ITGA6+ Human Testicular Cell Populations Acquire a Mesenchymal Rather than Germ Cell Transcriptional Signature during Long-Term Culture
Abstract
:1. Introduction
2. Results
2.1. Derivation of ITGA6+ Testicular Cell Fractions from Mixed Testicular Cell Cultures
2.2. Long-Term In Vitro Propagation of ITGA6 + PTCs Is Correlated with Distinct Transcriptional Changes
2.3. Long-Term Cultured ITGA6 + PTCs Possess Mesenchymal and Fibroblast Related Gene Expression Signatures
2.4. Analysis of Established In Vivo and In Vitro Testicular Cell Type Marker Genes
2.5. Cell Decomposition Analysis Points towards a Decrease in the Proportion of Spermatogonia in ITGA6 + PTCs over Culture Time Accompanied by an Increase in Mesenchymal Cells
3. Discussion
4. Materials and Methods
4.1. Ethics Statement
4.2. Patient Samples
4.3. Live Cell Imaging
4.4. RNA Isolation, Quality Control
4.5. cDNA Amplification
4.6. Library Preparation
4.7. Specification of Testicular Cell Type Marker Genes
4.8. Bioinformatics and Statistical Analyses
4.8.1. Count Data Normalization
4.8.2. Differential Gene Expression Analysis
4.8.3. Gene Set Enrichment Analysis
4.8.4. Cell Mix Decomposition Analysis
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Gene Set | # Genes | Direction | p-Value | FDR 1 |
---|---|---|---|---|
OXFORD_RALA_TARGETS_DN | 9 | Up | 1.79 × 10−6 | 0.0163 |
VERRECCHIA_RESPONSE_TO_TGFB1_C4 | 13 | Up | 1.53 × 10−4 | 0.357 |
VANHARANTA_UTERINE_FIBROID_UP | 41 | Up | 2.29 × 10−4 | 0.357 |
WEBER_METHYLATED_ICP_IN_FIBROBLAST | 17 | Down | 2.30 × 10−4 | 0.357 |
PID_INTEGRIN5_PATHWAY | 16 | Up | 2.35 × 10−4 | 0.357 |
JI_CARCINOGENESIS_BY_KRAS_AND_STK11_DN | 15 | Up | 3.09 × 10−4 | 0.36 |
CLASPER_LYMPHATIC_VESSELS_DURING_METASTASIS_DN | 35 | Up | 3.16 × 10−4 | 0.36 |
ONDER_CDH1_TARGETS_3_UP | 17 | Up | 4.35 × 10−4 | 0.385 |
INGRAM_SHH_TARGETS_DN | 59 | Up | 4.49 × 10-4 | 0.385 |
WEBER_METHYLATED_ICP_IN_SPERM_DN | 11 | Down | 5.71 × 10-4 | 0.385 |
SENESE_HDAC1_TARGETS_DN | 221 | Up | 5.78 × 10−4 | 0.385 |
BROWNE_HCMV_INFECTION_18HR_DN | 147 | Up | 6.47 × 10−4 | 0.385 |
WEBER_METHYLATED_ICP_IN_SPERM_UP | 6 | Down | 7.13 × 10−4 | 0.385 |
PID_INTEGRIN1_PATHWAY | 61 | Up | 7.29 × 10−4 | 0.385 |
ANASTASSIOU_CANCER_MESENCHYMAL_TRANSITION_SIGNATU | 60 | Up | 7.71 × 10−4 | 0.385 |
KEGG_LYSOSOME | 114 | Up | 8.18 × 10−4 | 0.385 |
SUZUKI_AMPLIFIED_IN_ORAL_CANCER | 11 | Up | 8.46 × 10−4 | 0.385 |
REACTOME_GLYCOSPHINGOLIPID_METABOLISM | 29 | Up | 1.00e × 10−3 | 0.388 |
KEGG_GLYCOSPHINGOLIPID_BIOSYNTHESIS_GANGLIO_SERIES | 14 | Up | 1.02 × 10−3 | 0.388 |
BROWNE_HCMV_INFECTION_48HR_DN | 432 | Up | 1.06 × 10−3 | 0.388 |
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Struijk, R.B.; Mulder, C.L.; van Daalen, S.K.M.; de Winter-Korver, C.M.; Jongejan, A.; Repping, S.; van Pelt, A.M.M. ITGA6+ Human Testicular Cell Populations Acquire a Mesenchymal Rather than Germ Cell Transcriptional Signature during Long-Term Culture. Int. J. Mol. Sci. 2020, 21, 8269. https://doi.org/10.3390/ijms21218269
Struijk RB, Mulder CL, van Daalen SKM, de Winter-Korver CM, Jongejan A, Repping S, van Pelt AMM. ITGA6+ Human Testicular Cell Populations Acquire a Mesenchymal Rather than Germ Cell Transcriptional Signature during Long-Term Culture. International Journal of Molecular Sciences. 2020; 21(21):8269. https://doi.org/10.3390/ijms21218269
Chicago/Turabian StyleStruijk, Robert B., Callista L. Mulder, Saskia K. M. van Daalen, Cindy M. de Winter-Korver, Aldo Jongejan, Sjoerd Repping, and Ans M. M. van Pelt. 2020. "ITGA6+ Human Testicular Cell Populations Acquire a Mesenchymal Rather than Germ Cell Transcriptional Signature during Long-Term Culture" International Journal of Molecular Sciences 21, no. 21: 8269. https://doi.org/10.3390/ijms21218269