Successful MRI-Guided Focused Ultrasound Thalamotomy after Ipsilateral Gamma Knife Radiosurgery for Essential Tremor: A Case Report with Video.

We report the case of a 67-year-old left-handed female patient with disabling medically refractory essential tremor who underwent successful right-sided magnetic resonance-guided focused ultrasound (MRgFUS) of the ventral intermediate nucleus after ipsilateral gamma knife radiosurgery (GKRS) thalamotomy performed 3 years earlier. The GKRS had a partial effect on her postural tremor without side effects, but there was no reduction of her kinetic tremor or improvement in her quality of life (QoL). The patient subsequently underwent a MRgFUS thalamotomy, which induced an immediate and marked reduction in both the postural and kinetic tremor components, with minor complications (left upper lip hypesthesia, dysmetria in her left hand, and slight gait ataxia). The MRgFUS-induced lesion was centered more medially than the GKRS-induced lesion and extended more posteriorly and inferiorly. The MRgFUS-induced lesion interrupted remaining fibers of the dentatorubrothalamic tract (DRTT). The functional improvement 1-year post-MRgFUS was significant due to a marked reduction of the patient's kinetic tremor. The QoL score (Quality of Life in Essential Tremor) improved by 88% and her Clinical Rating Scale for Tremor left hand score by 62%. The side effects persisted but were minor, with no impact on her QoL. The explanation for the superior efficacy of MRgFUS compared to GKRS in our patient could be due to either a poor response to the GKRS or to a better localization of the MRgFUS lesion with a more extensive interruption of DRTT fibers. In conclusion, MRgFUS can be a valuable therapeutic option after unsatisfactory GKRS, especially because MRgFUS has immediate clinical effectiveness, allowing intra-procedural test lesions and possible readjustment of the target if necessary.

Electrophysiological confrontation of Lead-DBS-based electrode localizations in patients with Parkinson's disease undergoing deep brain stimulation.

Microelectrode recordings (MERs) are often used during deep brain stimulation (DBS) surgeries to confirm the position of electrodes in patients with advanced Parkinson's disease. The present study focused on 32 patients who had undergone DBS surgery for advanced Parkinson's disease. The first objective was to confront the anatomical locations of intraoperative individual MERs as determined electrophysiologically with those determined postoperatively by image reconstructions. The second aim was to search for differences in cell characteristics among the three subthalamic nucleus (STN) subdivisions and between the STN and other identified subcortical structures. Using the DISTAL atlas implemented in the Lead-DBS image reconstruction toolbox, each MER location was determined postoperatively and attributed to specific anatomical structures (sensorimotor, associative or limbic STN; substantia nigra [SN], thalamus, nucleus reticularis polaris, zona incerta [ZI]). The STN dorsal borders determined intraoperatively from electrophysiology were then compared with the STN dorsal borders determined by the reconstructed images. Parameters of spike clusters (firing rates, amplitudes - with minimum amplitude of 60 µV -, spike durations, amplitude spectral density of ß-oscillations) were compared between structures (ANOVAs on ranks). Two hundred and thirty one MERs were analyzed (144 in 34 STNs, 7 in 4 thalami, 5 in 4 ZIs, 34 in 10 SNs, 41 others). The average difference in depth of the electrophysiological dorsal STN entry in comparison with the STN entry obtained with Lead-DBS was found to be of 0.1 mm (standard deviation: 0.8 mm). All 12 analyzed MERs recorded above the electrophysiologically-determined STN entry were confirmed to be in the thalamus or zona incerta. All MERs electrophysiologically attributed to the SN were confirmed to belong to this nucleus. However, 6/34 MERs that were electrophysiologically attributed to the ventral STN were postoperatively reattributed to the SN. Furthermore, 44 MERs of 3 trajectories, which were intraoperatively attributed to the STN, were postoperatively reattributed to the pallidum or thalamus. MER parameters seemed to differ across the STN, with higher spike amplitudes (H = 10.64, p < 0.01) and less prevalent ß-oscillations (H = 9.81, p < 0.01) in the limbic STN than in the sensorimotor and associative subdivisions. Some cells, especially in the SN, showed longer spikes with lower firing rates, in agreement with described characteristics of dopamine cells. However, these probabilistic electrophysiological signatures might become clinically less relevant with the development of image reconstruction tools, which deserve to be applied intraoperatively.

