The lateral entorhinal cortex is a hub for local and global dysfunction in early Alzheimer's disease states.

Functional network activity alterations are one of the earliest hallmarks of Alzheimer's disease (AD), detected prior to amyloidosis and tauopathy. Better understanding the neuronal underpinnings of such network alterations could offer mechanistic insight into AD progression. Here, we examined a mouse model (3xTgAD mice) recapitulating this early AD stage. We found resting functional connectivity loss within ventral networks, including the entorhinal cortex, aligning with the spatial distribution of tauopathy reported in humans. Unexpectedly, in contrast to decreased connectivity at rest, 3xTgAD mice show enhanced fMRI signal within several projection areas following optogenetic activation of the entorhinal cortex. We corroborate this finding by demonstrating neuronal facilitation within ventral networks and synaptic hyperexcitability in projection targets. 3xTgAD mice, thus, reveal a dichotomic hypo-connected:resting versus hyper-responsive:active phenotype. This strong homotopy between the areas affected supports the translatability of this pathophysiological model to tau-related, early-AD deficits in humans.

A triple-network organization for the mouse brain.

The triple-network model of psychopathology is a framework to explain the functional and structural neuroimaging phenotypes of psychiatric and neurological disorders. It describes the interactions within and between three distributed networks: the salience, default-mode, and central executive networks. These have been associated with brain disorder traits in patients. Homologous networks have been proposed in animal models, but their integration into a triple-network organization has not yet been determined. Using resting-state datasets, we demonstrate conserved spatio-temporal properties between triple-network elements in human, macaque, and mouse. The model predictions were also shown to apply in a mouse model for depression. To validate spatial homologies, we developed a data-driven approach to convert mouse brain maps into human standard coordinates. Finally, using high-resolution viral tracers in the mouse, we refined an anatomical model for these networks and validated this using optogenetics in mice and tractography in humans. Unexpectedly, we find serotonin involvement within the salience rather than the default-mode network. Our results support the existence of a triple-network system in the mouse that shares properties with that of humans along several dimensions, including a disease condition. Finally, we demonstrate a method to humanize mouse brain networks that opens doors to fully data-driven trans-species comparisons.

Silica Nanoparticle-Enhanced Fluorescent Sensor Array for Heavy Metal Ions Detection in Colloid Solution.

Sensitivity and detection limit are two vital factors that affect fluorophores-based sensing and imaging system. However, it remains a challenge to improve the sensitivity and detection limit of fluorophores, largely due to their limited response and photophysical properties. In this study, we report for the first time, a novel approach to enhance the sensitivity and detection limit of probes using silica nanoparticles, also known as silica nanoparticles-enhanced fluorescence (SiEF). SiEF can drastically improve the fluorescence intensities and detection limit of fluorophores. A SiEF-improved fluorescent sensor array for rapid and sensitive identification of different heavy metal ions is achieved, and a 3D spatial dispersion graph is obtained based on the SiEF-improved fluorescent sensor array, which provides a lower concentration dependent pattern than fluorophores alone, allowing qualitative, quantitative, and sensitive detection of heavy metal ions. Furthermore, with UV lamp irradiation of the sensor-metal ion mixtures, the output signals enable direct visual of heavy metal ions with low concentration. Thus, the SiEF approach provides a simple and practical strategy for fluorescent probes to improve their sensitivity and detection limit in analytes sensing.

Motion-induced change in emission (MICE) for developing fluorescent probes.

The need for detecting and labelling environmentally and biologically important analytes has driven considerable research efforts in developing fluorescent probes. During the sensing process, molecular motions (i.e., molecular rotations or vibrations) of a flexible fluorescent probe can be significantly altered by its embedding micro-environment or analyte, thereby leading to substantial changes in readout signals. Motion-induced change in emission (MICE) can be utilized as an effective sensing mechanism. However, in comparison to the well-understood sensing mechanisms, such as photo-induced electron transfer (PET), intramolecular charge transfer (ICT), aggregation-induced emission (AIE) and disaggregation-induced emission (DIE), MICE has not been systematically discussed to date. In this tutorial review, we will summarize the concept and mechanisms of MICE for developing single-molecular fluorescent probes, present unique advantages of MICE based sensors, demonstrate their various applications, and discuss technical challenges in this field. We expect that this review will promote a deeper understanding of MICE and facilitate the development of novel MICE based probes.

