The First Reported Case of Treating the Ultra-Central Thorax With Cone Beam Computed Tomography-Guided Stereotactic Adaptive Radiotherapy (CT-STAR).

Stereotactic body radiotherapy (SBRT) to the central and ultra-central thorax is associated with infrequent but potentially serious adverse events. Adaptive SBRT, which provides more precise treatment planning and inter-fraction motion management, may allow the delivery of ablative doses to ultra-central tumors with effective local control and improved toxicity profiles. Herein, we describe the first reported case of cone beam computed tomography (CBCT)-guided stereotactic adaptive radiotherapy (CT-STAR) in the treatment of ultra-central non-small cell lung cancer (NSCLC) in a prospective clinical trial (NCT05785845). An 80-year-old man with radiographically diagnosed early-stage NSCLC presented for definitive management of an enlarging ultra-central lung nodule. He was prescribed 55 Gy in five fractions with CT-STAR. A simulation was performed using four-dimensional CT, and patients were planned for treatment at end-exhale breath-hold. Treatment plans were generated using a strict isotoxicity approach, which prioritized organ at risk (OAR) constraints over target coverage. During treatment, daily CBCTs were acquired and used to generate adapted contours and treatment plans based on the patient's anatomy-of-the-day, all while the patient was on the treatment table. The initial and adapted plans were compared using dose-volume histograms, and the superior plan was selected for treatment. The adapted plan was deemed superior and used for treatment in three out of five fractions. The adapted plan provided improved target coverage in two fractions and resolved an OAR hard constraint violation in one fraction. We report the successful treatment of a patient with ultra-central NSCLC utilizing CT-STAR. This case report builds on previously published in silico data to support the viability and dosimetric advantages of CT-STAR in the ablative treatment of this challenging tumor location. Further data are needed to confirm the toxicity and efficacy of this technique.

Activation of non-classical Wnt signaling pathway effectively enhances HLA-A presentation in acute myeloid leukemia.

Objective: The escape from T cell-mediated immune surveillance is an important cause of death for patients with acute myeloid leukemia (AML). This study aims to identify clonal heterogeneity in leukemia progenitor cells and explore molecular or signaling pathways associated with AML immune escape. Methods: Single-cell RNA sequencing (scRNA-seq) was performed to identified AML-related cellular subsets, and intercellular communication was analyzed to investigate molecular mechanisms associated with AML immune escape. Bulk RNA sequencing (RNA-seq) was performed to screen differentially expressed genes (DEGs) related to hematopoietic stem cell progenitors (HSC-Prog) in AML, and critical ore signaling pathways and hub genes were found by Gene Set Enrichment Analysis (GSEA), Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. The mRNA level of the hub gene was verified using quantitative real-time PCR (qRT-PCR) and the protein level of human leukocyte antigen A (HLA-A) using enzyme-linked immuno sorbent assay (ELISA). Results: scRNA-seq analysis revealed a large heterogeneity of HSC-Prog across samples, and the intercellular communication analysis indicated a strong association between HSC-Prog and CD8+-T cells, and HSC-Prog also had an association with HLA-A. Transcriptome analysis identified 1748 DEGs, enrichment analysis results showed that non-classical wnt signaling pathway was associated with AML, and 4 pathway-related genes (RHOA, RYK, CSNK1D, NLK) were obtained. After qRT-PCR and ELISA validation, hub genes and HLA-A were found to be down-regulated in AML and up-regulated after activation of the non-classical Wnt signaling pathway. Conclusion: In this study, clonal heterogeneity of HSC-Prog cells in AML was identified, non-classical wnt signaling pathways associated with AML were identified, and it was verified that HLA-A could be upregulated by activation of non-classical wnt signaling, thereby increasing antigen presentation.

Unravelling tumour cell diversity and prognostic signatures in cutaneous melanoma through machine learning analysis.

