HSPA4 upregulation induces immune evasion via ALKBH5/CD58 axis in gastric cancer.

INTRODUCTION: Gastric cancer (GC) is one of the leading causes of cancer-related death worldwide. Recently, targeted therapies including PD1 (programmed cell death 1) antibodies have been used in advanced GC patients. However, identifying new biomarker for immunotherapy is still urgently needed. The objective of this study is to unveil the immune evasion mechanism of GC cells and identify new biomarkers for immune checkpoint blockade therapy in patients with GC. METHODS: Coimmunoprecipitation and meRIP were performed to investigate the mechanism of immune evasion of GC cells. Cocuture system was established to evaluate the cytotoxicity of cocultured CD8+ T cells. The clinical significance of HSPA4 upregulation was analyzed by multiplex fluorescent immunohistochemistry staining in GC tumor tissues. RESULTS: Histone acetylation causes HSPA4 upregulation in GC tumor tissues. HSPA4 upregulation increases the protein stability of m6A demethylase ALKBH5. ALKBH5 decreases CD58 in GC cells through m6A methylation regulation. The cytotoxicity of CD8+ T cells are impaired and PD1/PDL1 axis is activated when CD8+ T cells are cocultured with HSPA4 overexpressed GC cells. HSPA4 upregulation is associated with worse 5-year overall survival of GC patients receiving only surgery. It is an independent prognosis factor for worse survival of GC patients. In GC patients receiving the combined chemotherapy with anti-PD1 immunotherapy, HSPA4 upregulation is observed in responders compared with non-responders. CONCLUSION: HSPA4 upregulation causes the decrease of CD58 in GC cells via HSPA4/ALKBH5/CD58 axis, followed by PD1/PDL1 activation and impairment of CD8+ T cell's cytotoxicity, finally induces immune evasion of GC cells. HSPA4 upregulation is associated with worse overall survival of GC patients with only surgery. Meanwhile, HSPA4 upregulation predicts for better response in GC patients receiving the combined immunotherapy.

Lymphatic endothelial cell-mediated accumulation of CD177+Treg cells suppresses antitumor immunity in human esophageal squamous cell carcinoma.

Regulatory T (Treg) cells are critical in shaping an immunosuppressive microenvironment to favor tumor progression and resistance to therapies. However, the heterogeneity and function of Treg cells in esophageal squamous cell carcinoma (ESCC) remain underexplored. We identified CD177 as a tumor-infiltrating Treg cell marker in ESCC. Interestingly, expression levels of CD177 and PD-1 were mutually exclusive in tumor Treg cells. CD177+ Treg cells expressed high levels of IL35, in association with CD8+ T cell exhaustion, whereas PD-1+ Treg cells expressed high levels of IL10. Pan-cancer analysis revealed that CD177+ Treg cells display increased clonal expansion compared to PD-1+ and double-negative (DN) Treg cells, and CD177+ and PD-1+ Treg cells develop from the same DN Treg cell origin. Importantly, we found CD177+ Treg cell infiltration to be associated with poor overall survival and poor response to anti-PD-1 immunotherapy plus chemotherapy in ESCC patients. Finally, we found that lymphatic endothelial cells are associated with CD177+ Treg cell accumulation in ESCC tumors, which are also decreased after anti-PD-1 immunotherapy plus chemotherapy. Our work identifies CD177+ Treg cell as a tumor-specific Treg cell subset and highlights their potential value as a prognostic marker of survival and response to immunotherapy and a therapeutic target in ESCC.

Macrophage-expressed SRA ameliorates alcohol-induced liver injury by suppressing S-glutathionylation of Notch1 via recruiting thioredoxin.

