Masiulienė, Ieva

Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system that causes inflammatory and neurodegenerative lesions. Ectracellular vesicles are small membrane-bound structures secreted by all types of cells and found in various biological fluids. Currently, the diagnosis of MS is based on clinical, radiological and laboratory criteria, but especially in the early stages, these markers are often non-specific. For this reason, additional diagnostic markers are being sought that can increase diagnostic accuracy and complement existing MS diagnostic methods. One of the significant molecules that can increase diagnostic accuracy is microRNA (miRNA). MiRNAs are small, non-coding RNA molecules that regulate gene expression and can be used as circulating biomarkers for various diseases, including MS. MiRNAs were isolated from serum extracellular vesicles from 49 MS patients and 49 controls using „ExoRNeasy Midi Kit“. MiRNA concentration and quality were assessed by using „NanoDrop 2000“ and cDNA synthesis was performed using the “TaqMan® Advanced miRNA cDNA Synthesis Kit“. MiRNA expression was detected by using RTPCR method. RT-PCR results were analyzed with „QuantStudio™ Design & Analysis Software“. The obtained data were processed and statistically evaluated using the „IBM SPSS Statistics 29.0.2.0“ program. It was found that the expression of both studied miRNAs was significantly different between the MS and control groups of neurologically healthy individuals (p0.05). ROC curve analysis revealed the diagnostic and prognostic potential of the studied miRNAs, assessed by AUC values. The results obtained indicate that the studied miRNA-21-5p and miRNA-106a-p may be suitable as potential biomarkers in multiple sclerosis.

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system characterized by considerable clinical heterogeneity. Its diagnosis and prognosis remain challenging – although clinical, radiological, and laboratory criteria are applied, reliable biomarkers capable of enabling earlier detection, predicting relapses, or assessing therapeutic efficacy are still lacking. Moreover, treatment effectiveness is often limited by inter-individual variability in therapeutic response and the complexity of disease progression. One promising class of biomarkers is microRNAs (miRNAs), small non-coding RNA molecules that regulate gene expression at the transcriptional or post-transcriptional level. However, due to their broad diversity, identifying biologically relevant miRNAs requires integration of in silico and experimental approaches. Our study consisted of two major stages: (1) in silico miRNA modeling and (2) an experimental sequencing phase. In the silico phase, we computationally derived predicted miRNA expression profiles in the context of MS. MiRNA sequencing was performed using the Illumina platform on extracellular vesicle samples isolated from blood (9 MS patients and 13 healthy controls). Differential expression analysis was conducted using the DESeq2 package to evaluate miRNA expression profiles between MS patients and healthy individuals. Overlapping miRNAs identified through both in silico prediction and sequencing were compared to assess their biological relevance to MS. Four miRNAs predicted in silico were confirmed in sequencing data. Among them, hsa-miR-19a-3p showed the highest expression change (log₂FoldChange > 1), while hsa-miR-17-5p, hsa-miR-139-5p, and hsa-miR-20a-5p exhibited moderate but significant alterations. KEGG analysis revealed enrichment in pathways of neurodegeneration – multiple diseases, amyotrophic lateral sclerosis (ALS), and the MAPK signaling pathway, suggesting shared molecular mechanisms between MS and other neurodegenerative disorders. The integration of in silico modeling with high-throughput sequencing analysis effectively identifies biologically relevant miRNAs in multiple sclerosis and highlights their potential as biomarkers involved in neurodegenerative and inflammatory signaling pathways.

Multiple sclerosis (MS) is an inflammatory, demyelinating disease of the central nervous system that leads to progressive neurodegeneration and diverse neurological symptoms, most commonly affecting young and middle-aged women. MicroRNAs play key regulatory roles in immune function and myelin repair, making them potential markers of disease mechanisms and treatment response. MS is managed with disease-modifying therapies classified as first-line agents (e.g., interferon beta, glatiramer acetate) and second-line agents (e.g., natalizumab, fingolimod). However, reliable biomarkers capable of objectively assessing treatment effectiveness in individual patients are still lacking, underscoring the need to identify molecular indicators of therapeutic response. RNA was extracted from blood using the “MirVana™ miRNA Isolation Kit”. Nucleic acid concentration and purity were determined using a NanoDrop 2000 spectrophotometer. Complementary DNA was synthesized with “TaqMan® Advanced miRNA Assays”, and miRNA expression was quantified by real-time PCR. Resulting data were processed using “QuantStudio™ Design & Analysis Software”, and statistical analyses were performed in SPSS. Significant differences in the expression of both examined microRNAs were observed between treatment groups, with the highest levels detected in patients receiving secondline therapy (p < 0.05). A significant positive correlation between miR-204-5p and miR17-5p expression was also identified, most pronounced in the second-line treatment group (p < 0.05). MiR-204-5p and miR-17-5p showed distinct expression patterns across treatment groups, with both microRNAs most highly expressed in patients receiving second-line therapy. When comparing expression levels before and after both first-line and secondline treatment, miR-204-5p showed a slight increase, while miR-17-5p showed a slight decrease. However, these changes were not statistically significant. It was found that the interaction between the two microRNAs involved in the remyelination process was significant in patients treated with second-line therapies.