Melanoma, a highly malignant tumour, presents significant challenges due to its cellular heterogeneity, yet research on this aspect in cutaneous melanoma remains limited. In this study, we utilized single-cell data from 92,521 cells to explore the tumour cell landscape. Through clustering analysis, we identified six distinct cell clusters and investigated their differentiation and metabolic heterogeneity using multi-omics approaches. Notably, cytotrace analysis and pseudotime trajectories revealed distinct stages of tumour cell differentiation, which have implications for patient survival. By leveraging markers from these clusters, we developed a tumour cell-specific machine learning model (TCM). This model not only predicts patient outcomes and responses to immunotherapy, but also distinguishes between genomically stable and unstable tumours and identifies inflamed ('hot') versus non-inflamed ('cold') tumours. Intriguingly, the TCM score showed a strong association with TOMM40, which we experimentally validated as an oncogene promoting tumour proliferation, invasion and migration. Overall, our findings introduce a novel biomarker score that aids in selecting melanoma patients for improved prognoses and targeted immunotherapy, thereby guiding clinical treatment decisions.

Nuclear NME1 enhances the malignant behavior of A549 cells and impacts lung adenocarcinoma patient prognosis.

NME1 is a metastatic suppressor inconsistently reported to have multiple roles as both a promoter and inhibitor of cancer metastasis. Nevertheless, the specific mechanism behind these results is still unclear. We observed that A549 cells with stable transfer of NME1 into the nucleus (A549-nNm23-H1) exhibited significantly increased migration and invasion activity compared to vector control cells, which was further enhanced by over-expressing CYP24A1 (p < 0.001). NME1 demonstrated the ability to safely attach to and amplify the transcription activation of JUN, consequently leading to the up-regulation of CYP24A1. Analysis of clinical data showed a positive relationship between nuclear NME1 levels and CYP24A1 expression. Furthermore, they were positively associated with postoperative distant metastasis and negatively correlated with prognosis in those with early stage lung adenocarcinoma. In conclusion, the data presented provides a new understanding of the probable pathways by which nuclear NME1 facilitates tumor metastasis, establishing the groundwork for future prediction and treatment of tumor metastasis.

Successful Immune Reconstitution in a Patient with a TYK2 Deficiency after Allogeneic Stem Cell Transplantation from Unrelated Donors.

A boy with primary immunodeficiency, caused by a tyrosine kinase 2 (TYK2) mutation, presented with immune defects and a lifelong history of severe infections. Our aim was to determine whether allogeneic hematopoietic stem cell transplantation (HSCT) could restore the patient's immune defenses and reduce susceptibility to infection. In the absence of a suitable HLA-matched blood relative to act as a donor, the patient received an allogeneic HSCT from unrelated donors. The patient's clinical data were analyzed in the Children's Hospital of Chongqing Medical University (Chongqing, China) before transplantation and during the 4-year follow-up period using a combination of western blotting (e.g., TYK2 and STAT levels), qRT-PCR (e.g., T cell receptor rearrangement excision circles, kappa deletion element recombination circles, and TYK2 transcript levels), and flow cytometry (e.g., lymphocyte subpopulations and CD107α secretion). We found that HSCT significantly reduced the incidence of severe infections, restored normal TKY2 levels, and reversed defects such as impaired JAK/STAT signaling in response to interferon-α or interleukin-10 treatment. Although the patient did not develop acute graft-versus-host disease (GVHD) after transplantation, he did experience chronic GVHD symptoms in a number of organs, which were effectively managed. Our findings suggest that HSCT is a feasible strategy for reconstituting the immune system in TYK2-deficient patients; however, the factors associated with GVHD and autoimmune thyroiditis development in TYK2-deficient patients undergoing HSCT warrant further investigation.

Current understanding of ELF4 deficiency: a novel inborn error of immunity.

