Amyloid ß interferes with wound healing of brain microvascular endothelial cells by disorganizing the actin cytoskeleton.

Cerebral amyloid angiopathy (CAA) is a disease in which amyloid ß (Aß) is deposited in the cerebral blood vessels, reducing compliance, tearing and weakening of vessel walls, leading to cerebral hemorrhage. The mechanisms by which Aß leads to focal wall fragmentation and intimal damage are not well understood. We analyzed the motility of human brain microvascular endothelial cells (hBMECs) in real-time using a wound-healing assay. We observed the suppression of cell migration by visualizing Aß aggregation using quantum dot (QD) nanoprobes. In addition, using QD nanoprobes and a SiR-actin probe, we simultaneously observed Aß aggregation and F-actin organization in real-time for the first time. Aß began to aggregate at the edge of endothelial cells, reducing cell motility. In addition, Aß aggregation disorganized the actin cytoskeleton and induced abnormal actin aggregation. Aß aggregated actively in the anterior group, where cell motility was active. Our findings may be a first step toward explaining the mechanism by which Aß causes vascular wall fragility, bleeding, and rebleeding in CAA.

On-Site Evaluation of Constituent Content and Functionality of Perilla frutescens var. crispa Using Fluorescence Spectra.

Perilla frutescens leaves are hypothesized to possess antioxidant and amyloid-ß (Aß) aggregation inhibitory properties primarily due to their polyphenol-type compounds. While these bioactivities fluctuate daily, the traditional methods for quantifying constituent contents and functional properties are both laborious and impractical for immediate field assessments. To address this limitation, the present study introduces an expedient approach for on-site analysis, employing fluorescence spectra obtained through excitation light irradiation of perilla leaves. Standard analytical techniques were employed to evaluate various constituent contents (chlorophyl (Chl), total polyphenol content (TPC), total flavonoid content (TFC), and rosmarinic acid (RA)) and functional attributes (DPPH radical scavenging activity, ferric reducing antioxidant power (FRAP), oxygen radical absorbance capacity (ORAC), and Aß aggregation inhibitory activity). Correlations between the fluorescence spectra and these parameters were examined using normalized difference spectral index (NDSI), ratio spectral index (RSI), and difference spectral index (DSI) analyses. The resulting predictive model exhibited a high coefficient of determination, with R2 values equal to or greater than 0.57 for constituent contents and 0.49 for functional properties. This approach facilitates the convenient, simultaneous, and nondestructive monitoring of both the chemical constituents and the functional capabilities of perilla leaves, thereby simplifying the determination of optimal harvest times. The model derived from this method holds promise for real-time assessments, indicating its potential for the simultaneous evaluation of both constituents and functionalities in perilla leaves.

Effects of three microtubule-associated proteins (MAP2, MAP4, and Tau) on microtubules' physical properties and neurite morphology.

The physical properties of cytoskeletal microtubules have a multifaceted effect on the expression of their cellular functions. A superfamily of microtubule-associated proteins, MAP2, MAP4, and tau, promote the polymerization of microtubules, stabilize the formed microtubules, and affect the physical properties of microtubules. Here, we show differences in the effects of these three MAPs on the physical properties of microtubules. When microtubule-binding domain fragments of MAP2, tau, and three MAP4 isoforms were added to microtubules in vitro and observed by fluorescence microscopy, tau-bound microtubules showed a straighter morphology than the microtubules bound by MAP2 and the three MAP4 isoforms. Flexural rigidity was evaluated by the shape of the teardrop pattern formed when microtubules were placed in a hydrodynamic flow, revealing that tau-bound microtubules were the least flexible. When full-length MAPs fused with EGFP were expressed in human neuroblastoma (SH-SY5Y) cells, the microtubules in apical regions of protrusions expressing tau were straighter than in cells expressing MAP2 and MAP4. On the other hand, the protrusions of tau-expressing cells had the fewest branches. These results suggest that the properties of microtubules, which are regulated by MAPs, contribute to the morphogenesis of neurites.

Cultivation Factors That Affect Amyloid-ß Aggregation Inhibitory Activity in Perilla frutescens var. crispa.

