ABSTRACT
While the significance of N6-methyladenosine (m6A) in viral regulation has been extensively studied, the functions of 5-methylcytosine (m5C) modification in viral biology remain largely unexplored. In this study, we demonstrate that m5C is more abundant than m6A in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and provide a comprehensive profile of the m5C landscape of SARS-CoV-2 RNA. Knockout of NSUN2 reduces m5C levels in SARS-CoV-2 virion RNA and enhances viral replication. Nsun2 deficiency mice exhibited higher viral burden and more severe lung tissue damages. Combined RNA-Bis-seq and m5C-MeRIP-seq identified the NSUN2-dependent m5C-methylated cytosines across the positive-sense genomic RNA of SARS-CoV-2, and the mutations of these cytosines enhance RNA stability. The progeny SARS-CoV-2 virions from Nsun2 deficiency mice with low levels of m5C modification exhibited a stronger replication ability. Overall, our findings uncover the vital role played by NSUN2-mediated m5C modification during SARS-CoV-2 replication and propose a host antiviral strategy via epitranscriptomic addition of m5C methylation to SARS-CoV-2 RNA.
Subject(s)
COVID-19 , RNA, Viral , SARS-CoV-2 , Virus Replication , Virus Replication/genetics , Animals , SARS-CoV-2/genetics , SARS-CoV-2/pathogenicity , SARS-CoV-2/physiology , SARS-CoV-2/metabolism , RNA, Viral/genetics , RNA, Viral/metabolism , COVID-19/virology , COVID-19/pathology , Mice , Humans , Methylation , Virulence/genetics , 5-Methylcytosine/metabolism , 5-Methylcytosine/analogs & derivatives , Epigenesis, Genetic , Mice, Knockout , Adenosine/analogs & derivatives , Adenosine/metabolism , TranscriptomeABSTRACT
To solve the problems of poor surface quality and low tool life in conventional machining (CM) of AerMet100 steel, an experimental study was conducted in laser-assisted machining (LAM) of AerMet100 steel. The effects of laser power, cutting speed, feed rate, and depth of cut on the surface roughness of AerMet100 steel were studied based on a single-factor experiment. The degree of influence of each factor on the surface roughness was evaluated by analyses of variance and range in the orthogonal experiment, and the combination of process parameters for the optimal surface roughness was obtained. The order of influence was as follows: laser power > cutting speed > depth of cut > feed rate; the optimal combination of process parameters was laser power 200 W, cutting speed 56.5 m/min, feed rate 0.018 mm/rev, and depth of cut 0.3 mm. Compared to CM, the surface morphology of the workpiece under the optimization of LAM was relatively smooth and flat, the surface roughness Ra was 0.402 µm, which was reduced by 62.11%, the flank wear was reduced from 208.69 µm to 52.17 µm, there were no tipping or notches, and the tool life was significantly improved. The study shows that the LAM of AerMet100 steel has obvious advantages in improving surface quality and reducing tool wear.
ABSTRACT
A unique feature of coronaviruses is their utilization of self-encoded nonstructural protein 16 (nsp16), 2'-O-methyltransferase (2'-O-MTase), to cap their RNAs through ribose 2'-O-methylation modification. This process is crucial for maintaining viral genome stability, facilitating efficient translation, and enabling immune escape. Despite considerable advances in the ultrastructure of SARS-CoV-2 nsp16/nsp10, insights into its molecular mechanism have so far been limited. In this study, we systematically characterized the 2'-O-MTase activity of nsp16 in SARS-CoV-2, focusing on its dependence on nsp10 stimulation. We observed cross-reactivity between nsp16 and nsp10 in various coronaviruses due to a conserved interaction interface. However, a single residue substitution (K58T) in SARS-CoV-2 nsp10 restricted the functional activation of MERS-CoV nsp16. Furthermore, the cofactor nsp10 effectively enhanced the binding of nsp16 to the substrate RNA and the methyl donor S-adenosyl-l-methionine (SAM). Mechanistically, His-80, Lys-93, and Gly-94 of nsp10 interacted with Asp-102, Ser-105, and Asp-106 of nsp16, respectively, thereby effectively stabilizing the SAM binding pocket. Lys-43 of nsp10 interacted with Lys-38 and Gly-39 of nsp16 to dynamically regulate the RNA binding pocket and facilitate precise binding of RNA to the nsp16/nsp10 complex. By assessing the conformational epitopes of nsp16/nsp10 complex, we further determined the critical residues involved in 2'-O-MTase activity. Additionally, we utilized an in vitro biochemical platform to screen potential inhibitors targeting 2'-O-MTase activity. Overall, our results significantly enhance the understanding of viral 2'-O methylation process and mechanism, providing valuable targets for antiviral drug development.
