The development of smart natural delivery systems that can deliver several chemotherapeutics in a controlled manner is a promising idea to enhance cancer treatment efficacy. In this study, we designed and synthesized a smart, dual pH- and thermo-responsive nanocarrier system based on β-CD-g-BCN, which was further functionalized with MSP and copolymerized with NIPAAm and DMAEMA to yield the final β-CD-g-BCN@MSP@PND formulation. The nanocarriers indicated efficient encapsulation of two anticancer agents, MTX and CUR, with encapsulation efficiencies of 64.5% and 87%, respectively. Successful functionalization and thermal stability were validated by characterization using FTIR, FESEM, TEM, DLS, zeta potential, and TGA. The nanoparticles exhibited sharp LCST behavior (~ 40.5 °C), which facilitates temperature-sensitive drug release. Improved drug release under acidic (pH 5.8) and hyperthermic (40 °C) conditions, simulating the tumor, was confirmed by in vitro release studies. Based on cytotoxicity studies, the MTX/CUR-loaded system had strong synergistic effects (CI < 0.1) on MCF-7 cells, while blank nanoparticles showed negligible toxicity. The time-dependent cellular uptake of RhB-labeled nanocarriers was validated by fluorescence microscopy. Overall, the prepared dual-responsive nanocarrier shows great promise for effective breast cancer treatment. However, this study is limited to in vitro evaluations, and further in vivo studies are required to confirm the therapeutic potential of the developed nanocarrier.

The human microbiome provides a robust ecosystem for mucosal barrier function, immune regulation, metabolism, and resistance to pathogen infection. Dysbiosis causes inflammatory bowel disease, oral inflammation, wound damage, skin disease, and cancer-related immune dysregulation. Current probiotic therapy is limited by low survival during storage and administration, low oral bioavailability, short retention time, and insufficient targeting. Probiotic-hydrogel systems combine viable microorganisms with natural or synthetic polymer networks to promote probiotic protection, local retention, and controlled release. They can also incorporate prebiotics, engineered bacteria, oxygen-releasing agents, antioxidants, growth factors, drugs, or advanced manufacturing techniques such as microfluidics and 3D printing. Recent preclinical studies suggest that probiotic-hydrogel can modulate local microbes, suppress pathogens, control inflammation, support barrier repair, and promote tissue regeneration. However, most evidence is far from clinical, and human data are limited to a small number of oral and periodontal applications. In this review, we summarize the design strategies, encapsulation approaches, mechanistic frameworks, disease-based evidence, and translational challenges of probiotic-hydrogel. In particular, we paid attention to evidence level, safety, manufacturing, regulatory classification, product stability, clinical trial design. Such issues need to be well understood before probiotic-hydrogel can move from experimental platforms to clinically applicable microbiome treatments.

Bacterial cancer therapy has emerged as a promising strategy for the local delivery of anticancer agents to solid tumors because bacteria preferentially colonize the tumor microenvironment (TME). However, strategies for exploiting metabolic features of the TME to achieve tumor-selective expression of anticancer agents remain limited. Here, we present a tumor-responsive gene expression system in Escherichia coli for tumor-selective expression of the recombinant immunotoxin psp-TGFa-PE38 (TP), exploiting its intrinsic ability to sense metabolic cues in the TME. To identify such TME-associated metabolic cues, we examined tumor metabolites in CT26 tumor-bearing mice and confirmed that tumors contained > 3-fold lower glucose and > 5-fold higher fumarate levels than those in the liver. In response to this nutrient heterogeneity, E. coli upregulated genes involved in fumarate respiration, including frdA, dctA, dcuB, and aspA, in the DcuSR regulon. aspA was most upregulated (65-fold) in tumors, which is indicative of the strongest activity of its promoter in the TME. We used the aspA promoter (PaspA) as a tumor-specific switch to control TP expression, targeting secreted functional TP and inducing cytotoxicity in vitro. TP was specifically expressed by PaspA in tumors, reducing the tumor size by 2.8-fold and extending survival by 35 days in vivo. Overall, these results suggest that the potential of PaspA as a tumor-specific expression system.