Alzheimer's disease (AD) is thought to be caused by the deposition of amyloid-ß (Aß) in the brain. Aß begins to aggregate approximately 20 years before the expression of its symptoms. Previously, we developed a microliter-scale high-throughput screening (MSHTS) system for inhibitors against Aß aggregation using quantum dot nanoprobes. Using this system, we also found that plants in the Lamiaceae, particularly Perilla frutescens var. crispa, have high activity. The cultivation environment has the potential to enhance Aß aggregation inhibitory activity in plants by changing their metabolism. Here, we report on cultivation factors that affected the activity of P. frutescens var. crispa cultivated in three fields under different cultivation conditions. The results revealed that the activity of P. frutescens var. crispa harvested just before flowering was highest. Interestingly, the activity of wind-shielded plants that were cultivated to prevent exposure to wind, was reduced to 1/5th of plants just before flowering. Furthermore, activity just before flowering increased following appropriate nitrogen fertilization and at least one week of drying from the day before harvest. In addition, we confirmed that the P. frutescens var. crispa leaf extracts suppressed Aß-induced toxicity in nerve growth factor-differentiated PC12 cells. In this study, we demonstrated that flowering, wind, soil water content, and soil nitrogen content affected Aß aggregation inhibitory activity, necessary to suppress Aß neurotoxicity, in P. frutescens var. crispa extracts. This study provides practical cultivation methods for P. frutescens var. crispa with high Aß aggregation inhibitory activity for the prevention of AD.

Towards a comprehensive approach for characterizing cell activity in bright-field microscopic images.

When studying physical cellular response observed by light microscopy, variations in cell behavior are difficult to quantitatively measure and are often only discussed on a subjective level. Hence, cell properties are described qualitatively based on a researcher's impressions. In this study, we aim to define a comprehensive approach to estimate the physical cell activity based on migration and morphology based on statistical analysis of a cell population within a predefined field of view and timespan. We present quantitative measurements of the influence of drugs such as cytochalasin D and taxol on human neuroblastoma, SH-SY5Y cell populations. Both chemicals are well known to interact with the cytoskeleton and affect the cell morphology and motility. Being able to compute the physical properties of each cell for a given observation time, requires precise localization of each cell even when in an adhesive state, where cells are not visually differentiable. Also, the risk of confusion through contaminants is desired to be minimized. In relation to the cell detection process, we have developed a customized encoder-decoder based deep learning cell detection and tracking procedure. Further, we discuss the accuracy of our approach to quantify cell activity and its viability in regard to the cell detection accuracy.

Kinetics of amyloid accumulation in physiological viscosity.

Abnormal aggregation and accumulation of misfolded proteins are involved in the development of various forms of amyloidosis. Aggregates that accumulate in organs induce an inflammatory response and cytotoxicity, and lead to organ failure. Although protein accumulation around an affected area in the body is an important stage that is directly linked to the mechanism of pathogenesis, the kinetics of the accumulation of protein that precipitates while assembling is not well understood because 3D tracking of proteins in solution is difficult. Here, we analyzed the process of aggregation and accumulation of amyloid ß (Aß), which causes the development of Alzheimer's disease (AD), by real-time 3D imaging under physiological conditions using a quantum dot nanoprobe that we previously developed. 3D observations demonstrated that Aß aggregates with a diameter of several µm emerged in phosphate-buffered saline, gathered in a spiral-like step, and exhibited a mesh-like structure. Additionally, we found that the amount and size of aggregates decreased dramatically in 40% glycerol solution, mimicking the viscosity of human blood. We confirmed that fibrils in 40% glycerol exhibited an extremely short and tangled morphology and formed dense aggregates. Furthermore, numerical calculations revealed that several decades are required to fully develop the settling velocity and diameter of Aß aggregates in physiological conditions. This time span is consistent with the actual symptom progression of AD.

Amyloid ß aggregation induces human brain microvascular endothelial cell death with abnormal actin organization.

Cerebral amyloid angiopathy (CAA) is a disease in which amyloid ß (Aß) is deposited on the walls of blood vessels in the brain, making those walls brittle and causing cerebral hemorrhage. However, the mechanism underlying its onset is not well understood. The aggregation and accumulation of Aß cause the occlusion and fragility of blood vessels due to endothelial cell damage, breakdown of the blood-brain barrier, and replacement with elements constituting the blood vessel wall. In this study, we observed the effect of Aß on human primary brain microvascular endothelial cells (hBMECs) in real-time using quantum dot nanoprobes to elucidate the mechanism of vascular weakening by Aß. It was observed that Aß began to aggregate around hBMECs after the start of incubation and that the cells were covered with aggregates. Aß aggregates firmly anchored the cells on the plate surface, and eventually suppressed cell motility and caused cell death. Furthermore, Aß aggregation induced the organization of abnormal actin, resulting in a significant increase in intracellular actin dots over 10 µm2. These results suggest that the mechanism by which Aß forms a fragile vessel wall is as follows: Aß aggregation around vascular endothelial cells anchors them to the substrate, induces abnormal actin organization, and leads to cell death.