A Simple BODIPY-Based Viscosity Probe for Imaging of Cellular Viscosity in Live Cells.

Intracellular viscosity is a fundamental physical parameter that indicates the functioning of cells. In this work, we developed a simple boron-dipyrromethene (BODIPY)-based probe, BTV, for cellular mitochondria viscosity imaging by coupling a simple BODIPY rotor with a mitochondria-targeting unit. The BTV exhibited a significant fluorescence intensity enhancement of more than 100-fold as the solvent viscosity increased. Also, the probe showed a direct linear relationship between the fluorescence lifetime and the media viscosity, which makes it possible to trace the change of the medium viscosity. Furthermore, it was demonstrated that BTV could achieve practical applicability in the monitoring of mitochondrial viscosity changes in live cells through fluorescence lifetime imaging microscopy (FLIM).

Chemical Fluorescent Probe for Detection of Aß Oligomers.

Aggregation of amyloid ß-peptide (Aß) is implicated in the pathology of Alzheimer's disease (AD), with the soluble, Aß oligomeric species thought to be the critical pathological species. Identification and characterization of intermediate species formed during the aggregation process is crucial to the understanding of the mechanisms by which oligomeric species mediate neuronal toxicity and following disease progression. Probing these species proved to be extremely challenging, as evident by the lack of reliable sensors, due to their heterogeneous and transient nature. We describe here an oligomer-specific fluorescent chemical probe, BoDipy-Oligomer (BD-Oligo), developed through the use of the diversity-oriented fluorescent library approach (DOFLA) and high-content, imaging-based screening. This probe enables dynamic oligomer monitoring during fibrillogenesis in vitro and shows in vivo Aß oligomers staining possibility in the AD mice model.

A mitochondria-targeted ratiometric fluorescent probe to monitor endogenously generated sulfur dioxide derivatives in living cells.

Sulfur dioxide (SO2) can be endogenously produced by enzymes in mitochondria during oxidation of H2S or sulphur-containing amino acids, and plays important roles in several physiological processes. However, the design and synthesis of fluorescent probes which can detect mitochondrial SO2 and its derivatives in living cells still remain unresolved. Herein, we report the preparation of a lipophilic cationic dye 1 (Mito-Ratio-SO2), which targets the mitochondria in living cells and is sensitive to the presence of SO2 derivatives. The ratiometric probe Mito-Ratio-SO2 displays a 170 nm blue-shift in emission with two well-resolved emission bands upon addition of sulfite. Mechanistic studies indicate that three probe-SO2 adducts coexist after reaction, as supported by liquid chromatography and density function theory investigations. Importantly, the ratiometric probe is highly selective for sulfite over other bio-species including H2S. Fluorescence co-localization studies indicate that the probe localizes solely in the mitochondria of HeLa cells. Last but not least, fluorescent imaging of HeLa cells successfully demonstrates the detection of intrinsically generated intracellular SO2 derivatives in living cells.

New targets of molecular imaging in atherosclerosis: prehension of current status.

Atherosclerosis is an inflammatory disease in which immune mechanisms interact with diverse metabolic risk factors to initiate, propagate and activate lesions in the arterial wall. This process starts and evolves in response to cholesterol accumulation in arterial intima of the large and medium arteries. A number of modalities for inflammatory imaging in the arterial wall are currently in the stages of pre-clinical and clinical development. However, the identification of new targets that are linked to atherosclerosis to allow for dynamic ranges of diagnosis and prognosis still remains a challenge. In this review, we summarize the current status of atherosclerosis imaging aimed at macrophage-related, direct macrophage and non-macrophage as target molecules during different stages of atherogenesis. We also present activated macrophage, biofilm and amyloid fibrils as possible new targets for the development of non-invasive imaging probes to assess atherosclerosis.

