Cholesterol and matrisome pathways dysregulated in astrocytes and microglia.

The impact of apolipoprotein E ε4 (APOE4), the strongest genetic risk factor for Alzheimer's disease (AD), on human brain cellular function remains unclear. Here, we investigated the effects of APOE4 on brain cell types derived from population and isogenic human induced pluripotent stem cells, post-mortem brain, and APOE targeted replacement mice. Population and isogenic models demonstrate that APOE4 local haplotype, rather than a single risk allele, contributes to risk. Global transcriptomic analyses reveal human-specific, APOE4-driven lipid metabolic dysregulation in astrocytes and microglia. APOE4 enhances de novo cholesterol synthesis despite elevated intracellular cholesterol due to lysosomal cholesterol sequestration in astrocytes. Further, matrisome dysregulation is associated with upregulated chemotaxis, glial activation, and lipid biosynthesis in astrocytes co-cultured with neurons, which recapitulates altered astrocyte matrisome signaling in human brain. Thus, APOE4 initiates glia-specific cell and non-cell autonomous dysregulation that may contribute to increased AD risk.

Agreement in Squat Jump Force-Time Characteristics Between Smith Machine and Free-Weight Squat Jump Force-Time Characteristics.

ABSTRACT: Kotani, Y, Lake, J, Guppy, SN, Poon, W, Nosaka, K, and Haff, GG. Agreement in squat jump force-time characteristics between Smith machine and free-weight squat jump force-time characteristics. J Strength Cond Res 37(10): 1955-1962, 2023-The purpose of this study was to determine whether squat jump (SJ) force-velocity (FV) and load-velocity (LV) profiles created using free-weights agree with profiles created with a Smith machine. Fifteen resistance-trained male subjects (age = 26.4 ± 2.5 years; height = 1.75 ± 0.09 m; body mass = 82.6 ± 13.4 kg) participated in this study. All subjects completed 2 familiarization and 2 experimental sessions using both the Smith machine and free-weight SJs each separated by 48 hours. During the experimental trials, progressively loaded SJs were performed in a quasi-randomized block order with loads between 21 kg and 100% of the subject's body mass. Agreement between exercise mode was determined with a weighted least products regression analysis. No fixed or proportional bias was noted between exercise modes when using peak velocity (PV) and mean velocity (MV) to create an FV profile. There was no fixed and proportional bias present for the LV profile when the profile was created with PV. When the LV profile was calculated from MV, fixed and proportional bias were present, indicating that MVs were significantly different between exercise modes. In addition, the free-weight FV and LV profiles exhibited poor to good relative and good to poor absolute reliability. Furthermore, when created using the Smith machine, both profiles exhibited poor to moderate relative and absolute reliability. Based on these data, caution should be used when interpreting LV and FV profiles created with these 2 methods.

Effect of 3 Different Set Configurations on Kinematic Variables and Internal Loads During a Power Snatch Session.

ABSTRACT: Nagatani, T, Kendall, KL, Guppy, SN, Poon, WCK, and Haff, GG. Effect of 3 different set configurations on kinematic variables and internal loads during a power snatch session. J Strength Cond Res 37(10): 1929-1938, 2023-The aim of this study was to investigate the effect of 3 different set configurations on kinematic variables and internal loads during multiple sets performed with the power snatch. Ten strength-power athletes with at least 6 months of training experience performing the power snatch participated in this study, which consisted of 3 experimental protocols performed in a randomized repeated-measures design. The 3 protocols involved performing the power snatch for 3 sets of 5 repetitions at an average load of 75% 1 repetition maximum with a traditional (TRAD), cluster (CLU), or ascending cluster (A-CLU) protocol, where the training load was progressively increased across the set. Kinematic variables and internal loads (heart rate, blood lactate, and rate of perceived exertion) were measured during each protocol. The athletes maintained peak velocity (PV) and peak power (PP) and exhibited lower internal loads during CLU sets when compared with TRAD sets, whereas they displayed significant decreases in PV during TRAD sets. However, there were no statistically significant differences in PV and PP responses between the TRAD and CLU protocol. The athletes exhibited a significant decrease in PV, whereas PP was increased across each set in the A-CLU protocol, with lower internal loads observed compared with the TRAD protocol. Overall, the training loads used in this study do not appear to maximize the benefits of using CLU set during 3 sets of power snatches performed for 5 repetitions. In addition, A-CLU sets may potentially be useful as a means of maximizing the power output of the athlete.

