Systemic and skin-limited delayed-type drug hypersensitivity reactions associate with distinct resident and recruited T cell subsets.

Delayed-type drug hypersensitivity reactions are major causes of morbidity and mortality. The origin, phenotype, and function of pathogenic T cells across the spectrum of severity require investigation. We leveraged recent technical advancements to study skin-resident memory T cells (TRMs) versus recruited T cell subsets in the pathogenesis of severe systemic forms of disease, Stevens-Johnson syndrome/toxic epidermal necrolysis (SJS/TEN) and drug reaction with eosinophilia and systemic symptoms (DRESS), and skin-limited disease, morbilliform drug eruption (MDE). Microscopy, bulk transcriptional profiling, and single-cell RNA-sequencing (scRNA-Seq) plus cellular indexing of transcriptomes and epitopes by sequencing (CITE-Seq) plus T cell receptor sequencing (TCR-Seq) supported clonal expansion and recruitment of cytotoxic CD8+ T cells from circulation into skin along with expanded and nonexpanded cytotoxic CD8+ skin TRM in SJS/TEN. Comparatively, MDE displayed a cytotoxic T cell profile in skin without appreciable expansion and recruitment of cytotoxic CD8+ T cells from circulation, implicating TRMs as potential protagonists in skin-limited disease. Mechanistic interrogation in patients unable to recruit T cells from circulation into skin and in a parallel mouse model supported that skin TRMs were sufficient to mediate MDE. Concomitantly, SJS/TEN displayed a reduced Treg signature compared with MDE. DRESS demonstrated recruitment of cytotoxic CD8+ T cells into skin as in SJS/TEN, yet a pro-Treg signature as in MDE. These findings have important implications for fundamental skin immunology and clinical care.

Unusual clinical presentation of cervical extradural meningioma detected with neuromuscular ultrasound: A case report.

Extradural meningiomas are rare in the cervical region. A total of 70-77% of reported cases have occurred in the thoracic region. Tumors that occur in the cervical region may invade the adjacent nerve root and brachial plexus. Typically, diagnoses of extradural meningioma are made after patients present with signs of myelopathy, such as progressive paresis and numbness. In the current study, a 64-year-old male patient presented with neck pain, numbness and mild weakness in the left hand over a 6-month period. The general neurological examination was unremarkable, except for mild grasping weakness on the left side. Needle electromyography revealed complex repetitive discharges in the left 5 and 6th cervical paraspinal muscles. Neuromuscular ultrasound revealed a lesion over the left 7th cervical root, which enabled the early detection of an extradural meningioma before notable focal neurological defects developed. The patient underwent a subtotal tumor excision, followed by radiotherapy for residual tumor. Histopathological examination confirmed atypical meningioma.

A New Phenotype of TUBB4A Mutation in a Family With Adult-Onset Progressive Spastic Paraplegia and Isolated Hypomyelination Leukodystrophy: A Case Report and Literature Review.

Tubulin beta 4A class IVa (TUBB4A) spectrum disorders include autosomal dominant dystonia type 4 or hypomyelination with atrophy of the basal ganglia and cerebellum (H-ABC syndrome). However, in rare cases, only mild hypomyelination in the cortex with no basal ganglia atrophy may be observed. We report a case of a family with TUBB4A mutation and complicated hereditary spasticity paraplegia (HSP). We performed quadro whole-exome sequencing (WES) on the family to identify the causative gene of progressive spastic paraparesis with isolated hypomyelination leukodystrophy. We identified a novel TUBB4A p.F341L mutation, which was present in all three affected patients but absent in the unaffected father. The affected patients presented with adult-onset TUBB4A disorder, predominant spastic paraparesis with/without ataxia, and brain hypomyelination with no cognitive impairment or extrapyramidal symptoms. In the literature, HSP is considered a TUBB4A spectrum disorder.

Ultrasonography of abdominal muscles: Differential diagnosis of late-onset Pompe disease and myotonic dystrophy type 1.

