Kidney transplant recipients are vulnerable to ischemia-reperfusion injury during donor organ harvest and recipient allograft reperfusion. Ischemia-reperfusion injury of kidney transplantation is associated with delayed graft function and acute rejection, which are crucial factors affecting graft outcomes.1,2 Although graft survival and long-term prognosis are poor when delayed graft function and acute rejection occur,1,2,3 this does not necessarily lead to graft failure, depending on the severity of ischemia-reperfusion injury, delayed graft function, and the degree of recovery.4 Thus, strategies to reduce the risk of ischemia-reperfusion injury seem necessary for kidney transplantation.

Recently, there has been increasing recognition of the critical role of innate immunity in organ transplantation.5,6 Nucleotide‐binding domain‐like receptor protein 3 (NLRP3) inflammasome is closely associated with both ischemia-reperfusion injury and the innate immune system,7,8 and affects the outcomes of organ transplantation.6,9 One of the pathways involved in this pathogenesis is the contribution of the NLRP3 inflammasome to the excretion of inflammatory cytokines, particularly interleukin (IL)-1β and IL-18, resulting in an inflammatory response.10

Dexmedetomidine, a highly selective α2-adrenoreceptor agonist, has been shown to reduce renal ischemia-reperfusion injury through its sympatholytic, anti-inflammatory, hemodynamic stabilizing, and diuretic effects.11,12,13,14,15 Furthermore, a growing body of evidence supports its potential impact on the innate immunity via the inhibition of NLRP3 inflammasome formation.16,17 Nevertheless, the renoprotective effects of dexmedetomidine in kidney transplantation have not been explored.

Therefore, we conducted this prospective randomized controlled trial to investigate the effect of intraoperative dexmedetomidine infusion on the renal function and the development of delayed graft function in patients undergoing elective living donor kidney transplantation.

Methods

Patients

This study was conducted at Severance Hospital, Yonsei University Health System, Seoul, South Korea, approved by our institutional review board (4-2017-0767), and registered 31 October 2017 at http://www.ClinicalTrials.gov (NCT03327389). Written consent was obtained from all patients.

Between December 2017 and August 2020, 104 patients between 20 and 80 yr of age scheduled for elective living donor kidney transplantation were randomly assigned to either the dexmedetomidine or the control group in 1:1 ratio using a computer-generated number table by an investigator (J.H.P.) who was not involved in patient management. Group assignments were kept in sequentially numbered opaque, sealed envelopes that were opened by postanesthesia care unit nurses who were not involved in the study. The group assignment was revealed on postoperative day (POD) 7. From both groups, 30 patients were randomly selected at a 1:1 ratio for the measurement of plasma IL-1β and IL-18 levels. Patients were not included if they met at least one of the following criteria: severe sinus bradycardia (< 50 min-1), second- or third-degree atrioventricular block, left ventricular ejection fraction < 30%, exposure to dexmedetomidine in the past 30 days, or a history of severe allergy to dexmedetomidine or clonidine.

Intervention

In the dexmedetomidine group, dexmedetomidine 200 μg was added to 0.9% saline in 50 mL and was administered at a rate of 0.4 μg·kg-1·hr-1, starting immediately after anesthesia induction and until the end of surgery. In the control group, the equivalent volume of 0.9% saline was administered at the same rate and duration. The study drugs were prepared in identical syringes by an investigator who was not involved in patient management. The patients, surgeons and attending anesthesiologist involved in patient management, and outcome assessors were blinded to the group assignment. Data analysts were aware of the group allocation.

Primary outcome and secondary outcomes

The primary outcome was the serum creatinine levels on POD 7 between the dexmedetomidine and control groups. The secondary outcomes were changes in the serum creatinine level and estimated glomerular filtration rate (eGFR) up to six months; the incidence of delayed graft function; and changes in serum cystatin C, plasma IL-1β, and IL-18 levels during the perioperative period. Delayed graft function was defined as the need for dialysis within the first week after kidney transplantation.2

