Assessing Phytogenic and Chemogenic Silver Nanoparticles for Antibacterial Activity and Expedited Wound Recuperation.

Green silver nanoparticles (AgNPs) possess tremendous promise for diverse applications due to their versatile characteristics. Coriander and other plant extracts have become popular for greenly synthesizing AgNPs as an economical, biocompatible, cost-effective, and environmentally beneficial alternative to chemical processes. In this study, we synthesized AgNPs from coriander leaves and evaluated their antibacterial, anti-inflammatory, antioxidant, and wound-healing acceleration properties in comparison to chemically synthesized AgNPs. The zeta potentials of AgNPs extracted from green and chemical processes were -32.4 mV and -23.4 mV, respectively. TEM images showed a cuboidal shape of green and chemical AgNPs with a diameter of approximately 100 nm. The FTIR spectra of green AgNPs showed an extreme absorption peak at 3401 cm-1, which signifies O-H stretching vibrations, typically linked to hydroxyl groups. In vitro results elaborated that AgNPs from coriander exerted a stronger effect on anti-Klebsiella pneumoniae (KP) through interrupting cell integrity, generating ROS, depleting ATP, and exhibiting significant antioxidant activity, compared with AgNPs synthesized chemically. In vivo experiments showed that AgNPs from coriander, as opposed to chemically manufactured AgNPs, greatly accelerated the healing of wounds contaminated with Klebsiella pneumoniae bacteria by effectively eliminating the bacteria on the wounds and stimulating skin regeneration and the deposition of dense collagen. In vivo assays further demonstrated that green AgNPs effectively enhanced Klebsiella pneumoniae-infected wound healing by extenuating local inflammatory responses and up-regulating VEGF and CD31 expression. In conclusion, green AgNPs significantly alleviated the inflammation without significantly harming the organism.

Biomimetic Prussian blue nanocomplexes for chemo-photothermal treatment of triple-negative breast cancer by enhancing ICD.

Drug-induced immunogenic cell death (ICD) can efficiently inhibit tumor growth and recurrence through the release of tumor-associated antigens which activate both local and systemic immune responses. Pyroptosis has emerged as an effective means for inducing ICD; however, the development of novel pyroptosis inducers to specifically target tumor cells remains a pressing requirement. Herein, we report that Cinobufagin (CS-1), a main ingredient of Chansu, can effectively induce pyroptosis of triple-negative breast cancer (TNBC) cells, making it a potential therapeutic agent for this kind of tumor. However, the application of CS-1 in vivo is extremely limited by the high dosage/long-term usage and non-selectivity caused by systemic toxicity. To address these drawbacks, we developed a new nanomedicine by loading CS-1 into Prussian blue nanoparticles (PB NPs). The nanomedicine can release CS-1 in a photothermal-controlled manner inherited in PB NPs. Furthermore, hybrid membrane (HM) camouflage was adopted to improve the immune escape and tumor-targeting ability of this nanomedicine, as well. In vitro assays demonstrated that the chemo-photothermal combination treatment produced high-level ICD, ultimately fostering the maturation of dendritic cells (DCs). In vivo anti-tumor assessments further indicated that this strategy not only efficiently inhibited primary growth of MDA-MB-231 cells and 4T1 cells-bearing models but also efficiently attenuated distant tumor growth in 4T1 xenograft model. This was mechanistically achieved throuh the promotion of DCs maturation, infiltration of cytotoxic T lymphocyte into the tumor, and the inhibition of Treg cells. In summary, this work provides a novel strategy for efficient TNBC therapy by using nanomaterials-based multimodal nanomedicine through rational design.

Phyto-Synthesis, Characterization, and In Vitro Antibacterial Activity of Silver Nanoparticles Using Various Plant Extracts.

Aloe vera, Mentha arvensis (mint), Coriandrum sativum (coriander), and Cymbopogon citratus (lemongrass) leaf extracts were used to synthesize stable silver nanoparticles (Ag-NPs) by green chemistry. UV-vis spectrophotometry, X-ray diffraction (XRD), thermogravimetric analysis (TGA), scanning electron microscopy (SEM), and energy dispersive X-ray (EDX) spectroscopy techniques were used to characterize these biosynthesized nanoparticles. The data indicated that the silver nanoparticles were successfully synthesized, and the narrower particle size distribution was at 10-22 nm by maintaining a specific pH. As a short-term post-sowing treatment, Ag-NP solutions of different sizes (10 and 50 ppm) were introduced to mung bean seedlings, and the overall increase in plant growth was found to be more pronounced at 50 ppm concentration. The antibacterial activity of Ag-NPs was also investigated by disc diffusion test, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) test. The zones of inhibition (ZOI) were shown by Escherichia coli (E. coli) (1.9, 2.1, 1.7, and 2 mm), followed by Staphylococcus aureus (S. aureus) (1.8, 1.7, 1.6, and 1.9 mm), against coriander, mint, Aloe vera, and lemongrass, respectively. MIC and MBC values of E. coli, and S. aureus ranged from 7 to 8 µg/mL. Overall, this study demonstrates that Ag-NPs exhibit a strong antimicrobial activity and thus might be developed as a new type of antimicrobial agent for the treatment of bacterial infection.