Assessing preoperative hope and expectations related to functional neurosurgery: a new questionnaire.

BACKGROUND: Despite successful functional neurosurgery, patients suffering from epilepsy or Parkinson's disease may experience postoperative psychological distress and social maladjustments. Difficulties in coping with postoperative changes, even positive ones, have shown to be related to patients' presurgery cognitive representations (i.e., expectations, hope). The aim of this study was to develop an instrument assessing various key features of surgery outcomes' representations, namely the Preoperative Hope and Expectations Questionnaire (PHEQ). METHODS: Participants were patients (n = 50) diagnosed with Parkinson's disease (n = 25) or epilepsy (n = 25), candidates for functional neurosurgery (i.e., Deep brain stimulation, anterior temporal lobectomy). Two to three weeks before the planned surgery, they were administrated items assessing their actual state, preoperative expectations, and hope regarding surgery outcomes. They also completed measures assessing optimism, quality of life and mood. RESULTS: Exploratory analysis resulted in a 14-item version of the PHEQ composed of two factors (abstract representations, including psychological well-being and concrete representations, such as direct surgery outcomes). The PHEQ demonstrated high internal consistency and good convergent validity. Patients were more prone to express postoperative improvements in terms of hope rather than expectations. They generally focused on concrete rather than abstract features, although patients with Parkinson's disease had higher abstract future-oriented representations. CONCLUSIONS: The PHEQ presents satisfactory psychometric properties and may be considered as a reliable instrument for research and clinical practice.

Childhood-Onset Movement Disorders Can Mask a Primary Immunodeficiency: 6 Cases of Classical Ataxia-Telangiectasia and Variant Forms.

Ataxia-telangiectasia (A-T) is a neurodegenerative and primary immunodeficiency disorder (PID) characterized by cerebellar ataxia, oculocutaneous telangiectasia, immunodeficiency, progressive respiratory failure, and an increased risk of malignancies. It demands specialized care tailored to the individual patient's needs. Besides the classical ataxia-telangiectasia (classical A-T) phenotype, a variant phenotype (variant A-T) exists with partly overlapping but some distinctive disease characteristics. Here we present a case series of 6 patients with classical A-T and variant A-T, which illustrates the phenotypic variability of A-T that can present in childhood with prominent extrapyramidal features, with or without cerebellar ataxia. We report the clinical data, together with a detailed genotype description, immunological analyses, and related expression of the ATM protein. We show that the presence of some residual ATM kinase activity leads to the clinical phenotype variant A-T that differs from the classical A-T. Our data illustrate that the diagnosis of the variant form of A-T can be delayed and difficult, while early recognition of the variant form as well as the classical A-T is a prerequisite for providing a correct prognosis and appropriate rehabilitation and support, including the avoidance of diagnostic X-ray procedures, given the increased risk of malignancies and the higher risk for side effects of subsequent cancer treatment.

Multiomic Analyses of Dopaminergic Neurons Isolated from Human Substantia Nigra in Parkinson's Disease: A Descriptive and Exploratory Study.

Dopaminergic neurons (DA) of the substantia nigra pars compacta (SNpc) selectively and progressively degenerate in Parkinson's disease (PD). Until now, molecular analyses of DA in PD have been limited to genomic or transcriptomic approaches, whereas, to the best of our knowledge, no proteomic or combined multiomic study examining the protein profile of these neurons is currently available. In this exploratory study, we used laser capture microdissection to extract regions from DA in 10 human SNpc obtained at autopsy in PD patients and control subjects. Extracted RNA and proteins were identified by RNA sequencing and nanoliquid chromatography-mass spectrometry, respectively, and the differential expression between PD and control group was assessed. Qualitative analyses confirmed that the microdissection protocol preserves the integrity of our samples and offers access to specific molecular pathways. This multiomic analysis highlighted differential expression of 52 genes and 33 proteins, including molecules of interest already known to be dysregulated in PD, such as LRP2, PNMT, CXCR4, MAOA and CBLN1 genes, or the Aldehyde dehydrogenase 1 protein. On the other hand, despite the same samples were used for both analyses, correlation between RNA and protein expression was low, as exemplified by the CST3 gene encoding for the cystatin C protein. This is the first exploratory study analyzing both gene and protein expression of laser-dissected neuronal parts from SNpc in PD. Data are available via ProteomeXchange with identifier PXD024748 and via GEO with identifier GSE 169755.

