Negative Regulator Of Cell Proliferation Research Articles

Screening and identification of key biomarkers in prostate cancer using bioinformatics.

Prostate cancer (PCa) is the second most common cancer amongst males worldwide. In the current study, microarray datasets GSE3325 and GSE6919 from the Gene Expression Omnibus database were screened to identify candidate genes that are associated with the progression of PCa. A total of 273 differentially expressed genes (DEGs) were identified, which included 173 downregulated genes and 100 upregulated genes, and a protein-protein interaction network was constructed using Search Tool for the Retired of Interacting Genes. The enriched functions and pathways of the identified DEGs included cell adhesion, the negative regulation of cell proliferation, protein binding and focal adhesion. A total of 8 hub genes were identified, of which PDZ binding kinase, Krüppel-like factor 4, collagen type XII α-1 chain, RAP1A and RAP39B were indicated to be associated with the progression and recurrence of PCa. In conclusion, the DEGs and hub genes identified in the present study may aid in determining the molecular mechanisms associated with PCa carcinogenesis and progression.

MRNA expression profile of bronchoalveolar lavage fluid cells from patients with idiopathic pulmonary fibrosis and sarcoidosis.

Sarcoidosis and idiopathic pulmonary fibrosis (IPF) are two most frequent forms of interstitial lung diseases (ILDs). Cellular and biochemical composition of bronchoalveolar lavage fluid (BALf) was shown to reflect the fibrotic changes in the lung. However, the usefulness of BALf cellular profile evaluation in the diagnosis of ILDs is limited. The aim of the study was a multivariate, molecular analysis of BALf cells from IPF and sarcoidosis patients. Transcriptomic measurements were carried out using Affymetrix Human Gene 2.1 ST ArrayStrip in 21 samples: 9 IPF and 12 sarcoidosis. The mRNA expression for the most significantly differentiating genes was evaluated by real-time PCR in 32 samples (11 IPF and 21 sarcoidosis). The number of genes differentially expressed between IPF and sarcoidosis groups was 4832 (13359 probesets). Cluster analysis indicated that sarcoidosis BALf cells are characterized by increased mRNA expression of genes associated with ribosome biogenesis. Clusters formed by genes with changed mRNA expression in IPF samples were implicated in the processes of cell adhesion and migration, metalloproteinase expression and negative regulation of cell proliferation. The GO analysis indicated that predominant biological processes associated with the differential mRNA gene expression of BALf cells were upregulation of neutrophils in IPF and lymphocytes in sarcoidosis. Analysis of BALf from sarcoidosis and IPF showed highly different mRNA profile of cells. The most important biological processes observed at the molecular level in BALf cells were associated with ribosome biogenesis and proteasome apparatus in sarcoidosis and neutrophilic dysfunction in IPF.

Body Mass Index in Multiple Sclerosis modulates ceramide-induced DNA methylation and disease course.

BackgroundMultiple Sclerosis (MS) results from genetic predisposition and environmental variables, including elevated Body Mass Index (BMI) in early life. This study addresses the effect of BMI on the epigenome of monocytes and disease course in MS.MethodsFifty-four therapy-naive Relapsing Remitting (RR) MS patients with high and normal BMI received clinical and MRI evaluation. Blood samples were immunophenotyped, and processed for unbiased plasma lipidomic profiling and genome-wide DNA methylation analysis of circulating monocytes. The main findings at baseline were validated in an independent cohort of 91 therapy-naïve RRMS patients. Disease course was evaluated by a two-year longitudinal follow up and mechanistic hypotheses tested in human cell cultures and in animal models of MS.FindingsHigher monocytic counts and plasma ceramides, and hypermethylation of genes involved in negative regulation of cell proliferation were detected in the high BMI group of MS patients compared to normal BMI. Ceramide treatment of monocytic cell cultures increased proliferation in a dose-dependent manner and was prevented by DNA methylation inhibitors. The high BMI group of MS patients showed a negative correlation between monocytic counts and brain volume. Those subjects at a two-year follow-up showed increased T1 lesion load, increased disease activity, and worsened clinical disability. Lastly, the relationship between body weight, monocytic infiltration, DNA methylation and disease course was validated in mouse models of MS.InterpretationHigh BMI negatively impacts disease course in Multiple Sclerosis by modulating monocyte cell number through ceramide-induced DNA methylation of anti-proliferative genes.FundThis work was supported by funds from the Friedman Brain Institute, NIH, and Multiple Sclerosis Society.

