Autophagy modulation effect on homotypic transfer of intracellular components via tunneling nanotubes in mesenchymal stem cells.

BACKGROUND: Recent studies have proved the role of autophagy in mesenchymal stem cell (MSCs) function and regenerative properties. How and by which mechanism autophagy modulation can affect the juxtacrine interaction of MSCs should be addressed. Here, the role of autophagy was investigated in the formation of tunneling nanotubes (TNTs) and homotypic mitochondrial donation. METHODS: MSCs were incubated with 15 µM Metformin (Met) and/or 3 µM 3-methyladenine (3-MA) for 48 h. The formation of TNTs was assessed using bright-field and SEM images. The mitochondria density and ΔΨ values were monitored using flow cytometry analysis. Using RT-PCR and protein array, the close interaction and shared mediators between autophagy, apoptosis, and Wnt signaling pathways were also monitored. The total fatty acid profile was assessed using gas chromatography. RESULT: Data indicated the increase of TNT length and number, along with other cell projections after the induction of autophagy while these features were blunted in 3-MA-treated MSCs (p < 0.05). Western blotting revealed the significant reduction of Rab8 and p-FAK in 3-MA-treated MSCs (p < 0.05), indicating the inhibition of TNT assembly and vesicle transport. Likewise, the stimulation of autophagy increased autophagic flux and mitochondrial membrane integrity compared to 3-MA-treated MSCs. Despite these findings, protein levels of mitochondrial membrane Miro1 and 2 were unchanged after autophagy inhibition/stimulation (p > 0.05). We found that the inhibition/stimulation of autophagy can affect the protein, and transcription levels of several mediators related to Wnt and apoptosis signaling pathways involved in different cell bioactivities. Data confirmed the profound increase of mono and polyunsaturated/saturated fatty acid ratio in MSCs exposed to autophagy stimulator. CONCLUSIONS: In summary, autophagy modulation could affect TNT formation which is required for homotypic mitochondrial donation. Thus, the modulation of autophagy creates a promising perspective to increase the efficiency of cell-based therapies.

Transfer of cardiomyocyte-derived extracellular vesicles to neighboring cardiac cells requires tunneling nanotubes during heart development.

Rationale: Extracellular vesicles (EVs) are thought to mediate intercellular communication during development and disease. Yet, biological insight to intercellular EV transfer remains elusive, also in the heart, and is technically challenging to demonstrate. Here, we aimed to investigate biological transfer of cardiomyocyte-derived EVs in the neonatal heart. Methods: We exploited CD9 as a marker of EVs, and generated two lines of cardiomyocyte specific EV reporter mice: Tnnt2-Cre; double-floxed inverted CD9/EGFP and αMHC-MerCreMer; double-floxed inverted CD9/EGFP. The two mouse lines were utilized to determine whether developing cardiomyocytes transfer EVs to other cardiac cells (non-myocytes and cardiomyocytes) in vitro and in vivo and investigate the intercellular transport pathway of cardiomyocyte-derived EVs. Results: Genetic tagging of cardiomyocytes was confirmed in both reporter mouse lines and proof of concept in the postnatal heart showed that, a fraction of EGFP+/MYH1- non-myocytes exist firmly demonstrating in vivo cardiomyocyte-derived EV transfer. However, two sets of direct and indirect EGFP +/- cardiac cell co-cultures showed that cardiomyocyte-derived EGFP+ EV transfer requires cell-cell contact and that uptake of EGFP+ EVs from the medium is limited. The same was observed when co-cultiring with mouse macrophages. Further mechanistic insight showed that cardiomyocyte EV transfer occurs through type I tunneling nanotubes. Conclusion: While the current notion assumes that EVs are transferred through secretion to the surroundings, our data show that cardiomyocyte-derived EV transfer in the developing heart occurs through nanotubes between neighboring cells. Whether these data are fundamental and relate to adult hearts and other organs remains to be determined, but they imply that the normal developmental process of EV transfer goes through cell-cell contact rather than through the extracellular compartment.

Piggybacking functionalized DNA nanostructures into live-cell nuclei.

