Possible involvement of arylhydrocarbon receptor variants in TCDD-induced thymic atrophy and XRE-dependent transcriptional activity in Wistar Hannover GALAS rats

Wistar Hannover rats, which are maintained by three animal breeders in Japan, were examined to obtain basic data on reproductive and developmental parameters. Untreated pregnant females were terminated on gestational day 20, and the fetuses were removed by cesarian section. The fetuses were counted, weighed and examined for morphological abnormalities. There were few differences among the three stocks of Wistar Hannover rats on the numbers of implantations and live fetuses, sex ratio and fetal weights. The most common fetal abnormalities were the presence of left-sided umbilical arteries, supernumerary ribs and wavy ribs. The incidences of these abnormalities were different among the three stocks of Wistar Hannover rats. Our results provide important data which should be considered in the determination of which stock of rat is used in developmental studies.

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“…1994). Kawakami et al . (2009) found that the BrlHan:Wist@Jcl(GALAS) rat has three genotypes: AhR wt/wt , AhR wt/hw and AhR hw/hw .…”

Section: Resultsmentioning

confidence: 99%

Differences in spontaneous fetal abnormalities among three outbred stocks of Wistar Hannover rats in Japan

Takeuchi¹,

Okuda

Kasahara

et al. 2011

Congenital Anomalies

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“…Reverse transcription was performed with 1 μg of total RNA as the template and the PrimeScript ® RT reagent kit (Takara) according to the manufacturer's instructions. Gene expression was determined using SYBR ® green-based quantitative real-time PCR (Thermal Cycler Dice ® , Takara; Chen et al, 2009;Kawakami et al, 2009) with the following primers: HSF1-F, 5'-TGATGAAG-GGGAAGCAGGAGTG-3', and HSF1-R, 5'-GCTTGTT-GACGACTTTCTGTTGC-3', for the HSF1 gene; and GAPDH-F, 5'-GCACCGTCAAGGCTGAGAAC-3', and GAPDH-R, 5'-TGGTGAAGACGCCAGTGGA-3', for the GAPDH gene. The fold decreases in HSF1 mRNA levels were determined from standard curves after calibration of the assay.…”

Section: Sirna Transfection and Viability Assaymentioning

confidence: 99%

siRNA-mediated silencing of the gene for heat shock transcription factor 1 causes hypersensitivity to methylmercury in HEK293 cells

et al. 2011

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-Methylmercury is a hazardous heavy metal compound that can damage the central nervous system. The mechanisms of methylmercury toxicity and the biological defense mechanisms against it remain unclear. We employed gene knockdown using siRNA to search for transcription factors involved in the manifestation of methylmercury toxicity. We found that the inhibition of expression of the gene for heat shock transcription factor 1 (HSF1) sensitized HEK293 cells to methylmercury.

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“…6 941 cells/well in 6-well plates) were rinsed with 1 × PBS. Then, the total RNA was isolated using a FastPure ® RNA kit (Takara, Shiga, Japan), and first-strand cDNA synthesis was performed using the PrimeScrip ® RT reagent kit (Takara) in accordance with the manufacturer's protocol (Ikawa et al, 2008;Chen et al, 2009;Kawakami et al, 2009). We performed real-time PCR reactions (Thermal Cycler Dice ® , Takara) with the following primers: HOXB13-F, 5'-CTGGGTAGGTGGACAATTG-TA-3', and HOXB13-R, 5'-TACGCTGATGCCTGCT-GTCAA-3', for the HOXB13 gene; and GAPDH-F, 5'-GCACCGTCAAGGCTGAGAAC-3', and GAPDH-R, 5'-TGGTGAAGACGCCAGTGGA-3', for the GAP-DH gene.…”