Adipose-derived mesenchymal stromal cells improve hemodynamic function in pulmonary arterial hypertension: identification of microRNAs implicated in modulating endothelial function

Pulmonary arterial hypertension (PAH) is characterized by pulmonary arterial endothelial hyperproliferation and dysfunction. Restoration of endothelial function is a common goal of available treatments. In the present study, human adipose-derived mesenchymal stromal cells (ASCs) were co-cultured with monocrotaline pyrrole–treated human pulmonary arterial endothelial cells (HPAECs); increased proliferation of HPAECs and expression of vascular endothelial growth factor (VEGF) were observed. High throughput sequencing results showed that six microRNAs (miMNAs) of ASCs were significantly dysregulated. In monocrotaline-induced PAH rat models, ASC transplantation improved the right ventricle systolic pressure, right ventricle hypertrophy and pulmonary endothelium hyperproliferation, and four of the six miRNAs were validated in the lung tissue samples. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that these dysregulated miRNAs were involved in the regulation of transcription, signal transduction, negative regulation of cell proliferation through mitogen-activated protein kinase (MAPK) signaling pathway, Wnt signaling pathway, VEGF signaling pathway, cytokine-cytokine receptor interaction, regulation of actin cytoskeleton, transforming growth factor (TGF)-beta signaling pathway and P53 signaling pathway. Our data indicates that the unique six miRNA expression signature could be involved in the PAH endothelial repair by ASCs.

Discovery of targetable mutational signatures in advanced prostate cancer (aPC) using machine learning and next-generation sequencing (NGS) of circulating tumor DNA (ctDNA).

226 Background: Alterations (alts) of the AR gene are common in aPC and are associated with reduced responses to AR-axis inhibitors; optimal treatment is unknown. Genomic sequencing of ctDNA from aPCs may recapitulate truncal and branching alts across metastatic sites in heavily pretreated progressive aPC and facilitates identification of AR and other alts that contribute to aPC progression. Such data could lead to better understanding of aPC molecular biology and guide development of novel treatments. Methods: Two unsupervised machine learning algorithms, hierarchical clustering and principal components analysis, were used to evaluate mutational profiles of 2,679 plasma samples from 2,309 men with aPC. Samples were assessed by a validated ctDNA NGS panel that sequences 73 clinically relevant cancer genes (Guardant360, Redwood City, CA). Indels, amplifications, and fusions are evaluated in a subset of genes with high sensitivity and specificity. Chi-squared residual analysis and gene ontology enrichment were used to identify significantly co-altered molecular pathways. Results: Distinct, non-overlapping mutational signatures were observed in tumors with only AR amplification (amp) vs. only AR mutation (mut). Tumors with only AR amp, and those with both AR amp and mut, were strongly enriched for amps in the MAPK pathway (P = 3.62E-10, 4.92E-05, resp.). Tumors with only AR mut, and those with no AR alts, were strongly enriched for muts in negative regulators of cell proliferation (P = 1.09E-05, 3.30E-09, resp.). Muts further stratified to oxidative stress response in AR mut (P = 4.37E-05), and cell aging in no AR alts (P = 8.31E-07). Conclusions: In men with aPC, machine learning of ctDNA NGS data revealed recurrent mutational signatures that stratified according to the type of AR alts. These hypothesis-generating data require external validation but suggest that aPC may progress due to distinct mechanisms dependent on AR status. Future efforts to develop novel therapeutic approaches for aPC may need to consider the impact of the type of AR alt present and explore the interplay of AR with other altered pathways identified in this analysis.

Podocan affects C2C12 myogenic differentiation by enhancing Wnt/β-catenin signaling.

Podocan, a small leucine-rich repeat protein, is a negative regulator of cell proliferation. In this study, we demonstrated that podocan is involved in the differentiation of C2C12 murine myoblasts. Podocan expression increased with the progression of C2C12 differentiation. As expect, siRNA-mediated podocan knockdown inhibited C2C12 differentiation, as indicated by inhibition of MYOG, MYH2, and desmin expression, as well as reductions in the differentiation and fusion indices. Overexpression of podocan using dCas9 technology promoted C2C12 cell differentiation. In addition, supplementation of culture medium with podocan influenced C2C12 differentiation. Podocan knockdown reduced Wnt/β-catenin signaling, characterized by a reduction in the nuclear translocation of β-catenin, whereas podocan overexpression had the opposite effect. Furthermore, treatment with XAV939, an inhibitor of Wnt/β-catenin, reduced the podocan-mediated promotion of C2C12 differentiation. Induction of muscle injury in mice by bupivacaine administration suggested that podocan may play a role in muscle regeneration. In summary, our results suggest that podocan is required for normal C2C12 differentiation and that its role in myogenesis is mediated by the Wnt/β-catenin pathway.

