Biological Applications Research Articles

START: A Versatile Platform for Bacterial Ligand Sensing with Programmable Performances.

Recognition of signaling molecules for coordinated regulation of target genes is a fundamental process for biological systems. Cells often rely on transcription factors to accomplish these intricate tasks, yet the subtle conformational changes of protein structures, coupled with the complexity of intertwined protein interaction networks, pose challenges for repurposing these for bioengineering applications. This study introduces a novel platform for ligand-responsive gene regulation, termed START (Synthetic Trans-Acting Riboswitch with Triggering RNA). Inspired by the bacterial ligand sensing system, riboswitch, and the synthetic gene regulator, toehold switch, the START platform enables the implementation of synthetic biosensors for various ligands. Rational sequence design with targeted domain optimization yields high-performance STARTs with a dynamic range up to 67.29-fold and a tunable ligand sensitivity, providing a simple and intuitive strategy for sensor engineering. The START platform also exhibits modularity and composability to allow flexible genetic circuit construction, enabling seamless implementation of OR, AND, and NOT Boolean logic gates for multiple ligand inputs. The START design principle is capable of broadening the suite of synthetic biosensors for diverse chemical and protein ligands, providing a novel riboregulator chassis for synthetic biology and bioengineering applications.

Multi-resonance plasmonic refractive index sensor based on maze-shaped resonators for biological applications

Herein, a plasmonic refractive index (RI) sensor based on a metal–insulator–metal waveguide coupled with maze-shaped resonators is proposed and numerically investigated using finite element method. Various geometrical parameter impacts on the transmission spectrum are examined to optimize the sensor’s performance. Additionally, the effect of using SiO2 as a dielectric material instead of air has been investigated. The proposed sensor can achieve maximum RI sensitivity, figure of merit, and sensing resolution of 3340 nm RIU−1, 143.33 RIU−1, and 2.99 × 10−6 RIU, respectively, in the 500–3500 nm wavelength range. The designed structure is investigated for potential applications in different biological fields, including detecting cancer cells, determining blood hemoglobin (HB) levels, and glucose concentrations. This sensor can detect MCF-7 cancer cells with a maximum sensitivity of 3543 nm RIU−1 and can achieve the sensitivity of 0.407 nm·l g−1 for glucose concentration and 3329.41 nm RIU−1 for blood HB level. The structure presented in this study has promising specifications, making it suitable for use in optical integrated circuits, particularly in highly sensitive sensors.

Spirocyclic pyrrolidinyl nitroxides with exo-methylene substituents.

Nitroxides are stable organic radicals with exceptionally long lifetimes, which render them uniquely suitable as observable probes or polarising agents for spectroscopic investigation of biomolecular structure and dynamics. Radical-based probes for biological applications are ideally characterized by both robustness towards reductive degradation and beneficial electron spin relaxation parameters. These properties are largely influenced by the molecular structure of the nitroxide scaffold, and also by the conformations it prefers to adopt. In this study we present the synthesis of the first nitroxides based on a spirocyclic pyrrolidine scaffold with an exocyclic methylene substituent. The conformations adopted by these nitroxides were evaluated by X-ray crystallography, both with single nitroxide crystals and by inclusion of nitroxides in a microporous crystalline sponge. The kinetic and thermodynamic stability of the new nitroxides towards reduction was investigated by electron paramagnetic resonance (EPR) spectroscopy and cyclic voltammetry (CV). In combination with EPR measurements of electron spin relaxation properties, these results suggest that this new family of nitroxides can provide access to multifunctionalized probes and polarising agents suitable for use in biological environments at elevated temperatures.

