In-Depth Serum Proteomics Reveals the Trajectory of Hallmarks of Cancer in Hepatitis B Virus-Related Liver Diseases.

Hepatocellular carcinoma (HCC) is a prevalent cancer in China, with chronic hepatitis B (CHB) and liver cirrhosis (LC) being high-risk factors for developing HCC. Here, we determined the serum proteomes (762 proteins) of 125 healthy controls and Hepatitis B virus-infected CHB, LC, and HCC patients and constructed the first cancerous trajectory of liver diseases. The results not only reveal that the majority of altered biological processes were involved in the hallmarks of cancer (inflammation, metastasis, metabolism, vasculature, and coagulation) but also identify potential therapeutic targets in cancerous pathways (i.e., IL17 signaling pathway). Notably, the biomarker panels for detecting HCC in CHB and LC high-risk populations were further developed using machine learning in two cohorts comprised of 200 samples (discovery cohort = 125 and validation cohort = 75). The protein signatures significantly improved the area under the receiver operating characteristic curve of HCC (CHB discovery and validation cohort = 0.953 and 0.891, respectively; LC discovery and validation cohort = 0.966 and 0.818, respectively) compared to using the traditional biomarker, alpha-fetoprotein, alone. Finally, selected biomarkers were validated with parallel reaction monitoring mass spectrometry in an additional cohort (n = 120). Altogether, our results provide fundamental insights into the continuous changes of cancer biology processes in liver diseases and identify candidate protein targets for early detection and intervention.

Respiratory immune status and microbiome in recovered COVID-19 patients revealed by metatranscriptomic analyses.

Coronavirus disease 2019 (COVID-19) is currently a severe threat to global public health, and the immune response to COVID-19 infection has been widely investigated. However, the immune status and microecological changes in the respiratory systems of patients with COVID-19 after recovery have rarely been considered. We selected 72 patients with severe COVID-19 infection, 57 recovered from COVID-19 infection, and 65 with non-COVID-19 pneumonia, for metatranscriptomic sequencing and bioinformatics analysis. Accordingly, the differentially expressed genes between the infected and other groups were enriched in the chemokine signaling pathway, NOD-like receptor signaling pathway, phagosome, TNF signaling pathway, NF-kappa B signaling pathway, Toll-like receptor signaling pathway, and C-type lectin receptor signaling pathway. We speculate that IL17RD, CD74, and TNFSF15 may serve as disease biomarkers in COVID-19. Additionally, principal coordinate analysis revealed significant differences between groups. In particular, frequent co-infections with the genera Streptococcus, Veillonella, Gemella, and Neisseria, among others, were found in COVID-19 patients. Moreover, the random forest prediction model with differential genes showed a mean area under the curve (AUC) of 0.77, and KCNK12, IL17RD, LOC100507412, PTPRT, MYO15A, MPDZ, FLRT2, SPEG, SERPINB3, and KNDC1 were identified as the most important genes distinguishing the infected group from the recovered group. Agrobacterium tumefaciens, Klebsiella michiganensis, Acinetobacter pittii, Bacillus sp. FJAT.14266, Brevundimonas naejangsanensis, Pseudopropionibacterium propionicum, Priestia megaterium, Dialister pneumosintes, Veillonella rodentium, and Pseudomonas protegens were selected as candidate microbial markers for monitoring the recovery of COVID patients. These results will facilitate the diagnosis, treatment, and prognosis of COVID patients recovering from severe illness.

The expression of cuproptosis-related genes in hepatocellular carcinoma and their relationships with prognosis.

Background: The mechanism of cuproptosis has recently been reported in lipoylated proteins of the tricarboxylic acid (TCA) cycle. Besides, the role of copper was previously recognized in cancer progression. We evaluated the prognostic value of cuproptosis-related gene expression in hepatocellular carcinoma (HCC). Methods: Remarkable genes were selected both in differential expression analysis and Kaplan-Meier survival analysis from ninety-six cuproptosis-related genes using The Cancer Genome Atlas (TCGA) database. The relationships between clinical characteristics and gene expression were performed with Wilcoxon signed-rank test, Kruskal-Wallis test, and logistic regression. Clinicopathologic factors correlated with overall survival in HCCs conducting univariate and multivariate Cox regression analysis. Gene Expression Profiling Interactive Analysis 2 (GEPIA2) and Human Protein Atlas (HPA) databases were utilized to verify the results. Furthermore, Gene Set Enrichment Analysis (GSEA) identified the potential key pathways that dominate cuproptosis in HCC. Results: Elevated ATP7A, SLC25A3, SCO2, COA6, TMEM199, ATP6AP1, LIPT1, DLAT, PDHA1, MTF1, ACP1, FDX2, NUBP2, CIAPIN1, ISCA2 and NDOR1 expression, as well as declined AOC1, FDX1, MT-CO1, and ACO1 expression were significantly emerged in HCC tumor tissues and were significantly associated with HCCs poor survival. The expressions of screened cuproptosis-related genes were prominently related to clinical features. GSEA analysis reported many key signaling pathways (such as natural killer cell mediated cytotoxicity, TCA cycle, glutathione metabolism, ATP-binding cassette (ABC) transporters, Notch signaling pathway, ErbB signaling pathway, and metabolism of xenobiotics by cytochrome p450) were differentially enriched in HCCs with varying degrees of cuproptosis-related genes expression. Conclusions: The twenty cuproptosis-related genes might be utilized as new candidate prognostic biomarkers for HCC.

