Hyperplastic ovarian stromal cells express genes associated to tumor progression: a case study.

The current study presents the analysis of stromal cells obtained from an hyperplastic left-ovary of a Holstein cow. Cultured hyperplastic stromal cells displayed a fibroblast-like morphology and ceased proliferation after the 8th passage. The non-cancerous nature of stromal cells was confirmed by in vitro cell proliferation and migration assays. Negligible amounts of E2 were detected in the spent media of cultured stromal cells, which suggests that stromal cells were non-estradiol synthesizing cells. As revealed in immunofluorescence and gene expression analysis, the hyperplastic stromal cells explicitly expressed vimentin in their cytoskeleton. Upon hematoxylin staining, a highly dense population of stromal cells was observed in the stromal tissue of the hyperplastic ovary. To explore genome-wide alterations, mRNA microarray analysis was performed using Affymetrix Bovine Gene 1.0ST Arrays compared to normal ovarian derived stromal cells. The microarray identified 1396 differentially expressed genes, of which 733 were up- and 663 down-regulated in hyperplastic stromal cells. Importantly, asporin (ASPN) and vascular cell adhesion molecule 1 (VCAM1) were among the highly up-regulated genes. Higher expression of ASPN was also confirmed by immunohistochemistry and RT-qPCR analysis. Ingenuity pathway analysis (IPA) identified about 98 significantly enriched (-log (p value ≥ 1.3) canonical pathways, importantly of which the "Sirutin Signaling Pathway" and "Mitochondrial Dysfunction" were highly activated while "Oxidative phosphorylation" was inhibited. Additionally, higher proportion of hyperplastic stromal cells in the S-phase of cell cycle, could be attributed to higher expression levels of cell proliferation genes such as CCND2 and CDK6.

Early milk-feeding regimes in calves exert long-term effects on the development of ovarian granulosa cells.

BACKGROUND: Nutrition has not only an impact on the general wellbeing of an animal but can also affect reproductive processes. In cattle, feeding regimes can influence the age of puberty onset and alter gonadal development. We analyzed effects of different milk replacer (MR) feeding regimes during rearing on ovarian physiology with specific emphasis on the numbers as well as gene expression characteristics of granulosa cells (GCs) at the age of puberty onset. Two groups of calves received either 10% or 20% of bodyweight MR per day during their first 8 weeks. After weaning, both groups were fed the same mixed ration ad libitum until slaughter at 8 months. RESULTS: Animals of the 20% feeding group had a significantly higher body weight, but the proportion of animals having a corpus luteum at the time of slaughter was not different between groups, suggesting a similar onset of puberty. Calves of the 10% group showed a constant GC count regardless of the number of follicles (r = 0.23) whereas in the 20% group increasing numbers of GCs were detected with a higher follicle count (r = 0.71). As a first effort to find a possible molecular explanation for this unexpected limitation of GC numbers in the 10% group, we comparatively analyzed GC transcriptomes in both diet groups. The mRNA microarray analysis revealed a total of 557 differentially expressed genes comparing both groups (fold change > |1.5| and p < 0.05). OAS1X, MX2 and OAS1Z were among the top downregulated genes in the 20% vs. the 10% group, whereas top upregulated genes comprised BOLA and XCL1. All of these genes are known to be regulated by interferon. Subsequent signaling pathway analysis revealed the involvement of several immune response mechanisms in accordance with a number of interferons as upstream regulators. CONCLUSIONS: The results indicate that the plane of MR feeding in early life has an impact on the number and physiology of GCs later in life. This might influence the overall reproductive life initiated by the onset of puberty in cattle. In addition, the observed alterations in GCs of calves fed less MR might be a consequence of interferon regulated immunological pathways.

Transcriptome alterations in peripheral blood B cells of patients with multiple sclerosis receiving immune reconstitution therapy.

