Transplanting kidneys from CMV-seropositive donors to CMV-seronegative recipients is not associated with poorer renal allograft function or survival

Kevin McLaughlin1, Sabrina Sandhu1, Caren Wu1, Norman Muirhead2, David Hollomby2 and Anthony Jevnikar2

1 Department of Medicine, University of Calgary, Alberta and 2 Division of Nephrology, Department of Internal Medicine, London Health Science Centre and University of Western Ontario, London, Ontario, Canada

Correspondence and offprint requests to: Kevin McLaughlin, Division of Nephrology, Foothills Hospital, 1403 29th Street NW, Calgary, Alberta, T2N 4T3, Canada. Email: kevin.mclaughlin{at}calgaryhealthregion.ca



   Abstract
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Background. Cytomegalovirus (CMV)-seronegative recipients of renal allografts from CMV-seropositive donors (D+/R–) have a higher rate of acute rejection than other renal transplant recipients. A relationship between CMV infection/disease and chronic allograft nephropathy (CAN) has been proposed from animal studies, although human studies have been inconclusive. The objective of this study was to determine if CMV seromatching has an effect on renal allograft function and allograft survival.

Methods. A retrospective single centre study was carried out in 333 first cadaveric transplant recipients from January 1, 1991 to December 31, 1997. The primary end-point was creatinine clearance at 3 years post-transplant in groups based on CMV seromatching. The secondary end-point was renal allograft survival.

Results. Mean creatinine clearance 3 years post-transplant was 53.4 ml/min/1.73 m2 of body surface area. There was no significant difference in the mean creatinine clearance for groups formed on the basis of CMV seromatching. Delayed graft function and acute rejection were associated with a lower creatinine clearance at 3 years and reduced overall graft survival [hazard ratios 2.35 (1.56–3.54) (P<0.001) and 1.57 (1.0–2.46) (P = 0.046), respectively]. Considering the end-point of graft loss due to acute rejection (censoring for death with a functioning graft) identified the D+/R– group as having an increased hazard of graft loss due to acute rejection [hazard ratio 3.12 (1.16–8.57) (P = 0.024)].

Conclusions. The D+/R– group does not appear to have poorer renal allograft function 3 years post-transplant. This group does, however, have an increased risk of early allograft loss due to acute rejection.

Keywords: chronic allograft nephropathy; creatinine clearance; cytomegalovirus; renal transplantation



   Introduction
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Cytomegalovirus (CMV) disease incurs significant morbidity and health care costs in the setting of organ transplantation [1]. The risk of CMV infection and CMV disease following transplantation is highest in CMV-seronegative recipients of renal allografts from CMV-seropositive donors (D+/R–), i.e. recipients who have no immunity to the virus and receive a kidney from a donor who has been infected previously [2]. In addition to the direct effects of viral infection, there is also evidence of an association between CMV infection/disease and acute allograft rejection in the setting of renal and other solid organ transplantation [3–5]. The nature of this relationship is a matter of debate, with evidence supporting both a ‘forward’ relationship (CMV infection/disease precedes acute rejection) and a ‘backward’ relationship (CMV infection/disease follows acute rejection) [6]. We recently demonstrated that seromismatched recipients of renal allografts had a significantly higher rate of acute rejection than non-seromismatched recipients, supporting a forward relationship [7]. A relationship between CMV infection/disease and chronic allograft nephropathy (CAN) has been proposed, and several animal studies have demonstrated mechanisms by which CMV infection/disease may enhance the process of chronic allograft nephropathy [8–10]. It is unclear, however, if this effect is independent of the effects of acute rejection on the development of CAN.

The objective of this study was to determine if CMV matching has an effect on renal allograft function and allograft survival that is independent of its effect of acute rejection.