Dynamic and Isometric Force-Time Curve Characteristics Influencing Change of Direction Performance of State-Level Netball Players.

ABSTRACT: Smolarek, T, Haff, GG, Poon, WCK, Nagatani, T, Barley, OR, and Guppy, SN. Dynamic and isometric force-time curve characteristics influencing change of direction performance of state-level netball players. J Strength Cond Res 37(12): 2397-2404, 2023-Although multiple lower-body strength capacities are required to change direction rapidly, there is limited knowledge about the relative importance of these factors. Therefore, the purpose of this study was to assess the relationship between dynamic and isometric lower-body strength capacities and change of direction (COD) time in state-league netball players. Seventeen female athletes completed maximal isometric midthigh pull (IMTP), countermovement jump (CMJ), and modified 5-0-5 COD tests. Pearson's product moment correlations were used to determine the relationship between COD time and several IMTP and CMJ force-time curve characteristics. To assess the level of contribution of each force-time curve characteristic to COD time, multivariate-linear stepwise regression analyses were performed. A significant moderate correlation was noted between net relative peak force (PF) during the IMTP and COD time ( r = 0.488, p = 0.047), accounting for 23.8% of the variance in COD time. Moreover, concentric relative impulse during the CMJ was strongly correlated with COD time ( r = 0.718; p = 0.001), explaining 81.9% of the variance in COD time when combined with net relative braking PF in a stepwise regression. Based on these findings, female netball players who display higher concentric and isometric strength, as well as the ability to express higher impulses during the concentric phase of the CMJ, are likely to perform CODs faster. This may occur because COD requires the generation of greater propulsive forces, as well as reduced braking and contact times, along with greater isometric strength enabling effective repositioning of center of mass during COD tasks.

The effect of cold water immersion on the recovery of physical performance revisited: A systematic review with meta-analysis.

This review evaluated the effect of CWI on the temporal recovery profile of physical performance, accounting for environmental conditions and prior exercise modality. Sixty-eight studies met the inclusion criteria. Standardised mean differences were calculated for parameters assessed at <1, 1-6, 24, 48, 72 and ≥96 h post-immersion. CWI improved short-term recovery of endurance performance (p = 0.01, 1 h), but impaired sprint (p = 0.03, 1 h) and jump performance (p = 0.04, 6h). CWI improved longer-term recovery of jump performance (p < 0.01-0.02, 24 h and 96 h) and strength (p < 0.01, 24 h), which coincided with decreased creatine kinase (p < 0.01-0.04, 24-72 h), improved muscle soreness (p < 0.01-0.02, 1-72 h) and perceived recovery (p < 0.01, 72 h). CWI improved the recovery of endurance performance following exercise in warm (p < 0.01) and but not in temperate conditions (p = 0.06). CWI improved strength recovery following endurance exercise performed at cool-to-temperate conditions (p = 0.04) and enhanced recovery of sprint performance following resistance exercise (p = 0.04). CWI seems to benefit the acute recovery of endurance performance, and longer-term recovery of muscle strength and power, coinciding with changes in muscle damage markers. This, however, depends on the nature of the preceding exercise.

Reliability of the Squat Jump Force-Velocity and Load-Velocity Profiles.

