Publications by authors named "Toshinori Yoshida"

120 Publications

Disruption of postnatal neurogenesis and adult-stage suppression of synaptic plasticity in the hippocampal dentate gyrus after developmental exposure to sterigmatocystin in rats.

Toxicol Lett 2021 Oct 11;349:69-83. Epub 2021 Jun 11.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan; Cooperative Division of Veterinary Sciences, Graduate School of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan; Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan. Electronic address:

Exposure to sterigmatocystin (STC) raises concerns on developmental neurological disorders. The present study investigated the effects of maternal oral STC exposure on postnatal hippocampal neurogenesis of offspring in rats. Dams were exposed to STC (1.7, 5.0, and 15.0 ppm in diet) from gestational day 6 until day 21 post-delivery (weaning), and offspring were maintained without STC exposure until adulthood on postnatal day (PND) 77, in accordance with OECD chemical testing guideline Test No. 426. On PND 21, 15.0-ppm STC decreased type-3 neural progenitor cell numbers in the subgranular zone (SGZ) due to suppressed proliferation. Increased γ-H2AX-immunoreactive () cell numbers in the SGZ and Ercc1 upregulation and Brip1 downregulation in the dentate gyrus suggested induction of DNA double-strand breaks in SGZ cells. Upregulation of Apex1 and Ogg1 and downregulation of antioxidant genes downstream of NRF2-Keap1 signaling suggested induction of oxidative DNA damage. Increased p21 SGZ cell numbers and suppressed cholinergic signaling through CHRNB2-containing receptors in GABAergic interneurons suggested potential neurogenesis suppression mechanisms. Multiple mechanisms involving N-methyl-d-aspartate (NMDA) receptor-mediated glutamatergic signaling and various GABAergic interneuron subpopulations, including CHRNA7-expressing somatostatin interneurons activated by BDNF-TrkB signaling, may be involved in ameliorating the neurogenesis. Upregulation of Arc, Ptgs2, and genes encoding NMDA receptors and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors suggested synaptic plasticity facilitation. On PND 77, ARC granule cells decreased, and Nos2 was upregulated following 15.0 ppm STC exposure, suggesting oxidative stress-mediated synaptic plasticity suppression. Inverse pattern in gene expression changes in vesicular glutamate transporter isoforms, Slc17a7 and Slc17a6, from weaning might also be responsible for the synaptic plasticity suppression. The no-observed-adverse-effect level of maternal oral STC exposure for offspring neurogenesis was determined to be 5.0 ppm, translating to 0.34-0.85 mg/kg body weight/day.
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http://dx.doi.org/10.1016/j.toxlet.2021.06.006DOI Listing
October 2021

Anti-tumor effect of trametinib in bladder cancer organoid and the underlying mechanism.

Cancer Biol Ther 2021 May 26:1-15. Epub 2021 May 26.

Laboratory of Veterinary Pharmacology, Department of Veterinary Medicine, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Fuchu, Japan.

Bladder cancer (BC), a main neoplasm of urinary tract, is usually inoperable and unresponsive to chemotherapy. As a novel experimental model for muscle-invasive BC, we previously established a culture method of dog BC organoids. In the present study, the detailed in vitro and in vivo anti-tumor effects of trametinib were investigated by using this model. In each BC organoid strain, epidermal growth factor receptor (EGFR)/ERK signaling was upregulated compared with normal bladder cells. Trametinib even at a low concentration inhibited the cell viability of BC organoids and the activation of ERK through decreasing expression of c-Myc, ELK1, SIK1, and PLA2G4A. Trametinib arrested cell cycle of BC with few apoptosis. Dual treatment of BC organoids with trametinib and YAP inhibitor, verteporfin extremely inhibited the cell viability with apoptosis induction. Moreover, trametinib induced basal to luminal differentiation of BC organoids by upregulating luminal markers and downregulating basal ones. In vivo, trametinib decreased the tumor growth of BC organoids in mice and the xenograft-derived organoids from trametinib-administered mice showed enhanced sensitivity to carboplatin due to MSH2 upregulation. Our data suggested a new strategy of trametinib-YAP inhibitor or trametinib-carboplatin combination as a promising treatment of BC.
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http://dx.doi.org/10.1080/15384047.2021.1919004DOI Listing
May 2021

Squamous cell carcinoma in a digit of the hind limb with systemic metastasis in a 17-year-old female koala.

J Vet Med Sci 2021 Jul 26;83(6):994-996. Epub 2021 Apr 26.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan.

We encountered a case of cutaneous squamous cell carcinoma (SCC) in a 17-year-old female koala at a zoo. A fragile, papillary, elevated mass was found on the third digit of the right hind limb. SCC was identified histopathologically: squamous cell-like polygonal tumor cells showed a nest-like growth pattern with epidermal down growth, central keratinization and necrotic foci, and invaded dermal connective tissues. Metastatic lesions were observed in various organs, including the lung and axillary lymph node: in the lung, multiple metastatic foci similar to the primary lesion, and in the axillary lymph node, individual polygonal tumor cells infiltrated the sinusoids. Immunohistochemistry revealed that the tumor cells were positive for proliferating cell nuclear antigen, which exhibited 32-33% of labeling indices in the tumor cells. To our knowledge, this is the first report of a case of SCC in a digit of a koala.
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http://dx.doi.org/10.1292/jvms.20-0709DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8267191PMC
July 2021

Induction of cellular senescence as a late effect and BDNF-TrkB signaling-mediated ameliorating effect on disruption of hippocampal neurogenesis after developmental exposure to lead acetate in rats.

Toxicology 2021 05 20;456:152782. Epub 2021 Apr 20.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan; Cooperative Division of Veterinary Sciences, Graduate School of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan; Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan. Electronic address:

Lead (Pb) exposure causes cognitive deficits in children. The present study investigated the effect of developmental exposure to Pb acetate (PbAc) on postnatal hippocampal neurogenesis. Pregnant rats were administered drinking water containing 0, 2000, or 4000 ppm PbAc from gestational day 6 until day 21 post-delivery (weaning), and offspring were maintained without PbAc exposure until adulthood on postnatal day (PND) 77. There was a dose-related accumulation of Pb in the offspring brain at weaning, while Pb was mainly excreted in adulthood. In the hippocampus, metallothionein I/II immunoreactive glia were increased through adulthood as a neuroprotective response to accumulated Pb, accompanied by increased astrocyte and microglia numbers in adulthood, suggesting sustained neural damage. Gene expression changes suggested elevated oxidative stress at weaning and suppression of the antioxidant system in adulthood, as well as continued neuroinflammatory responses. At weaning, granule cell apoptosis was increased and numbers of type-3 neural progenitor cells (NPCs) were decreased. By contrast, type-2a and type-2b NPCs were increased, suggesting suppressed differentiation to type-3 NPCs. In adulthood, there were increased numbers of immature granule cells. In the hilus of the dentate gyrus, somatostatin interneurons were increased at weaning, while calbindin-D-29K interneurons were increased throughout adulthood, suggesting a strengthened interneuron regulatory system against the suppressed differentiation at weaning. In the dentate gyrus, Bdnf, Ntrk2, and Chrna7 gene expression were upregulated and numbers of hilar TrkB interneurons increased at weaning. These findings suggest activation of BDNF-TrkB signaling to increase somatostatin interneurons and promote cholinergic signaling, thus increasing later production of immature granule cells. In adulthood, Pcna and Apex1 gene expression were downregulated and Chek1 and cyclin-dependent kinase inhibitor expression were upregulated. Furthermore, there was an increase in γ-H2AX SGZ cells, suggesting induction of cellular senescence of SGZ cells due to Pb genotoxicity.
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http://dx.doi.org/10.1016/j.tox.2021.152782DOI Listing
May 2021

