Search Results (127)

Polyethylenimine (PEI) is a cationic polymer with a high density of amine groups suitable for strong electrostatic interactions with biological molecules to preserve their bioactivities during encapsulation and after delivery for biomedical applications. This review provides a comprehensive overview of PEI as a drug and gene carrier, describing its polymerization methods in both linear and branched forms while highlighting the processing methods to manufacture PEIs into drug carriers, such as nanoparticles, coatings, nanofibers, hydrogels, and films. These various PEI carriers enable applications in non-viral gene and small molecule drug deliveries. The structure–property relationships of PEI carriers are discussed with emphasis on how molecular weights, branching degrees, and surface modifications of PEI carriers impact biocompatibility, transfection efficiency, and cellular interactions. While PEI offers remarkable potential for drug and gene delivery, its clinical translation remains limited by challenges, including cytotoxicity, non-degradability, and serum instability. Our aim is to provide an understanding of PEI and the structure–property relationships of its carrier forms to inform future research directions that may enable safe and effective clinical use of PEI carriers for drug and gene delivery. Full article

Background: Transfection is vital for gene therapy, mRNA treatments, CAR-T cell therapy, and regenerative medicine. While viral vectors are effective, non-viral systems like lipid nanoparticles (LNPs) offer safer, more flexible alternatives. This work explores emerging non-viral transfection technologies to improve delivery efficiency and therapeutic outcomes. Methods: This review synthesizes the current literature and recent advancements in non-viral transfection technologies. It focuses on the mechanisms, advantages, and limitations of various delivery systems, including lipid nanoparticles, biodegradable polymers, electroporation, peptide-based carriers, and microfluidic platforms. Comparative analysis was conducted to evaluate their performance in terms of transfection efficiency, cellular uptake, biocompatibility, and potential for clinical translation. Several academic search engines and online resources were utilized for data collection, including Science Direct, PubMed, Google Scholar Scopus, the National Cancer Institute’s online portal, and other reputable online databases. Results: Non-viral systems demonstrated superior performance in delivering mRNA, siRNA, and antisense oligonucleotides, particularly in clinical applications. Biodegradable polymers and peptide-based systems showed promise in enhancing biocompatibility and targeted delivery. Electroporation and microfluidic systems offered precise control over transfection parameters, improving reproducibility and scalability. Collectively, these innovations address key challenges in gene delivery, such as stability, immune response, and cell-type specificity. Conclusions: The continuous evolution of transfection technologies is pivotal for advancing gene and cell-based therapies. Non-viral delivery systems, particularly LNPs and emerging platforms like microfluidics and biodegradable polymers, offer safer and more adaptable alternatives to viral vectors. These innovations are critical for optimizing therapeutic efficacy and enabling personalized medicine, immunotherapy, and regenerative treatments. Future research should focus on integrating these technologies to develop next-generation transfection platforms with enhanced precision and clinical applicability. Full article

Gene therapy is a groundbreaking strategy in regenerative medicine, enabling precise cellular behavior modulation for tissue repair. In situ nucleic acid delivery systems aim to directly deliver nucleic acids to target cells or tissues to realize localized genetic reprogramming and avoid issues like donor cell dependency and immune rejection. The key to success relies on biomaterial-engineered delivery platforms that ensure tissue-specific targeting and efficient intracellular transport. Viral vectors and non-viral carriers are strategically modified to enhance nucleic acid stability and cellular uptake, and integrate them into injectable or 3D-printed scaffolds. These scaffolds not only control nucleic acid release but also mimic native extracellular microenvironments to support stem cell recruitment and tissue regeneration. This review explores three key aspects: the mechanisms of gene editing in tissue repair; advancements in viral and non-viral vector engineering; and innovations in biomaterial scaffolds, including stimuli-responsive hydrogels and 3D-printed matrices. We evaluate scaffold fabrication methodologies, nucleic acid loading–release kinetics, and their biological impacts. Despite progress in spatiotemporal gene delivery control, challenges remain in balancing vector biocompatibility, manufacturing scalability, and long-term safety. Future research should focus on multifunctional “smart” scaffolds with CRISPR-based editing tools, multi-stimuli responsiveness, and patient-specific designs. This work systematically integrates the latest methodological advances, outlines actionable strategies for future investigations and advances clinical translation perspectives beyond the existing literature. Full article