Farm use of calcium hydroxide as an effective barrier against pathogens.

Livestock farming is affected by the occurrence of infectious diseases, but outbreaks can be prevented by proper sanitary control measures. Calcium hydroxide (Ca(OH)2), commonly called slaked lime, powder is traditionally used as a disinfectant to prevent infectious diseases in livestock. Since Ca(OH)2 can inactivate a wide variety of pathogens, has a small environmental impact, does not require a disinfection tank (i.e., can be spread directly on the ground) and is produced inexpensively worldwide, it is used for the prevention of epidemics on farms worldwide. Water is essential for the strong alkalinity that underlies its disinfecting effect, but it is unknown how much water is required under field conditions. In addition, Ca(OH)2 reacts with carbon dioxide in the environment, reducing its pH, but it is unclear how long its degradation takes under actual field use. Thus, we measured the water adsorption ability of Ca(OH)2-based disinfectants and its relation to disinfectant activity, as assessed by colony counts and live/dead staining and observation. We found that 15-20% (w/w) water in Ca(OH)2 was necessary for disinfection to occur in practice. Moreover, we found that the pH of Ca(OH)2 decreased within about two weeks to one month under actual use in practical conditions and lost its ability to disinfect. We further showed that granules prepared from Ca(OH)2 and zeolite maintained high alkalinity more than twice as long as calcium powder. These findings will help to establish a suitable method of applying Ca(OH)2 to protect farms from infectious diseases.

Mechanosensitive myosin II but not cofilin primarily contributes to cyclic cell stretch-induced selective disassembly of actin stress fibers.

Cells adapt to applied cyclic stretch (CS) to circumvent chronic activation of proinflammatory signaling. Currently, the molecular mechanism of the selective disassembly of actin stress fibers (SFs) in the stretch direction, which occurs at the early stage of the cellular response to CS, remains controversial. Here, we suggest that the mechanosensitive behavior of myosin II, a major cross-linker of SFs, primarily contributes to the directional disassembly of the actomyosin complex SFs in bovine vascular smooth muscle cells and human U2OS osteosarcoma cells. First, we identified that CS with a shortening phase that exceeds in speed the inherent contractile rate of individual SFs leads to the disassembly. To understand the biological basis, we investigated the effect of expressing myosin regulatory light-chain mutants and found that SFs with less actomyosin activities disassemble more promptly upon CS. We consequently created a minimal mathematical model that recapitulates the salient features of the direction-selective and threshold-triggered disassembly of SFs to show that disassembly or, more specifically, unbundling of the actomyosin bundle SFs is enhanced with sufficiently fast cell shortening. We further demonstrated that similar disassembly of SFs is inducible in the presence of an active LIM-kinase-1 mutant that deactivates cofilin, suggesting that cofilin is dispensable as opposed to a previously proposed mechanism.

Unidirectional cooperative binding of fimbrin actin-binding domain 2 to actin filament.

Fimbrin forms bundles of parallel actin filaments in filopodia, but it remains unclear how fimbrin forms well-ordered bundles. To address this issue, we focused on the cooperative interaction between the actin-binding domain of fimbrin and actin filaments. First, we loosely immobilized actin filaments on a glass surface via a positively charged lipid layer and observed the binding of GFP-fused actin-binding domain 2 of fimbrin using fluorescence microscopy. The actin-binding domain formed low-density clusters with unidirectional growth along actin filaments. When the actin filaments were tightly immobilized to the surface by increasing the charge density of the lipid layer, cluster formation was suppressed. This result suggests that the propagation of cooperative structural changes of actin filaments evoked by binding of the actin-binding domain was suppressed by a strong physical interaction with the glass surface. Interestingly, binding of the fimbrin actin-binding domain shortened the length of loosely immobilized actin filaments. Based on these results, we propose that fimbrin-actin interactions accompanied by unidirectional long-range allostery help the formation of well-ordered parallel actin filament bundles.