BACKGROUND: Multiple sclerosis (MS) is a chronic, inflammatory and neurodegenerative disease that leads to irreversible damage to the brain and spinal cord. The goal of so-called "immune reconstitution therapies" (IRTs) is to achieve long-term disease remission by eliminating a pathogenic immune repertoire through intense short-term immune cell depletion. B cells are major targets for effective immunotherapy in MS. OBJECTIVES: The aim of this study was to analyze the gene expression pattern of B cells before and during IRT (i.e., before B-cell depletion and after B-cell repopulation) to better understand the therapeutic effects and to identify biomarker candidates of the clinical response to therapy. METHODS: B cells were obtained from blood samples of patients with relapsing-remitting MS (n = 50), patients with primary progressive MS (n = 13) as well as healthy controls (n = 28). The patients with relapsing MS received either monthly infusions of natalizumab (n = 29) or a pulsed IRT with alemtuzumab (n = 15) or cladribine (n = 6). B-cell subpopulation frequencies were determined by flow cytometry, and transcriptome profiling was performed using Clariom D arrays. Differentially expressed genes (DEGs) between the patient groups and controls were examined with regard to their functions and interactions. We also tested for differences in gene expression between patients with and without relapse following alemtuzumab administration. RESULTS: Patients treated with alemtuzumab or cladribine showed on average a > 20% lower proportion of memory B cells as compared to before IRT. This was paralleled by profound transcriptome shifts, with > 6000 significant DEGs after adjustment for multiple comparisons. The top DEGs were found to regulate apoptosis, cell adhesion and RNA processing, and the most highly connected nodes in the network of encoded proteins were ESR2, PHB and RC3H1. Higher mRNA levels of BCL2, IL13RA1 and SLC38A11 were seen in patients with relapse despite IRT, though these differences did not pass the false discovery rate correction. CONCLUSIONS: We show that B cells circulating in the blood of patients with MS undergoing IRT present a distinct gene expression signature, and we delineated the associated biological processes and gene interactions. Moreover, we identified genes whose expression may be an indicator of relapse risk, but further studies are needed to verify their potential value as biomarkers.

Exploring Thiazolopyridine AV25R: Unraveling of Biological Activities, Selective Anti-Cancer Properties and In Silico Target and Binding Prediction in Hematological Neoplasms.

Thiazolopyridines are a highly relevant class of small molecules, which have previously shown a wide range of biological activities. Besides their anti-tubercular, anti-microbial and anti-viral activities, they also show anti-cancerogenic properties, and play a role as inhibitors of cancer-related proteins. Herein, the biological effects of the thiazolopyridine AV25R, a novel small molecule with unknown biological effects, were characterized. Screening of a set of lymphoma (SUP-T1, SU-DHL-4) and B- acute leukemia cell lines (RS4;11, SEM) revealed highly selective effects of AV25R. The selective anti-proliferative and metabolism-modulating effects were observed in vitro for the B-ALL cell line RS4;11. Further, we were able to detect severe morphological changes and the induction of apoptosis. Gene expression analysis identified a large number of differentially expressed genes after AV25R exposure and significant differentially regulated cancer-related signaling pathways, such as VEGFA-VEGFR2 signaling and the EGF/EGFR pathway. Structure-based pharmacophore screening approaches using in silico modeling identified potential biological AV25R targets. Our results indicate that AV25R binds with several proteins known to regulate cell proliferation and tumor progression, such as FECH, MAP11, EGFR, TGFBR1 and MDM2. The molecular docking analyses indicates that AV25R has a higher binding affinity compared to many of the experimentally validated small molecule inhibitors of these targets. Thus, here we present in vitro and in silico analyses which characterize, for the first time, the molecular acting mechanism of AV25R, including cellular and molecular biologic effects. Additionally, this predicted the target binding of the molecule, revealing a high affinity to cancer-related proteins and, thus, classified AVR25 for targeted intervention approaches.

Response of Osteoblasts on Amine-Based Nanocoatings Correlates with the Amino Group Density.

Increased life expectancy in industrialized countries is causing an increased incidence of osteoporosis and the need for bioactive bone implants. The integration of implants can be improved physically, but mainly by chemical modifications of the material surface. It was recognized that amino-group-containing coatings improved cell attachment and intracellular signaling. The aim of this study was to determine the role of the amino group density in this positive cell behavior by developing controlled amino-rich nanolayers. This work used covalent grafting of polymer-based nanocoatings with different amino group densities. Titanium coated with the positively-charged trimethoxysilylpropyl modified poly(ethyleneimine) (Ti-TMS-PEI), which mostly improved cell area after 30 min, possessed the highest amino group density with an N/C of 32%. Interestingly, changes in adhesion-related genes on Ti-TMS-PEI could be seen after 4 h. The mRNA microarray data showed a premature transition of the MG-63 cells into the beginning differentiation phase after 24 h indicating Ti-TMS-PEI as a supportive factor for osseointegration. This amino-rich nanolayer also induced higher bovine serum albumin protein adsorption and caused the cells to migrate slower on the surface after a more extended period of cell settlement as an indication of a better surface anchorage. In conclusion, the cell spreading on amine-based nanocoatings correlated well with the amino group density (N/C).

