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Gene expression analyses reveal potential mechanism of inorganic arsenic-induced apoptosis in zebrafish

JOURNAL OF APPLIED TOXICOLOGY 2023年第12期43卷 1872-1882页

作者： Silva, Camila S. S. Kudlyk, Tetyana Tryndyak, Volodymyr P. P. Twaddle, Nathan C. C. Robinson, Bonnie Gu, Qiang Beland, Frederick A. A. Fitzpatrick, Suzanne C. C. Kanungo, Jyotshna US FDA Div Biochem Toxicol Natl Ctr Toxicol Res Jefferson AR 72079 USA US FDA Div Neurotoxicol Natl Ctr Toxicol Res 3900 NCTR Rd Jefferson AR 72079 USA US FDA Off Ctr Director Ctr Food Safety & Appl Nutr College Pk MD USA

Our previous study showed that sodium arsenite (200 mg/L) affected the nervous system and induced motor neuron development via the Sonic hedgehog pathway in zebrafish larvae. To gain more insight into the effects of arsenite on other signaling pathways, including apoptosis, we have performed quantitative polymerase chain reaction array-based gene expression analyses. The 96-well array plates contained primers for 84 genes representing 10 signaling pathways that regulate several biological functions, including apoptosis. We exposed eggs at 5 h postfertilization until the 72 h postfertilization larval stage to 200 mg/L sodium arsenite. In the Janus kinase/signal transducers and activators of transcription, nuclear factor & kappa;-light-chain-enhancer of activated B cells, and Wingless/Int-1 signaling pathways, the expression of only one gene in each pathway was significantly altered. The expression of multiple genes was altered in the p53 and oxidative stress pathways. Sodium arsenite induced excessive apoptosis in the larvae. This compelled us to analyze specific genes in the p53 pathway, including cdkn1a, gadd45aa, and gadd45ba. Our data suggest that the p53 pathway is likely responsible for sodium arsenite-induced apoptosis. In addition, sodium arsenite significantly reduced global DNA methylation in the zebrafish larvae, which may indicate that epigenetic factors could be dysregulated after arsenic exposure. Together, these data elucidate potential mechanisms of arsenic toxicity that could improve understanding of arsenic's effects on human health.

关键词： apoptosis arsenic DNA methylation gene expression pcr array zebrafish

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Investigation of miRNAs that are effective in the pathogenesis of asthma

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JOURNAL OF ASTHMA 2023年第12期60卷 2145-2152页

作者： Coskunpinar, Ender Akcesme, Betul Tas, Sevgi Kalkanli Aynaci, Aysun Univ Hlth Sci Sch Med Dept Med Biol Istanbul Turkiye Univ Hlth Sci Sch Med Dept Immunol Istanbul Turkiye Istanbul Kanuni Sultan Suleyman Training & Res Ho Clin Chest Dis Istanbul Turkiye Univ Hlth Sci Sch Med Dept Med Biol Tibbiye Cad 38 TR-34668 Istanbul Turkiye

ObjectivesAsthma is a complex disease characterized by inflammation of the airways, involving epigenetic changes, in which genetic and environmental factors act together. MicroRNAs as candidate biomarkers stand out as target molecules in the diagnosis and treatment of immunological and inflammatory diseases. Our aim of this study is to identify miRNAs that are thought to be effective in the pathogenesis of allergic asthma and to reveal candidate biomarkers associated with the *** patients, aged between 18-80 years, who were diagnosed with allergic asthma and 18 healthy volunteers were included in the study. After the collection 2 mL of total blood from volunteers, RNA isolation and cDNA synthesis were performed. For miRNA profile screening, expression analysis was performed by real-time pcr method using miScript miRNA pcr array. GeneGlobe Data Analysis Center was used to evaluate dysregulated *** the allergic asthma group, 9 (18%) of the patients were male and 41 (82%) of them were female. In the control group, 7 (38.89%) were male and 11 (61.1%) were female (P:0.073). As a result of the research, the expression levels of miR-142-5p, miR-376c-3p and miR-22-3p were down-regulated, while miR-27b-3p, miR-26b-5p, miR-15b-5p and miR-29c-3p detected as *** results of our study suggest that miR142-5p, miR376c-3p and miR22-3p promote Ubiquitin-mediated proteolysis by inhibiting TGF-& beta;expression through a mechanism involving the p53 signaling pathway. The deregulated miRNAs may be used as a diagnostic and prognostic biomarker in asthma.

