ACS Applied Bio Materials最新文献

For activity-based sensing, where probes detect biological analytes through chemical reactivity, nucleic acid (NA) scaffolds represent an attractive platform to enhance biocompatibility and permit ratiometric analyte detection through the precise control of distance and orientation of donor/acceptor probes within the duplex framework for a turn-on FRET response. Herein, we present the first all-in-one NA FRET sensor (KR28) that is activated by H₂O₂ as a molecular trigger in human serum and in the nucleus of live cells. The 28-mer 2′-OMe-XNA hairpin (HP) KR28 features a boronic acid nucleobase surrogate (BA6HI) that serves as the H₂O₂ sensing element and is embedded into KR28 using an on-strand Aldol condensation approach. ipso-Hydroxylation of the BA6HI surrogate mediated by H₂O₂ furnishes an emissive phenolic product (PhOH6HI, λ_ex/λ_em = 390/490 nm) that serves as the FRET donor to a thiophene surrogate acceptor (Th6HI, λ_ex/λ_em = 530/580 nm) for a FRET efficiency ∼ 95%. The BA6HI-modified NA scaffolds can detect H₂O₂ in the low nM regime and localize in the nucleus of live cells for nuclear H₂O₂ FRET detection. Our work expands the function of NA scaffolds beyond their use as molecular recognition elements through noncovalent affinity interactions and demonstrates their potential to serve as activity-based sensors of ROS to probe the relationships between nuclear oxidative stress and disease states.

Extracellular vesicle (EV)-mediated transfer of biomolecules plays an essential role in intercellular communication and presents promising avenues for targeted drug delivery. Over the past decade, researchers have developed various approaches to modifying EV surfaces for targeting specific cells or tissues, including functionalization with targeting peptides to increase the specificity of drug delivery. Due to technical limitations, methods for characterizing the targeting moieties on the surface of small EVs (sEVs) are considerably restricted. To address these limitations and enhance the throughput capacity of sEV characterization, a dual-reporter platform was utilized to quantitatively assess the binding of tumor homing peptide (THP)-functionalized sEVs to breast cancer cells using bioluminescence assays and fluorescence microscopy. Twenty-four scrambled variants of the uPAR-binding peptide were designed for sEV engineering, and their uptake by MDA-MB-231 cells was evaluated in vitro. Our results revealed that amino acid scrambling generated both enhanced and reduced binding to cancer cells compared to the original peptide sequence. Furthermore, the data demonstrated that mechanical stimulation of EV producer HEK293FT cells enhanced the passive loading of methotrexate (MTX) into sEVs, but not large EVs, by increasing sEV production. By functionalizing MTX-loaded sEVs with a high-binding scrambled peptide, the delivery successfully surpassed the saturated free MTX uptake level in MDA-MB-231 cells, increasing cytotoxicity by 2.1-fold and providing a potent strategy for combating drug-resistant cancers. This study advances synthetic biology approaches to optimize tumor-targeted drug delivery, demonstrating that strategic peptide sequence scrambling can enhance targeting efficiency and drug delivery capabilities.

The rising prevalence of multidrug-resistant (MDR) bacterial infections highlights the urgent need for innovative antimicrobial therapies that are both effective and industrially scalable. We report a metal-free, facilely prepared, and biocompatible nanoplatform (TCPP-HF@Lec) designed for synergistic antibacterial treatment via light-triggered codelivery of singlet oxygen (¹O₂) and carbon monoxide (CO). This system coencapsulates the photosensitizer tetrakis(4-carboxyphenyl)porphyrin (TCPP) and the CO-releasing prodrug 3-hydroxyflavone (3-HF) within a lecithin-based nanoparticle, enabling straightforward preparation with potential for large-scale pharmaceutical production. Upon 660 nm irradiation, TCPP generates ¹O₂, which not only induces photodynamic cytotoxicity but also activates 3-HF via oxidative decarbonylation, achieving spatiotemporally controlled CO release. Comprehensive physicochemical characterization revealed uniform morphology, excellent colloidal stability, and efficient dual-agent loading. In vitro, TCPP-HF@Lec exhibited potent, concentration-dependent antibacterial activity against Gram-positive strains, including Staphylococcus aureus TISTR1466, Bacillus subtilis TISTR008, and methicillin-resistant Staphylococcus aureus (MRSA), achieving near-complete eradication under light irradiation. In contrast, Gram-negative bacteria (Escherichia coli TISTR780 and Pseudomonas aeruginosa TISTR781) showed negligible susceptibility, consistent with fluorescence imaging that revealed preferential nanoparticle uptake by Gram-positive bacteria lacking an outer membrane. By combining precise light-controlled activation with a production-friendly design, this dual-modality nanoplatform overcomes major limitations of conventional photodynamic therapy and CO-releasing molecules. TCPP-HF@Lec offers a promising approach for the scalable development of next-generation targeting antibacterial nanotherapeutics against MDR bacterial infections.

