Use of dried blood spots for monitoring inflammatory and nutritional biomarkers in the elderly.

OBJECTIVES: Blood microsampling, particularly dried blood spots (DBSs), is an attractive minimally-invasive approach that is well suited for home sampling and predictive medicine associated with longitudinal follow-up of the elderly. However, in vitro diagnostic quantification of biomarkers from DBS poses a major challenge. Clinical mass spectrometry can reliably quantify blood proteins in various research projects. Our goal here was to use mass spectrometry of DBS in a real-world clinical setting and compared it to the standard immunoassay method. We also sought to correlate DBS mass spectrometry measurements with clinical indices. METHODS: A clinical trial of diagnostic equivalence was conducted to compare conventional venous samples quantified by immunoassay and DBSs quantified by mass spectrometry in an elderly population. We assayed three protein biomarkers of nutritional and inflammatory status: prealbumin (transthyretin), C-reactive protein, and transferrin. RESULTS: The analysis of DBSs showed satisfactory variability and low detection limits. Statistical analysis confirmed that the two methods give comparable results at clinical levels of accuracy. In conclusion, we demonstrated, in a real-life setting, that DBSs can be used to measure prealbumin, CRP and transferrin, which are commonly used markers of nutritional status and inflammation in the elderly. However, there was no correlation with patient frailty for these proteins. CONCLUSIONS: Early detection and regular monitoring of nutritional and inflammatory problems using DBS appear to be clinically feasible. This could help resolve major public health challenges in the elderly for whom frailty leads to serious risks of health complications.

Diagnostic delay of multiple sclerosis: prevalence, determinants and consequences.

BACKGROUND: Early diagnosis and treatment of patients with multiple sclerosis (MS) are associated with better outcomes; however, diagnostic delays remain a major problem. OBJECTIVE: Describe the prevalence, determinants and consequences of delayed diagnoses. METHODS: This single-centre ambispective study analysed 146 adult relapsing-remitting MS patients (2016-2021) for frequency and determinants of diagnostic delays and their associations with clinical, cognitive, imaging and biochemical measures. RESULTS: Diagnostic delays were identified in 77 patients (52.7%), including 42 (28.7%) physician-dependent cases and 35 (24.0%) patient-dependent cases. Diagnosis was delayed in 22 (15.1%) patients because of misdiagnosis by a neurologist. A longer diagnostic delay was associated with trends towards greater Expanded Disability Status Scale (EDSS) scores (B = 0.03; p = 0.034) and greater z-score of the blood neurofilament light chain (B = 0.35; p = 0.031) at the time of diagnosis. Compared with patients diagnosed at their first clinical relapse, patients with a history of >1 relapse at diagnosis (n = 63; 43.2%) had a trend towards greater EDSS scores (B = 0.06; p = 0.006) and number of total (B = 0.13; p = 0.040) and periventricular (B = 0.06; p = 0.039) brain lesions. CONCLUSION: Diagnostic delays in MS are common, often determined by early misdiagnosis and associated with greater disease burden.

Optical coherence tomography versus other biomarkers: Associations with physical and cognitive disability in multiple sclerosis.

BACKGROUND: Optical coherence tomography (OCT) is a biomarker of neuroaxonal loss in multiple sclerosis (MS). OBJECTIVE: The objective was to assess the relative role of OCT, next to magnetic resonance imaging (MRI) and serum markers of disability in MS. METHODS: A total of 100 patients and 52 controls underwent OCT to determine peripapillary retinal nerve fiber layer (pRNFL) and ganglion cell-inner plexiform layers (GCIPL). Serum neurofilament light chain (sNfL), total lesion volume (TLV), and brain parenchymal fraction (BPF) were also assessed. The associations of OCT with disability were examined in linear regression models with correction for age, vision, and education. RESULTS: In patients, pRNFL was associated with the Symbol Digit Modalities Test (SDMT; p = 0.030). In the multivariate analysis including sNfL and MRI measures, pRNFL (ß = 0.19, p = 0.044) and TLV (ß = -0.24, p = 0.023) were the only markers associated with the SDMT. pRNFL (p < 0.001) and GCIPL (p < 0.001) showed associations with the Expanded Disability Status Scale (EDSS). In the multivariate analysis, GCIPL showed the strongest association with the EDSS (ß = -0.32, p < 0.001) followed by sNfL (ß = 0.18, p = 0.024). CONCLUSION: The associations of OCT measures with cognitive and physical disability were independent of serum and brain MRI markers of neuroaxonal loss. OCT can be an important tool for stratification in MS, while longitudinal studies using combinations of biomarkers are warranted.

