Liver proteome of mice with different genetic susceptibilities to the effects of fluoride

ABSTRACT A/J and 129P3/J mice strains have been widely studied over the last few years because they respond quite differently to fluoride (F) exposure. 129P3/J mice are remarkably resistant to the development of dental fluorosis, despite excreting less F in urine and having higher circulating F levels. These two strains also present different characteristics regardless of F exposure. Objective In this study, we investigated the differential pattern of protein expression in the liver of these mice to provide insights on why they have different responses to F. Material and Methods Weanling male A/J and 129P3/J mice (n=10 from each strain) were pared and housed in metabolic cages with ad libitum access to low-F food and deionized water for 42 days. Liver proteome profiles were examined using nLC-MS/MS. Protein function was classified by GO biological process (Cluego v2.0.7 + Clupedia v1.0.8) and protein-protein interaction network was constructed (PSICQUIC, Cytoscape). Results Most proteins with fold change were increased in A/J mice. The functional category with the highest percentage of altered genes was oxidation-reduction process (20%). Subnetwork analysis revealed that proteins with fold change interacted with Disks large homolog 4 and Calcium-activated potassium channel subunit alpha-1. A/J mice had an increase in proteins related to energy flux and oxidative stress. Conclusion This could be a possible explanation for the high susceptibility of these mice to the effects of F, since the exposure also induces oxidative stress.

Enamel crystals of mice susceptible or resistant to dental fluorosis: an AFM study

Objective: This study aimed to assess the overall apatite crystals profile in the enamel matrix of mice susceptible (A/J strain) or resistant (129P3/J strain) to dental fluorosis through analyses by atomic force microscopy (AFM). Material and Methods: Samples from the enamel matrix in the early stages of secretion and maturation were obtained from the incisors of mice from both strains. All detectable traces of matrix protein were removed from the samples by a sequential extraction procedure. The purified crystals (n=13 per strain) were analyzed qualitatively in the AFM. Surface roughness profile (Ra) was measured. Results: The mean (±SD) Ra of the crystals of A/J strain (0.58±0.15 nm) was lower than the one found for the 129P3/J strain (0.66±0.21 nm) but the difference did not reach statistical significance (t=1.187, p=0.247). Crystals of the 129P3/J strain (70.42±6.79 nm) were found to be significantly narrower (t=4.013, p=0.0013) than the same parameter measured for the A/J strain (90.42±15.86 nm). Conclusion: enamel crystals of the 129P3/J strain are narrower, which is indicative of slower crystal growth and could interfere in the occurrence of dental fluorosis. .

Betamethasone inhibits tumor development, microvessel density and prolongs survival in mice with a multiresistant adenocarcinoma TA3

Tumor resistance to traditional cancer treatments poses an important challenge to modern science. Thus, angiogenesis inhibition is an important emerging cancer treatment. Many drugs are tested and corticosteroids have shown interesting results. Herein we investigate the effect on microvessel density, survival time and tumoral volume of mice with TA3-MTX-R tumors. Twenty six mice were inoculated with lxlO6 tumor cells, 4-5 days after injection, six mice were injected with PBS (group A) and twenty mice were treated with p-met (group B). All animals from Group A died on day 22. Group B was divided into Bl (treated discontinued) and B2 (treated daily) and observed until day 88. All mice were processed for histo-immunohistochemical analysis and the blood vessels were counted. A decrease in microvessel density and tumoral volume and longer survival times were observed in the treated group. We propose that the antiangiogenic p-met effect explains, at least partially, its tumor inhibitory properties. As an important perspective, we will experimentally combine these strategies with those recently described by us with regard to the important antiangiogenic-antitumor effects of Trypanosoma cruzi calreticulin. Since the molecular targets of these strategies are most likely different, additive or synergic effects are envisaged.

