Prita R. Asih

The Effects of Testosterone Supplementation on Cognitive Functioning in Older Men.

Reduction in testosterone levels in men during aging is associated with cognitive decline and risk of dementia. Animal studies have shown benefits for testosterone supplementation in improving cognition and reducing Alzheimer’s disease pathology. In a randomized, placebo-controlled, crossover study of men with subjective memory complaint and low testosterone levels, we investigated whether testosterone treatment significantly improved performance on various measures of cognitive functioning. Forty-four men were administered a battery of neuropsychological tests to establish the baseline prior to being randomly divided into two groups. The first group (Group A) received 24 weeks of testosterone treatment (T treatment) followed by 4 weeks washout, and then 24 weeks of placebo (P); the second group (Group B) received the same treatments, in reverse order (Placebo, washout, and then T treatment). In group A (TèP), compared to baseline, there was a modest (1 point) but significant improvement in general cognitive functioning as measured by the Mini Mental State Examination (MMSE) following testosterone treatment. This improvement from baseline was sustained following the washout period and crossover to placebo treatment. Similar Mini Mental State Examination (MMSE) scores were observed when comparing testosterone treatment with placebo. In group B (PèT) a significant increase was observed from baseline following testosterone treatment and a trend towards an increase when compared to placebo treatment. Improvements in baseline depression scores (assessed by Geriatric Depression Scale) were observed following testosterone/placebo treatment in both groups, and no difference was observed when comparing testosterone with placebo treatment. Our findings indicate a modest improvement on global cognition with testosterone treatment. Larger clinical trials with a longer follow- up and with the inclusion of blood and brain imaging markers are now needed to conclusively determine the significance of testosterone treatment.

Testosterone replacement therapy in older male subjective memory complainers: Double-blind randomized crossover placebo-controlled clinical trial of physiological assessment and safety

Testosterone replacement therapy (TRT) has been investigated in older men as a preventative treatment against Alzheimers disease and dementia. However, previous studies have been contradictory. We assessed TRT physiological effects in 44 older men (aged 61 ± 7.7 years) with subjective memory complaints using a double blind, randomized, crossover, placebo-controlled study. Participants were randomized into 2 groups, one group received transdermal testosterone (50 mg) daily for 24 weeks, followed by a 4 week wash-out period, then 24 weeks of placebo; the other group received the reverse treatment. Blood evaluation revealed significant increases in total testosterone, free (calculated) testosterone, dihydrotestosterone, and a decrease in luteinizing hormone levels (p<0.001) following TRT. Although there were significant increases in red blood cell counts, hemoglobin and prostate specific antigen levels following TRT, they remained within normal ranges. No significant differences in plasma amyloid beta, estradiol, sex hormone binding globulin, insulin levels, body fat percentage, or body mass index were detected. This is the first carefully controlled study that has investigated the influence of TRT in Indonesian men on blood biomarkers linked to dementia risk. Our study suggests TRT is safe and well-tolerated in this Indonesian cohort, yet longitudinal studies with larger cohorts are needed to assess TRT further, and to establish whether TRT reduces dementia risk.

Alterations in erythrocyte fatty acid composition in preclinical Alzheimer's disease

Brain and blood fatty acids (FA) are altered in Alzheimer’s disease and cognitively impaired individuals, however, FA alterations in the preclinical phase, prior to cognitive impairment have not been investigated previously. The current study therefore evaluated erythrocyte FA in cognitively normal elderly participants aged 65–90 years via trans-methylation followed by gas chromatography. The neocortical beta-amyloid load (NAL) measured via positron emission tomography (PET) using ligand 18F-Florbetaben, was employed to categorise participants as low NAL (standard uptake value ratio; SUVR < 1.35, N = 65) and high NAL or preclinical AD (SUVR ≥ 1.35, N = 35) wherein, linear models were employed to compare FA compositions between the two groups. Increased arachidonic acid (AA, p < 0.05) and decreased docosapentaenoic acid (DPA, p < 0.05) were observed in high NAL. To differentiate low from high NAL, the area under the curve (AUC) generated from a ‘base model’ comprising age, gender, APOEε4 and education (AUC = 0.794) was outperformed by base model + AA:DPA (AUC = 0.836). Our findings suggest that specific alterations in erythrocyte FA composition occur very early in the disease pathogenic trajectory, prior to cognitive impairment. As erythrocyte FA levels are reflective of tissue FA, these alterations may provide insight into the pathogenic mechanism(s) of the disease and may highlight potential early diagnostic markers and therapeutic targets.

