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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, with50% 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.

In the mouse carcinogenicity study, an apparent increase in lung adenocarcinoma was observed in male mice at 7000 ppm. Based on the overall evaluation of toxicology, oncology, pathology and statistics, we concluded that the apparent increase in lung tumors is not relevant for evaluation of carcinogenicity of imiprothrin (Regul Toxicol Pharmacol, 105, 1–14, 2019). To investigate whether imiprothrin has any mitogenic effect on mouse Club cells, the present study examined its effects on replicative DNA synthesis of Club cells and lung histopathology in male mice treated with imiprothrin for 7 days at 3500 and 7000 ppm in the diet. Isoniazid, a known mouse lung mitogen and tumor inducer, was also examined at 1000 ppm in the diet as a positive control of Club cell mitogenesis and morphological changes. Neither imiprothrin nor isoniazid caused any necrotic changes in lung by light or electron microscopy. There were no increases observed in the bromodeoxyuridine (BrdU) labeling index in the imiprothrin groups, while there was a statistically significant increase in the BrdU labeling index in the isoniazid group. These findings demonstrate that imiprothrin does not induce mouse Club cell proliferation or morphologic changes, supporting our previous conclusion described above. Thus, imiprothrin should not be classified as a carcinogen. Furthermore, this study indicates that short-term studies focusing on cell proliferation can be reliable for predicting a lack of carcinogenic potential of test chemicals.

Preadipocyte factor-1 (pref-1) is specifically expressed in preadipocytes and acts as a gatekeeper of adipogenesis by maintaining the preadipocyte state and preventing adipocyte differentiation. We hypothesized that the breed differences of adipogenic capacity in cattle could be explained by the expression level of pref-1. In this experiment, we studied the expression level of the pref-1 gene and adipocyte cellularity in subcutaneous and mesenteric adipose tissues of Japanese Black (Wagyu) and Holstein fattening cattle. In subcutaneous adipose tissue, there were no significant differences in the pref-1 gene expression levels and adipocyte sizes between the breeds. In contrast, the expression level of the pref-1 gene in mesenteric adipose tissue of Holsteins was significantly higher than that of Wagyu. In addition, the size of mesenteric adipocytes in Holsteins was significantly smaller than that of Wagyu. These results indicate that the breed differences of fattening cattle affect the expression pattern of the pref-1 gene and adipocyte cellularity in a fat depot-specific manner.

We develop canonical correlation analysis in the context of two-step monotone incomplete data drawn from N"p"+"q(@m,@S), a multivariate normal population with mean @m and covariance matrix @S. Our data consist of n observations on each group and an additional N-n observations on only one group, where all observations are mutually independent. We perform the canonical correlation analysis using the maximum likelihood estimators, with the monotone incomplete data, of @m and @S. Further, we derive the asymptotic expansion of the distributions of the canonical correlations and the limiting distributions of the canonical vectors, and we compare them with the results of a typical canonical correlation.

We establish the Stein phenomenon in the context of two-step, monotone incomplete data drawn from N"p"+"q(@m,@S), a (p+q)-dimensional multivariate normal population with mean @m and covariance matrix @S. On the basis of data consisting of n observations on all p+q characteristics and an additional N-n observations on the last q characteristics, where all observations are mutually independent, denote by @[email protected]^ the maximum likelihood estimator of @m. We establish criteria which imply that shrinkage estimators of James-Stein type have lower risk than @[email protected]^ under Euclidean quadratic loss. Further, we show that the corresponding positive-part estimators have lower risk than their unrestricted counterparts, thereby rendering the latter estimators inadmissible. We derive results for the case in which @S is block-diagonal, the loss function is quadratic and non-spherical, and the shrinkage estimator is constructed by means of a nondecreasing, differentiable function of a quadratic form in @[email protected]^. For the problem of shrinking @[email protected]^ to a vector whose components have a common value constructed from the data, we derive improved shrinkage estimators and again determine conditions under which the positive-part analogs have lower risk than their unrestricted counterparts.

