Intraperitoneal alpha-radioimmunotherapy in mice using different specific activities

DEFF Research Database (Denmark)

Elgqvist, Jörgen; Andersson, Håkan; Haglund, Elin

2009-01-01

The aim of this study was to investigate the therapeutic efficacy of the alpha-radioimmunotherapy of ovarian cancer in mice, using different specific activities. This study was performed by using the monoclonal antibody, MX35 F(ab')(2), labeled with the alpha-particle-emitter, 211At.......The aim of this study was to investigate the therapeutic efficacy of the alpha-radioimmunotherapy of ovarian cancer in mice, using different specific activities. This study was performed by using the monoclonal antibody, MX35 F(ab')(2), labeled with the alpha-particle-emitter, 211At....

The novel δ opioid receptor agonist KNT-127 produces antidepressant-like and antinociceptive effects in mice without producing convulsions.

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Saitoh, Akiyoshi; Sugiyama, Azusa; Nemoto, Toru; Fujii, Hideaki; Wada, Keiji; Oka, Jun-Ichiro; Nagase, Hiroshi; Yamada, Mitsuhiko

2011-10-01

We previously reported that the δ opioid receptor (DOP) agonists SNC80 and TAN-67 produce potent antidepressant-like and antinociceptive effects in rodents. However, SNC80 produced convulsive effects. Recently, we succeeded in synthesizing a novel DOP agonist called KNT-127. The present study examined the convulsive, antidepressant-like, and antinociceptive effects of KNT-127 in mice. In contrast to SNC80, KNT-127 produced no convulsions at doses of up to 100mg/kg. In mice subjected to the forced swim test, a screening model for antidepressants, KNT-127 (1mg/kg, s.c.) significantly decreased the duration of immobility and increased the duration of swimming without influencing spontaneous locomotor activity. These behavioral changes were similar to that observed for the tricyclic antidepressant imipramine (6mg/kg). The antidepressant-like effect of KNT-127 in mice was antagonized by pretreatment with naltrindole (NTI), a selective DOP antagonist, or naltriben, a putative DOP(2) subtype antagonist. In addition, KNT-127 (3mg/kg, s.c.) significantly reduced the number of acetic acid-induced abdominal constrictions and the duration of licking time, respectively, in mice subjected to a writhing test and a formalin test. These antinociceptive effects were antagonized by pretreatment with either NTI or 7-benzylidenenaltrexone, a putative DOP(1) subtype antagonist. We propose that KNT-127 should be considered as a candidate compound for the development of DOP-based antidepressants and/or analgesics that lack convulsive effects. Crown Copyright © 2011. Published by Elsevier B.V. All rights reserved.

Phase-Specific Vocalizations of Male Mice at the Initial Encounter during the Courtship Sequence.

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Yui K Matsumoto

Full Text Available Mice produce ultrasonic vocalizations featuring a variety of syllables. Vocalizations are observed during social interactions. In particular, males produce numerous syllables during courtship. Previous studies have shown that vocalizations change according to sexual behavior, suggesting that males vary their vocalizations depending on the phase of the courtship sequence. To examine this process, we recorded large sets of mouse vocalizations during male-female interactions and acoustically categorized these sounds into 12 vocal types. We found that males emitted predominantly short syllables during the first minute of interaction, more long syllables in the later phases, and mainly harmonic sounds during mounting. These context- and time-dependent changes in vocalization indicate that vocal communication during courtship in mice consists of at least three stages and imply that each vocalization type has a specific role in a phase of the courtship sequence. Our findings suggest that recording for a sufficiently long time and taking the phase of courtship into consideration could provide more insights into the role of vocalization in mouse courtship behavior in future study.

FAKTOR MAKRO EKONOMI (VARIABEL CRR PADA RETURN PORTOFOLIO PASAR SAHAM DI INDONESIA SAAT BULLISH DAN BEARISH

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Elizabeth Lucky Maretha Sitinjak

2013-03-01

Full Text Available Stock market conditions in Indonesia since 1998 until 2008 is increasing with the 500 IDX Composite can be inflated into 2000s. This becomes interesting factor associated with macro-economic factors that affect the variabel CRR (Chen Roll and Ross. Researchers exam which we form portofiolio into 3 classes, large, medium, and large size companies from multiplying shares outstanding by its stock price. This research periodesasi long enough so that the portfolio is formed only by 15 issuers, this is caused by mergers or acquisitions from the issuer, the issuer is listed on the Stock Exchange from 1998-2008. However, with 11 years of data is very good for macro economic conditions in Indonesia. Independen variabels of this study consisted of changes in inflation expectations (DEIt, unexpected inflation (Uit, unexpected risk free rate (URFt, and the rate of economic growth (GMT have a significant effect on portfolio return of capital market conditions are bullish for all forms of portfolio. Meanwhile, bearish market conditions only for the portfolio of small and large sizes only. Economic growth rate did not significantly affect the three portfolios in the bearish market conditions, this is because the movement of our stock in Indonesia is still largely influenced by foreign investors.

The induction of specific immunity against Schistosoma japonicum by exposure of mice to ultraviolet attenuated cercariae

International Nuclear Information System (INIS)

Moloney, N.A.; Bickle, Q.D.; Webbe, G.

1985-01-01

Mice can be partially protected against Schistosoma japonicum by prior exposure to ultraviolet (UV)-attenuated infections which fail to survive to the adult stage and produce no overt pathology in the host. Optimum resistance was induced by parasites exposed to 40 seconds of UV, significantly lower levels of resistance being stimulated by both shorter and longer exposures. No consistent relationship between the degree of resistance induced and the number of irradiated cercariae given could be demonstrated and equivocal results were obtained when comparing the efficacy of single and multiple vaccinations. Vaccinations with UV-attenuated cercariae given intraperitoneally (i.p.) were as efficacious as those given percutaneously but mice were as or more resistant to challenges given by the i.p. route, the possible reasons are discussed. There was no observed delay in the migration of the challenge, vaccinated mice being as resistant when perfused 6 or 3.5 weeks after challenge. Vaccination was species specific since mice exposed to either UV-attenuated S. japonicum cercariae or gamma-attenuated S. mansoni cercariae were resistant to homologous but not heterologous challenge. (author)

Sex-specific effects of docosahexaenoic acid (DHA) on the microbiome and behavior of socially-isolated mice.

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Davis, Daniel J; Hecht, Patrick M; Jasarevic, Eldin; Beversdorf, David Q; Will, Matthew J; Fritsche, Kevin; Gillespie, Catherine H

2017-01-01

Dietary supplementation with the long-chain omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) has been shown to have a beneficial effect on reducing the symptoms associated with several neuropsychiatric conditions including anxiety and depression. However, the mechanisms underlying this effect remain largely unknown. Increasing evidence suggests that the vast repertoire of commensal bacteria within the gut plays a critical role in regulating various biological processes in the brain and may contribute to neuropsychiatric disease risk. The present study determined the contribution of DHA on anxiety and depressive-like behaviors through modulation of the gut microbiota in a paradigm of social isolation. Adult male and female mice were subjected to social isolation for 28days and then placed either on a control diet or a diet supplemented with 0.1% or 1.0% DHA. Fecal pellets were collected both 24h and 7days following the introduction of the new diets. Behavioral testing revealed that male mice fed a DHA diet, regardless of dose, exhibited reduced anxiety and depressive-like behaviors compared to control fed mice while no differences were observed in female mice. As the microbiota-brain-axis has been recently implicated in behavior, composition of microbial communities were analyzed to examine if these sex-specific effects of DHA may be associated with changes in the gut microbiota (GM). Clear sex differences were observed with males and females showing distinct microbial compositions prior to DHA supplementation. The introduction of DHA into the diet also induced sex-specific interactions on the GM with the fatty acid producing a significant effect on the microbial profiles in males but not in females. Interestingly, levels of Allobaculum and Ruminococcus were found to significantly correlate with the behavioral changes observed in the male mice. Predictive metagenome analysis using PICRUSt was performed on the fecal samples collected from males and

Glutamate receptor antibodies directed against AMPA receptors subunit 3 peptide B (GluR3B) can be produced in DBA/2J mice, lower seizure threshold and induce abnormal behavior.

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Ganor, Yonatan; Goldberg-Stern, Hadassa; Cohen, Ran; Teichberg, Vivian; Levite, Mia

2014-04-01

Anti-GluR3B antibodies (GluR3B Ab's), directed against peptide B/aa372-395 of GluR3 subunit of glutamate/AMPA receptors, are found in ∼35% of epilepsy patients, activate glutamate/AMPA receptors, evoke ion currents, kill neurons and damage the brain. We recently found that GluR3B Ab's also associate with neurological/psychiatric/behavioral abnormalities in epilepsy patients. Here we asked if GluR3B Ab's could be produced in DBA/2J mice, and also modulate seizure threshold and/or cause behavioral/motor impairments in these mice. DBA/2J mice were immunized with the GluR3B peptide in Complete Freund's Adjuvant (CFA), or with controls: ovalbumin (OVA), CFA, or phosphate-buffer saline (PBS). GluR3B Ab's and OVA Ab's were tested. Seizures were induced in all mice by the chemoconvulsant pentylenetetrazole (PTZ) at three time points, each time with less PTZ to avoid non-specific death. Behavior was examined in Open-Field, RotaRod and Grip tests. GluR3B Ab's were produced only in GluR3B-immunized mice, while OVA Ab's were produced only in OVA-immunized mice, showing high Ab's specificity. In GluR3B Ab's negative mice, seizure severity scores and percentages of animals developing generalized seizures declined in response to decreasing PTZ doses. In contrast, both parameters remained unchanged/high in the GluR3B Ab's positive mice, showing that these mice were more susceptible to seizures. The seizure scores associated significantly with the GluR3B Ab's levels. GluR3B Ab's positive mice were also more anxious in Open-Field test, fell faster in RotaRod test, and fell more in Grip test, compared to all the control mice. GluR3B Ab's are produced in DBA/2J mice, facilitate seizures and induce behavioral/motor impairments. This animal model can therefore serve for studying autoimmune epilepsy and abnormal behavior mediated by pathogenic anti-GluR3B Ab's. Copyright © 2014 Elsevier Ltd. All rights reserved.

Ketamine Does Not Produce Relief of Neuropathic Pain in Mice Lacking the β-Common Receptor (CD131)

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Swartjes, Maarten; Niesters, Marieke; Heij, Lara; Dunne, Ann; Aarts, Leon; Hand, Carla Cerami; Kim, Hyung-Suk; Brines, Michael; Cerami, Anthony; Dahan, Albert

2013-01-01

Neuropathic pain (NP) is a debilitating condition associated with traumatic, metabolic, autoimmune and neurological etiologies. Although the triggers for NP are diverse, there are common underlying pathways, including activation of immune cells in the spinal cord and up-regulation of the N-methyl-D-aspartate receptor (NMDAR). Ketamine, a well-known NDMAR antagonist, reduces neuropathic pain in a sustained manner. Recent study has shown that the novel 11-amino acid peptide erythropoietin derivative ARA290 produces a similar, long-lasting relief of NP. Here, we show that both drugs also have similar effects on the expression of mRNA of the NMDAR, as well as that of microglia, astrocytes and chemokine (C-C motif) ligand 2, all-important contributors to the development of NP. Although the effects of ketamine and ARA 290 on NP and its molecular mediators suggest a common mechanism of action, ARA 290 has no affinity for the NMDAR and acts specifically via the innate repair receptor (IRR) involved in tissue protection. We speculated therefore, that the IRR might be critically involved in the action of ketamine on neuropathic pain. To evaluate this, we studied the effects of ketamine and ARA 290 on acute pain, side effects, and allodynia following a spared nerve injury model in mice lacking the β-common receptor (βcR), a structural component of the IRR. Ketamine (50 mg/kg) and ARA 290 (30 µg/kg) produced divergent effects on acute pain: ketamine produced profound antinociception accompanied with psychomotor side effects, but ARA290 did not, in both normal and knock out mice. In contrast, while both drugs were antiallodynic in WT mice, they had no effect on NP in mice lacking the βcR. Together, these results show that an intact IRR is required for the effective treatment of NP with either ketamine or ARA 290, but is not involved in ketamine’s analgesic and side effects. PMID:23936499

Intestine-specific deletion of microsomal triglyceride transfer protein increases mortality in aged mice.

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Liang, Zhe; Xie, Yan; Dominguez, Jessica A; Breed, Elise R; Yoseph, Benyam P; Burd, Eileen M; Farris, Alton B; Davidson, Nicholas O; Coopersmith, Craig M

2014-01-01

Mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO) exhibit a complete block in chylomicron assembly together with lipid malabsorption. Young (8-10 week) Mttp-IKO mice have improved survival when subjected to a murine model of Pseudomonas aeruginosa-induced sepsis. However, 80% of deaths in sepsis occur in patients over age 65. The purpose of this study was to determine whether age impacts outcome in Mttp-IKO mice subjected to sepsis. Aged (20-24 months) Mttp-IKO mice and WT mice underwent intratracheal injection with P. aeruginosa. Mice were either sacrificed 24 hours post-operatively for mechanistic studies or followed seven days for survival. In contrast to young septic Mttp-IKO mice, aged septic Mttp-IKO mice had a significantly higher mortality than aged septic WT mice (80% vs. 39%, p = 0.005). Aged septic Mttp-IKO mice exhibited increased gut epithelial apoptosis, increased jejunal Bax/Bcl-2 and Bax/Bcl-XL ratios yet simultaneously demonstrated increased crypt proliferation and villus length. Aged septic Mttp-IKO mice also manifested increased pulmonary myeloperoxidase levels, suggesting increased neutrophil infiltration, as well as decreased systemic TNFα compared to aged septic WT mice. Blocking intestinal chylomicron secretion alters mortality following sepsis in an age-dependent manner. Increases in gut apoptosis and pulmonary neutrophil infiltration, and decreased systemic TNFα represent potential mechanisms for why intestine-specific Mttp deletion is beneficial in young septic mice but harmful in aged mice as each of these parameters are altered differently in young and aged septic WT and Mttp-IKO mice.

Intestine-specific deletion of microsomal triglyceride transfer protein increases mortality in aged mice.

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Zhe Liang

Full Text Available Mice with conditional, intestine-specific deletion of microsomal triglyceride transfer protein (Mttp-IKO exhibit a complete block in chylomicron assembly together with lipid malabsorption. Young (8-10 week Mttp-IKO mice have improved survival when subjected to a murine model of Pseudomonas aeruginosa-induced sepsis. However, 80% of deaths in sepsis occur in patients over age 65. The purpose of this study was to determine whether age impacts outcome in Mttp-IKO mice subjected to sepsis.Aged (20-24 months Mttp-IKO mice and WT mice underwent intratracheal injection with P. aeruginosa. Mice were either sacrificed 24 hours post-operatively for mechanistic studies or followed seven days for survival.In contrast to young septic Mttp-IKO mice, aged septic Mttp-IKO mice had a significantly higher mortality than aged septic WT mice (80% vs. 39%, p = 0.005. Aged septic Mttp-IKO mice exhibited increased gut epithelial apoptosis, increased jejunal Bax/Bcl-2 and Bax/Bcl-XL ratios yet simultaneously demonstrated increased crypt proliferation and villus length. Aged septic Mttp-IKO mice also manifested increased pulmonary myeloperoxidase levels, suggesting increased neutrophil infiltration, as well as decreased systemic TNFα compared to aged septic WT mice.Blocking intestinal chylomicron secretion alters mortality following sepsis in an age-dependent manner. Increases in gut apoptosis and pulmonary neutrophil infiltration, and decreased systemic TNFα represent potential mechanisms for why intestine-specific Mttp deletion is beneficial in young septic mice but harmful in aged mice as each of these parameters are altered differently in young and aged septic WT and Mttp-IKO mice.

Screening strategy to generate cell specific recombination: a case report with the RIP-Cre mice.

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Spinelli, Valeria; Martin, Céline; Dorchies, Emilie; Vallez, Emmanuelle; Dehondt, Hélène; Trabelsi, Mohamed-Sami; Tailleux, Anne; Caron, Sandrine; Staels, Bart

2015-10-01

Conditional gene knockout technology is a powerful tool to study the function of a gene in a specific tissue, organ or cell lineage. The most commonly used procedure applies the Cre-LoxP strategy, where the choice of the Cre driver promoter is critical to determine the efficiency and specificity of the system. However, a considered choice of an appropriate promoter does not always protect against the risk of unwanted recombination and the consequent deletion of the gene in other tissues than the desired one(s), due to phenomena of non-specific activation of the Cre transgene. Furthermore, the causes of these phenomena are not completely understood and this can potentially affect every strain of Cre-mice. In our study on the deletion of a same gene in two different tissues, we show that the incidence rate of non-specific recombination in unwanted tissues depends on the Cre driver strain, ranging from 100%, rendering it useless (aP2-Cre strain), to ~5%, which is still compatible with their use (RIP-Cre strain). The use of a simple PCR strategy conceived to detect this occurrence is indispensable when producing a tissue-specific knockout mouse. Therefore, when choosing the Cre-driver promoter, researchers not only have to be careful about its tissue-specificity and timing of activation, but should also include a systematical screening in order to exclude mice in which atypical recombination has occurred and to limit the unnecessary use of laboratory animals in uninterpretable experiments.

Behavioral Characteristics of Ubiquitin-Specific Peptidase 46-Deficient Mice

Science.gov (United States)

Imai, Saki; Kano, Makoto; Nonoyama, Keiko; Ebihara, Shizufumi

2013-01-01

We have previously identified Usp46, which encodes for ubiquitin-specific peptidase 46, as a quantitative trait gene affecting the immobility time of mice in the tail suspension test (TST) and forced swimming test. The mutation that we identified was a 3-bp deletion coding for lysine (Lys 92), and mice with this mutation (MT mice), as well as Usp46 KO mice exhibited shorter TST immobility times. Behavioral pharmacology suggests that the gamma aminobutyric acid A (GABAA) receptor is involved in regulating TST immobility time. In order to understand how far Usp46 controls behavioral phenotypes, which could be related to mental disorders in humans, we subjected Usp46 MT and KO mice to multiple behavioral tests, including the open field test, ethanol preference test, ethanol-induced loss of righting reflex test, sucrose preference test, novelty-suppressed feeding test, marble burying test, and novel object recognition test. Although behavioral phenotypes of the Usp46 MT and KO mice were not always identical, deficiency of Usp46 significantly affected performance in all these tests. In the open field test, activity levels were lower in Usp46 KO mice than wild type (WT) or MT mice. Both MT and KO mice showed lower ethanol preference and shorter recovery times after ethanol administration. Compared to WT mice, Usp46 MT and KO mice exhibited decreased sucrose preference, took longer latency periods to bite pellets, and buried more marbles in the sucrose preference test, novelty-suppressed feeding test, and marble burying test, respectively. In the novel object recognition test, neither MT nor KO mice showed an increase in exploration of a new object 24 hours after training. These findings indicate that Usp46 regulates a wide range of behavioral phenotypes that might be related to human mental disorders and provides insight into the function of USP46 deubiquitinating enzyme in the neural system. PMID:23472206

Behavioral characteristics of ubiquitin-specific peptidase 46-deficient mice.

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Saki Imai

Full Text Available We have previously identified Usp46, which encodes for ubiquitin-specific peptidase 46, as a quantitative trait gene affecting the immobility time of mice in the tail suspension test (TST and forced swimming test. The mutation that we identified was a 3-bp deletion coding for lysine (Lys 92, and mice with this mutation (MT mice, as well as Usp46 KO mice exhibited shorter TST immobility times. Behavioral pharmacology suggests that the gamma aminobutyric acid A (GABAA receptor is involved in regulating TST immobility time. In order to understand how far Usp46 controls behavioral phenotypes, which could be related to mental disorders in humans, we subjected Usp46 MT and KO mice to multiple behavioral tests, including the open field test, ethanol preference test, ethanol-induced loss of righting reflex test, sucrose preference test, novelty-suppressed feeding test, marble burying test, and novel object recognition test. Although behavioral phenotypes of the Usp46 MT and KO mice were not always identical, deficiency of Usp46 significantly affected performance in all these tests. In the open field test, activity levels were lower in Usp46 KO mice than wild type (WT or MT mice. Both MT and KO mice showed lower ethanol preference and shorter recovery times after ethanol administration. Compared to WT mice, Usp46 MT and KO mice exhibited decreased sucrose preference, took longer latency periods to bite pellets, and buried more marbles in the sucrose preference test, novelty-suppressed feeding test, and marble burying test, respectively. In the novel object recognition test, neither MT nor KO mice showed an increase in exploration of a new object 24 hours after training. These findings indicate that Usp46 regulates a wide range of behavioral phenotypes that might be related to human mental disorders and provides insight into the function of USP46 deubiquitinating enzyme in the neural system.

Formation and Expansion of Leukemia-Specific Chromosome Aberrations in Hematopoietic Cells of X-ray Irradiated Mice

International Nuclear Information System (INIS)

Ban, N.; Kai, M.; Kusama, T.

2004-01-01

C3H/He mice develop acute myeloid leukemia (AML) after whole-body irradiation, and typical chromosome 2 deletions are found in the leukemia cells. To investigate a process of the formation and the expansion of the AML-specific deletions, we have examined its frequency in primitive hematopoietic cells that could be the target of the leukemogenesis. Male C3H/He mice were exposed to 3Gy x-rays and sacrificed after certain periods of time. Bone marrow cells were collected from the femora and a single-cell suspension from each animal was divided into two parts. One part of the cell suspension was cultured in methylcellulose medium and metaphase spreads were prepared from each growing colony. The other part was sorted to obtain Lin+ and Lin Scal cells and those cells were scored with FISH for the AML-specific deletions. Karyotyping of the cultured cells detected signs of the delayed chromosomal instability, but an aberration involving chromosome 2 has not been found so far. FISH to the sorted cells, however, revealed the ANL-specific deletions could be produced in the primitive hematopoietic cells as an early event of radiation exposure. (Author) 16 refs

Identification of novel QTLs for isolate-specific partial resistance to Plasmodiophora brassicae in Brassica rapa.

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Jingjing Chen

Full Text Available Plasmodiophora brassicae, the causal agent of clubroot disease of the Brassica crops, is widespread in the world. Quantitative trait loci (QTLs for partial resistance to 4 different isolates of P. brassicae (Pb2, Pb4, Pb7, and Pb10 were investigated using a BC1F1 population from a cross between two subspecies of Brassica rapa, i.e. Chinese cabbage inbred line C59-1 as a susceptible recurrent parent and turnip inbred line ECD04 as a resistant donor parent. The BC1F2 families were assessed for resistance under controlled conditions. A linkage map constructed with simple sequence repeats (SSR, unigene-derived microsatellite (UGMS markers, and specific markers linked to published clubroot resistance (CR genes of B. rapa was used to perform QTL mapping. A total of 6 QTLs residing in 5 CR QTL regions of the B. rapa chromosomes A01, A03, and A08 were identified to account for 12.2 to 35.2% of the phenotypic variance. Two QTL regions were found to be novel except for 3 QTLs in the respective regions of previously identified Crr1, Crr2, and Crr3. QTL mapping results indicated that 1 QTL region was common for partial resistance to the 2 isolates of Pb2 and Pb7, whereas the others were specific for each isolate. Additionally, synteny analysis between B. rapa and Arabidopsis thaliana revealed that all CR QTL regions were aligned to a single conserved crucifer blocks (U, F, and R on 3 Arabidopsis chromosomes where 2 CR QTLs were detected in A. thaliana. These results suggest that some common ancestral genomic regions were involved in the evolution of CR genes in B. rapa.

Similar efficacy from specific and non-specific mineralocorticoid receptor antagonist treatment of muscular dystrophy mice.

Science.gov (United States)

Lowe, Jeovanna; Floyd, Kyle T; Rastogi, Neha; Schultz, Eric J; Chadwick, Jessica A; Swager, Sarah A; Zins, Jonathan G; Kadakia, Feni K; Smart, Suzanne; Gomez-Sanchez, Elise P; Gomez-Sanchez, Celso E; Raman, Subha V; Janssen, Paul M L; Rafael-Fortney, Jill A

2016-01-01

Combined treatment with an angiotensin-converting enzyme inhibitor and a mineralocorticoid receptor (MR) antagonist improved cardiac and skeletal muscle function and pathology in a mouse model of Duchenne muscular dystrophy. MR is present in limb and respiratory skeletal muscles and functions as a steroid hormone receptor. The goals of the current study were to compare the efficacy of the specific MR antagonist eplerenone with the non-specific MR antagonist spironolactone, both in combination with the angiotensin-converting enzyme inhibitor lisinopril. Three groups of n=18 dystrophin-deficient, utrophin-haploinsufficient male mice were given chow containing: lisinopril plus spironolactone, lisinopril plus eplerenone, or no drug, from four to 20 weeks-of-age. Eighteen C57BL/10 male mice were used as wild-type controls. In vivo measurements included cardiac magnetic resonance imaging, conscious electrocardiography, and grip strength. From each mouse in the study, diaphragm, extensor digitorum longus , and cardiac papillary muscle force was measured ex vivo , followed by histological quantification of muscle damage in heart, diaphragm, quadriceps, and abdominal muscles. MR protein levels were also verified in treated muscles. Treatment with specific and non-specific MR antagonists did not result in any adverse effects to dystrophic skeletal muscles or heart. Both treatments resulted in similar functional and pathological improvements across a wide array of parameters. MR protein levels were not reduced by treatment. These data suggest that spironolactone and eplerenone show similar effects in dystrophic mice and support the clinical development of MR antagonists for treating skeletal muscles in Duchenne muscular dystrophy.

Strain-specific aggressive behavior of male mice submitted to different husbandry procedures

NARCIS (Netherlands)

Loo, P.L.P. van; Meer, E. van der; Kruitwagen, C.L.J.J.; Koolhaas, J.M.; Zutphen, L.F.M. van; Baumans, V.

Severe aggression within groups of male laboratory mice can cause serious welfare problems. Previous experiments have shown that the transfer of specific olfactory cues during cage cleaning and the provision of nesting material decrease aggression and stress in group-housed male mice. In this study,

Association of Diet With Skin Histological Features in UV-B-Exposed Mice.

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Bhattacharyya, Tapan K; Hsia, Yvonne; Weeks, David M; Dixon, Tatiana K; Lepe, Jessica; Thomas, J Regan

2017-09-01

Long-term exposure to solar radiation produces deleterious photoaging of the skin. It is not known if diet can influence skin photoaging. To study the influence of a calorie-restricted diet and an obesity diet in mice exposed to long-term UV-B irradiation to assess if there is an association between diet and histopathological response to UV-B irradiation. In this animal model study in an academic setting, the dorsal skin of SKH1 hairless mice receiving normal, calorie-restricted, and obesity diets was exposed to UV-B irradiation 3 times a week for 10 weeks and were compared with corresponding controls. The mice were placed in the following groups, with 8 animals in each group: (1) intact control (C) with regular diet and no UV-B exposure, (2) intact control with UV-B exposure (CR), (3) calorie-restricted diet (CrC), (4) calorie-restricted diet with UV-B exposure (CrR), (5) obesity diet (OC), and (6) obesity diet with UV-B exposure (OR). The experiment was conducted during October through December 2013. Tissue processing and histological analysis were completed in 2016. Histomorphometric analysis was performed on paraffin-embedded skin sections stained by histological and immunohistochemical methods for estimation of epidermal thickness, epidermal proliferating cell nuclear antigen index, collagen I, elastic fibers, fibroblasts, mast cells, dermal cellularity, and adipose layer ratio. Changes in wrinkles were noted. Hairless female mice (age range, 6-8 weeks) were obtained. With a normal diet, changes from UV-B irradiation occurred in epidermal thickness, epidermal proliferating cell nuclear antigen index, collagen I, elastic fibers, fibroblasts, and mast cells, which were modestly influenced by an obesity diet. Calorie restriction influenced the skin in nonirradiated control animals, with higher values for most variables. After UV-B exposure in animals with calorie restriction, epidermal thickness was increased, but other variables were unaffected. Animals

Enhanced normal short-term human myelopoiesis in mice engineered to express human-specific myeloid growth factors.

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Miller, Paul H; Cheung, Alice M S; Beer, Philip A; Knapp, David J H F; Dhillon, Kiran; Rabu, Gabrielle; Rostamirad, Shabnam; Humphries, R Keith; Eaves, Connie J

2013-01-31

Better methods to characterize normal human hematopoietic cells with short-term repopulating activity cells (STRCs) are needed to facilitate improving recovery rates in transplanted patients.We now show that 5-fold more human myeloid cells are produced in sublethally irradiated NOD/SCID-IL-2Receptor-γchain-null (NSG) mice engineered to constitutively produce human interleukin-3, granulocyte-macrophage colony-stimulating factor and Steel factor (NSG-3GS mice) than in regular NSG mice 3 weeks after an intravenous injection of CD34 human cord blood cells. Importantly, the NSG-3GS mice also show a concomitant and matched increase in circulating mature human neutrophils. Imaging NSG-3GS recipients of lenti-luciferase-transduced cells showed that human cells being produced 3 weeks posttransplant were heterogeneously distributed, validating the blood as a more representative measure of transplanted STRC activity. Limiting dilution transplants further demonstrated that the early increase in human granulopoiesis in NSG-3GS mice reflects an expanded output of differentiated cells per STRC rather than an increase in STRC detection. NSG-3GS mice support enhanced clonal outputs from human short-term repopulating cells (STRCs) without affecting their engrafting efficiency. Increased human STRC clone sizes enable their more precise and efficient measurement by peripheral blood monitoring.

Formation of the accumulative human metabolite and human-specific glutathione conjugate of diclofenac in TK-NOG chimeric mice with humanized livers.

Science.gov (United States)

Kamimura, Hidetaka; Ito, Satoshi; Nozawa, Kohei; Nakamura, Shota; Chijiwa, Hiroyuki; Nagatsuka, Shin-ichiro; Kuronuma, Miyuki; Ohnishi, Yasuyuki; Suemizu, Hiroshi; Ninomiya, Shin-ichi

2015-03-01

3'-Hydroxy-4'-methoxydiclofenac (VI) is a human-specific metabolite known to accumulate in the plasma of patients after repeated administration of diclofenac sodium. Diclofenac also produces glutathione-conjugated metabolites, some of which are human-specific. In the present study, we investigated whether these metabolites could be generated in humanized chimeric mice produced from TK-NOG mice. After a single oral administration of diclofenac to humanized mice, the unchanged drug in plasma peaked at 0.25 hour and then declined with a half-life (t1/2) of 2.4 hours. 4'-Hydroxydiclofenac (II) and 3'-hydroxydiclofenac also peaked at 0.25 hour and were undetectable within 24 hours. However, VI peaked at 8 hours and declined with a t1/2 of 13 hours. When diclofenac was given once per day, peak and trough levels of VI reached plateau within 3 days. Studies with administration of II suggested VI was generated via II as an intermediate. Among six reported glutathione-conjugated metabolites of diclofenac, M1 (5-hydroxy-4-(glutathion-S-yl)diclofenac) to M6 (2'-(glutathion-S-yl)monoclofenac), we found three dichlorinated conjugates [M1, M2 (4'-hydroxy-3'-(glutathion-S-yl)diclofenac), and M3 (5-hydroxy-6-(glutathion-S-yl)diclofenac)], and a single monochlorinated conjugate [M4 (2'-hydroxy-3'-(glutathion-S-yl)monoclofenac) or M5 (4'-hydroxy-2'-(glutathion-S-yl)monoclofenac)], in the bile of humanized chimeric mice. M4 and M5 are positional isomers and have been previously reported as human-specific in vitro metabolites likely generated via arene oxide and quinone imine-type intermediates, respectively. The biliary monochlorinated metabolite exhibited the same mass spectrum as those of M4 and M5, and we discuss whether this conjugate corresponded to M4 or M5. Overall, humanized TK-NOG chimeric mice were considered to be a functional tool for the study of drug metabolism of diclofenac in humans. Copyright © 2015 by The American Society for Pharmacology and Experimental

Exopolysaccharide-producing probiotic Lactobacilli reduce serum cholesterol and modify enteric microbiota in ApoE-deficient mice.

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London, Lis E E; Kumar, Arun H S; Wall, Rebecca; Casey, Pat G; O'Sullivan, Orla; Shanahan, Fergus; Hill, Colin; Cotter, Paul D; Fitzgerald, Gerald F; Ross, R Paul; Caplice, Noel M; Stanton, Catherine

2014-12-01

Probiotic bacteria have been associated with a reduction in cardiovascular disease risk, a leading cause of death and disability. The aim of this study was to assess the impact of dietary administration of exopolysaccharide-producing probiotic Lactobacillus cultures on lipid metabolism and gut microbiota in apolipoprotein E (apoE)-deficient mice. First, we examined lipid metabolism in response to dietary supplementation with recombinant β-glucan-producing Lactobacillus paracasei National Food Biotechnology Centre (NFBC) 338 expressing the glycosyltransferase (Gtf) gene from Pediococcus parvulus 2.6 (GTF), and naturally exopolysaccharide-producing Lactobacillus mucosae Dairy Product Culture Collection (DPC) 6426 (DPC 6426) compared with the non-β-glucan-producing isogenic control strain Lactobacillus paracasei NFBC 338 (PNZ) and placebo (15% wt:vol trehalose). Second, we examined the effects on the gut microbiota of dietary administration of DPC 6426 compared with placebo. Probiotic Lactobacillus strains at 1 × 10(9) colony-forming units/d per animal were administered to apoE(-/-) mice fed a high-fat (60% fat)/high-cholesterol (2% wt:wt) diet for 12 wk. At the end of the study, aortic plaque development and serum, liver, and fecal variables involved in lipid metabolism were analyzed, and culture-independent microbial analyses of cecal content were performed. Total cholesterol was reduced in serum (P mice supplemented with GTF or DPC 6426 compared with the PNZ or placebo group, respectively. In addition, dietary intervention with GTF led to increased amounts of fecal cholesterol excretion (P mice. © 2014 American Society for Nutrition.

Strain-specific patterns of autonomic nervous system activity and heart failure susceptibility in mice.

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Shusterman, Vladimir; Usiene, Irmute; Harrigal, Chivonne; Lee, Joon Sup; Kubota, Toru; Feldman, Arthur M; London, Barry

2002-06-01

Transgenic mice are widely used to study cardiac function, but strain-dependent differences in autonomic nervous system activity (ANSA) have not been explored. We compared 1) short-term pharmacological responses of cardiac rhythm in FVB vs. C57Black6/SV129 wild-type mice and 2) long-term physiological dynamics of cardiac rhythm and survival in tumor necrosis factor (TNF)-alpha transgenic mice with heart failure (TNF-alpha mice) on defined backgrounds. Ambulatory telemetry electrocardiographic recordings and response to saline, adrenergic, and cholinergic agents were examined in FVB and C57Black6/SV129 mice. In FVB mice, baseline heart rate (HR) was higher and did not change after injection of isoproterenol or atropine but decreased with propranolol. In C57Black6/SV129 mice, HR did not change with propranolol but increased with isoproterenol or atropine. Mean HR, but not indexes of HR variability, was an excellent predictor of response to autonomic agents. The proportion of surviving animals was higher in TNF-alpha mice on an FVB background than on a mixed FVB/C57Black6 background. The homeostatic states of ANSA are strain specific, which can explain the interstrain differences in mean HR, pharmacological responses, and survival of animals with congestive heart failure. Strain-specific differences should be considered in selecting the strains of mice used for transgenic and gene targeting experiments.

Autoreactive T cells in MRL/Mpr-lpr/lpr mice. Characterization of the lymphokines produced and analysis of antigen-presenting cells required

International Nuclear Information System (INIS)

Weston, K.M.; Ju, S.T.; Lu, C.Y.; Sy, M.S.

1988-01-01

Lymph node cells from 4-wk-old MRL/Mp-lpr/lpr mice, but not from MRL/Mp-+/+ mice, when cultured in vitro for 5 to 7 days, will spontaneously proliferate and produce IL-2. We examined the expression of several cell surface Ag on lymph node cells from MRL/Mp-lpr/lpr mice before and after in vitro culture. There is an increase in the expression of Thy-1, L3T4, IL-2R, T cell activating protein, T cell receptor, and T3 complex on the surface of cultured cells. Cultured cells produced IL-3, IFN-gamma, and small but detectable amounts of IL-1 in addition to IL-2. Gamma irradiation of APC from young MRL/Mp-lpr/lpr mice or treatment of APC with a mAb (J11D) and C, completely abrogated their stimulatory capacity. These experiments suggest that B cells are the predominant APC responsible in the activation of autoreactive T cells in MRL/Mp-lpr/lpr mice. Lymph node cells from C57BL/6-lpr/lpr or C3H-lpr/lpr mice were unable to spontaneously proliferate or produce IL-2. Lymph node cells from (MRL/Mp-lpr/lpr x C57BL/6-lpr/lpr) F1 mice or (C3H-lpr/lpr x MRL/Mp-lpr/lpr) F1 mice did proliferate and produced IL-2 after in vitro culture. Using T cells from these F1 animals and APC from each parental haplotype, we found that APC from MRL/Mp-lpr/lpr mice induced more proliferation and greater amounts of IL-2, when compared to APC from F1 animals. APC from C57BL6-lpr/lpr mice or C3H-lpr/lpr were unable to induce spontaneous proliferation and IL-2 production. Therefore, B cells from MRL/Mp-lpr/lpr mice appear to possess unique features that enable them to activate autoreactive T cells more effectively than B cells from other mice bearing the lpr/lpr gene

Completely ES cell-derived mice produced by tetraploid complementation using inner cell mass (ICM deficient blastocysts.

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Duancheng Wen

Full Text Available Tetraploid complementation is often used to produce mice from embryonic stem cells (ESCs by injection of diploid (2n ESCs into tetraploid (4n blastocysts (ESC-derived mice. This method has also been adapted to mouse cloning and the derivation of mice from induced pluripotent stem (iPS cells. However, the underlying mechanism(s of the tetraploid complementation remains largely unclear. Whether this approach can give rise to completely ES cell-derived mice is an open question, and has not yet been unambiguously proven. Here, we show that mouse tetraploid blastocysts can be classified into two groups, according to the presence or absence of an inner cell mass (ICM. We designate these as type a (presence of ICM at blastocyst stage or type b (absence of ICM. ESC lines were readily derived from type a blastocysts, suggesting that these embryos retain a pluripotent epiblast compartment; whereas the type b blastocysts possessed very low potential to give rise to ESC lines, suggesting that they had lost the pluripotent epiblast. When the type a blastocysts were used for tetraploid complementation, some of the resulting mice were found to be 2n/4n chimeric; whereas when type b blastocysts were used as hosts, the resulting mice are all completely ES cell-derived, with the newborn pups displaying a high frequency of abdominal hernias. Our results demonstrate that completely ES cell-derived mice can be produced using ICM-deficient 4n blastocysts, and provide evidence that the exclusion of tetraploid cells from the fetus in 2n/4n chimeras can largely be attributed to the formation of ICM-deficient blastocysts.

Mild Social Stress in Mice Produces Opioid-Mediated Analgesia in Visceral but Not Somatic Pain States.

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Pitcher, Mark H; Gonzalez-Cano, Rafael; Vincent, Kathleen; Lehmann, Michael; Cobos, Enrique J; Coderre, Terence J; Baeyens, José M; Cervero, Fernando

2017-06-01

Visceral pain has a greater emotional component than somatic pain. To determine if the stress-induced analgesic response is differentially expressed in visceral versus somatic pain states, we studied the effects of a mild social stressor in either acute visceral or somatic pain states in mice. We show that the presence of an unfamiliar conspecific mouse (stranger) in an adjacent cubicle of a standard transparent observation box produced elevated plasma corticosterone levels compared with mice tested alone, suggesting that the mere presence of a stranger is stressful. We then observed noxious visceral or somatic stimulation-induced nociceptive behavior in mice tested alone or in mildly stressful conditions (ie, beside an unfamiliar stranger). Compared with mice tested alone, the presence of a stranger produced a dramatic opioid-dependent reduction in pain behavior associated with visceral but not somatic pain. This social stress-induced reduction of visceral pain behavior relied on visual but not auditory/olfactory cues. These findings suggest that visceral pain states may provoke heightened responsiveness to mild stressors, an effect that could interfere with testing outcomes during simultaneous behavioral testing of multiple rodents. In mice, mild social stress due to the presence of an unfamiliar conspecific mouse reduces pain behavior associated with noxious visceral but not somatic stimulation, suggesting that stress responsiveness may be enhanced in visceral pain versus somatic pain states. Published by Elsevier Inc.

Isoflurane produces sustained cardiac protection after ischemia-reperfusion injury in mice.

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Tsutsumi, Yasuo M; Patel, Hemal H; Lai, N Chin; Takahashi, Toshiyuki; Head, Brian P; Roth, David M

2006-03-01

Isoflurane reduces myocardial ischemia-reperfusion injury within hours to days of reperfusion. Whether isoflurane produces sustained cardiac protection has never been examined. The authors studied isoflurane-induced cardiac protection in the intact mouse after 2 h and 2 weeks of reperfusion and determined the dependence of this protection on adenosine triphosphate-dependent potassium channels and the relevance of this protection to myocardial function and apoptosis. Mice were randomly assigned to receive oxygen or isoflurane for 30 min with 15 min of washout. Some mice received mitochondrial (5-hydroxydecanoic acid) or sarcolemmal (HMR-1098) adenosine triphosphate-dependent potassium channel blockers with or without isoflurane. Mice were then subjected to a 30-min coronary artery occlusion followed by 2 h or 2 weeks of reperfusion. Infarct size was determined at 2 h and 2 weeks of reperfusion. Cardiac function and apoptosis were determined 2 weeks after reperfusion. Isoflurane did not change hemodynamics. Isoflurane reduced infarct size after reperfusion when compared with the control groups (27.7 +/- 6.3 vs. 41.7 +/- 6.4% at 2 h and 19.6 +/- 5.9 vs. 28.8 +/- 9.0% at 2 weeks). Previous administration of 5-hydroxydecanoic acid, but not HMR-1098, abolished isoflurane-induced cardiac protection. At 2 weeks, left ventricular end-diastolic diameter was decreased significantly and end-systolic pressure and maximum and minimum dP/dt were improved by isoflurane. Isoflurane-treated mice subjected to ischemia and 2 weeks of reperfusion showed less expression of proapoptotic genes, significantly decreased expression of cleaved caspase-3, and significantly decreased deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling-positive nuclei compared with the control group. Cardiac protection induced by isoflurane against necrotic and apoptotic cell death is associated with an acute memory period that is sustained and functionally relevant 2 weeks after

Interleukin 17-producing γδT cells promote hepatic regeneration in mice.

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Rao, Raghavendra; Graffeo, Christopher S; Gulati, Rishabh; Jamal, Mohsin; Narayan, Suchithra; Zambirinis, Constantinos P; Barilla, Rocky; Deutsch, Michael; Greco, Stephanie H; Ochi, Atsuo; Tomkötter, Lena; Blobstein, Reuven; Avanzi, Antonina; Tippens, Daniel M; Gelbstein, Yisroel; Van Heerden, Eliza; Miller, George

2014-08-01

Subsets of leukocytes synergize with regenerative growth factors to promote hepatic regeneration. γδT cells are early responders to inflammation-induced injury in a number of contexts. We investigated the role of γδT cells in hepatic regeneration using mice with disruptions in Tcrd (encodes the T-cell receptor δ chain) and Clec7a (encodes C-type lectin domain family 7 member a, also known as DECTIN1). We performed partial hepatectomies on wild-type C57BL/6, CD45.1, Tcrd(-/-), or Clec7a(-/-) mice. Cells were isolated from livers of patients and mice via mechanical and enzymatic digestion. γδT cells were purified by fluorescence-activated cell sorting. In mice, partial hepatectomy up-regulated expression of CCL20 and ligands of Dectin-1, which was associated with recruitment and activation of γδT cells and their increased production of interleukin (IL)-17 family cytokines. Recruited γδT cells induced production of IL-6 by antigen-presenting cells and suppressed expression of interferon gamma by natural killer T cells, promoting hepatocyte proliferation. Absence of IL-17-producing γδT cells or deletion of Dectin-1 prevented development of regenerative phenotypes in subsets of innate immune cells. This slowed liver regeneration and was associated with reduced expression of regenerative growth factors and cell cycle regulators. Conversely, exogenous administration of IL-17 family cytokines or Dectin-1 ligands promoted regeneration. More broadly, we found that γδT cells are required for inflammatory responses mediated by IL-17 and Dectin-1. γδT cells regulate hepatic regeneration by producing IL-22 and IL-17, which have direct mitogenic effects on hepatocytes and promote a regenerative phenotype in hepatic leukocytes, respectively. Dectin-1 ligation is required for γδT cells to promote hepatic regeneration. Copyright © 2014 AGA Institute. Published by Elsevier Inc. All rights reserved.

Disruption of NBS1 gene leads to early embryonic lethality in homozygous null mice and induces specific cancer in heterozygous mice

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Kurimasa, Akihiro; Burma, Sandeep; Henrie, Melinda; Ouyang, Honghai; Osaki, Mitsuhiko; Ito, Hisao; Nagasawa, Hatsumi; Little, John B.; Oshimura, Mitsuo; Li, Gloria C.; Chen, David J.

2002-04-15

Nijmegen breakage syndrome (NBS) is a rare autosomal recessive chromosome instability syndrome characterized by microcephaly, growth retardation, immunodeficiency, and cancer predisposition, with cellular features similar to that of ataxia telangiectasia (AT). NBS results from mutations in the mammalian gene Nbs1 that codes for a 95-kDa protein called nibrin, NBS1, or p95. To establish an animal model for NBS, we attempted to generate NBS1 knockout mice. However, NBS1 gene knockouts were lethal at an early embryonic stage. NBS1 homozygous(-/-) blastocyst cells cultured in vitro showed retarded growth and subsequently underwent growth arrest within 5 days of culture. Apoptosis, assayed by TUNEL staining, was observed in NBSI homozygous(-/-) blastocyst cells cultured for four days. NBSI heterozygous(+/-) mice were normal, and exhibited no specific phenotype for at least one year. However, fibroblast cells from NBSI heterozygous(+/-) mice displayed an enhanced frequency of spontaneous transformation to anchorage-independent growth as compared to NBS1 wild-type(+/+) cells. Furthermore, heterozygous(+/-) mice exhibited a high incidence of hepatocellular carcinoma after one year compared to wild-type mice, even though no significant differences in the incidence of other tumors such as lung adenocarcinoma and lymphoma were observed. Taken together, these results strongly suggest that NBS1 heterozygosity and reduced NBSI expression induces formation of specific tumors in mice.

Antitumor HPV E7-specific CTL activity elicited by in vivo engineered exosomes produced through DNA inoculation

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Di Bonito P

2017-06-01

Full Text Available Paola Di Bonito,1 Chiara Chiozzini,2 Claudia Arenaccio,2 Simona Anticoli,2 Francesco Manfredi,2 Eleonora Olivetta,2 Flavia Ferrantelli,2 Emiliana Falcone,3 Anna Ruggieri,3 Maurizio Federico2 1Department of Infectious, Parasitic and Immunomediated Diseases, Istituto Superiore di Sanità, Rome, Italy; 2National AIDS Center, Istituto Superiore di Sanità, Rome, Italy; 3Department of Veterinary Public Health and Food Safety, Istituto Superiore di Sanità, Rome, Italy Abstract: We recently proved that exosomes engineered in vitro to deliver high amounts of HPV E7 upon fusion with the Nefmut exosome-anchoring protein elicit an efficient anti-E7 cytotoxic T lymphocyte immune response. However, in view of a potential clinic application of this finding, our exosome-based immunization strategy was faced with possible technical difficulties including industrial manufacturing, cost of production, and storage. To overcome these hurdles, we designed an as yet unproven exosome-based immunization strategy relying on delivery by intramuscular inoculation of a DNA vector expressing Nefmut fused with HPV E7. In this way, we predicted that the expression of the Nefmut/E7 vector in muscle cells would result in a continuous source of endogenous (ie, produced by the inoculated host engineered exosomes able to induce an E7-specific immune response. To assess this hypothesis, we first demonstrated that the injection of a Nefmut/green fluorescent protein-expressing vector led to the release of fluorescent exosomes, as detected in plasma of inoculated mice. Then, we observed that mice inoculated intramuscularly with a vector expressing Nefmut/E7 developed a CD8+ T-cell immune response against both Nef and E7. Conversely, no CD8+ T-cell responses were detected upon injection of vectors expressing either the wild-type Nef isoform of E7 alone, most likely a consequence of their inefficient exosome incorporation. The production of immunogenic exosomes in the DNA

Endurance performance and energy metabolism during exercise in mice with a muscle-specific defect in the control of branched-chain amino acid catabolism.

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Xu, Minjun; Kitaura, Yasuyuki; Ishikawa, Takuya; Kadota, Yoshihiro; Terai, Chihaya; Shindo, Daichi; Morioka, Takashi; Ota, Miki; Morishita, Yukako; Ishihara, Kengo; Shimomura, Yoshiharu

2017-01-01

It is known that the catabolism of branched-chain amino acids (BCAAs) in skeletal muscle is suppressed under normal and sedentary conditions but is promoted by exercise. BCAA catabolism in muscle tissues is regulated by the branched-chain α-keto acid (BCKA) dehydrogenase complex, which is inactivated by phosphorylation by BCKA dehydrogenase kinase (BDK). In the present study, we used muscle-specific BDK deficient mice (BDK-mKO mice) to examine the effect of uncontrolled BCAA catabolism on endurance exercise performance and skeletal muscle energy metabolism. Untrained control and BDK-mKO mice showed the same performance; however, the endurance performance enhanced by 2 weeks of running training was somewhat, but significantly less in BDK-mKO mice than in control mice. Skeletal muscle of BDK-mKO mice had low levels of glycogen. Metabolome analysis showed that BCAA catabolism was greatly enhanced in the muscle of BDK-mKO mice and produced branched-chain acyl-carnitine, which induced perturbation of energy metabolism in the muscle. These results suggest that the tight regulation of BCAA catabolism in muscles is important for homeostasis of muscle energy metabolism and, at least in part, for adaptation to exercise training.

Endurance performance and energy metabolism during exercise in mice with a muscle-specific defect in the control of branched-chain amino acid catabolism.

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Minjun Xu

Full Text Available It is known that the catabolism of branched-chain amino acids (BCAAs in skeletal muscle is suppressed under normal and sedentary conditions but is promoted by exercise. BCAA catabolism in muscle tissues is regulated by the branched-chain α-keto acid (BCKA dehydrogenase complex, which is inactivated by phosphorylation by BCKA dehydrogenase kinase (BDK. In the present study, we used muscle-specific BDK deficient mice (BDK-mKO mice to examine the effect of uncontrolled BCAA catabolism on endurance exercise performance and skeletal muscle energy metabolism. Untrained control and BDK-mKO mice showed the same performance; however, the endurance performance enhanced by 2 weeks of running training was somewhat, but significantly less in BDK-mKO mice than in control mice. Skeletal muscle of BDK-mKO mice had low levels of glycogen. Metabolome analysis showed that BCAA catabolism was greatly enhanced in the muscle of BDK-mKO mice and produced branched-chain acyl-carnitine, which induced perturbation of energy metabolism in the muscle. These results suggest that the tight regulation of BCAA catabolism in muscles is important for homeostasis of muscle energy metabolism and, at least in part, for adaptation to exercise training.

Tetracycline-inducible system for regulation of skeletal muscle-specific gene expression in transgenic mice

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Grill, Mischala A.; Bales, Mark A.; Fought, Amber N.; Rosburg, Kristopher C.; Munger, Stephanie J.; Antin, Parker B.

2003-01-01

Tightly regulated control of over-expression is often necessary to study one aspect or time point of gene function and, in transgenesis, may help to avoid lethal effects and complications caused by ubiquitous over-expression. We have utilized the benefits of an optimized tet-on system and a modified muscle creatine kinase (MCK) promoter to generate a skeletal muscle-specific, doxycycline (Dox) controlled over-expression system in transgenic mice. A DNA construct was generated in which the codon optimized reverse tetracycline transactivator (rtTA) was placed under control of a skeletal muscle-specific version of the mouse MCK promoter. Transgenic mice containing this construct expressed rtTA almost exclusively in skeletal muscles. These mice were crossed to a second transgenic line containing a bi-directional promoter centered on a tet responder element driving both a luciferase reporter gene and a tagged gene of interest; in this case the calpain inhibitor calpastatin. Compound hemizygous mice showed high level, Dox dependent muscle-specific luciferase activity often exceeding 10,000-fold over non-muscle tissues of the same mouse. Western and immunocytochemical analysis demonstrated similar Dox dependent muscle-specific induction of the tagged calpastatin protein. These findings demonstrate the effectiveness and flexibility of the tet-on system to provide a tightly regulated over-expression system in adult skeletal muscle. The MCKrtTA transgenic lines can be combined with other transgenic responder lines for skeletal muscle-specific over-expression of any target gene of interest.

Estrogen deficiency heterogeneously affects tissue specific stem cells in mice

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Kitajima, Yuriko; Doi, Hanako; Ono, Yusuke; Urata, Yoshishige; Goto, Shinji; Kitajima, Michio; Miura, Kiyonori; Li, Tao-Sheng; Masuzaki, Hideaki

2015-01-01

Postmenopausal disorders are frequently observed in various organs, but their relationship with estrogen deficiency and mechanisms remain unclear. As tissue-specific stem cells have been found to express estrogen receptors, we examined the hypothesis that estrogen deficiency impairs stem cells, which consequently contributes to postmenopausal disorders. Six-week-old C57BL/6 female mice were ovariectomized, following which they received 17β-estradiol replacement or vehicle (control). Sham-operated mice were used as healthy controls. All mice were killed for evaluation 2 months after treatments. Compared with the healthy control, ovariectomy significantly decreased uterine weight, which was partially recovered by 17β-estradiol replacement. Ovariectomy significantly increased the numbers of c-kit-positive hematopoietic stem/progenitor cells in bone marrow, but impaired their capacity to grow mixed cell-type colonies in vitro. Estrogen replacement further increased the numbers of c-kit-positive hematopoietic stem/progenitor cells in bone marrow, without significantly affecting colony growth in vitro. The number of CD105-positive mesenchymal stem cells in bone marrow also significantly decreased after ovariectomy, but completely recovered following estrogen replacement. Otherwise, neither ovariectomy nor estrogen replacement changed the number of Pax7-positive satellite cells, which are a skeletal muscle-type stem cell. Estrogen deficiency heterogeneously affected tissue-specific stem cells, suggesting a likely and direct relationship with postmenopausal disorders. PMID:26245252

[Effect of vitamine A on mice immune response induced by specific periodontal pathogenic bacteria-immunization].

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Lin, Xiao-Ping; Zhou, Xiao-Jia; Liu, Hong-Li; DU, Li-Li; Toshihisa, Kawai

2010-12-01

The aim of this study was to investigate the effect of vitamine-A deficiency on the induction of specific periodontal pathogenic bacteria A. actinomycetetemcomitans(Aa) immunization. BALB/c mice were fed with vitamine A-depleted diet or control regular diet throughout the whole experiment period. After 2 weeks, immunized formalin-killed Aa to build immunized models, 6 weeks later, sacrificed to determine specific antibody-IgG, IgM and sub-class IgG antibody titers in serum, and concentration of IL-10, IFN-γ, TNF-α and RANKL in T cell supernatant were measured by ELISA and T cell proliferation was measured by cintilography. SPSS 11.5 software package was used for statistical analysis. The levels of whole IgG and IgM antibody which were immunized by Aa significantly elevated, non-immune group was unable to produce any antibody. Compared with Aa immunized+RD group, the level of whole IgG in Aa immunized+VAD group was significantly higher (Pvitamin-A diet can increase the immunized mice's susceptibility to periodontal pathogenic bacteria and trigger or aggravate immune inflammatory response. Adequate vitamin A is an important factor in maintaining body health. Supported by Natural Science Foundation of Liaoning Province (Grant No.20092139) and Science and Technology Program of Shenyang Municipality (Grant No.F10-149-9-32).

Liver-specific expression of the agouti gene in transgenic mice promotes liver carcinogenesis in the absence of obesity and diabetes

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Kuklin, Alexander [ORNL; Mynatt, Randall [ORNL; Klebig, Mitch [ORNL; Kiefer, Laura [Glaxo Wellcome, Research Triangle Park, NC; Wilkison, William O [Glaxo Wellcome, Research Triangle Park, NC; Woychik, Richard P [Jackson Laboratory, The, Bar Harbor, ME; Michaud III, Edward J [ORNL

2004-01-01

Background: The agouti protein is a paracrine factor that is normally present in the skin of many species of mammals. Agouti regulates the switch between black and yellow hair pigmentation by signalling through the melanocortin 1 receptor (Mc1r) on melanocytes. Lethal yellow (Ay) and viable yellow (Avy) are dominant regulatory mutations in the mouse agouti gene that cause the wild- ype protein to be produced at abnormally high levels throughout the body. Mice harboring these mutations exhibit a pleiotropic syndrome characterized by yellow coat color, obesity, hyperglycemia, hyperinsulinemia, and increased susceptibility to hyperplasia and carcinogenesis in numerous tissues, including the liver. The goal of this research was to determine if ectopic expression of the agouti gene in the liver alone is sufficient to recapitulate any aspect of this syndrome. For this purpose, we generated lines of transgenic mice expressing high levels of agouti in the liver under the regulatory control of the albumin promoter. Expression levels of the agouti transgene in the liver were quantified by Northern blot analysis. Functional agouti protein in the liver of transgenic mice was assayed by its ability to inhibit binding of the -melanocyte stimulating hormone ( MSH) to the Mc1r. Body weight, plasma insulin and blood glucose levels were analyzed in control and transgenic mice. Control and transgenic male mice were given a single intraperitoneal injection (10 mg/kg) of the hepatocellular carcinogen, diethylnitrosamine (DEN), at 15 days of age. Mice were euthanized at 36 or 40 weeks after DEN injection and the number of tumors per liver and total liver weights were recorded. Results: The albumin-agouti transgene was expressed at high levels in the livers of mice and produced a functional agouti protein. Albumin-agouti transgenic mice had normal body weights and normal levels of blood glucose and plasma insulin, but responded to chemical initiation of the liver with an increased number

Successful adaptation to ketosis by mice with tissue-specific deficiency of ketone body oxidation.

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Cotter, David G; Schugar, Rebecca C; Wentz, Anna E; d'Avignon, D André; Crawford, Peter A

2013-02-15

During states of low carbohydrate intake, mammalian ketone body metabolism transfers energy substrates originally derived from fatty acyl chains within the liver to extrahepatic organs. We previously demonstrated that the mitochondrial enzyme coenzyme A (CoA) transferase [succinyl-CoA:3-oxoacid CoA transferase (SCOT), encoded by nuclear Oxct1] is required for oxidation of ketone bodies and that germline SCOT-knockout (KO) mice die within 48 h of birth because of hyperketonemic hypoglycemia. Here, we use novel transgenic and tissue-specific SCOT-KO mice to demonstrate that ketone bodies do not serve an obligate energetic role within highly ketolytic tissues during the ketogenic neonatal period or during starvation in the adult. Although transgene-mediated restoration of myocardial CoA transferase in germline SCOT-KO mice is insufficient to prevent lethal hyperketonemic hypoglycemia in the neonatal period, mice lacking CoA transferase selectively within neurons, cardiomyocytes, or skeletal myocytes are all viable as neonates. Like germline SCOT-KO neonatal mice, neonatal mice with neuronal CoA transferase deficiency exhibit increased cerebral glycolysis and glucose oxidation, and, while these neonatal mice exhibit modest hyperketonemia, they do not develop hypoglycemia. As adults, tissue-specific SCOT-KO mice tolerate starvation, exhibiting only modestly increased hyperketonemia. Finally, metabolic analysis of adult germline Oxct1(+/-) mice demonstrates that global diminution of ketone body oxidation yields hyperketonemia, but hypoglycemia emerges only during a protracted state of low carbohydrate intake. Together, these data suggest that, at the tissue level, ketone bodies are not a required energy substrate in the newborn period or during starvation, but rather that integrated ketone body metabolism mediates adaptation to ketogenic nutrient states.

Establishment of clinically relevant radioresistant cell lines and their characteristics

International Nuclear Information System (INIS)

Fukumoto, Manabu; Kuwahara, Yoshikazu; Suzuki, Masatoshi

2014-01-01

Although radiotherapy is one of the major therapeutic modalities for eradicating malignant tumors, the existence of radioresistant cells remains one of the most critical obstacles. Standard radiotherapy consists of fractionated radiation (FR) of 2-Gy X-rays once a day, 5 days a week, over 60 Gy in total. To understand the characteristics of radioresistant cells and to develop more effective radiotherapy, we have established novel radioresistant cell lines by long-term (> 5 years) exposure to moderate doses of fractionated X-rays. While all the parental human cancer cells ceased, their radioresistant derivatives continue to proliferate with daily exposure to 2-Gy FR for more than 30 days. We have coined those cells as 'clinically relevant radioresistant' (CRR) cells. Transplanted tumors into nude mice were also CRR, indicating that CRR cell lines are powerful tools to improve cancer radiotherapy. We have shown that the suppression of autophagic cell death but not apoptosis was mainly involved in cellular radioresistance. An inhibitor of the mTOR pathway which enhances autophagy was effective to overcome CRR tumors induced in nude mice. But the underlined mechanism was not through the inhibition of autophagy. Guanine nucleotide-binding protein 1 (GBP1) over expression was necessary for maintaining the CRR phenotype, but radioresistant cells were not necessarily cancer stem cells (CSCs). Targeting GBP1 positive cancer cells may be a more efficient method in conquering cancer than targeting CSCs. Slight but significant radioresistance was acquired by 0.5 Gy/12 hrs of long-term FR exposures to parental cells for more than 31 days in accordance with cyclinD1 over expression. This acquired radioresistance (ARR) was stably maintained in the tumor cells even on 31 days after the cessation of 0.5-Gy FR. Present observations give a mechanistic insight for ARR of tumor cells through long-term FR exposure, and provide novel therapeutic targets for radiosensitization

Highly specific expression of luciferase gene in lungs of naive nude mice directed by prostate-specific antigen promoter

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Li Hongwei; Li Jinzhong; Helm, Gregory A.; Pan Dongfeng

2005-01-01

PSA promoter has been demonstrated the utility for tissue-specific toxic gene therapy in prostate cancer models. Characterization of foreign gene overexpression in normal animals elicited by PSA promoter should help evaluate therapy safety. Here we constructed an adenovirus vector (AdPSA-Luc), containing firefly luciferase gene under the control of the 5837 bp long prostate-specific antigen promoter. A charge coupled device video camera was used to non-invasively image expression of firefly luciferase in nude mice on days 3, 7, 11 after injection of 2 x 10 9 PFU of AdPSA-Luc virus via tail vein. The result showed highly specific expression of the luciferase gene in lungs of mice from day 7. The finding indicates the potential limitations of the suicide gene therapy of prostate cancer based on selectivity of PSA promoter. By contrary, it has encouraging implications for further development of vectors via PSA promoter to enable gene therapy for pulmonary diseases

Activation of specific cellular immunity toward murine leukemia in mice rejecting syngeneic somatic hybrid cells

International Nuclear Information System (INIS)

Liang, W.; Cohen, E.P.

1977-01-01

ASL-1 x LM(TK) - somatic hybrid cells form both H-2/sup a/ and H-2/sup k/ antigen complexes. After forming a localized tumor in syngeneic (A/J x C 3 H/HeJ)F 1 mice, they are rejected. Such mice are resistant to otherwise invariably lethal injections of ASL-1 cells, surviving for prolonged and, in some instances, indefinite periods. To examine the basis of immunity, the capacity of spleen cells from mice rejecting hybrid cells to stimulate the release of 51 Cr from labeled ASL-1 cells was investigated. Cells from the spleens of mice rejecting ASL-1 x LM(TK) - cells stimulated the release of 51 Cr from labeled ASL-1 cells, but not from Ehrlich ascites or P815 cells. Cells from mice injected with mitomycin-C-treated ASL-1 cells led to the release of 51 Cr from labeled ASL-1 cells as well, but the extent of 51 Cr release was approximately one-third as occurred in the presence of cells from hybrid cell-injected mice. Cells from noninjected mice or from mice injected with LM(TK) - cells failed to lead to the specific release of 51 Cr from ASL-1 cells. The presence of unlabeled ASL-1 cells, but not Ehrlich ascites cells, competitively inhibited the spleen cell-stimulated release of 51 Cr from labeled ASL-1 cells. Sera from A/J mice injected with mitomycin-C-treated ASL-1 cells contained antibodies specific for the tumor-associated antigen of ASL-1 cells

Oleanolic acid alters bile acid metabolism and produces cholestatic liver injury in mice

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Liu, Jie, E-mail: JLiu@kumc.edu [University of Kansas Medical Center, Kansas City, KS 66160 (United States); Zunyi Medical College, Zunyi 563003 (China); Lu, Yuan-Fu [University of Kansas Medical Center, Kansas City, KS 66160 (United States); Zunyi Medical College, Zunyi 563003 (China); Zhang, Youcai; Wu, Kai Connie [University of Kansas Medical Center, Kansas City, KS 66160 (United States); Fan, Fang [Cytopathology, University of Kansas Medical Center, Kansas City, KS 66160 (United States); Klaassen, Curtis D. [University of Kansas Medical Center, Kansas City, KS 66160 (United States)

2013-11-01

Oleanolic acid (OA) is a triterpenoids that exists widely in plants. OA is effective in protecting against hepatotoxicants. Whereas a low dose of OA is hepatoprotective, higher doses and longer-term use of OA produce liver injury. This study characterized OA-induced liver injury in mice. Adult C57BL/6 mice were given OA at doses of 0, 22.5, 45, 90, and 135 mg/kg, s.c., daily for 5 days, and liver injury was observed at doses of 90 mg/kg and above, as evidenced by increases in serum activities of alanine aminotransferase and alkaline phosphatase, increases in serum total bilirubin, as well as by liver histopathology. OA-induced cholestatic liver injury was further evidenced by marked increases of both unconjugated and conjugated bile acids (BAs) in serum. Gene and protein expression analysis suggested that livers of OA-treated mice had adaptive responses to prevent BA accumulation by suppressing BA biosynthetic enzyme genes (Cyp7a1, 8b1, 27a1, and 7b1); lowering BA uptake transporters (Ntcp and Oatp1b2); and increasing a BA efflux transporter (Ostβ). OA increased the expression of Nrf2 and its target gene, Nqo1, but decreased the expression of AhR, CAR and PPARα along with their target genes, Cyp1a2, Cyp2b10 and Cyp4a10. OA had minimal effects on PXR and Cyp3a11. Taken together, the present study characterized OA-induced liver injury, which is associated with altered BA homeostasis, and alerts its toxicity potential. - Highlights: • Oleanolic acid at higher doses and long-term use may produce liver injury. • Oleanolic acid increased serum ALT, ALP, bilirubin and bile acid concentrations. • OA produced feathery degeneration, inflammation and cell death in the liver. • OA altered bile acid homeostasis, affecting bile acid synthesis and transport.

Specific Inflammatory Stimuli Lead to Distinct Platelet Responses in Mice and Humans.

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Lea M Beaulieu

Full Text Available Diverse and multi-factorial processes contribute to the progression of cardiovascular disease. These processes affect cells involved in the development of this disease in varying ways, ultimately leading to atherothrombosis. The goal of our study was to compare the differential effects of specific stimuli--two bacterial infections and a Western diet--on platelet responses in ApoE-/- mice, specifically examining inflammatory function and gene expression. Results from murine studies were verified using platelets from participants of the Framingham Heart Study (FHS; n = 1819 participants.Blood and spleen samples were collected at weeks 1 and 9 from ApoE-/- mice infected with Porphyromonas gingivalis or Chlamydia pneumoniae and from mice fed a Western diet for 9 weeks. Transcripts based on data from a Western diet in ApoE-/- mice were measured in platelet samples from FHS using high throughput qRT-PCR.At week 1, both bacterial infections increased circulating platelet-neutrophil aggregates. At week 9, these cells individually localized to the spleen, while Western diet resulted in increased platelet-neutrophil aggregates in the spleen only. Microarray analysis of platelet RNA from infected or Western diet-fed mice at week 1 and 9 showed differential profiles. Genes, such as Serpina1a, Ttr, Fgg, Rpl21, and Alb, were uniquely affected by infection and diet. Results were reinforced in platelets obtained from participants of the FHS.Using both human studies and animal models, results demonstrate that variable sources of inflammatory stimuli have the ability to influence the platelet phenotype in distinct ways, indicative of the diverse function of platelets in thrombosis, hemostasis, and immunity.

Application of chimeric mice with humanized liver for study of human-specific drug metabolism.

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Bateman, Thomas J; Reddy, Vijay G B; Kakuni, Masakazu; Morikawa, Yoshio; Kumar, Sanjeev

2014-06-01

Human-specific or disproportionately abundant human metabolites of drug candidates that are not adequately formed and qualified in preclinical safety assessment species pose an important drug development challenge. Furthermore, the overall metabolic profile of drug candidates in humans is an important determinant of their drug-drug interaction susceptibility. These risks can be effectively assessed and/or mitigated if human metabolic profile of the drug candidate could reliably be determined in early development. However, currently available in vitro human models (e.g., liver microsomes, hepatocytes) are often inadequate in this regard. Furthermore, the conduct of definitive radiolabeled human ADME studies is an expensive and time-consuming endeavor that is more suited for later in development when the risk of failure has been reduced. We evaluated a recently developed chimeric mouse model with humanized liver on uPA/SCID background for its ability to predict human disposition of four model drugs (lamotrigine, diclofenac, MRK-A, and propafenone) that are known to exhibit human-specific metabolism. The results from these studies demonstrate that chimeric mice were able to reproduce the human-specific metabolite profile for lamotrigine, diclofenac, and MRK-A. In the case of propafenone, however, the human-specific metabolism was not detected as a predominant pathway, and the metabolite profiles in native and humanized mice were similar; this was attributed to the presence of residual highly active propafenone-metabolizing mouse enzymes in chimeric mice. Overall, the data indicate that the chimeric mice with humanized liver have the potential to be a useful tool for the prediction of human-specific metabolism of xenobiotics and warrant further investigation.

Cardiomyocyte specific deletion of Crif1 causes mitochondrial cardiomyopathy in mice.

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Juhee Shin

Full Text Available Mitochondria are key organelles dedicated to energy production. Crif1, which interacts with the large subunit of the mitochondrial ribosome, is indispensable for the mitochondrial translation and membrane insertion of respiratory subunits. To explore the physiological function of Crif1 in the heart, Crif1(f/f mice were crossed with Myh6-cre/Esr1 transgenic mice, which harbor cardiomyocyte-specific Cre activity in a tamoxifen-dependent manner. The tamoxifen injections were given at six weeks postnatal, and the mutant mice survived only five months due to hypertrophic heart failure. In the mutant cardiac muscles, mitochondrial mass dramatically increased, while the inner structure was altered with lack of cristae. Mutant cardiac muscles showed decreased rates of oxygen consumption and ATP production, suggesting that Crif1 plays a critical role in the maintenance of both mitochondrial structure and respiration in cardiac muscles.

Activation of inflammatory signaling by lipopolysaccharide produces a prolonged increase of voluntary alcohol intake in mice

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Blednov, Y.A.; Benavidez, J.M.; Geil, C.; Perra, S.; Morikawa, H.; Harris, R.A.

2011-01-01

Previous studies showed that mice with genetic predisposition for high alcohol consumption as well as human alcoholics show changes in brain expression of genes related to immune signaling. In addition, mutant mice lacking genes related to immune function show decreased alcohol consumption (Blednov et al., in press), suggesting that immune signaling promotes alcohol consumption. To test the possibility that activation of immune signaling will increase alcohol consumption, we treated mice with lipopolysaccaride (LPS; 1 mg/kg, i.p.) and tested alcohol consumption in the continuous two-bottle choice test. To take advantage of the long-lasting activation of brain immune signaling by LPS, we measured drinking beginning one week or one month after LPS treatment and continued the studies for several months. LPS produced persistent increases in alcohol consumption in C57/Bl6 J (B6) inbred mice, FVBxB6F1 and B6xNZBF1 hybrid mice, but not in FVB inbred mice. To determine if this effect of LPS is mediated through binding to TLR4, we tested mice lacking CD14, a key component of TLR4 signaling. These null mutants showed no increase of alcohol intake after treatment with LPS. LPS treatment decreased ethanol-conditioned taste aversion but did not alter ethanol-conditioned place preference (B6xNZBF1 mice). Electro-physiological studies of dopamine neurons in the ventral tegmental area showed that pretreatment of mice with LPS decreased the neuronal firing rate. These results suggest that activation of immune signaling promotes alcohol consumption and alters certain aspects of alcohol reward/aversion. PMID:21266194

Generation of transgenic mice producing fungal xylanase in the ...

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DR TONUKARI NYEROVWO

express exogenous digestive enzymes, since a single- stomached animal, such as a pig, can secret .... transgenic founder mice; 1 to15 are fifteen wild-type founder mice; M, marke; β-actin, endogenous control. (C) Identification of transgenic mice by ... 61.48±0.34%), gross energy digestibility (WT vs. TG = 68.79±0.51% vs.

Cis-acting sequences from a human surfactant protein gene confer pulmonary-specific gene expression in transgenic mice

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Korfhagen, T.R.; Glasser, S.W.; Wert, S.E.; Bruno, M.D.; Daugherty, C.C.; McNeish, J.D.; Stock, J.L.; Potter, S.S.; Whitsett, J.A. (Cincinnati College of Medicine, OH (USA))

1990-08-01

Pulmonary surfactant is produced in late gestation by developing type II epithelial cells lining the alveolar epithelium of the lung. Lack of surfactant at birth is associated with respiratory distress syndrome in premature infants. Surfactant protein C (SP-C) is a highly hydrophobic peptide isolated from pulmonary tissue that enhances the biophysical activity of surfactant phospholipids. Like surfactant phospholipid, SP-C is produced by epithelial cells in the distal respiratory epithelium, and its expression increases during the latter part of gestation. A chimeric gene containing 3.6 kilobases of the promoter and 5{prime}-flanking sequences of the human SP-C gene was used to express diphtheria toxin A. The SP-C-diphtheria toxin A fusion gene was injected into fertilized mouse eggs to produce transgenic mice. Affected mice developed respiratory failure in the immediate postnatal period. Morphologic analysis of lungs from affected pups showed variable but severe cellular injury confined to pulmonary tissues. Ultrastructural changes consistent with cell death and injury were prominent in the distal respiratory epithelium. Proximal components of the tracheobronchial tree were not severely affected. Transgenic animals were of normal size at birth, and structural abnormalities were not detected in nonpulmonary tissues. Lung-specific diphtheria toxin A expression controlled by the human SP-C gene injured type II epithelial cells and caused extensive necrosis of the distal respiratory epithelium. The absence of type I epithelial cells in the most severely affected transgenic animals supports the concept that developing type II cells serve as precursors to type I epithelial cells.