ABSTRACT
BACKGROUND: Anti-PD-1-based immunotherapy has limited benefits in patients with pancreatic cancer. Accumulating data indicate that natural products exert antitumor activity by remodeling the tumor immune microenvironment. It has been reported that neogambogic acid (NGA), an active natural monomer extracted from Garcinia, has anti-inflammatory and antitumor effects. Nevertheless, there are few systematic studies on the antitumor efficacy and immunomodulatory effects of NGA in pancreatic cancer. METHODS: An orthotopic mouse model of pancreatic cancer was established and were treated with different doses of NGA. Tumor growth and ascites were observed. Flow cytometry and immunohistochemistry (IHC) were used to investigate the tumor immune microenvironment. CD11b+ MDSCs were infused back into mice with pancreatic cancer to observe tumor progression after NGA treatment. Bone marrow cells were induced to differentiate into MDSCs, and the effects of NGA on MDSCs were analyzed and the underlying mechanism was explored. The effects of NGA combined with an anti-PD-1 antibody on pancreatic cancer were further tested. RESULTS: NGA significantly inhibited the tumor growth and improve ascites character in pancreatic cancer model mice. Flow cytometry and IHC analysis revealed that NGA decreased the MDSCs proportion and infiltration in the tumor microenvironment. Moreover, adoptive MDSCs largely attenuated the inhibitory effect of NGA on the progression of pancreatic cancer. In addition, we showed that NGA significantly promoted apoptosis and inhibited the differentiation, migration and immunosuppressive function of MDSCs and decreased level of STAT3 and p-STAT3. Furthermore, we demonstrated that NGA synergistically enhanced the efficacy of anti-PD-1 antibodies against pancreatic cancer. CONCLUSION: NGA inhibited the progression of pancreatic cancer by inhibiting MDSCs in the tumor microenvironment, and enhanced the efficacy of anti-PD-1 therapy in the treatment of pancreatic cancer.
Subject(s)
Immune Checkpoint Inhibitors , Myeloid-Derived Suppressor Cells , Pancreatic Neoplasms , Programmed Cell Death 1 Receptor , Tumor Microenvironment , Animals , Pancreatic Neoplasms/immunology , Pancreatic Neoplasms/drug therapy , Pancreatic Neoplasms/therapy , Pancreatic Neoplasms/pathology , Myeloid-Derived Suppressor Cells/immunology , Myeloid-Derived Suppressor Cells/drug effects , Mice , Tumor Microenvironment/drug effects , Tumor Microenvironment/immunology , Cell Line, Tumor , Programmed Cell Death 1 Receptor/antagonists & inhibitors , Programmed Cell Death 1 Receptor/immunology , Immune Checkpoint Inhibitors/pharmacology , Immune Checkpoint Inhibitors/therapeutic use , Humans , Mice, Inbred C57BL , Immunotherapy/methods , STAT3 Transcription Factor/metabolismABSTRACT
Background: Although previous studies have shown that the injection of contrast agents can improve image quality, the specific impact of this on T2-weighted fat-suppressed (T2 FS) and diffusion-weighted imaging (DWI) sequences in the diagnosis of breast cancer remains incompletely understood. In particular, there is insufficient research on how contrast agents affect the signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), and apparent diffusion coefficient (ADC) values within these sequences, and how these changes influence the diagnosis of benign and malignant breast tumors. Methods: Breast magnetic resonance images (MRI) were obtained from 178 consecutive patients on a 3T scanner. The SNR and CNR of lesions on T2 FS sequence were calculated before and after contrast agent injection and compared. Differences between pre- and post-contrast ADC in identifying different tumor types were compared using the Kruskal-Wallis H-test and the paired comparison test. The accuracy of ADC values between pre- and post-contrast in distinguishing benign and malignant breast masses was assessed using receiver operating characteristic (ROC) curves. Results: The SNR and CNR of T2 FS sequence increased after contrast injection, and especially for invasive cancer and benign tumor, the increase was significant. For DWI, there was a slight increase or decrease of ADC values after contrast injection, but the ADC values before and after contrast had a similar effect in identifying different types of tumors. In the ROC curve analysis for assessing benign and malignant breast tumors, the area under the curve (AUC) before and after contrast showed similar results. Conclusions: Contrast agent injection can improve the SNR and CNR of T2 FS sequence, thus providing higher quality images for the diagnosis of breast lesions. Furthermore, injection of contrast agent had little effect on the ability of ADC values to identify different types of lesions and both ADC values before and after the contrast agent were able to distinguish between benign and malignant tumors with almost the same accuracy.
