Broad Spectrum Antiviral and Antibiofilm Materials to Treat Hospital Acquired Infections

全世界每年有425万人死于急性呼吸道感染（Mayor， 2010年）。支气管炎和肺炎等感染通常用抗生素治疗，这使得急性呼吸道感染成为抗菌素耐药性（AMR）的主要驱动因素，鉴于耐药病原体的增加（O'Neill, 2016），急性呼吸道感染的治疗迫切需要抗生素的替代品。一氧化氮（NO）是一种有效的广谱抗菌素，是免疫系统对入侵病原体的反应。然而，其临床应用受到其反应性和半衰期短的阻碍，使其递送到感染部位往往具有挑战性。为了解决这一问题，人们开发了NO供体，如n -重氮双酸酯，以提高其稳定性和保质期。这种一氧化氮供体是由胺与一氧化氮气体在高压b[3]下反应形成的，由于其易于制造，通常用于生物材料的研究。明胶纳米颗粒（GNP）由于其生物相容性、可生物降解性和FDA批准而被用于许多药物输送系统。此外，由于明胶是一种多胺，它为n -重氮双酸酯的捆绑提供了多个结合位点。因此，我们制备了装载NO的明胶纳米颗粒（GNP/NO），用于控释NO治疗ARI，并评估了其抗菌效果、生物相容性和NO释放动力学。通过动态光散射（DLS）和扫描电子显微镜（SEM）分析证实，采用两步脱溶法合成GNP，得到了~225nm的均匀球形颗粒。通过傅里叶变换红外光谱（FT-IR）对负载n -重氮二酸酯（GNP/NO）的GNP进行了理化性质评估，证实了纳米颗粒的成功功能化。化学发光是用来确定无负载和释放动力学,证明国民生产总值/没有发布一个初始破裂,紧随其后的是持续释放24 h。国民生产总值的抗菌功效/不测试面板的微生物,铜绿假单胞菌、大肠杆菌、金黄色葡萄球菌和白色念珠菌在24小时内,显著减少(3)登录4 h和24 h后可行的增长对所有微生物测试。随后，采用ISO-10993间接（渗滤液）方案评估GNP/NO对L929和底特律562细胞系的细胞毒性，结果表明GNP/NO维持了70%以上的细胞活力。我们已经证明GNP/NO的成功开发，它在细胞相容性范围内表现出优异的抗菌性能，说明GNP/NO作为ARI治疗的潜力。在确保GNP/NO作为治疗急性呼吸道感染的有效方法的可行性之前，有必要进行进一步的研究以评估其在体内的作用。急性呼吸道感染是世界上第三大死亡原因。BMJ, 2010年。341: p. 6360.2。O'Neill， J.，《全球应对耐药感染：最终报告和建议》。2016.3. Coneski, p.n.，“通过与一氧化氮的厌氧反应竞争性形成n -重氮二酸酯和n -亚硝胺。”Org。让。, 2009年。11(23)。

Acute respiratory infections (ARIs) are responsible for 4.25 million deaths annually worldwide (Mayor, 2010). Infections such as bronchitis and pneumonia are commonly treated with antibiotics, making ARI a primary driver for antimicrobial resistance (AMR), and given the rise in antimicrobial-resistant pathogens (O'Neill, 2016), the treatment of ARIs necessitates a pressing need for alternatives to antibiotics. Nitric oxide (NO) is an effective broad-spectrum antimicrobial produced by the immune system in response to invading pathogens. However, its clinical application is hindered by its reactivity and short half-life, making its delivery to the site of infection often challenging. To address this issue, NO donors, such as N-diazeniumdiolates have been developed to improve its stability and shelf life. This NO donor is formed by the reaction of amines with NO gas under high pressure [3], and owing to its ease of fabrication, it is commonly used in biomaterial research. Gelatin nanoparticles (GNP) are used in many drug delivery systems owing to their biocompatibility, biodegradability and FDA approval. Furthermore, as gelatin is a polyamine, it provides multiple binding sites for the tethering of N-diazeniumdiolates. Therefore, we fabricated NO-loaded gelatin nanoparticles (GNP/NO) for the controlled release of NO for the treatment of ARI, and assessed their antimicrobial efficacy, biocompatibility, and NO release kinetics.A two-step desolvation method was used to synthesise GNP and produced homogenous spherical particles of ~225nm, confirmed by dynamic light scattering (DLS) and scanning electron microscopy (SEM) analysis. The physicochemical properties of the GNP loaded with N-diazeniumdiolates (GNP/NO) were assessed via Fourier-transform infrared spectroscopy (FT-IR), which confirmed the successful functionalisation of the nanoparticles. Chemiluminescence was used to determine the NO payload and release kinetics, demonstrating that GNP/NO released an initial NO burst, followed by a sustained release over 24 h. The antimicrobial efficacy of GNP/NO was tested against a panel of microorganisms, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus and Candida albicans over a 24 h period and revealed a significant reduction (up to 3 log) in viable growth after 4 h and 24 h against all tested microorganisms. Following this, the ISO-10993 indirect (leachate) protocol was used to assess the cytotoxicity of GNP/NO against both the L929 and Detroit 562 cell lines, which showed that GNP/NO maintained over 70% cell viability.We have demonstrated the successful development of GNP/NO, which exhibits excellent antimicrobial properties within a cytocompatible range, illustrating the potential of GNP/NO as a treatment for ARI. Further investigation is warranted to assess the in vivo effects of GNP/NO before ensuring its feasibility for use as an effective treatment for ARIs.References1. Mayor, S., Acute respiratory infections are world's third leading cause of death. BMJ, 2010. 341: p. c6360.2. O'Neill, J., Tackling drug-resistant infections globally: final report and recommendations. 2016.3. Coneski, P.N., 'Competitive formation of N-diazeniumdiolates and N-nitrosamines via anerobic reactions with nitric oxide.' Org. Let., 2009. 11(23).