Macrophage-targeted nanomedicine for the diagnosis and management of atherosclerosis.

Atherosclerosis is the primary cause of cardiovascular diseases, such as myocardial infarction and stroke, which account for the highest death toll worldwide. Macrophage is the major contributor to atherosclerosis progression, and therefore, macrophage-associated pathological process is considered an extremely important target for the diagnosis and treatment of atherosclerosis. However, the existing clinical strategies still have many bottlenecks and challenges in atherosclerosis's early detection and management. Nanomedicine, using various nanoparticles/nanocarriers for medical purposes, can effectively load therapeutic agents, significantly improve their stability and accurately deliver them to the atherosclerotic plaques. In this review, we summarized the latest progress of the macrophage-targeted nanomedicine in the diagnosis and treatment of atherosclerosis, and their potential applications and clinical benefits are also discussed.

High Resolution of Plasmonic Resonance Scattering Imaging with Deep Learning.

The dark-field microscopy (DFM) imaging technology has the advantage of a high signal-to-noise ratio, and it is often used for real-time monitoring of plasmonic resonance scattering and biological imaging at the single-nanoparticle level. Due to the limitation of the optical diffraction limit, it is still a challenging task to accurately distinguish two or more nanoparticles whose distance is less than the diffraction limit. Here, we propose a computational strategy based on a deep learning framework (NanoNet), which will realize the effective segmentation of the scattered light spots in diffraction-limited DFM images and obtain high-resolution plasmonic light scattering imaging. A small data set of DFM and the corresponding scanning electron microscopy (SEM) image pairs are used to learn for obtaining a highly resolved semantic imaging model using NanoNet, and thus highly resolved DFM images matching the resolution of those acquired using SEM can be obtained. Our method has the ability to transform diffraction-limited DFM images to highly resolved ones without adding a complex optical system. As a proof of concept, a highly resolved DFM image of living cells through the NanoNet technique is successfully made, opening up a new avenue for high-resolution optical nanoscopic imaging.

The aluminum distribution and translocation in two citrus species differing in aluminum tolerance.

BACKGROUND: Many citrus orchards of south China suffer from soil acidification, which induces aluminum (Al) toxicity. The Al-immobilization in vivo is crucial for Al detoxification. However, the distribution and translocation of excess Al in citrus species are not well understood. RESULTS: The seedlings of 'Xuegan' [Citrus sinensis (L.) Osbeck] and 'Shatianyou' [Citrus grandis (L.) Osbeck], that differ in Al tolerance, were hydroponically treated with a nutrient solution (Control) or supplemented by 1.0 mM Al3+ (Al toxicity) for 21 days after three months of pre-culture. The Al distribution at the tissue level of citrus species followed the order: lateral roots > primary roots > leaves > stems. The concentration of Al extracted from the cell wall (CW) of lateral roots was found to be about 8 to 10 times higher than in the lateral roots under Al toxicity, suggesting that the CW was the primary Al-binding site at the subcellular level. Furthermore, the Al distribution in CW components of the lateral roots showed that pectin had the highest affinity for binding Al. The relative expression level of genes directly relevant to Al transport indicated a dominant role of Cs6g03670.1 and Cg1g021320.1 in the Al distribution of two citrus species. Compared to C. grandis, C. sinensis had a significantly higher Al concentration on the CW of lateral roots, whereas remarkably lower Al levels in the leaves and stems. Furthermore, Al translocation revealed by the absorption kinetics of the CW demonstrated that C. sinensis had a higher Al retention and stronger Al affinity on the root CW than C. grandis. According to the FTIR (Fourier transform infrared spectroscopy) analysis, the Al distribution and translocation might be affected by a modification in the structure and components of the citrus lateral root CW. CONCLUSIONS: A higher Al-retention, mainly attributable to pectin of the root CW, and a lower Al translocation efficiency from roots to shoots contributed to a higher Al tolerance of C. sinensis than C. grandis. The aluminum distribution and translocation of two citrus species differing in aluminum tolerance were associated with the transcriptional regulation of genes related to Al transport and the structural modification of root CW.

Transformable Helical Self-Assembly for Cancerous Golgi Apparatus Disruption.