Elevated plasma ferritin in elderly individuals with high neocortical amyloid-β load

Ferritin, an iron storage and regulation protein, has been associated with Alzheimer’s disease (AD); however, it has not been investigated in preclinical AD, detected by neocortical amyloid-β load (NAL), before cognitive impairment. Cross-sectional analyses were carried out for plasma and serum ferritin in participants in the Kerr Anglican Retirement Village Initiative in Aging Health cohort. Subjects were aged 65–90 years and were categorized into high and low NAL groups via positron emission tomography using a standard uptake value ratio cutoff=1.35. Ferritin was significantly elevated in participants with high NAL compared with those with low NAL, adjusted for covariates age, sex, apolipoprotein E ɛ4 carriage and levels of C-reactive protein (an inflammation marker). Ferritin was also observed to correlate positively with NAL. A receiver operating characteristic curve based on a logistic regression of the same covariates, the base model, distinguished high from low NAL (area under the curve (AUC)=0.766), but was outperformed when plasma ferritin was added to the base model (AUC=0.810), such that at 75% sensitivity, the specificity increased from 62 to 71% on adding ferritin to the base model, indicating that ferritin is a statistically significant additional predictor of NAL over and above the base model. However, ferritin’s contribution alone is relatively minor compared with the base model. The current findings suggest that impaired iron mobilization is an early event in AD pathogenesis. Observations from the present study highlight ferritin’s potential to contribute to a blood biomarker panel for preclinical AD.

Multiple Mechanisms Linking Type 2 Diabetes and Alzheimer’s Disease: Testosterone as a Modifier

Evidence in support of links between type-2 diabetes mellitus (T2DM) and Alzheimers disease (AD) has increased considerably in recent years. AD pathological hallmarks include the accumulation of extracellular amyloid-β (Aβ) and intracellular hyperphosphorylated tau in the brain, which are hypothesized to promote inflammation, oxidative stress, and neuronal loss. T2DM exhibits many AD pathological features, including reduced brain insulin uptake, lipid dysregulation, inflammation, oxidative stress, and depression; T2DM has also been shown to increase AD risk, and with increasing age, the prevalence of both conditions increases. In addition, amylin deposition in the pancreas is more common in AD than in normal aging, and although there is no significant increase in cerebral Aβ deposition in T2DM, the extent of Aβ accumulation in AD correlates with T2DM duration. Given these similarities and correlations, there may be common underlying mechanism(s) that predispose to both T2DM and AD. In other studies, an age-related gradual loss of testosterone and an increase in testosterone resistance has been shown in men; low testosterone levels can also occur in women. In this review, we focus on the evidence for low testosterone levels contributing to an increased risk of T2DM and AD, and the potential of testosterone treatment in reducing this risk in both men and women. However, such testosterone treatment may need to be long-term, and would need regular monitoring to maintain testosterone at physiological levels. It is possible that a combination of testosterone therapy together with a healthy lifestyle approach, including improved diet and exercise, may significantly reduce AD risk.

