reported studies of circulating metal ion homeostasis. The role of gonadotropins in regulating metal ion homeostasis does however warrant further study.

Proteomics has advanced and identified various plasma proteins that may be implicated in lesions in brains from neurodegenerative individuals. Apolipoprotein E and amyloid beta are amongst the many candidate proteins identi- fied in common neurodegenerative disorders with effects by diet and nutriproteomics on the interactions of apo E and amyloid beta. The purpose of this review paper is to specify the role of proteins and lipoproteins and their effects on he- patic amyloid beta clearance that are important to brain amyloidosis. In recent and historical research provided in various research papers the blood brain barrier (BBB) is abnormal in neurodegenerative disease and proteins and oxidized lipids leak across the BBB. The improved scientific understanding of apo E/amyloid beta interactions identifies modulations in protein structure and lipid:protein interactions induced by nutrients and has become important to peripheral amyloid beta metabolism. Hepatic acute phase proteins induced by diet, inflammation and oxidative stress are connected to the patho- physiology of neurotoxic diseases. Oxidative stress induced by high cholesterol diets can cause electrostatic alterations in amyloid oligomers accompanied with alterations in oxidized lipids and acute phase proteins. Recent progress in pro- teomics with relevance to Alzheimer's disease has led to the identification of acute phase proteins and include pentraxins that regulate or reduce amyloid beta sizes that are related to membrane permeability and disruption. In this review re- search papers identify acute phase biomarkers that are involved in the alterations of amyloid beta oligomers in obesity and diabetes with increased BBB permeability and central nervous system disorders.

The incidence of diabetes is predicted to increase to 21% by 2050. Currently, one third of US adults are obese and over 11% of these individuals have diabetes. Due to the growing need for therapeutic intervention to control and/or stabilize this increase in the incidence of diabetes in Western communities, gaining a comprehensive understanding of the association between obesity and Type 2 diabetes has become increasingly important to diabetes research. The increased cell senescence associated with diabetes has been associated with the limited ability of cells to divide, with indication of telomere shortening and genomic instability of the cells. Obese individuals have shorter telomeres suggesting an inverse relationship between adiposity and telomere length. The implication that Type 2 diabetes has on biological aging is of particular interest since telomere shortening in obesity and diabetes has been associated with an early risk for dementia and even progression to Alzheimer’s disease (AD). Lifestyle, nutrition and longevity are closely related and cellular senescence has been associated with telomere shortening and connected to longevity. Diet, cholesterol lowering drugs and exercise that control food intake and glucose tolerance in aging and diabetic individuals, via connections between liver circadian clocks and the suprachiasmatic nucleus in the brain, also have been shown to alter telomere lengths. Lifestyle interventions, such as diets low in fat and exercise, target the rise in obesity and associated telomere shortening by delaying or preventing the onset of Type 2 diabetes. The implementation of these anti-aging therapies early in life may prevent calorie overload and activation of calorie sensitive genes such as Sirtuin 1 (Sirt1). This may maintain telomere length and the control of obesity, which is linked to cardiovascular disease, diabetes and accelerates aging and AD.

Obesity, especially at mid-life, is a major risk factor for atherosclerosis, insulin resistance and the metabolic syndrome, which in turn contribute to coronary artery disease (CAD), Type 2 diabetes and Alzheimer’s disease (AD). The rise in overweight and obesity in all societies is prompting intense research into the causes and effects of the condition. Obesity disrupts many body systems including glucose and lipid metabolism, circadian rhythms and liver function. It also causes or increases inflammation and oxidative stress. Within cells, the endoplasmic reticulum (ER) appears to be particularly susceptible to such metabolic disruption. Sirtuin 1 (Sirt1) and leptin have received attention recently as they are central regulatory factors for the body’s metabolic pathways which interact at particular levels, for example lipid and Abeta metabolism. This mini-review discusses recent findings concerning obesity, lipid metabolism and the role of Sirtuin 1 and how all influence the ER. A greater understanding of obesity and its effects on metabolic control systems of the body are required, to develop pharmacological, dietary and lifestyle changes that will reduce the incidence of CAD, Type 2 diabetes and AD.

