Molecular recognition of live methicillin-resistant staphylococcus aureus cells using DNA aptamers

AIM: To generate DNA-aptamers binding to Methicillinresistant Staphylococcus aureus(MRSA).METHODS: The Cell-Systematic Evolution of Ligands by Exponential Enrichment(SELEX) technology was used to run the selection against MRSA bacteria and develop target-specific aptamers. MRSA bacteria were targeted while Enterococcus faecalis bacteria were used for counter selection during that process. Binding assays to determine the right aptamer candidates as well as binding assays on clinical samples were performed through flow cytometry and analyzed using the FlowJ o software. The characterization of the aptamers was done by determination of their Kd values and determined by analysis of flow data at different aptamer concentration using Sigma Plot. Finally, the recognitionof the complex Gold-nanoparticle-aptamer to the bacteria cells was observed using transmission electron microscopy(TEM).RESULTS: During the cell-SELEX selection process, 17 rounds were necessary to generate enrichment of the pool. While the selection was run using fixed cells, it was shown that the binding of the pools with live cells was giving similar results. After sequencing and analysis of the two last pools, four sequences were identified to be aptamer candidates. The characterization of those aptamers showed that based on their Kd values, DTMRSA4 presented the best binding with a Kd value of 94.61 ± 18.82 nmol/L. A total of ten clinical samples of MRSA, S. aureus and Enterococcus faecalis were obtained to test those aptamers and determine their binding on a panel of samples. DTMRSA1 and DTMRSA3 showed the best results regarding their specificity to MRSA, DTMRSA1 being the most specific of all. Finally, those aptamers were coupled with gold-nanoparticle and their binding to MRSA cells was visualized through TEM showing that adduction of nanoparticles on the aptamers did not change their binding property.CONCLUSION: A total of four aptamers that bind to MRSA were obtained with Kd values ranking from 94 to 200 nmol/L.

Metal-Organic Framework Nanocarriers for Drug Delivery in Biomedical Applications

Investigation of metal–organic frameworks(MOFs)for biomedical applications has attracted much attention in recent years.MOFs are regarded as a promising class of nanocarriers for drug delivery owing to well-defined structure,ultrahigh surface area and porosity,tunable pore size,and easy chemical functionalization.In this review,the unique properties of MOFs and their advantages as nanocarriers for drug delivery in biomedical applications were discussed in the first section.Then,state-ofthe-art strategies to functionalize MOFs with therapeutic agents were summarized,including surface adsorption,pore encapsulation,covalent binding,and functional molecules as building blocks.In the third section,the most recent biological applications of MOFs for intracellular delivery of drugs,proteins,and nucleic acids,especially aptamers,were presented.Finally,challenges and prospects were comprehensively discussed to provide context for future development of MOFs as efficient drug delivery systems.

A process insight into production of ethyl levulinate via a stepwise fractionation

Ethyl levulinate(EL)is a key biomass-derived compounds due to its socio-economic benefits for the synthesis of commodity chemicals.Herein,we proposed an efficient one-step bamboo conversion to EL in ethanol,and a novel stepwise fractionation to purify EL and lignocellulose degradation products.A proton acid,due to its high catalytic efficiency,yielded 26.65%EL in 120 min at 200℃.The productions of ethyl glucoside and 5-ethoxymethylfurfural were analyzed in terms of by-products formation.To the best of our knowledge,there is no single report on catalyst for one step synthesis of EL directly from bamboo,as well as a stepwise fractionation to purify EL.Due to similar physiochemical properties in each fraction,the platform molecules could broaden a new paradigm of bamboo biomass utilization for renewable energy and value-added biochemicals.In addition,glucose,ethyl glucoside,corn starch,and microcrystalline cellulose were also investigated as substrates,so that the reaction intermediates of this one-pot procedure were identified and a possible reaction mechanism was proposed.