Scavenger receptor A (SRA) is preferentially expressed in macrophages and implicated as a multifunctional pattern recognition receptor for innate immunity. Hepatic macrophages play a primary role in the pathogenesis of alcoholic liver disease. Herein, we observed that SRA expression was significantly increased in the liver tissues of mice with alcohol-related liver injury. SRA-deficient (SRA-/-) mice developed more severe alcohol-induced liver disease than wild-type mice. Enhanced liver inflammation existed in alcohol-challenged SRA-/- mice and was associated with increased Notch activation in hepatic macrophages compared with wild-type control animals. Mechanistically, SRA directly bound with Notch1 and suppressed its S-glutathionylation, thereby inhibiting Notch pathway activation. Further, we determined that the SRA interacted with thioredoxin-1 (Trx-1), a redox-active protein. SRA inhibited Trx-1 dimerization and facilitated the interaction of Trx-1 with Notch1. Application of a Trx-1-specific inhibitory agent during macrophage stimulation abolished SRA-mediated regulation of the Notch pathway and its downstream targets. In summary, our study revealed that SRA plays a critical role in macrophage inflammatory response by targeting Notch1 for its glutathionylation. SRA-mediated negative regulation of Notch activation might serve as a novel therapeutic strategy for alcohol-induced liver injury.

CSE triggers ferroptosis via SIRT4-mediated GNPAT deacetylation in the pathogenesis of COPD.

BACKGROUND: It is now understood that ferroptosis plays a significant role in the progression of chronic obstructive pulmonary disease (COPD) induced by cigarette smoke extract (CSE). However, the mechanisms underlying this relationship remain largely unclear. METHODS: In this study, we established a COPD mouse model through exposure to cigarette smoke particulates, followed by H&E staining, analysis of bronchoalveolar lavage fluid, and immunohistochemistry assay. A549 cells were exposed to increasing concentrations of CSE, with the addition of the ferroptosis activator erastin or the inhibitor Fer-1. Cell viability, LDH (lactate dehydrogenase) release, inflammatory cytokines, total ROS (reactive oxygen species), and lipid ROS were measured using the corresponding assay kits. The acetylation level of GNPAT was determined through immunoprecipitation. We assessed the expression levels of molecules involved in plasmalogen biosynthesis (FAR1, AGPS, and GNPAT), GPX4, and SIRT4 using quantitative real-time PCR, western blot analysis, and immunofluorescence staining. RESULTS: CSE-induced lung tissue damage was initially observed, accompanied by oxidative stress, ferroptosis, and increased plasmalogen biosynthesis molecules (FAR1, AGPS, and GNPAT). CSE also induced ferroptosis in A549 cells, resulting in reduced cell viability, GSH, and GPX4 levels, along with increased LDH, ROS, MDA (malondialdehyde) levels, oxidized lipids, and elevated FAR1, AGPS, and GNPAT expression. Knockdown of GNPAT mitigated CSE-induced ferroptosis. Furthermore, we found that CSE regulated the acetylation and protein levels of GNPAT by modulating SIRT4 expression. Importantly, the overexpression of GNPAT countered the inhibitory effects of SIRT4 on ferroptosis. CONCLUSIONS: Our study revealed GNPAT could be deacetylated by SIRT4, providing novel insights into the mechanisms underlying the relationship between CSE-induced ferroptosis and COPD.

Synthesis, Surface Activity, and Foamability of Two Short-Chain Fluorinated Sulfonate Surfactants with Ether Bonds.

Fluorinated surfactants are widely used in many fields because of their excellent surface active properties, but their high stability has caused many environmental problems. With the ban and restriction of classical long-chain fluorinated surfactants such as perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) worldwide, the development and replacement of their alternatives is now a major challenge. How to reduce environmental persistence, bioaccumulation, and biotoxicity while maintaining high surface activity has become an important issue in the development of fluorinated surfactants. Using short-chain fluorinated surfactants is one of the important solutions to resolve the pollution of organic fluorinated compounds. In this article, we synthesized two short-chain fluorinated surfactants with ether bonds. One of them 6:2 FTESNa (2) used the perfluoroalkyl chain (n-C6F13-) and the other C72 FEESNa (4) used the fluoroether segment with six fluorinated carbons and two oxygens (CF3OCF(CF3)CF2OCF(CF3)). The surface activity, foam performance, and wettability of the two molecules were measured. The surface tensions at critical micelle concentration (γcmc) and the critical micelle concentration (cmc) of 2 and 4 were 17.6 mN/m (2.2 g/L) and 20.2 mN/m (4.6 g/L), respectively. Both of them were significantly superior to the surface activity of 6:2 FTSNa (7) which is one of the current alternatives for PFOS. Additionally, the foamability and foam stability of both 2 and 4 were better than that of 7. In the aspect of wettability on PTFE, that of 4 was greater than those of 2 and 7. In summary, this work provided a new choice for alternatives of PFOS and PFOA.