ABSTRACT: Kotani, Y, Lake, J, Guppy, SN, Poon, W, Nosaka, K, Hori, N, and Haff, GG. The reliability of the squat jump force-velocity and load-velocity profiles. J Strength Cond Res 36(11): 3000-3007, 2022-The purpose of this study was to investigate the between-session reliability of the squat jump force-velocity (FV) and load-velocity (LV) profiles. Eighteen subjects (age = 28.1 ± 4.8 years; height = 1.7 ± 9.7; body mass = 74.7 ± 12.8) who could back squat >1.5 times body mass participated in this study. Each subject completed a familiarization session, followed by 2 experimental sessions each separated by 72 hours. Subjects performed a series of squat jumps on a force plate against external loads between 0 and 100% of their body mass in a quasi-randomized block order. Intraclass correlation coefficient (ICC) and coefficient of variation (CV) were used to examine the between-session reliability. Peak velocity (PV) and mean velocity (MV) at each load were highly reliable (ICC >0.80, CV% <7.41, SEM <0.13 m·s -1 , smallest detectable difference [SDD] <0.31 m·s -1 , ES <0.21). Force-velocity profiles created with peak force and relative peak force resulted in poor to excellent reliability (ICC = 0.34-0.92, CV% = 11.9-26.3). When mean and relative mean forces were used to create FV profiles, there was poor to good reliability (ICC = 0.03-0.85, CV% = 18.1-39.4). When the LV profile was calculated with PV (ICC = 0.60-0.90, CV% = 7.9-16.9) or MV (ICC = 0.49-0.91, CV% = 11.1-23.4), there was poor to excellent reliability. There was no time effect found between sessions for both FV and LV profiles. The squat jump FV and LV profiles established with a force plate are not reliable. Therefore, these profiles are not recommended to be used to inform programming decisions.

Exosomal tau with seeding activity is released from Alzheimer's disease synapses, and seeding potential is associated with amyloid beta.

Synaptic transfer of tau has long been hypothesized from the human pathology pattern and has been demonstrated in vitro and in vivo, but the precise mechanisms remain unclear. Extracellular vesicles such as exosomes have been suggested as a mechanism, but not all tau is exosomal. The present experiments use a novel flow cytometry assay to quantify depolarization of synaptosomes by KCl after loading with FM2-10, which induces a fluorescence reduction associated with synaptic vesicle release; the degree of reduction in cryopreserved human samples equaled that seen in fresh mouse synaptosomes. Depolarization induced the release of vesicles in the size range of exosomes, along with tetraspanin markers of extracellular vesicles. A number of tau peptides were released, including tau oligomers; released tau was primarily unphosphorylated and C-terminal truncated, with Aß release just above background. When exosomes were immunopurified from release supernatants, a prominent tau band showed a dark smeared appearance of SDS-stable oligomers along with the exosomal marker syntenin-1, and these exosomes induced aggregation in the HEK tau biosensor assay. However, the flow-through did not seed aggregation. Size exclusion chromatography of purified released exosomes shows faint signals from tau in the same fractions that show a CD63 band, an exosomal size signal, and seeding activity. Crude synaptosomes from control, tauopathy, and AD cases demonstrated lower seeding in tauopathy compared to AD that is correlated with the measured Aß42 level. These results show that AD synapses release exosomal tau that is C-terminal-truncated, oligomeric, and with seeding activity that is enhanced by Aß. Taken together with previous findings, these results are consistent with a direct prion-like heterotypic seeding of tau by Aß within synaptic terminals, with subsequent loading of aggregated tau onto exosomes that are released and competent for tau seeding activity.

α-Synuclein in blood exosomes immunoprecipitated using neuronal and oligodendroglial markers distinguishes Parkinson's disease from multiple system atrophy.

