Receptor Interaction Pathway Research Articles

Genome-wide transcriptome architecture in a mouse model of Gulf War Illness

Gulf War Illness (GWI) is thought to be a chronic neuroimmune disorder caused by in-theater exposure during the 1990–1991 Gulf War. There is a consensus that the illness is caused by exposure to insecticides and nerve agent toxicants. However, the heterogeneity in both development of disease and clinical outcomes strongly suggests a genetic contribution. Here, we modeled GWI in 30 BXD recombinant inbred mouse strains with a combined treatment of corticosterone (CORT) and diisopropyl fluorophosphate (DFP). We quantified transcriptomes from 409 prefrontal cortex samples. Compared to the untreated and DFP treated controls, the combined treatment significantly activated pathways such as cytokine-cytokine receptor interaction and TNF signaling pathway. Protein-protein interaction analysis defined 6 subnetworks for CORT + DFP, with the key regulators being Cxcl1, Il6, Ccnb1, Tnf, Agt, and Itgam. We also identified 21 differentially expressed genes having significant QTLs related to CORT + DFP, but without evidence for untreated and DFP treated controls, suggesting regions of the genome specifically involved in the response to CORT + DFP. We identified Adamts9 as a potential contributor to response to CORT + DFP and found links to symptoms of GWI. Furthermore, we observed a significant effect of CORT + DFP treatment on the relative proportion of myelinating oligodendrocytes, with a QTL on Chromosome 5. We highlight three candidates, Magi2, Sema3c, and Gnai1, based on their high expression in the brain and oligodendrocyte. In summary, our results show significant genetic effects of the CORT + DFP treatment, which mirrors gene and protein expression changes seen in GWI sufferers, providing insight into the disease and a testbed for future interventions.

High-Throughput MicroRNA Profiling of Vitreoretinal Lymphoma: Vitreous and Serum MicroRNA Profiles Distinct from Uveitis.

Purpose: Vitreoretinal lymphoma (VRL) is a non-Hodgkin lymphoma of the diffuse large B cell type (DLBCL), which is an aggressive cancer causing central nervous system related mortality. The pathogenesis of VRL is largely unknown. The role of microRNAs (miRNAs) has recently acquired remarkable importance in the pathogenesis of many diseases including cancers. Furthermore, miRNAs have shown promise as diagnostic and prognostic markers of cancers. In this study, we aimed to identify differentially expressed miRNAs and pathways in the vitreous and serum of patients with VRL and to investigate the pathogenesis of the disease. Materials and Methods: Vitreous and serum samples were obtained from 14 patients with VRL and from controls comprising 40 patients with uveitis, 12 with macular hole, 14 with epiretinal membrane, 12 healthy individuals. The expression levels of 2565 miRNAs in serum and vitreous samples were analyzed. Results: Expression of the miRNAs correlated significantly with the extracellular matrix (ECM) ‒receptor interaction pathway in VRL. Analyses showed that miR-326 was a key driver of B-cell proliferation, and miR-6513-3p could discriminate VRL from uveitis. MiR-1236-3p correlated with vitreous interleukin (IL)-10 concentrations. Machine learning analysis identified miR-361-3p expression as a discriminator between VRL and uveitis. Conclusions: Our findings demonstrate that aberrant microRNA expression in VRL may affect the expression of genes in a variety of cancer-related pathways. The altered serum miRNAs may discriminate VRL from uveitis, and serum miR-6513-3p has the potential to serve as an auxiliary tool for the diagnosis of VRL.

Time- and Dose-Resolved Proteome of PM2.5-Exposure-Induced Lung Injury and Repair in Rats.

In recent years, airborne fine particulate matter (PM2.5) is drawing more public attention due to its various physicochemical features and causing pathological harm, as proven by epidemiological and clinical studies. However, the mechanism of PM2.5-exposure-induced lung injury has not been fully characterized. Here, we established a PM2.5-induced rat injury model for both short-term and long-term exposures at different concentrations. We employed the Fast-seq technique to profile 6316 proteins and the catTFRE approach to profile 387 transcription factors (TFs) in the lung tissue. In short-term exposure, we elucidated gradually upregulated proteins enriched in response to oxidative stress, phagosome, and the extracellular matrix (ECM)-receptor interaction pathway. Long-term exposure mainly showed the immune response pathway to be consisting of increased lymphocytes and cytokines. Intriguingly, we found that immune-related proteins were recoverable during short-term exposure. During the process of PM2.5 exposure, upregulated proteins presented dose-dependence in the lung, including stress response at low dose, minor immune response at middle dose, and severe inflammatory response at high dose. This data set provides a rich resource to facilitate the understanding of PM2.5-induced lung damage and repair mechanism.