Biomimetic nanoparticles loading with gamabutolin-indomethacin for chemo/photothermal therapy of cervical cancer and anti-inflammation.

A pro-nanodrug combinational strategy for efficient cervical cancer therapy with intrinsic tumor microenvironment (TME)-responsive elements and low side effects is highly desired. Here, a pro-nanodrug complexes with GSH and NIR responsive manner is reported to boost gamabufotalin induced chemo-photothermal therapy with the assistance of reprogrammed TME by indomethacin. In addition, hybrid cell membrane was used to endow nanocomplexes with the prolonging circulation time and high accumulation of drug at tumor tissue. Indomethacin activated by the high level GSH can attenuate tumor inflammation microenvironment triggered by PTT and sensitize tumor cells to gamabufotalin through inhibiting PGE2 secretion. The released low-dose gamabufotalin with low side effects can efficiently kill tumor cells by ROS production and COX-2 low expression. In vitro and in vivo assays demonstrated that strong anti-tumor activity of nanocomplexes in tumor-bearing mice through chemo-photothermal therapy, which was reflected by the eradication of cervical tumor and significant extension of survival time of mice.

Real-time monitoring and effector screening of APE1 based on rGO assisted DNA nanoprobe.

Human apurinic/pyrimidine endonuclease 1 (APE1) played a critical role in the occurrence, progress and prognosis of tumors through overexpression and subcellular localization. Thus, it has become an important target for enhancing the sensitivity of tumor cells to radiotherapy and chemotherapy. Therefore, detecting and imaging its intracellular activity is of great significance for inhibitor discovery, cancer diagnosis and therapy. In this work, using DNA-based nanoprobe, we developed a new method for monitor intracellular APE1 activity. The detecting system was consisted by single fluorophore labeled hairpin probe and reduced graphene oxide (rGO). The in vitro result showed that a liner response of the detection method ranged from 0.02 U/mL to 2 U/mL with a limit of detection of 0.02 U/mL. Furthermore, this strategy possessing high specificity was successfully applied for APE1-related inhibitor screening using intracellular fluorescence imaging. Panaxytriol, an effective inhibitor of APE1 activity, was screened from traditional Chinese medicine (TCM) and its effect on APE1 activity was monitored in real time in A549 cells. In summary, this sensitive and specific APE1 detection technology is expected to provide an assistance for APE1-related inhibitor screening and diseases diagnosis.

Biosafety and biocompatibility assessment of Prussian blue nanoparticles in vitro and in vivo.

Aim: To investigate the effects of the different morphological characteristics of Prussian blue nanoparticles (PB NPs) on their biocompatibility and biosafety. Materials & methods: PB NPs with different sizes, shapes and charges were synthesized and their biosafety and biocompatibility performance were systematically compared in vitro and in vivo. Results: Increased size and positive charge of PB NPs adversely affected cell viability, while improving their peroxidase activity and photothermal conversion efficiency. In vivo analysis demonstrated good biocompatibility of PB NPs, without retention in the organs, but increased size retarded their metabolism. Meanwhile, increased size and positive charge adversely affected hepatic and renal function. Conclusion: This comprehensive exploration of biosafety and biocompatibility provides strong evidences for the use of PB NPs as nanodrug carrier and/or imaging agent.

DNAzyme and rGO based fluorescence assay for Fpg activity analysis, drug screening, and bacterial imaging.

Due to the significant role of formamidopyrimidine DNA glycosylase (Fpg) in physiological processes and DNA oxidative damage-related diseases, it is essential to establish sensitive methods for monitoring the Fpg activity in vitro and in vivo so as to illustrate its concrete role in these events. In this work, a sensitive, simple and reliable fluorescence assay was developed by taking the advantages of DNAzyme assisted cascade signal amplification and ultra-high fluorescence quenching efficiency of reduced graphene oxide (rGO). This detection system consisted of DNAzyme, rGO and fluorescence probe allows the activity of Fpg to be detected in a linear range from 0 to 80 U/mL with a detection limit of 0.66 U/mL. With the help of this method, 11 natural compounds were screened, and 7 compounds were identified as activators of Fpg. More importantly, the developed assay was used to monitor the activity of Fpg through fluorescence imaging in living Escherichia coli for the first time. The imaging results visually demonstrated the dynamic activation effect of natural compound Ginsenoside Re on the Fpg of Escherichia coli. In summary, these results indicated that this DNAzyme and rGO based fluorescence assay provides a potent strategy for Fpg quantitative assay in vitro and real-time monitoring in living bacteria, which holds great potential for applying on biological study and Fpg-targeted drug screening.