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Animal models of COVID-19 facilitate the development of vaccines and antivirals against SARS-CoV-2. The efficacy of antivirals or vaccines may differ in different animal models with varied degrees of disease. Here, we introduce a mouse model expressing human angiotensin-converting enzyme 2 (ACE2). In this model, ACE2 with the human cytokeratin 18 promoter was knocked into the Hipp11 locus of C57BL/6J mouse by CRISPR - Cas9 (K18-hACE2 KI). Upon intranasal inoculation with high (3 × 105 PFU) or low (2.5 × 102 PFU) dose of SARS-CoV-2 wildtype (WT), Delta, Omicron BA.1, or Omicron BA.2 variants, all mice showed obvious infection symptoms, including weight loss, high viral loads in the lung, and interstitial pneumonia. 100% lethality was observed in K18-hACE2 KI mice infected by variants with a delay of endpoint for Delta and BA.1, and a significantly attenuated pathogenicity was observed for BA.2. The pneumonia of infected mice was accompanied by the infiltration of neutrophils and pulmonary fibrosis in the lung. Compared with K18-hACE2 Tg mice and HFH4-hACE2 Tg mice, K18-hACE2 KI mice are more susceptible to SARS-CoV-2. In the antivirals test, REGN10933 and Remdesivir had limited antiviral efficacies in K18-hACE2 KI mice upon the challenge of SARS-CoV-2 infections, while Nirmatrelvir, monoclonal antibody 4G4, and mRNA vaccines potently protected the mice from death. Our results suggest that the K18-hACE2 KI mouse model is lethal and stable for SARS-CoV-2 infection, and is practicable and stringent to antiviral development.
Subject(s)
Angiotensin-Converting Enzyme 2 , Antiviral Agents , COVID-19 , Disease Models, Animal , Mice, Inbred C57BL , SARS-CoV-2 , Animals , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/virology , Mice , SARS-CoV-2/genetics , SARS-CoV-2/immunology , SARS-CoV-2/drug effects , Antiviral Agents/pharmacology , Humans , Lung/virology , Lung/pathology , COVID-19 Drug Treatment , Keratin-18/genetics , Viral Load , Adenosine Monophosphate/analogs & derivatives , Adenosine Monophosphate/pharmacology , Adenosine Monophosphate/therapeutic use , Alanine/analogs & derivatives , Alanine/pharmacology , Gene Knock-In Techniques , Antibodies, Viral/immunology , Antibodies, Viral/blood , FemaleABSTRACT
Peritoneal metastasis is an important cause of high mortality and poor prognosis in colorectal cancer (CRC) patients. Therefore, the development of compounds with unique anti-CRC Peritoneal metastasis activities is urgently needed to improve the survival of CRC patients. Hydroxygenkwanin (HGK),a natural flavonoid compound, have been shown to display anti-inflammatory, antioxidant, antitumor, and immunoregulatory effects. Here, we employed CRC peritoneal metastasis mouse model with MC38 cells to examine the antitumor activity of HGK. The result showed that HGK not only inhibited peritoneal metastasis, but also significantly increased the proportion of M1-like macrophages while decreasing the proportion of M2-like macrophages within the tumor microenvironment (TME). Furthermore, we demonstrated that the inhibitory effect of HGK on peritoneal metastasis of CRC depended on macrophages in vitro and in vivo. Moreover, we revealed that HGK promoted the polarization of TAMs into M1-like macrophages and inhibited their polarization into M2-like macrophages in a LPS- or IL-4-induced bone marrow-derived macrophages (BMDMs) model and co-culture system. Finally, we also investigated the regulatory mechanism of HGK on TAMs polarization that HGK may active p-STAT5, p-NF-κB signaling in M1-like macrophages and inhibit p-STAT6, JMJD3, PPARγ expression in M2-like macrophages. Taken together, our findings suggest that HGK is a natural candidate for effective prevention of peritoneal metastasis in colorectal cancer, which provides a potential strategy for clinical treatment of colorectal cancer.