DNA origami nanostructures (DOs) are promising tools for applications including drug delivery, biosensing, detecting biomolecules, and probing chromatin substructures. Targeting these nanodevices to mammalian cell nuclei could provide impactful approaches for probing, visualizing, and controlling biomolecular processes within live cells. We present an approach to deliver DOs into live-cell nuclei. We show that these DOs do not undergo detectable structural degradation in cell culture media or cell extracts for 24 hours. To deliver DOs into the nuclei of human U2OS cells, we conjugated 30-nanometer DO nanorods with an antibody raised against a nuclear factor, specifically the largest subunit of RNA polymerase II (Pol II). We find that DOs remain structurally intact in cells for 24 hours, including inside the nucleus. We demonstrate that electroporated anti-Pol II antibody-conjugated DOs are piggybacked into nuclei and exhibit subdiffusive motion inside the nucleus. Our results establish interfacing DOs with a nuclear factor as an effective method to deliver nanodevices into live-cell nuclei.

The regulatory effect of TiO2 nanotubes loaded with graphene oxide on macrophage polarization in an inflammatory environment.

BACKGROUND: Excessive inflammation is a major cause of implant failure. The surface morphology, hydrophilicity, and loading of biomaterials are major properties modulating anti-inflammatory macrophage activation. This paper investigates the regulatory effects of modifying the surface of Titanium dioxide nanotubes (TNTs) with graphene oxide (GO) on the polarization of mouse monocyte macrophages (RAW264.7). METHODS: TNT was produced by the anodic oxidation of titanium. GO was subsequently electrodeposited on the TNT to obtain a TNT-GO composite. The samples were characterised through scanning electron microscopy (SEM), Raman spectroscopy, and X-ray diffraction. RAW264.7 cells were separately seeded onto the surface of three groups of samples: pure Ti, TNT, and TNT-GO. Under the condition of lipopolysaccharide stimulation, the influence of the sample surfaces on the gene expression profiles was investigated through RNA sequence analysis. In addition, cell spreading was observed through SEM, cell adhesion and proliferation were analysed using the CCK8 assay, and the expression of inflammation-related factors was investigated by ELISA and cellular immunofluorescence staining. The production of reactive oxygen species (ROS) in the RAW264.7 cells on the surface of the three groups was detected via immunofluorescence staining. RESULTS: The CCK8 results indicated that the adhesion and proliferation of the RAW264.7 cells were reduced on the TNT and TNT-GO surfaces. ELISA results revealed significant differences in the pro-inflammatory factors tumour necrosis factor-α and interleukin-6 secretion among the three groups at 24 h (p < 0.05). The secretion of pro-inflammatory factors significantly reduced and the expression of anti-inflammatory factor IL-10 increased on the TNT and TNT-GO surfaces. The RNA sequencing, ELISA, and cell immunofluorescence staining test results suggested that the inflammatory response of M1 polarization was reduced and the M2 polarization of macrophages was induced on the TNT-GO surface, which may be attributed to the reduction in ROS production. CONCLUSIONS: Under lipopolysaccharide stimulation, the inflammatory response of the RAW264.7 cells was reduced and the M2 polarization of macrophages was promoted on the TNT-GO surface, which may be caused by the reduced ROS production. Consequently, the designed TNT-GO material is promising for implants owing to its excellent inflammation regulation ability.

Ultrasensitive Photoelectrochemical Biosensor for Dual-miRNAs Detection Based on Molecular Logic Gates and Methylene Blue Sensitized ZnO@CdS@Au Nanorods.

The occurrence of cancer is often closely related to multiple tumor markers, so it is important to develop multitarget detection methods. By the proper design of the input signals and logical operations of DNA logic gates, detection and diagnosis of cancer at different stages can be achieved. For example, in the early stages, specific input signals can be designed to correspond to early specific tumor markers, thereby achieving early cancer detection. In the late stage, logic gates for multitarget detection can be designed to simultaneously detect multiple biomarkers to improve diagnostic accuracy and comprehensiveness. In this work, we constructed a dual-target-triggered DNA logic gate for anchoring DNA tetrahedra, where methylene blue was embedded in the DNA tetrahedra to sensitize ZnO@CdS@Au, achieving ultrasensitive detection of the target substance. We tested the response of AND and OR logic gates to the platform. For AND logic gates, the sensing platform only responds when both miRNAs are present. In the concentration range of 10 aM to 10 nM, the photoelectric signal gradually increases with an increase of the target concentration. Subsequently, we used OR logic gates for miRNA detection. Even if only one target exists, the sensing platform exhibits excellent performance. Similarly, within the concentration range of 10 aM to 10 nM, the photoelectric signal gradually increases with an increase of the target concentration. The minimum detection limit is 1.10 aM. Whether it is the need to detect multiple targets simultaneously or only one of them, we can achieve it by selecting the appropriate logic gate. This strategy holds promising application prospects in fields such as biosensing, medical diagnosis, and environmental monitoring.

