Functional Nanomaterials: From Structures to Biomedical Applications.

In recent decades, a number of functional nanomaterials have attracted a great amount of attention and exhibited excellent performance for biomedical and pharmaceutical applications [...].

Construction of a Rational-Designed Multifunctional Platform Based on a Fluorescence Resonance Energy Transfer Process for Simultaneous Detection of pH and Endogenous Peroxynitrite.

Peroxynitrite (ONOO-), a kind of reactive oxygen species, plays an indispensable role in many physiological processes. The stability and reactivity of ONOO- are significantly affected by the pH of the environment. A novel fluorescent probe RN-NA that can simultaneously respond to ONOO- and pH was proposed and constructed based on a rational-designed multifunctional fluorescence resonance energy transfer (FRET) platform. The RN-NA probe exhibited a remarkably different fluorescence change in response to ONOO- and pH. The fluorescence signals at 525 and 710 nm increased about 4-fold with a pH change from 8.0 to 3.0. The changes in fluorescence at 525 nm are mainly attributed to photo-induced electron transfer, and the fluorescence enhancement at 710 nm was mainly due to acid-induced open-closed circulation. In the presence of ONOO-, the fluorescence at 525 nm increased 5-fold, while the fluorescence at 710 nm was almost completely diminished. Up to 70-fold fluorescence enhancement was observed in the ratiometric channel F525/F710. In the cell imaging experiment, the intracellular pH was adjusted using H+/K+ ionophore and nigericin, and the endogenous ONOO- was generated by lipopolysaccharide (LPS) and γ-interferon (IFN-γ). The RN-NA probe can respond to cellular pH and endogenous ONOO- with remarkable fluorescence changes in both red/green and ratiometric channels.

Construction of a novel cell-trappable fluorescent probe for hydrogen sulfide (H2S) and its bio-imaging application.

Fluorescence detection of H2S in living organisms is greatly advantageous because it is nondestructive and can be used for in situ analysis. We have constructed a novel rhodamine analogue dye (Rho630) by extending the conjugated system of rhodamine to create a novel cell-trappable H2S fluorescent probe Rho630-AM-H2S with red light emission. Its application for H2S fluorescence detection in living HeLa cells and zebrafish was investigated. As expected, Rho630-AM-H2S showed a huge fluorescence turn-on response of about 20-fold at 630 nm and good selectivity toward H2S in solution. An MTT assay demonstrated that the probe showed negligible cytotoxicity in the concentrations typically used in fluorescence imaging experiments. Cell imaging experiments revealed that compared with compound 4 without cell-trappable unit modification, Rho630-AM-H2S exhibited remarkably enhanced cell penetration ability, as an enormous fluorescence signal increase was observed at the red channel within 5 min after Rho630-AM-H2S was incubated with HeLa cells. Finally, the probe Rho630-AM-H2S was used to detect H2S in living HeLa cells and zebrafish with great fluorescence enhancement in the red channel. Graphical abstract.

Remifentanil Protects against Lipopolysaccharide-Induced Inflammation through PARP-1/NF-κB Signaling Pathway.

Sepsis is a leading cause of death in patients with severe infection worldwide. Remifentanil is an ultra-short-acting, potent opioid analgesic. In the study, we aimed to investigate the role and underlying mechanism of remifentanil in lipopolysaccharide- (LPS-) induced inflammation in human aortic endothelial cells (HAECs). HAECs were pretreated with phosphate-buffered saline (PBS) or remifentanil (2.5 µM) for 30 min, then stimulated by LPS (10 µg/ml) for another 24 h. Poly(ADP-ribose) polymerase 1 (PARP-1) was inhibited by small interfering RNA (siRNA). Superoxide anion production and DNA damage were analyzed by dihydroethidium (DHE) staining and comet assay. The inducible nitric oxide synthase (iNOS), intercellular adhesion molecule 1 (ICAM-1), PARP-1, poly(ADP-ribose) (PAR), and nuclear factor-kappa B p65 (NF-κB p65) expressions were analyzed by RT-PCR or western blotting analysis. NF-κB p65 nuclear translocation was assessed by immunofluorescence. Compared with the control group, pretreatment with remifentanil significantly reduced superoxide anion production and DNA damage, with downregulation of iNOS, ICAM-1, and PARP-1 expressions as well as PAR expression. Moreover, pretreatment with PARP-1 siRNA or remifentanil inhibited LPS-induced NF-κB p65 expression and nuclear translocation. Remifentanil reduced LPS-induced inflammatory response through PARP-1/NF-κB signaling pathway. Remifentanil might be an optimal choice of analgesia in septic patients.

