The stem cell growth factor receptor KIT is not expressed on interstitial cells in bladder.

PubMed

Gevaert, Thomas; Ridder, Dirk De; Vanstreels, Els; Daelemans, Dirk; Everaerts, Wouter; Aa, Frank Van Der; Pintelon, Isabel; Timmermans, Jean-Pierre; Roskams, Tania; Steiner, Clara; Neuhaus, Jochen

2017-06-01

The mast/stem cell growth factor receptor KIT has long been assumed to be a specific marker for interstitial cells of Cajal (ICC) in the bladder, with possible druggable perspectives. However, several authors have challenged the presence of KIT + ICC in recent years. The aim of this study was therefore to attempt to clarify the conflicting reports on KIT expression in the bladder of human beings, rat, mouse and guinea pig and to elucidate the possible role of antibody-related issues and interspecies differences in this matter. Fresh samples were obtained from human, rat, mouse and guinea pig cystectomies and processed for single/double immunohistochemistry/immunofluorescence. Specific antibodies against KIT, mast cell tryptase (MCT), anoctamin-1 (ANO1) and vimentin were used to characterize the cell types expressing KIT. Gut (jejunum) tissue was used as an external antibody control. Our results revealed KIT expression on mast cells but not on ICC in human, rat, mouse and guinea pig bladder. Parallel immunohistochemistry showed KIT expression on ICC in human, rat, mouse and guinea pig gut, which confirmed the selectivity of the KIT antibody clones. In conclusion, we have shown that KIT + cells in human, rat, mouse and guinea pig bladder are mast cells and not ICC. The present report is important as it opposes the idea that KIT + ICC are present in bladder. In this perspective, functional concepts of KIT + ICC being involved in sensory and/or motor aspects of bladder physiology should be revised. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Metabolic inactivation of 2-oxiranylmethyl 2-ethyl-2,5-dimethylhexanoate (C10GE) in skin, lung and liver of human, rat and mouse.

PubMed

Boogaard, P J; van Elburg, P A; de Kloe, K P; Watson, W P; van Sittert, N J

1999-10-01

The inactivation of 2-oxiranylmethyl 2-ethyl-2,5-dimethylhexanoate (C10GE), one of the most abundant isomers of the epoxy-resin Carduras E-10 glycidyl ester, was studied in subcellular fractions of human, C3H mouse and F344 rat liver, lung and skin. C10GE is chemically very stable and resistant to aqueous hydrolysis, but it was rapidly metabolized in both cytosolic and microsomal fractions of all organs by epoxide hydrolase (EH)-catalysed hydrolysis of the epoxide moiety as well as carboxylesterase (CE)-catalysed hydrolysis of the ester bond. In cytosol the epoxide group was also efficiently conjugated with glutathione, catalysed by glutathione S-transferase (GST), but this conjugation was much less important than hydrolysis in human as well as rodent samples. Although CE-catalysed hydrolysis of C10GE would theoretically give rise to the formation of glycidol, a directly acting mutagen, it is highly unlikely that any significant level of glycidol would occur in vivo since reported rates of inactivation of glycidol exceed the total rate of hydrolysis of C10GE. The overall rates of inactivation in vitro decreased in the following order: mouse > rat > human. Scaling of the data in vitro to clearances in vivo suggests that the detoxifying capacity in the rodents is similar and about an order of magnitude greater than in human. Nevertheless, the rate of inactivation is 2-3 orders of magnitude greater than for simple epoxides such as butadiene monoxide and about one order of magnitude higher than for the diglycidyl ether of bisphenol A (BADGE). The transdermal penetration and metabolism of [14C]-C10GE was studied in fresh full-thickness mouse, and dermatomized human and rat skin. Of the total radioactivity applied on the skin, only 0.24+/-0.06 (SD), 1.8+/-0.2 and 6.8+/-0.6% penetrated through human, mouse and rat skin respectively. The corresponding apparent skin permeability constants were 0.81, 6.42 and 26.4 x 10(-6) cm/h. During transdermal penetration, [14C]-C10GE was extensively hydrolysed to the corresponding diol and the free acid. Only 0.01, 0.11 and 0.21]% of the applied dose was absorbed unchanged through the human, mouse and rat skin respectively.

Rat and mouse CD94 associate directly with the activating transmembrane adaptor proteins DAP12 and DAP10 and activate NK cell cytotoxicity.

PubMed

Saether, Per C; Hoelsbrekken, Sigurd E; Fossum, Sigbjørn; Dissen, Erik

2011-12-15

Signaling by the CD94/NKG2 heterodimeric NK cell receptor family has been well characterized in the human but has remained unclear in the mouse and rat. In the human, the activating receptor CD94/NKG2C associates with DAP12 by an ionic bond between oppositely charged residues within the transmembrane regions of NKG2C and DAP12. The lysine residue responsible for DAP12 association is absent in rat and mouse NKG2C and -E, raising questions about signaling mechanisms in these species. As a possible substitute, rat and mouse NKG2C and -E contain an arginine residue in the transition between the transmembrane and stalk regions. In this article, we demonstrate that, similar to their human orthologs, NKG2A inhibits, whereas NKG2C activates, rat NK cells. Redirected lysis assays using NK cells transfected with a mutated NKG2C construct indicated that the activating function of CD94/NKG2C did not depend on the transmembrane/stalk region arginine residue. Flow cytometry and biochemical analysis demonstrated that both DAP12 and DAP10 can associate with rat CD94/NKG2C. Surprisingly, DAP12 and DAP10 did not associate with NKG2C but instead with CD94. These associations depended on a transmembrane lysine residue in CD94 that is unique to rodents. Thus, in the mouse and rat, the ability to bind activating adaptor proteins has been transferred from NKG2C/E to the CD94 chain as a result of mutation events in both chains. Remarkable from a phylogenetic perspective, this sheds new light on the evolution and function of the CD94/NKG2 receptor family.

Bioinformatics approaches for cross-species liver cancer analysis based on microarray gene expression profiling

PubMed Central

Fang, H; Tong, W; Perkins, R; Shi, L; Hong, H; Cao, X; Xie, Q; Yim, SH; Ward, JM; Pitot, HC; Dragan, YP

2005-01-01

Background The completion of the sequencing of human, mouse and rat genomes and knowledge of cross-species gene homologies enables studies of differential gene expression in animal models. These types of studies have the potential to greatly enhance our understanding of diseases such as liver cancer in humans. Genes co-expressed across multiple species are most likely to have conserved functions. We have used various bioinformatics approaches to examine microarray expression profiles from liver neoplasms that arise in albumin-SV40 transgenic rats to elucidate genes, chromosome aberrations and pathways that might be associated with human liver cancer. Results In this study, we first identified 2223 differentially expressed genes by comparing gene expression profiles for two control, two adenoma and two carcinoma samples using an F-test. These genes were subsequently mapped to the rat chromosomes using a novel visualization tool, the Chromosome Plot. Using the same plot, we further mapped the significant genes to orthologous chromosomal locations in human and mouse. Many genes expressed in rat 1q that are amplified in rat liver cancer map to the human chromosomes 10, 11 and 19 and to the mouse chromosomes 7, 17 and 19, which have been implicated in studies of human and mouse liver cancer. Using Comparative Genomics Microarray Analysis (CGMA), we identified regions of potential aberrations in human. Lastly, a pathway analysis was conducted to predict altered human pathways based on statistical analysis and extrapolation from the rat data. All of the identified pathways have been known to be important in the etiology of human liver cancer, including cell cycle control, cell growth and differentiation, apoptosis, transcriptional regulation, and protein metabolism. Conclusion The study demonstrates that the hepatic gene expression profiles from the albumin-SV40 transgenic rat model revealed genes, pathways and chromosome alterations consistent with experimental and clinical research in human liver cancer. The bioinformatics tools presented in this paper are essential for cross species extrapolation and mapping of microarray data, its analysis and interpretation. PMID:16026603

Aromatic amine metabolism: immunochemical relationships of N-acetyltransferase and N,O-acyltransferase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Land, S.; Allaben, W.T.; King, C.M.

1986-05-01

Mutagenic and carcinogenic aromatic amines are acetylated in most organisms. Acetyl CoA and arylhydroxamic acids can serve as acetyl donors for N-Acetylation of amines to yield stable amides, or by O-acetylation of hydroxylamine derivatives to produce reactive metabolites that can react covalently with nucleic acid. Polyclonal antibodies against rat arylhydroxamic acid, N,O-acyltransferase (AHAT) have been compared for their abilities to react with this enzyme and the acetyl CoA-dependent N-acetyltransferase (NAT) of the rat, rabbit, hamster, mouse and human. Liver cytosols were treated with increasing quantities of antibodies from immune or control rabbits. Immune complexes were removed by treatment with proteinmore » A-Sepharose before assay of nucleic acid adduct formation by AHAT activation of N-hydroxy-2-acetylaminofluorene and the acetylation of 2-aminofluorene by NAT. Both rat activities, the AHAT of the hamster and the NAT of the mouse and human were removed by this treatment. No decrease in NAT activity of hamster, or of either rabbit cytosol activity was observed. Neither mouse nor human liver has appreciable AHAT activity. These data support the idea that AHAT and NAT of rat, AHAT of hamster and NAT of mouse and human liver are immunochemically related, but that NAT of the hamster is an immunochemically distinct peptide.« less

Species comparison of hepatic and pulmonary metabolism of benzene.

PubMed

Powley, M W; Carlson, G P

1999-12-06

Benzene is an occupational hazard and environmental toxicant found in cigarette smoke, gasoline, and the chemical industry. The major health concern associated with benzene exposure is leukemia. Studies using microsomal preparations from human, mouse, rabbit, and rat to determine species differences in the metabolism of benzene to phenol, hydroquinone and catechol, indicate that the rat is most similar, both quantitatively and qualitatively, to the human in pulmonary microsomal metabolism of benzene. With hepatic microsomes, rat is most similar to human in metabolite formation at the two lower concentrations examined (24 and 200 microM), while at the two higher concentrations (700 and 1000 microM) mouse is most similar in phenol formation. In all species, the enzyme system responsible for benzene metabolism approached saturation in hepatic microsomes but not in pulmonary microsomes. In pulmonary microsomes from mouse, rat, and human, phenol appeared to competitively inhibit benzene metabolism resulting in a greater proportion of phenol being converted to hydroquinone when the benzene concentration increased. The opposite effect was seen in hepatic microsomes. These findings support the hypothesis that the lung plays an important role in benzene metabolism, and therefore, toxicity.

The Relationship between Muscle Fiber Type-Specific PGC-1α Content and Mitochondrial Content Varies between Rodent Models and Humans

PubMed Central

Gouspillou, Gilles; Sgarioto, Nicolas; Norris, Brandon; Barbat-Artigas, Sébastien; Aubertin-Leheudre, Mylène; Morais, Jose A.; Burelle, Yan; Taivassalo, Tanja; Hepple, Russell T.

2014-01-01

PGC-1α regulates critical processes in muscle physiology, including mitochondrial biogenesis, lipid metabolism and angiogenesis. Furthermore, PGC-1α was suggested as an important regulator of fiber type determination. However, whether a muscle fiber type-specific PGC-1α content exists, whether PGC-1α content relates to basal levels of mitochondrial content, and whether such relationships are preserved between humans and classically used rodent models are all questions that have been either poorly addressed or never investigated. To address these issues, we investigated the fiber type-specific content of PGC-1α and its relationship to basal mitochondrial content in mouse, rat and human muscles using in situ immunolabeling and histochemical methods on muscle serial cross-sections. Whereas type IIa fibers exhibited the highest PGC-1α in all three species, other fiber types displayed a hierarchy of type IIx>I>IIb in mouse, type I = IIx> IIb in rat, and type IIx>I in human. In terms of mitochondrial content, we observed a hierarchy of IIa>IIx>I>IIb in mouse, IIa >I>IIx> IIb in rat, and I>IIa> IIx in human skeletal muscle. We also found in rat skeletal muscle that type I fibers displayed the highest capillarization followed by type IIa >IIx>IIb. Finally, we found in human skeletal muscle that type I fibers display the highest lipid content, followed by type IIa>IIx. Altogether, our results reveal that (i) the fiber type-specific PGC-1α and mitochondrial contents were only matched in mouse, (ii) the patterns of PGC-1α and mitochondrial contents observed in mice and rats do not correspond to that seen in humans in several respects, and (iii) the classical phenotypes thought to be regulated by PGC-1α do not vary exclusively as a function of PGC-1α content in rat and human muscles. PMID:25121500

Comparative analysis of genome maintenance genes in naked mole rat, mouse, and human.

PubMed

MacRae, Sheila L; Zhang, Quanwei; Lemetre, Christophe; Seim, Inge; Calder, Robert B; Hoeijmakers, Jan; Suh, Yousin; Gladyshev, Vadim N; Seluanov, Andrei; Gorbunova, Vera; Vijg, Jan; Zhang, Zhengdong D

2015-04-01

Genome maintenance (GM) is an essential defense system against aging and cancer, as both are characterized by increased genome instability. Here, we compared the copy number variation and mutation rate of 518 GM-associated genes in the naked mole rat (NMR), mouse, and human genomes. GM genes appeared to be strongly conserved, with copy number variation in only four genes. Interestingly, we found NMR to have a higher copy number of CEBPG, a regulator of DNA repair, and TINF2, a protector of telomere integrity. NMR, as well as human, was also found to have a lower rate of germline nucleotide substitution than the mouse. Together, the data suggest that the long-lived NMR, as well as human, has more robust GM than mouse and identifies new targets for the analysis of the exceptional longevity of the NMR. © 2015 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Generation and Characterization of a New Monoclonal Antibody Against CXCL4.

PubMed

Gao, Jing; Wu, Mingyuan; Gao, Jin; Wang, Xia; Zhang, Yang; Zhu, Shunying; Yu, Yan; Han, Wei

2015-04-01

CXCL4 plays important roles in numerous disease processes, which makes the CXCL4 signaling pathway a potential therapeutic target. In this study, we aimed to develop a neutralizing antibody against both human and mouse CXCL4. Rats were immunized with recombinant human CXCL4 (rhCXCL4). Hybridoma clones were created by fusion of the immunized rat spleen cells with mouse myeloma SP2/0 cells and screened using recombinant mouse CXCL4 (rmCXCL4) and rhCXCL4. The CXCL4 monoclonal antibody (CXCL4 MAb) produced by the 16D6-3 hybridoma clone was sequenced and characterized by Western blot and Biacore assays. It recognized both human and mouse CXCL4 with high affinity and neutralized the effect of rhCXCL4 in vitro. Thus, the antibody may be used in the studies of CXCL4 in murine disease models and as a template in the antibody humanization for clinical developments.

Identification and characterization of metabolites of ASP015K, a novel oral Janus kinase inhibitor, in rats, chimeric mice with humanized liver, and humans.

PubMed

Nakada, Naoyuki; Oda, Kazuo

2015-01-01

1. Here, we elucidated the structure of metabolites of novel oral Janus kinase inhibitor ASP015K in rats and humans and evaluated the predictability of human metabolites using chimeric mice with humanized liver (PXB mice). 2. Rat biological samples collected after oral dosing of (14)C-labelled ASP015K were examined using a liquid chromatography-radiometric detector and mass spectrometer (LC-RAD/MS). The molecular weight of metabolites in human and the liver chimeric mouse biological samples collected after oral dosing of non-labelled ASP015K was also investigated via LC-MS. Metabolites were also isolated from rat bile samples and analyzed using nuclear magnetic resonance. 3. Metabolic pathways of ASP015K in rats and humans were found to be glucuronide conjugation, methyl conjugation, sulfate conjugation, glutathione conjugation, hydroxylation of the adamantane ring and N-oxidation of the 1H-pyrrolo[2,3-b]pyridine ring. The main metabolite of ASP015K in rats was the glucuronide conjugate, while the main metabolite in humans was the sulfate conjugate. Given that human metabolites were produced by human hepatocytes in chimeric mice with humanized liver, this human model mouse was believed to be useful in predicting the human metabolic profile of various drug candidates.

Identification of rat Rosa26 locus enables generation of knock-in rat lines ubiquitously expressing tdTomato.

PubMed

Kobayashi, Toshihiro; Kato-Itoh, Megumi; Yamaguchi, Tomoyuki; Tamura, Chihiro; Sanbo, Makoto; Hirabayashi, Masumi; Nakauchi, Hiromitsu

2012-11-01

Recent discovery of a method for derivation and culture of germline-competent rat pluripotent stem cells (PSCs) enables generation of transgenic rats or knock-out rats via genetic modification of such PSCs. This opens the way to use rats, as is routine in mice, for analyses of gene functions or physiological features. In mouse or human, one widely used technique to express a gene of interest stably and ubiquitously is to insert that gene into the Rosa26 locus via gene targeting of PSCs. Rosa26 knock-in mice conditionally expressing a reporter or a toxin gene have contributed to tracing or ablation of specific cell lineages. We successfully identified a rat orthologue of the mouse Rosa26 locus. Insertion of tdTomato, a variant of red fluorescent protein, into the Rosa26 locus of PSCs of various rat strains allows ubiquitous expression of tdTomato. Through germline transmission of one Rosa26-tdTomato knock-in embryonic stem cell line, we also obtained tdTomato knock-in rats. These expressed tdTomato ubiquitously throughout their bodies, which indicates that the rat Rosa26 locus conserves functions of its orthologues in mouse and human. The new tools described here (targeting vectors, knock-in PSCs, and rats) should be useful for a variety of research using rats.

Saccharin and Cyclamate Inhibit Binding of Epidermal Growth Factor

NASA Astrophysics Data System (ADS)

Lee, L. S.

1981-02-01

The binding of 125I-labeled mouse epidermal growth factor (EGF) to 18 cell lines, including HeLa (human carcinoma), MDCK (dog kidney cells), HTC (rat hepatoma), K22 (rat liver), HF (human foreskin), GM17 (human skin fibroblasts), XP (human xeroderma pigmentosum fibroblasts), and 3T3-L1 (mouse fibroblasts), was inhibited by saccharin and cyclamate. The human cells were more sensitive to inhibition by these sweeteners than mouse or rat cells. EGF at doses far above the physiological levels reversed the inhibition in rodent cells but not in HeLa cells. In HeLa cells, the doses of saccharin and cyclamate needed for 50% inhibition were 3.5 and 9.3 mg/ml, respectively. Glucose, 2-deoxyglucose, sucrose, and xylitol did not inhibit EGF binding. Previous studies have shown that phorbol esters, strongly potent tumor promoters, also inhibit EGF binding to tissue culture cells. To explain the EGF binding inhibition by such greatly dissimilar molecules as phorbol esters, saccharin, and cyclamate, it is suggested that they operate through the activation of a hormone response control unit.

Neonicotinoid formaldehyde generators: possible mechanism of mouse-specific hepatotoxicity/hepatocarcinogenicity of thiamethoxam.

PubMed

Swenson, Tami L; Casida, John E

2013-02-04

Thiamethoxam (TMX), an important insecticide, is hepatotoxic and hepatocarcinogenic in mice but not rats. Studies of Syngenta Central Toxicology Laboratory on species specificity in metabolism established that TMX is a much better substrate for mouse liver microsomal CYPs than the corresponding rat or human enzymes in forming desmethyl-TMX (dm-TMX), which is also hepatotoxic, and clothianidin (CLO), which is not hepatotoxic or hepatocarcinogenic. They proposed that TMX hepatotoxicity/hepatocarcinogencity is due to dm-TMX and a further metabolite desmethyl-CLO (dm-CLO) (structurally analogous to a standard inducible nitric oxide synthase inhibitor) acting synergistically. The present study considers formation of formaldehyde (HCHO) and N-methylol intermediates as an alternative mechanism of TMX hepatotoxicity/hepatocarcinogenicity. Comparison of neonicotinoid metabolism by mouse, rat and human microsomes with NADPH showed two important points. First, TMX and dm-TMX yield more HCHO than any other commercial neonicotinoid. Second, mouse microsomes give much higher conversion than rat or human microsomes. These observations provide an alternative hypothesis of HCHO and N-methylol intermediates from CYP-mediated oxidative oxadiazinane ring cleavage as the bioactivated hepatotoxicants. However, the proposed mono-N-methylol CYP metabolites are not observed, possibly further reacting in situ. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Fenoterol functionally activates the β₃-adrenoceptor in human urinary bladder, comparison with rat and mouse: implications for drug discovery.

PubMed

Palea, Stefano; Rekik, Moèz; Rouget, Céline; Camparo, Philippe; Botto, Henri; Rischmann, Pascal; Lluel, Philippe; Westfall, Timothy D

2012-09-05

Fenoterol has been reported to be a potent and selective β(2)-adrenoceptor agonist and is currently used clinically to treat asthma. Electrical field stimulation (EFS) of isolated urinary bladder mimics the voiding contraction by stimulating parasympathetic nerves, resulting in neurogenic contractions. To determine if stimulation of β(2)-adrenoceptors can inhibit this response, fenoterol was tested against EFS-induced contractions in human isolated urinary bladder and compared with mouse and rat. Bladder strips were mounted in organ baths and reproducible contractions induced by EFS. Fenoterol was added cumulatively in the presence of the β(2)-adrenoceptor antagonist ICI118551 or the β(3)-adrenoceptor antagonist L-748337. Fenoterol inhibited neurogenic contractions in all three species in a concentration-dependent manner with pEC(50) values of 6.66 ± 0.11, 6.86 ± 0.06 and 5.71 ± 0.1 in human, mouse and rat respectively. In human bladder strips ICI118551 (100 nM) did not affect responses to fenoterol, while L-748337 (0.3-3 μM) produced rightward shifts of the concentration-response curves with a pA(2) value of 8.10. In mouse bladder strips ICI118551 (30 nM) blocked the inhibitory effect of fenoterol (pA(2)=8.80), while L-748337 (10 μM) inhibited the response with a pA(2) of 5.79. In rat bladder ICI118551 (30 nM) was without effect, while L-748,337 (10 μM) inhibited the response to fenoterol with a pA(2) of 5.40. From these results it is clear that fenoterol potently activates β(3)-adrenoceptors in human isolated urinary bladder to inhibit EFS-induced contractions. Fenoterol also activates β(3)-adrenoceptors in rat, but β(2)-adrenoceptors in mouse bladder to inhibit EFS-induced contractions. Copyright © 2012 Elsevier B.V. All rights reserved.

Precision-cut rat, mouse, and human intestinal slices as novel models for the early-onset of intestinal fibrosis.

PubMed

Pham, Bao Tung; van Haaften, Wouter Tobias; Oosterhuis, Dorenda; Nieken, Judith; de Graaf, Inge Anne Maria; Olinga, Peter

2015-04-01

Intestinal fibrosis (IF) is a major complication of inflammatory bowel disease. IF research is limited by the lack of relevant in vitro and in vivo models. We evaluated precision-cut intestinal slices (PCIS) prepared from human, rat, and mouse intestine as ex vivo models mimicking the early-onset of (human) IF. Precision-cut intestinal slices prepared from human (h), rat (r), and mouse (m) jejunum, were incubated up to 72 h, the viability of PCIS was assessed by ATP content and morphology, and the gene expression of several fibrosis markers was determined. The viability of rPCIS decreased after 24 h of incubation, whereas mPCIS and hPCIS were viable up to 72 h of culturing. Furthermore, during this period, gene expression of heat shock protein 47 and plasminogen activator inhibitor 1 increased in all PCIS in addition to augmented expression of synaptophysin in hPCIS, fibronectin (Fn2) and TGF-β1 in rPCIS, and Fn2 and connective tissue growth factor (Ctgf) in mPCIS. Addition of TGF-β1 to rPCIS or mPCIS induced the gene expression of the fibrosis markers Pro-collagen1a1, Fn2, and Ctgf in both species. However, none of the fibrosis markers was further elevated in hPCIS. We successfully developed a novel ex vivo model that can mimic the early-onset of fibrosis in the intestine using human, rat, and mouse PCIS. Furthermore, in rat and mouse PCIS, TGF-β1 was able to even further increase the gene expression of fibrosis markers. This indicates that PCIS can be used as a model for the early-onset of IF. © 2015 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

Determination of species-difference in microsomal metabolism of amitriptyline using a predictive MRM-IDA-EPI method.

PubMed

Lee, Ji-Yoon; Lee, Sang Yoon; Lee, KiHo; Oh, Soo Jin; Kim, Sang Kyum

2015-03-05

We investigated to compare species differences in amitriptyline (AMI) metabolism among mouse, rat, dog, and human liver microsomes. We developed a method for simultaneous determination of metabolic stability and metabolite profiling using predictive multiple reaction monitoring information-dependent acquisition-enhanced product ion (MRM-IDA-EPI) scanning. In the cofactor-dependent microsomal metabolism study, AMI was metabolized more rapidly in rat and human liver microsomes incubated with NADPH than UDPGA. AMI incubated with NADPH+UDPGA in rat, dog, or mouse liver microsomes disappeared rapidly with a half-life of 3.5, 8.4, or 9.2 min, respectively, but slowly in human liver microsomes with a half-life of 96 min. In total, 9, 10, 11, and 6 putative metabolites of AMI were detected in mouse, rat, dog, and human liver microsomes, respectively, based on mass spectrometric analyses. Kinetic analysis of metabolites in liver microsomes from each species over 120 min showed common metabolic routes of AMI, such as N-demethylation, hydroxylation, and glucuronidation, and subtle interspecies differences in AMI metabolism. The main metabolic routes in mouse, rat, dog, and human liver microsomes were hydroxylation followed by glucuronide conjugation, methyl hydroxylation, and N-demethylation, respectively. The MRM-IDA-EPI method can provide quantitative and qualitative information about metabolic stability and metabolite profiling simultaneously. Moreover, time course analysis of metabolites can not only eliminate false identification of metabolites, but also provide a rationale for proposed metabolic pathways. The MRM-IDA-EPI method combined with time course analysis of metabolites is useful for investigating drug metabolism at the early drug discovery stage. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Generation and Characterization of a New Monoclonal Antibody Against CXCL4

PubMed Central

Gao, Jing; Wu, Mingyuan; Gao, Jin; Wang, Xia; Zhang, Yang; Zhu, Shunying; Yu, Yan

2015-01-01

CXCL4 plays important roles in numerous disease processes, which makes the CXCL4 signaling pathway a potential therapeutic target. In this study, we aimed to develop a neutralizing antibody against both human and mouse CXCL4. Rats were immunized with recombinant human CXCL4 (rhCXCL4). Hybridoma clones were created by fusion of the immunized rat spleen cells with mouse myeloma SP2/0 cells and screened using recombinant mouse CXCL4 (rmCXCL4) and rhCXCL4. The CXCL4 monoclonal antibody (CXCL4 MAb) produced by the 16D6-3 hybridoma clone was sequenced and characterized by Western blot and Biacore assays. It recognized both human and mouse CXCL4 with high affinity and neutralized the effect of rhCXCL4 in vitro. Thus, the antibody may be used in the studies of CXCL4 in murine disease models and as a template in the antibody humanization for clinical developments. PMID:25897609

Pharmacological profiling of the TRPV3 channel in recombinant and native assays.

PubMed

Grubisha, Olivera; Mogg, Adrian J; Sorge, Jessica L; Ball, Laura-Jayne; Sanger, Helen; Ruble, Cara L A; Folly, Elizabeth A; Ursu, Daniel; Broad, Lisa M

2014-05-01

Transient receptor potential vanilloid subtype 3 (TRPV3) is implicated in nociception and certain skin conditions. As such, it is an attractive target for pharmaceutical research. Understanding of endogenous TRPV3 function and pharmacology remains elusive as selective compounds and native preparations utilizing higher throughput methodologies are lacking. In this study, we developed medium-throughput recombinant and native cellular assays to assess the detailed pharmacological profile of human, rat and mouse TRPV3 channels. Medium-throughput cellular assays were developed using a Ca(2+) -sensitive dye and a fluorescent imaging plate reader. Human and rat TRPV3 pharmacology was examined in recombinant cell lines, while the mouse 308 keratinocyte cell line was used to assess endogenous TRPV3 activity. A recombinant rat TRPV3 cellular assay was successfully developed after solving a discrepancy in the published rat TRPV3 protein sequence. A medium-throughput, native, mouse TRPV3 keratinocyte assay was also developed and confirmed using genetic approaches. Whereas the recombinant human and rat TRPV3 assays exhibited similar agonist and antagonist profiles, the native mouse assay showed important differences, namely, TRPV3 activity was detected only in the presence of potentiator or during agonist synergy. Furthermore, the native assay was more sensitive to block by some antagonists. Our findings demonstrate similarities but also notable differences in TRPV3 pharmacology between recombinant and native systems. These findings offer insights into TRPV3 function and these assays should aid further research towards developing TRPV3 therapies. © 2013 The British Pharmacological Society.

Zygote arrest 1 (Zar1) is an evolutionarily conserved gene expressed in vertebrate ovaries.

PubMed

Wu, Xuemei; Wang, Pei; Brown, Christopher A; Zilinski, Carolyn A; Matzuk, Martin M

2003-09-01

Zygote arrest 1 (ZAR1) is an ovary-specific maternal factor that plays essential roles during the oocyte-to-embryo transition. In mice, the Zar1 mRNA is detected as a 1.4-kilobase (kb) transcript that is synthesized exclusively in growing oocytes. To further understand the functions of ZAR1, we have cloned the orthologous Zar1 cDNA and/or genes for mouse, rat, human, frog, zebrafish, and pufferfish. The entire mouse Zar1 gene and a related pseudogene span approximately 4.0 kb, contain four exons, and map to adjacent loci on mouse chromosome 5. The human ZAR1 orthologous gene similarly consists of four exons and resides on human chromosome 4p12, which is syntenic with the mouse Zar1 chromosomal locus. Rat (Rattus norvegicus) and pufferfish (Fugu rubripes) Zar1 genes were recognized by database mining and deduced protein alignment analysis. The rat Zar1 gene also maps to a region that is syntenic with the mouse Zar1 gene locus on rat chromosome 14. Frog (Xenopus laevis) and zebrafish (Danio rerio) Zar1 orthologs were cloned by reverse transcription-polymerase chain reaction and rapid amplification of cDNA ends analysis of ovarian mRNA. Unlike mouse and human, the frog Zar1 is detected in multiple tissues, including lung, muscle, and ovary. The Zar1 mRNA appears in the cytoplasm of oocytes and persists until the tailbud stage during frog embryogenesis. Mouse, rat, human, frog, zebrafish, and pufferfish Zar1 genes encode proteins of 361, 361, 424, 295, 329, and 320 amino acids, respectively, and share 50.8%-88.1% amino acid identity. Regions of the N-termini of these ZAR1 orthologs show high sequence identity among these various proteins. However, the C-terminal 103 amino acids of these proteins, encoded by exons 2-4, contain an atypical eight-cysteine Plant Homeo Domain motif and are highly conserved, sharing 80.6%-98.1% identity among these species. These findings suggest that the carboxyl-termini of these ZAR1 proteins contain an important functional domain that is conserved through vertebrate evolution and that may be necessary for normal female reproduction in the transition from oocyte to embryonic life.

Analysis of Species-Selectivity of Human, Mouse and Rat Cytochrome P450 1A and 2B Subfamily Enzymes using Molecular Modeling, Docking and Dynamics Simulations.

PubMed

Karthikeyan, Bagavathy Shanmugam; Suvaithenamudhan, Suvaiyarasan; Akbarsha, Mohammad Abdulkader; Parthasarathy, Subbiah

2018-06-01

Cytochrome P450 (CYP) 1A and 2B subfamily enzymes are important drug metabolizing enzymes, and are highly conserved across species in terms of sequence homology. However, there are major to minor structural and macromolecular differences which provide for species-selectivity and substrate-selectivity. Therefore, species-selectivity of CYP1A and CYP2B subfamily proteins across human, mouse and rat was analyzed using molecular modeling, docking and dynamics simulations when the chiral molecules quinine and quinidine were used as ligands. The three-dimensional structures of 17 proteins belonging to CYP1A and CYP2B subfamilies of mouse and rat were predicted by adopting homology modeling using the available structures of human CYP1A and CYP2B proteins as templates. Molecular docking and dynamics simulations of quinine and quinidine with CYP1A subfamily proteins revealed the existence of species-selectivity across the three species. On the other hand, in the case of CYP2B subfamily proteins, no role for chirality of quinine and quinidine in forming complexes with CYP2B subfamily proteins of the three species was indicated. Our findings reveal the roles of active site amino acid residues of CYP1A and CYP2B subfamily proteins and provide insights into species-selectivity of these enzymes across human, mouse, and rat.

Human Fetal Testis Xenografts Are Resistant to Phthalate-Induced Endocrine Disruption

PubMed Central

Heger, Nicholas E; Hall, Susan J; Sandrof, Moses A; McDonnell, Elizabeth V; Hensley, Janan B; McDowell, Erin N; Martin, Kayla A; Gaido, Kevin W; Johnson, Kamin J

2012-01-01

Background: In utero exposure to endocrine-disrupting chemicals may contribute to testicular dysgenesis syndrome (TDS), a proposed constellation of increasingly common male reproductive tract abnormalities (including hypospadias, cryptorchidism, hypospermatogenesis, and testicular cancer). Male rats exposed in utero to certain phthalate plasticizers exhibit multinucleated germ cell (MNG) induction and suppressed steroidogenic gene expression and testosterone production in the fetal testis, causing TDS-consistent effects of hypospadias and cryptorchidism. Mice exposed to phthalates in utero exhibit MNG induction only. This disparity in response demonstrates a species-specific sensitivity to phthalate-induced suppression of fetal Leydig cell steroidogenesis. Importantly, ex vivo phthalate exposure of the fetal testis does not recapitulate the species-specific endocrine disruption, demonstrating the need for a new bioassay to assess the human response to phthalates. Objectives: In this study, we aimed to develop and validate a rat and mouse testis xenograft bioassay of phthalate exposure and examine the human fetal testis response. Methods: Fetal rat, mouse, and human testes were xenografted into immunodeficient rodent hosts, and hosts were gavaged with a range of phthalate doses over multiple days. Xenografts were harvested and assessed for histopathology and steroidogenic end points. Results: Consistent with the in utero response, phthalate exposure induced MNG formation in rat and mouse xenografts, but only rats exhibited suppressed steroidogenesis. Across a range of doses, human fetal testis xenografts exhibited MNG induction but were resistant to suppression of steroidogenic gene expression. Conclusions: Phthalate exposure of grafted human fetal testis altered fetal germ cells but did not reduce expression of genes that regulate fetal testosterone biosynthesis. PMID:22511013

Use of AN Eosinophil Specific Monoclonal Antibody in Assessing Eosinophil Function.

NASA Astrophysics Data System (ADS)

Minkoff, Marjorie Sue

A monoclonal antibody to an eosinophil specific determinant is very important in assessing eosinophil function during helminthic infection. Eosinophils induced by Schistosoma mansoni infection in BALB/c mice were used to induce C57B1/6 immunocytes for production of hybridomas secreting eosinophil monoclonal antibodies. These antibodies were shown to react with an eosinophil surface epitope but not with neutrophils or macrophages as determined by ELISA, immunodiffusion, immunofluorescence, and immunoblot assay. Affinity chromatography with eosinophil chemotactic factor-sepharose consistently selected out a { rm M_ R} 67,000 protein from solubilized eosinophil membrane antigens but not from neutrophil and macrophage antigens. In vitro studies showed that the eosinophil-specific monoclonal antibodies abrogated antibody-dependent eosinophil -mediated killing of S. mansoni schistosomula using mouse, rat or human eosinophils. Neutrophil and macrophage killing activities were unaffected. The monoclonal antibodies effected complement-dependent lysis of mouse and rat eosinophils but not of human eosinophils. ECF-treated eosinophils showed enhanced killing of schistosomula which was blocked by the monoclonal antibody. Murine and human eosinophils preincubated with monoclonal antibody exhibited decreased chemotaxis to ECF at optimal chemotactic concentrations. The monoclonal antibody also blocked eosinophil binding to ECF- sepharose beads. In vivo induction of peripheral blood eosinophilia by injection of S. mansoni eggs was suppressed by injections of monoclonal antibodies 2CD13 and 2QD45 in mouse and rat experimental models. Eosinophilia induced by keyhole limpet hemocyanin- cyclophosphamide treatment was also suppressed by monoclonal antibody in both murine and rat systems. Pulmonary granulomas in mice given egg injection and monoclonal antibody were smaller and contained fewer eosinophils than those granulomas from mice given eggs only. In immuno-biochemical studies, the monoclonal antibody 2QD45 specifically immunoprecipitated the {rm M_ R} 67,000 ECF-binding protein from ^{125}{rm I}-labeled mouse, rat, and human eosinophils as assessed by SDS-PAGE and autoradiography. Two-dimensional gel electrophoresis showed that this ECF-binding protein has a lower PI point than either mouse or bovine albumin.

A Rat Model for Human Ductal Carcinoma In Situ

DTIC Science & Technology

2006-09-01

complexes in early stages of mouse mammary preneoplasia. Cell Growth Differ 12:285-95. 14. Gudmundur Thordarson , Adrian V. Lee, Meghan McCarty...S.M., Thordarson , G., Collins, G., Talamantes, F., and Nandi, S. (1995) In vivo growth stimulation of collagen gel embedded normal human and mouse

Exploring human disease using the Rat Genome Database.

PubMed

Shimoyama, Mary; Laulederkind, Stanley J F; De Pons, Jeff; Nigam, Rajni; Smith, Jennifer R; Tutaj, Marek; Petri, Victoria; Hayman, G Thomas; Wang, Shur-Jen; Ghiasvand, Omid; Thota, Jyothi; Dwinell, Melinda R

2016-10-01

Rattus norvegicus, the laboratory rat, has been a crucial model for studies of the environmental and genetic factors associated with human diseases for over 150 years. It is the primary model organism for toxicology and pharmacology studies, and has features that make it the model of choice in many complex-disease studies. Since 1999, the Rat Genome Database (RGD; http://rgd.mcw.edu) has been the premier resource for genomic, genetic, phenotype and strain data for the laboratory rat. The primary role of RGD is to curate rat data and validate orthologous relationships with human and mouse genes, and make these data available for incorporation into other major databases such as NCBI, Ensembl and UniProt. RGD also provides official nomenclature for rat genes, quantitative trait loci, strains and genetic markers, as well as unique identifiers. The RGD team adds enormous value to these basic data elements through functional and disease annotations, the analysis and visual presentation of pathways, and the integration of phenotype measurement data for strains used as disease models. Because much of the rat research community focuses on understanding human diseases, RGD provides a number of datasets and software tools that allow users to easily explore and make disease-related connections among these datasets. RGD also provides comprehensive human and mouse data for comparative purposes, illustrating the value of the rat in translational research. This article introduces RGD and its suite of tools and datasets to researchers - within and beyond the rat community - who are particularly interested in leveraging rat-based insights to understand human diseases. © 2016. Published by The Company of Biologists Ltd.

Differences in betaine-homocysteine methyltransferase expression, ER stress response and liver injury between alcohol-fed mice and rats

PubMed Central

Shinohara, Masao; Ji, Cheng; Kaplowitz, Neil

2009-01-01

Chronic ethanol infusion resulted in greater serum ALT elevation, lipid accumulation, necroinflammation, and focal hepatic cell death in mice than rats. Mice exhibited a remarkable hyperhomocysteinemia but no increase was seen in rats. Similarly, a high methionine low folate diet (HMLF) induced less steatosis, serum ALT increase, and hyperhomocysteinemia in rats than in mice. Western blot analysis of betaine homocysteine methyltransferase (BHMT) expression showed that rats fed either ethanol or HMLF had significantly increased BHMT expression which did not occur in mice. Nuclear NFκB p65 was increased in mouse in response to alcohol feeding. The human BHMT promoter was repressed by homocysteine in mouse hepatocytes but not rat hepatocytes. BHMT induction was faster and greater in primary rat hepatocytes than mouse hepatocytes in response to exogenous homocysteine exposure. Mice fed ethanol i.g. exhibited an increase in GRP78 and IRE1 which was not seen in the rat and SREBP-1 was increased to a greater extent in mice than rats. Thus, rats are more resistant to ethanol induced steatosis, ER stress and hyperhomocysteinemia and this correlates with induction of BHMT in rats. These findings support the hypothesis that a critical factor in the pathogenesis of alcoholic liver injury is the enhanced ability of rat or impaired ability of mouse to up-regulate BHMT which prevents hyperhomocysteinemia, ER stress and liver injury. PMID:20069651

Activation properties of heterologously expressed mammalian TRPV2: evidence for species dependence.

PubMed

Neeper, Michael P; Liu, Yi; Hutchinson, Tasha L; Wang, Yan; Flores, Christopher M; Qin, Ning

2007-05-25

TRPV2 has been proposed as a potential pain target, in part due to its relatedness to the nociceptor TRPV1 and to its reported activation by noxious high temperatures (>52 degrees C). However, TRPV2 responses to heat as well as to the nonselective agonist 2-aminoethoxydiphenyl borate (2-APB) have not been universally reproduced in other laboratories, leading to debate about the activation properties of this channel. Here, we report the expression of rat, mouse, and human TRPV2 in HEK293 cells and the differential properties of their responses to heat and 2-APB. Expression of mouse or rat TRPV2 in HEK293 cells resulted in robust channel activation when induced by either temperature (>53 degrees C) or 2-APB. By contrast, expression of human TRPV2 did not lead to detectable activation by either of these stimuli. Human TRPV2 protein was expressed at levels comparable with those of rat TRPV2, exhibited similar surface localization and responded to a novelly identified TRPV2 agonist, Delta(9)-tetrahydrocannabinol, indicating that human TRPV2 is functionally expressed on the cell surface. Studies using deletion mutants and chimeras between rat and human TRPV2 indicated that both amino- and carboxyl-cytoplasmic termini of rat TRPV2 are important for responses to heat and 2-APB but can be supplied in trans to form an active channel. The present study not only confirms and extends previous reports demonstrating that rat and mouse TRPV2 respond to 2-APB and noxious heat but also indicates that further investigation will be required to elucidate TRPV2 activation and regulatory mechanisms.

Isolation, expression, and chromosomal localization of the human mitochondrial capsule selenoprotein gene (MCSP)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aho, Hanne; Schwemmer, M.; Tessmann, D.

1996-03-01

The mitochondrial capsule selenoprotein (MCS) (HGMW-approved symbol MCSP) is one of three proteins that are important for the maintenance and stabilization of the crescent structure of the sperm mitochondria. We describe here the isolation of a cDNA, the exon-intron organization, the expression, and the chromosomal localization of the human MCS gene. Nucleotide sequence analysis of the human and mouse MCS cDNAs reveals that the 5{prime}- and 3{prime}-untranslated sequences are more conserved (71%) than the coding sequences (59%). The open reading frame encodes a 116-amino-acid protein and lacks the UGA codons, which have been reported to encode the selenocysteines in themore » N-terminal of the deduced mouse protein. The deduced human protein shows a low degree of amino acid sequence identity to the mouse protein. The deduced human protein shows a low degree of amino acid sequence identity to the mouse protein (39%). The most striking homology lies in the dicysteine motifs. Northern and Southern zooblot analyses reveal that the MCS gene in human, baboon, and bovine is more conserved than its counterparts in mouse and rat. The single intron in the human MCS gene is approximately 6 kb and interrupts the 5{prime}-untranslated region at a position equivalent to that in the mouse and rat genes. Northern blot and in situ hybridization experiments demonstrate that the expression of the human MCS gene is restricted to haploid spermatids. The human gene was assigned to q21 of chromosome 1. 30 refs., 9 figs.« less

Cloning and characterization of the rat free fatty acid receptor GPR120: in vivo effect of the natural ligand on GLP-1 secretion and proliferation of pancreatic beta cells.

PubMed

Tanaka, Toshiki; Yano, Takeaki; Adachi, Tetsuya; Koshimizu, Taka-aki; Hirasawa, Akira; Tsujimoto, Gozoh

2008-06-01

We have recently found that GPR120, which is abundantly expressed in intestine, functions as a receptor for unsaturated long-chain free fatty acids (FFAs) and that GPR120 stimulation promotes the secretion of glucagons-like peptide-1 (GLP-1) in the mouse (Hirasawa et al., Nat Med 11:90-94, 2005). In this study, we cloned and characterized rat GPR120 (rGPR120), and then we examined the in vivo effects of acute and long-term administration of the natural ligand alpha-linolenic acid (alpha-LA). The cloned rat GPR120 complimentary DNA had a seven transmembrane structure, and a homology comparison of human, mouse, and rat GPR120 revealed that the rat GPR120 (rGPR120) shares 85 and 98% sequence identity with the human and mouse GPR120 proteins, respectively. The tissue distribution and ligand properties of rGPR120 were similar to those of mouse GPR120. In addition, alpha-LA provoked a transient increase in [Ca2+]i levels in HEK293 cells expressing rGPR120. Furthermore, administration of alpha-LA to the rat increased plasma GLP-1 levels, and long-term administration of alpha-LA led to proliferation of pancreatic beta cells, probably because of the enhanced GLP-1 secretion. These results show that rat GPR120 is a G-protein-coupled receptor whose ligand is a free fatty acid, and it may play an important role in the FFA-associated physiological responses.

Characterization of selective Calcium-Release Activated Calcium channel blockers in mast cells and T-cells from human, rat, mouse and guinea-pig preparations.

PubMed

Rice, Louise V; Bax, Heather J; Russell, Linda J; Barrett, Victoria J; Walton, Sarah E; Deakin, Angela M; Thomson, Sally A; Lucas, Fiona; Solari, Roberto; House, David; Begg, Malcolm

2013-03-15

Loss of function mutations in the two key proteins which constitute Calcium-Release Activated Calcium (CRAC) channels demonstrate the critical role of this ion channel in immune cell function. The aim of this study was to demonstrate that inhibition of immune cell activation could be achieved with highly selective inhibitors of CRAC channels in vitro using cell preparations from human, rat, mouse and guinea-pig. Two selective small molecule blockers of CRAC channels; GSK-5498A and GSK-7975A were tested to demonstrate their ability to inhibit mediator release from mast cells, and pro-inflammatory cytokine release from T-cells in a variety of species. Both GSK-5498A and GSK-7975A completely inhibited calcium influx through CRAC channels. This led to inhibition of the release of mast cell mediators and T-cell cytokines from multiple human and rat preparations. Mast cells from guinea-pig and mouse preparations were not inhibited by GSK-5498A or GSK-7975A; however cytokine release was fully blocked from T-cells in a mouse preparation. GSK-5498A and GSK-7975A confirm the critical role of CRAC channels in human mast cell and T-cell function, and that inhibition can be achieved in vitro. The rat displays a similar pharmacology to human, promoting this species for future in vivo research with this series of molecules. Together these observations provide a critical forward step in the identification of CRAC blockers suitable for clinical development in the treatment of inflammatory disorders. Copyright © 2013 Elsevier B.V. All rights reserved.

Characterization of 1-Aminobenzotriazole and Ketoconazole as Novel Inhibitors of Monoamine Oxidase (MAO): An In Vitro Investigation.

PubMed

Shaik, Abdul Naveed; LeDuc, Barbara W; Khan, Ansar A

2017-10-01

1-Aminobenzotriazole, a known time-dependent inhibitor of cytochrome P450 (CYP) enzymes, and ketoconazole, a strong inhibitor of the human CYP3A4 isozyme, are used as standard probe inhibitors to characterize the CYP and/or non-CYP-mediated metabolism of xenobiotics. In the present investigation, 1-Aminobenzotriazole and ketoconazole are characterized as potent monoamine oxidase (MAO) inhibitors in vitro using mouse, rat and human liver microsomes and S9 fractions. Inhibition potential of 1-aminobenzotriazole and ketoconazole was studied in mice, rat and human liver microsomes, S9 fractions, MAO-A and MAO-B expressed enzymes by monitoring the formation of 4-hydroxyquinoline (4-HQ) from kynuramine, a specific substrate of MAO by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Mechanism of MAO inhibition was studied by incubating varying concentration of kynuramine with mouse, rat and human S9 fractions at varying concentration of 1-aminobenzatriazole and ketoconazole and monitoring the formation of 4-HQ. 1-aminobenzotriazole and ketoconazole inhibited both MAO isozymes (MAO-A and MAO-B) with more specificity towards MAO-B. Kynuramine substrate kinetics in mouse, rat and human S9 fractions with varying 1-aminobenzotriazole and ketoconazole concentrations showed decreased maximum rate (V max ) for 4-HQ formation without affecting the Michaelis-Menten constant (K m ). A non-competitive inhibition model was constructed and inhibition constants (K i ) for 1-aminobenzotriazole (7.87 ± 0.61, 8.61 ± 0.92, 65.2 ± 1.61 µM for mice, rat and humans, respectively) and ketoconazole (0.12 ± 0.01, 2.04 ± 0.08, 5.52 ± 0.47 µM for mice, rat and humans, respectively) were determined. 1-Aminobenzotriazole and ketoconazole are characterized as non-competitive inhibitors of mice, rat and human MAO in vitro and the extent of their MAO inhibition potential is species specific. 1-Aminobenzotriazole or ketoconazole can be used as a probe inhibitor in vitro for screening the involvement of MAO-dependent metabolism of new chemical entities (NCE) in early drug discovery.

In Vitro Metabolism of 20(R)-25-Methoxyl-Dammarane-3, 12, 20-Triol from Panax notoginseng in Human, Monkey, Dog, Rat, and Mouse Liver Microsomes

PubMed Central

Li, Wei; Liu, Li; Sun, Baoshan; Guo, Zhenghong; Shi, Caihong; Zhao, Yuqing

2014-01-01

The present study characterized in vitro metabolites of 20(R)-25-methoxyl-dammarane-3β, 12β, 20-triol (20(R)-25-OCH3-PPD) in mouse, rat, dog, monkey and human liver microsomes. 20(R)-25-OCH3-PPD was incubated with liver microsomes in the presence of NADPH. The reaction mixtures and the metabolites were identified on the basis of their mass profiles using LC-Q/TOF and were quantified using triple quadrupole instrument by multiple reaction monitoring. A total of 7 metabolites (M1–M7) of the phase I metabolites were detected in all species. 25(R)-OCH3-PPD was metabolized by hydroxylation, dehydrogenation, and O-demethylation. Enzyme kinetic of 20(R)-25-OCH3-PPD metabolism was evaluated in rat and human hepatic microsomes. Incubations studies with selective chemical inhibitors demonstrated that the metabolism of 20(R)-25-OCH3-PPD was primarily mediated by CYP3A4. We conclude that 20(R)-25-OCH3-PPD was metabolized extensively in mammalian species of mouse, rat, dog, monkey, and human. CYP3A4-catalyzed oxygenation metabolism played an important role in the disposition of 25(R)-OCH3-PPD, especially at the C-20 hydroxyl group. PMID:24736630

Dietary modulation and structure prediction of rat mucosal pentraxin (Mptx) protein and loss of function in humans

PubMed Central

van der Meer-van Kraaij, Cindy; Siezen, Roland; Kramer, Evelien; Reinders, Marjolein; Blokzijl, Hans; van der Meer, Roelof

2007-01-01

Mucosal pentraxin (Mptx), identified in rats, is a short pentraxin of unknown function. Other subfamily members are Serum amyloid P component (SAP), C-reactive protein (CRP) and Jeltraxin. Rat Mptx mRNA is predominantly expressed in colon and in vivo is strongly (30-fold) regulated by dietary heme and calcium, modulators of colon cancer risk. This renders Mptx a potential nutrient sensitive biomarker of gut health. To support a role as biomarker, we examined whether the pentraxin protein structure is conserved, whether Mptx protein is nutrient-sensitively expressed and whether Mptx is expressed in mouse and human. Sequence comparison and 3D modelling showed that rat Mptx is highly homologous to the other pentraxins. The calcium-binding site and subunit interaction sites are highly conserved, while a loop deletion and charged residues contribute to a distinctive “top” face of the pentamer. In accordance with mRNA expression, Mptx protein is strongly down-regulated in rat colon mucosa in response to high dietary heme intake. Mptx mRNA is expressed in rat and mouse colon, but not in human colon. A stop codon at the beginning of human exon two indicates loss of function, which may be related to differences in intestinal cell turnover between man and rodents. PMID:18850182

A novel lens epithelium gene, LEP503, is highly conserved in different vertebrate species and is developmentally regulated in postnatal rat lens.

PubMed

Wen, Y; Sachs, G; Athmann, C

2000-02-01

The development of the lens is dependent on the proliferation of lens epithelial cells and their differentiation into fiber cells near the lens bow/equator. Identification of genes specifically expressed in the lens epithelial cells and their functions may provide insight into molecular events that regulate the processes of lens epithelial cell differentiation. In this study, a novel lens epithelium gene product, LEP503, identified from rat by a subtractive cDNA cloning strategy was investigated in the genome organization, mRNA expression and protein localization. The genomic sequences for LEP503 isolated from rat, mouse and human span 1754 bp, 1694 bp and 1895 bp regions encompassing the 5'-flanking region, two exons, one intron and 3'-flanking region. All exon-intron junction sequences conform to the GT/AG rule. Both mouse and human LEP503 genes show very high identity (93% for mouse and 79% for human) to rat LEP503 gene in the exon 1 that contains an open reading frame coding for a protein of 61 amino acid residues with a leucine-rich domain. The deduced protein sequences also show high identity (91% between mouse and rat and 77% between human and rat). Western blot shows that LEP503 is present as a specific approximately 6.9 kDa band in the water-insoluble-urea-soluble fraction of lens cortex where lens epithelium is included. Immuno-staining shows that LEP503 is localized in the epithelial cells along the entire anterior surface of rat lens. Developmentally, LEP503 is expressed at a low level at newborn, and then the expression level increases by about ten-fold around postnatal day 14 and remains at this high level for about 25 days before it drops back to the low level by postnatal day 84. These data suggest that the LEP503 may be an important lens epithelial cell gene involving the processes of epithelial cell differentiation. Copyright 2000 Academic Press.

Pharmacological profiling of the TRPV3 channel in recombinant and native assays

PubMed Central

Grubisha, Olivera; Mogg, Adrian J; Sorge, Jessica L; Ball, Laura-Jayne; Sanger, Helen; Ruble, Cara L A; Folly, Elizabeth A; Ursu, Daniel; Broad, Lisa M

2014-01-01

Background and Purpose Transient receptor potential vanilloid subtype 3 (TRPV3) is implicated in nociception and certain skin conditions. As such, it is an attractive target for pharmaceutical research. Understanding of endogenous TRPV3 function and pharmacology remains elusive as selective compounds and native preparations utilizing higher throughput methodologies are lacking. In this study, we developed medium-throughput recombinant and native cellular assays to assess the detailed pharmacological profile of human, rat and mouse TRPV3 channels. Experimental Approach Medium-throughput cellular assays were developed using a Ca2+-sensitive dye and a fluorescent imaging plate reader. Human and rat TRPV3 pharmacology was examined in recombinant cell lines, while the mouse 308 keratinocyte cell line was used to assess endogenous TRPV3 activity. Key Results A recombinant rat TRPV3 cellular assay was successfully developed after solving a discrepancy in the published rat TRPV3 protein sequence. A medium-throughput, native, mouse TRPV3 keratinocyte assay was also developed and confirmed using genetic approaches. Whereas the recombinant human and rat TRPV3 assays exhibited similar agonist and antagonist profiles, the native mouse assay showed important differences, namely, TRPV3 activity was detected only in the presence of potentiator or during agonist synergy. Furthermore, the native assay was more sensitive to block by some antagonists. Conclusions and Implications Our findings demonstrate similarities but also notable differences in TRPV3 pharmacology between recombinant and native systems. These findings offer insights into TRPV3 function and these assays should aid further research towards developing TRPV3 therapies. Linked Articles This article is part of a themed section on the pharmacology of TRP channels. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171.issue-10 PMID:23848361

Metabolism of Ginger Component [6]-Shogaol in Liver Microsomes from Mouse, Rat, Dog, Monkey, and Human

PubMed Central

Chen, Huadong; Soroka, Dominique; Zhu, Yingdong; Sang, Shengmin

2013-01-01

Scope There are limited data on the metabolism of [6]-shogaol, a major bioactive component of ginger. This study demonstrates metabolism of [6]-shogaol in liver microsomes from mouse, rat, dog, monkey, and human. Methods and results The in vitro metabolism of [6]-shogaol was compared among five species using liver microsomes from mouse, rat, dog, monkey, and human. Following incubations with [6]-shogaol, three major reductive metabolites 1-(4'-hydroxy-3'-methoxyphenyl)-4-decen-3-ol (M6), 1-(4′-hydroxy-3′-methoxyphenyl)-decan-3-ol (M9), and 1-(4'-hydroxy-3'-methoxyphenyl)-decan-3-one (M11), as well as two new oxidative metabolites (1E, 4E)-1-(4'-hydroxy-3'-methoxyphenyl)-deca-1,4-dien-3-one (M14) and (E)-1-(4'-hydroxy-3'-methoxyphenyl)-dec-1-en-3-one (M15) were found in all species. The kinetic parameters of M6 in liver microsomes from each respective species were quantified using Michaelis-Menten theory. A broad CYP-450 inhibitor, 1-aminobenzotriazole, precluded the formation of oxidative metabolites M14 and M15, and 18β-glycyrrhetinic acid, an aldo-keto reductase inhibitor, eradicated the formation of the reductive metabolites M6, M9, and M11 in all species. Metabolites M14 and M15 were tested for cancer cell growth inhibition and induction of apoptosis and both showed substantial activity, with M14 displaying greater potency than [6]-shogaol. Conclusion We conclude that [6]-shogaol is metabolized extensively in mammalian species mouse, rat, dog, monkey, and human, and that there are significant interspecies differences to consider when planning pre-clinical trials towards [6]-shogaol chemoprevention. PMID:23322474

Metabolism of ginger component [6]-shogaol in liver microsomes from mouse, rat, dog, monkey, and human.

PubMed

Chen, Huadong; Soroka, Dominique; Zhu, Yingdong; Sang, Shengmin

2013-05-01

There are limited data on the metabolism of [6]-shogaol (6S), a major bioactive component of ginger. This study demonstrates metabolism of 6S in liver microsomes from mouse, rat, dog, monkey, and human. The in vitro metabolism of 6S was compared among five species using liver microsomes from mouse, rat, dog, monkey, and human. Following incubations with 6S, three major reductive metabolites 1-(4'-hydroxy-3'-methoxyphenyl)-4-decen-3-ol (M6), 1-(4'-hydroxy-3'-methoxyphenyl)-decan-3-ol (M9), and 1-(4'-hydroxy-3'-methoxyphenyl)-decan-3-one (M11), as well as two new oxidative metabolites (1E,4E)-1-(4'-hydroxy-3'-methoxyphenyl)-deca-1,4-dien-3-one (M14) and (E)-1-(4'-hydroxy-3'-methoxyphenyl)-dec-1-en-3-one (M15) were found in all species. The kinetic parameters of M6 in liver microsomes from each respective species were quantified using Michaelis-Menten theory. A broad CYP-450 inhibitor, 1-aminobenzotriazole, precluded the formation of oxidative metabolites, M14 and M15, and 18β-glycyrrhetinic acid, an aldo-keto reductase inhibitor, eradicated the formation of the reductive metabolites M6, M9, and M11 in all species. Metabolites M14 and M15 were tested for cancer cell growth inhibition and induction of apoptosis and both showed substantial activity, with M14 displaying greater potency than 6S. We conclude that 6S is metabolized extensively in mammalian species mouse, rat, dog, monkey, and human, and that there are significant interspecies differences to consider when planning preclinical trials toward 6S chemoprevention. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Use of Terbinafine in Mouse and Rat Models of Pneumocystis carinii Pneumonia

PubMed Central

Walzer, Peter D.; Ashbaugh, Alan

2002-01-01

Terbinafine, an allylamine used to treat onychomycosis, has been reported to be active against rat Pneumocystis carinii in vitro and in vivo. By contrast, our in vitro data showed that the 50% inhibitory concentration of terbinafine against rat P. carinii is 3.7 μg/ml, a level that cannot be clinically achieved in serum. In the present study, terbinafine administered orally at doses of 20 to 400 mg/kg/day and 50 to 250 mg/kg/day was ineffective therapy for mouse and rat models of pneumocystosis, respectively. These results emphasize the complexities of P. carinii drug testing and the need for caution before considering studies in humans. PMID:11796365

Comparative Evaluation of Gemcabene and PPAR Ligands in Transcriptional Assays of Peroxisome Proliferator-Activated Receptors: Implication for the Treatment of Hyperlipidemia and Cardiovascular Disease.

PubMed

Bisgaier, Charles L; Oniciu, Daniela C; Srivastava, Rai Ajit K

2018-03-21

Gemcabene, a late-stage clinical candidate, has shown efficacy for LDL-C, non-HDL cholesterol, apoB, triglycerides and hsCRP reduction, all risk factors for cardiovascular disease (CVD). In rodents, gemcabene showed changes in targets, including apoC-III, apoA-I, peroxisomal enzymes, considered regulated via PPAR gene activation, suggesting a PPAR-mediated mechanism of action for the observed hypolipidemic effects observed in rodents and humans. In the current study, the gemcabene agonist activity against PPAR subtypes of human, rat and mouse were compared to known lipid lowering PPAR activators. Surprisingly, gemcabene showed no or little PPAR-α transactivation compared with reference agonists, which showed concentration-dependent transactivation against human PPAR-α of 2.4 to 30-fold (fenofibric acid), 17-fold (GW590735), and 2.3 to 25-fold (WY14643). These agents also showed robust transactivation of mouse and rat PPAR-α in a concentration-dependent manner. The known PPAR-δ agonists, GW1516, L165041 and GW0742, showed potent agonist activity against human, mouse and rat receptors (ranging from 165- to 396-fold). In contrast, gemcabene at the highest concentration tested (300 µM) showed no response in mouse and rat and a marginal response against human PPAR-δ receptors (3.2-fold). For PPAR-γ, gemcabene showed no agonist activity against all 3 species at 100 µM and marginal activity (3.6-5 fold) at 300 µM. In contrast, the known agonists, rosiglitazone, indomethacin and muraglitazar showed strong activation against the mouse, rat and human PPAR-γ receptors. No clear antagonist activity was observed with gemcabene against any PPAR-subtypes for all 3 species over a wide range of concentrations. In summary, the transactivation studies rule out gemcabene as a direct agonist or antagonist of PPAR-α, PPAR-γ, and PPAR-δ receptors of these three species. These data suggest that the peroxisomal effects observed in rodents and the lipid regulating effects observed in rodents and humans are not related to a direct activation of PPAR receptors by gemcabene.This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives License 4.0 (CCBY-NC-ND), where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal.

A detailed analysis of the erythropoietic control system in the human, squirrel, monkey, rat and mouse

NASA Technical Reports Server (NTRS)

Nordheim, A. W.

1985-01-01

The erythropoiesis modeling performed in support of the Body Fluid and Blood Volume Regulation tasks is described. The mathematical formulation of the species independent model, the solutions to the steady state and dynamic versions of the model, and the individual species specific models for the human, squirrel monkey, rat and mouse are outlined. A detailed sensitivity analysis of the species independent model response to parameter changes and how those responses change from species to species is presented. The species to species response to a series of simulated stresses directly related to blood volume regulation during space flight is analyzed.

Structural features of a close homologue of L1 (CHL1) in the mouse: a new member of the L1 family of neural recognition molecules.

PubMed

Holm, J; Hillenbrand, R; Steuber, V; Bartsch, U; Moos, M; Lübbert, H; Montag, D; Schachner, M

1996-08-01

We have identified a close homologue of L1 (CHL1) in the mouse. CHL1 comprises an N-terminal signal sequence, six immunoglobulin (Ig)-like domains, 4.5 fibronectin type III (FN)-like repeats, a transmembrane domain and a C-terminal, most likely intracellular domain of approximately 100 amino acids. CHL1 is most similar in its extracellular domain to chicken Ng-CAM (approximately 40% amino acid identity), followed by mouse L1, chicken neurofascin, chicken Nr-CAM, Drosophila neuroglian and zebrafish L1.1 (37-28% amino acid identity), and mouse F3, rat TAG-1 and rat BIG-1 (approximately 27% amino acid identity). The similarity with other members of the Ig superfamily [e.g. neural cell adhesion molecule (N-CAM), DCC, HLAR, rse] is 16-11%. The intracellular domain is most similar to mouse and chicken Nr-CAM, mouse and rat neurofascin (approximately 60% amino acid identity) followed by chicken neurofascin and Ng-CAM, Drosophila neuroglian and zebrafish L1.1 and L1.2 (approximately 40% amino acid identity). Besides the high overall homology and conserved modular structure among previously recognized members of the L1 family (mouse/human L1/rat NILE; chicken Ng-CAM; chicken/mouse Nr-CAM; Drosophila neuroglian; zebrafish L1.1 and L1.2; chicken/mouse neurofascin/rat ankyrin-binding glycoprotein), criteria characteristic of L1 were identified with regard to the number of amino acids between positions of conserved amino acid residues defining distances within and between two adjacent Ig-like domains and FN-like repeats. These show a collinearity in the six Ig-like domains and four adjacent FN-like repeats that is remarkably conserved between L1 and molecules containing these modules (designated the L1 family cassette), including the GPI-linked forms of the F3 subgroup (mouse F3/chicken F11/human CNTN1; rat BIG-1/mouse PANG; rat TAG-1/mouse TAX-1/chicken axonin-1). The colorectal cancer molecule (DCC), previously introduced as an N-CAM-like molecule, conforms to the L1 family cassette. Other structural features of CHL 1 shared between members of the L1 family are a high degree of N-glycosidically linked carbohydrates (approximately 20% of its molecular mass), which include the HNK-1 carbohydrate structure, and a pattern of protein fragments comprising a major 185 kDa band and smaller fragments of 165 and 125 kDa. As for the other L1 family members, predominant expression of CHL1 is observed in the nervous system and at later developmental stages. In the central nervous system CHL1 is expressed by neurons, but, in contrast to L1, also by glial cells. Our findings suggest a common ancestral L1-like molecule which evolved via gene duplication to generate a diversity of structurally and functionally distinct yet similar molecules.

Correlation of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine Neurotoxicity with Blood-Brain Barrier Monoamine Oxidase Activity

NASA Astrophysics Data System (ADS)

Kalaria, Rajesh N.; Mitchell, Mary Jo; Harik, Sami I.

1987-05-01

Systemic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) causes parkinsonism in humans and subhuman primates, but not in rats and many other laboratory animals; mice are intermediate in their susceptibility. Since MPTP causes selective dopaminergic neurotoxicity when infused directly into rat substantia nigra, we hypothesized that systemic MPTP may be metabolized by monoamine oxidase and/or other enzymes in rat brain capillaries and possibly other peripheral organs and thus prevented from reaching its neuronal sites of toxicity. We tested this hypothesis by assessing monoamine oxidase in isolated cerebral microvessels of humans, rats, and mice by measuring the specific binding of [3H]pargyline, an irreversible monoamine oxidase inhibitor, and by estimating the rates of MPTP and benzylamine oxidation. [3H]Pargyline binding to rat cerebral microvessels was about 10-fold higher than to human or mouse microvessels. Also, MPTP oxidation by rat brain microvessels was about 30-fold greater than by human microvessels; mouse microvessels yielded intermediate values. These results may explain, at least in part, the marked species differences in susceptibility to systemic MPTP. They also suggest the potential importance of ``enzyme barriers'' at the blood-brain interface that can metabolize toxins not excluded by structural barriers, and may provide biological bases for developing therapeutic strategies for the prevention of MPTP-induced neurotoxicity and other neurotoxic conditions including, possibly, Parkinson disease.

Concerns about the widespread use of rodent models for human risk assessments of endocrine disruptors

PubMed Central

Habert, René; Muczynski, Vincent; Grisin, Tiphany; Moison, Delphine; Messiaen, Sébastien; Frydman, René; Benachi, Alexandra; Delbes, Géraldine; Lambrot, Romain; Lehraiki, Abdelali; N'Tumba-Byn, Thierry; Guerquin, Marie-Justine; Levacher, Christine; Rouiller-Fabre, Virginie; Livera, Gabriel

2014-01-01

Fetal testis is a major target of endocrine disruptors (EDs). During the last 20 years, we have developed an organotypic culture system that maintains the function of the different fetal testis cell types and have used this approach as a toxicological test to evaluate the effects of various compounds on gametogenesis and steroidogenesis in rat, mouse and human testes. We named this test rat, mouse and human fetal testis assay. With this approach, we compared the effects of six potential EDs ((mono-(2-ethylhexyl) phthalate (MEHP), cadmium, depleted uranium, diethylstilboestrol (DES), bisphenol A (BPA) and metformin) and one signalling molecule (retinoic acid (RA)) on the function of rat, mouse and human fetal testis at a comparable developmental stage. We found that the response is similar in humans and rodents for only one third of our analyses. For instance, RA and MEHP have similar negative effects on gametogenesis in the three species. For another third of our analyses, the threshold efficient concentrations that disturb gametogenesis and/or steroidogenesis differ as a function of the species. For instance, BPA and metformin have similar negative effects on steroidogenesis in human and rodents, but at different threshold doses. For the last third of our analyses, the qualitative response is species specific. For instance, MEHP and DES affect steroidogenesis in rodents, but not in human fetal testis. These species differences raise concerns about the extrapolation of data obtained in rodents to human health risk assessment and highlight the need of rigorous comparisons of the effects in human and rodent models, when assessing ED risk. PMID:24497529

A physiologically based pharmacokinetic model for ethylene oxide in mouse, rat, and human.

PubMed

Fennell, T R; Brown, C D

2001-06-15

Ethylene oxide (EO) is widely used as a gaseous sterilant and industrial intermediate and is a direct-acting mutagen and carcinogen. The objective of these studies was to develop physiologically based pharmacokinetic (PB-PK) models for EO to describe the exposure-tissue dose relationship in rodents and humans. We previously reported results describing in vitro and in vivo kinetics of EO metabolism in male and female F344 rats and B6C3F1 mice. These studies were extended by determining the kinetics of EO metabolism in human liver cytosol and microsomes. The results indicate enzymatically catalyzed GSH conjugation via cytosolic glutathione S-transferase (cGST) and hydrolysis via microsomal epoxide hydrolase (mEH) occur in both rodents and humans. The in vitro kinetic constants were scaled to account for cytosolic (cGST) and microsomal (mEH) protein content and incorporated into PB-PK descriptions for mouse, rat, and human. Flow-limited models adequately predicted blood and tissue EO levels, disposition, and elimination kinetics determined experimentally in rats and mice, with the exception of testis concentrations, which were overestimated. Incorporation of a diffusion-limited description for testis improved the ability of the model to describe testis concentrations. The model accounted for nonlinear increases in blood and tissue concentrations that occur in mice on exposure to EO concentrations greater than 200 ppm. Species differences are predicted in the metabolism and exposure-dose relationship, with a nonlinear relationship observed in the mouse as a result of GSH depletion. These models represent an essential step in developing a mechanistically based EO exposure-dose-response description for estimating human risk from exposure to EO. Copyright 2001 Academic Press.

Comparative pharmacokinetics of perfluorononanoic acid in rat and mouse

EPA Science Inventory

Perfluorononanoic acid (PFNA) is a fluorinated organic chemical found at low levels in the environment, but is detectable in humans and wildlife. The present study compared the pharmacokinetic properties of PFNA in two laboratory rodent species. Male and female Sprague-Dawley rat...

Rats, cats, and elephants, but still no unicorn: induced pluripotent stem cells from new species.

PubMed

Trounson, Alan

2009-01-09

Two independent studies in this issue of Cell Stem Cell (Liao et al., 2009; Li et al., 2009) derive rat induced pluripotent stem cells (iPSCs). In one report, the method used results in rat and human iPSCs that exhibit phenotypic traits similar to mouse embryonic stem cells.

Review of the carcinogenic activity of diethanolamine and evidence of choline deficiency as a plausible mode of action.

PubMed

Leung, Hon-Wing; Kamendulis, Lisa M; Stott, William T

2005-12-01

Diethanolamine (DEA) is a chemical used widely in a number of industries and is present in many consumer products. Studies by the National Toxicology Program (NTP) have indicated that lifetime dermal exposure to DEA increased the incidence and multiplicity of liver tumors in mice, but not in rats. In addition, DEA was not carcinogenic when tested in the Tg.Ac transgenic mouse model. Short-term genotoxicity tests have yielded negative results. In view of these apparent inconsistencies, we have critically evaluated the NTP studies and other data relevant to assessing the carcinogenic potential of DEA. The available data indicate that DEA induces mouse liver tumors by a non-genotoxic mode of action that involves its ability to cause choline deficiency. The following experimental evidence supports this hypothesis. DEA decreased the hepatic choline metabolites and S-adenosylmethionine levels in mice, similar to those observed in choline-deficient mice. In contrast, DEA had no effect in the rat, a species in which it was not carcinogenic at a maximum tolerated dose level. In addition, a consistent dose-effect relationship had been established between choline deficiency and carcinogenic activity since all DEA dosages that induced tumors in the NTP studies were also shown to cause choline deficiency. DEA decreased phosphatidylcholine synthesis by blocking the cellular uptake of choline in vitro, but these events did not occur in the presence of excess choline. Finally, DEA induced transformation in the Syrian hamster embryo cells, increased S-phase DNA synthesis in mouse hepatocytes, and decreased gap junctional intracellular communication in primary cultured mouse and rat hepatocytes, but all these events were prevented with choline supplementation. Since choline is an essential nutrient in mammals, this mode of action is qualitatively applicable to humans. However, there are marked species differences in susceptibility to choline deficiency, with rats and mice being far more susceptible than other mammalian species including humans. These differences are attributed to quantitative differences in the enzyme kinetics controlling choline metabolism. The fact that DEA was carcinogenic in mice but not in rats also has important implications for human risk assessment. DEA has been shown to be less readily absorbed across rat and human skin than mouse skin. Since a no observed effect level for DEA-induced choline deficiency in mice has been established to be 10 mg/kg/d, this indicates that there is a critical level of DEA that must be attained in order to affect choline homeostasis. The lack of a carcinogenic response in rats suggests that exposure to DEA did not reach this critical level. Since rodents are far more sensitive to choline deficiency than humans, it can be concluded that the hepatocarcinogenic effect of DEA in mice is not predictive of similar susceptibility in humans.

Effects of clofibric acid on mRNA expression profiles in primary cultures of rat, mouse and human hepatocytes.

PubMed

Richert, Lysiane; Lamboley, Christelle; Viollon-Abadie, Catherine; Grass, Peter; Hartmann, Nicole; Laurent, Stephane; Heyd, Bruno; Mantion, Georges; Chibout, Salah-Dine; Staedtler, Frank

2003-09-01

The mRNA expression profile in control and clofibric acid (CLO)-treated mouse, rat, and human hepatocytes was analyzed using species-specific oligonucleotide DNA microarrays (Affymetrix). A statistical empirical Bayes procedure was applied in order to select the significantly differentially expressed genes. Treatment with the peroxisome proliferator CLO induced up-regulation of genes involved in peroxisome proliferation and in cell proliferation as well as down-regulation of genes involved in apoptosis in hepatocytes of rodent but not of human origin. CLO treatment induced up-regulation of microsomal cytochrome P450 4a genes in rodent hepatocytes and in two of six human hepatocyte cultures. In addition, genes encoding phenobarbital-inducible cytochrome P450s were also up-regulated by CLO in rodent and human hepatocyte cultures. Up-regulation of phenobarbital-inducible UDP-glucuronosyl-transferase genes by CLO was observed in both rat and human but not in mouse hepatocytes. CLO treatment induced up-regulation of L-fatty acid binding protein (L-FABP) gene in hepatocytes of both rodent and human origin. However, while genes of the cytosolic, microsomal, and mitochondrial pathways involved in fatty acid transport and metabolism were up-regulated by CLO in both rodent and human hepatocyte cultures, genes of the peroxisomal pathway of lipid metabolism were up-regulated in rodents only. An up-regulation of hepatocyte nuclear factor 1alpha (HNF1alpha) by CLO was observed only in human hepatocyte cultures, suggesting that this trans-activating factor may play a key role in the regulation of fatty acid metabolism in human liver as well as in the nonresponsiveness of human liver to CLO-induced regulation of cell proliferation and apoptosis.

Expression of glutathione peroxidase I gene in selenium-deficient rats.

PubMed Central

Reddy, A P; Hsu, B L; Reddy, P S; Li, N Q; Thyagaraju, K; Reddy, C C; Tam, M F; Tu, C P

1988-01-01

We have characterized a cDNA pGPX1211 encoding rat glutathione peroxidase I. The selenocysteine in the protein corresponded to a TGA codon in the coding region of the cDNA, similar to earlier findings in mouse and human genes, and a gene encoding the formate dehydrogenase from E. coli, another selenoenzyme. The rat GSH peroxidase I has a calculated subunit molecular weight of 22,155 daltons and shares 95% and 86% sequence homology with the mouse and human subunits, respectively. The 3'-noncoding sequence (greater than 930 bp) in pGPX1211 is much longer than that of the human sequences. We found that glutathione peroxidase I mRNA, but not the polypeptide, was expressed under nutritional stress of selenium deficiency where no glutathione peroxidase I activity can be detected. The failure of detecting any apoprotein for the glutathione peroxidase I under selenium deficiency and results published from other laboratories supports the proposal that selenium may be incorporated into the glutathione peroxidase I co-translationally. Images PMID:2838821

A gene catalogue of the Sprague-Dawley rat gut metagenome.

PubMed

Pan, Hudan; Guo, Ruijin; Zhu, Jie; Wang, Qi; Ju, Yanmei; Xie, Ying; Zheng, Yanfang; Wang, Zhifeng; Li, Ting; Liu, Zhongqiu; Lu, Linlin; Li, Fei; Tong, Bin; Xiao, Liang; Xu, Xun; Li, Runze; Yuan, Zhongwen; Yang, Huanming; Wang, Jian; Kristiansen, Karsten; Jia, Huijue; Liu, Liang

2018-05-01

Laboratory rats such as the Sprague-Dawley (SD) rats are an important model for biomedical studies in relation to human physiological or pathogenic processes. Here we report the first catalog of microbial genes in fecal samples from Sprague-Dawley rats. The catalog was established using 98 fecal samples from 49 SD rats, divided in 7 experimental groups, and collected at different time points 30 days apart. The established gene catalog comprises 5,130,167 non-redundant genes with an average length of 750 bp, among which 64.6% and 26.7% were annotated to phylum and genus levels, respectively. Functionally, 53.1%, 21.8%,and 31% of the genes could be annotated to KEGG orthologous groups, modules, and pathways, respectively. A comparison of rat gut metagenome catalogue with human or mouse revealed a higher pairwise overlap between rats and humans (2.47%) than between mice and humans (1.19%) at the gene level. Ninety-seven percent of the functional pathways in the human catalog were present in the rat catalogue, underscoring the potential use of rats for biomedical research.

Marker expression, behaviors, and responses vary in different lines of conditionally immortalized cultured podocytes

PubMed Central

Chittiprol, Seetharamaiah; Chen, Phylip; Petrovic-Djergovic, Danica; Eichler, Tad

2011-01-01

The state-of-the-art cultured podocyte is conditionally immortalized by expression of a temperature-sensitive mutant of the SV40 large-T antigen. These cultures proliferate at 33°C and differentiate at 37°C into arborized cells that more closely resemble in vivo podocytes. However, the degree of resemblance remains controversial. In this study, several parameters were measured in podocyte cell lines derived from mouse (JR, KE), human (MS), and rat (HK). In all lines, the quantities of NEPH1 and podocin proteins and NEPH1 and SYNPO mRNAs were comparable to glomeruli, while synaptopodin and nephrin proteins and NPHS1 and NPHS2 mRNAs were <5% of glomerular levels. Expression of Wilms' tumor-1 (WT1) mRNA in mouse lines was comparable to glomeruli, but rat and human lines expressed little WT1. Undifferentiated human and mouse lines had similar proliferation rates that decreased after differentiation, while the rate in rat cells remained constant. The motility of different lines varied as measured by both general motility and wound-healing assays. The toxicity of puromycin aminonucleoside was MS ∼ JR >> KE, and of doxorubicin was JR ∼ KE > MS, while HK cells were almost unaffected. Process formation was largely a result of contractile action after formation of lamellipodia. These findings demonstrate dramatic differences in marker expression, response to toxins, and motility between lines of podocytes from different species and even between similarly-derived mouse lines. PMID:21632959

Marker expression, behaviors, and responses vary in different lines of conditionally immortalized cultured podocytes.

PubMed

Chittiprol, Seetharamaiah; Chen, Phylip; Petrovic-Djergovic, Danica; Eichler, Tad; Ransom, Richard F

2011-09-01

The state-of-the-art cultured podocyte is conditionally immortalized by expression of a temperature-sensitive mutant of the SV40 large-T antigen. These cultures proliferate at 33°C and differentiate at 37°C into arborized cells that more closely resemble in vivo podocytes. However, the degree of resemblance remains controversial. In this study, several parameters were measured in podocyte cell lines derived from mouse (JR, KE), human (MS), and rat (HK). In all lines, the quantities of NEPH1 and podocin proteins and NEPH1 and SYNPO mRNAs were comparable to glomeruli, while synaptopodin and nephrin proteins and NPHS1 and NPHS2 mRNAs were <5% of glomerular levels. Expression of Wilms' tumor-1 (WT1) mRNA in mouse lines was comparable to glomeruli, but rat and human lines expressed little WT1. Undifferentiated human and mouse lines had similar proliferation rates that decreased after differentiation, while the rate in rat cells remained constant. The motility of different lines varied as measured by both general motility and wound-healing assays. The toxicity of puromycin aminonucleoside was MS ∼ JR > KE, and of doxorubicin was JR ∼ KE > MS, while HK cells were almost unaffected. Process formation was largely a result of contractile action after formation of lamellipodia. These findings demonstrate dramatic differences in marker expression, response to toxins, and motility between lines of podocytes from different species and even between similarly-derived mouse lines.

Physiologically based Pharmacokinetic Modeling of 1,4-Dioxane in Rats, Mice, and Humans

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sweeney, Lisa M.; Thrall, Karla D.; Poet, Torka S.

2008-01-01

ABSTRACT 1,4-Dioxane (CAS No. 123-91-1) is used primarily as a solvent or as a solvent stabilizer. It can cause lung, liver and kidney damage at sufficiently high exposure levels. Two physiologically-based pharmacokinetic (PBPK) models of 1,4-dioxane and its major metabolite, hydroxyethoxyacetic acid (HEAA), were published in 1990. These models have uncertainties and deficiencies that could be addressed and the model strengthened for use in a contemporary cancer risk assessment for 1,4-dioxane. Studies were performed to fill data gaps and reduce uncertainties pertaining to the pharmacokinetics of 1,4-dioxane and HEAA in rats, mice, and humans. Three types of studies were performed:partitionmore » coefficient measurements, blood time course in mice, and in vitro pharmacokinetics using rat, mouse, and human hepatocytes. Updated PBPK models were developed based on these new data and previously available data. The optimized rate of metabolism for the mouse was significantly higher than the value previously estimated. The optimized rat kinetic parameters were similar to those in the 1990 models. Only two human studies were identified. Model predictions were consistent with one study, but did not fit the second as well. In addition, a rat nasal exposure was completed. The results confirmed water directly contacts rat nasal tissues during drinking water under bioassays. Consistent with previous PBPK models, nasal tissues were not specifically included in the model. Use of these models will reduce the uncertainty in future 1,4-dioxane risk assessments.« less

Localization of peroxisome proliferator-activated receptor in mouse and rat-tissues and demonstration of its nuclear translocation in transfected cv-1 cells.

PubMed

Huang, Q; Yeldandi, A; Alvares, K; Ide, H; Reddy, J; Rao, M

1995-02-01

Hepatocarcinogenesis in rodents induced by nongenotoxic peroxisome proliferators is postulated to be a receptor-mediated process. The peroxisome proliferator-activated receptors (PPAR) are members of the steroid hormone receptor superfamily, which participate in ligand-dependent transcriptional activation of peroxisomal fatty acid beta oxidation enzyme system genes in liver parenchymal cells of rats and mice. In order to study the tissue distribution and cellular localization of PPAR, we raised polyclonal antibodies against PPAR using a recombinant rat PPAR (rPPAR) expressed as a glutathione-S-transferase-rPPAR fusion protein. On immunoblot analysis the antibodies specifically recognized a 55 kDa PPAR protein in rat, mouse and human liver homogenates. Immunoblotting also showed that in the mouse and rat, PPAR is expressed in liver, kidney and heart, and only weakly in brain and testis. Immunohistochemical localization in the rat and mouse revealed that PPAR is highly expressed in perivenular (i.e., those surrounding hepatic vein) hepatocytes and very weakly in the cytoplasm of remaining hepatocytes. In the kidney, PPAR was visualized predominantly in the p(3) segments of proximal convoluted tubular epithelium. CV-1 cells transiently transfected with rPPAR cDNA construct showed predominant cytoplasmic fluorescence; treatment of these cells with ciprofibrate, a peroxisome proliferator, resulted in the nuclear translocation of PPAR signal.

Validation of an automated counting procedure for phthalate-induced testicular multinucleated germ cells

EPA Science Inventory

In utero exposure to certain phthalate esters results in testicular toxicity, characterized at the tissue level by induction of multinucleated germ cells (MNGs) in rat, mouse, and human fetal testis. Phthalate exposures also result in a decrease in testicular testosterone in rats...

Evidence for triclosan-induced activation of human and rodent xenobiotic nuclear receptors.

PubMed

Paul, Katie B; Thompson, Jerry T; Simmons, Steven O; Vanden Heuvel, John P; Crofton, Kevin M

2013-10-01

The bacteriostat triclosan (2,4,4'-trichloro-2'-hydroxydiphenylether) (TCS) decreases rat serum thyroxine via putative nuclear receptor (NR) interaction(s) and subsequent transcriptional up-regulation of hepatic catabolism and clearance. However, due to the evolutionary divergence of the constitutive androstane and pregnane-X receptors (CAR, PXR), TCS-mediated downstream effects may be species-dependent. To test the hypothesis that TCS activates xenobiotic NRs across species, cell-based NR reporter assays were employed to assess potential activation of rat, mouse, and human PXR, and rat, mouse, and three splice variants of human CAR. TCS activated hPXR, acted as an inverse agonist of hCAR1, and as a weak agonist of hCAR3. TCS failed to activate rPXR in full-length receptor reporter assays, and instead acted as a modest inverse agonist of rCAR. Consistent with the rat data, TCS also failed to activate mPXR and was a modest inverse agonist of mCAR. These data suggest that TCS may interact with multiple NRs, including hPXR, hCAR1, hCAR3, and rCAR in order to potentially affect hepatic catabolism. Overall these data support the conclusion that TCS may interact with NRs to regulate hepatic catabolism and downstream thyroid hormone homeostasis in both rat and human models, though perhaps by divergent mechanisms. Published by Elsevier Ltd.

Changes in leptospirosis etiology in animals and humans.

PubMed

Vasylieva, Natalia; Andreychyn, Mykhaylo; Kravchuk, Yulia; Chervinska, Оlena; Iosyk, Iaryna

2017-12-23

Leptospirosis is endemic in Ternopil region. In Ukraine, the disease is registered in almost all regions, including the Ternopil region. The aim of the research is to study the regularities of epidemic and epizootic processes of leptospirosis, and the circulation of its pathogens among different sources (small mammals, animals) and humans. Etiologic spectrum of leptospirosis registered in Ternopil region in 1972-2016 among small mammals, farm animals and sick people was studied. Due to the analysis of pathogens circulation among different sources (small mammals, animals), as well as the annual morbidity in humans, it was proved that new leptospira serovars are endemic and brought into the regions mostly by farm animals. Farm animals introduce the infection to humans through the environment, sometimes within 3-5-years. The spread was observed of pathogen serovars, which are new in certain areas, among all types of mouse-like small mammals and rats. It was established that livestock and small mammals are parallel reservoirs. In the regions with endemic species, the structural modification in the etiology of leptospirosis in humans is caused by additional reservoirs among animals, as well as the circulation of other pathogen serovars that were absent in the main natural reservoir, i.e. mouse-like small mammals and rats. The constant monitoring of the population, contamination and carrier state of mouse-like small mammals, rats and farm animals, is required In order to predict the future epidemiological situation on leptospirosis among the population and to improve leptospirosis diagnosis.

Schistosoma mansoni Infection of Mice, Rats and Humans Elicits a Strong Antibody Response to a Limited Number of Reduction-Sensitive Epitopes on Five Major Tegumental Membrane Proteins

PubMed Central

Tremblay, Jacqueline M.; Oliveira, Sergio C.; Da’dara, Akram A.; Skelly, Patrick J.

2017-01-01

Schistosomiasis is a major disease of the developing world for which no vaccine has been successfully commercialized. While numerous Schistosoma mansoni worm antigens have been identified that elicit antibody responses during natural infections, little is known as to the identities of the schistosome antigens that are most prominently recognized by antibodies generated through natural infection. Non-reducing western blots probed with serum from schistosome-infected mice, rats and humans on total extracts of larval or adult schistosomes revealed that a small number of antigen bands predominate in all cases. Recognition of each of these major bands was lost when the blots were run under reducing condition. We expressed a rationally selected group of schistosome tegumental membrane antigens in insect host cells, and used the membrane extracts of these cells to unambiguously identify the major antigens recognized by S. mansoni infected mouse, rat and human serum. These results revealed that a limited number of dominant, reduction-sensitive conformational epitopes on five major tegumental surface membrane proteins: SmTsp2, Sm23, Sm29, SmLy6B and SmLy6F, are primary targets of mouse, rat and human S. mansoni infection sera antibodies. We conclude that, Schistosoma mansoni infection of both permissive (mouse) and non-permissive (rat) rodent models, as well as humans, elicit a dominant antibody response recognizing a limited number of conformational epitopes on the same five tegumental membrane proteins. Thus it appears that neither infecting schistosomula nor mature adult schistosomes are substantively impacted by the robust circulating anti-tegumental antibody response they elicit to these antigens. Importantly, our data suggest a need to re-evaluate host immune responses to many schistosome antigens and has important implications regarding schistosome immune evasion mechanisms and schistosomiasis vaccine development. PMID:28095417

Evaluation of 111In-labeled EPep and FibPep as tracers for fibrin SPECT imaging.

