Expression Microarray Analysis Research Articles

A novel orexin antagonist from a natural plant was discovered using zebrafish behavioural analysis.

Phenotypic screening is one of the most practical approaches to the identification of mediators of behaviour, since it is difficult to model brain function in vitro, at a cellular level. We used a zebrafish (Danio rerio) behavioural assay to discover novel, natural, neuroactive compounds. A zebrafish behavioural assay was performed for seven natural compounds, obtained from plants. The behavioural profiles were compared to those of known psychoactive drugs. We characterised a natural compound exhibiting a behaviour profile similar to that of suvorexant, using in silico, in vitro and microarray expression analysis. The behavioural analysis performed in this study classified central nervous system drugs according to their mechanism. Zebrafish treated with a natural compound, 8b-(4'-Hydroxytigloyloxy) costunolide (8b), showed behaviour profiles similar to those of zebrafish treated with suvorexant, a known orexin antagonist. This behavioural assay was validated using in silico and in vitro assays, which revealed that the new compound was a dual orexin receptor antagonist. In addition, transcriptome analysis suggested that 8b might regulate the nuclear factor-κB (NF-κB) related pathway. We conclude that zebrafish phenotypic screening, combined with in silico assays and gene expression profiling, is a useful strategy to discover and characterize novel therapeutic compounds, including natural products.

Screening differentially expressed genes of pancreatic cancer between Mongolian and Han people using bioinformatics technology

BackgroundTo screen and analyze differentially expressed genes in pancreatic carcinoma tissues taken from Mongolian and Han patients by Affymetrix Genechip. Methods: Pancreatic ductal cell carcinoma tissues were collected from the Mongolian and Han patients undergoing resection in the Second Affiliated Hospital of Nanchang University from March 2015 to May 2018 and the total RNA was extracted. Differentially expressed genes were selected from the total RNA qualified by Nanodrop 2000 and Agilent 2100 using Affymetrix and a cartogram was drawn; The gene ontology (GO) analysis and Pathway analysis were used for the collection and analysis of biological information of these differentially expressed genes. Finally, some differentially expressed genes were verified by real-time PCR.ResultsThrough the microarray analysis of gene expression, 970 differentially expressed genes were detected by comparing pancreatic cancer tissue samples between Mongolian and Han patients. A total of 257 genes were significantly up-regulated in pancreatic cancer tissue samples in Mongolian patients; while a total of 713 genes were down-regulated. In the Gene Ontology database, 815 differentially expressed genes were identified with clear GO classification, and CPB1 gene showed the highest increase in expression level (multiple difference: 31.76). The pathway analysis detected 28 signaling pathways that included these differentially expressed genes, involving a total of 178 genes. Among these pathways, the enrichment of differentially expressed genes in the FAK signaling pathway was the strongest and COL11A1 gene showed the highest multiple difference (multiple difference: 5.02). The expression of differentially expressed genes CPB1, COL11A1、ITGA4、BIRC3、PAK4、CPA1、CLPS、PIK3CG and HLA-DPA1 determined by real-time PCR were consistent with the results of gene microarray analysis.ConclusionsThe results of microarray analysis of gene expression profiles showed that there are a large number of differentially expressed genes in pancreatic cancer tissue samples comparing Mongolian and Han population. These genes are closely related to the cell proliferation, differentiation, invasion, metastasis and multi-drug resistance in pancreatic cancer. They are also involved in the regulation of multiple important signaling pathways in organisms.

SPRY4 regulates trophoblast proliferation and apoptosis via regulating IFN-γ-induced STAT1 expression and activation in recurrent miscarriage.

