Kannegieter Nynke M, Hesselink Dennis A, Dieterich Marjolein, de Graav Gretchen N, Kraaijeveld Rens, Rowshani Ajda T, Leenen Pieter J M, Baan Carla C
Departments of *Internal Medicine and †Immunology, Erasmus MC, University Medical Center Rotterdam, Rotterdam, the Netherlands.
Ther Drug Monit. 2017 Oct;39(5):463-471. doi: 10.1097/FTD.0000000000000426.
Monocytes significantly contribute to ischemia-reperfusion injury and allograft rejection after kidney transplantation. However, the knowledge about the effects of immunosuppressive drugs on monocyte activation is limited. Conventional pharmacokinetic methods for immunosuppressive drug monitoring are not cell type-specific. In this study, phosphorylation of 3 signaling proteins was measured to determine the pharmacodynamic effects of immunosuppression on monocyte activation in kidney transplant patients.
Blood samples from 20 kidney transplant recipients were monitored before and during the first year after transplantation. All patients received induction therapy with basiliximab, followed by tacrolimus (TAC), mycophenolate mofetil, and prednisolone maintenance therapy. TAC whole-blood predose concentrations were determined using an antibody-conjugated magnetic immunoassay. Samples were stimulated with phorbol 12-myristate 13-acetate (PMA)/ionomycin, and phosphorylation of p38MAPK, ERK, and Akt in CD14 monocytes was quantified by phospho-specific flow cytometry.
Phosphorylation of p38MAPK and Akt in monocytes of immunosuppressed recipients was lower after 360 days compared with before transplantation in the unstimulated samples [mean reduction in median fluorescence intensity 36%; range -28% to 77% for p-p38MAPK and 20%; range -22% to 53% for p-Akt; P < 0.05]. P-ERK was only decreased at day 4 after transplantation (mean inhibition 23%; range -52% to 73%; P < 0.05). At day 4, when the highest whole-blood predose TAC concentrations were measured, p-p38MAPK and p-Akt, but not p-ERK, correlated inversely with TAC (rs = -0.65; P = 0.01 and rs = -0.58; P = 0.03, respectively).
Immunosuppressive drug combination therapy partially inhibits monocyte activation pathways after kidney transplantation. This inhibition can be determined by phospho-specific flow cytometry, which enables the assessment of the pharmacodynamic effects of immunosuppressive drugs in a cell type-specific manner.
单核细胞在肾移植后的缺血再灌注损伤和同种异体移植排斥反应中起重要作用。然而，关于免疫抑制药物对单核细胞激活作用的了解有限。传统的免疫抑制药物监测药代动力学方法并非细胞类型特异性的。在本研究中，通过测量3种信号蛋白的磷酸化来确定免疫抑制对肾移植患者单核细胞激活的药效学作用。
对20例肾移植受者移植后第1年及移植前的血样进行监测。所有患者均接受巴利昔单抗诱导治疗，随后接受他克莫司（TAC）、霉酚酸酯和泼尼松龙维持治疗。采用抗体偶联磁免疫分析法测定TAC全血给药前浓度。样本用佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）/离子霉素刺激，通过磷酸化特异性流式细胞术对CD14单核细胞中p38MAPK、ERK和Akt的磷酸化进行定量分析。
与移植前相比，免疫抑制受者单核细胞中p38MAPK和Akt的磷酸化在360天后未刺激样本中降低[中位荧光强度平均降低36%；p-p38MAPK范围为-28%至77%，p-Akt为20%；范围为-22%至53%；P<0.05]。p-ERK仅在移植后第4天降低（平均抑制23%；范围为-52%至73%；P<0.05）。在第4天，当测量到最高全血给药前TAC浓度时，p-p38MAPK和p-Akt而非p-ERK与TAC呈负相关（rs=-0.65；P=0.01和rs=-0.58；P=0.03）。
免疫抑制联合药物治疗在肾移植后部分抑制单核细胞激活途径。这种抑制作用可以通过磷酸化特异性流式细胞术来确定，该方法能够以细胞类型特异性方式评估免疫抑制药物的药效学作用。