Golgi apparatus is a major subcellular organelle responsible for drug resistance. Golgi apparatus-targeted nanomechanical disruption provides an attractive approach for killing cancer cells by multimodal mechanism and avoiding drug resistance. Inspired by the poisonous twisted fibrils in Alzheimer's brain tissue and enhanced rigidity of helical structure in nature, we designed transformable peptide C6RVRRF4KY that can self-assemble into nontoxic nanoparticles in aqueous medium but transformed into left-handed helical fibrils (L-HFs) after targeting and furin cleavage in the Golgi apparatus of cancer cells. The L-HFs can mechanically disrupt the Golgi apparatus membrane, resulting in inhibition of cytokine secretion, collapse of the cellular structure, and eventually death of cancer cells. Repeated stimulation of the cancers by the precursors causes no acquired drug resistance, showing that mechanical disruption of subcellular organelle is an excellent strategy for cancer therapy without drug resistance. This nanomechanical disruption concept should also be applicable to multidrug-resistant bacteria and viruses.

Automated Plasmonic Resonance Scattering Imaging Analysis via Deep Learning.

Plasmonic nanoparticles, which have excellent local surface plasmon resonance (LSPR) optical and chemical properties, have been widely used in biology, chemistry, and photonics. The single-particle light scattering dark-field microscopy (DFM) imaging technique based on a color-coded analytical method is a promising approach for high-throughput plasmonic nanoparticle scatterometry. Due to the interference of high noise levels, accurately extracting real scattering light of plasmonic nanoparticles in living cells is still a challenging task, which hinders its application for intracellular analysis. Herein, we propose an automatic and high-throughput LSPR scatterometry technique using a U-Net convolutional deep learning neural network. We use the deep neural networks to recognize the scattering light of nanoparticles from background interference signals in living cells, which have a dynamic and complicated environment, and construct a DFM image semantic analytical model based on the U-Net convolutional neural network. Compared with traditional methods, this method can achieve higher accuracy, stronger generalization ability, and robustness. As a proof of concept, the change of intracellular cytochrome c in MCF-7 cells under UV light-induced apoptosis was monitored through the fast and high-throughput analysis of the plasmonic nanoparticle scattering light, providing a new strategy for scatterometry study and imaging analysis in chemistry.

Infusion of blood from mice displaying cerebral amyloidosis accelerates amyloid pathology in animal models of Alzheimer's disease.

Previous studies showed that injection of tissue extracts containing amyloid-ß (Aß) aggregates accelerate amyloid deposition in the brain of mouse models of Alzheimer's disease (AD) through prion-like mechanisms. In this study, we evaluated whether brain amyloidosis could be accelerated by blood infusions, procedures that have been shown to transmit prion diseases in animals and humans. Young transgenic mice infused with whole blood or plasma from old animals with extensive Aß deposition in their brains developed significantly higher levels brain amyloidosis and neuroinflammation compared to untreated animals or mice infused with wild type blood. Similarly, intra-venous injection of purified Aß aggregates accelerated amyloid pathology, supporting the concept that Aß seeds present in blood can reach the brain to promote neuropathological alterations in the brain of treated animals. However, an amyloid-enhancing effect of other factors present in the blood of donors cannot be discarded. Our results may help to understand the role of peripheral (amyloid-dependent or -independent) factors implicated in the development of AD and uncover new strategies for disease intervention.

Multifunctional aggregation-based fluorescent probe for visualizing intracellular calcium dynamic fluctuations.

