Cases of endophthalmitis caused by Candida albicans and Candida dubliniensis identified via internal transcribed spacer deep sequencing.

BACKGROUND: We report two cases of fungal endophthalmitis induced by Candida species identified based on internal transcribed spacer 1 (ITS1) sequencing. CASE PRESENTATION: In two cases, endophthalmitis was suspected, and the patients underwent pars plana vitrectomy. Case 1 was a 64-year-old woman with a history of cataract surgery 10 days prior. She had a history of anal primary melanoma, which metastasized throughout the body and subsequently relapsed. Vitreous culture and ITS-1 deep sequencing revealed the presence of the rare fungus, Candida dubliniensis. Case 2 was a 54-year-old man with a history of liver cancer and kidney failure. Culture methods and ITS1 deep sequencing both revealed the presence of Candida albicans. Both patients exhibited good visual prognoses after treatment with topical and systemic antibiotics. CONCLUSIONS: We present two cases of fungal endophthalmitis caused by two Candida species identified by both the culture method and ITS1 deep sequencing. The fungal pathogen was identified by ITS deep sequencing three days after sample submission; the culture method yielded results after 1 week. These findings support the applicability of ITS1 sequencing for timely pathogen identification for cases of fungal endophthalmitis and provide detailed taxonomic information at the species level.

Variability of macrolide-resistant profile in Mycobacterium avium complex pulmonary disease.

This single-center retrospective study aimed to analyze the variability of macrolide resistance (MR) in 68 patients with Mycobacterium avium complex pulmonary disease. Among 25 patients treated without macrolides, 13 (52%) reverted to macrolide-susceptible (MS) profiles. Only one (2%) of 43 patients who continued macrolide treatment showed this change. We compared 30 MR isolates with recent specimens. Among them, seven shifted to MS (five attributed to clonally related strains; two resulting from reinfection or polyclonal infection).

Neoself-antigens are the primary target for autoreactive T cells in human lupus.

Major histocompatibility complex class II (MHC-II) is the most significant genetic risk factor for systemic lupus erythematosus (SLE), but the nature of the self-antigens that trigger autoimmunity remains unclear. Unusual self-antigens, termed neoself-antigens, are presented on MHC-II in the absence of the invariant chain essential for peptide presentation. Here, we demonstrate that neoself-antigens are the primary target for autoreactive T cells clonally expanded in SLE. When neoself-antigen presentation was induced by deleting the invariant chain in adult mice, neoself-reactive T cells were clonally expanded, leading to the development of lupus-like disease. Furthermore, we found that neoself-reactive CD4+ T cells were significantly expanded in SLE patients. A high frequency of Epstein-Barr virus reactivation is a risk factor for SLE. Neoself-reactive lupus T cells were activated by Epstein-Barr-virus-reactivated cells through downregulation of the invariant chain. Together, our findings imply that neoself-antigen presentation by MHC-II plays a crucial role in the pathogenesis of SLE.

Gut Dysbiosis in Patients With Fontan Circulation.

BACKGROUND: The process underlying Fontan pathophysiology is multifactorial and may include gut dysbiosis (GD). We investigated the presence of GD and elucidated its correlation with Fontan pathophysiology. METHODS AND RESULTS: Gut microbiomes of 155 consecutive patients with Fontan pathophysiology and 44 healthy individuals were analyzed using 16S rRNA sequencing of bacterial DNA extracted from fecal samples. GD was evaluated on the basis of α and ß diversities of the gut microbiome and was compared with natural log-transformed C-reactive protein, hemodynamics, von Willebrand factor antigen (a bacterial translocation marker), Mac-2 binding protein glycosylation isomer (a liver fibrosis indicator), peak oxygen uptake, and heart failure hospitalization. Patients with Fontan exhibited GD in terms of α and ß diversities as compared with controls (P<0.01). Reduced α diversity was associated with a failed hemodynamic phenotype, hypoxia, high natural log-transformed C-reactive protein levels, and elevated von Willebrand factor antigen and Mac-2 binding protein glycosylation isomer levels (P<0.05-0.01). In addition to elevated von Willebrand factor antigen and hypoxia, decreased α diversity was independently correlated with a high natural log-transformed C-reactive protein level (P<0.05), which was associated with liver imaging abnormalities and a heightened risk of heart failure hospitalization (P<0.01 for both). CONCLUSIONS: Patients with Fontan pathophysiology exhibited GD compared with healthy individuals, and GD was linked to failed hemodynamics and systemic inflammation with a poor prognosis. Therefore, GD may play a pivotal role in a failing Fontan status, including Fontan-associated liver disease, through GD-associated systemic inflammation.

