Phantom limb pain (PLP) is a frequent and often persistent consequence of limb amputation, characterized by pain perceived in the absent limb. Despite decades of research, its biological basis remains incompletely understood, and available treatments often provide inconsistent relief. This reflects the complex and heterogeneous nature of phantom limb pain, which cannot be explained by a single anatomical site or pathological process. Current evidence suggests that phantom limb pain emerges from the interaction of changes occurring at multiple levels of the nervous system. Peripheral nerve injury associated with amputation induces molecular and cellular alterations that may influence early nociceptive signaling. These changes can interact with adaptive and maladaptive responses within the spinal cord, including altered synaptic transmission and neuron–glia interactions, which may facilitate sustained amplification of pain-related signals. At supraspinal levels, long-term adaptations within distributed neural networks involved in sensory, motor, and affective processing may contribute to the persistence of pain perceptions in the absence of ongoing peripheral input. Immune-related signaling and long-term regulation of gene expression further modulate these processes and may contribute to inter-individual variability. In this narrative review, we synthesize current experimental and clinical evidence addressing the molecular and cellular processes associated with phantom limb pain following lower limb amputation. Findings are integrated across peripheral, spinal, and supraspinal levels, with consideration of immune-related and regulatory influences. By highlighting areas of convergence, uncertainty, and existing knowledge gaps, this review aims to provide a coherent biological framework that may support future experimental and translational research in this challenging field.

Distinguishing relapsing–remitting multiple sclerosis (RRMS) from secondary-progressive disease (SPMS) remains a clinical challenge, as no single laboratory test reliably identifies the transition. We explored whether plasma extracellular-vesicle (EV) microRNAs could separate the two phenotypes. Four candidate EV-miRNAs: miR-30a-5p, miR-223-5p, miR-155-5p, and miR-146a-5p, were quantified by qRT-PCR in plasma-derived EVs from 13 healthy controls (HC), 7 RRMS, and 16 SPMS patients, normalized to miR-16-5p. EVs were characterized by electron microscopy, dynamic light scattering, and zeta potential, confirming vesicles of expected morphology, size, and negative surface charge. Two miRNAs were informative. miR-155-5p was significantly downregulated in both RRMS and SPMS versus controls but did not differentiate the phenotypes (AUC 0.52), behaving as a disease-general marker. In contrast, miR-223-5p was selectively reduced in SPMS, to approximately one-quarter of control abundance, and provided the only signal distinguishing progressive from relapsing disease (AUC 0.73; rank-biserial −0.46). miR-30a-5p and miR-146a-5p were uninformative. These findings, consistent with the circulating/EV literature, suggest EV-miR-223-5p as a candidate marker of the progressive phenotype and EV-miR-155-5p as a disease indicator. Given the small cohort and near-detection-limit measurements, the study is hypothesis-generating and requires validation in larger, longitudinal cohorts.

Background: Environmental factors are known to influence the clinical presentation of patients with multiple sclerosis. This study aims to compare the demographic and clinical characteristics of multiple sclerosis patients treated at two diverse geographical settings. Methods: A cross-sectional, observational cohort study was conducted in two MS centers: the Danish Multiple Sclerosis Center (DMSC) in Copenhagen, Denmark and the Regional MS Center in Târgu Mureș, Romania. We compared patients’ demographic and clinical characteristics between MS centers, including sex distribution, current age, MS onset age, latest EDSS scores, symptomatology at disease onset, MS phenotype and type of ongoing DMT. Results: In both cohorts, sex distribution was similar, with females constituting 69.2% in DMSC, and 65.7% in Târgu Mureș. Pyramidal symptoms at MS onset were predominant among Targu Mures patients (32.7%), while sensory symptoms were more frequent among DMSC patients (33%). Progressive forms of MS were more prevalent in Târgu Mureș (22.6%) compared to DMSC (9.9%). High-efficacy DMTs were on use by 58.3% patients in DMSC and only by 29.4% patients in Târgu Mureș, who were mostly on low-efficacy DMTs (54.4% vs. 12.4% in DMSC). Conclusions: The study highlights both shared and distinct characteristics of MS patients treated in these two centers. These findings underscore the importance of regional considerations in the management and treatment of MS.