Alzheimer’s disease: A journey from amyloid Peptides and Oxidative Stress, to Biomarker Technologies and Disease Prevention Strategies—Gains from AIBL and DIAN Cohort Studies

Worldwide there are over 46 million people living with dementia, and this number is expected to double every 20 years reaching about 131 million by 2050. The cost to the community and government health systems, as well as the stress on families and carers is incalculable. Over three decades of research into this disease have been undertaken by several research groups in Australia, including work by our original research group in Western Australia which was involved in the discovery and sequencing of the amyloid-β peptide (also known as Aβ or A4 peptide) extracted from cerebral amyloid plaques. This review discusses the journey from the discovery of the Aβ peptide in Alzheimer’s disease (AD) brain to the establishment of pre-clinical AD using PET amyloid tracers, a method now serving as the gold standard for developing peripheral diagnostic approaches in the blood and the eye. The latter developments for early diagnosis have been largely achieved through the establishment of the Australian Imaging Biomarker and Lifestyle research group that has followed 1,100 Australians for 11 years. AIBL has also been instrumental in providing insight into the role of the major genetic risk factor apolipoprotein E ɛ4, as well as better understanding the role of lifestyle factors particularly diet, physical activity and sleep to cognitive decline and the accumulation of cerebral Aβ.

Association of Plasma Neurofilament Light Chain with Neocortical Amyloid-β Load and Cognitive Performance in Cognitively Normal Elderly Participants

BACKGROUND The disruption of neurofilament, an axonal cytoskeletal protein, in neurodegenerative conditions may result in neuronal damage and its release into the cerebrospinal fluid and blood. In Alzheimers disease (AD), neurofilament light chain (NFL), a neurofilament subunit, is elevated in the cerebrospinal fluid and blood. OBJECTIVE Investigate the association of plasma NFL with preclinical-AD features, such as high neocortical amyloid-β load (NAL) and subjective memory complaints, and cognitive performance in cognitively normal older adults. METHODS Plasma NFL concentrations were measured employing the single molecule array platform in participants from the Kerr Anglican Retirement Village Initiative in Ageing Health cohort, aged 65- 90 years. Participants underwent a battery of neuropsychological testing to evaluate cognitive performance and were categorized as low NAL (NAL-, n = 65) and high NAL (NAL+, n = 35) assessed via PET, and further stratified into subjective memory complainers (SMC; nNAL- = 51, nNAL+ = 25) and non-SMC (nNAL- = 14, nNAL+ = 10) based on the Memory Assessment Clinic- Questionnaire. RESULTS Plasma NFL inversely correlated with cognitive performance. No significant difference in NFL was observed between NAL+ and NAL- participants; however, within APOEɛ4 non-carriers, higher NAL was observed in individuals with NFL concentrations within quartiles 3 and 4 (versus quartile 1). Additionally, within the NAL+ participants, SMC had a trend of higher NFL compared to non-SMC. CONCLUSION Plasma NFL is inversely associated with cognitive performance in elderly individuals. While plasma NFL may not reflect NAL in individuals with normal global cognition, the current observations indicate that onset of axonal injury, reflected by increased plasma NFL, within the preclinical phase of AD may contribute to the pathogenesis of AD.

Alterations in serum kynurenine pathway metabolites in individuals with high neocortical amyloid-β load: A pilot study

The kynurenine pathway (KP) is dysregulated in neuroinflammatory diseases including Alzheimer’s disease (AD), however has not been investigated in preclinical AD characterized by high neocortical amyloid-β load (NAL), prior to cognitive impairment. Serum KP metabolites were measured in the cognitively normal KARVIAH cohort. Participants, aged 65–90 y, were categorised into NAL+ (n = 35) and NAL− (n = 65) using a standard uptake value ratio cut-off = 1.35. Employing linear models adjusting for age and APOEε4, higher kynurenine and anthranilic acid (AA) in NAL+ versus NAL− participants were observed in females (kynurenine, p = 0.004; AA, p = 0.001) but not males (NALxGender, p = 0.001, 0.038, respectively). To evaluate the predictive potential of kynurenine or/and AA for NAL+ in females, logistic regressions with NAL+/− as outcome were carried out. After age and APOEε4 adjustment, kynurenine and AA were individually and jointly significant predictors (p = 0.007, 0.005, 0.0004, respectively). Areas under the receiver operating characteristic curves were 0.794 using age and APOEε4 as predictors, and 0.844, 0.866 and 0.871 when kynurenine, AA and both were added. Findings from the current study exhibit increased KP activation in NAL+ females and highlight the predictive potential of KP metabolites, AA and kynurenine, for NAL+. Additionally, the current study also provides insight into he influence of gender in AD pathogenesis.

