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At least 127 records · Page 7
All sourcesNASA NTRSDOE OSTI
DOE OSTI2016

2,3,7,8-Tetrachlorodibenzo-p-dioxin has both pro-carcinogenic and anti-carcinogenic effects on neuroendocrine prostate carcinoma formation in TRAMP mice

It is well established that the prototypical aryl hydrocarbon receptor (AHR) agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) can both cause and protect against carcinogenesis in non-transgenic rodents. But because these animals almost never develop prostate cancer with old age or after carcinogen exposure, whether AHR activation can affect cancer of the prostate remained unknown. We used animals designed to develop this disease, Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice, to investigate the potential role of AHR signaling in prostate cancer development. We previously reported that AHR itself has prostate tumor suppressive functions in TRAMP mice; i.e., TRAMP mice in which Ahr was knocked out developed neuroendocrine prostate carcinomas (NEPC) with much greater frequency than did those with both Ahr alleles. In the present study we investigated effects of AHR activation by three different xenobiotics. In utero and lactational TCDD exposure significantly increased NEPC tumor incidence in TRAMP males, while chronic TCDD treatment in adulthood had the opposite effect, a significant reduction in NEPC incidence. Chronic treatment of adult TRAMP mice with the low-toxicity selective AHR modulators indole-3-carbinol or 3,3′-diindolylmethane did not significantly protect against these tumors. Thus, we demonstrate, for the first time, that ligand-dependent activation of the AHR can alter prostate cancer incidence. The nature of the responses depended on the timing of AHR activation and ligand structures. - Highlights: • TRAMP mice model aggressive neuroendocrine prostate carcinomas in men • In utero/lactational TCDD exposure raised prostate cancer incidence in TRAMP mice. • TCDD treatment in adulthood lowered prostate cancer incidence in TRAMP mice. • No significant protection was seen in TRAMP mice given I3C or DIM in adulthood. • This is the first report that TCDD alters prostate cancer incidence in lab animals.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Bisphenol A impairs the memory function and glutamatergic homeostasis in a sex-dependent manner in mice: Beneficial effects of diphenyl diselenide

Bisphenol A (BPA) is a compound integrated in commodities, which consequently increases the human exposure to this toxicant. The deleterious effects of BPA exposure during periods of brain development have been documented mainly concerning the impairment in memory functions. Diphenyl diselenide (PhSe){sub 2}, an organoselenium compound, shows protective/restorative effects against memory deficits in experimental models. Thus, this study investigated the effects of (PhSe){sub 2} on the memory impairments induced by BPA exposure to male and female mice and the possible involvement of glutamatergic system in these effects. Three-week-old male and female Swiss mice received BPA (5 mg/kg), intragastrically, from 21st to 60th postnatal day. After, the animals were intragastrically treated with (PhSe){sub 2} (1 mg/kg) during seven days. The mice performed the behavioral memory tests and the [{sup 3}H] glutamate uptake and NMDA receptor subunits (2A and 2B) analyses were carried out in the hippocampus and cerebral cortex of mice. The results demonstrated that the BPA exposure induced impairment of object recognition memory in both sexes. However, it caused impairments in spatial memory in female and in the passive avoidance memory in male mice. Besides, BPA caused a decrease in the [{sup 3}H] glutamate uptake and NMDA receptor subunit levels in the cortical and hippocampal regions depending on the sex. Treatment with (PhSe){sub 2} reversed in a sex-independent manner the behavioral impairments and molecular alterations. In conclusion, BPA had a negative effect in different memory types as well as in the glutamatergic parameters in a sex-dependent manner and (PhSe){sub 2} treatment was effective against these alterations. - Highlights: • BPA exposure induced memory impairment in a sex dependent manner in mice. • (PhSe){sub 2} reversed memory impairments caused by BPA exposure in mice. • (PhSe){sub 2} modulated hippocampal and cortical [{sup 3}H]glutamate uptake in mice. • (PhSe){sub 2} restored the hippocampal and cortical NMDA receptor subunits levels.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2014

