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Tobacco mild green mosaic virus (TMGMV)-like nanocarriers were designed for gene delivery to plant cells. High aspect ratio TMGMVs were coated with a polycationic biopolymer, poly(allylamine) hydrochloride (PAH), to generate highly charged nanomaterials (TMGMV-PAH; 56.20 ± 4.7 mV) that efficiently load (1:6 TMGMV:DNA mass ratio) and deliver single-stranded and plasmid DNA to plant cells. The TMGMV-PAH were taken up through energy-independent mechanisms in Arabidopsis protoplasts. TMGMV-PAH delivered a plasmid DNA encoding a green fluorescent protein (GFP) to the protoplast nucleus (70% viability), as evidenced by GFP expression using confocal microscopy and Western blot analysis. TMGMV-PAH were inactivated (iTMGMV-PAH) using UV cross-linking to prevent systemic infection in intact plants. Inactivated iTMGMV-PAH-mediated pDNA delivery and gene expression of GFP in vivo was determined using confocal microscopy and RT-qPCR. Virus-like nanocarrier-mediated gene delivery can act as a facile and biocompatible tool for advancing genetic engineering in plants.

In this study, a small library of prednisolone and budesonide conjugated polyethylenimine (PEI) derivatives was prepared. Since nuclear entry is a major barrier for plasmid DNA (pDNA) delivery, it is hypothesized that glucocorticoid-conjugated PEIs that employ their specific receptors to enter the cell nucleus may increase the gene transfer ability of PEI. In this study, prednisolone and budesonide were conjugated onto low and high molecular weight PEI (1.8 and 25 kDa) with different degrees of substitution via different linkers. The glucocorticoid-polyethyleneimine (GC–PEI)/pDNA polyplexes were prepared, and their biophysical characteristics, plasmid delivery, and cell-induced toxicity were evaluated. The polyplexes demonstrated a substantial ability to condense plasmid DNA and the protection of nucleic acids against enzymatic degradation. The polyplexes’ size was in the range of 50–250 nm with zeta potentials between 10 and 30 mV. Interestingly, prednisolone-conjugated low molecular weight PEI could enhance the level of desired protein expression by up to 4 folds with no toxicity compared to the parent unmodified polymer. The results of this study show a promising approach to preparing gene carrier vehicles based on low molecular weight PEI with low toxicity. [ABSTRACT FROM AUTHOR]

Background: Gene therapy is garnering increasing support due to its potential for a "once-delivered, lifelong benefit." The limitations of traditional gene delivery methods have spurred the advancement of bionanomaterials. Despite this progress, a thorough analysis of the evolution, current state, key contributors, focal studies, and future directions of nanomaterials in gene delivery remains absent. Methods: This study scrutinizes articles from the Web of Science, spanning 1 January 2 000, to 31 December 2023, employing various online tools for analysis and visualization. Results: The 21st century has witnessed consistent growth in scholarly work in this domain globally, with notable contributions from China and the US. At the same time, the Chinese Academy of Sciences (CAS), Harvard University, and Massachusetts Institute of Technology (MIT) have emerged as the most productive institutions, with CAS's academician Weihong Tan becoming the field's leading author. While drug delivery and nanoparticles (NPs) have been central themes for two decades, the research focus has shifted from modifying NPs and ultrafine particles to exploring polymer-hybrid NPs, mRNA vaccines, immune responses, green synthesis, and CRISPR/Cas tools. Conclusions: This shift marks the transition from nanomaterials to bionanomaterials. The insights provided by this research offer a comprehensive overview of the field and valuable guidance for future investigations. GRAPHICAL ABSTRACT [ABSTRACT FROM AUTHOR]

Large volume bone defects that do not spontaneously heal despite surgical stabilization ("critical-sized" defects) remain a challenge to treat clinically. Recent research investigating bone regenerative implants made from 3D printed materials have shown promise as a potential alternative to current treatment methods, such as autografting, allografting, and multi-step surgical interventions. Recent work has shown that implanting 3D printed calcium phosphate cement (CPC) scaffolds loaded with bone morphogenetic protein-2 (BMP-2) can provide a one-step surgical intervention that has similar bone healing outcomes to a popular two-step intervention: the Masquelet technique. The aim of this study was to investigate whether a 3D printed CPC scaffold loaded with a lyophilized polyplex gene-delivery formulation could serve as an alternative to loading BMP-2 protein onto such scaffolds. We 3D printed CPC scaffolds, hardened them with multiple methods, and explored the impact of these hardening methods on surface texture, mechanical strength, osteogenic differentiation, and ion flux. We then gene-activated these materials with cationic polyplexes containing plasmid DNA encoding reporter genes to investigate transfection from the gene-activated scaffolds. We found that incubating CPC scaffolds in aqueous solutions after initial hardening in a humid environment could enhance scaffold mechanical strength (compressive strength of 21.28 MPa vs. 6.54 MPa) and osteogenic differentiation. We also found that when we increased the total surface area of the CPC material exposed to polyplex solutions, there was a reduction in transfection via adsorption of polyplexes to the CPC surface. This study shows that 3D printed, gene-activated CPC scaffolds are a promising avenue for future exploration in the field of bone regeneration, though the level of gene expression induced by the scaffolds must be improved. [ABSTRACT FROM AUTHOR]