Oral Dysbiosis and Inflammation in Parkinson's Disease.

BACKGROUND: Oral microbiota has largely escaped attention in Parkinson's disease (PD), despite its pivotal role in maintaining oral and systemic health. OBJECTIVE: The aim of our study was to examine the composition of the oral microbiota and the degree of oral inflammation in PD. METHODS: Twenty PD patients were compared to 20 healthy controls. Neurological, periodontal and dental examinations were performed as well as dental scaling and gingival crevicular fluid sampling for cytokines measurement (interleukine (IL)-1ß, IL-6, IL-1 receptor antagonist (RA), interferon-γ and tumor necrosis factor (TNF)-α). Two months later, oral microbiota was sampled from saliva and subgingival dental plaque. A 16S rRNA gene amplicon sequencing was used to assess bacterial communities. RESULTS: PD patients were in the early and mid-stage phases of their disease (Hoehn & Yahr 2-2.5). Dental and periodontal parameters did not differ between groups. The levels of IL-1ß and IL-1RA were significantly increased in patients compared to controls with a trend for an increased level of TNF-α in patients. Both saliva and subgingival dental plaque microbiota differed between patients and controls. Streptococcus mutans, Kingella oralis, Actinomyces AFQC_s, Veillonella AFUJ_s, Scardovia, Lactobacillaceae, Negativicutes and Firmicutes were more abundant in patients, whereas Treponema KE332528_s, Lachnospiraceae AM420052_s, and phylum SR1 were less abundant. CONCLUSION: Our findings show that the oral microbiome is altered in early and mid-stage PD. Although PD patients had good dental and periodontal status, local inflammation was already present in the oral cavity. The relationship between oral dysbiosis, inflammation and the pathogenesis of PD requires further study.

Illness Representations and Coping Strategies in Patients Treated with Deep Brain Stimulation for Parkinson's Disease.

There is a debate on possible alterations of self-identity following deep brain stimulation for neurological disorders including Parkinson's disease. Among the psychological variables likely to undergo changes throughout such a medical procedure, illness representations and coping strategies have not been the target of much research to this day. In order to remedy this, we investigated the dynamics of illness representations and coping strategies in an 18-month longitudinal study involving 45 patients undergoing deep brain stimulation for idiopathic Parkinson's disease. Two research hypotheses were formulated and investigated through repeated measures of ANOVAs and structural equation modelling with full information maximum likelihood and Bayesian estimations. Representations of Parkinson's disease as a cyclical condition and perception of control over the disease diminished after surgery. Use of instrumental coping strategies was not modified after deep brain stimulation. These changes were identified by SEM but not ANOVAs; their magnitude was nevertheless relatively small, implying general stability in representations. These findings suggest that psychological variables do not undergo major changes after deep brain stimulation for Parkinson's disease.

What Is the Best Electrophysiologic Marker of the Outcome of Subthalamic Nucleus Stimulation in Parkinson Disease?

OBJECTIVE: Deep brain stimulation of the subthalamic nucleus (STN) is advocated in patients with advanced Parkinson disease. Intraoperative microelectrode recordings (MER) and stimulation or imaging are applied to confirm electrode targeting. The study objective was to evaluate which intraoperative electrophysiologic marker, MER, stimulation, or local field potentials (LFP) was the most predictive of the clinical efficacy. METHODS: Efficacy was determined with lateralized motor scores of Movement Disorders Society-Unified Parkinson's Disease Rating Scale in 36 patients (OFF-drug/ON-stimulation 1 year after surgery vs. OFF-drug before surgery). Trajectory lengths in STN were determined from MER. Stimulation was increased up to the thresholds of first decrease, of complete suppression of rigidity, and of excitation of pyramidal motor tract. ß oscillations (11-31 Hz) were computed from LFP of the electrode macrocontact. Univariate and multivariate analyses were computed. RESULTS: Motor improvements were linked to trajectory lengths in STN (R2 = 0.17; P > 0.005). No significant relationship was found for thresholds of first decrease or suppression in rigidity or for motor tract excitation (R2 < 0.03, P > 0.05). Motor improvements were most linked to ß oscillation increases (R2 = 0.57, P < 0.005, linear regression; R2 = 0.84, P < 0.0001, post hoc sigmoid regression). ß oscillations appeared more predictive than length (ß: t = 5.4, P < 0.001; length: t = 2.70, P < 0.03). Improvements were also slightly predicted by preoperative scores (R2 = 0.13; P < 0.005). CONCLUSIONS: Motor improvements emerged as most related to ß oscillations, before trajectory length within the STN, whereas stimulation thresholds of rigidity or of motor tract excitation failed to show any relationship. The study encourages LFP measurement to confirm STN electrode location.