The development of a highly photostable and chemically stable zwitterionic near-infrared dye for imaging applications.

A novel zwitterionic near-infrared (NIR) dye, ZWCC, has been designed and synthesized. It shows significantly enhanced photostability and chemical stability compared to the existing zwitterionic NIR dye. In addition, the feasibility of labeling ZWCC with biological ligands was investigated and used in live cell imaging applications.

NeuO: a fluorescent chemical probe for live neuron labeling.

To address existing limitations in live neuron imaging, we have developed NeuO, a novel cell-permeable fluorescent probe with an unprecedented ability to label and image live neurons selectively over other cells in the brain. NeuO enables robust live neuron imaging and isolation in vivo and in vitro across species; its versatility and ease of use sets the basis for its development in a myriad of neuronal targeting applications.

Charge and charge-pair mutations alter the rate of assembly and structural properties of apolipoprotein C-II amyloid fibrils.

The misfolding, aggregation, and accumulation of proteins as amyloid fibrils is a defining characteristic of several debilitating diseases. Human apolipoprotein C-II (apoC-II) amyloid fibrils are representative of the fibrils formed by a number of plasma apolipoproteins implicated in amyloid-related disease. Previous structural analyses identified a buried charge pair between residues K30 and D69 within apoC-II amyloid fibrils. We have investigated the effects of amino acid substitutions of these residues on apoC-II fibril formation. Two point mutations of apoC-II, D69K and K30D, as well as a reversed ion-pair mutant containing both mutations (KDDK) were generated. Fibril formation by the double mutant, apoC-II KDDK, and apoC-II D69K was enhanced compared to that of wild-type (WT) apoC-II, while apoC-II K30D lacked the ability to form fibrils under standard conditions. Structural analyses showed that WT apoC-II, apoC-II D69K, and apoC-II KDDK fibrils have similar secondary structures and morphologies. Size distribution analyses revealed that apoC-II D69K fibrils have a broader range of fibril sizes while apoC-II KDDK fibrils showed an increased frequency of closed fibrillar loops. ApoC-II D69K fibrils exhibited reduced thioflavin T binding capacity compared to that of fibrils formed by WT apoC-II and apoC-II KDDK. These results indicate that specific charge and charge-pair mutations within apoC-II significantly alter the ability to form fibrils and that position 69 within apoC-II plays a key role in the rate-limiting step of apoC-II fibril formation.

High-efficiency in vitro and in vivo detection of Zn2+ by dye-assembled upconversion nanoparticles.

Development of highly sensitive and selective sensing systems of divalent zinc ion (Zn(2+)) in organisms has been a growing interest in the past decades owing to its pivotal role in cellular metabolism, apoptosis, and neurotransmission. Herein, we report the rational design and synthesis of a Zn(2+) fluorescent-based probe by assembling lanthanide-doped upconversion nanoparticles (UCNPs) with chromophores. Specifically, upconversion luminescence (UCL) can be effectively quenched by the chromophores on the surface of nanoparticles via a fluorescence resonant energy transfer (FRET) process and subsequently recovered upon the addition of Zn(2+), thus allowing for quantitative monitoring of Zn(2+). Importantly, the sensing system enables detection of Zn(2+) in real biological samples. We demonstrate that this chromophore-UCNP nanosystem is capable of implementing an efficient in vitro and in vivo detection of Zn(2+) in mouse brain slice with Alzheimer's disease and zebrafish, respectively.

The small molecule probe PT-Yellow labels the renal proximal tubules in zebrafish.

We report the development of a small fluorescent molecule, BDNCA3-D2, herein referred to as PT-Yellow. Soaking zebrafish embryos in PT-Yellow or intraperitoneal injection into adults results in non-toxic in vivo fluorescent labeling of the renal proximal tubules, the major site of blood filtrate reabsorption and a common target of injury in acute kidney injury. We demonstrate the applicability of this new compound as a rapid and simple readout for zebrafish kidney filtration and proximal tubule reabsorption function.

