DNA nanostructures for exploring cell-cell communication.

The process of coordinating between the same or multiple types of cells to jointly execute various instructions in a controlled and carefully regulated environment is a very appealing field. In order to provide clearer insight into the role of cell-cell interactions and the cellular communication of this process in their local communities, several interdisciplinary approaches have been employed to enhance the core understanding of this phenomenon. DNA nanostructures have emerged in recent years as one of the most promising tools in exploring cell-cell communication and interactions due to their programmability and addressability. Herein, this review is dedicated to offering a new perspective on using DNA nanostructures to explore the progress of cell-cell communication. After briefly outlining the anchoring strategy of DNA nanostructures on cell membranes and the subsequent dynamic regulation of DNA nanostructures, this paper highlights the significant contribution of DNA nanostructures in monitoring cell-cell communication and regulating its interactions. Finally, we provide a quick overview of the current challenges and potential directions for the application of DNA nanostructures in cellular communication and interactions.

A novel ß-cyclodextrin-assisted enhancement strategy for portable and sensitive detection of miR-21 in human serum.

Benefiting from our discovery that ß-cyclodextrin (ß-CD) could enhance the catalytic activity of invertase through hydrogen bonding to improve detection sensitivity, a highly sensitive and convenient biosensor for the detection of miR-21 was proposed, which is based on the simplicity of reading signals from a personal glucose meter (PGM), combined with self-assembled signal amplification probes and the performance of ß-CD as an enhancer. In the presence of miR-21, magnetic nanoparticle coupled capture DNA (MNPs-cDNA) could capture it and then connect assist DNA/H1-invertase (aDNA/H1) and self-assembled signal amplification probes (H1/H2) in turn. As a result, a "super sandwich" structure was formed. The invertase on MNPs-cDNA could catalyze the hydrolysis of sucrose to glucose and this catalytic process could be enhanced by ß-CD. The PGM signal exhibited a linear correlation with miR-21 concentration within the range of 25 pmol L-1 to 3 nmol L-1, and the detection limit was as low as 5 pmol L-1 with high specificity. Moreover, the recoveries were 103.82-124.65% and RSD was 2.59-6.43%. Furthermore, the biosensor was validated for the detection of miR-21 in serum, and the results showed that miR-21 levels in serum samples from patients with Diffuse Large B-Cell Lymphoma (DLBCL) (n = 12) were significantly higher than those from healthy controls (n = 12) (P < 0.001). Therefore, the ingenious combination of PGM-based signal reading, self-assembled signal amplification probes and ß-CD as an enhancer successfully constructed a convenient, sensitive and specific biosensing method, which is expected to be applied to clinical diagnosis.

Spatial Control of Receptor Dimerization Using Programmable DNA Nanobridge.

Receptor dimerization is an essential mechanism for the activation of most receptor tyrosine kinases by ligands. Thus, regulating the nanoscale spatial distribution of cell surface receptors is significant for studying both intracellular signaling pathways and cellular behavior. However, there are currently very limited methods for exploring the effects of modulating the spatial distribution of receptors on their function by using simple tools. Herein, we developed an aptamer-based double-stranded DNA bridge acting as "DNA nanobridge", which regulates receptor dimerization by changing the number of bases. On this basis, we confirmed that the different nanoscale arrangements of the receptor can influence receptor function and its downstream signals. Among them, the effect gradually changed from helping to activate to inhibiting as the length of DNA nanobridge increased. Hence, it can not only effectively inhibit receptor function and thus affect cellular behavior but also serve as a fine-tuning tool to get the desired signal activity. Our strategy is promising to provide insight into the action of receptors in cell biology from the perspective of spatial distribution.

Activatable Fluorescence-Encoded Nanoprobes Enable Simple Multiplexed RNA Imaging in Live Cells.