Aptamer-Based Cell-Surface Profiling with Single-Cell Resolution Enables Precise Cancer Characterization

Molecular profiling of cell-surface proteins is a powerful strategy for precise cancer diagnosis.While mass cytometry(MC)enables synchronous detection of over 40 cellular parameters,its full potential in disease classification is challenged by the limited types of recognition probes currently available.In this work,we synthesize a panel of heavy isotopeconjugated aptamers to profile cancer-associated signatures on the surface of hematological malignancy(HM)cells.Based on 15 molecular signatures,we performed cell-surface profiling that allowed the precise classification of 8 HM cell lines.Combined with machine-learning technology,this aptamer-based MC platform also achieved multiclass identification of HM subtypes in clinical sampleswith 100%accuracy in the training cohort and 80%accuracy in the test cohort.Therefore,we report an effective and practical strategy for precise cancer classification at the singlecell level,paving the way for its clinical use in the near future.

Self-assembled multifunctional DNA nanoflowers for the circumvention of multidrug resistance in targeted anticancer drug delivery

癌症化疗被它的副作用和 multidrug 抵抗(MDR ) 限制了。因此，指向了能围绕 MDR 的药交货系统被需要。这里，我们向 chemosensitive 和在白血病和乳癌房间模型围绕了 MDR 的 MDR 癌症房间为指向的药交货汇报多功能的 DNA nanoflowers (NF ) 。NF 经由卷圆复制， NF 为特定的癌症房间识别，为 bioimaging 的 fluorophores，和 doxorubicin (纪录影片) 用 aptamers 在期间被合并产生的 DNA 和镁焦磷酸盐的潜在的一起沉淀是自我装配的为药交货的有约束力的 DNA。NF 尺寸是悦耳的(在直径击倒 nm 到 200 ) ，并且浓密地包装的药绑定主题和多孔的 intrastructures 赋予 NF 以一个高装载药的能力(71.4% ， wt/wt ) 。尽管 Doxloaded NF (NF 纪录影片) 在生理的 pH 是稳定的，药版本在酸或基本的条件下面被便于。NF 交付纪录影片进目标 chemosensitive 和 MDR 癌症房间，阻止药流出并且提高在 MDR 房间的药保留。NF 纪录影片在两个目标 chemosensitive 房间和 MDR 房间，然而并非在 nontarget 房间导致有势力 cytotoxicity，并发地因此围绕 MDR 并且减少副作用。总的来说，这些 NF 为在指向的癌症治疗围绕 MDR 正在答应工具。

Zinc-substituted hemoglobin with specific drug binding sites and fatty acid resistance ability for enhanced photodynamic therapy

Precisely designed protein-based nanodrugs, as a kind of colloidal drug system, have attracted significant attention in tumor therapy because of their refined drug loading ratio, controlled delivery efficacy and natural biocompatibility. However, most drugs are conjugated to the protein carriers randomly without specific binding sites. Moreover, such sites could easily be replaced by lipophilic molecules in the physiological environment and result in low delivery efficiency. With strong and specific binding locations especially comparatively narrow spatial binding sites and nonflexible structure, hemin (FePPIX)-free hemoglobin or apohemoglobin (apoHb), as a natural metalloporphyrin protein carrier, represents great potential in bioapplication. Therefore, we herein introduce a folate acid (FA) modified, zinc-substituted hemoglobin (ZnPHb-FA) as a naturally occurring protein matrix-based photosensitizer for cancer photodynamic therapy (PDT). Noncovalent inserted ZnPPIX molecules in apoHb possess an extremely stable property and significant recovered photoproperties with superior biocompatibility and phototoxicity, both in vitro and in vivo. This stability was verified by molecular docking analysis and calculation of binding constant, representing a total of five drug binding sites of apoHb for ZnPPIX molecules, four of which are energetically favorable (△G value of -11.9 kcal/mol), and one which is energetically acceptable (△G value of -9 kcal/mol). Folate acid modification has been shown to efficiently enhance the internalization and retention time of ZnPHb nanodrug. ZnPHb-FA is also an efficient depressor of hemin oxygenase-1 (HO-1), which could, in turn, lower the antioxidant ability of cancer cells by decreasing the production of biiirublin. Results in vitro and in vivo both indicated that the firmly combination of apoHb and ZnPPIX described here represents a novel and efficient protein nanodrug systems for cancer therapy.