Sensitive RNase A detection and intracellular imaging using a natural compound-assisted tetrahedral DNA nanoprobe.

We report an intracellular imaging and assay nanoplatform for RNase A using a DNA tetrahedron-based fluorescent probe as a substrate. Importantly, a natural compound was used as an RNase A activity stimulator to improve the sensitivity. This platform provides an alternative for the diagnosis and prognosis of RNase A-related diseases and drug screening.

Endogenous Cys-Assisted GSH@AgNCs-rGO Nanoprobe for Real-Time Monitoring of Dynamic Change in GSH Levels Regulated by Natural Drug.

Glutathione (GSH) levels are closely related to the homeostasis of redox state which directly affects human disease occurrence by regulating cell apoptosis. Hence, real-time monitoring of dynamic changes in intracellular GSH levels is urgently needed for disease early diagnosis and evaluation of therapy efficiency. In this study, an endogenous cysteine (Cys)-assisted detection system based on GSH@AgNCs and reduced graphene oxide (rGO) with high sensitivity and specificity was developed for GSH detection. Compared with GSH, GSH@AgNCs with weaker affinity and bonding force was quite easier to extrude from the rGO surface when competing against GSH, leading to the obvious change in fluorescence signal. This phenomenon was termed as "a crowding out effect". Furthermore, the presence of Cys can improve GSH assay sensitivity by enhancing the quenching efficiency of rGO on the GSH@AgNCs. In vitro assay indicated that the efficiency of fluorescence recovery was positively related with GSH concentration in the range from 0 to 10 mM. In addition, the method was employed for real-time monitoring of the dynamic changes in GSH levels regulated by natural drugs. The imaging results showed that the natural compound 3 (C3) can downregulate GSH levels in HepG2 cells, which was accompanied by reactive oxygen species (ROS) release and apoptosis induction. Finally, the method was used to monitor the change of GSH levels in serum samples with chronic hepatitis B (CHB) infection. The results demonstrated that the occurrence and development of CHB may be positively correlated with GSH levels to some extent. Overall, the above results demonstrate the potential application of this new nanosystem in anticancer natural drug screening and clinical assay regarding GSH levels.

Daptomycin and AgNP co-loaded rGO nanocomposites for specific treatment of Gram-positive bacterial infection in vitro and in vivo.

In order to improve the stability of AgNPs and decrease the dosage of Daptomycin for killing bacteria, a reduced graphene oxide (rGO) was used for simultaneously anchoring AgNPs and Daptomycin to prepare rGO@Ag@Dap nanocomposites. In vitro experiments showed that the nanocomposites can efficiently kill four kinds of pathogenic bacteria, especially two kinds of Gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis) through damaging cell integrity, producing ROS, decreasing ATP and GSH and disrupting bacterial metabolism. Against Gram-positive bacteria, the rGO@Ag@Dap nanocomposites showed a cooperative antibacterial effect. Moreover, in vivo experiments showed that rGO@Ag@Dap can improve the healing of wounds infected with bacteria by efficiently killing the bacteria on the wounds and further promoting skin regeneration and dense collagen deposition. In summary, the above results suggest that the cooperative function of AgNPs with Daptomycin can significantly improve antibacterial efficiency against infectious diseases caused by bacteria, especially for therapies made ineffective due to the drug resistance of pathogenic bacteria.

RBC membrane camouflaged prussian blue nanoparticles for gamabutolin loading and combined chemo/photothermal therapy of breast cancer.