Benefiting from superior programmable performance and flexible design of DNA technologies, a variety of single-molecule RNA fluorescence imaging methodologies have been reported. However, the multiplexing capability is restricted owing to the spectral overlap of fluorophores. To overcome this limitation, some inspiring multiplex imaging strategies have been developed, but in practice, it remains challenging to achieve convenient and rapid imaging in live cells due to complex designs and additional pretreatments to increase cell permeability. Here, we report an activatable fluorescence-encoded nanoprobe (AFENP) strategy, through which fluorescence-encoded functional modules for qualitative analysis and activated nucleic acid assemblies functional modules for quantitative testing enable simple multiplexed RNA imaging in single live cells. As a proof of principle, by two distinguishable fluorophores (fluorescein and rhodamine B) and their seven distinctly differentiated intensity levels, self-assembled AFENP enables simplified and quick simultaneous in situ detection and imaging of seven types of targets in live single cells because the fluorescent quantitative signal is activated only in the presence of target avoiding the washing procedures and additional pretreatment to increase cell permeability is undesired. We expect that this practical single-cell analysis platform will be adopted for multiple gene expression analysis and imaging in live cells on account of its simplicity and multiplex capability.

Biomimetic-compartmented nanoprobe for in-situ imaging of iron storage and release from ferritin in cells.

Ferritin plays an important role in regulating the homeostasis of iron in cells by storing/releasing iron. Current methods usually explored the determination of iron content, but in-situ imaging of the iron storage/release from ferritin in cells cannot be achieved. Hence, an engineered self-assembled biomimetic-compartmented nanoprobe (APO@CDs) has been constructed. The protein shell of APO (apoferritin) acted as ion channel module to control iron ions entering/exiting ferritin cavity; the inner core of CDs (carbon dots) acted as signal module for iron ions response. Compared with CDs, the response sensitivity and specificity to iron ions (Fe3+) have been improved by using APO@CDs, and the cytotoxicity was significantly reduced. Additionally, compared with cells containing APO@CDs alone, the normalized fluorescence gray value of Fe3+-treated cells was significantly decreased (0.275), indicating that Fe3+ has effectively entered the ferritin. Furtherly, that of Fe3+-treated cells incubated with deferoxamine (DFO) was significantly enhanced (0.712), showing that Fe3+ was released from ferritin under the mediation of DFO. The results demonstrate that APO@CDs can be successfully applied to in-situ imaging of iron storage/release from ferritin in cells, providing a potential platform for the in-situ dynamic study of the iron storage/release in biomedical field.

Endocytosis and intracellular RNAs imaging of nanomaterials-based fluorescence probes.

Recently, using nanomaterials to enhance the endocytosis capability and sensitivity of probes for RNA imaging in living cells has gotten the attention of many researchers. Nanomaterials, as a reliable alternative to transfection reagents, could prevent nucleic acid probes from being degraded by DNase, and bring them into sub-cellular locations for efficient internalization. Therefore, nanomaterial-based fluorescent probes (NFPs) provide a promising sensing platform to realize in situ RNA detection and imaging, which can reveal the expression of RNA at single cell level and provide large amount of information about RNA spatial localization. Meanwhile, many RNAs are in low abundance in living cells, resulting in difficulty in sensitive detection. Thus, the incorporation of NFPs and signal amplification strategy offers a broader prospect for the detection of RNAs, that have been proven as predominant therapeutic targets or diagnostic biomarkers. Herein, the purpose of our review is to first introduce the general procedure of NFPs used for in situ RNA imaging and how nanomaterials deliver these probes into living cells. Further, we focused on different kinds of nanomaterials that are mainly used for sensitive detection of RNAs and those in low abundance, through different signal read-out modes.

Construction and bioapplications of aptamer-based dual recognition strategy.

Aptamer-based dual recognition strategy, using dual aptamers or the cooperation of aptamers with other recognition elements, can better utilize the advantages of each recognition molecule and increase the design flexibility to effectively overcome the limitations of a single molecule recognition strategy, thereby improving the sensitivity and selectivity and facilitating the regulation of biological process. Hence, this review systematically tracks the construction and application of dual aptamers recognition strategy in the versatile detection of protein biomarkers, pathogenic microorganisms, cancer cells, and the treatment of some diseases and, more importantly, in functional regulation and imaging of cell-surface protein receptors. Then, the cooperation of aptamers with other recognition elements are briefly introduced. Potential challenges facing this field have been highlighted, aiming to expand bioanalytical applications of aptamer-based dual or multiple recognition strategies and meet the growing demand for precision medicine.