The dysregulation of trophoblast functions is one of the leading causes of recurrent miscarriage (RM), which frustrates 1%-5% of couples of childbearing ages. Sprouty 4 (SPRY4) is considered as a tumour suppressor and exerts a negative role in cell viability. However, its role in regulating trophoblast behaviors at the maternal-fetal interface remains largely unknown. First-trimester villous samples were collected from RM patients and healthy controls (HCs) to determine the SPRY4 expression in human placenta during early pregnancy. The HTR8/SVneo cell line was introduced to clarify trophoblast cell functions via transfecting with specific short interfering RNA against SPRY4 or SPRY4-overexpressing lentivirus in vitro. In addition, gene expression microarray analysis was performed to explore the downstream molecules and pathways. Our results revealed that SPRY4 expression was significantly increased in the first-trimester cytotrophoblasts of RM patients compared with HCs. Furthermore, SPRY4 overexpression inhibited trophoblast proliferation and accelerated apoptosis in vitro, while SPRY4 knockdown reversed these effects. Mechanistically, IFN-γ -induced STAT1 expression and activation were involved in the regulation of trophoblast proliferation and apoptosis by SPRY4, and IFN-γ promoted SPRY4 expression and STAT1 phosphorylation through PI3K/AKT pathway. Additionally, both STAT1 and phosphorylated STAT (p-STAT) levels were also upregulated in trophoblasts from RM patients and positively correlated with SPRY4 expression. Our findings indicate that SPRY4 may act as a negative regulator of trophoblast functions through upregulating IFN-γ/PI3K/AKT-induced STAT1 activation. High levels of SPRY4 and STAT1 may contribute to RM development and progression, and blocking of either target could be a novel therapeutic strategy for RM patients.

Identification of differential expression genes, pathways and potential biomarkers for osteoarthritis using an integrative gene expression microarray analysis

Identification of differential expression genes, pathways and potential biomarkers for osteoarthritis using an integrative gene expression microarray analysis

Galectin-1 Overexpression Activates the FAK/PI3K/AKT/mTOR Pathway and Is Correlated with Upper Urinary Urothelial Carcinoma Progression and Survival.

Galectin-1 (GAL1) is a β-galactoside-binding protein involved in multiple aspects of tumorigenesis. However, the biological role of GAL1 in upper tract urothelial carcinoma (UTUC) has not been entirely understood. Herein, we investigated the oncological effects of GAL1 expression in tumor specimens and identified related gene alterations through molecular analysis of GAL1. Clinical parameter data and tumor specimens were collected from 86 patients with pT3N0M0 UTUC who had undergone radical nephroureterectomy. We analyzed the difference in survival by using Kaplan–Meier analyses and Cox proportional regression models and in GAL1 expression by using immunohistochemical (IHC) methods. Public genomic data from the Cancer Genome Atlas (TCGA) and GSE32894 data sets were analyzed for comparison. Using four urothelial carcinoma (UC) cell lines (BFTC-909, T24, RT4, and J82) as in vitro models, we evaluated the functions of GAL1 in UC cell growth, invasiveness, and migration and its role in downstream signaling pathways. The study population was classified into two groups, GAL1-high (n = 35) and GAL1-low (GAL1 n = 51), according to IHC interpretation. Univariate analysis revealed that high GAL1 expression was significantly associated with poor recurrence-free survival (RFS; p = 0.028) and low cancer-specific survival (CSS; p = 0.025). Multivariate analysis revealed that GAL1-high was an independent predictive factor for RFS (hazard ratio (HR) 2.43; 95% confidence interval (CI) 1.17–5.05, p = 0.018) and CSS (HR 4.04; 95% CI 1.25–13.03, p = 0.019). In vitro studies revealed that GAL1 knockdown significantly reduced migration and invasiveness in UTUC (BFTC-909) and bladder cancer cells (T24). GAL1 knockdown significantly reduced protein levels of matrix metalloproteinase-2 (MMP-2) and MMP-9, which increased tissue inhibitor of metalloproteinase-1 (TIMP-1) and promoted epithelial–mesenchymal transition (EMT). Through gene expression microarray analysis of GAL1 vector and GAL1-KD cells, we identified multiple significant signaling pathways including p53, Forkhead box O (FOXO), and phosphoinositide 3-kinase/protein kinase B (PI3K/AKT). We validated microarray results through immunoblotting, thus proving that downregulation of GAL1 reduced focal adhesion kinase (FAK), p-PI3K, p-AKT, and p-mTOR expression. We concluded that GAL1 expression was highly related to oncological survival in patients with locally advanced UTUC. GAL1 promoted UC invasion and metastasis by activating the FAK/PI3K/AKT/mTOR pathway.