"Print-to-pattern": Silk-Based Water Lithography.

The requirement of nontoxic and versatile manufacturing frameworks for biologically relevant applications has imposed significant constraints on the choice of functional materials and the complementary fabrication tools. In this context, silk is actively studied, thanks to its mechanical robustness, biocompatibility, wide availability, aqueous processing conditions, and ease of functionalization. The inherent matching between the water solubility of silk and the aqueous inks of the inkjet printing (IJP) process has derived a biofriendly and versatile "print-to-pattern" scheme-termed silk-based water lithography-toward scalable functional biomanufacturing. The deposition mode of IJP and the etching effect of silk film by water features a dual tone fabrication where functional molecules are dispensed additively, while the silk film is patterned in a "subtractive" fashion. Such versatility and scalability pave the way to a wide range of opportunities in the biomedical field.

Self-Powered Multifunctional Transient Bioelectronics.

Controllable degradation and excellent biocompatibility during/after a lifetime endow emerging transient electronics with special superiority in implantable biomedical applications. Currently, most of these devices need external power sources, limiting their real-world utilizations. Optimizing existing bioresorbable electronic devices requires natural-material-based construction and, more importantly, diverse or even all-in-one multifunctionalization. Herein, silk-based implantable, biodegradable, and multifunctional systems, self-powered with transient triboelectric nanogenerators (T2 ENGs), for real-time in vivo monitoring and therapeutic treatments of epileptic seizures, are reported. These T2 ENGs are of customizable in vitro/in vivo operating life and biomechanical sensitivity via the adjustments of silk molecular size, surface structuralization, and device configuration. Functions, such as drug delivery and structural-integrity optical readout (parallel to electronic signals), are enabled for localized anti-infection and noninvasive degradation indication, respectively. A proof-of-principle wireless system is built with mobile-device readout and "smart" treatment triggered by specific symptoms (i.e., epilepsy), exhibiting the practical potential of these silk T2 ENGs as self-powered, transient, and multifunctional implantable bioelectronic platforms.

Rational Design of a Robust Fluorescent Probe for the Detection of Endogenous Carbon Monoxide in Living Zebrafish Embryos and Mouse Tissue.

Carbon monoxide (CO) is one of the most important gaseous signal molecules in biological systems. However, the investigation of the functions of CO in living organisms is restricted by the lack of functional molecular tools. To address this critical challenge, we present herein the rational design, synthesis, and in vivo imaging studies of a powerful two-photon excited near-infrared fluorescent probe (1-Ac) for endogenous CO monitoring. The advantageous features of the new probe include high stability, low background fluorescence, large fluorescence enhancement, high sensitivity, and two-photon excitation with emission in the near-infrared region. Significantly, these merits of the probe enable the tracking of endogenous CO in zebrafish embryos and mouse tissues for the first time.

Mitochondria-Directed Fluorescent Probe for the Detection of Hydrogen Peroxide near Mitochondrial DNA.

It is important to detect hydrogen peroxide (H2O2) near mitochondrial DNA (mtDNA) because mtDNA is more prone to oxidative attack than nuclear DNA (nDNA). In this study, a mitochondria-targeted fluorescence probe, pep3-NP1, has been designed and synthesized. The probe contains a DNA-binding peptide, a H2O2 fluorescence reporter, and a positively charged red emissive styryl dye to facilitate accumulation in mitochondria. Due to groove binding of the peptide with DNA, the styryl dye of pep3-NP1 intercalated into the bases of DNA, leading to an increase in red fluorescence intensity (centered at 646 nm) and quantum yield. In this case, pep3-NP1 was a turn-on probe for labeling DNA. Subcellular locations of pep3-NP1 and MitoTracker suggested that pep3-NP1 mostly accumulated in the mitochondria of live cells. Namely, as an intracellular DNA marker, pep3-NP1 bound to mtDNA. In the presence of H2O2, pep3-NP1 emitted green fluorescence (centered at 555 nm). Thus, the ratio of green with red fluorescence of pep3-NP1 was suitable to reflect the change of the H2O2 level near mtDNA in living cells. The detecting limit for H2O2 was estimated at 2.9 and 5.0 µM in vitro and in cultured cells, respectively. The development of pep3-NP1 could help in studies to protect mtDNA from oxidative stress.

A Programmed DNA Marker Based on Bis(4-ethynyl-1,8-naphthalimide) and Three-Methane-Bridged Thiazole Orange.