Perioperative management

Upon arrival at the operating room, standard monitoring devices were applied. General anesthesia was induced with propofol 1.5-2 mg·kg-1 and remifentanil 0.3-0.5 μg·kg-1·min-1, and rocuronium 0.6 mg kg-1 was used for neuromuscular blockade. After endotracheal intubation, anesthesia was maintained with sevoflurane 1.5-2.5% in 50% oxygen in air, and a 0.1-0.3 μg·kg-1·min-1 remifentanil infusion. The depth of anesthesia was monitored by the patient state index (PSI) using a Sedline® electroencephalograph sensor (Masimo Corp., Irvine, CA, USA) and maintained at a PSI of 25-50. Subsequently, a radial artery catheter and an internal jugular central catheter were inserted. If the patient had a pre-existing hemodialysis catheter placed in an internal jugular or a subclavian vein, this was used. The radial arterial catheter was connected to FloTrac™ (Edwards Lifesciences LLC, Irvine, CA, USA) to monitor the cardiac index, stroke volume variation (SVV), and systemic vascular resistance index using the Vigileo™ monitor (Edwards Lifesciences LLC; software version 3.0). To maintain adequate perfusion, an acetate-buffered balanced crystalloid solution and total 750 mL of 5% albumin were administered throughout the surgery, with a final target mean arterial pressure > 90 mm Hg, central venous pressure (CVP) 10-15 mm Hg, and SVV < 6%. If hypotension occurred (defined as a reduction in mean arterial pressure below 20% of the preanesthetic value), acetate-buffered balanced crystalloid solution was infused until the target CVP and SVV were achieved. If response was not adequate, norepinephrine at 0.01 μg·kg-1·min-1 was started and titrated. After renal vascular anastomosis, furosemide 20 mg was administered to enhance diuresis. If requested by the surgeon, repeated doses of furosemide were administered. A hemoglobin level less than 8 g·dL-1 was used as the transfusion threshold.

Postoperative management followed the institutional standard protocol. Briefly, urine output was replaced (millilitre for millilitre) with an intravenous infusion of an acetate-buffered balanced crystalloid solution until POD 2, after which the fluid balance was targeted to be 500-1,000 mL negative using 0.45% saline. If the urine output was less than 100 mL·hr-1, intravenous furosemide was repeatedly administered with escalating doses from 20 mg at the surgeon’s discretion until the urine output exceed 100 mL·hr-1. For postoperative analgesia, an intravenous patient-controlled analgesia device (Accufuser plus®, P2015M, Woo Young Medical Co., Ltd., Chungbuk, South Korea) containing fentanyl 15 μg·kg-1 and ramosetron 0.3 mg in 0.9% saline with a total volume of 100 mL was connected at the time of skin closure. If patients requested additional analgesics, intravenous tramadol 50 mg was administered.

All donor nephrectomies and kidney transplantations were performed by the same nephrectomy team and transplantation team, respectively.

Assessment

The serum creatinine level was recorded one day before surgery, immediately after surgery, daily after surgery for seven days, and three and six months after surgery. Serum cystatin C levels were measured after anesthesia induction, immediately after surgery, and on POD 1 and 2. Estimated glomerular filtration rate_MDRD and eGFR_EPI were calculated using the modification of diet in renal disease (MDRD) and Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) formula, respectively. Plasma IL-1β and IL-18 were measured in 30 randomly selected patients after anesthesia induction, after renal vascular anastomosis, and 1 hour after surgery. Plasma levels of IL-1β and IL-18 were measured with a quantitative test using the human IL-1β and IL-18 ELISA kit (Abcam, Cambridge, UK) according to the manufacturer’s instructions.

Random urine creatinine, protein, and urine protein to creatinine (p/c) ratio were measured one day before surgery, immediately after surgery, and daily thereafter for seven days. Routine complete blood cell counts and serum chemistry profiles were collected at the abovementioned time points. C-reactive protein levels were measured one day before surgery and on POD 7.

Preoperative variables included demographic data, history of hypertension, diabetes, IgA nephropathy, duration of dialysis, preoperative left ventricular ejection fraction, medications, number of human leukocyte antigen mismatch, and ABO incompatibility. Intraoperative variables included anesthesia time, cold and warm ischemia time, transplanted kidney weight, use of norepinephrine, fluid balance, and transfusion requirement. Cold ischemia time was calculated from nephrectomy to when the kidney was taken from cold storage for transplantation. Warm ischemia time was calculated from when the kidney was taken from cold storage for transplantation to reperfusion with warm blood. Postoperative variables included heart rate, blood pressure, diuretic dose, fluid balance, incidence of biopsy-proven acute rejection, duration of hospital stay, reoperation, and graft loss and all-cause mortality within six months of transplantation. For heart rate and blood pressure, the lowest values of the day were collected.