Specificity of anti-tau antibodies when analyzing mice models of Alzheimer's disease: problems and solutions.

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Franck R Petry

Full Text Available Aggregates of hyperphosphorylated tau protein are found in a group of diseases called tauopathies, which includes Alzheimer's disease. The causes and consequences of tau hyperphosphorylation are routinely investigated in laboratory animals. Mice are the models of choice as they are easily amenable to transgenic technology; consequently, their tau phosphorylation levels are frequently monitored by Western blotting using a panel of monoclonal/polyclonal anti-tau antibodies. Given that mouse secondary antibodies can recognize endogenous mouse immunoglobulins (Igs and the possible lack of specificity with some polyclonal antibodies, non-specific signals are commonly observed. Here, we characterized the profiles of commonly used anti-tau antibodies in four different mouse models: non-transgenic mice, tau knock-out (TKO mice, 3xTg-AD mice, and hypothermic mice, the latter a positive control for tau hyperphosphorylation. We identified 3 tau monoclonal antibody categories: type 1, characterized by high non-specificity (AT8, AT180, MC1, MC6, TG-3, type 2, demonstrating low non-specificity (AT270, CP13, CP27, Tau12, TG5, and type 3, with no non-specific signal (DA9, PHF-1, Tau1, Tau46. For polyclonal anti-tau antibodies, some displayed non-specificity (pS262, pS409 while others did not (pS199, pT205, pS396, pS404, pS422, A0024. With monoclonal antibodies, most of the interfering signal was due to endogenous Igs and could be eliminated by different techniques: i using secondary antibodies designed to bind only non-denatured Igs, ii preparation of a heat-stable fraction, iii clearing Igs from the homogenates, and iv using secondary antibodies that only bind the light chain of Igs. All of these techniques removed the non-specific signal; however, the first and the last methods were easier and more reliable. Overall, our study demonstrates a high risk of artefactual signal when performing Western blotting with routinely used anti-tau antibodies, and proposes

Forebrain-Specific Loss of BMPRII in Mice Reduces Anxiety and Increases Object Exploration

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McBrayer, Zofeyah L.; Dimova, Jiva; Pisansky, Marc T.; Sun, Mu; Beppu, Hideyuki; Gewirtz, Jonathan C.; O’Connor, Michael B.

2015-01-01

To investigate the role of Bone Morphogenic Protein Receptor Type II (BMPRII) in learning, memory, and exploratory behavior in mice, a tissue-specific knockout of BMPRII in the post-natal hippocampus and forebrain was generated. We found that BMPRII mutant mice had normal spatial learning and memory in the Morris water maze, but showed significantly reduced swimming speeds with increased floating behavior. Further analysis using the Porsolt Swim Test to investigate behavioral despair did not ...

Hsp65-producing Lactococcus lactis prevents experimental autoimmune encephalomyelitis in mice by inducing CD4+LAP+ regulatory T cells

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Rezende, Rafael M.; Oliveira, Rafael P.; Medeiros, Samara R.; Gomes-Santos, Ana C.; Alves, Andrea C.; Loli, Flávia G.; Guimarães, Mauro A.F.; Amaral, Sylvia S.; da Cunha, André P.; Weiner, Howard L.; Azevedo, Vasco; Miyoshi, Anderson; Faria, Ana M.C.

2013-01-01

Heat shock proteins (Hsps) participate in the cellular response to stress and they are hiperexpressed in inflammatory conditions. They are also known to play a major role in immune modulation, controlling, for instance, autoimmune responses. In this study, we showed that oral administration of a recombinant Lactococcus lactis strain that produces and releases LPS-free Hsp65 prevented the development of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice. This was confirmed by the reduced inflammatory cell infiltrate and absence of injury signs in the spinal cord. The effect was associated with reduced IL-17 and increased IL-10 production in mesenteric lymph node and spleen cell cultures. Hsp65-producing-L. lactis-fed mice had a remarkable increase in the number of natural and inducible CD4+Foxp3+ regulatory T (Treg) cells and CD4+LAP+ (Latency-associated peptide) Tregs - which express the membrane-bound TGF-β - in spleen, inguinal and mesenteric lymph nodes as well as in spinal cord. Moreover, many Tregs co-expressed Foxp3 and LAP. In vivo depletion of LAP+ cells abrogated the effect of Hsp65-producing L. lactis in EAE prevention and worsened disease in medium-fed mice. Thus, Hsp65-L.lactis seems to boost this critical regulatory circuit involved in controlling EAE development in mice. PMID:22939403

Alterations in regulatory T cells induced by specific oligosaccharides improve vaccine responsiveness in mice.

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Marcel A Schijf

Full Text Available Prophylactic vaccinations are generally performed to protect naïve individuals with or without suppressed immune responsiveness. In a mouse model for Influenza vaccinations the specific alterations of CD4(+CD25(+Foxp3(+ regulatory T-cells (Tregs in the immune modulation induced by orally supplied oligosaccharides containing scGOS/lcFOS/pAOS was assessed. This dietary intervention increased vaccine specific DTH responses. In addition, a significant increased percentage of T-bet(+ (Th1 activated CD69(+CD4(+ T cells (p<0.001 and reduced percentage of Gata-3(+ (Th2 activated CD69(+CD4(+T cells (p<0.001 was detected in the mesenteric lymph nodes (MLN of mice receiving scGOS/lcFOS/pAOS compared to control mice. Although no difference in the number or percentage of Tregs (CD4(+Foxp3(+ could be determined after scGOS/lcFOS/pAOS intervention, the percentage of CXCR3 (+ /T-bet(+ (Th1-Tregs was significantly reduced (p<0.05 in mice receiving scGOS/lcFOS/pAOS as compared to mice receiving placebo diets. Moreover, although no absolute difference in suppressive capacity could be detected, an alteration in cytokine profile suggests a regulatory T cell shift towards a reducing Th1 suppression profile, supporting an improved vaccination response.These data are indicative for improved vaccine responsiveness due to reduced Th1 suppressive capacity in the Treg population of mice fed the oligosaccharide specific diet, showing compartmentalization within the Treg population. The modulation of Tregs to control immune responses provides an additional arm of intervention using alternative strategies possibly leading to the development of improved vaccines.

Specificity in Sociality: Mice and Prairie Voles Exhibit Different Patterns of Peer Affiliation

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Beery, Annaliese K.; Christensen, Jennifer D; Lee, Nicole S.; Blandino, Katrina L.

2018-01-01

Social behavior is often described as a unified concept, but highly social (group-living) species exhibit distinct social structures and may make different social decisions. Prairie voles (Microtus ochrogaster) are socially monogamous rodents that often reside in extended family groups, and exhibit robust preferences for familiar social partners (same- and opposite-sex) during extended choice tests, although short-term preferences are not known. Mice (Mus musculus) are gregarious and colonial, but in brief laboratory tests of social preference they typically prefer social novelty. This preference for novel vs. familiar peers may represent a species-specific difference in social decision-making between mice and prairie voles. However, the tests used to measure preferences in each species differ markedly in duration and degree of contact, such that the behaviors cannot be directly compared. We assessed whether social preferences for novelty or familiarity differed between mice and prairie voles of both sexes when assessed with matching protocols: the sociability/social preference test (SPT) typically used in mice (short, no direct contact), and the partner preference test (PPT) used in voles (long, direct contact). A subset of voles also underwent a PPT using barriers (long, no direct contact). In the short SPT, behavior did not differ between species. In the longer test, pronounced partner preferences emerged in prairie voles, but mice exhibited no social preferences and rarely huddled. No sex differences were evident in either test. Direct physical contact was required for partner preferences in huddling time in voles, but preference for the partner chamber was evident with or without contact. Both prairie voles and mice are social, but they exhibit important differences in the specificity and extent of their social behavior. While mice are often used to study social approach and other behaviors, voles are a more suitable species for the study of selective social

Specificity in Sociality: Mice and Prairie Voles Exhibit Different Patterns of Peer Affiliation

Directory of Open Access Journals (Sweden)

Annaliese K. Beery

2018-03-01

Full Text Available Social behavior is often described as a unified concept, but highly social (group-living species exhibit distinct social structures and may make different social decisions. Prairie voles (Microtus ochrogaster are socially monogamous rodents that often reside in extended family groups, and exhibit robust preferences for familiar social partners (same- and opposite-sex during extended choice tests, although short-term preferences are not known. Mice (Mus musculus are gregarious and colonial, but in brief laboratory tests of social preference they typically prefer social novelty. This preference for novel vs. familiar peers may represent a species-specific difference in social decision-making between mice and prairie voles. However, the tests used to measure preferences in each species differ markedly in duration and degree of contact, such that the behaviors cannot be directly compared. We assessed whether social preferences for novelty or familiarity differed between mice and prairie voles of both sexes when assessed with matching protocols: the sociability/social preference test (SPT typically used in mice (short, no direct contact, and the partner preference test (PPT used in voles (long, direct contact. A subset of voles also underwent a PPT using barriers (long, no direct contact. In the short SPT, behavior did not differ between species. In the longer test, pronounced partner preferences emerged in prairie voles, but mice exhibited no social preferences and rarely huddled. No sex differences were evident in either test. Direct physical contact was required for partner preferences in huddling time in voles, but preference for the partner chamber was evident with or without contact. Both prairie voles and mice are social, but they exhibit important differences in the specificity and extent of their social behavior. While mice are often used to study social approach and other behaviors, voles are a more suitable species for the study of

Decrease of virulence for BALB/c mice produced by continuous subculturing of Nocardia brasiliensis.

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Almaguer-Chávez, Janeth A; Welsh, Oliverio; Lozano-Garza, Hector G; Said-Fernández, Salvador; Romero-Díaz, Víktor J; Ocampo-Candiani, Jorge; Vera-Cabrera, Lucio

2011-10-26

Subculturing has been extensively used to attenuate human pathogens. In this work we studied the effect of continuous subculturing of Nocardia brasiliensis HUJEG-1 on virulence in a murine model. Nocardia brasiliensis HUJEG-1 was subcultured up to 130 times on brain heart infusion over four years. BALB/c mice were inoculated in the right foot pad with the bacteria subcultured 0, 40, 80, 100 and 130 times (T0, T40, T80 T100 and T130). The induction of resistance was tested by using T130 to inoculate a group of mice followed by challenge with T0 12 weeks later. Biopsies were taken from the newly infected foot-pad and immunostained with antibodies against CD4, CD8 and CD14 in order to analyze the in situ immunological changes. When using T40, T80 T100 and T130 as inoculums we observed lesions in 10, 5, 0 and 0 percent of the animals, respectively, at the end of 12 weeks. In contrast, their controls produced mycetoma in 80, 80, 70 and 60% of the inoculated animals. When studying the protection of T130, we observed a partial resistance to the infection. Immunostaining revealed an intense CD4+ lymphocytic and macrophage infiltrate in healing lesions. After 130 in vitro passages of N. brasiliensis HUJEG-1 a severe decrease in its virulence was observed. Immunization of BALB/c mice, with these attenuated cells, produced a state of partial resistance to infection with the non-subcultured isolate.

Decrease of virulence for BALB/c mice produced by continuous subculturing of Nocardia brasiliensis

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Ocampo-Candiani Jorge

2011-10-01

Full Text Available Abstract Background Subculturing has been extensively used to attenuate human pathogens. In this work we studied the effect of continuous subculturing of Nocardia brasiliensis HUJEG-1 on virulence in a murine model. Methods Nocardia brasiliensis HUJEG-1 was subcultured up to 130 times on brain heart infusion over four years. BALB/c mice were inoculated in the right foot pad with the bacteria subcultured 0, 40, 80, 100 and 130 times (T0, T40, T80 T100 and T130. The induction of resistance was tested by using T130 to inoculate a group of mice followed by challenge with T0 12 weeks later. Biopsies were taken from the newly infected foot-pad and immunostained with antibodies against CD4, CD8 and CD14 in order to analyze the in situ immunological changes. Results When using T40, T80 T100 and T130 as inoculums we observed lesions in 10, 5, 0 and 0 percent of the animals, respectively, at the end of 12 weeks. In contrast, their controls produced mycetoma in 80, 80, 70 and 60% of the inoculated animals. When studying the protection of T130, we observed a partial resistance to the infection. Immunostaining revealed an intense CD4+ lymphocytic and macrophage infiltrate in healing lesions. Conclusions After 130 in vitro passages of N. brasiliensis HUJEG-1 a severe decrease in its virulence was observed. Immunization of BALB/c mice, with these attenuated cells, produced a state of partial resistance to infection with the non-subcultured isolate.

Homeopathic pathogenetic trials produce specific symptoms different from placebo.

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Möllinger, Heribert; Schneider, Rainer; Walach, Harald

2009-04-01

Homeopathy uses information gathered from healthy volunteers taking homeopathic substances (pathogenetic trials) for clinical treatment. It is controversial whether such studies produce symptoms different from those produced by placebo. To test whether homeopathic preparations produce different symptoms than placebo in healthy volunteers. Three armed, double-blind, placebo controlled randomised experimental pathogenetic study in 25 healthy volunteers who took either one of two homeopathic remedies, Natrum muriaticum and Arsenicum album in 30CH or identical placebo. Main outcome parameter was the number of remedy-specific symptoms per group. On average, 6 symptoms typical for Arsenicum album were experienced by participants taking arsenicum album, 5 symptoms typical for Natrum muriaticum by those taking natrum muriaticum, and 11 non-specific symptoms by those in the placebo group. Differences were significant overall (Kruskall Wallis test, p = 0.0002,) and significantly different from placebo (Mann-Whitney test, p = 0.001). Homeopathic remedies produce different symptoms than placebo. Copyright (c) 2009 S. Karger AG, Basel.

Enhanced humoral and HLA-A2-restricted dengue virus-specific T-cell responses in humanized BLT NSG mice

Science.gov (United States)

Jaiswal, Smita; Pazoles, Pamela; Woda, Marcia; Shultz, Leonard D; Greiner, Dale L; Brehm, Michael A; Mathew, Anuja

2012-01-01

Dengue is a mosquito-borne viral disease of humans, and animal models that recapitulate human immune responses or dengue pathogenesis are needed to understand the pathogenesis of the disease. We recently described an animal model for dengue virus (DENV) infection using humanized NOD-scid IL2rγnull mice (NSG) engrafted with cord blood haematopoietic stem cells. We sought to further improve this model by co-transplantation of human fetal thymus and liver tissues into NSG (BLT-NSG) mice. Enhanced DENV-specific antibody titres were found in the sera of BLT-NSG mice compared with human cord blood haematopoietic stem cell-engrafted NSG mice. Furthermore, B cells generated during the acute phase and in memory from splenocytes of immunized BLT-NSG mice secreted DENV-specific IgM antibodies with neutralizing activity. Human T cells in engrafted BLT-NSG mice secreted interferon-γ in response to overlapping DENV peptide pools and HLA-A2 restricted peptides. The BLT-NSG mice will allow assessment of human immune responses to DENV vaccines and the effects of previous immunity on subsequent DENV infections. PMID:22384859

Activation of Kupffer Cells Is Associated with a Specific Dysbiosis Induced by Fructose or High Fat Diet in Mice.

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Gladys Ferrere

Full Text Available The increase consumption of fructose in diet is associated with liver inflammation. As a specific fructan substrate, fructose may modify the gut microbiota which is involved in obesity-induced liver disease. Here, we aimed to assess whether fructose-induced liver damage was associated with a specific dysbiosis, especially in mice fed a high fat diet (HFD. To this end, four groups of mice were fed with normal and HFD added or not with fructose. Body weight and glucose sensitivity, liver inflammation, dysbiosis and the phenotype of Kupffer cells were determined after 16 weeks of diet. Food intake was increased in the two groups of mice fed with the HFD. Mice fed with HFD and fructose showed a higher infiltration of lymphocytes into the liver and a lower inflammatory profile of Kupffer cells than mice fed with the HFD without fructose. The dysbiosis associated with diets showed that fructose specifically prevented the decrease of Mouse intestinal bacteria in HFD fed mice and increased Erysipelotrichi in mice fed with fructose, independently of the amount of fat. In conclusion, fructose, used as a sweetener, induced a dysbiosis which is different in presence of fat in the diet. Consequently, the activation of Kupffer cells involved in mice model of HFD-induced liver inflammation was not observed in an HFD/fructose combined diet. These data highlight that the complexity of diet composition could highly impact the development of liver lesions during obesity. Specific dysbiosis associated with the diet could explain that the progressions of liver damage are different.

Atypical patterns of neural infection produced in mice by drug-resistant strains of herpes simplex virus.

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Field, H J; Anderson, J R; Wildy, P

1982-03-01

Mice inoculated intracerebrally (i.c.) with a mutant strain of HSV were found to develop cataracts 1 to 2 months after inoculation. Cataract formation was subsequently shown to follow an acute retinitis which commenced within 1 week of inoculation. The mutant had been selected for high resistance to the nucleoside analogue acyclovir and has been shown previously to be defective in the induction of thymidine kinase and also to express an altered DNA polymerase. The LD50 for mice inoculated i.c. was greater than 10(5) p.f.u. compared with approx 7 p.f.u. for the parental strain. Studies of virus replication following i.c. inoculation with a sublethal dose of the mutant revealed that only small amounts of infectious virus were produced in the brain, but during a period from 6 to 12 days after inoculation vigorous replication occurred in retinal tissue, producing very high titres of virus.

Eosinophilic spleen colonies are produced in rat-marrow-transplanted but not in murine-marrow-transplanted mice

International Nuclear Information System (INIS)

Szabo, L.G.; Kelemen, E.

1988-01-01

Differential counts of about 5000 splenic clusters and colonies developing in whole-body-irradiated mice and rats were made, using semi-serial histological sections prepared 9 to 12 d after transplantation with bone marrow haemopoietic cells. The investigated mouse and rat spleens were from syngeneically, allogeneically, or xenogeneically transplanted recipients. Splenic eosinophil clusters were always found when rat eosinophil-producing progenitors were present in the inoculum, whereas murine inocula failed to produce splenic eosinophilic clusters even in the syngeneic mouse. The limiting factor in the production pf splenic eosinophilic clusters was the appropriate donor progenitor/committed stem cell itself. Changes in the percentages of eosinophil clusters with the number of injected cells and with increased doses of irradiation, as well as formation of rat eosinophil colonies in mice, as against mainly clusters in rats, themselves show that regulatory mechanisms of the recipients also play a role. These regulatory mechanisms cannot be attributed to the splenic microenvironment. (author)

Characterization and specificity of probiotics to prevent salmonella infection in mice

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Ana Andino

2014-08-01

Full Text Available Background: Probiotic strains of bacteria can prevent Salmonella from causing disease by preventing the pathogen from colonizing the intestines. Two strains of probiotics, Lactobacillus acidophilius and Pediococcus spp, that were obtained from poultry fecal samples have been shown to be efficacious in poultry. The objective of this study was to determine if these strains of probiotics could prevent salmonellosis in a mouse model. Methods: First, both strains of probiotics were evaluated for in vitro efficacy to inhibit the growth of and interfere with virulence gene regulation in Salmonella enterica. For in vivo efficacy, mice was used which models Typhoid illness. Mice were divided into 2 groups: Control and treatment, Lactobacillus and Pediococcus (LP; 108 Log CFU. Two experiments were conducted. In the first experiment, the mice were treated with LP in water for the first two days of the experiment and challenged with Salmonella at day three. In the second experiment, the LP treatment was given in the water for 10 days and challenge was performed on day 11. In both experiments, at day 20 post-challenge, all mice were sacrificed, intestinal tracts and organs removed and cultured for Salmonella. Results: The probiotic strains inhibited the growth of Salmonella and down-regulation of virulence genes was noted, but dependent on the strain of Salmonella being evaluated. For the in vivo experiment, the probiotics did not afford the mice protection from infection and increasing the length of time the probiotics were administered did not improve the efficacy of the probiotics. Conclusions: It appears that these strains of probiotic bacteria are effective against Salmonella in vitro. However, these isolates did not afford protection from Salmonella infection to mice which may be due to host specifity as these isolates were obtained from poultry

Specific anti-EL4-lymphoma immunity in mice cured 2 years earlier with doxorubicin and interleukin-2.

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Ehrke, M J; Verstovsek, S; Zaleskis, G; Ho, R L; Ujházy, P; Maccubbin, D L; Mihich, E

1996-05-01

This laboratory has reported the conditions for an effective, non-toxic, chemoimmunotherapy utilizing doxorubicin in combination with prolonged administration of interleukin-2 and the identification of the critical role of activated CD8+ T cells in the therapeutic effect. Mice (C57BL/6) cured in those studies have been followed for the remainder of their life spans. These mice, approximately 2 months of age when initially inoculated with syngeneic EL4 lymphoma, survived for more than 2 years, the normal life span of C57BL/6 mice. Mice 4 months old reinoculated with the EL4 cells all survived. At about 1 year of age mice were sacrificed and the ability of their thymocytes and splenocytes to develop specific CD8+ anti-EL4 activity was as high as it had been at the time of tumor rejection. At about 2 years of age EL4 was reimplanted into mice; all of them survived. These surviving mice, at 2 years 2 months of age, as well as a group of 2-year-old mice not rechallenged, were killed and functional antitumor activity and phenotype characteristics of various lymphocyte populations were determined in comparison to those of young and age-matched control mice. The phenotyping of the lymphocytes from the cured mice indicated very notable differences in subset distribution and increased CD44 expression. Functionally they developed high levels of anti-EL4 activity, which was ablated by combined treatment with monoclonal antibodies against CD8 and CD44, indicating the role of memory cells. Consistent with cells from aged mice, these same cell populations had a very reduced allogeneic responsiveness. It appears that cured mice have developed an immune memory specific for EL4.

Recipient micro-environment does not dictate the Igh-V restriction specificity of T cell suppressor inducer factor (TsiF) from allogeneic bone marrow chimera in mice

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Noguchi, M.; Ogasawara, M.; Iwabuchi, K.; Osgasawara, K.; Ishihara, T.; Good, R.A.; Morikawa, K.; Onoe, K.

1985-01-01

The authors have ascertained previously from a study of fully allogeneic irradiation chimeras in mice that the H-2 restriction of the suppressor factor (Ly-2 T suppressor factor) is determined by the post-thymic environment protected by the donor cells, rather than by the thymic environment of the recipient. In the present study, the author analyzed differentiation influences that determine the Igh restriction specificities of the suppressor inducer T cell factor(s) (TsiF) that are produced by Ly-1+ splenic T cells in fully allogeneic bone marrow chimeras in mice. AKR mice that had been lethally irradiated and reconstituted with B10 marrow cells, [B10----AKR] chimeras, produced Ly-1 TsiF after hyper-immunization with sheep erythrocytes (SRBC) which suppressed antigen--specifically the primary antibody responses to SRBC that were generated in cells of the same Igh-Vb haplotype of donor strain and not those generated in cells of the recipient Igh-Va type. Similar results were obtained when Ly-1 TsiF from [B6----BALB/c] and [BALB/c----B6] chimeras were analyzed. Furthermore, the Ly-1 TsiF from [BALB/c----B6] chimeras suppressed the primary antibody responses of both BALB/c [H-2d, Igh-Va, Igh-Ca] and BAB-14 (H-2d, Igh-Va, Igh-Cb), but not those of CAL-20 (H-2d, Igh-Vd, Igh-Cd). These results demonstrate clearly that the Ly-1 TsiF from allogeneic bone marrow chimeras are donor Igh-V-restricted and are not influenced by the recipient micro-environment, presumably that were provided by the thymuses of the recipient mice

Use of superoxide dismutase and catalase producing lactic acid bacteria in TNBS induced Crohn's disease in mice.

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LeBlanc, Jean Guy; del Carmen, Silvina; Miyoshi, Anderson; Azevedo, Vasco; Sesma, Fernando; Langella, Philippe; Bermúdez-Humarán, Luis G; Watterlot, Laurie; Perdigon, Gabriela; de Moreno de LeBlanc, Alejandra

2011-02-10

Reactive oxygen species are involved in various aspects of intestinal inflammation and tumor development. Decreasing their levels using antioxidant enzymes, such as catalase (CAT) or superoxide dismutase (SOD) could therefore be useful in the prevention of certain diseases. Lactic acid bacteria (LAB) are ideal candidates to deliver these enzymes in the gut. In this study, the anti-inflammatory effects of CAT or SOD producing LAB were evaluated using a trinitrobenzenesulfonic acid (TNBS) induced Crohn's disease murine model. Engineered Lactobacillus casei BL23 strains producing either CAT or SOD, or the native strain were given to mice before and after intrarectal administration of TNBS. Animal survival, live weight, intestinal morphology and histology, enzymatic activities, microbial translocation to the liver and cytokines released in the intestinal fluid were evaluated. The mice that received CAT or SOD-producing LAB showed a faster recovery of initial weight loss, increased enzymatic activities in the gut and lesser extent of intestinal inflammation compared to animals that received the wild-type strain or those that did not receive bacterial supplementation. Our findings suggest that genetically engineered LAB that produce antioxidant enzymes could be used to prevent or decrease the severity of certain intestinal pathologies. Copyright © 2010 Elsevier B.V. All rights reserved.

Brain-specific Crmp2 deletion leads to neuronal development deficits and behavioural impairments in mice.

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Zhang, Hongsheng; Kang, Eunchai; Wang, Yaqing; Yang, Chaojuan; Yu, Hui; Wang, Qin; Chen, Zheyu; Zhang, Chen; Christian, Kimberly M; Song, Hongjun; Ming, Guo-Li; Xu, Zhiheng

2016-06-01

Several genome- and proteome-wide studies have associated transcription and translation changes of CRMP2 (collapsing response mediator protein 2) with psychiatric disorders, yet little is known about its function in the developing or adult mammalian brain in vivo. Here we show that brain-specific Crmp2 knockout (cKO) mice display molecular, cellular, structural and behavioural deficits, many of which are reminiscent of neural features and symptoms associated with schizophrenia. cKO mice exhibit enlarged ventricles and impaired social behaviour, locomotor activity, and learning and memory. Loss of Crmp2 in the hippocampus leads to reduced long-term potentiation, abnormal NMDA receptor composition, aberrant dendrite development and defective synapse formation in CA1 neurons. Furthermore, knockdown of crmp2 specifically in newborn neurons results in stage-dependent defects in their development during adult hippocampal neurogenesis. Our findings reveal a critical role for CRMP2 in neuronal plasticity, neural function and behavioural modulation in mice.

Hormesis of specific IgG antibody to rabies virus in serum of mice irradiated with low dose γ-rays

International Nuclear Information System (INIS)

Liu Qingjie; Chen Deqing

1998-01-01

Objective: To explore the effect of low dose ionizing radiation on specific antibody in mouse serum. Methods: Kunming strain male mice, weighing 18-22 g, aged 6-8 weeks, were immunized intraperitoneally with rabies vaccine after exposure to cobalt-60 γ-rays. The specific IgG antibody against rabies virus in mouse serum was measured. Results: (1) The serum levels of specific IgG in mice irradiated with 5-30 cGy γ-rays were significantly elevated in comparison with those in control mice (P<0.01), the optimum stimulating dose being 10 cGy. (2) Exposure to 10 cGy caused significant enhancement and earlier emergence of the peak level of specific IgG in serum. (3) The hormesis of specific IgG to rabies virus induced by 10 cGy γ-rays could last one week at least. Conclusion: Low dose ionizing radiation can enhance the level of specific antibody in mouse serum, and this effect can last for one week at least

Human Polyclonal Antibodies Produced through DNA Vaccination of Transchromosomal Cattle Provide Mice with Post-Exposure Protection against Lethal Zaire and Sudan Ebolaviruses.

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Callie E Bounds

Full Text Available DNA vaccination of transchromosomal bovines (TcBs with DNA vaccines expressing the codon-optimized (co glycoprotein (GP genes of Ebola virus (EBOV and Sudan virus (SUDV produce fully human polyclonal antibodies (pAbs that recognize both viruses and demonstrate robust neutralizing activity. Each TcB was vaccinated by intramuscular electroporation (IM-EP a total of four times and at each administration received 10 mg of the EBOV-GPco DNA vaccine and 10 mg of the SUDV-GPco DNA vaccine at two sites on the left and right sides, respectively. After two vaccinations, robust antibody responses (titers > 1000 were detected by ELISA against whole irradiated EBOV or SUDV and recombinant EBOV-GP or SUDV-GP (rGP antigens, with higher titers observed for the rGP antigens. Strong, virus neutralizing antibody responses (titers >1000 were detected after three vaccinations when measured by vesicular stomatitis virus-based pseudovirion neutralization assay (PsVNA. Maximal neutralizing antibody responses were identified by traditional plaque reduction neutralization tests (PRNT after four vaccinations. Neutralizing activity of human immunoglobulins (IgG purified from TcB plasma collected after three vaccinations and injected intraperitoneally (IP into mice at a 100 mg/kg dose was detected in the serum by PsVNA up to 14 days after administration. Passive transfer by IP injection of the purified IgG (100 mg/kg to groups of BALB/c mice one day after IP challenge with mouse adapted (ma EBOV resulted in 80% protection while all mice treated with non-specific pAbs succumbed. Similarly, interferon receptor 1 knockout (IFNAR(-/- mice receiving the purified IgG (100 mg/kg by IP injection one day after IP challenge with wild type SUDV resulted in 89% survival. These results are the first to demonstrate that filovirus GP DNA vaccines administered to TcBs by IM-EP can elicit neutralizing antibodies that provide post-exposure protection. Additionally, these data describe

Cushing's syndrome and fetal features resurgence in adrenal cortex-specific Prkar1a knockout mice.

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Isabelle Sahut-Barnola

2010-06-01

Full Text Available Carney complex (CNC is an inherited neoplasia syndrome with endocrine overactivity. Its most frequent endocrine manifestation is primary pigmented nodular adrenocortical disease (PPNAD, a bilateral adrenocortical hyperplasia causing pituitary-independent Cushing's syndrome. Inactivating mutations in PRKAR1A, a gene encoding the type 1 alpha-regulatory subunit (R1alpha of the cAMP-dependent protein kinase (PKA have been found in 80% of CNC patients with Cushing's syndrome. To demonstrate the implication of R1alpha loss in the initiation and development of PPNAD, we generated mice lacking Prkar1a specifically in the adrenal cortex (AdKO. AdKO mice develop pituitary-independent Cushing's syndrome with increased PKA activity. This leads to autonomous steroidogenic genes expression and deregulated adreno-cortical cells differentiation, increased proliferation and resistance to apoptosis. Unexpectedly, R1alpha loss results in improper maintenance and centrifugal expansion of cortisol-producing fetal adrenocortical cells with concomitant regression of adult cortex. Our data provide the first in vivo evidence that loss of R1alpha is sufficient to induce autonomous adrenal hyper-activity and bilateral hyperplasia, both observed in human PPNAD. Furthermore, this model demonstrates that deregulated PKA activity favors the emergence of a new cell population potentially arising from the fetal adrenal, giving new insight into the mechanisms leading to PPNAD.

Disturbed α-Cell Function in Mice with β-Cell Specific Overexpression of Human Islet Amyloid Polypeptide

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Bo Ahrén

2008-01-01

Full Text Available Exogenous administration of islet amyloid polypeptide (IAPP has been shown to inhibit both insulin and glucagon secretion. This study examined α-cell function in mice with β-cell specific overexpression of human IAPP (hIAPP after an oral protein gavage (75 mg whey protein/mouse. Baseline glucagon levels were higher in transgenic mice (41±4.0 pg/mL, n=6 than in wildtype animals (19±5.1 pg/mL, n=5, P=.015. In contrast, the glucagon response to protein was impaired in transgenic animals (21±2.7 pg/mL in transgenic mice versus 38±5.7 pg/mL in wildtype mice at 15 minutes; P=.027. Baseline insulin levels did not differ between the groups, while the insulin response, as the glucagon response, was impaired after protein challenge (P=.018. Glucose levels were not different between the groups and did not change significantly after protein gavage. Acetaminophen was given through gavage to the animals (2 mg/mouse to estimate gastric emptying. The plasma acetaminophen profile was similar in the two groups of mice. We conclude that disturbances in glucagon secretion exist in mice with β-cell specific overexpression of human IAPP, which are not secondary to changes in gastric emptying. The reduced glucagon response to protein challenge may reflect a direct inhibitory influence of hIAPP on glucagon secretion.

Prenatal exposure to phencyclidine produces abnormal behaviour and NMDA receptor expression in postpubertal mice.

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Lu, Lingling; Mamiya, Takayoshi; Lu, Ping; Toriumi, Kazuya; Mouri, Akihiro; Hiramatsu, Masayuki; Kim, Hyoung-Chun; Zou, Li-Bo; Nagai, Taku; Nabeshima, Toshitaka

2010-08-01

Several studies have shown the disruptive effects of non-competitive N-methyl-d-aspartate (NMDA) receptor antagonists on neurobehavioural development. Based on the neurodevelopment hypothesis of schizophrenia, there is growing interest in animal models treated with NMDA antagonists at developing stages to investigate the pathogenesis of psychological disturbances in humans. Previous studies have reported that perinatal treatment with phencyclidine (PCP) impairs the development of neuronal systems and induces schizophrenia-like behaviour. However, the adverse effects of prenatal exposure to PCP on behaviour and the function of NMDA receptors are not well understood. This study investigated the long-term effects of prenatal exposure to PCP in mice. The prenatal PCP-treated mice showed hypersensitivity to a low dose of PCP in locomotor activity and impairment of recognition memory in the novel object recognition test at age 7 wk. Meanwhile, the prenatal exposure reduced the phosphorylation of NR1, although it increased the expression of NR1 itself. Furthermore, these behavioural changes were attenuated by atypical antipsychotic treatment. Taken together, prenatal exposure to PCP produced long-lasting behavioural deficits, accompanied by the abnormal expression and dysfunction of NMDA receptors in postpubertal mice. It is worth investigating the influences of disrupted NMDA receptors during the prenatal period on behaviour in later life.

Diet-induced obesity alters protein synthesis: Tissue-specific effects in fasted vs. fed mice

OpenAIRE

Anderson, Stephanie R.; Gilge, Danielle A.; Steiber, Alison L.; Previs, Stephen F.

2008-01-01

The influence of obesity on protein dynamics is not clearly understood. We have designed experiments to test the hypothesis that obesity impairs the stimulation of tissue-specific protein synthesis following nutrient ingestion. C57BL/6J mice were randomized into two groups: group 1 (control, n = 16) were fed a low-fat, high-carbohydrate diet and group 2 (experimental, n = 16) were fed a high-fat, low-carbohydrate diet ad libitum for 9 weeks. On the experiment day, all mice were fasted for 6 h...

Restoration of prostaglandin E2-producing splenic macrophages in 89Sr-treated mice with bone marrow from Corynebacterium parvum primed donors

International Nuclear Information System (INIS)

Shibata, Y.

1989-01-01

Administration of Corynebacterium parvum (CP), 56 mg/kg ip to CBA/J mice effected the induction of prostaglandin E2 (PGE2) producing macrophages (M phi) in the bone marrow and the spleen. Maximal release of PGE2 from M phi cultured in vitro with calcium ionophore A23187 for 2 h was reached by marrow M phi removed on 5 days after CP (450 ng/mg cell protein), and by splenic M phi 9 days after CP (400 ng/mg). Neither M phi population, however, yielded more than 6.0 ng/mg leukotriene C4. To assess ontogenic relationships mice were depleted of bone marrow and blood monocytes by iv injection of the bone-seeking isotope, 89Sr. CP was given at several points before or after bone marrow cell depletion. PGE2 production by splenic M phi harvested on day 9 after CP was profoundly impaired when CP was administered either concurrently with or 3 days after 89Sr. When CP was administered 1, 3, 5, and 7 days before 89Sr, however, the induction of PGE2-producing M phi in the spleen was unaffected. To determine whether bone marrow cells from CP-injected donors can restore PGE2-producing splenic M phi (PGSM) in 89Sr-mice, recipient mice which had and had not received CP 3 days after 89Sr were transfused with 5 x 10(6) syngeneic bone marrow cells from donor mice prepared at varying intervals after CP administration. The results clearly indicate the capacity of bone marrow cells harvested on either day 1 or 2 following CP to restore PGSM in CP-primed, but not unprimed, recipients

Parental resistance of irradiated mice to (CBAXM523)F/sub 1/ lymphocytes: the destiny of transplanted cells, duration of resistance and its specificity

Energy Technology Data Exchange (ETDEWEB)

Kondrat' eva, T K; Fontalin, L N; Nagurskaya, E V; Novikova, T K; Blandova, Z K; Chernousov, A D [Akademiya Meditsinskikh Nauk SSSR, Moscow. Inst. Ehpidemiologii i Mikrobiologii

1979-05-01

The immunologic reactivity of mouse (CBAxM523)F/sub 1/ lymphocytes to alien antigens (ram erythrocytes) in the organisms of lethally irradiated CBA mice was studied. If the irradiation, cell transfer and antigen test-injection were performed on the same day, the activity of the transplant was suppressed as compared to the syngenic system. If the intervals between these procedures increased to three days the activity of donor cells was recovered. The retransplantation of recipient spleen cells to the irradiated CBA and F/sub 1/ mice demonstrated the viability of the transplanted cells and the absence of their transadaptation to nonsyngenic microenvironment. The resistance of the recipients could be overcome specifically by preliminary injection of F/sub 1/ mouse cells in combination with cyclophosphamide or without it. The results obtained suggest to conclude, that the genetic parental resistance of CBA mice to F/sub 1/ mouse cells is caused by the immunologically competent recipient cells that are inactivated after three days following irradiation. They do not produce a cytotoxic effect on donor cells, but limit temporarily the activity of the latter.

Sepsis reveals compartment-specific responses in intestinal proliferation and apoptosis in transgenic mice whose enterocytes re-enter the cell cycle.

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Lyons, John D; Klingensmith, Nathan J; Otani, Shunsuke; Mittal, Rohit; Liang, Zhe; Ford, Mandy L; Coopersmith, Craig M

2017-12-01

Cell production and death are tightly regulated in the rapidly renewing gut epithelium, with proliferation confined to crypts and apoptosis occurring in villi and crypts. This study sought to determine how stress alters these compartmentalized processes. Wild-type mice made septic via cecal ligation and puncture had decreased crypt proliferation and increased crypt and villus apoptosis. Fabpi -TAg mice expressing large T-antigen solely in villi had ectopic enterocyte proliferation with increased villus apoptosis in unmanipulated animals. Septic fabpi -TAg mice had an unexpected increase in villus proliferation compared with unmanipulated littermates, whereas crypt proliferation was decreased. Cell cycle regulators cyclin D1 and cyclin D2 were decreased in jejunal tissue in septic transgenic mice. In contrast, villus and crypt apoptosis were increased in septic fabpi -TAg mice. To examine the relationship between apoptosis and proliferation in a compartment-specific manner, fabpi -TAg mice were crossed with fabpl -Bcl-2 mice, resulting in expression of both genes in the villus but Bcl-2 alone in the crypt. Septic bi-transgenic animals had decreased crypt apoptosis but had a paradoxical increase in villus apoptosis compared with septic fabpi -TAg mice, associated with decreased proliferation in both compartments. Thus, sepsis unmasks compartment-specific proliferative and apoptotic regulation that is not present under homeostatic conditions.-Lyons, J. D., Klingensmith, N. J., Otani, S., Mittal, R., Liang, Z., Ford, M. L., Coopersmith, C. M. Sepsis reveals compartment-specific responses in intestinal proliferation and apoptosis in transgenic mice whose enterocytes re-enter the cell cycle. © FASEB.

Immobilization induced osteopenia is strain specific in mice

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Andreas Lodberg

2015-06-01

Full Text Available Immobilization causes rapid and massive bone loss. By comparing Botulinum Toxin A (BTX-induced bone loss in mouse strains with different genetic backgrounds we investigated whether the genetic background had an influence on the severity of the osteopenia. Secondly, we investigated whether BTX had systemic effects on bone. Female mice from four inbred mouse strains (BALB/cJ, C57BL/6 J, DBA/2 J, and C3H/HeN were injected unilaterally with BTX (n = 10/group or unilaterally with saline (n = 10/group. Mice were euthanized after 21 days, and the bone properties evaluated using μCT, DXA, bone histomorphometry, and mechanical testing. BTX resulted in substantially lower trabecular bone volume fraction (BV/TV and trabecular thickness in all mouse strains. The deterioration of BV/TV was significantly greater in C57BL/6 J (−57% and DBA/2 J (−60% than in BALB/cJ (−45% and C3H/HeN (−34% mice. The loss of femoral neck fracture strength was significantly greater in C57BL/6 J (−47% and DBA/2 J (−45% than in C3H (−25% mice and likewise the loss of mid-femoral fracture strength was greater in C57BL/6 J (−17%, DBA/2 J (−12%, and BALB/cJ (−9% than in C3H/HeN (−1% mice, which were unaffected. Using high resolution μCT we found no evidence of a systemic effect on any of the microstructural parameters of the contralateral limb. Likewise, there was no evidence of a systemic effect on the bone strength in any mouse strain. We did, however, find a small systemic effect on aBMD in DBA/2 J and C3H/HeN mice. The present study shows that BTX-induced immobilization causes the greatest loss of cortical and trabecular bone in C57BL/6 J and DBA/2 J mice. A smaller loss of bone microstructure and fracture strength was seen in BALB/cJ mice, while the bone microstructure and fracture strength of C3H/HeN mice were markedly less affected. This indicates that BTX-induced loss of bone is mouse strain dependent. We found only minimal systemic

Evaluation of organ-specific glucose metabolism by 18F-FDG in insulin receptor substrate-1 (IRS-1) knockout mice as a model of insulin resistance

International Nuclear Information System (INIS)

Cheng, Chao; Nakamura, Akinobu; Minamimoto, Ryogo; Shinoda, Kazuaki; Tateishi, Ukihide; Terauchi, Yasuo; Inoue, Tomio; Goto, Atsuhi; Kadowaki, Takashi

2011-01-01

Insulin resistance (IR) is a physiological condition in which the body produces insulin but does not result in a sufficient biological effect. Insulin resistance is usually asymptomatic but is associated with health problems and is a factor in the metabolic syndrome. The aim of the present study is to clarify organ-specific insulin resistance in normal daily conditions using [ 18 F]-2-fluoro-2-deoxy-D-glucose ([ 18 F]-FDG). The biodistribution of [ 18 F]-FDG was examined in insulin receptor substrate-1 (IRS-1) knockout mice, an animal model of skeletal muscle insulin resistance, and C57BL/6J (wild-type) mice with and without insulin loading. Mice received 0.5 MBq of [ 18 F]-FDG injected into the tail vein, immediately followed by nothing (control cohorts) or an intraperitoneal injection of 1.5 mU/g body weight of human insulin as an insulin loading test. Blood glucose concentrations for all of the experimental animals were assessed at 0, 20, 40, and 60 min post-injection. The mice were subsequently killed, and tissue was collected for evaluation of [ 18 F]-FDG biodistribution. The radioactivity of each organ was measured using a gamma counter. In the absence of insulin, the blood glucose concentrations of wild-type mice (132±26 mg/dl) and IRS-1 knockout mice (134±18 mg/dl) were not significantly different. Blood glucose concentrations decreased following insulin administration, with lower concentrations in wild-type mice than in knockout mice at 20, 40, and 60 min. A statistically significant difference in [ 18 F]-FDG uptake between wild-type mice and IRS-1 knockout mice was confirmed in the heart, abdominal muscle, and femoral muscle. With insulin loading, [ 18 F]-FDG uptake in the heart, back muscle, and abdominal muscle was significantly increased compared to without insulin loading in both wild-type mice and knockout mice. Our results showed that IR significantly affected [ 18 F]-FDG uptake in the heart in normal daily conditions. IR was associated with

Impaired intervertebral disc development and premature disc degeneration in mice with notochord-specific deletion of CCN2.

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Bedore, Jake; Sha, Wei; McCann, Matthew R; Liu, Shangxi; Leask, Andrew; Séguin, Cheryle A

2013-10-01

Currently, our ability to treat intervertebral disc (IVD) degeneration is hampered by an incomplete understanding of disc development and aging. The specific function of matricellular proteins, including CCN2, during these processes remains an enigma. The aim of this study was to determine the tissue-specific localization of CCN proteins and to characterize their role in IVD tissues during embryonic development and age-related degeneration by using a mouse model of notochord-specific CCN2 deletion. Expression of CCN proteins was assessed in IVD tissues from wild-type mice beginning on embryonic day 15.5 to 17 months of age. Given the enrichment of CCN2 in notochord-derived tissues, we generated notochord-specific CCN2-null mice to assess the impact on the IVD structure and extracellular matrix composition. Using a combination of histologic evaluation and magnetic resonance imaging (MRI), IVD health was assessed. Loss of the CCN2 gene in notochord-derived cells disrupted the formation of IVDs in embryonic and newborn mice, resulting in decreased levels of aggrecan and type II collagen and concomitantly increased levels of type I collagen within the nucleus pulposus. CCN2-knockout mice also had altered expression of CCN1 (Cyr61) and CCN3 (Nov). Mirroring its role during early development, notochord-specific CCN2 deletion accelerated age-associated degeneration of IVDs. Using a notochord-specific gene targeting strategy, this study demonstrates that CCN2 expression by nucleus pulposus cells is essential to the regulation of IVD development and age-associated tissue maintenance. The ability of CCN2 to regulate the composition of the intervertebral disc suggests that it may represent an intriguing clinical target for the treatment of disc degeneration. Copyright © 2013 by the American College of Rheumatology.

Corporate Regional Responsibility - Warum engagieren sich Unternehmen gemeinsam für ihre Region?. Motive der kollektiven regionalen Verantwortungsübernahme von Unternehmen an den Beispielen des Initiativkreises Ruhr und der Wirtschaftsinitiative FrankfurtRheinMain

Science.gov (United States)

Schiek, Meike

2016-03-01

Companies can voluntarily participate in matters of regional developments, thereby accepting responsibility on a regional level. Referring to the concept of Corporate Social Responsibility (CSR), the term Corporate Regional Responsibility (CRR) is used to describe this behavior. Moreover, companies can form a CRR-corporation with other companies in order to take over a collective CRR. So far, the motives of companies for exercising collective CRR are unknown, thus, corporate resources can not be mobilized and utilized efficiently for regional developments. This article explores the subject of collective CRR and illustrates CRR motives using the example of the two CRR-cooperations Initiativkreis Ruhr and Wirtschaftsinitiative FrankfurtRheinMain.

Peripheral erythrocytes decrease upon specific respiratory challenge with grass pollen allergen in sensitized mice and in human subjects.

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Galateja Jordakieva

Full Text Available BACKGROUND AND AIMS: Specific hyper-responsiveness towards an allergen and non-specific airway hyperreactivity both impair quality of life in patients with respiratory allergic diseases. We aimed to investigate cellular responses following specific and non-specific airway challenges locally and systemically in i sensitized BALB/c mice challenged with grass pollen allergen Phl p 5, and in ii grass pollen sensitized allergic rhinitis subjects undergoing specific airway challenge in the Vienna Challenge Chamber (VCC. METHODS AND RESULTS: BALB/c mice (n = 20 were intraperitoneally immunized with grass pollen allergen Phl p 5 and afterwards aerosol challenged with either the specific allergen Phl p 5 (n = 10 or the non-specific antigen ovalbumin (OVA (n = 10. A protocol for inducing allergic asthma as well as allergic rhinitis, according to the united airway concept, was used. Both groups of exposed mice showed significantly reduced physical activity after airway challenge. Specific airway challenge further resulted in goblet cell hyperplasia, enhanced mucous secretion, intrapulmonary leukocyte infiltration and lymphoid follicle formation, associated with significant expression of IL-4, IL-5 and IL-13 in splenocytes and also partially in lung tissue. Concerning circulating blood cell dynamics, we observed a significant drop of erythrocyte counts, hemoglobin and hematocrit levels in both mouse groups, challenged with allergen or OVA. A significant decrease in circulating erythrocytes and hematocrit levels after airway challenges with grass pollen allergen was also found in grass pollen sensitized human rhinitis subjects (n = 42 at the VCC. The effects on peripheral leukocyte counts in mice and humans however were opposed, possibly due to the different primary inflammation sites. CONCLUSION: Our data revealed that, besides significant leukocyte dynamics, particularly erythrocytes are involved in acute hypersensitivity reactions to respiratory allergens

Hepcidin is elevated in mice injected with Mycoplasma arthritidis

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Kaplan Jerry

2009-11-01

Full Text Available Abstract Mycoplasma arthritidis causes arthritis in specific mouse strains. M. arthritidis mitogen (MAM, a superantigen produced by M. arthritidis, activates T cells by forming a complex between the major histocompatability complex II on antigen presenting cells and the T cell receptor on CD4+ T lymphocytes. The MAM superantigen is also known to interact with Toll-like receptors (TLR 2 and 4. Hepcidin, an iron regulator protein, is upregulated by TLR4, IL-6, and IL-1. In this study, we evaluated serum hepcidin, transferrin saturation, ferritin, IL-6, IL-1, and hemoglobin levels in M. arthritidis injected C3H/HeJ (TLR2+/+, TLR4-/- mice and C3H/HeSnJ (TLR2+/+, TLR4+/+ mice over a 21 day period. C3H/HeJ mice have a defective TLR4 and an inability to produce IL-6. We also measured arthritis severity in these mice and the amount of hepcidin transcripts produced by the liver and spleen. C3H/HeJ mice developed a more severe arthritis than that of C3H/HeSnJ mice. Both mice had an increase in serum hepcidin within three days after infection. Hepcidin levels were greater in C3H/HeJ mice despite a nonfunctioning TLR4 and low serum levels of IL-6. Splenic hepcidin production in C3H/HeJ mice was delayed compared to C3H/HeSnJ mice. Unlike C3H/HeSnJ mice, C3H/HeJ mice did not develop a significant rise in serum IL-6 levels but did develop a significant increase in IL-1β during the first ten days after injection. Both mice had an increase in serum ferritin but a decrease in serum transferrin saturation. In conclusion, serum hepcidin regulation in C3H/HeJ mice does not appear to be solely dependent upon TLR4 or IL-6.

Identification of NY-BR-1-specific CD4(+) T cell epitopes using HLA-transgenic mice.

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Gardyan, Adriane; Osen, Wolfram; Zörnig, Inka; Podola, Lilli; Agarwal, Maria; Aulmann, Sebastian; Ruggiero, Eliana; Schmidt, Manfred; Halama, Niels; Leuchs, Barbara; von Kalle, Christof; Beckhove, Philipp; Schneeweiss, Andreas; Jäger, Dirk; Eichmüller, Stefan B

2015-06-01

Breast cancer represents the second most common cancer type worldwide and has remained the leading cause of cancer-related deaths among women. The differentiation antigen NY-BR-1 appears overexpressed in invasive mammary carcinomas compared to healthy breast tissue, thus representing a promising target antigen for T cell based tumor immunotherapy approaches. Since efficient immune attack of tumors depends on the activity of tumor antigen-specific CD4(+) effector T cells, NY-BR-1 was screened for the presence of HLA-restricted CD4(+) T cell epitopes that could be included in immunological treatment approaches. Upon NY-BR-1-specific DNA immunization of HLA-transgenic mice and functional ex vivo analysis, a panel of NY-BR-1-derived library peptides was determined that specifically stimulated IFNγ secretion among splenocytes of immunized mice. Following in silico analyses, four candidate epitopes were determined which were successfully used for peptide immunization to establish NY-BR-1-specific, HLA-DRB1*0301- or HLA-DRB1*0401-restricted CD4(+) T cell lines from splenocytes of peptide immunized HLA-transgenic mice. Notably, all four CD4(+) T cell lines recognized human HLA-DR-matched dendritic cells (DC) pulsed with lysates of NY-BR-1 expressing human tumor cells, demonstrating natural processing of these epitopes also within the human system. Finally, CD4(+) T cells specific for all four CD4(+) T cell epitopes were detectable among PBMC of breast cancer patients, showing that CD4(+) T cell responses against the new epitopes are not deleted nor inactivated by self-tolerance mechanisms. Our results present the first NY-BR-1-specific HLA-DRB1*0301- and HLA-DRB1*0401-restricted T cell epitopes that could be exploited for therapeutic intervention against breast cancer. © 2014 UICC.

Probiotic Bifidobacterium breve induces IL-10-producing Tr1 cells in the colon.

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Seong Gyu Jeon

Full Text Available Specific intestinal microbiota has been shown to induce Foxp3(+ regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103(+ dendritic cells (DCs mediated B. breve-induced development of IL-10-producing T cells. CD103(+ DCs from Il10(-/-, Tlr2(-/-, and Myd88(-/- mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103(+ DCs failed to induce IL-10 production from co-cultured Il27ra(-/- T cells. B. breve treatment of Tlr2(-/- mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103(+ DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4(+ T cells from wild-type mice, but not Il10(-/- mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.

Probiotic Bifidobacterium breve induces IL-10-producing Tr1 cells in the colon.

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Jeon, Seong Gyu; Kayama, Hisako; Ueda, Yoshiyasu; Takahashi, Takuya; Asahara, Takashi; Tsuji, Hirokazu; Tsuji, Noriko M; Kiyono, Hiroshi; Ma, Ji Su; Kusu, Takashi; Okumura, Ryu; Hara, Hiromitsu; Yoshida, Hiroki; Yamamoto, Masahiro; Nomoto, Koji; Takeda, Kiyoshi

2012-01-01

Specific intestinal microbiota has been shown to induce Foxp3(+) regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103(+) dendritic cells (DCs) mediated B. breve-induced development of IL-10-producing T cells. CD103(+) DCs from Il10(-/-), Tlr2(-/-), and Myd88(-/-) mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103(+) DCs failed to induce IL-10 production from co-cultured Il27ra(-/-) T cells. B. breve treatment of Tlr2(-/-) mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103(+) DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4(+) T cells from wild-type mice, but not Il10(-/-) mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.

Characterization of anti-P monoclonal antibodies directed against the ribosomal protein-RNA complex antigen and produced using Murphy Roths large autoimmune-prone mice.

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Sato, H; Onozuka, M; Hagiya, A; Hoshino, S; Narita, I; Uchiumi, T

2015-02-01

Autoantibodies, including anti-ribosomal P proteins (anti-P), are thought to be produced by an antigen-driven immune response in systemic lupus erythematosus (SLE). To test this hypothesis, we reconstituted the ribosomal antigenic complex in vitro using human P0, phosphorylated P1 and P2 and a 28S rRNA fragment covering the P0 binding site, and immunized Murphy Roths large (MRL)/lrp lupus mice with this complex without any added adjuvant to generate anti-P antibodies. Using hybridoma technology, we subsequently obtained 34 clones, each producing an anti-P monoclonal antibody (mAb) that recognized the conserved C-terminal tail sequence common to all three P proteins. We also obtained two P0-specific monoclonal antibodies, but no antibody specific to P1, P2 or rRNA fragment. Two types of mAbs were found among these anti-P antibodies: one type (e.g. 9D5) reacted more strongly with the phosphorylated P1 and P2 than that with their non-phosphorylated forms, whereas the other type (e.g. 4H11) reacted equally with both phosphorylated and non-phosphorylated forms of P1/P2. Both 9D5 and 4H11 inhibited the ribosome/eukaryotic elongation factor-2 (eEF-2)-coupled guanosine triphosphate (GTP)ase activity. However, preincubation with a synthetic peptide corresponding to the C-terminal sequence common to all three P proteins, but not the peptide that lacked the last three C-terminal amino acids, mostly prevented the mAb-induced inhibition of GTPase activity. Thus, at least two types of anti-P were produced preferentially following the immunization of MRL mice with the reconstituted antigenic complex. Presence of multiple copies of the C-termini, particularly that of the last three C-terminal amino acid residues, in the antigenic complex appears to contribute to the immunogenic stimulus. © 2014 British Society for Immunology.

Characterization of anti-P monoclonal antibodies directed against the ribosomal protein–RNA complex antigen and produced using Murphy Roths large autoimmune-prone mice

Science.gov (United States)

Sato, H; Onozuka, M; Hagiya, A; Hoshino, S; Narita, I; Uchiumi, T

2015-01-01

Autoantibodies, including anti-ribosomal P proteins (anti-P), are thought to be produced by an antigen-driven immune response in systemic lupus erythematosus (SLE). To test this hypothesis, we reconstituted the ribosomal antigenic complex in vitro using human P0, phosphorylated P1 and P2 and a 28S rRNA fragment covering the P0 binding site, and immunized Murphy Roths large (MRL)/lrp lupus mice with this complex without any added adjuvant to generate anti-P antibodies. Using hybridoma technology, we subsequently obtained 34 clones, each producing an anti-P monoclonal antibody (mAb) that recognized the conserved C-terminal tail sequence common to all three P proteins. We also obtained two P0-specific monoclonal antibodies, but no antibody specific to P1, P2 or rRNA fragment. Two types of mAbs were found among these anti-P antibodies: one type (e.g. 9D5) reacted more strongly with the phosphorylated P1 and P2 than that with their non-phosphorylated forms, whereas the other type (e.g. 4H11) reacted equally with both phosphorylated and non-phosphorylated forms of P1/P2. Both 9D5 and 4H11 inhibited the ribosome/eukaryotic elongation factor-2 (eEF-2)-coupled guanosine triphosphate (GTP)ase activity. However, preincubation with a synthetic peptide corresponding to the C-terminal sequence common to all three P proteins, but not the peptide that lacked the last three C-terminal amino acids, mostly prevented the mAb-induced inhibition of GTPase activity. Thus, at least two types of anti-P were produced preferentially following the immunization of MRL mice with the reconstituted antigenic complex. Presence of multiple copies of the C-termini, particularly that of the last three C-terminal amino acid residues, in the antigenic complex appears to contribute to the immunogenic stimulus. PMID:25255895

Sex-specific mechanism of social hierarchy in mice.

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van den Berg, Wouter E; Lamballais, Sander; Kushner, Steven A

2015-05-01

The establishment of social hierarchies is a naturally occurring, evolutionarily conserved phenomenon with a well-established impact on fitness and health. Investigations of complex social group dynamics may offer novel opportunities for translational studies of autism spectrum disorder. Here we describe a robust behavioral paradigm using an automated version of the tube test. Isogenic groups of male and female mice establish linear social hierarchies that remain highly stable for at least 14 days, the longest interval tested. Remarkably, however, their social strategy is sex-specific: females primarily utilize intrinsic attributes, whereas males are strongly influenced by prior social experience. Using both genetic and pharmacological manipulations, we identify testosterone as a critical sex-specific factor for determining which social strategy is used. Males inheriting a null mutation of the sex-determining region Y (Sry) gene used a similar social cognitive strategy as females. In contrast, females with transgenic expression of Sry utilized a typically male social strategy. Analogously, castration of males and testosterone supplementation of females yielded similar outcomes, with a reversal of their social cognitive strategy. Together, our results demonstrate a sex-specific mechanism underlying social hierarchy, in which both males and females retain the functional capacity to adapt their social strategy. More generally, we expect the automated tube test to provide an important complementary approach for both fundamental and translational studies of social behavior.

Acid solution is a suitable medium for introducing QX-314 into nociceptors through TRPV1 channels to produce sensory-specific analgesic effects.

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He Liu

Full Text Available BACKGROUND: Previous studies have demonstrated that QX-314, an intracellular sodium channel blocker, can enter into nociceptors through capsaicin-activated TRPV1 or permeation of the membrane by chemical enhancers to produce a sensory-selective blockade. However, the obvious side effects of these combinations limit the application of QX-314. A new strategy for targeting delivery of QX-314 into nociceptors needs further investigation. The aim of this study is to test whether acidic QX-314, when dissolves in acidic solution directly, can enter into nociceptors through acid-activated TRPV1 and block sodium channels from the intracellular side to produce a sensory-specific analgesic effect. METHODOLOGY/PRINCIPAL FINDINGS: Acidic solution or noradrenaline was injected intraplantarly to induce acute pain behavior in mice. A chronic constrictive injury model was performed to induce chronic neuropathic pain. A sciatic nerve blockade model was used to evaluate the sensory-specific analgesic effects of acidic QX-314. Thermal and mechanical hyperalgesia were measured by using radiant heat and electronic von Frey filaments test. Spinal Fos protein expression was determined by immunohistochemistry. The expression of p-ERK was detected by western blot assay. Whole cell clamp recording was performed to measure action potentials and total sodium current in rats DRG neurons. We found that pH 5.0 PBS solution induced behavioral hyperalgesia accompanied with the increased expression of spinal Fos protein and p-ERK. Pretreatment with pH 5.0 QX-314, and not pH 7.4 QX-314, alleviated pain behavior, inhibited the increased spinal Fos protein and p-ERK expression induced by pH 5.0 PBS or norepinephrine, blocked sodium currents and abolished the production of action potentials evoked by current injection. The above effects were prevented by TRPV1 channel inhibitor SB366791, but not by ASIC channel inhibitor amiloride. Furthermore, acidic QX-314 employed adjacent to the

Spitzer mid-IR spectroscopy of powerful 2Jy and 3CRR radio galaxies. II. AGN power indicators and unification

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Dicken, D. [CEA-Saclay, F-91191 Gif-sur-Yvette (France); Tadhunter, C. [University of Sheffield, Hounsfield Road, Sheffield S3 7RH (United Kingdom); Morganti, R. [ASTRON, P.O. Box 2, 7990 AA Dwingeloo (Netherlands); Axon, D.; Robinson, A.; Magagnoli, M. [Rochester Institute of Technology, 84 Lomb Memorial Drive, Rochester, NY 14623 (United States); Kharb, P. [Indian Institute of Astrophysics, II Block, Koramangala, Bangalore 560034 (India); Ramos Almeida, C. [Instituto de Astrofisica de Canarias (IAC), C/V ia Lactea, s/n, E-38205 La Laguna, Tenerife (Spain); Mingo, B. [Department of Physics and Astronomy, University of Leicester, University Road, Leicester LE1 7RH (United Kingdom); Hardcastle, M. [School of Physics, Astronomy and Mathematics, University of Hertfordshire, College Lane, Hatfield AL10 9AB (United Kingdom); Nesvadba, N. P. H.; Singh, V. [Institut d' Astrophysique Spatiale, CNRS, Université Paris Sud, F-91405 Orsay (France); Kouwenhoven, M. B. N. [Kavli Institute for Astronomy and Astrophysics, Peking University, Yi He Yuan Lu 5, Haidian Qu, Beijing 100871 (China); Rose, M.; Spoon, H. [224 Space Sciences Building, Cornell University, Ithaca, NY 14853 (United States); Inskip, K. J. [Max Planck Institute for Astronomy, Königstuhl 17, D-69117 Heidelberg (Germany); Holt, J., E-mail: daniel.dicken@cea.fr [Leiden Observatory, Leiden University, P.O. Box 9513, 2300 RA Leiden (Netherlands)

2014-06-20

It remains uncertain which continuum and emission line diagnostics best indicate the bolometric powers of active galactic nuclei (AGNs), especially given the attenuation caused by the circumnuclear material and the possible contamination by components related to star formation. Here we use mid-IR spectra along with multiwavelength data to investigate the merit of various diagnostics of AGN radiative power, including the mid-IR [Ne III] λ25.89 μm and [O IV] λ25.89 μm fine-structure lines, the optical [O III] λ5007 forbidden line, and mid-IR 24 μm, 5 GHz radio, and X-ray continuum emission, for complete samples of 46 2Jy radio galaxies (0.05 < z < 0.7) and 17 3CRR FRII radio galaxies (z < 0.1). We find that the mid-IR [O IV] line is the most reliable indicator of AGN power for powerful radio-loud AGNs. By assuming that the [O IV] is emitted isotropically, and comparing the [O III] and 24 μm luminosities of the broad- and narrow-line AGNs in our samples at fixed [O IV] luminosity, we show that the [O III] and 24 μm emission are both mildly attenuated in the narrow-line compared to the broad-line objects by a factor of ≈2. However, despite this attenuation, the [O III] and 24 μm luminosities are better AGN power indicators for our sample than either the 5 GHz radio or the X-ray continuum luminosities. We also detect the mid-IR 9.7 μm silicate feature in the spectra of many objects but not ubiquitously: at least 40% of the sample shows no clear evidence for these features. We conclude that, for the majority of powerful radio galaxies, the mid-IR lines are powered by AGN photoionization.

Double transduction of a Cre/LoxP lentiviral vector: a simple method to generate kidney cell-specific knockdown mice.

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Nam, Bo Young; Kim, Dong Ki; Park, Jung Tak; Kang, Hye-Young; Paeng, Jisun; Kim, Seonghun; Park, Jimin; Um, Jae Eun; Oh, Hyung Jung; Han, Seung Hyeok; Yoo, Tae-Hyun; Kang, Shin-Wook

2015-12-15

In a lentivirus-based gene delivery system, the incorporated gene is continuously expressed for a long time. In this study, we devised a simple way to knock down a specific gene in a kidney cell-specific pattern in adult mice by lentivirus-assisted transfer of short hairpin RNA (shRNA). Kidney collecting duct (CD)-specific aquaporin-3 (AQP3)-knockdown mice were generated by consecutive injection of Hoxb7-Cre-expressing lentivirus (LV-Hoxb7 Cre) and loxP-AQP3 shRNA-expressing lentivirus (LV-loxP shAQP3) in adult C57BL6/J mice. LV-Hoxb7 Cre was designed to express mCherry, while LV-loxP shAQP3 was designed with a floxed enhanced green fluorescent protein (EGFP)-tagged stop sequence, and thus EGFP would be expressed only in the absence of Cre recombination. In mice treated with LV-Hoxb7 Cre alone, mCherry protein expression, which indicates the presence of Cre recombinase, occurred only in CD cells. However, LV-loxP shAQP3 injection alone resulted in an increase in EGFP expression in all kidney cells, indicating the transcription of the floxed region. When LV-Hoxb7 Cre and LV-loxP shAQP3 were sequentially transduced, EGFP expression was attenuated while mCherry expression was sustained in CD cells, demonstrating a CD cell-specific recombination of the floxed region. AQP3 expression in mice injected with LV-Hoxb7 Cre or LV-loxP shAQP3 alone did not differ, but consecutive injection of LV-Hoxb7 Cre and LV-loxP shAQP3 significantly reduced AQP3 expression in CD cells. However, the expression levels of AQP3 were not altered in other cell types. Double transduction of Cre- and loxP-based lentivirus can easily generate kidney cell-specific knockdown mice, and this method might be applicable to other species. Copyright © 2015 the American Physiological Society.

IL-18 Does not Increase Allergic Airway Disease in Mice When Produced by BCG

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Amniai, L.; Biet, F.; Marquillies, P.; Locht, C.; Pestel, J.; Tonnel, A.-B.; Duez, C.

2007-01-01

Whilst BCG inhibits allergic airway responses in murine models, IL-18 has adversary effects depending on its environment. We therefore constructed a BCG strain producing murine IL-18 (BCG-IL-18) and evaluated its efficiency to prevent an asthma-like reaction in mice. BALB/cByJ mice were sensitized (day (D) 1 and D10) by intraperitoneal injection of ovalbumin (OVA)-alum and primary (D20–22) and secondary (D62, 63) challenged with OVA aerosols. BCG or BCG-IL-18 were intraperitonealy administered 1 hour before each immunization (D1 and D10). BCG-IL-18 and BCG were shown to similarly inhibit the development of AHR, mucus production, eosinophil influx, and local Th2 cytokine production in BAL, both after the primary and secondary challenge. These data show that IL-18 did not increase allergic airway responses in the context of the mycobacterial infection, and suggest that BCG-IL-18 and BCG are able to prevent the development of local Th2 responses and therefore inhibit allergen-induced airway responses even after restimulation. PMID:18299704

Amphetamine sensitization in mice is sufficient to produce both manic- and depressive-related behaviors as well as changes in the functional connectivity of corticolimbic structures.

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Pathak, G; Ibrahim, B A; McCarthy, S A; Baker, K; Kelly, M P

2015-08-01

It has been suggested that amphetamine abuse and withdrawal mimics the diverse nature of bipolar disorder symptomatology in humans. Here, we determined if a single paradigm of amphetamine sensitization would be sufficient to produce both manic- and depressive-related behaviors in mice. CD-1 mice were subcutaneously dosed for 5 days with 1.8 mg/kg d-amphetamine or vehicle. On days 6-31 of withdrawal, amphetamine-sensitized (AS) mice were compared to vehicle-treated (VT) mice on a range of behavioral and biochemical endpoints. AS mice demonstrated reliable mania- and depression-related behaviors from day 7 to day 28 of withdrawal. Relative to VT mice, AS mice exhibited long-lasting mania-like hyperactivity following either an acute 30-min restraint stress or a low-dose 1 mg/kg d-amphetamine challenge, which was attenuated by the mood-stabilizers lithium and quetiapine. In absence of any challenge, AS mice showed anhedonia-like decreases in sucrose preference and depression-like impairments in the off-line consolidation of motor memory, as reflected by the lack of spontaneous improvement across days of training on the rotarod. AS mice also demonstrated a functional impairment in nest building, an ethologically-relevant activity of daily living. Western blot analyses revealed a significant increase in methylation of histone 3 at lysine 9 (H3K9), but not lysine 4 (H3K4), in hippocampus of AS mice relative to VT mice. In situ hybridization for the immediate-early gene activity-regulated cytoskeleton-associated protein (Arc) further revealed heightened activation of corticolimbic structures, decreased functional connectivity between frontal cortex and striatum, and increased functional connectivity between the amygdala and hippocampus of AS mice. The effects of amphetamine sensitization were blunted in C57BL/6J mice relative to CD-1 mice. These results show that a single amphetamine sensitization protocol is sufficient to produce behavioral, functional, and biochemical

Generation of healthy mice from gene-corrected disease-specific induced pluripotent stem cells.

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Guangming Wu

2011-07-01

Full Text Available Using the murine model of tyrosinemia type 1 (fumarylacetoacetate hydrolase [FAH] deficiency; FAH⁻/⁻ mice as a paradigm for orphan disorders, such as hereditary metabolic liver diseases, we evaluated fibroblast-derived FAH⁻/⁻-induced pluripotent stem cells (iPS cells as targets for gene correction in combination with the tetraploid embryo complementation method. First, after characterizing the FAH⁻/⁻ iPS cell lines, we aggregated FAH⁻/⁻-iPS cells with tetraploid embryos and obtained entirely FAH⁻/⁻-iPS cell-derived mice that were viable and exhibited the phenotype of the founding FAH⁻/⁻ mice. Then, we transduced FAH cDNA into the FAH⁻/⁻-iPS cells using a third-generation lentiviral vector to generate gene-corrected iPS cells. We could not detect any chromosomal alterations in these cells by high-resolution array CGH analysis, and after their aggregation with tetraploid embryos, we obtained fully iPS cell-derived healthy mice with an astonishing high efficiency for full-term development of up to 63.3%. The gene correction was validated functionally by the long-term survival and expansion of FAH-positive cells of these mice after withdrawal of the rescuing drug NTBC (2-(2-nitro-4-fluoromethylbenzoyl-1,3-cyclohexanedione. Furthermore, our results demonstrate that both a liver-specific promoter (transthyretin, TTR-driven FAH transgene and a strong viral promoter (from spleen focus-forming virus, SFFV-driven FAH transgene rescued the FAH-deficiency phenotypes in the mice derived from the respective gene-corrected iPS cells. In conclusion, our data demonstrate that a lentiviral gene repair strategy does not abrogate the full pluripotent potential of fibroblast-derived iPS cells, and genetic manipulation of iPS cells in combination with tetraploid embryo aggregation provides a practical and rapid approach to evaluate the efficacy of gene correction of human diseases in mouse models.

Immobilization induced osteopenia is strain specific in mice

DEFF Research Database (Denmark)

Lodberg, Andreas; Vegger, Jens Bay; Jensen, Michael Vinkel

2015-01-01

systemic effects on bone. Female mice from four inbred mouse strains (BALB/cJ, C57BL/6 J, DBA/2 J, and C3H/HeN) were injected unilaterally with BTX (n = 10/group) or unilaterally with saline (n = 10/group). Mice were euthanized after 21 days, and the bone properties evaluated using μCT, DXA, bone...... resolution μCT we found no evidence of a systemic effect on any of the microstructural parameters of the contralateral limb. Likewise, there was no evidence of a systemic effect on the bone strength in any mouse strain. We did, however, find a small systemic effect on aBMD in DBA/2 J and C3H/HeN mice...

Alteration of skin wound healing in keratinocyte-specific mediator complex subunit 1 null mice.

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Noguchi, Fumihito; Nakajima, Takeshi; Inui, Shigeki; Reddy, Janardan K; Itami, Satoshi

2014-01-01

MED1 (Mediator complex subunit 1) is a co-activator of various transcription factors that function in multiple transcriptional pathways. We have already established keratinocyte-specific MED1 null mice (Med1(epi-/-)) that develop epidermal hyperplasia. Herein, to investigate the function(s) of MED1 in skin wound healing, full-thickness skin wounds were generated in Med1(epi-/-) and age-matched wild-type mice and the healing process was analyzed. Macroscopic wound closure and the re-epithelialization rate were accelerated in 8-week-old Med1(epi-/-) mice compared with age-matched wild-type mice. Increased lengths of migrating epithelial tongues and numbers of Ki67-positive cells at the wounded epidermis were observed in 8-week-old Med1(epi-/-) mice, whereas wound contraction and the area of α-SMA-positive myofibroblasts in the granulation tissue were unaffected. Migration was enhanced in Med1(epi-/-) keratinocytes compared with wild-type keratinocytes in vitro. Immunoblotting revealed that the expression of follistatin was significantly decreased in Med1(epi-/-) keratinocytes. Moreover, the mitogen-activated protein kinase pathway was enhanced before and after treatment of Med1(epi-/-) keratinocytes with activin A in vitro. Cell-cycle analysis showed an increased ratio of S phase cells after activin A treatment of Med1(epi-/-) keratinocytes compared with wild-type keratinocytes. These findings indicate that the activin-follistatin system is involved in this acceleration of skin wound healing in 8-week-old Med1(epi-/-) mice. On the other hand, skin wound healing in 6-month-old Med1(epi-/-) mice was significantly delayed with decreased numbers of Ki67-positive cells at the wounded epidermis as well as BrdU-positive label retaining cells in hair follicles compared with age-matched wild-type mice. These results agree with our previous observation that hair follicle bulge stem cells are reduced in older Med1(epi-/-) mice, indicating a decreased contribution of hair

Alteration of skin wound healing in keratinocyte-specific mediator complex subunit 1 null mice.

