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Objective: Our study aimed to report the clinical features of ATTR-PN with TTR p.Ala117Ser mutation in mainland China. Methods: Thirteen patients from 13 different families diagnosed with p.Ala117Ser ATTR-PN were identified from three centres. Clinical and laboratory data were retrospectively retrieved for analysis. Results: The male/female ratio was 11:2. All patients showed late onset, with the age of onset at 57.8 ± 5.8 years. The initial symptom was numbness of the lower or upper extremities in 9 patients (69.2%). Paraesthesia was present in all patients. Eleven patients (84.6%) had autonomic dysfunction. Cardiac, renal, hepatic, and ocular dysfunctions were noted in 8 (61.5%), 1 (7.7%), 2 (15.4%), and 3 (23.1%) patients, respectively. Nerve conduction studies have shown axonal-type sensorimotor polyneuropathy. The decline in sensory nerve action potentials was more noticeable than in compound muscle action potentials. The nerve damage present in the lower limbs was more severe than that in the upper limbs. Nerve biopsy revealed positive Congo red staining in 7/10 patients (70%). Conclusion: Our study is the largest population report on this rare p.Ala117Ser mutation in mainland China.

ObjectiveAutoimmune encephalitis (AE) is a large category disorder urging antibody characterization. The aim was to identify a novel AE related autoantibody targeting an intracellular synaptic protein.MethodsSuspected AE patients had negative conventional antibodies screening but strong immunolabel signals on rat brain sections with the serum and cerebrospinal fluid (CSF) samples were considered burdening unkown antibody. Immunoprecipitation from the rat brain protein lysate followed by mass spectrometry analysis was used to identify the targeting antigen. Western blotting and/or cell-based assay (CBA) with antigen-overexpressing HEK293T cells were used for antibody specificity, epitope and IgG subtype determination. Patients with similar immunostaining pattern on rat brain sections were retrospectively screened for the antibody.ResultsThe antibody against collapsin response mediator protein 2 (CRMP2), a synaptic protein involved in axon guidance, was identified in a patient with suspected AE. The patient samples reactivated with HEK293T cell overexpressed CRMP2, rather than CRMP1, 3, 4, and 5. The patient samples mainly stained neuronal cytoplasm of the cortex, hippocampus and cerebellum Purkinje cells. This reactivity was eliminated by pre-immunoabsorption with CRMP2-overexpressing HEK293T cells. CRMP2 truncation experiments indicated that 536 amino acids at C-terminus was necessary for the epitope. Subtype analysis showed that anti-CRMP2 antibody was IgG4. Moreover, screening from 19 suspected AE patients led to identification of anti-CRMP2 antibody in another patient with a diagnosis of encephalomyelitis. The two patients responded to immunotherapy.ConclusionsThis study discovered a novel anti-CRMP2 antibody associated with AE. Testing of the antibody might be promising for AE diagnosis and treatment.

OBJECTIVE. This study aimed to determine whether inflow-based vascular-space-occupancy (iVASO) MRI could reproducibly quantify skeletal muscle perfusion and differentiate patients with dermatomyositis (DM) from healthy subjects. MATERIALS AND METHODS. A total of 25 patients with DM and 22 healthy volunteers underwent iVASO MRI in a 3-T MRI scanner. Maximum and mean arteriolar muscle blood volume (MBV) values of four subgroups of muscles (normal muscles, morphologically normal-appearing muscles, edematous muscles, and atrophic or fat-infiltrated muscles) were obtained. Maximum and mean arteriolar MBV values were compared among the different subgroups, and repeat testing was performed in 20 subjects to assess reproducibility. RESULTS. Compared with normal muscles in healthy subjects, morphologically normal-appearing muscles, edematous muscles, and atrophic or fat-infiltrated muscles in patients with DM showed a significant decrease of both maximum and mean arteriolar MBV (p < .001). Both parameters were significantly lower in atrophic or fat-infiltrated muscles than in morphologically normal-appearing and edematous muscles (p < .001). ROC AUCs for discriminating patients with DM from healthy volunteers were 0.842 and 0.812 for maximum and mean arteriolar MBV values, respectively. As a measure of test-retest studies, the intraclass correlation coefficients (ICCs) were 0.990 (95% CI, 0.986-0.993) and 0.990 (95% CI, 0.987-0.993) for maximum and mean arteriolar MBV, respectively. For interobserver reproducibility, the ICCs were 0.989 (95% CI, 0.986-0.991) and 0.980 (95% CI, 0.975-0.983), respectively. CONCLUSION. iVASO MRI can reproducibly quantify arteriolar MBV in the thigh and discriminate between healthy volunteers and patients with DM.