Plant genetic engineering is an evolving discipline that contributes to crop improvement by introducing desirable traits into crop plants, such as improved yield, enhanced nutrition value, and resistance to biotic and abiotic stresses. Plant transformation is carried out in two steps: Gene delivery into the plant cell and regeneration of the plant cell into the fertile plant. Gene delivery is an essential step in plant genetic transformation, and it is largely plant species-specific. Based on the mode of delivery the conventional plant gene delivery methods are divided into three main categories: biological (Agrobacterium-mediated transformation), physical (biolistic and electroporation), and chemical (Polyethylene glycol mediated and liposome-mediated gene delivery). Apart from species constraints, these methods have unique advantages and limitations, including random gene integration, low gene transfer efficiency, tissue damage, united gene alterations, time-consuming and labor-intensive plant regeneration protocols. Recent advancements in nanotechnology have introduced novel gene-delivery systems, utilizing micro and nanoparticles, which can overcome many limitations of conventional plant gene delivery methods by exhibiting superior transformation efficiency, demonstrate compatibility with biological systems, offer protection to different cargoes, and hold significant capability for enhancing plant regeneration. Nanoparticles are well recognized for its flexible size, shape, and cargo-binding properties, which enable them to surpass defensive primary cell wall barrier and it can be a promising candidate for plant gene delivery applications. However, delivering the nanoparticles and cargo complexes into plants is a critical step of the gene delivery process, and have not been thoroughly explored. In this review, we provide comprehensive insights into nano-delivery systems and detailed methods of introducing nanoparticle complexes into plant tissues. Further, we also discuss techniques such as syringe infiltration, vacuum infiltration, biolistic methods, magnetofection, ultrasound-mediated delivery, passive diffusion, cellular uptake, and spray method. This review serves as a valuable resource for advancing plant gene transformation using nanoparticles, offering guidance on the most effective delivery methods to enhance plant genetic engineering outcomes. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) have emerged as potential alternatives to conventional antibiotics due to their novelty and multiple mechanisms of action. Because they are peptides, AMPs are amenable to bioengineering and suitable for cloning and expression at large production scales. However, the efficient delivery of AMPs is an unaddressed issue, particularly due to their large size, possible toxicities, and the development of adverse immune responses. Here, we have reviewed the possibilities of adeno-associated virus (AAV)-based localized gene delivery of AMPs for the treatment of infectious diseases with a special focus on respiratory infections. By discussing the gene delivery mechanism of AAV and the accompanying technical and therapeutic challenges with AMPs, we describe a foundation that emphasizes the use of viral vector-based gene delivery of AMPs for disease treatment. [ABSTRACT FROM AUTHOR]

Alzheimer's disease is the most common form, accounting for 60–70% of 55 million dementia cases. Even though the precise pathophysiology of AD is not completely understood, clinical trials focused on antibodies targeting aggregated forms of β amyloid (Aβ) have demonstrated that reducing amyloid plaques can arrest cognitive decline in patients in the early stages of AD. In this study, we provide an overview of current research and innovations for controlled release from nano-biomaterial-assisted chimeric antigen receptor macrophage (CAR-M) therapeutic strategies targeted at AD. Nano-bio materials, such as iron-oxide nanoparticles (IONPs), can be made selectively (Hp-Hb/mannose) to bind and take up Aβ plaques like CAR-M cells. By using nano-bio materials, both the delivery and stability of CAR-M cells in brain tissue can be improved to overcome the barriers of the BBB and enhance therapeutic effects. By enhancing the targeting capabilities and stability of CAR-M cells, mRNA-loaded nano-biomaterials can significantly improve the efficacy of immunotherapy for plaque reduction in AD. This novel strategy holds promise for translating preclinical successes into clinical applications, potentially revolutionising the management of AD. [ABSTRACT FROM AUTHOR]

Nanoparticle (NP)-mediated gene delivery offers a promising alternative to traditional methods in plant biotechnology, facilitating genetic transformations with enhanced precision and efficiency. This review discusses key factors influencing NP efficacy, including plant cell wall composition, DNA/NP ratios, exposure time, cargo loading, and post-transformation assessments. We explore the challenges of NP cytotoxicity, transformation efficiency, and regeneration while addressing environmental impacts and regulatory considerations. We emphasize the potential for stimulus-responsive NPs and scalable delivery methods to optimize gene editing in agriculture. [ABSTRACT FROM AUTHOR]

