2′-Fucosyllactose (2′-FL) was the first commercially available Human Milk Oligosaccharide (HMO) supplemented into infant formula. Microbial synthesis has already become the main approach to produce 2′-FL and the modification of the synthesis pathway involves multiple strategies. In this study, GDP-L-fucose biosynthetic genes (gmd-fcl-manC-manB, GFCB) and α-1,2-fucosyltransferase were multi-integrated into the chromosome or plasmid-expressed to produce 2′-FL in E. coli. To further enhance the 2′-FL production, the expression levels of biosynthetic genes were fine-tuned, and manC was identified as the rate-limiting step. With the chromosome-integrated and plasmid-expressed GFCB and manC, the best microbial 2′-FL production was obtained. The titer reached 115.5 g/L in a 30- L bioreactor, with a productivity of 1.48 g/L/h. High 2′-FL titer was achieved merely via inactivation of two endogenous genes and overexpression of five key biosynthetic genes. This study revealed the importance of gene allocation and rate-limiting gene over-expression for HMO production.

Pollen allergy is a prevalent allergic disorder affecting a substantial proportion of the global population. Allergen-specific immunotherapy (ASIT) is the only disease-modifying treatment, acting by inducing regulatory T cells (Tregs) and Th1-based immunity. However, current ASIT approaches often elicit insufficient allergen-specific tolerance and fail to correct underlying immune dysregulation, thereby limiting clinical efficacy. Here, a combination adjuvant composed of aluminum oxyhydroxide and CpG oligodeoxynucleotides (Al-CpG) is developed and formulated with the T-cell epitope derived from the Artemisia allergen Art v 1 (rArt v 1). Al-CpG significantly enhances rArt v 1 uptake and presentation by bone marrow-derived dendritic cells (BMDCs) and promotes the secretion of IFN-γ and IL-10. In both therapeutic and prophylactic murine models of pollen allergy, Al-CpG vaccination significantly alleviated allergic symptoms, suppressed airway inflammation, and restored immune homeostasis. Mechanistically, Al-CpG induces a robust expansion of CD25+FoxP3+ Tregs in concert with IFN-γ-producing CD4+ T cells, indicative of coordinated regulatory and Th1 immune programming. Adoptive transfer of Al-CpG-conditioned BMDCs further demonstrates preferential expansion of stable, thymus-derived (Helios+FoxP3+) Tregs in draining lymph nodes and systemic immune reprogramming toward a regulated state in the spleen, as evidenced by increased Treg frequencies and CTLA-4 upregulation. Collectively, these findings identify Al-CpG as a safe and effective adjuvant platform that synergistically induces immune tolerance and immune deviation, highlighting its potential for next-generation pollen allergy vaccines and broader applications in the prevention and treatment of allergic diseases.

The low immunogenicity of peptide vaccines remains a critical challenge in immunotherapy. While peptide nanofibrils can function as self-delivery systems to activate antigen-presenting cells (APCs), most existing designs rely on complex covalent conjugation to β-sheet-forming motifs. Moreover, the diverse surface properties of various antigenic peptides complicate the rational selection of appropriate adjuvants. Herein, we develop a nanovaccine platform in which antigenic peptides self-assemble into nanofibrils without the need for exogenous β-sheet-forming sequences. Silica nanoparticles (SiO2 NPs) are nucleated and grown directly along these fibrils, resulting in SiO2@fibril nanovaccines with a unique "beads-on-a-string" morphology. Using amyloid-β(1-42) (Aβ42) and human papillomavirus (HPV) type 16 E7 (E7) as model antigens, we demonstrate that these nanovaccines significantly enhance the maturation and activation of bone marrow-derived dendritic cells (BMDCs). In an Alzheimer's disease animal model using APP/PS1 mice, SiO2@Aβ42 nanovaccines improve motor and cognitive function. Additionally, in an HPV animal model using TC-1 tumor-bearing mice, SiO2@E7 nanovaccines suppress tumor growth and increase survival rate. This strategy provides a universal and modular adjuvant platform for peptide-based nanovaccines.