A Zebrafish JAK1-A634D Gain-of-Function Model Provides New Insights into the Pathogenesis of Familial Hypereosinophilia

Introduction: JAK/STAT signaling is of major importance for hematopoiesis and immune function. A heterozygous JAK1 c.1901C>A de novo point mutation was identified in three members of a family (a mother and two male sons). These patients presented with a unique clinical phenotype of severe atopic dermatitis, markedly elevated peripheral blood eosinophil counts with eosinophilic infiltration of the liver and gastrointestinal tract, hepatosplenomegaly, and failure to thrive. Treatment with ruxolitinib, a JAK1/2 inhibitor, resulted in reduced eosinophilia and improved growth (Del Bel et al. JACI 2017). However, the role of this JAK1 mutation in hematopoiesis remains unclear.Methods: ubi-JAK1-WT or A634D-P2A-sfGFP-pA zebrafish transgenic lines were generated incorporating the human genes. Both of these constructs were injected into embryos, but the survival rate for the JAK1-A634D embryos was very low. Six hour post-fertilization (hpf) embryos expressing the mutant transgene were treated with 1 µM ruxolitinib for 24h, which dramatically increased survival. Whole-mount in situ hybridization (WISH) was used to assess changes in hematopoietic stem cells/blood cell populations due to wild-type or JAK1-A634D expression at 24, 28, 36 and 48 hpf using a battery of blood lineage specific probes. O-dianisidine staining was employed to evaluate hemoglobin levels. ImageJ cell counter pluggin was used for cell quantification. Gata1 and o-dianisidine staining intensity was analyzed using Ilastik and CellProfiler. Six groups of 30-50 embryos were analyzed by RNA sequencing (RNAseq): control, JAK1-WT and JAK1-A634D at 28 and 36 hpf. RNA was extracted using Trizol. RNAseq data was analyzed with edgeR and enrichment analysis was performed using DAVID 6.8.Results: JAK1-A634D was toxic to zebrafish embryos but they survived to sexual maturity post-ruxolitinib therapy. F1 embryos from JAK1-A634D founders with high expression of the transgene exhibited abnormal development (70% of embryos) which could be reduced to 12% of embryos following 24h exposure to 1 µM ruxolitinib. This rescue suggests that JAK1-A634D is highly active in zebrafish and its activity needs to be under tight regulation for normal development. Both JAK1-WT and A634D mutant fish demonstrated increased HSPCs (c-myb/runx1 expression) at 36 hpf (p<0.0001) through 48 hpf (A634D>WT>controls; p=0.0308 and p=0.0045, respectively). pu.1/spi1 early myeloid expression showed a similar expression pattern at 28 hpf, A634D>WT>controls; p<0.0001 for both). At 24 and 28 hpf, granulocytes expressing myeloperoxidase (mpx) were increased in both JAK1-WT and JAK1-A634D mutants, but particularly high in JAK1-A634D embryos at 48 hpf (p<0.0001). Macrophages marked by l-plastin (lcp) expression, showed similar cell numbers in JAK1-WT and controls at 24 and 28 hpf, but a significant decrease in JAK1-A634D embryos (p<0.0001). An increase of carboxypeptidase A5 positive mast cells was observed in both JAK1-WT and JAK1-A634D embryos at 48 hpf compared with controls (p<0.0001). We observed an increase of gata1a expression at 24 hpf in A634D embryos (p=0.0058, p=0.0065), but o-dianisidine staining at 48 hpf revealed a decrease in hemoglobin in both JAK1 lines (p<0.0001). The latter data is supported by RNAseq results revealing decreased expression of hemoglobin-associated genes at 36 hpf in both transgenic lines. RNAseq revealed a total of 365 genes differentially expressed, with enrichment for genes coding for proteosomal and ribosomal proteins. In the JAK-1WT we found 79 genes upregulated and 115 downregulated; in the JAK1-A634D, 94 genes were upregulated and 77 were downregulated. JAK-STAT pathway genes were upregulated in the A634D mutant (p<0.05), as well as a trend for genes associated with hematopoiesis (p=0.0633) and negative regulation of cell proliferation (p=0.0658).Conclusions: JAK1-A634D plays an important role in zebrafish development and specifically impacts the myeloid lineage, particularly the granulocyte branch, from which eosinophils arise. Ruxolitinib can revert the arrested development phenotype, demonstrating the specificity of this targeted therapy. Our model provides the first in vivo evidence of the effects of the JAK1-A634D mutation on hematopoiesis and will provide further insight in understanding the cause of familial hypereosinophilia. DisclosuresNo relevant conflicts of interest to declare.

