Cascade CRISPR/Cas12a and DSN for the electrochemical biosensing of miR-1246 in BC-derived exosomes.

MiR-1246 in breast cancer-derived exosomes was a promising biomarker for early diagnosis of breast cancer(BC). However, the low abundance, high homology and complex background interference make the accurate quantitative detection of miR-1246 facing great challenges. In this study, we developed an electrochemical biosensor based on the subtly combined of CRISPR/Cas12a, double-stranded specific nuclease(DSN) and magnetic nanoparticles(MNPs) for the detection of miR-1246 in BC-derived exosomes. Ascribed to the good synergistic effect of DSN, Cas12a and MNPs, the developed electrochemical biosensor exhibited excellent performance with the linear range from 500 aM to 5 pM, and the detection limit as low down to about 50 aM. The target-specific triggered enzyme-digest activity of DSN and Cas12a system, as well as the powerful separation ability of MNPs ensure the high specificity of developed electrochemical biosensor which can distinguish single base mismatches. In addition, the developed electrochemical biosensor has been successfully applied to detect miR-1246 in blood-derived exosomes and realize distinguishing the BC patients from the healthy individuals. It is expected that the well-designed biosensing platform will open up new avenues for clinical liquid biopsy and early screening of breast cancer, as well as provide deeper insights into clinical oncology treatment.

Emerging Drug Delivery Vectors: Engineering of Plant-Derived Nanovesicles and Their Applications in Biomedicine.

Extracellular vesicles can transmit intercellular information and transport biomolecules to recipient cells during various pathophysiological processes in the organism. Animal cell exosomes have been identified as potential nanodrugs delivery vehicles, yet they have some shortcomings such as high immunogenicity, high cytotoxicity, and complicated preparation procedures. In addition to exosomes, plant-derived extracellular vesicles (PDVs), which carry a variety of active substances, are another promising nano-transport vehicles emerging in recent years due to their stable physicochemical properties, wide source, and low cost. This work briefly introduces the collection and characterization of PDVs, then focuses on the application of PDVs as natural or engineered drug carriers in biomedicine, and finally discusses the development and challenges of PDVs in future applications.

Exogenous Spermidine and Amino-Ethoxyvinylglycine Improve Nutritional Quality via Increasing Amino Acids in Rice Grains.

Polyamines and ethylene are key regulators of the growth and development, quality formation, and stress response of cereal crops such as rice. However, it remains unclear whether the application of these regulators could improve the nutritional quality via increasing amino acids in rice grains. This study examined the role of exogenous polyamines and ethylene in regulating amino acid levels in the milled rice of earlier-flowered superior grain (SG) and later-flowered inferior grain (IG). Two rice varieties were field grown, and either 1 mmol L-1 spermidine (Spd) or 50 µmol L-1 amino-ethoxyvinylglycine (AVG) was applied to panicles at the early grain-filling stage. The control check (CK) was applied with deionized water. The results showed that the Spd or AVG applications significantly increased polyamine (spermine (Spm) and Spd) contents and decreased ethylene levels in both SG and IG and significantly increased amino acid levels in the milled rice of SG and IG relative to the CK. Collectively, the application of Spd or AVG can increase amino acid-based nutritional quality and grain yield via increasing polyamine (Spm and Spd) contents and reducing ethylene levels in both SG and IG of rice.

Multifunctional nanocarrier with self-catalytic production of nitric oxide for photothermal and gas-combined therapy of tumor.

Single treatment often faces the problem that it cannot completely eradicate tumor and inhibit the tumor metastasis. In order to overcome this shortcoming, multi-modal tumor treatment has attracted widespread attention. In the present article, based on ascorbyl palmitate (PA) and l-arginine (l-Arg), a multifunctional nanocarrier is designed for synergetic treatment of tumor with photothermal and nitric oxide (NO) gas therapy. Firstly, PA and l-Arg were self-assembled to form novel functional micelles, PL, with high biosafety using electrostatic interaction and hydrogen bonding. The functional micelles could self-catalyze to produce NO at the tumor site. Then, Ag2S quantum dots having fluorescence imaging and photothermal properties were encapsulated to obtain the nanocarrier, A@PL. The results show that A@PL had a hydrated size of around 78 nm and presented good stability within 30 d. Moreover, in vitro studies indicate that it was efficient with regards to NO self-generating capacity, whereas the photothermal conversion efficiency was as high as 34% under near-infrared light irradiation. The cytotoxicity results show that, when the concentration of A@PL was as high as 2 mM, the survival rate of 3 T3 cells was still 78.23%, proving that the probe has good safety characteristics. Fluorescence imaging results show that its maximum enrichment can be achieved at the tumor site after tail vein injection for 3 h, and out of the body after 24 h, indicating good internal circulation. The in vivo studies show that the rate of inhibition of tumor using the nanocarrier was as high as 98%, and almost overcame the problem of tumor recurrence caused by single treatment, thus presenting a significant tumor treatment effect. This new multifunctional nanocarrier with self-catalytic production of NO provides a new idea for the efficient treatment of tumors.