BACKGROUND: ELF4 deficiency has been recently recognized as a novel disorder within the spectrum of inborn errors of immunity (IEIs), specifically categorized as a "disease of immune dysregulation." Cases of this condition, reported by our team and others, are very limited worldwide. As such, our current knowledge of this new disease remains preliminary. This review aims to provide a brief overview of the clinical manifestations, pathogenesis, and treatment strategies for this novel IEI. DATA SOURCES: A comprehensive review was conducted after an extensive literature search in the PubMed/Medline database and websites concerning transcriptional factor ELF4 and reports concerning patients with ELF4 deficiency. Our search strategy was "ELF4 OR ETS-related transcription factor Elf-4 OR EL4-like factor 4 OR myeloid Elf-1-like factor" as of the time of manuscript submission. RESULTS: The current signature manifestations of ELF4 deficiency disorder are recurrent and prolonged oral ulcer, abdominal pain, and diarrhea in pediatric males. In some cases, immunodeficiency and autoimmunity can also be prominent. Targeted Sanger sequencing or whole exome sequencing can be used to detect variation in ELF4 gene. Western blotting for ELF4 expression of the patient's cells can confirm the pathogenic effect of the variant. To fully confirm the pathogenicity of the variant, further functional test is strongly advised. Glucocorticoid and biologics are the mainstream management of ELF4 deficiency disorder. CONCLUSIONS: Pediatric males presenting with recurring ulcerations in digestive tract epithelium with or without recurrent fever should be suspected of DEX. When atypical presentations are prominent, variations in ELF4 gene should be carefully evaluated functionally due to the complex nature of ELF4 function. Experience of treating DEX includes use of glucocorticoid and biologics and more precise treatment needs more patients to identify and further mechanistic study.

Comparison of the therapeutic effects of febuxostat combined with a low-purine diet and allopurinol combined with a low-purine diet on the improvement of gout patients.

OBJECTIVE: To compare the clinical efficacy of febuxostat combined with a low-purine diet versus allopurinol combined with a low-purine diet in the treatment of gout. METHODS: In this prospective controlled trial, 98 gout patients admitted to our hospital from February 2021 to December 2022 were enrolled as study subjects. Patients were randomly assigned to the study group (febuxostat combined with a low-purine diet) and the control group (allopurinol combined with a low-purine diet), with 49 patients in each group. The therapeutic effect was evaluated based on joint function and serum uric acid levels after treatment, and classified into three levels: markedly effective, effective, and ineffective. The levels of inflammatory factors, including tumor necrosis factor-a (TNF-a), cytokine interleukin-1beta (IL-1ß), and interleukin (IL)-18 (IL-18), were collected. The Numeric Rating Scale (NRS) was used to assess the degree of pain in patients. Clinical indicators before and 6 months after treatment were compared between the two groups. RESULTS: There was no statistically significant difference in age and gender between the two groups. After 6 months of treatment, the effective rate in the study group (48 cases, 97.96%) was higher than that in the control group (42 cases, 85.71%), with a statistically significant difference (p = .027). At the same time, the study group had significantly lower levels of serum uric acid (162.39 µmol/L ± 17.23 µmol/L vs. S198.32 µmol/L ± 18.34 µmol/L, p < .001), creatinine (87.39 mmol/L ± 9.76 mmol/L vs. 92.18 mmol/L ± 9.27 mmol/L, p = .014), total cholesterol (3.65 mmol/L ± 0.65 mmol/L vs. 4.76 mmol/L ± 0.73 mmol/L, p < .001), and triglycerides (1.76 mmol/L ± 0.32 mmol/L vs. 2.28 mmol/L ± 0.41 mmol/L, p < .001) compared to the control group, with statistically significant differences (p < .05). After treatment, the levels of inflammatory factors and degree of pain in the study group were significantly lower than those in the control group (all p < .05). During the treatment process, the incidence of adverse reactions in the study group (2 cases, 4.08%) was lower than that in the control group (9 cases, 18.37%), with a statistically significant difference (p = .025). CONCLUSION: Febuxostat combined with a low-purine diet can reduce inflammatory factors and alleviate the degree of pain in gout patients, significantly improving their clinical symptoms.

Fluoxetine Successfully Treats Intracranial Enterovirus E18 Infection in a Patient with CD79a Deficiency Arising from Segmental Uniparental Disomy of Chromosome 19.