Clonal landscape of autoantibody-secreting plasmablasts in COVID-19 patients.

Whereas severe COVID-19 is often associated with elevated autoantibody titers, the underlying mechanism behind their generation has remained unclear. Here we report clonal composition and diversity of autoantibodies in humoral response to SARS-CoV-2. Immunoglobulin repertoire analysis and characterization of plasmablast-derived monoclonal antibodies uncovered clonal expansion of plasmablasts producing cardiolipin (CL)-reactive autoantibodies. Half of the expanded CL-reactive clones exhibited strong binding to SARS-CoV-2 antigens. One such clone, CoV1804, was reactive to both CL and viral nucleocapsid (N), and further showed anti-nucleolar activity in human cells. Notably, antibodies sharing genetic features with CoV1804 were identified in COVID-19 patient-derived immunoglobulins, thereby constituting a novel public antibody. These public autoantibodies had numerous mutations that unambiguously enhanced anti-N reactivity, when causing fluctuations in anti-CL reactivity along with the acquisition of additional self-reactivities, such as anti-nucleolar activity, in the progeny. Thus, potentially CL-reactive precursors may have developed multiple self-reactivities through clonal selection, expansion, and somatic hypermutation driven by viral antigens. Our results revealed the nature of autoantibody production during COVID-19 and provided novel insights into the origin of virus-induced autoantibodies.

A plasmid-mediated type III secretion system associated with invasiveness and diarrheagenicity of Providencia rustigianii.

We have recently described a clinical isolate of Providencia rustigianii strain JH-1 carrying the genes for cytolethal distending toxin (CDT) in a conjugative plasmid. A cdtB mutant of strain JH-1, which lost CDT activity, was still found to retain invasiveness and diarrheagenicity. The strain was subjected to phenotypic and genetic analyses including whole genome sequencing (WGS) to explore the genetic determinants of the observed invasiveness and diarrheagenic properties. Analysis and annotation of WGS data revealed the presence of two distinct type III secretion systems (T3SS) in strain JH-1, one of which was located on the chromosome designated as cT3SS (3,992,833 bp) and the other on a mega-plasmid designated as pT3SS (168,819 bp). Comparative genomic analysis revealed that cT3SS is generally conserved in Providencia spp. but pT3SS was limited to a subset of Providencia spp., carrying cdt genes. Strain JH-1 was found to invade HeLa cells and induce fluid accumulation with characteristic pathological lesions in rabbit ileal loops. Remarkably, these phenomena were associated with the pT3SS but not cT3SS. The plasmid could be transferred by conjugation from strain JH-1 to other strains of P. rustigianii, Providencia rettgeri, and Escherichia coli with concomitant transfer of these virulence properties. This is the first report of a functional and mobile T3SS in P. rustigianii and its association with invasiveness and diarrheagenicity of this bacterium. These data suggest that P. rustigianii and other CDT-producing Providencia strains might carry T3SS and exert their diarrheagenic effect by exploiting the T3SS nano-machinery.IMPORTANCEThe precise mechanism of virulence of Providencia rustigianii is unclear, although some strains produce cytolethal distending toxin as a putative virulence factor. We have detected the presence of a type III secretion system (T3SS) for the first time on a plasmid in a P. rustigianii strain. Plasmid-mediated T3SS seems to be directly involved in virulence of P. rustigianii and may serve as a means of horizontal transfer of T3SS genes. Our results may have implication in understanding the mechanism of emergence of new pathogenic strains of P. rustigianii.

