Pathological findings with vacuoles in anti-mitochondrial antibody-positive inflammatory myopathy.

BACKGROUND: A few patients with inflammatory myopathy showed anti-mitochondrial antibody (AMA) positivity. This study aimed to report the clinical and pathological findings with vacuoles in 3 cases of such patients. METHODS: Three cases with myositis from the Myositis Clinical Database of Peking University First Hospital were identified with AMA positivity. Their clinical records were retrospectively reviewed and the data was extracted. All the 3 cases underwent muscle biopsy. RESULTS: Three middle-aged patients presented with chronic-onset weakness of proximal limbs, marked elevation of creatine kinase, and AMA-positivity. Two of the 3 cases meet the criteria of primary biliary cholangitis. All the 3 cases presented with cardiac involvement and proteinuria. Two cases developed type 2 respiratory failure. MRI of the thigh muscle showed multiple patches of edema bilaterally in both cases, mostly in the adductor magnus. Pathological findings include degeneration of muscle fibers, diffused MHC-I positivity, and complement deposits on cell membranes. Vacuoles without rims of different sizes were discovered under the membrane of the muscle fibers. A few RBFs were discovered in case 1, while a diffused proliferation of endomysium and perimysium was shown in case 2. CONCLUSIONS: AMA-positive inflammatory myopathy is a disease that could affect multiple systems. Apart from inflammatory changes, the pathological findings of muscle can also present vacuoles.

Advances in lymphatic metastasis of non-small cell lung cancer.

Lung cancer is a deeply malignant tumor with high incidence and mortality. Despite the rapid development of diagnosis and treatment technology, abundant patients with lung cancer are still inevitably faced with recurrence and metastasis, contributing to death. Lymphatic metastasis is the first step of distant metastasis and an important prognostic indicator of non-small cell lung cancer. Tumor-induced lymphangiogenesis is involved in the construction of the tumor microenvironment, except promoting malignant proliferation and metastasis of tumor cells, it also plays a crucial role in individual response to treatment, especially immunotherapy. Thus, this article reviews the current research status of lymphatic metastasis in non-small cell lung cancer, in order to provide some insights for the basic research and clinical and translational application in this field.

COVID-19 in patients with myasthenia gravis: a single-center retrospective study in China.

BACKGROUND: Coronavirus disease 2019 (COVID-19) has been a great concern since 2019. Patients with myasthenia gravis (MG) may be at higher risk of COVID-19 and a more severe disease course. We examined the associations between COVID-19 and MG. METHODS: This single-center retrospective cohort study involved 134 patients who were diagnosed with MG from June 2020 to November 2022 and followed up until April 2023. They were divided into a COVID-19 group and non-COVID-19 group. Logistic regression analysis was used to detect factors potentially associating COVID-19 with MG. RESULTS: Of the 134 patients with MG, 108 (80.6%) had COVID-19. A higher number of comorbidities was significantly associated with an increased risk of COVID-19 (p = 0.040). A total of 103 patients (95.4%) had mild/moderate COVID-19 symptoms, and 4 patients (3.7%) were severe/critical symptoms (including 2 deaths). Higher age (p = 0.036), use of rituximab (p = 0.037), tumors other than thymoma (p = 0.031), Hashimoto's thyroiditis (p = 0.011), more comorbidities (p = 0.002), and a higher baseline MG activities of daily living (MG-ADL) score (p = 0.006) were risk factors for severe COVID-19 symptoms. The MG-ADL score increased by ≥ 2 points in 16 (15.7%) patients. Dry cough and/or expectoration (p = 0.011), use of oral corticosteroids (p = 0.033), and use of more than one kind of immunosuppressant (p = 0.017) were associated with the increase of the post-COVID-19 MG-ADL score. CONCLUSION: Most patients with MG have a mild course of COVID-19. However, patients with older age, many comorbidities, a high MG-ADL score, and use of a variety of immunosuppressants during COVID-19 may be more prone to severe symptoms.

A novel deep intronic variant introduce dystrophin pseudoexon in Becker muscular dystrophy: A case report.