The pre BCR complex plays a crucial role in B cell production, and its successful expression marks the B cell differentiation from the pro-B to pre-B. The CD79a and CD79b mutations, encoding Igα and Igß respectively, have been identified as the cause of autosomal recessive agammaglobulinemia (ARA). Here, we present a case of a patient with a homozygous CD79a mutation, exhibiting recurrent respiratory infections, diarrhea, growth and development delay, unique facial abnormalities and microcephaly, as well as neurological symptoms including tethered spinal cord, sacral canal cyst, and chronic enteroviral E18 meningitis. Complete blockade of the early B cell development in the bone marrow of the patient results in the absence of peripheral circulating mature B cells. Whole exome sequencing revealed a Loss of Heterozygosity (LOH) of approximately 19.20Mb containing CD79a on chromosome 19 in the patient. This is the first case of a homozygous CD79a mutation caused by segmental uniparental diploid (UPD). Another key outcome of this study is the effective management of long-term chronic enteroviral meningitis using a combination of intravenous immunoglobulin (IVIG) and fluoxetine. This approach offers compelling evidence of fluoxetine's utility in treating enteroviral meningitis, particularly in immunocompromised patients.

Atypical familial hemophagocytic lymphohistiocytosis type 3 in children: A report of cases and literature review.

BACKGROUND: Familial hemophagocytic lymphohistiocytosis type 3 (FHL3) is caused by UNC13D variants. The clinical manifestations of FHL3 are highly diverse and complex. Some patients exhibit atypical or incomplete phenotypes, making accurate diagnosis difficult. Our study aimed to broaden the understanding of the atypical FHL3 clinical spectrum. METHODS: In our study, we analyzed in detail the clinical features of four Chinese patients with UNC13D variants. Additionally, we conducted a comprehensive review of the existing literature on previously reported atypical manifestations and summarized the findings. RESULTS: Two of our patients presented with muscle involvement, while the other two had hematological involvement; none of them met the diagnostic criteria for hemophagocytic lymphohistiocytosis (HLH). However, protein expression and functional analysis ultimately confirmed diagnostic criteria for FHL3 in all patients. From the literature we reviewed, many atypical FHL3 patients had neurological involvement, especially isolated neurological manifestations. At the same time, arthritis and hypogammaglobulinemia were also prone to occur. CONCLUSION: Our study highlights that the expression of the Munc13-4 protein may not fully indicate the pathogenicity of UNC13D variants, whereas CD107a analysis could be more sensitive for disease diagnosis. These findings contribute to a broader understanding of the FHL3 clinical spectrum and may offer new insights into the underlying pathogenesis of UNC13D variants. It is crucial to prioritize the timely and accurate diagnosis of atypical patients, as they may often be overlooked among individuals with rheumatic or hematological diseases.

Identification and Functional Analysis of a de novo IKZF3 Mutation in a Pediatric Patient with Combined Immunodeficiency.

AIOLOS, a vital member of the IKAROS protein family, plays a significant role in lymphocyte development and function through DNA binding and protein-protein interactions. Mutations in the IKZF3 gene, which encodes AIOLOS, lead to a rare combined immunodeficiency often linked with infections and malignancy. In this study, we evaluated a 1-year-4-month-old female patient presenting with recurrent infections, diarrhea, and failure to thrive. Laboratory investigations revealed decreased T lymphocyte and immunoglobulin levels. Through whole-exome and Sanger sequencing, we discovered a de novo mutation in IKZF3 (NM_012481; exon 5 c.571G > C, p.Gly191Arg), corresponding to the third DNA-binding zinc finger region of the encoded protein AIOLOS. Notably, the patient with the AIOLOS G191R mutation showed reduced recent thymic emigrants in naïve CD4+T cells compared to healthy counterparts of the same age, while maintaining normal levels of Th1, Th2, Th17, Treg, and Tfh cells. This mutation also resulted in decreased switched memory B cells and lower CD23 and IgM expression. In vitro studies revealed that AIOLOS G191R does not impact the expression of AIOLOS but compromises its stability, DNA binding and pericentromeric targeting. Furthermore, AIOLOS G191R demonstrated a dominant-negative effect over the wild-type protein. This case represents the first reported instance of a mutation in the third DNA-binding zinc finger region of AIOLOS highlighting its pivotal role in immune cell functionality.

Image quality characterization of an ultra-high-speed kilovoltage cone-beam computed tomography imaging system on an O-ring linear accelerator.