Deep brain stimulation-associated brain tissue imprints: a new in vivo approach to biological research in human Parkinson's disease.

BACKGROUND: Deep brain stimulation (DBS) of the subthalamic nucleus (STN) or the internal segment of the globus pallidus (GPi) has been established as a highly effective symptomatic therapy for Parkinson's disease (PD). An intriguing biological aspect related to the DBS procedure is that a temporary contact establishes between surgical instruments and the surrounding brain tissue. In this exploratory study, we took advantage of this unique context to harvest brain material adhering to the stylet routinely used during surgery, and to examine the biological value of these samples, here referred to as "brain tissue imprints" (BTIs). RESULTS: Nineteen BTIs from 12 STN- or GPi-electrode implanted patients were obtained in vivo during DBS surgery, without any modification of the surgical procedure. Immunofluorescence analyses confirmed that our approach allowed the harvesting of many neural cells including neurons harboring distinct neurotransmitter markers. Shotgun proteomic and transcriptomic analyses provided for the first time molecular information from DBS-associated brain samples, and confirmed the compatibility of this new type of sample with poly-omic approaches. The method appears to be safe and results consistent. CONCLUSIONS: We here propose BTIs as original and highly valuable brain samples, and DBS-related brain imprinting as a new conceptual approach to biological research in living patients with PD.

Acute and subacute drug-induced movement disorders.

Many pharmacological agents may induce a variety of movement disorders, including dystonia, tremor, parkinsonism, myoclonus and dyskinesia, with an acute, subacute or more chronic time course. Motor symptoms may be isolated or part of a more extensive cerebral or systemic condition, such as the neuroleptic malignant syndrome or the serotonin syndrome. Drug-induced movement disorders share a number of features that should make them easy to identify, including a clear temporal relationship between medication initiation and symptom onset, a dose-effect, and, with the exception of tardive syndromes, complete resolution after discontinuation of the offending agent. Diagnosis relies on a thorough medication history. Medications commonly involved include dopamine receptor blockers, antidepressants and anti-epileptics, among many others. Mechanisms underlying drug-induced movement disorders involve blockade, facilitation or imbalance of dopamine, serotonin, noradrenaline and cholinergic neurotransmission in the basal ganglia. The present review focuses on drug-induced movement disorders that typically develop as an acute (hours to days) or subacute (days to weeks) event, including acute dystonic reactions, akathisia, drug-induced parkinsonism, neuroleptic malignant syndrome, serotonin syndrome, parkinsonism-hyperpyrexia syndrome, drug-induced tremor, drug-induced hyperkinesias and movement disorders associated with the use of recreational drugs.

Comparison of gel-based methods for the detection of cerebrospinal fluid rhinorrhea.

Cerebrospinal fluid (CSF) rhinorrhea is a serious condition that may result in severe complications. Various laboratory tests, relying on the detection of CSF-specific proteins in nasal secretions, have been developed but diagnosis remains challenging. The aim of this study was to evaluate two new methods targeting either ß2-transferrin or beta-trace-protein. Rhinorrhea samples from patients suspected of CSF leakage (n=36) were analyzed using two-dimensional gel electrophoresis (2-DE) for CSF rhinorrhea diagnosis. Twelve patients with rhinorrhea strongly suggestive of a CSF leak also underwent a fluorescein test. The same cohort was retrospectively analyzed with a beta-trace protein immunoblot developed in-house (n=36) and a new commercial ß2-transferrin immunofixation assay (Sebia, Evry, France) (n=33). 2-DE was positive in 9 patients suffering from rhinorrhea following skull base fracture (n=3), post-surgery (n=4), or spontaneously (n=2). The 27 remaining cases were negative. These results were confirmed by the beta-trace protein immunoblot and ß2-transferrin immunofixation tests, except for one sample found negative with 2-DE but positive with the two other assays. Results from the three analytical methods were concordant with fluorescein tests. Beta-trace protein immunoblot and ß2-transferrin immunofixation assays are fast and reliable methods that allow detecting CSF leakage in nasal fluid with high sensitivity and specificity.