Serum tau protein elevation in migraine: a cross-sectional case-control study.

BACKGROUND: Migraine is a disorder associated with neuropeptide release, pain and inflammation. Tau protein has recently been linked to inflammatory diseases and can be influenced by neuropeptides such as CGRP, a key neurotransmitter in migraine. Here, we report serum concentrations of total-tau protein in migraine patients and healthy controls. METHODS: In this cross-sectional study, interictal blood samples from n = 92 patients with episodic migraine (EM), n = 93 patients with chronic migraine (CM), and n = 42 healthy matched controls (HC) were studied. We assessed serum total-tau protein (t-tau) and for comparison neurofilament light chain protein (NfL), glial fibrillary acidic protein (GFAP), and ubiquitin carboxy-terminal hydrolase L (UCH-L1) concentrations using the Neurology 4-plex kit, on a single molecule array HD-X Analyzer (Quanterix Corp Lexington, MA). Matched serum/cerebrospinal fluid (CSF) samples were used for post-hoc evaluations of a central nervous system (CNS) source of relevant findings. We applied non-parametric tests to compare groups and assess correlations. RESULTS: Serum t-tau concentrations were elevated in EM [0.320 (0.204 to 0.466) pg/mL] and CM [0.304 (0.158 to 0.406) pg/mL] patients compared to HC [0.200 (0.114 to 0.288) pg/mL] (p = 0.002 vs. EM; p = 0.025 vs. CM). EM with aura [0.291 (0.184 to 0.486 pg/mL); p = 0.013] and EM without aura [0.332 (0.234 to 0.449) pg/mL; p = 0.008] patients had higher t-tau levels than HC but did not differ between each other. Subgroup analysis of CM with/without preventive treatment revealed elevated t-tau levels compared to HC only in the non-prevention group [0.322 (0.181 to 0.463) pg/mL; p = 0.009]. T-tau was elevated in serum (p = 0.028) but not in cerebrospinal fluid (p = 0.760). In contrast to t-tau, all proteins associated with cell damage (NfL, GFAP, and UCH-L1), did not differ between groups. DISCUSSION: Migraine is associated with t-tau elevation in serum but not in the CSF. Our clinical study identifies t-tau as a new target for migraine research.

New Perspectives of Multiplex Mass Spectrometry Blood Protein Quantification on Microsamples in Biological Monitoring of Elderly Patients.

Blood microsampling combined with large panels of clinically relevant tests are of major interest for the development of home sampling and predictive medicine. The aim of the study was to demonstrate the practicality and medical utility of microsamples quantification using mass spectrometry (MS) in a clinical setting by comparing two types of microsamples for multiplex MS protein detection. In a clinical trial based on elderly population, we compared 2 µL of plasma to dried blood spot (DBS) with a clinical quantitative multiplex MS approach. The analysis of the microsamples allowed the quantification of 62 proteins with satisfactory analytical performances. A total of 48 proteins were significantly correlated between microsampling plasma and DBS (p < 0.0001). The quantification of 62 blood proteins allowed us to stratify patients according to their pathophysiological status. Apolipoproteins D and E were the best biomarker link to IADL (instrumental activities of daily living) score in microsampling plasma as well as in DBS. It is, thus, possible to detect multiple blood proteins from micro-samples in compliance with clinical requirements and this allows, for example, to monitor the nutritional or inflammatory status of patients. The implementation of this type of analysis opens new perspectives in the field of diagnosis, monitoring and risk assessment for personalized medicine approaches.

High serum neurofilament light chain levels correlate with brain atrophy and physical disability in multiple sclerosis.