Dissecting T-cell heterogeneity in esophageal squamous cell carcinoma reveals the potential role of LAIR2 in antitumor immunity.

Esophageal squamous cell carcinoma (ESCC), one of the most commonly diagnosed and lethal malignant diseases, has a complex tumor ecosystem. An obvious requirement for T-cell-mediated tumor control is the infiltration of tumor-reactive T cells into the tumor. Here, we obtained detailed T-cell compositions in both ESCC tumors and matched peripheral blood mononuclear cells (PBMCs) at single-cell resolution. We demonstrated that T cells in tumors and PBMCs had different compositions and functional states. ESCC tumors were rich in Treg and exhausted T cells but poor in cytotoxic and naïve T cells compared with PBMCs. The exhausted T cells showed higher exhausted signature in tumors than in PBMCs, while the cytotoxic T cells exhibited higher cytotoxic signature in PBMCs than in tumors. Our data indicated an immunosuppressive status and a defect at the level of T-cell priming in the tumor microenvironment. Leukocyte-associated Ig-like receptor-2 (LAIR2), a soluble collagen receptor that prevents the binding of human leukocyte-associated Ig-like receptor-1 (LAIR1) to collagens, was predominantly expressed in proliferating CD8+ T and Treg cells in tumors but in cytotoxic cells in PBMCs. LAIR2 could inhibit tumor metastasis, invasion, and collagen deposition via suppressing transforming growth factor-ß signaling. These findings revealed differential T-cell populations in tumors and PBMCs and provided convincing evidence that LAIR2 acted as a tumor suppressor.

Liver abscess complicated with multiple organ invasive infection caused by hematogenous disseminated hypervirulent Klebsiella pneumoniae: A case report.

Hypervirulent Klebsiella pneumoniae (hvKp) causes increasing infections in healthy individuals from the community. In severe cases, it can cause multiple organ infection with invasive metastasis of blood sources, seriously threatening the patients' life. Rapid and accurate diagnosis of the pathogen becomes the key to timely antibiotic treatment to improve the prognosis. This article reports a case of liver abscess complicated with multiple organ invasive infection caused by hematogenous-disseminated hvKp. K. pneumoniae was identified by culture and metagenomic next-generation sequencing (mNGS) using blood and liver abscess drainage fluid. The isolates from the two samples were subsequently identified with high homology (99.999%) by whole genome sequencing. In addition, multiple virulence genes were detected in the two isolates and the string test was positive, indicating hvKp with hypermucoviscosity phenotype. Multiple antibiotic treatments were given. The conditions of the patient were stable but the temperature remained high. Surgical drainage treatment was performed, and the patient's body temperature immediately dropped to normal. He finally recovered after 6 months of follow-up. mNGS using body fluids can facilitate the rapid diagnosis of pathogens. For hvKp infection, choosing a better antibiotic therapy and receiving surgical drainage can significantly improve the prognosis of the patient.

Human Adipose-Derived Mesenchymal Stem Cell-Based Microspheres Ameliorate Atherosclerosis Progression In Vitro.

