EK100 and Antrodin C Improve Brain Amyloid Pathology in APP/PS1 Transgenic Mice by Promoting Microglial and Perivascular Clearance Pathways

Alzheimer’s disease (AD) is characterized by the deposition of β-amyloid peptide (Aβ). There are currently no drugs that can successfully treat this disease. This study first explored the anti-inflammatory activity of seven components isolated from Antrodia cinnamonmea in BV2 cells and selected EK100 and antrodin C for in vivo research. APPswe/PS1dE9 mice were treated with EK100 and antrodin C for one month to evaluate the effect of these reagents on AD-like pathology by nesting behavior, immunohistochemistry, and immunoblotting. Ergosterol and ibuprofen were used as control. EK100 and antrodin C improved the nesting behavior of mice, reduced the number and burden of amyloid plaques, reduced the activation of glial cells, and promoted the perivascular deposition of Aβ in the brain of mice. EK100 and antrodin C are significantly different in activating astrocytes, regulating microglia morphology, and promoting plaque-associated microglia to express oxidative enzymes. In contrast, the effects of ibuprofen and ergosterol are relatively small. In addition, EK100 significantly improved hippocampal neurogenesis in APPswe/PS1dE9 mice. Our data indicate that EK100 and antrodin C reduce the pathology of AD by reducing amyloid deposits and promoting nesting behavior in APPswe/PS1dE9 mice through microglia and perivascular clearance, indicating that EK100 and antrodin C have the potential to be used in AD treatment.     

Metals in ALS TDP-43 Pathology

Amyotrophic lateral sclerosis (ALS), Alzheimer’s disease, Parkinson’s disease and similar neurodegenerative disorders take their toll on patients, caregivers and society. A common denominator for these disorders is the accumulation of aggregated proteins in nerve cells, yet the triggers for these aggregation processes are currently unknown. In ALS, protein aggregation has been described for the SOD1, C9orf72, FUS and TDP-43 proteins. The latter is a nuclear protein normally binding to both DNA and RNA, contributing to gene expression and mRNA life cycle regulation. TDP-43 seems to have a specific role in ALS pathogenesis, and ubiquitinated and hyperphosphorylated cytoplasmic inclusions of aggregated TDP-43 are present in nerve cells in almost all sporadic ALS cases. ALS pathology appears to include metal imbalances, and environmental metal exposure is a known risk factor in ALS. However, studies on metal-to-TDP-43 interactions are scarce, even though this protein seems to have the capacity to bind to metals. This review discusses the possible role of metals in TDP-43 aggregation, with respect to ALS pathology.     

Polymorphism of Alpha-Synuclein Amyloid Fibrils Depends on Ionic Strength and Protein Concentration

Protein aggregate formation is linked with multiple amyloidoses, including Alzheimer‘s and Parkinson‘s diseases. Currently, the understanding of such fibrillar structure formation and propagation is still not sufficient, the outcome of which is a lack of potent, anti-amyloid drugs. The environmental conditions used during in vitro protein aggregation assays play an important role in determining both the aggregation kinetic parameters, as well as resulting fibril structure. In the case of alpha-synuclein, ionic strength has been shown as a crucial factor in its amyloid aggregation. In this work, we examine a large sample size of alpha-synuclein aggregation reactions under thirty different ionic strength and protein concentration combinations and determine the resulting fibril structural variations using their dye-binding properties, secondary structure and morphology. We show that both ionic strength and protein concentration determine the structural variability of alpha-synuclein amyloid fibrils and that sometimes even identical conditions can result in up to four distinct types of aggregates.     

Ultra-Small ATP-Decorated Gold Nanoparticles for Targeting Amyloid Fibrils in Neurodegenerative Diseases

Ultrafine Gold nanoparticles (Au NPs) functionalized with various biomolecules constitute an alternative to antibodies as anti-amyloidogenic agents. However, generating stable ultrafine Au NPs with high surface activity is challenging. Here, the capacity of phosphate groups in biomolecules is used to stabilize Au NPs. The characteristics of Au NPs decorated with adenosine mono-, di-, and tri-phosphate are compared as well as adenosine and peptide nucleic acid-containing adenosine as controls. Among them, ATP-Au NPs are found to be superior having small size (2–4 nm) and stability (for several months) when analysed by spectroscopy and electron microscopy. Spectroscopy analysis also revealed that each ATP-stabilized Au NP is decorated with 7–8 molecules of ATP. ThT binding analysis and TEM imaging showed that the ATP-Au NPs efficiently prevented amyloid fibril formation in vitro by Aβ-42, α-Synuclein as well as by the Glucosylceramide metabolite, and disaggregated their pre-formed fibrils. NMR analysis revealed the interaction of the ATP-Au NPs with the amyloid fibrils. The ATP-Au NPs are safe toward cultured SH-SY5Y cells and when co-incubated with α-Synuclein amyloids inhibited their cytotoxicity and readily enter the cells to inhibit formation of amyloid fibrils within them. The results indicates the pharmacological potentials of ATP decorated Au NPs.     

