Plasma Neurofilament Light Levels Research Articles

A thorough analysis of the molecular mechanisms underlying the protein changes in the human cerebral cortex affected by Alzheimer's disease

Alzheimer's disease (AD) is a diverse condition with an intricate pathobiology. Cortex plays a role in regulating body movements, executive function, and cognitive control and serves as a hub for replaying information in the auditory and visual systems. There is a correlation between the rate of change in cortical thickness, cerebrospinal fluid, plasma neurofilament light levels, and cognitive function. However, cognitive impairment is typically closely associated with the decline of neurons and synapses in different regions of the cortex. Thus, more endeavor has been made to understand the pathophysiology of AD involved in the perturbation in the cortex, but fundamental molecular mechanisms related to protein changes that happen in the cortex region remain unanswered. We aimed to investigate the molecular mechanisms implicated in the etiology of Alzheimer's disease in eight distinct regions of the human cortex. Using mass-spectrometry-based proteomic data from 29 published studies until March 15, 2022, hub proteins and protein-protein interactions, signaling pathways, miRNAs, and transcription factors were analyzed. Nine proteins—GFAP, APP, HSPB1, CD44, CLU, RPH3A, VGF, CAPG, and CORO1A—were prevalent in all of the studies. APP, ALB, and HSP90AA1 were found in 4/8, 3/8, and 3/8 regions of the cortex, respectively. Although the putative proteins obtained from various areas of cortex differed, important molecular pathways were identified, including impaired neuronal systems, mitochondrial and ribosomal functions, activated innate immune systems and microglia, and activated NMDA receptors. The primary microRNA (hsa-miR-16-5p) and transcription factor (CREB3L3) were the main factors responsible for the etiology of AD related to protein changes. Focusing on the proteins, miRNAs, transcription factors, and molecular processes in the cortex plays a crucial role in managing AD.

Plasma neurofilament light as a promising biomarker in neuronal intranuclear inclusion disease.

Neuronal intranuclear inclusion disease (NIID) is a rare neurodegenerative disorder lacking reliable biomarkers. This study investigates plasma protein levels as potential biomarkers of disease severity and progression in NIID. In this study, we enrolled 30 NIID patients and 36 age- and sex-matched controls, following them for 1-2years. Plasma neurofilament light (NfL), glial fibrillary acidic protein (GFAP), ubiquitin carboxy-terminal hydrolase L1 (UCH-L1), and tau were measured using ultrasensitive single molecule array (Simoa) assays. Disease severity was evaluated with the Mini-Mental State Examination (MMSE), Montreal Cognitive Assessment (MoCA), Activities of Daily Living (ADL), and CNS symptom counts, in addition to neuroimaging data. Our study revealed that NIID patients has significantly higher plasma NfL (median, 35.2 vs. 8.61pg/mL, p < 0.001) and GFAP (102 vs. 79.0pg/mL, p = 0.010) levels compared to controls, with NfL emerging as a robust diagnostic marker (AUC = 0.956). NfL levels were notably higher in acute-onset NIID (77.5 vs. 28.8pg/mL, p = 0.001). NfL correlated strongly with disease severity, including MMSE (ρ = - 0.687, p < 0.001), MoCA (ρ = - 0.670, p < 0.001), ADL (ρ = 0.587, p = 0.001), CNS symptoms (ρ = 0.369, p = 0.045), and white matter hyperintensity volume (ρ = 0.620, p = 0.004). Higher baseline NfL (≥ 35.2pg/mL) associated with increased ADL scores, CNS symptoms, and white matter hyperintensity at follow-up. UCH-L1 and total tau levels showed no significant differences. Our results suggested the potential of NfL as a promising biomarker of disease severity and progression in NIID.

Genome‐wide association meta‐analysis of plasma neurofilament light levels in high‐risk adults with Down Syndrome

AbstractBackgroundOnly a few studies in the general population have investigated genetic contributions to variation in levels of neurofilament light (NfL), which is a general marker of axonal damage and is predictive of Alzheimer’s disease (AD). To our knowledge, no studies have examined this relation in people with Down Syndrome (DS), a population at high risk of developing AD. We performed a genome‐wide search for SNPs associated with plasma NfL levels. We paid special attention to genetic variants identified in the general population to be associated with AD risk and NfL to evaluate whether those variants contributed to AD in DS by altering NfL levels.MethodsWe performed a meta‐analysis using two datasets (N = 455): the omicsADDS and ABC‐DS studies. Levels of plasma NfL were measured using the Simoa platform and genotyping was performed using an Illumina Infinium General Screening Array v2. Imputation on autosomal chromosomes other than 21 was performed using the TOPMed Imputation Server. We first assessed the association between SNPs and plasma NfL levels in each dataset using a multivariable linear regression model that included age, sex, dementia status, and genetic ancestry. We then performed meta‐analysis using sample size weighting. We restricted our meta‐analysis to variants that had study‐specific P&lt;5×10−4, to minimize potential false positives.ResultsOur genome‐wide scan, after Bonferroni correction, identified 33 novel SNPs contributing to variation in plasma NfL (28 variants with elevated plasma NfL and 5 variants with decreased plasma NfL). Three AD‐risk variants previously identified in the general population (UNC5CL/rs10947943, USP6NL/rs7912495, FOXF1/rs16941239), were found to be associated with variation in plasma NfL levels at nominal Pmeta &lt; 0.05. Candidate variants associated with NfL in the general population were not replicated in the current meta‐analysis. The novel genes identified have been implicated in tau neuropathology, variation in lipid levels, insulin regulation, and neurotransmitter release.ConclusionThis genome‐wide meta‐analysis revealed multiple novel genetic loci associated with NfL levels in plasma in adults with DS. We additionally found supporting evidence for three known variants. The genes identified here may lead to insight into underlying mechanisms of neurodegeneration and AD in adults with DS.