Aggregation of Mouse Serum Amyloid A Protein Was Promoted by Amyloid-Enhancing Factors with the More Genetically Homologous Serum Amyloid A.

Amyloid A (AA) amyloidosis is a condition in which amyloid fibrils characterized by a linear morphology and a cross-ß structure accumulate and are deposited extracellularly in organs, resulting in chronic inflammatory diseases and infections. The incidence of AA amyloidosis is high in humans and several animal species. Serum amyloid A (SAA) is one of the most important precursor amyloid proteins and plays a vital step in AA amyloidosis. Amyloid enhancing factor (AEF) serves as a seed for fibril formation and shortens the onset of AA amyloidosis sharply. In this study, we examined whether AEFs extracted and purified from five animal species (camel, cat, cattle, goat, and mouse) could promote mouse SAA (mSAA) protein aggregation in vitro using quantum-dot (QD) nanoprobes to visualize the aggregation. The results showed that AEFs shortened and promoted mSAA aggregation. In addition, mouse and cat AEFs showed higher mSAA aggregation-promoting activity than the camel, cattle, and goat AEFs. Interestingly, homology analysis of SAA in these five animal species revealed a more similar amino acid sequence homology between mouse and cat than between other animal species. Furthermore, a detailed comparison of amino acid sequences suggested that it was important to mSAA aggregation-promoting activity that the 48th amino acid was a basic residue (Lys) and the 125th amino acid was an acidic residue (Asp or Glu). These data imply that AA amyloidosis exhibits higher transmission activity among animals carrying genetically homologous SAA gene, and may provide a new understanding of the pathogenesis of amyloidosis.

Comparison of AA Amyloid Fibril Morphology and Serum Amyloid A Gene Sequence in 5 Animal Species.

Amyloid fibrils are characterized by a linear morphology and a cross-ß structure. Polymorphic and multiple fibril morphologies can be found when amyloid fibrils are extracted from amyloid-laden tissue. In this study, we report on the purification and transmission electron microscopic analysis of amyloid fibrils from 5 different animal species (mouse, cow, goat, dog, and camel) with AA amyloidosis. The results show that amyloid fibrils had a linear morphology with a cross-structure and irregular length in vivo. Although the fibrils from these different species showed highly similar conformations, there were significant differences in fibril width and crossover distance. We analyzed the sequences of homologous amyloid proteins and serum amyloid A, an evolutionarily conserved protein and a major amyloid precursor. We found 78.23% homology between the most distant amyloid proteins. The findings suggested similar fibril width and crossover distance in different animal species that displayed high homology of amyloid protein sequences. Dog and camel, as well as goat and cow, showed high genetic homology and similar fibril morphology. These data indicate that the fibrils from different animal species have similar genetic homology and morphology, which may provide a better understanding of the pathogenesis of amyloidosis.

What factors determine the number of nonmuscle myosin II in the sarcomeric unit of stress fibers?

Actin stress fibers (SFs), a contractile apparatus in nonmuscle cells, possess a contractile unit that is apparently similar to the sarcomere of myofibrils in muscles. The function of SFs has thus often been addressed based on well-characterized properties of muscles. However, unlike the fixed number of myosin molecules in myofibrils, the number of nonmuscle myosin II (NMII) within the contractile sarcomeric unit in SFs is quite low and variable for some reason. Here we address what factors may determine the specific number of NMII in SFs. We suggest with a theoretical model that the number lies just in between the function of SFs for bearing cellular tension under static conditions and for promptly disintegrating upon forced cell shortening. We monitored shortening-induced disintegration of SFs in human osteosarcoma U2OS cells expressing mutants of myosin regulatory light chain that virtually regulates the interaction of NMII with actin filaments, and the behaviors observed were indeed consistent with the theoretical consequences. This situation-specific nature of SFs may allow nonmuscle cells to respond adaptively to mechanical stress to circumvent activation of pro-inflammatory signals as previously indicated, i.e., a behavior distinct from that of muscles that are basically specialized for exhibiting contractile activity.

Daurichromenic Acid from the Chinese Traditional Medicinal Plant Rhododendron dauricum Inhibits Sphingomyelin Synthase and Aß Aggregation.