Downregulation of histone methyltransferase SET8 inhibits progression of hepatocellular carcinoma

The expression of lysine methyltransferase SET8, which is involved in carcinogenesis of many types of human cancers through monomethylation of histone H4 lysine 20 (H4K20), is associated with the prognosis of hepatocellular carcinoma (HCC). We performed a functional analysis for SET8 to assess its effect on HCC progression. SET8 knockdown inhibited proliferation, migration and invasion of HCC cells. SET8 knockdown also inhibited tumour growth in a human xenograft mouse model. Overexpression of SET8 displayed the reverse effect, while treatment with the SET8 inhibitor UNC0379 produced an effect similar to SET8 knockdown. In addition, drug sensitivity testing in SET8-siRNA transfected HCC cells indicated that docetaxel inhibited cell growth dramatically, as demonstrated by the Cell Counting Kit-8 (CCK-8) assay. Furthermore, gene expression microarray analysis showed that genes altered after SET8 knockdown were clustered in pathways related to tumorigenesis and metastasis. Our data suggests that targeting SET8 for HCC therapy can inhibit the proliferation and invasion of HCC cells as well as increase their sensitivity to chemotherapy.

Identification of a robust functional subpathway signature for pancreatic ductal adenocarcinoma by comprehensive and integrated analyses

BackgroundPancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy and its mortality continues to rise globally. Because of its high heterogeneity and complex molecular landscapes, published gene signatures have demonstrated low specificity and robustness. Functional signatures containing a group of genes involved in similar biological functions may display a more robust performance.MethodsThe present study was designed to excavate potential functional signatures for PDAC by analyzing maximal number of datasets extracted from available databases with a recently developed method of FAIME (Functional Analysis of Individual Microarray Expression) in a comprehensive and integrated way.ResultsEleven PDAC datasets were extracted from GEO, ICGC and TCGA databases. By systemically analyzing these datasets, we identified a robust functional signature of subpathway (path:00982_1), which belongs to the drug metabolism-cytochrome P450 pathway. The signature has displayed a more powerful and robust capacity in predicting prognosis, drug response and chemotherapeutic efficacy for PDAC, particularly for the classical subtype, in comparison with published gene signatures and clinically used TNM staging system. This signature was verified by meta-analyses and validated in available cell line and clinical datasets with chemotherapeutic efficacy.ConclusionThe present study has identified a novel functional PDAC signature, which has the potential to improve the current systems for predicting the prognosis and monitoring drug response, and to serve a linkage to therapeutic options for combating PDAC. However, the involvement of path:00982_1 subpathway in the metabolism of anti-PDAC chemotherapeutic drugs, particularly its biological interpretation, requires a further investigation.55T42nE1wBcMSNQpJ96fRFVideo

Inter-Individual Variation in Response to Estrogen in Human Breast Explants.

Exposure to estrogen is strongly associated with increased breast cancer risk. While all women are exposed to estrogen, only 12% are expected to develop breast cancer during their lifetime. These women may be more sensitive to estrogen, as rodent models have demonstrated variability in estrogen sensitivity. Our objective was to determine individual variation in expression of estrogen receptor (ER) and estrogen-induced responses in the normal human breast. Human breast tissue from female donors undergoing reduction mammoplasty surgery were collected for microarray analysis of ER expression. To examine estrogen-induced responses, breast tissue from 23 female donors were cultured ex- vivo in basal or 10nM 17β-estradiol (E2) media for 4days. Expression of ER genes (ESR1 and ESR2) increased significantly with age. E2 induced consistent increases in global gene transcription, but expression of target genes AREG, PGR, and TGFβ2 increased significantly only in explants from nulliparous women. E2-treatment did not induce consistent changes in proliferation or radiation induced apoptosis. Responses to estrogen are highly variable among women and not associated with levels of ER expression, suggesting differences in intracellular signaling among individuals. The differences in sensitivity to E2-stimulated responses may contribute to variation in risk of breast cancer.