Nonalcoholic fatty liver disease (NAFLD) is the hepatic manifestation of metabolic syndrome as an arising health issue globally. Obesity and diabetes seem coherent with high NAFLD occurrence. Currently, its underlying mechanism of cause and progression remains to be elicited and effective therapy is unavailable. Huge investment on studying its pathogenesis and innovative therapy is underway. As mechanistic studies are more difficult or unethical to be performed in humans, animal model that recapitulates the human disease is urgently needed. Nonhuman primates (NHPs) with NAFLD can highly mimic the human disease. This study aimed to induce NAFLD by various diets (Table 1) in normoglycemic and diabetic NHPs with biomarker and histopathology assays to quantify NAFLD occurrence. Our data show that diabetic monkeys with high-fat-high-fructose (HFHF) diet developed NAFLD which could assemble to the human disease and be highly valuable for NAFLD research. However, low choline (LC), LC+10% fructose, LC+low methionine (LCM)+10% fructose diets for 31 weeks had no significant effects on liver function, except significant decrease in body weight and food intake in LCM+10%F group and some increase in liver histological scores in LC+10%F group (Table 1). In summary, normal diet with LC or LCM ± high fructose is hardly to induce NAFLD in normoglycemic NHPs, at least in 31 weeks. HFHF could induce or enhance NAFLD in diabetic NHPs. Disclosure J. Gao: Employee; Self; Crown Bioscience. D. Wang: Employee; Self; Crown Bioscience. Y. Ji: Employee; Self; Crown Bioscience. X. Zhang: Employee; Self; Crown Bioscience. C. Yang: None. J. Zhang: Employee; Self; Crown Bioscience. W. Ye: Employee; Self; Crown Bioscience. F. Shi: Employee; Self; Crown Bioscience. M. Jiang: Employee; Self; Crown Bioscience. Q. Jin: Employee; Self; Crown Bioscience. D. Zhang: None. X. Wang: None. W. Song: Employee; Self; Crown Bioscience. K. Chng: None. H. Jiang: Employee; Self; SYSE Bio-tec Co., Ltd. Y. Wang: None. Y. Xiao: Employee; Self; Crown Bioscience.

Background: Previous studies show that the high‐mobility group box protein 1 (HMGB1) and the toll‐like receptor 4 (TLR4) participate in systemic lupus erythematosus (SLE). The two molecules contribute to the occurrence and persistence of seizures in various disease conditions, such as epilepsy. Since seizures are one of the most severe complications associated with neuropsychiatric SLE (NPSLE), the current study aimed at investigating whether HMGB1 and TLR4 play any role in NPSLE related seizures. Methods: Data from 291 SLE patients and 100 healthy controls (HC) were prospectively collected from 2013 to 2018. The ELISA test was used to determine serum levels of HMGB1 for all patients and HC and cerebrospinal fluid (CSF) levels of NPSLE patients. The expression levels of TLR4 by the peripheral blood monocytes (PBMCs) were determined by real-time PCR of TLR4 mRNA. Binary logistic regression and ROC curve analysis were used to predict NPSLE. Results: Among the 291 SLE patients, 188 had active disease and were grouped into two, NPSLE (N=86) and Non-NPSLE (N=102) groups. Among the NPSLE patients, 21 had seizure disorders. Serum HMGB1 levels were increased in NPSLE (8.73±0.29 ng/mL) and were associated with disease activity (r=0.6527, P=0.000). Both serum and CSF HMGB1 levels in NPSLE patients with seizure disorders (9.59±0.63 and 2.90±2.29 ng/mL, respectively) were higher than in patients with other neuropsychiatric symptoms (8.45±0.33 and 2.56±1.70 ng/mL, respectively), though without significance. The gene expression of mRNA TLR4 in PBMCs was similar to serum HMGB1 in the investigated groups. Independent predictors of NPSLE were SLEDAI-2k (OR 1.25; 95% CI: 1.155–1.353), serum HMGB1 (OR 1.659; 95% CI: 1.266–2.175), and anti- Rib-P Ab (OR 3.296; 95% CI: 1.013–10.725). ROC curves for the above predictors had a large AUC (95% CI) of 0.936 (0.900–0.971), indicating a good prediction of NPSLE occurrence. Conclusions: The expression of HMGB1 and TLR4 was increased in NPSLE, but HMGB1 and TLR4 had minimal effect on NPSLE related seizures. The serum levels of HMGB1 were positively correlated with disease activity, and could, therefore, be a potential biomarker of NPSLE for use in future clinical practice.