Accumulation of cytolytic CD8{sup +} T cells in B16-melanoma and proliferation of mature T cells in TIS21-knockout mice after T cell receptor stimulation

In vivo and in vitro effects of TIS21 gene on the mature T cell activation and antitumor activities were explored by employing MO5 melanoma orthograft and splenocytes isolated from the TIS21-knockout (KO) mice. Proliferation and survival of mature T cells were significantly increased in the KO than the wild type (WT) cells, indicating that TIS21 inhibits the rate of mature T cell proliferation and its survival. In MO5 melanoma orthograft model, the KO mice recruited much more CD8{sup +} T cells into the tumors at around day 14 after tumor cell injection along with reduced tumor volumes compared with the WT. The increased frequency of granzyme B{sup +} CD8{sup +} T cells in splenocytes of the KO mice compared with the WT may account for antitumor-immunity of TIS21 gene in the melanoma orthograft. In contrast, reduced frequencies of CD107a{sup +} CD8{sup +} T cells in the splenocytes of KO mice may affect the loss of CD8{sup +} T cell infiltration in the orthograft at around day 19. These results indicate that TIS21 exhibits antiproliferative and proapoptotic effects in mature T cells, and differentially affects the frequencies of granzyme B{sup +} CD8{sup +} T-cells and CD107a{sup +} CD8{sup +} T-cells, thus transiently regulating in vivo anti-tumor immunity. - Highlights: • Constitutive expression of TIS21 in splenocytes and upregulation by TCR stimulation. • Proliferation of mature T-cells in spleen of TIS21KO mice after TCR stimulation. • Inhibition of cell death in mature T-cells of TIS21KO mice compared with the wild type. • Inhibition of melanoma growth in TIS21KO mice and CD8{sup +} T cell infiltration in tumor. • Reduction of CD 107{sup +}CD8{sup +} T cells, but increased granzyme B{sup +} CD8{sup +} T cells in TIS21KO mice.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Mini-peptide RPL41 attenuated retinal neovascularization by inducing degradation of ATF4 in oxygen-induced retinopathy mice

Highlights: • ER stress actively participates in RNV in OIR mice, in which ATF4 plays a vital role. • RPL41 inhibits the PERK/eIF2α/ATF4 pathway by phosphorylating and degrading ATF4 in mice retinas. • RPL41 is therapeutic implications for treating neovascular eye diseases. Endoplasmic reticulum (ER) stress signaling is activated in retinal degeneration disease. Activating transcription factor 4 (ATF4), an important mediator of the unfolded protein response (UPR), is a key element that maintains cell survival and proliferation in hypoxic conditions. Our previous studies showed that a small ribosomal protein L41 (RPL41) inhibits ATF4 by inducing its phosphorylation and degradation. In the present study, the effects of mini-peptide RPL41 on retinal neovascularization (RNV) in oxygen-induced retinopathy (OIR) mice was investigated. We induced OIR in C57BL/6 mice and obtained retinas from normoxia, OIR, OIR control (treated with PBS), and OIR treated (treated with RPL41) mice. Our results showed that ER stress signaling was activated and ATF4 was overexpressed in the retinas of OIR mice. After intravitreal injection of RPL41, the size of RNV and vaso-obliteration, and the number of preretinal neovascular cell nuclei in the retinas of OIR mice were significantly decreased. Western blot analysis and quantitative real-time polymerase chain reaction (qPCR) showed ATF4 and VEGF expression decreased after intravitreal injection of RPL41. Furthermore, the expression levels of inflammatory genes including TNF-α, IL-1β, and IL-6 were significantly decreased compared with the OIR control mice. In conclusion, RPL41 prevented pathologic neovascularization and exerted anti-inflammatory effects by degrading the important ER stress factor ATF4, thus, RPL41 could be a promising therapeutic agent for the treatment of neovascular eye diseases, especially retinopathy of prematurity (ROP).