The diagnosis of Parkinson's disease (PD) and atypical parkinsonian syndromes is difficult due to the lack of reliable, easily accessible biomarkers. Multiple system atrophy (MSA) is a synucleinopathy whose symptoms often overlap with PD. Exosomes isolated from blood by immunoprecipitation using CNS markers provide a window into the brain's biochemistry and may assist in distinguishing between PD and MSA. Thus, we asked whether α-synuclein (α-syn) in such exosomes could distinguish among healthy individuals, patients with PD, and patients with MSA. We isolated exosomes from the serum or plasma of these three groups by immunoprecipitation using neuronal and oligodendroglial markers in two independent cohorts and measured α-syn in these exosomes using an electrochemiluminescence ELISA. In both cohorts, α-syn concentrations were significantly lower in the control group and significantly higher in the MSA group compared to the PD group. The ratio between α-syn concentrations in putative oligodendroglial exosomes compared to putative neuronal exosomes was a particularly sensitive biomarker for distinguishing between PD and MSA. Combining this ratio with the α-syn concentration itself and the total exosome concentration, a multinomial logistic model trained on the discovery cohort separated PD from MSA with an AUC = 0.902, corresponding to 89.8% sensitivity and 86.0% specificity when applied to the independent validation cohort. The data demonstrate that a minimally invasive blood test measuring α-syn in blood exosomes immunoprecipitated using CNS markers can distinguish between patients with PD and patients with MSA with high sensitivity and specificity. Future optimization and validation of the data by other groups would allow this strategy to become a viable diagnostic test for synucleinopathies.

Changes in plasma hydroxyproline and plasma cell-free DNA concentrations after higher- versus lower-intensity eccentric cycling.

PURPOSE: We examined changes in plasma creatine kinase (CK) activity, hydroxyproline and cell-free DNA (cfDNA) concentrations in relation to changes in maximum voluntary isometric contraction (MVIC) torque and delayed-onset muscle soreness (DOMS) following a session of volume-matched higher- (HI) versus lower-intensity (LI) eccentric cycling exercise. METHODS: Healthy young men performed either 5 × 1-min HI at 20% of peak power output (n = 11) or 5 × 4-min LI eccentric cycling at 5% of peak power output (n = 9). Changes in knee extensor MVIC torque, DOMS, plasma CK activity, and hydroxyproline and cfDNA concentrations before, immediately after, and 24-72 h post-exercise were compared between groups. RESULTS: Plasma CK activity increased post-exercise (141 ± 73.5%) and MVIC torque decreased from immediately (13.3 ± 7.8%) to 48 h (6.7 ± 13.5%) post-exercise (P < 0.05), without significant differences between groups. DOMS was greater after HI (peak: 4.5 ± 3.0 on a 10-point scale) than LI (1.2 ± 1.0). Hydroxyproline concentration increased 40-53% at 24-72 h after both LI and HI (P < 0.05). cfDNA concentration increased immediately after HI only (2.3 ± 0.9-fold, P < 0.001), with a significant difference between groups (P = 0.002). Lack of detectable methylated HOXD4 indicated that the cfDNA was not derived from skeletal muscle. No significant correlations were evident between the magnitude of change in the measures, but the cfDNA increase immediately post-exercise was correlated with the maximal change in heart rate during exercise (r = 0.513, P = 0.025). CONCLUSION: Changes in plasma hydroxyproline and cfDNA concentrations were not associated with muscle fiber damage, but the increased hydroxyproline in both groups suggests increased collagen turnover. cfDNA may be a useful metabolic-intensity exercise marker.

Apolipoprotein E/Amyloid-ß Complex Accumulates in Alzheimer Disease Cortical Synapses via Apolipoprotein E Receptors and Is Enhanced by APOE4.

Apolipoprotein E (apoE) colocalizes with amyloid-ß (Aß) in Alzheimer disease (AD) plaques and in synapses, and evidence suggests that direct interactions between apoE and Aß are important for apoE's effects in AD. The present work examines the hypothesis that apoE receptors mediate uptake of apoE/Aß complex into synaptic terminals. Western blot analysis shows multiple SDS-stable assemblies in synaptosomes from human AD cortex; apoE/Aß complex was markedly increased in AD compared with aged control samples. Complex formation between apoE and Aß was confirmed by coimmunoprecipitation experiments. The apoE receptors low-density lipoprotein receptor (LDLR) and LDLR-related protein 1 (LRP1) were quantified in synaptosomes using flow cytometry, revealing up-regulation of LRP1 in early- and late-stage AD. Dual-labeling flow cytometry analysis of LRP1- and LDLR positives indicate most (approximately 65%) of LDLR and LRP1 is associated with postsynaptic density-95 (PSD-95)-positive synaptosomes, indicating that remaining LRP1 and LDLR receptors are exclusively presynaptic. Flow cytometry analysis of Nile red labeling revealed a reduction in cholesterol esters in AD synaptosomes. Dual-labeling experiments showed apoE and Aß concentration into LDLR and LRP1-positive synaptosomes, along with free and esterified cholesterol. Synaptic Aß was increased by apoE4 in control and AD samples. These results are consistent with uptake of apoE/Aß complex and associated lipids into synaptic terminals, with subsequent Aß clearance in control synapses and accumulation in AD synapses.