Interactions between CdTe quantum dots and plasma proteins: Kinetics, thermodynamics and molecular structure changes.

Environmental particulate matter, especially ultrafine particles (< 100 nm in diameter), can damage the endothelium and favor cardiovascular disease in the general population. With the wide application of nanomaterials, exposure to nanoscale particles (nanoparticles) in the environment is increasing. Systematic study of the interaction of nanoparticles with plasma proteins is critically important for understanding the cardiovascular toxicity of nanomaterials. We combined kinetics and thermodynamics information from surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) and conformational data from fluorescence spectroscopy and circular dichroism (CD) to explore the binding mechanism between cadmium telluride quantum dots (CdTe QDs) and plasma proteins. Special attention was paid to the interaction between CdTe QDs and coagulation-related proteins and the effects of CdTe QDs on protein conformation. The results showed that the binding affinities of CdTe QDs and plasma proteins depend on the nature of the protein and follow the order of fibrinogen (FIB)> plasminogen (PLG) > thrombin (TM) > metallothionein-II (MT-II) > human serum albumin (HSA). The interaction was primarily attributed to hydrophobic forces and the spontaneity of the occurrence of the interaction, and the protein secondary structures of FIB and PLG were changed significantly. The information gained in this study might shed light on the potential toxicity of QDs to the cardiovascular system.

Genome-wide Transcriptional Analysis of Oxidative Stress-related Genes and Pathways Induced by CdTe aqQDs in Mice.

Objective: Quantum dots (QDs) has widely applied in the field of science, whose potential toxic effect has increasingly become a focus concern we need pay attention to in public health. The purpose of this article was to explore the toxicity mechanism with oxidative damage from treatment with QDs at the molecular level through a gene microarray. Methods: Mice were administered aqueously synthesized cadmium telluride QDs (CdTe aqQDs) via intravenous tail injection of a 2 µmol/kg solution (based on the molar mass of Cd), and their kidneys were collected at 1 day in strict accordance with the programs used for treated mice. We determined the hierarchical clustering of expression ratios, enriched gene ontology (GO) terms and signaling pathways through gene microarray analysis and bioinformatics analysis in kidney tissue and screened the key enzyme genes, which were verified by real-time quantitative polymerase chain reaction (real-time qPCR). Results: Compared to control group, 459 lncRNAs (197 down-regulated and 262 up-regulated) and 256 mRNAs (103 down-regulated and 153 up-regulated) were differentially expressed. According to biological processes in enriched GO terms, the response to a redox state played a significant role in the biological processes involved altered genes. Pathway analysis showed that the signaling pathways that involved cytochrome P450 (CYP450) enzymes had a close relationship with QDs. Among these signaling pathways, gene expression profiling revealed that selected differentially expressed mRNAs (CYP19A1, CYP1B1, CYP11A1, CYP11B2, and CYP17A1 in the kidney and CYP19A1 and CYP1B1 in the liver) were validated by real-time qPCR, resulting in expression levels of CYP11A1, CYP11B2 and CYP17A1 in the kidney and CYP19A1 and CYP1B1 in the liver that were significantly increased, however in expression levels of CYP19A1 and CYP1B1 compared with control group in the kidney, there was no significant difference. Conclusions: Our results provide a foundation for and potential insight into the role of CYP450-related genes in QD-induced oxidative stress. QDs may produce a great deal of reactive oxygen species (ROS) by promoting high expression of CYP450 enzymes and accumulating steroid hormones, which may be an important toxicity mechanism for mediating oxidative stress and tissue damage.

A novel strategy to evaluate the degradation of quantum dots: identification and quantification of CdTe quantum dots and corresponding ionic species by CZE-ICP-MS.

In view of the significance and urgency of the speciation analysis of quantum dots (QDs) and their degradation products for clarifying their degradation rules and toxicity mechanisms, a method for the identification and quantification of CdTe QDs and corresponding ionic species in complex matrices was developed using capillary zone electrophoresis (CZE) coupled to inductively coupled plasma-mass spectrometry (ICP-MS). The quality assessment of commercial CdTe QDs and serum pharmacokinetics of synthesized CdTe QDs in rats were successfully undertaken using the developed CZE-ICP-MS method.

