Related Experiment Video
Updated: Aug 8, 2026
Generation of Human Alloantigen-specific T Cells from Peripheral Blood
Published on: November 21, 2014
Activation of human peripheral blood T cells does not lead to increased P-glycoprotein expression
X Y Mu1, M P Gosland, M M Bartik
1Department of Pharmacology, University of Kentucky, Lexington, USA.
Insights
P-glycoprotein (Pgp) expression on human lymphocytes is low, even after activation. This suggests Pgp does not significantly impact interleukin-2 (IL-2) secretion in activated T cells.
Area of Science:
- Immunology
- Cell Biology
- Pharmacology
Background:
- P-glycoprotein (Pgp) is involved in drug efflux.
- Its role in normal human lymphocyte function, particularly cytokine secretion, is unclear.
Purpose of the Study:
- To investigate the expression and function of Pgp in human lymphocytes.
- To determine Pgp's involvement in interleukin-2 (IL-2) secretion by activated T cells.
Main Methods:
- Two-color flow cytometry used to detect Pgp expression and IL-2 accumulation.
- Monoclonal antibodies (mAbs) 4E3 and MRK16 used to detect Pgp.
- Doxorubicin (DOX) uptake assays performed on tumor cells and lymphocytes.
Main Results:
- Pgp expression on resting lymphocytes was <1%, increasing to 3% upon mitogenic stimulation.
- Coexpression of Pgp and IL-2 was observed in <10% of IL-2 producing lymphocytes.
- mAb 4E3 showed high sensitivity and specificity for Pgp detection, correlating with DOX efflux.
Conclusions:
- Human T cells do not significantly up-regulate functional Pgp expression after activation.
- Pgp does not appear to play a major role in IL-2 secretion by activated T cells.
Abstract:
The expression of P-glycoprotein (Pgp) on normal human lymphocytes, and its drug exclusion capacity, implies that Pgp might be involved in cytokine secretion. We used two-color flow cytometry to detect simultaneously Pgp expression and IL-2 accumulation in resting and mitogen-activated human lymphocytes. Among resting lymphocytes from five healthy donors less than 1% were Pgp+ as determined by reactivity with the anti-Pgp monoclonal antibody (mAb) 4E3. The percentage of Pgp+ lymphocytes increased to 3% after 24 hr of mitogenic stimulation that induced maximal production of cytoplasmic IL-2. The percentage of lymphocytes that coexpressed membrane Pgp and cytoplasmic IL-2 accounted for < 10% of the total IL-2 producing lymphocytes. Finally, mitogen-induced cytoplasmic IL-2 accumulation was enhanced by stimulation in the presence of monensin but not the Pgp functional inhibitor verapamil. Because mAb 4E3 detected lower than expected numbers of Pgp+ lymphocytes, we compared the binding of mAbs MRK16 and 4E3 concomitant with doxorubicin (DOX)-uptake by K562 and R7 tumor cells and purified CD8+ lymphocytes. The MRK16 mAb was found to be sensitive but not very specific (30%). In contrast, the sensitivity of 4E3 was equivalent to MRK16 (98%) and was highly specific (98.5%). There was also a positive association between DOX efflux and the level of Pgp expression as detected by 4E3 but not MRK16. Thus, human T cells do not markedly up-regulate their expression of functional Pgp molecules as detected by mAb 4E3 following activation, suggesting that Pgp does not play a major role in IL-2 secretion by activated T cells.