A 28-day repeated oral dose toxicity study of enniatin complex in mice.

J Toxicol Sci 2021 ;46(4):157-165

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology.

Enniatins are so-called "emerging mycotoxins" that commonly occur in milligrams per kilogram levels in grains and their derived products, as well as in fish, dried fruits, nuts, spices, cocoa, and coffee. The present study investigated the 28-day repeated oral dose toxicity of enniatin complex in CD1(ICR) mice. Enniatin B, enniatin B1, and enniatin A1 at a ratio of 4:4:1 were administered to male and female mice at doses of 0 (vehicle controls), 0.8, 4, and 20 mg/kg body weight/day. In life parameters did not change during the study period, with the exception of slight reductions in food consumption in male mice administered 4 and 20 mg/kg and in female mice administered 20 mg/kg. Body and organ weights did not change, and no alterations in hematology, blood biochemistry, or histopathology parameters were observed at the end of the administration period. Thus, we determined that the no-observed-adverse-effect level of enniatin complex was 20 mg/kg/day for both sexes under the present experimental conditions.
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http://dx.doi.org/10.2131/jts.46.157DOI Listing
January 2021

Continuous exposure to amorphous formula of curcumin from the developmental stage facilitates anti-anxiety-like behavior and fear-extinction learning in rats.

Nutr Res 2021 01 29;85:99-118. Epub 2020 Oct 29.

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan; Cooperative Division of Veterinary Sciences, Graduate School of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan; Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan. Electronic address:

An amorphous formula of curcumin (CUR) has shown to enable an improved bioavailability after ingestion. The aim of this study was to investigate the hypothesis that exogenously administered CUR has an advantage in ameliorating post-traumatic stress disorder at low doses. To this end, Long-Evans rats were dietary exposed to CUR at 0.1% or 0.5% from gestational day 6 to postnatal day (PND) 74 or 77. Offspring exposed to 0.1% CUR revealed facilitation of anti-anxiety-like behavior in the open field test and fear-extinction learning tested during PND 62 to 74, increases in hippocampal granule cells expressing immediate-early gene proteins and a decrease in prelimbic cortical neurons expressing phosphorylated extracellular signal-regulated kinase 1/2 after the last trial of the fear-extinction learning test on PND 74. The constitutive gene expression levels of Gria1, Gria2, Grin2d, Slc17a6, and Slc17a7 were altered in the hippocampal dentate gyrus and amygdala on PND 77. These results suggest alterations in synaptic plasticity to strengthen neural circuits in promoting the behavioral effects by 0.1%-CUR. In contrast, 0.5% CUR revealed a lack of any of the changes in behavioral tests that were observed at 0.1%; however, this dose upregulated oxidative stress and neuroinflammation-related genes in the hippocampal dentate gyrus, and increased neural stem cells and proliferation activity of the subgranular zone in the dentate gyrus. These results suggest a possible preventive use of CUR at low doses in mitigating some stress disorders; however, excessively absorbed doses may prevent behavioral changes by inducing neuroinflammation that affects hippocampal neurogenesis involving neural stem cells.
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http://dx.doi.org/10.1016/j.nutres.2020.10.007DOI Listing
January 2021

Continuous exposure to α-glycosyl isoquercitrin from developmental stages to adulthood is necessary for facilitating fear extinction learning in rats.

J Toxicol Pathol 2020 Oct 13;33(4):247-263. Epub 2020 Aug 13.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan.

We previously reported that exposure to α-glycosyl isoquercitrin (AGIQ) from the fetal stage to adulthood facilitated fear extinction learning in rats. The present study investigated the specific AGIQ exposure period sufficient for inducing this behavioral effect. Rats were dietarily exposed to 0.5% AGIQ from the postweaning stage to adulthood (PW-AGIQ), the fetal stage to postweaning stage (DEV-AGIQ), or the fetal stage to adulthood (WP-AGIQ). Fear memory, anxiety-like behavior, and object recognition memory were assessed during adulthood. Fear extinction learning was exclusively facilitated in the WP-AGIQ rats. Synaptic plasticity-related genes showed a similar pattern of constitutive expression changes in the hippocampal dentate gyrus and prelimbic medial prefrontal cortex (mPFC) between the DEV-AGIQ and WP-AGIQ rats. However, WP-AGIQ rats revealed more genes constitutively upregulated in the infralimbic mPFC and amygdala than DEV-AGIQ rats, as well as FOS-immunoreactive neurons constitutively increased in the infralimbic cortex. Ninety minutes after the last fear extinction trial, many synaptic plasticity-related genes (encoding Ephs/Ephrins, glutamate receptors/transporters, and immediate-early gene proteins and their regulator, extracellular signal-regulated kinase 2 [ERK2]) were upregulated in the dentate gyrus and amygdala in WP-AGIQ rats. Additionally, WP-AGIQ rats exhibited increased phosphorylated ERK1/2 neurons in both the prelimbic and infralimbic cortices. These results suggest that AGIQ exposure from the fetal stage to adulthood is necessary for facilitating fear extinction learning. Furthermore, constitutive and learning-dependent upregulation of synaptic plasticity-related genes/molecules may be differentially involved in brain regions that regulate fear memory. Thus, new learning-related neural circuits for facilitating fear extinction can be established in the mPFC.
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http://dx.doi.org/10.1293/tox.2020-0025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7677619PMC
October 2020

Identification of gene targets of developmental neurotoxicity focusing on DNA hypermethylation involved in irreversible disruption of hippocampal neurogenesis in rats.

J Appl Toxicol 2021 Jul 4;41(7):1021-1037. Epub 2020 Nov 4.

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology, Tokyo, Japan.