Pollen allergy is a prevalent respiratory allergic disease, yet current allergen immunotherapy (AIT) remains limited by safety concerns and insufficient induction of durable immune tolerance. Here, we develop an engineered mRNA-LNP platform for allergen-specific tolerance induction by integrating allergen and delivery system optimization. A non-secretory mRNA encoding the major mugwort pollen allergen Art v 1 fused with invariant chain (mArt v 1) was designed to reduce IgE-mediated activation while promoting tolerogenic allergen processing. This antigen was delivered using a mannose-functionalized LNP formulation (mAr-C18mann) optimized for low inflammatory activation and enhanced immune regulation. The engineered platform promotes a tolerance-associated hepatic immune environment, regulates antigen-presenting cell responses, and enhances allergen-specific regulatory T-cell induction. In a murine model of pollen allergy, both mAr-C18mann and a simplified epitope-based variant (mdT-C18mann) attenuate allergic inflammation and allergen-specific IgE levels. Consistently, mAr-C18mann reprograms immune responses toward Treg- and Th1-dominated profiles, providing effective protection in a prophylactic allergy model. These findings support the mRNA-LNP platform as a modular next-generation AIT strategy for inducing durable immune tolerance to pollen allergy.

Messenger RNA (mRNA) vaccines, which leverage quick and customizable antigen production, have transformed immunization. However, their widespread use is still limited by challenges related to the natural instability of in vitro transcribed mRNA and the limitations of current delivery systems. Modern perspectives on stability extend beyond simple resistance to degradation. They now consider factors that can be modified through RNA structure and nanoscale environmental interactions. These interactions, which promote efficient translation, also affect the durability of these complexes against physical and chemical stresses. During formulation and storage, variables such as excipients, buffers, and solid-state architecture are essential for maintaining molecular integrity throughout manufacturing, transportation, and long-term preservation. As a result, stability serves as a key link between molecular design and overall vaccine efficacy, transforming a major obstacle into an area for strategic innovation. This review highlights recent advances at both the molecular and carrier levels aimed at developing thermostable, efficient, and highly effective mRNA vaccines, with a focus on improvements in their stability, storability, and delivery.

Aluminum adjuvants are crucial for the efficacy of inactivated and subunit vaccines. However, their effectiveness is often compromised during lyophilization, and the underlying mechanisms of freeze-drying-induced damage remain poorly defined. In this study, we mechanistically investigated the interfacial behavior of engineered amorphous aluminum hydroxyphosphate nanoparticles (AAHP NPs) during lyophilization. Through comprehensive physicochemical characterization at the ice-particle interface, we demonstrate that the surface charge of AAHP NPs governs their interfacial interactions with ice crystals. Charged AAHP NPs electrostatically repel ice recrystallization growth, thereby suppressing ice-crystal-induced physicochemical stress, mitigating structural damage, and markedly improving colloidal stability upon reconstitution. In a diphtheria toxoid vaccination model, when positively charged AAHP nanoparticles were formulated with 5% sucrose during lyophilization, antigen-specific humoral immune responses were maintained. Collectively, these results elucidate key mechanisms of lyophilization-induced damage to aluminum adjuvants and provide insights into rational design strategies for lyophilized vaccine formulations.

Nanoparticles (NPs) have emerged as ice-control materials to prevent damage caused by low-temperature-induced ice crystals. However, the mechanism of ice recrystallization inhibition (IRI) of NPs is poorly understood. A thorough understanding of the relationships between the physicochemical properties of nanomaterials and their IRI ability remains to be established. In this study, we prepared a comprehensive library of NPs, including Stöber silica, poly(lactic-co-glycolic acid) (PLGA), and Au NPs with tunable physicochemical properties, and examined their IRI functionalities. The results revealed that the surface free energy (SFE) of NPs was closely related to their IRI ability, which was further supported by molecular dynamics (MD) simulations. Our findings highlight the critical role of nanomaterials' SFE in mediating their IRI activity, advancing the fundamental understanding of the mechanism underlying NP-controlled ice recrystallization. This study provides a novel design strategy for developing nanoparticles as ice-control materials.