Using the axonal protein neurofilament light to distinguish psychiatric and neurodegenerative disorders across a program of clinical research studies

Introduction:There is an urgent need for clinical blood biomarkers which can rule in/out neurological disorders early in those with psychiatric symptoms, personality or behavioural changes and/or functional decline together with cognitive symptoms. The neuronal axonal protein neurofilament light (NfL) is released from damaged neuronal axons and can be measured in in blood and cerebrospinal fluid (CSF). We have undertaken a series of studies aimed at examining the clinical utility of blood and CSF NfL in assisting with the distinction between psychiatric and neurodegenerative / neurological disorders.Methods:Since 2016 we have measured blood and CSF NfL levels across multiple psychiatric and neurological populations recruited through Neuropsychiatry, Royal Melbourne Hospital and our collaborators (national and international). We have described our findings in a series of published studies. Data from our ongoing work, in larger cohorts and diagnostic groups, will be presented. The diagnostic groups include people with psychiatric disorders (schizophrenia, bipolar disorder, depression, functional neurological disorders), neurodegenerative disorders (Alzheimer’s disease, frontotemporal dementia, Huntington’s disease, Niemann-Pick Type C) and neurological disorders (e.g., epilepsy).Results:Our initial pilot study (n=129) found that CSF NfL was a promising biomarker in differentiating psychiatric from neurological disorders. In our larger follow up larger study (n=498) which included more diagnostic groups CSF NfL levels exhibited high accuracy (91%), sensitivity (92%), and specificity (87%) in differentiating psychiatric from neurological disorders, and distinguished behavioural variant frontotemporal dementia from frontal lobe syndrome phenocopies/mimics, with high accuracy. We have found that NfL is not elevated in people with treatment resistant schizophrenia compared to controls and is elevated in people with Niemann-Pick Type C compared to people with psychiatric disorders and controls. Further (unpublished) data has shown that these findings are replicated with plasma NfL levels across 400 further psychiatric, neurological and control participants.Conclusions:NfL is a highly promising biomarker which differentiates psychiatric from neurological disorders with high sensitivity and specificity. The translation of NfL levels into standard clinical practice could substantially improve the clinical diagnostic process in people with complex neuropsychiatric and cognitive disorders.

Relationship between generalization performance and plasma markers pTau231, pTau181 and NfL in cognitively healthy older African Americans

AbstractBackgroundPhosphorylated tau (pTau) in cerebrospinal fluid (CSF) is a biomarker of Alzheimer’s disease (AD) pathology. Recent studies confirm that pTau levels in plasma may provide a more cost‐effective and less invasive method of assessing AD pathology, which have shown to also be relevant in preclinical populations. Neurofilament light (NfL) is a measure of neuroaxonal damage associated with poorer cognitive outcomes. Generalization is a validated cognitive marker of hippocampal dysfunction, with individuals showing deficits years before overt cognitive impairment. Using a computer generalization task sensitive to medial temporal lobe (MTL) pathology, the current study examined the relationship between generalization performance and plasma pTau and NfL levels in a population with elevated AD risk, older African Americans.Method130 older, cognitively normal African Americans (M age = 71.60 years, SD = 6.58; M edu = 13.88 years, SD = 2.37; n = 98 women) provided blood samples and completed a hippocampal‐dependent cognitive task. Participants’ normal cognitive status was confirmed using the Mini‐Mental State Examination (M MMSE = 26.98, SD = 2.14) and generalization performance was assessed using the Acquired Equivalence Task. Plasma was extracted from blood samples to assess pTau181, pTau231, and NfL levels. Based on the sample median thresholds, participants were characterized as plasma tau positive (≥16.189 for pTau181 and ≥17.537 for pTau231).ResultParticipants positive for plasma pTau231 (n = 76) demonstrated worse generalization performance with increased pTau231 levels, B = ‐0.296, p = 0.024, as did participants positive for plasma pTau181, B = ‐0.247. p = 0.042, demonstrating that tau positivity status is related to MTL dysfunction. Moreover, pTau231 was more strongly associated with generalization performance, indicating that it is a more sensitive biomarker than pTau181 among preclinical AD populations. Higher levels of plasma NfL were also significantly associated with lower generalization performance B = ‐0.243. p = 0.008.ConclusionPlasma isoforms pTau181 and pTau231 may reflect different phases of tau progression, with plasma pTau231 shown to be better at capturing subtle MTL dysfunction. Plasma tau positivity status and measures of axonal damage along with lower generalization performance scores may be useful indicators of cognitive dysfunction in preclinical AD populations.