We have previously found that maternal exposure to 6-propyl-2-thiouracil (PTU), valproic acid (VPA), or glycidol (GLY) has a sustained or late effect on hippocampal neurogenesis at the adult stage in rat offspring. Herein, we searched for genes with hypermethylated promoter region and downregulated transcript level to reveal irreversible markers of developmental neurotoxicity. The hippocampal dentate gyrus of male rat offspring exposed maternally to PTU, VPA, or GLY was subjected to Methyl-Seq and RNA-Seq analyses on postnatal day (PND) 21. Among the genes identified, 170 were selected for further validation analysis of gene expression on PND 21 and PND 77 by real-time reverse transcription-PCR. PTU and GLY downregulated many genes on PND 21, reflecting diverse effects on neurogenesis. Furthermore, genes showing sustained downregulation were found after PTU or VPA exposure, reflecting a sustained or late effect on neurogenesis by these compounds. In contrast, such genes were not observed with GLY, probably because of the reversible nature of the effects. Among the genes showing sustained downregulation, Creb, Arc, and Hes5 were concurrently downregulated by PTU, suggesting an association with neuronal mismigration, suppressed synaptic plasticity, and reduction in neural stem and progenitor cells. Epha7 and Pvalb were also concurrently downregulated by PTU, suggesting an association with the reduction in late-stage progenitor cells. VPA induced sustained downregulation of Vgf and Dpysl4, which may be related to the aberrations in synaptic plasticity. The genes showing sustained downregulation may be irreversible markers of developmental neurotoxicity.
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http://dx.doi.org/10.1002/jat.4089DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8247304PMC
July 2021

Downregulation of low-density lipoprotein receptor class A domain-containing protein 4 (Ldlrad4) in the liver of rats treated with nongenotoxic hepatocarcinogen to induce transforming growth factor β signaling promoting cell proliferation and suppressing apoptosis in early hepatocarcinogenesis.

J Appl Toxicol 2020 11 28;40(11):1467-1479. Epub 2020 Jun 28.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, Tokyo, Japan.

We previously found downregulation of low-density lipoprotein receptor class A domain-containing protein 4 (LDLRAD4), a negative regulator of transforming growth factor (TGF)-β signaling, in glutathione S-transferase placental form (GST-P) expressing ( ) pre-neoplastic lesions produced by treatment with nongenotoxic hepatocarcinogens for up to 90 days in rats. Here, we investigated the relationship between LDLRAD4 downregulation and TGFβ signaling in nongenotoxic hepatocarcinogenesis. The transcripts of Tgfb and Hb-egf increased after ≥28 days of treatment. After 84 or 90 days, Snai1 increased transcripts and the subpopulation of GST-P foci downregulating LDLRAD4 co-expressed TGFβ1, phosphorylated EGFR, or phosphorylated AKT2, and downregulated PTEN, showing higher incidences than those in GST-P foci expressing LDLRAD4. The subpopulation of GST-P foci downregulating LDLRAD4 also co-expressed caveolin-1 or TACE/ADAM17, suggesting that disruptive activation of TGFβ signaling through a loss of LDLRAD4 enhances EGFR and PTEN/AKT-dependent pathways via caveolin-1-dependent activation of TACE/ADAM17 during nongenotoxic hepatocarcinogenesis. The numbers of c-MYC cells and PCNA cells were higher in LDLRAD4-downregulated GST-P foci than in LDLRAD4-expressing GST-P foci, suggesting a preferential proliferation of pre-neoplastic cells by LDLRAD4 downregulation. Nongenotoxic hepatocarcinogens markedly downregulated Nox4 after 28 days and later decreased cleaved caspase 3 cells in LDLRAD4-downregulated GST-P foci, suggesting an attenuation of apoptosis by LDLRAD4 downregulation through activation of the EGFR pathway. At the late hepatocarcinogenesis stage in a two-stage model, LDLRAD4 downregulation was higher in adenoma and carcinoma than in pre-neoplastic cell foci, suggesting a role of LDLRAD4 downregulation in tumor development. Our results suggest that nongenotoxic hepatocarcinogens cause disruptive activation of TGFβ signaling through downregulating LDLRAD4 toward carcinogenesis in the rat liver.
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http://dx.doi.org/10.1002/jat.3998DOI Listing
November 2020

Establishment of 2.5D organoid culture model using 3D bladder cancer organoid culture.

Sci Rep 2020 06 10;10(1):9393. Epub 2020 Jun 10.

Laboratory of Veterinary Pharmacology, Department of Veterinary Medicine, Faculty of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 183-8509, Japan.

Three-dimensional (3D) organoid culture holds great promises in cancer precision medicine. However, Matrigel and stem cell-stimulating supplements are necessary for culturing 3D organoid cells. It costs a lot of money and consumes more time and effort compared with 2D cultured cells. Therefore, the establishment of cheaper and Matrigel-free organoid culture that can maintain the characteristics of a part of 3D organoids is demanded. In the previous study, we established a dog bladder cancer (BC) 3D organoid culture system by using their urine samples. Here, we successfully isolated cells named "2.5D organoid" from multiple strains of dog BC 3D organoids using 2.5 organoid media. The cell proliferation speed of 2.5D organoids was faster than parental 3D organoid cells. The expression pattern of stem cell markers was close to 3D organoids. Injection of 2.5D organoid cells into immunodeficient mice formed tumors and showed the histopathological characteristics of urothelial carcinoma similar to the injection of dog BC 3D organoids. The 2.5D organoids had a similar sensitivity profile for anti-cancer drug treatment to their parental 3D organoids. These data suggest that our established 2.5D organoid culture method might become a reasonable and useful tool instead of 3D organoids in dog BC research and therapy.
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http://dx.doi.org/10.1038/s41598-020-66229-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7287130PMC
June 2020

Effect of dapagliflozin alone and in combination with insulin in a rat model of type 1 diabetes.

J Vet Med Sci 2020 Apr 11;82(4):467-474. Epub 2020 Mar 11.

Laboratory of Veterinary Pharmacology, Department of Veterinary Medicine, Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183-8509, Japan.

Dapagliflozin is a selective sodium-glucose cotransporter 2 (SGLT2) inhibitor; it reduces glucose reabsorption via the kidney and increases the glucose excretion in urine. This inhibitor functions through a unique insulin-independent mechanism, and is therefore a potential new approach for the treatment of hyperglycemia in patients with diabetes. In this study, we evaluated the effectiveness of the SGLT2 inhibitor, dapagliflozin, by using a rat model of type 1 diabetes. Type 1 diabetes was induced by a single intraperitoneal injection of 60 mg/kg streptozotocin (STZ). The STZ-induced rats showed marked hyperglycemia and other metabolic abnormalities. We clarified the hypoglycemic effect of the combination treatment of dapagliflozin with a low dose of insulin compared with dapagliflozin alone and insulin alone in 3-week and 8-week studies. Our results showed that dapagliflozin in combination with a low dose of insulin significantly lowered hyperglycemia, hypercholesterolemia, and hypertriglyceridemia. Furthermore, the antioxidant status and body weight were improved. In contrast, treatment with dapagliflozin alone did not improve the blood glucose levels, lipid profile, antioxidant status, or body weight. These findings suggested that in type 1 diabetes, dapagliflozin was effective in combination with a low dose of insulin; however, the administration of dapagliflozin alone did not achieve a significant effect.
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http://dx.doi.org/10.1292/jvms.19-0450DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7192721PMC
April 2020

Efficacy of primary liver organoid culture from different stages of non-alcoholic steatohepatitis (NASH) mouse model.