PURPOSE: The quality of on-board imaging systems, including cone-beam computed tomography (CBCT), plays a vital role in image-guided radiation therapy (IGRT) and adaptive radiotherapy. Recently, there has been an upgrade of the CBCT systems fused in the O-ring linear accelerators called HyperSight, featuring a high imaging performance. As the characterization of a new imaging system is essential, we evaluated the image quality of the HyperSight system by comparing it with Halcyon 3.0 CBCT and providing benchmark data for routine imaging quality assurance. METHODS: The HyperSight features ultra-fast scan time, a larger kilovoltage (kV) detector, a more substantial kV tube, and an advanced reconstruction algorithm. Imaging protocols in the two modes of operation, treatment mode with IGRT and the CBCT for planning (CBCTp) mode were evaluated and compared with Halcyon 3.0 CBCT. Image quality metrics, including spatial resolution, contrast resolution, uniformity, noise, computed tomography (CT) number linearity, and calibration error, were assessed using a Catphan and an electron density phantom and analyzed with TotalQA software. RESULTS: HyperSight demonstrated substantial improvements in contrast-to-noise ratio and noise in both IGRT and CBCTp modes compared to Halcyon 3.0 CBCT. CT number calibration error of HyperSight CBCTp mode (1.06%) closely matches that of a full CT scanner (0.72%), making it suitable for adaptive planning. In addition, the advanced hardware of HyperSight, such as ultra-fast scan time (5.9 s) or 2.5 times larger heat unit capacity, enhanced the clinical efficiency in our experience. CONCLUSIONS: HyperSight represented a significant advancement in CBCT imaging. With its image quality, CT number accuracy, and ultra-fast scans, HyperSight has a potential to transform patient care and treatment outcomes. The enhanced scan speed and image quality of HyperSight are expected to significantly improve the quality and efficiency of treatment, particularly benefiting patients.

Technical note: Preprocessing of portal images to improve image quality of VMAT-CT.

BACKGROUND: The concept of volumetric modulated arc therapy-computed tomography (VMAT-CT) was proposed more than a decade ago. However, its application has been very limited mainly due to the poor image quality. More specifically, the blurred areas in electronic portal imaging device (EPID) images collected during VMAT heavily degrade the image quality of VMAT-CT. PURPOSE: The goal of this study was to propose systematic methods to preprocess EPID images and improve the image quality of VMAT-CT. METHODS: Online region-based active contour method was introduced to binarize portal images. Multi-leaf collimator (MLC) motion modeling was developed to remove the MLC motion blur. Outlier filtering was then applied to replace the remaining artifacts with plausible data. To assess the impact of these preprocessing methods on the image quality of VMAT-CT, 44 clinical VMAT plans for several treatment sites (lung, esophagus, and head & neck) were delivered to a Rando phantom, and several real-patient cases were also acquired. VMAT-CT reconstruction was attempted for all the cases, and image quality was evaluated. RESULTS: All three preprocessing methods could effectively remove the blurred edges of EPID images. The combined preprocessing methods not only saved VMAT-CT from distortions and artifacts, but also increased the percentage of VMAT plans that can be reconstructed. CONCLUSIONS: The systematic preprocessing of portal images improves the image quality of VMAT-CT significantly, and facilitates the application of VMAT-CT as an effective image guidance tool.

A novel model incorporating chromatin regulatory factors for risk stratification, prognosis prediction, and characterization of the microenvironment in Wilms tumor.