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Fumihito Noguchi

Full Text Available MED1 (Mediator complex subunit 1 is a co-activator of various transcription factors that function in multiple transcriptional pathways. We have already established keratinocyte-specific MED1 null mice (Med1(epi-/- that develop epidermal hyperplasia. Herein, to investigate the function(s of MED1 in skin wound healing, full-thickness skin wounds were generated in Med1(epi-/- and age-matched wild-type mice and the healing process was analyzed. Macroscopic wound closure and the re-epithelialization rate were accelerated in 8-week-old Med1(epi-/- mice compared with age-matched wild-type mice. Increased lengths of migrating epithelial tongues and numbers of Ki67-positive cells at the wounded epidermis were observed in 8-week-old Med1(epi-/- mice, whereas wound contraction and the area of α-SMA-positive myofibroblasts in the granulation tissue were unaffected. Migration was enhanced in Med1(epi-/- keratinocytes compared with wild-type keratinocytes in vitro. Immunoblotting revealed that the expression of follistatin was significantly decreased in Med1(epi-/- keratinocytes. Moreover, the mitogen-activated protein kinase pathway was enhanced before and after treatment of Med1(epi-/- keratinocytes with activin A in vitro. Cell-cycle analysis showed an increased ratio of S phase cells after activin A treatment of Med1(epi-/- keratinocytes compared with wild-type keratinocytes. These findings indicate that the activin-follistatin system is involved in this acceleration of skin wound healing in 8-week-old Med1(epi-/- mice. On the other hand, skin wound healing in 6-month-old Med1(epi-/- mice was significantly delayed with decreased numbers of Ki67-positive cells at the wounded epidermis as well as BrdU-positive label retaining cells in hair follicles compared with age-matched wild-type mice. These results agree with our previous observation that hair follicle bulge stem cells are reduced in older Med1(epi-/- mice, indicating a decreased contribution of hair

Protective effect of a non specific inflammation on bone marrow protein synthesis in irradiated mice

International Nuclear Information System (INIS)

Herodin, F.; Roques, P.; Court, L.

1988-01-01

Gamma radiations exert a decrease in mouse bone marrow total protein synthesis. A non-specific inflammatory process induced with polyacrylamide microbeads stimulates spleen and marrow protein synthesis and protects the medullar protein synthesis in irradiated mice [fr

T-helper 17 and interleukin-17-producing lymphoid tissue inducer-like cells make different contributions to colitis in mice.

Science.gov (United States)

Ono, Yuichi; Kanai, Takanori; Sujino, Tomohisa; Nemoto, Yasuhiro; Kanai, Yasumasa; Mikami, Yohei; Hayashi, Atsushi; Matsumoto, Atsuhiro; Takaishi, Hiromasa; Ogata, Haruhiko; Matsuoka, Katsuyoshi; Hisamatsu, Tadakazu; Watanabe, Mamoru; Hibi, Toshifumi

2012-11-01

T helper (Th) 17 cells that express the retinoid-related orphan receptor (ROR) γt contribute to the development of colitis in mice, yet are found in normal and inflamed intestine. We investigated their development and functions in intestines of mice. We analyzed intestinal Th17 cells in healthy and inflamed intestinal tissues of mice. We analyzed expression of lymphotoxin (LT)α by Th17 cells and lymphoid tissue inducer-like cells. LTα(-/-) and RORγt(-/-) mice had significantly lower percentages of naturally occurring Th17 cells in the small intestine than wild-type mice. Numbers of CD3(-)CD4(+/-)interleukin-7Rα(+)c-kit(+)CCR6(+)NKp46(-) lymphoid tissue inducer-like cells that produce interleukin-17A were increased in LTα(-/-) and LTα(-/-) × recombination activating gene (RAG)-2(-/-) mice, compared with wild-type mice, but were absent from RORγt(-/-) mice. Parabiosis of wild-type and LTα(-/-) mice and bone marrow transplant experiments revealed that LTα-dependent gut-associated lymphoid tissue structures are required for generation of naturally occurring Th17 cells. However, when wild-type or LTα(-/-) CD4(+)CD45RB(high) T cells were transferred to RAG-2(-/-) or LTα(-/-)×RAG-2(-/-) mice, all groups, irrespective of the presence or absence of LTα on the donor or recipient cells, developed colitis and generated Th1, Th17, and Th17/Th1 cells. RAG-2(-/-) mice that received a second round of transplantation, with colitogenic but not naturally occurring Th17 cells, developed intestinal inflammation. The presence of naturally occurring Th17 cells in the colons of mice inhibited development of colitis after transfer of CD4(+)CD45RB(high) T cells and increased the numbers of Foxp3(+) cells derived from CD4(+)CD45RB(high) T cells. Gut-associated lymphoid tissue structures are required to generate naturally occurring Th17 cells that have regulatory activities in normal intestines of mice, but not for colitogenic Th17 and Th17/Th1 cells during inflammation

Astrocyte-Specific Overexpression of Insulin-Like Growth Factor-1 Protects Hippocampal Neurons and Reduces Behavioral Deficits following Traumatic Brain Injury in Mice.

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Sindhu K Madathil

Full Text Available Traumatic brain injury (TBI survivors often suffer from long-lasting cognitive impairment that stems from hippocampal injury. Systemic administration of insulin-like growth factor-1 (IGF-1, a polypeptide growth factor known to play vital roles in neuronal survival, has been shown to attenuate posttraumatic cognitive and motor dysfunction. However, its neuroprotective effects in TBI have not been examined. To this end, moderate or severe contusion brain injury was induced in mice with conditional (postnatal overexpression of IGF-1 using the controlled cortical impact (CCI injury model. CCI brain injury produces robust reactive astrocytosis in regions of neuronal damage such as the hippocampus. We exploited this regional astrocytosis by linking expression of hIGF-1 to the astrocyte-specific glial fibrillary acidic protein (GFAP promoter, effectively targeting IGF-1 delivery to vulnerable neurons. Following brain injury, IGF-1Tg mice exhibited a progressive increase in hippocampal IGF-1 levels which was coupled with enhanced hippocampal reactive astrocytosis and significantly greater GFAP levels relative to WT mice. IGF-1 overexpression stimulated Akt phosphorylation and reduced acute (1 and 3d hippocampal neurodegeneration, culminating in greater neuron survival at 10d after CCI injury. Hippocampal neuroprotection achieved by IGF-1 overexpression was accompanied by improved motor and cognitive function in brain-injured mice. These data provide strong support for the therapeutic efficacy of increased brain levels of IGF-1 in the setting of TBI.

Astrocyte-Specific Overexpression of Insulin-Like Growth Factor-1 Protects Hippocampal Neurons and Reduces Behavioral Deficits following Traumatic Brain Injury in Mice

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Madathil, Sindhu K.; Carlson, Shaun W.; Brelsfoard, Jennifer M.; Ye, Ping; D’Ercole, A. Joseph; Saatman, Kathryn E.

2013-01-01

Traumatic brain injury (TBI) survivors often suffer from long-lasting cognitive impairment that stems from hippocampal injury. Systemic administration of insulin-like growth factor-1 (IGF-1), a polypeptide growth factor known to play vital roles in neuronal survival, has been shown to attenuate posttraumatic cognitive and motor dysfunction. However, its neuroprotective effects in TBI have not been examined. To this end, moderate or severe contusion brain injury was induced in mice with conditional (postnatal) overexpression of IGF-1 using the controlled cortical impact (CCI) injury model. CCI brain injury produces robust reactive astrocytosis in regions of neuronal damage such as the hippocampus. We exploited this regional astrocytosis by linking expression of hIGF-1 to the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter, effectively targeting IGF-1 delivery to vulnerable neurons. Following brain injury, IGF-1Tg mice exhibited a progressive increase in hippocampal IGF-1 levels which was coupled with enhanced hippocampal reactive astrocytosis and significantly greater GFAP levels relative to WT mice. IGF-1 overexpression stimulated Akt phosphorylation and reduced acute (1 and 3d) hippocampal neurodegeneration, culminating in greater neuron survival at 10d after CCI injury. Hippocampal neuroprotection achieved by IGF-1 overexpression was accompanied by improved motor and cognitive function in brain-injured mice. These data provide strong support for the therapeutic efficacy of increased brain levels of IGF-1 in the setting of TBI. PMID:23826235

Depletion of alveolar macrophages in CD11c diphtheria toxin receptor mice produces an inflammatory response

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Roberts, Lydia M; Ledvina, Hannah E; Tuladhar, Shraddha; Rana, Deepa; Steele, Shaun P; Sempowski, Gregory D; Frelinger, Jeffrey A

2015-01-01

Alveolar macrophages play a critical role in initiating the immune response to inhaled pathogens and have been shown to be the first cell type infected following intranasal inoculation with several pathogens, including Francisella tularensis. In an attempt to further dissect the role of alveolar macrophages in the immune response to Francisella, we selectively depleted alveolar macrophages using CD11c.DOG mice. CD11c.DOG mice express the diphtheria toxin receptor (DTR) under control of the full CD11c promoter. Because mice do not express DTR, tissue restricted expression of the primate DTR followed by treatment with diphtheria toxin (DT) has been widely used as a tool in immunology to examine the effect of acute depletion of a specific immune subset following normal development. We successfully depleted alveolar macrophages via intranasal administration of DT. However, alveolar macrophage depletion was accompanied by many other changes to the cellular composition and cytokine/chemokine milieu in the lung that potentially impact innate and adaptive immune responses. Importantly, we observed a transient influx of neutrophils in the lung and spleen. Our experience serves as a cautionary note to other researchers using DTR mice given the complex changes that occur following DT treatment that must be taken into account when analyzing data. PMID:26029367

Cyclic GMP-AMP Displays Mucosal Adjuvant Activity in Mice

OpenAIRE

Škrnjug, Ivana; Guzmán, Carlos Alberto; Ruecker, Christine

2014-01-01

The recently discovered mammalian enzyme cyclic GMP-AMP synthase produces cyclic GMP-AMP (cGAMP) after being activated by pathogen-derived cytosolic double stranded DNA. The product can stimulate STING-dependent interferon type I signaling. Here, we explore the efficacy of cGAMP as a mucosal adjuvant in mice. We show that cGAMP can enhance the adaptive immune response to the model antigen ovalbumin. It promotes antigen specific IgG and a balanced Th1/Th2 lymphocyte response in immunized mice....

An Immunoglobulin G1 Monoclonal Antibody Highly Specific to the Wall of Cryptosporidium Oocysts

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Weir, C.; Vesey, G.; Slade, M.; Ferrari, B.; Veal, D. A.; Williams, K.

2000-01-01

The detection of Cryptosporidium oocysts in drinking water is critically dependent on the quality of immunofluorescent reagents. Experiments were performed to develop a method for producing highly specific antibodies to Cryptosporidium oocysts that can be used for water testing. BALB/c mice were immunized with six different antigen preparations and monitored for immunoglobulin G (IgG) and IgM responses to the surface of Cryptosporidium oocysts. One group of mice received purified oocyst walls, a second group received a soluble protein preparation extracted from the outside of the oocyst wall, and the third group received whole inactivated oocysts. Three additional groups were immunized with sequentially prepared oocyst extracts to provide for a comparison of the immune response. Mice injected with the soluble protein extract demonstrated an IgG response to oocysts surface that was not seen in the whole-oocyst group. Mice injected with whole oocysts showed an IgM response only, while mice injected with purified oocyst walls showed little increase in IgM or IgG levels. Of the additional reported preparations only one, BME (2-mercaptoethanol treated), produced a weak IgM response to the oocyst wall. A mouse from the soluble oocyst extract group yielding a high IgG response was utilized to produce a highly specific IgG1 monoclonal antibody (Cry104) specific to the oocyst surface. Comparative flow cytometric analysis indicated that Cry104 has a higher avidity and specificity to oocysts in water concentrates than other commercially available antibodies. PMID:10973448

Deficient CD4+ T cell priming and regression of CD8+ T cell functionality in virus-infected mice lacking a normal B cell compartment

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Christensen, Jan Pravsgaard; Kauffmann, Susanne Ørding; Thomsen, Allan Randrup

2003-01-01

of virus-specific CD4(+) T cells was markedly impaired in B(-/-) mice infected with either virus strain. Thus, our results indicate that B cells play an important role in antiviral immunity not only as Ab producers, but also in promoting an optimal and sustained T cell response. The T cell defects......In this study, we investigate the state of T cell-mediated immunity in B cell-deficient (B(-/-)) mice infected with two strains of lymphocytic choriomeningitis virus known to differ markedly in their capacity to persist. In B(-/-) C57BL mice infected with the more persisting virus, virus......-specific CD8(+) T cells are initially generated that are qualitatively similar to those in wild-type mice. However, although cell numbers are well sustained over time, the capacity to produce cytokines is rapidly impaired. In similarly infected B(-/-) BALB/c mice, virus-specific CD8(+) T cells are completely...

NOG-hIL-4-Tg, a new humanized mouse model for producing tumor antigen-specific IgG antibody by peptide vaccination.

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Yoshie Kametani

Full Text Available Immunodeficient mice transplanted with human peripheral blood mononuclear cells (PBMCs are promising tools to evaluate human immune responses to vaccines. However, these mice usually develop severe graft-versus-host disease (GVHD, which makes estimation of antigen-specific IgG production after antigen immunization difficult. To evaluate antigen-specific IgG responses in PBMC-transplanted immunodeficient mice, we developed a novel NOD/Shi-scid-IL2rγnull (NOG mouse strain that systemically expresses the human IL-4 gene (NOG-hIL-4-Tg. After human PBMC transplantation, GVHD symptoms were significantly suppressed in NOG-hIL-4-Tg compared to conventional NOG mice. In kinetic analyses of human leukocytes, long-term engraftment of human T cells has been observed in peripheral blood of NOG-hIL-4-Tg, followed by dominant CD4+ T rather than CD8+ T cell proliferation. Furthermore, these CD4+ T cells shifted to type 2 helper (Th2 cells, resulting in long-term suppression of GVHD. Most of the human B cells detected in the transplanted mice had a plasmablast phenotype. Vaccination with HER2 multiple antigen peptide (CH401MAP or keyhole limpet hemocyanin (KLH successfully induced antigen-specific IgG production in PBMC-transplanted NOG-hIL-4-Tg. The HLA haplotype of donor PBMCs might not be relevant to the antibody secretion ability after immunization. These results suggest that the human PBMC-transplanted NOG-hIL-4-Tg mouse is an effective tool to evaluate the production of antigen-specific IgG antibodies.

Islet-specific T cell clones transfer diabetes to nonobese diabetic (NOD) F1 mice.

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Peterson, J D; Pike, B; McDuffie, M; Haskins, K

1994-09-15

To investigate diabetes resistance to T cell-mediated disease transfer, we administered islet-specific T cell clones to the F1 progeny of nonobese diabetic (NOD) mice that were crossed with various nondiabetes-prone inbred mouse strains. We investigated four diabetogenic CD4+ T cell clones and all induced insulitis and full development of diabetes in (SWR x NOD)F1, (SJL x NOD)F1, and (C57BL/6 x NOD)F1 mice. In contrast, (BALB/c x NOD)F1 and (CBA x NOD)F1 mice were susceptible to disease transfer by some T cell clones but not others, and (C57/L x NOD)F1 mice seemed to be resistant to both insulitis and disease transfer by all of the clones tested. Disease induced by the T cell clones in susceptible F1 strains was age dependent and could only be observed in recipients younger than 13 days old. Full or partial disease resistance did not correlate with the presence or absence of I-E, different levels of Ag expression in islet cells, or differences in APC function. The results from this study suggest that there may be multiple factors contributing to susceptibility of F1 mice to T cell clone-mediated induction of diabetes, including non-MHC-related genetic background, the immunologic maturity of the recipient, and individual characteristics of the T cell clones.

Specific Macronutrients Exert Unique Influences on the Adipose-Liver Axis to Promote Hepatic Steatosis in Mice.

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Duwaerts, Caroline C; Amin, Amin M; Siao, Kevin; Her, Chris; Fitch, Mark; Beysen, Carine; Turner, Scott M; Goodsell, Amanda; Baron, Jody L; Grenert, James P; Cho, Soo-Jin; Maher, Jacquelyn J

2017-09-01

The factors that distinguish metabolically healthy obesity from metabolically unhealthy obesity are not well understood. Diet has been implicated as a determinant of the unhealthy obesity phenotype, but which aspects of the diet induce dysmetabolism are unknown. The goal of this study was to investigate whether specific macronutrients or macronutrient combinations provoke dysmetabolism in the context of isocaloric, high-energy diets. Mice were fed 4 high-energy diets identical in calorie and nutrient content but different in nutrient composition for 3 weeks to 6 months. The test diets contained 42% carbohydrate (sucrose or starch) and 42% fat (oleate or palmitate). Weight and glucose tolerance were monitored; blood and tissues were collected for histology, gene expression, and immunophenotyping. Mice gained weight on all 4 test diets but differed significantly in other metabolic outcomes. Animals fed the starch-oleate diet developed more severe hepatic steatosis than those on other formulas. Stable isotope incorporation showed that the excess hepatic steatosis in starch-oleate-fed mice derived from exaggerated adipose tissue lipolysis. In these mice, adipose tissue lipolysis coincided with adipocyte necrosis and inflammation. Notably, the liver and adipose tissue abnormalities provoked by starch-oleate feeding were reproduced when mice were fed a mixed-nutrient Western diet with 42% carbohydrate and 42% fat. The macronutrient composition of the diet exerts a significant influence on metabolic outcome, independent of calories and nutrient proportions. Starch-oleate appears to cause hepatic steatosis by inducing progressive adipose tissue injury. Starch-oleate phenocopies the effect of a Western diet; consequently, it may provide clues to the mechanism whereby specific nutrients cause metabolically unhealthy obesity.

Immunoglobulin gene expression and regulation of rearrangement in kappa transgenic mice

International Nuclear Information System (INIS)

Ritchie, K.A.

1986-01-01

Transgenic mice were produced by microinjection of the functionally rearranged immunoglobulin kappa gene from the myeloma MOPC-21 into the male pronucleus of fertilized mouse eggs, and implantation of the microinjected embryos into foster mothers. Mice that integrated the injected gene were detected by hybridizing tail DNA dots with radioactively labelled pBR322 plasmid DNA, which detects pBR322 sequences left as a tag on the microinjected DNA. Mice that integrated the injected gene (six males) were mated and the DNA, RNA and serum kappa chains of their offspring were analyzed. A rabbit anti-mouse kappa chain antiserum was also produced for use in detection of mouse kappa chains on protein blots. Hybridomas were produced from the spleen cells of these kappa transgenic mice to immortalize representative B cells and to investigate expression of the transgenic kappa gene, its effect on allelic exclusion, and its effect on the control of light chain gene rearrangement and expression. The results show that the microinjected DNA is integrated as concatamers in unique single or, rarely, two separate sites in the genome. The concatamers are composed of several copies (16 to 64) of injected DNA arranged in a head to tail fashion. The transgene is expressed into protein normally and in a tissue specific fashion. For the first time in these transgenic mice, all tissues contain a functionally rearranged and potentially expressible immunoglobulin gene. The transgene is expressed only in B cells and not in hepatocytes, for example. This indicates that rearrangement of immunoglobulin genes is necessary but not sufficient for the tissue specific expression of these genes by B cells

Antigenotoxic effect of Saccharomyces cerevisiae on the damage produced in mice fed with aflatoxin B(1) contaminated corn.

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Madrigal-Santillán, E; Madrigal-Bujaidar, E; Márquez-Márquez, R; Reyes, A

2006-12-01

The potential of Saccharomyces cerevisiae (Sc) was evaluated for reducing the micronucleated normochromatic erythrocytes (MNNE) rate in mice fed AFB(1) contaminated corn. The study included two groups fed AFB(1) contaminated corn (0.4 and 0.8 mg/kg), a control fed uncontaminated corn, another group fed uncontaminated corn and 0.3% of Sc (1 x 10(8) live cells/g), and two groups fed AFB(1) contaminated corn (0.4 and 0.8 mg/kg) plus 0.3% Sc. Weight and MNNE were determined weekly for six weeks. Subsequently, the same determinations were made for another three-week period, but in mice receiving only a normal diet, without AFB(1) and Sc. Results in the first period revealed the following: control and Sc fed mice had similar constant weight increase, and low MNNE rate; mice fed only AFB(1) showed weight decrease and significant MNNE increase; finally, Sc improved weight gain and reduced MNNE produced by AFB(1). In the second period, results exhibited a tendency similar to that of the previous phase in the control and Sc fed mice; the weight and MNNE values improved in the other groups. We also determined the capacity of Sc for adsorbing and modifying the mycotoxin structure. The mixture was filtered to obtain two phases, and AFB(1) content was measured. Sc revealed a potent adsorbent capacity; however, chromatographic determination suggested no structural modification.

Sex-specific phenotypes of hyperthyroidism and hypothyroidism in mice.

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Rakov, Helena; Engels, Kathrin; Hönes, Georg Sebastian; Strucksberg, Karl-Heinz; Moeller, Lars Christian; Köhrle, Josef; Zwanziger, Denise; Führer, Dagmar

2016-01-01

Thyroid dysfunction is more common in the female population, however, the impact of sex on disease characteristics has rarely been addressed. Using a murine model, we asked whether sex has an influence on phenotypes, thyroid hormone status, and thyroid hormone tissue response in hyper- and hypothyroidism. Hypo- and hyperthyroidism were induced in 5-month-old female and male wildtype C57BL/6N mice, by LoI/MMI/ClO4 (-) or T4 i.p. treatment over 7 weeks, and control animals underwent sham treatment (N = 8 animals/sex/treatment). Animals were investigated for impact of sex on body weight, food and water intake, body temperature, heart rate, behaviour (locomotor activity, motor coordination, and strength), liver function, serum thyroid hormone status, and cellular TH effects on gene expression in brown adipose tissue, heart, and liver. Male and female mice showed significant differences in behavioural, functional, metabolic, biochemical, and molecular traits of hyper- and hypothyroidism. Hyperthyroidism resulted in increased locomotor activity in female mice but decreased muscle strength and motor coordination preferably in male animals. Hypothyroidism led to increased water intake in male but not female mice and significantly higher serum cholesterol in male mice. Natural sex differences in body temperature, body weight gain, food and water intake were preserved under hyperthyroid conditions. In contrast, natural sex differences in heart rate disappeared with TH excess and deprivation. The variations of hyper- or hypothyroid traits of male and female mice were not explained by classical T3/T4 serum state. TH serum concentrations were significantly increased in female mice under hyperthyroidism, but no sex differences were found under eu- or hypothyroid conditions. Interestingly, analysis of expression of TH target genes and TH transporters revealed little sex dependency in heart, while sex differences in target genes were present in liver and brown adipose tissue

Comparison of antibody and cytokine responses to primary Giardia muris infection in H-2 congenic strains of mice.

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Venkatesan, P; Finch, R G; Wakelin, D

1996-11-01

The course of primary infections with Giardia muris differs between BALB and B10 H-2 congenic strains of mice. In the first 3 weeks of infection, there is a more rapid decline in intestinal trophozoite and fecal cyst counts in B10 strains than in BALB strains. To determine whether this difference could be explained by variation in specific antibody responses, both secretory immunoglobulin A (IgA) and serum antibody responses were compared between these strains. No significant differences in the timing, titer, or specificity of secretory or serum antibodies were found. However, on comparing specific anti-G. muris serum IgG subclass responses, we found that B10 strains produced IgG2a while BALB strains produced IgG1, suggesting differential involvement of T helper 1 and 2 subsets of lymphocytes. When cells harvested from mesenteric lymph nodes were stimulated with concanavalin A in vitro, both gamma interferon and interleukin-5 were secreted by cells from B10 mice, but only interleukin-5 was secreted by cells from BALB/c mice. Specific blockade of gamma interferon by monoclonal antibody administered to B10 mice resulted in an enhanced intensity of infection.

Circumsporozoite Protein-Specific Kd-Restricted CD8+ T Cells Mediate Protective Antimalaria Immunity in Sporozoite-Immunized MHC-I-Kd Transgenic Mice

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Jing Huang

2014-01-01

Full Text Available Although the roles of CD8+ T cells and a major preerythrocytic antigen, the circumsporozoite (CS protein, in contributing protective antimalaria immunity induced by radiation-attenuated sporozoites, have been shown by a number of studies, the extent to which these players contribute to antimalaria immunity is still unknown. To address this question, we have generated C57BL/6 (B6 transgenic (Tg mice, expressing Kd molecules under the MHC-I promoter, called MHC-I-Kd-Tg mice. In this study, we first determined that a single immunizing dose of IrPySpz induced a significant level of antimalaria protective immunity in MHC-I-Kd-Tg mice but not in B6 mice. Then, by depleting various T-cell subsets in vivo, we determined that CD8+ T cells are the main mediator of the protective immunity induced by IrPySpz. Furthermore, when we immunized (MHC-I-Kd-Tg × CS-Tg F1 mice with IrPySpz after crossing MHC-I-Kd-Tg mice with PyCS-transgenic mice (CS-Tg, which are unable to mount PyCS-specific immunity, we found that IrPySpz immunization failed to induce protective antimalaria immunity in (MHC-I-Kd-Tg × CS-Tg F1 mice, thus indicating the absence of PyCS antigen-dependent immunity in these mice. These results indicate that protective antimalaria immunity induced by IrPySpz in MHC-I-Kd-Tg mice is mediated by CS protein-specific, Kd-restricted CD8+ T cells.

Site-specific induction of nuclear anomalies (apoptotic bodies and micronuclei) by carcinogens in mice

International Nuclear Information System (INIS)

Ronen, A.; Heddle, J.A.

1984-01-01

The usefulness of nuclear anomalies (NA) as a short-term test for indication of carcinogens in the mouse colon has been suggested previously by experiments in which colon-specific carcinogens induced NA in the colon, whereas non-colon carcinogens were, in general, impotent in that organ. We have extended this work to other sites in the digestive tract of female C57BL/6 mice treated with gamma-rays, 1,2-dimethylhydrazine dihydrochloride, or N-methylnitrosourea. Each agent induced NA at all of the sites examined. The frequency of NA at different times after treatment depended upon both the agent used and the site examined. 1,2-Dimethylhydrazine dihydrochloride (which is known to induce tumors predominantly in the colon) induces NA with the highest efficiency (relative to gamma-rays) in the descending colon. N-Methylnitrosourea (which induces tumors mainly in the forestomach) induces NA with the highest efficiency in the forestomach. These results further support the usefulness of the assay in that the frequency of NA produced at the various sites by 1,2-dimethylhydrazine dihydrochloride and N-methylnitrosourea correlates with that found in the carcinogenicity studies

A novel system for tracking social preference dynamics in mice reveals sex- and strain-specific characteristics.

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Netser, Shai; Haskal, Shani; Magalnik, Hen; Wagner, Shlomo

2017-01-01

Deciphering the biological mechanisms underlying social behavior in animal models requires standard behavioral paradigms that can be unbiasedly employed in an observer- and laboratory-independent manner. During the past decade, the three-chamber test has become such a standard paradigm used to evaluate social preference (sociability) and social novelty preference in mice. This test suffers from several caveats, including its reliance on spatial navigation skills and negligence of behavioral dynamics. Here, we present a novel experimental apparatus and an automated analysis system which offer an alternative to the three-chamber test while solving the aforementioned caveats. The custom-made apparatus is simple for production, and the analysis system is publically available as an open-source software, enabling its free use. We used this system to compare the dynamics of social behavior during the social preference and social novelty preference tests between male and female C57BL/6J mice. We found that in both tests, male mice keep their preference towards one of the stimuli for longer periods than females. We then employed our system to define several new parameters of social behavioral dynamics in mice and revealed that social preference behavior is segregated in time into two distinct phases. An early exploration phase, characterized by high rate of transitions between stimuli and short bouts of stimulus investigation, is followed by an interaction phase with low transition rate and prolonged interactions, mainly with the preferred stimulus. Finally, we compared the dynamics of social behavior between C57BL/6J and BTBR male mice, the latter of which are considered as asocial strain serving as a model for autism spectrum disorder. We found that BTBR mice ( n  = 8) showed a specific deficit in transition from the exploration phase to the interaction phase in the social preference test, suggesting a reduced tendency towards social interaction. We successfully

Tau passive immunotherapy in mutant P301L mice: antibody affinity versus specificity.

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Cristina d'Abramo

Full Text Available The use of antibodies to treat neurodegenerative diseases has undergone rapid development in the past decade. To date, immunotherapeutic approaches to Alzheimer's disease have mostly targeted amyloid beta as it is a secreted protein that can be found in plasma and CSF and is consequently accessible to circulating antibodies. Few recent publications have suggested the utility of treatment of tau pathology with monoclonal antibodies to tau. Our laboratory has begun a systematic study of different classes of tau monoclonal antibodies using mutant P301L mice. Three or seven months old mutant tau mice were inoculated weekly with tau monoclonal antibodies at a dose of 10 mg/Kg, until seven or ten months of age were reached respectively. Our data strongly support the notion that in P301L animals treated with MC1, a conformational monoclonal antibody specific for PHF-tau, the rate of development of tau pathology is effectively reduced, while injecting DA31, a high affinity tau sequence antibody, does not exert such benefit. MC1 appears superior to DA31 in overall effects, suggesting that specificity is more important than affinity in therapeutic applications. Unfortunately the survival rate of the P301L treated mice was not improved when immunizing either with MC1 or PHF1, a high affinity phospho-tau antibody previously reported to be efficacious in reducing pathological tau. These data demonstrate that passive immunotherapy in mutant tau models may be efficacious in reducing the development of tau pathology, but a great deal of work remains to be done to carefully select the tau epitopes to target.

Altered consolidation of extinction-like inhibitory learning in genotype-specific dysfunctional coping fostered by chronic stress in mice.

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Campus, P; Maiolati, M; Orsini, C; Cabib, S

2016-12-15

Genetic and stress-related factors interact to foster mental disorders, possibly through dysfunctional learning. In a previous study we reported that a temporary experience of reduced food availability increases forced swim (FS)-induced helplessness tested 14days after a first experience in mice of the standard inbred C57BL/6(B6) strain but reduces it in mice of the genetically unrelated DBA/2J (D2) strain. Because persistence of FS-induced helplessness influences adaptive coping with stress challenge and involve learning processes the present study tested whether the behavioral effects of restricted feeding involved altered consolidation of FS-related learning. First, we demonstrated that restricted feeding does not influence behavior expressed on the first FS experience, supporting a specific effect on persistence rather then development of helplessness. Second, we found that FS-induced c-fos expression in the infralimbic cortex (IL) was selectively enhanced in food-restricted (FR) B6 mice and reduced in FR D2 mice, supporting opposite alterations of consolidation processes involving this brain area. Third, we demonstrated that immediate post-FS inactivation of IL prevents 24h retention of acquired helplessness by continuously free-fed mice of both strains, indicating the requirement of a functioning IL for consolidation of FS-related learning in either mouse strain. Finally, in line with the known role of IL in consolidation of extinction memories, we found that restricted feeding selectively facilitated 24h retention of an acquired extinction in B6 mice whereas impairing it in D2 mice. These findings support the conclusion that an experience of reduced food availability strain-specifically affects persistence of newly acquired passive coping strategies by altering consolidation of extinction-like inhibitory learning. Copyright © 2016 Elsevier B.V. All rights reserved.

Cardiac-specific inducible overexpression of human plasma membrane Ca2+ ATPase 4b is cardioprotective and improves survival in mice following ischemic injury.

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Sadi, Al Muktafi; Afroze, Talat; Siraj, M Ahsan; Momen, Abdul; White-Dzuro, Colin; Zarrin-Khat, Dorrin; Handa, Shivalika; Ban, Kiwon; Kabir, M Golam; Trivieri, Maria G; Gros, Robert; Backx, Peter; Husain, Mansoor

2018-03-30

Background: Heart failure (HF) is associated with reduced expression of plasma membrane Ca 2+ -ATPase 4 (PMCA4). Cardiac-specific overexpression of human PMCA4b in mice inhibited nNOS activity and reduced cardiac hypertrophy by inhibiting calcineurin. Here we examine temporally regulated cardiac-specific overexpression of hPMCA4b in mouse models of myocardial ischemia reperfusion injury (IRI) ex vivo , and HF following experimental myocardial infarction (MI) in vivo Methods and results: Doxycycline-regulated cardiomyocyte-specific overexpression and activity of hPMCA4b produced adaptive changes in expression levels of Ca 2+ -regulatory genes, and induced hypertrophy without significant differences in Ca 2+ transients or diastolic Ca 2+ concentrations. Total cardiac NOS and nNOS-specific activities were reduced in mice with cardiac overexpression of hPMCA4b while nNOS, eNOS and iNOS protein levels did not differ. hMPCA4b-overexpressing mice also exhibited elevated systolic blood pressure vs. controls, with increased contractility and lusitropy in vivo In isolated hearts undergoing IRI, hPMCA4b overexpression was cardioprotective. NO donor-treated hearts overexpressing hPMCA4b showed reduced LVDP and larger infarct size versus vehicle-treated hearts undergoing IRI, demonstrating that the cardioprotective benefits of hPMCA4b-repressed nNOS are lost by restoring NO availability. Finally, both pre-existing and post-MI induction of hPMCA4b overexpression reduced infarct expansion and improved survival from HF. Conclusions: Cardiac PMCA4b regulates nNOS activity, cardiac mass and contractility, such that PMCA4b overexpression preserves cardiac function following IRI, heightens cardiac performance and limits infarct progression, cardiac hypertrophy and HF, even when induced late post-MI. These data identify PMCA4b as a novel therapeutic target for IRI and HF. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Developmental Stage-Specific Manifestations of Absent TPO/c-MPL Signalling in Newborn Mice.

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Lorenz, Viola; Ramsey, Haley; Liu, Zhi-Jian; Italiano, Joseph; Hoffmeister, Karin; Bihorel, Sihem; Mager, Donald; Hu, Zhongbo; Slayton, William B; Kile, Benjamin T; Sola-Visner, Martha; Ferrer-Marin, Francisca

2017-12-01

Congenital amegakaryocytic thrombocytopaenia (CAMT) is a disorder caused by c-MPL mutations that impair thrombopoietin (TPO) signalling, resulting in a near absence of megakaryocytes (MKs). While this phenotype is consistent in adults, neonates with CAMT can present with severe thrombocytopaenia despite normal MK numbers. To investigate this, we characterized MKs and platelets in newborn c-MPL –/– mice. Liver MKs in c-MPL –/– neonates were reduced in number and size compared with wild-type (WT) age-matched MKs, and exhibited ultrastructural abnormalities not found in adult c-MPL –/– MKs. Platelet counts were lower in c-MPL –/– compared with WT mice at birth and did not increase over the first 2 weeks of life. In vivo biotinylation revealed a significant reduction in the platelet half-life of c-MPL –/– newborn mice (P2) compared with age-matched WT pups, which was not associated with ultrastructural abnormalities. Genetic deletion of the pro-apoptotic Bak did not rescue the severely reduced platelet half-life of c-MPL –/– newborn mice, suggesting that it was due to factors other than platelets entering apoptosis early. Indeed, adult GFP+ (green fluorescent protein transgenic) platelets transfused into thrombocytopenic c-MPL –/– P2 pups also had a shortened lifespan, indicating the importance of cell-extrinsic factors. In addition, neonatal platelets from WT and c-MPL –/– mice exhibited reduced P-selectin surface expression following stimulation compared with adult platelets of either genotype, and platelets from c-MPL –/– neonates exhibited reduced glycoprotein IIb/IIIa (GPIIb/IIIa) activation in response to thrombin compared with age-matched WT platelets. Taken together, our findings indicate that c-MPL deficiency is associated with abnormal maturation of neonatal MKs and developmental stage-specific defects in platelet function.

Properties of murine leukemia viruses produced by leukemic cells established from NIH Swiss mice with radiation-induced leukemia

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Okumoto, Masaaki; Nishikawa, Ryosuke; Takamori, Yasuhiko; Iwai, Yoshiaki; Iwai, Mineko [Radiation Center of Osaka Prefecture, Sakai (Japan); Imai, Shunsuke; Morimoto, Junji; Tsubura, Yoshihiko

1984-06-01

Three leukemic cell lines, designated NIH-RL1, NIH-RL2 and NFS-RL1, were established from spleen and thymuses of NIH Swiss and NFS mice with radiation-induced leukemia. The culture fluids of these cell lines contained RNA-dependent DNA polymerase (RDDP) activities associated with particles of buoyant density of 1.15-1.17 (g/cm/sup 3/). The divalent cation reqirement of these enzymes was characteristic for that of murine leukemia viruses. In competition radioimmunoassay, a major core protein, p30, was detected in culture fluid of each leukemic cell line. Competition curves of viral p30 produced by these cell lines revealed that these viruses were very similar to those of xenotropic viruses of NZB mice. These viruses were undetectable both by XC plaque assay using SC-1 cells as an indicator cell, and by mink S/sup +/L/sup -/ focus induction assay. These viruses also lacked productive infectivity to mink lung cells (CCL-64), and were nononcogenic in syngeneic mice when the viruses were intrathymically inoculated.

Diet-induced obesity in mice reduces the maintenance of influenza-specific CD8+ memory T cells.

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Karlsson, Erik A; Sheridan, Patricia A; Beck, Melinda A

2010-09-01

Obesity has been associated with increasing the risk for type 2 diabetes and heart disease, but its influence on the immune response to viral infection is understudied. Memory T cells generated during a primary influenza infection are important for protection against subsequent influenza exposures. Previously, we have demonstrated that diet-induced obese (DIO) mice have increased morbidity and mortality following secondary influenza infection compared with lean mice. To determine whether the problem resided in a failure to maintain functional, influenza-specific CD8(+) memory T cells, male DIO and lean mice were infected with influenza X-31. At 84 d postinfection, DIO mice had a 10% reduction in memory T cell numbers. This reduction may have resulted from significantly reduced memory T cell expression of interleukin 2 receptor beta (IL-2R beta, CD122), but not IL-7 receptor alpha (CD127), which are both required for memory cell maintenance. Peripheral leptin resistance in the DIO mice may be a contributing factor to the impairment. Indeed, leptin receptor mRNA expression was significantly reduced in the lungs of obese mice, whereas suppressor of cytokine signaling (Socs)1 and Socs3 mRNA expression were increased. It is imperative to understand how the obese state alters memory T cells, because impairment in maintenance of functional memory responses has important implications for vaccine efficacy in an obese population.

Forebrain-Specific Loss of BMPRII in Mice Reduces Anxiety and Increases Object Exploration.

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McBrayer, Zofeyah L; Dimova, Jiva; Pisansky, Marc T; Sun, Mu; Beppu, Hideyuki; Gewirtz, Jonathan C; O'Connor, Michael B

2015-01-01

To investigate the role of Bone Morphogenic Protein Receptor Type II (BMPRII) in learning, memory, and exploratory behavior in mice, a tissue-specific knockout of BMPRII in the post-natal hippocampus and forebrain was generated. We found that BMPRII mutant mice had normal spatial learning and memory in the Morris water maze, but showed significantly reduced swimming speeds with increased floating behavior. Further analysis using the Porsolt Swim Test to investigate behavioral despair did not reveal any differences in immobility between mutants and controls. In the Elevated Plus Maze, BMPRII mutants and Smad4 mutants showed reduced anxiety, while in exploratory tests, BMPRII mutants showed more interest in object exploration. These results suggest that loss of BMPRII in the mouse hippocampus and forebrain does not disrupt spatial learning and memory encoding, but instead impacts exploratory and anxiety-related behaviors.

Forebrain-Specific Loss of BMPRII in Mice Reduces Anxiety and Increases Object Exploration.

Directory of Open Access Journals (Sweden)

Zofeyah L McBrayer

Full Text Available To investigate the role of Bone Morphogenic Protein Receptor Type II (BMPRII in learning, memory, and exploratory behavior in mice, a tissue-specific knockout of BMPRII in the post-natal hippocampus and forebrain was generated. We found that BMPRII mutant mice had normal spatial learning and memory in the Morris water maze, but showed significantly reduced swimming speeds with increased floating behavior. Further analysis using the Porsolt Swim Test to investigate behavioral despair did not reveal any differences in immobility between mutants and controls. In the Elevated Plus Maze, BMPRII mutants and Smad4 mutants showed reduced anxiety, while in exploratory tests, BMPRII mutants showed more interest in object exploration. These results suggest that loss of BMPRII in the mouse hippocampus and forebrain does not disrupt spatial learning and memory encoding, but instead impacts exploratory and anxiety-related behaviors.

Caffeine/sleep-deprivation interaction in mice produces complex memory effects.

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Onaolapo, Olakunle J; Onaolapo, Adejoke Y; Akanmu, Moses A; Olayiwola, Gbola

2015-07-01

Sleep deprivation negatively impacts memory, causing deficits in memory processes. Of interest is any agent that can offset such deficits. Mice were given varying doses of caffeine for 14 days and then deprived of sleep for 6 hours by the 'gentle handling' method. Memory was assessed using the Novel Object Recognition Test and Y maze alternation. The study was designed to ascertain the impact of varying doses of caffeine combined with total sleep-deprivation on spatial and non spatial memory in mice. Adult Swiss Webster mice of both sexes were assigned to six groups viz., vehicle (distilled water), or one of five selected doses of caffeine (10, 20, 40, 80 and 120 mg/kg) for 14 days via the oral route. Open field novel object recognition test and Y maze spatial working memory tests were carried out on day 14. Results were analysed using multi-factorial ANOVA followed by Tukey HSD test and expressed as mean ± S.E.M, with p values less than 0.05 were considered statistically significant. Novel object recognition tests (NOR) revealed that pre-training and pre-test sleep deprivation and caffeine combination impaired non spatial and spatial memory in male and female mice. The study shows the complex interactions with memory that may arise when total sleep deprivation is superimposed on caffeine administration.

Modulation of ovomucoid-specific oral tolerance in mice fed plant extracts containing lectins

DEFF Research Database (Denmark)

Kjær, Tanja; Frøkiær, Hanne

2002-01-01

We investigated the effect of feeding extracts of four different legumes (red kidney bean (Phaseolus vulgaris), peanut (Arachis hypogaea), soyabean (Glycine max) and pea (Pisum sativum) on the specific immune response against a food protein. Mice were fed ovomucoid and the specific immune response...... influenced the immune response against ovomucoid; however, this was not as pronounced as for kidney bean and was only significant (Ppea extract was fed and peanut extract had a non-significant effect on induction of oral tolerance...... and on the general immune response. Plasma antibodies against kidney-bean lectin, but not against the three other legume lectins, were detected. Our current findings show that other dietary components can influence the specific immune response against food proteins. Various dietary components may thus contribute...

Sex-specific differences in hyperoxic lung injury in mice: Implications for acute and chronic lung disease in humans

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Lingappan, Krithika, E-mail: lingappa@bcm.edu [Department of Pediatrics, Section of Neonatology, Texas Children' s Hospital, Baylor College of Medicine, 1102 Bates Avenue, MC: FC530.01, Houston, TX 77030 (United States); Jiang, Weiwu; Wang, Lihua; Couroucli, Xanthi I. [Department of Pediatrics, Section of Neonatology, Texas Children' s Hospital, Baylor College of Medicine, 1102 Bates Avenue, MC: FC530.01, Houston, TX 77030 (United States); Barrios, Roberto [Department of Pathology and Genomic Medicine, The Methodist Hospital Physician Organization, 6565 Fannin Street, Suite M227, Houston, TX 77030 (United States); Moorthy, Bhagavatula [Department of Pediatrics, Section of Neonatology, Texas Children' s Hospital, Baylor College of Medicine, 1102 Bates Avenue, MC: FC530.01, Houston, TX 77030 (United States)

2013-10-15

Sex-specific differences in pulmonary morbidity in humans are well documented. Hyperoxia contributes to lung injury in experimental animals and humans. The mechanisms responsible for sex differences in the susceptibility towards hyperoxic lung injury remain largely unknown. In this investigation, we tested the hypothesis that mice will display sex-specific differences in hyperoxic lung injury. Eight week-old male and female mice (C57BL/6J) were exposed to 72 h of hyperoxia (FiO{sub 2} > 0.95). After exposure to hyperoxia, lung injury, levels of 8-iso-prostaglandin F{sub 2} alpha (8-iso-PGF 2α) (LC–MS/MS), apoptosis (TUNEL) and inflammatory markers (suspension bead array) were determined. Cytochrome P450 (CYP)1A expression in the lung was assessed using immunohistochemistry and western blotting. After exposure to hyperoxia, males showed greater lung injury, neutrophil infiltration and apoptosis, compared to air-breathing controls than females. Pulmonary 8-iso-PGF 2α levels were higher in males than females after hyperoxia exposure. Sexually dimorphic increases in levels of IL-6 (F > M) and VEGF (M > F) in the lungs were also observed. CYP1A1 expression in the lung was higher in female mice compared to males under hyperoxic conditions. Overall, our results support the hypothesis that male mice are more susceptible than females to hyperoxic lung injury and that differences in inflammatory and oxidative stress markers contribute to these sex-specific dimorphic effects. In conclusion, this paper describes the establishment of an animal model that shows sex differences in hyperoxic lung injury in a temporal manner and thus has important implications for lung diseases mediated by hyperoxia in humans. - Highlights: • Male mice were more susceptible to hyperoxic lung injury than females. • Sex differences in inflammatory markers were observed. • CYP1A expression was higher in females after hyperoxia exposure.

Sex-specific differences in hyperoxic lung injury in mice: Implications for acute and chronic lung disease in humans

International Nuclear Information System (INIS)

Lingappan, Krithika; Jiang, Weiwu; Wang, Lihua; Couroucli, Xanthi I.; Barrios, Roberto; Moorthy, Bhagavatula

2013-01-01

Sex-specific differences in pulmonary morbidity in humans are well documented. Hyperoxia contributes to lung injury in experimental animals and humans. The mechanisms responsible for sex differences in the susceptibility towards hyperoxic lung injury remain largely unknown. In this investigation, we tested the hypothesis that mice will display sex-specific differences in hyperoxic lung injury. Eight week-old male and female mice (C57BL/6J) were exposed to 72 h of hyperoxia (FiO 2 > 0.95). After exposure to hyperoxia, lung injury, levels of 8-iso-prostaglandin F 2 alpha (8-iso-PGF 2α) (LC–MS/MS), apoptosis (TUNEL) and inflammatory markers (suspension bead array) were determined. Cytochrome P450 (CYP)1A expression in the lung was assessed using immunohistochemistry and western blotting. After exposure to hyperoxia, males showed greater lung injury, neutrophil infiltration and apoptosis, compared to air-breathing controls than females. Pulmonary 8-iso-PGF 2α levels were higher in males than females after hyperoxia exposure. Sexually dimorphic increases in levels of IL-6 (F > M) and VEGF (M > F) in the lungs were also observed. CYP1A1 expression in the lung was higher in female mice compared to males under hyperoxic conditions. Overall, our results support the hypothesis that male mice are more susceptible than females to hyperoxic lung injury and that differences in inflammatory and oxidative stress markers contribute to these sex-specific dimorphic effects. In conclusion, this paper describes the establishment of an animal model that shows sex differences in hyperoxic lung injury in a temporal manner and thus has important implications for lung diseases mediated by hyperoxia in humans. - Highlights: • Male mice were more susceptible to hyperoxic lung injury than females. • Sex differences in inflammatory markers were observed. • CYP1A expression was higher in females after hyperoxia exposure

Gender-specific effects of endogenous testosterone: female alpha-estrogen receptor-deficient C57Bl/6J mice develop glomerulosclerosis.

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Elliot, S J; Berho, M; Korach, K; Doublier, S; Lupia, E; Striker, G E; Karl, M

2007-08-01

Young female mice on a C57Bl/6J (B6) background are considered glomerulosclerosis (GS)-resistant but aging B6 mice develop mild GS. Estrogen deficiency accelerates while estrogen replacement retards GS in young sclerosis-prone oligosyndactyly mutant mice on an ROP background. To explore the effects of sex hormones on glomerular structure and function in the context of gender and genetic background, we studied mice in which the estrogen-receptor (ER) genes alpha- or -beta were deleted (alpha- or betaER knockout (KO)) and crossed into the B6 background. We also studied ovariectomized (Ovx) B6 mice given testosterone. Male and female betaERKO and male alphaERKO mice had no glomerular dysfunction at 9 months of age; however, alphaERKO female mice displayed albuminuria and GS. Ovx prevented glomerular dysfunction in alphaERKO female mice by eliminating endogenous testosterone production while exogenous testosterone induced GS in Ovx B6 mice. Androgen receptor (AR) expression and function was found in microdissected glomeruli and cultured mesangial cells. Testosterone compared to placebo increased both AR expression and TGF-beta1 mRNA levels in glomeruli isolated from female B6 mice. Estrogen deficiency had no deleterious effects on the glomeruli in B6 mice. Our study shows that genetic traits strongly influence the GS-promoting effects of estrogen deficiency while testosterone induces GS in a gender-specific manner.

Effect of blood transfusion and cyclophosphamide on cardiac allograft survival in sensitized mice

International Nuclear Information System (INIS)

Lasek, Witold; Sora, Magdalena; Jakobisiak, Marek

1994-01-01

In the H-2-incompatible donor-recipient model in mice (BALB/c → CBA/H), combination of donor-specific blood transfusion (DST) on the day -9 before transplantation with both pre- and post transplant immunosuppression with cyclophosphamide (CY) interacted beneficially to produce significant donor-specific prolongation of cardiac graft survival. However, in recipients presensitized with donor-specific blood on the day -21, combination of DST with pre- and post transplant CY immunosuppression did not act synergistically and graft survival in this group did not differ from that in presentized mice treated with 2 doses of CY alone. (author). 21 refs, 1 fig., 3 tabs

Development of hepatocellular adenomas and carcinomas in mice with liver-specific G6Pase-α deficiency

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Roberta Resaz

2014-09-01

Full Text Available Glycogen storage disease type 1a (GSD-1a is caused by a deficiency in glucose-6-phosphatase-α (G6Pase-α, and is characterized by impaired glucose homeostasis and a high risk of developing hepatocellular adenomas (HCAs. A globally G6Pase-α-deficient (G6pc−/− mouse model that shows pathological features similar to those of humans with GSD-1a has been developed. These mice show a very severe phenotype of disturbed glucose homeostasis and rarely live beyond weaning. We generated liver-specific G6Pase-α-deficient (LS‑G6pc−/− mice as an alternative animal model for studying the long-term pathophysiology of the liver and the potential treatment strategies, such as cell therapy. LS‑G6pc−/− mice were viable and exhibited normal glucose profiles in the fed state, but showed significantly lower blood glucose levels than their control littermates after 6 hours of fasting. LS‑G6pc−/− mice developed hepatomegaly with glycogen accumulation and hepatic steatosis, and progressive hepatic degeneration. Ninety percent of the mice analyzed developed amyloidosis by 12 months of age. Finally, 25% of the mice sacrificed at age 10–20 months showed the presence of multiple HCAs and in one case late development of hepatocellular carcinoma (HCC. In conclusion, LS‑G6pc−/− mice manifest hepatic symptoms similar to those of human GSD-1a and, therefore, represent a valid model to evaluate long-term liver pathogenesis of GSD-1a.

Idiopathic paraproteinemia. II. Transplantation of the paraprotein- producing clone from old to young C57BL/KaLwRij mice

NARCIS (Netherlands)

Radl, J.; Glopper, E.de; Schuit, H.R.E.; Zurcher, C.

1979-01-01

Transplantation experiments in the C57BL/KaLwRij mouse model of idiopathic paraproteinemia (IP) showed that an IP-producing clone can be further propagated in young, lethally irradiated mice and also equally as well in nonirradiated recipients by a bone marrow and/or spleen cell transfer. The

Treatment with anti-IL-6 receptor antibody prevented increase in serum hepcidin levels and improved anemia in mice inoculated with IL-6–producing lung carcinoma cells

International Nuclear Information System (INIS)

Noguchi-Sasaki, Mariko; Sasaki, Yusuke; Shimonaka, Yasushi; Mori, Kazushige; Fujimoto-Ouchi, Kaori

2016-01-01

Hepcidin, a key regulator of iron metabolism, is produced mainly by interleukin-6 (IL-6) during inflammation. A mechanism linking cancer-related anemia and IL-6 through hepcidin production is suggested. To clarify the hypothesis that overproduction of IL-6 elevates hepcidin levels and contributes to the development of cancer-related anemia, we evaluated anti-IL-6 receptor antibody treatment of cancer-related anemia in an IL-6–producing human lung cancer xenograft model. Nude mice were subcutaneously inoculated with cells of the IL-6–producing human lung cancer cell line LC-06-JCK and assessed as a model of cancer-related anemia. Mice bearing LC-06-JCK were administered rat anti-mouse IL-6 receptor antibody MR16-1 and their serum hepcidin levels and hematological parameters were determined. LC-06-JCK–bearing mice developed anemia according to the production of human IL-6 from xenografts, with decreased values of hemoglobin, hematocrit, and mean corpuscular volume (MCV) compared to non–tumor-bearing (NTB) mice. LC-06-JCK–bearing mice showed decreased body weight and serum albumin with increased serum amyloid A. MR16-1 treatment showed significant inhibition of decreased body weight and serum albumin levels, and suppressed serum amyloid A level. There was no difference in tumor volume between MR16-1-treated mice and immunoglobulin G (IgG)-treated control mice. Decreased hemoglobin, hematocrit, and MCV in LC-06-JCK–bearing mice was significantly relieved by MR16-1 treatment. LC-06-JCK–bearing mice showed high red blood cell counts and erythropoietin levels as compared to NTB mice, whereas MR16-1 treatment did not affect their levels. Serum hepcidin and ferritin levels were statistically elevated in mice bearing LC-06-JCK. LC-06-JCK–bearing mice showed lower values of MCV, mean corpuscular hemoglobin (MCH), and serum iron as compared to NTB mice. Administration of MR16-1 to mice bearing LC-06-JCK significantly suppressed levels of both serum hepcidin and

Liver-specific Aquaporin 11 knockout mice show rapid vacuolization of the rough endoplasmic reticulum in periportal hepatocytes after amino acid feeding

DEFF Research Database (Denmark)

Rojek, Aleksandra; Füchtbauer, Ernst-Martin; Füchtbauer, Annette C.

2013-01-01

-specific Aqp11 KO mice, allowing us to study the role of AQP11 protein in liver of mice with normal kidney function. The unchallenged liver-specific Aqp11 KO mice have normal longevity, their livers appeared normal, and the plasma biochemistries revealed only a minor defect in lipid handling. Fasting......Aquaporin 11 (AQP11) is a protein channel expressed intracellularly in multiple organs, yet its physiological function is unclear. Aqp11 knockout (KO) mice die early due to malfunction of the kidney, a result of hydropic degeneration of proximal tubule cells. Here we report the generation of liver...... protein or larger doses of various amino acids. The fasting/refeeding challenge is associated with increased expression of markers of ER stress Grp78 and GADD153 and decreased glutathione levels, suggesting that ER stress may play role in the development of vacuoles in the AQP11-deficient hepatocytes. NMR...

Epitope mapping of the alpha-chain of the insulin-like growth factor I receptor using antipeptide antibodies.

Science.gov (United States)

Delafontaine, P; Ku, L; Ververis, J J; Cohen, C; Runge, M S; Alexander, R W

1994-12-01

Insulin-like growth factor I (IGF I) is an important mitogen for vascular smooth muscle cells (VSMC). The IGF I receptor (IGF IR) is a heterotetramer composed of two cross-linked extracellular alpha-chains and two membrane-spanning beta-chains that contain a tyrosine-kinase domain. It has a high degree of sequence similarity to the insulin receptor (IR), and the putative ligand-specific binding site has been localized to a cysteine-rich region (CRR) of the alpha-chain. To obtain insights into antigenic determinants of the IGF IR, we raised a panel of site-specific polyclonal antibodies against short peptide sequences N-terminal to and within the CRR. Several antibodies raised against linear epitopes within the CRR bound to solubilized and native rat and human IGF IR by ELISA, did not cross-react with IR, but unexpectedly failed to inhibit 125I-IGF I binding. A polyclonal antibody directed against a 48-amino acid synthetic peptide, corresponding to a region of the CRR postulated to be essential for ligand binding, failed to react with either solubilized, reduced or intact IGF IR. Three antibodies specific for the N-terminus of the alpha-chain reacted with solubilized and native IGF IR. One of these, RAB 6, directed against amino acids 38-44 of the IGF IR, inhibited 125I-IGF I binding to rat aortic smooth muscle cells (RASM) and to IGF IR/3T3 cells (overexpressing human IGF IR) by up to 45%. Immunohistochemical analysis revealed strong IGF IR staining in the medial smooth muscle cell layer of rat aorta. These findings are consistent with a model wherein conformational epitopes within the CRR and linear epitopes within the N-terminus of the alpha-chain contribute to the IGF I binding pocket. These antibodies should provide a valuable tool to study structure-function relationships and in vivo regulation of the IGF IR.

Effect of tumour mass animal gender on specific uptake of [99Tc] pertechnetate in the EMT6 tumor in mice

International Nuclear Information System (INIS)

Maddalena, D.J.; Snowdon, G.M.

1990-01-01

The effects of tumour mass and animal gender on tumour uptake of [ 99m Tc] pertechnetate were examined in balb/c mice bearing an EMT-6 tumour showing high affinity for pertechnetate. The injected dose per gram found in the tumours appears to be inversely related to the tumour mass with good correlation suggesting a high specific uptake. The tumours grown in female mice had greater uptakes and rates of uptake than those grown in the male mice. Studies carried out in animals treated with perchlorate to block active anion transport showed low pertechnetate uptake into the tumours suggesting that the high affinity of 99m TcO 4 - for the tumour was due to a specific 99m TcO 4 - transport mechanism. 15 refs., 3 tabs., 3 figs

Prenatal stress challenge impairs fetal lung development and asthma severity sex-specifically in mice.

Science.gov (United States)

Zazara, Dimitra E; Perani, Clara V; Solano, María E; Arck, Petra C

2018-02-01

Allergic asthma is an increasing health problem worldwide. Interestingly, prenatal challenges such as stress have been associated with an increased risk for asthma during childhood. The underlying pathogenesis of how prenatal stress increases the risk for asthma still remains unclear. Potential targets could be that the fetal immune ontogeny or fetal lung development are compromised by prenatal challenges. Here, we aimed to identify whether prenatal stress challenge affects fetal lung development in mice. C57BL/6 pregnant mice were challenged with sound stress and fetal lung development was assessed histologically. Whilst prenatal stress challenge did not profoundly affect lung development in male fetuses, it resulted in less extensive terminal sacs, surrounded by thicker mesenchymal tissue in female fetuses. Thus, prenatal stress disrupted fetal lung development sex-specifically. Interestingly, upon prenatal stress challenge, the airway hyperresponsiveness and eosinophilic inflammation- two hallmarks of asthma - were significantly increased in adult female offspring, whilst regulatory CD4+ T cells were reduced. These findings strongly underpin the sex-specific association between s challenged fetal development and a sex-specific altered severity of asthma in adult offspring. Our model now allows to identify maternal markers through which the risk for asthma and possible other diseases is vertically transferred before birth in response to challenges. Such identification then opens avenues for primary disease prevention. Copyright © 2017 Elsevier B.V. All rights reserved.

Tissue-specific down-regulation of RIPK 2 in Mycobacterium leprae-infected nu/nu mice

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Gue-Tae Chae

1992-01-01

Full Text Available RIPK 2 is adapter molecule in the signal pathway involved in Toll-like receptors. However, there has been no reported association between receptor-interacting serine/threonine kinase 2 (RIPK 2 expression and the infectious diseases involving mycobacterial infection. This study found that its expression was down-regulated in the footpads and skin but was up-regulated in the liver of Mycobacterium leprae-infected nu/nu mice compared with those of the M. leprae non-infected nu/nu mice. It was observed that the interlukin-12p40 and interferon-γ genes involved in the susceptibility of M. leprae were down-regulated in the skin but were up-regulated in the liver. Overall, this suggests that regulation of RIPK 2 expression is tissue-specifically associated with M. leprae infection.

Obligatory Role of EP1 Receptors in the Increase in Cerebral Blood Flow Produced by Hypercapnia in the Mice.

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Ken Uekawa

Full Text Available Hypercapnia induces potent vasodilation in the cerebral circulation. Although it has long been known that prostanoids participate in the cerebrovascular effects of hypercapnia, the role of prostaglandin E2 (PGE2 and PGE2 receptors have not been fully investigated. In this study, we sought to determine whether cyclooxygenase-1 (COX-1-derived PGE2 and EP1 receptors are involved in the cerebrovascular response induced by hypercapnia. Cerebral blood flow (CBF was recorded by laser-Doppler flowmetry in the somatosenasory cortex of anesthetized male EP1-/- mice and wild type (WT littermates. In WT mice, neocortical application of the EP1 receptor antagonist SC-51089 attenuated the increase in CBF elicited by systemic hypercapnia (pCO2 = 50-60 mmHg. SC-51089 also attenuated the increase in CBF produced by neocortical treatment of arachidonic acid or PGE2. These CBF responses were also attenuated in EP1-/- mice. In WT mice, the COX-1 inhibitor SC-560, but not the COX-2 inhibitor NS-398, attenuated the hypercapnic CBF increase. Neocortical application of exogenous PGE2 restored the attenuation in resting CBF and the hypercapnic response induced by SC-560. In contrast, exogenous PGE2 failed to rescue the attenuation both in WT mice induced by SC-51089 and EP1-/- mice, attesting to the obligatory role of EP1 receptors in the response. These findings indicate that the hypercapnic vasodilatation depends on COX-1-derived PGE2 acting on EP1 receptors and highlight the critical role that COX-1-derived PGE2 and EP1 receptors play in the hypercapnic regulation of the cerebral circulation.

Patent ductus arteriosus in mice with smooth muscle-specific Jag1 deletion

Science.gov (United States)

Feng, Xuesong; Krebs, Luke T.; Gridley, Thomas

2010-01-01

The ductus arteriosus is an arterial vessel that shunts blood flow away from the lungs during fetal life, but normally occludes after birth to establish the adult circulation pattern. Failure of the ductus arteriosus to close after birth is termed patent ductus arteriosus and is one of the most common congenital heart defects. Mice with smooth muscle cell-specific deletion of Jag1, which encodes a Notch ligand, die postnatally from patent ductus arteriosus. These mice exhibit defects in contractile smooth muscle cell differentiation in the vascular wall of the ductus arteriosus and adjacent descending aorta. These defects arise through an inability to propagate the JAG1-Notch signal via lateral induction throughout the width of the vascular wall. Both heterotypic endothelial smooth muscle cell interactions and homotypic vascular smooth muscle cell interactions are required for normal patterning and differentiation of the ductus arteriosus and adjacent descending aorta. This new model for a common congenital heart defect provides novel insights into the genetic programs that underlie ductus arteriosus development and closure. PMID:21068062

78 FR 35318 - Solicitation for a Cooperative Agreement-Corrections-Specific Resources for Extended Stakeholder...

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2013-06-12

... research. In addition, expertise in some aspect of media production (Web, mobile, broadcast, social media...). Registration in the CRR can be done online at the CCR Web site: http://www.bpn.gov/ccr . A CCR Handbook and... individual have the organizational capacity to achieve all project tasks? Are the proposed project management...

No beneficial effect of general and specific anti-inflammatory therapies on aortic dilatation in Marfan mice.

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Romy Franken

Full Text Available AIMS: Patients with Marfan syndrome have an increased risk of life-threatening aortic complications, mostly preceded by aortic dilatation. In the FBN1(C1039G/+ Marfan mouse model, losartan decreases aortic root dilatation. We recently confirmed this beneficial effect of losartan in adult patients with Marfan syndrome. The straightforward translation of this mouse model to man is reassuring to test novel treatment strategies. A number of studies have shown signs of inflammation in aortic tissue of Marfan patients. This study examined the efficacy of anti-inflammatory therapies in attenuating aortic root dilation in Marfan syndrome and compared effects to the main preventative agent, losartan. METHODS AND RESULTS: To inhibit inflammation in FBN1(C1039G/+ Marfan mice, we treated the mice with losartan (angiotensin II receptor type 1 inhibitor, methylprednisolone (corticosteroid or abatacept (T-cell-specific inhibitor. Treatment was initiated in adult Marfan mice with already existing aortic root dilatation, and applied for eight weeks. Methylprednisolone- or abatacept-treated mice did not reveal a reduction in aortic root dilatation. In this short time frame, losartan was the only treatment that significantly reduced aorta inflammation, transforming growth factor-beta (TGF-β signaling and aortic root dilatation rate in these adult Marfan mice. Moreover, the methylprednisolone-treated mice had significantly more aortic alcian blue staining as a marker for aortic damage. CONCLUSION: Anti-inflammatory agents do not reduce the aortic dilatation rate in Marfan mice, but possibly increase aortic damage. Currently, the most promising therapeutic drug in Marfan syndrome is losartan, by blocking the angiotensin II receptor type 1 and thereby inhibiting pSmad2 signaling.

Gender-specific induction of cytochrome P450s in nonylphenol-treated FVB/NJ mice

International Nuclear Information System (INIS)

Hernandez, Juan P.; Chapman, Laura M.; Kretschmer, Xiomara C.; Baldwin, William S.

2006-01-01

Nonylphenol (NP) is a breakdown product of nonylphenol ethoxylates, which are used in a variety of industrial, agricultural, household cleaning, and beauty products. NP is one of the most commonly found toxicants in the United States and Europe and is considered a toxicant of concern because of its long half-life. NP is an environmental estrogen that also activates the pregnane X-receptor (PXR) and in turn induces P450s. No study to date has examined the gender-specific effects of NP on hepatic P450 expression. We provided NP at 0, 50 or 75 mg/kg/day for 7 days to male and female FVB/NJ mice and compared their P450 expression profiles. Q-PCR was performed on hepatic cDNA using primers to several CYP isoforms regulated by PXR or its relative, the constitutive androstane receptor (CAR). In female mice, NP induced Cyp2b10 and Cyp2b13, and downregulated the female-specific P450s, Cyp3a41 and Cyp3a44. In contrast, male mice treated with NP showed increased expression of Cyp2a4, Cyp2b9, and Cyp2b10. Western blots confirmed induction of Cyp2b subfamily members in both males and females. Consistent with the Q-PCR data, Western blots showed dose-dependent downregulation of Cyp3a only in females and induction of Cyp2a only in males. The overall increase in female-predominant P450s in males (Cyp2a4, 2b9) and the decrease in female-predominant P450s in females (Cyp3a41, 3a44) suggest that NP is in part feminizing the P450 profile in males and masculinizing the P450 profile in females. Testosterone hydroxylation was also altered in a gender-specific manner, as testosterone 16α-hydroxylase activity was only induced in NP-treated males. In contrast, NP-treated females demonstrated a greater propensity for metabolizing zoxazolamine probably due to greater Cyp2b induction in females. In conclusion, NP causes gender-specific P450 induction and therefore exposure to NP may cause distinct pharmacological and toxicological effects in males compared to females

Distinctive cellular immunity in genetically susceptible BALB/c mice recovered from Leishmania major infection or after subcutaneous immunization with killed parasites

International Nuclear Information System (INIS)

Liew, F.Y.; Dhaliwal, J.S.

1987-01-01

Genetically susceptible BALB/c mice are refractory to further infection after recovery from Leishmania major infection after a sublethal dose of gamma-irradiation. In contrast, mice immunized with killed promastigotes s.c. develop exacerbated lesions after infection. Both groups of mice produce only a low level of specific antibody and no detectable cytotoxic T cells, but do have a strong antigen-specific DTH, which is adoptively transferable with Lyt-1+2-, L3T4+ T cells. Kinetic and histological studies revealed that mice immunized s.c. developed Jones-Mote hypersensitivity, peaking at 15 hr. with little mononuclear cell infiltration at the site of antigen administration; whereas mice that had recovered from infection developed tuberculin-type of reactivity, peaking at 24 to 48 hr, with intense mononuclear cell infiltration. Splenic T cells from recovered mice, when injected into the footpads of normal recipients together with live promastigotes, were able to retard lesion development; whereas T cells from s.c. immunized mice, when similarly transferred, accelerated disease progression. Antigen-specific culture supernatant of spleen cells from recovered mice also activated normal resident peritoneal macrophages to kill intracellular L. major amastigotes and tumor cells. Culture supernatants of spleen cells from s.c. immunized or normal mice were devoid of such activities. Part of the macrophage-activating potential can be inhibited by antibody specific for IFN-gamma. These results therefore demonstrate that whereas the Jones-Mote reaction is correlated with disease exacerbation, the tuberculin-type of DTH may be protective. Furthermore, in vivo immunity is directly related to the capacity of T cells to produce macrophage-activating factor

Schisandra chinensis produces the antidepressant-like effects in repeated corticosterone-induced mice via the BDNF/TrkB/CREB signaling pathway.

Science.gov (United States)

Yan, Tingxu; Xu, Mengjie; Wan, Shutong; Wang, Mengshi; Wu, Bo; Xiao, Feng; Bi, Kaishun; Jia, Ying

2016-09-30

The present study aimed to examine the antidepressant-like effects and the possible mechanisms of Schisandra chinensis on depressive-like behavior induced by repeated corticosterone injections in mice. Here we evaluated the effect of an ethanol extract of the dried fruit of S. chinensis (EESC) on BDNF/TrkB/CREB signaling in the hippocampus and the prefrontal cortex. Three weeks of corticosterone injections in mice resulted in depressive-like behavior, as indicated by the significant decrease in sucrose consumption and increase the immobility time in the forced swim test, but without any influence on the locomotor activity. Further, there was a significant increase in serum corticosterone level and a significant downregulation of BDNF/TrkB/CREB signaling pathway in the hippocampus and prefrontal cortex in CORT-treated mice. Treatment of mice with EESC (600mg/kg) significantly ameliorated all the behavioral and biochemical changes induced by corticosterone. Moreover, pharmacological inhibition of BDNF signaling by K252a abolished entirely the antidepressant-like effect triggered by chronic EESC treatment. These results suggest that EESC produces an antidepressant-like effect in CORT-induced depression in mice, which is possibly mediated, at least in part, by rectifying the stress-based hypothalamic-pituitary-adrenal (HPA) axis dysfunction paradigm and upregulation of BDNF/TrkB/CREB signaling pathway. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Contrasting effects of exercise and NOS inhibition on tissue-specific fatty acid and glucose uptake in mice.

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Rottman, Jeffrey N; Bracy, Deanna; Malabanan, Carlo; Yue, Zou; Clanton, Jeff; Wasserman, David H

2002-07-01

Isotopic techniques were used to test the hypothesis that exercise and nitric oxide synthase (NOS) inhibition have distinct effects on tissue-specific fatty acid and glucose uptakes in a conscious, chronically catheterized mouse model. Uptakes were measured using the radioactive tracers (125)I-labeled beta-methyl-p-iodophenylpentadecanoic acid (BMIPP) and deoxy-[2-(3)H]glucose (DG) during treadmill exercise with and without inhibition of NOS. [(125)I]BMIPP uptake at rest differed substantially among tissues with the highest levels in heart. With exercise, [(125)I]BMIPP uptake increased in both heart and skeletal muscles. In sedentary mice, NOS inhibition induced by nitro-L-arginine methyl ester (L-NAME) feeding increased heart and soleus [(125)I]BMIPP uptake. In contrast, exercise, but not L-NAME feeding, resulted in increased heart and skeletal muscle [2-(3)H]DG uptake. Significant interactions were not observed in the effects of combined exercise and L-NAME feeding on [(125)I]BMIPP and [2-(3)H]DG uptakes. In the conscious mouse, exercise and NOS inhibition produce distinct patterns of tissue-specific fatty acid and glucose uptake; NOS is not required for important components of exercise-associated metabolic signaling, or other mechanisms compensate for the absence of this regulatory mechanism.

Antigen-specific murine T cell clones produce soluble interleukin 2 receptor on stimulation with specific antigens

International Nuclear Information System (INIS)

Wagner, D.K.; York-Jolley, J.; Malek, T.R.; Berzofsky, J.A.; Nelson, D.L.

1986-01-01

In this study, monoclonal antibodies were used to the murine IL 2 receptor (IL 2R) termed 3C7 and 7D4, which bind to different epitopes on the murine IL 2R, to develop an ELISA to measure soluble murine IL 2R. Surprisingly, stimulated murine spleen cells not only expressed cell-associated IL 2R, but also produced a considerable level of cellfree IL 2R in the culture supernatant fluid. To assess the fine specificity of this response, myoglobin-immune murine T cell clones were stimulated with appropriate or inappropriate antigen and syngeneic or allogeneic presenting cells. Proliferation, measured by [ 3 H] thymidine incorporation, and levels of soluble IL 2R were determined at day 4. The production of soluble IL2R displayed the same epitope fine specificity, genetic restriction, and antigen dose-response as the proliferative response. Indeed, in some cases there was sharper discrimination of epitope specificity and genetic restriction with the soluble IL 2R levels. There was also reproducible clone-to-clone variation in the amount of soluble receptor produced in response to antigen among 12 T cell clones and lines tested. In time course experiments, proliferation was greatest at day 3, whereas soluble IL 2R levels continued to rise in subsequent days. To the authors' knowledge, this is the first demonstration of release of secretion of soluble IL 2R by murine T cells, and the first demonstration of the fine specificity and genetic restriction of the induction of soluble IL 2R by specific antigen

Tissue-Specific Expression of Monocarboxylate Transporters during Fasting in Mice

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Schutkowski, Alexandra; Wege, Nicole; Stangl, Gabriele I.; König, Bettina

2014-01-01

Monocarboxylates such as pyruvate, lactate and ketone bodies are crucial for energy supply of all tissues, especially during energy restriction. The transport of monocarboxylates across the plasma membrane of cells is mediated by monocarboxylate transporters (MCTs). Out of 14 known mammalian MCTs, six isoforms have been functionally characterized to transport monocarboxylates and short chain fatty acids (MCT1-4), thyroid hormones (MCT8, -10) and aromatic amino acids (MCT10). Knowledge on the regulation of the different MCT isoforms is rare. In an attempt to get more insights in regulation of MCT expression upon energy deprivation, we carried out a comprehensive analysis of tissue specific expression of five MCT isoforms upon 48 h of fasting in mice. Due to the crucial role of peroxisome proliferator-activated receptor (PPAR)-α as a central regulator of energy metabolism and as known regulator of MCT1 expression, we included both wildtype (WT) and PPARα knockout (KO) mice in our study. Liver, kidney, heart, small intestine, hypothalamus, pituitary gland and thyroid gland of the mice were analyzed. Here we show that the expression of all examined MCT isoforms was markedly altered by fasting compared to feeding. Expression of MCT1, MCT2 and MCT10 was either increased or decreased by fasting dependent on the analyzed tissue. MCT4 and MCT8 were down-regulated by fasting in all examined tissues. However, PPARα appeared to have a minor impact on MCT isoform regulation. Due to the fundamental role of MCTs in transport of energy providing metabolites and hormones involved in the regulation of energy homeostasis, we assumed that the observed fasting-induced adaptations of MCT expression seem to ensure an adequate energy supply of tissues during the fasting state. Since, MCT isoforms 1–4 are also necessary for the cellular uptake of drugs, the fasting-induced modifications of MCT expression have to be considered in future clinical care algorithms. PMID:25390336

Mice lacking liver-specific β-catenin develop steatohepatitis and fibrosis after iron overload.