Nanocarrier based on the assembly of protein and antisense oligonucleotide to combat multidrug resistance in tumor cells

Chemotherapy-induced multi-drug resistance(MDR) in tumors poses a huge challenge for clinical treatment of tumors. The downregulation of the multi-drug resistance relative protein, represented by P-glycoprotein(P-gp), can reverse MDR of cancer cells. In this study, we developed doxorubicin-loading nanocarrier based on the assembly of protein and antisense oligonucleotide(ASO) to combat MDR of cancer cells. The data demonstrate that the nanocarrier can efficiently deliver ASO to cytoplasm and downregulate the P-glycoprotein expression, subsequently improving the therapeutic effects of Dox in doxorubicin-resistant MCF-7/ADR cancer cells. The preparation is simple and effective, providing a powerful tool for gene delivery. Therefore, our nanocarrier shows high promise in cancer treatment.

Stable and unique graphitic Raman internal standard nanocapsules for surface-enhanced Raman spectroscopy quantitative analysis

Graphitic nanomaterials have unique, strong, and stable Raman vibrations that have been widely applied in chemistry and biomedicine. However, utilizing them as internal standards （ISs） to improve the accuracy of surface-enhanced Raman spectroscopy （SERS） analysis has not been attempted. Herein, we report the design of a unique IS nanostructure consisting of a large number of gold nanoparticles （AuNPs） decorated on multilayered graphitic magnetic nanocapsules （AGNs） to quantify the analyte and eliminate the problems associated with traditional ISs. The AGNs demonstrated a unique Raman band from the graphitic component, which was localized in the Raman silent region of the biomolecules, making them an ideal IS for quantitative Raman analysis without any background interference. The IS signal from the AGNs also indicated superior stability, even under harsh conditions. With the enhancement of the decorated AuNPs, the AGN nanostructures greatly improved the quantitative accuracy of SERS, in particular the exclusion of quantitative errors resulting from collection loss and non-uniform distribution of the analytes. The AGNs were further utilized for cell staining and Raman imaging, and they showed great promise for applications in biomedicine.

Recent progress of aptamer-drug conjugates in cancer therapy

Aptamers are single-stranded DNA or RNA sequences that can specifically bind with the target protein or molecule via specific secondary structures.Compared to antibody-drug conjugates(ADC),aptamer-drug conjugate(ApDC)is also an efficient,targeted drug for cancer therapy with a smaller size,higher chemical stability,lower immunogenicity,faster tissue penetration,and facile engineering.Despite all these advantages,several key factors have delayed the clinical translation of ApDC,such as in vivo off-target effects and potential safety issues.In this review,we highlight the most recent progress in the development of ApDC and discuss solutions to the problems noted above.

Molecular imaging:design mechanism and bioapplications

Molecular imaging is a non-invasive method to image and analyze the concentration and activity of functional biomolecules in cells or in vivo at molecular level,and plays an increasing role in deep understanding of biological processes,early and accurate diagnosis of diseases,and evaluation of treatment.Nowadays,numerous novel molecular imaging probes have been developed,involving every biomedical imaging modality,such as optical imaging,photoacoustic imaging,magnetic resonance imaging,single-photon-emission computed tomography,and positron emission tomography.In this review,we summarize the development of current state-of-the-art molecular imaging probes.We introduce the design strategies of molecular probes and detailed imaging modalities,and highlight the properties of probes and biomedical imaging applications in cells and in vivo,including disease diagnosis,drug tracking,and imaging-guided surgery.Then we discuss the perspectives and challenges in this emerging field.We expect this review could inspire more effective molecular imaging probes to be developed,achieving the goal towards clinical practices.