A simple fluorescent strategy for liver capillary labeling with carbon quantum dot-lectin nanoprobe.

Taking the hepatic sinusoid (HS) as the main delivery area of liver nutrients and metabolic waste, recognizing its structure is important for a deep understanding of liver function. In this paper, based on lycopersicon esculentum lectin (LEL), with targeting ability for endothelial cells, and carbon quantum dots (CQDs), with high biosafety, an LEL-coupled CQD immunofluorescence probe (CQD@LEL) that can label microvessels is designed and used for the fluorescence labeling and imaging of HS in liver tissue sections. The CQD size is approximately 2 nm. Blue fluorescence is emitted under excitation; its optimal excitation wavelength is 400 nm while the emission is at about 450 nm. Gel electrophoresis and capillary electrophoresis confirm that glutaraldehyde can couple LEL to CQD, and the obtained CQD@LEL retains the fluorescence property and has good stability. Optimization experiments show that its labeling effect is positively correlated with time and probe concentration for dyeing the blood vessels of mouse liver slices. In order to improve the effect further, a probe concentration of 0.17 mg mL-1 and incubation time of 3 h were chosen to label the liver tissue sections. The results show that the liver microvessels are formed by interstitial structures among the hepatic cords, and the HS presents a granular or patchy appearance. H&E and ultrathin section TEM show that the microvascular wall of the liver is composed of discontinuous endothelial cells, and there are Kupffer cells and other cells in the tubes, proving that our probe can clearly label the structure and morphology of liver microvessels. This work is of great significance for the visualization of HS.

CdTe@CdS quantum dots for labeling and imaging macrophages in liver frozen sections below the freezing point.

CdTe@CdS core-shell quantum dots with different particle sizes are synthesized by an aqueous method, and coating them with a CdS shell layer improves the quantum yield (36% → 59%) and fluorescence stability (37% → 77%) of CdTe@CdS quantum dots. When the KCl concentration (mass fraction) in the system is 15%, the CdTe@CdS quantum dot dispersion system remains in the liquid state at -20 °C, and the low temperature increases the fluorescence intensity. A QD-Ab probe is obtained after CdTe@CdS quantum dots are coupled with IgG; the circular dichroism shows that the IgG protein structure is not destroyed, while capillary electrophoresis, agarose gel electrophoresis and flow cytometry verify the conjugation efficiency. With rabbit anti-mouse EMR1 antibody as the primary antibody and QD-Ab as the secondary antibody, the hepatic macrophages in liver frozen sections are fluorescently labeled at -20 °C, and it is found that they are radially distributed in hepatic sinusoids with specific and highly efficient labeling; these results are verified by H&E staining and TEM. This technology can provide important technical support for in-depth understanding of the distribution of liver immune cells in the liver, and it can further provide a scientific basis to understand the relationship between the liver structure and function and pathological changes.

A self-assembled nanoplatform based on Ag2S quantum dots and tellurium nanorods for combined chemo-photothermal therapy guided by H2O2-activated near-infrared-II fluorescence imaging.