Intrahepatic exhausted antiviral immunity in an immunocompetent mouse model of chronic hepatitis B.

BACKGROUND & AIMS: Targeting exhausted immune systems would be a promising therapeutic strategy to achieve a functional cure for HBV infection in chronic hepatitis B (CHB) patients. However, animal models recapitulating the immunokinetics of CHB are very limited. We aimed to develop an immunocompetent mouse model of CHB for intrahepatic immune profiling. METHODS: CHB mice were created by intrahepatic delivery of the Sleeping Beauty transposon vector tandemly expressing the hepatitis B virus (HBV) genome and fumarylacetoacetate hydrolase (FAH) cDNA into C57BL/6J congenic FAH knockout mice via hydrodynamic tail vein injection. We profiled the viral and intrahepatic immune kinetics in CHB mice with or without treatment with recombinant IFNα or the hepatotropic Toll-like receptor 7 agonist SA-5 using single-cell RNA-seq. RESULTS: CHB mice exhibited sustained HBV viremia and persistent hepatitis. They showed intrahepatic expansion of exhausted CD8+ T (Tex) cells, the frequency of which was positively associated with viral load. Recruited macrophages increased in number but impaired inflammatory responses in the liver. The cytotoxicity of mature NK cells also increased in CHB mice. IFNα and SA-5 treatment both resulted in viral suppression with mild hepatic flares in CHB mice. While both treatments activated NK cells, SA-5 had the capacity to revitalize the impaired function of Tex cells and liver-recruited macrophages. CONCLUSION: Our novel CHB mouse model recapitulated the intrahepatic exhausted antiviral immunity in CHB patients, which might be able to be reinvigorated by a hepatotropic TLR7 agonist.

Full-length nanopore sequencing of circular RNA landscape in peripheral blood cells following sequential BNT162b2 mRNA vaccination.

Circular RNAs (circRNA) lack 5' or 3' ends; their unique covalently closed structures prevent RNA degradation by exonucleases. These characteristics provide circRNAs with high pharmaceutical stability and biostability relative to current standard-of-care linear mRNAs. CircRNA levels are reportedly associated with certain human diseases, making them novel disease biomarkers and a noncanonical class of therapeutic targets. In this study, the endogenous circRNAs underlying the response to BNT162b2 mRNA vaccination were evaluated. To this end, peripheral blood samples were subjected to full-length sequencing of circRNAs via nanopore sequencing and transcriptome sequencing. Fifteen samples, comprising pre-, first, and second vaccination cohorts, were obtained from five healthcare workers with no history of SARS-CoV-2 infection or previous vaccination. A total of 4706 circRNAs were detected; following full-length sequencing, 4217 novel circRNAs were identified as being specifically expressed during vaccination. These circRNAs were enriched in the binding motifs of stress granule assemblies and SARS-CoV-2 RNA binding proteins, namely poly(A) binding protein cytoplasmic 1 (PABPC1), pumilio RNA binding family member 1 (PUM1), and Y box binding protein 1 (YBX1). Moreover, 489 circRNAs were identified as previously reported miRNA sponges. The differentially expressed circRNAs putatively originated from plasma B cells compared to circRNAs reported in human blood single-cell RNA sequencing datasets. The pre- and post-vaccination differences observed in the circRNA expression landscape in response to the SARS-CoV-2 BNT162b2 mRNA vaccine.

Dysbiosis of the human skin mycobiome in patients receiving systemic IL-23 inhibitors.