Species of the genus Rhododendron have been used in traditional Chinese medicine, with the medicinal herb "Manshanfong" used as an expectorant and for the treatment of acute bronchitis. Daurichromenic acid (DCA), a constituent of Rhododendron dauricum, is a meroterpenoid with antibacterial, anti-HIV, and anti-inflammatory activities. However, the mechanisms underlying these pharmacologic activities are poorly understood. To develop new drugs based on DCA, more information is required regarding its interactions with biomolecules. The present study showed that DCA inhibits the activity of the enzyme sphingomyelin synthase, with an IC50 of 4 µM. The structure-activity relationships between DCA and sphingomyelin synthase were evaluated using derivatives and cyclized hongoquercin A. In addition, DCA was found to inhibit amyloid ß aggregation. These results may help in the design of effective drugs based on DCA.

Dietary Intake of Rosmarinic Acid Increases Serum Inhibitory Activity in Amyloid A Aggregation and Suppresses Deposition in the Organs of Mice.

Serum amyloid A (SAA) is one of the most important precursor amyloid proteins and plays a vital step in AA amyloidosis, although the underlying aggregation mechanism has not been elucidated. Since SAA aggregation is a key step in this pathogenesis, inhibitors are useful to prevent and treat AA amyloidosis, serving as tools to investigate the pathogenic mechanism. In this study, we showed that rosmarinic acid (RA), which is a well-known inhibitor of the aggregation of amyloid ß (Aß), displayed inhibitory activity against SAA aggregation in vitro using a microliter-scale high-throughput screening (MSHTS) system with quantum-dot nanoprobes. Therefore, we evaluated the amyloid aggregation inhibitory activity of blood and the deposition of SAA in organs by feeding mice with Melissa officinalis extract (ME) containing RA as an active substance. Interestingly, the inhibitory activity of ME-fed mice sera for SAA and Aß aggregation, measured with the MSHTS system, was higher than that of the control group. The amount of amyloid deposition in the organs of ME-fed mice was lower than that in the control group, suggesting that the SAA aggregation inhibitory activity of serum is associated with SAA deposition. These results suggest that dietary intake of RA-containing ME enhanced amyloid aggregation inhibitory activity of blood and suppressed SAA deposition in organs. This study also demonstrated that the MSHTS system could be applied to in vitro screening and to monitor comprehensive activity of metabolized foods adsorbed by blood.

Evaluation of Amyloid ß42 Aggregation Inhibitory Activity of Commercial Dressings by A Microliter-Scale High-Throughput Screening System Using Quantum-Dot Nanoprobes.

The aggregation and accumulation of amyloid ß (Aß) in the brain is a trigger of pathogenesis for Alzheimer's disease. Previously, we developed a microliter-scale high-throughput screening (MSHTS) system for Aß42 aggregation inhibitors using quantum-dot nanoprobes. The MSHTS system is seldom influenced by contaminants in samples and is able to directly evaluate Aß42 aggregation inhibitory activity of samples containing various compounds. In this study, to elucidate whether the MSHTS system could be applied to the evaluation of processed foods, we examined Aß42 aggregation inhibitory activity of salad dressings, including soy sauces. We estimated the 50% effective concentration (EC50) from serial diluted dressings. Interestingly, all 19 commercial dressings tested showed Aß42 aggregation inhibitory activity. It was suggested that EC50 differed by as much as 100 times between the dressings with the most (0.065 ± 0.020 v/v%) and least (6.737 ± 5.054 v/v%) inhibitory activity. The highest activity sample is traditional Japanese dressing, soy sauce. It is known that soy sauce is roughly classified into a heat-treated variety and a non-heat-treated variety. We demonstrated that non-heat-treated raw soy sauce exhibited higher Aß42 aggregation inhibitory activity than heat-treated soy sauce. Herein, we propose that MSHTS system can be applied to processed foods.

Three-dimensional real time imaging of amyloid ß aggregation on living cells.