Identification of methylated-differentially expressed genes and pathways in esophageal squamous cell carcinoma

Methylation, as an epigenetic modification, can affect gene expression and play a role in the occurrence and development of cancer. This research is devoted to discover methylated-differentially expressed genes (MDEGs) in esophageal squamous cell carcinoma (ESCC) and explore special associated pathways. We downloaded GSE51287 methylation profiles and GSE26886 expression profiles from GEO DataSets, and performed a comprehensive bioinformatics analysis. Totally, 19 hypermethylated, lowly expressed genes (Hyper-LGs) were identified, and involved in regulation of cell proliferation, phosphorus metabolic process and protein kinase activity. Meanwhile, 17 hypomethylated, highly expressed genes (Hypo-HGs) were participated in collagen catabolic process, metallopeptidase and cytokine activity. Pathway analysis determined that Hyper-LGs were enriched in arachidonic acid metabolism pathway, while Hypo-HGs were primarily associated with the cytokine-cytokine receptor interaction pathway. IL 6, MMP3, MMP9, SPP1 were identified as hub genes based on the PPI network that combined 7 ranked methods included in cytoHubba, and verification was performed in human tissues. Our integrated analysis identified many novel genetic lesions in ESCC and provides a crucial molecular foundation to improve our understanding of ESCC. Hub genes, including IL 6, MMP3, MMP9 and SPP1, could be considered for use as aberrant methylation-based biomarkers to facilitate the accurate diagnosis and therapy of ESCC.

Transcriptome profiling of digital flexor tendons after injury in a chicken model.

Background: Modulation of tendon healing remains a challenge because of our limited understanding of the tendon repair process. Therefore, we performed the present study to provide a global perspective of the gene expression profiles of tendons after injury and identify the molecular signals driving the tendon repair process.Results: The gene expression profiles of flexor digitorum profundus tendons in a chicken model were assayed on day 3, weeks 1, 2, 4, and 6 after injury using the Affymetrix microarray system. Principal component analysis (PCA) and hierarchical cluster analysis of the differentially expressed genes showed three distinct clusters corresponding to different phases of the tendon healing period. Gene ontology (GO) analysis identified regulation of cell proliferation and cell adhesion as the most enriched biological processes. Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis revealed that the cytokine–cytokine receptor interaction and extracellular matrix (ECM)–receptor interaction pathways were the most impacted. Weighted gene co-expression network analysis (WGCNA) demonstrated four distinct patterns of gene expressions during tendon healing. Cell adhesion and ECM activities were mainly associated with genes with drastic increase in expression 6 weeks after injury. The protein–protein interaction (PPI) networks were constructed to identify the key signaling pathways and hub genes involved.Conclusions: The comprehensive analysis of the biological functions and interactions of the genes differentially expressed during tendon healing provides a valuable resource to understand the molecular mechanisms underlying tendon healing and to predict regulatory targets for the genetic engineering of tendon repair.Tendon healing, Adhesion, Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Weighted Gene Co-expression Network Analysis, Protein–protein Interaction

The reproductive toxicity and potential mechanisms of combined exposure to dibutyl phthalate and diisobutyl phthalate in male zebrafish (Danio rerio)

