Skin Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
Biomaterials and Tissue Engineering Department, Stem Cell Division, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
Biomed Res Int. 2021 May 24;2021:5597758. doi: 10.1155/2021/5597758. eCollection 2021.
Investigating the viability and proliferative rates of fibroblast cells on human amniotic membrane (HAM) as a scaffold will be an important subject for further research. The aim of this study was to assess the fibroblast viability seeded on acellular HAM, since foreskin neonatal allogenic fibroblasts seeded on HAM accelerate the wound healing process.
Fibroblasts were retrieved from the foreskin of a genetically healthy male infant, and we exploited AM of healthy term neonates to prepare the amniotic scaffold for fibroblast transfer. After cell culture, preparation of acellular HAM, and seeding of cells on HAM based on the protocol, different methods including 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), 4',6-Diamidino-2-phenylindole dihydrochloride (DAPI), and propidium iodide (PI) staining were employed for assessment of fibroblast viability on HAM.
Based on the results obtained from the DAPI and PI staining, the percentage of viable cells in the former staining was clearly higher than that of the dead cells in the latter one. The results of DAPI and PI staining were in accordance with the findings of MTT assay, confirming that fibroblasts were viable and even proliferate on HAM.
Our findings showed the viability of fibroblasts seeded on the acellular HAM using MTT assay, DAPI, and PI staining; however, this study had some limitations. It would be an interesting subject for future research to compare the viability and proliferation rate of fibroblasts seeded on both cellular and acellular HAM.
研究成纤维细胞在人羊膜（HAM）作为支架的活力和增殖率将是进一步研究的重要课题。本研究旨在评估接种在去细胞化 HAM 上的成纤维细胞活力，因为新生儿包皮同种异体成纤维细胞接种在 HAM 上会加速伤口愈合过程。
从遗传健康男性婴儿的包皮中提取成纤维细胞，并利用健康足月新生儿的 AM 制备用于成纤维细胞转移的羊膜支架。根据方案进行细胞培养、制备去细胞化 HAM 和细胞接种后，采用 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）、4',6-二脒基-2-苯基吲哚二盐酸盐（DAPI）和碘化丙啶（PI）染色等多种方法评估 HAM 上成纤维细胞的活力。
根据 DAPI 和 PI 染色的结果，前者染色的活细胞百分比明显高于后者染色的死细胞百分比。DAPI 和 PI 染色的结果与 MTT 检测结果一致，证实成纤维细胞在 HAM 上是有活力的，甚至可以增殖。
我们的研究结果显示，MTT 检测、DAPI 和 PI 染色法均可检测到接种在去细胞化 HAM 上的成纤维细胞的活力；然而，本研究存在一些局限性。比较接种在细胞和去细胞化 HAM 上的成纤维细胞的活力和增殖率将是未来研究的一个有趣课题。