Gold nanoparticle-decorated MoSe_(2) nanosheets as highly effective peroxidase-like nanozymes for total antioxidant capacity assay

A visual colorimetric detection strategy is reported for total antioxidant capacity(TAC)assay by using 3,3',5,5'-tetramethylbenzidine(TMB)oxidation as chromogenic substrate based on gold nanoparticle-decorated MoSe_(2) nanosheets(Au@MoSe_(2)).Au@MoSe_(2) nanostructures exhibit high peroxidase-like activity and can catalyze H_(2)O_(2)to oxidize TMB.Based on inhibition effect of ascorbic acid(AA)on TMB oxidation,a facile and sensitive colorimetric method was developed for AA detection.Under optimal conditions,the proposed method showed a sensitivity for AA in a concentration range from 2 to 120μM and limit of detection was 0.41μM.Furthermore,the method was employed for TAC assay in actual samples,including commercial beverages and vitamin C tablets.This work represents a model in nanostructure design and will lead to further development of TAC assay in evaluation of antioxidant food quality.

Activatable and Self-Monitoring Hydrogen Sulfide-Based Molecular Senomorphics for Visualized Regulation of Cellular Senescence

Specific regulation of the senescence-associated secretory phenotype(SASP)is vital to block senescence-induced detrimental cellular plasticity.Recently,some chemical compounds called senomorphics have demonstrated such potential,but it remains challenging to achieve site-specific activation and real-time monitoring of the action of senomorphics,posing great obstacles for transformable applications.Here,we report a tailor-made hydrogen sulfide(H_(2)S)donor(Lyso-FH_(2)S-Gal)as a new class of molecule senomorphics for spatially controlled delivery of H_(2)S for visualization of regulation of cellular senescence.It comprises four functional moieties in a single molecular structure,including a lysosome-targeting group for cell recognition,a lysosomal enzyme-cleaved scaffold for site-specific activation,thiocarbamate as the H_(2)S precursor,and a switchable fluorophore for concurrent selfreporting of H_(2)S release and senescence imaging.Lyso-FH_(2)S-Gal exhibited remarkable response selectivity,sustained H_(2)S release,and 141-fold fluorescence enhancement.In cellular models,Lyso-FH_(2) S-Gal preferentially enriched in senescent cells over nonsenescent cells,and alleviated the levels of SASP and reactive oxygen species(ROS)in senescent cells,while remaining inert in nonsenescent cells.More impressively,it efficiently inhibited the SASPmediated crosstalk between senescent cells and surrounding nonsenescent cells,thereby preventing senescence propagation.This work offers a useful molecular tool with the hope for controlled intervention of senescence-related important biological processes.

Precise nanomedicine for intelligent therapy of cancer

Precise nanomedicine has been extensively explored for efficient cancer imaging and targeted cancer therapy, as evidenced by a few breakthroughs in their preclinical and clinical explorations. Here, we demonstrate the recent advances of intelligent cancer nanomedicine, and discuss the comprehensive understanding of their structure-function relationship for smart and efficient cancer nanomedicine including various imaging and therapeutic applications, as well as nanotoxicity. In particular, a few emerging strategies that have advanced cancer nanomedicine are also highlighted as the emerging focus such as tumor imprisonment, supramolecular chemotherapy, and DNA nanorobot. The challenge and outlook of some scientific and engineering issues are also discussed in future development. We wish to highlight these new progress of precise nanomedicine with the ultimate goal to inspire more successful explorations of intelligent nanoparticles for future clinical translations.

Recent progress in engineering near-infrared persistent luminescence nanoprobes for time-resolved biosensing/bioimaging

