Mitochondrial Biomarkers in the Omics Era: A Clinical-Pathophysiological Perspective.

Mitochondrial diseases (MDs) affect 4300 individuals, with different ages of presentation and manifestation in any organ. How defects in mitochondria can cause such a diverse range of human diseases remains poorly understood. In recent years, several published research articles regarding the metabolic and protein profiles of these neurogenetic disorders have helped shed light on the pathogenetic mechanisms. By investigating different pathways in MDs, often with the aim of identifying disease biomarkers, it is possible to identify molecular processes underlying the disease. In this perspective, omics technologies such as proteomics and metabolomics considered in this review, can support unresolved mitochondrial questions, helping to improve outcomes for patients.

Deoxyguanosine kinase deficiency: natural history and liver transplant outcome.

Autosomal recessive pathogenetic variants in the DGUOK gene cause deficiency of deoxyguanosine kinase activity and mitochondrial deoxynucleotides pool imbalance, consequently, leading to quantitative and/or qualitative impairment of mitochondrial DNA synthesis. Typically, patients present early-onset liver failure with or without neurological involvement and a clinical course rapidly progressing to death. This is an international multicentre study aiming to provide a retrospective natural history of deoxyguanosine kinase deficient patients. A systematic literature review from January 2001 to June 2023 was conducted. Physicians of research centres or clinicians all around the world caring for previously reported patients were contacted to provide followup information or additional clinical, biochemical, histological/histochemical, and molecular genetics data for unreported cases with a confirmed molecular diagnosis of deoxyguanosine kinase deficiency. A cohort of 202 genetically confirmed patients, 36 unreported, and 166 from a systematic literature review, were analyzed. Patients had a neonatal onset (≤ 1 month) in 55.7% of cases, infantile (>1 month and ≤ 1 year) in 32.3%, pediatric (>1 year and ≤18 years) in 2.5% and adult (>18 years) in 9.5%. Kaplan-Meier analysis showed statistically different survival rates (P < 0.0001) among the four age groups with the highest mortality for neonatal onset. Based on the clinical phenotype, we defined four different clinical subtypes: hepatocerebral (58.8%), isolated hepatopathy (21.9%), hepatomyoencephalopathy (9.6%), and isolated myopathy (9.6%). Muscle involvement was predominant in adult-onset cases whereas liver dysfunction causes morbidity and mortality in early-onset patients with a median survival of less than 1 year. No genotype-phenotype correlation was identified. Liver transplant significantly modified the survival rate in 26 treated patients when compared with untreated. Only six patients had additional mild neurological signs after liver transplant. In conclusion, deoxyguanosine kinase deficiency is a disease spectrum with a prevalent liver and brain tissue specificity in neonatal and infantile-onset patients and muscle tissue specificity in adult-onset cases. Our study provides clinical, molecular genetics and biochemical data for early diagnosis, clinical trial planning and immediate intervention with liver transplant and/or nucleoside supplementation.

Serum neurofilament light chain levels correlate with small fiber related parameters in patients with hereditary transthyretin amyloidosis with polyneuropathy (ATTRv-PN).

BACKGROUND: Recent evidence suggests that both serum neurofilament light chain (sNfL) levels and small fiber related diagnostic variables may be valuable disease biomarkers of hereditary transthyretin amyloidosis with polyneuropathy (ATTRv-PN). Our study aimed to explore the relations between sNfL and small fiber related skin biopsy and quantitative sensory testing (QST) parameters in a cohort of ATTRv-PN patients and pre-symptomatic carriers. METHODS: We retrospectively analyzed data from 13 ATTRv patients and 21 pre-symptomatic carriers who underwent sNfL dosage, skin biopsy, and QST, and analyzed correlations between sNFL, intraepidermal nerve fiber density (IENFD), and cold (CDT) and warm detection thresholds (WDT). RESULTS: Both sNfL and small fiber related parameters significantly differed between carriers and patients (sNfL: p < 0.0001; IENFD: p = 0.0008; CDT, WDT: < 0.0001). sNFL levels were normal in all carriers, altered in 85% of patients, negatively correlated with distal IENFD (r = -0.47, p = 0.005), and significantly correlated with CDT (r = -0.68; p < 0.0001) and WDT (r = 0.57; p < 0.0001). CONCLUSIONS: Our study showed that sNfL reliably discriminates symptomatic ATTRv-PN patients from pre-symptomatic carriers, and found significant relations between sNfL, skin biopsy, and QST small fiber related parameters, suggesting that sNfL might be a valuable biomarker of peripheral nerve involvement in ATTRv-PN and a supportive criterion for symptomatic disease transition.