MiR-23b-3p acts as a counter-response against skeletal muscle atrophy.

To investigate the role of microRNA (miRNA) in muscle atrophy, we performed microarray analysis of miRNA expression in skeletal muscles of Sham, orchiectomized (ORX) mice, and ORX mice treated with androgen and identified that the expression of miR-23b-3p in ORX mice was significantly higher than that in Sham mice (P = 0.007); however, miR-23b-3p expression in ORX mice treated with androgen was lower (P = 0.001). We also investigated the mechanism by which overexpression or knockdown of miR-23b-3p influences the expression of myosin heavy chain, muscle protein synthesis, ATP activity, and glucose uptake in C2C12 myotube cells. Moreover, we examined the serum miR-23b-3p levels among male subjects with type 2 diabetes and whether the serum miR-23b-3p levels could be a biomarker for muscle atrophy. The overexpression of miR-23b-3p in C2C12 myotube cells significantly upregulated the expression of myosin heavy chain, protein synthesis, ATP activity, and glucose uptake. Reporter assays raised a possible direct post-transcriptional regulation involving miR-23b-3p and the 3'-UTR of PTEN mRNA. Among subjects with type 2 diabetes, serum miR-23b-3p levels in the subjects with decreased muscle mass were significantly higher compared to the levels in the subjects without. Our results indicate that miR-23b-3p downregulates the expression of PTEN in myotube cells and induces the growth of myosin heavy chain. In addition, the serum level of miR-23b-3p can be used as a diagnostic marker for muscle atrophy.

Nuclear accumulation of MKL1 in luminal breast cancer cells impairs genomic activity of ERα and is associated with endocrine resistance

Estrogen receptor (ERα) is central in driving the development of hormone-dependent breast cancers. A major challenge in treating these cancers is to understand and overcome endocrine resistance. The Megakaryoblastic Leukemia 1 (MKL1, MRTFA) protein is a master regulator of actin dynamic and cellular motile functions, whose nuclear translocation favors epithelial-mesenchymal transition. We previously demonstrated that nuclear accumulation of MKL1 in estrogen-responsive breast cancer cell lines promotes hormonal escape. In the present study, we confirm through tissue microarray analysis that nuclear immunostaining of MKL1 is associated with endocrine resistance in a cohort of breast cancers and we decipher the underlining mechanisms using cell line models. We show through gene expression microarray analysis that the nuclear accumulation of MKL1 induces dedifferentiation leading to a mixed luminal/basal phenotype and suppresses estrogen-mediated control of gene expression. Chromatin immunoprecipitation of DNA coupled to high-throughput sequencing (ChIP-Seq) shows a profound reprogramming in ERα cistrome associated with a massive loss of ERα binding sites (ERBSs) generally associated with lower ERα-binding levels. Novel ERBSs appear to be associated with EGF and RAS signaling pathways. Collectively, these results highlight a major role of MKL1 in the loss of ERα transcriptional activity observed in certain cases of endocrine resistances, thereby contributing to breast tumor cells malignancy.

Altered gene expression in CHO cells following polyamine starvation.

To investigate the impact of polyamine deprivation on the transcriptome of CHO cells RESULTS: Polyamines play a central but poorly-understood role in cell proliferation. Most studies to date have utilised chemical inhibitors to probe polyamine function. Here we exploit the fact that CHO cells grown in serum-free medium have an absolute requirement for putrescine supplementation due to their deficiency in activity of the enzyme arginase. A gene expression microarray (Affymetrix) analysis of CHO-K1 cells starved of polyamines for 3days showed that cessation of growth, associated with increased G1/S transition and inhibition ofM/G1 transitionwas accompanied by increased mRNA levels of mitotic complex checkpoint genes (Mad2l1, Tkk, Bub1b) and in the transition of G1- to S-phase (such as Skp2 and Tfdp1). mRNAs associated with DNA homologous recombination and repair (including Fanconi's anaemia-related genes) and with RNA splicing were consistently increased. Alterations in mRNA levels for genes related to protein processing in the ER, to ER stress, and to p53-related and apoptosis pathways were also observed. mRNAs showing highest levels of fold-change included several which code for membrane-localised proteins and receptors (Thbs1, Tfrc1, Ackr3, Extl1). Growth-arrest induced by polyamine deprivation was associated with significant alterations in levels of mRNAs associated with cell cycle progression, DNA repair, RNA splicing, ER trafficking and membrane signalling as well as p53 and apoptosis-related pathways.