Comparison between high- and low-intensity eccentric cycling of equal mechanical work for muscle damage and the repeated bout effect.

PURPOSE: We compared high- and low-intensity eccentric cycling (ECC) with the same mechanical work for changes in muscle function and muscle soreness, and examined the changes after subsequent high-intensity ECC. METHODS: Twenty men performed either high-intensity ECC (1 min × 5 at 20% of peak power output: PPO) for two bouts separated by 2 weeks (H-H, n = 11), or low-intensity (4 min × 5 at 5% PPO) for the first and high-intensity ECC for the second bout (L-H, n = 9). Changes in indirect muscle damage markers were compared between groups and bouts. RESULTS: At 24 h after the first bout, both groups showed similar decreases in maximal isometric (70° knee angle, - 10.6 ± 11.8%) and isokinetic ( - 11.0 ± 8.2%) contraction torque of the knee extensors (KE), squat ( - 7.7 ± 10.4%) and counter-movement jump ( - 5.9 ± 8.4%) heights (p < 0.05). Changes in KE torque and jump height were smaller after the second than the first bout for both the groups (p < 0.05). Increases in plasma creatine kinase activity were small, and no significant changes in vastus lateralis or intermedius thickness nor ultrasound echo-intensity were observed. KE soreness with palpation was greater (p < 0.01) in H-H (peak: 4.2 ± 1.0) than L-H (1.4 ± 0.6) after the first bout, but greater in L-H (3.6 ± 0.9) than H-H (1.5 ± 0.5) after the second bout. This was also found for muscle soreness with squat, KE stretch and gluteal palpation. CONCLUSION: The high- and low-intensity ECC with matched mechanical work induced similar decreases in muscle function, but DOMS was greater after high-intensity ECC, which may be due to greater extracellular matrix damage and inflammation.

The adaptive immune system restrains Alzheimer's disease pathogenesis by modulating microglial function.

The innate immune system is strongly implicated in the pathogenesis of Alzheimer's disease (AD). In contrast, the role of adaptive immunity in AD remains largely unknown. However, numerous clinical trials are testing vaccination strategies for AD, suggesting that T and B cells play a pivotal role in this disease. To test the hypothesis that adaptive immunity influences AD pathogenesis, we generated an immune-deficient AD mouse model that lacks T, B, and natural killer (NK) cells. The resulting "Rag-5xfAD" mice exhibit a greater than twofold increase in ß-amyloid (Aß) pathology. Gene expression analysis of the brain implicates altered innate and adaptive immune pathways, including changes in cytokine/chemokine signaling and decreased Ig-mediated processes. Neuroinflammation is also greatly exacerbated in Rag-5xfAD mice as indicated by a shift in microglial phenotype, increased cytokine production, and reduced phagocytic capacity. In contrast, immune-intact 5xfAD mice exhibit elevated levels of nonamyloid reactive IgGs in association with microglia, and treatment of Rag-5xfAD mice or microglial cells with preimmune IgG enhances Aß clearance. Last, we performed bone marrow transplantation studies in Rag-5xfAD mice, revealing that replacement of these missing adaptive immune populations can dramatically reduce AD pathology. Taken together, these data strongly suggest that adaptive immune cell populations play an important role in restraining AD pathology. In contrast, depletion of B cells and their appropriate activation by T cells leads to a loss of adaptive-innate immunity cross talk and accelerated disease progression.