Expression Profiles Analysis Identification and Interaction of Key Genes and Micrornas in Hepatocellular Carcinoma

Background: Hepatocellular carcinoma is one of the most common malignant tumors, with rapid development and high malignancy. MicroRNAs have been reported to play important roles in hepatocellular carcinoma progression. Aim: To identify the key genes and miRNAs in HCC, and to explore their potential molecular mechanisms. Methods: Gene expression profiles of GSE15471 (mRNA profile) and GSE57555 (miRNA profile) were downloaded from gene expression omnibus, which were analysed using R software and bioconductor packages. The gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichments of DEGs were performed using the DAVID database, and the protein–protein interaction networks of the DEGs were constructed from the STRING database. In addition, targets of differentially expressed miRNAs were predicted by the online resource miRDB. Result: In total, 191 differentially expressed genes were identified, including 142 upregulated and 49 downregulated genes. Functional analysis revealed that these DEGs were associates with wound healing, endodermal cell-cell adhesion, activation of MAPK activity and negative regulation of cell proliferation. In addition, we identified five DEMs, which were upregulated and downregulated. hsa-miR-122-5p may target the PDK4, and hsa-miR-21-5p probably targets SPOCK1 and PAIP2B. Conclusions: We applied integrated bioinformatics to identify key pathogenic genes involved in hepatocellular carcinoma and provide new clues for further studies of hepatocellular carcinoma.

SALL2 represses cyclins D1 and E1 expression and restrains G1/S cell cycle transition and cancer-related phenotypes.

SALL2 is a poorly characterized transcription factor that belongs to the Spalt‐like family involved in development. Mutations on SALL2 have been associated with ocular coloboma and cancer. In cancers, SALL2 is deregulated and is proposed as a tumor suppressor in ovarian cancer. SALL2 has been implicated in stemness, cell death, proliferation, and quiescence. However, mechanisms underlying roles of SALL2 related to cancer remain largely unknown. Here, we investigated the role of SALL2 in cell proliferation using mouse embryo fibroblasts (MEFs) derived from Sall2 −/− mice. Compared to Sall2 +/+ MEFs, Sall2 −/− MEFs exhibit enhanced cell proliferation and faster postmitotic progression through G1 and S phases. Accordingly, Sall2 −/− MEFs exhibit higher mRNA and protein levels of cyclins D1 and E1. Chromatin immunoprecipitation and promoter reporter assays showed that SALL2 binds and represses CCND1 and CCNE1 promoters, identifying a novel mechanism by which SALL2 may control cell cycle. In addition, the analysis of tissues from Sall2 +/+ and Sall2 −/− mice confirmed the inverse correlation between expression of SALL2 and G1‐S cyclins. Consistent with an antiproliferative function of SALL2, immortalized Sall2 −/− MEFs showed enhanced growth rate, foci formation, and anchorage‐independent growth, confirming tumor suppressor properties for SALL2. Finally, cancer data analyses show negative correlations between SALL2 and G1‐S cyclins’ mRNA levels in several cancers. Altogether, our results demonstrated that SALL2 is a negative regulator of cell proliferation, an effect mediated in part by repression of G1‐S cyclins’ expression. Our results have implications for the understanding and significance of SALL2 role under physiological and pathological conditions.

Tumour cell dormancy as a contributor to the reduced survival of GBM patients who received standard therapy.

Glioblastoma multiforme (GBM) is a fatal cancer with varying life expectancy, even for patients undergoing the same standard therapy. Identification of differentially expressed genes in GBM patients with different survival rates may benefit the development of effective therapeutic strategies. In the present study, key pathways and genes correlated with survival in GBM patients were screened with bioinformatic analysis. Included in the study were 136eligible patients who had undertaken surgical resection of GBM followed by temozolomide (TMZ) chemoradiation and long-term therapy with TMZ. A total of 383differentially expressed genes (DEGs) related to GBM survival were identified. Gene Ontology and pathway enrichment analysis as well as hub gene screening and module analysis were performed. As expected, angiogenesis and migration of GBM cells were closely correlated with a poor prognosis. Importantly, the results also indicated that cell dormancy was an essential contributor to the reduced survival of GBM patients. Given the lack of specific targeted genes and pathways known to be involved in tumour cell dormancy, we proposed enriched candidate genes related to the negative regulation of cell proliferation, signalling pathways regulating pluripotency of stem cells and neuroactive ligand-receptor interaction, and 3hub genes (FTH1, GRM1 and DDIT3). Maintaining persistent cell dormancy or preventing tumour cells from entering dormancy during chemoradiation should be a promising therapeutic strategy.

