Prenatal test cohort of a modified rat comparative thyroid assay adding brain thyroid hormone measurements and histology but lowering group size appears able to detect disruption by sodium phenobarbital.

The Comparative Thyroid Assay (CTA, USEPA) is a screening test for thyroid hormone (TH) disruption in peripheral blood of dams and offspring. Recently, we began investigating feasible improvements to the CTA by adding examination of offspring brain TH concentrations and brain histopathology. In addition, we hypothesize that the number of animals required could be reduced by 50 % while still maintaining sensitivity to characterize treatment related changes in THs. Previously, we showed that the prenatal test cohort of the modified CTA could detect 1000 ppm sodium phenobarbital (NaPB)-induced suppression of brain T3 (by 9 %) and T4 (by 33 %) with no significant changes in serum T3 and T4 (less than 8 %). In the current study we expanded the dose response in a prenatal test cohort. Pregnant SD rats (N = 10/group) were exposed to 0, 1000 or 1500 ppm NaPB in the diet from gestational days (GD) 6 to GD20. Serum THs concentrations in GD20 dams together with serum/brain THs concentrations and brain histopathology in the GD20 fetuses were examined. NaPB dose-dependently suppressed serum T3 (up to -26 %) and T4 (up to -44 %) in dams, with suppression of T3 in serum (up to -26 %) and brain (up to -18 %) and T4 in serum (up to -26 %) and brain (up to -29 %) of fetuses but without clear dose dependency. There were no remarkable findings that deviated significantly from controls in GD20 fetal brain by qualitative histopathology. Overall, the present study suggests that the prenatal test cohort of this modified CTA is able to detect the expected fetal TH disruptions by prenatal exposure to NaPB, while also reducing the number of animals used by 50 %, consistent with the results of our previous study. These findings add to the suggestion that lowering group sizes and adding endpoints may be a useful alternative to the original CTA design.

Dysfunction of sinus macrophages in tumor-bearing host induces resistance to immunotherapy.

Sinus macrophages in draining lymph nodes (DLNs) are involved in anti-tumor immune reactions. CD169 (Sialoadhesin, Siglec-1) is expressed on sinus macrophages and is considered a surrogate marker for the immunostimulatory phenotype of macrophages. In this study, the significance of sinus macrophages in immunotherapy was evaluated using mouse models. Treatment with anti-programmed death-ligand 1 (PD-L1) antibody suppressed the subcutaneous tumor growth of MC38 and E0771 cells but was not effective against MB49 and LLC tumors. Decreased cytotoxic T-lymphocyte (CTL) infiltration in tumor tissues and CD169 expression in sinus macrophages were observed in MB49 and LLC cells compared to corresponding parameters in MC38 and E0771 cells. The anti-tumor effects of the anti-PD-L1 antibody on MC38 and E0771 cells were abolished when sinus macrophages in DLNs were depleted, suggesting that sinus macrophages are involved in the therapeutic effect of the anti-PD-L1 antibody. Naringin activated sinus macrophages. Naringin inhibited tumor growth in MB49- and LLC-bearing mice but did not affect that in MC38- and E0771-bearing mice. The infiltration of CTLs in tumor tissues and their activation were increased by naringin, and this effect was impaired when sinus macrophages were depleted. Combination therapy with naringin and anti-PD-L1 antibody suppressed MB49 tumor growth. In conclusion, CD169-positive sinus macrophages in DLNs are critical for anti-tumor immune responses, and naringin suppresses tumor growth by activating CD169-positive sinus macrophages and anti-tumor CTL responses. The activation status of sinus macrophages has been suggested to differ among tumor models, and this should be investigated in future studies.

Smooth muscle hamartoma of the lungs in a Wistar Hannover rat.

Hamartomas are tumor-like masses comprising disorganized normal tissue elements. To date, spontaneous hamartomas have been reported in several organs and tissues in rodents but not in the lungs. Here, we report the first case of a hamartoma in the lungs of a 108-week-old female Wistar Hannover rat. Grossly, a white spot, 7 mm in diameter, was observed on the costal surface of the left lung. Histopathologically, the nodular lesions adjacent to the bronchioles comprised mature smooth muscle cells. The lesion was not encapsulated and spread along the alveolar walls and ducts without compression of the surrounding tissue. In the nodules, elastic fibers enclosed small lumens lined with factor VIII-related antigen-positive endothelial cells. This structure suggested that the nodule mimicked an artery. Moreover, structural abnormalities were observed within the bronchioles and arterioles owing to the increased number of smooth muscle cells in the surrounding tissues. These features suggested that this was a case of tissue malformation rather than a neoplasm, leading to the diagnosis of a smooth muscle hamartoma of the lung.