A genetic variant associated with multiple sclerosis inversely affects the expression of CD58 and microRNA-548ac from the same gene.

Genome-wide association studies have identified more than 200 genetic variants to be associated with an increased risk of developing multiple sclerosis (MS). Still, little is known about the causal molecular mechanisms that underlie the genetic contribution to disease susceptibility. In this study, we investigated the role of the single-nucleotide polymorphism (SNP) rs1414273, which is located within the microRNA-548ac stem-loop sequence in the first intron of the CD58 gene. We conducted an expression quantitative trait locus (eQTL) analysis based on public RNA-sequencing and microarray data of blood-derived cells of more than 1000 subjects. Additionally, CD58 transcripts and mature hsa-miR-548ac molecules were measured using real-time PCR in peripheral blood samples of 32 MS patients. Cell culture experiments were performed to evaluate the efficiency of Drosha-mediated stem-loop processing dependent on genotype and to determine the target genes of this underexplored microRNA. Across different global populations and data sets, carriers of the MS risk allele showed reduced CD58 mRNA levels but increased hsa-miR-548ac levels. We provide evidence that the SNP rs1414273 might alter Drosha cleavage activity, thereby provoking partial uncoupling of CD58 gene expression and microRNA-548ac production from the shared primary transcript in immune cells. Moreover, the microRNA was found to regulate genes, which participate in inflammatory processes and in controlling the balance of protein folding and degradation. We thus uncovered new regulatory implications of the MS-associated haplotype of the CD58 gene locus, and we remind that paradoxical findings can be encountered in the analysis of eQTLs upon data aggregation. Our study illustrates that a better understanding of RNA processing events might help to establish the functional nature of genetic variants, which predispose to inflammatory and neurological diseases.

Expression Profiling Suggests Loss of Surface Integrity and Failure of Regenerative Repair as Major Driving Forces for Chronic Obstructive Pulmonary Disease Progression.

Chronic obstructive pulmonary disease (COPD) poses a major risk for public health, yet remarkably little is known about its detailed pathophysiology. Definition of COPD as nonreversible pulmonary obstruction revealing more about spatial orientation than about mechanisms of pathology may be a major reason for this. We conducted a controlled observational study allowing for simultaneous assessment of clinical and biological development in COPD. Sixteen healthy control subjects and 104 subjects with chronic bronchitis, with or without pulmonary obstruction at baseline, were investigated. Using both the extent of and change in bronchial obstruction as main scoring criteria for the analysis of gene expression in lung tissue, we identified 410 genes significantly associated with progression of COPD. One hundred ten of these genes demonstrated a distinctive expression pattern, with their functional annotations indicating participation in the regulation of cellular coherence, membrane integrity, growth, and differentiation, as well as inflammation and fibroproliferative repair. The regulatory pattern indicates a sequentially unfolding pathology that centers on a two-step failure of surface integrity commencing with a loss of epithelial coherence as early as chronic bronchitis. Decline of regenerative repair starting in Global Initiative for Chronic Obstructive Lung Disease stage I then activates degradation of extracellular-matrix hyaluronan, causing structural failure of the bronchial wall that is only resolved by scar formation. Although they require independent confirmation, our findings provide the first tangible pathophysiological concept of COPD to be further explored.Clinical trial registered with www.clinicaltrials.gov (NCT00618137).

Genome wide effects of oleic acid on cultured bovine granulosa cells: evidence for the activation of pathways favoring folliculo-luteal transition.

