O-glycan Biosynthesis Research Articles

Loss of the disease-associated glycosyltransferase Galnt3 alters Muc10 glycosylation and the composition of the oral microbiome

Loss of the disease-associated glycosyltransferase Galnt3 alters Muc10 glycosylation and the composition of the oral microbiome

Integrated analysis of lncRNA-associated ceRNA network identified potential regulatory interactions in osteosarcoma.

This study aimed to identify potential therapeutic targets in osteosarcoma (OS) through the network analysis of competing endogenous RNAs (ceRNAs). The differentially expressed miRNAs (DEMIs) and mRNAs (DEMs) were identified between OS cell lines and human mesenchymal stem cells (hMSCs) from the data deposited under GSE70415 using limma package. Functional analysis of DEMs was performed using DAVID and clusterProfiler to identify significantly enriched Gene Ontology biological processes and KEGG pathways, respectively. The DEMI-DEM interaction network was constructed using Cytoscape. LncRNA–miRNA interactions were predicted using starBase database. The ceRNA regulatory network was constructed by integrating mRNAs, miRNAs, and lncRNAs, and functional enrichment analysis was performed for the genes involved. The analysis revealed a total of 326 DEMs and 54 DEMIs between OS cells and hMSCs. We identified several novel therapeutic targets involved in the progression and metastasis of OS, such as CBX7, RAD9A, SNHG7 and miR-34a-5p. The miRNA, miR-543 (target gene: CBX7) was found to be associated with the pathway Mucin type O-glycan biosynthesis. Using the ceRNA network, we established the following regulatory interactions: NEAT1/miR-543/CBX7, SNHG7/miR-34a-5p/RAD9A, and XIST/miR-34a-5p/RAD9A. CBX7, RAD9A, lncRNA SNHG7, miR-543, and miR-34a-5p may be explored as novel therapeutic targets for treatment of OS.

Design, Synthesis and Bioactivity of Core 1 O-glycan and its Derivative on Human Gut Microbiota

Background: Disaccharide core 1 (Galβ1-3GalNAc) is a common O-glycan structure in nature. Biochemical studies have confirmed that the formation of the core 1 structure is an important initial step in O-glycan biosynthesis and it is of great importance for human body. Objective: Our study will provide meaningful and useful sights for O-glycan synthesis and their bioassay. And all the synthetic glycosides would be used as intermediate building blocks in the scheme developed for oligosaccharide construction. Methods: In this article, we firstly used chemical procedures to prepare core 1 and its derivative, and a novel disaccharide was efficiently synthesized. The structures of the synthesized compounds were elucidated and confirmed by 1H NMR, 13C NMR and MS. Then we employed three human gut symbionts belonging to Bacteroidetes, a predominantphyla in the distal gut, as models to study the bioactivity of core 1 and its derivative on human gut microbiota. Results: According to our results, both core 1 and derivative could support the growth of B. fragilis, especially the core 1 derivative, while failed to support the growth of B. thetaiotaomicron and B. ovatus. Conclusion: This suggested that the B. fragilis might have the specificity glycohydrolase to cut the glycosidic bond for acquiring monosaccharide.

Label-free quantitative proteomic analysis of molting-related proteins of Trichinella spiralis intestinal infective larvae

Molting is a key step for body-size expansion and environmental adaptation of parasitic nematodes, and it is extremely important for Trichinella spiralis growth and development, but the molting mechanism is not fully understood. In this work, label-free LC–MS/MS was used to determine the proteome differences between T. spiralis muscle larvae (ML) at the encapsulated stage and intestinal infective larvae (IIL) at the molting stage. The results showed that a total of 2885 T. spiralis proteins were identified, 323 of which were differentially expressed. These proteins were involved in cuticle structural elements, regulation of cuticle synthesis, remodeling and degradation, and hormonal regulation of molting. These differential proteins were also involved in diverse intracellular pathways, such as fatty acid biosynthesis, arachidonic acid metabolism, and mucin type O-glycan biosynthesis. qPCR results showed that five T. spiralis genes (cuticle collagen 14, putative DOMON domain-containing protein, glutamine synthetase, cathepsin F and NADP-dependent isocitrate dehydrogenase) had significantly higher transcriptional levels in 10 h IIL than ML (P < 0.05), which were similar to their protein expression levels, suggesting that they might be T. spiralis molting-related genes. Identification and characterization of T. spiralis molting-related proteins will be helpful for developing vaccines and new drugs against the early enteral stage of T. spiralis.