Near-infrared-triggered plasmonic regulation and cardiomyocyte-based biosensing system for in vitro bradyarrhythmia treatment.

Bradyarrhythmia, a life-threatening cardiovascular disease, is an increasing burden for the healthcare system. Currently, surgery, implanted device, and drug are introduced to treat the bradyarrhythmia in clinical practice. However, these conventional therapeutic strategies suffer from the invasive surgery, power supply, or drug side effect, respectively, hence developing the alternative therapeutic strategy is necessarily imperative. Here, a convenient and effective strategy to treat the bradyarrhythmia is proposed using near-infrared-triggered Au nanorod (NR) based plasmonic photothermal effect (PPE). Moreover, electrophysiology of cardiomyocytes is dynamically monitored by the integrated biosensing-regulating system during and after the treatment. Cardiomyocyte-based bradyarrhythmia recover rhythmic for a long time by regulating plasmonic photothermal effect. Furthermore, the regulatory mechanism is qualitatively investigated to verify the significant thermal stimulation in the recovery process. This study establishes a reliable platform for long-term recording and evaluation of mild photothermal therapy for bradyarrhythmia in vitro, offering an efficient and non-invasive strategy for the potential clinical applications.

TiO2 nanotubes with customized diameters for local drug delivery systems.

In this study, we evaluated the drug release behavior of diameter customized TiO2 nanotube layers fabricated by anodization with various applied voltage sequences: conventional constant applied potentials of 20 V (45 nm) and 60 V (80 nm), a 20/60 V stepped potential (50 nm [two-diameter]), and a 20-60 V swept potential (49 nm [full-tapered]) (values in parentheses indicate the inner tube diameter at the top part of nanotube layers). The structures of the 50 nm (two-diameter) and 49 nm (full-tapered) samples had smaller inner diameters at the top part of nanotube layers than that of the 80 nm sample, while the outer diameters at the bottom part of nanotube layers were almost the same size as the 80 nm sample. The 80 nm sample, which had the largest nanotube diameter and length, exhibited the greatest burst release, followed by the 50 nm (two-diameter), 49 nm (full-tapered), and 45 nm samples. The initial burst released drug amounts and release rates from the 50 nm (two-diameter) and 49 nm (full-tapered) samples were significantly suppressed by the smaller tube top. On the other hand, the largest proportion of the slow released drug amount to the total released drug amount was observed for the 50 nm (two-diameter) sample. Thus, 50 nm (two-diameter) achieved suppressed initial burst release and large storage capacity. Therefore, this study has, for the first time, applied TiO2 nanotube layers with modulated diameters (two-diameter and full-tapered) to the realization of a localized drug delivery system (LDDS) with customized drug release properties.

Stable peptide-assembled nanozyme mimicking dual antifungal actions.

Natural antimicrobial peptides (AMPs) and enzymes (AMEs) are promising non-antibiotic candidates against antimicrobial resistance but suffer from low efficiency and poor stability. Here, we develop peptide nanozymes which mimic the mode of action of AMPs and AMEs through de novo design and peptide assembly. Through modelling a minimal building block of IHIHICI is proposed by combining critical amino acids in AMPs and AMEs and hydrophobic isoleucine to conduct assembly. Experimental validations reveal that IHIHICI assemble into helical ß-sheet nanotubes with acetate modulation and perform phospholipase C-like and peroxidase-like activities with Ni coordination, demonstrating high thermostability and resistance to enzymatic degradation. The assembled nanotubes demonstrate cascade antifungal actions including outer mannan docking, wall disruption, lipid peroxidation and subsequent ferroptotic death, synergistically killing >90% Candida albicans within 10 min on disinfection pad. These findings demonstrate an effective de novo design strategy for developing materials with multi-antimicrobial mode of actions.