Most pathogenic DMD variants are detectable and interpretable by standard genetic testing for dystrophinopthies. However, approximately 1∼3% of dystrophinopthies patients still do not have a detectable DMD variant after standard genetic testing, most likely due to structural chromosome rearrangements and/or deep intronic pseudoexon-activating variants. Here, we report on a boy with a suspected diagnosis of Becker muscular dystrophy (BMD) who remained without a detectable DMD variant after exonic DNA-based standard genetic testing. Dystrophin mRNA studies and genomic Sanger sequencing were performed in the boy, followed by in silico splicing analyses. We successfully detected a novel deep intronic disease-causing variant in the DMD gene (c.2380 + 3317A > T), which consequently resulting in a new dystrophin pseudoexon activation through the enhancement of a cryptic donor splice site. The patient was therefore genetically diagnosed with BMD. Our case report further emphasizes the significant role of disease-causing splicing variants within deep intronic regions in genetically undiagnosed dystrophinopathies.

Clinical and genetic interpretation of uncertain DMD missense variants: evidence from mRNA and protein studies.

BACKGROUND: Pathogenic missense variants in the dystrophin (DMD) gene are rarely reported in dystrophinopathies. Most DMD missense variants are of uncertain significance and their pathogenicity interpretation remains complicated. We aimed to investigate whether DMD missense variants would cause aberrant splicing and re-interpret their pathogenicity based on mRNA and protein studies. METHODS: Nine unrelated patients who had an elevated serum creatine kinase level with or without muscle weakness were enrolled. They underwent a detailed clinical, imaging, and pathological assessment. Routine genetic testing and muscle-derived mRNA and protein studies of dystrophin and sarcoglycan genes were performed in them. RESULTS: Three of the 9 patients presented with a Duchenne muscular dystrophy (DMD) phenotype and the remaining 6 patients had a suspected diagnosis of Becker muscular dystrophy (BMD) or sarcoglycanopathy based on their clinical and pathological characteristics. Routine genetic testing detected only 9 predicted DMD missense variants in them, of which 6 were novel and interpreted as uncertain significance. Muscle-derived mRNA studies of sarcoglycan genes didn't reveal any aberrant transcripts in them. Dystrophin mRNA studies confirmed that 3 predicted DMD missense variants (c.2380G > C, c.4977C > G, and c.5444A > G) were in fact splicing and frameshift variants due to aberrant splicing. The 9 DMD variants were re-interpreted as pathogenic or likely pathogenic based on mRNA and protein studies. Therefore, 3 patients with DMD splicing variants and 6 patients with confirmed DMD missense variants were diagnosed with DMD and BMD, respectively. CONCLUSION: Our study highlights the importance of muscle biopsy and aberrant splicing for clinical and genetic interpretation of uncertain DMD missense variants.

Encephalitis-like episodes with cortical edema and enhancement in patients with neuronal intranuclear inclusion disease.

OBJECTIVES: Neuronal intranuclear inclusion disease (NIID) exhibited significant clinical heterogeneities. However, the clinical features, radiographic changes, and prognosis of patients with encephalitis-like NIID have yet to be systematically elucidated. METHODS: Clinical data including medical history, physical examination, and laboratory examinations were collected and analyzed. Skin and sural nerve biopsies were conducted on the patient. Repeat-primed PCR (RP-PCR) and fluorescence amplicon length PCR (AL-PCR) were used to detect the expansion of CGG repeat. We also reviewed the clinical and genetic data of NIID patients with cortical enhancement. RESULTS: A 54-year-old woman presented with encephalitis-like NIID, characterized by severe headache and agitative psychiatric symptoms. The brain MRI showed cortical swelling in the temporo-occipital lobes and significant enhancement of the cortical surface and dura, but without hyperintensities along the corticomedullary junction on diffusion-weighted image (DWI). A biopsy of the sural nerve revealed a demyelinating pathological change. The intranuclear inclusions were detected in nerve and skin tissues using the p62 antibody and electron microscopy. RP-PCR and AL-PCR unveiled the pathogenic expansion of CGG repeats in the NOTCH2NLC gene. A review of the literature indicated that nine out of the 16 patients with cortical lesions and linear enhancement exhibited encephalitis-like NIID. CONCLUSION: This study indicated that patients with encephalitis-like NIID typically exhibited headache and excitatory psychiatric symptoms, often accompanied by cortical edema and enhancement of posterior lobes, and responded well to glucocorticoid treatment. Furthermore, some patients may not exhibit hyperintensities along the corticomedullary junction on DWI, potentially leading to misdiagnosis.

Associations of sleep-related variables with reverse dipping patterns of blood pressure in α-synucleinopathies.

