Gene Expression Analysis of Troglitazone Reveals Its Impact on Multiple Pathways in Cell Culture: A Case for In Vitro Platforms Combined with Gene Expression Analysis for Early (Idiosyncratic) Toxicity Screening

2006 ◽  
Vol 25 (2) ◽  
pp. 85-94 ◽  
Author(s):  
Gordon Vansant ◽  
Patrick Pezzoli ◽  
Robert Saiz ◽  
Aaron Birch ◽  
Chris Duffy ◽  
...  
Keyword(s):  
Gene Expression ◽  
Type 2 Diabetes ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Peroxisome Proliferator ◽  
Toxicity Screening ◽  
The Impact

Peroxisome proliferator-activated receptor gamma (PPAR γ) agonists of the thiazolidinedione family are used for the treatment of type 2 diabetes mellitus due to their ability to reduce glucose and lipid levels in patients with this disease. Three thiazolidinediones that were approved for treatment are Rezulin (troglitazone), Avandia (rosiglitazone), and Actos (pioglitazone). Troglitazone was withdrawn from the market due to idiosyncratic drug toxicity. Rosiglitazone and pioglitazone are still on the market for the treatment of type 2 diabetes. The authors present data from a gene expression screen that compares the impact these three compounds have in rats, in rat hepatocytes, and in the clone 9 rat liver cell line. The authors monitored the changes in expression in multiple genes, including those related to xenobiotic metabolism, proliferation, DNA damage, oxidative stress, apoptosis, and inflammation. Compared to the other two compounds, troglitazone had a significant impact on many of the pathways monitored in vitro although no major perturbation was detected in vivo. The changes detected predict not only general toxicity but potential mechanisms of toxicity. Based on gene expression analysis, the authors propose there is not just one but multiple ways troglitazone could be toxic, depending on a patient’s environment and genetic makeup, including immune response-related toxicity.

scholarly journals Effects of Vitrification on the Blastocyst Gene Expression Profile in a Porcine Model

2021 ◽  
Vol 22 (3) ◽  
pp. 1222
Author(s):  
Cristina Cuello ◽  
Cristina A. Martinez ◽  
Josep M. Cambra ◽  
Inmaculada Parrilla ◽  
Heriberto Rodriguez-Martinez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Analysis ◽  
Survival Rates ◽  
Control Group ◽  
P Value ◽  
Transcriptome Profile ◽  
Embryo Viability ◽  
The Impact

This study was designed to investigate the impact of vitrification on the transcriptome profile of blastocysts using a porcine (Sus scrofa) model and a microarray approach. Blastocysts were collected from weaned sows (n = 13). A total of 60 blastocysts were vitrified (treatment group). After warming, vitrified embryos were cultured in vitro for 24 h. Non-vitrified blastocysts (n = 40) were used as controls. After the in vitro culture period, the embryo viability was morphologically assessed. A total of 30 viable embryos per group (three pools of 10 from 4 different donors each) were subjected to gene expression analysis. A fold change cut-off of ±1.5 and a restrictive threshold at p-value < 0.05 were used to distinguish differentially expressed genes (DEGs). The survival rates of vitrified/warmed blastocysts were similar to those of the control (nearly 100%, n.s.). A total of 205 (112 upregulated and 93 downregulated) were identified in the vitrified blastocysts compared to the control group. The vitrification/warming impact was moderate, and it was mainly related to the pathways of cell cycle, cellular senescence, gap junction, and signaling for TFGβ, p53, Fox, and MAPK. In conclusion, vitrification modified the transcriptome of in vivo-derived porcine blastocysts, resulting in minor gene expression changes.

scholarly journals The Wnt signaling pathway effector TCF7L2 is upregulated by insulin and represses hepatic gluconeogenesis

2012 ◽  
Vol 303 (9) ◽  
pp. E1166-E1176 ◽  
Author(s):  
Wilfred Ip ◽  
Weijuan Shao ◽  
Yu-ting Alex Chiang ◽  
Tianru Jin
Keyword(s):  
Gene Expression ◽  
Type 2 Diabetes ◽  
Transcription Factor ◽  
Wnt Signaling ◽  
Wnt Signaling Pathway ◽  
Glucose Production ◽  
Hepatic Gluconeogenesis