The association between several autophagy-related genes and their prognostic values in hepatocellular carcinoma: a study on the foundation of TCGA, GEPIA and HPA databases.

BACKGROUND: The purpose of this study was to investigate the relationship between the expression of autophagy-related genes and prognosis in hepatocellular carcinoma (HCC). METHODS AND RESULTS: We selected three autophagy-related genes (ATG3, ATG7, and ATG9A) from gene expression data of liver cancer patients in The Cancer Genome Atlas (TCGA) database by Kaplan-Meier survival analysis, univariate and multivariate Cox regression analysis, and Gene Set Enrichment Analysis (GSEA). Human Protein Atlas (HPA) and Gene Expression Profiling Interactive Analysis (GEPIA) databases were applied to testify the credibility of our results. The expression levels of ATG3, ATG7, and ATG9A were verified by real-time quantitative PCR (RT-qPCR) in normal liver cells (L02) and three HCC cell lines (HepG2, Hep3b, and Li-7). Data analysis results from TCGA showed high ATG3, ATG7, ATG9A expression in HCC tumor tissues. Kaplan-Meier survival analysis showed that the survival rate of the high expression group of ATG3, ATG7, and ATG9A was all significantly lower than the low expression group. GSEA analysis showed that many signaling pathways (such as the regulation of autophagy, glycine serine and threonine metabolism, pathways in cancer, mitogen-activated protein kinase (MAPK) signaling pathway, mammalian target of rapamycin (mTOR) signaling pathway, as well as P53 signaling pathway) were differentially enriched in HCCs with ATG3, ATG7, and ATG9A expression. GEPIA and RT-qPCR also identified that the mRNA expression level of ATG3, ATG7, and ATG9A in normal liver cells were significantly lower than in HCC cells. High protein expression of ATG3, ATG7, and ATG9A was displayed in HCCs from the HPA database. CONCLUSIONS: The ATG3, ATG7, ATG9A might be utilized as prognostic biomarkers for liver cancer.

Corrigendum: Genomic Characteristics and Pan-Genome Analysis of Rhodococcus equi.

[This corrects the article DOI: 10.3389/fcimb.2022.807610.].

Inhibitor screening using microarray identifies the high capacity of neutralizing antibodies to Spike variants in SARS-CoV-2 infection and vaccination.

Rationale: Mutations of SARS-CoV-2, which is responsible for coronavirus disease 2019 (COVID-19), could impede drug development and reduce the efficacy of COVID-19 vaccines. Here, we developed a multiplexed Spike-ACE2 Inhibitor Screening (mSAIS) assay that can measure the neutralizing effect of antibodies across numerous variants of the coronavirus's Spike (S) protein simultaneously. Methods: The SARS-CoV-2 spike variant protein microarrays were prepared by printing 72 S variants onto a chemically-modified glass slides. The neutralization potential of purified anti-S antibodies and serum from convalescent COVID-19 patients and vaccinees to S variants were assessed with the mSAIS assay. Results: We identified new S mutations that are sensitive and resistant to neutralization. Serum from both infected and vaccinated groups with a high titer of neutralizing antibodies (NAbs) displayed a broader capacity to neutralize S variants than serum with low titer NAbs. These data were validated using serum from a large vaccinated cohort (n = 104) with a tiled S peptide microarray. In addition, similar results were obtained using a SARS-CoV-2 pseudovirus neutralization assay specific for wild-type S and five prevalent S variants (D614G, B.1.1.7, B.1.351, P.1, B.1.617.2), thus demonstrating that high antibody diversity is associated with high NAb titers. Conclusions: Our results demonstrate the utility of the mSAIS platform in screening NAbs. Moreover, we show that heterogeneous antibody populations provide a more protective effect against S variants, which may help direct COVID-19 vaccine and drug development.

Genomic Characteristics Revealed Plasmid-Mediated Pathogenicity and Ubiquitous Rifamycin Resistance of Rhodococcus equi.

