Exploration and validation of the effects of robust co-expressed immune-related genes on immune infiltration patterns and prognosis in laryngeal cancer

ObjectivesLaryngeal cancer is a common malignant tumor that originates from the larynx, yet its molecular mechanisms have not been thoroughly explored. The purpose of this study was to identify and evaluate immune-related genes in laryngeal cancer through gene co-expression networks, which may serve as biomarkers for its immunotherapy.MethodsWe applied ESTIMATE to evaluate the immune-infiltration landscape of tumor microenvironment. The co-expression networks were constructed by weighted gene co expression network analysis (WGCNA) and compared with the existing human immune related genes (IRGs) to determine the co-expressed IRGs. GSVA combined with CIBERSORT and ssGSEA illustrated the correlation of hub genes and immune infiltration patterns. TIDE algorithm and Subclass mapping evaluated the function of hub genes in predicting immune function and immunotherapeutic sensitivity. The pRRophetic was employed in the sensitivity prediction of chemotherapeutic drugs.ResultsA total of 23 co-expressed IRGs were identified and showed robust expression characteristics. These genes were significantly related to immune infiltration patterns, immune function and sensitivity prediction of immunotherapy and chemotherapeutic drugs for laryngeal cancer patients. Genetic alteration in somatic mutation level and related pathways were also revealed.ConclusionThe 23 co-expressed IRGs may act as immunotherapeutic biomarkers and potential therapeutic targets for laryngeal cancer with certain expression robustness. The molecular mechanisms deserve further investigation, which will guide clinical treatment in the future.     

Analysis of long noncoding RNA-associated competing endogenous RNA network in glucagon-like peptide-1 receptor agonist-mediated protection in β cells

BACKGROUND Long noncoding RNAs(lncRNAs) and mRNAs are widely involved in various physiological and pathological processes. The use of glucagon-like peptide-1 receptor agonists(GLP-1 RAs) is a novel therapeutic strategy that could promote insulin secretion and decrease the rate of β-cell apoptosis in type 2 diabetes mellitus(T2 DM) patients. However, the specific lncRNAs and mRNAs and their functions in these processes have not been fully identified and elucidated.AIM To identify the lncRNAs and mRNAs that are involved in the protective effect of GLP-1 RA in β cells, and their roles.METHODS Rat gene microarray was used to screen differentially expressed(DE) lncRNAs and mRNAs in β cells treated with geniposide, a GLP-1 RA. Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analyses were performed to assess the underlying functions of DE mRNAs. Hub mRNAs were filtered using the STRING database and the Cytoscape plugin, CytoHubba. In order to reveal the regulatory relationship between lncRNAs and hub mRNAs, their co-expression network was constructed based on the Pearson coefficient of DE lncRNAs and mRNAs, and competing endogenous RNA(ceRNA) mechanism was explored through miRanda and TargetScan databases.RESULTS We identified 308 DE lncRNAs and 128 DE mRNAs with a fold change filter of ≥ 1.5 and P value 0.05. GO and KEGG pathway enrichment analyses indicated that the most enriched terms were G-protein coupled receptor signaling pathway, inflammatory response, calcium signaling pathway, positive regulation of cell proliferation, and ERK1 and ERK2 cascade. Pomc, Htr2 a, and Agtr1 a were screened as hub mRNAs using the STRING database and the Cytoscape plugin, CytoHubba. This result was further verified using SwissTargetPrediction tool. Through the co-expression network and competing endogenous(ceRNA) mechanism, we identified seven lncRNAs(NONRATT027738, NONRATT027888, NONRATT030038, etc.) co-expressed with the three hub mRNAs(Pomc, Htr2 a, and Agtr1 a) based on the Pearson coefficient of the expression levels. These lncRNAs regulated hub mRNA functions by competing with six miRNAs(rno-miR-5132-3 p, rno-miR-344 g, rno-miR-3075, etc.) via the ceRNA mechanism. Further analysis indicated that lncRNA NONRATT027738 interacts with all the three hub mRNAs, suggesting that it is at a core position within the ceRNA network.CONCLUSION We have identified key lncRNAs and mRNAs, and highlighted here how they interact through the ceRNA mechanism to mediate the protective effect of GLP-1 RA in β cells.     