Certain single nucleotide polymorphisms (SNPs) in transcription factor 7-like 2 (TCF7L2) are strongly associated with the risk of type 2 diabetes. TCF7L2 and β-catenin (β-cat) form the bipartite transcription factor cat/TCF in stimulating Wnt target gene expression. cat/TCF may also mediate the effect of other signaling cascades, including that of cAMP and insulin in cell-type specific manners. As carriers of TCF7L2 type 2 diabetes risk SNPs demonstrated increased hepatic glucose production, we aimed to determine whether TCF7L2 expression is regulated by nutrient availability and whether TCF7L2 and Wnt regulate hepatic gluconeogenesis. We examined hepatic Wnt activity in the TOPGAL transgenic mouse, assessed hepatic TCF7L2 expression in mice upon feeding, determined the effect of insulin on TCF7L2 expression and β-cat Ser675 phosphorylation, and investigated the effect of Wnt activation and TCF7L2 knockdown on gluconeogenic gene expression and glucose production in hepatocytes. Wnt activity was observed in pericentral hepatocytes in the TOPGAL mouse, whereas TCF7L2 expression was detected in human and mouse hepatocytes. Insulin and feeding stimulated hepatic TCF7L2 expression in vitro and in vivo, respectively. In addition, insulin activated β-cat Ser675 phosphorylation. Wnt activation by intraperitoneal lithium injection repressed hepatic gluconeogenic gene expression in vivo, whereas lithium or Wnt-3a reduced gluconeogenic gene expression and glucose production in hepatic cells in vitro. Small interfering RNA-mediated TCF7L2 knockdown increased glucose production and gluconeogenic gene expression in cultured hepatocytes. These observations suggest that Wnt signaling and TCF7L2 are negative regulators of hepatic gluconeogenesis, and TCF7L2 is among the downstream effectors of insulin in hepatocytes.

375 Gene expression analysis of galactosamine-induced hepatotoxicity in-vivo and in-vitro

2003 ◽  
Vol 144 ◽  
pp. s102
Author(s):  
H. Hildebrand ◽  
G. Kempka ◽  
H. Ellinger ◽  
B. Stuart ◽  
B. Wahle ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Analysis ◽  
Gene Expression Analysis

Role of the snake venom toxin jararhagin in proinflammatory pathogenesis: In vitro and in vivo gene expression analysis of the effects of the toxin

2005 ◽  
Vol 441 (1) ◽  
pp. 1-15 ◽  
Author(s):  
Paul Gallagher ◽  
Yongde Bao ◽  
Solange M.T. Serrano ◽  
Gavin D. Laing ◽  
R. David G. Theakston ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Analysis ◽  
Snake Venom ◽  
Gene Expression Analysis ◽  
Venom Toxin ◽  
Snake Venom Toxin

Human and murine amniotic fluid c-Kit+Lin− cells display hematopoietic activity

Blood ◽  
2009 ◽  
Vol 113 (17) ◽  
pp. 3953-3960 ◽  
Author(s):  
Andrea Ditadi ◽  
Paolo de Coppi ◽  
Olivier Picone ◽  
Laetitia Gautreau ◽  
Rim Smati ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Amniotic Fluid ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Lymphoid Cells ◽  
Immunocompromised Hosts ◽  
Hematopoietic Activity

Abstract We have isolated c-Kit+Lin− cells from both human and murine amniotic fluid (AF) and investigated their hematopoietic potential. In vitro, the c-Kit+Lin− population in both species displayed a multilineage hematopoietic potential, as demonstrated by the generation of erythroid, myeloid, and lymphoid cells. In vivo, cells belonging to all 3 hematopoietic lineages were found after primary and secondary transplantation of murine c-Kit+Lin− cells into immunocompromised hosts, thus demonstrating the ability of these cells to self-renew. Gene expression analysis of c-Kit+ cells isolated from murine AF confirmed these results. The presence of cells with similar characteristics in the surrounding amnion indicates the possible origin of AF c-Kit+Lin− cells. This is the first report showing that cells isolated from the AF do have hematopoietic potential; our results support the idea that AF may be a new source of stem cells for therapeutic applications.