Background: Human cytomegalovirus (CMV) is the most common cause of viral congenital infections worldwide. The development of effective vaccines against human CMV infection and disease is a high priority. Attenuated Salmonella are attractive oral vaccine vectors against human diseases because they can be administrated orally. Methods: In this study, an attenuated Salmonella strain was generated as an oral vaccine vector for the delivery and expression of the M78 protein of murine cytomegalovirus (MCMV). Using the MCMV infection of mice as the CMV infection model, we characterized the immune responses and protection induced by the constructed Salmonella-based vaccine. Results: The generated Salmonella-based vaccine, v-M78, which contained an M78 expression plasmid construct, carried out gene transfer efficiently for M78 expression and showed little pathogenicity and virulence in mice. In orally vaccinated mice, v-M78 induced anti-MCMV serum IgG and mucosal IgA responses and also elicited anti-MCMV T cell responses. Furthermore, mice immunized with v-M78 were protected from intraperitoneal and intranasal challenges with MCMV. The v-M78 vaccination reduced the titers of the challenged viruses in spleens, livers, lungs, and salivary glands. Conclusions: These results provide the first direct evidence that a Salmonella-based vaccine expressing M78 elicits strong humoral and cellular immune responses and induces immune protection against MCMV infection. Furthermore, our study demonstrates the potential of using Salmonella-based oral vaccines against CMV infection. [ABSTRACT FROM AUTHOR]

As a popular tool for gene function characterization and gene therapy, RNA interference (RNAi)-based gene silencing has been increasingly explored for potential applications to control invasive species. At least two major hurdles exist when applying this approach to invasive plants: (1) the design and screening of species- and gene-specific biomacromolecules (i.e., gene-silencing agents or GSAs) made of DNA, RNA, or peptides that can suppress the expression of target genes efficiently, and (2) the delivery vehicle needed to penetrate plant cell walls and other physical barriers (e.g., leaf cuticles). In this study, we investigated the cell-penetrating peptide (CPP)-mediated delivery of multiple types of GSAs (e.g., double-stranded RNA (dsRNA), artificial microRNA (amiRNA), and antisense oligonucleotide (ASO)) to knock down a putative phytoene desaturase (PDS) gene in the invasive common reed (Phragmites australis spp. australis). Both microscopic and quantitative gene expression evidence demonstrated the CPP-mediated internalization of GSA cargos and transient suppression of PDS expression in both treated and systemic leaves up to 7 days post foliar application. Although various GSA combinations and application rates and frequencies were tested, we observed limitations, including low gene-silencing efficiency and a lack of physiological trait alteration, likely owing to low CPP payload capacity and the incomplete characterization of the PDS-coding genes (e.g., the recent discovery of two PDS paralogs) in P. australis. Our work lays a foundation to support further research toward the development of convenient, cost-effective, field-deployable, and environmentally benign gene-silencing technologies for invasive P. australis management. [ABSTRACT FROM AUTHOR]

In recent years, factors such as the postponement of childbearing and the relaxation of the childbearing policy have led to an increase in the proportion of cesarean sections and other intrauterine surgeries among pregnant women, further increasing the incidence of uterine scars. Currently, there is a lack of effective clinical treatment methods for uterine scars. In this study, a suture loaded with gene medicine was designed for the repair of uterine scars. Specifically, the non-viral vector Lipo8000 was first used to form a complex solution with the plasmid TGF-β3. Then, it was mixed and adsorbed with the surgical sutures pretreated with recombinant human type III collagen (RhCol III). In vitro experiments confirmed that RhCol III and the plasmid were successfully loaded onto the sutures and could be released and expressed. In vivo experiments were carried out using a rat model simulating uterine scars. The section results showed that compared with the scar model group, the expression level of TGF-β3 in the RhCol III+TGF-β3 group increased by 39%, the expression level of TGF-β1 decreased by 62.8%, and the fibrosis rate decreased by 16.8%, which has a positive effect on the prevention of uterine scars. This study integrates the therapeutic medicine into the sutures, ensuring that the medicine can come into contact with the wound site after suturing. Moreover, RhCol III and the gene medicine work synergistically to promote the repair of uterine wounds. [ABSTRACT FROM AUTHOR]

Schematic representation of co-delivery liposomes for the treatment of malignant tumors. [Display omitted] • Efficient co-loading of PROTAC and siRNA into novel liposomes. • Enhancement of PROTAC drug solubility and endocytosis via a novel lipid nanodelivery system. • Synergistic action of PROTAC and siRNA for enhanced degradation of BCL-xL target protein. • Augmented tumor therapeutic effects through synergistic interaction of PROTAC and siRNA. Targeted elimination of damaged or overexpressed proteins within the tumor serves a pivotal role in regulating cellular function and restraining tumor cell growth. Researchers have been striving to identify safer and more effective methods for protein removal. Here, we propose the synergistic employment of a small molecule degrading agent (PROTAC) and siRNA to attain enhanced protein clearance efficiency and tumor therapeutic effects. Co-delivery liposomes were prepared to facilitate the efficient encapsulation of PROTAC and siRNA. Specifically, the cationic liposome significantly improved the solubility of the insoluble PROTAC (DT2216). The cationic polymer (F-PEI) achieved efficient encapsulation of the nucleic acid drug, thereby promoting endocytosis and enhancing the therapeutic impact of the drug. Both in vivo and in vitro experiments demonstrated remarkable degradation of target proteins and inhibition of tumor cells by the co-delivery system. In conclusion, the co-delivery liposomes furnished a nano-delivery system proficient in effectively encapsulating both hydrophilic and hydrophobic drugs, thereby presenting a novel strategy for targeted combination therapy in treating tumors. [ABSTRACT FROM AUTHOR]