Simultaneous Determination of Selected Steroids with Neuroactive Effects in Human Serum by Ultrahigh-Performance Liquid Chromatography-Tandem Mass Spectrometry.

Neuroactive steroids are a group of steroid molecules that are involved in the regulation of functions of the nervous system. The nervous system is not only the site of their action, but their biosynthesis can also occur there. Neuroactive steroid levels depend not only on the physiological state of an individual (person's sex, age, diurnal variation, etc.), but they are also affected by various pathological processes in the nervous system (some neurological and psychiatric diseases or injuries), and new knowledge can be gained by monitoring these processes. The aim of our research was to develop and validate a comprehensive method for the simultaneous determination of selected steroids with neuroactive effects in human serum. The developed method enables high throughput and a sensitive quantitative analysis of nine neuroactive steroid substances (pregnenolone, progesterone, 5α-dihydroprogesterone, allopregnanolone, testosterone, 5α-dihydrotestosterone, androstenedione, dehydroepiandrosterone, and epiandrosterone) in 150 µL of human serum by ultrahigh-performance liquid chromatography with tandem mass spectrometry. The correlation coefficients above 0.999 indicated that the developed analytical procedure was linear in the range of 0.90 nmol/L to 28.46 µmol/L in human serum. The accuracy and precision of the method for all analytes ranged from 83 to 118% and from 0.9 to 14.1%, respectively. This described method could contribute to a deeper understanding of the pathophysiology of various diseases. Similarly, it can also be helpful in the search for new biomarkers and diagnostic options or therapeutic approaches.

Prognostic relevance of the C-X-C motif chemokine ligand 13 and interleukin-8 in predicting the transition from clinically isolated syndrome to multiple sclerosis.

The initial phase of multiple sclerosis (MS), often known as clinically isolated syndrome (CIS), is a critical period for identifying individuals at high risk of progressing to full-blown MS and initiating timely treatment. In this study, we aimed to evaluate the prognostic value of C-X-C motif chemokine ligand 13 (CXCL13) and interleukin-8 (IL-8) as potential markers for CIS patients' conversion to MS. Our study encompassed patients with CIS, those with relapsing-remitting MS (RRMS), and control subjects, with sample analysis conducted on both cerebrospinal fluid (CSF) and serum. Patients were categorized into four groups: CIS-CIS (no MS development within 2 years), CIS-RRMS (conversion to RRMS within 2 years), RRMS (already diagnosed), and a control group (CG) with noninflammatory central nervous system disorders. Results showed significantly elevated levels of CXCL13 in CSF across all patient groups compared with the CG (p < 0.0001, Kruskal-Wallis test). Although CXCL13 concentrations were slightly higher in the CIS-RRMS group, statistical significance was not reached. Similarly, significantly higher levels of IL-8 were detected in CSF samples from all patient groups compared with the CG (p < 0.0001, Kruskal-Wallis test). Receiver operating characteristic analysis in the CIS-RRMS group identified both CXCL13 (area under receiver operating characteristic curve = .959) and IL-8 (area under receiver operating characteristic curve = .939) in CSF as significant predictors of CIS to RRMS conversion. In conclusion, our study suggests a trend towards elevated CSF IL-8 and CSF CXCL13 levels in CIS patients progressing to RRMS. These findings emphasize the importance of identifying prognostic markers to guide appropriate treatment strategies for individuals in the early stages of MS.

Patients with Neurodegenerative Proteinopathies Exhibit Altered Tryptophan Metabolism in the Serum and Cerebrospinal Fluid.

Some pathological conditions affecting the human body can also disrupt metabolic pathways and thus alter the overall metabolic profile. Knowledge of metabolic disturbances in specific diseases could thus enable the differential diagnosis of otherwise similar conditions. This work therefore aimed to comprehensively characterize changes in tryptophan metabolism in selected neurodegenerative diseases. Levels of 18 tryptophan-related neuroactive substances were determined by high throughput and sensitive ultrahigh-performance liquid chromatography-tandem mass spectrometry in time-linked blood serum and cerebrospinal fluid samples from 100 age-matched participants belonging to five cohorts: healthy volunteers (n = 21) and patients with Lewy body disease (Parkinson's disease and dementia with Lewy bodies; n = 31), four-repeat tauopathy (progressive supranuclear palsy and corticobasal syndrome; n = 10), multiple system atrophy (n = 13), and Alzheimer's disease (n = 25). Although these conditions have different pathologies and clinical symptoms, the discovery of new biomarkers is still important. The most statistically significant differences (with p-values of ≤0.05 to ≤0.0001) between the study cohorts were observed for three tryptophan metabolites: l-kynurenine in cerebrospinal fluid and 3-hydroxy-l-kynurenine and 5-hydroxy-l-tryptophan in blood serum. This led to the discovery of distinctive correlation patterns between the profiled cerebrospinal fluid and serum metabolites that could provide a basis for the differential diagnosis of neurodegenerative tauopathies and synucleinopathies. However, further large-scale studies are needed to determine the direct involvement of these metabolites in the studied neuropathologies, their response to medication, and their potential therapeutic relevance.

