Identification of Key Genes Associated with Tumor Microenvironment Infiltration and Survival in Gastric Adenocarcinoma via Bioinformatics Analysis.

OBJECTIVE: Gastric carcinoma (GC) is the fifth most commonly diagnosed cancer and the third leading cause of cancer-related deaths globally. The tumor microenvironment plays a significant role in the pathogenesis, prognosis, and response to immunotherapy. However, the immune-related molecular mechanisms underlying GC remain elusive. Bioinformatics analysis of the gene expression of GC and paracancerous healthy tissues from the same patient was performed to identify the key genes and signaling pathways, as well as their correlation to the infiltration of the tumor microenvironment (TME) by various immune cells related to GC development. METHODS: We employed GSE19826, a gene expression profile from the Gene Expression Omnibus (GEO), for our analysis. Functional enrichment analysis of Differentially Expressed Genes (DEGs) was conducted using the Gene Ontology and Kyoto Encyclopedia of Genes and Genomes database. RESULTS: Cytoscape software facilitated the identification of nine hub DEGs, namely, FN1, COL1A1, COL1A2, THBS2, COL3A1, COL5A1, APOE, SPP1, and BGN. Various network analysis algorithms were applied to determine their high connectivity. Among these hub genes, FN1, COL1A2, THBS2, COL3A1, COL5A1, and BGN were found to be associated with a poor prognosis for GC patients. Subsequent analysis using the TIMER database revealed the infiltration status of the TME concerning the overexpression of these six genes. Specifically, the abovementioned genes demonstrated direct correlations with cancer-associated fibroblasts, M1 and M2 macrophages, myeloid-derived suppressor cells, and activated dendritic cells. CONCLUSION: Our findings suggest that the identified hub genes, particularly BGN, FN1, COL1A2, THBS2, COL3A1, and COL5A1, play crucial roles in GC prognosis and TME cell infiltration. This comprehensive analysis enhances our understanding of the molecular mechanisms underlying GC development and may contribute to the identification of potential therapeutic targets and prognostic markers for GC patients.

Apoptotic Gastritis in Melanoma Patients Treated With PD-1-Based Immune Checkpoint Inhibition - Clinical and Histopathological Findings Including the Diagnostic Value of Anti-Caspase-3 Immunohistochemistry.

BACKGROUND: Gastritis induced by checkpoint inhibitors (CPI) is a rare but severe drug-related side effect. The reference standard for confirming CPI-associated gastritis (CPI-assGastritis) is histopathological assessment; however, the histopathological features of CPI-assGastritis are not yet adequately defined. MATERIALS AND METHODS: Gastric biopsies of melanoma patients with histopathologically suspected CPI-assGastritis were compared with gastric biopsies of patients with inflammation free gastric mucosa (IFGM), type A, B, and C gastritis with respect to apoptosis count and predominant histopathological features. Immunohistochemical anti-caspase-3 staining was performed to identify apoptosis. Quantification was performed by manually counting the number of apoptotic events per 10 high-power fields (HPF). Clinical symptoms, treatment, and follow-up data of patients with CPI-assGastritis were examined. The nonparametric Mann-Whitney U test was used for statistical testing. RESULTS: Five melanoma patients (three women, two men; median age: 45 years) were treated with PD-1-based CPI. The patients reported epigastric pain, weight loss, nausea, and vomiting. Histologically, the patients with CPI-assGastritis showed a partly lymphocytic, partly granulocytic inflammatory infiltrate. Manual counting of apoptotic cells in biopsy tissue slides stained against caspase 3 revealed a median of 6 apoptotic events/10 HPF (95% CI, 2.75-17.30) in the patients with CPI-assGastritis. Results for the comparison cohort (patients n = 21) were a median of 1 apoptotic event/10 HPF (95% CI, 0.5-4.5) for type-A gastritis (six patients), a median of 2 apoptotic events/10 HPF (95% CI, 0-4.5) for type-B gastritis (five patients), and no apoptosis for IFGM and type-C gastritis (five patients). Patients with CPI-assGastritis had a significantly higher apoptosis count than patients with IFGM (p<0.01), type A (p<0.05), B (p<0.05), and C gastritis (p<0.01). None of the CPI-assGastritis biopsies showed evidence of Helicobacter pylori. All CPI-assGastritis patients responded to systemic treatment with corticosteroids. CONCLUSION: CPI-assGastritis manifests with nonspecific symptoms but histologically shows a high number of apoptotic events, which can best be visualized by anti-caspase-3 immunohistochemistry. This histopathological feature may help to histologically differentiate CPI-assGastritis from other forms of gastritis and inform decision-making regarding its optimal management.

Mutations in Hepatitis D Virus Allow It to Escape Detection by CD8+ T Cells and Evolve at the Population Level.