Diverse Chiral Nanotubes Assembled from Identical DNA Strands.

DNA nanotubes with controllable geometries hold a wide range of interdisciplinary applications. When preparing DNA nanotubes of varying widths or distinct chirality, existing methods require repeatedly designing and synthesizing specific DNA sequences, which can be costly and laborious. Here, we proposed an intercalator-assisted DNA tile assembly method which enables the production of DNA nanotubes of diverse widths and chirality using identical DNA strands. Through adjusting the concentration of intercalators during assembly, the twisting direction and extent of DNA tiles could be modulated, leading to the formation of DNA nanotubes featuring controllable widths and chirality. Moreover, through introducing additional intercalators and secondary annealing, right-handed nanotubes could be reconfigured into distinct left-handed nanotubes. We expect that this method could be universally applied to modulating the self-assembly pathways of various DNA tiles and other chiral materials, advancing the landscape of DNA tile assembly.

Potentiality of phosphide-based nanotubes for breast cancer detection: A DFT investigation.

Breast cancer is one of the most basilisk cancers for women due to its high mortality rate which can be prevented drastically with early-stage detection. In this work, the adsorption mechanism of two volatile organic compounds that are present in the breath of breast cancer patients, 2-Methyloctane and 3, 3-Dimethylpentane, has been investigated on aluminum phosphide nanotubes (AlPNT) and gallium phosphide nanotubes (GaPNT) in order to understand their feasibility as sensor materials to diagnosis breast cancer at early stage. We have used the quantum mechanical approach by employing density functional theory using B3LYP-D3 hybrid potential for noncovalent interaction along with the LanL2DZ basis in the Gaussian 09 software package. The adsorption properties analyses suggest that GaPNT exhibits better sensing behavior as well as proclaims 12.6% greater adsorption energy for 2-Methyloctane and 9.4% greater adsorption energy for 3, 3-Dimethylpentane than AlPNT. Other structural and electric properties analyses satisfy this conclusion and suggest that GaPNT exhibits higher stability than AlPNT and could possibly be a potential candidate for developing biosensors to detect breast cancer at the preliminary stages.

Stimulus-Responsive Four-Stranded DNA Nanoring Assembly to Host Multiple Nanosilver Clusters for Cooperatively Enhanced Fluorescence Biosensing.

Exploring the ability of four-stranded DNA nanorings (fsDNRs) to host multiple nanosilver clusters (NAgCs) for cooperatively amplifiable fluorescence biosensing to a specific initiator (tI*) is fascinating. By designing three DNA single strands and three analogous stem-loop hairpins, we developed a functional fsDNR through sequential cross-opening and overlapped hybridization. Note that a substrate strand (SS) was programmed with six modules: two severed splits (sT and sT') of NAgCs template, two sequestered segments by a middle unpaired spacer, and a partition for tI*-recognizable displacement, while sT and sT' were also tethered in two ends of three hairpins. At first, a triple dsDNA complex with stimulus-responsiveness was formed to guide the specific binding to tI*, while the exposed toehold of the SS activated the forward cascade hybridization of three hairpins, until the ring closure in the tailored self-assembly pathway for forming the fsDNR. The resulting four duplexes forced each pair of sT/sT' to be merged as the parent template in four nicks, guiding the preferential synthesis of four clusters in the shared fsDNR, thereby cooperatively amplifying the green fluorescence signal for sensitive assay of tI*. Meanwhile, the topological conformation of fsDNR can be stabilized by the as-formed cluster adducts to rivet the pair of two splits in the nicks. Benefitting from the self-enhanced effect of multiple emitters, this label-free fluorescent sensing strategy features simplicity, rapidity, and high on-off contrast, without involving complicated nucleic acid amplifiers.

Polyethyleneimine-mediated assembly of DNA nanotubes for KRAS siRNA delivery in lung adenocarcinoma therapy.

Self-assembled DNA nanostructures hold great promise in biosensing, drug delivery and nanomedicine. Nevertheless, challenges like instability and inefficiency in cellular uptake of DNA nanostructures under physiological conditions limit their practical use. To tackle these obstacles, this study proposes a novel approach that integrates the cationic polymer polyethyleneimine (PEI) with DNA self-assembly. The hypothesis is that the positively charged linear PEI can facilitate the self-assembly of DNA nanostructures, safeguard them against harsh conditions and impart them with the cellular penetration characteristic of PEI. As a demonstration, a DNA nanotube (PNT) was successfully synthesized through PEI mediation, and it exhibited significantly enhanced stability and cellular uptake efficiency compared to conventional Mg2+-assembled DNA nanotubes. The internalization mechanism was further found to be both clathrin-mediated and caveolin-mediated endocytosis, influenced by both PEI and DNA. To showcase the applicability of this hybrid nanostructure for biomedical settings, the KRAS siRNA-loaded PNT was efficiently delivered into lung adenocarcinoma cells, leading to excellent anticancer effects in vitro. These findings suggest that the PEI-mediated DNA assembly could become a valuable tool for future biomedical applications.

A near-infrared plasmonic biosensor for detection of morphine and codeine in biological samples based on the end-to-end assembly of modified gold nanorods.

The analytical determination of opiates in biological samples is a critical mission and remains a challenge for almost all judicial and clinical drug testing panels due to their high abuse potential. Based on the high sensitivity of the longitudinal surface plasmon resonance (LSPR) peak of gold nanorods (AuNRs), we successfully developed a novel and simple refractive index sensing platform for detection of morphine (MOR) and codeine (COD) by means of 2-amino-5-mercapto-1,3,4-thiadiazole functionalized gold nanorods (AMTD-AuNRs) in aqueous solution, which is, to the best of our knowledge, the first report on the assay of MOR and COD using AuNRs. AMTD molecules strongly anchor onto the tips of AuNRs via the mercapto group and subsequent hydrogen-bonding interactions between AMTD and the analytes induced end-to-end chain assembly of AuNRs and a consequent decrease of the LSPR absorption band at 850 nm along with a bathochromic shift and emergence of a new hybridized plasmon mode at 1050 nm which was characterized using a Vis-NIR spectrophotometer. After systematic optimization, the absorbance ratio (A1050/A850) was proportional to the concentration of MOR in the ranges of 0.08-5 µM and 0.2-8 µM for COD without any significant effect from possible interferents. Furthermore, detection limits of 40 and 62 nM were achieved for MOR and COD, respectively, which are much lower than the cut-off level of 2000 ng mL-1 for opiates in urine samples set by the Substance and Abuse Mental Health Services Administration (SAMHSA). Eventually, as proof-of-applicability, human urine and blood serum samples spiked with MOR and COD were analyzed and excellent recoveries ranging from 94.4 to 108.9% were obtained, demonstrating the successful applicability of the designed refractive index probe in real biological specimens.

Urchin-like CoP3/Cu3P heterostructured nanorods supported on a 3D porous copper foam for high-performance non-enzymatic electrochemical dopamine sensors.

In this study, we developed a high-performance non-enzymatic electrochemical sensor based on urchin-like CoP3/Cu3P heterostructured nanorods supported on a three-dimensional porous copper foam, namely, CoP3/Cu3P NRs/CF, for the detection of dopamine. Benefiting from the promising intrinsic catalytic activities of CoP3 and Cu3P, urchin-like microsphere structures, and a large electrochemically active surface area for exposing numerous accessible catalytic active sites, the proposed CoP3/Cu3P NRs/CF shows extraordinary electrochemical response towards the electrocatalytic oxidation of dopamine. As a result, the CoP3/Cu3P NRs/CF sensing electrode has a broad detection window (from 0.2 to 2000 µM), low detection limit (0.51 µM), high electrochemical sensitivity (0.0105 mA µM-1 cm-2), excellent selectivity towards dopamine in the coexistence of some interfering species, and good stability for dopamine determination. More importantly, the CoP3/Cu3P NRs/CF catalyst also exhibits excellent catalytic activity, sensitivity, and selectivity for dopamine detection under simulated human body conditions at a physiological pH of 7.25 (0.1 M PBS) at 36.6 °C.