BACKGROUND: Metabolic stress, as negative energy balance on one hand or obesity on the other hand can lead to increased levels of free fatty acids in the plasma and follicular fluid of animals and humans. In an earlier study, we showed that increased oleic acid (OA) concentrations affected the function of cultured bovine granulosa cells (GCs). Here, we focus on genome wide effects of increased OA concentrations. RESULTS: Our data showed that 413 genes were affected, of which 197 were down- and 216 up-regulated. Specifically, the expression of FSH-regulated functional key genes, CCND2, LHCGR, INHA and CYP19A1 and 17-ß-estradiol (E2) production were reduced by OA treatment, whereas the expression of the fatty acid transporter CD36 was increased and the morphology of the cells was changed due to lipid droplet accumulation. Bioinformatic analysis revealed that associated pathways of the putative upstream regulators "FSH" and "Cg (choriogonadotropin)" were inhibited and activated, respectively. Down-regulated genes are over-represented in GO terms "reproductive structure/system development", "ovulation cycle process", and "(positive) regulation of gonadotropin secretion", whereas up-regulated genes are involved in "circulatory system development", "vasculature development", "angiogenesis" or "extracellular matrix/structure organization". CONCLUSIONS: From these data we conclude that besides inhibiting GC functionality, increased OA levels seemingly promote angiogenesis and tissue remodelling, thus suggestively initiating a premature fulliculo-luteal transition. In vivo this may lead to impeded folliculogenesis and ovulation, and cause sub-fertility.

Proteostasis regulators modulate proteasomal activity and gene expression to attenuate multiple phenotypes in Fabry disease.

The lysosomal storage disorder Fabry disease is characterized by a deficiency of the lysosomal enzyme α-Galactosidase A. The observation that missense variants in the encoding GLA gene often lead to structural destabilization, endoplasmic reticulum retention and proteasomal degradation of the misfolded, but otherwise catalytically functional enzyme has resulted in the exploration of alternative therapeutic approaches. In this context, we have investigated proteostasis regulators (PRs) for their potential to increase cellular enzyme activity, and to reduce the disease-specific accumulation of the biomarker globotriaosylsphingosine in patient-derived cell culture. The PRs also acted synergistically with the clinically approved 1-deoxygalactonojirimycine, demonstrating the potential of combination treatment in a therapeutic application. Extensive characterization of the effective PRs revealed inhibition of the proteasome and elevation of GLA gene expression as paramount effects. Further analysis of transcriptional patterns of the PRs exposed a variety of genes involved in proteostasis as potential modulators. We propose that addressing proteostasis is an effective approach to discover new therapeutic targets for diseases involving folding and trafficking-deficient protein mutants.

The contact system proteases play disparate roles in streptococcal sepsis.

Sepsis causes an activation of the human contact system, an inflammatory response mechanism against foreign surfaces, proteins and pathogens. The serine proteases of the contact system, factor XII and plasma kallikrein, are decreased in plasma of septic patients, which was previously associated with an unfavorable outcome. However, the precise mechanisms and roles of contact system factors in bacterial sepsis are poorly understood. We, therefore, studied the physiological relevance of factor XII and plasma kallikrein in a mouse model of experimental sepsis. We show that decreased plasma kallikrein concentration in septic mice is a result of reduced mRNA expression plasma prekallikrein gene, indicating that plasma kallikrein belong to negative acute phase proteins. Investigations regarding the pathophysiological function of contact system proteases during sepsis revealed different roles for factor XII and plasma kallikrein. In vitro, factor XII decelerated bacteria induced fibrinolysis, whereas plasma kallikrein supported it. Remarkably, depletion of plasma kallikrein (but not factor XII) by treatment with antisense-oligonucleotides, dampens bacterial dissemination and growth in multiple organs in the mouse sepsis model. These findings identify plasma kallikrein as a novel host pathogenicity factor in Streptococcus pyogenes sepsis.

L-lactate induces specific genome wide alterations of gene expression in cultured bovine granulosa cells.

BACKGROUND: Previously, we could show that L-lactate affects cultured bovine granulosa cells (GC) in a specific manner driving the cells into an early pre-ovulatory phenotype. Here we studied genome wide effects in L-lactate-treated GC to further elucidate the underlying mechanisms that are responsible for the L-lactate induced transformation. Cultured estrogen producing GC treated either with L-lactate or vehicle control were subjected to mRNA microarray analysis. RESULTS: The analysis revealed 487 differentially expressed clusters, representing 461 annotated genes. Of these, 333 (= 318 genes) were identified as up- and 154 (= 143 genes) as down-regulated. As the top up-regulated genes we detected TXNIP, H19 and AHSG as well as our previously established marker transcripts RGS2 and PTX3. The top down-regulated genes included VNN1, SLC27A2 and GFRA1, but also MYC and the GC marker transcript CYP19A1. Pathway analysis with differentially expressed genes indicated "cAMP-mediated signaling" and "Axon guidance signaling" among the most affected pathways. Furthermore, estradiol, progesterone and Vegf were identified as potential upstream regulators. An effector network analysis by IPA provided first hints that processes of "angiogenesis" and "vascularization", but also "cell movement" appeared to be activated, whereas "organismal death" was predicted to be inhibited. CONCLUSIONS: Our data clearly show that L-lactate alters gene expression in cultured bovine GC in a broad, but obviously specific manner. Pathway analysis revealed that the mode of L-lactate action in GC initiates angiogenic processes, but also migratory events like cell movement and axonal guidance signaling, thus supporting the transformation of GC into an early luteal phenotype.