Hospital admissions from the emergency department of adult patients affected by myopathies.

BACKGROUND AND PURPOSE: Myopathies are associated with classic signs and symptoms, but also with possible life-threatening complications that may require assistance in an emergency setting. This phenomenon is understudied in the literature. We aimed to assess the presentation, management, and outcomes of clinical manifestations potentially related to a muscle disorder requiring referral to the adult emergency department (ED) and hospitalization. METHODS: Anonymized patient data retrieved using the International Classification of Diseases, Ninth Revision codes related to muscle disorders over 4 years were retrospectively analyzed. Medical reports were evaluated to extract demographic and clinical variables, along with outcomes. Two groups were defined based on the presence (known diagnosis [KD] group) or absence (unknown diagnosis [UD] group) of a diagnosed muscle disorder at arrival. RESULTS: A total of 244 patients were included, 51% of whom were affected by a known myopathy, predominantly limb-girdle muscular dystrophies and myotonic dystrophies. The main reasons for ED visits in the KD group were respiratory issues, worsening of muscle weakness, and gastrointestinal problems. Heart complications were less prevalent. In the UD group, 27 patients received a new diagnosis of a specific primary muscle disorder after the ED access, mostly an inflammatory myopathy. Death during hospitalization was recorded in 26 patients, with a higher rate in the KD group and in patients affected by mitochondrial and inflammatory myopathies. Sepsis and dyspnea were associated with increased death risk. CONCLUSIONS: Respiratory complications are the most common reason for myopathic patients accessing the ED, followed by gastrointestinal issues. Infections are severe threats and, once hospitalized, these patients have relatively high mortality.

Serum Neurofilament and Free Light Chain Levels in Patients Undergoing Treatment for Chronic Inflammatory Demyelinating Polyneuropathy.

Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is an immune-mediated disorder affecting the peripheral nervous system. Despite the established diagnostic criteria, monitoring disease activity and treatment remains challenging. To address this limitation, we investigated serum neurofilament light chain (sNfL) and serum free light chains (sFLCs) as potential biomarkers. A total of 32 CIDP patients undergoing immunoglobulin therapy and 32 healthy controls enrolled in the present study, and agreed to have their blood plasma sNfL and sFLCs analyzed, while CIDP severity was assessed through the modified Rankin Scale (mRS) and the Overall Neuropathy Limitations Scale (ONLS). In line with the immunoglobulin treatment aimed at limiting neuronal damage administered to the majority of patients, sNfL levels did not exhibit significant differences between the two groups. However, CIDP patients showed significantly elevated sFLC and sFLC ratios, while the marker levels did not correlate with the clinical scores. The study confirms the potential of sFLCs as a sensitive biomarker of inflammatory processes in CIDP. Additionally, the present study results regarding neurofilaments strengthen the role of sNfL in monitoring CIDP treatments, confirming the effectiveness of immunoglobulin therapy. Overall, our results demonstrate how combining these markers can lead to better patient characterization for improved treatment.

Characterization of D4Z4 alleles and assessment of de novo cases in Facioscapulohumeral dystrophy (FSHD) in a cohort of Italian families.

Facioscapulohumeral dystrophy (FSHD) is an autosomal dominant disease, although 10%-30% of cases are sporadic. However, this percentage may include truly de novo patients (carrying a reduced D4Z4 allele that is not present in either of the parents) and patients with apparently sporadic disease resulting from mosaicism, non-penetrance, or complex genetic situations in either patients or parents. In this study, we characterized the D4Z4 Reduced Alleles (DRA) and evaluated the frequency of truly de novo cases in FSHD1 in a cohort of DNA samples received consecutively for FSHD-diagnostic from 100 Italian families. The D4Z4 testing revealed that 60 families reported a DRA compatible with FSHD1 (1-10 RU). The DRA co-segregated with the disease in most cases. Five families with truly de novo cases were identified, suggesting that this condition may be slightly lower (8%) than previously reported. In addition, D4Z4 characterization in the investigated families showed 4% of mosaic cases and 2% with translocations. This study further highlighted the importance of performing family studies for clarifying apparently sporadic FSHD cases, with significant implications for genetic counseling, diagnosis, clinical management, and procreative choices for patients and families.