We hypothesized that dietary roughage level would alter the expression levels of adipogenic transcription factors in adipose tissue of Japanese black (Wagyu) steers. Steers were fed whole crop rice silage at three levels: (1) high-roughage feeding group, fed 8 kg silage and 5 kg concentrate (HR); (2) middle roughage feeding group, fed 5 kg silage and 6 kg concentrate (MR); and (3) low roughage feeding group, fed 2 kg silage and 7 kg concentrate (LR) from 22 to 30 months of age. In subcutaneous adipose tissue, there were no significant differences in the expression of the adipogenic transcription factors and adipocyte size among feeding groups. In mesenteric adipose tissue, the expression of C/EBPα in the LR and MR groups was significantly higher than that in the HR group. Adipocyte size in the mesenteric adipose tissue of the LR group was significantly larger than that of the HR group. In intermuscular adipose tissue, the expression of C/EBPβ-LAP in the LR group was significantly higher than that in the HR group, and the expression of C/EBPβ-LIP in the LR and MR groups was significantly higher than that in the HR group. Adipocyte size in the intermuscular adipose tissue of the LR and MR groups was significantly smaller than that of the HR group. These results suggest that dietary roughage revel affects the adipose tissue depot-specific differences in C/EBP family expression pattern and adipocyte cellularity in Wagyu steers.

High doses of Metofluthrin (MTF) have been shown to produce liver tumours in rats by a mode of action (MOA) involving activation of the constitutive androstane receptor leading to liver hypertrophy, induction of cytochrome P450 (CYP) forms and increased cell proliferation. The aim of this study was to compare the effects of MTF with those of the known rodent liver tumour promoter phenobarbital (PB) on the induction CYP2B forms and replicative DNA synthesis in cultured rat and human hepatocytes. Treatment with 50 microM MTF and 50 microM PB for 72 h increased CYP2B1 mRNA levels in male Wistar rat hepatocytes and CYP2B6 mRNA levels in human hepatocytes. Replicative DNA synthesis was determined by incorporation of 5-bromo-2'-deoxyuridine over the last 24 h of a 48 h treatment period. Treatment with 10-1000 microM MTF and 100-500 microM PB resulted in significant increases in replicative DNA synthesis in rat hepatocytes. While replicative DNA synthesis was increased in human hepatocytes treated with 5-50 ng/ml epidermal growth factor or 5-100 ng/ml hepatocyte growth factor, treatment with MTF and PB had no effect. These results demonstrate that while both MTF and PB induce CYP2B forms in both species, MTF and PB only induced replicative DNA synthesis in rat and not in human hepatocytes. These results provide further evidence that the MOA for MTF-induced rat liver tumour formation is similar to that of PB and some other non-genotoxic CYP2B form inducers and that the key event of increased cell proliferation would not occur in human liver.

In this experiment, we studied the effects of breed differences on the protein expression of adipogenic transcription factors, the C/EBP family (C/EBPα, C/EBPβ-LAP, C/EBPβ-LIP and C/EBPδ) and PPARγ, in the adipose tissues of Japanese Black (Wagyu) and Holstein steers from various anatomical sites (subcutaneous, intermuscular, and mesenteric) at different fattening periods (19 and 24 months of age). The expression of C/EBPβ-LAP and C/EBPα in the mesenteric fat tissue of Wagyu at 19 months of age was significantly higher than that of Holstein. The expression of C/EBPδ in the subcutaneous, intermuscular and mesenteric fat tissue of Wagyu at 19 months of age was significantly higher than that of Holstein. The plasma insulin concentrations of Wagyu steers at 19 months of age tended to be higher than those of Holstein. No significant differences in the expression of the adipogenic transcription factors and plasma insulin concentration were observed between the breeds at 24 months of age. These results suggest the existence of breed difference on the expression of the C/EBP family between fattening Wagyu and Holstein steers at 19 months of age, whereas breed difference might have disappeared before 24 months of age.