Dietary Butyrate, an HDAC Inhibitor, Primes Muscle Satellite Cells for Differentiation by Altering the Epigenetic Landscape

Satellite cells (SC) are a heterogeneous population of skeletal muscle resident stem cells that are responsible for postnatal muscle growth and repair. Satellite cells lie in a state of quiescence along the muscle fiber where they will activate, proliferate, and either self‐renew or differentiate. With the investigation into the genomic regulation of satellite cell fate, we now have an idea as to how the satellite cell epigenome governs myogenesis. Specifically, changes in the chromatin structure and the regulators that direct these changes modulate SC activation, proliferation, and differentiation. Interestingly, histone deacetylase (HDAC) inhibitors have proven to be effective at enhancing the myogenic program of SC, but their precise mode of action in altering the epigenetic landscape of SC remains undetermined. Our objective was to determine how a HDAC inhibitor, butyrate, promotes myogenic differentiation. Twenty female neonatal piglets (24±8hrs old) were fed a standard milk replacer or one supplemented with 0.50% (on a dry‐matter basis) spray dried Tributyrin (TB), a butyrate pro‐drug, for 17d. Satellite cells from TB supplemented piglets showed a decrease in the expression of EZH2 (enhancer of zeste homolog 2), the methyltransferase subunit of the PRC2 (polycomb repressive complex 2), just prior to being induced to differentiate. ChIP‐Seq analysis revealed that SC from TB treated piglets resulted in a global reduction of the repressive chromatin mark H3K27me3. Of the 532 differentially enriched regions (DER), TB treatment decreased binding at 358 sites, specifically those sites associated with genes involved in myogenic differentiation, negative regulation of cell proliferation, and micro‐RNAs normally upregulated during differentiation. ChIP‐Seq analysis for global enrichment of H3K27ac did not reveal any numerical differences in DER resulting from TB treatment. Given that EZH2 is responsible for the deposition of the repressive H3K27me3 mark, SC were transfected with siRNA targeting the EZH2 transcript (siEZH2), treated with 0.5 mM sodium butyrate (NaBu), or both (siEZH2 + NaBu) to determine if this was the primary mechanism through which butyrate alters SC myogenesis. Treatment with NaBu and siEZH2 reduced EZH2 expression in an additive manner. Treatment with NaBu alone and with siEZH2 significantly reduced Pax7 and MyoD expression after 24h of treatment while siEZH2 treatment had no effect on the expression of these myogenic transcription factors. Immunostaining for the contractile protein myosin heavy chain (MyHC) revealed an increase in MyHC+ cells from the siEZH2 treatment group after 72h of differentiation; however, these cells did not form multinucleated myotubes. The epigenetic changes caused by dietary TB supplementation resulted in reduced EZH2 and EZH2 trimethylation of H3K27. Taken together, these results indicate that while EZH2 ablation may force myogenic differentiation, butyrate may operate through a parallel mechanism to enhance differentiation. These findings help support the notion that butyrate may be used to enhance muscle growth and could prove useful at treating myopathic conditions.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Identification of novel drug targets for diamond-blackfan anemia based on RPS19 gene mutation using protein-protein interaction network

BackgroundDiamond-Blackfan anemia (DBA) is a congenital erythroid aplasia that usually presents in infancy. In order to explore the molecular mechanisms of wild and mutated samples from DBA patients were exposed to bioinformatics investigation. Biological network of differentially expressed genes was constructed. This study aimed to identify novel therapeutic signatures in DBA and uncovered their mechanisms. The gene expression dataset of GSE14335 was used, which consists of 6 normal and 4 diseased cases. The gene ontology (GO), as well as Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed, and then protein–protein interaction (PPI) network of the identified differentially expressed genes (DEGs) was constructed by Cytoscape software.ResultsA total of 607 DEGs were identified in DBA, including 433 upregulated genes and 174 downregulated genes. GO analysis results showed that upregulated DEGs were significantly enriched in biological processes, negative regulation of transcription from RNA polymerase II promoter, chemotaxis, inflammatory response, immune response, positive regulation of cell proliferation, negative regulation of cell proliferation, response to mechanical stimulus, positive regulation of cell migration, response to lipopolysaccharide, and defence response. KEGG pathway analysis revealed the TNF signalling pathway, Osteoclast differentiation, Chemokine signalling pathway, Cytokine -cytokine receptor interaction, Rheumatoid arthritis, Biosynthesis of amino acids, Biosynthesis of antibiotics and Glycine, serine and threonine metabolism. The top 10 hub genes, AKT1, IL6, NFKB1, STAT3, STAT1, RAC1, EGR1, IL8, RELA, RAC3, mTOR and CCR2 were identified from the PPI network and sub-networks.ConclusionThe present study flagged that the identified DEGs and hub genes enrich our understanding of the molecular mechanisms underlying the development of DBA, and might shine some lights on identifying molecular targets and diagnostic biomarkers for DBA.