Evaluation of a five-gene signature associated with stromal infiltration for diffuse large B-cell lymphoma

BACKGROUNDDiffuse large B-cell lymphoma (DLBCL) is a common non-Hodgkin lymphoma. The development of immunotherapy greatly improves the patient prognosis but there are some exceptions. Thus, screening for better biomarkers for prognostic evaluation could contribute to the treatment of DLBCL patients.AIMTo screen the novel mediators involved in the development of DLBCL.METHODSThe {"type":"entrez-geo","attrs":{"text":"GSE60","term_id":"60"}}GSE60 dataset was applied to identify the differentially expressed genes (DEGs) in DLBCL, and the principal components analysis plot was used to determine the quality of the included samples. The protein-protein interactions were analyzed by the STRING tool. The key hub genes were entered into to the GEPIA database to determine their expressions in DLBCL. Furthermore, these hub gene alterations were analyzed in cBioportal. The UALCAN portal was employed to analyze the expression of the hub genes in different stages of DLBCL. The Estimation of Stromal and Immune cells in Malignant Tumor tissues using Expression data Score was conducted to evaluate the correlation between the gene expression and tumor purity. The gene-gene correlation analysis was conducted in the GEPIA. The stromal score analysis was conducted in TIMER to confirm the correlation between the gene expression and infiltrated stromal cells. The correlation between the indicated genes and infiltration level of cancer-associated fibroblasts (CAFs) was also completed in TIMER with two methods, MCP-Counter and Tumor immune dysfunction and exclusion. The correlation between fibronectin (FN1) protein level and secreted protein acidic and cysteine-rich (SPARC) messenger ribonucleic acid expression was confirmed in the cBioportal.RESULTSThe top 20 DEGs in DLBCL were identified, and the principal components analysis plot confirmed the quality of the significant DEGs. The pairwise correlation coefficient analysis among all samples showed that these DEGs have a certain co-expression pattern. The DEGs were subjected to STRING to identify the hub genes, alpha-2-macroglobulin (A2M), cathepsin B (CTSB), FN1, matrix metallopeptidase 9 (MMP9), and SPARC. The five hub genes were confirmed to be overexpressed in DLBCL. The cBioportal portal detected these five hub genes that had gene alteration, including messenger ribonucleic acid high amplification and missense mutation, and the gene alteration percentages of A2M, FN1, CTSB, MMP9, and SPARC were 5%, 8%, 5%, 2.7%, and 5%, respectively. Furthermore, the five hub genes had a potential positive correlation with tumor stage. The correlation analysis between the five genes and tumor purity confirmed that the five genes were overexpressed in DLBCL and had a positive correlation with the development of DLBCL. More interestingly, the five genes had a significant correlation with the stromal infiltration scores. The correlation analysis between the fives genes and CAFs also showed a significant value, among which the top two genes, FN1 and SPARC, had a remarkable co-expression pattern.CONCLUSIONThe top DEGs were identified, and the five hub genes were overexpressed in DLBCL. Furthermore, the gene alterations were confirmed and the positive correlation with tumor purity revealed the overexpression of the five genes and close association with the development of DLBCL. More interestingly, the five genes were positively correlated with stromal infiltration, especially in CAFs. The top two genes, FN1 and SPARC, showed a co-expression pattern, which indicates their potential as novel therapeutic targets for DLBCL.     

Cytokeratin expression in normal salivary glands and in cystadenolymphomas demonstrated by monoclonal antibodies against selective cytokeratin polypeptides

Summary The distribution of selective cytokeratin polypeptides, vimentin, and glial fibrillary protein (GFP) in 5 human cystadenolymphomas of the parotid gland was compared with normal human parotid (n=5) and submandibular (n=4) glands using a panel of monoclonal antibodies against diverse and selective cytokeratin polypeptides, vimentin and glial fibrillary protein (GFP). A biotin-streptavidin method was used on cryostat sections. The immunocytochemical finding of identical cytokeratin polypeptides Nos. 7, 8, 18 and 19 and basal cells selectively labeled by the monoclonal antibody KS 8.58, in both the epithelial part of the cystadenolymphomas and in the duct epithelium of the parotid gland, confirms the hypothesis that the epithelial compartment of cystadenolymphomas is derived from the duct system. The triple expression of cytokeratin, vimentin and GFP in myoepithelial cells of the parotid gland is discussed.     