Gene expression in IFN-gamma-activated murine macrophages

Macrophages are critical for natural immunity and play a central role in specific acquired immunity. The IFN-gamma activation of macrophages derived from A/J or BALB/c mice yielded two different patterns of antiviral state in murine hepatitis virus 3 infection, which were related to a down-regulation of the main virus receptor. Using cDNA hybridization to evaluate mRNA accumulation in the cells, we were able to identify several genes that are differently up- or down-regulated by IFN-gamma in A/J (267 and 266 genes, respectively, up- and down-regulated) or BALB/c (297 and 58 genes, respectively, up- and down-regulated) mouse macrophages. Macrophages from mice with different genetic backgrounds behave differently at the molecular level and comparison of the patterns of non-activated and IFN-gamma-activated A/J or BALB/c mouse macrophages revealed, for instance, an up-regulation and a down-regulation of genes coding for biological functions such as enzymatic reactions, nucleic acid synthesis and transport, protein synthesis, transport and metabolism, cytoskeleton arrangement and extracellular matrix, phagocytosis, resistance and susceptibility to infection and tumors, inflammation, and cell differentiation or activation. The present data are reported in order to facilitate future correlation of proteomic/transcriptomic findings as well as of results obtained from a classical approach for the understanding of biological phenomena. The possible implication of the role of some of the gene products relevant to macrophage biology can now be further scrutinized. In this respect, a down-regulation of the main murine hepatitis virus 3 receptor gene was detected only in IFN-gamma-activated macrophages of resistant mice.

Inhibition of DMBA-croton oil two-stage mouse skin carcinogenesis by diphenylmethyl selenocyanate through modulation of cutaneous oxidative stress and inhibition of nitric oxide production.

Selenium, an essential micronutrient, plays important roles against different diseases, including several types of cancer. In the present study, antioxidative and chemopreventive properties of a synthetic organoselenium compound, diphenylmethyl selenocyanate, were evaluated with a 7,12-dimethylbenz (a) anthracene - croton oil induced two-stage mouse skin carcinogenesis model. The compound was administered orally to carcinogen-treated mice at two different non-toxic doses, 2mg/kg. b.w. and 3mg/kg. b.w. Significant inhibition in the incidence of papilloma formation (53-80%) as well as in the cumulative numbers of papillomas per papilloma bearing mouse were observed in the treated groups as compared to the carcinogen control group. The compound was also found to upregulate significantly different phase II detoxifying enzymes such as glutathione-S-transferase (p<0.01) and superoxide dismutase (p<0.01) in skin cytosol when measured after 15 days and also after 12 weeks of the first 7,12-dimethylbenz (a) anthracene treatment. Lipid peroxidation measured with reference to thiobarbituric acid reactive substances in skin microsomes was significantly inhibited (p<0.05) in a dose dependent manner by diphenylmethyl selenocyanate. Considerable inhibition of the level of nitric oxide production in peritoneal macrophages was observed after 12 weeks (p<0.05). Thus the compound appears to exert chemopreventive activity in terms of papilloma formation, which may be through modulation of cutaneous lipid peroxidation, the phase II detoxifying enzyme system and nitric oxide production.

Combined therapy of interleukin-12 and interleukin-18 against Cryptococcus neoformans infection in a murine model / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To explore adverse effects of combined treatment of interleukin-12 (IL-12) and interleukin-18 (IL-18) against cryptococcosis in a murine model.</p><p><b>METHODS</b>Infected mice were treated with a combination of IL-12 and IL-18. Their body weight and intake of water and food were observed and recorded. Serum levels of leptin were detected with an enzyme-linked immuno sorbent assay (ELISA).</p><p><b>RESULTS</b>In the combined treatment group, the intake volume of water and food were reduced, leading to weight loss and undetectable levels of leptin in the serum. These adverse effects were more profound in mice that had received higher doses of cytokines, which sometimes led to a fatal outcome. There was a significant difference compared with the control group. Neutralization of endogenous tumor necrosis factor-alpha (TNF-alpha) by its specific mAb did not alter the wasting effect of this treatment.</p><p><b>CONCLUSIONS</b>The combined IL-12/IL-18 treatment may cause a number of adverse effects independent of TNF-alpha and leptin synthesis. Further investigations for resolving these adverse effects are required before clinical application of these cytokines.</p>

Free radical scavenging and metal chelation by Tinospora cordifolia, a possible role in radioprotection.

Aqueous extract of T. cordifolia inhibited Fenton (FeSO4) reaction and radiation mediated 2-deoxyribose degradation in a dose dependent fashion with an IC50 value of 700 microg/ml for both Fenton and radiation mediated 2-DR degradation. Similarly, it showed a moderate but dose dependent inhibition of chemically generated superoxide anion at 500 microg/ml concentration and above with an IC50 value of 2000 microg/ml. Aqueous extract inhibited the formation of Fe2+-bipiridyl complex and formation of comet tail by chelating Fe2+ ions in a dose dependent manner with an IC50 value of 150 microg/ml for Fe2+-bipirydyl formation and maximally 200 microg/ml for comet tail formation, respectively. The extract inhibited ferrous sulphate mediated lipid peroxidation in a dose-dependent manner with an IC50 value of 1300 microg/ml and maximally (70%) at 2000 microg/ml. The results reveal that the direct and indirect antioxidant actions of T. cordifolia probably act in corroboration to manifest the overall radioprotective effects.