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Preziosi, Morgan E; Singh, Sucha; Valore, Erika V; Jung, Grace; Popovic, Branimir; Poddar, Minakshi; Nagarajan, Shanmugam; Ganz, Tomas; Monga, Satdarshan P

2017-08-01

Iron overload disorders such as hereditary hemochromatosis and iron loading anemias are a common cause of morbidity from liver diseases and increase risk of hepatic fibrosis and hepatocellular carcinoma (HCC). Treatment options for iron-induced damage are limited, partly because there is lack of animal models of human disease. Therefore, we investigated the effect of iron overload in liver-specific β-catenin knockout mice (KO), which are susceptible to injury, fibrosis and tumorigenesis following chemical carcinogen exposure. Iron overload diet was administered to KO and littermate control (CON) mice for various times. To ameliorate an oxidant-mediated component of tissue injury, N-Acetyl-L-(+)-cysteine (NAC) was added to drinking water of mice on iron overload diet. KO on iron diet (KO +Fe) exhibited remarkable inflammation, followed by steatosis, oxidative stress, fibrosis, regenerating nodules and occurrence of occasional HCC. Increased injury in KO +Fe was associated with activated protein kinase B (AKT), ERK, and NF-κB, along with reappearance of β-catenin and target gene Cyp2e1, which promoted lipid peroxidation and hepatic damage. Addition of NAC to drinking water protected KO +Fe from hepatic steatosis, injury and fibrosis, and prevented activation of AKT, ERK, NF-κB and reappearance of β-catenin. The absence of hepatic β-catenin predisposes mice to hepatic injury and fibrosis following iron overload, which was reminiscent of hemochromatosis and associated with enhanced steatohepatitis and fibrosis. Disease progression was notably alleviated by antioxidant therapy, which supports its chemopreventive role in the management of chronic iron overload disorders. Lack of animal models for iron overload disorders makes it hard to study the disease process for improving therapies. Feeding high iron diet to mice that lack the β-catenin gene in liver cells led to increased inflammation followed by fat accumulation, cell death and wound healing that mimicked

Multiple factors interact to produce responses resembling spectrum of human disease in Campylobacter jejuni infected C57BL/6 IL-10-/- mice

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Wolf John E

2009-03-01

Full Text Available Abstract Background Campylobacter jejuni infection produces a spectrum of clinical presentations in humans – including asymptomatic carriage, watery diarrhea, and bloody diarrhea – and has been epidemiologically associated with subsequent autoimmune neuropathies. This microorganism is genetically variable and possesses genetic mechanisms that may contribute to variability in nature. However, relationships between genetic variation in the pathogen and variation in disease manifestation in the host are not understood. We took a comparative experimental approach to explore differences among different C. jejuni strains and studied the effect of diet on disease manifestation in an interleukin-10 deficient mouse model. Results In the comparative study, C57BL/6 interleukin-10-/- mice were infected with seven genetically distinct C. jejuni strains. Four strains colonized the mice and caused disease; one colonized with no disease; two did not colonize. A DNA:DNA microarray comparison of the strain that colonized mice without disease to C. jejuni 11168 that caused disease revealed that putative virulence determinants, including loci encoding surface structures known to be involved in C. jejuni pathogenesis, differed from or were absent in the strain that did not cause disease. In the experimental study, the five colonizing strains were passaged four times in mice. For three strains, serial passage produced increased incidence and degree of pathology and decreased time to develop pathology; disease shifted from watery to bloody diarrhea. Mice kept on an ~6% fat diet or switched from an ~12% fat diet to an ~6% fat diet just before infection with a non-adapted strain also exhibited increased incidence and severity of disease and decreased time to develop disease, although the effects of diet were only statistically significant in one experiment. Conclusion C. jejuni strain genetic background and adaptation of the strain to the host by serial passage

Adoptive transfer of EBV specific CD8+ T cell clones can transiently control EBV infection in humanized mice.

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Olga Antsiferova

2014-08-01

Full Text Available Epstein Barr virus (EBV infection expands CD8+ T cells specific for lytic antigens to high frequencies during symptomatic primary infection, and maintains these at significant numbers during persistence. Despite this, the protective function of these lytic EBV antigen-specific cytotoxic CD8+ T cells remains unclear. Here we demonstrate that lytic EBV replication does not significantly contribute to virus-induced B cell proliferation in vitro and in vivo in a mouse model with reconstituted human immune system components (huNSG mice. However, we report a trend to reduction of EBV-induced lymphoproliferation outside of lymphoid organs upon diminished lytic replication. Moreover, we could demonstrate that CD8+ T cells against the lytic EBV antigen BMLF1 can eliminate lytically replicating EBV-transformed B cells from lymphoblastoid cell lines (LCLs and in vivo, thereby transiently controlling high viremia after adoptive transfer into EBV infected huNSG mice. These findings suggest a protective function for lytic EBV antigen-specific CD8+ T cells against EBV infection and against virus-associated tumors in extra-lymphoid organs. These specificities should be explored for EBV-specific vaccine development.

D-Aspartate Modulates Nociceptive-Specific Neuron Activity and Pain Threshold in Inflammatory and Neuropathic Pain Condition in Mice

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Serena Boccella

2015-01-01

Full Text Available D-Aspartate (D-Asp is a free D-amino acid found in the mammalian brain with a temporal-dependent concentration based on the postnatal expression of its metabolizing enzyme D-aspartate oxidase (DDO. D-Asp acts as an agonist on NMDA receptors (NMDARs. Accordingly, high levels of D-Asp in knockout mice for Ddo gene (Ddo−/− or in mice treated with D-Asp increase NMDAR-dependent processes. We have here evaluated in Ddo−/− mice the effect of high levels of free D-Asp on the long-term plastic changes along the nociceptive pathway occurring in chronic and acute pain condition. We found that Ddo−/− mice show an increased evoked activity of the nociceptive specific (NS neurons of the dorsal horn of the spinal cord (L4–L6 and a significant decrease of mechanical and thermal thresholds, as compared to control mice. Moreover, Ddo gene deletion exacerbated the nocifensive responses in the formalin test and slightly reduced pain thresholds in neuropathic mice up to 7 days after chronic constriction injury. These findings suggest that the NMDAR agonist, D-Asp, may play a role in the regulation of NS neuron electrophysiological activity and behavioral responses in physiological and pathological pain conditions.

Cognitive assessment of mice strains heterozygous for cell-adhesion genes reveals strain-specific alterations in timing.

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Gallistel, C R; Tucci, Valter; Nolan, Patrick M; Schachner, Melitta; Jakovcevski, Igor; Kheifets, Aaron; Barboza, Luendro

2014-03-05

We used a fully automated system for the behavioural measurement of physiologically meaningful properties of basic mechanisms of cognition to test two strains of heterozygous mutant mice, Bfc (batface) and L1, and their wild-type littermate controls. Both of the target genes are involved in the establishment and maintenance of synapses. We find that the Bfc heterozygotes show reduced precision in their representation of interval duration, whereas the L1 heterozygotes show increased precision. These effects are functionally specific, because many other measures made on the same mice are unaffected, namely: the accuracy of matching temporal investment ratios to income ratios in a matching protocol, the rate of instrumental and classical conditioning, the latency to initiate a cued instrumental response, the trials on task and the impulsivity in a switch paradigm, the accuracy with which mice adjust timed switches to changes in the temporal constraints, the days to acquisition, and mean onset time and onset variability in the circadian anticipation of food availability.

Effects of Early-Life Stress on Social and Anxiety-Like Behaviors in Adult Mice: Sex-Specific Effects

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Natalya P. Bondar

2018-01-01

Full Text Available Stressful events in an early postnatal period have critical implications for the individual’s life and can increase later risk for psychiatric disorders. The aim of this study was to investigate the influence of early-life stress on the social behavior of adult male and female mice. C57Bl/6 mice were exposed to maternal separation (MS, 3 h once a day or handling (HD, 15 min once a day on postnatal day 2 through 14. Adult male and female mice were tested for social behavior in the social interaction test and for individual behavior in the plus-maze and open-field tests. Female mice exposed to maternal separation had increased social behavior and increased anxiety. MS male mice had no changes in social behavior but had significantly disrupted individual behavior, including locomotor and exploratory activity. Handling had positive effects on social behavior in males and females and decreased anxiety in males. Our results support the hypothesis that brief separation of pups from their mothers (handling, which can be considered as moderate stress, may result in future positive changes in behavior. Maternal separation has deleterious effects on individual behavior and significant sex-specific effects on social behavior.

Human pluripotent stem cell-derived erythropoietin-producing cells ameliorate renal anemia in mice.

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Hitomi, Hirofumi; Kasahara, Tomoko; Katagiri, Naoko; Hoshina, Azusa; Mae, Shin-Ichi; Kotaka, Maki; Toyohara, Takafumi; Rahman, Asadur; Nakano, Daisuke; Niwa, Akira; Saito, Megumu K; Nakahata, Tatsutoshi; Nishiyama, Akira; Osafune, Kenji

2017-09-27

The production of erythropoietin (EPO) by the kidneys, a principal hormone for the hematopoietic system, is reduced in patients with chronic kidney disease (CKD), eventually resulting in severe anemia. Although recombinant human EPO treatment improves anemia in patients with CKD, returning to full red blood cell production without fluctuations does not always occur. We established a method to generate EPO-producing cells from human induced pluripotent stem cells (hiPSCs) by modifying previously reported hepatic differentiation protocols. These cells showed increased EPO expression and secretion in response to low oxygen conditions, prolyl hydroxylase domain-containing enzyme inhibitors, and insulin-like growth factor 1. The EPO protein secreted from hiPSC-derived EPO-producing (hiPSC-EPO) cells induced the erythropoietic differentiation of human umbilical cord blood progenitor cells in vitro. Furthermore, transplantation of hiPSC-EPO cells into mice with CKD induced by adenine treatment improved renal anemia. Thus, hiPSC-EPO cells may be a useful tool for clarifying the mechanisms of EPO production and may be useful as a therapeutic strategy for treating renal anemia. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Characterization of an immunodominant cancer-specific O-glycopeptide epitope in murine podoplanin (OTS8)

DEFF Research Database (Denmark)

Steentoft, Catharina; Schjoldager, Katrine T; Cló, Emiliano

2010-01-01

antibody 237, developed to a spontaneous murine fibrosarcoma, was shown to be directed to murine podoplanin (OTS8) with truncated Tn O-glycans. Our understanding of such cancer-specific auto-antibodies to truncated glycoforms of glycoproteins is limited. Here we have investigated immunogenicity...... of a chemoenzymatically produced Tn-glycopeptide derived from the putative murine podoplanin O-glycopeptide epitope. We found that the Tn O-glycopeptide was highly immunogenic in mice and produced a Tn-glycoform specific response with no reactivity against unglycosylated peptides or the O-glycopeptide with extended O......-glycan (STn and T glycoforms). The immunodominant epitope was strictly dependent on the peptide sequence, required Tn at a specific single Thr residue (Thr(77)), and antibodies to the epitope were not found in naive mice. We further tested a Tn O-glycopeptide library derived from human podoplanin...

Visceral hyperalgesia induced by forebrain-specific suppression of native Kv7/KCNQ/M-current in mice

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Bian Xiling

2011-10-01

Full Text Available Abstract Background Dysfunction of brain-gut interaction is thought to underlie visceral hypersensitivity which causes unexplained abdominal pain syndromes. However, the mechanism by which alteration of brain function in the brain-gut axis influences the perception of visceral pain remains largely elusive. In this study we investigated whether altered brain activity can generate visceral hyperalgesia. Results Using a forebrain specific αCaMKII promoter, we established a line of transgenic (Tg mice expressing a dominant-negative pore mutant of the Kv7.2/KCNQ2 channel which suppresses native KCNQ/M-current and enhances forebrain neuronal excitability. Brain slice recording of hippocampal pyramidal neurons from these Tg mice confirmed the presence of hyperexcitable properties with increased firing. Behavioral evaluation of Tg mice exhibited increased sensitivity to visceral pain induced by intraperitoneal (i.p. injection of either acetic acid or magnesium sulfate, and intracolon capsaicin stimulation, but not cutaneous sensation for thermal or inflammatory pain. Immunohistological staining showed increased c-Fos expression in the somatosensory SII cortex and insular cortex of Tg mice that were injected intraperitoneally with acetic acid. To mimic the effect of cortical hyperexcitability on visceral hyperalgesia, we injected KCNQ/M channel blocker XE991 into the lateral ventricle of wild type (WT mice. Intracerebroventricular injection of XE991 resulted in increased writhes of WT mice induced by acetic acid, and this effect was reversed by co-injection of the channel opener retigabine. Conclusions Our findings provide evidence that forebrain hyperexcitability confers visceral hyperalgesia, and suppression of central hyperexcitability by activation of KCNQ/M-channel function may provide a therapeutic potential for treatment of abdominal pain syndromes.

Therapeutic Efficacy of Vectored PGT121 Gene Delivery in HIV-1-Infected Humanized Mice.

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Badamchi-Zadeh, Alexander; Tartaglia, Lawrence J; Abbink, Peter; Bricault, Christine A; Liu, Po-Ting; Boyd, Michael; Kirilova, Marinela; Mercado, Noe B; Nanayakkara, Ovini S; Vrbanac, Vladimir D; Tager, Andrew M; Larocca, Rafael A; Seaman, Michael S; Barouch, Dan H

2018-04-01

Broadly neutralizing antibodies (bNAbs) are being explored for HIV-1 prevention and cure strategies. However, administration of purified bNAbs poses challenges in resource-poor settings, where the HIV-1 disease burden is greatest. In vivo vector-based production of bNAbs represents an alternative strategy. We investigated adenovirus serotype 5 (Ad5) and adeno-associated virus serotype 1 (AAV1) vectors to deliver the HIV-1-specific bNAb PGT121 in wild-type and immunocompromised C57BL/6 mice as well as in HIV-1-infected bone marrow-liver-thymus (BLT) humanized mice. Ad5.PGT121 and AAV1.PGT121 produced functional antibody in vivo Ad5.PGT121 produced PGT121 rapidly within 6 h, whereas AAV1.PGT121 produced detectable PGT121 in serum by 72 h. Serum PGT121 levels were rapidly reduced by the generation of anti-PGT121 antibodies in immunocompetent mice but were durably maintained in immunocompromised mice. In HIV-1-infected BLT humanized mice, Ad5.PGT121 resulted in a greater reduction of viral loads than did AAV1.PGT121. Ad5.PGT121 also led to more-sustained virologic control than purified PGT121 IgG. Ad5.PGT121 afforded more rapid, robust, and durable antiviral efficacy than AAV1.PGT121 and purified PGT121 IgG in HIV-1-infected humanized mice. Further evaluation of vector delivery of HIV-1 bNAbs is warranted, although approaches to prevent the generation of antiantibody responses may also be required. IMPORTANCE Broadly neutralizing antibodies (bNAbs) are being explored for HIV-1 prevention and cure strategies, but delivery of purified antibodies may prove challenging. We investigated adenovirus serotype 5 (Ad5) and adeno-associated virus serotype 1 (AAV1) vectors to deliver the HIV-1-specific bNAb PGT121. Ad5.PGT121 afforded more rapid, robust, and durable antiviral efficacy than AAV1.PGT121 and purified PGT121 IgG in HIV-1-infected humanized mice. Copyright © 2018 Badamchi-Zadeh et al.

Immunization with 60 kD Ro peptide produces different stages of preclinical autoimmunity in a Sjögren's syndrome model among multiple strains of inbred mice.

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Kurien, B T; Dsouza, A; Igoe, A; Lee, Y J; Maier-Moore, J S; Gordon, T; Jackson, M; Scofield, R H

2013-07-01

Sjögren's syndrome is a chronic illness manifested characteristically by immune injury to the salivary and lacrimal glands, resulting in dry mouth/eyes. Anti-Ro [Sjögren's syndrome antigen A (SSA)] and anti-La [Sjögren's syndrome antigen B (SSB)] autoantibodies are found frequently in Sjögren's subjects as well as in individuals who will go on to develop the disease. Immunization of BALB/c mice with Ro60 peptides results in epitope spreading with anti-Ro and anti-La along with lymphocyte infiltration of salivary glands similar to human Sjögren's. In addition, these animals have poor salivary function/low saliva volume. In this study, we examined whether Ro-peptide immunization produces a Sjögren's-like illness in other strains of mice. BALB/c, DBA-2, PL/J, SJL/J and C57BL/6 mice were immunized with Ro60 peptide-274. Sera from these mice were studied by immunoblot and enzyme-linked immunosorbent assay for autoantibodies. Timed salivary flow was determined after pharmacological stimulation, and salivary glands were examined pathologically. We found that SJL/J mice had no immune response to the peptide from Ro60, while C57BL/6 mice produced antibodies that bound the peptide but had no epitope spreading. PL/J mice had epitope spreading to other structures of Ro60 as well as to La, but like C57BL/6 and SJL/J had no salivary gland lymphocytic infiltration and no decrement of salivary function. DBA-2 and BALB/c mice had infiltration but only BALB/c had decreased salivary function. The immunological processes leading to a Sjögren's-like illness after Ro-peptide immunization were interrupted in a stepwise fashion in these differing mice strains. These data suggest that this is a model of preclinical disease with genetic control for epitope spreading, lymphocytic infiltration and glandular dysfunction. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

Frequency-specific corticofugal modulation of the dorsal cochlear nucleus in mice.

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Kong, Lingzhi; Xiong, Colin; Li, Liang; Yan, Jun

2014-01-01

The primary auditory cortex (AI) modulates the sound information processing in the lemniscal subcortical nuclei, including the anteroventral cochlear nucleus (AVCN), in a frequency-specific manner. The dorsal cochlear nucleus (DCN) is a non-lemniscal subcortical nucleus but it is tonotopically organized like the AVCN. However, it remains unclear how the AI modulates the sound information processing in the DCN. This study examined the impact of focal electrical stimulation of AI on the auditory responses of the DCN neurons in mice. We found that the electrical stimulation induced significant changes in the best frequency (BF) of DCN neurons. The changes in the BFs were highly specific to the BF differences between the stimulated AI neurons and the recorded DCN neurons. The DCN BFs shifted higher when the AI BFs were higher than the DCN BFs and the DCN BFs shifted lower when the AI BFs were lower than the DCN BFs. The DCN BFs showed no change when the AI and DCN BFs were similar. Moreover, the BF shifts were linearly correlated to the BF differences. Thus, our data suggest that corticofugal modulation of the DCN is also highly specific to frequency information, similar to the corticofugal modulation of the AVCN. The frequency-specificity of corticofugal modulation does not appear limited to the lemniscal ascending pathway.

Dual specificity phosphatase 6 deficiency is associated with impaired systemic glucose tolerance and reversible weight retardation in mice.

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Katrin Pfuhlmann

Full Text Available Here, we aimed to investigate the potential role of DUSP6, a dual specificity phosphatase, that specifically inactivates extracellular signal-regulated kinase (ERK, for the regulation of body weight and glucose homeostasis. We further assessed whether metabolic challenges affect Dusp6 expression in selected brain areas or white adipose tissue. Hypothalamic Dusp6 mRNA levels remained unchanged in chow-fed lean vs. high fat diet (HFD fed obese C57Bl/6J mice, and in C57Bl/6J mice undergoing prolonged fasting or refeeding with fat free diet (FFD or HFD. Similarly, Dusp6 expression levels were unchanged in selected brain regions of Lepob mice treated with 1 mg/kg of leptin for 6 days, compared to pair-fed or saline-treated Lepob controls. Dusp6 expression levels remained unaltered in vitro in primary adipocytes undergoing differentiation, but were increased in eWAT of HFD-fed obese C57Bl/6J mice, compared to chow-fed lean controls. Global chow-fed DUSP6 KO mice displayed reduced body weight and lean mass and slightly increased fat mass at a young age, which is indicative for early-age weight retardation. Subsequent exposure to HFD led to a significant increase in lean mass and body weight in DUSP6 deficient mice, compared to WT controls. Nevertheless, after 26 weeks of high-fat diet exposure, we observed comparable body weight, fat and lean mass in DUSP6 WT and KO mice, suggesting overall normal susceptibility to develop obesity. In line with the increased weight gain to compensate for early-age weight retardation, HFD-fed DUSP6 KO displayed increased expression levels of anabolic genes involved in lipid and cholesterol metabolism in the epididymal white adipose tissue (eWAT, compared to WT controls. Glucose tolerance was perturbed in both chow-fed lean or HFD-fed obese DUSP6 KO, compared to their respective WT controls. Overall, our data indicate that DUSP6 deficiency has limited impact on the regulation of energy metabolism, but impairs systemic

Skeletal muscle-specific HMG-CoA reductase knockout mice exhibit rhabdomyolysis: A model for statin-induced myopathy.

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Osaki, Yoshinori; Nakagawa, Yoshimi; Miyahara, Shoko; Iwasaki, Hitoshi; Ishii, Akiko; Matsuzaka, Takashi; Kobayashi, Kazuto; Yatoh, Shigeru; Takahashi, Akimitsu; Yahagi, Naoya; Suzuki, Hiroaki; Sone, Hirohito; Ohashi, Ken; Ishibashi, Shun; Yamada, Nobuhiro; Shimano, Hitoshi

2015-10-23

HMG-CoA reductase (HMGCR) catalyzes the conversion of HMG-CoA to mevalonic acid (MVA); this is the rate-limiting enzyme of the mevalonate pathway that synthesizes cholesterol. Statins, HMGCR inhibitors, are widely used as cholesterol-reducing drugs. However, statin-induced myopathy is the most adverse side effect of statins. To eludicate the mechanisms underlying statin the myotoxicity and HMGCR function in the skeletal muscle, we developed the skeletal muscle-specific HMGCR knockout mice. Knockout mice exhibited postnatal myopathy with elevated serum creatine kinase levels and necrosis. Myopathy in knockout mice was completely rescued by the oral administration of MVA. These results suggest that skeletal muscle toxicity caused by statins is dependent on the deficiencies of HMGCR enzyme activity and downstream metabolites of the mevalonate pathway in skeletal muscles rather than the liver or other organs. Copyright © 2015 Elsevier Inc. All rights reserved.

Secretory IgM Exacerbates Tumor Progression by Inducing Accumulations of MDSCs in Mice.

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Tang, Chih-Hang Anthony; Chang, Shiun; Hashimoto, Ayumi; Chen, Yi-Ju; Kang, Chang Won; Mato, Anthony R; Del Valle, Juan R; Gabrilovich, Dmitry I; Hu, Chih-Chi Andrew

2018-06-01

Chronic lymphocytic leukemia (CLL) cells can secrete immunoglobulin M. However, it is not clear whether secretory IgM (sIgM) plays a role in disease progression. We crossed the Eμ-TCL1 mouse model of CLL, in which the expression of human TCL1 oncogene was driven by the V(H) promoter-Ig(H)-Eμ enhancer, with MD4 mice whose B cells produced B-cell receptor (membrane-bound IgM) and sIgM with specificity for hen egg lysozyme (HEL). CLL cells that developed in these MD4/Eμ-TCL1 mice reactivated a parental Ig gene allele and secreted IgM, and did not recognize HEL. The MD4/Eμ-TCL1 mice had reduced survival, increased myeloid-derived suppressor cells (MDSC), and decreased numbers of T cells. We tested whether sIgM could contribute to the accumulation of MDSCs by crossing μS -/- mice, which could not produce sIgM, with Eμ-TCL1 mice. The μS -/- /Eμ-TCL1 mice survived longer than Eμ-TCL1 mice and developed decreased numbers of MDSCs which were less able to suppress proliferation of T cells. We targeted the synthesis of sIgM by deleting the function of XBP-1s and showed that targeting XBP-1s genetically or pharmacologically could lead to decreased sIgM, accompanied by decreased numbers and reduced functions of MDSCs in MD4/Eμ-TCL1 mice. Additionally, MDSCs from μS -/- mice grafted with Lewis lung carcinoma were inefficient suppressors of T cells, resulting in slower tumor growth. These results demonstrate that sIgM produced by B cells can upregulate the functions of MDSCs in tumor-bearing mice to aggravate cancer progression. In a mouse model of CLL, production of secretory IgM led to more MDSCs, fewer T cells, and shorter survival times for the mice. Thus, secretory IgM may aggravate the progression of this cancer. Cancer Immunol Res; 6(6); 696-710. ©2018 AACR . ©2018 American Association for Cancer Research.

Antivenom reversal of biochemical alterations induced by black scorpion Heterometrus fastigiousus Couzijn venom in mice

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MK Chaubey

2009-01-01

Full Text Available In the present study, Heterometrus fastigiousus venom (HFV was employed as antigen to produce species-specific scorpion antivenom (SAV in albino mice (NIH strain. To determine SAV efficacy, it was pre-incubated with 10 LD50 of HFV and then injected subcutaneously into mice. Subsequently, mortality was observed after 24 hours. Minimum effective dose (MED was 12.5 LD50 of HFV/mL of SAV. SAV effectiveness to reverse HFV-induced biochemical alterations in mice was analyzed by challenge method. Simultaneously, mice received subcutaneously 40% of 24-hour-LD50 of HFV and intravenously SAV. After four hours, changes in serum glucose, free amino acids, uric acids, pyruvic acid, cholesterol, total protein, alkaline phosphatase, acid phosphatase, lactic dehydrogenase and glutamate-pyruvate transaminase enzyme level were determined. Treatment with species-specific SAV resulted in the reversal of HFV-induced biochemical alterations.

Effects of PPARs agonists on cardiac metabolism in littermate and cardiomyocyte-specific PPAR-γ-knockout (CM-PGKO mice.

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Michelangela Barbieri

Full Text Available Understanding the molecular regulatory mechanisms controlling for myocardial lipid metabolism is of critical importance for the development of new therapeutic strategies for heart diseases. The role of PPARγ and thiazolidinediones in regulation of myocardial lipid metabolism is controversial. The aim of our study was to assess the role of PPARγ on myocardial lipid metabolism and function and differentiate local/from systemic actions of PPARs agonists using cardiomyocyte-specific PPARγ -knockout (CM-PGKO mice. To this aim, the effect of PPARγ, PPARγ/PPARα and PPARα agonists on cardiac function, intra-myocyte lipid accumulation and myocardial expression profile of genes and proteins, affecting lipid oxidation, uptake, synthesis, and storage (CD36, CPT1MIIA, AOX, FAS, SREBP1-c and ADPR was evaluated in cardiomyocyte-specific PPARγ-knockout (CM-PGKO and littermate control mice undergoing standard and high fat diet (HFD. At baseline, protein levels and mRNA expression of genes involved in lipid uptake, oxidation, synthesis, and accumulation of CM-PGKO mice were not significantly different from those of their littermate controls. At baseline, no difference in myocardial lipid content was found between CM-PGKO and littermate controls. In standard condition, pioglitazone and rosiglitazone do not affect myocardial metabolism while, fenofibrate treatment significantly increased CD36 and CPT1MIIA gene expression. In both CM-PGKO and control mice submitted to HFD, six weeks of treatment with rosiglitazone, fenofibrate and pioglitazone lowered myocardial lipid accumulation shifting myocardial substrate utilization towards greater contribution of glucose. In conclusion, at baseline, PPARγ does not play a crucial role in regulating cardiac metabolism in mice, probably due to its low myocardial expression. PPARs agonists, indirectly protect myocardium from lipotoxic damage likely reducing fatty acids delivery to the heart through the actions on adipose

Decreased spontaneous activity in AMPK alpha 2 muscle specific kinase dead mice is not caused by changes in brain dopamine metabolism

DEFF Research Database (Denmark)

Møller, Lisbeth Liliendal Valbjørn; Sylow, Lykke; Gøtzsche, Casper René

2016-01-01

was tested in an open field test. Furthermore, we investigated maximal running capacity and voluntary running over a period of 19 days. AMPK α2 KD mice ran 30% less in daily distance compared to WT. Furthermore, AMPK α2 KD mice showed significantly decreased locomotor activity in the open field test compared...... through alterations of the brain dopamine levels specifically in the striatal region. To test this hypothesis, transgenic mice overexpressing an inactivatable dominant negative α2 AMPK construct (AMPK α2 KD) in muscles and littermate wildtype (WT) mice were tested. AMPK α2 KD mice have impaired running...... capacity and display reduced voluntary wheel running activity. Striatal content of dopamine and its metabolites were measured under basal physiological conditions and after cocaine-induced dopamine efflux from the ventral striatum by in vivo microdialysis. Moreover, cocaine-induced locomotor activity...

Neuron-specific caveolin-1 overexpression improves motor function and preserves memory in mice subjected to brain trauma.

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Egawa, Junji; Schilling, Jan M; Cui, Weihua; Posadas, Edmund; Sawada, Atsushi; Alas, Basheer; Zemljic-Harpf, Alice E; Fannon-Pavlich, McKenzie J; Mandyam, Chitra D; Roth, David M; Patel, Hemal H; Patel, Piyush M; Head, Brian P

2017-08-01

Studies in vitro and in vivo demonstrate that membrane/lipid rafts and caveolin (Cav) organize progrowth receptors, and, when overexpressed specifically in neurons, Cav-1 augments neuronal signaling and growth and improves cognitive function in adult and aged mice; however, whether neuronal Cav-1 overexpression can preserve motor and cognitive function in the brain trauma setting is unknown. Here, we generated a neuron-targeted Cav-1-overexpressing transgenic (Tg) mouse [synapsin-driven Cav-1 (SynCav1 Tg)] and subjected it to a controlled cortical impact model of brain trauma and measured biochemical, anatomic, and behavioral changes. SynCav1 Tg mice exhibited increased hippocampal expression of Cav-1 and membrane/lipid raft localization of postsynaptic density protein 95, NMDA receptor, and tropomyosin receptor kinase B. When subjected to a controlled cortical impact, SynCav1 Tg mice demonstrated preserved hippocampus-dependent fear learning and memory, improved motor function recovery, and decreased brain lesion volume compared with wild-type controls. Neuron-targeted overexpression of Cav-1 in the adult brain prevents hippocampus-dependent learning and memory deficits, restores motor function after brain trauma, and decreases brain lesion size induced by trauma. Our findings demonstrate that neuron-targeted Cav-1 can be used as a novel therapeutic strategy to restore brain function and prevent trauma-associated maladaptive plasticity.-Egawa, J., Schilling, J. M., Cui, W., Posadas, E., Sawada, A., Alas, B., Zemljic-Harpf, A. E., Fannon-Pavlich, M. J., Mandyam, C. D., Roth, D. M., Patel, H. H., Patel, P. M., Head, B. P. Neuron-specific caveolin-1 overexpression improves motor function and preserves memory in mice subjected to brain trauma. © FASEB.

Evaluating genome-wide DNA methylation changes in mice by Methylation Specific Digital Karyotyping

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Maruoka Shuichiro

2008-12-01

Full Text Available Abstract Background The study of genome-wide DNA methylation changes has become more accessible with the development of various array-based technologies though when studying species other than human the choice of applications are limited and not always within reach. In this study, we adapted and tested the applicability of Methylation Specific Digital Karyotyping (MSDK, a non-array based method, for the prospective analysis of epigenetic changes after perinatal nutritional modifications in a mouse model of allergic airway disease. MSDK is a sequenced based method that allows a comprehensive and unbiased methylation profiling. The method generates 21 base pairs long sequence tags derived from specific locations in the genome. The resulting tag frequencies determine in a quantitative manner the methylation level of the corresponding loci. Results Genomic DNA from whole lung was isolated and subjected to MSDK analysis using the methylation-sensitive enzyme Not I as the mapping enzyme and Nla III as the fragmenting enzyme. In a pair wise comparison of the generated mouse MSDK libraries we identified 158 loci that are significantly differentially methylated (P-value = 0.05 after perinatal dietary changes in our mouse model. Quantitative methylation specific PCR and sequence analysis of bisulfate modified genomic DNA confirmed changes in methylation at specific loci. Differences in genomic MSDK tag counts for a selected set of genes, correlated well with changes in transcription levels as measured by real-time PCR. Furthermore serial analysis of gene expression profiling demonstrated a dramatic difference in expressed transcripts in mice exposed to perinatal nutritional changes. Conclusion The genome-wide methylation survey applied in this study allowed for an unbiased methylation profiling revealing subtle changes in DNA methylation in mice maternally exposed to dietary changes in methyl-donor content. The MSDK method is applicable for mouse models

Dose-specific transcriptional responses in thyroid tissue in mice after 131I administration

International Nuclear Information System (INIS)

Rudqvist, Nils; Schüler, Emil; Parris, Toshima Z.; Langen, Britta; Helou, Khalil; Forssell-Aronsson, Eva

2015-01-01

Introduction: In the present investigation, microarray analysis was used to monitor transcriptional activity in thyroids in mice 24 h after 131 I exposure. The aims of this study were to 1) assess the transcriptional patterns associated with 131 I exposure in normal mouse thyroid tissue and 2) propose biomarkers for 131 I exposure of the thyroid. Methods: Adult BALB/c nude mice were i.v. injected with 13, 130 or 260 kBq of 131 I and killed 24 h after injection (absorbed dose to thyroid: 0.85, 8.5, or 17 Gy). Mock-treated mice were used as controls. Total RNA was extracted from thyroids and processed using the Illumina platform. Results: In total, 497, 546, and 90 transcripts were regulated (fold change ≥ 1.5) in the thyroid after 0.85, 8.5, and 17 Gy, respectively. These were involved in several biological functions, e.g. oxygen access, inflammation and immune response, and apoptosis/anti-apoptosis. Approximately 50% of the involved transcripts at each absorbed dose level were dose-specific, and 18 transcripts were commonly detected at all absorbed dose levels. The Agpat9, Plau, Prf1, and S100a8 gene expression displayed a monotone decrease in regulation with absorbed dose, and further studies need to be performed to evaluate if they may be useful as dose-related biomarkers for 131I exposure. Conclusion: Distinct and substantial differences in gene expression and affected biological functions were detected at the different absorbed dose levels. The transcriptional profiles were specific for the different absorbed dose levels. We propose that the Agpat9, Plau, Prf1, and S100a8 genes might be novel potential absorbed dose-related biomarkers to 131 I exposure of thyroid. Advances in knowledge: During the recent years, genomic techniques have been developed; however, they have not been fully utilized in nuclear medicine and radiation biology. We have used RNA microarrays to investigate genome-wide transcriptional regulations in thyroid tissue in mice after low

The role of CD80/CD86 in generation and maintenance of functional virus-specific CD8+ T cells in mice infected with lymphocytic choriomeningitis virus

DEFF Research Database (Denmark)

Grujic, Mirjana; Bartholdy, Christina; Remy, Melissa

2010-01-01

Lymphocytic choriomeningitis virus (LCMV)-specific CD8(+) T cell responses are considered to be independent of CD28-B7 costimulation. However, the LCMV-specific response has never been evaluated in B7.1/B7.2(-/-) mice. For this reason, we decided to study the T cell response in B7.1/B7.2(-/-) mice......, but no chronic infection. Taken together, these results indicate that B7 costimulation is required for induction and maintenance of LCMV-specific CD8(+) T cell memory, irrespective of the LCMV strain used for priming. However, the erosion of CD8(+) T cell memory in B7.1/B7.2(-/-) mice was more pronounced...

Gender-specific reduction of hepatic Mrp2 expression by high-fat diet protects female mice from ANIT toxicity

International Nuclear Information System (INIS)

Kong, Bo; Csanaky, Iván L.; Aleksunes, Lauren M.; Patni, Meghan; Chen, Qi; Ma, Xiaochao; Jaeschke, Hartmut; Weir, Scott; Broward, Melinda; Klaassen, Curtis D.; Guo, Grace L.

2012-01-01

Emerging evidence suggests that feeding a high-fat diet (HFD) to rodents affects the expression of genes involved in drug transport. However, gender-specific effects of HFD on drug transport are not known. The multidrug resistance-associated protein 2 (Mrp2, Abcc2) is a transporter highly expressed in the hepatocyte canalicular membrane and is important for biliary excretion of glutathione-conjugated chemicals. The current study showed that hepatic Mrp2 expression was reduced by HFD feeding only in female, but not male, C57BL/6J mice. In order to determine whether down-regulation of Mrp2 in female mice altered chemical disposition and toxicity, the biliary excretion and hepatotoxicity of the Mrp2 substrate, α-naphthylisothiocyanate (ANIT), were assessed in male and female mice fed control diet or HFD for 4 weeks. ANIT-induced biliary injury is a commonly used model of experimental cholestasis and has been shown to be dependent upon Mrp2-mediated efflux of an ANIT glutathione conjugate that selectively injures biliary epithelial cells. Interestingly, HFD feeding significantly reduced early-phase biliary ANIT excretion in female mice and largely protected against ANIT-induced liver injury. In summary, the current study showed that, at least in mice, HFD feeding can differentially regulate Mrp2 expression and function and depending upon the chemical exposure may enhance or reduce susceptibility to toxicity. Taken together, these data provide a novel interaction between diet and gender in regulating hepatobiliary excretion and susceptibility to injury. -- Highlights: ► High-fat diet decreases hepatic Mrp2 expression only in female but not in male mice. ► HFD significantly reduces early-phase biliary ANIT excretion in female mice. ► HFD protects female mice against ANIT-induced liver injury.

Variant-specific immunity to Plasmodium berghei in pregnant mice

DEFF Research Database (Denmark)

Megnekou, Rosette; Hviid, Lars; Staalsoe, Trine

2009-01-01

for recrudescence-type IEs are related to the protection of pregnant mice from maternal anemia, low birth weight, and decreased litter size. We conclude that the model replicates many of the key parasitological and immunological features of PAM, although the P. berghei genome does not encode proteins homologous...... to the P. falciparum erythrocyte membrane protein 1 adhesins, which are of key importance in P. falciparum malaria. The study of P. berghei malaria in pregnant, immune mice can be used to gain significant new insights regarding malaria pathogenesis and immunity in general and regarding PAM in particular....

CRR and monetary control

OpenAIRE

Michael R. Pakko

1985-01-01

A review of the history of the lagged reserve requirements- contemporaneous reserve requirements controversy and a discussion of the relative importance of reserve timing under various operating procedures.

Self-recognition specificity expressed by T cells from nude mice. Absence of detectable Ia-restricted T cells in nude mice that do exhibit self-K/D-restricted T cell responses

International Nuclear Information System (INIS)

Kruisbeek, A.M.; Davis, M.L.; Matis, L.A.; Longo, D.L.

1984-01-01

The presence in athymic nude mice of precursor T cells with self-recognition specificity for either H-2 K/D or H-2 I region determinants was investigated. Chimeras were constructed of lethally irradiated parental mice receiving a mixture of F1 nude mouse (6-8 wk old) spleen and bone marrow cells. The donor inoculum was deliberately not subjected to any T cell depletion procedure, so that any potential major histocompatibility complex-committed precursor T cells were allowed to differentiate and expand in the normal parental recipients. 3 mo after reconstitution, the chimeras were immunized with several protein antigens in complete Freund's adjuvant in the footpads and their purified draining lymph node T cells tested 10 d later for ability to recognize antigen on antigen-presenting cells of either parental haplotype. Also, their spleen and lymph node cells were tested for ability to generate a cytotoxic T lymphocyte (CTL) response to trinitrophenyl (TNP)-modified stimulator cells of either parental haplotype. It was demonstrated that T cell proliferative responses of these F1(nude)----parent chimeras were restricted solely to recognizing parental host I region determinants as self and expressed the Ir gene phenotype of the host. In contrast, CTL responses could be generated (in the presence of interleukin 2) to TNP-modified stimulator cells of either parental haplotype. Thus these results indicate that nude mice which do have CTL with self-specificity for K/D region determinants lack proliferating T cells with self-specificity for I region determinants. These results provide evidence for the concepts that development of the I region-restricted T cell repertoire is strictly an intrathymically determined event and that young nude mice lack the unique thymic elements responsible for education of I region-restricted T cells

Bifidobacterium adolescentis Exerts Strain-Specific Effects on Constipation Induced by Loperamide in BALB/c Mice

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Linlin Wang

2017-02-01

Full Text Available Constipation is one of the most common gastrointestinal complaints worldwide. This study was performed to determine whether Bifidobacterium adolescentis exerts inter-strain differences in alleviating constipation induced by loperamide in BALB/c mice and to analyze the main reasons for these differences. BALB/c mice underwent gavage with B. adolescentis (CCFM 626, 667, and 669 once per day for 17 days. The primary outcome measures included related constipation indicators, and the secondary outcome measures were the basic biological characteristics of the strains, the concentration changes of short-chain fatty acids in feces, and the changes in the fecal flora. B. adolescentis CCFM 669 and 667 relieved constipation symptoms by adhering to intestinal epithelial cells, growing quickly in vitro and increasing the concentrations of propionic and butyric acids. The effect of B. adolescentis on the gut microbiota in mice with constipation was investigated via 16S rRNA metagenomic analysis. The results revealed that the relative abundance of Lactobacillus increased and the amount of Clostridium decreased in the B. adolescentis CCFM 669 and 667 treatment groups. In conclusion, B. adolescentis exhibits strain-specific effects in the alleviation of constipation, mostly due to the strains’ growth rates, adhesive capacity and effects on the gut microbiome and microenvironment.

Comparison of the virulence of exopolysaccharide-producing Prevotella intermedia to exopolysaccharide non-producing periodontopathic organisms.

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Yamanaka, Takeshi; Yamane, Kazuyoshi; Furukawa, Tomoyo; Matsumoto-Mashimo, Chiho; Sugimori, Chieko; Nambu, Takayuki; Obata, Noboru; Walker, Clay B; Leung, Kai-Poon; Fukushima, Hisanori

2011-08-25

Evidence in the literature suggests that exopolysaccharides (EPS) produced by bacterial cells are essential for the expression of virulence in these organisms. Secreted EPSs form the framework in which microbial biofilms are built. This study evaluates the role of EPS in Prevotella intermedia for the expression of virulence. This evaluation was accomplished by comparing EPS-producing P. intermedia strains 17 and OD1-16 with non-producing P. intermedia ATCC 25611 and Porphyromonas gingivalis strains ATCC 33277, 381 and W83 for their ability to induce abscess formation in mice and evade phagocytosis. EPS-producing P. intermedia strains 17 and OD1-16 induced highly noticeable abscess lesions in mice at 107 colony-forming units (CFU). In comparison, P. intermedia ATCC 25611 and P. gingivalis ATCC 33277, 381 and W83, which all lacked the ability to produce viscous materials, required 100-fold more bacteria (109 CFU) in order to induce detectable abscess lesions in mice. Regarding antiphagocytic activity, P. intermedia strains 17 and OD1-16 were rarely internalized by human polymorphonuclear leukocytes, but other strains were readily engulfed and detected in the phagosomes of these phagocytes. These results demonstrate that the production of EPS by P. intermedia strains 17 and OD1-16 could contribute to the pathogenicity of this organism by conferring their ability to evade the host's innate defence response.

Pancreas-Specific Sirt1-Deficiency in Mice Compromises Beta-Cell Function without Development of Hyperglycemia.

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Pinho, Andreia V; Bensellam, Mohammed; Wauters, Elke; Rees, Maxine; Giry-Laterriere, Marc; Mawson, Amanda; Ly, Le Quan; Biankin, Andrew V; Wu, Jianmin; Laybutt, D Ross; Rooman, Ilse

2015-01-01

Sirtuin 1 (Sirt1) has been reported to be a critical positive regulator of glucose-stimulated insulin secretion in pancreatic beta-cells. The effects on islet cells and blood glucose levels when Sirt1 is deleted specifically in the pancreas are still unclear. This study examined islet glucose responsiveness, blood glucose levels, pancreatic islet histology and gene expression in Pdx1Cre; Sirt1ex4F/F mice that have loss of function and loss of expression of Sirt1 specifically in the pancreas. We found that in the Pdx1Cre; Sirt1ex4F/F mice, the relative insulin positive area and the islet size distribution were unchanged. However, beta-cells were functionally impaired, presenting with lower glucose-stimulated insulin secretion. This defect was not due to a reduced expression of insulin but was associated with a decreased expression of the glucose transporter Slc2a2/Glut2 and of the Glucagon like peptide-1 receptor (Glp1r) as well as a marked down regulation of endoplasmic reticulum (ER) chaperones that participate in the Unfolded Protein Response (UPR) pathway. Counter intuitively, the Sirt1-deficient mice did not develop hyperglycemia. Pancreatic polypeptide (PP) cells were the only other islet cells affected, with reduced numbers in the Sirt1-deficient pancreas. This study provides new mechanistic insights showing that beta-cell function in Sirt1-deficient pancreas is affected due to altered glucose sensing and deregulation of the UPR pathway. Interestingly, we uncovered a context in which impaired beta-cell function is not accompanied by increased glycemia. This points to a unique compensatory mechanism. Given the reduction in PP, investigation of its role in the control of blood glucose is warranted.

Pancreas-Specific Sirt1-Deficiency in Mice Compromises Beta-Cell Function without Development of Hyperglycemia.

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Andreia V Pinho

Full Text Available Sirtuin 1 (Sirt1 has been reported to be a critical positive regulator of glucose-stimulated insulin secretion in pancreatic beta-cells. The effects on islet cells and blood glucose levels when Sirt1 is deleted specifically in the pancreas are still unclear.This study examined islet glucose responsiveness, blood glucose levels, pancreatic islet histology and gene expression in Pdx1Cre; Sirt1ex4F/F mice that have loss of function and loss of expression of Sirt1 specifically in the pancreas.We found that in the Pdx1Cre; Sirt1ex4F/F mice, the relative insulin positive area and the islet size distribution were unchanged. However, beta-cells were functionally impaired, presenting with lower glucose-stimulated insulin secretion. This defect was not due to a reduced expression of insulin but was associated with a decreased expression of the glucose transporter Slc2a2/Glut2 and of the Glucagon like peptide-1 receptor (Glp1r as well as a marked down regulation of endoplasmic reticulum (ER chaperones that participate in the Unfolded Protein Response (UPR pathway. Counter intuitively, the Sirt1-deficient mice did not develop hyperglycemia. Pancreatic polypeptide (PP cells were the only other islet cells affected, with reduced numbers in the Sirt1-deficient pancreas.This study provides new mechanistic insights showing that beta-cell function in Sirt1-deficient pancreas is affected due to altered glucose sensing and deregulation of the UPR pathway. Interestingly, we uncovered a context in which impaired beta-cell function is not accompanied by increased glycemia. This points to a unique compensatory mechanism. Given the reduction in PP, investigation of its role in the control of blood glucose is warranted.

Organ specific mapping of in vivo redox state in control and cigarette smoke-exposed mice using EPR/NMR co-imaging

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Caia, George L.; Efimova, Olga V.; Velayutham, Murugesan; El-Mahdy, Mohamed A.; Abdelghany, Tamer M.; Kesselring, Eric; Petryakov, Sergey; Sun, Ziqi; Samouilov, Alexandre; Zweier, Jay L.

2014-01-01

In vivo mapping of alterations in redox status is important for understanding organ specific pathology and disease. While electron paramagnetic resonance imaging (EPRI) enables spatial mapping of free radicals, it does not provide anatomic visualization of the body. Proton MRI is well suited to provide anatomical visualization. We applied EPR/NMR co-imaging instrumentation to map and monitor the redox state of living mice under normal or oxidative stress conditions induced by secondhand cigarette smoke (SHS) exposure. A hybrid co-imaging instrument, EPRI (1.2 GHz) / proton MRI (16.18 MHz), suitable for whole-body co-imaging of mice was utilized with common magnet and gradients along with dual EPR/NMR resonators that enable co-imaging without sample movement. The metabolism of the nitroxide probe, 3–carbamoyl–proxyl (3-CP), was used to map the redox state of control and SHS-exposed mice. Co-imaging allowed precise 3D mapping of radical distribution and reduction in major organs such as the heart, lungs, liver, bladder and kidneys. Reductive metabolism was markedly decreased in SHS-exposed mice and EPR/NMR co-imaging allowed quantitative assessment of this throughout the body. Thus, in vivo EPR/NMR co-imaging enables in vivo organ specific mapping of free radical metabolism and redox stress and the alterations that occur in the pathogenesis of disease. PMID:22296801

Dengue envelope-based 'four-in-one' virus-like particles produced using Pichia pastoris induce enhancement-lacking, domain III-directed tetravalent neutralising antibodies in mice.

Science.gov (United States)

Rajpoot, Ravi Kant; Shukla, Rahul; Arora, Upasana; Swaminathan, Sathyamangalam; Khanna, Navin

2018-06-05

Dengue is a significant public health problem worldwide, caused by four antigenically distinct mosquito-borne dengue virus (DENV) serotypes. Antibodies to any given DENV serotype which can afford protection against that serotype tend to enhance infection by other DENV serotypes, by a phenomenon termed antibody-dependent enhancement (ADE). Antibodies to the viral pre-membrane (prM) protein have been implicated in ADE. We show that co-expression of the envelope protein of all four DENV serotypes, in the yeast Pichia pastoris, leads to their co-assembly, in the absence of prM, into tetravalent mosaic VLPs (T-mVLPs), which retain the serotype-specific antigenic integrity and immunogenicity of all four types of their monomeric precursors. Following a three-dose immunisation schedule, the T-mVLPs elicited EDIII-directed antibodies in mice which could neutralise all four DENV serotypes. Importantly, anti-T-mVLP antibodies did not augment sub-lethal DENV-2 infection of dengue-sensitive AG129 mice, based on multiple parameters. The 'four-in-one' tetravalent T-mVLPs possess multiple desirable features which may potentially contribute to safety (non-viral, prM-lacking and ADE potential-lacking), immunogenicity (induction of virus-neutralising antibodies), and low cost (single tetravalent immunogen produced using P. pastoris, an expression system known for its high productivity using simple inexpensive media). These results strongly warrant further exploration of this vaccine candidate.

Protection against Fasciola gigantica infection in mice by vaccination with recombinant juvenile-specific cathepsin L.

Science.gov (United States)

Sansri, Veerawat; Meemon, Krai; Changklungmoa, Narin; Kueakhai, Pornanan; Chantree, Pathanin; Chaichanasak, Pannigan; Lorsuwannarat, Natcha; Itagaki, Tadashi; Sobhon, Prasert

2015-03-24

Fasciola gigantica cathepsin L1H (FgCatL1H) is one of the major cathepsin L released by juveniles of F. gigantica to aid in the invasion of host's tissues. Due to its high sequence similarity with other cathepsin L (CatL) isoforms of late stage F. gigantica, it was considered to be a good vaccine candidate that can block all CatL-mediated protease activities and affect juveniles as well as adult parasites. In this study, recombinant proFgCatL1H protein expressed in yeast, Pichia pastoris, system was mixed with Freund's adjuvants and used to subcutaneously immunize mice that were later challenged with metacercariae of F. gigantica. The percentage of worm protection in the rproFgCatL1H-vaccinated mice compared to the non-immunized and adjuvant control mice were approximately 62.7% and 66.1%, respectively. Anti-rproFgCatL1H antisera collected from vaccinated mice reacted specifically with rproFgCatL1H and other cathepsin L isoforms of F. gigantica, but the antibodies did not cross react with antigens from other trematode and nematode parasites, including Eurytrema pancreaticum, Opisthorchis viverrini, Fischoederius cobboldi, Cotylophoron cotylophorum, Gigantocotyle explanatum, Paramphistomum cervi, and Setaria labiato-papillosa. The levels of IgG1 and IgG2a in mouse sera increased significantly at two weeks after immunization and were highest during the sixth to eighth weeks after immunization. The IgG1 level was higher than IgG2a at all periods of immunization, implicating the dominance of the Th2 response. The levels of IgG1 and IgG2a in the immune sera were shown to be strongly correlated with the numbers of worm recovery, and the correlation coefficient was higher for IgG1. The levels of serum aspartate aminotransferase and alanine transaminase were significantly lower in the sera of rproFgCatL1H-vaccinated mice than in the infected control mice indicating a lower degree of liver damage. This study demonstrated a high potential of FgCatL1H vaccine, and its

Formulation of the bivalent prostate cancer vaccine with surgifoam elicits antigen-specific effector T cells in PSA-transgenic mice.

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Karan, Dev

2017-10-13

We previously developed and characterized an adenoviral-based prostate cancer vaccine for simultaneous targeting of prostate-specific antigen (PSA) and prostate stem cell antigen (PSCA). We also demonstrated that immunization of mice with the bivalent vaccine (Ad 5 -PSA+PSCA) inhibited the growth of established prostate tumors. However, there are multiple challenges hindering the success of immunological therapies in the clinic. One of the prime concerns has been to overcome the immunological tolerance and maintenance of long-term effector T cells. In this study, we further characterized the use of the bivalent vaccine (Ad 5 -PSA+PSCA) in a transgenic mouse model expressing human PSA in the mouse prostate. We demonstrated the expression of PSA analyzed at the mRNA level (by RT-PCR) and protein level (by immunohistochemistry) in the prostate lobes harvested from the PSA-transgenic (PSA-Tg) mice. We established that the administration of the bivalent vaccine in surgifoam to the PSA-Tg mice induces strong PSA-specific effector CD8 + T cells as measured by IFN-γ secretion and in vitro cytotoxic T-cell assay. Furthermore, the use of surgifoam with Ad 5 -PSA+PSCA vaccine allows multiple boosting vaccinations with a significant increase in antigen-specific CD8 + T cells. These observations suggest that the formulation of the bivalent prostate cancer vaccine (Ad 5 -PSA+PSCA) with surgifoam bypasses the neutralizing antibody response, thus allowing multiple boosting. This formulation is also helpful for inducing an antigen-specific immune response in the presence of self-antigen, and maintains long-term effector CD8 + T cells. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Time-specific blockade of PDGFR with Imatinib (Glivec®) causes cataract and disruption of lens fiber cells in neonatal mice.

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Zhou, Yin-Pin; He, Yang-Tao; Chen, Cheng-Li; Ji, Jun; Niu, Jian-Qin; Wang, Han-Zhi; Li, Shi-Feng; Huang, Lan; Mei, Feng

2011-03-01

This study aimed at investigating the response of lens epithelial cells in postnatal mice to Imatinib (Glivec®, a potent inhibitor of platelet-derived growth factor receptor (PDGFR)) treatment. Mouse eyes were sampled 10 days after administration of Imatinib (0.5 mg·g(-1)·day(-1)) for 3 days, at either 7, 14, or 21 days postpartum. Structural changes of lens were revealed by routine H.E. staining. Levels of proliferation and apoptosis were revealed by BrdU incorporation and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, respectively, and immunofluorescent staining with anti-PDGFRα antibody was carried out on the sections of eyeball. PDGFRα and p-PDGFRαprotein levels were evaluated by Western blot. Our results indicated that administration of Imatinib led to blockade of PDGFR signaling. Formation of cataracts was found only in those mice where treatment started from 7 days postpartum (P7), but was not observed in those samples from P14 nor P21. Fiber cells were disorganized in cataract lens core as observed histologically, and migration of epithelial cells was also inhibited. No apoptosis was detected with the TUNEL method. Our results indicated blockade of PDGFR at the neonatal stage (P7) would lead to cataracts and lens fiber cells disorganization, suggesting that PDGFR signaling plays a time-specific and crucial role in the postnatal development of lens in the mouse, and also may provide a new approach to produce a congenital cataract animal model.

In Vivo Imaging of Prostate Cancer Tumors and Metastasis Using Non-Specific Fluorescent Nanoparticles in Mice

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Coralie Genevois

2017-12-01

Full Text Available With the growing interest in the use of nanoparticles (NPs in nanomedicine, there is a crucial need for imaging and targeted therapies to determine NP distribution in the body after systemic administration, and to achieve strong accumulation in tumors with low background in other tissues. Accumulation of NPs in tumors results from different mechanisms, and appears extremely heterogeneous in mice models and rather limited in humans. Developing new tumor models in mice, with their low spontaneous NP accumulation, is thus necessary for screening imaging probes and for testing new targeting strategies. In the present work, accumulation of LipImageTM 815, a non-specific nanosized fluorescent imaging agent, was compared in subcutaneous, orthotopic and metastatic tumors of RM1 cells (murine prostate cancer cell line by in vivo and ex vivo fluorescence imaging techniques. LipImageTM 815 mainly accumulated in liver at 24 h but also in orthotopic tumors. Limited accumulation occurred in subcutaneous tumors, and very low fluorescence was detected in metastasis. Altogether, these different tumor models in mice offered a wide range of NP accumulation levels, and a panel of in vivo models that may be useful to further challenge NP targeting properties.

Lack of significant metabolic abnormalities in mice with liver-specific disruption of 11β-hydroxysteroid dehydrogenase type 1.

LENUS (Irish Health Repository)

Lavery, Gareth G

2012-07-01

Glucocorticoids (GC) are implicated in the development of metabolic syndrome, and patients with GC excess share many clinical features, such as central obesity and glucose intolerance. In patients with obesity or type 2 diabetes, systemic GC concentrations seem to be invariably normal. Tissue GC concentrations determined by the hypothalamic-pituitary-adrenal (HPA) axis and local cortisol (corticosterone in mice) regeneration from cortisone (11-dehydrocorticosterone in mice) by the 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme, principally expressed in the liver. Transgenic mice have demonstrated the importance of 11β-HSD1 in mediating aspects of the metabolic syndrome, as well as HPA axis control. In order to address the primacy of hepatic 11β-HSD1 in regulating metabolism and the HPA axis, we have generated liver-specific 11β-HSD1 knockout (LKO) mice, assessed biomarkers of GC metabolism, and examined responses to high-fat feeding. LKO mice were able to regenerate cortisol from cortisone to 40% of control and had no discernible difference in a urinary metabolite marker of 11β-HSD1 activity. Although circulating corticosterone was unaltered, adrenal size was increased, indicative of chronic HPA stimulation. There was a mild improvement in glucose tolerance but with insulin sensitivity largely unaffected. Adiposity and body weight were unaffected as were aspects of hepatic lipid homeostasis, triglyceride accumulation, and serum lipids. Additionally, no changes in the expression of genes involved in glucose or lipid homeostasis were observed. Liver-specific deletion of 11β-HSD1 reduces corticosterone regeneration and may be important for setting aspects of HPA axis tone, without impacting upon urinary steroid metabolite profile. These discordant data have significant implications for the use of these biomarkers of 11β-HSD1 activity in clinical studies. The paucity of metabolic abnormalities in LKO points to important compensatory effects by HPA

Cardiomyocyte-Specific Ablation of Med1 Subunit of the Mediator Complex Causes Lethal Dilated Cardiomyopathy in Mice.

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Jia, Yuzhi; Chang, Hsiang-Chun; Schipma, Matthew J; Liu, Jing; Shete, Varsha; Liu, Ning; Sato, Tatsuya; Thorp, Edward B; Barger, Philip M; Zhu, Yi-Jun; Viswakarma, Navin; Kanwar, Yashpal S; Ardehali, Hossein; Thimmapaya, Bayar; Reddy, Janardan K

2016-01-01

Mediator, an evolutionarily conserved multi-protein complex consisting of about 30 subunits, is a key component of the polymerase II mediated gene transcription. Germline deletion of the Mediator subunit 1 (Med1) of the Mediator in mice results in mid-gestational embryonic lethality with developmental impairment of multiple organs including heart. Here we show that cardiomyocyte-specific deletion of Med1 in mice (csMed1-/-) during late gestational and early postnatal development by intercrossing Med1fl/fl mice to α-MyHC-Cre transgenic mice results in lethality within 10 days after weaning due to dilated cardiomyopathy-related ventricular dilation and heart failure. The csMed1-/- mouse heart manifests mitochondrial damage, increased apoptosis and interstitial fibrosis. Global gene expression analysis revealed that loss of Med1 in heart down-regulates more than 200 genes including Acadm, Cacna1s, Atp2a2, Ryr2, Pde1c, Pln, PGC1α, and PGC1β that are critical for calcium signaling, cardiac muscle contraction, arrhythmogenic right ventricular cardiomyopathy, dilated cardiomyopathy and peroxisome proliferator-activated receptor regulated energy metabolism. Many genes essential for oxidative phosphorylation and proper mitochondrial function such as genes coding for the succinate dehydrogenase subunits of the mitochondrial complex II are also down-regulated in csMed1-/- heart contributing to myocardial injury. Data also showed up-regulation of about 180 genes including Tgfb2, Ace, Atf3, Ctgf, Angpt14, Col9a2, Wisp2, Nppa, Nppb, and Actn1 that are linked to cardiac muscle contraction, cardiac hypertrophy, cardiac fibrosis and myocardial injury. Furthermore, we demonstrate that cardiac specific deletion of Med1 in adult mice using tamoxifen-inducible Cre approach (TmcsMed1-/-), results in rapid development of cardiomyopathy and death within 4 weeks. We found that the key findings of the csMed1-/- studies described above are highly reproducible in TmcsMed1-/- mouse heart

Specific Physical Exercise Improves Energetic Metabolism in the Skeletal Muscle of Amyotrophic-Lateral- Sclerosis Mice

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Céline Desseille

2017-10-01

Full Text Available Amyotrophic Lateral Sclerosis is an adult-onset neurodegenerative disease characterized by the specific loss of motor neurons, leading to muscle paralysis and death. Although the cellular mechanisms underlying amyotrophic lateral sclerosis (ALS-induced toxicity for motor neurons remain poorly understood, growing evidence suggest a defective energetic metabolism in skeletal muscles participating in ALS-induced motor neuron death ultimately destabilizing neuromuscular junctions. In the present study, we report that a specific exercise paradigm, based on a high intensity and amplitude swimming exercise, significantly improves glucose metabolism in ALS mice. Using physiological tests and a biophysics approach based on nuclear magnetic resonance (NMR, we unexpectedly found that SOD1(G93A ALS mice suffered from severe glucose intolerance, which was counteracted by high intensity swimming but not moderate intensity running exercise. Furthermore, swimming exercise restored the highly ALS-sensitive tibialis muscle through an autophagy-linked mechanism involving the expression of key glucose transporters and metabolic enzymes, including GLUT4 and glyceraldehyde-3-phosphate dehydrogenase (GAPDH. Importantly, GLUT4 and GAPDH expression defects were also found in muscles from ALS patients. Moreover, we report that swimming exercise induced a triglyceride accumulation in ALS tibialis, likely resulting from an increase in the expression levels of lipid transporters and biosynthesis enzymes, notably DGAT1 and related proteins. All these data provide the first molecular basis for the differential effects of specific exercise type and intensity in ALS, calling for the use of physical exercise as an appropriate intervention to alleviate symptoms in this debilitating disease.

Cyclooxygenase-2 mediates the febrile response of mice to interleukin-1beta.

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Li, S; Ballou, L R; Morham, S G; Blatteis, C M

2001-08-10

Various lines of evidence have implicated cyclooxygenase (COX)-2 as a modulator of the fever induced by the exogenous pyrogen lipopolysaccharide (LPS). Thus, treatment with specific inhibitors of COX-2 suppresses the febrile response without affecting basal body (core) temperature (T(c)). Furthermore, COX-2 gene-ablated mice are unable to develop a febrile response to intraperitoneal (i.p.) LPS, whereas their COX-1-deficient counterparts produce fevers not different from their wild-type (WT) controls. To extend the apparently critical role of COX-2 for LPS-induced fevers to fevers produced by endogenous pyrogens, we studied the thermal responses of COX-1- and COX-2 congenitally deficient mice to i.p. and intracerebroventricular (i.c.v.) injections of recombinant murine (rm) interleukin (IL)-1beta. We also assessed the effects of one selective COX-1 inhibitor, SC-560, and two selective COX-2 inhibitors, nimesulide (NIM) and dimethylfuranone (DFU), on the febrile responses of WT and COX-1(-/-) mice to LPS and rmIL-1beta, i.p. Finally, we verified the integrity of the animals' responses to PGE2, i.c.v. I.p. and i.c.v. rmIL-1beta induced similar fevers in WT and COX-1 knockout mice, but provoked no rise in the T(c)s of COX-2 null mutants. The fever produced in WT mice by i.p. LPS was not affected by SC-560, but it was attenuated and abolished by NIM and DFU, respectively, while that caused by i.p. rmIL-1beta was converted into a T(c) fall by DFU. There were no differences in the responses to i.c.v. PGE2 among the WT and COX knockout mice. These results, therefore, further support the notion that the production of PGE2 in response to pyrogens is critically dependent on COX-2 expression.

Non-specific immunization against babesiosis

International Nuclear Information System (INIS)

Cox, F.E.G.

1980-01-01

The rodent babesias, Babesia rodhaini and the less virulent B. microti, are useful models with which to study immunity to and immunization against babesiosis. In contrast with the difficulty in inducing specific immunity to these parasites it is comparatively easy to induce non-specific immunity by prior exposure to related and unrelated intra-erythrocytic protozoa, micro-organisms such as Mycobacterium bovis (BCG) and Corynebacterium parvum, microbial extracts and muramyl dipeptide. This non-specific immunity is long lasting and extremely effective. It is characterized by the facts that (a) it occurs early in the infection at the height of the first peak of parasitaemia, and (b) it involves the intra-erythrocytic death of the parasites. After the primary parasitaemia has resolved, some parasites continue to persist at a low level and when introduced into clean mice produce only low-level 'attenuated' infections in these. Non-specific immunity is not equally effective in all strains of mice. It is suggested that immunity to babesiosis, and infections caused by other intra-erythrocytic protozoa, involves two mechanisms, the first non-specific and the second specific. The actual balance between these two mechanisms varies from parasite to parasite and from host to host. An effective vaccine would have to be based on an understanding of the roles of non-specific immunity in the actual disease under consideration, and would ideally combine an adjuvant that would also stimulate non-specific immunity and an attenuated strain of parasite that would induce a specific response. (author)

Spontaneous Pancreatitis Caused by Tissue-Specific Gene Ablation of Hhex in MiceSummary

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Mark J. Ferreira

2015-09-01

Full Text Available Background & Aims: Perturbations in pancreatic ductal bicarbonate secretion cause chronic pancreatitis. The physiologic mechanism of ductal secretion is known, but its transcriptional control is not. We determine the role of the transcription factor hematopoietically expressed homeobox protein (Hhex in ductal secretion and pancreatitis. Methods: We derived mice with pancreas-specific, Cre-mediated Hhex gene ablation to determine the requirement of Hhex in the pancreatic duct in early life and in adult stages. Histologic and immunostaining analyses were used to detect the presence of pathology. Pancreatic primary ductal cells were isolated to discover differentially expressed transcripts upon acute Hhex ablation on a cell autonomous level. Results: Hhex protein was detected throughout the embryonic and adult ductal trees. Ablation of Hhex in pancreatic progenitors resulted in postnatal ductal ectasia associated with acinar-to-ductal metaplasia, a progressive phenotype that ultimately resulted in chronic pancreatitis. Hhex ablation in adult mice, however, did not cause any detectable pathology. Ductal ectasia in young mice did not result from perturbation of expression of Hnf6, Hnf1β, or the primary cilia genes. RNA-seq analysis of Hhex-ablated pancreatic primary ductal cells showed mRNA levels of the G-protein coupled receptor natriuretic peptide receptor 3 (Npr3, implicated in paracrine signaling, up-regulated by 4.70-fold. Conclusions: Although Hhex is dispensable for ductal cell function in the adult, ablation of Hhex in pancreatic progenitors results in pancreatitis. Our data highlight the critical role of Hhex in maintaining ductal homeostasis in early life and support ductal hypersecretion as a novel etiology of pediatric chronic pancreatitis. Keywords: Npr3, Pancreatic Ducts, Primary Cilia

Kidney epithelium specific deletion of kelch-like ECH-associated protein 1 (Keap1) causes hydronephrosis in mice.

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Noel, Sanjeev; Arend, Lois J; Bandapalle, Samatha; Reddy, Sekhar P; Rabb, Hamid

2016-08-02

Transcription factor Nrf2 protects from experimental acute kidney injury (AKI) and is promising to limit progression in human chronic kidney disease (CKD) by upregulating multiple antioxidant genes. We recently demonstrated that deletion of Keap1, the endogenous inhibitor of Nrf2, in T lymphocytes significantly protects from AKI. In this study, we investigated the effect of Keap1 deletion on Nrf2 mediated antioxidant response in the renal tubular epithelial cells. We deleted Keap1 exon 2 and 3 in the renal tubular epithelial cells by crossing Ksp-Cre mice with Keap1 floxed (Keap1 (f/f)) mice. Deletion of Keap1 gene in the kidney epithelial cells of Ksp-Keap1 (-/-) mice and its effect on Nrf2 target gene expression was performed using PCR and real-time PCR respectively. Histological evaluation was performed on H&E stained sections. Complete blood count, serum and urine analysis were performed to assess systemic effects of defective kidney development. Student's T test was used to determine statistical difference between the groups. Ksp-Cre resulted in the deletion of Keap1 exon 2 and 3 and subsequent upregulation of Nrf2 target genes, Nqo1, Gclm and Gclc in the kidney epithelial cells of Ksp-Keap1 (-/-) mice at baseline. Renal epithelial cell specific deletion of Keap1 in Ksp-Keap1 (-/-) mice caused marked renal pelvic expansion and significant compression of medullary parenchyma consistent with hydronephrosis in both (3 month-old) males and females. Kidneys from 6 month-old Ksp-Keap1 (-/-) mice showed progressive hydronephrosis. Hematological, biochemical and urinary analysis showed significantly higher red blood cell count (p = 0.04), hemoglobin (p = 0.01), hematocrit (p = 0.02), mean cell volume (p = 0.02) and mean cell hemoglobin concentration (p = 0.003) in Ksp-Keap1 (-/-) mice in comparison to Keap1 (f/f) mice. These unexpected findings demonstrate that Keap1 deletion in renal tubular epithelial cells results in an abnormal kidney

Insulin, IGF-1, and GH Receptors Are Altered in an Adipose Tissue Depot-Specific Manner in Male Mice With Modified GH Action.

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Hjortebjerg, Rikke; Berryman, Darlene E; Comisford, Ross; Frank, Stuart J; List, Edward O; Bjerre, Mette; Frystyk, Jan; Kopchick, John J

2017-05-01

Growth hormone (GH) is a determinant of glucose homeostasis and adipose tissue (AT) function. Using 7-month-old transgenic mice expressing the bovine growth hormone (bGH) gene and growth hormone receptor knockout (GHR-/-) mice, we examined whether changes in GH action affect glucose, insulin, and pyruvate tolerance and AT expression of proteins involved in the interrelated signaling pathways of GH, insulinlike growth factor 1 (IGF-1), and insulin. Furthermore, we searched for AT depot-specific differences in control mice. Glycated hemoglobin levels were reduced in bGH and GHR-/- mice, and bGH mice displayed impaired gluconeogenesis as judged by pyruvate tolerance testing. Serum IGF-1 was elevated by 90% in bGH mice, whereas IGF-1 and insulin were reduced by 97% and 61% in GHR-/- mice, respectively. Igf1 RNA was increased in subcutaneous, epididymal, retroperitoneal, and brown adipose tissue (BAT) depots in bGH mice (mean increase ± standard error of the mean in all five depots, 153% ± 27%) and decreased in all depots in GHR-/- mice (mean decrease, 62% ± 4%). IGF-1 receptor expression was decreased in all AT depots of bGH mice (mean decrease, 49% ± 6%) and increased in all AT depots of GHR-/- mice (mean increase, 94% ± 8%). Insulin receptor expression was reduced in retroperitoneal, mesenteric, and BAT depots in bGH mice (mean decrease in all depots, 56% ± 4%) and augmented in subcutaneous, retroperitoneal, mesenteric, and BAT depots in GHR-/- mice (mean increase: 51% ± 1%). Collectively, our findings indicate a role for GH in influencing hormone signaling in AT in a depot-dependent manner. Copyright © 2017 Endocrine Society.

HdhQ111 Mice Exhibit Tissue Specific Metabolite Profiles that Include Striatal Lipid Accumulation

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Carroll, Jeffrey B.; Deik, Amy; Fossale, Elisa; Weston, Rory M.; Guide, Jolene R.; Arjomand, Jamshid; Kwak, Seung; Clish, Clary B.; MacDonald, Marcy E.

2015-01-01

The HTT CAG expansion mutation causes Huntington’s Disease and is associated with a wide range of cellular consequences, including altered metabolism. The mutant allele is expressed widely, in all tissues, but the striatum and cortex are especially vulnerable to its effects. To more fully understand this tissue-specificity, early in the disease process, we asked whether the metabolic impact of the mutant CAG expanded allele in heterozygous B6.HdhQ111/+ mice would be common across tissues, or whether tissues would have tissue-specific responses and whether such changes may be affected by diet. Specifically, we cross-sectionally examined steady state metabolite concentrations from a range of tissues (plasma, brown adipose tissue, cerebellum, striatum, liver, white adipose tissue), using an established liquid chromatography-mass spectrometry pipeline, from cohorts of 8 month old mutant and wild-type littermate mice that were fed one of two different high-fat diets. The differential response to diet highlighted a proportion of metabolites in all tissues, ranging from 3% (7/219) in the striatum to 12% (25/212) in white adipose tissue. By contrast, the mutant CAG-expanded allele primarily affected brain metabolites, with 14% (30/219) of metabolites significantly altered, compared to wild-type, in striatum and 11% (25/224) in the cerebellum. In general, diet and the CAG-expanded allele both elicited metabolite changes that were predominantly tissue-specific and non-overlapping, with evidence for mutation-by-diet interaction in peripheral tissues most affected by diet. Machine-learning approaches highlighted the accumulation of diverse lipid species as the most genotype-predictive metabolite changes in the striatum. Validation experiments in cell culture demonstrated that lipid accumulation was also a defining feature of mutant HdhQ111 striatal progenitor cells. Thus, metabolite-level responses to the CAG expansion mutation in vivo were tissue specific and most evident

HdhQ111 Mice Exhibit Tissue Specific Metabolite Profiles that Include Striatal Lipid Accumulation.

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Jeffrey B Carroll

Full Text Available The HTT CAG expansion mutation causes Huntington's Disease and is associated with a wide range of cellular consequences, including altered metabolism. The mutant allele is expressed widely, in all tissues, but the striatum and cortex are especially vulnerable to its effects. To more fully understand this tissue-specificity, early in the disease process, we asked whether the metabolic impact of the mutant CAG expanded allele in heterozygous B6.HdhQ111/+ mice would be common across tissues, or whether tissues would have tissue-specific responses and whether such changes may be affected by diet. Specifically, we cross-sectionally examined steady state metabolite concentrations from a range of tissues (plasma, brown adipose tissue, cerebellum, striatum, liver, white adipose tissue, using an established liquid chromatography-mass spectrometry pipeline, from cohorts of 8 month old mutant and wild-type littermate mice that were fed one of two different high-fat diets. The differential response to diet highlighted a proportion of metabolites in all tissues, ranging from 3% (7/219 in the striatum to 12% (25/212 in white adipose tissue. By contrast, the mutant CAG-expanded allele primarily affected brain metabolites, with 14% (30/219 of metabolites significantly altered, compared to wild-type, in striatum and 11% (25/224 in the cerebellum. In general, diet and the CAG-expanded allele both elicited metabolite changes that were predominantly tissue-specific and non-overlapping, with evidence for mutation-by-diet interaction in peripheral tissues most affected by diet. Machine-learning approaches highlighted the accumulation of diverse lipid species as the most genotype-predictive metabolite changes in the striatum. Validation experiments in cell culture demonstrated that lipid accumulation was also a defining feature of mutant HdhQ111 striatal progenitor cells. Thus, metabolite-level responses to the CAG expansion mutation in vivo were tissue specific and

Leukemogenic Ptpn11 allele causes defective erythropoiesis in mice.