Section: Toxicogenomics/proteomics Reportmentioning

confidence: 99%

Silencing of the gene for homeobox protein HOXB13 by siRNA confers resistance to methylmercury on HEK293 cells

et al. 2010

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-Methylmercury is an environmental pollutant that causes severe central nervous system disorders. We searched for transcription factors involved in the development of methylmercury toxicity and found that the decreased expression of a homeobox protein, HOXB13, in HEK293 cells conferred strong resistance to methylmercury. Key words: Methylmercury, Homeobox protein, HOXB13, Transcription factorCorrespondence: Akira Naganuma (E-mail: naganuma@mail.pharm.tohoku.ac.jp) Toxicogenomics/proteomics ReportThe Journal of Toxicological Sciences (J. Toxicol. Sci.) Vol.35, No.6, 941-944, 2010 Vol. 35 No. 6 941 cells/well in 6-well plates) were rinsed with 1 × PBS. Then, the total RNA was isolated using a FastPure ® RNA kit (Takara, Shiga, Japan), and first-strand cDNA synthesis was performed using the PrimeScrip ® RT reagent kit (Takara) in accordance with the manufacturer's protocol (Ikawa et al., 2008;Chen et al., 2009;Kawakami et al., 2009). We performed real-time PCR reactions (Thermal Cycler Dice ® , Takara) with the following primers: HOXB13-F, 5'-CTGGGTAGGTGGACAATTG-TA-3', and HOXB13-R, 5'-TACGCTGATGCCTGCT-GTCAA-3', for the HOXB13 gene; and GAPDH-F, 5'-GCACCGTCAAGGCTGAGAAC-3', and GAPDH-R, 5'-TGGTGAAGACGCCAGTGGA-3', for the GAP-DH gene. The fold decreases in HOXB13 mRNA levels were determined from standard curves after calibration of the assay. RESULTS AND DISCUSSIONWe selected about 1,000 proteins that are expected to function as transcription factors from DBD, a transcription factor prediction database (http://www.transcriptionfactor.org). Two double-stranded siRNAs targeting protein-coding genes were designed and introduced into HEK293 cells. The cells with introduced siRNAs were cultured for 48 hr in the presence of methylmercuric chloride (4 μM), which suppresses the proliferation of normal cells by about 60%. HOXB13 was identified as a factor whose reduction conferred strong methylmercury resistance to the cells. We introduced double-stranded siRNAs (siRNA Nos. 1 and 2) targeting HOXB13 into HEK293 cells. Cells harboring HOXB13 siRNA No. 1 exhibited slightly stronger methylmercury resistance than control cells. Cells harboring HOXB13 siRNA No. 2 exhibited marked methylmercury resistance (Fig. 1A). Cells harboring both siRNAs exhibited stronger methylmercury resistance than those harboring HOXB13 siRNA No. 2 alone (Fig. 1A) and the lowest HOXB13 mRNA level (about 16% of the level in control cells; Fig. 1B). HOXB13 mRNA levels in the cells harboring HOXB13 siRNA No. 1 or 2 were about 64% and 25% of the levels in the control cells, respectively (Fig. 1B). These results suggest that HOXB13 increases methylmercury toxicity. Decreased HOXB13 expression had little effect on the susceptibility of the cells to three metallic compounds other than methylmercury (Fig. 2). This suggests that HOXB13 may specifically increase methylmercury toxicity. We introduced HOXB13 with a V5 tag fused at the C terminus (HOXB13-V5) into HEK293 cells to investigate the intracellular distribution of HOXB13 and found HOXB13 predomi...

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Possible involvement of arylhydrocarbon receptor variants in TCDD-induced thymic atrophy and XRE-dependent transcriptional activity in Wistar Hannover GALAS rats

Cited by 14 publications

References 31 publications

Differences in spontaneous fetal abnormalities among three outbred stocks of Wistar Hannover rats in Japan

Differences in spontaneous fetal abnormalities among three outbred stocks of Wistar Hannover rats in Japan

siRNA-mediated silencing of the gene for heat shock transcription factor 1 causes hypersensitivity to methylmercury in HEK293 cells

Silencing of the gene for homeobox protein HOXB13 by siRNA confers resistance to methylmercury on HEK293 cells

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