Titania (TiO2) nanotube surfaces doped with zinc and strontium for improved cell compatibility.

Titanium-based orthopedic implants are gaining popularity in recent years due to their excellent biocompatibility, superior corrosion resistance and lightweight properties. However, these implants often fail to perform effectively due to poor osseointegration. Nanosurface modification approaches may help to resolve this problem. In this work, TiO2 nanotube (NT) arrays were fabricated on commercially available pure titanium (Ti) surfaces by anodization and annealing. Then, zinc (Zn) and strontium (Sr), important for cell signaling, were doped on the NT surface by hydrothermal treatment. This very simple method of Zn and Sr doping takes less time and energy compared to other complicated techniques. Different surface characterization tools such as scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), energy-dispersive X-ray spectroscopy (EDS), static water contact angle, X-ray diffraction (XRD) and nanoindentation techniques were used to evaluate the modified surfaces. Then, adipose derived stem cells (ADSCs) were cultured with the surfaces to evaluate cell adhesion, proliferation, and growth on the surfaces. After that, the cells were differentiated towards osteogenic lineage to evaluate alkaline phosphatase (ALP) activity, osteocalcin expression, and calcium phosphate mineralization. Results indicate that NT surfaces doped with Zn and Sr had significantly enhanced ADSC adhesion, proliferation, growth, and osteogenic differentiation compared to an unmodified surface, thus confirming the enhanced performance of these surfaces.

Halloysite nanotubes enhanced polyimide/oxidized-lignin nanofiber separators for long-cycling lithium metal batteries.

The high energy density and robust cycle properties of lithium-ion batteries contribute to their extensive range of applications. Polyolefin separators are often used for the purpose of storing electrolytes, hence ensuring the efficient internal ion transport. Nevertheless, the electrochemical performance of lithium-ion batteries is constrained by its limited interaction with electrolytes and poor capacity for cation transport. This work presents the preparation of a new bio-based nanofiber separator by combining oxidized lignin (OL) and halloysite nanotubes (HNTs) with polyimide (PI) using an electrospinning technique. Analysis was conducted to examine and compare the structure, morphology, thermal characteristics, and EIS of the separator with those of commercially available polypropylene separator (PP). The results indicate that the PI@OL and PI-OL@ 10 % HNTs separators exhibit higher lithium ion transference number and ionic conductivity. Moreover, the use of HNTs successfully impeded the proliferation of lithium dendrites, hence exerting a beneficial impact on both the cycle performance and multiplier performance of the battery. Consequently, after undergoing 300 iterations, the battery capacity of LiFePO4|PI-OL@ 10 % HNTs|Li stays at 92.1 %, surpassing that of PP (86.8 %) and PI@OL (89.6 %). These findings indicate that this new bio-based battery separator (PI-OL@HNTs) has the great potential to serve as a substitute for the commonly used PP separator in lithium metal batteries.

In Situ Self-Growth of a ZnO Nanorod Array on Nonwoven Fabrics for Empowering Superhydrophobic and Antibacterial Features.

ZnO nanorod nonwoven fabrics (ZNRN) were developed through hydrothermal synthesis to facilitate the prevention of the transmission of respiratory pathogens. The superhydrophobicity and antibacterial properties of ZNRN were improved through the response surface methodology. The synthesized material exhibited significant water repellency, indicated by a water contact angle of 163.9°, and thus demonstrated antibacterial rates of 91.8% for Escherichia coli (E. coli) and 79.75% for Staphylococcus aureus (S. aureus). This indicated that E. coli with thinner peptidoglycan may be more easily killed than S. aureus. This study identified significant effects of synthesis conditions on the antibacterial effectiveness, with comprehensive multivariate analyses elucidating the underlying correlations. In addition, the ZnO nanorod structure of ZNRN was characterized through SEM and XRD analyses. It endows the properties of superhydrophobicity (thus preventing bacteria from adhering to the ZNRN surface) and antibacterial capacity (thus damaging cells through the puncturing of these nanorods). Consequently, the alignment of two such features is desired to help support the development of personal protective equipment, which assists in avoiding the spread of respiratory infections.

Plasmonics nanorod biosensor for in situ intracellular detection of gene expression biomarkers in intact plant systems.