60 APPLIED LIFE SCIENCES↗
DOE OSTI2022

Short term treatment with a cocktail of rapamycin, acarbose and phenylbutyrate delays aging phenotypes in mice

Pharmaceutical intervention of aging requires targeting multiple pathways, thus there is rationale to test combinations of drugs targeting different but overlapping processes. In order to determine if combining drugs shown to extend lifespan and healthy aging in mice would have greater impact than any individual drug, a cocktail diet containing 14 ppm rapamycin, 1000 ppm acarbose, and 1000 ppm phenylbutyrate was fed to 20-month-old C57BL/6 and HET3 4-way cross mice of both sexes for three months. Mice treated with the cocktail showed a sex and strain-dependent phenotype consistent with healthy aging including decreased body fat, improved cognition, increased strength and endurance, and decreased age-related pathology compared to mice treated with individual drugs or control. The severity of age-related lesions in heart, lungs, liver, and kidney was consistently decreased in mice treated with the cocktail compared to mice treated with individual drugs or control, suggesting an interactive advantage of the three drugs. This study shows that a combination of three drugs, each previously shown to enhance lifespan and health span in mice, is able to delay aging phenotypes in middle-aged mice more effectively than any individual drug in the cocktail over a 3-month treatment period.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Intermittent supplementation with fisetin improves arterial function in old mice by decreasing cellular senescence

Cellular senescence and the senescence-associated secretory phenotype (SASP) contribute to age-related arterial dysfunction, in part, by promoting oxidative stress and inflammation, which reduce the bioavailability of the vasodilatory molecule nitric oxide (NO). In the present study, we assessed the efficacy of fisetin, a natural compound, as a senolytic to reduce vascular cell senescence and SASP factors and improve arterial function in old mice. We found that fisetin decreased cellular senescence in human endothelial cell culture. In old mice, vascular cell senescence and SASP-related inflammation were lower 1 week after the final dose of oral intermittent (1 week on—2 weeks off—1 weeks on dosing) fisetin supplementation. Old fisetin-supplemented mice had higher endothelial function. Leveraging old p16-3MR mice, a transgenic model allowing genetic clearance of p16 INK4A -positive senescent cells, we found that ex vivo removal of senescent cells from arteries isolated from vehicle- but not fisetin-treated mice increased endothelium-dependent dilation, demonstrating that fisetin improved endothelial function through senolysis. Enhanced endothelial function with fisetin was mediated by increased NO bioavailability and reduced cellular- and mitochondrial-related oxidative stress. Arterial stiffness was lower in fisetin-treated mice. Ex vivo genetic senolysis in aorta rings from p16-3MR mice did not further reduce mechanical wall stiffness in fisetin-treated mice, demonstrating lower arterial stiffness after fisetin was due to senolysis. Lower arterial stiffness with fisetin was accompanied by favorable arterial wall remodeling. The findings from this study identify fisetin as promising therapy for clinical translation to target excess cell senescence to treat age-related arterial dysfunction.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2011

Biological Effects and Adaptive Response from Single and Repeated Computed Tomography Scans in Reticulocytes and Bone Marrow of C57BL/6 Mice

This study investigated the biological effects and adaptive responses induced by single and repeated in vivo computed tomography (CT) scans. We postulated that, through the induction of low-level oxidative stress, repeated low-dose CT scans (20 mGy, 2 days/week, 10 weeks) could protect mice (C57BL/6) from acute effects of high-dose radiation (1 Gy, 2 Gy). The micronucleated reticulocyte (MN-RET) count increased linearly after exposure to single CT scans of doses ranging from 20 to 80 mGy (P< 0.033). Ten weeks of repeated CT scans (total dose 400 mGy) produced a slight reduction in spontaneous MN-RET levels relative to levels in sham CT-scanned mice (P<0.04). Decreases of nearly 10% in gamma-H2AX fluorescence levels were observed in the repeated CT-scanned mice after an in vitro challenge dose of 1 Gy (P<0.017) and 2 Gy (P<0.026). Spontaneous apoptosis levels (caspase 3 and 7 activation) were also significantly lower in the repeated CTscanned mice than the sham CT-scanned mice (P < 0.01). In contrast, mice receiving only a single CT scan showed a 19% elevation in apoptosis (P < 0.02) and a 10% increase in gamma- H2AX fluorescence levels after a 2-Gy challenge (P < 0.05) relative to sham CT controls. Overall, repeated CT scans seemed to confer resistance to larger doses in mice, whereas mice exposed to single CT scans exhibited transient genotoxicity, enhanced apoptosis, and characteristics of radiation sensitization.