Synthesis and systematic evaluation of dark resonance energy transfer (DRET)-based library and its application in cell imaging.

In this paper, we report a new strategy for constructing a dye library with large Stokes shifts. By coupling a dark donor with BODIPY acceptors of tunable high quantum yield, a novel dark resonance energy transfer (DRET)-based library, named BNM, has been synthesized. Upon excitation of the dark donor (BDN) at 490 nm, the absorbed energy is transferred to the acceptor (BDM) with high efficiency, which was tunable in a broad range from 557 nm to 716 nm, with a high quantum yield of up to 0.8. It is noteworthy to mention that the majority of the non-radiative energy loss of the donor was converted into the acceptor's fluorescence output with a minimum leak of donor emission. Fluorescence imaging tested in live cells showed that the BNM compounds are cell-permeable and can also be employed for live-cell imaging. This is a new library which can be excited through a dark donor allowing for strong fluorescence emission in a wide range of wavelengths. Thus, the BNM library is well suited for high-throughput screening or multiplex experiments in biological applications by using a single laser excitation source.

NeuO for Neuronal Labeling in Zebrafish.

We report the application of our newly developed fluorescent probe 3-(benzylamino)-4,4-difluoro-5-(4-propyl-1H-1,2,3-triazol-1-yl)-8-(4-(2-hydroxyacetamido)phen-yl)-4-bora-3a,4a-diaza-s-indacene (NeuO) to label and image live neurons in zebrafish. Immersing zebrafish embryos in NeuO or injecting NeuO into zebrafish brain ventricles results in nontoxic in vivo neuronal labeling. We demonstrate the applicability of NeuO and envisage the potential of this compound as a rapid and simple labeling reagent for studying neuron development and degeneration.

An artificial tongue fluorescent sensor array for identification and quantitation of various heavy metal ions.

Herein, a small-molecule fluorescent sensor array for rapid identification of seven heavy metal ions was designed and synthesized, with its sensing mechanism mimicking that of a tongue. The photoinduced electron transfer and intramolecular charge transfer mechanism result in combinatorial interactions between sensor array and heavy metal ions, which lead to diversified fluorescence wavelength shifts and emission intensity changes. Upon principle component analysis (PCA), this result renders clear identification of each heavy metal ion on a 3D spatial dispersion graph. Further exploration provides a concentration-dependent pattern, allowing both qualitative and quantitative measurements of heavy metal ions. On the basis of this information, a "safe-zone" concept was proposed, which provides rapid exclusion of versatile hazardous species from clean water samples based on toxicity characteristic leaching procedure standards. This type of small-molecule fluorescent sensor array could open a new avenue for multiple heavy metal ion detection and simplified water quality analysis.

Live cells imaging using a turn-on FRET-based BODIPY probe for biothiols.

We designed a red-emitting turn-on FRET-based molecular probe 1 for selective detection of cysteine and homocysteine. Probe 1 shows significant fluorescence enhancement after cleavage of the 2, 4-dinitrobenzensulfonyl (DNBS) unit from the fluorophore upon thiols treatment. The precursor of probe 1, BNM153, is a moderate quantum yield FRET dye which contributes a minimum emission leakage from its donor part. We synthesized this assembly by connecting a low quantum yield (less than 1%) BODIPY donor to a high quantum yield BODIPY acceptor via a 1, 3-triazine bridge system. It is noteworthy that the majority of the non-radiative energy loss of donor (BDN) was converted to the acceptor (BDM)'s fluorescence output with minimum leaks of donor emission. The fluorescence sensing mechanism of probe 1 was illustrated by fluorescence spectroscopy, kinetic measurements, HPLC-MS analysis and DFT calculations. Probe 1 is pH-independent at the physiological pH range. Finally, live cells imaging demonstrated the utility of probe 1 as a biosensor for thiols.