关键词： Asthma miRNA biomarker pcr array

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pcr array analysis of gene expression profiles in chronic active Epstein- Barr virus infection

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MICROBES AND INFECTION 2014年第7期16卷 581-586页

作者： Murakami, Masanao Hashida, Yumiko Imajoh, Masayuki Maeda, Akihiko Kamioka, Mikio Senda, Yasutaka Sato, Tetsuya Fujieda, Mikiya Wakiguchi, Hiroshi Daibata, Masanori Kochi Univ Kochi Med Sch Dept Microbiol & Infect Kochi 7838505 Japan Kochi Univ Kochi Med Sch Dept Pediat Kochi 7838505 Japan Kochi Univ Kochi Med Sch Dept Lab Med Kochi 7838505 Japan Kochi Univ Kochi Med Sch Fac Med Kochi 7838505 Japan

To determine the host cellular gene expression profiles in chronic active Epstein Barr virus infection (CAEBV), peripheral blood samples were obtained from three patients with CAEBV and investigated using a pcr array analysis that focused on T-cell/B-cell activation. We identified six genes with expression levels that were tenfold higher in CAEBV patients compared with those in healthy controls. These results were verified by quantitative reverse transcription-pcr. We identified four highly upregulated genes, i.e., IL-10, IL-2, IFNGRI, and INHBA. These genes may be involved in inflammatory responses and cell proliferation, and they may contribute to the development and progression of CAEBV. (C) 2014 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

关键词： Epstein-Barr virus CAEBV Gene profile pcr array Cytokine

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Molecular toxicological alterations in the mouse hearts induced by sub-chronic thiazolidinedione drugs administration

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FUNDAMENTAL & CLINICAL PHARMACOLOGY 2022年第1期36卷 143-149页

作者： Jarrar, Yazun Bashir Jarrar, Qais Abaalkhail, Sara J. Moh'd Kalloush, Hanin Naser, Wisam Zihlif, Malek Al Shhab, Mohammad El Madani, Abdulla Jamous, Yahya Lee, Su-Jun Al Zaytoonah Univ Jordan Dept Pharm Amman Jordan Al Israa Univ Dept Pharmaceut Sci Amman Jordan Univ Jordan Fac Med Dept Pharmacol Amman Jordan King AbdulAziz City Sci & Technol Nanotechnol Ctr Riyadh Saudi Arabia Inje Univ Coll Med Dept Pharmacol Pusan South Korea

Thiazolidinediones are well-known anti-diabetic drugs. However, they are not widely used due to their cardiotoxic effects. Therefore, in this study, we aimed to determine the molecular toxicological alterations induced in the mouse hearts after thiazolidinedione administration. Balb/c mice received doses clinically equivalent to those given to humans of the most commonly used thiazolidinediones, pioglitazone, and rosiglitazone for 30 days. After that, RNA samples were isolated from the hearts. The mRNA expression of cytochrome (cyp) p450 genes that synthesize the cardiotoxic 20-hydroxyeicosatetraenoic acid (20-HETE) in addition to 92 cardiotoxicity biomarker genes were analyzed using quantitative polymerase chain reaction array technique. The analysis demonstrated that thiazolidinediones caused a significant upregulation (p < 0.5) of the mRNA expression of cyp1a1, cyp4a12, itpr1, ccl7, ccr1, and b2 m genes. In addition, thiazolidinediones caused a significant (p < 0.05) downregulation of the mRNA expression of adra2a, bsn, col15a1, fosl1, Il6, bpifa1, plau, and reg3b genes. The most affected gene was itpr1 gene, which was upregulated by pioglitazone and rosiglitazone by sevenfold and 3.5-fold, respectively. In addition, pioglitazone caused significant upregulation of (p < 0.05) hamp, ppbp, psma2, sik1, timp1, and ucp1 genes, which were not affected significantly (p > 0.05) by rosiglitazone administration. In conclusion, this study showed that thiazolidinediones induce toxicological molecular alterations in the mouse hearts, such as the induction of cyp450s that synthesize 20-HETE, chemokine activation, inflammatory responses, blood clotting, and oxidative stress. These findings may help us understand the mechanism of cardiotoxicity involved in thiazolidinedione administration.