Two large conjugated naphthalimide derivatives with or without three-methane-bridged thiazole orange (TO3; i.e., compounds 1 a and 2 a, respectively) were designed and synthesized. The fluorescence of the naphthalimide group in compound 1 a at λ=532 nm initially decreased and that for the TO3 group at λ=655 nm increased sequentially upon adding Salmon testes (St) DNA. In contrast, without the TO3 group, the fluorescence intensity of compound 2 a monotonously decreased in response to the addition of DNA. The non-monotonic change in the fluorescence for compound 1 a could be divided into two linear sections with two different wavelengths in the range of 0

A highly sensitive ratiometric fluorescent probe for the detection of cytoplasmic and nuclear hydrogen peroxide.

As a marker for oxidative stress and a second messenger in signal transduction, hydrogen peroxide (H2O2) plays an important role in living systems. It is thus critical to monitor the changes in H2O2 in cells and tissues. Here, we developed a highly sensitive and versatile ratiometric H2O2 fluorescent probe (NP1) based on 1,8-naphthalimide and boric acid ester. In response to H2O2, the ratio of its fluorescent intensities at 555 and 403 nm changed 1020-fold within 200 min. The detecting limit of NP1 toward H2O2 is estimated as 0.17 µM. It was capable of imaging endogenous H2O2 generated in live RAW 264.7 macrophages as a cellular inflammation response, and especially, it was able to detect H2O2 produced as a signaling molecule in A431 human epidermoid carcinoma cells through stimulation by epidermal growth factor. This probe contains an azide group and thus has the potential to be linked to various molecules via the click reaction. After binding to a Nuclear Localization Signal peptide, the peptide-based combination probe (pep-NP1) was successfully targeted to nuclei and was capable of ratiometrically detecting nuclear H2O2 in living cells. These results indicated that NP1 was a highly sensitive ratiometric H2O2 dye with promising biological applications.

Solvent induced helical aggregation in the self-assembly of cholesterol tailed platinum complexes.

Three alkynylplatinum(ii) bipyridyl complexes in which two cholesterol groups are combined with a bipyridyl group via alkyl chains and amido bonds were designed and synthesized. The complexes have different lengths of ethylene glycol chains at the para-position of 1-phenylethyne. All three complexes can self-assemble to gel networks in DMSO, while only the morphology of 1a without an ether chain shows a well-defined right-handed helical structure in layer packing mode. However, 1c with long ethylene glycol chains forms perfect regular left-handed helical structures in aqueous ethanol solution while the volume percentage of water is less than 5% (v/v). As the ratio of water increases, the chirality changes from a left-handed helix to a right-handed helix and the packing mode alters from a monolayer structure to a hexagonal structure. As the ratio of water further increases to greater than 50% (v/v), the structure of the assembly finally transforms into bilayer vesicles. The process of the morphology transition is traced by circular dichroism spectra, powder X-ray diffraction, SEM and TEM images. The result indicates that a polar solvent (water) acts as a trigger to change the self-assembly of the chiral structures of the complex due to the strong hydrophobic interaction between cholesterol groups and the balance of the hydrophobicity and hydrophilicity of the solvent environment.

(-)-Menthol based thixotropic hydrogel and its application as a universal antibacterial carrier.

A kind of novel hydrogelator based on (-)-menthol, a traditional cooling compound, tailed by an amino acid derivate through an alkyl chain, has been designed and synthesized. The hydrogelator containing an l-lysine can form a stable hydrogel with thixotropic character in a large pH range. An interesting feature is that the viscoelastic character of the hydrogel can be enhanced by mechanical force. The mechanism of the self-assembly process was investigated by means of IR, SEM, AFM and X-ray diffraction. The formation of three dimensional multiporous networks through acid base interactions and strong double hydrogen bonding between amino acids is proposed to be the driving force for the construction of the stable hydrogel. As a result, the hydrogelator can further gelate aqueous solutions of some confirmed antibacterial agents such as Zn(2+) and a series of water soluble organic antibiotic medicines like lincomycin, amoxicillin, etc., in such a unique way that the concentration of the antibacterial agents loaded into the hydrogel can be tuned to a large extent. The antimicrobial susceptibility of the hydrogels loaded with Zn(2+) or lincomycin is much more effective than that of the corresponding aqueous solution tested by the Oxford cup method. Furthermore, the hydrogelator is completely innoxious to living cells by measurement of MTT assay. Thus, the hydrogel can be developed as a universal carrier for antibacterial agents and may also be widely used in the fields of cell culture, tissue engineering, or drug delivery systems.