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Tatiana Usenko

Full Text Available Src homology 2 (SH2 domain-containing phosphatase 2 (SHP2, encoded by PTPN11, regulates signaling networks and cell fate in many tissues. Expression of oncogenic PTPN11 in the hematopoietic compartment causes myeloproliferative neoplasm (MPN in humans and mice. However, the stage-specific effect(s of mutant Ptpn11 on erythroid development have remained unknown. We found that expression of an activated, leukemogenic Ptpn11 allele, Ptpn11D61Y, specifically in the erythroid lineage causes dyserythropoiesis in mice. Ptpn11D61Y progenitors produce excess cKIT+ CD71+ Ter119- cells and aberrant numbers of cKITl° CD71+ erythroblasts. Mutant erythroblasts show elevated activation of ERK, AKT and STAT3 in response to EPO stimulation, and MEK inhibitor treatment blocks Ptpn11D61Y-evoked erythroid hyperproliferation in vitro. Thus, the expression of oncogenic Ptpn11 causes dyserythropoiesis in a cell-autonomous manner in vivo.

RhD Specific Antibodies Are Not Detectable in HLA-DRB11501* Mice Challenged with Human RhD Positive Erythrocytes

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Lidice Bernardo

2014-01-01

Full Text Available The ability to study the immune response to the RhD antigen in the prevention of hemolytic disease of the fetus and newborn has been hampered by the lack of a mouse model of RhD immunization. However, the ability of transgenic mice expressing human HLA DRB11501* to respond to immunization with purified RhD has allowed this question to be revisited. In this work we aimed at inducing anti-RhD antibodies by administering human RhD+ RBCs to mice transgenic for the human HLA DRB11501* as well as to several standard inbred and outbred laboratory strains including C57BL/6, DBA1/J, CFW(SW, CD1(ICR, and NSA(CF-1. DRB11501* mice were additionally immunized with putative extracellular immunogenic RhD peptides. DRB11501* mice immunized with RhD+ erythrocytes developed an erythrocyte-reactive antibody response. Antibodies specific for RhD could not however be detected by flow cytometry. Despite this, DRB11501* mice were capable of recognizing immunogenic sequences of Rh as injection with Rh peptides induced antibodies reactive with RhD sequences, consistent with the presence of B cell repertoires capable of recognizing RhD. We conclude that while HLA DRB11501* transgenic mice may have the capability of responding to immunogenic sequences within RhD, an immune response to human RBC expressing RhD is not directly observed.

Bone marrow function. I. Peripheral T cells are responsible for the increased auto-antiidiotype response of older mice

International Nuclear Information System (INIS)

Kim, Y.T.; Goidl, E.A.; Samarut, C.; Weksler, M.E.; Thorbecke, G.J.; Siskind, G.W.

1985-01-01

After immunization with trinitrophenyl (TNP)-Ficoll, mice produced both anti-TNP antibodies and auto-anti-idiotype (auto-anti-Id) antibodies specific for the anti-TNP antibody. Older animals produced more auto-anti-Id than did young animals. When mice were exposed to a normally lethal dose of irradiation while their bone marrow (BM) was partially shielded, they survived and slowly (6 wk) regained immune function, as indicated by the number of nucleated cells in their spleen and the in vitro primary plaque-forming cell (PFC) response of their spleen cells to TNP-treated aminoethylated polyacrylamide beads. Recovery is presumably the result of repopulation of the peripheral lymphoid system by cells originating in the BM. By enzyme-linked immunosorbent assay (ELISA), and by hapten-augmentable PFC assay, the authors show that, after recovery from irradiation with their BM shielded, old animals produce low auto-anti-Id responses, like those of young animals. The transfer of splenic T cells into mice irradiated with their BM shielded provided evidence that the magnitude of the auto-anti-Id response is controlled by the peripheral T cells. Thus, mice that received splenic T cells from aged donors produced high levels of auto-anti-Id while those that received splenic T cells from young donors produce low levels of auto-anti-Id

Diminished primary and secondary influenza virus-specific CD8(+) T-cell responses in CD4-depleted Ig(-/-) mice

DEFF Research Database (Denmark)

Riberdy, J M; Christensen, Jan Pravsgaard; Branum, K

2000-01-01

Optimal expansion of influenza virus nucleoprotein (D(b)NP(366))-specific CD8(+) T cells following respiratory challenge of naive Ig(-/-) microMT mice was found to require CD4(+) T-cell help, and this effect was also observed in primed animals. Absence of the CD4(+) population was consistently...

Immunizations with hepatitis B viral antigens and a TLR7/8 agonist adjuvant induce antigen-specific immune responses in HBV-transgenic mice

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Ying Wang

2014-12-01

Conclusions: Immunization with CL097-conjugated HBV-Ag reversed immune tolerance in HBV-Tg mice and induced antigen-specific immune responses. TLR7/8 agonists appear to be potent adjuvants for the induction of antigen-specific Th1 responses in an immune tolerant state.

Microbiota-Derived Metabolic Factors Reduce Campylobacteriosis in Mice.

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Sun, Xiaolun; Winglee, Kathryn; Gharaibeh, Raad Z; Gauthier, Josee; He, Zhen; Tripathi, Prabhanshu; Avram, Dorina; Bruner, Steven; Fodor, Anthony; Jobin, Christian

2018-05-01

Campylobacter jejuni, a prevalent foodborne bacterial pathogen, exploits the host innate response to induce colitis. Little is known about the roles of microbiota in C jejuni-induced intestinal inflammation. We investigated interactions between microbiota and intestinal cells during C jejuni infection of mice. Germ-free C57BL/6 Il10 -/- mice were colonized with conventional microbiota and infected with a single dose of C jejuni (10 9 colony-forming units/mouse) via gavage. Conventional microbiota were cultured under aerobic, microaerobic, or anaerobic conditions and orally transplanted into germ-free Il10 -/- mice. Colon tissues were collected from mice and analyzed by histology, real-time polymerase chain reaction, and immunoblotting. Fecal microbiota and bile acids were analyzed with 16S sequencing and high-performance liquid chromatography with mass spectrometry, respectively. Introduction of conventional microbiota reduced C jejuni-induced colitis in previously germ-free Il10 -/- mice, independent of fecal load of C jejuni, accompanied by reduced activation of mammalian target of rapamycin. Microbiota transplantation and 16S ribosomal DNA sequencing experiments showed that Clostridium XI, Bifidobacterium, and Lactobacillus were enriched in fecal samples from mice colonized with microbiota cultured in anaerobic conditions (which reduce colitis) compared with mice fed microbiota cultured under aerobic conditions (susceptible to colitis). Oral administration to mice of microbiota-derived secondary bile acid sodium deoxycholate, but not ursodeoxycholic acid or lithocholic acid, reduced C jejuni-induced colitis. Depletion of secondary bile acid-producing bacteria with antibiotics that kill anaerobic bacteria (clindamycin) promoted C jejuni-induced colitis in specific pathogen-free Il10 -/- mice compared with the nonspecific antibiotic nalidixic acid; colitis induction by antibiotics was associated with reduced level of luminal deoxycholate. We identified a

Loss of hypothalamic corticotropin-releasing hormone markedly reduces anxiety behaviors in mice

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Zhang, Rong; Asai, Masato; Mahoney, Carrie E; Joachim, Maria; Shen, Yuan; Gunner, Georgia; Majzoub, Joseph A

2016-01-01

A long-standing paradigm posits that hypothalamic corticotropin-releasing hormone (CRH) regulates neuroendocrine functions such as adrenal glucocorticoid release, while extra-hypothalamic CRH plays a key role in stressor-triggered behaviors. Here we report that hypothalamus-specific Crh knockout mice (Sim1CrhKO mice, created by crossing Crhflox with Sim1Cre mice) have absent Crh mRNA and peptide mainly in the paraventricular nucleus of the hypothalamus (PVH) but preserved Crh expression in other brain regions including amygdala and cerebral cortex. As expected, Sim1CrhKO mice exhibit adrenal atrophy as well as decreased basal, diurnal and stressor-stimulated plasma corticosterone secretion and basal plasma ACTH, but surprisingly, have a profound anxiolytic phenotype when evaluated using multiple stressors including open field, elevated plus maze, holeboard, light-dark box, and novel object recognition task. Restoring plasma corticosterone did not reverse the anxiolytic phenotype of Sim1CrhKO mice. Crh-Cre driver mice revealed that PVHCrh fibers project abundantly to cingulate cortex and the nucleus accumbens shell, and moderately to medial amygdala, locus coeruleus, and solitary tract, consistent with the existence of PVHCrh-dependent behavioral pathways. Although previous, nonselective attenuation of CRH production or action, genetically in mice and pharmacologically in humans, respectively, has not produced the anticipated anxiolytic effects, our data show that targeted interference specifically with hypothalamic Crh expression results in anxiolysis. Our data identify neurons that express both Sim1 and Crh as a cellular entry point into the study of CRH-mediated, anxiety-like behaviors and their therapeutic attenuation. PMID:27595593

Plasmid DNA initiates replication of yellow fever vaccine in vitro and elicits virus-specific immune response in mice

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Tretyakova, Irina; Nickols, Brian; Hidajat, Rachmat; Jokinen, Jenny; Lukashevich, Igor S.; Pushko, Peter

2014-01-01

Yellow fever (YF) causes an acute hemorrhagic fever disease in tropical Africa and Latin America. To develop a novel experimental YF vaccine, we applied iDNA infectious clone technology. The iDNA represents plasmid that encodes the full-length RNA genome of 17D vaccine downstream from a cytomegalovirus (CMV) promoter. The vaccine was designed to transcribe the full-length viral RNA and to launch 17D vaccine virus in vitro and in vivo. Transfection with 10 ng of iDNA plasmid was sufficient to start replication of vaccine virus in vitro. Safety of the parental 17D and iDNA-derived 17D viruses was confirmed in AG129 mice deficient in receptors for IFN-α/β/γ. Finally, direct vaccination of BALB/c mice with a single 20 μg dose of iDNA plasmid resulted in seroconversion and elicitation of virus-specific neutralizing antibodies in animals. We conclude that iDNA immunization approach combines characteristics of DNA and attenuated vaccines and represents a promising vaccination strategy for YF. - Highlights: • The iDNA ® platform combines advantages of DNA and live attenuated vaccines. • Yellow fever (YF) 17D vaccine was launched from iDNA plasmid in vitro and in vivo. • Safety of iDNA-generated 17D virus was confirmed in AG129 mice. • BALB/c mice seroconverted after a single-dose vaccination with iDNA. • YF virus-neutralizing response was elicited in iDNA-vaccinated mice

Plasmid DNA initiates replication of yellow fever vaccine in vitro and elicits virus-specific immune response in mice

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Tretyakova, Irina; Nickols, Brian; Hidajat, Rachmat [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States); Jokinen, Jenny; Lukashevich, Igor S. [Department of Pharmacology and Toxicology, School of Medicine, Center for Predictive Medicine and Emerging Infectious Diseases, University of Louisville, Louisville, KY (United States); Pushko, Peter, E-mail: ppushko@medigen-usa.com [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States)

2014-11-15

Yellow fever (YF) causes an acute hemorrhagic fever disease in tropical Africa and Latin America. To develop a novel experimental YF vaccine, we applied iDNA infectious clone technology. The iDNA represents plasmid that encodes the full-length RNA genome of 17D vaccine downstream from a cytomegalovirus (CMV) promoter. The vaccine was designed to transcribe the full-length viral RNA and to launch 17D vaccine virus in vitro and in vivo. Transfection with 10 ng of iDNA plasmid was sufficient to start replication of vaccine virus in vitro. Safety of the parental 17D and iDNA-derived 17D viruses was confirmed in AG129 mice deficient in receptors for IFN-α/β/γ. Finally, direct vaccination of BALB/c mice with a single 20 μg dose of iDNA plasmid resulted in seroconversion and elicitation of virus-specific neutralizing antibodies in animals. We conclude that iDNA immunization approach combines characteristics of DNA and attenuated vaccines and represents a promising vaccination strategy for YF. - Highlights: • The iDNA{sup ®} platform combines advantages of DNA and live attenuated vaccines. • Yellow fever (YF) 17D vaccine was launched from iDNA plasmid in vitro and in vivo. • Safety of iDNA-generated 17D virus was confirmed in AG129 mice. • BALB/c mice seroconverted after a single-dose vaccination with iDNA. • YF virus-neutralizing response was elicited in iDNA-vaccinated mice.

Podocyte specific knock out of selenoproteins does not enhance nephropathy in streptozotocin diabetic C57BL/6 mice

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Carlson Bradley A

2008-07-01

Full Text Available Abstract Background Selenoproteins contain selenocysteine (Sec, commonly considered the 21st genetically encoded amino acid. Many selenoproteins, such as the glutathione peroxidases and thioredoxin reductases, protect cells against oxidative stress by functioning as antioxidants and/or through their roles in the maintenance of intracellular redox balance. Since oxidative stress has been implicated in the pathogenesis of diabetic nephropathy, we hypothesized that selenoproteins protect against this complication of diabetes. Methods C57BL/6 mice that have a podocyte-specific inability to incorporate Sec into proteins (denoted in this paper as PodoTrsp-/- and control mice were made diabetic by intraperitoneal injection of streptozotocin, or were injected with vehicle. Blood glucose, body weight, microalbuminuria, glomerular mesangial matrix expansion, and immunohistochemical markers of oxidative stress were assessed. Results After 3 and 6 months of diabetes, control and PodoTrsp-/- mice had similar levels of blood glucose. There were no differences in urinary albumin/creatinine ratios. Periodic acid-Schiff staining to examine mesangial matrix expansion also demonstrated no difference between control and PodoTrsp-/- mice after 6 months of diabetes, and there were no differences in immunohistochemical stainings for nitrotyrosine or NAD(PH dehydrogenase, quinone 1. Conclusion Loss of podocyte selenoproteins in streptozotocin diabetic C57BL/6 mice does not lead to increased oxidative stress as assessed by nitrotyrosine and NAD(PH dehydrogenase, quinone 1 immunostaining, nor does it lead to worsening nephropathy.

Age- and region-specific imbalances of basal amino acids and monoamine metabolism in limbic regions of female Fmr1 knock-out mice.

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Gruss, Michael; Braun, Katharina

2004-07-01

The Fragile X syndrome, a common form of mental retardation in humans, originates from the loss of expression of the Fragile X mental retardation gene leading to the absence of the encoded Fragile X mental retardation protein 1 (FMRP). A broad pattern of morphological and behavioral abnormalities is well described for affected humans as well as Fmr1 knock-out mice, a transgenic animal model for the human Fragile X syndrome. In the present study, we examined neurochemical differences between female Fmr1 knock-out and wildtype mice with particular focus on neurotransmission. Significant age- and region-specific differences of basal tissue neurotransmitter and metabolite levels measured by high performance liquid chromatography were found. Those differences were more numerous in juvenile animals (postnatal day (PND) 28-31) compared to adults (postnatal day 209-221). In juvenile female knock-out mice, especially aspartate and taurine were increased in cortical regions, striatum, cerebellum, and brainstem. Furthermore, compared to the wildtype animals, the juvenile knock-out mice displayed an increased level of neuronal inhibition in the hippocampus and brainstem reflected by decreased ratios of (aspartate + glutamate)/(taurine + GABA), as well as an increased dopamine (DA) turnover in cortical regions, striatum, and hippocampus. These results provide the first evidence that the lack of FMRP expression in female Fmr1 knock-out mice is accompanied by age-dependent, region-specific alterations in brain amino acids, and monoamine turnover, which might be related to the reported synaptical and behavioural alterations in these animals.

Acetaminophen-induced liver damage in mice is associated with gender-specific adduction of peroxiredoxin-6

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Isaac Mohar

2014-01-01

Full Text Available The mechanism by which acetaminophen (APAP causes liver damage evokes many aspects drug metabolism, oxidative chemistry, and genetic-predisposition. In this study, we leverage the relative resistance of female C57BL/6 mice to APAP-induced liver damage (AILD compared to male C57BL/6 mice in order to identify the cause(s of sensitivity. Furthermore, we use mice that are either heterozygous (HZ or null (KO for glutamate cysteine ligase modifier subunit (Gclm, in order to titrate the toxicity relative to wild-type (WT mice. Gclm is important for efficient de novo synthesis of glutathione (GSH. APAP (300 mg/kg, ip or saline was administered and mice were collected at 0, 0.5, 1, 2, 6, 12, and 24 h. Male mice showed marked elevation in serum alanine aminotransferase by 6 h. In contrast, female WT and HZ mice showed minimal toxicity at all time points. Female KO mice, however, showed AILD comparable to male mice. Genotype-matched male and female mice showed comparable APAP–protein adducts, with Gclm KO mice sustaining significantly greater adducts. ATP was depleted in mice showing toxicity, suggesting impaired mitochondria function. Indeed, peroxiredoxin-6, a GSH-dependent peroxiredoxin, was preferentially adducted by APAP in mitochondria of male mice but rarely adducted in female mice. These results support parallel mechanisms of toxicity where APAP adduction of peroxiredoxin-6 and sustained GSH depletion results in the collapse of mitochondria function and hepatocyte death. We conclude that adduction of peroxiredoxin-6 sensitizes male C57BL/6 mice to toxicity by acetaminophen.

Subregion-Specific Proteomic Signature in the Hippocampus for Recognition Processes in Adult Mice

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Lukas M. von Ziegler

2018-03-01

Full Text Available Summary: The hippocampal formation is a brain structure essential for higher-order cognitive functions. It has a complex anatomical organization and cellular composition, and hippocampal subregions have different properties and functional roles. In this study, we used SWATH-MS to determine whether the proteomes of hippocampus areas CA1 and CA3 can explain the commonalities or specificities of these subregions in basal conditions and after recognition memory. We show that the proteomes of areas CA1 and CA3 are largely different in basal conditions and that differential changes and dynamics in protein expression are induced in these areas after recognition of an object or object location. While changes are consistent across both recognition paradigms in area CA1, they are not in area CA3, suggesting distinct proteomic responses in areas CA1 and CA3 for memory formation. : How does the proteome differ in hippocampus areas CA1 and CA3? von Ziegler et al. identify the proteomes of areas CA1 and CA3 and characterize their dynamics during different recognition processes in adult mice. Keywords: hippocampus, areas CA1 and CA3, proteome, dynamics, object memory, object location memory, mass spectrometry, SWATH-MS, mice, bioinformatic tools

Genes involved in the astrocyte-neuron lactate shuttle (ANLS) are specifically regulated in cortical astrocytes following sleep deprivation in mice.

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Petit, Jean-Marie; Gyger, Joël; Burlet-Godinot, Sophie; Fiumelli, Hubert; Martin, Jean-Luc; Magistretti, Pierre J

2013-10-01

There is growing evidence indicating that in order to meet the neuronal energy demands, astrocytes provide lactate as an energy substrate for neurons through a mechanism called "astrocyte-neuron lactate shuttle" (ANLS). Since neuronal activity changes dramatically during vigilance states, we hypothesized that the ANLS may be regulated during the sleep-wake cycle. To test this hypothesis we investigated the expression of genes associated with the ANLS specifically in astrocytes following sleep deprivation. Astrocytes were purified by fluorescence-activated cell sorting from transgenic mice expressing the green fluorescent protein (GFP) under the control of the human astrocytic GFAP-promoter. 6-hour instrumental sleep deprivation (TSD). Animal sleep research laboratory. Young (P23-P27) FVB/N-Tg (GFAP-GFP) 14Mes/J (Tg) mice of both sexes and 7-8 week male Tg and FVB/Nj mice. Basal sleep recordings and sleep deprivation achieved using a modified cage where animals were gently forced to move. Since Tg and FVB/Nj mice displayed a similar sleep-wake pattern, we performed a TSD in young Tg mice. Total RNA was extracted from the GFP-positive and GFP-negative cells sorted from cerebral cortex. Quantitative RT-PCR analysis showed that levels of Glut1, α-2-Na/K pump, Glt1, and Ldha mRNAs were significantly increased following TSD in GFP-positive cells. In GFP-negative cells, a tendency to increase, although not significant, was observed for Ldha, Mct2, and α-3-Na/K pump mRNAs. This study shows that TSD induces the expression of genes associated with ANLS specifically in astrocytes, underlying the important role of astrocytes in the maintenance of the neuro-metabolic coupling across the sleep-wake cycle.

Podocyte-specific deletion of Rac1 leads to aggravation of renal injury in STZ-induced diabetic mice

International Nuclear Information System (INIS)

Ishizaka, Masanori; Gohda, Tomohito; Takagi, Miyuki; Omote, Keisuke; Sonoda, Yuji; Oliva Trejo, Juan Alejandro; Asao, Rin; Hidaka, Teruo; Asanuma, Katsuhiko; Horikoshi, Satoshi; Tomino, Yasuhiko

2015-01-01

Rac1, a GTPase of the Rho subfamily, has a crucial role in cytoskeletal architecture, as well as the regulation of cell migration and growth. However, renal injury in mice with podocyte-specific deletion of Rac1 has yet to be elucidated fully due to conflicting findings. Herein, we identified a possible role for Rac1 in podocytes of streptozotocin- (STZ) induced diabetic mice. The urinary albumin/creatinine ratio (ACR) in the knockout (KO) group was significantly higher than that in the wild type (WT) group at any week of age. A more marked ACR increase was observed in STZ/KO group than STZ/WT group, although ACR did increase with weeks of age in both diabetic groups. The kidney sections from diabetic mice revealed a glomerular hypertrophy with mesangial expansion, but there was no appreciable difference in glomerular findings under a light microscope between STZ/WT and STZ/KO mice. However, an electron microscopy analysis revealed that regardless of the presence or absence of diabetes, both KO (KO and STZ/KO) groups had a higher rate of foot process effacement compared with both WT (WT and STZ/WT) groups. The expression levels of the slit diaphragm protein, podocin, was reduced with the induction of diabetes, and the levels in the STZ/KO group experienced a further reduction compared with the STZ/WT group. The number of WT1-positive cells in the STZ/KO group was more significantly decreased than that in the other three groups. In contrast, the numbers of cleaved caspase 3- and TUNEL-positive cells in the glomeruli of the STZ/KO group were more increased than those in the STZ/WT group. Thus, this study provides evidence that podocyte-specific deletion of Rac1 results in morphological alteration in podocytes, and that the induction of apoptosis or decreased expression of the slit diaphragm proteins by hyperglycemic stimuli are associated with the progression of diabetic nephropathy.

Podocyte-specific deletion of Rac1 leads to aggravation of renal injury in STZ-induced diabetic mice

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Ishizaka, Masanori [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Gohda, Tomohito, E-mail: goda@juntendo.ac.jp [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Takagi, Miyuki; Omote, Keisuke; Sonoda, Yuji [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Oliva Trejo, Juan Alejandro [Laboratory for Kidney Research (TMK Project), Medical Innovation Center, Kyoto University Graduate School of Medicine, 53 Shogoin Kawaharacho, Sakyo-ku, Kyoto 606-8397 (Japan); Asao, Rin; Hidaka, Teruo [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Asanuma, Katsuhiko [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Laboratory for Kidney Research (TMK Project), Medical Innovation Center, Kyoto University Graduate School of Medicine, 53 Shogoin Kawaharacho, Sakyo-ku, Kyoto 606-8397 (Japan); Horikoshi, Satoshi [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Tomino, Yasuhiko [Division of Nephrology, Department of Internal Medicine, Juntendo University Faculty of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421 (Japan); Medical Corporation SHOWAKAI, 3-12-12 Nishishinjuku, Shinjuku-ku, Tokyo 160-0023 (Japan)

2015-11-20

Rac1, a GTPase of the Rho subfamily, has a crucial role in cytoskeletal architecture, as well as the regulation of cell migration and growth. However, renal injury in mice with podocyte-specific deletion of Rac1 has yet to be elucidated fully due to conflicting findings. Herein, we identified a possible role for Rac1 in podocytes of streptozotocin- (STZ) induced diabetic mice. The urinary albumin/creatinine ratio (ACR) in the knockout (KO) group was significantly higher than that in the wild type (WT) group at any week of age. A more marked ACR increase was observed in STZ/KO group than STZ/WT group, although ACR did increase with weeks of age in both diabetic groups. The kidney sections from diabetic mice revealed a glomerular hypertrophy with mesangial expansion, but there was no appreciable difference in glomerular findings under a light microscope between STZ/WT and STZ/KO mice. However, an electron microscopy analysis revealed that regardless of the presence or absence of diabetes, both KO (KO and STZ/KO) groups had a higher rate of foot process effacement compared with both WT (WT and STZ/WT) groups. The expression levels of the slit diaphragm protein, podocin, was reduced with the induction of diabetes, and the levels in the STZ/KO group experienced a further reduction compared with the STZ/WT group. The number of WT1-positive cells in the STZ/KO group was more significantly decreased than that in the other three groups. In contrast, the numbers of cleaved caspase 3- and TUNEL-positive cells in the glomeruli of the STZ/KO group were more increased than those in the STZ/WT group. Thus, this study provides evidence that podocyte-specific deletion of Rac1 results in morphological alteration in podocytes, and that the induction of apoptosis or decreased expression of the slit diaphragm proteins by hyperglycemic stimuli are associated with the progression of diabetic nephropathy.

Production of Monoclonal Antibodies Specific for Progesterone, Estradiole by Simultaneous Injection of Different Steroids

OpenAIRE

YÜCEL, Fatima ŞAHİNGÖZ

2014-01-01

We report here the development of hybrid cells producing monoclonal antibodies specific for two different steroid hormones with mixed immunization using hybridoma technology. BALB/c mice were immunized with a mixture of three steroid antigens: progesterone, estradiole and testosterone linked to bovine serum albumine. These mice were used for fusion. In the two fusion experiments, ELISA tests showed that among 645 wells only 2 hybrids reacted with progesterone (MAM 3C2, MAM 3E3) and one o...

Antioxidant rich grape pomace extract suppresses postprandial hyperglycemia in diabetic mice by specifically inhibiting alpha-glucosidase

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Hogan Shelly

2010-08-01

Full Text Available Abstract Background Postprandial hyperglycemia is an early defect of type 2 diabetes and one of primary anti-diabetic targets. Treatment of postprandial hyperglycemia can be achieved by inhibiting intestinal α-glucosidase, the key enzyme for oligosaccharide digestion and further glucose absorption. Grape pomace is winemaking byproduct rich in bioactive food compounds such as phenolic antioxidants. This study evaluated the anti-diabetic potential of two specific grape pomace extracts by determining their antioxidant and anti-postprandial hyperglycemic activities in vitro and in vivo. Methods The extracts of red wine grape pomace (Cabernet Franc and white wine grape pomace (Chardonnay were prepared in 80% ethanol. An extract of red apple pomace was included as a comparison. The radical scavenging activities and phenolic profiles of the pomace extracts were determined through the measurement of oxygen radical absorbance capacity, DPPH radical scavenging activity, total phenolic content and flavonoids. The inhibitory effects of the pomace extracts on yeast and rat intestinal α-glucosidases were determined. Male 6-week old C57BLKS/6NCr mice were treated with streptozocin to induce diabetes. The diabetic mice were then treated with vehicle or the grape pomace extract to determine whether the oral intake of the extract can suppress postprandial hyperglycemia through the inhibition of intestinal α-glucosidases. Results The red grape pomace extract contained significantly higher amounts of flavonoids and phenolic compounds and exerted stronger oxygen radical absorbance capacity than the red apple pomace extract. Both the grape pomace extracts but not the apple pomace extract exerted significant inhibition on intestinal α-glucosidases and the inhibition appears to be specific. In the animal study, the oral intake of the grape pomace extract (400 mg/kg body weight significantly suppressed the postprandial hyperglycemia by 35% in streptozocin

Antioxidant rich grape pomace extract suppresses postprandial hyperglycemia in diabetic mice by specifically inhibiting alpha-glucosidase.

Science.gov (United States)

Hogan, Shelly; Zhang, Lei; Li, Jianrong; Sun, Shi; Canning, Corene; Zhou, Kequan

2010-08-27

Postprandial hyperglycemia is an early defect of type 2 diabetes and one of primary anti-diabetic targets. Treatment of postprandial hyperglycemia can be achieved by inhibiting intestinal α-glucosidase, the key enzyme for oligosaccharide digestion and further glucose absorption. Grape pomace is winemaking byproduct rich in bioactive food compounds such as phenolic antioxidants. This study evaluated the anti-diabetic potential of two specific grape pomace extracts by determining their antioxidant and anti-postprandial hyperglycemic activities in vitro and in vivo. The extracts of red wine grape pomace (Cabernet Franc) and white wine grape pomace (Chardonnay) were prepared in 80% ethanol. An extract of red apple pomace was included as a comparison. The radical scavenging activities and phenolic profiles of the pomace extracts were determined through the measurement of oxygen radical absorbance capacity, DPPH radical scavenging activity, total phenolic content and flavonoids. The inhibitory effects of the pomace extracts on yeast and rat intestinal α-glucosidases were determined. Male 6-week old C57BLKS/6NCr mice were treated with streptozocin to induce diabetes. The diabetic mice were then treated with vehicle or the grape pomace extract to determine whether the oral intake of the extract can suppress postprandial hyperglycemia through the inhibition of intestinal α-glucosidases. The red grape pomace extract contained significantly higher amounts of flavonoids and phenolic compounds and exerted stronger oxygen radical absorbance capacity than the red apple pomace extract. Both the grape pomace extracts but not the apple pomace extract exerted significant inhibition on intestinal α-glucosidases and the inhibition appears to be specific. In the animal study, the oral intake of the grape pomace extract (400 mg/kg body weight) significantly suppressed the postprandial hyperglycemia by 35% in streptozocin-induced diabetic mice following starch challenge. This is the

Glial and tissue-specific regulation of Kynurenine Pathway dioxygenases by acute stress of mice

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Carlos R. Dostal

2017-12-01

Full Text Available Stressors activate the hypothalamic-pituitary-adrenal (HPA axis and immune system eliciting changes in cognitive function, mood and anxiety. An important link between stress and altered behavior is stimulation of the Kynurenine Pathway which generates neuroactive and immunomodulatory kynurenines. Tryptophan entry into this pathway is controlled by rate-limiting indoleamine/tryptophan 2,3-dioxygenases (DOs: Ido1, Ido2, Tdo2. Although implicated as mediating changes in behavior, detecting stress-induced DO expression has proven inconsistent. Thus, C57BL/6J mice were used to characterize DO expression in brain-regions, astrocytes and microglia to characterize restraint-stress-induced DO expression. Stress increased kynurenine in brain and plasma, demonstrating increased DO activity. Of three Ido1 transcripts, only Ido1-v1 expression was increased by stress and within astrocytes, not microglia, indicating transcript- and glial-specificity. Stress increased Ido1-v1 only in frontal cortex and hypothalamus, indicating brain-region specificity. Of eight Ido2 transcripts, Ido2-v3 expression was increased by stress, again only within astrocytes. Likewise, stress increased Tdo2-FL expression in astrocytes, not microglia. Interestingly, Ido2 and Tdo2 transcripts were not correspondingly induced in Ido1-knockout (Ido1KO mice, suggesting that Ido1 is necessary for the central DO response to acute stress. Unlike acute inflammatory models resulting in DO induction within microglia, only astrocyte DO expression was increased by acute restraint-stress, defining their unique role during stress-dependent activation of the Kynurenine Pathway. Keywords: Stress, Ido, Tdo, Kynurenine, Astrocyte, Liver

Whole-Retina Reduced Electrophysiological Activity in Mice Bearing Retina-Specific Deletion of Vesicular Acetylcholine Transporter.

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Jake Bedore

Full Text Available Despite rigorous characterization of the role of acetylcholine in retinal development, long-term effects of its absence as a neurotransmitter are unknown. One of the unanswered questions is how acetylcholine contributes to the functional capacity of mature retinal circuits. The current study investigates the effects of disrupting cholinergic signalling in mice, through deletion of vesicular acetylcholine transporter (VAChT in the developing retina, pigmented epithelium, optic nerve and optic stalk, on electrophysiology and structure of the mature retina.A combination of electroretinography, optical coherence tomography imaging and histological evaluation assessed retinal integrity in mice bearing retina- targeted (embryonic day 12.5 deletion of VAChT (VAChTSix3-Cre-flox/flox and littermate controls at 5 and 12 months of age. VAChTSix3-Cre-flox/flox mice did not show any gross changes in nuclear layer cellularity or synaptic layer thickness. However, VAChTSix3-Cre-flox/flox mice showed reduced electrophysiological response of the retina to light stimulus under scotopic conditions at 5 and 12 months of age, including reduced a-wave, b-wave, and oscillatory potential (OP amplitudes and decreased OP peak power and total energy. Reduced a-wave amplitude was proportional to the reduction in b-wave amplitude and not associated with altered a-wave 10%-90% rise time or inner and outer segment thicknesses.This study used a novel genetic model in the first examination of function and structure of the mature mouse retina with disruption of cholinergic signalling. Reduced amplitude across the electroretinogram wave form does not suggest dysfunction in specific retinal cell types and could reflect underlying changes in the retinal and/or extraretinal microenvironment. Our findings suggest that release of acetylcholine by VAChT is essential for the normal electrophysiological response of the mature mouse retina.

Differential androgenesis in gamma irradiated mice

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Kim, Jihyang; Yoon, Yongdal [Hanyang Univ., Seoul (Korea, Republic of); Kim, Jin Kyu [Korea Atomic Energy Research Institute, Taejon (Korea, Republic of)

2002-07-01

The Leydig cells of the testis account for at least 75% of the total testosterone produced in the normal adult male. Whereas the production of estrogen from androgen is catalyzed by aromatase cytochrome P450, which is found in many tissues, including gonad, brain, adipose tissue, bone, and heart. The gamma-irradiation causes the impairment of spermatogenesis and steroidogenesis in male mice. The present study was performed to analyze changes in testosterone concentrations and expression of steroidogenic enzyme of mice after whole body gamma-irradiation. Eight-week-old male ICR mice were irradiated with 6.5 or 10 Gy. At days 1, 2, 3, 4, and 5 after irradiation, testes were removed and processed for paraffin sections and isolation of mRNA. We calculated the gonad index from body and testis weight, and checked the testis volume. Hormonal analysis was performed by means of radioimmunoassay (RIA) in serum and intratesticular fluid. Semiquantitative reverse transcription polymerase chain reaction (RT-PCR) was used to evaluate the expression kinetics of the apoptotic gene and the cytochrome P450 aromatase gene after irradiation. In gamma-irradiated mice, the body weight reduced in comparison to that of the control group. Therefore, gonad indices increased. The testosterone concentrations in serum and intratesticular fluid were significantly reduced. RT- PCR data represented that the expression of Fas, Fas ligand, and aromatase cytochrome P450 showed the specific patterns against control groups. These results indicated that gamma- irradiation of adult mice induced the alteration of androgenesis and suggested that might counteract the spermatogenesis.

Antibody hyporesponsiveness in resistant BALB/cJ mice intracorneally infected with Pseudomonas aeruginosa.

Science.gov (United States)

Berk, R S; Preston, M; Montgomery, I N; Hazlett, L D; Tse, H Y

Six- to eight-week-old BALB/cJ (and BALB/cPi) mice were found able to restore corneal clarity within 3 to 4 weeks after intracorneal infection with Pseudomonas aeruginosa strain 19660. However, enzyme-linked immunosorbent assay (ELISA) for serum immunoglobulins directed specifically against P. aeruginosa indicated that the mice were initially non- or hypo-responsive for IgG and IgA over the 4-week holding period. Low IgM titers could be detected 1 week after infection, but tended to decrease with time. When the mice were re-infected by using the contralateral control eye, then the serum IgG levels began to gradually increase as the time interval following the secondary infection increased from 1 to 3 weeks. Re-infected mice did not show a significantly increased rate of corneal clarity restoration with time, when compared to the corneas of mice receiving only a primary infection despite the presence of serum antibodies specific to P. aeruginosa. When congenic mice of the BALB/c background carrying the DBA/2N Idh/Pep-3 locus found on chromosome 1 were intracorneally infected, they tended to restore corneal clarity at approximately the same rate as the BALB/cJ mice. However, the congenic mice mounted a faster and substantially greater magnitude of serum IgM and IgG response during the primary infection than BALB/cJ mice receiving either a primary and/or secondary infection. IgG subclass studies with the congenic mice indicated that serum IgG1, and IgG2a to a lesser extent, were the primary immunoglobulins produced and no major shift in subclass was noted with time during the primary infection.(ABSTRACT TRUNCATED AT 250 WORDS)

Apolipoprotein E-specific innate immune response in astrocytes from targeted replacement mice

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Montine Thomas J

2006-04-01

Full Text Available Abstract Background Inheritance of the three different alleles of the human apolipoprotein (apo E gene (APOE are associated with varying risk or clinical outcome from a variety of neurologic diseases. ApoE isoform-specific modulation of several pathogenic processes, in addition to amyloid β metabolism in Alzheimer's disease, have been proposed: one of these is innate immune response by glia. Previously we have shown that primary microglia cultures from targeted replacement (TR APOE mice have apoE isoform-dependent innate immune activation and paracrine damage to neurons that is greatest with TR by the ε4 allele (TR APOE4 and that derives from p38 mitogen-activated protein kinase (p38MAPK activity. Methods Primary cultures of TR APOE2, TR APOE3 and TR APOE4 astrocytes were stimulated with lipopolysaccharide (LPS. ApoE secretion, cytokine production, and nuclear factor-kappa B (NF-κB subunit activity were measured and compared. Results Here we showed that activation of primary astrocytes from TR APOE mice with LPS led to TR APOE-dependent differences in cytokine secretion that were greatest in TR APOE2 and that were associated with differences in NF-κB subunit activity. Conclusion Our results suggest that LPS activation of innate immune response in TR APOE glia results in opposing outcomes from microglia and astrocytes as a result of TR APOE-dependent activation of p38MAPK or NF-κB signaling in these two cell types.

Iron oxide nanoparticles modulate heat shock proteins and organ specific markers expression in mice male accessory organs

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Sundarraj, Kiruthika; Raghunath, Azhwar; Panneerselvam, Lakshmikanthan; Perumal, Ekambaram, E-mail: ekas2009@buc.edu.in

2017-02-15

With increased industrial utilization of iron oxide nanoparticles (Fe{sub 2}O{sub 3}-NPs), concerns on adverse reproductive health effects following exposure have been immensely raised. In the present study, the effects of Fe{sub 2}O{sub 3}-NPs exposure in the seminal vesicle and prostate gland were studied in mice. Mice were exposed to two different doses (25 and 50 mg/kg) of Fe{sub 2}O{sub 3}-NPs along with the control and analyzed the expressions of heat shock proteins (HSP60, HSP70 and HSP90) and organ specific markers (Caltrin, PSP94, and SSLP1). Fe{sub 2}O{sub 3}-NPs decreased food consumption, water intake, and organo-somatic index in mice with elevated iron levels in serum, urine, fecal matter, seminal vesicle and prostate gland. FTIR spectra revealed alterations in the functional groups of biomolecules on Fe{sub 2}O{sub 3}-NPs treatment. These changes are accompanied by increased lactate dehydrogenase levels with decreased total protein and fructose levels. The investigation of oxidative stress biomarkers demonstrated a significant increase in reactive oxygen species, nitric oxide, lipid peroxidation, protein carbonyl content and glutathione peroxidase with a concomitant decrement in the glutathione and ascorbic acid in the male accessory organs which confirmed the induction of oxidative stress. An increase in NADPH-oxidase-4 with a decrease in glutathione-S-transferase was observed in the seminal vesicle and prostate gland of the treated groups. An alteration in HSP60, HSP70, HSP90, Caltrin, PSP94, and SSLP1 expression was also observed. Moreover, accumulation of Fe{sub 2}O{sub 3}-NPs brought pathological changes in the seminal vesicle and prostate gland of treated mice. These findings provide evidence that Fe{sub 2}O{sub 3}-NPs could be an environmental risk factor for reproductive disease. - Highlights: • Fe{sub 2}O{sub 3}-NPs caused adverse effects on the seminal vesicle and prostate gland of mice • Heat shock proteins (Hsp60, 70 and 90) were

Propagation of senescent mice using nuclear transfer embryonic stem cell lines.

Science.gov (United States)

Mizutani, Eiji; Ono, Tetsuo; Li, Chong; Maki-Suetsugu, Rinako; Wakayama, Teruhiko

2008-09-01

Senescent mice are often infertile, and the cloning success rate decreases with age, making it almost impossible to produce cloned progeny directly from such animals. In this study, we tried to produce offspring from such "unclonable" senescent mice using nuclear transfer techniques. Donor fibroblasts were obtained from the tail tips of mice aged up to 2 years and 9 months. Although most attempts failed to produce cloned mice by direct somatic cell nuclear transfer, we managed to establish nuclear transfer embryonic stem (ntES) cell lines from all aged mice with an establishment rate of 10-25%, irrespective of sex or strain. Finally, cloned mice were obtained from these ntES cells by a second round of nuclear transfer. In addition, healthy offspring was obtained from all aged donors via germline transmission of ntES cells in chimeric mice. This technique is thus applicable to the propagation of a variety of animals, irrespective of age or fertile potential.

Superoxide produced by Kupffer cells is an essential effector in concanavalin A-induced hepatitis in mice.

Science.gov (United States)

Nakashima, Hiroyuki; Kinoshita, Manabu; Nakashima, Masahiro; Habu, Yoshiko; Shono, Satoshi; Uchida, Takefumi; Shinomiya, Nariyoshi; Seki, Shuhji

2008-12-01

Although concanavalin A (Con-A)-induced experimental hepatitis is thought to be induced by activated T cells, natural killer T (NKT) cells, and cytokines, precise mechanisms are still unknown. In the current study, we investigated the roles of Kupffer cells, NKT cells, FasL, tumor necrosis factor (TNF), and superoxide in Con-A hepatitis in C57BL/6 mice. Removal of Kupffer cells using gadolinium chloride (GdCl(3)) from the liver completely inhibited Con-A hepatitis, whereas increased serum TNF and IFN-gamma levels were not inhibited at all. Unexpectedly, anti-FasL antibody pretreatment did not inhibit Con-A hepatitis, whereas it inhibited hepatic injury induced by a synthetic ligand of NKT cells, alpha-galactosylceramide. Furthermore, GdCl(3) pretreatment changed neither the activation-induced down-regulation of NK1.1 antigens as well as T cell receptors of NKT cells nor the increased expression of the CD69 activation antigen of hepatic T cells. CD68(+) Kupffer cells greatly increased in proportion in the early phase after Con-A injection; this increase was abrogated by GdCl(3) pretreatment. Anti-TNF antibody (Ab) pretreatment did not inhibit the increase of Kupffer cells, but it effectively suppressed superoxide/reactive oxygen production from Kupffer cells and the resulting hepatic injury. Conversely, depletion of NKT cells in mice by NK1.1 Ab pretreatment did suppress both the increase of CD68(+) Kupffer cells and Con-A hepatitis. Consistently, the diminution of oxygen radicals produced by Kupffer cells by use of free radical scavengers greatly inhibited Con-A hepatitis without suppressing cytokine production. However, adoptive transfer experiments also indicate that a close interaction/cooperation of Kupffer cells with NKT cells is essential for Con-A hepatitis. Superoxide produced by Kupffer cells may be the essential effector in Con-A hepatitis, and TNF and NKT cells support their activation and superoxide production.

Generation of Oxtr cDNA(HA)-Ires-Cre Mice for Gene Expression in an Oxytocin Receptor Specific Manner.

Science.gov (United States)

Hidema, Shizu; Fukuda, Tomokazu; Hiraoka, Yuichi; Mizukami, Hiroaki; Hayashi, Ryotaro; Otsuka, Ayano; Suzuki, Shingo; Miyazaki, Shinji; Nishimori, Katsuhiko

2016-05-01

The neurohypophysial hormone oxytocin (OXT) and its receptor (OXTR) have critical roles in the regulation of pro-social behaviors, including social recognition, pair bonding, parental behavior, and stress-related responses. Supporting this hypothesis, a portion of patients suffering from autism spectrum disorder have mutations, such as single nucleotide polymorphisms, or epigenetic modifications in their OXTR gene. We previously reported that OXTR-deficient mice exhibit pervasive social deficits, indicating the critical role of OXTR in social behaviors. In the present study, we generated Oxtr cDNA(HA)-Ires-Cre knock-in mice, expressing both OXTR and Cre recombinase under the control of the endogenous Oxtr promoter. Knock-in cassette of Oxtr cDNA(HA)-Ires-Cre consisted of Oxtr cDNA tagged with the hemagglutinin epitope at the 3' end (Oxtr cDNA(HA)), internal ribosomal entry site (Ires), and Cre. Cre was expressed in the uterus, mammary gland, kidney, and brain of Oxtr cDNA(HA)-Ires-Cre knock-in mice. Furthermore, the distribution of Cre in the brain was similar to that observed in Oxtr-Venus fluorescent protein expressing mice (Oxtr-Venus), another animal model previously generated by our group. Social behavior of Oxtr cDNA(HA)-Ires-Cre knock-in mice was similar to that of wild-type animals. We demonstrated that this construct is expressed in OXTR-expressing neurons specifically after an infection with the recombinant adeno-associated virus carrying the flip-excision switch vector. Using this system, we showed the transport of the wheat-germ agglutinin tracing molecule from the OXTR-expressing neurons to the innervated neurons in knock-in mice. This study might contribute to the monosynaptic analysis of neuronal circuits and to the optogenetic analysis of neurons expressing OXTR. © 2015 Wiley Periodicals, Inc.

The Involvement of Mitochondrial Membrane Potential in Cross-Resistance Between Radiation and Docetaxel

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Kuwahara, Yoshikazu [Department of Radiation Biology and Medicine, Faculty of Medicine, Tohoku Medical and Pharmaceutical University, Sendai (Japan); Department of Pathology, Institute of Development, Aging and Cancer, Tohoku University, Sendai (Japan); Roudkenar, Mehryar Habibi; Suzuki, Masatoshi; Urushihara, Yusuke; Fukumoto, Motoi [Department of Pathology, Institute of Development, Aging and Cancer, Tohoku University, Sendai (Japan); Saito, Yohei [Department of Radiopharmacy, Tohoku Medical and Pharmaceutical University, Sendai (Japan); Fukumoto, Manabu, E-mail: manabu.fukumoto.a8@tohoku.ac.jp [Department of Pathology, Institute of Development, Aging and Cancer, Tohoku University, Sendai (Japan); Department of Molecular Pathology, Tokyo Medical University, Tokyo (Japan)

2016-11-01

Purpose: To understand the molecular mechanisms underlying cancer cell radioresistance, clinically relevant radioresistant (CRR) cells that continue to proliferate during exposure to 2 Gy/day X-rays for more than 30 days were established. A modified high-density survival assay for anticancer drug screening revealed that CRR cells were resistant to an antimicrotubule agent, docetaxel (DTX). The involvement of reactive oxygen species (ROS) from mitochondria (mtROS) in the cross-resistance to X-rays and DTX was studied. Methods and Materials: Sensitivity to anticancer agents was determined by a modified high-density cell survival or water-soluble tetrazolium salt assay. DTX-induced mtROS generation was determined by MitoSOX red staining. JC-1 staining was used to visualize mitochondrial membrane potential. DTX-induced DNA double-strand breaks were determined by γ-H2AX staining. To obtain mitochondrial DNA-lacking (ρ{sup 0}) cells, the cells were cultured for 3 to 4 weeks in medium containing ethidium bromide. Results: Treatment with DTX increased mtROS in parental cells but not in CRR cells. DTX induced DNA double-strand breaks in parental cells. The mitochondrial membrane potential of CRR cells was lower in CRR cells than in parental cells. Depletion of mtDNA induced DTX resistance in parental cells. Treatment with dimethyl sulfoxide also induced DTX resistance in parental cells. Conclusions: The mitochondrial dysfunction observed in CRR cells contributes to X-ray and DTX cross-resistance. The activation of oxidative phosphorylation in CRR cells may represent an effective approach to overcome radioresistant cancers. In general, the overexpression of β-tubulin or multidrug efflux pumps is thought to be involved in DTX resistance. In the present study, we discovered another DTX resistant mechanism by investigating CRR cells.

Ovalbumin-coated pH-sensitive microneedle arrays effectively induce ovalbumin-specific antibody and T-cell responses in mice.

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van der Maaden, Koen; Varypataki, Eleni Maria; Romeijn, Stefan; Ossendorp, Ferry; Jiskoot, Wim; Bouwstra, Joke

2014-10-01

The aim of this work was to study the applicability of antigen-coated pH-sensitive microneedle arrays for effective vaccination strategies. Therefore, a model antigen (ovalbumin) was coated onto pH-sensitive (pyridine-modified) microneedle arrays to test pH-triggered antigen release by applying the coated arrays onto ex vivo human skin, and by conducting a dermal immunization study in mice. The release of antigen into ex vivo human skin from the coated microneedles was determined by using radioactively labeled ovalbumin. To investigate the induction of antigen-specific IgG, and CD4(+) and CD8(+) T-cell responses, BALB/c mice were immunized with antigen-coated pH-sensitive microneedles by the 'coat and poke' approach. These responses were compared to responses induced by the 'poke and patch' approach, and subcutaneous and intradermal vaccination with classic hypodermic needles. The pH-sensitive microneedle arrays were efficiently coated with ovalbumin (95% coating efficiency) and upon application of six microneedle arrays 4.27 of 7 μg ovalbumin was delivered into the skin, showing a release efficiency of 70%. In contrast, the 'poke and patch' approach led to a delivery of only 6.91 of 100 μg ovalbumin (7% delivery efficiency). Immunization by means of ovalbumin-coated microneedles resulted in robust CD4(+) and CD8(+) T-cell responses comparable to those obtained after subcutaneous or intradermal immunization with conventional needles. Moreover, it effectively induced IgG responses; however, it required prime-boost immunizations before antibodies were produced. In conclusion, antigen delivery into ex vivo human skin by antigen-coated pH-sensitive microneedle arrays is more efficient than the 'poke-and-patch' approach and in vivo vaccination studies show the applicability of pH-sensitive microneedles for the induction of both T cell and B cell responses. Copyright © 2014 Elsevier B.V. All rights reserved.

MRP-1 expression levels determine strain-specific susceptibility to sodium arsenic-induced renal injury between C57BL/6 and BALB/c mice

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Kimura, Akihiko; Ishida, Yuko; Wada, Takashi; Yokoyama, Hitoshi; Mukaida, Naofumi; Kondo, Toshikazu

2005-01-01

To clarify the pathophysiological mechanism underlying acute renal injury caused by acute exposure to arsenic, we subcutaneously injected both BALB/c and C57BL/6 mice with sodium arsenite (NaAs; 13.5 mg/kg). BALB/c mice exhibited exaggerated elevation of serum blood urea nitrogen (BUN) and creatinine (CRE) levels, compared with C57BL/6 mice. Moreover, half of BALB/c mice died by 24 h, whereas all C57BL/6 mice survived. Histopathological examination on kidney revealed severe hemorrhages, acute tubular necrosis, neutrophil infiltration, cast formation, and disappearance of PAS-positive brush borders in BALB/c mice, later than 10 h. These pathological changes were remarkably attenuated in C57BL/6 mice, accompanied with lower intrarenal arsenic concentrations, compared with BALB/c mice. Among heavy metal inducible proteins including multidrug resistance-associated protein (MRP)-1, multidrug resistance gene (MDR)-1, metallothionein (MT)-1, and arsenite inducible, cysteine- and histidine-rich RNA-associated protein (AIRAP), intrarenal MDR-1, MT-1, and AIRAP gene expression was enhanced to a similar extent in both strains, whereas NaAs challenge augmented intrarenal MRP-1 mRNA and protein expression levels in C57BL/6 but not BALB/c mice. Moreover, the administration of a specific inhibitor of MRP-1, MK-571, significantly exaggerated acute renal injury in C57BL/6 mice. Thus, MRP-1 is crucially involved in arsenic efflux and eventually prevention of acute renal injury upon acute exposure to NaAs

Improved motor performance in Dyt1 ΔGAG heterozygous knock-in mice by cerebellar Purkinje-cell specific Dyt1 conditional knocking-out.

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Yokoi, Fumiaki; Dang, Mai Tu; Li, Yuqing

2012-05-01

Early-onset generalized torsion dystonia (dystonia 1) is an inherited movement disorder caused by mutations in DYT1 (TOR1A), which codes for torsinA. Most patients have a 3-base pair deletion (ΔGAG) in one allele of DYT1, corresponding to a loss of a glutamic acid residue (ΔE) in the C-terminal region of the protein. Functional alterations in basal ganglia circuits and the cerebellum have been reported in dystonia. Pharmacological manipulations or mutations in genes that result in functional alterations of the cerebellum have been reported to have dystonic symptoms and have been used as phenotypic rodent models. Additionally, structural lesions in the abnormal cerebellar circuits, such as cerebellectomy, have therapeutic effects in these models. A previous study has shown that the Dyt1 ΔGAG heterozygous knock-in (KI) mice exhibit motor deficits in the beam-walking test. Both Dyt1 ΔGAG heterozygous knock-in (KI) and Dyt1 Purkinje cell-specific knockout (Dyt1 pKO) mice exhibit dendritic alterations of cerebellar Purkinje cells. Here, Dyt1 pKO mice exhibited significantly less slip numbers in the beam-walking test, suggesting better motor performance than control littermates, and normal gait. Furthermore, Dyt1 ΔGAG KI/Dyt1 pKO double mutant mice exhibited significantly lower numbers of slips than Dyt1 ΔGAG heterozygous KI mice, suggesting Purkinje-cell specific knockout of Dyt1 wild-type (WT) allele in Dyt1 ΔGAG heterozygous KI mice rescued the motor deficits. The results suggest that molecular lesions of torsinA in Purkinje cells by gene therapy or intervening in the signaling pathway downstream of the cerebellar Purkinje cells may rescue motor symptoms in dystonia 1. Copyright © 2012 Elsevier B.V. All rights reserved.

Generating Chimeric Mice by Using Embryos from Nonsuperovulated BALB/c Mice Compared with Superovulated BALB/c and Albino C57BL/6 Mice.

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Esmail, Michael Y; Qi, Peimin; Connor, Aurora Burds; Fox, James G; García, Alexis

2016-01-01

The reliable generation of high-percentage chimeras from gene-targeted C57BL/6 embryonic stem cells has proven challenging, despite optimization of cell culture and microinjection techniques. To improve the efficiency of this procedure, we compared the generation of chimeras by using 3 different inbred, albino host, embryo-generating protocols: BALB/cAnNTac (BALB/c) donor mice superovulated at 4 wk of age, 12-wk-old BALB/c donor mice without superovulation, and C57BL/6NTac-Tyr(tm1Arte) (albino B6) mice superovulated at 4 wk of age. Key parameters measured included the average number of injectable embryos per donor, the percentage of live pups born from the total number of embryos transferred to recipients, and the number of chimeric pups with high embryonic-stem-cell contribution by coat color. Although albino B6 donors produced significantly more injectable embryos than did BALB/c donors, 12-wk-old BALB/c donor produced high-percentage (at least 70%) chimeras more than 2.5 times as often as did albino B6 mice and 20 times more efficiently than did 4-wk-old BALB/c donors. These findings clearly suggest that 12-wk-old BALB/c mice be used as blastocyst donors to reduce the number of mice used to generate each chimera, reduce the production of low-percentage chimeras, and maximize the generation of high-percentage chimeras from C57BL/6 embryonic stem cells.

Structure, Function, and Evolution of Rice Centromeres

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Jiang, Jiming

2010-02-04

The centromere is the most characteristic landmark of eukaryotic chromosomes. Centromeres function as the site for kinetochore assembly and spindle attachment, allowing for the faithful pairing and segregation of sister chromatids during cell division. Characterization of centromeric DNA is not only essential to understand the structure and organization of plant genomes, but it is also a critical step in the development of plant artificial chromosomes. The centromeres of most model eukaryotic species, consist predominantly of long arrays of satellite DNA. Determining the precise DNA boundary of a centromere has proven to be a difficult task in multicellular eukaryotes. We have successfully cloned and sequenced the centromere of rice chromosome 8 (Cen8), representing the first fully sequenced centromere from any multicellular eukaryotes. The functional core of Cen8 spans ~800 kb of DNA, which was determined by chromatin immunoprecipitation (ChIP) using an antibody against the rice centromere-specific H3 histone. We discovered 16 actively transcribed genes distributed throughout the Cen8 region. In addition to Cen8, we have characterized eight additional rice centromeres using the next generation sequencing technology. We discovered four subfamilies of the CRR retrotransposon that is highly enriched in rice centromeres. CRR elements are constitutively transcribed and different CRR subfamilies are differentially processed by RNAi. These results suggest that different CRR subfamilies may play different roles in the RNAi-mediated pathway for formation and maintenance of centromeric chromatin.

Vγ4+ T Cells: A Novel IL-17-Producing γδ T Subsets during the Early Phase of Chlamydial Airway Infection in Mice

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Li-da Sun

2018-01-01

Full Text Available Our previous studies showed that γδ T cells provided immune protection against Chlamydial muridarum (Cm, an obligate intracellular strain of chlamydia trachomatis, lung infection by producing abundant IL-17. In this study, we investigated the proliferation and activation of lung γδ T cell subsets, specifically the IL-17 and IFNγ production by them following Cm lung infection. Our results found that five γδ T cell subsets, Vγ1+ T, Vγ2+ T, Vγ4+ T, Vγ5+ T, and Vγ6+ T, expressed in lungs of naïve mice, while Cm lung infection mainly induced the proliferation and activation of Vγ4+ T cells at day 3 p.i., following Vγ1+ T cells at day 7 p.i. Cytokine detection showed that Cm lung infection induced IFNγ secretion firstly by Vγ4+ T cells at very early stage (day 3 and changed to Vγ1+ T cells at midstage (day 7. Furthermore, Vγ4+ T cell is the main γδ T cell subset that secretes IL-17 at the very early stage of Cm lung infection and Vγ1+ T cell did not secrete IL-17 during the infection. These findings provide in vivo evidence that Vγ4+T cells are the major IL-17 and IFNγ-producing γδ T cell subsets at the early period of Cm lung infection.

A Postnatal Diet Containing Phospholipids, Processed to Yield Large, Phospholipid-Coated Lipid Droplets, Affects Specific Cognitive Behaviors in Healthy Male Mice.

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Schipper, Lidewij; van Dijk, Gertjan; Broersen, Laus M; Loos, Maarten; Bartke, Nana; Scheurink, Anton Jw; van der Beek, Eline M

2016-06-01

Infant cognitive development can be positively influenced by breastfeeding rather than formula feeding. The composition of breast milk, especially lipid quality, and the duration of breastfeeding have been linked to this effect. We investigated whether the physical properties and composition of lipid droplets in milk may contribute to cognitive development. From postnatal day (P) 16 to P44, healthy male C57BL/6JOlaHsd mice were fed either a control or a concept rodent diet, in which the dietary lipid droplets were large and coated with milk phospholipids, resembling more closely the physical properties and composition of breast milk lipids. Thereafter, all mice were fed an AIN-93M semisynthetic rodent diet. The mice were subjected to various cognitive tests during adolescence (P35-P44) and adulthood (P70-P101). On P102, mice were killed and brain phospholipids were analyzed. The concept diet improved performance in short-term memory tasks that rely on novelty exploration during adolescence (T-maze; spontaneous alternation 87% in concept-fed mice compared with 74% in mice fed control diet; P diet. Brain phospholipid composition at P102 was not different between diet groups. Exposure to a diet with lipids mimicking more closely the structure and composition of lipids in breast milk improved specific cognitive behaviors in mice. These data suggest that lipid structure should be considered as a relevant target to improve dietary lipid quality in infant milk formulas. © 2016 American Society for Nutrition.

RETINOIC ACID INDUCTION OF CLEFT PALATE IN EGF AND TGF-ALPHA KNOCKOUT MICE: STAGE SPECIFIC INFLUENCES OF GROWTH FACTOR EXPRESSION

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ABBOTT, B. D., LEFFLER, K.E. AND BUCKALEW, A.R, Reproductive Toxicology Division, NHEERL, ORD, US EPA, Research Triangle Park, North Carolina. Retinoic acid induction of cleft palate (CP) in EGF and TGF knockout mice: Stage specific influences of growth factor expression.<...

Obesity development in neuron-specific lipoprotein lipase deficient mice is not responsive to increased dietary fat content or change in fat composition.

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Wang, Hong; Taussig, Matthew D; DiPatrizio, Nicholas V; Bruce, Kimberley; Piomelli, Daniele; Eckel, Robert H

2016-07-01

We have previously reported that mice with neuron-specific LPL deficiency (NEXLPL-/-) become obese by 16weeks of age on chow. Moreover, these mice had reduced uptake of triglyceride (TG)-rich lipoprotein-derived fatty acids and lower levels of n-3 long chain polyunsaturated fatty acids (n-3 PUFAs) in the hypothalamus. Here, we asked whether increased dietary fat content or altered dietary composition could modulate obesity development in NEXLPL-/- mice. Male NEXLPL-/- mice and littermate controls (WT) were randomly assigned one of three synthetic diets; a high carbohydrate diet (HC, 10% fat), a high-fat diet (HF, 45% fat), or a HC diet supplemented with n-3 PUFAs (HCn-3, 10% fat, Lovaza, GSK®). After 42weeks of HC feeding, body weight and fat mass were increased in the NEXLPL-/- mice compared to WT. WT mice fed a HF diet displayed typical diet-induced obesity, but weight gain was only marginal in HF-fed NEXLPL-/- mice, with no significant difference in body composition. Dietary n-3 PUFA supplementation did not prevent obesity in NEXLPL-/- mice, but was associated with differential modifications in hypothalamic gene expression and PUFA concentration compared to WT mice. Our findings suggest that neuronal LPL is involved in the regulation of body weight and composition in response to either the change in quantity (HF feeding) or quality (n-3 PUFA-enriched) of dietary fat. The precise role of LPL in lipid sensing in the brain requires further investigation. Copyright © 2016 Elsevier Inc. All rights reserved.

X-Ray-Induced Damage to the Submandibular Salivary Glands in Mice: An Analysis of Strain-Specific Responses

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Kamiya, Mana; Kawase, Tomoyuki; Hayama, Kazuhide; Tsuchimochi, Makoto; Okuda, Kazuhiro; Yoshie, Hiromasa

2015-01-01

Abstract Radiation therapy for head and neck cancers often causes xerostomia (dry mouth) by acutely damaging the salivary glands through the induction of severe acute inflammation. By contrast, the mechanism underlying the X-ray-induced delayed salivary dysfunction is unknown and has attracted increasing attention. To identify and develop a mouse model that distinguishes the delayed from the acute effects, we examined three different mouse strains (C57BL/6, ICR, and ICR-nu/nu) that showed distinct T-cell activities to comparatively analyze their responses to X-ray irradiation. Three strains were irradiated with X-rays (25 Gy), and functional changes of the submandibular glands were examined by determining pilocarpine-induced saliva secretion. Structural changes were evaluated using histopathological and immunohistochemical examinations of CD3, cleaved poly (ADP-ribose) polymerase (PARP), and Bcl-xL. In C57BL/6 mice, the X-ray irradiation induced acute inflammation accompanied by severe inflammatory cell infiltration at 4 days postirradiation, causing substantial destruction and significant dysfunction at 2 weeks. Fibrotic repair was observed at 16 weeks. In ICR-nu/nu mice, the inflammation and organ destruction were much milder than in the other mice strains, but increased apoptotic cells and a significant reduction in salivary secretion were observed at 4 and 8 weeks and beyond, respectively. These results suggest that in C57BL/6 mice, X-ray-induced functional and structural damage to the salivary glands is caused mainly by acute inflammation. By contrast, although neither acute inflammation nor organ destruction was observed in ICR-nu/nu mice, apoptotic cell death preceded the dysfunction in salivary secretion in the later phase. These data suggest that the X-ray-irradiated ICR-nu/nu mouse may be a useful animal model for developing more specific therapeutic methods for the delayed dysfunction of salivary glands. PMID:26309806

RhoE deficiency produces postnatal lethality, profound motor deficits and neurodevelopmental delay in mice.

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Enric Mocholí

Full Text Available Rnd proteins are a subfamily of Rho GTPases involved in the control of actin cytoskeleton dynamics and other cell functions such as motility, proliferation and survival. Unlike other members of the Rho family, Rnd proteins lack GTPase activity and therefore remain constitutively active. We have recently described that RhoE/Rnd3 is expressed in the Central Nervous System and that it has a role in promoting neurite formation. Despite their possible relevance during development, the role of Rnd proteins in vivo is not known. To get insight into the in vivo function of RhoE we have generated mice lacking RhoE expression by an exon trapping cassette. RhoE null mice (RhoE gt/gt are smaller at birth, display growth retardation and early postnatal death since only half of RhoE gt/gt mice survive beyond postnatal day (PD 15 and 100% are dead by PD 29. RhoE gt/gt mice show an abnormal body position with profound motor impairment and impaired performance in most neurobehavioral tests. Null mutant mice are hypoactive, show an immature locomotor pattern and display a significant delay in the appearance of the hindlimb mature responses. Moreover, they perform worse than the control littermates in the wire suspension, vertical climbing and clinging, righting reflex and negative geotaxis tests. Also, RhoE ablation results in a delay of neuromuscular maturation and in a reduction in the number of spinal motor neurons. Finally, RhoE gt/gt mice lack the common peroneal nerve and, consequently, show a complete atrophy of the target muscles. This is the first model to study the in vivo functions of a member of the Rnd subfamily of proteins, revealing the important role of Rnd3/RhoE in the normal development and suggesting the possible involvement of this protein in neurological disorders.

Collective Recycling Responsibility in Closed-Loop Fashion Supply Chains with a Third Party: Financial Sharing or Physical Sharing?

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Jiajia Nie

2013-01-01

Full Text Available We develop three closed-loop supply chain models where manufacturers can utilize financial or physical support to push a third party to collect the used fashion product for remanufacturing. We first examine two strategies for the collective recycling responsibility (CRR, namely, the financial sharing (FS and the physical sharing (PS, using the model with no CRR as a benchmark. After that, we conduct a detailed comparison among the three models in terms of the retail price, demand, return rate, and the profits received by the supply chain members. With this study, we find the following. (i The FS or PS support offered by the manufacturer to the third party will result in a lower retail price and a higher demand. (ii The optimal return rate with PS scheme is always higher than that without the CRR, and the one with FS scheme is at least as high as that without the CRR. (iii All the members in the closed-loop supply chain can always benefit from the CRR. In addition, (iv which scheme of the FS, PS, or no CRR is the best for the supply chain members will depend heavily on the transfer price of the used product.

Ontogeny of SERT Expression and Antidepressant-like Response to Escitalopram in Wild-Type and SERT Mutant Mice.

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Mitchell, Nathan C; Gould, Georgianna G; Koek, Wouter; Daws, Lynette C

2016-08-01

Depression is a disabling affective disorder for which the majority of patients are not effectively treated. This problem is exacerbated in children and adolescents for whom only two antidepressants are approved, both of which are selective serotonin reuptake inhibitor (SSRIs). Unfortunately SSRIs are often less effective in juveniles than in adults; however, the mechanism(s) underlying age-dependent responses to SSRIs is unknown. To this end, we compared the antidepressant-like response to the SSRI escitalopram using the tail suspension test and saturation binding of [(3)H]citalopram to the serotonin transporter (SERT), the primary target of SSRIs, in juvenile [postnatal day (P)21], adolescent (P28), and adult (P90) wild-type (SERT+/+) mice. In addition, to model individuals carrying low-expressing SERT variants, we studied mice with reduced SERT expression (SERT+/-) or lacking SERT (SERT-/-). Maximal antidepressant-like effects were less in P21 mice relative to P90 mice. This was especially apparent in SERT+/- mice. However, the potency for escitalopram to produce antidepressant-like effects in SERT+/+ and SERT+/- mice was greater in P21 and P28 mice than in adults. SERT expression increased with age in terminal regions and decreased with age in cell body regions. Binding affinity values did not change as a function of age or genotype. As expected, in SERT-/- mice escitalopram produced no behavioral effects, and there was no specific [(3)H]citalopram binding. These data reveal age- and genotype-dependent shifts in the dose-response for escitalopram to produce antidepressant-like effects, which vary with SERT expression, and may contribute to the limited therapeutic response to SSRIs in juveniles and adolescents. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

CD90-positive cells, an additional cell population, produce laminin α2 upon transplantation to dy3k/dy3k mice

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Fukada, So-ichiro; Yamamoto, Yukiko; Segawa, Masashi; Sakamoto, Kenta; Nakajima, Mari; Sato, Masaki; Morikawa, Daisuke; Uezumi, Akiyoshi; Miyagoe-Suzuki, Yuko; Takeda, Shin'ichi; Tsujikawa, Kazutake; Yamamoto, Hiroshi

2008-01-01

Laminin α2 is a component of skeletal and cardiac muscle basal lamina. A defect of the laminin α2 chain leads to severe congenital muscular dystrophy (MDC1A) in humans and dy/dy mice. Myogenic cells including myoblasts, myotubes, and myofibers in skeletal muscle are a possible source of the laminin α2 chain, and myogenic cells are thus proposed as a cell source for congenital muscular dystrophy therapy. However, we observed production of laminin α2 in non-myogenic cells of normal mice, and we could enrich these laminin α2-producing cells in CD90 + cell fractions. Intriguingly, the number of CD90 + cells increased dramatically during skeletal muscle regeneration in mice. This fraction did not include myogenic cells but exhibited a fibroblast-like phenotype. Moreover, these cells were resident in skeletal muscle, not derived from bone marrow. Finally, the production of laminin α2 in CD90 + cells was not dependent on fusion with myogenic cells. Thus, CD90 + cells are a newly identified additional cell fraction that increased during skeletal muscle regeneration in vivo and could be another cell source for therapy for lama2-deficient muscular dystrophy

GABAA Receptors Containing ρ1 Subunits Contribute to In Vivo Effects of Ethanol in Mice

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Blednov, Yuri A.; Benavidez, Jillian M.; Black, Mendy; Leiter, Courtney R.; Osterndorff-Kahanek, Elizabeth; Johnson, David; Borghese, Cecilia M.; Hanrahan, Jane R.; Johnston, Graham A. R.; Chebib, Mary; Harris, R. Adron

2014-01-01

GABAA receptors consisting of ρ1, ρ2, or ρ3 subunits in homo- or hetero-pentamers have been studied mainly in retina but are detected in many brain regions. Receptors formed from ρ1 are inhibited by low ethanol concentrations, and family-based association analyses have linked ρ subunit genes with alcohol dependence. We determined if genetic deletion of ρ1 in mice altered in vivo ethanol effects. Null mutant male mice showed reduced ethanol consumption and preference in a two-bottle choice test with no differences in preference for saccharin or quinine. Null mutant mice of both sexes demonstrated longer duration of ethanol-induced loss of righting reflex (LORR), and males were more sensitive to ethanol-induced motor sedation. In contrast, ρ1 null mice showed faster recovery from acute motor incoordination produced by ethanol. Null mutant females were less sensitive to ethanol-induced development of conditioned taste aversion. Measurement of mRNA levels in cerebellum showed that deletion of ρ1 did not change expression of ρ2, α2, or α6 GABAA receptor subunits. (S)-4-amino-cyclopent-1-enyl butylphosphinic acid (“ρ1” antagonist), when administered to wild type mice, mimicked the changes that ethanol induced in ρ1 null mice (LORR and rotarod tests), but the ρ1 antagonist did not produce these effects in ρ1 null mice. In contrast, (R)-4-amino-cyclopent-1-enyl butylphosphinic acid (“ρ2” antagonist) did not change ethanol actions in wild type but produced effects in mice lacking ρ1 that were opposite of the effects of deleting (or inhibiting) ρ1. These results suggest that ρ1 has a predominant role in two in vivo effects of ethanol, and a role for ρ2 may be revealed when ρ1 is deleted. We also found that ethanol produces similar inhibition of function of recombinant ρ1 and ρ2 receptors. These data indicate that ethanol action on GABAA receptors containing ρ1/ρ2 subunits may be important for specific effects of ethanol in vivo. PMID:24454882

GABAA receptors containing ρ1 subunits contribute to in vivo effects of ethanol in mice.

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Yuri A Blednov

Full Text Available GABAA receptors consisting of ρ1, ρ2, or ρ3 subunits in homo- or hetero-pentamers have been studied mainly in retina but are detected in many brain regions. Receptors formed from ρ1 are inhibited by low ethanol concentrations, and family-based association analyses have linked ρ subunit genes with alcohol dependence. We determined if genetic deletion of ρ1 in mice altered in vivo ethanol effects. Null mutant male mice showed reduced ethanol consumption and preference in a two-bottle choice test with no differences in preference for saccharin or quinine. Null mutant mice of both sexes demonstrated longer duration of ethanol-induced loss of righting reflex (LORR, and males were more sensitive to ethanol-induced motor sedation. In contrast, ρ1 null mice showed faster recovery from acute motor incoordination produced by ethanol. Null mutant females were less sensitive to ethanol-induced development of conditioned taste aversion. Measurement of mRNA levels in cerebellum showed that deletion of ρ1 did not change expression of ρ2, α2, or α6 GABAA receptor subunits. (S-4-amino-cyclopent-1-enyl butylphosphinic acid ("ρ1" antagonist, when administered to wild type mice, mimicked the changes that ethanol induced in ρ1 null mice (LORR and rotarod tests, but the ρ1 antagonist did not produce these effects in ρ1 null mice. In contrast, (R-4-amino-cyclopent-1-enyl butylphosphinic acid ("ρ2" antagonist did not change ethanol actions in wild type but produced effects in mice lacking ρ1 that were opposite of the effects of deleting (or inhibiting ρ1. These results suggest that ρ1 has a predominant role in two in vivo effects of ethanol, and a role for ρ2 may be revealed when ρ1 is deleted. We also found that ethanol produces similar inhibition of function of recombinant ρ1 and ρ2 receptors. These data indicate that ethanol action on GABAA receptors containing ρ1/ρ2 subunits may be important for specific effects of ethanol in vivo.

A developmental stage-specific switch from DAZL to BOLL occurs during fetal oogenesis in humans, but not mice.

