Biomaterial and Tissue Engineering Laboratory, Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati , Guwahati 781039, Assam, India.
Biological and Chemical Sciences Section, Life Sciences Division, Institute of Advanced Study in Science and Technology (IASST) , Guwahati 781035, Assam, India.
ACS Appl Mater Interfaces. 2016 Aug 24;8(33):21236-49. doi: 10.1021/acsami.6b08285. Epub 2016 Aug 10.
An osteoarthritis pandemic has accelerated exploration of various biomaterials for cartilage reconstruction with a special emphasis on silk fibroin from mulberry (Bombyx mori) and non-mulberry (Antheraea assamensis) silk worms. Retention of positive attributes of the agarose standard and nullification of its negatives are central to the current agarose/silk fibroin hydrogel design. In this study, hydrogels of mulberry and non-mulberry silk fibroin blended with agarose were fabricated and evaluated in vitro for two weeks for cartilaginous tissue formation. The fabricated hydrogels were physicochemically characterized and analyzed for cell viability, proliferation, and extra cellular matrix deposition. The amalgamation of silk fibroin with agarose impacted the pore size, as illustrated by field emission scanning electron microscopy studies, swelling behavior, and in vitro degradation of the hydrogels. Fourier transform infrared spectroscopy results indicated the blend formation and confirmed the presence of both components in the fabricated hydrogels. Rheological studies demonstrated enhanced elasticity of blended hydrogels with G' > G″. Biochemical analysis revealed significantly higher levels of sulfated glycosaminoglycans (sGAGs) and collagen (p ≤ 0.01) in blended hydrogels. More specifically, the non-mulberry silk fibroin blend showed sGAG and collagen content (∼1.5-fold) higher than that of the mulberry blend (p ≤ 0.05). Histological and immunohistochemical analyses further validated the enhanced deposition of sGAG and collagen, indicating maintenance of chondrogenic phenotype within constructs after two weeks of culture. Real-time PCR analysis further confirmed up-regulation of cartilage-specific aggrecan, sox-9 (∼1.5-fold) and collagen type II (∼2-fold) marker genes (p ≤ 0.01) in blended hydrogels. The hydrogels demonstrated immunocompatibility, which was evidenced by minimal in vitro secretion of tumor necrosis factor-α (TNF-α) by murine macrophages. Taken together, the results suggest promising attributes of blended hydrogels and particularly the non-mulberry silk fibroin/agarose blends as alternative biomaterial for cartilage tissue engineering.
骨关节炎的流行加速了对各种生物材料的探索，以用于软骨重建，特别关注来自桑蚕（Bombyx mori）和非桑蚕（Antheraea assamensis）的丝素纤维。保留琼脂糖标准的积极属性并消除其负面属性是当前琼脂糖/丝素纤维水凝胶设计的核心。在这项研究中，制备了桑蚕丝素纤维和非桑蚕丝素纤维与琼脂糖混合的水凝胶，并在体外进行了为期两周的软骨组织形成评估。对制备的水凝胶进行了物理化学特性分析，并对细胞活力、增殖和细胞外基质沉积进行了分析。丝素纤维与琼脂糖的结合影响了孔径，场发射扫描电子显微镜研究、溶胀行为和水凝胶的体外降解表明了这一点。傅里叶变换红外光谱结果表明了混合物的形成，并证实了所制备的水凝胶中两种成分的存在。流变学研究表明，混合水凝胶的弹性增强，G' > G″。生化分析显示，混合水凝胶中硫酸化糖胺聚糖（sGAGs）和胶原蛋白的水平显著升高（p ≤ 0.01）。具体而言，非桑蚕丝素纤维的混合物显示出 sGAG 和胶原蛋白含量（约 1.5 倍）高于桑蚕丝素纤维混合物（p ≤ 0.05）。组织学和免疫组织化学分析进一步验证了 sGAG 和胶原蛋白的沉积增加，表明在培养两周后构建体中保持了软骨表型。实时 PCR 分析进一步证实了软骨特异性聚集蛋白聚糖、sox-9（约 1.5 倍）和胶原蛋白 II（约 2 倍）标记基因的上调（p ≤ 0.01）。水凝胶表现出免疫相容性，这可以通过小鼠巨噬细胞体外分泌的肿瘤坏死因子-α（TNF-α）最小化来证明。总之，这些结果表明混合水凝胶具有有前景的特性，特别是非桑蚕丝素纤维/琼脂糖混合水凝胶作为软骨组织工程的替代生物材料。