Dibutyl phthalate (DBP) and diisobutyl phthalate (DiBP) are phthalate compounds frequently detected in the environment. Despite increasing awareness of their toxicity in human and animals, the male reproductive toxicity of their combined exposure remains elusive. The purposes of this study were to investigate whether combined exposure to DBP and DiBP could induce male reproductive toxicity, and to explore the potential toxicological mechanisms. Adult male zebrafish were exposed to DBP (11, 113 and 1133 μg L−1), DiBP (10, 103 and 1038 μg L−1) and their mixtures (Mix) (11 + 10, 113 + 103, 1133 + 1038 μg L−1) for 30 days, and their effects on plasma hormone secretion, testis histology and transcriptomics were examined. Highest concentrations of Mix exposure caused greater imbalance ratio of T/E2 and more severe structural damage to testis than single exposure. These effects were consistent with the testis transcriptome analysis for which 4570 genes were differentially expressed in Mix exposure, while 2795 and 1613 genes were differentially expressed in DBP and DiBP, respectively. KEGG pathway analysis showed that both single and combined exposure of DBP and DiBP could affect cytokine-cytokine receptor interaction. The difference was that combined exposure could also affect steroid hormone synthesis, extracellular matrix receptor interaction, retinol metabolism, and PPAR signaling pathways. These results demonstrated that combined exposure to DBP and DiBP could disrupt spermatogenesis and elicit male reproductive toxicity in zebrafish.

Upregulated C-C Motif Chemokine Ligand 2 Promotes Ischemic Stroke via Chemokine Signaling Pathway

This study aims to evaluate the potential effect and the underlying mechanism of C-C motif chemokine ligand 2 (CCL2) in ischemic stroke. An integrated bioinformatics analysis was performed to identify the differentially expressed (DE) genes and their related pathways in ischemic stroke. Invivo study of a rat model of middle cerebral artery occlusion (MCAO) was further established to assess the effect of CCL2 on severity of neurologic impairments. The expression levels of proinflammatory cytokines were also evaluated using the ELISA assay, and Western blot was also used to determine the expression of CCL2 and other DE proteins in the related pathways. A total of 88 DE genes were identified from the microarray dataset of ischemic stroke. The bioinformatics analysis revealed that CCL2 was highly expressed in ischemic stroke tissue and promoted the ischemic stroke progression via activation of the chemokine signaling pathway and cytokine-cytokine receptor interaction pathway. The invivo study of the ischemic stroke rat model also showed that the CCL2 expression was elevated in the MCAO/R rats, with significant neurological impairments and ischemic infarct area in the brain tissue being observed. The administration of CCL2 inhibitors significantly inhibited the inflammatory response, attenuated the neurological impairments, and decreased the ischemic infarct area in the MCAO/R rats. Furthermore, the downregulation of CCL2 also inhibited the expression of the pathway-related proteins including CCL7, CCR2, CXCL16, and TNF-α. These results indicate that the CCL2/chemokine signaling pathway is responsible for ischemic stroke progression and might represent a potential therapeutic target for ischemic stroke treatment.

MRNA and microRNA Profiles in the Amygdala Are Relevant to Susceptibility and Resilience to Psychological Stress Induced in Mice.

Severe or prolonged stress increases the risk for developing psychopathological disorders. An individual's perception of stress exposure varies greatly, as do its consequences. Numerous individuals demonstrate resilience to psychological stress. The mRNA and microRNA profiles of stress susceptibility and resilience to induced psychological stress in the amygdala remain to be elucidated. In this work, psychological stress was induced in an observer mouse by witnessing a similar individual under attack by an aggressor. After 5 days of psychological stress, the degree of fear memory and anxiety in mice was measured by a social interaction test and elevated plus-maze (EPM) test. mRNA and microRNA profiles were quantified by high-throughput sequencing in amygdala tissue harvested from Control, Susceptible and Resilient mice. In the amygdala of Susceptible versus Resilient mice, the upregulation of peptide, thyrotropin-releasing hormone, ECM receptors, glutamatergic synapse, cytokine-cytokine receptor interaction, long-term depression, PI3K-Akt, oxytocin, GnRH, HIF-1, estrogen, and calcium signaling pathways may be related to psychological stress-induced susceptibility, and their downregulation may be related to resilience. The downregulation of adrenergic synapse, adherens junction, Wnt, sphingolipid, B cell receptor, cAMP, Rap1, and Toll-like receptor signaling pathways may be related to psychological stress-induced susceptibility, and the upregulation may be related to resilience. Results by sequencing of mRNA and microRNA profiles are consistent, in which some are validated by qRT-PCR and dual-luciferase reporter assay. Susceptibility and resilience induced by psychological stresses are caused by the imbalanced regulation of different synapses and signaling pathways in the amygdala.