From relative to absolute quantification of tryptic peptides with tandem mass tags: application to cerebrospinal fluid.

Quantification is a major task in proteomics. Among the different analytical strategies to enable peptide and protein quantification, tagging with isotopic labels has emerged as a practical, versatile, and efficient alternative. In particular, isobaric labels, such as TMT or iTRAQ, are now widely employed to make relative comparison of the protein amounts in separate biological samples with tandem mass spectrometry (MS/MS). We used herein a shotgun proteomic approach based on labelling with tandem mass tags (TMTs) for the relative quantification of proteins, and the absolute quantification of their tryptic peptides in human cerebrospinal fluid (CSF). First, the comparison of ante- and post-mortem CSF samples was carried out for the discovery of protein marker candidates of brain-damage disorders. Second, tryptic peptides representative of these candidates were measured in CSF using reporter-ion calibration curves. These works highlighted the advantages and limitations of such strategies for quantification purposes in proteomics.

Pluripotent stem cells as new drugs? The example of Parkinson's disease.

Cell replacement therapy is a widely discussed novel concept of medical treatment. The increased knowledge in the stem cell field, particularly pluripotent stem cells, potentially provides powerful tools for this therapeutic concept. A large number of disease characterized by the loss of functional cells are potential candidates for cell replacement therapy and, in this regards, Parkinson's disease is of particular interest. It is one of the most prevalent neurodegenerative diseases caused by the loss of dopaminergic neurons in the Substantia nigra pars compacta. Pharmacological therapies are valuable but suffer from the progressive decline of efficacy as the disease progresses. Cell therapy application has emerged about two decades ago as a valid therapeutic alternative and recent advances in stem cell research suggest that pluripotent stem cell transplantation may be a promising approach to replace degenerated neurons in Parkinson's disease. Various sources of pluripotent stem cells (PSC) currently tested in animal models of Parkinson's disease have proven their efficacy in relieving symptoms and restoring damaged brain function. This review summarizes and discusses the important challenges that actually must be solved before the first studies of PSC transplantation can be undertaken into humans.

Long-term cognitive profile and incidence of dementia after STN-DBS in Parkinson's disease.

An effect of subthalamic nucleus deep brain stimulation (STN-DBS) on cognition has been suspected but long-term observations are lacking. The aim of this study was to evaluate the long-term cognitive profile and the incidence of dementia in a cohort of Parkinson's disease (PD) patients treated by STN-DBS. 57 consecutive patients were prospectively assessed by the mean of a neuropsychological battery over 3 years after surgery. Dementia (DSM-IV) and UPDRS I to IV were recorded. 24.5% of patients converted to dementia over 3 years (incidence of 89 of 1,000 per year). This group of patients cognitively continuously worsened over 3 years up to fulfilling dementia criteria (PDD). The rest of the cohort remained cognitively stable (PD) over the whole follow-up. Preoperative differences between PDD and PD included older age (69.2 +/- 5.8 years; 62.6 +/- 8 years), presence of hallucinations and poorer executive score (10.1 +/- 5.9; 5.5 +/- 4.4). The incidence of dementia over 3 years after STN-DBS is similar to the one reported in medically treated patients. The PDD presented preoperative risk factors of developing dementia similar to those described in medically treated patients. These observations suggest dementia being secondary to the natural evolution of PD rather than a direct effect of STN-DBS.

Long-duration response to levodopa in patients with advanced Parkinson disease treated with subthalamic deep brain stimulation.

BACKGROUND: Long-duration response (LDR) to levodopa is supposed to decrease with Parkinson disease (PD) progression, but direct observation of this response in advanced PD has never been performed. OBJECTIVE: To study the LDR to levodopa in patients with advanced PD treated with subthalamic deep brain stimulation (DBS). DESIGN AND SETTING: We studied 30 consecutive patients with PD who underwent subthalamic DBS. One group had no antiparkinsonian treatment since surgery (no-levodopa group), whereas medical treatment had to be reinitiated in the other group (levodopa group). MAIN OUTCOME MEASURE: Motor subscale score of the Unified Parkinson's Disease Rating Scale. RESULTS: Compared with preoperative assessment, evaluation 6 months postoperatively with DBS turned off for 3 hours found a worsening of the motor subscale score of the Unified Parkinson's Disease Rating Scale in the no-levodopa group. This worsening being absent in the levodopa group, it probably reflected the loss of the LDR to levodopa in the no-levodopa group. When DBS was turned on, postoperative motor subscale scores of the Unifid Parkinson's Disease Rating Scale in both groups were similar to preoperative scores while receiving medication, suggesting that subthalamic DBS compensated for the short-duration response and LDR to levodopa. CONCLUSIONS: Our results suggest that the LDR to levodopa remains significant even in advanced PD, and that subthalamic DBS compensates for the short-duration response and LDR to levodopa.