Brain sex steroids are derived from both peripheral (primarily gonadal) and local (neurosteroids) sources and are crucial for neurogenesis, neural differentiation and neural function. The mechanism(s) regulating the production of neurosteroids is not understood. To determine whether hypothalamic-pituitary-gonadal axis components previously detected in the extra-hypothalamic brain comprise a feedback loop to regulate neuro-sex steroid (NSS) production, we assessed dynamic changes in expression patterns of steroidogenic acute regulatory (StAR) protein, a key regulator of steroidogenesis, and key hypothalamic-pituitary-gonadal endocrine receptors, by modulating peripheral sex hormone levels in female mice. Ovariectomy (OVX; high serum gonadotropins, low serum sex steroids) had a differential effect on StAR protein levels in the extrahypothalamic brain; increasing the 30- and 32-kDa variants but decreasing the 37-kDa variant and is indicative of cholesterol transport into mitochondria for steroidogenesis. Treatment of OVX animals with E(2), P(4), or E(2) + P(4) for 3 days, which decreases OVX-induced increases in GnRH/gonadotropin production, reversed this pattern. Suppression of gonadotropin levels in OVX mice using the GnRH agonist leuprolide acetate inhibited the processing of the 37-kDa StAR protein into the 30-kDa StAR protein, confirming that the differential processing of brain StAR protein is regulated by gonadotropins. OVX dramatically suppressed extra-hypothalamic brain gonadotropin-releasing hormone 1 receptor expression, and was further suppressed in E(2)- or P(4)-treated OVX mice. Together, these data indicate the existence of endocrine and autocrine/paracrine feedback loops that regulate NSS synthesis. Further delineation of these feedback loops that regulate NSS production will aid in developing therapies to maintain brain sex steroid levels and cognition.

Studies in transgenic mice bearing mutated human Alzheimer disease (AD) genes show that active vaccination with the amyloid β (Aβ) protein or passive immunization with anti-Aβ antibodies has beneficial effects on the development of disease. Although a trial of Aβ vaccination in humans was halted because of autoimmune meningoencephalitis, favorable effects on Aβ deposition in the brain and on behavior were seen. Conflicting results have been observed concerning the relationship of circulating anti-Aβ antibodies and AD. Although these autoantibodies are thought to arise from exposure to Aβ, it is also possible that homologous proteins may induce antibody synthesis. We propose that the long-standing presence of anti-Aβ antibodies or antibodies to immunogens homologous to the Aβ protein may produce protective effects. The amino acid sequence of the potato virus Y (PVY) nuclear inclusion b protein is highly homologous to the immunogenic N-terminal region of Aβ. PVY infects potatoes and related crops worldwide. Here, we show through immunocytochemistry, enzyme-linked immunosorbent assay, and NMR studies that mice inoculated with PVY develop antibodies that bind to Aβ in both neuritic plaques and neurofibrillary tangles, whereas antibodies to material from uninfected potato leaf show only modest levels of background immunoreactivity. NMR data show that the anti-PVY antibody binds to Aβ within the Phe4–Ser8 and His13–Leu17 regions. Immune responses generated from dietary exposure to proteins homologous to Aβ may induce antibodies that could influence the normal physiological processing of the protein and the development or progression of AD.