Persistent luminescence nanoprobes (PLNPs) can remain luminescent after ceasing excitation.Due to the ultra-long decay time of persistent luminescence (PersL),autofluorescence interference can be efficiently eliminated by collecting PersL signal after autofluorescence decays completely,thus the imaging contrast and sensing sensitivity can be significantly improved.Since near-infrared (NIR) light shows reduced scattering and absorption coefficient in penetrating biological organs or tissues,near-infrared persistent luminescence nanoprobes (NIR PLNPs) possess deep tissue penetration and offer a bright prospect in the areas of in vivo biosensing/bioimaging.In this review,we firstly summarize the design of different types of NIR PLNPs for biosensing/bioimaging,such as transition metal ions-doped NIR PLNPs,lanthanide ions-doped NIR PLNPs,organic molecules-based NIR PLNPs,and semiconducting polymer self-assembled NIR PLNPs.Notably,organic molecules-based NIR PLNPs and semiconductor self-assembled NIR PLNPs,for the first time,were introduced to the review of PLNPs.Secondly,the effects of different types of charge carriers on NIR PersL and luminescence decay of NIR PLNPs are significantly emphasized so as to build up an in-depth understanding of their luminescence mechanism.It includes the regulation of valence band and conduction band of different host materials,alteration of defect types,depth and concentration changes caused by ion doping,effective radiation transitions and energy transfer generated by different luminescence centers.Given the design and potential of NIR PLNPs as long-lived luminescent materials,the current challenges and future perspective in this rapidly growing field are also discussed.

Gold nanorod-photosensitizer conjugate with extracellular pH-driven tumor targeting ability for photothermal/photodynamic therapy

氯 e6-pHLIP <候补选手class=“ a-plus-plus ”> ss -AuNRs,金使nanorod具有感光性结合包含 pH (低)有给予细胞外的 pH 的一张二硫化物契约的插入肽( pHLIP )( pH <候补选手class=“ a-plus-plus ”> e )-driven 肿瘤指向能力，成功地为光力学的 bimodal 和 photothermal 治疗被开发了。在这 bimodal 治疗，氯 e6 (Ce6 ) ，第二代的 photosensitizer (PS ) ，被用于光力学的治疗(太平洋夏季时间) 。金 nanorods (AuNRs ) 作为 Ce6 的 nanocarrier 和 quencher 为 photothermal 治疗(PTT ) 并且也被用作一个过高热代理人。pHLIPss 作为一根驾驶 pHe 的指向探针被设计在低 pH 在癌症房间提高 Ce6 和 AuNRs 的累积。在 Ce6-pHLIP < 潜水艇 class= “ a-plus-plus ” > ss -AuNRs, Ce6 接近并且由 AuNRs 熄灭了，引起小太平洋夏季时间效果。当暴露了到正常生理的 pH 时 7.4， Ce6-pHLIP < 潜水艇 class= “ a-plus-plus ” > ss -AuNRs 泛泛地与房间膜联系。然而，曾经暴露了到酸的 pH 6.2， pHLIP 活跃地插入到房间膜，和 conjugates 是进房间的 translocated。当这发生时， Ce6 由于二硫化物与 AuNRs 分开细胞内部的谷胱甘肽(GSH ) 引起的契约劈开，和汗衫氧在光之上为太平洋夏季时间被生产照耀。作为单个 PTT 代理人，另外， AuNRs 能由提高的浸透和保留(EPR ) 效果在肿瘤提高 PS 的累积。因此由我们的数据显示了当暴露了到酸的 pH 时， Ce6-pHLIP < 潜水艇 class= “ a-plus-plus ” > ss -AuNRs 能为癌症治疗完成 synergistic PTT/PDT bimodality。

Bioconjugated Silica Nanoparticles: Development and Applications

Advanced bioanalysis,including accurate quantitation,has driven the need to understand biology and medicine at the molecular level.Bioconjugated silica nanoparticles have the potential to address this emerging challenge.Particularly intriguing diagnostic and therapeutic applications in cancer and infectious disease as well as uses in gene and drug delivery,have also been found for silica nanoparticles.In this review,we describe the synthesis,bioconjugation,and applications of silica nanoparticles in different bioanalysis formats,such as selective tagging,barcoding,and separation of a wide range of biomedically important targets.Overall,we envisage that further development of these nanoparticles will provide a variety of advanced tools for molecular biology,genomics,proteomics and medicine.