Introduction: Axial muscles are involved earlier and to a greater extent in late-onset Pompe disease (LOPD) than in myotonic muscular dystrophy type 1 (DM1). We aimed to evaluate abdominal muscles in LOPD compared in DM1 using muscle ultrasonography. Methods: Patients with LOPD (n = 3), DM1 (n = 10), and age- and gender-matched healthy subjects (n = 34) were enrolled for muscle ultrasonography. Patients with LOPD and DM1 were 20 to 59 years of age with a disease duration ranging between 7 and 30 years. A multifrequency linear transducer was used to evaluate quality and thickness in the abdominal muscles and extremities. Results: The quantitative muscle echo score revealed a higher Z score in abdominal muscles in Patients with LOPD (scores were relatively normal for the biceps and flexor digitorum groups). Patients with LOPD had significantly lower abdominal muscle thickness than patients with DM1. Abdominal muscle strength was significantly correlated with the muscle echogenicity, trunk impairment scale, and trunk control test. The extremities' sum score was correlated with the total Medical Research Council score. Discussion: The increased quantitative muscle score in abdominal muscles, sparing the biceps and flexor digitorum groups, may offer differential diagnosis between LOPD and DM1. Ultrasound can easily access abdominal muscles and investigate muscle echogenicity and thickness. A quantitative approach using muscle echogenicity rather than muscle thickness may provide a greater correlation with trunk muscle function.

Diagnosis and Clinical Features in Autoimmune-Mediated Movement Disorders.

Movement disorders are common manifestations in autoimmune-mediated encephalitis. This group of diseases is suspected to be triggered by infection or neoplasm. Certain phenotypes correlate with specific autoantibody-related neurological disorders, such as orofacial-lingual dyskinesia with N-methyl-D-aspartate receptor encephalitis and faciobrachial dystonic seizures with leucine-rich glioma-inactivated protein 1 encephalitis. Early diagnosis and treatment, especially for autoantibodies targeting neuronal surface antigens, can improve prognosis. In contrast, the presence of autoantibodies against intracellular neuronal agents warrants screening for underlying malignancy. However, early clinical diagnosis is challenging because these diseases can be misdiagnosed. In this article, we review the distinctive clinical phenotypes, magnetic resonance imaging findings, and current treatment options for autoimmune-mediated encephalitis.

Cross-Sectional Area Reference Values for Sonography of Peripheral Nerves in Taiwanese Adults.

Background: Neuromuscular ultrasound is a complementary technology that aids in the diagnosis of peripheral neuropathy. The interpretation of neuromuscular ultrasound results requires the use of accurate normative cross-sectional area (CSA) reference values. This study aims to provide CSA reference values specific to Taiwanese adults for Sonography of peripheral nerves in the upper and lower extremities. Methods: The study cohort included 66 healthy subjects (36 women; 30 men). A linear probe was used to measure the CSA of the median, ulnar, radial, tibial, sural, and peroneal nerves at multiple sites. These data were analyzed to determine standard ranges for the CSA at each site (reference range = mean ± 2 × SD) and identify correlations between the CSA and patient characteristics. Results: Normative CSA ranges were determined for all the assessed nerve sites, revealing that the nerve sizes in this Taiwanese population were smaller than Caucasian populations but comparable to those reported for other Asian cohorts. Men tended to have larger nerves than women, even after adjusting for height and weight. The size of ulnar nerve in the cubital tunnel and the peroneal nerve in the popliteal fossa correlated negatively with increasing age. The nerve size correlated positively with increasing weight and BMI at several sites, correlation of median nerve in the forearm with weight and BMI was significant after multiple testing. Significant correlation was also found between size of ulnar nerve in cubital tunnel and decreasing height. Conclusion: We provide reference ranges for neuromuscular ultrasound CSA values for the upper and lower extremities that are specific to the Taiwanese population. These reference values may be useful for evaluating peripheral neuropathy in Taiwanese subjects.

Peripheral host T cells survive hematopoietic stem cell transplantation and promote graft-versus-host disease.

Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality in hematopoietic stem cell transplantation (HSCT). Donor T cells are key mediators in pathogenesis, but a contribution from host T cells has not been explored, as conditioning regimens are believed to deplete host T cells. To evaluate a potential role for host T cells in GVHD, the origin of skin and blood T cells was assessed prospectively in patients after HSCT in the absence of GVHD. While blood contained primarily donor-derived T cells, most T cells in the skin were host derived. We next examined patient skin, colon, and blood during acute GVHD. Host T cells were present in all skin and colon acute GVHD specimens studied, yet were largely absent in blood. We observed acute skin GVHD in the presence of 100% host T cells. Analysis demonstrated that a subset of host T cells in peripheral tissues were proliferating (Ki67+) and producing the proinflammatory cytokines IFN-γ and IL-17 in situ. Comparatively, the majority of antigen-presenting cells (APCs) in tissue in acute GVHD were donor derived, and donor-derived APCs were observed directly adjacent to host T cells. A humanized mouse model demonstrated that host skin-resident T cells could be activated by donor monocytes to generate a GVHD-like dermatitis. Thus, host tissue-resident T cells may play a previously unappreciated pathogenic role in acute GVHD.

POLG R964C and GBA L444P mutations in familial Parkinson's disease: Case report and literature review.

Polymerase gamma (POLG) is an enzyme responsible for the replication and repair of mitochondrial DNA. Mutations in POLG may cause variable clinical manifestations, including parkinsonism, epilepsy, cerebellar ataxia, neuropathy, and progressive external ophthalmoplegia. However, mutations of this gene are rare in patients with typical Parkinson's disease (PD). We report a man (current age: 59 years) without any underlying disease presenting with right-hand tremor at the age of 39 years, followed by slow movement, rigidity, and postural instability. He developed motor fluctuation and levodopa-induced dyskinesia 8 years later. At the age of 58 years, cognitive decline and visual hallucination ensued; he was institutionalized thereafter. We used multiplex ligation-dependent probe amplification, which demonstrated no large deletions or duplications of relevant PD genes. Next, targeted sequencing panel covering 51 genes causative for PD was applied for the proband; it revealed a heterozygous missense substitution R964C in POLG and a heterozygous missense substitution L444P in GBA. The patient's father, who had been diagnosed as having PD and type 2 diabetes mellitus at the age of 70 years, demonstrated identical mutations. This is the first report of familial PD combined with POLG R964C and GBA L444P mutations. Two pathogenic gene mutations potentially cause double hit in pathological neurodegeneration. This finding extends our understanding of the PD genotype-phenotype correlation.

Predictive Factors for Early Initiation of Artificial Feeding in Patients With Sporadic Creutzfeldt-Jakob Disease.

Background: Akinetic mutism has often been used as the predictor of sporadic Creutzfeldt-Jacob disease (sCJD) endpoints, but it may be difficult for general physcians to assess. Nasogastric (NG) tube insertion is indicated for many neurodegenerative diseases with a clinical course of swallowing failure, and can be more easily identified than akinetic mutism by general physicians. Therefore, the aim of this study was to identify whether there are predictive factors for early initiation of artificial feeding in patients with sCJD who require enteral nutrition due to swallowing failure. Methods: We retrospectively reviewed the medical records of all patients diagnosed with probable sCJD who were admitted to the neurology ward at a medical center in Taiwan from January 2002 to July 2017. We used Pearson's chi-squared test to detect the correlation of initial symptoms, neurological signs, brain magnetic resonance imaging (MRI), electroencephalography (EEG), and increased levels of 14-3-3 protein in cerebrospinal fluid (CSF) analysis. The Cox proportional hazards model was used to detect prognostic factors for early initiation of NG tube insertion in sCJD patients. Results: The onset age ranged from 51 to 83 years, and mostly ranged from 60 to 79 years. Akinetic mutism was correlated with pyramidal tract signs, myoclonus, and extrapyramidal signs. Furthermore, myoclonus was revealed to be associated with pyramidal tract signs. Multivariate Cox regression analysis showed that myoclonus and elevated CSF levels of 14-3-3 protein are predictive of early NG insertion. Conclusions: Increased levels of 14-3-3 protein in CSF and the presence of myoclonus at diagnosis are predictive of early swallowing difficulty and indicate rapid deterioration in probable sCJD. In addition to akinetic mutism, early initiation of artificial feeding can be used to predict early deterioration in sCJD.

CHD1L Regulated PARP1-Driven Pluripotency and Chromatin Remodeling During the Early-Stage Cell Reprogramming.