Reproductive hormones have been demonstrated to modulate both gap and tight junction protein expression in the ovary and other reproductive tissues, however the effects of changes in reproductive hormones on the selective permeability of the blood-brain barrier (BBB) remain unclear. Age-related declines in BBB integrity correlate with the loss of serum sex steroids and increase in gonadotropins with menopause/andropause. To examine the effect of reproductive senescence on BBB permeability and gap and tight junction protein expression/localization, female mice at 3 months of age were either sham operated (normal serum E2 and gonadotropins), ovariectomized (low serum E2 and high serum gonadotropins) or ovariectomized and treated with the GnRH agonist leuprolide acetate (low serum E2 and gonadotropins). Ovariectomy induced a 2.2-fold increase in Evan's blue dye extravasation into the brain. The expression and localization of the cytoplasmic membrane-associated tight junction protein zona occludens 1 (ZO-1) in microvessels was not altered among groups indicating that the increased paracellular permeability was not due to changes in this tight junction protein. However, ovariectomy induced a redistribution of the gap junction protein connexin-43 (Cx43) such that immunoreactivity relocalized from along the extracellular microvascular endothelium to become associated with endothelial cells. An increase in Cx43 expression in the mouse brain following ovariectomy was suppressed in ovariectomized animals treated with leuprolide acetate, indicating that serum gonadotropins rather than sex steroids were modulating Cx43 expression. These results suggest that elevated serum gonadotropins following reproductive senescence may be one possible cause of the loss of selective permeability of the BBB at this time. Furthermore, these findings implicate Cx43 in mediating changes in BBB permeability, and serum gonadotropins in the cerebropathophysiology of age-related neurodegenerative diseases such as stroke and Alzheimer's disease.

Leuprolide acetate is a synthetic nonapeptide that is a potent gonadotropin-releasing hormone receptor (GnRHR) agonist used for diverse clinical applications, including the treatment of prostate cancer, endometriosis, uterine fibroids, central precocious puberty and in vitro fertilization techniques. As its basic mechanism of action, leuprolide acetate suppresses gonadotrope secretion of luteinizing hormone and follicle-stimulating hormone that subsequently suppresses gonadal sex steroid production. In addition, leuprolide acetate is presently being tested for the treatment of Alzheimer's disease, polycystic ovary syndrome, functional bowel disease, short stature, premenstrual syndrome and even as an alternative for contraception. Mounting evidence suggests that GnRH agonist suppression of serum gonadotropins may also be important in many of the clinical applications described above. Moreover, the presence of GnRHR in a multitude of non-reproductive tissues including the recent discovery of GnRHR expression in the hippocampi and cortex of the human brain indicates that GnRH analogs such as leuprolide acetate may also act directly via tissue GnRHRs to modulate (brain) function. Thus, the molecular mechanisms underlying the therapeutic effect of GnRH analogs in the treatment of these diseases may be more complex than originally thought. These observations also suggest that the potential uses of GnRH analogs in the modulation of GnRH signaling and treatment of disease has yet to be fully realized.

Gonadotropin-releasing hormone receptor I (GnRHR I) has been localized to the limbic system of the rat brain, although the functional consequences of GnRH signaling through these receptors is unknown. In this paper, we characterize the expression of GnRHR I in the human hippocampus and cortex, and the functionality of GnRHR I in human neuroblastoma cells. Robust GnRHR I immunoreactivity was detected in the cell body as well as along the apical dendrites of pyramidal neurons in the CA2, CA1, and end plate, but was clearly lower in the subiculum of the hippocampus. Immunolabeling was also evident in cortical neurons, including those located in the entorhinal cortex and occipitotemporal gyrus but was not observed within the granular layer of the dentate gyrus. No differences in immunohistochemical staining were observed between control and Alzheimer’s disease brain. GnRHR I mRNA and protein (mature, immature, and other variant) expression was detected in human neuroblastoma cells (M17, SH-SY5Y) and rat embryonic primary neurons and varied with differentiation and GnRH treatment. Since GnRHR I was expressed by extrapituitary cells, and hypothalamic GnRH I secretion markedly increases post-menopause/andropause, we treated human M17 neuroblastoma cells cultured in serum-free conditions with GnRH I for 6 h and measured LH expression. M17 neuroblastoma cells express LHβ mRNA, while immunoblot analysis indicated the presence of three LH variants (approximately 30, 47, and 60 kDa) that were upregulated by low concentrations of GnRH I, but down-regulated at higher GnRH I concentrations. LH expression was also found to increase in differentiating embryonic rat primary cortical neurons. Our results demonstrate that neurons expressing GnRHR I are functional, responding to GnRH I by upregulating LH production. Post-reproductive surges in GnRH I secretion may explain the accumulation of LH in pyramidal neurons of the aged human and rat.

