scholarly journals Distinct Gene Expression Profiling after Infection of Immature Human Monocyte-Derived Dendritic Cells by the Attenuated Poxvirus Vectors MVA and NYVAC

2007 ◽  
Vol 81 (16) ◽  
pp. 8707-8721 ◽  
Author(s):  
Susana Guerra ◽  
José Luis Nájera ◽  
José Manuel González ◽  
Luis A. López-Fernández ◽  
Nuria Climent ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dendritic Cells ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Human Monocyte ◽  
Type I ◽  
Mrna Levels ◽  
Antigen Uptake

ABSTRACT Although recombinants based on the attenuated poxvirus vectors MVA and NYVAC are currently in clinical trials, the nature of the genes triggered by these vectors in antigen-presenting cells is poorly characterized. Using microarray technology and various analysis conditions, we compared specific changes in gene expression profiling following MVA and NYVAC infection of immature human monocyte-derived dendritic cells (MDDC). Microarray analysis was performed at 6 h postinfection, since these viruses induced extensive cytopathic effects, rRNA breakdown, and apoptosis at late times postinfection. MVA- and NYVAC-infected MDDC shared upregulation of 195 genes compared to uninfected cells: MVA specifically upregulated 359 genes, and NYVAC upregulated 165 genes. Microarray comparison of NYVAC and MVA infection revealed 544 genes with distinct expression patterns after poxvirus infection and 283 genes specifically upregulated after MVA infection. Both vectors upregulated genes for cytokines, cytokine receptors, chemokines, chemokine receptors, and molecules involved in antigen uptake and processing, including major histocompatibility complex genes. mRNA levels for interleukin 12β (IL-12β), beta interferon, and tumor necrosis factor alpha were higher after MVA infection than after NYVAC infection. The expression profiles of transcription factors such as NF-κB/Rel and STAT were regulated similarly by both viruses; in contrast, OASL, MDA5, and IRIG-I expression increased only during MVA infection. Type I interferon, IL-6, and Toll-like receptor pathways were specifically induced after MVA infection. Following MVA or NYVAC infection in MDDC, we found similarities as well as differences between these virus strains in the expression of cellular genes with immunological function, which should have an impact when these vectors are used as recombinant vaccines.

scholarly journals Three Years Follow-Up Study Showed Prenatal Methamphetamine Exposure May Cause Chronic Abnormalities In Transcriptomic Pattern

2021 ◽  
Author(s):  
Arvin Haghighatfard ◽  
Soha Seifollahi ◽  
Pegah Rajabi ◽  
Niloofar Rahmani ◽  
Rojin Ghannadzadeh
Keyword(s):  
Gene Expression ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Rna Extraction ◽  
Childbearing Age ◽  
Methamphetamine Use ◽  
Methamphetamine Use Disorder

Abstract Background: The high rate of methamphetamine use disorder among young adults and women of childbearing age makes it imperative to clarify the long-term effects of Methamphetamine exposure on the offspring. Behavioral and cognitive problems had been reported in children with parental Methamphetamine exposure (PME). The present study aimed to assess the acute and chronic effects of PME in molecular regulations and gene expression profiles of children during their first years of life.Methods: All subjects were recruited before birth, and sampling was conducted from the first ten days of birth, twelve months, twenty months, and thirty-six months of age. Finally, 2658 children with PME and 3573 normal children had been finished the follow-up. RNA extraction was operated from blood samples and gene expression profiling was conducted by using the Affymetrix GeneChip Human Genome U133 plus 2.0 Array Platform. Gene expression data were confirmed by Real-time PCR. Results: Gene expression profiling during thirty-six months showed several constant mRNA level alterations in children with PME compared with normal. These genes are involved in several gene ontologies and pathways involved with the immune system, neuronal functions, and bioenergetic metabolism. It seems that Methamphetamine use disorder before and during the pregnancy period may affect the expression profile of children, and these changes could remain years after birth. Affected genes have some similarities with the gene expression patterns of addiction, psychiatric disorders, neurodevelopmental disabilities, and immune deficiencies. Conclusion: Findings may shed light on the molecular effects of prenatal methamphetamine exposure and may lead to new psychological and somatic caring protocols for these children based on their potential abnormalities.

scholarly journals Prenatal Methamphetamine Exposure could Cause Chronic Abnormalities in Transcriptomic Pattern; an Extended Follow-up Cohort Study