INTRODUCTION: The reverse dipping blood pressure (BP) pattern is very common in α-synucleinopathies. We aimed to explore the associations of sleep-related variables with abnormal BP circadian rhythms in Parkinson's disease (PD) and multiple system atrophy (MSA). METHODS: A total of 126 patients, 76 with PD and 50 with MSA, were included. All participants underwent ambulatory BP monitoring and full-night polysomnography (PSG). We analyzed abnormal dipping patterns and sleep-related parameters, including moderate to severe obstructive sleep apnea (OSA), rapid eye movement behavior disorder (RBD), average oxygen saturation (SaO2%), lowest SaO2%, duration of SaO2% <90%, and apnea-hypopnea index (AHI). Binary logistic regression was performed to explore the associations between paraclinical variables, sleep-related variables, and reverse dipping patterns. RESULTS: Reverse dipping patterns were predominant in patients with PD (58.5 %) and MSA (68.0 %). Patients with MSA had higher AHI, RBD, and lower average SaO2% than those with PD. Taking both diseases together as a whole group of α-synucleinopathies, logistic regression analysis indicates the Hoehn-Yahr stage (odds ratio [OR] = 2.00 for reverse systolic and 2.34 for reverse diastolic dipping patterns), moderate to severe OSA (OR = 2.71 for reverse systolic and 2.53 for reverse diastolic dipping patterns), average SaO2% (OR = 1.35 for reverse systolic dipping patterns), and male sex (OR = 2.70 for reverse diastolic dipping patterns) were independently associated with reverse dipping patterns. CONCLUSIONS: Reverse dipping patterns were common in patients with PD and MSA. Hoehn-Yahr stage, moderate to severe OSA, average SaO2%, and male sex were associated with reverse dipping patterns in α-synucleinopathy.

Decreased retinal vascular density is associated with cognitive impairment in CADASIL: an optical coherence tomography angiography study.

PURPOSE: This study aimed to assess alterations in retinal vascular density in cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) patients using optical coherence tomography angiography (OCTA) and investigate their association with MRI and cognitive features. METHODS: Twenty-five patients with CADASIL and forty healthy controls were evaluated by Cirrus HD-OCT 5000 with AngioPlex OCTA to determine changes in macular retinal vasculature. Retinal vasculature parameters between two groups were compared. The MRI lesion burden and neuropsychological scales were also examined in patients. The association between OCTA parameters and MRI/cognitive features was evaluated using partial Spearman rank correlation. RESULTS: The vessel density and perfusion density of whole image in macular region (vessel density: t = - 2.834, p = 0.005; perfusion density: t = - 2.691, p = 0.007) were significantly decreased in patients with CADASIL. Moreover, vessel density of whole image in macular region was negatively associated with Fazekas scores (ρ = - 0.457; p = 0.025) and the number of lacunar infractions (ρ = - 0.425, p = 0.038) after adjustment for age. Decreased macular vessel density and perfusion density of whole image were also associated with MoCA scores (vessel density: ρ = 0.542, p = 0.006; perfusion density: ρ = 0.478, p = 0.018) and other domain-specific neuropsychological tests (p < 0.05) after adjustment for age. CONCLUSION: Decreased retinal vascular density was associated with increased MRI lesion burden and cognitive impairment in patients with CADASIL. Our findings suggest that the degree of retinal vascular involvement, as demonstrated by OCTA, may be consistent with the severity of MRI lesions and the degree of cognitive impairment in patients.

Enhanced dewaterability of food waste digestate by biochar/potassium ferrate treatments: Performance and mechanisms.

The combined process of biochar (BC) and potassium ferrate (PF) offers a fascinating technique for efficient dewatering of digestate. However, the effects of BC/PF treatment on the dewaterability and mechanisms of FWD are still unknown. This study aimed to reveal the impact mechanisms of BC/PF treatment on digestate dewatering performance. Experimental results indicated that BC/PF treatment significantly enhanced the dewaterability of digestate, with the minimum specific resistance to filtration of (1.05 ± 0.02) × 1015 m·kg-1 and water content of 57.52 ± 0.51% being obtained at the concentrations of 0.018 g·g-1 total solid (TS) BC300 and 0.20 g·g-1 TS PF, which were 8.60% and 13.59% lower than PF treatment, respectively. BC/PF treatment proficiently reduced the fractal dimension, bound water content, apparent viscosity, and gel-like network structure strength of digestate, as well as increased the floc size and zeta potential of digestate. BC/PF treatment promoted the conversion of extracellular polymeric substances (EPS) fractions from inner EPS to soluble EPS, increased the fluorescence intensity of the dissolved compounds, and enhanced the hydrophobicity of proteins. Mechanisms investigations showed that BC/PF enhanced dewatering through non-reactive oxygen species pathways, i.e., via strong oxidative intermediate irons species Fe(V)/Fe(IV). BC/PF treatment enhanced the solubilization of nutrients, the inactivation of fecal coliforms, and the mitigation of heavy metal toxicity. The results suggested that BC/PF treatment is an effective digestate dewatering technology which can provide technological supports to the closed-loop treatment of FWD.