Rhodococcus equi is a zoonotic pathogen that can cause fatal disease in patients who are immunocompromised. At present, the epidemiology and pathogenic mechanisms of R. equi infection are not clear. This study characterized the genomes of 53 R. equi strains from different sources. Pan-genome analysis showed that all R. equi strains contained 11481 pan genes, including 3690 core genes and 602 ~ 1079 accessory genes. Functional annotation of pan genome focused on the genes related to basic lifestyle, such as the storage and expression of metabolic and genetic information. Phylogenetic analysis based on pan-genome showed that the R. equi strains were clustered into six clades, which was not directly related to the isolation location and host source. Also, a total of 84 virulence genes were predicted in 53 R. equi strains. These virulence factors can be divided into 20 categories related to substance metabolism, secreted protein and immune escape. Meanwhile, six antibiotic resistance genes (RbpA, tetA (33), erm (46), sul1, qacEdelta 1 and aadA9) were detected, and all strains carried RbpA related to rifamycin resistance. In addition, 28 plasmids were found in the 53 R. equi strains, belonging to Type-A (n = 14), Type-B (n = 8) and Type-N (n = 6), respectively. The genetic structures of the same type of plasmid were highly similar. In conclusion, R. equi strains show different genomic characteristics, virulence-related genes, potential drug resistance and virulence plasmid structures, which may be conducive to the evolution of its pathogenesis.

The Effect and Mechanism of LINC00663 on the Biological Behavior of Glioma.

Glioma is the most frequent primary malignant brain tumor, which is characterized by high incidence and mortality, with a poor prognosis. Numerous studies have revealed the abnormal expression of long non-coding RNAs in gliomas. This study explored the effects and potential mechanism of LINC00663 in glioma. The LINC00663 levels and their prognostic values were analyzed from the GEO databases using bioinformatics. Also, LINC00663 expression in tissue samples and cell lines was measured using qRT-PCR. The roles of LINC00663 in glioma were confirmed using CCK8, EdU assay as well as Transwell tests. Moreover, the influences of LINC00663 on the AKT/mTOR signal cascades were detected using western blotting assay. LINC00663 expression was higher in both glioma tissues and cell lines than that in the normal brain tissues and human astrocytes. High expression of LINC00663 led to the low overall survival rate of patients with glioma. LINC00663 knockdown notably restrained cell proliferation, migration, and invasion abilities by decreasing the activation of AKT and mTOR. This study indicated that LINC00663 might have a cancer-promoting role in accelerating glioma development and progression through regulating AKT/mTOR pathway.

Aryl Hydrocarbon Receptor Promotes IL-10 Expression in Inflammatory Macrophages Through Src-STAT3 Signaling Pathway.

The aryl hydrocarbon receptor (AhR) is an important immune regulator with a role in inflammatory response. However, the role of AhR in IL-10 production by inflammatory macrophages is currently unknown. In this study, we investigated LPS-induced IL-10 expression in macrophages from AhR-KO mice and AhR-overexpressing RAW264.7 cells. AhR was highly expressed after LPS stimulation through NF-κB pathway. Loss of AhR resulted in reduced IL-10 expression in LPS-induced macrophages. Moreover, the IL-10 expression was elevated in LPS-induced AhR-overexpressing RAW264.7 cells. Maximal IL-10 expression was dependent on an AhR non-genomic pathway closely related to Src and STAT3. Furthermore, AhR-associated Src activity was responsible for tyrosine phosphorylation of STAT3 and IL-10 expression by inflammatory macrophages. Adoptive transfer of AhR-expressing macrophages protected mice against LPS-induced peritonitis associated with high IL-10 production. In conclusion, we identified the AhR-Src-STAT3-IL-10 signaling pathway as a critical pathway in the immune regulation of inflammatory macrophages, It suggests that AhR may be a potential therapeutic target in immune response.

[Exogenous agmatine inhibits lipopolysaccharide-induced activation and dysfunction of human umbilical vein endothelial cells].