Subject(s)
Colorectal Neoplasms , Mice, Inbred C57BL , Peritoneal Neoplasms , Tumor-Associated Macrophages , Animals , Colorectal Neoplasms/pathology , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/metabolism , Mice , Peritoneal Neoplasms/secondary , Peritoneal Neoplasms/drug therapy , Peritoneal Neoplasms/prevention & control , Tumor-Associated Macrophages/drug effects , Tumor-Associated Macrophages/metabolism , Cell Line, Tumor , Signal Transduction/drug effects , Tumor Microenvironment/drug effects , NF-kappa B/metabolism , Humans , MaleABSTRACT
Previous studies through targeted mutagenesis of K-D-K-E motif have demonstrated that 2'-O-MTase activity is essential for efficient viral replication and immune evasion. However, the K-D-K-E catalytic motif of 2'-O-MTase is highly conserved across numerous viruses, including flaviviruses, vaccinia viruses, coronaviruses, and extends even to mammals. Here, we observed a stronger 2'-O-MTase activity in SARS-CoV-2 compared to SARS-CoV, despite the presence of a consistently active catalytic center. We further identified critical residues (Leu-36, Asn-138 and Ile-153) which served as determinants of discrepancy in 2'-O-MTase activity between SARS-CoV-2 and SARS-CoV. These residues significantly enhanced the RNA binding affinity of 2'-O-MTase and boosted its versatility toward RNA substrates. Of interest, a triple substitution (Leu36 â Ile36, Asn138 â His138, Ile153 â Leu153, from SARS-CoV-2 to SARS-CoV) within nsp16 resulted in a proportional reduction in viral 2'-O-methylation and impaired viral replication. Furthermore, it led to a significant upregulation of type I interferon (IFN-I) and proinflammatory cytokines both in vitro and vivo, relying on the cooperative sensing of melanoma differentiation-associated protein 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2). In conclusion, our findings demonstrated that alterations in residues other than K-D-K-E of 2'-O-MTase may affect viral replication and subsequently influence pathogenesis. Monitoring changes in nsp16 residues is crucial as it may aid in identifying and assessing future alteration in viral pathogenicity resulting from natural mutations occurring in nsp16.
Subject(s)
COVID-19 , Methyltransferases , SARS-CoV-2 , Virus Replication , Humans , SARS-CoV-2/genetics , SARS-CoV-2/enzymology , SARS-CoV-2/pathogenicity , COVID-19/virology , COVID-19/genetics , Methyltransferases/genetics , Methyltransferases/metabolism , Methyltransferases/chemistry , Virus Replication/genetics , RNA, Viral/genetics , RNA, Viral/metabolism , RNA, Viral/chemistry , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/metabolism , Severe acute respiratory syndrome-related coronavirus/genetics , Severe acute respiratory syndrome-related coronavirus/enzymology , Severe acute respiratory syndrome-related coronavirus/pathogenicity , Animals , Interferon-Induced Helicase, IFIH1/genetics , Interferon-Induced Helicase, IFIH1/metabolismABSTRACT
During enteric nervous system (ENS) development, pioneering wavefront enteric neural crest cells (ENCCs) initiate gut colonization. However, the molecular mechanisms guiding their specification and niche interaction are not fully understood. We used single-cell RNA sequencing and spatial transcriptomics to map the spatiotemporal dynamics and molecular landscape of wavefront ENCCs in mouse embryos. Our analysis shows a progressive decline in wavefront ENCC potency during migration and identifies transcription factors governing their specification and differentiation. We further delineate key signaling pathways (ephrin-Eph, Wnt-Frizzled, and Sema3a-Nrp1) utilized by wavefront ENCCs to interact with their surrounding cells. Disruptions in these pathways are observed in human Hirschsprung's disease gut tissue, linking them to ENS malformations. Additionally, we observed region-specific and cell-type-specific transcriptional changes in surrounding gut tissues upon wavefront ENCC arrival, suggesting their role in shaping the gut microenvironment. This work offers a roadmap of ENS development, with implications for understanding ENS disorders.