62 RADIOLOGY AND NUCLEAR MEDICINE↗
DOE OSTI2011

Particle Rate and Host Accelerator Beam Loss on the MICE Experiment

A study is presented of particle rates in the MICE Muon Beamline and their relationship to beam loss produced in ISIS. A brief overview of neutrino physics is presented, together with a discussion on the Neutrino Factory as a motivation for MICE. An overview of MICE itself is then presented, highlighting the need for a systematic understanding of the relationship between the MICE target parameters, ISIS beam loss, and MICE particle rate. The variation of beam loss with target depth is examined and observed to be non-linear. The variation of beam loss with respect to the target dip time in the ISIS cycle is examined and observed to be approximately linear for dip times between 11.1 ms and 12.6 ms after ISIS injection, before tailing at earlier dip times. The variation of beam loss with particle rate is also observed to follow an approximately linear relationship from 0.05 V.ms to 4.7 V.ms beam loss, with a further strong indication that this continues up to 7.1 V.ms. Particle identification using time-of-flight data is used to give an insight into the relative abundances of each particle species present in the MICE beam. Estimates of muon rate are then produced as a function of beam loss. At a level of 2 V.ms beam loss ~10.9 muons per spill for a 3.2 ms spill with negative π → μ optics, and ~31.1 muons per 1 ms spill with positive π → μ optics are observed. Simulations using the ORBIT particle tracking code of the beam loss distributions around the ISIS ring, caused by the MICE target, are also presented and the implications for MICE running discussed.

43 PARTICLE ACCELERATORS↗
DOE OSTI2018

A Novel Non-Parametric Density Estimation Approach to Measuring Muon Ionization Cooling and Reverse Emittance Exchange in the MICE Experiment

The Muon Ionization Cooling Experiment (MICE) located at Rutherford Ap- pleton Laboratory in the U.K. has demonstrated muon ionization beam cooling for the rst time. A beam of muons in MICE is produced from high-energy proton beam collision with a xed target, generating pions which in turn decay into muons. Pion-decay muons, thus, are tertiary particles and, as a result, occupy a large vol- ume in position{momentum phase space. To t the muon beam into smaller and more cost-eective accelerating devices, muon beam phase{space volume needs to be reduced (beam cooling). Ionization beam cooling, which before MICE has never been demonstrated experimentally for muons, is the only technique fast enough to be used for muons within their short lifetime. Ionization cooling occurs when muons traverse an absorbing material and lose momentum through ionization energy loss. The cooling eect in MICE is measured using two scintillating-ber tracking detec- tors. These trackers, one upstream and one downstream of the absorber, reconstruct and measure the position and momentum coordinates of each muon. Given the pre- cision MICE needed to demonstrate beam cooling, it is necessary to develop analysis tools that can account for any eects that may lead to inaccurate measurement of cooling, such as non{linear eects in beam optics. Non{parametric density estima- tion techniques, such as kernel density estimation (KDE), provide a basis for creating analysis tools that are robust against these eects, directly calculating the muon beam phase-space density and volume for demonstrating beam cooling. This thesis focuses on the application of KDE to the measurement of beam cooling in MICE. The KDE technique is validated using known distributions and is applied to sim- ulated and experimental MICE data corresponding to the various magnet, optics, and absorber congurations. Using the KDE technique, muon beam cooling in the four{dimensional transverse phase space, as well as reverse emittance exchange using MICE data have been demonstrated.