2021 ◽  
Author(s):  
Arvin Haghighatfard ◽  
Soha Seifollahi ◽  
Pegah Rajabi ◽  
Niloofar Rahmani ◽  
Rojin Ghannad zadeh
Keyword(s):  
Gene Expression ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Rna Extraction ◽  
High Rate ◽  
Childbearing Age ◽  
Methamphetamine Abuse

Abstract BackgroundThe high rate of methamphetamine abuse among young adults and women of childbearing age makes it imperative to clarify the long-term effects of Methamphetamine exposure on the offspring. Behavioral and cognitive problems had reported in children with parental Methamphetamine exposure (PME). The present study aimed to assess the acute and chronic effects of PME in molecular regulations and gene expression profiles of children during their first years of life.ResultsAll subjects were recruited before birth, and sampling was conducted from the first ten days of birth, twelve months, twenty months and thirty-six months of age. Finally, 2658 children with PME and 3573 normal children had been finished the follow-up. RNA extraction was operated from blood samples and gene expression profiling was conducted by using the Affymetrix GeneChip Human Genome U133 plus 2.0 Array Platform. Gene expression data were confirmed by Real-time PCR. Gene expression profiling during thirty-six months showed several constant mRNA level alterations in children with PME compared with normal. These genes are involved in several gene ontology and pathways involved with the immune system, neuronal functions and bioenergetic metabolism. It seems that Methamphetamine abuse before and during the pregnancy period may affect the expression profile of children, and these changes could be remain years after birth. Affected genes have some similarities to the gene expression patterns of addiction, psychiatric disorders, neurodevelopmental disabilities and immune deficiencies. ConclusionFindings may shed light on the molecular effects of prenatal methamphetamine exposure and may lead to new psychological and somatic caring protocols for these children based on their potential abnormalities.

All Clinically Relevant Leukemia Subtypes Can Be Diagnosed and Classified Based Solely on Gene Expression Profiling with an Accuracy of 95.1%: A Study on 1337 Adult Patients.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 143-143
Author(s):  
Torsten Haferlach ◽  
Alexander Kohlmann ◽  
Susanne Schnittger ◽  
Martin Dugas ◽  
Sylvia Merk ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Profiling ◽  
Sensitivity And Specificity ◽  
Diagnostic Tool ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Second Step ◽  
Support Vector ◽  
Therapeutic Decisions

Abstract So far, comprehensive diagnosis of leukemia requires a combination of cytomorphology, immunophenotyping, and genetic methods. We aimed at developing a new diagnostic tool based solely on gene expression profiling to accurately predict all clinically relevant subtypes of leukemia in adults and to distinguish these from normal bone marrow. Therefore, we analyzed samples from 1337 untreated patients at diagnosis and healthy donors using oligonucleotide microarrays. The first series of 937 cases was hybridized to HG-U133A+B microarrays (Affymetrix). The following 13 subgroups were included: 620 AML (42 t(15;17); 38 t(8;21); 49 inv(16); 47 t(11q23); 75 complex aberrant karyotype; 193 normal karyotype; 176 other cytogenetic abn.); 152 ALL (26 Pro-B-ALL/t(11q23); 12 ALL-t(8;14); 32 T-ALL; 82 c-ALL/Pre-B-ALL); 75 CML, 45 CLL, and 45 bone marrows from healthy volunteers or non-leukemia pts. (nBM). For each disease entity the top 100 differentially expressed genes were calculated in a one-versus-all (OVA) approach. Class prediction was performed using support vector machines (SVM). Prediction accuracy was estimated by 10-fold cross validation (CV) and assessed for robustness in a resampling approach. 891 of the 937 samples (95.1%) were correctly classified (10-fold CV). A resampling approach with 2/3 training and 1/3 test cohort (100 runs of SVM) confirmed this high accuracy (median, 93.8%). In particular, a median of 100% sensitivity and specificity was achieved for AML with t(15;17), t(8;21), and inv(16), as well as for Pro-B-ALL/t(11q23), and CLL. The median specificity was at least 99.7% in all subgroups except for AML normal/other (median specificity, 93.7%). In a second step T-ALL cases were separated into cortical and immature ones (accuracy, 84.4%) and c-ALL/Pre-B-ALL into cases with and without t(9;22) (accuracy, 82.9%). The second prospective series comprized 400 unselected cases which were hybridized to the new generation HG-U133 Plus 2.0 microarrays (Affymetrix). To validate the diagnostic accuracy of our approach these cases were processed blinded in parallel to routine diagnostic work-up and classified based on the gene expression signatures discovered in the first series described above. Applying a first classification step as described above the 13 different diagnoses were predicted with an accuracy of 94.5%. Failures were mostly due to misclassification into biologically related subgroups, e.g. AML with del(5q) aberrations classified as AML with complex aberrant karyotype. In the second step (separation of the two T-ALL subtypes, and c-ALL/Pre-B-ALL with or without t(9;22)) accuracies of 100% and 70.6% respectively were achieved. In conclusion, we were able to identify within a routine diagnostic workflow distinct expression profiles for all clinically and prognostically relevant adult leukemia subtypes and their discrimination from nBM based only on gene expression data. Accuracy, sensitivity, and specificity were higher than achieved with each of the gold standard techniques alone used today. Thus, gene expression patterns analyzed by microarrays qualify as a diagnostic tool in a routine setting for leukemia diagnosis and classification and may guide relevant therapeutic decisions.