关键词： 20‐ HETE cardiotoxicity molecular alteration pcr array thiazolidinediones

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The Stage-specific Testicular Germ Cell Apoptotic Response to Low-dose X-irradiation and 2,5-hexanedione Combined Exposure. I: Validation of the Laser Capture Microdissection Method for qRT-pcr array Application

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TOXICOLOGIC PATHOLOGY 2014年第8期42卷 1221-1228页

作者： Catlin, Natasha R. Huse, Susan M. Boekelheide, Kim Brown Univ Dept Pathol & Lab Med Providence RI 02912 USA

Over the past decade, laser capture microdissection (LCM) has grown as a tool for gene expression profiling of small numbers of cells from tumor samples and of specific cell populations in complex tissues. LCM can be used to study toxicant effects on selected cell populations within the testis at different stages of spermatogenesis. There are several LCM-related hurdles to overcome, including issues inherent to the method itself, as well as biases that result from amplifying the LCM-isolated RNA. Many technical issues associated with the LCM method are addressed here, including increasing RNA yield and obtaining more accurate quantification of RNA yields. We optimized the LCM method optimized to generate RNA quantities sufficient for quantitative reverse transcription polymerase chain reaction (qRT-pcr) array analysis without amplification and were able to validate the method through direct comparison of results from unamplified and amplified RNA from individual samples. The addition of an amplification step for gene expression studies using LCM RNA resulted in a bias, especially for low abundance transcripts. Although the amplification bias was consistent across samples, researchers should use caution when comparing results generated from amplified and unamplified LCM RNA. Here, we have validated the use of LCM-derived RNA with the qRT-pcr array, improving our ability to investigate cell-type and stage-specific responses to toxicant exposures.

关键词： laser capture microdissection testis amplification pcr array

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Primate-specific 82-kDa choline acetyltransferase attenuates progression of Alzheimer's disease-like pathology in the APP^NL-G-F knock-in mouse model

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SCIENTIFIC REPORTS 2024年第1期14卷 1-17页

作者： Alqot, Hadir E. Rylett, Rebecca Jane Univ Western Ontario Robarts Res Inst Schulich Sch Med & Dent Dept Physiol & Pharmacol London ON N6A 5K8 Canada

Alzheimer's disease (AD) is characterized by amyloidosis, neuroinflammation, cholinergic dysfunction and cognitive impairment. In AD, the cholinergic neuronal marker choline acetyltransferase (ChAT) is reduced and the primate-specific nuclear isoform, 82-kDa ChAT, is mislocalized to cytoplasm. Cell-based studies suggest a role for 82-kDa ChAT in regulating expression of AD-related genes with potential reductions in beta-amyloid (A beta) levels. To study this further, we crossed transgenic mice expressing human 82-kDa ChAT with the AD mouse model APP(NL-G-F) and used molecular techniques and neurobehavioral tests to study the impact of 82-kDa ChAT on AD pathology. These mice had altered expression of genes linked to A beta clearance and inflammation, and reduced cognitive decline, amyloidosis and gliosis. These effects were inversely related to age and A beta plaque load and correlated best with 82-kDa ChAT localized to nuclei of neurons. The study suggests a role for 82-kDa ChAT in decreasing AD-like pathology.

关键词： 82-kDa choline acetyltransferase APP(NL-G-F) mice Alzheimer's disease beta-Amyloid clearance Microglia Neurobehavioral testing pcr array Immunofluorescence

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Focused cancer pathway analysis revealed unique therapeutic targets in retinoblastoma

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MEDICAL ONCOLOGY 2024年第7期41卷 168-168页

作者： Balaji, Sekaran Rao, Anindita Saraswathi, Karuvel Kannan Nagarajan, Rathinavel Sethu Santhi, Radhakrishnan Kim, Usha Muthukkaruppan, Veerappan Vanniarajan, Ayyasamy Aravind Med Res Fdn Dept Mol Genet 1 Anna Nagar Madurai 625020 Tamil Nadu India Aravind Eye Hosp Dept Pathol Madurai 625020 Tamil Nadu India Aravind Eye Hosp Dept Orbit Oculoplasty & Ocular Oncol Madurai 625020 Tamil Nadu India Aravind Med Res Fdn Dept Immunol & Stem Cell Biol Madurai 625020 Tamil Nadu India Alagappa Univ Dept Mol Biol Aravind Med Res Fdn Karaikkudi 630003 Tamil Nadu India