Atherosclerosis (AS) is a chronic inflammatory disease associated with lipid deposition, which could be converted into acute clinical events by thrombosis or plaque rupture. Adipose-derived mesenchymal stem cell (ADSC)-encapsulated repair units could be an effective cure for the treatment of AS patients. In this study, we encapsulate human adipose-derived mesenchymal stem cells (hADSCs) in collagen microspheres to fabricate stem cell repair units. Besides, we show that encapsulation in collagen microspheres and cultured in vitro for 14 days maintain the viability and stemness of hADSCs. Moreover, we generate AS progression model and niche in vitro by combining hyperlipemia serum of AS patients with AS cell models. We further systematically demonstrate that hADSC-based microspheres could ameliorate AS progression by inhibiting oxidative stress injury, cell apoptosis, endothelial dysfunction, inflammation, and lipid accumulation. In addition, we perform transcriptomic analysis and functional studies to demonstrate how hADSCs (three dimensional cultured in microspheres) respond to AS niche compared with healthy microenvironment. These findings reveal a role for ADSC-based microspheres in the treatment of AS and provide new ideas for stem cell therapy in cardiovascular disease. The results may have implications for improving the efficiency of hADSC therapies by illuminating the mechanisms of hADSCs exposed in special pathological niche.

HERC2 promotes inflammation-driven cancer stemness and immune evasion in hepatocellular carcinoma by activating STAT3 pathway.

BACKGROUND: Hepatic inflammation is a common initiator of liver diseases and considered as the primary driver of hepatocellular carcinoma (HCC). However, the precise mechanism of inflammation-induced HCC development and immune evasion remains elusive and requires extensive investigation. This study sought to identify the new target that is involved in inflammation-related liver tumorigenesis. METHODS: RNA-sequencing (RNA-seq) analysis was performed to identify the differential gene expression signature in primary human hepatocytes treated with or without inflammatory stimulus. A giant E3 ubiquitin protein ligase, HECT domain and RCC1-like domain 2 (HERC2), was identified in the analysis. Prognostic performance in the TCGA validation dataset was illustrated by Kaplan-Meier plot. The functional role of HERC2 in HCC progression was determined by knocking out and over-expressing HERC2 in various HCC cells. The precise molecular mechanism and signaling pathway networks associated with HERC2 in HCC stemness and immune evasion were determined by quantitative real-time PCR, immunofluorescence, western blot, and transcriptomic profiling analyses. To investigate the role of HERC2 in the etiology of HCC in vivo, we applied the chemical carcinogen diethylnitrosamine (DEN) to hepatocyte-specific HERC2-knockout mice. Additionally, the orthotopic transplantation mouse model of HCC was established to determine the effect of HERC2 during HCC development. RESULTS: We found that increased HERC2 expression was correlated with poor prognosis in HCC patients. HERC2 enhanced the stemness and PD-L1-mediated immune evasion of HCC cells, which is associated with the activation of signal transducer and activator of transcription 3 (STAT3) pathway during the inflammation-cancer transition. Mechanically, HERC2 coupled with the endoplasmic reticulum (ER)-resident protein tyrosine phosphatase 1B (PTP1B) and limited PTP1B translocation from ER to ER-plasma membrane junction, which ameliorated the inhibitory role of PTP1B in Janus kinase 2 (JAK2) phosphorylation. Furthermore, HERC2 knockout in hepatocytes limited hepatic PD-L1 expression and ameliorated HCC progression in DEN-induced mouse liver carcinogenesis. In contrast, HERC2 overexpression promoted tumor development and progression in the orthotopic transplantation HCC model. CONCLUSION: Our data identified HERC2 functions as a previously unknown modulator of the JAK2/STAT3 pathway, thereby promoting inflammation-induced stemness and immune evasion in HCC.

Iron carbide nanoparticles supported on an N-doped carbon porous framework as a bifunctional material for electrocatalytic oxygen reduction and supercapacitors.

Highly active and durable bifunctional materials are of pivotal importance for energy conversion and storage devices, yet a comprehensive understanding of their geometric and electronic influence on electrochemical activity is urgently needed. Fe-N-C materials with physical and chemical structural merits are considered as one of the promising candidates for efficient oxygen reduction reaction electrocatalysts and supercapacitor electrodes. Herein, Fe3C nanoparticles supported on a porous N-doped carbon framework (denoted as Fe3C/PNCF) were readily prepared by one-step chemical vapor deposition under the assistance of a NaCl salt template. The experiment results revealed that the as-synthesized Fe3C/PNCF nanocomposites successfully displayed attractive electrocatalytic oxygen reduction reaction (ORR) activity comparable to that of the Pt/C catalyst (E1/2 of 0.84 V and 0.83 V, respectively), and a superior capacitance of 385.3 F g-1 under 1 A g-1 for a supercapacitor. It's proposed that the increased pyridinic and graphitic N coordination on the hydrophilic porous framework provides more electrochemical active surface area for the storage and transport of electrolyte ions. Additionally, an appropriate d-band center created by the optimized adsorption function endows Fe3C/PNCF with excellent electrochemical properties. The results confirmed that the integration strategy of porous heterogeneous structure and accessible active sites balanced the complex relationship between geometry, electronic structure, and electrochemical activity. Our research provides a facile approach for fabricating multi-functional nanomaterials applicable in both ORR and supercapacitors in the future.