Testicular transcriptional signatures associated with high fertility.

Factors of high fertility are poorly described. The majority of transgenic or knockout models with a reproductive phenotype are subfertile or infertile phenotypes. Few genotypes have been linked to improved reproductive performance (0.2%) or increased litter size (1%). In this study, we used a unique mouse model, fertility line FL1, selected for 'high fertility' for more than 170 generations. This strain has almost doubled the number of littermates as well as their total birth weight accompanied by an elevated ovulation rate and increased numbers of corpora lutea compared to a randomly mated and unselected control line (Ctrl). Here, we investigate whether the gonadal tissue of FL1 males are affected by 'co-evolution' after more than 40 years of female-focused selection. Using microarrays, we analysed the testicular transcriptome of the FL1 and Ctrl mice. These data were also compared with previously published female gonadal transcriptional alterations. We detected alterations in testicular gene expression, which are partly associated with female reproductive performance. Thus, female-focused selection for litter size has not only affected the female side, but also has been manifested in transcriptional alterations on the male gonadal organ. This suggests consequences for the entire mouse lines in the long run and emphasizes the perspective of inevitably considering both genders about mechanisms of high fertility.

Copy Number Alterations with Prognostic Potential in Clear Cell Renal Cell Carcinoma.

OBJECTIVES: To detect chromosomal aberrations in a genome-wide manner with potential value for prognosis in groups of patients with different histopathological grading in clear cell renal carcinoma (ccRCC). MATERIAL AND METHODS: We performed a copy number alteration analysis using the Affymetrix platform and SNP 6.0 mapping arrays with samples from 48 ccRCC-patients. The data analysis was done using 3 different Software Platforms: Affymetrix Genotyping Console (version 4.1.3.840) and 2 open-source packages for validation: PennCNV and PICNIC. RESULTS: Consistent changes were found to divide the tumors into 4 groups: first group showed typical losses on 3p, second group losses on 3p plus gains on 5q, third group gains on chromosome 7 plus losses on chromosome 8; fourth group did not show any major changes. We selected the affected genes with the highest consistency and identified 13 different genes mapping in the SNP 6.0 results and Kyoto Encyclopedia of Genes and Genomes. Remarkable for further consideration were the phosphatidylinositol 3-kinase pathway, BRAF, MET, EGLN1; growth factors, for example, HGF, PGF and TGFB2. CONCLUSION: A multimodal approach with a well-defined workflow for detecting genomic aberrations by using array technologies and comparing the findings with different comprehensive databases may provide insights into functional tumor processes and help to identify potential new targets for more individualized future treatment.

Role of Bone Morphogenetic Protein 7 (BMP7) in the Modulation of Corneal Stromal and Epithelial Cell Functions.

In the cornea, healing of the wounded avascular surface is an intricate process comprising the involvement of epithelial, stromal and neuronal cell interactions. These interactions result to the release of various growth factors that play prominent roles during corneal wound healing response. Bone morphogenetic proteins (BMPs) are unique multi-functional potent growth factors of the transforming growth factor-beta (TGF-β) superfamily. Treatment of corneal epithelial cells with substance P and nerve growth factor resulted to an increase in the expression of BMP7 mRNA. Since BMP7 is known to modulate the process of corneal wound healing, in this present study, we investigated the influence of exogenous rhBMP7 on human corneal epithelial cell and stromal cell (SFs) function. To obtain a high-fidelity expression profiling of activated biomarkers and pathways, transcriptome-wide gene-level expression profiling of epithelial cells in the presence of BMP7 was performed. Gene ontology analysis shows BMP7 stimulation activated TGF-β signaling and cell cycle pathways, whereas biological processes related to cell cycle, microtubule and intermediate filament cytoskeleton organization were significantly impacted in corneal epithelial cells. Scratch wound healing assay showed increased motility and migration of BMP7 treated epithelial cells. BMP7 stimulation studies show activation of MAPK cascade proteins in epithelial cells and SFs. Similarly, a difference in the expression of claudin, Zink finger E-box-binding homeobox 1 was observed along with phosphorylation levels of cofilin in epithelial cells. Stimulation of SFs with BMP7 activated them with increased expression of α-smooth muscle actin. In addition, an elevated phosphorylation of epidermal growth factor receptor following BMP7 stimulation was also observed both in corneal epithelial cells and SFs. Based on our transcriptome analysis data on epithelial cells and the results obtained in SFs, we conclude that BMP7 contributes to epithelial-to-mesenchymal transition-like responses and plays a role equivalent to TGF-β in the course of corneal wound healing.