scholarly journals Antimicrobial Effect of Roselle (Hibiscus sabdariffa L.) Water Fraction against Pseudomonas aeruginosa using Drosophila Infection Model

2021 ◽  
Vol 11 (5) ◽  
pp. 12877-12885
Keyword(s):  
Gene Expression ◽  
Pseudomonas Aeruginosa ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Mrna Levels ◽  
Antibacterial Compounds ◽  
Water Fraction ◽  
Survival Assay ◽  
The Impact

Pseudomonas aeruginosa is one of the most common pathogenic bacteria that cause nosocomial infection. Unfortunately, the irrational use of antibiotics has created a surge in P. aeruginosa resistance nowadays. To overcome this situation, new antibacterial compounds are urgently needed. One of the potential sources to obtain such antibacterial compounds is roselle calyx. This research was carried out using two experimental approaches, survival assay and gene expression analysis, to examine the in vivo antibacterial effect of water fraction of roselle calyx (WFR) against Pseudomonas aeruginosa in Drosophila model of infection. Survival assay was used to demonstrate the impact of treatment on the lifespan of the infected host. The measurement of immune-related Dpt mRNA levels by reverse-transcriptase quantitative PCR (RT-qPCR) was used to assess whether immunostimulation is involved in the antibacterial protection of WFR against P. aeruginosa. The result demonstrated that WFR at concentrations of 0.8% and 2% were able to enhance P. aeruginosa-infected flies' survival. Furthermore, gene expression analysis showed the insignificant difference between WFR-treated flies and healthy control flies at all tested concentrations, implying the non-involvement of Imd-Dpt-mediated pathway immunity in the antipseudomonal protection of WFR. Taken together, our data suggested the in vivo antibacterial activity of WFR against P. aeruginosa in the fruit fly model of infection.

scholarly journals New Shuttle Vectors for Real-Time Gene Expression Analysis in Multidrug-Resistant Acinetobacter Species: In Vitro and In Vivo Responses to Environmental Stressors

2019 ◽  
Vol 85 (18) ◽  
Author(s):  
Massimiliano Lucidi ◽  
Daniela Visaggio ◽  
Elisa Prencipe ◽  
Francesco Imperi ◽  
Giordano Rampioni ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Multidrug Resistant ◽  
Antibiotic Selection ◽  
Content Type ◽  
Shuttle Vectors ◽  
Reporter Systems

ABSTRACT The Acinetobacter genus includes species of opportunistic pathogens and harmless saprophytes. The type species, Acinetobacter baumannii, is a nosocomial pathogen renowned for being multidrug resistant (MDR). Despite the clinical relevance of infections caused by MDR A. baumannii and a few other Acinetobacter spp., the regulation of their pathogenicity remains elusive due to the scarcity of adequate genetic tools, including vectors for gene expression analysis. Here, we report the generation and testing of a series of Escherichia coli-Acinetobacter promoter-probe vectors suitable for gene expression analysis in Acinetobacter spp. These vectors, named pLPV1Z, pLPV2Z, and pLPV3Z, carry both gentamicin and zeocin resistance markers and contain lux, lacZ, and green fluorescent protein (GFP) reporter systems downstream of an extended polylinker, respectively. The presence of a toxin-antitoxin gene system and the high copy number allow pLPV plasmids to be stably maintained even without antibiotic selection. The pLPV plasmids can easily be introduced by electroporation into MDR A. baumannii belonging to the major international lineages as well as into species of the Acinetobacter calcoaceticus-A. baumannii complex. The pLPV vectors have successfully been employed to study the regulation of stress-responsive A. baumannii promoters, including the DNA damage-inducible uvrABC promoter, the ethanol-inducible adhP and yahK promoters, and the iron-repressible promoter of the acinetobactin siderophore biosynthesis gene basA. A lux-tagged A. baumannii ATCC 19606T strain, carrying the iron-responsive pLPV1Z::PbasA promoter fusion, allowed in vivo and ex vivo monitoring of the bacterial burden in the Galleria mellonella infection model. IMPORTANCE The short-term adaptive response to environmental cues greatly contributes to the ecological success of bacteria, and profound alterations in bacterial gene expression occur in response to physical, chemical, and nutritional stresses. Bacteria belonging to the Acinetobacter genus are ubiquitous inhabitants of soil and water though some species, such as Acinetobacter baumannii, are pathogenic and cause serious concern due to antibiotic resistance. Understanding A. baumannii pathobiology requires adequate genetic tools for gene expression analysis, and to this end we developed user-friendly shuttle vectors to probe the transcriptional responses to different environmental stresses. Vectors were constructed to overcome the problem of antibiotic selection in multidrug-resistant strains and were equipped with suitable reporter systems to facilitate signal detection. By means of these vectors, the transcriptional response of A. baumannii to DNA damage, ethanol exposure, and iron starvation was investigated both in vitro and in vivo, providing insights into A. baumannii adaptation during stress and infection.