Transcriptome analysis reveals seven key immune pathways of Japanese flounder (Paralichthys olivaceus) involved in megalocytivirus infection

Megalocytivirus is a serious viral pathogen to many farmed fish including Japanese flounder (Paralichthys olivaceus). In this study, in order to systematically identify host immune genes induced by megalocytivirus infection, we examined the transcription profiles of flounder infected by megalocytivirus for 2, 6, and 8 days. Compared with uninfected fish, virus-infected fish exhibited 1242 differentially expressed genes (DEGs), with 225, 275, and 877 DEGs occurring at 2, 6, and 8 days post infection, respectively. Of these DEGs, 728 were upregulated and 659 were downregulated. The majority of DEGs were time-specific and formed four distinct expression profiles well correlated with the time of infection. The DEGs were classified into diverse Gene Ontology (GO) functional terms and enriched in 27 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, approximately one third of which were related to immunity. Weighted co-expression network analysis (WGCNA) was used to identify 16 key immune DEGs belonging to seven immune pathways (RIG-I-like receptor signaling pathway, JAK-STAT signaling pathway, TLR signaling pathway, cytokine-cytokine receptor interaction, phagosome, apoptosis, and p53 signaling pathway). These pathways interacted extensively and formed complicated networks. This study provided a global picture of megalocytivirus-induced gene expression profiles of flounder at the transcriptome level and uncovered a set of key immune genes and pathways closely linked to megalocytivirus infection. These results provided a set of targets for future delineation of the key factors implicated in the anti-megalocytivirus immunity of flounder.

Comparative Transcriptome Analysis Suggests Key Roles for 5-Hydroxytryptamlne Receptors in Control of Goose Egg Production.

To date, research on poultry egg production performance has only been conducted within inter or intra-breed groups, while those combining both inter- and intra-breed groups are lacking. Egg production performance is known to differ markedly between Sichuan white goose (Anser cygnoides) and Landes goose (Anser anser). In order to understand the mechanism of egg production performance in geese, we undertook this study. Here, 18 ovarian stromal samples from both Sichuan white goose and Landes goose at the age of 145 days (3 individuals before egg production initiation for each breed) and 730 days (3 high- and low egg production individuals during non-laying periods for each breed) were collected to reveal the genome-wide expression profiles of ovarian mRNAs and lncRNAs between these two geese breeds at different physiological stages. Briefly, 58, 347, 797, 777, and 881 differentially expressed genes (DEGs) and 56, 24, 154, 105, and 224 differentially expressed long non-coding RNAs (DElncRNAs) were found in LLD vs. HLD (low egg production Landes goose vs. high egg production Landes goose), LSC vs. HSC (low egg production Sichuan White goose vs. high egg production Sichuan white goose), YLD vs. YSC (young Landes goose vs. young Sichuan white goose), HLD vs. HSC (high egg production Landes goose vs. high egg production Sichuan white goose), and LLD vs. LSC (low egg production Landes goose vs. low egg production Sichuan white goose) groups, respectively. Functional enrichment analysis of these DEGs and DElncRNAs suggest that the “neuroactive ligand–receptor interaction pathway” is crucial for egg production, and particularly, members of the 5-hydroxytryptamine receptor (HTR) family affect egg production by regulating ovarian metabolic function. Furthermore, the big differences in the secondary structures among HTR1F and HTR1B, HTR2B, and HTR7 may lead to their different expression patterns in goose ovaries of both inter- and intra-breed groups. These results provide novel insights into the mechanisms regulating poultry egg production performance.

Abnormal Peripheral Neutrophil Transcriptome in Newly Diagnosed Type 2 Diabetes Patients.

