Background: DNA methylation patterns are initiated by de novo DNA methyltransferases DNMT3a/3b adding methyl groups to CG dinucleotides in the hypomethylated genome of early embryos. These patterns are faithfully maintained by DNMT1 during DNA replication to ensure epigenetic inheritance across generations. However, this two-step model is based on limited data. Results: We generated base-resolution DNA methylomes for a series of DNMT knockout embryonic stem cells, with deep coverage at highly repetitive elements. We show that DNMT1 and DNMT3a/3b activities work complementarily and simultaneously to establish symmetric CG methylation and CHH (H = A, T or C) methylation. DNMT3a/3b can add methyl groups to daughter strands after each cycle of DNA replication. We also observe an unexpected division of labor between DNMT1 and DNMT3a/3b in suppressing retrotransposon long terminal repeats and long interspersed elements, respectively. Our data suggest that mammalian cells use a specific CG density threshold to predetermine methylation levels in wild-type cells and the magnitude of methylation reduction in DNMT knockout cells. Only genes with low CG density can be induced or, surprisingly, suppressed in the hypomethylated genome. Lastly, we do not find any association between gene body methylation and transcriptional activity. Conclusions: We show the concerted actions of DNMT enzymes in the establishment and maintenance of methylation patterns. The finding of distinct roles of DNMT1-dependent and -independent methylation patterns in genome stability and regulation of transcription provides new insights for understanding germ cell development, neuronal diversity, and transgenerational epigenetic inheritance and will help to develop next-generation DNMT inhibitors.

Ewing’s sarcoma (ES) and primitive neuroectodermal tumors (PNET) are closely related tumors. Although soft tissue ES/PNET are common in clinical practice, they are rare in the small intestine. Because of the absence of characteristic clinical symptoms, they are easily misdiagnosed as other benign or malignant diseases. Here, we present the case of a 16-year-old female who complained of anemia and interval hematochezia. Her serum test results showed only a slight elevation of CA-125 and a low level of hemoglobin. Computer tomography and magnetic resonance imaging revealed a cystic and solid mass in the lower abdominal quadrant and pelvic region, which prompted suspicion of a malignant gastrointestinal stromal tumor of the small intestine. After resection, the tumor’s histology and immunohistochemistry (positive for CD99, vimentin and synaptophysin) results suggested ES/PNET. Fluorescent in situ hybridization tests proved the breakpoint rearrangement of the EWSR1 gene in chr 22.Ultrastructural analysis revealed neurosecretory and glycogen granules in the tumor cell cytoplasm. Together, these data supported the diagnosis of a rare case of localized ES/PNET in the small intestine without adjuvant chemo- or radiotherapy. To our knowledge, this is the first report from China of a primary small bowel ES/PNET in the English-language literature. In addition, on the basis of findings from previous publications and the current case, the optimal treatment for localized gastrointestinal ES/PNET is discussed.

Background. B7 family members and ligands have been identified as critical checkpoints in orchestrating the immune response during sepsis. V‐domain Ig suppressor of T cell activation (VISTA) is a new inhibitory immune checkpoint involved in restraining T cell response. Previous studies demonstrated that VISTA engagement on T cells and myeloid cells could transmit inhibitory signals, resulting in reduced activation and function. The current study was designed to determine the potential therapeutic effects of a high‐affinity anti‐VISTA antibody (clone MH5A) in a murine model of sepsis. Methods. Polymicrobial sepsis was induced in male C57BL/6 mice via cecal ligation and puncture. Expression profiles of VISTA on T lymphocytes and macrophage were examined at 24 and 72 h postsurgery. The effects of anti‐VISTA mAb on the 7‐day survival, lymphocyte apoptosis, cytokine expression, bacterial burden, and vital organ damage were determined. Furthermore, the effects of anti‐VISTA mAb on CD3⁺ T cell apoptosis and macrophage activation were determined in vitro. Results. VISTA was substantially expressed on T cells and macrophages in sham‐operated mice; septic peritonitis did not induce significant changes in the expression profiles. Treatment with MH5A improved the survival of septic mice, accompanied by reduced lymphocyte apoptosis, decreased cytokine expression, and enhanced bacterial clearance. Engagement of VISTA receptor with MH5A mitigated CD3⁺ T cell apoptosis cultured from CLP mice and suppressed LPS‐induced cytokine production by macrophage in vitro. Conclusion. The present study identified VISTA as a novel immune checkpoint in the regulation of T cell and macrophage response during sepsis. Modulation of the VISTA pathway might offer a promising opportunity in the immunotherapy for sepsis.