Comparative analysis of hyperspectral Image reconstruction using deep learning for agricultural and biological applications

Hyperspectral imaging (HSI) has become a key technology for non-invasive quality evaluation in various fields, offering detailed insights through spatial and spectral data. Despite its efficacy, the complexity and high cost of HSI systems have hindered their widespread adoption. This study addressed these challenges by exploring deep learning-based hyperspectral image reconstruction from RGB (Red, Green, Blue) images, particularly for agricultural products. Specifically, different hyperspectral reconstruction algorithms, such as Hyperspectral Convolutional Neural Network - Dense (HSCNN-D), High-Resolution Network (HRNET), and Multi-Scale Transformer Plus Plus (MST++), were compared to assess the dry matter content of sweet potatoes. Among the tested reconstruction methods, HRNET demonstrated superior performance, achieving the lowest mean relative absolute error (MRAE) of 0.07, root mean square error (RMSE) of 0.03, and the highest peak signal-to-noise ratio (PSNR) of 32.28 decibels (dB). Some key features were selected using the genetic algorithm (GA), and their importance was interpreted using explainable artificial intelligence (XAI). Partial least squares regression (PLSR) models were developed using the RGB, reconstructed, and ground truth (GT) data. The visual and spectra quality of these reconstructed methods was compared with GT data, and predicted maps were generated. The results revealed the prospect of deep learning-based hyperspectral image reconstruction as a cost-effective and efficient quality assessment tool for agricultural and biological applications.

Towards Self-regeneration: Exploring the Limits of Protein Synthesis in the Protein Synthesis Using Recombinant Elements (PURE) Cell-free Transcription-Translation System.

Self-regeneration is a key function of living systems that needs to be recapitulated in vitro to create a living synthetic cell. A major limiting factor for protein self-regeneration in the PURE cell-free transcription-translation system is its high protein concentration, which far exceeds the system's protein synthesis rate. Here, we were able to drastically reduce the nonribosomal PURE protein concentration up to 97.3% while increasing protein synthesis efficiency. Although crowding agents were not effective in the original PURE formulation, we found that in highly dilute PURE formulations, addition of 6% dextran considerably increased protein synthesis rate and total protein yield. These new PURE formulations will be useful for many cell-free synthetic biology applications, and we estimate that PURE can now support the complete self-regeneration of all 36 nonribosomal proteins, which is a critical step toward the development of a universal biochemical constructor and living synthetic cell.

Water-Soluble Alumina-Coated Indium Phosphide Core-Shell Quantum Dots with Efficient Deep-Red Emission Beyond 700nm.

Solution-processed colloidal III-V semiconductor-based quantum dots (QDs) represent promising and environmentally-friendly alternatives to Cd-based QDs in the realms of optoelectronics and biological applications. While InP-based core-shell QDs have demonstrated efficientlight-emitting diode (LED) performance in the visible region, achieving deep-red emission (above 700nm) with a narrow linewidth has proven challenging. Herein, the study presents a novel strategy for synthesizing InP/ZnSe/ZnS core-shell-shell QDs tailored for emission in the first biological transparency window. The resulting QDs exhibit an emission wavelength up to 725nm with a narrow peak full width at half maximum (FWHM) down to 107 meV (45nm). To enhance the biocompatibility and chemical stability of the QDs, their surface is further capped with a layer of amorphous alumina resulting in an InP/ZnSe/ZnS/Al2O3 heterostructure. This surface passivation not only ensures environmental- and photostability but also enhances the photoluminescence quantum yield (PLQY). The alumina capping enables the aqueous phase transfer via surface ligand exchange using mercaptopropionic acid (MPA) while maintaining the initial quantum yield. The resulting QDs demonstrate a significant potential for advancing next-generation optoelectronic technologies and bio-applications.

Assessing next-generation sequencing-based computational methods for predicting transcriptional regulators with query gene sets.

This article provides an in-depth review of computational methods for predicting transcriptional regulators (TRs) with query gene sets. Identification of TRs is of utmost importance in many biological applications, including but not limited to elucidating biological development mechanisms, identifying key disease genes, and predicting therapeutic targets. Various computational methods based on next-generation sequencing (NGS) data have been developed in the past decade, yet no systematic evaluation of NGS-based methods has been offered. We classified these methods into two categories based on shared characteristics, namely library-based and region-based methods. We further conducted benchmark studies to evaluate the accuracy, sensitivity, coverage, and usability of NGS-based methods with molecular experimental datasets. Results show that BART, ChIP-Atlas, and Lisa have relatively better performance. Besides, we point out the limitations of NGS-based methods and explore potential directions for further improvement.