Biomaterials 2020 04 27;237:119823. Epub 2020 Jan 27.

Laboratory of Veterinary Pharmacology, Department of Veterinary Medicine, Faculty of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu, Tokyo, 183-8509, Japan.

Non-alcoholic steatohepatitis (NASH) is associated with liver fibrosis and cirrhosis, which eventually leads to hepatocellular carcinoma. Although several animal models were developed to understand the mechanisms of NASH pathogenesis and progression, it remains obscure. A 3D organoid culture system can recapitulate organ structures and maintain gene expression profiles of original tissues. We therefore tried to generate liver organoids from different degrees [defined as mild (NASH A), moderate (NASH B) and severe (NASH C)] of methionine- and choline-deficient diet-induced NASH model mice and analyzed the difference of their architecture, cell components, organoid-forming efficacy, and gene expression profiles. Organoids from each stage of NASH model mice were successfully generated. Interestingly, epithelial-mesenchymal transition was observed in NASH C organoids. Expression of Collagen I and an activated hepatic stellite cell marker, α-sma was upregulated in the liver organoids from NASH B and C mice. The analysis of RNA sequencing revealed that several novel genes were upregulated in all NASH liver organoids. These results suggest that our generated liver organoids from different stages of NASH diseased mice might become a useful tool for in vitro studies of the molecular mechanism of NASH development and also for identifying novel biomarkers for early diagnosis of NASH disease.
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http://dx.doi.org/10.1016/j.biomaterials.2020.119823DOI Listing
April 2020

Developmental exposure to diacetoxyscirpenol reversibly disrupts hippocampal neurogenesis by inducing oxidative cellular injury and suppressed differentiation of granule cell lineages in mice.

Food Chem Toxicol 2020 Feb 10;136:111046. Epub 2019 Dec 10.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan; Cooperative Division of Veterinary Sciences, Graduate School of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan; Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan. Electronic address:

To investigate the developmental exposure effect of diacetoxyscirpenol (DAS) on postnatal hippocampal neurogenesis, pregnant ICR mice were provided a diet containing DAS at 0, 0.6, 2.0, or 6.0 ppm from gestational day 6 to day 21 on weaning after delivery. Offspring were maintained through postnatal day (PND) 77 without DAS exposure. On PND 21, neural stem cells (NSCs) and all subpopulations of proliferating progenitor cells were suggested to decrease in number in the subgranular zone (SGZ) at ≥ 2.0 ppm. At 6.0 ppm, increases of SGZ cells showing TUNEL, metallothionein-I/II, γ-H2AX or malondialdehyde, and transcript downregulation of Ogg1, Parp1 and Kit without changing the level of double-stranded DNA break-related genes were observed in the dentate gyrus. This suggested induction of oxidative DNA damage of NSCs and early-stage progenitor cells, which led to their apoptosis. Cdkn2a, Rb1 and Trp53 downregulated transcripts, which suggested an increased vulnerability to DNA damage. Hilar PVALB GABAergic interneurons decreased and Grin2a and Chrna7 were downregulated, which suggested suppression of type-2-progenitor cell differentiation. On PND 77, hilar RELN interneurons increased at ≥ 2.0 ppm; at 6.0 ppm, RELN-related Itsn1 transcripts were upregulated and ARC granule cells decreased. Increased RELN signals may ameliorate the response to the decreases of NSCs and ARC-mediated synaptic plasticity. These results suggest that DAS reversibly disrupts hippocampal neurogenesis by inducing oxidative cellular injury and suppressed differentiation of granule cell lineages. The no-observed-adverse-effect level of DAS for offspring neurogenesis was determined to be 0.6 ppm (0.09-0.29 mg/kg body weight/day).
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http://dx.doi.org/10.1016/j.fct.2019.111046DOI Listing
February 2020

Differential responses on energy metabolic pathway reprogramming between genotoxic and non-genotoxic hepatocarcinogens in rat liver cells.

J Toxicol Pathol 2019 Oct 28;32(4):261-274. Epub 2019 Jul 28.

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan.

To clarify difference in the responses on the reprogramming of metabolism toward carcinogenesis between genotoxic and non-genotoxic hepatocarcinogens in the liver, rats were repeatedly administered genotoxic hepatocarcinogens (-nitrosodiethylamine, aflatoxin B, -nitrosopyrrolidine, or carbadox) or non-genotoxic hepatocarcinogens (carbon tetrachloride, thioacetamide, or methapyrilene hydrochloride) for 28, 84, or 90 days. Non-genotoxic hepatocarcinogens revealed transcript expression changes suggestive of suppressed mitochondrial oxidative phosphorylation (OXPHOS) after 28 days and increased glutathione -transferase placental form-positive (GST-P) foci downregulating adenosine triphosphate (ATP) synthase subunit beta, mitochondrial precursor (ATPB), compared with genotoxic hepatocarcinogens after 84 or 90 days, suggesting that non-genotoxic hepatocarcinogens are prone to suppress OXPHOS from the early stage of treatment, which is in contrast to genotoxic hepatocarcinogens. Both genotoxic and non-genotoxic hepatocarcinogens upregulated glycolytic enzyme genes and increased cellular membrane solute carrier family 2, facilitated glucose transporter member 1 (GLUT1) expression in GST-P foci for up to 90 days, suggesting induction of a metabolic shift from OXPHOS to glycolysis at early hepatocarcinogenesis by hepatocarcinogens unrelated to genotoxic potential. Non-genotoxic hepatocarcinogens increased c-MYC cells after 28 days and downregulated after 84 or 90 days, suggesting a commitment to enhanced metabolic shift and cell proliferation. Genotoxic hepatocarcinogens also enhanced c-MYC activation-related metabolic shift until 84 or 90 days. In addition, both genotoxic and non-genotoxic hepatocarcinogens upregulated glutaminolysis-related or , or both, after 28 days and induced liver cell foci immunoreactive for neutral amino acid transporter B(0) (SLC1A5) in the subpopulation of GST-P foci after 84 or 90 days, suggesting glutaminolysis-mediated facilitation of cell proliferation toward hepatocarcinogenesis. These results suggest differential responses between genotoxic and non-genotoxic hepatocarcinogens on reprogramming of energy metabolic pathways toward carcinogenesis in liver cells from the early stage of hepatocarcinogen treatment.
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http://dx.doi.org/10.1293/tox.2019-0048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6831489PMC
October 2019

Extraskeletal chondrosarcoma in the abdominal cavity of a cow.