43 PARTICLE ACCELERATORS↗
DOE OSTI2023

Topography of respiratory tract and gut microbiota in mice with influenza A virus infection

Introduction Influenza A virus (IAV)-induced dysbiosis may predispose to severe bacterial superinfections. Most studies have focused on the microbiota of single mucosal surfaces; consequently, the relationships between microbiota at different anatomic sites in IAV-infected mice have not been fully studied. Methods We characterized respiratory and gut microbiota using full-length 16S rRNA gene sequencing by Nanopore sequencers and compared the nasopharyngeal, oropharyngeal, lung and gut microbiomes in healthy and IAV-infected mice. Results The oropharyngeal, lung and gut microbiota of healthy mice were dominated by Lactobacillus spp., while nasopharyngeal microbiota were comprised primarily of Streptococcus spp. However, the oropharyngeal, nasopharyngeal, lung, and gut microbiota of IAV-infected mice were dominated by Pseudomonas, Escherichia , Streptococcus , and Muribaculum spp., respectively. Lactobacillus murinus was identified as a biomarker and was reduced at all sites in IAV-infected mice. The microbiota composition of lung was more similar to that of the nasopharynx than the oropharynx in healthy mice. Discussion These findings suggest that the main source of lung microbiota in mice differs from that of adults. Moreover, the similarity between the nasopharyngeal and lung microbiota was increased in IAV-infected mice. We found that IAV infection reduced the similarity between the gut and oropharyngeal microbiota. L. murinus was identified as a biomarker of IAV infection and may be an important target for intervention in post-influenza bacterial superinfections.

Chen, Qichao↗
NASA NTRS1987

Effect of treatment with interferon-gamma and concanavalin A on the course of infection of mice with Salmonella typhimurium strain LT-2

The effect of pretreatment of mice with 34 units/day, for five days, of interferon-gamma (IFN-gamma) on the course of infection with LD50 of Salmonella typhimurium strain LT-2 was assessed, using two IFN preparations: (1) a hybridoma supernatant fluid containing concanavalin-A-induced IFN-gamma activity and (2) pure murine IFN-gamma produced by recombinant DNA technology. The hybridoma supernatant-treated Salmonella-infected mice were found to die faster than mice treated only with Salmonella. Pure murine IFN-gamma was found to protect infected mice significantly, with 95 percent of mice surviving LD50 infection. In contrast, the Salmonella-infected mice treated with hybridoma supernatant were found to die faster than the Salmonella-infected untreated controls. Mice treated with concanavalin A alone prior to infection with S. typhimurium died more quickly than the untreated infected controls, suggesting that contamination with concanavalin A had a detrimental effect on mice survival.

Gould, Cheryl L.↗
DOE OSTI2011

Lentivirus-ABCG1 instillation reduces lipid accumulation and improves lung compliance in GM-CSF knock-out mice

Highlights: Black-Right-Pointing-Pointer Lentivirus-ABCG1 reduces lipid accumulation in lungs of GM-CSF knock-out mice. Black-Right-Pointing-Pointer Up-regulation of ABCG1 improves lung function. Black-Right-Pointing-Pointer Upregulation of ABCG1 improves surfactant metabolism. -- Abstract: We have shown decreased expression of the nuclear transcription factor, peroxisome proliferator-activated receptor-gamma (PPAR{gamma}) and the PPAR{gamma}-regulated ATP-binding cassette transporter G1 (ABCG1) in alveolar macrophages from patients with pulmonary alveolar proteinosis (PAP). PAP patients also exhibit neutralizing antibodies to granulocyte-macrophage colony stimulating factor (GM-CSF), an upregulator of PPAR{gamma}. In association with functional GM-CSF deficiency, PAP lung is characterized by surfactant-filled alveolar spaces and lipid-filled alveolar macrophages. Similar pathology characterizes GM-CSF knock-out (KO) mice. We reported previously that intratracheal instillation of a lentivirus (lenti)-PPAR{gamma} plasmid into GM-CSF KO animals elevated ABCG1 and reduced alveolar macrophage lipid accumulation. Here, we hypothesized that instillation of lenti-ABCG1 might be sufficient to decrease lipid accumulation and improve pulmonary function in GM-CSF KO mice. Animals received intratracheal instillation of lenti-ABCG1 or control lenti-enhanced Green Fluorescent Protein (eGFP) plasmids and alveolar macrophages were harvested 10 days later. Alveolar macrophage transduction efficiency was 79% as shown by lenti-eGFP fluorescence. Quantitative PCR analyses indicated a threefold (p = 0.0005) increase in ABCG1 expression with no change of PPAR{gamma} or ABCA1 in alveolar macrophages of lenti-ABCG1 treated mice. ABCG1 was unchanged in control lenti-eGFP and PBS-instilled groups. Oil Red O staining detected reduced intracellular neutral lipid in alveolar macrophages from lenti-ABCG1 treated mice. Extracellular cholesterol and phospholipids were also decreased as shown by analysis of bronchoalveolar lavage fluid. Lung compliance was diminished in untreated GMCSF KO mice but improved significantly after lenti-ABCG1 treatment. Data demonstrate that in vivo instillation of lenti-ABCG1 in GM-CSF KO mice is sufficient to restore pulmonary homeostasis by: (1) upregulating ABCG1; (2) reducing intra and extracellular lipids; and (3) improving lung function. Results suggest that the ABCG1 lipid transporter is the key downstream target of GM-CSF-induced PPAR{gamma} necessary for surfactant catabolism.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2012

