Narayanan Kannan Badri, Bhaskar Rakesh, Han Sung Soo
School of Chemical Engineering, Yeungnam University, 280 Daehak-Ro, Gyeongsan, Gyeongbuk 38541, Republic of Korea; Research Institute of Cell Culture, Yeungnam University, 280 Daehak-Ro, Gyeongsan, Gyeongbuk 38541, Republic of Korea.
School of Chemical Engineering, Yeungnam University, 280 Daehak-Ro, Gyeongsan, Gyeongbuk 38541, Republic of Korea; Research Institute of Cell Culture, Yeungnam University, 280 Daehak-Ro, Gyeongsan, Gyeongbuk 38541, Republic of Korea.
Tissue Cell. 2025 Apr;93:102734. doi: 10.1016/j.tice.2025.102734. Epub 2025 Jan 10.
Numerous naturally occurring biological structures have inspired the development of innovative biomaterials for a wide range of applications. Notably, the nanotopographical architectures found in natural materials have been leveraged in biomaterial design to enhance cell adhesion and proliferation and improve tissue regeneration for biomedical applications. In this study, we fabricated three-dimensional (3D) chitin-glucan micro/nanofibrous fungal-based spheres coated with collagen (type I) to mimic the native extracellular matrix (ECM) microenvironment. These collagen-coated fungal nano/microfibrous spheres (C-FNS) were utilized to construct 3D scaffolded spheroids of human fibroblasts through suspension culture for tissue engineering and regenerative medicine. The particle sizes of C-FNS ranged from 1.4 to 3.25 µm (average: 2.27 ± 0.38 µm), with a porosity of 81.17 %. Field emission-scanning electron microscopy (FE-SEM) revealed that C-FNS comprised continuous chitin-glucan fibers with an average diameter of 363 ± 61 nm (range: 203-512 nm), exhibiting a highly interconnected structure. The reduced arithmetic average roughness (Ra) and root mean square roughness (Rq) values of C-FNS compared to uncoated FNS suggested that collagen coating reduced surface roughness, resulting in a smoother surface that enhanced hydrophilicity, crucial for mammalian cell adhesion and spheroid formation. Moreover, the in vitro cytocompatibility of C-FNS with fibroblasts was evaluated using a resazurin-based PrestoBlue assay, which demonstrated a time-dependent increase in the metabolic activity of C-FNS/fibroblast spheroids during suspension culture for up to 14 days. FE-SEM images of C-FNS/fibroblast spheroids further revealed enhanced adhesion and proliferation of fibroblasts on the nano/microfibrous mycelial architecture, accompanied by the secretion of ECM components and formation of multilayered cell sheets over the 14-day culture period. Similarly, an assessment of the hemocompatibility of C-FNS with erythrocytes revealed the non-hemolytic properties of the biomaterial. Overall, the interaction between collagen-coated fungal chitin-glucan nano/microfibrous structures and mammalian cells holds significant potential for the development of novel, sustainable biomaterials with tailored properties for a myriad of biomedical applications, including tissue engineering, regenerative medicine, drug screening, and wound healing.
许多天然存在的生物结构激发了用于广泛应用的创新生物材料的开发。值得注意的是，天然材料中发现的纳米拓扑结构已被用于生物材料设计，以增强细胞粘附和增殖，并改善生物医学应用中的组织再生。在本研究中，我们制备了涂有I型胶原蛋白的三维（3D）几丁质-葡聚糖微/纳米纤维真菌基球体，以模拟天然细胞外基质（ECM）微环境。这些胶原蛋白包被的真菌纳米/微纤维球体（C-FNS）用于通过悬浮培养构建人成纤维细胞的3D支架球体，用于组织工程和再生医学。C-FNS的粒径范围为1.4至3.25μm（平均：2.27±0.38μm），孔隙率为81.17%。场发射扫描电子显微镜（FE-SEM）显示，C-FNS由连续的几丁质-葡聚糖纤维组成，平均直径为363±61nm（范围：203-512nm），呈现高度互连的结构。与未包被的FNS相比，C-FNS的算术平均粗糙度（Ra）和均方根粗糙度（Rq）值降低，表明胶原蛋白包被降低了表面粗糙度，从而产生了更光滑的表面，增强了亲水性，这对哺乳动物细胞粘附和球体形成至关重要。此外，使用基于刃天青的PrestoBlue测定法评估了C-FNS与成纤维细胞的体外细胞相容性，结果表明在长达14天的悬浮培养期间，C-FNS/成纤维细胞球体的代谢活性呈时间依赖性增加。C-FNS/成纤维细胞球体的FE-SEM图像进一步显示，在14天的培养期内，成纤维细胞在纳米/微纤维菌丝结构上的粘附和增殖增强，同时伴随着ECM成分的分泌和多层细胞片的形成。同样，对C-FNS与红细胞的血液相容性评估显示了该生物材料的非溶血特性。总体而言，胶原蛋白包被的真菌几丁质-葡聚糖纳米/微纤维结构与哺乳动物细胞之间的相互作用对于开发具有定制特性的新型可持续生物材料具有巨大潜力，可用于多种生物医学应用，包括组织工程、再生医学、药物筛选和伤口愈合。
