Department of Renal Medicine, The University of Sydney at Westmead Hospital, Westmead, NSW, Australia
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Abstract |
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Methods. We performed a cross-sectional study of haemodialysis (HD) and peritoneal dialysis (PD) patients in order to study potential determinants of carbamylated Hb (CarbHb) and to investigate the relationship between CarbHb and established measures of dialysis dose/adequacy by multivariate analysis.
Results. In 80 HD patients, CarbHb was independently predicted by post-dialysis urea (r=0.40, P<0.01), serum albumin (r=0.24, P<0.05) and serum bicarbonate (r=-0.40, P<0.05). No correlation was found between CarbHb and measures of dialysis dose/adequacy (Kt/V, urea reduction ratio, weekly dialysis duration, and normalized protein catabolic rate (nPCR)). In 42 PD patients, serum urea was the only significant independent predictor of CarbHb (r=-0.51, P=0.001). No relationship was found between CarbHb and Kt/V, corrected creatinine clearance (CrCl) or nPCR in PD patients.
Conclusions. Serum urea is the most consistent independent predictor of CarbHb in dialysis patients. This association in combination with the lack of a relationship with conventional measures of dialysis dose and a positive relationship with serum albumin suggest that a single measurement of CarbHb is unlikely to be a useful indicator of the adequacy of dialysis.
Keywords: adequacy; carbamylated; dialysis; haemoglobin; uraemia; urea
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Introduction |
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Because the process of Hb carbamylation is irreversible for the life of the Hb protein, the extent of Hb carbamylation is an indicator of the extent of exposure to urea, analogous to the glycosylation of Hb in diabetic patients. In view of this, it has been suggested [5] that CarbHb may be a marker of both uraemia and the adequacy of dialysis therapy. Although widespread agreement on the necessity of monitoring dialysis dose exists, there remains significant contention as to the most appropriate measure [6,7]. The widespread use of measurements of urea removal is largely based on studies showing their relationship with morbidity and mortality [8,9]; however, the survival rates obtained with long slow dialysis [6] indicate that other factors besides urea clearance are important for determining patient outcome. Although not previously examined in peritoneal dialysis (PD) patients, earlier studies in haemodialysis (HD) patients have shown a correlation between CarbHb and dialysis dose measured by Kt/V [5] and the urea reduction ratio (URR) [5,10], leading to the suggestion that CarbHb may be an alternative simplified index of uraemic control and the adequacy of dialysis. The true utility of CarbHb as a predictor of mortality and morbidity in dialysis patients remains unknown, however, as there are no published longitudinal studies.
In order to examine potential determinants of CarbHb and its relationship to current measures of dialysis dose or adequacy, we conducted a cross-sectional analysis of a population of HD and PD patients.
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Subjects and methods |
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Dialysis method
HD patients in this study were undergoing thrice-weekly low-flux maintenance dialysis. HD was performed with modified cellulose (Baxter, Illinois, USA) or polysulphone (Fresenius AG, Bad Homburg, Germany) membranes, using a bicarbonate-based buffer of 40 mmol/l. Treatment times varied between 4 and 6 h per session (mean 5.1 h). PD patients were undergoing maintenance PD at home using a dextrose-containing dialysate (Baxter, Illinois, USA).
Dialysis adequacy
HD patients Blood was drawn at the start of the mid-week dialysis session for routine estimation of pre-dialysis serum urea, albumin, phosphate, and bicarbonate. Post-dialysis urea was measured at the same session using the slow-flow method. This involved sampling blood from the arterial line 2 min after reducing the blood pump speed to 50 ml/min without an ultrafiltration gradient. The proportional reduction of urea (URR) was calculated from these two measurements.
Values for a single-pool variable volume Kt/V were calculated using the logarithmic estimate of Daugirdas [12]:
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Normalized protein catabolic rate (nPCR) was estimated using the nomographic method of Daugirdas and Depner [14].