Due to the non-targeted release of anti-cancer agent gamabufotalin (CS-6), conventional chemotherapy using this drug can cause serious side effects, which accordingly result in poor therapeutic efficiency. Recently, the development of smart nanodrug systems has attracted more and more attention due to their significant advantages of high loading efficiency, controllable release behavior and targeted accumulation at tumor sites. In this study, a nanodrug system named as HA@RBC@PB@CS-6 NPs (HRPC) was constructed. In this system, Prussian blue nanoparticles (PB NPs) with hollow porous structure were used as the carrier for CS-6 and photothermal sensitizer simultaneously. The result indicated that the encapsulation of erythrocyte membrane on the PB NPs prolonged the blood circulation life to 10 h and improved the immune evasion ability for more than 60%, as well, which is beneficial for the targeting molecule (HA) to achieve high concentration accumulation of HRPCs at tumor sites. Moreover, we also disclosed that loading drug of CS-6 performed its ultra-strong anti-tumor function partly through markedly suppressing the expression of HSP70, which conversely amplified the efficiency of photothermal therapy. The in vivo study demonstrated the outstanding performance of HRPC in synergistic photothermal/chemotherapy of cancer without side effect to normal tissues.

A novel fluorescence method for activity assay and drug screening of T4 PNK by coupling rGO with ligase reaction.

T4 polynucleotide kinase (PNK) is the primary member of the 5'-kinase family that can transfer the γ-phosphate residue of ATP to the 5'-hydroxyl group of oligonucleotides. In this article, using the differential quenching ability of reduced graphene oxide (rGO) towards the fluorophore-labeled DNA probe, we propose a novel method for detecting T4 PNK activity assisted by ligase reaction. Under the optimized conditions, the detection limit of T4 PNK was estimated to be 0.0002 U µL-1 in the linear region of 0.001 U µL-1-0.1 U µL-1. Additionally, the developed method was used to screen regulators of T4 PNK from natural compounds. The compound f isolated from the root of Kadsura coccinea (Lem.) A.C. Smith was found to stimulate T4 PNK activity in a concentration-dependent manner in vitro. Finally, the method was used to monitor the relation of T4 PNK activity with pelvic inflammatory disease (PID). The results demonstrated that the development of this disease could inhibit T4 PNK activity to some extent. In summary, the above data indicate that the method not only provides a universal platform for monitoring T4 PNK activity, but also shows great potential to be used in drug screening and clinic diagnosis.

An erythrocyte membrane coated mimetic nano-platform for chemo-phototherapy and multimodal imaging.

The tumor variability and low efficiency associated with conventional chemical drugs provide an impetus to develop drug-carrying systems with targeted accumulation and controllable release behavior. Herein, DOX-loaded Prussian blue (PB) nano-composites are developed after coating with erythrocyte membrane (EM) and modifying with folic acid (FA). In these nano-composites, PB nanoparticles mixture with different shapes were adopted to improve the photothermal performance, which is highly helpful for cancer photothermal ablation and controllable drug release. In addition, the nano-composites were endowed with high biocompatibility, immune evading capacity, pH-/photo-responsive release behavior, and markedly prolonged blood circulation time, which was reflected by a 99.6% cervical tumor growth inhibition value (TGI) in vivo. Meanwhile, they functioned as multimodal bioimaging agents for photothermal, fluorescence, and photoacoustic imaging of tumors. The reported strategy can be applied for personalized therapy of various tumors by modifying the tumor-targeting molecule on the surface of nanoparticles.

The Role of Iron Competition in the Antagonistic Action of the Rice Endophyte Streptomyces sporocinereus OsiSh-2 Against the Pathogen Magnaporthe oryzae.

Rice blast caused by Magnaporthe oryzae severely impacts global rice yield stability. The rice endophyte Streptomyces sporocinereus OsiSh-2, with strong antagonistic activity towards M. oryzae, has been reported in our previous study. To decipher the model of the antagonistic action of OsiSh-2 towards M. oryzae, we compared the iron-capturing abilities of these two strains. The cultivation of OsiSh-2 and a M. oryzae strain under iron-rich and iron-starved conditions showed that M. oryzae depended more on iron supplementation for growth and development than did OsiSh-2. Genomic analysis of the S. sporocinereus and M. oryzae species strains revealed that they might possess different iron acquisition strategies. The actinobacterium OsiSh-2 is likely to favor siderophore utilization compared to the fungus M. oryzae. In addition, protein annotations found that OsiSh-2 contains the highest number of the siderophore biosynthetic gene clusters among the 13 endophytic actinomycete strains and 13 antifungal actinomycete strains that we compared, indicating the prominent siderophore production potential of OsiSh-2. Additionally, we verified that OsiSh-2 could excrete considerably more siderophores than Guy11 under iron-restricted conditions and displayed greater Fe3+-reducing activity during iron-supplemental conditions. Measurements of the iron mobilization between the antagonistic OsiSh-2 and Guy11 showed that the iron concentration is higher around OsiSh-2 than around Guy11. In addition, adding iron near OsiSh-2 could decrease the antagonism of OsiSh-2 towards Guy11. Our study revealed that the antagonistic capacity displayed by OsiSh-2 towards M. oryzae was related to the competition for iron. The highly efficient iron acquisition system of OsiSh-2 may offer valuable insight for the biocontrol of rice blast.