BACKGROUND: Systemic inhibition of pro-inflammatory cytokines affects the skin microbiome; however, the impact of systemic anti-inflammatory therapy on the skin fungal microbiome is poorly understood. To examine the effects of cytokine inhibition on the fungal community on human skin and oral mucosa, we analyzed the composition of the skin mycobiome before and after IL-23 inhibition. METHODS: The study enrolled 15 psoriasis patients. Swab samples were collected from the psoriasis-free skin of antecubital fossa, post-auricular, and the tongue surface before and after 16 weeks of treatment with anti-IL-23 antibodies. Fungal DNA was sequenced by ITS1 metagenomic analysis, and taxonomic classification was performed. RESULTS: Data from samples collected from the antecubital fossa revealed that the α diversity of the skin mycobiome decreased significantly after treatment with anti-IL-23 antibodies (p = 0.0120). Fungal DNAs were not amplified in 6/15 swab samples after 16 weeks of IL-23 inhibition; by contrast, sufficiently detected in all 15 samples before treatment (p = 0.0554). A comparison of 9/15 paired samples containing well-detected reads revealed that the percentage of genus Malassezia in the mycobiome fell significantly after treatment with IL-23 inhibitors (before, 29.3% ± 9.9%; after; 8.5% ± 3.4%, p = 0.0137). The mycobiome on post-auricular skin and on the tongue surface showed no marked changes after IL-23 inhibition. CONCLUSIONS: Taken together, the data suggest that inhibition of systemic IL-23 provokes dysbiosis of the mycobiome at the antecubital fossa skin, a finding characterized by reduced fungal diversity and a reduction in the percentage of the genus Malassezia.

Paraclostridium tenue Causing an Anaerobic Brain Abscess Identified by Whole-Metagenome Sequencing: A Case Report.

When treating anaerobic brain abscesses, healthcare professionals often face the difficulty of identifying the causal pathogens, necessitating empiric therapies with uncertain efficacy. We present the case of a 57-year-old woman who was admitted to our hospital with a fever and headache. Brain magnetic resonance imaging revealed a hemorrhagic lesion with wall enhancement at the left hemisphere on contrast-enhanced T1-weighted imaging. Cerebrospinal fluid examination showed pleocytosis (23 cells/µL), an elevated protein level (125 mg/dL), and decreased glucose level (51 mg/dL; blood glucose was 128 mg/dL). Intracerebral hemorrhage accompanied by a brain abscess was clinically suspected. The patient received empirical treatment with intravenous meropenem and vancomycin for 2 weeks. However, conventional bacterial culture tests failed to identify the pathogen. We then performed shotgun sequencing and ribosomal multilocus sequence typing, which identified Paraclostridium tenue. Based on this finding, we de-escalated to benzylpenicillin potassium for 4 weeks, leading to a 2.5-year remission of the anaerobic brain abscess. Therefore, Paraclostridium can be a causative pathogen for brain abscesses. Furthermore, whole-metagenome sequencing is a promising method for detecting rare pathogens that are not identifiable by conventional bacterial culture tests. This approach enables more targeted treatment and contributes to achieving long-term remission in clinical settings.

Spatial transcriptomics elucidates medulla niche supporting germinal center response in myasthenia gravis-associated thymoma.

Myasthenia gravis (MG) is etiologically associated with thymus abnormalities, but its pathology in the thymus remains unclear. In this study, we attempt to narrow down the features associated with MG using spatial transcriptome analysis of thymoma and thymic hyperplasia samples. We find that the majority of thymomas are constituted by the cortical region. However, the small medullary region is enlarged in seropositive thymomas and contains polygenic enrichment and MG-specific germinal center structures. Neuromuscular medullary thymic epithelial cells, previously identified as MG-specific autoantigen-producing cells, are enriched in the cortico-medullary junction. The medulla is characterized by a specific chemokine pattern and immune cell composition, including migratory dendritic cells and effector regulatory T cells. Similar germinal center structures and immune microenvironments are also observed in the thymic hyperplasia medulla. This study shows that the medulla and junction areas are linked to MG pathology and provides insights into future MG research.

Identification of causative fungus from sterile abscess using metagenomics followed by in situ hybridization.