Novel TUBA4A variant causes congenital myopathy with focal myofibrillar disorganisation.

BACKGROUND: Congenital myopathies are a clinical, histopathological and genetic heterogeneous group of inherited muscle disorders that are defined on peculiar architectural abnormalities in the muscle fibres. Although there have been at least 33 different genetic causes of the disease, a significant percentage of congenital myopathies remain genetically unresolved. The present study aimed to report a novel TUBA4A variant in two unrelated Chinese patients with sporadic congenital myopathy. METHODS: A comprehensive strategy combining laser capture microdissection, proteomics and whole-exome sequencing was performed to identify the candidate genes. In addition, the available clinical data, myopathological changes, the findings of electrophysiological examinations and thigh muscle MRIs were also reviewed. A cellular model was established to assess the pathogenicity of the TUBA4A variant. RESULTS: We identified a recurrent novel heterozygous de novo c.679C>T (p.L227F) variant in the TUBA4A (NM_006000), encoding tubulin alpha-4A, in two unrelated patients with clinicopathologically diagnosed sporadic congenital myopathy. The prominent myopathological changes in both patients were muscle fibres with focal myofibrillar disorganisation and rimmed vacuoles. Immunofluorescence showed ubiquitin-positive TUBA4A protein aggregates in the muscle fibres with rimmed vacuoles. Overexpression of the L227F mutant TUBA4A resulted in cytoplasmic aggregates which colocalised with ubiquitin in cellular model. CONCLUSION: Our findings expanded the phenotypic and genetic manifestations of TUBA4A as well as tubulinopathies, and added a new type of congenital myopathy to be taken into consideration in the differential diagnosis.

Geographic differences in pharmacotherapy patterns and outcomes of acute ischemic stroke in China.

BACKGROUND: Vast economic and healthcare status discrepancies exist among regions in China, contributing to different treatment patterns. This study was aimed to investigate the current status of pharmacotherapy for acute ischemic stroke (AIS) and outcomes in China and explore the geographic variation in stroke care. METHODS: This study was a multicenter prospective registry study, which collected the data of patients with AIS from 80 hospitals in 46 cities in 2015-2017 across China. Poor functional outcome defined as a modified Rankin Scale score of 3-6 was assessed at 3 and 12 months. Multivariate logistic regression was used. RESULTS: Among 9973 eligible patients, the number of receiving intravenous thrombolysis (IVT), antiplatelet agents, anticoagulants, statin and human urinary kallidinogenase was 429 (4.3%), 9363 (93.9%), 1063 (10.7%), 6828 (74.7%) and 5112 (51.2%), respectively. Multivariable analysis showed IVT use in northeastern was significantly more frequent than in eastern region (OR = 3.17, 95% CI, 2.53-3.99), while the antiplatelets agents use were less frequent (OR = 0.46, 95%CI: 0.38-0.57). The proportions of poor outcomes at 3 and 12 months were 20.7% and 15.8%, respectively. Multivariate analysis showed AIS patients from northeastern and central region had significantly lower risk of poor outcome at month 3 and 12 than those from eastern region (all P < 0.05). CONCLUSIONS: There was a low IVT use and a high antiplatelet agent and statin use for AIS in China. The pharmacotherapy and prognosis of AIS had variation by geographic region. TRIAL REGISTRATION: This study was registered with ClinicalTrials.gov (NCT02470624).

A new pseudoexon activation due to ultrarare branch point formation in Duchenne muscular dystrophy.