This article presents the synthesis and comprehensive investigation of the static and dynamic magnetization properties of large-scale PEG-400-coated superparamagnetic Zn_xMn_1–xFe₂O₄ (0 ≤ × ≤ 0.8) nanoparticles, highlighting their potential use as magnetic carriers in magnetic fluid hyperthermia (MFH). The Rietveld analysis of the X-ray diffraction spectra confirmed that the surface-functionalized core nanoparticles exhibit a single-phase spinel structure at the nanoscale, ranging from 15.4 to 11.2 nm. The use of PEG-400 as a hydrophilic shell over Mn–Zn ferrite enhances colloidal stability, as evidenced by the elevated zeta potential (ζ) values ranging from −40 to −26 mV. This enhancement reflects an increase in electrostatic repulsion among Zn_xMn_1–xFe₂O₄ nanoparticles, making them well-suited for formulating viscoelastic and water-based magnetic fluids for hyperthermia applications. The AC inductive heating efficiency for magnetic hyperthermia was systematically investigated as a function of particle concentration, applied alternating magnetic field (AMF), and radiofrequency. To optimize hyperthermic performance, AMF strengths of 9.5 kA/m, 18.3 kA/m, and 25.4 kA/m were applied at corresponding constant frequencies of 586.4 kHz, 154.8 kHz, and 103 kHz, using nanoparticle concentrations of 3 and 6 mg/mL. The optimal heating performance, with maximum specific absorption rate and intrinsic loss power (ILP) values of 273.4 W/g and 5.271 nHm²/kg, respectively, was achieved at 18.3 kA/m and 154.8 kHz for a 3 mg/mL concentration, which was comparable to clinically approved magnetic hyperthermia fluids (ILP range: 0.15–3.1 nHm²/kg). At the highest tested frequency (586.4 kHz), the system deviated from linear response theory, further enhancing heating efficiency due to nonlinear Brownian and Neel relaxation processes. The Zn_xMn_1–xFe₂O₄ MNPs exhibit good biocompatibility (95–70% cell viability) with tested concentrations of 50, 100, 250, 500, and 1000 μM over HeLa cell lines.

Pathogenic bacterial infections, which can perpetuate a harmful cycle of inflammation and hinder wound healing. Consequently, constructing a multifunctional strategy that can both eradicate bacteria and alleviate excessive inflammation holds great significance for wound healing. Herein, this study developed multifunctional metal-phenolic nanopreparations (Quer-Fe NPs). Through the one-pot coordination of quercetin (Quer) and Fe, Quer-Fe NPs possess outstanding photothermal properties and reactive oxygen species scavenging capability. After the photothermal destruction of the biofilm, Quer-Fe NPs can ultimately exhibit good broad-spectrum antibacterial effects against Staphylococcus aureus (99.23%), Escherichia coli (90.34%), and Candida albicans (72.62%). RNA sequencing indicates that under the photothermal treatment of Quer-Fe NPs, it can interfere with the bacterial metabolic process and genetic material repair process, affect bacterial proliferation and biofilm diffusion, thereby achieving excellent antibacterial outcomes. Additionally, Quer-Fe NPs can also upregulate the anti-inflammatory genes and downregulate the pro-inflammatory genes in macrophages, and promote the polarization of macrophages from M1 to M2 to relieve inflammation. The in vivo wound healing treatment experiment demonstrates that this nanoformulation can accelerate the wound healing process. In this groundbreaking study, an ingeniously contrived minimalist methodology was formulated to synthesize multifunctional metal-phenolic nanozymes. These nanozymes incorporate highly efficacious photothermal antibacterial activity, bacterium-ensnaring capabilities, along with anti-inflammatory attributes, thereby spotlighting their prodigious potential in the remediation of bacterial infections.