A novel fluorescent label based on biological fluores-cent nanoparticles and its application in cell recognition

Uniform-sized fluorescent nanoparticles have been prepared by employing silica as the shell and a highly luminescent dye complex of ruthenium ion and bipyridyl, tris(2,2 ’-bipyridyl) dichlororuthenium( II) hexahydrate as the core of the nanoparticles. A novel fluorescent label method is proposed, which is based on the biological fluorescent nanoparticles on the foundation of nanotechnology, biotechnology and fluorescent label technology. In comparison with the conventional fluorophores as fluorescent labels such as fluorescein isothiocyanate (FITC) label, this new label shows more superiority in photochemical stability, detection sensitivity and application scope for the biomedicine research. SmIgG+ B lymphocytes isolated from the circulating blood of human beings can be easily recognized by using this new fluorescent label.

Fabrication of superstable gold nanorod-carbon nanocapsule as a molecule loading material

In this work, we fabricated a monodisperse nanocomposite by coating gold nanorods(Au NRs) with a layer of biocompatible, stable carbon, obtaining Au NR@Carbon core–shell nanocapsules, which without any functionalization could be used as a molecule loading material due to its high surface areas. In this system, the Au NR core had a high-absorption cross section for conversion of near-infrared light to heat, which could be explored for local hyperthermia. The carbon shell, which was biocompatible and stable even under concentrated acidic and alkaline conditions, was able to adsorb molecules with p–p interactions or electrostatic interactions. In comparison with Au NR@Si O2, Au NR@Carbon nanocapsules demonstrate the following merits:(1) simple and green synthesis method,(2) far more stable with respect to high-temperature stability and(3) larger molecule loading capacity,which indicate great potential in the biomedical applications.

Highly Fluorescent Dye-Doped Silica Nanoparticles Increase Flow Cytometry Sensitivity for Cancer Cell Monitoring

Early and accurate diagnosis and treatment of cancer depend on rapid,sensitive,and selective detection of tumor cells.Current diagnosis of cancers,especially leukemia,relies on histology and fl ow cytometry using single dye-labeled antibodies.However,this combination may not lead to high signal output,which can hinder detection,especially when the probes have relatively weak affi nities or when the receptor is expressed in a low concentration on the target cell surface.To solve these problems,we have developed a novel method for sensitive and rapid detection of cancer cells using dye-doped silica nanoparticles(NPs)which increases detection sensitivity in fl ow cytometry analyses between 10-and 100-fold compared to standard methods.Our NPs are~60 nm in size and can encapsulate thousands of individual dye molecules within their matrix.We have extensively investigated surface modifi cation strategies in order to make the NPs suitable for selective detection of cancer cells using fl ow cytometry.The NPs are functionalized with polyethylene glycol(PEG)to prevent nonspecifi c interactions and with neutravidin to allow universal binding with biotinylated molecules.By virtue of their reliable and selective detection of target cancer cells,these NPs have demonstrated their promising usefulness in conventional fl ow cytometry.Moreover,they have shown low background signal,high signal enhancement,and effi cient functionalization,either with antibody-or aptamer-targeting moieties.

The influence of physiological environment on the targeting effect of aptamer-guided gold nanoparticles

Aptamer guided nano medicine shows great promise in targeted cancer therapies.However the loss of targeting capacity duri ng in vivo or clinical trials has largely hindered its popularity and there are no systematic studies to elucidate the causes.Herein,we investigated such loss of targeting capacity by examining how the physiological milieu affected targeting effect.Aptamer functionalized gold nanoparticle(AuNP)was chosen as the model and exposed to human blood serum that is used to mimic physiological milieu.Dynamic light scattering(DLS),flow cytometry and label-free liquid chromatography tan dem mass spectrometry(LC-MS/MS)were employed to determi ne variations of NPs'surface chemistry and biological identities changes after serum exposure.Results showed that the targeting ability loss was caused by protein corona blocking,replacement and enzymatic cleavage of surface aptamer targeting ligands.Noteworthy,the aggregation issue is critical for the smaller NPs.Analysis of the protein corona profile indicated the accumulation of immune-related proteins on the surface of aptamer-conjugated NPs,which could induce immune response,resulting in rapid clearanee of NPs.