Natural polysaccharides are versatile building blocks for regenerative biomaterials because of their structural diversity, generally favorable biocompatibility and biodegradability, chemical tunability, and capacity for biologically relevant interactions. Advances in biotechnology and materials engineering have expanded their role from primarily structural matrices to multifunctional platforms that can influence cellular, immune, and biochemical processes during tissue repair. This review presents an integrated biological engineering framework linking polysaccharide source and molecular architecture with extraction and purification, chemical functionalization, crosslinking, advanced fabrication, biological recognition, and translational performance. Particular emphasis is placed on structure-function relationships, stereospecific glycan-receptor interactions, immune-instructive biomaterials, protein-mediated cell-material interactions, microbiome-responsive systems, and emerging artificial intelligence-assisted approaches for biomaterial design. The importance of material purity is also critically considered, particularly the potential for residual endotoxin contamination to confound interpretation of immunomodulatory responses and compromise reproducibility. Representative applications in skin, musculoskeletal, cardiovascular, neural, and other regenerative systems illustrate how material composition, scaffold architecture, biofunctionalization, and manufacturing strategy collectively influence biological performance. Finally, challenges associated with scalable manufacturing, batch-to-batch reproducibility, sterilization, storage stability, quality control, regulatory translation, and clinical implementation are examined. By integrating molecular design, biological mechanisms, fabrication, and translational requirements, this review provides engineering principles for the development of adaptive polysaccharide biomaterials and identifies priorities for advancing reproducible and clinically translatable regenerative technologies.

Engineering compact bacterial promoters that integrate two chemical inputs would simplify biosensors, synthetic circuits, and dynamic metabolic control, but such promoters remain difficult to build because a second operator can increase partial-state leak, reduce the fully induced state, and create sequence- or reporter-context effects. Here we engineered 12 compact Marionette-derived two-input promoter architectures in Escherichia coli by combining defined template scaffolds with added operators, then quantified 19 reporter-specific four-state truth tables under a predefined operational criterion for chemical-input AND behaviour. Single-input template-promoter controls in matched reporter contexts helped separate weak parent-scaffold output from effects introduced by the second operator. Nine curated architectures passed in at least one reporter, and selected constructs delivered high-utility AND responses, including [11]/max-off separation up to > 1,367. The full pass-fail dataset showed that a statistically higher [11] state does not necessarily create a useful switch. Practical utility depended mainly on suppressing [10] and [01] leakage, matching the template scaffold to the inserted operator, and controlling long-operator orientation and local sequence context. Reciprocal architectures behaved differently, and 4 of 7 dual-reporter architectures changed operational classification with reporter context. Thus, this work delivers a compact set of candidate Marionette-compatible two-input promoter parts and a source-traceable design framework for engineering bacterial chemical-input AND switches.

The current study presents the development of a Cetuximab (CTX)-functionalized chitosan-polydopamine (CS-PDA) hydrogel co-delivering calcium ions (Ca[2 +]) and Gefitinib (GFT) for EGFR-targeted therapy against esophageal cancer through the induction of Ca[2+] mediated cell death and potential immune modulatory effects. FTIR analysis confirmed the functional groups present in the fabricated hydrogel, while UV-Vis spectroscopy verified successful drug incorporation, and SEM analysis revealed a porous morphology suitable for efficient drug encapsulation. The hydrogel shows high drug loading efficiencies of 78.5 ± 2.3% for Ca[2 +] and 81.6 ± 3.1% for GFT, with pH-responsive drug release reaching up to 70% under acidic conditions. NIR-activated CTX-CS-pDA@Ca[2 +]/GFT hydrogel showed superior cytotoxicity in KYSE-140 cells, with significantly lower IC50 values than free GFT. Further, JC-1 and AO/EB staining confirmed mitochondrial depolarization and apoptosis induction, while flow cytometry demonstrated that CTX-CS-pDA@Ca[2 +]/GFT + NIR markedly induced apoptosis in KYSE-140 cells by combining EGFR inhibition, Ca[2 +] -mediated stress, and NIR-enhanced ROS generation, resulting in reduced cell viability and increased apoptotic populations. Calcium imaging further validated intracellular Ca[2 +] accumulation indicated enhanced Ca[2+]mediated cellular stress in treated cells. In vivo evaluation in a xenograft mouse model revealed tumor inhibition rates of 42%, 24%, 60%, 78%, and 92% for GFT, CS-pDA, CS-pDA@Ca[2 +]/GFT, CTX-CS-pDA@Ca[2 +]/GFT, and CTX-CS-pDA@Ca[2 +]/GFT + NIR, respectively. The NIR-activated CTX-CS-pDA@Ca[2 +]/GFT formulation achieved the most pronounced tumor regression with negligible impact on body weight, confirming its superior therapeutic efficacy and biosafety. Biochemical analyses of serum ALP, SGOT, urea, and creatinine levels confirmed the biocompatibility and safety of the fabricated hydrogel. Histopathological examination of excised tumor tissues supported the induction of apoptosis and tissue necrosis in treated groups. Furthermore, qRT-PCR and Western blot analyses demonstrated upregulation of Bax and cleaved caspase-3 and downregulation of Bcl-2 and EGFR/p-AKT signaling, supporting the dual mechanism of action. Besides, CTX-CS-pDA@Ca[2 +]/GFT + NIR significantly upregulated pro-inflammatory cytokines (IL-6, TNF-α, and IFN-γ), indicating potential immunomodulatory activity associated with cytokine expression changes driven by targeted delivery, Ca[2 +] -mediated stress, and NIR-triggered activation. Overall, this multifunctional hydrogel demonstrates strong potential as a localized and targeted therapeutic strategy for EGFR-overexpressing esophageal cancer through synergistic EGFR inhibition, Ca[2 +] -mediated cytotoxicity, and potential immunomodulatory effects.