Deep Coverage Tissue and Cellular Proteomics Revealed IL-1β Can Independently Induce the Secretion of TNF-Associated Proteins from Human Synoviocytes.

Synovitis is a key contributor to the inflammatory environment in osteoarthritis (OA) joints. Currently, the biological therapy of OA is not satisfactory in multiple single-target trials on anti-TNF agents, or IL-1 antagonists. Systems biological understanding of the phosphorylation state in OA synovium is warranted to direct further therapeutic strategies. Therefore, in this study, we compared the human synovial phosphoproteome of the OA with the acute joint fracture subjects. We found that OA synovium had significantly more phosphoproteins, and 82 phosphoproteins could only be specifically found in all the OA samples. Differentially expressed proteins of the OA synovium were focusing on endoplasmic reticulum-/Golgi-associated secretion and negative regulation of cell proliferation, which was verified through an IL-1β-treated human synoviocyte (HS) in vitro model. With data-independent acquisition-based mass spectrometry, we found that IL-1β could induce HS to secrete proteins that were significantly associated with the endosomal/vacuolar pathway, endoplasmic reticulum/Golgi secretion, complement activation, and collagen degradation. Especially, we found that while specifically suppressing HS endocytosis, IL-1β could activate the secretion of 25 TNF-associated proteins, and the change of SERPINE2 and COL3A1 secretion was verified by immunoblotting. In conclusion, our results suggest that OA synovium has a polarized phosphoproteome to inhibit proliferation and maintain active secretion of HS, whereas IL-1β alone can transform HS to produce a synovitis-associated secretome, containing numerous TNF-associated secretory proteins in a TNF-independent mode.

Identification of Key Genes and Pathways in Tongue Squamous Cell Carcinoma Using Bioinformatics Analysis.

BackgroundTongue squamous cell carcinoma (TSCC) is a major type of oral cancers and has remained an intractable cancer over the past decades. The aim of this study was to identify differentially expressed genes (DEGs) during TSCC and reveal their potential mechanisms.Material/MethodsThe gene expression profiles of GSE13601 were downloaded from the GEO database. The GSE13601 dataset contains 57 samples, including 31 tongue SCC samples and 26 matched normal mucosa samples. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) enrichment analyses were performed; Cytoscape software was used for the protein-protein interaction (PPI) network and module analysis of the DEGs.ResultsWe identified a total of 1,050 upregulated DEGs (uDEGs) and 702 downregulated DEGs (dDEGs) of TSCC. The GO analysis results showed that uDEGs were significantly enriched in the following biological processes (BP): signal transduction, positive or negative regulation of cell proliferation, and negative regulation of cell proliferation. The dDEGs were significantly enriched in the following biological processes: signal transduction, cell adhesion, and apoptotic process. The KEGG pathway analysis showed that uDEGs were enriched in metabolic pathways, pathways in cancer, and PI3K-Akt signaling pathway, while the dDEGs were enriched in focal adhesion and ECM-receptor interaction. The top centrality hub genes RAC1, APP, EGFR, KNG1, AGT, and HRAS were identified from the PPI network. Module analysis revealed that TSCC was associated with significant pathways, including neuroactive ligand-receptor interaction, calcium signaling pathway, and chemokine signaling pathway.ConclusionsThe present study identified key genes and signal pathways, which deepen our understanding of the molecular mechanisms of carcinogenesis and development of the disease, and might be used as diagnostic and therapeutic molecular biomarkers for TSCC.

TMEM16A regulates portal vein smooth muscle cell proliferation in portal hypertension.

The aim of the present study was to elucidate the effect of transmembrane protein 16A (TMEM16A) on portal vein smooth muscle cell (PVSMC) proliferation associated with portal vein remodeling in portal hypertension (PHT). Sprague-Dawley rats were subjected to bile duct ligation to establish a rat model of liver cirrhosis and PHT. Sham-operated animals served as controls. At 8 weeks after bile duct ligation, the extent of liver fibrosis and the portal vein wall thickness were assessed using hematoxylin-eosin staining. The protein expression levels of TMEM16A, extracellular signal-regulated kinase 1 and 2 (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2) in the portal vein were detected by immunohistochemistry and western blotting. In vitro, the lentivirus vectors were constructed and transfected into PVSMCs to upregulate the expression of TMEM16A. Isolated rat primary PVSMCs were treated with a small molecule inhibitor of TMEM16A, T16A-inhA01. Cell cycle was detected by flow cytometry. The activity of TMEM16A in the portal vein isolated from bile duct ligated rats was decreased, while the expression level of p-ERK1/2 was increased. However, in vitro, upregulation of TMEM16A promoted the proliferation PVSMCs, while inhibition of TMEM16A channels inhibited the proliferation of PVSMCs. The results indicated that TMEM16A contributes to PVSMCs proliferation in vitro, but in vivo, it may be a negative regulator of cell proliferation influenced by numerous factors.