Ectopic pancreatic acinar cell carcinoma in the thoracic cavity of F344 rat.

Ectopic pancreatic tissue can occasionally cause inflammation, hemorrhage, stenosis, and invagination, similar to normal pancreatic tissue; however, tumorigenesis is rare. This case report describes an ectopically observed pancreatic acinar cell carcinoma in the thoracic cavity of a female Fischer (F344/DuCrlCrlj) rat. Histopathologically, polygonal tumor cells with periodic acid-Schiff-positive cytoplasmic eosinophilic granules showed solid proliferation and infrequently formed acinus-like structures. Immunohistochemically, the tumor cells were positive for cytokeratin, trypsin, and human B-cell leukemia/lymphoma 10, which specifically reacted with pancreatic acinar cells, and negative for vimentin and human α-smooth muscle actin. Ectopic pancreas develops in the submucosa of the gastrointestinal tract; however, there are few reports of its development and neoplasia in the thoracic cavity. To the best of our knowledge, this is the first report of ectopic pancreatic acinar cell carcinoma in the thoracic cavity of a rat.

iPS cell-derived model to study the interaction between tissue macrophage and HIV-1.

Despite effective antiretroviral therapy, HIV-1 persists in cells, including macrophages, which is an obstacle to cure. However, the precise role of macrophages in HIV-1 infection remains unclear because they reside in tissues that are not easily accessible. Monocyte-derived macrophages are widely used as a model in which peripheral blood monocytes are cultured and differentiated into macrophages. However, another model is needed because recent studies revealed that most macrophages in adult tissues originate from the yolk sac and fetal liver precursors rather than monocytes, and the embryonic macrophages possess a self-renewal (proliferating) capacity that monocyte-derived macrophages lack. Here, we show that human induced pluripotent stem cell-derived immortalized macrophage-like cells are a useful self-renewing macrophage model. They proliferate in a cytokine-dependent manner, retain macrophage functions, support HIV-1 replication, and exhibit infected monocyte-derived macrophage-like phenotypes, such as enhanced tunneling nanotube formation and cell motility, as well as resistance to a viral cytopathic effect. However, several differences are also observed between monocyte-derived macrophages and induced pluripotent stem cell-derived immortalized macrophage-like cells, most of which can be explained by the proliferation of induced pluripotent stem cell-derived immortalized macrophage-like cells. For instance, proviruses with large internal deletions, which increased over time in individuals receiving antiretroviral therapy, are enriched more rapidly in induced pluripotent stem cell-derived immortalized macrophage-like cells. Interestingly, inhibition of viral transcription by HIV-1-suppressing agents is more obvious in induced pluripotent stem cell-derived immortalized macrophage-like cells. Collectively, our present study proposes that the model of induced pluripotent stem cell-derived immortalized macrophage-like cells is suitable for mimicking the interplay between HIV-1 and self-renewing tissue macrophages, the newly recognized major population in most tissues that cannot be fully modeled by monocyte-derived macrophages alone.

Gene expression analysis of antioxidant and DNA methylation on the rat liver after 4-week wood preservative chromated copper arsenate exposure.

Our previous 4-week repeated dose toxicity study showed that wood preservative chromated copper arsenate (CCA) induced hepatocellular hypertrophy accompanied by biochemical hepatic dysfunction and an increase in oxidative stress marker, 8-hydroxydeoxyguanosine, in female rats. To further explore the molecular mechanisms of CCA hepatotoxicity, we analyzed 10%-buffered formalin-fixed liver samples from female rats for cell proliferation, apoptosis, and protein glutathionylation and conducted microarray analysis on frozen liver samples from female rats treated with 0 or 80 mg/kg/day of CCA. Chemical analysis revealed that dimethylated arsenical was the major metabolite in liver tissues of male and female rats. CCA increase labeling indices of proliferating cell nuclear antigen and decrease terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling accompanied with increased expression of protein glutathionylation, indicating a decrease in glutathione (GSH) in hepatocytes of female rats. Microarray analysis revealed that CCA altered gene expression of antioxidants, glutathione-S-transferase (GST), heat shock proteins and ubiquitin-proteasome pathway, cell proliferation, apoptosis, DNA methylation, cytochrome P450, and glucose and lipid metabolism in female rats. Increased expression of GSTs, including Gsta2, Gsta3, Mgst1, and Cdkn1b (p27), and decreased expression of the antioxidant Mt1, and DNA methylation Dnmt1, Dnmt3a, and Ctcf were confirmed in the liver of female rats in a dose-dependent manner. Methylation status of the promoter region of the Mt1 was not evidently changed between control and treatment groups. The results suggested that CCA decreased GSH and altered the expression of several genes, including antioxidants, GST, and DNA methylation, followed by impaired cell proliferation in the liver of female rats.