Senescence is characterized neurologically by a decline in cognitive function, which we propose is the result of degenerative processes initiated by the dysregulation of the hypothalamic-pituitary-gonadal (HPG) axis with menopause and andropause. Compelling epidemiologic evidence to support this assertion includes the increased prevalence of Alzheimer disease (AD) in women, the correlation of serum HPG hormones with disease and the decreased incidence, and delay in the onset of AD following hormone replacement therapy. Dysregulation of the axis at this time leads to alterations in the concentrations of all serum HPG hormones (decreased neuronal sex steroid signaling, but increased neuronal gonadotropin releasing hormone, luteinizing hormone, and activin signaling). Hormones of the HPG axis, receptors for which are present in the adult brain, are important regulators of cell proliferation and differentiation during growth and development. Based on this, we propose that dysregulated HPG hormone signaling with menopause/andropause leads to the abortive reentry of differentiated neurons into the cell cycle via a process we term "dyosis." Interestingly, the major biochemical and neuropathologic changes reported for the AD brain also are intimately associated with neuron division: altered AbetaPP metabolism, Abeta deposition, tau phosphorylation, mitochondrial alterations, chromosomal replication, synapse loss, and death of differentiated neurons. Recent evidence supports the premise that AD-related biochemical changes are likely the combined result of increased mitotic signaling by gonadotropins and GnRH, decreased differentiative and neuroprotective signaling via sex steroids, and increased differentiative signaling via activins. This results in a hormonal milieu that is permissive of cell cycle reentry but does not allow completion of metaphase. Partial resetting of the axis following administration of normal endogenous sex steroids delays the onset and decreases the incidence of AD. Ideally, supplementation with HPG hormones should mimic closely the serum concentrations of all HPG hormones in reproductive men and cycling women to prevent dyotic signaling and attempted neuron division.

Introduction Alzheimer’s disease (AD) is a growing socioeconomic problem worldwide. Early diagnosis and prevention of this devastating disease have become a research priority. Consequently, the identification of clinically significant and sensitive blood biomarkers for its early detection is very important. Apolipoprotein E (APOE) is a well-known and established genetic risk factor for late-onset AD; however, the impact of the protein level on AD risk is unclear. We assessed the utility of plasma ApoE protein as a potential biomarker of AD in the large, well-characterised Australian Imaging, Biomarkers and Lifestyle Study of Ageing (AIBL) cohort. Methods Total plasma ApoE levels were measured at 18-month follow-up using a commercial bead-based enzyme-linked immunosorbent assay: the Luminex xMAP human apolipoprotein kit. ApoE levels were then analysed between clinical classifications (healthy controls, mild cognitive impairment (MCI) and AD) and correlated with the data available from the AIBL cohort, including but not limited to APOE genotype and cerebral amyloid burden. Results A significant decrease in ApoE levels was found in the AD group compared with the healthy controls. These results validate previously published ApoE protein levels at baseline obtained using different methodology. ApoE protein levels were also significantly affected, depending on APOE genotypes, with ε2/ε2 having the highest protein levels and ε4/ε4 having the lowest. Plasma ApoE levels were significantly negatively correlated with cerebral amyloid burden as measured by neuroimaging. Conclusions ApoE is decreased in individuals with AD compared with healthy controls at 18-month follow-up, and this trend is consistent with our results published at baseline. The influence of APOE genotype and sex on the protein levels are also explored. It is clear that ApoE is a strong player in the aetiology of this disease at both the protein and genetic levels.