PubMed

Starmans, Lucas W E; van Duijnhoven, Sander M J; Rossin, Raffaella; Berben, Monique; Aime, Silvio; Daemen, Mat J A P; Nicolay, Klaas; Grüll, Holger

2013-11-04

Fibrin targeting is an attractive strategy for nuclear imaging of thrombosis, atherosclerosis and cancer. Recently, FibPep, an (111)In-labeled fibrin-binding peptide, was established as a tracer for fibrin SPECT imaging and was reported to allow sensitive detection of minute thrombi in mice using SPECT. In this study, we developed EPep, a novel (111)In-labeled fibrin-binding peptide containing the fibrin-binding domain of the clinically verified EP-2104R peptide, and sought to compare the potential of EPep and FibPep as tracers for fibrin SPECT imaging. In vitro, both EPep and FibPep showed high stability in serum, but were less stable in liver and kidney homogenate assays. Both peptide probes displayed comparable affinities toward human and mouse derived fibrin (Kd ≈ 1 μM), and similarly to FibPep, EPep showed fast blood clearance, low nontarget uptake and high thrombus uptake (6.8 ± 1.2% ID g(-1)) in a mouse carotid artery thrombosis model. Furthermore, EPep showed a similar affinity toward rat derived fibrin (Kd ≈ 1 μM), displayed high thrombus uptake in a rat carotid artery thrombosis model (0.74 ± 0.39% ID g(-1)), and allowed sensitive detection of thrombosis in rats using SPECT. In contrast, FibPep displayed a significantly lower affinity toward rat derived fibrin (Kd ≈ 14 μM) and low uptake in rat thrombi (0.06 ± 0.02% ID g(-1)) and did not allow clear visualization of carotid artery thrombosis in rats using SPECT. These results were confirmed ex vivo by autoradiography, which showed a 7-fold higher ratio of activity in the thrombus over the contralateral carotid artery for EPep in comparison to FibPep. These findings suggest that the FibPep binding fibrin epitope is not fully homologous between humans and rats, and that preclinical rat models of disease should not be employed to gauge the clinical potential of FibPep. In conclusion, both peptides showed approximately similar metabolic stability and affinity toward human and mouse derived fibrin, and displayed high thrombus uptake in a mouse carotid artery thrombosis model. Therefore, both EPep and FibPep are promising fibrin targeted tracers for translation into clinical settings to serve as novel tools for molecular imaging of fibrin.

A selective and potent CXCR3 antagonist SCH 546738 attenuates the development of autoimmune diseases and delays graft rejection

PubMed Central

2012-01-01

Background The CXCR3 receptor and its three interferon-inducible ligands (CXCL9, CXCL10 and CXCL11) have been implicated as playing a central role in directing a Th1 inflammatory response. Recent studies strongly support that the CXCR3 receptor is a very attractive therapeutic target for treating autoimmune diseases, such as rheumatoid arthritis, multiple sclerosis and psoriasis, and to prevent transplant rejection. We describe here the in vitro and in vivo pharmacological characterizations of a novel and potent small molecule CXCR3 antagonist, SCH 546738. Results In this study, we evaluated in vitro pharmacological properties of SCH 546738 by radioligand receptor binding and human activated T cell chemotaxis assays. In vivo efficacy of SCH 546738 was determined by mouse collagen-induced arthritis, rat and mouse experimental autoimmune encephalomyelitis, and rat cardiac transplantation models. We show that SCH 546738 binds to human CXCR3 with a high affinity of 0.4 nM. In addition, SCH 546738 displaces radiolabeled CXCL10 and CXCL11 from human CXCR3 with IC50 ranging from 0.8 to 2.2 nM in a non-competitive manner. SCH 546738 potently and specifically inhibits CXCR3-mediated chemotaxis in human activated T cells with IC90 about 10 nM. SCH 546738 attenuates the disease development in mouse collagen-induced arthritis model. SCH 546738 also significantly reduces disease severity in rat and mouse experimental autoimmune encephalomyelitis models. Furthermore, SCH 546738 alone achieves dose-dependent prolongation of rat cardiac allograft survival. Most significantly, SCH 546738 in combination with CsA supports permanent engraftment. Conclusions SCH 546738 is a novel, potent and non-competitive small molecule CXCR3 antagonist. It is efficacious in multiple preclinical disease models. These results demonstrate that therapy with CXCR3 antagonists may serve as a new strategy for treatment of autoimmune diseases, including rheumatoid arthritis and multiple sclerosis, and to prevent transplant rejection. PMID:22233170

Comparative histology of mouse, rat, and human pelvic ligaments.

PubMed

Iwanaga, Ritsuko; Orlicky, David J; Arnett, Jameson; Guess, Marsha K; Hurt, K Joseph; Connell, Kathleen A

2016-11-01

The uterosacral (USL) and cardinal ligaments (CL) provide support to the uterus and pelvic organs, and the round ligaments (RL) maintain their position in the pelvis. In women with pelvic organ prolapse (POP), the connective tissue, smooth muscle, vasculature, and innervation of the pelvic support structures are altered. Rodents are commonly used animal models for POP research. However, the pelvic ligaments have not been defined in these animals. In this study, we hypothesized that the gross anatomy and histological composition of pelvic ligaments in rodents and humans are similar. We performed an extensive literature search for anatomical and histological descriptions of the pelvic support ligaments in rodents. We also performed anatomical dissections of the pelvis to define anatomical landmarks in relation to the ligaments. In addition, we identified the histological components of the pelvic ligaments and performed quantitative analysis of the smooth muscle bundles and connective tissue of the USL and RL. The anatomy of the USL, CL, and RL and their anatomical landmarks are similar in mice, rats, and humans. All species contain the same cellular components and have similar histological architecture. However, the cervical portion of the mouse USL and RL contain more smooth muscle and less connective tissue compared with rat and human ligaments. The pelvic support structures of rats and mice are anatomically and histologically similar to those of humans. We propose that both mice and rats are appropriate, cost-effective models for directed studies in POP research.

Derivation of mouse embryonic stem cell lines from tyrosine hydroxylase reporter mice crossed with a human SNCA transgenic mouse model of Parkinson's disease.

PubMed

Chumarina, Margarita; Azevedo, Carla; Bigarreau, Julie; Vignon, Clémentine; Kim, Kwang-Soo; Li, Jia-Yi; Roybon, Laurent

2017-03-01

Mouse embryonic stem cell (mESC) lines were derived by crossing heterozygous transgenic (tg) mice expressing green fluorescent protein (GFP) under the control of the rat tyrosine hydroxylase (TH) promoter, with homozygous alpha-synuclein (aSYN) mice expressing human mutant SNCA A53T under the control of the mouse Prion promoter (MoPrP), or wildtype (WT) mice. The expression of GFP and human aSYN was validated by immunocytochemistry in midbrain neuron cultures upon differentiation of mESC lines using stromal cell-derived inducing activity. These mESC lines can help to study the impact of human aSYN expression in neurons and oligodendrocytes, and also trace GFP-expressing midbrain neurons. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Transdifferentiated rat pancreatic progenitor cells (AR42J-B13/H) respond to phenobarbital in a rat hepatocyte-specific manner.

PubMed

Osborne, M; Haltalli, M; Currie, R; Wright, J; Gooderham, N J

2016-07-01

Phenobarbital (PB) is known to produce species-specific effects in the rat and mouse, being carcinogenic in certain mouse strains, but only in rats if treated after a DNA damaging event. PB treatment in the rat and mouse also produces disparate effects on cell signalling and miRNA expression profiles. These responses are induced by short term and prolonged PB exposure, respectively, with the latter treatments being difficult to examine mechanistically in primary hepatocytes due to rapid loss of the original hepatic phenotype and limited sustainability in culture. Here we explore the rat hepatocyte-like B13/H cell line as a model for hepatic response to PB exposure in both short-term and longer duration treatments. We demonstrate that PB with Egf treatment in the B13/H cells resulted in a significant increase in Erk activation, as determined by the ratio of phospho-Erk to total Erk, compared to Egf alone. We also show that an extended treatment with PB in the B13/H cells produces a miRNA response similar to that seen in the rat in vivo, via the time-dependent induction of miR-182/96. Additionally, we confirm that B13/H cells respond to Car activators in a typical rat-specific manner. These data suggest that the B13/H cells produce temporal responses to PB that are comparable to those reported in short-term primary rat hepatocyte cultures and in the longer term are similar to those in the rat in vivo. Finally, we also show that Car-associated miR-122 expression is decreased by PB treatment in B13/H cells, a PB-induced response that is common to the rat, mouse and human. We conclude that the B13/H cell system produces a qualitative response comparable to the rat, which is different to the response in the mouse, and that this model could be a useful tool for exploring the functional consequences of PB-sensitive miRNA changes and resistance to PB-mediated tumours in the rat. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

A New Orally Active, Aminothiol Radioprotector-Free of Nausea and Hypotension Side Effects at Its Highest Radioprotective Doses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Soref, Cheryl M.; Hacker, Timothy A.; Fahl, William E., E-mail: fahl@oncology.wisc.edu

Purpose: A new aminothiol, PrC-210, was tested for orally conferred radioprotection (rats, mice; 9.0 Gy whole-body, which was otherwise lethal to 100% of the animals) and presence of the debilitating side effects (nausea/vomiting, hypotension/fainting) that restrict use of the current aminothiol, amifostine (Ethyol, WR-2721). Methods and Materials: PrC-210 in water was administered to rats and mice at times before irradiation, and percent-survival was recorded for 60 days. Subcutaneous (SC) amifostine (positive control) or SC PrC-210 was administered to ferrets (Mustela putorius furo) and retching/emesis responses were recorded. Intraperitoneal amifostine (positive control) or PrC-210 was administered to arterial cannulated rats tomore » score drug-induced hypotension. Results: Oral PrC-210 conferred 100% survival in rat and mouse models against an otherwise 100% lethal whole-body radiation dose (9.0 Gy). Oral PrC-210, administered by gavage 30-90 min before irradiation, conferred a broad window of radioprotection. The comparison of PrC-210 and amifostine side effects was striking because there was no retching or emesis in 10 ferrets treated with PrC-210 and no induced hypotension in arterial cannulated rats treated with PrC-210. The tested PrC-210 doses were the ferret and rat equivalent doses of the 0.5 maximum tolerated dose (MTD) PrC-210 dose in mice. The human equivalent of this mouse 0.5 MTD PrC-210 dose would likely be the highest PrC-210 dose used in humans. By comparison, the mouse 0.5 MTD amifostine dose, 400 {mu}g/g body weight (equivalent to the human amifostine dose of 910 mg/m{sup 2}), when tested at equivalent ferret and rat doses in the above models produced 100% retching/vomiting in ferrets and 100% incidence of significant, progressive hypotension in rats. Conclusions: The PrC-210 aminothiol, with no detectable nausea/vomiting or hypotension side effects in these preclinical models, is a logical candidate for human drug development to use in healthy humans in a wide variety of radioprotection settings, including medical radiation, space travel, and nuclear accidents.« less

Structure of novel rat major histocompatibility complex class II genes RT1.Ha and Hb

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arimura, Yutaka; Tang, Wei Ran; Koda, Toshiaki

1995-03-01

We have cloned the novel rat MHC class II genes, RT1.Ha and Hb, which are homologous to human HLA-DPA and DPB. RT1.Hb is a pseudogene, whereas RT1.Ha is apparently intact and may have transcriptional potential. In addition, with an RT1.Ha probe, we detecteda single Southern hybridization band in the genome of the mouse. This finding may aford an opportunity to analyze the HLA-DPA homologue in the mouse genome. 18 refs., 4 figs., 1 tab.

A new chapter in the bisphenol A story: bisphenol S and bisphenol F are not safe alternatives to this compound.

PubMed

Eladak, Soria; Grisin, Tiphany; Moison, Delphine; Guerquin, Marie-Justine; N'Tumba-Byn, Thierry; Pozzi-Gaudin, Stéphanie; Benachi, Alexandra; Livera, Gabriel; Rouiller-Fabre, Virginie; Habert, René

2015-01-01

Bisphenol A (BPA) is a widely studied typical endocrine-disrupting chemical, and one of the major new issues is the safe replacement of this commonly used compound. Bisphenol S (BPS) and bisphenol F (BPF) are already or are planned to be used as BPA alternatives. With the use of a culture system that we developed (fetal testis assay [FeTA]), we previously showed that 10 nmol/L BPA reduces basal testosterone secretion of human fetal testis explants and that the susceptibility to BPA is at least 100-fold lower in rat and mouse fetal testes. Here, we show that addition of LH in the FeTA system considerably enhances BPA minimum effective concentration in mouse and human but not in rat fetal testes. Then, using the FeTA system without LH (the experimental conditions in which mouse and human fetal testes are most sensitive to BPA), we found that, as for BPA, 10 nmol/L BPS or BPF is sufficient to decrease basal testosterone secretion by human fetal testes with often nonmonotonic dose-response curves. In fetal mouse testes, the dose-response curves were mostly monotonic and the minimum effective concentrations were 1,000 nmol/L for BPA and BPF and 100 nmol/L for BPS. Finally, 10,000 nmol/L BPA, BPS, or BPF reduced Insl3 expression in cultured mouse fetal testes. This is the first report describing BPS and BPF adverse effects on a physiologic function in humans and rodents. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Human brain factor 1, a new member of the fork head gene family

DOE Office of Scientific and Technical Information (OSTI.GOV)

Murphy, D.B.; Wiese, S.; Burfeind, P.

1994-06-01

Analysis of cDNA clones that cross-hybridized with the fork head domain of the rat HNF-3 gene family revealed 10 cDNAs from human fetal brain and human testis cDNA libraries containing this highly conserved DNA-binding domain. Three of these cDNAs (HFK1, HFK2, and HFK3) were further analyzed. The cDNA HFK1 has a length of 2557 nucleotides and shows strong homology at the nucleotide level (91.2%) to brain factor 1 (BF-1) from rat. The HFK1 cDNA codes for a putative 476 amino acid protein. The homology to BF-1 from rat in the coding region at the amino acid level is 87.5%. Themore » fork head homologous region includes 111 amino acids starting at amino acid 160 and has a 97.5% homology to BF-1. Southern hybridization revealed that HFK1 is highly conserved among mammalian species and possibly birds. Northern analysis with total RNA from human tissues and poly(A)-rich RNA from mouse revealed a 3.2-kb transcript that is present in human and mouse fetal brain and in adult mouse brain. In situ hybridization with sections of mouse embryo and human fetal brain reveals that HFK1 expression is restricted to the neuronal cells in the telencepthalon, with strong expression being observed in the developing dentate gyrus and hippocampus. HFK1 was chromosomally localized by in situ hybridization to 14q12. The cDNA clones HFK2 and HFK3 were analyzed by restriction analysis and sequencing. HFK2 and HFK3 were found to be closely related but different from HFK1. Therefore, it would appear that HFK1, HFK2, HFK3, and BF-1 form a new fork head related subfamily. 33 refs., 6 figs.« less

Rat organic solute carrier protein 1 (rOscp1) mediated the transport of organic solutes in Xenopus laevis oocytes: isolation and pharmacological characterization of rOscp1.

PubMed

Izuno, Hisanori; Kobayashi, Yasuna; Sanada, Yutaka; Nihei, Daisuke; Suzuki, Masako; Kohyama, Noriko; Ohbayashi, Masayuki; Yamamoto, Toshinori

2007-09-22

Rat organic solute carrier protein 1 (rOscp1) was isolated from a rat testis cDNA library. Isolated rOscp1 cDNA consisted of 1089 base pairs that encoded a 363-amino acid protein, and the amino acid sequence was 88% and 93% identical to that of human OSCP1 (hOSCP1) and mouse Oscp1 (mOscp1), respectively. The message for rOscp1 is highly detected in rat testis. When expressed in X. oocytes, rOscp1 mediated the high affinity transport of p-aminohippurate (PAH) with a Km value of 15.7+/-1.9 microM, and rOscp1-mediated organic solutes were exhibited in time- and Na+-independent manners. rOscp1 also transported various structurally heterogenous compounds such as testosterone, dehydroepiandrosterone sulfate (DHEA-S), and taurocholate with some differences in substrate specificity compared with hOSCP1. Immunohistochemical analysis revealed that the rOscp1 protein is localized in the basal membrane side of Sertoli cells as observed in mouse testis [Kobayashi et al., 2007; Kobayashi, Y., Tsuchiya, A., Hayashi, T., Kohyama, N., Ohbayashi, M., Yamamoto, T., 2007. Isolation and characterization of polyspecific mouse organic solute carrier protein 1 (mOscp1). Drug Metabolism and Disposition 35 (7), 1239-1245]. Thus, the present results indicate that a newly isolated cDNA clone, rOscp1, is a polyspecific organic solute carrier protein with some differences in substrate specificity compared with human and mouse OSCP1.

Naloxegol, an opioid antagonist with reduced CNS penetration: Mode-of-action and human relevance for rat testicular tumours.

PubMed

Andersson, Håkan; Mitchard, Terri; Johnson, Nakpangi; Floettmann, Eike

2017-08-15

Naloxegol is an opioid antagonist which has been developed for the treatment of patients with opioid induced constipation. In the nonclinical safety program naloxegol was shown to have a very benign toxicity profile. In the rat, but not the mouse, 2-year carcinogenicity study a change in tumour pattern with an increase in testicular Leydig cell tumours (LCT) was observed after dosing at high (supra-pharmacological) concentrations. To establish the basis of the increase in LCT and to assess its potential relevance to humans, studies to exclude and potentially identify mode-of-action (MoA) were performed. A genotoxic mechanism was ruled out following negative results in the Ames, mouse lymphoma, and micronucleus assays. An effect on androgen metabolism was excluded since the treatment of rats with naloxegol for 14days did not result in any induction of CYP protein levels. It was demonstrated that administration of centrally restricted opioid antagonists naloxegol or methylnaltrexone at high doses induced an increase in LH release with no clear increase in testosterone, in contrast to the centrally acting opioid antagonist naloxone, which showed marked increases in both LH and testosterone. LCT due to increased LH stimulation is common in rats but not documented in humans. Collectively, the lack of genotoxicity signal, the lack of androgen effect, the increase in LH secretion in rats, which is no considered to be relevant for LCT formation in humans, and high margins to clinical exposures, the observed increase in LCT in the rat is not expected to be clinically relevant. Copyright © 2017 Elsevier Inc. All rights reserved.

Characterisation of Cdkl5 transcript isoforms in rat.

PubMed

Hector, Ralph D; Dando, Owen; Ritakari, Tuula E; Kind, Peter C; Bailey, Mark E S; Cobb, Stuart R

2017-03-01

CDKL5 deficiency is a severe neurological disorder caused by mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5). The predominant human CDKL5 brain isoform is a 9.7kb transcript comprised of 18 exons with a large 6.6kb 3'-untranslated region (UTR). Mammalian models of CDKL5 disorder are currently limited to mouse, and little is known about Cdkl5 in other organisms used to model neurodevelopmental disorders, such as rat. In this study we characterise, both bioinformatically and experimentally, the rat Cdkl5 gene structure and its associated transcript isoforms. New exonic regions, splice sites and UTRs are described, confirming the presence of four distinct transcript isoforms. The predominant isoform in the brain, which we name rCdkl5_1, is orthologous to the human hCDKL5_1 and mouse mCdkl5_1 isoforms and is the most highly expressed isoform across all brain regions tested. This updated gene model of Cdkl5 in rat provides a framework for studies into its protein products and provides a reference for the development of molecular therapies for testing in rat models of CDKL5 disorder. Copyright © 2016 Elsevier B.V. All rights reserved.

Recent mouse and rat methods for the study of experimental oral candidiasis.

PubMed

Costa, Anna C B P; Pereira, Cristiane A; Junqueira, Juliana C; Jorge, Antonio O C

2013-07-01

The Candida genus expresses virulence factors that, when combined with immunosuppression and other risk factors, can cause different manifestations of oral candidiasis. The treatment of mucosal infections caused by Candida and the elucidation of the disease process have proven challenging. Therefore, the study of experimentally induced oral candidiasis in rats and mice is useful to clarify the etiopathology of this condition, improve diagnosis, and search for new therapeutic options because the disease process in these animals is similar to that of human candidiasis lesions. Here, we describe and discuss new studies involving rat and mouse models of oral candidiasis with respect to methods for inducing experimental infection, methods for evaluating the development of experimental candidiasis, and new treatment strategies for oral candidiasis.

Comparison of animal discs used in disc research to human lumbar disc: torsion mechanics and collagen content.

PubMed

Showalter, Brent L; Beckstein, Jesse C; Martin, John T; Beattie, Elizabeth E; Espinoza Orías, Alejandro A; Schaer, Thomas P; Vresilovic, Edward J; Elliott, Dawn M

2012-07-01

Experimental measurement and normalization of in vitro disc torsion mechanics and collagen content for several animal species used in intervertebral disc research and comparing these with the human disc. To aid in the selection of appropriate animal models for disc research by measuring torsional mechanical properties and collagen content. There is lack of data and variability in testing protocols for comparing animal and human disc torsion mechanics and collagen content. Intervertebral disc torsion mechanics were measured and normalized by disc height and polar moment of inertia for 11 disc types in 8 mammalian species: the calf, pig, baboon, goat, sheep, rabbit, rat, and mouse lumbar discs, and cow, rat, and mouse caudal discs. Collagen content was measured and normalized by dry weight for the same discs except the rat and the mouse. Collagen fiber stretch in torsion was calculated using an analytical model. Measured torsion parameters varied by several orders of magnitude across the different species. After geometric normalization, only the sheep and pig discs were statistically different from human discs. Fiber stretch was found to be highly dependent on the assumed initial fiber angle. The collagen content of the discs was similar, especially in the outer annulus where only the calf and goat discs were statistically different from human. Disc collagen content did not correlate with torsion mechanics. Disc torsion mechanics are comparable with human lumbar discs in 9 of 11 disc types after normalization by geometry. The normalized torsion mechanics and collagen content of the multiple animal discs presented are useful for selecting and interpreting results for animal disc models. Structural organization of the fiber angle may explain the differences that were noted between species after geometric normalization.

Comparison of Animal Discs Used in Disc Research to Human Lumbar Disc: Torsion Mechanics and Collagen Content

PubMed Central

Showalter, Brent L.; Beckstein, Jesse C.; Martin, John T.; Beattie, Elizabeth E.; Orías, Alejandro A. Espinoza; Schaer, Thomas P.; Vresilovic, Edward J.; Elliott, Dawn M.

2012-01-01

Study Design Experimental measurement and normalization of in vitro disc torsion mechanics and collagen content for several animal species used in intervertebral disc research and comparing these to the human disc. Objective To aid in the selection of appropriate animal models for disc research by measuring torsional mechanical properties and collagen content. Summary of Background Data There is lack of data and variability in testing protocols for comparing animal and human disc torsion mechanics and collagen content. Methods Intervertebral disc torsion mechanics were measured and normalized by disc height and polar moment of inertia for 11 disc types in 8 mammalian species: the calf, pig, baboon, goat, sheep, rabbit, rat, and mouse lumbar, and cow, rat, and mouse caudal. Collagen content was measured and normalized by dry weight for the same discs except the rat and mouse. Collagen fiber stretch in torsion was calculated using an analytical model. Results Measured torsion parameters varied by several orders of magnitude across the different species. After geometric normalization, only the sheep and pig discs were statistically different from human. Fiber stretch was found to be highly dependent on the assumed initial fiber angle. The collagen content of the discs was similar, especially in the outer annulus where only the calf and goat discs were statistically different from human. Disc collagen content did not correlate with torsion mechanics. Conclusion Disc torsion mechanics are comparable to human lumbar discs in 9 of 11 disc types after normalization by geometry. The normalized torsion mechanics and collagen content of the multiple animal discs presented is useful for selecting and interpreting results for animal models of the disc. Structural composition of the disc, such as initial fiber angle, may explain the differences that were noted between species after geometric normalization. PMID:22333953

Evaluation of the adequacy of published studies of low-dose effects of bisphenol A on the rodent prostate for use in human risk assessment.

PubMed

Milman, Harry A; Bosland, Maarten C; Walden, Paul D; Heinze, John E

2002-06-01

Studies conducted in our laboratories and by others found no consistent correlation between prostate size, prostate pathology, or the development of prostate cancer under a variety of experimental conditions. Furthermore, an evaluation of eight published studies that were conducted in mice and rats following in utero exposure by oral treatment of dams with low levels of bisphenol A (BPA) and that focused on the prostate identified several discrepancies that affect their adequacy for use in human risk assessment. For example, there was inadequate reporting of the purity of BPA and the animal supplier used, and housing of offspring was not the same among the studies. In addition, there were differences between studies with mice and rats in exposure regimen, route of exposure, and numbers of dams or pups used per BPA dose group. Poor inter- and intraspecies correlation (i.e., mouse to rat or between mouse or rat strains) further complicates the ability to use results from these studies to predict potential prostate effects in humans. Thus, we conclude that a finding of increased prostate weight in rodent studies with perinatal exposure in the absence of associated pathologic and/or functional changes is meaningless and not indicative of a potential adverse effect in humans. Copyright 2002 Elsevier Science (USA)

Calcium-binding protein from mouse Ehrlich ascites-tumour cells is homologous to human calcyclin.

PubMed Central

Kuźnicki, J; Filipek, A; Hunziker, P E; Huber, S; Heizmann, C W

1989-01-01

A Ca2+-binding protein was purified from mouse Ehrlich ascites-tumour cells. The protein forms monomers and disulphide-linked dimers, which can be separated by reverse-phase h.p.l.c. A partial amino acid sequence analysis demonstrated that the protein has an EF-hand structure. A striking homology was found to rat and human calcyclin (a member of the S-100 protein family), which is possibly involved in cell-cycle regulation. Images Fig. 1. Fig. 2. PMID:2597136

Molecular Biology: Conference on Genetic Engineering Techniques (2nd) Held in London (United Kingdom) on 20-21 November 1986.

DTIC Science & Technology

1987-05-27

system in Chinese t-PA to be a serine protease of 327 amino ovary hamster cells. Precise yields from acids in length. The protein appears, high-level...ham- ster or mouse cell line, allowing the differentiation of human and hamster or ________ mouse clones by hybridization with total human DNA or...appropriate lo- functional protein when transferred into cation downstream of a strong promoter in baby hamster kidney (BHK) cells or rat place of one or

Activation and inhibition of mouse muscle and neuronal nicotinic acetylcholine receptors expressed in Xenopus oocytes.

PubMed

Papke, Roger L; Wecker, Lynn; Stitzel, Jerry A

2010-05-01

Transgenic mouse models with nicotinic acetylcholine receptor (nAChR) knockouts and knockins have provided important insights into the molecular substrates of addiction and disease. However, most studies of heterologously expressed neuronal nAChR have used clones obtained from other species, usually human or rat. In this work, we use mouse clones expressed in Xenopus oocytes to provide a relatively comprehensive characterization of the three primary classes of nAChR: muscle-type receptors, heteromeric neuronal receptors, and homomeric alpha7-type receptors. We evaluated the activation of these receptor subtypes with acetylcholine and cytisine-related compounds, including varenicline. We also characterized the activity of classic nAChR antagonists, confirming the utility of mecamylamine and dihydro-beta-erythroidine as selective antagonists in mouse models of alpha3beta4 and alpha4beta2 receptors, respectively. We also conducted an in-depth analysis of decamethonium and hexamethonium on muscle and neuronal receptor subtypes. Our data indicate that, as with receptors cloned from other species, pairwise expression of neuronal alpha and beta subunits in oocytes generates heterogeneous populations of receptors, most likely caused by variations in subunit stoichiometry. Coexpression of the mouse alpha5 subunit had varying effects, depending on the other subunits expressed. The properties of cytisine-related compounds are similar for mouse, rat, and human nAChR, except that varenicline produced greater residual inhibition of mouse alpha4beta2 receptors than with human receptors. We confirm that decamethonium is a partial agonist, selective for muscle-type receptors, but also note that it is a nondepolarizing antagonist for neuronal-type receptors. Hexamethonium was a relatively nonselective antagonist with mixed competitive and noncompetitive activity.

Activation and Inhibition of Mouse Muscle and Neuronal Nicotinic Acetylcholine Receptors Expressed in Xenopus Oocytes

PubMed Central

Wecker, Lynn; Stitzel, Jerry A.

2010-01-01

Transgenic mouse models with nicotinic acetylcholine receptor (nAChR) knockouts and knockins have provided important insights into the molecular substrates of addiction and disease. However, most studies of heterologously expressed neuronal nAChR have used clones obtained from other species, usually human or rat. In this work, we use mouse clones expressed in Xenopus oocytes to provide a relatively comprehensive characterization of the three primary classes of nAChR: muscle-type receptors, heteromeric neuronal receptors, and homomeric α7-type receptors. We evaluated the activation of these receptor subtypes with acetylcholine and cytisine-related compounds, including varenicline. We also characterized the activity of classic nAChR antagonists, confirming the utility of mecamylamine and dihydro-β-erythroidine as selective antagonists in mouse models of α3β4 and α4β2 receptors, respectively. We also conducted an in-depth analysis of decamethonium and hexamethonium on muscle and neuronal receptor subtypes. Our data indicate that, as with receptors cloned from other species, pairwise expression of neuronal α and β subunits in oocytes generates heterogeneous populations of receptors, most likely caused by variations in subunit stoichiometry. Coexpression of the mouse α5 subunit had varying effects, depending on the other subunits expressed. The properties of cytisine-related compounds are similar for mouse, rat, and human nAChR, except that varenicline produced greater residual inhibition of mouse α4β2 receptors than with human receptors. We confirm that decamethonium is a partial agonist, selective for muscle-type receptors, but also note that it is a nondepolarizing antagonist for neuronal-type receptors. Hexamethonium was a relatively nonselective antagonist with mixed competitive and noncompetitive activity. PMID:20100906

Production and characterization of monoclonal antibodies against conserved epitopes of P-selectin (CD62P).

PubMed

Massaguer, A; Engel, P; Pérez-del-Pulgar, S; Bosch, J; Pizcueta, P

2000-08-01

P-selectin (CD62P) is an adhesion molecule expressed on the activated endothelium and activated platelets that is involved in the initial attachment of leukocytes to inflamed vascular endothelium. Blocking monoclonal antibodies (mAbs) and P-selectin-deficient mice have shown that P-selectin is a potential target in anti-inflammatory therapy. Most mAbs against P-selectin do not bind to conserved epitopes, including the ligand-binding region, since P-selectin from mammalian species shares high amino acid sequence homology. The aim of this study was to generate a novel panel of anti-P-selectin mAbs against the conserved epitopes present in several animal species. To produce these mAbs, P-selectin-deficient mice were immunized with a pre-B-cell line transfected with human P-selectin cDNA. Twelve mouse mAbs that recognize human P-selectin were obtained. Individual mAbs that bound to human, rat, mouse, rabbit and pig activated platelets were characterized by flow-cytometry, immunohistochemistry, adhesion assays and immunoprecipitation. Four of these mAbs (P-sel.KO.2.3, P-sel.KO.2.4, P-sel.KO.2.7 and P-sel.KO.2.12) cross-reacted with human, rat and mouse P-selectin. Another three mAbs (P-sel.KO.2.2, P-sel.KO.2.11 and P-sel.KO.2.12) blocked the attachment of HL60 cells to P-selectin-transfected COS cells, demonstrating that these mAbs inhibit P-selectin-mediated adhesion. MAb cross-blocking experiments showed that these three mAbs bind to very close and overlapping epitopes. An ELISA assay using mAbs P-sel.KO.2.3 and P-sel.KO.2.12 was designed to measure soluble rat, mouse and human P-selectin. These anti-P-selectin mAbs are unique since they recognize common epitopes conserved during mammalian evolution and they may be useful for studying P-selectin function in inflammatory models in various species.

Loss of MeCP2 in the rat models regression, impaired sociability and transcriptional deficits of Rett syndrome

PubMed Central

Veeraragavan, Surabi; Wan, Ying-Wooi; Connolly, Daniel R.; Hamilton, Shannon M.; Ward, Christopher S.; Soriano, Sirena; Pitcher, Meagan R.; McGraw, Christopher M.; Huang, Sharon G.; Green, Jennie R.; Yuva, Lisa A.; Liang, Agnes J.; Neul, Jeffrey L.; Yasui, Dag H.; LaSalle, Janine M.; Liu, Zhandong; Paylor, Richard; Samaco, Rodney C.

2016-01-01

Mouse models of the transcriptional modulator Methyl-CpG-Binding Protein 2 (MeCP2) have advanced our understanding of Rett syndrome (RTT). RTT is a ‘prototypical’ neurodevelopmental disorder with many clinical features overlapping with other intellectual and developmental disabilities (IDD). Therapeutic interventions for RTT may therefore have broader applications. However, the reliance on the laboratory mouse to identify viable therapies for the human condition may present challenges in translating findings from the bench to the clinic. In addition, the need to identify outcome measures in well-chosen animal models is critical for preclinical trials. Here, we report that a novel Mecp2 rat model displays high face validity for modelling psychomotor regression of a learned skill, a deficit that has not been shown in Mecp2 mice. Juvenile play, a behavioural feature that is uniquely present in rats and not mice, is also impaired in female Mecp2 rats. Finally, we demonstrate that evaluating the molecular consequences of the loss of MeCP2 in both mouse and rat may result in higher predictive validity with respect to transcriptional changes in the human RTT brain. These data underscore the similarities and differences caused by the loss of MeCP2 among divergent rodent species which may have important implications for the treatment of individuals with disease-causing MECP2 mutations. Taken together, these findings demonstrate that the Mecp2 rat model is a complementary tool with unique features for the study of RTT and highlight the potential benefit of cross-species analyses in identifying potential disease-relevant preclinical outcome measures. PMID:27365498

Peptidase-3 (Pep-3), dipeptidase variant in the rat homologous to mouse pep-3 (Dip-1) and human PEP-c.

PubMed

Womack, J E; Cramer, D V

1980-10-01

Starch gel electrophoresis and histochemical staining with L-leucyl-L-tyrosine have revealed genetic variation for dipeptidase in Rattus norvegicus. The tissue distribution, substrate specificity, and heterozygous expression as a monmeric protein suggest homology of the variant peptidase to human PEP-C and mouse Pep-3 (Dip-1). We propose Peptidase-3 (Pep-3) as a name for this autosomal locus in the rat. The allele responsible for slower (less anodal) electrophoretic migration is designated Pep-3a and is characteristic of strain ACI/Pit. A faster (more anodal) electrophoretic mobility is the product of the Pep-3b allele in strain F344/Pit. Twenty-five additional inbred strains carry Pep-3a and 16 others carry Pep-3b. Wild rats trapped in Pittsburgh were polymorphic for this locus. Alleles at Pep-3 segregated independently of c (linkage group I), a (linkage group IV), RT2 and Es-1 (linkage group V), h (linkage group VI), and RTI (linkage group VIII).

Inhibitor(s) of the classical complement pathway in mouse serum limit the utility of mice as experimental models of neuromyelitis optica.

PubMed

Ratelade, Julien; Verkman, A S

2014-11-01

Neuromyelitis optica (NMO) is an inflammatory demyelinating disease of the central nervous system in which anti-aquaporin-4 (AQP4) autoantibodies (AQP4-IgG) cause damage to astrocytes by complement-dependent cytotoxicity (CDC). Various approaches have been attempted to produce NMO lesions in rodents, some involving genetically modified mice with altered immune cell function. Here, we found that mouse serum strongly inhibits complement from multiple species, preventing AQP4-IgG-dependent CDC. Effects of mouse serum on complement activation were tested in CDC assays in which AQP4-expressing cells were incubated with AQP4-IgG and complement from different species. Biochemical assays and mass spectrometry were used to characterize complement inhibitor(s) in mouse serum. Sera from different strains of mice produced almost no AQP4-IgG-dependent CDC compared with human, rat and guinea pig sera. Remarkably, addition of mouse serum prevented AQP4-IgG-dependent CDC caused by human, rat or guinea pig serum, with 50% inhibition at <5% mouse serum. Hemolysis assays indicated that the inhibitor(s) in mouse serum target the classical and not the alternative complement pathway. We found that the complement inhibitor(s) in mouse serum were contained in a serum fraction purified with protein-A resin; however, the inhibitor was not IgG as determined using serum from IgG-deficient mice. Mass spectrometry on the protein A-purified fraction produced several inhibitor candidates. The low intrinsic complement activity of mouse serum and the presence of complement inhibitor(s) limit the utility of mouse models to study disorders, such as NMO, involving the classical complement pathway. Copyright © 2014 Elsevier Ltd. All rights reserved.

FORMATION OF HEMOGLOBIN AND ALBUMIN ADDUCTS OF BENZENE OXIDE IN MOUSE, RAT, AND HUMAN BLOOD

EPA Science Inventory

Little is known about the formation and disposition of benzene oxide (BO), the initial metabolite arising from oxidation of benzene by cytochrome P450. In this study, reactions of BO with hemoglobin (Hb) and albumin (Alb) were investigated in blood from B6C3F1 mice, F344 rats, ...

Antitumor effect of novel anti-podoplanin antibody NZ-12 against malignant pleural mesothelioma in an orthotopic xenograft model.

PubMed

Abe, Shinji; Kaneko, Mika Kato; Tsuchihashi, Yuki; Izumi, Toshihiro; Ogasawara, Satoshi; Okada, Naoto; Sato, Chiemi; Tobiume, Makoto; Otsuka, Kenji; Miyamoto, Licht; Tsuchiya, Koichiro; Kawazoe, Kazuyoshi; Kato, Yukinari; Nishioka, Yasuhiko

2016-09-01

Podoplanin (aggrus) is highly expressed in several types of cancers, including malignant pleural mesothelioma (MPM). Previously, we developed a rat anti-human podoplanin mAb, NZ-1, and a rat-human chimeric anti-human podoplanin antibody, NZ-8, derived from NZ-1, which induced antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity against podoplanin-positive MPM cell lines. In this study, we showed the antitumor effect of NZ-1, NZ-8, and NZ-12, a novel rat-human chimeric anti-human podoplanin antibody derived from NZ-1, in an MPM orthotopic xenograft SCID mouse model. Treatment with NZ-1 and rat NK (CD161a(+) ) cells inhibited the growth of tumors and the production of pleural effusion in NCI-H290/PDPN or NCI-H226 orthotopic xenograft mouse models. NZ-8 and human natural killer (NK) (CD56(+) ) cells also inhibited tumor growth and pleural effusion in MPM orthotopic xenograft mice. Furthermore, NZ-12 induced potent ADCC mediated by human MNC, compared with either NZ-1 or NZ-8. Antitumor effects were observed following treatment with NZ-12 and human NK (CD56(+) ) cells in MPM orthotopic xenograft mice. In addition, combined immunotherapy using the ADCC activity of NZ-12 mediated by human NK (CD56(+) ) cells with pemetrexed, led to enhanced antitumor effects in MPM orthotopic xenograft mice. These results strongly suggest that combination therapy with podoplanin-targeting immunotherapy using both NZ-12 and pemetrexed might provide an efficacious therapeutic strategy for the treatment of MPM. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

76 FR 51031 - Registration; Cancellation Order for Rodenticide Products That Have Expired

Federal Register 2010, 2011, 2012, 2013, 2014

2011-08-17

...--Registrations and Product Names EPA Registration No. Product name 47629-14 Difenacoum Rat and Mouse Pellets (consumer use only). 47629-16 Difenacoum Rat and Mouse Block (consumer use only). 47629-17 Difenacoum Rat and Mouse Place Packs (consumer use only). 47629-11 Bromethalin Rat & Mouse Block. 47629-13...

Species Differences in Human and Rodent PEPT2-Mediated Transport of Glycylsarcosine and Cefadroxil in Pichia Pastoris Transformants

PubMed Central

Song, Feifeng; Hu, Yongjun; Jiang, Huidi

2017-01-01

The proton-coupled oligopeptide transporter PEPT2 (SLC15A2) plays an important role in the disposition of di/tripeptides and peptide-like drugs in kidney and brain. However, unlike PEPT1 (SLC15A1), there is little information about species differences in the transport of PEPT2-mediated substrates. The purpose of this study was to determine whether PEPT2 exhibited a species-dependent uptake of glycylsarcosine (GlySar) and cefadroxil using yeast Pichia pastoris cells expressing cDNA from human, mouse, and rat. In such a system, the functional activity of PEPT2 was evaluated with [3H]GlySar as a function of time, pH, substrate concentration, and specificity, and with [3H]cefadroxil as a function of concentration. We observed that the uptake of GlySar was pH-dependent with an optimal uptake at pH 6.5 for all three species. Moreover, GlySar showed saturable uptake kinetics, with Km values in human (150.6 µM) > mouse (42.8 µM) ≈ rat (36.0 µM). The PEPT2-mediated uptake of GlySar in yeast transformants was specific, being inhibited by di/tripeptides and peptide-like drugs, but not by amino acids and nonsubstrate compounds. Cefadroxil also showed a saturable uptake profile in all three species, with Km values in human (150.8 μM) > mouse (15.6 μM) ≈ rat (11.9 μM). These findings demonstrated that the PEPT2-mediated uptake of GlySar and cefadroxil was specific, species dependent, and saturable. Furthermore, based on the Km values, mice appeared similar to rats but both were less than optimal as animal models in evaluating the renal reabsorption and pharmacokinetics of peptides and peptide-like drugs in humans. PMID:27836942

Behavioral Phenotyping Assays for Genetic Mouse Models of Neurodevelopmental, Neurodegenerative, and Psychiatric Disorders.

PubMed

Sukoff Rizzo, Stacey J; Crawley, Jacqueline N

2017-02-08

Animal models offer heuristic research tools to understand the causes of human diseases and to identify potential treatments. With rapidly evolving genetic engineering technologies, mutations identified in a human disorder can be generated in the mouse genome. Phenotypic outcomes of the mutation are then explicated to confirm hypotheses about causes and to discover effective therapeutics. Most neurodevelopmental, neurodegenerative, and psychiatric disorders are diagnosed primarily by their prominent behavioral symptoms. Mouse behavioral assays analogous to the human symptoms have been developed to analyze the consequences of mutations and to evaluate proposed therapeutics preclinically. Here we describe the range of mouse behavioral tests available in the established behavioral neuroscience literature, along with examples of their translational applications. Concepts presented have been successfully used in other species, including flies, worms, fish, rats, pigs, and nonhuman primates. Identical strategies can be employed to test hypotheses about environmental causes and gene × environment interactions.

Recent mouse and rat methods for the study of experimental oral candidiasis

PubMed Central

Costa, Anna CBP; Pereira, Cristiane A; Junqueira, Juliana C; Jorge, Antonio OC

2013-01-01

The Candida genus expresses virulence factors that, when combined with immunosuppression and other risk factors, can cause different manifestations of oral candidiasis. The treatment of mucosal infections caused by Candida and the elucidation of the disease process have proven challenging. Therefore, the study of experimentally induced oral candidiasis in rats and mice is useful to clarify the etiopathology of this condition, improve diagnosis, and search for new therapeutic options because the disease process in these animals is similar to that of human candidiasis lesions. Here, we describe and discuss new studies involving rat and mouse models of oral candidiasis with respect to methods for inducing experimental infection, methods for evaluating the development of experimental candidiasis, and new treatment strategies for oral candidiasis. PMID:23715031

In vitro metabolism of a novel JNK inhibitor tanzisertib: interspecies differences in oxido-reduction and characterization of enzymes involved in metabolism.

PubMed

Atsriku, Christian; Hoffmann, Matthew; Moghaddam, Mehran; Kumar, Gondi; Surapaneni, Sekhar

2015-01-01

1. In vitro metabolism of Tanzisertib [(1S,4R)-4-(9-((S)tetrahydrofuran-3-yl)-8-(2,4,6-trifluorophenylamino)-9H-purin-2-ylamino) cyclohexanol], a potent, selective c-Jun amino-terminal kinase (JNK) inhibitor, was investigated in mouse, rat, rabbit, dog, monkey and human hepatocytes over 4 h. The extent of metabolism of [(14)C]tanzisertib was variable, with <10% metabolized in dog and human, <20% metabolized in rabbit and monkey and >75% metabolized in rat and mouse. Primary metabolic pathways in human and dog hepatocytes, were direct glucuronidation and oxidation of cyclohexanol to a keto metabolite, which was subsequently reduced to parent or cis-isomer, followed by glucuronidation. Rat and mouse produced oxidative metabolites and cis-isomer, including direct glucuronides and sulfates of tanzisertib and cis-isomer. 2. Enzymology of oxido-reductive pathways revealed that human aldo-keto reductases AKR1C1, 1C2, 1C3 and 1C4 were responsible for oxido-reduction of tanzisertib, CC-418424 and keto tanzisertib. Characterizations of enzyme kinetics revealed that AKR1C4 had a high affinity for reduction of keto tanzisertib to tanzisertib compared to other isoforms. These results demonstrate unique stereoselectivity of the reductive properties documented by human AKR1C enzymes for the same substrate. 3. Characterization of UGT isoenzymes in glucuronidation of tanzisertib and CC-418424 revealed that, tanzisertib glucuronide was catalyzed by: UGT1A1, 1A4, 1A10 and 2B4, while CC-418424 glucuronidation was catalyzed by UGT2B4 and 2B7.

Cross-species pharmacokinetic comparison from mouse to man of a second-generation antisense oligonucleotide, ISIS 301012, targeting human apolipoprotein B-100.

PubMed

Yu, Rosie Z; Kim, Tae-Won; Hong, An; Watanabe, Tanya A; Gaus, Hans J; Geary, Richard S

2007-03-01

The pharmacokinetics of a 2'-O-(2-methoxyethyl)-modified oligonucleotide, ISIS 301012 [targeting human apolipoprotein B-100 (apoB-100)], was characterized in mouse, rat, monkey, and human. Plasma pharmacokinetics following parental administration was similar across species, exhibiting a rapid distribution phase with t(1/2alpha) of several hours and a prolonged elimination phase with t(1/2beta) of days. The prolonged elimination phase represents equilibrium between tissues and circulating drug due to slow elimination from tissues. Absorption was nearly complete following s.c. injection, with bioavailability ranging from 80 to 100% in monkeys. Plasma clearance scaled well across species as a function of body weight alone, and this correlation was improved when corrected for plasma protein binding. In all of the animal models studied, the highest tissue concentrations of ISIS 301012 were observed in kidney and liver. Urinary excretion was less than 3% in monkeys and human in the first 24 h. ISIS 301012 is highly bound to plasma proteins, probably preventing rapid removal by renal filtration. However, following 25 mg/kg s.c. administration in mouse and 5-mg/kg i.v. bolus administration in rat, plasma concentrations of ISIS 301012 exceeded their respective protein binding capacity. Thus, urinary excretion increased to 16% or greater within the first 24 h. Albeit slow, urinary excretion of ISIS 301012 and its shortened metabolites is the ultimate elimination pathway of this compound, as demonstrated by 32% of dose recovered in total excreta by 14 days in a rat mass balance study. The pharmacokinetics of ISIS 301012 in human is predictable from the pharmacokinetics measured in animals. The pharmacokinetic properties of ISIS 301012 provide guidance for clinical development and support infrequent dose administration.

Ultrasonic vocalizations: a tool for behavioural phenotyping of mouse models of neurodevelopmental disorders

PubMed Central

Scattoni, Maria Luisa; Crawley, Jacqueline; Ricceri, Laura

2009-01-01

In neonatal mice ultrasonic vocalizations have been studied both as an early communicative behavior of the pup-mother dyad and as a sign of an aversive affective state. Adult mice of both sexes produce complex ultrasonic vocalization patterns in different experimental/social contexts. All these vocalizations are becoming an increasingly valuable assay for behavioral phenotyping throughout the mouse life-span and alterations of the ultrasound patterns have been reported in several mouse models of neurodevelopmental disorders. Here we also show that the modulation of vocalizations by maternal cues (maternal potentiation paradigm) – originally identified and investigated in rats - can be measured in C57Bl/6 mouse pups with appropriate modifications of the rat protocol and can likely be applied to mouse behavioral phenotyping. In addition we suggest that a detailed qualitative evaluation of neonatal calls together with analysis of adult mouse vocalization patterns in both sexes in social settings, may lead to a greater understanding of the communication value of vocalizations in mice. Importantly, both neonatal and adult USV altered patterns can be determined during the behavioural phenotyping of mouse models of human neurodevelopmental and neuropsychiatric disorders, starting from those in which deficits in communication are a primary symptom. PMID:18771687

Increased trabecular bone and improved biomechanics in an osteocalcin-null rat model created by CRISPR/Cas9 technology.

PubMed

Lambert, Laura J; Challa, Anil K; Niu, Aidi; Zhou, Lihua; Tucholski, Janusz; Johnson, Maria S; Nagy, Tim R; Eberhardt, Alan W; Estep, Patrick N; Kesterson, Robert A; Grams, Jayleen M

2016-10-01

Osteocalcin, also known as bone γ-carboxyglutamate protein (Bglap), is expressed by osteoblasts and is commonly used as a clinical marker of bone turnover. A mouse model of osteocalcin deficiency has implicated osteocalcin as a mediator of changes to the skeleton, endocrine system, reproductive organs and central nervous system. However, differences between mouse and human osteocalcin at both the genome and protein levels have challenged the validity of extrapolating findings from the osteocalcin-deficient mouse model to human disease. The rat osteocalcin (Bglap) gene locus shares greater synteny with that of humans. To further examine the role of osteocalcin in disease, we created a rat model with complete loss of osteocalcin using the CRISPR/Cas9 system. Rat osteocalcin was modified by injection of CRISPR/Cas9 mRNA into the pronuclei of fertilized single cell Sprague-Dawley embryos, and animals were bred to homozygosity and compound heterozygosity for the mutant alleles. Dual-energy X-ray absorptiometry (DXA), glucose tolerance testing (GTT), insulin tolerance testing (ITT), microcomputed tomography (µCT), and a three-point break biomechanical assay were performed on the excised femurs at 5 months of age. Complete loss of osteocalcin resulted in bones with significantly increased trabecular thickness, density and volume. Cortical bone volume and density were not increased in null animals. The bones had improved functional quality as evidenced by an increase in failure load during the biomechanical stress assay. Differences in glucose homeostasis were observed between groups, but there were no differences in body weight or composition. This rat model of complete loss of osteocalcin provides a platform for further understanding the role of osteocalcin in disease, and it is a novel model of increased bone formation with potential utility in osteoporosis and osteoarthritis research. © 2016. Published by The Company of Biologists Ltd.

Inhibition of bile salt transport by drugs associated with liver injury in primary hepatocytes from human, monkey, dog, rat, and mouse

PubMed Central

Zhang, Jie; He, Kan; Cai, Lining; Chen, Yu-Chuan; Yang, Yifan; Shi, Qin; Woolf, Thomas F.; Ge, Weigong; Guo, Lei; Borlak, Jürgen; Tong, Weida

2018-01-01

Interference of bile salt transport is one of the underlying mechanisms for drug-induced liver injury (DILI). We developed a novel bile salt transport activity assay involving in situ biosynthesis of bile salts from their precursors in primary human, monkey, dog, rat, and mouse hepatocytes in suspension as well as LC-MS/MS determination of extracellular bile salts transported out of hepatocytes. Glycine- and taurine-conjugated bile acids were rapidly formed in hepatocytes and effectively transported into the extracellular medium. The bile salt formation and transport activities were time– and bile-acid-concentration–dependent in primary human hepatocytes. The transport activity was inhibited by the bile salt export pump (BSEP) inhibitors ketoconazole, saquinavir, cyclosporine, and troglitazone. The assay was used to test 86 drugs for their potential to inhibit bile salt transport activity in human hepatocytes, which included 35 drugs associated with severe DILI (sDILI) and 51 with non-severe DILI (non-sDILI). Approximately 60% of the sDILI drugs showed potent inhibition (with IC50 values <50 μM), but only about 20% of the non-sDILI drugs showed this strength of inhibition in primary human hepatocytes and these drugs are associated only with cholestatic and mixed hepatocellular cholestatic (mixed) injuries. The sDILI drugs, which did not show substantial inhibition of bile salt transport activity, are likely to be associated with immune-mediated liver injury. Twenty-four drugs were also tested in monkey, dog, rat and mouse hepatocytes. Species differences in potency were observed with mouse being less sensitive than other species to inhibition of bile salt transport. In summary, a novel assay has been developed using hepatocytes in suspension from human and animal species that can be used to assess the potential for drugs and/or drug-derived metabolites to inhibit bile salt transport and/or formation activity. Drugs causing sDILI, except those by immune-mediated mechanism, are highly associated with potent inhibition of bile salt transport. PMID:27000539

Inhibition of bile salt transport by drugs associated with liver injury in primary hepatocytes from human, monkey, dog, rat, and mouse.

PubMed

Zhang, Jie; He, Kan; Cai, Lining; Chen, Yu-Chuan; Yang, Yifan; Shi, Qin; Woolf, Thomas F; Ge, Weigong; Guo, Lei; Borlak, Jürgen; Tong, Weida

2016-08-05

Interference of bile salt transport is one of the underlying mechanisms for drug-induced liver injury (DILI). We developed a novel bile salt transport activity assay involving in situ biosynthesis of bile salts from their precursors in primary human, monkey, dog, rat, and mouse hepatocytes in suspension as well as LC-MS/MS determination of extracellular bile salts transported out of hepatocytes. Glycine- and taurine-conjugated bile acids were rapidly formed in hepatocytes and effectively transported into the extracellular medium. The bile salt formation and transport activities were time‒ and bile-acid-concentration‒dependent in primary human hepatocytes. The transport activity was inhibited by the bile salt export pump (BSEP) inhibitors ketoconazole, saquinavir, cyclosporine, and troglitazone. The assay was used to test 86 drugs for their potential to inhibit bile salt transport activity in human hepatocytes, which included 35 drugs associated with severe DILI (sDILI) and 51 with non-severe DILI (non-sDILI). Approximately 60% of the sDILI drugs showed potent inhibition (with IC50 values <50 μM), but only about 20% of the non-sDILI drugs showed this strength of inhibition in primary human hepatocytes and these drugs are associated only with cholestatic and mixed hepatocellular cholestatic (mixed) injuries. The sDILI drugs, which did not show substantial inhibition of bile salt transport activity, are likely to be associated with immune-mediated liver injury. Twenty-four drugs were also tested in monkey, dog, rat and mouse hepatocytes. Species differences in potency were observed with mouse being less sensitive than other species to inhibition of bile salt transport. In summary, a novel assay has been developed using hepatocytes in suspension from human and animal species that can be used to assess the potential for drugs and/or drug-derived metabolites to inhibit bile salt transport and/or formation activity. Drugs causing sDILI, except those by immune-mediated mechanism, are highly associated with potent inhibition of bile salt transport. Published by Elsevier Ireland Ltd.

Effect of Culture Conditions on Viability of Mouse and Rat Embryos Developed in Vitro

PubMed Central

Popova, Elena; Bader, Michael; Krivokharchenko, Alexander

2011-01-01

Currently in vitro culture of mouse preimplantation embryos has become a very important technique to investigate different mechanisms of early embryogenesis. However, there is a big difference in the preimplantation development between mammalian species. Despite close relatedness to mice, in vitro cultivation of rat preimplantation embryos is still delicate and needs further investigation and optimizations. In this study we have compared the in vitro developmental potential of mouse and rat embryos cultured at different culture conditions in parallel experiments. Interestingly, mouse zygotes developed in vitro until blastocyst stage even in inadequate medium without any phosphates and with low osmolarity which was formulated especially for cultivation of rat embryos. Rat parthenotes and zygotes developed in M16 medium formulated for mouse embryos only till 2-cell stage and further development is blocked completely at this stage. Moreover, developmental ability of rat embryos in vitro was significantly lower in comparison with mouse even in special rat mR1ECM medium. Mouse and rat embryos at 2-cell stage obtained in vivo developed until blastocyst stages significantly more efficiently compared to zygotes. Culture of mouse zygotes in glass capillaries resulted in a significantly higher rate of morula and blastocyst development compared with dishes. The Well-of-the-Well system resulted in a significant improvement when compared with dishes for the culture of rat zygotes only until morula stage. Reduced oxygen tension increased the developmental rate of rat but not mouse zygotes until blastocyst stage. This study demonstrates that development of early preimplantation embryos is altered by different culture conditions and show strong differences even between two related species such as mice and rats. Therefore, for understanding the fundamental mechanisms of early mammalian development it is very important to use embryos of various species. PMID:24710194

Cycloheximide: No Ordinary Bitter Stimulus

PubMed Central

Hettinger, Thomas P.; Formaker, Bradley K.; Frank, Marion E.

2007-01-01

Cycloheximide (CyX), a toxic antibiotic with a unique chemical structure generated by the actinomycete, Streptomyces griseus, has emerged as a primary focus of studies on mammalian bitter taste. Rats and mice avoid it at concentrations well below the thresholds for most bitter stimuli and T2R G-protein-coupled receptors specific for CyX with appropriate sensitivity are identified for those species. Like mouse and rat, golden hamsters, Mesocricetus auratus, also detected and rejected micromolar levels of CyX, although 1 mM CyX failed to activate the hamster chorda tympani nerve. Hamsters showed an initial tolerance for 500 μM CyX, but after that, avoidance of CyX dramatically increased, plasticity not reported for rat or mouse. As the hamster lineage branches well before division of the mouse-rat lineage in evolutionary time, differences between hamster and mouse-rat reactions to CyX are not surprising. Furthermore, unlike hamster LiCl-induced learned aversions, the induced CyX aversion neither specifically nor robustly generalized to other non-ionic bitter stimuli; and unlike adverse reactions to other chemosensory stimuli, aversions to CyX were not mollified by adding a sweetener. Thus, CyX is unlike other bitter stimuli. The gene for the high-affinity CyX receptor is a member of a cluster of 5 orthologous T2R genes that are likely rodent specific; this “CyX clade” is found in the mouse, rat and probably hamster, but not in the human or rabbit genome. The rodent CyX-T2R interaction may be one of multiple lineage-specific stimulus-receptor interactions reflecting a response to a particular environmental toxin. The combination of T2R multiplicity, species divergence and gene duplication results in diverse ligands for multiple species-specific T2R receptors, which confounds definition of ‘bitter’ stimuli across species. PMID:17400304

Expression of acid-sensing ion channels and selection of reference genes in mouse and naked mole rat.

PubMed

Schuhmacher, Laura-Nadine; Smith, Ewan St John

2016-12-13

Acid-sensing ion channels (ASICs) are a family of ion channels comprised of six subunits encoded by four genes and they are expressed throughout the peripheral and central nervous systems. ASICs have been implicated in a wide range of physiological and pathophysiological processes: pain, breathing, synaptic plasticity and excitotoxicity. Unlike mice and humans, naked mole-rats do not perceive acid as a noxious stimulus, even though their sensory neurons express functional ASICs, likely an adaptation to living in a hypercapnic subterranean environment. Previous studies of ASIC expression in the mammalian nervous system have often not examined all subunits, or have failed to adequately quantify expression between tissues; to date there has been no attempt to determine ASIC expression in the central nervous system of the naked mole-rat. Here we perform a geNorm study to identify reliable housekeeping genes in both mouse and naked mole-rat and then use quantitative real-time PCR to estimate the relative amounts of ASIC transcripts in different tissues of both species. We identify RPL13A (ribosomal protein L13A) and CANX (calnexin), and β-ACTIN and EIF4A (eukaryotic initiation factor 4a) as being the most stably expressed housekeeping genes in mouse and naked mole-rat, respectively. In both species, ASIC3 was most highly expressed in dorsal root ganglia (DRG), and ASIC1a, ASIC2b and ASIC3 were more highly expressed across all brain regions compared to the other subunits. We also show that ASIC4, a proton-insensitive subunit of relatively unknown function, was highly expressed in all mouse tissues apart from DRG and hippocampus, but was by contrast the lowliest expressed ASIC in all naked mole-rat tissues.

Architectural assessment of rhesus macaque pelvic floor muscles: comparison for use as a human model.

PubMed

Stewart, Amanda M; Cook, Mark S; Esparza, Mary C; Slayden, Ov D; Alperin, Marianna

2017-10-01

Animal models are essential to further our understanding of the independent and combined function of human pelvic floor muscles (PFMs), as direct studies in women are limited. To assure suitability of the rhesus macaque (RM), we compared RM and human PFM architecture, the strongest predictor of muscle function. We hypothesized that relative to other models, RM best resembles human PFM. Major architectural parameters of cadaveric human coccygeus, iliococcygeus, and pubovisceralis (pubococcygeus + puborectalis) and corresponding RM coccygeus, iliocaudalis, and pubovisceralis (pubovaginalis + pubocaudalis) were compared using 1- and 2-way analysis of variance (ANOVA) with post hoc testing. Architectural difference index (ADI), a combined measure of functionally relevant structural parameters predictive of length-tension, force-generation, and excursional muscle properties was used to compare PFMs across RM, rabbit, rat, and mouse. RM and human PFMs were similar with respect to architecture. However, the magnitude of similarity varied between individual muscles, with the architecture of the most distinct RM PFM, iliocaudalis, being well suited for quadrupedal locomotion. Except for the pubovaginalis, RM PFMs inserted onto caudal vertebrae, analogous to all tailed animals. Comparison of the PFM complex architecture across species revealed the lowest, thus closest to human, ADI for RM (1.9), followed by rat (2.0), mouse (2.6), and rabbit (4.7). Overall, RM provides the closest architectural representation of human PFM complex among species examined; however, differences between individual PFMs should be taken into consideration. As RM is closely followed by rat with respect to PFM similarity with humans, this less-sentient and substantially cheaper model is a good alternative for PFM studies.

Cellular Localization of Aquaporin-1 in the Human and Mouse Trigeminal Systems

PubMed Central

Gu, Minxia; Marshall, Charles; Ding, Jiong; Hu, Gang; Xiao, Ming

2012-01-01

Previous studies reported that a subpopulation of mouse and rat trigeminal neurons express water channel aquaporin-1 (AQP1). In this study we make a comparative investigation of AQP1 localization in the human and mouse trigeminal systems. Immunohistochemistry and immunofluorescence results showed that AQP1 was localized to the cytoplasm and cell membrane of some medium and small-sized trigeminal neurons. Additionally, AQP1 was found in numerous peripheral trigeminal axons of humans and mice. In the central trigeminal root and brain stem, AQP1 was specifically expressed in astrocytes of humans, but was restricted to nerve fibers within the central trigeminal root and spinal trigeminal tract and nucleus in mice. Furthermore, AQP1 positive nerve fibers were present in the mucosal and submucosal layers of human and mouse oral tissues, but not in the muscular and subcutaneous layers. Fluorogold retrograde tracing demonstrated that AQP1 positive trigeminal neurons innervate the mucosa but not skin of cheek. These results reveal there are similarities and differences in the cellular localization of AQP1 between the human and mouse trigeminal systems. Selective expression of AQP1 in the trigeminal neurons innervating the oral mucosa indicates an involvement of AQP1 in oral sensory transduction. PMID:23029502

Clinical significance of the correlation between PLCE 1 and PRKCA in esophageal inflammation and esophageal carcinoma

PubMed Central

Ma, Ming; Zhang, Shanshan; Zhang, Yongmeng; Yuan, Ming; Liu, Bing; Yang, Yiqiong; Cui, Wen; Ansong, Emmanuel; Dong, Huali; Macias, Virgilia; Yang, Wancai

2017-01-01

Esophagitis and Barrett's esophagus are linked to esophageal squamous cell carcinoma and adenocarcinoma, respectively. However, the underlying mechanisms are still unclear. This study analyzed the expression levels of and correlation between PLCE1 and PRKCA in human esophagitis, carcinogen NMBA-induced rat esophagus, PLCE1 genetic deficient mouse esophageal epithelial tissues and human esophageal cancer cell line, integrated with Online oncology data sets. We found that the expression levels of both PLCE1 and PRKCA were significantly elevated in human esophagitis, esophageal squamous cell carcinoma, Barrett's esophagus, esophageal adenocarcinoma and in NMBA-treated rat esophageal epithelia. However, PRKCA and cytokines were significantly downregulated in PLCE1-deficient mouse esophageal epithelia, and knockdown of PLCE1 in human esophageal cancer cells led to reduction of PRKCA and cytokines. Finally, high expression of both PLCE1 and PRKCA is significantly associated with poor outcomes of the patients with esophageal cancers. In conclusion, this study defined the initiation and progression of esophageal inflammation and malignant transformation, in which the positive correlation of PLCE1 and PRKCA exhibits critical clinical significance. PMID:28402280

Liver natural killer cells: subsets and roles in liver immunity

PubMed Central

Peng, Hui; Wisse, Eddie; Tian, Zhigang

2016-01-01

The liver represents a frontline immune organ that is constantly exposed to a variety of gut-derived antigens as a result of its unique location and blood supply. With a predominant role in innate immunity, the liver is enriched with various innate immune cells, among which natural killer (NK) cells play important roles in host defense and in maintaining immune balance. Hepatic NK cells were first described as ‘pit cells' in the rat liver in the 1970s. Recent studies of NK cells in mouse and human livers have shown that two distinct NK cell subsets, liver-resident NK cells and conventional NK (cNK) cells, are present in this organ. Here, we review liver NK cell subsets in different species, revisiting rat hepatic pit cells and highlighting recent progress related to resident NK cells in mouse and human livers, and also discuss the dual roles of NK cells in liver immunity. PMID:26639736

RatMap--rat genome tools and data.

PubMed

Petersen, Greta; Johnson, Per; Andersson, Lars; Klinga-Levan, Karin; Gómez-Fabre, Pedro M; Ståhl, Fredrik

2005-01-01

The rat genome database RatMap (http://ratmap.org or http://ratmap.gen.gu.se) has been one of the main resources for rat genome information since 1994. The database is maintained by CMB-Genetics at Goteborg University in Sweden and provides information on rat genes, polymorphic rat DNA-markers and rat quantitative trait loci (QTLs), all curated at RatMap. The database is under the supervision of the Rat Gene and Nomenclature Committee (RGNC); thus much attention is paid to rat gene nomenclature. RatMap presents information on rat idiograms, karyotypes and provides a unified presentation of the rat genome sequence and integrated rat linkage maps. A set of tools is also available to facilitate the identification and characterization of rat QTLs, as well as the estimation of exon/intron number and sizes in individual rat genes. Furthermore, comparative gene maps of rat in regard to mouse and human are provided.

RatMap—rat genome tools and data

PubMed Central

Petersen, Greta; Johnson, Per; Andersson, Lars; Klinga-Levan, Karin; Gómez-Fabre, Pedro M.; Ståhl, Fredrik

2005-01-01

The rat genome database RatMap (http://ratmap.org or http://ratmap.gen.gu.se) has been one of the main resources for rat genome information since 1994. The database is maintained by CMB–Genetics at Göteborg University in Sweden and provides information on rat genes, polymorphic rat DNA-markers and rat quantitative trait loci (QTLs), all curated at RatMap. The database is under the supervision of the Rat Gene and Nomenclature Committee (RGNC); thus much attention is paid to rat gene nomenclature. RatMap presents information on rat idiograms, karyotypes and provides a unified presentation of the rat genome sequence and integrated rat linkage maps. A set of tools is also available to facilitate the identification and characterization of rat QTLs, as well as the estimation of exon/intron number and sizes in individual rat genes. Furthermore, comparative gene maps of rat in regard to mouse and human are provided. PMID:15608244

Rat astrocytes are more supportive for mouse OPC self-renewal than mouse astrocytes in culture.

PubMed

Cheng, Xuejun; Xie, Binghua; Qi, Jiajun; Zhao, Xiaofeng; Zhang, Zunyi; Qiu, Mengsheng; Yang, Junlin

2017-09-01

Mouse primary oligodendrocyte precursor cells (OPCs) are increasingly used to study the molecular mechanisms underlying the phenotype changes in oligodendrocyte differentiation and axonal myelination observed in transgenic or mutant mouse models. However, mouse OPCs are much more difficult to be isolated by the simple dissociation culture of brain tissues than their rat counterparts. To date, the mechanisms underlying the species difference in OPC preparation remain obscure. In this study, we showed that astrocytes from rats have a stronger effect than those from mouse in promoting OPC proliferation and survival in vitro. Mouse astrocytes displayed significantly weaker viability in culture and reduced potential in maintaining OPC self-renewal, as confirmed by culturing OPCs with conditioned media from rat or mouse astrocytes. These results explained the reason for why stratified cultures of OPCs and astrocytes are difficult to be achieved in mouse CNS tissues. Based on these findings, we adopted inactivated rat astrocytes as feeder cells to support the self-renewal of mouse cortical OPCs and preparation of high-purity mouse OPCs. © 2016 Wiley Periodicals, Inc. Develop Neurobiol 77: 907-916, 2017. © 2016 Wiley Periodicals, Inc.

A preliminary regional PBPK model of lung metabolism for improving species dependent descriptions of 1,3-butadiene and its metabolites.

PubMed

Campbell, Jerry; Van Landingham, Cynthia; Crowell, Susan; Gentry, Robinan; Kaden, Debra; Fiebelkorn, Stacy; Loccisano, Anne; Clewell, Harvey

2015-08-05

1,3-Butadiene (BD), a volatile organic chemical (VOC), is used in synthetic rubber production and other industrial processes. It is detectable at low levels in ambient air as well as in tobacco smoke and gasoline vapors. Inhalation exposures to high concentrations of BD have been associated with lung cancer in both humans and experimental animals, although differences in species sensitivity have been observed. Metabolically active lung cells such as Pulmonary Type I and Type II epithelial cells and club cells (Clara cells)(1) are potential targets of BD metabolite-induced toxicity. Metabolic capacities of these cells, their regional densities, and distributions vary throughout the respiratory tract as well as between species and cell types. Here we present a physiologically based pharmacokinetic (PBPK) model for BD that includes a regional model of lung metabolism, based on a previous model for styrene, to provide species-dependent descriptions of BD metabolism in the mouse, rat, and human. Since there are no in vivo data on BD pharmacokinetics in the human, the rat and mouse models were parameterized to the extent possible on the basis of in vitro metabolic data. Where it was necessary to use in vivo data, extrapolation from rat to mouse was performed to evaluate the level of uncertainty in the human model. A kidney compartment and description of downstream metabolism were also included in the model to allow for eventual use of available urinary and blood biomarker data in animals and humans to calibrate the model for estimation of BD exposures and internal metabolite levels. Results from simulated inhalation exposures to BD indicate that incorporation of differential lung region metabolism is important in describing species differences in pulmonary response and that these differences may have implications for risk assessments of human exposures to BD. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

A preliminary regional PBPK model of lung metabolism for improving species dependent descriptions of 1,3-butadiene and its metabolites

DOE PAGES

Campbell, Jerry; Van Landingham, Cynthia; Crowell, Susan; ...

2015-06-12

1,3-Butadiene (BD), a volatile organic chemical (VOC), is used in synthetic rubber production and other industrial processes. It is detectable at low levels in ambient air as well as in tobacco smoke and gasoline vapors. Inhalation exposures to high concentrations of BD have been associated with lung cancer in both humans and experimental animals, although differences in species sensitivity have been observed. Metabolically active lung cells such as Pulmonary Type I and Type II epithelial cells and club cells (Clara cells) 1 are potential targets of BD metabolite-induced toxicity. Metabolic capacities of these cells, their regional densities, and distributions varymore » throughout the respiratory tract as well as between species and cell types. Here we present a physiologically based pharmacokinetic (PBPK) model for BD that includes a regional model of lung metabolism, based on a previous model for styrene, to provide species-dependent descriptions of BD metabolism in the mouse, rat, and human. Since there are no in vivo data on BD pharmacokinetics in the human, the rat and mouse models were parameterized to the extent possible on the basis of in vitro metabolic data. Where it was necessary to use in vivo data, extrapolation from rat to mouse was performed to evaluate the level of uncertainty in the human model. A kidney compartment and description of downstream metabolism were also included in the model to allow for eventual use of available urinary and blood biomarker data in animals and humans to calibrate the model for estimation of BD exposures and internal metabolite levels. Results from simulated inhalation exposures to BD indicate that incorporation of differential lung region metabolism is important in describing species differences in pulmonary response and that these differences may have implications for risk assessments of human exposures to BD.« less

A preliminary regional PBPK model of lung metabolism for improving species dependent descriptions of 1,3-butadiene and its metabolites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Campbell, Jerry; Van Landingham, Cynthia; Crowell, Susan

1,3-Butadiene (BD), a volatile organic chemical (VOC), is used in synthetic rubber production and other industrial processes. It is detectable at low levels in ambient air as well as in tobacco smoke and gasoline vapors. Inhalation exposures to high concentrations of BD have been associated with lung cancer in both humans and experimental animals, although differences in species sensitivity have been observed. Metabolically active lung cells such as Pulmonary Type I and Type II epithelial cells and club cells (Clara cells) 1 are potential targets of BD metabolite-induced toxicity. Metabolic capacities of these cells, their regional densities, and distributions varymore » throughout the respiratory tract as well as between species and cell types. Here we present a physiologically based pharmacokinetic (PBPK) model for BD that includes a regional model of lung metabolism, based on a previous model for styrene, to provide species-dependent descriptions of BD metabolism in the mouse, rat, and human. Since there are no in vivo data on BD pharmacokinetics in the human, the rat and mouse models were parameterized to the extent possible on the basis of in vitro metabolic data. Where it was necessary to use in vivo data, extrapolation from rat to mouse was performed to evaluate the level of uncertainty in the human model. A kidney compartment and description of downstream metabolism were also included in the model to allow for eventual use of available urinary and blood biomarker data in animals and humans to calibrate the model for estimation of BD exposures and internal metabolite levels. Results from simulated inhalation exposures to BD indicate that incorporation of differential lung region metabolism is important in describing species differences in pulmonary response and that these differences may have implications for risk assessments of human exposures to BD.« less

Conserved and Divergent Molecular and Anatomic Features of Human and Mouse Nephron Patterning.

PubMed

Lindström, Nils O; Tran, Tracy; Guo, Jinjin; Rutledge, Elisabeth; Parvez, Riana K; Thornton, Matthew E; Grubbs, Brendan; McMahon, Jill A; McMahon, Andrew P

2018-03-01

The nephron is the functional unit of the kidney, but the mechanism of nephron formation during human development is unclear. We conducted a detailed analysis of nephron development in humans and mice by immunolabeling, and we compared human and mouse nephron patterning to describe conserved and divergent features. We created protein localization maps that highlight the emerging patterns along the proximal-distal axis of the developing nephron and benchmark expectations for localization of functionally important transcription factors, which revealed unanticipated cellular diversity. Moreover, we identified a novel nephron subdomain marked by Wnt4 expression that we fate-mapped to the proximal mature nephron. Significant conservation was observed between human and mouse patterning. We also determined the time at which markers for mature nephron cell types first emerge-critical data for the renal organoid field. These findings have conceptual implications for the evolutionary processes driving the diversity of mammalian organ systems. Furthermore, these findings provide practical insights beyond those gained with mouse and rat models that will guide in vitro efforts to harness the developmental programs necessary to build human kidney structures. Copyright © 2018 by the American Society of Nephrology.

Dissection of the Mouse Pancreas for Histological Analysis and Metabolic Profiling.

PubMed

Veite-Schmahl, Michelle J; Regan, Daniel P; Rivers, Adam C; Nowatzke, Joseph F; Kennedy, Michael A

2017-08-19

We have been investigating the pancreas specific transcription factor, 1a cre-recombinase; lox-stop-lox- Kristen rat sarcoma, glycine to aspartic acid at the 12 codon (Ptf1a cre/+ ;LSL-Kras G12D/+ ) mouse strain as a model of human pancreatic cancer. The goal of our current studies is to identify novel metabolic biomarkers of pancreatic cancer progression. We have performed metabolic profiling of urine, feces, blood, and pancreas tissue extracts, as well as histological analyses of the pancreas to stage the cancer progression. The mouse pancreas is not a well-defined solid organ like in humans, but rather is a diffusely distributed soft tissue that is not easily identified by individuals unfamiliar with mouse internal anatomy or by individuals that have little or no experience performing mouse organ dissections. The purpose of this article is to provide a detailed step-wise visual demonstration to guide novices in the removal of the mouse pancreas by dissection. This article should be especially valuable to students and investigators new to research that requires harvesting of the mouse pancreas by dissection for metabolic profiling or histological analyses.

The role of vertebrate models in understanding craniosynostosis.

PubMed

Holmes, Greg

2012-09-01

Craniosynostosis (CS), the premature fusion of cranial sutures, is a relatively common pediatric anomaly, occurring in isolation or as part of a syndrome. A growing number of genes with pathologic mutations have been identified for syndromic and nonsyndromic CS. The study of human sutural material obtained post-operatively is not sufficient to understand the etiology of CS, for which animal models are indispensable. The similarity of the human and murine calvarial structure, our knowledge of mouse genetics and biology, and ability to manipulate the mouse genome make the mouse the most valuable model organism for CS research. A variety of mouse mutants are available that model specific human CS mutations or have CS phenotypes. These allow characterization of the biochemical and morphological events, often embryonic, which precede suture fusion. Other vertebrate organisms have less functional genetic utility than mice, but the rat, rabbit, chick, zebrafish, and frog provide alternative systems in which to validate or contrast molecular functions relevant to CS.

Lentiviral vectors containing mouse Csf1r control elements direct macrophage-restricted expression in multiple species of birds and mammals

PubMed Central

Pridans, Clare; Lillico, Simon; Whitelaw, Bruce; Hume, David A

2014-01-01

The development of macrophages requires signaling through the lineage-restricted receptor Csf1r. Macrophage-restricted expression of transgenic reporters based upon Csf1r requires the highly conserved Fms-intronic regulatory element (FIRE). We have created a lentiviral construct containing mouse FIRE and promoter. The lentivirus is capable of directing macrophage-restricted reporter gene expression in mouse, rat, human, pig, cow, sheep, and even chicken. Rat bone marrow cells transduced with the lentivirus were capable of differentiating into macrophages expressing the reporter gene in vitro. Macrophage-restricted expression may be desirable for immunization or immune response modulation, and for gene therapy for lysosomal storage diseases and some immunodeficiencies. The small size of the Csf1r transcription control elements will allow the insertion of large “cargo” for applications in gene therapy and vaccine delivery. PMID:26015955

Rodent Control Pesticide Safety Review

EPA Pesticide Factsheets

EPA completed a safety review in 2008 of rat and mouse poison products. Many companies' products have our safety measures that reduce risks to humans, pets, and non-target wildlife. Products must be sold with bait stations that securely contain the poison.

Immunotargeting and cloning of two CD34 variants exhibiting restricted expression in adult rat endothelia in vivo.

PubMed

Testa, Jacqueline E; Chrastina, Adrian; Oh, Phil; Li, Yan; Witkiewicz, Halina; Czarny, Malgorzata; Buss, Tim; Schnitzer, Jan E

2009-08-01

Mapping protein expression of endothelial cells (EC) in vivo is fundamental to understanding cellular function and may yield new tissue-selective targets. We have developed a monoclonal antibody, MAb J120, to a protein expressed primarily in rat lung and heart endothelium. The antigen was identified as CD34, a marker of hematopoietic stem cells and global marker of endothelial cells in human and mouse tissues. PCR-based cloning identified two CD34 variant proteins, full length and truncated, both of which are expressed on luminal endothelial cell plasma membranes (P) isolated from lung. Truncated CD34 predominated in heart P, and neither variant was detected in P from kidney or liver. CD34 in lung was readily accessible to (125)I-J120 inoculated intravenously, and immunohistochemistry showed strong CD34 expression in lung EC. Few microvessels stained in heart and kidney, and no CD34 was detected in vessels of other organs or in lymphatics. We present herein the first complete sequence of a rat CD34 variant and show for the first time that the encoded truncated variant is endogenously expressed on EC in vivo. We also demonstrate that CD34 expression in rat EC, unlike mouse and human, is restricted in its distribution enabling quite specific lung targeting in vivo.

Absolute measurement of species differences in sodium taurocholate cotransporting polypeptide (NTCP/Ntcp) and its modulation in cultured hepatocytes.

PubMed

Qiu, Xi; Bi, Yi-An; Balogh, Larissa M; Lai, Yurong

2013-09-01

Species differences among membrane transporters can be remarkable and difficult to properly assess by conventional methods. Herein, we employed the first use of stable isotope labeling in mammals or stable isotope-labeled peptides combined with mass spectrometry to identify species differences in sodium taurocholate cotransporting polypeptide (NTCP/Ntcp) protein expression in liver tissue and to characterize the modulation of protein expression in sandwich-cultured human (SCHH) and rat hepatocytes (SCRH). The lower limit of quantification was established to be 5 fmol on column with a standard curve that was linear up to 2000 fmol. The accuracy and precision were evaluated with three quality control samples and known amounts of synthetic proteotypic peptides that were spiked into the membrane protein extracts. The overall relative error and coefficient of variation were less than 10%. The expression of Ntcp in mouse and rat was significant higher than that in human (five-fold) and monkey (two-fold) and ranked as mouse > rat > monkey > human. In the cultured hepatocytes, although significant downregulation of Ntcp expression in SCRH at day 5 after the culture was detected, NTCP expression in SCHH was comparable to the suspension hepatocytes. The results suggested that NTCP/Ntcp modulation in cultured hepatocytes is species specific. Copyright © 2013 Wiley Periodicals, Inc.

Adenosine transport systems on dissociated brain cells from mouse, guinea-pig, and rat

DOE Office of Scientific and Technical Information (OSTI.GOV)

Johnston, M.E.; Geiger, J.D.

1990-09-01

The kinetics and sodium dependence of adenosine transport were determined using an inhibitor-stop method on dissociated cell body preparations obtained from mouse, guinea-pig and rat brain. Transport affinity (KT) values for the high affinity adenosine transport systems KT(H) were significantly different between these three species; mean +/- SEM values were 0.34 +/- 0.1 in mouse, 0.9 +/- 0.2 in rat, and 1.5 +/- 0.5 microM in guinea-pig. The KT values for the low affinity transport system KT(L) were not different between the three species. Brain cells from rat displayed a significantly greater maximal capacity to accumulate (3H)adenosine (Vmax) than didmore » mouse or guinea-pig for the high affinity system, or than did mouse for the low affinity system. When sodium chloride was replaced in the transport medium with choline chloride, the KT(H) values for guinea-pig and rat were both increased by approximately 100%; only in rat did the change reach statistical significance. The sodium-dependence of adenosine transport in mouse brain was clearly absent. The differences between KT(H) values in mouse and those in guinea-pig or rat were accentuated in the absence of sodium. The differences in kinetic values, ionic requirements, and pharmacological characteristics between adenosine transporters in CNS tissues of mouse, guinea-pig and rat may help account for some of the variability noted among species in terms of their physiological responses to adenosine.« less

Cloning and characterization of the mouse alpha1C/A-adrenergic receptor gene and analysis of an alpha1C promoter in cardiac myocytes: role of an MCAT element that binds transcriptional enhancer factor-1 (TEF-1).

PubMed

O'Connell, T D; Rokosh, D G; Simpson, P C

2001-05-01

alpha1-Adrenergic receptor (AR) subtypes in the heart are expressed by myocytes but not by fibroblasts, a feature that distinguishes alpha1-ARs from beta-ARs. Here we studied myocyte-specific expression of alpha1-ARs, focusing on the subtype alpha1C (also called alpha1A), a subtype implicated in cardiac hypertrophic signaling in rat models. We first cloned the mouse alpha1C-AR gene, which consisted of two exons with an 18 kb intron, similar to the alpha1B-AR gene. The receptor coding sequence was >90% homologous to that of rat and human. alpha1C-AR transcription in mouse heart was initiated from a single Inr consensus sequence at -588 from the ATG; this and a putative polyadenylation sequence 8.5 kb 3' could account for the predominant 11 kb alpha1C mRNA in mouse heart. A 5'-nontranscribed fragment of 4.4 kb was active as a promoter in cardiac myocytes but not in fibroblasts. Promoter activity in myocytes required a single muscle CAT (MCAT) element, and this MCAT bound in vitro to recombinant and endogenous transcriptional enhancer factor-1. Thus, alpha1C-AR transcription in cardiac myocytes shares MCAT dependence with other cardiac-specific genes, including the alpha- and beta-myosin heavy chains, skeletal alpha-actin, and brain natriuretic peptide. However, the mouse alpha1C gene was not transcribed in the neonatal heart and was not activated by alpha1-AR and other hypertrophic agonists in rat myocytes, and thus differed from other MCAT-dependent genes and the rat alpha1C gene.

Monoclonal antibodies against the rat liver glucocorticoid receptor.

PubMed Central

Okret, S; Wikström, A C; Wrange, O; Andersson, B; Gustafsson, J A

1984-01-01

Splenic cells from one BALB/c mouse and one C57/BL mouse, immunized with purified rat liver glucocorticoid receptor (GR), were fused with the mouse myeloma cell line Sp 2/0-Ag 14. Screening for production of anti-GR-antibodies by the hybridomas was carried out with an enzyme-linked immunosorbent assay, using partially purified rat liver GR as antigen. Further screening was by a second-antibody immunoprecipitation assay using [3H]triamcinolone acetonide-GR complex from rat liver cytosol as tracer. Hybridomas from 10 different microplate wells, positive in both assays, were successfully cloned by the limiting dilution method to monoclonality. The different origins of the monoclonal antibodies were confirmed by their various isoelectric points when analyzed by isoelectric focusing. Four of the monoclonal hybridoma cell lines secreted IgM antibodies; two, IgG1; three, IgG2a; and one, IgG2b. The GR-antibody complex was identified in glycerol density gradients by a shift of the 4S GR to an 8.5S or 19S GR-antibody complex when incubated with monoclonal IgG or IgM antibody, respectively. The 10 monoclonal antibodies recognized different determinants on the GR, all situated on that domain of the receptor that is separate from the ligand and DNA-binding domains. Also, the cross-reactivity to the mouse liver GR varied among the monoclonal antibodies. No cross-reactivity was observed to the human lymphocytic GR. NaDodSO4 electrophoresis of a 0.5% pure GR preparation followed by immunoblotting using one of the monoclonal antibodies identified a single peptide with a molecular weight of 94,000, identical to the purified rat liver GR. Images PMID:6200880

Age-related retention of fiber cell nuclei and nuclear fragments in the lens cortices of multiple species

PubMed Central

Pendergrass, William; Zitnik, Galynn; Urfer, Silvan R.

2011-01-01

Purpose To determine the differences between species in the retention of lens fiber cell nuclei and nuclear fragments in the aging lens cortex and the relationship of nuclear retention to lens opacity. For this purpose old human, monkey, dog, and rat lenses were compared to those of three strains of mouse. We also investigated possible mechanisms leading to nuclear retention. Methods Fixed specimens of the species referred to above were obtained from immediate on site sacrifice of mice and rats, or from recently fixed lenses of other species, dogs, monkeys, and humans, obtained from collaborators. The retention of undegraded nuclei and nuclear fragments was graded 1–4 from histologic observation. All species lenses were examined microscopically in fixed sections stained with hematoxylin and eosin (H&E) or 4',6-diamidino-2-phenylindole (DAPI). Slit lamp observations were made only on the mice and rats before sacrifice and lens fixation. Values of 0 to 4 (clear lens to cataract) were given to degree of opacity. MRNA content in young versus old C57BL/6 mouse lenses was determined by quantitative PCR (qPCR) for DNase II-like acid DNase (DLAD) and other proteins. DLAD protein was determined by immunofluorescence of fixed eye sections. Results In old C57BL/6 and DBA mice and, to a lesser degree, in old CBA mice and old Brown Norway (BN) rats lenses were seen to contain a greatly expanded pool of unresolved whole nuclei or fragments of nuclei in differentiating lens fiber cells. This generally correlated with increased slit lamp opacities in these mice. Most old dog lenses also had an increase in retained cortical nuclei, as did a few old humans. However, a second rat strain, BNF1, in which opacity was quite high had no increase in retained nuclei with age nor did any of the old monkeys, indicating that retained nuclei could not be a cause of opacity in these animals. The nuclei and nuclear fragments were located at all levels in the outer cortex extending inward from the lens equator and were observable by the DAPI. These nuclei and nuclear fragments were seen from 12 months onward in all C57BL/6 and DBA/2 mice and to a lesser degree in the CBA, increasing in number and in space occupancy with increasing age. Preliminary results suggest that retention of nuclei in the C57BL/6 mouse is correlated with an age-related loss of DLAD from old lenses. Conclusions A very marked apparently light refractive condition caused by retained cortical nuclei and nuclear fragments is present in the lens cortices, increasing with age in the three strains of mice examined and in one of two strains of rats (BN). This condition was also seen in some old dogs and a few old humans. It may be caused by an age-related loss of DLAD, which is essential for nuclear DNA degradation in the lens. However, this condition does not develop in old BNF1 rats, or old monkeys and is only seen sporadically in humans. Thus, it can not be a universal cause for age related lens opacity or cataract presence, although it develops concurrently with opacity in mice. This phenomenon should be considered when using the old mouse as a model for human age-related cataract. PMID:22065920

Cloning, sequencing and expression in MEL cells of a cDNA encoding the mouse ribosomal protein S5.

PubMed

Vanegas, N; Castañeda, V; Santamaría, D; Hernández, P; Schvartzman, J B; Krimer, D B

1997-06-05

We describe the isolation and characterization of a cDNA encoding the mouse S5 ribosomal protein. It was isolated from a MEL (murine erythroleukemia) cell cDNA library by differential hybridization as a down regulated sequence during HMBA-induced differentiation. Northern series analysis showed that S5 mRNA expression is reduced 5-fold throughout the differentiation process. The mouse S5 mRNA is 760 bp long and encodes for a 204 amino acid protein with 94% homology with the human and rat S5.

EMBRYONIC PALATAL RESPONSES TO TERATOGENS IN SERUM-FREE ORGAN CULTURE

EPA Science Inventory

This study examines development of rat, mouse and human embryonic palates in submerged, serum-free organ culture. he concentration-response profiles for retinoic acid (RA), triamcinolone (TRI), hydrocortisone (HC), dexamethasone (DEX), and 2,3,7,11- tetrachlorodibenzo-p-dioxin (T...

Proteomic Assessment of Poultry Spermatozoa

USDA-ARS?s Scientific Manuscript database

Fully characterizing the protein composition of spermatozoa is the first step in utilizing proteomics to delineate the function of sperm proteins. To date, sperm proteome maps have been partially developed for the human, mouse, rat, bull and several invertebrates. Here we report the first proteomic...

Temporal profiles and 2-dimensional oxy-, deoxy-, and total-hemoglobin somatosensory maps in rat versus mouse cortex

PubMed Central

Prakash, Neal; Biag, Jonathan D.; Sheth, Sameer A.; Mitsuyama, Satoshi; Theriot, Jeremy; Ramachandra, Chaithanya; Toga, Arthur W.