PARP1 and poly(ADP-ribosyl)ation (PARylation) have been shown to be essential for the initial steps of cellular reprogramming. However, the mechanism underlying PARP1/PARylation-regulated activation of pluripotency loci remains undetermined. Here, we demonstrate that CHD1L, a DNA helicase, possesses chromatin remodeling activity and interacts with PARP1/PARylation in regulating pluripotency during reprogramming. We found that this interaction is mediated through the interplay of the CHD1L macro-domain and the PAR moiety of PARylated-PARP1. Chromatin immunoprecipitation assays demonstrated the co-occupancy of CHD1L and PARP1 at Pou5f1, Nanog, and Esrrb pluripotency loci. Knockdown of CHD1L significantly blocked the binding activity of PARP1 at pluripotency loci and inhibited the efficiency of PARP1-driven reprogramming. Notably, we found that CHD1L-promoted reprogramming requires both a PARP1-interacting domain and DNA helicase activity, partly contributing to the chromatin-remodeling states of pluripotency loci. Taken together, these results identify CHD1L as a key chromatin remodeler involved in PARP1/PARylation-regulated early-stage reprogramming and pluripotency in stem cells.

Biological evaluation of secondary metabolites from the roots of Myrica adenophora.

Bioassay-guided fractionation of the roots of Myrica adenophora led to isolation of 24 known compounds and hitherto unknown compounds, including three A-type proanthocyanidins [adenodimerins A-C], two esters of sucrose [myricadenins A and B ], and the phenolic glycoside 6'-O-galloyl orbicularin. Spectroscopic analyses were used to determine their structures. Adenodimerin A, myricananin C, and myricetin showed strong 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activities, with SC50 values of 7.9, 16.3, and 15.9 µM, respectively. Adenodimerin A, myricanone, myricananin C, (-)-myricanol, myricanol 11-O-ß-D-glucopyranoside, and myricetin showed stronger 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS) radical scavenging activities than the positive control, with SC50 values of 7.5, 19.6, 12.0, 22.3, 19.6, and 15.6 µM, respectively. 5-Deoxymyricanone, porson, 12-hydroxymyricanone (-)-myricanol, and (+)-galeon exhibited anti-tubercular activity against Mycobacterium tuberculosis H37Rv in vitro and MICs values of 25.8, 40.0, 35.8, 30.0, and 15.0 µg/mL, respectively. Myricadenin A, myricanone, myricananin C, and (-)-myricanol exhibited anti-inflammatory activities in the iNOS assay with EC50 values of 18.1, 1.00, 13.0, and 7.5 µM, respectively.

Melamine induces human renal proximal tubular cell injury via transforming growth factor-ß and oxidative stress.

Although several reports have failed to observe adverse subchronic renal effects following relatively high melamine exposure, the safety of low and continuous melamine exposure is still debatable. Recent studies suggest that long-term, low-dose melamine exposure is associated with an increased risk of urolithiasis, which has been linked to chronic kidney disease (CKD). CKD is a consequence of nephron loss and is associated with the interaction of inflammation, oxidative stress, and transforming growth factor-ß (TGF-ß), which increases extracellular matrix genes and cell apoptosis with progression to fibrosis and end-stage renal disease. Thus far, information is still lacking regarding the influence of melamine at the gene and protein levels, which are activated at a much earlier phase than the occurrence of the renal morphological change. In this study, we stimulated human renal proximal tubular HK-2 cells with melamine (0, 125, 250, 500, or 1000 µg/ml) for different time intervals and observed its effects on several well-documented molecular mechanisms of CKD. Here, we demonstrate that melamine can activate mitogen-activated protein kinases, NFκB, and reactive oxygen species, which results in the upregulation of interleukin-6, monocyte chemoattractant protein-1, vascular cell adhesion molecule-1, and TGF-ß1 in HK-2 cells. The melamine-stimulated overexpression of TGF-ß1 not only promotes fibronectin production but also leads to decreased antiapoptotic (bcl-2, bcl-xl)/proapoptotic (bad, bax) protein ratio, increased caspase-3 and caspase-9 activities, and eventually HK-2 cell apoptosis. Our study suggests that melamine exposure may be a risk factor for the chronic loss of tubular cells and may ultimately lead to tubulointerstitial damage.

DDA3 stabilizes microtubules and suppresses neurite formation.

We have previously shown that DDA3 - also known as proline/serine-rich coiled-coil protein 1 (PSRC1) - is a microtubule-associated protein that promotes cell growth by stimulating the ß-catenin pathway. Here, we report that DDA3 can bundle and stabilize microtubules in vivo and in vitro. We found that overexpression of DDA3 increased the abundance of acetylated and tyrosinated microtubules. We employed PC12 and N2a cell lines, as well as cultured hippocampal neurons, and demonstrated that overexpression of DDA3 suppressed neurite/axon outgrowth, whereas its depletion accelerated neurite/axon formation and elongation. Knockdown of DDA3 reduced ß3-tubulin levels in N2a cells, which contributed to the spontaneous neurite formation caused by DDA3 depletion. Consistent with its role in suppressing neuritogenesis, DDA3 was downregulated during induced neuronal differentiation. Moreover, expression of DDA3 was detected in the rat brain at embryonic (E) day E15 and in the cortical region at E17, the period of active neurogenesis. Levels of cortical DDA3 decreased at the beginning of E19, when active neuritogenesis is completed. Overall our results demonstrate that DDA3 is a so-far-unknown microtubule-stabilizing protein that is involved in regulating neurite formation and elongation.

Suppression of Glutamine:fructose-6-phosphate amidotransferase-1 inhibits adipogenesis in 3T3-L1 adipocytes.

O-linked N-acetylglucosamine (O-GlcNAc) protein modification has been implicated in the regulation of signaling pathways, cell function, and gene expression. Glutamine:fructose-6-phosphate amidotransferase-1 (GFAT-1) is the rate-limiting enzyme in the hexosamine biosynthetic pathway (HBP), which generates the sugar nucleotide UDP-GlcNAc, where this nucleotide acts as the donor for O-GlcNAc modification. In this study, we determined whether GFAT-1 regulates adipogenesis in adipocytes. 3T3-L1 preadipocytes were differentiated using medium containing high glucose, insulin, dexamethasone, and isobutylmethylxanthine. Cells were harvested 4, 8, and 12 h and 1, 2, 3, 4, 6, and 8 days after the initiation of differentiation. Global level of O-GlcNAc modification increased 4 h after induction and persisted for 8 days of observation. GFAT-1 mRNA and protein expression was also upregulated beginning 4 h after induction. Pharmacological inhibition of GFAT-1 or GFAT-1 siRNA treatment blocked the increase in O-GlcNAcylation and the formation of lipid droplets in adipocytes. GFAT-1 may regulate the expression of C/EBPß, PPARγ, SREBP-1, fatty acid synthase, S3-12, perilipin, or adipophilin during adipogenesis. Our results suggest that GFAT-1 plays a critical role in modulating adipogenesis via the regulation of protein O-GlcNAcylation in adipocytes.

Targeting autophagy enhances BO-1051-induced apoptosis in human malignant glioma cells.

PURPOSE: BO-1051 is an N-mustard derivative that is conjugated with DNA-affinic 9-anilinoacridine. Since BO-1051 was reported to have strong anticancer activity, we investigated the effect and underlying mechanism of BO-1051 in human glioma cell lines. METHODS: Human glioma cell lines U251MG and U87MG were studied with BO-1051 or the combination of BO-1051 and autophagic inhibitors. Growth inhibition was assessed by MTT assay. Apoptosis was measured by annexin V staining followed by flow cytometry and immunoblotting for apoptosis-related molecules. Induction of autophagy was detected by acridine orange labeling, electron microscopy, LC3 localization and its conversion. Transfection of shRNA was used to determine the involvement of Beclin1 in apoptotic cell death. RESULTS: MTT assay showed that BO-1051 suppressed the viability of four glioma cell lines (U251MG, U87MG, GBM-3 and DBTRG-05MG) in a dose-dependent manner. The IC(50) values of BO-1051 for the glioma cells were significantly lower than the values for primary neurons cultures and normal fibroblast cells. Moreover, BO-1051 not only induced apoptotic cell death, but also enhanced autophagic flux via inhibition of Akt/mTOR and activation of Erk1/2. Importantly, suppression of autophagy by 3-methyladenine or bafilomycin A1 significantly increased BO-1051-induced apoptotic cell death in U251MG and U87MG cells. In addition, the proportion of apoptotic cells after BO-1051 treatment was enhanced by co-treatment with shRNA against Beclin1. CONCLUSIONS: BO-1051 induced both apoptosis and autophagy, and inhibition of autophagy significantly augmented the cytotoxic effect of BO-1051. Thus, a combination of BO-1051 and autophagic inhibitors offers a potentially new therapeutic modality for the treatment of malignant glioma.

Betel nut extract and arecoline block insulin signaling and lipid storage in 3T3-L1 adipocytes.

According to several population-based studies, betel nut chewing is associated with metabolic syndrome and diabetes in British South Asians and Taiwanese. However, the underlying molecular mechanism is not yet clear. Arecoline is an alkaloid-type natural product found in betel nuts. Our aim was to clarify the influence of betel nut extract and arecoline on lipid accumulation and insulin signaling in adipocytes. We found that betel nut extract and arecoline blocked lipid storage in 3T3-L1 adipocytes. The possible mechanism may function by inhibiting the expression of the insulin receptor, glucose transporter-4, fatty acid synthase, and the lipid droplet proteins perilipin and adipophilin. In addition, betel nut extract and arecoline increased the basal level of IRS-1 serine(307) phosphorylation and decreased insulin-stimulated IRS-1 tyrosine, Akt, and PI3 kinase phosphorylation. In conclusion, betel nut extract and arecoline have diabetogenic potential on adipocytes that may result in insulin resistance and diabetes at least in part via the obstruction of insulin signaling and the blockage of lipid storage.

Rad is a p53 direct transcriptional target that inhibits cell migration and is frequently silenced in lung carcinoma cells.

The p53 tumor suppressor exerts its function mainly as a transcriptional activator. Here we show that the Ras-related small GTPase Rad, an inhibitor of Rho kinase, is a direct transcriptional target of p53. Expression of Rad messenger RNA (mRNA) and protein was induced by DNA damage in a p53-dependent manner. The -2934/-2905-bp Rad promoter region, to which p53 bound, was required for p53-mediated Rad gene activation. Treatment by DNA damaging agents increased p53 occupancy and histone acetylation in the region of Rad promoter containing the p53-binding site. Expression of Rad diminished the inhibitory phosphorylation at Ser3 of cofilin, a regulator of actin dynamics, and suppressed migration and invasiveness of cancer cells. Knockdown of Rad promoted cell migration and alleviated the p53-mediated migration suppression. Frequent loss of Rad mRNA and protein expression was observed in non-small cell lung carcinoma tissues. Together our results reveal a mechanism that p53 may inhibit cell migration by disrupting actin dynamics via Rad activation and implicate a tumor suppressor role of Rad in lung cancer.

Low-energy visible light irradiation modulates immune responses induced by epicutaneous sensitization with protein antigen.

Epicutaneous sensitization has been an important route for protein allergen sensitization in atopic disease. Although the skin is irradiated by sunlight daily, the influence of visible light on epicutaneous sensitization has not been explored. In this study, by using a well-established murine protein-patch model, we show that low-energy visible light (LEVL) irradiation could differentially modulate the predominant Th2 immune response induced by epicutaneous sensitization with protein antigen. When the induced Th2 response was strong, as usually observed in BALB/c mice, LEVL irradiation suppressed the response. In contrast, LEVL irradiation enhanced the weaker Th2 response in C57BL/6 mice. Increased IL-18 and decreased TGF-beta expression in draining lymph nodes after LEVL irradiation was observed in BALB/c mice, but not in C57BL/6 mice. LEVL irradiation also enhanced IL-18 expression in skin and reduced the downregulation of CD24 expression on epidermal Langerhans cells in draining lymph nodes of BALB/c mice. Collectively, these results provide evidence for immunomodulatory effects of LEVL irradiation and will help us develop a useful strategy for prevention of allergen sensitization.