DNMT3A mutants provide proliferating advantage with augmentation of self-renewal activity in the pathogenesis of AML in KMT2A-PTD-positive leukemic cells

Acute myeloid leukemia (AML) with partial tandem duplication of histone-lysine N-methyltransferase 2A (KMT2A-PTD) is a subtype of AML and is associated with adverse survival, yet the molecular pathogenesis of KMT2A-PTD is not fully understood. DNA methyltransferase 3A (DNMT3A) is mutated in various myeloid neoplasms including AML, especially at the Arg882. Recently, it has been found that DNMT3A mutations frequently coexisted with KMT2A-PTD and are associated with inferior outcomes. We aimed to understand the biological role of DNMT3A mutation in KMT2A-PTD-positive cells. Herein, we found that overexpression of DNMT3A mutants (MT) in KMT2A-PTD-positive EOL-1 cells augmented cell proliferation and clonogenicity. Serial colony replating assays indicated that DNMT3A-MT increased the self-renewal ability of Kmt2a-PTD-expressing mouse bone marrow cells with immature morphology. At 10 months post bone marrow transplantation, mice with the combined Kmt2a-PTD and DNMT3A-MT showed hepatosplenomegaly and leukocytosis with a shorter latency compared to control and DNMT3A-wild-type. Gene expression microarray analyses of bone marrow samples from human AML with KMT2A-PTD/DNMT3A-MT showed a stem cell signature and myeloid hematopoietic lineage with dysregulation of HOXB gene expression. In addition, human bone marrow AML cells carrying KMT2A-PTD/DNMT3A-MT showed abnormal growth and augmented self-renewal activity in primary cell culture. The present study provides information underlying the pathogenic role of DNMT3A-MT with KMT2A-PTD in proliferating advantage with augmentation of self-renewal activity in human leukemia, which may help to better understand the disease and to design better therapy for AML patients with these mutations.

Bitter melon fruit extract affects hepatic expression of the genes involved in inflammation and fatty acid metabolism in ob/ob mice

Background: The fruit of bitter melon (Momordica charantia L.) has traditionally been used as a crude drug to treat obesity and diabetes mellitus. Previously, we reported that an ethyl acetate (EtOAc)-soluble fraction of bitter melon fruit extract showed anti-inflammatory effects on interleukin-1β-treated rat hepatocytes. The liver plays a pivotal role to maintain homeostasis of glucose and lipid. It is speculated that the administration of an EtOAc-soluble fraction of a bitter melon fruit extract (BMFA) may change the expression profile of hepatic genes involved in inflammation, glucose metabolism, and lipid metabolism.Objective: We aimed to evaluate the effects of BMFA on the gene expression in the liver of ob/ob mice, which are deficient in leptin and are used as a diabetes mellitus model. The changes may be related to hepatic steatosis and obesity.Methods: Male 8-week-old ob/ob mice (average body weight, 40.9 g) were fed a standard diet with and without BMFA (ob/ob+BMFA and ob/ob–BMFA groups, respectively; 4 mice/group) for seven days. The livers were excised to prepare total RNA for microarray expression analyses. Signal ratios (i.e., ob/ob+BMFA versus ob/ob–BMFA group) were calculated to compare gene expression.Results: The mRNA levels of 2,632 genes were significantly changed in the livers of ob/ob+BMFA mice when compared with those of ob/ob–BMFA mice. As expected, proinflammatory cytokine and chemokine genes were significantly downregulated by BMFA. The acyl-Coenzyme A thioesterase 3 gene in fatty acid oxidation was upregulated in ob/ob+BMFA liver, whereas free fatty acid receptor 2 and solute carrier family 27 member 3 genes were downregulated. Most genes involved in glycolysis and gluconeogenesis were not affected by BMFA. In contrast, the level of peroxisome proliferator-activated receptor γ coactivator 1α mRNA significantly increased in ob/ob+BMFA mice, suggesting that BMFA administration may reduce insulin resistance in ob/ob mice.Conclusion: Our study suggested that BMFA may possess anti-inflammatory effects and enhance fatty acid metabolism in ob/ob mouse liver. These data imply the possibility that EtOAc-soluble constituents in bitter melon fruit may improve lipid accumulation in the liver during obesity and diabetes mellitus, as well as non-alcoholic fatty liver disease.Keywords: Momordica charantia, gene expression, inflammation, diabetes mellitus, non-alcoholic fatty liver disease.