J Vet Med Sci 2019 Dec 23;81(12):1749-1752. Epub 2019 Oct 23.

Shibaura Meat Sanitary Inspection Center, Tokyo Metropolitan Government, 2-7-19 Konan, Minato-ku, Tokyo 163-8001, Japan.

A 25-month-old female crossbred cow presented with astasia, emaciation, and stunted growth. Macroscopic examination revealed a large mass in the abdominal cavity, approximately 100 × 30 × 30 cm. Microscopic examination revealed that the mass consisted of multilobular mature and immature cartilaginous matrices with chondrocytic cells, surrounded by spindle to pleomorphic mesenchymal tumor cells. The cartilaginous matrices consisted of hyaline and elastic cartilages, as confirmed with Azan stain, and Victoria Blue and Van Gieson stain. Immunohistochemistry revealed that the chondrocytic and mesenchymal cells both expressed S-100. The tumor was diagnosed as an extraskeletal chondrosarcoma in the abdominal cavity of this cow.
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http://dx.doi.org/10.1292/jvms.19-0203DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943319PMC
December 2019

Lack of preventive effect of maternal exposure to α-glycosyl isoquercitrin and α-lipoic acid on developmental hypothyroidism-induced aberrations of hippocampal neurogenesis in rat offspring.

J Toxicol Pathol 2019 Jul 31;32(3):165-180. Epub 2019 May 31.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan.

Hypothyroidism during the developmental stage induces disruption of hippocampal neurogenesis in later life, as well as inducing oxidative stress in the brain. The present study investigated the preventive effect of co-exposure to an antioxidant on disruptive neurogenesis induced by developmental exposure to anti-thyroid agent in rats. For this purpose, we used two antioxidants, α-glycosyl isoquercitrin (AGIQ) and α-lipoic acid (ALA). Mated female Sprague Dawley rats were either untreated (control) or treated with 12 ppm 6-propyl-2-thiouracil (PTU), an anti-thyroid agent, in drinking water from gestational day 6 to postnatal day (PND) 21, the latter group being subjected to feeding basal diet alone or diet containing AGIQ at 5,000 ppm or ALA at 2,000 ppm during PTU exposure. On PND 21, PTU-exposed offspring showed reductions in a broad range of granule cell lineage subpopulations and a change in the number of GABAergic interneuron subpopulations. Co-exposure of AGIQ or ALA with PTU altered the transcript levels of many genes across multiple functions, suggestive of enhancement of synaptic plasticity and neurogenesis. Nevertheless, immunohistochemical results did not support these changes. PTU exposure and co-exposure of AGIQ or ALA with PTU did not alter the hippocampal lipid peroxidation level. The obtained results suggest a possibility that thyroid hormone depletion itself primarily disrupts neurogenesis and that oxidative stress may not be involved in the disruption during development. Transcript expression changes of many genes caused by antioxidants may be the result of neuroprotective actions of antioxidants rather than their antioxidant activity. However, no preventive effect on neurogenesis suggested impairment of protein synthesis via an effect on mRNA translation due to hypothyroidism.
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http://dx.doi.org/10.1293/tox.2019-0018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6682556PMC
July 2019

Immunohistochemical expression of autophagosome markers LC3 and p62 in preneoplastic liver foci in high fat diet-fed rats.

J Toxicol Sci 2019 ;44(8):565-574

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology.

Nonalcoholic fatty liver disease (NAFLD) is characterized by excessive deposition of droplets in hepatocytes. Patients with NAFLD can be at risk for nonalcoholic steatohepatitis, which can lead to hepatocellular carcinoma. Autophagy is a cellular pathway that is crucial for survival and homeostasis, and which protects against pathophysiological changes like obesity and cancer. We determined the expression of autophagy markers in preneoplastic hepatic lesions and the effects of an autophagy repressor chloroquine (CQ) or inducer amiodarone (AM) in a steatosis-related hepatocarcinogenesis model. Male F344 rats were fed a control diet or high fat diet (HFD), and subjected to initiation and promotion steps with N-nitrosodiethylamine injection at week 0 and a partial hepatectomy at week 3. Several HFD-fed rats were administered 0.1% CQ and 0.5% AM in their drinking water during week 2 and 8. CQ and AM did not improve HFD-induced obesity. AM, but not CQ, significantly decreased the number of glutathione S-transferase placental form-positive preneoplastic liver foci in the liver. Autophagosome markers LC3 and the LC3-binding protein p62 were heterogeneously expressed in the preneoplastic foci. CQ might inhibit autophagy by significantly increased p62/LC3 ratio, while AM might have a potential of inducing autophagy by showing an increased gene expression of the autophagy regulator, Atg5. These results suggest that preneoplastic lesions express autophagosome markers and that AM might decrease steatosis-related early hepatocarcinogenesis by potentially inducing autophagy in HFD-fed rats, while inhibition of autophagy by CQ did not alter the hepatocarcinogenesis. However, an immunohistochemical trial revealed a technical limitation in detecting autophagosome markers because there were variations in each preneoplastic lesion.
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http://dx.doi.org/10.2131/jts.44.565DOI Listing
December 2019

Spontaneous Age-Related Histopathological Changes in Microminipigs.

Toxicol Pathol 2019 10 23;47(7):817-832. Epub 2019 Jul 23.

Swine and Poultry Department, Shizuoka Prefectural Research Institute of Animal Industry, Swine and Poultry Research Center, Shizuoka, Japan.

Microminipigs have become an attractive animal model for toxicology and pharmacology studies and for human disease models, owing to their manageable size. Although there are numerous reports of spontaneous age-related lesions in mice, rats, dogs, and monkeys, those in minipigs are scarce. In the present study, spontaneous age-related histopathological changes were investigated using 37 microminipigs (20 males and 17 females) that were 6 months to 10 years of age. Abnormal deposits of materials were evident in several animals from 6 years of age, and these deposits included amyloid in the renal medulla, thyroid gland, and adrenal gland, hyaline droplets in glomeruli, and fibrillar inclusions in neurons. Arterial sclerosing changes (intimal thickening, intimal proliferation, and medial mineralization) and proliferative lesions (hyperplasia of hepatocytes, follicular cells, Leydig cells, and uterine endometrial glands) were present at 4 years of age and beyond. Renal adenoma, uterine leiomyoma, and Leydig cell tumor were observed in several microminipigs. Moreover, glomerulosclerosis, renal interstitial fibrosis, thymic involution, and adrenocortical cell vacuolation were common in aging microminipigs. Since knowledge of age-related changes is helpful for pathologists, the basic information obtained in this study will be a useful reference for all future toxicity evaluations in microminipigs.
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http://dx.doi.org/10.1177/0192623319861350DOI Listing
October 2019

Effects of radiation based on whole-body irradiation in HTLV-1-infected mice.