Feasibility study for a downsized comparative thyroid assay with measurement of brain thyroid hormones and histopathology in rats: Case study with 6-propylthiouracil and sodium phenobarbital at high dose.

Concern has been raised that thyroid hormone disruptors (THDs) may potentially interfere with the developing brain, but effects of mild suppression of maternal THs by environmental contaminants on neonatal brain development are not fully understood. The comparative thyroid assay (CTA) is a screening test for offspring THDs, but it requires several animals and is criticized that reliance on serum THs alone as predictive markers of brain malfunction is inadequate. To verify feasibility of the downsized CTA but additional examination of brain THs levels and histopathology, we commenced internal-validation studies. This paper presents the data of the study where 6-propylthiouracil (6-PTU, 10 ppm) and sodium phenobarbital (NaPB, 1000 ppm) were dosed by feeding from gestational days (GD)6-20, and from GD6 to lactation day 21. The modified CTA detected 6-PTU-induced severe (>70%) suppression of serum THs in dams, with >50% suppressed serum/brain TH levels in offspring and brain heterotopia in postnatal day 21 pups. The modified CTA also detected NaPB-induced mild (<35%) suppression of serum THs in dams, with mild (<35%) reduction of serum/brain TH levels in fetuses but not in pups. These findings suggest that the modified CTA may have a potential as a screening test for offspring THDs.

Inhibitory and Stimulatory Effects of IL-32 on HIV-1 Infection.

The proinflammatory cytokine IL-32 is elevated in the plasma and tissues of HIV-1-infected individuals. However, its significance in HIV-1 infection remains unclear because IL-32 inhibits and stimulates viral production in monocyte-derived macrophages (MDMs) and CD4+ T cells, respectively. In this study, we initially found that the inhibitory effect on human MDMs depends on SAMHD1, a dNTP triphosphohydrolase that inhibits viral reverse transcription. IL-32 increased the unphosphorylated active form of SAMHD1, which was consistent with the reduced expression of the upstream cyclin-dependent kinases. Indeed, IL-32 lost its anti-HIV-1 activity in MDMs when SAMHD1 was depleted. These results explain why IL-32 inhibits HIV-1 in MDMs but not CD4+ T cells, because SAMHD1 restricts HIV-1 in noncycling MDMs but not in cycling CD4+ T cells. Another unique feature of IL-32 is the induction of the immunosuppressive molecule IDO1, which is beneficial for HIV-1 infection. In this study, we found that IL-32 also upregulates other immunosuppressive molecules, including PD-L1, in MDMs. Moreover, IL-32 promoted the motility of MDMs, which potentially facilitates intercellular HIV-1 transmission. Our findings indicate that IL-32 has both the direct inhibitory effect on HIV-1 production in MDMs and the indirect stimulatory effects through phenotypic modulation of MDMs, and they suggest that the stimulatory effects may outweigh the inhibitory effect because the window for IL-32 to inhibit HIV-1 is relatively confined to SAMHD1-mediated reverse transcription suppression in the viral life cycle.

The potential of organoids in toxicologic pathology: role of toxicologic pathologists in in vitro chemical hepatotoxicity assessment.

The development of in vitro toxicity assessment methods using cultured cells has gained popularity for promoting animal welfare in animal experiments. Herein, we briefly discuss the current status of hepatoxicity assessment using human- and rat-derived hepatocytes; we focus on the liver organoid method, which has been extensively studied in recent years, and discuss how toxicologic pathologists can use their knowledge and experience to contribute to the development of in vitro chemical hepatotoxicity assessment methods for drugs, pesticides, and chemicals. We also propose how toxicological pathologists should assess toxicity regarding the putative distribution of undifferentiated and differentiated cells in the organoid when liver organoids are observed in hematoxylin and eosin-stained specimens. This was done while considering the usefulness and limitations of in vitro studies for toxicologic pathology assessment.

Metronidazole enhances steatosis-related early-stage hepatocarcinogenesis in high fat diet-fed rats through DNA double-strand breaks and modulation of autophagy.