Leukemia-initiating cells (LICs) are responsible for the initiation, development, and relapse of leukemia. The identification of novel therapeutic LIC targets is critical to curing leukemia. In this report, we reveal that junctional adhesion molecule 3 (JAM3) is highly enriched in both mouse and human LICs. Leukemogenesis is almost completely abrogated upon Jam3 deletion during serial transplantations in an MLL-AF9-induced murine acute myeloid leukemia model. In contrast, Jam3 deletion does not affect the functions of mouse hematopoietic stem cells. Moreover, knockdown of JAM3 leads to a dramatic decrease in the proliferation of both human leukemia cell lines and primary LICs. JAM3 directly associates with LRP5 to activate the downstream PDK1/AKT pathway, followed by the downregulation of GSK3β and activation of β-catenin/CCND1 signaling, to maintain the self-renewal ability and cell cycle entry of LICs. Thus, JAM3 may serve as a functional LIC marker and play an important role in the maintenance of LIC stemness through unexpected LRP5/PDK1/AKT/GSK3β/β-catenin/CCND1 signaling pathways but not via its canonical role in cell junctions and migration. JAM3 may be an ideal therapeutic target for the eradication of LICs without influencing normal hematopoiesis.

Purpose The application of metagenomic next-generation sequencing (mNGS) in the diagnosis of tuberculous meningitis (TBM) remains poorly characterized. Here, we retrospectively analyzed data from patients with TBM who had taken both mNGS and conventional tests including culture of Mycobacterium tuberculosis (MTB), polymerase chain reaction (PCR) and acid-fast bacillus (AFB) stain, and the sensitivity and specificity of these methods were compared. Methods We retrospectively recruited TBM patients admitted to the hospital between December 2015 and October 2018. The first collection of cerebrospinal fluid (CSF) samples underwent both mNGS and conventional tests. In addition, patients with bacterial/cryptococcal meningitis or viral meningoencephalitis were mNGS positive controls, and a patient with auto-immune encephalitis was an mNGS negative control. Results Twenty three TBM patients were classified as 12 definite and 11 clinical diagnoses, which were based on clinical manifestations, pathogen evidence, CSF parameters, brain imaging, and treatment response. The mNGS method identified sequences of Mycobacterium tuberculosis complex (MBTC) from 18 samples (18/23, 78.26%). In patients with definite TBM, the sensitivity of mNGS, AFB, PCR, and culture to detect MTB in the first CSF samples were 66.67, 33.33, 25, and 8.33%, respectively. The specificity of each method was 100%. Among the four negative mNGS cases (4/23, 17.39%), three turned out positive by repeated AFB stain. The agreement of mNGS with the total of conventional methods was 44.44% (8/18). Combination of mNGS and conventional methods increased the detection rate to 95.65%. One patient was diagnosed as complex of TBM and cryptococcal meningitis, in which AFB stain and cryptococcal antigen enzyme immunoassay were positive and the DNA of Cryptococcus neoformans was detected by mNGS. Conclusion Our study indicates that mNGS is an alternative method to detect the presence of mycobacterial DNA in CSF samples from patients with TBM and deserves to be applied as a front-line CSF test.

BackgroundInfectious encephalitis and meningitis are often treated empirically without identification of the causative pathogen. Metagenomic next-generation sequencing (mNGS) is a high throughput technology that enables the detection of pathogens independent of prior clinical or laboratory information.MethodsThe present study was a multicentre prospective evaluation of mNGS of cerebrospinal fluid (CSF) for the diagnosis of suspected central nervous system infections.ResultsA total of 276 patients were enrolled in this study between Jan 1, 2017 and Jan 1, 2018. Identification of an etiologic pathogen in CSF by mNGS was achieved in 101 patients (36.6%). mNGS detected 11 bacterial species, 7 viral species, 2 fungal species, and 2 parasitic species. The five leading positive detections were varicella-zoster virus (17), Mycobacterium tuberculosis (14), herpes simplex virus 1 (12), Epstein-Barr virus (12), and Cryptococcus neoformans (7). False positives occurred in 12 (4.3%) patients with bacterial infections known to be widespread in hospital environments. False negatives occurred in 16 (5.8%) patients and included bacterial, viral and fungal aetiologies.ConclusionsmNGS of CSF is a powerful diagnostic method to identify the pathogen for many central nervous system infections.