Utilizing neurodegenerative markers for the diagnostic evaluation of amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by progressive deterioration of upper and lower motor neurons. A definitive diagnostic test or biomarker for ALS is currently unavailable, leading to a diagnostic delay following the onset of initial symptoms. Our study focused on cerebrospinal fluid (CSF) concentrations of clusterin, tau protein, phosphorylated tau protein, and beta-amyloid1-42 in ALS patients and a control group. METHODS: Our study involved 54 ALS patients and 58 control subjects. Among the ALS patients, 14 presented with bulbar-onset ALS, and 40 with limb-onset ALS. We quantified biomarker levels using enzyme-linked immunosorbent assay (ELISA) and compared the results using the Mann-Whitney U-test. RESULTS: Significant elevations in neurodegenerative markers, including tau protein (p < 0.0001), phosphorylated tau protein (p < 0.0001), and clusterin (p = 0.038), were observed in ALS patients compared to controls. Elevated levels of tau protein and phosphorylated tau protein were also noted in both bulbar and limb-onset ALS patients. However, no significant difference was observed for beta-amyloid1-42. ROC analysis identified tau protein (AUC = 0.767) and p-tau protein (AUC = 0.719) as statistically significant predictors for ALS. CONCLUSION: Our study demonstrates that neurodegenerative marker levels indicate an ongoing neurodegenerative process in ALS. Nonetheless, the progression of ALS cannot be predicted solely based on these markers. The discovery of a specific biomarker could potentially complement existing diagnostic criteria for ALS.

Cerebrospinal fluid and blood serum biomarkers in neurodegenerative proteinopathies: A prospective, open, cross-correlation study.

Neurodegenerative diseases are a broad heterogeneous group affecting the nervous system. They are characterized, from a pathophysiological perspective, by the selective involvement of a subpopulation of nerve cells with a consequent clinical picture of a disease. Clinical diagnoses of neurodegenerative diseases are quite challenging and often not completely accurate because of their marked heterogeneity and frequently overlapping clinical pictures. Efforts are being made to define sufficiently specific and sensitive markers for individual neurodegenerative diseases or groups of diseases in order to increase the accuracy and speed of clinical diagnosis. Thus said, this present research aimed to identify biomarkers in the cerebrospinal fluid (CSF) and serum (α-synuclein [α-syn], tau protein [t-tau], phosphorylated tau protein [p-tau], ß-amyloid [Aß], clusterin, chromogranin A [chromogrA], cystatin C [cyst C], neurofilament heavy chains [NFH], phosphorylated form of neurofilament heavy chains [pNF-H], and ratio of tau protein/amyloid beta [Ind tau/Aß]) that could help in the differential diagnosis and differentiation of the defined groups of α-synucleinopathies and four-repeat (4R-) tauopathies characterized by tau protein isoforms with four microtubule-binding domains. In this study, we analyzed a cohort of 229 patients divided into four groups: (1) Parkinson's disease (PD) + dementia with Lewy bodies (DLB) (n = 82), (2) multiple system atrophy (MSA) (n = 25), (3) progressive supranuclear palsy (PSP) + corticobasal syndrome (CBS) (n = 30), and (4) healthy controls (HC) (n = 92). We also focused on analyzing the biomarkers in relation to each other with the intention of determining whether they are useful in distinguishing among individual proteinopathies. Our results indicate that the proposed set of biomarkers, when evaluated in CSF, is likely to be useful for the differential diagnosis of MSA versus 4RT. However, these biomarkers do not seem to provide any useful diagnostic information when assessed in blood serum.

Quantitative Determination of Endogenous Tetrahydroisoquinolines, Potential Parkinson's Disease Biomarkers, in Mammals.

Current diagnostic options for Parkinson's disease are very limited and primarily based on characteristic clinical symptoms. Thus, there are urgent needs for reliable biomarkers that enable us to diagnose the disease in the early stages, differentiate it from other atypical Parkinsonian syndromes, monitor its progression, increase knowledge of its pathogenesis, and improve the development of potent therapies. A promising group of potential biomarkers are endogenous tetrahydroisoquinoline metabolites, which are thought to contribute to the multifactorial etiology of Parkinson's disease. The aim of this critical review is to highlight trends and limitations of available traditional and modern analytical techniques for sample pretreatment (extraction and derivatization procedures) and quantitative determination of tetrahydroisoquinoline derivatives in various types of mammalian fluids and tissues (urine, plasma, cerebrospinal fluid, brain tissue, liver tissue). Particular attention is paid to the most sensitive and specific analytical techniques, involving immunochemistry and gas or liquid chromatography coupled with mass spectrometric, fluorescence, or electrochemical detection. The review also includes a discussion of other relevant agents proposed and tested in Parkinson's disease.