Multiple sclerosis (MS) is an autoimmune disorder characterized by demyelination of neurons in the central nervous system, resulting in axonal damage and subsequent motor, sensory, and cognitive dysfunctions. Current therapeutic strategies primarily aim to suppress immune activation and infiltration. Consequently, elucidating the biological mechanisms underlying myelin repair – remyelination – has become a central focus of MS research. One of the potential regulators of the remyelination process is microRNAs, which are involved in the differentiation of oligodendrocyte precursor cells into mature oligodendrocytes. It remains unclear whether disease-modifying therapies influence the expression of miRNAs associated with remyelination in the blood. The aim of this study is to investigate the expression of miR-204-5p and miR-17-5p involved in the remyelination process – both before and after treatment – and healthy controls, in order to identify treatment-related differences. The expression of miRNAs were analyzed in extracellular vesicles isolated from the blood serum of three groups: a control group (n=25), untreated MS patients (n=18), and MS patients treated with second-line disease-modifying therapies prior to the study (n=24). MiRNA isolation was performed using the ExoRNeasy Midi Kit (Qiagen), and cDNA synthesis was conducted with the TaqMan Advanced miRNA cDNA Synthesis Kit (AB). QPCR was performed using the QuantStudioTM 3 System. Statistical analyses were performed using GraphPad Prism and SPSS software. A statistically significant difference in miR-17-5p expression was observed between the control group and MS patients treated with second-line disease-modifying therapies (p=0.0415). The diagnostic performance of miR-17-5p for distributing these two groups was moderate (AUC=0.6719; 95% CI: 0.5154-0.8283). MiR-17-5p expression was significantly higher in MS patients treated with second-line disease-modifying therapies compared with the control group. These findings suggest a potential association between miR-17-5p expression and treatment-induced modulation of remyelination processes in MS.

Multiple sclerosis (MS) is a complex autoimmune disorder driven by diverse molecular, genetic, and immunological processes, and characterized by diverse clinical course forms. The identification of reliable biomarkers is essential for improving our understanding of MS disease mechanisms and for developing more advanced diagnostic and prognostic strategies. microRNA expression profiling represents one of the most promising approaches for revealing molecular alterations associated with MS pathogenesis. The aim of this study was to identify potential MS biomarkers by applying sequencingbased expression analysis and machine-learning methods using untreated samples. MiRNA sequencing was performed on samples from 9 MS patients and 13 healthy controls. Following data quality control, principal component analysis (PCA) was applied to evaluate sample structure and group separation. Differential expression analysis was conducted using DESeq2, with results visualized through a volcano plot and heatmap. In parallel, sPLS-DA was employed to identify features contributing most strongly to group discrimination. Molecules overlapping between DESeq2 and sPLS-DA results were further evaluated using boxplots and univariate logistic regression models. PCA analysis showed a clear separation between the MS and control groups based on miRNA expression. DESeq2 identified miRNAs that were significantly differentially expressed. sPLS-DA further narrowed down the set of features with strong ability to distinguish the groups. All miRNAs selected by sPLS-DA overlapped with those found by DESeq2, supporting their stability and biological relevance. Logistic regression showed that even single miRNAs from this set could significantly separate the two groups. The integrated DESeq2 and sPLS-DA analysis identified a group of miRNAs that were statistically different between MS patients and healthy controls and showed strong discriminatory ability. The overlapping miRNAs represent strong candidates for potential diagnostic or prognostic MS biomarkers. The analysis was performed using untreated samples, making the results more reliable and providing a solid basis for future validation studies.