J Radiat Res 2019 10;60(5):705-708

Faculty of Bioscience, Nagahama Institute of Bioscience and Technology, 1266 Tamura, Nagahama, Shiga, Japan.

Adult T-cell leukemia is one of the life-threatening diseases that occur in individuals infected with human T-cell leukemia virus type 1 (HTLV-1). Clinical trials of hematopoietic stem cell transplantation therapy are being performed in addition to chemotherapy; however, neither is satisfactory. As a pretreatment for transplantation, anticancer drugs or whole-body irradiation is used to decrease the number of HTLV-1-infected cells, but there are numerous side effects. Therefore, in the present study, using a mouse model of HTLV-1 infection, the long-term survival and number of infected cells in the reservoir organ were investigated in order to determine the effect of γ-irradiation on HTLV-1-infected mice in vivo. There was no improvement in the survival period following γ-irradiation in the γ-irradiated group after HTLV-1 infection when compared with the HTLV-1-infected group. It was also found that the incidence of splenomegaly was ≥80% in the HTLV-1-infected and γ-irradiated group, which was significantly higher than that in the HTLV-1-infected mice. The tissue morphology in the spleen became non-uniform because of γ-rays. Importantly, the number of infected cells in the spleen was increased 4.1-fold in the HTLV-1-infected and γ-irradiated mice compared with that in the HTLV-1-infected mice. Careful consideration might be necessary when using whole-body irradiation in patients with HTLV-1 infection.
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http://dx.doi.org/10.1093/jrr/rrz050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6805971PMC
October 2019

Clinical and pathological characteristics of acute myelogenous leukemia in a female koala with diabetes mellitus.

J Vet Med Sci 2019 Aug 2;81(8):1229-1233. Epub 2019 Jul 2.

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan.

A female koala presented with hyperglycemia related to diabetes mellitus diagnosed at 9 years and treated with insulin. She presented with nasal hemorrhage, anemia, leukocytosis, and tachypnea at 10 years. A blood smear examination revealed scattered, atypical large myeloid cells and a clinical diagnosis of myelogenous leukemia was made. White blood cell count reached a maximum of 295 × 10/µl, with evidence of severe regenerative anemia and thrombocytopenia. Grossly, systemic lymph node enlargement, fragile liver with hemorrhage, and bloody ascites were observed. Histopathologically, atypical myeloid cells, including myelocytic and metamyelocytic cells, were scattered in the vasculature and surrounding tissues throughout the organs. The patient was infected with a koala retrovirus, which might have caused the myelogenous leukemia.
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http://dx.doi.org/10.1292/jvms.19-0006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6715919PMC
August 2019

Establishment of a novel experimental model for muscle-invasive bladder cancer using a dog bladder cancer organoid culture.

Cancer Sci 2019 Sep 23;110(9):2806-2821. Epub 2019 Jul 23.

Laboratory of Veterinary Pharmacology, Department of Veterinary Medicine, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Fuchu, Japan.

In human and dogs, bladder cancer (BC) is the most common neoplasm affecting the urinary tract. Dog BC resembles human muscle-invasive BC in histopathological characteristics and gene expression profiles, and could be an important research model for this disease. Cancer patient-derived organoid culture can recapitulate organ structures and maintains the gene expression profiles of original tumor tissues. In a previous study, we generated dog prostate cancer organoids using urine samples, however dog BC organoids had never been produced. Therefore we aimed to generate dog BC organoids using urine samples and check their histopathological characteristics, drug sensitivity, and gene expression profiles. Organoids from individual BC dogs were successfully generated, expressed urothelial cell markers (CK7, CK20, and UPK3A) and exhibited tumorigenesis in vivo. In a cell viability assay, the response to combined treatment with a range of anticancer drugs (cisplatin, vinblastine, gemcitabine or piroxicam) was markedly different in each BC organoid. In RNA-sequencing analysis, expression levels of basal cell markers (CK5 and DSG3) and several novel genes (MMP28, CTSE, CNN3, TFPI2, COL17A1, and AGPAT4) were upregulated in BC organoids compared with normal bladder tissues or two-dimensional (2D) BC cell lines. These established dog BC organoids might be a useful tool, not only to determine suitable chemotherapy for BC diseased dogs but also to identify novel biomarkers in human muscle-invasive BC. In the present study, for the 1st time, dog BC organoids were generated and several specifically upregulated organoid genes were identified. Our data suggest that dog BC organoids might become a new tool to provide fresh insights into both dog BC therapy and diagnostic biomarkers.
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http://dx.doi.org/10.1111/cas.14118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6726682PMC
September 2019

Twenty-eight-day repeated oral doses of sodium valproic acid increases neural stem cells and suppresses differentiation of granule cell lineages in adult hippocampal neurogenesis of postpubertal rats.

Toxicol Lett 2019 Sep 11;312:195-203. Epub 2019 May 11.

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan; Institute of Global Innovation Research, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan. Electronic address:

Developmental exposure to valproic acid (VPA), a model compound for experimental autism, has shown to primarily target GABAergic interneuron subpopulations in hippocampal neurogenesis of rat offspring. The VPA-exposed animals had revealed late effects on granule cell lineages, involving progenitor cell proliferation and synaptic plasticity. To investigate the possibility whether hippocampal neurogenesis in postpubertal rats in a protocol of 28-day repeated exposure is affected in relation with the property of a developmental neurotoxicant by developmental exposure, VPA was orally administered to 5-week-old male rats at 0, 200, 800 and 900 mg/kg body weight/day for 28 days. At 900 mg/kg, GFAP cells increased in number, but DCX cells decreased in number in the granule cell lineages. Moreover, CHRNB2 cells and NeuN postmitotic neurons decreased in number in the hilus of the dentate gyrus. Transcript level examined at 900 mg/kg in the dentate gyrus was increased with Kit, but decreased with Dpsyl3, Btg2, Pvalb and Chrnb2. These results suggest that VPA increased type-1 stem cells in relation to the activation of SCF-KIT signaling and suppression of BTG2-mediated antiproliferative effect on stem cells. VPA also decreased type-3 progenitor cells and immature granule cells probably in relation with PVALB interneuron hypofunction and reduced CHRNB2 interneuron subpopulation in the hilus, as well as with suppression of BTG2-mediated terminal differentiation of progenitor cells. Thus, the disruption pattern of VPA by postpubertal exposure was different from developmental exposure. However, disruption itself can be detected, suggesting availability of hippocampal neurogenesis in detecting developmental neurotoxicants in a 28-day toxicity study.
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http://dx.doi.org/10.1016/j.toxlet.2019.05.013DOI Listing
September 2019

Ameliorating effect of postweaning exposure to antioxidant on disruption of hippocampal neurogenesis induced by developmental hypothyroidism in rats.