Using a radioimmunoassay (RIA) with high specificity and sensitivity (1 pg/tube) for acetylcholine (ACh), we have been able to measure the ACh content in samples from the bacteria, archaea and eucarya domains of the universal phylogenetic tree. We found detectable levels of ACh to be ubiquitous in bacteria (e.g., Bacillus subtilis), archaea (e.g., Thermococcus kodakaraensis KOD1), fungi (e.g., shiitake mushroom and yeast), plants (e.g., bamboo shoot and fern) and animals (e.g., bloodworm and lugworm). The levels varied considerably, however, with the highest ACh content detected in the top portion of bamboo shoot (2.9 μmol/g), which contained about 80 times that found in rat brain. In addition, using the method of Fonnum, various levels of ACh-synthesizing activity also were detected, a fraction of which was catalyzed by a choline acetyltransferase (ChAT)-like enzyme (sensitive to bromoACh, a selective ChAT inhibitor) in T. kodakaraensis KOD1 (15%), bamboo shoot (91%) and shiitake mushroom (51%), bloodworm (91%) and lugworm (81%). Taken together, these findings demonstrate the ubiquitous expression of ACh and ACh-synthesizing activity among life forms without nervous systems, and support the notion that ACh has been expressed and may be active as a local mediator and modulator of physiological functions since the early beginning of life.

In canonical correlation analysis, we are interested in testing whether the @ath canonical correlation coefficient is some number, especially the first canonical correlation coefficient. In this paper, we try the permutation test in canonical correlation analysis and suggest some test statistics.

Although benomyl and its metabolite carbendazim have been shown to adversely affect male reproduction, the mechanisms of action do not appear to involve the endocrine system. However, few studies have been conducted using currently proposed tests specifically focused on endocrine disruption. Here, potential estrogen- and androgen-mediated activity of benomyl was therefore investigated in vitro and in vivo. Benomyl and carbendazim proved negative for agonistic and antagonistic activity in reporter gene assays for the human estrogen receptor alpha and androgen receptor. In uterotrophic and Hershberger assays using Crj:CD(SD)IGS rats, benomyl (100, 300 or 1000 mg/kg/day, p.o., N = 6) did not exert agonistic effects. However, the highest dose decreased uterine weights in the uterotrophic assay, and decreased weights of some androgen-related tissues of castrated rats receiving a testosterone propionate (TP, 0.2 mg/kg) injection in the Hershberger assay; the effects were less severe than those with p,p'-DDE (100 mg/kg/day). When 4 mg/kg/day of TP was injected, decrease of organ weights due to benomyl was attenuated but still observed. Thus, its influence in some tissues was more potent than that of p,p'-DDE. Benomyl had no apparent effects on serum androgen levels. Microarray analysis of the gene expression profile in the ventral prostate of TP-injected castrated rats treated with benomyl indicated clear differences from the patterns observed with p,p'-DDE and flutamide. Taken together, these findings suggest the decreased organ weights observed in vivo to be caused by mechanisms that are not steroid-receptor-mediated, such as interfering with assembly of microtubules by benomyl. The study furthermore suggests that functional genomics may provide a reliable evidence for accurate categorization of test chemicals.

A new E-beam excited plasma reactor was developed for sub-100-nm etching, which had an interface film between an E-beam source and a reaction chamber. The plasma was excited by an E-beam that was transmitted through a 1-μm polyimide interface film. This ultra thin polyimide film was fabricated by a new method of spin coating on a copper plate with evaporated aluminium film. The maximum transmitted E-beam current was 3 mA for a 30-mm φ interface window, which was enough to induce plasma. A fine pattern of 50-nm lines and spaces was successfully fabricated by CF4 plasma etching on a Si wafer through a resist mask.