Calcium ion (Ca2+) is an indispensable second messenger in living organisms. The impaired Ca2+ handling can induce many diseases. In this paper, we developed a simple and effective method to encapsulate a coumarin-based Ca2+ probe ((E)-2-hydroxy-N'-((7-hydroxy-2-oxo-2H-chromen-8-yl)methylene)-2-phenylacetohydrazide, CPM) into nanoparticles (NPs), and CPM NPs with blue fluorescence were obtained, whose maximum excitation and maximum emission wavelengths were characterized at 365 nm and 450 nm, respectively. The CPM NPs show significant fluorescence enhancement toward Ca2+ over other metal ions, with a limit of determination (LOD) of 0.04 µM. To optimize the optical property of the NPs, CPM and curcumin, which were introduced as the Förster resonance energy transfer (FRET) donor and acceptor, respectively, were co-encapsulated, and bright green CPM@Cur NPs with large stokes shift and narrow emission band width were constructed. Due to their low cytotoxicity and excellent stability, CPM NPs and CPM@Cur NPs were further successfully used to discriminate the primary aortic smooth muscle cells isolated from mice with abnormal Ca2+ homeostasis from their littermate controls. It is worth noting that CPM@Cur NPs exhibit stronger fluorescence signal and diminished background interference, which make them have great potential in the Ca2+ monitoring during biological processes. This strategy opens a new way to synthesize NPs with high brightness and has a potential application prospect in composite sensing and intracellular imaging. CPM@Cur NPs are developed and applied in biological sensing and intracellular Ca2+ imaging, as well as discriminating the cells with abnormal calcium homeostasis.

Multifunctional Single-Layered Graphene Quantum Dots Used for Diagnosis of Mitochondrial Malfunction-Related Diseases.

Mitochondria are critical organelles in eukaryotes that efficiently generate adenosine 5'-triphosphate (ATP) for various biological activities, and any defect in the process of ATP synthesis may lead to mitochondrial dysfunction and directly link to a variety of medical disorders. Monitoring the ATP variations in cells is key for innovative early diagnosis of mitochondrial diseases. Herein, multifunctional single-layered graphene quantum dots (s-GQDs) with bright green emission were constructed, which exhibit strong binding affinity for ATP and good mitochondria targeting ability. Using the proposed s-GQDs, we successfully discriminated the primary smooth muscle cells isolated from the transgenic mouse (heterozygote sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) 2 C674S knock-in mouse) with mitochondrial disorders or their littermate controls, indicating s-GQDs as promising probes for the study of cell metabolism and mitochondrial malfunction-related diseases, and targeting endoplasmic reticulum stress is an effective way to modulate metabolic pathways relevant to SERCA 2 inactivity mitochondrial dysfunction.

The role of lyases, NblA and NblB proteins and bilin chromophore transfer in restructuring the cyanobacterial light-harvesting complex‡.

Phycobilisomes are large light-harvesting complexes attached to the stromal side of thylakoids in cyanobacteria and red algae. They can be remodeled or degraded in response to changing light and nutritional status. Both the core and the peripheral rods of phycobilisomes contain biliproteins. During biliprotein biosynthesis, open-chain tetrapyrrole chromophores are attached covalently to the apoproteins by dedicated lyases. Another set of non-bleaching (Nb) proteins has been implicated in phycobilisome degradation, among them NblA and NblB. We report in vitro experiments with lyases, biliproteins and NblA/B which imply that the situation is more complex than currently discussed: lyases can also detach the chromophores and NblA and NblB can modulate lyase-catalyzed binding and detachment of chromophores in a complex fashion. We show: (i) NblA and NblB can interfere with chromophorylation as well as chromophore detachment of phycobiliprotein, they are generally inhibitors but in some cases enhance the reaction; (ii) NblA and NblB promote dissociation of whole phycobilisomes, cores and, in particular, allophycocyanin trimers; (iii) while NblA and NblB do not interact with each other, both interact with lyases, apo- and holo-biliproteins; (iv) they promote synergistically the lyase-catalyzed chromophorylation of the ß-subunit of the major rod component, CPC; and (v) they modulate lyase-catalyzed and lyase-independent chromophore transfers among biliproteins, with the core protein, ApcF, the rod protein, CpcA, and sensory biliproteins (phytochromes, cyanobacteriochromes) acting as potential traps. The results indicate that NblA/B can cooperate with lyases in remodeling the phycobilisomes to balance the metabolic requirements of acclimating their light-harvesting capacity without straining the overall metabolic economy of the cell.

Size dependent anti-invasiveness of silver nanoparticles in lung cancer cells.

The molecular mechanism of cancer cell death caused by silver nanoparticles (AgNPs) of different sizes is investigated. Compared with the larger nanoparticles, 13 nm AgNPs significantly inhibit the migration and invasiveness of lung adenocarcinoma A549 cells, induce elevated reactive oxygen species and lead to NF-κB directed cellular apoptosis.