Elevated Expression of Lumican in Lung Cancer Cells Promotes Bone Metastasis through an Autocrine Regulatory Mechanism

Background: The microarray analysis of whole-genome expression indicated that the gene encoding the protein lumican, which is associated with extracellular matrix (ECM) interaction, was highly expressed in osteotropic lung cancer cell lines with an enhanced capacity of bone metastasis. Methods: The expression of lumican in the osteotropic lung cancer cells was downregulated, and the in vitro migration, invasion, and adhesion of cancer cells to ECM components, and the in vivo bone metastasis capacity of these cells were examined. Exogenous lumican was provided to study the autocrine regulation mechanism of lumican in the bone metastasis of lung cancer cells. Results: Transfection with lumican-specific short hairpin RNA (shRNA) in the osteotropic lung cancer cells reduced the establishment of in vivo bone metastasis, but not lung metastasis. Reduction in the expression of lumican also decreased the attachment of lung osteotropic cancer cells to several components of the ECM and suppressed cell migration and invasion in vitro. Exogenous lumican restored these reduced capacities of lumican knockdown cells and promoted the seeding of lung cancer cells in the bone microenvironment. Conclusions: These results suggested that lumican promotes the metastasis of lung cancer cells to the bones via an autocrine regulatory mechanism, and blocking this interaction may provide a new therapeutic approach to reduce bone metastasis in cases of lung cancer.

Generation and characterization of a mitotane-resistant adrenocortical cell line.

Mitotane is the only drug approved for the therapy of adrenocortical carcinoma (ACC). Its clinical use is limited by the occurrence of relapse during therapy. To investigate the underlying mechanisms in vitro, we here generated mitotane-resistant cell lines. After long-term pulsed treatment of HAC-15 human adrenocortical carcinoma cells with 70 µM mitotane, we isolated monoclonal cell populations of treated cells and controls and assessed their respective mitotane sensitivities by MTT assay. We performed exome sequencing and electron microscopy, conducted gene expression microarray analysis and determined intracellular lipid concentrations in the presence and absence of mitotane. Clonal cell lines established after pulsed treatment were resistant to mitotane (IC50 of 102.2 ± 7.3 µM (n = 12) vs 39.4 ± 6.2 µM (n = 6) in controls (biological replicates, mean ± s.d., P = 0.0001)). Unlike nonresistant clones, resistant clones maintained normal mitochondrial and nucleolar morphology during mitotane treatment. Resistant clones largely shared structural and single nucleotide variants, suggesting a common cell of origin. Resistance depended, in part, on extracellular lipoproteins and was associated with alterations in intracellular lipid homeostasis, including levels of free cholesterol, as well as decreased steroid production. By gene expression analysis, resistant cells showed profound alterations in pathways including steroid metabolism and transport, apoptosis, cell growth and Wnt signaling. These studies establish an in vitro model of mitotane resistance in ACC and point to underlying molecular mechanisms. They may enable future studies to overcome resistance in vitro and improve ACC treatment in vivo.

Comparative Expression Profile Analysis of Apoptosis-Related miRNA and Its Target Gene in Leishmania major Infected Macrophages.