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Jing He

Full Text Available The Deleted in Azoospermia gene family encodes three germ cell-specific RNA-binding proteins (DAZ, DAZL and BOLL that are essential for gametogenesis in diverse species. Targeted disruption of Boll in mice causes male-specific spermiogenic defects, but females are apparently fertile. Overexpression of human BOLL promotes the derivation of germ cell-like cells from genetically female (XX, but not male (XY human ES cells however, suggesting a functional role for BOLL in regulating female gametogenesis in humans. Whether BOLL is expressed during oogenesis in mammals also remains unclear. We have therefore investigated the expression of BOLL during fetal oogenesis in humans and mice. We demonstrate that BOLL protein is expressed in the germ cells of the human fetal ovary, at a later developmental stage than, and almost mutually-exclusive to, the expression of DAZL. Strikingly, BOLL is downregulated, and DAZL re-expressed, as primordial follicles form, revealing BOLL expression to be restricted to a narrow window during fetal oogenesis. By quantifying the extent of co-expression of DAZL and BOLL with markers of meiosis, we show that this window likely corresponds to the later stages of meiotic prophase I. Finally, we demonstrate that Boll is also transiently expressed during oogenesis in the fetal mouse ovary, but is simultaneously co-expressed within the same germ cells as Dazl. These data reveal significant similarities and differences between the expression of BOLL homologues during oogenesis in humans and mice, and raise questions as to the validity of the Boll(-/- mouse as a model for understanding BOLL function during human oogenesis.

The MICE Online Systems

CERN Multimedia

CERN. Geneva

2012-01-01

The Muon Ionization Cooling Experiment (MICE) is designed to test transverse cooling of a muon beam, demonstrating an important step along the path toward creating future high intensity muon beam facilities. Protons in the ISIS synchrotron impact a titanium target, producing pions which decay into muons that propagate through the beam line to the MICE cooling channel. Along the beam line, particle identification (PID) detectors, scintillating fiber tracking detectors, and beam diagnostic tools identify and measure individual muons moving through the cooling channel. The MICE Online Systems encompass all tools; including hardware, software, and documentation, within the MLCR (MICE Local Control Room) that allow the experiment to efficiently record high quality data. Controls and Monitoring (C&M), Data Acquisition (DAQ), Online Monitoring and Reconstruction, Data Transfer, and Networking all fall under the Online Systems umbrella. C&M controls all MICE systems including the target, conventional an...

The contribution of Chlamydia-specific CD8⁺ T cells to upper genital tract pathology.

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Vlcek, Kelly R; Li, Weidang; Manam, Srikanth; Zanotti, Brian; Nicholson, Bruce J; Ramsey, Kyle H; Murthy, Ashlesh K

2016-02-01

Genital chlamydial infections lead to severe upper reproductive tract pathology in a subset of untreated women. We demonstrated previously that tumor necrosis factor (TNF)-α-producing CD8(+) T cells contribute significantly to chlamydial upper genital tract pathology in female mice. In addition, we observed that minimal chlamydial oviduct pathology develops in OT-1 transgenic (OT-1) mice, wherein the CD8(+) T-cell repertoire is restricted to recognition of the ovalbumin peptide Ova(257-264), suggesting that non-Chlamydia-specific CD8(+) T cells may not be responsible for chlamydial pathogenesis. In the current study, we evaluated whether antigen-specific CD8(+) T cells mediate chlamydial pathology. Groups of wild-type (WT) C57BL/6J, OT-1 mice, and OT-1 mice replete with WT CD8(+) T cells (1 × 10(6) cells per mouse intravenously) were infected intravaginally with C. muridarum (5 × 10(4) IFU/mouse). Serum total anti-Chlamydia antibody and total splenic anti-Chlamydia interferon (IFN)-γ and TNF-α responses were comparable among the three groups of animals. However, Chlamydia-specific IFN-γ and TNF-α production from purified splenic CD8(+) T cells of OT-1 mice was minimal, whereas responses in OT-1 mice replete with WT CD8(+) T cells were comparable to those in WT animals. Vaginal chlamydial clearance was comparable between the three groups of mice. Importantly, the incidence and severity of oviduct and uterine horn pathology was significantly reduced in OT-1 mice but reverted to WT levels in OT-1 mice replete with WT CD8(+) T cells. Collectively, these results demonstrate that Chlamydia-specific CD8(+) T cells contribute significantly to upper genital tract pathology.

Altered tumor growth in vivo after immunization of mice with antitumor antibodies

International Nuclear Information System (INIS)

Gorczynski, R.M.; Kennedy, M.; Polidoulis, I.; Price, G.B.

1984-01-01

A comparison has been made between the growth patterns of two spontaneously appearing mammary adenocarcinomas in murine bone marrow radiation chimeras and in mice preimmunized with monoclonal antibodies (MAb) detecting embryo-associated antigenic determinants. A correlation was seen between the ability of the embryo-immunized chimeras to produce cytotoxic antibody to the tumors, as assessed by an antibody-dependent cellular cytotoxic assay, and the permissiveness of the mice for growth of a tumor transplant. In addition, mice deliberately preimmunized with cytotoxic MAb (antibody-dependent cellular cytotoxic assay) allowed more rapid growth specifically of that tumor earlier found to be most sensitive to the MAb used for immunization. By comparing the changing antigenic phenotype of tumor cells serially passaged through different immunized, nonimmunized mice, evidence was found suggesting that immunization could cause either antigen modulation of transferred tumor cells or a (transient) selective advantage to antigenically discrete subpopulations within the heterogeneous tumor population. Finally, a study has been made of the growth pattern of tumor cells transplanted into mice immunized with rabbit antibodies directed against the murine MAb. In this case, tumor growth was slowed preferentially for the tumor reactive with the specific MAb, and again, predictable changes in the antigenic spectrum of tumor cells harvested from these animals were observed. Our overall findings are interpreted in terms of the involvement of networks of antibodies reacting with embryo-associated antigens in the regulation of growth of the murine mammary adenocarcinomas studied

Increased intracellular Th1 cytokines in scid mice with inflammatory bowel disease

DEFF Research Database (Denmark)

Bregenholt, S; Claesson, Mogens Helweg

1998-01-01

Severe combined immunodeficient (scid) mice engrafted with small pieces of full thickness gut wall from immunocompetent syngenic donors develop a chronic and lethal colitis. Lymphocytes from the lamina propria of engrafted mice were analyzed for phorbol ester/ionomycin-induced cytokine production...... by intracellular staining. A 4-5-fold increase in the fraction of IFN-gamma-producing CD4+ lamina propria T cells was found in moderately and severely diseased mice when compared to healthy congenic C.B-17 control mice. The number of IL-2-producing T cells was increased by approximately 2-fold when comparing mice...... suffering from severe disease to healthy control mice. The fraction of TNF-alpha positive CD4+ T cells was increased by a factor of two in both moderately and severely diseased mice. When analyzing Th2 cytokines, it was found that the levels of IL-4-producing CD4+ T cells was not altered in diseased animals...

Lymphatic endothelial cells efferent to inflamed joints produce iNOS and inhibit lymphatic vessel contraction and drainage in TNF-induced arthritis in mice.

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Liang, Qianqian; Ju, Yawen; Chen, Yan; Wang, Wensheng; Li, Jinlong; Zhang, Li; Xu, Hao; Wood, Ronald W; Schwarz, Edward M; Boyce, Brendan F; Wang, Yongjun; Xing, Lianping

2016-03-12

In this study, we sought to determine the cellular source of inducible nitric oxide synthase (iNOS) induced in lymphatic endothelial cells (LECs) in response to tumor necrosis factor (TNF), the effects of iNOS on lymphatic smooth muscle cell (LSMC) function and on the development of arthritis in TNF-transgenic (TNF-Tg) mice, and whether iNOS inhibitors improve lymphatic function and reduce joint destruction in inflammatory erosive arthritis. We used quantitative polymerase chain reactions, immunohistochemistry, histology, and near-infrared imaging to examine (1) iNOS expression in podoplanin + LECs and lymphatic vessels from wild-type (WT) and TNF-Tg mice, (2) iNOS induction by TNF in WT LECs, (3) the effects of iNOS inhibitors on expression of functional muscle genes in LSMCs, and (4) the effects of iNOS inhibitors on lymphatic vessel contraction and drainage, as well as the severity of arthritis, in TNF-Tg mice. LECs from TNF-Tg mice had eight fold higher iNOS messenger RNA levels than WT cells, and iNOS expression was confirmed immunohistochemically in podoplanin + LECs in lymphatic vessels from inflamed joints. TNF (0.1 ng/ml) increased iNOS levels 40-fold in LECs. LSMCs cocultured with LECs pretreated with TNF had reduced expression of functional muscle genes. This reduction was prevented by ferulic acid, which blocked nitric oxide production. Local injection of L-N(6)-(1-iminoethyl)lysine 5-tetrazole-amide into inflamed paws of TNF-Tg mice resulted in recovery of lymphatic vessel contractions and drainage. Treatment of TNF-Tg mice with ferulic acid reduced synovial inflammation as well as cartilage and bone erosion, and it also restored lymphatic contraction and drainage. iNOS is produced primarily by LECs in lymphatic vessel efferent from inflamed joints of TNF-Tg mice in response to TNF and inhibits LSMC contraction and lymph drainage. Ferulic acid represents a potential new therapy to restore lymphatic function and thus improve inflammatory

The maximum specific hydrogen-producing activity of anaerobic mixed cultures: definition and determination

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Mu, Yang; Yang, Hou-Yun; Wang, Ya-Zhou; He, Chuan-Shu; Zhao, Quan-Bao; Wang, Yi; Yu, Han-Qing

2014-06-01

Fermentative hydrogen production from wastes has many advantages compared to various chemical methods. Methodology for characterizing the hydrogen-producing activity of anaerobic mixed cultures is essential for monitoring reactor operation in fermentative hydrogen production, however there is lack of such kind of standardized methodologies. In the present study, a new index, i.e., the maximum specific hydrogen-producing activity (SHAm) of anaerobic mixed cultures, was proposed, and consequently a reliable and simple method, named SHAm test, was developed to determine it. Furthermore, the influences of various parameters on the SHAm value determination of anaerobic mixed cultures were evaluated. Additionally, this SHAm assay was tested for different types of substrates and bacterial inocula. Our results demonstrate that this novel SHAm assay was a rapid, accurate and simple methodology for determining the hydrogen-producing activity of anaerobic mixed cultures. Thus, application of this approach is beneficial to establishing a stable anaerobic hydrogen-producing system.

The maximum specific hydrogen-producing activity of anaerobic mixed cultures: definition and determination.

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Mu, Yang; Yang, Hou-Yun; Wang, Ya-Zhou; He, Chuan-Shu; Zhao, Quan-Bao; Wang, Yi; Yu, Han-Qing

2014-06-10

Fermentative hydrogen production from wastes has many advantages compared to various chemical methods. Methodology for characterizing the hydrogen-producing activity of anaerobic mixed cultures is essential for monitoring reactor operation in fermentative hydrogen production, however there is lack of such kind of standardized methodologies. In the present study, a new index, i.e., the maximum specific hydrogen-producing activity (SHAm) of anaerobic mixed cultures, was proposed, and consequently a reliable and simple method, named SHAm test, was developed to determine it. Furthermore, the influences of various parameters on the SHAm value determination of anaerobic mixed cultures were evaluated. Additionally, this SHAm assay was tested for different types of substrates and bacterial inocula. Our results demonstrate that this novel SHAm assay was a rapid, accurate and simple methodology for determining the hydrogen-producing activity of anaerobic mixed cultures. Thus, application of this approach is beneficial to establishing a stable anaerobic hydrogen-producing system.

Production of knock-in mice in a single generation from embryonic stem cells.

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Ukai, Hideki; Kiyonari, Hiroshi; Ueda, Hiroki R

2017-12-01

The system-level identification and analysis of molecular networks in mammals can be accelerated by 'next-generation' genetics, defined as genetics that does not require crossing of multiple generations of animals in order to achieve the desired genetic makeup. We have established a highly efficient procedure for producing knock-in (KI) mice within a single generation, by optimizing the genome-editing protocol for KI embryonic stem (ES) cells and the protocol for the generation of fully ES-cell-derived mice (ES mice). Using this protocol, the production of chimeric mice is eliminated, and, therefore, there is no requirement for the crossing of chimeric mice to produce mice that carry the KI gene in all cells of the body. Our procedure thus shortens the time required to produce KI ES mice from about a year to ∼3 months. Various kinds of KI ES mice can be produced with a minimized amount of work, facilitating the elucidation of organism-level phenomena using a systems biology approach. In this report, we describe the basic technologies and protocols for this procedure, and discuss the current challenges for next-generation mammalian genetics in organism-level systems biology studies.

Fructans from aged garlic extract produce a delayed immunoadjuvant response to ovalbumin antigen in BALB/c mice.

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Chandrashekar, Puthanapura M; Venkatesh, Yeldur P

2012-02-01

Garlic (Allium sativum) is known for its innumerable biological activities including immunomodulation. Aged garlic extract (AGE), an odorless garlic preparation, has been shown to have superior immunomodulatory properties over raw garlic extract. Although garlic is a very rich source of fructans (17%, fresh weight basis), AGE contains only 0.22% of raw garlic fructans. Aged garlic fructans (AGF) have recently been shown to possess immunomodulatory activities in vitro. Natural adjuvants capable of eliciting better immune response of a model antigen are important in developing newer vaccines. In the present study, the adjuvant activity of AGF has been investigated in BALB/c mice using ovalbumin (OVA, 30 µg) as an experimental antigen. The body weights of animals did not change significantly indicating that the administration of garlic fructans is well-tolerated. AGF produce a significant humoral (serum IgG) response to OVA in BALB/c mice administered mucosally by either intranasal or oral route--a delayed response appearing on 50th day at a dose of 30 µg AGF by intranasal route. However, the serum IgG response was seen earlier on 35th day at a dose of 100 µg AGF by oral route. Higher concentrations of AGF (>50 µg) were inhibitory for adjuvant activity by intranasal administration. These observations indicate that AGF display immunoadjuvant activity for a test antigen though the humoral immune response is delayed.

The acute glucose lowering effect of specific GPR120 activation in mice is mainly driven by glucagon-like peptide 1.

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Linda Sundström

Full Text Available The mechanism behind the glucose lowering effect occurring after specific activation of GPR120 is not completely understood. In this study, a potent and selective GPR120 agonist was developed and its pharmacological properties were compared with the previously described GPR120 agonist Metabolex-36. Effects of both compounds on signaling pathways and GLP-1 secretion were investigated in vitro. The acute glucose lowering effect was studied in lean wild-type and GPR120 null mice following oral or intravenous glucose tolerance tests. In vitro, in GPR120 overexpressing cells, both agonists signaled through Gαq, Gαs and the β-arrestin pathway. However, in mouse islets the signaling pathway was different since the agonists reduced cAMP production. The GPR120 agonists stimulated GLP-1 secretion both in vitro in STC-1 cells and in vivo following oral administration. In vivo GPR120 activation induced significant glucose lowering and increased insulin secretion after intravenous glucose administration in lean mice, while the agonists had no effect in GPR120 null mice. Exendin 9-39, a GLP-1 receptor antagonist, abolished the GPR120 induced effects on glucose and insulin following an intravenous glucose challenge. In conclusion, GLP-1 secretion is an important mechanism behind the acute glucose lowering effect following specific GPR120 activation.

Transgenic mice expressing human glucocerebrosidase variants: utility for the study of Gaucher disease.

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Sanders, Angela; Hemmelgarn, Harmony; Melrose, Heather L; Hein, Leanne; Fuller, Maria; Clarke, Lorne A

2013-08-01

Gaucher disease is an autosomal recessively inherited storage disorder caused by deficiency of the lysosomal hydrolase, acid β-glucosidase. The disease manifestations seen in Gaucher patients are highly heterogeneous as is the responsiveness to therapy. The elucidation of the precise factors responsible for this heterogeneity has been challenging as the development of clinically relevant animal models of Gaucher disease has been problematic. Although numerous murine models for Gaucher disease have been described each has limitations in their specific utility. We describe here, transgenic murine models of Gaucher disease that will be particularly useful for the study of pharmacological chaperones. We have produced stable transgenic mouse strains that individually express wild type, N370S and L444P containing human acid β-glucosidase and show that each of these transgenic lines rescues the lethal phenotype characteristic of acid β-glucosidase null mice. Both the N370S and L444P transgenic models show early and progressive elevations of tissue sphingolipids with L444P mice developing progressive splenic Gaucher cell infiltration. We demonstrate the potential utility of these new transgenic models for the study of Gaucher disease pathogenesis. In addition, since these mice produce only human enzyme, they are particularly relevant for the study of pharmacological chaperones that are specifically targeted to human acid β-glucosidase and the common mutations underlying Gaucher disease. Copyright © 2013 Elsevier Inc. All rights reserved.

A neuron-specific deletion of the microRNA-processing enzyme DICER induces severe but transient obesity in mice.

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Géraldine M Mang

Full Text Available MicroRNAs (miRNAs are small, non-coding RNA molecules that regulate gene expression post-transcriptionally. MiRNAs are implicated in various biological processes associated with obesity, including adipocyte differentiation and lipid metabolism. We used a neuronal-specific inhibition of miRNA maturation in adult mice to study the consequences of miRNA loss on obesity development. Camk2a-CreERT2 (Cre+ and floxed Dicer (Dicerlox/lox mice were crossed to generate tamoxifen-inducible conditional Dicer knockouts (cKO. Vehicle- and/or tamoxifen-injected Cre+;Dicerlox/lox and Cre+;Dicer+/+ served as controls. Four cohorts were used to a measure body composition, b follow food intake and body weight dynamics, c evaluate basal metabolism and effects of food deprivation, and d assess the brain transcriptome consequences of miRNA loss. cKO mice developed severe obesity and gained 18 g extra weight over the 5 weeks following tamoxifen injection, mainly due to increased fat mass. This phenotype was highly reproducible and observed in all 38 cKO mice recorded and in none of the controls, excluding possible effects of tamoxifen or the non-induced transgene. Development of obesity was concomitant with hyperphagia, increased food efficiency, and decreased activity. Surprisingly, after reaching maximum body weight, obese cKO mice spontaneously started losing weight as rapidly as it was gained. Weight loss was accompanied by lowered O2-consumption and respiratory-exchange ratio. Brain transcriptome analyses in obese mice identified several obesity-related pathways (e.g. leptin, somatostatin, and nemo-like kinase signaling, as well as genes involved in feeding and appetite (e.g. Pmch, Neurotensin and in metabolism (e.g. Bmp4, Bmp7, Ptger1, Cox7a1. A gene cluster with anti-correlated expression in the cerebral cortex of post-obese compared to obese mice was enriched for synaptic plasticity pathways. While other studies have identified a role for miRNAs in obesity, we

Branched-chain amino acid (BCAA) supplementation enhances adaptability to exercise training of mice with a muscle-specific defect in the control of BCAA catabolism.

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Xu, Minjun; Kitaura, Yasuyuki; Shindo, Daichi; Shimomura, Yoshiharu

2018-03-01

Branched-chain α-keto acid dehydrogenase (BCKDH) kinase (BDK) suppresses the branched-chain amino acid (BCAA) catabolism by inactivation of the BCKDH complex. The muscle-specific BDK-deficient (BDK-mKO) mice showed accelerated BCAA oxidation in muscle and decreased endurance capacity after training (Xu et al. PLoS One. 12 (2017) e0180989). We here report that BCAA supplementation overcompensated endurance capacity in BDK-mKO mice after training.

A liver-specific defect of Acyl-CoA degradation produces hyperammonemia, hypoglycemia and a distinct hepatic Acyl-CoA pattern.

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Nicolas Gauthier

Full Text Available Most conditions detected by expanded newborn screening result from deficiency of one of the enzymes that degrade acyl-coenzyme A (CoA esters in mitochondria. The role of acyl-CoAs in the pathophysiology of these disorders is poorly understood, in part because CoA esters are intracellular and samples are not generally available from human patients. We created a mouse model of one such condition, deficiency of 3-hydroxy-3-methylglutaryl-CoA lyase (HL, in liver (HLLKO mice. HL catalyses a reaction of ketone body synthesis and of leucine degradation. Chronic HL deficiency and acute crises each produced distinct abnormal liver acyl-CoA patterns, which would not be predictable from levels of urine organic acids and plasma acylcarnitines. In HLLKO hepatocytes, ketogenesis was undetectable. Carboxylation of [2-(14C] pyruvate diminished following incubation of HLLKO hepatocytes with the leucine metabolite 2-ketoisocaproate (KIC. HLLKO mice also had suppression of the normal hyperglycemic response to a systemic pyruvate load, a measure of gluconeogenesis. Hyperammonemia and hypoglycemia, cardinal features of many inborn errors of acyl-CoA metabolism, occurred spontaneously in some HLLKO mice and were inducible by administering KIC. KIC loading also increased levels of several leucine-related acyl-CoAs and reduced acetyl-CoA levels. Ultrastructurally, hepatocyte mitochondria of KIC-treated HLLKO mice show marked swelling. KIC-induced hyperammonemia improved following administration of carglumate (N-carbamyl-L-glutamic acid, which substitutes for the product of an acetyl-CoA-dependent reaction essential for urea cycle function, demonstrating an acyl-CoA-related mechanism for this complication.

Detection of herpes simplex virus-specific DNA sequences in latently infected mice and in humans.

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Efstathiou, S; Minson, A C; Field, H J; Anderson, J R; Wildy, P

1986-02-01

Herpes simplex virus-specific DNA sequences have been detected by Southern hybridization analysis in both central and peripheral nervous system tissues of latently infected mice. We have detected virus-specific sequences corresponding to the junction fragment but not the genomic termini, an observation first made by Rock and Fraser (Nature [London] 302:523-525, 1983). This "endless" herpes simplex virus DNA is both qualitatively and quantitatively stable in mouse neural tissue analyzed over a 4-month period. In addition, examination of DNA extracted from human trigeminal ganglia has shown herpes simplex virus DNA to be present in an "endless" form similar to that found in the mouse model system. Further restriction enzyme analysis of latently infected mouse brainstem and human trigeminal DNA has shown that this "endless" herpes simplex virus DNA is present in all four isomeric configurations.

Regulation of allergic airway inflammation by adoptive transfer of CD4+ T cells preferentially producing IL-10.

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Matsuda, Masaya; Doi, Kana; Tsutsumi, Tatsuya; Fujii, Shinya; Kishima, Maki; Nishimura, Kazuma; Kuroda, Ikue; Tanahashi, Yu; Yuasa, Rino; Kinjo, Toshihiko; Kuramoto, Nobuyuki; Mizutani, Nobuaki; Nabe, Takeshi

2017-10-05

Anti-inflammatory pharmacotherapy for asthma has mainly depended on the inhalation of glucocorticoids, which non-specifically suppress immune responses. If the anti-inflammatory cytokine interleukin (IL)-10 can be induced by a specific antigen, asthmatic airway inflammation could be suppressed when individuals are exposed to the antigen. The purpose of this study was to develop cellular immunotherapeutics for atopic diseases using IL-10-producing CD4 + T cells. Spleen cells isolated from ovalbumin (OVA)-sensitized mice were cultured with the antigen, OVA and growth factors, IL-21, IL-27 and TGF-β for 7 days. After the 7-day culture, the CD4 + T cells were purified using a murine CD4 magnetic beads system. When the induced CD4 + T cells were stimulated by OVA in the presence of antigen-presenting cells, IL-10 was preferentially produced in vitro. When CD4 + T cells were adoptively transferred to OVA-sensitized mice followed by intratracheal OVA challenges, IL-10 was preferentially produced in the serum and bronchoalveolar lavage fluid in vivo. IL-10 production coincided with the inhibition of eosinophilic airway inflammation and epithelial mucus plugging. Most of the IL-10-producing CD4 + T cells were negative for Foxp3 and GATA-3, transcription factors of naturally occurring regulatory T cells and Th2 cells, respectively, but double positive for LAG-3 and CD49b, surface markers of inducible regulatory T cells, Tr1 cells. Collectively, most of the induced IL-10-producing CD4 + T cells could be Tr1 cells, which respond to the antigen to produce IL-10, and effectively suppressed allergic airway inflammation. The induced Tr1 cells may be useful for antigen-specific cellular immunotherapy for atopic diseases. Copyright © 2017 Elsevier B.V. All rights reserved.

Deletion of a coordinate regulator of type 2 cytokine expression in mice

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Mohrs, Markus; Blankespoor, Catherine M.; Wang, Zhi-En; Loots, Gaby G.; Hadeiba, Husein; Shinkai, Kanade; Rubin, Edward M.; Locksley, Richard M.

2001-07-30

Mechanisms underlying the differentiation of stable T helper subsets will be important in understanding how discrete types of immunity develop in response to different pathogens. An evolutionarily conserved {approx}400 base pair non-coding sequence in the IL-4/IL-13 intergenic region, designated CNS-1, was deleted in mice. The capacity to develop Th2 cells was compromised in vitro and in vivo in the absence of CNS-1. Despite the profound effect in T cells, mast cells from CNS-1-deleted mice maintained their capacity to produce IL-4. A T cell-specific element critical for optimal expression of type 2 cytokines may represent evolution of a regulatory sequence exploited by adaptive immunity.

Liver-specific rescuing of CEACAM1 reverses endothelial and cardiovascular abnormalities in male mice with null deletion of Ceacam1 gene

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Lucia Russo

2018-03-01

Full Text Available Objective: Mice with global null mutation of Ceacam1 (Cc1−/−, display impairment of insulin clearance that causes hyperinsulinemia followed by insulin resistance, elevated hepatic de novo lipogenesis, and visceral obesity. In addition, they manifest abnormal vascular permeability and elevated blood pressure. Liver-specific rescuing of Ceacam1 reversed all of the metabolic abnormalities in Cc1−/−liver+ mice. The current study examined whether Cc1−/− male mice develop endothelial and cardiac dysfunction and whether this relates to the metabolic abnormalities caused by defective insulin extraction. Methods and results: Myography studies showed reduction of agonist-stimulated nitric oxide production in resistance arterioles in Cc1−/−, but not Cc1−/−liver+ mice. Liver-based rescuing of CEACAM1 also attenuated the abnormal endothelial adhesiveness to circulating leukocytes in parallel to reducing plasma endothelin-1 and recovering plasma nitric oxide levels. Echocardiography studies revealed increased septal wall thickness, cardiac hypertrophy and reduced cardiac performance in Cc1−/−, but not Cc1−/−xliver+ mice. Insulin signaling experiments indicated compromised IRS1/Akt/eNOS pathway leading to lower nitric oxide level, and activated Shc/MAPK pathway leading to more endothelin-1 production in the aortae and hearts of Cc1−/−, but not Cc1−/−xliver+ mice. The increase in the ratio of endothelin-1 receptor A/B indicated an imbalance in the vasomotor activity of Cc1−/− mice, which was normalized in Cc1−/−xliver+ mice. Conclusions: The data underscore a critical role for impaired CEACAM1-dependent hepatic insulin clearance pathways and resulting hyperinsulinemia and lipid accumulation in aortae and heart in regulating the cardiovascular function. Keywords: Insulin clearance, Hyperinsulinemia, Insulin resistance, Endothelial function, Cardiomyopathy

Eplerenone ameliorates the phenotypes of metabolic syndrome with NASH in liver-specific SREBP-1c Tg mice fed high-fat and high-fructose diet.

Science.gov (United States)

Wada, Tsutomu; Miyashita, Yusuke; Sasaki, Motohiro; Aruga, Yusuke; Nakamura, Yuto; Ishii, Yoko; Sasahara, Masakiyo; Kanasaki, Keizo; Kitada, Munehiro; Koya, Daisuke; Shimano, Hitoshi; Tsuneki, Hiroshi; Sasaoka, Toshiyasu

2013-12-01

Because the renin-angiotensin-aldosterone system has been implicated in the development of insulin resistance and promotion of fibrosis in some tissues, such as the vasculature, we examined the effect of eplerenone, a selective mineralocorticoid receptor (MR) antagonist, on nonalcoholic steatohepatitis (NASH) and metabolic phenotypes in a mouse model reflecting metabolic syndrome in humans. We adopted liver-specific transgenic (Tg) mice overexpressing the active form of sterol response element binding protein-1c (SREBP-1c) fed a high-fat and fructose diet (HFFD) as the animal model in the present study. When wild-type (WT) C57BL/6 and liver-specific SREBP-1c Tg mice grew while being fed HFFD for 12 wk, body weight and epididymal fat weight increased in both groups with an elevation in blood pressure and dyslipidemia. Glucose intolerance and insulin resistance were also observed. Adipose tissue hypertrophy and macrophage infiltration with crown-like structure formation were also noted in mice fed HFFD. Interestingly, the changes noted in both genotypes fed HFFD were significantly ameliorated with eplerenone. HFFD-fed Tg mice exhibited the histological features of NASH in the liver, including macrovesicular steatosis and fibrosis, whereas HFFD-fed WT mice had hepatic steatosis without apparent fibrotic changes. Eplerenone effectively ameliorated these histological abnormalities. Moreover, the direct suppressive effects of eplerenone on lipopolysaccharide-induced TNFα production in the presence and absence of aldosterone were observed in primary-cultured Kupffer cells and bone marrow-derived macrophages. These results indicated that eplerenone prevented the development of NASH and metabolic abnormalities in mice by inhibiting inflammatory responses in both Kupffer cells and macrophages.

Disruption of Transient Serotonin Accumulation by Non-Serotonin-Producing Neurons Impairs Cortical Map Development

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Xiaoning Chen

2015-01-01

Full Text Available Polymorphisms that alter serotonin transporter SERT expression and functionality increase the risks for autism and psychiatric traits. Here, we investigate how SERT controls serotonin signaling in developing CNS in mice. SERT is transiently expressed in specific sets of glutamatergic neurons and uptakes extrasynaptic serotonin during perinatal CNS development. We show that SERT expression in glutamatergic thalamocortical axons (TCAs dictates sensory map architecture. Knockout of SERT in TCAs causes lasting alterations in TCA patterning, spatial organizations of cortical neurons, and dendritic arborization in sensory cortex. Pharmacological reduction of serotonin synthesis during the first postnatal week rescues sensory maps in SERTGluΔ mice. Furthermore, knockdown of SERT expression in serotonin-producing neurons does not impair barrel maps. We propose that spatiotemporal SERT expression in non-serotonin-producing neurons represents a determinant in early life genetic programming of cortical circuits. Perturbing this SERT function could be involved in the origin of sensory and cognitive deficits associated with neurodevelopmental disorders.

Datasets in Gene Expression Omnibus used in the study ORD-020969: Genomic effects of androstenedione and sex-specific liver cancer susceptibility in mice

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U.S. Environmental Protection Agency — Datasets in Gene Expression Omnibus used in the study ORD-020969: Genomic effects of androstenedione and sex-specific liver cancer susceptibility in mice. This...

Calorie restriction and dwarf mice in gerontological research.

Science.gov (United States)

McKee Alderman, J; DePetrillo, Michael A; Gluesenkamp, Angela M; Hartley, Antonia C; Verhoff, S Veronica; Zavodni, Katherine L; Combs, Terry P

2010-01-01

What aging process is delayed by calorie restriction (CR) and mutations that produce long-lived dwarf mice? From 1935 until 1996, CR was the only option for increasing the maximum lifespan of laboratory rodents. In 1996, the mutation producing the Ames dwarf mouse (Prop-1(-/-)) was reported to increase lifespan. Since 1996, other gene mutations that cause dwarfism or lower body weight have been reported to increase the lifespan of mice. The recent discovery of long-lived mutant dwarf mice provides an opportunity to investigate common features between CR and dwarf models. Both CR and dwarf mutations increase insulin sensitivity. Elevated insulin sensitivity reduces oxidative stress, a potential cause of aging. The elevation of liver insulin sensitivity by the hormone adiponectin in CR and long-lived dwarf mice can lower endogenous glucose production and raise fatty acid oxidation. Adiponectin reduction of plasma glucose in CR and long-lived dwarf mice can thereby lower age-related increases in oxidative damage and cancer. Copyright 2009 S. Karger AG, Basel.

Mouse mammary tumor viruses expressed by RIII/Sa mice with a high incidence of mammary tumors interact with the Vβ-2- and Vβ-8-specific T cells during viral infection

International Nuclear Information System (INIS)

Uz-Zaman, Taher; Ignatowicz, Leszek; Sarkar, Nurul H.

2003-01-01

The mouse mammary tumor viruses (MMTVs) that induce mammary adenocarcinomas in mice are transmitted from mother to offspring through milk. MMTV infection results in the deletion of specific T cells as a consequence of interaction between the MMTV-encoded superantigen (Sag) and specific Vβ chains of the T cell receptor. The specificity and kinetics of T cell deletion for a number of highly oncogenic MMTVs, such as C3H- and GR-MMTVs, have been studied in great detail. Some work has also been done with the MMTVs expressed in two substrains of RIII mice, BR6 and RIIIS/J, but the nature of the interaction between T cells and the virus(es) that the parental RIII-strain of mice express has not been investigated. Since RIII mice (designated henceforth as RIII/Sa) have a very high incidence (90-98%) of mammary tumors, and they have been extensively used in studies of the biology of mammary tumor development, we have presently determined the pattern of Vβ-T cell deletion caused by RIII/Sa-MMTV-Sag(s) during viral infection. T cells were isolated from lymph nodes and thymus of young RIII/Sa mice, as well as from BALB/c (BALB/cfRIII/Sa), C57BL (C57BLfRIII/Sa), and RIIIS/J (RIIIS/JfRIII/Sa) mice after they were infected with RIII/Sa-MMTV(s) by foster nursing. The composition of the T cells was analyzed by FACS using a panel of monoclonal antibodies specific to a variety of Vβs. Our results show that milk-borne RIII/Sa-MMTV(s) infection leads to the deletion of CD4 + Vβ-2, and to a lesser extent Vβ-8 bearing peripheral and central T cells in RIII/Sa, RIIIS/J, BALB/c, and C57BL mice. Our results are in contrast to the findings that C3H-, GR-, and BR6-MMTVs delete Vβ-14- and/or Vβ-15-specific T cells

Acute toxicity of Psilocybe cubensis (Ear. Sing., Strophariaceae, aqueous extract in mice

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Thiago Berti Kirsten

Full Text Available Psilocybe cubensis (Ear. Sing., Strophariaceae, is a hallucinogen mushroom that has been used since the old times by humans, causing several psychotic effects. P. cubensis contains two tryptamine derivates: psilocybin and psilocin, agonists of the 5-HT2 receptor (serotonin. The main objective of this study was to investigate the acute toxicity effects of P. cubensis aqueous extract (PCAE administration in mice. Male and female adult Swiss mice received PCAE 0.1 mL/10 g i.p., and were observed individually, directly in a glass box and in an open-field. In relation to the data of the control group, PCAE-treated animals presented: an increased gnawing, appearance of wet-dog shakes and a decreased locomotion and rearing frequencies after 29-38 min. Also a clear gender difference was detected, being female mice more sensible to the PCAE than males. It was suggested that PCAE administration produced specific effects on mice behaviors, characteristic of drugs which interfere on central serotonergic and dopaminergic systems. Finally, the observational methods here employed were efficient to evaluate the toxic effects of the extract.

Gender-Specific Neuroimmunoendocrine Response to Treadmill Exercise in 3xTg-AD Mice

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Lydia Giménez-Llort

2010-01-01

Full Text Available The 3xTg-AD mouse develops a progressive Alzheimer's disease- (AD- like brain pathology that causes cognitive- and neuropsychiatric-like symptoms of dementia. Since its neuroimmunoendocrine axis is likewise impaired, this mouse is also useful for modelling complex age-related neurodegeneration. This study analyzed behavioral, physiological, neurochemical, pathological and immunoendocrine alterations in male and female 3xTg-AD mice and assayed the effects of a short therapy of forced physical exercise at the moderate pathology stage of 6 months of age. Gender effects were observed in most AD-related pathology and dysfunctions. Five weeks of treadmill training produced beneficial effects, such as the reduction of brain oxidative stress and GABA-A receptor dysfunction in males and improvement of sensorimotor function in females. In both sexes, exercise decreased the brain amyloid 42/40 ratio levels. The results highlight the importance of analyzing experimental therapies in both mouse model genders in order to improve our understanding of the disease and develop more appropriate therapies.

Modulation of Specific Sensory Cortical Areas by Segregated Basal Forebrain Cholinergic Neurons Demonstrated by Neuronal Tracing and Optogenetic Stimulation in Mice.

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Chaves-Coira, Irene; Barros-Zulaica, Natali; Rodrigo-Angulo, Margarita; Núñez, Ángel

2016-01-01

Neocortical cholinergic activity plays a fundamental role in sensory processing and cognitive functions. Previous results have suggested a refined anatomical and functional topographical organization of basal forebrain (BF) projections that may control cortical sensory processing in a specific manner. We have used retrograde anatomical procedures to demonstrate the existence of specific neuronal groups in the BF involved in the control of specific sensory cortices. Fluoro-Gold (FlGo) and Fast Blue (FB) fluorescent retrograde tracers were deposited into the primary somatosensory (S1) and primary auditory (A1) cortices in mice. Our results revealed that the BF is a heterogeneous area in which neurons projecting to different cortical areas are segregated into different neuronal groups. Most of the neurons located in the horizontal limb of the diagonal band of Broca (HDB) projected to the S1 cortex, indicating that this area is specialized in the sensory processing of tactile stimuli. However, the nucleus basalis magnocellularis (B) nucleus shows a similar number of cells projecting to the S1 as to the A1 cortices. In addition, we analyzed the cholinergic effects on the S1 and A1 cortical sensory responses by optogenetic stimulation of the BF neurons in urethane-anesthetized transgenic mice. We used transgenic mice expressing the light-activated cation channel, channelrhodopsin-2, tagged with a fluorescent protein (ChR2-YFP) under the control of the choline-acetyl transferase promoter (ChAT). Cortical evoked potentials were induced by whisker deflections or by auditory clicks. According to the anatomical results, optogenetic HDB stimulation induced more extensive facilitation of tactile evoked potentials in S1 than auditory evoked potentials in A1, while optogenetic stimulation of the B nucleus facilitated either tactile or auditory evoked potentials equally. Consequently, our results suggest that cholinergic projections to the cortex are organized into segregated

Sex-Specific Diurnal Immobility Induced by Forced Swim Test in Wild Type and Clock Gene Deficient Mice

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Ningyue Li

2015-03-01

Full Text Available Objective: The link between alterations in circadian rhythms and depression are well established, but the underlying mechanisms are far less elucidated. We investigated the circadian characteristics of immobility behavior in wild type (WT mice and mice with mutations in core Clock genes. Methods: All mice were tested with forced swim test (FST at 4 h intervals. Results: These experiments revealed significant diurnal rhythms associated with immobility behavior in both male and female WT mice with sex-different circadian properties. In addition, male mice showed significantly less immobility during the night phase in comparison to female mice. Female Per1Brdm1 mice also showed significant rhythmicity. However, the timing of rhythmicity was very different from that observed in female wild type mice. Male Per1Brdm1 mice showed a pattern of rhythmicity similar to that of wild type mice. Furthermore, female Per1Brdm1 mice showed higher duration of immobility in comparison to male Per1Brdm1 mice in both daytime and early night phases. Neither Per2Brdm1 nor ClockΔ19 mice showed significant rhythmicity, but both female Per2Brdm1 and ClockΔ19 mice had lower levels of immobility, compared to males. Conclusions: This study highlights the differences in the circadian characteristics of immobility induced by FST in WT, ClockΔ19, Per1, and Per2 deficient mice.

Cyp26b1 within the growth plate regulates bone growth in juvenile mice

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Minegishi, Yoshiki; Sakai, Yasuo; Yahara, Yasuhito; Akiyama, Haruhiko; Yoshikawa, Hideki; Hosokawa, Ko; Tsumaki, Noriyuki

2014-01-01

Highlights: • Retinoic acid and Cyp26b1 were oppositely localized in growth plate cartilage. • Cyp26b1 deletion in chondrocytes decreased bone growth in juvenile mice. • Cyp26b1 deletion reduced chondrocyte proliferation and growth plate height. • Vitamin A-depletion partially reversed growth plate abnormalities caused by Cyp26b1 deficiency. • Cyp26b1 regulates bone growth by controlling chondrocyte proliferation. - Abstract: Retinoic acid (RA) is an active metabolite of vitamin A and plays important roles in embryonic development. CYP26 enzymes degrade RA and have specific expression patterns that produce a RA gradient, which regulates the patterning of various structures in the embryo. However, it has not been addressed whether a RA gradient also exists and functions in organs after birth. We found localized RA activities in the diaphyseal portion of the growth plate cartilage were associated with the specific expression of Cyp26b1 in the epiphyseal portion in juvenile mice. To disturb the distribution of RA, we generated mice lacking Cyp26b1 specifically in chondrocytes (Cyp26b1 Δchon cKO). These mice showed reduced skeletal growth in the juvenile stage. Additionally, their growth plate cartilage showed decreased proliferation rates of proliferative chondrocytes, which was associated with a reduced height in the zone of proliferative chondrocytes, and closed focally by four weeks of age, while wild-type mouse growth plates never closed. Feeding the Cyp26b1 cKO mice a vitamin A-deficient diet partially reversed these abnormalities of the growth plate cartilage. These results collectively suggest that Cyp26b1 in the growth plate regulates the proliferation rates of chondrocytes and is responsible for the normal function of the growth plate and growing bones in juvenile mice, probably by limiting the RA distribution in the growth plate proliferating zone

A Novel Closed-head Model of Mild Traumatic Brain Injury Caused by Primary Overpressure Blast to the Cranium Produces Sustained Emotional Deficits in Mice

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Scott A Heldt

2014-01-01

Full Text Available Emotional disorders are a common outcome from mild traumatic brain injury (TBI in humans, but their pathophysiological basis is poorly understood. We have developed a mouse model of closed-head blast injury using an air pressure wave delivered to a small area on one side of the cranium, which we have used to create mild TBI. We found that 20-psi blasts in 3-month old C57BL/6 male mice yielded no obvious behavioral or histological evidence of brain injury, while 25-40 psi blasts produced transient anxiety in an open field arena but little histological evidence of brain damage. By contrast, 50-60 psi blasts resulted in anxiety-like behavior in an open field arena that became more evident with time after blast. In additional behavioral tests conducted 2-8 weeks after blast, 50-60 psi mice also demonstrated increased acoustic startle, perseverance of learned fear, and enhanced contextual fear, as well as depression-like behavior and diminished prepulse inhibition. We found no evident cerebral pathology, however, and only scattered axonal degeneration in brain sections from 50-60 psi mice 3-8 weeks after blast. Thus, the TBI caused by single 50-60 psi blasts in mice exhibits the minimal neuronal loss coupled to diffuse axonal injury characteristic of human mild TBI. A reduction in the abundance of a subpopulation of excitatory projection neurons in basolateral amygdala enriched in Thy1 was, however, observed. The reported link of this neuronal population to fear suppression suggests their damage by mild TBI may contribute to the heightened anxiety and fearfulness observed after blast in our mice. Our overpressure air blast model of concussion in mice will enable further studies of the mechanisms underlying the diverse emotional deficits seen after mild TBI.

Strain-specific induction of experimental autoimmune prostatitis (EAP) in mice.

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Jackson, Christopher M; Flies, Dallas B; Mosse, Claudio A; Parwani, Anil; Hipkiss, Edward L; Drake, Charles G

2013-05-01

Prostatitis, a clinical syndrome characterized by pelvic pain and inflammation, is common in adult males. Although several induced and spontaneous murine models of prostatitis have been explored, the role of genetic background on induction has not been well-defined. Using a standard methodology for the induction of experimental autoimmune prostatitis (EAP), we investigated both acute and chronic inflammation on several murine genetic backgrounds. In our colony, nonobese diabetic (NOD) mice evinced spontaneous prostatitis that was not augmented by immunization with rat prostate extract (RPE). In contrast, the standard laboratory strain Balb/c developed chronic inflammation in response to RPE immunization. Development of EAP in other strains was variable. These data suggest that Balb/c mice injected with RPE may provide a useful model for chronic prostatic inflammation. Copyright © 2012 Wiley Periodicals, Inc.

Vaccination with dengue virus-like particles induces humoral and cellular immune responses in mice

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Zhang Quanfu

2011-06-01

Full Text Available Abstract Background The incidence of dengue, an infectious disease caused by dengue virus (DENV, has dramatically increased around the world in recent decades and is becoming a severe public health threat. However, there is currently no specific treatment for dengue fever, and licensed vaccine against dengue is not available. Vaccination with virus-like particles (VLPs has shown considerable promise for many viral diseases, but the effect of DENV VLPs to induce specific immune responses has not been adequately investigated. Results By optimizing the expression plasmids, recombinant VLPs of four antigenically different DENV serotypes DENV1-4 were successfully produced in 293T cells. The vaccination effect of dengue VLPs in mice showed that monovalent VLPs of each serotype stimulated specific IgG responses and potent neutralizing antibodies against homotypic virus. Tetravalent VLPs efficiently enhanced specific IgG and neutralizing antibodies against all four serotypes of DENV. Moreover, vaccination with monovalent or tetravalent VLPs resulted in the induction of specific cytotoxic T cell responses. Conclusions Mammalian cell expressed dengue VLPs are capable to induce VLP-specific humoral and cellular immune responses in mice, and being a promising subunit vaccine candidate for prevention of dengue virus infection.

β-cell-specific IL-2 therapy increases islet Foxp3+Treg and suppresses type 1 diabetes in NOD mice.

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Johnson, Mark C; Garland, Alaina L; Nicolson, Sarah C; Li, Chengwen; Samulski, R Jude; Wang, Bo; Tisch, Roland

2013-11-01

Interleukin-2 (IL-2) is a critical cytokine for the homeostasis and function of forkhead box p3-expressing regulatory T cells (Foxp3(+)Tregs). Dysregulation of the IL-2-IL-2 receptor axis is associated with aberrant Foxp3(+)Tregs and T cell-mediated autoimmune diseases such as type 1 diabetes. Treatment with recombinant IL-2 has been reported to enhance Foxp3(+)Tregs and suppress different models of autoimmunity. However, efficacy of IL-2 therapy is dependent on achieving sufficient levels of IL-2 to boost tissue-resident Foxp3(+)Tregs while avoiding the potential toxic effects of systemic IL-2. With this in mind, adeno-associated virus (AAV) vector gene delivery was used to localize IL-2 expression to the islets of NOD mice. Injection of a double-stranded AAV vector encoding IL-2 driven by a mouse insulin promoter (dsAAVmIP-IL2) increased Foxp3(+)Tregs in the islets but not the draining pancreatic lymph nodes. Islet Foxp3(+)Tregs in dsAAVmIP-IL2-treated NOD mice exhibited enhanced fitness marked by increased expression of Bcl-2, proliferation, and suppressor function. In contrast, ectopic IL-2 had no significant effect on conventional islet-infiltrating effector T cells. Notably, β-cell-specific IL-2 expression suppressed late preclinical type 1 diabetes in NOD mice. Collectively, these findings demonstrate that β-cell-specific IL-2 expands an islet-resident Foxp3(+)Tregs pool that effectively suppresses ongoing type 1 diabetes long term.

Interleukin-10 is produced by a specific subset of taste receptor cells and critical for maintaining structural integrity of mouse taste buds.

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Feng, Pu; Chai, Jinghua; Zhou, Minliang; Simon, Nirvine; Huang, Liquan; Wang, Hong

2014-02-12

Although inflammatory responses are a critical component in defense against pathogens, too much inflammation is harmful. Mechanisms have evolved to regulate inflammation, including modulation by the anti-inflammatory cytokine interleukin-10 (IL-10). Previously we have shown that taste buds express various molecules involved in innate immune responses, including the proinflammatory cytokine tumor necrosis factor (TNF). Here, using a reporter mouse strain, we show that taste cells also express the anti-inflammatory cytokine IL-10. Remarkably, IL-10 is produced by only a specific subset of taste cells, which are different from the TNF-producing cells in mouse circumvallate and foliate taste buds: IL-10 expression was found exclusively in the G-protein gustducin-expressing bitter receptor cells, while TNF was found in sweet and umami receptor cells as reported previously. In contrast, IL-10R1, the ligand-binding subunit of the IL-10 receptor, is predominantly expressed by TNF-producing cells, suggesting a novel cellular hierarchy for regulating TNF production and effects in taste buds. In response to inflammatory challenges, taste cells can increase IL-10 expression both in vivo and in vitro. These findings suggest that taste buds use separate populations of taste receptor cells that coincide with sweet/umami and bitter taste reception to modulate local inflammatory responses, a phenomenon that has not been previously reported. Furthermore, IL-10 deficiency in mice leads to significant reductions in the number and size of taste buds, as well as in the number of taste receptor cells per taste bud, suggesting that IL-10 plays critical roles in maintaining structural integrity of the peripheral gustatory system.

Intrathymic selection of NK1.1+α/β T cell antigen receptor (TCR)+ cells in transgenic mice bearing TCR specific for chicken ovalbumin and restricted to I-Ad

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Iwabuchi, Chikako; Iwabuchi, Kazuya; Nakagawa, Ken-ichi; Takayanagi, Toshiaki; Nishihori, Hiroki; Tone, Saori; Ogasawara, Kazumasa; Good, Robert A.; Onoé, Kazunori

1998-01-01

Generation and negative selection of NK1.1+α/β T cell receptor (TCR)+ thymocytes were analyzed using TCR-transgenic (B10.D2 × DO10)F1 and (C57BL/6 × DO10)F1 mice and Rag-1−/−/DO10 mice, which had been established by breeding and backcrossing between Rag-1−/− and DO10 mice. Almost all T cells from these mice were shown to bear Vα13/Vβ8.2 that is specific for chicken ovalbumin (cOVA) and restricted to I-Ad. A normal proportion of the NK1.1+ Vα13/Vβ8.2+ thymocytes was generated in these mice. Ho...

Insulin signaling displayed a differential tissue-specific response to low-dose dihydrotestosterone in female mice.

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Andrisse, Stanley; Billings, Katelyn; Xue, Ping; Wu, Sheng

2018-04-01

Hyperandrogenemia and hyperinsulinemia are believed to play prominent roles in polycystic ovarian syndrome (PCOS). We explored the effects of low-dose dihydrotestosterone (DHT), a model of PCOS, on insulin signaling in metabolic and reproductive tissues in a female mouse model. Insulin resistance in the energy storage tissues is associated with type 2 diabetes. Insulin signaling in the ovaries and pituitary either directly or indirectly stimulates androgen production. Energy storage and reproductive tissues were isolated and molecular assays were performed. Livers and white adipose tissue (WAT) from DHT mice displayed lower mRNA and protein expression of insulin signaling intermediates. However, ovaries and pituitaries of DHT mice exhibited higher expression levels of insulin signaling genes/proteins. Insulin-stimulated p-AKT levels were blunted in the livers and WAT of the DHT mice but increased or remained the same in the ovaries and pituitaries compared with controls. Glucose uptake decreased in liver and WAT but was unchanged in pituitary and ovary of DHT mice. Plasma membrane GLUTs were decreased in liver and WAT but increased in ovary and pituitary of DHT mice. Skeletal muscle insulin-signaling genes were not lowered in DHT mice compared with control. DHT mice did not display skeletal muscle insulin resistance. Insulin-stimulated glucose transport increased in skeletal muscles of DHT mice compared with controls. DHT mice were hyperinsulinemic. However, the differential mRNA and protein expression pattern was independent of hyperinsulinemia in cultured hepatocytes and pituitary cells. These findings demonstrate a differential effect of DHT on the insulin-signaling pathway in energy storage vs. reproductive tissues independent of hyperinsulinemia.

The mannosylated extracellular domain of Her2/neu produced in P. pastoris induces protective antitumor immunity

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Dimitriadis, Alexios; Gontinou, Chrysanthi; Baxevanis, Constantin N; Mamalaki, Avgi

2009-01-01

Her2/neu is overexpressed in various human cancers of epithelial origin and is associated with increased metastatic potential and poor prognosis. Several attempts have been made using the extracellular domain of Her2/neu (ECD/Her2) as a prophylactic vaccine in mice with no success in tumor prevention. The extracellular domain of Her2/neu (ECD/Her2) was expressed in yeast P. pastoris, in a soluble highly mannosylated form. The immune response of the immunization with this recombinant ECD/Her2 was analyzed using immunoprecipitation and western blot analysis, proliferation and cytotoxicity assays as well as specific tumor growth assays. Mannosylated ECD/Her2 elicited a humoral response with HER2/neu specific antibodies in vaccinated mice, which were able to reduce the proliferation rate of cancer cells in vitro. Moreover, it elicited a cellular response with Her2/neu-specific CTL capable of lysing tumor cells, in vitro. When immunized Balb/c and HHD mice were challenged with Her2/neu-overexpressing cells, tumor growth was inhibited. Here we report on the efficacy of the extracellular domain of human Her2/neu produced in yeast P. pastoris, which confers mannosylation of the protein, to act as a potent anti-tumor vaccine against Her2/neu overexpressing tumors. Specific cellular and humoral responses were observed as well as efficacy

Loss of neutrophil polarization in colon carcinoma liver metastases of mice with an inducible, liver-specific IGF-I deficiency.

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Rayes, Roni F; Milette, Simon; Fernandez, Maria Celia; Ham, Boram; Wang, Ni; Bourdeau, France; Perrino, Stephanie; Yakar, Shoshana; Brodt, Pnina

2018-03-20

The growth of cancer metastases in the liver depends on a permissive interaction with the hepatic microenvironment and neutrophils can contribute to this interaction, either positively or negatively, depending on their phenotype. Here we investigated the role of IGF-I in the control of the tumor microenvironment in the liver, using mice with a conditional, liver-specific, IGF-I deficiency (iLID) induced by a single tamoxifen injection. In mice that had a sustained (3 weeks) IGF-I deficiency prior to the intrasplenic/portal inoculation of colon carcinoma MC-38 cells, we observed an increase in neutrophil accumulation in the liver relative to controls. However, unlike controls, these neutrophils did not acquire the (anti-inflammatory) tumor-promoting phenotype, as evidenced by retention of high ICAM-1 expression and nitric oxide production and low CXCR4, CCL5, and VEGF expression and arginase production, all characteristic of the (pro-inflammatory) phenotype. This coincided with an increase in apoptotic tumor cells and reduced metastasis. Neutrophils isolated from these mice also had reduced IGF-IR expression levels. These changes were not observed in iLID mice with a short-term (2 days) IGF-I depletion, despite a 70% reduction in their circulating IGF-I levels, indicating that a sustained IGF-I deficiency was necessary to alter the neutrophil phenotype. Similar results were obtained with the highly metastatic Lewis lung carcinoma subline H-59 cells and in mice injected with an IGF-Trap that blocks IGF-IR signaling by reducing ligand bioavailability. Our results implicate the IGF axis in neutrophil polarization and the induction of a pro-metastatic microenvironment in the liver.

A shorter and more specific oral sensitization-based experimental model of food allergy in mice.

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Bailón, Elvira; Cueto-Sola, Margarita; Utrilla, Pilar; Rodríguez-Ruiz, Judith; Garrido-Mesa, Natividad; Zarzuelo, Antonio; Xaus, Jordi; Gálvez, Julio; Comalada, Mònica

2012-07-31

Cow's milk protein allergy (CMPA) is one of the most prevalent human food-borne allergies, particularly in children. Experimental animal models have become critical tools with which to perform research on new therapeutic approaches and on the molecular mechanisms involved. However, oral food allergen sensitization in mice requires several weeks and is usually associated with unspecific immune responses. To overcome these inconveniences, we have developed a new food allergy model that takes only two weeks while retaining the main characters of allergic response to food antigens. The new model is characterized by oral sensitization of weaned Balb/c mice with 5 doses of purified cow's milk protein (CMP) plus cholera toxin (CT) for only two weeks and posterior challenge with an intraperitoneal administration of the allergen at the end of the sensitization period. In parallel, we studied a conventional protocol that lasts for seven weeks, and also the non-specific effects exerted by CT in both protocols. The shorter protocol achieves a similar clinical score as the original food allergy model without macroscopically affecting gut morphology or physiology. Moreover, the shorter protocol caused an increased IL-4 production and a more selective antigen-specific IgG1 response. Finally, the extended CT administration during the sensitization period of the conventional protocol is responsible for the exacerbated immune response observed in that model. Therefore, the new model presented here allows a reduction not only in experimental time but also in the number of animals required per experiment while maintaining the features of conventional allergy models. We propose that the new protocol reported will contribute to advancing allergy research. Copyright © 2012 Elsevier B.V. All rights reserved.

A new mutation, gld, that produces lymphoproliferation and autoimmunity in C3H/HeJ mice.

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Roths, J B; Murphy, E D; Eicher, E M

1984-01-01

A newly discovered autosomal recessive mutation, generalized lymphoproliferative disease (gld), in the C3H/HeJ strain of mice, determines the development of early onset massive lymphoid hyperplasia with autoimmunity. Significant lymph node enlargement is apparent as early as 12 wk of age. By 20 wk, lymph nodes are 50-fold heavier than those of coisogenic C3H/HeJ-+/+ mice. There is a concomitant increase in the numbers of peripheral blood lymphocytes. Analysis of C3H-gld lymph node lymphocyte subsets by immunofluorescence indicates an increase in numbers of B cells, T cells, and null (Thy-1-, sIg-) lymphocytes by 6-, 15-, and 33-fold compared with congeneic control mice. Serologically, gld/gld mice develop antinuclear antibodies (including anti-dsDNA), thymocyte-binding autoantibody, and hypergammaglobulinemia with major increases in several immunoglobulin isotypes. Mutant gld mice live only one-half as long as normal controls (12 and 23 mo, respectively). Interstitial pneumonitis was found in virtually all C3H-gld mice autopsied when moribund. Although immune complexes were detected in the glomerulus by immunofluorescence techniques, only 14% of the autopsied mice had significant lupus-like nephritis. Vascular disease was not found. The pattern of early onset massive lymph node enlargement, hypergammaglobulinemia, and production of antinuclear autoantibodies resembles the basic abnormal phenotype induced by the lpr (lymphoproliferation) mutation. The mutations gld and lpr are not allelic. Linkage studies indicate that gld is located between Pep-3 and Lp on chromosome 1. This new mutation adds another genetically well-defined model to the list of murine lymphoproliferative/autoimmune disorders that may be exploited to gain a clearer understanding of immunoregulatory defects and for identifying common pathogenetic factors involved in systemic autoimmune diseases.

Toxicity of Parasporin-4 and Health Effects of Pro-parasporin-4 Diet in Mice

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Shiro Okumura

2014-07-01

Full Text Available Parasporin-4 (PS4 is an aerolysin-type β-pore-forming toxin produced by Bacillus thuringiensis strain A1470. It exhibits specific cytotoxicity against human cancer cell lines; therefore, it is expected to be useful for the diagnosis and treatment of particular types of cancer cells. We examined the acute toxicity of PS4 on ICR mice. The LD50 value was 160 μg/kg by a subcutaneous route. Potassium, ammonium, magnesium ion, creatinine, and urea nitrogen decreased in urine by the injection of PS4. Simultaneously, creatinine and urea nitrogen in mice serum increased. These results imply that PS4 impairs kidney function in mice. PS4 is obtained from Pro-parasporin-4 (ProPS4 by processing, and ProPS4 is produced by recombinant Escherichia coli as the inclusion body. The inclusion body of ProPS4 can be solubilized in a weak acid solution and activated by pepsin, implying that it would be solubilized and activated in the stomach of mammals after oral administration. Thus, the influence of the oral administration of it by C57BL/6J mice was examined. Although ProPS4 was activated to PS4 in the mouse digestive tract, any serious health hazard was not observed and there was no significant difference in body weight change.

Protective specific immunity induced by doxorubicin plus TNF-alpha combination treatment of EL4 lymphoma-bearing C57BL/6 mice.

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Ehrke, M J; Verstovsek, S; Maccubbin, D L; Ujházy, P; Zaleskis, G; Berleth, E; Mihich, E

2000-07-01

The therapeutic efficacy of a single (day 8), moderate dose (4 mg/kg, i.v.) of doxorubicin (DOX, Adriamycin) combined with recombinant human TNF-alpha (3 different doses and 5 different schedules, i.v.) was evaluated in C57BL/6 mice bearing an implant (s.c.) of the DOX-sensitive, TNF-alpha-resistant EL4 lymphoma. In parallel to monitoring survival, the levels of several host anti-tumor cytolytic effector functions of splenocytes and thymocytes were evaluated throughout the treatment period and in long-term survivors (LTS). DOX treatment alone resulted in a moderate (approx. 20%) increase in life span but no cures. TNF-alpha alone, at any tested dose or schedule, had little or no positive effect on survival. The combinations of DOX and TNF-alpha were only slightly better than DOX alone with respect to the time to death of mice that died (approx. 29% increase); however, each of the combinations involving 1,000 U TNF-alpha/injection produced a fraction (20% to 80%) of LTS. The host defense activities examined included those of splenic and thymic cytolytic T lymphocytes (CTL) and lymphokine-activated killer cells as well as splenic tumoricidal macrophages. Although most activities were modulated by tumor growth and/or treatment, only CTL responsiveness appeared to correlate with survival. CTL activity in the treated groups with LTS was significantly higher than in control groups late in the treatment period. Finally, ex vivo analyses of splenocytes and thymocytes together with the rejection of implanted tumor at 17 months established that LTS displayed specific long-term immune memory. Copyright 2000 Wiley-Liss, Inc.

Conditioned social preference, but not place preference, produced by intranasal oxytocin in female mice.

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Kosaki, Yutaka; Watanabe, Shigeru

2016-04-01

Oxytocin (OT) has been implicated in a variety of mammalian reproductive and social behaviors, and the use of intranasal OT for clinical purposes is on the rise. However, basic actions of OT, including the rewarding or reinforcing properties of the drug, are currently not fully understood. In this study, the authors investigated whether intranasally administered OT has different reinforcing properties for social and nonsocial stimuli and whether such effects are variable between male and female subjects. Conditioned social preference (CSP) and conditioned place preference (CPP) paradigms were used to examine social and nonsocial reinforcing properties of OT. In CSP, the presence of a same-sex unfamiliar conspecific was repeatedly paired with intranasal OT, while a different conspecific was associated with saline. The reinforcing effect of OT was assessed in a postconditioning choice test under a drug-free condition. In CPP, the 2 conspecifics were replaced with nonsocial black and white compartments. The authors found that intranasal OT (12 μg) in females supported the formation of CSP (Experiment 1) but not CPP (Experiment 3). Neither CSP (Experiment 2) nor CPP (Experiment 4) was formed in males. Extended conditioning with higher dose OT (36 μg), however, abolished the initial CSP in females and produced an aversion to the OT-paired stimulus mouse. Experiment 5 indicated that it was the repeated administrations rather than the higher dose that produced the abolition of the original preference. Overall, the current results demonstrate for the first time a sex- and stimulus-dependent reinforcing property of intranasal OT in mice. (PsycINFO Database Record (c) 2016 APA, all rights reserved).

Deletions, Inversions, Duplications: Engineering of Structural Variants using CRISPR/Cas in Mice

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Katerina Kraft

2015-02-01

Full Text Available Structural variations (SVs contribute to the variability of our genome and are often associated with disease. Their study in model systems was hampered until now by labor-intensive genetic targeting procedures and multiple mouse crossing steps. Here we present the use of CRISPR/Cas for the fast (10 weeks and efficient generation of SVs in mice. We specifically produced deletions, inversions, and also duplications at six different genomic loci ranging from 1.1 kb to 1.6 Mb with efficiencies up to 42%. After PCR-based selection, clones were successfully used to create mice via aggregation. To test the practicability of the method, we reproduced a human 500 kb disease-associated deletion and were able to recapitulate the human phenotype in mice. Furthermore, we evaluated the regulatory potential of a large genomic interval by deleting a 1.5 Mb fragment. The method presented permits rapid in vivo modeling of genomic rearrangements.

Neutrophil trails guide influenza-specific CD8+ T cells in the airways

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Lim, Kihong; Hyun, Young-Min; Lambert-Emo, Kris; Capece, Tara; Bae, Seyeon; Miller, Richard; Topham, David J.; Kim, Minsoo

2016-01-01

During viral infections, chemokines guide activated effector T cells to infection sites. However, the cells responsible for producing these chemokines and how such chemokines recruit T cells is unknown. Here, we show that the early recruitment of neutrophils into influenza-infected trachea is essential for CD8+ T cell-mediated immune protection in mice. We observed that migrating neutrophils leave behind long-lasting trails that are enriched in the chemokine CXCL12. Experiments with granulocyte-specific CXCL12 conditional knock-out mice and a CXCR4 antagonist revealed that CXCL12 derived from neutrophil trails is critical for virus-specific CD8+ T cell recruitment and effector functions. Collectively, these results suggest neutrophils deposit long-lasting, chemokine-containing trails, which may provide both chemotactic and haptotactic cues for efficient CD8+ T cell migration and localization in influenza-infected tissues. PMID:26339033

Iron oxide nanoparticles modulate heat shock proteins and organ specific markers expression in mice male accessory organs.

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Sundarraj, Kiruthika; Raghunath, Azhwar; Panneerselvam, Lakshmikanthan; Perumal, Ekambaram

2017-02-15

With increased industrial utilization of iron oxide nanoparticles (Fe 2 O 3 -NPs), concerns on adverse reproductive health effects following exposure have been immensely raised. In the present study, the effects of Fe 2 O 3 -NPs exposure in the seminal vesicle and prostate gland were studied in mice. Mice were exposed to two different doses (25 and 50 mg/kg) of Fe 2 O 3 -NPs along with the control and analyzed the expressions of heat shock proteins (HSP60, HSP70 and HSP90) and organ specific markers (Caltrin, PSP94, and SSLP1). Fe 2 O 3 -NPs decreased food consumption, water intake, and organo-somatic index in mice with elevated iron levels in serum, urine, fecal matter, seminal vesicle and prostate gland. FTIR spectra revealed alterations in the functional groups of biomolecules on Fe 2 O 3 -NPs treatment. These changes are accompanied by increased lactate dehydrogenase levels with decreased total protein and fructose levels. The investigation of oxidative stress biomarkers demonstrated a significant increase in reactive oxygen species, nitric oxide, lipid peroxidation, protein carbonyl content and glutathione peroxidase with a concomitant decrement in the glutathione and ascorbic acid in the male accessory organs which confirmed the induction of oxidative stress. An increase in NADPH-oxidase-4 with a decrease in glutathione-S-transferase was observed in the seminal vesicle and prostate gland of the treated groups. An alteration in HSP60, HSP70, HSP90, Caltrin, PSP94, and SSLP1 expression was also observed. Moreover, accumulation of Fe 2 O 3 -NPs brought pathological changes in the seminal vesicle and prostate gland of treated mice. These findings provide evidence that Fe 2 O 3 -NPs could be an environmental risk factor for reproductive disease. Copyright © 2017 Elsevier Inc. All rights reserved.

Ontogeny and localization of the cells produce IL-2 in healthy animals.

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Yamamoto, Mutsumi; Seki, Yoichi; Iwai, Kazuyuki; Ko, Iei; Martin, Alicia; Tsuji, Noriko; Miyagawa, Shuji; Love, Robert B; Iwashima, Makio

2013-03-01

IL-2 is a growth factor for activated T cells and is required for maintenance of naturally arising regulatory T cells (nTregs). Mice defective in IL-2/IL-2 receptor signaling pathways have impaired nTregs and suffer from lymphoproliferative disorders, suggesting that IL-2 is present and functional in healthy animals. However, the cellular source of IL-2 is currently unknown. To determine which cells produce IL-2 in healthy animals, we established mice carrying cre gene knock in at the il-2 locus (termed IL-2(cre)). When IL-2(cre) mice were crossed with EGFP reporter mice, EGFP was exclusively expressed by a fraction of CD4 T cells present in both lymphoid and non-lymphoid tissues. Live imaging of IL-2(cre) mice that carry the luciferase reporter showed concentrated localization of luciferase(+) cells in Peyer's patches. These cells were not observed in new born mice but appeared within 3days after birth. Reduction of antigen receptor repertoire by transgene expression reduced their number, indicating that recognition of environmental antigens is necessary for generation of these IL-2 producers in healthy animals. A substantial fraction of EGFP(+) cells also produce IL-10 and IFN-γ, a characteristic profile of type 1 regulatory T cells (Tr1). The data suggest that a group of Tr1 cells have addition roles in immune homeostasis by producing IL-2 along with other cytokines and help maintaining Tregs. Copyright © 2012 Elsevier Ltd. All rights reserved.

Enzyme immunoassay for measurement of murine plasminogen activator inhibitor-1, employing a specific antibody produced by the DNA vaccine method.

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Yamada, Takayuki; Takagi, Akira; Takeshita, Kyosuke; Yamamoto, Koji; Ito, Masafumi; Matsushita, Tadashi; Murate, Takashi; Saito, Hidehiko; Kojima, Tetsuhito

2003-01-01

We developed a sensitive immunoassay to determine the concentration of mouse plasminogen activator inhibitor-1. The assay was a non-competitive sandwich enzyme-linked immunosorbent assay (ELISA) based on the production of a specific polyclonal antibody against mouse plasminogen activator inhibitor type-1 (PAI-1) used both as a trapping and detecting antibody. This antibody was raised in a rabbit by direct introduction of the expression vector plasmid DNA encoding mouse PAI-1, instead of conventional immunization with the purified protein. The standard curve was constructed with a recombinant glutathione S-transferase (GST)-mouse PAI-1 fusion protein (GST-mPAI-1) and dose-response of the assay was linear for GST-mPAI-1 between 6.25 and 100 pM. In order to assess the consistency of the assay, we measured PAI-1 antigen in normal mouse pooled plasma several times. We found that the intra-assay and inter-assay coefficients of variation (CV) were 4.8% and 9.2%, respectively, indicating that the ELISA would be sufficiently repeatable and reproducible. In this assay, lipopolysaccharide (LPS)-injected mice showed substantially higher levels (22-fold) of plasma PAI-1 antigen than did control mice (12.5+/-2.4 vs. 0.58+/-0.16 nM), similar to results reported elsewhere. Taken together, the DNA vaccine method is extremely useful for preparing specific antibodies against mouse PAI-1, which can be utilized to establish the ELISA and analyze the profile of PAI-1 distributions in mice under various conditions. This approach might also be useful for immunological investigation of other coagulation factors and related proteins.

Dengue virus infection induces broadly cross-reactive human IgM antibodies that recognize intact virions in humanized BLT-NSG mice.

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Jaiswal, Smita; Smith, Kenneth; Ramirez, Alejandro; Woda, Marcia; Pazoles, Pamela; Shultz, Leonard D; Greiner, Dale L; Brehm, Michael A; Mathew, Anuja

2015-01-01

The development of small animal models that elicit human immune responses to dengue virus (DENV) is important since prior immunity is a major risk factor for developing severe dengue disease. This study evaluated anti-DENV human antibody (hAb) responses generated from immortalized B cells after DENV-2 infection in NOD-scid IL2rγ(null) mice that were co-transplanted with human fetal thymus and liver tissues (BLT-NSG mice). DENV-specific human antibodies predominantly of the IgM isotype were isolated during acute infection and in convalescence. We found that while a few hAbs recognized the envelope protein produced as a soluble recombinant, a number of hAbs only recognized epitopes on intact virions. The majority of the hAbs isolated during acute infection and in immune mice were serotype-cross-reactive and poorly neutralizing. Viral titers in immune BLT-NSG mice were significantly decreased after challenge with a clinical strain of dengue. DENV-specific hAbs generated in BLT-NSG mice share some of the characteristics of Abs isolated in humans with natural infection. Humanized BLT-NSG mice provide an attractive preclinical platform to assess the immunogenicity of candidate dengue vaccines. © 2014 by the Society for Experimental Biology and Medicine.

Fucosylation Deficiency in Mice Leads to Colitis and Adenocarcinoma.

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Wang, Yiwei; Huang, Dan; Chen, Kai-Yuan; Cui, Min; Wang, Weihuan; Huang, Xiaoran; Awadellah, Amad; Li, Qing; Friedman, Ann; Xin, William W; Di Martino, Luca; Cominelli, Fabio; Miron, Alex; Chan, Ricky; Fox, James G; Xu, Yan; Shen, Xiling; Kalady, Mathew F; Markowitz, Sanford; Maillard, Ivan; Lowe, John B; Xin, Wei; Zhou, Lan

2017-01-01

De novo synthesis of guanosine diphosphate (GDP)-fucose, a substrate for fucosylglycans, requires sequential reactions mediated by GDP-mannose 4,6-dehydratase (GMDS) and GDP-4-keto-6-deoxymannose 3,5-epimerase-4-reductase (FX or tissue specific transplantation antigen P35B [TSTA3]). GMDS deletions and mutations are found in 6%-13% of colorectal cancers; these mostly affect the ascending and transverse colon. We investigated whether a lack of fucosylation consequent to loss of GDP-fucose synthesis contributes to colon carcinogenesis. FX deficiency and GMDS deletion produce the same biochemical phenotype of GDP-fucose deficiency. We studied a mouse model of fucosylation deficiency (Fx-/- mice) and mice with the full-length Fx gene (controls). Mice were placed on standard chow or fucose-containing diet (equivalent to a control fucosylglycan phenotype). Colon tissues were collected and analyzed histologically or by enzyme-linked immunosorbent assays to measure cytokine levels; T cells also were collected and analyzed. Fecal samples were analyzed by 16s ribosomal RNA sequencing. Mucosal barrier function was measured by uptake of fluorescent dextran. We transplanted bone marrow cells from Fx-/- or control mice (Ly5.2) into irradiated 8-week-old Fx-/- or control mice (Ly5.1). We performed immunohistochemical analyses for expression of Notch and the hes family bHLH transcription factor (HES1) in colon tissues from mice and a panel of 60 human colorectal cancer specimens (27 left-sided, 33 right-sided). Fx-/- mice developed colitis and serrated-like lesions. The intestinal pathology of Fx-/- mice was reversed by addition of fucose to the diet, which restored fucosylation via a salvage pathway. In the absence of fucosylation, dysplasia appeared and progressed to adenocarcinoma in up to 40% of mice, affecting mainly the right colon and cecum. Notch was not activated in Fx-/- mice fed standard chow, leading to decreased expression of its target Hes1. Fucosylation deficiency

Male bovine GH transgenic mice have decreased adiposity with an adipose depot-specific increase in immune cell populations.