表皮生长因子受体和环氧化酶-2在前列腺癌中的共同表达和意义

目的:探讨表皮生长因子受体(Epidermal growth factor receptor,EGFR)和环氧化酶-2(Cyclooxyge-nase-2,COX-2)在前列腺癌(Prostate cancer,PCa)中的共同表达和临床意义.方法:采用免疫组织化学法(SP)测定48例PCa中EGFR和COX-2阳性表达情况,按免疫组织化学结果分为EGFR(-)COX-2(-)、EGFR(-)COX-2( )、EGFR( )COX-2(-)和EGFR( )COX-2( )四组,分析它们表达与临床病理和预后的关系.结果:EGFR、COX-2阳性在PCa中为34例(70.8%)、21例(45.8%),两者的表达在PCa中无相关性(r＝0.04,P＝0.80),EGFR和COX-2共同表达有16例(33.3%),和Gleason评分、PSA水平有关(P<0.05),5年复发率83.1%,显著高于EGFR( )COX-2(-)(27.8%)、EGFR(-)COX 2( )(16.7%)和EGFR(-)COX-2(-)(12.5%)(P<0.05);13例转移PCa中有10例为EGFR( )COX-2( )(62.5%),显著高于其他三组(P<0.05).结论:EGFR和COX-2在PCa中表达无相关性,但其共同表达可以作为判断PCa预后的重要标志.     

Co-expressing GP5 and M proteins under different promoters in recombinant modified vaccinia virus ankara (rMVA)-based vaccine vector enhanced the humoral and cellular immune responses of porcine reproductive and respiratory syndrome virus (PRRSV)

The porcine reproductive and respiratory syndrome virus (PRRSV) has three major structural proteins which designated as GP5, M, and N. Protein GP5 and M have been considered very important to arouse the humoral and cellular immune responses against PRRSV infection and proposed to be the excellent candidate proteins in the design of PRRS bioengineering vaccine. There were some attempts on expressing GP5 or M in DNA vaccine and adenovirus to arouse humoral and cellular immune responses, but few papers have been reported on that the immune response can be difference because of the expression patterns of GP5 and M proteins in the recombinant virus. In this article, four recombinant viruses that expressed GP5 and M proteins of PRRSV in the modified vaccinia virus ankara (MVA) with different expression patterns were made. In these recombinant virus (rMVAs), GP5 and M proteins were expressed in MVA in the same virus but under the control of two promoters (rMVA-GP5/M), or as a fusion protein under one promoter (rMVA-GP5-M), or separately (rMVA-GP5 and rMVA-M). The humoral and cellular immune responses for the four recombinant viruses were evaluated with mouse model. Every mouse was inoculated with 5 × 10⁵ TCID₅₀ of the different rMVAs and boosted 3 weeks later. Neutralizing antibody titers for each group were detected with virus neutralization test assay weekly after the primary inoculation for 13 weeks to evaluate the humoral immune response. The production of gamma interferon (IFN-γ), interleukin-2 (IL-2), and interleukin-4 (IL-4) was detected in splenocytes of rMVA-inoculated mice at 30, 60, and 90 days post inoculation to evaluate the cellular immune response. Results showed that rMVA-GP5 and rMVA-M cannot induce obvious humoral and cellular immune responses; rMVA-GP5-M inoculated group developed better immune responses than rMVA-GP5 and rMVA-M inoculated groups; however, mice inoculated with rMVA-GP5/M maintained the strongest cellular response against PRRS and consistently enhanced the anti-PRRSV humoral responses. The strategy of co-expressing PRRSV GP5 and M protein in MVA under the control of different promoters might be an attractive method for future PRRSV vaccine design. Co-first authors have same contribution for this article.     

Amygdala-kindled seizures increase the expression of corticotropin-releasing factor (CRF) and CRF-binding protein in GABAergic interneurons of the dentate hilus

Kindling, a model of temporal lobe epilepsy, induces a number of neuropeptides including corticotropin-releasing factor (CRF). CRF itself can produce limbic seizures which resemble kindling in some aspects. However, tolerance to the convulsant effects of CRF develops rapidly. Hypothetically, this could be explained should seizures also induce the CRF-binding protein (CRF-BP), which has been postulated to restrict the actions of CRF. Therefore, in the present study, we used in situ hybridization to examine the effects of amygdala-kindled seizures on the mRNA levels of CRF and CRF-BP. Kindled seizures markedly elevated CRF and CRF-BP in the dentate gyrus of rats. CRF and CRF-BP were induced almost exclusively in GABAergic interneurons of the dentate hilus. The CRF and CRF-BP interneurons also expressed neuropeptide Y but not cholecystokinin. CRF appeared to have an excitatory role in the dentate gyrus as it decreased the afterhyperpolarization of dentate granule neurons. These results suggest that CRF may contribute to the development of amygdala kindling. However, the compensatory induction of CRF-BP may serve to limit the excitatory effects of CRF in the dentate gyrus.     