Inhalable immunization is a multidimensional colloidal delivery process, involving aerosol particle transport in the airways and interaction with mucus at the mucosal interface. Primarily, the impact of airway anatomy, airflow, and particle's physicochemical properties on aerosol deposition and deep lung delivery are highly desired. In this study, a human airway geometry model was constructed, and the airflow field distribution and turbulence characteristics based on airway anatomy were visualized through in silico simulation. The dimensionless numbers of Stokes number (Stk) and Schmidt number (Sc) were introduced to mechanistically demonstrate the particle deposition under the impact of multiple parameters, including inlet airflow rate (Q), particle size (dp), and particle density (rho). With the increasement of Stk and Sc, the mechanism transitioned from Brownian diffusion- to inertial impact-dominated deposition. The response surface methodology (RSM) analysis indicated that the Q and interaction term between particle size and inlet airflow rate (dpQ) were the most critical parameters that dominated deep lung transportation fraction (DLF). The proposed regression equation provided quantitative design guidance for parameter of airflow and particle. This in silico methodology provides a rational predictive design strategy for inhaled aerosol formulations for both therapeutic and prophylactic applications.

The stability of protein antigens during production, storage, and transportation is critical for determining the effectiveness and safety of vaccines. However, protein antigens are susceptible to thermal and mechanical stresses, which can lead to aggregation and decreased immunogenicity. This study utilized alkyl glucosides to analyze the relationship between the structural characteristics of alkyl glucosides and their effects on the biophysical properties of proteins by using antigens, including bovine serum albumin (BSA), human papillomavirus virus-like particles (HPV VLPs), and tetanus toxoid (TT), through comprehensive biophysical analysis. Among the surfactants tested, N-dodecyl-β-d-maltoside (MalC12), with its longer hydrophobic chain and dual-sugar headgroup, was particularly effective in preserving antigen stability. MalC12 formed a dense protective layer through hydrophobic interactions, reducing heat-induced aggregation and maintaining the proteins' secondary structures. Under mechanical stress, MalC12 decreased protein adsorption at interfaces, minimized the risk of protein unfolding, and enhanced hydrophobic interactions with the proteins. In an HPV vaccine model, MalC12 effectively preserved HPV immunogenicity by preventing thermal-induced aggregation. These findings highlight the importance of surfactant characteristics in preventing protein destabilization and offer valuable insights for designing excipients that enhance antigen stability and immunogenicity in vaccine formulations.

Recent progress in stem cell therapy has demonstrated the therapeutic potential of intravenous stem cell infusions for treating the life-threatening lung disease of pulmonary fibrosis (PF). However, it is confronted with limitations, such as a lack of control over cellular function and rapid clearance by the host after implantation. In this study, we developed an innovative PF therapy through tracheal administration of microfluidic-templated stem cell-laden microcapsules, which effectively reversed the progression of inflammation and fibrotic injury. Our findings highlight that hydrogel microencapsulation can enhance the persistence of donor mesenchymal stem cells (MSCs) in the host while driving MSCs to substantially augment their therapeutic functions, including immunoregulation and matrix metalloproteinase (MMP)-mediated extracellular matrix (ECM) remodeling. We revealed that microencapsulation activates the MAPK signaling pathway in MSCs to increase MMP expression, thereby degrading overexpressed collagen accumulated in fibrotic lungs. Our research demonstrates the potential of hydrogel microcapsules to enhance the therapeutic efficacy of MSCs through cell-material interactions, presenting a promising yet straightforward strategy for designing advanced stem cell therapies for fibrotic diseases.

Nanotechnology-based photodynamic (PDT) and photothermal (PTT) therapies offer precise treatment by generating reactive oxygen species (ROS) and localized hyperthermia. However, photosensitizer nanodrugs suffer from rapid degradation and limited tumor targeting. To overcome these challenges, we have developed a dual-nanoparticle drug delivery platform based on bowl-shaped polymer vesicles, which were loaded in the protective nanocavity with nanofibers composed of a histidine dipeptide-photosensitizer conjugate. The stomatocytes were surface modified with mannose glycopolymers to induce targeting capacity. This dual nanoparticle platform exhibited enhanced tumor imaging, improved targeting, and increased cellular uptake in tumor cells and 3D tumor spheroids. Under 660 nm laser irradiation, it demonstrated excellent PTT and PDT performance, with significant ROS generation and temperature elevation. This advanced nanomedicine platform, therefore, offers a promising approach for precision cancer therapy.