Microarray analysis to identify the similarities and differences of pathogenesis between aortic occlusive disease and abdominal aortic aneurysm.

Objectives Expression profile of GSE57691 was analyzed to identify the similarities and differences between aortic occlusive disease and abdominal aortic aneurysm. Methods The expression profile of GSE57691 was downloaded from Gene Expression Omnibus database, including 20 small abdominal aortic aneurysm samples, 29 large abdominal aortic aneurysm samples, 9 aortic occlusive disease samples, and 10 control samples. Using the limma package in R, the differentially expressed genes were screened. Followed by enrichment analysis was performed for the differentially expressed genes using database for annotation, visualization, and integrated discovery online tool. Based on string online tool and Cytoscape software, protein-protein interaction network and module analyses were carried out. Moreover, integrated TF platform database and Cytoscape software were used for constructing transcriptional regulatory networks. Results As a result, 1757, 354, and 396 differentially expressed genes separately were identified in aortic occlusive disease, large abdominal aortic aneurysm, and small abdominal aortic aneurysm samples. UBB was significantly enriched in proteolysis related pathways with a high degree in three groups. SPARCL1 was another gene shared by these groups and regulated by NFIA, which had a high degree in transcriptional regulatory network. ACTB, a significant upregulated gene in abdominal aortic aneurysm samples, could be regulated by CLIC4, which was significantly enriched in cell motions. ACLY and NFIB were separately identified in aortic occlusive disease and small abdominal aortic aneurysm samples, and separately enriched in lipid metabolism and negative regulation of cell proliferation. Conclusions The downregulated UBB, NFIA, and SPARCL1 might play key roles in both aortic occlusive disease and abdominal aortic aneurysm, while the upregulated ACTB might only involve in abdominal aortic aneurysm. ACLY and NFIB were specifically involved in aortic occlusive disease and small abdominal aortic aneurysm separately.

Identification of differentially expressed genes regulated by molecular signature in breast cancer-associated fibroblasts by bioinformatics analysis.

Breast cancer is a severe risk to public health and has adequately convoluted pathogenesis. Therefore, the description of key molecular markers and pathways is of much importance for clarifying the molecular mechanism of breast cancer-associated fibroblasts initiation and progression. Breast cancer-associated fibroblasts gene expression dataset was downloaded from Gene Expression Omnibus database. A total of nine samples, including three normal fibroblasts, three granulin-stimulated fibroblasts and three cancer-associated fibroblasts samples, were used to identify differentially expressed genes (DEGs) between normal fibroblasts, granulin-stimulated fibroblasts and cancer-associated fibroblasts samples. The gene ontology (GO) and pathway enrichment analysis was performed, and protein-protein interaction (PPI) network of the DEGs was constructed by NetworkAnalyst software. Totally, 190 DEGs were identified, including 66 up-regulated and 124 down-regulated genes. GO analysis results showed that up-regulated DEGs were significantly enriched in biological processes (BP), including cell-cell signalling and negative regulation of cell proliferation; molecular function (MF), including insulin-like growth factor II binding and insulin-like growth factor I binding; cellular component (CC), including insulin-like growth factor binding protein complex and integral component of plasma membrane; the down-regulated DEGs were significantly enriched in BP, including cell adhesion and extracellular matrix organization; MF, including N-acetylgalactosamine 4-sulfate 6-O-sulfotransferase activity and calcium ion binding; CC, including extracellular space and extracellular matrix. WIKIPATHWAYS analysis showed the up-regulated DEGs were enriched in myometrial relaxation and contraction pathways. WIKIPATHWAYS, REACTOME, PID_NCI and KEGG pathway analysis showed the down-regulated DEGs were enriched endochondral ossification, TGF beta signalling pathway, integrin cell surface interactions, beta1 integrin cell surface interactions, malaria and glycosaminoglycan biosynthesis-chondroitin sulfate/dermatan sulphate. The top 5 up-regulated hub genes, CDKN2A, MME, PBX1, IGFBP3, and TFAP2C and top 5 down-regulated hub genes VCAM1, KRT18, TGM2, ACTA2, and STAMBP were identified from the PPI network, and subnetworks revealed these genes were involved in significant pathways, including myometrial relaxation and contraction pathways, integrin cell surface interactions, beta1 integrin cell surface interaction. Besides, the target hsa-mirs for DEGs were identified. hsa-mir-759, hsa-mir-4446-5p, hsa-mir-219a-1-3p and hsa-mir-26a-5p were important miRNAs in this study. We pinpoint important key genes and pathways closely related with breast cancer-associated fibroblasts initiation and progression by a series of bioinformatics analysis on DEGs. These screened genes and pathways provided for a more detailed molecular mechanism underlying breast cancer-associated fibroblasts occurrence and progression, holding promise for acting as molecular markers and probable therapeutic targets.