Epstein-Barr Virus-encoded Latent Membrane Protein 1 Co-expresses with Epidermal Growth Factor Receptor in Nasopharyngeal Carcinoma

Latent membrane protein 1 (LMP-1) is the only Epstein-Barr virus (EBV)-encoded oncogenic protein that has been detected in nasopharyngeal carcinoma (NPC), a cancer that is closely associated with EBV. Previous in-vitro studies have demonstrated that LMP-1 can upregulate epidermal growth factor receptor (EGFR) in epithelial cells. It was not established whether this cellular effect exists in NPC. To assess the association between LMP-1 and EGFR in NPC tissues, 60 NPC specimens were examined by immunohistochemistry using anti-LMP-1 antibody (CS 1–4) and anti-EGFR antibodies (EGFR 1, EGFR 1005). The results revealed that 41 (68.3%) specimens were immunopositive for LMP-1 and 44 (73.3%) specimens over-expressed EGFR. Morphologically, the expressions of LMP-1 and EGFR were homogeneously distributed in the tumor nests. In addition, the correlation between LMP-1 and EGFR was statistically significant (P<0.001, χ² test, d.f.=1). To elucidate further the correlation between LMP-1 and EGFR in vivo and in situ , an indirect dual immunofluorescence assay was conducted, using secondary antibodies conjugated with fluorescein isothiocyanate (FITC) or indocarbocyanine (Cy3). The results disclosed an intimate co-expression of LMP-1 and EGFR. In summary, the data indicate that over-expression of EGFR is a common phenomenon in NPC, and that EGFR is co-expressed with LMP-1 in NPC. Thus, EBV may play a role in the tumorigenesis of NPC through the effects of LMP-1 and EGFR.     

Meta-analysis of Gene Expression Data Identifies Causal Genes for Prostate Cancer

Prostate cancer is a leading cause of death in male populations across the globe. With the advent of geneexpression arrays, many microarray studies have been conducted in prostate cancer, but the results have variedacross different studies. To better understand the genetic and biologic mechanisms of prostate cancer, we conducteda meta-analysis of two studies on prostate cancer. Eight key genes were identified to be differentially expressedwith progression. After gene co-expression analysis based on data from the GEO database, we obtained a coexpressedgene list which included 725 genes. Gene Ontology analysis revealed that these genes are involvedin actin filament-based processes, locomotion and cell morphogenesis. Further analysis of the gene list shouldprovide important clues for developing new prognostic markers and therapeutic targets.     

Analysis of Indoleamine 2-3 Dioxygenase (IDO) and EGFR Coexpression in Breast Cancer Tissue by Immunohistochemistry

Background: To determine the amount of co-expression of IDO and EGFR in breast cancer patients.Materials and Methods:In order to obtain the distribution of co-expression of IDO and EGFR in breast cancer,we tested 110 breast cancer paraffin tissue blocks with immunohistochemical methods. Then we investigated therelationship between the diagnostic and pathologic characteristics (tumor size, lymph node status, histologicgrade, the gene expression of ER, PR, HER2, p53, Ki67 and PCNA) with the situation of co-expression of IDOand EGFR by reviewing the medical records of 32 breast cancer patients. Results: Among 110 breast cancers, 32cases demonstrated IDO and EGFR co-expression (29.1%), IDO and EGFR synchronous co-expression beingfound in 19.1% and asynchronous in 10.0%. Conclusions: IDO and EGFR were co-expressed in breast cancer,including synchronous and asynchronous co-expression. The results suggest that considering IDO and EGFRas two indicators for breast cancer treatment or prognosis analysis provides a potential option of individualtreatment for the portion of breast cancer patients with co-expression of IDO and EGFR.