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Benencia, Fabian; Harshman, Stephanie; Duran-Ortiz, Silvana; Lubbers, Ellen R; List, Edward O; Householder, Lara; Al-Naeeli, Mawadda; Liang, Xiaoyu; Welch, Lonnie; Kopchick, John J; Berryman, Darlene E

2015-05-01

White adipose tissue (WAT) is composed of mature adipocytes and a stromal vascular fraction (SVF), which contains a variety of cells, including immune cells that vary among the different WAT depots. Growth hormone (GH) impacts immune function and adiposity in an adipose depot-specific manner. However, its effects on WAT immune cell populations remain unstudied. Bovine GH transgenic (bGH) mice are commonly used to study the in vivo effects of GH. These giant mice have an excess of GH action, impaired glucose metabolism, decreased adiposity, increased lean mass, and a shortened lifespan. Therefore, the purpose of this study was to characterize the WAT depot-specific differences in immune cell populations in the presence of excess GH in vivo. Three WAT depots were assessed: inguinal (sc), epididymal (EPI), and mesenteric (MES). Subcutaneous and MES bGH WAT depots showed a significantly higher number of total SVF cells, yet only MES bGH WAT had higher leukocyte counts compared with control samples. By means of flow cytometry analysis of the SVF, we detected greater macrophage and regulatory T-cell infiltration in sc and MES bGH WAT depots compared with controls. However, no differences were observed in the EPI WAT depot. RNA-sequencing confirmed significant alterations in pathways related to T-cell infiltration and activation in the sc depot with fewer significant changes in the EPI bGH WAT depot. These findings collectively point to a previously unrecognized role for GH in influencing the distribution of WAT immune cell populations in a depot-specific manner.

Aging-associated renal disease in mice is fructokinase dependent.

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Roncal-Jimenez, Carlos A; Ishimoto, Takuji; Lanaspa, Miguel A; Milagres, Tamara; Hernando, Ana Andres; Jensen, Thomas; Miyazaki, Makoto; Doke, Tomohito; Hayasaki, Takahiro; Nakagawa, Takahiko; Marumaya, Shoichi; Long, David A; Garcia, Gabriela E; Kuwabara, Masanari; Sánchez-Lozada, Laura G; Kang, Duk-Hee; Johnson, Richard J

2016-10-01

Aging-associated kidney disease is usually considered a degenerative process associated with aging. Recently, it has been shown that animals can produce fructose endogenously, and that this can be a mechanism for causing kidney damage in diabetic nephropathy and in association with recurrent dehydration. We therefore hypothesized that low-level metabolism of endogenous fructose might play a role in aging-associated kidney disease. Wild-type and fructokinase knockout mice were fed a normal diet for 2 yr that had minimal (renal injury was amplified by provision of high-salt diet for 3 wk, as noted by the presence of glomerular hypertrophy, mesangial matrix expansion, and alpha smooth muscle actin expression, and with segmental thrombi. Fructokinase knockout mice were protected from renal injury both at baseline and after high salt intake (3 wk) compared with wild-type mice. This was associated with higher levels of active (phosphorylated serine 1177) endothelial nitric oxide synthase in their kidneys. These studies suggest that aging-associated renal disease might be due to activation of specific metabolic pathways that could theoretically be targeted therapeutically, and raise the hypothesis that aging-associated renal injury may represent a disease process as opposed to normal age-related degeneration.

Effects of Specific Multi-Nutrient Enriched Diets on Cerebral Metabolism, Cognition and Neuropathology in AβPPswe-PS1dE9 Mice

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Jansen, Diane; Zerbi, Valerio; Arnoldussen, Ilse A. C.; Wiesmann, Maximilian; Rijpma, Anne; Fang, Xiaotian T.; Dederen, Pieter J.; Mutsaers, Martina P. C.; Broersen, Laus M.; Lütjohann, Dieter; Miller, Malgorzata; Joosten, Leo A. B.; Heerschap, Arend; Kiliaan, Amanda J.

2013-01-01

Recent studies have focused on the use of multi-nutrient dietary interventions in search of alternatives for the treatment and prevention of Alzheimer's disease (AD). In this study we investigated to which extent long-term consumption of two specific multi-nutrient diets can modulate AD-related etiopathogenic mechanisms and behavior in 11-12-month-old AβPPswe-PS1dE9 mice. Starting from 2 months of age, male AβPP-PS1 mice and wild-type littermates were fed either a control diet, the DHA+EPA+UMP (DEU) diet enriched with uridine monophosphate (UMP) and the omega-3 fatty acids docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), or the Fortasyn® Connect (FC) diet enriched with the DEU diet plus phospholipids, choline, folic acid, vitamins and antioxidants. We performed behavioral testing, proton magnetic resonance spectroscopy, immunohistochemistry, biochemical analyses and quantitative real-time PCR to gain a better understanding of the potential mechanisms by which these multi-nutrient diets exert protective properties against AD. Our results show that both diets were equally effective in changing brain fatty acid and cholesterol profiles. However, the diets differentially affected AD-related pathologies and behavioral measures, suggesting that the effectiveness of specific nutrients may depend on the dietary context in which they are provided. The FC diet was more effective than the DEU diet in counteracting neurodegenerative aspects of AD and enhancing processes involved in neuronal maintenance and repair. Both diets elevated interleukin-1β mRNA levels in AβPP-PS1 and wild-type mice. The FC diet additionally restored neurogenesis in AβPP-PS1 mice, decreased hippocampal levels of unbound choline-containing compounds in wild-type and AβPP-PS1 animals, suggesting diminished membrane turnover, and decreased anxiety-related behavior in the open field behavior. In conclusion, the current data indicate that specific multi-nutrient diets can influence AD

Effects of specific multi-nutrient enriched diets on cerebral metabolism, cognition and neuropathology in AβPPswe-PS1dE9 mice.

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Diane Jansen

Full Text Available Recent studies have focused on the use of multi-nutrient dietary interventions in search of alternatives for the treatment and prevention of Alzheimer's disease (AD. In this study we investigated to which extent long-term consumption of two specific multi-nutrient diets can modulate AD-related etiopathogenic mechanisms and behavior in 11-12-month-old AβPPswe-PS1dE9 mice. Starting from 2 months of age, male AβPP-PS1 mice and wild-type littermates were fed either a control diet, the DHA+EPA+UMP (DEU diet enriched with uridine monophosphate (UMP and the omega-3 fatty acids docosahexaenoic acid (DHA and eicosapentaenoic acid (EPA, or the Fortasyn® Connect (FC diet enriched with the DEU diet plus phospholipids, choline, folic acid, vitamins and antioxidants. We performed behavioral testing, proton magnetic resonance spectroscopy, immunohistochemistry, biochemical analyses and quantitative real-time PCR to gain a better understanding of the potential mechanisms by which these multi-nutrient diets exert protective properties against AD. Our results show that both diets were equally effective in changing brain fatty acid and cholesterol profiles. However, the diets differentially affected AD-related pathologies and behavioral measures, suggesting that the effectiveness of specific nutrients may depend on the dietary context in which they are provided. The FC diet was more effective than the DEU diet in counteracting neurodegenerative aspects of AD and enhancing processes involved in neuronal maintenance and repair. Both diets elevated interleukin-1β mRNA levels in AβPP-PS1 and wild-type mice. The FC diet additionally restored neurogenesis in AβPP-PS1 mice, decreased hippocampal levels of unbound choline-containing compounds in wild-type and AβPP-PS1 animals, suggesting diminished membrane turnover, and decreased anxiety-related behavior in the open field behavior. In conclusion, the current data indicate that specific multi-nutrient diets can

Surface protein Adr2 of Rickettsia rickettsii induced protective immunity against Rocky Mountain spotted fever in C3H/HeN mice.

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Gong, Wenping; Xiong, Xiaolu; Qi, Yong; Jiao, Jun; Duan, Changsong; Wen, Bohai

2014-04-11

Rickettsia rickettsii is the pathogen of Rocky Mountain spotted fever (RMSF), a life-threatening tick-transmitted infection. Adr2 was a surface-exposed adhesion protein of R. rickettsii and its immunoprotection against RMSF was investigated in mice. Recombinant Adr2 (rAdr2) was used to immunize C3H/HeN mice, and the rickettsial loads in organs of the mice were detected after challenge with R. rickettsii. The levels of specific antibodies of sera from the immunized mice were determined and the sera from immunized mice were applied to neutralize R. rickettsii. Proliferation and cytokine secretion of CD4(+) and CD8(+) T cells isolated from R. rickettsii-infected mice were also assayed after rAdr2 stimulation. After R. rickettsii challenge, the rickettsial loads in spleens, livers, and lungs were significantly lower and the impairment degrees of these organs in rAdr2-immunized mice were markedly slighter, compared with those in negative control mice. The ratio of specific IgG2a/IgG1 of rAdr2-immunized mice kept increasing during the immunization. After treatment with rAdr2-immunized sera, the total number of R. rickettsii organisms adhering and invading host cells was significantly lower than that treated with PBS-immunized sera. Interferon-γ secretion by CD4(+) or CD8(+) T cells and tumor necrosis factor-α secretion by CD4(+) T cells from R. rickettsii-infected mice were respectively significantly greater than those from uninfected mice after rAdr2 stimulation. Adr2 is a protective antigen of R. rickettsii. Protection offered by Adr2 is mainly dependent on antigen-specific cell-mediated immune responses, including efficient activity of CD4(+) and CD8(+) T cells to produce great amount of TNF-α and/or IFN-γ as well as rapid increase of specific IgG2a, which synergistically activate and opsonize host cells to killing intracellular rickettsiae. Copyright © 2014 Elsevier Ltd. All rights reserved.

Gene expression signatures of radiation response are specific, durable and accurate in mice and humans.

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Sarah K Meadows

2008-04-01

Full Text Available Previous work has demonstrated the potential for peripheral blood (PB gene expression profiling for the detection of disease or environmental exposures.We have sought to determine the impact of several variables on the PB gene expression profile of an environmental exposure, ionizing radiation, and to determine the specificity of the PB signature of radiation versus other genotoxic stresses. Neither genotype differences nor the time of PB sampling caused any lessening of the accuracy of PB signatures to predict radiation exposure, but sex difference did influence the accuracy of the prediction of radiation exposure at the lowest level (50 cGy. A PB signature of sepsis was also generated and both the PB signature of radiation and the PB signature of sepsis were found to be 100% specific at distinguishing irradiated from septic animals. We also identified human PB signatures of radiation exposure and chemotherapy treatment which distinguished irradiated patients and chemotherapy-treated individuals within a heterogeneous population with accuracies of 90% and 81%, respectively.We conclude that PB gene expression profiles can be identified in mice and humans that are accurate in predicting medical conditions, are specific to each condition and remain highly accurate over time.

Clonal analysis of T-cell responses to herpes simplex virus: isolation, characterization and antiviral properties of an antigen-specific helper T-cell clone.

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Leung, K N; Nash, A A; Sia, D Y; Wildy, P

1984-12-01

A herpes simplex virus (HSV)-specific long-term T-cell clone has been established from the draining lymph node cells of BALB/c mice; the cells required repeated in vitro restimulation with UV-irradiated virus. The established T-cell clone expresses the Thy-1 and Lyt-1+2,3- surface antigens. For optimal proliferation of the cloned cells, both the presence of specific antigen and an exogenous source of T-cell growth factor are required. The proliferative response of the cloned T cells was found to be virus-specific but it did not distinguish between HSV-1 and HSV-2. Adoptive cell transfer of the cloned T cells helped primed B cells to produce anti-herpes antibodies: the response was antigen-specific and cell dose-dependent. The clone failed to produce a significant DTH reaction in vivo, but did produce high levels of macrophage-activating factor. Furthermore, the T-cell clone could protect from HSV infection, as measured by a reduction in local virus growth, and by enhanced survival following the challenge of mice with a lethal dose of virus. The mechanism(s) whereby this clone protects in vivo is discussed.

SPECT imaging of D2 dopamine receptors and endogenous dopamine release in mice

International Nuclear Information System (INIS)

Jongen, Cynthia; Bruin, Kora de; Booij, Jan; Beekman, Freek

2008-01-01

The dopamine D 2 receptor (D2R) is important in the mediation of addiction. [ 123 I]iodobenzamide (IBZM), a SPECT ligand for the D2R, has been used for in vivo studies of D2R availability in humans, monkeys, and rats. Although mouse models are important in the study of addiction, [ 123 I]IBZM has not been used in mice SPECT studies. This study evaluates the use of [ 123 I]IBZM for measuring D2R availability in mice. Pharmacokinetics of [ 123 I]IBZM in mice were studied with pinhole SPECT imaging after intravenous (i.v.) injection of [ 123 I]IBZM (20, 40, and 70 MBq). In addition, the ability to measure the release of endogenous dopamine after amphetamine administration with [ 123 I]IBZM SPECT was investigated. Thirdly, i.v. administration, the standard route of administration, and intraperitoneal (i.p.) administration of [ 123 I]IBZM were compared. Specific binding of [ 123 I]IBZM within the mouse striatum could be clearly visualized with SPECT. Peak specific striatal binding ratios were reached around 90 min post-injection. After amphetamine administration, the specific binding ratios of [ 123 I]IBZM decreased significantly (-27.2%; n=6; p=0.046). Intravenous administration of [ 123 I]IBZM led to significantly higher specific binding than i.p. administration of the same dose. However, we found that i.v. administration of a dose of 70 MBq [ 123 I]IBZM might result in acute ethanol intoxication because ethanol is used as a preparative aid for the routine production of [ 123 I]IBZM. Imaging of D2R availability and endogenous dopamine release in mice is feasible using [ 123 I]IBZM single pinhole SPECT. Using commercially produced [ 123 I]IBZM, a dose of 40 MBq injected i.v. can be recommended. (orig.)

Aberrant phenotypes of transgenic mice expressing dimeric human erythropoietin

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Yun Seong-Jo

2012-01-01

Full Text Available Abstract Background Dimeric human erythropoietin (dHuEPO peptides are reported to exhibit significantly higher biological activity than the monomeric form of recombinant EPO. The objective of this study was to produce transgenic (tg mice expressing dHuEPO and to investigate the characteristics of these mice. Methods A dHuEPO-expressing vector under the control of the goat beta-casein promoter, which produced a dimer of human EPO molecules linked by a 2-amino acid peptide linker (Asp-Ile, was constructed and injected into 1-cell fertilized embryos by microinjection. Mice were screened using genomic DNA samples obtained from tail biopsies. Blood samples were obtained by heart puncture using heparinized tubes, and hematologic parameters were assessed. Using the microarray analysis tool, we analyzed differences in gene expression in the spleens of tg and control mice. Results A high rate of spontaneous abortion or death of the offspring was observed in the recipients of dHuEPO embryos. We obtained 3 founder lines (#4, #11, and #47 of tg mice expressing the dHuEPO gene. However, only one founder line showed stable germline integration and transmission, subsequently establishing the only transgenic line (#11. We obtained 2 F1 mice and 3 F2 mice from line #11. The dHuEPO protein could not be obtained because of repeated spontaneous abortions in the tg mice. Tg mice exhibited symptoms such as short lifespan and abnormal blood composition. The red blood cell count, white blood cell count, and hematocrit levels in the tg mice were remarkably higher than those in the control mice. The spleens of the tg mice (F1 and F2 females were 11- and -21-fold larger than those of the control mice. Microarray analysis revealed 2,672 spleen-derived candidate genes; more genes were downregulated than upregulated (849/764. Reverse transcriptase-polymerase chain reaction (RT-PCR and quantitative real-time PCR (qRT-PCR were used for validating the results of the microarray

Cardiac overexpression of Mammalian enabled (Mena) exacerbates heart failure in mice.

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Belmonte, Stephen L; Ram, Rashmi; Mickelsen, Deanne M; Gertler, Frank B; Blaxall, Burns C

2013-09-15

Mammalian enabled (Mena) is a key regulator of cytoskeletal actin dynamics, which has been implicated in heart failure (HF). We have previously demonstrated that cardiac Mena deletion produced cardiac dysfunction with conduction abnormalities and hypertrophy. Moreover, elevated Mena expression correlates with HF in human and animal models, yet the precise role of Mena in cardiac pathophysiology is unclear. In these studies, we evaluated mice with cardiac myocyte-specific Mena overexpression (TTA/TgTetMena) comparable to that observed in cardiac pathology. We found that the hearts of TTA/TgTetMena mice were functionally and morphologically comparable to wild-type littermates, except for mildly increased heart mass in the transgenic mice. Interestingly, TTA/TgTetMena mice were particularly susceptible to cardiac injury, as these animals experienced pronounced decreases in ejection fraction and fractional shortening as well as heart dilatation and hypertrophy after transverse aortic constriction (TAC). By "turning off" Mena overexpression in TTA/TgTetMena mice either immediately prior to or immediately after TAC surgery, we discovered that normalizing Mena levels eliminated cardiac hypertrophy in TTA/TgTetMena animals but did not preclude post-TAC cardiac functional deterioration. These findings indicate that hearts with increased levels of Mena fare worse when subjected to cardiac injury and suggest that Mena contributes to HF pathophysiology.

A non-specific effect associated with conditional transgene expression based on Cre-loxP strategy in mice.

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Linghua Qiu

Full Text Available Transgenes flanked by loxP sites have been widely used to generate transgenic mice where the transgene expression can be controlled spatially and temporally by Cre recombinase. Data from this approach has led to important conclusions in cancer, neurodevelopment and neurodegeneration. Using this approach to conditionally express micro RNAs (miRNAs in mice, we found that Cre-mediated recombination in neural progenitor cells caused microcephaly in five of our ten independent transgenic lines. This effect was not associated with the types or the quantity of miRNAs being expressed, nor was it associated with specific target knockdown. Rather, it was correlated with the presence of multiple tandem transgene copies and inverted (head-to-head or tail-to-tail transgene repeats. The presence of these inverted repeats caused a high level of cell death in the ventricular zone of the embryonic brain, where Cre was expressed. Therefore, results from this Cre-loxP approach to generate inducible transgenic alleles must be interpreted with caution and conclusions drawn in previous reports may need reexamination.

Skeletal muscles of aged male mice fail to adapt following contractile activity.

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Vasilaki, A; Iwanejko, L M; McArdle, F; Broome, C S; Jackson, M J; McArdle, A

2003-04-01

Skeletal muscle adapts rapidly following exercise by the increased production of heat-shock proteins (HSPs). The aim of this study was to examine the ability of muscle from adult and aged mice to produce HSPs following non-damaging exercise. Adult and aged B6XSJL mice were anaesthetized and their hind limbs were subjected to isometric contractions. At different time points, muscles were analysed for HSP production by Western and Northern blotting and by electrophoretic mobility-shift assay. HSP protein and mRNA levels in muscles from adult mice increased significantly following exercise. This was not evident in muscles of aged mice. In contrast, binding of the transcription factor heat-shock factor 1 (HSF1) was not grossly altered in muscles of aged mice compared with adult mice. The data suggest that the inability of muscles of aged mice to produce HSPs appears to be due to alterations during gene transcription.

Deletion of PEA-15 in mice is associated with specific impairments of spatial learning abilities

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Hale Gregory

2009-11-01

Full Text Available Abstract Background PEA-15 is a phosphoprotein that binds and regulates ERK MAP kinase and RSK2 and is highly expressed throughout the brain. PEA-15 alters c-Fos and CREB-mediated transcription as a result of these interactions. To determine if PEA-15 contributes to the function of the nervous system we tested mice lacking PEA-15 in a series of experiments designed to measure learning, sensory/motor function, and stress reactivity. Results We report that PEA-15 null mice exhibited impaired learning in three distinct spatial tasks, while they exhibited normal fear conditioning, passive avoidance, egocentric navigation, and odor discrimination. PEA-15 null mice also had deficient forepaw strength and in limited instances, heightened stress reactivity and/or anxiety. However, these non-cognitive variables did not appear to account for the observed spatial learning impairments. The null mice maintained normal weight, pain sensitivity, and coordination when compared to wild type controls. Conclusion We found that PEA-15 null mice have spatial learning disabilities that are similar to those of mice where ERK or RSK2 function is impaired. We suggest PEA-15 may be an essential regulator of ERK-dependent spatial learning.

Vocal ontogeny in neotropical singing mice (Scotinomys.

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Polly Campbell

Full Text Available Isolation calls produced by dependent young are a fundamental form of communication. For species in which vocal signals remain important to adult communication, the function and social context of vocal behavior changes dramatically with the onset of sexual maturity. The ontogenetic relationship between these distinct forms of acoustic communication is surprisingly under-studied. We conducted a detailed analysis of vocal development in sister species of Neotropical singing mice, Scotinomys teguina and S. xerampelinus. Adult singing mice are remarkable for their advertisement songs, rapidly articulated trills used in long-distance communication; the vocal behavior of pups was previously undescribed. We recorded 30 S. teguina and 15 S. xerampelinus pups daily, from birth to weaning; 23 S. teguina and 11 S. xerampelinus were recorded until sexual maturity. Like other rodent species with poikilothermic young, singing mice were highly vocal during the first weeks of life and stopped vocalizing before weaning. Production of first advertisement songs coincided with the onset of sexual maturity after a silent period of ≧2 weeks. Species differences in vocal behavior emerged early in ontogeny and notes that comprise adult song were produced from birth. However, the organization and relative abundance of distinct note types was very different between pups and adults. Notably, the structure, note repetition rate, and intra-individual repeatability of pup vocalizations did not become more adult-like with age; the highly stereotyped structure of adult song appeared de novo in the first songs of young adults. We conclude that, while the basic elements of adult song are available from birth, distinct selection pressures during maternal dependency, dispersal, and territorial establishment favor major shifts in the structure and prevalence of acoustic signals. This study provides insight into how an evolutionarily conserved form of acoustic signaling provides

Visualization of the human CD4+ T-cell response in humanized HLA-DR4-expressing NOD/Shi-scid/γcnull (NOG) mice by retrogenic expression of the human TCR gene

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Takahashi, Takeshi; Katano, Ikumi; Ito, Ryoji; Ito, Mamoru

2015-01-01

Highlights: • β-Lactoglobulin (BLG) specific TCR genes were introduced to human HSC by retrovirus. • Human HSC with BLG-specific TCR were transplanted into NOG-HLA-DR4 I-A −/− mice. • BLG-specific TCR induced positive selection of thymocytes. • BLG-specific TCR positive CD4 + T cells mediated immune responses in humanized mice. - Abstract: The development of severe immunodeficient mouse strains containing various human genes, including cytokines or HLA, has enabled the reconstitution of functional human immune systems after transplantation of human hematopoietic stem cells (HSC). Accumulating evidence has suggested that HLA-restricted antigen-specific human T-cell responses can be generated in these humanized mice. To directly monitor immune responses of human CD4 + T cells, we introduced β-lactoglobulin (BLG)-specific T cell receptor (TCR) genes derived from CD4 + T-cell clones of cow-milk allergy patients into HSCs, and subsequently transplanted them into NOG-HLA-DR4 transgenic/I-Aβ deficient mice (NOG-DR4/I-A o ). In the thymus, thymocytes with BLG-specific TCR preferentially differentiated into CD4 + CD8 − single-positive cells. Adoptive transfer of mature CD4 + T cells expressing the TCR into recipient NOG-DR4/I-A o mice demonstrated that human CD4 + T cells proliferated in response to antigenic stimulation and produced IFN-γ in vivo, suggesting that functional T-cell reactions (especially Th1-skewed responses) were induced in humanized mice

Mechanisms of dietary response in mice and primates: a role for EGR1 in regulating the reaction to human-specific nutritional content.

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Kai Weng

Full Text Available Humans have a widely different diet from other primate species, and are dependent on its high nutritional content. The molecular mechanisms responsible for adaptation to the human diet are currently unknown. Here, we addressed this question by investigating whether the gene expression response observed in mice fed human and chimpanzee diets involves the same regulatory mechanisms as expression differences between humans and chimpanzees.Using mouse and primate transcriptomic data, we identified the transcription factor EGR1 (early growth response 1 as a putative regulator of diet-related differential gene expression between human and chimpanzee livers. Specifically, we predict that EGR1 regulates the response to the high caloric content of human diets. However, we also show that close to 90% of the dietary response to the primate diet found in mice, is not observed in primates. This might be explained by changes in tissue-specific gene expression between taxa.Our results suggest that the gene expression response to the nutritionally rich human diet is partially mediated by the transcription factor EGR1. While this EGR1-driven response is conserved between mice and primates, the bulk of the mouse response to human and chimpanzee dietary differences is not observed in primates. This result highlights the rapid evolution of diet-related expression regulation and underscores potential limitations of mouse models in dietary studies.

Lipid metabolism and body composition in Gclm(-/-) mice

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Kendig, Eric L. [Department of Environmental Health, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States); Center for Environmental Genetics, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States); Chen, Ying [Department of Pharmaceutical Sciences, School of Pharmacy, University of Colorado Denver, Aurora, CO 80045 (United States); Krishan, Mansi; Johansson, Elisabet; Schneider, Scott N. [Department of Environmental Health, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States); Genter, Mary Beth; Nebert, Daniel W. [Department of Environmental Health, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States); Center for Environmental Genetics, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States); Shertzer, Howard G., E-mail: shertzhg@ucmail.uc.edu [Department of Environmental Health, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States); Center for Environmental Genetics, University of Cincinnati Medical Center, P.O. Box 670056, Cincinnati, OH 45267 (United States)

2011-12-15

In humans and experimental animals, high fat diets (HFD) are associated with risk factors for metabolic diseases, such as excessive weight gain and adiposity, insulin resistance and fatty liver. Mice lacking the glutamate-cysteine ligase modifier subunit gene (Gclm(-/-)) and deficient in glutathione (GSH), are resistant to HFD-mediated weight gain. Herein, we evaluated Gclm-associated regulation of energy metabolism, oxidative stress, and glucose and lipid homeostasis. C57BL/6J Gclm(-/-) mice and littermate wild-type (WT) controls received a normal diet or an HFD for 11 weeks. HFD-fed Gclm(-/-) mice did not display a decreased respiratory quotient, suggesting that they are unable to process lipid for metabolism. Although dietary energy consumption and intestinal lipid absorption were unchanged in Gclm(-/-) mice, feeding these mice an HFD did not produce excess body weight nor fat storage. Gclm(-/-) mice displayed higher basal metabolic rates resulting from higher activities of liver mitochondrial NADH-CoQ oxidoreductase, thus elevating respiration. Although Gclm(-/-) mice exhibited strong systemic and hepatic oxidative stress responses, HFD did not promote glucose intolerance or insulin resistance. Furthermore, HFD-fed Gclm(-/-) mice did not develop fatty liver, likely resulting from very low expression levels of genes encoding lipid metabolizing enzymes. We conclude that Gclm is involved in the regulation of basal metabolic rate and the metabolism of dietary lipid. Although Gclm(-/-) mice display a strong oxidative stress response, they are protected from HFD-induced excessive weight gain and adipose deposition, insulin resistance and steatosis. -- Highlights: Black-Right-Pointing-Pointer A high fat diet does not produce body weight and fat gain in Gclm(-/-) mice. Black-Right-Pointing-Pointer A high fat diet does not induce steatosis or insulin resistance in Gclm(-/-) mice. Black-Right-Pointing-Pointer Gclm(-/-) mice have high basal metabolism and mitochondrial

Immunobiology of congenitally athymic-asplenic mice

International Nuclear Information System (INIS)

Gershwin, M.E.; Ahmed, A.; Ikeda, R.M.; Shifrine, M.; Wilson, F.

1978-01-01

A study has been made of congenitally athymic-asplenic mice obtained by the mating of nude by hereditarily asplenic (Dh/+) mice. The mice survived for up to 9 months, under specific pathogen-free conditions, with no evidence for increased risk of spontaneous neoplasia. Although lymphocyte surface markers and sera immunoglobulin levels of athymic-asplenic mice were similar to those of their nude and asplenic littermates, there were a number of major immunologic differences. The athymic-asplenic mice appeared more immunologically compromised than nude mice. There was an elevated rate of growth and a lower inoculated cell threshold needed for successful transplantation of a human malignant melanoma. There was no evidence for auto-antibody production in mice up to 9 months of age. Congenitally athymic-asplenic mice can be used for a variety of studies in which other immunologically deprived mouse mutants are desired. (author)

The Cstf2t Polyadenylation Gene Plays a Sex-Specific Role in Learning Behaviors in Mice.

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Jaryse C Harris

Full Text Available Polyadenylation is an essential mechanism for the processing of mRNA 3' ends. CstF-64 (the 64,000 Mr subunit of the cleavage stimulation factor; gene symbol Cstf2 is an RNA-binding protein that regulates mRNA polyadenylation site usage. We discovered a paralogous form of CstF-64 called τCstF-64 (Cstf2t. The Cstf2t gene is conserved in all eutherian mammals including mice and humans, but the τCstF-64 protein is expressed only in a subset of mammalian tissues, mostly testis and brain. Male mice that lack Cstf2t (Cstf2t-/- mice experience disruption of spermatogenesis and are infertile, although female fertility is unaffected. However, a role for τCstF-64 in the brain has not yet been determined. Given the importance of RNA polyadenylation and splicing in neuronal gene expression, we chose to test the hypothesis that τCstF-64 is important for brain function. Male and female 185-day old wild type and Cstf2t-/- mice were examined for motor function, general activity, learning, and memory using rotarod, open field activity, 8-arm radial arm maze, and Morris water maze tasks. Male wild type and Cstf2t-/- mice did not show differences in learning and memory. However, female Cstf2t-/- mice showed significantly better retention of learned maze tasks than did female wild type mice. These results suggest that τCstf-64 is important in memory function in female mice. Interestingly, male Cstf2t-/- mice displayed less thigmotactic behavior than did wild type mice, suggesting that Cstf2t may play a role in anxiety in males. Taken together, our studies highlight the importance of mRNA processing in cognition and behavior as well as their established functions in reproduction.

Enhanced expression of Nrf2 in mice attenuates the fatty liver produced by a methionine- and choline-deficient diet

International Nuclear Information System (INIS)

Zhang, Yu-Kun Jennifer; Yeager, Ronnie L.; Tanaka, Yuji; Klaassen, Curtis D.

2010-01-01

Oxidative stress has been proposed as an important promoter of the progression of fatty liver diseases. The current study investigates the potential functions of the Nrf2-Keap1 signaling pathway, an important hepatic oxidative stress sensor, in a rodent fatty liver model. Mice with no (Nrf2-null), normal (wild type, WT), and enhanced (Keap1 knockdown, K1-kd) expression of Nrf2 were fed a methionine- and choline-deficient (MCD) diet or a control diet for 5 days. Compared to WT mice, the MCD diet-caused hepatosteatosis was more severe in the Nrf2-null mice and less in the K1-kd mice. The Nrf2-null mice had lower hepatic glutathione and exhibited more lipid peroxidation, whereas the K1-kd mice had the highest amount of glutathione in the liver and developed the least lipid peroxidation among the three genotypes fed the MCD diet. The Nrf2 signaling pathway was activated by the MCD diet, and the Nrf2-targeted cytoprotective genes Nqo1 and Gstα1/2 were induced in WT and even more in K1-kd mice. In addition, Nrf2-null mice on both control and MCD diets exhibited altered expression profiles of fatty acid metabolism genes, indicating Nrf2 may influence lipid metabolism in liver. For example, mRNA levels of long chain fatty acid translocase CD36 and the endocrine hormone Fgf21 were higher in livers of Nrf2-null mice and lower in the K1-kd mice than WT mice fed the MCD diet. Taken together, these observations indicate that Nrf2 could decelerate the onset of fatty livers caused by the MCD diet by increasing hepatic antioxidant and detoxification capabilities.

Increased infectivity of anchorless mouse scrapie prions in transgenic mice overexpressing human prion protein.

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Race, Brent; Phillips, Katie; Meade-White, Kimberly; Striebel, James; Chesebro, Bruce

2015-06-01

Prion protein (PrP) is found in all mammals, mostly as a glycoprotein anchored to the plasma membrane by a C-terminal glycosylphosphatidylinositol (GPI) linkage. Following prion infection, host protease-sensitive prion protein (PrPsen or PrPC) is converted into an abnormal, disease-associated, protease-resistant form (PrPres). Biochemical characteristics, such as the PrP amino acid sequence, and posttranslational modifications, such as glycosylation and GPI anchoring, can affect the transmissibility of prions as well as the biochemical properties of the PrPres generated. Previous in vivo studies on the effects of GPI anchoring on prion infectivity have not examined cross-species transmission. In this study, we tested the effect of lack of GPI anchoring on a species barrier model using mice expressing human PrP. In this model, anchorless 22L prions derived from tg44 mice were more infectious than 22L prions derived from C57BL/10 mice when tested in tg66 transgenic mice, which expressed wild-type anchored human PrP at 8- to 16-fold above normal. Thus, the lack of the GPI anchor on the PrPres from tg44 mice appeared to reduce the effect of the mouse-human PrP species barrier. In contrast, neither source of prions induced disease in tgRM transgenic mice, which expressed human PrP at 2- to 4-fold above normal. Prion protein (PrP) is found in all mammals, usually attached to cells by an anchor molecule called GPI. Following prion infection, PrP is converted into a disease-associated form (PrPres). While most prion diseases are species specific, this finding is not consistent, and species barriers differ in strength. The amino acid sequence of PrP varies among species, and this variability affects prion species barriers. However, other PrP modifications, including glycosylation and GPI anchoring, may also influence cross-species infectivity. We studied the effect of PrP GPI anchoring using a mouse-to-human species barrier model. Experiments showed that prions produced by

Cyclic GMP-AMP displays mucosal adjuvant activity in mice.

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Ivana Škrnjug

Full Text Available The recently discovered mammalian enzyme cyclic GMP-AMP synthase produces cyclic GMP-AMP (cGAMP after being activated by pathogen-derived cytosolic double stranded DNA. The product can stimulate STING-dependent interferon type I signaling. Here, we explore the efficacy of cGAMP as a mucosal adjuvant in mice. We show that cGAMP can enhance the adaptive immune response to the model antigen ovalbumin. It promotes antigen specific IgG and a balanced Th1/Th2 lymphocyte response in immunized mice. A characteristic of the cGAMP-induced immune response is the slightly reduced induction of interleukin-17 as a hallmark of Th17 activity--a distinct feature that is not observed with other cyclic di-nucleotide adjuvants. We further characterize the innate immune stimulation activity in vitro on murine bone marrow-derived dendritic cells and human dendritic cells. The observed results suggest the consideration of cGAMP as a candidate mucosal adjuvant for human vaccines.

Cyclic GMP-AMP displays mucosal adjuvant activity in mice.

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Škrnjug, Ivana; Guzmán, Carlos Alberto; Rueckert, Christine; Ruecker, Christine

2014-01-01

The recently discovered mammalian enzyme cyclic GMP-AMP synthase produces cyclic GMP-AMP (cGAMP) after being activated by pathogen-derived cytosolic double stranded DNA. The product can stimulate STING-dependent interferon type I signaling. Here, we explore the efficacy of cGAMP as a mucosal adjuvant in mice. We show that cGAMP can enhance the adaptive immune response to the model antigen ovalbumin. It promotes antigen specific IgG and a balanced Th1/Th2 lymphocyte response in immunized mice. A characteristic of the cGAMP-induced immune response is the slightly reduced induction of interleukin-17 as a hallmark of Th17 activity--a distinct feature that is not observed with other cyclic di-nucleotide adjuvants. We further characterize the innate immune stimulation activity in vitro on murine bone marrow-derived dendritic cells and human dendritic cells. The observed results suggest the consideration of cGAMP as a candidate mucosal adjuvant for human vaccines.

Chronic Giardia muris infection in anti-IgM-treated mice. I. Analysis of immunoglobulin and parasite-specific antibody in normal and immunoglobulin-deficient animals.

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Snider, D P; Gordon, J; McDermott, M R; Underdown, B J

1985-06-01

To investigate the role of B cells and antibody in the immune response of mice to the murine intestinal parasite Giardia muris, we used mice treated from birth with rabbit anti-IgM antisera (aIgM). Such mice developed in serum and in gut secretions extreme Ig deficiency (IgM, IgA, and IgG) relative to control animals. The aIgM-treated mice showed no anti-G. muris antibody in serum or in gut wash material. Infections of G. muris in these mice were chronic, with a high load of parasite present in the small bowel, as reflected by prolonged cyst excretion (greater than 11 wk) and high trophozoite counts. In contrast, normal, untreated mice or NRS-treated animals developed anti-parasite IgA and IgG antibody in serum, demonstrated IgA antibody against the parasite in gut washings, and expelled the parasite within 9 wk. These effects of aIgM treatment on the murine response to primary infection with G. muris were demonstrated in two strains of mice: BALB/c and (C57BL/6 X C3H/He) F1. It was also observed that the response to G. muris infection in untreated animals was characterized by higher than normal total secretion of IgA into the gut and a concomitant increase in the serum polymeric IgA level. Mice treated with aIgM had a marked decrease of both monomeric and polymeric IgA in serum, and little detectable IgA in the intestinal lumen. These experiments provide the first demonstration that anti-IgM treatment suppresses a specific intestinal antibody response to antigen, and provide evidence that B cells and antibody play a role in the development of an effective response to a primary infection with G. muris in mice.

Health surveillance of specific pathogen-free and conventionally-housed mice and rats in Korea.

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Seok, Seunghyeok; Park, Jonghwan; Cho, Suna; Baek, Minwon; Lee, Huiyoung; Kim, Dongjae; Yang, Kihwa; Jang, Dongdeuk; Han, Beomseok; Nam, Kitaek; Park, Jaehak

2005-01-01

The present study contains information about proper microbiological monitoring of laboratory animals' health and the standardization of microbiological monitoring methods in Korea. Microbiological quality control for laboratory animals, composed of biosecurity and health surveillance, is essential to guard against research complications and public health dangers that have been associated with adventitious infections. In this study, one hundred and twenty-two mice and ninety rats from laboratory animal breeding companies and one animal facility of the national universities in Korea were monitored in 2000-2003. Histopathologically, thickening of the alveolar walls and lymphocytic infiltration around the bronchioles were observed in mice and rats from microbiologically contaminated facilities. Cryptosporidial oocysts were observed in the gastric pits of only conventionally-housed mice and rats. Helicobacter spp. infection was also detected in 1 of 24 feces DNA samples in mice and 9 of 40 feces DNA samples in rats by PCR in 2003, but they were not Helicobacter hepaticus. This paper describes bacteriological, parasitological, and virological examinations of the animals.

Subcutaneous or oral immunization of mice with Lactococcus lactis expressing F4 fimbrial adhesin FaeG.

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Liu, Shujie; Li, Yongming; Xu, Ziwei; Wang, Yicheng

2013-01-01

Enterotoxigenic Escherichia coli (ETEC) is one of the most common causes of diarrhea in neonatal and postweaning piglets. Fimbrial adhesion of ETEC has been considered an important colonization factor with antigenicity. To safely and effectively deliver the F4 (K88) fimbrial adhesin FaeG to the immune system, we have previously constructed the secretory expression vector pNZ8112-faeG, and FaeG was produced in cytoplasmic form in Lactococcus lactis. In this work, BALB/c mice were immunized with recombinant L. lactis to further determine the immunogenicity of recombinant FaeG (rFaeG) via the subcutaneous or oral route. Subcutaneous immunization in mice with recombinant L. lactis induced a significant increase in the F4-specific serum IgG titer and the number of antibody-secreting cells (ASCs) in the spleen. Oral immunization of mice with recombinant L. lactis induced mucosal and systemic F4-specific immune responses and increased the number of ASCs in the spleen, mesenteric lymph nodes and Peyer's patches. High-dose (2.8 × 10(11) CFU) recombinant strains and adjuvant cholera toxin B subunit enhanced specific mucosal immune responses. The results suggest the feasibility of delivering rFaeG expressed in L. lactis to the immune system in order to induce an F4-specific immune response.

Bacterially produced recombinant influenza vaccines based on virus-like particles.

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Andrea Jegerlehner

Full Text Available Although current influenza vaccines are effective in general, there is an urgent need for the development of new technologies to improve vaccine production timelines, capacities and immunogenicity. Herein, we describe the development of an influenza vaccine technology which enables recombinant production of highly efficient influenza vaccines in bacterial expression systems. The globular head domain of influenza hemagglutinin, comprising most of the protein's neutralizing epitopes, was expressed in E. coli and covalently conjugated to bacteriophage-derived virus-like particles produced independently in E.coli. Conjugate influenza vaccines produced this way were used to immunize mice and found to elicit immune sera with high antibody titers specific for the native influenza hemagglutinin protein and high hemagglutination-inhibition titers. Moreover vaccination with these vaccines induced full protection against lethal challenges with homologous and highly drifted influenza strains.

Free energy landscape and cooperatively rearranging region in a hard sphere glass

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Yoshidome, Takashi; Yoshimori, Akira; Odagaki, Takashi

2007-08-01

Exploiting the density functional theory, we calculate the free energy landscape (FEL) of the hard sphere glass in three dimensions. From the FEL, we estimate the number of the particles in the cooperatively rearranging region (CRR). We find that the density dependence of the number of the particles in the CRR is expressed as a power law function of the density. Analyzing the relaxation process in the CRR, we also find that the string motion is the elementary process for the structural relaxation, which leads to the natural definition of the simultaneously rearranging region as the particles displaced in the string motion.

Sex-Specific Life Course Changes in the Neuro-Metabolic Phenotype of Glut3 Null Heterozygous Mice: Ketogenic Diet Ameliorates Electroencephalographic Seizures and Improves Sociability.

Science.gov (United States)

Dai, Yun; Zhao, Yuanzi; Tomi, Masatoshi; Shin, Bo-Chul; Thamotharan, Shanthie; Mazarati, Andrey; Sankar, Raman; Wang, Elizabeth A; Cepeda, Carlos; Levine, Michael S; Zhang, Jingjing; Frew, Andrew; Alger, Jeffry R; Clark, Peter M; Sondhi, Monica; Kositamongkol, Sudatip; Leibovitch, Leah; Devaskar, Sherin U

2017-04-01

We tested the hypothesis that exposure of glut3+/- mice to a ketogenic diet ameliorates autism-like features, which include aberrant behavior and electrographic seizures. We first investigated the life course sex-specific changes in basal plasma-cerebrospinal fluid (CSF)-brain metabolic profile, brain glucose transport/uptake, glucose and monocarboxylate transporter proteins, and adenosine triphosphate (ATP) in the presence or absence of systemic insulin administration. Glut3+/- male but not female mice (5 months of age) displayed reduced CSF glucose/lactate concentrations with no change in brain Glut1, Mct2, glucose uptake or ATP. Exogenous insulin-induced hypoglycemia increased brain glucose uptake in glut3+/- males alone. Higher plasma-CSF ketones (β-hydroxybutyrate) and lower brain Glut3 in females vs males proved protective in the former while enhancing vulnerability in the latter. As a consequence, increased synaptic proteins (neuroligin4 and SAPAP1) with spontaneous excitatory postsynaptic activity subsequently reduced hippocampal glucose content and increased brain amyloid β1-40 deposition in an age-dependent manner in glut3+/- males but not females (4 to 24 months of age). We then explored the protective effect of a ketogenic diet on ultrasonic vocalization, sociability, spatial learning and memory, and electroencephalogram seizures in male mice (7 days to 6 to 8 months of age) alone. A ketogenic diet partially restored sociability without affecting perturbed vocalization, spatial learning and memory, and reduced seizure events. We conclude that (1) sex-specific and age-dependent perturbations underlie the phenotype of glut3+/- mice, and (2) a ketogenic diet ameliorates seizures caused by increased cortical excitation and improves sociability, but fails to rescue vocalization and cognitive deficits in glut3+/- male mice. Copyright © 2017 Endocrine Society.

Generation of Recombinant Monoclonal Antibodies from Immunised Mice and Rabbits via Flow Cytometry and Sorting of Antigen-Specific IgG+ Memory B Cells.

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Dale O Starkie

Full Text Available Single B cell screening strategies, which avoid both hybridoma fusion and combinatorial display, have emerged as important technologies for efficiently sampling the natural antibody repertoire of immunized animals and humans. Having access to a range of methods to interrogate different B cell subsets provides an attractive option to ensure large and diverse panels of high quality antibody are produced. The generation of multiple antibodies and having the ability to find rare B cell clones producing IgG with unique and desirable characteristics facilitates the identification of fit-for-purpose molecules that can be developed into therapeutic agents or research reagents. Here, we describe a multi-parameter flow cytometry single-cell sorting technique for the generation of antigen-specific recombinant monoclonal antibodies from single IgG+ memory B cells. Both mouse splenocytes and rabbit PBMC from immunised animals were used as a source of B cells. Reagents staining both B cells and other unwanted cell types enabled efficient identification of class-switched IgG+ memory B cells. Concurrent staining with antigen labelled separately with two spectrally-distinct fluorophores enabled antigen-specific B cells to be identified, i.e. those which bind to both antigen conjugates (double-positive. These cells were then typically sorted at one cell per well using FACS directly into a 96-well plate containing reverse transcriptase reaction mix. Following production of cDNA, PCR was performed to amplify cognate heavy and light chain variable region genes and generate transcriptionally-active PCR (TAP fragments. These linear expression cassettes were then used directly in a mammalian cell transfection to generate recombinant antibody for further testing. We were able to successfully generate antigen-specific recombinant antibodies from both the rabbit and mouse IgG+ memory B cell subset within one week. This included the generation of an anti-TNFR2 blocking

Department of Homeland Security Cyber Resilience Review (Case Study)

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2014-01-23

operational stress and c-Ues. TheCRRseek:stoelidtthecurrentstateof cyber seruritymanagementpracticesfromkeyc.yber .securitypersonnel...Institutionalization in the CRR Processes  are   acculturated ,   defined,   measured,   and   governed   Maturity indictor levels (MIL) are used in...processes that •  produce consistent results over time •  are retained during times of stress Level 0-Incomplete Level 1-Performed Level 2

Cyp26b1 within the growth plate regulates bone growth in juvenile mice

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Minegishi, Yoshiki [Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Department of Plastic and Reconstructive Surgery, University of Fukui Hospital, 23-3 Matsuokashimoaizuki, Eiheiji-cho, Yoshida-gun, Fukui 910-1193 (Japan); Department of Plastic Surgery, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871 (Japan); Sakai, Yasuo [Department of Plastic Surgery, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871 (Japan); Department of Plastic Surgery, Bellland General Hospital, 500-3 Higashiyama Naka-ku, Sakai, Osaka 599-8247 (Japan); Yahara, Yasuhito [Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Akiyama, Haruhiko [Department of Orthopaedic Surgery, Gifu University Graduate School of Medicine, 1-1 Yanagito, Gifu 501-1194 (Japan); Yoshikawa, Hideki [Department of Orthopaedic Surgery, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871 (Japan); Hosokawa, Ko [Department of Plastic Surgery, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871 (Japan); Tsumaki, Noriyuki, E-mail: ntsumaki@cira.kyoto-u.ac.jp [Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507 (Japan); Japan Science and Technology Agency, CREST, Tokyo 102-0075 (Japan)

2014-11-07

Highlights: • Retinoic acid and Cyp26b1 were oppositely localized in growth plate cartilage. • Cyp26b1 deletion in chondrocytes decreased bone growth in juvenile mice. • Cyp26b1 deletion reduced chondrocyte proliferation and growth plate height. • Vitamin A-depletion partially reversed growth plate abnormalities caused by Cyp26b1 deficiency. • Cyp26b1 regulates bone growth by controlling chondrocyte proliferation. - Abstract: Retinoic acid (RA) is an active metabolite of vitamin A and plays important roles in embryonic development. CYP26 enzymes degrade RA and have specific expression patterns that produce a RA gradient, which regulates the patterning of various structures in the embryo. However, it has not been addressed whether a RA gradient also exists and functions in organs after birth. We found localized RA activities in the diaphyseal portion of the growth plate cartilage were associated with the specific expression of Cyp26b1 in the epiphyseal portion in juvenile mice. To disturb the distribution of RA, we generated mice lacking Cyp26b1 specifically in chondrocytes (Cyp26b1{sup Δchon} cKO). These mice showed reduced skeletal growth in the juvenile stage. Additionally, their growth plate cartilage showed decreased proliferation rates of proliferative chondrocytes, which was associated with a reduced height in the zone of proliferative chondrocytes, and closed focally by four weeks of age, while wild-type mouse growth plates never closed. Feeding the Cyp26b1 cKO mice a vitamin A-deficient diet partially reversed these abnormalities of the growth plate cartilage. These results collectively suggest that Cyp26b1 in the growth plate regulates the proliferation rates of chondrocytes and is responsible for the normal function of the growth plate and growing bones in juvenile mice, probably by limiting the RA distribution in the growth plate proliferating zone.

Liver regeneration in mice bearing a transplanted hepatoma.

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Badran, A F; Moreno, F R; Echave Llanos, J M

1984-01-01

The hepatocyte mitotic index curve in hepatectomized hepatoma-bearing mice, rises earlier, has a greater amplitude and is less synchronized than that of normal hepatectomized mice. This indicates a stimulation (more mitosis in a shorter time period) produced by the presence of the tumors. The sinusoid litoral cells mitotic index curve in hepatectomized hepatoma-bearing mice appears earlier and is much less synchronized than that of normal hepatectomized mice. Nevertheless both curves have the same amplitude for the whole sampling period and the early stimulation is quickly compensated by lower values (apparent inhibition) appearing in the resting (light) period.

SPECT imaging of D{sub 2} dopamine receptors and endogenous dopamine release in mice

Energy Technology Data Exchange (ETDEWEB)

Jongen, Cynthia [University Medical Center Utrecht, Image Sciences Institute, Q0S.459, P.O. Box 85500, Utrecht (Netherlands); Bruin, Kora de; Booij, Jan [University of Amsterdam, Academic Medical Center, Department of Nuclear Medicine, Amsterdam (Netherlands); Beekman, Freek [University Medical Center Utrecht, Image Sciences Institute, Q0S.459, P.O. Box 85500, Utrecht (Netherlands); University Medical Center Utrecht, Department of Neuroscience and Pharmacology, Utrecht (Netherlands); Technical University Delft, Department R3, Section Radiation, Detection and Matter, Delft (Netherlands)

2008-09-15

The dopamine D{sub 2} receptor (D2R) is important in the mediation of addiction. [{sup 123}I]iodobenzamide (IBZM), a SPECT ligand for the D2R, has been used for in vivo studies of D2R availability in humans, monkeys, and rats. Although mouse models are important in the study of addiction, [{sup 123}I]IBZM has not been used in mice SPECT studies. This study evaluates the use of [{sup 123}I]IBZM for measuring D2R availability in mice. Pharmacokinetics of [{sup 123}I]IBZM in mice were studied with pinhole SPECT imaging after intravenous (i.v.) injection of [{sup 123}I]IBZM (20, 40, and 70 MBq). In addition, the ability to measure the release of endogenous dopamine after amphetamine administration with [{sup 123}I]IBZM SPECT was investigated. Thirdly, i.v. administration, the standard route of administration, and intraperitoneal (i.p.) administration of [{sup 123}I]IBZM were compared. Specific binding of [{sup 123}I]IBZM within the mouse striatum could be clearly visualized with SPECT. Peak specific striatal binding ratios were reached around 90 min post-injection. After amphetamine administration, the specific binding ratios of [{sup 123}I]IBZM decreased significantly (-27.2%; n=6; p=0.046). Intravenous administration of [{sup 123}I]IBZM led to significantly higher specific binding than i.p. administration of the same dose. However, we found that i.v. administration of a dose of 70 MBq [{sup 123}I]IBZM might result in acute ethanol intoxication because ethanol is used as a preparative aid for the routine production of [{sup 123}I]IBZM. Imaging of D2R availability and endogenous dopamine release in mice is feasible using [{sup 123}I]IBZM single pinhole SPECT. Using commercially produced [{sup 123}I]IBZM, a dose of 40 MBq injected i.v. can be recommended. (orig.)

Teratogenic and embryolethal effects in mice of fission-spectrum neutrons and γ-rays

International Nuclear Information System (INIS)

Cairnie, A.B.; Grahn, D.; Rayburn, H.B.; Williamson, F.S.; Brown, R.J.

1974-01-01

Fission-spectrum neutrons from the Janus reactor at Argonne National Laboratory were compared with γ-rays in terms of their relative biological effectiveness (RBE) for embryolethal and teratogenic effects in mice. No evidence was found of any processes that were abnormally sensitive to neutrons. The RBE for killing embryos and producing abnormal embryos or specific abnormalities was between 2 and 3. This is close to the values found in other systems for processes involving cell killing. (U.S.)

Growth hormone modulates hypothalamic inflammation in long-lived pituitary dwarf mice.

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Sadagurski, Marianna; Landeryou, Taylor; Cady, Gillian; Kopchick, John J; List, Edward O; Berryman, Darlene E; Bartke, Andrzej; Miller, Richard A

2015-12-01

Mice in which the genes for growth hormone (GH) or GH receptor (GHR(-/-) ) are disrupted from conception are dwarfs, possess low levels of IGF-1 and insulin, have low rates of cancer and diabetes, and are extremely long-lived. Median longevity is also increased in mice with deletion of hypothalamic GH-releasing hormone (GHRH), which leads to isolated GH deficiency. The remarkable extension of longevity in hypopituitary Ames dwarf mice can be reversed by a 6-week course of GH injections started at the age of 2 weeks. Here, we demonstrate that mutations that interfere with GH production or response, in the Snell dwarf, Ames dwarf, or GHR(-/-) mice lead to reduced formation of both orexigenic agouti-related peptide (AgRP) and anorexigenic proopiomelanocortin (POMC) projections to the main hypothalamic projection areas: the arcuate nucleus (ARH), paraventricular nucleus (PVH), and dorsomedial nucleus (DMH). These mutations also reduce hypothalamic inflammation in 18-month-old mice. GH injections, between 2 and 8 weeks of age, reversed both effects in Ames dwarf mice. Disruption of GHR specifically in liver (LiGHRKO), a mutation that reduces circulating IGF-1 but does not lead to lifespan extension, had no effect on hypothalamic projections or inflammation, suggesting an effect of GH, rather than peripheral IGF-1, on hypothalamic development. Hypothalamic leptin signaling, as monitored by induction of pStat3, is not impaired by GHR deficiency. Together, these results suggest that early-life disruption of GH signaling produces long-term hypothalamic changes that may contribute to the longevity of GH-deficient and GH-resistant mice. © 2015 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

Effects of repeated treatment with MDMA on working memory and behavioural flexibility in mice.

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Viñals, Xavier; Maldonado, Rafael; Robledo, Patricia

2013-03-01

Repeated administration of 3,4-methylenedioxymethamphetamine (MDMA) produces dopaminergic neurotoxicity in mice. However, it is still not clear whether this exposure induces deficits in cognitive processing related to specific subsets of executive functioning. We evaluated the effects of neurotoxic and non-neurotoxic doses of MDMA (0, 3 and 30 mg/kg, twice daily for 4 days) on working memory and attentional set-shifting in mice, and changes in extracellular levels of dopamine (DA) in the striatum. Treatment with MDMA (30 mg/kg) disrupted performance of acquired operant alternation, and this impairment was still apparent 5 days after the last drug administration. Decreased alternation was not related to anhedonia because no differences were observed between groups in the saccharin preference test under similar experimental conditions. Correct responding on delayed alternation was increased 1 day after repeated treatment with MDMA (30 mg/kg), probably because of general behavioural quiescence. Notably, the high dose regimen of MDMA impaired attentional set-shifting related to an increase in total perseveration errors. Finally, basal extracellular levels of DA in the striatum were not modified in mice repeatedly treated with MDMA with respect to controls. However, an acute challenge with MDMA (10 mg/kg) failed to increase DA outflow in mice receiving the highest MDMA dose (30 mg/kg), corroborating a decrease in the functionality of DA transporters. Seven days after this treatment, the effects of MDMA on DA outflow were recovered. These results suggest that repeated neurotoxic doses of MDMA produce lasting impairments in recall of alternation behaviour and reduce cognitive flexibility in mice. © 2012 The Authors, Addiction Biology © 2012 Society for the Study of Addiction.

Reversal of hyperglycemia in mice by using human expandable insulin-producing cells differentiated from fetal liver progenitor cells

Science.gov (United States)

Zalzman, Michal; Gupta, Sanjeev; Giri, Ranjit K.; Berkovich, Irina; Sappal, Baljit S.; Karnieli, Ohad; Zern, Mark A.; Fleischer, Norman; Efrat, Shimon

2003-06-01

Beta-cell replacement is considered to be the most promising approach for treatment of type 1 diabetes. Its application on a large scale is hindered by a shortage of cells for transplantation. Activation of insulin expression, storage, and regulated secretion in stem/progenitor cells offers novel ways to overcome this shortage. We explored whether fetal human progenitor liver cells (FH) could be induced to differentiate into insulin-producing cells after expression of the pancreatic duodenal homeobox 1 (Pdx1) gene, which is a key regulator of pancreatic development and insulin expression in beta cells. FH cells possess a considerable replication capacity, and this was further extended by introduction of the gene for the catalytic subunit of human telomerase. Immortalized FH cells expressing Pdx1 activated multiple beta-cell genes, produced and stored considerable amounts of insulin, and released insulin in a regulated manner in response to glucose. When transplanted into hyperglycemic immunodeficient mice, the cells restored and maintained euglycemia for prolonged periods. Quantitation of human C-peptide in the mouse serum confirmed that the glycemia was normalized by the transplanted human cells. This approach offers the potential of a novel source of cells for transplantation into patients with type 1 diabetes.

Exposure to chronic variable social stress during adolescence alters affect-related behaviors and adrenocortical activity in adult male and female inbred mice.

Science.gov (United States)

Caruso, Michael J; Kamens, Helen M; Cavigelli, Sonia A

2017-09-01

Rodent models provide valuable insight into mechanisms that underlie vulnerability to adverse effects of early-life challenges. Few studies have evaluated sex differences in anxiogenic or depressogenic effects of adolescent social stress in a rodent model. Furthermore, adolescent stress studies often use genetically heterogeneous outbred rodents which can lead to variable results. The current study evaluated the effects of adolescent social stress in male and female inbred (BALB/cJ) mice. Adolescent mice were exposed to repeat cycles of alternating social isolation and social novelty for 4 weeks. Adolescent social stress increased anxiety-related behaviors in both sexes and depression-related behavior in females. Locomotion/exploratory behavior was also decreased in both sexes by stress. Previously stressed adult mice produced less basal fecal corticosteroids than controls. Overall, the novel protocol induced sex-specific changes in anxiety- and depression-related behaviors and corticoid production in inbred mice. The chronic variable social stress protocol used here may be beneficial to systematically investigate sex-specific neurobiological mechanisms underlying adolescent stress vulnerability where genetic background can be controlled. © 2017 Wiley Periodicals, Inc.

Epidermal dysplasia and abnormal hair follicles in transgenic mice overexpressing homeobox gene MSX-2.

Science.gov (United States)

Jiang, T X; Liu, Y H; Widelitz, R B; Kundu, R K; Maxson, R E; Chuong, C M

1999-08-01

The homeobox gene Msx-2 is expressed specifically in sites of skin appendage formation. To explore its part in skin morphogenesis, we produced transgenic mice expressing Msx-2 under the control of the cytomegalovirus promoter. The skin of these transgenic mice was flaky, exhibiting desquamation and shorter hairs. Histologic analysis showed thickened epidermis with hyperproliferation, which was restricted to the basal layer. Hyperkeratosis was also evident. A wide zone of suprabasal cells were misaligned and coexpressed keratins 14 and 10. There was reduced expression of integrin beta 1 and DCC in the basal layer. Hair follicles were misaligned with a shrunken matrix region. The dermis showed increased cellularity and empty vacuoles. We suggest that Msx-2 is involved in the growth control of skin and skin appendages.

Cell type-specific deficiency of c-kit gene expression in mutant mice of mi/mi genotype.

Science.gov (United States)

Isozaki, K.; Tsujimura, T.; Nomura, S.; Morii, E.; Koshimizu, U.; Nishimune, Y.; Kitamura, Y.

1994-01-01

The mi locus of mice encodes a novel member of the basic-helix-loop-helix-leucine zipper protein family of transcription factors (hereafter called mi factor). In addition to microphthalmus, osteopetrosis, and lack of melanocytes, mice of mi/mi genotype are deficient in mast cells. Since the c-kit receptor tyrosine kinase plays an important role in the development of mast cells, and since the c-kit expression by cultured mast cells from mi/mi mice is deficient in both mRNA and protein levels, the mast cell deficiency of mi/mi mice has been attributed at least in part to the deficient expression of c-kit. However, it remained to be examined whether the c-kit expression was also deficient in tissues of mi/mi mice. In the present study, we examined the c-kit expression by mi/mi skin mast cells using in situ hybridization and immunohistochemistry. Moreover, we examined the c-kit expression by various cells other than mast cells in tissues of mi/mi mice. We found that the c-kit expression was deficient in mast cells but not in erythroid precursors, testicular germ cells, and neurons of mi/mi mice. This suggested that the regulation of the c-kit transcription by the mi factor was dependent on cell types. Mice of mi/mi genotype appeared to be a useful model to analyze the function of transcription factors in the whole-animal level. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7524330

The transcriptional repressor Zbtb20 is essential for specification of hippocampal projection neurons and territory in mice

DEFF Research Database (Denmark)

Rosenthal, Eva Helga

for specification of both hippocampal pyramidal neurons and territory in a mouse knockout model. Homozygous Zbtb20-/- mice are viable at birth, but display dwarfism and die during the first month of postnatal life. Characterization of the Zbtb20-/- brain phenotype reveals a small vestigial hippocampus...... with a dramatic change in the molecular patterning of the subiculum and Ammon’s horn. In absence of Zbtb20, the pattern of expression of distinct molecular markers was altered at four borders: retrosplenial cortex/subiculum, subiculum/CA1, CA1/CA2, and CA2/CA3, leading to a replacement of Ammon’s horn...

Expert system to produce a welding procedure specification

International Nuclear Information System (INIS)

Fukuda, S.; Morita, H.; Yamauchi, Y.; Tsuji, S.

1993-01-01

This paper describes the outline of the expert system for producing a welding procedure specification for a pressure vessel which has been developed in the 5 year project of Nagasaki Prefecture financially supported by MITI (Ministry of International Trade and Industry). The aim of this project is to reduce the burden of welding engineers because requirements are betting more and more severe and products are getting more and more complicated and diversified. In order to reduce the burden of a welding engineer, the Project has shown that the following facts must be taken into account in addition to the usual procedures of developing an expert system. The inference must be processed in accordance with the lines of reasoning of a welding engineer. It must be remembered that these lines differ from engineer to engineer. Thus, a very flexible inference mechanism is called for. The constraints are propagated in any direction. The mechanism to deal with this kind of constraint propagation is called for. The user wishes not only to follow the lines of reasoning but also to get the whole perspective during the reasoning process. The search space must be made visible to the user. Thus, the range must be clearly shown in the spreadsheet and in the graphics. And their correspondence must be established. Hierarchical inference mechanism is preferred

Novel chimeric virus-like particles vaccine displaying MERS-CoV receptor-binding domain induce specific humoral and cellular immune response in mice.

Science.gov (United States)

Wang, Chong; Zheng, Xuexing; Gai, Weiwei; Wong, Gary; Wang, Hualei; Jin, Hongli; Feng, Na; Zhao, Yongkun; Zhang, Weijiao; Li, Nan; Zhao, Guoxing; Li, Junfu; Yan, Jinghua; Gao, Yuwei; Hu, Guixue; Yang, Songtao; Xia, Xianzhu

2017-04-01

Middle East respiratory syndrome coronavirus (MERS-CoV) has continued spreading since its emergence in 2012 with a mortality rate of 35.6%, and is a potential pandemic threat. Prophylactics and therapies are urgently needed to address this public health problem. We report here the efficacy of a vaccine consisting of chimeric virus-like particles (VLP) expressing the receptor binding domain (RBD) of MERS-CoV. In this study, a fusion of the canine parvovirus (CPV) VP2 structural protein gene with the RBD of MERS-CoV can self-assemble into chimeric, spherical VLP (sVLP). sVLP retained certain parvovirus characteristics, such as the ability to agglutinate pig erythrocytes, and structural morphology similar to CPV virions. Immunization with sVLP induced RBD-specific humoral and cellular immune responses in mice. sVLP-specific antisera from these animals were able to prevent pseudotyped MERS-CoV entry into susceptible cells, with neutralizing antibody titers reaching 1: 320. IFN-γ, IL-4 and IL-2 secreting cells induced by the RBD were detected in the splenocytes of vaccinated mice by ELISpot. Furthermore, mice inoculated with sVLP or an adjuvanted sVLP vaccine elicited T-helper 1 (Th1) and T-helper 2 (Th2) cell-mediated immunity. Our study demonstrates that sVLP displaying the RBD of MERS-CoV are promising prophylactic candidates against MERS-CoV in a potential outbreak situation. Copyright © 2017 Elsevier B.V. All rights reserved.

Induction of antigen-specific immunity by pH-sensitive carbonate apatite as a potent vaccine carrier

International Nuclear Information System (INIS)

Hebishima, Takehisa; Tada, Seiichi; Takeshima, Shin-nosuke; Akaike, Toshihiro; Ito, Yoshihiro; Aida, Yoko

2011-01-01

Highlights: ► To develop effective vaccine, we examined the effects of CO 3 Ap as an antigen carrier. ► OVA contained in CO 3 Ap was taken up by BMDCs more effectively than free OVA. ► OVA-immunized splenocytes was activated by OVA contained in CO 3 Ap effectively. ► OVA contained in CO 3 Ap induced strong OVA-specific immune responses to C57BL/6 mice. ► CO 3 Ap is promising antigen carrier for the achievement of effective vaccine. -- Abstract: The ability of carbonate apatite (CO 3 Ap) to enhance antigen-specific immunity was examined in vitro and in vivo to investigate its utility as a vaccine carrier. Murine bone marrow-derived dendritic cells took up ovalbumin (OVA) containing CO 3 Ap more effectively than free OVA. Interestingly, mice immunized with OVA-containing CO 3 Ap produced OVA-specific antibodies more effectively than mice immunized with free OVA. Furthermore, immunization of C57BL/6 mice with OVA-containing CO 3 Ap induced the proliferation and antigen-specific production of IFN-γ by splenocytes more strongly than immunization with free OVA. Moreover, no significant differences were detected in the induction of delayed-type hypersensitivity responses, an immune reaction involving an antigen-specific, cell-mediated immune response between OVA-containing CO 3 Ap and OVA-containing alumina salt (Alum), suggesting that CO 3 Ap induced cell-mediated immune response to the same degree as Alum, which is commonly used for clinical applications. This study is the first to demonstrate the induction of antigen-specific immune responses in vivo by CO 3 Ap.

Plasmid DNA initiates replication of yellow fever vaccine in vitro and elicits virus-specific immune response in mice.

Science.gov (United States)

Tretyakova, Irina; Nickols, Brian; Hidajat, Rachmat; Jokinen, Jenny; Lukashevich, Igor S; Pushko, Peter

2014-11-01

Yellow fever (YF) causes an acute hemorrhagic fever disease in tropical Africa and Latin America. To develop a novel experimental YF vaccine, we applied iDNA infectious clone technology. The iDNA represents plasmid that encodes the full-length RNA genome of 17D vaccine downstream from a cytomegalovirus (CMV) promoter. The vaccine was designed to transcribe the full-length viral RNA and to launch 17D vaccine virus in vitro and in vivo. Transfection with 10 ng of iDNA plasmid was sufficient to start replication of vaccine virus in vitro. Safety of the parental 17D and iDNA-derived 17D viruses was confirmed in AG129 mice deficient in receptors for IFN-α/β/γ. Finally, direct vaccination of BALB/c mice with a single 20 μg dose of iDNA plasmid resulted in seroconversion and elicitation of virus-specific neutralizing antibodies in animals. We conclude that iDNA immunization approach combines characteristics of DNA and attenuated vaccines and represents a promising vaccination strategy for YF. Copyright © 2014 Elsevier Inc. All rights reserved.

ß-cell specific overexpression of suppressor of cytokine signalling-3 does not protect against multiple low dose streptozotocin induced type 1 diabetes in mice

DEFF Research Database (Denmark)

Börjesson, A; Rønn, S G; Karlsen, A E

2011-01-01

We investigated the impact of ß-cell specific overexpression of suppressor of cytokine signalling-3 (SOCS-3) on the development of multiple low dose streptozotocin (MLDSTZ) induced Type 1 diabetes and the possible mechanisms involved. MLDSTZ treatment was administered to RIP-SOCS-3 transgenic......RNA in islet cells and secretion of IL-1Ra into culture medium. MLDSTZ treatment caused gradual hyperglycemia both in the wt mice and in the transgenic mice with the latter tending to be more sensitive. In vitro experiments on wt and transgenic islets did not reveal any differences in sensitivity to damaging...

Tolerance induction between two different strains of parental mice prevents graft-versus-host disease in haploidentical hematopoietic stem cell transplantation to F1 mice

International Nuclear Information System (INIS)

Guo, Yixian; Zhang, Lanfang; Wan, Suigui; Sun, Xuejing; Wu, Yongxia; Yu, Xue-Zhong; Xia, Chang-Qing

2014-01-01

Highlights: • Injection of UVB-irradiated iDCs induces alloantigen tolerance. • This alloantigen tolerance may be associated regulatory T cell induction. • Tolerant mice serve as bone marrow donors reduces GVHD to their F1 recipients in allo-HSCT. • Tolerance is maintained in F1 recipients for long time post HSCT. - Abstract: Haploidentical hematopoietic stem cell transplantation (Haplo-HSCT) has been employed worldwide in recent years and led to favorable outcome in a group of patients who do not have human leukocyte antigen (HLA)-matched donors. However, the high incidence of severe graft-versus-host disease (GVHD) is a major problem for Haplo-HSCT. In the current study, we performed a proof of concept mouse study to test whether induction of allogeneic tolerance between two different parental strains was able to attenuate GVHD in Haplo-HSCT to the F1 mice. We induced alloantigen tolerance in C3H mice (H-2k) using ultraviolet B (UVB) irradiated immature dendritic cells (iDCs) derived from the cultures of Balb/c bone marrow cells. Then, we performed Haplo-HSCT using tolerant C3H mice as donors to F1 mice (C3H × Balb/c). The results demonstrated that this approach markedly reduced GVHD-associated death and significantly prolonged the survival of recipient mice in contrast to the groups with donors (C3H mice) that received infusion of non-UVB-irradiated DCs. Further studies showed that there were enhanced Tregs in the tolerant mice and alloantigen-specific T cell response was skewed to more IL-10-producing T cells, suggesting that these regulatory T cells might have contributed to the attenuation of GVHD. This study suggests that it is a feasible approach to preventing GVHD in Haplo-HSCT in children by pre-induction of alloantigen tolerance between the two parents. This concept may also lead to more opportunities in cell-based immunotherapy for GVHD post Haplo-HSCT

Tolerance induction between two different strains of parental mice prevents graft-versus-host disease in haploidentical hematopoietic stem cell transplantation to F1 mice

Energy Technology Data Exchange (ETDEWEB)

Guo, Yixian; Zhang, Lanfang; Wan, Suigui; Sun, Xuejing; Wu, Yongxia [Department of Hematology, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Yu, Xue-Zhong [Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, SC 29425 (United States); Xia, Chang-Qing, E-mail: cqx65@yahoo.com [Department of Hematology, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China)

2014-04-18

Highlights: • Injection of UVB-irradiated iDCs induces alloantigen tolerance. • This alloantigen tolerance may be associated regulatory T cell induction. • Tolerant mice serve as bone marrow donors reduces GVHD to their F1 recipients in allo-HSCT. • Tolerance is maintained in F1 recipients for long time post HSCT. - Abstract: Haploidentical hematopoietic stem cell transplantation (Haplo-HSCT) has been employed worldwide in recent years and led to favorable outcome in a group of patients who do not have human leukocyte antigen (HLA)-matched donors. However, the high incidence of severe graft-versus-host disease (GVHD) is a major problem for Haplo-HSCT. In the current study, we performed a proof of concept mouse study to test whether induction of allogeneic tolerance between two different parental strains was able to attenuate GVHD in Haplo-HSCT to the F1 mice. We induced alloantigen tolerance in C3H mice (H-2k) using ultraviolet B (UVB) irradiated immature dendritic cells (iDCs) derived from the cultures of Balb/c bone marrow cells. Then, we performed Haplo-HSCT using tolerant C3H mice as donors to F1 mice (C3H × Balb/c). The results demonstrated that this approach markedly reduced GVHD-associated death and significantly prolonged the survival of recipient mice in contrast to the groups with donors (C3H mice) that received infusion of non-UVB-irradiated DCs. Further studies showed that there were enhanced Tregs in the tolerant mice and alloantigen-specific T cell response was skewed to more IL-10-producing T cells, suggesting that these regulatory T cells might have contributed to the attenuation of GVHD. This study suggests that it is a feasible approach to preventing GVHD in Haplo-HSCT in children by pre-induction of alloantigen tolerance between the two parents. This concept may also lead to more opportunities in cell-based immunotherapy for GVHD post Haplo-HSCT.

Rhox8 Ablation in the Sertoli Cells Using a Tissue-Specific RNAi Approach Results in Impaired Male Fertility in Mice.

Science.gov (United States)

Welborn, Joshua P; Davis, Matthew G; Ebers, Steven D; Stodden, Genna R; Hayashi, Kanako; Cheatwood, Joseph L; Rao, Manjeet K; MacLean, James A

2015-07-01

The reproductive homeobox X-linked, Rhox, genes encode transcription factors that are selectively expressed in reproductive tissues. While there are 33 Rhox genes in mice, only Rhox and Rhox8 are expressed in Sertoli cells, suggesting that they may regulate the expression of somatic-cell gene products crucial for germ cell development. We previously characterized Rhox5-null mice, which are subfertile, exhibiting excessive germ cell apoptosis and compromised sperm motility. To assess the role of Rhox8 in Sertoli cells, we used a tissue-specific RNAi approach to knockdown RHOX8 in vivo, in which the Rhox5 promoter was used to drive Rhox8-siRNA transgene expression in the postnatal Sertoli cells. Western and immunohistochemical analysis confirmed Sertoli-specific knockdown of RHOX8. However, other Sertoli markers, Gata1 and Rhox5, maintained normal expression patterns, suggesting that the knockdown was specific. Interestingly, male RHOX8-knockdown animals showed significantly reduced spermatogenic output, increased germ cell apoptosis, and compromised sperm motility, leading to impaired fertility. Importantly, our results revealed that while some RHOX5-dependent factors were also misregulated in Sertoli cells of RHOX8-knockdown animals, the majority were not, and novel putative RHOX8-regulated genes were identified. This suggests that while reduction in levels of RHOX5 and RHOX8 in Sertoli cells elicits similar phenotypes, these genes are not entirely redundant. Taken together, our study underscores the importance of Rhox genes in male fertility and suggests that Sertoli cell-specific expression of Rhox5 and Rhox8 is critical for complete male fertility. © 2015 by the Society for the Study of Reproduction, Inc.