Autoantibodies Specific for Galactose-Deficient IgA1 in IgA Vasculitis With Nephritis

IntroductionPatients with IgA nephropathy (IgAN) have elevated serum levels of galactose-deficient IgA1 (Gd-IgA1) that are bound by Gd-IgA1–specific autoantibodies in pathogenic immune complexes. Renal biopsy histopathologic features of IgA vasculitis (IgAV) with nephritis (IgAV-N) are similar to those of IgAN. Mucosal infections often are associated with clinical onset and exacerbation in both diseases. We investigated whether patients with IgAV-N share pathogenic characteristics of IgAN.MethodsWe generated IgA1- and IgG-secreting cell lines from Epstein-Barr virus (EBV)–immortalized cells of patients with IgAV without nephritis (IgAV-woN), IgAV-N, and IgAN and from healthy individuals. Sera and cell-culture supernatants were used for analysis of Gd-IgA1 and Gd-IgA1–specific IgG autoantibodies.ResultsIgA1-producing cells from patients with IgAV-N, like cells from patients with IgAN, secreted more Gd-IgA1 than did cells from patients with IgAV-woN or healthy control subjects, in agreement with elevated serum Gd-IgA1 levels in patients with IgAV-N and IgAN. IgA1-producing cells from patients with IgAV-N had altered expression of genes involved in O-glycan biosynthesis: decreased for core 1 synthase (glycoprotein-N-acetylgalactosamine 3-β-galactosyltransferase 1; C1GALT1) and C1GALT1 Specific Chaperone 1 (C1GALTC1; COSMC) and elevated for N-acetylgalactosaminide α-2,6-sialyltransferase 2 (ST6GALNAC2). Levels of Gd-IgA1–specific IgG in sera and supernatants of IgG-producing cells were similar for patients with IgAV-N and IgAN and higher than those for IgAV-woN patients or healthy control subjects. Moreover, patients with IgAV-N who had active disease, manifested by hematuria and substantial proteinuria, had higher serum levels of Gd-IgA1–specific IgG autoantibodies than did patients with IgAV-N who had inactive disease.ConclusionSerum levels and cellular production of Gd-IgA1 and Gd-IgA1–specific IgG autoantibodies were elevated in patients with IgAV-N, supporting the hypothesis that IgAV-N and IgAN share pathogenic features.

The effects of resveratrol feeding and exercise training on the skeletal muscle function and transcriptome of aged rats.

This study investigated the effects of resveratrol feeding and exercise training on the skeletal muscle function and transcriptome of aged rats. Male SD rats (25 months old) were divided into the control group (Old), the daily exercise training group (Trained), and the resveratrol feeding group (Resveratrol). After 6 weeks of intervention, the body mass, grip strength, and gastrocnemius muscle mass were determined, and the muscle samples were analyzed by transcriptome sequencing. The differentially expressed genes were analyzed followed by GO enrichment analysis and KEGG analysis. The Old group showed positive increases in body mass, while both the Trained and Resveratrol groups showed negative growth. No significant differences in the gastrocnemius muscle index and absolute grip strength were found among the three groups. However, the relative grip strength was higher in the Trained group than in the Old group. Only 21 differentially expressed genes were identified in the Trained group vs. the Old group, and 12 differentially expressed genes were identified in the Resveratrol group vs. the Old group. The most enriched GO terms in the Trained group vs. the Old group were mainly associated with RNA metabolic processes and transmembrane transporters, and the significantly upregulated KEGG pathways included mucin-type O-glycan biosynthesis, drug metabolism, and pyrimidine metabolism. The most enriched GO terms in the Resveratrol group vs. the Old group were primarily associated with neurotransmitter transport and synaptic vesicle, and the upregulated KEGG pathways included synaptic vesicle cycle, nicotine addiction, retinol metabolism, insulin secretion, retrograde endocannabinoid signaling, and glutamatergic synapse. Neither exercise training nor resveratrol feeding has a notable effect on skeletal muscle function and related gene expression in aged rats. However, both exercise training and resveratrol feeding have strong effects on weight loss, which is beneficial for reducing the exercise loads of the elderly.