黑果枸杞花青素对Aβ_(42)致痴呆模型大鼠记忆力及抗氧化活性研究

采用双侧海马CA1区注射Aβ42构建阿尔茨海默症(Alzheimer′s disease,AD)动物模型,通过穿梭箱被动回避记录行为学数据;以大鼠血清和脑组织中的总超氧化物歧化酶(T-SOD)、过氧化氢酶(CAT)活力及还原型谷胱甘肽(GSH)、丙二醛(MDA)和蛋白质羰基(PC)含量作为评价指标,分析黑果枸杞花青素(OPC)样品对AD模型大鼠记忆力以及体内抗氧化活性的影响。结果表明,模型组动物的记忆能力显著下降,而黑果枸杞花青素组可改善AD模型大鼠的记忆损伤;同时,发现灌胃剂量为80 mg/kg的黑果枸杞花青素组能显著提高AD大鼠血清和脑组织中T-SOD、CAT活力和GSH含量,并降低MDA和蛋白质羰基含量水平。综合上述结果可知,本研究所述黑果枸杞花青素具有良好的增强体内抗氧化活性和提高AD大鼠记忆力的作用,并具有预防AD的潜在功效。     

Synthesis and pharmacological evaluation of 8- and 9-substituted benzolactam-v8 derivatives as potent ligands for protein kinase C, a therapeutic target for Alzheimer's disease

A central element in the pathophysiology of Alzheimer's disease (AD) is the formation of amyloid plaques, which result from abnormal processing of the amyloid precursor protein (APP). The processing of APP is largely provided by three key enzymes, namely the alpha-, beta-, and gamma-secretases. As the latter two contribute to the formation of neurotoxic Abeta fragments while alpha-secretase does not, a decrease in the amyloidogenic products can be brought about either by inhibition of the beta- and gamma-secretases or through the activation of alpha-secretase. It is now known that the activation of protein kinase C (PKC) enhances alpha-secretase activity and therefore represents a possible target for the development of agents urgently needed for the treatment of this devastating neurodegenerative disorder. In the present study, new benzolactam-V8-based PKC activators were synthesized and tested for their binding affinity toward PKCalpha. All compounds tested showed binding values in the nanomolar concentration range. In accordance with previous publications, 9-substitution dramatically increased PKC binding affinity in comparison with the corresponding 8-substituted analogues. In addition to the location of the side chain on the aromatic ring, the binding affinities of these benzolactams were found to depend on the orientation, length, and electronic properties of this appendage. An interesting decrease in binding affinity was found for the 9-thienyl analogue 13, suggesting adverse electronic interactions of the sulfur atom with PKC or parts of the cellular membrane.     

Head-to-tail cyclization of a heptapeptide eliminates its cytotoxicity and significantly increases its inhibition effect on amyloid β-protein fibrillation and cytotoxicity

Amyloid-β (Aβ) protein aggregation is the main hallmark of Alzheimer’s disease (AD). Inhibition of Aβ fibrillation is thus a promising therapeutic approach to the prevention and treatment of AD. Recently, we designed a heptapeptide inhibitor, LVFFARK (LK7). LK7 shows a promising inhibitory capability on Aβ fibrillation, but is prone to self-assembling and displays high cytotoxicity, which would hinder its practical application. Herein, we modified LK7 by a head-to-tail cyclization and obtained a cyclic LK7 (cLK7). cLK7 exhibits a different self-assembly behavior from LK7, and has higher stability against proteolysis than LK7 and little cytotoxicity to SH-SY5Y cells. Thermodynamic analysis revealed that both LK7 and cLK7 could bind to Aβ₄₀ by electrostatic interactions, hydrogen bonding and hydrophobic interactions, but the binding affinity of cLK7 for Aβ₄₀ (K_D = 4.96 µmol/L) is six times higher than that of LK7 (K_D = 32.2 µmol/L). The strong binding enables cLK7 to stabilize the secondary structure of Aβ₄₀ and potently inhibit its nucleation, fibrillation and cytotoxicity at extensive concentration range, whereas LK7 could only moderately inhibit Aβ₄₀ fibrillation and cytotoxicity at low concentrations. The findings indicate that the peptide cyclization is a promising approach to enhance the performance of peptide-based amyloid inhibitors.