2007-01-01

Background Mechanisms of neurovascular coupling—the relationship between neuronal chemoelectrical activity and compensatory metabolic and hemodynamic changes—appear to be preserved across species from rats to humans despite differences in scale. However, previous work suggests that the highly cellular dense mouse somatosensory cortex has different functional hemodynamic changes compared to other species. Methods We developed novel hardware and software for 2-dimensional optical spectroscopy (2DOS). Optical changes at four simultaneously recorded wavelengths were measured in both rat and mouse primary somatosensory cortex (S1) evoked by forepaw stimulation to create four spectral maps. The spectral maps were converted to maps of deoxy-, oxy-, and total-hemoglobin (HbR, HbO, and HbT) concentration changes using the modified Beer-Lambert law and phantom HbR and HbO absorption spectra. Results Functional hemodynamics were different in mouse versus rat neocortex. On average, hemodynamics were as expected in rat primary somatosensory cortex (S1): the fractional change in the log of HbT concentration increased monophasically 2 s after stimulus, whereas HbO changes mirrored HbR changes, with HbO showing a small initial dip at 0.5 s followed by a large increase 3.0 s post stimulus. In contrast, mouse S1 showed a novel type of stimulus-evoked hemodynamic response, with prolonged, concurrent, monophasic increases in HbR and HbT and a parallel decrease in HbO that all peaked 3.5–4.5 s post stimulus onset. For rats, at any given time point the average size and shape of HbO and HbR forepaw maps were the same, whereas surface veins distorted the shape of the HbT map. For mice, HbO, HbR, and HbT forepaw maps were generally the same size and shape at any post-stimulus time point. Conclusions 2DOS using image splitting optics is feasible across species for brain mapping and quantifying the map topography of cortical hemodynamics. These results suggest that during physiologic stimulation, different species and/or cortical architecture may give rise to different hemodynamic changes during neurovascular coupling. PMID:17574868

A Metadata description of the data in "A metabolomic comparison of urinary changes in type 2 diabetes in mouse, rat, and human."

PubMed Central

2011-01-01

Background Metabolomics is a rapidly developing functional genomic tool that has a wide range of applications in diverse fields in biology and medicine. However, unlike transcriptomics and proteomics there is currently no central repository for the depositing of data despite efforts by the Metabolomics Standard Initiative (MSI) to develop a standardised description of a metabolomic experiment. Findings In this manuscript we describe how the MSI description has been applied to a published dataset involving the identification of cross-species metabolic biomarkers associated with type II diabetes. The study describes sample collection of urine from mice, rats and human volunteers, and the subsequent acquisition of data by high resolution 1H NMR spectroscopy. The metadata is described to demonstrate how the MSI descriptions could be applied in a manuscript and the spectra have also been made available for the mouse and rat studies to allow others to process the data. Conclusions The intention of this manuscript is to stimulate discussion as to whether the MSI description is sufficient to describe the metadata associated with metabolomic experiments and encourage others to make their data available to other researchers. PMID:21801423

Inhibition of colony-stimulating-factor-1 signaling in vivo with the orally bioavailable cFMS kinase inhibitor GW2580.

PubMed

Conway, James G; McDonald, Brad; Parham, Janet; Keith, Barry; Rusnak, David W; Shaw, Eva; Jansen, Marilyn; Lin, Peiyuan; Payne, Alan; Crosby, Renae M; Johnson, Jennifer H; Frick, Lloyd; Lin, Min-Hwa Jasmine; Depee, Scott; Tadepalli, Sarva; Votta, Bart; James, Ian; Fuller, Karen; Chambers, Timothy J; Kull, Frederick C; Chamberlain, Stanley D; Hutchins, Jeff T

2005-11-01

Colony-stimulating-factor-1 (CSF-1) signaling through cFMS receptor kinase is increased in several diseases. To help investigate the role of cFMS kinase in disease, we identified GW2580, an orally bioavailable inhibitor of cFMS kinase. GW2580 completely inhibited human cFMS kinase in vitro at 0.06 microM and was inactive against 26 other kinases. GW2580 at 1 microM completely inhibited CSF-1-induced growth of mouse M-NFS-60 myeloid cells and human monocytes and completely inhibited bone degradation in cultures of human osteoclasts, rat calvaria, and rat fetal long bone. In contrast, GW2580 did not affect the growth of mouse NS0 lymphoblastoid cells, human endothelial cells, human fibroblasts, or five human tumor cell lines. GW2580 also did not affect lipopolysaccharide (LPS)-induced TNF, IL-6, and prostaglandin E2 production in freshly isolated human monocytes and mouse macrophages. After oral administration, GW2580 blocked the ability of exogenous CSF-1 to increase LPS-induced IL-6 production in mice, inhibited the growth of CSF-1-dependent M-NFS-60 tumor cells in the peritoneal cavity, and diminished the accumulation of macrophages in the peritoneal cavity after thioglycolate injection. Unexpectedly, GW2580 inhibited LPS-induced TNF production in mice, in contrast to effects on monocytes and macrophages in vitro. In conclusion, GW2580's selective inhibition of monocyte growth and bone degradation is consistent with cFMS kinase inhibition. The ability of GW2580 to chronically inhibit CSF-1 signaling through cFMS kinase in normal and tumor cells in vivo makes GW2580 a useful tool in assessing the role of cFMS kinase in normal and disease processes.

Phospholipase A2 in experimental allergic bronchitis: a lesson from mouse and rat models.

PubMed

Mruwat, Rufayda; Yedgar, Saul; Lavon, Iris; Ariel, Amiram; Krimsky, Miron; Shoseyov, David

2013-01-01

Phospholipases A2 (PLA2) hydrolyzes phospholipids, initiating the production of inflammatory lipid mediators. We have previously shown that in rats, sPLA2 and cPLA2 play opposing roles in the pathophysiology of ovalbumin (OVA)-induced experimental allergic bronchitis (OVA-EAB), an asthma model: Upon disease induction sPLA2 expression and production of the broncho-constricting CysLTs are elevated, whereas cPLA2 expression and the broncho-dilating PGE2 production are suppressed. These were reversed upon disease amelioration by treatment with an sPLA2 inhibitor. However, studies in mice reported the involvement of both sPLA2 and cPLA2 in EAB induction. To examine the relevance of mouse and rat models to understanding asthma pathophysiology. OVA-EAB was induced in mice using the same methodology applied in rats. Disease and biochemical markers in mice were compared with those in rats. As in rats, EAB in mice was associated with increased mRNA of sPLA2, specifically sPLA2gX, in the lungs, and production of the broncho-constricting eicosanoids CysLTs, PGD2 and TBX2 in bronchoalveolar lavage (BAL). In contrast, EAB in mice was associated also with elevated cPLA2 mRNA and PGE2 production. Yet, treatment with an sPLA2 inhibitor ameliorated the EAB concomitantly with reverting the expression of both cPLA2 and sPLA2, and eicosanoid production. In both mice and rats sPLA2 is pivotal in OVA-induced EAB. Yet, amelioration of asthma markers in mouse models, and human tissues, was observed also upon cPLA2 inhibition. It is plausible that airway conditions, involving multiple cell types and organs, require the combined action of more than one, essential, PLA2s.

The Trw Type IV Secretion System of Bartonella Mediates Host-Specific Adhesion to Erythrocytes

PubMed Central

Vayssier-Taussat, Muriel; Le Rhun, Danielle; Deng, Hong Kuan; Biville, Francis; Cescau, Sandra; Danchin, Antoine; Marignac, Geneviève; Lenaour, Evelyne; Boulouis, Henri Jean; Mavris, Maria; Arnaud, Lionel; Yang, Huanming; Wang, Jing; Quebatte, Maxime; Engel, Philipp; Saenz, Henri; Dehio, Christoph

2010-01-01

Bacterial pathogens typically infect only a limited range of hosts; however, the genetic mechanisms governing host-specificity are poorly understood. The α-proteobacterial genus Bartonella comprises 21 species that cause host-specific intraerythrocytic bacteremia as hallmark of infection in their respective mammalian reservoirs, including the human-specific pathogens Bartonella quintana and Bartonella bacilliformis that cause trench fever and Oroya fever, respectively. Here, we have identified bacterial factors that mediate host-specific erythrocyte colonization in the mammalian reservoirs. Using mouse-specific Bartonella birtlesii, human-specific Bartonella quintana, cat-specific Bartonella henselae and rat-specific Bartonella tribocorum, we established in vitro adhesion and invasion assays with isolated erythrocytes that fully reproduce the host-specificity of erythrocyte infection as observed in vivo. By signature-tagged mutagenesis of B. birtlesii and mutant selection in a mouse infection model we identified mutants impaired in establishing intraerythrocytic bacteremia. Among 45 abacteremic mutants, five failed to adhere to and invade mouse erythrocytes in vitro. The corresponding genes encode components of the type IV secretion system (T4SS) Trw, demonstrating that this virulence factor laterally acquired by the Bartonella lineage is directly involved in adherence to erythrocytes. Strikingly, ectopic expression of Trw of rat-specific B. tribocorum in cat-specific B. henselae or human-specific B. quintana expanded their host range for erythrocyte infection to rat, demonstrating that Trw mediates host-specific erythrocyte infection. A molecular evolutionary analysis of the trw locus further indicated that the variable, surface-located TrwL and TrwJ might represent the T4SS components that determine host-specificity of erythrocyte parasitism. In conclusion, we show that the laterally acquired Trw T4SS diversified in the Bartonella lineage to facilitate host-restricted adhesion to erythrocytes in a wide range of mammals. PMID:20548954

Molecular cloning of the mouse gene coding for {alpha}{sub 2}-macroglobulin and targeting of the gene in embryonic stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Umans, L.; Serneels, L.; Hilliker, C.

1994-08-01

The authors have cloned the mouse gene coding for {alpha}{sub 2}-macroglobulin in overlapping {lambda} clones and have analyzed its structure. The gene contains 36 exons, coding for the 4.8-kb cDNA that we cloned previously. Including putative control elements in the 5{prime} flanking region, the gene covers about 45 kb. A region of 3.8 kb, stretching from 835 bases upstream of the cDNA start site to exon 4, including all intervening sequences, was sequenced completely. The analysis demonstrated that the putative promoter region of the mouse A2M gene differed considerably from the known promoter sequences of the human A2M gene andmore » of the rat acute-phas A2M gene. Comparison of the exon-intron structure of all known genes of the A2M family confirmed that the rat acute phase A2M gene is more closely related to the human gene than to the mouse A2M gene. To generate mice with the A2M gene inactivated, an insertion type of construct containing 7.5 kb of genomic DNA of the mouse strain 129/J, encompassing exons 16 to 19, was synthesized. A hygromycin marker gene was embedded in intron 17. After electroporation, 198 hygromycin-resistant ES cell lines were isolated and analyzed by Southern blotting. Five ES cell lines were obtained with one allele of the mouse A2M gene targeted by this insertion construct, demonstrating that the position and the characteristics of the vector served the intended goal.« less

Metabolism of a new positive inotropic agent, 3,4-dihydro-6-[4-(3,4-dimethoxybenzoyl)-1-piperazinyl]-2(1H)- quinolinone (OPC-8212) in the rat, mouse, dog, monkey and human.

PubMed

Miyamoto, G; Sasabe, H; Tominaga, N; Uegaki, N; Tominaga, M; Shimizu, T

1988-10-01

1. After OPC-8212 was orally given to rats, mice, dogs, monkeys and humans, its metabolites were identified by n.m.r. and mass spectrometry, and their concentrations in the plasma, urine and faeces of these species were measured by high-performance liquid chromatography (h.p.l.c.). 2. Hydrolysis of the amide group, oxidation and cleavage of the piperazine ring, O-demethylation of the methoxy group, and conjugation were proposed as metabolic pathways of OPC-8212. 3. In rats, mice and monkeys given OPC-8212 orally, metabolites M-1 to M-6 were detected in the plasma, urine and faeces, while M-1, -4, -5 and M-6 were detected in dogs, and M-1, M-3, M-4, M-5 and M-6 were detected in humans. 4. Conjugates of metabolites M-6 and M-7, with glucuronic acid and sulphuric acid, were observed in the urine of rats and humans.

Dentin and pulp sense cold stimulus.

PubMed

Tokuda, Masayuki; Tatsuyama, Shoko; Fujisawa, Mari; Morimoto-Yamashita, Yoko; Kawakami, Yoshiko; Shibukawa, Yoshiyuki; Torii, Mistuso

2015-05-01

Dentin hypersensitivity is a common symptom, and recent convergent evidences have reported transient receptor potential (TRP) channels in odontoblasts act as mechanical and thermal molecular sensor, which detect stimulation applied on the exposed dentin surface, to drive multiple odontoblastic cellular functions, such as sensory transduction and/or dentin formation. In the present study, we confirmed expression of TRP melastatin subfamily member-8 (TRPM8) channels in primary cultured cells derived from human dental pulp cells (HPCs) and mouse odontoblast-lineage cells (OLCs) as well as in dentin matrix protein-1 (DMP-1) and dentin sialoprotein (DSP) positive acutely isolated rat odontoblasts from dental pulp tissue slice culture by immunohistochemical analyses. In addition, we detected TRPM8 channel expression on HPCs and OLCs by RT-PCR and Western blotting analyses. These results indicated that both odontoblasts and dental pulp cells express TRPM8 channels in rat, mouse and human, and therefore we hypothesize they may contribute as cold sensor in tooth. Copyright © 2015 Elsevier Ltd. All rights reserved.

Identification of a mouse synaptic glycoprotein gene in cultured neurons.

PubMed

Yu, Albert Cheung-Hoi; Sun, Chun Xiao; Li, Qiang; Liu, Hua Dong; Wang, Chen Ran; Zhao, Guo Ping; Jin, Meilei; Lau, Lok Ting; Fung, Yin-Wan Wendy; Liu, Shuang

2005-10-01

Neuronal differentiation and aging are known to involve many genes, which may also be differentially expressed during these developmental processes. From primary cultured cerebral cortical neurons, we have previously identified various differentially expressed gene transcripts from cultured cortical neurons using the technique of arbitrarily primed PCR (RAP-PCR). Among these transcripts, clone 0-2 was found to have high homology to rat and human synaptic glycoprotein. By in silico analysis using an EST database and the FACTURA software, the full-length sequence of 0-2 was assembled and the clone was named as mouse synaptic glycoprotein homolog 2 (mSC2). DNA sequencing revealed transcript size of mSC2 being smaller than the human and rat homologs. RT-PCR indicated that mSC2 was expressed differentially at various culture days. The mSC2 gene was located in various tissues with higher expression in brain, lung, and liver. Functions of mSC2 in neurons and other tissues remain elusive and will require more investigation.

Survival of Free and Encapsulated Human and Rat Islet Xenografts Transplanted into the Mouse Bone Marrow

PubMed Central

Meier, Raphael P. H.; Seebach, Jörg D.; Morel, Philippe; Mahou, Redouan; Borot, Sophie; Giovannoni, Laurianne; Parnaud, Geraldine; Montanari, Elisa; Bosco, Domenico; Wandrey, Christine; Berney, Thierry; Bühler, Leo H.; Muller, Yannick D.

2014-01-01

Bone marrow was recently proposed as an alternative and potentially immune-privileged site for pancreatic islet transplantation. The aim of the present study was to assess the survival and rejection mechanisms of free and encapsulated xenogeneic islets transplanted into the medullary cavity of the femur, or under the kidney capsule of streptozotocin-induced diabetic C57BL/6 mice. The median survival of free rat islets transplanted into the bone marrow or under the kidney capsule was 9 and 14 days, respectively, whereas that of free human islets was shorter, 7 days (bone marrow) and 10 days (kidney capsule). Infiltrating CD8+ T cells and redistributed CD4+ T cells, and macrophages were detected around the transplanted islets in bone sections. Recipient mouse splenocytes proliferated in response to donor rat stimulator cells. One month after transplantation under both kidney capsule or into bone marrow, encapsulated rat islets had induced a similar degree of fibrotic reaction and still contained insulin positive cells. In conclusion, we successfully established a small animal model for xenogeneic islet transplantation into the bone marrow. The rejection of xenogeneic islets was associated with local and systemic T cell responses and macrophage recruitment. Although there was no evidence for immune-privilege, the bone marrow may represent a feasible site for encapsulated xenogeneic islet transplantation. PMID:24625569

Single Body Parts are Processed by Individual Neurons in the Mouse Dorsolateral Striatum

PubMed Central

Coffey, Kevin R.; Nader, Miles; West, Mark O.

2016-01-01

Interest in the dorsolateral striatum (DLS) has generated numerous scientific studies of its neuropathologies, as well as its roles in normal sensorimotor integration and learning. Studies are informed by knowledge of DLS functional organization, the guiding principle being its somatotopic afferent projections from primary somatosensory (S1) and motor (M1) cortices. The potential to connect behaviorally relevant function to detailed structure is elevated by mouse models, which have access to extensive genetic neuroscience tool kits. Remaining to be demonstrated, however, is whether the correspondence between S1/M1 corticostriatal terminal distributions and the physiological properties of DLS neurons demonstrated in rats and non-human primates exists in mice. Given that the terminal distribution of S1/M1 projections to the DLS in mice is similar to that in rats, we studied whether firing rates (FRs) of DLS neurons in awake, behaving mice are related to activity of individual body parts. MSNs exhibited robust, selective increases in FR during movement or somatosensory stimulation of single body parts. Properties of MSNs, including baseline FRs, locations, responsiveness to stimulation, and proportions of responsive neurons were similar to properties observed in rats. Future studies can be informed by the present demonstration that the mouse lateral striatum functions as a somatic sensorimotor sector of the striatum and appears to be a homolog of the primate putamen, as demonstrated in rats (Carelli and West, 1991). PMID:26827625

The truth about mouse, human, worms and yeast

PubMed Central

2004-01-01

Genome comparisons are behind the powerful new annotation methods being developed to find all human genes, as well as genes from other genomes. Genomes are now frequently being studied in pairs to provide cross-comparison datasets. This 'Noah's Ark' approach often reveals unsuspected genes and may support the deletion of false-positive predictions. Joining mouse and human as the cross-comparison dataset for the first two mammals are: two Drosophila species, D. melanogaster and D. pseudoobscura; two sea squirts, Ciona intestinalis and Ciona savignyi; four yeast (Saccharomyces) species; two nematodes, Caenorhabditis elegans and Caenorhabditis briggsae; and two pufferfish (Takefugu rubripes and Tetraodon nigroviridis). Even genomes like yeast and C. elegans, which have been known for more than five years, are now being significantly improved. Methods developed for yeast or nematodes will now be applied to mouse and human, and soon to additional mammals such as rat and dog, to identify all the mammalian protein-coding genes. Current large disparities between human Unigene predictions (127,835 genes) and gene-scanning methods (45,000 genes) still need to be resolved. This will be the challenge during the next few years. PMID:15601543

The truth about mouse, human, worms and yeast.

PubMed

Nelson, David R; Nebert, Daniel W

2004-01-01

Genome comparisons are behind the powerful new annotation methods being developed to find all human genes, as well as genes from other genomes. Genomes are now frequently being studied in pairs to provide cross-comparison datasets. This 'Noah's Ark' approach often reveals unsuspected genes and may support the deletion of false-positive predictions. Joining mouse and human as the cross-comparison dataset for the first two mammals are: two Drosophila species, D. melanogaster and D. pseudoobscura; two sea squirts, Ciona intestinalis and Ciona savignyi; four yeast (Saccharomyces) species; two nematodes, Caenorhabditis elegans and Caenorhabditis briggsae; and two pufferfish (Takefugu rubripes and Tetraodon nigroviridis). Even genomes like yeast and C. elegans, which have been known for more than five years, are now being significantly improved. Methods developed for yeast or nematodes will now be applied to mouse and human, and soon to additional mammals such as rat and dog, to identify all the mammalian protein-coding genes. Current large disparities between human Unigene predictions (127,835 genes) and gene-scanning methods (45,000 genes) still need to be resolved. This will be the challenge during the next few years.

Mouse scrapie responsive gene 1 (Scrg1): genomic organization, physical linkage to sap30, genetic mapping on chromosome 8, and expression in neuronal primary cell cultures.

PubMed

Dron, M; Tartare, X; Guillo, F; Haik, S; Barbin, G; Maury, C; Tovey, M; Dandoy-Dron, F

2000-11-15

We have previously reported a transcript of a novel mouse gene (Scrg1) with increased expression in transmissible spongiform encephalopathies and the cloning of the human mRNA analogue. In this paper, we present the genomic organization of the mouse and human SCRG1 loci, which exhibit a high degree of conservation. The genes are composed of three exons; the two downstream exons contain the protein coding region. The mouse gene is expressed in brain tissue essentially as a 0.7-kb message but also as a minor 2.6-kb mRNA. We have sequenced 20 kb of DNA at the mouse Scrg1 locus and found that the longer transcript is the prolongation of the 0.7-kb mRNA to a polyadenylation site located about 2 kb further downstream. Sequencing revealed that the mouse Scrg1 gene is physically linked to Sap30, a gene that encodes a protein of the histone deacetylase complex, and genetic linkage mapping assigned the localization of Scrg1 to chromosome 8 between Ant1 and Hmg2. Northern blot analysis showed that Scrg1 is under strict developmental control in mouse embryo and is expressed by cells of neuronal origin in vitro. Comparison of the rat, mouse, and human SCRG1 proteins identified a box of 35 identical contiguous amino acids and a characteristic cysteine distribution pattern defining a new protein signature. Copyright 2000 Academic Press.

Identification and Validation of Plasma Biomarkers in California Sea Lions

DTIC Science & Technology

2014-04-24

of domoic acid in bodily fluids (urine, feces, milk , aqueous humor) provided additional DAT confirmation in some cases. Acute DAT cases were...include, Panda, Dog, Human, Mouse, Sea Lion, Rat, Pig, and Cow in an effort to maximize discoverability, yet minimize false discovery. The criteria

Monitoring transcription initiation activities in rat and dog.

PubMed

Lizio, Marina; Mukarram, Abdul Kadir; Ohno, Mizuho; Watanabe, Shoko; Itoh, Masayoshi; Hasegawa, Akira; Lassmann, Timo; Severin, Jessica; Harshbarger, Jayson; Abugessaisa, Imad; Kasukawa, Takeya; Hon, Chung Chau; Carninci, Piero; Hayashizaki, Yoshihide; Forrest, Alistair R R; Kawaji, Hideya

2017-11-28

The promoter landscape of several non-human model organisms is far from complete. As a part of FANTOM5 data collection, we generated 13 profiles of transcription initiation activities in dog and rat aortic smooth muscle cells, mesenchymal stem cells and hepatocytes by employing CAGE (Cap Analysis of Gene Expression) technology combined with single molecule sequencing. Our analyses show that the CAGE profiles recapitulate known transcription start sites (TSSs) consistently, in addition to uncover novel TSSs. Our dataset can be thus used with high confidence to support gene annotation in dog and rat species. We identified 28,497 and 23,147 CAGE peaks, or promoter regions, for rat and dog respectively, and associated them to known genes. This approach could be seen as a standard method for improvement of existing gene models, as well as discovery of novel genes. Given that the FANTOM5 data collection includes dog and rat matched cell types in human and mouse as well, this data would also be useful for cross-species studies.

Transgenic rats with green, red, and blue fluorescence: powerful tools for bioimaging, cell trafficking, and differentiation

NASA Astrophysics Data System (ADS)

Murakami, Takashi; Kobayashi, Eiji

2005-04-01

The rat represents a perfect animal for broadening medical experiments, because its physiology has been well understood in the history of experimental animals. In addition, its larger body size takes enough advantage for surgical manipulation, compared to the mouse. Many rat models mimicking human diseases, therefore, have been used in a variety of biomedical studies including physiology, pharmacology, transplantation, and immunology. In an effort to create the specifically designed rats for biomedical research and regenerative medicine, we have developed the engineered rat system on the basis of transgenic technology and succeeded in establishing various transgenic rat strains. The transgenic rats with green fluorescent protein (GFP) were generated in the two different strains (Wistar and Lewis), in which GFP is driven under the chicken beta-actin promoter and cytomegalovirus enhancer (CAG promoter). Their GFP expression levels were different in each organ, but the Lewis line expressed GFP strongly and ubiquitously in most of the organs compared with that of Wistar. For red fluorescence, DsRed2 was transduced to the Wistar rats: one line specifically expresses DsRed2 in the liver under the mouse albumin promoter, another is designed for the Cre/LoxP system as the double reporter rat (the initial DsRed2 expression turns on GFP in the presence of Cre recombinase). LacZ-transgenic rats represent blue color, and LacZ is driven the CAG (DA) or ROSA26 promoter (Lewis). Our unique transgenic rats" system highlights the powerful performance for the elucidation of many cellular processes in regenerative medicine, leading to innovative medical treatments.

Photoacoustic Imaging of Epilepsy

DTIC Science & Technology

2013-04-01

mouse brain with the skin and skull intact,” Opt. Lett. 28(19), 1739–1741 (2003). 5. Q. Zhang, Z. Liu, P. R. Carney, Z. Yuan, H. Chen, S. N. Roper, and...imaging at centimeter scale depths. To date PAT has been applied to the detection of breast cancer, skin cancer and osteoarthritis in humans [1–3...the hemodynamic changes and reveal the 3D structures in the rat brain. Two small rats (~40g) were imaged with intact skull and skin but hairs on the

Human umbilical cord mesenchymal stromal cells suppress MHC class II expression on rat vascular endothelium and prolong survival time of cardiac allograft

PubMed Central

Qiu, Ying; Yun, Mark M; Han, Xia; Zhao, Ruidong; Zhou, Erxia; Yun, Sheng

2014-01-01

Background: Human umbilical cord mesenchymal stromal cells (UC-MSCs) have low immunogenicity and immune regulation. To investigate immunomodulatory effects of human UC-MSCs on MHC class II expression and allograft, we transplanted heart of transgenic rats with MHC class II expression on vascular endothelium. Methods: UC-MSCs were obtained from human umbilical cords and confirmed with flow cytometry analysis. Transgenic rat line was established using the construct of human MHC class II transactivator gene (CIITA) under mouse ICAM-2 promoter control. The induced MHC class II expression on transgenic rat vascular endothelial cells (VECs) was assessed with immunohistological staining. And the survival time of cardiac allograft was compared between the recipients with and without UC-MSC transfusion. Results: Flow cytometry confirmed that the human UC-MSCs were positive for CD29, CD44, CD73, CD90, CD105, CD271, and negative for CD34 and HLA-DR. Repeated infusion of human UC-MSCs reduced MHC class II expression on vascular endothelia of transplanted hearts, and increased survival time of allograft. The UC-MSCs increased regulatory cytokines IL10, transforming growth factor (TGF)-β1 and suppressed proinflammatory cytokines IL2 and IFN-γ in vivo. The UC-MSC culture supernatant had similar effects on cytokine expression, and decreased lymphocyte proliferation in vitro. Conclusions: Repeated transfusion of the human UC-MSCs reduced MHC class II expression on vascular endothelia and prolonged the survival time of rat cardiac allograft. PMID:25126177

Evidence for the Involvement of Xenobiotic-responsive Nuclear Receptors in Transcriptional Effects Upon Perfluoroalkyl Acid Exposure in Diverse Species.

EPA Science Inventory

Humans and other species have detectable body burdens of a number of perfluorinated alkyl acids (PFAA) including perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS). In mouse and rat liver these compounds elicit transcriptional and phenotypic effects similar to pe...

Evidence for the Involvement of Xenobiotic-response Nuclear Receptors in Transcriptional Effects Upon Perfluroroalkyl Acid Exposure in Diverse Species

EPA Science Inventory

Humans and ecological species have been found to have detectable body burdens of a number of perfluorinated alkyl acids (PFAA) including perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS). In mouse and rat liver these compounds elicit transcriptional and phenotyp...

77 FR 52240 - Pendimethalin; Pesticide Tolerances

Federal Register 2010, 2011, 2012, 2013, 2014

2012-08-29

..., identified by docket ID number EPA-HQ-OPP-2011-0521, by one of the following methods: Federal eRulemaking... relationship of the results of the studies to human risk. EPA has also considered available information... subchronic rat and mouse studies was manifested as alterations in thyroid hormones, increased thyroid weight...

Gene Expression Profiling in the Liver and Lung of Perfluorooctane Sulfonate-Exposed Mouse Fetuses: Comparison to Changes Induced by Exposure to Perfluorooctanoic Acid

EPA Science Inventory

Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) are environmental contaminants found in the tissues of humans and wildlife. They are activators of peroxisome proliferator-activated receptor-alpha (PPARα) and exhibit hepatocarcinogenic potential in rats. PFOS...

COMPARATIVE ANATOMY OF MAMMALIAN RESPIRATORY TRACTS: THE NASOPHARYNGEAL REGION AND THE TRACHEOBRONCHIAL REGION

EPA Science Inventory

Silicone rubber casts of the respiratory tract are used in morphological studies of the human, baboon, rhesus monkey, dog, rabbit, guinea pig, rat, hamster, and mouse. n these studies, the trachea of the specimen was opened by tracheotomy, and silicone rubber (734 RTV) was introd...

A Review of the Molecular Mechanisms of Chemically-Induced Neoplasia in Rat and Mouse Models in National Toxicology Program Bioassays and Their Relevance to Human Cancer

PubMed Central

Hoenerhoff, Mark J.; Hong, Hue Hua; Ton, Tai-Vu; Lahousse, Stephanie A.; Sills, Robert C.

2012-01-01

Tumor response in the B6C3F1 mouse, F344 rat, and other animal models following exposure to various compounds provides evidence that people exposed to these or similar compounds may be at risk for developing cancer. Although tumors in rodents and humans are often morphologically similar, underlying mechanisms of tumorigenesis are often unknown and may be different between the species. Therefore, the relevance of an animal tumor response to human health would be better determined if the molecular pathogenesis were understood. The underlying molecular mechanisms leading to carcinogenesis are complex and involve multiple genetic and epigenetic events and other factors. To address the molecular pathogenesis of environmental carcinogens, we examine rodent tumors (e.g., lung, colon, mammary gland, skin, brain, mesothelioma) for alterations in cancer genes and epigenetic events that are associated with human cancer. Our NTP studies have identified several genetic alterations in chemically induced rodent neoplasms that are important in human cancer. Identification of such alterations in rodent models of chemical carcinogenesis caused by exposure to environmental contaminants, occupational chemicals, and other compounds lends further support that they are of potential human health risk. These studies also emphasize the importance of molecular evaluation of chemically induced rodent tumors for providing greater public health significance for NTP evaluated compounds. PMID:19846892

Molecular cloning, sequence identification and tissue expression profile of three novel sheep (Ovis aries) genes - BCKDHA, NAGA and HEXA.

PubMed

Liu, G Y; Gao, S Z

2009-01-01

The complete coding sequences of three sheep genes- BCKDHA, NAGA and HEXA were amplified using the reverse transcriptase polymerase chain reaction (RT-PCR), based on the conserved sequence information of the mouse or other mammals. The nucleotide sequences of these three genes revealed that the sheep BCKDHA gene encodes a protein of 313 amino acids which has high homology with the BCKDHA gene that encodes a protein of 447 amino acids that has high homology with the Branched chain keto acid dehydrogenase El, alpha polypeptide (BCKDHA) of five species chimpanzee (93%), human (96%), crab-eating macaque (93%), bovine (98%) and mouse (91%). The sheep NAGA gene encodes a protein of 411 amino acids that has high homology with the alpha-N-acetylgalactosaminidase (NAGA) of five species human (85%), bovine (94%), mouse (91%), rat (83%) and chicken (74%). The sheep HEXA gene encodes a protein of 529 amino acids that has high homology with the hexosaminidase A(HEXA) of five species bovine (98%), human (84%), Bornean orangután (84%), rat (80%) and mouse (81%). Finally these three novel sheep genes were assigned to GenelDs: 100145857, 100145858 and 100145856. The phylogenetic tree analysis revealed that the sheep BCKDHA, NAGA, and HEXA all have closer genetic relationships to the BCKDHA, NAGA, and HEXA of bovine. Tissue expression profile analysis was also carried out and results revealed that sheep BCKDHA, NAGA and HEXA genes were differentially expressed in tissues including muscle, heart, liver, fat, kidney, lung, small and large intestine. Our experiment is the first to establish the primary foundation for further research on these three sheep genes.

Rodent heart failure models do not reflect the human circulating microRNA signature in heart failure.

PubMed

Vegter, Eline L; Ovchinnikova, Ekaterina S; Silljé, Herman H W; Meems, Laura M G; van der Pol, Atze; van der Velde, A Rogier; Berezikov, Eugene; Voors, Adriaan A; de Boer, Rudolf A; van der Meer, Peter

2017-01-01

We recently identified a set of plasma microRNAs (miRNAs) that are downregulated in patients with heart failure in comparison with control subjects. To better understand their meaning and function, we sought to validate these circulating miRNAs in 3 different well-established rat and mouse heart failure models, and correlated the miRNAs to parameters of cardiac function. The previously identified let-7i-5p, miR-16-5p, miR-18a-5p, miR-26b-5p, miR-27a-3p, miR-30e-5p, miR-199a-3p, miR-223-3p, miR-423-3p, miR-423-5p and miR-652-3p were measured by means of quantitative real time polymerase chain reaction (qRT-PCR) in plasma samples of 8 homozygous TGR(mREN2)27 (Ren2) transgenic rats and 8 (control) Sprague-Dawley rats, 6 mice with angiotensin II-induced heart failure (AngII) and 6 control mice, and 8 mice with ischemic heart failure and 6 controls. Circulating miRNA levels were compared between the heart failure animals and healthy controls. Ren2 rats, AngII mice and mice with ischemic heart failure showed clear signs of heart failure, exemplified by increased left ventricular and lung weights, elevated end-diastolic left ventricular pressures, increased expression of cardiac stress markers and reduced left ventricular ejection fraction. All miRNAs were detectable in plasma from rats and mice. No significant differences were observed between the circulating miRNAs in heart failure animals when compared to the healthy controls (all P>0.05) and no robust associations with cardiac function could be found. The previous observation that miRNAs circulate in lower levels in human patients with heart failure could not be validated in well-established rat and mouse heart failure models. These results question the translation of data on human circulating miRNA levels to experimental models, and vice versa the validity of experimental miRNA data for human heart failure.

Cloning, cell-type specificity, and regulatory function of the mouse alpha(1B)-adrenergic receptor promoter.

PubMed

Zuscik, M J; Piascik, M T; Perez, D M

1999-12-01

The functionality of a 3422-base pair promoter fragment from the mouse alpha(1B)-adrenergic receptor (alpha(1B)AR) gene was examined. This fragment, cloned from a mouse genomic library, was found to have significant sequence homology to the known human and rat alpha(1B)AR promoters. However, the consensus motif of several key cis-acting elements is not conserved among the rat, human, and mouse genes, suggesting species specificity. Confirming fidelity of the murine promoter, robust in vitro expression of a chloramphenicol acetyltransferase (CAT) reporter was detected in known alpha(1B)AR-expressing BC(3)H1, NB41A3, and DDT(1)MF-2 cells transiently transfected with a promoter-CAT construct. Conversely, minimal CAT expression was detected in known alpha(1B)AR-negative RAT-1 and R3T3 cells. These findings were extended by transfecting the same promoter-CAT construct into various primary cell types. In support of the hypothesis that alpha(1)ARs are differentially expressed in the smooth muscle of the vasculature, primary cultures of superior mesenteric and renal artery vascular smooth muscle cells showed significantly stronger CAT expression than did vascular smooth muscle cells derived from pulmonary, femoral, and iliac arteries. Primary osteoblastic bone-forming cells, which are known to be alpha(1B)AR negative, showed minimal CAT expression. Indicating regulatory function through cis-acting elements, RAT-1, R3T3, NB41A3, BC(3)H1, and DDT(1)MF2 cells transfected with the promoter-CAT construct all showed increased CAT production when challenged with forskolin or hypoxic conditions. Additionally, tissue-specific regulation of the promoter was observed when cells were simultaneously challenged with both forskolin and hypoxia. These results collectively demonstrate that a 3.4-kb PvuII fragment of the murine alpha(1B)AR gene promoter can: 1) drive tissue-specific production of a CAT reporter in both clonal and primary cell lines; and 2) confer tissue-specific regulation of that CAT reporter when induced by challenge with forskolin and/or hypoxic conditions.

Generation and Characterization of Siglec-F-Specific Monoclonal Antibodies.

PubMed

Shahmohammadi-Farid, Sima; Ghods, Roya; Jeddi-Tehrani, Mahmood; Bayat, Ali-Ahmad; Mojtabavi, Nazanin; Razavi, Alireza; Zarnani, Amir-Hassan

2017-12-01

Siglec-F (SF) is a surface glycoprotein expressed by mouse eosinophils and induces caspase- and mitochondria-dependent apoptosis after engagement with its cognate ligand or specific antibodies. This targeting eosinophils by monoclonal antibodies may help diverse diseases associated with increased frequency of eosinophils including allergy and asthma. In this paper, production of murine and rat monoclonal antibodies (mAbs) against Siglec-F has been addressed. Balb/c mice were immunized with siglec-F1 (SF1) and siglec-F2 (SF2) synthetic peptides conjugated to a carrier protein. Rats were immunized with Chinese hamster ovary CHO cells overexpressing Siglec-F (CHO-SF) or with Siglec-F-human immunoglobulin FC fusion protein (CHO-SF-Ig). Hybridomas were produced by standard protocol and screened for their reactivity by enzyme-linked immunosorbent assay (ELISA), western blotting (WB), and flow cytometry. In parallel, polyclonal antibodies were generated in New Zealand White rabbits immunized with SF1 and SF2 peptides. Three mouse and three rat mAbs were generated against synthetic peptides and SF-Ig, respectively. All mouse monoclonal and rabbit polyclonal antibodies reacted well with immunizing molecules in ELISA and detected specific band of Siglec-F in WB. However, they failed to detect native molecule in flow cytometry analysis. Quite the contrary, rat mAbs did not reacted with the denatured protein in WB, instead exhibited significant reactivity with CHO-SF cells in flow cytometry. Based on the heavily glycosylated nature of Siglec-F, it seems that generation of anti-SF antibodies able to detect native protein needs a properly folded molecule for immunization. Monoclonal antibodies reported here are invaluable tools for studying linear and conformation epitopes of SF and tracing mouse eosinophils.

Xenobiotic-metabolizing enzymes in the skin of rat, mouse, pig, guinea pig, man, and in human skin models.

PubMed

Oesch, F; Fabian, E; Guth, K; Landsiedel, R

2014-12-01

The exposure of the skin to medical drugs, skin care products, cosmetics, and other chemicals renders information on xenobiotic-metabolizing enzymes (XME) in the skin highly interesting. Since the use of freshly excised human skin for experimental investigations meets with ethical and practical limitations, information on XME in models comes in the focus including non-human mammalian species and in vitro skin models. This review attempts to summarize the information available in the open scientific literature on XME in the skin of human, rat, mouse, guinea pig, and pig as well as human primary skin cells, human cell lines, and reconstructed human skin models. The most salient outcome is that much more research on cutaneous XME is needed for solid metabolism-dependent efficacy and safety predictions, and the cutaneous metabolism comparisons have to be viewed with caution. Keeping this fully in mind at least with respect to some cutaneous XME, some models may tentatively be considered to approximate reasonable closeness to human skin. For dermal absorption and for skin irritation among many contributing XME, esterase activity is of special importance, which in pig skin, some human cell lines, and reconstructed skin models appears reasonably close to human skin. With respect to genotoxicity and sensitization, activating XME are not yet judgeable, but reactive metabolite-reducing XME in primary human keratinocytes and several reconstructed human skin models appear reasonably close to human skin. For a more detailed delineation and discussion of the severe limitations see the "Overview and Conclusions" section in the end of this review.

Gain in Transcriptional Activity by Primate-specific Coevolution of Melanoma Antigen-A11 and Its Interaction Site in Androgen Receptor*

PubMed Central

Liu, Qiang; Su, Shifeng; Blackwelder, Amanda J.; Minges, John T.; Wilson, Elizabeth M.

2011-01-01

Male sex development and growth occur in response to high affinity androgen binding to the androgen receptor (AR). In contrast to complete amino acid sequence conservation in the AR DNA and ligand binding domains among mammals, a primate-specific difference in the AR NH2-terminal region that regulates the NH2- and carboxyl-terminal (N/C) interaction enables direct binding to melanoma antigen-A11 (MAGE-11), an AR coregulator that is also primate-specific. Human, mouse, and rat AR share the same NH2-terminal 23FQNLF27 sequence that mediates the androgen-dependent N/C interaction. However, the mouse and rat AR FXXLF motif is flanked by Ala33 that evolved to Val33 in primates. Human AR Val33 was required to interact directly with MAGE-11 and for the inhibitory effect of the AR N/C interaction on activation function 2 that was relieved by MAGE-11. The functional importance of MAGE-11 was indicated by decreased human AR regulation of an androgen-dependent endogenous gene using lentivirus short hairpin RNAs and by the greater transcriptional strength of human compared with mouse AR. MAGE-11 increased progesterone and glucocorticoid receptor activity independently of binding an FXXLF motif by interacting with p300 and p160 coactivators. We conclude that the coevolution of the AR NH2-terminal sequence and MAGE-11 expression among primates provides increased regulatory control over activation domain dominance. Primate-specific expression of MAGE-11 results in greater steroid receptor transcriptional activity through direct interactions with the human AR FXXLF motif region and indirectly through steroid receptor-associated p300 and p160 coactivators. PMID:21730049

Interspecies scaling of a camptothecin analogue: human predictions for intravenous topotecan using animal data.

PubMed

Ahlawat, P; Srinivas, N R

2008-11-01

As a class, camptothecin analogues via market entry of topotecan and irinotecan, have shown promise for the treatment of various solid tumours. Topotecan, in particular, was chosen as the substrate for allometric scaling and prediction of human parameter values for both total clearance (CL) and volume of distribution (V(ss)). The availability of published data in mouse, rat, dog, and monkey paved the way for interspecies scaling via allometry. Although it appeared that at a minimum mouse, rat, and dog would reasonably fit in a three-species allometry scale-up, the inclusion of monkey data enabled a better prediction of the human parameter values for total topotecan-e.g., CL: allometric equation: 1.5234W(0.7865); predicted value = 43.04 l h(-1): observed CL = 24-53 l h(-1); V(ss): allometric equation: 1.1939W(1.0208); predicted value = 91.29 litres: observed V(ss) = 66-146 litres. The proximity of the allometric exponent values of CL (0.7885) and V(ss) (1.0208) to the suggested values of 0.75 and 1.00 was not only encouraging, but also confirmed the applicability of interspecies scaling approach for topotecan. The data suggest that allometric scaling approaches with suitable correction factors could potentially be used to predict the human pharmacokinetics of novel CPT analogues prospectively.

Genomic sequences of murine gamma B- and gamma C-crystallin-encoding genes: promoter analysis and complete evolutionary pattern of mouse, rat and human gamma-crystallins.

PubMed

Graw, J; Liebstein, A; Pietrowski, D; Schmitt-John, T; Werner, T

1993-12-22

The murine genes, gamma B-cry and gamma C-cry, encoding the gamma B- and gamma C-crystallins, were isolated from a genomic DNA library. The complete nucleotide (nt) sequences of both genes were determined from 661 and 711 bp, respectively, upstream from the first exon to the corresponding polyadenylation sites, comprising more than 2650 and 2890 bp, respectively. The new sequences were compared to the partial cDNA sequences available for the murine gamma B-cry and gamma C-cry, as well as to the corresponding genomic sequences from rat and man, at both the nt and predicted amino acid (aa) sequence levels. In the gamma B-cry promoter region, a canonical CCAAT-box, a TATA-box, putative NF-I and C/EBP sites were detected. An R-repeat is inserted 366 bp upstream from the transcription start point. In contrast, the gamma C-cry promoter does not contain a CCAAT-box, but some other putative binding sites for transcription factors (AP-2, UBP-1, LBP-1) were located by computer analysis. The promoter regions of all six gamma-cry from mouse, rat and human, except human psi gamma F-cry, were analyzed for common sequence elements. A complex sequence element of about 70-80 bp was found in the proximal promoter, which contains a gamma-cry-specific and almost invariant sequence (crygpel) of 14 nt, and ends with the also invariant TATA-box. Within the complex sequence element, a minimum of three further features specific for the gamma A-, gamma B- and gamma D/E/F-cry genes can be defined, at least two of which were recently shown to be functional. In addition to these four sequence elements, a subtype-specific structure of inverted repeats with different-sized spacers can be deduced from the multiple sequence alignment. A phylogenetic analysis based on the promoter region, as well as the complete exon 3 of all gamma-cry from mouse, rat and man, suggests separation of only five gamma-cry subtypes (gamma A-, gamma B-, gamma C-, gamma D- and gamma E/F-cry) prior to species separation.

Single body parts are processed by individual neurons in the mouse dorsolateral striatum.

PubMed

Coffey, Kevin R; Nader, Miles; West, Mark O

2016-04-01

Interest in the dorsolateral striatum (DLS) has generated numerous scientific studies of its neuropathologies, as well as its roles in normal sensorimotor integration and learning. Studies are informed by knowledge of DLS functional organization, the guiding principle being its somatotopic afferent projections from primary somatosensory (S1) and motor (M1) cortices. The potential to connect behaviorally relevant function to detailed structure is elevated by mouse models, which have access to extensive genetic neuroscience tool kits. Remaining to be demonstrated, however, is whether the correspondence between S1/M1 corticostriatal terminal distributions and the physiological properties of DLS neurons demonstrated in rats and non-human primates exists in mice. Given that the terminal distribution of S1/M1 projections to the DLS in mice is similar to that in rats, we studied whether firing rates (FRs) of DLS neurons in awake, behaving mice are related to activity of individual body parts. MSNs exhibited robust, selective increases in FR during movement or somatosensory stimulation of single body parts. Properties of MSNs, including baseline FRs, locations, responsiveness to stimulation, and proportions of responsive neurons were similar to properties observed in rats. Future studies can be informed by the present demonstration that the mouse lateral striatum functions as a somatic sensorimotor sector of the striatum and appears to be a homolog of the primate putamen, as demonstrated in rats (Carelli and West, 1991). Copyright © 2016 Elsevier B.V. All rights reserved.

E3024, 3-but-2-ynyl-5-methyl-2-piperazin-1-yl-3,5-dihydro-4H-imidazo[4,5-d]pyridazin-4-one tosylate, is a novel, selective and competitive dipeptidyl peptidase-IV inhibitor.

PubMed

Yasuda, Nobuyuki; Nagakura, Tadashi; Inoue, Takashi; Yamazaki, Kazuto; Katsutani, Naruo; Takenaka, Osamu; Clark, Richard; Matsuura, Fumiyoshi; Emori, Eita; Yoshikawa, Seiji; Kira, Kazunobu; Ikuta, Hironori; Okada, Toshimi; Saeki, Takao; Asano, Osamu; Tanaka, Isao

2006-10-24

Dipeptidyl peptidase IV (DPP-IV) inhibitors are expected to become a useful new class of anti-diabetic agent. The aim of the present study is to characterize the in vitro and in vivo profile of E3024, 3-but-2-ynyl-5-methyl-2-piperazin-1-yl-3,5-dihydro-4H-imidazo[4,5-d]pyridazin-4-one tosylate, which is a novel imidazopyridazinone-derived DPP-IV inhibitor. E3024 inhibited recombinant human and mouse DPP-IV with IC50 values of approximately 100 nM. E3024 inhibited DPP-IV in human, mouse, rat and canine plasma with IC50 values of 140 to 400 nM. In contrast, E3024 did not inhibit DPP-8 or DPP-9 activity. Kinetic analysis indicated that E3024 is a competitive DPP-IV inhibitor. In Zucker fa/fa rats, E3024 (1 mg/kg) reduced glucose excursion after glucose load, with increases in plasma insulin and active glucagon-like peptide-1 levels. In fasted rats, this compound did not cause hypoglycemia. In a rat 4-week toxicological study, no notable changes were found at doses up to 750 mg/kg. The present preclinical studies indicate that E3024 is a novel selective DPP-IV inhibitor with anti-diabetic effects and a good safety profile.

Comparison of TCDD-elicited genome-wide hepatic gene expression in Sprague–Dawley rats and C57BL/6 mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nault, Rance; Kim, Suntae; Zacharewski, Timothy R., E-mail: tzachare@msu.edu

2013-03-01

Although the structure and function of the AhR are conserved, emerging evidence suggests that downstream effects are species-specific. In this study, rat hepatic gene expression data from the DrugMatrix database (National Toxicology Program) were compared to mouse hepatic whole-genome gene expression data following treatment with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). For the DrugMatrix study, male Sprague–Dawley rats were gavaged daily with 20 μg/kg TCDD for 1, 3 and 5 days, while female C57BL/6 ovariectomized mice were examined 1, 3 and 7 days after a single oral gavage of 30 μg/kg TCDD. A total of 649 rat and 1386 mouse genes (|fold change| ≥more » 1.5, P1(t) ≥ 0.99) were differentially expressed following treatment. HomoloGene identified 11,708 orthologs represented across the rat Affymetrix 230 2.0 GeneChip (12,310 total orthologs), and the mouse 4 × 44K v.1 Agilent oligonucleotide array (17,578 total orthologs). Comparative analysis found 563 and 922 orthologs differentially expressed in response to TCDD in the rat and mouse, respectively, with 70 responses associated with immune function and lipid metabolism in common to both. Moreover, QRTPCR analysis of Ceacam1, showed divergent expression (induced in rat; repressed in mouse) functionally consistent with TCDD-elicited hepatic steatosis in the mouse but not the rat. Functional analysis identified orthologs involved in nucleotide binding and acetyltransferase activity in rat, while mouse-specific responses were associated with steroid, phospholipid, fatty acid, and carbohydrate metabolism. These results provide further evidence that TCDD elicits species-specific regulation of distinct gene networks, and outlines considerations for future comparisons of publicly available microarray datasets. - Highlights: ► We performed a whole-genome comparison of TCDD-regulated genes in mice and rats. ► Previous species comparisons were extended using data from the DrugMatrix database. ► Less than 15% of TCDD-regulated orthologs were common to mice and rats. ► Considerations for the comparison of publicly available datasets are described.« less

Safety assessment of boron by application of new uncertainty factors and their subdivision.

PubMed

Hasegawa, Ryuichi; Hirata-Koizumi, Mutsuko; Dourson, Michael L; Parker, Ann; Ono, Atsushi; Hirose, Akihiko

2013-02-01

The available toxicity information for boron was reevaluated and four appropriate toxicity studies were selected in order to derive a tolerable daily intake (TDI) using newly proposed uncertainty factors (UFs) presented in Hasegawa et al. (2010). No observed adverse effect levels (NOAELs) of 17.5 and 8.8 mgB/kg/day for the critical effect of testicular toxicity were found in 2-year rat and dog feeding studies. Also, the 95% lower confidence limit of the benchmark doses for 5% reduction of fetal body weight (BMDL(05)) was calculated as 44.9 and 10.3 mgB/kg/day in mouse and rat developmental toxicity studies, respectively. Measured values available for differences in boron clearance between rats and humans and variability in the glomerular filtration rate (GFR) in pregnant women were used to derive chemical specific UFs. For the remaining uncertainty, newly proposed default UFs, which were derived from the latest applicable information with a probabilistic approach, and their subdivided factors for toxicokinetic and toxicodynamic variability were applied. Finally, overall UFs were calculated as 68 for rat testicular toxicity, 40 for dog testicular toxicity, 247 for mouse developmental toxicity and 78 for rat developmental toxicity. It is concluded that 0.13 mgB/kg/day is the most appropriate TDI for boron, based on rat developmental toxicity. Copyright © 2012 Elsevier Inc. All rights reserved.

The human is an exception to the evolutionarily-conserved phenomenon of pre-fertilization zona pellucida resistance to proteolysis induced by oviductal fluid.

PubMed

Mondéjar, I; Avilés, M; Coy, P

2013-03-01

Is zona pellucida (ZP) resistance to proteolysis, induced by oviductal fluid (OF), a mechanism common to species other than the pig and cow? ZP resistance to proteolysis induced by OF was observed in the mouse, rat, hamster, rabbit, sheep, goat, pig and cow, but not in humans. Oviductal ZP resistance to proteolysis occurs in the pig and cow where it influences the incidence of fertilization and polyspermy. The effect is observed after incubation of ZP in OFs from pig (pOF), cow (cOF), rabbit (rOF) and sheep (sOF). Oocytes from nine different species, including ungulates, rodents, lagomorphs and primates were incubated in rOF, sOF, gOF, cOF, pOF and human oviductal fluid (hOF). ZP digestion times for the matured oocytes of these nine species, without any treatment or incubated in 5 (mouse, rat, hamster, rabbit, cow, ewe and goat) or 6 (pig and humans) of the OFs collected were compared using three replicates per treatment and at least three oocytes per replicate. In vivo matured oocytes from rat, hamster, mouse, rabbit and humans, in vitro matured oocytes from cow, goat, ewe and pig and rOF, cOF, gOF, sOF, pOF and human (hOF) were collected and processed for the study. Oocytes from each species were incubated in the different OFs for 30 min. The resistance of the ZP of the oocytes to enzymatic digestion in a pronase solution (0.5% in PBS) was measured and registered as ZP digestion time. rOF increased ZP resistance to proteolytic digestion in the range of between 96 and 720 h for any of the species tested, whereas the corresponding increase in human ZP was only 1 min. OFs from the remaining species also had a significant effect, with variations among the cross-species experiments (P < 0.05). hOF, which was only tested on human and porcine oocytes, had no effect on ZP chemical hardening. Measurements of ZP digestion times are not of extreme accuracy and errors of a few seconds can be assumed in the experimental data. However, when differences are in the range of hours among treatments, variations measured in seconds do not alter the robustness of the findings. Human oocytes and OF were of limited access, compared with oocytes from species collected in slaughterhouses. OFs from mouse, rat and hamster were not tested due to the small size of the genital tract in these species and the small volume of fluid available. Since oviductal modification of ZP resistance to proteolytic digestion has been demonstrated to influence fertilization and this pre-fertilization mechanism is considered to contribute to the control of polyspermy, the apparent absence of this mechanism in humans suggests that the regulation of polyspermy depends mainly on other mechanisms, most probably of cortical granule origin. Investigation into a possible relationship between the lack of oviductal ZP hardening in human oocytes and the existence of tubal ectopic pregnancies in this species is proposed. This work was supported by the Spanish Ministry of Science and Innovation and FEDER, Grant AGL2009-12512-C02-01-02. The authors declare no competing interest.

RECENT ADVANCES IN ARSENIC CARCINOGENESIS: MODES OF ACTION, ANIMAL MODEL SYSTEMS AND METHYLATED ARSENIC METABOLITES

EPA Science Inventory

Abstract:

Recent advances in our knowledge of arsenic carcinogenesis include the development of rat or mouse models for all human organs in which inorganic arsenic is known to cause cancer -skin, lung, urinary bladder, liver and kidney. Tumors can be produced from eit...

Molecular dynamics in drug design: new generations of compstatin analogs.

PubMed

Tamamis, Phanourios; López de Victoria, Aliana; Gorham, Ronald D; Bellows-Peterson, Meghan L; Pierou, Panayiota; Floudas, Christodoulos A; Morikis, Dimitrios; Archontis, Georgios

2012-05-01

We report the computational and rational design of new generations of potential peptide-based inhibitors of the complement protein C3 from the compstatin family. The binding efficacy of the peptides is tested by extensive molecular dynamics-based structural and physicochemical analysis, using 32 atomic detail trajectories in explicit water for 22 peptides bound to human, rat or mouse target protein C3, with a total of 257 ns. The criteria for the new design are: (i) optimization for C3 affinity and for the balance between hydrophobicity and polarity to improve solubility compared to known compstatin analogs; and (ii) development of dual specificity, human-rat/mouse C3 inhibitors, which could be used in animal disease models. Three of the new analogs are analyzed in more detail as they possess strong and novel binding characteristics and are promising candidates for further optimization. This work paves the way for the development of an improved therapeutic for age-related macular degeneration, and other complement system-mediated diseases, compared to known compstatin variants. © 2012 John Wiley & Sons A/S.

Dipeptidyl peptidase-4 greatly contributes to the hydrolysis of vildagliptin in human liver.

PubMed

Asakura, Mitsutoshi; Fujii, Hideaki; Atsuda, Koichiro; Itoh, Tomoo; Fujiwara, Ryoichi

2015-04-01

The major metabolic pathway of vildagliptin in mice, rats, dogs, and humans is hydrolysis at the cyano group to produce a carboxylic acid metabolite M20.7 (LAY151), whereas the major metabolic enzyme of vildagliptin has not been identified. In the present study, we determined the contribution rate of dipeptidyl peptidase-4 (DPP-4) to the hydrolysis of vildagliptin in the liver. We performed hydrolysis assay of the cyano group of vildagliptin using mouse, rat, and human liver samples. Additionally, DPP-4 activities in each liver sample were assessed by DPP-4 activity assay using the synthetic substrate H-glycyl-prolyl-7-amino-4-methylcoumarin (Gly-Pro-AMC). M20.7 formation rates in liver microsomes were higher than those in liver cytosol. M20.7 formation rate was significantly positively correlated with the DPP-4 activity using Gly-Pro-AMC in liver samples (r = 0.917, P < 0.01). The formation of M20.7 in mouse, rat, and human liver S9 fraction was inhibited by sitagliptin, a selective DPP-4 inhibitor. These findings indicate that DPP-4 is greatly involved in vildagliptin hydrolysis in the liver. Additionally, we established stable single expression systems of human DPP-4 and its R623Q mutant, which is the nonsynonymous single-nucleotide polymorphism of human DPP-4, in human embryonic kidney 293 (HEK293) cells to investigate the effect of R623Q mutant on vildagliptin-hydrolyzing activity. M20.7 formation rate in HEK293 cells expressing human DPP-4 was significantly higher than that in control HEK293 cells. Interestingly, R623Q mutation resulted in a decrease of the vildagliptin-hydrolyzing activity. Our findings might be useful for the prediction of interindividual variability in vildagliptin pharmacokinetics. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

Fractionation and reconstitution of factors required for accurate transcription of mammalian ribosomal RNA genes: identification of a species-dependent initiation factor.

PubMed Central

Mishima, Y; Financsek, I; Kominami, R; Muramatsu, M

1982-01-01

Mouse and human cell extracts (S100) can support an accurate and efficient transcription initiation on homologous ribosomal RNA gene (rDNA) templates. The cell extracts were fractionated with the aid of a phosphocellulose column into four fractions (termed A, B, C and D), including one containing a major part of the RNA polymerase I activity. Various reconstitution experiments indicate that fraction D is an absolute requirement for the correct and efficient transcription initiation by RNA polymerase I on both mouse and human genes. Fraction B effectively suppresses random initiation on these templates. Fraction A appears to further enhance the transcription which takes place with fractions C and D. Although fractions A, B and C are interchangeable between mouse and human extracts, fraction D is not; i.e. initiation of transcription required the presence of a homologous fraction D for both templates. The factor(s) in fraction D, however, is not literally species-specific, since mouse D fraction is capable of supporting accurate transcription initiation on a rat rDNA template in the presence of all the other fractions from human cell extract under the conditions where human D fraction is unable to support it. We conclude from these experiments that a species-dependent factor in fraction D plays an important role in the initiation of rDNA transcription in each animal species. Images PMID:7177852

Cell cycle, differentiation and tissue-independent expression of ribosomal protein L37.

PubMed

Su, S; Bird, R C

1995-09-15

A unique human cDNA (hG1.16) that encodes a mRNA of 450 nucleotides was isolated from a subtractive library derived from HeLa cells. The relative expression level of hG1.16 during different cell-cycle phases was determined by Northern-blot analysis of cells synchronized by double-thymidine block and serum deprivation/refeeding. hG1.16 was constitutively expressed during all phases of the cell cycle, including the quiescent phase when even most constitutively expressed genes experience some suppression of expression. The expression level of hG1.16 did not change during terminal differentiation of myoblasts to myotubes, during which cells become permanently post-mitotic. Examination of other tissues revealed that the relative expression level of hG1.16 was constitutive in all embryonic mouse tissues examined, including brain, eye, heart, kidney, liver, lung and skeletal muscle. This was unusual in that expression was not down-modulated during differentiation and did not vary appreciably between tissue types. Analysis by inter-species Northern-blot analysis revealed that hG1.16 was highly conserved among all vertebrates studied (from fish to humans but not in insects). DNA sequence analysis of hG1.16 revealed a high level of similarity to rat ribosomal protein L37, identifying hG1.16 as a new member of this multigene family. The deduced amino acid sequence of hG1.16 was identical to rat ribosomal protein L37 that contained 97 amino acids, many of which are highly positively charged (15 arginine and 14 lysine residues with a predicted M(r) of 11,065). hG1.16 protein has a single C2-C2 zinc-finger-like motif which is also present in rat ribosomal protein L37. Using primers designed from the sequence of hG1.16, unique bovine and rat cDNAs were also isolated by 5'-rapid-amplification of cDNA ends. DNA sequences of bovine and rat G1.16, clones were 92.8% and 92.2% similar to human G1.16 while the deduced amino acid sequences derived from bovine and rat cDNAs each differed by a single amino acid from the sequence of hG1.16 and the published rat L37 sequence. Southern-blot analysis revealed that hG1.16 exists in multiple copies in human, rat and mouse genomes. These G1.16 clones encode unique human, rat and bovine members of the ribosomal protein L37 gene family, which are constitutively expressed even during transitions from quiescence to active cell proliferation or terminal differentiation, in all tissues and all vertebrates investigated.

Cloning of rat amelotin and localization of the protein to the basal lamina of maturation stage ameloblasts and junctional epithelium.

PubMed

Moffatt, Pierre; Smith, Charles E; St-Arnaud, René; Simmons, Darrin; Wright, J Timothy; Nanci, Antonio

2006-10-01

Formation of tooth enamel is a very complex process in which a specific set of proteins secreted by ameloblasts play a primordial role. As part of a screening procedure to identify novel proteins secreted by EO (enamel organ) cells of rat incisors, we isolated a partial cDNA fragment (EO-017) that is the homologue of the recently described mouse Amtn (amelotin) gene [Iwasaki, Bajenova, Somogyi-Ganss, Miller, Nguyen, Nourkeyhani, Gao, Wendel and Ganss (2005) J. Dent. Res. 84, 1127-1132]. Presented herein is the cloning of rat and pig full-length cDNAs with their deduced protein sequences. Detailed expression profiling by Northern-blot analysis and RT (reverse transcriptase)-PCR on rat and mouse tissues revealed highest expression in the mandible, more specifically in the maturation stage of the EO. Among all tissues tested, low expression was detected only in periodontal ligament, lung, thymus and gingiva. In silico analyses revealed that the Amtn gene is highly conserved in seven other mammals, but is absent from fish, birds and amphibians. The Amtn protein is enriched in proline, leucine, glutamine and threonine (52% of total) and contains a perfectly conserved protein kinase CK2 phosphorylation site. Transient transfection experiments in HEK-293 cells (human embryonic kidney cells) showed that secreted Amtn is post-translationally modified possibly through O-linked oligosaccharides on threonine residues. In concordance with its predominant expression site, immunofluorescence localization within the rat and mouse mandibles revealed Amtn localized to the basal lamina of maturation stage ameloblasts of incisors and unerupted molars. Intense Amtn protein expression was also detected in the internal basal lamina of junctional epithelium in molars. The peculiar and unique cellular localization of Amtn suggests a role in cell adhesion.

Human hepatocytes support the hypertrophic but not the hyperplastic response to the murine nongenotoxic hepatocarcinogen sodium phenobarbital in an in vivo study using a chimeric mouse with humanized liver.

PubMed

Yamada, Tomoya; Okuda, Yu; Kushida, Masahiko; Sumida, Kayo; Takeuchi, Hayato; Nagahori, Hirohisa; Fukuda, Takako; Lake, Brian G; Cohen, Samuel M; Kawamura, Satoshi

2014-11-01

High doses of sodium phenobarbital (NaPB), a constitutive androstane receptor (CAR) activator, have been shown to produce hepatocellular tumors in rodents by a mitogenic mode of action (MOA) involving CAR activation. The effect of 1-week dietary treatment with NaPB on liver weight and histopathology, hepatic CYP2B enzyme activity and CYP2B/3A mRNA expression, replicative DNA synthesis and selected genes related to cell proliferation, and functional transcriptomic and metabolomic analyses was studied in male CD-1 mice, Wistar Hannover (WH) rats, and chimeric mice with human hepatocytes. The treatment of chimeric mice with 1000-1500-ppm NaPB resulted in plasma levels around 3-5-fold higher than those observed in human subjects given therapeutic doses of NaPB. NaPB produced dose-dependent increases in hepatic CYP2B activity and CYP2B/3A mRNA levels in all animal models. Integrated functional metabolomic and transcriptomic analyses demonstrated that the responses to NaPB in the human liver were clearly different from those in rodents. Although NaPB produced a dose-dependent increase in hepatocyte replicative DNA synthesis in CD-1 mice and WH rats, no increase in replicative DNA synthesis was observed in human hepatocyte-originated areas of chimeric mice. In addition, treatment with NaPB had no effect on Ki-67, PCNA, GADD45β, and MDM2 mRNA expression in chimeric mice, whereas significant increases were observed in CD-1 mice and/or WH rats. However, increases in hepatocyte replicative DNA synthesis were observed in chimeric mice both in vivo and in vitro after treatment epidermal growth factor. Thus, although NaPB could activate CAR in both rodent and human hepatocytes, NaPB did not increase replicative DNA synthesis in human hepatocytes of chimeric mice, whereas it was mitogenic to rat and mouse hepatocytes. As human hepatocytes are refractory to the mitogenic effects of NaPB, the MOA for NaPB-induced rodent liver tumor formation is thus not relevant for humans. © The Author 2014. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Inflammation, Prostate Cancer and Negative Regulation of Androgen Receptor Expression

DTIC Science & Technology

2009-05-01

Chem. 282(32):23716-24 2. Taganov, K.D., Boldin , M.P., Chang, K.J., and Baltimore, D. (2006) NF-kappaB- dependent induction of microRNA miR-146, an...produced by the rat liver nuclear extract revealed four protein-binding sites (Fig. 5A). Site I is especially note- worthy because it includes an NF-B-like...liver nuclear extract . B, Sequence conservation of rat and mouse AR promoters with the human AR promoter at 144 to 204. An NF-B-like element in

Olfaction variation in mouse husbandry and its implications for refinement and standardization: UK survey of animal scents.

PubMed

López-Salesansky, Noelia; Mazlan, Nur H; Whitfield, Lucy E; Wells, Dominic J; Burn, Charlotte C

2016-10-01

Olfaction plays a crucial role in mouse communication, providing information about genetic identity, physiological status of conspecifics and alerting mice to potential predators. Scents of animal origin can trigger physiological and behavioural responses that could affect experimental responses and impact positively or negatively on mouse welfare. Additionally, differing olfactory profiles could help explain variation in results between laboratories. A survey was sent to animal research units in the UK to investigate potential transfer of scents of animal origin during routine husbandry procedures, and responses were obtained from animal care workers and researchers using mice in 51 institutions. The results reveal great diversity between animal units regarding the relevant husbandry routines covered. Most [71%] reported housing non-breeding male and female mice in the same room, with 76% reporting that hands were not washed and gloves not changed between handling male and female mice. The most commonly reported species housed in the same facility as mice was the rat (91%), and 41% of respondents were aware that scents from rats could affect mice. Changing of gloves between handling mice and other species was reported by 79% of respondents. Depending on the aspect considered, between 18 and 33% of respondents believed human and non-human animal odours would strongly affect mouse physiology, behaviour or standardization, while approximately 32-54% believed these effects would be weak. This indicates uncertainty regarding the significance of these factors. Understanding and controlling these practices could reduce unwanted variability in experimental results and maximize welfare. © The Author(s) 2015.

Relationship Among Tau Antigens Isolated from Various Lines of Simian Virus 40-Transformed Cells

PubMed Central

Simmons, Daniel T.; Martin, Malcolm A.; Mora, Peter T.; Chang, Chungming

1980-01-01

In addition to the virus-specified tumor antigens, simian virus 40-transformed cells contain at least one other protein which can be immunoprecipitated with serum from animals bearing simian virus 40-induced tumors. This protein, which is designated Tau antigen, has an apparent molecular weight of 56,000 as determined by electrophoresis on acrylamide gels. The relationship among Tau antigens isolated from different lines of simian virus 40-transformed cells was examined by comparing the methionine-labeled tryptic peptides of these proteins by two-dimensional fingerprinting on thin-layer cellulose plates. In this fashion, we initially determined that the Tau antigens isolated from three different lines of transformed mouse cells were very similar. Second, we found that Tau antigen isolated from a line of rat transformants was closely related, but not identical, to the mouse cell Tau antigens. Approximately 70% of their methionine peptides comigrated in two dimensions. Finally, we showed that Tau antigen isolated from a line of transformed human cells was only partially related to the mouse and rat proteins. About 40% of the methionine peptides of the human protein were also contained in the Tau antigens from the other two species. These results strongly indicate that the Tau antigens isolated from these various simian virus 40-transformed cell lines contain common amino acid sequences. Images PMID:6247503

Identification, characterization and potent antitumor activity of ECO-4601, a novel peripheral benzodiazepine receptor ligand.

PubMed

Gourdeau, Henriette; McAlpine, James B; Ranger, Maxime; Simard, Bryan; Berger, Francois; Beaudry, Francis; Farnet, Chris M; Falardeau, Pierre

2008-05-01

ECO-4601 is a structurally novel farnesylated dibenzodiazepinone discovered through DECIPHER technology, Thallion's proprietary drug discovery platform. The compound was shown to have a broad cytotoxic activity in the low micromolar range when tested in the NCI 60 cell line panel. In the work presented here, ECO-4601 was further evaluated against brain tumor cell lines. Preliminary mechanistic studies as well as in vivo antitumor evaluation were performed. Since ECO-4601 has a benzodiazepinone moiety, we first investigated if it binds the central and/or peripheral benzodiazepine receptors. ECO-4601 was tested in radioligand binding assays on benzodiazepine receptors obtained from rat hearts. The ability of ECO-4601 to inhibit the growth of CNS cancers was evaluated on a panel of mouse, rat and human glioma cell lines using a standard MTT assay. Antitumor efficacy studies were performed on gliomas (rat and human), human breast and human prostate mouse tumor xenografts. Antitumor activity and pharmacokinetic analysis of ECO-4601 was evaluated following intravenous (i.v.), subcutaneous (s.c.), and intraperitoneal (i.p.) bolus administrations. ECO-4601 was shown to bind the peripheral but not the central benzodiazepine receptor and inhibited the growth of CNS tumor cell lines. Bolus s.c. and i.p. administration gave rise to low but sustained drug exposure, and resulted in moderate to significant antitumor activity at doses that were well tolerated. In a rat glioma (C6) xenograft model, ECO-4601 produced up to 70% tumor growth inhibition (TGI) while in a human glioma (U-87MG) xenograft, TGI was 34%. Antitumor activity was highly significant in both human hormone-independent breast (MDA-MB-231) and prostate (PC-3) xenografts, resulting in TGI of 72 and 100%, respectively. On the other hand, i.v. dosing was followed by rapid elimination of the drug and was ineffective. Antitumor efficacy of ECO-4601 appears to be associated with the exposure parameter AUC and/or sustained drug levels rather than C (max). These in vivo data constitute a rationale for clinical studies testing prolonged continuous administration of ECO-4601.

Atherosclerotic lesions in mouse and man: is it the same disease?

PubMed

Bentzon, Jacob Fog; Falk, Erling

2010-10-01

Genetically-engineered mice with hyperlipidemia are the most widely used atherosclerosis models today, but recent advances in transgenesis open the possibility to create new models in alternative species, such as the rat and pig. It seems relevant at this point in time to review some of the strengths and weaknesses of the mouse. The histology of lesion development in mouse and man has more similarities than differences, and comparative genetics show that many mechanisms of murine and human atherogenesis are shared. Unfortunately, the most feared complication of human atherosclerosis, that is, plaque rupture and thrombosis, occur extremely rarely in mice. This is a major problem. Most patients today are not treated before symptoms ensue, and at this late stage of the disease, mechanisms identified during plaque development in the mouse may not be very important. Murine atherosclerosis models are highly valuable for identifying atherogenic mechanisms that can be targeted by preventive medicine. However, models with thrombotic complications and large animal models suitable for interventional procedures and imaging would be more supportive for current clinical practice and are highly wanted.

An analysis of pharmaceutical experience with decades of rat carcinogenicity testing: support for a proposal to modify current regulatory guidelines.

PubMed

Sistare, Frank D; Morton, Daniel; Alden, Carl; Christensen, Joel; Keller, Douglas; Jonghe, Sandra De; Storer, Richard D; Reddy, M Vijayaraj; Kraynak, Andrew; Trela, Bruce; Bienvenu, Jean-Guy; Bjurström, Sivert; Bosmans, Vanessa; Brewster, David; Colman, Karyn; Dominick, Mark; Evans, John; Hailey, James R; Kinter, Lewis; Liu, Matt; Mahrt, Charles; Marien, Dirk; Myer, James; Perry, Richard; Potenta, Daniel; Roth, Arthur; Sherratt, Philip; Singer, Thomas; Slim, Rabih; Soper, Keith; Fransson-Steen, Ronny; Stoltz, James; Turner, Oliver; Turnquist, Susan; van Heerden, Marjolein; Woicke, Jochen; DeGeorge, Joseph J

2011-06-01

Data collected from 182 marketed and nonmarketed pharmaceuticals demonstrate that there is little value gained in conducting a rat two-year carcinogenicity study for compounds that lack: (1) histopathologic risk factors for rat neoplasia in chronic toxicology studies, (2) evidence of hormonal perturbation, and (3) positive genetic toxicology results. Using a single positive result among these three criteria as a test for outcome in the two-year study, fifty-two of sixty-six rat tumorigens were correctly identified, yielding 79% test sensitivity. When all three criteria were negative, sixty-two of seventy-six pharmaceuticals (82%) were correctly predicted to be rat noncarcinogens. The fourteen rat false negatives had two-year study findings of questionable human relevance. Applying these criteria to eighty-six additional chemicals identified by the International Agency for Research on Cancer as likely human carcinogens and to drugs withdrawn from the market for carcinogenicity concerns confirmed their sensitivity for predicting rat carcinogenicity outcome. These analyses support a proposal to refine regulatory criteria for conducting a two-year rat study to be based on assessment of histopathologic findings from a rat six-month study, evidence of hormonal perturbation, genetic toxicology results, and the findings of a six-month transgenic mouse carcinogenicity study. This proposed decision paradigm has the potential to eliminate over 40% of rat two-year testing on new pharmaceuticals without compromise to patient safety.

Muro-Neuro-Urodynamics; a Review of the Functional Assessment of Mouse Lower Urinary Tract Function.

PubMed

Ito, Hiroki; Pickering, Anthony E; Igawa, Yasuhiko; Kanai, Anthony J; Fry, Christopher H; Drake, Marcus J

2017-01-01

Background: Mouse urodynamic tests are fundamental to understanding normal lower urinary tract (LUT) function. These experiments also contribute to our understanding of neurological dysfunction, pathophysiological processes, and potential mechanisms of therapy. Objectives: Systematic assessment of published evidence on urodynamics, advantages and limitations of different urodynamic measurements in mice, and consideration of potential implications for the clinical field. Methods: A search using specific search-terms for urodynamic studies and mice was conducted on PubMed (from inception to 1 July 2016). Results: We identified 55 studies examining or describing mouse neuro-urodynamics. We summarize reported features of mouse urodynamic function deriving from frequency-volume chart (FVC) measurements, voiding spot assays, filling cystometry, and pressure-flow studies. Similarly, an influence of the diurnal cycle on voiding is observed in mice and should be considered when interpreting rodent urodynamic studies, especially FVC measurements and voiding spot assays. Anaesthesia, restraint conditions, or filling rate influence mouse neuro-urodynamics. Mouse cystometric studies have observed intravesical pressure oscillations that accompany urine flow, attributed to high frequency opening and closing of the urethra. This characterization is not seen in other species, except rats. In contrast to human clinical urodynamics, the terminology of these examinations has not been standardized although many rodent urodynamic studies have been described. Conclusion: Mice have many anatomical and physiological similarities to humans and they are generally cost effective, and allow investigation of the effects of aging because of their short lifespan. There are some differences between mouse and human urodynamics. These must be considered when interpreting LUT function in mice, and translational value of murine disease models.

A Replication-incompetent Rift Valley Fever Vaccine: Chimeric Virus-like Particles Protect Mice and Rats Against Lethal Challenge

PubMed Central

Mandell, Robert B.; Koukuntla, Ramesh; Mogler, Laura J. K.; Carzoli, Andrea K.; Freiberg, Alexander N.; Holbrook, Michael R.; Martin, Brian K.; Staplin, William R.; Vahanian, Nicholas N.; Link, Charles J.; Flick, Ramon

2009-01-01

Virus-like particles (VLPs) present viral antigens in a native conformation and are effectively recognized by the immune system and therefore are considered as suitable and safe vaccine candidates against many viral diseases. Here we demonstrate that chimeric VLPs containing Rift Valley fever virus (RVFV) glycoproteins GN and GC, nucleoprotein N and the gag protein of Moloney murine leukemia virus represent an effective vaccine candidate against Rift Valley fever, a deadly disease in humans and livestock. Long-lasting humoral and cellular immune responses are demonstrated in a mouse model by the analysis of neutralizing antibody titers and cytokine secretion profiles. Vaccine efficacy studies were performed in mouse and rat lethal challenge models resulting in high protection rates. Taken together, these results demonstrate that replication-incompetent chimeric RVF VLPs are an efficient RVFV vaccine candidate. PMID:19932911

Functional cardiac magnetic resonance microscopy

NASA Astrophysics Data System (ADS)

Brau, Anja Christina Sophie

2003-07-01

The study of small animal models of human cardiovascular disease is critical to our understanding of the origin, progression, and treatment of this pervasive disease. Complete analysis of disease pathophysiology in these animal models requires measuring structural and functional changes at the level of the whole heart---a task for which an appropriate non-invasive imaging method is needed. The purpose of this work was thus to develop an imaging technique to support in vivo characterization of cardiac structure and function in rat and mouse models of cardiovascular disease. Whereas clinical cardiac magnetic resonance imaging (MRI) provides accurate assessment of the human heart, the extension of cardiac MRI from humans to rodents presents several formidable scaling challenges. Acquiring images of the mouse heart with organ definition and fluidity of contraction comparable to that achieved in humans requires an increase in spatial resolution by a factor of 3000 and an increase in temporal resolution by a factor of ten. No single technical innovation can meet the demanding imaging requirements imposed by the small animal. A functional cardiac magnetic resonance microscopy technique was developed by integrating improvements in physiological control, imaging hardware, biological synchronization of imaging, and pulse sequence design to achieve high-quality images of the murine heart with high spatial and temporal resolution. The specific methods and results from three different sets of imaging experiments are presented: (1) 2D functional imaging in the rat with spatial resolution of 175 mum2 x 1 mm and temporal resolution of 10 ms; (2) 3D functional imaging in the rat with spatial resolution of 100 mum 2 x 500 mum and temporal resolution of 30 ms; and (3) 2D functional imaging in the mouse with spatial resolution down to 100 mum2 x 1 mm and temporal resolution of 10 ms. The cardiac microscopy technique presented here represents a novel collection of technologies capable of acquiring routine high-quality images of murine cardiac structure and function with minimal artifacts and markedly higher spatial resolution compared to conventional techniques. This work is poised to serve a valuable role in the evaluation of cardiovascular disease and should find broad application in studies ranging from basic pathophysiology to drug discovery.

Aurora-A as a Modifier of Breast Cancer Risk in BRCA 1/2 Mutation Carriers

DTIC Science & Technology

2007-06-01

Dieter Schaefer, Institute of Human Genetics, University of Frankfurt, Frankfurt, Germany; Norbert Arnold, University of Schleswig- Holstein , Campus...Intron 2 Opossum Mouse Rat Cow Dog Intron 1 Figure 3 | The FGFR2 locus. a, Map of the whole FGFR2 gene, viewed relative to common SNPs on HapMap

A Single Amino Acid Difference between Mouse and Human 5-Lipoxygenase Activating Protein (FLAP) Explains the Speciation and Differential Pharmacology of Novel FLAP Inhibitors.

PubMed

Blevitt, Jonathan M; Hack, Michael D; Herman, Krystal; Chang, Leon; Keith, John M; Mirzadegan, Tara; Rao, Navin L; Lebsack, Alec D; Milla, Marcos E

2016-06-10

5-Lipoxygenase activating protein (FLAP) plays a critical role in the metabolism of arachidonic acid to leukotriene A4, the precursor to the potent pro-inflammatory mediators leukotriene B4 and leukotriene C4 Studies with small molecule inhibitors of FLAP have led to the discovery of a drug binding pocket on the protein surface, and several pharmaceutical companies have developed compounds and performed clinical trials. Crystallographic studies and mutational analyses have contributed to a general understanding of compound binding modes. During our own efforts, we identified two unique chemical series. One series demonstrated strong inhibition of human FLAP but differential pharmacology across species and was completely inactive in assays with mouse or rat FLAP. The other series was active across rodent FLAP, as well as human and dog FLAP. Comparison of rodent and human FLAP amino acid sequences together with an analysis of a published crystal structure led to the identification of amino acid residue 24 in the floor of the putative binding pocket as a likely candidate for the observed speciation. On that basis, we tested compounds for binding to human G24A and mouse A24G FLAP mutant variants and compared the data to that generated for wild type human and mouse FLAP. These studies confirmed that a single amino acid mutation was sufficient to reverse the speciation observed in wild type FLAP. In addition, a PK/PD method was established in canines to enable preclinical profiling of mouse-inactive compounds. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

An interspecies comparison of mercury inhibition on muscarinic acetylcholine receptor binding in the cerebral cortex and cerebellum

DOE Office of Scientific and Technical Information (OSTI.GOV)

Basu, Niladri; Department of Natural Resource Sciences, McGill University, Ste.-Anne-de-Bellevue, Quebec, H9X 3V9; Stamler, Christopher J.

2005-05-15

Mercury (Hg) is a ubiquitous pollutant that can disrupt neurochemical signaling pathways in mammals. It is well documented that inorganic Hg (HgCl{sub 2}) and methyl Hg (MeHg) can inhibit the binding of radioligands to the muscarinic acetylcholine (mACh) receptor in rat brains. However, little is known concerning this relationship in specific anatomical regions of the brain or in other species, including humans. The purpose of this study was to explore the inhibitory effects of HgCl{sub 2} and MeHg on [{sup 3}H]-quinuclidinyl benzilate ([{sup 3}H]-QNB) binding to the mACh receptor in the cerebellum and cerebral cortex regions from human, rat, mouse,more » mink, and river otter brain tissues. Saturation binding curves were obtained from each sample to calculate receptor density (B {sub max}) and ligand affinity (K {sub d}). Subsequently, samples were exposed to HgCl{sub 2} or MeHg to derive IC50 values and inhibition constants (K {sub i}). Results demonstrate that HgCl{sub 2} is a more potent inhibitor of mACh receptor binding than MeHg, and the receptors in the cerebellum are more sensitive to Hg-mediated mACh receptor inhibition than those in the cerebral cortex. Species sensitivities, irrespective of Hg type and brain region, can be ranked from most to least sensitive: river otter > rat > mink > mouse > humans. In summary, our data demonstrate that Hg can inhibit the binding [{sup 3}H]-QNB to the mACh receptor in a range of mammalian species. This comparative study provides data on interspecies differences and a framework for interpreting results from human, murine, and wildlife studies.« less

Differences in social interaction- vs. cocaine reward in mouse vs. rat.

PubMed

Kummer, Kai K; Hofhansel, Lena; Barwitz, Constanze M; Schardl, Aurelia; Prast, Janine M; Salti, Ahmad; El Rawas, Rana; Zernig, Gerald

2014-01-01

We previously developed rat experimental models based on the conditioned place preference (CPP) paradigm in which only four 15-min episodes of dyadic social interaction with a sex- and weight-matched male Sprague Dawley (SD) rat (1) reversed CPP from cocaine to social interaction despite continuing cocaine training, and (2) prevented the reacquisition/re-expression of cocaine CPP. In a concurrent conditioning schedule, pairing one compartment with social interaction and the other compartment with 15 mg/kg cocaine injections, rats spent the same amount of time in both compartments and the most rewarding sensory component of the composite stimulus social interaction was touch (taction). In the present study, we validated our experimental paradigm in C57BL/6 mice to investigate if our experimental paradigm may be useful for the considerable number of genetically modified mouse models. Only 71% of the tested mice developed place preference for social interaction, whereas 85% of the rats did. Accordingly, 29% of the mice developed conditioned place aversion (CPA) to social interaction, whereas this was true for only 15% of the rats. In support of the lesser likelihood of mice to develop a preference for social interaction, the average amount of time spent in direct contact was 17% for mice vs. 79% for rats. In animals that were concurrently conditioned for social interaction vs. cocaine, the relative reward strength for cocaine was 300-fold higher in mice than in rats. Considering that human addicts regularly prefer drugs of abuse to drug-free social interaction, the present findings suggest that our experimental paradigm of concurrent CPP for cocaine vs. social interaction is of even greater translational power if performed in C57BL/6 mice, the genetic background for most transgenic rodent models, than in rats.

Mammalian cDNA Library from the NIH Mammalian Gene Collection (MGC) | Office of Cancer Genomics

Cancer.gov

The MGC provides the research community full-length clones for most of the defined (as of 2006) human and mouse genes, along with selected clones of cow and rat genes. Clones were designed to allow easy transfer of the ORF sequences into nearly any type of expression vector. MGC provides protein ‘expression-ready’ clones for each of the included human genes. MGC is part of the ORFeome Collaboration (OC).

Isolation and Growth of Prostate Stem Cells and Establishing Cancer Cell Lines from Human Prostate Tumors

DTIC Science & Technology

2009-05-01

contaminating rat UGSE cells ; and regions of host mouse glands were either from circulating pluripotent stem cells or local epithelial cells which were...CONTRACT NUMBER Isolation and Growth of Prostate Stem Cells and Establishing Cancer Cell Lines from Human Prostate Tumors 5b. GRANT NUMBER 81WXH...NOTES 14. ABSTRACT The objective of this proposal was to isolate, grow, and characterize normal prostate stem cells and establish new prostate

Gene discovery in the hamster: a comparative genomics approach for gene annotation by sequencing of hamster testis cDNAs

PubMed Central

Oduru, Sreedhar; Campbell, Janee L; Karri, SriTulasi; Hendry, William J; Khan, Shafiq A; Williams, Simon C

2003-01-01

Background Complete genome annotation will likely be achieved through a combination of computer-based analysis of available genome sequences combined with direct experimental characterization of expressed regions of individual genomes. We have utilized a comparative genomics approach involving the sequencing of randomly selected hamster testis cDNAs to begin to identify genes not previously annotated on the human, mouse, rat and Fugu (pufferfish) genomes. Results 735 distinct sequences were analyzed for their relatedness to known sequences in public databases. Eight of these sequences were derived from previously unidentified genes and expression of these genes in testis was confirmed by Northern blotting. The genomic locations of each sequence were mapped in human, mouse, rat and pufferfish, where applicable, and the structure of their cognate genes was derived using computer-based predictions, genomic comparisons and analysis of uncharacterized cDNA sequences from human and macaque. Conclusion The use of a comparative genomics approach resulted in the identification of eight cDNAs that correspond to previously uncharacterized genes in the human genome. The proteins encoded by these genes included a new member of the kinesin superfamily, a SET/MYND-domain protein, and six proteins for which no specific function could be predicted. Each gene was expressed primarily in testis, suggesting that they may play roles in the development and/or function of testicular cells. PMID:12783626

Comparative metabolism of honokiol in mouse, rat, dog, monkey, and human hepatocytes.

PubMed

Jeong, Hyeon-Uk; Kim, Ju-Hyun; Kong, Tae Yeon; Choi, Won Gu; Lee, Hye Suk

2016-04-01

Honokiol has antitumor, antioxidative, anti-inflammatory, and antithrombotic effects. Here we aimed to identify the metabolic profile of honokiol in mouse, rat, dog, monkey, and human hepatocytes and to characterize the enzymes responsible for the glucuronidation and sulfation of honokiol. Honokiol had a high hepatic extraction ratio in all five species, indicating that it was extensively metabolized. A total of 32 metabolites, including 17 common and 15 different metabolites, produced via glucuronidation, sulfation, and oxidation of honokiol allyl groups were tentatively identified using liquid chromatography-high resolution quadrupole Orbitrap mass spectrometry. Glucuronidation of honokiol to M8 (honokiol-4-glucuronide) and M9 (honokiol-2'-glucuronide) was the predominant metabolic pathway in hepatocytes of all five species; however, interspecies differences between 4- and 2'-glucuronidation of honokiol were observed. UGT1A1, 1A8, 1A9, 2B15, and 2B17 played major roles in M8 formation, whereas UGT1A7 and 1A9 played major roles in M9 formation. Human cDNA-expressed SULT1C4 played a major role in M10 formation (honokiol-2'-sulfate), whereas SULT1A1*1, 1A1*2, and 1A2 played major roles in M11 formation (honokiol-4-sulfate). In conclusion, honokiol metabolism showed interspecies differences.

Cellular biophysics during freezing of rat and mouse sperm predicts post-thaw motility.

PubMed

Hagiwara, Mie; Choi, Jeung Hwan; Devireddy, Ramachandra V; Roberts, Kenneth P; Wolkers, Willem F; Makhlouf, Antoine; Bischof, John C

2009-10-01

Though cryopreservation of mouse sperm yields good survival and motility after thawing, cryopreservation of rat sperm remains a challenge. This study was designed to evaluate the biophysics (membrane permeability) of rat in comparison to mouse to better understand the cooling rate response that contributes to cryopreservation success or failure in these two sperm types. In order to extract subzero membrane hydraulic permeability in the presence of ice, a differential scanning calorimeter (DSC) method was used. By analyzing rat and mouse sperm frozen at 5 degrees C/min and 20 degrees C/min, heat release signatures characteristic of each sperm type were obtained and correlated to cellular dehydration. The dehydration response was then fit to a model of cellular water transport (dehydration) by adjusting cell-specific biophysical (membrane hydraulic permeability) parameters L(pg) and E(Lp). A "combined fit" (to 5 degrees C/min and 20 degrees C/min data) for rat sperm in Biggers-Whitten-Whittingham media yielded L(pg) = 0.007 microm min(-1) atm(-1) and E(Lp) = 17.8 kcal/mol, and in egg yolk cryopreservation media yielded L(pg) = 0.005 microm min(-1) atm(-1) and E(Lp) = 14.3 kcal/mol. These parameters, especially the activation energy, were found to be lower than previously published parameters for mouse sperm. In addition, the biophysical responses in mouse and rat sperm were shown to depend on the constituents of the cryopreservation media, in particular egg yolk and glycerol. Using these parameters, optimal cooling rates for cryopreservation were predicted for each sperm based on a criteria of 5%-15% normalized cell water at -30 degrees C during freezing in cryopreservation media. These predicted rates range from 53 degrees C/min to 70 degrees C/min and from 28 degrees C/min to 36 degrees C/min in rat and mouse, respectively. These predictions were validated by comparison to experimentally determined cryopreservation outcomes, in this case based on motility. Maximum motility was obtained with freezing rates between 50 degrees C/min and 80 degrees C/min for rat and at 20 degrees C/min with a sharp drop at 50 degrees C/min for mouse. In summary, DSC experiments on mouse and rat sperm yielded a difference in membrane permeability parameters in the two sperm types that, when implemented in a biophysical model of water transport, reasonably predict different optimal cooling rate outcomes for each sperm after cryopreservation.