BACKGROUND & AIMS: Hepatitis D virus (HDV) superinfection in patients with hepatitis B virus (HBV) is associated with rapid progression to liver cirrhosis and hepatocellular carcinoma. Treatment options are limited, and no vaccine is available. Although HDV-specific CD8+ T cells are thought to control the virus, little is known about which HDV epitopes are targeted by virus-specific CD8+ T cells or why these cells ultimately fail to control the infection. We aimed to define how HDV escapes the CD8+ T-cell-mediated response. METHODS: We collected plasma and DNA samples from 104 patients with chronic HDV and HBV infection at medical centers in Europe and the Middle East, sequenced HDV, typed human leukocyte antigen (HLA) class I alleles from patients, and searched for polymorphisms in HDV RNA associated with specific HLA class I alleles. We predicted epitopes in HDV that would be recognized by CD8+ T cells and corresponded with the identified virus polymorphisms in patients with resolved (n = 12) or chronic (n = 13) HDV infection. RESULTS: We identified 21 polymorphisms in HDV that were significantly associated with specific HLA class I alleles (P < .005). Five of these polymorphisms were found to correspond to epitopes in HDV that are recognized by CD8+ T cells; we confirmed that CD8+ T cells in culture targeted these HDV epitopes. HDV variant peptides were only partially cross-recognized by CD8+ T cells isolated from patients, indicating that the virus had escaped detection by these cells. These newly identified HDV epitopes were restricted by relatively infrequent HLA class I alleles, and they bound most frequently to HLA-B. In contrast, frequent HLA class I alleles were not associated with HDV sequence polymorphisms. CONCLUSIONS: We analyzed sequences of HDV RNA and HLA class I alleles that present epitope peptides to CD8+ T cells in patients with persistent HDV infection. We identified polymorphisms in the HDV proteome that associate with HLA class I alleles. Some variant peptides in epitopes from HDV were only partially recognized by CD8+ T cells isolated from patients; these could be mutations that allow HDV to escape the immune response, resulting in persistent infection. HDV escape from the immune response was associated with uncommon HLA class I alleles, indicating that HDV evolves, at the population level, to evade recognition by common HLA class I alleles.

A Mass Spectrometry-Based Profiling of Interactomes of Viral DDB1- and Cullin Ubiquitin Ligase-Binding Proteins Reveals NF-κB Inhibitory Activity of the HIV-2-Encoded Vpx.

Viruses and hosts are situated in a molecular arms race. To avoid morbidity and mortality, hosts evolved antiviral restriction factors. These restriction factors exert selection pressure on the viruses and drive viral evolution toward increasingly efficient immune antagonists. Numerous viruses exploit cellular DNA damage-binding protein 1 (DDB1)-containing Cullin RocA ubiquitin ligases (CRLs) to induce the ubiquitination and subsequent proteasomal degradation of antiviral factors expressed by their hosts. To establish a comprehensive understanding of the underlying protein interaction networks, we performed immuno-affinity precipitations for a panel of DDB1-interacting proteins derived from viruses such as mouse cytomegalovirus (MCMV, Murid herpesvirus [MuHV] 1), rat cytomegalovirus Maastricht MuHV2, rat cytomegalovirus English MuHV8, human cytomegalovirus (HCMV), hepatitis B virus (HBV), and human immunodeficiency virus (HIV). Cellular interaction partners were identified and quantified by mass spectrometry (MS) and validated by classical biochemistry. The comparative approach enabled us to separate unspecific interactions from specific binding partners and revealed remarkable differences in the strength of interaction with DDB1. Our analysis confirmed several previously described interactions like the interaction of the MCMV-encoded interferon antagonist pM27 with STAT2. We extended known interactions to paralogous proteins like the interaction of the HBV-encoded HBx with different Spindlin proteins and documented interactions for the first time, which explain functional data like the interaction of the HIV-2-encoded Vpr with Bax. Additionally, several novel interactions were identified, such as the association of the HIV-2-encoded Vpx with the transcription factor RelA (also called p65). For the latter interaction, we documented a functional relevance in antagonizing NF-κB-driven gene expression. The mutation of the DDB1 binding interface of Vpx significantly impaired NF-κB inhibition, indicating that Vpx counteracts NF-κB signaling by a DDB1- and CRL-dependent mechanism. In summary, our findings improve the understanding of how viral pathogens hijack cellular DDB1 and CRLs to ensure efficient replication despite the expression of host restriction factors.

Successful treatment of cerebral aspergillosis: case report of a patient with T-cell large granular lymphocytic leukemia (T-LGL).