Introduction. Invasive fungal infections require early diagnosis for treatment. Microscopic observation of biopsy and blood culture is the gold standard for the identification of the causative fungus, but it is difficult to identify the causative pathogen by a sterile abscess biopsy. Case Presentation. We present a case report of breakthrough invasive trichosporonosis in a 65-year-old Japanese male with acute myeloid leukaemia receiving antifungal prophylaxis. Blood cultures showed no fungal growth, and a liver biopsy and a removed spleen with abscess showed fragmented fungi, but no fungal identification was possible. This report demonstrates that retrospective analyses were able to identify the causative fungus. Conclusion. We narrowed down the candidate fungi by deep sequencing of the ITS1 region of fungal genome and confirmed that the fungus observed in the specimen was Trichosporon asahii by in situ hybridization using a DNA probe targeting 26S rRNA.

Significance of signal recognition particle 9 nuclear translocation: Implications for pancreatic cancer prognosis and functionality.

Signal recognition particles (SRPs) are essential for regulating intracellular protein transport and secretion. Patients with tumors with high SRP9 expression tend to have a poorer overall survival. However, to the best of our knowledge, no reports have described the relationship between SRP9 localization and prognosis in pancreatic cancer. Thus, the present study aimed to investigate this relationship. Immunohistochemical staining for SRP9 using excised specimens from pancreatic cancer surgery cases without preoperative chemotherapy or radiotherapy showed that SRP9 was preferentially expressed in the nucleus of the cancerous regions in some cases, which was hardly detected in other cases, indicating that SRP9 was transported to the nucleus in the former cases. To compare the prognosis of patients with SRP9 nuclear translocation, patients were divided into two groups: Those with a nuclear translocation rate of >50% and those with a nuclear translocation rate of ≤50%. The nuclear translocation rate of >50% group had a significantly better recurrence­free survival than the nuclear translocation rate of ≤50% group (P=0.037). Subsequent in vitro experiments were conducted; notably, the nuclear translocation rate of SRP9 was reduced under amino acid­deficient conditions, suggesting that multiple factors are involved in this phenomenon. To further study the function of SRP9 nuclear translocation, in vitro experiments were performed by introducing SRP9 splicing variants (v1 and v2) and their deletion mutants lacking C­terminal regions into MiaPaCa pancreatic cancer cells. The results demonstrated that both splicing variants showed nuclear translocation regardless of the C­terminal deletions, suggesting the role of the N­terminal regions. Given that SRP9 is an RNA­binding protein, the study of RNA immunoprecipitation revealed that signaling pathways involved in cancer progression and protein translation were downregulated in nuclear­translocated v1 and v2. Undoubtedly, further studies of the nuclear translocation of SRP9 will open an avenue to optimize the precise evaluation and therapeutic control of pancreatic cancer.

Far-East Asian Toxoplasma isolates share ancestry with North and South/Central American recombinant lineages.

Toxoplasma gondii is a global protozoan pathogen. Clonal lineages predominate in Europe, North America, Africa, and China, whereas highly recombinant parasites are endemic in South/Central America. Far East Asian T. gondii isolates are not included in current global population genetic structure analyses at WGS resolution. Here we report a genome-wide population study that compared eight Japanese and two Chinese isolates against representative worldwide T. gondii genomes using POPSICLE, a novel population structure analyzing software. Also included were 7 genomes resurrected from non-viable isolates by target enrichment sequencing. Visualization of the genome structure by POPSICLE shows a mixture of Chinese haplogroup (HG) 13 haploblocks introgressed within the genomes of Japanese HG2 and North American HG12. Furthermore, two ancestral lineages were identified in the Japanese strains; one lineage shares a common ancestor with HG11 found in both Japanese strains and North American HG12. The other ancestral lineage, found in T. gondii isolates from a small island in Japan, is admixed with genetically diversified South/Central American strains. Taken together, this study suggests multiple ancestral links between Far East Asian and American T. gondii strains and provides insight into the transmission history of this cosmopolitan organism.

Deletion of the mRNA endonuclease Regnase-1 promotes NK cell anti-tumor activity via OCT2-dependent transcription of Ifng.