Subject(s)
Cell Movement , Enteric Nervous System , Neural Crest , Signal Transduction , Animals , Neural Crest/metabolism , Neural Crest/cytology , Mice , Enteric Nervous System/metabolism , Enteric Nervous System/embryology , Enteric Nervous System/cytology , Embryo, Mammalian/metabolism , Embryo, Mammalian/cytology , Cell Differentiation , Gene Expression Regulation, Developmental , Hirschsprung Disease/genetics , Hirschsprung Disease/metabolism , Hirschsprung Disease/pathology , HumansABSTRACT
BACKGROUND: Recent studies have demonstrated that APOC1 is associated with cancer progression, exerting cancer-promoting and immune infiltration-promoting effects. Nevertheless, there is currently no report on the presence of APOC1 in ovarian cancer (OV). METHOD: In this study, we conducted data analysis using the GEO and TCGA databases. We conducted a thorough bioinformatics analysis to investigate the function of APOC1 in OV, utilizing various platforms including cBioPortal, STRING, GeneMANIA, LinkedOmics, GSCALite, TIMER, and CellMarker. Additionally, we performed immunohistochemical staining on tissue microarrays and conducted in vitro cellular assays to validate our findings. RESULT: Our findings reveal that APOC1 expression is significantly upregulated in OV compared to normal tissues. Importantly, patients with high APOC1 levels show a significantly poorer prognosis. Furthermore, our study demonstrated that APOC1 exerted a crucial function in promoting the capacity of ovarian cancer cells to proliferate, migrate, and invade. Additionally, we have identified that genes co-expressed with APOC1 are primarily associated with adaptive immune responses. Notably, the levels of APOC1 in OV exhibit a correlation with the presence of M2 Tumor-associated Macrophages (TAMs). CONCLUSION: APOC1 emerges as a promising prognostic biomarker for OV and exhibits a significant association with M2 TAMs in OV.
Subject(s)
Ovarian Neoplasms , Female , Humans , Biomarkers , Macrophages , Ovarian Neoplasms/genetics , PrognosisABSTRACT
Objective: Due to the information-rich nature of positron emission tomography/computed tomography (PET/CT) images, the authors hope to explore radiomics features that could distinguish metastatic lymph nodes (LNs) from hypermetabolic benign LNs, in addition to conventional indicators. Methods: PET/CT images of 106 patients with early-stage cervical cancer from 2019 to 2021 were retrospectively analyzed. The tumor lesions and LN regions of PET/CT images were outlined with SeeIt, and then radiomics features were extracted. The least absolute shrinkage and selection operator (LASSO) was used to select features. The final selected radiomics features of LNs were used as predictors to construct a machine learning model to predict LN metastasis. Results: The authors determined two morphological coefficient characteristics of cervical lesions (shape - major axis length and shape - mesh volume), one first order characteristics of LNs (first order - 10 percentile) and two gray-level co-occurrence matrix (GLCM) characteristics of LNs (GLCM - id and GLCM - inverse variance) were closely related to LN metastasis. Finally, a neural network was constructed based on the radiomic features of the LNs. The area under the curve of receiver operating characteristic (AUC-ROC) of the model was 0.983 in the training set and 0.860 in the test set. Conclusion: The authors constructed and demonstrated a neural network based on radiomics features of PET/CT to evaluate the risk of single LN metastasis in early-stage cervical cancer.
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PURPOSE: To investigate the diagnostic efficacy of T1ρ dispersion and Gd-EOB-DTPAenhanced T1mapping in the identification of early liver fibrosis (LF) and non-alcoholic steatohepatitis (NASH) in a non-alcoholic fatty liver disease (NAFLD) rabbit model induced by a high-fat diet using histopathological findings as the standard reference. METHODS: A total of sixty rabbits were randomly allocated into the standard control group (n = 12) and the NAFLD model groups (8 rabbits per group) corresponding to different high-fat high cholesterol diet feeding weeks. All rabbits underwent noncontrast transverse T1ρ mapping with varying spin-locking frequencies (FSL = 0 Hz and 500 Hz), native T1 mapping, and Gd-EOB-DTPA-enhanced T1 mapping during the hepatobiliary phase. The histopathological findings were assessed based on the NASH CRN Scoring System. Statistical analyses were conducted using the intraclass correlation coefficient, analysis of variance, multiple linear regression, and receiver operating characteristics. RESULTS: Except for native T1, T1ρ, T1ρ dispersion, HBP T1, and â³T1 values significantly differed among different liver fibrosis groups (F = 14.414, 18.736, 10.15, and 9.799, respectively; all P < 0.05). T1ρ, T1ρ dispersion, HBP T1, and â³T1 values also exhibited significant differences among different NASH groups (F = 4.138, 4.594, 21.868, and 22.678, respectively; all P < 0.05). In the multiple regression analysis, liver fibrosis was the only factor that independently influenced T1ρ dispersion (R2 = 0.746, P = 0.000). Among all metrics, T1ρ dispersion demonstrated the best area under curve (AUC) for identifying early LF (≥ F1 stage) and significant LF (≥ F2 stage) (AUC, 0.849 and 0.916, respectively). The performance of â³T1 and HBP T1 (AUC, 0.948 and 0.936, respectively) were better than that of T1ρ and T1ρ dispersion (AUC, 0.762 and 0.769, respectively) for diagnosing NASH. CONCLUSION: T1â´ dispersion may be suitable for detecting liver fibrosis in the complex background of NAFLD, while Gd-EOB-DTPA enhanced T1 mapping is superior to nonenhanced T1â´ mapping (T1â´ and T1â´ dispersion) for identifying NASH.