Automated Sorting of Monocytes from Whole Blood for Reproducible Gene Expression Profiling.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3840-3840
Author(s):  
Carsten Poggel ◽  
Timo Adams ◽  
Sabine Martin ◽  
Carola Pickel ◽  
Nicole Prahl ◽  
...  
Keyword(s):  
Gene Expression ◽  
Blood Cell ◽  
Gene Expression Profiling ◽  
Whole Blood ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Gene Expression Profiles ◽  
Cell Types ◽  
Specific Gene

Abstract Microarray-based gene expression profiling has been used to develop clinically relevant molecular classifiers for many different diseases. Furthermore, it has been shown for various chronic diseases that specific gene expression patterns are reflected at the level of blood cells. However, blood is a complex tissue comprising numerous cell types. Therefore, the contribution of rare cell types to a whole blood expression profile might not be detected and a substantial proportion of what is usually reported as “up-regulation” or “down-regulation” might actually be the result of a shift in cell populations and not of a true regulatory process. In order to circumvent these problems, several techniques have been established to analyze purified subpopulations rather than whole blood samples. Previously, it has been shown, for example, that reproducible gene expression profiles can be generated by positive selection of blood cell subsets from PBMCs1. As the preparation of PBMCs by, for example, Ficoll is time-consuming, inconvenient, and not amenable to automation, we have set up a combined direct whole blood cell separation and gene expression profiling protocol. By using Whole Blood CD14 MicroBeads in combination with the autoMACS Pro™ Separator, the separation protocol generally allowed enrichment of monocytes from whole blood within 30 min with purities higher than 90%. In combination with the depletion of neutrophils, the major source of contaminating RNA, purities increased to over 95% for all tested blood donors. Monocytes included the CD14bright/CD16− as well as the CD14dim/CD16+ populations. To assess the reproducibility of gene expression profiles and the influence of several experimental parameters, monocytes were sorted from 5 ml whole blood. RNA was extracted and hybridized to microarrays and the Pearson correlation coefficients of pairwise comparisons were calculated. Technical repeats of monocyte analysis from blood donated at different days showed a higher correlation coefficient than whole blood RNA. Blood storage at room temperature resulted in a strong deregulation of many genes, whereas blood stored at 4°C showed minimal changes, which is in agreement with previous studies. Skipping the centrifugation step, which is used to remove unbound MicroBeads did not alter the gene expression profiles. Incubation of sorted cells in PrepProtect™ Stabilization Buffer showed no alteration of gene expression thus enabling the shipping of cells without liquid nitrogen. Monocytes play a crucial role in diseases like atherosclerosis. Our rapid and simple protocol for combined direct cell sorting from whole blood and gene expression profiling of monocytes might help to ease the discovery of new biomarkers and to screen and monitor patients. 1 Lyons et al., BMC Genomics (2007), 8:64.

scholarly journals Sequential gene expression profiling in the mouse spleen during 14 d feeding withLactobacillus brevisKB290

2014 ◽  
Vol 111 (11) ◽  
pp. 1957-1966 ◽  
Author(s):  
Yuichiro Fukui ◽  
Erika Sasaki ◽  
Nobuo Fuke ◽  
Yuji Nakai ◽  
Tomoko Ishijima ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cytotoxic Activity ◽  
Gene Expression Profiling ◽  
Dna Microarray ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Killer Cell ◽  
Mrna Levels ◽  
Gene Set ◽  
Go Terms