BACKGROUND AND PURPOSE: Serum neurofilament light chain (sNfL) is a promising biomarker of neuroaxonal damage in persons with multiple sclerosis (pwMS). In cross-sectional studies, sNfL has been associated with disease activity and brain magnetic resonance imaging (MRI) changes; however, it is still unclear to what extent in particular high sNfL levels impact on subsequent disease evolution. METHODS: sNfL was quantified by an ultrasensitive single molecule array (Simoa) in 199 pwMS (median age = 34.2 years, 64.3% female) and 49 controls. All pwMS underwent 3-T MRI to assess global and compartmental normalized brain volumes, T2-lesion load, and cortical mean thickness. Follow-up data and serum samples were available in 144 pwMS (median follow-up time = 3.8 years). Linear and binary logistic models were used to estimate the independent contribution of sNfL for changes in MRI and Expanded Disability Status Scale (EDSS). Age-corrected sNfL z-scores from a normative database of healthy controls were used for sensitivity analyses. RESULTS: High sNfL levels at baseline were associated with atrophy measures of the whole brain (standardized beta coefficient ßj = -0.352, p < 0.001), white matter (ßj = -0.229, p = 0.007), thalamus (ßj = -0.372, p = 0.004), and putamen (ßj = -1.687, p = 0.012). pwMS with high levels of sNfL at baseline and follow-up had a greater risk of EDSS worsening (p = 0.007). CONCLUSIONS: Already single time point elevation of sNfL has a distinct effect on brain volume changes over a short-term period, and repeated high levels of sNfL indicate accumulating physical disability. Serial assessment of sNfL may provide added value in the clinical management of pwMS.

GnRH replacement rescues cognition in Down syndrome.

At the present time, no viable treatment exists for cognitive and olfactory deficits in Down syndrome (DS). We show in a DS model (Ts65Dn mice) that these progressive nonreproductive neurological symptoms closely parallel a postpubertal decrease in hypothalamic as well as extrahypothalamic expression of a master molecule that controls reproduction-gonadotropin-releasing hormone (GnRH)-and appear related to an imbalance in a microRNA-gene network known to regulate GnRH neuron maturation together with altered hippocampal synaptic transmission. Epigenetic, cellular, chemogenetic, and pharmacological interventions that restore physiological GnRH levels abolish olfactory and cognitive defects in Ts65Dn mice, whereas pulsatile GnRH therapy improves cognition and brain connectivity in adult DS patients. GnRH thus plays a crucial role in olfaction and cognition, and pulsatile GnRH therapy holds promise to improve cognitive deficits in DS.

Serum Neurofilament Light Chain Levels in the Intensive Care Unit: Comparison between Severely Ill Patients with and without Coronavirus Disease 2019.

There is emerging evidence for multifarious neurological manifestations of coronavirus disease 2019 (COVID-19), but little is known regarding whether they reflect structural damage to the nervous system. Serum neurofilament light chain (sNfL) is a specific biomarker of neuronal injury. We measured sNfL concentrations of 29 critically ill COVID-19 patients, 10 critically ill non-COVID-19 patients, and 259 healthy controls. After adjusting for neurological comorbidities and age, sNfL concentrations were higher in patients with COVID-19 versus both comparator groups. Higher sNfL levels were associated with unfavorable short-term outcome, indicating that neuronal injury is common and pronounced in critically ill patients. ANN NEUROL 2021;89:610-616.

Neuroprotective associations of apolipoproteins A-I and A-II with neurofilament levels in early multiple sclerosis.

BACKGROUND: The role of cholesterol homeostasis in neuroaxonal injury in multiple sclerosis is not known. OBJECTIVE: The objective of the study is to investigate the associations of cerebrospinal fluid (CSF) and serum neurofilament light chain levels (CSF-NfL and sNfL, respectively), which are biomarkers of neuroaxonal injury, with cholesterol biomarkers at the clinical onset of multiple sclerosis. METHODS: sNfL, serum cholesterol profile (total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol), serum apolipoprotein (Apo) levels (ApoA-I, ApoA-II, ApoB, and ApoE), and albumin quotient were obtained for 133 patients (63% female, age: 29.9 ± 8.0 years) during the first demyelinating event. CSF-NfL was available for 103 (77%) patients. RESULTS: CSF-NfL and sNfL were negatively associated with serum ApoA-II (P = .005, P < .001) and positively associated with albumin quotient (P < .001, P < .0001). In addition, higher CSF-NfL was associated with lower serum ApoA-I (P = .009) levels and higher sNfL was associated with lower high-density lipoprotein cholesterol (P = .010). In stepwise regression, age (P = .045), serum ApoA-II (P = .022), and albumin quotient (P < .001) were associated with CSF-NfL; albumin quotient (P = .002) and ApoA-II (P = .001) were associated with sNfL. Path analysis identified parallel pathways from ApoA-II (P = .009) and albumin quotient (P < .001) to the sNfL outcome that were mediated by CSF-NfL (P < .001). The associations of CSF-NfL with ApoA-I (P = .014) and ApoA-II (P = .015) and sNfL with ApoA-II (P < .001) remained significant after adjusting for number of contrast-enhancing lesions and T2 lesion volume. CONCLUSION: Lower serum ApoA-II and ApoA-I levels are associated with greater neuroaxonal injury as measured by CSF-NfL.