This study investigates the multifunctional potential of ZnO/CuO/Clay heterojunction nanocomposites (NCs) synthesized via the solution combustion method. Six NCs were prepared by varying ZnO/CuO ratios within two clay molar fractions (0.25 and 0.5 mol). Structural and compositional analyses (field emission scanning electron microscopy (FE-SEM), energy-dispersive X-ray spectroscopy (EDS), inductively coupled plasma optical emission spectroscopy (ICP-OES), dynamic light scattering (DLS), and Brunauer–Emmett–Teller (BET)) confirmed successful heterojunction formation, uniform elemental distribution, stable colloidal behavior, and a mesoporous nanostructure. Ultraviolet-visible diffuse reflectance spectroscopy (UV–vis DRS) revealed enhanced visible-light absorption with increasing CuO and decreasing clay content, thereby improving the NC’s optical characteristics and resulting in enhanced photocatalytic performance. Band gap measurements revealed CuO’s band gap narrowing effect, while ZnO and clay increased it. Antibacterial assays against Escherichia coli and Staphylococcus aureus showed significantly enhanced activity, with lower MIC values observed for NCs containing 0.25 mol clay. This behavior can be attributed to the smaller particle size, improved nanoparticle (NP) dispersion, reduced aggregation, increased porosity, and greater active surface area of these NCs compared to those with 0.5 mol clay. Transmission electron microscopy (TEM) imaging confirmed membrane disruption as a key antibacterial mechanism, supported by reactive oxygen species (ROS) generation, ion release, and synergistic interaction with clay nanosheets. Cytotoxicity tests on cancerous HT-29 cells demonstrated dose- and time-dependent behavior for the 0.75CuO/0.25Clay NC and dose-dependent behavior for the 0.75ZnO/0.25Clay NC, both with minimal toxicity to normal HFF cells. Antioxidant evaluation showed significant 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity (57%), comparable to ascorbic acid (60.2%). Overall, these results highlight solution combustion-synthesized ZnO/CuO/Clay NCs as promising bioactive materials for photocatalytic, antibacterial, anticancer, and antioxidant applications in medicine, food packaging, and environmental remediation.

The ongoing threat of antimicrobial-resistant (AMR) bacteria and the growing population of AMR bacteria have inspired research into alternative antimicrobial agents. Previous studies have shown clinically relevant bactericidal effects of the molecule nitric oxide (NO). Not only has extensive research proven its antimicrobial effect, but bacteria have also been shown to be less likely to become resistant to NO. Numerous studies have also demonstrated that NO is compatible with commercially available antibiotic drugs, enhancing antimicrobial effects. However, these drugs are not always readily available or easily manufactured. This study proposes combining NO with naturally sourced antibacterial agents, namely bacteriophages. This combination of NO with bacteriophages in solution demonstrated an 82 ± 1.7% killing efficiency against its target pathogen, Escherichia coli, and a 74 ± 2.9% reduction in Methicillin-resistant Staphylococcus aureus throughout a 12 h growth curve, indicating significant potential for further development as a broad-spectrum antimicrobial combination therapy.