SRA Suppresses Antiviral Innate Immune Response in Macrophages by Limiting TBK1 K63 Ubiquitination via Deubiquitinase USP15.

The innate immune system is the first line of host defense against microbial infections. During virus infection, pattern recognition receptors (PRRs) are engaged to detect specific viral components, such as viral RNA or DNA, and regulate the innate immune response in the infected cells or immune cells. Our previous study demonstrated that scavenger receptor A (SRA), an important innate PRR, impaired the anti-hepatitis B virus (HBV) response in hepatocytes. Given that SRA is primarily expressed in macrophages, here, we assessed the function of SRA expressed in macrophages in response to RNA or DNA viral infection. SRA-deficient (SRA-/-) mice showed reduced susceptibility to viral infection caused by vesicular stomatitis virus (VSV) or herpes simplex virus 1 (HSV-1). In the virus-infected SRA-/- mice, compared with their wild-type (WT) counterparts, we observed low amounts of virus accompanied by enhanced interferon (IFN) production. Furthermore, SRA significantly inhibited the phosphorylation of TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3). We provided biochemical evidence showing that SRA directly interacts with the N-terminal kinase domain (KD) of TBK1, resulting in the limitation of its K63-linked ubiquitination. Moreover, we demonstrated that SRA negatively regulates the activity of TBK1 by promoting the recruitment of ubiquitin-specific protease 15 (USP15) to deubiquitinate TBK1. In summary, we have identified the connection between SRA and the TBK1/IRF3 signaling pathway in macrophages, indicating a critical role of SRA in the regulation of host antiviral immunity. IMPORTANCE During virus infection, PRRs are engaged to detect specific viral components, such as viral RNA or DNA, and regulate the innate immune response in the infected cells or other immune cells. We reported that deficiency of SRA, an important innate PRR, promoted IRF3 activation, type I IFN production, and innate antiviral responses against RNA and DNA viruses in vivo and in vitro. Furthermore, the biochemical analysis showed that SRA directly interacts with the KD domain of TBK1 and limits its K63-linked polyubiquitination, reducing TBK1 activation. Further analyses determined that SRA is a modulator for TBK1 activation via the recruitment of USP15, which delineated a previously unrecognized function for SRA in innate antiviral immunity.

Fucoidan inhibits EGFR redistribution and potentiates sorafenib to overcome sorafenib-resistant hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide. Although sorafenib is a standard first-line molecule-targeted drug against advanced HCC, the drug resistance development and adverse side effects usually limit its efficacy. This study investigated the effect of fucoidan on the sorafenib sensitivity of sorafenib-resistant human HCC cell line HepG2-SR established by long-time exposure of HepG2 to sorafenib. We demonstrated fucoidan combined with sorafenib synergistically promoted apoptosis and cell cycle arrest whereas inhibited cell migration in HepG2-SR cells. This combination treatment effectively suppressed the cellular epithelial growth factor receptor (EGFR) nuclear distribution and downstream gene transcription. Interestingly, fucoidan bound the cell surface EGFR, dampening EGFR translocation to lipid raft and further nuclear distribution, restoring the sorafenib sensitivity in HepG2-SR cells. Blocking fucoidan-EGFR interaction using EGFR antibody restrained the enhanced anti-tumor effects upon the combined administration. Besides, EGFR knockdown abolished the combination treatment-improved anti-tumor efficacy. This combination also suppressed in vivo xenograft tumor growth in nude mice. Our present study uncovered that fucoidan overcame sorafenib resistance in HCC via its interaction with cell membrane EGFR and further suppression of EGFR redistribution and downstream signaling in sorafenib-resistant cells. Overall, current results suggest that simultaneous treatment of fucoidan and sorafenib might serve as a potential therapeutic strategy against sorafenib-resistant HCC.