Background:Cutaneous Leishmaniasis (CL) is an emerging uncontrollable and neglected infectious disease worldwide including Iran. The aim of this study was to investigate the expression profile of apoptosis-related miRNA and its target gene in macrophages.Methods:This study was carried out in the Department of Medical Parasitology and Mycology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran from January 2016 to November 2018. Applying literature reviews, bioinformatics software, and microarray expression analysis, we selected miRNA-24-3p interfering in apoptosis pathway. The expression profile of this miRNA and target gene were investigated in Leishmania major (MRHO/IR/75/ER)-infected primary and RAW 264.7 macrophages (IBRC-C10072) compared with non-infected macrophages (control group) using quantitative Real-time PCR.Results:Results of bioinformatics analysis showed that miR-24-3p as anti-apoptotic miRNA inhibits pro-apoptotic genes (Caspases 3 and 7). Microarray expression data presented in Gene Expression Omnibus (GEO) revealed a significant difference in the expression level of selected miRNA and its target gene between two groups. QRT-PCR results showed that the expression of miR-24-3p was upregulated in L. major infectioned macrophages that approved the results of bioinformatics and microarray analysis.Conclusion:Parasite can alter miRNAs expression pattern in the host cells to establish infection and its survival. Alteration in miRNAs levels likely plays an important role in regulating macrophage functions following L. major infection. These results could highlight current understanding and new insights concerning the gene expression in macrophages during leishmaniasis and will help to development of novel strategies for control and treatment of CL.

LAMC1 is a prognostic factor and a potential therapeutic target in endometrial cancer.

ObjectiveWith the emerging significance of genetic profiles in the management of endometrial cancer, the identification of tumor-driving genes with prognostic value is a pressing need. The LAMC1 gene, encoding the laminin subunit gamma 1 (LAMC1) protein, has been reported to be involved in the progression of various malignant tumors. In this study, we aimed to investigate the role of LAMC1 in endometrial cancer and elucidate the underlying mechanism.MethodsWe evaluated the immunohistochemical expression of LAMC1 in atypical endometrial hyperplasia and endometrial cancer. Within the endometrial cancer cases, we analyzed the association of LAMC1 overexpression with clinicopathological factors and prognosis. Furthermore, to indentify genes influenced by LAMC1 overexpression, we transfected HEC50B and SPAC-S cells with siRNA targeting LAMC1 and conducted microarray gene expression assays.ResultsWhile none of the atypical endometrial hyperplasia specimens exhibited LAMC1 overexpression, endometrial cancer possessed a significantly higher LAMC1 overexpression rate. LAMC1 overexpression was strongly associated with histological type, lymphovascular space invasion, lymph node metastasis, advanced International Federation of Gynecology and Obstetrics stage, and poor overall survival in endometrial cancer. Gene expression microarray analysis identified 8 genes correlated with tumor progression (LZTFL1, TAPT1, SEL1L, PAQR6, NME7, TMEM109, CCDC58, and ANKRD40) that were commonly influenced in HEC50B and SPAC-S by LAMC1 silencing.ConclusionLAMC1 overexpression is a potent biomarker for identifying endometrial cancer patients needing aggressive adjuvant therapy. We elucidated 8 candidate genes that may mediate progression of LAMC1 overexpressing cancer. Further investigation of the underlying mechanism should lead to the discovery of new therapeutic targets.

Embryonic response to high beta-hydroxybutyrate (BHB) levels in postpartum dairy cows

Cows at the beginning of lactation often do not meet their energy needs by feeding and therefore mobilize body fat, which produces ketone bodies, including β-hydroxybutyrate (BHB). They are nevertheless usually inseminated around 60 d postpartum, when they are still in this characteristic period of energy deficit. The aim of this study was to observe the effects of negative energy balance on embryo quality and to identify ways to improve the fertility of dairy cows. Holstein cows (n = 18) grouped as high or low BHB based on blood measurement at day 45 postpartum were estrus-synchronized and treated with follicle-stimulating hormone to obtain multiple follicle development, induced to ovulate and inseminated with sexed semen around day 60 postpartum. Of the 290 embryos collected over 16 mo, 159 were of quality I to IV. Based on microarray analysis of gene expression, exposure to an energy deficit metabolic environment (high BHB) during early development appeared to modify signaling by the mTOR and sirtuins pathways in the embryo, implying mitochondrial dysfunction and inhibition of transcription, leading to slower cell division, thus programming the embryo to be more energy efficient. Altered methylation markers suggested that such coping mechanisms might persist into adulthood.