Aim There are increasing evidence demonstrating that neutrophil-mediated inflammation plays a role in the etiology of type 2 diabetes. However, the molecular mechanisms by which neutrophils contribute to type 2 diabetes remain largely unknown. The aim of the present work was to identify possible changes in circulating neutrophils to better elucidate neutrophil involvement in human type 2 diabetes. Methods Patients newly diagnosed with type 2 diabetes (n = 5) and age- and sex-matched healthy controls (n = 5) were recruited. Neutrophils were purified from type 2 diabetes patients and controls, and RNA sequencing (RNA-seq) was used for comprehensive transcriptome analysis. Differentially expressed genes (DEGs) were screened, and Gene Ontology (GO) and KEGG pathway enrichment analyses were performed. Real-time polymerase chain reaction (qPCR) was used for validation in external samples of type 2 diabetes patients (n = 8) and healthy controls (n = 8). Results Gene expression analysis showed that, compared with neutrophils from healthy controls, there were 1990 upregulated DEGs and 1314 downregulated DEGs in neutrophils from type 2 diabetes patients. GO analysis demonstrated that the DEGs were mainly involved in myeloid leukocyte activation, T cell activation, adaptive immunity, and cytokine production. The top 20 enriched KEGG pathways included the cytokine-cytokine receptor interaction pathway, NF-κB signaling pathway, cell adhesion molecules, and chemokine signaling pathway. Furthermore, qPCR of genes related to neutrophil activation revealed that the expression of SELL, SELP, CXCR1, and S100A8 was significantly increased in neutrophils from type 2 diabetes patients compared with that in neutrophils from controls. Conclusions Our study reveals an abnormal activation of circulating neutrophils at the transcriptome level in type 2 diabetes patients. These findings suggest a potential involvement of neutrophil dysfunction in the pathologic process of type 2 diabetes and provide insight into potential therapeutic targets for type 2 diabetes.

PGN and LTA from Staphylococcus aureus Induced Inflammation and Decreased Lactation through Regulating DNA Methylation and Histone H3 Acetylation in Bovine Mammary Epithelial Cells.

Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) are the most common pathogens of mastitis, and S. aureus generally causes subclinical mastitis which is more persistent and resistant to treatment. Peptidoglycan (PGN) and lipoteichoic acid (LTA) are cell wall components of S. aureus. Although the roles of PGN and LTA in causing inflammation are well studied, the epigenetic mechanisms of the effects of PGN and LTA on the inflammation and lactation remain poorly understood. This study characterized the gene expression profiling by RNA sequencing and investigated DNA methylation and histone acetylation in relation to inflammation and lactation in the immortalized bovine mammary epithelial cell line (MAC-T). The cells were cultured for 24 h with neither PGN nor LTA (CON), PGN (30 μg/mL), LTA (30 μg/mL), and PGN (30 μg/mL) + LTA (30 μg/mL), respectively. The number of differentially expressed genes (DEGs) and the expression of proinflammatory factors including interleukin (IL)-1β, IL-6, IL-8, chemokine (C-X-C motif) ligand (CXCL)1, and CXCL6 of the treatments increased in the following order: CON < PGN < LTA < PGN + LTA, and the DEGs mainly enriched on the cytokine-cytokine receptor interaction and chemokine signaling pathway. LTA and PGN + LTA induced hypomethylation of global DNA by suppressing DNA methyltransferase (DNMT) activity. PGN and LTA, alone or combined, decreased the mRNA expression of casein genes (CSN1S1, CSN2, and CSN3) and the expression of two caseins (CSN2 and CSN3), and reduced histone H3 acetylation by suppressing histone acetyltransferase (HAT) activity and promoting histone deacetylase (HDAC) activity. Collectively, this study revealed that PGN and LTA induced inflammation probably due to decreasing DNA methylation through regulating DNMT activity, and decreased lactation possibly through reducing histone H3 acetylation by regulating HAT and HDAC activity in bovine mammary epithelial cells.