Fluorescence turn-on detection of Sn2+ in live eukaryotic and prokaryotic cells.

Sn(2+) is usually added to toothpaste to prevent dental plaque and oral disease. However, studies of its physiological role and bacteriostatic mechanism are restricted by the lack of versatile Sn(2+) detection methods applicable to live cells, including Streptococcus mutans. Here we report two Sn(2+) fluorescent probes containing a rhodamine B derivative as a fluorophore, linked via the amide moiety to N,N-bis(2-hydroxyethyl)ethylenediamine (R1) and tert-butyl carbazate group (R2), respectively. These probes can selectively chelate Sn(2+) and show marked fluorescence enhancement due to the ring open reaction of rhodamine induced by Sn(2+) chelation. The probes have high sensitivity and selectivity for Sn(2+) in the presence of various relevant metal ions. Particularly, both R1 and R2 can target lysosomes, and R2 can probe Sn concentrations in lysosomes with rather acidic microenvironment. Furthermore, these two probes have low toxicity and can be used as imaging probes for monitoring Sn(2+) not only in live KB cells (eukaryotic) but also in Streptococcus mutans cells (prokaryotic), which is a useful tool to study the physiological function of Sn(2+) in biological systems.

Near-Infrared Fluorescent Probe with pH- and Viscosity-Switchable Performance for the Detection of Thrombi in Live Animals and Organs.

Blood viscosity changes and blood clots are high-impact diseases, but the pathogenic mechanisms and detection methods are still limited. Due to the complexity of the cellular microenvironment, viscosity is a key factor in regulating the behavior of mitochondria and lysosomes in cells. Conventional fluorescence probes are highly restrictive for complex viscosity detection in live animals. Therefore, we developed two near-infrared fluorescence probes, QL1 and QL2, with dual responses to the pH and viscosity. Notably, QL2 has two maximum fluorescence emissions at 680 and 750 nm, when excitation by 580 and 700 nm, respectively. QL2 exhibited both a pH and viscosity switchable fluorescence response. The two emission peaks exhibited a reverse change trend: the fluorescence at 680 nm decreased by 90%, and the fluorescence at 750 nm increased by about 5-fold with pH from 2 to 10. Meanwhile, both emission peaks show remarkable fluorescence enhancement toward viscosity change, with 185 and 32 times enhancement, respectively. The sensing mechanism and spectral changes are confirmed by DFT calculations. QL2 was further used for viscosity imaging in live cells, zebrafish, and live animals. Most importantly, QL2 is able to successfully track changes in blood clots in live mice and organs, thus enabling the study of blood clots in cerebral strokes and the underlying pathological mechanisms.

Strategies for measuring concentrations and forms of amyloid-ß peptides.

Alzheimer's disease (AD) is affecting more and more people worldwide without the effective treatment, while the existed pathological mechanism has been confirmed barely useful in the treatment. Amyloid-ß peptide (Aß), a main component of senile plaque, is regarded as the most promising target in AD treatment. Aß clearance from AD brain seems to be a reliably therapeutic strategy, as the two exited drugs, GV-971 and aducanumab, are both developed based on it. However, doubt still exists. To exhaustive expound on the pathological mechanism of Aß, rigorous analyses on the concentrations and aggregation forms are essential. Thus, it is attracting broad attention these years. However, most of the sensors have not been used in pathological studies, as the lack of the bridge between analytical chemist and pathologists. In this review, we made a brief introduce on Aß-related pathological mechanism included in ß-amyloid hypothesis to elucidate the detection conditions of sensor methods. Furthermore, a summary of the sensor methods was made, which were based on Aß concentrations and form detections that have been developed in the past 10 years. As the greatest number of the sensors were built on fluorescent spectroscopy, electrochemistry, and Roman spectroscopy, detailed elucidation on them was made. Notably, the aggregation process is another important factor in revealing the progress of AD and developing the treatment methods, so the sensors on monitoring Aß aggregation processes were also summarized.

A fluorescent probe for detecting H2O2 and delivering H2S in lysosomes and its application in maintaining the redox environments.