Limited infiltration and activity of natural killer (NK) and T cells within the tumor microenvironment (TME) correlate with poor immunotherapy responses. Here, we examined the role of the endonuclease Regnase-1 on NK cell anti-tumor activity. NK cell-specific deletion of Regnase-1 (Reg1ΔNK) augmented cytolytic activity and interferon-gamma (IFN-γ) production in vitro and increased intra-tumoral accumulation of Reg1ΔNK-NK cells in vivo, reducing tumor growth dependent on IFN-γ. Transcriptional changes in Reg1ΔNK-NK cells included elevated IFN-γ expression, cytolytic effectors, and the chemokine receptor CXCR6. IFN-γ induced expression of the CXCR6 ligand CXCL16 on myeloid cells, promoting further recruitment of Reg1ΔNK-NK cells. Mechanistically, Regnase-1 deletion increased its targets, the transcriptional regulators OCT2 and IκBζ, following interleukin (IL)-12 and IL-18 stimulation, and the resulting OCT2-IκBζ-NF-κB complex induced Ifng transcription. Silencing Regnase-1 in human NK cells increased the expression of IFNG and POU2F2. Our findings highlight NK cell dysfunction in the TME and propose that targeting Regnase-1 could augment active NK cell persistence for cancer immunotherapy.

Turicibacter faecis sp. nov., isolated from faeces of heart failure mouse model.

Strain TC023T, a Gram-positive, long, rod-shaped, spore-forming anaerobe, was isolated from the faeces of a heart failure mouse model. The strain formed greyish-white coloured colonies with a convex elevation on brain-heart infusion medium supplemented with 0.1 % sodium taurocholate, incubated at 37 °C for 2 days. Taxonomic analysis based on the 16S rRNA gene sequence showed that TC023T belonged to the genus Turicibacter, and was closely related to Turicibacter bilis MMM721T (97.6 %) and Turicibacter sanguinis MOL361T (97.4 %). The whole genome of the strain has a G+C content of 37.3 mol%. The average nucleotide identity and genome-to-genome distance between TC023T and Turicibacter bilis MMM721T were 77.6 % and 24.3 %, respectively, and those with Turicibacter sanguinis MOL361T were 75.4 % and 24.3 %, respectively. These genotypic, phenotypic, and biochemical analyses indicated that the isolate represents a novel species in the genus Turicibacter, and the name Turicibacter faecis sp. nov. is proposed. The type strain is TC023T (RIMD 2002001T=TSD 372T).

Regulation of muscle hypertrophy through granulin: Relayed communication among mesenchymal progenitors, macrophages, and satellite cells.

Skeletal muscles exert remarkable regenerative or adaptive capacities in response to injuries or mechanical loads. However, the cellular networks underlying muscle adaptation are poorly understood compared to those underlying muscle regeneration. We employed single-cell RNA sequencing to investigate the gene expression patterns and cellular networks activated in overloaded muscles and compared these results with those observed in regenerating muscles. The cellular composition of the 4-day overloaded muscle, when macrophage infiltration peaked, closely resembled that of the 10-day regenerating muscle. In addition to the mesenchymal progenitor-muscle satellite cell (MuSC) axis, interactome analyses or targeted depletion experiments revealed communications between mesenchymal progenitors-macrophages and macrophages-MuSCs. Furthermore, granulin, a macrophage-derived factor, inhibited MuSC differentiation, and Granulin-knockout mice exhibited blunted muscle hypertrophy due to the premature differentiation of overloaded MuSCs. These findings reveal the critical role of granulin through the relayed communications of mesenchymal progenitors, macrophages, and MuSCs in facilitating efficient muscle hypertrophy.

Periportal macrophages protect against commensal-driven liver inflammation.