Chemical compositions and pharmacological effects of essential oil from the fruit of Zanthoxylum limonella.

Examination of chemical compositions of essential oil distilled from the fruit of Zanthoxylum limonella Alston (Rutaceae) revealed the presence of 33 chemical components. Evaluation of the oil composition was achieved by GC/MS analysis. Limonene (31.09%), terpin-4-ol (13.94%) and sabinene (9.13%) were found to be the major components. Effects of essential oil have been performed in isolated guinea pig ileum, rat thoracic aorta and conscious mice. The essential oil at the concentration of 7.68 x 10(-5)-1.92 x 10(-3) microl/25 ml, produced dose-dependent contraction of the isolated rat thoracic aorta. These contractions were significantly reduced by pretreatment with prazosin (1 x 10(-7) M) and verapamil (1 x 10(-7) M). Its contraction was abolished in calcium free Krebs solution. Contractile response to the volatile oil (2 x 10(-6)-5.12 x 10(-4) microl/25 ml) was examined in isolated guinea pig ileum, it evoked ileal contraction in concentration-dependent manner and the contractions were suppressed after exposure to chlorpheniramine (1 x 10(-7) M) cypoheptadine (1 x 10(-7) M) atropine (1 x 10(-7) M) and verapamil (1 x 10(-7) M). Therefore, it could be concluded that the essential oil from the fruit of Zanthoxylum limonella possessed stimulation effect on different smooth muscle preparations by non-specific mechanisms. It involved the non receptor and receptor-mediated mechanism. Gastrointestinal stimulant effect of the essential oil was confirmed in intact mice since the oil significantly increased black ink movement from the stomach to ileo-caecal junction after oral feeding.

Construcción de una biblioteca genómica de Leishmania amazonensis y su expresión en músculo de ratones BALB/c / Building of a genomic library of Leishmania amazonensis and its expression in BALB/c miceïs muscle

Se construyó una biblioteca genómica de Leishmania amazonensis mediante el vector pcDNA3, con promotor de expresión en células eucariotas, con el objetivo de contribuir a la aplicación de la tecnología de inmunización con ácidos nucleicos en la leishmaniosis. Para demostrar la expresión de la genoteca en el músculo de ratones inmunizados con esta, se realizó la técnica de inmunofluorescencia indirecta. Como anticuerpo primario se utilizó una mezcla de sueros con alto título antileishmania, de una zona donde predomina la infección con L. braziliensis. Se obtuvo una biblioteca con 80 porciento de clones recombinantes. Se demostró la expresión de determinantes antigénicos en el músculo de ratones BALB/c inmunizados, según resultados de la inmunofluorescencia

Role of ketamine in convulsions.

It can be concluded from the present study that ketamine showed dose-dependent anticonvulsant effect on MES in mice. It is presumed that this anticonvulsant effect of ketamine could be due to blockade of excitatory amino acid NMDA receptors. It was potentiated by anticonvulsants like diazepam and DPH but was found to be insensitive to naloxone. These findings suggest the involvement of NMDA receptors and their antagonists in epilepsy. Ketamine thus can be given as add-on therapy in refractory cases, and may prove to be useful as an anticonvulsant in future.

Camundongos isogênicos A/SNELL, BALB/c e C57BL/6 como modelos biológicos de sepsis intra-abdominal / Inbred mice A/SNELL, BALB/c and C57L/6 as animal models of intra-abdominal sepsis

Com o objetivo de desenvolver modelos biológicos alternativos de sepsis intra-abdominal para estudos experimentais, foram infectados camundongos isogênicos A/ SNELL, BALB/c e C57BL/6, com a bactéria "Escherichia coli", cepa ATCC 117755, sorotipo H7:01:K1. O resultado da determinaçào da DL50 mostrou que para camundongos da linhagem C57BL/6 a dose (3E-5,85 UFC) foi 14 vezes maior do que para camundongos A/SNELL (3E-5,68 UFC), e 12 vezes maior do que para camundongos BALB/c (3xE-5,77). Os exames histopatológicos de orgãos da cavidade abdominal, juntamente com análise do número de leucócitos polimorfonucleares encontrados no lavado peritoneal e em esfregaço sangüíneo, representam parâmetros que podem ser utilizados para avaliação de sepsis intra-abdominal em estudos experimentais (au)