J Toxicol Sci 2019 ;44(5):357-372

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology.

Developmental hypothyroidism as a model of autism spectrum disorders disrupts hippocampal neurogenesis through the adult stage. The present study investigated the ameliorating effect of postweaning exposure to antioxidant on the hypothyroidism-induced disruptive neurogenesis. Mated female Sprague-Dawley rats were treated with 0 or 10 ppm 6-propyl-2-thiouracil (PTU) as an anti-thyroid agent in drinking water from gestational day 6 to postnatal day (PND) 21 on weaning. PTU-exposed male offspring were fed either basal diet, diet containing α-glycosyl isoquercitrin (AGIQ) at 5,000 ppm or α-lipoic acid (ALA) at 1,000 ppm as an antioxidant from PND 21 to PND 77. PTU-exposure decreased DCX and NeuN granule cell lineage subpopulations, synaptic plasticity-related FOS granule cells, and hilar PVALB and GAD67 GABAergic interneurons, increased hilar SST and CALB2 interneurons, and upregulated Gria3, Otx2, and antioxidant enzyme genes in the dentate gyrus on PND 77. These results suggest disruption of neurogenesis remained in relation with increase of oxidative stress and compensatory responses to the disruption at the adult stage. AGIQ recovered expression of some antioxidant enzyme genes and was effective for restoration of NeuN postmitotic granule cells and PVALB and SST interneurons. In contrast, ALA was effective for restoration of all interneuron subpopulations, as well as postmitotic granule cells, and upregulated Grin2a that may play a role for the restoration. Both antioxidants recovered expression of Otx2 and AGIQ-alone recovered Gria3, suggesting a reversal of disruptive neurogenesis by compensatory responses. Thus, postweaning antioxidant exposure may be effective for ameliorating developmental hypothyroidism-induced disruptive neurogenesis by restoring the function of regulatory system.
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http://dx.doi.org/10.2131/jts.44.357DOI Listing
September 2019

Histological Changes of the Testicular Interstitium during Postnatal Development in Microminipigs.

Toxicol Pathol 2019 06 10;47(4):469-482. Epub 2019 Feb 10.

1 Swine and Poultry Department, Swine and Poultry Research Center, Shizuoka Prefectural Research Institute of Animal Industry, Kikugawa, Shizuoka, Japan.

Microminipigs have become an attractive animal model for the toxicology- and pharmacology-related studies because of their manageable size. In this study, the development of the testicular interstitium and steroidogenesis in microminipigs, from 0 to 12 months of age, were investigated. Testicular interstitium was mostly composed of two types of Leydig cells (large and small Leydig cells) and a few macrophages and mast cells. Large Leydig cells were observed in the peritubular area throughout all the ages. Small Leydig cells were present in the interlobular and subcapsular areas at an early age and then gradually converted into large Leydig cells. Testicular composition of the Leydig cells began to increase after 3 months of age, when spermatogenesis was completed, and reached approximately 35% at 12 months. Steroidogenic enzymes in Leydig cells were detected by immunohistochemistry from 0 month of age. Serum testosterone levels increased substantially from 1.5 to 4.5 months of age, which coincided well with the age of sexual development previously reported in microminipigs. Because the interstitial space of the testis has dramatic variations between species, the basic information obtained in the present study will be a useful reference for all the future toxicity evaluations in microminipigs.
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http://dx.doi.org/10.1177/0192623319827477DOI Listing
June 2019

Aberrant epigenetic gene regulation in hippocampal neurogenesis of mouse offspring following maternal exposure to 3,3'-iminodipropionitrile.

J Toxicol Sci 2019 ;44(2):93-105

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology.

Maternal exposure to 3,3'-iminodipropionitrile (IDPN) affects hippocampal neurogenesis in mouse offspring, with biphasic disruption, which facilitates neurogenesis during exposure and reduces the broad range of the granule cell lineage population at the adult stage. The present study investigated the epigenetically hypermethylated and downregulated genes related to the IDPN-induced disrupted neurogenesis. Mated female mice were treated with IDPN at 0 or 1200 ppm in drinking water from gestational day 6 to postnatal day (PND) 21 on weaning. The hippocampal dentate gyrus of male offspring on PND 21 was subjected to methyl-capture sequencing and real-time reverse transcription-PCR analyses, followed by validation analyses on DNA methylation. Three genes, Edc4, Kiss1 and Mrpl38, were identified as those showing promoter-region hypermethylation and transcript downregulation, with Mrpl38 sustaining the changes through PND 77. Immunohistochemically, MRPL38, a mitochondrial ribosomal protein, revealed an irreversible decrease in the number of immunoreactive interneurons in the dentate gyrus hilar region, suggesting a causal relationship with the long-lasting effect on neurogenesis by the impaired migration due to mitochondrial dysfunction of interneurons, which regulate the differentiation and survival of granule cell lineages. Downregulation of Edc4 may also be responsible for decreased neurogenesis on PND 77 owing to a mechanism involving interleukin-6 downregulation via processing body dysfunction. Downregulation of Kiss1 may be responsible for the facilitation of neurogenesis during IDPN-exposure due to decreased glutamatergic neurotransmission and also for suppressed neurogenesis on PND 77 due to decreased expression of immediate-early genes, which play a crucial role in the maintenance of cell differentiation or plasticity.
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http://dx.doi.org/10.2131/jts.44.93DOI Listing
February 2019

Expression Characteristics of Genes Hypermethylated and Downregulated in Rat Liver Specific to Nongenotoxic Hepatocarcinogens.

Toxicol Sci 2019 05;169(1):122-136

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, Fuchu-shi, Tokyo, Japan.