Identification of altered microRNAs in retinas of mice with oxygen-induced retinopathy.

To identify disease-related miRNAs in retinas of mice with oxygen-induced retinopathy (OIR), and to explore their potential roles in retinal pathological neovascularization. The retinal miRNA expression profile in mice with OIR and room air controls at postnatal day 17 (P17) were determined through miRNA microarray analysis. Several miRNAs were significantly up- and down-regulated in retinas of mice with OIR compared to controls by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Two databases including Targetscan7.1 and MirdbV5 were used to predict target genes that associated with those significantly altered miRNAs in retinas of mice with OIR. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were also conducted to identify possible biological functions of the target genes. In comparison with room air controls, 3 and 8 miRNAs were significantly up- and down-regulated, respectively, in retinas of mice with OIR. The qRT-PCR data confirmed that mmu-miR-350-3p and mmu-miR-202-3p were significantly up-regulated, while mmu-miR-711 and mmu-miR-30c-1-3p were significantly down-regulated in mice with OIR compared to controls. GO analysis demonstrated that the identified target genes were related to functions such as cellular macromolecule metabolic process. KEGG pathway analysis showed a group of pathways, such as Wnt signaling pathway, transcriptional misregulation in cancer, Mucin type O-glycan biosynthesis, and mitogen-activated protein kinase (MAPK) signaling pathway might be involved in pathological process of retinal neovascularization. Our findings suggest that the differentially expressed miRNAs in retinas of mice with OIR might provide potential therapeutic targets for treating retinal neovascularization.

Bioinformatic Analysis of Key Genes in Cardiotoxic Compound Induced Long QT Interval

Background: Long QT (LQT) syndrome is an inherited disease with various triggering factors. The present study investigated the genetic role in medication induced LQT by conducting integrated bioinformatics analysis. Methods: The gene expression dataset of GSE59927 and GSE59905 were downloaded from the Gene Expression Omnibus (GEO) database. LQT associated medications with genetic informations were identified by crossing reference the list of cardiotoxic compounds in GSE dataset with the list of drugs with known risk of LQT. Common differentially expressed genes (DEGs) were extracted and compared with target genes of cardiac tissue specific microRNA. The overlapped mRNA were considered as tissue specific DEGs and employed to construct the regulatory network with microRNAs. The PPI network was built to identify the key genes. Findings: A total of 15 cardiac tissue specific DEGs in 14 different medications were identified and substantially enriched in the pathways of 'Calcium signaling pathway, Mucin type O-Glycan biosynthesis, Primary bile acid biosynthesis'. With the PPI network, 'Nrcam, Dlgap1, Gnal' were identified as key genes. Interpretation: The identification of key genes and related pathways in the present study provided genetic insight into the acquired LQT syndrome and may facilitate therapeutic individualization. Funding Statement: The authors state: No funding was received. Declaration of Interests: The authors declare that they have no competing interests, and all authors confirm its accuracy. Ethics Approval Statement: The analyses were based on previously published studies. Thus no ethical approval was required.