OBJECTIVE: To investigate whether exogenous agmatine inhibits lipopolysaccharide (LPS)-induced activation and dysfunction of human umbilical vein endothelial cells (HUVECs) by modulating nuclear factor-κB (NF-κB) and MAPK signal pathways and the production of reactive oxygen species (ROS). METHODS: Cultured HUVECs were treated with agmatine at the optimized concentration of 1.0 mmolγL, LPS (10 µgγmL), and LPS + agmatine, with or without pretreatment with the inhibitors of NF-κB (PDTC), p38 (SB203580), and ERK (PD98059) for 1 h. The levels of soluble vascular cell adhesion molecule 1 (VCAM-1), soluble intercellular adhesion molecule 1 (sICAM-1), soluble E-selectin and monocyte chemoattractant protein 1 (MCP-1) in the supernatant were determined using ELISA, and their mRNA expressions, along with heme oxygenase-1 (HO-1), and NAD(P)H: quinone oxidoreductase 1 (NQO-1), were assessed using real-time PCR. ROS production in the cells was determined using 2, 7-dichlorofluoresce in diacetate (DCFH-DA) as the fluorescence probe. The protein expressions of VCAM-1, ICAM-1, p65, phospho-p65 (p-p65), IκBα, p-IκBα, ERK, p-ERK, p38, p-p38, JNK, and p-JNK were detected using Western blotting. RESULTS: LPS stimulation for 6 and 24 h significantly increased the levels of sVCAM-1, sICAM-1, sE-selectin and MCP-1 in the supernatant, intracellular ROS production, and the mRNA expressions of these molecules (P<0.05). Intervention with 1 mmolγL agmatine, similar with pretreatment with p38, ERK and NF-κB inhibitors, obviously inhibited such effects of LPS in HUVECs (P<0.05). Agmatine significantly up-regulated the mRNA expression of HO-1 (P<0.05), inhibited LPS-induced phosphorylation of p38, ERK, nuclear p65 and cytoplasmic IκBα, and up-regulated the protein expression of cytoplasmic IκBα. CONCLUSION: Agmatine inhibits LPS-induced activation and dysfunction of HUVECs by modulating NF-κB and MAPK signal pathways to down-regulate the expressions of adhesion molecules and chemokines and by up-regulating the expression of HO-1 to reduce ROS production.

[Establishment of multiple organ dysfunction syndrome early warning score in patients with severe trauma and its clinical significance: a multicenter study].

OBJECTIVE: To explore the risk factors of multiple organ dysfunction syndrome (MODS) in severe trauma patients, put forward a new warning scoring system of MODS, and to provide a more accurate scoring method for doctors to judge the clinical condition and prognosis of patients. METHODS: Clinical data of 342 patients with severe trauma admitted to intensive care unit (ICU) of the Affiliated Hospital of Zunyi Medical College and Daping Hospital of the Third Military Medical University from January 1st, 2015 to December 31st, 2016 were retrospectively analyzed. The patients were divided into MODS groups (n = 251) and non-MODS group (n = 91) according to clinical outcomes. The clinical data of patients, including gender, age, heart rate (HR) and blood pressure within 24 hours after admission to the hospital, indicators of blood routine and blood biochemistry, severity of disease, severity of trauma, whether received the emergency intubation or surgery within 24 hours or not, whether developed sepsis or acute respiratory distress syndrome (ARDS) during hospitalization, were recorded, and univariate analysis was conducted. The indicators with statistical significance found by univariate analysis were enrolled in multivariate Logistic regression analysis, and the risk factors for MODS in patients with severe trauma were screened and assigned, and the final total score was MODS warning score. Receiver operating characteristic (ROC) curve was plotted to evaluate MODS warning score for predicting the occurrence of MODS in patients with severe trauma. RESULTS: Compared with non-MODS group, HR, Na+, serum creatinine (SCr), activated partial thromboplastin time (APTT), injury severity score (ISS), new injury severity score (NISS), acute physiology and chronic health evaluation II (APACHE II) score and sequential organ failure assessment (SOFA) score in MODS group were significantly increased, pH value, red blood cell (RBC), platelet (PLT), albumin (Alb) and Glasgow coma score (GCS) were remarkably decreased, and multiple injury, the patients with shock at admission, blood transfusion, central venous catheter, emergency intubation and infection were also increased, and more patients suffered from sepsis and ARDS. Multivariate Logistic regression analysis showed that the number of injured places equal or more than 2, shock at admission, APACHE II score ≥ 15, SOFA score ≥ 4 and APTT > 40 s were risk factors for MODS in patients with severe trauma, with total MODS warning score of 7.5. ROC curve analysis showed that the area under ROC curve (AUC) of MODS warning score for predicting MODS in patients with severe trauma was 0.822, which was significantly higher than that of APACHE II score (AUC = 0.698, P < 0.001), APTT (AUC = 0.693, P < 0.001) and SOFA score (AUC = 0.770, P = 0.025). When the cut-off value of MODS warning score was 2.5, the sensitivity was 61.35%, the specificity was 90.11%, and Youden index was 0.515. CONCLUSIONS: MODS warning score is composed of five factors, including the number of injured places, shock at admission, APACHE II score, SOFA score and APTT, which could be regarded as early warning score system for predicting MODS in patients with severe trauma. MODS warning score can be more comprehensive and timely to assess the possibility of MODS and prognosis of patients with severe trauma, and the prediction result is better than the single use of APTT, APACHE II or SOFA score.