Neuropathic pain is a significant global clinical issue that poses substantial challenges to both public health and the economy due to its complex underlying mechanisms. It has emerged as a serious health concern worldwide. Recent studies involving dorsal root ganglion (DRG) stimulation have provided strong evidence supporting its effectiveness in alleviating chronic pain and its potential for sustaining long-term pain relief. In addition to that, there has been ongoing research with clinical evidence relating to the role of small non-coding ribonucleic acids known as microRNAs in regulating gene expressions affecting pain signals. The signal pathway involves alterations in neuronal excitation, synaptic transmission, dysregulated signaling, and subsequent pro-inflammatory response activation and pain development. When microRNAs are dysregulated in the dorsal root ganglia neurons, they polarize macrophages from anti-inflammatory M2 to inflammatory M1 macrophages causing pain signal generation. By reversing this polarization, a therapeutic activity can be induced. However, the direct delivery of these nucleotides has been challenging due to limitations such as rapid clearance, degradation, and reduction in half-life. Therefore, safe and efficient carrier vehicles are fundamental for microRNA delivery. Here, we present a comprehensive analysis of miRNA-based nano-systems for chronic neuropathic pain, focusing on their impact in dorsal root ganglia. This review provides a critical evaluation of various delivery platforms, including viral, polymeric, lipid-based, and inorganic nanocarriers, emphasizing their therapeutic potential as well as their limitations in the treatment of chronic neuropathic pain. Innovative strategies such as hybrid nanocarriers and stimulus-responsive systems are also proposed to enhance the prospects for clinical translation. Serving as a roadmap for future research, this review aims to guide the development and optimization of miRNA-based therapies for effective and sustained neuropathic pain management. Full article

Linear DNA constructs are used in gene delivery and therapy application due to their capacity of integration into the mammalian genome, offering stable transgene expression. Compared to circular plasmids, linear DNA also has the advantage that its dimension and steric hindrance are directly correlated to the length of the nucleotide chain. These considerations make linear DNA an effective choice for gene delivery pilot studies, where formulations and transfection efficiency calculations are studied considering the nucleic acid dimensions. Meanwhile, the development of DNA–chitosan nanoparticles (NPs) has gained significant interest for their potential in nucleic acid delivery, especially as non-viral gene delivery systems and for embedding linear DNA fragments, as well as gene delivery to the lung. This study explored an easy polyelectrolyte complexing preparation of linear DNA-loaded chitosan nanoparticles. Among the different formulations of nanoparticles prepared, the optimal one exhibited a size of approximately 290 nm, an encapsulation efficiency of 86% and a zeta potential of 25 mV. Additionally, this study examined how the concentration of DNA in solution influenced nanoparticle formation, encapsulation efficiency and particle size. In particular, transient transfection of the chitosan–linear DNA fragment complex, encoding for green fluorescent protein (GFP), was conducted in human pulmonary distal lung cells (NCI-H441 cells), demonstrating successful cellular internalization and protein expression. These studies highlight the potential of DNA–chitosan NPs in nucleic acid delivery, particularly for pulmonary applications. Future works will focus on formulating the achieved carrier into an inhalable dosage form to improve its translational application. Full article

CRISPR/Cas9 has revolutionized genome editing by enabling precise and efficient genetic modifications across multiple biological systems. Despite its growing therapeutic potential, key challenges remain in mitigating off-target effects, minimizing immunogenicity, and improving the delivery of CRISPR components into target cells. This review provides an integrated analysis of physical, viral, and non-viral delivery systems, highlighting recent advances in the use of lipid nanoparticles, polymeric carriers, and hybrid platforms. We also examine an often overlooked factor: the aggregation behavior of the Cas9 protein, which may interfere with cellular uptake, the encapsulation efficiency, and nuclear localization. By comparing delivery platforms and their reported editing outcomes, we identify critical physicochemical parameters that influence therapeutic success. Finally, we propose standardized methods to assess Cas9 encapsulation and aggregation and discuss translational barriers such as manufacturing scalability and regulatory requirements. These insights aim to guide the development of safer and more effective CRISPR/Cas9-based therapies. Full article

Clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR-Cas9), an emerging gene-editing technology, has recently gained rapidly increasing attention. However, the lack of efficient delivery vectors to deliver CRISPR-Cas9 to specific cells or tissues has hindered the translation of this biotechnology into clinical applications. Chemically synthesized nanoparticles (NPs), as attractive non-viral delivery platforms for CRISPR-Cas9, have been extensively investigated because of their unique characteristics, such as controllable size, high stability, multi-functionality, bio-responsive behavior, biocompatibility, and versatility in chemistry. In this review, the key considerations for the precise design of chemically synthesized-based nanoparticles include efficient encapsulation, cellular uptake, the targeting of specific tissues and cells, endosomal escape, and controlled release. We discuss cutting-edge strategies to integrate chemical modifications into non-viral nanoparticles that guide the CRISPR-Cas9 genome-editing machinery to specific edits. We also highlighted the rationale of intelligent nanoparticle design. In particular, we have summarized promising functional groups and molecules that can effectively optimize carrier function. In addition, this review focuses on advances in the widespread application of NPs delivery in the biomedical fields to promote the development of safe, specific, and efficient NPs for delivering CRISPR-Cas9 systems, providing references for accelerating their clinical translational applications. Full article

Background/Objectives: This study aims to gain insights into the antimicrobial and antiherpetic activity of hyperforin-rich Hypericum perforatum L. (HP) extract using nanostructured lipid carriers (NLCs) as delivery platforms. Methods: Two established NLC specimens, comprising glyceryl behenate and almond oil or borage oil, and their extract-loaded counterparts (HP-NLCs) were utilized. Their minimal bactericidal/fungicidal concentrations (MBC; MFC) were investigated against Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 10145, Klebsiella pneumoniae ATCC 10031, and Candida albicans ATCC 10231. The anti-herpesvirus (HSV-1) potential was evaluated concerning antiviral and virucidal activity and impact on viral adsorption. Results: The borage oil-based extract-loaded nanodispersion (HP-NLC2) exhibited pronounced microbicidal activity against S. aureus (MBC 6.3 mg/mL), K. pneumoniae (MBC 97.7 µg/mL), and C. albicans (MFC < 48.8 µg/mL), unlike the almond oil-containing sample (HP-NLC1), which showed only weak inhibition of the fungal growth. HP-NLC2 was found to be less cytotoxic and to suppress HSV-1 replication slightly more than HP-NLC1, but generally, the effects were weak. Neither the empty lipid nanoparticles nor the HP extract-loaded carriers expressed activity against E. coli, P. aeruginosa, the HSV-1 extracellular virions, or viral adhesion. Conclusions: It could be concluded that both HP-NLC samples revealed only minor antiherpetic potential of the hyperforin-rich extract, but HP-NLC2 demonstrated significant antibacterial and antimycotic activity. Therefore, the latter was featured as a more convenient HP-carrier system for nano-designed dermal pharmaceutical formulations. Such a thorough investigation of hyperforin-determined anti-HSV-1 effects and antibacterial and antimycotic properties, being the first of its kind, contributes to the fundamental knowledge of HP and reveals new perspectives for the utilization, limitations, and therapeutic designation of its non-polar components. Full article

Background/Objectives: Non-viral vectors have gained recognition for their ability to enhance the safety of gene delivery processes. Among these, polyethyleneimine (PEI) stands out as the most widely utilized cationic polymer due to its accessibility. Traditional methods of modifying PEI, such as ligand conjugation, chemical derivatization, and cross-linking, are associated with intricate preparation procedures, limited transfection efficiency, and suboptimal biocompatibility. Methods: In this investigation, enhanced transfection efficiency was achieved through the straightforward physical blending of PEI carriers with spermine. Results: Transfection assays explored the maximal enhancement potential conferred by spermine, alongside further methodological refinements aimed at optimizing transfection efficacy, showcasing a potential increase of up to 40.7%. Through the comparison of different addition sequences of spermine, the optimal complex PEI/Spermine/DNA for transfection efficiency was selected. Characterization of PEI/Spermine/DNA revealed that, compared to PEI/DNA, its particle size increased to approximately 150 nm. Molecular dynamics simulation results revealed that spermine can enhance the interaction between PEI and DNA, thereby forming a system with lower energy and greater stability. Mechanistic inquiries studies also disclosed that spermine augments the endosomal escape capability of PEI carriers without altering pathways involved in the cellular uptake of gene nanoparticles, thereby facilitating heightened gene expression. Conclusions: PEI-Sper emerges as a promising non-viral vector for gene delivery, distinguished by its simplicity in preparation, cost-effectiveness, and superior transfection efficiency. Full article