The liver is the main gateway from the gut, and the unidirectional sinusoidal flow from portal to central veins constitutes heterogenous zones, including the periportal vein (PV) and the pericentral vein zones1-5. However, functional differences in the immune system in each zone remain poorly understood. Here intravital imaging revealed that inflammatory responses are suppressed in PV zones. Zone-specific single-cell transcriptomics detected a subset of immunosuppressive macrophages enriched in PV zones that express high levels of interleukin-10 and Marco, a scavenger receptor that sequesters pro-inflammatory pathogen-associated molecular patterns and damage-associated molecular patterns, and consequently suppress immune responses. Induction of Marco+ immunosuppressive macrophages depended on gut microbiota. In particular, a specific bacterial family, Odoribacteraceae, was identified to induce this macrophage subset through its postbiotic isoallolithocholic acid. Intestinal barrier leakage resulted in inflammation in PV zones, which was markedly augmented in Marco-deficient conditions. Chronic liver inflammatory diseases such as primary sclerosing cholangitis (PSC) and non-alcoholic steatohepatitis (NASH) showed decreased numbers of Marco+ macrophages. Functional ablation of Marco+ macrophages led to PSC-like inflammatory phenotypes related to colitis and exacerbated steatosis in NASH in animal experimental models. Collectively, commensal bacteria induce Marco+ immunosuppressive macrophages, which consequently limit excessive inflammation at the gateway of the liver. Failure of this self-limiting system promotes hepatic inflammatory disorders such as PSC and NASH.

Squamous cell carcinoma initially occurring on the tongue dorsum: a case series report with molecular analysis.

BACKGROUND: Squamous cell carcinoma (SCC) of the dorsum of the tongue is extremely rare, and it clinically resembles various benign lesions. Somatic mutations in TP53 and some driver genes were implicated in the development of SCC; however, the somatic genetic characteristics of dorsal tongue SCC remain unknown. With a detailed analysis of gene mutations in dorsal tongue SCC, we aimed to better understand its biology. METHODS: Four cases of SCC initially occurring on the tongue dorsum were evaluated for clinical and histological findings and immunohistochemical expression of p53 and p16. Gene mutations were analyzed using next-generation sequencing with a custom panel of driver genes. RESULTS: We retrospectively investigated 557 cases of tongue SCC, and only four cases of SCC initially occurred on the tongue dorsum. The four patients (cases 1-4) were one woman and three men with a mean age of 53.75 years (range: 15-74 years). Histological analysis revealed well-differentiated SCC. Through molecular analysis, we identified pathogenic somatic mutations, namely, TP53 p.C176F (c.527G > T) in case 3 and TP53 p.R282W (c.844 C > T) in case 4. No pathogenic variants were identified in the PI3K/AKT or RAS/RAF pathways. The p53 immunohistochemical examination revealed a wild-type expression pattern in cases 1-3 and strong expression in case 4. The results of p16 immunostaining were negative in all cases. CONCLUSIONS: We described four previously unreported genetic characteristics of dorsal tongue SCC. Somatic TP53 mutations may contribute to the development of a subset of dorsal tongue SCC; however, more cases with genetic analysis need to be accumulated.

High N6-methyladenosine-activated TCEAL8 mRNA is a novel pancreatic cancer marker.

N6-methyladenosine (m6A) is an RNA modification involved in RNA processing and widely found in transcripts. In cancer cells, m6A is upregulated, contributing to their malignant transformation. In this study, we analyzed gene expression and m6A modification in cancer tissues, ducts, and acinar cells derived from pancreatic cancer patients using MeRIP-seq. We found that dozens of RNAs highly modified by m6A were detected in cancer tissues compared with ducts and acinar cells. Among them, the m6A-activated mRNA TCEAL8 was observed, for the first time, as a potential marker gene in pancreatic cancer. Spatially resolved transcriptomic analysis showed that TCEAL8 was highly expressed in specific cells, and activation of cancer-related signaling pathways was observed relative to TCEAL8-negative cells. Furthermore, among TCEAL8-positive cells, the cells expressing the m6A-modifying enzyme gene METTL3 showed co-activation of Notch and mTOR signaling, also known to be involved in cancer metastasis. Overall, these results suggest that m6A-activated TCEAL8 is a novel marker gene involved in the malignant transformation of pancreatic cancer.