Nonalcoholic fatty liver disease is a hepatic disorder with deposition of fat droplets and has a high risk of progression to steatosis-related hepatitis and irreversible hepatic cancer. Metronidazole (MNZ) is an antiprotozoal and antimicrobial agent widely used to treat patients infected with anaerobic bacteria and intestinal parasites; however, MNZ has also been shown to induce liver tumors in rodents. To investigate the effects of MNZ on steatosis-related early-stage hepatocarcinogenesis, male rats treated with N-nitrosodiethylamine following 2/3 hepatectomy at week 3 were received a control basal diet, high fat diet (HFD), or HFD containing 0.5% MNZ. The HFD induced obesity and steatosis in the liver, accompanied by altered expression of Pparg and Fasn, genes related to lipid metabolism. MNZ increased nuclear translocation of lipid metabolism-related transcription factor peroxisome proliferator-activated receptor gamma in hepatocytes, together with altered liver expression of lipid metabolism genes (Srebf1, Srebf2, Pnpla2). Furthermore, MNZ significantly increased the number of preneoplastic liver foci, accompanied by DNA double-strand breaks and late-stage autophagy inhibition, as reflected by increased levels of γ-H2AX, LC3, and p62. Therefore, MNZ could induce steatosis-related hepatocarcinogenesis by inducing DNA double-strand breaks and modulating autophagy in HFD-fed rats.

M-Sec induced by HTLV-1 mediates an efficient viral transmission.

Human T-cell leukemia virus type 1 (HTLV-1) infects target cells primarily through cell-to-cell routes. Here, we provide evidence that cellular protein M-Sec plays a critical role in this process. When purified and briefly cultured, CD4+ T cells of HTLV-1 carriers, but not of HTLV-1- individuals, expressed M-Sec. The viral protein Tax was revealed to mediate M-Sec induction. Knockdown or pharmacological inhibition of M-Sec reduced viral infection in multiple co-culture conditions. Furthermore, M-Sec knockdown reduced the number of proviral copies in the tissues of a mouse model of HTLV-1 infection. Phenotypically, M-Sec knockdown or inhibition reduced not only plasma membrane protrusions and migratory activity of cells, but also large clusters of Gag, a viral structural protein required for the formation of viral particles. Taken together, these results suggest that M-Sec induced by Tax mediates an efficient cell-to-cell viral infection, which is likely due to enhanced membrane protrusions, cell migration, and the clustering of Gag.

Declining neutrophil production despite increasing G-CSF levels is associated with chronic inflammation in elderly rhesus macaques.

Aging is characterized by a loss of bone marrow hematopoietic tissue, systemic chronic inflammation, and higher susceptibility to infectious and noninfectious diseases. We previously reported the tightly regulated kinetics and massive daily production of neutrophils during homeostasis in adult rhesus macaques aged 3 to 19 yr (equivalent to approximately 10 to 70 yr of age in humans). In the current study, we observed an earlier release of recently dividing neutrophils from bone marrow and greater in-group variability of neutrophil kinetics based on in vivo BrdU labeling in a group of older rhesus macaques of 20-26 yr of age. Comparing neutrophil numbers and circulating cytokine levels in rhesus macaques spanning 2 to 26 yr of age, we found a negative correlation between age and blood neutrophil counts and a positive correlation between age and plasma G-CSF levels. Hierarchic clustering analysis also identified strong associations between G-CSF with the proinflammatory cytokines, IL-1ß and MIP-1α. Furthermore, neutrophils from older macaques expressed less myeloperoxidase and comprised higher frequencies of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) compared to the young adult macaques. In summary, we observed an earlier release from bone marrow and a reduced production of neutrophils despite the increased levels of plasma G-CSF, especially in the elderly rhesus macaques. This lower neutrophil production capacity associated with increased production of proinflammatory cytokines as well as an earlier release of less mature neutrophils and PMN-MDSCs may contribute to the chronic inflammation and greater susceptibility to infectious and noninfectious diseases during aging.

Comparison of predictors for terminal disease progression in simian immunodeficiency virus/simian-HIV-infected rhesus macaques.