The intracellular developmental processes in plants, particularly concerning lignin polymer formation and biomass production are regulated by microRNAs (miRNAs). MiRNAs including miR397b are important for developing efficient and cost-effective biofuels. However, traditional methods of monitoring miRNA expression, like PCR, are time-consuming, require sample extraction, and lack spatial and temporal resolution, especially in real-world conditions. We present a novel approach using plasmonics nanosensing to monitor miRNA activity within living plant cells without sample extraction. Plasmonic biosensors using surface-enhanced Raman scattering (SERS) detection offer high sensitivity and precise molecular information. We used the Inverse Molecular Sentinel (iMS) biosensor on unique silver-coated gold nanorods (AuNR@Ag) with a high-aspect ratio to penetrate plant cell walls for detecting miR397b within intact living plant cells. MiR397b overexpression has shown promise in reducing lignin content. Thus, monitoring miR397b is essential for cost-effective biofuel generation. This study demonstrates the infiltration of nanorod iMS biosensors and detection of non-native miRNA 397b within plant cells for the first time. The investigation successfully demonstrates the localization of nanorod iMS biosensors through TEM and XRF-based elemental mapping for miRNA detection within plant cells of Nicotiana benthamiana. The study integrates shifted-excitation Raman difference spectroscopy (SERDS) to decrease background interference and enhance target signal extraction. In vivo SERDS testing confirms the dynamic detection of miR397b in Arabidopsis thaliana leaves after infiltration with iMS nanorods and miR397b target. This proof-of-concept study is an important stepping stone towards spatially resolved, intracellular miRNA mapping to monitor biomarkers and biological pathways for developing efficient renewable biofuel sources.

Scalable drop-casting construction of light-addressable photoelectrochemical biosensor on laser-induced graphene electrode arrays for high-throughput drug screening.

A drop-casting method for the scalable construction of a solar cell-type light-addressable photoelectrochemical (PEC) sensor on commercial phenol resin (PR) plates is reported. The sensor was fabricated by laser writing of addressable laser-induced graphene (LIG) electrode arrays on PR plates with ring-disc dual-electrode cell configurations using a 405 nm laser machine. Beneficial from the good hydrophilicity of PR-based LIG and the excellent film formation of bismuth sulfide nanorods (Bi2S3 NRs), uniform Bi2S3 photovoltaic films can be reproducibly deposited onto the LIG disc photoanode array via drop casting modification, which show a sensitive photocurrent response toward thiocholine (TCl) when the ring cathode array was coated with Ag/AgCl. An acetylcholinesterase (AChE)-based PEC biosensor was therefore constructed by a similar drop-casting modification method. The resulting biosensor exhibits good sensitivity toward an AChE inhibitor, i.e., galantamine hydrobromide (GH), with a calibration range of 10-300 µM and a detection limit of 7.33 µM (S/N = 3). Moreover, the biosensor possesses good storage stability, which can achieve the high-throughput screening of AChE inhibitor drugs from traditional Chinese medicines (TCMs). The present work thus demonstrates the promising application of LIG technology in constructing light-addressable PEC sensing devices with high performance and low cost.

Membrane nanotubes transform into double-membrane sheets at condensate droplets.

Cellular membranes exhibit a multitude of highly curved morphologies such as buds, nanotubes, cisterna-like sheets defining the outlines of organelles. Here, we mimic cell compartmentation using an aqueous two-phase system of dextran and poly(ethylene glycol) encapsulated in giant vesicles. Upon osmotic deflation, the vesicle membrane forms nanotubes, which undergo surprising morphological transformations at the liquid-liquid interfaces inside the vesicles. At these interfaces, the nanotubes transform into cisterna-like double-membrane sheets (DMS) connected to the mother vesicle via short membrane necks. Using super-resolution (stimulated emission depletion) microscopy and theoretical considerations, we construct a morphology diagram predicting the tube-to-sheet transformation, which is driven by a decrease in the free energy. Nanotube knots can prohibit the tube-to-sheet transformation by blocking water influx into the tubes. Because both nanotubes and DMSs are frequently formed by cellular membranes, understanding the formation and transformation between these membrane morphologies provides insight into the origin and evolution of cellular organelles.