This study examined hypermethylated and downregulated genes specific to carbon tetrachloride (CCl4) by Methyl-Seq analysis combined with expression microarray analysis in the liver of rats treated with CCl4 or N-nitrosodiethylamine (DEN) for 28 days, by excluding those with DEN. Among 52 genes, Ldlrad4, Proc, Cdh17, and Nfia were confirmed to show promoter-region hypermethylation by methylation-specific quantitative PCR analysis on day 28. The transcript levels of these 4 genes decreased by real-time reverse transcription-PCR analysis in the livers of rats treated with nongenotoxic hepatocarcinogens for up to 90 days compared with untreated controls and genotoxic hepatocarcinogens. Immunohistochemically, LDLRAD4 and PROC showed decreased immunoreactivity, forming negative foci, in glutathione S-transferase placental form (GST-P)+ foci, and incidences of LDLRAD4- and PROC- foci in GST-P+ foci induced by treatment with nongenotoxic hepatocarcinogens for 84 or 90 days were increased compared with those with genotoxic hepatocarcinogens. In contrast, CDH17 and NFIA responded to hepatocarcinogens without any relation to the genotoxic potential of carcinogens. All 4 genes did not respond to renal carcinogens after treatment for 28 days. Considering that Ldlrad4 is a negative regulator of transforming growth factor-β signaling, Proc participating in p21WAF1/CIP1 upregulation by activation, Cdh17 inducing cell cycle arrest by gene knockdown, and Nfia playing a role in a tumor-suppressor, all these genes may be potential in vivo epigenetic markers of nongenotoxic hepatocarcinogens from the early stages of treatment in terms of gene expression changes. LDLRAD4 and PROC may have a role in the development of preneoplastic lesions produced by nongenotoxic hepatocarcinogens.
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http://dx.doi.org/10.1093/toxsci/kfz027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6484883PMC
May 2019

Developmental Exposure of Mice to T-2 Toxin Increases Astrocytes and Hippocampal Neural Stem Cells Expressing Metallothionein.

Neurotox Res 2019 Apr 29;35(3):668-683. Epub 2018 Nov 29.

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan.

We previously reported that developmental exposure to T-2 toxin caused transient disruption of the hippocampal neurogenesis targeting neural stem cells (NSCs) and early-stage progenitor cells involving oxidative stress on weaning in mouse offspring. The present study examined metallothionein (MT) expression changes and their cellular identity in brain regions of these animals. T-2 toxin at 0, 1, 3, and 9 mg/kg was given in the diet of maternal mice from gestational day 6 to postnatal day (PND) 21 on weaning. Offspring were maintained through PND 77 without T-2 toxin exposure. Male offspring were analyzed. Immunohistochemically, MT-I/II cells increased in the subgranular zone (SGZ) of the dentate gyrus and cerebral cortex at ≥ 3 mg/kg and in the hilus of the dentate gyrus, corpus callosum, and cerebellum at 9 mg/kg on PND 21, suggestive of operation of cytoprotective function against oxidative stress throughout the brain. Double immunohistochemistry analysis revealed MT-I/II SGZ cells to be NSCs and MT-I/II cells in other brain regions to be astrocytes as toxicity targets of T-2 toxin. Phosphorylated STAT3 cell numbers increased only in the cerebellum in parallel with the increase of GFAP astrocytes at 9 mg/kg, suggesting a STAT3-mediated transcriptional GFAP upregulation in cerebellar astrocytes. In the dentate gyrus, Il1a, Il1r1, and Mt2 increased transcripts at 9 mg/kg, suggesting activation of the IL-1 signaling cascade, possibly causing MT-II upregulation. The increase of MT-I/II cells in all brain regions disappeared or was suppressed below the control level on PND 77, suggesting a recovery from the T-2 toxin-induced oxidative stress.
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http://dx.doi.org/10.1007/s12640-018-9981-4DOI Listing
April 2019

Differential impacts of mineralocorticoid receptor antagonist potassium canrenoate on liver and renal changes in high fat diet-mediated early hepatocarcinogenesis model rats.

J Toxicol Sci 2018 ;43(10):611-621

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology.

Mineralocorticoid receptor (MR)/NADPH oxidase (NOX) signaling is involved in the development of obesity, insulin resistance, and renal diseases; however, the role of this signaling on steatotic preneoplastic liver lesions is not fully elucidated. We determined the effects of the MR antagonist potassium canrenoate (PC) on MR/NOX signaling in hepatic steatosis and preneoplastic glutathione S-transferase placental form (GST-P)-positive liver foci. Rats were subjected to a two-stage hepatocarcinogenesis model and fed with basal diet or high fat diet (HFD) that was co-administered with PC alone or in combination with the antioxidant alpha-glycosyl isoquercitrin (AGIQ). PC reduced obesity and renal changes (basophilic tubules that expressed MR and p22phox) but did not affect blood glucose tolerance and non-alcoholic fatty liver disease activity score (NAS) in HFD-fed rats. However, the drug increased the area of GST-P-positive liver foci that expressed MR and p22phox as well as increased expression of NOX genes (p22phox, Poldip2, and NOX4). PC in combination with AGIQ had the potential of inhibiting the effects of PC on the area of GST-P-positive liver foci and the effects were associated with increasing expression of an anti-oxidative enzyme (Catalase). The results suggested that MR/NOX signaling might be involved in development of preneoplastic liver foci and renal basophilic changes in HFD-fed rats; however, the impacts of PC were different in each organ.
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http://dx.doi.org/10.2131/jts.43.611DOI Listing
November 2018

Concise Commentary: Quercetin Flavonoid of the Month or IBD Therapy?

Dig Dis Sci 2018 12;63(12):3305-3306

Laboratory of Veterinary Pathology, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo, 183-8509, Japan.

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http://dx.doi.org/10.1007/s10620-018-5269-zDOI Listing
December 2018

A case of rapid recurrence of apocrine ductal carcinoma originating from the oral scent gland of a Richardson's ground squirrel ().

J Toxicol Pathol 2018 Jul 29;31(3):189-193. Epub 2018 Mar 29.

Laboratory of Veterinary Pathology, Division of Animal Life Science, Institute of Agriculture, Tokyo University of Agriculture and Technology, 3-5-8 Saiwai-cho, Fuchu-shi, Tokyo 183-8509, Japan.

A 3-year-old female Richardson's ground squirrel developed a subcutaneous mass at the left oral angle. Seven days after removal of the mass, the mass recurred and metastasized to the cervical lymph node. Histologically, the primary mass was subdivided by fibrous trabeculae into various-sized neoplastic cell lobules showing a solid growth pattern with frequent mitoses and sometimes forming intracytoplasmic lumina. Large to medium-sized lobules formed a central cyst plugged by comedo necrosis. Neoplastic cells showed infiltrative subcutaneous growth. In the recurrent tumor, tubular structures lacking apparent apocrine secretion appeared within the solid growth portion. Neutrophil infiltration was evident within the tubules and intracytoplasmic lumina. Neoplastic cells were diffusely immunopositive for AE1/AE3 pan-cytokeratin (CK) in all lobules and focally positive for CAM5.2 CK in the lobules forming a central cyst and/or tubular structures, but they entirely lacked positivity for the periodic acid Schiff reaction. Ki-67-positive proliferating neoplastic cells were higher in numbers with the recurrent tumor than with the primary tumor. In addition, phosphorylated c-MYC immunoreactivity was observed in neoplastic cell nuclei, distinctly at the portion of invasive growth. Thus, the present case was diagnosed as apocrine ductal carcinoma originating from the oral scent gland, which typically shows highly aggressive biological behavior.
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http://dx.doi.org/10.1293/tox.2017-0071DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6077161PMC
July 2018
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