Serum neurofilament light chain: a promising early diagnostic biomarker for hereditary transthyretin amyloidosis?

BACKGROUND AND PURPOSE: Hereditary transthyretin amyloidosis (ATTRv) is a life-threatening disease caused by mutations in the gene encoding transthyretin (TTR). The recent therapeutic advances have underlined the importance of easily accessible, objective biomarkers of both disease onset and progression. Preliminary evidence suggests a potential role in this respect for neurofilament light chain (NfL). In this study, the aim was to determine serum NfL (sNfL) levels in a late-onset ATTRv population and evaluate whether it might represent a reliable biomarker of disease onset (i.e., 'conversion' from the asymptomatic status to symptomatic disease in TTR mutation carriers). METHODS: In all, 111 individuals harbouring a pathogenic TTR variant (61 symptomatic ATTRv patients and 50 presymptomatic carriers) were consecutively enrolled. Fifty healthy volunteers were included as the control group. Ella™ apparatus was used to assess sNfL levels. RESULTS: Serum NfL levels were increased in ATTRv patients compared to both presymptomatic carriers and healthy controls, whilst not differing between carriers and healthy controls. An sNfL cut-off of 37.10 pg/mL could discriminate between asymptomatic and symptomatic individuals with high diagnostic accuracy (area under the curve 0.958; p < 0.001), sensitivity (81.4%) and specificity (100%). CONCLUSIONS: Serum NfL seems to be a promising biomarker of peripheral nerve involvement in ATTRv amyloidosis and might become a reliable, objective measure to detect the transition from the presymptomatic stage to the onset of symptomatic disease. Further longitudinal studies are needed to confirm such a role and determine whether it could equally represent a biomarker of disease progression and response to therapy.

Current Evidence Supporting the Role of Immune Response in ATTRv Amyloidosis.

Hereditary transthyretin (ATTRv) amyloidosis with polyneuropathy, also known as familial amyloid polyneuropathy (FAP), represents a progressive, heterogeneous, severe, and multisystemic disease caused by pathogenic variants in the TTR gene. This autosomal-dominant neurogenetic disorder has an adult onset with variable penetrance and an inconstant phenotype, even among subjects carrying the same mutation. Historically, ATTRv amyloidosis has been viewed as a non-inflammatory disease, mainly due to the absence of any mononuclear cell infiltration in ex vivo tissues; nevertheless, a role of inflammation in its pathogenesis has been recently highlighted. The immune response may be involved in the development and progression of the disease. Fibrillary TTR species bind to the receptor for advanced glycation end products (RAGE), probably activating the nuclear factor κB (NF-κB) pathway. Moreover, peripheral blood levels of several cytokines, including interferon (IFN)-gamma, IFN-alpha, IL-6, IL-7, and IL-33, are altered in the course of the disease. This review summarizes the current evidence supporting the role of the immune response in ATTRv amyloidosis, from the pathological mechanisms to the possible therapeutic implications.

Hearing Impairment and Neuroimaging Results in Mitochondrial Diseases.