Late Blood Levels of Neurofilament Light Correlate With Outcome in Patients With Traumatic Brain Injury.

Neurofilament light (NF-L) is an axonal protein that has shown promise as a traumatic brain injury (TBI) biomarker. Serum NF-L shows a rather slow rise after injury, peaking after 1-2 weeks, although some studies suggest that it may remain elevated for months after TBI. The aim of this study was to examine if plasma NF-L levels several months after the injury correlate with functional outcome in patients who have sustained TBIs of variable initial severity. In this prospective study of 178 patients with TBI and 40 orthopedic injury controls, we measured plasma NF-L levels in blood samples taken at the follow-up appointment on average 9 months after injury. Patients with TBI were divided into two groups (mild [mTBI] vs. moderate-to-severe [mo/sTBI]) according to the severity of injury assessed with the Glasgow Coma Scale upon admission. Recovery and functional outcome were assessed using the Extended Glasgow Outcome Scale (GOSE). Higher levels of NF-L at the follow-up correlated with worse outcome in patients with moderate-to-severe TBI (Spearman's rho = -0.18; p < 0.001). In addition, in computed tomography-positive mTBI group, the levels of NF-L were significantly lower in patients with GOSE 7-8 (median 18.14; interquartile range [IQR] 9.82, 32.15) when compared with patients with GOSE <7 (median 73.87; IQR 32.17, 110.54; p = 0.002). In patients with mTBI, late NF-L levels do not seem to provide clinical benefit for late-stage assessment, but in patients with initially mo/sTBI, persistently elevated NF-L levels are associated with worse outcome after TBI and may reflect ongoing brain injury.

SIMOA-based analysis of plasma NFL levels in MCI and AD patients: a systematic review and meta-analysis

BackgroundThe single-molecule array assay (SIMOA)-based detection of neurofilament light (NFL) chain could be useful in diagnosing mild cognitive impairment (MCI) and Alzheimer’s disease (AD). This meta-analysis aimed to evaluate the circulating concentration of NFL in AD and MCI patients compared with healthy controls using the SIMOA technique.MethodsTo this end, Google Scholar, PubMed, Scopus, Web of Science, and the reference lists of relevant articles were systematically searched for studies reporting serum NFL chain levels in healthy controls, MCI, and AD patients. Appropriate statistical methods were employed to achieve the study purpose.ResultsFifteen eligible studies including 3086 patients were pooled out of a total of 347 publications. Fixed effect model analysis showed that NFL chain level was significantly higher in the serum of patients with MCI (0.361 SMD, 95% CI, 0.286–0.435, p = 0.000, I2 = 49.179) and AD (0.808 SMD, 95% CI, 0.727–0.888, p = 0.000, I2 = 39.433) compared with healthy individuals. The analysis also showed that the NFL chain levels in plasma were significantly different between patients with MCI and AD (0.436 SMD, 95% CI, 0.359–0.513, p = 0.000, I2 = 37.44). The overall heterogeneity of the studies was modest.ConclusionsThis study highlights the potential of serum NFL chain detected using SIMOA in differentiating MCI, AD, and healthy controls.

Pre-Analytical Variables Influencing Stability of Blood-Based Biomarkers of Neuropathology.

Sample collection and preanalytical protocols may significantly impact the results of large-scale epidemiological studies incorporating blood-based biomarkers of neuropathology. To evaluate the stability and assay variability of several blood-based biomarkers of neuropathology for common preanalytical conditions. We collected serum and plasma samples from 41 participants and evaluated the effect of processing delay of up to 72 h when stored at 4∘C, three freeze-thaw cycles, and a combination of 48-h processing delay when stored at 4∘C and three freeze-thaw cycles on biomarker stability. Using the Simoa assay (Quanterix Inc.), we measured amyloid-β 40 (Aβ40), amyloid-β 42 (Aβ42), neurofilament light (NfL), glial fibrillary acidic protein (GFAP), and phosphorylated tau 181 (p-tau-181). We found that Aβ40 and Aβ42 levels significantly decreased after a 24-h processing delay in both plasma and serum samples, and a single freeze-thaw cycle (p < 0.0001). Nevertheless, serum Aβ42/40 ratio remained stable with a processing delay up to 48 h while plasma Aβ42/40 ratio showed only small but significant increase with a delay up to 72 h. Both plasma and serum GFAP and NfL levels were only modestly affected by processing delay and freeze-thaw cycles. Plasma p-tau-181 levels notably increased with a 24-, 48-, and 72-h processing delay, but remained stable in serum. Intra-individual variation over two weeks was minimal for all biomarkers and their levels were substantially lower in serum when compared with plasma. These results suggest that standardizing preanalytical variables will allow robust measurements of biomarkers of neuropathology in population studies.