   Subjects and methods
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
This was a retrospective study of patients transplanted at a single centre. The primary end-point was the square root of creatinine clearance 3 years after renal transplant in groups based on CMV seromatching. The secondary end-point was renal allograft survival. The study population comprised all first cadaveric transplant recipients from January 1, 1991 to December 31, 1997. The standard induction immunosuppression regime comprised cyclosporin and prednisone. Patients who were considered to be at high immunological risk received biological induction with antithymocyte globulin (ATGAM) or OKT3. Some patients with delayed graft function were also switched to biological induction. Azathioprine was added to the immunosuppression regime if the patient had an episode of acute rejection. Patients who were CMV seromismatched received CMV hyperimmune globulin (CMVHIG) for the first 4 months post-transplant. Prophylactic antiviral therapy was not given during this time period. Creatinine clearance was determined for each individual using the Cockcroft–Gault equation [11]. Acute rejection was either biopsy proven according to standard criteria or diagnosed clinically if serum creatinine rose ≥25% from baseline and improved following three consecutive daily doses of intravenous 250 mg methylprednisolone [12]. Renal allograft biopsy was performed in all cases where the serum creatinine did not improve after methylprednisolone treatment. All cases of graft loss due to acute rejection were confirmed histologically. Data collected included age, sex, the degree of donor–recipient human leukocyte antigen (HLA) matching, recipient panel-reactive antibody (PRA) percentage, cold ischaemic time, use of biological induction, CMV viral serology of donor and recipient, delayed graft function (requirement for dialysis beyond 24 h after transplantation), weight and serum creatinine at 3 years post-transplant.

As a dependent variable, creatinine clearance did not have a symmetrical distribution and a variety of transformations were tried to create a symmetrical distribution. The square root transformation yielded a distribution that approximated a normal distribution and was, therefore, used as the dependent variable. A multiple linear regression model was used to study the effects of dichotomous and continuous independent explanatory variables on the continuous dependent variable square root of creatinine clearance. Kaplan–Meier survival analysis using a log rank test was used to compare renal allograft survival for groups based on CMV seromatching. A Cox's proportional hazard model was used to identify variables influencing allograft survival. Variables included in the models were either known to affect the creatinine clearance or the risk of allograft loss from previous studies, or were considered as potentially significant from univariate logistic regression (i.e. P<0.2). The model considered effect modification and/or confounding by two and/or three variable interactions. Manual backward stepwise elimination of the least significant variable, starting with three variable interaction terms, was done based on a combination of clinical and statistical importance. Analyses were done with STATA (Stata Corporation, College Station, TX).



   Results
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Three hundred and thirty-three patients (216 males) underwent first cadaveric renal transplant during this time period. Mean age (±SD) at the time of transplantation was 46.6 years (±12.6). Mean cold ischaemic time was 24.7 h (±7.9). Fifty patients (15%) received a biological induction agent and 111 patients (33%) had delayed graft function. One hundred and ninety-four (58.3%) patients had at least one acute rejection episode. Mean creatinine clearance 3 years post-transplant was 53.4 ml/min/1.73 m2 body surface area (BSA). Graft survival [±95% confidence interval (CI)] 3 years post-transplant was 78% (72.7–82.0). Table 1 shows the variables affecting the creatinine clearance at 3 years by simple linear regression. In this table, the groups formed on the basis of donor and recipient CMV serostatus are: donor negative/recipient positive (D–/R+); donor positive/recipient negative (D+/R–); donor positive/recipient positive (D+/R+); and donor negative/recipient negative (D–/R–). In the models, the four CMV groups were considered as three ‘dummy’ variables with the D–/R– group as the control and one dummy variable representing each of the other three groups. Despite not reaching the required level of significance by univariate analysis, CMV serostatus was entered into the multiple linear regression model as this was the primary variable under study. All interaction terms were non-significant. The final model contained three variables of which two, delayed graft function and acute rejection, had an independent and significant effect on the square root of creatinine clearance at 3 years. Compared with those who had no delayed graft function, individuals who experienced delayed graft function had a reduction in creatinine clearance at 3 years of –5.42 ml/min/1.73 m2 BSA. Compared with those who had no acute rejection, individuals who experienced acute rejection had a reduction in creatinine clearance at 3 years of –5.39 ml/min/1.73 m2 BSA. These data are shown in Table 2. The use of biological induction remained in the final model although the effect failed to reach the level of statistical significance.


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Table 1. Variables affecting creatinine clearance at 3 years post-transplant by simple linear regression

 

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Table 2. Variables affecting creatinine clearance at 3 years post-transplant by multiple linear regression

 
Using Kaplan–Meier's survival analysis and considering the end-point of graft loss from any cause, there was a trend towards poorer graft survival for the D+/R– group, but the difference did not reach statistical significance (P = 0.09). This is shown in Figure 1. In the Cox proportional hazard model, all interaction terms were non-significant. The final model identified two variables, delayed graft function and acute rejection, that were associated with graft loss. The hazard ratios for delayed graft function and acute rejection were 2.35 (1.56–3.54) (P<0.001) and 1.57 (1.0–2.46) (P = 0.046), respectively. Considering the end-point of graft loss due to acute rejection (censoring for death with a functioning graft) revealed poorer graft survival in the D+/R– group (P = 0.017). This is shown in Figure 2. In the Cox proportional hazard model, all interaction terms were non-significant and the final model identified D+/R– as being associated with graft loss with a hazard ratio of 3.12 (1.16–8.57) (P = 0.024).