A nanoplatform based on Ag2S quantum dots (QDs) and tellurium nanorods (TeNRs) was developed for combined chemo-photothermal therapy guided by H2O2-activated near-infrared (NIR)-II fluorescence imaging. Polypeptide PC10AGRD-modified TeNRs and Ag2S QDs were co-encapsulated in 4T1 cell membrane to prepare a nanoplatform (CCM@AT). Ag2S QDs and TeNRs in the CCM@AT were used as a fluorescence probe and photosensitizer, and a chemotherapeutic prodrug and quenching agent to quench the fluorescence of Ag2S QDs, respectively. After the CCM@AT was specifically targeted to the tumor site, the TeNRs were dissolved by the high concentration of H2O2 at the tumor site to light up the fluorescence of Ag2S QDs for NIR-II fluorescence imaging. In addition, the generated toxic TeO66- molecules decreased ATP production by selective cancer chemotherapy, which is beneficial for photothermal therapy. The elevated temperature due to photothermal therapy in turn promoted the chemical reaction in chemotherapy. In vitro and in vivo toxicity results showed that the CCM@AT possesses high biocompatibility. Compared to single photothermal therapy and chemotherapy, the synergistic chemo-photothermal therapy can effectively suppress the growth of 4T1 tumor. This all-in-one nanoplatform provides a boulevard for the combination therapy of tumors guided by NIR-II fluorescence imaging. STATEMENT OF SIGNIFICANCE: NIR-II fluorescence imaging shows the characteristics of low tissue absorption, reflection, and scattering, which can greatly reduce the influence of autofluorescence in vivo. However, the non-negligible effect of autofluorescence is still observed in fluorescence imaging in vivo. Therefore, there is an urgent need to develop a strategy of controlled release of fluorescence for accurate imaging and tumor therapy. Here, Ag2S quantum dots (QDs) with NIR-II fluorescence emission and good photothermal conversion efficiency are used as a fluorescence probe and photosensitizer, and tellurium nanorods (TeNRs) are used as a chemotherapeutic prodrug and quenching agent to quench the fluorescence of Ag2S QDs. This multiple nanoplatform provides an inspiration for the combination therapy of tumor guided by NIR-II fluorescence imaging.

Tumor Microenvironment-Activated Theranostics Nanozymes for Fluorescence Imaging and Enhanced Chemo-Chemodynamic Therapy of Tumors.

Chemodynamic therapy (CDT) is widely explored for tumor-specific therapy by converting endogenous H2O2 to lethal ·OH to destroy cancer cells. However, ·OH scavenging by glutathione (GSH) and insufficient intratumoral H2O2 levels seriously hinder the application of CDT. Herein, we reported the fabrication of copper ion-doped ZIF-8 loaded with gold nanozymes and doxorubicin hydrochloride (DOX) for the chemotherapy and CDT synergistic treatment of tumors with the assistance of tumor microenvironment (TME)-activated fluorescence imaging. The Cu2+-doped ZIF-8 shell was gradually degraded to release DOX and gold nanoclusters responding to the acidic TME. The fluorescence signal of the tumor region was acquired after the quenched fluorescence of the gold nanoclusters by Cu2+ and DOX by aggregation-induced quenching was turned on because of the interaction of GSH with Cu2+ and the release of free DOX. The Cu2+ ions could deplete the GSH via redox reactions and the generated Cu+ could convert internal H2O2 to ·OH for tumor CDT. The chemotherapeutic effect of DOX was strengthened through drug efflux inhibition and drug sensitivity increase due to the consumption of GSH and ·OH burst. Moreover, DOX could raise the level of H2O2 and augment the effect of CDT. In addition, the fluorescent gold nanoclusters not only served as a peroxidase to convert H2O2 to ·OH but also employed as an oxidase to consume GSH, resulting in the amplification of chemotherapy and CDT. This work presents an approach to construct tumor microenvironment-activated theranostic probes without external stimuli and to achieve the tumor elimination through cascade reactions and synergistic treatment.

Labeling of liver cells with CdSe/ZnS quantum dot-based fluorescence probe below freezing point.

In this paper, CdSe/ZnS quantum dots (QDs) with particle size of 5.5 ~ 9.3 nm were synthesized, and the fluorescence emission ranged from 545 ~ 616 nm. When the volume fraction of ethanol was 30%, the water-soluble QD dispersion system remained liquid under -20 °C freezing conditions, the fluorescence intensity increased with a decrease in temperature, and the quantum yield reached 79% at -20 °C. The endothelial cell adhesion molecule CD31 antibody (anti-CD31) was used as the primary antibody, QDs were coupled with IgG as the secondary antibody (QD-Ab), and effective labeling of hepatic sinusoid endothelial cells was achieved at -20 °C. Fluorescence imaging and flow cytometry analysis showed that the labeling efficiency was as high as 97%, indicating that QDs have an important application prospect in microscopic section tomography of the liver.

