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Enhancing the Purity and Intrinsic Properties of Ovine Dermal Papilla Cells through Flow Cytometry Sorting and Cellular Interactions |
Yuan Gou1 
, Tingyan Hu1
, Lijuan Zhu1
, Xiaoyang Lv2,3
, Yutao Li4
, Rui Su5
, Zhenghai Song6
, Shanhe Wang1,3,*
, Wei Sun1,2,3,*
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1Department of College of Animal Science and Technology, Yangzhou University, Yangzhou, China
2Department of Joint International Research Laboratory of Agriculture and Agri-Product Safety, Ministry of Education of China, Yangzhou University, Yangzhou 225009, China
3Department of International Joint Research Laboratory in Universities of Jiangsu Province of China for Domestic Animal Germplasm Resources and Genetic Improvement, Yangzhou University, , Yangzhou 225009, China., China
4Department of CSIRO Agriculture and Food, 306 Carmody Rd, St Lucia, Brisbane, QLD 4067, Australia
5Department of Suzhou Sheep Breeding Farm, Qingyang Road, Jiangling Street, Wujiang District, Suzhou City, Jiangsu 215200, China., China
6Department of Dongshan Animal Epidemic Prevention Station, Wuzhong District, Suzhou, China |
Correspondence:
Shanhe Wang,Email: shanhe12315@163.com
Wei Sun, Tel: ＋86-13952750912, Fax: ＋86-13952750912, Email: dkxmsunwei@163.com
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Received: 15 November 2024 • Revised: 11 February 2025 • Accepted: 1 April 2025 |
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| Abstract |
Objective
Dermal Papilla Cells (DPCs) play a crucial role in regulating hair follicle development and serve as a valuable in vitro model for screening and analyzing the genes associated with this process. However, current methods for isolating ovine DPCs primarily rely on mechanical techniques, which present several limitations. The aim of this study is to establish a method for isolating and culturing ovine DPCs with high purity and retains their intrinsic properties.
Methods
We identified sheep DPC membrane-specific genes using single-cell transcriptomic data, validated by immunostaining and flow cytometry. Antibody-labelled DPCs were isolated, cultured, and assessed via FACS, comparing their purity with conventional mechanical isolation. Mechanically isolated and flow-sorted DPCs were analyzed through agglutination, CCK-8, and EDU staining. Furthermore, we examined the biological properties of isolated DPCs in conditioned media using CCK-8, EDU, and qRT-PCR assays.
Results
PDGFRA was identified as a marker for ovine DPCs. Flow cytometry showed that PDGFRA-labeled DPCs made up 1.54% of the hair follicle cell population, with 1.92% live DPCs obtained via FACS. The isolated DPCs demonstrated agglutination and were positive for ALP, Versican, and α-SMA. Antibody labeling yielded higher DPC purity compared to mechanical isolation, highlighting its efficiency.Accordingly, the addition of conditioned media from mechanically isolated DPCs significantly enhanced agglutination, cell viability, proliferation, and inductive capacity of the sorted DPCs. However, as the number of passages increased, the sorted DPCs demonstrated significant disadvantages in cell agglutination, proliferation rate, and viability compared to mechanically isolated DPCs. Accordingly, the addition of conditioned media from mechanically isolated DPCs significantly enhanced agglutination, cell viability, proliferation, and inductive capacity of the sorted DPCs.
Conclusion
This study highlights the effectiveness of antibody labeling and flow cytometry for isolating functionally pure DPCs, as well as the potential of conditioned media to maintain the functional properties of these cells.
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| Keywords:
Dermal Papilla Cells; PDGFRA; Hair follicle; Flow Cytometry; Cellular Interactions |
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