Comparative study of different molecular weight pullulan productions by Aureobasidium pullulans CGMCC No.11602.

Pullulan productions by Aureobasidium pullulans CGMCC No.11602 were conducted using the initial culture (IC) medium and the optimized culture (OC) medium, respectively, in which pullulan with significantly different molecular weights was obtained. Under the IC medium condition, the pullulan molecular weight (M w) reached 288,403Da with a yield of 64.12g/L after 96h fermentation period. However, the pullulan molecular weight was much higher (M w, 3,715,352Da), while the yield of pullulan was lower (40.12g/L) using the OC medium. The FTIR spectra indicated that pullulan produced using the IC and OC medium were similar. Transcriptome analysis showed that a total of 871 differentially expressed genes (DEGs) were screened and "N-glycan biosynthesis" and "other types of O-glycan biosynthesis" were the most highly represented differential metabolic pathways (DMPs). Specifically, the genes involved in the two DMPs consistently pointed to glucosyltransferase genes (GTF), all of which were up-regulated in the OC medium when compared with those in the IC medium. Further studies showed that the activity and the relative quantity (RQ) of GTF transcription were remarkable higher, which were coincident with the slower decrease in the molecular weight of pullulan in the OC medium than those in the IC medium during the late stage of fermentation. The results indicated that GTF may be involved in the regulation of pullulan molecular weight.

Expression patterns of novel circular RNAs in chicken cells after avian leukosis virus subgroup J infection

Avian leukosis virus subgroup J (ALV-J) is an oncogenic retrovirus that causes severe economic losses to the poultry industry worldwide. Circular RNAs (circRNAs) are a class of non-coding RNAs that has been described in various biological systems and pathogenic processes. However, the immune mechanisms in response to circRNAs remain unknown. In this study, high-throughput transcriptome sequencing was used to detect circRNAs present in chicken macrophage (HD11) and chick embryo fibroblast (CEF) cells infected with ALV-J. We identified 7684 circRNAs from diverse genomic locations in CEF and HD11 after ALV-J infection, these RNAs showed complex expression patterns that differed based on the cells type and infection time. In total, 302 differentially expressed (DE) circRNAs and 164 DE circRNAs were identified in CEF and HD11 after ALV-J infection, respectively. CircRNA7419—associated with KDM4C— and circRNA6679 and circRNA6680—associated with TNFAIP6— were involved in the immune response upon ALV-J infection in CEF. Host genes were analyzed through further bioinformatics analysis. The result confirmed that a large number of DE circRNAs corresponded to several immune-associated or tumor-associated terms and pathways, such as Mucin type O-Glycan biosynthesis, MAPK signaling pathway, B cell receptor signaling, and Wnt signaling pathway in CEF, as well as Jak-STAT signaling pathway, apoptosis, and MAPK signaling pathway in HD11. CircRNAs related to the B cell receptor signaling pathway in CEF, and the Jak-STAT signaling pathway in HD11, were selected for circRNA-miRNA interaction network analyses. Our study indicates that circRNAs expression was altered by ALV-J infection in both CEF and HD11, and may play a key role in the progression of ALV-J infection.

Mechanism of Low Expression of miR-30a-5p on Epithelial-Mesenchymal Transition and Metastasis in Ovarian Cancer.

Metastasis of ovarian cancer is regulated by microRNAs. This study focused on the effects of miR-30a-5p on ovarian cancer migration and invasion. Our results showed that the miR-30a-5p and mucin type O-glycan biosynthesis are closely related to ovarian cancer, and that miR-30a-5p was downregulated in ovarian cancer cells. miR-30a-5p overexpression reduced cell viability and inhibited migration and invasion in HO-8910 and HO-8910PM cells. S phase kinase-associated protein 2 (SKP2), B cell lymphoma 9 (BCL9), and NOTHC1 are direct target genes of miR-30a-5p. MTDH, SKP2, BCL9, and NOTCH1 genes were overexpressed in ovarian cancer cells, and they are direct target genes of miR-30a-5p. miR-30a-5p overexpression inhibited epithelial-mesenchymal transition (EMT) process, while upregulation of SKP2, BCL9, and NOTCH1 gene expression levels reduced the inhibition of EMT process by miR-30a-5p. miR-30a-5p was lowly expressed in ovarian cancer, and such a phenomenon is related to ovarian cancer metastasis. miR-30a-5p might inhibit the migration and invasion of ovarian cancer cells by downregulating the expression of SKP2, BCL9, and NOTCH1 genes.