Plasma NfL as a prognostic biomarker for enriching HD-ISS stage 1 categorisation: a cross-sectional study

The recently proposed Huntington's Disease Integrated Staging System (HD-ISS) categorises individuals with the Huntintin genetic mutation into disease progression cohorts based on quantitative neuroimaging, cognitive, and functional markers for research purposes. Unfortunately, many research studies do not collect quantitative neuroimaging data, and so the authors of the HD-ISS have subsequently provided approximated cohort thresholds based on disease and clinical data alone. However, these are rough proxies that aim to maximise stage separation, and should not be considered as 1:1 substitutes for the HD-ISS. Notably, no wet biomarker met the stringent criteria required to be considered a landmark for HD-ISS categorisation. We have previously shown that levels of plasma neurofilament light (NfL), a neuronal marker associated with axonal injury, are associated with predicted years to clinical motor diagnosis (CMD). Our objective in the current study was to determine whether HD-ISS categorisation, particularly for stages prior to CMD, could be improved with consideration of plasma NfL levels.A total of 290 blood samples, and clinical measures, were collected from participants across all HD-ISS stages: n = 50 [Stage 0], n = 64 [Stage 1], n = 63 [Stage 2], n = 63 [Stage 3], as well as 50 healthy controls. Plasma NfL levels were measured using a Meso Scale Discovery assay.Cohorts differed by age, cognitive function, CAG repeat length, and select UHDRS measures. Plasma NfL levels also differed significantly across cohorts. Approximately 50% of Stage 1 participants had plasma NfL levels indicative of predicted CMD within ten years.Our findings suggest that plasma NfL levels may have use in enriching Stage 1 membership into sub-groups that are less than, and within, predicted 10 years until CMD.This work was supported by the National Institutes of Health (NS111655 to E.A.T.); the UCSD Huntington's Disease Society of America Center of Excellence; and the UCSD Shiley-Marcos Alzheimer's Disease Research Center (NIH-NIA P30 AG062429).

Neurofilament Light Chain as a Potential Biomarker in Plasma for Alzheimer's Disease and Mild Cognitive Impairment: A Systematic Review and a Meta-Analysis.

Plasma neurofilament light (NfL) is an intermediate filamentous protein involved in stabilizing axonal structure and promoting axon growth. Recent clinical studies have reported increased NfL levels in the plasma of Alzheimer's disease (AD) patients and patients with mild cognitive impairment (MCI). This study used meta-analysis to evaluate the potential of plasma NfL as a biomarker for patients with AD and MCI. PubMed, Embase, and Web of Science databases were systematically searched for studies of plasma NfL levels in AD and MCI, and a meta-analysis was employed to identify whether it was suited as a reliable biomarker and discrimination of healthy controls. A total of 24 published articles that included 2397 AD and 3242 MCI patients were analysed. The level of plasma NfL was significantly increased in patients with AD and MCI when compared with healthy control subjects (standard mean difference [SMD]: 14.33 [12.42-16.24], z = 14.71, p < 0.00001; SMD: 4.95 [3.82-6.80], z = 8.59, p < 0.00001) and higher in AD patients than MCI patients (SMD: 9.32 [8.07-10.57], z = 14.62, p < 0.00001). Meta-regression analysis showed a negative relationship between Mini-Mental State Examination (MMSE) scores and plasma NfL levels in MCI patients (slope = -0.399 [95% confidence interval (CI): -0.518 to -0.281], p < 0.05). The meta-analysis suggested that NfL levels increased in the plasma of patients with AD and MCI and were associated with cognitive decline. Results provide the clinical evidence to support plasma NfL as a cognitive biomarker for AD and MCI.

Sex differences in blood biomarkers and cognitive performance in individuals with autosomal dominant Alzheimer's disease.

Plasma tau phosphorylated at threonine 217 (P-tau217) and neurofilament light (NfL) have emerged as markers of Alzheimer's disease (AD) pathology. Few studies have examined the role of sex in plasma biomarkers in sporadic AD, yielding mixed findings, and none in autosomal dominant AD. We examined the effects of sex and age on plasma P-tau217 and NfL, and their association with cognitive performance in a cross-sectional study of 621 Presenilin-1 E280A mutation carriers (PSEN1) and non-carriers. As plasma P-tau217 levels increase, cognitively unimpaired female carriers showed better cognitive performance than cognitively unimpaired male carriers. Yet, as disease progresses, female carriers had a greater plasma NfL increase than male carriers. There were no sex differences in the association between age and plasma biomarkers among non-carriers. Our findings suggest that, among PSEN1 mutation carriers, females had a greater rate of neurodegeneration than males, yet it did not predict cognitive performance. We examined sex differences in plasma P-tau217 and NfL in Presenilin-1 E280A (PSEN1) mutation carriers and non-carriers. Female carriers had a greater plasma NfL increase, but not P-tau217, than male carriers. As plasma P-tau217 levels increase, cognitively unimpaired female carriers showed better cognitive performance than cognitively unimpaired male carriers. The interaction effect of sex by plasma NfL levels did not predict cognition among carriers.