Synaptic Amyloid-ß Oligomers Precede p-Tau and Differentiate High Pathology Control Cases.

Amyloid-ß (Aß) and hyperphosphorylated tau (p-tau) aggregates form the two discrete pathologies of Alzheimer disease (AD), and oligomeric assemblies of each protein are localized to synapses. To determine the sequence by which pathology appears in synapses, Aß and p-tau were quantified across AD disease stages in parietal cortex. Nondemented cases with high levels of AD-related pathology were included to determine factors that confer protection from clinical symptoms. Flow cytometric analysis of synaptosome preparations was used to quantify Aß and p-tau in large populations of individual synaptic terminals. Soluble Aß oligomers were assayed by a single antibody sandwich enzyme-linked immunosorbent assay. Total in situ Aß was elevated in patients with early- and late-stage AD dementia, but not in high pathology nondemented controls compared with age-matched normal controls. However, soluble Aß oligomers were highest in early AD synapses, and this assay distinguished early AD cases from high pathology controls. Overall, synapse-associated p-tau did not increase until late-stage disease in human and transgenic rat cortex, and p-tau was elevated in individual Aß-positive synaptosomes in early AD. These results suggest that soluble oligomers in surviving neocortical synaptic terminals are associated with dementia onset and suggest an amyloid cascade hypothesis in which oligomeric Aß drives phosphorylated tau accumulation and synaptic spread. These results indicate that antiamyloid therapies will be less effective once p-tau pathology is developed.

Pre-synaptic C-terminal truncated tau is released from cortical synapses in Alzheimer's disease.

The microtubule-associated protein tau has primarily been associated with axonal location and function; however, recent work shows tau release from neurons and suggests an important role for tau in synaptic plasticity. In our study, we measured synaptic levels of total tau using synaptosomes prepared from cryopreserved human postmortem Alzheimer's disease (AD) and control samples. Flow cytometry data show that a majority of synaptic terminals are highly immunolabeled with the total tau antibody (HT7) in both AD and control samples. Immunoblots of synaptosomal fractions reveal increases in a 20 kDa tau fragment and in tau dimers in AD synapses, and terminal-specific antibodies show that in many synaptosome samples tau lacks a C-terminus. Flow cytometry experiments to quantify the extent of C-terminal truncation reveal that only 15-25% of synaptosomes are positive for intact C-terminal tau. Potassium-induced depolarization demonstrates release of tau and tau fragments from pre-synaptic terminals, with increased release from AD compared to control samples. This study indicates that tau is normally highly localized to synaptic terminals in cortex where it is well-positioned to affect synaptic plasticity. Tau cleavage may facilitate tau aggregation as well as tau secretion and propagation of tau pathology from the pre-synaptic compartment in AD. Results demonstrate the abundance of tau, mainly C-terminal truncated tau, in synaptic terminals in aged control and in Alzheimer's disease (AD) samples. Tau fragments and dimers/oligomers are prominent in AD synapses. Following depolarization, tau release is potentiated in AD nerve terminals compared to aged controls. We hypothesize (i) endosomal release of the different tau peptides from AD synapses, and (ii) together with phosphorylation, fragmentation of synaptic tau exacerbates tau aggregation, synaptic dysfunction, and the spread of tau pathology in AD. Aß = amyloid-beta.

Human neural stem cells improve cognition and promote synaptic growth in two complementary transgenic models of Alzheimer's disease and neuronal loss.

