Markers in cerebrospinal fluid provide clues to a person’s risk of Alzheimer’s disease, but such tests still fall short of a definitive diagnosis. Two recent papers propose new markers to improve the predictive power of CSF testing. Each presents a potential biomarker that tracks with cognition and sharpens diagnosis in preliminary studies. In the May 19 Nature Communications, researchers led by Ashley Bush at the University of Melbourne, Australia, report that high CSF ferritin, a protein that reflects the iron load in the brain, predicts earlier progression to dementia. Meanwhile, in the May 5 Translational Psychiatry, researchers led by Sergio Ferreira and Rogerio Panizzutti at the Federal University of Rio de Janeiro, Brazil, proposed the neuromodulator D-serine as a marker of Alzheimer’s. In a small study, patients had fivefold higher CSF levels of this amino acid than age-matched controls.
If the findings hold up in larger studies, both molecules would make attractive markers, as they are relatively easy to measure reliably in CSF, noted Henrik Zetterberg at the University of Gothenburg, Sweden, who was not involved in either study. Zetterberg was even more intrigued by the clues to disease mechanisms provided by the data, and said both findings should stimulate more basic research.
The Australian researchers have long been interested in the link between brain iron and Alzheimer’s. Some histological and MRI studies detect more iron in the brains of patients, while others report changes in its oxidation state (see Jellinger et al., 1990; Bartzokis et al., 2004; Quintana et al., 2006). Since ferritin is the main carrier of iron in the body, elevated CSF ferritin might indicate a higher brain iron burden. Previous studies gave mixed results, however, with only some reporting high CSF ferritin in AD patients (see Kuiper et al., 1994; Craig-Schapiro et al., 2011; Paterson et al., 2014).
To resolve this, joint first authors Scott Ayton and Noel Faux analyzed data from the Alzheimer’s Disease Neuroimaging Initiative, which included ferritin as a CSF analyte. In agreement with previous negative findings, ferritin levels were similar between 91 cognitively healthy controls, 144 people clinically diagnosed with mild cognitive impairment, and 67 AD patients. However, the authors saw something interesting when they looked at cognition: The amount of ferritin correlated inversely with a person’s cognitive scores, regardless of their diagnosis. The authors had to use different cognitive tests to detect decline in AD patients versus those at earlier stages. For AD patients, people in the highest tertile of ferritin levels scored about three points worse on the ADAS-Cog13 than did those in the lowest. The magnitude of the cognitive difference was similar to that seen in people with a high CSF tau/Aβ42 ratio. Likewise, controls and MCI patients who had the highest levels of ferritin scored lower on a verbal learning test than their peers with less ferritin. Unlike tau/Aβ42, however, ferritin levels did not associate with a higher rate of cognitive decline over time. Instead, ferritin correlated with a set amount of cognitive loss at all stages of the disease.
The authors took this data to mean that high CSF ferritin predicted earlier progression to dementia. They calculated that an elevation of ferritin by one standard deviation over the mean of the baseline population corresponded to a 9½-month-earlier diagnosis of AD. As has been seen in other studies, high CSF tau/Aβ42 and low CSF ApoE independently predicted faster disease progression (see Toledo et al., 2014). Combining ferritin with these markers improved the sensitivity and specificity of an AD diagnosis, the authors claim.
“This is the first time that ferritin has been reported as being of diagnostic value in Alzheimer’s disease. It looks to be as valuable as the gold-standard CSF biomarkers,” Bush told Alzforum. He is attempting to replicate the results in a separate cohort of MCI and AD patients. He is also comparing ferritin levels across several neurodegenerative diseases to find out if the marker is specific for AD. Brain iron accumulation has also been reported in Parkinson’s disease.
Commentators found the implications of the data intriguing. “This well-executed study points to a widespread change in iron metabolism in Alzheimer’s disease that is associated with cognition,” said George Perry at the University of Texas, San Antonio. “It advances research in this area substantially.”
The data underscore a potentially toxic effect of brain iron, Zetterberg said. He noted that when blood leaks into brain tissue, it causes a rare disease known as superficial CNS siderosis. Iron accumulates in the cortex, and tau becomes hyperphosphorylated. Left untreated, the condition leads to neurodegeneration and dementia. The disorder is seven times more common in AD patients than in the general population (see Feb 2014 news). “Iron poisoning of the brain may injure neurons and reduce function,” Zetterberg said. In the ADNI data, high baseline ferritin levels correlated with greater atrophy of the hippocampus over the next six years, supporting a role in neurodegeneration.