OBJECTIVES: CD4+ T-cell decline and increasing virus levels are considered hallmarks of HIV/AIDS pathogenesis but we previously demonstrated in rhesus macaques that tissue macrophage destruction by simian immunodeficiency virus (SIV) infection associated with increased monocyte turnover also appear to impact pathogenesis. It remains unclear, however, which factors best predict onset of terminal disease progression and survival time. The objective of this study, therefore, was to directly compare these co-variates of infection for predicting survival times in retrospective studies of SIV/simian-HIV (SHIV)-infected adult rhesus macaques. METHODS: Rhesus macaques were infected with various strains of SIV/SHIV and evaluated longitudinally for monocyte turnover, CD4+ T-cell loss, plasma viral load, and SIV/SHIV strain. Correlation analyses and machine learning algorithm modeling were applied to compare relative contributions of each of the co-variates to survival time. RESULTS: All animals with AIDS-related clinical signs requiring euthanasia exhibited increased monocyte turnover regardless of CD4+ T-cell level, viral strain, or plasma viral load. Regression analyses and machine learning algorithms indicated a stronger correlation and contribution between increased monocyte turnover and reduced survival time than between CD4+ T-cell decline, plasma viral load, or virus strain and reduced survival time. Decision tree modeling categorized monocyte turnover of 13.2% as the initial significant threshold that best predicted decreased survival time. CONCLUSION: These results demonstrate that monocytes/macrophages significantly affect HIV/SIV pathogenesis outcomes. Monocyte turnover analyses are not currently feasible in humans, so there is a need to identify surrogate biomarkers reflecting tissue macrophage damage that predict HIV infection disease progression.

An extraskeletal osteosarcoma in the auricle of a Wistar Hannover rat.

An extraskeletal osteosarcoma was detected in the auricle of a 110-week-old female Wistar Hannover rat. Grossly, the tumor, measuring 15 mm in size, was observed in the subcutis as a solid and hard mass. Histologically, the majority of the mass comprised mature, compact bone. It was surrounded by neoplastic cells showing a variety of histologies, such as sarcoma, not otherwise specified, and myxosarcoma away from the bone-forming region. However, these different histological regions were considered to be components of a single bone tumor, based on the common expression of osterix and a similar mixture of constituent cells in each region. The tumor was diagnosed as an extraskeletal osteosarcoma because of the presence of infiltrative growth and abnormal mitosis and its development in the auricle without attachment to the skeleton. The present case is a rare histological type of an extraskeletal osteosarcoma with independent and different histological elements in rats.

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

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 × 102/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.

Shifting Dynamics of Intestinal Macrophages during Simian Immunodeficiency Virus Infection in Adult Rhesus Macaques.

The intestinal tract is a primary barrier to invading pathogens and contains immune cells, including lymphocytes and macrophages. We previously reported that CD163+CD206- (single-positive [SP]) interstitial macrophages of the lung are short-lived and succumb early to SIV infection. Conversely, CD163+CD206+ (double-positive [DP]) alveolar macrophages are long-lived, survive after SIV infection, and may contribute to the virus reservoir. This report characterizes analogous populations of macrophages in the intestinal tract of rhesus macaques (Macaca mulatta) with SIV/AIDS. By flow cytometry analysis, immunofluorescence staining, and confocal microscopy, CD163+CD206+ DP macrophages predominated in the lamina propria of uninfected animals, compared with CD163+CD206- SP macrophages, which predominated in the lamina propria in animals with SIV infection that were exhibiting AIDS. In submucosal areas, CD163+CD206+ DP macrophages predominated in both SIV-infected and uninfected macaques. Furthermore, BrdU-labeled CD163+CD206+ DP and CD163+CD206- SP macrophages recently arriving in the colon, which are both presumed to be shorter-lived, were observed to localize only in the lamina propria. Conversely, longer-lived CD163+CD206+ DP macrophages that retained dextran at least 2 mo after in vivo administration localized exclusively in the submucosa. This suggests that CD163+CD206+ DP intestinal macrophages of the lamina propria were destroyed after SIV infection and replaced by immature CD163+CD206- SP macrophages, whereas longer-lived CD163+CD206+ DP macrophages remained in the submucosa, supporting their potential role as an SIV/HIV tissue reservoir. Moreover, the DP macrophages in the submucosa, which differ from lamina propria DP macrophages, may be missed from pinch biopsy sampling, which may preclude detecting virus reservoirs for monitoring HIV cure.

Subacute oral administration of folic acid elicits anti-inflammatory response in a mouse model of allergic dermatitis.