Mitochondrial diseases (MDs) are heterogeneous genetic disorders characterized by mitochondrial DNA (mtDNA) defects, involving tissues highly dependent on oxidative metabolism: the inner ear, brain, eye, skeletal muscle, and heart. We describe adult patients with genetically defined MDs, characterizing hearing function and neuroimaging results. We enrolled 34 patients (mean age: 50.02 ± 15 years, range: 18-75 years; 20 females and 14 males) classified in four groups: MELAS, MIDD, PEO, and Encephalopathy/Polyneuropathy. Audiological evaluations included psychoacoustical tests (pure-tone and speech audiometry), electrophysiological tests (Auditory Brainstem Responses, ABRs), and Impedenzometry. Neuroimaging evaluations considered global MRI abnormalities or structural brain changes. In total, 19/34 patients carried the m.3243A > G mutation (6 affected by MELAS, 12 affected by MIDD, and 1 affected by PEO); 11 had an mtDNA deletion (all affected by PEO); 3 had nuclear genes associated with MDs (POLG1 and OPA1); and 1 patient had an mtDNA deletion without an identified nuclear gene defect (affected by PEO). Sensory neural, bilateral, and symmetrical hearing loss was present in 25 patients (73.5%) to different degrees: 9 mild, 9 moderate, 5 severe, and 2 profound. The severe/profound and mild hearing losses were associated with pantonal and high-frequency audiograms, respectively. Instead, moderate hearing losses were associated with both high-frequency (five cases) and pantonal (five cases) audiogram shapes. In addition, 21/25 patients showed a cochlear site of lesion (84%), and 4/25 (16%) showed a retrocochlear site. We found global MRI abnormalities or structural brain changes in 26/30 subjects (86.6%): 21 had white matter abnormalities, 15 had cortical atrophy, 10 had subcortical atrophy, 8 had basal nuclei involvement or cerebellar atrophy, 4 had stroke-like lesions or laminar necrosis, and 1 had cysts or vacuolated lesions. We concluded that genetic alterations are associated with different clinical presentations for both auditory function and neuroradiological findings. There is no fixed relationship between genotype and phenotype for the clinical conditions analyzed.

Quantitative sensory testing and skin biopsy findings in late-onset ATTRv presymptomatic carriers: Relationships with predicted time of disease onset (PADO).

INTRODUCTION: Hereditary transthyretin amyloidosis polyneuropathy (ATTRv-PN) presymptomatic carriers often show preclinical abnormalities at small fiber-related diagnostic tests. However, no validated biomarker is currently available to use for presymptomatic carriers' follow-up, thus helping therapeutic decision making. Our study aimed at assessing nerve conduction study (NCS), quantitative sensory testing (QST), and skin biopsy parameters in a large cohort of late-onset ATTRv presymptomatic carriers and to evaluate whether they correlated with predicted age of disease onset (PADO). METHODS: Late-onset ATTRv presymptomatic carriers were consecutively enrolled and underwent NCS, QST, and skin biopsy with intraepidermal nerve fiber density (IENFD) evaluation from a distal and a proximal site. Douleur Neuropathique-4 (DN4) and Small Fiber Neuropathy-Symptoms Inventory (SFN-SIQ) were used to assess painful and small fiber neuropathy-related symptoms. PADO and time-to-PADO (delta-PADO) were estimated for each carrier, and correlations with diagnostic test measures were analyzed. RESULTS: Forty presymptomatic ATTRv subjects were enrolled. Twenty carriers (50%) had distal IENFD reduction, with a non-length-dependent distribution in 73% of cases. Eleven subjects (27.5%) had cold and/or warm detection threshold (CDT and/or WDT) abnormalities at QST. Delta-PADO positively correlated with sural sensory nerve action potential (SNAP) amplitude (r = .416, p = .004), and z-values of QST parameters like CDT (r = .314, p = .028), WDT (r = -.294, p = .034), and mechanical detection threshold (MDT; r = -.382, p = .012). Simple linear regression models showed a linear relation between delta-PADO and sural SAP, CDT, and MDT. CONCLUSIONS: Our findings confirm that IENFD reduction and QST abnormalities may occur early in ATTRv presymptomatic carriers, often with a non-length-dependent pattern. However, only sural SAP amplitude and QST parameters correlated with delta-PADO, suggesting that serial combined QST and NCS evaluation could be useful in ATTRv presymptomatic carriers' follow-up.

Digital health and Clinical Patient Management System (CPMS) platform utility for data sharing of neuromuscular patients: the Italian EURO-NMD experience.