Fabrication of heterocellular spheroids with controllable core-shell structure using inertial focusing effect for scaffold-free 3D cell culture models

Three-dimensional (3D) cell culture models capable of emulating the biological functions of natural tissues are pivotal in tissue engineering and regenerative medicine. Despite progress, the fabrication ofin vitroheterocellular models that mimic the intricate structures of natural tissues remains a significant challenge. In this study, we introduce a novel, scaffold-free approach leveraging the inertial focusing effect in rotating hanging droplets for the reliable production of heterocellular spheroids with controllable core-shell structures. Our method offers precise control over the core-shell spheroid's size and geometry by adjusting the cell suspension density and droplet morphology. We successfully applied this technique to create hair follicle organoids, integrating dermal papilla cells within the core and epidermal cells in the shell, thereby achieving markedly enhanced hair inducibility compared to mixed-structure models. Furthermore, we have developed melanoma tumor spheroids that accurately mimic the dynamic interactions between tumor and stromal cells, showing increased invasion capabilities and altered expressions of cellular adhesion molecules and proteolytic enzymes. These findings underscore the critical role of cellular spatial organization in replicating tissue functionalityin vitro. Our method represents a significant advancement towards generating heterocellular spheroids with well-defined architectures, offering broad implications for biological research and applications in tissue engineering.

Shining New Light on Biological Systems: Luminescent Transition Metal Complexes for Bioimaging and Biosensing Applications.

Luminescence imaging is a powerful and versatile technique for investigating cell physiology and pathology in living systems, making significant contributions to life science research and clinical diagnosis. In recent years, luminescent transition metal complexes have gained significant attention for diagnostic and therapeutic applications due to their unique photophysical and photochemical properties. In this Review, we provide a comprehensive overview of the recent development of luminescent transition metal complexes for bioimaging and biosensing applications, with a focus on transition metal centers with a d6, d8, and d10 electronic configuration. We elucidate the structure-property relationships of luminescent transition metal complexes, exploring how their structural characteristics can be manipulated to control their biological behavior such as cellular uptake, localization, biocompatibility, pharmacokinetics, and biodistribution. Furthermore, we introduce the various design strategies that leverage the interesting photophysical properties of luminescent transition metal complexes for a wide variety of biological applications, including autofluorescence-free imaging, multimodal imaging, organelle imaging, biological sensing, microenvironment monitoring, bioorthogonal labeling, bacterial imaging, and cell viability assessment. Finally, we provide insights into the challenges and perspectives of luminescent transition metal complexes for bioimaging and biosensing applications, as well as their use in disease diagnosis and treatment evaluation.

Achieving Ultrasound-Excited Emission with Organic Mechanoluminescent Materials.

Unlike traditional photoluminescence (PL), mechanoluminescence (ML) achieved under mechanical excitation demonstrates unique characteristics such as high penetrability, spatial resolution, and signal-to-background ratio (SBR) for bioimaging applications. However, bioimaging with organic mechanoluminescent materials remains challenging because of the shallow penetration depth of ML with short emission wavelengths and the absence of a suitable mechanical force to generate ML in vivo. To resolve these issues, the present paper reports the achievement of ultrasound (US)-excited fluorescence and phosphorescence from purely organic luminogens for the first time with emission wavelengths extending to the red/NIR region, with the penetrability of the US-excited emission being considerably higher than that of PL. Consequently, US-excited subcutaneous phosphorescence imaging can be achieved using a mechanoluminescent-luminogen-based capsule device with a quantified intensity of 9.15 ± 1.32 × 104 p s-1 cm-2 sr-1 and an SBR of 24. Moreover, the US-excited emission can be adequately tuned using the packing modes of the conjugated skeletons, dipole orientation of mechanoluminescent luminogens, and strength and direction of intermolecular interactions. Overall, this study innovatively expands the kind of excitation sources and the emission wavelengths of organic mechanoluminescent materials, paving the way for practical biological applications based on US-excited emission.