Alzheimer's disease (AD) is a progressive disorder of the brain that gradually decreases thinking, memory, and language abilities. The aggregation process of amyloid ß (Aß) is a key step in the expression of its neurocytotoxicity and development of AD because Aß aggregation and accumulation around neuronal cells induces cell death. However, the molecular mechanism underlying the neurocytotoxicity and cell death by Aß aggregation has not been clearly elucidated. In this study, we successfully visualized real-time process of Aß42 aggregation around living cells by applying our established QD imaging method. 3D observations using confocal laser microscopy revealed that Aß42 preferentially started to aggregate at the region where membrane protrusions frequently formed. Furthermore, we found that inhibition of actin polymerization using cytochalasin D reduced aggregation of Aß42 on the cell surface. These results indicate that actin polymerization-dependent cell motility is responsible for the promotion of Aß42 aggregation at the cell periphery. 3D observation also revealed that the aggregates around the cell remained in that location even if cell death occurred, implying that amyloid plaques found in the AD brain grew from the debris of dead cells that accumulated Aß42 aggregates.

Long-Range and Directional Allostery of Actin Filaments Plays Important Roles in Various Cellular Activities.

A wide variety of uniquely localized actin-binding proteins (ABPs) are involved in various cellular activities, such as cytokinesis, migration, adhesion, morphogenesis, and intracellular transport. In a micrometer-scale space such as the inside of cells, protein molecules diffuse throughout the cell interior within seconds. In this condition, how can ABPs selectively bind to particular actin filaments when there is an abundance of actin filaments in the cytoplasm? In recent years, several ABPs have been reported to induce cooperative conformational changes to actin filaments allowing structural changes to propagate along the filament cables uni- or bidirectionally, thereby regulating the subsequent binding of ABPs. Such propagation of ABP-induced cooperative conformational changes in actin filaments may be advantageous for the elaborate regulation of cellular activities driven by actin-based machineries in the intracellular space, which is dominated by diffusion. In this review, we focus on long-range allosteric regulation driven by cooperative conformational changes of actin filaments that are evoked by binding of ABPs, and discuss roles of allostery of actin filaments in narrow intracellular spaces.

Microtubule elongation along actin filaments induced by microtubule-associated protein 4 contributes to the formation of cellular protrusions.

Actin-microtubule crosstalk is implicated in the formation of cellular protrusions, but the mechanism remains unclear. In this study, we examined the regulation of cell protrusion involving a ubiquitously expressed microtubule-associated protein (MAP) 4, and its superfamily proteins, neuronal MAP2 and tau. Fluorescence microscopy revealed that these MAPs bound to F-actin and microtubules simultaneously, and formed F-actin/microtubule hybrid bundles. The hybrid bundle-forming activity was in the order of MAP2 > MAP4 ≫ tau. Interestingly, the microtubule assembly-promoting activity of MAP4 and MAP2, but not of tau, was upregulated by their interaction with F-actin. When MAP4 was overexpressed in NG108-15 cells, the number of cell processes and maximum process length of each cell increased significantly by 28% and 30%, respectively. Super-resolution microscopy revealed that 95% of microtubules in cell processes colocalized with F-actin, and MAP4 was always found in their vicinity. These results suggest that microtubule elongation along F-actin induced by MAP4 contributes to the formation of cellular protrusions. Since MAP4, MAP2 and tau had different crosstalk activity between F-actin and microtubules, it is likely that the functional differentiation of these MAPs is a driving force for neural evolution, causing significant changes in cell morphology.

Real-Time 3D Imaging and Inhibition Analysis of Various Amyloid Aggregations Using Quantum Dots.

Amyloidosis refers to aggregates of protein that accumulate and are deposited as amyloid fibrils into plaques. When these are detected in organs, they are the main hallmark of Alzheimer's disease, Parkinson's disease, and other related diseases. Recent medical advances have shown that many precursors and proteins can induce amyloidosis even though the mechanism of amyloid aggregation and the relationship of these proteins to amyloidosis remains mostly unclear. In this study, we report the real-time 3D-imaging and inhibition analysis of amyloid ß (Aß), tau, and α-synuclein aggregation utilizing the affinity between quantum dots (QD) and amyloid aggregates. We successfully visualized these amyloid aggregations in real-time using fluorescence microscopy and confocal microscopy simply by adding commercially available QD. The observation by transmission electron microscopy (TEM) showed that QD particles bound to all amyloid fibrils. The 3D-imaging with QD revealed differences between amyloid aggregates composed of different amyloid peptides that could not be detected by TEM. We were also able to quantify the inhibition activities of these proteins by rosmarinic acid, which has high activity for Aß aggregation, from fluorescence micrographs as half-maximal effective concentrations. These imaging techniques with QD serve as quick, easy, and powerful tools to understand amyloidosis and to discover drugs for therapies.