MicroRNA-608 promotes apoptosis via BRD4 downregulation in pancreatic ductal adenocarcinoma.

The prognosis of pancreatic ductal adenocarcinoma (PDAC) remains poor even among patients with the same Tumor-Node-Metastasis stage. Thus, it is necessary to identify biomarkers that can accurately predict outcomes. There is accumulating evidence suggesting that microRNA (miR) expression influences overall survival (OS) time in patients with PDAC, via the regulation of tumor suppressor genes and oncogene expression. Specifically, miR-608 expression is hypothesized to regulate PDAC progression via the downregulation of bromodomain-containing protein 4 (BRD4) expression and the promotion of cell apoptosis. The present study aimed to investigate this theory. Thus, whole genome expression microarray analysis was performed on three patient samples with OS time >30 months, and compared with three samples with <12 months, in order to identify differentially expressed miRNAs (DEMs), via EdgeR analysis. A total of 591 DEMs were identified that exhibited a fold change >1, including 390 upregulated and 201 downregulated genes. Subsequently, 10 DEMs were identified using quantitative PCR in a different population of 68 tissues, collected from patients with PDAC. Notably, a high level of miRNA-608 expression was associated with longer OS times (P<0.05). Bioinformatics analysis was then performed to predict the molecular mechanism underlying the regulation of cell apoptosis by miRNA-608, and a dual-luciferase assay determined that overexpression of mimics in the Panc-1 and Bxpc-3 pancreatic cancer cell lines increased levels of apoptosis compared with the control. Additionally, high miRNA-608 expression decreased the protein level of BRD4. A luciferase reporting assay was used to elucidate whether miRNA-608 may directly inhibit the expression of BRD4 by binding to the 3′-untranslated region of its mRNA in the same cell lines. A subsequent rescue experiment indicated that the upregulation of BRD4 may reverse the apoptosis-promoting effect induced by miRNA-608. In summary, the present study revealed that miRNA-608 promotes apoptosis in PDAC via the negative regulation of BRD4. The results of the present study provide a theoretical basis that may improve the prediction of prognosis in patients with PDAC, and also indicate an opportunity to develop individualized treatment and investigate novel therapeutics that target these mechanisms.

Analysis of Long Non-Coding RNA Expression Profile and Functional Study of LOC389332 in Early Gastric Cancer.

BackgroundLong non-coding RNAs (LncRNAs) could potentially function as diagnostic markers for gastric carcinoma. Nevertheless, the expression profile and biological feature of LncRNAs in early gastric cancer (EGC) remains to be explored.Material/MethodsLncRNA expression microarray analysis was performed on 6 paired EGC tissues. One deregulated LncRNA, LOC389332, was validated using a quantitative reverse-transcription polymerase chain reaction (qRT-PCR) assay using independent tissue samples and cell lines. The Cell Counting Kit-8 (CCK-8) assay and wound healing assay were conducted to evaluate its influences on the proliferation and migration of gastric cancer cells. LncRNA expression microarray and gene ontology (GO) analysis were also performed on the LOC389332 knockdown cell line model to explore the molecular feature of LOC389332 in gastric carcinoma.ResultsThe LncRNA expression profiling showed that 72 LncRNAs were significantly differentially expressed in EGC tissues. The results in the validation phase revealed that LOC389332 was remarkably overexpressed in gastric carcinoma tissues, precancerous lesions, and gastric cancer cells. Functional study showed that knockdown of LOC389332 expression could inhibit cell proliferation and migration. LncRNA expression microarray on the LOC389332 knockdown cell line model revealed that 393 mRNAs were differentially expressed. The GO enrichment analysis indicated that the downregulated genes were mainly associated with cell membrane function, signal transmission process, and cell adhesion process.ConclusionsThe LncRNA expression profile between EGC and gastritis tissues was significantly different. LOC389332 was potential non-coding oncogenes in gastric cancer, and it may perform its function through altering cell membrane function, signal transmission, and cell adhesion.