Subject(s)
Non-alcoholic Fatty Liver Disease , Animals , Rabbits , Non-alcoholic Fatty Liver Disease/pathology , Liver/diagnostic imaging , Liver/pathology , Contrast Media , Magnetic Resonance Imaging , Gadolinium DTPA , Liver Cirrhosis/pathology , Risk AssessmentABSTRACT
Interspecific variations in phenotypic plasticity of trees that are affected by climate change may alter the ecosystem function of forests. Seedlings of four common tree species (Castanopsis fissa, Michelia macclurei, Dalbergia odorifera and Ormosia pinnata) in subtropical plantations of southern China were grown in the field under rainout shelters and subjected to changing precipitation (48 L of water every 4 days in the dry season, 83 L of water every 1 day in the wet season; 4 g m-2 year-1 of nitrogen (N)), low N deposition (48 L of water every 2 days in the dry season, 71 L of water every 1 day in the wet season; 8 g m-2 year-1 N), high N deposition (48 L of water every 2 days in the dry season, 71 L of water every 1 day in the wet season; 10 g m-2 year-1 N) and their interactive effects. We found that the changes in seasonal precipitation reduced the light-saturated photosynthetic rate (Asat) for C. fissa due to declining area-based foliar N concentrations (Na). However, we also found that the interactive effects of changing precipitation and N deposition enhanced Asat for C. fissa by increasing foliar Na concentrations, suggesting that N deposition could alleviate N limitations associated with changing precipitation. Altered precipitation and high N deposition reduced Asat for D. odorifera by decreasing the maximum electron transport rate for RuBP regeneration (Jmax) and maximum rate of carboxylation of Rubisco (Vcmax). Ormosia pinnata under high N deposition exhibited increasing Asat due to higher stomatal conductance and Vcmax. The growth of D. odorifera might be inhibited by changes in seasonal precipitation and N deposition, while O. pinnata may benefit from increasing N deposition in future climates. Our study provides an important insight into the selection of tree species with high capacity to tolerate changing precipitation and N deposition in subtropical plantations.
Subject(s)
Ecosystem , Trees , Trees/physiology , Seedlings , Seasons , Photosynthesis , Nitrogen , WaterABSTRACT
Streptococcus pneumoniae is an opportunistic pathogen that causes substantial illness and death among children worldwide. The genetic backgrounds of pneumococci that cause infection versus asymptomatic carriage vary substantially. To determine the evolutionary mechanisms of opportunistic pathogenicity, we conducted a genomic surveillance study in China. We collected 783 S. pneumoniae isolates from infected and asymptomatic children. By using a 2-stage genomewide association study process, we compared genomic differences between infection and carriage isolates to address genomic variation associated with pathogenicity. We identified 8 consensus k-mers associated with adherence, antimicrobial resistance, and immune modulation, which were unevenly distributed in the infection isolates. Classification accuracy of the best k-mer predictor for S. pneumoniae infection was good, giving a simple target for predicting pathogenic isolates. Our findings suggest that S. pneumoniae pathogenicity is complex and multifactorial, and we provide genetic evidence for precise targeted interventions.