Pieter Jelle Visser at VU University Medical Center in Amsterdam agreed. “Given the relatively strong correlation of high ferritin with increased CSF tau and the weak association with increased CSF Aβ42, it may be possible that ferritin reflects neurodegeneration rather than an event associated with the onset of the disease. The current evidence seems insufficient to recommend studies on lowering brain iron, but rather emphasizes the need for further cross-sectional and longitudinal studies to define the role of iron in AD,” he wrote to Alzforum (see full comment below).
The study raises numerous questions. What causes brain iron to rise? Bush and colleagues claim that APP and tau help export iron from neurons, suggesting that dysfunction of these molecules could allow the metal to build up in AD (see Feb 2012 news). Zetterberg suggested that microbleeds, which often occur in blood vessels peppered with amyloid, might help kick off iron accumulation. One surprise in the data was that CSF ApoE levels correlated strongly but inversely with ferritin levels. The effect was particularly strong in people who carried an ApoE4 allele; they had lower CSF ApoE and higher CSF ferritin than their peers. This hints at a role for ApoE in iron metabolism.
The Brazilian study took a different tack in the race for biomarkers, focusing on D-serine, an enantiomer of the common L-amino acid. Serine racemase converts the L to the D form and is expressed by neurons, astrocytes, and microglia. D-serine is abundant in mammalian brains. It acts as a co-agonist with glutamate, binding to the glycine site of NMDA receptors to help activate these channels, which are essential for learning and memory (for review, see Wolosker et al., 2008). In Alzheimer’s, extrasynaptic NMDA receptors may become overactive, leading to synapse damage and neuron death. The approved AD drug memantine blocks these channels. In animal models, D-serine contributes to excitotoxicity at NMDA receptors (see Dec 2011 conference news; Mustafa et al., 2010).
To investigate D-serine in Alzheimer’s pathology, joint first authors Caroline Madeira and Mychael Lourenco examined postmortem brains from 17 AD patients and 12 controls. They found a twofold elevation of D-serine in the patients’ hippocampuses and parietal cortices, but not in their occipital cortices, a region relatively spared by AD.
The authors wondered if the excess would show up in CSF, as suggested by one previous study (see Fisher et al., 1998). In a pilot study on 21 AD patients, the authors saw fivefold higher levels of D-serine than in 10 age-matched controls, and about twofold higher levels than in nine patients with hydrocephalus and another nine with depression. High D-serine correlated with poorer performance on the Mini-Mental State Exam and a worse Clinical Dementia Rating, regardless of a person’s diagnosis, suggesting an effect on cognition.
Moreover, D-serine analysis improved AD diagnosis in this small study. In the Brazilian cohort, the standard CSF biomarker tau/Aβ42 detected Alzheimer’s with 81 percent sensitivity and 94 percent specificity, and adding D-serine into the equation improved sensitivity to 96 percent and specificity to 100 percent. To validate the findings, Ferreira collected CSF from another 160 AD patients in Brazil, and will compare their levels of D-serine and other neurotransmitters to those of age-matched controls. Zetterberg suggested that it would be valuable to measure D-serine in other neurodegenerative diseases such as Parkinson’s as well, to determine if this marker is specific for AD.
What might explain such extreme levels of D-serine? To investigate, the authors used cell cultures. Adding synthetic Aβ42 oligomers to hippocampal neuron cultures for 24 hours pumped up levels of D-serine, as well as mRNA and protein levels of serine racemase, the enzyme that produces D-serine. Transgenic APPPS1 mice also displayed high levels of D-serine and serine racemase in the hippocampus compared to littermate controls. The results indicate that Aβ oligomers somehow turn up expression of serine racemase. Possibly, this is a compensatory response, Ferreira speculated. NMDA receptor levels fall early in AD, which may cause microglia to boost serine levels in an attempt to restore signaling. “The data support this notion that NMDA receptor signaling becomes deregulated in the AD brain,” Ferreira said.
The findings agree with a previous study from Steve Barger at the University of Arkansas for Medical Sciences in Little Rock. He reported that Aβ stimulates cultured microglia to make more serine racemase, and the resulting boost in D-serine amplifies toxicity (see Wu et al., 2004). “Together, the data suggest that serine racemase and its product provide a mechanistic link between neuroinflammation and neurodegeneration,” Barger wrote to Alzforum (see full comment below).—Madolyn Bowman Rogers
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