Breast Milk Jaundice: Effect of 3α 20β-pregnanediol on Bilirubin Conjugation by Human Liver

PubMed Central

Adlard, B. P. F.; Lathe, G. H.

1970-01-01

The effect of 3α,20β-pregnanediol and other steroids on bilirubin conjugation was examined using liver tissue from human and four other species. Neither 3α,20β-pregnanediol nor 3α,20β-pregnanediol inhibited conjugation by human liver slices or by solubilized human liver microsomes. 3α,20β-pregnanediol is unlikely to be the inhibitor causing breast milk jaundice. Oestriol inhibited conjugation by human liver slices. A comparison of species indicated that the response of the human liver slice system to steroids resembles that of the rabbit and guinea-pig rather than the rat or mouse. PMID:4246186

Shifts in an invasive rodent community favoring black rats (Rattus rattus) following restoration of native forest

USGS Publications Warehouse

Shiels, Aaron B.; Medeiros, Arthur C.; von Allmen, Erica I.

2017-01-01

One potential, unintended ecological consequence accompanying forest restoration is a shift in invasive animal populations, potentially impacting conservation targets. Eighteen years after initial restoration (ungulate exclusion, invasive plant control, and out planting native species) at a 4 ha site on Maui, Hawai'i, we compared invasive rodent communities in a restored native dry forest and adjacent non-native grassland. Quarterly for 1 year, we trapped rodents on three replicate transects (107 rodent traps) in each habitat type for three consecutive nights. While repeated trapping may have reduced the rat (Black rat, Rattus rattus) population in the forest, it did not appear to reduce the mouse (House mouse, Mus musculus) population in the grassland. In unrestored grassland, mouse captures outnumbered rat captures 220:1, with mice averaging 54.9 indiv./night versus rats averaging 0.25 indiv./night. In contrast, in restored native forest, rat captures outnumbered mouse captures by nearly 5:1, averaging 9.0 indiv./night versus 1.9 indiv./night for mice. Therefore, relatively recent native forest restoration increased Black rat abundance and also increased their total biomass in the restored ecosystem 36-fold while reducing House mouse biomass 35-fold. Such a community shift is worrisome because Black rats pose a much greater threat than do mice to native birds and plants, perhaps especially to large-seeded tree species. Land managers should be aware that forest restoration (i.e. converting grassland to native forest) can invoke shifts in invasive rodent populations, potentially favoring Black rats. Without intervention, this shift may pose risks for intended conservation targets and modify future forest restoration trajectories.

A comparative evaluation of the tissue responses associated with polymeric implants in the rat and mouse.

PubMed

Kidd, Kameha R; Dal Ponte, Donny B; Kellar, Robert S; Williams, Stuart K

2002-03-15

End product application is an important consideration when evaluating a material in an in vivo setting (Didisheim, Cardiovasc Pathol 1993;2:1S-2S). Small animal models allow high through-put evaluation of biocompatability. Previous preclinical evaluations have often used a rat subcutaneous model for the characterization of material-tissue interaction. Recent advances in genetic manipulation have provided mouse models with selective expression of a wide range of critical proteins. The rat model does not have many of the resources (i.e., knockouts, SCID, nude) that are present in mouse strains. The availability of these mice provides a resource to delineate the mechanisms regulating the healing associated with implants. However, before the mouse models can be used, they must be validated with respect to their ability to accurately assess tissue responses to materials. In this study the tissue responses after the implantation of expanded polytetrafluoroethylene (ePTFE) were compared between rat and mouse. Discs of ePTFE (30-microm internodal distance) were implanted in subcutaneous and epididymal fat tissue of rats (Sprague-Dawley) and mice (129-SVJ). After 5 weeks the samples were removed and evaluated for vascular density, inflammation, and fibrous encapsulation. No difference in the vessel density was observed within the peri-implant subcutaneous and adipose tissue or within the porous material. However, a significant difference was found in the number of activated macrophages and giant cells between these two species. Implants in the rat exhibited greater numbers of activated inflammatory cells in the peri-implant tissue. The data indicate that the mouse and rat provide a comparable model for evaluating angiogenesis and neovascularization associated with synthetic porous implants. Copyright 2001 John Wiley & Sons, Inc. J Biomed Mater Res 59: 682-689, 2002

In Vitro Transformation of Rat and Mouse Cells by DNA from Simian Virus 40

PubMed Central

Abrahams, P. J.; van der Eb, A. J.

1975-01-01

Primary rat kidney cells and mouse 3T3 cells can be transformed by DNA of simian virus 40 when use is made of the calcium technique (Graham and van der Eb, 1973). The transformation assay in primary rat cells is reproducible, but the dose response is not linear. PMID:166204

Propagation of Human Hepatocytes in uPA/SCID Mice: Producing Chimeric Mice with Humanized Liver.

PubMed

Ohshita, Hiroki; Tateno, Chise

2017-01-01

Primary or cryopreserved human hepatocytes (h-heps) have been used as the gold standard for in vitro metabolism and hepatotoxicity studies; however, the supply of h-heps is limited and they cannot grow in vitro. We achieved approximately 1000-fold propagation of h-heps in the liver of albumin promoter/enhancer-driven urokinase-type plasminogen activator transgenic/severe combined immunodeficiency disease (uPA/SCID) mice with genetically induced liver disease and immunodeficiency. When h-heps are transplanted into the uPA/SCID mouse liver via the spleen, the h-heps engraft in the mouse liver, resulting in its repopulation with h-heps. We have named this model "chimeric mouse with humanized liver, PXB-mouse ® ." Fresh h-heps can be isolated from the chimeric mice (PXB-cells ® ) and have been used for in vitro studies.The efficacy and safety of chemical entities for use in humans are estimated using experimental animals such as rats and mice. The drug development of many chemical entities has been halted because of metabolic differences between humans and animals during clinical studies. Therefore, chimeric mice with humanized liver have been used to predict human-type metabolism and safety conditions for h-heps. In addition, until recently there were no suitable hepatitis B or C virus (HBV or HCV) susceptible animal models aside from chimpanzees. Chimeric mice are the sole persistent infectious small animal model for HBV and HCV and they have been used to investigate the efficacy of new anti-HBV or HCV agents.In this chapter, we describe a method for producing chimeric mice with humanized liver using uPA/SCID mice.

Distribution of vesicular glutamate transporter 1 (VGLUT1) in the mouse esophagus.

PubMed

Kraus, T; Neuhuber, W L; Raab, M

2007-08-01

In rat and mouse esophagus, vesicular glutamate transporter 2 (VGLUT2) has been demonstrated to identify vagal intraganglionic laminar endings (IGLEs); this has recently also been shown for VGLUT1 in rat esophagus. In this study, we have investigated the distribution of VGLUT1 in the mouse esophagus and compared these results with the recently published data from the rat esophagus. Unexpectedly, we have discovered that VGLUT1 mostly fails to identify IGLEs in the mouse esophagus. This is surprising, since the distribution of VGLUT2 shows comparable results in both species. Confocal imaging has revealed substantial colocalization of VGLUT1 immunoreactivity (-ir) with cholinergic and nitrergic/peptidergic markers within the myenteric neuropil and in both cholinergic and nitrergic myenteric neuronal cell bodies. VGLUT1 and cholinergic markers have also been colocalized in fibers of the muscularis mucosae, whereas VGLUT1 and nitrergic markers have never been colocalized in fibers of the muscularis mucosae, although this does occur in fibers of the muscularis running to motor endplates. Thus, VGLUT1 is contained in the nitrergic innervation of mouse esophageal motor endplates, another difference from the rat esophagus. VGLUT1-ir is therefore present in extrinsic and intrinsic innervation of the mouse esophagus, but the significant differences from the rat indicate species variations concerning the distribution of VGLUTs in the peripheral nervous system.

Based on an analysis of mode of action, styrene-induced mouse lung tumors are not a human cancer concern.

PubMed

Cruzan, George; Bus, James S; Andersen, Melvin E; Carlson, Gary P; Banton, Marcy I; Sarang, Satinder S; Waites, Robbie

2018-06-01

Based on 13 chronic studies, styrene exposure causes lung tumors in mice, but no tumor increases in other organs in mice or rats. Extensive research into the mode of action demonstrates the key events and human relevance. Key events are: metabolism of styrene by CYP2F2 in mouse lung club cells to ring-oxidized metabolites; changes in gene expression for metabolism of lipids and lipoproteins, cell cycle and mitotic M-M/G1 phases; cytotoxicity and mitogenesis in club cells; and progression to preneoplastic/neoplastic lesions in lung. Although styrene-7,8-oxide (SO) is a common genotoxic styrene metabolite in in vitro studies, the data clearly demonstrate that SO is not the proximate toxicant and that styrene does not induce a genotoxic mode of action. Based on complete attenuation of styrene short-term and chronic toxicity in CYP2F2 knockout mice and similar attenuation in CYP2F1 (humanized) transgenic mice, limited metabolism of styrene in human lung by CYP2F1, 2 + orders of magnitude lower SO levels in human lung compared to mouse lung, and lack of styrene-related increase in lung cancer in humans, styrene does not present a risk of cancer to humans. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Transcranial Near-Infrared Laser Transmission (NILT) Profiles (800 nm): Systematic Comparison in Four Common Research Species

PubMed Central

Lapchak, Paul A.; Boitano, Paul D.; Butte, Pramod V.; Fisher, David J.; Hölscher, Thilo; Ley, Eric J.; Nuño, Miriam; Voie, Arne H.; Rajput, Padmesh S.

2015-01-01

Background and Purpose Transcranial near-infrared laser therapy (TLT) is a promising and novel method to promote neuroprotection and clinical improvement in both acute and chronic neurodegenerative diseases such as acute ischemic stroke (AIS), traumatic brain injury (TBI), and Alzheimer’s disease (AD) patients based upon efficacy in translational animal models. However, there is limited information in the peer-reviewed literature pertaining to transcranial near-infrared laser transmission (NILT) profiles in various species. Thus, in the present study we systematically evaluated NILT characteristics through the skull of 4 different species: mouse, rat, rabbit and human. Results Using dehydrated skulls from 3 animal species, using a wavelength of 800nm and a surface power density of 700 mW/cm2, NILT decreased from 40.10% (mouse) to 21.24% (rat) to 11.36% (rabbit) as skull thickness measured at bregma increased from 0.44 mm in mouse to 0.83 mm in rat and then 2.11 mm in rabbit. NILT also significantly increased (p<0.05) when animal skulls were hydrated (i.e. compared to dehydrated); but there was no measurable change in thickness due to hydration. In human calvaria, where mean thickness ranged from 7.19 mm at bregma to 5.91 mm in the parietal skull, only 4.18% and 4.24% of applied near-infrared light was transmitted through the skull. There was a slight (9.2-13.4%), but insignificant effect of hydration state on NILT transmission of human skulls, but there was a significant positive correlation between NILT and thickness at bregma and parietal skull, in both hydrated and dehydrated states. Conclusion This is the first systematic study to demonstrate differential NILT through the skulls of 4 different species; with an inverse relationship between NILT and skull thickness. With animal skulls, transmission profiles are dependent upon the hydration state of the skull, with significantly greater penetration through hydrated skulls compared to dehydrated skulls. Using human skulls, we demonstrate a significant correlation between thickness and penetration, but there was no correlation with skull density. The results suggest that TLT should be optimized in animals using novel approaches incorporating human skull characteristics, because of significant variance of NILT profiles directly related to skull thickness. PMID:26039354

Genomic Imprinting of the M6P/IGF2 Receptor: A Novel Breast Cancer Susceptibility Mechanism

DTIC Science & Technology

2000-07-01

i.e. echidna and platypus ), marsupials (i.e. opossum) and eutherian mammals (i.e. mouse, rat, pig, cow, bat, flying lemur, tree shrew, ringtail lemur...and humans). Our findings demonstrate that M6P/IGF2R is not imprinted in the egg-laying platypus and echidna, whereas it is imprinted in the opossum

Investigation of the metabolic biotransformation of substance P in liver microsomes by liquid chromatography quadrupole ion trap mass spectrometry.

PubMed

Pailleux, Floriane; Lemoine, Jérôme; Beaudry, Francis

2013-01-01

Substance P (SP) belongs to the tachykinin family and plays an essential role in pain transmission and in neurogenic inflammation. It can be detected in the central and peripheral nervous systems. The objectives of this study were to establish SP metabolic stability in liver microsomes in three species (rat, mouse and human), and identify and characterize SP metabolites by LC-MS/MS. Endogenous peptide metabolism is not well documented and this is particularly true for neuropeptides participating in neurogenic inflammation. In vitro, T(1/2) results in pooled liver microsomes were 9.2, 5.6 and 18.6 min for rat, mouse and human liver microsomes, respectively. Five major SP metabolites were identified and quantified, including C-terminal SP fragments SP(3-11) , SP(5-11) , SP(6-11) , SP(8-11) as well as N-terminal fragment SP(1-7) . The results suggest significant differences between species in SP metabolite concentrations. Consequently, the metabolic profile of each species is distinctive and may have a significant impact on biomolecular mechanisms involved in specific pathophysiological changes. Copyright © 2012 John Wiley & Sons, Ltd.

Exposure to perfluorooctane sulfonate during pregnancy in rat and mouse. II: postnatal evaluation

EPA Science Inventory

The postnatal effects of in utero exposure to perfluorooctane sulfonate (PFOS, C8F17SO3-) were evaluated in the rat and mouse. Pregnant Sprague-Dawley rats were given 1, 2, 3, 5, or 10 mg/kg PFOS daily by gavage from gestation day (GD) 2 to GD 21; pregnant CD-1 mice were treated ...

A defect in the inflammation-primed macrophage-activation cascade in osteopetrotic rats.

PubMed

Yamamoto, N; Lindsay, D D; Naraparaju, V R; Ireland, R A; Popoff, S N

1994-05-15

Macrophages were activated by administration of lysophosphatidylcholine (lyso-Pc) or dodecylglycerol (DDG) to wild-type rats but not in osteopetrotic (op) mutant rats. In vitro treatment of wild-type rat peritoneal cells with lyso-Pc or DDG efficiently activated macrophages whereas treatment of op mutant rat peritoneal cells with lyso-Pc or DDG did not activate macrophages. The inflammation-primed macrophage activation cascade in rats requires participation of B lymphocytes and vitamin D binding protein (DBP). Lyso-Pc-inducible beta-galactosidase of wild-type rat B lymphocytes can convert DBP to the macrophage-activating factor (MAF), whereas B lymphocytes of the op mutant rats were shown to be deficient in lyso-Pc-inducible beta-galactosidase. DBP is conserved among mammalian species. Treatment of human DBP (Gc1 protein) with commercial glycosidases yields an extremely high titrated MAF as assayed on mouse and rat macrophages. Because the enzymatically generated MAF (GcMAF) bypasses the role of lymphocytes in macrophage activation, the op mutant rat macrophages were efficiently activated by administration of a small quantity (100 pg/rat) of GcMAF. Likewise, in vitro treatment of op rat peritoneal cells with as little as 40 pg GcMAF/ml activated macrophages.

A Hemorrhagic Factor (Apicidin) Produced by Toxic Fusarium Isolates from Soybean Seeds

PubMed Central

Park, Jun-Suk; Lee, Kyung-Rim; Kim, Jin-Cheol; Lim, Sun-Hee; Seo, Jeong-Ah; Lee, Yin-Won

1999-01-01

Fifty-two isolates of Fusarium species were obtained from soybean seeds from various parts of Korea and identified as Fusarium oxysporum, F. moniliforme, F. semitectum, F. solani, F. graminearum, or F. lateritium. These isolates were grown on autoclaved wheat grains and examined for toxicity in a rat-feeding test. Nine cultures were toxic to rats. One of these, a culture of Fusarium sp. strain KCTC 16677, produced apicidin, an antiprotozoal agent that caused toxic effects in rats (including body weight loss; hemorrhage in the stomach, intestines, and bladder; and finally death) when rats were fed diets supplemented with 0.05 and 0.1% apicidin. The toxin was toxic to brine shrimp (the 50% lethal concentration was 40 μg/ml) and was weakly cytotoxic to human and mouse tumor cell lines. PMID:9872769

Rat chromosome 1: regional localization of seven genes (Slc9a3, Srd5a1, Esr, Tcp1, Grik5, Tnnt3, Jak2) and anchoring of the genetic linkage map to the cytogenetic map.

PubMed

Szpirer, C; Szpirer, J; Tissir, F; Stephanova, E; Vanvooren, P; Kurtz, T W; Iwai, N; Inagami, T; Pravenec, M; Kren, V; Klinga-Levan, K; Levan, G

1997-09-01

Seven genes were regionally localized on rat Chromosome (Chr) 1, from 1p11 to 1q42, and two of these genes were also included in a linkage map. This mapping work integrates the genetic linkage map and the cytogenetic map, and allows us to orient the linkage map with respect to the centromere, and to deduce the approximate position of the centromere in the linkage map. These mapping data also indicate that the Slc9a3 gene, encoding the Na+/H+ exchanger 3, is an unlikely candidate for the blood pressure loci assigned to rat Chr 1. These new localizations expand comparative mapping between rat Chr 1 and mouse or human chromosomes.

Genetic variation of an acid phosphatase (Acp-2) in the laboratory rat: possible homology with mouse AP-1 and human ACP2.

PubMed

Bender, K; Bissbort, S; Kuhn, A; Nagel, M; Günther, E

1986-02-01

A genetic locus controlling the electrophoretic mobility of an acid phosphatase in the rat (Rattus norvegicus) is described. The locus, designed Acp-2, is not expressed in erythrocytes but is expressed in all other tissues studied. The product of Acp-2 hydrolyzes a wide variety of phosphate monoesters and is inhibited by L(+)-tartaric acid. Inbred rat strains have fixed either allele Acp-2a or allele Acp-2b. Codominant expression is observed in the respective F1 hybrids. Backcross progenies revealed the expected 1:1 segregation ratio. Possible loose linkage was found between the Acp-2 and the Pep-3 gene loci at a recombination frequency of 0.36 +/- 0.06.

Oncolytic Rat Parvovirus H-1PV, a Candidate for the Treatment of Human Lymphoma: In Vitro and In Vivo Studies

PubMed Central

Angelova, Assia L; Aprahamian, Marc; Balboni, Ginette; Delecluse, Henri-Jacques; Feederle, Regina; Kiprianova, Irina; Grekova, Svitlana P; Galabov, Angel S; Witzens-Harig, Mathias; Ho, Anthony D; Rommelaere, Jean; Raykov, Zahari

2009-01-01

The incidence of lymphomas developing in both immunocompetent and immunosuppressed patients continues to steadily increase worldwide. Current chemotherapy and immunotherapy approaches have several limitations, such as severe side toxicity and selection of resistant cell variants. Autonomous parvoviruses (PVs), in particular the rat parvovirus H-1PV, have emerged as promising anticancer agents. Although it is apathogenic in humans, H-1PV has been shown to infect and suppress various rat and human tumors in animal models. In this study, we demonstrate the capacity of H-1PV for efficiently killing, through necrosis, cell cultures originating from Burkitt's lymphoma (BL), while sparing normal B lymphocytes. The cytotoxic effect was generally accompanied by a productive H-1PV infection. Remarkably, parvovirus-based monotherapy efficiently suppressed established BL at an advanced stage in a severe combined immunodeficient (SCID) mouse model of the disease. The data show for the first time that an oncolytic parvovirus deserves further consideration as a potential tool for the treatment of some non-Hodgkin B-cell lymphomas, including those resistant to apoptosis induction by rituximab. PMID:19367260

[Genome loses all 5-methylcytosine a life span. How is this connected with accumulation of mutations during aging?].

PubMed

Mazin, A L

1993-01-01

The 5-methylcytosine (5mC) content in liver DNA has been determined for rats of different age. The rate of the 5mC loss from DNA is maximal in pre- and neonatal rats, 1.28% of reduction of the 5mC content per day, then it decreases to 0.33% and becomes minimal and constant in adult rats, 0.028% per day. During pregnancy and the first 15 days of postnatal development rat genome loses 49% of all 5mC. Within the next 45 days 15% of 5mC disappears, and during maximal rat life span, about four years, 39% of the genomic 5mC may be lost. Thus, it has been found for the first time that the animal genome loses practically all 5mC residues during the life span. Analysis of the literature data shows that for embryos the rate of the 5mC loss from DNA proves to be higher than that for adult animals by 96 times for mice, 69-for rats and 28-for cows. The rate of embryonal DNA hypomethylation may be inversely proportional to the pregnancy duration of species. In adult animals the rate inversely correlates with their maximal life span and accounts for the 5mC loss from DNA of a mouse by 0.028%, of a rat by 0.024%, of a hamster by 0.007%, of a cow by 0.004% and of a human being by 0.0005% per day. During the entire ontogenesis, the genome of a mouse loses 93% of all 5mC residues, that of a rat-101% and of a cow-88%. The age-dependent loss of 5mC from DNA is also typical for cell lines aging in vitro. It is constant, as a rule, and correlates with the number of cell population doublings (PD). The removal of all 5mC from DNA corresponds to 70-130 PD for human, 40-60 PD-for hamster and 6 PD- for mouse cells. In immortal lines the level of DNA methylation is stable or grows with age. A possible mechanism of an age-related 5mC loss from DNA is discussed. DNA hypomethylation may result from 5mC deamination directly at the moment of replicative DNA methylation and subsequent reparation of the G.T mispairs which leads to accumulation of the 5mC-->T+C substitutions in the genome with each cell division. So DNA methylation may serve as an ideal mechanism for counting cell divisions in vivo and in vitro.(ABSTRACT TRUNCATED AT 400 WORDS)

Recognition of the Species of Origin of Cells in Culture by Mixed Agglutination

PubMed Central

Coombs, R. R. A.; Daniel, Mary R.; Gurner, B. W.; Kelus, A.

1961-01-01

Preliminary experiment on the mixed agglutination reaction suggests that this reaction will afford a useful method for identifying the species of origin of cells maintained in culture. The reaction depends on the presence of antigens characteristic of the species, common to both tissue cells and red cells. Culture cells derived from man, ox, pig and rat could be distinguished one from the other. Fibroblasts of the mouse may be differentiated from those of the rat by means of a rat anti-mouse red-cell serum or a mouse anti-rat red-cell serum. Experiments are reported on trial absorption procedures to render the sera completely species-specific in their reactions. ImagesFIG. 1 PMID:13695283

A Mouse Model to Evaluate the Impact of Species, Sex, and Lipid Load on Lymphatic Drug Transport

PubMed Central

Caliph, Suzanne M.; Nguyen, Gary; Tso, Patrick; Charman, William N.

2014-01-01

Purpose To establish a lymph-cannulated mouse model, and use the model to investigate the impact of lipid dose on exogenous and endogenous lipid recruitment, and drug transport, into the lymph of males versus females. Finally, lymphatic transport and drug absorption in the mouse were compared to other pre-clinical models (rats/dogs). Methods Animals were orally or intraduodenally administered 1.6 mg/kg halofantrine in low or high 14C-lipid doses. For bioavailability calculation, animals were intravenuosly administered halofantrine. Lymph or blood samples were taken and halofantrine, triglyceride, phospholipid and 14C-lipid concentrations measured. Results Lymphatic lipid transport increased linearly with lipid dose, was similar across species and in male/female animals. In contrast, lymphatic transport of halofantrine differed markedly across species (dogs>rats>mice) and plateaued at higher lipid doses. Lower bioavailability appeared responsible for some species differences in halofantrine lymphatic transport; however other systematic differences were involved. Conclusions A contemporary lymph-cannulated mouse model was established which will enable investigation of lymphatic transport in transgenic and disease models. The current study found halofantrine absorption and lymphatic transport are reduced in small animals. Future analyses will investigate mechanisms involved, and if similar trends occur for other drugs, to establish the most relevant model(s) to predict lymphatic transport in humans. PMID:23430484

Bioactivation of food genotoxicants 5-hydroxymethylfurfural and furfuryl alcohol by sulfotransferases from human, mouse and rat: a comparative study.

PubMed

Sachse, Benjamin; Meinl, Walter; Sommer, Yasmin; Glatt, Hansruedi; Seidel, Albrecht; Monien, Bernhard H

2016-01-01

5-Hydroxymethylfurfural (HMF) and furfuryl alcohol (FFA) are moderately potent rodent carcinogens that are present in thermally processed foodstuffs. The carcinogenic effects were hypothesized to originate from sulfotransferase (SULT)-mediated bioactivation yielding DNA-reactive and mutagenic sulfate esters, a confirmed metabolic pathway of HMF and FFA in mice. It is known that orthologous SULT forms substantially differ in substrate specificity and tissue distribution. This could influence HMF- and FFA-induced carcinogenic effects. Here, we studied HMF and FFA sulfoconjugation by 30 individual SULT forms of humans, mice and rats. The catalytic efficiencies (k cat/K M) of HMF sulfoconjugation of human SULT1A1 (13.7 s(-1) M(-1)), mouse Sult1a1 (15.8 s(-1) M(-1)) and 1d1 (4.8 s(-1) M(-1)) and rat Sult1a1 (5.3 s(-1) M(-1)) were considerably higher than those of all other SULT forms investigated (≤0.73 s(-1 )M(-1)). FFA sulfoconjugation was monitored using adenosine as a nucleophilic scavenger for the reactive 2-sulfoxymethylfuran (t 1/2 = 20 s at 37 °C). The resulting adduct N (6)-((furan-2-yl)methyl)-adenosine (N (6)-MF-A) was quantified by isotope-dilution UPLC-MS/MS. The rates of N (6)-MF-A formation showed that hSULT1A1 and its orthologues in mice and rats were also the most important contributors to FFA sulfoconjugation in each of the species. Taken together, the catalytic capacity of hSULT1A1 is comparable to that of mSult1a1 in mice, the species in which carcinogenic effects of HMF and FFA were detected. This is of primary concern due to the expression of hSULT1A1 in many different tissues.

Identification of adipocyte adhesion molecule (ACAM), a novel CTX gene family, implicated in adipocyte maturation and development of obesity

PubMed Central

2004-01-01

Few cell adhesion molecules have been reported to be expressed in mature adipocytes, and the significance of cell adhesion process in adipocyte biology is also unknown. In the present study, we identified ACAM (adipocyte adhesion molecule), a novel homologue of the CTX (cortical thymocyte marker in Xenopus) gene family. ACAM cDNA was isolated during PCR-based cDNA subtraction, and its mRNA was shown to be up-regulated in WATs (white adipose tissues) of OLETF (Otsuka Long–Evans Tokushima fatty) rats, an animal model for Type II diabetes and obesity. ACAM, 372 amino acids in total, has a signal peptide, V-type (variable) and C2-type (constant) Ig domains, a single transmembrane segment and a cytoplasmic tail. The amino acid sequence in rat is highly homologous to mouse (94%) and human (87%). ACAM mRNA was predominantly expressed in WATs in OLETF rats, and increased with the development of obesity until 30 weeks of age, which is when the peak of body mass is reached. Western blot analysis revealed that ACAM protein, approx. 45 kDa, was associated with plasma membrane fractions of mature adipocytes isolated from mesenteric and subdermal adipose deposits of OLETF rats. Up-regulation of ACAM mRNAs in obesity was also shown in WATs of genetically obese db/db mice, diet-induced obese ICR mice and human obese subjects. In primary cultured mouse and human adipocytes, ACAM mRNA expression was progressively up-regulated during differentiation. Several stably transfected Chinese-hamster ovary K1 cell lines were established, and the quantification of ACAM mRNA and cell aggregation assay revealed that the degree of homophilic aggregation correlated well with ACAM mRNA expression. In summary, ACAM may be the critical adhesion molecule in adipocyte differentiation and development of obesity. PMID:15563274

Point: From animal models to prevention of colon cancer. Systematic review of chemoprevention in min mice and choice of the model system.

PubMed

Corpet, Denis E; Pierre, Fabrice

2003-05-01

The Apc(Min/+) mouse model and the azoxymethane (AOM) rat model are the main animal models used to study the effect of dietary agents on colorectal cancer. We reviewed recently the potency of chemopreventive agents in the AOM rat model (D. E. Corpet and S. Tache, Nutr. Cancer, 43: 1-21, 2002). Here we add the results of a systematic review of the effect of dietary and chemopreventive agents on the tumor yield in Min mice. The review is based on the results of 179 studies from 71 articles and is displayed also on the internet http://corpet.net/min.(2) We compared the efficacy of agents in the Min mouse model and the AOM rat model, and found that they were correlated (r = 0.66; P < 0.001), although some agents that afford strong protection in the AOM rat and the Min mouse small bowel increase the tumor yield in the large bowel of mutant mice. The agents included piroxicam, sulindac, celecoxib, difluoromethylornithine, and polyethylene glycol. The reason for this discrepancy is not known. We also compare the results of rodent studies with those of clinical intervention studies of polyp recurrence. We found that the effect of most of the agents tested was consistent across the animal and clinical models. Our point is thus: rodent models can provide guidance in the selection of prevention approaches to human colon cancer, in particular they suggest that polyethylene glycol, hesperidin, protease inhibitor, sphingomyelin, physical exercise, epidermal growth factor receptor kinase inhibitor, (+)-catechin, resveratrol, fish oil, curcumin, caffeate, and thiosulfonate are likely important preventive agents.

CONFOCAL LASER SCANNING MICROSCOPY OF APOPTOSIS IN WHOLE MOUSE AND RAT OVARIES

EPA Science Inventory

Confocal Laser Scanning Microscopy of Apoptosis in Whole Mouse and Rat Ovaries. Robert M. Zucker Susan C. Jeffay and Sally D. Perreault Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, Research ...

Binge alcohol alters PNPLA3 levels in liver through epigenetic mechanism involving histone H3 acetylation.

PubMed

Restrepo, Ricardo J; Lim, Robert W; Korthuis, Ronald J; Shukla, Shivendra D

2017-05-01

The human PNPLA3 (patatin-like phospholipase domain-containing 3) gene codes for a protein which is highly expressed in adipose tissue and liver, and is implicated in lipid homeostasis. While PNPLA3 protein contains regions homologous to functional lipolytic proteins, the regulation of its tissue expression is reflective of lipogenic genes. A naturally occurring genetic variant of PNPLA3 in humans has been linked to increased susceptibility to alcoholic liver disease. We have examined the modulatory effect of alcohol on PNPLA3 protein and mRNA expression as well as the association of its gene promoter with acetylated histone H3K9 by chromatin immunoprecipitation (ChIP) assay in rat hepatocytes in vitro, and in vivo in mouse and rat models of acute binge, chronic, and chronic followed by acute binge ethanol administration. Protein expression of PNPLA3 was significantly increased by alcohol in all three models used. PNPLA3 mRNA also increased, albeit to a varying degree. ChIP assay using H3AcK9 antibody showed increased association with the promoter of PNPLA3 in hepatocytes and in mouse liver. This was less evident in rat livers in vivo except under chronic treatment. It is concluded for the first time that histone acetylation plays a role in the modulation of PNPLA3 levels in the liver exposed to binge ethanol both in vitro and in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

Binge alcohol alters PNPLA3 levels in liver through epigenetic mechanism involving histone H3 acetylation

PubMed Central

Restrepo, Ricardo J.; Lim, Robert W.; Korthuis, Ronald J.; Shukla, Shivendra D.

2017-01-01

The human PNPLA3 (patatin-like phospholipase domain-containing 3) gene codes for a protein which is highly expressed in adipose tissue and liver, and is implicated in lipid homeostasis. While PNPLA3 protein contains regions homologous to functional lipolytic proteins, the regulation of its tissue expression is reflective of lipogenic genes. A naturally occurring genetic variant of PNPLA3 in humans has been linked to increased susceptibility to alcoholic liver disease. We have examined the modulatory effect of alcohol on PNPLA3 protein and mRNA expression as well as the association of its gene promoter with acetylated histone H3K9 by chromatin immunoprecipitation (ChIP) assay in rat hepatocytes in vitro, and in vivo in mouse and rat models of acute binge, chronic, and chronic followed by acute binge ethanol administration. Protein expression of PNPLA3 was significantly increased by alcohol in all three models used. PNPLA3 mRNA also increased, albeit to a varying degree. ChIP assay using H3AcK9 antibody showed increased association with the promoter of PNPLA3 in hepatocytes and in mouse liver. This was less evident in rat livers in vivo except under chronic treatment. It is concluded for the first time that histone acetylation plays a role in the modulation of PNPLA3 levels in the liver exposed to binge ethanol both in vitro and in vivo. PMID:28433418

Glyphosate rodent carcinogenicity bioassay expert panel review.

PubMed

Williams, Gary M; Berry, Colin; Burns, Michele; de Camargo, Joao Lauro Viana; Greim, Helmut

2016-09-01

Glyphosate has been rigorously and extensively tested for carcinogenicity by administration to mice (five studies) and to rats (nine studies). Most authorities have concluded that the evidence does not indicate a cancer risk to humans. The International Agency for Research on Cancer (IARC), however, evaluated some of the available data and concluded that glyphosate probably is carcinogenic to humans. The expert panel convened by Intertek assessed the findings used by IARC, as well as the full body of evidence and found the following: (1) the renal neoplastic effects in males of one mouse study are not associated with glyphosate exposure, because they lack statistical significance, strength, consistency, specificity, lack a dose-response pattern, plausibility, and coherence; (2) the strength of association of liver hemangiosarcomas in a different mouse study is absent, lacking consistency, and a dose-response effect and having in high dose males only a significant incidence increase which is within the historical control range; (3) pancreatic islet-cell adenomas (non-significant incidence increase), in two studies of male SD rats did not progress to carcinomas and lacked a dose-response pattern (the highest incidence is in the low dose followed by the high dose); (4) in one of two studies, a non-significant positive trend in the incidence of hepatocellular adenomas in male rats did not lead to progression to carcinomas; (5) in one of two studies, the non-significant positive trend in the incidence of thyroid C-cell adenomas in female rats was not present and there was no progression of adenomas to carcinomas at the end of the study. Application of criteria for causality considerations to the above mentioned tumor types and given the overall weight-of-evidence (WoE), the expert panel concluded that glyphosate is not a carcinogen in laboratory animals.

Nonclinical safety of the sodium-glucose cotransporter 2 inhibitor empagliflozin.

PubMed

Bogdanffy, Matthew S; Stachlewitz, Robert F; van Tongeren, Susan; Knight, Brian; Sharp, Dale E; Ku, Warren; Hart, Susan Emeigh; Blanchard, Kerry

2014-01-01

Empagliflozin, a selective inhibitor of the renal tubular sodium-glucose cotransporter 2, was developed for treatment of type 2 diabetes mellitus. Nonclinical safety of empagliflozin was studied in a battery of tests to support global market authorization. Safety pharmacology studies indicated no effect of empagliflozin on measures of respiratory or central nervous system function in rats or cardiovascular safety in telemeterized dogs. In CD-1 mouse, Wistar Han rat, or beagle dogs up to 13, 26, or 52 weeks of treatment, respectively, empagliflozin exhibited a toxicity profile consistent with secondary supratherapeutic pharmacology related to glucose loss and included decreased body weight and body fat, increased food consumption, diarrhea, dehydration, decreased serum glucose and increases in other serum parameters reflective of increased protein catabolism, gluconeogenesis, and electrolyte imbalances, and urinary changes such as polyuria and glucosuria. Microscopic changes were consistently observed in kidney and included tubular nephropathy and interstitial nephritis (dog), renal mineralization (rat) and tubular epithelial cell karyomegaly, single cell necrosis, cystic hyperplasia, and hypertrophy (mouse). Empagliflozin was not genotoxic. Empagliflozin was not carcinogenic in female mice or female rats. Renal adenoma and carcinoma were induced in male mice only at exposures 45 times the maximum clinical dose. These tumors were associated with a spectrum of nonneoplastic changes suggestive of a nongenotoxic, cytotoxic, and cellular proliferation-driven mechanism. In male rats, testicular interstitial cell tumors and hemangiomas of the mesenteric lymph node were observed; both tumors are common in rats and are unlikely to be relevant to humans. These studies demonstrate the nonclinical safety of empagliflozin. © The Author(s) 2014.

Gene Transfer and Molecular Cloning of the Human NGF Receptor

NASA Astrophysics Data System (ADS)

Chao, Moses V.; Bothwell, Mark A.; Ross, Alonzo H.; Koprowski, Hilary; Lanahan, Anthony A.; Buck, C. Randall; Sehgal, Amita

1986-04-01

Nerve growth factor (NGF) and its receptor are important in the development of cells derived from the neural crest. Mouse L cell transformants have been generated that stably express the human NGF receptor gene transfer with total human DNA. Affinity cross-linking, metabolic labeling and immunoprecipitation, and equilibrium binding with 125I-labeled NGF revealed that this NGF receptor had the same size and binding characteristics as the receptor from human melanoma cells and rat PC12 cells. The sequences encoding the NGF receptor were molecularly cloned using the human Alu repetitive sequence as a probe. A cosmid clone that contained the human NGF receptor gene allowed efficient transfection and expression of the receptor.

Monoclonal antibodies to human vitamin D-binding protein.

PubMed Central

Pierce, E A; Dame, M C; Bouillon, R; Van Baelen, H; DeLuca, H F

1985-01-01

Monoclonal antibodies to vitamin D-binding protein isolated from human serum have been produced. The antibodies obtained have been shown to be specific for human vitamin D-binding protein by three independent assays. The antibodies recognize human vitamin D-binding protein specifically in an enzyme-linked immunosorbent assay. Human vitamin D-binding protein is detected specifically in both pure and crude samples by a radiometric immunosorbent assay (RISA) and by an immunoprecipitation assay. The anti-human vitamin D-binding protein antibodies cross-react with monkey and pig vitamin D-binding protein, but not with vitamin D-binding protein from rat, mouse, or chicken, as determined by the RISA and immunoprecipitation assays. Images PMID:3936035

Species-specific vesicular monoamine transporter 2 (VMAT2) expression in mammalian pancreatic beta cells: implications for optimising radioligand-based human beta cell mass (BCM) imaging in animal models

PubMed Central

Hartwig, N. R.; Kalmbach, N.; Klietz, M.; Anlauf, M.; Eiden, L. E.; Weihe, E.

2014-01-01

Aims/hypothesis Imaging of beta cell mass (BCM) is a major challenge in diabetes research. The vesicular monoamine transporter 2 (VMAT2) is abundantly expressed in human beta cells. Radiolabelled analogues of tetrabenazine (TBZ; a low-molecular-weight, cell-permeant VMAT2-selective ligand) have been employed for pancreatic islet imaging in humans. Since reports on TBZ-based VMAT2 imaging in rodent pancreas have been fraught with confusion, we compared VMAT2 gene expression patterns in the mouse, rat, pig and human pancreas, to identify appropriate animal models with which to further validate and optimise TBZ imaging in humans. Methods We used a panel of highly sensitive VMAT2 antibodies developed against equivalently antigenic regions of the transporter from each species in combination with immunostaining for insulin and species-specific in situ hybridisation probes. Individual pancreatic islets were obtained by laser-capture microdissection and subjected to analysis of mRNA expression of VMAT2. Results The VMAT2 protein was not expressed in beta cells in the adult pancreas of common mouse or rat laboratory strains, in contrast to its expression in beta cells (but not other pancreatic endocrine cell types) in the pancreas of pigs and humans. VMAT2- and tyrosine hydroxylase co-positive (catecholaminergic) innervation was less abundant in humans than in rodents. VMAT2-positive mast cells were identified in the pancreas of all species. Conclusions/interpretation Primates and pigs are suitable models for TBZ imaging of beta cells. Rodents, because of a complete lack of VMAT2 expression in the endocrine pancreas, are a ‘null’ model for assessing interference with BCM measurements by VMAT2-positive mast cells and sympathetic innervation in the pancreas. PMID:23404442

Early tumor growth in metastatic organs influenced by the microenvironment is an important factor which provides organ specificity of colon cancer metastasis.

PubMed

Hara, Y; Ogata, Y; Shirouzu, K

2000-12-01

We have previously demonstrated that liver metastases in nude mice and lung metastases in nude rats occurred specifically, when KM12SM human colon carcinoma cells were inoculated orthotopically into the cecal wall of nude mice and rats. To clarify the relationship between the tumor growth potential in the metastatic organs and the metastatic organ preference in these two metastatic models, we have evaluated the in vitro cell growth activities affected by the organ conditioned medium (CM) from the liver and lung, and the in vivo growth activities of the ectopic implanted tumors in the liver and lung. The tumorigenicity of the ectopic implanted tumors was 100% in mouse liver, 33% in rat liver, 50% in mouse lung, and 75% in rat lung. The crude liver CM of the animals showed inhibitory activities for KM12SM cell growth in a dosage-dependent manner, and the crude lung CM stimulated KM12SM cell growth. The liver CM of nude mice inhibited the KM12SM cell growth more strongly compared with the CM of nude rats, and the lung CM of nude rats was more strongly stimulated compared with the CM of nude mice. The liver CM of nude mice had non-heparin binding factors, which stimulated or inhibited KM12SM cell growth, in a molecular weight range of 50 to 100 kDa. By contrast, the liver CM of nude rats showed no growth stimulating activity for KM12SM cells. These results suggest that the metastatic organ specificity of KM12SM cells may depend on the early tumor growth influenced by the microenvironment in metastatic organs.

Insights from Human/Mouse genome comparisons

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pennacchio, Len A.

2003-03-30

Large-scale public genomic sequencing efforts have provided a wealth of vertebrate sequence data poised to provide insights into mammalian biology. These include deep genomic sequence coverage of human, mouse, rat, zebrafish, and two pufferfish (Fugu rubripes and Tetraodon nigroviridis) (Aparicio et al. 2002; Lander et al. 2001; Venter et al. 2001; Waterston et al. 2002). In addition, a high-priority has been placed on determining the genomic sequence of chimpanzee, dog, cow, frog, and chicken (Boguski 2002). While only recently available, whole genome sequence data have provided the unique opportunity to globally compare complete genome contents. Furthermore, the shared evolutionary ancestrymore » of vertebrate species has allowed the development of comparative genomic approaches to identify ancient conserved sequences with functionality. Accordingly, this review focuses on the initial comparison of available mammalian genomes and describes various insights derived from such analysis.« less

IN VIVO ANTI-INFLAMMATORY EFFECTS OF TARAXASTEROL AGAINST ANIMAL MODELS

PubMed Central

Wang, Ying; Li, Guan-Hao; Liu, Xin-Yu; Xu, Lu; Wang, Sha-Sha; Zhang, Xue-Mei

2017-01-01

Background: Traditional Chinese medicine Taraxacum officinale has been widely used to treat various inflammatory diseases. Taraxasterol is one of the main active components isolated from Taraxacum officinale. Recently, we have demonstrated that taraxasterol has the in vitro anti-inflammatory effects. This study aims to determine the in vivo anti-inflammatory effects of taraxasterol against animal models. Materials and Methods: Anti-inflammatory effects were assessed in four animal models by using dimethylbenzene-induced mouse ear edema, carrageenan-induced rat paw edema, acetic acid-induced mouse vascular permeability and cotton pellet-induced rat granuloma tests. Results: Our results demonstrated that taraxasterol dose-dependently attenuated dimethylbenzene-induced mouse ear edema and carrageenan-induced rat paw edema, decreased acetic acid-induced mouse vascular permeability and inhibited cotton pellet-induced rat granuloma formation. Conclusion: Our finding indicates that taraxasterol has obvious in vivo anti-inflammatory effects against animal models. It will provide experimental evidences for the traditional use of Taraxacum officinale and taraxasterol in inflammatory diseases. PMID:28480383

Discovery of LY2457546: a multi-targeted anti-angiogenic kinase inhibitor with a novel spectrum of activity and exquisite potency in the acute myelogenous leukemia-Flt-3-internal tandem duplication mutant human tumor xenograft model.

PubMed

Burkholder, Timothy P; Clayton, Joshua R; Rempala, Mark E; Henry, James R; Knobeloch, John M; Mendel, David; McLean, Johnathan A; Hao, Yan; Barda, David A; Considine, Eileen L; Uhlik, Mark T; Chen, Yuefeng; Ma, Liandong; Bloem, Laura J; Akunda, Jacqueline K; McCann, Denis J; Sanchez-Felix, Manuel; Clawson, David K; Lahn, Michael M; Starling, James J

2012-06-01

LY2457546 is a potent and orally bioavailable inhibitor of multiple receptor tyrosine kinases involved in angiogenic and tumorigenic signalling. In biochemical and cellular assays, LY2457546 demonstrates potent activity against targets that include VEGFR2 (KDR), PDGFRβ, FLT-3, Tie-2 and members of the Eph family of receptors. With activities against both Tie2 and Eph receptors, LY2457546 possesses an activity profile that distinguishes it from multikinase inhibitors. When compared head to head with sunitinib, LY2457546 was more potent for inhibition of endothelial tube formation in an in vitro angiogenesis co-culture model with an intermittent treatment design. In vivo, LY2457546 inhibited VEGF-driven autophosphorylation of lung KDR in the mouse and rat in a dose and concentration dependent manner. LY2457546 was well tolerated and exhibited efficacy in a 13762 syngeneic rat mammary tumor model in both once and twice daily continuous dosing schedules and in mouse human tumor xenograft models of lung, colon, and prostate origin. Additionally, LY2457546 caused complete regression of well-established tumors in an acute myelogenous leukemia (AML) FLT3-ITD mutant xenograft tumor model. The observed efficacy that was displayed by LY2457546 in the AML FLT3-ITD mutant tumor model was superior to sunitinib when both were evaluated using equivalent doses normalized to in vivo inhibition of pKDR in mouse lung. LY2457546 was well tolerated in non-clinical toxicology studies conducted in rats and dogs. The majority of the toxicities observed were similar to those observed with other multi-targeted anti-angiogenic kinase inhibitors (MAKs) and included bone marrow hypocellularity, hair and skin depigmentation, cartilage dysplasia and lymphoid organ degeneration and necrosis. Thus, the unique spectrum of target activity, potent in vivo anti-tumor efficacy in a variety of rodent and human solid tumor models, exquisite potency against a clinically relevant model of AML, and non-clinical safety profile justify the advancement of LY2457546 into clinical testing.

The d4 gene family in the human genome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chestkov, A.V.; Baka, I.D.; Kost, M.V.

1996-08-15

The d4 domain, a novel zinc finger-like structural motif, was first revealed in the rat neuro-d4 protein. Here we demonstrate that the d4 domain is conserved in evolution and that three related genes form a d4 family in the human genome. The human neuro-d4 is very similar to rat neuro-d4 at both the amino acid and the nucleotide levels. Moreover, the same splice variants have been detected among rat and human neuro-d4 transcripts. This gene has been localized on chromosome 19, and two other genes, members of the d4 family isolated by screening of the human genomic library at lowmore » stringency, have been mapped to chromosomes 11 and 14. The gene on chromosome 11 is the homolog of the ubiquitously expressed mouse gene ubi-d4/requiem, which is required for cell death after deprivation of trophic factors. A gene with a conserved d4 domain has been found in the genome of the nematode Caenorhabditis elegans. The conservation of d4 proteins from nematodes to vertebrates suggests that they have a general importance, but a diversity of d4 proteins expressed in vertebrate nervous systems suggests that some family members have special functions. 11 refs., 2 figs.« less

Role of the gastrointestinal ecosystem in the development of Type 1 Diabetes

PubMed Central

Daft, Joseph G.; Lorenz, Robin G.

2015-01-01

A new emphasis has been put on the role of the gastrointestinal (GI) ecosystem in autoimmune diseases; however, there is limited knowledge about its role in type 1 diabetes (T1D). Distinct differences have been observed in intestinal permeability, epithelial barrier function, commensal microbiota, and mucosal innate and adaptive immunity of patients and animals with T1D, when compared to healthy controls. The non-obese diabetic (NOD) mouse and the BioBreeding diabetes prone (BBdp) rat are the most commonly used models to study T1D pathogenesis. With the increasing awareness of the importance of the GI ecosystem in systemic disease, it is critical to understand the basics, as well as the similarities and differences between rat and mouse models and human patients. This review examines the current knowledge of the role of the GI ecosystem in T1D and indicates the extensive opportunities for further investigation that could lead to biomarkers and therapeutic interventions for disease prevention and/or modulation. PMID:25952017

Preclinical pharmacokinetics of the novel PI3K inhibitor GDC-0941 and prediction of its pharmacokinetics and efficacy in human.

PubMed

Salphati, Laurent; Pang, Jodie; Plise, Emile G; Chou, Bilin; Halladay, Jason S; Olivero, Alan G; Rudewicz, Patrick J; Tian, Qingping; Wong, Susan; Zhang, Xiaolin

2011-12-01

The phosphatidylinositol 3-kinase (PI3K) pathway is a major determinant of cell cycling and proliferation. Its deregulation is associated with the development of many cancers. GDC-0941, a potent and selective inhibitor of PI3K, was characterised preclinically in in vitro and in vivo studies. Plasma protein binding was extensive, with free fraction less than 7%, and blood-to-plasma ratio ranged from 0.6 to 1.2 among the species tested. GDC-0941 human hepatic clearance was predicted to be moderate by liver microsomal incubations. GDC-0941 had high permeability in Madin-Darby canine kidney cells. The clearance of GDC-0941 was high in mouse (63.7 mL/min/kg), rat (49.3 mL/min/kg) and cynomolgus monkey (58.6 mL/min/kg), and moderate in dog (11.9 mL/min/kg). The volume of distribution ranged from 2.52 L/kg in rat to 2.94 L/kg in monkey. Oral bioavailability ranged from 18.6% in monkey to 77.9% in mouse. Predicted human clearance and volume of distribution using allometry were 6 mL/min/kg and 2.9 L/kg, respectively. The human efficacious doses were predicted based on results from preclinical pharmacokinetic studies and xenograft models. GDC-0941 preclinical characterisation and predictions of its properties in human supported its progression towards clinical development. GDC-0941 is currently in phase II clinical trials.

Malignant pericytes expressing GT198 give rise to tumor cells through angiogenesis.

PubMed

Zhang, Liyong; Wang, Yan; Rashid, Mohammad H; Liu, Min; Angara, Kartik; Mivechi, Nahid F; Maihle, Nita J; Arbab, Ali S; Ko, Lan

2017-08-01

Angiogenesis promotes tumor development. Understanding the crucial factors regulating tumor angiogenesis may reveal new therapeutic targets. Human GT198 ( PSMC3IP or Hop2) is an oncoprotein encoded by a DNA repair gene that is overexpressed in tumor stromal vasculature to stimulate the expression of angiogenic factors. Here we show that pericytes expressing GT198 give rise to tumor cells through angiogenesis. GT198 + pericytes and perivascular cells are commonly present in the stromal compartment of various human solid tumors and rodent xenograft tumor models. In human oral cancer, GT198 + pericytes proliferate into GT198 + tumor cells, which migrate into lymph nodes. Increased GT198 expression is associated with increased lymph node metastasis and decreased progression-free survival in oral cancer patients. In rat brain U-251 glioblastoma xenografts, GT198 + pericytes of human tumor origin encase endothelial cells of rat origin to form mosaic angiogenic blood vessels, and differentiate into pericyte-derived tumor cells. The net effect is continued production of glioblastoma tumor cells from malignant pericytes via angiogenesis. In addition, activation of GT198 induces the expression of VEGF and promotes tube formation in cultured U251 cells. Furthermore, vaccination using GT198 protein as an antigen in mouse xenograft of GL261 glioma delayed tumor growth and prolonged mouse survival. Together, these findings suggest that GT198-expressing malignant pericytes can give rise to tumor cells through angiogenesis, and serve as a potential source of cells for distant metastasis. Hence, the oncoprotein GT198 has the potential to be a new target in anti-angiogenic therapies in human cancer.

CARCINOGENICITY OF BROMODICHLOROMETHANE ADMINISTERED IN DRINKING WATER TO THE MALE F344/N RAT AND B6C3F, MOUSE

EPA Science Inventory

CARCINOGENICITY OF BROMODICHLOROMETHANE ADMINISTERED IN DRINKING WATER TO THE MALE F344/N RAT AND B6C3F1 MOUSE.

Bromodichloromethane (BDCM) has been shown to produce kidney and large bowel tumors in both male and female F344/N rats, kidney tumors in male B6C3F 1 mice and ...

Cloning of a cDNA encoding rat aldehyde dehydrogenase with high activity for retinal oxidation.

PubMed

Bhat, P V; Labrecque, J; Boutin, J M; Lacroix, A; Yoshida, A

1995-12-12

Retinoic acid (RA), an important regulator of cell differentiation, is biosynthesized from retinol via retinal by a two-step oxidation process. We previously reported the purification and partial amino acid (aa) sequence of a rat kidney aldehyde dehydrogenase (ALDH) isozyme that catalyzed the oxidation of 9-cis and all-trans retinal to corresponding RA with high efficiency [Labrecque et al. Biochem. J. 305 (1995) 681-684]. A rat kidney cDNA library was screened using a 291-bp PCR product generated from total kidney RNA using a pair of oligodeoxyribonucleotide primers matched with the aa sequence. The full-length rat kidney ALDH cDNA contains a 2315-bp (501 aa) open reading frame (ORF). The aa sequence of rat kidney ALDH is 89, 96 and 87% identical to that of the rat cytosolic ALDH, the mouse cytosolic ALDH and human cytosolic ALDH, respectively. Northern blot and RT-PCR-mediated analysis demonstrated that rat kidney ALDH is strongly expressed in kidney, lung, testis, intestine, stomach and trachea, but weakly in the liver.

RNA PROFILES IN RAT AND MOUSE EPIDIDYMAL SPERMATOZOA

EPA Science Inventory

RNA PROFILES IN RAT AND MOUSE EPIDIDYMAL SPERMATOZOA
Kary E. Thompson1, Hongzu Ren1, Judith E. Schmid1 and David J. Dix1
1Reproductive Toxicology Division, NHEERL, ORD, U.S. Environmental Protection Agency, RTP, NC.

Mature spermatozoa are transcriptionally inactive...

Studies of styrene, styrene oxide and 4-hydroxystyrene toxicity in CYP2F2 knockout and CYP2F1 humanized mice support lack of human relevance for mouse lung tumors.

PubMed

Cruzan, G; Bus, J; Hotchkiss, J; Sura, R; Moore, C; Yost, G; Banton, M; Sarang, S

2013-06-01

Styrene (S) is lung tumorigenic in mice but not in rats. S and its alkene-oxidized metabolite styrene oxide (SO) were not lung toxic in CYP2F2(-/-) [knockout] mice, indicating S-induced mouse lung tumors are mediated through mouse-specific CYP2F2-generated ring-oxidized metabolite(s) in lung bronchioles. The human relevance of the CYP2F MOA was assessed by insertion of a human CYP2F1, 2A13, 2B6 transgene into CYP2F2(-/-) mice; CYP2F1 expression and activity were confirmed in the transgenic (TG) mice. No evidence of cytotoxicity or increased cell proliferation (BrdU labeling) was seen in TG mice treated with either S or SO (200mg/kg/day ip for 5days). In contrast to S and SO, 4HS (105mg/kg/day ip for 5days) increased BrdU labeling 5-10-fold in WT mice, <3-fold increase in KO mice and 2-4-fold in TG mice. The limited response of 4HS in KO and TG mice may result from intrinsic toxicity or from further metabolism; regardless of the MOA, these findings indicate that the CYP2F-mediated tumorigenic MOA in WT mice is not operative for S, SO, or for 4HS putatively derived from metabolism of S by CYP2F1 in humans, and thus S-induced mouse lung tumors are unlikely to be relevant to human risk. Copyright © 2013. Published by Elsevier Inc.

A dinucleotide motif in oligonucleotides shows potent immunomodulatory activity and overrides species-specific recognition observed with CpG motif.

PubMed

Kandimalla, Ekambar R; Bhagat, Lakshmi; Zhu, Fu-Gang; Yu, Dong; Cong, Yan-Ping; Wang, Daqing; Tang, Jimmy X; Tang, Jin-Yan; Knetter, Cathrine F; Lien, Egil; Agrawal, Sudhir

2003-11-25

Bacterial and synthetic DNAs containing CpG dinucleotides in specific sequence contexts activate the vertebrate immune system through Toll-like receptor 9 (TLR9). In the present study, we used a synthetic nucleoside with a bicyclic heterobase [1-(2'-deoxy-beta-d-ribofuranosyl)-2-oxo-7-deaza-8-methyl-purine; R] to replace the C in CpG, resulting in an RpG dinucleotide. The RpG dinucleotide was incorporated in mouse- and human-specific motifs in oligodeoxynucleotides (oligos) and 3'-3-linked oligos, referred to as immunomers. Oligos containing the RpG motif induced cytokine secretion in mouse spleen-cell cultures. Immunomers containing RpG dinucleotides showed activity in transfected-HEK293 cells stably expressing mouse TLR9, suggesting direct involvement of TLR9 in the recognition of RpG motif. In J774 macrophages, RpG motifs activated NF-kappa B and mitogen-activated protein kinase pathways. Immunomers containing the RpG dinucleotide induced high levels of IL-12 and IFN-gamma, but lower IL-6 in time- and concentration-dependent fashion in mouse spleen-cell cultures costimulated with IL-2. Importantly, immunomers containing GTRGTT and GARGTT motifs were recognized to a similar extent by both mouse and human immune systems. Additionally, both mouse- and human-specific RpG immunomers potently stimulated proliferation of peripheral blood mononuclear cells obtained from diverse vertebrate species, including monkey, pig, horse, sheep, goat, rat, and chicken. An immunomer containing GTRGTT motif prevented conalbumin-induced and ragweed allergen-induced allergic inflammation in mice. We show that a synthetic bicyclic nucleotide is recognized in the C position of a CpG dinucleotide by immune cells from diverse vertebrate species without bias for flanking sequences, suggesting a divergent nucleotide motif recognition pattern of TLR9.

Resveratrol Metabolism in a Non-Human Primate, the Grey Mouse Lemur (Microcebus murinus), Using Ultra-High-Performance Liquid Chromatography–Quadrupole Time of Flight

PubMed Central

Menet, Marie-Claude; Marchal, Julia; Dal-Pan, Alexandre; Taghi, Méryam; Nivet-Antoine, Valérie; Dargère, Delphine; Laprévote, Olivier; Beaudeux, Jean-Louis; Aujard, Fabienne; Epelbaum, Jacques; Cottart, Charles-Henry

2014-01-01

The grey mouse lemur (Microcebus murinus) is a non-human primate used to study the ageing process. Resveratrol is a polyphenol that may increase lifespan by delaying age-associated pathologies. However, no information about resveratrol absorption and metabolism is available for this primate. Resveratrol and its metabolites were qualitatively and quantitatively analyzed in male mouse-lemur plasma (after 200 mg.kg−1 of oral resveratrol) by ultra-high performance liquid chromatography (UHPLC), coupled to a quadrupole-time-of-flight (Q-TOF) mass spectrometer used in full-scan mode. Data analyses showed, in MSE mode, an ion common to resveratrol and all its metabolites: m/z 227.072, and an ion common to dihydro-resveratrol metabolites: m/z 229.08. A semi-targeted study enabled us to identify six hydrophilic resveratrol metabolites (one diglucurono-conjugated, two monoglucurono-conjugated, one monosulfo-conjugated and two both sulfo- and glucurono-conjugated derivatives) and three hydrophilic metabolites of dihydro-resveratrol (one monoglucurono-conjugated, one monosulfo-conjugated, and one both sulfo- and glucurono-conjugated derivatives). The presence of such metabolites has been already detected in the mouse, rat, pig, and humans. Free resveratrol was measurable for several hours in mouse-lemur plasma, and its two main metabolites were trans-resveratrol-3-O-glucuronide and trans-resveratrol-3-sulfate. Free dihydro-resveratrol was not measurable whatever the time of plasma collection, while its hydrophilic metabolites were present at 24 h after intake. These data will help us interpret the effect of resveratrol in mouse lemurs and provide further information on the inter-species characteristics of resveratrol metabolism. PMID:24663435

Correction of mouse ornithine transcarbamylase deficiency by gene transfer into the germ line.

PubMed Central

Cavard, C; Grimber, G; Dubois, N; Chasse, J F; Bennoun, M; Minet-Thuriaux, M; Kamoun, P; Briand, P

1988-01-01

The sparse fur with abnormal skin and hair (Spf-ash) mouse is a model for the human X-linked hereditary disorder, ornithine transcarbamylase (OTC) deficiency. In Spf-ash mice, both OTC mRNA and enzyme activity are 5% of control values resulting in hyperammonemia, pronounced orotic aciduria and an abnormal phenotype characterized by growth retardation and sparse fur. Using microinjection, we introduced a construction containing rat OTC cDNA linked to the SV40 early promoter into fertilized eggs of Spf-ash mice. The expression of the transgene resulted in the development of a transgenic mouse whose phenotype and orotic acid excretion are fully normalized. Thus, the possibility of correcting hereditary enzymatic defect by gene transfer of heterologous cDNA coding for the normal enzyme has been demonstrated. Images PMID:3162766

Role of muscarinic receptor subtypes in central antinociception.

PubMed Central

Bartolini, A.; Ghelardini, C.; Fantetti, L.; Malcangio, M.; Malmberg-Aiello, P.; Giotti, A.

1992-01-01

1. The ability to modify the pain threshold by the two M1-muscarinic agonists: McN-A-343 and AF-102B and by the specific M2-agonist arecaidine was examined in mice and rats by using three different noxious stimuli: chemical (writhing test), thermic (hot-plate test) and mechanical (paw pressure test). 2. In the mouse hot-plate test McN-A-343 (20-50 micrograms per mouse i.c.v.) and AF-102B (1-10 mg kg-1 i.p.) produced significant antinociception which was prevented by atropine (1 microgram per mouse i.c.v.) and by the two selective M1 antagonists: pirenzepine (0.01 micrograms per mouse i.c.v.) and dicyclomine (0.08 micrograms per mouse i.c.v. or 10 mg kg-1 i.p.) but not by the specific M2-antagonist AFDX-116 (0.1 micrograms per mouse i.c.v.), naloxone (1 mg kg-1 i.p.) or by the acetylcholine (ACh) depletor hemicholinium-3 (HC-3) (1 micrograms per mouse i.c.v.). McN-A-343 and AF-102B were able to increase the pain threshold also in the mouse acetic acid writhing test and in rat paw pressure test. These antinociceptive effects were completely prevented by dicyclomine (0.08 micrograms per mouse i.c.v. or 10 mg kg-1 i.p.) but not by AFDX-116 (0.1 microgram per mouse or rat i.c.v.). 3. In contrast with the M1-agonists, the M2-agonist arecaidine (0.1-2 micrograms per mouse or rat i.c.v.) did not induce antinociception in all three analgesic tests. However, arecaidine, at the same i.c.v. doses, was able to reduce the pain threshold in the hot-plate and paw pressure tests.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1375858

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buchholz, B. A.

The project has two main goals: 1) Identify the types of adducts naphthalene (NA) forms with DNA and 2) determine whether adduct formation correlates with site selective tumor formation in defined subcompartments of the respiratory tract (respiratory and olfactory nasal epithelium and airways of mice, rats and rhesus monkeys). Five tasks are associated with the completion of the goals. Task 1: Contracting and Animal Use Approvals. IACUC and ACURO approvals are complete. The subcontract with UC Davis (UCD) was executed in December 2014. Task 2: Perform In Vitro Study for Goal 1. Rat and mouse samples exposures completed. Monkey samplesmore » need to be exposed in next quarter. Task 3: Perform In Vitro Study for Goal 2. Mouse and rat ex vivo exposures completed. Monkey samples need to be completed in the next quarter. Task 4: Sample Preparation and Analysis. Mouse and Rat Goal 2 samples completed. Monkey samples remain to be done for Goal 2. Rat samples completed for Goal 1. Mouse and Monkey samples for Goal 1 need to be completed. Task 5: Data Interpretation and Reporting. Poster will be presented at 2016 Society of Toxicology Meeting. Outline for paper on adduct formation complete and similar to poster for SOT meeting.« less

Kras, Egfr, and Tp53 Mutations in B6C3F1/N Mouse and F344/NTac Rat Alveolar/Bronchiolar Carcinomas Resulting from Chronic Inhalation Exposure to Cobalt Metal

PubMed Central

Hong, Hue-Hua L.; Hoenerhoff, Mark J.; Ton, Thai-Vu; Herbert, Ronald A.; Kissling, Grace E.; Hooth, Michelle J.; Behl, Mamta; Witt, Kristine L.; Smith-Roe, Stephanie L.; Sills, Robert C.; Pandiri, Arun R.

2015-01-01

Rodent lung tumors are morphologically similar to a subtype of human lung adenocarcinomas. The objective of this study was to evaluate Kras, Egfr and Tp53 mutations, which are relevant to human lung cancer, in cobalt metal dust (CMD) induced alveolar/bronchiolar tumors of B6C3F1/N mice and F344/NTac rats. Kras mutations were detected in 67% (mice) and 31% (rats) of CMD-induced lung tumors, and were predominantly exon 1 codon 12 G to T transversions (80% in mice and 57% in rats). Egfr mutations were detected in 17% (both mice and rats) of CMD-induced lung tumors, and were predominantly in exon 20 with 50% G to A transitions (mice and rats). Tp53 mutations were detected in 19% (mice) and 23% (rats) of CMD-induced lung tumors and were predominantly in exon 5 (mice, 69% transversions) and exon 6 (rats, all transitions). No mutations were observed for these genes in spontaneous lung tumors or normal lungs from untreated controls. Ames assays indicated that CMD is mutagenic in the absence but not in the presence of S9 mix. Thus, the mutation data (G to T transversions) and Ames assay results suggest that oxidative damage to DNA may be a contributing factor in CMD-induced pulmonary carcinogenesis in rodents. PMID:26059825

Rodent models of congenital and hereditary cataract in man.

PubMed

Tripathi, B J; Tripathi, R C; Borisuth, N S; Dhaliwal, R; Dhaliwal, D

1991-01-01

Because the organogenesis and physiology of the lens are essentially similar in various mammals, an understanding of the etiology and pathogenesis of the formation of cataract in an animal model will enhance our knowledge of cataractogenesis in man. In this review, we summarize the background, etiology, and pathogenesis of cataracts that occur in rodents. The main advantages of using rodent mutants include the well-researched genetics of the animals and the comparative ease of breeding of large litters. Numerous rodent models of congenital and hereditary cataracts have been studied extensively. In mice, the models include the Cts strain, Fraser mouse, lens opacity gene (Lop) strain, Lop-2 and Lop-3 strains, Philly mouse, Nakano mouse, Nop strain, Deer mouse, Emory mouse, Swiss Webster strain, Balb/c-nct/nct mouse, and SAM-R/3 strain. The rat models include BUdR, ICR, Sprague-Dawley, and Wistar rats, the spontaneously hypertensive rat (SHR), the John Rapp inbred strain of Dahl salt-sensitive rat, as well as WBN/Kob, Royal College of Surgeons (RCS), and Brown-Norway rats. Other proposed models for the study of hereditary cataract include the degu and the guinea pig. Because of the ease of making clinical observations in vivo and the subsequent availability of the intact lens for laboratory analyses at different stages of cataract formation, these animals provide excellent models for clinicopathologic correlations, for monitoring of the natural history of the aging process and of metabolic defects, as well as for investigations on the effect of cataract-modulating agents and drugs, including the prospect of gene therapy.

A Physiologically Based Kinetic Model of Rat and Mouse Gestation: Disposition of a Weak Acid

EPA Science Inventory

A physiologically based toxicokinetic model of gestation in the rat mouse has been developed. The model is superimposed on the normal growth curve for nonpregnant females. It describes the entire gestation period including organogenesis. The model consists of uterus, mammary tiss...

Exposure to perfluorooctane sulfonate during pregnancy in rat and mouse. I: maternal and prenatal evaluations

EPA Science Inventory

Abstract: The maternal and developmental toxicities of perfluorooctane sulfonate (PFOS, C8F17SO3-) were evaluated in the rat and mouse. PFOS is an environmentally persistent compound used as a surfactant and occurs as a degradation product of both perfluorooctane sulfonyl fluorid...

Characterization of motor units in behaving adult mice shows a wide primary range

PubMed Central

Ritter, Laura K.; Tresch, Matthew C.; Heckman, C. J.; Manuel, Marin

2014-01-01

The mouse is essential for genetic studies of motor function in both normal and pathological states. Thus it is important to consider whether the structure of motor output from the mouse is in fact analogous to that recorded in other animals. There is a striking difference in the basic electrical properties of mouse motoneurons compared with those in rats, cats, and humans. The firing evoked by injected currents produces a unique frequency-current (F-I) function that emphasizes recruitment of motor units at their maximum force. These F-I functions, however, were measured in anesthetized preparations that lacked two key components of normal synaptic input: high levels of synaptic noise and neuromodulatory inputs. Recent studies suggest that the alterations in the F-I function due to these two components are essential for recreating firing behavior of motor units in human subjects. In this study we provide the first data on firing patterns of motor units in the awake mouse, focusing on steady output in quiet stance. The resulting firing patterns did not match the predictions from the mouse F-I behaviors but instead revealed rate modulation across a remarkably wide range (10–60 Hz). The low end of the firing range may be due to changes in the F-I relation induced by synaptic noise and neuromodulatory inputs. The high end of the range may indicate that, unlike other species, quiet standing in the mouse involves recruitment of relatively fast-twitch motor units. PMID:24805075

Characterization of motor units in behaving adult mice shows a wide primary range.

PubMed

Ritter, Laura K; Tresch, Matthew C; Heckman, C J; Manuel, Marin; Tysseling, Vicki M

2014-08-01

The mouse is essential for genetic studies of motor function in both normal and pathological states. Thus it is important to consider whether the structure of motor output from the mouse is in fact analogous to that recorded in other animals. There is a striking difference in the basic electrical properties of mouse motoneurons compared with those in rats, cats, and humans. The firing evoked by injected currents produces a unique frequency-current (F-I) function that emphasizes recruitment of motor units at their maximum force. These F-I functions, however, were measured in anesthetized preparations that lacked two key components of normal synaptic input: high levels of synaptic noise and neuromodulatory inputs. Recent studies suggest that the alterations in the F-I function due to these two components are essential for recreating firing behavior of motor units in human subjects. In this study we provide the first data on firing patterns of motor units in the awake mouse, focusing on steady output in quiet stance. The resulting firing patterns did not match the predictions from the mouse F-I behaviors but instead revealed rate modulation across a remarkably wide range (10-60 Hz). The low end of the firing range may be due to changes in the F-I relation induced by synaptic noise and neuromodulatory inputs. The high end of the range may indicate that, unlike other species, quiet standing in the mouse involves recruitment of relatively fast-twitch motor units. Copyright © 2014 the American Physiological Society.

Species differences in the formation of vabicaserin carbamoyl glucuronide.

PubMed

Tong, Zeen; Chandrasekaran, Appavu; DeMaio, William; Jordan, Ronald; Li, Hongshan; Moore, Robin; Poola, Nagaraju; Burghart, Peter; Hultin, Theresa; Scatina, JoAnn

2010-04-01

Vabicaserin is a potent 5-hydroxtryptamine 2C full agonist with therapeutic potential for a wide array of psychiatric disorders. Metabolite profiles indicated that vabicaserin was extensively metabolized via carbamoyl glucuronidation after oral administration in humans. In the present study, the differences in the extent of vabicaserin carbamoyl glucuronide (CG) formation in humans and in animals used for safety assessment were investigated. After oral dosing, the systemic exposure ratios of CG to vabicaserin were approximately 12 and up to 29 in monkeys and humans, respectively, and the ratios of CG to vabicaserin were approximately 1.5 and 1.7 in mice and dogs, respectively. These differences in systemic levels of CG are likely related to species differences in the rate and extent of CG formation and elimination. Whereas CG was the predominant circulating metabolite in humans and a major metabolite in mice, dogs, and monkeys, it was a relatively minor metabolite in rats, in which oxidative metabolism was the major metabolic pathway. Although the CG was not detected in plasma or urine of rats, approximately 5% of the dose was excreted in bile as CG in the 24-h collection postdose, indicating the rat had the metabolic capability of producing the CG. In vitro, in a CO(2)-enriched environment, the CG was the predominant metabolite in dog and human liver microsomes, a major metabolite in monkey and mice, and only a very minor metabolite in rats. Carbamoyl glucuronidation and hydroxylation had similar contributions to vabicaserin metabolism in mouse and monkey liver microsomes. However, only trace amounts of CG were formed in rat liver microsomes, and other metabolites were more prominent than the CG. In conclusion, significant differences in the extent of formation of the CG were observed among the various species examined. The exposure ratios of CG to vabicaserin were highest in humans, followed by monkeys, then mice and dogs, and lowest in rats, and the in vitro metabolite profiles generally correlated well with the in vivo metabolites.

Identification of human cytochrome P450 enzymes involved in the metabolism of IN-1130, a novel activin receptor-like kinase-5 (ALK5) inhibitor.

PubMed

Kim, Y W; Kim, Y K; Kim, D-K; Sheen, Y Y

2008-05-01

1. The in vitro metabolism of 3-((5-(6-methylpyridin-2-yl)-4-(quinoxalin-6-yl)-1H-imidazol-2-yl)methyl)benzamide (IN-1,130), a selective activin receptor-like kinase-5 (ALK5) inhibitor and a candidate drug for fibrotic disease, was studied. 2. The cytochrome P450s (CYPs) responsible for metabolism of IN-1,130 in liver microsomes of rat, mouse, dog, monkey and human, and in human CYP supersomestrade mark, were identified using specific CYP inhibitors. The order of disappearance of IN-1,130 in various liver microsomal systems studied was as follows: monkey, mouse, rat, human, and dog. 3. Five distinct metabolites (M1-M5) were identified in all the above microsomes and their production was substantially inhibited by CYP inhibitors such as SKF-525A and ketoconazole. Among nine human CYP supersomestrade mark examined, CYP3A4, CYP2C8, CYP2D6 1, and CYP2C19 were involved in the metabolism of IN-1,130, and the production of metabolites were significantly inhibited by specific CYP inhibitors. IN-1,130 disappeared fastest in CYP2C8 supersomes. CYP3A4 produced four metabolites of IN-1,130 (M1-M4), whereas supersomes expressing human FMO cDNAs, such as FMO1, FMO3, and FMO5, produced no metabolites. 4. Hence, it is concluded that metabolism of IN-1,130 is mediated by CYP3A4, CYP2C8, CYP2D6 1, and CYP2C19.

The neonatal Fc receptor (FcRn) binds independently to both sites of the IgG homodimer with identical affinity.

PubMed

Abdiche, Yasmina Noubia; Yeung, Yik Andy; Chaparro-Riggers, Javier; Barman, Ishita; Strop, Pavel; Chin, Sherman Michael; Pham, Amber; Bolton, Gary; McDonough, Dan; Lindquist, Kevin; Pons, Jaume; Rajpal, Arvind

2015-01-01

The neonatal Fc receptor (FcRn) is expressed by cells of epithelial, endothelial and myeloid lineages and performs multiple roles in adaptive immunity. Characterizing the FcRn/IgG interaction is fundamental to designing therapeutic antibodies because IgGs with moderately increased binding affinities for FcRn exhibit superior serum half-lives and efficacy. It has been hypothesized that 2 FcRn molecules bind an IgG homodimer with disparate affinities, yet their affinity constants are inconsistent across the literature. Using surface plasmon resonance biosensor assays that eliminated confounding experimental artifacts, we present data supporting an alternate hypothesis: 2 FcRn molecules saturate an IgG homodimer with identical affinities at independent sites, consistent with the symmetrical arrangement of the FcRn/Fc complex observed in the crystal structure published by Burmeister et al. in 1994. We find that human FcRn binds human IgG1 with an equilibrium dissociation constant (KD) of 760 ± 60 nM (N = 14) at 25°C and pH 5.8, and shows less than 25% variation across the other human subtypes. Human IgG1 binds cynomolgus monkey FcRn with a 2-fold higher affinity than human FcRn, and binds both mouse and rat FcRn with a 10-fold higher affinity than human FcRn. FcRn/IgG interactions from multiple species show less than a 2-fold weaker affinity at 37°C than at 25°C and appear independent of an IgG's variable region. Our in vivo data in mouse and rat models demonstrate that both affinity and avidity influence an IgG's serum half-life, which should be considered when choosing animals, especially transgenic systems, as surrogates.

Characterization of rodent liver and kidney AVP receptors: pharmacologic evidence for species differences.

PubMed

Tahara, A; Tsukada, J; Ishii, N; Tomura, Y; Wada, K; Kusayama, T; Yatsu, T; Uchida, W; Tanaka, A

1999-10-22

Radioligand binding studies with [3H]vasopressin (AVP) were used to determine the affinities of AVP receptor agonists and antagonists for mouse liver and kidney plasma membrane preparations. Both membrane preparations exhibited one class of high-affinity binding site. AVP ligand binding inhibition studies confirmed that mouse liver binding sites belong to the V1A subtype while kidney binding sites belong to the V2 receptor subtype. The affinity of each ligand for mouse V1A receptors was very similar to that for rat V1A receptors, showing differences in Ki values of less than 3-fold. In contrast, several peptide (d(CH2)5Tyr(Me)AVP) and nonpeptide (OPC-21268 and SR 49059) ligands had different affinities for mouse and rat kidney V2 receptors, with differences in Ki values ranging from 14- to 17-fold. These results indicate that mouse and rat kidney V2 receptors show significant pharmacologic differences.

Identification and characterization of a Fc receptor activity on the Toxoplasma gondii tachyzoite.

PubMed

Vercammen, M; el Bouhdidi, A; Ben Messaoud, A; de Meuter, F; Bazin, H; Dubremetz, J F; Carlier, Y

1998-01-01

The Immunoglobulin (Ig) binding capacity of Toxoplasma gondii tachyzoites was investigated using fluorescence flow-cytometry analysis. Polyclonal mouse, human and rat immunoglobulins without specific anti-Toxoplasma activity bound to parasites in a concentration-dependent manner, saturating them at circulating serum concentrations. The immunoglobulin class and subclass specificity of binding was investigated using irrelevant monoclonal antibodies. IgM, IgA and IgG reacted with the parasite membrane. The attachment of mouse IgM to the parasite surface was hampered by mouse IgG1, IgG2a, IgG2b and IgG3. The binding of mouse IgG was proportionally reduced with increasing concentrations of mouse monoclonal IgM. The binding of murine immunoglobulin was diminished when in presence of human IgG. Purified Fc- but not Fab portions of immunoglobulins, fixed to parasites. Using labelled calibrated beads, the Ig binding capacity of parasites was estimated to be 6900 +/- 500 sites per tachyzoite. The Kd of the T. gondii Fc Receptor (FcR) activity was determined at 1.4 +/- 0.1 microM (mean +/- SEM). Such FcR activity was reduced by phospholipase C, trypsin and pronase treatment of the parasites. These data show a low affinity FcR activity on T. gondii tachyzoites which recognizes Ig of different species and isotypes and is likely supported by a glycosyl-phosphatidylinositol (GPI)-anchored surface protein of the parasite.

GDF10 Is a Signal for Axonal Sprouting and Functional Recovery after Stroke

PubMed Central

Li, S; Nie, EH; Yin, Y; Benowitz, LI; Tung, S; Vinters, HV; Bahjat, FR; Stenzel-Poore, MP; Kawaguchi, R; Coppola, G; Carmichael, ST

2016-01-01

Stroke produces a limited process of neural repair. Axonal sprouting in cortex adjacent to the infarct is part of this recovery process, but the signal that initiates axonal sprouting is not known. Growth and Differentiation Factor 10 (GDF10) is induced in peri-infarct neurons in mouse, non-human primate and human. GDF10 promotes axonal outgrowth in vitro in mouse, rat and human neurons through TGFβRI/II signaling. Using pharmacogenetic gain and loss of function studies, GDF10 produces axonal sprouting and enhanced functional recovery after stroke; knocking down GDF10 blocks axonal sprouting and reduces recovery. RNA-seq from peri-infarct cortical neurons indicates that GDF10 downregulates PTEN and upregulates PI3 kinase signaling and induces specific axonal guidance molecules. Unsupervised genome-wide association analysis of the GDF10 transcriptome shows that it is not related to neurodevelopment but may partially overlap with other CNS injury patterns. GDF10 is a stroke-induced signal for axonal sprouting and functional recovery. PMID:26502261

Differences in Butadiene Adduct Formation between Rats and Mice Not Due to Selective Inhibition of CYP2E1 by Butadiene Metabolites

PubMed Central

Pianalto, Kaila M.; Hartman, Jessica H.; Boysen, Gunnar; Miller, Grover P.

2013-01-01

CYP2E1 metabolizes 1,3-butadiene (BD) into genotoxic and possibly carcinogenic 1,2-epoxy-3-butene (EB), 1,2:3,4-diepoxybutane (DEB), and 1,2-epoxy-3,4-butanediol (EB-diol). The dose response of DNA and protein adducts derived from BD metabolites increase linearly at low BD exposures and then saturate at higher exposures in rats, but not mice. It was hypothesized that differences in adduct formation between rodents reflect more efficient BD oxidation in mice than rats. Herein, we assessed whether BD-derived metabolites selectively inhibit rat but not mouse CYP2E1 activity using B6C3F1 mouse and Fisher 344 rat liver microsomes. Basal CYP2E1 activities toward 4-nitrophenol were similar between rodents. Through IC50 studies, EB was the strongest inhibitor (IC50 54 μM, mouse; 98 μM, rat), BD-diol considerably weaker (IC50 1200 μM, mouse; 1000 μM, rat), and DEB inhibition nonexistent (IC50 >25 mM). Kinetic studies showed that in both species EB and BD-diol inhibited 4-nitrophenol oxidation through two-site mechanisms in which inhibition constants reflected trends observed in IC50 studies. None of the reactive epoxide metabolites inactivated CYP2E1 irreversibly. Thus, there was no selective inhibition or inactivation of rat CYP2E1 by BD metabolites relative to mouse Cyp2e1, and it can be inferred that CYP2E1 activity toward BD between rodent species would similarly not be impacted by the presence of BD metabolites. Inhibition of CYP2E1 by BD metabolites is then not responsible for the reported species difference in BD metabolism, formation of BD-derived DNA and protein adducts, mutagenicity and tumorigenesis. PMID:24021170

Differences in butadiene adduct formation between rats and mice not due to selective inhibition of CYP2E1 by butadiene metabolites.

PubMed

Pianalto, Kaila M; Hartman, Jessica H; Boysen, Gunnar; Miller, Grover P

2013-11-25

CYP2E1 metabolizes 1,3-butadiene (BD) into genotoxic and possibly carcinogenic 1,2-epoxy-3-butene (EB), 1,2:3,4-diepoxybutane (DEB), and 1,2-epoxy-3,4-butanediol (EB-diol). The dose response of DNA and protein adducts derived from BD metabolites increases linearly at low BD exposures and then saturates at higher exposures in rats, but not mice. It was hypothesized that differences in adduct formation between rodents reflect more efficient BD oxidation in mice than rats. Herein, we assessed whether BD-derived metabolites selectively inhibit rat but not mouse CYP2E1 activity using B6C3F1 mouse and Fisher 344 rat liver microsomes. Basal CYP2E1 activities toward 4-nitrophenol were similar between rodents. Through IC50 studies, EB was the strongest inhibitor (IC50 54μM, mouse; 98μM, rat), BD-diol considerably weaker (IC50 1200μM, mouse; 1000μM, rat), and DEB inhibition nonexistent (IC50>25mM). Kinetic studies showed that in both species EB and BD-diol inhibited 4-nitrophenol oxidation through two-site mechanisms in which inhibition constants reflected trends observed in IC50 studies. None of the reactive epoxide metabolites inactivated CYP2E1 irreversibly. Thus, there was no selective inhibition or inactivation of rat CYP2E1 by BD metabolites relative to mouse Cyp2e1, and it can be inferred that CYP2E1 activity toward BD between rodent species would similarly not be impacted by the presence of BD metabolites. Inhibition of CYP2E1 by BD metabolites is then not responsible for the reported species difference in BD metabolism, formation of BD-derived DNA and protein adducts, mutagenicity and tumorigenesis. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Myelin-oligodendrocyte glycoprotein is a member of a subset of the immunoglobulin superfamily encoded within the major histocompatibility complex

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pham-Dinh, D.; Dautigny, A.; Mattei, M.G.

1993-09-01

Myelin/oligodendrocyte glycoprotein (MOG) is found on the surface of myelinating oligodendrocytes and external lamellae of myelin sheaths in the central nervous system, and it is target antigen in experimental autoimmune encephalomyelitis and multiple sclerosis. The authors have isolated bovine, mouse, and rat MOG cDNA clones and shown that the developmental pattern of MOG expression in the rat central nervous system coincides with the late stages of myelination. The amino-terminal, extracellular domain of MOG has characteristics of an immunoglobulin variable domain and is 46% and 41% identical with the amino terminus of bovine butyrophilin (expressed in the lactating mammary gland) andmore » B-G antigens of the chicken major histocompatibility complex (MHC), respectively; these proteins thus form a subset of the immunoglobulin superfamily. The homology between MOG and B-G extends beyond their structure and genetic mapping to their ability to induce strong antibody responses and has implications for the role of MOG in pathological, autoimmune conditions. The authors colocalized the MOG and BT genes to the human MHC on chromosome 6p21.3-p22. The mouse MOG gene was mapped to the homologous band C of chromosome 17, within the M region of the mouse MHC. 38 refs., 6 figs.« less

Liver repopulation by c-Met-positive stem/progenitor cells isolated from the developing rat liver.

PubMed

Suzuki, Atsushi; Zheng, Yun-wen; Fukao, Katashi; Nakauchi, Hiromitsu; Taniguchi, Hideki

2004-01-01

Self-renewing stem cells responsible for tissue or organ development and regeneration have been recently described. To isolate such cells using flow cytometry, it should be required to find molecules expressing on their cell surfaces. We have previously reported that, on cells fulfilling the criteria for hepatic stem cells, the hepatocyte growth factor receptor c-Met is expressed specifically in the developing mouse liver. In this study, to determine whether c-Met is an essential marker for hepatic stem cells in other animal strains, we examined the potential for in vivo liver-repopulation in sorted fetal rat-derived c-Met+ cells using the retrorsine model. Using flow cytometry and monoclonal antibodies for c-Met and leukocyte common antigen CD45, fetal rat liver cells were fractionated according to the expression of these molecules. Then, cells in each cell subpopulation were sorted and transplanted into the retrorsine-treated adult rats with two-third hepatectomy. At 9 months post transplant, frequency of liver-repopulation was examined by qualitative and quantitative analyses. When we transplanted c-Met+ CD45- sorted cells, many donor-derived cells formed colonies that included mature hepatocytes expressing albumin and containing abundant glycogen in their cytoplasm. In contrast, c-Met- cells and CD45+ cells could not repopulate damaged recipient livers. High enrichment of liver-repopulating cells was conducted by sorting of c-Met+ cells from the developing rat liver. This result suggests that c-Met/HGF interaction plays a crucial role for stem cell growth, differentiation, and self-renewal in rat liver organogenesis. Since the c-Met is also expressed in the fetal mouse-derived hepatic stem cells, this molecule could be expected to be an essential marker for such cell population in the various animal strains, including human.

Litigation Technical Support and Services, Rocky Mountain Arsenal. Biota Remedial Investigation, Version 3.2. Volume 1

DTIC Science & Technology

1989-05-01

significantly. while isodrin is an analog of endrtr and is converted metabolically to that compound. Analyses were also performed for I *l.-dichloro-2,2-bis...abundant are the deer mouse, prairie and meadow voles, and Ord’s kangaroo rat. Larger, more conspicuous rodents include the black-tailed prairie dog...meadow vole, Ord’s kangaroo rat, hispid pocket mouse, and silky pocket mouse(MKE, 1988). Tall weedy forb$, yucca, sand sagebrush, and cattails are

Computational modeling of nanoscale and microscale particle deposition, retention and dosimetry in the mouse respiratory tract.

PubMed

Asgharian, B; Price, O T; Oldham, M; Chen, Lung-Chi; Saunders, E L; Gordon, T; Mikheev, V B; Minard, K R; Teeguarden, J G

2014-12-01

Comparing effects of inhaled particles across rodent test systems and between rodent test systems and humans is a key obstacle to the interpretation of common toxicological test systems for human risk assessment. These comparisons, correlation with effects and prediction of effects, are best conducted using measures of tissue dose in the respiratory tract. Differences in lung geometry, physiology and the characteristics of ventilation can give rise to differences in the regional deposition of particles in the lung in these species. Differences in regional lung tissue doses cannot currently be measured experimentally. Regional lung tissue dosimetry can however be predicted using models developed for rats, monkeys, and humans. A computational model of particle respiratory tract deposition and clearance was developed for BALB/c and B6C3F1 mice, creating a cross-species suite of available models for particle dosimetry in the lung. Airflow and particle transport equations were solved throughout the respiratory tract of these mice strains to obtain temporal and spatial concentration of inhaled particles from which deposition fractions were determined. Particle inhalability (Inhalable fraction, IF) and upper respiratory tract (URT) deposition were directly related to particle diffusive and inertial properties. Measurements of the retained mass at several post-exposure times following exposure to iron oxide nanoparticles, micro- and nanoscale C60 fullerene, and nanoscale silver particles were used to calibrate and verify model predictions of total lung dose. Interstrain (mice) and interspecies (mouse, rat and human) differences in particle inhalability, fractional deposition and tissue dosimetry are described for ultrafine, fine and coarse particles.

Metabolism and disposition of 2-ethylhexyl-p-methoxycinnamate following oral gavage and dermal exposure in Harlan Sprague Dawley rats and B6C3F1/N mice and in hepatocytes in vitro.