Does subthalamic nucleus deep brain stimulation really improve quality of life in Parkinson's disease?

We investigated the impact of subthalamic nucleus (STN) deep brain stimulation (DBS) on quality of life (QOL) in patients with advanced Parkinson's disease, as self-assessed before and after surgery by completing the Parkinson's Disease Questionnaire (PDQ39). In addition to this prospective evaluation, we asked patients postoperatively to evaluate their preoperative QOL. In the prospective assessment, results showed that patients perceived a general improvement of QOL after the STN DBS. However, when evaluated retrospectively, they tended to overestimate their preoperative functioning, therefore obscuring the improvement found prospectively. This observation highlights the impact of the method used on obtained results when assessing the effects of STN DBS.

PARK7 and nucleoside diphosphate kinase A as plasma markers for the early diagnosis of stroke.

BACKGROUND: Plasma markers for stroke could be useful in diagnosis and prognosis and in prediction of response of stroke patients to therapy. PARK7 and nucleoside diphosphate kinase A (NDKA) are increased in human postmortem cerebrospinal fluid (CSF), a model of global brain insult, suggesting that measurement in CSF and, more importantly, in plasma may be useful as a biomarker of stroke. METHODS: We used ELISA to measure PARK7 and NDKA in plasma in 3 independent European and North American retrospective studies encompassing a total of 622 stroke patients and 165 control individuals. RESULTS: Increases in both biomarkers were highly significant, with sensitivities of 54%-91% for PARK7 and 70%-90% for NDKA and specificities of 80%-97% for PARK7 and 90%-97% for NDKA. The concentrations of both biomarkers increased within 3 h of stroke onset. CONCLUSIONS: PARK7 and NDKA may be useful plasma biomarkers for the early diagnosis of stroke. In addition, this study demonstrated the utility of analysis of postmortem CSF proteins as a first step in the discovery of plasma markers of ischemic brain injury.

Truncated cystatin C in cerebrospiral fluid: Technical [corrected] artefact or biological process?

Cystatin C, a low molecular weight cysteine proteinase inhibitor present in human body fluids at physiological concentrations, is more expressed in cerebrospinal fluid (CSF) than in plasma. Mass spectrometric characterization showed that after 3 months of storage of human CSF at -20 degrees C, cystatin C was cleaved in the peptide bond between R8 and L9 and lost its eight N-termini amino acids, whereas this cleavage did not occur when stored at -80 degrees C. This truncation occurred in all CSF samples studied irrespective of the underlying neurological status, indicating a storage-related artefact rather than a physiological or pathological processing of the protein. These results stress the importance of optimal preanalytical storage conditions of any sample prior to proteomics studies.

Cystatin C as a potential cerebrospinal fluid marker for the diagnosis of Creutzfeldt-Jakob disease.

The definite diagnosis of Creutzfeldt-Jakob disease (CJD), the most common form of human prion diseases, relies upon neuropathological data usually obtained at autopsy. In living patients, the diagnosis, based on suggestive clinical features and EEG abnormalities, can be aided by the detection of altered levels of isoforms of the 14-3-3 protein in the cerebrospinal fluid (CSF). However, the validity of this test has been recently challenged and the search for other, more reliable biomarkers for CJD remains highly desirable. The present study describes the identification of a new potential surrogate marker in the CSF of CJD-affected patients. A preliminary study employing surface-enhanced laser desorption/ionization-time of flight (SELDI-TOF) technology highlighted a protein at 13.4 kDa in a small group (n = 8) of CJD-affected patients. Further analysis aimed at identifying this protein using cationic exchange chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed it to be cystatin C. Additional immunoblot assays confirmed that the level of cystatin C was significantly increased (p