Leaching-Free durable antibacterial composite films constructed via delicate balance between potent contact sterilization and low adhesion

Durable antibacterial surfaces are often required high bactericidal ability and low bacterial adhesion property, but the potent bactericidal surfaces, especially based on the contact bactericidal mechanism, are often paradoxically intended to attract bacteria to adhere. In this work, a delicate balance between the potent contact bactericidal action and low bacterial adhesion was successfully achieved in quaternized fluoropolymer/SiO2 composite films (QFP/SiO2 CFs) through judicious selection of a common fluorinated monomer, a common quaternary ammonium monomer, and a common inert inorganic nanoparticle, to acquire suitable surface energy and chain mobility to ensure low bacterial adhesion, as well as to allow appropriate surface-exposed quaternary ammonium units to synergistically enhance the contact bactericidal action with the specific fluorinated side chains. The QFP/SiO2 CFs were conveniently fabricated through miniemulsion polymerization and followed squeegeeing technique. The QFP/SiO2 CF with an optimized composition exhibited extremely low initial adhesion of bacteria, potent contact bactericidal action, superior long-term antibacterial performance, easy sonication-assistant regeneration for multiple usage, and low cytotoxicity with non-leachable attribute. This work may guide the design and efficient fabrication of leaching-free durable antibacterial surfaces for biological applications, as well as provide an effective strategy to balance contact bactericidal action and low bacteria adhesion for durable antibacterial performance.

Supramolecular Polymer Co-Assembled Multifunctional Chiral Hybrid Hydrogels with Adhesive, Self-Healing and Antibacterial Properties

Amino acid-derived self-assembled nanofibers comprising supramolecular chiral hydrogels with unique physiochemical characteristics are highly demanded biomaterials for various biological applications. However, their narrow functionality often limits practical use, necessitating the development of biomaterials with multiple features within a single system. Herein, chiral co-assembled hybrid hydrogel systems termed LPH-EGCG and DPH-EGCG were constructed by co-assembling L/DPFEG gelators with epigallocatechin gallate (EGCG) followed by cross-linking with polyvinyl alcohol (PVA) and hyaluronic acid (HA). The developed hybrid hydrogels exhibit superior mechanical strength, self-healing capabilities, and adhesive properties, owing to synergistic non-covalent interactions. Integrating hydrophilic polymers enhances the system’s capacity to demonstrate favorable swelling characteristics. Furthermore, the introduction of EGCG facilitated the hybrid gels to display notable antibacterial properties against both Gram-positive and Gram-negative bacterial strains, alongside showcasing strong antioxidant capabilities. In vitro investigation demonstrated enhanced cell adhesion and migration with the LPH-EGCG system in comparison to DPH-EGCG, thus emphasizing the promising prospects of these hybrid hydrogels in advanced tissue engineering applications.

Solvent-free synthesis of TiO2 nanoparticles embedded MWCNTs with enhanced photocatalytic activity for bacterial disinfection

Sustainable energy has been widely researched to address issues caused by population growth and rapid industrialization. Among them, photocatalysis using solar energy has been considered promising. We developed a hybrid photocatalyst by combining titanium dioxide (TiO2) with carbon nanotubes (CNTs), known for their excellent adsorption performance. A unique structure encapsulating TiO2 nanoparticles (NPs) within the inner space of multi-walled carbon nanotubes (MWCNTs) was developed via a facile, solvent-free method, conforming to green chemistry principles. Except purification and modification of the CNTs, the particle formation process is solvent-free, making this method remarkably simple and environmentally friendly. This arrangement enhances stability and recyclability, maintaining over 90 % efficiency up to the 3rd cycle. X-ray diffraction (XRD) analysis confirmed that the TiO2 NPs exhibited a crystalline phase, forming exclusively anatase. The defects in the outer walls of the MWCNTs in the composite, coupled with their large specific surface area and dangling bonds, endow them with high adsorption capacity and photocatalytic activity when combined with TiO2 NPs. The MWCNTs@TiO2 composites achieved over 99 % inactivation of Escherichia coli in 60 min and bacteriophage MS2 in 30 min, along with dye adsorption/decomposition. This method offers a distinctive platform for photocatalytic degradation in biological applications and water treatment.