Deep-intronic variants that create or enhance a splice site are increasingly reported as a significant cause of monogenic diseases. However, deep-intronic variants that activate pseudoexons by affecting a branch point are extremely rare in monogenic diseases. Here, we describe a novel deep-intronic DMD variant that created a branch point in a Duchenne muscular dystrophy (DMD) patient. A 7.0-year-old boy was enrolled because he was suspected of DMD based on his clinical, muscle imaging, and pathological features. Routine genetic testing did not discover a pathogenic DMD variant. We then performed muscle-derived dystrophin mRNA analysis and detected an aberrant pseudoexon-containing transcript. Further genomic Sanger sequencing and bioinformatic analyses revealed a novel deep-intronic splicing variant in DMD (NM_004006.2:c.5325+1759G>T), which created a new branch point sequence and thus activated a new dystrophin pseudoexon (NM_004006.2:r.5325_5326ins5325+1779_5325+1855). Our study highlights the significant role of branch point alterations in the pathogenesis of monogenic diseases.

HMGB1-mediated transcriptional activation of circadian gene TIMELESS contributes to endometrial cancer progression through Wnt-ß-catenin pathway.

TIMELESS (TIM) is a circadian gene which is implicated in the regulation of daily rhythm, DNA replication and repair, and cancer initiation and progression. Nevertheless, the role of TIM in endometrial cancer (EC) development is largely unknown. Bioinformatics analysis showed that TIM was aberrantly up-regulated in EC tissues and positively correlated with clinical or histological grade of EC. Functional studies showed that TIM knockdown reduced EC cell viability and restrained EC cell migration in vitro, as well as blocked xenograft tumor growth in vivo. Mechanistically, HMGB1 transcriptionally up-regulated TIM expression in EC cells. In addition, TIM could activate the transcription of the canonical Wnt ligand WNT8B, and TIM depletion could reduce the malignant potential of EC cells largely by targeting and down-regulating WNT8B. As a conclusion, HMGB1/TIM/WNT8B signal cascade was identified in this study for the first time. HMGB1 exerted its oncogenic role by activating the transcription of TIM, leading to the activation of Wnt signaling and EC progression.

Abnormal Ocular Movement in the Early Stage of Multiple-System Atrophy With Predominant Parkinsonism Distinct From Parkinson's Disease.

BACKGROUND AND PURPOSE: The eye-movement examination can be applied as a noninvasive method to identify multiple-system atrophy (MSA). Few studies have investigated eye movements during the early stage of MSA with predominant parkinsonism (MSA-P). We aimed to determine the characteristic oculomotor changes in the early stage of MSA-P. METHODS: We retrospectively selected 17 patients with MSA-P and 40 with Parkinson's disease (PD) with disease durations of less than 2 years, and 40 age-matched healthy controls (HCs). Oculomotor performance in the horizontal direction was measured in detail using videonystagmography. RESULTS: We found that the proportions of patients with MSA-P and PD exhibiting abnormal eye movements were 82.4% and 77.5%, respectively, which were significantly higher than that in the HCs (47.5%, p<0.05). Compared with HCs, patients with MSA-P presented significantly higher abnormal proportions of fixation and gaze-holding (17.6% vs. 0%), without-fixation (47.1% vs. 0%), prolonged latency in reflexive saccades (29.4% vs. 5.0%), memory-guided saccades (93.3% vs. 10.0%), and catch-up saccades in smooth-pursuit movement (SPM, 41.2% vs. 0) (all p<0.05). Compared with those with PD, patients with MSA-P presented a significantly higher proportion of catch-up saccades in SPM (41.2% vs. 2.5%, p<0.001). CONCLUSIONS: MSA-P presented the characteristic of catch-up saccades in SPM in the early stage, which may provide some value in differentiating MSA-P from PD.

Aquaporin 4 Mediates the Effect of Iron Overload on Hydrocephalus After Intraventricular Hemorrhage.