Plasma Neurofilament Light (NfL) associated with retest learning effects on ambulatory cognitive assessments of processing speed: Results from the Einstein Aging Study (EAS)

AbstractBackgroundMeasurement burst designs, using ambulatory cognitive assessments facilitate the dissociation of retest effects from optimal (“asymptotic”) performance. Dissociating these features via cognitive process modeling may reveal indices of subtle cognitive impairment that signal risk for neurocognitive decline. We investigated cross‐sectional associations between plasma NfL (a biomarker of neurodegeneration), retest improvements, and optimal performance on the Symbol Match task, which assesses processing speed by response time (RT), in a racially/ethnically diverse community‐based EAS Study cohort.MethodAnalyses included 283 EAS participants free of dementia (Mean age = 77.4 years, SD = 4.8, Range = 70‐91; 67% Female; 46% Non‐Hispanic White, 40% Non‐Hispanic Black). Among them, 82 met the Jak/Bondi criteria of mild cognitive impairment (MCI). Participants completed a smartphone cognitive battery, resulting in up to 96 assessment sessions (six times per day for 16 days). Plasma NfL concentration was measured using a Single molecule array assay. A mixed‐effect negative exponential model was used to examine the associations between plasma NfL and indices of subtle cognitive impairment adjusting for covariates including age, gender, education, and race/ethnicity. Retest improvement was measured by the exponential decrease in RT across sessions and optimal performance was measured by asymptotic RT (fastest performance).ResultOn average, the RT of the Symbol Match task was 3.4 seconds (SD = 0.9). Mean plasma NfL level was 23.9 pg/mL (SD = 17.5). Higher plasma NfL levels were associated with a reduced retest improvement rate per session (estimate = ‐0.011, 95% confidence interval (‐0.013, ‐0.008) corresponding to one SD increase in plasma NfL, p&lt;0.001). No significant association between plasma NfL and optimal cognitive performance was found. Stratified analyses showed NfL’s association with retest improvement was larger in non‐Hispanic Whites than in non‐Hispanic Blacks (‐0.009 vs ‐0.002), although formal testing of the interaction between NfL and race was not significant. Results remained very similar excluding participants with MCI.ConclusionHigher plasma NfL was associated with a reduced retest improvement rate on a smartphone processing speed task, highlighting its potential value for identifying subtle cognitive impairment. Longitudinal data from larger samples, a broader array of biomarkers and of ambulatory cognitive tests, are required to fully explore these issues.

Association of Chronic Kidney Disease With Plasma NfL and Other Biomarkers of Neurodegeneration: The H70 Birth Cohort Study in Gothenburg.

Studies associate chronic kidney disease (CKD) with neurodegeneration. This study investigated the relationship between kidney function, blood, CSF, and structural brain MRI markers of neurodegeneration in a sample including individuals with and without CKD. Participants from the Gothenburg H70 Birth Cohort Study, with data on plasma neurofilament light (P-NfL), estimated glomerular filtration rate (eGFR), and structural brain MRI were included. Participants were invited to also have the CSF collected. The primary endpoint of this study was to determine any association between CKD and P-NfL. Secondary endpoints included cross-sectional associations between CKD, eGFR, and CSF-derived and MRI-derived markers of neurodegeneration and Alzheimer disease (AD) pathology (MRI: cortical thickness, hippocampal volume, lateral ventricle volume, and white matter lesion volume; CSF: β-amyloid (Aβ) 42, Aβ42/40, Aβ42/p-tau, t-tau, p-tau, and NfL). Participants with P-NfL and eGFR at baseline were re-examined on eGFR, 5.5 (5.3-6.1) years (median; IQR) after the first visit, and the predictive value of P-NfL levels on incident CKD was estimated longitudinally, using a Cox proportional hazards model. We included 744 participants, 668 without CKD (age 71 [70-71] years, 50% males) and 76 with CKD (age 71 [70-71] years, 39% males). Biomarkers from the CSF were analyzed in 313 participants. A total of 558 individuals returned for a re-examination of eGFR (75% response rate, age 76 [76; 77] years, 48% males, 76 new cases of CKD). Participants with CKD had higher P-NfL levels than those with normal kidney function (median; 18.8 vs 14.1 pg/mL, p < 0.001), while MRI and CSF markers were similar between the groups. P-NfL was independently associated with CKD after adjustment for confounding variables, including hypertension and diabetes (OR; 3.231, p < 0.001), in a logistic regression model. eGFR and CSF Aβ 42/40: R = 0.23, p = 0.004 correlated in participants with Aβ42 pathology. P-NfL levels in the highest quartile were associated with incident CKD at follow-up (HR; 2.39 [1.21: 4.72]). In a community-based cohort of 70-year olds, P-NfL was associated with both prevalent and incident CKD, while CSF and/or imaging measures did not differ by CKD status. Participants with CKD and dementia presented similar levels of P-NfL.

Plasma levels of phosphorylated tau and neurofilament light chain as potential biomarkers for Alzheimer’s disease: A biochemical analysis in Pakistani population