We examined whether the repeated oral administration with a high dose of l -3-(3,4-dihydroxyphenyl)-alanine ( l -DOPA) in 0.5% carboxymethyl cellulose increases serum luteinizing hormone (LH) levels in male rats. Serum LH levels were increased 4 h after a single administration of 1000 mg/kg l -DOPA to male rats, and returned to control levels within 8 h after administration. Four hours after a single administration, serum LH levels were significantly increased by l -DOPA at 1000 mg/kg, but not at 20, 100 or 200 mg/kg. Decreases in body weight and relative weight of the prostate were observed after 7 and 14 days of administration of 1000 mg/kg per day l -DOPA, but no changes were observed in weight of the testis, epididymis or seminal vesicle. The administration of l -DOPA at 500 or 1000 mg/kg per day for 7 or 14 days resulted in increased basal serum LH levels and decreased basal serum prolactin levels 24 h after the last administration. Serum testosterone levels tended to be higher in treated than in control rats. The levels of two metabolites of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), in rats treated with 500 mg/kg per day tended to be slightly higher than those in control rats after 7 days of administration. Levels of DA, DOPAC and HVA were significantly increased after 7 and 14 days of administration of 1000 mg/kg per day and after 14 days of administration of 500 mg/kg per day. The level of norepinephrine, but not its metabolite 3-methoxy-4-hydroxyphenylglycol, was significantly increased after only 7 days of administration of 1000 mg/kg per day. No significant changes were observed in levels of 5-hydroxytryptamine or its metabolite 5-hydroxyindole-3-acetic acid with administration of 500 or 1000 mg/kg per day. These findings suggest that a prolonged treatment with a high dose of l -DOPA in male rats induces release of LH from the pituitary, resulting in sustained elevation of LH levels in peripheral circulation.

Studies were performed to examine the mechanism by which testicular Leydig cell tumors are induced in rats by administration of the antimicrobial agent oxolinic acid (1-ethyl-1,4-dihydro-6,7-methylenedioxy-4-oxo-3-quinolinecarboxylic acid). In these studies, the effects of oxolinic acid on serum levels of luteinizing hormone (LH), testosterone, and prolactin and the binding of testosterone to prostatic androgen receptors were examined. In a long-term hormonal study, male Wistar rats were fed a diet containing oxolinic acid at 0, 100, 1000, or 3000 ppm for 104 weeks. A statistically significant increase in serum LH levels was observed at 1000 and 3000 ppm, but no dose of oxolinic acid had a significant effect on serum testosterone levels. Serum LH levels were no longer elevated above control levels within 2 weeks of cessation of the administration of oxolinic acid. Oxolinic acid was found to have no effect on the rate of clearance of exogenous LH from the circulation. Serum prolactin levels were decreased by the administration of oxolinic acid. The increase in serum LH induced by oxolinic acid was completely blocked by the intraperitoneal injection of the dopamine antagonist haloperidol (2 mg/kg). In addition, no significant affinity of oxolinic acid for androgen receptors was found in an in vitro study. These findings suggest that: (1) oxolinic acid induces Leydig cell tumors in rats by chronically stimulating the release of LH from the pituitary, (2) the mechanism of stimulating the release of LH involves facilitation of the dopaminergic systems in the hypothalamus-pituitary axis, and (3) oxolinic acid has no effect on androgen-mediated feedback inhibition.

We have measured heat capacities of 4 He and 3 He films adsorbed on a new mesoporous material with the pore diameter 27 A called HMM-2, which has an ordered 3D network of pores. The substrate is an organic–inorganic hybrid material made of C2H4 and Si2O3, and consists of cages connected in the HCP structure (a=55.4 A , c=88.6 A ) . When adsorptions are above one layer, heat capacities are qualitatively different between 3 He and 4 He . It indicates that the Bose fluid of the 4 He film appears at the second layer.