129 POST-HATCHING DEVELOPMENT AND GENE EXPRESSION OF IN VITRO-PRODUCED BOVINE EMBRYOS IN RECIPIENT UTERUS AFTER MULTIPLE TRANSFER

2013 ◽  
Vol 25 (1) ◽  
pp. 212
Author(s):  
G. Machado ◽  
A. Ferreira ◽  
I. Pivato ◽  
A. Fidelis ◽  
J. F. Srpicigo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
T Test ◽  
Molecular Profile ◽  
Control Group ◽  
Fluid Medium ◽  
Uterine Flushing

This study aimed to compare post-hatching development of Day 7 in vitro and in vivo embryos cultured in recipient uterus until Day 14. For producing in vitro embryos (IVP), oocytes were matured, fertilized (Day 0) and cultured in vitro for 6 days (Day 7) in synthetic oviduct fluid medium supplemented with 5% of fetal bovine serum and incubated at 39°C in 5% CO2 in air. At Day 7, part of IVP blastocysts was transferred to recipient uterus and part was stored for gene expression analysis. As a control group, in vivo embryos were produced after ovarian stimulation, insemination and uterine flushing on Day 7 post insemination. Similarly to the IVP embryos, part of embryos was transferred to recipient uterus and part was stored for gene expression analysis. Day 7 in vivo (n = 53) and IVP (n = 64) expanded blastocysts were transferred to synchronized recipients (10/horn) and were collected by uterine flushing 7 days after transfer (Day 14). Recovered embryos were measured using Motic Image Plus software and evaluated for presence and size of embryonic disc (ED). A trophoblast biopsy was removed and stored for gene expression analysis. For the molecular profile evaluation of Day 7 and Day 14 in vivo and in vitro embryos, 8 genes related with placentation, implantation, oxidative stress, and glucose metabolism (PLAC8, CD9, GLUT-1, GLUT-3, KRT8, MnSOD, HSP70, and INFT, respectively) were quantified by RT-qPCR using ΔΔCT method and CYC-A gene as endogenous control. The recovery rate of Day 14 embryos, analyzed by chi-square test, was higher (P < 0.05) for in vitro than for in vivo embryos, being 50.0% (64/128) and 38.6% (53/137), respectively. No differences (P > 0.05; t-test) were observed in embryo length when comparing Day 14 in vitro (19.1 ± 2.4 mm) and in vivo embryos (24.2 ± 3.7 mm). ED was detected in 25% (16/64) of in vitro and in 26% (14/53) of in vivo embryos. No differences were found (P > 0.05; t-test) in diameter between the two types of embryos (0.3 ± 0.0 mm/in vitro and 0.3 ± 0.0 mm/in vivo). Regarding gene expression, Day 7 IVP embryos showed higher (P < 0.05, Mann–Whitney test) expression of HSP70 and SCL2A1 than in vivo embryos. However, at Day 14 no differences between embryos were observed in transcript levels for any of the studied genes. Therefore, the present study showed that although differences in Day 7 in vitro embryos were observed at the molecular level compared to in vivo counterpart, after transfer to the uterine environment, they showed similar morphology and gene expression profile. These results highlight the importance of evaluating embryos produced by assisted reproductive techniques in later stages of development to have a more precise evaluation of their quality. Financial support: Embrapa, CNPq, CAPES.

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