The National Institute on Aging-Alzheimer's Association's research framework in 2018 proposed a molecular construct for the diagnosis of Alzheimer’s disease (AD). Nonetheless, the clinical exclusionary strategy is still the mainstay of AD diagnosis in Pakistan. We looked at the plasma levels of amyloid beta-42 (Aβ-42), phosphorylated tau (P-tau), and neurofilament light (NFL) in patients with Alzheimer's clinical syndrome (ACS) and healthy controls (HC) from the Pakistani population to keep pace with the global efforts towards establishing accessible and affordable biochemical diagnostic markers for AD in Pakistan.Consultant neurologists screened patients who presented with cognitive impairment to three large tertiary care hospitals in Karachi, and after receiving informed consent, recruited participants with ACS and HC from the same facilities. We collected 5cc of blood in EDTA tubes along with demographic and lifestyle information of the subjects. Plasma aliquots were stored at −80°C after centrifugation. For analysis it was thawed at 4℃ and levels of the three proteins were measured through ELISA.Data from 28 ACS patients and 28 age matched healthy controls were evaluated. Among demographic factors, education and depression were related with health status (p = 0.03 and 0.003, respectively). NFL and P-tau mean values demonstrated a significant difference between the ACS and control groups (p = 0.003 and 0.006), however Aβ42 did not (p = 0.114). ROC analysis showed that plasma P-tau and NFL, with AUCs of 0.717 and 0.735, respectively, could substantially distinguish ACS from the HC group (p = 0.007 and 0.003, respectively). Both plasma P-tau (r = −0.389; p = 0.004) and NFL (r = −0.424; p = 0.001) levels were significantly and negatively correlated with individuals' MMSE scores. NFL and plasma P-tau show promise in differentiating AD patients from healthy individuals. However, similar larger studies are needed to validate our findings.

Using Plasma NfL and GFAP to Monitor Response to Anti-CD20 Treatment in MS (P1-3.010)

<h3>Objective:</h3> To compare changes in plasma concentrations of Neurofilament light (NfL) and Glial Fibrillary Acidic Protein (GFAP) in individuals with relapsing multiple sclerosis (RMS) vs progressive multiple sclerosis (PMS), treated with anti-CD20 immunotherapy (rituximab or ocrelizumab) <h3>Background:</h3> Blood biomarkers, especially NfL, can augment clinical and radiographic data in MS treatment monitoring. Few studies have investigated longitudinal change in NfL or GFAP in PMS patients taking anti-CD20s or compared changes in these biomarkers between RMS and PMS. <h3>Design/Methods:</h3> Subjects were selected by MS diagnosis, anti-CD20 treatment for at least 6 months, and presence of multiple blood samples in our Center’s Biorepository. Demographic/clinical information was extracted by chart review. SIMOA plasma assays of NfL and GFAP were conducted at baseline and follow-up (between 3 and 12 months) on Quanterix SR-X. Biomarker concentrations were log transformed. Summary statistics and longitudinal regression analyses, correcting for age, were generated. <h3>Results:</h3> 37 subjects and 67 samples were analyzed (n=7 had no follow-up samples between 3 and 12 months). 64.9% were female. RMS (n=19) had mean(SD) age 38.7(9.2). PMS (n=18) had mean age 56.1(10.4). Age-adjusted mean concentrations of NfL and GFAP did not differ significantly between RMS and PMS at baseline. Geometric mean NfL declined between baseline (6.8, 95%CI 4.2–11.0) and follow-up (4.9, 95%CI 3.3–7.3) in RMS but stayed stable in PMS. Geometric mean GFAP increased between baseline (52.4, 95%CI 35.2–77.9) and follow-up (60.8, 95%CI 40.4–91.4) in PMS but stayed stable in RMS. The changes for both NfL and GFAP differed statistically significantly between MS types. <h3>Conclusions:</h3> With anti-CD20 treatment, plasma NfL levels decreased in RMS by 27.6% while staying stable in PMS; GFAP levels increased in PMS by 16% while remaining stable in RMS. Results correlate with clinical evidence showing greater benefit of anti-CD20 immunotherapy in RMS compared to PMS. Future studies will include more subjects, longer observation, and different immunotherapies. <b>Disclosure:</b> Dr. Gross has received personal compensation in the range of $5,000-$9,999 for serving as an Expert Witness for AP Expert Group. The institution of Stefan Sillau has received research support from Alzheimer’s Association. The institution of Stefan Sillau has received research support from Hewitt Family Foundation; State of Colorado. The institution of Stefan Sillau has received research support from PCORI. The institution of Stefan Sillau has received research support from NINR. The institution of Stefan Sillau has received research support from Michael J. Fox Foundation. The institution of Stefan Sillau has received research support from Department of Defense. The institution of Stefan Sillau has received research support from Colorado Department of Public Health and Environment. The institution of Stefan Sillau has received research support from Benign Essential Blepharospasm Research Foundation. Stefan Sillau has a non-compensated relationship as a Statistician with Novartis that is relevant to AAN interests or activities. Stefan Sillau has a non-compensated relationship as a Statistician with Biogen that is relevant to AAN interests or activities. Sean Selva has nothing to disclose. Mrs. Ritchie has nothing to disclose.