Sample size calculation

The sample size calculation was based on the primary outcome. This study was powered to detect a 0.3 mg·dL-1 decrease in the serum creatinine level in the dexmedetomidine group compared with the control group. There is no generally accepted definition for renal injury in patients undergoing kidney transplantation because of difficulties identifying an established baseline of renal function,18 so we assumed that a difference of 0.3 mg·dL-1 in the serum creatinine level between groups would be meaningful according to the clinical significance19 and Kidney Disease: Improving Global Outcomes criteria.20 Therefore, the sample size was calculated based on the previous results in patients undergoing living donor kidney transplantation in our institute, in which the mean (standard deviation [SD]) serum creatinine level was 1.13 (0.51) mg·dL-1 on POD 7. Forty-six patients were required in each group to obtain a power of 80% with an alpha of 0.05. Considering a 10% dropout rate, we decided to enrol 52 patients in each group.

Statistical analysis

Continuous variables are shown as mean (SD) or median [interquartile range]. Dichotomous variables are expressed as number of patients (percentage). Continuous variables were compared using the Mann–Whitney U test or independent t test, based on the normality of the residuals tested by the Shapiro–Wilk test. Dichotomous variables were compared using the Fisher’s exact tests. The confidence interval was calculated based on Student t distribution. A linear mixed model with patient indicator as the random effect, and group, time, and group-by-time as the fixed effects was used to analyze repeatedly measured variables such as cystatin C, serum creatinine, inflammatory mediators, and percentage of neutrophil count. Two independent t tests were used for power analysis. SPSS 24 (SPSSFW, SPSS, IBM, Armonk, NY, USA), SAS (version 9.4; SAS Institute Inc., Cary, NC, USA), and PASS (version 12; NCSS, LLC, Kaysville, UT, USA) were used for statistical analyses. P values less than 0.05 were considered statistically significant for the primary outcome. P values less than 0.01 were considered statistically significant for the secondary outcomes.

Results

Of the 104 patients, one patient in the dexmedetomidine group was excluded because they underwent an additional surgery after kidney transplantation (Fig. 1). Patient demographics and donor details are summarized in Table 1. None of the donors had diabetes.

Fig. 1
figure 1

Patient enrolment in the study

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Table 1 Patient baseline characteristics
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Changes in the renal function and biomarkers of inflammation are shown in Fig. 2 and Table 2. Postoperatively, serum creatinine levels continued to decrease throughout the study period. The mean (SD) serum creatinine level on POD 7 was comparable between the groups (dexmedetomidine vs control: 1.11 [0.87] mg·dL-1 vs 1.06 [0.73] mg·dL-1; mean difference, 0.05 mg·dL-1; 95% confidence interval (CI), -0.27 to 0.36; P = 0.77, Fig. 2A). Delayed graft function occurred in one patient in each group (odds ratio, 1.020; P > 0.99). Serum cystatin C levels during the first two postoperative days and serum creatinine levels, eGFR, and urine output during the first seven postoperative days were also comparable without significant group × time interactions (Table 2, Fig. 2B-D). Serum creatinine levels and eGFR at three and six months after surgery remained similar to those on POD 7 and were not different between the groups (Fig. 2B-D). Plasma IL-1β and IL-18 levels were comparable between the groups throughout the study period. Other biomarkers reflecting degree of inflammation were not significantly different between the groups without significant group × time interactions (Table 2).

Fig. 2
figure 2

Perioperative changes in: A = serum creatinine levels; B = urine output; C = eGFR_MDRD; D = eGFR_EPI. Data are presented as mean (standard deviation) and analyzed using independent t tests, based on the normality of the residuals tested by the Shapiro–Wilk test. Pgroup x time = P value of the group and time interaction obtained by the linear mixed model. *Compared with the baseline value (all, P < 0.05). eGFR_MDRD = estimated glomerular filtration rate calculated using the Modification of Diet in Renal Disease (MDRD); eGFR_EPI = estimated glomerular filtration rate calculated using the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) formula; POD = postoperative days

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Table 2 Changes in renal function and biomarkers of inflammation
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Intraoperative data including duration of warm and cold ischemia, transplanted kidney weight, amount of packed red blood cells transfused, and fluid balance were similar between the groups. Intraoperative vasopressor requirement, hemodynamic data and diuretic usage were not significantly different between the groups. Emergence time from anesthesia was not significantly different between the groups (median, 13 vs 10 min; P = 0.05) (Table 3).

Table 3 Intraoperative data
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Postoperative data were comparable (Table 4). Biopsy-proven acute rejection occurred in 13 and 14 patients in the dexmedetomidine and the control groups, respectively (odds ratio, 0.929; 95% CI, 0.386 to 2.236; P = 0.87). Patients with biopsy-proven acute rejection were treated with plasmapheresis and/or intravenous immunoglobulin and/or rituximab and/or steroid pulse therapy at a similar treatment rate. Three patients in the dexmedetomidine group and one patient in the control group required dialysis after POD 7 (odds ratio, 3.188; 95% CI, 0.320 to 31.705; P = 0.36). All patients received a similar immunosuppressive regimen, consisting of tacrolimus with prednisone and mycophenolate mofetil.