PubMed

Fennell, Timothy R; Mathews, James M; Snyder, Rodney W; Hong, Yan; Watson, Scott L; Black, Sherry R; McIntyre, Barry S; Waidyanatha, Suramya

2017-11-23

1. 2-Ethylhexyl-p-methoxycinnamate (EHMC) is commonly used as an ingredient in sunscreens, resulting in potential oral and dermal exposure in humans. 2. Clearance and metabolism of EHMC in hepatocytes and disposition and metabolism of EHMC in rodents following oral (8-800 mg/kg) intravenous (IV) (8 mg/kg) or dermal (0.8-80 mg/kg representing 0.1-10% formulation concentration) exposure to [ 14 C]EHMC were investigated in rats and mice. 3. EHMC was rapidly cleared from rat and mouse hepatocytes (half-life ≤3.16 min) and less rapidly (half-life ≤48 min) from human hepatocytes. 4. [ 14 C]EHMC was extensively absorbed and excreted primarily in urine by 72 h after oral administration to rats (65-80%) and mice (63-72%). Oral doses to rats were excreted to a lesser extent (3-8%) in feces and as CO 2 (1-4%). Radioactive residues in tissues were <1% of the dose. There were no sex or species differences in disposition in rats. 5. Following dermal application, 34-42% of an 8-mg/kg dose was absorbed in rats, and 54-62% in mice in 72-h. 6. Among numerous urinary metabolites associated with hydrolysis of the ester, two potential reproductive and developmental toxicants, 2-ethylhexanol and 2-ethylhexanoic acid were produced by metabolism of EHMC.

Optimisation of the differing conditions required for bone formation in vitro by primary osteoblasts from mice and rats

PubMed Central

ORRISS, ISABEL R.; HAJJAWI, MARK O.R.; HUESA, CARMEN; MACRAE, VICKY E.; ARNETT, TIMOTHY R.

2014-01-01

The in vitro culture of calvarial osteoblasts from neonatal rodents remains an important method for studying the regulation of bone formation. The widespread use of transgenic mice has created a particular need for a reliable, simple method that allows the differentiation and bone-forming activity of murine osteoblasts to be studied. In the present study, we established such a method and identified key differences in optimal culture conditions between mouse and rat osteoblasts. Cells isolated from neonatal rodent calvariae by collagenase digestion were cultured for 14–28 days before staining for tissue non-specific alkaline phosphatase (TNAP) and bone mineralisation (alizarin red). The reliable differentiation of mouse osteoblasts, resulting in abundant TNAP expression and the formation of mineralised ‘trabecular-shaped’ bone nodules, occurred only following culture in α minimum essential medium (αMEM) and took 21–28 days. Dexamethasone (10 nM) inhibited bone mineralisation in the mouse osteoblasts. By contrast, TNAP expression and bone formation by rat osteoblasts were observed following culture in both αMEM and Dulbecco’s modified Eagle’s medium (DMEM) after approximately 14 days (although ~3-fold more effectively in αMEM) and was strongly dependent on dexamethasone. Both the mouse and rat osteoblasts required ascorbate (50 μg/ml) for osteogenic differentiation and β-glycerophosphate (2 mM) for mineralisation. The rat and mouse osteoblasts showed similar sensitivity to the well-established inhibitors of mineralisation, inorganic pyrophosphate (PPi) and adenosine triphosphate (ATP; 1–100 μM). The high efficiency of osteogenic differentiation observed following culture in αMEM, compared with culture in DMEM possibly reflects the richer formulation of the former. These findings offer a reliable technique for inducing mouse osteoblasts to form bone in vitro and a more effective method for culturing bone-forming rat osteoblasts. PMID:25200658

Immunocompetent syngeneic cotton rat tumor models for the assessment of replication-competent oncolytic adenovirus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Steel, Jason C.; Morrison, Brian J.; Mannan, Poonam

Oncolytic adenoviruses as a treatment for cancer have demonstrated limited clinical activity. Contributing to this may be the relevance of preclinical animal models used to study these agents. Syngeneic mouse tumor models are generally non-permissive for adenoviral replication, whereas human tumor xenograft models exhibit attenuated immune responses to the vector. The cotton rat (Sigmodon hispidus) is susceptible to human adenovirus infection, permissive for viral replication and exhibits similar inflammatory pathology to humans with adenovirus replicating in the lungs, respiratory passages and cornea. We evaluated three transplantable tumorigenic cotton rat cell lines, CCRT, LCRT and VCRT as models for the studymore » of oncolytic adenoviruses. All three cells lines were readily infected with adenovirus type-5-based vectors and exhibited high levels of transgene expression. The cell lines supported viral replication demonstrated by the induction of cytopathogenic effect (CPE) in tissue culture, increase in virus particle numbers and assembly of virions seen on transmission electron microscopy. In vivo, LCRT and VCRT tumors demonstrated delayed growth after injection with replicating adenovirus. No in vivo antitumor activity was seen in CCRT tumors despite in vitro oncolysis. Adenovirus was also rapidly cleared from the CCRT tumors compared to LCRT and VCRT tumors. The effect observed with the different cotton rat tumor cell lines mimics the variable results of human clinical trials highlighting the potential relevance of this model for assessing the activity and toxicity of oncolytic adenoviruses.« less

Mouse chromosomal mapping of a murine leukemia virus integration region (Mis-1) first identified in rat thymic leukemia.

PubMed Central

Jolicoeur, P; Villeneuve, L; Rassart, E; Kozak, C

1985-01-01

We have previously identified a region of genomic DNA which constitutes the site of frequent provirus integration in rat thymomas induced by Moloney murine leukemia virus (Lemay and Jolicoeur, Proc. Natl. Acad. Sci. USA 81:38-42, 1984). This genetic locus is now designated Mis-1 (Moloney integration site). Cellular sequences homologous to Mis-1 are present in mouse DNA. Using a series of hamster-mouse somatic cell hybrids, we mapped the Mis-1 locus to mouse chromosome 15. Frequent chromosome 15 aberrations have been described in mouse thymomas. Mis-1 represents a putative new oncogene which might be involved in the initiation or maintenance or both of these neoplasms. Images PMID:4068142

Characterization of Differential Cocaine Metabolism in Mouse and Rat through Metabolomics-Guided Metabolite Profiling

PubMed Central

Yao, Dan; Shi, Xiaolei; Wang, Lei; Gosnell, Blake A.

2013-01-01

Rodent animal models have been widely used for studying neurologic and toxicological events associated with cocaine abuse. It is known that the mouse is more susceptible to cocaine-induced hepatotoxicity (CIH) than the rat. However, the causes behind this species-dependent sensitivity to cocaine have not been elucidated. In this study, cocaine metabolism in the mouse and rat was characterized through LC-MS-based metabolomic analysis of urine samples and were further compared through calculating the relative abundance of individual cocaine metabolites. The results showed that the levels of benzoylecgonine, a major cocaine metabolite from ester hydrolysis, were comparable in the urine from the mice and rats treated with the same dose of cocaine. However, the levels of the cocaine metabolites from oxidative metabolism, such as N-hydroxybenzoylnorecgonine and hydroxybenzoylecgonine, differed dramatically between the two species, indicating species-dependent cocaine metabolism. Subsequent structural analysis through accurate mass analysis and LC-MS/MS fragmentation revealed that N-oxidation reactions, including N-demethylation and N-hydroxylation, are preferred metabolic routes in the mouse, while extensive aryl hydroxylation reactions occur in the rat. Through stable isotope tracing and in vitro enzyme reactions, a mouse-specific α-glucoside of N-hydroxybenzoylnorecgonine and a group of aryl hydroxy glucuronides high in the rat were identified and structurally elucidated. The differences in the in vivo oxidative metabolism of cocaine between the two rodent species were confirmed by the in vitro microsomal incubations. Chemical inhibition of P450 enzymes further revealed that different P450-mediated oxidative reactions in the ecgonine and benzoic acid moieties of cocaine contribute to the species-dependent biotransformation of cocaine. PMID:23034697

Reactive oxygen species and fatigue-induced prolonged low-frequency force depression in skeletal muscle fibres of rats, mice and SOD2 overexpressing mice.

PubMed

Bruton, Joseph D; Place, Nicolas; Yamada, Takashi; Silva, José P; Andrade, Francisco H; Dahlstedt, Anders J; Zhang, Shi-Jin; Katz, Abram; Larsson, Nils-Göran; Westerblad, Håkan

2008-01-01

Skeletal muscle often shows a delayed force recovery after fatiguing stimulation, especially at low stimulation frequencies. In this study we focus on the role of reactive oxygen species (ROS) in this fatigue-induced prolonged low-frequency force depression. Intact, single muscle fibres were dissected from flexor digitorum brevis (FDB) muscles of rats and wild-type and superoxide dismutase 2 (SOD2) overexpressing mice. Force and myoplasmic free [Ca(2+)] ([Ca(2+)](i)) were measured. Fibres were stimulated at different frequencies before and 30 min after fatigue induced by repeated tetani. The results show a marked force decrease at low stimulation frequencies 30 min after fatiguing stimulation in all fibres. This decrease was associated with reduced tetanic [Ca(2+)](i) in wild-type mouse fibres, whereas rat fibres and mouse SOD2 overexpressing fibres instead displayed a decreased myofibrillar Ca(2+) sensitivity. The SOD activity was approximately 50% lower in wild-type mouse than in rat FDB muscles. Myoplasmic ROS increased during repeated tetanic stimulation in rat fibres but not in wild-type mouse fibres. The decreased Ca(2+) sensitivity in rat fibres could be partially reversed by application of the reducing agent dithiothreitol, whereas the decrease in tetanic [Ca(2+)](i) in wild-type mouse fibres was not affected by dithiothreitol or the antioxidant N-acetylcysteine. In conclusion, we describe two different causes of fatigue-induced prolonged low-frequency force depression, which correlate to differences in SOD activity and ROS metabolism. These findings may have clinical implications since ROS-mediated impairments in myofibrillar function can be counteracted by reductants and antioxidants, whereas changes in SR Ca(2+) handling appear more resistant to interventions.

Guinea Pig Chymase Is Leucine-specific

PubMed Central

Caughey, George H.; Beauchamp, Jeremy; Schlatter, Daniel; Raymond, Wilfred W.; Trivedi, Neil N.; Banner, David; Mauser, Harald; Fingerle, Jürgen

2008-01-01

To explore guinea pigs as models of chymase biology, we cloned and expressed the guinea pig ortholog of human chymase. In contrast to rats and mice, guinea pigs appear to express just one chymase, which belongs to the α clade, like primate chymases and mouse mast cell protease-5. The guinea pig enzyme autolyzes at Leu residues in the loop where human chymase autolyzes at Phe. In addition, guinea pig α-chymase selects P1 Leu in a combinatorial peptide library and cleaves Ala-Ala-Pro-Leu-4-nitroanilide but has negligible activity toward substrates with P1 Phe and does not cleave angiotensin I. This contrasts with human chymase, which cleaves after Phe or Tyr, prefers P1 Phe in peptidyl 4-nitroanilides, and avidly hydrolyzes angiotensin I at Phe8 to generate bioactive angiotensin II. The guinea pig enzyme also is inactivated more effectively by α1-antichymotrypsin, which features P1 Leu in the reactive loop. Unlike mouse, rat, and hamster α-chymases, guinea pig chymase lacks elastase-like preference for P1 Val or Ala. Partially humanized A216G guinea pig chymase acquires human-like P1 Phe- and angiotensin-cleaving capacity. Molecular models suggest that the wild type active site is crowded by the Ala216 side chain, which potentially blocks access by bulky P1 aromatic residues. On the other hand, the guinea pig pocket is deeper than in Val-selective chymases, explaining the preference for the longer aliphatic side chain of Leu. These findings are evidence that chymase-like peptidase specificity is sensitive to small changes in structure and provide the first example of a vertebrate Leu-selective peptidase. PMID:18353771

Use of the hyperinsulinemic euglycemic clamp to assess insulin sensitivity in guinea pigs: dose response, partitioned glucose metabolism, and species comparisons.

PubMed

Horton, Dane M; Saint, David A; Owens, Julie A; Gatford, Kathryn L; Kind, Karen L

2017-07-01

The guinea pig is an alternate small animal model for the study of metabolism, including insulin sensitivity. However, only one study to date has reported the use of the hyperinsulinemic euglycemic clamp in anesthetized animals in this species, and the dose response has not been reported. We therefore characterized the dose-response curve for whole body glucose uptake using recombinant human insulin in the adult guinea pig. Interspecies comparisons with published data showed species differences in maximal whole body responses (guinea pig ≈ human < rat < mouse) and the insulin concentrations at which half-maximal insulin responses occurred (guinea pig > human ≈ rat > mouse). In subsequent studies, we used concomitant d-[3- 3 H]glucose infusion to characterize insulin sensitivities of whole body glucose uptake, utilization, production, storage, and glycolysis in young adult guinea pigs at human insulin doses that produced approximately half-maximal (7.5 mU·min -1 ·kg -1 ) and near-maximal whole body responses (30 mU·min -1 ·kg -1 ). Although human insulin infusion increased rates of glucose utilization (up to 68%) and storage and, at high concentrations, increased rates of glycolysis in females, glucose production was only partially suppressed (~23%), even at high insulin doses. Fasting glucose, metabolic clearance of insulin, and rates of glucose utilization, storage, and production during insulin stimulation were higher in female than in male guinea pigs ( P < 0.05), but insulin sensitivity of these and whole body glucose uptake did not differ between sexes. This study establishes a method for measuring partitioned glucose metabolism in chronically catheterized conscious guinea pigs, allowing studies of regulation of insulin sensitivity in this species. Copyright © 2017 the American Physiological Society.

Molecular cloning and expression of the CRISP family of proteins in the boar.

PubMed

Vadnais, Melissa L; Foster, Douglas N; Roberts, Kenneth P

2008-12-01

The family of mammalian cysteine-rich secretory proteins (CRISP) have been well characterized in the rat, mouse, and human. Here we report the molecular cloning and expression analysis of CRISP1, CRISP2, and CRISP3 in the boar. A partial sequence published in the National Center for Biotechnology Information (NCBI) database was used to derive the full-length sequences for CRISP1 and CRISP2 using rapid amplification of cDNA ends. RT-PCR confirmed the expression of these mRNAs in the boar reproductive tract, and real time RT-PCR showed CRISP1 to be highly expressed throughout the epididymis, with CRISP2 highly expressed in the testis. A search of the porcine genomic sequence in the NCBI database identified a BAC (CH242-199E6) encoding the CRISP1 gene. This BAC is derived from porcine Chromosome 7 and is syntenic with the regions of the mouse, rat, and human genomes encoding the CRISP gene family. This BAC was found to encode a third CRISP protein with a predicted amino acid sequence of high similarity to human CRISP3. Using RT-PCR we show that CRISP3 expression in the boar reproductive tract is confined to the prostate. Recombinant porcine (rp) CRISP2 protein was produced and purified. When incubated with capacitated boar sperm, rpCRISP2 induced an acrosome reaction, consistent with its demonstrated ability to alter the activity of calcium channels.

Generating gene knockout rats by homologous recombination in embryonic stem cells

PubMed Central

Tong, Chang; Huang, Guanyi; Ashton, Charles; Li, Ping; Ying, Qi-Long

2013-01-01

We describe here a detailed protocol for generating gene knockout rats by homologous recombination in embryonic stem (ES) cells. This protocol comprises the following procedures: derivation and expansion of rat ES cells, construction of gene-targeting vectors, generation of gene-targeted rat ES cells and, finally, production of gene-targeted rats. The major differences between this protocol and the classical mouse gene-targeting protocol include ES cell culture methods, drug selection scheme, colony picking and screening strategies. This ES cell–based gene-targeting technique allows sophisticated genetic modifications to be performed in the rat, as many laboratories have been doing in the mouse for the past two decades. Recently we used this protocol to generate Tp53 (also known as p53) gene knockout rats. The entire process requires ~1 year to complete, from derivation of ES cells to generation of knockout rats. PMID:21637202

Yoso-tama-no-kakehashi; the first Japanese guidebook on raising rats.

PubMed

Kuramoto, Takashi

2011-01-01

During the 18th century, raising the "nezumi" rodent became so prevalent in Japan that two guidebooks were published on the topic. The first guidebook was entitled Yoso-tama-no-kakehashi (1775) and the second was entitled Chinganso-date-gusa (1787). It remains unclear in these texts whether the term nezumi was used to refer to the rat (Rattus norvegicus) or the mouse (Mus musculus). In this review, I explore Yoso-tama-no-kakehashi (English translation: A bridge to obtaining novel jewel-like nezumi). It was written by the owner of "Shunpo-do" and comprises two volumes; the first is 34 pages in length and the second is 14 pages. It introduces the nezumi and then provides details on novel varieties and the methods that were used to raise them. The nezumi dwells in peoples' homes. It is noteworthy that the "norako" species is classified in the same group as the nezumi. The norako is smaller than the nezumi. Its alias is "hatsuka-nezumi", a term which is still used in Japan today when referring to the mouse. This indicates that when the guidebook was written people distinguished the rat from the mouse by identifying the rat using the word nezumi and the mouse using the word norako. Moreover, I recently confirmed that the rat varieties which are introduced in Yoso-tama-no-kakehashi, such as "white", "spotted", "black bear-like", "deer-spotted", and "cracked-mark", can be found in modern laboratory rats. Taken together, it is very likely that the term nezumi is used to refer to the rat in Yoso-tama-no-kakehashi and that this was indeed a guidebook on the rat.

Electrochemical treatment of mouse and rat fibrosarcomas with direct current

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chou, C.K.; McDougall, J.A.; Ahn, C.

1997-03-01

Electrochemical treatment (ECT) of cancer utilizes direct current to produce chemical changes in tumors. ECT has been suggested as an effective alternative local cancer therapy. However, a methodology is not established, and mechanisms are not well studied. In vivo studies were conducted to evaluate the effectiveness of ECT on animal tumor models. Radiation-induced fibrosarcomas were implanted subcutaneously in 157 female C3H/HeJ mice. Larger rat fibrosarcomas were implanted on 34 female Fisher 344 rats. When the spheroidal tumors reached 10 mm in the mice, two to five platinum electrodes were inserted into the tumors at various spacings and orientations. Ten ratsmore » in a pilot group were treated when their ellipsoidal tumors were about 25 mm long; electrode insertion was similar to the later part of the mouse study; i.e., two at the base and two at the center. A second group of 24 rats was treated with six or seven electrodes when their tumors were about 20 mm long; all electrodes were inserted at the tumor base. Of the 24 rats, 12 of these were treated once, 10 were treated twice, and 2 were treated thrice. All treated tumors showed necrosis and regression for both mice and rats; however, later tumor recurrence reduced long-term survival. When multiple treatments were implemented, the best 3 month mouse tumor cure rate was 59.3%, and the best 6 month rat tumor cure rate was 75.0%. These preliminary results indicate that ECT is effective on the radiation-induced fibrosarcoma (RIF-1) mouse tumor and rat fibrosarcoma. The effectiveness is dependent on electrode placement and dosage.« less

A physiologically based pharmacokinetic model for atrazine and its main metabolites in the adult male C57BL/6 mouse

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin Zhoumeng; Interdisciplinary Toxicology Program, University of Georgia, Athens, GA 30602; Fisher, Jeffrey W.

Atrazine (ATR) is a chlorotriazine herbicide that is widely used and relatively persistent in the environment. In laboratory rodents, excessive exposure to ATR is detrimental to the reproductive, immune, and nervous systems. To better understand the toxicokinetics of ATR and to fill the need for a mouse model, a physiologically based pharmacokinetic (PBPK) model for ATR and its main chlorotriazine metabolites (Cl-TRIs) desethyl atrazine (DE), desisopropyl atrazine (DIP), and didealkyl atrazine (DACT) was developed for the adult male C57BL/6 mouse. Taking advantage of all relevant and recently made available mouse-specific data, a flow-limited PBPK model was constructed. The ATR andmore » DACT sub-models included blood, brain, liver, kidney, richly and slowly perfused tissue compartments, as well as plasma protein binding and red blood cell binding, whereas the DE and DIP sub-models were constructed as simple five-compartment models. The model adequately simulated plasma levels of ATR and Cl-TRIs and urinary dosimetry of Cl-TRIs at four single oral dose levels (250, 125, 25, and 5 mg/kg). Additionally, the model adequately described the dose dependency of brain and liver ATR and DACT concentrations. Cumulative urinary DACT amounts were accurately predicted across a wide dose range, suggesting the model's potential use for extrapolation to human exposures by performing reverse dosimetry. The model was validated using previously reported data for plasma ATR and DACT in mice and rats. Overall, besides being the first mouse PBPK model for ATR and its Cl-TRIs, this model, by analogy, provides insights into tissue dosimetry for rats. The model could be used in tissue dosimetry prediction and as an aid in the exposure assessment to this widely used herbicide.« less

KRAS-driven lung adenocarcinoma: combined DDR1/Notch inhibition as an effective therapy

PubMed Central

Ambrogio, Chiara; Nadal, Ernest; Villanueva, Alberto; Gómez-López, Gonzalo; Cash, Timothy P; Barbacid, Mariano; Santamaría, David

2016-01-01

Understanding the early evolution of cancer heterogeneity during the initial steps of tumorigenesis can uncover vulnerabilities of cancer cells that may be masked at later stages. We describe a comprehensive approach employing gene expression analysis in early lesions to identify novel therapeutic targets and the use of mouse models to test synthetic lethal drug combinations to treat human Kirsten rat sarcoma viral oncogene homologue (KRAS)-driven lung adenocarcinoma. PMID:27843638

A sense oligonucleotide to inducible nitric oxide synthase mRNA increases the survival rate of rats in septic shock.

PubMed

Okuyama, Tetsuya; Nakatake, Richi; Kaibori, Masaki; Okumura, Tadayoshi; Kon, Masanori; Nishizawa, Mikio

2018-01-30

Natural antisense transcripts (asRNAs) that do not encode proteins are transcribed from rat, mouse, and human genes, encoding inducible nitric oxide synthase (iNOS), which catalyzes the production of the inflammatory mediator nitric oxide (NO). In septic shock, NO is excessively produced in hepatocytes and macrophages. The iNOS asRNA interacts with and stabilizes iNOS mRNA. We found that single-stranded 'sense' oligonucleotides corresponding to the iNOS mRNA sequence reduced iNOS mRNA levels by interfering with the mRNA-asRNA interactions in rat hepatocytes. The iNOS sense oligonucleotides that were substituted with phosphorothioate bonds and locked nucleic acids efficiently decreased the levels of iNOS mRNA and iNOS protein. In this study, the gene expression patterns in the livers of two endotoxemia model rats with acute liver failure were compared. Next, we optimized the sequence and modification of the iNOS sense oligonucleotides in interleukin 1β-treated rat hepatocytes. When a sense oligonucleotide was simultaneously administered with d-galactosamine and bacterial lipopolysaccharide (LPS) to rats, their survival rate significantly increased compared to the rats administered d-galactosamine and LPS alone. In the livers of the sense oligonucleotide-administered rats, apoptosis in the hepatocytes markedly decreased. These results suggest that natural antisense transcript-targeted regulation technology using iNOS sense oligonucleotides may be used to treat human inflammatory diseases, such as sepsis and septic shock. Copyright © 2017 Elsevier Inc. All rights reserved.

Changes in consensus arylamine N-acetyltransferase (NAT) gene nomenclature

PubMed Central

Hein, David W.; Boukouvala, Sotiria; Grant, Denis M.; Minchin, Rodney F.; Sim, Edith

2008-01-01

Changes in consensus arylamine N-acetyltransferase (NAT) gene nomenclature determined at the 2007 international NAT workshop include: 1) Alleles in all species except mouse and rat are all uppercase. For mouse and rat, the first letter is upper case followed by lower case. 2) The nomenclature system is now species-specific. Thus, NAT2*1 (chicken), NAT2*2 & NAT2*3 (rabbit), Nat2*8 Nat2*9, Nat2*22 & Nat2*23 (mouse), NAT2*15, NAT2*16A & NAT2*16B (Syrian hamster), and NAT2*20, NAT2*21A & NAT2*21B (rat) are retired and renumbered within a species. A species modifier incorporated into the allele designation is written in upper case Roman font, e.g., (MOUSE)Nat1*1 is now the reference Nat1 allele in mouse; and 3) The NAT website also can now be accessed at a webbalias address: http://N-acetyltransferasenomenclature.louisville.edu. New NAT alleles should continue to be submitted to the NAT nomenclature committee for inclusion on the website to ensure proper categorization and to continue consistency in nomenclature. PMID:18334921

Novel and existing data for a future physiological toxicokinetic model of ethylene and its metabolite ethylene oxide in mouse, rat, and human.

PubMed

Filser, Johannes Georg; Artati, Anna; Li, Qiang; Pütz, Christian; Semder, Brigitte; Klein, Dominik; Kessler, Winfried

2015-11-05

The olefin ethylene is a ubiquitously found gas. It originates predominantly from plants, combustion processes and industrial sources. In mammals, inhaled ethylene is metabolized by cytochrome P450-dependent monooxygenases, particularly by cytochrome P450 2E1, to ethylene oxide, an epoxide that directly alkylates proteins and DNA. Ethylene oxide was mutagenic in vitro and in vivo in insects and mammals and carcinogenic in rats and mice. A physiological toxicokinetic model is a most useful tool for estimating the ethylene oxide burden in ethylene-exposed rodents and humans. The only published physiological toxicokinetic model for ethylene and metabolically produced ethylene oxide is discussed. Additionally, existing data required for the development of a future model and for testing its predictive accuracy are reviewed and extended by new gas uptake studies with ethylene and ethylene oxide in B6C3F1 mice and with ethylene in F344 rats. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Clearance of Human IgG1-Sensitised Red Blood Cells In Vivo in Humans Relates to the In Vitro Properties of Antibodies from Alternative Cell Lines

PubMed Central

Armour, Kathryn L.; Smith, Cheryl S.; Ip, Natasha C. Y.; Ellison, Cara J.; Kirton, Christopher M.; Wilkes, Anthony M.; Williamson, Lorna M.; Clark, Michael R.

2014-01-01

We previously produced a recombinant version of the human anti-RhD antibody Fog-1 in the rat myeloma cell line, YB2/0. When human, autologous RhD-positive red blood cells (RBC) were sensitised with this IgG1 antibody and re-injected, they were cleared much more rapidly from the circulation than had been seen earlier with the original human-mouse heterohybridoma-produced Fog-1. Since the IgG have the same amino acid sequence, this disparity is likely to be due to alternative glycosylation that results from the rat and mouse cell lines. By comparing the in vitro properties of YB2/0-produced Fog-1 IgG1 and the same antibody produced in the mouse myeloma cell line NS0, we now have a unique opportunity to pinpoint the cause of the difference in ability to clear RBC in vivo. Using transfected cell lines that express single human FcγR, we showed that IgG1 made in YB2/0 and NS0 cell lines bound equally well to receptors of the FcγRI and FcγRII classes but that the YB2/0 antibody was superior in FcγRIII binding. When measuring complexed IgG binding, the difference was 45-fold for FcγRIIIa 158F, 20-fold for FcγRIIIa 158V and approximately 40-fold for FcγRIIIb. The dissimilarity was greater at 100-fold in monomeric IgG binding assays with FcγRIIIa. When used to sensitise RBC, the YB2/0 IgG1 generated 100-fold greater human NK cell antibody-dependent cell-mediated cytotoxicity and had a 103-fold advantage over the NS0 antibody in activating NK cells, as detected by CD54 levels. In assays of monocyte activation and macrophage adherence/phagocytosis, where FcγRI plays major roles, RBC sensitised with the two antibodies produced much more similar results. Thus, the alternative glycosylation profiles of the Fog-1 antibodies affect only FcγRIII binding and FcγRIII-mediated functions. Relating this to the in vivo studies confirms the importance of FcγRIII in RBC clearance. PMID:25302805

Clearance of human IgG1-sensitised red blood cells in vivo in humans relates to the in vitro properties of antibodies from alternative cell lines.

PubMed

Armour, Kathryn L; Smith, Cheryl S; Ip, Natasha C Y; Ellison, Cara J; Kirton, Christopher M; Wilkes, Anthony M; Williamson, Lorna M; Clark, Michael R

2014-01-01

We previously produced a recombinant version of the human anti-RhD antibody Fog-1 in the rat myeloma cell line, YB2/0. When human, autologous RhD-positive red blood cells (RBC) were sensitised with this IgG1 antibody and re-injected, they were cleared much more rapidly from the circulation than had been seen earlier with the original human-mouse heterohybridoma-produced Fog-1. Since the IgG have the same amino acid sequence, this disparity is likely to be due to alternative glycosylation that results from the rat and mouse cell lines. By comparing the in vitro properties of YB2/0-produced Fog-1 IgG1 and the same antibody produced in the mouse myeloma cell line NS0, we now have a unique opportunity to pinpoint the cause of the difference in ability to clear RBC in vivo. Using transfected cell lines that express single human FcγR, we showed that IgG1 made in YB2/0 and NS0 cell lines bound equally well to receptors of the FcγRI and FcγRII classes but that the YB2/0 antibody was superior in FcγRIII binding. When measuring complexed IgG binding, the difference was 45-fold for FcγRIIIa 158F, 20-fold for FcγRIIIa 158V and approximately 40-fold for FcγRIIIb. The dissimilarity was greater at 100-fold in monomeric IgG binding assays with FcγRIIIa. When used to sensitise RBC, the YB2/0 IgG1 generated 100-fold greater human NK cell antibody-dependent cell-mediated cytotoxicity and had a 103-fold advantage over the NS0 antibody in activating NK cells, as detected by CD54 levels. In assays of monocyte activation and macrophage adherence/phagocytosis, where FcγRI plays major roles, RBC sensitised with the two antibodies produced much more similar results. Thus, the alternative glycosylation profiles of the Fog-1 antibodies affect only FcγRIII binding and FcγRIII-mediated functions. Relating this to the in vivo studies confirms the importance of FcγRIII in RBC clearance.

Covalent modification of proteins by cocaine

NASA Astrophysics Data System (ADS)

Deng, Shi-Xian; Bharat, Narine; Fischman, Marian C.; Landry, Donald W.

2002-03-01

Cocaine covalently modifies proteins through a reaction in which the methyl ester of cocaine acylates the -amino group of lysine residues. This reaction is highly specific in vitro, because no other amino acid reacts with cocaine, and only cocaine's methyl ester reacts with the lysine side chain. Covalently modified proteins were present in the plasma of rats and human subjects chronically exposed to cocaine. Modified endogenous proteins are immunogenic, and specific antibodies were elicited in mouse and detected in the plasma of human subjects. Covalent modification of proteins could explain cocaine's autoimmune effects and provide a new biochemical approach to cocaine's long-term actions.

Automated evaluation of liver fibrosis in thioacetamide, carbon tetrachloride, and bile duct ligation rodent models using second-harmonic generation/two-photon excited fluorescence microscopy.

PubMed

Liu, Feng; Chen, Long; Rao, Hui-Ying; Teng, Xiao; Ren, Ya-Yun; Lu, Yan-Qiang; Zhang, Wei; Wu, Nan; Liu, Fang-Fang; Wei, Lai

2017-01-01

Animal models provide a useful platform for developing and testing new drugs to treat liver fibrosis. Accordingly, we developed a novel automated system to evaluate liver fibrosis in rodent models. This system uses second-harmonic generation (SHG)/two-photon excited fluorescence (TPEF) microscopy to assess a total of four mouse and rat models, using chemical treatment with either thioacetamide (TAA) or carbon tetrachloride (CCl 4 ), and a surgical method, bile duct ligation (BDL). The results obtained by the new technique were compared with that using Ishak fibrosis scores and two currently used quantitative methods for determining liver fibrosis: the collagen proportionate area (CPA) and measurement of hydroxyproline (HYP) content. We show that 11 shared morphological parameters faithfully recapitulate Ishak fibrosis scores in the models, with high area under the receiver operating characteristic (ROC) curve (AUC) performance. The AUC values of 11 shared parameters were greater than that of the CPA (TAA: 0.758-0.922 vs 0.752-0.908; BDL: 0.874-0.989 vs 0.678-0.966) in the TAA mice and BDL rat models and similar to that of the CPA in the TAA rat and CCl 4 mouse models. Similarly, based on the trends in these parameters at different time points, 9, 10, 7, and 2 model-specific parameters were selected for the TAA rats, TAA mice, CCl 4 mice, and BDL rats, respectively. These parameters identified differences among the time points in the four models, with high AUC accuracy, and the corresponding AUC values of these parameters were greater compared with those of the CPA in the TAA rat and mouse models (rats: 0.769-0.894 vs 0.64-0.799; mice: 0.87-0.93 vs 0.739-0.836) and similar to those of the CPA in the CCl 4 mouse and BDL rat models. Similarly, the AUC values of 11 shared parameters and model-specific parameters were greater than those of HYP in the TAA rats, TAA mice, and CCl 4 mouse models and were similar to those of HYP in the BDL rat models. The automated evaluation system, combined with 11 shared parameters and model-specific parameters, could specifically, accurately, and quantitatively stage liver fibrosis in animal models.

Role of the gastrointestinal ecosystem in the development of type 1 diabetes.

PubMed

Daft, Joseph G; Lorenz, Robin G

2015-09-01

A new emphasis has been put on the role of the gastrointestinal (GI) ecosystem in autoimmune diseases; however, there is limited knowledge about its role in type 1 diabetes (T1D). Distinct differences have been observed in intestinal permeability, epithelial barrier function, commensal microbiota, and mucosal innate and adaptive immunity of patients and animals with T1D, when compared with healthy controls. The non-obese diabetic (NOD) mouse and the BioBreeding diabetes prone (BBdp) rat are the most commonly used models to study T1D pathogenesis. With the increasing awareness of the importance of the GI ecosystem in systemic disease, it is critical to understand the basics, as well as the similarities and differences between rat and mouse models and human patients. This review examines the current knowledge of the role of the GI ecosystem in T1D and indicates the extensive opportunities for further investigation that could lead to biomarkers and therapeutic interventions for disease prevention and/or modulation. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Abnormal notochord branching is associated with foregut malformations in the adriamycin treated mouse model.

PubMed

Hajduk, Piotr; Sato, Hideaki; Puri, Prem; Murphy, Paula

2011-01-01

Oesophageal atresia (OA) and tracheooesophageal fistula (TOF) are relatively common human congenital malformations of the foregut where the oesophagus does not connect with the stomach and there is an abnormal connection between the stomach and the respiratory tract. They require immediate corrective surgery and have an impact on the future health of the individual. These abnormalities are mimicked by exposure of rat and mouse embryos in utero to the drug adriamycin. The causes of OA/TOF during human development are not known, however a number of mouse mutants where different signalling pathways are directly affected, show similar abnormalities, implicating multiple and complex signalling mechanisms. The similarities in developmental outcome seen in human infants and in the adriamycin treated mouse model underline the potential of this model to unravel the early embryological events and further our understanding of the processes disturbed, leading to such abnormalities. Here we report a systematic study of the foregut and adjacent tissues in embryos treated with adriamycin at E7 and E8 and analysed between E9 and E12, comparing morphology in 3D in 149 specimens. We describe a spectrum of 8 defects, the most common of which is ventral displacement and branching of the notochord (in 94% of embryos at E10) and a close spatial correspondence between the site of notochord branching and defects of the foregut. In addition gene expression analysis shows altered dorso-ventral foregut patterning in the vicinity of notochord branches. This study shows a number of features of the adriamycin mouse model not previously reported, implicates the notochord as a primary site of disturbance in such abnormalities and underlines the importance of the model to further address the mechanistic basis of foregut congenital abnormalities.

Abnormal Notochord Branching Is Associated with Foregut Malformations in the Adriamycin Treated Mouse Model

PubMed Central

Hajduk, Piotr; Sato, Hideaki; Puri, Prem; Murphy, Paula

2011-01-01

Oesophageal atresia (OA) and tracheooesophageal fistula (TOF) are relatively common human congenital malformations of the foregut where the oesophagus does not connect with the stomach and there is an abnormal connection between the stomach and the respiratory tract. They require immediate corrective surgery and have an impact on the future health of the individual. These abnormalities are mimicked by exposure of rat and mouse embryos in utero to the drug adriamycin. The causes of OA/TOF during human development are not known, however a number of mouse mutants where different signalling pathways are directly affected, show similar abnormalities, implicating multiple and complex signalling mechanisms. The similarities in developmental outcome seen in human infants and in the adriamycin treated mouse model underline the potential of this model to unravel the early embryological events and further our understanding of the processes disturbed, leading to such abnormalities. Here we report a systematic study of the foregut and adjacent tissues in embryos treated with adriamycin at E7 and E8 and analysed between E9 and E12, comparing morphology in 3D in 149 specimens. We describe a spectrum of 8 defects, the most common of which is ventral displacement and branching of the notochord (in 94% of embryos at E10) and a close spatial correspondence between the site of notochord branching and defects of the foregut. In addition gene expression analysis shows altered dorso-ventral foregut patterning in the vicinity of notochord branches. This study shows a number of features of the adriamycin mouse model not previously reported, implicates the notochord as a primary site of disturbance in such abnormalities and underlines the importance of the model to further address the mechanistic basis of foregut congenital abnormalities. PMID:22132119

Studies on the Detection, Expression, Glycosylation, Dimerization, and Ligand Binding Properties of Mouse Siglec-E*

PubMed Central

Siddiqui, Shoib; Schwarz, Flavio; Springer, Stevan; Khedri, Zahra; Yu, Hai; Deng, Lingquan; Verhagen, Andrea; Naito-Matsui, Yuko; Jiang, Weiping; Kim, Daniel; Zhou, Jie; Ding, Beibei; Chen, Xi; Varki, Nissi; Varki, Ajit

2017-01-01

CD33-related Siglecs are a family of proteins widely expressed on innate immune cells. Binding of sialylated glycans or other ligands triggers signals that inhibit or activate inflammation. Immunomodulation by Siglecs has been extensively studied, but relationships between structure and functions are poorly explored. Here we present new data relating to the structure and function of Siglec-E, the major CD33-related Siglec expressed on mouse neutrophils, monocytes, macrophages, and dendritic cells. We generated nine new rat monoclonal antibodies specific to mouse Siglec-E, with no cross-reactivity to Siglec-F. Although all antibodies detected Siglec-E on transfected human HEK-293T cells, only two reacted with mouse bone marrow neutrophils by flow cytometry and on spleen sections by immunohistochemistry. Moreover, whereas all antibodies recognized Siglec-E-Fc on immunoblots, binding was dependent on intact disulfide bonds and N-glycans, and only two antibodies recognized native Siglec-E within spleen lysates. Thus, we further investigated the impact of Siglec-E homodimerization. Homology-based structural modeling predicted a cysteine residue (Cys-298) in position to form a disulfide bridge between two Siglec-E polypeptides. Mutagenesis of Cys-298 confirmed its role in dimerization. In keeping with the high level of 9-O-acetylation found in mice, sialoglycan array studies indicate that this modification has complex effects on recognition by Siglec-E, in relationship to the underlying structures. However, we found no differences in phosphorylation or SHP-1 recruitment between dimeric and monomeric Siglec-E expressed on HEK293A cells. Phylogenomic analyses predicted that only some human and mouse Siglecs form disulfide-linked dimers. Notably, Siglec-9, the functionally equivalent human paralog of Siglec-E, occurs as a monomer. PMID:27920204

Negligible Colon Cancer Risk from Food-Borne Acrylamide Exposure in Male F344 Rats and Nude (nu/nu) Mice-Bearing Human Colon Tumor Xenografts

PubMed Central

Raju, Jayadev; Roberts, Jennifer; Sondagar, Chandni; Kapal, Kamla; Aziz, Syed A.; Caldwell, Don; Mehta, Rekha

2013-01-01

Acrylamide, a possible human carcinogen, is formed in certain carbohydrate-rich foods processed at high temperature. We evaluated if dietary acrylamide, at doses (0.5, 1.0 or 2.0 mg/kg diet) reflecting upper levels found in human foods, modulated colon tumorigenesis in two rodent models. Male F344 rats were randomized to receive diets without (control) or with acrylamide. 2-weeks later, rats in each group received two weekly subcutaneous injections of either azoxymethane (AOM) or saline, and were killed 20 weeks post-injections; colons were assessed for tumors. Male athymic nude (nu/nu) mice bearing HT-29 human colon adenocarcinoma cells-derived tumor xenografts received diets without (control) or with acrylamide; tumor growth was monitored and mice were killed 4 weeks later. In the F344 rat study, no tumors were found in the colons of the saline-injected rats. However, the colon tumor incidence was 54.2% and 66.7% in the control and the 2 mg/kg acrylamide-treated AOM-injected groups, respectively. While tumor multiplicity was similar across all diet groups, tumor size and burden were higher in the 2 mg/kg acrylamide group compared to the AOM control. These results suggest that acrylamide by itself is not a “complete carcinogen”, but acts as a “co-carcinogen” by exacerbating the effects of AOM. The nude mouse study indicated no differences in the growth of human colon tumor xenografts between acrylamide-treated and control mice, suggesting that acrylamide does not aid in the progression of established tumors. Hence, food-borne acrylamide at levels comparable to those found in human foods is neither an independent carcinogen nor a tumor promoter in the colon. However, our results characterize a potential hazard of acrylamide as a colon co-carcinogen in association with known and possibly other environmental tumor initiators/promoters. PMID:24040114

Proceedings of the 2010 National Toxicology Program Satellite Symposium

PubMed Central

Adams, E. Terence; Auerbach, Scott; Blackshear, Pamela E.; Bradley, Alys; Gruebbel, Margarita M.; Little, Peter B.; Malarkey, David; Maronpot, Robert; McKay, Jennifer S.; Miller, Rodney A.; Moore, Rebecca R.; Morrison, James P.; Nyska, Abraham; Ramot, Yuval; Rao, Deepa; Suttie, Andrew; Wells, Monique Y.; Willson, Gabrielle A.; Elmore, Susan A.

2011-01-01

The 2010 annual National Toxicology Program (NTP) Satellite Symposium, entitled “Pathology Potpourri,” was held in Chicago, Illinois, in advance of the scientific symposium sponsored jointly by the Society of Toxicologic Pathology (STP) and the International Federation of Societies of Toxicologic Pathologists (IFSTP). The goal of the annual NTP Symposium is to present current diagnostic pathology or nomenclature issues to the toxicologic pathology community. This article presents summaries of the speakers' presentations, including diagnostic or nomenclature issues that were presented, along with select images that were used for voting or discussion. Some topics covered during the symposium included a comparison of rat and mouse hepatocholangiocarcinoma, a comparison of cholangiofibrosis and cholangiocarcinoma in rats, a mixed pancreatic neoplasm with acinar and islet cell components, an unusual preputial gland tumor, renal hyaline glomerulopathy in rats and mice, eosinophilic substance in the nasal septum of mice, INHAND nomenclature for proliferative and nonproliferative lesions of the CNS/PNS, retinal gliosis in a rat, fibroadnexal hamartoma in rats, intramural plaque in a mouse, a treatment-related chloracne-like lesion in mice, and an overview of mouse ovarian tumors. PMID:21177527

Species-specific susceptibility to cannabis-induced convulsions.

PubMed

Whalley, Benjamin J; Lin, Hong; Bell, Lynne; Hill, Thomas; Patel, Amesha; Gray, Roy A; Elizabeth Roberts, C; Devinsky, Orrin; Bazelot, Michael; Williams, Claire M; Stephens, Gary J

2018-02-19

Numerous claims are made for cannabis' therapeutic utility upon human seizures, but concerns persist about risks. A potential confounder is the presence of both Δ 9 -tetrahydrocannabinol (THC), variously reported to be pro- and anticonvulsant, and cannabidiol (CBD), widely confirmed as anticonvulsant. Therefore, we investigated effects of prolonged exposure to different THC/CBD cannabis extracts on seizure activity and associated measures of endocannabinoid (eCB) system signalling. Cannabis extract effects on in vivo neurological and behavioural responses, and on bioanalyte levels, were measured in rats and dogs. Extract effects on seizure activity were measured using electroencephalography telemetry in rats. eCB signalling was also investigated using radioligand binding in cannabis extract-treated rats and treatment-naïve rat, mouse, chicken, dog and human tissue. Prolonged exposure to cannabis extracts caused spontaneous, generalized seizures, subserved by epileptiform discharges in rats, but not dogs, and produced higher THC, but lower 11-hydroxy-THC (11-OH-THC) and CBD, plasma concentrations in rats versus dogs. In the same rats, prolonged exposure to cannabis also impaired cannabinoid type 1 receptor (CB 1 receptor)-mediated signalling. Profiling CB 1 receptor expression, basal activity, extent of activation and sensitivity to THC suggested interspecies differences in eCB signalling, being more pronounced in a species that exhibited cannabis extract-induced seizures (rat) than one that did not (dog). Sustained cannabis extract treatment caused differential seizure, behavioural and bioanalyte levels between rats and dogs. Supporting radioligand binding data suggest species differences in eCB signalling. Interspecies variations may have important implications for predicting cannabis-induced convulsions from animal models. © 2018 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.

Translational Mouse Models of Autism: Advancing Toward Pharmacological Therapeutics

PubMed Central

Kazdoba, Tatiana M.; Leach, Prescott T.; Yang, Mu; Silverman, Jill L.; Solomon, Marjorie

2016-01-01

Animal models provide preclinical tools to investigate the causal role of genetic mutations and environmental factors in the etiology of autism spectrum disorder (ASD). Knockout and humanized knock-in mice, and more recently knockout rats, have been generated for many of the de novo single gene mutations and copy number variants (CNVs) detected in ASD and comorbid neurodevelopmental disorders. Mouse models incorporating genetic and environmental manipulations have been employed for preclinical testing of hypothesis-driven pharmacological targets, to begin to develop treatments for the diagnostic and associated symptoms of autism. In this review, we summarize rodent behavioral assays relevant to the core features of autism, preclinical and clinical evaluations of pharmacological interventions, and strategies to improve the translational value of rodent models of autism. PMID:27305922

Xenogeneic spermatogenesis following transplantation of hamster germ cells to mouse testes.

PubMed

Ogawa, T; Dobrinski, I; Avarbock, M R; Brinster, R L

1999-02-01

It was recently demonstrated that rat spermatogenesis can occur in the seminiferous tubules of an immunodeficient recipient mouse after transplantation of testis cells from a donor rat. In the present study, hamster donor testis cells were transplanted to mice to determine whether xenogeneic spermatogenesis would result. The hamster diverged at least 16 million years ago from the mouse and produces spermatozoa that are larger than, and have a shape distinctly different from, those of the mouse. In four separate experiments with a total of 13 recipient mice, hamster spermatogenesis was identified in the testes of each mouse. Approximately 6% of the tubules examined demonstrated xenogeneic spermatogenesis. In addition, cryopreserved hamster testis cells generated spermatogenesis in recipients. However, abnormalities were noted in hamster spermatids and acrosomes in seminiferous tubules of recipient mice. Hamster spermatozoa were also found in the epididymis of recipient animals, but these spermatozoa generally lacked acrosomes, and heads and tails were separated. Thus, defects in spermiogenesis occur in hamster spermatogenesis in the mouse, which may reflect a limited ability of endogenous mouse Sertoli cells to support fully the larger and evolutionarily distant hamster germ cell. The generation of spermatogenesis from frozen hamster cells now adds this species to the mouse and rat, in which spermatogonial stem cells also can be cryopreserved. This finding has immediate application to valuable animals of many species, because the cells could be stored until suitable recipients are identified or culture techniques devised to expand the stem cell population.

Food habits of rodents inhabiting arid and semi-arid ecosystems of central New Mexico

USGS Publications Warehouse

Hope, Andrew G.; Parmenter, Robert R.

2007-01-01

In this study, we describe seasonal dietary composition for 15 species of rodents collected in all major habitats on the Sevilleta National Wildlife Refuge (Socorro County) in central New Mexico. A comprehensive literature review of food habits for these species from throughout their distribution also is provided. We collected rodents in the field during winter, spring and late summer in 1998 from six communities: riparian cottonwood forest; piñon-juniper woodland; juniper-oak savanna; mesquite savanna; short-grass steppe; and Chihuahuan Desert scrubland. Rodents included Spermophilus spilosoma (Spotted Ground Squirrel), Perognathus flavescens (Plains Pocket Mouse), Perognathus flavus (Silky Pocket Mouse), Dipodomys merriami (Merriam’s Kangaroo Rat), Dipodomys ordii (Ord’s Kangaroo Rat), Dipodomys spectabilis (Banner-tailed Kangaroo Rat), Reithrodontomys megalotis (Western Harvest Mouse), Peromyscus boylii (Brush Mouse), Peromyscus eremicus (Cactus Mouse), Peromyscus leucopus (White-footed Mouse), Peromyscus truei (Piñon Mouse), Onychomys arenicola (Mearn’s Grasshopper Mouse), Onychomys leucogaster (Northern Grasshopper Mouse), Neotoma albigula/leucodon (White-throated Woodrats), and Neotoma micropus (Southern Plains Woodrat). We collected stomach contents of all species, and cheek-pouch contents of heteromyids, and quantified them in the laboratory. We determined seasonal diets in each habitat by calculating mean percentage volumes of seeds, arthropods and green vegetation (plant leaves and stems) for each species of rodent. Seeds consumed by each rodent were identified to genus, and often species, and quantified by frequency counts. Comparisons of diets between and among species of rodents, seasons, and ecosystems were also examined. We provide an appendix of all plant taxa documented.

NS6180, a new KCa3.1 channel inhibitor prevents T-cell activation and inflammation in a rat model of inflammatory bowel disease

PubMed Central

Strøbæk, D; Brown, DT; Jenkins, DP; Chen, Y-J; Coleman, N; Ando, Y; Chiu, P; Jørgensen, S; Demnitz, J; Wulff, H; Christophersen, P

2013-01-01

Background and Purpose The KCa3.1 channel is a potential target for therapy of immune disease. We identified a compound from a new chemical class of KCa3.1 inhibitors and assessed in vitro and in vivo inhibition of immune responses. Experimental Approach We characterized the benzothiazinone NS6180 (4-[[3-(trifluoromethyl)phenyl]methyl]-2H-1,4-benzothiazin-3(4H)-one) with respect to potency and molecular site of action on KCa3.1 channels, selectivity towards other targets, effects on T-cell activation as well as pharmacokinetics and inflammation control in colitis induced by 2,4-dinitrobenzene sulfonic acid, a rat model of inflammatory bowel disease (IBD). Key Results NS6180 inhibited cloned human KCa3.1 channels (IC50 = 9 nM) via T250 and V275, the same amino acid residues conferring sensitivity to triarylmethanes such as like TRAM-34. NS6180 inhibited endogenously expressed KCa3.1 channels in human, mouse and rat erythrocytes, with similar potencies (15–20 nM). NS6180 suppressed rat and mouse splenocyte proliferation at submicrolar concentrations and potently inhibited IL-2 and IFN-γ production, while exerting smaller effects on IL-4 and TNF-α and no effect on IL-17 production. Antibody staining showed KCa3.1 channels in healthy colon and strong up-regulation in association with infiltrating immune cells after induction of colitis. Despite poor plasma exposure, NS6180 (3 and 10 mg·kg−1 b.i.d.) dampened colon inflammation and improved body weight gain as effectively as the standard IBD drug sulfasalazine (300 mg·kg−1 q.d.). Conclusions and Implications NS6180 represents a novel class of KCa3.1 channel inhibitors which inhibited experimental colitis, suggesting KCa3.1 channels as targets for pharmacological control of intestinal inflammation. PMID:22891655

Localization of organ-specific antigens in the nervous system of the rat.

PubMed

Weinrauder, H; Lach, B

1977-08-16

Localization of organ-specific brain antigens in the central nervous system of the rat has been studied by means of indirect immunofluorescence. Rabbit antiserum against homogenate of rat brain, previously absorbed with normal serum and homogenates of rat organs (kidney, liver, spleen), reacted with the water-soluble antigens of rat brain prepared by extraction with phosphate buffer (pH 7.3) and ultracentrifugation at 50 000 X g to give one band in the immunodiffusion test and 2--3 precipitation arcs in immunoelectrophoresis. There was also a positive reaction with peripheral nerve. The antigen was detectable in all regions of the CNS. Cells with distinct cytoplasmic immunofluorescence were most frequently observed in cerebellar white matter, pons, cerebellar pedunculi, longitudinal tracts of the brain stem. Positive immunofluorecence reaction has appeared in the outer plexiform layer and granular layer of the retina, satelite cells of the spinal root ganglia and Schwann cells. A similar reaction was observed in human, mouse and guinea pig brain slices. Both the morphological and immunochemical reactions are indicative of glial localization of this antigen.

The multidrug resistance 1 gene Abcb1 in brain and placenta: comparative analysis in human and guinea pig.

PubMed

Pappas, Jane J; Petropoulos, Sophie; Suderman, Matthew; Iqbal, Majid; Moisiadis, Vasilis; Turecki, Gustavo; Matthews, Stephen G; Szyf, Moshe

2014-01-01

The Multidrug Resistance 1 (MDR1; alternatively ABCB1) gene product P-glycoprotein (P-gp), an ATP binding cassette transporter, extrudes multiple endogenous and exogenous substrates from the cell, playing an important role in normal physiology and xenobiotic distribution and bioavailability. To date, the predominant animal models used to investigate the role of P-gp have been the mouse and rat, which have two distinct genes, Abcb1a and Abcb1b. In contrast, the human has a single gene, ABCB1, for which only a single isoform has been validated. We and others have previously shown important differences between Abcb1a and Abcb1b, limiting the extrapolation from rodent findings to the human. Since the guinea pig has a relatively long gestation, hemomonochorial placentation and neuroanatomically mature offspring, it is more similar to the human, and may provide a more comparable model for investigating the regulation of P-gp in the brain and placenta, however, to date, the Abcb1 gene in the guinea pig remains to be characterized. The placenta and fetal brain are barrier sites that express P-gp and that play a critical role of protection of the fetus and the fetal brain from maternally administered drugs and other xenobiotics. Using RNA sequencing (RNA-seq), reverse transcription-polymerase chain reaction (RT-PCR) and quantitative PCR (QPCR) to sequence the expressed isoforms of guinea pig Abcb1, we demonstrate that like the human, the guinea pig genome contains one gene for Abcb1 but that it is expressed as at least three different isoforms via alternative splicing and alternate exon usage. Further, we demonstrate that these isoforms are more closely related to human than to rat or mouse isoforms. This striking, overall similarity and evolutionary relatedness between guinea pig Abcb1 and human ABCB1 indicate that the guinea pig represents a relevant animal model for investigating the function and regulation of P-gp in the placenta and brain.

Isolation and sequence of partial cDNA clones of human L1: homology of human and rodent L1 in the cytoplasmic region.

PubMed

Harper, J R; Prince, J T; Healy, P A; Stuart, J K; Nauman, S J; Stallcup, W B

1991-03-01

We have isolated cDNA clones coding for the human homologue of the neuronal cell adhesion molecule L1. The nucleotide sequence of the cDNA clones and the deduced primary amino acid sequence of the carboxy terminal portion of the human L1 are homologous to the corresponding sequences of mouse L1 and rat NILE glycoprotein, with an especially high sequences identity in the cytoplasmic regions of the proteins. There is also protein sequence homology with the cytoplasmic region of the Drosophila cell adhesion molecule, neuroglian. The conservation of the cytoplasmic domain argues for an important functional role for this portion of the molecule.

Tofogliflozin, a potent and highly specific sodium/glucose cotransporter 2 inhibitor, improves glycemic control in diabetic rats and mice.

PubMed

Suzuki, Masayuki; Honda, Kiyofumi; Fukazawa, Masanori; Ozawa, Kazuharu; Hagita, Hitoshi; Kawai, Takahiro; Takeda, Minako; Yata, Tatsuo; Kawai, Mio; Fukuzawa, Taku; Kobayashi, Takamitsu; Sato, Tsutomu; Kawabe, Yoshiki; Ikeda, Sachiya

2012-06-01

Sodium/glucose cotransporter 2 (SGLT2) is the predominant mediator of renal glucose reabsorption and is an emerging molecular target for the treatment of diabetes. We identified a novel potent and selective SGLT2 inhibitor, tofogliflozin (CSG452), and examined its efficacy and pharmacological properties as an antidiabetic drug. Tofogliflozin competitively inhibited SGLT2 in cells overexpressing SGLT2, and K(i) values for human, rat, and mouse SGLT2 inhibition were 2.9, 14.9, and 6.4 nM, respectively. The selectivity of tofogliflozin toward human SGLT2 versus human SGLT1, SGLT6, and sodium/myo-inositol transporter 1 was the highest among the tested SGLT2 inhibitors under clinical development. Furthermore, no interaction with tofogliflozin was observed in any of a battery of tests examining glucose-related physiological processes, such as glucose uptake, glucose oxidation, glycogen synthesis, hepatic glucose production, glucose-stimulated insulin secretion, and glucosidase reactions. A single oral gavage of tofogliflozin increased renal glucose clearance and lowered the blood glucose level in Zucker diabetic fatty rats. Tofogliflozin also improved postprandial glucose excursion in a meal tolerance test with GK rats. In db/db mice, 4-week tofogliflozin treatment reduced glycated hemoglobin and improved glucose tolerance in the oral glucose tolerance test 4 days after the final administration. No blood glucose reduction was observed in normoglycemic SD rats treated with tofogliflozin. These findings demonstrate that tofogliflozin inhibits SGLT2 in a specific manner, lowers blood glucose levels by increasing renal glucose clearance, and improves pathological conditions of type 2 diabetes with a low hypoglycemic potential.

Human Usher 1B/mouse shaker-1: the retinal phenotype discrepancy explained by the presence/absence of myosin VIIA in the photoreceptor cells.

PubMed

el-Amraoui, A; Sahly, I; Picaud, S; Sahel, J; Abitbol, M; Petit, C

1996-08-01

Usher syndrome type 1 (USH1) associates severe congenital deafness, vestibular dysfunction and progressive retinitis pigmentosa leading to blindness. The gene encoding myosin VIIA is responsible for USH1B. Mutations in the murine orthologous gene lead to the shaker-1 phenotype, which manifests cochlear and vestibular dysfunction, without any retinal defect. To address this phenotypic discrepancy, the expression of myosin VIIA in retinal cells was analyzed in human and mouse during embryonic development and adult life. In the human embryo, myosin VIIA was present first in the pigment epithelium cells, and later in these cells as well as in the photoreceptor cells. In the adult human retina, myosin VIIA was present in both cell types. In contrast, in mouse, only pigment epithelium cells expressed the protein throughout development and adult life. Myosin VIIA was also found to be absent in the photoreceptor cells of other rodents (rat and guinea-pig), whereas these cells expressed the protein in amphibians, avians and primates. These observations suggest that retinitis pigmentosa of USH1B results from a primary rod and cone defect. The USH1B/shaker-1 paradigm illustrates a species-specific cell pattern of gene expression as a possible cause for the discrepancy between phenotypes involving defective orthologous genes in man and mouse. Interestingly, in the photoreceptor cells, myosin VIIA is mainly localized in the inner and base of outer segments as well as in the synaptic ending region where it is co-localized with the synaptic vesicles. Therefore, we suggest that myosin VIIA might play a role in the trafficking of ribbon-synaptic vesicle complexes and the renewal processes of the outer photoreceptor disks.

Remarkable sequence conservation of the last intron in the PKD1 gene.

PubMed

Rodova, Marianna; Islam, M Rafiq; Peterson, Kenneth R; Calvet, James P

2003-10-01

The last intron of the PKD1 gene (intron 45) was found to have exceptionally high sequence conservation across four mammalian species: human, mouse, rat, and dog. This conservation did not extend to the comparable intron in pufferfish. Pairwise comparisons for intron 45 showed 91% identity (human vs. dog) to 100% identity (mouse vs. rat) for an average for all four species of 94% identity. In contrast, introns 43 and 44 of the PKD1 gene had average pairwise identities of 57% and 54%, and exons 43, 44, and 45 and the coding region of exon 46 had average pairwise identities of 80%, 84%, 82%, and 80%. Intron 45 is 90 to 95 bp in length, with the major region of sequence divergence being in a central 4-bp to 9-bp variable region. RNA secondary structure analysis of intron 45 predicts a branching stem-loop structure in which the central variable region lies in one loop and the putative branch point sequence lies in another loop, suggesting that the intron adopts a specific stem-loop structure that may be important for its removal. Although intron 45 appears to conform to the class of small, G-triplet-containing introns that are spliced by a mechanism utilizing intron definition, its high sequence conservation may be a reflection of constraints imposed by a unique mechanism that coordinates splicing of this last PKD1 intron with polyadenylation.

Tissue-specific DNA methylation is conserved across human, mouse, and rat, and driven by primary sequence conservation.

PubMed

Zhou, Jia; Sears, Renee L; Xing, Xiaoyun; Zhang, Bo; Li, Daofeng; Rockweiler, Nicole B; Jang, Hyo Sik; Choudhary, Mayank N K; Lee, Hyung Joo; Lowdon, Rebecca F; Arand, Jason; Tabers, Brianne; Gu, C Charles; Cicero, Theodore J; Wang, Ting

2017-09-12

Uncovering mechanisms of epigenome evolution is an essential step towards understanding the evolution of different cellular phenotypes. While studies have confirmed DNA methylation as a conserved epigenetic mechanism in mammalian development, little is known about the conservation of tissue-specific genome-wide DNA methylation patterns. Using a comparative epigenomics approach, we identified and compared the tissue-specific DNA methylation patterns of rat against those of mouse and human across three shared tissue types. We confirmed that tissue-specific differentially methylated regions are strongly associated with tissue-specific regulatory elements. Comparisons between species revealed that at a minimum 11-37% of tissue-specific DNA methylation patterns are conserved, a phenomenon that we define as epigenetic conservation. Conserved DNA methylation is accompanied by conservation of other epigenetic marks including histone modifications. Although a significant amount of locus-specific methylation is epigenetically conserved, the majority of tissue-specific DNA methylation is not conserved across the species and tissue types that we investigated. Examination of the genetic underpinning of epigenetic conservation suggests that primary sequence conservation is a driving force behind epigenetic conservation. In contrast, evolutionary dynamics of tissue-specific DNA methylation are best explained by the maintenance or turnover of binding sites for important transcription factors. Our study extends the limited literature of comparative epigenomics and suggests a new paradigm for epigenetic conservation without genetic conservation through analysis of transcription factor binding sites.

Phospholipid epitopes for mouse antibodies against bromelain-treated mouse erythrocytes.

PubMed Central

Kawaguchi, S

1987-01-01

The reactivity of mouse antibodies against bromelain-treated mouse erythrocytes (BrMRBC) with phospholipid epitopes was assessed by ELISA, using four clones of monoclonal anti-BrMRBC antibodies that had idiotypes distinct from one another. The four antibodies could bind to low-density lipoproteins (LDL) from human and chicken, but not to LDL from mouse and rat. As to liposomes of natural phospholipids, all the clones reacted with liposomes of phosphatidylcholine, and some of them could react with liposomes of sphingomyelin, phosphatidylglycerol, phosphatidylic acid or cardiolipin. For liposomes of synthetic phosphatidylcholine with different fatty acids, the length of carbon chains and the number of unsaturated carbon chains of the fatty acids markedly affected the binding of each monoclonal antibody to the liposomes. The addition of dicetyl phosphate or stearylamine to phosphatidylcholine liposomes changed the reactivity of the liposomes. These results support the view that mouse anti-BrMRBC antibodies can recognize appropriately spaced phosphorylcholine residues on the surface of phospholipid liposomes, LDL and cells. The four clones had similar capacities for binding to LDL as well as to BrMRBC, but they had obviously different capacities for binding to phospholipid liposomes; the epitopes on phospholipid liposomes used in the present study were not so perfect as to react well with every anti-BrMRBC antibody. PMID:2443446

AFPep: an anti-breast cancer peptide that is orally active.

PubMed

Bennett, James A; DeFreest, Lori; Anaka, Ikenna; Saadati, Hamid; Balulad, Sujata; Jacobson, Herbert I; Andersen, Thomas T

2006-07-01

We have synthesized a cyclic nonapeptide (AFPep) that is effective, after being administered by parenteral routes, for the treatment or the prevention of breast cancer. To test the hypothesis that AFPep remains safe and efficacious after oral administration, three different whole-animal bioassays were utilized, and the mechanism by which AFPep functions was investigated. Using a human breast cancer xenograft model in mice for therapeutic activity, a carcinogen-induced breast cancer model in rats for prevention efficacy, and a mouse uterus growth inhibition model of anti-estrogenic activity, AFPep was administered by oral gavage (p.o.) and its effects compared to those following intraperitoneal (i.p.) and subcutaneous (s.c.) administration. Toxicity studies evaluated body weights and organ weights in mice and rats receiving AFPep. Preliminary mechanistic studies were carried out in T47D human breast cancer cells growing in culture and evaluated the effect of AFPep on estrogen-stimulated cell growth, phosphorylation of the estrogen receptor (ER), and on level of ER-related kinases. Orally administered AFPep stopped the growth of human tumor xenografts in mice, decreased the incidence and multiplicity of breast cancers in carcinogen-exposed rats, and inhibited the estrogen-stimulated growth of mouse uteri. In each of these systems, orally administered AFPep produced an effect similar to that obtained for AFPep administered by either i.p or s.c. routes. In rodents, no evidence of toxicity was seen for the peptide, even at very high doses. In culture, AFPep inhibited the estrogen-stimulated growth, but not the basal growth, of T47D cells, and it inhibited the estrogen-stimulated phosphorylation of Serine 118 in the ER of these cells, which was not explainable by early changes in ER-related kinases. Chronic oral administration of AFPep appears to be safe and effective for the treatment or prevention of breast cancer in animal models.

Immune recognition of botulinum neurotoxin B: antibody-binding regions on the heavy chain of the toxin.

PubMed

Dolimbek, Behzod Z; Steward, Lance E; Aoki, K Roger; Atassi, M Zouhair

2008-02-01

The purpose of this work was to map the continuous regions recognized by human, horse and mouse anti-botulinum neurotoxin B (BoNT/B) antibodies (Abs). We synthesized a panel of sixty 19-residue peptides (peptide C31 was 24 residues) that overlapped consecutively by 5 residues and together encompassed the entire heavy chain of BoNT/B (H/B, residues 442-1291). Abs from the three host species recognized similar, but not identical, peptides. There were also peptides recognized by two or only by one host species. Where a peptide was recognized by Abs of more than one host species, these Abs were at different levels among the species. Human, horse and mouse Abs bound, although in different amounts, to regions within peptides 736-754, 778-796, 848-866, 932-950, 974-992, 1058-1076 and 1128-1146. Human and horse Abs bound to peptides 890-908 and 1170-1188. Human and mouse Abs recognized peptides 470-488/484-502 overlap, 638-656, 722-740, 862-880, 1030-1048, 1072-1090, 1240-1258 and 1268-1291. We concluded that the antigenic regions localized with the three antisera are quite similar, exhibiting in some cases a small shift to the left or to the right. This is consistent with what is known about protein immune recognition. In the three-dimensional structure, the regions recognized on H/B by anti-BoNT/B Abs occupied surface locations and analysis revealed no correlation between these surface locations and surface electrostatic potential, hydrophilicity, hydrophobicity, or temperature factor. A region that bound mouse Abs overlapped with a recently defined site on BoNT/B that binds to mouse and rat synaptotagmin II, thus providing a molecular explanation for the blocking (protecting) activity of these Abs. The regions thus localized afford candidates for incorporation into a synthetic vaccine design.

Modulation of neuronal sodium channels by the sea anemone peptide BDS-I

PubMed Central

Liu, Pin; Jo, Sooyeon

2012-01-01

Blood-depressing substance I (BDS-I), a 43 amino-acid peptide from sea anemone venom, is used as a specific inhibitor of Kv3-family potassium channels. We found that BDS-I acts with even higher potency to modulate specific types of voltage-dependent sodium channels. In rat dorsal root ganglion (DRG) neurons, 3 μM BDS-I strongly enhanced tetrodotoxin (TTX)-sensitive sodium current but weakly inhibited TTX-resistant sodium current. In rat superior cervical ganglion (SCG) neurons, which express only TTX-sensitive sodium current, BDS-I enhanced current elicited by small depolarizations and slowed decay of currents at all voltages (EC50 ∼ 300 nM). BDS-I acted with exceptionally high potency and efficacy on cloned human Nav1.7 channels, slowing inactivation by 6-fold, with an EC50 of approximately 3 nM. BDS-I also slowed inactivation of sodium currents in N1E-115 neuroblastoma cells (mainly from Nav1.3 channels), with an EC50 ∼ 600 nM. In hippocampal CA3 pyramidal neurons (mouse) and cerebellar Purkinje neurons (mouse and rat), BDS-I had only small effects on current decay (slowing inactivation by 20–50%), suggesting relatively weak sensitivity of Nav1.1 and Nav1.6 channels. The biggest effect of BDS-I in central neurons was to enhance resurgent current in Purkinje neurons, an effect reflected in enhancement of sodium current during the repolarization phase of Purkinje neuron action potentials. Overall, these results show that BDS-I acts to modulate sodium channel gating in a manner similar to previously known neurotoxin receptor site 3 anemone toxins but with different isoform sensitivity. Most notably, BDS-I acts with very high potency on human Nav1.7 channels. PMID:22442564

Toxicity and Carcinogenicity Studies of Ginkgo biloba extract in Rat and Mouse: Liver, Thyroid, and Nose are Targets

PubMed Central

Rider, Cynthia V.; Nyska, Abraham; Cora, Michelle C.; Kissling, Grace E.; Smith, Cynthia; Travlos, Gregory S.; Hejtmancik, Milton R.; Fomby, Laurene M.; Colleton, Curtis A.; Ryan, Michael J.; Kooistra, Linda; Morrison, James P.; Chan, Po C.

2014-01-01

Ginkgo biloba extract (GBE) is a popular herbal supplement that is used to improve circulation and brain function. In spite of widespread human exposure to relatively high doses over potentially long periods of time, there is a paucity of data from animal studies regarding the toxicity and carcinogenicity associated with GBE. In order to fill this knowledge gap, three-month and two-year toxicity and carcinogenicity studies with GBE administered by oral gavage to B6C3F1/N mice and F344/N rats were performed as part of the National Toxicology Program’s Dietary Supplements and Herbal Medicines Initiative. The targets of GBE treatment were the liver, thyroid, and nose. These targets were consistent across exposure period, sex, and species, albeit with varying degrees of effect observed among studies. Key findings included a notably high incidence of hepatoblastomas in male and female mice and evidence of carcinogenic potential in the thyroid gland of both mice and rats. Various nonneoplastic lesions were observed beyond control levels in the liver, thyroid gland, and nose of rats and mice administered GBE. Although these results cannot be directly extrapolated to humans, the findings fill an important data gap in assessing risk associated with GBE use. PMID:23960164

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buchholz, Bruce A.

The project has two main goals: 1) Identify the types of adducts naphthalene (NA) forms with DNA and 2) determine whether adduct formation correlates with site selective tumor formation in defined subcompartments of the respiratory tract (respiratory and olfactory nasal epithelium and airways of mice, rats and rhesus monkeys). Five tasks are associated with the completion of the goals. Task 1: Contracting and Animal Use Approvals. IACUC and ACURO approvals are complete, The subcontract with UC Davis (UCD) was executed in December 2014. Task 2: Perform In Vitro Study for Goal 1. Rat samples exposed and in freezer while adductmore » standards are being made. Mouse samples need to be exposed in next quarter. Task 3: Perform In Vitro Study for Goal 2. Mouse ex vivo samples completed. Rat and monkey samples need to be completed in the next quarter. Task 4: Sample Preparation and Analysis. Mouse Goal 2 samples completed. Other samples remain to be done. Task 5: Data Interpretation and Reporting. Need rat data to write paper on adduct formation.« less

[Values of the micronucleus test on animal epithelial cells exposed to titanium dioxide].

PubMed

Iurchenko, V V; Krivtsova, E K; Iuretseva, N A; Tul'skaia, E A; Mamonov, R A; Zholdakova, Z I; Sinitsyna, O O; Mal'tseva, M M; Pankratova, G P; Sycheva, L P

2011-01-01

The genetic safety of titanium dioxide (TD)-containing foods and cosmetic products has been little investigated. The study evaluated the mutagenic activity of TD in the micronucleus test with animal visceral mucosal epithelial cells. Two simethicone-coated anatase samples (mean size 160 and 33.2 nm) were inserted into the mouse stomach in doses of 40-200-1000 mg/kg seven times and applied as an ingredient of 10 and 25% cream (doses 250 and 625 mg/kg, respectively) to the hair-sheared rat skin once for 4 hours. Analysis of cytogenetic disorders (micronuclei, protrusions, and the atypical form of the nucleus) revealed no mutagenic properties of TD on the mucosal epithelium of the mouse and rat intestine, mouse prostomach, and rat uterine bladder. Enhanced mitotic activity was observed in all the study tissues after exposure of both samples to TD given in some or in all (in the rat urinary bladder mucosal epithelium) doses.

Oracle, a novel PDZ-LIM domain protein expressed in heart and skeletal muscle.

PubMed

Passier, R; Richardson, J A; Olson, E N

2000-04-01

In order to identify novel genes enriched in adult heart, we performed a subtractive hybridization for genes expressed in mouse heart but not in skeletal muscle. We identified two alternative splicing variants of a novel PDZ-LIM domain protein, which we named Oracle. Both variants contain a PDZ domain at the amino-terminus and three LIM domains at the carboxy-terminus. Highest homology of Oracle was found with the human and rat enigma proteins in the PDZ domain (62 and 61%, respectively) and in the LIM domains (60 and 69%, respectively). By Northern hybridization analysis, we showed that expression is highest in adult mouse heart, low in skeletal muscle and undetectable in other adult mouse tissues. In situ hybridization in mouse embryos confirmed and extended these data by showing high expression of Oracle mRNA in atrial and ventricular myocardial cells from E8.5. From E9.5 low expression of Oracle mRNA was detectable in myotomes. These data suggest a role for Oracle in the early development and function of heart and skeletal muscle.

Obeticholic acid raises LDL-cholesterol and reduces HDL-cholesterol in the Diet-Induced NASH (DIN) hamster model.

PubMed

Briand, François; Brousseau, Emmanuel; Quinsat, Marjolaine; Burcelin, Rémy; Sulpice, Thierry

2018-01-05

The use of rat and mouse models limits the translation to humans for developing novel drugs targeting nonalcoholic steatohepatitis (NASH). Obeticholic acid (OCA) illustrates this limitation since its dyslipidemic effect in humans cannot be observed in these rodents. Conversely, Golden Syrian hamsters have a lipoprotein metabolism mimicking human dyslipidemia since it does express the cholesteryl ester transfer protein (CETP). We therefore developed a Diet-Induced NASH (DIN) hamster model and evaluated the impact of OCA. Compared with chow fed controls, hamsters fed for 20 weeks with a free-choice (FC) diet, developed obesity, insulin resistance, dyslipidemia and NASH (microvesicular steatosis, inflammation, hepatocyte ballooning and perisinusoidal to bridging fibrosis). After 20 weeks of diet, FC fed hamsters were treated without or with obeticholic acid (15mg/kg/day) for 5 weeks. Although a non-significant trend towards higher dietary caloric intake was observed, OCA significantly lowered body weight after 5 weeks of treatment. OCA significantly increased CETP activity and LDL-C levels by 20% and 27%, and reduced HDL-C levels by 20%. OCA blunted hepatic gene expression of Cyp7a1 and Cyp8b1 and reduced fecal bile acids mass excretion by 64% (P < 0.05). Hamsters treated with OCA showed a trend towards higher scavenger receptor Class B type I (SR-BI) and lower LDL-receptor hepatic protein expression. OCA reduced NAS score for inflammation (P < 0.01) and total NAS score, although not significantly. Compared to mouse and rat models, the DIN hamster replicates benefits and side effects of OCA as observed in humans, and should be useful for evaluating novel drugs targeting NASH. Copyright © 2017 Elsevier B.V. All rights reserved.

Somatic cell hybrid mapping on mouse chromosome 11 (MMU11): Assignment of markers relative to two breakpoints in band D

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morris, D.J.; Robinson, T.J.; Adler, I.D.

1993-02-01

Mouse [times] rat somatic cell hybrids were generated by fusing mouse cell lines that are heterozygous for reciprocal translocations involving the T42H and T9Ad breakpoints on mouse chromosome 11 (MMU11) to a thymidine kinase-negative (Tk[sup [minus

Port d’Entrée for Respiratory Infections – Does the Influenza A Virus Pave the Way for Bacteria?

PubMed Central

Siemens, Nikolai; Oehmcke-Hecht, Sonja; Mettenleiter, Thomas C.; Kreikemeyer, Bernd; Valentin-Weigand, Peter; Hammerschmidt, Sven

2017-01-01

Bacterial and viral co-infections of the respiratory tract are life-threatening and present a global burden to the global community. Staphylococcus aureus, Streptococcus pneumoniae, and Streptococcus pyogenes are frequent colonizers of the upper respiratory tract. Imbalances through acquisition of seasonal viruses, e.g., Influenza A virus, can lead to bacterial dissemination to the lower respiratory tract, which in turn can result in severe pneumonia. In this review, we summarize the current knowledge about bacterial and viral co-infections of the respiratory tract and focus on potential experimental models suitable for mimicking this disease. Transmission of IAV and pneumonia is mainly modeled by mouse infection. Few studies utilizing ferrets, rats, guinea pigs, rabbits, and non-human primates are also available. The knowledge gained from these studies led to important discoveries and advances in understanding these infectious diseases. Nevertheless, mouse and other infection models have limitations, especially in translation of the discoveries to humans. Here, we suggest the use of human engineered lung tissue, human ex vivo lung tissue, and porcine models to study respiratory co-infections, which might contribute to a greater translation of the results to humans and improve both, animal and human health. PMID:29312268

Localization of alpha-dystroglycan on the podocyte: from top to toe.

PubMed

Vogtländer, Nils P J; Dijkman, Henry; Bakker, Marinka A H; Campbell, Kevin P; van der Vlag, Johan; Berden, Jo H M

2005-11-01

alpha-Dystroglycan (DG) is a negatively charged membrane-associated glycoprotein that links the cytoskeleton to the extracellular matrix. Previously, we described that alpha-DG covers the whole podocyte cell membrane in the rat. However, our finding was challenged by the description of a strictly basolateral localization in human kidney biopsies, using a different antibody against alpha-DG. Therefore, we studied the exact localization of glomerular alpha-DG by using these two antibodies in both species. The studies were performed by using monoclonal antibodies (MoAbs) IIH6 and VIA4.1 in immunofluorescence, confocal microscopy, and immunoelectron microscopy on both rat and human kidney sections, as well as on cultured mouse podocytes. The apical localization of alpha-DG on podocytes was more dominant than the basolateral localization. The basolateral staining with MoAb VIA4.1 was more pronounced than that of MoAb IIH6. With both MoAbs, the staining in rat kidneys was more prominent, in comparison to human kidneys. We conclude that alpha-DG is expressed at both the basolateral and apical sides of the podocyte. This localization suggests that alpha-DG plays a dual role in the maintenance of the unique architecture of podocytes by its binding to the glomerular basement membrane, and in the maintenance of the integrity of the filtration slit, respectively.

A neuropeptide ligand of the G protein-coupled receptor GPR103 regulates feeding, behavioral arousal, and blood pressure in mice.

PubMed

Takayasu, Shinobu; Sakurai, Takeshi; Iwasaki, Satoshi; Teranishi, Hitoshi; Yamanaka, Akihiro; Williams, S Clay; Iguchi, Haruhisa; Kawasawa, Yuka Imamura; Ikeda, Yukio; Sakakibara, Iori; Ohno, Kousaku; Ioka, Ryoichi X; Murakami, Saori; Dohmae, Naoshi; Xie, Jian; Suda, Toshihiro; Motoike, Toshiyuki; Ohuchi, Takashi; Yanagisawa, Masashi; Sakai, Juro

2006-05-09

Here, we report the isolation and characterization of an endogenous peptide ligand of GPR103 from rat brains. The purified peptide was found to be the 43-residue RF-amide peptide QRFP. We also describe two mouse homologues of human GPR103, termed mouse GPR103A and GPR103B. QRFP binds and activates the human GPR103, as well as mouse GPR103A and GPR103B, with nanomolar affinities in transfected cells. Systematic in situ hybridization analysis in mouse brains showed that QRFP is expressed exclusively in the periventricular and lateral hypothalamus, whereas the two receptor mRNAs are distinctly localized in various brain areas without an overlap to each other. When administered centrally in mice, QRFP induced feeding behavior, accompanied by increased general locomotor activity and metabolic rate. QRFP-induced food intake was abolished by preadministration of BIBP3226, a specific antagonist for the Y1 neuropeptide Y receptor. Hypothalamic prepro-QRFP mRNA expression was up-regulated upon fasting and in genetically obese ob/ob and db/db mice. Central QRFP administration also evoked highly sustained elevation of blood pressure and heart rate. Our findings suggest that QRFP and GPR103A/B may regulate diverse neuroendocrine and behavioral functions and implicate this neuropeptide system in metabolic syndrome.

Selective binding of lectins to normal and neoplastic urothelium in rat and mouse bladder carcinogenesis models.

PubMed

Zupančič, Daša; Kreft, Mateja Erdani; Romih, Rok

2014-01-01

Bladder cancer adjuvant intravesical therapy could be optimized by more selective targeting of neoplastic tissue via specific binding of lectins to plasma membrane carbohydrates. Our aim was to establish rat and mouse models of bladder carcinogenesis to investigate in vivo and ex vivo binding of selected lectins to the luminal surface of normal and neoplastic urothelium. Male rats and mice were treated with 0.05 % N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in drinking water and used for ex vivo and in vivo lectin binding experiments. Urinary bladder samples were also used for paraffin embedding, scanning electron microscopy and immunofluorescence labelling of uroplakins. During carcinogenesis, the structure of the urinary bladder luminal surface changed from microridges to microvilli and ropy ridges and the expression of urothelial-specific glycoproteins uroplakins was decreased. Ex vivo and in vivo lectin binding experiments gave comparable results. Jacalin (lectin from Artocarpus integrifolia) exhibited the highest selectivity for neoplastic compared to normal urothelium of rats and mice. The binding of lectin from Amaranthus caudatus decreased in rat model and increased in mouse carcinogenesis model, indicating interspecies variations of plasma membrane glycosylation. Lectin from Datura stramonium showed higher affinity for neoplastic urothelium compared to the normal in rat and mouse model. The BBN-induced animal models of bladder carcinogenesis offer a promising approach for lectin binding experiments and further lectin-mediated targeted drug delivery research. Moreover, in vivo lectin binding experiments are comparable to ex vivo experiments, which should be considered when planning and optimizing future research.

MECHANISTIC CONSIDERATIONS FOR HUMAN RELEVANCE OF CANCER HAZARD OF DI(2-ETHYLHEXYL) PHTHALATE

PubMed Central

Rusyn, Ivan; Corton, J. Christopher

2012-01-01

Di(2-ethylhexyl) phthalate (DEHP) is a peroxisome proliferator agent that is widely used as a plasticizer to soften polyvinylchloride plastics and non-polymers. Both occupational (e.g., by inhalation during its manufacture and use as a plasticizer of polyvinylchloride) and environmental (medical devices, contamination of food, or intake from air, water and soil) routes of exposure to DEHP are of concern for human health. There is sufficient evidence for carcinogenicity of DEHP in the liver in both rats and mice; however, there is little epidemiological evidence on possible associations between exposure to DEHP and liver cancer in humans. Data are available to suggest that liver is not the only target tissue for DEHP-associated toxicity and carcinogenicity in both humans and rodents. The debate regarding human relevance of the findings in rats or mice has been informed by studies on the mechanisms of carcinogenesis of the peroxisome proliferator class of chemicals, including DEHP. Important additional mechanistic information became available in the past decade, including, but not limited to, sub-acute, sub-chronic and chronic studies with DEHP in peroxisome proliferator-activated receptor (PPAR) α-null mice, as well as experiments utilizing several transgenic mouse lines. Activation of PPARα and the subsequent downstream events mediated by this transcription factor represent an important mechanism of action for DEHP in rats and mice. However, additional data from animal models and studies in humans exposed to DEHP from the environment suggest that multiple molecular signals and pathways in several cell types in the liver, rather than a single molecular event, contribute to the cancer in rats and mice. In addition, the toxic and carcinogenic effects of DEHP are not limited to liver. The International Agency for Research on Cancer working group concluded that the human relevance of the molecular events leading to cancer elicited by DEHP in several target tissues (e.g., liver and testis) in rats and mice can not be ruled out and DEHP was classified as possibly carcinogenic to humans (Group 2B). PMID:22198209

Molecular Determinants of the Human α2C-Adrenergic Receptor Temperature-Sensitive Intracellular Traffic

PubMed Central

Pullikuth, Ashok K.; Guidry, Jessie J.

2015-01-01

The human α2C-adrenergic receptor (α2C-AR) is localized intracellularly at physiologic temperature. Decreasing the environmental temperature strongly stimulates the receptor transport to the cell surface. In contrast, rat and mouse α2C-AR plasma membrane levels are less sensitive to decrease in temperature, whereas the opossum α2C-AR cell surface levels are not changed in these conditions. Structural analysis demonstrated that human α2C-AR has a high number of arginine residues in the third intracellular loop and in the C-terminus, organized as putative RXR motifs. Although these motifs do not affect the receptor subcellular localization at 37°C, deletion of the arginine clusters significantly enhanced receptor plasma membrane levels at reduced temperature. We found that this exaggerated transport of the human receptor is mediated by two functional arginine clusters, one in the third intracellular loop and one in the C-terminus. This effect is mediated by interactions with COPI vesicles, but not by 14-3-3 proteins. In rat α2C-AR, the arginine cluster from the third intracellular loop is shifted to the left due to three missing residues. Reinsertion of these residues in the rat α2C-AR restored the same temperature sensitivity as in the human receptor. Proteomic and coimmunoprecipitation experiments identified pontin as a molecule having stronger interactions with human α2C-AR compared with rat α2C-AR. Inhibition of pontin activity enhanced human receptor plasma membrane levels and signaling at 37°C. Our results demonstrate that human α2C-AR has a unique temperature-sensitive traffic pattern within the G protein–coupled receptor class due to interactions with different molecular chaperones, mediated in part by strict spatial localization of specific arginine residues. PMID:25680754

Teratology studies in the mouse.

PubMed

Marsden, Edward; Leroy, Mariline

2013-01-01

The rat is the routine species of choice as the rodent model for regulatory safety testing of xenobiotics such as medicinal products, food additives, and other chemicals. However, the rat is not always suitable for pharmacological, toxicological, immunogenic, pharmacokinetic, or even practical reasons. Under such circumstances, the mouse offers an alternative for finding a suitable rodent model acceptable to the regulatory authorities. Since all essential routes of administration are possible, the short reproductive cycle and large litter size of the mouse make it a species well adapted for use in teratology studies. Given that good quality animals, including virgin mated females, can be acquired relatively easily and inexpensively, the mouse has been used in reproductive toxicity studies for decades and study protocols are well established.

78 FR 8410 - Thiacloprid; Pesticide Tolerances

Federal Register 2010, 2011, 2012, 2013, 2014

2013-02-06

...) were noted in dog, rat, and mouse studies. Increased prostate weight and prostatic hypertrophy were... acetylcholine receptors. In the neurotoxicity studies in rats, there was a reduction in motor and locomotor... uterine tumors in rats, thyroid follicular adenomas in rat and ovarian tumors in mice. A cancer slope...

Super-resolution imaging of subcortical white matter using stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI)

PubMed Central

Hainsworth, A. H.; Lee, S.; Patel, A.; Poon, W. W.; Knight, A. E.

2018-01-01

Aims The spatial resolution of light microscopy is limited by the wavelength of visible light (the ‘diffraction limit’, approximately 250 nm). Resolution of sub-cellular structures, smaller than this limit, is possible with super resolution methods such as stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI). We aimed to resolve subcellular structures (axons, myelin sheaths and astrocytic processes) within intact white matter, using STORM and SOFI. Methods Standard cryostat-cut sections of subcortical white matter from donated human brain tissue and from adult rat and mouse brain were labelled, using standard immunohistochemical markers (neurofilament-H, myelin-associated glycoprotein, glial fibrillary acidic protein, GFAP). Image sequences were processed for STORM (effective pixel size 8–32 nm) and for SOFI (effective pixel size 80 nm). Results In human, rat and mouse, subcortical white matter high-quality images for axonal neurofilaments, myelin sheaths and filamentous astrocytic processes were obtained. In quantitative measurements, STORM consistently underestimated width of axons and astrocyte processes (compared with electron microscopy measurements). SOFI provided more accurate width measurements, though with somewhat lower spatial resolution than STORM. Conclusions Super resolution imaging of intact cryo-cut human brain tissue is feasible. For quantitation, STORM can under-estimate diameters of thin fluorescent objects. SOFI is more robust. The greatest limitation for super-resolution imaging in brain sections is imposed by sample preparation. We anticipate that improved strategies to reduce autofluorescence and to enhance fluorophore performance will enable rapid expansion of this approach. PMID:28696566

Super-resolution imaging of subcortical white matter using stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI).

PubMed

Hainsworth, A H; Lee, S; Foot, P; Patel, A; Poon, W W; Knight, A E

2018-06-01

The spatial resolution of light microscopy is limited by the wavelength of visible light (the 'diffraction limit', approximately 250 nm). Resolution of sub-cellular structures, smaller than this limit, is possible with super resolution methods such as stochastic optical reconstruction microscopy (STORM) and super-resolution optical fluctuation imaging (SOFI). We aimed to resolve subcellular structures (axons, myelin sheaths and astrocytic processes) within intact white matter, using STORM and SOFI. Standard cryostat-cut sections of subcortical white matter from donated human brain tissue and from adult rat and mouse brain were labelled, using standard immunohistochemical markers (neurofilament-H, myelin-associated glycoprotein, glial fibrillary acidic protein, GFAP). Image sequences were processed for STORM (effective pixel size 8-32 nm) and for SOFI (effective pixel size 80 nm). In human, rat and mouse, subcortical white matter high-quality images for axonal neurofilaments, myelin sheaths and filamentous astrocytic processes were obtained. In quantitative measurements, STORM consistently underestimated width of axons and astrocyte processes (compared with electron microscopy measurements). SOFI provided more accurate width measurements, though with somewhat lower spatial resolution than STORM. Super resolution imaging of intact cryo-cut human brain tissue is feasible. For quantitation, STORM can under-estimate diameters of thin fluorescent objects. SOFI is more robust. The greatest limitation for super-resolution imaging in brain sections is imposed by sample preparation. We anticipate that improved strategies to reduce autofluorescence and to enhance fluorophore performance will enable rapid expansion of this approach. © 2017 British Neuropathological Society.