Magnetic immunoassay using CdSe/ZnS quantum dots as fluorescent probes to detect the level of DNA methyltransferase 1 in human serum sample.

BACKGROUND: DNA methyltransferase 1 (DNMT1), a dominant enzyme responsible for the transfer of a methyl group from the universal methyl donor to the 5-position of cytosine residues in DNA, is essential for mammalian development and closely related to cancer and a variety of age-related chronic diseases. DNMT1 has become a useful biomarker in early disease diagnosis and a potential therapeutic target in cancer therapy and drug development. However, till now, most of the studies on DNA methyltransferase (MTase) detection have focused on the prokaryote MTase and its activity. METHODS: A magnetic fluorescence-linked immunosorbent assay (FLISA) using CdSe/ZnS quantum dots as fluorescent probes was proposed for the rapid and sensitive detection of the DNMT1 level in this study. Key factors that affect the precision and accuracy of the determination of DNMT1 were optimized. RESULTS: Under the optimal conditions, the limit of detection was 0.1 ng/mL, the linear range was 0.1-1,500 ng/mL, the recovery was 91.67%-106.50%, and the relative standard deviations of intra- and inter-assays were respectively 5.45%-11.29% and 7.03%-11.25%. The cross-reactivity rates with DNA methyltransferases 3a and 3b were only 4.0% and 9.4%, respectively. Furthermore, FLISA was successfully used to detect the levels of DNMT1 in human serum samples, and compared with commercial enzyme-linked immunosorbent assay (ELISA) kits. The results revealed that there was a good correlation between FLISA and commercial ELISA kits (correlation coefficient r=0.866, p=0.001). The linear scope of FLISA was broader than ELISA, and the measurement time was much shorter than ELISA kits. CONCLUSION: These indicated that the proposed FLISA method was sensitive and high throughput and can quickly screen the level of DNMT1 in serum samples.

Dual signal amplification strategy for high-sensitivity detection of copper species in bio-samples with a tunable dynamic range.

A facile and sensitive method with a tunable dynamic range has been proposed for the detection of Cu2+ based on the self-cleavage of Cu2+-specific DNAzyme and the Cu2+-based inhibition of HRP activity, and this method was applied to evaluate the copper species in healthy people and WD patients.

Dose and time effect of CdTe quantum dots on antioxidant capacities of the liver and kidneys in mice.

Although quantum dot (QD)-induced toxicity occurs due to free radicals, generation of oxidative stress mediated by reactive oxygen species (ROS) formation is considered an important mechanism. However, free radical mechanisms are essentially difficult to elucidate at the molecular level because most biologically relevant free radicals are highly reactive and short-lived, making them difficult to directly detect, especially in vivo. Antioxidants play an important role in preventing or, in most cases, limiting the damage caused by ROS. Healthy people and animals possess many endogenous antioxidative substances that scavenge free radicals in vivo to maintain the redox balance and genome integrity. The antioxidant capacity of an organism is highly important but seldom studied. In this study, the dose and time effects of CdTe QDs on the antioxidant capacities of the liver and kidneys were investigated in mice using the electron paramagnetic resonance (EPR) spin-trapping technique. We found that the liver and kidneys of healthy mice contain specific antioxidant capacities that scavenge ·OH and ·O2-. Furthermore, oxidative stress markers (superoxide dismutase [SOD], catalase [CAT], glutathione peroxidase [GPx], glutathione [GSH] and malondialdehyde [MDA]) were examined. In dose course studies, the free radical scavenging efficiencies of the liver and kidneys were found to gradually decrease with increasing concentration of CdTe QD exposure. The activities and levels of SOD, CAT, GPx and MDA were observed to increase in treated groups, whereas those of GSH were reduced. The time course studies revealed that the QD-induced antioxidant efficiency reduction was time dependent with GSH decrease and could recover after a period of time. These experimental results offer new information on QD toxicity in vivo. Specifically, CdTe QDs can deplete GSH to reduce the elimination ability of the liver and kidneys for ·OH and ·O2-, thus inducing oxidative damage to tissues.