Sensitive Detection of RNase A Activity and Collaborative Drug Screening Based on rGO and Fluorescence Probe.

In addition to being an important object in theoretical and experimental studies in enzymology, RNase A also plays an important role in the development of many kinds of diseases by regulating various physiological or pathological processes, including cell growth, proliferation, differentiation, and invasion. Thus, it can be used as a useful biomarker for disease theranostics. Here, a simple, sensitive, and low-cost assay for RNase A was constructed by combining a fluorogenic substrate with reduced graphene oxide (rGO). The method with detection limit of 0.05 ng/mL was first applied for RNase A targeted drug screening, and 14 natural compounds were identified as activators of this enzyme. Then, it was applied to detect the effect of drug treatment and Hepatitis B virus (HBV) infection on RNase A activity. The results indicated that RNase A level in tumor cells was upregulated by G-10 and Chikusetsusaponin V in a concentration-dependent manner, while the average level of RNase A in the HBV infection group was significantly inhibited compared with that in the control group. Furthermore, the concentration-dependent inhibitory effect of heavy metal ions on RNase A was observed using the method and the results indicated that Ba2+, Co2+, Pb2+, As3+, and Cu2+ inhibited RNase A activity with IC50 values of 93.7 µM (Ba2+), 90.9 µM (Co2+), 110.6 µM (Pb2+), 171.5 µM (As3+), and 165.1 µM (Cu2+), respectively. In summary, considering the benefits of rapidity and high sensitivity, the method is practicable for RNase A assay in biosamples and natural compounds screening in vitro and in vivo.

PEGylated mBPEI-rGO nanocomposites facilitate hepotocarcinoma treatment combining photothermal therapy and chemotherapy.

Despite chemotherapy has been widely used for tumor therapy, the serious side effect is still a major challenge. Recently, two dimensional nanomaterial-based drug delivery systems have attracted wide concern due to their high drug loading and low side effect. In addition, some kinds of nanomaterials can directly act as a photosensitizer to induce cancer destruction. In this study, we developed a drug delivery system of mixture of high/low molecular weight branched polyethylenimine-polyethylene glycol-reduced graphene oxide (mBPEI-PEG-rGO) using reduced graphene oxide as matrix. A model drug of doxorubicin (DOX) was loaded on the nanocomposites with the efficiency of 81% and the release rate of more than 50% at acidic environment. In vitro experiments indicated that mBPEI-PEG-rGO-DOX with enhanced stability and biocompatibility efficiently delivered and released DOX into cells mainly through micropinocytosis and killed SMMC-7721 cells by inducing reactive oxygen species (ROS) and cell apoptosis. Furthermore, in vivo experiments indicated that the combination of intratumoral injection of mBPEI-PEG-rGO-DOX and local laser irradiation nearly ablated hepatocarcinoma. In conclusion, this new drug delivery system provided an alternative for combinational photothermal and chemotherapy against hepatocarcinoma.

An effective thermal therapy against cancer using an E-jet 3D-printing method to prepare implantable magnetocaloric mats.

Magnetic hyperthermia has been rapidly developed as a potential cancer treatment in recent years. Artificially induced hyperthermia close to a tumor can raise the temperature to 45°C causing tumor cell death. Herein, we introduce a novel method for rapid preparation of anti-cancer magnetocaloric PCL/Fe3 O4 mats capable of high-performance hyperthermia using E-jet 3D printing technology. Our 3D printed mats not only maintained the heating efficiency of traditional techniques for magnetic hyperthermia but also prolonged the effective therapy in vivo. When Fe3 O4 nanoparticles (NPs) were used in mats at a concentration of 6 mmol/L, 0.07 g PCL/Fe3 O4 mats were able to increase the temperature peripherally to 45°C under an alternating magnetic field (AMF) within 45 min. Moreover, the reproducibility experiment indicated that the maximum temperature was achieved following repeated heating and cooling cycles. Cell toxicity tests showed a high cell death rate during one treatment cycle. In vivo experiments indicated clear signs of tumor growth inhibitory and prolonged survival time of tumor-bearing mice after 4 weeks of treatment. The present magnetic mats may be a potential candidate for a novel heat-generating substrate for localized hyperthermia cancer therapy. Furthermore, the main advantage of such implantable magnetic mats is the local and precise delivery of Fe3 O4 NPs, ideal for the hyperthermia treatment of easily accessible tumors. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1827-1841, 2018.