Nagaraj Athmika, Subramaniyan Yuvarajan, Surya Suprith, Rekha Punchappady Devasya
Division of Microbiology and Biotechnology, Yenepoya Research Centre, Yenepoya (Deemed to be University), University Road, Deralakatte, Mangalore, 575018, India.
Advanced Surgical Skill ENhancement Division (ASSEND), Yenepoya (Deemed to Be University), Deralakatte, Mangalore, 575018, India.
Appl Biochem Biotechnol. 2024 Nov;196(11):8190-8213. doi: 10.1007/s12010-024-04966-8. Epub 2024 May 3.
Bacterial exopolysaccharides (EPS) are an emerging class of biopolymers with extensive applications in different fields due to their versatile physico-chemical and biological properties. The role of EPS in healing of different wound types is gaining interest in the tissue engineering sector. Burn is one of the devitalizing injuries that causes greater physical harm and can be fatal. Appropriate treatment modalities have to be followed for faster healing outcomes and to minimize the risk. In this study, a bacterial EPS (EPS-H29) from the marine bacterium Halomonas malpeensis YU-PRIM-29 was used to treat the burn wound in vivo. The biochemical and structural characterizations of EPS-H29 were carried out using standard methods. In addition, FE-SEM, conformational, rheological, and HP-GPC analyses were carried out. In vitro biocompatibility of EPS-H29 was studied in human dermal fibroblasts (HDFs) and keratinocytes (HaCaT). Scratch assay was used to study the wound healing in vitro. For in vivo evaluation, burn wound (second-degree) was created on Wistar albino rats and treated with EPS-H29 along with appropriate control groups. The total sugar and protein contents of EPS-H29 were 72.0 ± 1.4% and 4.0 ± 0.5%, respectively, with a molecular weight of 5.2 × 10 Da. The lyophilized samples exhibited porous surface features, and in solution, it showed triple helical conformation and shear thickening behavior. In vitro cell-based assays showed biocompatibility of EPS-H29 up to 200 μg/mL concentration. At a concentration up to 50 μg/mL, EPS-H29 promoted cell proliferation. Significant increase in the HDF cell migration was evident with EPS-H29 (15 μg/mL) treatment in vitro and induced significantly higher (p ≤ 0.0001) closure of the scratch area (90.3 ± 1.1%), compared to the control (84.3 ± 1.3%) at 24 h. Enhanced expression of Ki-67 was associated with the cell proliferative activities of EPS-H29. The animals treated with EPS-H29 showed improved healing of burn wounds with significantly higher wound contraction rate (80.6 ± 9.4%) compared to the positive control (54.6 ± 8.0%) and untreated group (49.2 ± 3.7%) with histopathological evidence of epidermal tissue formation at 15 days of treatment. These results demonstrate the biocompatibility and burn wound healing capability of EPS-H29 and its potential as an effective topical agent for the burn wound care.
细菌胞外多糖(EPS)是一类新兴的生物聚合物,由于其多样的物理化学和生物学特性,在不同领域有着广泛的应用。EPS在不同类型伤口愈合中的作用在组织工程领域正引起越来越多的关注。烧伤是一种极具破坏性的损伤,会造成更大的身体伤害甚至可能致命。必须采用适当的治疗方式以实现更快的愈合效果并将风险降至最低。在本研究中,来自海洋细菌马尔佩恩斯盐单胞菌YU-PRIM-29的一种细菌EPS(EPS-H29)被用于体内治疗烧伤创面。使用标准方法对EPS-H29进行了生化和结构表征。此外,还进行了场发射扫描电子显微镜(FE-SEM)、构象、流变学和高效凝胶渗透色谱(HP-GPC)分析。在人皮肤成纤维细胞(HDFs)和角质形成细胞(HaCaT)中研究了EPS-H29的体外生物相容性。划痕试验用于研究体外伤口愈合情况。为了进行体内评估,在Wistar白化大鼠身上制造了二度烧伤创面,并用EPS-H29进行治疗,同时设置了适当的对照组。EPS-H29的总糖和蛋白质含量分别为72.0±1.4%和4.0±0.5%,分子量为5.2×10 Da。冻干样品呈现出多孔表面特征,在溶液中表现出三螺旋构象和剪切增稠行为。基于细胞的体外试验表明,EPS-H29在浓度高达200μg/mL时具有生物相容性。在浓度高达50μg/mL时,EPS-H29促进细胞增殖。在体外,用EPS-H29(15μg/mL)处理后,HDF细胞迁移显著增加,在24小时时诱导的划痕区域闭合率(90.3±1.1%)明显高于对照组(84.3±1.3%)(p≤0.0001)。Ki-67的表达增强与EPS-H29的细胞增殖活性相关。与阳性对照组(54.6±8.0%)和未治疗组(49.2±3.7%)相比,用EPS-H29治疗的动物烧伤创面愈合情况改善,伤口收缩率显著更高(80.6±9.4%),在治疗15天时具有表皮组织形成的组织病理学证据。这些结果证明了EPS-H29的生物相容性和烧伤创面愈合能力及其作为烧伤创面护理有效局部用药的潜力。