Shear flow induced changes in apolipoprotein C-II conformation and amyloid fibril formation.

The misfolding and self-assembly of proteins into amyloid fibrils that occur in several debilitating diseases are affected by a variety of environmental factors, including mechanical factors associated with shear flow. We examined the effects of shear flow on amyloid fibril formation by human apolipoprotein C-II (apoC-II). Shear fields (150, 300, and 500 s(-1)) accelerated the rate of apoC-II fibril formation (1 mg/mL) approximately 5-10-fold. Fibrils produced at shear rates of 150 and 300 s(-1) were similar to the twisted ribbon fibrils formed in the absence of shear, while at 500 s(-1), tangled ropelike structures were observed. The mechanism of the shear-induced acceleration of amyloid fibril formation was investigated at low apoC-II concentrations (50 µg/mL) where fibril formation does not occur. Circular dichroism and tryptophan fluorescence indicated that shear induced an irreversible change in apoC-II secondary structure. Fluorescence resonance energy transfer experiments using the single tryptophan residue in apoC-II as the donor and covalently attached acceptors showed that shear flow increased the distance between the donor and acceptor molecules. Shear-induced higher-order oligomeric species were identified by sedimentation velocity experiments using fluorescence detection, while fibril seeding experiments showed that species formed during shear flow are on the fibril formation pathway. These studies suggest that physiological shear flow conditions and conditions experienced during protein manufacturing can exert significant effects on protein conformation, leading to protein misfolding, aggregation, and amyloid fibril formation.

Apolipoproteins and amyloid fibril formation in atherosclerosis.

Amyloid fibrils arise from the aggregation of misfolded proteins into highly-ordered structures. The accumulation of these fibrils along with some non-fibrillar constituents within amyloid plaques is associated with the pathogenesis of several human degenerative diseases. A number of plasma apolipoproteins, including apolipoprotein (apo) A-I, apoA-II, apoC-II and apoE are implicated in amyloid formation or influence amyloid formation by other proteins. We review present knowledge of amyloid formation by apolipoproteins in disease, with particular focus on atherosclerosis. Further insights into the molecular mechanisms underlying their amyloidogenic propensity are obtained from in vitro studies which describe factors affecting apolipoprotein amyloid fibril formation and interactions. Additionally, we outline the evidence that amyloid fibril formation by apolipoproteins might play a role in the development and progression of atherosclerosis, and highlight possible molecular mechanisms that could contribute to the pathogenesis of this disease.

High-affinity amphipathic modulators of amyloid fibril nucleation and elongation.

The misfolding and aggregation of proteins to form amyloid fibrils are associated with a number of debilitating, age-related diseases. Many of the proteins that form amyloid in vivo are lipid-binding proteins, accounting for the significant impact of lipids on the rate of formation and morphology of amyloid fibrils. To systematically investigate the effect of lipid-like compounds, we screened a range of amphipathic lipids and detergents for their effect on amyloid fibril formation by human apolipoprotein (apo) C-II. The initial screen, conducted using a set of amphiphiles at half critical micelle concentration, identified several activators and inhibitors that were selected for further analysis. Sedimentation analysis and circular dichroism studies of apoC-II at low, non-fibril-forming concentrations (0.05 mg/ml) revealed that all of the inhibitors induced the formation of apoC-II dimers enriched in α-helical content while the activators promoted the formation of stable apoC-II tetramers with increased ß-structure. Kinetic analysis identified modulators of apoC-II fibril formation that were effective at concentrations as low as 10 µM, corresponding to a modulator-to-apoC-II ratio of approximately 1:10. Delayed addition of the test compounds after fibril formation had commenced allowed the effects of selected amphiphiles on fibril elongation to be determined separately from their effects on fibril nucleation. The results indicated that specific amphiphiles induce structural changes in apoC-II that cause separate and independent effects on fibril nucleation and elongation. Low-molecular-weight amphipathic lipids and detergents may serve as useful, stage-specific modulators of protein self-assembly and fibril formation in disease-prevention strategies.