Specificity and role of anti-Trypanosoma cruzi clearance antibodies

Two strains of Trypanosoma Cruzi (Y and CL) were used to study the specificity and role of anti-T. cruzi clearance antibodies. Clearance antibodies were only induced after immunization with living blood-stream trypomastigotes (Btrys) but not with dead parasites. Btrys of either strain were readily cleared from the circulation after passive immunization with anti-Y or anti-CL scrum provided that the homologous strain was used. CL or Y Btrys sensitized in vitro with the homologous or heterologous antiserum and transferred to normal mice were cleared from the circulation only when the homologous antiserum was used. Clearance antibodies were removed from serum by absorption with the homologous but not with the heterologous strain. Clearance antibodies were removed from serum by absorption with living Btrys but not with fixed parasites. These results suggest that: a) the parasite epitopes involved in the clearance are peculiar to each strain, b) the clearance antibodies are specific to these epitopes, and c) a proper conformation of the parasite antigens is required for the induction and effector activity of the clearance antibodies.

Reduced sensory neuron regeneration by C57BL/6J mice

The tube repair method was used to study peripheral nerve regeneration in five different inbred mouse strains. The sciatic nerve of male adult mice of the C57BL/6J,DBA/1J, C3H/HeJ, BALB/cJ and A/J strains (N=3) was cut and both proximal and distal nerve stumps were inserted into a polyethylene tube leaving a 4-mm nerve gap. After 6 weeks the tubes containing the regenerated nerve cables were processed for total myelinated axon counts. C57BL/6J mice regenerated significantly fewer myelinated axons (1024 + or - 178, mean + or - SEM) compared to the BALB/cJ (1618+ or - 64), a/j (1788 + OR - 95), dba/1j(2168 + OR - 296) OR c3h/hEj (3468 + OR - 36) strains. Horseradish peroxidase was applied 3 mm distal to be tube 4 and 40 weeks after tube implantation to further characterize the reduced regenerative response of C57BL/6J mice. Labeled sensory and somatic motor neurons were counted in the spinal cord and L4,5,6 dorsal root ganglia (DRG), respectively. Sciatic nerves from four intact C57BL/6J mice were processed in the same fashion and used as normal controls. No significant difference in the number of motor neurons was detected between the experimental (4 weeks = 663 + or - 13) animals. However, there were fewer labeled neurons in the DRG of the operated group (4 weeks = 1163 + or - 167; 40 weeks = 2574 + or - 104) compared to the control group (4211 + or - 96). These results indicated that sensory neurons are responsible for the diminished regenerative response in C57BL/6J mice after peripheral nerve transection. Neurotrophic factors may be implicated in the reduced response, although no systematic study has been done to quantify either trophic factors or their receptor synthesis in different mouse strains during Wallerian degeneration. Diminished peripheral nerve fiber regeneration makes the C57BL/6J mouse strain an ideal experimental model to evaluate the effects of exogenous substances on nerve repair

Role of platelets and complement in the clearance of epimastigote forms of Trypanosoma cruzy

1. Trypanosoma cruzi epimastigote forms are very rapidly removed from the circulation of normal and C5-deficient mice. Depletion of C3 by cobra venom factor results in a significant delay in parasite clearance. 2. During parasite clearance there is a significant decrease in the number of circulating platelets and parasite clearance is considerably delayed in thrombocytopenic animals. 3. In vitro incubation of epimastigote forms with normal mouse serum leads to the formation of parasite clumps provided that platelets are present. Innactivation of factor B or depletion of C3 prevents this phenomenon. 4. When epimastigotes are incubated with normal mouse serum they absorb one or more factors required for their aggregation with platelets. 5. It is suggested that in mice T. cruzi epimastigote forms are removed from circulation by the alternative pathway of complement activation and that both C3 and platelets are required for parasite clearance

Antiviral activity of interferon gamma in vivo during mouse hepatitis virus infection

A/J mice became resistant to experimental MHV3 infection after immunization with UV-inactivated MHV3 (0 percent mortality, 0/10). Depletion of interferon (IFN) gamma-producing CD4+ T lymphocytes with monoclonal antibodies to CD4+ led to susceptibility to virus infection (60 percent of mortality, 6/10). The resistance to MHV3 infection of CD4+ T lymphocyte-depleted-A/J mice was restored by treatment with 1000 U of IFN gamma on days -1, 0, 1, 2, 3 and 4 (10 percent of mortality, 1/10). The low virus titers observed in resistant mice (controls or CD4+ depleted plus IFN gamma treated) were cleared 6 days after infection and the virus titers observed among susceptible mice (CD4+ depleted) increased gradually and peaked on day 6, when the animals died. Previous data, taken together with the direct evidence presented in this paper, provide strong evidence supporting the concept of an in vivo antiviral role of IFN gamma through a central action on the mechanisms of resistance to MHV3 infection