A dimensionless ordered pull-through model of the mammalian lens epithelium evidences scaling across species and explains the age-dependent changes in cell density in the human lens

PubMed Central

Wu, Jun Jie; Wu, Weiju; Tholozan, Frederique M.; Saunter, Christopher D.; Girkin, John M.; Quinlan, Roy A.

2015-01-01

We present a mathematical (ordered pull-through; OPT) model of the cell-density profile for the mammalian lens epithelium together with new experimental data. The model is based upon dimensionless parameters, an important criterion for inter-species comparisons where lens sizes can vary greatly (e.g. bovine (approx. 18 mm); mouse (approx. 2 mm)) and confirms that mammalian lenses scale with size. The validated model includes two parameters: β/α, which is the ratio of the proliferation rate in the peripheral and in the central region of the lens; and γGZ, a dimensionless pull-through parameter that accounts for the cell transition and exit from the epithelium into the lens body. Best-fit values were determined for mouse, rat, rabbit, bovine and human lens epithelia. The OPT model accounts for the peak in cell density at the periphery of the lens epithelium, a region where cell proliferation is concentrated and reaches a maximum coincident with the germinative zone. The β/α ratio correlates with the measured FGF-2 gradient, a morphogen critical to lens cell survival, proliferation and differentiation. As proliferation declines with age, the OPT model predicted age-dependent changes in cell-density profiles, which we observed in mouse and human lenses. PMID:26236824

The Status, Quality, and Expansion of the NIH Full-Length cDNA Project: The Mammalian Gene Collection (MGC)

PubMed Central

2004-01-01

The National Institutes of Health's Mammalian Gene Collection (MGC) project was designed to generate and sequence a publicly accessible cDNA resource containing a complete open reading frame (ORF) for every human and mouse gene. The project initially used a random strategy to select clones from a large number of cDNA libraries from diverse tissues. Candidate clones were chosen based on 5′-EST sequences, and then fully sequenced to high accuracy and analyzed by algorithms developed for this project. Currently, more than 11,000 human and 10,000 mouse genes are represented in MGC by at least one clone with a full ORF. The random selection approach is now reaching a saturation point, and a transition to protocols targeted at the missing transcripts is now required to complete the mouse and human collections. Comparison of the sequence of the MGC clones to reference genome sequences reveals that most cDNA clones are of very high sequence quality, although it is likely that some cDNAs may carry missense variants as a consequence of experimental artifact, such as PCR, cloning, or reverse transcriptase errors. Recently, a rat cDNA component was added to the project, and ongoing frog (Xenopus) and zebrafish (Danio) cDNA projects were expanded to take advantage of the high-throughput MGC pipeline. PMID:15489334

Cloning and characterization of full-length mouse thymidine kinase 2: the N-terminal sequence directs import of the precursor protein into mitochondria.

PubMed Central

Wang, L; Eriksson, S

2000-01-01

The subcellular localization of mitochondrial thymidine kinase (TK2) has been questioned, since no mitochondrial targeting sequences have been found in cloned human TK2 cDNAs. Here we report the cloning of mouse TK2 cDNA from a mouse full-length enriched cDNA library. The mouse TK2 cDNA codes for a protein of 270 amino acids, with a 40-amino-acid presumed N-terminal mitochondrial targeting signal. In vitro translation and translocation experiments with purified rat mitochondria confirmed that the N-terminal sequence directed import of the precursor TK2 into the mitochondrial matrix. A single 2.4 kb mRNA transcript was detected in most tissues examined, except in liver, where an additional shorter (1.0 kb) transcript was also observed. There was no correlation between the tissue distribution of TK2 activity and the expression of TK2 mRNA. Full-length mouse TK2 protein and two N-terminally truncated forms, one of which corresponds to the mitochondrial form of TK2 and a shorter form corresponding to the previously characterized recombinant human TK2, were expressed in Escherichia coli and affinity purified. All three forms of TK2 phosphorylated thymidine, deoxycytidine and 2'-deoxyuridine, but with different kinetic efficiencies. A number of cytostatic pyrimidine nucleoside analogues were also tested and shown to be good substrates for the various forms of TK2. The active form of full-length mouse TK2 was a dimer, as judged by Superdex 200 chromatography. These results enhance our understanding of the structure and function of TK2, and may help to explain the mitochondrial disorder, mitochondrial neurogastrointestinal encephalomyopathy. PMID:11023833

Blocking antibodies induced by immunization with a hypoallergenic parvalbumin mutant reduce allergic symptoms in a mouse model of fish allergy

PubMed Central

Freidl, Raphaela; Gstoettner, Antonia; Baranyi, Ulrike; Swoboda, Ines; Stolz, Frank; Focke-Tejkl, Margarete; Wekerle, Thomas; van Ree, Ronald; Valenta, Rudolf; Linhart, Birgit

2017-01-01

Background Fish is a frequent elicitor of severe IgE-mediated allergic reactions. Beside avoidance, there is currently no allergen-specific therapy available. Hypoallergenic variants of the major fish allergen, parvalbumin, for specific immunotherapy based on mutation of the 2 calcium-binding sites have been developed. Objectives This study sought to establish a mouse model of fish allergy resembling human disease and to investigate whether mouse and rabbit IgG antibodies induced by immunization with a hypoallergenic mutant of the major carp allergen protect against allergic symptoms in sensitized mice. Methods C3H/HeJ mice were sensitized with recombinant wildtype Cyp c 1 or carp extract by intragastric gavage. Antibody, cellular immune responses, and epitope specificity in sensitized mice were investigated by ELISA, rat basophil leukemia assay, T-cell proliferation experiments using recombinant wildtype Cyp c 1, and overlapping peptides spanning the Cyp c 1 sequence. Anti-hypoallergenic Cyp c 1 mutant mouse and rabbit sera were tested for their ability to inhibit IgE recognition of Cyp c 1, Cyp c 1–specific basophil degranulation, and Cyp c 1–induced allergic symptoms in the mouse model. Results A mouse model of fish allergy mimicking human disease regarding IgE epitope recognition and symptoms as close as possible was established. Administration of antisera generated in mice and rabbits by immunization with a hypoallergenic Cyp c 1 mutant inhibited IgE binding to Cyp c 1, Cyp c 1–induced basophil degranulation, and allergic symptoms caused by allergen challenge in sensitized mice. Conclusions Antibodies induced by immunization with a hypoallergenic Cyp c 1 mutant protect against allergic reactions in a murine model of fish allergy. PMID:27876628

Blocking antibodies induced by immunization with a hypoallergenic parvalbumin mutant reduce allergic symptoms in a mouse model of fish allergy.

PubMed

Freidl, Raphaela; Gstoettner, Antonia; Baranyi, Ulrike; Swoboda, Ines; Stolz, Frank; Focke-Tejkl, Margarete; Wekerle, Thomas; van Ree, Ronald; Valenta, Rudolf; Linhart, Birgit

2017-06-01

Fish is a frequent elicitor of severe IgE-mediated allergic reactions. Beside avoidance, there is currently no allergen-specific therapy available. Hypoallergenic variants of the major fish allergen, parvalbumin, for specific immunotherapy based on mutation of the 2 calcium-binding sites have been developed. This study sought to establish a mouse model of fish allergy resembling human disease and to investigate whether mouse and rabbit IgG antibodies induced by immunization with a hypoallergenic mutant of the major carp allergen protect against allergic symptoms in sensitized mice. C3H/HeJ mice were sensitized with recombinant wildtype Cyp c 1 or carp extract by intragastric gavage. Antibody, cellular immune responses, and epitope specificity in sensitized mice were investigated by ELISA, rat basophil leukemia assay, T-cell proliferation experiments using recombinant wildtype Cyp c 1, and overlapping peptides spanning the Cyp c 1 sequence. Anti-hypoallergenic Cyp c 1 mutant mouse and rabbit sera were tested for their ability to inhibit IgE recognition of Cyp c 1, Cyp c 1-specific basophil degranulation, and Cyp c 1-induced allergic symptoms in the mouse model. A mouse model of fish allergy mimicking human disease regarding IgE epitope recognition and symptoms as close as possible was established. Administration of antisera generated in mice and rabbits by immunization with a hypoallergenic Cyp c 1 mutant inhibited IgE binding to Cyp c 1, Cyp c 1-induced basophil degranulation, and allergic symptoms caused by allergen challenge in sensitized mice. Antibodies induced by immunization with a hypoallergenic Cyp c 1 mutant protect against allergic reactions in a murine model of fish allergy. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Comparative toxicogenomic analysis of oral Cr(VI) exposure effects in rat and mouse small intestinal epithelia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kopec, Anna K.; Thompson, Chad M.; Kim, Suntae

2012-07-15

Continuous exposure to high concentrations of hexavalent chromium [Cr(VI)] in drinking water results in intestinal tumors in mice but not rats. Concentration-dependent gene expression effects were evaluated in female F344 rat duodenal and jejunal epithelia following 7 and 90 days of exposure to 0.3–520 mg/L (as sodium dichromate dihydrate, SDD) in drinking water. Whole-genome microarrays identified 3269 and 1815 duodenal, and 4557 and 1534 jejunal differentially expressed genes at 8 and 91 days, respectively, with significant overlaps between the intestinal segments. Functional annotation identified gene expression changes associated with oxidative stress, cell cycle, cell death, and immune response that weremore » consistent with reported changes in redox status and histopathology. Comparative analysis with B6C3F1 mouse data from a similarly designed study identified 2790 differentially expressed rat orthologs in the duodenum compared to 5013 mouse orthologs at day 8, and only 1504 rat and 3484 mouse orthologs at day 91. Automated dose–response modeling resulted in similar median EC{sub 50}s in the rodent duodenal and jejunal mucosae. Comparative examination of differentially expressed genes also identified divergently regulated orthologs. Comparable numbers of differentially expressed genes were observed at equivalent Cr concentrations (μg Cr/g duodenum). However, mice accumulated higher Cr levels than rats at ≥ 170 mg/L SDD, resulting in a ∼ 2-fold increase in the number of differentially expressed genes. These qualitative and quantitative differences in differential gene expression, which correlate with differences in tissue dose, likely contribute to the disparate intestinal tumor outcomes. -- Highlights: ► Cr(VI) elicits dose-dependent changes in gene expression in rat intestine. ► Cr(VI) elicits less differential gene expression in rats compared to mice. ► Cr(VI) gene expression can be phenotypically anchored to intestinal changes. ► Species-specific and divergent changes are consistent with species-specific tumors.« less

Characterization of the synthesis and expression of the GTA-kinase from transformed and normal rodent cells.

PubMed

Kerr, M; Fischer, J E; Purushotham, K R; Gao, D; Nakagawa, Y; Maeda, N; Ghanta, V; Hiramoto, R; Chegini, N; Humphreys-Beher, M G

1994-08-02

The murine transformed cell line YC-8 and beta-adrenergic receptor agonist (isoproternol) treated rat and mouse parotid gland acinar cells ectopically express cell surface beta 1-4 galactosyltransferase during active proliferation. This activity is dependent upon the expression of the GTA-kinase (p58) in these cells. Using total RNA, cDNA clones for the protein coding region of the kinase were isolated by reverse transcriptase-PCR cloning. DNA sequence analysis failed to show sequence differences with the normal homolog from mouse cells although Southern blot analysis of YC-8, and a second cell line KI81, indicated changes in the restriction enzyme digestion profile relative to murine cell lines which do not express cell surface galactosyltransferase. The rat cDNA clone from isoproterenol-treated salivary glands showed a high degree of protein and nucleic acid sequence homology to the GTA-kinase from both murine and human sources. Northern blot analysis of YC-8 and a control cell line LSTRA revealed the synthesis of a major 3.0 kb mRNA from both cell lines plus the unique expression of a 4.5 kb mRNA in the YC-8 cells. Reverse transcriptase-PCR of LSTRA and YC-8 confirmed the increased steady state levels of the GTA-kinase mRNA in YC-8. In the mouse, induction of cell proliferation by isoproterenol resulted in a 50-fold increase in steady state mRNA levels for the kinase over the low level of expression in quiescent cells. Expression of the rat 3' untranslated region in rat parotid cells in vitro led to an increased rate of DNA synthesis, cell number an ectopic expression of cell surface galactosyltransferase in the sense orientation. Antisense expression or vector alone did not alter growth characteristics of acinar cells. A polyclonal antibody monospecific to a murine amino terminal peptide sequence revealed a uniform distribution of GTA-kinase over the cytoplasm of acinar and duct cells of control mouse parotid glands. However, upon growth stimulation, kinase was detected primarily in a perinuclear and nuclear immunostaining pattern. Western blot analysis confirmed a translocation from a cytoplasmic localization in both LSTRA and quiescent salivary cells to a membrane-associated localization in YC-8 and proliferating salivary cells.

Palisade Endings Are a Constant Feature in the Extraocular Muscles of Frontal-Eyed, But Not Lateral-Eyed, Animals.

PubMed

Blumer, Roland; Maurer-Gesek, Barbara; Gesslbauer, Bernhard; Blumer, Michael; Pechriggl, Elisabeth; Davis-López de Carrizosa, María A; Horn, Anja K; May, Paul J; Streicher, Johannes; de la Cruz, Rosa R; Pastor, Ángel M

2016-02-01

To test whether palisade endings are a general feature of mammalian extraocular muscles (EOMs). Thirteen species, some frontal-eyed (human, monkey, cat, and ferret), and others lateral-eyed (pig, sheep, calf, horse, rabbit, rat, mouse, gerbil, and guinea pig) were analyzed. Palisade endings were labeled by using different combinations of immunofluorescence techniques. Three-dimensional reconstructions of immunolabeled palisade endings were done. In all frontal-eyed species, palisade endings were a consistent feature in the rectus EOMs. Their total number was high and they exhibited an EOM-specific distribution. In particular, the number of palisade endings in the medial recti was significantly higher than in the other rectus muscles. In the lateral-eyed animals, palisade endings were infrequent and, when present, their total number was rather low. They were only found in ungulates (sheep, calf, pig, and horse) and in rabbit. In rodents (rat, guinea pig, mouse, and gerbil) palisade endings were found infrequently (e.g., rat) or were completely absent. Palisade endings in frontal-eyed species and in some lateral-eyed species (pig, sheep, calf, and horse) had a uniform morphology. They generally lacked α-bungarotoxin staining, with a few exceptions in primates. Palisade endings in other lateral-eyed species (rabbit and rat) exhibited a simplified morphology and bound α-bungarotoxin. Palisade endings are not a universal feature of mammalian EOMs. So, if they are proprioceptors, not all species require them. Because in frontal-eyed species, the medial rectus muscle has the highest number of palisade endings, they likely play a special role in convergence.

Palisade Endings Are a Constant Feature in the Extraocular Muscles of Frontal-Eyed, But Not Lateral-Eyed, Animals

PubMed Central

Blumer, Roland; Maurer-Gesek, Barbara; Gesslbauer, Bernhard; Blumer, Michael; Pechriggl, Elisabeth; Davis-López de Carrizosa, María A.; Horn, Anja K.; May, Paul J.; Streicher, Johannes; de la Cruz, Rosa R.; Pastor, Ángel M.

2016-01-01

Purpose To test whether palisade endings are a general feature of mammalian extraocular muscles (EOMs). Methods Thirteen species, some frontal-eyed (human, monkey, cat, and ferret), and others lateral-eyed (pig, sheep, calf, horse, rabbit, rat, mouse, gerbil, and guinea pig) were analyzed. Palisade endings were labeled by using different combinations of immunofluorescence techniques. Three-dimensional reconstructions of immunolabeled palisade endings were done. Results In all frontal-eyed species, palisade endings were a consistent feature in the rectus EOMs. Their total number was high and they exhibited an EOM-specific distribution. In particular, the number of palisade endings in the medial recti was significantly higher than in the other rectus muscles. In the lateral-eyed animals, palisade endings were infrequent and, when present, their total number was rather low. They were only found in ungulates (sheep, calf, pig, and horse) and in rabbit. In rodents (rat, guinea pig, mouse, and gerbil) palisade endings were found infrequently (e.g., rat) or were completely absent. Palisade endings in frontal-eyed species and in some lateral-eyed species (pig, sheep, calf, and horse) had a uniform morphology. They generally lacked α-bungarotoxin staining, with a few exceptions in primates. Palisade endings in other lateral-eyed species (rabbit and rat) exhibited a simplified morphology and bound α-bungarotoxin. Conclusions Palisade endings are not a universal feature of mammalian EOMs. So, if they are proprioceptors, not all species require them. Because in frontal-eyed species, the medial rectus muscle has the highest number of palisade endings, they likely play a special role in convergence. PMID:26830369

Interspecies differences in metabolism of arsenic by cultured primary hepatocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Drobna, Zuzana; Walton, Felecia S.; Harmon, Anne W.

2010-05-15

Biomethylation is the major pathway for the metabolism of inorganic arsenic (iAs) in many mammalian species, including the human. However, significant interspecies differences have been reported in the rate of in vivo metabolism of iAs and in yields of iAs metabolites found in urine. Liver is considered the primary site for the methylation of iAs and arsenic (+3 oxidation state) methyltransferase (As3mt) is the key enzyme in this pathway. Thus, the As3mt-catalyzed methylation of iAs in the liver determines in part the rate and the pattern of iAs metabolism in various species. We examined kinetics and concentration-response patterns for iAsmore » methylation by cultured primary hepatocytes derived from human, rat, mice, dog, rabbit, and rhesus monkey. Hepatocytes were exposed to [{sup 73}As]arsenite (iAs{sup III}; 0.3, 0.9, 3.0, 9.0 or 30 nmol As/mg protein) for 24 h and radiolabeled metabolites were analyzed in cells and culture media. Hepatocytes from all six species methylated iAs{sup III} to methylarsenic (MAs) and dimethylarsenic (DMAs). Notably, dog, rat and monkey hepatocytes were considerably more efficient methylators of iAs{sup III} than mouse, rabbit or human hepatocytes. The low efficiency of mouse, rabbit and human hepatocytes to methylate iAs{sup III} was associated with inhibition of DMAs production by moderate concentrations of iAs{sup III} and with retention of iAs and MAs in cells. No significant correlations were found between the rate of iAs methylation and the thioredoxin reductase activity or glutathione concentration, two factors that modulate the activity of recombinant As3mt. No associations between the rates of iAs methylation and As3mt protein structures were found for the six species examined. Immunoblot analyses indicate that the superior arsenic methylation capacities of dog, rat and monkey hepatocytes examined in this study may be associated with a higher As3mt expression. However, factors other than As3mt expression may also contribute to the interspecies differences in the hepatocyte capacity to methylate iAs.« less

Mechanism for Clastogenic Activity of Naphthalene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buchholz, Bruce A.

2016-06-24

Naphthalene incubations form DNA adducts in vitro in a dose dependent manner in both mouse and rat tissues. Rodent tissue incubations with naphthalene indicate that naphthalene forms as many DNA adducts as Benzo(a)pyrene, a known DNA binding carcinogen. The mouse airway has the greatest number of DNA adducts, corresponding to the higher metabolic activation of naphthalene in this location. Both rat tissues, the rat olfactory (tumor target) and the airways (non-tumor target), have similar levels of NA-DNA adducts, indicating that short term measures of initial adduct formation do not directly correlate with sites of tumor formation in the NTP bioassays.

The final stage of cholinergic differentiation occurs below inner hair cells during development of the rodent cochlea.

PubMed

Bergeron, Adam L; Schrader, Angela; Yang, Dan; Osman, Abdullah A; Simmons, Dwayne D

2005-12-01

To gain further insights into the cholinergic differentiation of presynaptic efferent terminals in the inner ear, we investigated the expression of the high-affinity choline transporter (ChT1) in comparison to other presynaptic and cholinergic markers. In the adult mammalian cochlea, cholinergic axons from medial olivocochlear (OC) neurons form axosomatic synapses with outer hair cells (OHCs), whereas axons from lateral OC neurons form axodendritic synapses on afferent fibers below inner hair cells (IHCs). Mouse brain and cochlea homogenates reveal at least two ChT1 isoforms: a nonglycosylated approximately 73 kDa protein and a glycosylated approximately 45 kDa protein. In mouse brain, ChT1 is preferentially expressed by neurons in periolivary regions of the superior olive consistent with the location of medial OC neurons. In the adult mouse cochlea, ChT1-positive terminals are located almost exclusively below OHCs consistent with a medial OC innervation. Between postnatal day 2 (P2) and P4, ChT1-positive terminals are below IHCs and occur after the expression of growth-associated protein 43, synapsin, and the vesicular acetylcholine transporter. By P15, ChT1-positive terminals are mostly on OHCs. Accounting for differences in gestational age, the developmental expression of ChT1 in the rat cochlea is similar to the mouse. However, in older rats ChT1-positive terminals are below IHCs and OHCs. In both rat and mouse, our observations indicate that the onset of ChT1 expression occurs after efferent terminals are below IHCs and express other presynaptic and cholinergic markers. In the mouse, but not in the rat, ChT1 may preferentially identify medial OC neurons.

Expression of Receptors for Tetanus Toxin and Monoclonal Antibody A2B5 by Pancreatic Islet Cells

NASA Astrophysics Data System (ADS)

Eisenbarth, G. S.; Shimizu, K.; Bowring, M. A.; Wells, S.

1982-08-01

Studies of the reaction of antibody A2B5 and tetanus toxin with pancreatic islet cells, islet cell tumors, and other human amine precursor uptake and decarboxylation (APUD) tumors are described. By indirect immunofluorescence, antibody A2B5 and tetanus toxin were shown to specifically bind to the plasma membrane of human, rat, chicken, and mouse islet cells. The binding of antibody A2B5 to the cell surface of living islet cells has allowed isolation of these cells from a suspension of pancreatic cells by using a fluorescence-activated cell sorter. In studies designed to determine whether tetanus toxin and antibody A2B5 bound to the same surface antigen, A2B5 and tetanus toxin did not compete for binding to normal islet cells, a human islet cell tumor, or a rat islet cell tumor. In addition to binding to islet cell tumors, antibody A2B5 reacts with frozen sections, isolated cells, and cell lines of neural, neural crest, and APUD origin.

A Reverse-Translational Approach to Bipolar Disorder: Rodent and human studies in the Behavioral Pattern Monitor

PubMed Central

Young, Jared W.; Minassian, Arpi; Paulus, Martin P.; Geyer, Mark A.; Perry, William

2007-01-01

Mania is the defining feature of Bipolar Disorder (BD). There has been limited progress in understanding the neurobiological underpinnings of BD mania and developing novel therapeutics, in part due to a paucity of relevant animal models with translational potential. Hyperactivity is a cardinal symptom of mania, traditionally measured in humans using observer-rated scales. Multivariate assessment of unconditioned locomotor behavior using the rat Behavioral Pattern Monitor (BPM) developed in our laboratory has shown that hyperactivity includes complex multifaceted behaviors. The BPM has been used to demonstrate differential effects of drugs on locomotor activity and exploratory behavior in rats. Studies of genetically engineered mice in a mouse BPM have confirmed its utility as a cross-species tool. In a “reverse-translational” approach to this work, we developed the human BPM to characterize motor activity in BD patients. Increased activity, object interactions, and altered locomotor patterns provide multidimensional phenotypes to model in the rodent BPM. This unique approach to modeling BD provides an opportunity to identify the neurobiology underlying BD mania and test novel antimanic agents. PMID:17706782

Livestock in biomedical research: history, current status and future prospective.

PubMed

Polejaeva, Irina A; Rutigliano, Heloisa M; Wells, Kevin D

2016-01-01

Livestock models have contributed significantly to biomedical and surgical advances. Their contribution is particularly prominent in the areas of physiology and assisted reproductive technologies, including understanding developmental processes and disorders, from ancient to modern times. Over the past 25 years, biomedical research that traditionally embraced a diverse species approach shifted to a small number of model species (e.g. mice and rats). The initial reasons for focusing the main efforts on the mouse were the availability of murine embryonic stem cells (ESCs) and genome sequence data. This powerful combination allowed for precise manipulation of the mouse genome (knockouts, knockins, transcriptional switches etc.) leading to ground-breaking discoveries on gene functions and regulation, and their role in health and disease. Despite the enormous contribution to biomedical research, mouse models have some major limitations. Their substantial differences compared with humans in body and organ size, lifespan and inbreeding result in pronounced metabolic, physiological and behavioural differences. Comparative studies of strategically chosen domestic species can complement mouse research and yield more rigorous findings. Because genome sequence and gene manipulation tools are now available for farm animals (cattle, pigs, sheep and goats), a larger number of livestock genetically engineered (GE) models will be accessible for biomedical research. This paper discusses the use of cattle, goats, sheep and pigs in biomedical research, provides an overview of transgenic technology in farm animals and highlights some of the beneficial characteristics of large animal models of human disease compared with the mouse. In addition, status and origin of current regulation of GE biomedical models is also reviewed.

Gut Immune Maturation Depends on Colonization with a Host-Specific Microbiota

PubMed Central

Chung, Hachung; Pamp, Sünje J.; Hill, Jonathan A.; Surana, Neeraj K.; Edelman, Sanna M.; Troy, Erin B.; Reading, Nicola C.; Villablanca, Eduardo J.; Wang, Sen; Mora, Jorge R.; Umesaki, Yoshinori; Mathis, Diane; Benoist, Christophe; Relman, David A.; Kasper, Dennis L.

2012-01-01

SUMMARY Gut microbial induction of host immune maturation exemplifies host-microbe mutualism. We colonized germ-free (GF) mice with mouse microbiota (MMb) or human microbiota (HMb) to determine whether small intestinal immune maturation depends on a coevolved host-specific microbiota. Gut bacterial numbers and phylum abundance were similar in MMb and HMb mice, but bacterial species differed, especially the Firmicutes. HMb mouse intestines had low levels of CD4+ and CD8+ T cells, few proliferating T cells, few dendritic cells, and low antimicrobial peptide expression–all characteristics of GF mice. Rat microbiota also failed to fully expand intestinal T cell numbers in mice. Colonizing GF or HMb mice with mouse-segmented filamentous bacteria (SFB) partially restored T cell numbers, suggesting that SFB and other MMb organisms are required for full immune maturation in mice. Importantly, MMb conferred better protection against Salmonella infection than HMb. A host-specific microbiota appears to be critical for a healthy immune system. PMID:22726443

A monoclonal IgM smooth muscle antibody reactive with fibroblast stress fibres produced by immunization with Treponema pallidum.

PubMed Central

Strugnell, R A; Underwood, J R; Clarke, F M; Pedersen, J S; Chalmers, P J; Faine, S; Toh, B H

1983-01-01

A monoclonal IgM smooth muscle antibody secreted by a hybrid (MMI-1) of mouse plasmacytoma NS-1 with spleen cells from mouse immunized with Treponema pallidum was detected by indirect immunofluorescence tests on frozen tissue sections and on acetone fixed monolayers of rat and human fibroblasts. The antibody did not react with acetone fixed smears of T. pallidum but reacted with smooth muscle fibres and with striations of skeletal and cardiac muscle. In non-muscle cells, the antibody stained liver in a 'polygonal' pattern, thymus with accentuated staining of the thymic medulla, renal glomeruli and the brush border and peritubular fibrils of renal tubules. In fibroblast monolayers, the antibody stained stress fibres in an interrupted pattern. Immunoblotting with muscle proteins and the antibody showed labelling of a 100K molecule. The cellular distribution of the mouse monoclonal antibody is similar to that obtained with anti-actin antibody suggesting that the corresponding antigen may be an actin binding protein. Images Fig. 3 PMID:6347470

Differentiation of the SH-SY5Y Human Neuroblastoma Cell Line

PubMed Central

Shipley, Mackenzie M.; Mangold, Colleen A.; Szpara, Moriah L.

2016-01-01

Having appropriate in vivo and in vitro systems that provide translational models for human disease is an integral aspect of research in neurobiology and the neurosciences. Traditional in vitro experimental models used in neurobiology include primary neuronal cultures from rats and mice, neuroblastoma cell lines including rat B35 and mouse Neuro-2A cells, rat PC12 cells, and short-term slice cultures. While many researchers rely on these models, they lack a human component and observed experimental effects could be exclusive to the respective species and may not occur identically in humans. Additionally, although these cells are neurons, they may have unstable karyotypes, making their use problematic for studies of gene expression and reproducible studies of cell signaling. It is therefore important to develop more consistent models of human neurological disease. The following procedure describes an easy-to-follow, reproducible method to obtain homogenous and viable human neuronal cultures, by differentiating the chromosomally stable human neuroblastoma cell line, SH-SY5Y. This method integrates several previously described methods1-4 and is based on sequential removal of serum from media. The timeline includes gradual serum-starvation, with introduction of extracellular matrix proteins and neurotrophic factors. This allows neurons to differentiate, while epithelial cells are selected against, resulting in a homogeneous neuronal culture. Representative results demonstrate the successful differentiation of SH-SY5Y neuroblastoma cells from an initial epithelial-like cell phenotype into a more expansive and branched neuronal phenotype. This protocol offers a reliable way to generate homogeneous populations of neuronal cultures that can be used for subsequent biochemical and molecular analyses, which provides researchers with a more accurate translational model of human infection and disease. PMID:26967710

Differentiation of the SH-SY5Y Human Neuroblastoma Cell Line.

PubMed

Shipley, Mackenzie M; Mangold, Colleen A; Szpara, Moriah L

2016-02-17

Having appropriate in vivo and in vitro systems that provide translational models for human disease is an integral aspect of research in neurobiology and the neurosciences. Traditional in vitro experimental models used in neurobiology include primary neuronal cultures from rats and mice, neuroblastoma cell lines including rat B35 and mouse Neuro-2A cells, rat PC12 cells, and short-term slice cultures. While many researchers rely on these models, they lack a human component and observed experimental effects could be exclusive to the respective species and may not occur identically in humans. Additionally, although these cells are neurons, they may have unstable karyotypes, making their use problematic for studies of gene expression and reproducible studies of cell signaling. It is therefore important to develop more consistent models of human neurological disease. The following procedure describes an easy-to-follow, reproducible method to obtain homogenous and viable human neuronal cultures, by differentiating the chromosomally stable human neuroblastoma cell line, SH-SY5Y. This method integrates several previously described methods(1-4) and is based on sequential removal of serum from media. The timeline includes gradual serum-starvation, with introduction of extracellular matrix proteins and neurotrophic factors. This allows neurons to differentiate, while epithelial cells are selected against, resulting in a homogeneous neuronal culture. Representative results demonstrate the successful differentiation of SH-SY5Y neuroblastoma cells from an initial epithelial-like cell phenotype into a more expansive and branched neuronal phenotype. This protocol offers a reliable way to generate homogeneous populations of neuronal cultures that can be used for subsequent biochemical and molecular analyses, which provides researchers with a more accurate translational model of human infection and disease.

The human phospholamban gene: structure and expression.

PubMed

McTiernan, C F; Frye, C S; Lemster, B H; Kinder, E A; Ogletree-Hughes, M L; Moravec, C S; Feldman, A M

1999-03-01

Phospholamban, through modulation of sarcoplasmic reticulum calcium-ATPase activity, is a key regulator of cardiac diastolic function. Alterations in phospholamban expression may define parameters of muscle relaxation. In experimental animals, phospholamban is differentially expressed in various striated and smooth muscles, and within the four chambers of the heart. Decreased phospholamban expression within the heart during heart failure has also been observed. Furthermore, regulatory elements of mammalian phospholamban genes remain poorly defined. To extend these studies to humans, we (1) characterized phospholamban expression in various human organs, (2) isolated genomic clones encoding the human phospholamban gene, and (3) prepared human phospholamban promoter/luciferase reporter constructs and performed transient transfection assays to begin identification of regulatory elements. We observed that human ventricle and quadriceps displayed high levels of phospholamban transcripts and proteins, with markedly lower expression observed in smooth muscles, while the right atria also expressed low levels of phospholamban. The human phospholamban gene structure closely resembles that reported for chicken, rabbit, rat, and mouse. Comparison of the human to other mammalian phospholamban genes indicates a marked conservation of sequence for at least 217 bp upstream of the transcription start site, which contains conserved motifs for GATA, CP1/NFY, M-CAT-like, and E-box elements. Transient transfection assays with a series of plasmids containing deleted 5' flanking regions (between -2530 and -66 through +85) showed that sequences between -169 and the CP1-box at -93 were required for maximal promoter activity in neonatal rat cardiomyocytes. Activity of these reporters in HeLa cells was markedly lower than that observed in rat cardiomyocytes, suggesting at least a partial tissue selectivity of these reporter constructs.

Fatty acid binding protein-1 (FABP1) and the human FABP1 T94A Variant: Roles in the Endocannabinoid System and Dyslipidemias

PubMed Central

Schroeder, Friedhelm; McIntosh, Avery L.; Martin, Gregory G.; Huang, Huan; Landrock, Danilo; Chung, Sarah; Landrock, Kerstin K.; Dangott, Lawrence J.; Li, Shengrong; Kaczocha, Martin; Murphy, Eric J.; Atshaves, Barbara P.; Kier, Ann B.

2017-01-01

The first discovered member of the mammalian FABP family, liver fatty acid binding protein (FABP1, L-FABP), occurs at high cytosolic concentration in liver, intestine and in the case of humans also in kidney. While the rat FABP1 is well studied, the extent these findings translate to human FABP1 is not clear—especially in view of recent studies showing that endocannabinoids and cannabinoids represent novel rat FABP1 ligands and FABP1 gene ablation impacts the hepatic endocannabinoid system, known to be involved in non-alcoholic fatty liver (NAFLD) development. Although not detectable in brain, FABP1 ablation nevertheless also impacts brain endocannabinoids. Despite overall tertiary structure similarity, human FABP1 differs significantly from rat FABP1 in secondary structure, much larger ligand binding cavity, and affinities/specificities for some ligands. Moreover, while both mouse and human FABP1 mediate ligand induction of peroxisome proliferator activated receptor-α, (PPARα), they differ markedly in pattern of genes induced. This is critically important because a highly prevalent human SNP (26–38% minor allele frequency and 8.3±1.9% homozygous) results in a FABP1 T94A substitution that further accentuates these species differences. The human FABP1 T94A variant is associated with altered body mass index (BMI), clinical dyslipidemias (elevated plasma triglycerides and LDL cholesterol), atherothrombotic cerebral infarction, and non-alcoholic fatty liver disease (NAFLD). Resolving human FABP1 and the T94A variant’s impact on the endocannabinoid and cannabinoid system is an exciting challenge due to the importance of this system on hepatic lipid accumulation as well as behavior, pain, inflammation, and satiety. PMID:27117865

Fatty Acid Binding Protein-1 (FABP1) and the Human FABP1 T94A Variant: Roles in the Endocannabinoid System and Dyslipidemias.

PubMed

Schroeder, Friedhelm; McIntosh, Avery L; Martin, Gregory G; Huang, Huan; Landrock, Danilo; Chung, Sarah; Landrock, Kerstin K; Dangott, Lawrence J; Li, Shengrong; Kaczocha, Martin; Murphy, Eric J; Atshaves, Barbara P; Kier, Ann B

2016-06-01

The first discovered member of the mammalian FABP family, liver fatty acid binding protein (FABP1, L-FABP), occurs at high cytosolic concentration in liver, intestine, and in the case of humans also in kidney. While the rat FABP1 is well studied, the extent these findings translate to human FABP1 is not clear-especially in view of recent studies showing that endocannabinoids and cannabinoids represent novel rat FABP1 ligands and FABP1 gene ablation impacts the hepatic endocannabinoid system, known to be involved in non-alcoholic fatty liver (NAFLD) development. Although not detectable in brain, FABP1 ablation nevertheless also impacts brain endocannabinoids. Despite overall tertiary structure similarity, human FABP1 differs significantly from rat FABP1 in secondary structure, much larger ligand binding cavity, and affinities/specificities for some ligands. Moreover, while both mouse and human FABP1 mediate ligand induction of peroxisome proliferator activated receptor-α (PPARα), they differ markedly in pattern of genes induced. This is critically important because a highly prevalent human single nucleotide polymorphism (SNP) (26-38 % minor allele frequency and 8.3 ± 1.9 % homozygous) results in a FABP1 T94A substitution that further accentuates these species differences. The human FABP1 T94A variant is associated with altered body mass index (BMI), clinical dyslipidemias (elevated plasma triglycerides and LDL cholesterol), atherothrombotic cerebral infarction, and non-alcoholic fatty liver disease (NAFLD). Resolving human FABP1 and the T94A variant's impact on the endocannabinoid and cannabinoid system is an exciting challenge due to the importance of this system in hepatic lipid accumulation as well as behavior, pain, inflammation, and satiety.

The Guinea Pig as a Model for Sporadic Alzheimer’s Disease (AD): The Impact of Cholesterol Intake on Expression of AD-Related Genes

PubMed Central

Ong, Daniel; Wijaya, Linda; Laws, Simon M.; Taddei, Kevin; Newman, Morgan; Lardelli, Michael; Martins, Ralph N.; Verdile, Giuseppe

2013-01-01

We investigated the guinea pig, Cavia porcellus, as a model for Alzheimer’s disease (AD), both in terms of the conservation of genes involved in AD and the regulatory responses of these to a known AD risk factor - high cholesterol intake. Unlike rats and mice, guinea pigs possess an Aβ peptide sequence identical to human Aβ. Consistent with the commonality between cardiovascular and AD risk factors in humans, we saw that a high cholesterol diet leads to up-regulation of BACE1 (β-secretase) transcription and down-regulation of ADAM10 (α-secretase) transcription which should increase release of Aβ from APP. Significantly, guinea pigs possess isoforms of AD-related genes found in humans but not present in mice or rats. For example, we discovered that the truncated PS2V isoform of human PSEN2, that is found at raised levels in AD brains and that increases γ-secretase activity and Aβ synthesis, is not uniquely human or aberrant as previously believed. We show that PS2V formation is up-regulated by hypoxia and a high-cholesterol diet while, consistent with observations in humans, Aβ concentrations are raised in some brain regions but not others. Also like humans, but unlike mice, the guinea pig gene encoding tau, MAPT, encodes isoforms with both three and four microtubule binding domains, and cholesterol alters the ratio of these isoforms. We conclude that AD-related genes are highly conserved and more similar to human than the rat or mouse. Guinea pigs represent a superior rodent model for analysis of the impact of dietary factors such as cholesterol on the regulation of AD-related genes. PMID:23805206

Acetaminophen-induced liver injury in rats and mice: Comparison of protein adducts, mitochondrial dysfunction, and oxidative stress in the mechanism of toxicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

McGill, Mitchell R.; Williams, C. David; Xie, Yuchao

2012-11-01

Acetaminophen (APAP) overdose is the most common cause of acute liver failure in the West. In mice, APAP hepatotoxicity can be rapidly induced with a single dose. Because it is both clinically relevant and experimentally convenient, APAP intoxication has become a popular model of liver injury. Early data demonstrated that rats are resistant to APAP toxicity. As a result, mice are the preferred species for mechanistic studies. Furthermore, recent work has shown that the mechanisms of APAP toxicity in humans are similar to mice. Nevertheless, some investigators still use rats. New mechanistic information from the last forty years invites amore » reevaluation of the differences between these species. Comparison may provide interesting insights and confirm or exclude the rat as an option for APAP studies. To this end, we treated rats and mice with APAP and measured parameters of liver injury, APAP metabolism, oxidative stress, and activation of the c-Jun N-terminal kinase (JNK). Consistent with earlier data, we found that rats were highly resistant to APAP toxicity. Although overall APAP metabolism was similar in both species, mitochondrial protein adducts were significantly lower in rats. Accordingly, rats also had less oxidative stress. Finally, while mice showed extensive activation and mitochondrial translocation of JNK, this could not be detected in rat livers. These data support the hypothesis that mitochondrial dysfunction is critical for the development of necrosis after APAP treatment. Because mitochondrial damage also occurs in humans, rats are not a clinically relevant species for studies of APAP hepatotoxicity. Highlights: ► Acetaminophen overdose causes severe liver injury only in mice but not in rats. ► APAP causes hepatic GSH depletion and protein adduct formation in rats and mice. ► Less protein adducts were measured in rat liver mitochondria compared to mouse. ► No oxidant stress, peroxynitrite formation or JNK activation was present in rats. ► The limited mitochondrial adducts in rats are insufficient to trigger cell necrosis.« less

Muscularity as a function of species, sex and age in small mammals

NASA Technical Reports Server (NTRS)

Pace, N.; Rahlmann, D. F.; Smith, A. H.

1984-01-01

Changes in the body skeletal muscle mass SMM (measured as a function of the ratio between the body creatine mass and the fat-free muscle creatine), and in muscularity (expressed as the ratio of SMM to fat-free body mass) were studied as functions of age, sex, and species in mouse, rat, hamster, guinea pig, and rabbit. Six animals of each sex were examined in eight age cohorts ranging from 1 to 24 months. Both species and age factors affect SMM. Strong sexual dimorphism in the SMM changes with age was displayed by mouse, rat, and guinea pig, whereas the hamster and rabbit were statistically monomorphic. The mouse, rat, and hamster attain a maximal SMM at about 1 year of age, whereas in the guinea pig and rabbit the decrease in SMM starts after 2 years. The value of muscularity reached a peak at age of 2-3 months in all animals of both sexes, with a pronounced difference among the species. The mouse emerged as the most muscular, while the guinea pig the least muscular, of all species.

MOUSE LIVER TUMOR DATA: ASSESSMENT OF CARCINOGENIC ACTIVITY

EPA Science Inventory

A significant number of chemicals have been shown to be carcinogenic in mouse liver while lacking carcinogenic activity in other organs or tissues of mice or rats. The review focus on the reasons for the unique susceptibility of the mouse liver to these carcinogens, and the exten...

Nevirapine bioactivation and covalent binding in the skin.

PubMed

Sharma, Amy M; Klarskov, Klaus; Uetrecht, Jack

2013-03-18

Nevirapine (NVP) treatment is associated with serious skin rashes that appear to be immune-mediated. We previously developed a rat model of this skin rash that is immune-mediated and is very similar to the rash in humans. Treatment of rats with the major NVP metabolite, 12-OH-NVP, also caused the rash. Most idiosyncratic drug reactions are caused by reactive metabolites; 12-OH-NVP forms a benzylic sulfate, which was detected in the blood of animals treated with NVP or 12-OH-NVP. This sulfate is presumably formed in the liver; however, the skin also has significant sulfotransferase activity. In this study, we used a serum against NVP to detect covalent binding in the skin of rats. There was a large artifact band in immunoblots of whole skin homogenates that interfered with detection of covalent binding; however, when the skin was separated into dermal and epidermal fractions, covalent binding was clearly present in the epidermis, which is also the location of sulfotransferases. In contrast to rats, treatment of mice with NVP did not result in covalent binding in the skin or skin rash. Although the reaction of 12-OH-NVP sulfate with nucleophiles such as glutathione is slow, incubation of this sulfate with homogenized human and rat skin led to extensive covalent binding. Incubations of 12-OH-NVP with the soluble fraction from a 9,000g centrifugation (S9) of rat or human skin homogenate in the presence of 3'-phosphoadenosine-5'-phosphosulfate (PAPS) produced extensive covalent binding, but no covalent binding was detected with mouse skin S9, which suggests that the reason mice do not develop a rash is that they lack the required sulfotransferase. This is the first study to report covalent binding of NVP to rat and human skin. These data provide strong evidence that covalent binding of NVP in the skin is due to 12-OH-NVP sulfate, which is likely responsible for NVP-induced skin rash. Sulfation may represent a bioactivation pathway for other drugs that cause a skin rash.

Hyperoxia reduces insulin release and induces mitochondrial dysfunction with possible implications for hyperoxic treatment of neonates.

PubMed

Hals, Ingrid; Ohki, Tsuyoshi; Singh, Rinku; Ma, Zuheng; Björklund, Anneli; Balasuriya, Chandima; Scholz, Hanne; Grill, Valdemar

2017-10-01

We previously showed that hyperoxia in vitro negatively affects beta cells of the rat. Here, we tested for possible clinical significance as well as mitochondrial interactions by hyperoxia, using human islets (function and viability), INS-1 832/13 cells (mitochondrial metabolism), and mouse neonates (effects in vivo). Lastly, we assessed relevant parameters in a cohort of individuals born preterm and then exposed to hyperoxia. Human islets and INS-1 832/13 cells were exposed to 24 h of hyperoxia (90-92% oxygen). Mouse neonates were subjected to 5 days of continuous hyperoxia. Individuals born preterm were evaluated in terms of glucose homeostasis and beta cell function by HbA1c and the HOMA2 formula. In human islets, hyperoxia significantly reduced glucose-stimulated insulin secretion by 42.2 ± 5.3% and viability assessed by MTT by 22.5 ± 5.4%. Hyperoxia down-regulated mitochondrial complex II by 21 ± 5% and upregulated complex III by 26 ± 10.1% and complex IV by 37 ± 10.6%. Partly similar effects on mitochondrial complexes were found in hyperoxia-exposed INS-1 832/13 cells. Exposure to hyperoxia swiftly reduced oxygen consumption in these cells and increased mitochondrial uncoupling. Hyperoxia transiently but significantly reduced insulin release in mouse neonates. Individuals born preterm displayed higher HbA1c versus controls, as well as insulin resistance. Thus, hyperoxia exerts negative effects in vitro on human beta cells and results indicate inhibitory effects on insulin secretion in vivo in mouse neonates. Negative effects may be lessened by the demonstrated swift and profound mitochondrial adaptability. Our findings open the possibility that hyperoxia could negatively affect beta cells of preterm human neonates. © 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

The endogenous and reactive depression subtypes revisited: integrative animal and human studies implicate multiple distinct molecular mechanisms underlying major depressive disorder.

PubMed

Malki, Karim; Keers, Robert; Tosto, Maria Grazia; Lourdusamy, Anbarasu; Carboni, Lucia; Domenici, Enrico; Uher, Rudolf; McGuffin, Peter; Schalkwyk, Leonard C

2014-05-07

Traditional diagnoses of major depressive disorder (MDD) suggested that the presence or absence of stress prior to onset results in either 'reactive' or 'endogenous' subtypes of the disorder, respectively. Several lines of research suggest that the biological underpinnings of 'reactive' or 'endogenous' subtypes may also differ, resulting in differential response to treatment. We investigated this hypothesis by comparing the gene-expression profiles of three animal models of 'reactive' and 'endogenous' depression. We then translated these findings to clinical samples using a human post-mortem mRNA study. Affymetrix mouse whole-genome oligonucleotide arrays were used to measure gene expression from hippocampal tissues of 144 mice from the Genome-based Therapeutic Drugs for Depression (GENDEP) project. The study used four inbred mouse strains and two depressogenic 'stress' protocols (maternal separation and Unpredictable Chronic Mild Stress) to model 'reactive' depression. Stress-related mRNA differences in mouse were compared with a parallel mRNA study using Flinders Sensitive and Resistant rat lines as a model of 'endogenous' depression. Convergent genes differentially expressed across the animal studies were used to inform candidate gene selection in a human mRNA post-mortem case control study from the Stanley Brain Consortium. In the mouse 'reactive' model, the expression of 350 genes changed in response to early stresses and 370 in response to late stresses. A minimal genetic overlap (less than 8.8%) was detected in response to both stress protocols, but 30% of these genes (21) were also differentially regulated in the 'endogenous' rat study. This overlap is significantly greater than expected by chance. The VAMP-2 gene, differentially expressed across the rodent studies, was also significantly altered in the human study after correcting for multiple testing. Our results suggest that 'endogenous' and 'reactive' subtypes of depression are associated with largely distinct changes in gene-expression. However, they also suggest that the molecular signature of 'reactive' depression caused by early stressors differs considerably from that of 'reactive' depression caused by late stressors. A small set of genes was consistently dysregulated across each paradigm and in post-mortem brain tissue of depressed patients suggesting a final common pathway to the disorder. These genes included the VAMP-2 gene, which has previously been associated with Axis-I disorders including MDD, bipolar depression, schizophrenia and with antidepressant treatment response. We also discuss the implications of our findings for disease classification, personalized medicine and case-control studies of MDD.

From engineering to editing the rat genome.

PubMed

Meek, Stephen; Mashimo, Tomoji; Burdon, Tom

2017-08-01

Since its domestication over 100 years ago, the laboratory rat has been the preferred experimental animal in many areas of biomedical research (Lindsey and Baker The laboratory rat. Academic, New York, pp 1-52, 2006). Its physiology, size, genetics, reproductive cycle, cognitive and behavioural characteristics have made it a particularly useful animal model for studying many human disorders and diseases. Indeed, through selective breeding programmes numerous strains have been derived that are now the mainstay of research on hypertension, obesity and neurobiology (Okamoto and Aoki Jpn Circ J 27:282-293, 1963; Zucker and Zucker J Hered 52(6):275-278, 1961). Despite this wealth of genetic and phenotypic diversity, the ability to manipulate and interrogate the genetic basis of existing phenotypes in rat strains and the methodology to generate new rat models has lagged significantly behind the advances made with its close cousin, the laboratory mouse. However, recent technical developments in stem cell biology and genetic engineering have again brought the rat to the forefront of biomedical studies and enabled researchers to exploit the increasingly accessible wealth of genome sequence information. In this review, we will describe how a breakthrough in understanding the molecular basis of self-renewal of the pluripotent founder cells of the mammalian embryo, embryonic stem (ES) cells, enabled the derivation of rat ES cells and their application in transgenesis. We will also describe the remarkable progress that has been made in the development of gene editing enzymes that enable the generation of transgenic rats directly through targeted genetic modifications in the genomes of zygotes. The simplicity, efficiency and cost-effectiveness of the CRISPR/Cas gene editing system, in particular, mean that the ability to engineer the rat genome is no longer a limiting factor. The selection of suitable targets and gene modifications will now become a priority: a challenge where ES culture and gene editing technologies can play complementary roles in generating accurate bespoke rat models for studying biological processes and modelling human disease.

Interspecific in vitro assay for the chimera-forming ability of human pluripotent stem cells.

PubMed

Masaki, Hideki; Kato-Itoh, Megumi; Umino, Ayumi; Sato, Hideyuki; Hamanaka, Sanae; Kobayashi, Toshihiro; Yamaguchi, Tomoyuki; Nishimura, Ken; Ohtaka, Manami; Nakanishi, Mahito; Nakauchi, Hiromitsu

2015-09-15

Functional assay limitations are an emerging issue in characterizing human pluripotent stem cells (PSCs). With rodent PSCs, chimera formation using pre-implantation embryos is the gold-standard assay of pluripotency (competence of progeny to differentiate into all three germ layers). In human PSCs (hPSCs), however, this can only be monitored via teratoma formation or in vitro differentiation, as ethical concerns preclude generation of human-human or human-animal chimeras. To circumvent this issue, we developed a functional assay utilizing interspecific blastocyst injection and in vitro culture (interspecies in vitro chimera assay) that enables the development and observation of embryos up to headfold stage. The assay uses mouse pre-implantation embryos and rat, monkey and human PSCs to create interspecies chimeras cultured in vitro to the early egg-cylinder stage. Intra- and interspecific chimera assays with rodent PSC lines were performed to confirm the consistency of results in vitro and in vivo. The behavior of chimeras developed in vitro appeared to recapitulate that of chimeras developed in vivo; that is, PSC-derived cells survived and were integrated into the epiblast of egg-cylinder-stage embryos. This indicates that the interspecific in vitro chimera assay is useful in evaluating the chimera-forming ability of rodent PSCs. However, when human induced PSCs (both conventional and naïve-like types) were injected into mouse embryos and cultured, some human cells survived but were segregated; unlike epiblast-stage rodent PSCs, they never integrated into the epiblast of egg-cylinder-stage embryos. These data suggest that the mouse-human interspecies in vitro chimera assay does not accurately reflect the early developmental potential/process of hPSCs. The use of evolutionarily more closely related species as host embryos might be necessary to evaluate the developmental potency of hPSCs. © 2015. Published by The Company of Biologists Ltd.

Chemical synthesis and orexigenic activity of rat/mouse relaxin-3.

PubMed

Hossain, Mohammed Akhter; Smith, Craig M; Ryan, Philip J; Büchler, Elena; Bathgate, Ross A D; Gundlach, Andrew L; Wade, John D

2013-06-01

The insulin-like peptide, relaxin-3 was first identified just a decade ago via a genomic database search and is now recognized to be a key neuropeptide with several roles including the regulation of arousal, stress responses and neuroendocrine homeostasis. It also has significant potential as a drug to treat stress and obesity. Its actions are mediated via its cognate G protein-coupled receptor, RXFP3, which is found in abundant numbers in the brain. However, much remains to be determined with respect to the mechanism of neurological action of this peptide. Consequently, the chemical synthesis of the rat and mouse (which share identical primary structures) two-chain, three disulfide peptide was undertaken and the resulting peptide subjected to detailed in vitro and in vivo assay. Use of efficient solid-phase synthesis methods provided the two regioselectively S-protected A- and B-chains which were readily combined via sequential disulfide bond formation. The synthetic rat/mouse relaxin-3 was obtained in high purity and good overall yield. It demonstrated potent orexigenic activity in rats in that central intracerebroventricular infusion led to significantly increased food intake and water drinking.

A Comparative Study on Aflatoxin B1 Metabolism in Mice and Rats

PubMed Central

Steyn, M.; Pitout, M. J.; Purchase, I. F. H.

1971-01-01

In vivo metabolic studies on rats and mice revealed a marked difference in the fluorescent compounds produced after ingestion of aflatoxin B1. The mouse converted aflatoxin B1 to three unknown fluorescent compounds, designated x1, x2 and x3 and the known aflatoxin M1, while the rat was only capable of producing aflatoxin M1. The results suggested that metabolites x1, x2, x3 and aflatoxin M1 were not part of a major metabolic pathway, but produced independently. These unknown yellowish-green fluorescent compounds did not seem to be conjugated with sulphate or glucuronic acid. In vitro incubations of various mouse liver cell fractions with aflatoxin B1 showed that metabolites x1, x2, x3 and aflatoxin M1, could only be produced by the microsomal fraction and that NADPH was needed as a co-factor. The differences in aflatoxin metabolism by mice and rats are discussed in relation to the apparent resistance of the mouse to the carcinogenic effects of this toxin. PMID:4398926

Tyrosine-Based 1-(S)-[3-hydroxy-2-(phosphonomethoxy)propyl]cytosine and -adenine ((S)-HPMPC and (S)-HPMPA) Prodrugs: Synthesis, Stability, Antiviral Activity and in Vivo Transport Studies

PubMed Central

Zakharova, Valeria M.; Serpi, Michaela; Krylov, Ivan S.; Peterson, Larryn W.; Breitenbach, Julie M.; Borysko, Katherine Z.; Drach, John C.; Collins, Mindy; Hilfinger, John M.; Kashemirov, Boris A.; McKenna, Charles E.

2011-01-01

Eight novel single amino acid (6–11) and dipeptide (12, 13) tyrosine P-O esters of cyclic cidofovir ((S)-cHPMPCa, 4) and its cyclic adenine analog ((S)-cHPMPA, 3) were synthesized and evaluated as prodrugs. In vitro IC50 values for the prodrugs vs vaccinia, cowpox, human cytomegalo- and herpes simplex type 1 viruses were similar to those for the parent drugs ((S)-HPMPC, 2, (S)-HPMPA, 1; IC50 0.3 – 30 µM); there were no cytoxicity with KB or HFF cells at ≤ 100 µM. The prodrugs exhibited a wide range of half-lives in rat intestinal homogenate at pH 6.5 (<30 – 1732 min) with differences of 3–10× between phostonate diastereomers. The tyrosine-alkylamide derivatives of 3 and 4 were the most stable. (L)-Tyr-NHiBu cHPMPA (11) was converted in rat or mouse plasma solely to two active metabolites and had significantly enhanced oral bioavailability vs parent drug 1 in a mouse model (39 % vs <5 %). PMID:21812420

Development of the Nonobese Diabetic Mouse and Contribution of Animal Models for Understanding Type 1 Diabetes.

PubMed

Mullen, Yoko

2017-04-01

In 1974, the discovery of a mouse and a rat that spontaneously developed hyperglycemia led to the development of 2 autoimmune diabetes models: nonobese diabetic (NOD) mouse and Bio-Breeding rat. These models have contributed to our understanding of autoimmune diabetes, provided tools to dissect autoimmune islet damage, and facilitated development of early detection, prevention, and treatment of type 1 diabetes. The genetic characterization, monoclonal antibodies, and congenic strains have made NOD mice especially useful.Although the establishment of the inbred NOD mouse strain was documented by Makino et al (Jikken Dobutsu. 1980;29:1-13), this review will focus on the not-as-well-known history leading to the discovery of a glycosuric female mouse by Yoshihiro Tochino. This discovery was spearheaded by years of effort by Japanese scientists from different disciplines and dedicated animal care personnel and by the support of the Shionogi Pharmaceutical Company, Osaka, Japan. The history is based on the early literature, mostly written in Japanese, and personal communications especially with Dr Tochino, who was involved in diabetes animal model development and who contributed to the release of NOD mice to the international scientific community. This article also reviews the scientific contributions made by the Bio-Breeding rat to autoimmune diabetes.

Bone pain caused by swelling of mouse ear capsule static xylene and effects on rat models of cervical spondylosis

NASA Astrophysics Data System (ADS)

Zhang, Xuhui; Xia, Lei; Hao, Shaojun; Chen, Weiliang; Guo, Junyi; Ma, Zhenzhen; Wang, Huamin; Kong, Xuejun; Wang, Hongyu; Zhang, Zhengchen

2018-04-01

To observe the effect of intravenous bone pain Capsule on the ear of mice induced by xylene, swelling of rat models of cervical spondylosis. Weighing 18 ˜ 21g 50 mice, male, were randomly divided into for five groups, which were fed with service for bone pain static capsule suspension, Jingfukang granule suspension 0.5%CMC liquid and the same volume of. Respectively to the mice ear drop of xylene 0.05 ml, 4h after cervical dislocation, the mice were sacrificed and the cut two ear, rapid analytical balance weighing, and calculate the ear swelling degree and the other to take the weight of 200 - 60 250g male SD rats, were randomly divided into for 6 groups, 10 rats in each group, of which 5 groups made cervical spondylosis model. Results: with the blank group than bone pain static capsule group and Jingfukang granule group can significantly reduce mouse auricular dimethylbenzene swelling, significantly reduce ear swelling degree (P < 0.01); the successful establishment of the rat model of cervical spondylosis. With the model group ratio, large, medium and small dose of bone pain static capsule group, Jingfukang granule group (P < 0.01) angle of swash plate of rats increased significantly, the high and middle dose of bone pain static capsule group, Jingfukang granule group can significantly reduce the rat X-ray scores (P < 0.05). Bone pain static capsule can significantly reduce mouse auricular dimethylbenzene swelling. The bone pain capsule has a good effect on the rat model of cervical spondylosis.

A combined solenoid-surface RF coil for high-resolution whole-brain rat imaging on a 3.0 Tesla clinical MR scanner.

PubMed

Underhill, Hunter R; Yuan, Chun; Hayes, Cecil E

2010-09-01

Rat brain models effectively simulate a multitude of human neurological disorders. Improvements in coil design have facilitated the wider utilization of rat brain models by enabling the utilization of clinical MR scanners for image acquisition. In this study, a novel coil design, subsequently referred to as the rat brain coil, is described that exploits and combines the strengths of both solenoids and surface coils into a simple, multichannel, receive-only coil dedicated to whole-brain rat imaging on a 3.0 T clinical MR scanner. Compared with a multiturn solenoid mouse body coil, a 3-cm surface coil, a modified Helmholtz coil, and a phased-array surface coil, the rat brain coil improved signal-to-noise ratio by approximately 72, 61, 78, and 242%, respectively. Effects of the rat brain coil on amplitudes of static field and radiofrequency field uniformity were similar to each of the other coils. In vivo, whole-brain images of an adult male rat were acquired with a T(2)-weighted spin-echo sequence using an isotropic acquisition resolution of 0.25 x 0.25 x 0.25 mm(3) in 60.6 min. Multiplanar images of the in vivo rat brain with identification of anatomic structures are presented. Improvement in signal-to-noise ratio afforded by the rat brain coil may broaden experiments that utilize clinical MR scanners for in vivo image acquisition. 2010 Wiley-Liss, Inc.

Effect of NASA light-emitting diode irradiation on wound healing.

PubMed

Whelan, H T; Smits, R L; Buchman, E V; Whelan, N T; Turner, S G; Margolis, D A; Cevenini, V; Stinson, H; Ignatius, R; Martin, T; Cwiklinski, J; Philippi, A F; Graf, W R; Hodgson, B; Gould, L; Kane, M; Chen, G; Caviness, J

2001-12-01

The purpose of this study was to assess the effects of hyperbaric oxygen (HBO) and near-infrared light therapy on wound healing. Light-emitting diodes (LED), originally developed for NASA plant growth experiments in space show promise for delivering light deep into tissues of the body to promote wound healing and human tissue growth. In this paper, we review and present our new data of LED treatment on cells grown in culture, on ischemic and diabetic wounds in rat models, and on acute and chronic wounds in humans. In vitro and in vivo (animal and human) studies utilized a variety of LED wavelength, power intensity, and energy density parameters to begin to identify conditions for each biological tissue that are optimal for biostimulation. LED produced in vitro increases of cell growth of 140-200% in mouse-derived fibroblasts, rat-derived osteoblasts, and rat-derived skeletal muscle cells, and increases in growth of 155-171% of normal human epithelial cells. Wound size decreased up to 36% in conjunction with HBO in ischemic rat models. LED produced improvement of greater than 40% in musculoskeletal training injuries in Navy SEAL team members, and decreased wound healing time in crew members aboard a U.S. Naval submarine. LED produced a 47% reduction in pain of children suffering from oral mucositis. We believe that the use of NASA LED for light therapy alone, and in conjunction with hyperbaric oxygen, will greatly enhance the natural wound healing process, and more quickly return the patient to a preinjury/illness level of activity. This work is supported and managed through the NASA Marshall Space Flight Center-SBIR Program.

A physiologically based toxicokinetic model for inhaled ethylene and ethylene oxide in mouse, rat, and human.

PubMed

Filser, Johannes Georg; Klein, Dominik

2018-04-01

Ethylene (ET) is the largest volume organic chemical. Mammals metabolize the olefin to ethylene oxide (EO), another important industrial chemical. The epoxide alkylates macromolecules and has mutagenic and carcinogenic properties. In order to estimate the EO burden in mice, rats, and humans resulting from inhalation exposure to gaseous ET or EO, a physiological toxicokinetic model was developed. It consists of the compartments lung, richly perfused tissues, kidneys, muscle, fat, arterial blood, venous blood, and liver containing the sub-compartment endoplasmic reticulum. Modeled ET metabolism is mediated by hepatic cytochrome P450 2E1, EO metabolism by hepatic microsomal epoxide hydrolase or cytosolic glutathione S-transferase in various tissues. EO is also spontaneously hydrolyzed or conjugated with glutathione. The model was validated on experimental data collected in mice, rats, and humans. Modeled were uptake by inhalation, wash-in-wash-out effect in the upper respiratory airways, distribution into tissues and organs, elimination via exhalation and metabolism, and formation of 2-hydroxyethyl adducts with hemoglobin and DNA. Simulated concentration-time courses of ET or EO in inhaled (gas uptake studies) or exhaled air, and of EO in blood during exposures to ET or EO agreed excellently with measured data. Predicted levels of adducts with DNA and hemoglobin, induced by ET or EO, agreed with reported levels. Exposures to 10000 ppm ET were predicted to induce the same adduct levels as EO exposures to 3.95 (mice), 5.67 (rats), or 0.313 ppm (humans). The model is concluded to be applicable for assessing health risks from inhalation exposure to ET or EO. Copyright © 2017 The Author(s). Published by Elsevier B.V. All rights reserved.

The contribution of Ca2+ signaling and Ca2+ sensitivity to the regulation of airway smooth muscle contraction is different in rats and mice.

PubMed

Bai, Yan; Sanderson, Michael J

2009-06-01

To determine the relative contributions of Ca(2+) signaling and Ca(2+) sensitivity to the contractility of airway smooth muscle cells (SMCs), we compared the contractile responses of mouse and rat airways with the lung slice technique. Airway contraction was measured by monitoring changes in airway lumen area with phase-contrast microscopy, whereas changes in intracellular calcium concentration ([Ca(2+)](i)) of the SMCs were recorded with laser scanning microscopy. In mice and rats, methacholine (MCh) or serotonin induced concentration-dependent airway contraction and Ca(2+) oscillations in the SMCs. However, rat airways demonstrated greater contraction compared with mice, in response to agonist-induced Ca(2+) oscillations of a similar frequency. Because this indicates that rat airway SMCs have a higher Ca(2+) sensitivity compared with mice, we examined Ca(2+) sensitivity with Ca(2+)-permeabilized airway SMCs in which the [Ca(2+)](i) was experimentally controlled. In the absence of agonists, high [Ca(2+)](i) induced a sustained contraction in rat airways but only a transient contraction in mouse airways. This sustained contraction of rat airways was relaxed by Y-23672, a Rho kinase inhibitor, but not affected by GF-109203X, a PKC inhibitor. The subsequent exposure of Ca(2+)-permeabilized airway SMCs, with high [Ca(2+)](i), to MCh elicited a further contraction of rat airways and initiated a sustained contraction of mouse airways, without changing the [Ca(2+)](i) of the SMCs. Collectively, these results indicate that airway SMCs of rats have a substantially higher innate Ca(2+) sensitivity than mice and that this strongly influences the transduction of the frequency of Ca(2+) oscillations into the contractility of airway SMCs.

Rift Valley fever virus infection in golden Syrian hamsters.

PubMed

Scharton, Dionna; Van Wettere, Arnaud J; Bailey, Kevin W; Vest, Zachary; Westover, Jonna B; Siddharthan, Venkatraman; Gowen, Brian B

2015-01-01

Rift Valley fever virus (RVFV) is a formidable pathogen that causes severe disease and abortion in a variety of livestock species and a range of disease in humans that includes hemorrhagic fever, fulminant hepatitis, encephalitis and blindness. The natural transmission cycle involves mosquito vectors, but exposure can also occur through contact with infected fluids and tissues. The lack of approved antiviral therapies and vaccines for human use underlies the importance of small animal models for proof-of-concept efficacy studies. Several mouse and rat models of RVFV infection have been well characterized and provide useful systems for the study of certain aspects of pathogenesis, as well as antiviral drug and vaccine development. However, certain host-directed therapeutics may not act on mouse or rat pathways. Here, we describe the natural history of disease in golden Syrian hamsters challenged subcutaneously with the pathogenic ZH501 strain of RVFV. Peracute disease resulted in rapid lethality within 2 to 3 days of RVFV challenge. High titer viremia and substantial viral loads were observed in most tissues examined; however, histopathology and immunostaining for RVFV antigen were largely restricted to the liver. Acute hepatocellular necrosis associated with a strong presence of viral antigen in the hepatocytes indicates that fulminant hepatitis is the likely cause of mortality. Further studies to assess the susceptibility and disease progression following respiratory route exposure are warranted. The use of the hamsters to model RVFV infection is suitable for early stage antiviral drug and vaccine development studies.

Cloning and expression of sheep renal K-CI cotransporter-1.

PubMed

Zhang, Jin J; Misri, Sandeep; Adragna, Norma C; Gagnon, Kenneth B E; Fyffe, Robert E W; Lauf, Peter K

2005-01-01

Sheep K-Cl cotransporter-1(shKCC1) cDNA was cloned from kidney by RT-PCR with an open reading frame of 3258 base pairs exhibiting 92%, 90%, 88% and 87% identity with pig, rabbit and human, rat and mouse KCC1 cDNAs, respectively, encoding an approximately 122 kDa polypeptide of 1086-amino acids. Hydropathy analysis reveals the familiar KCC1 topology with 12 transmembrane domains (TMDs) and the hydrophilic NH2-terminal (NTD) and COOH-terminal (CTD) domains both at the cytoplasmic membrane face. However, shKCC1 has two rather than one large extracellular loops (ECL): ECL3 between TMDs 5 and 6, and ECL6, between TMDs 11 and 12. The translated shKCC1 protein differs in 12 amino acid residues from other KCC1s, mainly within the NTD, ECL3, ICL4, ECL6, and CTD. Notably, a tyrosine residue at position 996 replaces aspartic acid conserved in all other species. Human embryonic kidney (HEK293) cells and mouse NIH/3T3 fibroblasts, transiently transfected with shKCCI-cDNA, revealed the glycosylated approximately 150 kDa proteins by Western blots and positive immunofluorescence-staining with polyclonal rabbit anti-ratKCC1 antibodies. ShKCC1 was functionally expressed in NIH/3T3 cells by an elevated basal Cl-dependent K influx measured with Rb as K-congener that was stimulated three-fold by the KCC-activator N-ethylmaleimide. Copyright (c) 2005 S. Karger AG, Basel.

Exploring the radiosynthesis and in vitro characteristics of [68 Ga]Ga-DOTA-Siglec-9.

PubMed

Jensen, Svend B; Käkelä, Meeri; Jødal, Lars; Moisio, Olli; Alstrup, Aage K O; Jalkanen, Sirpa; Roivainen, Anne

2017-07-01

Vascular adhesion protein 1 is a leukocyte homing-associated glycoprotein, which upon inflammation rapidly translocates from intracellular sources to the endothelial cell surface. It has been discovered that the cyclic peptide residues 283-297 of sialic acid-binding IgG-like lectin 9 (Siglec-9) "CARLSLSWRGLTLCPSK" bind to vascular adhesion protein 1 and hence makes the radioactive analogues of this compound ([ 68 Ga]Ga-DOTA-Siglec-9) interesting as a noninvasive visualizing marker of inflammation. Three different approaches to the radiosynthesis of [ 68 Ga]Ga-DOTA-Siglec-9 are presented and compared with previously published methods. A simple, robust radiosynthesis of [ 68 Ga]Ga-DOTA-Siglec-9 with a yield of 62% (non decay-corrected) was identified, and it had a radiochemical purity >98% and a specific radioactivity of 35 MBq/nmol. Furthermore, the protein binding and stability of [ 68 Ga]Ga-DOTA-Siglec-9 were analyzed in vitro in mouse, rat, rabbit, pig, and human plasma and compared with in vivo pig results. The plasma in vitro protein binding of [ 68 Ga]Ga-DOTA-Siglec-9 was the lowest in the pig followed by rabbit, human, rat, and mouse. It was considerably higher in the in vivo pig experiments. The in vivo stability in pigs was lower than the in vitro stability. Despite considerable species differences, the observed characteristics of [ 68 Ga]Ga-DOTA-Siglec-9 are suitable as a positron emission tomography tracer. Copyright © 2017 John Wiley & Sons, Ltd.

TXNIP regulates myocardial fatty acid oxidation via miR-33a signaling.

PubMed

Chen, Junqin; Young, Martin E; Chatham, John C; Crossman, David K; Dell'Italia, Louis J; Shalev, Anath

2016-07-01

Myocardial fatty acid β-oxidation is critical for the maintenance of energy homeostasis and contractile function in the heart, but its regulation is still not fully understood. While thioredoxin-interacting protein (TXNIP) has recently been implicated in cardiac metabolism and mitochondrial function, its effects on β-oxidation have remained unexplored. Using a new cardiomyocyte-specific TXNIP knockout mouse and working heart perfusion studies, as well as loss- and gain-of-function experiments in rat H9C2 and human AC16 cardiomyocytes, we discovered that TXNIP deficiency promotes myocardial β-oxidation via signaling through a specific microRNA, miR-33a. TXNIP deficiency leads to increased binding of nuclear factor Y (NFYA) to the sterol regulatory element binding protein 2 (SREBP2) promoter, resulting in transcriptional inhibition of SREBP2 and its intronic miR-33a. This allows for increased translation of the miR-33a target genes and β-oxidation-promoting enzymes, carnitine octanoyl transferase (CROT), carnitine palmitoyl transferase 1 (CPT1), hydroxyacyl-CoA dehydrogenase/3-ketoacyl-CoA thiolase/enoyl-CoA hydratase-β (HADHB), and AMPKα and is associated with an increase in phospho-AMPKα and phosphorylation/inactivation of acetyl-CoA-carboxylase. Thus, we have identified a novel TXNIP-NFYA-SREBP2/miR-33a-AMPKα/CROT/CPT1/HADHB pathway that is conserved in mouse, rat, and human cardiomyocytes and regulates myocardial β-oxidation. Copyright © 2016 the American Physiological Society.

Comparative study of the primary cilia in thyrocytes of adult mammals

PubMed Central

Utrilla, J C; Gordillo-Martínez, F; Gómez-Pascual, A; Fernández-Santos, J M; Garnacho, C; Vázquez-Román, V; Morillo-Bernal, J; García-Marín, R; Jiménez-García, A; Martín-Lacave, I

2015-01-01

Since their discovery in different human tissues by Zimmermann in 1898, primary cilia have been found in the vast majority of cell types in vertebrates. Primary cilia are considered to be cellular antennae that occupy an ideal cellular location for the interpretation of information both from the environment and from other cells. To date, in mammalian thyroid gland, primary cilia have been found in the thyrocytes of humans and dogs (fetuses and adults) and in rat embryos. The present study investigated whether the existence of this organelle in follicular cells is a general event in the postnatal thyroid gland of different mammals, using both immunolabeling by immunofluorescence and electron microscopy. Furthermore, we aimed to analyse the presence of primary cilia in various thyroid cell lines. According to our results, primary cilia are present in the adult thyroid gland of most mammal species we studied (human, pig, guinea pig and rabbit), usually as a single copy per follicular cell. Strikingly, they were not found in rat or mouse thyroid tissues. Similarly, cilia were also observed in all human thyroid cell lines tested, both normal and neoplastic follicular cells, but not in cultured thyrocytes of rat origin. We hypothesize that primary cilia could be involved in the regulation of normal thyroid function through specific signaling pathways. Nevertheless, further studies are needed to shed light on the permanence of these organelles in the thyroid gland of most species during postnatal life. PMID:26228270

High-throughput quantitation of amino acids in rat and mouse biological matrices using stable isotope labeling and UPLC-MS/MS analysis.

PubMed

Takach, Edward; O'Shea, Thomas; Liu, Hanlan

2014-08-01

Quantifying amino acids in biological matrices is typically performed using liquid chromatography (LC) coupled with fluorescent detection (FLD), requiring both derivatization and complete baseline separation of all amino acids. Due to its high specificity and sensitivity, the use of UPLC-MS/MS eliminates the derivatization step and allows for overlapping amino acid retention times thereby shortening the analysis time. Furthermore, combining UPLC-MS/MS with stable isotope labeling (e.g., isobaric tag for relative and absolute quantitation, i.e., iTRAQ) of amino acids enables quantitation while maintaining sensitivity, selectivity and speed of analysis. In this study, we report combining UPLC-MS/MS analysis with iTRAQ labeling of amino acids resulting in the elution and quantitation of 44 amino acids within 5 min demonstrating the speed and convenience of this assay over established approaches. This chromatographic analysis time represented a 5-fold improvement over the conventional HPLC-MS/MS method developed in our laboratory. In addition, the UPLC-MS/MS method demonstrated improvements in both specificity and sensitivity without loss of precision. In comparing UPLC-MS/MS and HPLC-MS/MS results of 32 detected amino acids, only 2 amino acids exhibited imprecision (RSD) >15% using UPLC-MS/MS, while 9 amino acids exhibited RSD >15% using HPLC-MS/MS. Evaluating intra- and inter-assay precision over 3 days, the quantitation range for 32 detected amino acids in rat plasma was 0.90-497 μM, with overall mean intra-day precision of less than 15% and mean inter-day precision of 12%. This UPLC-MS/MS assay was successfully implemented for the quantitative analysis of amino acids in rat and mouse plasma, along with mouse urine and tissue samples, resulting in the following concentration ranges: 0.98-431 μM in mouse plasma for 32 detected amino acids; 0.62-443 μM in rat plasma for 32 detected amino acids; 0.44-8590μM in mouse liver for 33 detected amino acids; 0.61-1241 μM in mouse kidney for 37 detected amino acids; and 1.39-1,681 μM in rat urine for 34 detected amino acids. The utility of the assay was further demonstrated by measuring and comparing plasma amino acid levels between pre-diabetic Zucker diabetic fatty rats (ZDF/Gmi fa/fa) and their lean littermates (ZDF/Gmi fa/?). Significant differences (P<0.001) in 9 amino acid concentrations were observed, with the majority ranging from a 2- to 5-fold increase in pre-diabetic ZDF rats on comparison with ZDF lean rats, consistent with previous literature reports. Copyright © 2014 Elsevier B.V. All rights reserved.

Comparative immunology of allergic responses.

PubMed

Gershwin, Laurel J

2015-01-01

Allergic responses occur in humans, rodents, non-human primates, avian species, and all of the domestic animals. These responses are mediated by immunoglobulin E (IgE) antibodies that bind to mast cells and cause release/synthesis of potent mediators. Clinical syndromes include naturally occurring asthma in humans and cats; atopic dermatitis in humans, dogs, horses, and several other species; food allergies; and anaphylactic shock. Experimental induction of asthma in mice, rats, monkeys, sheep, and cats has helped to reveal mechanisms of pathogenesis of asthma in humans. All of these species share the ability to develop a rapid and often fatal response to systemic administration of an allergen--anaphylactic shock. Genetic predisposition to development of allergic disease (atopy) has been demonstrated in humans, dogs, and horses. Application of mouse models of IgE-mediated allergic asthma has provided evidence for a role of air pollutants (ozone, diesel exhaust, environmental tobacco smoke) in enhanced sensitization to allergens.

New products tissue-engineering in the treatment of spinal cord injury

NASA Astrophysics Data System (ADS)

Bolshakov, I. N.; Sergienko, V. I.; Kiselev, S. L.; Lagarkova, M. A.; Remigaylo, A. A.; Mihaylov, A. A.; Prokopenko, S. V.

2015-11-01

In the treatment of patients with complicated spinal cord injury the Russian Health spends about one million rubles for each patient in the acute and the interim period after the injury. The number of complicated spinal cord injury is different in geographical areas Russian Federation from 30 to 50 people per 1 million that is affected by the year 5600. Applied to the present surgical and pharmacological techniques provide unsatisfactory results or minimally effective treatment. Transplantation of 100 thousand neuronal mouse predecessors (24 rats) or human neuronal predecessors (18 rats) in the anatomical gap rat spinal cord, followed by analysis of neurological deficit. The neuro-matrix implantation in the rat spinal cord containing 100 thousand neuronal precursors hESC, repeatable control neuro-matrix transplantation, non-cell mass, eliminating neurological deficit for 14 weeks after transplantation about 5-9 points on the scale of the BBB. The cultivation under conditions in vitro human induced pluripotent stem cells on collagen-chitosan matrix (hIPSC) showed that neurons differentiated from induced pluripotent stem cells grown on scaffolds as compact groups and has no neurites. Cells do not penetrate into the matrix during long-term cultivation and formed near the surface of the spherical structures resembling neurospheres. At least 90% of the cells were positive for the neuronal marker tubulin b3. Further studies should be performed to examine the compatibility of neuronal cultures and matrices.

Comparison between surrogate indexes of insulin sensitivity/resistance and hyperinsulinemic euglycemic clamp estimates in rats

PubMed Central

Muniyappa, Ranganath; Chen, Hui; Muzumdar, Radhika H.; Einstein, Francine H.; Yan, Xu; Yue, Lilly Q.; Barzilai, Nir

2009-01-01

Assessing insulin resistance in rodent models gives insight into mechanisms that cause type 2 diabetes and the metabolic syndrome. The hyperinsulinemic euglycemic glucose clamp, the reference standard for measuring insulin sensitivity in humans and animals, is labor intensive and technically demanding. A number of simple surrogate indexes of insulin sensitivity/resistance have been developed and validated primarily for use in large human studies. These same surrogates are also frequently used in rodent studies. However, in general, these indexes have not been rigorously evaluated in animals. In a recent validation study in mice, we demonstrated that surrogates have a weaker correlation with glucose clamp estimates of insulin sensitivity/resistance than in humans. This may be due to increased technical difficulties in mice and/or intrinsic differences between human and rodent physiology. To help distinguish among these possibilities, in the present study, using data from rats substantially larger than mice, we compared the clamp glucose infusion rate (GIR) with surrogate indexes, including QUICKI, HOMA, 1/HOMA, log (HOMA), and 1/fasting insulin. All surrogates were modestly correlated with GIR (r = 0.34–0.40). Calibration analyses of surrogates adjusted for body weight demonstrated similar predictive accuracy for GIR among all surrogates. We conclude that linear correlations of surrogate indexes with clamp estimates and predictive accuracy of surrogate indexes in rats are similar to those in mice (but not as substantial as in humans). This additional rat study (taken with the previous mouse study) suggests that application of surrogate insulin sensitivity indexes developed for humans may not be appropriate for determining primary outcomes in rodent studies due to intrinsic differences in metabolic physiology. However, use of surrogates may be appropriate in rodents, where feasibility of clamps is an obstacle and measurement of insulin sensitivity is a secondary outcome. PMID:19706785

Protective effects of papaverine salicylate in mouse ear dermatitis and PAF-induced rat paw oedema.

PubMed

de Bernardis, E; Leonardi, G; Caruso, A; Cutuli, V M; Amico-Roxas, M

1994-08-01

Papaverine salicylate (MR-800) has been tested as a topical antiinflammatory agent in several models of skin inflammation in rodents, such as mouse ear dermatitis induced by croton oil, cantharidin or zymosan, and rat paw oedema induced by PAF. MR-800 exerted a dose-dependent inhibitory activity in all assays, when equimolar doses of sodium salicylate or papaverine were less effective, suggesting the existence of a favourable synergism between salicylate and papaverine.

Relationship of Metabolism and Cell Proliferation to the Mode of Action of Fluensulfone-Induced Mouse Lung Tumors. II: Additional Mechanistic Studies.

PubMed

Strupp, Christian; Bomann, Werner; Cohen, Samuel M; Weber, Klaus

2016-12-01

Fluensulfone is a nematicide for agricultural use. Chronic dietary exposure led to bronchiolo-alveolar hyperplasia and bronchiolo-alveolar adenomas in CD-1 mice but not in rats. Genotoxicity could be excluded as a mode of action (MOA). An earlier publication (Strupp, C., Banas, D. A., Cohen, S. M., Gordon, E. B., Jaeger, M., and Weber, K. (2012). Relationship of metabolism and cell proliferation to the mode of action of fluensulfone-induced mouse lung tumors: analysis of their human relevance using the IPCS framework. Toxicol. Sci. 128, 284-294.) reported MOA studies identifying the following key events: increased metabolism of fluensulfone by CYP2f2 in mouse lung Club cells, followed by local proliferation, finally leading to adenoma formation. Human lung microsomes were found not to metabolize fluensulfone. The Joint FAO/WHO Meeting on Pesticide Residues has reviewed the previous data and concluded that the MOA is plausible however some areas of uncertainty were identified. This publication provides additional data to address these. New cell proliferation studies in mice showed that the MOA is functionally independent of sex. A threshold of cell proliferation in Club cells correlating with the dose response for adenoma formation was shown. CYP2f2 knockout mice did not react to fluensulfone exposure with cell proliferation like wild-type mice, confirming the key role of this enzyme. The collective data for fluensulfone were evaluated according to the International Programme on Chemical Safety (IPCS) Mode of Action Framework which leads to the conclusion that the mouse-specific lung tumors after fluensulfone are not relevant to humans. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Specificity and rate of human and mouse liver and plasma phosphatidylcholine synthesis analyzed in vivo[S

PubMed Central

Pynn, Christopher J.; Henderson, Neil G.; Clark, Howard; Koster, Grielof; Bernhard, Wolfgang; Postle, Anthony D.

2011-01-01

Phosphatidylcholine (PC) synthesis by the direct cytidine diphosphate choline (CDP-choline) pathway in rat liver generates predominantly mono- and di-unsaturated molecular species, while polyunsaturated PC species are synthesized largely by the phosphatidylethanolamine-N-methyltransferase (PEMT) pathway. Although altered PC synthesis has been suggested to contribute to development of hepatocarcinoma and nonalcoholic steatohepatitis, analysis of the specificity of hepatic PC metabolism in human patients has been limited by the lack of sensitive and safe methodologies. Here we incorporated a deuterated methyl-d9-labled choline chloride, to quantify biosynthesis fluxes through both of the PC synthetic pathways in vivo in human volunteers and compared these fluxes with those in mice. Rates and molecular specificities of label incorporated into mouse liver and plasma PC were very similar and strongly suggest that label incorporation into human plasma PC can provide a direct measure of hepatic PC synthesis in human subjects. Importantly, we demonstrate for the first time that the PEMT pathway in human liver is selective for polyunsaturated PC species, especially those containing docosahexaenoic acid. Finally, we present a multiple isotopomer distribution analysis approach, based on transfer of deuterated methyl groups to S-adenosylmethionine and subsequent sequential methylations of PE, to quantify absolute flux rates through the PEMT pathway that are applicable to studies of liver dysfunction in clinical studies. PMID:21068006

Esculin exhibited anti-inflammatory activities in vivo and regulated TNF-α and IL-6 production in LPS-stimulated mouse peritoneal macrophages in vitro through MAPK pathway.

PubMed

Niu, Xiaofeng; Wang, Yu; Li, Weifeng; Zhang, Hailin; Wang, Xiumei; Mu, Qingli; He, Zehong; Yao, Huan

2015-12-01

Esculin, a coumarinic derivative found in Aesculus hippocastanum L. (Horse-chestnut), has been reported to have potent anti-inflammatory properties. The present study is designed to investigate the protective effects of esculin on various inflammation models in vivo and in vitro and to clarify the possible mechanism. Induced-animal models of inflammation and lipopolysaccharide (LPS)-challenged mouse peritoneal macrophages were used to examine the anti-inflammatory activity of esculin. In present study, xylene-induced mouse ear edema, carrageenan-induced rat paw edema, and carrageenan-induced mouse pleurisy were attenuated by esculin. In vitro, the pro-inflammatory cytokine levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in supernatant were reduced by esculin. Meanwhile, we found that esculin significantly inhibited LPS-induced activation of mitogen-activated protein kinase (MAPK) pathway in peritoneal macrophages. These results suggest that esculin has potent anti-inflammatory activities in vivo and in vitro, which may involve the inhibition of the MAPK pathway. Esculin may be a promising preventive agent for inflammatory diseases in human. Copyright © 2015 Elsevier B.V. All rights reserved.

Subretinal delivery and electroporation in pigmented and nonpigmented adult mouse eyes

PubMed Central

Nickerson, John M.; Goodman, Penny; Chrenek, Micah A.; Johnson, Christiana J.; Berglin, Lennart; Redmond, T. Michael.; Boatright, Jeffrey H.

2013-01-01

Subretinal injection offers one of the best ways to deliver many classes of drugs, reagents, cells and treatments to the photoreceptor, Müller, and retinal pigment epithelium (RPE) cells of the retina. Agents delivered to this space are placed within microns of the intended target cell, accumulating to high concentrations because there is no dilution due to transport processes or diffusion. Dilution in the interphotoreceptor space (IPS) is minimal because the IPS volume is only 10-20 microliters in the human eye and less than 1 microliter in the mouse eye. For gene delivery purposes, we wished to transfect the cells adjacent to the IPS in adult mouse eyes. Others transfect these cells in neonatal rats to study the development of the retina. In both neonates and adults, electroporation is found to be effective Here we describe the optimization of electroporation conditions for RPE cells in the adult mouse eye with naked plasmids. However, both techniques, subretinal injection and electroporation, present some technical challenges that require skill on the part of the surgeon to prevent untoward damage to the eye. Here we describe methods that we have used for the past ten years (1). PMID:22688698

The Peptide Near the C Terminus Regulates Receptor CAR Nuclear Translocation Induced by Xenochemicals in Mouse Liver

PubMed Central

Zelko, Igor; Sueyoshi, Tatsuya; Kawamoto, Takeshi; Moore, Rick; Negishi, Masahiko

2001-01-01

In response to phenobarbital (PB) and other PB-type inducers, the nuclear receptor CAR translocates to the mouse liver nucleus (T. Kawamoto et al., Mol. Cell. Biol. 19:6318–6322, 1999). To define the translocation mechanism, fluorescent protein-tagged human CAR (hCAR) was expressed in the mouse livers using the in situ DNA injection and gene delivery systems. As in the wild-type hCAR, the truncated receptor lacking the C-terminal 10 residues (i.e., AF2 domain) translocated to the nucleus, indicating that the PB-inducible translocation is AF2 independent. Deletion of the 30 C-terminal residues abolished the receptor translocation, and subsequent site-directed mutagenesis delineated the PB-inducible translocation activity of the receptor to the peptide L313GLL316AEL319. Ala mutations of Leu313, Leu316, or Leu319 abrogated the translocation of CAR in the livers, while those of Leu312 or Leu315 did not affect the nuclear translocation. The leucine-rich peptide dictates the nuclear translocation of hCAR in response to various PB-type inducers and appears to be conserved in the mouse and rat receptors. PMID:11283262

Differential Regulation of Endosomal GPCR/β-Arrestin Complexes and Trafficking by MAPK*

PubMed Central

Khoury, Etienne; Nikolajev, Ljiljana; Simaan, May; Namkung, Yoon; Laporte, Stéphane A.

