Carbon Monoxide-based Rebreathing Method and Bioimpedance in Hemodialysis Patients

Aim:

1. To combine a carbon monoxide rebreathing method and bioimpedance which allows to objectively measure the amount of fluid in the bloodstream (intravascular) and throughout the body, and thereby test the actual amount of fluid in the body when patients are hydrated according to their clinical dry weight.
2. To validate carbon monoxide rebreathing method in hemodialysis patients compared against a radioactive labeling of erythrocytes and albumin (the gold standard).
3. To investigate whether hemoglobin is a valid marker of anemia in this patient population by measuring the erythrocyte volume and hemoglobin mass by the carbon monoxide rebreathing method and correlating this to the hemoglobin concentration.

Background:

Anemia and overhydration are challenging conditions in patients on hemodialysis. Overhydration is associated with hypertension, cardiomyopathy and increased mortality. In assessing the patient's volume status, blood pressure, body weight and peripheral edema are used as indicators. To achieve a more accurate estimate, these measures have been combined with various techniques, such as relative changes in blood volume (measured with blood volume sensor (BVS)) and bioimpedance. These methods are used to varying degrees in everyday clinical practice.

In case of overhydration, fluid is distributed intravascularly and can thereby expand blood volume. The primary clinical problem with an expanded blood volume is hypertension and increased risk of cardiovascular complications. Measurement of the intravascular volume can therefore have great clinical relevance, but currently there are no direct measures for this implemented in the clinical daily routine. Various techniques have been used for research purposes. These include fluorescent dilution techniques, radioactive labeling of erythrocytes and albumin, as well as infusion of dialysate during dialysis where changes in BVS measurements can be used to calculate the intravascular volumes.

Bioimpedance can be used to measure the total water phase and the intra- and extracellular volume. The method has been shown to be useful in the diagnosis of overhydration in hemodialysis patients. Overhydration detected by bioimpedance in hemodialysis patients has been shown to correlate to increased mortality. Bioimpedance is a non-invasive, painless and practically useful way of assessing the volume status.

Anemia is another significant challenge in patients on hemodialysis. The majority of patients on hemodialysis have a significant decrease in endogenous erythropoietin production, which causes anemia and requires treatment with an erythrocyte stimulant (ESA). The basis for anemia diagnosis and the decision on ESA treatment in hemodialysis patients is based on pre-dialysis hemoglobin (measured at dialysis start-up). Upon overhydration, fluid distributed to the intravascular space will lead to dilated plasma volume (PV) and dilution of hemoglobin. This dilution and thus artificial decrease in hemoglobin makes pre-dialysis hemoglobin difficult to interpret in patients on dialysis. Time-averaged hemoglobin, calculated on the basis of pre-dialysis and post-dialysis hemoglobin, has been shown to correlate better with the actual hemoglobin of hemodialysis patients, but the value has not gained ground in practice.

The carbon monoxide rebreathing method is a well-proven technique for determining intravascular volume, (blood volume, BV), PV and red blood cell volume (RBCV). In renally healthy patients, the method is validated against a radioactive labeling technique (gold standard) where high correlation (r = 0.97) has been demonstrated between the two methods. The carbon monoxide rebreathing method has also been shown to be able to detect even very small changes in blood volume. The method is safe and requires 6 minutes of carbon monoxide inhalation. A prerequisite for using the carbon monoxide rebreathing method is a normal diffusion of gas over the lungs, which allows uptake into the blood. Dialysis patients may be significantly overhydrated and possible accumulation of water in the alveoli may decrease the diffusion of carbon monoxide and affect the method. Against this background, the method is to be validated against a radioactive labeling technique.

The validity of the carbon monoxide rebreathing method in hemodialysis patients has not been performed which is important if the method is to be implemented clinically and for research purposes.

Method:

Case-control study with 25 patients in each group. The case group consists of hemodialysis patients and the control group consists of healthy individuals with eGFR above 60 ml/min/1.73m2. Subjects were matched on gender, age (+/- 5 years) and (weight +/- 10kg).

Hemodialysis group: 25 hemodialysis patients are included. Immediately before the start of dialysis, the total body water is measured by bioimpedance and blood samples are taken. A normal hemodialysis lasts 3-4 hours and during this period the patient undergoes fluid extraction corresponding to their estimated dry weight. 30 minutes after the dialysis is completed, the total boby water is measured with bioimpedance and the blood volume, plasma volume and erythrocyte volume are measured by carbon monoxide rebreathing method. In 10 participants, blood volume, plasma volume and the erythrocyte volume are measured by a radioactive labeling of erythrocytes and albumin, where blood is taken at the beginning of dialysis and cleaned at the end of dialysis. Hemoglobin and hematocrit are measured pre-dialysis, post-dialysis and 30 minutes after cessation of dialysis. Participation in the study involving the radioactive labeling of erythrocytes and albumin may be omitted if a participant does not wish the study.

Control group: Includes 25 patients with eGFR above 60 ml/min/1.73m2 and without severe heart failure, liver failure or lung disease. In all participants, the intravascular volume is measured by carbon monoxide rebreathing method and the total body water by bioimpedance. In 10 participants, blood volume is measured by a radioactive label of erythrocytes and albumin. Hemoglobin and other standard samples are measured.

If a participant after inclusion must be withdrawn, the patient is replaced by a new participant until a number of 25 participants in each group is achieved. If a matched participant cannot be found for an included participant, the person in question is withdrawn and replaced by a new one.

Statistics:

Blood volume in patients hemodialysis has been found to be 73.4 ml / kg (SD 15 ml / kg) and from the predicted normal blood volume has been found to be 59.0 ml / kg (SD 9.3 ml / kg). In a two-sided unpaired t-test with α = 0.05, SD = 15 and a power of 90%, a sample size of 23 patients in each group will show a significant difference. A dropout rate of approximately 2 patients is assumed and a total of 25 are included in each group.

After completion of the study and data completion the results are analysed according to primary and secondary endpoints. Results are reported as mean values with confidence interval or median and interquartile range. Data are analysed with parametric (normally distributed data) or non-parametric statistics (non-normal distributed data). A 95 % confidence interval is accepted as statistically significant (p < 0.05). Analysis are performed as a per protocol analysis.