Bioinformatics analysis of fibroblasts exposed to TGF‑β at the early proliferation phase of wound repair.

The aim of the current study was to identify gene signatures during the early proliferation stage of wound repair and the effect of TGF-β on fibroblasts and reveal their potential mechanisms. The gene expression profiles of GSE79621 and GSE27165 were obtained from GEO database. Differentially expressed genes (DEGs) were identified using Morpheus and co-expressed DEGs were selected using Venn Diagram. Gene ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of DEGs were performed using the Database for Annotation, Visualization and Integrated Discovery (DAVID) online tool. Protein-protein interaction (PPI) networks of the DEGs were constructed using Cytoscape software. PPI interaction network was divided into subnetworks using the MCODE algorithm and the function of the top one module was analyzed using DAVID. The results revealed that upregulated DEGs were significantly enriched in biological process, including the Arp2/3 complex-mediated actin nucleation, positive regulation of hyaluronan cable assembly, purine nucleobase biosynthetic process, de novo inosine monophosphate biosynthetic process, positive regulation of epithelial cell proliferation, whereas the downregulated DEGs were enriched in the regulation of blood pressure, negative regulation of cell proliferation, ossification, negative regulation of gene expression and type I interferon signaling pathway. KEGG pathway analysis showed that the upregulated DEGs were enriched in shigellosis, pathogenic Escherichia coli infection, the mitogen-activated protein kinase signaling pathway, Ras signaling pathway and bacterial invasion of epithelial cells. The downregulated DEGs were enriched in systemic lupus erythematosus, lysosome, arachidonic acid metabolism, thyroid cancer and allograft rejection. The top 10 hub genes were identified from the PPI network. The top module analysis revealed that the included genes were involved in ion channel, neuroactive ligand-receptor interaction pathway, purine metabolism and intestinal immune network for IgA production pathway. The functional analysis revealed that TGF-β may promote fibroblast migration and proliferation and defend against microorganisms at the early proliferation stage of wound repair. Furthermore, these results may provide references for chronic wound repair.

The Arabidopsis thaliana NGATHA1 transcription factor acts as a promoter of a general differentiation program and a carpel identity factor

The Arabidopsis thaliana NGATHA (NGA) family is a four-membered subclade of the B3-type transcription factor superfamily, and plays an essential role in the style development. NGAs also act as negative regulators of cell proliferation in leaf and petal growth. The notion that NGAs might act as promoters of a general differentiation program has been proposed to explain the involvement of NGAs in these seemingly discrete functions. Here, we tested this notion by activating NGA1 transcription in the expression domains of Arabidopsis thaliana Meristem Layer 1, AINTEGUMENTA, SHOOTMERISTEMLESS, CLAVATA3 (CLV3), and WUSCHEL (WUS), where cells remain mostly undifferentiated and pluripotent. We found that those specific cells expressing NGA1 lost their meristematic competence, becoming terminally differentiated. We also found that the inflorescence meristems of the plants expressing NGA1 in the CLV3 domain failed to develop floral meristems and differentiated into a terminal carpeloid cylinder without any other floral organs; similarly, the floral meristems of the plants expressing NGA1 in the WUS domain all developed into carpeloid cylinders with no other floral organs. Our results not only support the notion that NGA1 seems to be a promoter of a general differentiation program, but also raise the possibility that NGA1 may act as a carpel identity factor.

FOXO in B-cell lymphopoiesis and B cell neoplasia.

FOX O family transcription factors are important for differentiation and function of multiple cell types. In B lymphocytes they play a critical role. The activity of FOXOs is directly regulated both by signaling from B cell receptor (BCR) and cytokine receptors. FOXO1 action controls the transition between differentiation stages of B cell development. In comparison to other FOXO family members, FOXO1 plays a superior role in the regulation of early stages of B-cell differentiation. Although being known as a negative regulator of cell proliferation and therefore potential tumor suppressor, FOXO1 is downregulated only in Hodgkin lymphoma (HL) subtypes. In non-Hodgkin lymphoma (NHL) entities its expression is maintained at significant levels, raising the question on the role of FOXO-transcription factors in the proliferation and survival programs in the process of B cell differentiation as well as their contribution to the oncogenic programs of B-cell lymphomas. In particular, we discuss molecular mechanisms that might determine the switch between pro-apoptotic and pro-survival effects of FOXO1 and their interplay with specific differentiation programs.