Alzheimer's disease (AD) is the most prevalent age-related neurodegenerative disorder, affecting over 35 million people worldwide. Pathologically, AD is characterized by the progressive accumulation of ß-amyloid (Aß) plaques and neurofibrillary tangles within the brain. Together, these pathologies lead to marked neuronal and synaptic loss and corresponding impairments in cognition. Current treatments, and recent clinical trials, have failed to modify the clinical course of AD; thus, the development of novel and innovative therapies is urgently needed. Over the last decade, the potential use of stem cells to treat cognitive impairment has received growing attention. Specifically, neural stem cell transplantation as a treatment for AD offers a novel approach with tremendous therapeutic potential. We previously reported that intrahippocampal transplantation of murine neural stem cells (mNSCs) can enhance synaptogenesis and improve cognition in 3xTg-AD mice and the CaM/Tet-DT(A) model of hippocampal neuronal loss. These promising findings prompted us to examine a human neural stem cell population, HuCNS-SC, which has already been clinically tested for other neurodegenerative disorders. In this study, we provide the first evidence that transplantation of research grade HuCNS-SCs can improve cognition in two complementary models of neurodegeneration. We also demonstrate that HuCNS-SC cells can migrate and differentiate into immature neurons and glia and significantly increase synaptic and growth-associated markers in both 3xTg-AD and CaM/Tet-DTA mice. Interestingly, improvements in aged 3xTg-AD mice were not associated with altered Aß or tau pathology. Rather, our findings suggest that human NSC transplantation improves cognition by enhancing endogenous synaptogenesis. Taken together, our data provide the first preclinical evidence that human NSC transplantation could be a safe and effective therapeutic approach for treating AD.

ß-Amyloid (Aß) oligomers impair brain-derived neurotrophic factor retrograde trafficking by down-regulating ubiquitin C-terminal hydrolase, UCH-L1.

We previously found that BDNF-dependent retrograde trafficking is impaired in AD transgenic mouse neurons. Utilizing a novel microfluidic culture chamber, we demonstrate that Aß oligomers compromise BDNF-mediated retrograde transport by impairing endosomal vesicle velocities, resulting in impaired downstream signaling driven by BDNF/TrkB, including ERK5 activation, and CREB-dependent gene regulation. Our data suggest that a key mechanism mediating the deficit involves ubiquitin C-terminal hydrolase L1 (UCH-L1), a deubiquitinating enzyme that functions to regulate cellular ubiquitin. Aß-induced deficits in BDNF trafficking and signaling are mimicked by LDN (an inhibitor of UCH-L1) and can be reversed by increasing cellular UCH-L1 levels, demonstrated here using a transducible TAT-UCH-L1 strategy. Finally, our data reveal that UCH-L1 mRNA levels are decreased in the hippocampi of AD brains. Taken together, our data implicate that UCH-L1 is important for regulating neurotrophin receptor sorting to signaling endosomes and supporting retrograde transport. Further, our results support the idea that in AD, Aß may down-regulate UCH-L1 in the AD brain, which in turn impairs BDNF/TrkB-mediated retrograde signaling, compromising synaptic plasticity and neuronal survival.

Levels of soluble apolipoprotein E/amyloid-ß (Aß) complex are reduced and oligomeric Aß increased with APOE4 and Alzheimer disease in a transgenic mouse model and human samples.

Human apolipoprotein E (apoE) isoforms may differentially modulate amyloid-ß (Aß) levels. Evidence suggests physical interactions between apoE and Aß are partially responsible for these functional effects. However, the apoE/Aß complex is not a single static structure; rather, it is defined by detection methods. Thus, literature results are inconsistent and difficult to interpret. An ELISA was developed to measure soluble apoE/Aß in a single, quantitative method and was used to address the hypothesis that reduced levels of soluble apoE/Aß and an increase in soluble Aß, specifically oligomeric Aß (oAß), are associated with APOE4 and AD. Previously, soluble Aß42 and oAß levels were greater with APOE4 compared with APOE2/APOE3 in hippocampal homogenates from EFAD transgenic mice (expressing five familial AD mutations and human apoE isoforms). In this study, soluble apoE/Aß levels were lower in E4FAD mice compared with E2FAD and E3FAD mice, thus providing evidence that apoE/Aß levels isoform-specifically modulate soluble oAß clearance. Similar results were observed in soluble preparations of human cortical synaptosomes; apoE/Aß levels were lower in AD patients compared with controls and lower with APOE4 in the AD cohort. In human CSF, apoE/Aß levels were also lower in AD patients and with APOE4 in the AD cohort. Importantly, although total Aß42 levels decreased in AD patients compared with controls, oAß levels increased and were greater with APOE4 in the AD cohort. Overall, apoE isoform-specific formation of soluble apoE/Aß modulates oAß levels, suggesting a basis for APOE4-induced AD risk and a mechanistic approach to AD biomarkers.