ROLE OF TYROSYLPROTEIN SULFOTRANSFERASE 2 (TPST2) IN PANCREATIC CANCER

Pancreatic cancer is an aggressive disease which has become the third leading cause for cancer deaths in the United States. Tyrosylprotein sulfotransferase 2 (TPST2) is a post‐translational modification enzyme which locates in the trans‐Golgi apparatus and catalyzes the sulfation of proteins’ tyrosine residues. Moreover, tyrosine sulfation has been shown to play a critical role in protein‐protein interaction. By analyzing TCGA data, we found that the expression of several known substrates of TPST2 were affected in pancreatic cancer. However, whether TPST2 affects cancer cell behavior is unclear. To determine the role of TPST2 in pancreatic cancer, MTT cell growth assay and soft agar colony formation assay were performed to assess cancer cell proliferation in MiaPACA2 and PANC‐1 human pancreatic cancer cell lines. Migration and invasion of cancer cells in vitro were evaluated by transwell assay. CRISPR‐Cas9 was utilized to establish TPST2 KO stable cell lines. Total RNA‐seq analysis was performed to analyze the transcriptome in the TPST2 KO and control cells. We showed that knockdown of TPST2 inhibited the proliferation of pancreatic cancer cells both in vitro and in vivo. Knockdown of TPST2 also decreased migration and invasion of pancreatic cancer cells in vitro. RNA‐seq analysis revealed that the extracellular matrix (ECM) receptor interaction pathway was significantly downregulated in TPST2 KO cells. We further demonstrated that expression of intergrin beta 4 (ITGB4), a protein involved in ECM receptor interaction pathway, was decreased when TPST2 was knocked‐down or inhibited. Our studies have uncovered the role of protein sulfation in pancreatic cancer and may establish TPST2 as a therapeutic target for pancreatic cancer.Support or Funding InformationThis work was supported in part by National Institutes of Health grants DK083952, ES023438 to Wen Xie.

APEC infection affects cytokine–cytokine receptor interaction and cell cycle pathways in chicken trachea

Avian pathogenic Escherichia coli (APEC) can lead to extraintestinal disease in avian species via respiratory tract infection. However, the regulatory mechanism of APEC on the pathogenicity of chicken trachea epithelium remains unknown. In this study, we examined pathological changes in chicken trachea at different infection times (4, 8, 12 and 24 h). The RNA sequencing of APEC infection group and the PBS group (negative control) of chicken trachea epithelium were analysed. Our studies revealed that the oedema, heterophil infiltration and hyperaemia appeared at 8 and 12 h post APEC infection. And the hyperaemia phenomenon and heterophilic granulocyte infiltration disappeared at 24 h post infection. Then RNA sequencing showed many genes were dynamically expressed in the APEC infection group. At 4, 8 and 12 h post infection, the mRNA of differentially expressed genes were enriched by cytokine–cytokine receptor interaction and the toll-like receptor signalling pathway. The cell cycle pathway was enriched at 24 h post infection. Altogether, these findings suggest that APEC infection induces pathological change in the chicken trachea, the mRNA of differentially expressed genes participating in inflammation and hyperplasia signalling pathways. Which not only provide more evidence for regulatory mechanism of APEC on the pathogenicity of chicken trachea epithelium, but also facilitate the effective management of APEC infections in poultry through trachea.

Application of NanoString technologies in angioimmunoblastic T cell lymphoma.

Angioimmunoblastic T-cell lymphoma (AITL) is an aggressive disease. Most cancer diagnoses are determined by anatomical histology. Therefore, many samples are stored in FFPE blocks for H&E staining. However, RNAs extracted from the FFPE block have a high level of fragmentation, making it difficult to perform accurate DEG analysis using RNA sequencing. To overcome fragmented RNA's drawback in NGS application, we applied the NanoString nCounter® technique of hybridization method that can be used for DEG analysis without PCR amplification. We characterized the gene expression profiling of AITLs though transcriptome analysis based on the nCounter® PanCancer IO 360™ Panel and NanoString platform. To perform the analysis of differential expression gene (DEG) profiles in AITLs, we compared the NanoString data from eight AITL patients with a healthy control donor. Ninety-one genes were up-regulated and six genes were down-regulated in AITLs compared to control. The Gene Ontology (GO) analysis of 97-DEGs revealed that they were closely related to cytokine, MAPK cascade, leukocyte differentiation, and immune response, suggesting that this affect the immune system. In addition, KEGG analysis revealed that AITL DEGs were found to be highly involved in cytokine-cytokine receptor interaction and PI3K-Akt signaling pathway. We believe that comprehensive multiplex studies, along with NanoString analysis, may be helpful to understand the molecular mechanisms of AITL, including mutations, gene expression, and protein expression studies.

Oncogenic Network and Hub Genes for Natural Killer/T-Cell Lymphoma Utilizing WGCNA.