MicroRNA-mediated Regulation of Mucin-type O-glycosylation Pathway: A Putative Mechanism of Salivary Gland Dysfunction in Sjögren Syndrome.

To investigate microRNA (miRNA) that is potentially implicated in primary Sjögren syndrome (pSS)-related salivary hypofunction in labial salivary glands and to study miRNA-mediated mechanisms underlying oral dryness and altered rheology, focusing on the mucin O-glycosylation pathway. We performed miRNA expression profiling in minor salivary gland samples of patients with pSS presenting a different impairment in their unstimulated salivary flow rate. A computational in silico analysis was performed to identify genes and pathways that might be modulated by the deregulated miRNA that we had identified. To confirm in silico analysis, expression levels of genes encoding for glycosyltransferases and glycan-processing enzymes were investigated using Human Glycosylation-RT2 Profiler PCR Array. Among 754 miRNA analyzed, we identified 126 miRNA that were significantly deregulated in pSS compared to controls, with a trend that was inversely proportional with the impairment of salivary flow rates. An in silico approach pinpointed that several upregulated miRNA in patients with pSS target important genes in the mucin O-glycosylation. We confirmed this prediction by quantitative real-time PCR, highlighting the downregulation of some glycosyltransferase and glycosidase genes in pSS samples compared to controls, such as GALNT1, responsible for mucin-7 glycosylation. Collectively, our data suggest that the expression of different predicted miRNA-target genes in the mucin type O-glycan biosynthesis pathway is altered in pSS patients with low salivary flow and that the miRNA expression profile could influence the glycosidase expression levels and consequently the rheology in pSS.

Differential expression patterns of specific long noncoding RNAs and competing endogenous RNA network in alopecia areata.

Long noncoding RNAs (lncRNAs) regulate gene expression by acting with microRNAs (miRNAs) and indirectly interact with messenger RNA (mRNAs). However, the roles of specific lncRNA and its related competing endogenous RNAs (ceRNA) network in alopecia areata (AA) are not fully understood. The blood lncRNA profiles were obtained by microarray from 10 samples, including five alopecia areata samples and five normal samples. Based on bioinformatics generated from miRcode, starBase, and miRTarBase, we constructed an lncRNA-miRNA-mRNA network (ceRNA network) in alopecia areata. We found 154 differentially expressed lncRNAs and 46 differentially expressed genes (DEGs). The functional enrichment indicated that the DEGs mainly regulated the pathways of focal adhesion, Mucin type O-glycan biosynthesis, and so on. The differentially expressed lncRNA (DElncRNA) involved in the pathway of thyronamine and iodothyronamine metabolism and so on. Through integrated lncRNA-mRNA and miRNA-mRNA pairs, the ceRNA network was constructed, thereafter, six ceRNA subnetworks were identified and subnetwork 1 were found to be significantly associated with the occurrence of alopecia areata. Our results showed blood lncRNA expression patterns and a complex ceRNA network in alopecia areata. However, futher studies on blood and tissue verification of these lncRNAs and relative pathways are needed.

MicroRNAs regulating mucin type O-glycan biosynthesis and transforming growth factor β signaling pathways in nasal mucosa of patients with chronic rhinosinusitis with nasal polyps in Northern China.