Peutz-Jeghers syndrome (PJS) is a rare, autosomal dominant disorder caused by germline mutations of STK11/LKB1, with an increased risk of tumors at multiple sites. Intraductal oncocytic papillary neoplasm (IOPN) is a unique subtype of intraductal papillary neoplasm of the bile duct (IPNB) defined by a premalignant neoplasm with intraductal papillary or villous growth of biliary-type epithelium. IOPN has a distinct mutation profile compared with both IPNB and intraductal papillary mucinous neoplasm (IPMN). We herein describe the case of a 44-year-old woman who presented as polyps in the intestinal lumen of sigmoid colon and a 3.1 × 2.1 cm mass in the left lobe of liver. Gross feature revealed a cystic papillary mass and the neoplasm had a clear boundary with the surrounding liver tissue. Histology revealed complex papillary structures, a small amount of fine fibrovascular cores and immunohistochemistry showed extensive positive for MUC5AC, MUC6, CD117. Therefore, histological and immunohistochemical examination of the liver tumor suggested the diagnosis of IOPN. Next-generation sequencing (NGS) revealed other than STK11 germline mutation, the tumor also harbors GNAS somatic mutation at codon 478 and EGFR amplification. To our knowledge, this is the first report of IOPN arising in PJS. This case enlarges the spectrum of PJS related tumors and genetic rearrangements in IOPN.

The glioma tumor microenvironment (TME) exhibits profound heterogeneity that drives tumor progression and therapy resistance. By integrating single-cell RNA sequencing (eleven samples) and spatial transcriptomics (two samples), the cellular components of the glioma microenvironment were deconvoluted, revealing tumor-associated foam cells (TAFCs) as the most abundant and centrally connected subtype. The high expression of two prognostic candidate genes, growth differentiation factor 8 (GDF-8, also known as myostatin, MSTN) and transcription factor 12 (TCF12), in TAFCs was found to be correlated with poor overall survival. These two genes were associated with M2 macrophage infiltration, altered cholesterol homeostasis, and immunosuppressive signaling. Regulatory network and pathway analyses, based on computational motif enrichment and co-expression analysis, linked them to ribosome, Notch signaling, DNA repair, and cell-cycle pathways. Pseudotime trajectories revealed dynamic expression during differentiation. Additionally, drug sensitivity prediction analysis demonstrated that MSTN expression was significantly associated with sensitivity to paclitaxel and VE-822, while TCF12 expression showed potential associations with sensitivity to cytarabine, olaparib, Wee1 inhibitor, paclitaxel, and VE-822. Logistic regression analysis combining clinical parameters with MSTN and TCF12 expression effectively achieved risk stratification for glioma, with higher composite scores predicting worse 2- and 3-year survival outcomes. Calibration curves demonstrated high consistency between nomogram-predicted overall survival probabilities and actual observed outcomes. Immunofluorescence confirmed upregulated expression of MSTN and TCF12 in glioma tissues and their co-localization with macrophages. In conclusion, this study identified TAFCs as the central cells in the glioma microenvironment, with their signature genes MSTN and TCF12 representing candidate immunometabolic signatures associated with macrophage-mediated immunosuppression and metabolic reprogramming in glioma, suggesting their potential as biomarkers for patient stratification and as targets for immunometabolic therapies.