BACKGROUND: Wilms tumor, also known as nephroblastoma, a pediatric most-frequent malignant-kidney tumor, may be regulated and influenced by transcriptional and epigenetic mechanisms. Chromatin regulatory factors (CRs) play key roles in epigenetic regulation. The present study aimed to explore the involvement of CRs in the development of nephroblastoma. METHODS: RNA-sequencing and clinical information of nephroblastoma samples were obtained by downloading data from the TARGET database. The Limma package was utilized to perform differential expression analysis of genes (DEGs) between the tumor group and the control group. A Venn map was used for intersection of differential genes and CRs and to perform Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis of DEGs using the clusterProfiler package. LASSO and Cox analyses were used to construct CR-related risk models and were evaluated based on clinical parameters. A receiver operating characteristic curve was employed to assess the diagnostic performance of risk model. Furthermore, we used a single-sample gene set enrichment analysis algorithm for immune cell infiltration analysis. Finally, to confirm the transcriptome expression of pivotal genes in human nephroblastoma cell lines, a quantitative real-time PCR was employed. RESULTS: Fifteen key CRs were obtained through analysis in nephroblastoma and then the risk model based on 13 important CRs was constructed using the transcriptome data of nephroblastoma. Using the risk model, pediatric nephroblastoma patients were stratified into high- and low-risk groups based on their individual risk scores. The risk score of CRs can predict adverse outcomes in pediatric nephroblastoma, and this gene cluster is closely related to various immunity characteristics of nephroblastoma. Moreover, the nephroblastoma cell line exhibited higher expression levels of prognostic genes (VRK1, ARNTL, RIT1, PRDM6, and TSPY1) compared to the HEK293 T cell line. CONCLUSIONS: The risk characteristics derived from CRs have tremendous significance in predicting prognosis and guiding clinical classification and intervention strategies for pediatric nephroblastoma.

Peroxisome proliferator-activated receptors: A key link between lipid metabolism and cancer progression.

Lipids represent the essential components of membranes, serve as fuels for high-energy processes, and play crucial roles in signaling and cellular function. One of the key hallmarks of cancer is the reprogramming of metabolic pathways, especially abnormal lipid metabolism. Alterations in lipid uptake, lipid desaturation, de novo lipogenesis, lipid droplets, and fatty acid oxidation in cancer cells all contribute to cell survival in a changing microenvironment by regulating feedforward oncogenic signals, key oncogenic functions, oxidative and other stresses, immune responses, or intercellular communication. Peroxisome proliferator-activated receptors (PPARs) are transcription factors activated by fatty acids and act as core lipid sensors involved in the regulation of lipid homeostasis and cell fate. In addition to regulating whole-body energy homeostasis in physiological states, PPARs play a key role in lipid metabolism in cancer, which is receiving increasing research attention, especially the fundamental molecular mechanisms and cancer therapies targeting PPARs. In this review, we discuss how cancer cells alter metabolic patterns and regulate lipid metabolism to promote their own survival and progression through PPARs. Finally, we discuss potential therapeutic strategies for targeting PPARs in cancer based on recent studies from the last five years.

Generation of a human iPSC line (CHCMUi002-A) from peripheral blood mononuclear cells from a chronic granulomatous disease patient.

Chronic granulomatous disease (CGD) is a rare X-linked recessive primary immunodeficiency disease (PID). Herein, a human induced pluripotent stem cell (iPSC) line was generated from the peripheral blood mononuclear cells (PBMCs) of a CGD patient with a mutation (c.785_786delTT) in the CYBB gene. These iPSCs showed the expression of pluripotency markers, the ability to differentiate into three germ layers. They offer a promising technique for studying the pathogenesis and conducting drug screening for CGD patients.

Establishment and validation of a nomogram to select patients with metastatic sarcomatoid renal cell carcinoma suitable for cytoreductive radical nephrectomy.

Introduction: Metastatic renal cell carcinoma (mRCC) with sarcomatoid features has a poor prognosis. Cytoreductive radical nephrectomy (CRN) can improve prognosis, but patient selection is unclear. This study aimed to develop a prediction model for selecting patients suitable for CRN. Materials and methods: Patients with a diagnosis of mRCC with sarcomatoid features in the Surveillance, Epidemiology, and End Results (SEER) database between 2010 and 2015 were retrospectively reviewed. CRN benefit was defined as a survival time longer than the median overall survival (OS) in patients who did not receive CRN. A prediction nomogram was established and validated using the SEER cohort (training and internal validation) and an external validation cohort. Results: Of 900 patients with sarcomatoid mRCC, 608 (67.6%) underwent CRN. OS was longer in the CRN group than in the non-CRN group (8 vs. 6 months, hazard ratio (HR) = 0.767, p = 0.0085). In the matched CRN group, 124 (57.7%) patients survived >6 months after the surgery and were considered to benefit from CRN. Age, T-stage, systematic therapy, metastatic site, and lymph nodes were identified as independent factors influencing OS after CRN, which were included in the prediction nomogram. The monogram performed well on the training set (area under the receiver operating characteristic (AUC) curve = 0.766, 95% confidence interval (CI): 0.687-0.845), internal validation set (AUC = 0.796, 95% CI: 0.684-0.908), and external validation set (AUC = 0.911, 95% CI: 0.831-0.991). Conclusions: A nomogram was constructed and validated with good accuracy for selecting patients with sarcomatoid mRCC suitable for CRN.

