Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets
Healthy human skin tissue is often used as a control for comparison to diseased skin in patients with skin pathologies, including skin cancers or other inflammatory conditions such as atopic dermatitis or psoriasis. Although non-affected skin from these patients is a more appropriate choice for comp...
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| Format: | Article |
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MDPI AG
2024-01-01
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| Series: | Cells |
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| Online Access: | https://www.mdpi.com/2073-4409/13/3/266 |
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| author | Terri Clister Rosalyn M. Fey Zachary R. Garrison Cristian D. Valenzuela Anna Bar Justin J. Leitenberger Rajan P. Kulkarni |
| author_facet | Terri Clister Rosalyn M. Fey Zachary R. Garrison Cristian D. Valenzuela Anna Bar Justin J. Leitenberger Rajan P. Kulkarni |
| author_sort | Terri Clister |
| collection | DOAJ |
| description | Healthy human skin tissue is often used as a control for comparison to diseased skin in patients with skin pathologies, including skin cancers or other inflammatory conditions such as atopic dermatitis or psoriasis. Although non-affected skin from these patients is a more appropriate choice for comparison, there is a paucity of studies examining such tissue. This lack is exacerbated by the difficulty of processing skin tissue for experimental analysis. In addition, choosing a processing protocol for skin tissue which preserves cell viability and identity while sufficiently dissociating cells for single-cell analysis is not a trivial task. Here, we compare three digestion methods for human skin tissue, evaluating the cell yield and viability for each protocol. We find that the use of a sequential dissociation method with multiple enzymatic digestion steps produces the highest cell viability. Using single-cell sequencing, we show this method results in a relative increase in the proportion of non-antigen-presenting mast cells and CD8 T cells as well as a relative decrease in the proportion of antigen-presenting mast cells and KYNU<sup>+</sup> CD4 T cells. Overall, our findings support the use of this sequential digestion method on freshly processed human skin samples for optimal cell yield and viability. |
| first_indexed | 2024-03-08T03:59:18Z |
| format | Article |
| id | doaj.art-1fade0e795d8433eaebe867b965f2f5d |
| institution | Directory Open Access Journal |
| issn | 2073-4409 |
| language | English |
| last_indexed | 2024-03-08T03:59:18Z |
| publishDate | 2024-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Cells |
| spelling | doaj.art-1fade0e795d8433eaebe867b965f2f5d2024-02-09T15:09:48ZengMDPI AGCells2073-44092024-01-0113326610.3390/cells13030266Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell SubsetsTerri Clister0Rosalyn M. Fey1Zachary R. Garrison2Cristian D. Valenzuela3Anna Bar4Justin J. Leitenberger5Rajan P. Kulkarni6Department of Dermatology, Oregon Health & Science University, Portland, OR 97239, USADepartment of Dermatology, Oregon Health & Science University, Portland, OR 97239, USADepartment of Dermatology, Oregon Health & Science University, Portland, OR 97239, USADepartment of Surgery, Oregon Health & Science University, Portland, OR 97239, USADepartment of Dermatology, Oregon Health & Science University, Portland, OR 97239, USADepartment of Dermatology, Oregon Health & Science University, Portland, OR 97239, USADepartment of Dermatology, Oregon Health & Science University, Portland, OR 97239, USAHealthy human skin tissue is often used as a control for comparison to diseased skin in patients with skin pathologies, including skin cancers or other inflammatory conditions such as atopic dermatitis or psoriasis. Although non-affected skin from these patients is a more appropriate choice for comparison, there is a paucity of studies examining such tissue. This lack is exacerbated by the difficulty of processing skin tissue for experimental analysis. In addition, choosing a processing protocol for skin tissue which preserves cell viability and identity while sufficiently dissociating cells for single-cell analysis is not a trivial task. Here, we compare three digestion methods for human skin tissue, evaluating the cell yield and viability for each protocol. We find that the use of a sequential dissociation method with multiple enzymatic digestion steps produces the highest cell viability. Using single-cell sequencing, we show this method results in a relative increase in the proportion of non-antigen-presenting mast cells and CD8 T cells as well as a relative decrease in the proportion of antigen-presenting mast cells and KYNU<sup>+</sup> CD4 T cells. Overall, our findings support the use of this sequential digestion method on freshly processed human skin samples for optimal cell yield and viability.https://www.mdpi.com/2073-4409/13/3/266tissue digestionscRNA-seqskin cancermelanomaimmune cell compositionT cells |
| spellingShingle | Terri Clister Rosalyn M. Fey Zachary R. Garrison Cristian D. Valenzuela Anna Bar Justin J. Leitenberger Rajan P. Kulkarni Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets Cells tissue digestion scRNA-seq skin cancer melanoma immune cell composition T cells |
| title | Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets |
| title_full | Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets |
| title_fullStr | Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets |
| title_full_unstemmed | Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets |
| title_short | Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets |
| title_sort | optimization of tissue digestion methods for characterization of photoaged skin by single cell rna sequencing reveals preferential enrichment of t cell subsets |
| topic | tissue digestion scRNA-seq skin cancer melanoma immune cell composition T cells |
| url | https://www.mdpi.com/2073-4409/13/3/266 |
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