Chronic rhinosinusitis with nasal polyps (CRSwNP) is a persistent sinonasal mucosa inflammatory disease. MicroRNAs (miRNAs) that are involved in the pathogenesis of CRSwNP in Northern China remain unknown. A miRCURY™ LNA Array was used to analyze miRNA profiles in nasal mucosa tissues of CRSwNP patients (n = 19) and healthy controls (n = 10). Subsequent pathways were predicted by DIANA-mirPath software. Five upregulated miRNAs, including miR-210-5p, miR-3178, miR-585-3p, miR-3146, and miR-320e, and 19 downregulated miRNAs, including miR-32-3p, miR-1299, miR-3196, miR-3924, and miR-548e-3p, were differentially expressed (p < 0.05, fold change >2) in tissues of CRSwNP vs controls. Utilizing the Kyoto Encyclopedia of Genes and Genomes database (KEGG), which is an online database for pathway mapping, mucin type O-glycan biosynthesis pathway was significantly enriched in upregulated miRNAs. Transforming growth factor-beta (TGF-β), transient receptor potential (TRP) channels, and the mitogen-activated protein kinase (MAPK) signaling pathway were significantly linked to downregulated miRNAs. The mucin type O-glycan biosynthesis pathway and TGF-β signaling pathway are regulated by miRNAs, which could be our focus in the future studies.

T-Synthase Deficiency Enhances Oncogenic Features in Human Colorectal Cancer Cells via Activation of Epithelial-Mesenchymal Transition.

Background Immature truncated O-glycans such as Tn antigen are frequently detected in human colorectal cancer (CRC); however, the precise pathological consequences of Tn antigen expression on CRC are unknown. T-synthase is the key enzyme required for biosynthesis of mature O-glycans. Here we investigated the functional roles of Tn antigen expression mediated by T-synthase deficiency in CRC cells. Methods To knock out T-synthase, we used CRISPR-Cas9 technology to target C1GALT1, the gene encoding T-synthase, in a CRC cell line (HCT116). Deletion of T-synthase was confirmed by western blotting, and expression of Tn antigen was determined by flow cytometry in HCT116 cells. We then assessed the biological effects of T-synthase deficiency on oncogenic behaviors in HCT116 cells. Furthermore, we analyzed the mechanistic role of T-synthase deficiency in cancer cells by determining the epithelial-mesenchymal transition (EMT) pathway. Results We showed that forced knockout of T-synthase in HCT116 cells significantly induced Tn antigen expression, which represented the occurrence of aberrant O-glycosylation. Loss of T-synthase significantly enhanced cell proliferation and adhesion, as well as migration and invasiveness in culture. More importantly, we demonstrated that T-synthase deficiency directly induced classical EMT characteristics in cancer cells. E-cadherin, a typical epithelial cell marker, was markedly decreased in T-synthase knockout HCT 116 cells, accompanied by an enhanced expression of mesenchymal markers including snail and fibronectin (FN). Conclusions These findings indicate that T-synthase deficiency in CRC cells not only is responsible for aberrant O-glycosylation, but also triggers the molecular process of EMT pathway, which may translate to increased invasiveness and metastasis in cancers.

Glycosylation pathways at the ocular surface.

Glycosylation is a major form of enzymatic modification of organic molecules responsible for multiple biological processes in an organism. The biosynthesis of glycans is controlled by a series of glycosyltransferases, glycosidases and glycan-modifying enzymes that collectively assemble and process monosaccharide moieties into a diverse array of structures. Many studies have provided insight into various pathways of glycosylation at the ocular surface, such as those related to the biosynthesis of mucin-type O-glycans and N-glycans on proteins, but many others still remain largely unknown. This review provides an overview of the different classes of glycans described at the ocular surface focusing on their biosynthetic pathways and biological relevance. A precise understanding of these pathways under physiological and pathological conditions could help identify biomarkers and novel targets for therapeutic intervention.