A single-cell atlas of immunocytes in the spleen of a mouse model of Wiskott-Aldrich syndrome.

Wiskott-Aldrich syndrome (WAS) is a disorder characterized by rare X-linked genetic immune deficiency with mutations in the Was gene, which is specifically expressed in hematopoietic cells. The spleen plays a major role in hematopoiesis and red blood cell clearance. However, to date, comprehensive analyses of the spleen in wild-type (WT) and WASp-deficient (WAS-KO) mice, especially at the transcriptome level, have not been reported. In this study, single-cell RNA sequencing (scRNA-seq) was adopted to identify various types of immune cells and investigate the mechanisms underlying immune deficiency. We identified 30 clusters and 10 major cell subtypes among 11,269 cells; these cell types included B cells, T cells, dendritic cells (DCs), natural killer (NK) cells, monocytes, macrophages, granulocytes, stem cells and erythrocytes. Moreover, we evaluated gene expression differences among cell subtypes, identified differentially expressed genes (DEGs), and performed enrichment analyses to identify the reasons for the dysfunction in these different cell populations in WAS. Furthermore, some key genes were identified based on a comparison of the DEGs in each cell type involved in specific and nonspecific immune responses, and further analysis showed that these key genes were previously undiscovered pathology-related genes in WAS-KO mice. In summary, we present a landscape of immune cells in the spleen of WAS-KO mice based on detailed data obtained at single-cell resolution. These unprecedented data revealed the transcriptional characteristics of specific and nonspecific immune cells, and the key genes were identified, laying a foundation for future studies of WAS, especially studies into novel and underexplored mechanisms that may improve gene therapies for WAS.

A circRNA-based ceRNA network shows its diagnostic value in non-small-cell lung cancer.

BACKGROUND: Numerous studies have reported the vital roles of circular RNA (circRNA)-based competitive endogenous RNA (ceRNA) regulatory networks in cancers. Here, we established a non-small-cell lung cancer (NSCLC)-related circRNA-miRNA-mRNA axis and estimated its diagnostic value in NSCLC. METHODS: The circ_0061235-miR-3180-5p-PPM1L axis was constructed by small RNA deep sequencing, bioinformatics databases, and preliminary testing. The serum levels of the selected circ_0061235, miR-3180-5p, and PPM1L were quantified using quantitative polymerase chain reaction. Receiver operating characteristic analyses were conducted to evaluate the diagnostic power. RESULTS: The levels of circ_0061235, miR-3180-5p, and PPM1L showed close correlations according to the ceRNA regulation rule. They were significantly dysregulated in NSCLC and showed the diagnostic ability to discriminate between healthy and NSCLC, and remarkably, between benign lung tumors and NSCLC. Additionally, the down-regulated levels of hsa_circ_0061235, the up-regulated levels of miR-3180-5p, and the decreased levels of PPM1L were correlated to more aggressive features of NSCLC, such as lymph node metastasis, distant metastasis, and higher stages. Intriguingly, compared to the single circ_0061235, miR-3180-5p, PPM1L, and traditional tumor markers, the diverse combinations of circ_0061235, miR-3180-5p, and PPM1L showed much higher sensitivity and specificity to differentiate greater or lesser severity of NSCLC. GO annotation and KEGG pathway analyses revealed the underlying role of the circ_0061235-miR-3180-5p-PPM1L axis in NSCLC. CONCLUSIONS: We established a specific circRNA-miRNA-mRNA network with higher sensitivity and specificity to diagnose NSCLC, particularly more aggressive NSCLC, providing a new strategy for further developing tumor biomarkers.