Coacervate-based artificial cells have gained significant attraction in synthetic biology for their ability to mimic life-like functions such as compartmentalization, selective molecular uptake, and the hosting of biochemical reactions. However, the incorporation of motility, a key feature of natural cells, remains underexplored. This is mainly caused by the dynamic character of coacervates, which hampers their stability and limits control over functional motile components within the structure. In this contribution, we have been able to address this gap by physically anchoring gold nanoparticles (AuNPs)-coated nanomotors at the coacervate interface in combination with a terpolymer membrane. The positively charged coacervates promoted the assembly of negatively charged nanomotors on their surfaces via electrostatic interactions. By costabilizing the coacervates with a terpolymer membrane, patches of nanomotors were firmly immobilized on the coacervates' surface and the stability of coacervates was preserved during motion performance. The distribution of nanomotors shifted from spotted distribution, patchy distribution, to almost full coverage upon increasing nanomotors concentration. Optimal motile behavior was found when achieving a patchy coverage of nanomotors at the interface, which enabled the system to become a light-driven micromotor platform through the surface plasmon thermal effect of the AuNPs. Remarkably, the motion dynamics of these coacervate droplets could be modulated by tuning nanomotors' density on the surface, coacervates' size, and laser light intensity. This study provides a first example of a coacervate system, which is stabilized by a combination of nanoparticles and a terpolymer membrane, of which their motility is effectively transferred to the artificial cell structure.

[This corrects the article DOI: 10.1016/j.mtbio.2025.102529.].

The acellular pertussis (aP) vaccine has increasingly replaced the whole-cell pertussis (wP) vaccine due to its superior safety profile. However, the aP vaccine is less effective at preventing infection and transmission of Bordetella pertussis, highlighting the need for more effective aP vaccines. Current aP vaccines do not elicit the robust cellular immunity necessary to eliminate intracellular bacteria and do not induce sufficient mucosal immunity to prevent bacterial colonization in the upper respiratory tract. Incorporating novel adjuvants represents a promising avenue for the future development of pertussis vaccines. Nevertheless, there remains a significant gap in understanding the application of novel adjuvants. In this article, we summarize the currently approved pertussis vaccines, focusing on the types of antigens and adjuvants used, and discuss the mechanisms of novel adjuvants. This provides valuable insights into the roles of adjuvants in pertussis vaccines, laying a foundation for designing next-generation pertussis vaccines with improved adjuvant systems.

Although proven to be promising for CO 2 capture from flue gas, maintaining superior separation efficiency of CO 2 ‐philic membranes under widely differing humidities remains highly challenging to date. Targeting high‐efficiency and humidity‐resistant flue gas separation, in this study, a multi‐scale structure optimization protocol is pioneered to fabricate highly (200)‐oriented 55 nm‐thick MIL‐140A membrane. On one hand, employing ʟ‐histidine as modulator retards crystallization kinetics and inhibits multilamellar‐stacking crystal growth, warranting formation of MIL‐140A nanosheets and ultrathin oriented membrane; on the other hand, bulky ʟ‐histidine segments coordinates in MIL‐140A framework not only induced distorted pore configuration toward precise discrimination of CO 2 from N 2 but also serves as reactive‐carriers toward CO 2 ‐facilitated diffusion. Relying on facilitated diffusion mechanism, the CO 2 /N 2 selectivity of obtained MIL‐140A membrane reached 79.0 under high‐humid environments, which is 32.1% higher than that in dry environments; moreover, the membrane exhibited stable CO 2 /N 2 separation performance over a wide humidity range due to its intrinsic hydrophobicity, showing great promise in practical flue gas separation.

Surfactant replacement has been studied as a supportive therapy for managing COVID-19-induced acute respiratory distress syndrome. The clinical applications require biophysical understanding of the molecular mechanisms behind SARS-CoV-2-induced surfactant inhibition. Although SARS-CoV-2 is known to attack alveolar type II epithelial cells, it is unknown whether the virus can directly interact with the pulmonary surfactant film adsorbed at the alveolar surface. The virus utilizes its spike (S) protein, consisting of two functional subunits (S1 and S2), to bind to the host cell membrane and mediate subsequent membrane fusion. We hypothesize that these two subunits may differentially interact with pulmonary surfactant, resulting in distinct effects on surfactant inhibition. The biophysical impact of recombinant S1 and S2 subunit proteins on a bovine-extracted natural pulmonary surfactant film was investigated with combined constrained drop surfactometry and atomic force microscopy. Our findings revealed that the S2 subunit, in contrast to the S1 subunit, selectively induces surfactant inhibition, evidenced by its capacity in reducing dynamic surface activity and causing domain fusion in surfactant monolayers. These results contribute novel insights into the biophysical mechanisms underlying surfactant inhibition in SARS-CoV-2-induced acute respiratory distress syndrome and may hold translational implications for advancing surfactant therapy to manage COVID-19.