Subject(s)
Biological Evolution , Streptococcus pneumoniae , Child , Humans , Streptococcus pneumoniae/genetics , China/epidemiology , Genome-Wide Association Study , Genetic VariationABSTRACT
Chronic hepatitis B virus (HBV) infection is a major cause of liver cirrhosis and liver cancer, despite strong prevention and treatment efforts. The study of the epigenetic modification of HBV has become a research hotspot, including the N6-methyladenosine (m6A) modification of HBV RNA, which plays complex roles in the HBV life cycle. In addition to m6A modification, 5-methylcytosine (m5C) is another major modification of eukaryotic mRNA. In this study, we explored the roles of m5C methyltransferase and demethyltransferase in the HBV life cycle. The results showed that m5C methyltransferase NSUN2 deficiency could negatively regulate the expression of HBV while m5C demethyltransferase TET2 deficiency positively regulates the expression of HBV. Subsequently, we combined both in vitro bisulfite sequencing and high-throughput bisulfite sequencing methods to determine the distribution and stoichiometry of m5C modification in HBV RNA. Two sites: C2017 and C131 with the highest-ranking methylation rates were identified, and mutations at these two sites could lead to the decreased expression and replication of HBV, while the mutation of the "fake" m5C site had no effect. Mechanistically, NSUN2-mediated m5C modification promotes the stability of HBV RNA. In addition, compared with wild-type HepG2-NTCP cells and primary human hepatocytes, the replication level of HBV after NSUN2 knockdown decreased, and the ability of the mutant virus to infect and replicate in wild-type HepG2-NTCP cells and PHHs was substantially impaired. Similar results were found in the experiments using C57BL/6JGpt-Nsun2+/- mice. Interestingly, we also found that HBV expression and core protein promoted the endogenous expression of NSUN2, which implied a positive feedback loop. In summary, our study provides an accurate and high-resolution m5C profile of HBV RNA and reveals that NSUN2-mediated m5C modification of HBV RNA positively regulates HBV replication by maintaining RNA stability.
Subject(s)
Hepatitis B virus , Hepatitis B, Chronic , Animals , Humans , Mice , Hepatitis B virus/genetics , Hepatitis B, Chronic/genetics , Methyltransferases/genetics , Mice, Inbred C57BL , RNAABSTRACT
BACKGROUND: Radical hysterectomy (RH) is commonly used to treat early-stage cervical cancer in women of childbearing age and sexual dysfunction due to postoperative vaginal shortening is a major concern. The impact of intraoperative vaginoplasty on prognosis and quality of sexual life in patients with early-stage cervical cancer remains controversial and lacks high-level evidence. However, there are few reports on vaginoplasty after RH to lengthen vagina in patients. This prospective, multi-center, randomized controlled trial aims to explore the impact of peritoneal vaginoplasty with or without ovarian transposition after laparoscopic RH on sexual dysfunction in patients with early-stage cervical cancer. METHODS: Eligible patients will be randomly assigned (1:1) to receive peritoneal vaginoplasty or not. The primary evaluation indicators are female sexual function index (FSFI) and male sexual satisfaction scale. The secondary evaluation indicators include EORTC QLQ-CX24, 2-year overall survival (OS), 5-year OS, 2-year progression-free survival (PFS), 5-year PFS and surgery-related complications. The trial will enroll 368 patients from 6 hospitals in China over a 3-year period and follow up for 5 years. TRIAL REGISTRATION: Chinese Clinical Trial Registry Identifier: ChiCTR2000040610.
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Cervical cancer (CC) is the fourth most gynecological malignancy in the world. The identification of predictive markers can provide a basis for personalized treatment and prognostic evaluation. Our aim was to identify a new predictive marker of epiregulin (EREG) gene and explore its functional characteristics of CC and other cancer types. Differentially highly expressed genes were obtained from Gene Expression Omnibus (GEO) databases. Key genes can be verified by the Cancer Genome Atlas (TCGA) and Genotype Tissue Expression (GTEx) data, and the functions of these genes were investigated through gene ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Survival analysis were performed to determine the key genes (EREG) related to the prognosis of CC. Then, the expression difference of EREG between tumor and normal tissue was evaluated by real-time polymerase chain reaction (PCR), western blotting, and immunohistochemistry. The relationship between EREG and prognosis of patients, immune microenvironment, immune checkpoint, immune therapy and angiogenesis was discussed in 33 tumor types. Finally, the regulatory mechanism of EREG on human umbilical vein endothelial cells (HUVECs) was also explored. The differential analysis results from multiple databases showed that EREG was significantly highly expressed in CC, which was further verified in Hela and Siha cell lines. Then, Survival analysis revealed that EREG was associated with the prognosis of CC and other tumor types, and high EREG expression was significantly associated with poor prognosis. In addition, in almost all tumor types, the expression of EREG was related to immune cells infiltration, immune checkpoint genes expression and immunotherapy. Further analysis exhibited that high EREG expression can promote the high expression of angiogenesis related genes. The experimental data demonstrated that EREG could promote the proliferative, migration, invasive and tube formation of HUVECs by interacting with receptors, such as epidermal growth factor receptor (EGFR and ERBB4). EREG may be an independent prognostic marker for predicting tumor prognosis and immunotherapy response of various cancers, and may be a potential target of tumor anti-angiogenic therapy in CC.