A Deep Learning Model for Accurate Diagnosis of Infection Using Antibody Repertoires.

The adaptive immune receptor repertoire consists of the entire set of an individual's BCRs and TCRs and is believed to contain a record of prior immune responses and the potential for future immunity. Analyses of TCR repertoires via deep learning (DL) methods have successfully diagnosed cancers and infectious diseases, including coronavirus disease 2019. However, few studies have used DL to analyze BCR repertoires. In this study, we collected IgG H chain Ab repertoires from 276 healthy control subjects and 326 patients with various infections. We then extracted a comprehensive feature set consisting of 10 subsets of repertoire-level features and 160 sequence-level features and tested whether these features can distinguish between infected individuals and healthy control subjects. Finally, we developed an ensemble DL model, namely, DL method for infection diagnosis (https://github.com/chenyuan0510/DeepID), and used this model to differentiate between the infected and healthy individuals. Four subsets of repertoire-level features and four sequence-level features were selected because of their excellent predictive performance. The DL method for infection diagnosis outperformed traditional machine learning methods in distinguishing between healthy and infected samples (area under the curve = 0.9883) and achieved a multiclassification accuracy of 0.9104. We also observed differences between the healthy and infected groups in V genes usage, clonal expansion, the complexity of reads within clone, the physical properties in the α region, and the local flexibility of the CDR3 amino acid sequence. Our results suggest that the Ab repertoire is a promising biomarker for the diagnosis of various infections.

A Novel Multi-Port Containment System for Laparoscopic Power Morcellation to Prevent Tumoral Spread: A Retrospective Cohort Study.

OBJECTIVE: To report a novel multi-port containment (NMC) system for laparoscopic power morcellation to prevent tumoral spread and to evaluate its safety, validity, and feasibility. METHODS: This retrospective study included women who underwent laparoscopic myomectomy (LM) between January 2014 and August 2020 at a single academic institution. The NMC system was used in the study group (n = 193); the control group underwent unprotected LM (n = 1753). RESULTS: After 1:1 propensity score matching, no significant differences in the baseline characteristics were observed between 193 matched pairs. Bag damages were detected in two cases in the study group before morcellation, and the NMC systems were replaced. There were no significant differences between the two groups in terms of the complications, total operative time, estimated blood loss, or postoperative hospitalization duration. In the study group, all operations were completed and no system rupture or leakage was observed. The median follow-up times were 21 and 54 months in the study and control groups, respectively. There was no peritoneal tissue spread in the study group. However, three (3/5, 0.6%) and six (6/1,753, 0.3%) patients in the control group experienced malignant and benign peritoneal tissue spread, respectively. CONCLUSION: The NMC system for laparoscopic power morcellation is valid, safe, and feasible for preventing a tumor spread.

The promoter hypermethylation of SULT2B1 accelerates esophagus tumorigenesis via downregulated PER1.