Šioje apžvalgoje aptariami klinikiniai tyrimai ir atvejai, kuriais tirtas monokloninių antikūnų prieš su kalcitonino genu susijusį peptidą (angl. calcitonin gene-related peptide, CGRP) efektyvumas gydant ne migreninius galvos skausmus. Remiantis naujausiais straipsniais, apžvelgta galvos skausmų patofiziologija ir CGRP reikšmė patogenezėje. Atrinktuose klinikiniuose tyrimuose ir atvejų analizėse nagrinėtas galkanezumabo, fremanezumabo ir erenumabo efektyvumas gydant klasterinį galvos skausmą, trišakio nervo neuralgiją bei potrauminį galvos skausmą. Pastaruoju metu daugėja duomenų, kad CGRP dalyvauja įvairių pirminių ir antrinių galvos skausmo ligų patogenezėje. Monokloninių antikūnų prieš CGRP efektyvumas gydant klasterinį galvos skausmą ir trišakio nervo neuralgiją šiuo metu yra grindžiamas klinikiniais tyrimais ir atvejų analizėmis, tačiau reikia tolesnių klinikinių atsitiktinai parinktų imčių placebu kontroliuojamų tyrimų.

Background and Objectives: Recent findings suggest that neurodegeneration starts early in the course of multiple sclerosis (MS) and significantly contributes to the progression of patients’ disability. Tau is a microtubule-binding protein that is known to play a role in the pathophysiology of many neurodegenerative disorders. Newly emerging data on tau protein-induced neurodegenerative processes and its possible involvement in MS suggest that it may be involved in the pathology of early-stage MS. Therefore, this study aimed to test this hypothesis in patients with newly diagnosed MS. Materials and Methods: Cerebrospinal fluid (CSF) was collected from 19 patients with newly diagnosed MS and 19 control subjects. All MS patients underwent neurological examination, lumbar punction, and brain magnetic resonance imaging (MRI). CSF concentrations of total and phosphorylated tau (phospho-tau-181) protein were measured using commercial enzyme-linked immunosorbent assay kits. Results: The total tau concentration was significantly higher in the CSF of MS patients compared to controls (141.67 pg/mL, IQR 77.79–189.17 and 68.77 pg/mL, IQR 31.24–109.17, p = 0.025). In MS patients, the total tau protein positively correlated with total CSF protein (r = 0.471, p = 0.048). Significantly higher total tau concentration was measured in MS patients with higher lesion load in brain MRI (≥9 versus <9 lesions; 168.33 pg/mL, IQR 111.67–222.32 and 73.33 pg/mL, IQR -32.13–139.29-, p = 0.021). The CSF concentration of phospho-tau-181 protein was below the detection limit in both MS and control subjects. Conclusions: The concentration of total tau protein level is elevated, whereas phospho-tau-181 is undetectable in the CSF of patients with early-stage MS.

Išsėtinė sklerozė (IS) - tai lėtinė centrinės nervų sistemos (CNS) liga, kuri dažniausiai diagnozuojama jauno amžiaus suaugusiesiems. Per pastaruosius dešimtmečius atsiradę nauji gydymo metodai kardinaliai pakeitė šių pacientų ligos prognozę ir gyvenimo kokybę, tačiau taip pat iškėlė ir naujų iššūkių, prognozuojant ligos eigą, nustatant ligos aktyvumą prieš išsivystant naujai negalią sunkinančiai neurologinei simptomatikai ir siekiant laiku paskirti ligos eigą modifikuojantį gydymą individualiam pacientui tinkamiausiu vaistu. Viena iš priemonių, galinčių padėti atsakyti į šiuos klausimus, galėtų būti laboratoriniai IS biožymenys. Be jau gerai žinomų ir klinikinėje praktikoje naudojamų oligokloninių juostų (OGJ) ir imunoglobulino G indekso, atrasti ir kiti rodikliai, galintys padėti nustatyti CNS vykstančius uždegiminius ir neurodegeneracinius procesus. Kaip galimas naujas diagnostinis IS biožymuo moksliniuose tyrimuose išskiriamos kappa lengvosios laisvosios grandinės ir K-indeksas, kuris pasižymi panašiu jautrumu ir specifiškumu kaip OGJ. Taip pat atrasti biožymenys, kurie gali padėti diferencijuoti kliniškai izoliuotą sindromą nuo IS ir diferencijuoti IS ligos eigą. Šiuo metu į chitinazę-3 panašaus baltymo 1 (angl. chitinase-3-like protein 1) koncentracija yra vienintelis žymuo, kurio ištyrimas smegenų skystyje gali padėti atskirti IS nuo kliniškai izoliuoto sindromo. Glijos fibrilinio rūgštinio baltymo (angl. glial fibrillary acidic protein) nustatymas smegenų skystyje ir kraujo serume gali padėti diferencijuoti pirminę progresuojančią IS nuo recidyvuojančios remituojančios ligos eigos. Naudin¬giausiu ligos aktyvumo ir gydymo efektyvumo monitoravimo biožymeniu yra laikoma serumo neurofilamentų lengvųjų grandinių koncentracija. Šiame straipsnyje aptariami perspektyviausi IS diagnostikos, ligos aktyvumo ir atsako į gydymą biožymenys.