BACKGROUND: Iron overload plays an important role in hydrocephalus development following intraventricular hemorrhage (IVH). Aquaporin 4 (AQP4) participates in the balance of cerebrospinal fluid secretion and absorption. The current study investigated the role of AQP4 in the formation of hydrocephalus caused by iron overload after IVH. METHODS: There were three parts to this study. First, Sprague-Dawley rats received an intraventricular injection of 100 µl autologous blood or saline control. Second, rats had IVH and were treated with deferoxamine (DFX), an iron chelator, or vehicle. Third, rats had IVH and were treated with 2-(nicotinamide)-1,3,4-thiadiazole (TGN-020), a specific AQP4 inhibitor, or vehicle. Rats underwent T2-weighted and T2* gradient-echo magnetic resonance imaging to assess lateral ventricular volume and intraventricular iron deposition at 7, 14, and 28 days after intraventricular injection and were then euthanized. Real-time quantitative polymerase chain reaction, western blot analysis, and immunofluorescence analyses were conducted on the rat brains to evaluate the expression of AQP4 at different time points. Hematoxylin and eosin-stained brain sections were obtained to assess the ventricular wall damage on day 28. RESULTS: Intraventricular injection of autologous blood caused a significant ventricular dilatation, iron deposition, and ventricular wall damage. There was increased AQP4 mRNA and protein expression in the periventricular tissue in IVH rats through day 7 to day 28. The DFX treatment group had a lower lateral ventricular volume and less intraventricular iron deposition and ventricular wall damage than the vehicle-treated group after IVH. The expression of AQP4 protein in periventricular tissue was also inhibited by DFX on days 14 and 28 after IVH. The use of TGN-020 attenuated hydrocephalus development after IVH and inhibited the expression of AQP4 protein in the periventricular tissue between day 14 and day 28 without a significant effect on intraventricular iron deposition or ventricular wall damage. CONCLUSIONS: AQP4 located in the periventricular area mediated the effect of iron overload on hydrocephalus after IVH.

Multifunctional hydrogel for synergistic reoxygenation and chemo/photothermal therapy in metastatic breast cancer recurrence and wound infection.

Metastatic recurrence and postoperative wound infection are two major challenges for breast cancer patients. In this study, a multifunctional responsive hydrogel system was developed for synergistic reoxygenation and chemo/photothermal therapy in metastatic breast cancer and wound infection. The hydrogel system was obtained by cross-linking Prussian blue-modified N-carboxyethyl chitosan (PBCEC) and oxidized sodium alginate using the amino and aldehyde groups on the polysaccharides, resulting in the formation of responsive dynamic imine bonds. Conditioned stimulation (e.g., acid microenvironment) enabled the controlled swelling of hydrogels as well as subsequent slow release of loaded doxorubicin (DOX). Additionally, this hydrogel system decomposed endogenous reactive oxygen species into oxygen to relieve the hypoxic tumor microenvironment and promote the healing of infected-wounds. Both in vitro and in vivo experiments demonstrated the synergistic reoxygenation and chemo/photothermal effects of the PB/DOX hydrogel system against metastatic breast cancer and its recurrence, as well as postoperative wound infection. Thus, the combination of reoxygenation and chemo/photothermal therapy represents a novel strategy for treating and preventing tumor recurrence and associated wound infection.

The clinical, myopathological, and genetic analysis of 155 Chinese mitochondrial ophthalmoplegia patients with mitochondrial DNA single large deletions.

BACKGROUND: Progressive external ophthalmoplegia (PEO) is a common subtype of mitochondrial encephalomyopathy. OBJECTIVE: The study aimed to investigate the relationship between mitochondrial DNA (mtDNA) abnormalities, muscle pathology, and clinical manifestations in Chinese patients with single large-scale mtDNA deletion presenting with PEO. METHODS: This is a retrospective single-center study. Patients with PEO who had a single large deletion in mitochondrial DNA were included in this study. The associations were analyzed between mtDNA deletion patterns, myopathological changes, and clinical characteristics. RESULTS: In total, 155 patients with mitochondrial PEO carrying single large-scale mtDNA mutations were enrolled, including 137 chronic progressive external ophthalmoplegia (CPEO) and 18 Kearns-Sayre syndrome (KSS) patients. The onset ages were 9.61 ± 4.12 in KSS and 20.15 ± 9.06 in CPEO. The mtDNA deletions ranged from 2225 bp to 9131 bp, with m.8470_13446del being the most common. The KSS group showed longer deletions than the CPEO group (p = 0.004). Additionally, a higher number of deleted genes encoding respiratory chain complex subunits (p = 0.001) and tRNA genes (p = 0.009) were also observed in the KSS group. A weak negative correlation between the mtDNA deletion size and ages of onset (p < 0.001, r = -0.369) was observed. The proportion of ragged red fibers, ragged blue fibers, and cytochrome c negative fibers did not correlate significantly with onset ages (p > 0.05). However, a higher percentage of abnormal muscle fibers corresponds to an increased prevalence of exercise intolerance, limb muscle weakness, dysphagia, and cerebellar ataxia. CONCLUSION: We reported a large Chinese cohort consisting of mitochondrial PEO patients with single large-scale mtDNA deletions. Our results demonstrated that the length and locations of mtDNA deletions may influence onset ages and clinical phenotypes. The severity of muscle pathology could not only indicate diagnosis but also may be associated with clinical manifestations beyond the extraocular muscles.

