Near-infrared fluorophores with absolute aggregation-caused quenching and negligible fluorescence re-illumination for in vivo bioimaging of nanocarriers

Environment-responsive fluorophores with aggregation-caused quenching(ACQ)properties have been applied to track nanocarriers with reduced artefacts caused by unbound or free fluorophores but suffer from incomplete fluorescence quenching and significant re-illumination,which undermine bioimaging accuracy.Herein,through structural modifications to reinforce the hydrophobicity,planarity and rigidity of fluorophores with an aza-BODIPY framework,probes featuring absolute ACQ(aACQ)and negligible re-illumination are developed and evaluated in various nanocarriers.aACQ probes,FD-B21 and FD-C7,exhibit near-infrared emission,high quantum yield,photostability,water sensitivity,and negligible re-illumination in blood,plasma and 1%Tween-80 in contrast to ACQ probe P2 and conventional probe DiR.All nanocarriers can be labeled efficiently by the tested fluorophores.Polymeric micelles(PMs)labeled by different aACQ probes manifest similar biodistribution patterns,which however differ from that of DiR-labeled PMs and could be ascribed to the appreciable re-illumination of DiR.Significantly lower re-illumination is also found in aACQ probes(2%-3%)than DiR(20%-40%)in Caco-2,Hela,and Raw264.7 cells.Molecular dynamics simulations unravel the molecular mechanisms behind aggregation and re-illumination,supporting the hypothesis of planarity dependency.It is concluded that aACQ fluorophores demonstrate excellent water sensitivity and negligible fluorescence re-illumination,making themselves useful tools for more accurate bioimaging of nanocarriers.

Cytidine Mediated AuAg Nanoclusters as Bright Fluorescent Probe for Tumor Imaging in vivo

Noble metal clusters is an emerging class of fluorescent probes, avoiding most of the drawbacks of common fluorescent compounds, and they are simple to prepare and have good water solubility, good biocompatibility and excellent fluorescence properties. In this study, we have explored the synthesis of the cytidine mediated gold-silver nanoclusters （AuAg NCs） and applied it for both in vitro cellular imaging and tumor in vivo detection. Experimental results show that the as-prepared AuAg NCs can be used as a sensitive fluorescent probe for cancer cells/tissue de- tection. Especially, it is evident that under the relevant light irradiation with the wavelength of 488 nm, obviously bright fluorescence signal could be readily detected from focus location of inoculating tumor mouse, implying its possible application for the effective in vivo tumor bioimaging.

NIR-IIb excitable bright polymer dots with deep-red emission for in vivo through-skull three-photon fluorescence bioimaging

It is of great significance to study the brain structure and function in deep-tissue for neuroscience research and bio-medical applications because of the urgent demand for precise theranostics.Three-photon fluorescence microscopic(3PFM)bioimaging excited by the light in near-infrared IIb(NIR-IIb,1,500–1,700 nm)spectral region is one of the most promising imaging techniques with the advantages of high spatial resolution,large imaging depth,and reduced scattering.Herein,a type of NIR-IIb light excitable deep-red emissive semiconducting polymer dots(P-dots)with bright 3PF and large three-photon absorption cross-section(σ3)at 1,550 nm was prepared.Then the P-dots were functionalized with polystyrene polymer polystyrene graft ethylene oxide functionalized with carboxyl groups(PS-PEG-COOH)and modified with NH2-poly(ethylene glycol)(PEG)to synthesis photochemically stable and biocompatible P-dots nanoparticles(NPs).Further the P-dots NPs were utilized for in vivo 3PFM bioimaging of cerebral vasculature with and without the brain skull under 1,550 nm femtosecond(fs)laser excitation.In vivo 3PFM bioimaging of the mice cerebral vasculature at various vertical depths was obtained.Moreover,a vivid three-dimensional structure of the mice vascular architecture beneath the skull was reconstructed.At the depth of 350μm beneath the brain skull,3.8μm blood vessels could still be clearly recognized.NIR-IIb excitable P-dots assisted 3PFM bioimaging has great potential in accurate deep tissue bioimaging.

Bio-assembled smart nanocapsules for targeted delivery of KRAS shRNA and cancer cell bioimage

The five-year survival rate for pancreatic cancer is less than 5%. However, the current clinical multimodal therapy combined with first-line chemotherapy drugs only increases the patient’s median survival from 5.0 months to 7.2 months. Consequently, a new strategy of cancer treatments is urgently needed to overcome this high-fatality disease. Through a series of biometric analyses, we found that KRAS is highly expressed in the tumor of pancreatic cancer patients, and this high expression is closely related to the poor prognosis of patients. It shows that inhibiting the expression of KRAS has great potential in gene therapy for pancreatic cancer. Given those above, we have exploited the possibility of targeted delivery of KRAS shRNA with the intelligent and bio-responsive nanomedicine to detect the special oxidative stress microenvironment of cancer cells and realize efficient cancer theranostics. Our observations demonstrate that by designing the smart self-assembled nanocapsules of melanin with fluorescent nanoclusters we can readily achieve the bio-recognition and bioimaging of cancer cells in biological solution or serum.The self-assembled nanocapsules can make a significant bio-response to the oxidative stress microenvironment of cancer cells and generate fluorescent zinc oxide Nanoclusters in situ for targeted cell bioimaging. Moreover, it can also readily facilitate cancer cell suppression through the targeted delivery of KRAS shRNA and low-temperature hyperthermia. This raises the possibility to provide a promising theranostics platform and self-assembled nanomedicine for targeted cancer diagnostics and treatments through special oxidative stress-responsive effects of cancer cells.