Conditional expression of constitutively active estrogen receptor {alpha} in chondrocytes impairs longitudinal bone growth in mice

Highlights: Black-Right-Pointing-Pointer Conditional transgenic mice expressing constitutively active estrogen receptor {alpha} (caER{alpha}) in chondrocytes were developed. Black-Right-Pointing-Pointer Expression of caER{alpha} in chondrocytes impaired longitudinal bone growth in mice. Black-Right-Pointing-Pointer caER{alpha} affects chondrocyte proliferation and differentiation. Black-Right-Pointing-Pointer This mouse model is useful for understanding the physiological role of ER{alpha}in vivo. -- Abstract: Estrogen plays important roles in the regulation of chondrocyte proliferation and differentiation, which are essential steps for longitudinal bone growth; however, the mechanisms of estrogen action on chondrocytes have not been fully elucidated. In the present study, we generated conditional transgenic mice, designated as caER{alpha}{sup ColII}, expressing constitutively active mutant estrogen receptor (ER) {alpha} in chondrocytes, using the chondrocyte-specific type II collagen promoter-driven Cre transgenic mice. caER{alpha}{sup ColII} mice showed retardation in longitudinal growth, with short bone lengths. BrdU labeling showed reduced proliferation of hypertrophic chondrocytes in the proliferating layer of the growth plate of tibia in caER{alpha}{sup ColII} mice. In situ hybridization analysis of type X collagen revealed that the maturation of hypertrophic chondrocytes was impaired in caER{alpha}{sup ColII} mice. These results suggest that ER{alpha} is a critical regulator of chondrocyte proliferation and maturation during skeletal development, mediating longitudinal bone growth in vivo.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2012

Mice carrying a complete deletion of the talin2 coding sequence are viable and fertile

Highlights: Black-Right-Pointing-Pointer Mice lacking talin2 are viable and fertile with only a mildly dystrophic phenotype. Black-Right-Pointing-Pointer Talin2 null fibroblasts show no major defects in proliferation, adhesion or migration. Black-Right-Pointing-Pointer Maintaining a colony of talin2 null mice is difficult indicating an underlying defect. -- Abstract: Mice homozygous for several Tln2 gene targeted alleles are viable and fertile. Here we show that although the expression of talin2 protein is drastically reduced in muscle from these mice, other tissues continue to express talin2 albeit at reduced levels. We therefore generated a Tln2 allele lacking the entire coding sequence (Tln2{sup cd}). Tln2{sup cd/cd} mice were viable and fertile, and the genotypes of Tln2{sup cd/+} intercrosses were at the expected Mendelian ratio. Tln2{sup cd/cd} mice showed no major difference in body mass or the weight of the major organs compared to wild-type, although they displayed a mildly dystrophic phenotype. Moreover, Tln2{sup cd/cd} mouse embryo fibroblasts showed no obvious defects in cell adhesion, migration or proliferation. However, the number of Tln2{sup cd/cd} pups surviving to adulthood was variable suggesting that such mice have an underlying defect.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2013

Recovery from diabetes in neonatal mice after a low-dose streptozotocin treatment