Role of sialic acid-containing glycans of matrix metalloproteinase-9 (MMP-9) in the interaction between MMP-9 and staphylococcal superantigen-like protein 5.

Staphylococcal superantigen-like proteins (SSL) show no superantigenic activity but have recently been considered to act as immune suppressors. It was previously reported that SSL5 bound to P-selectin glycoprotein ligand-1 (PSGL-1) and matrix metalloproteinase (MMP)-9, leading to inhibition of leukocyte adhesion and invasion. These interactions were suggested to depend on sialic acid-containing glycans of MMP-9, but the roles of sialic acids in the interaction between SSL5 and MMP-9 are still controversial. In the present study, we prepared recombinant glutathione S-transferase-tagged SSL5 (GST-SSL5) and analyzed its binding capacity to MMP-9 by pull-down assay after various modifications of its carbohydrate moieties. We observed that GST-SSL5 specifically bound to MMP-9 from a human monocytic leukemia cell line (THP-1 cells) and inhibited its enzymatic activity in a concentration-dependent manner. After MMP-9 was treated with neuraminidase, its binding activity towards GST-SSL5 was markedly decreased. Furthermore, recombinant MMP-9 produced by sialic acid-deficient Lec2 mutant cells showed much lower affinity for SSL5 than that produced by wild-type CHO-K1 cells. Treatment of MMP-9 with PNGase F to remove N-glycan resulted in no significant change in the GST-SSL5/MMP-9 interaction. In contrast, the binding of GST-SSL5 to MMP-9 secreted from THP-1 cells cultured in the presence of an inhibitor for the biosynthesis of O-glycan (benzyl-GalNAc) was weaker than the binding of GST-SSL5 to MMP-9 secreted from untreated cells. These results strongly suggest the importance of the sialic acid-containing O-glycans of MMP-9 for the interaction of MMP-9 with GST-SSL5.

Revealing the action mechanisms of dexamethasone on the birth weight of infant using RNA-sequencing data of trophoblast cells.

Dexamethasone (DEX) could induce low birth weight of infant, and low birth weight has close associations with glucocorticoid levels, insulin resistance, hypertension, and metabolic syndrome in adulthood. This study was designed to reveal the action mechanisms of DEX on the birth weight of infant.Using quantitative real-time polymerase chain reaction (qRT-PCR), trophoblast cells of human placenta were identified and the optimum treatment time of DEX were determined. Trophoblast cells were treated by DEX (DEX group) or ethanol (control group) (each group had 3 samples), and then were performed with RNA-sequencing. Afterward, the differentially expressed genes (DEGs) were identified by R package, and their potential functions were successively enriched using DAVID database and Enrichr method. Followed by protein–protein interaction (PPI) network was constructed using Cytoscape software. Using Enrichr method and TargetScan software, the transcription factors (TFs) and micorRNAs (miRNAs) targeted the DEGs separately were predicted. Based on MsigDB database, gene set enrichment analysis (GSEA) was performed.There were 391 DEGs screened from the DEX group. Upregulated SRR and potassium voltage-gated channel subfamily J member 4 (KCNJ4) and downregulated GALNT1 separately were enriched in PDZ (an acronym of PSD-95, Dlg, and ZO-1) domain binding and Mucin type O-glycan biosynthesis. In the PPI network, CDK2 and CDK4 had higher degrees. TFs ATF2 and E2F4 and miRNA miR-16 were predicted for the DEGs. Moreover, qRT-PCR analysis confirmed that SRR and KCNJ4 were significantly upregulated.These genes might affect the roles of DEX in the birth weight of infant, and might be promising therapeutic targets for reducing the side effects of DEX.