Table 4 Postoperative data
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Postoperative fluid balance and hemodynamic data were also comparable between groups (Electronic Supplementary Material, eTable).

Discussion

In this prospective randomized controlled trial, intraoperative dexmedetomidine infusion produced neutral effects on renal function and the development of delayed graft function in patients undergoing elective living donor kidney transplantation. Dexmedetomidine did not mitigate an increase in inflammatory cytokines, including plasma IL-1β and IL-18, or affect the incidence of acute rejection and postoperative outcomes.

Dexmedetomidine, a commonly used adjuvant anesthetic, has shown renoprotective effects in various animal models of renal ischemia-reperfusion injury.14,16 Dexmedetomidine provides hemodynamic stability, increases glomerular filtration by inhibiting catecholamine and renin secretion,21,22 and attenuates systemic inflammatory responses and inflammatory mediators.23 Additionally, in a recent study, a protective effect of dexmedetomidine against renal ischemia-reperfusion injury was suggested to be mediated through an enhanced autophagy, which decreased activation of the NLRP3 inflammasome.16

Contrary to experimental data, however, the renoprotective effects of dexmedetomidine for surgical patients are controversial.12,13,24,25,26 Dexmedetomidine reduced the incidence of postoperative acute kidney injury in cardiac sugery patients,12,13 while it failed to reduce postoperative acute kidney injury or improve early renal function in other surgical patient groups.25,26 Differences in the degree of renal ischemia-reperfusion injury, including the occurrence of cold ischemia caused by a direct renal pedicle clamping, could be considered possible contributing factors to the inconsistent effects of dexmedetomidine. Notably, most prospective clinical trials addressing the effects of dexmedetomidine on renal function excluded patients with reduced renal function.12,13,24 Only one randomized controlled study including patients with a serum creatinine level between 1.5 and 2 mg·dL-1 reported a neutral effect of dexmedetomidine.27 In this context, kidney transplantation surgery performed in patients with renal impairment undergoing cold kidney ischemia might be a unique opportunity to examine the renoprotective effect of dexmedetomidine. We therefore conducted this study to investigate the effects of dexmedetomidine on renal function in living donor kidney recipients with a concomitant examination of possible mechanisms.

In this prospective study, we did not observe any beneficial effects of dexmedetomidine in terms of renal function or the incidence of delayed graft function. After surgery, the biomarkers reflecting renal function such as serum creatinine, cystatin C, urine output, and urine p/c ratio were improved in both groups without intergroup differences. With respect to the inflammasome-related markers, which we hypothesized to be an important mechanism of action of dexmedetomidine, white blood cell counts increased immediately after surgery to POD 2, and then decreased thereafter. Postoperative plasma IL-1β and IL-18 levels were not significantly higher than baseline values, except IL-18 levels after vascular anastomosis in the dexmedetomidine group. There were no significant differences between the groups throughout the study period.

In association with the neutral effects of dexmedetomidine in this study, the adequate dose of dexmedetomidine for providing renal protection in kidney transplantation might be the first issue to be reviewed, although there has been no clear guidance on the proper dose of dexmedetomidine that provides renoprotective effects without hemodynamic compromise in this subset of surgery. Organ protective effects of dexmedetomidine have been reported to be dose-dependent,24 and prolonged use of dexmedetomidine with postoperative infusion may be efficacious for protecting renal function against acute kidney injury in previous studies.12,13 Considering the prolonged sedative effect of dexmedetomidine in patients with severe renal impairment,28 in the current study, dexmedetomidine was infused without an initial loading dose, and the chosen dose of dexmedetomidine was the same or less than those used in previous studies to provide hemodynamic stability and overall safety.12,24,25,29 Dexmedetomidine produced no statistically different prolongation in emergence time from anesthesia (based on a prespecified P value for secondary outcomes < 0.01) or significant hemodynamic derangements. Nonetheless, regarding the patients’ renal function, relatively short operation time, and the need for immediate emergence after surgery and postoperative transfer to the ward, increasing the dose of dexmedetomidine to obtain renoprotective effects requires careful evaluation.