Background: The Epstein–Barr virus (EBV) infects more than 90 percent of the human population. In pediatric patients, the innate immune response against EBV primary infection plays a key role. Monocytes and macrophages can have distinct functions depending on the microenvironment surrounding them. At least three monocyte subpopulations can be differentiated depending on membrane protein expression: classical (C, CD14++CD16−), intermediate (I, CD14++CD16+), and non-classical (NC, CD14+CD16++). They also modulate T and B lymphocyte activation/inhibition through the expression of costimulatory molecules such as CD80, CD86, and PD-L1. Yet, little is known about monocytes’ role in EBV infection. Methods: Peripheral blood and tonsil biopsies of EBV primary infected (PI) patients, healthy carriers (HCs), and patients undergoing reactivation (R) were studied. Results: Classical monocytes prevailed in all infectious statuses. Tonsillar CD163 positively correlated with CD163 expression in NC monocytes in HCs. PD-L1+ cells in the tonsil positively correlated with PD-L1 expression in NC monocytes. LMP-1 viral latent protein presented a positive correlation with PD-L1, CD163, and CD206 expression in the NC subpopulation. Conclusions: Our results evidence the predominant role of I and NC monocytes’ response against EBV infection. Furthermore, the viral oncoprotein LMP-1 could be involved in the expression of regulatory proteins in I and NC monocytes. Full article

Among the physical decontamination methods, treatment with ultraviolet (UV) radiation is a suitable means of preventing viral infections. Mercury vapor lamps (254 nm) used for room decontamination are potentially damaging to human skin (radiation) and harmful to the environment (mercury). Therefore, other UV-C wavelengths (100–280 nm) may be effective for virus inactivation on skin without damaging it, e.g., far-UV-C radiation with a wavelength of 233 nm, which is absorbed in the outer layer of the skin and thus does not reach the deeper layers of the skin. For room disinfection, 275 nm UV-C LED lamps could be a more environmentally friendly alternative, since toxic mercury is avoided. A carrier test using multiple viruses was used to determine the TCID₅₀/mL value on stainless steel, PVC, and glass carriers. In addition to the inactivation kinetics (233 nm), the necessary UV-C dose for 4 lg inactivation (275 nm) was investigated. The impact of irradiance on the inactivation efficacy was also assessed. The inactivation of the viruses was a function of the radiation dose. UV-C-radiation at 233 nm (80 mJ/cm²) inactivated from 1.49 ± 0.08 to 4.28 ± 0.18 lg depending on the virus used. To achieve a 4 lg inactivation (275 nm) for enveloped viruses, doses of up to 70 mJ/cm² (SuHV-1) were sufficient. For non-enveloped viruses, a maximum dose of 600 mJ/cm² (MS2) was necessary. Enveloped viruses were inactivated with lower doses compared to non-enveloped viruses. Higher radiation doses were required for inactivation at 275 nm in comparison to 254 nm. A more environmentally friendly alternative to mercury vapor lamps is available with 275 nm LED emitters. Radiation at 233 nm could serve as an additional prophylactic or therapeutic measure for virus inactivation in direct contact with human skin. Full article

Interleukin-12 (IL-12) is considered to be a promising cytokine for enhancing an antitumor immune response; however, recombinant IL-12 has shown significant toxicity and limited efficacy in early clinical trials. Recently, many strategies for delivering IL-12 to tumor tissues have been developed, such as modifying IL-12, utilizing viral vectors, non-viral vectors, and cellular vectors. Previous studies have found that the fusion of IL-12 with extracellular matrix proteins, collagen, and immune factors is a way to enhance its therapeutic potential. In addition, studies have demonstrated that viral vectors are a good platform, and a variety of viruses such as oncolytic viruses, adenoviruses, and poxviruses have been used to deliver IL-12—with testing previously conducted in various cancer models. The local expression of IL-12 in tumors based on viral delivery avoids systemic toxicity while inducing effective antitumor immunity and acting synergistically with other therapies without compromising safety. In addition, lipid nanoparticles are currently considered to be the most mature drug delivery system. Moreover, cells are also considered to be drug carriers because they can effectively deliver therapeutic substances to tumors. In this article, we will systematically discuss the anti-tumor effects of IL-12 on its own or in combination with other therapies based on different delivery strategies. Full article