proteins was quantitated using a l abel-free protein quantification method, and these were competed with the concentrations of A b42, pTau181, and tTau. Results: The MMSE score in AD patients was 21.3 6 4.9, and the average CDR scores were 0.86 0.3. All of the CSF samples were sufficient to establish the diagnostic cut-off value of the Ab42/pTau181 and Ab42/ tTau ratios in our laboratory.Fifteen of the 724 protein samples demonstrated a significant correlation with at least one of the AD biomarkers (p < 0.05). Seven of these proteins were previously identified as AD biomarkers, whereas eight were newly found. T he direction of change in the levels of Ab42 was found to be opposite to those of the levels of the Tau proteins in all analytes. Regarding the Ab42 levels, four proteins revealed a significant correlation. Another four proteins demonstrated a significant correlation with both pTau181 and tTau concentrations. We found three proteins to have a significant relationship only with pTau181, whereas two proteins showed a significant correlation only with tTau levels. Interestingly, t here were no analytes that demonstrated a correlation with both Ab42 and Tau proteins simultaneously. The remaining proteins showed a distinct correlation with the combined ratio of Ab42/ pTau181 or Ab42/tTau; however, the relationship with the individual concentrations was statistically negligible. Conclusions: Through our work, we identified new candidate CSF biomarkers of AD. Considering their biological roles, these candidate markers are suggested to be intimately related to AD pathophysiology. Further investigations are needed to identify their values as surrogate biomarkers or therapeutic targets in AD.

Transactive response DNA-binding protein of 43 kDa (TDP-43), an RNA and DNA binding protein involved in transcriptional repression, RNA splicing and RNA metabolism during the stress response, is the major component of neuronal inclusions in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin inclusions, now referred to as FTLD-TDP. While initially thought to be relatively specific to ALS and FTLD-TDP, TDP-43 pathology has now been detected in a number of other neurodegenerative diseases, many associated with tau pathology, including Guam Parkinson dementia complex and Alzheimer's disease (AD). TDP-43 pathology is detected in 25% to 50% of AD cases, especially those with more severe clinical phenotype and greater Alzheimer type pathology, as well as AD cases with hippocampal sclerosis (HS). HS is characterized by selective neuronal loss affecting CA1 sector of the hippocampus, and most cases of HS, with or without AD, have TDP-43 pathology. Whether TDP-43 pathology is merely an incidental finding in AD or actually contributing to the more severe clinical phenotype remains unresolved. Presence of TDP-43 in normal elderly, who are at increased risk for AD, would strengthen the argument that it is not merely a secondary or incidental finding in end stage AD. Limited studies suggest that TDP-43 pathology is infrequent in neurologically normal elderly (3% or less). We provide an overview of what is known about TDP-43 in AD, normal aging and in other disorders and suggest that TDP-43 proteinopathies be considered in two classes - primary and secondary.

Alterations in iron metabolism or oxidative damage in response to hypoxic incidents have been examined following re-oxygenation of the hypoxic tissue. To understand the consequences of decreased tissue oxygen on iron load, metal-catalyzed redox activity and oxidative modifications in isolation from re-oxygenation, the present study exposed mice to either normoxia, or mild hypoxia (380 Torr; approximately 10% normobaric oxygen) where the tissue was not allowed to re-oxygenate prior to examination. Brain, liver and skeletal muscle were examined for Fe3+ load, metal-catalyzed redox activity and oxidative modifications to proteins (N(epsilon)-(carboxymethyl)lysine), lipids (4-hydroxynonenal pyrrole) and nucleic acids (8-hydroxyguanosine). Hypoxia induced a 43% increase in the iron content of the liver (P < 0.001) as determined by ICP-MS and a 3.8-fold increase in Fe3+ load (P < 0.001) as determined by Perl's stain. There was a corresponding 2-fold increase in metal-catalyzed redox activity (P < 0.01) in the liver, but no change in the expression of oxidative markers. In contrast, non-significant increases in Fe3+ and metal-catalyzed redox activity were observed in the cerebral cortex, and molecular and granular layers of the hippocampus and cerebellum. Interestingly, hypoxia significantly decreased oxidative modifications to proteins and lipids, but not nucleic acids in most brain regions examined. In addition, hypoxia did not alter the Fe content of skeletal muscle, or the contents of Zn, Cu, Ni or Mn in liver, skeletal muscle, cerebral cortex or hippocampus. Together, these results indicate that there is a tighter regulation of iron metabolism in the brain than the liver, which limits the redistribution of Fe3+ following hypoxia.