Isolation and characterization of a new SHED cell line as a standard source for stem cell research and clinical translation

IF 2.5 4区生物学 Q1 ANATOMY & MORPHOLOGY Tissue & cell Pub Date : 2025-04-01 Epub Date: 2024-11-30 DOI:10.1016/j.tice.2024.102649

Niloufar Hosseini , Ezatolah Kazeminejad , Morteza Oladnabi , Ayyoob Khosravi

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Abstract

Background and aims

Stem cells from human exfoliated deciduous teeth (SHED) are multi-potent mesenchymal stem/stromal cells (MSCs) and are inspected a favorable, non-invasive source beneficial to stem cell-mediated regeneration of damaged tissues. Our aim was to establish and characterize a non-immortalized SHED cell line as an accessible resource and novel platform for stem cell research and tissue regeneration studies.

Methods

A Healthy exfoliated deciduous molar was extracted from a 12-year-old girl and shipped to an animal cell culture laboratory. Outgrowing primary cells from explanted small pulp tissues were monitored daily and characterized after passage 3 both morphologically and functionally. The SHED cell line was characterized by calculation of doubling time, cytogenetic analyses, STR analysis, adherence to cell culture flasks under standard cell culture media, and immunophenotypic analysis of specific MSC markers (CD90+, CD73+, CD34- and CD45-) using flow cytometry method. Differentiation potential to osteoblast, adipocyte, and chondrocyte was evaluated under standard differentiation media Expression of OCT-4 and NANOG genes was also assessed using RT-PCR method.

Results

After the third day, SHED cells were visible. SHED cells were subcultured when they reached 90 % confluence after approximately 17 days. The doubling time of SHED cells was forty seven hours. SHED immunophenotyping showed the high expression level of CD90 (99.2 %) and CD73 (45.9 %), and approximately no expression of CD34 (0.079 %) and CD45 (0.19 %). The human origin, female gender and chromosomal normality of SHED cells was confirmed by cytogenetic analysis.

The STR matching analysis showed that SHED cells are well-identified and authentic. No genetic instability and cross-contamination were observed in SHED cells.

Conclusions

This study provides a new SHED cell line with a normal karyotype and all the characteristics of MSCs, which can be used as a favorable model cell line in biomedical research and a promising source for clinical translation.

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一个新的SHED细胞系的分离和特性作为干细胞研究和临床翻译的标准来源。

背景和目的：人脱落乳牙（SHED）干细胞是一种多能间充质干细胞/基质细胞（MSCs），被认为是干细胞介导的损伤组织再生的有利的非侵入性来源。我们的目标是建立和表征一种非永生化的SHED细胞系，作为干细胞研究和组织再生研究的可获取资源和新平台。方法：从一名12岁女孩身上提取一颗脱落的健康乳牙，并送往动物细胞培养实验室。从小牙髓组织中取出的原代细胞每天进行监测，并在传代3后进行形态学和功能表征。利用流式细胞术对SHED细胞系进行倍增时间计算、细胞遗传学分析、STR分析、在标准细胞培养基下与细胞培养瓶的粘附以及特异性MSC标志物（CD90+、CD73+、CD34-和CD45-）的免疫表型分析。在标准分化培养基下评估成骨细胞、脂肪细胞和软骨细胞的分化潜力，并采用RT-PCR法评估OCT-4和NANOG基因的表达。结果：第3天可见SHED细胞。约17天后，当细胞融合度达到90% %时进行传代培养。SHED细胞倍增时间为47小时。SHED免疫表型分析显示CD90（99.2 %）和CD73（45.9 %）高表达，CD34（0.079 %）和CD45（0.19 %）几乎不表达。细胞遗传学分析证实了SHED细胞的人类起源、女性性别和染色体正常。STR匹配分析表明，SHED细胞被很好地识别和真实。在SHED细胞中未观察到遗传不稳定性和交叉污染。结论：本研究提供了一种新的SHED细胞系，它具有正常核型和间充质干细胞的所有特征，可作为生物医学研究的良好模型细胞系和临床翻译的良好来源。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Tissue & cell 医学-解剖学与形态学

CiteScore

3.90

自引率

0.00%

发文量

234

期刊介绍： Tissue and Cell is devoted to original research on the organization of cells, subcellular and extracellular components at all levels, including the grouping and interrelations of cells in tissues and organs. The journal encourages submission of ultrastructural studies that provide novel insights into structure, function and physiology of cells and tissues, in health and disease. Bioengineering and stem cells studies focused on the description of morphological and/or histological data are also welcomed. Studies investigating the effect of compounds and/or substances on structure of cells and tissues are generally outside the scope of this journal. For consideration, studies should contain a clear rationale on the use of (a) given substance(s), have a compelling morphological and structural focus and present novel incremental findings from previous literature.