BACKGROUND: Invasive aspergillosis involving patients with neutropenia or severe immunosuppression, such as patients with hematologic malignancies is associated with high mortality. Patients with T-cell large granular lymphocytic leukemia (T-LGL) on the other hand are considered to be less vulnerable for severe opportunistic fungal infection as their course of disease is chronic and marked by less violent cytopenia then in e.g. Aplastic Anemia. Only neutropenia is regarded as independent risk factor for severe opportunistic infection in T-LGL patients. CASE PRESENTATION: We report a case of a 53 year old patient with T-LGL, Immune-Thrombocytopenia (ITP) and combined antibody deficiency, who presented with fever and reduced general condition. The patient revealed a complicated infection involving the lungs and later the brain, with the presentation of vomiting and seizures. Broad microbiological testing of blood-, lung- and cerebrospinal fluid samples was inconclusive. In the absence of mycological proof, Aspergillus infection was confirmed by pathological examination of a brain specimen and finally successfully treated with liposomal amphotericin B and voriconazole, adopting a long-term treatment scheme. CONCLUSIONS: Beyond typical problems in the clinical practice involving fungal infections and hematologic malignancies, this case of invasive aspergillosis in a patient with T-LGL illustrates caveats in diagnosis, therapy and follow-up. Our data support careful ambulatory monitoring for patients with T-LGL, even in the absence of neutropenia. Especially those patients with combined hematologic malignancies and immune defects are at risk. Long-term treatment adhesion for 12 months with sufficient drug levels was necessary for sustained clearance from infection.

Immunosuppression with mTOR inhibitors prevents the development of donor-specific antibodies after liver transplant.

BACKGROUND: Donor-specific antibodies (DSAs) are an important cause of complications after solid organ transplant. Risk factors and, thus, strategies for preventing DSA development are not well defined. METHODS: The DSA status of 400 patients who underwent liver transplant (LT) at the outpatient clinic of the University Hospital Essen was determined. Human leukocyte antigen (HLA) antibodies were detected by single-antigen bead technology. The strength of DSAs was reported as mean fluorescence intensity. RESULTS: Detectable DSAs were found in 74 (18.5%) patients and significantly more often in patients who underwent LT for autoimmune liver disease than for all other indications (29.3%; P=.022), but significantly less often found in patients who underwent LT for hepatocellular carcinoma (7.6%, P=.005). The incidence of DSAs increased with time after LT, and the risk was generally higher for female patients. The frequency of DSA detection was significantly lower (10.6%) for patients receiving immunosuppressive treatment with mammalian target of rapamycin (mTOR) inhibitors than for those receiving other regimens (20.5%; P=.025). CONCLUSION: Autoimmune liver diseases, female sex, and time of more than 8 years since LT predispose patients to the development of DSAs. Immunosuppression with the mTOR inhibitor everolimus protects against DSA development after liver transplant.

"Activated" STAT proteins: a paradoxical consequence of inhibited JAK-STAT signaling in cytomegalovirus-infected cells.

We have previously characterized mouse CMV (MCMV)-encoded immune-evasive IFN signaling inhibition and identified the viral protein pM27 as inducer of proteasomal degradation of STAT2. Extending our analysis to STAT1 and STAT3, we found that MCMV infection neither destabilizes STAT1 protein nor prevents STAT1 tyrosine Y701 phosphorylation, nuclear translocation, or the capability to bind γ-activated sequence DNA-enhancer elements. Unexpectedly, the analysis of STAT3 revealed an induction of STAT3 Y705 phosphorylation by MCMV. In parallel, we found decreasing STAT3 protein amounts upon MCMV infection, although STAT3 expression normally is positive autoregulative. STAT3 phosphorylation depended on the duration of MCMV infection, the infectious dose, and MCMV gene expression but was independent of IFNAR1, IL-10, IL-6, and JAK2. Although STAT3 phosphorylation did not require MCMV immediate early 1, pM27, and late gene expression, it was restricted to MCMV-infected cells and not transmitted to bystander cells. Despite intact STAT1 Y701 phosphorylation, IFN-γ-induced target gene transcription (e.g., IRF1 and suppressor of cytokine signaling [SOCS] 1) was strongly impaired. Likewise, the induction of STAT3 target genes (e.g., SOCS3) by IL-6 was also abolished, indicating that MCMV antagonizes STAT1 and STAT3 despite the occurrence of tyrosine phosphorylation. Consistent with the lack of SOCS1 induction, STAT1 phosphorylation was prolonged upon IFN-γ treatment. We conclude that the inhibition of canonical STAT1 and STAT3 target gene expression abrogates their intrinsic negative feedback loops, leading to accumulation of phospho-tyrosine-STAT3 and prolonged STAT1 phosphorylation. These findings challenge the generalization of tyrosine-phosphorylated STATs necessarily being transcriptional active and document antagonistic effects of MCMV on STAT1/3-dependent target gene expression.