Selection for female traits of high fertility affects male reproductive performance and alters the testicular transcriptional profile.

BACKGROUND: Many genes important for reproductive performance are shared by both sexes. However, fecundity indices are primarily based on female parameters such as litter size. We examined a fertility mouse line (FL2), which has a considerably increased number of offspring and a total litter weight of 180% compared to a randomly bred control line (Ctrl) after more than 170 generations of breeding. In the present study, we investigated whether there might be a parallel evolution in males after more than 40 years of breeding in this outbred mouse model. RESULTS: Males of the fertility mouse line FL2 showed reduced sperm motility performance in a 5 h thermal stress experiment and reduced birth rate in the outbred mouse line. Transcriptional analysis of the FL2 testis showed the differential expression of genes associated with steroid metabolic processes (Cyp1b1, Cyp19a1, Hsd3b6, and Cyp21a1) and female fecundity (Gdf9), accompanied by 150% elevated serum progesterone levels in the FL2 males. Cluster analysis revealed the downregulation of genes of the kallikrein-related peptidases (KLK) cluster located on chromosome 7 in addition to alterations in gene expression with serine peptidase activity, e.g., angiotensinogen (Agt), of the renin-angiotensin system essential for ovulation. Although a majority of functional annotations map to female reproduction and ovulation, these genes are differentially expressed in FL2 testis. CONCLUSIONS: These data indicate that selection for primary female traits of increased litter size not only affects sperm characteristics but also manifests as transcriptional alterations of the male side likely with direct long-term consequences for the reproductive performance of the mouse line.

Induction of altered gene expression profiles in cultured bovine granulosa cells at high cell density.

BACKGROUND: In previous studies it has been shown that bovine granulosa cells (GC) cultured at a high plating density dramatically change their physiological and molecular characteristics, thus resembling an early stage of luteinization. During the present study, these specific effects on the GC transcriptome were comprehensively analysed to clarify the underlying mechanisms. METHODS: GC were cultured in serum free medium with FSH and IGF-1 stimulation at different initial plating density. The estradiol and progesterone production was determined by radioimmunoassays and the gene expression profiles were analysed by mRNA microarray analysis after 9 days. The data were statistically analysed and the abundance of selected, differentially expressed transcripts was re-evaluated by qPCR. Bioinformatic pathway analysis of density affected transcripts was done using Ingenuity Pathway Analysis. RESULTS: The data showed that at high plating density the expression of 1510 annotated genes, represented by 1575 transcript clusters, showed highly altered expression levels. Nearly two-thirds were up- and one third down-regulated. Within the top up-regulated genes VNN2, RGS2 and PTX3 could be identified, as well as HBA or LOXL2. Down-regulated genes included important key genes of folliculogenesis like CYP19A1 and FSHR. Ingenuity pathway analysis identified "AMPK signaling" as well as "cAMP-mediated signaling" as major pathways affected by the alteration of the expression profile. Main putative upstream regulators were TGFB1 and VEGF, thus indicating a connection with cell differentiation and angiogenesis. A detailed cluster analysis revealed one single cluster that was highly associated with the upstream regulator beta-estradiol. Within this cluster key genes of steroid biosynthesis were not included, but instead, other genes importantly involved in follicular development, like OXT and VEGFA as well as the three most down-regulated genes TXNIP, PAG11 and ARRDC4 were identified. CONCLUSIONS: From these data we hypothesize that high density conditions induce a stage of differentiation in cultured GC that is similar to early post-LH conditions in vivo. Furthermore we hypothesize that specific cell-cell-interactions led to this differentiation including transformations necessary to promote angiogenesis.

Global Transcriptional Response of Human Liver Cells to Ethanol Stress of Different Strength Reveals Hormetic Behavior.