Jasmonates Alleviate the Harm of High-Temperature Stress During Anthesis to Stigma Vitality of Photothermosensitive Genetic Male Sterile Rice Lines.

Using photothermosensitive genic male sterile (PTSGMS) rice (Oryza sativa L.) lines to produce hybrids can obtain great heterosis. However, PTSGMS rice lines exhibit low stigma vitality when high-temperature (HT) stress happens during anthesis. Jasmonates (JAs) are novel phytohormones and play vital roles in mediating biotic and abiotic stresses. Little is known, however, if and how JAs could alleviate the harm of HT stress during anthesis to the stigma vitality of PTSGMS lines. This study investigated the question. Two PTSGMS lines and one restorer line of rice were pot-grown and subjected to normal temperature and HT stress during anthesis. The stigma exertion rate, sigma fresh weight, stigma area, contents of JAs, hydrogen peroxide (H2O2), and ascorbic acid (AsA), activity of catalase in stigmas, and the number of pollens germinated on the stigma of PTSGMS lines were determined. The results showed that a rice line with higher JAs content in the stigma under HT stress showed lower H2O2 content, higher AsA content and catalase activity in stigmas, larger stigma area, heavier stigma fresh weight, more pollens germinated on the stigma, and higher fertilization and seed-setting and rates. Applying methyl JAs during anthesis to rice panicles decreased the accumulation of reactive oxygen species and enhanced stigma vitality, thereby increasing fertilization and seed-setting rates of the hybrids of PTSGMS rice lines under HT stress. The results demonstrate that JAs attenuate the injury of HT stress to the stigma vitality of PTSGMS rice lines through enhancing antioxidant ability.

Quantitative analysis of various targets based on aptamer and functionalized Fe3O4@graphene oxide in dairy products using pregnancy test strip and smartphone.

Pregnancy test strips are one of the most mature and widely used commercial lateral flow devices used to determine pregnancy. Being a simple and rapid detection method, human chorionic gonadotropin (hCG) was used with different aptamers (hCG-apt) as probes for the detection of metal ions, small organic molecules, and proteins. Quantitative detection of target analytes was achieved using a smartphone app and a portable device developed in our laboratory. The results showed detection ranges of 1 nM-1 µM, 0.1 nM-10 µM and 32 nM-500 nM for Pb2+, chloramphenicol, and ß-lactoglobulin, respectively, and the corresponding visual detection limits in dairy products were 5 nM, 1 nM and 50 nM, respectively. Based on these results, rapid detection of multiple analytes can be realized through aptamer modification, thereby broadening the application range of commercial lateral flow devices for analysis of food chemistry.

Injectable polypeptide-engineered hydrogel depot for amplifying the anti-tumor immune effect induced by chemo-photothermal therapy.

The immunosuppressive tumor microenvironment has caused great obstacles to tumor immunotherapy, especially where less tumor-associated antigens are released from tumor sites. Herein, a Ag2S QD/DOX/Bestatin@PC10ARGD genetically engineered polypeptide hydrogel PC10ARGD as a sustained-release material was developed for mammary carcinoma treatment. A near-infrared silver sulfide (Ag2S) QD as a photosensitizer was encapsulated into the hydrophobic cavity formed by the self-assembly of the polypeptide nanogel (PC10ARGD) for photothermal therapy. The water-soluble drug DOX and Bestatin were integrated into the PC10ARGD hydrogel. The photothermal effect could trigger the sustained release of the DOX, which could be applied to initiate in situ vaccination. Bestatin as an immune-adjuvant drug could amplify the body's immune function. The results of in vivo therapy tests exhibited that the Ag2S QD/DOX/Bestatin@PC10ARGD hydrogel with laser irradiation could activate anti-tumor immune effects that inhibit the growth of primary tumors and distal lung metastatic nodules. Meanwhile, a safer lower-temperature with multiple laser irradiation treatment strategy exhibited more effective tumor-killing performance (84.4% tumor inhibition rate) and promoted the penetration of immune cells into the tumor tissue. The CD8+ and CD4+ cytotoxic T cells ratio was increased by 5.3 and 10 times, respectively, thus exhibiting a good prognostic signal. The multifunctional polypeptide hydrogel as a green manufacturing and engineering material is promising to serve as a cancer vaccine for anticancer applications.