The Red-/Green-Switching GAF3 of Cyanobacteriochrome Slr1393 from Synechocystis sp. PCC6803 Regulates the Activity of an Adenylyl Cyclase.

Cyanobacteriochromes (CBCRs) are photoreceptors in cyanobacteria that present a bilin chromophore-binding GAF domain as a photochromic element to control the activity of a downstream enzyme or regulator. CBCR Slr1393 from Synechocystis PCC 6803 carries three GAF domains, but only the third one binds phycocyanobilin covalently. Slr1393 shows photochromicity between red and green absorbing states and regulates a C-terminally located histidine kinase. In this work, we fused this third GAF domain to an adenylyl cyclase (AC) from Microcoleus chthonoplastes PCC7420 that in its genuine form is under blue-light control from a LOV domain. A series of RGS-AC variants were constructed with various lengths of the linkers between RGS and AC. Assays in vitro and in living Escherichia coli cells (AC-deletion mutant) demonstrated that the activity of AC was light regulated, namely, the red-light-converted form of RGSΔ14-Δ4AC (in vitro) was about three times more active than the green-light-converted form. Expression of the fusion protein RGSΔ14-Δ4AC in vivo again showed highest light regulation with at least threefold amplification of the AC function. In some experiments, even tenfold higher activity was observed, which indicated that the protein, if expressed under in vivo conditions, was part of the E. coli physiological conditions and thereby subjected to more complex and variable regulation through other E. coli inherent factors.

Altered sarco(endo)plasmic reticulum calcium adenosine triphosphatase 2a content: Targets for heart failure therapy.

Sarco(endo)plasmic reticulum calcium adenosine triphosphatase is responsible for transporting cytosolic calcium into the sarcoplasmic reticulum and endoplasmic reticulum to maintain calcium homeostasis. Sarco(endo)plasmic reticulum calcium adenosine triphosphatase is the dominant isoform expressed in cardiac tissue, which is regulated by endogenous protein inhibitors, post-translational modifications, hormones as well as microRNAs. Dysfunction of sarco(endo)plasmic reticulum calcium adenosine triphosphatase is associated with heart failure, which makes sarco(endo)plasmic reticulum calcium adenosine triphosphatase a promising target for heart failure therapy. This review summarizes current approaches to ameliorate sarco(endo)plasmic reticulum calcium adenosine triphosphatase function and focuses on phospholamban, an endogenous inhibitor of sarco(endo)plasmic reticulum calcium adenosine triphosphatase, pharmacological tools and gene therapies.

Prognostic significance of long non-coding RNA MALAT1 for predicting the recurrence and metastasis of gallbladder cancer and its effect on cell proliferation, migration, invasion, and apoptosis.

The objective of this study is to explore the role of MALAT1 as a molecular indicator in predicting the recurrence, metastasis, and prognosis of gallbladder cancer (GBC) and its effect on the proliferation, invasion, migration, and apoptosis of GBC cells in vitro. GBC tissues and adjacent normal tissues were collected from 102 patients. MALAT1 short hairpin RNA (shRNA) plasmids were first constructed to transfect the GBC-SD cells. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) was applied to detect MALAT1 expression. CCK-8 assay, flow cytometry, and Transwell assay were applied to testify the cell proliferation, cell cycle, apoptosis, invasion, and migration. A receiver operating characteristic (ROC) curve was used to evaluate the values of MALAT1 in GBC recurrence, metastasis, and prognosis. COX regression analysis was applied to analyze the independent influencing factors of GBC patients' survival status. ROC curve results showed that the MALAT1 expression could be a predictor of the GBC recurrence, metastasis, and prognosis. According to the COX regression analysis, MALAT1 expression, tumor size, and TNM stage were independent influencing factors of GBC patients' survival condition. Compared with the GBC-SD cells transfected with empty plasmids, those transfected with MALAT1 shRNA plasmids showed higher apoptosis rates, weakened proliferation, migration, and invasion. In conclusion, our findings demonstrate that lncRNA MALAT1 can be considered as an indicator for evaluating the recurrence, metastasis, and prognosis of GBC patients. We also demonstrate how the overexpression of MALAT1 confers an oncogenic function in GBC.