Anti-neutrophil cytoplasmic antibodies (ANCA)-associated vasculitides (AAVs) are a group of autoimmune diseases that can affect many vital organs and tissues, as well as the nervous system. When peripheral nerves are affected, one of the clinical features of AAVs can be vasculitic neuropathies (VNs). This usually causes asymmetric weakness and numbness in the distal parts of extremities, frequently followed by pain. Nerve conduction study (NCS) reveals axonal loss in multiple individual nerves, whereas a length-dependent process may also be observed. As AAV is a rare diagnostic entity and can manifest with a vast variety of symptoms, it is seldom involved in the initial diagnostic work-up of polyneuropathy. Moreover, the only definitive diagnostic method for VN is peripheral nerve biopsy. Therefore, AAV represents one of the greatest difficulties in the differential diagnosis of neuropathies. In this article, we present a clinical case of a 73-year-old patient admitted to the hospital due to blurry vision, headache, and limb weakness. The findings of the neurological examination were consistent with polyneuropathy. A thorough examination was performed to determine the exact diagnosis. However, due to the lack of typical symptoms, VN was diagnosed only after acute renal failure evolved. This case demonstrates the many symptoms that AAV can present with and the importance of a thorough differential diagnosis for multiple mononeuropathies.

Introduction: Multiple sclerosis (MS) is a widely spread and debilitating disease with 2.8 million people worldwide currently affected (Walton et al., 2020). Even in a relatively short period of time of 2 years at least one third of MS patients report disability progression (Pellegrini et al., 2020), however the exact pathogenesis of the disease still remains incompletely understood. Cerebrospinal Fluid Oligoclonal Bands (CSF OCBs) can be found in up to 85% of MS patients (Garg et al., 2015), nowadays they play a key role in the diagnostics of MS while presenting a prognostic value as well (Thompson et al., 2017). Objectives/Aims: The study aimed to evaluate the association between CSF OCBs status and clinical features in patients with MS in Lithuania. Methods: The selection of 200 MS patients treated in Lithuanian University of Health Sciences (LUHS) Kaunas Clinics was performed in order to find associations between CSF OCBs status and various disease features. The data was acquired from outpatient records and a retrospective analysis was performed. Variables included in the selection were patient age (at diagnosis and first symptoms), sex, course of the disease, EDSS (Expanded Status Disability Scale) scores at the first (EDSS1) and the last (EDSS2) visit, MRI data and CSF OCBs status. Results were seen as statisctically signifcant when p<0.05. Results: Out of 200 reviewed MS cases 65 (32.5%) were male patients and 135 (67.5%) were female. Mean age of diagnosis was 36.4 ±11.668. Mean age at first symptoms was 32.13 ±10.790. Positive CSF OCBs were found in 151 (75.5%) of the cases. There were no statistically significant associations between OCBs status and sex, as well as the course of the disease. OCBs positive MS patients (median=35 (18-74)) were diagnosed with MS earlier than OCBs negative patients (median=39 (20-59), U=3084, p=0.039) No relations were found between OCBs positivity results and patient age at the symptom onset, EDSS scores of first and last visit and its change through time. The majority of patients (81.3%) with spinal cord lesions were OCBs positive, meaning that spinal cord lesions were more frequently found in OCBs positive patients (χ2=4.473, p=0.034). Other MRI lesion localizations as well as the progression and number of MRI lesions were not associated with OCBs status. Conclusion: OCBs positive patients were diagnosed with MS earlier and had spinal cord lesions more frequently than OCBs negative patients.