PD patients Blood was drawn at the time of dialysis adequacy for routine measurement of serum urea, albumin, phosphate, and bicarbonate. Estimations of Kt/V (weekly), corrected creatinine clearance (CrCl), and residual renal function (weekly urea clearance) were derived from 24-h collections of spent dialysate and urine; nPCRs were calculated from total urea removal using the Randerson equation [15] with a correction for estimated body surface area.
Measurement of CarbHb
Blood for measurement of CarbHb was collected in 5-ml tubes containing lithium-heparin. CarbHb was measured by the method described by Kwan et al. [16]. Briefly, carbamyl valine was released from washed erythrocytes after hydrolysis with a mixture of acetic and hydrochloric acid. Valine hydantoin (VH), formed after the spontaneous dissociation of carbamyl valine, was extracted and quantitated by high-performance liquid chromatography, with results expressed as µg VH/g Hb.
Statistical methods
Statistical analysis was performed using a computer-based statistics program (Statistical Package for Interactive Data Analysis, Statistical Computing Laboratory, Sydney, Australia). Using CarbHb as the dependent variable, univariate linear regression was performed on all potential predictors using Pearson's or Spearman's correlation tests as appropriate. The t test was used to compare dichotomous variables and patient subgroups. Because of the likelihood that some of the univariate correlates were co-dependent, multivariate analysis was used to model CarbHb. Multivariate data are expressed as ß coefficient, standard error (SE), and probability.
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Results |
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Because of co-dependency of many of the variables measured, multivariate analysis with backward elimination was performed to model CarbHb. With this analysis, the strongest independent predictors were post-dialysis urea, serum albumin, and serum bicarbonate (Figure 1). Pre-dialysis urea and nPCR were not independently predictive of CarbHb, presumably because of their strong relationship to post-dialysis urea. The results of this multivariate analysis are shown in Table 3
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PD patients
CarbHb was collected in a total of 42 PD patients. CarbHb levels as well as patient characteristics, adequacy measurements and biochemical parameters for these patients are shown in Table 1. As expected, CarbHb levels in these patients were significantly higher than the range established for patients with normal renal function.
Univariate analysis (Table 4) identified a statistically significant correlation between CarbHb and serum urea (r=0.51, P=0.001) and nPCR (r=0.52, P=0.01). No correlation was found for patient factors (age (P=0.68), gender (P=0.43), months of dialysis (P=0.97)), adequacy measures (weekly Kt/V (P=0.78), corrected CrCl (P=0.58)), or biochemical parameters (albumin (P=0.43), phosphate (P=0.86), bicarbonate (P=0.75)). The extent of residual renal function measured by urea clearance similarly did not correlate with CarbHb levels (P=0.13).
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Using multivariate analysis (Table 5), serum urea was the only independent predictor of CarbHb identified (Figure 2
), presumably because of the significant co-dependence of urea and nPCR.
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Discussion |
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The correlation with urea in both groups of patients is not unexpected, given the concentration and time dependence of the carbamylation process in vitro [1]. This result is in agreement with previous studies of HD patients, which showed a relationship between CarbHb and both pre-dialysis urea [1,10,17] and the time-averaged concentration of urea (TACurea) [5,11,17]. Although TACurea was not formally measured in the HD patients in this study, a crude estimate of TACurea based on the average of pre- and post-dialysis urea levels was in fact the best urea-based predictor of CarbHb (r=0.48, P<0.001), and further improved mathematical modelling of CarbHb when incorporated into the multivariate analysis in place of post-dialysis urea. For PD patients, the relationship between CarbHb and urea is supported by the findings of a previous smaller study [2]. In PD patients the correlation coefficient for urea and CarbHb is slightly higher than in HD patients, presumably as urea levels in PD patients are a better estimate of TACurea.
In this study, CarbHb levels were found to be significantly higher in HD patients with higher serum albumin, a finding that has not been previously described. The reason for this relationship is unclear; however, it may relate to higher rates of urea generation in patients who are better nourished (and possibly better dialysed)a view that is supported by the positive correlation between CarbHb and the derived urea generation rate in a previous study by Kwan [17]. This finding somewhat weakens the case for CarbHb as a measure of adequacy, if lower levels of CarbHb are associated both with lower urea levels and lower levels of serum albumin, an established predictor of patient mortality [8,18]. This relationship did not appear in the PD patients in this analysis, possibly because of the smaller number of patients studied. Alternatively, it has been suggested that albumin levels are less useful as a marker of nutritional status in PD patients [19,20], due to the loss of albumin in the peritoneal effluent.