BACKGROUND: The development of e-health technologies for teleconsultation and exchange of knowledge is one of the core purposes of European Reference Networks (ERNs), including the ERN EURO-NMD for rare neuromuscular diseases. Within ERNs, the Clinical Patient Management System (CPMS) is a web-based platform that seeks to boost active collaboration within and across the network, implementing data sharing. Through CPMS, it is possible to both discuss patient cases and to make patients' data available for registries and databases in a secure way. In this view, CPMS may be considered a sort of a temporary storage for patients' data and an effective tool for data sharing; it facilitates specialists' consultation since rare diseases (RDs) require multidisciplinary skills, specific, and outstanding clinical experience. Following European Union (EU) recommendation, and to promote the use of CPMS platform among EURO-NMD members, a twelve-month pilot project was set up to train the 15 Italian Health Care Providers (HCPs). In this paper, we report the structure, methods, and results of the teaching course, showing that tailored, ERN-oriented, training can significantly enhance the profitable use of the CPMS. RESULTS: Throughout the training course, 45 professionals learned how to use the many features of the CPMS, eventually opening 98 panels of discussion-amounting to 82% of the total panels included in the EURO-NMD. Since clinical, genetic, diagnostic, and therapeutic data of patients can be securely stored within the platform, we also highlight the importance of this platform as an effective tool to discuss and share clinical cases, in order to ease both case solving and data storing. CONCLUSIONS: In this paper, we discuss how similar course could help implementing the use of the platform, highlighting strengths and weaknesses of e-health for ERNs. The expected result is the creation of a "map" of neuromuscular patients across Europe that might be improved by a wider use of CPMS.

Clinical, neurophysiological and serological clues for the diagnosis of neuromyotonia and distinction from cramp-fasciculation syndrome.

Neuromyotonia and cramp-fasciculation syndrome diagnosis currently relies on neurophysiological examination. In this study we investigated the clinical features and neural antibody profile of patients with neuromyotonia and cramp-fasciculation syndrome to assess the diagnostic value of serological testing. Available sera from adult patients with electromyography-defined neuromyotonia and cramp-fasciculation syndrome were tested for neural antibodies by indirect immunofluorescence on mouse brain sections and live cell-based assays. Forty patients were included, 14 with neuromyotonia and 26 with cramp-fasciculation syndrome. Neural antibodies were detected in 10/10 neuromyotonia sera, most commonly against contactin-associated protein 2 (7/10, 70%), and in 1/20 (5%) cramp-fasciculation syndrome sera. Clinical myokymia, hyperhidrosis, and paresthesia or neuropathic pain were more common in neuromyotonia and mostly associated with contactin-associated protein 2 antibodies. Central nervous system involvement was present in 4/14 (29%) neuromyotonia patients. A tumor was detected in 13/14 (93%) neuromyotonia patients (thymoma, 13), and in 4/26 (15%) with cramp-fasciculation syndrome (thymoma, 1; other neoplasms, 3). Twenty-one/27 (78%) patients achieved a significant improvement or complete remission. Our findings highlight clinical, neurophysiological and serological clues that can be useful in the diagnosis of neuromyotonia and cramp-fasciculation syndrome. Antibody testing is valuable for neuromyotonia diagnosis, while its usefulness in cramp-fasciculation syndrome confirmation is limited.

A coordinated multiorgan metabolic response contributes to human mitochondrial myopathy.

Mitochondrial diseases are a heterogeneous group of monogenic disorders that result from impaired oxidative phosphorylation (OXPHOS). As neuromuscular tissues are highly energy-dependent, mitochondrial diseases often affect skeletal muscle. Although genetic and bioenergetic causes of OXPHOS impairment in human mitochondrial myopathies are well established, there is a limited understanding of metabolic drivers of muscle degeneration. This knowledge gap contributes to the lack of effective treatments for these disorders. Here, we discovered fundamental muscle metabolic remodeling mechanisms shared by mitochondrial disease patients and a mouse model of mitochondrial myopathy. This metabolic remodeling is triggered by a starvation-like response that evokes accelerated oxidation of amino acids through a truncated Krebs cycle. While initially adaptive, this response evolves in an integrated multiorgan catabolic signaling, lipid store mobilization, and intramuscular lipid accumulation. We show that this multiorgan feed-forward metabolic response involves leptin and glucocorticoid signaling. This study elucidates systemic metabolic dyshomeostasis mechanisms that underlie human mitochondrial myopathies and identifies potential new targets for metabolic intervention.

Factors associated with the severity of COVID-19 outcomes in people with neuromuscular diseases: Data from the International Neuromuscular COVID-19 Registry.