BACKGROUND: The liver is the major site for alcohol metabolism in the body and therefore the primary target organ for ethanol (EtOH)-induced toxicity. In this study, we investigated the in vitro response of human liver cells to different EtOH concentrations in a perfused bioartificial liver device that mimics the complex architecture of the natural organ. METHODS: Primary human liver cells were cultured in the bioartificial liver device and treated for 24 hours with medium containing 150 mM (low), 300 mM (medium), or 600 mM (high) EtOH, while a control culture was kept untreated. Gene expression patterns for each EtOH concentration were monitored using Affymetrix Human Gene 1.0 ST Gene chips. Scaled expression profiles of differentially expressed genes (DEGs) were clustered using Fuzzy c-means algorithm. In addition, functional classification methods, KEGG pathway mapping and also a machine learning approach (Random Forest) were utilized. RESULTS: A number of 966 (150 mM EtOH), 1,334 (300 mM EtOH), or 4,132 (600 mM EtOH) genes were found to be differentially expressed. Dose-response relationships of the identified clusters of co-expressed genes showed a monotonic, threshold, or nonmonotonic (hormetic) behavior. Functional classification of DEGs revealed that low or medium EtOH concentrations operate adaptation processes, while alterations observed for the high EtOH concentration reflect the response to cellular damage. The genes displaying a hormetic response were functionally characterized by overrepresented "cellular ketone metabolism" and "carboxylic acid metabolism." Altered expression of the genes BAHD1 and H3F3B was identified as sufficient to classify the samples according to the applied EtOH doses. CONCLUSIONS: Different pathways of metabolic and epigenetic regulation are affected by EtOH exposition and partly undergo hormetic regulation in the bioartificial liver device. Gene expression changes observed at high EtOH concentrations reflect in some aspects the situation of alcoholic hepatitis in humans.

ADAM23 promotes neuronal differentiation of human neural progenitor cells.

BACKGROUND: ADAM23 is widely expressed in the embryonic central nervous system and plays an important role in tissue formation. RESULTS: In this study, we showed that ADAM23 contributes to cell survival and is involved in neuronal differentiation during the differentiation of human neural progenitor cells (hNPCs). Upregulation of ADAM23 in hNPCs was found to increase the number of neurons and the length of neurite, while its downregulation decreases them and triggers cell apoptosis. RNA microarray analysis revealed mechanistic insights into genes and pathways that may become involved in multiple cellular processes upon up- or downregulation of ADAM23. CONCLUSIONS: Our results suggest that ADAM23 regulates neuronal differentiation by triggering specific signaling pathways during hNPC differentiation.

SLCO1B1*5 polymorphism (rs4149056) is associated with chemotherapy-induced amenorrhea in premenopausal women with breast cancer: a prospective cohort study.

BACKGROUND: Because inheritance is recognized as playing a role in age at menarche and natural menopause, the development of chemotherapy-induced amenorrhea (CIA) might depend on inherited genetic factors; however, studies that explore such a correlation are few and have received scant attention. Given the importance of this topic we conducted a comprehensive genotype study in young women (≤45 years) with early-stage breast cancer. METHODS: Our approach tested the effect of variant polymorphisms in drug metabolism enzymes (DMEs) using a predesigned pharmacogenomics panel (TaqMan® OpenArray®, Life Technologies GmbH, Darmstadt, Germany) in premenopausal women (n = 50). Patients received contemporary chemotherapy; in all cases a cyclophosphamide-based regimen with a dose of at least 500 mg/m(2) for six cycles. CIA was considered to be present in women with no resumption of menstrual bleeding within 12 months after completion of chemotherapy or goserelin. RESULTS: Twenty-six patients (52 %) showed CIA during follow-up whereas 24 women (48 %) remained premenopausal. Of all the DMEs studied, only the SLCO1B1*5 (rs4149056) genotype was associated with the development of CIA (P = 0.017). Of the 26 patients who were homozygous for the T/T allele SLCO1B1*5, 18 (69.2 %) developed CIA compared with 8 (30.8 %) of the 22 patients who were heterozygous (C/T allele). The association of heterozygous SLCO1B1*5 allele (OR 0.038; 95%CI: 0.05-0.92) with a lower risk of developing CIA remained significant in a binary logistic regression analysis that include age, SLCO1B1*5 allele variants, and goserelin therapy. CONCLUSIONS: Patient age and SLCO1B1*5 allele variants predict the likelihood of young women with breast cancer developing CIA.