TIM-3 Expression Is Downregulated on Human NK Cells in Response to Cancer Targets in Synergy with Activation
Abstract
:1. Introduction
2. Materials and Methods
2.1. Isolation and Purification of Human NK Cells from Peripheral Blood
2.2. Cell Culture
2.3. Evaluation of NK Cell Receptor Expression
2.4. Evaluation of Cytokine, Media and Supplement Stimulation of NK Cells
2.5. Cytotoxicity Assay
2.6. Flow Cytometry
2.7. CD107a Degranulation Assay
2.8. IFNγ Secretion Assay
2.9. Bioinformatics Analysis of GBM Patient Data
2.10. Statistical Analysis
2.11. Ethics Statement
3. Results
3.1. NK Cells Downregulate Expression of TIM-3 in Response to Cancer Targets
3.2. Media Composition Contributes to Changes in TIM-3 Expression on Activated NK Cells
3.3. IL-15 Alone Is not Responsible for Downregulation in TIM-3 Expression
3.4. Human Serum Contributes to Changes in TIM-3 Expression on NK Cells
3.5. Downregulation in TIM-3 Expression Correlates to Impaired NK Cell Cytotoxicity
3.6. GBM Patient Data Show Correlation between TIM-3 Expression and NK Cell Function
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
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Name | Composition |
---|---|
RPMI II | RPMI 1640 + 10% FBS + 1%Pen/strep + 50 ng/mL 4-1BBL + 10 ng/mL rhIL-15 + 500 IU/mL rhIL-2 + 25 ng/mL rhIL-21 |
RPMI III | RPMI 1640 + 10% FBS + 1%Pen/strep + 50 ng/mL 4-1BBL + 10 ng/mL rhIL-15 + 500 IU/mL rhIL-2 + 50 ng/mL rhIL-21 |
OpT without (w/o) Supplements | OpTmizer Basal Medium + 0% OpTmizer supplement + 5% hAB serum + 1% Pen/strep + 0.2 mM l-glutamine + 10 ng/mL rhIL-15 + 500 IU/mL rhIL-2 + and 25 ng/mL rhIL-21 |
OpT w/o IL-15 | OpTmizer Basal Medium + 2.6% OpTmizer supplement + 5% hAB serum + 1% Pen/strep + 0.2 mM l-glutamine + 0 ng/mL rhIL-15 + 500 IU/mL rhIL-2 + and 25 ng/mL rhIL-21 |
OpT-RPMI 1640 | RPMI 1640 + 2.6% OpTmizer supplement + 5% hAB serum + 1% Pen/strep + 0.2 mM l-glutamine + 10 ng/mL rhIL-15 + 500 IU/mL rhIL-2 + and 25 ng/mL rhIL-21 |
OpT-FBS | OpTmizer Basal Medium + 2.6% OpTmizer supplement + 5% FBS + 1% Pen/strep + 0.2 mM l-glutamine + 10 ng/mL rhIL-15 + 500 IU/mL rhIL-2 + and 25 ng/mL rhIL-21 |
RPMI-hAB | RPMI 1640 + 10% hAB serum + 1%Pen/strep + 50 ng/mL 4-1BBL + 500 IU/mL rhIL-2 + 50 ng/mL rhIL-21 |
Medium Component | I | II | III | IV | V | VI | VII | VIII |
---|---|---|---|---|---|---|---|---|
RPMI | + | + | + | + | ||||
OpT Basal | + | + | + | + | ||||
OpT Supp | + | + | + | + | ||||
AB Serum | + | + | + | + | + | |||
FBS | + | + | + | |||||
IL-2 | + | + | + | + | + | + | + | + |
IL-15 | + | + | + | + | + | |||
IL-21 | + | + | + | + | + | + | + | + |
TIM-3 MFI decrease | YES | NO | NO | YES | YES | YES | YES | YES |
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Dao, T.N.; Utturkar, S.; Atallah Lanman, N.; Matosevic, S. TIM-3 Expression Is Downregulated on Human NK Cells in Response to Cancer Targets in Synergy with Activation. Cancers 2020, 12, 2417. https://doi.org/10.3390/cancers12092417
Dao TN, Utturkar S, Atallah Lanman N, Matosevic S. TIM-3 Expression Is Downregulated on Human NK Cells in Response to Cancer Targets in Synergy with Activation. Cancers. 2020; 12(9):2417. https://doi.org/10.3390/cancers12092417
Chicago/Turabian StyleDao, Tram N., Sagar Utturkar, Nadia Atallah Lanman, and Sandro Matosevic. 2020. "TIM-3 Expression Is Downregulated on Human NK Cells in Response to Cancer Targets in Synergy with Activation" Cancers 12, no. 9: 2417. https://doi.org/10.3390/cancers12092417