Role of Prion Replication in the Strain-dependent Brain Regional Distribution of Prions.

One intriguing feature of prion diseases is their strain variation. Prion strains are differentiated by the clinical consequences they generate in the host, their biochemical properties, and their potential to infect other animal species. The selective targeting of these agents to specific brain structures have been extensively used to characterize prion strains. However, the molecular basis dictating strain-specific neurotropism are still elusive. In this study, isolated brain structures from animals infected with four hamster prion strains (HY, DY, 139H, and SSLOW) were analyzed for their content of protease-resistant PrP(Sc) Our data show that these strains have different profiles of PrP deposition along the brain. These patterns of accumulation, which were independent of regional PrP(C) production, were not reproduced by in vitro replication when different brain regions were used as substrate for the misfolding-amplification reaction. On the contrary, our results show that in vitro replication efficiency depended exclusively on the amount of PrP(C) present in each part of the brain. Our results suggest that the variable regional distribution of PrP(Sc) in distinct strains is not determined by differences on prion formation, but on other factors or cellular pathways. Our findings may contribute to understand the molecular mechanisms of prion pathogenesis and strain diversity.

Highly Photoluminescent Molybdenum Oxide Quantum Dots: One-Pot Synthesis and Application in 2,4,6-Trinitrotoluene Determination.

As a well-studied transition-metal semiconductor material, MoOx has a wider band gap than molybdenum disulfide (MoS2), and its property varies dramatically for the existence of several different allotropes and suboxide phases of molybdenum oxides (MoOx, x < 3). In this manuscript, a one-pot method possessing the advantages of one pot, easily prepared, rapid, and environmentally friendly, has been developed for facile synthesis of highly photoluminescent MoOx quantum dots (MoOx QDs), in which commercial molybdenum disulfide (MoS2) powder and hydrogen peroxide (H2O2) are employed as the precursor and oxidant, respectively. The obtained MoOx QDs can be further utilized as an efficient photoluminescent probe, and a new turn-off sensor is developed for 2,4,6-trinitrotoluene (TNT) determination based on the fact that the photoluminescence of MoOx QDs can be quenched by the Meisenheimer complexes formed in the strong alkali solution through the inner filter effect (IFE). Under the optimal conditions, the decreased photoluminescence of MoOx QDs shows a good linear relationship to the concentration of TNT ranging from 0.5 to 240.0 µM, and the limit of detection was 0.12 µM (3σ/k). With the present turn-off sensor, TNT in river water samples can be rapidly and selectively detected without tedious sample pretreatment processes.

Strain-dependent profile of misfolded prion protein aggregates.

Prions are composed of the misfolded prion protein (PrP(Sc)) organized in a variety of aggregates. An important question in the prion field has been to determine the identity of functional PrP(Sc) aggregates. In this study, we used equilibrium sedimentation in sucrose density gradients to separate PrP(Sc) aggregates from three hamster prion strains (Hyper, Drowsy, SSLOW) subjected to minimal manipulations. We show that PrP(Sc) aggregates distribute in a wide range of arrangements and the relative proportion of each species depends on the prion strain. We observed a direct correlation between the density of the predominant PrP(Sc) aggregates and the incubation periods for the strains studied. The relative presence of PrP(Sc) in fractions of different sucrose densities was indicative of the protein deposits present in the brain as analyzed by histology. Interestingly, no association was found between sensitivity to proteolytic degradation and aggregation profiles. Therefore, the organization of PrP molecules in terms of the density of aggregates generated may determine some of the particular strain properties, whereas others are independent from it. Our findings may contribute to understand the mechanisms of strain variation and the role of PrP(Sc) aggregates in prion-induced neurodegeneration.

Nanosilver-based surface-enhanced Raman spectroscopic determination of DNA methyltransferase activity through real-time hybridization chain reaction.