Folic acid (FA) deficiency is associated with several health problems, including megaloblastic anemia and fetal neural tube defects. Therefore, supplementation with FA is strongly recommended by governments worldwide. Recent published reports indicate that FA functions in immune system maintenance. The main objective of this study is to examine possible anti-inflammatory and antipruritic effects of FA using a mouse model of allergic dermatitis. The mouse model was developed by repetitive sensitization to the Th2-type hapten toluene-2,4-diisocyanate (TDI). During the development of allergic dermatitis, FA was orally administered to the mice at doses of 8, 160, 1000 or 10,000 µg/day for 5 weeks. The ear swelling response and scratching behavior were monitored after the TDI challenge. Serum, ear tissue and auricular lymph node samples were isolated for further analysis 24 h after the TDI challenge. The ear swelling response was reduced in a dose-dependent manner by FA administration, and a significant change was observed at a concentration of 10,000-µg/day group. Comparable results were obtained through histological evaluation and cytokine level measurement in the ear tissue samples. Oral administration of FA exhibited the inhibitory effect on T-cell infiltration and T-cell-related cytokine production in auricular lymph nodes. Scratching behavior was not altered by FA administration. The in vivo evidence was corroborated by in vitro results, which showed that FA treatment significantly interfered with T-cell proliferation in a dose-dependent manner. Our findings imply that subacute oral administration of FA elicits an anti-inflammatory response, mainly through inhibition of T-cell proliferation.

Human leukocyte antigen-associated gag and nef polymorphisms in HIV-1 subtype A/E-infected individuals in Vietnam.

Numbers of HLA-associated polymorphisms have been reported on HIV-1 subtypes B and C, but few on other subtypes. Here, we analyzed HLA-associated gag and nef polymorphisms in HIV-1 subtype A/E prevalent in Vietnam. We determined HLA-A, B and C genotypes in 179 HIV-1-infected Vietnamese by next generation sequencing and analyzed proviral genome sequences in 144 of them, showing that 142 of the 144 were subtype A/E. Analysis revealed HLA-associated subtype A/E gag and nef polymorphisms at nineteen residues including those newly determined. Accumulation of these data would contribute to our understanding of HIV-1 subtype A/E and host immune interaction.

Spironolactone in Combination with α-glycosyl Isoquercitrin Prevents Steatosis-related Early Hepatocarcinogenesis in Rats through the Observed NADPH Oxidase Modulation.

Administration of the diuretic, spironolactone (SR), can inhibit chronic liver diseases. We determined the effects of SR alone or in combination with the antioxidant α-glycosyl isoquercitrin (AGIQ) on hyperlipidemia- and steatosis-related precancerous lesions in high-fat diet (HFD)-fed rats subjected to a two-stage hepatocarcinogenesis model. Rats were fed with control basal diet or HFD, which was administered with SR alone or in combination with an antioxidant AGIQ in drinking water. An HFD increased body weight, intra-abdominal fat (adipose) tissue weight, and plasma lipids, which were reduced by coadministration of SR and AGIQ. SR and AGIQ coadministration also reduced hepatic steatosis and preneoplastic glutathione S-transferase placental form-positive foci, in association with decrease in NADPH oxidase (NOX) subunit p22phox-positive cells and an increase in active-caspase-3-positive cells in the foci. Hepatic gene expression analysis revealed that the coadministration of SR and AGIQ altered mRNA levels of lipogenic enzymes ( Scd1 and Fasn), antioxidant-related enzymes ( Catalase), NOX component ( P67phox), and anti-inflammatory transcriptional factor ( Pparg). Our results indicated that SR in combination with AGIQ had the potential of suppressing hyperlipidemia- and steatosis-related early hepatocarcinogenesis through the reduced expression of NOX subunits.

Spontaneous malignant myoid thymoma in an aged female Fischer 344 rat.

A whitish mass approximately 30 mm in diameter was noted in the anterior mediastinum of a 67-week-old female Fischer 344 rat. Histopathologically, two types of tumor cells were identified on the basis of morphologic features: epithelial tumor cells with a tubular or cord-like growth pattern and rhabdomyosarcomatous tumor cells characterized by the presence of cross-striations. Immunohistochemically, the epithelial tumor cells reacted positively for cytokeratin AE1/AE3, and some reacted positively for p63, which is expressed in normal thymic epithelial cells. The rhabdomyosarcomatous tumor cells stained positively for desmin, sarcomeric actin, and S-100 protein, which coincides with the stainability of normal thymic myoid cells. Since the tumor was also found to have malignant features such as high proliferative activity, cytologic atypia, and necrotic behavior, it was diagnosed as a malignant myoid thymoma. We believe that this is the first case report of such a tumor in a rodent.