P-glycoprotein, FK-binding Protein-12, and the Intracellular Tacrolimus Concentration in T-lymphocytes and Monocytes of Kidney Transplant Recipients

Suwasin Udomkarnjananun*, Marith I. Francke, Marjolein Dieterich, Daan Van De Velde, Nicolle H.R. Litjens, Karin Boer, Brenda C.M. De Winter, Carla C. Baan, Dennis A. Hesselink

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

6 Citations (Scopus)
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Abstract

Background. Transplant recipients may develop rejection despite having adequate tacrolimus whole blood predose concentrations (C0). The intra-immune cellular concentration is potentially a better target than C0. However, little is known regarding intracellular tacrolimus concentration in T-lymphocytes and monocytes. We investigated the tacrolimus concentrations in both cell types and their relation with the expression and activity of FK-binding protein (FKBP)-12 and P-glycoprotein (P-gp). Methods. T-lymphocytes and monocytes were isolated from kidney transplant recipients followed by intracellular tacrolimus concentration measurement. FKBP-12 and P-gp were quantified with Western blot, flow cytometry, and the Rhodamine-123 assay. Interleukin-2 and interferon-γ in T-lymphocytes were measured to quantify the effect of tacrolimus. Results. Tacrolimus concentration in T-lymphocytes was lower than in monocytes (15.3 [8.5-33.4] versus 131.0 [73.5-225.1] pg/million cells; P < 0.001). The activity of P-gp (measured by Rhodamine-123 assay) was higher in T-lymphocytes than in monocytes. Flow cytometry demonstrated a higher expression of P-gp (normalized mean fluorescence intensity 1.5 [1.2-1.7] versus 1.2 [1.1-1.4]; P = 0.012) and a lower expression of FKBP-12 (normalized mean fluorescence intensity 1.3 [1.2-1.7] versus 1.5 [1.4-2.0]; P = 0.011) in T-lymphocytes than monocytes. Western blot confirmed these observations. The addition of verapamil, a P-gp inhibitor, resulted in a 2-fold higher intra-T-cell tacrolimus concentration. This was accompanied by a significantly fewer cytokine-producing cells. Conclusions. T-lymphocytes have a higher activity of P-gp and lower concentration of the FKBP-12 compared with monocytes. This explains the relatively lower tacrolimus concentration in T-lymphocytes. The addition of verapamil prevents loss of intracellular tacrolimus during the cell isolation process and is required to ensure adequate intracellular concentration measurement.

Original languageEnglish
Pages (from-to)382-391
Number of pages10
JournalTransplantation
Volume107
Issue number2
DOIs
Publication statusPublished - 1 Feb 2023

Bibliographical note

Funding Information:
S.U. received a grant from the King Chulalongkorn Memorial Hospital and the Thai Red Cross Society for conducting research at the Erasmus MC.

Publisher Copyright:
© Copyright 2022 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited.

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