Cadmium nanoclusters in a protein matrix： Synthesis, characterization, and application in targeted drug delivery and cellular imaging

Biotemplated metal nanoclusters have garnered much attention owing to their wide range of potential applications in biosensing, bioimaging, catalysis, and nanomedicine. Here, we report the synthesis of stable, biocompatible, watersoluble, and highly fluorescent bovine serum albumin-templated cadmium nanoclusters （CdNcs） through a facile one-pot green method. We covalently conjugated hyaluronic acid （HA） to the CdNcs to form a pH-responsive, tumor- targeting theranostic nanocarrier with a sustained release profile for doxorubicin （DOX）, a model anticancer drug. The nanocarrier showed a DOX encapsulation efficiency of about 75.6%. DOX release profiles revealed that 74% of DOX was released at pH 5.3, while less than 26% of DOX was released at pH 7.4 within the same 24-h period. The nanocarrier selectively recognized MCF-7 breast cancer cells expressing CD44, a cell surface receptor for HA, whereas no such recognition was observed with HA receptor-negative HEK293 cells. Biocompatibility of the nanocarrier was evaluated through cytotoxicity assays with HEK293 and MCF-7 ceils. The nanocarrier exhibited very low to no cytotoxicity, whereas the DOX-loaded nanocarrier showed considerable cellular uptake and enhanced MCF-7 breast cancer cell-killing ability. We also confirmed the feasibility of using the highly fluorescent nanoconjugate for bioimaging of MCF-7 and HeLa cells. The superior targeted drug delivery efficacy, cellular imaging capability, and low cytotoxicity position this nanoconjugate as an exciting new nanoplatform with promising biomedical applications.

Bio-safety assessment of carbon quantum dots, N-doped and folic acid modified carbon quantum dots: A systemic comparison

The carbon quantum dots(CQDs)and their functionalized materials are promising in biomedical field because of their unique properties;meanwhile,a growing concern has been raised about the potential toxicity of these modified materials in biosystem.In this study,we synthesized original CQDs and two common functionalized CQDs including N-doped CQDs(NCQDs)and folic acid-modified CQDs(FACQDs),and compared the toxicity and biocompatibility with each other in vitro and in vivo.L929,C6 and normal cell MDCK were selected to detect the adverse reaction of these materials in vitro.No acute toxicity or obvious changes were noted from in vitro cytotoxicity studies with the dose of these CQD materials increasing to a high concentration at 1 mg/mL.Among these materials,the FA-CQDs show a much lower toxicity.Moreover,in vivo toxicity studies were performed on the nude mice for 15 days.The experimental animals in 10 or 15 mg/kg groups were similar with animals treated by phosphate buffer solution(PBS)after 15 days.The results of the multifa rious biochemical parameters also suggest that the functionalized products of CQDs do not influence the biological indicators at feasible concentration.Our findings in vitro and in vivo through toxicity tests demonstrate that CQDs and their modified materials are safe for future biological applications.

Intelligent bio-assembly imaging-guided platform for real-time bacteria sterilizing and infectious therapy

Bacterial infection is rising as a threatening health issue.Because of the present delay in early diagnosis of bacterial diseases as well as the abuse of antibiotics,it has become a vital issue in the development of in-time detection and therapy of bacterial infections.Herein,we designed a multifunctional nanotheranostics platform based on the unique micro-environment of bacterial infections to achieve specific bioimaging and simultaneous inactivation of the target bacteria.We showed that in bacterial infections,the metal precursors(i.e.,HAuCl_(4),FeCl_(2),and herring sperm DNA)could be readily bio-self-assembled to multifunctional nanoclusters(NCs)that exhibit luminescence,in which AuCl_(4)-was biosynthesized via reductive biomolecules such as NADPH to the fluorescent AuNCs.The DNA may assist as an encapsulation and delivery vector,and Fe^(2+)served as a fluorescence intensifier and reduced reactive oxygen species(ROS)to produce the iron oxides.While the bacteria were being visualized,the microenvironment-responsive NCs were enabled to sterilize bacteria efficiently due to electrostatic effect,cell membrane destruction,inhibition of biofilm formation,and ROS accumulation.Besides,the bio-responsive self-assembled NCs complexes contributed to accelerating bacteria-infected wound healing and showed negligible side effects in long-term toxicity tests in vivo.Also,intracellular molecules involved in microenvironmental response were investigated.The work may become an effective strategy for the detection and real-time sterilization of intractable bacterial infections.