Systematic assessment of plasma biomarkers in spinocerebellar ataxia

Background and objectivesPlasma neurofilament light (NfL), glial fibrillary acidic protein (GFAP), phosphorylated-tau (p-tau), and β-amyloid (Aβ) have emerged as promising markers in several neurodegenerative disorders, but whether they can be used as biomarkers in spinocerebellar ataxias (SCA) is yet to be determined. This study aimed to identify sensitive plasma markers for SCA and investigate their effectiveness in tracking ataxia severity, cognition, non-motor symptoms, and brain atrophy. MethodsThis observational study recruited consecutive participants from Huashan Hospital and the CABLE study from November 2019. Patients with SCA were genetically diagnosed, grouped according to the ataxia severity, and compared with healthy older individuals and patients with multiple system atrophy type C (MSA-C). Plasma NfL, GFAP, p-tau, and Aβ levels were measured by Simoa in all participants. Analysis of covariance, Spearman correlation, and multivariable regression were used to explore candidate markers in SCA. ResultsA total of 190 participants (60 SCA, 56 MSA-C, and 74 healthy controls) were enrolled. Plasma NfL level increased early in the pre-ataxic stage of SCA (32.23 ± 3.07 vs. 11.41 ± 6.62 pg/mL in controls), was positively associated with the ataxia severity (r = 0.45, P = 0.005) and CAG repeat length (r = 0.51, P = 0.001), varied among the different SCA subtypes (39.57 ± 13.50 pg/mL in SCA3, which was higher than 28.17 ± 8.02 pg/mL in SCA2, 17.08 ± 6.78 pg/mL in SCA8, and 24.44 ± 18.97 pg/mL in rare SCAs; P < 0.05), and was associated with brainstem atrophy. NfL alone (area under the curve [AUC] 0.867) or combined with p-tau181 and Aβ (AUC 0.929), showed excellent performance in discriminating SCA patients from controls. Plasma GFAP distinguished SCA from MSA-C with moderate accuracy (AUC > 0.700) and correlated with cognitive performance and cortical atrophy. Changes in levels of p-tau181 and Aβ were observed in SCA patients compared to controls. They were both correlated with cognition, while Aβ was also associated with non-motor symptoms, such as anxiety and depression. DiscussionPlasma NfL may serve as a sensitive biomarker for SCA, and its level is elevated in the pre-ataxic stage. The different performance of NfL and GFAP indicates differences in the underlying neuropathology of SCA and MSA-C. Moreover, amyloid markers may be useful for detecting memory dysfunction and other non-motor symptoms in SCA.

Evaluation of plasma levels of NFL, GFAP, UCHL1 and tau as Parkinson's disease biomarkers using multiplexed single molecule counting

Objective biomarkers for Parkinson’s Disease (PD) could aid early and specific diagnosis, effective monitoring of disease progression, and improved design and interpretation of clinical trials. Although alpha-synuclein remains a biomarker candidate of interest, the multifactorial and heterogenous nature of PD highlights the need for a PD biomarker panel. Ideal biomarker candidates include markers that are detectable in easily accessible samples, (ideally blood) and that reflect the underlying pathological process of PD. In the present study, we explored the diagnostic and prognostic PD biomarker potential of the SIMOA neurology 4-plex-A biomarker panel, which included neurofilament light (NFL), glial fibrillary acid protein (GFAP), tau and ubiquitin C-terminal hydrolase L1 (UCHL-1). We initially performed a serum vs plasma comparative study to determine the most suitable blood-based matrix for the measurement of these proteins in a multiplexed assay. The levels of NFL and GFAP in plasma and serum were highly correlated (Spearman rho-0.923, p < 0.0001 and rho = 0.825, p < 0.001 respectively). In contrast, the levels of tau were significantly higher in plasma compared to serum samples (p < 0.0001) with no correlation between sample type (Spearman p > 0.05). The neurology 4-plex-A panel, along with plasma alpha-synuclein was then assessed in a cross-sectional cohort of 29 PD patients and 30 controls. Plasma NFL levels positively correlated with both GFAP and alpha-synuclein levels (rho = 0.721, p < 0.0001 and rho = 0.390, p < 0.05 respectively). As diagnostic biomarkers, the control and PD groups did not differ in their mean NFL, GFAP, tau or UCHL-1 plasma levels (t test p > 0.05). As disease state biomarkers, motor severity (MDS-UPDRS III) correlated with increased NFL (rho = 0.646, p < 0.0001), GFAP (rho = 0.450, p < 0.05) and alpha-synuclein levels (rho = 0.406, p < 0.05), while motor stage (Hoehn and Yahr) correlated with increased NFL (rho = 0.455, p < 0.05) and GFAP (rho = 0.549, p < 0.01) but not alpha-synuclein levels (p > 0.05). In conclusion, plasma was determined to be most suitable blood-based matrix for multiplexing the neurology 4-plex-A panel. Given their correlation with motor features of PD, NFL and GFAP appear to be promising disease state biomarker candidates and further longitudinal validation of these two proteins as blood-based biomarkers for PD progression is warranted.

Effect of renal function on the diagnostic performance of plasma biomarkers for Alzheimer's disease.