2014-01-01

β-Arrestins are signaling adaptors that bind to agonist-occupied G protein-coupled receptors (GPCRs) and target them for endocytosis; however, the mechanisms regulating receptor/β-arrestin complexes and trafficking in endosomes, remain ill defined. Here we show, in live cells, differential dynamic regulation of endosomal bradykinin B2 receptor (B2R) complexes with either β-arrestin-1 or -2. We find a novel role for MAPK in the B2R/β-arrestin-2 complex formation, receptor trafficking and signaling mediated by an ERK1/2 regulatory motif in the hinge domain of the rat β-arrestin-2 (PET178P), but not rat β-arrestin-1 (PER177P). While the ERK1/2 regulatory motif is conserved between rat and mouse β-arrestin-2, it is surprisingly not conserved in human β-arrestin-2 (PEK178P). However, mutation of lysine 178 to threonine is sufficient to confer MAPK sensitivity to the human β-arrestin-2. Furthermore, substitution for a phosphomimetic residue in both the rat and the human β-arrestin-2 (T/K178D) significantly stabilizes B2R/β-arrestin complexes in endosomes, delays receptor recycling to the plasma membrane and maintains intracellular MAPK signaling. Similarly, the endosomal trafficking of β2-adrenergic, angiotensin II type 1 and vasopressin V2 receptors was altered by the β-arrestin-2 T178D mutant. Our findings unveil a novel subtype specific mode of MAPK-dependent regulation of β-arrestins in intracellular trafficking and signaling of GPCRs, and suggest differential endosomal receptor/β-arrestin-2 signaling roles among species. PMID:25016018

DOE Office of Scientific and Technical Information (OSTI.GOV)

Buckpitt, Alan, E-mail: arbuckpitt@ucdavis.edu; Morin, Dexter; Murphy, Shannon

Naphthalene produces species and cell selective injury to respiratory tract epithelial cells of rodents. In these studies we determined the apparent K{sub m}, V{sub max}, and catalytic efficiency (V{sub max}/K{sub m}) for naphthalene metabolism in microsomal preparations from subcompartments of the respiratory tract of rodents and non-human primates. In tissues with high substrate turnover, major metabolites were derived directly from naphthalene oxide with smaller amounts from conjugates of diol epoxide, diepoxide, and 1,2- and 1,4-naphthoquinones. In some tissues, different enzymes with dissimilar K{sub m} and V{sub max} appeared to metabolize naphthalene. The rank order of V{sub max} (rat olfactory epitheliummore » > mouse olfactory epithelium > murine airways ≫ rat airways) correlated well with tissue susceptibility to naphthalene. The V{sub max} in monkey alveolar subcompartment was 2% that in rat nasal olfactory epithelium. Rates of metabolism in nasal compartments of the monkey were low. The catalytic efficiencies of microsomes from known susceptible tissues/subcompartments are 10 and 250 fold higher than in rat airway and monkey alveolar subcompartments, respectively. Although the strong correlations between catalytic efficiencies and tissue susceptibility suggest that non-human primate tissues are unlikely to generate metabolites at a rate sufficient to produce cellular injury, other studies showing high levels of formation of protein adducts support the need for additional studies. - Highlights: • Naphthalene is metabolized with high catalytic efficiency in susceptible tissue. • Naphthalene is metabolized at low catalytic efficiency in non-susceptible tissue. • Respiratory tissues of the non human primate metabolize naphthalene slowly.« less

Translational Rodent Models for Research on Parasitic Protozoa—A Review of Confounders and Possibilities

PubMed Central

Ehret, Totta; Torelli, Francesca; Klotz, Christian; Pedersen, Amy B.; Seeber, Frank

2017-01-01

Rodents, in particular Mus musculus, have a long and invaluable history as models for human diseases in biomedical research, although their translational value has been challenged in a number of cases. We provide some examples in which rodents have been suboptimal as models for human biology and discuss confounders which influence experiments and may explain some of the misleading results. Infections of rodents with protozoan parasites are no exception in requiring close consideration upon model choice. We focus on the significant differences between inbred, outbred and wild animals, and the importance of factors such as microbiota, which are gaining attention as crucial variables in infection experiments. Frequently, mouse or rat models are chosen for convenience, e.g., availability in the institution rather than on an unbiased evaluation of whether they provide the answer to a given question. Apart from a general discussion on translational success or failure, we provide examples where infections with single-celled parasites in a chosen lab rodent gave contradictory or misleading results, and when possible discuss the reason for this. We present emerging alternatives to traditional rodent models, such as humanized mice and organoid primary cell cultures. So-called recombinant inbred strains such as the Collaborative Cross collection are also a potential solution for certain challenges. In addition, we emphasize the advantages of using wild rodents for certain immunological, ecological, and/or behavioral questions. The experimental challenges (e.g., availability of species-specific reagents) that come with the use of such non-model systems are also discussed. Our intention is to foster critical judgment of both traditional and newly available translational rodent models for research on parasitic protozoa that can complement the existing mouse and rat models. PMID:28638807

Translational Rodent Models for Research on Parasitic Protozoa-A Review of Confounders and Possibilities.

PubMed

Ehret, Totta; Torelli, Francesca; Klotz, Christian; Pedersen, Amy B; Seeber, Frank

2017-01-01

Rodents, in particular Mus musculus , have a long and invaluable history as models for human diseases in biomedical research, although their translational value has been challenged in a number of cases. We provide some examples in which rodents have been suboptimal as models for human biology and discuss confounders which influence experiments and may explain some of the misleading results. Infections of rodents with protozoan parasites are no exception in requiring close consideration upon model choice. We focus on the significant differences between inbred, outbred and wild animals, and the importance of factors such as microbiota, which are gaining attention as crucial variables in infection experiments. Frequently, mouse or rat models are chosen for convenience, e.g., availability in the institution rather than on an unbiased evaluation of whether they provide the answer to a given question. Apart from a general discussion on translational success or failure, we provide examples where infections with single-celled parasites in a chosen lab rodent gave contradictory or misleading results, and when possible discuss the reason for this. We present emerging alternatives to traditional rodent models, such as humanized mice and organoid primary cell cultures. So-called recombinant inbred strains such as the Collaborative Cross collection are also a potential solution for certain challenges. In addition, we emphasize the advantages of using wild rodents for certain immunological, ecological, and/or behavioral questions. The experimental challenges (e.g., availability of species-specific reagents) that come with the use of such non-model systems are also discussed. Our intention is to foster critical judgment of both traditional and newly available translational rodent models for research on parasitic protozoa that can complement the existing mouse and rat models.

Exposure to perfluorooctane sulfonate during pregnancy in rat and mouse. I: maternal and prenatal evaluations.

PubMed

Thibodeaux, Julie R; Hanson, Roger G; Rogers, John M; Grey, Brian E; Barbee, Brenda D; Richards, Judy H; Butenhoff, John L; Stevenson, Lisa A; Lau, Christopher

2003-08-01

The maternal and developmental toxicities of perfluorooctane sulfonate (PFOS, C8F17SO3-) were evaluated in the rat and mouse. PFOS is an environmentally persistent compound used as a surfactant and occurs as a degradation product of both perfluorooctane sulfonyl fluoride and substituted perfluorooctane sulfonamido components found in many commercial and consumer applications. Pregnant Sprague-Dawley rats were given 1, 2, 3, 5, or 10 mg/kg PFOS daily by gavage from gestational day (GD) 2 to GD 20; CD-1 mice were similarly treated with 1, 5, 10, 15, and 20 mg/kg PFOS from GD 1 to GD 17. Controls received 0.5% Tween-20 vehicle (1 ml/kg for rats and 10 ml/kg for mice). Maternal weight gain, food and water consumption, and serum chemistry were monitored. Rats were euthanized on GD 21 and mice on GD 18. PFOS levels in maternal serum and in maternal and fetal livers were determined. Maternal weight gains in both species were suppressed by PFOS in a dose-dependent manner, likely attributed to reduced food and water intake. Serum PFOS levels increased with dosage, and liver levels were approximately fourfold higher than serum. Serum thyroxine (T4) and triiodothyronine (T3) in the PFOS-treated rat dams were significantly reduced as early as one week after chemical exposure, although no feedback response of thyroid-stimulating hormone (TSH) was observed. A similar pattern of reduction in T4 was also seen in the pregnant mice. Maternal serum triglycerides were significantly reduced, particularly in the high-dose groups, although cholesterol levels were not affected. In the mouse dams, PFOS produced a marked enlargement of the liver at 10 mg/kg and higher dosages. In the rat fetuses, PFOS was detected in the liver but at levels nearly half of those in the maternal counterparts, regardless of administered doses. In both rodent species, PFOS did not alter the numbers of implantations or live fetuses at term, although small deficits in fetal weight were noted in the rat. A host of birth defects, including cleft palate, anasarca, ventricular septal defect, and enlargement of the right atrium, were seen in both rats and mice, primarily in the 10 and 20 mg/kg dosage groups, respectively. Our results demonstrate both maternal and developmental toxicity of PFOS in the rat and mouse.

Cytokine stimulation of MUC4 expression in human female reproductive tissue carcinoma cell lines and endometrial cancer.

PubMed

Chapela, Patricia J; Broaddus, Russell R; Hawkins, Shannon M; Lessey, Bruce A; Carson, Daniel D

2015-11-01

MUC4, a transmembrane glycoprotein, interferes with cell adhesion, and promotes EGFR signaling in cancer. Studies in rat models have demonstrated steroid hormonal regulation of endometrial MUC4 expression. In this study, qRT-PCR screening of mouse tissues determined that Muc4 mRNA also was robustly expressed in mouse uteri. Previous studies from our labs have demonstrated MUC4 mRNA was expressed at levels <1% of MUC1 mRNA in human endometrium and endometriotic tissue. Multiple human endometrial adenocarcinoma cell lines were assayed for MUC4 mRNA expression revealing extremely low basal expression in the Ishikawa, RL-95-2, AN3CA, and KLE lines. Moderate to high expression was observed in HEC50 and HEC-1A cells. MUC4 mRNA expression was not affected by progesterone and/or estrogen treatment, but was greatly stimulated at both mRNA and protein levels by proinflammatory cytokines (IFN-γ and TNF-α), particularly when used in combination. In endometrial tissue, MUC4 mRNA levels did not change significantly between normal or cancerous samples; although, a subset of patients with grade 1 and 2 tumors displayed substantially higher expression. Likewise, immunostaining of human endometrial adenocarcinoma tissues revealed little to no staining in many patients (low MUC4), but strong staining in some patients (high MUC4) independent of cancer grade. In cases where staining was observed, it was heterogeneous with some cells displaying robust MUC4 expression and others displaying little or no staining. Collectively, these observations demonstrate that while MUC4 is highly expressed in the mouse uterus, it is not a major mucin in normal human endometrium. Rather, MUC4 is a potential marker of endometrial adenocarcinoma in a subset of patients. © 2015 Wiley Periodicals, Inc.

Heterophilic binding of the adhesion molecules poliovirus receptor and immunoglobulin superfamily 4A in the interaction between mouse spermatogenic and Sertoli cells.

PubMed

Wakayama, Tomohiko; Sai, Yoshimichi; Ito, Akihiko; Kato, Yukio; Kurobo, Miho; Murakami, Yoshinori; Nakashima, Emi; Tsuji, Akira; Kitamura, Yukihiko; Iseki, Shoichi

2007-06-01

The cell adhesion protein immunoglobulin superfamily 4A (IGSF4A) is expressed on the surfaces of spermatogenic cells in the mouse testis. During spermatogenesis, IGSF4A is considered to bind to the surface of Sertoli cells in a heterophilic manner. To identify this unknown partner of IGSF4A, we generated rat monoclonal antibodies against the membrane proteins of mouse Sertoli cells grown in primary culture. Using these monoclonal antibodies, we isolated a clone that immunostained Sertoli cells and reacted with the product of immunoprecipitation of the homogenate of mouse testis with anti-IGSF4A antibody. Subsequently, to identify the Sertoli cell membrane protein that is recognized by this monoclonal antibody, we performed expression cloning of a cDNA library from the mouse testis. As a result, we identified poliovirus receptor (PVR), which is another IGSF-type cell adhesion molecule, as the binding partner of IGSF4A. The antibodies raised against PVR and IGSF4A immunoprecipitated both antigens in the homogenate of mouse testis. Immunoreactivity for PVR was present in Sertoli cells but not in spermatogenic cells at all stages of spermatogenesis. Overexpression of PVR in TM4, a mouse Sertoli cell line, increased more than three-fold its capacity to adhere to Tera-2, which is a human cell line that expresses IGSF4A. These findings suggest that the heterophilic binding of PVR to IGSF4A is responsible, at least in part, for the interaction between Sertoli and spermatogenic cells during mouse spermatogenesis.

TRIADIMEFON INDUCES RAT THYROID TUMORS THROUGH A NON-TSH MEDIATED MODE OF ACTION

EPA Science Inventory

Conazoles are a class of fungicides used as agricultural and pharmaceutical products which inhibit ergosterol biosynthesis. Members of this class are hepatotoxic and cause mouse hepatocellular tumors and/or rat thyroid follicular cell tumors. Triadimefon-induced rat thyroid tumor...

Development and testing of a mouse simulated space flight model

NASA Technical Reports Server (NTRS)

Sonnenfeld, Gerald

1987-01-01

The development and testing of a mouse model for simulating some aspects of weightlessness that occurs during space flight, and the carrying out of immunological experiments on animals undergoing space flight is examined. The mouse model developed was an antiorthostatic, hypokinetic, hypodynamic suspension model similar to one used with rats. The study was divided into two parts. The first involved determination of which immunological parameters should be observed on animals flown during space flight or studied in the suspension model. The second involved suspending mice and determining which of those immunological parameters were altered by the suspension. Rats that were actually flown in Space Shuttle SL-3 were used to test the hypotheses.

Investigation of a thiazolidinone derivative as an allosteric modulator of follicle stimulating hormone receptor: evidence for its ability to support follicular development and ovulation.

PubMed

Sriraman, Venkataraman; Denis, Deborah; de Matos, Daniel; Yu, Henry; Palmer, Stephen; Nataraja, Selva

2014-05-15

FSH signalling through its cognate receptor is critical for follicular development and ovulation. An earlier study had documented thiazolidinone derivatives to activate FSH receptor expressed in CHO cells and rat granulosa cells; however development of this compound for clinical use was halted for unobvious reasons. The objective of the current study is to extend the previous investigations in detail on the ability of thiazolidinone derivative (henceforth referred to as Compound 5) to activate FSH signalling and learn the barriers that preclude development of this derivative for clinical purposes. Our results demonstrate that the Compound 5 in a dose-dependent manner stimulated cAMP production, activated AKT and ERK signalling pathways and induced estradiol production in cultured rat granulosa cells. Compound 5 also caused dose-dependent increase in estradiol production from human granulosa cells. In increasingly more complex in vitro systems, Compound 5 was able to induce the expansion of mouse cumulus-oocyte-complex and support in vitro development of mouse preantral follicle to preovulatory stage and release of oocyte from the follicle. In vivo, the compound stimulated preovulatory follicular development and ovulation in immature rats. Pharmacokinetic and safety investigations reveal poor oral availability and genotoxicity. Together, our results document Compound 5 to act as a FSHR allosteric modulator but have poor pharmacological properties for development of an oral FSH receptor modulator. Copyright © 2014 Elsevier Inc. All rights reserved.

A novel small-molecule thienoquinolin urea transporter inhibitor acts as a potential diuretic.

PubMed

Li, Fei; Lei, Tianluo; Zhu, Juanjuan; Wang, Weiling; Sun, Yi; Chen, Jihui; Dong, Zixun; Zhou, Hong; Yang, Baoxue

2013-06-01

Urea transporters (UTs) are a family of membrane channel proteins that are specifically permeable to urea and play an important role in intrarenal urea recycling and in urine concentration. Using an erythrocyte osmotic lysis assay, we screened a small-molecule library for inhibitors of UT-facilitated urea transport. A novel class of thienoquinolin UT-B inhibitors were identified, of which PU-14 had potent inhibition activity on human, rabbit, rat, and mouse UT-B. The half-maximal inhibitory concentration of PU-14 on rat UT-B-mediated urea transport was ∼0.8 μmol/l, and it did not affect urea transport in mouse erythrocytes lacking UT-B but inhibited UT-A-type urea transporters, with 36% inhibition at 4 μmol/l. PU-14 showed no significant cellular toxicity at concentrations up to its solubility limit of 80 μmol/l. Subcutaneous delivery of PU-14 (at 12.5, 50, and 100 mg/kg) to rats caused an increase of urine output and a decrease of the urine urea concentration and subsequent osmolality without electrolyte disturbances and liver or renal damages. This suggests that PU-14 has a diuretic effect by urea-selective diuresis. Thus, PU-14 or its analogs might be developed as a new diuretic to increase renal fluid clearance in diseases associated with water retention without causing electrolyte imbalance. PU-14 may establish 'chemical knockout' animal models to study the physiological functions of UTs.

Knockout of the aryl hydrocarbon receptor results in distinct hepatic and renal phenotypes in rats and mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harrill, Joshua A.; Hukkanen, Renee R.; Lawson, Marie

2013-10-15

The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor which plays a role in the development of multiple tissues and is activated by a large number of ligands, including 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In order to examine the roles of the AHR in both normal biological development and response to environmental chemicals, an AHR knockout (AHR-KO) rat model was created and compared with an existing AHR-KO mouse. AHR-KO rats harboring either 2-bp or 29-bp deletion mutation in exon 2 of the AHR were created on the Sprague–Dawley genetic background using zinc-finger nuclease (ZFN) technology. Rats harboring either mutation type lacked expressionmore » of AHR protein in the liver. AHR-KO rats were also insensitive to thymic involution, increased hepatic weight and the induction of AHR-responsive genes (Cyp1a1, Cyp1a2, Cyp1b1, Ahrr) following acute exposure to 25 μg/kg TCDD. AHR-KO rats had lower basal expression of transcripts for these genes and also accumulated ∼ 30–45-fold less TCDD in the liver at 7 days post-exposure. In untreated animals, AHR-KO mice, but not AHR-KO rats, had alterations in serum analytes indicative of compromised hepatic function, patent ductus venosus of the liver and persistent hyaloid arteries in the eye. AHR-KO rats, but not AHR-KO mice, displayed pathological alterations to the urinary tract: bilateral renal dilation (hydronephrosis), secondary medullary tubular and uroepithelial degenerative changes and bilateral ureter dilation (hydroureter). The present data indicate that the AHR may play significantly different roles in tissue development and homeostasis and toxicity across rodent species. - Highlights: • An AHR knockout rat was generated on a Sprague–Dawley outbred background. • AHR-KO rats lack expression of AHR protein. • AHR-KO rats are insensitive to TCDD-mediated effects. • Data suggests difference in the role of AHR in tissue development of rats and mice. • Abnormalities in vascular development are observed in AHR-KO mouse, but not rat. • Renal pathology is observed in AHR-KO rat, but not mouse.« less

cDNA cloning of the murine PEX gene implicated in X-linked hypophosphatemia and evidence for expression in bone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Du, L.; Desbarats, M.; Viel, J.

1996-08-15

The recently identified human PEX g ene apparently encodes for a neutral endopeptidase that is mutated in patients with X-linked hypophosphatemia. The 3{prime} and 5{prime} ends of the coding region of PEX have not been cloned, nor has the tissue expression of the gene been identified. Here we report the isolation and characterization of the complete open reading frame of the mouse Pex gene and the demonstration of its expression in bone. Mouse Pex cDNA is predicted to encode a protein of 749 amino acids with 95% identity to the available human PEX sequence and significant homology to members ofmore » the membrane-bound metalloendopeptidase family. Northern blot analysis revealed a 6.6-kb transcript in bone and in cultured osteoblasts from normal mice that was not detectable in samples from the Hyp mouse, the murine homolog of human X-linked hypophosphatemia. Pex transcripts were, however, detectable in Hyp bone by RT-PCR amplification. Of particular interest, a cDNA clone from rat incisor shows 93% sequence identity to the 5{prime} end of Pex cDNA, suggesting that Pex may be expressed in another calcified tissue, the tooth. The association of impaired mineralization of bone and teeth and disturbed renal phosphate reabsorption with altered expression of Pex suggests that the Pex gene product may play a critical role in these processes. 47 refs., 2 figs., 1 tab.« less

A conserved gene family encodes transmembrane proteins with fibronectin, immunoglobulin and leucine-rich repeat domains (FIGLER)

PubMed Central

Munfus, Delicia L; Haga, Christopher L; Burrows, Peter D; Cooper, Max D

2007-01-01

Background In mouse the cytokine interleukin-7 (IL-7) is required for generation of B lymphocytes, but human IL-7 does not appear to have this function. A bioinformatics approach was therefore used to identify IL-7 receptor related genes in the hope of identifying the elusive human cytokine. Results Our database search identified a family of nine gene candidates, which we have provisionally named fibronectin immunoglobulin leucine-rich repeat (FIGLER). The FIGLER 1–9 genes are predicted to encode type I transmembrane glycoproteins with 6–12 leucine-rich repeats (LRR), a C2 type Ig domain, a fibronectin type III domain, a hydrophobic transmembrane domain, and a cytoplasmic domain containing one to four tyrosine residues. Members of this multichromosomal gene family possess 20–47% overall amino acid identity and are differentially expressed in cell lines and primary hematopoietic lineage cells. Genes for FIGLER homologs were identified in macaque, orangutan, chimpanzee, mouse, rat, dog, chicken, toad, and puffer fish databases. The non-human FIGLER homologs share 38–99% overall amino acid identity with their human counterpart. Conclusion The extracellular domain structure and absence of recognizable cytoplasmic signaling motifs in members of the highly conserved FIGLER gene family suggest a trophic or cell adhesion function for these molecules. PMID:17854505

Cu,Zn-superoxide dismutase is lower and copper chaperone CCS is higher in erythrocytes of copper-deficient rats and mice.

PubMed

West, Elizabeth C; Prohaska, Joseph R

2004-09-01

Discovery of a sensitive blood biochemical marker of copper status would be valuable for assessing marginal copper intakes. Rodent models were used to investigate whether erythrocyte concentrations of copper,zinc-superoxide dismutase (SOD), and the copper metallochaperone for SOD (CCS) were sensitive to dietary copper changes. Several models of copper deficiency were studied in postweanling male Holtzman rats, male Swiss Webster mice offspring, and both rat and mouse dams. Treatment resulted in variable but significantly altered copper status as evaluated by the presence of anemia, and lower liver copper and higher liver iron concentrations in copper-deficient compared with copper-adequate animals. Associated with this copper deficiency were consistent reductions in immunoreactive SOD and robust enhancements in CCS. In most cases, the ratio of CCS:SOD was several-fold higher in red blood cell extracts from copper-deficient compared with copper-adequate rodents. Determination of red cell CCS:SOD may be useful for assessing copper status of humans.

Magmas Overexpression Inhibits Staurosporine Induced Apoptosis in Rat Pituitary Adenoma Cell Lines

PubMed Central

Gentilin, Erica; Minoia, Mariella; Molè, Daniela; delgi Uberti, Ettore C.; Zatelli, Maria Chiara

2013-01-01

Magmas is a nuclear gene that encodes for the mitochondrial import inner membrane translocase subunit Tim16. Magmas is overexpressed in the majority of human pituitary adenomas and in a mouse ACTH-secreting pituitary adenoma cell line. Here we report that Magmas is highly expressed in two out of four rat pituitary adenoma cell lines and its expression levels inversely correlate to the extent of cellular response to staurosporine in terms of apoptosis activation and cell viability. Magmas over-expression in rat GH/PRL-secreting pituitary adenoma GH4C1 cells leads to an increase in cell viability and to a reduction in staurosporine-induced apoptosis and DNA fragmentation, in parallel with the increase in Magmas protein expression. These results indicate that Magmas plays a pivotal role in response to pro-apoptotic stimuli and confirm and extend the finding that Magmas protects pituitary cells from staurosporine-induced apoptosis, suggesting its possible involvement in pituitary adenoma development. PMID:24069394

The Multidrug Resistance 1 Gene Abcb1 in Brain and Placenta: Comparative Analysis in Human and Guinea Pig

PubMed Central

Pappas, Jane J.; Petropoulos, Sophie; Suderman, Matthew; Iqbal, Majid; Moisiadis, Vasilis; Turecki, Gustavo; Matthews, Stephen G.; Szyf, Moshe

2014-01-01

The Multidrug Resistance 1 (MDR1; alternatively ABCB1) gene product P-glycoprotein (P-gp), an ATP binding cassette transporter, extrudes multiple endogenous and exogenous substrates from the cell, playing an important role in normal physiology and xenobiotic distribution and bioavailability. To date, the predominant animal models used to investigate the role of P-gp have been the mouse and rat, which have two distinct genes, Abcb1a and Abcb1b. In contrast, the human has a single gene, ABCB1, for which only a single isoform has been validated. We and others have previously shown important differences between Abcb1a and Abcb1b, limiting the extrapolation from rodent findings to the human. Since the guinea pig has a relatively long gestation, hemomonochorial placentation and neuroanatomically mature offspring, it is more similar to the human, and may provide a more comparable model for investigating the regulation of P-gp in the brain and placenta, however, to date, the Abcb1 gene in the guinea pig remains to be characterized. The placenta and fetal brain are barrier sites that express P-gp and that play a critical role of protection of the fetus and the fetal brain from maternally administered drugs and other xenobiotics. Using RNA sequencing (RNA-seq), reverse transcription-polymerase chain reaction (RT-PCR) and quantitative PCR (QPCR) to sequence the expressed isoforms of guinea pig Abcb1, we demonstrate that like the human, the guinea pig genome contains one gene for Abcb1 but that it is expressed as at least three different isoforms via alternative splicing and alternate exon usage. Further, we demonstrate that these isoforms are more closely related to human than to rat or mouse isoforms. This striking, overall similarity and evolutionary relatedness between guinea pig Abcb1 and human ABCB1 indicate that the guinea pig represents a relevant animal model for investigating the function and regulation of P-gp in the placenta and brain. PMID:25353162

Proceedings of the 2013 National Toxicology Program Satellite Symposium

PubMed Central

Elmore, Susan A.; Boyle, Michael C.; Boyle, Molly H.; Cora, Michelle C.; Crabbs, Torrie A.; Cummings, Connie A.; Gruebbel, Margarita M.; Johnson, Crystal L.; Malarkey, David E.; McInnes, Elizabeth F.; Nolte, Thomas; Shackelford, Cynthia C.; Ward, Jerrold M.

2014-01-01

The 2013 annual National Toxicology Program (NTP) Satellite Symposium, entitled “Pathology Potpourri” was held in Portland, Oregon in advance of the Society of Toxicologic Pathology's 32nd annual meeting. The goal of the NTP Symposium is to present current diagnostic pathology or nomenclature issues to the toxicologic pathology community. This article presents summaries of the speakers' presentations, including diagnostic or nomenclature issues that were presented, along with select images that were used for audience voting and discussion. Some lesions and topics covered during the symposium included a caudal tail vertebra duplication in mice; nephroblastematosis in rats; ectopic C cell tumor in a hamster; granular cell aggregates/tumor in the uterus of a hamster; Pneumocystis carinii in the lung of a rat; iatrogenic chronic inflammation in the lungs of control rats; hepatoblastoma arising within an adenoma in a mouse; humoral hypercalcemia of benignancy in a transgenic mouse; acetaminophen induced hepatoxicity in rats; electron microscopy images of iatrogenic intraerythrocytic inclusions in transgenic mice; questionable hepatocellular degeneration/cell death/artifact in rats; atypical endometrial hyperplasia in rats; malignant mixed Müllerian tumors/carcinosarcomas in rats; differential diagnoses of proliferative lesions the intestine of rodents; and finally obstructive nephropathy caused by melamine poisoning in a rat. PMID:24334674

Airway disease phenotypes in animal models of cystic fibrosis.

PubMed

McCarron, Alexandra; Donnelley, Martin; Parsons, David

2018-04-02

In humans, cystic fibrosis (CF) lung disease is characterised by chronic infection, inflammation, airway remodelling, and mucus obstruction. A lack of pulmonary manifestations in CF mouse models has hindered investigations of airway disease pathogenesis, as well as the development and testing of potential therapeutics. However, recently generated CF animal models including rat, ferret and pig models demonstrate a range of well characterised lung disease phenotypes with varying degrees of severity. This review discusses the airway phenotypes of currently available CF animal models and presents potential applications of each model in airway-related CF research.

Taxonomy, genetic organization, and life cycle of Pneumocystis carinii.

PubMed

Cushion, M T

1998-12-01

Pneumocystis carinii was initially misidentified as a protozoan parasite. Recent molecular and biochemical analyses provide unequivocal evidence for placement of P. carinii with the fungi, and that P. carinii is most likely an ascomycete. Genetic investigations further show that P. carinii derived from different mammalian hosts (human, rat, mouse, and ferret) exhibit considerable chromosomal and gene sequence divergence indicating that they are likely of different species. The life cycle of P. carinii has not been definitively established, but available evidence is reviewed in light of classification of this organism as a fungus.

Imaging Tumor Cell Movement In Vivo

PubMed Central

Entenberg, David; Kedrin, Dmitriy; Wyckoff, Jeffrey; Sahai, Erik; Condeelis, John; Segall, Jeffrey E.

2013-01-01

This unit describes the methods that we have been developing for analyzing tumor cell motility in mouse and rat models of breast cancer metastasis. Rodents are commonly used both to provide a mammalian system for studying human tumor cells (as xenografts in immunocompromised mice) as well as for following the development of tumors from a specific tissue type in transgenic lines. The Basic Protocol in this unit describes the standard methods used for generation of mammary tumors and imaging them. Additional protocols for labeling macrophages, blood vessel imaging, and image analysis are also included. PMID:23456602

Ups and Downs of Viagra: Revisiting Ototoxicity in the Mouse Model

PubMed Central

Au, Adrian; Stuyt, John Gerka; Chen, Daniel; Alagramam, Kumar

2013-01-01

Sildenafil citrate (Viagra), a phosphodiesterase 5 inhibitor (PDE5i), is a commonly prescribed drug for erectile dysfunction. Since the introduction of Viagra in 1997, several case reports have linked Viagra to sudden sensorineural hearing loss. However, these studies are not well controlled for confounding factors, such as age and noise-induced hearing loss and none of these reports are based on prospective double-blind studies. Further, animal studies report contradictory data. For example, one study (2008) reported hearing loss in rats after long-term and high-dose exposure to sildenafil citrate. The other study (2012) showed vardenafil, another formulation of PDE5i, to be protective against noise-induced hearing loss in mice and rats. Whether or not clinically relevant doses of sildenafil citrate cause hearing loss in normal subjects (animals or humans) is controversial. One possibility is that PDE5i exacerbates age-related susceptibility to hearing loss in adults. Therefore, we tested sildenafil citrate in C57BL/6J, a strain of mice that displays increased susceptibility to age-related hearing loss, and compared the results to those obtained from the FVB/N, a strain of mice with no predisposition to hearing loss. Six-week-old mice were injected with the maximum tolerated dose of sildenafil citrate (10 mg/kg/day) or saline for 30 days. Auditory brainstem responses (ABRs) were recorded pre- and post injection time points to assess hearing loss. Entry of sildenafil citrate in the mouse cochlea was confirmed by qRT-PCR analysis of a downstream target of the cGMP-PKG cascade. ABR data indicated no statistically significant difference in hearing between treated and untreated mice in both backgrounds. Results show that the maximum tolerated dose of sildenafil citrate administered daily for 4 weeks does not affect hearing in the mouse. Our study gives no indication that Viagra will negatively impact hearing and it emphasizes the need to revisit the issue of Viagra related ototoxicity in humans. PMID:24244454

Ups and downs of Viagra: revisiting ototoxicity in the mouse model.

PubMed

Au, Adrian; Stuyt, John Gerka; Chen, Daniel; Alagramam, Kumar

2013-01-01

Sildenafil citrate (Viagra), a phosphodiesterase 5 inhibitor (PDE5i), is a commonly prescribed drug for erectile dysfunction. Since the introduction of Viagra in 1997, several case reports have linked Viagra to sudden sensorineural hearing loss. However, these studies are not well controlled for confounding factors, such as age and noise-induced hearing loss and none of these reports are based on prospective double-blind studies. Further, animal studies report contradictory data. For example, one study (2008) reported hearing loss in rats after long-term and high-dose exposure to sildenafil citrate. The other study (2012) showed vardenafil, another formulation of PDE5i, to be protective against noise-induced hearing loss in mice and rats. Whether or not clinically relevant doses of sildenafil citrate cause hearing loss in normal subjects (animals or humans) is controversial. One possibility is that PDE5i exacerbates age-related susceptibility to hearing loss in adults. Therefore, we tested sildenafil citrate in C57BL/6J, a strain of mice that displays increased susceptibility to age-related hearing loss, and compared the results to those obtained from the FVB/N, a strain of mice with no predisposition to hearing loss. Six-week-old mice were injected with the maximum tolerated dose of sildenafil citrate (10 mg/kg/day) or saline for 30 days. Auditory brainstem responses (ABRs) were recorded pre- and post injection time points to assess hearing loss. Entry of sildenafil citrate in the mouse cochlea was confirmed by qRT-PCR analysis of a downstream target of the cGMP-PKG cascade. ABR data indicated no statistically significant difference in hearing between treated and untreated mice in both backgrounds. Results show that the maximum tolerated dose of sildenafil citrate administered daily for 4 weeks does not affect hearing in the mouse. Our study gives no indication that Viagra will negatively impact hearing and it emphasizes the need to revisit the issue of Viagra related ototoxicity in humans.

Borders and Comparative Cytoarchitecture of the Perirhinal and Postrhinal Cortices in an F1 Hybrid Mouse

PubMed Central

Beaudin, Stephane A.; Singh, Teghpal; Agster, Kara L.

2013-01-01

We examined the cytoarchitectonic and chemoarchitectonic organization of the cortical regions associated with the posterior rhinal fissure in the mouse brain, within the framework of what is known about these regions in the rat. Primary observations were in a first-generation hybrid mouse line, B6129PF/J1. The F1 hybrid was chosen because of the many advantages afforded in the study of the molecular and cellular bases of learning and memory. Comparisons with the parent strains, the C57BL6/J and 129P3/J are also reported. Mouse brain tissue was processed for visualization of Nissl material, myelin, acetyl cholinesterase, parvalbumin, and heavy metals. Tissue stained for heavy metals by the Timm’s method was particularly useful in the assignment of borders and in the comparative analyses because the patterns of staining were similar across species and strains. As in the rat, the areas examined were parcellated into 2 regions, the perirhinal and the postrhinal cortices. The perirhinal cortex was divided into areas 35 and 36, and the postrhinal cortex was divided into dorsal (PORd) and ventral (PORv) subregions. In addition to identifying the borders of the perirhinal cortex, we were able to identify a region in the mouse brain that shares signature features with the rat postrhinal cortex. PMID:22368084

Exon 11 skipping of SCN10A coding for voltage-gated sodium channels in dorsal root ganglia

PubMed Central

Schirmeyer, Jana; Szafranski, Karol; Leipold, Enrico; Mawrin, Christian; Platzer, Matthias; Heinemann, Stefan H

2014-01-01

The voltage-gated sodium channel NaV1.8 (encoded by SCN10A) is predominantly expressed in dorsal root ganglia (DRG) and plays a critical role in pain perception. We analyzed SCN10A transcripts isolated from human DRGs using deep sequencing and found a novel splice variant lacking exon 11, which codes for 98 amino acids of the domain I/II linker. Quantitative PCR analysis revealed an abundance of this variant of up to 5–10% in human, while no such variants were detected in mouse or rat. Since no obvious functional differences between channels with and without the exon-11 sequence were detected, it is suggested that SCN10A exon 11 skipping in humans is a tolerated event. PMID:24763188

Transplanting Human Skin Grafts onto Nude Mice to Model Skin Scars.

PubMed

Ding, Jie; Tredget, Edward E

2017-01-01

Hypertrophic scar (HTS) is a common outcome of deep dermal wound healing mainly followed mechanical, chemical, and thermal injuries in the skin. Because of the lack of the most effective prevention and treatment, it is particularly important to establish an ideal dermal animal model for improving the understanding of the pathogenesis and exploring therapeutic approaches of HTS. Compared to other dermal fibrotic animal models in rabbits, red Duroc pigs, guinea pigs, rats, and mice, the approach that uses normal human split-thickness skin grafted onto nude or other immunodeficient mice which develop scars that resemble human HTS offers the advantages of lower cost, easier manipulation, and shorter research period. In this chapter, we will introduce the detailed procedures to create the ideal dermal fibrotic mouse model.

Glutamatergic and Dopaminergic Neurons in the Mouse Ventral Tegmental Area

PubMed Central

Yamaguchi, Tsuyoshi; Qi, Jia; Wang, Hui-Ling; Zhang, Shiliang; Morales, Marisela

2014-01-01

The ventral tegmental area (VTA) comprises dopamine (DA), GABA and glutamate (Glu) neurons. Some rat VTA Glu neurons, expressing vesicular glutamate transporter 2 (VGluT2), co-express tyrosine hydroxylase (TH). While transgenic mice are now being used in attempts to determine the role of VGluT2/TH neurons in reward and neuronal signaling, such neurons have not been characterized in mouse tissue. By cellular detection of VGluT2-mRNA and TH-immunoreactivity (TH-IR), we determined the cellular expression of VGluT2-mRNA within VTA TH-IR neurons in the mouse. We found that some mouse VGluT2 neurons co-expressed TH-IR, but their frequency was lower than in the rat. To determine whether low expression of TH mRNA or TH-IR accounts for this low frequency, we evaluated VTA cellular co-expression of TH-transcripts and TH-protein. Within the medial aspects of the VTA, some neurons expressed TH mRNA but lacked TH-IR; among them a subset co-expressed VGluT2 mRNA. To determine if lack of VTA TH-IR was due to TH trafficking, we tagged VTA TH neurons by cre-inducible expression of mCherry in TH::Cre mice. By dual immunofluorescence, we detected axons containing mCherry, but lacking TH-IR, in the lateral habenula, indicating that mouse low frequency of VGluT2 mRNA (+)/TH-IR (+) neurons is due to lack of synthesis of TH protein, rather than TH-protein trafficking. In conclusion, VGluT2 neurons are present in the rat and mouse VTA, but they differ in the populations of VGluT2/TH and TH neurons. We reveal that under normal conditions, the translation of TH protein is suppressed in the mouse mesohabenular TH neurons. PMID:25572002

Anatomical Variation of the Tarsus in Common Inbred Mouse Strains.

PubMed

Richbourg, Heather A; Martin, Matthew J; Schachner, Emma R; McNulty, Margaret A

2017-03-01

Rodent models are used for a variety of orthopedic research applications; however, anatomy references include mostly artistic representations. Advanced imaging techniques, including micro-computed tomography (microCT), can provide more accurate representations of subtle anatomical characteristics. A recent microCT atlas of laboratory mouse (Mus musculus) anatomy depicts the central and tarsal bone III (T3) as a single bone, differing from previous references. Fusion of tarsal bones is generally characterized as pathological secondary to mutations associated with growth factors, and normal variation has not been documented in the mouse tarsus. Therefore, it is unclear if this fusion is a normal or a pathological characteristic. The aim of this study is to characterize the tarsus of the laboratory mouse and compare it to the rat and selected outgroup species (i.e., white-footed mouse) via microCT and histology to determine if the central and T3 are separate or fused into a single bone. Laboratory mice (C57/Bl6 [n = 17] and BalbC [n = 2]) and rats (n = 5) were scanned with microCT. A representative laboratory mouse from each strain was evaluated histologically via serial sagittal sections through the mid-tarsus. General pedal anatomy was similar between all species; however, the central and T3 bones were fused in all laboratory mice but not the rat or white-footed mouse. A band of hyaline cartilage was identified within the fused bone of the laboratory mice. We conclude that the fusion found is a normal characteristic in laboratory mice, but timing of the fusion remains ambiguous. Anat Rec, 300:450-459, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Role of cytochrome P450 IA2 in acetanilide 4-hydroxylation as determined with cDNA expression and monoclonal antibodies.

PubMed

Liu, G; Gelboin, H V; Myers, M J

1991-02-01

The role of P450 IA2 in the hydroxylation of acetanilide was examined using an inhibitory monoclonal antibody (MAb) 1-7-1 and vaccinia cDNA expression producing murine P450 IA1 (mIA1), murine P450 IA2 (mIA2), or human P450 IA2 (hIA2). Acetanilide hydroxylase (AcOH) activity was measured using an HPLC method with more than 500-fold greater sensitivity than previously described procedures. This method, which does not require the use of radioactive acetanilide, was achieved by optimizing both the gradient system and the amount of enzyme needed to achieve detection by uv light. MAb 1-7-1 inhibits up to 80% of the AcOH activity in both rat liver microsomes and cDNA expressed mouse and human P450 IA2. MAb 1-7-1, which recognizes both P450 IA1 and P450 IA2, completely inhibits the aryl hydrocarbon hydroxylase (AHH) activity of cDNA expressed in IA1. The inhibition of only 80% of the AHH activity present in MC liver microsomes by MAb 1-7-1 suggests that additional P450 forms are contributing to the overall AHH activity present in methylcholanthrene (MC)-liver microsomes as MAb 1-7-1 almost completely inhibits the AHH activity of expressed mIA1. Maximal inhibition of IA2 by 1-7-1 results in an 80% decrease in acetanilide hydroxylase activity in both liver microsomes and expressed mouse and human IA2. The capacity of MAb 1-7-1 to produce identical levels of inhibition of acetanilide hydroxylase activity in rat MC microsomes (80%) and in expressed mouse (81%) and human P450 IA2 (80%) strongly suggests that P450 IA2 is the major and perhaps the only enzyme responsible for the metabolism of acetanilide. These results demonstrate the complementary utility of monoclonal antibodies and cDNA expression for defining the contribution of specific P450 enzymes to the metabolism of a given substrate. This complementary approach allows for a more precise determination of the inhibitory capacity of MAb with respect to the metabolic capacity of the target P450.

Expression profile and distribution of Efhc1 gene transcript during rodent brain development.

PubMed

Conte, Fábio F; Ribeiro, Patrícia A O; Marchesini, Rafael B; Pascoal, Vinícius D B; Silva, Joelcimar M; Oliveira, Amanda R; Gilioli, Rovílson; Sbragia, Lourenço; Bittencourt, Jackson C; Lopes-Cendes, Iscia

2009-09-01

One of the putative causative genes for juvenile myoclonic epilepsy (JME) is EFHC1. We report here the expression profile and distribution of Efhc1 messenger RNA (mRNA) during mouse and rat brain development. Real-time polymerase chain reaction revealed that there is no difference in the expression of Efhc1 mRNA between right and left hemispheres in both species. In addition, the highest levels of Efhc1 mRNA were found at intra-uterine stages in mouse and in adulthood in rat. In common, there was a progressive decrease in Efhc1 expression from 1-day-old neonates to 14-day-old animals in both species. In situ hybridization studies showed that rat and mouse Efhc1 mRNAs are expressed in ependymal cells of ventricle walls. Our findings suggest that Efhc1 expression is more important during initial phases of brain development and that at this stage it could be involved in key developmental mechanisms underlying JME.

Identification of allantoin, uric acid, and indoxyl sulfate as biochemical indicators of filth in food packaging by LC.

PubMed

Carlson, M; Thompson, R D

2001-01-01

A liquid chromatographic (LC) method was developed for the determination of allantoin, uric acid, and indoxyl sulfate in mammalian urine contaminated packaging material including paper bagging, corrugated cardboard, grayboard, and burlap bagging. The procedure involves solvent extraction and isolation of the 3 analytes by reversed-phase LC with ultraviolet detection at 225 nm for allantoin and 286 nm for uric acid and indoxyl sulfate. The composition of authentic mammalian urine such as mouse, rat, cat, dog, and human were also determined with regard to the 3 compounds of interest. A linear concentration range of 0.11-20.4, 0.02-10.0, and 0.04-30.0 microg/mL was obtained for allantoin, uric acid, and indoxyl sulfate, respectively. Limits of detection (LOD) and quantitation (LOQ) were 0.0104 and 0.0345 microg/mL for allantoin; 0.0018 and 0.0060 microg/mL for uric acid; and 0.0049 and 0.0165 microg/mL for indoxyl sulfate, respectively. Interday relative standard deviation values for a mixture of standard allantoin, uric acid, and indoxyl sulfate (n = 5) were 0.97, 0.80, and 0.94%, respectively. Analyte composition for 5 types of authentic mammalian urine varied from 0.19-6.88 mg/mL allantoin; 0.08-0.57 mg/mL uric acid; and 0.03-0.78 mg/mL indoxyl sulfate. Analyte content for 8 samples including 2 samples each for paper, cardboard, grayboard, and burlap bagging each contaminated with mouse or rat urine ranged from

Hepatocyte nuclear factor-4alpha is a central transactivator of the mouse Ntcp gene.

PubMed

Geier, Andreas; Martin, Ina V; Dietrich, Christoph G; Balasubramaniyan, Natarajan; Strauch, Sonja; Suchy, Frederick J; Gartung, Carsten; Trautwein, Christian; Ananthanarayanan, Meenakshisundaram

2008-08-01

Sodium taurocholate cotransporting polypeptide (Ntcp) is the major uptake system for conjugated bile acids. Deletions of hepatocyte nuclear factor (HNF)-1alpha and retinoid X receptor-alpha:retinoic acid receptor-alpha binding sites in the mouse 5'-flanking region corresponding to putatively central regulatory elements of rat Ntcp do not significantly reduce promoter activity. We hypothesized that HNF-4alpha, which is increasingly recognized as a central regulator of hepatocyte function, may directly transactivate mouse (mNtcp). A 1.1-kb 5'-upstream region including the mouse Ntcp promoter was cloned and compared with the rat promoter. In contrast to a moderate 3.5-fold activation of mNtcp by HNF-1alpha, HNF-4alpha cotransfection led to a robust 20-fold activation. Deletion analysis of mouse and rat Ntcp promoters mapped a conserved HNF-4alpha consensus site at -345/-326 and -335/-316 bp, respectively. p-475bpmNtcpLUC is not transactivated by HNF-1alpha but shows a 50-fold enhanced activity upon cotransfection with HNF-4alpha. Gel mobility shift assays demonstrated a complex of the HNF-4alpha-element formed with liver nuclear extracts that was blocked by an HNF-4alpha specific antibody. HNF-4alpha binding was confirmed by chromatin immunoprecipitation. Using Hepa 1-6 cells, HNF-4alpha-knockdown resulted in a significant 95% reduction in NTCP mRNA. In conclusion, mouse Ntcp is regulated by HNF-4alpha via a conserved distal cis-element independently of HNF-1alpha.

Molokhia (Corchorus olitorius L.) extract suppresses transformation of the aryl hydrocarbon receptor induced by dioxins.

PubMed

Nishiumi, Shin; Yabushita, Yoshiyuki; Fukuda, Itsuko; Mukai, Rie; Yoshida, Ken-Ichi; Ashida, Hitoshi

2006-02-01

Dioxins enter the body mainly through diet and cause the various toxicological effects by binding to the cytosolic aryl hydrocarbon receptor (AhR) followed by its transformation. In recent reports, it has been shown that certain natural compounds suppress AhR transformation in vitro. In this study, we demonstrated that ethanolic extract from molokhia, known as Egyptian spinach, showed the strongest suppressive effect on AhR transformation induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in cell-free system using rat hepatic cytosol among 41 kinds of extracts from vegetables and fruits. The molokhia extract also suppressed TCDD-induced AhR transformation in mouse hepatoma Hepa-1c1c7 cells and in intestinal permeability system constructed with human colon adenocarcinoma Caco-2 cells and human hepatoma HepG2 cells. Moreover, oral administration of the molokhia extract (100mg/kg body weight) decreased 3-methylcholanthrene-induced AhR transformation to the control level by inhibiting translocation of the AhR from cytosol into the nucleus in the liver of rats. The molokhia extract-administered rat liver showed a tolerance to TCDD-induced AhR transformation by ex vivo experiment. These results indicate that molokhia is an attractive food for isolation and identification of a natural antagonist for the AhR.

GATE Simulations of Small Animal SPECT for Determination of Scatter Fraction as a Function of Object Size

NASA Astrophysics Data System (ADS)

Konik, Arda; Madsen, Mark T.; Sunderland, John J.

2012-10-01

In human emission tomography, combined PET/CT and SPECT/CT cameras provide accurate attenuation maps for sophisticated scatter and attenuation corrections. Having proven their potential, these scanners are being adapted for small animal imaging using similar correction approaches. However, attenuation and scatter effects in small animal imaging are substantially less than in human imaging. Hence, the value of sophisticated corrections is not obvious for small animal imaging considering the additional cost and complexity of these methods. In this study, using GATE Monte Carlo package, we simulated the Inveon small animal SPECT (single pinhole collimator) scanner to find the scatter fractions of various sizes of the NEMA-mouse (diameter: 2-5.5 cm , length: 7 cm), NEMA-rat (diameter: 3-5.5 cm, length: 15 cm) and MOBY (diameter: 2.1-5.5 cm, length: 3.5-9.1 cm) phantoms. The simulations were performed for three radionuclides commonly used in small animal SPECT studies:99mTc (140 keV), 111In (171 keV 90% and 245 keV 94%) and 125I (effective 27.5 keV). For the MOBY phantoms, the total Compton scatter fractions ranged (over the range of phantom sizes) from 4-10% for 99mTc (126-154 keV), 7-16% for 111In (154-188 keV), 3-7% for 111In (220-270 keV) and 17-30% for 125I (15-45 keV) including the scatter contributions from the tungsten collimator, lead shield and air (inside and outside the camera heads). For the NEMA-rat phantoms, the scatter fractions ranged from 10-15% (99mTc), 17-23% 111In: 154-188 keV), 8-12% (111In: 220-270 keV) and 32-40% (125I). Our results suggest that energy window methods based on solely emission data are sufficient for all mouse and most rat studies for 99mTc and 111In. However, more sophisticated methods may be needed for 125I.

The “curved lead pathway” method to enable a single lead to reach any two intracranial targets

NASA Astrophysics Data System (ADS)

Ding, Chen-Yu; Yu, Liang-Hong; Lin, Yuan-Xiang; Chen, Fan; Lin, Zhang-Ya; Kang, De-Zhi

2017-01-01

Deep brain stimulation is an effective way to treat movement disorders, and a powerful research tool for exploring brain functions. This report proposes a “curved lead pathway” method for lead implantation, such that a single lead can reach in sequence to any two intracranial targets. A new type of stereotaxic system for implanting a curved lead to the brain of human/primates was designed, the auxiliary device needed for this method to be used in rat/mouse was fabricated and verified in rat, and the Excel algorithm used for automatically calculating the necessary parameters was implemented. This “curved lead pathway” method of lead implantation may complement the current method, make lead implantation for multiple targets more convenient, and expand the experimental techniques of brain function research.

L-threo 3,4-dihydroxyphenylserine treatment during mouse perinatal and rat postnatal development does not alter the impact of dietary copper deficiency

PubMed Central

Pyatskowit, Joshua W.; Prohaska, Joseph R.

2009-01-01

Dietary copper (Cu) deficiency was induced perinatally in Swiss Albino mice and postnatally in male Holtzman rats to investigate the effect of L-threo 3,4-dihydroxyphenylserine (DOPS) on pup survival and catecholamine levels in a 2 × 2 factorial design. Mouse dams were placed on one of four treatments 14 days after mating and rats at postnatal day 19 (P19). Treatments were Cu-adequate (Cu +) and Cu-deficient (Cu −) diets with or without DOPS (1 mg/ml) in the drinking water. Mouse pups were killed at P14 and rats at P49. Mortality in Cu − pups was 46% and not significantly improved by DOPS, 39%. A repeat study with mice adding ascorbic acid in the water with DOPS showed no improvement. Compared to Cu + animals, Cu − animals were smaller, anemic and had a 92% reduction in liver Cu. DOPS treatment made no improvement to and in some cases exacerbated the Cu deficiency. Catecholamine levels measured in heart and brain by LCEC showed decreased NE levels and increased DA levels in Cu − animals compared to controls. DOPS treatment did not alter this pattern. Although DOPS was present in treated animal’s tissues, survival in mice and catecholamine levels in mice and rats were not altered by the 1 mg/ml dose of DOPS. PMID:16117185

Glycosaminoglycan Chain of Dentin Sialoprotein Proteoglycan

PubMed Central

Zhu, Q.; Sun, Y.; Prasad, M.; Wang, X.; Yamoah, A.K.; Li, Y.; Feng, J.; Qin, C.

2010-01-01

Dentin sialophosphoprotein (DSPP) is processed into dentin sialoprotein (DSP) and dentin phosphoprotein. A molecular variant of rat DSP, referred to as “HMW-DSP”, has been speculated to be a proteoglycan form of DSP. To determine if HMW-DSP is the proteoglycan form of DSP and to identify the glycosaminoglycan side-chain attachment site(s), we further characterized HMW-DSP. Chondroitinase ABC treatment reduced the migration rate for portions of rat HMW-DSP to the level of DSP. Disaccharide analysis showed that rat HMW-DSP contains glycosaminoglycan chains made of chondroitin-4-sulfate and has an average of 31-32 disaccharides/mol. These observations confirmed that HMW-DSP is the proteoglycan form of DSP (renamed “DSP-PG”). Edman degradation and mass spectrometric analyses of tryptic peptides from rat DSP-PG, along with substitution analyses of candidate Ser residues in mouse DSPP, confirmed that 2 glycosaminoglycan chains are attached to Ser241 and Ser253 in the rat, or Ser242 and Ser254 in the mouse DSPP sequence. PMID:20400719

Glycosaminoglycan chain of dentin sialoprotein proteoglycan.

PubMed

Zhu, Q; Sun, Y; Prasad, M; Wang, X; Yamoah, A K; Li, Y; Feng, J; Qin, C

2010-08-01

Dentin sialophosphoprotein (DSPP) is processed into dentin sialoprotein (DSP) and dentin phosphoprotein. A molecular variant of rat DSP, referred to as "HMW-DSP", has been speculated to be a proteoglycan form of DSP. To determine if HMW-DSP is the proteoglycan form of DSP and to identify the glycosaminoglycan side-chain attachment site(s), we further characterized HMW-DSP. Chondroitinase ABC treatment reduced the migration rate for portions of rat HMW-DSP to the level of DSP. Disaccharide analysis showed that rat HMW-DSP contains glycosaminoglycan chains made of chondroitin-4-sulfate and has an average of 31-32 disaccharides/mol. These observations confirmed that HMW-DSP is the proteoglycan form of DSP (renamed "DSP-PG"). Edman degradation and mass spectrometric analyses of tryptic peptides from rat DSP-PG, along with substitution analyses of candidate Ser residues in mouse DSPP, confirmed that 2 glycosaminoglycan chains are attached to Ser(241) and Ser(253) in the rat, or Ser(242) and Ser(254) in the mouse DSPP sequence.

EXPERIMENTAL USEFULNESS OF THE KANGAROO RAT

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haley, T.J.

1963-09-13

The kangaroo rat is readily tamed and has certain characteristics that make it unique and of interest in highly specialized research programs. Studies were conducted on its ability to exist on a dried diet with only a bare minimum of water and that obtained from succulent plants. Hematological studies indicate that the kangaroo rat exhibits a different hematological distribution of cells than the mouse or rat. The lymphocyte constitutes 81.4% of the total leokocytes. The hematocrit has a value of 46 to 48 in spite of the high degree of water conservation practiced by the animal. The response to ionizingmore » radiation of this species does not differ from that reported for the mouse or rat. Behavior studies indicate that the digging characteristics of the kangaroo rat are similar to those of the gerbil. Furthermore, the animal shows definite psychotic tendencies under the influence of psychotomimetics like LSD-25 and psilocybin. An evaluation of the physiological responses of isolated tissues from this animal as well as its responses to anesthetics is being undertaken to evaluate its further usefulness in the laboratory. (auth)« less

Species and interspecies radioimmunoassays for rat type C virus p30: interviral comparisons and assay of human tumor extracts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Charman, H.P.; White, M.H.; Rahman, R.

1976-01-01

The major internal protein, p30, of rat type C virus (RaLV) was purified and utilized to establish intra- and interspecies radioimmunoassays. Three rat viruses were compared in homologous and heterologous intraspecies assays with no evidence of type specificity. The only heterologous viruses to give inhibition in these species assays were the feline (FeLV) and hamster (HaLV) type C viruses; these reactions were incomplete and required high virus concentrations. An interspecies assay using a goat antiserum prepared after sequentially immunizing with FeLV, RD 114, and woolly monkey virus p30's and labeled RaLV p30 was inhibited by all mammalian type C viruses,more » although preferentially by RaLV, FeLV, and HaLV. Thus, as in a previously reported assay developed with HaLV p30, rat, hamster, and cat p30's seem more closely related to each other than to mouse type C virus p30. High levels of specific antigen were found in all cell lines producing rat virus, whereas embryonic tissues from several rat strains and cell lines considered virus-free based on other tests were negative for p30. Rats bearing tumors containing Moloney murine sarcoma virus (RaLV) did not contain free circulating antibody to RaLV p30. Fifty-one human tumor extracts (including two tumor cell lines) were tested for activity in the RaLV species and 47 in the interspecies assays after Sephadex gel filtration and pooling of material in the 15,000- to 40,000-molecular-weight range. At a sensitivity level of 7 ng/ml (0.7 ng/assay) in the interspecies assay, all human tissues, with one exception, were negative. The one positive result is considered nonspecific based on proteolysis of the labeled antigen. Input tissue protein of the purified tumor extracts averaged 1.9 mg/ml with a range of less than 0.025 to 22 mg/ml. Tissues from NIH Swiss mice processed in the same manner were positive in the interspecies assay but negative in the intraspecies RaLV assay. (auth)« less

Cloning and expression of the rat homologue of the Huntington disease gene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schmitt, I.; Epplen, J.T.; Riess, O.

1994-09-01

Huntington`s disease (HD) is an autosomal dominant neurodegenerative disorder which is manifested usually in adult life. The age of onset is variable and leads to progressive symptoms including involuntary choreatic movements and various cognitive and psychiatric disturbances. Recently, a gene (IT15) was cloned containing a (CAG){sub n} repeat which is elongated and unstable in HD patients. IT15 is widely expressed in human tissues but unrelated to any known deduced protein sequence. To further investigate the HD gene, 15 rat cDNA libraries were screened. 24 clones have been identified covering the Huntingtin gene. Comparison of the Huntingtin gene between human andmore » rat revealed homologies between 80% and 87% at the DNA level and about 90% at the protein level. These analyses will help to define biologically important sequence regions, e.g., via evolutionary conservation. One clone contains the (CAG){sub n} repeat which consists of eight triplets compared to seven triplets in the mouse and a median of 17 in human. As in humans there are two transcripts arising from differential 3{prime}-polyadenylation. In the 3{prime}UTR a stretch of about 280 bp is exchanged for a 250 bp fragment with no homology in rodents and man. The cDNA clones are currently used to study Huntingtin gene expression during development in rodent tissues. RNA in situ hybridization of embryonic sections shows predominant signals in all neuronal tissues. In contrast to previously published data Huntingtin mRNA expression in testis is increased in spermatocytes vs. spermatogonia.« less

Experimental Diabetes Mellitus in Different Animal Models

PubMed Central

Al-awar, Amin; Veszelka, Médea; Szűcs, Gergő; Attieh, Zouhair; Murlasits, Zsolt; Török, Szilvia; Pósa, Anikó; Varga, Csaba

2016-01-01

Animal models have historically played a critical role in the exploration and characterization of disease pathophysiology and target identification and in the evaluation of novel therapeutic agents and treatments in vivo. Diabetes mellitus disease, commonly known as diabetes, is a group of metabolic disorders characterized by high blood glucose levels for a prolonged time. To avoid late complications of diabetes and related costs, primary prevention and early treatment are therefore necessary. Due to its chronic symptoms, new treatment strategies need to be developed, because of the limited effectiveness of the current therapies. We overviewed the pathophysiological features of diabetes in relation to its complications in type 1 and type 2 mice along with rat models, including Zucker Diabetic Fatty (ZDF) rats, BB rats, LEW 1AR1/-iddm rats, Goto-Kakizaki rats, chemically induced diabetic models, and Nonobese Diabetic mouse, and Akita mice model. The advantages and disadvantages that these models comprise were also addressed in this review. This paper briefly reviews the wide pathophysiological and molecular mechanisms associated with type 1 and type 2 diabetes, particularly focusing on the challenges associated with the evaluation and predictive validation of these models as ideal animal models for preclinical assessments and discovering new drugs and therapeutic agents for translational application in humans. PMID:27595114

The Disease Portals, disease-gene annotation and the RGD disease ontology at the Rat Genome Database.

PubMed

Hayman, G Thomas; Laulederkind, Stanley J F; Smith, Jennifer R; Wang, Shur-Jen; Petri, Victoria; Nigam, Rajni; Tutaj, Marek; De Pons, Jeff; Dwinell, Melinda R; Shimoyama, Mary

2016-01-01

The Rat Genome Database (RGD;http://rgd.mcw.edu/) provides critical datasets and software tools to a diverse community of rat and non-rat researchers worldwide. To meet the needs of the many users whose research is disease oriented, RGD has created a series of Disease Portals and has prioritized its curation efforts on the datasets important to understanding the mechanisms of various diseases. Gene-disease relationships for three species, rat, human and mouse, are annotated to capture biomarkers, genetic associations, molecular mechanisms and therapeutic targets. To generate gene-disease annotations more effectively and in greater detail, RGD initially adopted the MEDIC disease vocabulary from the Comparative Toxicogenomics Database and adapted it for use by expanding this framework with the addition of over 1000 terms to create the RGD Disease Ontology (RDO). The RDO provides the foundation for, at present, 10 comprehensive disease area-related dataset and analysis platforms at RGD, the Disease Portals. Two major disease areas are the focus of data acquisition and curation efforts each year, leading to the release of the related Disease Portals. Collaborative efforts to realize a more robust disease ontology are underway. Database URL:http://rgd.mcw.edu. © The Author(s) 2016. Published by Oxford University Press.

Liquid Chromatography Combined with Mass Spectrometry Utilising High-Resolution, Exact Mass, and Multi-Stage Fragmentation for the Identification of Oxysterols in Rat Brain

PubMed Central

Karu, Kersti; Hornshaw, Martin; Woffendin, Gary; Bodin, Karl; Hamberg, Mats; Alvelius, Gunvor; Sjövall, Jan; Turton, John; Wang, Yuqin; Griffiths, William J.

2008-01-01

In man the brain accounts for about 20% of the body's free cholesterol, most of which is synthesised de novo in brain. To maintain cholesterol balance throughout life, cholesterol becomes metabolised to 24S-hydroxycholesterol principally in neurons. In mouse, rat, and probably human, metabolism to 24S-hydroxycholesterol accounts for about 50% of cholesterol turnover, however, the route by which the remainder is turned over has yet to be elucidated. Here we describe a novel liquid chromatography (LC) – multi-stage fragmentation mass spectrometry (MSn) methodology for the identification, with high sensitivity (low pg), of cholesterol metabolites in rat brain. The methodology includes derivatisation to enhance ionisation, exact mass analysis at high-resolution to identify potential metabolites, and LC-MS3 to allow their characterisation. 24S-Hydroxycholesterol was confirmed as a major oxysterol in rat brain, while other oxysterols identified for the first time in brain included 24,25-, 24,27-, 25,27-, 6,24, 7α,25-, and 7α,27-dihydroxycholesterols. In addition, 3β-hydroxy-5-oxo-5,6-secocholestan-6-al and its aldol, two molecules linked to amyloidogenesis of proteins, were characterised in rat brain. PMID:17251593

The endogenous and reactive depression subtypes revisited: integrative animal and human studies implicate multiple distinct molecular mechanisms underlying major depressive disorder

PubMed Central

2014-01-01

Background Traditional diagnoses of major depressive disorder (MDD) suggested that the presence or absence of stress prior to onset results in either ‘reactive’ or ‘endogenous’ subtypes of the disorder, respectively. Several lines of research suggest that the biological underpinnings of ‘reactive’ or ‘endogenous’ subtypes may also differ, resulting in differential response to treatment. We investigated this hypothesis by comparing the gene-expression profiles of three animal models of ‘reactive’ and ‘endogenous’ depression. We then translated these findings to clinical samples using a human post-mortem mRNA study. Methods Affymetrix mouse whole-genome oligonucleotide arrays were used to measure gene expression from hippocampal tissues of 144 mice from the Genome-based Therapeutic Drugs for Depression (GENDEP) project. The study used four inbred mouse strains and two depressogenic ‘stress’ protocols (maternal separation and Unpredictable Chronic Mild Stress) to model ‘reactive’ depression. Stress-related mRNA differences in mouse were compared with a parallel mRNA study using Flinders Sensitive and Resistant rat lines as a model of ‘endogenous’ depression. Convergent genes differentially expressed across the animal studies were used to inform candidate gene selection in a human mRNA post-mortem case control study from the Stanley Brain Consortium. Results In the mouse ‘reactive’ model, the expression of 350 genes changed in response to early stresses and 370 in response to late stresses. A minimal genetic overlap (less than 8.8%) was detected in response to both stress protocols, but 30% of these genes (21) were also differentially regulated in the ‘endogenous’ rat study. This overlap is significantly greater than expected by chance. The VAMP-2 gene, differentially expressed across the rodent studies, was also significantly altered in the human study after correcting for multiple testing. Conclusions Our results suggest that ‘endogenous’ and ‘reactive’ subtypes of depression are associated with largely distinct changes in gene-expression. However, they also suggest that the molecular signature of ‘reactive’ depression caused by early stressors differs considerably from that of ‘reactive’ depression caused by late stressors. A small set of genes was consistently dysregulated across each paradigm and in post-mortem brain tissue of depressed patients suggesting a final common pathway to the disorder. These genes included the VAMP-2 gene, which has previously been associated with Axis-I disorders including MDD, bipolar depression, schizophrenia and with antidepressant treatment response. We also discuss the implications of our findings for disease classification, personalized medicine and case-control studies of MDD. PMID:24886127

Functional human antibody CDR fusions as long-acting therapeutic endocrine agonists.

PubMed

Liu, Tao; Zhang, Yong; Liu, Yan; Wang, Ying; Jia, Haiqun; Kang, Mingchao; Luo, Xiaozhou; Caballero, Dawna; Gonzalez, Jose; Sherwood, Lance; Nunez, Vanessa; Wang, Danling; Woods, Ashley; Schultz, Peter G; Wang, Feng

2015-02-03

On the basis of the 3D structure of a bovine antibody with a well-folded, ultralong complementarity-determining region (CDR), we have developed a versatile approach for generating human or humanized antibody agonists with excellent pharmacological properties. Using human growth hormone (hGH) and human leptin (hLeptin) as model proteins, we have demonstrated that functional human antibody CDR fusions can be efficiently engineered by grafting the native hormones into different CDRs of the humanized antibody Herceptin. The resulting Herceptin CDR fusion proteins were expressed in good yields in mammalian cells and retain comparable in vitro biological activity to the native hormones. Pharmacological studies in rodents indicated a 20- to 100-fold increase in plasma circulating half-life for these antibody agonists and significantly extended in vivo activities in the GH-deficient rat model and leptin-deficient obese mouse model for the hGH and hLeptin antibody fusions, respectively. These results illustrate the utility of antibody CDR fusions as a general and versatile strategy for generating long-acting protein therapeutics.

Fatty acids activate a chimera of the clofibric acid-activated receptor and the glucocorticoid receptor.

PubMed Central

Göttlicher, M; Widmark, E; Li, Q; Gustafsson, J A

1992-01-01

Peroxisome proliferators such as clofibric acid, nafenopin, and WY-14,643 have been shown to activate PPAR (peroxisome proliferator-activated receptor), a member of the steroid nuclear receptor superfamily. We have cloned the cDNA from the rat that is homologous to that from the mouse [Issemann, I. & Green, S. (1990) Nature (London) 347, 645-650], which encodes a 97% similar protein with a particularly well-conserved putative ligand-binding domain. To search for physiologically occurring activators, we established a transcriptional transactivation assay by stably expressing in CHO cells a chimera of rat PPAR and the human glucocorticoid receptor that activates expression of the placental alkaline phosphatase reporter gene under the control of the mouse mammary tumor virus promoter. Testing of compounds related to lipid metabolism or peroxisomal proliferation revealed that 150 microM concentrations of arachidonic or linoleic acid but not of dehydroepiandrosterone, cholesterol, or 25-hydroxy-cholesterol, activate the receptor chimera. In addition, saturated fatty acids induce the reporter gene. Shortening the chain length to n = 6 or introduction of an omega-terminal carboxylic group abolished the activation potential of the fatty acid. In conclusion, the present results indicate that fatty acids can regulate gene expression mediated by a member of the steroid nuclear receptor superfamily. Images PMID:1316614

DNA sequence responsible for the amplification of adjacent genes.

PubMed

Pasion, S G; Hartigan, J A; Kumar, V; Biswas, D K

1987-10-01

A 10.3-kb DNA fragment in the 5'-flanking region of the rat prolactin (rPRL) gene was isolated from F1BGH(1)2C1, a strain of rat pituitary tumor cells (GH cells) that produces prolactin in response to 5-bromodeoxyuridine (BrdU). Following transfection and integration into genomic DNA of recipient mouse L cells, this DNA induced amplification of the adjacent thymidine kinase gene from Herpes simplex virus type 1 (HSV1TK). We confirmed the ability of this "Amplicon" sequence to induce amplification of other linked or unlinked genes in DNA-mediated gene transfer studies. When transferred into the mouse L cells with the 10.3-5'rPRL gene sequence of BrdU-responsive cells, both the human growth hormone and the HSV1TK genes are amplified in response to 5-bromodeoxyuridine. This observation is substantiated by BrdU-induced amplification of the cotransferred bacterial Neo gene. Cotransfection studies reveal that the BrdU-induced amplification capability is associated with a 4-kb DNA sequence in the 5'-flanking region of the rPRL gene of BrdU-responsive cells. These results demonstrate that genes of heterologous origin, linked or unlinked, and selected or unselected, can be coamplified when located within the amplification boundary of the Amplicon sequence.

Cardiorespiratory alterations in rodents experimentally envenomed with Hadruroides lunatus scorpion venom.

PubMed

Costal-Oliveira, Fernanda; Guerra-Duarte, Clara; Oliveira, Maira Souza; Castro, Karen Larissa Pereira de; Lopes-de-Sousa, Leticia; Lara, Aline; Gomes, Enéas Ricardo de Morais; Bonilla, Cesar; Guatimosim, Sílvia; Melo, Marília Martins; Chávez-Olórtegui, Carlos

2017-01-01

Hadruroides lunatus is the most abundant scorpion species in the Peruvian central coast, where most of the accidents involving humans are registered. In spite of its prevalence, there are only very few studies on H. lunatus envenomation. The aim of the present study was to analyze the cardiorespiratory alterations caused by H. lunatus envenomation in rodents. Wistar rats injected with H. lunatus scorpion venom were submitted to electrocardiography. After euthanasia, rat lungs were collected and histopathologically analyzed. Mouse cardiomyocytes were used to perform immunofluorescence and calcium transient assays. Data were analyzed by ANOVA or Student's t-test. The significance level was set at p < 0.05. It was observed that H. lunatus venom increased heart rate and caused arrhythmia, thereby impairing the heart functioning. Lungs of envenomed animals showed significant alterations, such as diffuse hemorrhage. In addition, immunofluorescence showed that H. lunatus venom was capable of binding to cardiomyocytes. Furthermore, mouse ventricular cardiomyocytes incubated with H. lunatus venom showed a significant decrease in calcium transient, confirming that H. lunatus venom exerts a toxic effect on heart. Our results showed that H. lunatus venom is capable of inducing cardiorespiratory alterations, a typical systemic effect of scorpionism, stressing the importance of medical monitoring in envenomation cases.

Interaction of human, rat, and mouse immunoglobulin A (IgA) with Staphylococcal superantigen-like 7 (SSL7) decoy protein and leukocyte IgA receptor.

PubMed

Wines, Bruce D; Ramsland, Paul A; Trist, Halina M; Gardam, Sandra; Brink, Robert; Fraser, John D; Hogarth, P Mark

2011-09-23

Host survival depends on an effective immune system and pathogen survival on the effectiveness of immune evasion mechanisms. Staphylococcus aureus utilizes a number of molecules to modulate host immunity, including the SSL family of which SSL7 binds IgA and inhibits Fcα receptor I (FcαRI)-mediated function. Other Gram-positive bacterial pathogens produce IgA binding proteins, which, similar to SSL7, also bind the Fc at the CH2/CH3 interface (the junction between constant domains 2 and 3 of the heavy chain). The opposing activities of the host FcαRI-IgA receptor ligand pair and the pathogen decoy proteins select for host and pathogen variants, which exert stronger protection or evasion, respectively. Curiously, mouse but not rat IgA contains a putative N-linked glycosylation site in the center of this host receptor and pathogen-binding site. Here, we demonstrate that this site is glycosylated and that the effect of amino acid changes and glycosylation of the CH2/CH3 interface inhibits interaction with the pathogen IgA binding protein SSL7, while maintaining binding of pIgR, essential to the biosynthesis and transport of SIgA.

Identification and characterisation of the angiotensin converting enzyme-3 (ACE3) gene: a novel mammalian homologue of ACE

PubMed Central

Rella, Monika; Elliot, Joann L; Revett, Timothy J; Lanfear, Jerry; Phelan, Anne; Jackson, Richard M; Turner, Anthony J; Hooper, Nigel M

2007-01-01

Background Mammalian angiotensin converting enzyme (ACE) plays a key role in blood pressure regulation. Although multiple ACE-like proteins exist in non-mammalian organisms, to date only one other ACE homologue, ACE2, has been identified in mammals. Results Here we report the identification and characterisation of the gene encoding a third homologue of ACE, termed ACE3, in several mammalian genomes. The ACE3 gene is located on the same chromosome downstream of the ACE gene. Multiple sequence alignment and molecular modelling have been employed to characterise the predicted ACE3 protein. In mouse, rat, cow and dog, the predicted protein has mutations in some of the critical residues involved in catalysis, including the catalytic Glu in the HEXXH zinc binding motif which is Gln, and ESTs or reverse-transcription PCR indicate that the gene is expressed. In humans, the predicted ACE3 protein has an intact HEXXH motif, but there are other deletions and insertions in the gene and no ESTs have been identified. Conclusion In the genomes of several mammalian species there is a gene that encodes a novel, single domain ACE-like protein, ACE3. In mouse, rat, cow and dog ACE3, the catalytic Glu is replaced by Gln in the putative zinc binding motif, indicating that in these species ACE3 would lack catalytic activity as a zinc metalloprotease. In humans, no evidence was found that the ACE3 gene is expressed and the presence of deletions and insertions in the sequence indicate that ACE3 is a pseudogene. PMID:17597519

miRWalk--database: prediction of possible miRNA binding sites by "walking" the genes of three genomes.

PubMed

Dweep, Harsh; Sticht, Carsten; Pandey, Priyanka; Gretz, Norbert

2011-10-01

MicroRNAs are small, non-coding RNA molecules that can complementarily bind to the mRNA 3'-UTR region to regulate the gene expression by transcriptional repression or induction of mRNA degradation. Increasing evidence suggests a new mechanism by which miRNAs may regulate target gene expression by binding in promoter and amino acid coding regions. Most of the existing databases on miRNAs are restricted to mRNA 3'-UTR region. To address this issue, we present miRWalk, a comprehensive database on miRNAs, which hosts predicted as well as validated miRNA binding sites, information on all known genes of human, mouse and rat. All mRNAs, mitochondrial genes and 10 kb upstream flanking regions of all known genes of human, mouse and rat were analyzed by using a newly developed algorithm named 'miRWalk' as well as with eight already established programs for putative miRNA binding sites. An automated and extensive text-mining search was performed on PubMed database to extract validated information on miRNAs. Combined information was put into a MySQL database. miRWalk presents predicted and validated information on miRNA-target interaction. Such a resource enables researchers to validate new targets of miRNA not only on 3'-UTR, but also on the other regions of all known genes. The 'Validated Target module' is updated every month and the 'Predicted Target module' is updated every 6 months. miRWalk is freely available at http://mirwalk.uni-hd.de/. Copyright © 2011 Elsevier Inc. All rights reserved.

Targeted BikDD expression kills androgen-dependent and castration-resistant prostate cancer cells

PubMed Central

Xie, Xiaoming; Kong, Yanan; Tang, Hailin; Yang, Lu; Hsu, Jennifer L; Hung, Mien-Chie

2014-01-01

Targeted gene therapy is a promising approach for treating prostate cancer after the discovery of prostate cancer-specific promoters such as prostate-specific antigen, rat probasin, and human glandular kallikrein. However, these promoters are androgen-dependent, and after castration or androgen ablation therapy, they become much less active or sometimes inactive. Importantly, the disease will inevitably progress from androgen-dependent (ADPC) to castration-resistant prostate cancer (CRPC) at which treatments fail and high mortality ensues. Therefore, it is critical to develop a targeted gene therapy strategy that is effective in both ADPC and CRPC to eradicate recurrent prostate tumors. The human telomerase reverse transcriptase-VP16-Gal4-WPRE integrated systemic amplifier composite (T-VISA) vector we previously developed which targets transgene expression in ovarian and breast cancer is also active in prostate cancer. To further improve its effectiveness based on androgen response in ADPC progression, the ARR2 element (two copies of androgen response region from rat probasin promoter) was incorporated into T-VISA to produce AT-VISA. Under androgen analog (R1881) stimulation, the activity of AT-VISA was increased to a level greater than or comparable to the cytomegalovirus (CMV) promoter in ADPC and CRPC cells, respectively. Importantly, AT-VISA demonstrated little or no expression in normal cells. Systemic administration of AT-VISA-BikDD encapsulated in liposomes repressed prostate tumor growth and prolonged mouse survival in orthotopic animal models as well as in the transgenic adenocarcinoma mouse prostate model, indicating that AT-VISA-BikDD has therapeutic potential to treat ADPC and CRPC safely and effectively in preclinical setting. PMID:24785255

Novel Monoclonal Antibodies Recognizing Human Prostate-Specific Membrane Antigen (PSMA) as Research and Theranostic Tools.

PubMed

Nováková, Zora; Foss, Catherine A; Copeland, Benjamin T; Morath, Volker; Baranová, Petra; Havlínová, Barbora; Skerra, Arne; Pomper, Martin G; Barinka, Cyril

2017-05-01

Prostate-specific membrane antigen (PSMA) is a validated target for the imaging and therapy of prostate cancer. Here, we report the detailed characterization of four novel murine monoclonal antibodies (mAbs) recognizing human PSMA as well as PSMA orthologs from different species. Performance of purified mAbs was assayed using a comprehensive panel of in vitro experimental setups including Western blotting, immunofluorescence, immunohistochemistry, ELISA, flow cytometry, and surface-plasmon resonance. Furthermore, a mouse xenograft model of prostate cancer was used to compare the suitability of the mAbs for in vivo applications. All mAbs demonstrate high specificity for PSMA as documented by the lack of cross-reactivity to unrelated human proteins. The 3F11 and 1A11 mAbs bind linear epitopes spanning residues 226-243 and 271-288 of human PSMA, respectively. 3F11 is also suitable for the detection of PSMA orthologs from mouse, pig, dog, and rat in experimental setups where the denatured form of PSMA is used. 5D3 and 5B1 mAbs recognize distinct surface-exposed conformational epitopes and are useful for targeting PSMA in its native conformation. Most importantly, using a mouse xenograft model of prostate cancer we show that both the intact 5D3 and its Fab fragment are suitable for in vivo imaging. With apparent affinities of 0.14 and 1.2 nM as determined by ELISA and flow cytometry, respectively, 5D3 has approximately 10-fold higher affinity for PSMA than the clinically validated mAb J591 and, therefore, is a prime candidate for the development of next-generation theranostics to target PSMA. Prostate 77:749-764, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Nestin in the epididymis is expressed in vascular wall cells and is regulated during postnatal development and in case of testosterone deficiency.

PubMed

Reckmann, Ansgar N; Tomczyk, Claudia U M; Davidoff, Michail S; Michurina, Tatyana V; Arnhold, Stefan; Müller, Dieter; Mietens, Andrea; Middendorff, Ralf

2018-01-01

Vascular smooth muscle cells (SMCs), distinguished by the expression of the neuronal stem cell marker nestin, may represent stem cell-like progenitor cells in various organs including the testis. We investigated epididymal tissues of adult nestin-GFP mice, rats after Leydig cell depletion via ethane dimethane sulfonate (EDS), rats and mice during postnatal development and human tissues. By use of Clarity, a histochemical method to illustrate a three-dimensional picture, we could demonstrate nestin-GFP positive cells within the vascular network. We localized nestin in the epididymis in proliferating vascular SMCs by colocalization with both smooth muscle actin and PCNA, and it was distinct from CD31-positive endothelial cells. The same nestin localization was found in the human epididymis. However, nestin was not found in SMCs of the epididymal duct. Nestin expression is high during postnatal development of mouse and rat and down-regulated towards adulthood when testosterone levels increase. Nestin increases dramatically in rats after Leydig cell ablation with EDS and subsequently low testosterone levels. Interestingly, during this period, the expression of androgen receptor in the epididymis is low and increases until nestin reaches normal levels of adulthood. Here we show that nestin, a common marker for neuronal stem cells, is also expressed in the vasculature of the epididymis. Our results give new insights into the yet underestimated role of proliferating nestin-expressing vascular SMCs during postnatal development and repair of the epididymis.

Gene transfer with a vector expressing Maxi-K from a smooth muscle-specific promoter restores erectile function in the aging rat.

PubMed

Melman, A; Biggs, G; Davies, K; Zhao, W; Tar, M T; Christ, G J

2008-03-01

Previous reports have demonstrated that gene transfer with the alpha, or pore-forming, subunit of the human Maxi-K channel (hSlo) restores the decline in erectile capacity observed in established rat models of diabetes and aging. Preliminary data from a human clinical trial also showed safety and potential efficacy in 11 men treated with the same plasmid construct expressing the Maxi-K channel. In all instances, the original plasmid was driven by the heterologous cytomegalovirus promoter which is broadly active in a wide variety of cell and tissue types. To more precisely determine the contribution of the corporal myocyte to the observed physiological effects in vivo, we report here our initial work using a distinct vector (pSMAA-hSlo) in which hSlo gene expression was driven off the mouse smooth muscle alpha-actin (SMAA) promoter. Specifically, older rats, with diminished erectile capacity, were given a single intracorporal injection with either 100 mug pVAX-hSlo or 10, 100 or 1000 mug pSMAA-hSlo, or vector or vehicle alone. Significantly increased intracavernous pressure (ICP) responses to cavernous nerve stimulation were observed for all doses of both plasmids encoding hSlo, relative to control injections. These data confirm and extend previous observations to document that smooth muscle cell-specific expression of hSlo in corporal tissue is both necessary and sufficient to restore erectile function in aging rats.

Induction of ovulation by a potent, orally active, low molecular weight agonist (Org 43553) of the luteinizing hormone receptor.

PubMed

van de Lagemaat, R; Timmers, C M; Kelder, J; van Koppen, C; Mosselman, S; Hanssen, R G J M

2009-03-01

In assisted reproductive technology, human chorionic gonadotrophin (hCG) is administered subcutaneously for the induction of oocyte maturation and ovulation. Our efforts to develop orally bioavailable luteinizing hormone (LH) receptor agonists have led to the discovery of Org 43553, a low molecular weight (LMW) LH receptor (LH-R) agonist. Org 43553 was tested in vitro and in vivo in pre-clinical pharmacological models to demonstrate efficacy and oral availability. Org 43553 is a potent stimulator of the human LH-R in vitro (EC(50) 3.7 nM). In primary mouse Leydig cells, Org 43553 stimulated testosterone production. Pharmacokinetic analyses showed high oral bioavailability in rats (79%) and dogs (44%) with a shorter half-life compared with hCG (3.4 versus 5.6 h in the rat). Ovulation induction by Org 43553 was demonstrated in immature mice as well as in cyclic rats after single-dose oral administration (50 mg/kg). The ovulated oocytes were of good quality as demonstrated by successful fertilization and implantation of normal embryos. In male rats, testosterone production was substantially induced after oral administration. Org 43553 is the first LMW LH-R mimetic with demonstrated in vivo efficacy upon oral administration and could therefore replace subcutaneously administered hCG. The elimination half-life of Org 43553 is substantially shorter than hCG, which could potentially represent a clinical benefit in reducing the risk of ovarian hyperstimulation syndrome (OHSS).

The clinical and immunological significance of GAD-specific autoantibody and T-cell responses in type 1 diabetes.

PubMed

Boettler, Tobias; Pagni, Philippe P; Jaffe, Rachel; Cheng, Yang; Zerhouni, Peter; von Herrath, Matthias

2013-08-01

Antigen-specific interventions are desirable approaches in Type 1 Diabetes (T1D) as they can alter islet-specific autoimmunity without systemic side effects. Glutamic acid decarboxylase of 65 kDa (GAD65) is a major autoantigen in type 1 diabetes (T1D) and GAD-specific autoimmunity is a common feature of T1D in humans but also in mouse models of the disease. In humans, administration of the GAD65 protein in an alum formulation has been shown to reduce C-peptide decline in recently diagnosed patients, however, these observations were not confirmed in subsequent phase II/III clinical trials. As GAD-based immune interventions in different formulations have successfully been employed to prevent the establishment of T1D in mouse models of T1D, we sought to analyze the efficacy of GAD-alum treatment and the effects on the GAD-specific immune response in two different mouse models of T1D. Consistent with the latest clinical trials, mice treated with GAD-alum were not protected from diabetes, although GAD-alum induced a GAD-specific Th2-deviated immune response in transgenic rat insulin promoter-glycoprotein (RIP-GP) mice. These observations underline the importance of a thorough, preclinical evaluation of potential drugs before the initiation of clinical trials. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

Characterization of species-related differences in the pharmacology of tachykinin NK receptors 1, 2 and 3.

PubMed

Leffler, Agnes; Ahlstedt, Ingela; Engberg, Susanna; Svensson, Arne; Billger, Martin; Oberg, Lisa; Bjursell, Magnus K; Lindström, Erik; von Mentzer, Bengt

2009-05-01

Tachykinin NK receptors (NKRs) differ to a large degree among species with respect to their affinities for small molecule antagonists. The aims of the present study were to clone NKRs from gerbil (NK2R and NK3R) and dog (NK1R, NK2R and NK3R) in which the sequence was previously unknown and to investigate the potency of several NKR antagonists at all known human, dog, gerbil and rat NKRs. The NKR protein coding sequences were cloned and expressed in CHO cells. The inhibitory concentrations of selective and non-selective NKR antagonists were determined by inhibition of agonist-induced mobilization of intracellular Ca2+. Receptor homology models were constructed based on the rhodopsin crystal structure to investigate and identify the antagonist binding sites and interaction points in the transmembrane (TM) regions of the NKRs. Data collected using the cloned dog NK1R confirmed that the dog NK1R displays similar pharmacology as the human and the gerbil NK1R, but differs greatly from the mouse and the rat NK1R. Despite species-related amino acid (AA) differences located close to the antagonist binding pocket of the NK2R, they did not affect the potency of the antagonists ZD6021 and saredutant. Two AA differences located close to the antagonist binding site of NK3R likely influence the NK3R antagonist potency, explaining the 3-10-fold decrease in potency observed for the rat NK3R. For the first time, detailed pharmacological experiments in vitro with cloned NKRs demonstrate that not only human, but also dog and gerbil NKR displays similar antagonist pharmacology while rat diverges significantly with respect to NK1R and NK3R.

Stem cell potency and the ability to contribute to chimeric organisms.

PubMed

Polejaeva, Irina; Mitalipov, Shoukhrat

2013-03-01

Mouse embryonic chimeras are a well-established tool for studying cell lineage commitment and pluripotency. Experimental chimeras were successfully produced by combining two or more preimplantation embryos or by introducing into host embryo cultured pluripotent embryonic stem cells (ESCs). Chimera production using genetically modified ESCs became the method of choice for the generation of knockout or knockin mice. Although the derivation of ESCs or ESC-like cells has been reported for other species, only mouse and rat pluripotent stem cells have been shown to contribute to germline-competent chimeras, which is the defining feature of ESCs. Herein, we describe different approaches employed for the generation of embryonic chimeras, define chimera-competent cell types, and describe cases of spontaneous chimerism in humans. We also review the current state of derivation of pluripotent stem cells in several species and discuss outcomes of various chimera studies when such cells are used.

Wound Regeneration Deficit in Rats Correlates with Low Morphogenetic Potential and Distinct Transcriptome Profile of Epidermis.

PubMed

Guerrero-Juarez, Christian F; Astrowski, Aliaksandr A; Murad, Rabi; Dang, Christina T; Shatrova, Vera O; Astrowskaja, Aksana; Lim, Chae Ho; Ramos, Raul; Wang, Xiaojie; Liu, Yuchen; Lee, Hye-Lim; Pham, Kim T; Hsi, Tsai-Ching; Oh, Ji Won; Crocker, Daniel; Mortazavi, Ali; Ito, Mayumi; Plikus, Maksim V

2018-06-01

Large excisional wounds in mice prominently regenerate new hair follicles (HFs) and fat, yet humans are deficient for this regenerative behavior. Currently, wound-induced regeneration remains a clinically desirable, but only partially understood phenomenon. We show that large excisional wounds in rats across seven strains fail to regenerate new HFs. We compared wound transcriptomes between mice and rats at the time of scab detachment, which coincides with the onset of HF regeneration in mice. In both species, wound dermis and epidermis share core dermal and epidermal transcriptional programs, respectively, yet prominent interspecies differences exist. Compared with mice, rat epidermis expresses distinct transcriptional and epigenetic factors, markers of epidermal repair, hyperplasia, and inflammation, and lower levels of WNT signaling effectors and regulators. When recombined on the surface of excisional wounds with vibrissa dermal papillae, partial-thickness skin grafts containing distal pelage HF segments, but not interfollicular epidermis, readily regenerated new vibrissa-like HFs. Together, our findings establish rats as a nonregenerating rodent model for excisional wound healing and suggest that low epidermal competence and associated transcriptional profile may contribute to its regenerative deficiency. Future comparison between rat and mouse may lend further insight into the mechanism of wounding-induced regeneration and causes for its deficit. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Protection against methotrexate toxicity by a soybean protein- and omega-3 fatty acid-containing diet: comparative study with a casein-containing diet.

PubMed

Mitsugi, Kenji; Nakamura, Tsuyoshi; Kashiwabara, Norio; Ariyama, Hiroshi; Tanaka, Risa; Baba, Eishi; Nakamura, Minoru; Harada, Mine; Nakano, Shuji

2004-07-01

Effects of two clinically used liquid diets on toxicity and antitumor activity of methotrexate (MTX) were investigated in Sprague-Dawley (SD) rats and tumor-bearing mice, respectively. Diets tested were commercially available formulas enriched either with soybean and omega-3 fatty acids or with casein. The soybean-containing diet offered significant protection against MTX toxicity in rats compared with the casein-containing diet, completely alleviating MTX-induced anorexia, diarrhea, and weight loss, when ingested as the sole diet and fed 7 days prior to and 7 days following intraperitoneal MTX injection. As a result, 90% of rats were alive on soybean-containing diet while all rats were dead on casein-containing diet. Histologic examination of the small intestine of MTX-treated rats revealed that the soybean-containing diet significantly prevented loss of mucosal villus tips compared to the casein-containing diet. Pharmocokinetic examination indicated that plasma MTX concentrations were similar for rats in the two diet-defined groups. No significant differences were observed between diets in survival of mice injected with L1210 mouse leukemia cells and subsequently with MTX. Thus the soybean-containing diet appeared to be superior to the casein-containing diet in preventing gastrointestinal toxicity while preserving antitumor activity. A soybean diet enriched in omega-3 fatty acids may be a useful adjunct to MTX treatment of human cancers.