Regarding the timing of administration, treating dexmedetomidine before renal ischemia-reperfusion injury rather than after ischemia-reperfusion injury was reported to be more effective.15,23 In the current study, dexmedetomidine was also administered immediately after anesthesia induction as a pretreatment to prevent renal ischemia-reperfusion injury in kidney recipients. In terms of the grafted kidney, however, ischemic injury begins at the time of organ harvest.30 Therefore, pretreatment with dexmedetomidine in the recipient alone is probably insufficient to mitigate renal ischemia-reperfusion injury, which is an inherent limitation in the clinical study of organ transplantation. Whether proper donor interventions before organ harvest or using preservation solution reduces ischemic injury needs to be further evaluated.

In kidney recipients with an increased risk of renal injury,31 intraoperative dexmedetomidine alone might not be sufficient to prevent renal ischemia-reperfusion injury. Approximately 90% of our patients had hypertension, 80% were under dialysis before transplantation, 60% received desensitization therapy, and 40% were ABO incompatible. In the context of cardiac surgery, dexmedetomidine was particularly effective at reducing renal ischemia-reperfusion injury in patients with preoperative normal renal function or mild chronic kidney disease, whereas it failed to show beneficial effects in patients with reduced preoperative renal function.32 Similar contradictory clinical results between patients undergoing cardiac surgery and kidney transplantation have been reported with erythropoietin treatment.33,34

In this study, dexmedetomidine did not affect the levels of plasma IL-1β and IL-18, which are markers of NLRP3 inflammasome activation and innate immunity.35 Activation of the NLRP3 inflammasome has a pathogenic role in renal ischemia-reperfusion injury, suggesting that inhibition of NLRP3 reduces renal tubular injury, fibrosis, and inflammation.36 Dexmedetomidine has been shown to inhibit the activation of NLRP3 inflammasome.16 In the present study, however, IL-1β and IL-18 levels were comparable throughout kidney transplantation in both groups. As reported in dialyzed chronic kidney patients,35,37 pre-existing increased NLRP3 inflammasome activity and elevated levels of IL-1β and IL-18 may have attenuated the effects of dexmedetomidine on the NLRP3 inflammasome in the present study. In addition, although IL-18 levels increased after vascular anastomosis compared with the baseline value in the dexmedetomidine group, this intragroup difference does not seem to be clinically meaningful, considering the large sampling variability in relation to the small sample size.

Although delayed graft function, defined as requiring dialysis within seven days after kidney transplantation, is one of the most important clinical entities determining the prognosis in the early phase after kidney transplantation,1,2,18 practical barriers have limited studying delayed graft function as a primary end point. Physicians’ subjective experience might be involved in deciding whether to dialyze kidney recipients, even within the same transplant team. In this study, the incidence of delayed graft function was 2% in both groups, which was lower than the reported prevalence (4-10%) in living donor kidney transplantation.1 Serum creatinine has been used as the most practical and sensitive predictor of graft outcome.19,38 Even mild differences in very early serum creatinine levels have a significant impact on long-term graft survival,39 and a 0.3 mg·dL-1 increase in the serum creatinine level was reported to double the risk of long-term graft loss.39

The limitations of this study are as follows. First, this study appears to be underpowered to detect the originally anticipated difference in mean creatinine concentration of 0.3 mg·dL-1. The measured SD for the primary outcome (0.73 and 0.87 mg·dL-1) was considerably wider than the anticipated SD used for sample size calculation (0.51 mg·dL-1). This increased variability likely diminished the statistical power for detecting the projected difference in mean creatinine concentrations. Second, patients received antibiotics and anti-fungal agents in the perioperative period according to an institutional standard protocol. Although there were no intergroup differences regarding their usage, we cannot determine the influence of these medications on the results because the effects of these drugs on dexmedetomidine are unknown. Third, plasma concentration of dexmedetomidine may not have reached high enough concentrations to convey the hypothesized effects because we used intraoperative dexmedetomidine infusion without an initial loading dose to avoid delayed recovery, and discontinued dexmedetomidine at the end of surgery. In previous studies reporting the beneficial effect of dexmedetomidine on renal function,12,24,40 a loading dose of dexmedetomidine was administered or continuously infused after surgery. In addition, considering the elimination half-life of dexmedetomidine (two to three hours), it would not have reached steady-state (five halve times) within the operation time (mean, 6 hr).41 Finally, the fact that this is a single-centre study and that the hard end point could not be established is another limitation of this study.

In conclusion, intraoperative dexmedetomidine infusion did not produce any beneficial effects on renal function or delayed graft function in patients undergoing elective living donor kidney transplantation.