Intracellular amyloid and the neuronal origin of Alzheimer neuritic plaques.

Genetic analysis of familial forms of Alzheimer's disease (AD) causally links the proteolytic processing of the amyloid precursor protein (APP) and AD. However, the specific type of amyloid and mechanisms of amyloid pathogenesis remain unclear. We conducted a detailed analysis of intracellular amyloid with an aggregation specific conformation dependent monoclonal antibody, M78, raised against fibrillar Aß42. M78 immunoreactivity colocalizes with Aß and the carboxyl terminus of APP (APP-CTF) immunoreactivities in perinuclear compartments at intermediate times in 10month 3XTg-AD mice, indicating that this represents misfolded and aggregated protein rather than normally folded APP. At 12months, M78 immunoreactivity also accumulates in the nucleus. Neuritic plaques at 12months display the same spatial organization of centrally colocalized M78, diffuse chromatin and neuronal nuclear NeuN staining surrounded by peripheral M78 and APP-CTF immunoreactivity as observed in neurons, indicating that neuritic plaques arise from degenerating neurons with intracellular amyloid immunoreactivity. The same staining pattern was observed in neuritic plaques in human AD brains, showing elevated intracellular M78 immunoreactivity at intermediate stages of amyloid pathology (Braak A and B) compared to no amyloid pathology and late stage amyloid pathology (Braak 0 and C, respectively). These results indicate that intraneuronal protein aggregation and amyloid accumulation is an early event in AD and that neuritic plaques are initiated by the degeneration and death of neurons by a mechanism that may be related to the formation of extracellular traps by neutrophils.

ABCC9 gene polymorphism is associated with hippocampal sclerosis of aging pathology.

Hippocampal sclerosis of aging (HS-Aging) is a high-morbidity brain disease in the elderly but risk factors are largely unknown. We report the first genome-wide association study (GWAS) with HS-Aging pathology as an endophenotype. In collaboration with the Alzheimer's Disease Genetics Consortium, data were analyzed from large autopsy cohorts: (#1) National Alzheimer's Coordinating Center (NACC); (#2) Rush University Religious Orders Study and Memory and Aging Project; (#3) Group Health Research Institute Adult Changes in Thought study; (#4) University of California at Irvine 90+ Study; and (#5) University of Kentucky Alzheimer's Disease Center. Altogether, 363 HS-Aging cases and 2,303 controls, all pathologically confirmed, provided statistical power to test for risk alleles with large effect size. A two-tier study design included GWAS from cohorts #1-3 (Stage I) to identify promising SNP candidates, followed by focused evaluation of particular SNPs in cohorts #4-5 (Stage II). Polymorphism in the ATP-binding cassette, sub-family C member 9 (ABCC9) gene, also known as sulfonylurea receptor 2, was associated with HS-Aging pathology. In the meta-analyzed Stage I GWAS, ABCC9 polymorphisms yielded the lowest p values, and factoring in the Stage II results, the meta-analyzed risk SNP (rs704178:G) attained genome-wide statistical significance (p = 1.4 × 10(-9)), with odds ratio (OR) of 2.13 (recessive mode of inheritance). For SNPs previously linked to hippocampal sclerosis, meta-analyses of Stage I results show OR = 1.16 for rs5848 (GRN) and OR = 1.22 rs1990622 (TMEM106B), with the risk alleles as previously described. Sulfonylureas, a widely prescribed drug class used to treat diabetes, also modify human ABCC9 protein function. A subsample of patients from the NACC database (n = 624) were identified who were older than age 85 at death with known drug history. Controlling for important confounders such as diabetes itself, exposure to a sulfonylurea drug was associated with risk for HS-Aging pathology (p = 0.03). Thus, we describe a novel and targetable dementia risk factor.