Hydrogen peroxide (H2O2) is a reactive oxygen species (ROS) that can be used as a marker for the occurrence of oxidative stress in the organism. Lysosomes serve as intracellular digestive sites, and when the concentration of H2O2 in them is abnormal, lysosomal function is often impaired, leading to the development of diseases. Hydrogen sulfide (H2S) acts as a gaseous signaling molecule that scavenges H2O2 from cells and tissues, thereby maintaining the redox environment of the body. However, most of the reported hydrogen peroxide fluorescent probes so far can only detect H2O2, but cannot maintain the intracellular redox environment. In this paper, an H2O2 fluorescent probe LN-HOD with lysosomal targeting properties was designed and synthesized by combining the H2O2 recognition site with a naphthylamine fluorophore via a thiocarbamate moiety. The probe has the advantages of large Stokes shift (110 nm), high sensitivity and good H2S release capability. The probe LN-HOD can be used to detect H2O2 in cells, zebrafish and plant roots. In addition, LN-HOD detects changes in the concentration of H2O2 in plant roots when Arabidopsis is stressed by cadmium ion (Cd2+). And through its ability to release H2S, it can help to remove excess H2O2 and maintain the redox environment in cells, zebrafish and plant roots. The present work provides new ideas for the detection and assisted removal of H2O2, which contributes to the in-depth study of the cellular microenvironment in organisms.

Near-infrared dual-response fluorescent probe for detection of N2H4 and intracellular viscosity changes in biological samples and various water samples.

N2H4 is a common raw material used in the production of pesticides and has good water solubility, so it may contaminate water sources and eventually enter living organisms, causing serious health problems. Viscosity is an important indicator of the cellular microenvironment and an early warning signal for many diseases. The high reactivity of hydrazine depletes glutathione (GSH) in hepatocytes, causing oxidative stress ultimately leading to significant changes in intracellular viscosity and even death. Therefore, it is particularly important to develop an effective method to detect N2H4 and viscosity in environmental and biological systems. On this basis, we developed two fluorescent probes, BDD and BHD, based on xanthene and 2-benzothiazole acetonitrile. The experimental results show that BHD and BDD have good imaging capabilities for N2H4 in cells, zebrafish and Arabidopsis. BHD and BDD also showed sensitive detection and fluorescence enhancement in the near-infrared region when the intracellular viscosity was changed. Notably, the probe BDD has also successfully imaged N2H4 in a variety of real water samples.

Construction of a pH- and viscosity-switchable near-infrared fluorescent probe and its imaging application.

Viscosity is a parameter used to measure the fluidity of liquids and a key indicator in evaluating the states of body fluid in biological tissues and lesions. Most traditional detection methods have many drawbacks such as a short emission wavelength and interference by background fluorescence. Inspired by the multiple double bond structure of retinal, a novel pH and viscosity dual-response fluorescent probe (Rh-TR) was constructed in this study. Rh-TR exhibited two emission signals centered at 510 and 660 nm. As the pH of the phosphate-buffered saline increased, the fluorescence at 510 nm increased by about 124-fold, while the change in fluorescence at 660 nm was not obvious. When detecting the change in viscosity using the probe, the fluorescence at 510 nm decreased by about 85 %, while the fluorescence at 660 nm increased by over 20-fold. The probe also showed high selectivity and little toxicity. As demonstrated by the biological imaging experiment, the probe successfully imaged changes in the pH and viscosity of cells and in a live animal model of zebrafish. Considering the unique structure of Rh-TR with retinal and its pH- and viscosity-switchable spectral property, the probe may find further application in detecting viscosity-related diseases and industrial detection.

A chitosan-based near-infrared ratiometric fluorescent nanoprobe created by molecular assembly with applications in hypochlorous acid detection in live mouse.

Hypochlorous acid (HClO/ClO-) is a key reactive oxidative species (ROS) in the body. The HClO/ClO- concentrations are imbalanced during cancer formation due to the ROS stress response. This paper introduces a novel chitosan-based self-calibration fluorescent nanoprobe (ChCyNil) constructed by molecular assembly for the ratiometric detection of HClO/ClO-. Two chromophores with different fluorescence characteristics and HClO/ClO- sensitivity were labeled on chitosan, and nanoparticles were prepared by a self-assembly strategy for HClO/ClO- detection. ChCyNil exhibits several advantages, such as dual near-infrared emissions at 670 nm and 845 nm, tunable fluorescence intensity, self-calibration fluorescence, and good biocompatibility, improving its accuracy in HClO/ClO- detection. Our study confirmed that ChCyNil exhibits a well-assembled spheroidal nanostructure and good photophysical properties in solution. The fluorescence imaging properties were further proved by detecting endogenous HClO/ClO- produced by LPS/PMA stimuli in cells and zebrafish. In addition, ChCyNil was used to detect the fluorescence behavior of HClO/ClO- in tumors of live mice. The successful design and fabrication of ChCyNil have presented a new strategy for constructing detection tools with improved fluorescence properties for HClO/ClO- in live animals.