Synthesis of hydroxyapatite and strontium-substituted hydroxyapatite for bone replacement and osteoporosis treatment

Hydroxyapatite (HAp) is an inorganic component exhibiting bioactivity similar to that of natural bone. However, it is not resorbed by osteoclasts during bone remodelling due to its lack of bio-resorption property. This can be enhanced by the substitution of other element presented in bone mineral. In this research work, hydroxyapatite (HAp) and strontium-substituted hydroxyapatite (Sr-HAp) were synthesized by a precipitation method. Calcium nitrate tetra hydrate [Ca(NO3)2•4H2O], disodium hydrogen phosphate (Na2HPO4), and Strontium nitrate [Sr(NO3)2] were used as Ca, PO4 and Sr sources, respectively. Molar ratio Ca/P=1.67 was used to synthesize HAp, where (Ca+Sr)/P=1.67 was used to synthesize strontium substituted-HAp (Sr-HAp). The reaction was carried out at room temperature. The results show that pure HAp and Sr-HAp were formed with nanometer-sized particles. Sr substitution in the HAp lattice results in an increase in both the lattice disorder and crystal aspect ratio. The results of in vitro bioactive testing using simulated bodily fluid also showed that both HAp and Sr-HAp have high bioactive, with the Sr-HAp sample having the greater bioactive. Therefore, HAp and Sr-HAp have great potential for biological applications.

Bioelectret Materials and Their Bioelectric Effects for Tissue Repair: A Review.

Biophysical and clinical medical studies have confirmed that biological tissue lesions and trauma are related to the damage of an intrinsic electret (i.e., endogenous electric field), such as wound healing, embryonic development, the occurrence of various diseases, immune regulation, tissue regeneration, and cancer metastasis. As exogenous electrical signals, such as conductivity, piezoelectricity, ferroelectricity, and pyroelectricity, bioelectroactives can regulate the endogenous electric field, thus controlling the function of cells and promoting the repair and regeneration of tissues. Materials, once polarized, can harness their inherent polarized static electric fields to generate an electric field through direct stimulation or indirect interactions facilitated by physical signals, such as friction, ultrasound, or mechanical stimulation. The interaction with the biological microenvironment allows for the regulation and compensation of polarized electric signals in damaged tissue microenvironments, leading to tissue regeneration and repair. The technique shows great promise for applications in the field of tissue regeneration. In this paper, the generation and change of the endogenous electric field and the regulation of exogenous electroactive substances are expounded, and the latest research progress of the electret and its biological effects in the field of tissue repair include bone repair, nerve repair, drug penetration promotion, wound healing, etc. Finally, the opportunities and challenges of electret materials in tissue repair were summarized. Exploring the research and development of new polarized materials and the mechanism of regulating endogenous electric field changes may provide new insights and innovative methods for tissue repair and disease treatment in biological applications.

Mitochondria-targeted and photo-activated CO release for synergistic photodynamic therapy

In photodynamic therapy, organelle-targeted strategies are allowed to selectively impair specific cell functions, leading to efficiently eliminating cancer cells. Carbon monoxide (CO), as a highly toxic gaseous signaling molecule, often plays an active role in enhancing the sensitivity of cancer cells during cancer treatment, making cells prone to apoptosis. Therefore, we report a photosensitizer Cy-Mn, which possesses the capability of photoactivatable CO releasing and 1O2 generation. Furthermore, during the treatment, Cy-Mn has perfect synergistic effect on the cancer cells for enhanced photodynamic therapy. Under 760nm laser irradiation, Cy-Mn released manganese ions, which altered the permeability transition pore of the mitochondria membrane, reduced the mitochondrial membrane potential, and exhibited severe phototoxicity. Additionally, we investigated the imaging of Cy-Mn in vivo, and found Cy-Mn displayed excellent fluorescence imaging capabilities, which further promotes its biological applications in imaging-guided PDT.

Genetic Engineering and Innovative Cultivation Strategies for Enhancing the Lutein Production in Microalgae

Carotenoids, with their diverse biological activities and potential pharmaceutical applications, have garnered significant attention as essential nutraceuticals. Microalgae, as natural producers of these bioactive compounds, offer a promising avenue for sustainable and cost-effective carotenoid production. Despite the ability to cultivate microalgae for its high-value carotenoids with health benefits, only astaxanthin and β-carotene are produced on a commercial scale by Haematococcus pluvialis and Dunaliella salina, respectively. This review explores recent advancements in genetic engineering and cultivation strategies to enhance the production of lutein by microalgae. Techniques such as random mutagenesis, genetic engineering, including CRISPR technology and multi-omics approaches, are discussed in detail for their impact on improving lutein production. Innovative cultivation strategies are compared, highlighting their advantages and challenges. The paper concludes by identifying future research directions, challenges, and proposing strategies for the continued advancement of cost-effective and genetically engineered microalgal carotenoids for pharmaceutical applications.