Natural killer (NK)/T-cell lymphoma (NKTCL) is a subtype of non-Hodgkin lymphoma with aggressive progression and poor prognosis. The molecular mechanisms of NKTCL have not been well-studied. Herein, we revealed the lymphoma-associated dysregulated genes and signaling pathways or biological processes in NKTCL. We characterized that the extracellular matrix (ECM) receptor interaction pathway and T-cell receptor signaling pathway were the main dysregulated pathways in NKTCL by Gene Ontology (GO) analysis and pathway enrichment analysis. By using weighted gene co-expression network analysis (WGCNA), the gene co-expression network of NKTCL (SRP049695) was constructed, and hub genes (LMO3, GRB14) were identified. In addition, another Gene Expression Omnibus (GEO) dataset (GSE69406) was used to validate these hub genes. Furthermore, these hub genes were identified and validated by survival analysis (GSE90597). These results provided novel insights into the pathogenesis of NKTCL. Of particular interest, LMO3 and GRB14 might be potential oncoproteins and biomarkers for the diagnosis and treatment of NKTCL.

Indirectly stimulation of DCs by Ganoderma atrum polysaccharide in intestinal-like Caco-2/DCs co-culture model based on RNA-seq

Abstract Our previous study has demonstrated that Ganoderma atrum polysaccharide (PSG-1) activated the immune response of dendritic cells (DCs) by direct contacting with surface receptors. However, as a kind of macromolecule compound, PSG-1 cannot directly stimulate DCs via passing through the intestine epithelial cells. To investigate the role of PSG-1 in the interaction between the intestinal epithelium and its internal immune cells, the intestinal-like Caco-2/DCs co-culture model was established, then all transcripts of DCs with or without treatment of PSG-1 were globally screened by RNA-seq. According to the RNA-seq analysis, cytokine receptor interaction, NF-κB and MAPK signaling pathway were three important signal pathways annotate to the environmental information processing. Moreover, cytokine and related proteins expression were in DCs indirectly stimulated by PSG-1 were raised. These above findings indicated that PSG-1 indirectly affected the immune function of DCs in intestinal-like Caco-2/DCs co-culture model, in which NF-κB and MAPK pathway played essential roles.

Time-Dependent Response of Human Deciduous Tooth-Derived Dental Pulp Cells Treated with TheraCal LC: Functional Analysis of Gene Interactions Compared to MTA.

Pulp capping material should facilitate hard tissue regeneration on the injured pulp tissue. TheraCal LC (TC) was recently developed. Although TC has shown reliable clinical outcomes after direct pulp capping, there are still remaining concerns regarding its detrimental effect on pulp cells. Therefore, this study aimed to identify the gene expression of human deciduous tooth-derived dental pulp cells exposed to TC compared to mineral trioxide aggregate (MTA). The cells were cultured and exposed to TC and MTA for 24 and 72 h. Next, total RNA was isolated. QuantSeq 3′ mRNA-sequencing was used to examine differentially expressed genes (DEGs) in exposed to TC and MTA. Functional analysis of DEGs was performed using bioinformatics analysis. In gene ontology (GO) functional enrichment analysis, cells in TC for 24 h presented significantly enriched immune response (p < 0.001) and inflammatory response (p < 0.01) compared to MTA. TC showed enriched positive regulation of cell migration at 72 h (p < 0.001). In Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, neuroactive ligand–receptor interaction (p = 1.19 × 10−7) and calcium signaling pathway (p = 2.96 × 10−5) were confirmed in the shared DEGs in TC. In conclusion, DEGs in TC may be involved in pathways associated with osteoclastogenesis and osteoclastic differentiation.