Retinoblastoma (RB) is a pediatric cancer of the eye that occurs in 1/15000 live births worldwide. Albeit RB is initiated by the inactivation of RB1 gene, the disease progression relies largely on transcriptional alterations. Therefore, evaluating gene expression is vital to unveil the therapeutic targets in RB management. In this study, we employed an RT2 Profiler (TM) pcr array for a focused analysis of 84 cancer-specific genes in RB. An interaction network was built with gene expression data to identify the dysregulated pathways in RB. The key transcript alterations identified in 13 tumors by RT2 Profiler (TM) pcr array was further validated in 15 tumors by independent RT-qpcr. Out of 84 cancer-specific genes, 68 were dysregulated in RB tumors. Among the 68 genes, 23 were chosen for further analysis based on statistical significance and abundance across multiple tumors. Pathway analysis of altered genes showed the frequent perturbations of cell cycle, angiogenesis and apoptotic pathways in RB. Notably, upregulation of MCM2, MKI67, PGF, WEE1, CDC20 and downregulation of COX5A were found in all the tumors. Western blot confirmed the dysregulation of identified targets at protein levels as well. These alterations were more prominent in invasive RB, correlating with the disease pathogenesis. Our molecular analysis thus identified the potential therapeutic targets for improving retinoblastoma treatment. We also suggest that pcr array can be used as a tool for rapid and cost-effective gene expression analysis.

关键词： Retinoblastoma pcr array Pathway analysis Gene expression Cancer-related pathways Therapeutic targets

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Cumulus cell microRNA expression when LH is added to the ovarian stimulation protocol: a pilot study

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REPRODUCTIVE BIOMEDICINE ONLINE 2021年第6期43卷 1070-1077页

作者： Vigo, Francieli Maria Fraietta, Renato Rodrigues, Fernanda Carvalho, Cristina, V Bonetti, Tatiana C. S. Motta, Eduardo L. A. Univ Fed Sao Paulo Dept Ginecol Escola Paulista Med UNIFESP EPM Rua Napoleao Barros608 Vila Clementino BR-04024002 Sao Paulo SP Brazil Univ Fed Sao Paulo Setor Integrad Reprod Humana UNIFESP EPM Escola Paulista MedDept Cirurgia Rua Botucatu725 Vila Vila Clementino BR-04024002 Sao Paulo SP Brazil Huntington Med Reprod Av Republ Libano 529 Ibirapuera BR-04501000 Sao Paulo SP Brazil

Research question: Recombinant FSH administration in ovarian stimulation for IVF is a standard procedure, whereas the role of LH is controversial. MicroRNAs (mRNA) are small endogenous non-coding transcripts that are involved in the regulation of many cellular processes, including foliculogenesis and gonadotrophin function. The aim was to study the possible role of miRNA in ovarian follicular development in groups having different ovarian stimulation protocols. Are there different miRNA expression profiles in cumulus cells of infertile women undergoing IVF? What are the regulated pathways? Design: This prospective observational study included 13 patients who fulfilled the following inclusion criteria: younger than 38 years of age;a tubal infertility factor;a male factor;or idiopathic infertility. This is a pilot study in which the patients were aleatory enrolled into two groups: seven in FSH group (recombinant FSH, 225 IU) and six in FSH plus LH group (recombinant FSH, 150 IU + recombinant LH, 75 IU). The granulosa cells obtained from the follicular ovarian retrieval were analysed using polyerase chain reaction. Results were analysed using DIANA Tools, an online bioinformatics tool. Results: Among the 84 microRNAs evaluated, 11 were differentially expressed between the groups, all of which were upregulated in the FSH plus LH group, compared with the FSH group. Differentially expressed miRNA profiles are related to oestrogen signalling, oocyte meiosis and pluripotent cells regulation. Conclusion: miRNA overexpression in the FSH plus LH group is consistent with the independent and fundamental role of LH in folliculogenesis, leading to a distinct molecular response between groups.