Some lactic acid bacteria play an important role in the immune system with potential benefits to the host. However, detailed mechanisms of immune modulation exerted by probiotics remain to be clarified. Since immune response changes in a time-related manner in some cases, we monitored changes in mRNA levels in the spleen of mice during 14 d feeding withLactobacillus brevisKB290 (KB290). Female BALB/c mice, aged 9 weeks, commenced a diet containing KB290 (3 × 109colony-forming units/g) or starch for a period of 1, 4, 7 or 14 d. Cytotoxic activity of the resulting splenocytes against YAC-1 cells was measured using flow cytometry. The activity was found to be significantly higher in the treated group on days 1 and 7. The highest activity appeared on day 4, but was not statistically significantly different. Gene expression profiles were analysed using DNA microarray. Gene Ontology (GO) terms related to the immune process were significantly enriched in the up-regulated gene set on days 1, 4 and 7, and GO terms related to the cellular process were enriched in the down-regulated gene set on days 4 and 7. Although the up-regulated genes involved in antigen processing and presentation for stimulation of CD8+cytotoxic T cells were not observed on day 14, some genes involved in T-cell and natural killer cell activation remained up-regulated until day 14. For the majority of the genes tested, RT-PCR analysis was used to verify the results obtained from the DNA microarray analysis. The sequential gene expression profiling reflected changes in cytotoxic activity during KB290 feeding.

Gene expression analysis of the pro-oestrous-stage rat uterus reveals neuroligin 2 as a novel steroid-regulated gene

2004 ◽  
Vol 16 (8) ◽  
pp. 763 ◽  
Author(s):  
Han-Seung Kang ◽  
Chae-Kwan Lee ◽  
Ju-Ran Kim ◽  
Seong-Jin Yu ◽  
Sung-Goo Kang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Oestrous Cycle ◽  
Mrna Levels ◽  
Rat Uterus ◽  
Ovx Rats

In the present study, differential gene expression in the uteri of ovariectomised (OVX) and pro-oestrous rats (OVX v. pro-oestrus pair) was investigated using cDNA expression array analysis. Differential uterine gene expression in OVX rats and progesterone (P4)-injected OVX rats (OVX v. OVX + P4 pair) was also examined. The uterine gene expression profiles of these two sets of animals were also compared for the effects of P4 treatment. RNA samples were extracted from uterine tissues and reverse transcribed in the presence of [α32P]-dATP. Membrane sets of rat arrays were hybridised with cDNA probe sets. Northern blot analysis was used to validate the relative gene expression patterns obtained from the cDNA array. Of the 1176 cDNAs examined, 23 genes showed significant (>two-fold) changes in expression in the OVX v. pro-oestrus pair. Twenty of these genes were upregulated during pro-oestrus compared with their expression in the OVX rat uterus. In the OVX v. OVX + P4 pair, 22 genes showed significant (>two-fold) changes in gene expression. Twenty of these genes were upregulated in the OVX + P4 animals. The genes for nuclear factor I–XI, afadin, neuroligin 2, semaphorin Z, calpain 4, cyclase-associated protein homologue, thymosin β-4X and p8 were significantly upregulated in the uteri of the pro-oestrus and OVX + P4 rats of both experimental pairs compared with the OVX rat uteri. These genes appear to be under the control of P4. One of the most interesting findings of the present study is the unexpected and marked expression of the neuroligin 2 gene in the rat uterus. This gene is expressed at high levels in the central nervous system and acts as a nerve cell adhesion factor. According to Northern blot analysis, neuroligin 2 gene expression was higher during the pro-oestrus and metoestrus stages than during the oestrus and dioestrus stages of the oestrous cycle. In addition, neuroligin 2 mRNA levels were increased by both 17β-oestradiol (E2) and P4, although P4 administration upregulated gene expression to a greater extent than injection of E2. These results indicate that neuroligin 2 gene expression in the rat uterus is under the control of both E2 and P4, which are secreted periodically during the oestrous cycle.

scholarly journals 256CONSTRUCTION OF A BOVINE CDNA ARRAY TO MONITOR GENE EXPRESSION PROFILES IN BOVINE PREIMPLANTATION EMBRYOS

2004 ◽  
Vol 16 (2) ◽  
pp. 248
Author(s):  
C. Wrenzycki ◽  
T. Brambrink ◽  
D. Herrmann ◽  
J.W. Carnwath ◽  
H. Niemann
Keyword(s):  
Gene Expression ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Cdna Array ◽  
Preimplantation Embryos ◽  
Average Insert Size ◽  
Rt Pcr ◽  
Public Data