Circulating extracellular vesicle-associated microRNAs (EV-miRNAs) are emerging as promising peripheral biomarkers in multiple sclerosis (MS). This prospective, observational pilot study was conceived as a hypothesis-generating investigation to characterize the expression profile of four candidate EV-miRNAs (miR-223-5p, miR-155-5p, miR-30a-5p, and miR-146a-5p) within an EV-enriched plasma fraction. The cohort comprised 16 patients with secondary progressive MS (SPMS) undergoing siponimod therapy, 13 age- and sex-matched healthy controls (HCs), and 7 patients with relapsing–remitting MS (RRMS) included as an exploratory comparator. Quantification was performed by quantitative real-time PCR employing the ΔΔC_t methodology, with miR-16-5p as the endogenous normalizer. Analyses were conducted cross-sectionally and longitudinally, the latter within a paired subgroup of 11 SPMS patients evaluated at baseline and after twelve months of uninterrupted treatment. Cross-sectional comparisons demonstrated a significant downregulation of EV-miR-223-5p in SPMS patients relative to HCs (fold-change [FC] = 0.26; FDR q = 0.026), whereas EV-miR-155-5p was significantly reduced in both the SPMS (FC = 0.35; FDR q = 0.033) and RRMS (FC = 0.28; FDR q = 0.046) cohorts compared with HCs. No significant intergroup differences were observed for EV-miR-30a-5p or EV-miR-146a-5p. Longitudinal assessment revealed no significant modulation of any target EV-miRNA following one year of siponimod therapy. These preliminary observations should be interpreted with caution, given the exploratory nature and modest cohort size. Importantly, the isolation of total plasma EVs does not permit resolution of the specific cellular provenance of the observed signals, nor does it capture their downstream functional consequences. Nevertheless, the selective downregulation of EV-miR-223-5p and EV-miR-155-5p may tentatively suggest candidate molecular signatures warranting further interrogation. Adequately powered studies incorporating cell-specific EV sorting and paired cerebrospinal fluid sampling will be required to substantiate these signals and clarify their potential utility in monitoring disease progression and therapeutic response in progressive MS.

Multiple sclerosis (MS) is an incurable disease of the central nervous system that affects young adults and determines significant disability. The past decade has seen a change in the treatment options, moving toward disease-modifying therapies. cladribine (Cladribine) tablets are the only short-course oral immune reconstitution therapy approved for highly active relapsing MS (RMS). Romanian authorities approved the full reimbursement in July 2021. We conducted a multicenter, observational, post-marketing, real-world cohort study in Romania that included all patients with MS (PwMS) who completed at least one cycle of CLD with at least three months of follow-up as of September 2024. Demographic variables and MS clinical characteristics regarding each patient were recorded. No Evidence of Disease Activity (NEDA-3) was assessed at one and two years after CLD initiation. The primary outcome was the proportion and characteristics of PwMS achieving NEDA-3 at 12 and 24 months after CLD initiation and the identification of predictors of failure in achieving NEDA-3. A total of 348 PwMS cases were included, of which CLD was the first DMT option in 129 (37.1%). At one year, 76.74% of the patients with complete available data fulfilled all criteria, achieving NEDA-3 status, with similar proportions between previously DMT-treated PwMS and naïve PwMS. In the univariate analysis, the onset of long-tract involvement, as compared to infratentorial (OR = 6.32, p = 0.01), and the number of Gd-enhancing lesions (OR = 1.12, p = 0.02), were significantly associated with increased odds of not achieving NEDA-3 at one year. In the multivariate model, long tracts (OR = 10.28, p = 0.006), higher baseline EDSS (OR = 1.39, p = 0.01), and longer disease duration (OR = 0.92, p = 0.04) were independent predictors. The initiation of CLD treatment was associated with achieving NEDA-3 at one year in >70% of cases. CLD demonstrated an improvement in MS control, irrespective of the patients' demographics, RMS history, or prior DMT exposure. CLD treatment has a favorable safety profile and a high level of patient adherence.