Reconciling Estimates of Cell Proliferation from Stable Isotope Labeling Experiments

Area of Science:

• Immunology
• Biochemistry
• Isotope Tracing

Background:

• Stable isotope labeling is crucial for in vivo lymphocyte kinetics quantification.
• Discrepancies exist between deuterated glucose (D2-glucose) and deuterated water (D2O) labeling methods, impacting immune function studies.
• Previous studies have not directly compared D2-glucose and D2O labeling under identical conditions.

Purpose of the Study:

• To directly compare D2-glucose and D2O stable isotope labeling techniques for lymphocyte kinetics.
• To identify the cause of discrepancies in proliferation rate estimates between the two methods.
• To improve the accuracy and reliability of lymphocyte kinetic measurements.

Main Methods:

• Performed parallel in vitro and murine in vivo labeling experiments using identical protocols with both D2-glucose and D2O.
• Analyzed normalization accuracy in blood glucose enrichment for D2-glucose studies.
• Re-analyzed published human studies using corrected normalization methods.

Main Results:

• Intrinsic biochemical differences between D2-glucose and D2O do not explain proliferation rate discrepancies.
• In vivo D2-glucose labeling estimates are susceptible to normalization errors due to variable blood glucose enrichment.
• Corrected normalization reduced D2-glucose proliferation estimates and slightly increased D2O estimates, aligning the methods.

Conclusions:

• Normalization accuracy is critical for reliable in vivo lymphocyte kinetics using stable isotope labeling.
• The D2-glucose method requires careful normalization to ensure accurate proliferation rate estimation.
• This study reconciles discrepancies between D2-glucose and D2O labeling, enhancing confidence in lymphocyte kinetics quantification.