BACKGROUND: Esophageal cancer is currently the eighth most common tumor in the world and a leading cause of cancer death. SULT2B1 plays crucial roles in tumorigenesis. The purpose of this study is to explore the role of SULT2B1 in esophageal squamous cell carcinoma (ESCC). METHODS: The expression of SULT2B1 and its clinicopathological characteristics were evaluated in ESCC cohorts. Bisulfite genomic sequencing and methylation specific PCR were used to detect the promoter hypermethylation of the SULT2B1 gene. The effects of SULT2B1 on the biological characters of ESCC cells were identified on functional assays. Subcutaneous xenograft models revealed the role of SULT2B1 in vivo with tumor growth. RNA-Seq analysis and qRT-PCR were performed to recognize the targeted effect of SULT2B1 on PER1. RESULTS: SULT2B1 was not expressed or at a low level in most patients with ESCC or in ESCC cell lines, and this was accompanied by poor clinical prognosis. Furthermore, the downregulation of SULT2B1 occurred in promoter hypermethylation. According to the functional results, overexpression of SULT2B1 could inhibit tumoral proliferation in vitro and retard tumor growth in vivo, whereas SULT2B1 knockdown could accelerate ESCC progression. Mechanistically, SULT2B1 targeted PER1 at the mRNA level during post-transcriptional regulation. Finally, PER1 was verified as a suppressor and poor-prognosis factor in ESCC. CONCLUSIONS: SULT2B1 loss is a consequence owing to its ability to promote hypermethylation. In addition, it serves as a suppressor and poor-prognosis factor because of the post-transcriptional regulation of PER1 in ESCC.

Comprehensive single-cell sequencing reveals the stromal dynamics and tumor-specific characteristics in the microenvironment of nasopharyngeal carcinoma.

The tumor microenvironment (TME) of nasopharyngeal carcinoma (NPC) harbors a heterogeneous and dynamic stromal population. A comprehensive understanding of this tumor-specific ecosystem is necessary to enhance cancer diagnosis, therapeutics, and prognosis. However, recent advances based on bulk RNA sequencing remain insufficient to construct an in-depth landscape of infiltrating stromal cells in NPC. Here we apply single-cell RNA sequencing to 66,627 cells from 14 patients, integrated with clonotype identification on T and B cells. We identify and characterize five major stromal clusters and 36 distinct subpopulations based on genetic profiling. By comparing with the infiltrating cells in the non-malignant microenvironment, we report highly representative features in the TME, including phenotypic abundance, genetic alternations, immune dynamics, clonal expansion, developmental trajectory, and molecular interactions that profoundly influence patient prognosis and therapeutic outcome. The key findings are further independently validated in two single-cell RNA sequencing cohorts and two bulk RNA-sequencing cohorts. In the present study, we reveal the correlation between NPC-specific characteristics and progression-free survival. Together, these data facilitate the understanding of the stromal landscape and immune dynamics in NPC patients and provides deeper insights into the development of prognostic biomarkers and therapeutic targets in the TME.

Bronchoscopy for diagnosis of COVID-19 with respiratory failure: A case report.

BACKGROUND: Although the imaging features of coronavirus disease 2019 (COVID-19) are starting to be well determined, what actually occurs within the bronchi is poorly known. Here, we report the processes and findings of bronchoscopy in a patient with COVID-19 accompanied by respiratory failure. CASE SUMMARY: A 65-year-old male patient was admitted to the Hainan General Hospital on February 3, 2020 for fever and shortness of breath for 13 d that worsened for the last 2 d. The severe acute respiratory syndrome coronavirus 2 nucleic acid test was positive. Routine blood examination on February 28 showed a white blood cell count of 11.02 × 109/L, 86.9% of neutrophils, 6.4% of lymphocytes, absolute lymphocyte count of 0.71 × 109/L, procalcitonin of 2.260 ng/mL, and C-reactive protein of 142.61 mg/L. Oxygen saturation was 46% at baseline and turned to 94% after ventilation. The patient underwent video bronchoscopy. The tracheal cartilage ring was clear, and no deformity was found in the lumen. The trachea and bilateral bronchi were patent, while the mucosa was with slight hyperemia; no neoplasm or ulcer was found. Moderate amounts of white gelatinous secretions were found in the dorsal segment of the left inferior lobe, and the bronchial lumen was patent after sputum aspiration. The right inferior lobe was found with hyperemia and mucosal erosion, with white gelatinous secretion attachment. The patient's condition did not improve after the application of therapeutic bronchoscopy. CONCLUSION: For patients with COVID-19 and respiratory failure, bronchoscopy can be performed under mechanical ventilation to clarify the airway conditions. Protection should be worn during the process. Considering the risk of infection, it is not necessary to perform bronchoscopy in the mild to moderate COVID-19 patients.

DNase1L3 suppresses hepatocellular carcinoma growth via inhibiting complement autocrine effect.