BACKGROUND AND PURPOSE: Clinical outcome information on patients with neuromuscular diseases (NMDs) who have been infected with SARS-CoV-2 is limited. The aim of this study was to determine factors associated with the severity of COVID-19 outcomes in people with NMDs. METHODS: Cases of NMD, of any age, and confirmed/presumptive COVID-19, submitted to the International Neuromuscular COVID-19 Registry up to 31 December 2021, were included. A mutually exclusive ordinal COVID-19 severity scale was defined as follows: (1) no hospitalization; (2) hospitalization without oxygenation; (3) hospitalization with ventilation/oxygenation; and (4) death. Multivariable ordinal logistic regression analyses were used to estimate odds ratios (ORs) for severe outcome, adjusting for age, sex, race/ethnicity, NMD, comorbidities, baseline functional status (modified Rankin scale [mRS]), use of immunosuppressive/immunomodulatory medication, and pandemic calendar period. RESULTS: Of 315 patients from 13 countries (mean age 50.3 [±17.7] years, 154 [48.9%] female), 175 (55.5%) were not hospitalized, 27 (8.6%) were hospitalized without supplemental oxygen, 91 (28.9%) were hospitalized with ventilation/supplemental oxygen, and 22 (7%) died. Higher odds of severe COVID-19 outcomes were observed for: age ≥50 years (50-64 years: OR 2.4, 95% confidence interval [CI] 1.33-4.31; >64 years: OR 4.16, 95% CI 2.12-8.15; both vs. <50 years); non-White race/ethnicity (OR 1.81, 95% CI 1.07-3.06; vs. White); mRS moderately severe/severe disability (OR 3.02, 95% CI 1.6-5.69; vs. no/slight/moderate disability); history of respiratory dysfunction (OR 3.16, 95% CI 1.79-5.58); obesity (OR 2.24, 95% CI 1.18-4.25); ≥3 comorbidities (OR 3.2, 95% CI 1.76-5.83; vs. ≤2; if comorbidity count used instead of specific comorbidities); glucocorticoid treatment (OR 2.33, 95% CI 1.14-4.78); and Guillain-Barré syndrome (OR 3.1, 95% CI 1.35-7.13; vs. mitochondrial disease). CONCLUSIONS: Among people with NMDs, there is a differential risk of COVID-19 outcomes according to demographic and clinical characteristics. These findings could be used to develop tailored management strategies and evidence-based recommendations for NMD patients.

Telemedicine in Neuromuscular Diseases During Covid-19 Pandemic: ERN-NMD European Survey.

BACKGROUND: Telemedicine (TM) contributes to bridge the gap between healthcare facilities and patients' homes with neuromuscular disease (NMD) because of mobility issues. However, its deployment is limited due to difficulties evaluating subtle neurological signs such as mild weakness or sensory deficits. The COVID-19 pandemic has disrupted healthcare delivery worldwide, necessitating rapid measures implementation by health care providers (HCPs) to protect patients from acquiring SARS-CoV-2 while maintaining the best care and treatment. OBJECTIVES: Given the challenges faced by remote healthcare assistance of NMD patients, we aim to evaluate the use of TM in NMD during the COVID-19 pandemic. METHODS: Based on the Model for Assessment-of-Telemedicine-Applications (MAST), we conducted a survey amongst clinicians of the ERN EURO NMD (European-Reference-Network-for-Rare-Neuromuscular-Diseases). RESULTS: Based on 42 responses over 76 expected ones, our results show that the COVID-19 pandemic significantly increased the number of HCPs using TM (from 60% to 100%). The TM types most used during the COVID-19 period are teleconsultation and consultation by phone, particularly in the context of symptoms worsening in NMD patients with COVID-19 infection. Most European HCPs were satisfied when using TM but as a complementary option to physical consultations. Many responses addressed the issue of technical aspects needing improvement, particularly for elderly patients who need caregivers' assistance for accessing the TM platform. CONCLUSIONS: TM has been essential during COVID-19, but its use still presents some limitations for NMD patients with cognitive deficits or for first-time diagnosis. Thus, TM should be used as complement to, rather than substitute, for face-to-face consultations.