Expression Signature and Role of miR-30d-5p in Non-Small Cell Lung Cancer: a Comprehensive Study Based on in Silico Analysis of Public Databases and in Vitro Experiments

Background/Aims: The purpose of this study was to probe the clinico-pathological significance and the underlying mechanism of miR-30d-5p expression in non-small cell lung cancer (NSCLC). Methods: We initially examined the level of miR-30d-5p expression in NSCLC and non-cancer tissues using RT-qPCR. Then, a series of validation analyses including a meta-analysis of data from microarray chips in Gene Expression Omnibus (GEO), data mining of the cancer genome atlas (TCGA) and an integrated meta-analysis incorporating GEO microarray chips, TCGA data, in-house RT-qPCR and literature studies were performed to examine the clinico-pathological value of miR-30d-5p expression in NSCLC. In vitro experiments were further conducted to investigate the impact of miR-30d-5p on NSCLC cell growth. The molecular mechanism by which miR-30d-5p regulates the pathogenesis of NSCLC was probed through a bioinformatics analysis of its target genes. Moreover, dual luciferase reporter assay was conducted to verify the targeting regulatory relationship between miR-30d-5p and CCNE2. Results: Based on results from RT-qPCR, GEO meta-analysis, TCGA data mining and the integrated meta-analysis incorporating GEO microarray chips, TCGA data, in-house RT-qPCR and literature studies, miR-30d-5p expression was decreased in NSCLC tissues, and patients with NSCLC who presented with lower miR-30d-5p expression tended to display an advanced clinical progression. Significant pathways including the Mucin type O-glycan biosynthesis pathway, cell cycle pathway and cysteine and methionine metabolism pathway (all P< 0.05) revealed potential roles of the target genes of miR-30d-5p in the oncogenesis of NSCLC. Results from in vitro experiments indicated that miR-30d-5p could attenuate proliferation and viability of NSCLC cells. Among the 12 identified hub genes, nine genes including E2F3, CCNE2, SKP2, CDK6, TFDP1, LDHA, GOT2, DNMT3B and ST6GALNAC1 were validated by Pearson’s correlation test and the human protein atlas (HPA) database as targets of miR-30d-5p with higher probability. Specifically, dual luciferase reporter assay confirmed that CCNE2 was directly targeted by miR-30d-5p. Conclusion: In summary, miR-30d-5p expression is decreased in NSCLC, and it might play the role as tumor suppressor in NSCLC by regulating target genes.

Altered O-glycosylation is associated with inherent radioresistance and malignancy of human laryngeal carcinoma

Radioresistance (inherent or acquired) remains a major obstacle affecting the clinical outcome of radiotherapy for laryngeal carcinoma. Results from our laboratory and other groups suggest that aberrant glycosylation contributes to cancer acquired radioresistance. However, the role of glycosylation in inherent radioresistance of laryngeal carcinoma has not been fully uncovered. In this study, we investigated the glycan profiling of the inherent radioresistant (Hep-2max) and radiosensitive (Hep-2min) cell lines using lectin microarray analysis. The results revealed that the radioresistant cell line Hep-2max presented higher core 1-type O-glycans than the sensitive one. Further analysis of the O-glycan regulation by benzyl-α-GalNAc application in Hep-2max cells showed partial inhibition of the O-glycan biosynthesis and increased radiosensitivity. In addition, core 1 β1, 3-galactosyltransferase (C1GALT1) overexpression in Hep-2min cells enhanced cell migration, invasion, and radioresistance. Conversely, knockdown of C1GALT1 in Hep-2max cells was able to suppress these malignant phenotypes. Moreover, mechanistic investigations showed that C1GALT1 modified the O-glycans on integrin β1 and regulated its activity. The glycosylation-mediated radioresistance was further inhibited by anti-integrin β1 blocking antibody. Importantly, we also observed that core 1-type O-glycans expression was correlated with advanced tumor stage, metastasis, and poor survival of laryngeal carcinoma patients. These findings suggest that altered O-glycosylation can lead to the inherent radioresistance and progression, and therefore may be important for enhancing the efficacy of radiotherapy in laryngeal carcinoma.