Dataset Information

Preservation of epithelial progenitor cells from collagenase-digested oral mucosa during ex vivo cultivation.

ABSTRACT: To avoid xenogeneic infection, we report a novel protocol for producing animal-derived component-free oral mucosal epithelial cells (OMECs) sheet for transplantation, in which collagenase was used to replace dispase II/trypsin-EDTA for digesting oral mucosal tissue, and human platelet-derived PLTMax to replace fetal bovine serum. The resulting epithelial aggregates were expanded on de-epithelialized amniotic membranes without 3T3 feeder cells, and serum-free EpiLife was used to reduce contamination by submucosal mesenchymal cells. The OMEC sheets thus generated showed similar positive keratin 3/76-positive and keratin 8-negative staining patterns compared with those generated by the original protocol. Colony formation efficiency assay, BrdU label retention assay, and p63 and p75^NTR immunostaining results indicated that higher proliferative potentials and more progenitor cells were preserved by the modified protocol. TaqMan array analysis revealed that the transcription of integrin-linked kinase (ILK) was up-regulated along with an increase in β-catenin signaling and its downstream cell cycle modulators, cyclin D1 and p27^KIP1. Furthermore, ILK silencing led to the inhibition of nuclear β-catenin accumulation, suppressed p63 expression, and reduced the expression of cyclin D1 and p27^KIP1; these observations suggest that ILK/β-catenin pathway may be involved in cell proliferation regulation during the ex vivo expansion of OMECs for transplantation purposes.

SUBMITTER: Hsueh YJ

PROVIDER: S-EPMC5099970 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Preservation of epithelial progenitor cells from collagenase-digested oral mucosa during ex vivo cultivation.

Hsueh Yi-Jen YJ Huang Shiang-Fu SF Lai Jui-Yang JY Ma Shih-Chieh SC Chen Hung-Chi HC Wu Sung-En SE Wang Tze-Kai TK Sun Chi-Chin CC Ma Kevin Sheng-Kai KS Chen Jan-Kan JK Lai Chyong-Huey CH Ma David Hui-Kang DH

Scientific reports 20161108

To avoid xenogeneic infection, we report a novel protocol for producing animal-derived component-free oral mucosal epithelial cells (OMECs) sheet for transplantation, in which collagenase was used to replace dispase II/trypsin-EDTA for digesting oral mucosal tissue, and human platelet-derived PLTMax to replace fetal bovine serum. The resulting epithelial aggregates were expanded on de-epithelialized amniotic membranes without 3T3 feeder cells, and serum-free EpiLife was used to reduce contaminat ...[more]

PMID: 27824126

Similar Datasets

Project description:BackgroundThere is a substantial need for ex vivo cartilage damage models to assess new emerging cartilage repair strategies. Ex vivo cartilage explant models have the advantages of achieving standardized and reproducible experimental conditions while maintaining the cells in their native tissue environment. This study aimed to establish a bovine cartilage damage model to evaluate the safety and efficacy of novel cartilage repair therapies. We hypothesized that combining transient exposure to matrix-degrading enzymes with mechanical loading on bovine cartilage would simulate cartilage damage.MethodsPrior to mechanical load, bovine osteochondral plugs underwent a brief 5-minutes treatment with collagenase to induce mild cartilage damage by disrupting the collagen network. To induce a moderate cartilage damage, aggrecanase 1 and aggrecanase 2 were additionally applied to the cartilage for 40 min post-collagenase treatment to degrade aggrecan. Data was analyzed using ANOVA or the Friedman test.ResultsObservations revealed a statistically significant loss of sulphated glycosaminoglycan (sGAG) using both Collagenase Treatment (CT) and Collagenase and Aggrecanase Treatment (CAT), while chondrocytes viability was maintained. Both treatments resulted in a significantly elevated release of inflammation markers during the initial two days, including IL6 and nitric oxide. Collagenase treatment also significantly increased neo-epitopes of aggrecan compared to the untreated plugs at day 7, suggesting endogenous aggrecanase activation upon collagen network disruption. The additional effect of mechanical loading on cartilage degeneration was also explored in the CT group. Mildly damaged cartilage treated solely with collagenase could withstand 1 h per day of cyclical load, at 10-20% compression of cartilage thickness combined with interfacial shear at 25 degrees. However, higher compression levels (20-40% of cartilage thickness) with the same shear stress regimen led to a significant increase in surface chondrocyte death, with no evidence of TUNEL staining.ConclusionsThis study establishes a promising model for evaluating cartilage repair strategies, and screening anti-catabolic drugs, particularly overload-related cartilage damage.

Dataset Information

Preservation of epithelial progenitor cells from collagenase-digested oral mucosa during ex vivo cultivation.

Publications

Preservation of epithelial progenitor cells from collagenase-digested oral mucosa during ex vivo cultivation.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets