Drug hypersensitivity such as severe cutaneous adverse reactions (SCAR), including Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN), could be life-threatening. Here, we enroll SCAR ...patients to investigate the T cell receptor (TCR) repertoire by next-generation sequencing. A public αβTCR is identified from the cytotoxic T lymphocytes of patients with carbamazepine-SJS/TEN, with its expression showing drug/phenotype-specificity and an bias for HLA-B*15:02. This public αβTCR has binding affinity for carbamazepine and its structural analogs, thereby mediating the immune response. Adoptive transfer of T cell expressing this public αβTCR to HLA-B*15:02 transgenic mice receiving oral administration of carbamazepine induces multi-organ injuries and symptoms mimicking SCAR, including hair loss, erythema, increase of inflammatory lymphocytes in the skin and blood, and liver and kidney dysfunction. Our results not only demonstrate an essential role of TCR in the immune synapse mediating SCAR, but also implicate potential clinical applications and development of therapeutics.
The combination of EGF, CHIR99021, A83-01, SB431542, VPA, and Y27632 (EGF/CASVY) facilitates the derivation of trophoblast stem (TS) cells from human blastocysts and first-trimester, but not term, ...cytotrophoblasts. The mechanism underlying this chemical induction of TS cells remains elusive. Here we demonstrate that the induction efficiency of cytotrophoblast is determined by functional antagonism of the placental transcription factor GCM1 and the stemness regulator ΔNp63α. ΔNp63α reduces GCM1 transcriptional activity, whereas GCM1 inhibits ΔNp63α oligomerization and autoregulation. EGF/CASVY cocktail activates ΔNp63α, thereby partially inhibiting GCM1 activity and reverting term cytotrophoblasts into stem cells. By applying hypoxia condition, we can further reduce GCM1 activity and successfully induce term cytotrophoblasts into TS cells. Consequently, we identify mitochondrial creatine kinase 1 (CKMT1) as a key GCM1 target crucial for syncytiotrophoblast differentiation and reveal decreased CKMT1 expression in preeclampsia. Our study delineates the molecular underpinnings of trophoblast stemness and differentiation and an efficient method to establish TS cells from term placentas.
Stress-induced phosphoprotein-1 (STIP1)-a heat shock protein (HSP)70/HSP90 adaptor protein-is commonly overexpressed in malignant cells, where it controls proliferation via multiple signaling ...pathways, including JAK2/STAT3. We have previously shown that STIP1 stabilizes the protein tyrosine kinase JAK2 in cancer cells via HSP90 binding. In this study, we demonstrate that STIP1 may act as a substrate for JAK2 and that phosphorylation of tyrosine residues 134 and 152 promoted STIP1 protein stability, induced its nuclear-cytoplasmic shuttling, and promoted its secretion into the extracellular space. We also found that JAK2-mediated STIP1 phosphorylation enhanced cell viability and increased resistance to cisplatin-induced cell death. Conversely, interference STIP1 with JAK2 interaction-attained either through site-directed mutagenesis or the use of cell-penetrating peptides-decreased JAK2 protein levels, ultimately leading to cell death. On analyzing human ovarian cancer specimens, JAK2 and STIP1 expression levels were found to be positively correlated with each other. Collectively, these results indicate that JAK2-mediated phosphorylation of STIP-1 is critical for sustaining the JAK2/STAT3 signaling pathway in cancer cells.
Karyopherins, the nucleocytoplasmic transporters, participate in multiple RNA silencing stages by transporting associated proteins into the nucleus. Importin α is a member of karyopherins and has ...been reported to facilitate virus infection via nuclear import of viral proteins. Unlike other RNA viruses, silencing of importin α2 (α2i) by virus‐induced gene silencing (VIGS) boosted the titre of bamboo mosaic virus (BaMV) in protoplasts, and inoculated and systemic leaves of Nicotiana benthamiana. The enhanced BaMV accumulation in importin α2i plants was linked to reduced levels of RDR6‐dependent secondary virus‐derived small‐interfering RNAs (vsiRNAs). Small RNA‐seq revealed importin α2 silencing did not affect the abundance of siRNAs derived from host mRNAs but significantly reduced the 21 and 22 nucleotide vsiRNAs in BaMV‐infected plants. Deletion of BaMV TGBp1, an RNA silencing suppressor, compromised importin α2i‐mediated BaMV enhancement. Moreover, silencing of importin α2 upregulated NbAGO10a, a proviral protein recruited by TGBp1 for BaMV vsiRNAs clearance, but hindered the nuclear import of NbAGO10a. Taken together, these results indicate that importin α2 acts as a negative regulator of BaMV invasion by controlling the expression and nucleocytoplasmic shuttling of NbAGO10a, which removes vsiRNAs via the TGBp1‐NbAGO10a‐SDN1 pathway. Our findings reveal the hidden antiviral mechanism of importin α2 in countering BaMV infection in N. benthamiana.
Silencing of importin α's enhances BaMV accumulation but reduces its virus‐derived small‐interfering RNAs (vsiRNAs) via NbAGO10a and SDN1‐mediated small RNA clearance in Nicotiana benthamiana.
Abstract
Background
Severe cutaneous adverse drug reactions (SCARs) are a group of serious clinical conditions caused by immune reaction to certain drugs. The allelic variance of human leukocyte ...antigens of HLA-B*13:01 has been strongly associated with hypersensitivities induced by dapsone (DDS). T-cell receptor mediated activation of cytotoxic T lymphocytes (CTLs) has also been suggested to play an essential role in pathogenesis of SCARs. However, HLA-B*13:01-DDS-TCR immune synapse that plays role in drug-induced hypersensitivity syndrome (DIHS) associated T cells activation remains uncharacterized.
Methods
To investigate the molecular mechanisms for HLA-B*13:01 in the pathogenesis of Dapsone-induced drug hypersensitivity (DDS-DIHS), we performed crystallization and expanded drug-specific CTLs to analyze the pathological role of DDS-DIHS.
Results
Results showed the crystal structure of HLA-B*13:01-beta-2-microglobulin (β2M) complex at 1.5 Å resolution and performed mutation assays demonstrating that I118 or I119, and R121 of HLA-B*13:01 were the key residues that mediate the binding of DDS. Subsequent single-cell TCR and RNA sequencing indicated that TCRs composed of paired TRAV12-3/TRBV28 clonotype with shared CDR3 region specifically recognize HLA-B*13:01-DDS complex to trigger inflammatory cytokines associated with DDS-DIHS.
Conclusion
Our study identified the novel p-i-HLA/TCR as the model of interaction between HLA-B*13:01, DDS and the clonotype-specific TCR in DDS-DIHS.
Graphical Abstract
Adenomyosis is a condition characterised by the invasion of endometrial tissues into the uterine myometrium, the molecular pathogenesis of which remains incompletely elucidated. Lesion profiling with ...next-generation sequencing (NGS) can lead to the identification of previously unanticipated causative genes and the detection of therapeutically actionable genetic changes. Using an NGS panel that included 275 cancer susceptibility genes, this study examined the occurrence and frequency of somatic mutations in adenomyotic tissue specimens collected from 17 women. Extracted DNA was enriched using targeted formalin-fixed paraffin-embedded tissue cores prior to the identification of lesion-specific variants. The results revealed that KRAS and AT-rich interactive domain 1A (ARID1A) were the two most frequently mutated genes (mutation frequencies: 24% and 12%, respectively). Notably, endometrial atypical hyperplasia did not involve adenomyotic areas. We also identified, for the first time, two potentially pathogenic mutations in the F-box/WD repeat-containing protein 7 (FBXW7) and cohesin subunit SA-2 (STAG2) genes. These findings indicate that mutations in the KRAS, ARID1A, FBXW7 and STAG2 genes may play a critical role in the pathogenesis of adenomyosis. Additional studies are needed to assess whether the utilisation of oncogenic driver mutations can inform the surveillance of patients with adenomyosis who had not undergone hysterectomy.
Impact statement
What is already known on this subject? Although somatic point mutations in the KRAS oncogene have been recently detected in adenomyosis, the molecular underpinnings of this condition remains incompletely elucidated. Lesion profiling with next-generation sequencing (NGS) can lead to the identification of previously unanticipated causative genes and the detection of therapeutically actionable genetic changes.
What do the results of this study add? The results of NGS revealed that KRAS and AT-rich interactive domain 1A (ARID1A) were the two most frequently mutated genes (mutation frequencies: 24% and 12%, respectively). We also identified, for the first time, two potentially pathogenic mutations in the F-box/WD repeat-containing protein 7 (FBXW7) and cohesin subunit SA-2 (STAG2) genes.
What are the implications of these findings for clinical practice and/or further research? The utilisation of oncogenic driver mutations has the potential to inform the surveillance of patients with adenomyosis who had not undergone hysterectomy.
Background Cardiac hypertrophy is associated with abnormal electrophysiology and increased arrhythmia risk. This study assessed whether candesartan cilexetil, an angiotensin II type 1 receptor ...blocker, could suppress arrhythmogenecity by attenuating cardiac electrical remodeling and calcium mishandling in rats with pressure‐overload hypertrophy. Methods and Results Male Sprague‐Dawley rats were randomly subjected to abdominal aorta banding or sham procedure and received either candesartan cilexetil (3.0 mg/kg per day) or vehicle by gavage for 5 weeks. Pressure overload was characterized by compensated left ventricular (LV) hypertrophy and fibrosis, increased LV pressure and its decay time, and prolonged corrected QT interval, all of which were attenuated by candesartan cilexetil treatment. Candesartan cilexetil–treated banded rat hearts displayed shorter QT intervals and lower vulnerability to atrial and ventricular tachyarrhythmias than vehicle‐treated banded hearts. Candesartan cilexetil prevented banding‐induced prolonged action potential duration and reduced the occurrence of triggered activity in LV papillary muscles. In addition, the prolonged time to 50% cell relengthening and calcium transient decay time were normalized in LV myocytes from candesartan cilexetil–treated banded rats, along with a normalization of decreased SERCA2a (sarcoendoplasmic reticulum calcium‐ATPase) expression in LV tissues. Furthermore, candesartan cilexetil normalized depressed transient outward potassium current densities and protein and mRNA levels of both voltage‐gated potassium 4.2 and 4.3 channel subunits (Kv4.2 and Kv4.3) in banded rats. Conclusions Candesartan cilexetil protects the heart from pressure overload‐induced adverse electrical remodeling by preserving potassium channel densities. In addition, calcium handling and its molecular regulation also improved after treatment. These beneficial effects may contribute to a lower susceptibility to arrhythmias in hearts from candesartan cilexetil–treated pressure‐overloaded rats.
The antiepileptic drug phenytoin can cause cutaneous adverse reactions, ranging from maculopapular exanthema to severe cutaneous adverse reactions, which include drug reactions with eosinophilia and ...systemic symptoms, Stevens-Johnson syndrome, and toxic epidermal necrolysis. The pharmacogenomic basis of phenytoin-related severe cutaneous adverse reactions remains unknown.
To investigate the genetic factors associated with phenytoin-related severe cutaneous adverse reactions.
Case-control study conducted in 2002-2014 among 105 cases with phenytoin-related severe cutaneous adverse reactions (n=61 Stevens-Johnson syndrome/toxic epidermal necrolysis and n=44 drug reactions with eosinophilia and systemic symptoms), 78 cases with maculopapular exanthema, 130 phenytoin-tolerant control participants, and 3655 population controls from Taiwan, Japan, and Malaysia. A genome-wide association study (GWAS), direct sequencing of the associated loci, and replication analysis were conducted using the samples from Taiwan. The initial GWAS included samples of 60 cases with phenytoin-related severe cutaneous adverse reactions and 412 population controls from Taiwan. The results were validated in (1) 30 cases with severe cutaneous adverse reactions and 130 phenytoin-tolerant controls from Taiwan, (2) 9 patients with Stevens-Johnson syndrome/toxic epidermal necrolysis and 2869 population controls from Japan, and (3) 6 cases and 374 population controls from Malaysia.
Specific genetic factors associated with phenytoin-related severe cutaneous adverse reactions.
The GWAS discovered a cluster of 16 single-nucleotide polymorphisms in CYP2C genes at 10q23.33 that reached genome-wide significance. Direct sequencing of CYP2C identified missense variant rs1057910 (CYP2C9*3) that showed significant association with phenytoin-related severe cutaneous adverse reactions (odds ratio, 12; 95% CI, 6.6-20; P=1.1 × 10(-17)). The statistically significant association between CYP2C9*3 and phenytoin-related severe cutaneous adverse reactions was observed in additional samples from Taiwan, Japan, and Malaysia. A meta-analysis using the data from the 3 populations showed an overall odds ratio of 11 (95% CI, 6.2-18; z=8.58; P < .00001) for CYP2C9*3 association with phenytoin-related severe cutaneous adverse reactions. Delayed clearance of plasma phenytoin was detected in patients with severe cutaneous adverse reactions, especially CYP2C9*3 carriers, providing a functional link of the associated variants to the disease.
This study identified CYP2C variants, including CYP2C9*3, known to reduce drug clearance, as important genetic factors associated with phenytoin-related severe cutaneous adverse reactions.
Maxillary sinus fungal balls (MSFBs) mostly occur in older individuals and demonstrate female predominance. Early diagnosis is important to avoid treatment delays. Intralesional hyperdensity (IH) ...indicates the presence of heavy metal deposition within fungal hyphae and has been the most specific characteristic of MSFB on computed tomography (CT). For those without IH on CT, the diagnosis of MSFB remains challenging. This study aimed to characterize clinical presentation of MSFB with and without IH and to study factors contributing to MSFB with no IH formation. We retrospectively identified 588 patients with MSFB. The clinical characteristics and CT findings were reviewed. Patients with unilateral MSFB had a mean age of 57.4 years and demonstrated female predominance (64.63%). The female-to-male ratio was highest at 51-60 years (2.02) and rose to 2.60 in MSFB with IH only. Compared to those with IH, MSFB without IH was significantly more common in males (OR = 2.49), in those with diabetes mellitus (DM) (OR = 1.87), adjacent maxillary odontogenic pathology (OR = 1.75). Complete opacification on CT was less common in MSFB without IH (OR = 0.60). Patients with MSFB without IH were more likely to have DM, no female predominance, adjacent maxillary odontogenic pathology, and partial opacification of the sinus, compared to those with IH. These may be helpful in better understanding of the formation of MSFBs without IH, early identification of them and prevention of post-operative recurrence.
Translocase of outer mitochondrial membrane 40 (
) is located in the outer membrane of mitochondria.
is essential for protein import into mitochondria.
genetic variants are believed to increase the ...risk of Alzheimer's disease (AD) in different populations. In this study, three exonic variants (rs772262361, rs157581, and rs11556505) and three intronic variants (rs157582, rs184017, and rs2075650) of the
gene were identified from Taiwanese AD patients using next-generation sequencing. Associations between the three
exonic variants and AD susceptibility were further evaluated in another AD cohort. Our results showed that rs157581 (c.339T > C, p.Phe113Leu, F113L) and rs11556505 (c.393C > T, p.Phe131Leu, F131L) were associated with an increased risk of AD. We further utilized cell models to examine the role of TOMM40 variation in mitochondrial dysfunction that causes microglial activation and neuroinflammation. When expressed in BV2 microglial cells, the AD-associated mutant (F113L) or (F131L) TOMM40 induced mitochondrial dysfunction and oxidative stress-induced activation of microglia and NLRP3 inflammasome. Pro-inflammatory TNF-α, IL-1β, and IL-6 released by mutant (F113L) or (F131L) TOMM40-activated BV2 microglial cells caused cell death of hippocampal neurons. Taiwanese AD patients carrying TOMM40 missense (F113L) or (F131L) variants displayed an increased plasma level of inflammatory cytokines IL-6, IL-18, IL-33, and COX-2. Our results provide evidence that TOMM40 exonic variants, including rs157581 (F113L) and rs11556505 (F131L), increase the AD risk of the Taiwanese population. Further studies suggest that AD-associated mutant (F113L) or (F131L) TOMM40 cause the neurotoxicity of hippocampal neurons by inducing the activation of microglia and NLRP3 inflammasome and the release of pro-inflammatory cytokines.
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