An in situ synthesis of silver nanoparticle-loaded genetically engineered polypeptide nanogels for antibacterial and wound healing applications.

The fact that the wounds infected by bacteria are difficult to heal is a major health issue. Herein, we synthesized silver nanoparticle-loaded polypeptide nanogels via an in situ method using UV irradiation, which is a relatively green and simple method. The size of the nanogel and silver nanoparticles could be regulated by changing the concentrations of the polypeptide and silver ions, respectively. Because the polypeptide PC10ARGD was histidine-rich and biodegradable, the as-synthesized silver nanogels exhibited low toxicity and good biocompatibility. The in vitro antibacterial experiments showed that the silver nanogels presented excellent antibacterial activity against both Gram-negative and Gram-positive bacteria. Moreover, silver nanogels had a good effect as an antibacterial and wound healing agent in vivo. These findings provide a new strategy and theoretical basis for the synthesis and application of silver nanoparticles.

Ion current rectification in combination with ion current saturation.

Over the decades, nanochannels have been widely used for single molecule detection, smart sensors, and energy transfer and storage based on its unique ion transport properties. Although various ion transport phenomena of nanochannels have been reported, the discovery of new ion transport phenomena is still of great significance for understanding material transport of nanochannels and development of nanodevices with unique working capabilities. This article reports a novel nanochannel ion transport phenomenon - ion current rectification in combination with ion current saturation (ICR-S), which arised from a mesoporous titania conical microplug generated in situ in the glass micropipette tip cavity by space confinement evaporation. The ion current of forward voltage is greater than that of reverse voltage, and the saturation currents appear in both the forward and reverse voltages, the ratio of forward and reverse saturation current can reaches to 10. In addition, the influence of pH, ionic strength, and micropipette angle on ICR-S is also investigated.

Antitumor immunity triggered by photothermal therapy and photodynamic therapy of a 2D MoS2 nanosheet-incorporated injectable polypeptide-engineered hydrogel combinated with chemotherapy for 4T1 breast tumor therapy.

A multifunctional PC10A/DOX/MoS2 hydrogel was designed and prepared for chemotherapy/photothermal therapy/photodynamic therapy of 4T1 tumor, and the immune responses triggered by photothermal and photodynamic effect of MoS2 nanosheet in the hydrogel were also studied. Positively charged DOX and negatively charged PC10A were loaded on the surface of MoS2 nanosheet through layer-by-layer method to prepare hybrid PC10A/DOX/MoS2 nanoparticles. PC10A/DOX/MoS2 nanoparticles were dispersed in PC10A hydrogel to prepare PC10A/DOX/MoS2 hydrogel. 2D MoS2 nanosheet in the hydrogel was simultaneously utilized as photothermal agent and photodynamic agent for the generation of hyperthermia and reactive oxygen species, respectively. This PC10A/DOX/MoS2 hydrogel was injectable and possessed excellent biocompatibility. The results of in vivo tumor-bearing mice experiments showed that a remarkably enhance tumor inhibition was observed by the combination of chemo-photothermal-photodynamic therapy compared with photothermal therapy, photodynamic therapy, or chemotherapy alone. In addition, the results of in vivo therapy exhibited that the PC10A/DOX/MoS2 hydrogel with laser irradiation could activate antitumor immune effects to suppress the growth of primary 4T1 breast tumors and distal lung metastatic nodules. Therefore, these results demonstrated that the PC10A/DOX/MoS2 hydrogel was promising to be utilized in antitumor immunity therapy triggered by photothermal therapy and photodynamic therapy for malignant tumor.

Ion Current Rectification in High-Salt Environment from Mesoporous TiO2 Microplug in Situ Grown at the Tip of a Micropipette Induced by Space-Confined Evaporation.

