Metabolic effects of albumin therapy in acute lung injury measured by proton nuclear magnetic resonance spectroscopy of plasma: a pilot study

Abstract

Objective: Improved means to monitor and guide interventions could be useful in the intensive care unit. Metabolomic analysis with bioinformatics is used to understand mechanisms and identify biomarkers of disease development and progression. This pilot study evaluated plasma proton nuclear magnetic resonance spectroscopy as a means to monitor metabolism following albumin administration in acute lung injury patients.

Design: This study was conducted on plasma samples from six albumin-treated and six saline-treated patients from a larger double-blind trial. The albumin group was administered 25 g of 25% human albumin in 0.9% saline every 8 hrs for a total of nine doses over 72 hrs. A 0.9% concentration of saline was used as a placebo. Blood samples were collected immediately before, 1 hr after, and 4 hrs after the albumin/saline administration for the first, fourth, and seventh doses (first dose of each day for 3 days). Samples were analyzed by proton nuclear magnetic resonance spectroscopy, and spectra were analyzed by principal component analysis and biostatistical methods.

Interventions: None.

Measurements and main results: After 1 day of albumin therapy, changes in small molecules, including amino acids and plasma lipids, were evident with principal component analysis. Differences remained 3 days after the last albumin administration. Analysis of data along with spectra from healthy controls showed that spectra for patients receiving albumin had a trajectory toward the spectra observed for healthy individuals while those of the placebo controls did not.

Conclusion: The data suggest that metabolic changes detected by proton nuclear magnetic resonance spectroscopy and the bioinformatics tool may be a useful approach to clinical research, especially in acute lung injury.

Figure 1. Experimental design to measure effects of albumin treatment in acute lung injury

Twelve mechanically ventilated patients with ALI/ARDS whose serum total protein concentrations were <6 g/dL were studied. Patients were equally randomly allocated to receive furosemide with albumin or furosemide with placebo for 72 h. Each patient was administered albumin or placebo every 8 h for a total of 9 doses indicated by upward arrows. Blood samples (a, b and c) were collected surrounding the 1^st (Day 1), 4^th (Day 2), and 7^th (Day 3) doses, immediately prior to, 1 h after and 4 h after the albumin/placebo administration as indicated by the downward arrows. A sample was also collected in the morning on Day 7, i.e., 3 d after the last dose.

Figure 2. PCA of ¹H-NMR spectra of plasma samples from albumin- or placebo-treated acute lung injury patients

Samples were color-coded green for the baseline samples taken on Day 1 just prior to the first dose of albumin or placebo. Samples from patients after receiving albumin are color-coded red and those receiving placebo are color-coded blue. A. PCA of mean-centered data, where the first 3 Principal Components (PC) accounted for 90% of the variance, showed some separation according to albumin treatment. This analysis is biased toward high abundance features, and the corresponding loading plot showed that lipids, which have a relatively high abundance in the ¹H-NMR spectrum. B. Better separation by PCA was obtained following variance scaling to reduce bias toward high-abundance signals. The corresponding loading plot showed that spectral regions associated with lipids and amino acids contributed to the separation.

Figure 3. PCA of samples according to day of treatment

All samples were plotted in A–C. A. On Day 1, there was no separation of albumin-treated (red) or placebo-treated (blue) samples from the baseline samples (green) taken before initiation of treatment. The samples of other days were color-coded cyan. B. On Day 2, samples from albumin-treated patients were clearly separated from placebo-treated patients. C. On Day 3, samples from albumin-treated patients were separated from placebo-treated patients. Data were analyzed with variance scaling.

Figure 4. PCA of ¹H-NMR spectral data for albumin-treated patients on Day 7 compared to baseline samples taken before the first albumin or placebo treatment

Samples were color coded green for baseline, blue for placebo and black for albumin on Day 7. Results showed that 3 days after last albumin treatment, metabolic differences showed a trend in patients receiving albumin. Corresponding plot for patients receiving placebo showed no separation. Data were analyzed with mean centered.

Figure 5. ¹H-NMR spectra of plasma from albumin-treated patients shows a trajectory toward spectra obtained from healthy individuals

PCA of plasma obtained at baseline (green), in patients after receiving placebo (blue), and patients after receiving albumin (red) were combined with data obtained from fasting, healthy individuals after equilibration to a nutritionally adequate diet (black). Results show that albumin treatment resulted in a trajectory from the baseline conditions toward the healthy profiles. Data were analyzed with variance scaling.

Figure 6. Comparison of selected ¹H-NMR spectral regions between albumin- and placebo-treated patients

Analyses were performed on spectral regions which contributed to separation in PCA analyses and also some selected regions associated with plasma metabolites of interest. Spectral peaks were integrated and analyzed by repeated measures ANOVA. Open circle represented albumin treated group and close circle was placebo group. Results for albumin effects, time effects and albumin*time effects are given with panel descriptions. Because this was a pilot study and specific time-dependent differences were of interest, t-tests for specific sample times were performed and significance at p < 0.05 are identified in the panels by an asterisk (*). A. 0.66 ppm, HDL; albumin (p=0.46), time <0.05 and albumin*time (p=0.09). B. 1.25,1.29 ppm, LDL, VLDL; albumin (p=0.67), time (p=0.49) and albumin*time (p=0.49). C. 1.33 ppm, lactate; albumin (p=0.39), time (p=0.063) and albumin*time (p=0.26). D. 1.00 ppm, isoleucine; albumin (p=0.34), time <0.05 and albumin*time (p=0.50). E. 0.97,1.02 ppm, valine; (p=0.32), time <0.05 and albumin*time (p=0.46). F. 1.46 ppm, alanine; albumin (p=0.28), time <0.05 and albumin*time (p=0.36). G. 7.77 ppm, 1-methylhistidine (Me-His); albumin (p=0.057), time <0.05 and albumin*time (p=0.10). H. 2.08,2.09 ppm, glutamine, glutamate (Gln, Glu); albumin (p=0.45), time (p=0.23) and albumin*time (p=0.43). I. 5.23 ppm, glucose; albumin (p=0.35), time (p=0.43) and albumin*time (p=0.40). J. 2.89 ppm, albumin (lysyl) (Albumin-lys); albumin (p=0.30), time (p=0.11) and albumin*time (p=0.40).