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Fig. 1. Renal allograft survival for CMV seromismatched and non-seromismatched groups considering all causes of graft loss.

 


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Fig. 2. Renal allograft survival for CMV seromismatched and non-seromismatched groups considering only graft loss due to acute rejection.

 


   Discussion
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
The primary objective of this study was to determine if CMV seromatching has an effect on renal allograft function that is independent of its effect of acute rejection. In the absence of histological data (protocol biopsies were not performed during this time period), creatinine clearance was used as a marker of allograft function and as a surrogate for CAN [13]. The results observed here suggest that CMV-seronegative recipients of renal allografts from CMV-seropositive donors do not have poorer renal allograft function 3 years post-transplant. This result should, however, not be considered in isolation as the creatinine clearance measurements reflect only values from patients who had functioning grafts at 3 years. The importance of this is underscored by the fact that the D+/R– group have an increased risk of allograft loss due to acute rejection. Overall, the results of our previous and present studies on the effects of CMV seromismatching would suggest that the D+/R– group has an increased risk of acute renal allograft rejection and early graft loss due to acute rejection. If, however, the allograft is not lost due to acute rejection, the allograft function appears to be similar to that of other groups as judged by either creatinine clearance or graft survival (note that the survival curves in Figure 2 are parallel after the initial post-transplant period).

Despite the evidence from animal studies that CMV infection may enhance and/or accelerate the histological changes associated with CAN, human studies using either histological or functional markers of CAN have failed to confirm this association [14–16]. The explanation for the discrepant results between animal studies and clinical observations is unclear. The first possibility to consider is that the present (and previous) studies were underpowered. Assuming an {alpha} of 0.05, the present study had sufficient power (>0.8) to detect a 13% (or 7.2 ml/min/1.73 m2 BSA) difference in creatinine clearance between the D+/R– group and the other groups assuming that the mean creatinine clearance in the other groups was 54.4 (±21.9). Assuming an {alpha} of 0.05 and power 0.8, a study based upon the observed difference in creatinine clearance of 6.5% (or 3.6 ml/min/1.73 m2 BSA) would require a sample size of almost 1400 patients. A second possible explanation for the negative findings is that functional markers alone are too insensitive to detect differences in CAN. Despite the prognostic importance of abnormal functional parameters, it has been shown previously that the histological markers of both acute allograft rejection and CAN may precede changes in functional parameters [13,17–19]. This explanation cannot be excluded in the absence of histological data, although it is noteworthy that in the study by Helantera et al., no difference was observed in the 6-month protocol biopsies of patients with and without CMV infection [15]. A third possible explanation is that the effects of CMV seromismatching were negated due to performance bias. As part of our treatment protocol, recipients who were CMV seromismatched received prophylactic CMVHIG. As has been demonstrated in several reports, immunoglobulin preparations have multiple immunomodulatory effects [20]. Mechanisms by which immunoglobulin may attenuate diseases mediated by antigen-specific immunoglobulin G (IgG) include Fc{gamma} receptor blockade, anti-idiotype antibody activity and downregulation of specific autoreactive B cells [20]. As such, the mechanisms by which CMV disease may accelerate CAN, such as via the production of anti-endothelial cell antibodies, may have been attenuated by CMVHIG. The study reported by Akposso also included pre-emptive treatment (using gancyclovir), which may partly explain their negative results [16]. Another potential source of performance bias was the individualization of immunosuppression. The use of biological induction in patients with delayed graft function or considered to be at high immunological risk (which may explain the trend towards lower creatinine clearance in patients given biological induction) and the addition of a third immunosuppressive agent following acute rejection may have attenuated the effect of some explanatory variables on allograft function. This may also have attenuated the interaction between variables, such as the synergistic effect of CMV infection and cold ischaemic time observed in animal models [21].

The principal findings of this study were that the D+/R– group have an increased risk of early allograft loss due to acute rejection, but did not have poorer allograft function 3 years post-transplant. While the magnitude of the impact of CMV seromatching on the development of CAN may have been underestimated by the absence of histological data and the performance bias that is inherent in studies that are observations of clinical practice, these results would suggest that transplant allocation on the basis of CMV matching cannot be supported on the basis of clinical data.



   References
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 

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Received for publication: 20. 2.04
Accepted in revised form: 22.10.04





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