This protocol was developed to assess in vivo osteogenesis of 3D-printed, selectively polymerized β-tricalcium phosphate (SP-βTCP) scaffolds placed between the periosteum and native bone of sheep scapulae. The protocol spans the entire development process of scaffold design, infusion, implantation and explant analysis. 3D printed SP-βTCP scaffolds of variable pore size were infused with combinations of gelatin methacryloyl and autologous or allogeneic adipose-derived stem cells (ADSCs), and placed within plasma-treated polyetherketone bioreactor chambers manufactured by laser sintering. These were implanted for 12 weeks on the left sheep scapula and 16 weeks on the right, followed by explantation and scanning using micro computed tomography (μCT). Images were analyzed using Imalytics Preclinical software via six key steps: (i) experimental chamber selection, (ii) chamber isolation, (iii) scaffold thresholding, (iv) bone thresholding, (v) volume generation and (vi) verification. This protocol delineated the border between bone and scaffold material, allowing for reliable quantification of bone formation. ADSCs and scaffolds with a smaller pore size were associated with superior bone formation, regardless of cell origin. These results demonstrate the utility of this protocol in analyzing μCT images in situations where high-density biomaterials degrade at variable rates.

The unprecedented use of conventional, commercial fertilizers has degraded soil health and enhanced aquatic pollution, and is inefficient in improving crop yield and productivity, suggesting the need for environmentally friendly alternatives. The current work entails the fabrication of customizable 3D-printed nanoparticle (NPs)-based micronutrient-releasing systems designed for improving plant growth parameters, nutritional aspects, and productivity. A 3D-printed precision agriculture platform was designed with various nanoparticles embedded in a gelatin matrix with engineered release profiles through a varied degree of cross-linking (0.2–1% cross-linker). The developed systems present a relatively faster release of Zn and relatively slow release of Fe and Mn nanoparticles, respectively, targeting various growth stages in wheat plants (Triticum aestivum). 3D-printed micronutrient fertilizers (MnFts) showed an improved swelling of 340%, with high water retention until 24 h, and slow, sustained release of micronutrients such as Mn, Fe, and Zn NPs for 7 days in aqueous media and 15 days in the soil medium. In this study, 3D-printed MnFts show enhancement in various growth stages of wheat plants (Triticum aestivum) (14.2% shoot length, 40.7% root length, 27.3% chlorophyll content, and 40% root volume increase), grain characteristics (∼50% more grains), total proteins (35.5% increase), pigments (32.3% increase), antioxidant enzymes (40.2% increase), and NPs content in roots, grain, and shoots. The pre- and post-treatment of the soil with 3D-printed MnFts did not affect the inherent soil microbial communities, suggesting the released Mn, Fe, and Zn NPs and degraded 3D-printed structures are nontoxic. The customizable 3D-printed structures with an engineered release profile of micronutrients targeting different growth stages of plants improve plant productivity and show no toxicity toward the soil microbial community, suggesting its potential for scalable adaptation in replacing conventional fertilizers for sustainable agriculture and environments.

Rapid and accurate detection of procalcitonin (PCT), a major biomarker for bacterial infections and sepsis, remains a pressing need in clinical diagnostics because sepsis progresses rapidly and may initially present with nonspecific or even subtle symptoms. Herein, we report a CRISPR-Cas12a-based fluorescence biosensing platform for ultrasensitive detection of PCT. The platform employs antibody-functionalized magnetic beads (MBs) for specific protein enrichment and antibody- and oligonucleotide- dual-functionalized gold nanoparticles (AuNPs) for high-density DNA payload. After sandwich complex formation with the target PCT, a programmed ssDNA strand is released by thermal denaturation, which then activates Cas12a collateral cleavage, thereby generating a fluorescence signal. Thorough physicochemical characterizations, including zeta potential, dynamic light scattering, UV–vis spectroscopy, and TEM, were carried out to confirm the successful functionalization of MBs and AuNPs. The developed PCT sensor was highly sensitive with a limit of detection (LOD) reaching 3 pg/mL. Moreover, the biosensor exhibited an excellent specificity toward PCT against clinically relevant interferents such as C-reactive protein (CRP), interleukin-2β (IL-2β), interleukin-6 (IL-6), human serum albumin (HSA), and bovine serum albumin (BSA), and simulated serum sample analysis was successfully carried out with the recoveries ranging from 108 to 122%. The PCT sensing technique developed in this work offers the potential to be expanded to construct a multiplexing platform for simultaneous detection of multiple biomarker species for early and accurate disease diagnosis.