Cerebrospinal fluid–based kinetic biomarkers of axonal transport in monitoring neurodegeneration

Abstract

Progress in neurodegenerative disease research is hampered by the lack of biomarkers of neuronal dysfunction. We here identified a class of cerebrospinal fluid-based (CSF-based) kinetic biomarkers that reflect altered neuronal transport of protein cargo, a common feature of neurodegeneration. After a pulse administration of heavy water ( 2 H 2 O), distinct, newly synthesized 2 H-labeled neuronal proteins were transported to nerve terminals and secreted, and then appeared in CSF. In 3 mouse models of neurodegeneration, distinct 2 H-cargo proteins displayed delayed appearance and disappearance kinetics in the CSF, suggestive of aberrant transport kinetics. Microtubule-modulating pharmacotherapy normalized CSF-based kinetics of affected 2 H-cargo proteins and ameliorated neurodegenerative symptoms in mice. After 2 H 2 O labeling, similar neuronal transport deficits were observed in CSF of patients with Parkinson's disease (PD) compared with non-PD control subjects, which indicates that these biomarkers are translatable and relevant to human disease. Measurement of transport kinetics may provide a sensitive method to monitor progression of neurodegeneration and treatment effects.

In vivo kinetic approach reveals slow SOD1 turnover in the CNS

“…Misfolded SOD1 G93A had extremely rapid kinetics in the liver, faster than those of the total soluble SOD1 G93A pool and much faster than those of the misfolded protein pool in the spinal cord. Interestingly, misfolded SOD1 G93A in the liver peaked higher than the peripheral plasma-free 13 C 6 -leucine in the spinal cord (15.9 days) and cortex (9.3 days) was 2.8 to 9 times slower than the half-life in the liver (1.8 days) or kidneys (3.4 days).…”

“…The development of stable isotope-labeling kinetics (SILK) has enabled the study of protein turnover rates in vivo using a safe, stable isotope amino acid tracer that is incorporated into newly synthesized proteins and can be quantitatively measured by mass spectrometry -a technique that has been highly successful in studies of amyloid-β in human cerebral spinal fluid (CSF) and in brains from animal models (5)(6)(7)(8)(9)(10)(11)(12). In these studies, a relatively short (9-hour) intravenous infusion of the stable isotope 13 C 6 -leucine resulted in adequate labeling of the rapidly turned over amyloid-β (half-life of 8 hours). However, the measurement of the turnover of long-lived proteins is challenging, especially in humans.…”

“…The labeling time needed becomes too long for an intravenous approach, and sample collection must be carried out over many days instead of hours. Indeed, D 2 O-labeling studies in both animal models and human subjects have demonstrated that long-term labeling is necessary to measure proteins with a half-life on the order of days (13,14). Here, we report the development of a SILK method using a stable isotope-labeled amino acid to measure the turnover of a long-lived protein in rodents and in human CSF.…”

“…The calculation of protein turnover by administering stable isotope-labeled amino acids or D 2 O over time has been demonstrated in numerous cell culture models and in human CSF and plasma proteins (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)23). Stable isotope-labeled amino acids are biologically identical to their naturally occurring counterparts and, unlike radiolabeled amino acids, are innocuous to both the system being studied and the experimental environment.…”

Protein kinetic signatures of the remodeling heart following isoproterenol stimulation

“…Measuring protein turnover in vivo entails additional technical challenges including label delivery and tolerance, determination of precursor enrichment, and data interpretation (3,12,16). Stable isotope labeling using deuterium oxide ( 2 H 2 O) tracers has shown great potential for tracing protein turnover in mammals (17)(18)(19)(20)(21). However, widespread adjudged to capture the turnover of most cardiac proteins (16).…”