This study found a weak negative association between CarbHb and bicarbonate in HD patients, as has been previously reported [10]. Bicarbonate was measured in 33 of the 80 patients, but this group did not appear statistically distinct in terms of any of the other measured variables. This result suggests an effect of acidosis independent of urea to increase CarbHb. The cause of this finding is unclear, but as previous studies in HD patients have associated acidosis with increased nPCR and dietary protein intake [21], it is possible that this effect may relate to increased generation of urea in acidotic HD patients.
In both groups of dialysis patients, treatment with rHuEpo did not appear to effect CarbHb levels significantly. In the study by Davenport et al. [5] the group receiving rHuEpo had both significantly lower CarbHb levels and significantly higher reticulocyte counts, a result attributed to the contribution of newly synthesized and unexposed Hb proteins to the total Hb pool. Interestingly, we found that the stability of Hb over the 3 months prior to CarbHb measurement did not differ in the group receiving rHuEpo therapy either in HD or in PD patients, possibly explaining the lack of association found. If changes in Hb production and turnover do impact significantly on the levels of CarbHb, a further level of complexity is added to the interpretation of CarbHb levels that may impair its use in monitoring uraemia in dialysis patients.
This study found no relationship between CarbHb and conventional measures of dialysis dose in either HD or PD patients. This result is at variance with previous smaller studies of HD patients in which a relationship was found between CarbHb and both Kt/V [5] and the URR [5,10], and has not been previously described in PD patients. Reasons for the discordance between this result and those of earlier studies in HD patients are unclear, despite similar levels of CarbHb. Significantly, the achieved Kt/V and URR were higher in this study than in previous studies where a correlation was found, and it is possible that a relationship between CarbHb and measurements of urea removal may be weaker at a higher achieved dose. It is also possible that the composition of the dialysis membranes will contribute to the observed differences, as this study used modified cellulose or polysulphone membranes as opposed to the cellulose acetate membranes of earlier studies. Finally, earlier studies did not submit the correlation to multivariate analysis, possibly leading to the error of confounding.
There are therefore several lines of evidence that would argue against a significant role for CarbHb as a measure of dialysis adequacy. Firstly, CarbHb has a consistent relationship with serum urea in the majority of the dialysis patient series reported to date. Urea is a poor surrogate as a uraemic toxin, both because of the fact that its rate of generation varies with the nutritional state of the patient and because both high and low levels of urea are associated with excess mortality in dialysis patients [22]. Secondly, CarbHb does not appear to relate to conventional measures of dialysis adequacy in HD or PD patients, which, although not without their own limitations, have been shown to predict patient outcome. CarbHb levels also do not provide information relating to the dose of dialysis which may lead to modification of either the technique or dose of dialysis if levels are inadequate. Lastly, CarbHb levels appear to be lower in HD patients with a lower serum albumin, which is a known predictor of patient outcome [8,18]. The utility of CarbHb as a predictor of patient outcome has not been examined in detail, however. The available information comes from two previous small comparisons of CarbHb levels in patients dying over a short period of follow-up compared with their surviving counterparts [5,10]. The results of these analyses were conflicting (Davenport found higher levels of CarbHb in patients who died, and Tarif found no difference); however, the small numbers of patients examined makes the interpretation of these results hazardous. Large longitudinal studies of sufficient power to resolve a relationship between repeated measures of CarbHb and outcome are not available.
In summary, CarbHb is a marker of uraemia in both HD and PD patients, but several lines of evidence suggest that a single measure is unlikely to be a useful index of dialysis adequacy. Whether CarbHb has any utility as a predictor of patient outcome or is just a surrogate measure of urea will remain unclear until large, longitudinal studies are performed.
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Acknowledgments |
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Notes |
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References |
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