Array technology is a widely used tool for gene expression profiling, providing the possibility to monitor expression levels of an unlimited number of genes in various biological systems including preimplantation embryos. The objective of the present study was to develop and validate a bovine cDNA array and to compare expression profiles of embryos derived from different origins. A bovine blastocyst cDNA library was generated. Poly(A+)RNA was extracted from in vitro-produced embryos using a Dynabead mRNA purification kit. First-strand synthesis was performed with SacIT21 primer followed by randomly primed second-strand synthesis with a DOP primer mix (Roche) and a global PCR with 35 cycles using SacIT21 and DOP primers. Complementary DNA fragments from 300 to 1500bp were extracted from the gel and normalized via reassoziation and hydroxyapatite chromatography. Resulting cDNAs were digested with SacI and XhoI, ligated into a pBKs vector, and transfected into competent bacteria (Stratagene). After blue/white colony selection, plasmids were extracted and the inserts were subjected to PCR using vector specific primers. Average insert size was determined by size idenfication on agarose gels stained with ethidium bromide. After purification via precipitation and denaturation, 192 cDNA probes were double-spotted onto a nylon membrane and bound to the membrane by UV cross linking. Amplified RNA (aRNA) probes from pools of three or single blastocysts were generated as described recently (Brambrink et al., 2002 BioTechniques, 33, 3–9) and hybridized to the membranes. Expression profiles of in vitro-produced blastocysts cultured in either SOF plus BSA or TCM plus serum were compared with those of diploid parthenogenetic ones generated by chemical activation. Thirty-three probes have been sequenced and, after comparison with public data bases, 26 were identified as cDNAs or genes. Twelve out of 192 (6%) seem to be differentially expressed within the three groups;; 7/12 (58%) were down-regulated, 3/12 (25%) were up-regulated in SOF-derived embryos, and 2/12 (20%) were up-regulated in parthenogenetic blastocysts compared to their in vitro-generated counterparts. Three of these genes involved in calcium signaling (calmodulin, calreticulin) and regulation of actin cytoskeleton (destrin) were validated by semi-quantitative RT-PCR (Wrenzycki et al., 2001 Biol. Reprod. 65, 309–317) employing poly(A+) RNA from a single blastocyst as starting material. No differences were detected in the relative abundance of the analysed gene transcripts within the different groups. These findings were confirmed employing the aRNA used for hybridization in RT-PCR and showed a good representativity of the selected transcripts. Results indicate that it is possible to construct a homologous cDNA array which could be used for gene expression profiling of bovine preimplantation embryos. Supported by the Deutsche Forschungsgemeinschaft (DFG Ni 256/18-1).

scholarly journals Effectiveness of Gene Expression Profiling for Response Prediction of Rectal Adenocarcinomas to Preoperative Chemoradiotherapy

2005 ◽  
Vol 23 (9) ◽  
pp. 1826-1838 ◽  
Author(s):  
B. Michael Ghadimi ◽  
Marian Grade ◽  
Michael J. Difilippantonio ◽  
Sudhir Varma ◽  
Richard Simon ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Profiling ◽  
Expression Profiling ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Response Prediction ◽  
Preoperative Chemoradiotherapy ◽  
Response To Therapy ◽  
Phase Iii ◽  
Correct Prediction

Purpose There is a wide spectrum of tumor responsiveness of rectal adenocarcinomas to preoperative chemoradiotherapy ranging from complete response to complete resistance. This study aimed to investigate whether parallel gene expression profiling of the primary tumor can contribute to stratification of patients into groups of responders or nonresponders. Patients and Methods Pretherapeutic biopsies from 30 locally advanced rectal carcinomas were analyzed for gene expression signatures using microarrays. All patients were participants of a phase III clinical trial (CAO/ARO/AIO-94, German Rectal Cancer Trial) and were randomized to receive a preoperative combined-modality therapy including fluorouracil and radiation. Class comparison was used to identify a set of genes that were differentially expressed between responders and nonresponders as measured by T level downsizing and histopathologic tumor regression grading. Results In an initial set of 23 patients, responders and nonresponders showed significantly different expression levels for 54 genes (P < .001). The ability to predict response to therapy using gene expression profiles was rigorously evaluated using leave-one-out cross-validation. Tumor behavior was correctly predicted in 83% of patients (P = .02). Sensitivity (correct prediction of response) was 78%, and specificity (correct prediction of nonresponse) was 86%, with a positive and negative predictive value of 78% and 86%, respectively. Conclusion Our results suggest that pretherapeutic gene expression profiling may assist in response prediction of rectal adenocarcinomas to preoperative chemoradiotherapy. The implementation of gene expression profiles for treatment stratification and clinical management of cancer patients requires validation in large, independent studies, which are now warranted.

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