Lipid nanoparticles (LNPs) have emerged as leading non-viral carriers for messenger RNA (mRNA) delivery in clinical applications. Overcoming challenges in safe and effective mRNA delivery to target tissues and cells, along with controlling release from the delivery vehicle, remains pivotal in mRNA-based therapies. This review elucidates the structure of LNPs, the mechanism for mRNA delivery, and the targeted delivery of LNPs to various cells and tissues, including leukocytes, T-cells, dendritic cells, Kupffer cells, hepatic endothelial cells, and hepatic and extrahepatic tissues. Here, we discuss the applications of mRNA–LNP vaccines for the prevention of infectious diseases and for the treatment of cancer and various genetic diseases. Although challenges remain in terms of delivery efficiency, specific tissue targeting, toxicity, and storage stability, mRNA–LNP technology holds extensive potential for the treatment of diseases. Full article

Disease’s severity, mortality rates, and common failures to achieve clinical improvement during the unprecedented COVID-19 pandemic exposed the emergency need for new antiviral therapeutics with higher efficacy and fewer adverse effects. This study explores the potential to encapsulate multi-component plant extracts in liposomes as optimized delivery systems and to verify if they exert inhibitory effects against human seasonal betacoronavirus OC43 (HCoV-OC43) in vitro. The selection of Sambucus nigra, Potentilla reptans, Allium sativum, Aesculus hippocastanum, and Glycyrrhiza glabra L. plant extracts was based on their established pharmacological and antiviral properties. The physicochemical characterization of extract-loaded liposomes was conducted by DLS and electrokinetics. Encapsulated amounts of the extract were evaluated based on the total flavonoid content (TFC) and total polyphenol content (TPC) by colorimetric methods. The BALB 3T3 neutral red uptake (NRU) phototoxicity/cytotoxicity assay was used to estimate compounds’ safety. Photo irritation factors (PIFs) of the liposomes containing extracts were <2 which assigned them as non-phototoxic substances. The antiviral capacities of liposomes containing medicinal plant extracts against HCoV-OC43 were measured by the cytopathic effect inhibition test in susceptible HCT-8 cells. The antiviral activity increased by several times compared to “naked” extracts’ activity reported previously. A. hippocastanum extract showed 16 times higher inhibitory properties reaching a selectivity index (SI) of 58.96. Virucidal and virus-adsorption effects were investigated using the endpoint dilution method and ∆lgs comparison with infected and untreated controls. The results confirmed that nanoparticles do not directly affect the viral surface or cell membrane, but only serve as carriers of the active substances and the observed protection is due solely to the intracellular action of the extracts. Full article

Genetic constructs must be delivered selectively to target tissues and intracellular compartments at the necessary concentrations in order to achieve the maximum therapeutic effect in gene therapy. Development of targeted carriers for non-viral delivery of nucleic acids into cells, including those in muscle, which is one of the most challenging tissues to transfect in vivo, remains a topical issue. We have studied ternary complexes of plasmid DNA and an arginine–histidine-rich peptide-based carrier coated with a glutamate–histidine-rich polymer bearing skeletal muscle targeting peptide (SMTP) for the gene delivery to muscle tissue. The relaxation of the ternary complexes after polyanion treatment was assessed using the ethidium bromide displacement assay. The developed polyplexes were used to transfect C2C12 myoblasts in full-media conditions, followed by analysis of their toxic properties using the Alamar Blue assay and expression analysis of lacZ and GFP reporter genes. After delivering plasmids containing the GFP and lacZ genes into the femoral muscles of mdx mice, which are model of Duchenne muscular dystrophy, GFP fluorescence and β-galactosidase activity were detected. We observed that the modification of ternary polyplexes with 10 mol% of SMTP ligand resulted in a 2.3-fold increase in lacZ gene expression when compared to unmodified control polyplexes in vivo. Thus, we have demonstrated that the developed DNA/carrier complexes and SMTP-modified coating are nontoxic, are stable against polyanion-induced relaxation, and can provide targeted gene delivery to muscle cells and tissues. The results of this study are useful for a range of therapeutic applications, from immunization to amelioration of inherited neuromuscular diseases. Full article