In this work, in situ growth of a titanium dioxide microplug (TDMP) having mesoporous channels at the tip of a glass micropipette induced by space-confined evaporation is reported. Moreover, clear ion current rectification (ICR) of a single-material nanopore in a saturated potassium chloride solution is observed for the first time. TDMP presents an asymmetrical channel structure with the top and bottom apertures of 12.3 ± 6.1 and 42.6 ± 19.7 nm, respectively. TDMP exhibits outstanding ICR capability as the ions get transported through it due to the applied potential. The values for the rectification coefficient (r = log2|I+1 V/I-1 V|) in a saturated KCl solution under acidic (pH of 3.0) and alkaline (pH of 10.0) environments are 1.32 and -0.84, respectively. The intensity and direction of ICR can be adjusted by pH or through the modification of citric acid. Meanwhile, the length and ion transport behavior of TDMP under different growth conditions (time and diameter) were also investigated. TDMP with asymmetric mesoporous channels, maintaining ICR in a saturated salt solution, is expected to expand the application of nanopores in high-salt environments. Furthermore, growth of mesoporous material in the micropipette facilitates the miniaturization of the nanopore device, which further promotes its application potential.

Hollow gold nanoshells-incorporated injectable genetically engineered hydrogel for sustained chemo-photothermal therapy of tumor.

BACKGROUND: Combined therapy has demonstrated to be an effective strategy for cancer therapy. Herein, an injectable hydrogel based on the genetically engineered polypeptide and hollow gold nanoshells (HAuNS) has been developed for chemo-photothermal therapy of HepG2 tumor. METHODS: PC10A/DOX/HAuNS nanogel was prepared with layer-by-layer through the adsorption of DOX and PC10A successively. DOX with positive charge and PC10A with negative charge were coated step by step onto the surface of negatively charged HAuNS. The multifunctional hydrogel PC10A/DOX/HAuNS were prepared via dissolving hybrid PC10A/DOX/HAuNS nanogel in polypeptide PC10A. Chemotherapy drug DOX in the PC10A/DOX/HAuNS hydrogel was absorbed on the HAuNS and directly embedded in the PC10A hydrogel, which contributes to sequentially release of the drug. Specifically, DOX adsorbed on the HAuNS could be released slowly for sustainable chemotherapy. RESULTS: The PC10A/DOX/HAuNS hydrogel could pass 26-gauge needle without clogging, indicating that it is injectable. In addition, the PC10A/DOX/HAuNS hydrogel possessed outstanding photothermal effect and photothermal stability. In both in vitro cell and in vivo tumor-bearing mice experiments, a remarkably enhance tumor inhibition was observed by the combined therapy of chemo-photothermal therapy compared with photothermal therapy or chemotherapy alone. CONCLUSIONS: The combined chemotherapy and photothermal therapy of PC10A/DOX/HAuNS hydrogels could significantly improve the therapeutic effect. Therefore, the multifunctional hydrogel PC10A/DOX/HAuNS is promising to provide a new strategy for sustained chemo-photothermal therapy.

An injectable hybrid hydrogel based on a genetically engineered polypeptide for second near-infrared fluorescence/photoacoustic imaging-monitored sustained chemo-photothermal therapy.

An injectable multifunctional hydrogel based on an engineered coiled-coil polypeptide, Ag2S quantum dots (QDs), and paclitaxel (PTX) has been developed for sustained chemo-photothermal therapy. Oil-soluble Ag2S QDs and PTX were first loaded into nanogels formed with engineered polypeptide PC10A by ultrasonic treatment to prepare PC10A/Ag2S QD/PTX nanogels. The multifunctional PC10A/Ag2S QD/PTX hydrogels were prepared by dissolving the PC10A/Ag2S QD/PTX nanogels into the PC10A hydrogel. The PC10A/Ag2S QD/PTX hydrogel can be injected directly into the site of tumors. In vitro and in vivo toxicity results showed that the PC10A/Ag2S QD/PTX hydrogel presented excellent biocompatibility. Compared with single near-infrared photothermal therapy and chemotherapy, the combined therapy could effectively suppress the growth of SKOV3 ovarian carcinoma tumor. In addition, real-time monitoring of the in vivo degradation of the PC10A/Ag2S QD/PTX hydrogel was achieved by near-infrared fluorescence imaging and photoacoustic imaging. These results demonstrated that this injectable multifunctional PC10A/Ag2S QD/PTX hydrogel has the potential as a theranostic platform for sustained cancer treatments.