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Background: RNA methylation is a significant form of post-transcriptional modification that has been implicated in various diseases, including cancers. One prominent type of RNA methylation is 5-Methylcytosine (m5C), which primarily regulates RNA stability, transcription, and translation. However, the role of m5C-related gene regulation in cell adhesion within uterine corpus endometrial carcinoma (UCEC) remains unexplored. Therefore, the objective of this study was to investigate the association between RNA m5C methylation and UCEC and develop a prognostic predictive model to forecast survival outcomes in UCEC patients. Methods: The RNA datasets were acquired from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. The dataset was used to explore the interaction relationships of m5C regulators in UCEC. Unsupervised clustering analysis identified clusters with distinct m5C modification patterns. Different clusters underwent Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment level analysis to investigate the effects of pathways related to m5C methylation, which were further validated through in vitro cellular experiments. A prognostic predictive model was developed using the least absolute shrinkage and selection operator (LASSO) and multivariate regression analysis. Results: Two clusters with distinct m5C modification patterns were identified using unsupervised cluster analysis. Furthermore, the prognosis of cluster 2 was found to be worse. Enrichment analysis showed alterations in cell adhesion-related pathways in both clusters, as well as differences between the clusters. Through this analysis, we identified 25 genes with significant prognostic value. Finally, a prognostic predictive model comprising NSUN2 and YBX1 was constructed. Conclusions: In conclusion, diverse m5C modification patterns display distinct cell adhesion properties in UCEC, which are correlated with prognosis and offer significant potential as prognostic markers for UCEC assessment. We developed a prognostic predictive model to accurately predict the prognosis of UCEC.
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RNA methyltransferase nucleolar protein p120 (NOP2), commonly referred to as NOP2/Sun RNA methyltransferase family member 1 (NSUN1), is involved in cell proliferation and is highly expressed in various cancers. However, its role in high-grade serous ovarian cancer (HGSOC) remains unclear. Our study investigated the expression of NOP2 in HGSOC tissues and normal fimbria tissues, and found that NOP2 was significantly upregulated in HGSOC tissues. Our experiments showed that NOP2 overexpression promoted cell proliferation in vivo and in vitro and increased the migration and invasion ability of HGSOC cells in vitro. Furthermore, we identified Rap guanine nucleotide exchange factor 4 (RAPGEF4) as a potential downstream target of NOP2 in HGSOC. Finally, our findings suggest that the regulation of NOP2 and RAPGEF4 may depend on m5C methylation levels.
Subject(s)
Ovarian Neoplasms , RNA , Humans , Female , Methyltransferases/genetics , Ovarian Neoplasms/genetics , Cell Proliferation , Nuclear Proteins/metabolism , Guanine Nucleotide Exchange Factors , tRNA Methyltransferases/genetics , tRNA Methyltransferases/metabolismABSTRACT
In this study, we analyzed high-frequency data pertaining to phytoplankton and environmental factors in Jiaozhou Bay from December 10, 2004, to December 10, 2005 and from July 26, 2020, to August 1, 2021. Compared with 2004-2005, the abundance of phytoplankton during 2020-2021 presented a "two-peak pattern" of annual variation, and the number of species decreased significantly. The ecological types were dominated by eurythermal species, while cold-water species declined. The overall abundance showed a downward trend, and the peak period moved to a time with lower water temperature. This could be attributed to the increase in sea surface temperature. The increase in water temperature and intensification of eutrophication made the emergence of dominant species more inclined to be single; the diversity and stability of the community structure decreased. Generalized additive model (GAM) and network analysis showed that temperature was the main driving factor influencing the phytoplankton community structure, especially during the peak period. In addition, nitrogen and phosphorus were important factors influencing species composition and competitive advantage of phytoplankton.