Grammomys surdaster, the Natural Host for Plasmodium berghei Parasites, as a Model to Study Whole-Organism Vaccines Against Malaria.

PubMed

Conteh, Solomon; Anderson, Charles; Lambert, Lynn; Orr-Gonzalez, Sachy; Herrod, Jessica; Robbins, Yvette L; Carter, Dariyen; Karhemere, Stomy Bin Shamamba; Pyana, Pati; Büscher, Philippe; Duffy, Patrick E

2017-04-01

AbstractInbred mice are commonly used to test candidate malaria vaccines, but have been unreliable for predicting efficacy in humans. To establish a more rigorous animal model, we acquired African woodland thicket rats of the genus Grammomys , the natural hosts for Plasmodium berghei . Thicket rats were acquired and identified as Grammomys surdaster by skull and teeth measurements and mitochondrial DNA genotyping. Herein, we demonstrate that thicket rats are highly susceptible to infection by P. berghei , and moderately susceptible to Plasmodium yoelii and Plasmodium chabaudi : 1-2 infected mosquito bites or 25-100 sporozoites administered by intravenous injection consistently resulted in patent parasitemia with P. berghei , and resulted in patent parasitemia with P. yoelii and P. chabaudi strains for at least 50% of animals. We then assessed efficacy of whole-organism vaccines to induce sterile immunity, and compared the thicket rat model to conventional mouse models. Using P. berghei ANKA radiation-attenuated sporozoites, and P. berghei ANKA and P. yoelii chemoprophylaxis vaccination approaches, we found that standard doses of vaccine sufficient to protect laboratory mice for a long duration against malaria challenge, are insufficient to protect thicket rats, which require higher doses of vaccine to achieve even short-term sterile immunity. Thicket rats may offer a more stringent and pertinent model for evaluating whole-organism vaccines.

Rat PPAR delta contains a CGG triplet repeat and is prominently expressed in the thalamic nuclei.

PubMed

Xing, G; Zhang, L; Zhang, L; Heynen, T; Yoshikawa, T; Smith, M; Weiss, S; Detera-Wadleigh, S

1995-12-26

We have isolated a new rat sequence containing motifs of a nuclear hormone receptor from a brain cDNA library. The deduced amino acid sequence encoded by the cDNA clone showed a strong homology to the human NUCI and the mouse peroxisome proliferator activated receptor delta (PPAR delta). We therefore refer to this new clone as rat PPAR delta (rPPAR delta). The new feature of rPPAR delta is a 14 CGG triplet repeat on the 5' untranslated region, not previously reported in either NUCI or mPPAR delta. We found that rPPAR delta was expressed as a 3.5-kb transcript which showed a wide distribution in adult rat tissues. Abundant expression was detected in brain, heart, skeletal muscle, kidney and lung. Weaker expression was noted in the liver, spleen and testis. To determine the specific brain localization of rPPAR delta we performed in situ hybridization analysis. Prominent expression was observed in the thalamus, particularly in the posterior part of the ventral medial nucleus, a site responsive to pain and cold stress. These results raise the possibility that PPAR delta might play a role in modulating response to thermal and pain sensations.

Developmental history and application of CRISPR in human disease.

PubMed

Liang, Puping; Zhang, Xiya; Chen, Yuxi; Huang, Junjiu

2017-06-01

Genome-editing tools are programmable artificial nucleases, mainly including zinc-finger nucleases, transcription activator-like effector nucleases and clustered regularly interspaced short palindromic repeat (CRISPR). By recognizing and cleaving specific DNA sequences, genome-editing tools make it possible to generate site-specific DNA double-strand breaks (DSBs) in the genome. DSBs will then be repaired by either error-prone nonhomologous end joining or high-fidelity homologous recombination mechanisms. Through these two different mechanisms, endogenous genes can be knocked out or precisely repaired/modified. Rapid developments in genome-editing tools, especially CRISPR, have revolutionized human disease models generation, for example, various zebrafish, mouse, rat, pig, monkey and human cell lines have been constructed. Here, we review the developmental history of CRISPR and its application in studies of human diseases. In addition, we also briefly discussed the therapeutic application of CRISPR in the near future. Copyright © 2017 John Wiley & Sons, Ltd.

Species differences in the metabolism of benzene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Henderson, R.F.

1996-12-01

The pathways of metabolism of benzene appear to be qualitatively similar in all species studied thus far. However, there are quantitative differences in the fraction of benzene metabolized by the different pathways. These species differences become important for risk assessments based on animal data. Mice have a greater overall capacity to metabolize benzene than rats or primates, based on mass balance studies conducted in vivo using radiolabled benzene. Mice and monkeys metabolize more of the benzene to hydroquinone metabolites than do rats or chimpanzees, especially at low doses. Nonhuman primates metabolize less of the benzene to muconic acid than domore » rodents or humans. In all species studied, a greater proportion of benzene is converted to hydroquinone and ring-breakage metabolites at low doses than at high doses. This finding should be considered in attempting to extrapolate the toxicity of benzene observed at high doses to predicted toxicity at low doses. Because ring-breakage metabolites and hydroquinone have both been implicated in the toxicity of benzene, the higher formation of those metabolites in the mouse may partially explain why mice are more sensitive to benzene than are rats. Metabolism of benzene in humans, the species of interest, does not exactly mimic that of any animal species studied. More information on the urinary and blood metabolites of occupationally exposed people is required to determine the fractional conversion of benzene to putative toxic metabolites and the degree of variability present in human subjects. 12 refs., 4 tabs.« less

Transgenic rat model of childhood-onset dermatitis by overexpressing telomerase reverse transcriptase (TERT).

PubMed

Kaneko, Ryosuke; Sato, Atsuko; Hamada, Shun; Yagi, Takeshi; Ohsawa, Ichiro; Ohtsuki, Mamitaro; Kobayashi, Eiji; Hirabayashi, Masumi; Murakami, Takashi

2016-08-01

Childhood-onset dermatitis is one of the most common skin disorders in children. Although various mouse models that mirror aspects of dermatitis have become available, there is still a need for an animal model that develops dermatitis in childhood and is more suitable for performing tissue transplantation experiments. There is emerging evidence that peripheral blood T lymphocytes from patients with dermatitis have significantly increased telomerase activity. Here, we developed telomerase reverse transcriptase (TERT)-expressing transgenic (Tg) rats that spontaneously developed eczematous skin inflammation in childhood. Newborn TERT-Tg rats developed visible dermatitis in 56 % of cases, and the skin lesions microscopically showed spongiosis and acanthosis with infiltration of lymphocytes, eosinophils and mast cells. TERT-Tg rats with dermatitis exhibited increased CD4 (2.5-fold) and CD8 (fivefold) T cell numbers compared with dermatitis-free TERT-Tg rats. Stronger TERT activity was observed in the peripheral lymphocytes of dermatitis-positive TERT-Tg rats than those of dermatitis-free TERT-Tg rats. RT-PCR analysis revealed that IL-4 was markedly elevated in the spleen of dermatitis-positive TERT-Tg rats, and that interferon-gamma was increased in the dermatitis lesions. Moreover, skin grafting of TERT-Tg rats with dermatitis onto T cell-deficient nude rats demonstrated that the inflamed skin lesions could not be maintained. Taken together, the results suggest that TERT activation in T lymphocytes is one of the potential predisposing factors for dermatitis. Moreover, our results demonstrated that the TERT-Tg rats mirror aspects of human childhood-onset dermatitis and that these animals represent a potential animal model system for studying childhood-onset dermatitis.

Human Health Effects of Biphenyl: Key Findings and Scientific Issues.

PubMed

Li, Zheng; Hogan, Karen A; Cai, Christine; Rieth, Susan

2016-06-01

In support of the Integrated Risk Information System (IRIS), the U.S. Environmental Protection Agency (EPA) has evaluated the human health hazards of biphenyl exposure. We review key findings and scientific issues regarding expected human health effects of biphenyl. Scientific literature from 1926 through September 2012 was critically evaluated to identify potential human health hazards associated with biphenyl exposure. Key issues related to the carcinogenicity and noncancer health hazards of biphenyl were examined based on evidence from experimental animal bioassays and mechanistic studies. Systematic consideration of experimental animal studies of oral biphenyl exposure took into account the variety of study designs (e.g., study sizes, exposure levels, and exposure durations) to reconcile differing reported results. The available mechanistic and toxicokinetic evidence supports the hypothesis that male rat urinary bladder tumors arise through urinary bladder calculi formation but is insufficient to hypothesize a mode of action for liver tumors in female mice. Biphenyl and its metabolites may induce genetic damage, but a role for genotoxicity in biphenyl-induced carcinogenicity has not been established. The available health effects data for biphenyl provides suggestive evidence for carcinogenicity in humans, based on increased incidences of male rat urinary bladder tumors at high exposure levels and on female mouse liver tumors. Kidney toxicity is also a potential human health hazard of biphenyl exposure. Li Z, Hogan KA, Cai C, Rieth S. 2016. Human health effects of biphenyl: key findings and scientific issues. Environ Health Perspect 124:703-712; http://dx.doi.org/10.1289/ehp.1509730.

Developmental toxicity evaluation of inhaled tertiary amyl methyl ether in mice and rats.

PubMed

Welsch, Frank; Elswick, Barbara; James, R Arden; Marr, Melissa C; Myers, Christina B; Tyl, Rochelle W

2003-01-01

This evaluation was part of a much more comprehensive testing program to characterize the mammalian toxicity potential of the gasoline oxygenator additive tertiary amyl methyl ether (TAME), and was initiated upon a regulatory agency mandate. A developmental toxicity hazard identification study was conducted by TAME vapor inhalation exposure in two pregnant rodent species. Timed-pregnant CD(Sprague-Dawley) rats and CD-1 mice, 25 animals per group, inhaled TAME vapors containing 0, 250, 1500 or 3500 ppm for 6 h a day on gestational days 6-16 (mice) or 6-19 (rats). The developmental toxicity hazard potential was evaluated following the study design draft guidelines and end points proposed by the United States Environmental Protection Agency. Based on maternal body weight changes during pregnancy, the no-observable-adverse-effect level (NOAEL) was 250 ppm for maternal toxicity in rats and 1500 ppm for developmental toxicity in rats using the criterion of near-term fetal body weights. In mice, more profound developmental toxicity was present than in rats, at both 1500 and 3500 ppm. At the highest concentration, mouse litters revealed more late fetal deaths, significantly reduced fetal body weights per litter and increased incidences of cleft palate (classified as an external malformation), as well as enlarged lateral ventricles of the cerebrum (a visceral variation). At 1500 ppm, mouse fetuses also exhibited an increased incidence of cleft palate and the dam body weights were reduced. Therefore, the NOAEL for the mouse maternal and developmental toxicity was 250 ppm under the conditions of this study. Copyright 2003 John Wiley & Sons, Ltd.

In vivo and in vitro gene transfer to mammalian somatic cells by particle bombardment.

PubMed Central

Yang, N S; Burkholder, J; Roberts, B; Martinell, B; McCabe, D

1990-01-01

Chimeric chloramphenicol acetyltransferase and beta-galactosidase marker genes were coated onto fine gold particles and used to bombard a variety of mammalian tissues and cells. Transient expression of the genes was obtained in liver, skin, and muscle tissues of rat and mouse bombarded in vivo. Similar results were obtained with freshly isolated ductal segments of rat and human mammary glands and primary cultures derived from these explants. Gene transfer and transient expression were also observed in eight human cell culture lines, including cells of epithelial, endothelial, fibroblast, and lymphocyte origin. Using CHO and MCF-7 cell cultures as models, we obtained stable gene transfer at frequencies of 1.7 x 10(-3) and 6 x 10(-4), respectively. The particle bombardment technology thus provides a useful means to transfer foreign genes into a variety of mammalian somatic cell systems. The method is applicable to tissues in vivo as well as to isolated cells in culture and has proven effective with all cell or tissue types tested thus far. This technology may therefore prove to be applicable in various aspects of gene therapy. Images PMID:2175906

Estradiol alters body temperature regulation in the female mouse.

PubMed

Krajewski-Hall, Sally J; Blackmore, Elise M; McMinn, Jessi R; Rance, Naomi E

2018-01-01

Hot flushes are due to estrogen withdrawal and characterized by the episodic activation of heat dissipation effectors. Recent studies (in humans and rats) have implicated neurokinin 3 (NK 3 ) receptor signaling in the genesis of hot flushes. Although transgenic mice are increasingly used for biomedical research, there is limited information on how 17β-estradiol and NK 3 receptor signaling alters thermoregulation in the mouse. In this study, a method was developed to measure tail skin temperature (T SKIN ) using a small data-logger attached to the surface of the tail, which, when combined with a telemetry probe for core temperature (T CORE ), allowed us to monitor thermoregulation in freely-moving mice over long durations. We report that estradiol treatment of ovariectomized mice reduced T CORE during the light phase (but not the dark phase) while having no effect on T SKIN or activity. Estradiol also lowered T CORE in mice exposed to ambient temperatures ranging from 20 to 36°C. Unlike previous studies in the rat, estradiol treatment of ovariectomized mice did not reduce T SKIN during the dark phase. Subcutaneous injections of an NK 3 receptor agonist (senktide) in ovariectomized mice caused an acute increase in T SKIN and a reduction in T CORE , consistent with the activation of heat dissipation effectors. These changes were reduced by estradiol, suggesting that estradiol lowers the sensitivity of central thermoregulatory pathways to NK 3 receptor activation. Overall, we show that estradiol treatment of ovariectomized mice decreases T CORE during the light phase, reduces the thermoregulatory effects of senktide and modulates thermoregulation differently than previously described in the rat.

An assessment of the effects of serotonin 6 (5-HT6) receptor antagonists in rodent models of learning.

PubMed

Lindner, Mark D; Hodges, Donald B; Hogan, John B; Orie, Anitra F; Corsa, Jason A; Barten, Donna M; Polson, Craig; Robertson, Barbara J; Guss, Valerie L; Gillman, Kevin W; Starrett, John E; Gribkoff, Valentin K

2003-11-01

Antagonists of serotonin 6 (5-HT6) receptors have been reported to enhance cognition in animal models of learning, although this finding has not been universal. We have assessed the therapeutic potential of the specific 5-HT6 receptor antagonists 4-amino-N-(2,6-bis-methylamino-pyrimidin-4-yl)-benzenesulfonamide (Ro 04-6790) and 5-chloro-N-(4-methoxy-3-piperazin-1-yl-phenyl)-3-methyl-2-benzothiophenesulfonamide (SB-271046) in rodent models of cognitive function. Although mice express the 5-HT6 receptor and the function of this receptor has been investigated in mice, all reports of activity with 5-HT6 receptor antagonists have used rat models. In the present study, receptor binding revealed that the pharmacological properties of the mouse receptor are different from the rat and human receptor: Ro 04-6790 does not bind to the mouse 5-HT6 receptor, so all in vivo testing included in the present report was conducted in rats. We replicated previous reports that 5-HT6 receptor antagonists produce a stretching syndrome previously shown to be mediated through cholinergic mechanisms, but Ro 04-6790 and SB-271046 failed to attenuate scopolamine-induced deficits in a test of contextual fear conditioning. We also failed to replicate the significant effects reported previously in both an autoshaping task and in a version of the Morris water maze. The results of our experiments are not consistent with previous reports that suggested that 5-HT6 antagonists might have therapeutic potential for cognitive disorders.

Nuclear Export Factor CRM1 Interacts with Nonstructural Proteins NS2 from Parvovirus Minute Virus of Mice

PubMed Central

Bodendorf, Ursula; Cziepluch, Celina; Jauniaux, Jean-Claude; Rommelaere, Jean; Salomé, Nathalie

1999-01-01

The nonstructural NS2 proteins of autonomous parvoviruses are known to act in a host cell-dependent manner and to play a role in viral DNA replication, efficient translation of viral mRNA, and/or encapsidation. Their exact function during the parvovirus life cycle remains, however, still obscure. We report here the characterization of the interaction with the NS2 proteins from the parvovirus minute virus of mice (MVM) and rat as well as mouse homologues of the human CRM1 protein, a member of the importin-beta family recently identified as an essential nuclear export factor. Using the two-hybrid system, we could detect the interaction between the carboxy-terminal region of rat CRM1 and each of the three isoforms of NS2 (P [or major], Y [or minor], and L [or rare]). NS2 proteins were further shown to interact with the full-length CRM1 by coimmunoprecipitation experiments using extracts from both mouse and rat cell lines. Our data show that CRM1 preferentially binds to the nonphosphorylated isoforms of NS2. Moreover, we observed that the treatment of MVM-infected cells with leptomycin B, a drug that specifically inhibits the CRM1-dependent nuclear export pathway, leads to a drastic accumulation of NS2 proteins in the nucleus. Both NS2 interaction with CRM1 and nuclear accumulation upon leptomycin B treatment strongly suggest that these nonstructural viral proteins are actively exported out of the nuclei of infected cells via a CRM1-mediated nuclear export pathway. PMID:10438867

The Fecal Viral Flora of Wild Rodents

PubMed Central

Phan, Tung G.; Kapusinszky, Beatrix; Wang, Chunlin; Rose, Robert K.; Lipton, Howard L.; Delwart, Eric L.

2011-01-01

The frequent interactions of rodents with humans make them a common source of zoonotic infections. To obtain an initial unbiased measure of the viral diversity in the enteric tract of wild rodents we sequenced partially purified, randomly amplified viral RNA and DNA in the feces of 105 wild rodents (mouse, vole, and rat) collected in California and Virginia. We identified in decreasing frequency sequences related to the mammalian viruses families Circoviridae, Picobirnaviridae, Picornaviridae, Astroviridae, Parvoviridae, Papillomaviridae, Adenoviridae, and Coronaviridae. Seventeen small circular DNA genomes containing one or two replicase genes distantly related to the Circoviridae representing several potentially new viral families were characterized. In the Picornaviridae family two new candidate genera as well as a close genetic relative of the human pathogen Aichi virus were characterized. Fragments of the first mouse sapelovirus and picobirnaviruses were identified and the first murine astrovirus genome was characterized. A mouse papillomavirus genome and fragments of a novel adenovirus and adenovirus-associated virus were also sequenced. The next largest fraction of the rodent fecal virome was related to insect viruses of the Densoviridae, Iridoviridae, Polydnaviridae, Dicistroviriade, Bromoviridae, and Virgaviridae families followed by plant virus-related sequences in the Nanoviridae, Geminiviridae, Phycodnaviridae, Secoviridae, Partitiviridae, Tymoviridae, Alphaflexiviridae, and Tombusviridae families reflecting the largely insect and plant rodent diet. Phylogenetic analyses of full and partial viral genomes therefore revealed many previously unreported viral species, genera, and families. The close genetic similarities noted between some rodent and human viruses might reflect past zoonoses. This study increases our understanding of the viral diversity in wild rodents and highlights the large number of still uncharacterized viruses in mammals. PMID:21909269

Human mammary microenvironment better regulates the biology of human breast cancer in humanized mouse model.

PubMed

Zheng, Ming-Jie; Wang, Jue; Xu, Lu; Zha, Xiao-Ming; Zhao, Yi; Ling, Li-Jun; Wang, Shui

2015-02-01

During the past decades, many efforts have been made in mimicking the clinical progress of human cancer in mouse models. Previously, we developed a human breast tissue-derived (HB) mouse model. Theoretically, it may mimic the interactions between "species-specific" mammary microenvironment of human origin and human breast cancer cells. However, detailed evidences are absent. The present study (in vivo, cellular, and molecular experiments) was designed to explore the regulatory role of human mammary microenvironment in the progress of human breast cancer cells. Subcutaneous (SUB), mammary fat pad (MFP), and HB mouse models were developed for in vivo comparisons. Then, the orthotopic tumor masses from three different mouse models were collected for primary culture. Finally, the biology of primary cultured human breast cancer cells was compared by cellular and molecular experiments. Results of in vivo mouse models indicated that human breast cancer cells grew better in human mammary microenvironment. Cellular and molecular experiments confirmed that primary cultured human breast cancer cells from HB mouse model showed a better proliferative and anti-apoptotic biology than those from SUB to MFP mouse models. Meanwhile, primary cultured human breast cancer cells from HB mouse model also obtained the migratory and invasive biology for "species-specific" tissue metastasis to human tissues. Comprehensive analyses suggest that "species-specific" mammary microenvironment of human origin better regulates the biology of human breast cancer cells in our humanized mouse model of breast cancer, which is more consistent with the clinical progress of human breast cancer.

Experimental Models of Vaginal Candidiasis and Their Relevance to Human Candidiasis

PubMed Central

Sobel, Jack D.

2016-01-01

Vulvovaginal candidiasis (VVC) is a high-incidence disease seriously affecting the quality of life of women worldwide, particularly in its chronic, recurrent forms (RVVC), and with no definitive cure or preventive measure. Experimental studies in currently used rat and mouse models of vaginal candidiasis have generated a large mass of data on pathogenicity determinants and inflammation and immune responses of potential importance for the control of human pathology. However, reflection is necessary about the relevance of these rodent models to RVVC. Here we examine the chemical, biochemical, and biological factors that determine or contrast the forms of the disease in rodent models and in women and highlight the differences between them. We also appeal for approaches to improve or replace the current models in order to enhance their relevance to human infection. PMID:26883592

Effect of Synthetic Matrix Metalloproteinase Inhibitors on Lipopolysaccharide-Induced Blood-Brain Barrier Opening in Rodents: Differences in Response Based on Strains and Solvents

PubMed Central

Rosenberg, Gary A.; Estrada, Eduardo Y.; Mobashery, Shahriar

2007-01-01

Matrix metalloproteinase inhibitors (MMPIs) reduce blood-brain barrier (BBB) disruption and prevent cell death. Animal models of multiple sclerosis, cerebral ischemia and hemorrhage, and bacterial meningitis respond to treatment with MMPIs. We have used the intracerebral injection of lipopolysaccharide (LPS) in rat, which induces MMP production and results in a delayed opening of the BBB, to screen MMPIs to identify therapeutic agents. We hypothesized that the mouse would respond similarly to LPS and that the mouse/LPS model of BBB damage would be more useful for screening of MMPIs. Therefore, we adapted the rat LPS model to the mouse and compared the response to LPS and treatment with MMPIs. Wistar-Kyoto rats (WKY) and three strains of mice had stereotactic injections of LPS into the caudate. 14C-sucrose was used to measure permeability of the BBB 24 hours after injection. Initially, we tested three broad-spectrum MMPIs in the rat, BB-1101, BB-94, and BB-2293, and a MMP-2 selective inhibitor, IW449; both BB-1101 and BB-94 significantly suppressed LPS-induced BBB damage (p<0.05). In the 3 mouse strains, C57/BL6, C57/BL10, and C57/BL10HIIIR2, LPS significantly opened the BBB in C57/BL6, and it was the only strain that showed a reduction in BBB permeability with BB-94. Treatment with methylprednisolone and several broad spectrum MMPIs, including BB-1101, were ineffective in the C57/BL6. There was a significant reduction in BBB permeability seen with 10% dimethyl sulfoxide (DMSO) alone, which was used to dissolve the selective MMP-2 and -9 inhibitor, SB-3CT. The tetracycline derivative, minocycline, reduced the BBB injury in mouse by blocking the production of MMP-9. Our results show variability in rats and mice to LPS and MMPIs, which most likely is based on genetic make-up. Understanding these differences may provide important clues that could guide selection of MMPIs in treatment of neurological diseases. PMID:17184743

The regulation and regulatory role of collagenase in bone

NASA Technical Reports Server (NTRS)

Partridge, N. C.; Walling, H. W.; Bloch, S. R.; Omura, T. H.; Chan, P. T.; Pearman, A. T.; Chou, W. Y.

1996-01-01

Interstitial collagenase plays an important role in both the normal and pathological remodeling of collagenous extracellular matrices, including skeletal tissues. The enzyme is a member of the family of matrix metalloproteinases. Only one rodent interstitial collagenase has been found but there are two human enzymes, human collagenase-1 and -3, the latter being the homologue of the rat enzyme. In developing rat and mouse bone, collagenase is expressed by hypertrophic chondrocytes, osteoblasts, and osteocytes, a situation that is replicated in a fracture callus. Cultured osteoblasts derived from neonatal rat calvariae show greater amounts of collagenase transcripts late in differentiation. These levels can be regulated by parathyroid hormone (PTH), retinoic acid, and insulin-like growth factors, as well as the degree of matrix mineralization. Much of the work on collagenase in bone has been derived from studies on the rat osteosarcoma cell line, UMR 106-01. All bone-resorbing agents stimulate these cells to produce collagenase mRNA and protein, with PTH being the most potent stimulator. Determination of secreted levels of collagenase has been difficult because UMR cells, normal rat osteoblasts, and rat fibroblasts possess a scavenger receptor that removes the enzyme from the extracellular space, internalizes and degrades it, thus imposing another level of control. PTH can also regulate the abundance of the receptor as well as the expression and synthesis of the enzyme. Regulation of the collagenase gene by PTH appears to involve the cAMP pathway as well as a primary response gene, possibly Fos, which then contributes to induction of the collagenase gene. The rat collagenase gene contains an activator protein-1 sequence that is necessary for basal expression, but other promoter regions may also participate in PTH regulation. Thus, there are many levels of regulation of collagenase in bone perhaps constraining what would otherwise be a rampant enzyme.

Active vibrissal sensing in rodents and marsupials

PubMed Central

Mitchinson, Ben; Grant, Robyn A.; Arkley, Kendra; Rankov, Vladan; Perkon, Igor; Prescott, Tony J.

2011-01-01

In rats, the long facial whiskers (mystacial macrovibrissae) are repetitively and rapidly swept back and forth during exploration in a behaviour known as ‘whisking’. In this paper, we summarize previous evidence from rats, and present new data for rat, mouse and the marsupial grey short-tailed opossum (Monodelphis domestica) showing that whisking in all three species is actively controlled both with respect to movement of the animal's body and relative to environmental structure. Using automatic whisker tracking, and Fourier analysis, we first show that the whisking motion of the mystacial vibrissae, in the horizontal plane, can be approximated as a blend of two sinusoids at the fundamental frequency (mean 8.5, 11.3 and 7.3 Hz in rat, mouse and opossum, respectively) and its second harmonic. The oscillation at the second harmonic is particularly strong in mouse (around 22 Hz) consistent with previous reports of fast whisking in that species. In all three species, we found evidence of asymmetric whisking during head turning and following unilateral object contacts consistent with active control of whisker movement. We propose that the presence of active vibrissal touch in both rodents and marsupials suggests that this behavioural capacity emerged at an early stage in the evolution of therian mammals. PMID:21969685

Ectoparasites and other epifaunistic arthropods of sympatric cotton mice and golden mice: comparisons and implications for vector-borne zoonotic diseases.

PubMed

Durden, Lance A; Polur, Ram N; Nims, Todd; Banks, Craig W; Oliver, James H

2004-12-01

Ectoparasite and epifaunistic arthropod biodiversity and infestation parameters were compared between 2 sympatric small rodent species, the cotton mouse (Peromyscus gossypinus (Le Conte)) and golden mouse (Ochrotomys nuttalli (Harlan)), in southern Georgia from 1992 to 2003. Because the cotton mouse is known to be a reservoir of more vector-borne zoonotic pathogens than the golden mouse, we hypothesized that it would be parasitized by more ectoparasites that are known to be vectors of these pathogens. Cotton mice (n = 202) were parasitized by 19 species of arthropods, whereas golden mice (n = 46) were parasitized by 12 species. Eleven species of arthropods were recovered from both host species, whereas 7 were recorded only from cotton mice, and 1 species only from golden mice. Infestation prevalences (percent of mice parasitized) were significantly higher for 1 species of arthropod (the tropical rat mite Ornithonyssus bacoti (Hirst)) infesting cotton mice and for 4 species (the flea Peromyscopsylla scotti Fox and the mites Glycyphagus hypudaei Koch, Androlaelaps casalis (Berlese), and Androlaelaps fahrenholzi (Berlese)) infesting golden mice. Mean intensities (mean per infested mouse) were significantly higher for 2 species (the flea Orchopeas leucopus (Baker) and the blacklegged tick Ixodes scapularis Say) infesting cotton mice and for 2 species (G. hypudaei and A. fahrenholzi) infesting golden mice. Ectoparasites that are known to be vectors of zoonotic pathogens were significantly more common on cotton mice than on golden mice. These ectoparasites included the rhopalopsyllid flea Polygenis gwyni (Fox), a vector of the agent of murine typhus; I. scapularis, the principal vector of the agents of Lyme borreliosis, human granulocytic ehrlichiosis, and human babesiosis; and O. bacoti, a laboratory vector of several zoonotic pathogens. However, 2 species of ixodid ticks that can transmit zoonotic pathogens were recovered from both host species. These were the American dog tick Dermacentor variabilis (Say), the principal vector of the agent of Rocky Mountain spotted fever in eastern North America, and Ixodes minor Neumann, an enzootic vector of the agent of Lyme borreliosis. Overall, the cotton mouse was parasitized by significantly more ectoparasites that are known to be vectors of zoonotic pathogens than was the golden mouse. These data support the hypothesis that the cotton mouse has greater epidemiological importance for zoonotic vector-borne pathogen transmission than does the golden mouse.

Effects of Acer okamotoanum sap on the function of polymorphonuclear neutrophilic leukocytes in vitro and in vivo.

PubMed

An, Beum-Soo; Kang, Ji-Houn; Yang, Hyun; Yang, Mhan-Pyo; Jeung, Eui-Bae

2013-02-01

Sap is a plant fluid that primarily consists of water and small amounts of mineral elements, sugars, hormones and other nutrients. Acer mono (A. mono) is an endemic Korean mono maple which was recently suggested to have health benefits due to its abundant calcium and magnesium ion content. In the present study, we examined the effects of sap from Acer okamotoanum (A. okamotoanum) on the phagocytic response of mouse neutrophils in vivo and rat and canine neutrophils in vitro. We tested the regulation of phagocytic activity, oxidative burst activity (OBA) and the levels of filamentous polymeric actin (F-actin) in the absence and presence of dexamethasone (DEX) in vitro and in vivo. Our results showed that DEX primarily reduced OBA in the mouse neutrophils, and that this was reversed in the presence of the sap. By contrast, the phagocytic activity of the mouse cells was not regulated by either DEX or the sap. Rat and canine polymorphonuclear neutrophilic leukocytes (PMNs) responded in vitro to the sap in a similar manner by increasing OBA. However, regulation of phagocytic activity by the sap was different between the species. In canine PMNs, phagocytic activity was enhanced by the sap at a high dose, while it did not significantly modulate this activity in rat PMNs. These findings suggest that the sap of A. okamotoanum stimulates neutrophil activity in the mouse, rat and canine by increasing OBA in vivo and in vitro, and thus may have a potential antimicrobial effect in the PMNs of patients with infections.

Oral LD50 toxicity modeling and prediction of per- and polyfluorinated chemicals on rat and mouse.

PubMed

Bhhatarai, Barun; Gramatica, Paola

2011-05-01

Quantitative structure-activity relationship (QSAR) analyses were performed using the LD(50) oral toxicity data of per- and polyfluorinated chemicals (PFCs) on rodents: rat and mouse. PFCs are studied under the EU project CADASTER which uses the available experimental data for prediction and prioritization of toxic chemicals for risk assessment by using the in silico tools. The methodology presented here applies chemometrical analysis on the existing experimental data and predicts the toxicity of new compounds. QSAR analyses were performed on the available 58 mouse and 50 rat LD(50) oral data using multiple linear regression (MLR) based on theoretical molecular descriptors selected by genetic algorithm (GA). Training and prediction sets were prepared a priori from available experimental datasets in terms of structure and response. These sets were used to derive statistically robust and predictive (both internally and externally) models. The structural applicability domain (AD) of the models were verified on 376 per- and polyfluorinated chemicals including those in REACH preregistration list. The rat and mouse endpoints were predicted by each model for the studied compounds, and finally 30 compounds, all perfluorinated, were prioritized as most important for experimental toxicity analysis under the project. In addition, cumulative study on compounds within the AD of all four models, including two earlier published models on LC(50) rodent analysis was studied and the cumulative toxicity trend was observed using principal component analysis (PCA). The similarities and the differences observed in terms of descriptors and chemical/mechanistic meaning encoded by descriptors to prioritize the most toxic compounds are highlighted.

Comparative effects of sub-stimulating concentrations of non-human versus human Luteinizing Hormones (LH) or chorionic gonadotropins (CG) on adenylate cyclase activation by forskolin in MLTC cells.

PubMed

Nguyen, Thi-Mong Diep; Filliatreau, Laura; Klett, Danièle; Combarnous, Yves

2018-05-15

We have compared various Luteinizing Hormone (LH) and Chorionic Gonadotropin (CG) preparations from non-human and human species in their ability to synergize with 10 µM forskolin (FSK) for cyclic AMP intracellular accumulation, in MLTC cells. LH from rat pituitary as well as various isoforms of pituitary ovine, bovine, porcine, equine and human LHs and equine and human CG were studied. In addition, recombinant human LH and CG were also compared with the natural human and non-human hormones. Sub-stimulating concentrations of all LHs and CGs (2-100 pM) were found to stimulate cyclic AMP accumulation in MLTC cells in the presence of an also non-stimulating FSK concentration (10 µM). Like rat LH, the most homologous available hormone for mouse MLTC cells, all non-human LHs and CG exhibit a strong potentiating effect on FSK response. The human, natural and recombinant hLH and hCG also do so but in addition, they were found to elicit a permissive effect on FSK stimulation. Indeed, when incubated alone with MLTC cells at non-stimulating concentrations (2-70 pM) hLH and hCG permit, after being removed, a dose-dependent cyclic AMP accumulation with 10 µM FSK. Our data show a clearcut difference between human LH and CG compared to their non-human counterparts on MLTC cells adenylate cyclase activity control. This points out the risk of using hCG as a reference ligand for LHR in studies using non-human cells. Copyright © 2018 Elsevier Inc. All rights reserved.

DA-6034 Induces [Ca(2+)]i Increase in Epithelial Cells.

PubMed

Yang, Yu-Mi; Park, Soonhong; Ji, Hyewon; Kim, Tae-Im; Kim, Eung Kweon; Kang, Kyung Koo; Shin, Dong Min

2014-04-01

DA-6034, a eupatilin derivative of flavonoid, has shown potent effects on the protection of gastric mucosa and induced the increases in fluid and glycoprotein secretion in human and rat corneal and conjunctival cells, suggesting that it might be considered as a drug for the treatment of dry eye. However, whether DA-6034 induces Ca(2+) signaling and its underlying mechanism in epithelial cells are not known. In the present study, we investigated the mechanism for actions of DA-6034 in Ca(2+) signaling pathways of the epithelial cells (conjunctival and corneal cells) from human donor eyes and mouse salivary gland epithelial cells. DA-6034 activated Ca(2+)-activated Cl(-) channels (CaCCs) and increased intracellular calcium concentrations ([Ca(2+)]i) in primary cultured human conjunctival cells. DA-6034 also increased [Ca(2+)]i in mouse salivary gland cells and human corneal epithelial cells. [Ca(2+)]i increase of DA-6034 was dependent on the Ca(2+) entry from extracellular and Ca(2+) release from internal Ca(2+) stores. Interestingly, these effects of DA-6034 were related to ryanodine receptors (RyRs) but not phospholipase C/inositol 1,4,5-triphosphate (IP3) pathway and lysosomal Ca(2+) stores. These results suggest that DA-6034 induces Ca(2+) signaling via extracellular Ca(2+) entry and RyRs-sensitive Ca(2+) release from internal Ca(2+) stores in epithelial cells.

Osteoblast Specific Overexpression of Human Interleukin-7 Rescues the Bone Mass Phenotype of Interleukin-7 Deficient Female Mice

PubMed Central

Aguila, Hector L.; Mun, Se Hwan; Kalinowski, Judith; Adams, Douglas J.; Lorenzo, Joseph A.; Lee, Sun-Kyeong

2012-01-01

Interleukin-7 is a critical cytokine for lymphoid development and a direct inhibitor of in vitro osteoclastogenesis in murine bone marrow cultures. To explore the role of IL-7 in bone, we generated transgenic mouse lines bearing the 2.3 Kb rat collagen 1A1 promoter driving the expression of human IL-7 specifically in osteoblasts. In addition we crossed these mice with IL-7 deficient mice to determine if the alterations in lymphopoiesis, bone mass and osteoclast formation observed in the IL-7 KO mice could be rescued by osteoblast-specific overexpression of IL-7. Here we show that mice overexpressing human IL-7 in the osteoblast lineage demonstrated increased trabecular bone volume in vivo by µCT and decreased osteoclast formation in vitro. Furthermore, targeted overexpression of IL-7 in osteoblasts rescued the osteopenic bone phenotype and B cell development of IL-7 KO mice but did not have an effect on T lymphopoiesis, which occurs in the periphery. The bone phenotypes in IL-7 KO mice and targeted IL-7 overexpressing mouse models were observed only in females. These results likely reflect both a direct inhibitory effects of IL-7 on osteoclastogenesis in vivo and gender specific differences in responses to IL-7. PMID:22258693

Effect of diphenyl ether herbicides and oxadiazon on porphyrin biosynthesis in mouse liver, rat primary hepatocyte culture and HepG2 cells.

PubMed

Krijt, J; van Holsteijn, I; Hassing, I; Vokurka, M; Blaauboer, B J

1993-01-01

The effects of the herbicides fomesafen, oxyfluorfen, oxadiazon and fluazifop-butyl on porphyrin accumulation in mouse liver, rat primary hepatocyte culture and HepG2 cells were investigated. Ten days of herbicide feeding (0.25% in the diet) increased the liver porphyrins in male C57B1/6J mice from 1.4 +/- 0.6 to 4.8 +/- 2.1 (fomesafen) 16.9 +2- 2.9 (oxyfluorfen) and 25.9 +/- 3.1 (oxadiazon) nmol/g wet weight, respectively. Fluazifop-butyl had no effect on liver porphyrin metabolism. Fomesafen, oxyfluorfen and oxadiazon increased the cellular porphyrin content of rat hepatocytes after 24 h of incubation (control, 3.2 pmol/mg protein, fomesafen, oxyfluorfen and oxadiazon at 0.125 mM concentration 51.5, 54.3 and 44.0 pmol/mg protein, respectively). The porphyrin content of HepG2 cells increased from 1.6 to 18.2, 10.6 and 9.2 pmol/mg protein after 24 h incubation with the three herbicides. Fluazifop-butyl increased hepatic cytochrome P450 levels and ethoxy- and pentoxyresorufin O-dealkylase (EROD and PROD) activity, oxyfluorfen increased PROD activity. Peroxisomal palmitoyl CoA oxidation increased after fomesafen and fluazifop treatment to about 500% of control values both in mouse liver and rat hepatocytes. Both rat hepatocytes and HepG2 cells can be used as a test system for the porphyrogenic potential of photobleaching herbicides.

Evidence of NK1 and NK2 Tachykinin Receptors and their Involvement in Histamine Release in a Murine Mast Cell Line

DTIC Science & Technology

1992-01-01

either human p ~ulmo(nary,. Delectaible in the absence of estrmcclular CaCI’. i’Potent 4.23ug/105 cells, or rat peritoneal mast cells. bousbesin...ABSTRACT (Maximum 200 words) Abstract-Binding of )kH substance P (SP) and histamine release were examined using a cloned mouse mast cell line SP binding...the cells with the NK2 antagonist peptide A reduced NKA-induced histamine release ID.Arg’,D.Phe’,D-Trp 0 3 .Leu t )nsu b s tance P , a putative SP

The transcription factor encyclopedia.

PubMed

Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I; Bolotin, Eugene; Ticoll, Amy; Cheung, Warren A; Zhang, Xiao Yu Cindy; Dickman, Christopher T D; Fulton, Debra L; Lim, Jonathan S; Schnabl, Jake M; Ramos, Oscar H P; Vasseur-Cognet, Mireille; de Leeuw, Charles N; Simpson, Elizabeth M; Ryffel, Gerhart U; Lam, Eric W-F; Kist, Ralf; Wilson, Miranda S C; Marco-Ferreres, Raquel; Brosens, Jan J; Beccari, Leonardo L; Bovolenta, Paola; Benayoun, Bérénice A; Monteiro, Lara J; Schwenen, Helma D C; Grontved, Lars; Wederell, Elizabeth; Mandrup, Susanne; Veitia, Reiner A; Chakravarthy, Harini; Hoodless, Pamela A; Mancarelli, M Michela; Torbett, Bruce E; Banham, Alison H; Reddy, Sekhar P; Cullum, Rebecca L; Liedtke, Michaela; Tschan, Mario P; Vaz, Michelle; Rizzino, Angie; Zannini, Mariastella; Frietze, Seth; Farnham, Peggy J; Eijkelenboom, Astrid; Brown, Philip J; Laperrière, David; Leprince, Dominique; de Cristofaro, Tiziana; Prince, Kelly L; Putker, Marrit; del Peso, Luis; Camenisch, Gieri; Wenger, Roland H; Mikula, Michal; Rozendaal, Marieke; Mader, Sylvie; Ostrowski, Jerzy; Rhodes, Simon J; Van Rechem, Capucine; Boulay, Gaylor; Olechnowicz, Sam W Z; Breslin, Mary B; Lan, Michael S; Nanan, Kyster K; Wegner, Michael; Hou, Juan; Mullen, Rachel D; Colvin, Stephanie C; Noy, Peter John; Webb, Carol F; Witek, Matthew E; Ferrell, Scott; Daniel, Juliet M; Park, Jason; Waldman, Scott A; Peet, Daniel J; Taggart, Michael; Jayaraman, Padma-Sheela; Karrich, Julien J; Blom, Bianca; Vesuna, Farhad; O'Geen, Henriette; Sun, Yunfu; Gronostajski, Richard M; Woodcroft, Mark W; Hough, Margaret R; Chen, Edwin; Europe-Finner, G Nicholas; Karolczak-Bayatti, Magdalena; Bailey, Jarrod; Hankinson, Oliver; Raman, Venu; LeBrun, David P; Biswal, Shyam; Harvey, Christopher J; DeBruyne, Jason P; Hogenesch, John B; Hevner, Robert F; Héligon, Christophe; Luo, Xin M; Blank, Marissa Cathleen; Millen, Kathleen Joyce; Sharlin, David S; Forrest, Douglas; Dahlman-Wright, Karin; Zhao, Chunyan; Mishima, Yuriko; Sinha, Satrajit; Chakrabarti, Rumela; Portales-Casamar, Elodie; Sladek, Frances M; Bradley, Philip H; Wasserman, Wyeth W

2012-01-01

Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130 mini review articles on pertinent human, mouse and rat TFs. Notable features of the TFe website include a high-quality PDF generator and web API for programmatic data retrieval. TFe aims to rapidly educate scientists about the TFs they encounter through the delivery of succinct summaries written and vetted by experts in the field. TFe is available at http://www.cisreg.ca/tfe.

Evaulation of cancer and non-cancer effects of cumene ...

EPA Pesticide Factsheets

Cumene, also known as isopropyl benzene, is a volatile liquid. We have systematically reviewed published literature to evaluate cancer and noncancer effects of cumene. Cumene, readily absorbed via inhalation is distributed in several tissues, metabolized extensively by cytochrome P-450 isozymes within hepatic and extra-hepatic tissues and excreted through urine. Although, there are no epidemiological cancer studies for humans, chronic inhalation exposure studies in rat and mouse have shown increased nasal lesions including atrophy, basal cell hyperplasia, atypical hyperplasia and hyperplasia of the olfactory epithelium glands. To present the information at the Society of Toxicology Meeting.

Distribution of histaminergic neuronal cluster in the rat and mouse hypothalamus.

PubMed

Moriwaki, Chinatsu; Chiba, Seiichi; Wei, Huixing; Aosa, Taishi; Kitamura, Hirokazu; Ina, Keisuke; Shibata, Hirotaka; Fujikura, Yoshihisa

2015-10-01

Histidine decarboxylase (HDC) catalyzes the biosynthesis of histamine from L-histidine and is expressed throughout the mammalian nervous system by histaminergic neurons. Histaminergic neurons arise in the posterior mesencephalon during the early embryonic period and gradually develop into two histaminergic substreams around the lateral area of the posterior hypothalamus and the more anterior peri-cerebral aqueduct area before finally forming an adult-like pattern comprising five neuronal clusters, E1, E2, E3, E4, and E5, at the postnatal stage. This distribution of histaminergic neuronal clusters in the rat hypothalamus appears to be a consequence of neuronal development and reflects the functional differentiation within each neuronal cluster. However, the close linkage between the locations of histaminergic neuronal clusters and their physiological functions has yet to be fully elucidated because of the sparse information regarding the location and orientation of each histaminergic neuronal clusters in the hypothalamus of rats and mice. To clarify the distribution of the five-histaminergic neuronal clusters more clearly, we performed an immunohistochemical study using the anti-HDC antibody on serial sections of the rat hypothalamus according to the brain maps of rat and mouse. Our results confirmed that the HDC-immunoreactive (HDCi) neuronal clusters in the hypothalamus of rats and mice are observed in the ventrolateral part of the most posterior hypothalamus (E1), ventrolateral part of the posterior hypothalamus (E2), ventromedial part from the medial to the posterior hypothalamus (E3), periventricular part from the anterior to the medial hypothalamus (E4), and diffusely extended part of the more dorsal and almost entire hypothalamus (E5). The stereological estimation of the total number of HDCi neurons of each clusters revealed the larger amount of the rat than the mouse. The characterization of histaminergic neuronal clusters in the hypothalamus of rats and mice may provide useful information for further investigations. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Hair-bundle proteomes of avian and mammalian inner-ear utricles

PubMed Central

Wilmarth, Phillip A.; Krey, Jocelyn F.; Shin, Jung-Bum; Choi, Dongseok; David, Larry L.; Barr-Gillespie, Peter G.

2015-01-01

Examination of multiple proteomics datasets within or between species increases the reliability of protein identification. We report here proteomes of inner-ear hair bundles from three species (chick, mouse, and rat), which were collected on LTQ or LTQ Velos ion-trap mass spectrometers; the constituent proteins were quantified using MS2 intensities, which are the summed intensities of all peptide fragmentation spectra matched to a protein. The data are available via ProteomeXchange with identifiers PXD002410 (chick LTQ), PXD002414 (chick Velos), PXD002415 (mouse Velos), and PXD002416 (rat LTQ). The two chick bundle datasets compared favourably to a third, already-described chick bundle dataset, which was quantified using MS1 peak intensities, the summed intensities of peptides identified by high-resolution mass spectrometry (PXD000104; updated analysis in PXD002445). The mouse bundle dataset described here was comparable to a different mouse bundle dataset quantified using MS1 intensities (PXD002167). These six datasets will be useful for identifying the core proteome of vestibular hair bundles. PMID:26645194

Effect of Iboga Alkaloids on µ-Opioid Receptor-Coupled G Protein Activation

PubMed Central

Antonio, Tamara; Childers, Steven R.; Rothman, Richard B.; Dersch, Christina M.; King, Christine; Kuehne, Martin; Bornmann, William G.; Eshleman, Amy J.; Janowsky, Aaron; Simon, Eric R.; Reith, Maarten E. A.; Alper, Kenneth

2013-01-01

Objective The iboga alkaloids are a class of small molecules defined structurally on the basis of a common ibogamine skeleton, some of which modify opioid withdrawal and drug self-administration in humans and preclinical models. These compounds may represent an innovative approach to neurobiological investigation and development of addiction pharmacotherapy. In particular, the use of the prototypic iboga alkaloid ibogaine for opioid detoxification in humans raises the question of whether its effect is mediated by an opioid agonist action, or if it represents alternative and possibly novel mechanism of action. The aim of this study was to independently replicate and extend evidence regarding the activation of μ-opioid receptor (MOR)-related G proteins by iboga alkaloids. Methods Ibogaine, its major metabolite noribogaine, and 18-methoxycoronaridine (18-MC), a synthetic congener, were evaluated by agonist-stimulated guanosine-5´-O-(γ-thio)-triphosphate ([35S]GTPγS) binding in cells overexpressing the recombinant MOR, in rat thalamic membranes, and autoradiography in rat brain slices. Results And Significance In rat thalamic membranes ibogaine, noribogaine and 18-MC were MOR antagonists with functional Ke values ranging from 3 uM (ibogaine) to 13 uM (noribogaine and 18MC). Noribogaine and 18-MC did not stimulate [35S]GTPγS binding in Chinese hamster ovary cells expressing human or rat MORs, and had only limited partial agonist effects in human embryonic kidney cells expressing mouse MORs. Ibogaine did not did not stimulate [35S]GTPγS binding in any MOR expressing cells. Noribogaine did not stimulate [35S]GTPγS binding in brain slices using autoradiography. An MOR agonist action does not appear to account for the effect of these iboga alkaloids on opioid withdrawal. Taken together with existing evidence that their mechanism of action also differs from that of other non-opioids with clinical effects on opioid tolerance and withdrawal, these findings suggest a novel mechanism of action, and further justify the search for alternative targets of iboga alkaloids. PMID:24204784

Effect of Iboga alkaloids on µ-opioid receptor-coupled G protein activation.

PubMed

Antonio, Tamara; Childers, Steven R; Rothman, Richard B; Dersch, Christina M; King, Christine; Kuehne, Martin; Bornmann, William G; Eshleman, Amy J; Janowsky, Aaron; Simon, Eric R; Reith, Maarten E A; Alper, Kenneth

2013-01-01

The iboga alkaloids are a class of small molecules defined structurally on the basis of a common ibogamine skeleton, some of which modify opioid withdrawal and drug self-administration in humans and preclinical models. These compounds may represent an innovative approach to neurobiological investigation and development of addiction pharmacotherapy. In particular, the use of the prototypic iboga alkaloid ibogaine for opioid detoxification in humans raises the question of whether its effect is mediated by an opioid agonist action, or if it represents alternative and possibly novel mechanism of action. The aim of this study was to independently replicate and extend evidence regarding the activation of μ-opioid receptor (MOR)-related G proteins by iboga alkaloids. Ibogaine, its major metabolite noribogaine, and 18-methoxycoronaridine (18-MC), a synthetic congener, were evaluated by agonist-stimulated guanosine-5´-O-(γ-thio)-triphosphate ([(35)S]GTPγS) binding in cells overexpressing the recombinant MOR, in rat thalamic membranes, and autoradiography in rat brain slices. In rat thalamic membranes ibogaine, noribogaine and 18-MC were MOR antagonists with functional Ke values ranging from 3 uM (ibogaine) to 13 uM (noribogaine and 18MC). Noribogaine and 18-MC did not stimulate [(35)S]GTPγS binding in Chinese hamster ovary cells expressing human or rat MORs, and had only limited partial agonist effects in human embryonic kidney cells expressing mouse MORs. Ibogaine did not did not stimulate [(35)S]GTPγS binding in any MOR expressing cells. Noribogaine did not stimulate [(35)S]GTPγS binding in brain slices using autoradiography. An MOR agonist action does not appear to account for the effect of these iboga alkaloids on opioid withdrawal. Taken together with existing evidence that their mechanism of action also differs from that of other non-opioids with clinical effects on opioid tolerance and withdrawal, these findings suggest a novel mechanism of action, and further justify the search for alternative targets of iboga alkaloids.

Protection by meningococcal outer membrane protein PorA-specific antibodies and a serogroup B capsular polysaccharide-specific antibody in complement-sufficient and C6-deficient infant rats.

PubMed

Toropainen, Maija; Saarinen, Leena; Vidarsson, Gestur; Käyhty, Helena

2006-05-01

The relative contributions of antibody-induced complement-mediated bacterial lysis and antibody/complement-mediated phagocytosis to host immunity against meningococcal infections are currently unclear. Further, the in vivo effector functions of antibodies may vary depending on their specificity and Fc heavy-chain isotype. In this study, a mouse immunoglobulin G2a (mIgG2a) monoclonal antibody (MN12H2) to meningococcal outer membrane protein PorA (P1.16), its human IgG subclass derivatives (hIgG1 to hIgG4), and an mIgG2a monoclonal antibody (Nmb735) to serogroup B capsular polysaccharide (B-PS) were evaluated for passive protection against meningococcal serogroup B strain 44/76-SL (B:15:P1.7,16) in an infant rat infection model. Complement component C6-deficient (PVG/c-) rats were used to assess the importance of complement-mediated bacterial lysis for protection. The PorA-specific parental mIgG2a and the hIgG1 to hIgG3 derivatives all induced efficient bactericidal activity in vitro in the presence of human or infant rat complement and augmented bacterial clearance in complement-sufficient HsdBrlHan:WIST rats, while the hIgG4 was unable to do so. In C6-deficient PVG/c- rats, lacking complement-mediated bacterial lysis, the augmentation of bacterial clearance by PorA-specific mIgG2a and hIgG1 antibodies was impaired compared to that in the syngeneic complement-sufficient PVG/c+ rat strain. This was in contrast to the case for B-PS-specific mIgG2a, which conferred similar protective activity in both rat strains. These data suggest that while anti-B-PS antibody can provide protection in the infant rats without membrane attack complex formation, the protection afforded by anti-PorA antibody is more dependent on the activation of the whole complement pathway and subsequent bacterial lysis.

Peromyscus as a model system for human hepatitis C: An opportunity to advance our understanding of a complex host parasite system.

PubMed

Vandegrift, Kurt J; Critchlow, Justin T; Kapoor, Amit; Friedman, David A; Hudson, Peter J

2017-01-01

Worldwide, there are 185 million people infected with hepatitis C virus and approximately 350,000 people die each year from hepatitis C associated liver diseases. Human hepatitis C research has been hampered by the lack of an appropriate in vivo model system. Most of the in vivo research has been conducted on chimpanzees, which is complicated by ethical concerns, small sample sizes, high costs, and genetic heterogeneity. The house mouse system has led to greater understanding of a wide variety of human pathogens, but it is unreasonable to expect Mus musculus to be a good model system for every human pathogen. Alternative animal models can be developed in these cases. Ferrets (influenza), cotton rats (human respiratory virus), and woodchucks (hepatitis B) are all alternative models that have led to a greater understanding of human pathogens. Rodent models are tractable, genetically amenable and inbred and outbred strains can provide homogeneity in results. Recently, a rodent homolog of hepatitis C was discovered and isolated from the liver of a Peromyscus maniculatus. This represents the first small mammal (mouse) model system for human hepatitis C and it offers great potential to contribute to our understanding and ultimately aid in our efforts to combat this serious public health concern. Peromyscus are available commercially and can be used to inform questions about the origin, transmission, persistence, pathology, and rational treatment of hepatitis C. Here, we provide a disease ecologist's overview of this new virus and some suggestions for useful future experiments. Copyright © 2016. Published by Elsevier Ltd.

Microspectroscopic Study of Liposome-to-cell Interaction Revealed by Förster Resonance Energy Transfer.

PubMed

Yefimova, Svetlana L; Kurilchenko, Irina Yu; Tkacheva, Tatyana N; Kavok, Nataliya S; Todor, Igor N; Lukianova, Nataliya Yu; Chekhun, Vasyl F; Malyukin, Yuriy V

2014-03-01

We report the Förster resonance energy transfer (FRET)-labeling of liposomal vesicles as an effective approach to study in dynamics the interaction of liposomes with living cells of different types (rat hepatocytes, rat bone marrow, mouse fibroblast-like cells and human breast cancer cells) and cell organelles (hepatocyte nuclei). The in vitro experiments were performed using fluorescent microspectroscopic technique. Two fluorescent dyes (DiO as the energy donor and DiI as an acceptor) were preloaded in lipid bilayers of phosphatidylcholine liposomes that ensures the necessary distance between the dyes for effective FRET. The change in time of the donor and acceptor relative fluorescence intensities was used to visualize and trace the liposome-to-cell interaction. We show that FRET-labeling of liposome vesicles allows one to reveal the differences in efficiency and dynamics of these interactions, which are associated with composition, fluidity, and metabolic activity of cell plasma membranes.

CYP1A1 and CYP1A2 expression: Comparing 'humanized' mouse lines and wild-type mice; comparing human and mouse hepatoma-derived cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Uno, Shigeyuki; Endo, Kaori; Ishida, Yuji

2009-05-15

Human and rodent cytochrome P450 (CYP) enzymes sometimes exhibit striking species-specific differences in substrate preference and rate of metabolism. Human risk assessment of CYP substrates might therefore best be evaluated in the intact mouse by replacing mouse Cyp genes with human CYP orthologs; however, how 'human-like' can human gene expression be expected in mouse tissues? Previously a bacterial-artificial-chromosome-transgenic mouse, carrying the human CYP1A1{sub C}YP1A2 locus and lacking the mouse Cyp1a1 and Cyp1a2 orthologs, was shown to express robustly human dioxin-inducible CYP1A1 and basal versus inducible CYP1A2 (mRNAs, proteins, enzyme activities) in each of nine mouse tissues examined. Chimeric mice carryingmore » humanized liver have also been generated, by transplanting human hepatocytes into a urokinase-type plasminogen activator(+/+){sub s}evere-combined-immunodeficiency (uPA/SCID) line with most of its mouse hepatocytes ablated. Herein we compare basal and dioxin-induced CYP1A mRNA copy numbers, protein levels, and four enzymes (benzo[a]pyrene hydroxylase, ethoxyresorufin O-deethylase, acetanilide 4-hydroxylase, methoxyresorufin O-demethylase) in liver of these two humanized mouse lines versus wild-type mice; we also compare these same parameters in mouse Hepa-1c1c7 and human HepG2 hepatoma-derived established cell lines. Most strikingly, mouse liver CYP1A1-specific enzyme activities are between 38- and 170-fold higher than human CYP1A1-specific enzyme activities (per unit of mRNA), whereas mouse versus human CYP1A2 enzyme activities (per unit of mRNA) are within 2.5-fold of one another. Moreover, both the mouse and human hepatoma cell lines exhibit striking differences in CYP1A mRNA levels and enzyme activities. These findings are relevant to risk assessment involving human CYP1A1 and CYP1A2 substrates, when administered to mice as environmental toxicants or drugs.« less

Sulforaphane is not an effective antagonist of the human pregnane X-receptor in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Poulton, Emma Jane; Department of Environmental and Occupational Health Sciences, University of Washington; Levy, Lisa

2013-01-01

Sulforaphane (SFN), is an effective in vitro antagonist of ligand activation of the human pregnane and xenobiotic receptor (PXR). PXR mediated CYP3A4 up-regulation is implicated in adverse drug-drug interactions making identification of small molecule antagonists a desirable therapeutic goal. SFN is not an antagonist to mouse or rat PXR in vitro; thus, normal rodent species are not suitable as in vivo models for human response. To evaluate whether SFN can effectively antagonize ligand activation of human PXR in vivo, a three-armed, randomized, crossover trial was conducted with 24 healthy adults. The potent PXR ligand — rifampicin (300 mg/d) was givenmore » alone for 7 days in arm 1, or in daily combination with 450 μmol SFN (Broccoli Sprout extract) in arm 2; SFN was given alone in arm 3. Midazolam as an in vivo phenotype marker of CYP3A was administered before and after each treatment arm. Rifampicin alone decreased midazolam AUC by 70%, indicative of the expected increase in CYP3A4 activity. Co-treatment with SFN did not reduce CYP3A4 induction. Treatment with SFN alone also did not affect CYP3A4 activity in the cohort as a whole, although in the subset with the highest basal CYP3A4 activity there was a statistically significant increase in midazolam AUC (i.e., decrease in CYP3A4 activity). A parallel study in humanized PXR mice yielded similar results. The parallel effects of SFN between humanized PXR mice and human subjects demonstrate the predictive value of humanized mouse models in situations where species differences in ligand-receptor interactions preclude the use of a native mouse model for studying human ligand-receptor pharmacology. -- Highlights: ► The effects of SFN on PXR mediated CYP3A4 induction in humanized PXR mice and humans were examined. ► SFN had no effect on rifampicin mediated CYP3A4 induction in humans or humanized mice. ► SFN had a modest effect on basal CYP3A4 activity among subjects with higher baseline activity. ► Humanized PXR mice were generally predictive of the in vivo human response.« less

Proceedings of the 2016 National Toxicology Program Satellite Symposium

PubMed Central

Elmore, Susan A.; Chen, Vivian S.; Hayes-Bouknight, Schantel; Hoane, Jessica S.; Janardhan, Kyathanahalli; Kooistra, Linda H.; Nolte, Thomas; Szabo, Kathleen A.; Willson, Gabrielle A.; Wolf, Jeffrey C.; Malarkey, David E.

2016-01-01

The 2016 annual National Toxicology Program (NTP) Satellite Symposium, entitled “Pathology Potpourri” was held in San Diego, California, at the Society of Toxicologic Pathology’s (STP) 35th annual meeting. The goal of this symposium was to present and discuss challenging diagnostic pathology and/or nomenclature issues. This article presents summaries of the speakers’ talks, along with select images that were used by the audience for voting and discussion. Some lesions and topics covered during the symposium included malignant glioma and histiocytic sarcoma in the rodent brain; a new statistical method designed for histopathology data evaluation; uterine stromal/glandular polyp in a rat; malignant plasma cell tumor in a mouse brain; Schwann cell proliferative lesions in rat hearts; axillary schwannoma in a cat; necrosis and granulomatous inflammation in a rat brain; adenoma/carcinoma in a rat adrenal gland; hepatocyte maturation defect and liver/spleen hematopoietic defects in an embryonic mouse; distinguishing malignant glioma, malignant mixed glioma and malignant oligodendroglioma in the rat; comparison of mammary gland whole mounts and histopathology from mice; and discussion of the International Harmonization of Nomenclature and Diagnostic Criteria (INHAND) collaborations. PMID:27821709

Proceedings of the 2016 National Toxicology Program Satellite Symposium.

PubMed

Elmore, Susan A; Chen, Vivian S; Hayes-Bouknight, Schantel; Hoane, Jessica S; Janardhan, Kyathanahalli; Kooistra, Linda H; Nolte, Thomas; Szabo, Kathleen A; Willson, Gabrielle A; Wolf, Jeffrey C; Malarkey, David E

2017-01-01

The 2016 annual National Toxicology Program Satellite Symposium, entitled "Pathology Potpourri" was held in San Diego, CA, at the Society of Toxicologic Pathology's (STP) 35th annual meeting. The goal of this symposium was to present and discuss challenging diagnostic pathology and/or nomenclature issues. This article presents summaries of the speakers' talks, along with select images that were used by the audience for voting and discussion. Some lesions and topics covered during the symposium included malignant glioma and histiocytic sarcoma in the rodent brain; a new statistical method designed for histopathology data evaluation; uterine stromal/glandular polyp in a rat; malignant plasma cell tumor in a mouse brain; Schwann cell proliferative lesions in rat hearts; axillary schwannoma in a cat; necrosis and granulomatous inflammation in a rat brain; adenoma/carcinoma in a rat adrenal gland; hepatocyte maturation defect and liver/spleen hematopoietic defects in an embryonic mouse; distinguishing malignant glioma, malignant mixed glioma, and malignant oligodendroglioma in the rat; comparison of mammary gland whole mounts and histopathology from mice; and discussion of the International Harmonization of Nomenclature and Diagnostic Criteria collaborations.

CYP1A1 and CYP1A2 expression: Comparing ‘humanized’ mouse lines and wild-type mice; comparing human and mouse hepatoma-derived cell lines

PubMed Central

Uno, Shigeyuki; Endo, Kaori; Ishida, Yuji; Tateno, Chise; Makishima, Makoto; Yoshizato, Katsutoshi; Nebert, Daniel W.

2009-01-01

Human and rodent cytochrome P450 (CYP) enzymes sometimes exhibit striking species-specific differences in substrate preference and rate of metabolism. Human risk assessment of CYP substrates might therefore best be evaluated in the intact mouse by replacing mouse Cyp genes with human CYP orthologs; however, how “human-like” can human gene expression be expected in mouse tissues? Previously a bacterial-artificial-chromosome-transgenic mouse, carrying the human CYP1A1_CYP1A2 locus and lacking the mouse Cyp1a1 and Cyp1a2 orthologs, was shown to express robustly human dioxin-inducible CYP1A1 and basal versus inducible CYP1A2 (mRNAs, proteins, enzyme activities) in each of nine mouse tissues examined. Chimeric mice carrying humanized liver have also been generated, by transplanting human hepatocytes into a urokinase-type plasminogen activator(+/+)_severe-combined-immunodeficiency (uPA/SCID) line with most of its mouse hepatocytes ablated. Herein we compare basal and dioxin-induced CYP1A mRNA copy numbers, protein levels, and four enzymes (benzo[a]pyrene hydroxylase, ethoxyresorufin O-deethylase, acetanilide 4-hydroxylase, methoxyresorufin O-demethylase) in liver of these two humanized mouse lines versus wild-type mice; we also compare these same parameters in mouse Hepa-1c1c7 and human HepG2 hepatoma-derived established cell lines. Most strikingly, mouse liver CYP1A1-specific enzyme activities are between 38- and 170-fold higher than human CYP1A1-specific enzyme activities (per unit of mRNA), whereas mouse versus human CYP1A2 enzyme activities (per unit of mRNA) are within 2.5-fold of one another. Moreover, both the mouse and human hepatoma cell lines exhibit striking differences in CYP1A mRNA levels and enzyme activities. These findings are relevant to risk assessment involving human CYP1A1 and CYP1A2 substrates, when administered to mice as environmental toxicants or drugs. PMID:19285097

IMPROVED TEMPERATURE GRADIENT FOR MONITORING BEHAVIORAL THERMOREGULATION IN THE RAT

EPA Science Inventory

Past studies have found that the laboratory rat placed In a temperature gradient prefers temperatures that are markedly below its lower critical ambient temperature (LCT), whereas other rodents (e.g., mouse, hamster, and guinea pig) generally select thermal environments associate...

HBV life cycle is restricted in mouse hepatocytes expressing human NTCP.

PubMed

Li, Hanjie; Zhuang, Qiuyu; Wang, Yuze; Zhang, Tianying; Zhao, Jinghua; Zhang, Yali; Zhang, Junfang; Lin, Yi; Yuan, Quan; Xia, Ningshao; Han, Jiahuai

2014-03-01

Recent studies have revealed that human sodium taurocholate cotransporting polypeptide (SLC10A1 or NTCP) is a functional cellular receptor for hepatitis B virus (HBV). However, whether human NTCP can support HBV infection in mouse hepatocyte cell lines has not been clarified. Because an HBV-permissible mouse model would be helpful for the study of HBV pathogenesis, it is necessary to investigate whether human NTCP supports the susceptibility of mouse hepatocyte cell lines to HBV. The results show that exogenous human NTCP expression can render non-susceptible HepG2 (human), Huh7 (human), Hepa1-6 (mouse), AML-12 (mouse) cell lines and primary mouse hepatocyte (PMH) cells susceptible to hepatitis D virus (HDV) which employs HBV envelope proteins. However, human NTCP could only introduce HBV susceptibility in human-derived HepG2 and Huh7 cells, but not in mouse-derived Hepa1-6, AML-12 or PMH cells. These data suggest that although human NTCP is a functional receptor that mediates HBV infection in human cells, it cannot support HBV infection in mouse hepatocytes. Our study indicated that the restriction of HBV in mouse hepatocytes likely occurs after viral entry but prior to viral transcription. We have excluded the role of mouse hepatocyte nuclear factors in the restriction of the HBV life cycle and showed that knockdown or inhibition of Sting, TBK1, IRF3 or IRF7, the components of the anti-viral signaling pathways, had no effect on HBV infection in mouse hepatocytes. Therefore, murine restriction factors that limit HBV infection need to be identified before a HBV-permissible mouse line can be created.

Small animal models of cardiovascular disease: tools for the study of the roles of metabolic syndrome, dyslipidemia, and atherosclerosis.

PubMed

Russell, James C; Proctor, Spencer D

2006-01-01

Cardiovascular disease, the leading cause of death in much of the modern world, is the common symptomatic end stage of a number of distinct diseases and, therefore, is multifactorial and polygenetic in character. The two major underlying causes are disorders of lipid metabolism and metabolic syndrome. The ability to develop preventative and ameliorative treatments will depend on animal models that mimic human disease processes. The focus of this review is to identify suitable animal models and insights into cardiovascular disease achieved to date using such models. The ideal animal model of cardiovascular disease will mimic the human subject metabolically and pathophysiologically, will be large enough to permit physiological and metabolic studies, and will develop end-stage disease comparable to those in humans. Given the complex multifactorial nature of cardiovascular disease, no one species will be suitable for all studies. Potential larger animal models are problematic due to cost, ethical considerations, or poor pathophysiological comparability to humans. Rabbits require high-cholesterol diets to develop cardiovascular disease, and there are no rabbit models of metabolic syndrome. Spontaneous mutations in rats provide several complementary models of obesity, hyperlipidemia, insulin resistance, and type 2 diabetes, one of which spontaneously develops cardiovascular disease and ischemic lesions. The mouse, like normal rats, is characteristically resistant to cardiovascular disease, although genetically altered strains respond to cholesterol feeding with atherosclerosis, but not with end-stage ischemic lesions. The most useful and valid species/strains for the study of cardiovascular disease appear to be small rodents, rats, and mice. This fragmented field would benefit from a consensus on well-characterized appropriate models for the study of different aspects of cardiovascular disease and a renewed emphasis on the biology of underlying diseases.

PCR detection and quantitation of predominant anaerobic bacteria in human and animal fecal samples.

PubMed Central

Wang, R F; Cao, W W; Cerniglia, C E

1996-01-01

PCR procedures based on 16S rRNA gene sequences specific for 12 anaerobic bacteria that predominate in the human intestinal tract were developed and used for quantitative detection of these species in human (adult and baby) feces and animal (rat, mouse, cat, dog, monkey, and rabbit) feces. Fusobacterium prausnitzii, Peptostreptococcus productus, and Clostridium clostridiiforme had high PCR titers (the maximum dilutions for positive PCR results ranged from 10(-3) to 10(-8)) in all of the human and animal fecal samples tested. Bacteroides thetaiotaomicron, Bacteroides vulgatus, and Eubacterium limosum also showed higher PCR titers (10(-2) to 10(-6)) in adult human feces. The other bacteria tested, including Escherichia coli, Bifidobacterium adolescentis, Bifidobacterium longum, Lactobacillus acidophilus, Eubacterium biforme, and Bacteroides distasonis, were either at low PCR titers (less than 10(-2)) or not detected by PCR. The reported PCR procedure including the fecal sample preparation method is simplified and rapid and eliminates the DNA isolation steps. PMID:8919784

The clinical implications of mouse models of enhanced anxiety

PubMed Central

Sartori, Simone B; Landgraf, Rainer; Singewald, Nicolas

2011-01-01

Mice are increasingly overtaking the rat model organism in important aspects of anxiety research, including drug development. However, translating the results obtained in mouse studies into information that can be applied in clinics remains challenging. One reason may be that most of the studies so far have used animals displaying ‘normal’ anxiety rather than ‘psychopathological’ animal models with abnormal (elevated) anxiety, which more closely reflect core features and sensitivities to therapeutic interventions of human anxiety disorders, and which would, thus, narrow the translational gap. Here, we discuss manipulations aimed at persistently enhancing anxiety-related behavior in the laboratory mouse using phenotypic selection, genetic techniques and/or environmental manipulations. It is hoped that such models with enhanced construct validity will provide improved ways of studying the neurobiology and treatment of pathological anxiety. Examples of findings from mouse models of enhanced anxiety-related behavior will be discussed, as well as their relation to findings in anxiety disorder patients regarding neuroanatomy, neurobiology, genetic involvement and epigenetic modifications. Finally, we highlight novel targets for potential anxiolytic pharmacotherapeutics that have been established with the help of research involving mice. Since the use of psychopathological mouse models is only just beginning to increase, it is still unclear as to the extent to which such approaches will enhance the success rate of drug development in translating identified therapeutic targets into clinical trials and, thus, helping to introduce the next anxiolytic class of drugs. PMID:21901080

Minimum datasets to establish a CAR-mediated mode of action for rodent liver tumors.

PubMed

Peffer, Richard C; LeBaron, Matthew J; Battalora, Michael; Bomann, Werner H; Werner, Christoph; Aggarwal, Manoj; Rowe, Rocky R; Tinwell, Helen

2018-04-16

Methods for investigating the Mode of Action (MoA) for rodent liver tumors via constitutive androstane receptor (CAR) activation are outlined here, based on current scientific knowledge about CAR and feedback from regulatory agencies globally. The key events (i.e., CAR activation, altered gene expression, cell proliferation, altered foci and increased adenomas/carcinomas) can be demonstrated by measuring a combination of key events and associative events that are markers for the key events. For crop protection products, a primary dataset typically should include a short-term study in the species/strain that showed the tumor response at dose levels that bracket the tumorigenic and non-tumorigenic dose levels. The dataset may vary depending on the species and the test compound. As examples, Case Studies with nitrapyrin (in mice) and metofluthrin (in rats) are described. Based on qualitative differences between the species, the key events leading to tumors in mice or rats by this MoA are not operative in humans. In the future, newer approaches such as a CAR biomarker signature approach and/or in vitro CAR3 reporter assays for mouse, rat and human CAR may eventually be used to demonstrate a CAR MoA is operative, without the need for extensive additional studies in laboratory animals. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Immunolocalization of ciliary neurotrophic factor receptor alpha (CNTFRalpha) in mammalian photoreceptor cells.

PubMed

Beltran, William A; Rohrer, Hermann; Aguirre, Gustavo D

2005-04-01

To characterize the site of expression of the alpha subunit of the receptor for ciliary neurotrophic factor (CNTFRalpha) in the retina of a variety of mammalian species, and determine whether CNTFRalpha is localized to photoreceptor cells. The cellular distribution of CNTFRalpha(protein) was examined by immunocytochemistry in the adult retinas of several mammalian species that included mouse, rat, dog, cat, sheep, pig, horse, monkey, and human. Developing retinas from 3-day-old and 6-day-old rats were also included in this study. The molecular weight of CNTFRalpha in rat, dog, cat, pig, and human retinas was determined by immunoblotting. CNTFRalpha immunolabeling was present in the retina of all species. A common pattern was observed in all species, and represented labeling of the nerve fiber layer (NFL), ganglion cell layer (GCL), inner plexiform layer (IPL), inner nuclear layer (INL), and outer plexiform layer (OPL). CNTFRalpha did not immunolocalize to photoreceptor cells in both adult and developing rodent retinas, but was consistently observed in both rods and cones of non-rodent species. The molecular weight of CNTFRalpha in mammalian retinas was approximately 61-64 kDa. These findings highlight a significant difference in the expression of CNTFRalpha in the retina of rodent and non-rodent mammalian species. The expression of CNTFRalpha by rods and cones in non-rodent species may suggest a direct mechanism of action if CNTF administration results in photoreceptor rescue.

Intravitreal Injection of Proinsulin-Loaded Microspheres Delays Photoreceptor Cell Death and Vision Loss in the rd10 Mouse Model of Retinitis Pigmentosa.

PubMed

Isiegas, Carolina; Marinich-Madzarevich, Jorge A; Marchena, Miguel; Ruiz, José M; Cano, María J; de la Villa, Pedro; Hernández-Sánchez, Catalina; de la Rosa, Enrique J; de Pablo, Flora

2016-07-01

The induction of proinsulin expression by transgenesis or intramuscular gene therapy has been shown previously to retard retinal degeneration in mouse and rat models of retinitis pigmentosa (RP), a group of inherited conditions that result in visual impairment. We investigated whether intraocular treatment with biodegradable poly (lactic-co-glycolic) acid microspheres (PLGA-MS) loaded with proinsulin has cellular and functional neuroprotective effects in the retina. Experiments were performed using the Pde6brd10 mouse model of RP. Methionylated human recombinant proinsulin (hPI) was formulated in PLGA-MS, which were administered by intravitreal injection on postnatal days (P) 14 to 15. Retinal neuroprotection was assessed at P25 by electroretinography, and by evaluating outer nuclear layer (ONL) cellular preservation. The attenuation of photoreceptor cell death by hPI was determined by TUNEL assay in cultured P22 retinas, as well as Akt phosphorylation by immunoblotting. We successfully formulated hPI PLGA-MS to deliver the active molecule for several weeks in vitro. The amplitude of b-cone and mixed b-waves in electroretinographic recording was significantly higher in eyes injected with hPI-PLGA-MS compared to control eyes. Treatment with hPI-PLGA-MS attenuated photoreceptor cell loss, as revealed by comparing ONL thickness and the number of cell rows in this layer in treated versus untreated retinas. Finally, hPI prevented photoreceptor cell death and increased AktThr308 phosphorylation in organotypic cultured retinas. Retinal degeneration in the rd10 mouse was slowed by a single intravitreal injection of hPI-PLGA-MS. Human recombinant proinsulin elicited a rapid and effective neuroprotective effect when administered in biodegradable microspheres, which may constitute a future potentially feasible delivery method for proinsulin-based treatment of RP.

Neuroprotection with a brain-penetrating biologic tumor necrosis factor inhibitor.

PubMed

Zhou, Qing-Hui; Sumbria, Rachita; Hui, Eric Ka-Wai; Lu, Jeff Zhiqiang; Boado, Ruben J; Pardridge, William M

2011-11-01

Biologic tumor necrosis factor (TNF)-α inhibitors do not cross the blood-brain barrier (BBB). A BBB-penetrating TNF-α inhibitor was engineered by fusion of the extracellular domain of the type II human TNF receptor (TNFR) to the carboxyl terminus of the heavy chain of a mouse/rat chimeric monoclonal antibody (MAb) against the mouse transferrin receptor (TfR), and this fusion protein is designated cTfRMAb-TNFR. The cTfRMAb-TNFR fusion protein and etanercept bound human TNF-α with high affinity and K(D) values of 374 ± 77 and 280 ± 80 pM, respectively. Neuroprotection in brain in vivo after intravenous administration of the fusion protein was examined in a mouse model of Parkinson's disease. Mice were also treated with saline or a non-BBB-penetrating TNF decoy receptor, etanercept. After intracerebral injection of the nigral-striatal toxin, 6-hydroxydopamine, mice were treated every other day for 3 weeks. Treatment with the cTfRMAb-TNFR fusion protein caused an 83% decrease in apomorphine-induced rotation, a 67% decrease in amphetamine-induced rotation, a 82% increase in vibrissae-elicited forelimb placing, and a 130% increase in striatal tyrosine hydroxylase (TH) enzyme activity. In contrast, chronic treatment with etanercept, which does not cross the BBB, had no effect on neurobehavior or striatal TH enzyme activity. A bridging enzyme-linked immunosorbent assay specific for the cTfRMAb-TNFR fusion protein showed that the immune response generated in the mice was low titer. In conclusion, a biologic TNF inhibitor is neuroprotective after intravenous administration in a mouse model of neurodegeneration, providing that the TNF decoy receptor is reengineered to cross the BBB.

CYP2F2-generated metabolites, not styrene oxide, are a key event mediating the mode of action of styrene-induced mouse lung tumors.

PubMed

Cruzan, G; Bus, J; Hotchkiss, J; Harkema, J; Banton, M; Sarang, S

2012-02-01

Styrene induces lung tumors in mice but not in rats. Although metabolism of styrene to 7,8-styrene oxide (SO) by CYP2E1 has been suggested as a mediator of styrene toxicity, lung toxicity is not attenuated in CYP2E1 knockout mice. However, styrene and/or SO metabolism by mouse lung Clara cell-localized CYP2F2 to ring-oxidized cytotoxic metabolite(s) has been postulated as a key metabolic gateway responsible for both lung toxicity and possible tumorigenicity. To test this hypothesis, the lung toxicity of styrene and SO was evaluated in C57BL/6 (WT) and CYP2F2⁻/⁻ knockout mice treated with styrene (400 mg/kg/day, gavage, or 200 or 400 mg/kg/day, ip) or S- or R-SO (200 mg/kg/day, ip) for 5 days. Styrene treated WT mice displayed significant necrosis and exfoliation of Clara cells, and cumulative BrdU-labeling index of S-phase cells was markedly increased in terminal bronchioles of WT mice exposed to styrene or S- or RSO. In contrast, Clara and terminal bronchiole cell toxicity was not observed in CYP2F2⁻/⁻ mice exposed to either styrene or SO. This study clearly demonstrates that the mouse lung toxicity of both styrene and SO is critically dependent on metabolism by CYP2F2. Importantly, the human isoform of CYP2F, CYP2F1, is expressed at much lower levels and likely does not catalyze significant styrene metabolism, supporting the hypothesis that styrene-induced mouse lung tumors may not quantitatively, or possibly qualitatively, predict lung tumor potential in humans. Copyright © 2011 Elsevier Inc. All rights reserved.

Oral recombinant human or mouse lactoferrin reduces Mycobacterium tuberculosis TDM induced granulomatous lung pathology.

PubMed

Hwang, Shen-An; Kruzel, Marian L; Actor, Jeffrey K

2017-02-01

Trehalose 6'6-dimycolate (TDM) is the most abundant glycolipid on the cell wall of Mycobacterium tuberculosis (MTB). TDM is capable of inducing granulomatous pathology in mouse models that resembles those induced by MTB infection. Using the acute TDM model, this work investigates the effect of recombinant human and mouse lactoferrin to reduce granulomatous pathology. C57BL/6 mice were injected intravenously with TDM at a dose of 25 μg·mouse -1 . At day 4 and 6, recombinant human or mouse lactoferrin (1 mg·(100 μL) -1 ·mouse -1 ) were delivered by gavage. At day 7 after TDM injection, mice were evaluated for lung pathology, cytokine production, and leukocyte populations. Mice given human or mouse lactoferrin had reduced production of IL-12p40 in their lungs. Mouse lactoferrin increased IL-6 and KC (CXCL1) in lung tissue. Increased numbers of macrophages were observed in TDM-injected mice given human or mouse lactoferrin. Granulomatous pathology, composed of mainly migrated leukocytes, was visually reduced in mice that received human or mouse lactoferrin. Quantitation of granulomatous pathology demonstrated a significant decrease in mice given human or mouse lactoferrin compared with TDM control mice. This report is the first to directly compare the immune modulatory effects of both heterologous recombinant human and homologous mouse lactoferrin on the development of TDM-induced granulomas.

Normal radial migration and lamination are maintained in dyslexia-susceptibility candidate gene homolog Kiaa0319 knockout mice.

PubMed

Martinez-Garay, Isabel; Guidi, Luiz G; Holloway, Zoe G; Bailey, Melissa A G; Lyngholm, Daniel; Schneider, Tomasz; Donnison, Timothy; Butt, Simon J B; Monaco, Anthony P; Molnár, Zoltán; Velayos-Baeza, Antonio

2017-04-01

Developmental dyslexia is a common disorder with a strong genetic component, but the underlying molecular mechanisms are still unknown. Several candidate dyslexia-susceptibility genes, including KIAA0319, DYX1C1, and DCDC2, have been identified in humans. RNA interference experiments targeting these genes in rat embryos have shown impairments in neuronal migration, suggesting that defects in radial cortical migration could be involved in the disease mechanism of dyslexia. Here we present the first characterisation of a Kiaa0319 knockout mouse line. Animals lacking KIAA0319 protein do not show anatomical abnormalities in any of the layered structures of the brain. Neurogenesis and radial migration of cortical projection neurons are not altered, and the intrinsic electrophysiological properties of Kiaa0319-deficient neurons do not differ from those of wild-type neurons. Kiaa0319 overexpression in cortex delays radial migration, but does not affect final neuronal position. However, knockout animals show subtle differences suggesting possible alterations in anxiety-related behaviour and in sensorimotor gating. Our results do not reveal a migration disorder in the mouse model, adding to the body of evidence available for Dcdc2 and Dyx1c1 that, unlike in the rat in utero knockdown models, the dyslexia-susceptibility candidate mouse homolog genes do not play an evident role in neuronal migration. However, KIAA0319 protein expression seems to be restricted to the brain, not only in early developmental stages but also in adult mice, indicative of a role of this protein in brain function. The constitutive and conditional knockout lines reported here will be useful tools for further functional analyses of Kiaa0319.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bhattacharyya, M.H.; Larsen, R.P.; Oldham, R.D.

The fraction of plutonium absorbed after oral administration of Pu(VI) to 24-h-fasted mice was 19 X 10(-4), 13-fold higher than in fed mice, 1.4 X 10(-4). We have investigated the relevance of the high gastrointestinal (GI) absorption value for the 24-h-fasted animals in setting drinking water standards for humans. When fasting was initiated at the beginning of the active phase of the mouse's daily activity cycle (when they would normally eat), plutonium GI absorption rose from 2.8 X 10(-4) at zero-time to a level typical of the 24-h-fasted mouse after only 2 h of fasting. In contrast, in mice allowedmore » to eat for 4 h into their active phase prior to initiation of the fast (meal-fed mice), 8 h of fasting were required before GI absorption rose to a level similar to that of the 24-h-fasted mouse. The fraction of plutonium retained after gavage administration of Pu(VI) to 1-day-old rats was 74 X 10(-4), 70-fold higher than the value for fed adults. Retention after GI absorption in neonates remained 30- to 70-fold higher than in adults until weaning. One week after weaning, the fraction absorbed and retained by fed weanling rats was the same as that for fed adults, 1 X 10(-4). Drinking water standards for plutonium have been set based on GI absorption values for fed adult animals. The 10- to 100-fold increases in plutonium absorption in young and fasted animals reported by ourselves and others, and the rapid rise to fasted levels of absorption at the start of the animal's active phase, indicate that consideration should be given to elevated levels of plutonium absorption in young and fasted individuals.« less

Human cationic amino acid transporter hCAT-3 is preferentially expressed in peripheral tissues.

PubMed

Vékony, N; Wolf, S; Boissel, J P; Gnauert, K; Closs, E I

2001-10-16

At least five distinct carrier proteins form the family of mammalian cationic amino acid transporters (CATs). We have cloned a cDNA containing the complete coding region of human CAT-3. hCAT-3 is glycosylated and localized to the plasma membrane. Transport studies in Xenopus laevis oocytes revealed that hCAT-3 is selective for cationic L-amino acids and exhibits a maximal transport activity similar to other CAT proteins. The apparent substrate affinity and sensitivity to trans-stimulation of hCAT-3 resembles most closely hCAT-2B. This is in contrast to rat and murine CAT-3 proteins that have been reported to display a very low activity and to be inhibited by neutral and anionic L-amino acids as well as D-arginine (Hosokawa, H., et al. (1997) J. Biol. Chem. 272, 8717-8722; Ito, K., and Groudine, M. (1997) J. Biol. Chem. 272, 26780-26786). Also, in adult rat and mouse, CAT-3 has been found exclusively in central neurons. Human CAT-3 expression is not restricted to the brain, in fact, by far the highest expression was found in thymus. Also in other peripheral tissues, hCAT-3 expression was equal to or higher than in most brain regions, suggesting that hCAT-3 is not a neuron-specific transporter.

In Vitro Plasma Stability, Permeability and Solubility of Mercaptoacetamide Histone Deacetylase Inhibitors

PubMed Central

Konsoula, Roula; Jung, Mira

2008-01-01

Histone deacetylase inhibitors (HDACIs) are emerging as a new class of therapeutic agents with potent antitumor activities in a broad spectrum of human cancers. In this study, the in vitro plasma stability, permeability, solubility, and lipophilicity (logD) of two mercaptoacetamide-based HDACIs (coded as W2 and S2) were evaluated and compared to Vorinostat (SAHA). The results demonstrated that the compounds manifested high solubility in HCl (pH 1.2) but lower in PBS (pH 7.4) than SAHA. Moreover, mercaptoacetamide-based HDACIs exhibited higher lipophilicity values compared to SAHA. The permeability of these compounds was evaluated using the Caco-2 cell monolayer as a model of the intestinal mucosa. The Caco-2 studies revealed that the compounds S2 and W2 are highly permeable with apparent permeability coefficients (Papp) in the apical to basolateral direction of 7.33 × 10−6 and 15.0 × 10−6 cm/s, respectively. The in vitro stability was determined in human, mouse, porcine and rat plasma. Data showed that the compound W2 is more stable in human and rat plasma and the S2 is more stable in all plasma species than SAHA. Taken together, these results indicate that the mercaptoacetamide-based HDACIs possess favorable solubility, lipophilicity, permeability and plasma stability features. PMID:18562136

The Complexity of Alcohol Drinking: Studies in Rodent Genetic Models

PubMed Central

Phillips, Tamara J.; Belknap, John K.

2012-01-01

Risk for alcohol dependence in humans has substantial genetic contributions. Successful rodent models generally attempt to address only selected features of the human diagnosis. Most such models target the phenotype of oral administration of alcohol solutions, usually consumption of or preference for an alcohol solution versus water. Data from rats and mice for more than 50 years have shown genetic influences on preference drinking and related phenotypes. This paper summarizes some key findings from that extensive literature. Much has been learned, including the genomic location and possible identity of several genes influencing preference drinking. We report new information from congenic lines confirming QTLs for drinking on mouse chromosomes 2 and 9. There are many strengths of the various phenotypic assays used to study drinking, but there are also some weaknesses. One major weakness, the lack of drinking excessively enough to become intoxicated, has recently been addressed with a new genetic animal model, mouse lines selectively bred for their high and intoxicating blood alcohol levels after a limited period of drinking in the circadian dark. We report here results from a second replicate of that selection and compare them with the first replicate. PMID:20552264

Oxytocin and cardioprotection in diabetes and obesity.

PubMed

Jankowski, Marek; Broderick, Tom L; Gutkowska, Jolanta

2016-06-07

Oxytocin (OT) emerges as a drug for the treatment of diabetes and obesity. The entire OT system is synthesized in the rat and human heart. The direct myocardial infusion with OT into an ischemic or failing heart has the potential to elicit a variety of cardioprotective effects. OT treatment attenuates cardiomyocyte (CMs) death induced by ischemia-reperfusion by activating pro-survival pathways within injured CMs in vivo and in isolated cells. OT treatment reduces cardiac apoptosis, fibrosis, and hypertrophy. The OT/OT receptor (OTR) system is downregulated in the db/db mouse model of type 2 diabetes which develops genetic diabetic cardiomyopathy (DC) similar to human disease. We have shown that chronic OT treatment prevents the development of DC in the db/db mouse. In addition, OT stimulates glucose uptake in both cardiac stem cells and CMs, and increases cell resistance to diabetic conditions. OT may help replace lost CMs by stimulating the in situ differentiation of cardiac stem cells into functional mature CMs. Lastly, adult stem cells amenable for transplantation such as MSCs could be preconditioned with OT ex vivo and implanted into the injured heart to aid in tissue regeneration through direct differentiation, secretion of protective and cardiomyogenic factors and/or their fusion with injured CMs.