Plasma and CSF biomarkers for the diagnosis of Alzheimer's disease in adults with Down syndrome: a cross-sectional study.

BACKGROUND: Diagnosis of Alzheimer's disease in Down syndrome is challenging because of the absence of validated diagnostic biomarkers. We investigated the diagnostic performance of plasma and CSF biomarkers in this population. METHODS: We did a cross-sectional study of adults aged 18 years and older with Down syndrome enrolled in a population-based health plan in Catalonia, Spain. Every person with Down syndrome assessed in the health plan was eligible to enter the Down Alzheimer Barcelona Neuroimaging Initiative, and those with a plasma or CSF sample available were included in this study. Participants underwent neurological and neuropsychological examination and blood sampling, and a subset underwent a lumbar puncture. Adults with Down syndrome were classified into asymptomatic, prodromal Alzheimer's disease, or Alzheimer's disease dementia groups by investigators masked to biomarker data. Non-trisomic controls were a convenience sample of young (23-58 years) healthy people from the Sant Pau Initiative on Neurodegeneration. Amyloid-ß (Aß)1-40, Aß1-42, total tau (t-tau), 181-phosphorylated tau (p-tau; only in CSF), and neurofilament light protein (NfL) concentrations were measured in plasma with a single molecule array assay and in CSF with ELISA. Plasma and CSF biomarker concentrations were compared between controls and the Down syndrome clinical groups. Diagnostic performance was assessed with receiver operating characteristic curve analyses between asymptomatic participants and those with prodromal Alzheimer's disease and between asymptomatic participants and those with Alzheimer's disease dementia. FINDINGS: Between Feb 1, 2013, and Nov 30, 2017, we collected plasma from 282 participants with Down syndrome (194 asymptomatic, 39 prodromal Alzheimer's disease, 49 Alzheimer's disease dementia) and 67 controls; CSF data were available from 94 participants (54, 18, and 22, respectively) and all 67 controls. The diagnostic performance of plasma biomarkers was poor (area under the curve [AUC] between 0·53 [95% CI 0·44-0·62] and 0·74 [0·66-0·82]) except for plasma NfL concentrations, which had an AUC of 0·88 (0·82-0·93) for the differentiation of the asymptomatic group versus the prodromal Alzheimer's disease group and 0·95 (0·92-0·98) for the asymptomatic group versus the Alzheimer's disease dementia group. In CSF, except for Aß1-40 concentrations (AUC 0·60, 95% CI 0·45-0·75), all biomarkers had a good performance in the asymptomatic versus prodromal Alzheimer's disease comparison: AUC 0·92 (95% CI 0·85-0·99) for Aß1-42, 0·81 (0·69-0·94) for t-tau, 0·80 (0·67-0·93) for p-tau, and 0·88 (0·79-0·96) for NfL. Performance of the CSF biomarkers was optimal in the asymptomatic versus Alzheimer's disease dementia comparison (AUC ≥0·90 for all except Aß1-40 [0·59, 0·45-0·72]). Only NfL concentrations showed a strong correlation between plasma and CSF biomarker concentrations in participants with Down syndrome (rho=0·80; p<0·0001). INTERPRETATION: Plasma NfL and CSF biomarkers have good diagnostic performance to detect Alzheimer's disease in adults with Down syndrome. Our findings support the utility of plasma NfL for the early detection of Alzheimer's disease in Down syndrome in clinical practice and clinical trials. FUNDING: Institute of Health Carlos III, Fundació La Marató de TV3, Fundació Bancaria Obra Social La Caixa, Fundació Catalana Síndrome de Down, and Fundació Víctor Grífols i Lucas.