Ebola virus-like particles produced in insect cells exhibit dendritic cell stimulating activity and induce neutralizing antibodies

International Nuclear Information System (INIS)

Ye Ling; Lin Jianguo; Sun Yuliang; Bennouna, Soumaya; Lo, Michael; Wu Qingyang; Bu Zhigao; Pulendran, Bali; Compans, Richard W.; Yang Chinglai

2006-01-01

Recombinant baculoviruses (rBV) expressing Ebola virus VP40 (rBV-VP40) or GP (rBV-GP) proteins were generated. Infection of Sf9 insect cells by rBV-VP40 led to assembly and budding of filamentous particles from the cell surface as shown by electron microscopy. Ebola virus-like particles (VLPs) were produced by coinfection of Sf9 cells with rBV-VP40 and rBV-GP, and incorporation of Ebola GP into VLPs was demonstrated by SDS-PAGE and Western blot analysis. Recombinant baculovirus infection of insect cells yielded high levels of VLPs, which were shown to stimulate cytokine secretion from human dendritic cells similar to VLPs produced in mammalian cells. The immunogenicity of Ebola VLPs produced in insect cells was evaluated by immunization of mice. Analysis of antibody responses showed that most of the GP-specific antibodies were of the IgG2a subtype, while no significant level of IgG1 subtype antibodies specific for GP was induced, indicating the induction of a Th1-biased immune response. Furthermore, sera from Ebola VLP immunized mice were able to block infection by Ebola GP pseudotyped HIV virus in a single round infection assay, indicating that a neutralizing antibody against the Ebola GP protein was induced. These results show that production of Ebola VLPs in insect cells using recombinant baculoviruses represents a promising approach for vaccine development against Ebola virus infection

Sex-specific phenotypes of hyperthyroidism and hypothyroidism in mice

OpenAIRE

Rakov, Helena; Engels, Kathrin; H?nes, Georg Sebastian; Strucksberg, Karl-Heinz; Moeller, Lars Christian; K?hrle, Josef; Zwanziger, Denise; F?hrer, Dagmar

2016-01-01

Background Thyroid dysfunction is more common in the female population, however, the impact of sex on disease characteristics has rarely been addressed. Using a murine model, we asked whether sex has an influence on phenotypes, thyroid hormone status, and thyroid hormone tissue response in hyper- and hypothyroidism. Methods Hypo- and hyperthyroidism were induced in 5-month-old female and male wildtype C57BL/6N mice, by LoI/MMI/ClO4 ? or T4 i.p. treatment over 7?weeks, and control animals unde...

Liver-Derived Insulin-Like Growth Factor-I is Involved in the Regulation of Blood Pressure in Mice

DEFF Research Database (Denmark)

Tivesten, Asa; Bollano, Entela; Andersson, Irene

2002-01-01

IGF-I has been suggested to be of importance for cardiovascular structure and function, but the relative role of locally produced and liver-derived endocrine IGF-I remains unclear. Using the Cre-LoxP recombination system, we have previously created transgenic mice with a liver-specific, inducible...... IGF-I knockout (LI-IGF-I-/-). To examine the role of liver-derived IGF-I in cardiovascular physiology, liver-derived IGF-I was inactivated at 4 wk of age, resulting in a 79% reduction of serum IGF-I levels. At 4 months of age, systolic blood pressure (BP) was increased in LI-IGF-I-/- mice...... to endothelial dysfunction associated with increased expression of endothelin-1 and impaired vasorelaxation of resistance vessels. In conclusion, our findings suggest that liver-derived IGF-I is involved in the regulation of BP in mice....

Frontal Lobe Contusion in Mice Chronically Impairs Prefrontal-Dependent Behavior.

Directory of Open Access Journals (Sweden)

Austin Chou

Full Text Available Traumatic brain injury (TBI is a major cause of chronic disability in the world. Moderate to severe TBI often results in damage to the frontal lobe region and leads to cognitive, emotional, and social behavioral sequelae that negatively affect quality of life. More specifically, TBI patients often develop persistent deficits in social behavior, anxiety, and executive functions such as attention, mental flexibility, and task switching. These deficits are intrinsically associated with prefrontal cortex (PFC functionality. Currently, there is a lack of analogous, behaviorally characterized TBI models for investigating frontal lobe injuries despite the prevalence of focal contusions to the frontal lobe in TBI patients. We used the controlled cortical impact (CCI model in mice to generate a frontal lobe contusion and studied behavioral changes associated with PFC function. We found that unilateral frontal lobe contusion in mice produced long-term impairments to social recognition and reversal learning while having only a minor effect on anxiety and completely sparing rule shifting and hippocampal-dependent behavior.

Development of a lion-specific interferon-gamma assay.

Science.gov (United States)

Maas, M; van Kooten, P J S; Schreuder, J; Morar, D; Tijhaar, E; Michel, A L; Rutten, V P M G

2012-10-15

The ongoing spread of bovine tuberculosis (BTB) in African free-ranging lion populations, for example in the Kruger National Park, raises the need for diagnostic assays for BTB in lions. These, in addition, would be highly relevant for zoological gardens worldwide that want to determine the BTB status of their lions, e.g. for translocations. The present study concerns the development of a lion-specific IFN-γ assay, following the production and characterization of monoclonal antibodies specific for lion interferon-gamma (IFN-γ). Recombinant lion IFN-γ (rLIFN-γ) was produced in mammalian cells and used to immunize mice to establish hybridoma cell lines producing monoclonal antibodies. These were used to develop a sensitive, lion IFN-γ-specific capture ELISA, able to detect rLIFN-γ to the level of 160 pg/ml. Recognition of native lion IFN-γ was shown in an initial assessment of supernatants of mitogen stimulated whole blood cultures of 11 known BTB-negative lions. In conclusion, the capture ELISA shows potential as a diagnostic assay for bovine tuberculosis in lions. Preliminary results also indicate the possible use of the test for other (feline) species. Copyright © 2012 Elsevier B.V. All rights reserved.

Dopamine D3 receptor knockout mice exhibit abnormal nociception in a sex-different manner.

Science.gov (United States)

Liu, Peng; Xing, Bo; Chu, Zheng; Liu, Fei; Lei, Gang; Zhu, Li; Gao, Ya; Chen, Teng; Dang, Yong-Hui

2017-07-01

Pain is a complex and subjective experience. Previous studies have shown that mice lacking the dopamine D3 receptor (D3RKO) exhibit hypoalgesia, indicating a role of the D3 receptor in modulation of nociception. Given that there are sex differences in pain perception, there may be differences in responses to nociceptive stimuli between male and female D3RKO mice. In the current study, we examined the role of the D3 receptor in modulating nociception in male and female D3RKO mice. Acute thermal pain was modeled by hot-plate test. This test was performed at different temperatures including 52°C, 55°C, and 58°C. The von Frey hair test was applied to evaluate mechanical pain. And persistent pain produced by peripheral tissue injury and inflammation was modeled by formalin test. In the hot-plate test, compared with wild-type (WT) mice, D3RKO mice generally exhibited longer latencies at each of the three temperatures. Specially, male D3RKO mice showed hypoalgesia compared with male WT mice when the temperature was 55°C, while for the female mice, there was a statistical difference between genotypes when the test condition was 52°C. In the von Frey hair test, both male and female D3RKO mice exhibited hypoalgesia. In the formalin test, the male D3RKO mice displayed a similar nociceptive behavior as their sex-matched WT littermates, whereas significantly depressed late-phase formalin-induced nociceptive behaviors were observed in the female mutants. These findings indicated that the D3 receptor affects nociceptive behaviors in a sex-specific manner and that its absence induces more analgesic behavior in the female knockout mice. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Cooperative rearranging region size and free volume in As-Se glasses

Energy Technology Data Exchange (ETDEWEB)

Saiter, A; Saiter, J-M [Laboratoire PBM, UMR 6522, LECAP, Institut des Materiaux de Rouen, Universite de Rouen, Faculte des Sciences, Avenue de l' Universite BP 12, 76801 Saint Etienne du Rouvray (France); Golovchak, R; Shpotyuk, M; Shpotyuk, O [Lviv Scientific Research Institute of Materials of SRC ' Carat' , 202, Stryjska street, Lviv, UA-79031 (Ukraine)

2009-02-18

Glasses of the As-Se system have been used as model objects of the covalent disordered inorganic polymers to investigate the correlation between the cooperative rearranging region (CRR) size determined at the glass transition temperature and the free volume fraction in the glassy state. The CRR size has been determined using temperature modulated differential scanning calorimetry data according to Donth's approach, while the free volume fraction in the investigated materials has been estimated using positron annihilation lifetime spectroscopy data. The obtained results testify that the appearance of open-volume defects greater than 80 A{sup 3} leads to a significant decrease in the CRR size.

Cooperative rearranging region size and free volume in As-Se glasses

International Nuclear Information System (INIS)

Saiter, A; Saiter, J-M; Golovchak, R; Shpotyuk, M; Shpotyuk, O

2009-01-01

Glasses of the As-Se system have been used as model objects of the covalent disordered inorganic polymers to investigate the correlation between the cooperative rearranging region (CRR) size determined at the glass transition temperature and the free volume fraction in the glassy state. The CRR size has been determined using temperature modulated differential scanning calorimetry data according to Donth's approach, while the free volume fraction in the investigated materials has been estimated using positron annihilation lifetime spectroscopy data. The obtained results testify that the appearance of open-volume defects greater than 80 A 3 leads to a significant decrease in the CRR size.

Ramifide resonators for cyclotrons

International Nuclear Information System (INIS)

Smirnov, Yu.V.

2000-01-01

The resonators with the conductors ramified form for cyclotrons are systematized and separated into the self-contained class - the ramified resonators for cyclotrons (Carr). The ramified resonators are compared with the quarter-wave and half-wave nonramified resonators, accomplished from the transmitting lines fragments. The CRR are classified into two types: ones with the additional structural element, switched in parallel and in series. The CRR may include several additional structural elements. The CRR calculations may be concluded by analytical methods - the method of matrix calculation or the method of telegraph equations and numerical methods - by means of the ISFEL3D, MAFIA and other programs [ru

Rhox8 Ablation in the Sertoli Cells Using a Tissue-Specific RNAi Approach Results in Impaired Male Fertility in Mice1

Science.gov (United States)

Welborn, Joshua P.; Davis, Matthew G.; Ebers, Steven D.; Stodden, Genna R.; Hayashi, Kanako; Cheatwood, Joseph L.; Rao, Manjeet K.; MacLean, James A.

2015-01-01

The reproductive homeobox X-linked, Rhox, genes encode transcription factors that are selectively expressed in reproductive tissues. While there are 33 Rhox genes in mice, only Rhox and Rhox8 are expressed in Sertoli cells, suggesting that they may regulate the expression of somatic-cell gene products crucial for germ cell development. We previously characterized Rhox5-null mice, which are subfertile, exhibiting excessive germ cell apoptosis and compromised sperm motility. To assess the role of Rhox8 in Sertoli cells, we used a tissue-specific RNAi approach to knockdown RHOX8 in vivo, in which the Rhox5 promoter was used to drive Rhox8-siRNA transgene expression in the postnatal Sertoli cells. Western and immunohistochemical analysis confirmed Sertoli-specific knockdown of RHOX8. However, other Sertoli markers, Gata1 and Rhox5, maintained normal expression patterns, suggesting that the knockdown was specific. Interestingly, male RHOX8-knockdown animals showed significantly reduced spermatogenic output, increased germ cell apoptosis, and compromised sperm motility, leading to impaired fertility. Importantly, our results revealed that while some RHOX5-dependent factors were also misregulated in Sertoli cells of RHOX8-knockdown animals, the majority were not, and novel putative RHOX8-regulated genes were identified. This suggests that while reduction in levels of RHOX5 and RHOX8 in Sertoli cells elicits similar phenotypes, these genes are not entirely redundant. Taken together, our study underscores the importance of Rhox genes in male fertility and suggests that Sertoli cell-specific expression of Rhox5 and Rhox8 is critical for complete male fertility. PMID:25972016

Engineering Specificity and Function of Therapeutic Regulatory T Cells

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Jenny L. McGovern

2017-11-01

Full Text Available Adoptive therapy with polyclonal regulatory T cells (Tregs has shown efficacy in suppressing detrimental immune responses in experimental models of autoimmunity and transplantation. The lack of specificity is a potential limitation of Treg therapy, as studies in mice have demonstrated that specificity can enhance the therapeutic potency of Treg. We will discuss that vectors encoding T cell receptors or chimeric antigen receptors provide an efficient gene-transfer platform to reliably produce Tregs of defined antigen specificity, thus overcoming the considerable difficulties of isolating low-frequency, antigen-specific cells that may be present in the natural Treg repertoire. The recent observations that Tregs can polarize into distinct lineages similar to the Th1, Th2, and Th17 subsets described for conventional T helper cells raise the possibility that Th1-, Th2-, and Th17-driven pathology may require matching Treg subsets for optimal therapeutic efficacy. In the future, genetic engineering may serve not only to enforce FoxP3 expression and a stable Treg phenotype but it may also enable the expression of particular transcription factors that drive differentiation into defined Treg subsets. Together, established and recently developed gene transfer and editing tools provide exciting opportunities to produce tailor-made antigen-specific Treg products with defined functional activities.

Intranasal delivery of E-selectin reduces atherosclerosis in ApoE-/- mice.

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Xinhui Li

Full Text Available Mucosal tolerance to E-selectin prevents stroke and protects against ischemic brain damage in experimental models of stroke studying healthy animals or spontaneously hypertensive stroke-prone rats. A reduction in inflammation and neural damage was associated with immunomodulatory or "tolerogenic" responses to E-selectin. The purpose of the current study on ApoE deficient mice is to assess the capacity of this stroke prevention innovation to influence atherosclerosis, a major underlying cause for ischemic strokes; human E-selectin is being translated as a potential clinical prevention strategy for secondary stroke. Female ApoE-/- mice received intranasal delivery of E-selectin prior to (pre-tolerization or simultaneously with initiation of a high-fat diet. After 7 weeks on the high-fat diet, lipid lesions in the aorta, serum triglycerides, and total cholesterol were assessed as markers of atherosclerosis development. We also assessed E-selectin-specific antibodies and cytokine responses, in addition to inflammatory responses that included macrophage infiltration of the aorta and altered gene expression profiles of aortic mRNA. Intranasal delivery of E-selectin prior to initiation of high-fat chow decreased atherosclerosis, serum total cholesterol, and expression of the leucocyte chemoattractant CCL21 that is typically upregulated in atherosclerotic lesions of ApoE-/- mice. This response was associated with the induction of E-selectin specific cells producing the immunomodulatory cytokine IL-10 and immunosuppressive antibody isotypes. Intranasal administration of E-selectin generates E-selectin specific immune responses that are immunosuppressive in nature and can ameliorate atherosclerosis, a major risk factor for ischemic stroke. These results provide additional preclinical support for the potential of induction of mucosal tolerance to E-selectin to prevent stroke.

Heat production during contraction in skeletal muscle of hypothyroid mice

International Nuclear Information System (INIS)

Leijendekker, W.J.; van Hardeveld, C.; Elzinga, G.

1987-01-01

The effect of hypothyroidism on tension-independent and -dependent heat produced during a twitch and a tetanic contraction of extensor digitorum longus (EDL) and soleus muscle of mice was examined. The amount of heat produced during a twitch and the rate of heat development during a tetanus of EDL and soleus were measured at and above optimal length. The effect of hypothyroidism on force production was 2+ cycling, these findings suggest that ATP splitting due to the Ca 2+ cycling is reduced in hypothyroid mice. This conclusion was strengthened by the observation that the oxalate-supported 45 Ca 2+ -uptake activity and 45 Ca 2+ -loading capacity of muscle homogenates from hypothyroid mice were reduced, respectively, to 51 and to 65% in soleus and to 63 and 73% in EDL muscle as compared with euthyroid mice. The tension-dependent rate of heat development during a tetanus was also decreased in soleus muscle of hypothyroid mice. This suggests a lower rate of ATP hydrolysis related to cross-bridge cycling in this muscle due to the hypothyroid state

Factors related to resistance to hematopoietic death in mice

International Nuclear Information System (INIS)

Mori, Nobuko; Okumoto, Masaaki; Yonezawa, Morio; Nishikawa, Ryosuke; Takamori, Yasuhiko; Esaki, Kozaburo.

1994-01-01

Mouse strain difference in the radiosensitivity to hematopoietic death is thought to be determined by several factors besides radiosensitivity and the initial number of hematopoietic stem cells. Factors related to the survival of mice exposed to X-irradiation were analyzed using BALB/cHeA and STS/A strains whose LD 50/30 values differ markedly (BALB/cHeA, 5.55 Gy; STS/A, 8.45 Gy). STS/A mice exposed to 4 Gy of X-irradiation showed a small reduction but rapid recovery of blood cells (leukocytes, erythrocytes, and thrombocytes) when compared with BALB/cHeA mice. The survival of endogenous and exogenous CFU-S was much higher, by a magnitude of one log or more, in STS/A mice than those in BALB/cHeA mice; whereas the initial numbers of femoral CFU-S were similar for the two strains. The recovery of exogenous CFU-S was much more rapid in STS/A mice than it was in BALB/cHeA mice after 4 Gy of X-irradiation. Furthermore, spleen colonies produced by the transfusion of STS/A marrow cells into syngeneic recipients were significantly larger than those produced by BALB/cHeA marrow cells, regardless of whether the mice used for sources of marrow cells had been irradiated. But, there was no such difference when unirradiated marrow cells from the two strains were transfused into (BALB/cHeA X STS/A) F 1 recipients. These results indicate the possible contribution of a host factor (s) that stimulates the growth of spleen colonies after radiation to the radioresistance of STS/A mice, in addition to the primary effect of higher number of survivals of endogenous and exogenous CFU-S in STS/A mice. (author)

The hypoxic cytotoxin SR 4233 increases the effectiveness of radioimmunotherapy in mice with human non-Hodgkin's lymphoma xenografts.

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Wilder, R B; McGann, J K; Sutherland, W R; Waller, E K; Minchinton, A I; Goris, M L; Knox, S J

1994-01-01

To determine if either the hypoxic cell radiosensitizer etanidazole (SR 2508) or the hypoxic cytotoxin SR 4233 could improve the effectiveness of radioimmunotherapy. LC4 (an IgG1 monoclonal antibody directed toward malignant T cells) and MB-1 (an irrelevant isotype-matched control antibody) were injected intraperitoneally into severe combined immunodeficient phenotype mice with human cutaneous T cell lymphoma xenografts in order to determine the distribution of the antibodies in the tumors and normal tissues as a function of time. Computerized-pO2-histography was used to measure the median oxygen tension in the tumors. Tumor-bearing mice were treated with: (a) LC4; (b) 90Y-LC4; (c) 90Y-MB-1; (d) whole body irradiation delivered via an external 137Cs source; (e) etanidazole and 90Y-LC4; (f) SR 4233 and 90Y-LC4; (g) etanidazole; and (h) SR 4233. An additional group of mice received no treatment and served as controls. A tumor growth delay assay was used to assess the effectiveness of the different treatment regimens. LC4 accumulated in the tumors to a significantly greater extent than MB-1 (p LC4 by itself was able to produce a minor decrease in tumor size (control vs. LC4; p = 0.001). 90Y-LC4 produced greater tumor growth delay than LC4 alone (LC4 vs. 90Y-LC4; p = 0.01); however, the Yttrium-90 caused neutropenia and weight loss. The 90Y-labeled tumor-specific and non-specific antibodies both exerted greater tumor growth delay than externally delivered whole body irradiation (p LC4 (90Y-LC4 vs etanidazole and 90Y-LC4, p = 0.13). SR 4233, on the other hand, did enhance the tumor growth delay produced by 90Y-LC4 (90Y-LC4 vs. SR 4233 and 90Y-LC4, p = 0.046). The neutropenia and weight loss caused by 90Y-LC4 were exacerbated slightly (< 10%) by the administration of SR 4233. A first generation hypoxic cytotoxin, SR 4233, was able to enhance the tumor growth delay produced by radioimmunotherapy in severe combined immunodeficient phenotype mice with human cutaneous T cell

3D Modelling and Printing Technology to Produce Patient-Specific 3D Models.

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Birbara, Nicolette S; Otton, James M; Pather, Nalini

2017-11-10

A comprehensive knowledge of mitral valve (MV) anatomy is crucial in the assessment of MV disease. While the use of three-dimensional (3D) modelling and printing in MV assessment has undergone early clinical evaluation, the precision and usefulness of this technology requires further investigation. This study aimed to assess and validate 3D modelling and printing technology to produce patient-specific 3D MV models. A prototype method for MV 3D modelling and printing was developed from computed tomography (CT) scans of a plastinated human heart. Mitral valve models were printed using four 3D printing methods and validated to assess precision. Cardiac CT and 3D echocardiography imaging data of four MV disease patients was used to produce patient-specific 3D printed models, and 40 cardiac health professionals (CHPs) were surveyed on the perceived value and potential uses of 3D models in a clinical setting. The prototype method demonstrated submillimetre precision for all four 3D printing methods used, and statistical analysis showed a significant difference (p3D printed models, particularly using multiple print materials, were considered useful by CHPs for preoperative planning, as well as other applications such as teaching and training. This study suggests that, with further advances in 3D modelling and printing technology, patient-specific 3D MV models could serve as a useful clinical tool. The findings also highlight the potential of this technology to be applied in a variety of medical areas within both clinical and educational settings. Copyright © 2017 Australian and New Zealand Society of Cardiac and Thoracic Surgeons (ANZSCTS) and the Cardiac Society of Australia and New Zealand (CSANZ). Published by Elsevier B.V. All rights reserved.

Induction of Interleukin-10 Producing Dendritic Cells As a Tool to Suppress Allergen-Specific T Helper 2 Responses

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Stefan Schülke

2018-03-01

Full Text Available Dendritic cells (DCs are gatekeepers of the immune system that control induction and polarization of primary, antigen-specific immune responses. Depending on their maturation/activation status, the molecules expressed on their surface, and the cytokines produced DCs have been shown to either elicit immune responses through activation of effector T cells or induce tolerance through induction of either T cell anergy, regulatory T cells, or production of regulatory cytokines. Among the cytokines produced by tolerogenic DCs, interleukin 10 (IL-10 is a key regulatory cytokine limiting und ultimately terminating excessive T-cell responses to microbial pathogens to prevent chronic inflammation and tissue damage. Because of their important role in preventing autoimmune diseases, transplant rejection, allergic reactions, or in controlling chronic inflammation DCs have become an interesting tool to modulate antigen-specific immune responses. For the treatment of allergic inflammation, the aim is to downregulate allergen-specific T helper 2 (Th2 responses and the associated clinical symptoms [allergen-driven Th2 activation, Th2-driven immunoglobulin E (IgE production, IgE-mediated mast cell and basophil activation, allergic inflammation]. Here, combining the presentation of allergens by DCs with a pro-tolerogenic, IL-10-producing phenotype is of special interest to modulate allergen-specific immune responses in the treatment of allergic diseases. This review discusses the reported strategies to induce DC-derived IL-10 secretion for the suppression of allergen-specific Th2-responses with a focus on IL-10 treatment, IL-10 transduction, and the usage of both whole bacteria and bacteria-derived components. Interestingly, while IL-10-producing DCs induced either by IL-10 treatment or IL-10 transduction are arrested in an immature/semi-mature state, treatment of DCs with live or killed bacteria as well as isolated bacterial components results in the induction of

Tissue Specific Expression Of Sprouty1 In Mice Protects Against High Fat Diet Induced Fat Accumulation, Bone Loss, And Metabolic Dysfunction

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Urs, Sumithra; Henderson, Terry; Le, Phuong; Rosen, Clifford J.; Liaw, Lucy

2012-01-01

We recently characterized Sprouty1 (Spry1), a growth factor signaling inhibitor as a regulator of marrow progenitor cells promoting osteoblast differentiation at the expense of adipocytes. Adipose tissue specific Spry1 expression in mice resulted in increased bone mass and reduced body fat while conditional knockout of Spry1 had the opposite effect with decreased bone and increased body fat. Because Spry1 suppresses normal fat development, we tested the hypothesis that Spry1 expression prevents high fat diet-induced obesity, bone loss, and associated lipid abnormalities and demonstrate that Spry1 has a long-term protective effect on mice fed a high caloric diet. We studied diet-induced obesity in mice with fatty acid binding promoter (aP2)-driven expression or conditional knockout of Spry1 in adipocytes. Phenotyping was performed by whole body dual-energy X-ray absorptiometry, microCT, histology and blood analysis. In conditional Spry1 null mice, high fat diet increased body fat by 40%, impaired glucose regulation, and led to liver steatosis. However, over-expression of Spry1 led to 35% lower body fat, reduced bone loss, and normal metabolic function compared to single transgenics. This protective phenotype was associated with decreased circulating insulin (70%) and leptin (54%) compared to controls on a high fat diet. Additionally, Spry1 expression decreased adipose tissue inflammation by 45%. We show that conditional Spry1 expression in adipose tissue protects against high fat diet-induced obesity and associated bone loss. PMID:22142492

Response rates in studies of couples coping with cancer: a systematic review.

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Dagan, Meirav; Hagedoorn, Mariët

2014-08-01

Recruiting couples for psychological studies can be challenging. This brief report is the first to examine the average couples' response rate and to systematically review the quality of reporting of couples' response rate in studies of couples coping with cancer. A systematic review (1980-2011) was conducted, including 83 studies meeting the inclusion criteria of being published in peer-reviewed journals, describing quantitative findings using a cross-sectional or longitudinal design. Overall reporting was unsatisfactory in more than half of the included studies. As a consequence, the couples' response rate (CRR; all analyzed couples divided by the number of eligible partnered patients/couples approached) could be calculated for only 33 samples. This CRR varied considerably across studies from 25% to 90% (CRRM = 58%, SD = 17%). The rates reported in the articles (M = 65%) were often higher than the average CRR (CRRM = 57%) of these samples. This systematic review revealed incomplete reporting of response rate. Therefore, it cannot be firmly concluded that the average CRR reported is representative for all studies on couples coping with cancer. Finally, the figures presented, which are often more favorable than the CRR, may create the impression that the sample is more representative of the target population than it actually is. This has consequences for implementing the findings of such studies into practice. The results are critically discussed, and recommendations for improvement are provided.

Early-life nutrition modulates the epigenetic state of specific rDNA genetic variants in mice.

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Holland, Michelle L; Lowe, Robert; Caton, Paul W; Gemma, Carolina; Carbajosa, Guillermo; Danson, Amy F; Carpenter, Asha A M; Loche, Elena; Ozanne, Susan E; Rakyan, Vardhman K

2016-07-29

A suboptimal early-life environment, due to poor nutrition or stress during pregnancy, can influence lifelong phenotypes in the progeny. Epigenetic factors are thought to be key mediators of these effects. We show that protein restriction in mice from conception until weaning induces a linear correlation between growth restriction and DNA methylation at ribosomal DNA (rDNA). This epigenetic response remains into adulthood and is restricted to rDNA copies associated with a specific genetic variant within the promoter. Related effects are also found in models of maternal high-fat or obesogenic diets. Our work identifies environmentally induced epigenetic dynamics that are dependent on underlying genetic variation and establishes rDNA as a genomic target of nutritional insults. Copyright © 2016, American Association for the Advancement of Science.

Hexachlorobenzene stimulates uroporphyria in low affinity AHR mice without increasing CYP1A2

International Nuclear Information System (INIS)

Gorman, Nadia; Trask, Heidi S.; Robinson, Susan W.; Sinclair, Jacqueline F.; Gerhard, Glenn S.; Smith, Andrew G.; Sinclair, Peter R.

2007-01-01

Hexachlorobenzene (HCB), a weak ligand of the aryl hydrocarbon receptor (AHR), causes hepatic uroporphyrin (URO) accumulation (uroporphyria) in humans and animals. CYP1A2 has been shown to be necessary in the development of uroporphyria in mice. Using mice expressing the low affinity form of the AH receptor (AHRd), we investigated whether the enhancement of uroporphyria by HCB involves an obligatory increase in CYP1A2 as measured by specific enzyme assays and immunoblotting. We compared the ability of HCB, in combination with iron dextran and the porphyrin precursor, 5-aminolevulinate (ALA), to cause uroporphyria in a strain of mice (C57BL/6) which expresses the high affinity form of the receptor (AHRb 1 ), with three strains of mice (SWR and two 129 sublines) expressing the low affinity AHRd. In C57BL/6 mice, HCB-enhanced uroporphyria was associated with a doubling of CYP1A2. HCB treatment produced uroporphyria in iron-loaded mice expressing AHRd, even though there was little or no increase in CYP1A2. Cyp1a2(-/-) mice in a 129 background were completely resistant to HCB-induced uroporphyria, and female Hfe(-/-) 129 mice, in which the levels of hepatic CYP1A2 were half of those of the male levels, responded poorly. The effect of exogenous iron, administered in the form of iron dextran, on HCB enhancement of uroporphryia could be replicated utilizing the endogenous hepatic iron accumulated in 129 Hfe(-/-) mice. In conclusion, some minimal basal expression of CYP1A2 is essential for HCB-mediated enhancement of uroporphyria, but increases in CYP1A2 above that level are not essential

Defective B cell response to T-dependent immunization in lupus-prone mice

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Niu, Haitao; Sobel, Eric S.; Morel, Laurence

2009-01-01

Lupus anti-nuclear Abs show the characteristics of Ag-driven T cell-dependent (TD) humoral responses. If autoAgs elicit the same response as exogenous Ags, lupus should enhance humoral responses to immunization. Blunted responses to various immunizations have, however, been reported in a significant portion of lupus patients. In this study, we show that lupus-prone B6.Sle1.Sle2.Sle3 (B6.TC) mice produce significantly less Ab in response to TD immunization than congenic controls, while producing significantly more total Ig. This blunted Ab response to TD Ag could be reconstituted with B6.TC B and CD4+ T cells. Multiple defects were found in the B6.TC response to NP-KLH as compared to total Ig, including a smaller percentage of B cells participating to the NP-response, a reduced entry into germinal centers, and highly defective production of NP-specific long-lived plasma cells in the bone marrow. B6.TC plasma cells expressed reduced levels of FcγRIIb, which suggests that reduced apoptosis in resident plasma cells prevents the establishment of newly-formed NP-specific plasma cells in bone marrow niches. Overall, these results show that lupus-prone mice responded differently to auto- and exogenous antigens and suggest that low FcγRIIb, hypergammaglobulinemia and high autoantibody production would be predictive of a poor response to immunization in lupus patients. PMID:18924209

Inhibition of serotonin but not norepinephrine transport during development produces delayed, persistent perturbations of emotional behaviors in mice.

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Ansorge, Mark S; Morelli, Emanuela; Gingrich, Jay A

2008-01-02

Serotonin (5-HT) acts as a neurotransmitter, but also modulates brain maturation during early development. The demonstrated influence of genetic variants on brain function, personality traits, and susceptibility to neuropsychiatric disorders suggests a critical importance of developmental mechanisms. However, little is known about how and when developmentally perturbed 5-HT signaling affects circuitry and resulting behavior. The 5-HT transporter (5-HTT) is a key regulator of extracellular 5-HT levels and we used pharmacologic strategies to manipulate 5-HTT function during development and determine behavioral consequences. Transient exposure to the 5-HTT inhibitors fluoxetine, clomipramine, and citalopram from postnatal day 4 (P4) to P21 produced abnormal emotional behaviors in adult mice. Similar treatment with the norepinephrine transporter (NET) inhibitor, desipramine, did not adversely affect adult behavior, suggesting that 5-HT and norepinephrine (NE) do not share the same effects on brain development. Shifting our period of treatment/testing to P90/P185 failed to mimic the effect of earlier exposure, demonstrating that 5-HT effects on adult behavior are developmentally specific. We have hypothesized that early-life perturbations of 5-HT signaling affect corticolimbic circuits that do not reach maturity until the peri-adolescent period. In support of this idea, we found that abnormal behaviors resulting from postnatal fluoxetine exposure have a post-pubescent onset and persist long after reaching adult age. A better understanding of the underlying 5-HT sensitive circuits and how they are perturbed should lead to new insights into how various genetic polymorphisms confer their risk to carriers. Furthermore, these studies should help determine whether in utero exposure to 5-HTT blocking drugs poses a risk for behavioral abnormalities in later life.

Outer Membrane Vesicle Vaccines from Biosafe Surrogates Prevent Acute Lethal Glanders in Mice

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Michael H. Norris

2018-01-01

Full Text Available Burkholderia mallei is a host-adapted Gram-negative mammalian pathogen that causes the severe disease glanders. Glanders can manifest as a rapid acute progression or a chronic debilitating syndrome primarily affecting solipeds and humans in close association with infected animals. In USA, B. mallei is classified as one of the most important bacterial biothreat agents. Presently, there is no licensed glanders vaccine available for humans or animals. In this work, outer membrane vesicles (OMVs were isolated from three attenuated biosafe bacterial strains, Burkholderia pseudomallei Bp82, B. thailandensis E555, and B. thailandensis TxDOH and used to vaccinate mice. B. thailandensis OMVs induced significantly higher antibody responses that were investigated. B. mallei specific serum antibody responses were of higher magnitude in mice vaccinated with B. thailandensis OMVs compared to levels in mice vaccinated with B. pseudomallei OMVs. OMVs derived from biosafe strains protected mice from acute lethal glanders with vesicles from the two B. thailandensis strains affording significant protection (>90% up to 35 days post-infection with some up to 60 days. Organ loads from 35-day survivors indicated bacteria colonization of the lungs, liver, and spleen while those from 60 days had high CFUs in the spleens. The highest antibody producing vaccine (B. thailandensis E555 OMVs also protected C57BL/6 mice from acute inhalational glanders with evidence of full protection.

Oral Immunization with Recombinant Norwalk Virus-Like Particles Induces a Systemic and Mucosal Immune Response in Mice

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Ball, Judith M.; Hardy, Michele E.; Atmar, Robert L.; Conner, Margaret E.; Estes, Mary K.

1998-01-01

Recombinant Norwalk virus-like particles (rNV VLPs) produced in insect cells were evaluated as an oral immunogen in CD1 and BALB/c mice by monitoring rNV-specific serum total and subclass immunoglobulin G (IgG) and intestinal IgA responses. Dose and kinetics of response were evaluated in the presence and absence of the mucosal adjuvant cholera toxin (CT). rNV-specific serum IgG and intestinal IgA were detected in the absence of CT, and the number of responders was not significantly different ...

Neutrophil trails guide influenza-specific CD8⁺ T cells in the airways.

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Lim, Kihong; Hyun, Young-Min; Lambert-Emo, Kris; Capece, Tara; Bae, Seyeon; Miller, Richard; Topham, David J; Kim, Minsoo

2015-09-04

During viral infections, chemokines guide activated effector T cells to infection sites. However, the cells responsible for producing these chemokines and how such chemokines recruit T cells are unknown. Here, we show that the early recruitment of neutrophils into influenza-infected trachea is essential for CD8(+) T cell-mediated immune protection in mice. We observed that migrating neutrophils leave behind long-lasting trails that are enriched in the chemokine CXCL12. Experiments with granulocyte-specific CXCL12 conditionally depleted mice and a CXCR4 antagonist revealed that CXCL12 derived from neutrophil trails is critical for virus-specific CD8(+) T cell recruitment and effector functions. Collectively, these results suggest that neutrophils deposit long-lasting, chemokine-containing trails, which may provide both chemotactic and haptotactic cues for efficient CD8(+) T cell migration and localization in influenza-infected tissues. Copyright © 2015, American Association for the Advancement of Science.

Reduced immune responses in chimeric mice engrafted with bone marrow cells from mice with airways inflammation.

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Scott, Naomi M; Ng, Royce L X; McGonigle, Terence A; Gorman, Shelley; Hart, Prue H

2015-11-01

During respiratory inflammation, it is generally assumed that dendritic cells differentiating from the bone marrow are immunogenic rather than immunoregulatory. Using chimeric mice, the outcomes of airways inflammation on bone marrow progenitor cells were studied. Immune responses were analyzed in chimeric mice engrafted for >16 weeks with bone marrow cells from mice with experimental allergic airways disease (EAAD). Responses to sensitization and challenge with the allergen causing inflammation in the bone marrow-donor mice were significantly reduced in the chimeric mice engrafted with bone marrow cells from mice with EAAD (EAAD-chimeric). Responses to intranasal LPS and topical fluorescein isothiocyanate (non-specific challenges) were significantly attenuated. Fewer activated dendritic cells from the airways and skin of the EAAD-chimeric mice could be tracked to the draining lymph nodes, and may contribute to the significantly reduced antigen/chemical-induced hypertrophy in the draining nodes, and the reduced immune responses to sensitizing allergens. Dendritic cells differentiating in vitro from the bone marrow of >16 weeks reconstituted EAAD-chimeric mice retained an ability to poorly prime immune responses when transferred into naïve mice. Dendritic cells developing from bone marrow progenitors during airways inflammation are altered such that daughter cells have reduced antigen priming capabilities.

CCL28 induces mucosal homing of HIV-1-specific IgA-secreting plasma cells in mice immunized with HIV-1 virus-like particles.

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Veronica Rainone

Full Text Available Mucosae-associated epithelial chemokine (MEC or CCL28 binds to CCR3 and CCR10 and recruits IgA-secreting plasma cells (IgA-ASCs in the mucosal lamina propria. The ability of this chemokine to enhance migration of IgA-ASCs to mucosal sites was assessed in a mouse immunization model using HIV-1(IIIB Virus-like particles (VLPs. Mice receiving either HIV-1(IIIB VLPs alone, CCL28 alone, or the irrelevant CCL19 chemokine were used as controls. Results showed a significantly increased CCR3 and CCR10 expression on CD19(+ splenocytes of HIV-1(IIIB VPL-CCL28-treated mice. HIV-1 Env-specific IFN-γ, IL-4 and IL-5 production, total IgA, anti-Env IgA as well as gastro-intestinal mucosal IgA-secreting plasma cells were also significantly augmented in these mice. Notably, sera and vaginal secretions from HIV-1(IIIB VLP-CCL28-treated mice exhibited an enhanced neutralizing activity against both a HIV-1/B-subtype laboratory strain and a heterologous HIV-1/C-subtype primary isolate. These data suggest that CCL28 could be useful in enhancing the IgA immune response that will likely play a pivotal role in prophylactic HIV vaccines.

Fshb-iCre mice are efficient and specific Cre deleters for the gonadotrope lineage.

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Wang, Huizhen; Hastings, Richard; Miller, William L; Kumar, T Rajendra

2016-01-05

Genetic analysis of development and function of the gonadotrope cell lineage within mouse anterior pituitary has been greatly facilitated by at least three currently available Cre strains in which Cre was either knocked into the Gnrhr locus or expressed as a transgene from Cga and Lhb promoters. However, in each case there are some limitations including CRE expression in thyrotropes within pituitary or ectopic expression outside of pituitary, for example in some populations of neurons or gonads. Hence, these Cre strains often pose problems with regard to undesirable deletion of alleles in non-gonadotrope cells, fertility and germline transmission of mutant alleles. Here, we describe generation and characterization of a new Fshb-iCre deleter strain using 4.7 kb of ovine Fshb promoter regulatory sequences driving iCre expression exclusively in the gonadotrope lineage within anterior pituitary. Fshb-iCre mice develop normally, display no ectopic CRE expression in gonads and are fertile. When crossed onto a loxP recombination-mediated red to green color switch reporter mouse genetic background, in vivo CRE recombinase activity is detectable in gonadotropes at more than 95% efficiency and the GFP-tagged gonadotropes readily purified by fluorescence activated cell sorting. We demonstrate the applicability of this Fshb-iCre deleter strain in a mouse model in which Dicer is efficiently and selectively deleted in gonadotropes. We further show that loss of DICER-dependent miRNAs in gonadotropes leads to profound suppression of gonadotropins resulting in male and female infertility. Thus, Fshb-iCre mice serve as a new genetic tool to efficiently manipulate gonadotrope-specific gene expression in vivo. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Proteomic analysis of skeletal muscle in insulin-resistant mice: response to 6-week aerobic exercise.

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Hairui Yuan

Full Text Available Aerobic exercise has beneficial effects on both weight control and skeletal muscle insulin sensitivity through a number of specific signaling proteins. To investigate the targets by which exercise exerts its effects on insulin resistance, an approach of proteomic screen was applied to detect the potential different protein expressions from skeletal muscle of insulin-resistant mice after prolonged aerobic exercise training and their sedentary controls. Eighteen C57BL/6 mice were divided into two groups: 6 mice were fed normal chow (NC and 12 mice were fed high-fat diet (HFD for 10 weeks to produce an IR model. The model group was then subdivided into HFD sedentary control (HC, n = 6 and HFD exercise groups (HE, n = 6. Mice in HE group underwent 6 weeks of treadmill running. After 6 weeks, mice were sacrificed and skeletal muscle was dissected. Total protein (n = 6, each group was extracted and followed by citrate synthase, 2D proteome profile analysis and immunoblot. Fifteen protein spots were altered between the NC and HC groups and 23 protein spots were changed between the HC and HE groups significantly. The results provided an array of changes in protein abundance in exercise-trained skeletal muscle and also provided the basis for a new hypothesis regarding the mechanism of exercise ameliorating insulin resistance.

Biochemical behavior of Trypanosoma cruzi strains isolated from mice submitted to specific chemotherapy

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Jesila Pinto M. Marretto

1994-12-01

Full Text Available To investigate the influence of chemotherapy on the biochemical beha vior of Trypanosoma cruzi strains, three groups of mice were infected with one of three strains of T. cruzi of different biological and isoenzymic patterns (Peruvian, 21 SF and Colombian strains. Each group was subdivided into subgroups: 1 - treated with nifurtimox; 2 - treated with benznidazole and 3 - untreated infected controls. At the end of treatment, that lasted for 90 days, xenodiagnosis, sub inoculation of blood into new born mice and haemoculture were performed as tests of cure. From the positive tests, 22 samples of T. cruzi were isolated from all subgroups. Electrophoretic analysis of the isoenzymes PGM, GP1, ALAT and AS AT failed to show any difference between parasite strains isolated from treated and untreated mice, which indicates that no detectable clonal selection or parasite genetic markers alterations concerning the isoenzymes analysed have been determined by treatment with drugs of recognized antiparasitic effect, suggesting stability of the phenotypic characteristics of the three biological types of T. cruzi strains.

Efficient replication, and evolution of Sindbis virus genomes with non-canonical 3'A/U-rich elements (NC3ARE) in neonatal mice.

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James, Frederick D; Hietala, Katie A; Eldar, Dganit; Guess, Tiffany E; Cone, Cecil; Mundell, Nathan A; Mundall, Nathan; Barnett, Joey V; Raju, Ramaswamy

2007-12-01

Sindbis virus (SIN) is a mosquito-transmitted animal RNA virus. We previously reported that SIN genomes lacking a canonical 19 nt 3'CSE undergo novel repair processes in BHK cells to generate a library of stable atypical SIN genomes with non-canonical 3'A/U-rich elements (NC3AREs) adjacent to the 3' poly(A) tail [1]. To determine the stability and evolutionary pressures on the SIN genomes with NC3AREs to regain a 3'CSE, five representative SIN isolates and a wild type SIN were tested in newborn mice. The key findings of this study are: (a) all six SIN isolates, including those that have extensive NC3AREs in the 3'NTRs, replicate well and produce high titer viremia in newborn mice; (b) 7-9 successive passages of these isolates in newborn mice produced comparable levels of viremia; (c) while all isolates produced only small-sized plaques during primary infection in animals, both small- and large-sized plaques were generated in all other passages; (d) polymerase stuttering occurs on select 3' oligo(U) motifs to add more U residues within the NC3AREs; (e) the S3-8 isolate with an internal UAUUU motif in the 3'poly(A) tail maintains this element even after 9 passages in animals; (f) despite differences in 3'NTRs and variable tissue distribution, all SIN isolates appear to produce similar tissue pathology in infected animals. Competition experiments with wt SIN and atypical SIN isolates in BHK cells show dominance of wt SIN. As shown for BHK cells in culture, the 3'CSE of the SIN genome is not required for virus replication and genome stability in live animals. Since the NC3AREs of atypical SIN genomes are not specific to SIN replicases, alternate RNA motifs of alphavirus genome must confer specificity in template selection. These studies fulfill the need to confirm the long-term viability of atypical SIN genomes in newborn mice and offer a basis for exploring the use of atypical SIN genomes in biotechnology.

In vitro radionuclide therapy and in vivo scintigraphic imaging of alpha fetoprotein producing hepatocellular carcinoma by targeted sodium iodide symporter gene expression

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Kim, Kwang Il; Lee, Yong Jin; Lee, Tae Sup; Song, Inho; Cheon, Gi Jeong; Lim, Sang Moo; Kang, Joo Hyun [Korea Institute of Radiological and Medical and Medical Sciences, Seoul (Korea, Republic of); Chung, June Key [Seoul National Univ. College of Medicine, Seoul (Korea, Republic of)

2012-03-15

This study aimed to develop a gene expression targeting method for specific imaging and therapy of alpha fetoprotein (AFP) producing hepatocellular carcinoma (HCC) cells, using an adenovirus vector containing the human sodium/iodide symporter (hNIS) gene driven by an AFP enhancer/promoter. The recombinant adenovirus vector, AdAFPhNIS (containing the hNIS gene driven by human AFP enhancer/promoter) was prepared. After in vitro infection by the adenovirus, hNIS gene expression in AFP producing cells and in AFP nonproducing cells was investigated using {sup 125}I uptake assay and semi quantitative reverse transcription polymerase chain reaction (RT-PCR). The killing effect of {sup 131}I vitro clonogenic assay. In addition, tumor bearing mice were intravenously injected with the adenovirus, and scintigraphic images were obtained. The expression of hNIS was efficiently demonstrated by {sup 125}I uptake assay in AFP producing cells, but not in AFP nonproducing cells. AFP producing HCC targeted gene expression was confirmed at the mRNA level. Furthermore, in vitro clonogenic assay showed that hNIS gene expression induced by AdAFPhNIS infection in AFP producing cells caused more sensitivity to {sup 131}I than that in AFP nonproducing cells. Injected intravenously in HuH-7 tumor xenografts mice by adenovirus, the functional hNIS gene expression was confirmed in tumor by in vivo scintigraphic imaging. An AFP producing HCC was targeted with an adenovirus vector containing the hNIS gene using the AFP enhancer/promoter in vitro and in vivo. These findings demonstrate that AFP producing HCC specific molecular imaging and radionuclide gene therapy are feasible using this recombinant adenovirus vector system.

In vitro radionuclide therapy and in vivo scintigraphic imaging of alpha fetoprotein producing hepatocellular carcinoma by targeted sodium iodide symporter gene expression

International Nuclear Information System (INIS)

Kim, Kwang Il; Lee, Yong Jin; Lee, Tae Sup; Song, Inho; Cheon, Gi Jeong; Lim, Sang Moo; Kang, Joo Hyun; Chung, June Key

2012-01-01

This study aimed to develop a gene expression targeting method for specific imaging and therapy of alpha fetoprotein (AFP) producing hepatocellular carcinoma (HCC) cells, using an adenovirus vector containing the human sodium/iodide symporter (hNIS) gene driven by an AFP enhancer/promoter. The recombinant adenovirus vector, AdAFPhNIS (containing the hNIS gene driven by human AFP enhancer/promoter) was prepared. After in vitro infection by the adenovirus, hNIS gene expression in AFP producing cells and in AFP nonproducing cells was investigated using 125 I uptake assay and semi quantitative reverse transcription polymerase chain reaction (RT-PCR). The killing effect of 131 I vitro clonogenic assay. In addition, tumor bearing mice were intravenously injected with the adenovirus, and scintigraphic images were obtained. The expression of hNIS was efficiently demonstrated by 125 I uptake assay in AFP producing cells, but not in AFP nonproducing cells. AFP producing HCC targeted gene expression was confirmed at the mRNA level. Furthermore, in vitro clonogenic assay showed that hNIS gene expression induced by AdAFPhNIS infection in AFP producing cells caused more sensitivity to 131 I than that in AFP nonproducing cells. Injected intravenously in HuH-7 tumor xenografts mice by adenovirus, the functional hNIS gene expression was confirmed in tumor by in vivo scintigraphic imaging. An AFP producing HCC was targeted with an adenovirus vector containing the hNIS gene using the AFP enhancer/promoter in vitro and in vivo. These findings demonstrate that AFP producing HCC specific molecular imaging and radionuclide gene therapy are feasible using this recombinant adenovirus vector system

Sex-specific hippocampal 5-hydroxymethylcytosine is disrupted in response to acute stress.

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Papale, Ligia A; Li, Sisi; Madrid, Andy; Zhang, Qi; Chen, Li; Chopra, Pankaj; Jin, Peng; Keleş, Sündüz; Alisch, Reid S

2016-12-01

Environmental stress is among the most important contributors to increased susceptibility to develop psychiatric disorders. While it is well known that acute environmental stress alters gene expression, the molecular mechanisms underlying these changes remain largely unknown. 5-hydroxymethylcytosine (5hmC) is a novel environmentally sensitive epigenetic modification that is highly enriched in neurons and is associated with active neuronal transcription. Recently, we reported a genome-wide disruption of hippocampal 5hmC in male mice following acute stress that was correlated to altered transcript levels of genes in known stress related pathways. Since sex-specific endocrine mechanisms respond to environmental stimulus by altering the neuronal epigenome, we examined the genome-wide profile of hippocampal 5hmC in female mice following exposure to acute stress and identified 363 differentially hydroxymethylated regions (DhMRs) linked to known (e.g., Nr3c1 and Ntrk2) and potentially novel genes associated with stress response and psychiatric disorders. Integration of hippocampal expression data from the same female mice found stress-related hydroxymethylation correlated to altered transcript levels. Finally, characterization of stress-induced sex-specific 5hmC profiles in the hippocampus revealed 778 sex-specific acute stress-induced DhMRs some of which were correlated to altered transcript levels that produce sex-specific isoforms in response to stress. Together, the alterations in 5hmC presented here provide a possible molecular mechanism for the adaptive sex-specific response to stress that may augment the design of novel therapeutic agents that will have optimal effectiveness in each sex. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Producing infectious enterovirus type 71 in a rapid strategy

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Qin E-De

2010-06-01

Full Text Available Abstract Background Enterovirus 71 (EV71 is an etiologic agent of hand-foot-and-mouth disease (HFMD, and recent HFMD epidemics worldwide have been associated with a severe form of brainstem encephalitis associated with pulmonary edema and high case-fatality rates. EV71 contains a positive-sense single-stranded genome RNA of approximately 7400 bp in length which encodes a polyprotein with a single open reading frame (ORF, which is flanked by untranslated regions at both the 5' and 3' ends. Results A long distance RT-PCR assay was developed to amplify the full length genome cDNA of EV71 by using specific primes carrying a SP6 promoter. Then the in vitro synthesized RNA transcripts from the RT-PCR amplicons were then transfected into RD cells to produce the rescued virus. The rescued virus was further characterized by RT-PCR and indirect fluorescent-antibody (IFA assay in comparison with the wild type virus. The rescued viruses were infectious on RD cells and neurovirulent when intracerebrally injected into suckling mice. Conclusions Thus, we established a rapid method to produce the infectious full length cDNA of EV71 directly from RNA preparations and specific mutations can be easily engineered into the rescued enterovirus genome by this method.

Male mice song syntax depends on social contexts and influences female preferences

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Jonathan eChabout

2015-04-01

Full Text Available In 2005 Holy & Guo advanced the idea that male mice produce ultrasonic vocalizations (USV with some features similar to courtship songs of songbirds. Since then, studies showed that male mice emit USV songs in different contexts (sexual and other and possess a multisyllabic repertoire. Debate still exists for and against plasticity in their vocalizations. But the use of a multisyllabic repertoire can increase potential flexibility and information, in how elements are organized and recombined, namely syntax. In many bird species, modulating song syntax has ethological relevance for sexual behavior and mate preferences. In this study we exposed adult male mice to different social contexts and developed a new approach of analyzing their USVs based on songbird syntax analysis. We found that male mice modify their syntax, including specific sequences, length of sequence, repertoire composition, and spectral features, according to stimulus and social context. Males emit longer and simpler syllables and sequences when singing to females, but more complex syllables and sequences in response to fresh female urine. Playback experiments show that the females prefer the complex songs over the simpler ones. We propose the complex songs are to lure females in, whereas the directed simpler sequences are used for direct courtship. These results suggest that although mice have a much more limited ability of song modification, they could still be used as animal models for understanding some vocal communication features that songbirds are used for.

Reduction of the immunostainable length of the hippocampal dentate granule cells’ primary cilia in 3xAD-transgenic mice producing human Aβ1-42 and tau

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Chakravarthy, Balu; Gaudet, Chantal; Ménard, Michel; Brown, Leslie; Atkinson, Trevor; LaFerla, Frank M.; Ito, Shingo; Armato, Ubaldo; Dal Prà, Ilaria; Whitfield, James

2012-01-01

Highlights: ► Aβ and tau-induced neurofibrillary tangles play a key role in Alzheimer’s disease. ► Aβ 1-42 and mutant tau protein together reduce the primary cilium length. ► This shortening likely reduces cilium-dependent neurogenesis and memory function. ► This provides a model of an Aβ/tau targeting of a neuronal signaling organelle. -- Abstract: The hippocampal dentate gyrus is one of the two sites of continuous neurogenesis in adult rodents and humans. Virtually all dentate granule cells have a single immobile cilium with a microtubule spine or axoneme covered with a specialized cell membrane loaded with receptors such as the somatostatin receptor 3 (SSTR3), and the p75 neurotrophin receptor (p75 NTR ). The signals from these receptors have been reported to stimulate neuroprogenitor proliferation and the post-mitotic maturation of newborn granule cells into functioning granule cells. We have found that in 6–24-months-old triple transgenic Alzheimer’s disease model mice (3xTg-AD) producing both Aβ 1-42 and the mutant human tau protein tau P301L, the dentate granule cells still had immunostainable SSTR3- and p75 NTR -bearing cilia but they were only half the length of the immunostained cilia in the corresponding wild-type mice. However, the immunostainable length of the granule cell cilia was not reduced either in 2xTg-AD mice accumulating large amounts of Aβ 1-42 or in mice accumulating only a mutant human tau protein. Thus it appears that a combination of Aβ 1-42 and tau protein accumulation affects the levels of functionally important receptors in 3xTg-AD mice. These observations raise the important possibility that structural and functional changes in granule cell cilia might have a role in AD.

Inositol- and folate-resistant neural tube defects in mice lacking the epithelial-specific factor Grhl-3.

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Ting, Stephen B; Wilanowski, Tomasz; Auden, Alana; Hall, Mark; Voss, Anne K; Thomas, Tim; Parekh, Vishwas; Cunningham, John M; Jane, Stephen M

2003-12-01

The neural tube defects (NTDs) spina bifida and anencephaly are widely prevalent severe birth defects. The mouse mutant curly tail (ct/ct) has served as a model of NTDs for 50 years, even though the responsible genetic defect remained unrecognized. Here we show by gene targeting, mapping and genetic complementation studies that a mouse homolog of the Drosophila grainyhead (grh) gene, grainyhead-like-3 (Grhl3), is a compelling candidate for the gene underlying the curly tail phenotype. The NTDs in Grhl3-null mice are more severe than those in the curly tail strain, as the Grhl3 alleles in ct/ct mice are hypomorphic. Spina bifida in ct/ct mice is folate resistant, but its incidence can be markedly reduced by maternal inositol supplementation periconceptually. The NTDs in Grhl3-/- embryos are also folate resistant, but unlike those in ct/ct mice, they are resistant to inositol. These findings suggest that residual Grhl3 expression in ct/ct mice may be required for inositol rescue of folate-resistant NTDs.

Potential role of masting by introduced bamboos in deer mice (Peromyscus maniculatus population irruptions holds public health consequences.

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Melissa C Smith

Full Text Available We hypothesized that the ongoing naturalization of frost/shade tolerant Asian bamboos in North America could cause environmental consequences involving introduced bamboos, native rodents and ultimately humans. More specifically, we asked whether the eventual masting by an abundant leptomorphic ("running" bamboo within Pacific Northwest coniferous forests could produce a temporary surfeit of food capable of driving a population irruption of a common native seed predator, the deer mouse (Peromyscus maniculatus, a hantavirus carrier. Single-choice and cafeteria-style feeding trials were conducted for deer mice with seeds of two bamboo species (Bambusa distegia and Yushania brevipaniculata, wheat, Pinus ponderosa, and native mixed diets compared to rodent laboratory feed. Adult deer mice consumed bamboo seeds as readily as they consumed native seeds. In the cafeteria-style feeding trials, Y. brevipaniculata seeds were consumed at the same rate as native seeds but more frequently than wheat seeds or rodent laboratory feed. Females produced a median litter of 4 pups on a bamboo diet. Given the ability of deer mice to reproduce frequently whenever food is abundant, we employed our feeding trial results in a modified Rosenzweig-MacArthur consumer-resource model to project the population-level response of deer mice to a suddenly available/rapidly depleted supply of bamboo seeds. The simulations predict rodent population irruptions and declines similar to reported cycles involving Asian and South American rodents but unprecedented in deer mice. Following depletion of a mast seed supply, the incidence of Sin Nombre Virus (SNV transmission to humans could subsequently rise with dispersal of the peridomestic deer mice into nearby human settlements seeking food.

JTC801 Induces pH-dependent Death Specifically in Cancer Cells and Slows Growth of Tumors in Mice.

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Song, Xinxin; Zhu, Shan; Xie, Yangchun; Liu, Jiao; Sun, Lingyi; Zeng, Dexing; Wang, Pengcheng; Ma, Xiaochao; Kroemer, Guido; Bartlett, David L; Billiar, Timothy R; Lotze, Michael T; Zeh, Herbert J; Kang, Rui; Tang, Daolin

2018-04-01

lineage kinase domain like pseudokinase [MLKL]), or ferroptosis (degradation of glutathione peroxidase 4 [GPX4]). Inhibitors of apoptosis (Z-VAD-FMK), necroptosis (necrosulfonamide), ferroptosis (ferrostatin-1), or autophagy (hydroxychloroquine) did not prevent JTC801-induced death of PANC1 or MiaPaCa2 cells. The cytotoxic effects of JTC801 in immortalized fibroblast cell lines was not affected by disruption of genes that promote apoptosis (Bax -/- /Bak -/- cells), necroptosis (Ripk1 -/- , Ripk3 -/- , or Mlkl -/- cells), ferroptosis (Gpx4 -/- cells), or autophagy (Atg3 -/- , Atg5 -/- , Atg7 -/- , or Sqstm1 -/- cells). We found JTC801 to induce a pH-dependent form cell death (alkaliptosis) in cancer cells but not normal cells (hepatocytes, bone marrow CD34 + progenitor cells, peripheral blood mononuclear cells, or dermal fibroblasts) or healthy tissues of C57BL/6 mice. JTC801 induced alkaliptosis in cancer cells by activating NF-κB, which repressed expression of the carbonic anhydrase 9 gene (CA9), whose product regulates pH balance in cells. In analyses of Cancer Genome Atlas data and tissue microarrays, we associated increased tumor level of CA9 mRNA or protein with shorter survival times of patients with pancreatic, kidney, or lung cancers. Knockdown of CA9 reduced the protective effects of NF-κB inhibition on JTC801-induced cell death and intracellular alkalinization in PANC1 and MiaPaCa2 cell lines. Oral administration of JTC801 inhibited growth of xenograft tumors (from PANC1, MiaPaCa2, SK-MEL-28, PC-3, 786-0, SF-295, HCT116, OV-CAR3, and HuH7 cells), orthotropic tumors (from KPC cells), lung metastases (from KPC cells) of mice, and slowed growth of tumors in KCH mice. In a screen of agents that interact with GPCR pathways, we found JTC801 to induce pH-dependent cell death (alkaliptosis) specifically in cancer cells such as PDAC cells, by reducing expression of CA9. Levels of CA9 are increased in human cancer tissues. JTC801 might be developed for treatment of

Visualization of the human CD4{sup +} T-cell response in humanized HLA-DR4-expressing NOD/Shi-scid/γc{sup null} (NOG) mice by retrogenic expression of the human TCR gene

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Takahashi, Takeshi, E-mail: takeshi-takahashi@ciea.or.jp; Katano, Ikumi; Ito, Ryoji; Ito, Mamoru

2015-01-02

Highlights: • β-Lactoglobulin (BLG) specific TCR genes were introduced to human HSC by retrovirus. • Human HSC with BLG-specific TCR were transplanted into NOG-HLA-DR4 I-A{sup −/−} mice. • BLG-specific TCR induced positive selection of thymocytes. • BLG-specific TCR positive CD4{sup +} T cells mediated immune responses in humanized mice. - Abstract: The development of severe immunodeficient mouse strains containing various human genes, including cytokines or HLA, has enabled the reconstitution of functional human immune systems after transplantation of human hematopoietic stem cells (HSC). Accumulating evidence has suggested that HLA-restricted antigen-specific human T-cell responses can be generated in these humanized mice. To directly monitor immune responses of human CD4{sup +} T cells, we introduced β-lactoglobulin (BLG)-specific T cell receptor (TCR) genes derived from CD4{sup +} T-cell clones of cow-milk allergy patients into HSCs, and subsequently transplanted them into NOG-HLA-DR4 transgenic/I-Aβ deficient mice (NOG-DR4/I-A{sup o}). In the thymus, thymocytes with BLG-specific TCR preferentially differentiated into CD4{sup +}CD8{sup −} single-positive cells. Adoptive transfer of mature CD4{sup +} T cells expressing the TCR into recipient NOG-DR4/I-A{sup o} mice demonstrated that human CD4{sup +} T cells proliferated in response to antigenic stimulation and produced IFN-γ in vivo, suggesting that functional T-cell reactions (especially Th1-skewed responses) were induced in humanized mice.

AAV-mediated pancreatic overexpression of Igf1 counteracts progression to autoimmune diabetes in mice.

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Mallol, Cristina; Casana, Estefania; Jimenez, Veronica; Casellas, Alba; Haurigot, Virginia; Jambrina, Claudia; Sacristan, Victor; Morró, Meritxell; Agudo, Judith; Vilà, Laia; Bosch, Fatima

2017-07-01

Type 1 diabetes is characterized by autoimmune destruction of β-cells leading to severe insulin deficiency. Although many improvements have been made in recent years, exogenous insulin therapy is still imperfect; new therapeutic approaches, focusing on preserving/expanding β-cell mass and/or blocking the autoimmune process that destroys islets, should be developed. The main objective of this work was to test in non-obese diabetic (NOD) mice, which spontaneously develop autoimmune diabetes, the effects of local expression of Insulin-like growth factor 1 (IGF1), a potent mitogenic and pro-survival factor for β-cells with immunomodulatory properties. Transgenic NOD mice overexpressing IGF1 specifically in β-cells (NOD-IGF1) were generated and phenotyped. In addition, miRT-containing, IGF1-encoding adeno-associated viruses (AAV) of serotype 8 (AAV8-IGF1-dmiRT) were produced and administered to 4- or 11-week-old non-transgenic NOD females through intraductal delivery. Several histological, immunological, and metabolic parameters were measured to monitor disease over a period of 28-30 weeks. In transgenic mice, local IGF1 expression led to long-term suppression of diabetes onset and robust protection of β-cell mass from the autoimmune insult. AAV-mediated pancreatic-specific overexpression of IGF1 in adult animals also dramatically reduced diabetes incidence, both when vectors were delivered before pathology onset or once insulitis was established. Transgenic NOD-IGF1 and AAV8-IGF1-dmiRT-treated NOD animals had much less islet infiltration than controls, preserved β-cell mass, and normal insulinemia. Transgenic and AAV-treated islets showed less expression of antigen-presenting molecules, inflammatory cytokines, and chemokines important for tissue-specific homing of effector T cells, suggesting IGF1 modulated islet autoimmunity in NOD mice. Local expression of Igf1 by AAV-mediated gene transfer counteracts progression to diabetes in NOD mice. This study suggests a

Sensing vascularization of ex-vivo produced oral mucosal equivalent (EVPOME) skin grafts in nude mice using optical spectroscopy

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Vishwanath, Karthik; Gurjar, Rajan; Kuo, Shiuhyang; Fasi, Anthony; Kim, Roderick; Riccardi, Suzannah; Feinberg, Stephen E.; Wolf, David E.

2014-03-01

Repair of soft tissue defects of the lips as seen in complex maxillofacial injuries, requires pre-vascularized multi-tissue composite grafts. Protocols for fabrication of human ex-vivo produced oral mucosal equivalents (EVPOME) composed of epithelial cells and a dermal equivalent are available to create prelaminated flaps for grafting in patients. However, invivo assessment of neovascularization of the buried prelaminated flaps remains clinically challenging. Here, we use diffuse reflectance spectroscopy (DRS) and diffuse correlation spectroscopy (DCS) to non-invasively quantify longitudinal changes in the vessel density and blood-flow within EVPOME grafts implanted in the backs of SCID mice and subsequently to determine the utility of these optical techniques for assessing vascularization of implanted grafts. 20 animals were implanted with EVPOME grafts (1x1x0.05 cm3) in their backs. DRS and DCS measurements were obtained from each animal both atop the graft site and far away from the graft site, at one week post-implantation, each week, for four consecutive weeks. DRS spectra were analyzed using an inverse Monte Carlo model to extract tissue absorption and scattering coefficients, which were then used to extract blood flow information by fitting the experimental DCS traces. There were clear differences in the mean optical parameters (averaged across all mice) at the graft site vs. the off-site measurements. Both the total hemoglobin concentration (from DRS) and the relative blood flow (from DCS) peaked at week 3 at the graft site and declined to the off-site values by week 4. The optical parameters remained relatively constant throughout 4 weeks for the off-site measurements.

TAM receptor knockout mice are susceptible to retinal autoimmune induction.

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Ye, Fei; Li, Qiutang; Ke, Yan; Lu, Qingjun; Han, Lixia; Kaplan, Henry J; Shao, Hui; Lu, Qingxian

2011-06-16

TAM receptors are expressed mainly by dendritic cells and macrophages in the immune system, and mice lacking TAM receptors develop systemic autoimmune diseases because of inefficient negative control of the cytokine signaling in those cells. This study aims to test the susceptibility of the TAM triple knockout (tko) mice to the retina-specific autoantigen to develop experimental autoimmune uveoretinitis (EAU). TAM tko mice that were or were not immunized with interphotoreceptor retinoid-binding protein (IRBP) peptides were evaluated for retinal infiltration of the macrophages and CD3(+) T cells by immunohistochemistry, spontaneous activation of CD4(+) T cells, and memory T cells by flow cytometry and proliferation of IRBP-specific CD4(+) T cells by [(3)H]thymidine incorporation assay. Ocular inflammation induced by IRBP peptide immunization and specific T cell transfer were observed clinically by funduscopy and confirmed by histology. Tko mice were found to have less naive, but more activated, memory T cells, among which were exhibited high sensitivity to ocular IRBP autoantigens. Immunization with a low dose of IRBP and adoptive transfer of small numbers of IRBP-specific T cells from immunized tko mice caused the infiltration of lymphocytes, including CD3(+) T cells, into the tko retina. Mice without TAM receptor spontaneously develop IRBP-specific CD4(+) T cells and are more susceptible to retinal autoantigen immunization. This TAM knockout mouse line provides an animal model with which to study the role of antigen-presenting cells in the development of T cell-mediated uveitis.

Premature Aging Phenotype in Mice Lacking High-Affinity Nicotinic Receptors: Region-Specific Changes in Layer V Pyramidal Cell Morphology.

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Konsolaki, Eleni; Skaliora, Irini

2015-08-01

The mechanisms by which aging leads to alterations in brain structure and cognitive deficits are unclear. Α deficient cholinergic system has been implicated as one of the main factors that could confer a heightened vulnerability to the aging process, and mice lacking high-affinity nicotinic receptors (β2(-/-)) have been proposed as an animal model of accelerated cognitive aging. To date, however, age-related changes in neuronal microanatomy have not been studied in these mice. In the present study, we examine the neuronal structure of yellow fluorescent protein (YFP(+)) layer V neurons in 2 cytoarchitectonically distinct cortical regions in wild-type (WT) and β2(-/-) animals. We find that (1) substantial morphological differences exist between YFP(+) cells of the anterior cingulate cortex (ACC) and primary visual cortex (V1), in both genotypes; (2) in WT animals, ACC cells are more susceptible to aging compared with cells in V1; and (3) β2 deletion is associated with a regionally and temporally specific increase in vulnerability to aging. ACC cells exhibit a prematurely aged phenotype already at 4-6 months, whereas V1 cells are spared in adulthood but strongly affected in old animals. Collectively, our data reveal region-specific synergistic effects of aging and genotype and suggest distinct vulnerabilities in V1 and ACC neurons. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Lipid metabolism and body composition in Gclm(−/−) mice

International Nuclear Information System (INIS)

Kendig, Eric L.; Chen, Ying; Krishan, Mansi; Johansson, Elisabet; Schneider, Scott N.; Genter, Mary Beth; Nebert, Daniel W.; Shertzer, Howard G.

2011-01-01

In humans and experimental animals, high fat diets (HFD) are associated with risk factors for metabolic diseases, such as excessive weight gain and adiposity, insulin resistance and fatty liver. Mice lacking the glutamate–cysteine ligase modifier subunit gene (Gclm(−/−)) and deficient in glutathione (GSH), are resistant to HFD-mediated weight gain. Herein, we evaluated Gclm-associated regulation of energy metabolism, oxidative stress, and glucose and lipid homeostasis. C57BL/6J Gclm(−/−) mice and littermate wild-type (WT) controls received a normal diet or an HFD for 11 weeks. HFD-fed Gclm(−/−) mice did not display a decreased respiratory quotient, suggesting that they are unable to process lipid for metabolism. Although dietary energy consumption and intestinal lipid absorption were unchanged in Gclm(−/−) mice, feeding these mice an HFD did not produce excess body weight nor fat storage. Gclm(−/−) mice displayed higher basal metabolic rates resulting from higher activities of liver mitochondrial NADH-CoQ oxidoreductase, thus elevating respiration. Although Gclm(−/−) mice exhibited strong systemic and hepatic oxidative stress responses, HFD did not promote glucose intolerance or insulin resistance. Furthermore, HFD-fed Gclm(−/−) mice did not develop fatty liver, likely resulting from very low expression levels of genes encoding lipid metabolizing enzymes. We conclude that Gclm is involved in the regulation of basal metabolic rate and the metabolism of dietary lipid. Although Gclm(−/−) mice display a strong oxidative stress response, they are protected from HFD-induced excessive weight gain and adipose deposition, insulin resistance and steatosis. -- Highlights: ► A high fat diet does not produce body weight and fat gain in Gclm(−/−) mice. ► A high fat diet does not induce steatosis or insulin resistance in Gclm(−/−) mice. ► Gclm(−/−) mice have high basal metabolism and mitochondrial oxygen consumption. �

Enhancing specific-antibody production to the ragB vaccine with GITRL that expand Tfh, IFN-γ(+ T cells and attenuates Porphyromonas gingivalis infection in mice.

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Dong Zheng

Full Text Available The outer membrane protein RagB is one of the major virulence factors of the periodontal pathogen Porphyromonas gingivalis (P. gingivalis. In order to induce protective immune response against P. gingivalis infection, an mGITRL gene-linked ragB DNA vaccine (pIRES-ragB-mGITRL was constructed. Six-week-old female BALB/c mice were immunized with pIRES-ragB-mGITRL through intramuscular injection and then challenged by subcutaneous injection in the abdomen with P. gingivalis. RagB-specific antibody-forming cells were evaluated by an Enzyme-linked immunosorbent spot, and specific antibody was determined by enzyme-linked immunosorbent assay. In addition, the frequencies of Tfh and IFN-γ(+ T cells in spleen were measured using flow cytometer, and the levels of IL-21 and IFN-γ mRNA or proteins were detected by real time RT-PCR or ELISA. The data showed that the mGITRL-linked ragB DNA vaccine induced higher levels of RagB-specific IgG in serum and RagB-specific antibody-forming cells in spleen. The frequencies of Tfh and IFN-γ(+ T cells were obviously expanded in mice immunized by pIRES-ragB-mGITRL compared with other groups (pIRES or pIRES-ragB . The levels of Tfh and IFN-γ(+ T cells associated cytokines were also significantly increased in pIRES-ragB-mGITRL group. Therefore, the mice immunized with ragB plus mGITRL showed the stronger resistant to P. gingivalis infection and a significant reduction of the lesion size caused by P. gingivalis infection comparing with other groups. Taken together, our findings demonstrated that intramuscular injection of DNA vaccine ragB together with mGITRL induced protective immune response dramatically by increasing Tfh and IFN-γ(+ T cells and antibody production to P. gingivalis.

Ontogeny of Contextual Fear Memory Formation, Specificity, and Persistence in Mice

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Akers, Katherine G.; Arruda-Carvalho, Maithe; Josselyn, Sheena A.; Frankland, Paul W.

2012-01-01

Pinpointing the precise age when young animals begin to form memories of aversive events is valuable for understanding the onset of anxiety and mood disorders and for detecting early cognitive impairment in models of childhood-onset disorders. Although these disorders are most commonly modeled in mice, we know little regarding the development of…

Recombinant Mycobacterium bovis BCG producing IL-18 reduces IL-5 production and bronchoalveolar eosinophilia induced by an allergic reaction.

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Biet, F; Duez, C; Kremer, L; Marquillies, P; Amniai, L; Tonnel, A-B; Locht, C; Pestel, J

2005-08-01

Allergic reactions occur through the exacerbated induction of a Th2 cell type expression profile and can be prevented by agents favoring a Th1 profile. Bacillus Calmette-Guérin (BCG) is able to induce high IFN-gamma levels and has been shown to decrease experimentally induced allergy. The induction of IFN-gamma is mediated by interleukin (IL)-12 known to be secreted upon mycobacterial infections and can be enhanced by IL-18 acting in synergy with IL-12. We evaluated the ability of a recombinant BCG strain producing IL-18 (rBCG) to modify the Th2 type responses in a murine model of ovalbumin (OVA)-dependent allergic reaction. Mice were injected intraperitoneally or intranasally with OVA at days 0 and 15 and exposed to an OVA aerosol challenge at days 29, 30, 31 and 34. At days 0 and 15, two additional groups of mice received OVA together with 5 x 10(6) colony forming units of either rBCG or nonrecombinant BCG. A time-course analysis of OVA-specific immunoglobulin (Ig)E, IgG1 and IgG2a levels indicated no significant difference between the three groups of mice. However, following in vitro stimulation with OVA, lymph node cells from rBCG-treated mice produced less IL-5 and more IFN-gamma than those of mice injected with nonrecombinant BCG. In addition, 48 h after the last OVA challenge, a strong reduction of bronchoalveolar eosinophilia was found in the rBCG-injected mice compared to the nontreated or nonrecombinant BCG-treated groups. These results indicate that the production of IL-18 by rBCG may enhance the immunomodulatory properties of BCG that suppress pulmonary Th2 responses and, in particular, decrease airway eosinophilia.