An instructional design strategy for optimization of GelMA hydrogels material properties

Gelatin methacryloyl (GelMA) hydrogels have become a key biomaterial in various biomedical fields, such as tissue regeneration engineering and drug delivery, due to their exceptional biocompatibility and customizable physical and chemical characteristics. The resemblance of GelMA to the extracellular matrix and its ability to be cured with light make it a popular choice for researchers. This research delves into how the material properties of GelMA hydrogels, specifically the density of the molecular network (DMN), impact their performance in different applications. By preparing GelMA hydrogels with different degrees of substitution (DS) and concentrations, and utilizing these attributes as key measurable parameters for determining DMN, we conducted a comprehensive evaluation of properties relevant to biological applications. We also suggest strategies for optimizing these properties. Through the application of the DMN theory, we show that precise control over material properties allows for tailored customization of GelMA hydrogels for specific tissue engineering and drug delivery purposes. This study offers valuable insights into the optimization of GelMA hydrogel properties to advance the field of tissue engineering and improve customization of cellular microenvironments.

Gelatin and lipidoid integrate to create gelasomes to enhance siRNA delivery with low toxicity

RNAi therapeutics possess the potential to cure many uncurable human diseases. For instance, RNAi therapeutics using liposomes showed remarkable survival benefits in patients with liver diseases. However, the extension of liposomes to deliver RNA to cure other ailments has largely been unsuccessful. Therefore, researchers are focusing on designing and testing different combinations of materials for versatile RNA delivery applications. Yet, an efficient and safe RNA delivery platform has not been identified. In this work, we have developed a new class of RNA-delivery vehicle called “Gelasomes,” using an incongruous combination of gelatin and lipidoid to exploit each material's unique properties while overcoming their inherent limitations. The low in vivo toxicity of Gelasomes is attributed to the exterior gelatin layers that shield the exposure of cationic lipidoid-siRNA clusters and yet present a biocompatible surface. Indeed, toxicity studies in mice indicate that repeated administration of Gelasomes (up to 48 mg/kg BW) is well-tolerated with no notable changes in body weight, hematology, or serum chemistry. Interestingly, the gelatin outer layer efficiently protects siRNA from serum degradation (48 h), preserving its functionality beyond two months of storage. Notably, Gelasomes possess dual siRNA conjugation modes, i.e., electrostatic binding with lipidoid core and covalent attachment to gelatin surface. The bivalency coupled with lipidoids' high transfection efficiency rendered Gelasomes with remarkably high gene silencing efficiency (>90 %) at very low treatment doses in vitro (40 μg/mL). In vivo studies further confirmed the high gene silencing ability of Gelasomes in non-small cell lung tumor mouse models. This new platform is tunable on all fronts: size, degree of surface coating, and biomolecule functionalization. Truncating the lipidoid C14-tail to a C8-tail yielded Gelasomes of reduced size. As lipidoids with different carbon lengths are synthesizable, we can develop a library of Gelasomes with different sizes. The surface coating with less gelatin resulted in high transfection efficiency at low doses of Gelasomes. The structure of Gelasomes offers chemical handles to couple target-specific molecules like antibodies to tune their properties for efficient biological application.

Research Progress of Cyanine-based Near-Infrared Fluorescent Probes for Biological Application.

Cyanine-based near-infrared (NIR) fluorescent probes have played vital roles in biological application due to their low interference from background fluorescence, deep tissue penetration, high sensitivity, and minimal photodamage to biological samples. They are widely utilized in molecular recognition, medical diagnosis, biomolecular detection, and biological imaging. Herein, we provide a review of recent advancements in cyanine-based NIR fluorescent probes for the detection of pH, cells, tumor as well as their application in photothermal therapy (PTT) and photodynamic therapy (PDT).