Abstract P5-13-05: Gene expression changes with neoadjuvant hormonal treatment

Abstract Breast cancers (BC) expressing estrogen and/or progesterone receptors, referred to as hormone receptor (HR)-positive represent the largest molecular subset of breast tumors. Fortunately, estrogen receptor expression on the cell surface serves as a drug target for endocrine therapy. In HR+, human epidermal growth factor receptor-2 (HER2) negative BC, neoadjuvant endocrine therapy (NET) is being increasingly used. While NET has been shown to reduce tumor cell proliferation causing apoptosis, up to 20% of tumors do not respond. Currently there are no validated predictive gene signatures distinguishing responders from non-responders to NET. Furthermore, there are limited data on the effects of different NET regimens in vivo and how they vary between responders and non-responders. Methods: We conducted a retrospective translational study in patients who were treated with NET at our institution. This study was approved by our institutional review board. Paired pre-treatment and surgical samples were collected from 24 patients along with clinico-pathological information. Responders were defined as those with a preoperative endocrine prognostic index (PEPI) score of 0. RNA isolation was performed using the Roche HighPure FFPET RNA Isolation spin-column kit. Purified RNA was hybridized with the PanCancer Code Set and finally the cartridge was scanned on the nCounter Digital Analyzer. Raw counts from each gene were imported into the nSolver Analysis. Log2 normalized data obtained from Nanostring nCounter analysis was analyzed for differentially expressed genes between pre- and post-treatment samples. The bioinformatics analysis was carried out using nCounter analysis software. Results: The median age at diagnosis was 64 years and NET used was anastrozole alone or in combination with fulvestrant. At baseline the majority of tumors were invasive ductal carcinomas (n=18), grade II (n=18), with a Ki67% &amp;gt;15% (n=16). Median duration of treatment with NET was 5 months (range 2-10 months). Nine tumors were defined as responders to NET and 15 were non-responders. In all of the 24 paired samples, we found 123 genes with a statistically significant change in expression (defined by greater than two-fold change and a false discovery rate adjusted p-value &amp;lt;0.05) in the surgical specimen compared to pre-treatment biopsy. Genes in the Janus kinase-signal transducers and activators of transcription (Jak-STAT) signaling pathway, mitogen-activated protein kinase (MAPK) signaling pathway, RAS signaling, cytosine-cytosine receptor interaction and the Phosphatidylinositol-3-kinase/Akt/mammalian target of rapamycin (PI3K-Akt) signaling pathway were found to be upregulated whereas genes in cell cycle pathway were downregulated in the surgical specimens after NET. Comparing pre-treatment samples of responders to non-responders 25 genes involving PI3K-Akt, Notch signaling and Fanconi anemia pathway were differentially expressed. Conclusions: In our retrospective translational study we found that NET upregulates genes in the Jak-STAT, MAPK, RAS, cytosine-cytosine receptor interaction and PI3K-Akt signaling pathways and downregulates cell cycle pathway genes independent of type or duration of NET. We also found multiple genes involving the PI3K-Akt and Notch pathway were differentially expressed in pre-treatment specimens among responders compared to non-responders. While the sample size comparing responders versus non-responders was inadequate, our results suggest some early trends which warrant further investigation. Citation Format: Darien E Reed, Ritu Pandey, Jennifer M Segar, Michele Chu, Lauren LeBeau, Pavani Chalasani. Gene expression changes with neoadjuvant hormonal treatment [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P5-13-05.

Identification of an immune-related risk signature for predicting prognosis in clear cell renal cell carcinoma.

Immune status affects the initiation and progression of clear cell renal cell carcinoma (ccRCC), the most common subtype of renal cell carcinoma. In this study, we identified an immune-related, five-gene signature that improves survival prediction in ccRCC. Patients were classified as high- and low-risk based on the signature risk score. Survival analysis showed differential prognosis, while principal component analysis revealed distinctly different immune phenotypes between the two risk groups. High-risk patients tended to have advanced stage, higher grade disease, and poorer prognoses. Functional enrichment analysis showed that the signature genes were mainly involved in the cytokine-cytokine receptor interaction pathway. Moreover, we found that tumors from high-risk patients had higher relative abundance of T follicular helper cells, regulatory T cells, and M0 macrophages, and higher expression of PD-1, CTLA-4, LAG3, and CD47 than low-risk patients. This suggests our gene signature may not only serve as an indicator of tumor immune status, but may be a promising tool to select high-risk patients who may benefit from immune checkpoint inhibitor therapy. Multivariate Cox regression analysis showed that the signature remained an independent prognostic factor after adjusting for clinicopathological variables, while prognostic accuracy was further improved after integrating clinical parameters into the analysis.