Highlights: ► We monitored long-term beta cell regeneration in neonatal mice treated with low dose STZ. ► Low-dose STZ neonatal female mice recovered blood glucose in 150 days. ► Glucose intolerance of the STZ treated mice significantly improved in 150 days. -- Abstract: Administration of streptozotocin (STZ) induces destruction of β-cells and is widely used as an experimental animal model of type I diabetes. In neonatal rat, after low-doses of STZ-mediated destruction of β-cells, β-cells regeneration occurs and reversal of hyperglycemia was observed. However, in neonatal mice, β-cell regeneration seems to occur much slowly compared to that observed in the rat. Here, we described the time dependent quantitative changes in β-cell mass during a spontaneous slow recovery of diabetes induced in a low-dose STZ mice model. We then investigated the underlying mechanisms and analyzed the cell source for the recovery of β-cells. We showed here that postnatal day 7 (P7) female mice treated with 50 mg/kg STZ underwent the destruction of a large proportion of β-cells and developed hyperglycemia. The blood glucose increased gradually and reached a peak level at 500 mg/dl on day 35–50. This was followed by a spontaneous regeneration of β-cells. A reversal of non-fasting blood glucose to the control value was observed within 150 days. However, the mice still showed impaired glucose tolerance on day 150 and day 220, although a significant improvement was observed on day 150. Quantification of the β-cell mass revealed that the β-cell mass increased significantly between day 100 and day 150. On day 150 and day 220, the β-cell mass was approximately 23% and 48.5% of the control, respectively. Of the insulin-positive cells, 10% turned out to be PCNA-positive proliferating cells. Our results demonstrated that, β-cell duplication is one of the cell sources for β-cell regeneration.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2014

Mammalian target of rapamycin is essential for cardiomyocyte survival and heart development in mice

Highlights: • mTOR is a critical regulator of many biological processes yet its function in heart is not well understood. • MCK-Cre/Mtor{sup flox/flox} mice were established to delete Mtor in cardiomyocytes. • The mTOR-mKO mice developed normally but die prematurely within 5 weeks after birth due to heart disease. • The mTOR-mKO mice had dilated myocardium and increased cell death. • mTOR-mKO hearts had reduced expression of metabolic genes and activation of mTOR target proteins. - Abstract: Mammalian target of rapamycin (mTOR) is a critical regulator of protein synthesis, cell proliferation and energy metabolism. As constitutive knockout of Mtor leads to embryonic lethality, the in vivo function of mTOR in perinatal development and postnatal growth of heart is not well defined. In this study, we established a muscle-specific mTOR conditional knockout mouse model (mTOR-mKO) by crossing MCK-Cre and Mtor{sup flox/flox} mice. Although the mTOR-mKO mice survived embryonic and perinatal development, they exhibited severe postnatal growth retardation, cardiac muscle pathology and premature death. At the cellular level, the cardiac muscle of mTOR-mKO mice had fewer cardiomyocytes due to apoptosis and necrosis, leading to dilated cardiomyopathy. At the molecular level, the cardiac muscle of mTOR-mKO mice expressed lower levels of fatty acid oxidation and glycolysis related genes compared to the WT littermates. In addition, the mTOR-mKO cardiac muscle had reduced Myh6 but elevated Myh7 expression, indicating cardiac muscle degeneration. Furthermore, deletion of Mtor dramatically decreased the phosphorylation of S6 and AKT, two key targets downstream of mTORC1 and mTORC2 mediating the normal function of mTOR. These results demonstrate that mTOR is essential for cardiomyocyte survival and cardiac muscle function.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2014

Mitogen-activated protein kinase kinase 1/2 inhibition and angiotensin II converting inhibition in mice with cardiomyopathy caused by lamin A/C gene mutation