关键词： MicroRNA Ovarian stimulation pcr array Recombinant LH

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Effects of acetyl L-carnitine on zebrafish embryos: Phenotypic and gene expression studies

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JOURNAL OF APPLIED TOXICOLOGY 2021年第2期41卷 256-264页

作者： Gu, Qiang Ali, Syed F. Kanungo, Jyotshna US FDA Div Neurotoxicol Natl Ctr Toxicol Res 3900 NCTR Rd Jefferson AR 72079 USA

Acetyl L-carnitine (ALCAR), a dietary supplement and an antioxidant, plays a vital role in the bioenergetic process that produces ATP. Although there are reports on antioxidant toxicity, there is no information on the potential toxicity of ALCAR. Here, using zebrafish embryos, we explored whether ALCAR modulated ATP synthesis, generation of reactive oxygen species (ROS) and expression of specific genes related to major signaling pathways that control metabolism, growth, differentiation, apoptosis and oxidative stress. First, we show that ALCAR elicits a physiologic response, as ATP levels increased after ALCAR treatment. Simultaneously, an increase in the expression of ROS, a by-product of ATP synthesis, was observed in the ALCAR-treated embryos. Consistent with higher ROS expression, the level of cysteine, a precursor of glutathione, was significantly reduced. ALCAR did not have any drastic effect on overall development and heart rate. Polymerase chain reaction-based gene expression array analyses showed no significant change in the expression of 83 genes related to 10 major signaling pathways including: the transforming growth factor beta (TGF beta), Wingless and Int-1 (Wnt), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kappa B), Janus kinase/signal transducers and activators of transcription (JAK/STAT), p53, Notch, Hedgehog, Peroxisome proliferator-activated receptor (PPAR), oxidative stress, and hypoxia pathways. Our results show that the expression of 83 genes related to these major signaling pathways did not change significantly.

关键词： acetyl L-carnitine developmental toxicity pcr array reactive oxygen species (ROS) zebrafish

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Superior mechanical recovery in male and female MRL/MpJ tendons is associated with a unique genetic profile

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JOURNAL OF ORTHOPAEDIC RESEARCH 2021年第6期39卷 1344-1354页

作者： George, Nisha S. Bell, Rebecca Paredes, J. J. Taub, Peter J. Andarawis-Puri, Nelly Icahn Sch Med Mt Sinai Leni & Peter W May Dept Orthopaed New York NY 10029 USA Cornell Univ Sibley Sch Mech & Aerosp Engn Ithaca NY 14853 USA Cornell Univ Nancy E & Peter C Meinig Sch Biomed Engn Ithaca NY USA Hosp Special Surg Res Div New York NY USA

Tendon ruptures heal by forming a mechanically inferior scar. We have shown that male Murphy Roths large (MRL/MpJ) mice exhibit improved tendon healing, suggesting that they can inform biological mechanisms that lead to effective tendon healing. As sex impacts healing, we assessed the effect of sex on tendon healing in MRL/MpJ and normal healer C57BL/6 (B6) mice and compared the associated biological environment with identify genes that may be integral to the improved healing outcome. We hypothesized that (a) male MRL/MpJ mice will heal with improved mechanical properties compared to females;and (b) that regenerative tendon healing will be associated with decreased fibrotic pathways, decreased inflammation, and increased activity of matrix metalloproteinases (MMPs). A midsubstance punch was introduced, and tendons were harvested after (a) 1 or 7 days for profiling of 84 genes;(b) 7 or 14 days for the assessment of MMP-2 and MMP-9 activity;and (c) 6 weeks for mechanical assessment. MRL/MpJ tendons healed with the better restoration of mechanical properties than B6 tendons. Sex did not affect the mechanical properties of healing B6 or MRL/MpJ tendons. Comparison of the gene expression profiles in the context of the mechanical outcome revealed several differences between MRL/MpJ and B6 tendon healing, including, lower inflammation, an earlier higher expression of TGF-beta-related genes that diminish by 7 days, and genes associated with enhanced cell migration in MRL/MpJ in comparison to B6 tendons. We expect that the timecourse and expression levels of these genes in scarless MRL/MpJ tendon healing represent the balanced environment that leads to improved tendon healing.

关键词： pcr array regeneration scarless healing tendon injury

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