Selective spatiotemporal vulnerability of central nervous system neurons to pathologic TAR DNA-binding protein 43 in aged transgenic mice

Amyotrophic lateral sclerosis (ALS) is a rapidly progressing and fatal disease characterized by muscular atrophy because of loss of upper and lower motor neurons. Histopathologically, most patients with ALS have abnormal cytoplasmic accumulation and aggregation of the nuclear RNA-regulating protein TAR DNA-binding protein 43 (TDP-43). Pathogenic mutations in the TARDBP gene that encode TDP-43 have been identified in familial ALS. We have previously reported transgenic mice with neuronal expression of human TDP-43 carrying the pathogenic A315T mutation (iTDP-43A315T mice), presenting with early-onset motor deficits in adolescent animals. Here, we analyzed aged iTDP-43A315T mice, focusing on the spatiotemporal profile and progression of neurodegeneration in upper and lower motor neurons. Magnetic resonance imaging and histologic analysis revealed a differential loss of upper motor neurons in a hierarchical order as iTDP-43A315T mice aged. Furthermore, we report progressive gait problems, profound motor deficits, and muscle atrophy in aged iTDP-43A315T mice. Despite these deficits and TDP-43 pathologic disorders in lower motor neurons, stereological analysis did not show cell loss in spinal cords. Taken together, neuronal populations in aging iTDP-43A315T mice show differential susceptibility to the expression of human TDP-43A315T.

ELEVATED KYNURENINE AND ANTHRANILIC ACID LEVELS IN ELDERLY FEMALES WITH HIGH NEOCORTICAL AMYLOID-BETA LOAD

compared to non-carriers. To determine whether APOE ε4 associates with altered blood mitochondrial biomarkers from AD subjects, we measured mitochondrial biomarkers in platelets and lymphocytes from ADAPOE ε4 carriers and non-carriers.Methods: Platelet mitochondria COX and citrate synthase (CS) Vmax activity were measured in 22 APOE ε4 carrier and 20 non-carrier subjects. Lymphocyte mitochondrial biomarkers (JC1, MitoSox, MitoTracker /Annexin V) were measured in 14 APOE ε4 carrier and 12 non-carrier sujbects. All subjects had AD by McKhann et al. criteria and were CDR 0.5 or 1. 40 ml of blood was collected in tubes containing acid-citrate-dextrose anticoagulant. Plateletrich plasma and buffy coat were isolated by centrifugation, which was centrifuged to pellet platelets and lymphocytes. Lymphocytes were stained with Mitotracker Red/Annexin V, MitoSox Red, or JC-1 and flurosecence was measured via flow cytometry. Mitochondria were isolated from platelets via nitrogen cavitation and centrifugation. The platelet mitochondria were resuspended, and added to a cuvette to spectrophotometrically determine the rate of conversion of reduced cytochrome c to its oxidized form (for COX), and the formation of 5-thio-2-nitrobenzoate (for CS). For COX, the resulting pseudo-first order rate constant (sec) was normalized to protein to yield the final activity value (sec/ mg protein) or to the CS activity (COX/CS). The CS rate was normalized to mg protein to yield the final activity (nmol/mg/mg protein) Group means were compared using Student’s T-test or One-Way ANOVA (posthoc LSD). Results: Mean CS and COX Vmax activities (normalized to protein content) were lower in the APOE ε4 carriers (p<0.05). As a result COX/CS ratios were comparable between groups. Lymphocyte Annexin V was significantly higher in APOE ε4 carriers (p<0.05). A trend for increased lymphocyte MitoSox and increased red/green JC1 ratio in APOE ε4 carriers was also observed. Conclusions: Our data support a relationship between APOE genotype and mitochondrial function. Studies to elucidate molecular pathways affected by APOE genotype in AD lymphocytes are in progress.