Several blood-based biomarkers are promising to be used in the diagnosis of Alzheimer's disease (AD) including Aβ42/40, p-tau181, and neurofilament light (NfL). The kidney is associated with the clearance of proteins. It is crucial to evaluate the effect of renal function on the diagnostic performance of these biomarkers before clinical implementation, which is important for the establishment of reference ranges and the interpretation of results. This study is a cross-sectional analysis based on ADNI cohort. Renal function was determined by the estimated glomerular filtration rate (eGFR). Plasma Aβ42/40 was measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Plasma p-tau181 and NfL were analyzed by Single Molecule array (Simoa) technique. [18F] florbetapir-PET (Aβ-PET) was used as a reference standard to estimate the brain amyloid load. The cutoff of Aβ-PET positivity was defined as ≥1.11. Linear regression models were used to investigate the associations of continuous eGFR with each plasma biomarker separately. The diagnostic accuracies of plasma biomarkers for positive brain amyloid across different renal function groups were analyzed by Receiver operating characteristic (ROC) curve. Youden-Index was used to determine the cutoff levels. A total of 645 participants were included in this study. The levels and diagnostic performance of Aβ42/40 were not affected by renal function. eGFR was only found negatively associated with p-tau181 levels in Aβ-PET negetive sample (β = -0.09, p = 0.039). eGFR was found negatively associated with NfL levels both in whole sample and Aβ-PET stratified groups (β = -0.27, p < 0.001 in whole sample; β = -0.28, p = 0.004 in A-; β = -0.27, p < 0.001 in A+). The diagnostic accuracies of p-tau181 and NfL were not affected by renal function. But the cutoff values of p-tau181 and NfL changed in participants with mild to moderate eGFR decline compared to participants with normal eGFR. Plasma Aβ42/40 was a robust biomarker for AD which was not affected by renal function. Plasma p-tau181 and NfL levels were affected by renal function, specific reference values of them should be considered in populations with different renal function stages.

Acute Blood Levels of Neurofilament Light Indicate One-Year White Matter Pathology and Functional Impairment in Repetitive Mild Traumatic Brain Injured Mice.

Mild traumatic brain injury (mTBI) mostly causes transient symptoms, but repeated (r)mTBI can lead to neurodegenerative processes. Diagnostic tools to evaluate the presence of ongoing occult neuropathology are lacking. In a mouse model of rmTBI, we investigated MRI and plasma biomarkers of brain damage before chronic functional impairment arose. Anesthetized adult male and female C57BL/6J mice were subjected to rmTBI or a sham procedure. Sensorimotor deficits were evaluated up to 12 months post-injury in SNAP and Neuroscore tests. Cognitive function was assessed in the novel object recognition test at six and 12 months. Diffusion tensor imaging (DTI) and structural magnetic resonance imaging (MRI) were performed at six and 12 months to examine white matter and structural damage. Plasma levels of neurofilament light (NfL) were assessed longitudinally up to 12 months. Brain histopathology was performed at 12 months. Independent groups of mice were used to examine the effects of 2-, 7- and 14-days inter-injury intervals on acute plasma NfL levels and on hyperactivity. Twelve months after an acute transient impairment, sensorimotor functions declined again in rmTBI mice (p < 0.001 vs sham), but not earlier. Similarly, rmTBI mice showed memory impairment at 12 (p < 0.01 vs sham) but not at 6 months. White matter damage examined by DTI was evident in rmTBI mice at both six and 12 months (p < 0.001 vs sham). This was associated with callosal atrophy (p < 0.001 vs sham) evaluated by structural MRI. Plasma NfL at one week was elevated in rmTBI (p < 0.001 vs sham), and its level correlated with callosal atrophy at 12 months (Pearson r = 0.72, p < 0.01). Histopathology showed thinning of the corpus callosum and marked astrogliosis in rmTBI mice. The NfL levels were higher in mice subjected to short (2 days) compared with longer (7 and 14 days) inter-injury intervals (p < 0.05), and this correlated with hyperactivity in mice (Pearson r = 0.50; p < 0.05). These findings show that rmTBI causes white matter pathology detectable by MRI before chronic functional impairment. Early quantification of plasma NfL correlates with the degree of white matter atrophy one year after rmTBI and can serve to monitor the brain's susceptibility to a second mTBI, supporting its potential clinical application to guide the return to practice in sport-related TBI.

Longitudinal decline of plasma neurofilament light levels after antiretroviral initiation in people living with HIV.

This retrospective follow-up study aims to investigate the dynamic longitudinal change of plasma neurofilament light (NfL) levels after antiretroviral therapy (ART) initiation in a cohort of people living with human immunodeficiency virus (HIV) (PWH). We tested a convenience sample of 116 patients from the NORTHIV study. Plasma NfL levels-measured using Single molecule array (Simoa) technology-as well as other laboratory parameters were collected at baseline, weeks 4, 48, 96, and 144. Linear mixed-effects models were estimated to evaluate longitudinal change over time. Baseline CD4+ T-cell levels, CDC classification, and HIV RNA levels were considered. Models were adjusted by age, sex, treatment regimen, and baseline serum creatinine levels. Plasma NfL levels were higher at baseline and also declined faster during the follow-up for participants with CD4+ count <100 cells/µl compared with >100 cells/µl. No significant difference was found between the CD4+ strata 100-199 and 200-499/µl. Participants with CDC classification stages B and C had higher levels of plasma NfL at baseline, as well as faster decline compared with participants with stage A. No significant main effects or change over time was found in baseline HIV RNA levels, treatment regimen, or sex. Plasma NfL is a sensitive biomarker to assess ongoing central nervous system injury in PWH. Plasma NfL concentrations decline relatively fast following ART initiation and then stabilize after 48 weeks. Plasma NfL concentrations are associated with CD4+ count and stage of HIV disease. No correlations were seen with different ART regimens.