Pathologic changes in neuronal intranuclear inclusion disease are linked to aberrant FUS interaction under hyperosmotic stress.

CGG repeat expansion in NOTCH2NLC is the genetic cause of neuronal intranuclear inclusion disease (NIID). Previous studies indicated that the CGG repeats can be translated into polyglycine protein (N2CpolyG) which was toxic to neurons by forming intranuclear inclusions (IIs). However, little is known about the factors governing polyG IIs formation as well as its molecular pathogenesis. Considering that neurogenetic disorders usually involve interactions between genetic and environmental stresses, we investigated the effect of stress on the formation of IIs. Our results revealed that under hyperosmotic stress, N2CpolyG translocated from the cytoplasm to the nucleus and formed IIs in SH-SY5Y cells, recapitulating the pathological hallmark of NIID patients. Furthermore, N2CpolyG interacted/ co-localized with an RNA-binding protein FUS in the IIs of cellular model and NIID patient tissues, thereby disrupting stress granule formation in cytoplasm under hyperosmotic stress. Consequently, dysregulated expression of microRNAs was found both in NIID patients and cellular model, which could be restored by FUS overexpression in cultured cells. Overall, our findings indicate a mechanism of stress-induced pathological changes as well as neuronal damage, and a potential strategy for the treatment of NIID.

A novel biomarker of fibrofatty replacement in dystrophinopathies identified by integrating transcriptome, magnetic resonance imaging, and pathology data.

BACKGROUND: We aimed to analyse genome-wide transcriptome differences between Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) patients and identify biomarkers that correlate well with muscle magnetic resonance imaging (MRI) and histological fibrofatty replacement in both patients, which have not been reported. METHODS: One hundred and one male patients with dystrophinopathies (55 DMD and 46 BMD) were enrolled. Muscle-derived genome-wide RNA-sequencing was performed in 31 DMD patients, 29 BMD patients, and 11 normal controls. Fibrofatty replacement was scored on muscle MRI and histological levels in all patients. A unique pipeline, single-sample gene set enrichment analysis combined with Spearman's rank correlations (ssGSEA-Cor) was developed to identify the most correlated gene signature for fibrofatty replacement. Quantitative real-time PCR (qRT-PCR) analysis, western blot analysis, and single-nucleus RNA-sequencing (snRNA-seq) were performed in the remaining patients to validate the most correlated gene signature. RESULTS: Comparative transcriptomic analysis revealed that 31 DMD muscles were characterized by a significant increase of inflammation/immune response and extracellular matrix remodelling compared with 29 BMD muscles (P < 0.05). The ssGSEA-Cor pipeline revealed that the gene set of CDKN2A and CDKN2B was the most correlated gene signature for fibrofatty replacement (histological rs  = 0.744, P < 0.001; MRI rs  = 0.718, P < 0.001). Muscle qRT-PCR confirmed that CDKN2A mRNA expression in both 15 DMD (median = 25.007, P < 0.001) and 12 BMD (median = 5.654, P < 0.001) patients were significantly higher than that in controls (median = 1.101), while no significant difference in CDKN2B mRNA expression was found among DMD, BMD, and control groups. In the 27 patients, muscle CDKN2A mRNA expression respectively correlated with muscle MRI (rs  = 0.883, P < 0.001) and histological fibrofatty replacement (rs  = 0.804, P < 0.001) and disease duration (rs  = 0.645, P < 0.001) and North Star Ambulatory Assessment total scores (rs  = -0.698, P < 0.001). Muscle western blot analysis confirmed that both four DMD (median = 2.958, P < 0.05) and four BMD (median = 1.959, P < 0.01) patients had a significantly higher level of CDKN2A protein expression than controls (median = 1.068). The snRNA-seq analysis of two DMD muscles revealed that CDKN2A was mainly expressed in fibro-adipogenic progenitors, satellite cells, and myoblasts. CONCLUSIONS: We identify CDKN2A expression as a novel biomarker of fibrofatty replacement, which might be a new target for antifibrotic therapy in dystrophinopathies.