Expansion microscopy (ExM) is developing rapidly to explore biological structures, both towards nanoscale imaging and the wider-scale analysis of cells and tissues. ExM is based on embedding samples in swellable hydrogels, which then expand to enable superior microscopy performance with conventional microscopes. However, imaging precision is limited by the stability of the hydrogels, which are typically regarded as less robust than conventional samples. Here, we show that hydrogels based on N, N-dimethylacrylamide (DMAA) remain stable for several years, without a perceivable loss of structure. No special measures of stabilization are necessary for the gel matrix, but post-fixation is required to maintain the integrity of antibody labeling, when post-expansion labeling procedures are employed. Otherwise, antibodies slowly lose epitope binding over years, corrupting both signal intensity and resolution. We conclude that ExM hydrogels are substantially more robust than previously envisioned, enhancing signal intensity and enabling years-long performance, with minimal care. Moreover, this paves the avenue to re-imaging, inter-lab sample exchange, and retrospective analyses, which could be employed in medical applications ranging from diagnostic validation and biomarker discovery to comparative studies across time points or cohorts. This approach extends ExM from a single-use technique to a platform compatible with sustained experimental workflows.

Wounds damage the skin, mucosal membranes, or underlying tissues, disrupting normal structure and function. Natural therapeutic agents, particularly plant extracts, have been widely used in wound management due to their antimicrobial and regenerative properties. Here, we report a novel multifunctional wound dressing by integrating Petroselinum crispum (parsley, PS) extract and zinc ascorbate-based metal-organic frameworks (MOFs) into a polyvinyl alcohol (PVA) nanofibrous matrix. The resulting MOF/PS-loaded PVA nanofibers (NFs) combine biodegradability, biocompatibility, and enhanced bioactivity for wound healing. The optimized formulation (PVA/MOF5/PS30-NFs) exhibited strong antibacterial activity, with inhibition zones of ~ 250 mm[2], total phenolic content of 1.41 mg GAE/g, and antioxidant IC50 values of 15.17 and 11.11 mg/3mL at 2 and 24 h, respectively. Cytocompatibility tests confirmed excellent cell viability (> 100%). In-vitro assays demonstrated enhanced fibroblast migration and edge contraction, with substantial wound closure after 48 h. In-vivo studies showed complete (> 100%) wound closure by day 15, surpassing controls (~ 80%), with the fastest healing trajectory and early acceleration phase observed. To our knowledge, this is the first report of this composition and application. These findings highlight MOF/PS-loaded PVA nanofibrous mats as a promising platform for advanced wound-healing therapies. CLINICAL TRIAL REGISTRATION: Not applicable.

Efficient genetic engineering of lactic acid bacteria remains technically challenging due to their thick peptidoglycan cell wall, low transformation efficiency, strain-specific restriction-modification systems, and sensitivity to Cas9-induced double-strand breaks. In this study, we adapted an established CRISPR/Cas9 approach for the targeted disruption of plnD, a key negative regulatory gene within the plantaricin quorum-sensing network of Lactiplantibacillus plantarum 8P-A3 through extensive optimization of transformation and genome-editing conditions. The genetically modified strain exhibited upregulation of plnA, plnE, and plnF, accompanied by elevated antimicrobial activity. These findings underscore the feasibility of rationally reconfiguring a quorum-sensing-associated regulatory circuit and provide a practical strategy for successful genetic engineering in L. plantarum for elevated bacteriocin production.