Organoids are three-dimensional, self-organizing microtissues obtained from stem cells, primary tissues, or patient-derived tumors through in vitro culture, which are invaluable models for studying organ development, disease mechanism modelling, drug screening, and regenerative medicine. Nanomaterials, with unique surface properties and excellent biocompatibility, have emerged as powerful tools in biomedical research, particularly in the fields of organoid construction (e.g., microenvironment modulation and advanced engineering cultivation) and cryopreservation (e.g., intracellular delivery, nanowarming, and ice-inhibition). This review comprehensively discusses the recent advancements in nanomaterial-based strategies for organoid fabrication and cryopreservation while addressing their challenges such as biocompatibility, scalability and long-term safety. Finally, we envision the prospects in the development of advanced nanomaterial-based platforms for organoid construction and cryopreservation with higher biocompatibility and standardization.

The preferred method for Escherichia coli genome editing relies on Cas9 from Streptococcus pyogenes (SpCas9) and λ-Red recombinase. Although SpCas9 is currently the most active RNA-guided DNA endonuclease, a significant number of escapers are often observed, making it inefficient across different sites, particularly when inserting large fragments. In this study, we identified two RAGATH RNA-associated DNA endonucleases (RADs) derived from IS607 transposons. Both of them exhibited high cleavage activity in E. coli. When combined with λ-Red recombinase, they achieved editing efficiencies approaching 100%. Even at target sites where SpCas9 exhibited low editing efficiency, RADs maintained efficiencies ranging from 57% to 94%. Moreover, RADs exhibited higher efficiencies in inserting large fragments in certain cases compared to SpCas9. Taken together, these RAD-based genome editing tools provide viable alternatives to SpCas9, particularly for challenging targets and/or large fragment insertions.

Collective motion, exemplified by swarming insects, flocking birds, and bacterial colonies, emerges from the synchronized actions and velocity adjustments of individual units. Inspired by these natural phenomena, researchers have sought to replicate and harness collective behavior in swarms of self-propelled micro/nanomotors. This endeavor is of great importance for the development of multicomponent adaptive systems. Comprising numerous interacting elements, collective swarms exhibit emergent behaviors that capitalize on multi-motor cooperation, enabling highly efficient, flexible, and robust group performance. Understanding and engineering these behaviors are essential for translating them into practical applications. This review examines the driving forces underlying collective motion, outlines various modes of swarm formation across different dimensions, and highlights representative applications. By linking driving forces, system dimensionality, and functional implementation, it provides a comprehensive perspective on this field.

An attractive strategy in cancer cell therapy is to employ motile nanoparticles that can actively search for their target. Herein, we introduce mannosylated compartmentalized cross-linked enzyme-driven nanomotors (c-CLEnM), which exhibit specific and efficient targeting of Hep G2 cells through elevated autonomous motion. In this design, we constructed biodegradable bowl-shaped stomatocytes encapsulating the enzymes glucose oxidase (GOx) and catalase (CAT) within their nanocavity. A subsequent enzyme crosslinking reaction was performed to guarantee their stability. Furthermore, the c-CLEnM were surface modified with a mannose-functional glycopolymer, enabling binding with receptors expressed on Hep G2 cells. Interestingly, the targeting ligands on the nanomotors not only improved their specificity toward cancer cells but also enhanced motility. Compared to the non-mannosylated nanomotors, mannosylated c-CLEnM exhibited enhanced motion and higher targeting efficiency to cells in glucose-containing ionic environments. The unexpected acceleration in speed resulted from the surface modification of these nanomotors with a glycopolymer layer, which increased the zeta potential and created a shielding effect that mitigated the influence of the surrounding ions. This nanomotor design highlights the synergistic effect of functional glycopolymer modification on cellular uptake, adding an additional level of control to nanomotors for application in cancer therapy.