In this manuscript, a nanosilver enhanced SERS strategy was successfully constructed for the determination of DNA methyltransferase activity in soulution combined with hybridization chain reaction (HCR). The proposed method was mainly on the basis of excellent separation ability of magnetic microparticles (MMPs), HCR as signal amplification unit and assembled AgNPs as enhancement substrate. In the presence of M. SssI MTase, the duplex sequence (5'-CCGG-3') tethered to MMPs was methylated, which cannot be cleaved by HpaII endonuclease. The resulted DNA skeleton captured on MMPs then triggered the HCR reaction, generated a polymerized and extended symmetrical sequence, in which more biotin terminal was available for the conjugation of AgNPs-SA, leading to significantly amplified SERS response. When it was used to analyze M. SssI activity, a linear equation ∆ISERS=1215.32+446.80 cM.SssI was obtained with the M. SssI activity ranged from 0.1 to 10.0 U with the correlation coefficient (r(2)) of 0.97. The most important advantage of this method is the combination of SERS and HCR in solution for the first time and its good selectivity, which enabled the detection of even one-base mismatched sequence. The new assay method holds great promising application to be a versatile platform for sensitive, high-throughput detection, and the screening of new anticancer drugs on DNA MTase.

Coomassie brilliant blue R-250 as a new surface-enhanced Raman scattering probe for prion protein through a dual-aptamer mechanism.

Surface-enhanced Raman scattering (SERS) spectra, which can provide large information about trace amount of chemical and biological species have been widely performed as a well-established tool in complex biological system. In this work, coomassie brilliant blue (R-250) with high affinity to proteins and high Raman activity was employed as a Raman reporter to probe prion protein (PrP) through a dual-aptamer mechanism, and thus an original strategy for PrP determination was proposed, which showed great potential to turn on the SERS response through specific recognition of anti-prion aptamers towards the target protein. Aptamers (Apt1 and Apt 2) recognizing distinct epitopes of PrP with high affinity were first conjugated to Ag@Si NPs, and Ag@Si-PrP/R-250-Ag@Si conjugates were obtained in the presence of PrP/R-250, inducing dramatically enhanced Raman signal. SERS responses enhanced with increasing amount of PrP and a linear equation of ISERS=6729.7+3091.2 cPrP was obtained in the range of 3.0-12.0×10(-9)M with the determination coefficient of 0.988. The proposed strategy is simple, rapid, and high specificity to probe protein-aptamer recognition in the solution.

Aggregate-depleted brain fails to induce Aß deposition in a mouse model of Alzheimer's disease.

Recent studies in animal models of Alzheimer's disease (AD) show that amyloid-beta (Aß) misfolding can be transmissible; however, the mechanisms by which this process occurs have not been fully explored. The goal of this study was to analyze whether depletion of aggregates from an AD brain suppresses its in vivo "seeding" capability. Removal of aggregates was performed by using the Aggregate Specific Reagent 1 (ASR1) compound which has been previously described to specifically bind misfolded species. Our results show that pre-treatment with ASR1-coupled magnetic beads reduces the in vivo misfolding inducing capability of an AD brain extract. These findings shed light respect to the active principle responsible for the prion-like spreading of Alzheimer's amyloid pathology and open the possibility of using seeds-capturing reagents as a promising target for AD treatment.

Brains from non-Alzheimer's individuals containing amyloid deposits accelerate Aß deposition in vivo.

BACKGROUND: One of the main features of Alzheimer's disease (AD) is the presence of Aß deposits, which accumulate in the brain years before the onset of symptoms. We and others have demonstrated that cerebral Aß-amyloidosis can be induced in vivo by administration of AD-brain extracts into transgenic mice. However, it is currently unknown whether amyloid formation can be induced using extracts from individuals harboring Aß deposits, but not clinical disease. RESULTS: In this study we analyzed the amyloid-inducing capability of samples from individuals affected by mild cognitive impairment (MCI) and Non-Demented persons with Alzheimer's disease Neuropathology (NDAN). Our results show that inoculation of transgenic mice with MCI and NDAN brain samples accelerated Aß pathology in a similar way as extracts from confirmed AD. CONCLUSIONS: This data demonstrate that the sole presence of Aß aggregates in a given sample, regardless of the clinical condition, is capable to accelerate Aß deposition in vivo. These findings indicate that the amyloid-inducing activity may be present in the brain of people during pre-symptomatic or a-symptomatic stages of AD.