Highlights: • Both ACE and MEK1/2 inhibition are beneficial on cardiac function in Lmna cardiomyopathy. • MEK1/2 inhibitor has beneficial effects beyond ACE inhibition for Lmna cardiomyopathy. • These results provide further preclinical rationale for a clinical trial of a MEK1/2 inhibitor. - Abstract: Background: Mutations in the LMNA gene encoding A-type nuclear lamins can cause dilated cardiomyopathy with or without skeletal muscular dystrophy. Previous studies have shown abnormally increased extracellular signal-regulated kinase 1/2 activity in hearts of Lmna{sup H222P/H222P} mice, a small animal model. Inhibition of this abnormal signaling activity with a mitogen-activated protein kinase kinase 1/2 (MEK1/2) inhibitor has beneficial effects on heart function and survival in these mice. However, such treatment has not been examined relative to any standard of care intervention for dilated cardiomyopathy or heart failure. We therefore examined the effects of an angiotensin II converting enzyme (ACE) inhibitor on left ventricular function in Lmna{sup H222P/H222P} mice and assessed if adding a MEK1/2 inhibitor would provide added benefit. Methods: Male Lmna{sup H222P/H222P} mice were treated with the ACE inhibitor benazepril, the MEK1/2 inhibitor selumetinib or both. Transthoracic echocardiography was used to measure left ventricular diameters and fractional shortening was calculated. Results: Treatment of Lmna{sup H222P/H222P} mice with either benazepril or selumetinib started at 8 weeks of age, before the onset of detectable left ventricular dysfunction, lead to statistically significantly increased fractional shortening compared to placebo at 16 weeks of age. There was a trend towards a great value for fractional shortening in the selumetinib-treated mice. When treatment was started at 16 weeks of age, after the onset of left ventricular dysfunction, the addition of selumetinib treatment to benazepril lead to a statistically significant increase in left ventricular fractional shortening at 20 weeks of age. Conclusions: Both ACE inhibition and MEK1/2 inhibition have beneficial effects on left ventricular function in Lmna{sup H222P/H222P} mice and both drugs together have a synergistic benefit when initiated after the onset of left ventricular dysfunction. These results provide further preclinical rationale for a clinical trial of a MEK1/2 inhibitor in addition to standard of care in patients with dilated cardiomyopathy caused by LMNA mutations.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Sustained expression of GLP-1 receptor differentially modulates β-cell functions in diabetic and nondiabetic mice

Glucagon-like peptide 1 (GLP-1) has been shown to play important roles in maintaining β-cell functions, such as insulin secretion and proliferation. While expression levels of GLP-1 receptor (Glp1r) are compromised in the islets of diabetic rodents, it remains unclear when and to what degree Glp1r mRNA levels are decreased during the progression of diabetes. In this study, we performed real-time PCR with the islets of db/db diabetic mice at different ages, and found that the expression levels of Glp1r were comparable to those of the islets of nondiabetic db/misty controls at the age of four weeks, and were significantly decreased at the age of eight and 12 weeks. To investigate whether restored expression of Glp1r affects the diabetic phenotypes, we generated the transgenic mouse model Pdx1{sup PB}-CreER{sup TM}; CAG-CAT-Glp1r (βGlp1r) that allows for induction of Glp1r expression specifically in β cells. Whereas the expression of exogenous Glp1r had no measurable effect on glucose tolerance in nondiabetic βGlp1r;db/misty mice, βGlp1r;db/db mice exhibited higher glucose and lower insulin levels in blood on glucose challenge test than control db/db littermates. In contrast, four weeks of treatment with exendin-4 improved the glucose profiles and increased serum insulin levels in βGlp1r;db/db mice, to significantly higher levels than those in control db/db mice. These differential effects of exogenous Glp1r in nondiabetic and diabetic mice suggest that downregulation of Glp1r might be required to slow the progression of β-cell failure under diabetic conditions. - Highlights: • Expression levels of incretin receptors were significantly decreased in diabetic db/db islets after the age of eight weeks. • A transgenic mouse model expressing Glp1r specifically in β cells was generated. • Exogenous expression of Glp1r in β cells did not affect metabolic profiles in nondiabetic mice. • Sustained expression of Glp1r in diabetic db/db β cells deteriorated glucose profiles, but significantly improved insulin secretion after treatment with a GLP-1 analog.

60 APPLIED LIFE SCIENCES↗