We previously reported familial amyotrophic lateral sclerosis (FALS) of 11 years duration in a 57-year-old woman, who received artificial ventilation for 5 years prior to death and exhibited widespread multisystem degeneration and neurofilamentous aggregates, so-called conglomerate inclusions (CIs). In the present study, we re-evaluated this autopsied patient (proband) with further immunohistochemical observation as well as mutational analysis of the superoxide dismutase 1 (SOD1) gene. A review of the clinical features of the proband's family revealed five affected members (including the proband) over two successive generations who showed marked variability in clinical presentation, such as the age at onset. The proband was found to harbor a heterozygous missense mutation in exon 4 (I104F) of the SOD1 gene. In the brain and spinal cord, SOD1-positive neuronal cytoplasmic inclusions (NCIs) were found to be more widely distributed than CIs, the latter being weakly positive for SOD1. No Lewy body-like hyaline inclusions were found. This is considered to be the first description of an autopsy case of FALS with an I104F SOD1 gene mutation, suggesting that combination of marked intra-familial clinical variability and multisystem degeneration with occurrence of CIs and SOD1-positive NCIs is a characteristic feature of FALS with this SOD1 gene mutation.

Bunina bodies (BBs) are small eosinophilic neuronal cytoplasmic inclusions (NCIs) found in the remaining lower motor neurons (LMNs) of patients with sporadic amyotrophic lateral sclerosis (SALS), being a specific feature of the cellular pathology. We examined a case of SALS, unassociated with TDP-43 or C9ORF72 mutation, of 12 years duration in a 75-year-old man, who had received artificial respiratory support for 9 years, and showed widespread multisystem degeneration with TDP-43 pathology. Interestingly, in this patient, many NCIs reminiscent of BBs were observed in the oculomotor nucleus, medullary reticular formation and cerebellar dentate nucleus. As BBs in the cerebellar dentate nucleus have not been previously described, we performed ultrastructural and immunohistochemical studies of these NCIs to gain further insight into the nature of BBs. In each region, the ultrastructural features of these NCIs were shown to be identical to those of BBs previously described in LMNs. These three regions and the relatively well preserved sacral anterior horns (S1 and S2) and facial motor nucleus were immunostained with antibodies against cystatin C (CC) and TDP-43. Importantly, it was revealed that BBs exhibiting immunoreactivity for CC were a feature of LMNs, but not of non-motor neurons, and that in the cerebellar dentate nucleus, the ratio of neurons with BBs and TDP-43 inclusions/neurons with BBs was significantly lower than in other regions. These findings suggest that the occurrence of BBs with CC immunoreactivity is intrinsically associated with the particular cellular properties of LMNs, and that the mechanism responsible for the formation of BBs is distinct from that for TDP-43 inclusions.

Type 2 episodic ataxia (EA2) is the most common subtype among a group of rare hereditary syndromes characterized by recurrent attacks of ataxia. More than 60 mutations and several gene rearrangements due to large deletions in CACNA1A gene have been reported so far for the cause of EA2. Because CACNA1A gene is a large gene containing 47 exons and there is no hot spot mutation, direct sequencing will be a challenge in clinical genetic testing. In this study, we used next generation sequencing technology to identify a novel nonsense mutation of CACNA1A (p.Tyr1957Ter, NP_001120693.1) resulting in truncated protein without 305 amino acids in the c-terminus. Sanger sequencing confirmed the heterozygous mutation of CACNA1A in a Chinese family with 11 affected individuals. Affected individuals experienced recurrent attacks with or without nystagmus, dysarthria, seizure, myokymia, dystonia, weakness, blurred vision, visual field defects, diplopia, migraine, dizziness, nausea and vomiting, sweating and abdominal pain. This is the first report of EA2 in a Chinese family that carries a novel mutation in CACNA1A gene and had abdominal pain as a novel phenotype associated with EA2.