Hepatocellular carcinoma (HCC) is one of the most aggressive types of cancer and currently lacks effective treatment strategies. The present study revealed that deoxyribonuclease 1 like 3 (DNase1L3) expression levels were significantly downregulated in numerous types of gastrointestinal cancer, and especially in HCC. Tissue microarrays were further used to illustrate that DNase1L3 expression levels were frequently downregulated in HCC tissues compared with normal liver tissues. In addition, DNase1L3 expression levels were identified to be significantly associated with tumor size (p=0.0028), tumor thrombus formation (p<0.01), and a poorer overall survival (p=0.005) and disease-free survival (p=0.006) of HCC. Gene Ontology functional term enrichment analysis of biological processes discovered that DNase1L3 was significantly associated with complement activation. Further studies demonstrated that the ectopic expression of DNase1L3 suppressed cell growth and inhibited the PI3K/AKT signaling pathway activation following C3a receptor agonist treatment. In conclusion, the findings of the present study suggested, for the first time, that DNase1L3 may serve as a biomarker for the prognosis of patients with HCC, and may suppress HCC growth via inhibiting the PI3K/AKT signaling pathway.

Laminin γ2-mediating T cell exclusion attenuates response to anti-PD-1 therapy.

PD-1/PD-L1 blockade therapies provide notable clinical benefits for patients with advanced cancers, but the factors influencing the effectiveness of the treatment remain incompletely cataloged. Here, the up-regulation of laminin γ2 (Ln-γ2) predicted the attenuated efficacy of anti-PD-1 drugs and was associated with unfavorable outcomes in patients with lung cancer or esophageal cancer. Furthermore, Ln-γ2 was transcriptionally activated by transforming growth factor-ß1 (TGF-ß1) secreted from cancer-associated fibroblasts via JNK/AP1 signaling, which blocked T cell infiltration into the tumor nests by altering the expression of T cell receptors. Coadministration of the TGF-ß receptor inhibitor galunisertib and chemotherapy drugs provoked vigorous antitumor activity of anti-PD-1 therapy in mouse tumor models. Therefore, Ln-γ2 may represent a useful biomarker to optimize clinical decisions and predict the response of cancer patients to treatment with anti-PD-1 drugs.

Tumor Fibroblast-Derived FGF2 Regulates Expression of SPRY1 in Esophageal Tumor-Infiltrating T Cells and Plays a Role in T-cell Exhaustion.

T-cell exhaustion was initially identified in chronic infection in mice and was subsequently described in humans with cancer. Although the distinct signature of exhausted T (TEX) cells in cancer has been well investigated, the molecular mechanism of T-cell exhaustion in cancer is not fully understood. Using single-cell RNA sequencing, we report here that TEX cells in esophageal cancer are more heterogeneous than previously clarified. Sprouty RTK signaling antagonist 1 (SPRY1) was notably enriched in two subsets of exhausted CD8+ T cells. When overexpressed, SPRY1 impaired T-cell activation by interacting with CBL, a negative regulator of ZAP-70 tyrosine phosphorylation. Data from the Tumor Immune Estimation Resource revealed a strong correlation between FGF2 and SPRY1 expression in esophageal cancer. High expression of FGF2 was evident in fibroblasts from esophageal cancer tissue and correlated with poor overall survival. In vitro administration of FGF2 significantly upregulated expression of SPRY1 in CD8+ T cells and attenuated T-cell receptor-triggered CD8+ T-cell activation. A mouse tumor model confirmed that overexpression of FGF2 in fibroblasts significantly upregulated SPRY1 expression in TEX cells, impaired T-cell cytotoxic activity, and promoted tumor growth. Thus, these findings identify FGF2 as an important regulator of SPRY1 expression involved in establishing the dysfunctional state of CD8+ T cells in esophageal cancer. SIGNIFICANCE: These findings reveal FGF2 as an important regulator of SPRY1 expression involved in establishing the dysfunctional state of CD8+ T cells and suggest that inhibition of FGF2 has potential clinical value in ESCC. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/24/5583/F1.large.jpg.