Reduction of the accumulation of viral mRNA and of cellular rRNA synthesis in MHV3-infected macrophages of resistant mice

1. After MHV3 infection, only macrophages from resistant A/J mice partially restricted virus growth compared to those from susceptible BALB/c mice (2 logs of difference in virus titer). 2. Cellular ribosomal ribonucleic acid (rRNA) synthesis by MHV3-infected macrophages was decreased only in A/J mouse macrophages as indicated by accumulation of the 28S rRNA fraction. 3. The accumulation of viral messenger ribonucleic acids (mRNAs) in MHV3-infected macrophages was also reduced in A/J mouse macrophages compared to BALB/c mice. 4. In pulse-chase experiments of viral protein synthesis, the appearance, glycosylation and cleavage of glycoprotein S, as well as the metabolism of nucleoprotein N were delayed in A/J mouse macrophages. 5. These data show that MHV3 infection of A/J mouse macrophages induced an imbalanced accumulation of the 28S fraction of rRNA. Furthermore the synthesis of mRNAs correlated with viral protein synthesis in both A/J and BALB/c macrophages, but was delayed in A/J mice. 6. These results suggest that the partial restriction of MHV3 replication in macrophages of resistant A/J mice may take place during or before the mRNA synthesis, although it is correlated with the appearance, glycosylation, cleavage and metabolism of viral proteins

Effect of some chemical radioprotectors on mouse bone marrow.

Effect of HT, AET and Se on mice bone marrow has been studied by counting bone marrow micronucleated cells and endogenous spleen colony count (CFU-S). Combination of HT and AET used as a radioprotector has not caused any significant variation in any of the parameter studied when administered once, it increases bone marrow micronucleated cells and decreases CFU-S slightly after daily administration for 7 days. The individual constituent of the combination administered singly does not increase micronucleated cell number. Seven consecutive doses of HT + AET and same in combination with Se enhances micronucleated cells to a higher level. Daily injection of Se alone up to 7 days also causes an increase in micronucleated cells upto same level. CFU-S pool does not show any significant change in number of bone marrow cells through out the study except in the groups where animals were treated with Se.

Mouse hepatitis virus 3 and interferon gamma binding to extracted macrophage proteins correlate with virus growth in A/J and BALB/c mice

1. After immunization, adult A/J mice are resistant and BALB/c mice are susceptible to MHV3 infection. After IFN gamma activation, only macrophages originating from A/J mice were able to partially restrict MHV3 growth. 2. When the binding of MHV3 and interferon (IFN) gamma to solubilized cytoplasmic and membrane macrophage proteins of mice was determined by ELISA, there was more binding of MHV3 to proteins extracted from BALB/c macrophages than to proteins extracted from A/J macrophages. When the proteins were obtained from IFN gamma-activated macrophages, decreased MHV3 binding was observed only in proteins originating from A/J macrophages. 3. ELISA showed a comparable binding of IFN gamma to A/J or BALB/c macrophage proteins. When the proteins were obtained from IFN gamma-activated macrophages, only IFN gamma-binding to A/J macrophage proteins was increased. 4. The results indicate a different expression and IFN gamma modulation of MHV3 receptors in macrophages from A/J and BALB/c mice, which directly correlated with their acquired resistance or susceptibility to MHV3 infection

Trypanosoma cruzi and experimental Chagas' disease: characterization of a stock isolated from a patient with associated digestive and cardiac form

Um novo estoque de Trypanosoma cruzi isolado de paciente chagásico crônico, com a forma digestiva e cardiaca da doença, foi caracterizado através de infecçäo experimental em camundongos isogênicos A/Sn suscetíveis à infecçäo chagásica. As curvas de parasitemia mostraram picos de até 1,7x10**6 parasitas/ml näo se observando mortalidade até 300 dias após infecçäo. anticorpos da classe IgM foram encontrados na fase aguda até 40 dias e também na fase crônica e IgG foi detectada nas fases aguda e crônica. O exame histopatológico mostrou miotropismo para músculo liso do tubo digestivo e cardíaco